Study of long non-coding RNA and mitochondrial dysfunction in diabetic rats.

Diabetes mellitus (DM) is a worldwide health problem. The Micro- and macro-vascular complications are the major causes of morbidity and mortality of DM. Molecular regulation of mitochondrial fission/fusion cycles is being studied, but the results were not conclusive. The aim of this study is to investigate the possible functional role of lncRNA H19 and its relation to mitofusin-2 (Mfn-2) gene expression in diabetic rats with cardiac and renal complications. Streptozotocin-induced diabetic male, albino rats and a matched control group were investigated. Cardiac weights, blood pressure and ECG were recorded. Biochemical evaluation of cardiac and renal functions was performed. Molecular determination of lncRNA H19 and Mfn-2 gene expression and histological examination by light and electron microscopy for cardiac and renal tissues were performed. Diabetic rats showed a significant increase of left ventricle weight/whole body weight ratio, R wave voltage, and a significant decrease of blood pressure, heart rate, and P wave voltage. At the molecular level, lncRNA H19 and Mfn-2 mRNA showed altered expression with a statistically significant downregulation of Mfn-2 mRNA expression in renal tissues. In conclusion, the changes in lncRNA H19 and Mfn-2 mRNA expression may help better understanding of the pathogenesis of cardiac and renal dysfunctions associated with type 1 DM.

CRISPR/Cas9 mediated knock-out of VPREB1 gene induces a cytotoxic effect in myeloma cells.

BACKGROUND: Multiple Myeloma (MM) is a heterogeneous, hematological neoplasm that accounts 2% of all cancers. Although, autologous stem cell transplantation and chemotherapy are currently the most effective therapy, it carries a notable hazards, in addition for being non curative. Recently, the Clustered Regular Interspaced Short Palindromic Repeats (CRISPR-cas9) has been successfully tried at the experimental level, for the treatment of several hematological malignancies. OBJECTIVES: We aimed to investigate the in-vitro effect of CRISPR-cas9-mediated knock-out of V-set pre B-cell surrogate light chain 1"VPREB1" gene on the malignant proliferation of primary cultured myeloma cells. METHODS: Bioinformatics' analysis was performed to explore the gene expression profile of MM, and the VPREB1 gene was selected as a target gene for this study. We knocked-out the VPREB1 gene in primary cultured myeloma cells using CRISPR-cas9, the VPREB1 gene editing efficacy was verified by determining VPREB1 gene expression at both the mRNA and protein levels by qPCR and immunofluorescence, respectively. Furthermore, the cytotoxic effect on primary myeloma cells proliferation was evaluated using cytotoxicity assay. RESULTS: There was a statistically significant reduction of both VPREB1 mRNA and protein expression levels (p<0.01). knock-out of VPREB1 gene in myeloma cell line resulted in a statistically significant reduction of myeloma cell proliferation. CONCLUSION: CRISPR-cas9-mediated knock-out of VPREB1 gene is effective for inhibiting the proliferation of primary myeloma cells. This would provide a basis for a promising therapeutic strategy for patients with multiple myeloma.

Genotype-phenotype correlation of 33 patients with maple syrup urine disease.

Maple syrup urine disease (MSUD) is a rare autosomal recessive inherited disorder due to defects in the branched-chain α-ketoacid dehydrogenase complex (BCKDC). MSUD varies in severity and its clinical spectrum is quite broad, ranging from mild to severe phenotypes. Thirty-three MSUD patients were recruited into this study for molecular genetic variant profiling and genotype-phenotype correlation. Except for one patient, all other patients presented with the classic neonatal form of the disease. Seventeen different variants were detected where nine were novel. The detected variants spanned across the entire BCKDHA, BCKDHB and DBT genes. All variants were in homozygous forms. The commonest alterations were nonsense and frameshift variants, followed by missense variants. For the prediction of variant's pathogenicity, we used molecular modeling and several in silico tools including SIFT, Polyphen2, Condel, and Provean. In addition, six other tools were used for the prediction of the conservation of the variants' sites including Eigen-PC, GERP++, SiPhy, PhastCons vertebrates and primates, and PhyloP100 rank scores. Herein, we presented a comprehensive characterization of a large cohort of patients with MSUD. The clinical severity of the variants' phenotypes was well correlated with the genotypes. The study underscores the importance of the use of in silico analysis of MSUD genotypes for the prediction of the clinical outcomes in patients with MSUD.

Percutaneous embolization of cystic duct stump leak following failed endoscopic management.

A case of a 79-year-old man, status post laparoscopic cholecystectomy with a drainage catheter placed at the gallbladder fossa is presented. The case was complicated postoperatively by abdominal pain and bilious discharge from the drainage catheter. Endoscopic retrograde cholangio-pancreatography demonstrated leakage through the cystic duct stump into the gallbladder fossa. Placement of a covered metal stent endoscopically failed to seal the leak. We performed percutaneous embolization of the cystic duct stump using a combination of coils and gelatin sponge through the drainage catheter in the gallbladder fossa. To our knowledge, this technique has not been previously described in the literature.

A proteomic analysis of excreted and circulating proteins from obese patients following two different weight-loss strategies.

Bariatric surgery is the most successful therapeutic approach to weight loss, but how it leads to weight loss, and how it resolves obesity-related complications, including type-2 diabetes, are poorly understood. This study, comprising two groups of individuals, one on a low-calorie diet (n = 5) and one undergoing bariatric surgery (n = 7), used both targeted and untargeted proteomic approaches to determine changes in protein levels pre- and post-intervention (i.e. 3-6 months later). Changes were observed in both circulating and excreted proteins following weight loss. Targeted multiplexed biochip arrays measured 12 plasma peptides/proteins involved in metabolism and inflammation: C-peptide, ferritin, interleukin-6, interleukin-1 alpha, resistin, insulin, tumor necrosis factor alpha, leptin, plasminogen-activator inhibitor-1, adiponectin, cystatin C, and C-reactive protein. Following a low-calorie diet, plasma insulin and C-reactive protein levels were significantly reduced (P = 0.045 and P = 0.030, respectively); adiponectin increased and leptin decreased following surgery (P = 0.014 and P = 0.005, respectively). Untargeted proteomic analysis employing 2D difference in-gel electrophoresis (DIGE) showed 28 protein spots with ≥1.5-fold changes in expression following weight loss by a low-calorie diet; comparison of pre- and post-intervention urine samples from the bariatric surgery group showed changes in excretion of 110 protein spots. The combination of targeted protein analysis by multiplexed arrays and an exploratory (i.e. an unbiased or discovery) proteomic assessment of hundreds of proteins offers valuable insights into the mechanistic differences between alternative weight-loss strategies. This is a powerful hypothesis-generating approach to study complex, multifactorial syndromes such as obesity. The findings that arise from these studies can then be validated in targeted, hypothesis-directed investigations.

Hypoglycemic effects of date seed extract. Possible mechanism of action, and potential therapeutic implications.

OBJECTIVE: To investigate the possible mechanism, by which an extract from date seeds exert its hypoglycemic effect. METHODS: This study was performed at the Anatomy Department, College of Medicine, King Saud University, Riyadh, Kingdom of Saudi Arabia from May to December 2012. Eighty rats were divided into 4 groups. Group 1 received no treatment. Group 2 received daily ingestions of 10 ml of date seed extract for 8 weeks. Animals of groups 3 and 4 were made diabetic by streptozotocin injection, and were given daily subcutaneous injections of 3 IU/day of insulin for 8 weeks. Group 4 received, in addition, daily ingestions of 10 ml of seed extracts. Rats were sacrificed, and the sera were separated for estimation of serum C-peptide levels. Pancreatic tissues were processed for histological study of the islet cells, immunohistochemical study for insulin secretion and image analysis for insulin quantification. RESULTS: Mean serum C-peptide level was significantly higher in group 4 compared to group 3. Pancreatic islets from rats of group 3 showed weak immunoreactivity for insulin, while those of group 4 showed strong immunoreactivity in some hypertrophied beta cells. Immunopositive cells were detected in the wall of interlobular ducts and in centroacinar cells of pancreas only in group 4. Quantification of insulin immunoreactivity showed a marked reduction in islet size and extent of insulin immunoreactivity in diabetic compared to control groups. CONCLUSION: Date seed extracts may stimulate endogenous insulin secretion through extra-islet sources.

Differential patterns of serum concentration and adipose tissue expression of chemerin in obesity: adipose depot specificity and gender dimorphism.

Chemerin, a recognized chemoattractant, is expressed in adipose tissue and plays a role in adipocytes differentiation and metabolism. Gender- and adipose tissue-specific differences in human chemerin expression have not been well characterized. Therefore, these differences were assessed in the present study. The body mass index (BMI) and the circulating levels of chemerin and other inflammatory, adiposity and insulin resistance markers were assessed in female and male adults of varying degree of obesity. Chemerin mRNA expression was also measured in paired subcutaneous and visceral adipose tissue samples obtained from a subset of the study subjects. Serum chemerin concentrations correlated positively with BMI and serum leptin levels and negatively with high density lipoprotein (HDL)-cholesterol levels. No correlation was found between serum chemerin concentrations and fasting glucose, total cholesterol, low density lipoprotein (LDL)-cholesterol, triglycerides, insulin, C-reactive protein or adiponectin. Similarly, no relation was observed with the homeostasis model assessment for insulin resistance (HOMA-IR) values. Gender- and adipose tissue-specific differences were observed in chemerin mRNA expression levels, with expression significantly higher in women than men and in subcutaneous than visceral adipose tissue. Interestingly, we found a significant negative correlation between circulating chemerin levels and chemerin mRNA expression in subcutaneous fat. Among the subjects studied, circulating chemerin levels were associated with obesity markers but not with markers of insulin resistance. At the tissue level, fat depot-specific differential regulation of chemerin mRNA expression might contribute to the distinctive roles of subcutaneous vs. visceral adipose tissue in human obesity.