beta1 integrin and collagen matrix interactions regulate the survival of cells by associating with focal adhesion kinase (FAK) and initiating MAPK/ERK signalling, but little is known about these signalling pathways during human fetal islet ontogeny. The purpose of this study was to investigate whether beta1 integrin/FAK activation of the MAPK/ERK pathway regulates human fetal islet cell expression of endocrine cell markers and survival. Isolated human (18-21 weeks fetal age) islet-epithelial cell clusters, cultured on collagen I, were examined using beta1 integrin blocking antibody, beta1 integrin siRNA and FAK expression vector. Perturbing beta1 integrin function in the human fetal islet-epithelial cell clusters resulted in a marked decrease in cell adhesion, in parallel with a reduction in the number of cells expressing PDX-1, insulin and glucagon (p < 0.05). beta1 integrin blockade disorganized focal adhesion contacts in the PDX-1(+) cells and decreased activation of FAK and ERK1/2 signalling in parallel with an increase in expression of cleaved caspases 9 and 3 (p < 0.01). Similar results were obtained following an siRNA knock-down of beta1 integrin expression. In contrast, over-expression of FAK not only increased phospho-ERK and the expression of PDX-1, insulin and glucagon (p < 0.05) but also abrogated the decreases in phospho-ERK and PDX-1 by beta1 integrin blockade. This study demonstrates that activation of the FAK/ERK signalling cascade by beta1 integrin is involved in the differentiation and survival of human fetal pancreatic islet cells.

Recent evidence has shown that stem cell factor (SCF) and its receptor, c-Kit, have an important role in pancreatic islet development by promoting islet cell differentiation and proliferation. In this study, we examined the role of c-Kit and SCF in the differentiation and proliferation of insulin-and glucagon-producing cells using a human pancreatic duct cell line (PANC-1). Our study showed that increased expression of endocrine cell markers (such as insulin and glucagon) and transcription factors (such as PDX-1 and PAX-6) coincided with a decrease in CK19 þ and c-Kit þ cells (Po0.001) during PANC-1 cell differentiation, determined by immunofluorescence and qRT-PCR. Cells cultured with exogenous SCF showed an increase in insulin þ (26%) and glucagon þ (35%) cell differentiation (Po0.01), an increase in cell proliferation (Po0.05) and a decrease in cell apoptosis (Po0.01). siRNA knockdown of c-Kit resulted in a decrease in endocrine cell differentiation with a reduction in PDX-1 and insulin mRNA, as well as the number of cells immunostaining for PDX-1 and insulin. Taken together, these results show that c-Kit/SCF interactions are involved in mediating islet-like cluster formation and islet-like cell differentiation in a human pancreatic duct cell line. Islet transplantation is a promising method for the cure of diabetes.1 However, the limited availability of cadaveric donors greatly hampers the application of this method. 2,3Therefore, multiple alternative methods are being explored to generate insulin-producing cells in vitro.4,5 A better understanding of the factors responsible for islet cell growth and differentiation is important to develop an unlimited supply of islets.c-Kit is a receptor tyrosine kinase that binds stem cell factor (SCF). It has been well established that c-Kit/SCF interaction is critical for the survival and development of stem cells involved in hematopoiesis, pigmentation and reproduction. 6 Recently, studies in the human and rat fetal pancreas have shown that c-Kit is expressed in islets and its ligand SCF is present in the pancreatic mesenchyme. 7-9Oberg-Welsh and Welsh 7 reported that c-Kit/SCF interaction promoted an increase in insulin content, indicating its role in the maturation of pancreatic b-cells. We recently examined the c-Kit expression pattern in the developing rat 10 and human fetal pancreas, 9,11 as well as in epithelial monolayers derived from postnatal rat islets. 12 Our studies showed pronounced changes in the distribution and population dynamics of c-Kit-expressing cells during islet development and differentiation, suggesting that c-Kit may have an important role.9,10 c-Kit þ epithelial monolayers derived from postnatal rat islets can give rise to new b-cells that secrete insulin in a glucose-responsive manner.12 Furthermore, a study of c-Kit WÀv mutant mice indicated that a lack of functional c-Kit receptors affected b-cell mass and disrupted b-cell maturation and function.13 Taken together, these reports suggest that c-Kit and its ligand, SCF, are important partners...

Background: Haemolysis is defined as the release of cellular components of erythrocytes and other blood cells into the extracellular space of blood. These cellular components can cause interference in laboratory measurements, potassium being a commonly measured analyte to be affected. A number of factors have been implicated in the aetiology of haemolysis. We undertook this study to enable us to identify and hence rectify causes of haemolysis in samples from patients on acute medical and surgical wards. Methods: We performed a prospective study of 353 blood sampling events during February and March 2007. A proforma was used to obtain detailed information of each blood-taking episode. Information from the proforma was linked to the incidence of haemolysis obtained from the hospital computer system. Results: The incidence of haemolysis among the samples studied was 6.5%. While staff group, method of sampling, tourniquet time and number of attempts at venepuncture were each univariately associated with haemolysis, stepwise logistic regression resulted in a final model which only included tourniquet time (odds ratio for haemolysis if tourniquet time .1 min was 19.5 [95% confidence interval [CI] 5.6-67.4%]). Conclusion: Tourniquet time of more than a minute is associated with a significant increase in risk of haemolysis. Advice on tourniquet time is included in phlebotomy training within the hospital; hence a campaign of appropriately channelled continuing education on this issue may be successful in reducing the haemolysis rate.

BackgroundThe aim of this pilot study was to identify proteins associated with advancement of colon cancer (CC).MethodsA quantitative proteomics approach was used to determine the global changes in the proteome of primary colon cancer from patients with non-cancer normal colon (NC), non-adenomatous colon polyp (NAP), non-metastatic tumor (CC NM) and metastatic tumor (CC M) tissues, to identify up- and down-regulated proteins. Total protein was extracted from each biopsy, trypsin-digested, iTRAQ-labeled and the resulting peptides separated using strong cation exchange (SCX) and reverse-phase (RP) chromatography on-line to electrospray ionization mass spectrometry (ESI-MS).ResultsDatabase searching of the MS/MS data resulted in the identification of 2777 proteins which were clustered into groups associated with disease progression. Proteins which were changed in all disease stages including benign, and hence indicative of the earliest molecular perturbations, were strongly associated with spliceosomal activity, cell cycle division, and stromal and cytoskeleton disruption reflecting increased proliferation and expansion into the surrounding healthy tissue. Those proteins changed in cancer stages but not in benign, were linked to inflammation/immune response, loss of cell adhesion, mitochondrial function and autophagy, demonstrating early evidence of cells within the nutrient-poor solid mass either undergoing cell death or adjusting for survival. Caveolin-1, which decreased and Matrix metalloproteinase-9, which increased through the three disease stages compared to normal tissue, was selected to validate the proteomics results, but significant patient-to-patient variation obfuscated interpretation so corroborated the contradictory observations made by others.ConclusionNevertheless, the study has provided significant insights into CC stage progression for further investigation.

Background/aim: Diabetic foot ulcers and related complications are a major cause of morbidity and hospital admissions. Our aim was to evaluate the risk factors associated with poor outcome in diabetic foot ulcers. Materials and methods:A prospective study was conducted on patients with diabetic foot ulceration attending the Madinah Teaching Hospital from June 2014 to December 2015. Potential risk factors and laboratory test results at presentation were recorded and their association with outcome (healing vs. amputation) was analyzed using IBM SPSS Statistics for Windows, Version 22.0.Results: In total, 112 patients were studied during our study period. The majority of the patients were male (60.7%) and aged 50 years and older (62.5%). Regarding the outcome, 68% healed completely, 27.7% underwent amputation, and 4.5% died during this period. Patient age of 50 and older, long duration of diabetes (>10 years), rural origin, and heel ulcers were significantly associated with poor outcome (P < 0.05). Conclusion:Patients with diabetes should have a detailed annual foot examination; those having risk factors for poor outcome require more frequent foot care, patient education, and early referral to tertiary care centers.