Current scenario and challenges ahead in application of spermatogonial stem cell technology in livestock.

PURPOSE: Spermatogonial stem cells (SSCs) are the source for the mature male gamete. SSC technology in humans is mainly focusing on preserving fertility in cancer patients. Whereas in livestock, it is used for mining the factors associated with male fertility. The review discusses the present status of SSC biology, methodologies developed for in vitro culture, and challenges ahead in establishing SSC technology for the propagation of superior germplasm with special reference to livestock. METHOD: Published literatures from PubMed and Google Scholar on topics of SSCs isolation, purification, characterization, short and long-term culture of SSCs, stemness maintenance, epigenetic modifications of SSCs, growth factors, and SSC cryopreservation and transplantation were used for the study. RESULT: The fine-tuning of SSC isolation and culture conditions with special reference to feeder cells, growth factors, and additives need to be refined for livestock. An insight into the molecular mechanisms involved in maintaining stemness and proliferation of SSCs could facilitate the dissemination of superior germplasm through transplantation and transgenesis. The epigenetic influence on the composition and expression of the biomolecules during in vitro differentiation of cultured cells is essential for sustaining fertility. The development of surrogate males through gene-editing will be historic achievement for the foothold of the SSCs technology. CONCLUSION: Detailed studies on the species-specific factors regulating the stemness and differentiation of the SSCs are required for the development of a long-term culture system and in vitro spermatogenesis in livestock. Epigenetic changes in the SSCs during in vitro culture have to be elucidated for the successful application of SSCs for improving the productivity of the animals.

Supraphysiological concentration of urea affects the functional competence of Holstein-Friesian (Bos taurus) sperm.

To understand the effects of urea on sperm functional attributes, fresh bull semen (n = 12) was subjected to four different concentrations (mg/mL) of urea to mimic the physiological (0.04 and 0.13), supraphysiological (0.43) concentrations and control (0 mg/mL). Sperm membrane integrity, kinematics, chromatin integrity, and mitochondrial membrane potential were assessed at different time points (before incubation, 0, 1, 2, and 4 h) of incubation. The concentration of urea in serum and seminal plasma was estimated and correlated with the ejaculate rejection rate and sperm functional attributes. The relative expression of urea transporter gene transcripts (UT-A and UT-B) was assessed in sperm and testis (control) using real-time PCR. The supraphysiological concentration of urea affected sperm kinematics, viability, functional membrane integrity, and acrosome integrity within 1 h of incubation (p < 0.05). Sperm head area decreased (p < 0.05) at 0 h and subsequently increased at 1 h of incubation in all media except supraphysiological (0.43 mg/dL) concentration of urea. Seminal plasma urea concentration showed a significant negative correlation with sperm motility, membrane integrity, and mitochondrial membrane potential (p < 0.05), but had a positive correlation with the ejaculate rejection rate (r = 0.69). Relative expression of the urea transporter genes revealed that UT-A was expressed only in the testis. In contrast, UT-B was expressed in both the testis and sperm, suggesting UT-B's role in regulating urea transport in sperm. At a supraphysiological level, urea adversely affected sperm functional attributes, osmoadaptation and may affect fertility.