Stem Cell-Based Technologies Of Assisted Reproduction In The Cat And Dog
Spermatogonial stem cells (SSC) exist on the basement membrane of the seminiferous tubules in the testis throughout a male animal?s life. These cells self-renew to maintain a stem cell population while producing daughter cells that differentiate into sperm. Because of their relatively undistinguishable morphology, very little was known about them until testis xenografting and spermatogonial stem cell transplantation (SSCT) were developed. Both technologies are promising tools to preserve male genetic information as well as to study the characteristics of SSC. In this dissertation, I investigate testis xenografting and SSCT in cats and dogs, which are important animal models for biomedical research. First, I examine the effect of donor age on spermatogenesis in feline xenografts and the ability of xenograft-derived sperm to support embryo development. In addition, I compare the outcomes of xenografting using testis tissue from two additional species of carnivore, dogs and ferrets. My studies indicate that the optimal donor age was just prior to onset of puberty and xenograft sperm had poor fertility compared to control sperm, necessitating further investigation. Furthermore, the comparative study demonstrates differences in the ability of testicular tissue from different species of carnivore to establish spermatogenesis following testicular xenografting. Second, I describe my development of all three major steps of SSCT in the cat and dog including depleting endogenous male germ cells in recipients, isolating male germ cells from donors and injecting the isolated germ cells. My studies provide a foundation to perform SSCT in the cat, and I was successful at performing SSCT in the dog. Finally, I examine multiple steps to develop transgenesis in dogs, which will enhance their utility as genetic models. This study includes 1) methodology for the accurate staging of dog blastocysts, 2) attempted derivation of canine embryonic stem cells (cESC), 3) manipulation of blastocysts, and 4) transfer of canine embryos. Our results demonstrate that canine transgenesis mediated via cESC and/or SSCT are technically feasible. These studies examine the potential of stem cell-based technologies in the preservation of male genetic information and biomedical research. In addition, my work provides a foundation to achieve transgenesis in the dog.
Cat and Dog Reproduction
dissertation or thesis