大黄灵仙方对胆固醇结石豚鼠模型胆囊Cajal间质细胞中scf/c-kit信号通路的影响

肖丽君; 甘苡榕; 刘春丽; 李承积; 杨文; 庞浇安; 滕金豪; 俞渊

doi:10.3969/j.issn.1001-5256.2023.02.019

大黄灵仙方对胆固醇结石豚鼠模型胆囊Cajal间质细胞中scf/c-kit信号通路的影响

DOI: 10.3969/j.issn.1001-5256.2023.02.019

1.
广西中医药大学研究生学院，南宁 530001
2.
广西中医药大学第一附属医院肝胆外科，南宁 530023

基金项目:

广西自然科学基金项目 (2020GXNSFAA238012);

2021年“岐黄工程”高层次人才团队培育项目 (2021006);

2020年广西中医药大学第一附属医院院级博士启动基金项目 (2020BS004);

广西中医药大学2021年研究生教育创新计划项目 (YCXJ2021068)

伦理学声明：本研究方案于2021年5月25日经由广西中医药大学第一附属医院实验动物伦理委员会审批，批号：DW20191017-023，符合实验室动物管理与使用准则。

利益冲突声明：本研究不存在研究者、伦理委员会成员以及与公开研究成果有关的利益冲突。

作者贡献声明：肖丽君负责课题设计，资料分析，拟定写作思路，撰写论文；甘苡榕参与资料分析，修改论文；刘春丽、李承积、杨文、庞浇安、滕金豪负责文献检索，收集数据及数据统计分析；俞渊负责指导撰写文章并最后定稿。

详细信息

通信作者:
俞渊，doctoryuyuan@163.com(ORCID：0000-0001-9662-8959)

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出版历程
- 收稿日期: 2022-07-13
- 录用日期: 2022-08-18
- 出版日期: 2023-02-20

Effect of Dahuang Lingxian prescription on the scf/c-kit signaling pathway in gallbladder interstitial cells of Cajal in a guinea pig model of cholesterol gallstone

1.
Graduate School of Guangxi University of Chinese Medicine, Nanning 530001, China
2.
Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi University of Chinese Medicine, Nanning 530023, China

Research funding:

Guangxi Natural Science Foundation Project (2020GXNSFAA238012);

2021 "Qihuang Project" High Level Talent Team Training Project (2021006);

2020 Hospital-level Doctoral Start-up Fund Project of the First Affiliated Hospital of Guangxi University of Traditional Chinese Medicine (2020BS004);

2021 Postgraduate Education Innovation Plan Project of Guangxi University of traditional Chinese medicine (YCXJ2021068)

More Information

Corresponding author: YU Yuan, doctoryuyuan@163.com (ORCID: 0000-0001-9662-8959)

摘要

摘要: 目的观察在大黄灵仙方的调控下干细胞生长因子(scf)、酪氨酸激酶受体(c-kit)的表达水平，探讨其影响胆囊动力学改变的可能机制，为大黄灵仙方预防胆石症的发生及复发提供理论依据。方法将45只SPF级健康雄性豚鼠随机分为正常组、模型组、中药组，其中正常组予正常饲料喂养，模型组、中药组予高脂致石饲料喂养，喂养8周后分别从各组中随机抽取5只豚鼠，肉眼观察下结石形成超过4只，则判断为模型建立成功。造模成功后中药组予大黄灵仙方灌胃，模型组予等体积生理盐水灌胃，连续灌胃给药8周后取豚鼠胆囊组织，HE染色观察观察胆囊组织的病理学改变，Western blot测定胆囊组织中TNFα的表达水平，免疫组化测定胆囊平滑肌组织中scf、c-kit的蛋白表达水平。符合正态分布的计量资料多组间比较采用单因素方差分析，进一步两两比较采用LSD多重比较法。结果 HE染色显示模型组胆囊组织炎症明显，中药组胆囊组织内炎症程度较模型组明显减轻。Western blot结果表明在模型组中，胆囊组织中TNFα的表达水平最高，中药组次之，正常组最低(P值均＜0.05)；免疫组化结果表明豚鼠胆囊平滑肌组织中scf、c-kit蛋白在正常组、中药组中的表达水平显著高于模型组(P值均＜0.05)。结论大黄灵仙方能增强胆囊动力功能，其作用机制可能与上调胆囊Cajal间质细胞中的scf、c-kit信号通路有关。
- 大黄灵仙方 /
- 胆石 /
- Cajal小肠细胞 /
- 信号传导 /
- 豚鼠
Abstract: Objective To investigate the expression levels of scf and c-kit under the regulation of Dahuang Lingxian prescription and the possible mechanism of its effect on gallbladder dynamics, and to provide a theoretical basis for Dahuang Lingxian prescription in preventing the development and recurrence of cholesterol gallstone. Methods A total of 45 specific pathogen-free healthy male guinea pigs were randomly divided into normal group, model group, and traditional Chinese medicine (TCM) group. The guinea pigs in the normal group were fed with normal diet, and those in the model group and the TCM group were fed with high-fat lithogenic diet. After 8 weeks of feeding, 5 guinea pigs were randomly selected from each group, and successful modeling was determined if gallstone was observed with the naked eye in more than 4 guinea pigs. After successful modeling, the guinea pigs in the TCM group were given Dahuang Lingxian prescription by gavage, and those in the model group were given an equal volume of normal saline by gavage. After 8 consecutive weeks of administration by gavage, gallbladder tissue samples were collected, and HE staining was used to observe the pathological changes of gallbladder tissue; Western blot was used to measure the expression level of tumor necrosis factor-α (TNF-α) in gallbladder tissue; immunohistochemistry was used to measure the protein expression levels of scf and c-kit in gallbladder smooth muscle tissue. A one-way analysis of variance was used for comparison of normally distributed continuous data between multiple groups, and the least significant difference multiple comparison method was used for further comparison between two groups. Results HE staining showed marked inflammation of gallbladder tissue in the model group, and compared with the model group, the TCM group had a significantly lower degree of inflammation. Western blot showed that the model group had the highest expression level of TNF-α in gallbladder tissue, followed by the TCM group and the normal group (P < 0.05); immunohistochemistry showed that compared with the model group, the normal group and the TCM group had significantly higher protein expression levels of scf and c-kit in gallbladder smooth muscle tissue (P < 0.05). Conclusion Dahuang Lingxian prescription can enhance the dynamic function of the gallbladder, possibly by upregulating the scf/c-kit signaling pathway in interstitial cells of Cajal in gallbladder.
- Dahuang Lingxian Formula /
- Gallstones /
- Interstitial Cells of Cajal /
- Signal Transduction /
- Guinea Pigs

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图 1 豚鼠体质量的变化情况

Figure 1. Changes in body mass of guinea pigs

下载: 全尺寸图片幻灯片

图 2 各组豚鼠胆囊组织的病理情况(HE染色，×200)

注：a，正常组；b，模型组；c，中药组。

Figure 2. Pathological changes of gallbladder tissues of guinea pigs in each group (HE staining, ×200)

下载: 全尺寸图片幻灯片

图 3 大黄灵仙方干预下TNFα的表达情况

Figure 3. Expression of TNFα under the intervention of Dahuang lingxian formula

下载: 全尺寸图片幻灯片

图 4 大黄灵仙方对TNFα表达水平的影响

Figure 4. Effect of Dahuang lingxian formula on TNFα expression

下载: 全尺寸图片幻灯片

图 5 大黄灵仙方干预下TNF、scf、c-kit的表达情况(免疫组化，×100)

Figure 5. Expression of TNFα, scf and c-kit under the intervention of Dahuang lingxian formula (immunohisto-chemistry, ×100)

下载: 全尺寸图片幻灯片

表 1 大黄灵仙方对TNFα、scf、c-kit表达水平的影响

Table 1. Effect of Dahuang Lingxian formula on the expression of TNFα, scf and c-kit

组别	TNFα	c-kit	SCF
正常组	3.48±1.79¹⁾²⁾	29.35±8.97¹⁾²⁾	28.11±5.63¹⁾²⁾
模型组	26.42±3.87	13.32±2.01	15.46±3.14
中药组	17.35±2.75¹⁾	21.11±2.72¹⁾	23.08±6.41¹⁾
F值	108.91	14.70	10.28
P值	＜0.05	＜0.05	＜0.05
注：与模型组比较，1)P＜0.05；与中药组比较，2)P＜0.05。

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