脂多糖诱导的炎症对雄性大鼠精子质量的影响及其可能机制探索

Impact of lipopolysaccharide-induced inflammation on sperm quality in male rats and its possible mechanisms

目的:研究脂多糖(LPS)诱导的炎症对雄性大鼠附睾精子质量的影响,初步探讨其可能的机制。方法:36只雄性SD大鼠(400~450 g)随机分为4组,每组9只。对照组大鼠腹膜内注射无菌生理盐水,余下3组大鼠腹膜内注射LPS(5 mg/kg,溶于无菌生理盐水中),分别于注药12 h(LPS 12 h组)、24 h(LPS 24 h组)和72 h(LPS 72 h组)后麻醉处死。取其附睾检测附睾尾精子活动率及精子数目,并计算精子相对计数值(10~6个/100 mg附睾重);检测附睾组织匀浆中丙二醛(MDA)和超氧化物歧化酶(SOD)活性来反映附睾氧化应激状态;流式细胞术检测睾丸生精细胞凋亡率。结果:(1)与对照组(68.01±1.80)%相比,LPS 12 h组大鼠附睾尾精子活动率[(62.28±4.06)%]明显降低(P﹤0.05),而LPS 24 h组(45.35±3.39)%以及LPS 72 h组附睾尾精子活动率(34.85±4.42)%极显著降低(P﹤0.01);与对照组(38.94±4.08)×10~6个/100 mg相比,LPS 12 h组大鼠附睾尾精子相对计数[(37.15±2.54)×10~6个/100 mg]减少,但无统计学差异(P>0.05),而LPS 24 h组[(31.97±2.81)×10~6个/100 mg]和LPS 72 h组[(28.60±4.03)×10~6个/100 mg]极显著减少(P﹤0.01)。(2)与对照组(4.66±1.49)nmol/mg prot相比,LPS 12 h组[(15.95±3.26)nmol/mg prot]和LPS 24 h组[(12.93±2.54)nmol/mg prot]大鼠附睾MDA含量极显著升高(P﹤0.01),而LPS 72 h组[(9.67±1.68)nmol/mg prot]明显升高(P﹤0.05);与对照组(879.335±105.089)U/mg prot相比,LPS 12 h组大鼠附睾组织匀浆中SOD活性[(729.265±93.783)U/mg prot]降低,但无统计学差异(P>0.05),LPS 24 h组[(694.126±58.530)U/mg prot]和LPS 72 h组[(655.352±115.215)U/mg prot]显著降低(P﹤0.05)。(3)流式细胞术结果显示,与对照组(4.21±1.67)%相比,LPS 12 h组大鼠睾丸生精细胞凋亡率(11.01±3.30)%明显增加(P﹤0.05),而LPS 24 h组[(23.88±4.58)%]和LPS 72 h组[(41.28±2.28)%]极显著增加(P﹤0.01)。结论:LPS(5 mg/kg)腹膜内注射诱导炎症呈时间依赖性损害雄性大鼠生殖细胞与附睾精子质量,可能与其生殖系统氧化应激损伤和凋亡有关。

Objective: To investigate the influence of lipopolysaccharide(LPS)-induced inflammation on sperm quality in male rats and its possible mechanisms. Methods: Thirty-six male SD rats were randomly divided into groups A(control), B(12 h LPS), C(24 h LPS) and D(72 h LPS), the former group injected intraperitoneally with sterile saline, and the latter three with LPS at 5 mg/kg and sacrificed at 12, 24 and 72 hours respectively after treatment. Then the left epididymides of the rats were harvested for detection of the sperm count and motility in the cauda epididymis, measurement of the relative sperm count value, examined the content of malondialdehyde(MDA) and activity of superoxide dismutase(SOD) in the epididymal plasma, and determined the apoptosis rate of spermatogenic cells by flow cytometry(FCM). Results: Compared with the parameters in group A, sperm motility in the cauda epididymis was decreased in groups B([68.01 ± 1.80]% vs [62.28 ± 4.06]%, P < 0.05), C([68.01 ± 1.80]% vs [45.35 ± 3.39]%, P < 0.05) and D([68.01 ± 1.80]% vs [34.85 ± 4.42]%, P < 0.01), and so was the sperm count in the cauda epididymis([38.94 ± 4.08] vs [37.15 ± 2.54] ×10~6/100 mg, P > 0.05;[38.94 ± 4.08] vs [31.97 ± 2.81] ×10~6/100 mg, P < 0.05;[38.94 ± 4.08] vs [28.60 ± 4.03] ×10~6/100 mg, P < 0.01), while the content of MDA in the epididymal plasma significantly increased([4.66 ± 1.49] vs [15.95 ± 3.26] nmol/mg prot, P < 0.01;[4.66 ± 1.49] vs [12.93 ± 2.54] nmol/mg prot, P < 0.01;[4.66 ± 1.49] vs [9.67 ± 1.68] nmol/mg prot, P < 0.05), the activity of SOD reduced([879.335 ± 105.089] vs [729.265 ± 93.783] U/mg prot, P > 0.05;[879.335 ± 105.089] vs [694.126 ± 58.530] U/mg prot, P < 0.05;[879.335 ± 105.089] vs [655.352 ± 115.215] U/mg prot, P < 0.05), and the apoptosis rate of spermatogenic cells dramatically elevated([4.21 ± 1.67]% vs [11.01 ± 3.30]%, P < 0.05;[4.21 ± 1.67]% vs [23.88 ± 4.58]%, P < 0.01;[4.21 ± 1.67]% vs [41.28 ± 2.28]%, P < 0.01). Conclusion: LPS-induced inflammation damages spermatogenic cells and the quality of epididymal sperm in male rats in a time-dependent manner, which is associated with oxidative stress injury and cell apoptosis in the reproductive system.