The Role of Epigenetic Regulation and Pluripotency-Related MicroRNAs in Differentiation of Pancreatic Stem Cells to Beta Cells

In this study, we aimed to research the effects of class-I HDACs and glucose on differentiation of pancreatic islet derived mesenchymal stem cells (PI-MSCs) to beta cells. Beta cell differentiation determined by flow cytometric analysis and gene expression levels of PDX1, PAX4, PAX6, NKX6.1, NGN3, INS2, and GLUT2. As a result the valproic acid, is an inhibitor of class-I HDACs, caused the highest beta cell differentiation in PI-MSCs. However, the cells in this group were at early stages of differentiation. Glucose co-administration to this group carried the differentiation to higher levels, but these newly formed beta cells were not functional. Moreover, reduction in the levels of pluripotency factors that Oct3/4, c-Myc, and Nanog were parallel to beta cell differentiation. Also, the levels of HDAC1 and acetylated H3/H4 were increased and methylated H3 was decreased by VPA treatment. In addition, we have detected over expression in genes of miR-18a-5p, miR-19b-5p, miR-30d-3p, miR-124, miR-146a-5p, miR-184, miR-335, and miR-433-5p in parallel to beta cell differentiation. As the conclusion, this study is important for understanding the epigenetic mechanism that controls the beta cell differentation and it suggests new molecules that can be used for diagnosis, and treatment of diabetes. J. Cell. Biochem. 119: 455-467, 2018. (c) 2017 Wiley Periodicals, Inc

4-Methylcatechol prevents streptozotocin-induced acute kidney injury through modulating NGF/TrkA and ROS-related Akt/GSK3 beta/beta-catenin pathways

Nerve growth factor (NGF) has been shown to protect the viability of kidney cells in acute phase of renal damage. However, since the half-life of NGF is very short, it is too large to pass the blood-brain barrier and rapidly transported to the liver for catabolizing its use in therapy is limited. 4-Methylcatechol (4MC) is a substance that increases NGF synthesis in many tissues. This study aimed to investigate the protective effects of 4MC against acute renal injury induced by streptozotocin (STZ). We have investigated the profibrotic, proinflarnmatory, oxidative changes in STZ-induced acute renal damage and the possible role of the NGF/TrkA system and Akt/GSK beta/beta-catenin pathway in this mechanism. Experiment was designed as to be started with injection of 4MC for 10 days as a single dose (10 mu g/kg) per day and to be terminated after 4 h of a single dose (75 mg/kg) STZ injection. As the result, 4MC pre-treatment decreased kidney damage, ROS production, the renal levels of TGF beta 1, CD68, tumor necrosis factor-alpha and interleukin 1 beta. Moreover, 4MC pre-treatment increased levels of NGF and its receptor TrkA, p-Akt (Thr308), p-GSK3 beta (Ser9) and nuclear beta-catenin. These data suggest that 4MC prevents the development of STZ-induced renal damage by suppressing ROS production and inflammation via Akt/GSK3 beta/beta-catenin pathway which may be stimulated by NGF/TrkA signaling. Therefore, 4MC can be suggested as a potential agent for the prevention of acute renal injury

Tub and beta-catenin play a key role in insulin and leptin resistance-induced pancreatic beta-cell differentiation

The aim of this study was to investigate the molecular mechanism of pancreatic islet-derived mesenchymal stem cell (PID-MSC) differentiation into beta-cells in the presence of insulin and leptin resistance stimulators. We determined that beta-cell differentiation was stimulated by glucose, insulin, and leptin. Co-administration of insulin and leptin resulted in greater, at a further stage of differentiation but non-functional beta-cell formation. The levels of p-AKT(Ser473) did not change; SOCS3, PTP1B, p-IRS1(Ser307), PTEN levels increased and p-IRS1(Try) levels decreased due to insulin and leptin co-administration. These findings suggest that co-administration of insulin and leptin to PID-MSCs results in the development of both insulin and leptin resistance together. We showed that this differentiation signaling is mainly mediated by AKT/GSK-3 beta/beta-catenin and Tub. Moreover, beta-catenin and Tub were linked to each other in the nucleus under this condition. Furthermore, we found that Tub and beta-catenin contributes to insulin production by increasing the expression of transcription factors by binding to the promoter regions of ins1, ins2, and pdx1 genes. In addition, Tub is also bound to the promoter region of the MafA gene. These findings demonstrate that when insulin and leptin resistance develop together in rat PID-MSCs beta-cell differentiation increases markedly via beta-catenin and Tub. New therapeutic agents that inhibit AKT/GSK-3 beta/beta-catenin and in particular Tub may help prevent the development or retard the progression of type 2 diabetes

UR-144, synthetic cannabinoid receptor agonist, induced cardiomyoblast toxicity mechanism comprises cytoplasmic Ca2+ and DAPK1 related autophagy and necrosis

UR-144, a cannabinoid receptor agonist, is widely used alone or in combination with other synthetic cannabinoids (SCs) all over the world. At overdose, cardiovascular symptoms have been reported and the underlying molecular mechanisms of these adverse effects are not known. It is highly important to clarify the toxic effects of UR-144 for the treatment of poisoning. In the present study, the molecular mechanism of cytotoxic effects of UR-144 is evaluated on a cardiomyoblastic cell line using WST-1 and LDH assays. Apoptosis/necrosis, autophagy, and ROS (reactive oxygen species) levels were determined using flow cytometry. Cytoplasmic Ca2+ levels were measured by using a fluorogenic calcium-binding dye. Released and cytoplasmic troponin T levels, a specific marker of cardiotoxicity, were examined with western blot. For the evaluation of the role of DAPK1, on UR-144-induced cell death, DAPK1 activity and DAPK1 protein level were investigated. Its cytotoxic effects increased in a dose-dependent manner for WST-1 and LDH assays, while membrane damage, one of the signs of necrotic cell death, was more remarkable than damage to mitochondria. Cytoplasmic Ca2+ levels rose after high-dose UR-144 treatment and inhibition of DAPK1 activity ameliorated UR-144-induced cytotoxicity. Released troponin T significantly increased at a dose of 200 mu M. ROS and total antioxidant capacity of cells were both reduced following high dose UR-144 treatment. The results indicated that UR-144-induced autophagic and necrotic cell death might be a consequence of elevated cytoplasmic Ca2+ levels and DAPK1 activation. However, in vivo/clinical studies are needed to identify molecular mechanisms of cardiotoxic effects of UR-144

2D and 3D cultured human umbilical cord-derived mesenchymal stem cell-conditioned medium has a dual effect in type 1 diabetes model in rats: immunomodulation and beta-cell regeneration

Abstract Background Type 1 diabetes (T1D) is a T-cell-mediated autoimmune disease characterized by the irreversible destruction of insulin-producing β-cells in pancreatic islets. Helper and cytotoxic T-cells and cytokine production, which is impaired by this process, take a synergetic role in β-cell destruction, and hyperglycemia develops due to insulin deficiency in the body. Mesenchymal stem cells (MSCs) appear like an excellent therapeutic tool for autoimmune diseases with pluripotent, regenerative, and immunosuppressive properties. Paracrine factors released from MSCs play a role in immunomodulation by increasing angiogenesis and proliferation and suppressing apoptosis. In this context, the study aims to investigate the therapeutic effects of MSC’s secretomes by conditioned medium (CM) obtained from human umbilical cord-derived MSCs cultured in 2-dimensional (2D) and 3-dimensional (3D) environments in the T1D model. Methods First, MSCs were isolated from the human umbilical cord, and the cells were characterized. Then, two different CMs were prepared by culturing MSCs in 2D and 3D environments. The CM contents were analyzed in terms of total protein, IL-4, IL-10, IL-17, and IFN-λ. In vivo studies were performed in Sprague-Dawley-type rats with an autoimmune T1D model, and twelve doses of CM were administered intraperitoneally for 4 weeks within the framework of a particular treatment model. In order to evaluate immunomodulation, the Treg population was determined in lymphocytes isolated from the spleen after sacrification, and IL-4, IL-10, IL-17, and IFN-λ cytokines were analyzed in serum. Finally, β-cell regeneration was evaluated immunohistochemically by labeling Pdx1, Nkx6.1, and insulin markers, which are critical for the formation of β-cells. Results Total protein and IL-4 levels were higher in 3D-CM compared to 2D-CM. In vivo results showed that CMs induce the Treg population and regulate cytokine release. When the immunohistochemical results were evaluated together, it was determined that CM application significantly increased the rate of β-cells in the islets. This increase was at the highest level in the 3D-CM applied group. Conclusion The dual therapeutic effect of MSC-CM on immunomodulation and homeostasis/regeneration of β-cells in the T1D model has been demonstrated. Furthermore, this effect could be improved by using 3D scaffolds for culturing MSCs while preparing CM

Aspartame induces cancer stem cell enrichment through p21, NICD and GLI1 in human PANC-1 pancreas adenocarcinoma cells

This study aimed to investigate the molecular effects of the common natural sugar glucose and artificial sweetener aspartame on cancer stem cell (CSC) population and cancer aggressiveness of PANC-1 human pancreas adenocarcinoma cells. According to our findings while aspartame exposure significantly increased the CSC population, high glucose had no effect on it. The epithelial-mesenchymal transition marker N-cadherin increased only in the aspartame group. The findings indicate that a high level of glucose exposure does not effect the invasion and migration of PANC-1 cells, while aspartame increases both of these aggressiveness criteria. The findings also suggest that a high concentration of glucose maintains CSC population through induction of nuclear Oct3/4 and differentiation to parental cells via increasing cytoplasmic c-myc. Aspartame exposure to PANC-1 cells activated AKT and deactivated GSK3 beta by increasing levels of ROS and cytoplasmic Ca+2, respectively, through T1R2/T1R3 stimulation. Then p-GSK3 beta(Ser9) boosted the CSC population by increasing pluripotency factors Oct3/4 and c-myc via NICD, GLI1 and p21. In the aspartame group, T1R1 silencing further increased the CSC population but decreased cell viability and suppressed the p21, NICD and GLI activation. The presence and amount of T1R subunits in the membrane fraction of PANC-1 cells are demonstrated for the first time in this study, as is the regulatory effect of T1R1's on CSC population. In conclusion, the present study demonstrated that long-term aspartame exposure increases CSC population and tumor cell aggressiveness through p21, NICD, GLI1. Moreover, while aspartame had no tumorigenic effect, it could potentially advance an existing tumor