A New Immunodeficient Pigmented Retinal Degenerate Rat Strain To Study Transplantation of Human Cells

Abstract

Purpose: To develop an immunodeficient rat model of retinal degeneration (RD) that will not reject transplanted human cells (“RD nude rats”).

Methods: Female homozygous S334ter line 3 rats (Tg S334ter3Lav; mutant human S334ter opsin transgene) were crossed with male pigmented NIH nude rats (NTac:NIH-Whn; mutation in the Foxn1 gene; no T-cells). The offspring after several crosses and back-crosses were homozygous for Foxn1 and homo- or heterozygous for S334ter (“RD nude rats”). Immunodeficiency was tested by transplanting sheets of hESC-derived neural progenitor cells to the subretinal space of 12 RD nude rats (39-45d old), and, as a control, 8 homozygous standard nu/nu rats (43-51d old). Rats were housed in sterile microisolator cages, sacrificed between 8d and 6 months after surgery, and eye sections analyzed by immunohistochemistry for human, neuronal and glial markers. The new strain was donated to the Rat Research Resource Center (RRRC) and re-derived to a pathogen-free status for distribution to interested investigators (RRRC# 539 SD- Foxn1 Tg(S334ter)3LavRrrc, available at www.rrrc.us.).

Results: Homozygous nude females are poor breeders. Therefore, heterozygous Foxn1, Tg S334ter positive females need to be bred with homozygous Foxn1, Tg S334ter positive males. This provides homozygous nude Tg S334ter positive rats suitable for study. Genotyping assays for both the Foxn1 mutation and the S334ter transgene were developed for accurate genotyping of animals. Foxn1 homozygous rats are hairless whereas Foxn1 heterozygous rats have hair. After transplantation to RD and standard nude rats, hESC-derived neural progenitor cells differentiated to neuronal and glial cells, and migrated extensively from the transplant sheets all over the host retina. Migration was much more extensive in RD than in standard nude rats. Already 8d after transplantation, donor neuronal processes were found in the host inner plexiform layer. In addition, host glial cells extended processes into the transplants. Transplants were observed up to 6 months. The host retina showed the same photoreceptor degeneration pattern as in the immunocompetent S334ter-3 rats. Recipient survival after surgery was very good, with only one rat dying prematurely.

Conclusions: This newly created rat model is extremely useful to test the effect of human cell transplantation on restoration of vision without interference of immunosuppression.

Keywords: 696 retinal degenerations: hereditary • 741 transplantation • 740 transgenics/knock-outs