Differentiation of human mesenchymal stem cells into insulin producing cells by using A lentiviral vector carrying PDX1

Type I diabetes is an immunologically-mediated devastation of insulin producing cells [IPCs] in the pancreatic islet. Stem cells that produce beta-cells are a new promising tool. Adult stem cells such as mesenchymal stem cells [MSCs] are self renewing multi potent cells showing capabilities to differentiate into ectodermal, mesodermal and endodermal tissues. Pancreatic and duodenal homeobox factor 1 [PDX1] is a master regulator gene required for embryonic development of the pancreas and is crucial for normal pancreatic islets activities in adults. We induced the over-expression of the PDX1 gene in human bone marrow MSCs [BM-MSCs] by Lenti-PDX1 in order to generate IPCs. Next, we examine the ability of the cells by measuring insulin/c-peptide production and INSULIN and PDX1 gene expressions. After transduction, MSCs changed their morphology at day 5 and gradually differentiated into IPCs. INSULIN and PDX1 expressions were confirmed by real time polymerase chain reaction [RT-PCR] and immunostaining. IPC secreted insulin and C-peptide in the media that contained different glucose concentrations. MSCs differentiated into IPCs by genetic manipulation. Our result showed that lentiviral vectors could deliver PDX1 gene to MSCs and induce pancreatic differentiation

[Plasma levels of osteocalcin and retinol binding protein-4 in patients with medullary thyroid carcinoma]

Thyroid carcinoma is the most frequent malignant tumor of the endocrine system in human body and accounts for nearly 1% of all cancers. Medullary thyroid carcinoma is the third frequent of thyroid cancer and accounts about 5-8% of thyroid cancer. Osteocalcin, known as a Bone Gamma-carboxyglutamic Acid-containing Protein [BGLAP], is the most non collagenous protein. Retinol binding proteins are the family of proteins that have diverse actions but mainly transport retinol in human body. In this study to evaluate effect of existence medullary thyroid carcinoma on metabolism of bone and adipose tissue, plasma level of two mentioned proteins had analyzed. Population in this study consists of 46 individuals with medullary thyroid carcinoma and 44 healthy subjects referred individuals to Research Institute for Endocrine Sciences, Shahid Beheshti University of Medical Sciences. People with the disease after diagnosis of medullary thyroid carcinoma and pathologically confirmed by biopsy in the initial stages of the study were called. After informed consent, 10 ml of blood from the antecubital vein of left hand in sitting position obtained and after centrifugation, plasma was isolated from all samples until analyzed kept in the freezer. Plasma levels of hormones were measured by sandwich type ELISA method. Obtained results were analyzed by SPSS version 16 with independent t-test method. Mean plasma level of osteocalcin in patients was 33.1 +/- 3.5 and in healthy subjects was 12.5 +/- 1.2 ng/ml [Mean +/- SD] and Odds Ratio [OR] value was 1.04. In patients, mean plasma level of retinol binding protein was 82.5 +/- 2.7 and in healthy subjects was 22.8 +/- 1.6 mg/ml and OR value was 2.1. The confidence level considered at 95%. These differences of plasma levels were statistically significant [P=0.001]. According to difference between plasma levels of osteocalcin and retinol binding protein-4 in patients suffered of medullary thyroid carcinoma comparison with normal subjects, it can be said that, probably medullary thyroid carcinoma has effect on bone and adipose tissue metabolism, so osteocalcin and retinol binding protein-4 hormones have potential to be used for confirmation of diagnosis or following treatment of medullary thyroid carcinoma

[Effects of eccentric and concentric exercises on some blood biochemical parameters in patients with type 2 diabetes]

About 7% of world populations suffer from diabetes disease and its widespread complications. There are several studies about the beneficial effects of aerobic and resistance exercises on blood glucose and lipids levels, insulin sensitivity in type 2 diabetic patients. However, there is no study to investigate the effects of eccentric exercise in these patients. This study was designed to compare the effect of eccentric and concentric exercises on some of the blood biochemical parameters in type 2 of diabetic patients. 28 type 2 diabetic patients were assigned randomly in one of the two experimental eccentric and concentric groups. Before and after control and also after intervention, blood sugers, HbA1c, lipids and body mass index were measured. The results showed that eccentric and concentric exercises result in a significant decrease in blood sugar, HbA1c, and blood lipids. Eccentric exercise training also significantly reduced blood sugar levels and HbA1c compared to the concentric exercise [P<0.0001]. Moreover, no significant changes were found in blood lipids levels in regard to the type of exercise. Our findings show that eccentric exercise is more effective than concentric exercise with respect to reducing and controlling blood glucose levels in type 2 diabetic patients

Tehranolide could shift the immune response towards Th1 and modulate the intra-tumor infiltrated T regulatory cells

Artemisia diffusa contains a new type of sesquiterpene lactone with an endoperoxide group [Tehranolide]. Due to the existing similarity between the structures of Tehranolide and Artemisinin, it was hypothesized that Tehranolide would have similar effects as Artemisinin. In this study, the immunotherapeutic effectiveness of Tehranolide was investigated by direct intra-tumoral injection. Tehranolide was purified from Artemisia diffusa, and its effect on the tumor volume was investigated. The splenocyte proliferation, shifting of cytokine profile, and the presence of naturally-occurring CD4+CD25+Foxp3+ Treg cells were assessed to describe the anti-tumor immune response. Analysis of immune response showed that, intra-tumoral injection of Tehranolide decreased the rate of tumor growth compared to control group. Furthermore, the proliferative response of mice treated with Tehranolide was enhanced. In comparison with the control group, production of both IL-4 and IFN-gamma was induced [p<0.05]. The results indicated a decrease in tumor CD4+CD25+Foxp3+ T lymphocytes in the Tehranolide-treated group compared to the control group. Treatment of tumors with Tehranolide attenuated CD4+CD25+Foxp3+ Treg cell-mediated immune suppression and elicited a persistent anti-tumor immunity against cancer

Immune response following oral immunization with BCG encapsulated in alginate microspheres

Different methods have been used for BCG vaccination. Alginate microspheres are useful in delivery of vaccines to the gastrointestinal tract by oral route. To compare the immune response following oral microencapsulated and subcutaneous [SC] route administration of BCG vaccine in BALB/c mice. Alginate microspheres were produced by an internal emulsification method within olive oil. Four groups of mice were studied, including two groups receiving oral gavages of microencapsulated and free BCG, one receiving SC injection of BCG, and a control group. T cell proliferation, specific anti-BCG total IgG, and IgG subclasses [IgG1 and IgG2a] were compared between groups 5 and 12 weeks after vaccination. The best result was achieved using oral microencapsulated form in comparison with oral BCG alone. Delivery of oral BCG with alginate microspheres is an effective way to induce immune response in BALB/c mice

Immunobiological consequences of sulfur mustard contamination

Sulfur mustard has been employed in chemical warfare in certain regions including Iran. The short and long term biological effects of sulfur mustard contamination have been studied in both basic and clinical aspects. Sulfur mustard has been shown to induce a vast array of pathological effects in affected persons. In addition to skin, lung, eyes and gastrointestinal disturbances, sulfur mustard has been shown to induce hematological complications and a severe suppression of the immune system. The short and long term immunological [both cellular and humoral], hematological, genetic and biochemical consequences of persons exposed to sulfur mustard are extensively reviewed here. The long term complications of these patients indicate the need to develop effective preventive and therapeutic strategies in the clinic. These strategies may be based upon immunopotentiating intervention and therapy

Multilineage differentiation activity by the human umbilical vein-derived mesenchymal stem cells

Mesenchymal stem cells [MSC] are a very promising transplantable stem cell source for a variety of cell replacement therapies. As the main source of MSC is bone marrow [BM], most of studies have been done on BM-derived MSC [BM-MSC]. Umbilical cord [UC]-derived MSC [UC-MSC] which are recently introduced, is one of the good alternative source for these cells. The objective of this study was to isolate and characterize UC-MSC from human UC veins and studying of their potential to differentiate into various cell types such as fat, bone, cartilage and neuronal cells. In this way, a cell population was isolated from 20 UC veins using a solution of 0.1% collagenase type IV. After identification of isolated cells by immunocytochemical and flow cytometry methods, these cells were exposured with adipogenic, osteogenic, chondrogenic and neurogenic agents. Resulted differentiated cells were detected using specific staining for each lineage and room temperature [RT]-PCR. Immunophenotypically, this cell population was positive for CD73, CD 166, CD 105 markers and alpha-smooth muscle actin and negative for CD31, CD34, CD49d markers, von Willebrand factor and smooth muscle myosin. Exposure of these cells to adipogenic, osteogenic, chondrogenic and neurogenic agents resulted in morphological changes followed by lineage-specific staining for each differentiated cell type. RT-PCR analysis showed that these differentiated cells express fat, bone, cartilage and neuronal markers, respectively. Altogether, these findings indicate that UC-MSC possess morphological, immunophenotypical and cell differentiation capacities similar to BM-MSC and so they can be used more extensively in cell based therapy protocols and in vitro differentiation study models

KatG mutation of isoniazid-resistant isolated from tuberculosis patients

The emergence of drug-resistant strains of Mycobacterium tuberculosis [MTB] is an increasing problem in developed and developing countries. The aims of the present study were to identify various types of mutations in katG region from 28 MDR strains isolated from sputum of tuberculosis patients. Twenty-eight rifampin-resistant strains isolated from sputum of patients with active pulmonary tuberculosis were obtained from various geographic regions of Iran. Drug susceptibility was determined by using the BACTEC system. DNA extraction, standard PCR identification, katG gene amplification, DNA sequencing and analysis were done. There was no mutation in 2 strains. In 20 strains, mutation was shown to be in codon 315. Three types of mutations were detected consisting of AGC-ACC [Ser-Thr] [80%], AGC-AGG [Ser-Arg] [5%] and AGC-AAC[Ser-Asn] [15%].Furthermore, one type of mutation was seen in codons 311, 299, and 323. Twelve strains showed one mutation in codon 315 [46%], 7 strains 2 mutations [27%], 5 isolate 3 mutations [19%] and in 2 strains 4 mutations [8%] were observed in different codons. Nine silent mutations was demonstrated in codon 311[GAC1TAC]. This research demonstrated that mutations were mostly seen in codons 315 and 299 indicating resistance to isoniazide

Isolation, culture and characterization of postnatal human umbilical vein-derived mesenchymal stem cells

On the basis of reports that mesenchymal stem cells [MSCs] can be isolated from the placenta/umbilical cord stroma, the present study was undertaken to isolate and characterize MSCs from the human umbilical cord veins. In this investigation, a cell population was isolated which was derived from the endothelium/subendothelium layers of 20 umbilical cord veins obtained from term deliveries using a solution of 0.1% collagenase type IV. Results suggest that these cells possess morphological, immunophenotypical and cell differentiation capacities similar to the bone marrow-derived mesenchymal stem cells [MSCs]. The isolated cell population has fibroblastoid morphology which upon proper stimulation gives rise to adipocytes, osteocytes and chondrocytes in culture. Immunophenotypically, this cell population is positive for CD54, CD29, CD73, CD49e, CD166, CD105, CD13, and CD44 markers and alpha-smooth muscle actin and negative for CD31, CD45, CD49d, and CD34 markers, von Willebrand factor [vWF] and smooth muscle myosin [MySM]. Altogether, these findings indicate that umbilical cord obtained from term deliveries is an important source of MSCs which could have an important application in cell therapy protocols

apoptotic effect of extracellular zinc sequestration on HT29/219 and SW742 cell lines

Zn [II] is an important regulator of caspase-3, as well as an antioxidant, microtubule stabilizer, growth cofactor, and anti-inflammatory agent. Over the past 30 years, many researchers have demonstrated the important role of Zn [II] in a variety of physiological processes, including growth and development, maintenance and priming of the immune system, and in tissue repair and regeneration. In this study, we present evidence that chelation of extracellular zinc by diethylenetriaminepentacetic acid [DTPA] in different concentrations causes cell death in carcinoma cell lines, HT29/219 and SW742. Hoechst 33258 staining revealed that cell death was mainly by apoptosis. Additionally, significant increases in the activity of caspase-3 and -9 were observed in both cell lines. Caspase-8 activation was negligible in both cell lines. The cytotoxicity/apoptotic effect of DTPA was inhibited significantly by Zn [II], Cu [II] and N-Acetyl-L-Cysteine [NAC] [P<0.05]. Therefore, DTPA, the membrane-impermeable metal ion chelator, induces apoptosis through the depletion of extracellular zinc ion

Effects of glucosamine on pancreatic glucokinase and hexokinase activities and their relationship to insulin secretion from isolated islets of normal and type 2 diabetic rat

Glucokinase serves as a glucose sensor in pancreatic beta-cells and plays a key role in glucose homeostasis and glucose-stimulated insulin secretion [GSIS]. In the present study we examined the effect of glucosamine, a glucokinase inhibitor, on the pancreatic glucokinase and hexokinase activities and on insulin secretion from freshly rat pancreatic islets of Langerhans. Insulin concentration was measured by rat insulin ELISA kit. The pancreatic islets from normal and type 2 diabetic [nSTZ] rats were isolated by collagenase digestion method. Glucose phosphorylation was quantitated by measuring the rate of glucose-6-phosphate formation in the fluorometric assay. Insulin secretion from hand-picked islets was evaluated in static incubation system. Insulin concentration was measured by rat insulin ELISA kit. Our findings demonstrate that glucosamine in a dose dependent manner, reduced glucokinase activity in islet extract, but had no effect on hexokinase activity. The glucose-stimulated insulin secretion was inhibited by glucosamine but it had no effect on the basal insulin secretion. In diabetic rats glucokinase was decreased while the basal insulin secretion and the activity of hexokinase were higher than normals. Based on results obtained from the present study, the assumption could be made that the decrease in the activity of glucokinase of pancreatic islets could be related to the impaired glucose stimulated insulin secretion. The increase in basal insulin secretion of diabetic rats may be due to an increase in pancreatic hexokinase activity

Collagen as adherent substratum and inducer of dorsal root ganglia outgrowth

Neurite outgrowth from dorsal root ganglion [DRG] explants is a method of evaluating neurotrophic activity of growth factors. When complete medium containing collagen was supplemented with nerve growth factor [NGF] DRG outgrowth was observed after 18 h. In the absence of NGF and in the presence of collagen, the DRG outgrowth took place after 72 h. In wells not supplemented with collagen gel in substratum, no DRG outgrowth was observed. Partially, DRG differentiation was observed in the presence of NGF. In the absence of NGF and collagen, there was no DRG outgrowth detected. It seems that, in some circumstances, cells degenerated by DRG may be an indication of an apoptosis phenomenon. Therefore, we suggested that collagen as a substratum is more effective than NGF