Research of Influence on Inflammation of Xieheyin on PCOS-IR Mice Based on Intestinal Mucosal Barrier and TLR4/NF-<italic style="font-style: italic">κ</italic>B/NLRP3 Pathway

Xue-fei XU; En-li WANG; Tian-ye YANG; Jing-chun YAO; Xin GONG; Wen-yue CHEN; Xin-yue LIU; Guang-ci DI; Xiu LI; Qing-ling REN

doi:10.13422/j.cnki.syfjx.20211738

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Research of Influence on Inflammation of Xieheyin on PCOS-IR Mice Based on Intestinal Mucosal Barrier and TLR4/NF-κB/NLRP3 Pathway

Pharmacology | 更新时间：2021-07-28

- Research of Influence on Inflammation of Xieheyin on PCOS-IR Mice Based on Intestinal Mucosal Barrier and TLR4/NF-κB/NLRP3 Pathway
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 27, Issue 17, Pages: 90-98(2021)
- 作者机构：
  
  1.南京中医药大学附属医院，南京 210029
  2.中药制药共性技术国家重点实验室，山东临沂 371300
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20211738
  CLC： R2-0;R271.1;R33;R364.5
- Received：02 June 2021，
  
  Published Online：07 July 2021，
  
  Published：05 September 2021
- 稿件说明：
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徐雪飞,王恩力,杨田野等.基于肠黏膜屏障及TLR4/NF-κB/NLRP3信号通路探讨燮和饮改善PCOS-IR小鼠炎症反应的影响[J].中国实验方剂学杂志,2021,27(17):90-98.

XU Xue-fei,WANG En-li,YANG Tian-ye,et al.Research of Influence on Inflammation of Xieheyin on PCOS-IR Mice Based on Intestinal Mucosal Barrier and TLR4/NF-κB/NLRP3 Pathway[J].Chinese Journal of Experimental Traditional Medical Formulae,2021,27(17):90-98.
徐雪飞,王恩力,杨田野等.基于肠黏膜屏障及TLR4/NF-κB/NLRP3信号通路探讨燮和饮改善PCOS-IR小鼠炎症反应的影响[J].中国实验方剂学杂志,2021,27(17):90-98. DOI： 10.13422/j.cnki.syfjx.20211738.

XU Xue-fei,WANG En-li,YANG Tian-ye,et al.Research of Influence on Inflammation of Xieheyin on PCOS-IR Mice Based on Intestinal Mucosal Barrier and TLR4/NF-κB/NLRP3 Pathway[J].Chinese Journal of Experimental Traditional Medical Formulae,2021,27(17):90-98. DOI： 10.13422/j.cnki.syfjx.20211738.

摘要

目的

探讨燮和饮对改善肥胖型多囊卵巢综合征胰岛素抵抗（PCOS-IR），减缓炎症反应的可能作用机制。

方法

将60只SPF级雌性C57BL/6J小鼠随机抽取10只，并设为正常组，其余小鼠均采用粪菌混悬液（2 g·kg

-1

）联合来曲唑溶液（0.002 g·kg

-1

）灌胃4周建立PCOS-IR模型。将模型复制成功的动物随机分为模型组，二甲双胍组（0.25 g·kg

-1

）及燮和饮低、中、高剂量组（10，20，40 g·kg

-1

），给予相应剂量的药物灌胃，每天1次，连续治疗4周。除空白组外，其余各组小鼠在治疗期间持续灌胃粪菌混悬液和来曲唑溶液以维持模型。动物每周称1次体质量；治疗结束后测量空腹血糖（FBG）；酶联免疫吸附测定法（ELISA）检测血清睾酮（T），促卵泡生成素（FSH），促黄体生成素（LH），空腹胰岛素（FINS）水平，计算LH/FSH及胰岛素抵抗指数（HOMA-IR）值；摘取子宫和双侧卵巢称质量后固定，苏木素-伊红（HE）染色观察卵巢组织病理形态学变化；蛋白免疫印迹法（Western blot）检测小鼠结肠组织中Toll样受体4/核转录因子-

B/NOD样受体蛋白3（TLR4/NF-

B/NLRP3）信号通路及下游炎症相关因子半胱氨酸天冬氨酸水解酶-1（Caspase-1），白细胞介素-1

（IL-1

），紧密连接蛋白-1（ZO-1），闭合蛋白（occludin）表达水平。

结果

与正常组比较，模型组小鼠体质量显著升高（

0.01）；血清FBG，FINS和HOMA-IR明显升高（

0.05）；血清T，LH/FSH显著升高（

0.01）；子宫脏器比显著降低（

0.01），卵巢脏器比显著升高（

0.01），镜下可见卵巢中囊性扩张卵泡及闭锁小卵泡数量明显增多，黄体数量显著减少，卵泡颗粒细胞层减少，卵泡膜显著增厚；结肠组织中ZO-1，occludin的相对表达量显著下调（

0.01），炎症相关蛋白TLR4，NF-

B，NLRP3，Caspase-1，IL-1

表达量明显上调（

0.05）。与模型组比较，燮和饮低、中、高剂量组小鼠体质量显著降低（

0.01）；燮和饮中、高剂量组血清FBG，FINS，HOMA-IR明显降低（

0.05）；高剂量组血清T，LH/FSH显著降低（

0.01）；卵巢脏器比明显降低（

0.05），子宫脏器比显著升高（

0.05），卵巢中囊状卵泡明显减少，内见黄体，卵巢颗粒细胞正常排列，卵泡膜略增厚；高剂量组结肠组织TLR4，NF-

B，NLRP3，Caspase-1，IL-1

蛋白表达显著下调（

0.01），ZO-1，occludin蛋白表达量显著上调（

0.01）。

结论

燮和饮调控肠道TLR4/NF-

B/NLRP3信号通路，减少下游炎症因子释放，通过重建肠黏膜屏障功能及抑制肠道炎症反应，改善肥胖和IR，从而逆转PCOS-IR。

Abstract

Objective

To investigate the possible mechanism of Xieheyin in alleviating obese polycystic ovary syndrome with insulin resistance（PCOS-IR）and reducing inflammatory response.

Method

Ten of sixty SPF femlae C57BL/6J mice were randomly selected as the normal group，and the rest mice were given letrozole 0.002 g·kg

-1

combined with fecal suspension 2 g·kg

-1

for 28 consecutive days to establish model of PCOS-IR.The mice that were successfully modeled were randomized into the model group，metformin group（0.25 g·kg

-1

），and low（10 g·kg

-1

），medium（20 g·kg

-1

），and high-dose（40 g·kg

-1

）Xieheyin groups，and administered with the corresponding drugs by gavage，once a day，for four consecutive weeks. Except the normal control group， the mice in the other groups were continuously given fecal suspension combined with letrozole solution to maintain the model during the treatment. The mice were weighed once a week.Levels of fasting blood glucose （FBG） were detected by blood glucose test strips.And enzyme-linked immunosorbent assay （ELISA） method was used to detect serum testosterone（T），follicle stimulating hormone（FSH），luteinizing hormone（LH），fasting insulin（FINS）level，and LH/FSH and Homeostasis model assesment of insulin resistance （HOMA-IR） were calculated．The uterus and ovaries were weighed and fixed.Hematoxylin-eosin（HE）staining was used to observe ovarian tissue pathology morphology. Western blot was used to detect the expression levels of tight junction key molecular zonula occludens 1（ZO-1），occludin in colon tissues，and the expression levels of Toll-like receptor 4/nuclear factor kappa B/Nod-like receptor protein 3（TLR4/NF-

B/NLRP3）signaling pathway and inflammation associated proteins cysteinyl aspartate specific proteinase-1（Caspase-1） and interleukin-1

（IL-1

） in colon tissues.

Result

Compared with normal control group，the body weight of mice in the model control group increased significantly（

0.01）. Serum FINS，FBG，HOMA-IR，T，LH/FSH were significantly increased（

0.05，

0.01）. The uterine organ ratio were decreased significantly（

0.01），while the ovarian organ ratio were significantly increased（

0.01）. The number of atresia follicles and cystic dilatation follicles increased significantly，and the number of corpus luteum significantly decreased，the thickness of follicular granulosa cells also decreased，while the white membrane thickness of the ovary increased. Tight junction related ZO-1，occludin proteins in colon tissues were all decreased（

0.01）.The relative expression levels of inflammation-related protein IL-1

，Caspase-1 and TLR4/NF-

B/NLRP3 target protein signaling pathway were significantly increased（

0.05）.Compared with model control group， the body weight of mice in the low，middle and high dose Xieheyin group decreased significantly（

0.01）. The serum T，LH/FSH，FINS，FBG，HOMA-IR were significantly decreased（

0.05，

0.01）. The uterine organ ratio were increased（

0.05），while the ovarian organ ratio were decreased（

0.05）. The number of cystic follicles decreased and corpus luteum increased，the thickness of follicular granulosa cells increased and be arranged normally，while the white membrane thickness of the ovary increased slightly. The expressions of ZO-1，occludin proteins were increased（

0.01）. The expression levels of IL-1

，Caspase-1 and TLR4/NF-

B/NLRP3 target protein in the high dose group were significantly decreased（

0.01）.

Conclusion

Xieheyin could activate intestinal TLR4/NF-

B/NLRP3 signaling pathway，inhibit pro-inflammatory factor secretion，improve obesity and IR，which was correlated with rebuilding intestinal mucosal barrier and inhibiting intestinal inflammation．

关键词

Keywords

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⁰

Research of Influence on Inflammation of Xieheyin on PCOS-IR Mice Based on Intestinal Mucosal Barrier and TLR4/NF-κB/NLRP3 Pathway

DOI：10.13422/j.cnki.syfjx.20211738

摘要

Abstract

关键词

Keywords

references