活血补肾法调控慢性盆腔炎模型大鼠抗炎抗氧化及细胞凋亡相关机制的实验研究
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摘要
目的:通过观察慢性盆腔炎模型大鼠血清、子宫组织中炎症细胞因子、抗氧化相关指标及细胞凋亡的变化,探讨活血补肾法治疗慢性盆腔炎(CPID)的抗炎、抗氧化及细胞凋亡机制。
方法:以Wistar雌性大鼠为实验对象,采用混合细菌加机械损伤方法,复制慢性盆腔炎动物模型。将56只大鼠随机分为正常组、假手术组、模型组、盆腔炎颗粒大、中、小剂量组及盆炎净颗粒组,酶联免疫吸附测定法(ELISA)检测各组大鼠血清的炎症细胞因子水平,包括白介素-6(IL-6)、白介素-4(IL-4)及黄嘌呤氧化酶法测定超氧化物歧化酶(SOD)、硫代巴比妥酸法测定丙二醛(MDA);大鼠子宫形态学观察后,免疫组织化学法S-P法检测各组大鼠子宫组织Caspase-3的表达;逆转录-聚合酶链反应法(RT-PCR)检测大鼠子宫组织Fas/FasLmRNA和p53mRNA的表达。
结果:慢性盆腔炎模型大鼠血清前炎症细胞因子IL-6水平较正常组大鼠升高,抗炎症细胞因子IL-4水平较正常组大鼠降低,血清中抗氧化相关指标SOD较正常对照组大鼠降低,MDA较正常对照组升高,慢性盆腔炎模型大鼠子宫组织中Caspase-3、Fas/FasLmRNA和p53mRNA的表达较正常组大鼠升高,差异均有极显著性(P<0.01)。药物干预后,各治疗组大鼠血清前炎症细胞因子IL-6水平较模型组下降,抗炎症细胞因子IL-4水平则均较模型组升高;血清中SOD的含量较模型组显著升高(P<0.01),血清中MDA含量较模型组显著降低(P<0.01),盆腔炎颗粒各组血清中SOD及MDA浓度相关系数为﹣0.798,呈显著负相关性(P<0.01)。各治疗组大鼠子宫组织中Caspase-3、Fas/FasLmRNA和p53mRNA的表达与模型组相比较均增强(P<0.05)。其中,盆腔炎颗粒大剂量组IL-6、IL-4及SOD和MDA水平均已接近正常对照组水平(P>0.05)。除IL-6和Caspase-3外,盆腔炎颗粒大剂量组的作用均明显优于盆炎净颗粒组(P<0.01)。除IL-4外,盆腔炎颗粒中、大剂量组效果优于盆腔炎颗粒小剂量组(P<0.05)。盆腔炎颗粒各剂量组子宫组织FasmRNA及FasLmRNA表达相关系数为0.894,呈显著正相关性(P<0.01)。
结论:活血补肾法通过对慢性盆腔炎模型大鼠血清炎症细胞因子、抗氧化相关指标的调节,使机体恢复免疫平衡,有效抑制慢性盆腔炎模型大鼠炎症,加快清除自由基,减少脂质类物质的堆积,从而减轻盆腔内组织器官的慢性损伤。活血补肾法通过增强慢性盆腔炎模型大鼠子宫组织中相关凋亡指标的表达,诱导炎细胞凋亡,抑制炎细胞生长,从而减轻炎症反应。
Objective : To explore the immunoregulation mechanism of the treatment of promoting blood circulation and tonifying the kidney on the rats with chronic pelvic inflammatory disease(CPID) by observing the changes of inflammatory cytokines, antioxygen and control of cell apoptosis.
Methods :Female Wistar rats were used and the CPID model was established with mixed bacteria injection combined with mechanic injury. Fifty-six rats were randomly divided into normal group, sham operation group, model group, Penqiangyan granule large , medium and small dose groups and Penyanjing granule group. The histomorphological changes of the rats' uteri were observed using light microscope. The enzyme-linked immunosorbent assay (ELISA) was applied to measure the concentration of pro-inflammatory cytokines (IL-6) and anti-inflammatory cytokines (IL-4)in serum of rats in each group; The method of xanthinoxidase were applied to measure concentration of SOD.The method of thio-malonylurea were applied to measure the concentration of MDA.The immunochemistry method was used to detect the expression of Caspase-3 in rats’uterine tissue.The expression of Fas/FasLmRNA and p53mRNA were measured by using RT-PCR
Results:The serous levels of pro-inflammatory cytokines (IL-6) in model group were significantly higher than those in normal group(P<0.01); the serous levels of anti-inflammatory cytokines(IL-4) in model group were significantly lower than those in normal group. The serous levels of MDA in model group were significantly higher than those in normal group(P<0.01);the serous levels of SOD in model group were significantly lower than those in normal group(P<0.01).The uterine tissure expression of Caspase-3, Fas/FasLmRNA and p53mRNA were all significantly increased than those in normal group.After treatment ,the serous levels of pro-inflammatory cytokine(IL-6)in all treatment group were significantly lower than those in model group,meanwhile the anti-inflammatory cytokin(eIL-4)in all treatment group were elevated .The serous levels of MDA in all treatment groupwere significantly decreased than those in model group(P<0.01);the serous levels of SOD in all treatment group were significantly increased than those in model group(P<0.01).The coefficient correlation of the serous levels of SOD and MDA in all Penqiangyan granule groups was 0.798,showing the significant negative correlation(P<0.01). The uterine tissure expression of Caspase-3, Fas/FasLmRNA as well as p53mRNA in all treatment group were significantly increased compared with model group(P<0.01). IL-6、IL-4、SOD and MDA in Penqiangyan granule large dose group were similar to the levels of normal group(P>0.05).Except for IL-6 and Caspase-3 Penqiangyan granule large dose group significantly outweighed the Penyanjing granule group(P<0.01). Except for IL-4 Penqiangyan granule large、medium dose groups significantly outweighed the Penqiangyan granule small dose group(P<0.05).The coefficient correlation of the uterine tissure expression of FasmRNA and FasLmRNA in all Penqiangyan granule groups was 0.894,showing the significant positive correlation(P<0.01).
Conclusions: The method of promoting blood circulation and tonifying the kidney could adjust the levels of the inflammatory cytokines and the corresponding antioxygen indices,thus restore the immune balance effectively so as to inhibit inflammation,speeding up eliminating free radicles,decrease the lipid,so as to relieve the chronic injury of pelvic tissures and organs.The method of promoting blood circulation and tonifying the kidney could strengthen and induce the expression of cell apoptosis,inhibit the growth of inflammatory corpuscles,thus ease the inflammatory reaction.
方法:以Wistar雌性大鼠为实验对象,采用混合细菌加机械损伤方法,复制慢性盆腔炎动物模型。将56只大鼠随机分为正常组、假手术组、模型组、盆腔炎颗粒大、中、小剂量组及盆炎净颗粒组,酶联免疫吸附测定法(ELISA)检测各组大鼠血清的炎症细胞因子水平,包括白介素-6(IL-6)、白介素-4(IL-4)及黄嘌呤氧化酶法测定超氧化物歧化酶(SOD)、硫代巴比妥酸法测定丙二醛(MDA);大鼠子宫形态学观察后,免疫组织化学法S-P法检测各组大鼠子宫组织Caspase-3的表达;逆转录-聚合酶链反应法(RT-PCR)检测大鼠子宫组织Fas/FasLmRNA和p53mRNA的表达。
结果:慢性盆腔炎模型大鼠血清前炎症细胞因子IL-6水平较正常组大鼠升高,抗炎症细胞因子IL-4水平较正常组大鼠降低,血清中抗氧化相关指标SOD较正常对照组大鼠降低,MDA较正常对照组升高,慢性盆腔炎模型大鼠子宫组织中Caspase-3、Fas/FasLmRNA和p53mRNA的表达较正常组大鼠升高,差异均有极显著性(P<0.01)。药物干预后,各治疗组大鼠血清前炎症细胞因子IL-6水平较模型组下降,抗炎症细胞因子IL-4水平则均较模型组升高;血清中SOD的含量较模型组显著升高(P<0.01),血清中MDA含量较模型组显著降低(P<0.01),盆腔炎颗粒各组血清中SOD及MDA浓度相关系数为﹣0.798,呈显著负相关性(P<0.01)。各治疗组大鼠子宫组织中Caspase-3、Fas/FasLmRNA和p53mRNA的表达与模型组相比较均增强(P<0.05)。其中,盆腔炎颗粒大剂量组IL-6、IL-4及SOD和MDA水平均已接近正常对照组水平(P>0.05)。除IL-6和Caspase-3外,盆腔炎颗粒大剂量组的作用均明显优于盆炎净颗粒组(P<0.01)。除IL-4外,盆腔炎颗粒中、大剂量组效果优于盆腔炎颗粒小剂量组(P<0.05)。盆腔炎颗粒各剂量组子宫组织FasmRNA及FasLmRNA表达相关系数为0.894,呈显著正相关性(P<0.01)。
结论:活血补肾法通过对慢性盆腔炎模型大鼠血清炎症细胞因子、抗氧化相关指标的调节,使机体恢复免疫平衡,有效抑制慢性盆腔炎模型大鼠炎症,加快清除自由基,减少脂质类物质的堆积,从而减轻盆腔内组织器官的慢性损伤。活血补肾法通过增强慢性盆腔炎模型大鼠子宫组织中相关凋亡指标的表达,诱导炎细胞凋亡,抑制炎细胞生长,从而减轻炎症反应。
Objective : To explore the immunoregulation mechanism of the treatment of promoting blood circulation and tonifying the kidney on the rats with chronic pelvic inflammatory disease(CPID) by observing the changes of inflammatory cytokines, antioxygen and control of cell apoptosis.
Methods :Female Wistar rats were used and the CPID model was established with mixed bacteria injection combined with mechanic injury. Fifty-six rats were randomly divided into normal group, sham operation group, model group, Penqiangyan granule large , medium and small dose groups and Penyanjing granule group. The histomorphological changes of the rats' uteri were observed using light microscope. The enzyme-linked immunosorbent assay (ELISA) was applied to measure the concentration of pro-inflammatory cytokines (IL-6) and anti-inflammatory cytokines (IL-4)in serum of rats in each group; The method of xanthinoxidase were applied to measure concentration of SOD.The method of thio-malonylurea were applied to measure the concentration of MDA.The immunochemistry method was used to detect the expression of Caspase-3 in rats’uterine tissue.The expression of Fas/FasLmRNA and p53mRNA were measured by using RT-PCR
Results:The serous levels of pro-inflammatory cytokines (IL-6) in model group were significantly higher than those in normal group(P<0.01); the serous levels of anti-inflammatory cytokines(IL-4) in model group were significantly lower than those in normal group. The serous levels of MDA in model group were significantly higher than those in normal group(P<0.01);the serous levels of SOD in model group were significantly lower than those in normal group(P<0.01).The uterine tissure expression of Caspase-3, Fas/FasLmRNA and p53mRNA were all significantly increased than those in normal group.After treatment ,the serous levels of pro-inflammatory cytokine(IL-6)in all treatment group were significantly lower than those in model group,meanwhile the anti-inflammatory cytokin(eIL-4)in all treatment group were elevated .The serous levels of MDA in all treatment groupwere significantly decreased than those in model group(P<0.01);the serous levels of SOD in all treatment group were significantly increased than those in model group(P<0.01).The coefficient correlation of the serous levels of SOD and MDA in all Penqiangyan granule groups was 0.798,showing the significant negative correlation(P<0.01). The uterine tissure expression of Caspase-3, Fas/FasLmRNA as well as p53mRNA in all treatment group were significantly increased compared with model group(P<0.01). IL-6、IL-4、SOD and MDA in Penqiangyan granule large dose group were similar to the levels of normal group(P>0.05).Except for IL-6 and Caspase-3 Penqiangyan granule large dose group significantly outweighed the Penyanjing granule group(P<0.01). Except for IL-4 Penqiangyan granule large、medium dose groups significantly outweighed the Penqiangyan granule small dose group(P<0.05).The coefficient correlation of the uterine tissure expression of FasmRNA and FasLmRNA in all Penqiangyan granule groups was 0.894,showing the significant positive correlation(P<0.01).
Conclusions: The method of promoting blood circulation and tonifying the kidney could adjust the levels of the inflammatory cytokines and the corresponding antioxygen indices,thus restore the immune balance effectively so as to inhibit inflammation,speeding up eliminating free radicles,decrease the lipid,so as to relieve the chronic injury of pelvic tissures and organs.The method of promoting blood circulation and tonifying the kidney could strengthen and induce the expression of cell apoptosis,inhibit the growth of inflammatory corpuscles,thus ease the inflammatory reaction.
引文
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