Analysis of zinc content,zinc-containing enzyme activity and zinc transporter expression during testicular development in rats
Objective To investigate the role and mechanism of zinc homeostasis in testicular development by detecting zinc content and zinc transporter(ZnT)expression at different developmental stages of rat testis.Methods Male SD rats were randomly divided into four groups including4-week age group(n=10),8-week age group(n=10),12-week age group(n=10)and 32-week age group(n=10).Fresh spermatozoa from the tail of epididymis of rats were collected and their number and motility were measured.The histopathological changes of testis were observed by HE staining and microscope.Serum testosterone was determined by chemiluminescence method.The content of zinc in testicular tissue was determined by inductively coupled plasma mass spectrometry.The level of zinc ion in testis was detected by Zinquin probe.Zinc-containing enzyme activity(ALP,ADH,LDH)was analyzed by enzyme-linked immunosorbent assay.mRNA and protein expression of zinc transporters ZnT1-9 in testicular tissue were detected by real-time fluorescence quantitative PCR and Western Blot.Results Spermatozoa appeared in the testis of the 8-week-old rats,and the total number of spermatozoa was increased with the increase of age.Sperm motility and testosterone level of rats in 8-week and 12-week age groups were all significantly higher than those of rats in 4-week and 32-week age groups(P＜0.05).The level of free zinc ion in testis tissue in 8,12 and 32-week age groups was significantly higher than that of 4-week age group.The activity of ALP in testis of 12 and 32 weeks of age was significantly lower than that of 4 and 8 weeks of age(P＜0.05).The levels of ZnT3 mRNA and protein in testis of rats at 8,12 and 32 weeks of age were significantly higher than those of rats at 4 weeks of age(P＜0.05),and the levels of ZnT4 mRNA in testis of rats at 8,12 and 32 weeks of age were significantly higher than those of rats at 4 weeks of age(P＜0.05).The levels of ZnT8 mRNA and protein in testis in 12 and 32 weeks of age groups were significantly higher than those in 4 and 8 weeks of age groups(P＜0.05).Conclusion Zinc-containing enzymeand zinc transporter(ZnT)change dynamically during testicular development in rats,which provides a basis for further investigation on the effect of zinc homeostasis on male reproductive system development.
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