Establishment Of The Light-induced Retinal Degeneration Model In Miniature Porcine And Evaluate The Model In Vivo Study After Cell Transplantation

Objective：To establish light-induced retinal degeneration model in Guangxi Bama miniature pigsand to determine the changes of histology and function. In addition，to discuss the properprotocol of sub-retinal space injection of transplanting human embryonic stem cell(hESC)derived retinal pigment epithelium(RPE) cells on light-induced retinal degenerationminiature porcine model and to evaluate the safety, the changes of retinal structure andfunction afterward，providing laboratory evidence to evaluate the possibility of stem celltherapy in retinal degeneration diseases.Methods：1. Establishment and identification of the light-induced retinal degeneration model inGuangxi Bama miniature pigs.Guangxi Bama miniature pigs were exposed to white lamp continuously more thanthree months and the retinas were determined by optical coherence tomography (OCT)，full-field electroretinogram (fERG)，multifocal electroretinogram (mfERG)，as compared toHE staining and electron microscopy(ECM)in vitro study.2. Sub-retinal space transplantation of hESC derived RPE cells in porcine retinaldegeneration model and the changes of retinal structure and function were examined byophthalmoscope，B ultrasound, OCT，fERG and mfERG.Results：1. Post-light exposure, OCT examination showed the thickness of retinal ILM-RPElayer and ONL layer became significantly thinner；amplitudes of dark-adapted combinedrod–cone response’s a waves、b waves，OPS waves and light-adapted b waves, p1waves ofthe fERG significantly decreased；average amplitude density of p1waves of the mfERGsignificantly lower；outer nuclear layer was significantly thinner in retinal HE staining. Transmission electron microscope showed the mitochondria of retinal photoreceptor cellsafter light exposure were swelling， the cristae were disrupted， the vacuolar weredegenerated and the structure of retinal pigment epithelial cell microvilli was damaged. Invivo results were consistent with in vitro study.2. The central and posterior vitreous were removed by using a localized three-port parsplana vitrectomy approach and the hESC derived RPE cells were transplanted by sub-retinalspace injection. B ultrasound result showed cataract has been observed in two weeks on onepig in batch one, after improving the technology and surgical equipment, no complicationsappeared on three pigs in batch two；OCT result showed the retinal pigment epithelial cellscould survive after sub-retinal space transplantation and integrate with the recipient porcineretina. Moreover, there are no significant changes in retinal function according to fERG andmfERG examination.Conclusion：1. In vivo study showed continuous light exposure could change the histology andfunction of porcine retina，it could be used to identify the large animal retinal degenerationmodel.2. The study also proved the sub-retinal space injection technique was suitable forhESC derived RPE cells transplantation.3. In vivo study also showed hESC derived RPE cells could survive aftertransplantation and integrate with the host retina up to1month, retinal function has nosignificant changes after light exposure.