Expression Localization And Histological Analysis Of LYZL4,LYZL6 And PCNA In Yak Testis

The spermatogenesis of mammals is regulated by many factors.In order to ensure the quality of semen,it is necessary to ensure the smooth progress of the spermatogenesis.Studies have reported that LYZL4,LYZL6 and PCNA proteins play an important role in the normal development of male testis and spermatogenesis.However,the research on these genes mainly stays in the model animal mouse and human.nowdays,the research on these genes in yak is still in a vacant state.Therefore,exploring the regulation mechanism of LYZL4,LYZL6 and PCNA in yak testis has great significance for elucidating the molecular regulation mechanism of yak spermatogenesis.In this study,three healthy,6 months old,18 months old,30 months old and 72 months old yak male animals were selected.The expression mechanisms of LYZL4,LYZL6 and PCNA genes in different months old yak testes were studied by fluorescence real-time PCR and Western Blot.The paraffin section were used to reveal the development and changes of spermatogenic tubules and mesenchymal cells in the testis of yak different age.The immunohistochemical method was used to study the immunopositive distribution of LYZL4,LYZL6 and PCNA proteins in the testes of different months yak,so as to understand the regulation mechanism of these three genes in the testis of yak.The main findings are as follows:1.There were significant changes in the number of cells in different months(P<0.01),but only the spermatogonial cells,spermatocytes and interstitial cells had no significant difference in length and length(P>0.05),supporting cells and fines.The long and short diameters of the seminiferous tubes were extremely significant(P<0.01).Through the observation of tissue sections,it was found that as the age of yak increased,the seminiferous tubules became more closely arranged,and the lumen of the seminiferous tubules increased significantly.At the same time,spermatogenic cells at all levels are gradually increasing,reaching the maximum when reaching 72 months of age,and then there is a slowing trend.It is indicated that 72-month-old is the best breeding period for yak,indicating that spermatogenic cells and seminiferous tubules at all levels change regularly with age.2.The expression of LYZL4 gene in 72 months old yak testis was significantly higher than that in 6 months old,18 months old and 30 months old(P<0.01),the expression of genes increased with the growth of the age,high expression at sexual maturity and adult yak.LYZL4 protein mainly locate in round spermatid and mesenchymal cells of yak testicular tissue,and higher expression in the maturation period,protein expression levels at 30 and 72 months old were significantly higher than those at 6 and 18 months old(P < 0.01).These indicate that LYZL4 is involved in the process of spermatogenesis and plays an important regulatory role in the development and maturation of yak testis.3.The expression of LYZL6 was mainly in the testis tissue of adult yak,while the expression of LYZL6 in the testis of pre-mature yak was relatively low.There was a significant difference in the expression of testis between 72 months old testes and 6,18,and 30 months old testes(P<0.01);LYZL6 protein is localized in primary spermatocytes,round spermatid and the apical and medial regions of mature epididymal sperm.The expression of pre-mature protein was significantly lower than that after sexual maturation(P<0.01).It is indicated that LYZL6 regulates spermatogenesis in the testis of yak mainly by affecting theproliferation of spermatocytes.4.The expression of PCNA gene is highly expressed in sexual maturity and testicular tissues of adult yak.The expression levels of pre-mature and sexual maturity are very different,and the expression levels in 30 months old and 72 months old yak testes was significantly higher than 6 and 18 months old(P<0.01).The expression level of PCNA protein in testis after sexual maturity was significantly higher than that before sexual maturity(P<0.01),and the protein expression level at 30 months old was significantly higher than that at 72 months old in yak testis,immunohistochemical detection showed that it was mainly located in primary spermatocytes,secondary spermatocytes,and some supporting cells,and there was no significant difference in the positive signal strength among different age groups(P<0.01).It is suggested that PCNA regulates spermatogenesis by affecting testis mesenchymal cell proliferation and spermatocyte division.