Proteomic analysis of effect of hyperthermia on spermatogenesis in adult male mice.

We characterized cellular and molecular mechanisms involved in spermatogenesis following short-term heat exposure of murine testis. For these studies, we utilized a proteomic approach with two-dimensional gel electrophoresis (2DE) analyses and mass spectroscopic identification of proteins with altered expression in mouse testes at different times after heat shock. We established a proteome reference map from 7-wk-old mouse testis linked to a federated proteome database. We used these tools to analyze quantitative variations in the tissue over a time course of 0.5, 2, 6, and 12 h following heat exposure. We separated 108 protein spots expressed differentially between the heat shock tissues and the control mouse testes. Of these spots, we identified 36 by comparing with the control reference map. We then focused on the heterogeneous nuclear ribonucleoproteins (hnRNPs) and the chaperonins containing t-complex polypeptide-1 (CCT). Further analysis in this heat-shocked model suggests numerous potential mechanisms for heat shock-induced spermatogenic disorder.

[Cloning and expression of human IL-1R II gene].

AIM: To clone human IL-1R II cDNA and construct its recombinant retrovirus vector so as to explore its role in IL-1R II related diseases. METHODS: Human IL-1R II cDNA was amplified by RT-PCR from peripheral blood mononuclear cells (PBMCs) and inserted into the vector PET22b to construct recombinant vector PET22b-IL-R II. The recombinant was transfected into E. coli BL21 and expressed under IPTG induction. Expressed products were detected by Western blot. In addition, human IL-1R-II cDNA was subcloned into retrovirus vector LZRSPBMN and transfected into 293 cells by calcium phosphate precipitation. IL-1R II expression was detected by immunohistochemical staining. RESULTS: IL-1R II cDNA with 1,203 bp was amplified by RT-PCR from human PBMCs. The recombinant of this cDNA could be expressed in E. coli,which was confirmed by Western blot results. Immunohistochemistry detection showed IL-1R II protein was expressed in 293 cells. CONCLUSION: Human IL-1R II gene was cloned successfully. PET22b-IL-1R II and LZR-IL-1R II were constructed and the recombinant protein IL-1R II was expressed in E.coli BL21. The results reported herein lay the foundation for further research on the role of IL-1R II in certain diseases.