Cell Reprogramming And Transplantation For Treatment Of Optic Nerve Injury

Regenerative medicine aims to create healthy and functional cells of the same type to replace diseased or damaged tissues. Two major problems need to be solved urgently are the shortage of functional cells available for transplantation and the risk of immune rejection. Successful reprogramming of adult fibroblasts into induced pluripotent stem cells (iPSCs) using defined transcription factors gives a potentially approach to generate functional cells from the patient himself. Rather than switching the fate of differentiated cells going backward and then going forward, there might be a shortcut directly converting one adult cell type into another without going through the pluripotent state. Recent pioneering works show that this method is feasible. Cells generated via direct reprogramming of somatic cells are probably not tumorigenic, and give new possibilities for disease modeling and cellular repair.Here, we show that a combination of Mash1, Math5 and RAX was capable of converting human fibroblasts from the scalp tissue of patients with traumatic brain injury (TBI) into functional hiRGC cells. Twenty days after delievering Mash1, Math5 and RAX, the conversion rate was 12.1%. Electrophysiology results showed that hiRGC cells had normal neuron function. Besides, hiRGC cells maintained normal karyotype during the conversion. Their p53 expression remained low and stable, which showed that hiRGC was not tumorigenic.Then, fluorescence-activated cell sorting (FACS) was performed using PSA-NCAM, a marker for immature neurons or neuronal progenitors. The percentage of GFP+/PSA-NCAM+ cells was 5.41%, containing more than 85% GFP+ population. Patch-clamp recordings demonstrated that, these cells expressed voltage-gated ion channels. The spontaneous action potentials generated in these cells was also discovered. We transplanted human GFP+ iRGC cells into the retina of nude mice. We also demonstrate that hiRGC cells were able to survive, migrate into the ratina after been grafted.To our knowledge, this is the first time human fibroblast was converted directly into human retinal ganglion cell and transplanted into the retina tissue of nude mice. Moreover, we have confirmed the safety of human iRGC cells in vivo by observing 5 months, neither teratoma nor neural tumor emerged.