L-Ascorbic acid affect the DNA methytransferase expression in Mouse Embryonic Fibroblasts

Introduction: Induced pluripotent stem cells (iPSCs) can be generated from different source of cells with different efficiencies. Two DNA methyltransferases DNMT1 and DNMT3A have been shown to regulate epigenetically the gene expression involved in cell viability and reprogramming. L-ascorbic acid (L-AA) is a chemical factor that can accelerate reprogramming.  Here, we sought to investigate the effect of L-AA on DNMT1 and DNMT3A expressions. Materials and Methods: First, mouse embryonic fibroblasts at passage 3 were cultured in the presence of 10 µg/ml L-AA days for 5 days. Then, DNMT1 and DNMT3A expressions were determined using real-time PCR at days 3 and 5. Results: It was showed that L-AA could enhance DNMT-1 expression which involve in cell viability and decrease the DNMT3A which involve in cell differentiation. Conclusion: The results therefore suggest a new insight into L-AA mechanism impact on reprogramming process

Significant Changes of 5-Hydroxytriptamine 3A Receptor Gene Expression in Peripheral Blood Mononuclear Cells of Allergic Asthmatic Patients

Asthma is a chronic inflammatory disorder of the airways. The stress is a factor for asthma which indicates a disorder in the function of communicational mediators of nervous and  immunological systems  such  as  neurotransmitters.  A  study  indicated  that  blood serotonin concentration increases in asthmatic patients. Other study indicates that one kind of the serotonin  receptors, named 5HT3A, on PBMCs causes secretion of series of pro- inflammatory cytokines which play important  roles in allergic asthma disease. Thus, we evaluated the ratio expression level of 5HT3A subtype receptors in asthma. The Peripheral Blood Mononuclear Cells were separated from whole blood of 30 allergic asthmatic patients and 30 normal controls by a gradient density centrifugation technique, then the total cellular RNA was extracted and the cDNA was synthesized. This process was followed by real-time PCR using primer pairs specific for 5-hydroxytryptamine 3A subtype receptor mRNA and beta-actin as internal control. Results  revealed  that  relative  gene  expression  of  5-hydroxytryptamine 3A  subtype receptor  increased significantly in Peripheral Blood Mononuclear  Cells of  patients  with asthma in comparison with normal individuals. To conclude, considering 5-hydroxytryptamine 3A subtype receptor role in accomplishment of asthma symptoms, this increase in its expression may exacerbate the seriousness of asthma disease

Comparison of Osteogenic Differentiation Potential of Induced Pluripotent Stem Cells and Buccal Fat Pad Stem Cells on 3D-Printed HA/β-TCP Collagen-Coated Scaffolds

Production of a 3D bone construct with high-yield differentiated cells using an appropriate cell source provides a reliable strategy for different purposes such as therapeutic screening of the drugs. Although adult stem cells can be a good source, their application is limited due to invasive procedure of their isolation and low yield of differentiation. Patient-specific human-induced pluripotent stem cells (hiPSCs) can be an alternative due to their long-term self-renewal capacity and pluripotency after several passages, resolving the requirement of a large number of progenitor cells. In this study, a new biphasic 3D-printed collagen-coated HA/β-TCP scaffold was fabricated to provide a 3D environment for the cells. The fabricated scaffolds were characterized by the 3D laser scanning digital microscopy, X-ray diffraction, Fourier transform infrared spectroscopy, and mechanical test. Then, the osteogenesis potential of the hiPSC-seeded scaffolds was investigated compared to the buccal fat pad stem cell (BFPSC)-seeded scaffolds through in vitro and in vivo studies. In vitro results demonstrated up-regulated expressions of osteogenesis-related genes of RUNX2, ALP, BMP2, and COL1 compared to the BFPSC-seeded scaffolds. In vivo results on calvarial defects in the rats confirmed a higher bone formation in the hiPSC-seeded scaffolds compared to the BFPSC-seeded groups. The immunofluorescence assay also showed higher expression levels of collagen I and osteocalcin proteins in the hiPSC-seeded scaffolds. It can be concluded that using the hiPSC-seeded scaffolds can lead to a high yield of osteogenesis, and the hiPSCs can be used as a superior stem cell source compared to BFPSCs for bone-like construct bioengineering

Enhanced Cardiac Differentiation of Human Cardiovascular Disease Patient-Specific Induced Pluripotent Stem Cells by Applying Unidirectional Electrical Pulses Using Aligned Electroactive Nanofibrous Scaffolds

In the embryonic heart, electrical impulses propagate in a unidirectional manner from the sinus venosus and appear to be involved in cardiogenesis. In this work, aligned and random polyaniline/polyetersulfone (PANI/PES) nanofibrous scaffolds doped by Camphor-10-sulfonic acid (β) (CPSA) were fabricated via electrospinning and used to conduct electrical impulses in a unidirectional and multidirectional fashion, respectively. A bioreactor was subsequently engineered to apply electrical impulses to cells cultured on PANI/PES scaffolds. We established cardiovascular disease-specific induced pluripotent stem cells (CVD-iPSCs) from the fibroblasts of patients undergoing cardiothoracic surgeries. The CVD-iPSCs were seeded onto the scaffolds, cultured in cardiomyocyte-inducing factors, and exposed to electrical impulses for 1 h/day, over a 15-day time period in the bioreactor. The application of the unidirectional electrical stimulation to the cells significantly increased the number of cardiac Troponin T (cTnT+) cells in comparison to multidirectional electrical stimulation using random fibrous scaffolds. This was confirmed by real-time polymerase chain reaction for cardiac-related transcription factors (<i>NKX2.5, GATA4</i>, and <i>NPPA</i>) and a cardiac-specific structural gene (<i>TNNT2</i>). Here we report for the first time that applying electrical pulses in a unidirectional manner mimicking the unidirectional wave of electrical stimulation in the heart, could increase the derivation of cardiomyocytes from CVD-iPSCs