Role Of Smads In Spermatogenesis Of Infertile Rats With Kidney-yang Deficiency And Effect Of Wen Yang Sheng Jing Soup

The transforming growth factor-β (TGF-β) superfamily widely exists in the tissues of a wide spectrum of species, participating many biological events such as cell proliferation, differentiation and substance metabolism. Smads are the intracellular signaling molecules of TGF-β superfamily, including R-Smads, Co-Smads and I-Smads. R-Smads are activated by TGF-β receptor complexes, then combine with Co-Smads and enter the nucleus. Nuclear Smads complexes regulate the transcription of target genes through the recognition of different Smads binding proteins to exert the biological effects of TGF-β. The main function of I-Smads is to suppress the signal transduction of TGF-β. At present it is known that TGF-β/Smads take an important regulatory role in spermatogenesis, development, maturation and testicular function, but their roles in dyszoospermia are far from clear. It is thought that the kidney is concerned with the procreation in Chinese traditional medicine. Deficiency of the Kidney yang can influence directly and indirectly thespermatogenetic function of testis, and induce infertility, which is relevant to the disorder of hypothalamus-pituitary gland-testis axis in various components and degrees. The traditional Chinese medicine may adjust the function of this target gland axis and help it to recover the balance. At present, the research on mechanism of infertility induced by Kidney-yang deficiency and treatment with the traditional Chinese medicine mostly concentrate on morphology and function of hypothalamus-pituitary gland-testis axis. The role of Smads in spermatogenesis of infertile rats with Kidney-yang deficiency and effect of the traditional Chinese medicine have not been reported yet.In this study, we examined the expression of Smads, the level of T, FSH and LH, the change of testis morphology and sperm quantity with immunohistochemistry, Western blots, RT-PCR, electron microscope and radioimmunity on the foundation of infertile male rat model induced by adenine, in order to discuss the role of Smads in spermatogenesis of infertile rats with Kidney-yang deficiency and the effect of Wen Yang Sheng Jing soup. 1. The serum level of T, LH and FSH, the sperm quantity in epididymis and the change of testis morphology were examined in the infertile male rat model III group. The contents of T were obviously decreased, LH and FSH were remarkably increased. The sperm quantity in epididymis was reduced. Partial seminiferous tubule became degenerated, distorted, shrinked, collapsed, and the number of sperm was reduced, while the membrane of spermatogonium was breakaged, mitochondria spread out of cell, mitochondria of sperm was vacuolated and swollen obviously, the membrane of Leydig cell bursted and organelles spread in cytoplasm. The rats appeared polyuria, polydipsia, depressed, less eating, body weight loss, color pattern withered, less activityand scrunch, which are typical performance of Kidney-yang deficiency in Chinese traditional medicine. It confirmed that the infertile male rat model induced by adenine is similar to human Kidney-yang deficiency, which is one kind of ideal model of male Kidney-yang deficiency.2. The change of Smads expression was observed. The results of immunohistochemistry showed that Smad 1 and Smad 2 mainly presented in spermatogenic cells while Smad 2 also presented in Leydig cell, and Smad 4 presented in Leydig cell. The results of Western bolts showed that Smad 1, Smad 2 and Smad 4 all presented in rat testis. Similiar with the results of immunohistochemistry, the expression of Smad 1 was decreased in testis, whereas Smad 2 and Smad 4 were increased. This change of expression may directly or indirectly influence spermatogenesis of infertile male rats with Kidney-yang deficiency induced by adenine, which demonstrated that male sterile induced by Kidney-yang deficiency closely correlate with TGF-J5 signaling molecules.3. The effect of Wen Yang Sheng Jing soup on expression of Smads in rats with Kidney-yang deficiency was observed. The body weight and sperm quantity increased remarkably in the group of Wen Yang Sheng Jing soup, the performance of Kidney-yang deficiency alleviated obviously. The content of T increased, FSH and LH decreased. The damage of testis structure was reduced obviously. The results of immunohistochemistry showed that the expression of Smad 1 in rat testis in the group of Wen Yang Sheng Jing soup was higher than that in the group of Kidney-yang deficiency, whereas expression of Smad 2 and Smad 4 were reduced. Semi-quantitive RT-PCR results showed that expression of Smad 1 mRNA was decreased in the group...