Progressive accumulation of n-9 PUFAs in testicular lipids during ex vivo tissue maintenance

Abstract

Spermatogenesis has been achieved in vitro using a gas-liquid interphase culture system. In this setting, it is possible to follow lipid metabolism to know its role during the spermatogenic process, thus gathering potentially useful information for ex vivo spermatogenesis biotechnology. We observed a relationship between the progression of spermatogenesis in both, in vivo and ex vivo, at cytological and histological level and the gene expression of enzymes involved in fatty acid desaturation, elongation and transport. The aim of this study was to examine and extend whether developmental changes that occur in vivo in testicular lipids with long-chain (C18- C22) and very long-chain (≥ C24) polyunsaturated fatty acids (PUFA) also occur ex vivo in explants of neonatal testes in culture. Explants from 6-day old mice cultured for 22 days showed that differentiation proceed from spermatogonial stem cells up to haploid round spermatids. Notably, after 44 days in culture we were able to detect some spermatozoa, although still scarce. Interestingly, like in vivo, total lipids from explants increased their proportion of PUFA during the period in culture. In addition to the common n-6 and n-3 PUFAs (22:5n-6, 22:6n-3, 24:4 and 24:5) we observed that unusual n9 PUFAs (20:3n-9 and 22:4n-9) were accumulated in the explants. The 22:5n-6/20:4n-6 ratio in the glycerophospholipids was increased during the first 22 days associated to the appearance of haploid spermatids and then it remains unchanged until day 44. In addition, we noted that in culture the testicular tissue accumulated neutral lipids, mainly, triacylglycerides (TAG) and cholesterol esters (CE) that contained a high proportion of n-9 PUFAs. In vivo, the increase of neutral lipids occurs in testicular tissue associated with spermatogenesis impairment. The presence of the unusual fatty acids of the n-9 serie in the explants suggests that the culture system does not ensure the provision of essential fatty acids to the tissue. Addressing this point could improve the rate of gamete production in this system