輔仁大學 醫學系1, 生物醫學暨藥學研究所2
The apoptosis pathway involved in the spermatogenesis of CFTR-deficient mice
Guan-Ru Chen1, Han-Sun Chiang2, Yi-No Wu1
School of Medicine1, Graduate Institute of Biomedical and Pharmaceutical Science2, Fu Jen Catholic University, New Taipei City, Taiwan
Purpose: Cystic fibrosis transmembrane conductance regulator (CFTR) is an apical membrane chloride channel and responsible for anion secretion in the lungs, pancreas and male reproductive tract. Loss of CFTR activity causes the dehydration of the apical membrane and male infertility. However, the underlying mechanisms remain elusive. The aim of this study is to determine the role of apoptosis pathway in CFTR deletion during spermatogenesis.
Materials and Methods: CF mice (Cftrtm1Unc) carried the S486X mutation were obtained by Jackson Laboratory. Breeding of Cftr heterozygous mice yielded offspring of all three genotypes including Cftr-/-, Cftr+/-, and Cftr+/+ male mice. We calculated the genotype ratios and number of offspring of male Cftr+/− and female Cftr+/+ breeders. Mice survived beyond 6 weeks and 8 weeks of age were used in this experiment. Then we calculated the sperm count and measured the weight of their reproductive organs and body weight. The reproductive tract tissue were determined the apoptosis marker (caspase-3, caspase-7, and caspase-9) expression by immunofluorescence staining and western blot.
Results: We found that Cftr-/- male mice owns a lower breeding rate and have less offspring in comparison with Cftr+/+ male mice. We also found that the testis and epididymis of 6-week-old Cftr-/- mice is significantly lighter than the Cftr+/+ ones. However, these organs of 8-week-old Cftr-/- mice have the same weight as the 8-week-old Cftr+/+ control group mice. Moreover, the expression of cleaved caspase-3 is elevated in testis germ cells of Cftr-/- mice, with a stage-specific pattern. In Cftr-/- mice, caspase-3 is excessively expressed in stage V-VIII spermatocytes and spermatids. Besides the effector caspase, caspase-3, we found that the expression of caspase-9 and caspase-7 is also altered by the depletion of CFTR.
Conclusions: In conclusion, we found the correlation between the depletion of CFTR and the pathway of apoptosis. We expect these results to give us a better knowing on the reasons why non-CBAVD with CFTR mutation patients are infertile, and possibly give us the vision to seek new treatments for infertile patients.