Non-viral direct reprogramming strategies for the creation of insulin producing cells for type 1 diabetes
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Date
2020-12
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The Ohio State University
Abstract
According to the CDC and the American Diabetes Association (ADA), Diabetes affects approximately 10% of the United States population and totals $327 Billion in annual costs. Type 1 Diabetes (T1D) is an autoimmune disorder resulting in loss of pancreatic beta-cells which leads to an insulin deficiency and affects approximately 1.6 million Americans. Patients with T1D diabetes rely on careful glucose monitoring and insulin injections typically 3-4 times a day. Poor patient adherence to this process can lead to dysglycemia and a multitude of other health complications. Current advancements in cell therapies show promise in the development of insulin producing cells (IPCs) which could be used to treat T1D. However, the use of stem cells and direct reprogramming methods face several clinical hurdles (e.g, tumorigenicity, biosafety concerns). Here, we show the creation of IPCs both in vitro and in vivo using electroporation-based direct reprogramming approaches, avoiding those translational hurdles.
Human dermal fibroblasts were transfected, in vitro, with a combination of pro-IPC genes (TCF3, Pdx1, Mafa, NeuroG3, and Pax4) or a sham plasmid (pCMV6). The transfected cells were cultured in differentiation media for 3 weeks. Results indicate that co-transfection of TCF3 along with Pdx1+NeuroG3+Mafa (T+PNM) or Pdx1+NeuroG3+Mafa+Pax4 (T+PNMP) lead to morphology and phenotypic changes in the transfected fibroblast compared to other experimental groups.
The dorsal skin of Streptozotocin (STZ)-treated mice were tissue transfected by electroporation with a pro-insulinogenic gene cocktail (TCF3, Pdx1, Mafa, and NeuroG3) or a sham plasmid. Fasting glucose and weight measurements were used to assess insulin deficiency for up to 11 weeks post-transfection. Results indicate that transfection on day 1 with TCF3 followed by a co-transfection on day 7 with Pdx1+NeuroG3+Mafa (T1+PNM7) resulted in a lower net increase in fasting blood glucose compared to week one in the STZ-mice compared to sham plasmid. These two studies indicate the potential use of electroporation as a direct cell reprogramming approach to produce IPCs and account for loss of beta-cell mass in T1D patients.
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Keywords
Electroporation, Cell Therapies, Diabetes, Regenerative Medicine, Personalized Medicine