Proteomics Analysis Of Rat Caput Epidiymal Early Microenvironment Affected By Heat Stress

Objective To establish the male reproductive system in rat model of heat stress, to provide experimental evidence for epididymal sperm maturation and male contraception research:Proteomics technology to identify epididymis heat stress-related protein expression profiles of heat stress for epididymal microenvironment early impact,provide experimental evidence for the protection of the male reproductive system and a suitable ambient temperature of the male reproductive system.Methods 1.The 47 rats were divided into three parts: Take three of the rat epididymal rat(caput) protein expression profiling; take 12 rats were divided into four groups: normal control group, 42 30 min group, 42 60 min group and 42 120 min groups of three.32 ℃ ℃ ℃male SD rats(300-400g) divided into two groups, heat stress group 16 and control group 16.Heat stress model established by the testes of rats immersed in 42 ℃water bath in a warm bath at different times;The rats in the control group to establish a water bath at room temperature in the control group. 2.The use of sperm motility analyzer for analysis.3.Observing morphology changes of epididymis after HE tissue staining by light microscope after heat stress. 4. Proteomics platform:By two-dimensional gel electrophoresis of heat stress proteins and the control group, the establishment of protein expression profiles, and then use the Image Master 2D Flatinum software to scan and look for protein gel dot upregulated and downregulated. Finally, take the dot decolorized gel eluted protein, the last point with the target by 5800 MALDI-TOF / TOF mass spectrometry identification of differentially expressed proteins, establish differential protein expression profile.Utilization bioinformatics tools(Gene Ontology, PIE, DAVID) analyze the differences and the biological function of proteins involved in the molecular biology processes. 5.Regulated proteins were selected after heat stress proteins and reduced protein key differences, immunohistochemistry and Western blot analysis to verify the expression and proteomics trends are the same. 6. Cell apoptosis was detected epididymis tissue after heat stress apoptosis.Results 1.Sperm motility analysis showed that 42 ℃ water bath time exceeds 60 min sperm motility in rats begins to significantly decrease,so this study in a warm bath 60 min rat epididymis heat stress model; 2.Through the establishment of proteomics expression profiles in rat epididymal head, identified 150 protein; Heat stress-related protein spots were identified 92, the use of MALDI-TOF / TOF mass spectrometry were identified 21 non-redundant protein differences, which increase heat stress-related protein 11, 10 down-regulated protein. 3.HE staining showed epididymis tissue heat stress group relative to the control group, no significant morphological changes, but heat stress group had epididymal epithelium nuclear vacuoles.4. Immunohistochemistry and Western blot verification significantly decreased Prdx6 and Clu expression in heat stress group, Sod1 expression trend in increased expression of heat stress group differences. 5. Tunel detected that heat stress group apoptosis compared with the control group was significantly more obvious.Conclusion Construction of the rat epididymis of acute heat stress model, identified by proteomics heat stress-related protein expression profile provides useful information for the physiological function of the epididymal sperm maturation and research.