五子衍宗丸对少弱精子症模型大鼠精子线粒体膜电位及超微结构影响

目的:研究五子衍宗丸对少弱精子症模型大鼠精子线粒体膜电位(MMP)水平及线粒体超微结构的影响。方法:取体重为200～220 g雄性SD大鼠60只,随机分成正常组,模型组,对照组(黄精赞育胶囊组),五子衍宗丸低、中、高剂量组,除正常组外,其他各组大鼠灌服雷公藤多苷30 mg/(kg.d)],连续8周,制备少弱精子症模型。造模结束后,正常组、模型组给予等容量蒸馏水10 ml/(kg.d)],对照组给予黄精赞育胶囊溶液3.01 g/(kg.d)],五子衍宗丸各组分别给予水提液,低剂量组2.30 g生药/(kg.d)],中剂量组4.60 g生药/(kg.d)],高剂量组9.20 g生药/(kg.d)],连续30 d。末次给药后30 min,采用荧光流式细胞技术(JC-1染色)测定精子MMP水平(JC-1+%),并通过透射电镜观察精子线粒体超微结构的改变。结果:①MMP:JC-1+%和强度分别为:正常组70.80±4.92、4 360±945;模型组33.77±6.19、1 4685±496;对照组56.34±10.35、3 277±895;五子衍宗丸低剂量组40.80±10.40、2 016±767;中剂量组59.40±6.51、3 897±643;高剂量组60.71±7.81、3 371±6467。造模各组大鼠精子线粒体JC-1+%及强度均明显下降,与正常组比较差异有显著性意义(P均<0.05);连续给药30 d后,给药各组均能明显提高精子MMP,增加JC-1+%,除低剂量组外,其他用药各组与模型组比较差异有显著性意义(P均<0.05)。②精子线粒体超微结构:雷公藤造模后,精子外膜松散、变性,线粒体肿胀、大小不一、线粒体膜不完整,轴丝结构不清或出现断裂。给予五子衍宗丸30 d后,精子外膜及线粒体膜结构完整,减少线粒体肿胀,轴丝及微管结构基本正常。结论:雷公藤多苷能降低精子MMP水平,破坏线粒体的结构。五子衍宗丸能明显提高少弱精子症模型大鼠精子MMP水平,减轻精子线粒体结构损伤。保护精子线粒体结构与功能的完整是五子衍宗丸治疗少弱精子症的机制之一。

Wuzi Yanzong Pills increases sperm mitochondrial membrane potential and protects its ultrastructure in oligoasthenotspermia model rats

Objective: To study the effects of Wuzi Yanzong Pills(WYP) on sperm mitochondrial membrane potential(MMP) and its ultrastructure in oligoasthenotspermia model rats.Methods: Oligoasthenotspermia models were made in 50 male rats weighing 200-220 g by intragastric administration of Tripterygium Glucosides at 30 mg per kg per d for 8 weeks,and then equally allocated to a model control,a Huangjing Zanyu Capsule(HZC) control,a low-dose WYP,a medium-dose WYP,and a high-dose WYP group.Another 10 age-matched normal male rats were included as normal controls.The rats in the model and normal control groups were given intragastrically distilled water at 10 ml/kg,those in the HZC group administered HZC at 3.01 g/kg,and those in the low-,medium-and high-dose WYP groups medicated with WYP at 2.30,4.60 and 9.20 g/kg,respectively,once daily for 30 days.At 30 minutes after the last administration,we detected the sperm MMP by JC-1 fluorescent staining and flow cytometry,and examined the sperm ultrastructure under the JEM-1230 transmission electron microscope.Results: JC-1+% and its fluorescence intensity were(33.77±6.19)% and 1 468±496 in the model control,(56.34±10.35)% and 3 277±895 in the HZC control,(40.80±10.40)% and 2 016±767 in the low-dose WYP,(59.40±6.51)% and 3 897±643 in the medium-dose WYP,and(60.71±7.81)% and 3 371±647 in the high-dose WYP group,significantly reduced in comparison with(70.80±4.92)% and 4 360±945 in the normal control group(P<0.05),but remarkably higher in the medium-and high-dose WYP groups than in the model controls(P<0.05).After modeling,the sperm membrane was loose and degenerated,the mitochondria swelling,variously sized and with incomplete membrane,and the axonemal structure unclear or ruptured.After 30 days of WYP administration,compared with the model control group,the rats exhibited integrated sperm membrane and mitochondrial membrane,reduced mitochondrial swelling and basically normal axonemal and microtubular structures.Conclusion: Tripterygium Glucosides could decrease the sperm mitochondrial membrane potential and damage the mitochondrial structure,while WYP could significantly increase the sperm mitochondrial membrane potential and reduce the sperm mitochondrial structure damage.The protection of the integrity of sperm mitochondrial structure and function is one of the mechanisms of WYP acting on oligoasthenotspermia.