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  • Our gene chip analysis also revealed

    2018-10-20

    Our gene chip analysis also revealed a robust upregulation of metallothionein (MT) family members that are involved in metal metabolism and detoxification, as well as reactive oxygen species (ROS) scavenging. Overexpression of MT has been shown to protect mice from streptozotocin (STZ)-induced β cell damage and diabetes (Chen et al., 2001) and to improve islet graft survival following transplantation by protecting islets from hypoxia (Li et al., 2004). Therefore, the robust induction of MT1/2 in grafts from rats may have contributed to the improved function of hESC-derived β cytochrome p450 inhibitors by providing protection from the generally hypoxic transplant environment. Engrafted endocrine cells were more highly vascularized in rats than in mice, which may also contribute to protecting the developing β cells from hypoxia and providing more rapid delivery of nutrients for promoting secretagogue-induced secretory responses. We also demonstrated that MT1 expression is highly inducible in rodent and human β cell lines by various cell stress pathways, including exposure to ROS or cytokines. Interestingly, ROS stimulation has been shown recently to enhance β cell differentiation in pancreatic explants ex vivo, and reducing ROS production in vivo resulted in decreased β cell differentiation (Hoarau et al., 2014). Grafts from mice produced ∼6-fold more IL-8 than rats, a proinflammatory cytokine that is highly secreted from human islets after culture and into the serum of human islet transplant recipients (Citro et al., 2012). Notably, peri-transplantation administration of reparixin (an IL-8 receptor inhibitor) improved islet graft function and survival in both mice and humans (Citro et al., 2012). Therefore, reducing IL-8 secretion by hESC-derived grafts or preventing IL-8 action post-transplantation may be beneficial to progenitor cell development. Taken together, these studies provide several clues for understanding the accelerated development of hESC-derived, glucose-responsive β cells within rats, including improved integration of the ECM and vasculature with engrafted cells; induction of metallothionein proteins as a protective response to cellular stress; exposure to lower circulating glucose levels in vivo; higher endogenous thyroid hormone levels, which may contribute to induction of MAFA; and reduced IL-8 secretion by grafts. Clearly further work is required to elucidate the exact mechanism(s) underlying the in vivo development of hESC-derived β cells. These studies also highlight the susceptibility of pancreatic progenitor cells to variation within the host environment. Implantation of progenitor cells into closely related but different species revealed substantial differences in the fate choice between pancreatic endocrine versus exocrine lineages as well as α versus β cell lineages. Because the maturation environment may differ amongst human recipients, future studies should continue to investigate the role of host physiology on progenitor cell differentiation and function following transplantation. Efforts to generate bona fide mature human β cells in vitro (Pagliuca et al., 2014; Rezania et al., 2014; Russ et al., 2015) should also be pursued to minimize the maturation window required in vivo.
    Experimental Procedures
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    Acknowledgments
    Introduction Ataxia-telangiectasia (A-T) is caused by the loss of function of the ATM gene, usually as a heterozygotic combination of two allelic mutations. In most cases, ATM mutations lead to expression of truncated proteins through early termination, but there are a large number of variations in mutation sites (Concannon and Gatti, 1997; Jacquemin et al., 2012). Studies of functional properties of A-T missense mutations found varying ATM protein levels and abnormal enrichment of cytoplasmic ATM protein (Jacquemin et al., 2012). Mouse models of A-T have been limited to standard knockout strategies that are normally homozygotic (Barlow et al., 1996; Xu et al., 1996). Because mutations may lead to expression of a portion of the ATM protein or even a full-length ATM protein lacking key regulatory sites, there is a possibility of partial function or dominant-negative function in A-T. For this reason, we sought to develop a collection of human A-T induced pluripotent stem cells (iPSC) for functional studies.