Reddy K V R, Sukanya D, Patgaonkar M S, Selvaakumar C
Division of Molecular Immunology, National Institute for Research in Reproductive Health (NIRRH), Indian Council of Medical Research (ICMR), Jehangir Merwanji Street, Parel, Mumbai 400 012, India.
Int J Pept. 2012;2012:782019. doi: 10.1155/2012/782019. Epub 2012 Mar 14.
Antimicrobial peptides (AMP's) protect epithelial surfaces including epididymis against pathogens and play a key role in orchestrating various defensive responses. Recently, we have identified one such AMP, rabbit epididymal hemoglobin-β subuit (REHbβP) from the epididymal fluid of rabbit, Oryctologus cuniculus. The demonstration of a protective role of REHbβP in epididymal epithelial cells (EPEC's) led us to investigate: (1) the identification of LPS interactive domain in REHbβP, and (2) whether the REHbβP of rabbit origin mediates vaginal cellular immune responses of another species (human). HeLa-S3, human vaginal epithelial cells (hVECs) were exposed to LPS or the LPS-stimulated cells treated with REHbβP or neutral peptide, nREHbβP. Effect of LPS and cytokines (IL-6 and IL-1α) and chemokines (IL-8, MCP-1) levels was determined in the culture supernatants. In response to the LPS, hVECs synthesized these mediators and the levels were significantly higher than controls. This enhancing effect was ameliorated when the LPS-induced hVECs were treated with REHbβP. Similar results were obtained on NF-κB protein and hBD-1 mRNA expression. Confocal microscopy studies revealed that REHbβP attenuated the LPS-induced internalization of E. coli by macrophages. The chemotaxis studies performed using Boyden chamber Transwell assay, which showed elevated migration of U937 cells when the supernatants of LPS-induced hVECs were used, and the effect was inhibited by REHbβP. REHbβP was found to be localized on the acrosome of rabbit spermatozoa, suggesting its role in sperm protection beside sperm function. In conclusion, REHbβP may have the potential to develop as a therapeutic agent for reproductive tract infections (RTI's).
抗菌肽（AMP's）可保护包括附睾在内的上皮表面免受病原体侵害，并在协调各种防御反应中发挥关键作用。最近，我们从穴兔（Oryctologus cuniculus）的附睾液中鉴定出了一种这样的AMP，即兔附睾血红蛋白-β亚基（REHbβP）。REHbβP在附睾上皮细胞（EPEC's）中保护作用的证明促使我们进行以下研究：（1）鉴定REHbβP中与脂多糖（LPS）相互作用的结构域，以及（2）兔源的REHbβP是否介导另一个物种（人类）的阴道细胞免疫反应。将人阴道上皮细胞（hVECs）HeLa-S3暴露于LPS或用REHbβP或中性肽nREHbβP处理LPS刺激的细胞。测定培养上清液中LPS、细胞因子（IL-6和IL-1α）和趋化因子（IL-8、MCP-1）的水平。响应LPS时，hVECs合成这些介质，其水平显著高于对照组。当用REHbβP处理LPS诱导的hVECs时，这种增强作用得到改善。在NF-κB蛋白和人β-防御素-1（hBD-1）mRNA表达上也得到了类似结果。共聚焦显微镜研究表明，REHbβP减弱了LPS诱导的巨噬细胞对大肠杆菌的内化作用。使用Boyden小室Transwell分析法进行的趋化性研究表明，当使用LPS诱导的hVECs的上清液时，U937细胞的迁移增加，而这种作用被REHbβP抑制。发现REHbβP定位于兔精子的顶体上，表明其除了在精子功能方面外，在精子保护中也发挥作用。总之，REHbβP可能有潜力发展成为一种治疗生殖道感染（RTI's）的治疗剂。
