The pluripotency gene regulatory network of porcine induced pluripotent stem cells(piPSCs), especially in epigenetics, remains elusive. To determine the biological function of epigenetics, we cultured piPSCs in different culture conditions. We found that activation of pluripotent gene-and pluripotency-related pathways requires the erasure of H3K9 methylation modification which was further influenced by mouse embryonic fibroblast (MEF) served feeder. By dissecting the dynamic change of H3K9 methylation during loss of pluripotency, we demonstrated that the H3K9 demethylases KDM3A and KDM3B regulated global H3K9me2/me3 level and that their co-depletion led to the collapse of the pluripotency gene regulatory network.Immunoprecipitation-mass spectrometry (IP-MS) provided evidence that KDM3A and KDM3B formed a complex to perform H3K9 demethylation. The genome-wide regulation analysis revealed that OCT4 (O) and SOX2 (S), the core pluripotency transcriptional activators, maintained the pluripotent state of piPSCs depending on the H3K9 hypomethylation. Further investigation revealed that O/S cooperating with histone demethylase complex containing KDM3A and KDM3B promoted pluripotency 2 of 18 | ZHU et al.

Self-renewal and differentiation of male germline stem cells (mGSCs) provide the basic function for continual spermatogenesis. Studies of in vitro culture of germline stem cells are important and meaningful for basic biological research and practical application. Growth factors, such as GDNF, bFGF, CSF1, and EGF, could maintain the self-renewal of mGSCs. Insulin-like growth factor 1 (IGF-1), an important growth factor, and its pathway have been reported to maintain the survival of several types of stem cells and play important roles in male reproduction. However, the mechanism through which the IGF-1 pathway acts to regulate the self-renewal of mGSCs remains unclear. We analyzed the effect of IGF-1 on the proliferation and apoptosis of bovine mGSCs. We evaluated the expression profile of long noncoding RNA (LncRNA) H19 in bovine and mouse tissues. Moreover, we investigated whether LncRNA H19 could regulate the IGF-1 pathway. Results showed that IGF-1 could activate the phosphorylation of AKT and ERK signaling pathways, and the IGF-1 pathway played an important role in regulating the proliferation and apoptosis of bovine mGSCs. The proliferation rate of mGSCs decreased, whereas the apoptosis rate of mGSCs increased when the IGF-1 receptor (IGF-1R) was blocked using the IGF-1R-specific inhibitor (picropodophyllin). LncRNA H19 could regulate the IGF-1 signaling pathway and, consequently, the proliferation and apoptosis of mGSCs. The number of cells in the seminiferous tubule decreased when H19 was interfered by injecting a virus-containing supernatant. Hence, LncRNA H19 participated in the regulation of the proliferation and apoptosis of mGSCs via the IGF-1 signaling pathway.

Background Mesenchymal stem cells (MSCs) are promising candidates for tissue regeneration and disease treatment. However, long-term in vitro passaging leads to stemness loss of MSCs, resulting in failure of MSC therapy. This study investigated whether the combination of melatonin and human umbilical cord mesenchymal stem cells (hUC-MSCs) was superior to hUC-MSCs alone in ameliorating high-fat diet and streptozocin (STZ)-induced type II diabetes mellitus (T2DM) in a mouse model. Methods Mice were divided into four groups: normal control (NC) group; T2DM group; hUC-MSCs treatment alone (UCMSC) group and pretreatment of hUC-MSCs with melatonin (UCMSC/Mel) group. Results RNA sequence analysis showed that certain pathways, including the signaling pathway involved in the regulation of cell proliferation signaling pathway, were regulated by melatonin. The blood glucose levels of the mice in the UCMSC and UCMSC/Mel treatment groups were significantly reduced compared with the T2DM group without treatment (P < 0.05). Furthermore, hUC-MSCs enhance the key factor in the activation of the PI3K/Akt pathway in T2DM mouse hepatocytes. Conclusion The pretreatment of hUC-MSCs with melatonin partly boosted cell efficiency and thereby alleviated impaired glycemic control and insulin resistance. This study provides a practical strategy to improve the application of hUC-MSCs in diabetes mellitus and cytotherapy. Graphical abstract Overview of the PI3K/AKT signaling pathway. (A) Underlying mechanism of UCMSC/Mel inhibition of hyperglycemia and insulin resistance T2DM mice via regulation of PI3K/AKT pathway. hUC-MSCs stimulates glucose uptake and improves insulin action thus should inhibition the clinical signs of T2DM, through activation of the p-PI3K/Akt signaling pathway and then regulates glucose transport through activating AS160. UCMSC/Mel increases p53-dependent expression of BCL2, and inhibit BAX and Capase3 protein activation. Leading to the decrease in apoptosis. (B) Melatonin modulated PI3K/AKT signaling pathway. Melatonin activated PI3K/AKT response pathway through binding to MT1and MT2 receptor. Leading to the increase in hUC-MSCs proliferation, migration and differentiation. → (Direct stimulatory modification); ┴ ( Direct Inhibitory modification); → ┤ (Multistep inhibitory modification); ↑ (Up regulate); ↓ (Down regulate); PI3K (Phosphoinositide 3-Kinase); AKT ( protein kinase B); PDK1 (Phosphoinositide-dependent protein kinase 1); IR, insulin receptor; GLUT4 ( glucose transporter type 4); ROS (reactive oxygen species); BCL-2 (B-cell lymphoma-2); PDK1 (phosphoinositide-dependent kinase 1) BAX (B-cell lymphoma-2-associated X protein); PCNA (Proliferating cell nuclear antigen); Cell cycle-associated proteins (KI67, cyclin A, cyclin E)

Objectives The establishment of porcine pluripotent stem cells (pPSCs) is still a critical topic. However, all pPSCs were failed to contribute to efficient chimeric pig and were extremely sensitive to changes of culture conditions. This study aimed to investigate the role of BCL2 in pPSCs and further explain the mechanism. Materials and Methods Porcine BCL2 gene was cloned and overexpressed in porcine induce pluripotent stem cells (piPSCs). Digital RNA‐seq was performed to explain the mechanism of anti‐apoptosis. Finally, the cells carrying BCL2 were injected into mouse early embryo to evaluate its chimeric ability. Results Here, we found that overexpression of porcine BCL2 gene significantly improved the survivability of piPSCs and the efficiency of embryonic chimerism, and did not wreck the pluripotency of piPSCs. Furthermore, the Digital RNA‐seq analysis revealed that BCL2, as a downstream gene of the PI3K signal pathway, enhanced the expression of PI3K signal pathway receptors, such as FGFR2, and further promoted oxidoreductases activity and lipid metabolism, thus maintaining the survival and pluripotency of piPSCs. Conclusion Our data not only suggested that porcine BCL2 gene could enhance the survivability and chimeric ability of pPSCs, but also explained the positive feedback mechanism in this process, providing strong support for the chimeric experiment of pPSCs.

Cell fate is precisely regulated in the developmental process and human disease. Since the discovery of Mendelian, gene regulation models are continuously developing. 1-4 In the beginning, the transcriptional models were linear. 5-8 In the 1980s, the first enhancer was identified which can increase gene expression by 200-fold. 9 With technological development, the study of chromatin is expanded into three dimensions (3D). Distal enhanced elements transmit the activation signal to the promoter by chromosome folding. 10,11 These advances further explained the accuracy of gene expression regulation, enriching theories of transcriptional models. In 2013, Young and colleagues came up with the concept of the super enhancer (SE), as the region in the embryonic stem (ES) cells where Oct4, Sox2 and Nanog co-bind, 12,13 with a huge region of the genome. 14-16 Later, more and more biological macromolecules were found in the formation of SEs, and more biological functions were found to be controlled by them. 17-21 Based on the