Qatar Foundation Annual Research Forum Volume 2013 Issue 1

The expanding economy of Qatar in the last two decades has attracted immigrants, often from countries with poor socio-economic standards. Many arrive with patent intestinal parasitic infections, and recent analyses have indicated consistently rising trends in the prevalence of some infections. We have used several approaches for quantifying the prevalence of parasitic infections brought into the country by immigrant workers. Using data from 2009 we compared prevalence of infection in newly arrived immigrants with that in long-term residents from the same nations (matching countries among which both newly arrived immigrants and long-term residents were represented) and found that intestinal parasitic infections fell from 26.5% among the newly arrived workers to 16.5% among residents, the biggest drop being among helminth infections (from 20% to 9.2%). Helminth infections among newly arrived workers varied significantly by region of origin, and hookworm infections in particular were frequent among the Nepalese (18.9%) but in contrast to all other regions, did not fall markedly after acquisition of residency status (17.5%). Protozoan infections changed little overall, because some species increased while others declined: there was a significant increase in the prevalence of Blastocystis hominis among residents compared with immigrants and a concomitant declined in both Giardia duodenalis and Entamoebae histolytica/dispar. More recently we analyzed another data-set of 18,563 hospital records of subjects from 57 countries who in the period from 2005 to 2011 sought medical assistance for a variety of ailments, to enable trends to be identified across a seven year period. We found that overall 8.6% were infected with one or more species of parasite, but in contrast to the earlier years, in the last three years there were falling trends of prevalence (from 13.4% in 2009 to 4.9% in 2011) providing some optimism that parasitic infections among the resident immigrants have begun to decline. We identified also geographic regions from which resident workers still maintain relatively high prevalences of helminth infections despite their long-term residence in Qatar. Workers from Nepal were the most likely to carry hookworm infections (19.7%) followed by other W. Asian nationals including in descending order those from Bangladesh (8.3%), Sri Lanka (6.8%) and India (4.4%). Helminth infections are probably acquired abroad during visits to their home villages, whilst protozoan infections are reinforced by transmission in Qatar, possibly in the poorer areas of the state where immigrant workers live. Our results have clearly identified high risk groups among the many foreign immigrant workers and have clear implications for the health authorities. The two outstanding problems that now require further attention are the still relatively high prevalence rates with hookworms among the Nepalese and other Asian workers and the rising trend in the prevalence of B. hominis with host age among both Qataris and foreign residents. Our attention is currently focused on unravelling the reasons for the relatively high prevalence of hookworm infections among resident Asians, and especially the Nepalese, in the hope that eventually the decline in these infections can be accelerated even more markedly than has been observed in recent years.

Correlation between quality of life and disease severity in children with atopic dermatitis in the state of Qatar Background: Atopic dermatitis (AD) is an itchy, chronic or chronically relapsing, inflammatory skin condition which is most prevalent in childhood. It is known to affect 10-20% of children. The prevalence of AD has increased during the past 3 decades and is probably due to modification of lifestyles and environment. Atopic dermatitis (AD) can affect the behavior, life and development of children. Little is known regarding the effect of AD on the quality of life (QoL) in children of the Gulf region and especially in the state of Qatar. Objectives: To study the impact of AD on the QoL in children and its relation to disease severity in the state of Qatar. Methods: This study was conducted on 45 patients with atopic dermatitis aged from 5 to 16 years who attended the outpatient clinic of the Dermatology Department, Rummilah and Al Khor Hospitals, Hamad Medical Corporation, Doha, Qatar. The UK diagnostic criteria of AD were used for the diagnosis of atopic dermatitis. Disease severity was assessed using modified SCORAD (SCORing Atopic Dermatitis) measuring only objective criteria .The Children Dermatology Life Quality Index (CDLQI) was used to quantify the impact of AD on children's quality of life. SPSS 14.0 statistical package has been used for the analysis. Correlation coefficient (Pearson) was calculated to see associations between variables. Results: There were a positive significant correlation between objective SCORAD and each question from 1 to 10 as well as the total score of CDLQI except for question 2, the strongest relation between objective SCORAD and CDLQI was found in question 1 (symptoms) and question 9 (sleep). It was statistically evident that as the SCORAD index increases the CDLQI increases. Conclusion: The study has shown adverse effects of AD on children QoL especially on symptoms and feelings and a positive correlation between CDLQI and modified SCORAD which denote the effect of AD on the lives and development of children. We recommend the use of CDLQI in research work as an additional subjective measure to the clinical objective scoring tools used in chronic skin diseases. Health economists and caregivers should put the impact of skin diseases (non-life threatening diseases) on the QoL of patients, especially children and its implications for psychology, development, social functioning and school outcome.

Malignant tumors metabolize glucose through glycolysis to lactic acid and produce ATP at an accelerated rate when compared to normal cells, a phenotype detected clinically using Positron Emission Tomography (PET). Out of the four available isoforms, Hexokinase 2 (HK2) is the major expressing isoform in cancers specially those that metastasize and kill their human host. In malignant cells, HK2 not only improves the cell's energy supply but also protects cancer cells against apoptosis through direct interaction with the mitochondria and Voltage Dependent Anion Channel 1 (VDAC1). Here we report the 3D crystal structure of the human HK2 in complex with glucose and glucose-6-phosphate, the first enzyme in the glycolytic pathway. The N- and C-terminal domains are catalytically active and linked by a long seven turn a-helix. Each domain contains an active site that is sandwiched between a small and large sub-domain. Cancer patients with high HK2 gene expression level showed a worse prognosis and aggressive character. Therefore, HK2 is an ideal therapeutic target to inhibit cancer proliferation and tumor destruction. Very limited literature describes HK2 and detailed understanding of the HK2 structure and function is needed to characterize its mechanism, to understand its role in cancer metabolism and to develop new drug treatments to slow the rate of cancer proliferation.

The foodborne and waterborne disease continues to pose significant health problems in the Middle East. This region specifically Arabic speaking countries, imports large amounts of food products from all over the world. At the same time, this region exports a variety of food commodities to many countries worldwide. The increased international food trade, changes in agriculture production, food processing, and consumer demand for wholesome safe food have led to new food safety challenges which have impacted the food safety systems. There are country- and intra-country/subregional specific based food safety systems. The Gulf Cooperation Council (GCC) is an example of a subregional coalition aimed at harmonizing their country-members' food safety standards based on Codex Alimentarius. There are a number of challenges and potential opportunities available for the Middle Eastern countries to enhance their food safety systems. These challenges include: 1) domestic and imported food inspection, 2) laboratory capacity and staff training, 3) foodborne surveillance systems, and 4) scientific research to support policies and regulations. Given these challenges, there are a number of opportunities to enhance food safety in the Middle East. These include: 1) Training of inspectors for domestic and imported food and making available a detailed and comprehensive manual to follow as a guideline in their daily work. This guideline would be a valuable aid especially to those with limited training. In addition, standard uniform methods for sampling and data collection would be needed, 2) building research institutions for food safety and quality research that supports laboratory capacity, surveillance, and data analysis, and 3) better risk assessment and communication is needed to focus on highest foodborne disease risks and to link the strategies to reduce those risks across the full food production chain. Special consideration should be given to certain situations in specific countries to overcome barriers of non-collaboration among disciplines and responsible agencies.

Anti-allergic effects by arginase inhibitors: role of nitric oxide Herman Meurs¹, Ramadan Sopi¹´², Mirjam Simoons¹, Paul Jackson³, Harm Maarsingh¹´³ ¹Department of Molecular Pharmacology, Groningen Research Institute for Pharmacy and Groningen Research Institute for Asthma and COPD, University of Groningen, Groningen, The Netherlands, ²Faculty of Medicine, University of Prishtina, Prishtina, Kosovo, ³Department of Pharmaceutical Sciences, Gregory School of Pharmacy, Palm Beach Atlantic University, West Palm Beach, FL, USA Using animal models for acute and chronic asthma, we demonstrated that inhaled arginase inhibitors attenuate allergen-induced early and late asthmatic reactions, airway hyperresponsiveness, airway inflammation and airway remodeling. Moreover, it was found that arginase inhibition greatly reduces the sensitivity of the airways to the inhaled allergen, suggesting that arginase inhibitors may have an anti-allergic effect by inhibition of mediator release from mast cells. Since arginase activity regulates nitric oxide (NO) synthesis via competition with NO synthase (NOS) for the common substrate L-arginine, and activation of mast cells is inhibited by NO, we hypothesized that the anti-allergic effect of arginase inhibition involves increased NO synthesis. This hypothesis was investigated in precision-cut lung slices from actively ovalbumin (OVA)-sensitized guinea pigs, by using video-assisted microscopy to measure allergen (OVA)-induced airway constriction. OVA induced a dose-dependent airway constriction with a mean maximal effect (Emax) of 98% and a mean EC50 of 0.039 µg/ml. The arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH, 1 µM) significantly reduced the sensitivity towards OVA by >100-fold to 5 µg/ml, whereas the Emax of OVA was reduced to 50%. The effects of ABH were mimicked by the arginase inhibitor N?-hydroxy-nor-L-arginine (nor-NOHA, 5 µM). The NOS inhibitor N?-nitro-L-arginine methyl ester (L-NAME, 100 µM) largely reversed the inhibitory effect of ABH, indicating the involvement of NO. Remarkably, inhibition of arginase and/or NOS did not affect airway constriction towards exogenous histamine, the primary contractile mediator released by mast cells upon allergen challenge. This indicates that ABH reduces the histamine release from mast cells rather than the responsiveness to this mediator. In conclusion, inhibition of arginase greatly reduces airway responsiveness towards allergen by increased NO synthesis, most likely preventing mediator release from mast cells and subsequent airway constriction. Supported by the European Erasmus Mundus partnership JoinEU-SEE

Abstract The aim of this study is to examine the bioactive compounds from secondary metabolites of the fungus Rhizoctonia solani against some selected strains of pathogenic bacteria. The fungus grown in a fermented culture and the filtrate was extracted in different chemical solvents. The fungal crude extract was used to examine the bioactivity against six strains of bacteria using disc diffusion method. The results showed that the fungal extract exhibited highest growth inhibition against E. coli (34 mm inhibition zone) and S. aureus (38 mm). The minimal inhibitory concentration (MIC) values were 12.5 ug/ml and 6.5 ug/ml against E. coli and S. aureus, respectively. The fungal crude extract did not show any toxicity against human red blood. A comparison between bioactivity of the fungal extract with five commercial drugs against the tested bacteria showed that the fungal extract exhibited a significant bacterial growth inhibition than the commercial drugs. Meantime, the fungal extract showed an antitumor bioactivity by using tumor cell-line of Hep2 at low concentration (0.05 mg/ml). Also the fungal extract revealed an antioxidant action by using Fe II reduction test. Chemical analysis by using FT-IR, HNMR, and GC-Mass techniques indicated that the fungal extract composed of two main chemical compounds; Malonic acid, 2, 4-dimethyl-3-yl ethyl ester with a chemical formula C12H22O4 and Molecular weight of 230 kd and Pentanedioic,2-oxo-dimethyl ester with a chemical formula C7H10O5 and molecular weight of 174 kd. It can be concluded that the fungus R. solani possesses a potential bioactive compounds as a promising agents in medical therapy.

Osteopontin regulates hypoxic induction of NHE1 activity in H9C2 cells. Soumaya Bouchoucha, Fatima Mraiche Hypoxia, an important component of ischemia, is the result of an imbalance between oxygen supply and demand. Severe impairment of oxygen supply may result in a maladaptive cardiac phenotype. During myocardial ischemia, the Na+/H+ exchanger isoform 1 (NHE1), the main regulator of pHi, has been shown to be increased. It has been shown recently that NHE1 elevation increases Osteopontin (OPN) expression, a matricellular protein and proinflammatory cytokine that has been reported to have a protective role in ischemic injury. In this study we tested the hypothesis that hypoxic induction of NHE1 expression and subsequent alterations in H9c2 cells are mediated by OPN. H9c2 cardiomyoblasts were infected with GFP, NHE1 in the presence and absence of OPN adenoviruses following a hypoxic insult using cobalt chloride (CoCl2). Western blotting was performed to investigate the expression of OPN and NHE1. NHE1 activity was measured using BCECF-AM. H9c2 infected cells exposed to hypoxia were characterized by measuring cell viability. Inducing hypoxia for 6 hours caused a significant decrease in cell viability in H9c2 cells (GFP: 87±2.5% Vs 70.76±8.17% GFP + hypoxia; P<0.05). In NHE1 + hypoxia infected cells, viability was significantly lower than the control (NHE1 + hypoxia: 69.36±13.71% Vs 100.0 GFP + hypoxia; P<0.05). Interestingly, when overexpressing both OPN and NHE1+hypoxia, cell viability tended to increase. Compared to the control groups, CoCl2 increased the rate of recovery, which reflects the activity of NHE1, in all groups. These results were accompanied by an increase in OPN expression in the NHE1+ hypoxia infected cells (NHE1 + hypoxia: 167.36±10.01% Vs. 100.0 GFP + hypoxia; P<0.05), which validates that NHE1 activates the expression of OPN under hypoxic conditions. These results reveal a suspected role of osteopontin in protecting cell against ischemic injury owing to the increase of NHE1 expression and activity. Taken together, identified osteopontin may function as a survival factor against hypoxic induction of NHE1 cell death. Keywords: NHE1, Osteopontin, hypoxia.

Background: Most of available literature on atrial fibrillation (AF) is based on studies in the developed world and included mainly Caucasian patients. Data about AF in other ethnicities is very limited. The aim of the current study is to evaluate the effect of hypertension (HTN) in Middle-eastern Arab and South-Asian patients hospitalized with AF from a 20 year national registry in a Middle-Eastern country. Methods: Retrospective analysis of all patients hospitalized with AF in Qatar from 1991 through 2010 was made. Patients were divided into two groups according to the presence or absence of HTN on presentation. Clinical characteristics and outcomes were analyzed. Results: During the 20-years period, 3850 patients were hospitalized for AF; 1483 (38.3%) had HTN on presentation while 2367 (61.5%) had no HTN. HTN patients were 11 years older, had significantly more prevalence of diabetes mellitus, chronic kidney disease and dyslipidemia on presentation. Underlying coronary artery disease and heart failure were significantly more common in patients with HTN while valvular and rheumatic heart disease was more common in non-HTN patients. The in-hospital mortality and stroke rates were significantly higher in HTN patients (5.3% versus 3.5%, and 0.7% versus 0.2% respectively, p= 0.001) [table]. Conclusions: Our study demonstrates that in a cohort of Middle-eastern Arab and South-Asian patients hospitalized with AF, HTN is significantly associated with worse in-hospital outcomes. Our study underscores the need to study AF in different ethnicities.

Blood doping still remains as a performance enhancement strategy misused by athletes . Blood doping is the practice of boosting the number of red blood cells in the bloodstream by means of either blood transfusion of an individual or by administration of erythropoietin, in order to increase the oxygen carrying capacity of the blood. There is no direct method currently for the detection of autologous transfusion, leaving this an open issue in antidoping strategies. Recently it was reported that some proteins might represent suitable markers for in vitro aging of erythrocytes, particularly those proteins that can be detected in the membrane. Thus, this study was conducted on erythrocytes isolated from fresh and 20-day old blood samples of the same individual, with a particular focus on the erythrocyte membrane bound proteins. Isolated proteins were separated into 12 fractions on the Gelfree system, digested with trypsin and separated using a nanoLC. Eluted peptides were then spotted on a MALDI plate with MALDI spotter and detected on MALDI-TOF/TOF instrument. Afterwards, the MS/MS data for these peptides were identified using SwissProt and Mascot database search. The results have shown significant differences in the peptides when comparing fresh and stored blood samples, revealing several potential targets that can be used as markers for blood doping detection. The next step is to prove the reproducibility of the results on a larger number of specimens as an initial validation step.

Polycyclic Aromatic Hydrocarbons (PAH) are an important class of chemical carcinogens formed as a result of incomplete combustion; it is therefore found in grilled foods, tobacco smoke and in general, the environment. Previous studies have shown exposure of smokers and second hand smokers (SHS) towards a whole host of PAHs and more than 500 of these have been demonstrated in tobacco smoke. As it is ubiquitous in the environment, exposure may result from various sources; for cigarette smokers a number of these PAHs markers have been correlated to tobacco smoke exposure while others like benzo(a)pyrene has been associated with environmental and occupational exposures. Tobacco smoke from narghile or "shisha" has been shown to contain approximately 50 times more PAHs compared to cigarette smoke; this phenomenally high exposure can significantly increase risks of narghile smokers towards the toxic carcinogenic effects of this class of compounds. In this study, the smoke condensates in shisha water was extracted and analysed for 16 targeted PAHs using Selected Ion Monitoring (SIM) mode on the gas chromatograph mass spectrometer (GCMS). As part of the establishment of a validated quantitated method, the LOD (limit of detection), LOQ (limit of quantification), linearity, reproducibility and precision as well as robustness were determined for all 16 PAHs. The PAHs were quantitated in 20 shisha water collected from 20 different locations; as PAHs in shisha water may be directly related to the amount of charcoal and flavourings as well as amount of condensate, the profiles were found to be vastly/relatively different. Included is a discussion of these relative differences and their implications for shisha users. Finally, preliminary work is presented for the discovery of more toxic nitrated PAH in the shisha water condensate.

Background: Individuals with diabetic peripheral neuropathy (DPN) frequently suffer from concomitant impaired proprioception and postural instability. Conventional exercise training has been demonstrated effective in improving balance; however these exercises are often repetitive, causing patients to lose interest and not complete the long term rehabilitation process. In addition, these programs do not incorporate visual feedback targeting joint perception, which is an integral mechanism that helps compensate for impaired proprioception among DPN. Methods: This is a prospective randomized control trial study. Participants were randomized either in intervention or sham. The intervention group received twice per week an innovative exercise program based on virtual reality for 4 weeks. Their gait and balance were assessed at baseline prior initiating the intervention and after 4 weeks. Sham group were not included in balance training but their balance and gait were assessed at study visit and 4 weeks later. An innovative exercise program based on combination of wearable sensors technology and virtual reality was designed. The wearable sensors allow measuring lower extremity joint position in real-time which were used for the purpose of animation and real-time visual feedback to subject during exercise. Exercise program includes a series of ankle point-to-point reaching tasks in different direction as well as crossing a series of virtual obstacle with different heights, which were appeared on a computer screen in front of participants, Figure 1. Results: Forty-one eligible subjects have been recruited to date. However, the results of 15 participants (Age: 56.3±4.9, BMI: 30±15 m/Kg2) who completed the exercise program have been reported. Participants were diagnosed with diabetes by primary care and were confirmed to have peripheral neuropathy. The preliminary results suggest that active group reduced ankle sway by 76% (2.82±2.8deg to 0.66±0.47deg), hip sway by 81% (7.96±9deg to 1.48±1.2deg) and center of mass (CoM) sway by 76% (0.69±0.7deg to 0.16±0.11deg) during eyes open balance assessment. Similar reductions during eyes closed assessment were observed with reductions of 50%, 24% and 45% for ankle, hip and CoM sway. Conclusion: The current research implemented a novel balance rehabilitation strategy based on virtual-reality. Our methodology included wearable sensors and interactive user interface for real-time visual feedback based on ankle joint motion, similar to video gaming environment for compensating impaired joint proprioception. Findings support that visual feedback generated from ankle joint coupled with motor learning may be effective in improving postural control and gait among DPN patients.

Endometriosis is a debilitating medical condition in females in which endometrial glands and stroma appear and flourish in areas outside the uterine cavity and walls, most commonly in the ovaries. Incidence in general population is 5-15% and around 30% in infertile women. Usual age group is 20-30years and patients present with dysmenorrhoea, dyspareunia, dyschezia, infertility and pelvic pain. We report two interesting cases of stage IV endometriosis who presented as bilateral complex ovarian masses which simulated ovarian cancer on ultrasound. MRI pelvis for both these cases diagnosed stage IV Endometriosis.

Type I interferon α/β (IFN-α/β) cytokines are produced by the host cell upon sensing viral proteins and nucleic acids as pathogen associated molecular patterns (PAMPs) and thereby elicit an antiviral response that represents the frontline of innate immunity against a virus infection. Among the signatures of the "non self" presence, double-stranded RNA (dsRNA) is the most potent PAMP in triggering the IFN-α/β-based antiviral response. dsRNA is synthesized during the course of infection by RNA viruses as a byproduct of replication and transcription, or it may be originated through the panhandle self-pairing occurring at viral genome ends. dsRNA is targeted by specific pathogen recognition receptors (PRRs), of which the class of cellular helicases termed as RIG-I-like receptors (RLRs) is the subset deputed to its prompt detection in the cytosol. This is the reason why avoiding dsRNA recognition by RLRs is a strategy adopted by RNA viruses to inhibit the IFN-α/β induction. Moreover, in most cases, such circumvention of the innate immune antiviral response relies on the function of virally-encoded proteins that act binding viral dsRNA to impede its interaction with RLRs. To date, proteins displaying dsRNA binding-mediated IFN-antagonism have been described for several RNA viruses that are highly lethal to humans. Among these there are emerging pathogens that recently affected Middle East and the Arabian Peninsula, such as influenza A viruses and - putatively - also the newly identified SARS-related coronavirus. Therefore, given the role of IFN-inhibiting proteins as key determinants of virulence and pathogenesis, characterization of their dsRNA binding function is of urgent need, and disruption of their interaction with viral dsRNA is an attractive and promising target for the development of effective antiviral agents. Within this picture, we have recently developed a new in vitro assay, namely the Magnetic Pull Down (MPD), as a method for measuring dsRNA binding activity of viral proteins. Relying on the properties of the paramagnetic TALON Dynabeads, an His-tagged recombinant protein can be stably coated to beads in this assay, and subsequently incubated with a labeled dsRNA substrate. Next, application of a magnetic field allows the precipitation and further isolation of a quantifiable dsRNA:protein complex. By using as a paradigm model a full length recombinant version of the Ebola virus VP35 (EBOVrVP35) protein - a potent dsRNA binding-dependent IFN-α/β suppressor - we characterized its dsRNA binding function and validated the MPD assay for antiviral screening. Results showed that EBOV rVP35 binds to 3H-radiolabeled in vitro transcribed dsRNA molecules of different length (500-50bp) with very high affinity and KD values that are within the low nanomolar range. Moreover, the formation of EBOV rVP35:dsRNA complex was inhibited in this assay by using the tester compound auryntricarboxylic acid, which showed an IC50 value of 50μg/mL. In summary, the MPD assay herein described provides a straightforward tool i) to identify viral IFN-antagonists displaying dsRNA binding activity; ii) to quantitatively characterize dsRNA binding function by measuring the dsRNA:protein complex formation and iii) to screen for antiviral agents inhibiting such interaction, either they are synthetic compounds or natural herbal extracts.

Introduction: The objective of the "Italian Study on gene-environment interactions in lymphoma etiology", hereafter named the Italian GxE Study, is to identify individual conditions and external factors associated with lymphoma aetiology, the response to therapy and survival, as possible targets for preventive action and possible drivers of new individualized therapeutic strategies. Methods: This objective is being pursued by recruiting up to 1000 incident lymphoma cases and 1000 controls in six Italian areas: central and southern Sardinia, Florence, Bari and Taranto, Verona, Novara and Perugia. After signing the informed consent form, cases and controls will donate a blood sample and respond to a detailed questionnaire on health history, lifestyle factors, and occupational and environmental exposures of interest. The interaction between genetic polymorphisms, epigenetic conditions, and the most prevalent lifestyle and occupational exposures will be explored using the case-control epidemiological study design. Survival analysis will be conducted in relation to the therapeutic protocol, lifestyle variables, and gene polymorphisms using a modified proportional hazard model. Study design: Here are the main features of the Italian GxE study: 1. Genome Wide Scan of patients and controls DNA, which aims to evaluate several million SNPs, some functionally known, and other in strict linkage disequilibrium with genes in loci encoding for known proteins, using last generation high throughput platforms (Illumina (R) Human660W). 2. Use of the Telepathology Network to reach maximum consensus among the pathologists' panel collaborating in this project so achieving the maximum diagnostic precision, by applying the most recent lymphoma classification schemes consistently across the study areas. 3. Detailed assessment of occupational and environmental exposure, thanks to the experts in retrospective exposure assessment participating in our project and the continuous refinement of the questionnaires which have been used in international studies for the last two decades. Precision in defining external exposures is of paramount importance to effectively explore gene-environment interactions in the aetiology of the different lymphoma subtypes. 4. Analysis of response to therapy and survival of lymphoma patients in relation to their genetic features, which will allow to design future individualized therapeutic schemes, by identifying responders to specific chemotherapeutic agents based upon their polymorphisms of genes implicated in xenobiotic metabolism, DNA repair, release of inflammatory cytokines, as well as other yet to be identified. Conclusion: The Italian GxE Study contributes to the International InterLymph Consortium (http://epi.grants.cancer.gov/InterLymph/), which creates the necessary synergy to inquire into lymphoma etiology in general, and its individual subtypes in particular.

الفكرة هي اعتبار عقل الطفل لا يوجد به البرنامج التشغيلي ي للعقل أو أن برنامج التشغيل الأولي لا يعمل كما يجب وهو فرضية أن عقل الطفل عندما يولد يكون عقله محمل ببرنامج مكتسب من الجينات وضعه الخالق سبحانه وتعالى نصه * انتبه إلى ما هو مكرر في بيئتك * انتبه إلى المثيرات البصرية التي تتكرر * انتبه إلى المثيرات السمعية التي تتكرر * سوف تتعرف على نفسك ومن حولك في حالة الطفل تم إهماله في الشهور الأولى مثل ترك الطفل ليشاهد قنوات الأطفال في أول 6 أشهر أو أن الدماغ لم يحفز فيصبح اقل نشاطا في السنوات اللاحقة فيكون اهتمام العقل بالأشياء التي اعتاد عليها فقط مثل اللعب الفردي بدون هدف الاهتمام الأكبر بقنوات الأطفال مع عدم الاهتمام بأي جديد وتكون سرعة التعلم بطيئة جدا بدليل انه لا يستجيب لنداء أمه 80 في المية من أطفال التوحد هم حالات طيف توحد ناتج من ترك الطفل أمام التلفزيون في العام الأول أو عدم تفاعل مع الطفل مثال تركه لخادمة غير عربية وبذلك يحدث خلل في البرنامج التشغيلي لعقل الطفل وهو ما يجعل الطفل كأنه فاصل عن الشبكة الخارجية لا يستجيب للنداء لا يطيل النظر ينعزل - وهذه الطريقة تصلح بنسبة مرتفعة بمثل هذه الحالات الطريقة هي وضع برنامج لعقل طفل التوحد يختلف هذا البرنامج باختلاف ما ينقصه لإعادة تحفيز البرنامج الأصلي الناتج من جينات الطفل هذا البرنامج يساعد على أن يعرف الطفل نفسه من هو ومن حوله ويبدأ من الصفر بداية التعلم ولكن بشخصية أخرى ويصبح على الفطرة طريقة البرمجة سريعة و مشجعة بفضل الله وسريعة قد تكون هذا العام احدي الطرق الرئيسية لكسر سمات التوحد في العالم طفلا عمره الزمني 4 سنوات وعمره العقلي 2 سنه سيبدأ إعادة تشغيل للعقل بعمر سنتين وكأننا قمنا بإعادة تشغيل الجهاز reset ويقوم بإعادة تحفيز البرنامج الأصلي فينتبه إلى ماهو مكرر سينتبه إلى المثيرات البصرية والسمعية وتنشط جميع الحواس ذاتيا فيبدأ العقل بضبط كيماويا ت العقل وهرمونات الجسم ذاتيا فيبدأ بالتعلم ويقود الدماغ الى اكتساب المعرفة وتختفي تدريجيا أعراض سمات التوحد وتزداد سرعة التعلم ويجب تكثيف برامج التعلم له ليستطيع تعويض ما فاته الاستجابة لبعض الأطفال في الأسبوع الأول حتى مشاكل الهضم سواء إمساك أو إسهال أما إذا كان العقل لا يحتوي على البرنامج التشغيلي أصلا نتيجة خلل ما أو نتيجة لمرض وراثي أو كان سبب التوحد خلل في الجينات نتيجة تعرض الأب أو الأم لأدوية قبل الحمل أو أن مخ الطفل لم يكتمل فطريقة البرمجة إلى الآن لم تعطي نتائج طريقة العلاج إرسال الاسئلة لمعرفة سلوك الطفل في العام الأول الأدوية المستعملة التشخيص بعد الإجابة على الأسئلة ومعرفة هل تصلح الطريقة أم لا يتم كتابة البرنامج الأول ويرسل إلى الأسرة يتم تسجيل البرنامج بصوت الأب والأم و إرساله لنا نقوم بعمل فلترة و دمج ذبذبات مكافئة لموجات الدماغ الثيتا سواء كانت طبيعية أم من أجهزة مع دراسة مراحل النوم وإعادة الملف الصوتي ليسمعه الطفل في أوقات محدده منها أثناء النوم المميزات تصلح لجميع اطفال العالم تم التطبيق على طفل مقيم بأمريكا على طفل مقيم بايطاليا باللغة الايطالية وتسجيله بصوت الام تم تطبيق البرنامج على طفل كردستان العراق تم التطبيق على عشرات الاطفال من السعودية ومصر السودان المغرب تونس لا نحتاج الى رؤية الطفل

Background & Objectives Aneurysm is a condition in which an abnormal dilation occurs in the wall of an artery or vein. Since cerebral aneurysm morphology is considered as a potential surrogate of rupture, clinicians often desire its segmentation before any pre-operative or interventional planning is performed. Also, segmentation of such images offers the potential of identifying the right treatment for a wide range of medical conundrums. This paper presents a simple and fast automatic algorithm based on active contour technique for cerebral aneurysm segmentation. Since the performance of active contour is highly dependent on the placement of the initial contour, the objective of this work is to automate this placement process to get the desired contour quickly. Methods: The active contour approach is a prime candidate for practical exploitation because active contours make effective use of specific prior information about objects and this makes them inherently efficient algorithms. In this work, we utilize gradient vector flow (GVF) snake, which is an extension of the active contour because it efficiently converges to boundary concavities. However, computational time remains a concern; therefore, we try to place the initial contour as close to the desired boundary of the object in the medical image as possible. For this, we use Otsu's adaptive thresholding method that is based on the histogram of the intensity distribution of the input image. As a post-processing step, the extracted contour is smoothened using linear regression. Results: The proposed method has been implemented on angiography datasets obtained from two different sources. The angiography CT datasets are collected from University of College of London and Hamad Medical Corporation (512x512x405 pixels at circa 0.5mm resolution). The original, ground truth and segmented intracranial aneurysms are shown in Figure 1, which clearly reflects the potential of the proposed algorithm. The average time taken for segmentation is 2 s per slice (without optimization) on MATLAB 7.5 on a PC with Pentium 4, 3 GHz dual core processor. Conclusion: Although our work might be a simplified approach, we envision that it could represent a necessary first step towards improving the performance of active contour that is dependent on the initial contour in a clinical setting.

Spinal muscular atrophy (SMA) is an autosomal recessive disease affecting mainly the alpha motor neurons in the ventral horn of the spinal cord leading to muscular atrophy and paralysis. Although SMA is one of the leading genetic causes of infant mortality, no effective treatment is currently available. Gene delivery studies of Glial Derived Neurotrophic (GDNF) in Parkinson&#39;s disease (PD) have demonstrated positive impacts in vitro studies, PD models and early phases of clinical studies. In light of that, the objective of this work is to assess the efficiency of GDNF in preventing the neuromuscular pathology in SMA. Our specific aims are to: i) generate GDNF expressing vectors for in vitro and in vivo transduction; ii) Evaluate the impact of GDNF on axonal growth in SMN-deficient motor neurons in vitro; iii) generate a pre-clinical efficacy proof-of-concept in the SMNΔ7 mouse model of SMA. Because lentivirus (LV) is known to be efficient for in vitro transduction, an LV‐SIN‐PGK‐GDNF plasmid was used to express human GDNF. In order to assess the efficacy of the produced viral prep, it was later used to transduce HEK293T cells. Preliminary analysis of protein levels of the GDNF using western blot and ELISA showed an increase in the LV-GDNF transduced cells in comparison to un-transduced cells. Experiments are ongoing to evaluate the effects of the LV-GDNF on the axonal growth in SMN-deficient primary motor neuron cells. For in vivo studies, self-complementary adeno-associated virus of serotype 9 expressing GDNF gene (scAAV-GDNF) was produced as scAAV9 has been shown to have low pathogenicity and remarkable capacity to transduce both neurons and astrocytes in vivo. Preliminary transfection and transduction assays have shown increase in GDNF expression in the HEK293 transfected and HELA transduced cells. However, further work is being performed to confirm these results. Also, pre-clinical efficacy proof of concept will be performed using SMNDelta7 mouse model via scAAV-GDNF injections. This is going to be followed by evaluating the impacts on the survival and the phenotype of the injected mice. To this end, this work represents an essential step to validate the ability of the LV-GDNF and the scAAV-GDNF to transduce cells and produce the desired GDNF. Ongoing work will give insights on the efficacy of using GDNF gene delivery as a therapeutic approach for SMA.

Cancer develops as a result of a defective apoptotic system. Anticancer drugs mainly aim to modulate apoptosis. Multiple publications strongly support the link between the elevation of the intracellular calcium concentration and the induction of apoptosis. Cisplatin was one of the pioneers of the metal based anticancer drugs. Many tumors have been treated with cisplatin and other platin-based anti-cancer drugs like carboplatin, and oxaliplatin. Over the decades metal-based anti-cancer treatment has come a long way with the introduction of newer drugs with less cytotoxicity and different biologic targets. A gold-based complex, Auranofin was introduced to treat rheumatoid arthritis but is also a successful anti-cancer drug. Auranofin is currently undergoing clinical trials in treating patients with epithelial ovarian, primary peritoneal, or fallopian tube cancer. Here we test the induction of apoptotis by Auranofin on MCF-7 breast cancer cell-line and compare this results with the elevation of the intracellular calcium concentration. Methods Breast cancer cells obtained from ATCC were grown in DMEM containing 10% FBS at 370C in a carbon dioxide incubator. Varying concentrations of the compound ranging from 0.1µM to 10µM were tested. Intracellular calcium was recorded in cells treated with different concentration of the drug and a dose response for the apoptosis and cell viability was established. Calcium channel modulators were applied to see the source of calcium upon drug treatment which may help in improving the efficiency of the drug. Cell viability was tested by a standard MTS assay after 24 hours of treatment with Auranofin. Parallel, apoptosis and necrosis was measured by flow cytometry, here the cells were stained with 7 AAD and Sr Flica. Cells for calcium imaging were loaded with the calcium sensitive dye Fluo-4 AM. Fluorescence of the selected cells were recorded every 1min for 2 to 3 hours. To see the source of intracellular calcium rise, calcium channel modulators 2-APB, caffeine, cyclosporine a mitochondrial permeability transition pore inhibitor were used. ROS was monitored by spectrophotometry using fluorescent dye di hydro calcein, Results MTS assay showed a concentration-dependent decrease in cell viability. IC50 was below 6.25µM. The flow cytometry analysis showed four different population of cells, viable, early apoptotic, late apoptotic and necrotic. 30% higher or more apoptosis were observed in concentration above 6.25µM. Apoptosis was 98% when higher concentrations were used. Necrotic cells remained below 16% in all the concentrations. Calcium imaging experiments showed a time and concentration dependent elevation of the intracellular calcium concentration. After 2 hours of application fluorescent intensity corresponding to intracellular calcium increased for concentrations 0.1µM, 1µM, 5µM and 10µM respectively (given 100% as control). Viability proportionally decreased when intracellular calcium was elevated. Blockers targeting PM, ER and mitochondria such as caffeine, cyclosporine 2-APB, nimodipine showed no significant changes. It was noticed that 1mM melatonin was able to decrease the ROS production induced by Auranofin. Conclusion: Auranofin induces apoptosis and cell death. The triggered concentration dependent increase of intracellular calcium might be a crucial factor for the induction of apoptosis. More experiments are required to establish the source of calcium.

The computational advances in recent times have led to analysis of the complex interaction of blood flow through elastic artery. This aids in prognosing the progression and behaviour of cardiovascular diseases like atherosclerosis, aneurysms etc. In the present investigation, an idealistic 66% eccentric stenosed common carotid artery is analyzed under normal blood pressure condition based on clinical data obtained from ultrasound measurements and 3D FSI model is generated in ANSYS, commercially available Finite Element Analysis software. The blood flow is assumed to be incompressible, homogenous and Newtonian, while artery wall is assumed to behave linearly elastic. The two-way sequentially coupled transient FSI analysis is performed using FSI solver in ANSYS-10.0 for three pulse cycles and haemodynamic parameters such as flow pattern, Wall Shear Stress, pressure contours and arterial wall deformation are studied at throat and downstream of the stenosis. Further, the flow changes with varying the severity of stenosis is also attempted to investigate the changes in flow behaviour by varying the percentage of stenosis in terms of 75%, 80%, 85%, 90% and 95%. Comparison of results concludes that, with the increase in severity of stenosis, the flow changes abruptly causing a significant increase in velocity and WSS at throat region, while highly complex flow is observed in the downstream of stenosis leading to formation of eddies. During flow deceleration, flow is further highly turbulent due to the effect of back pressure and considerably altering the flow behaviour in the downstream end. The results obtained agree very well with available clinical observation during various stenosed conditions.

Purpose: Pancreatic ductal adenocarcinoma (PDAC) has one of the worst prognoses of all malignancies. At time of diagnosis, 80% of patients with PDAC have unresectable tumors, and conventional therapies are nearly ineffective. The relative poor efficacy of chemo/radiotherapy against PDAC indicates that the development of new therapeutic strategies is a crucial step to improve the survival of patients with this disease. This includes the modification of established therapies, the testing of new targets and the improved delivery of drugs to their target. Riproximin (Rpx) is a new plant agent, which was isolated from the plant Ximenia Americana. Riproximin was classified as a ribosome inactivating protein (RIP) of type II and its anticancer activity has been recently demonstrated in vitro and in vivo Methods: In vivo experiments The rat pancreatic cancer cell line (ASML) was used for its property to mimic liver metastasis in nude rats, as shown before (Eyol et al). Tumor-bearing rats were treated intravenously with different concentrations of Rpx as single agent or in combination with gemcitabine (GEM) or dinaline (DIN). Ripx was administered twice weekly at doses ranging from 500&#181;g/kg to 1500&#181;g/kg. GEM and DIN were administered to animals for a period of 2 weeks either intravenously at a dose of 50mg/kg or perorally at a 5 times weekly dose of 10mg/kg. 98 representative apoptosis genes were analyzed by quantitative-RT/PCR in with Rpx treated cells in comparison to untreated cells. Results: The intravenous administration of Rpx (1500&#181;g/kg and 500&#181;g/kg) reduced the tumor growth significantly by 61% and 67%, respectively (p=0.00174 -p= 0.00024). The survival rate was also significantly increased (21.8 days for riproximin treated versus 17.6 days in untreated control rats; P=0.01349) after the intravenous administration of Rpx. Rats, which were treated with the high dose of Rpx showed no further reduction in tumor size, when compared with rats treated with the low dose. In comparison, no significant effect on tumor size or on survival was observed after treatment with gemcitabine or dinaline. The combination therapy with Rpx and GEM as well as Rpx and DIN produced a significant reduction in mean tumor size when compared with the corresponding untreated control group but it was not superior over the single therapy with Rpx. 17 apoptosis gens have been found to be modulated after exposure to Rpx. Conclusion: Taken together the results of our in vivo experiment suggest that riproximin has a clear potential for pancreatic cancer treatment. Further experiments are required for optimizing the combination therapy with other antineoplastic agents.

Patient engagement in their care process, vis-&#224;-vis self-management programs is an important element of the patient&#39;s longitudinal care plan, where the patient is encouraged and expected to achieve self-efficacy in the self-management of the disease through a regime of educational and behavioral modification strategies. To ensure the effectiveness of self-management programs, it is important that the proposed self-management interventions are (a) personalized to the unique needs and constraints of the patient; (b) based on sound theoretical health models; (c) based on validated health and behavior assessment tools to determine the patient&#39;s physical and behavioral dispositions; and (d) readily accessible to the patient through a ubiquitous medium, such as smart phones or the web. In this paper, we present a novel personalized self-management framework that delivers personalized health educational interventions to empower, educate and engage patients/individuals through self-observation, barrier identification, goal setting and action planning to achieve behavioral self-efficacy and self-regulation so that individuals can self-manage their condition. Our personalized self-management framework is guided by Social Cognition Theory, whereby have ensured that self-management programs for chronic disease management not just focus on changing the patient&#39;s awareness of the disease, rather they focus on enhancing the ability of the patient to make the right choices to achieve effective disease management. We present a three-stage personalized self-management framework that comprises: Stage 1: A high-level characterization of an individual with respect to a specific health outcome using validated assessment tools; Stage 2: A behavioral categorization of the individual based on his/her levels of self-efficacy, motivation and self-regulation, etc.; Stage 3: Use the personalized profile of the individual to tailor generic educational and self-management to develop a personalized self-management program that comprises personalized strategies to counter the challenges and barriers faced by the individual to achieving positive self-efficacy and self-regulation which in turn will lead to positive health outcomes. Our personalized self-management framework features (a) a novel self-management oriented individual profiling mechanism that takes into account both the health and psychosocial characteristics of an individual to generate his/her holistic profile; (b) a semantic web based knowledge model that captures the theoretical foundations of the SCT in terms of a Self-Management Program Personalization (SPP) ontology; (c) a semantic web based personalization tool that uses a logic-based execution engine that reasons over the SPP ontology, based on an individual&#39;s profile, to generate personalized self-management interventions; and (d) a mobile messaging platform to deliver the personalized self-management interventions and to monitor the patient&#39;s compliance using smart phones. We take a semantic web approach in designing the personalization approach whereby we have developed the SPP ontology to (a) model the theoretical framework of SCT in terms of SCT concepts; (b) model health assessment tools; (c) model the personalization rules that integrate the health and SCT models with the educational messages to generate a personalized self-management program. We have demonstrated the novel integration of health models, educational content and behavior change strategies to design self-management programs for cardiac risk factors, where the program is delivered through a mobile app.

Background: People with diabetes are at risk of developing foot ulcers (DFUs), which accounts for significant morbidity and mortality. Damage caused from repeated foot loading and temperature changes during walking may go undetected by patients due to loss of lower-extremity sensations (peripheral neuropathy). Clinical assessment on the other hand needs considerable time. Therefore, a feasible assessment of pressure and temperature is vital to measure pre-ulcerative inflammation and predict DFUs. Current study is aimed to validate effectiveness of an innovative smart textile based on fiber optics which allows measuring simultaneously plantar pressure, plantar temperature, and lower extremity joint angles. Methods: The proposed technology is based on optical fiber glass that propagates of light. Multiple sensors were juxtaposed on the length of the fiber which was integrated in a comfortable sock -SmartSox (Novinoor LLC, IL, USA) - Figure 1. Based on changes in wavelength of light, mechanical changes in environment including changes in temperature, pressure and joint angles are measured. The sensors were integrated in anatomical regions of interest including heel, mid-foot, 1st and 5th metatarsal heads, and under and over big toe. The later sensor allows measuring hallux range of motion in addition to temperature and pressure. To validate the accuracy of the designed prototype, 21 volunteers diagnosed with diabetes were recruited. Subjects were asked to walk a predetermined route of 200 steps in their normal or prescribed shoes. Plantar foot thermal images were taken by a thermal camera system twice: once at baseline after a five-minute temperature acclimation period and once after each walking trial. To determine the number of steps taken, subjects were asked to wear a gait analyzer system (LEGSys™, Biosensics LLC, MA, USA) in addition to SmartSox. To validate the accuracy of plantar pressure, a computerized pressure insole, F-scan (TekScan® Inc, MA, USA) was used as a gold standard. Results: Twenty-one patients with diabetes were recruited (Age: 57.8±7.9 years, BMI: 31.6±8.0 kg/m2, VPT: 26.8±15 volt, 68% diagnosed with peripheral neuropathy). All subjects perceived the SmartSox as comfortable and no problems were observed during walking assessment. Some fibers were however broken during wearing the socks in particular for patients who worn removable cast walker. A significant correlation was observed between the pressure profile measured by SmartSox and F-Scan under different anatomical regions of interest (r>0.6, p<0.05). A similar agreement between SmartSox and thermal camera was observed for changes in plantar temperature during walking. Conclusions: This study demonstrates the proofs of concept of an innovative smart textile for assessing simultaneously the key parameters associated with risk of foot ulcer in patients with diabetes. Fragility of the current prototype is considered as one of its major weakness for routine clinical assessment and thus further improvement in design of fiber and sock is needed. Additional study is required to address whether the measured parameters can be used to predict and better management of diabetic foot ulcer.

Modulation of calcium homeostasis plays a key role in regulating fundamental cellular processes, including gene transcription, cell proliferation, differentiation, and apoptosis. Store-operated calcium entry (SOCE) is the major pathway in non-excitable cells for extracellular Ca2+ influx across the plasma membrane and is regulated by the content of the intracellular Ca2+ stores. Following store depletion, the Ca2+ sensor in ER, stromal interaction molecule 1 (STIM1) clusters in ER regions close to the plasma membrane and recruits Orai1, which is a Ca2+ selective channel resulting in SOCE. Despite the significant knowledge in understanding STIM1 subcellular distribution dynamics, little is known about the trafficking of Orai1. Our laboratory previously showed that in Xenopus leavis oocytes Orai1 shuttles between plasma membrane and endosomal compartments, and that it internalizes during meiosis. However, the subcellular distribution and trafficking of Orai1 in mammalian cells is not fully understood. In this study we show that at steady state around 46% of Orai1 is on the surface of the plasma membrane of CHO cells, while the remaining 54% localizes intracellularly, suggesting that Orai1 constitutively recycles between the two compartments. Our time lapse imaging shows continuous shuttling of Orai1 to and from the PM with an exocytosis rate T1/2 of around 5 minutes. We also measured the endocytic rate of Orai1 at 5% min-1. To identify the domain within Orai1 required for its trafficking, we generated deletions of either the N- or C-terminus of Orai1. Deletion of N-terminus does not affect Orai1 trafficking to the cell membrane in CHO cells. In contrast deletion of the whole C-terminus (257-301) significantly altered Orai1 trafficking to the cell membrane. Deletion of amino acids 285-301 of Orai1 C-terminus does not have any impact on Orai1 trafficking to PM, suggesting that the information necessary for Orai1 trafficking to PM is located in the segment 257-285 of Orai1 C-terminus. In this study we determined, for the first time, the percentage of Orai1 on PM of mammalian cells and showed through time lapse-imaging that it constitutively recycles between PM and intracellular compartments. Our results also suggest, for the first time, that the amino acid region (257-285) of Orai1 C-terminus is essential for its plasma membrane trafficking.

Gastric bypass surgery is the most effective treatment for obesity, achieving both sustained weight loss and improved insulin sensitivity. However, following Roux-en-Y gastric bypass surgery (RYGB) some patients develop debilitating nausea and vomiting (N/V) persisting for years. The aim of this study was to determine if the N/V following RYGB is due to elevated systemic GLP-1 levels and whether GLP-1 directly mediates secretion of adipokines, such as leptin. 42 female non-diabetic subjects were studied in the fasting and post-prandial state and divided into 5 groups according to BMI and presence of N/V post-operatively. The effect of GLP-1 on adipose tissue leptin secretion in vitro was measured. Subjects with N/V post-RYGB surgery had significantly elevated fasting GLP-1 levels compared to post-operative subjects without N/V symptoms, and to morbidly obese (MO), obese and lean subjects not undergoing surgery. However, weight loss, as well as systemic levels of glucose, insulin and post-prandial GLP-1 was similar in all post-operative subjects. Despite similar BMI (P = 0.86) and fasting adiponectin levels, leptin was significantly lower in subjects with N/V compared to those without N/V (P = 0.04). Leptin secretion from subcutaneous adipose tissue in vitro was significantly inhibited by GLP-1 (0.1-1.0 nM; n = 6). Persistent N/V following RYGB surgery is associated with elevated fasting GLP-1 and lower leptin levels, perhaps as a consequence of GLP-1 mediated inhibition of leptin secretion from adipose tissue. GLP-1 antagonists may alleviate these symptoms, but, adverse effects on weight maintenance and insulin sensitivity need to be considered.

Epithelial ovarian carcinoma (EOC) is an aggressive neoplasm that mainly metastasizes to organs of the peritoneal cavity. This event is mediated by molecular mechanisms that remain elusive. Cell adhesion molecules play a key role in tumor invasion, metastasis and drug resistance. The conventional in vitro two-dimensional cell culture models are not sufficient to explain the exact mechanism behind tumor invasion, migration and anoikis resistance events. The objective of the present study is to analyze the role of cell adhesion molecules in tumor invasion metastasis and drug resistance using multiple phenotyping approach. Five ovarian cancer cell lines PA1, SW626, CAOV3, SKOV3 and OVCAR3 were selected for the study. Cell line phenotyping was conducted using cultured cells in an anchorage independent method that utilized an ultra low attachment plate for mimicking anoikis resistance in vitro. Second type phenotyping was done for sorting highly invasive phenotype by selecting cells that can pass through Boyden chamber (8 micron pore size upper chamber membrane). Development of drug-resistant cell lines were achieved by growing cells in culture media containing standard chemotherapeutic agents such as Taxol and Carboplatin. Cells were selected according to their ability to attach different ECM and cell adhesion molecule coated chambers. A real time PCR array of 29 genes involved in cell adhesion, drug resistance and EMT (epithelial-mesenchymal transition) were also analyzed and gene expression analysis conducted. The invasive potential of PA1 (teratocarcinoma) seems to be higher than other cell lines followed by SKOV3 cells. PA1 cells form embryoid bodies while culturing in in anchorage independent condition and exhibit an elevated level of expression of myc and TGF beta. Cell viability assay also shows that PA1 is the most sensitive cell line against carboplatin and taxol. Gene expression levels of cell adhesion molecules were altered among the phenotypes. Anoikis resistant cells show altered levels of expression in EPCAM, Collagen 6, CD24, vimentin and N-cadherin. These results suggest the cell lines are heterogeneous in nature and three dimensional culture model mimics the in vivo tumor model for anoikis resistance and EMT. To conclude, this study shows selection of various phenotypes of heterogeneous cancer cell lines can help decipher the role of cell adhesion molecules in ovarian cancer invasion and drug resistance. These experiments will give an overall view of the role of cell adhesion molecules in different ovarian cancer types in vitro.

Asperger syndrome is one of the Autism Spectrum Disorders (ASD's), characterized by significant difficulties in social interaction and nonverbal communication, alongside restricted and repetitive patterns of behavior and interests. ASD's have a strong and complex genetic basis that cannot be distinguished by the clinical presentation. A South African family with an affected father and three affected sons all with characteristics of Asperger syndrome including repetitive routine physical gestures and Sensory Processing Disorder was studied for the possible identification of the responsible genetic factor(s). Due to the significant proof of heritability and the extreme heterogeneity of Asperger syndrome, next generation sequencing was performed on all members of this family to extrapolate monoallelic variations in the affected father that have been inherited by all three of the affected sons probably through an autosomal dominant pattern of inheritance. These variations, validated by Sanger sequencing, were analyzed, prioritizing significant changes in the encoded protein. Variants that segregate with the affected individuals include one deletion in C17orf80, an unidentified protein expressed in the brain, (c.1745_1748delGTAA), three missense mutations that change highly conserved amino acids in PARK2, which codes for a component of a multiprotein E3 ubiquitin ligase complex that targets proteins for degradation also known to cause juvenile Parkinson disease (c.110 C>T, p.Pro37Leu), FAT1, member of a large cadherin family required for cell-cell association and actin organization, (c.2563 C>A p.Gly855Arg), and OR4C6, an olfactory receptor protein coding gene, (c.293 A>C p.Gln98Pro) and one nonsense mutation introducing a premature stop codon in HYAL4, a hyaluronidase that intracellularly degrades hyaluronan, one of the major glycosaminoglycans in the extracellular matrix (c.628 C>T p.Arg210STOP). The direct link of these previously reported variants to Asperger syndrome is yet unknown, however some of these genes such as PARK2, HYAL4 and FAT1 have previously been reported to be in involved in brain function and development, indicating a possible role in the onset of Asperger syndrome.

This study evaluated the effectiveness of paravertebral block as alternative anesthetic technique for extracorporeal shock wave lithotripsy (ESWL) procedure. Fifty patients with renal stones, aged 20 to 60 years, were randomly allocated into two groups; twenty five patients in group P; received unilateral paravertebral block from T8 through L1 with injection of 5ml 0.5%bupivacaine and 25 patients in group L; received local infiltration by bupivacaine 0.25%(2mg/kg) into the 30 cm2 area after localizing the stones site, 10 minutes before the session. 10 mm visual analogue scale (VAS) was used to evaluate pain every 10 minutes during the session. At the end of the procedure, total doses of rescue analgesia, the number of shockwaves, their power and the total duration of shockwave treatment were recorded. After completion of the procedure the patient was assessed for pain and nausea in the post anesthesia Care Unit (PACU) using the Visual Analog Scale. Patient's satisfaction and time needed to discharge patients to home also were recorded. Time to do the anesthetic technique was significantly higher (p<0.001) in group-P than group-L, it was 12.7±2.3 minutes versus 6.9±1.9 minutes respectively, intraoperative rescue analgesia by fentanyl was lesser (P<0.001) in group-P than group-L, 26.7 ± 6.32mcg versus 78.6±5.41mcg, respectively, also time interval between ends of the procedure till discharge to home was significantly higher (P<0.001) in group-P than group-L, it was 99±17 minuets versus 133±31minuits respectively. VAS was not significant difference between both groups either intraoperative or postoperative in first hour. Patient's satisfaction was significantly higher (P<0.05) in group-P than group-L, it was 8.8±1.1 versus 6.1±0.6, respectively. Adverse events were lesser, but not significant in group-P than in group-L. Two patients (8%) in group-L and one patient (4%) in the group-P experienced episodes of postoperative nausea and vomiting (PONV). Paravertebral block is effective alternative anesthesia for outpatient lithotripsy; multiple level paravertebral blocks provide an optimal anesthetic condition, with acceptable adverse events for ESWL. And providing proper analgesia during the procedure and in first hour after finishing of the procedure, early discharge to home and providing better patient's satisfactions.

Abstract: Background: While promising, cationic lipid-mediated gene delivery can still benefit from improvements in lipid design and lipid-DNA (lipoplex) formulation. The putative mechanism of cellular lipoplex uptake is believed to occur by endocytosis, where the key influential factors are lipoplex size and morphology; lamellar and inverted hexagonal. Lamellar lipoplexes offer superior protection to the DNA cargo, while the inverted hexagonal phase best facilitates endosomal escape. Ideally, the initial lipoplex packaging would have the lamellar phase upon uptake, followed by a phase transition to hexagonal, facilitating cargo release into the cytosol. The cationic lipid structure defines its molecular packing parameter, S, which in turn controls the lipid phase transition. A molar weighted average packing parameter (Smix) for the overall cationic and neutral co-lipid mixture within a lipoplex formulation is predictive of a lamellar (S&lt;1) or hexagonal (S&gt;1) phase lipoplex. Objectives: Pyridinium-based cationic lipids represent a class of non-viral vectors that have shown promise in gene delivery. The objective of this study was to test the influence of lipid shape on lipoplex phase structure through small-angle x-ray diffraction (SAXD), and correlate shape with transfection efficiency. Lipoplexes co-formulated with pyridinium lipid, (16:0)(11:1) having a calculated shape parameter, S, of 1.08 and the commercial cationic vector, EPC (S=0.94) were predicted to undergo a packing transition from lamellar to hexagonal as the ratio of (16:0)(11:1) / EPC is increased. A fixed amount of neutral co-lipid was employed (1,2-dioleoyl-sn-glycero-3-phosphoethanolamine, DOPE: S=1.01; or cholesterol, Chol: S=1.20). Given that cholesterol is a higher-S lipid than DOPE, the lipoplex phase transition from lamellar to hexagonal was anticipated to occur at a lower ratio of (16:0)(11:1) to EPC. Methods: Liposomes were prepared from pyridinium-based cationic lipids in combination with EPC and co-lipid, DOPE or cholesterol. Lipoplexes were then formulated by incubating the liposomes with plasmid DNA at various N/P (+/-) molar charge ratios, and subsequently characterized by gel retardation, DNAse I degradation, biocompatibility and β-galactosidase (β-gal) transfection assays using Chinese Hamster Ovarian (CHO-K1) cells. Lastly, lipoplexes at N/P molar charge ratio 3 (only) were analyzed by SAXS at the synchrotron in Grenoble, France. Results: The SAXD results revealed that the (16:0)(11:1)/EPC/DOPE-DNA lipoplex formulations underwent a lamellar to hexagonal packing transition when the (16:0)(11:1)/EPC molar ratio was increased from 1:2 to 1:1, where the Smix increased from 1.01 to 1.04. However, (16:0)(11:1)/EPC/Chol-DNA lipoplex formulations underwent a lamellar to hexagonal transition when the (16:0)(11:1)/EPC molar ratio increased from 1:1 to 2:1; when Smix increased from 1.11 to 1.13. For both DOPE and Chol containing lipoplexes, the greatest transfection was found at (16:0)(11:1) to EPC ratios below 2:1, at an N/P molar charge ratio of 3. Conclusion: The lamellar to hexagonal packing transition, as determined by SAXD, for the lipid-DNA lipoplexes composed of the (16:0)(11:1)/EPC mixture occurred as predicted by our Smix calculations when DOPE was employed as co-lipid. The same was not observed when cholesterol was employed co-lipid. Finally, superior lipoplex transfection correlated with lamellar packing.

Introduction: Mild Brain Injury or concussion is frequently observed during contact sports such as football and soccer. If it remains undetected, a repeat concussion can lead to long term consequences because of the vulnerable brain tissue damage. Balance Error Scoring System (BESS) test is a widely used test to detect concussion. It requires the athlete with suspected concussion to maintain his/her position in three different stances, that are, both legs, single leg and tandem, and has a total score ranging from 0 to 30. The recommended way of performing this test is to do it barefoot in clinical or semi-clinical settings. Such conditions are however difficult to achieve during an ongoing match, and athletes would like to be near the field with their cleats on - situations often found on soccer fields in eastern countries. Objective: Therefore, we aimed to assess the reliability of BESS test in different field conditions. This research is in line with Qatar National Research Strategy 2012 pillars, H.E.1.9 (prevention of brain Injury) and H.E 1.10 (control of sports injuries). Methods: This study was conducted under the auspices of McGill Sports Emergency Medicine Clinic. Athletes from soccer and football teams were approached on the field during practice games. After informed consents, they performed BESS test in three conditions, that were, barefoot, on turf with cleats and on hard surface with cleats. Each athlete was rated by three observers independently of each other. We computed mean difference in total BESS scores with 95% confidence intervals (95%CI). Comparison of total BESS scores under different conditions as well as inter-observer reliability was assessed using intraclass correlation coefficient (ICC).. Results: We recruited 49 athletes from football (n=39) and soccer (n=10) teams in this study. Thirty nine of them were male, 10 were females. Average age was 21.1 years (standard deviation [SD]=1.9). We found that total BESS scores were significantly different (P&lt;0.001) between barefoot and the two conditions with cleats-on: 2.2 (95%CI=1.6, 2.8) for turf and 2.0 (95%CI=1.4, 2.6) for hard surface. Concordances of barefoot with turf (ICC=0.47, P=0.02) and hard surface (ICC=0.51, P=0.01) conditions were moderate. A moderate to high inter-observer reliability (0.60≥ICC≤0.75) was observed for BESS test under three conditions. Conclusion: These findings show that BESS test has a fair reliability under different conditions, and may be useful in screening concussion on the field. However, cut-off of BESS should be reduced by 1 to 2 points if it is applied on the field with cleats.

Diabetic nephropathy (DN), a serious complication of diabetes, is characterized by hyperfiltration, hypertrophy, extracellular matrix accumulation, fibrosis and proteinuria leading to loss of renal function. In renal hypertrophy, tubules increase in size and accumulate extracellular matrix and are also associated with alterations in renal sodium handling as well as hypertension; processes linked by involvement of the arachidonic acid (AA) metabolites 20-HETE and EETs. This study aims to determine the specific AA-metabolizing CYP450 isoforms present in proximal tubules (PT) that are altered by high glucose (HG) in cultured PTs, and in an animal model of diabetes. It intends to investigate the effects of alterations in CYP isoforms and/or AA-metabolite levels in DN. This work will investigate the mechanism of PTs injury and the effect of inhibition of AA-metabolites in vitro and will also get insight onto the cross-talk between CYP450 isoforms and other sources of Reactive Oxygen Species (ROS). Immunohistochemistry, hypertrophy, apoptosis, fibrosis, ROS generation, 20-HETE and EET formation, CYP4A and Nox protein expression, and mRNA levels were measured in vitro and in vivo. In our study, we show that exposure of rats proximal tubular epithelial cells to high glucose (HG) resulted in increased extracellular matrix accumulation and hypertrophy. HG treatment increased ROS production and was associated with alteration in CYPs 4A and 2C11 expression concomitant with alteration in 20-HETE and EETs formation. HG-induced tubular injury were blocked by HET0016, an inhibitor of CYPs 4A. In contrast, inhibition of EETs promoted the effects of HG on cultured proximal tubular cells. Our results also show that alteration in CYPs 4A and 2C expression and 20HETE and EETs formation regulates the activation of the mTOR/p70S6Kinase pathway, known to play a major role in the development of DN. To assess the significance of our in vitro findings, in vivo experiments were performed. Type 1 diabetic rats were used to assess the levels of different cytochromes as well as the levels of injury in these rats. In this study, we demonstrate that rats with streptozotocin-induced diabetes develop renal hypertrophy and increased fibronectin expression concomitant with an increase in CYP4A expression and a decrease in CYP2C expression. These observations were paralled by an alteration in 20-HETE and EETs productions. These results were also paralleled by an increase in reactive oxygen species (ROS) production and NADPH oxidase activity. Treatment of diabetic rats with HET0016, selective inhibitor of CYP 4A, prevented all these changes. In contrast, treatment of diabetic rats with MsPPOH, a potent inhibitor of EETs formation worsens the injury seen in the kidneys of the diabetic rats. Our results indicate that hyperglycemia in diabetes has a significant effect on the expression of AA-metabolizing CYPs, manifested by increased AA metabolism, and might thus alter kidney function through alteration of type and amount of AA metabolites; this pathway is through an oxidative stress-dependant mechanism.

Background and Aim: The prevalence of type-2 diabetes (T2D) has doubled in the last three decades and is still rising at an alarming rate, thereby posing a major challenge to global health. MicroRNAs (miRNAs) are a class of short RNAs, which play an important role in regulating physiological processes in diabetes. These small RNAs post-transcriptionally suppress mRNA target expression and, therefore, modulation of specific miRNAs may prove to be a promising strategy to treat diabetes. However, the potential roles of the different microRNAs in the eitiology of diabetes and diabetes-related complications are not yet completely understood. In the present study, we aimed to explore the role of miR221/ 222 in diabetes. Materials and Methods: Mouse microvascular endothelial cells (MMECs) were cultured for 48 hours under conditions designed to mimic the mileu of type 2 diabetic mice: normal glucose (NG=11mM) and high glucose (HG=40mM). The levels of miRNA221/222 expression were then analysed by real-time PCR. MMECs were transfected with miR221/222 inhibitors and mir221/222 mimics and then cells were exposed to normal/ high glucose in the presence or absence of metformin. Expression of miRNA 221/222 were again analysed and compared. The effects of miR 221/222 expression on eNOS, phospho-eNOS and eNOS monomer/dimer protein levels were determined through western blotting. Results: We found that exposure to high levels of glucose, which mimics hyperglycaemia conditions, induced expression of miR-221 and miR-222. Treatment of metformin partially restored this HG-induced high expression of miRNA. Moreover, metformin showed an additive effect with miR221/222 inhibitors to inhibit miR 221/222 expression in hyperglycaemic condition. Furthermore, we observed that protein levels of total eNOS and phospho-eNOS decreases in HG in comparison with NG. The eNOS/P-eNOS protein levels were restored upon inhibition of miR221/222, thus indicating correlation of these micro-RNAs with eNOS signalling. Results were inversely validated by using miR mimics (overexpression). Conclusion: These findings suggest that miR221/222 may offer a new therapeutic strategy for treatment of endothelial dysfunction in diabetic patients or may work as a therapeutic modulator. The project is supported by UREP 13-116-3-024

Background: Expanding adipose tissue in obesity requires effective angiogenesis and vasoreactivity to combat hypoxia and its consequences, such as insulin resistance and type-2 diabetes. While recent evidence suggests that the adipose tissue is highly angiogenic and inflammed, the tissue arteriolar vasoreactivity has been less investigated. As a consequence of the inflammation the omental adipose tissue (OAT) also synthesizes greater levels of vasoconstrictive molecules, such as cytokines and catecholamines, compared to the sub-cutaneous (SAT) depot, and these are likely to impact on the maintenance of vascular tone leading to greater susceptibility to hypoxia. This study compared the contractile responses of OAT and SAT arterioles from insulin-resistant (IR) and insulin-sensitive (IS) morbidly obese non-diabetic Qatari patients. Methods: Segments of arterioles (ID ~240-250 µm) isolated from SAT and OAT obtained from obese non-diabetic Qatari patients (age ~29 years, BMI ~40kg.m-2), undergoing bariatric weight reduction surgery, were mounted on a dual wire myograph (510A) and investigated for noradrenergic responsiveness. Cumulative concentration-response curves were constructed for noradrenaline (10-9 -10-5 M). Curves were also constructed for potassium chloride (KCl, 1-70 mM). Results: OM arterioles from IR patients were significantly less sensitive to NA compared with SAT arterioles from same patients (log EC50 -5.9±0.2 vs. -6.5±0.1, p<0.05). Maximum NA contractile response was also attenuated in OAT compared with SAT vessels (p<0.05) from these patients. On the other hand, KCl curves were comparable for OAT and SAT vessels from the same patients. In addition, no differences in noradrenergic sensitivity were observed between OAT and SAT vessels from IS patients. Conclusions: The data suggest that insulin resistance selectively alters noradrenergic responsiveness of OAT arterioles compared with SAT vessels from morbidly obese non-diabetic Qataris. Differences in adrenoceptor density/function may underlie these depot-specific responses. Studies on the disease specific differences need further investigation.

Purpose The first stage of this QNRF funded study includes a series of key informants' interviews with stakeholders in the PHC sector in Lebanon and Qatar to formulate an understanding of the recruitment and retention strategies of health human resources (HHR) and understand the obstacles and challenges that they face. This abstract reports on the findings obtained in Lebanon, as data collection remains ongoing in Qatar. Participants & Methods Study utilized a qualitative design involving semi-structured key informant interviews with stakeholders in the PHC sector. An initial list of key stakeholders was acquired from a review of public and private information sources while ensuring maximum variability across institutions, disciplines and geographical locations. Overall, 22 key informants participated in the study. They included decision and policy makers in the PHC sector, managers/directors of PHCCs, human resources coordinators in PHCCs, physicians and nurses, and academicians. Analysis Thematic and content analysis, using Nvivo 8, was conducted on the data collected from the key informant interviews. After coding the responses into similar concepts, axial coding allowed for the emergence of five comprehensive themes: perception of PHC, PHC services, recruitment of HHR in PHC, retention of HHR in PHC, and recruitment and retention in rural areas. Results Responses of stakeholders with regards to the definition of PHC revealed a lack of a unified understanding of the concept. With regards to services offered at PHCCs, there was a consensus that there is a deficiency in various services, most notably was mental health, as well as various preventive functions such as vaccination, women's health, and health behavior modification. Thematic analysis identified a number of impediments to the recruitment of HHR, mainly related to shortage in the overall supply of HHR, of qualified HHR, and HHR gender imbalances. Despite recruitment strategies in place, factors including financial constraints and poor leadership/management hinder the effectiveness of recruitment efforts. Although stakeholders report an acceptable retention of HHR, they relate turnover to poor working environment and lack of professional development. There was consensus that challenges faced are more pronounced in PHCCs of rural areas. Conclusions The study findings reveal that the current status of recruitment and retention within the PHC sector is not conducive to a solid and stable workforce. There is an evident need for the establishment of a unified contextualized definition of PHC to be applied across all PHCCs operating nationally. Moreover, the adoption of a system's thinking approach is crucial for PHC capacity building, in which the existent structure is better supported by qualified personnel. Extensive efforts need to be exerted towards directing health care professionals to the PHC field especially in rural areas, while concurrently enhancing the working conditions within PHCCs. Accordingly, essential services may be more adequately provided to the community. Of particular importance is the integration of mental health services into community care. Decision and policy makers are urged to reflect upon the recommendations developed in order to not only stabilize the PHC workforce but to also ensure the longevity of its services.

Sickle Cell Anaemia is an, autosomal, genetically-inherited, blood disorder, arising due to a point mutation in the bases coding for the sixth amino-acid of the β-chain of haemoglobin. The effects of the mutation begin to play role at the event of translation; during which the mutated haemoglobin (HbS) is synthesised. Here a hydrophobic protein residue (valine) is incorporated at the position of a hydrophilic residue (glutamic acid) in the growing protein chain. However, its orientation imitates that of the hydrophilic residue as otherwise seen in the wild type haemoglobin (HbA). Due to this change the hydrophobic side-chain of the amino-acid (valine) is prevented from being buried within the hydrophobic core of the protein and remains exposed to the surface. The resulting HbS thus displays a hydrophobic and unstable character with a tendency to polymerise with other HbS molecules causing the Red Blood Cells (RBC) to take a characteristic sickle shape. Through this research initiative, an attempt was made to unfold the protein to an extent that was sufficient to expose its hydrophobic core, thereby allowing it to engulf the side-chain through the formation of hydrophobic interactions. Thus the resultant modified protein would display an overall stable character, mimicking the wild-type HbA in spite of its mutational status. As a preliminary analysis, partial unfolding and refolding experiments were carried out on HbA molecules to determine whether the quaternary structure of haemoglobin would be retained. These experiments were monitored through Circular Dichroism (CD) Spectrophotometry. Partial unfolding was targeted by treating the protein with different concentrations of a mild denaturant (dimethyl sulfoxide). Unfolding was arrested at different stages of time (12, 24, 36, 48, 60, 72 hours) by the introduction of an organic solvent (chloroform) that induces precipitation of the protein. The modified protein was then tested for changes in hydrophobic character and stability by Reverse Phase Chromatography using C18 columns. Tests for solubility and aggregation were also performed spectrophotometrically. All tests were carried out under both oxygenated and deoxygenated conditions. It was observed that modified haemoglobin molecules arrested at 36 and 48 hours displayed a change in structural conformation. However, CD spectrophotometric analysis confirmed that they did not refold to resemble the wild-type HbA . Chromatographic results showed that the modified protein developed an overall neutral character, while spectrophotometric analysis proved that the molecules were insoluble in potassium phosphate buffer (pH 7) under both oxygenated and deoxygenated conditions. The study proved that partial unfolding up to 36 to 48 hours was sufficient to expose the hydrophobic core to trigger interactions that causes the haemoglobin molecule to attain an overall neutral and stable character. However, further analysis is ongoing to control the refolding of the protein to more closely match its native state. The study is also experimenting with other models of denaturants as well as more refined methods to arrest unfolding in order to improve solubility of the protein.

Autoinflammatory disorders are a group of Mendelian disorders characterized by seemingly unprovoked inflammatory bouts without high-titer autoantibodies or antigen-specific T-cells and are probably due to defects in the innate immunity. We here report on a 4-year old Arabic child with the clinical presentation of an autoinflammatory disorder, namely Pyogenic Arthritis, Pyoderma Gangrenosum and Acne (PAPA) syndrome. The presentation includes abscess formation after immunization and recurrent mono-articular acute arthritis in various joints that responded favorably to systemic glucocorticosteroids, albeit without acne or pyoderma gangrenosum. The mutation analysis of the child identified a novel de novo mutation in PSTPIP1, the gene responsible for PAPA syndrome. We recommend that the diagnosis of PAPA syndrome should be entertained in the differential diagnosis of patients with recurrent sterile pyogenic arthritis prior to the development of pyoderma gangrenosum or acne in order to initiate a timely management of the disorder.

The Yamanaka factors (Oct4, sox2, Klf4, cMyc) used to produce induced pluripotent stem cells (iPSC) have become a staple of stem cell biology and ChIP-seq studies have led to an understanding of the genomic landscape of how these as well as other recently discovered factors transduce their effects to form iPSC. A conundrum that previously existed was that the Sox family has high homology and yet various members drive completely different cell fates. My lab in collaboration with Larry Stanton&#39;s lab at the Genome Institute of Singapore unraveled part of the mystery by identifying and validating a single residue within the hmg domain of the Sox family responsible for a specific interaction with Oct4 (pou5F1 domain) which then decides specific cell fates. In addition a novel genomic motif was discovered which has a single base difference between the Oct4 and Sox binding sites. Using this knowledge we were able to transform the function of pluripotent Sox2 into an endodermal TF and endodermal Sox17 into a pluripotent TF. In fact the mutated Sox17 TF was even more efficient in forming iPSC (compared to Sox2) which was not expected. Recently we investigated the C-terminal activation domains of these Sox family members and discovered that they play a major role in the level of activation of iPSC. Conversion of Sox17 into a Pluripotency Reprogramming Factor by re-engineering its Association with Oct4 on DNA. Ralf Jauch, Irene Aksoy, Andrew Paul Hutchins, Calista Keow Leng Ng, Xian Feng Tian, Jiaxuan Chen, Paaventhan Palasingam, Paul Robson, Lawrence W. Stanton and Prasanna R Kolatkar. Stem Cells. 2011. Jun;29(6):940-51. Oct4 switches partnering from Sox2 to Sox17 to reinterpret the enhancer code and specify endoderm. Aksoy I, Jauch R, Chen J, Dyla M, Divakar U, Bogu GK, Teo R, Leng Ng CK, Herath W, Lili S, Hutchins AP, Robson P, Kolatkar PR, Stanton LW. EMBO J. 2013. 32(7): 938-53. Sox transcription factors require selective interactions with Oct4 and specific transactivation functions to mediate reprogramming. Aksoy, I., Jauch, R., Eras, V., Bin, A.C-W., Chen, J., Divakar, U., Ng, C. K-L., Kolatkar, P.R., Stanton, L.W. Stem Cells. 2013.

Background. Chronic myelogenous leukemia (CML) is the most common myeloproliferative disease accounting for ~15% to 20% of all cases of leukemia (1-1.5/100.000 cases per year). It originates from a pluripotent bone marrow stem cell in which a t(9;22) results in the production of BCR/ABL fusion protein which has a constitutive tyrosine kinase activity and deregulates signal transduction pathways. Phosphorylation of key residues is required for the full transforming activity of BCR/ABL; for this reason much attention has been focused on the role of phosphatases, natural regulators of the tyrosine kinase signaling. We have previously reported that protein tyrosine phosphatase receptor type ? (PTPRG) is a tumor suppressor gene which interacts with BCR/ABL, inhibits downstream signaling events and is downregulated in CML. Whitin the QNRF research project NPRP 4-157-3-052 we analyzed the expression levels of PTPRG gene in 32 CML patients at diagnosis and following TKI treatment aiming at the evaluation of the clinical impact of PTPRG dowregulation in CML. Methods: Thirtytwo patients diagnosed with CML in chronic phase and 13 untreated patients diagnosed with philadelphia-negative myelod disorders as control group were included in the study. The study was approved by the Local Ethics Committee, and informed consent in accordance with declaration of Helsinki was obtained from each patient. The expression level of the PTPRG gene was evaluated by a sybr-green absolute quantification RT-PCR assay in 2 samples from each patient, taken at diagnosis and following TKI treatment using the beta-Actin (ACTB) housekeeping gene for normalization. Results were expressed as PTPRG/ACTB ratio and were validated using predesigned TaqMan quantitative RT-PCR assays for PTPRG and ABL1 genes. BCR-ABL1 transcript was quantified by realtime RT-PCR according to the European Leukemia Net guidelines. Statistical analysis and comparisons were performed using the SPSS-software. Results: PTPRG transcript was undetectable in 11/32 (34,3%) CML samples at diagnosis and the median levels of PTPRG mRNA were significantly lower in CML samples at diagnosis compared to the non-CML control group (0,44%, range 0-0,37 vs 6,29%, range 0,09-52; p=0.02). On the contrary, PTPRG mRNA was detected at variable levels, ranging from 0.17 to 30%, in 29/32 follow up samples, taken at different time points of treatment. Differences in PTPRG gene expression levels between CML samples before and after treatment were statistically significant (p=0,027). Quantitative RT-PCR for PTPRG has been also set up at the Hematology center, NCCCR, Doha, Qatar. Preliminary results showed that mean levels of PTPRG mRNA were comparable to the italian group of patients. No statistically significant correlation was observed between PTPRG levels and clinical/biological factors. Interestingly, 2 of the 3 patients showing higher level of PTPRG mRNA at diagnosis than in the follow up sample showed resistance to TKI treatment. Conclusions: We found a down regulation of PTPRG in a high percentage of CML patients and a recovery of its expression upon treatment with TKIs. Deregulated expression of PTPRG phosphatase underline its role as a tumor suppressor gene in CML and highlights its potential use as a new bio-marker of disease potentially usable in association with BCR/ABL1.

Background &amp; objectives: Vascular complications account for much of the morbidity of obesity. The proportion of Qatari population that is overweight or obese is one of the highest in the world. With this comes the increased risk of blood vessel dysfunction and predisposition to type 2 diabetes and hypertension. The vascular endothelium is often the first target of the negative impact of obesity. It is however unclear how the heterogeneity in the metabolic status of obese individuals (ie metabolically healthy [MHO] vs. Pathologically obese [PO]) will impact on endothelial function, particularly in a relatively young obese population, as in Qatar. This study investigated endothelium-dependent relaxation of small arteries embedded in the visceral (omental) and subcutaneous adipose tissues in morbid obesity with varying metabolic status. Methods: Arteries were isolated from abdominal omental (OM) and subcutaneous (SC) fat collected from consented Qatari patients undergoing bariatric surgery for weight reduction. The arteries (normalized luminal diameter ~250 &#181;m for SC and ~ 240 &#181;m for OM) were cut into segments (~2 mm) and mounted on a dual wire Myograph (510A) for measurement of isometric tension. Cumulative concentration-response curves were constructed for acetylcholine (1- 30000 nM, the classical endothelium-dependent relaxant) in the absence or presence of Nω-Nitro-L-arginine methyl ester (L-NAME,100 &#181;M, nitric oxide [NO] synthase inhibitor) on initial tone generated with noradrenaline (5 &#181;M). Relaxation to sodium nitroprusside (SNP, an NO donor) was also recorded. Results: There were no differences in age (~32 years), blood glucose (~5.6 mmol/L) and body mass index (BMI , ~ 43.4 Kg.m-2) between the MHO and PO patients . Insulin levels were 3 vs 19 &#181;U/ml for MHO vs PO patients and their indices of insulin resistance (HOMA) were 1 vs 5 respectively. In general, relaxation to Ach was significantly attenuated in OM vessels (Emax 44&#177;8 %) compared with SC vessels (Emax 78&#177;4 %, p&lt;0.01) from same patients. In contrast, relaxation to SNP was greater in the OM compared with the SC vessels. When Ach relaxation of the OM vessels were separated according to the patients&#39; metabolic status, the MHO patients had significantly improved result compared with PO patients. On the other hand, relaxation of SC vessels from both groups of patients were comparable. In both vessel types, L-NAME caused a right-ward shift in the Ach curves. Conclusions: These results suggest that the metabolic status of obese Qatari patients has bearing on the physiology of their microvascular endothelium. The data also demonstrate that early changes in endothelial vasomotor function are depot-specific, being more marked in OM compared with SC vessels of pathologically obese patients.

Abstract The use of herbal and nutrition supplements is widespread. It has been reported that about 80% of the world's population use herbal medicines [1, 2, 3, 4]. Very few supplement use studies have been conducted in the Gulf Cooperation Council (GCC) and neighboring nations (5,6). Supplement use data among young adults is scarce. In one study aimed at determining the use of supplements among athletes in Qatar (5), over 60% of the study participants reported using vitamin supplements. Data regarding attitudes on the use of supplements among young adults is almost non-existent. The data on their perceptions about alternative medicine practitioners is also inadequate. We thus conducted a survey in which we examined the a) prevalence of supplement use among college students in Qatar, and b) their perceptions about the use of supplements and alternative medicine practitioners. We have previously presented our study findings regarding the use of supplements (7). At this meeting we will present our study findings concerning the perceptions of college students about the a) effectiveness and safety of supplements and b) alternative medicine practitioners. References 1. Eisenberg, DM, Davis, RB, Ettner, SL et al. Trends in alternative medicine use in the United States, 1990-1997: results of a follow-up national survey. JAMA 280: 1569-1575, 1998. 2. Farnsworth, NR, Akerele, O, Bingel, AS., Socjata, DD, Eno, Z. Medicinal plants in therapy. Bull World Health Organization 1985: 63: 965-981. 3. Gesler, WM. Therapeutic landscape medical issues in light of the new cultural geography. Soc.Sci. Med. 1992; 34: 735-746. 4. Rafferty, AP, McGee, HB, Miller, CE, Reyes, M. Prevalence of complementary and alternative medicine use: state-specific estimates from the 2001 Behavioural Risk Factor Surveillance System. Am. J. Public Health 92: 1598-1600, 2002. 5. Knez WL and Peake JM, The prevalence of vitamin supplementation in ultraendurance triathletes. Int J Sport Nutr Exerc Metab. 2010 Dec;20(6):507-14. 6. Abu-Irmaileh, BE, Affi, F.U. Herbal medicine in Jordan with special emphasis on commonly used herbs. J. Ethnopharmacology 89: 193-197, 2003. 7. Mamtani R, MacRae B, Mahfoud Z, Cheema S , El Hajj M, Lopez T and Lowenfels A. Use of herbal and nutrition supplements among college students in Qatar. American Society of Clinical Nutrition, 2013, Dubai. Acknowledgement: This work is supported by the Biomedical Research Program at Weill Cornell Medical College in Qatar, a program funded by Qatar Foundation.

Introduction: Diabetes mellitus (DM) is a disease with severe complications and major health/economic impacts. It is a leading cause of morbidity and mortality worldwide, with an estimated 346 million adults being affected in year 2011. World Health Organization (WHO) projects indicated that diabetes death will increase by two thirds between 2008 and 2030. The WHO estimated that 80% of the populations of developing countries rely on traditional medicines, mostly plant drugs, for their primary health care needs. Diabetes is an example of a disease that has been treated with plant medicines. Our study evaluated ethanolic and aqueous extracts of selected Sudanese plants that are traditionally used to treat diabetes; these are: Ambrosia maritima, Ammi visnaga, Acacia senegal, Sesamum indicum, Nigella sativa and Foeniculum vulgare. The plants extracts were tested for their glycogen phosphorylase inhibition, toxicity and antioxidant activity. Materials & Methods: Ethanolic and aqueous extracts prepared from leaves of Ambrosia maritime, fruits of Foeniculum vulgare and Ammi visnaga, exudates of Acacia Senegal, and seeds of Sesamum indicum and Nigella sativa were investigated. The antioxidant properties of the extracts were tested using (DPPH) photometric and Iron Chelating Assays. The enzymatic inhibition of glycogen phosphorylase (GP) activity was monitored using multiskan spectrum (Thermo-Scientific). GP activity was measured in the direction of glycogen synthesis by the release of phosphate from glucose-1-phosphate. Brine Shrimp Lethality Test was also used to determine plants toxicity. Results and Discussion: Free radicals are formed in diabetes by glucose oxidation, nonenzymatic glycation of proteins and subsequent oxidative degradation of glycated proteins. Abnormally high levels of free radicals can lead to damage of cellular organelles and enzymes, increased lipid peroxidation and development of insulin resistance. These consequences of oxidative stress can promote the development of complications of diabetes mellitus. In this study all plant extracts with exception of Acacia senegal exhibited significant antioxidant activity in DPPH free radical scavenging assay. This may support the traditional usage of these plants to improve complications that caused by diabetes mellitus. Nigella sativa aqueous extract showed no toxicity on Brine shrimp Lethality Test, while its ethanolic extract was toxic. All other extracts are toxic and ethanolic extracts of Foeniculum vulgare and Ammi visnaga exhibit the highest toxicity. Results of this study did not show any significant inhibition of glycogen phosphorylase, but extracts of these plants may act on one of other enzymatic reactions that involved in carbohydrate metabolism and improved glucose homeostasis. Conclusion and Recommondation: Changes in oxidative stress and effects of antioxidants in diabetes management should be considered, and hopefully, further research into the pathophysiology of oxidative stress and the role of antioxidant therapy will lead to appropriately-designed clinical trials in which the promise of antioxidant therapy will be realized. Extraction processes and usage doses should be monitored. Further work is underway to test the plant extracts on diabetes-induced mice. Key words: Diabetes mellitus, medicinal plants, antioxidant activity, glycogen phosphorylase, brine shrimp

Rationale Despite improvements in oral health care during the last decade, caries and other tooth structure defects still represent major global public health concerns. To date, no index has satisfied records of different tooth structure defects that could be present in the same mouth and in the same tooth. Available indices provide data on some defects separately. Exposing decision makers to deficient information leaves them unaware of the high levels of untreated defects. Objective Our long term objective is to develop a method for valid and reliable evaluation of tooth structure defects. Such a method would have a broader utility in the implementation of cost effective preventive, non-operative and therapeutic measures to sustain oral health. Specific aims of the current study was to describe: 1) Different tooth structure defects in a random sample for the target population ( Qataris and Egyptians); 2) determine the most prevalent defect(s) in this population and 3)identify the most frequently affected teeth subject to these defects Materials and methods We carried out a cross sectional study to address the stated objectives. A random sample was recruited from 2 subpopulations ( Qatar and Egypt). Each study unit received thorough clinical assessment and each tooth was scored accordingly (table 1 shows clinical evaluation criteria). Data was entered in the data base and statistical methods were used. The association between condition and nationality was performed using the Chi square test/ Fischer's exact test. Results All individuals(93) showed evidence of tooth structure defect(s). Table 2 shows the frequency and percent distribution according to teeth structure defects (fig a, b). Apparently, decay is the most common defect in this target population (87%) followed by filled teeth (57%). The least common defects are abrasion and abfraction. A remarkable finding was that hypoplasia and hypo calcification are common in this population. These 2 conditions render teeth susceptible to caries. Among Qataris, the most common defect was decay followed by missed teeth(wisdom probably due to age) which is a similar pattern observed in Egyptians . Qataris are less likely to experience fractured teeth in comparison to Egyptians (odds ratio = 0.06 , p<0.001). Qataris are four times more likely to experience hypoplasia in comparison to Egyptians ( odds ratio 3.7 and the p value is 0.07). However, Qataris are 1.4 times more likely to experience hypo calcification compared to Egyptians but the odds ratio is not significant (p value 0.5). In this population it was common to see that teeth have more than one tooth structure defect which was not captured in the previous indices. Molars and premolars were most susceptible to decay whereas anterior teeth showed hypo calcification in Qataris whereas for Egyptians, molars and premolars were mostly decayed (tables 3 and 4) Conclusions To our knowledge, this is the first attempt to demonstrate the need to develop a thorough evaluation method that captures all possible tooth structure defects likely to occur in the same mouth and within the same tooth. The results urges the development of a valid and reliable index.

Background and objectives: A high prevalence of insulin resistance (IR) amongst normal weight Qatari individuals may contribute to progression of type II diabetes and account for the current epidemic. Factors contributing to this increased risk of IR and its associated co-morbidities, especially in the absence of obesity, are still under investigation. Therefore the objectives of this pilot study were to characterize components of IR in normal weight Qatari individuals and to investigate the potential molecular mechanisms underlying this phenotype. Methods: Non-diabetic lean/overweight Qatari subjects were recruited and anthropometric measures including body weight (kg), height (m) and blood pressure recorded, along with determination of systemic lipids, glucose, insulin and adipokines. Subjects were dichotomized into IR and IS groups based on their HOMA index (fasting plasma glucose < 6.8 mmol/l and insulin levels < 6.5 miU/ml). The expression of 84 most abundantly expressed and characterized miRNA species was profiled in peripheral blood samples. Target genes of miRNA let7 were determined using Human let-7a Targets PCR Array. Results: When subjects (29+/-6.8 years old, BMI of 23+/-4.9 kg/m-2) were stratified into two groups based on their HOMA index (IS 1.3 (1-1.6) and IR 2.2 (1.6-2.7), IR (66%) was only associated with higher insulin levels {(IS 5.3 (5.2-5.5) vs IR 9.5 (7.7-12.1) u/ml, p <0.01)} with no significant differences in blood pressure, lipids profile or adipokines levels. Among 84 profiled miRNA species, the expression of 7 miRNA varied significantly between the two groups; among these were four members of let7 family (g/b/c/f). Three potential target genes of let7 exhibited significant variable expression between the two groups, including dual specificity protein phosphatase 1 (DUSP1). Conclusions: Prevalence of IR among young, lean/overweight Qatari individuals is alarmingly high (66%). This study has revealed let7 miRNA as a potential target mediating this phenotype. This miRNA has been shown previously to play an essential role in adipogenesis. The targets of let7 with an ability to regulate adipogenesis is currently being investigated to confer functionality.

Hematological Malignancies (e.g., leukemia and lymphoma) are among the top 5 causes of cancer death in the Middle Eastern Countries, including Qatar, Kuwait and Saudi Arabia. The monoclonal antibody rituximab, directed at the CD20 antigen, has become an essential drug for the treatment of Non Hodgkin Lymphoma (NHL). Although transient B cells depletion frequently occurs after rituximab treatment, it usually resolves after 6-9 months. Nevertheless, high frequency of non-neutropenic infections and persistent hypogammaglobulinaemia during follow-up period have been recently reported. However, impaired humoral response to the recall and primary antigens was found in NHL patients during (or few months after), rituximab treatment. Influenza vaccination is generally recommended in lymphoma patients, but few data are available about the activity of this vaccine after rituximab-containing regimens (RCR). It is presently unclear whether patients treated with RCR regain normal immunocompetence after achievement of complete remission. We presently combined data of 3 sequential studies conduced at our Institutions assessing the humoral response to seasonal influenza vaccination (2008/2009, 2009/2010 and 2011/2012 seasons; RIT-01, RIT-02, and RIT-03 studies, respectively) in NHL patients in complete remission (CR) for at least 6 months after treatment with rituximab-containing regimens (RCR). Response was evaluated by hemagglutinin inhibition assay 3 or 4 weeks after vaccination. The following (inactivated) vaccine formulations were used: virosomal vaccine (RIT-01; N=31), MF-59 adjuvanted vaccine (RIT-02; N=14), and intradermal vaccine (RIT-03; N=22). Data were compared with those from age-matched cohorts of healthy volunteers (HV). In the RIT-03 study, cancer patients who received chemotherapy without rituximab (CWR) more than 6 months before study entry were also evaluated. In the RIT-02 study, patients also received two doses of pandemic H1N1 vaccine. To determine early transcriptional changes predictive of immunoresponsiveness and to determine differences in innate immunity activation among patients treated with RCR, HV, and patients treated with CWR, PBMC were collected just before and 1 day after vaccination (RIT-03 study). Whole-genome gene expression analysis of these samples using microarray analysis is currently ongoing. We found that the intradermal vaccination is associated with dramatic transcriptomic changes in PBMC, already detectable 24 hours after vaccination. These changes underlie modulation of innate response (eg, interferon stimulated genes). Changes after influenza vaccination differ among CWR, CRC, and healthy volunteers. Overall, we found that patients treated with RCR have a significant lack of humoral response to both recall and na&#239;ve influenza antigens as compared with HV and with cancer patients treated with CWR. This impairment persisted even long time (&gt; 6 years) after last rituximab administration and was associated with depletion of CD27+ memory B cells and hypogammaglobulinemia. Therefore, patients previously treated with RCR should be strictly monitored during influenza epidemic season. Intradermal vaccination seems to induce a stronger response as compared with the other two formulations.

Background: Traumatic brain injury (TBI) and related death rates vary worldwide. Objectives: To evaluate the incidence, causes and outcome of TBI in Qatar. Method: A retrospective review of all TBIs admitted to the trauma center between January 2008 and December 2011 was performed. Patients' demographics, mechanism of injury, morbidity, and mortality were analyzed. Results: A total of 1665 patients with TBI were admitted, the majority were males (92%) with a mean age of 28±16 years. The common mechanism of injury was motor vehicle crashes (MVCs)and falls from height (51% and 35%, respectively). TBI was incidentally higher in young adults (34%) and middle age group (21%). The most frequent injuries were brain contusion (40%) followed with subarachnoid (25%), subdural (24%), and epidural hemorrhage (18%). The mortality rate was 11% among TBI patients. Mortality rates were 8% and 12% among adolescents and young adults, respectively. The highest mortality rate was observed in elderly patients (35%). Head AIS, ISS and age were independent predictors for mortality. Conclusion: In Qatar,TBI is reported in around 27% of all the trauma admissions;mostly due to MVCs and is associated with high mortality. Elderly are the most vulnerable group. particularly in the older group. Public awareness and injury prevention campaigns should target young population.

Objectives: Dizziness is a relatively common medical complaint that that rarely requires hospitalization for work-up and management. Whether gender related differences exist in the etiologies and outcome of patients hospitalized with dizziness is unknown. The aim of this study was to compare women and men presenting with dizziness in a real-world population. Methods: Retrospective analysis of all patients hospitalized with dizziness in Qatar to the cardiology service from 1991 through 2010 was made. Patients were divided into two groups according to gender. Clinical characteristics, management and outcomes were analyzed. Results: During the 20-years period, 1578 patients were hospitalized with dizziness; 404 women (25.7%) and 1173 men (74.3%). Women had significantly more prevalence of hypertension (46.9% vs. 31.1%, P=0.001) and diabetes mellitus (39.8% vs. 31.1% P=0.001) compared to men. Cardiac arrhythmia was the most common underlying diagnosis and was significantly more common in women than men (40% vs. 28.4%; P= 0.001), whereas acute coronary syndromes were significantly less common in women (13.6 vs. 25.9%; P= 0.001). The in-hospital mortality rate was significantly higher in women with dizziness compared to men (5.7% vs. 3.2%; P=0.02) [table]. Conclusions: Our study demonstrates that women hospitalized with dizziness have worse in-hospital outcome and different underlying etiologies compared to men. Further prospective research is warranted to confirm our observations in other registries.

Failure to offer evidence based structured diabetes educational program impacts negatively on the effectiveness of diabetes management and leads to increased morbidity and mortality. We assessed the efficacy of a nurse led group based structured diabetes education in improving glycemic and metabolic parameters among Arab type 2 diabetic patients compared to usual care Methods This was a parallel group randomized trial in adult Arab type 2 diabetic patients living in Qatar. Subjects were randomized to nurse led structured diabetes educational program ,or to usual care .The primary outcome was the improvement in HbA1c and other metabolic parameters including lipid profile ,albumin/creatinine ratio ,blood pressure and body mass index.. Patients were invited to attend four 2-hour sessions of self-efficacy improvement education .Outcomes were assessed at base line and 12 months later. The primary analysis was an intention to treat. Results Participation of the intervention was shown after 12months to have led to a statistically significant improvements in HbA1c(-0.55 m mol /L ,p=0.012),F.B.S(-0.92 m mol/L ,p=0.022),B.M.I(-1.70,p=0.001) and albumin/creatinine ratio(-3.09,p<0.001) but not in the control arm. conclusion Inclusion in the Nurse led intervention by adult Arab type 2 diabetic subjects was shown 12-months post intervention to have led to enhanced glycemic and metabolic parameters including body mass index and blood pressure .The success of this trial justify the feasibility and generalization of this intervention to neighboring Arabian Gulf countries who have similar Arab population with identical cultural beliefs and practices.

Facilitated LMA fiberoptic intubation while keeping on ventilation in ischemic heart patient with unexpected difficult airway Authors: Mahmoud Abdalla MD. Hesham Ewila,MD. Hany Osman, MD. Abdulrashed Pattah MD. Institution: HMC, heart hospital, DOHA QATAR Difficult tracheal intubation remains an important cause of mortality and morbidity during general anesthesia, especially in ischemic cardiac patients where hypoxia rapidly compromise myocardial function and may induce dysrrhythmias. We report 68 years old male was admitted to Qatar heart center for elective CABG with poor left ventricular function EF 30 -35 % .No signs of difficult intubation were appreciated preoperatively. Intraoperatively LMA was inserted due to unexpected difficult airway after failing of 3 optimized trials of intubation. Endotracheal fiberoptic intubation through LMA also failed as patient rapidly desaturated. We attached T piece to the distal end of LMA, keeping patient ventilated through the side port of T piece, fiberoptic intubation achieved through pre cut plastic venous cap 0.5 inch which was attached to distal end of LMA. Endotracheal tube with ID 6.5 mm was inserted over the fiberscope then exchanged over a ventilating bougie to 8.5 mm tube. No significant changes in heart rate, blood pressure or oxygen saturation were appreciated throughout the procedure time. This maneuver allowed us to secure the airway while keeping on ventilation without compromising the poorly reserved cardiac function or exposing the patient to hypoxia or hypercarbia. Key Words: fiberoptic bronchoscope, unexpected difficult intubation, laryngeal mask airway. Ischemic heart disease

Household spices comprising, chili, Kashmiri chili hot, Kashmiri chili mild, basil, oregano, ginger, curry, cumin, turmeric, tandoori masala, garam masala, black pepper, garlic and coriander, were collected from local markets in Doha, Qatar, during 2012, and were surveyed for the presence of potentially harmful mycoflora and for contamination with aflatoxins B1, B2, G1, and G2 by high-performance liquid chromatography (HPLC). Among the tested spice samples, chili powder showed the highest presence of fungal propagules, while ginger, curry and garlic samples did not present any fungal contamination. A total of 120 isolates, mostly belonging to Aspergillus and Penicillium genera, were collected and 33 representative species were identified by amplification and sequencing of the internal transcribed spacer (ITS) region. Aspergillus flavus, Aspergillus nomius and Aspergillus niger were the most dominant. Thirty-seven Aspergillus strains were screened for their potential to produce aflatoxins using biochemical and molecular tools. Upon these methods, only 9 A. flavus strains showed both fluorescence and amplification with all the three primers targeting aflP, aflM and aflR genes. Aflatoxins were detected in five spices (black pepper, chili, tandoori masala. turmeric and garam masala), and with the exception of garam masala, the tested samples of turmeric, black pepper, tandoori masala and chili powder exceeded B1 and ⁄or total aflatoxin maximum levels. Our results demonstrate the potential for mycotoxin biosynthesis by fungi contaminating imported spice products.

Phospholipids in cells are mainly synthesized in the cytoplasmic leaflet of the endoplasmic reticulum (ER) and the newly made polar lipids must flip-flop rapidly across biological membranes to sustain cellular life. But this 'flipping' is energetically costly as well as its translocation rate is low. As a solution to this, cells have membrane proteins that function as lipid transporters - 'flippases', proteins that facilitates the rapid, bi-directional, energy-independent flip-flop of phospholipids between the cytosolic and lumenal leaflets of the ER membrane. Flippases have been known to play a key role in membrane stability as well as in the mechanism by which cells avoid being killed by macrophages. Candidate flippases have been implicated in human diseases that include intrahepatic cholestasis, angelman syndrome, autism, tangier disease, macular dystrophy and adrenoleukodystrophy. Establishing the primary function of candidate flippases and how they contribute to cell function and human disease is becoming a central issue in biology. Although the flippases that operate at the plasma membrane of eukaryotes at the expense of ATP hydrolysis resulting in unidirectional lipid flipping have been identified, the ATP-independent bi-directional flippases that translocate lipids in specialized compartments such as the ER have not yet been identified at their molecular level. The objective of the current study is to identify ER flippases in yeast Saccharomyces cerevisiae using a quantitative proteomics approach based on stable isotope labeling by amino acids in cell culture (SILAC). Yeast cells were grown in synthetic medium supplemented with either 'light' or 'heavy' lysine. Proteins extracted from unlabeled (light) cells were further fractionated by velocity sedimentation in a glycerol gradient and flippase activity of each fraction was quantified by a phospholipid reconstitution-based procedure. An aliquot of labeled (heavy) extract (containing equal amount of protein by weight) was added to each light fraction. The mixed fractions were then subjected to in-gel digestion followed by quantitative proteomic analysis using mass spectrometry. The data obtained were processed using MaxQuant followed by Spearman correlation analysis for identification of proteins with enrichment profiles matching that of the activity profile. The potential flippase candidates were tested for their activity using genomically tagged yeast strains.