The Expression And Localization Analysis Of AURKA In HPGA Of Male Mouse (Mus Musculus) And Its Effect On Androgens Expression

Aurora kinase A(AURKA),one of the protein kinases,regulates mitotic protein kinase in cell cycle.Previous studies showed AURKA may be involved in regulating the development and maturation of germ cells.Studies have shown that AURKA is a target gene of AR and DHT to promote sperm maturationby combining with intracellular AR.This is of great significance for the study of animal reproduction and related diseases,but the specific function of AURKA remains unclear.In the present study,the expression level and localization AURKA in hypothalamus,pituitary and testis in male mice with 8 weeks were analyzed using semi-quantitative interpretation PCR(RT-PCR),Western blotting(WB),hematoxylin-eosin staining(H.E.)and immunohistochemistry(IHC).The designed specific mouse knock down sequence of AURKA and AURKA overexpression vector was transfected into the primary cultured sertoli cells.Immunofluorescence(IF)was used to detect the localized expression of AURKA in adult mice sertoli cells;The expressed variation of AURKA and key enzymes in the androgen synthesis pathway steroidogenic acute regulatory protein(StAR)?cholesterolside-chain cleavage cytochrome P450 enzyme(CYP11A1)? 17?-hydroxylase(CYP17A1)?3?-hydroxysteroid dehydrogenase(3?-HSD)?5?-reductase type 1(SRD5A1)mRNA and proteins was detected using the quantitative real-time PCR(qRT-PCR)and WB.The concentration of T and DHT in sertoli cells treated with or without AURKA-siRNA and pIRES2-EGFP-AURKA was determined using the enzyme-linked immunosorbent assay(ELISA).1.The results showed that AURKA mRNA and protein were expressed in the HPGA tissues of male mice.Compared to the expression of AURKA in hypothalamus and pituitary tissues,the highest expression level of AURKA mRNA and protein presented in testis(P<0.01).The strongest immuno-positive protein of AURKA showed in sertoli cells and sperm;2.The results showed that AURKA is expressed in the cytoplasm and nucleus of adult mice sertoli cells;3.The expression level of AURKA mRNA and protein was down-regulated(P<0.05),whereas the expression level of key enzymes in the androgen synthesis pathway mRNA and protein was significantly up-regulated and the the concentration of T and DHT increased(P<0.05)in adult mice sertoli cells 24 h after the interference with AURKA-siRNA;4.The expression level of AURKA mRNA and protein was up-regulated(P<0.01),whereas the expression level of key enzymes in the androgen synthesis pathway mRNA and protein was significantly down-regulated(P<0.05)and the the concentration of T and DHT decresae(P<0.05)in adult mice sertoli cells 72 h and 96 h after the interference with pIRES2-EGFP-AURKA.Those results suggested the expression of StAR?CYP11A1?CYP17A1?3?-HSD?SRD5A1 and T and DHT synthesis were negatively regulated by AURKA.This data provides some theoretical basis for understanding the function of AURKA in the reproductive processes of male animals.