Development and Assessment of a P23H Rhodopsin Transgenic Mini-Swine Model of Retinitis Pigmentosa

Abstract

Purpose: : The Pro23His (P23H) rhodopsin mutation underlies the most common form of autosomal dominant Retinitis Pigmentosa (RP) in humans. We used somatic cell nuclear transfer (SCNT) to develop a large animal transgenic model of this type of RP and characterized the functional phenotype of six founders, using the electroretinogram (ERG).

Methods: : Fetal fibroblasts of SLA defined NIH miniature swine (c/c MHC haplotype) were genetically modified by integrating a human genome fragment containing a P23H mutation in the rhodopsin gene along with putative regulatory regions. Multiple cells lines representing different integration sites were utilized for SCNT. A portable mini-Ganzfeld ERG system (HMsERG, RetVetCorp, Columbia, MO), with the standard ISCEV ERG dark- and light-adapted protocols, was used to evaluate the phenotype from six transgenic founders at 4 months of age, and two were re-evaluated at 6 months of age. A clinical examination of the fundus also was performed by indirect opthalmoscopy.

Results: : Six founders were produced and PCR verified that all carried the human DNA fragment. Clinical examinations showed no obvious fundus abnormalities. Compared to age-matched control Yucatan miniature swine, transgenic animals had abnormal dark-adapted and mixed rod/cone ERG responses. The severity of the phenotype varied across the founders. One animal had no dark-adapted ERG response at two months of age, and the mixed rod/cone response as well as the light-adapted response was severely attenuated. Less severe phenotypes were observed in other animals, including one swine that had an abnormal dark-adapted ERG response, but a normal light-adapted response.

Conclusions: : Like other animals in which P23H rhodopsin has been mutated, the transgenic mini-swine exhibits a range of rod/cone dysfunction phenotypes. This represents a new large animal model of human RP and the variation in the severity of the phenotype among the founding lines can be exploited to develop therapeutic approaches to restore vision loss due to photoreceptor degeneration.

Keywords: transgenics/knock-outs • retinal degenerations: cell biology • electroretinography: clinical