Toxicological effects of bioactive peptide fractions obtained from Bothrops jararaca snake venom on the structure and function of mouse seminiferous epithelium

Abstract Background: Pathogenesis of Bothrops envenomations is complex and despite numerous studies on the effects of this snake venom on various biological systems, relatively little is known about such effects on the male reproductive system. In the present study, the toxicological outcomes of the low molecular weight fraction (LMWF) of B. jararaca snake venom - containing a range of bioactive peptides - were investigated on the dynamics and structure of the seminiferous epithelium and 15P-1 Sertoli cells viability. Methods: LMWF (5 µg/dose per testis) venom was administered in male Swiss mice by intratesticular (i.t.) injection. Seven days after this procedure, the testes were collected for morphological and morphometric evaluation, distribution of claudin-1 in the seminiferous epithelium by immunohistochemical analyses of testes, and the nitric oxide (NO) levels were evaluated in the total extract of the testis protein. In addition, the toxicological effects of LMWF and crude venom (CV) were analyzed on the 15P-1 Sertoli cell culture. Results: LMWF induced changes in the structure and function of the seminiferous epithelium without altering claudin-1 distribution. LMWF effects were characterized especially by lost cells in the adluminal compartment of epithelium (spermatocytes in pachytene, preleptotene spermatocytes, zygotene spermatocytes, and round spermatid) and different stages of the seminiferous epithelium cycle. LMWF also increased the NO levels in the total extract of the testis protein and was not cytotoxic in concentrations and time tested in the present study. However, CV showed cytotoxicity at 10 μg/mL from 6 to 48 h of treatment. Conclusions: The major finding of the present study was that the LMWF inhibited spermatozoa production; principally in the spermiogenesis stage without altering claudin-1 distribution in the basal compartment. Moreover, NO increased by LMWF induce open of complexes junctions and release the germ cells of the adluminal compartment to the seminiferous tubule.

摘要
背景：Bothrops蛇毒的致病机制复杂，尽管有众多研究探讨该蛇毒对各种生物系统的影响，但对于其对男性生殖系统的影响了解相对有限。在本研究中，我们调查了B. jararaca蛇毒低分子量组分（LMWF）——其中含有多种生物活性肽——对生精上皮的动态结构和15P-1 Sertoli细胞生存能力的毒理学效应。
方法：将LMWF（每个睾丸5 µg/剂量）通过睾丸内（i.t.）注射给予雄性瑞士小鼠。在此操作后七天，收集睾丸进行形态学和形态计量学评估，通过睾丸免疫组化分析评估生精上皮中claudin-1的分布，并评估睾丸蛋白总提取物中的硝酸氧化酶（NO）水平。此外，还分析了LMWF和粗毒（CV）对15P-1 Sertoli细胞培养的毒理学效应。
结果：LMWF在不改变claudin-1分布的情况下，诱导了生精上皮的结构和功能变化。LMWF的作用特点在于上皮的近腔室中细胞丢失（包括处于有丝分裂期的精细胞、前期精细胞、中期精细胞和圆形精原细胞）以及生精上皮周期不同阶段的变化。LMWF还增加了睾丸蛋白总提取物中的NO水平，并在本研究的浓度和时间范围内未表现出细胞毒性。然而，CV在10 μg/mL的浓度下，从6小时到48小时的治疗过程中表现出细胞毒性。
结论：本研究的主要发现是LMWF抑制了精子生成；主要在精子形成阶段，且未改变基底部claudin-1的分布。此外，LMWF引起的NO增加导致复合连接开放并释放近腔室中的生精细胞至生精小管。