CAJ | 학술논문

目的：建立吸烟小鼠模型，检测小鼠精子特异性钙离子通道蛋白CatSper在RNA和蛋白水平表达的变化，分析与精子活力之间的关系，为吸烟导致雄性生殖损害的分子机制研究提供可靠的实验数据。方法：建立吸烟小鼠模型，运用非连续密度梯度离心分离精子并进行活力检测，RT-PCR 检测小鼠精子mRNA表达的变化，免疫组织细胞化学技术观察小鼠CatSper蛋白的表达，免疫印迹进一步对CatSper蛋白表达的变化趋势进行统计学分析。结果：正常对照组，轻、中和重度吸烟组（a ＋ b）级精子百分率[（87．6±16．2）％vs （72．9±13．0）％vs （68．1±16．0）％vs （44．6±15．0）％，P＜0．05]、CatSper mRNA相对表达量（0．915±0．202 vs 0．712±0．121 vs 0．522±0．126 vs 0．258±0．177，P＜0．05）、免疫印迹相对表达量（0．882±0．208 vs 0．693±0．202 vs 0．471±0．251 vs 0．263±0．198，P＜0．05）均依次降低，免疫组化结果显示吸烟使 CatSper 蛋白在睾丸组织中表达降低；相关性分析结果表明（a＋b）级精子百分率与CatSper mRNA和蛋白呈正相关。结论：吸烟明显抑制小鼠钙离子通道蛋白 CatSper 的表达，精子活力与 CatSper 的表达呈正相关，提示 CatSper表达降低可能是吸烟导致雄性精子活力损伤的分子机制之一。
목적：건립흡연소서모형，검측소서정자특이성개리자통도단백CatSper재RNA화단백수평표체적변화，분석여정자활력지간적관계，위흡연도치웅성생식손해적분자궤제연구제공가고적실험수거。방법：건립흡연소서모형，운용비련속밀도제도리심분리정자병진행활력검측，RT-PCR 검측소서정자mRNA표체적변화，면역조직세포화학기술관찰소서CatSper단백적표체，면역인적진일보대CatSper단백표체적변화추세진행통계학분석。결과：정상대조조，경、중화중도흡연조（a ＋ b）급정자백분솔[（87．6±16．2）％vs （72．9±13．0）％vs （68．1±16．0）％vs （44．6±15．0）％，P＜0．05]、CatSper mRNA상대표체량（0．915±0．202 vs 0．712±0．121 vs 0．522±0．126 vs 0．258±0．177，P＜0．05）、면역인적상대표체량（0．882±0．208 vs 0．693±0．202 vs 0．471±0．251 vs 0．263±0．198，P＜0．05）균의차강저，면역조화결과현시흡연사 CatSper 단백재고환조직중표체강저；상관성분석결과표명（a＋b）급정자백분솔여CatSper mRNA화단백정정상관。결론：흡연명현억제소서개리자통도단백 CatSper 적표체，정자활력여 CatSper 적표체정정상관，제시 CatSper표체강저가능시흡연도치웅성정자활력손상적분자궤제지일。
Objective To provide reliable experimental data for molecular mechanism research on male reproductive impairment by establishing the mice model of smoking to detect the expression of specific cation channel CatSper mRNA and protein in sperm, and analyze its relation with sperm motility. Methods Mice model of smoking was established, sperm samples were separated by Percoll discontinuous density gradients and sperm motility was detected, mice sperm CatSper mRNA was determined by RT-PCR, the distribution of CatSper protein was observed by immunocytochemical technique , CatSper protein level was detected by western blotting. Results In normal control group, mild smoking group, moderate smoking group, and severe smoking group, the proportion of sperm motility (a + b) grade were (87.6 ± 16.2)%vs. (72.9 ± 13.0)%vs. (68.1 ± 16.0)%vs. (44.6 ± 15.0)%(P<0.05), the relative contents of CatSper mRNA were 0.915 ± 0.202 vs. 0.712 ± 0.121 vs. 0.522 ± 0.126 vs. 0.258 ± 0.177 (P<0.05), the relative contents of CatSper protein were 0.882 ± 0.208 vs. 0.693 ± 0.202 vs. 0.471 ± 0.251 vs. 0.263 ± 0.198 (P < 0.05). CatSper protein expression was depressed by smoking in testis testified by immunocytochemical technique, and the results indicated significant negative correlation between CatSper mRNA/protein and the proportion of sperm motility (a+ b) grade. Conclusions The expression of cation channel CatSper was significant inhibited by smoking with a positive correlation with sperm motility , which suggest that the lower expression of CatSper may be involved in the molecular mechanism of male reproductions impairment by smoking.