采用优化家兔阴道刺激模型评价乙酸阴道热敏凝胶剂的阴道免疫炎症毒性研究

Study on Using the Optimized Vaginal Irritation Model of the Rabbit to Evaluate the Vaginal Mucosal Immuno-inflammatory Toxicity of the Acetic Acid Vaginal Thermosensitive in Situ Gel

目的采用优化家兔阴道刺激模型评价乙酸阴道热敏凝胶剂(acetic acid vaginal thermosensitive in situ gel,Acid-TISG)对家兔阴道黏膜的免疫炎症毒性。方法将24只♀家兔随机分为空白凝胶组、Acid-TISG组(12.11 mg·m L^(-1))、壬苯醇醚(nonoxynol-9,N-9)凝胶组(5.6 mg·m L^(-1)),阴道给予3 m L凝胶,每天1次,连续10 d,给药前和给药后24 h收集家兔阴道灌洗液(cervicovaginal lavages,CVLs),检测CVL中白介素6(interleukin-6,IL-6)和肿瘤坏死因子α(tumor necrosis factorα,TNF-α)的含量。家兔处死后取阴道组织做病理学检查,HE染色观察家兔阴道黏膜组织损伤并进行评分。运用免疫组化技术检测阴道黏膜内白细胞共同抗原-CD45RB和核增殖抗原(proliferating cell nuclear antigen,PCNA)的表达。结果与空白凝胶组相比,Acid-TISG组黏膜上皮完整,无明显炎症浸润、充血和水肿,N-9凝胶组黏膜上皮出现脱落,固有层炎性细胞明显增多(P<0.05);空白凝胶组和Acid-TISG组阴道黏膜无明显CD45RB阳性表达,N-9凝胶组阴道黏膜固有层出现较多的CD45RB阳性表达细胞。与空白凝胶组相比,Acid-TISG组的阴道黏膜PCNA表达增多,但无显著性差异,N-9凝胶组的阴道黏膜PCNA阳性表达细胞明显增多(P<0.05)。与给药前相比,给药后24 h时CVL中IL-6和TNF-α含量在空白凝胶组和Acid-TISG组无明显改变,N-9凝胶组明显升高(P<0.05)。结论 Acid-TISG对家兔阴道黏膜无明显免疫炎症毒性,优化的家兔阴道刺激模型可用来评价生殖道外用药物对阴道黏膜的免疫炎症毒性。

OBJECTIVE To evaluate the immuno-inflammatory toxicity of vaginal mucosa of acetic acid vaginal thermosensitive in situ gel（Acid-TISG） by using the optimized vaginal irritation model in the rabbits. METHODS Twenty-four rabbits were divided randomly into the blank gel group, Acid-TISG group（12.11 mg·m L-1） and positive nonoxynol-9（N-9） group（5.6 mg·m L-1）. Gel was administered through vagina to rabbits at 3 m L per day for 10 d. Cervicovaginal lavages（CVLs） were collected at pre-administration and 24 h after the last administration. The concentrations of interleukin-6（IL-6） and tumor necrosis factor-α（TNF-α） in the CVLs were detected. The pathological examination of the vaginal tissues was conducted, and the vaginal irritation was assessed and scored. The expression of leukocyte common antigen-CD45 RB and proliferating cell nuclear antigen（PCNA） in the vaginal mucosa was detected by the immuno-histochemistry（IHC） assay. RESULTS The vaginal mucosal epithelium in the rabbits of the Acid-TISG group was intact with no obvious inflammation infiltration, congestion and edema; while the exfoliation of mucosal epithelium cells and the increase of the inflammatory cells were obviously observed in N-9 gel group（P〈0.05）. There was no obvious positive expression of CD45 RB cells both in the blank gel group and the Acid-TISG group, but the positive expression of CD45 RB cells increased significantly in the N-9 gel group（P〈0.05）. The concentrations of IL-6 and TNF-α in CVLs, were significantly increased in the N-9 gel group（P〈0.05） 24 h after administration while there were no significantly increase in both blank gel group and the Acid-TISG group compared with predose. CONCLUSION Acid-TISG is unlikely to cause vaginal mucosal irritation in the rabbits. The optimized rabbit vaginal irritation model can be used to evaluate the topical toxicity of the candidate vaginal drugs.