目的 构建慢性骨盆疼痛综合征前列腺炎(chronic prostatitis /chronic pelvic pain syndrome,CP/CPPS)C57BL/6小鼠模型,探索其机械痛阈和自噬相关微管轻链蛋白LC3和底物蛋白p62表达水平随造模时间的变化规律,为CP/CPPS疼痛及自噬水平研究提供动物实验依据.方法 将36只雄性C57BL/6小鼠随机均分为空白组、对照组和模型组,模型组小鼠皮下多点注射大鼠前列腺蛋白提取液和完全弗式佐剂的混悬液,建立CP/CPPS小鼠模型.通过HE染色观察前列腺病理变化,运用Von Frey纤维丝测定骨盆区域机械痛阈,通过免疫组化染色检测LC3和p62表达水平,运用Image Pro Plus 6.0软件计算平均光密度.结果 HE染色可见模型组小鼠出现慢性前列腺炎,表现为不同程度的上皮增生和淋巴细胞侵润,且实验后第6个月前列腺出现上皮内瘤(prostatic intraepithelial neoplasia,PIN),表现为基底膜消失和细胞核异形性明显等,空白组和对照组则表现为正常组织学形态.与空白组及对照组相比,模型组机械痛阈随造模时间延长逐渐降低[初始痛阈值为(0.35±0.154)g,第22周为(0.008±0.000)g],差异有显著性(P〈0.05).其LC3和p62表达水平逐渐增高[LC3,p62平均光密度值分别为,第1个月:(2.767±0.464)%,(2.872±1.642)%;第6个月:(13.501±1.900)%,(9.070±0.490)%],差异有显著性(P〈0.05).结论 成功建立了CP/CPPS模型,且造模后第6个月出现PIN.模型组小鼠机械痛阈随造模时间的延长逐渐降低,LC3和p62表达逐渐增高,表明CP/CPPS炎症微环境促进疼痛产生及加剧,并提高小鼠前列腺自噬水平,与PIN的发生发展密切相关.
Objective To observe the changes of mechanical pain thresholds and autophagy related proteins microtubule-associated protein 1 light chain 3 (LC3) and sequestosome 1 (SQSTM1 also known as p62) expression levels in the C57BL/6 mouse models of chronic prostatitis/ chronic pelvic pain syndrome (CP/CPPS),and provide animal experimental evidence for CP/CPPS pain and autophagy study.Methods 36 male C57BL/6 mice were randomly divided into three groups: the model group,control group and na(i)ve group.The CP/CPPS model was established by subcutaneous injection in the lower abdomen region with suspension liquid,containing protein extract of male SD rat prostate gland and complete Freund adjuvant.At 1month and 6 months after modeling,the mice were sacrificed and prostate tissues were harvested for histological examination using HE staining.Mechanical tactile hyperalgesia was measured with von Frey filaments.The autophagy-related proteins LC3 and p62 expression levels were detected by immunohistochemistry,respectively.The average IOD was measured by Image Pro Plus 6.0,and the statistical analysis was performed with GraphPad Prism 5 software.Results The histopathology showed the appearance of chronic prostatitis in the model group,representing hyperplasia and lymphocytic infiltration to a different degree and lasted for 6 months after modeling.Moreover,prostate intraepithelial neoplasia (PIN) appeared in the model group at 6 months after modeling,characterized by the disappearence of basement membrane and obvious nuclear abnormality,while the control and na(i)ve groups showed normal histology during the 1-6 months.Compared with the control and na(i)ve groups,the mechanical pain threshold in the model group was significantly decreased along with the time from (0.353±0.154) g at 0 week to (0.008±0.00) g at 22 weeks (P〈0.05).The average IOD of LC3 and p62 expression in the model group was significantly increased with timing from [(2.767±0.464)%,(2.872±1.642)%] at 1month to