Effect of <italic style="font-style: italic">Dahuang Lingxian Formula</italic>（大黄灵仙方）on miRNA-30b Expression and Inflammation-fibrosis Related Factors in Biliary Duct Tissue of Cholecystitis Model Rats

JU Yanfei; CHEN Yalu; MENG Jianlin; LAO Yongcai; WANG Qingjian

doi:10.13288/j.11-2166/r.2023.18.014

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Effect of Dahuang Lingxian Formula（大黄灵仙方）on miRNA-30b Expression and Inflammation-fibrosis Related Factors in Biliary Duct Tissue of Cholecystitis Model Rats

更新时间：2023-09-12

- Effect of Dahuang Lingxian Formula（大黄灵仙方）on miRNA-30b Expression and Inflammation-fibrosis Related Factors in Biliary Duct Tissue of Cholecystitis Model Rats
- Journal of Traditional Chinese Medicine Vol. 64, Issue 18, Pages: 1908-1915(2023)
- 作者机构：
  
  1.广西中医药大学，广西壮族自治区南宁市青秀区五合大道13号，530200
  2.广西中医药大学第一附属医院
- 作者简介：
- 基金信息：
- DOI：10.13288/j.11-2166/r.2023.18.014
  CLC：
- Received：12 April 2023，
  
  Revised：2023-06-02，
  
  Published：17 September 2023
- 稿件说明：
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居燕飞,陈雅璐,蒙健林等.大黄灵仙方对胆道炎症模型大鼠胆管组织miRNA-30b表达及炎症-纤维化相关因子的影响[J].中医杂志,2023,64(18):1908-1915.

JU Yanfei,CHEN Yalu,MENG Jianlin,et al.Effect of Dahuang Lingxian Formula（大黄灵仙方）on miRNA-30b Expression and Inflammation-fibrosis Related Factors in Biliary Duct Tissue of Cholecystitis Model Rats[J].Journal of Traditional Chinese Medicine,2023,64(18):1908-1915.
居燕飞,陈雅璐,蒙健林等.大黄灵仙方对胆道炎症模型大鼠胆管组织miRNA-30b表达及炎症-纤维化相关因子的影响[J].中医杂志,2023,64(18):1908-1915. DOI： 10.13288/j.11-2166/r.2023.18.014.

JU Yanfei,CHEN Yalu,MENG Jianlin,et al.Effect of Dahuang Lingxian Formula（大黄灵仙方）on miRNA-30b Expression and Inflammation-fibrosis Related Factors in Biliary Duct Tissue of Cholecystitis Model Rats[J].Journal of Traditional Chinese Medicine,2023,64(18):1908-1915. DOI： 10.13288/j.11-2166/r.2023.18.014.

摘要

目的

探讨大黄灵仙方减轻胆道炎症防治胆石症的可能作用机制。

方法

将50只SD大鼠随机分为空白组、模型组、大黄灵仙组、大黄灵仙+空白抑制剂组、大黄灵仙+抑制剂组，每组10只。除空白组外，其余各组均采用脂多糖（LPS）诱导构建大鼠胆道炎症动物模型。大黄灵仙组、大黄灵仙+空白抑制剂组、大黄灵仙+抑制剂组三组大鼠制备胆道炎症模型前3天开始中药灌胃，给药剂量为320 mg/（kg·d），空白组、模型组大鼠给予2 ml/100 g蒸馏水灌胃，每天2次，连续干预6天。采用HE染色观察各组胆管病理形态改变，免疫组化法检测胆管核因子κB（NF-κB）蛋白表达，ELISA法检测大鼠血清白细胞介素1β（IL-1β）和肿瘤坏死因子α（TNF-α）表达水平，实时PCR法检测胆管组织中微小核糖核酸-30b（miRNA-30b）及转化生长因子β1（TGF-β1）、核苷酸结合寡聚化结构域样受体蛋白3（NLRP3）和骨形态发生蛋白2（BMP2）mRNA表达，Western blot法检测TGF-β1、NLRP3以及BMP2蛋白表达量。

结果

与空白组比较，模型组大鼠胆管结构异常，汇管区内可见大量淋巴细胞、浆细胞浸润以及胆小管扩张，NF-κB蛋白阳性表达增加，出现核浸润，血清炎症因子IL-1β、TNF-α表达明显升高，胆管组织中TGF-β1、NLRP3、BMP2 mRNA以及蛋白表达量显著增加，miRNA-30b表达水平明显下降（

＜0.01）。与模型组比较，大黄灵仙组和大黄灵仙+空白抑制剂组胆管病理形态均改善，炎性细胞浸润减少，NF-κB阳性表达降低，同时核浸润减轻，血清炎症因子IL-1β、TNF-α表达水平下降，胆管组织中TGF-β1、NLRP3、BMP2 mRNA以及蛋白表达量均降低，miRNA-30b表达水平均明显升高（

＜0.05或

＜0.01），而大黄灵仙+抑制剂组各指标改善不明显（

＞0.05）。大黄灵仙组和大黄灵仙+空白抑制剂组各指标差异亦无统计学意义（

＞0.05）。

结论

大黄灵仙方可能通过调控miRNA-30b及炎症-纤维化相关因子表达，起到延缓胆石症病程进展的作用。

Abstract

Objective

To explore the possible mechanism of reducing cholecystitis and preventing cholelithiasis by

Dahuang Lingxian Formula

（大黄灵仙方， DLF）.

Methods

Fifty SD rats were randomly divided into blank group， model group， DLF group， DLF + blank inhibitor group， and DLF + inhibitor group， with 10 rats in each group. The rat model of

holecystitis was established by lipopolysaccharide （LPS） induction in all the groups except for the blank group. Rats in DLF group， DLF + blank inhibitor group and DLF + inhibitor group received intragastric administration of 320 mg/ （kg·d） of DLF 3 days before the preparation of

holecystitis model， while those in blank group and model group were given 2 ml/100 g of distilled water by gastric， twice a day， for 6 consecutive days. Hematoxylin-eosin （HE） staining was used to observe the histopathologic changes of bile duct tissues in each group. The expression of nuclear factor κB （NF-κB） protein in bile duct tissues was detected by immunohistochemistry. The expression levels of interleukin1β （IL-1β） and tumor necrosis factor α （TNF-α） in serum of rats were detected by enzym-linked immunosorbent assay （ELISA）. The mRNA expression levels of miRNA-30b， transforming growth factor β1 （TGF-β1）， nucleotide-binding oligomerization domain-like receptor protein 3 （NLRP3） and bone morphogenetic protein 2 （BMP2） in bile duct tissues were detected by real-time PCR， and the protein expression levels of TGF-β1， NLRP3 and BMP2 were detected by Western blot.

Results

Compared to those in the blank group， the structure of the bile duct in the model group was abnormal， and a large number of lymphocytes， plasma cell infiltration and bile canaliculi dilation were seen in the portal area； the positive expression of NF-κB protein increased； there was nuclear infiltration； the expressions of serum inflammatory factors IL-1β and TNF-α， as well as the mRNA and protein expressions of TGF-β1， NLRP3 and BMP2 in bile duct tissue significantly increased， while the expression level of miRNA-30b significantly decreased （

＜0.01）. Compared to those in the model group， the pathological morphology of the bile ducts in the DLF group and DLF + blank inhibitor group was improved， and the infiltration of inflammatory cells was reduced， with decreased positive expression of NF-κB and nuclear infiltration； expression levels of serum inflammatory factors IL-1β and TNF-α， and the mRNA and protein expressions of TGF-β1， NLRP3 and BMP2 in bile duct tissue decreased， while the expression level of miRNA-30b significantly increased （

＜0.05 or

＜0.01）. Compared to the model group， those indicators in the DLF + inhibitor group was not significantly improved （

＞0.05）. There was no significant difference in the indicators between the DLF group and the DLF + blank inhibitor group （

＞0.05）.

Conclusion

DLF

may play a role in delaying the progression of cholelithiasis by regulating the expression of miRNA-30b and inflammation-fibrosis related factors.

关键词

Keywords

references

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Effect of Dahuang Lingxian Formula（大黄灵仙方）on miRNA-30b Expression and Inflammation-fibrosis Related Factors in Biliary Duct Tissue of Cholecystitis Model Rats

DOI：10.13288/j.11-2166/r.2023.18.014

摘要

Abstract

关键词

Keywords

references