Inheritable Model of Male Pseudohermaphroditism in Canine Somatic Cell Nuclear Transfer.

Abstract

Normal mammalian sex differentiation depends upon three genetically controlled steps: chromosomal sex determination, gonadal differentiation, and phenotypical sex development. However, two case studies have reported anomalies in the sexual differentiation of mouse and wolf that have occurred during SCNT procedure for unknown reasons. This evokes questions about the early event of reprogramming in SCNT when the chromosomal sex is clearly determined in genetic donor cells, which can never be overcome by technical inflexibilities of sex.During our canine somatic cell nuclear transfer (SCNT) using skin fibroblasts from a male adult as a genetic donor, we report here the birth of one sex reversed cloned dog out of 6 healthy live offspring. Chromosomal, genetic, and histological analyses revealed that the sex-reversed clone has a 78 XY, SRY positive with female external genitalia, which includes an oviduct and a uterus. Interestingly enough, the bilateral testis was in the shape of a hypotrophic ovary and was positioned in the location of normal ovaries. A complete bicornuate uterus was also present. Histological analyses showed that the testis consists of a gonocyte free semiferous tubule-like structure that was stained positive with vimentin, and inhibin alpha, the sertoli cell markers.To study further whether it can be reversible or not, we recloned him, and the result was that the sex reversal condition remained. Thus we believe that we have established the cell line for canine model of inheritable male pseudohermaphroditism and its sex differentiation mechanism. However, we have not observed any opposite sex reversal cases in female genotypes. Further molecular analyses on SRY, SOX-9, MISIIR, AR showed no mutations in their full or partial sequences, which were associated with disorders of sex development in previous reports. In summary, this report describes the first SRY positive XY sex reversal with an almost complete female phenotype including clinical signs; ultrasonographic findings in the abdomen; plasma concentrations of testosterone, oestradiol and progesterone before after administration of GnRH; cytogenetic analysis; PCR analysis for presence of the SRY gene and histology of the genital tract. The results will be discussed with regard to possible causes of the sex reversal. (poster)