加味少腹逐瘀汤调控EGFR/PI3K/AKT信号通路诱导子宫内膜细胞自噬的作用机制

杨亚玲; 王婉润; 张作良; 王家兴; 林祥羽; 武权生

doi:10.3969/j.issn.1006-2157.2024.09.002

您当前的位置：

首页 >

文章列表页 >

加味少腹逐瘀汤调控EGFR/PI3K/AKT信号通路诱导子宫内膜细胞自噬的作用机制

“中医药防治妇科疾病”专题 | 更新时间：2024-10-09

- 加味少腹逐瘀汤调控EGFR/PI3K/AKT信号通路诱导子宫内膜细胞自噬的作用机制
- The mechanism of modified Shaofu Zhuyu Decoction regulating the EGFR/PI3K/AKT signaling pathway to induce autophagy in endometrial cells
- 北京中医药大学学报 2024年47卷第9期页码：1191-1202
- 作者机构：
  
  1.甘肃中医药大学附属医院　兰州　730000
  2.甘肃中医药大学
- 作者简介：
  
  杨亚玲，女，硕士，副主任医师
  #张作良，男，博士，住院医师，主要研究方向：中医妇科基础及临床研究，E-mail：zzllanzhou@126.com
- 基金信息：
  
  国家自然科学基金项目(82260949);甘肃省自然科学基金项目(23JRRA1199);甘肃省中医药科研项目(GZKP-2022-22);兰州市科技计划项目(2021-1-97);甘肃中医药大学附属医院2021院内一般项目(gzfy-2021-03)
- DOI：10.3969/j.issn.1006-2157.2024.09.002
  中图分类号： R285.5
- 纸质出版日期：2024-09-30，
  
  网络出版日期：2024-08-21，
  
  收稿日期：2024-05-14，
- 稿件说明：
移动端阅览
杨亚玲, 王婉润, 张作良, 等. 加味少腹逐瘀汤调控EGFR/PI3K/AKT信号通路诱导子宫内膜细胞自噬的作用机制[J]. 北京中医药大学学报, 2024,47(9):1191-1202.

YANG Yaling, WANG Wanrun, ZHANG Zuoliang, et al. The mechanism of modified Shaofu Zhuyu Decoction regulating the EGFR/PI3K/AKT signaling pathway to induce autophagy in endometrial cells[J]. Journal of Beijing University of Traditional Chinese Medicine, 2024,47(9):1191-1202.
杨亚玲, 王婉润, 张作良, 等. 加味少腹逐瘀汤调控EGFR/PI3K/AKT信号通路诱导子宫内膜细胞自噬的作用机制[J]. 北京中医药大学学报, 2024,47(9):1191-1202. DOI： 10.3969/j.issn.1006-2157.2024.09.002.

YANG Yaling, WANG Wanrun, ZHANG Zuoliang, et al. The mechanism of modified Shaofu Zhuyu Decoction regulating the EGFR/PI3K/AKT signaling pathway to induce autophagy in endometrial cells[J]. Journal of Beijing University of Traditional Chinese Medicine, 2024,47(9):1191-1202. DOI： 10.3969/j.issn.1006-2157.2024.09.002.

摘要

目的

从表皮生长因子受体(EGFR)/磷酸肌醇3－激酶(PI3K)/蛋白激酶B(AKT)信号通路介导异位子宫内膜组织细胞自噬角度探讨加味少腹逐瘀汤治疗子宫内膜异位症的潜在分子机制。

方法

72只雌性SD大鼠按照随机数字表法分为空白组、模型组、孕三烯酮组及加味少腹逐瘀汤高、中、低剂量组，每组12只。采用自体移植法构建子宫内膜异位症模型。孕三烯酮组大鼠灌胃孕三烯酮混悬液0.25 mg/(kg·d)，加味少腹逐瘀汤高、中、低剂量组

大鼠分别灌胃加味少腹逐瘀汤水煎剂30、15、7.5 g/(kg·d)，空白组及模型组大鼠灌胃等量生理盐水，连续4周。给药结束后，每只大鼠给予缩宫素2 U诱发宫缩，观察大鼠疼痛情况；记录大鼠异位子宫内膜组织的质量和体积；苏木素－伊红(HE)染色法观察各组大鼠子宫内膜组织病理变化；透射电镜观察各组大鼠子宫内膜组织超微结构；酶联免疫吸附测定法检测各组大鼠血清中雌二醇(E

)、孕酮(P)、白细胞介素－1β(IL-β)、白细胞介素－6(IL-6)、肿瘤坏死因子－α(TNF-α)、转化生长因子－β(TGF-β)、表皮生长因子(EGF)、EGFR含量；免疫荧光法检测各组大鼠子宫内膜组织中重组人自噬效应蛋白Beclin 1(Beclin-1)、微管相关蛋白1轻链3(LC3B)平均荧光强度；蛋白质印迹法检测各组大鼠子宫内膜组织中EGFR、PI3K、磷酸化磷酸肌醇3－激酶(p-PI3K)、AKT、磷酸化蛋白激酶B(p-AKT)的蛋白表达水平；实时荧光PCR法检测EGFR、PI3K、AKT mRNA表达水平。

结果

与空白组比较，模型组大鼠出现典型扭体反应，腹壁可见异位子宫内膜组织，HE结果显示，异位内膜组织增厚，间质增生，腺体扩张；超微结构仅见自噬小体，部分视野未见明显自噬结构；大鼠血清E

、IL-6、IL-1β、TNF-α、TGF-β、EGF、EGFR含量升高，P含量降低；自噬相关蛋白Beclin-1、LC3B平均荧光强度降低；EGFR、PI3K、AKT、p-PI3K、p-AKT蛋白表达升高；EGFR、PI3K、AKT mRNA表达升高(均

0.05)。与模型组比较，孕三烯酮组及加味少腹逐瘀汤高、中剂量组大鼠扭体次数减少、扭体反应潜伏时间延长；异位子宫内膜组织体积及质量减小；HE结果显示，病理损伤程度减轻；超微结构不同程度出现溶酶体、自噬溶酶体、自噬小体结构；血清E

、IL-6、IL-1β、TNF-α、TGF-β、EGF、EGFR含量降低，P含量升高；自噬相关蛋白Beclin-1、LC3B平均荧光强度升高；EGFR、PI3K、AKT、p-PI3K、p-AKT蛋白表达降低；EGFR、PI3K、AKT mRNA表达降低(均

0.05)。

结论

加味少腹逐瘀汤治疗子宫内膜异位症的机制可能与诱导EGFR/PI3K/AKT信号通路介导的异位子宫内膜组织自噬、减轻炎性反应有关。

Abstract

Objective

To investigate the potential molecular mechanism of modified

Shaofu Zhuyu

Decoction in treating endometriosis from the perspective of autophagy in ectopic endometrial tissue cells mediated by the epidermal growth factor receptor (EGFR)/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway.

Methods

Seventy-two female SD rats were divided into the blank group

the model group

the gestrinone group and the modified

Shaofu Zhuyu

Decoction high-

medium-

and low- dose groups according to the random number table method

12 rats in each group. Construction of an endometriosis model used auto-transplantation method. The rats in the gestrinone group were gavaged with 0.25 mg/(kg·d) of gestrinone suspension

the rats in

the modified

Shaofu Zhuyu

Decoction high-

medium-

and low- dose groups were gavaged with 30

and 7.5 g/(kg·d) of modified

Shaofu Zhuyu

Decoction

respectively

and the rats in the blank and model groups were gavaged with an equal amount of physiological saline

respectively. 4 weeks of continuous treatment. At the end of drug administration

2 U of oxytocin was given to each rat to induce contractions

and the pain of the rats was observed; the weight and volume of ectopic endometrial tissue of the rats were recorded. Hematoxylin-eosin (HE) staining method was used to observe the pathological changes of rat endometrial tissue in each group; transmission electron microscope was used to observe the ultrastructure of rat endometrial tissue. Enzyme-linked immunosorbent assay was used to detect the serum levels of oestradiol (E

)

progesterone (P)

interleukin-1β (IL-β)

interleukin-6 (IL-6)

tumour necrosis factor-α (TNF-α)

transforming growth factor-β (TGF-β)

epidermal growth factor (EGF)

and EGFR in the rat serum of each group. Mean fluorescence intensity of autophagy-associated protein yeast Atg6 homologue (Beclin-1) and microtubule-associated protein 1 light chain 3 (LC3B) in endometrial tissues of rat endometrium in each group by immunofluorescence assay. Protein expression levels of EGFR

PI3K

phosphorylated phosphatidylinositol 3-kinase (p-PI3K)

AKT

and phosphorylated protein kinase B (p-AKT) in endometrial tissues of rats in each group were examined by protein blotting. Real-time fluorescence PCR was used to detect EGFR

PI3K

and AKT mRNA expression levels.

Results

Compared with the blank group

the rats in the model group showed typical torsion reaction

ectopic endometrial tissue was visible in the abdominal wall

and the HE result showed thickening of the ectopic endometrial tissue

mesenchymal hyperplasia

and glandular dilatation; only autophagic vesicles were seen in the ultrastructure

and no obvious autophagic str

uctures were seen in some of the fields of view; Serum E

IL-6

IL-1β

TNF-α

TGF-β

EGF

and EGFR levels were elevated and P levels were decreased in rats; autophagy-related proteins Beclin-1 and LC3B decreased in average fluorescence intensity; protein expressions of EGFR

PI3K

AKT

p-PI3K

p-AKT was elevated; and mRNA expressions of EGFR

PI3K

AKT was elevated (

0.05). Compared with the model group

the gestrinone group and modified

Shaofu Zhuyu

Decoction high-

medium- dose groups showed a decrease in the number of torsion and a prolongation of the latency time of the torsion response

a decrease in the volume and mass of ectopic endometrial tissues

a decrease in the degree of pathological damage as shown by HE

and the appearance of lysosomes

autophagolysosomes and autophagic vesicles in different degrees in the ultrastructure; Serum E

IL-6

IL-1β

TNF-α

TGF-β

EGF

EGFR levels were decreased

and P levels were increased; the mean fluorescence intensity of autophagy-related proteins Beclin-1 and LC3B was increased; the protein expression of EGFR

PI3K

AKT

p-PI3K

and p-AKT was decreased; and the mRNA expression of EGFR

PI3K

and AKT was decreased (

0.05).

Conclusion

The mechanism of modified modified

Shaofu Zhuyu

Decoction for the treatment of endometriosis may be related to the induction of autophagy of ectopic endometrial tissues mediated by the EGFR/PI3K/AKT signaling pathway and the attenuation of inflammatory responses.

关键词

子宫内膜异位症加味少腹逐瘀汤表皮生长因子受体(EGFR)/磷酸肌醇3－激酶(PI3K)/蛋白激酶B(AKT)信号通路自噬炎症大鼠

Keywords

endometriosismodified Shaofu Zhuyu Decoctionepidermal growth factor receptor (EGFR)/phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathwayautophagyinflammationrats

references

PEIRIS AN, CHALJUB E, MEDLOCK D. Endometriosis[J]. JAMA, 2018，320(24)：2608.

TAYLOR HS, KPTLYAR AM, FLORES VA. Endometriosis is a chronic systemic disease: clinical challenges and novel innovations[J]. Lancet, 2021，397(10276)：839－852.

DE ZIEGLER D, BORGHESE B, CHAPRON C. Endometriosis and infertility: pathophysiology and management[J]. Lancet, 2010, 376(9742): 730－738.

MCCALLION A, NASIRZADEH Y, LINGEGOWDA H, et al. Estrogen mediates inflammatory role of mast cells in endometriosis pathophysiology[J]. Front Immunol, 2022, 13: 961599.

CARBONE G, NELSON K, BAUMGARTNER C, et al. Endometriosis: cell death and cell signaling machinery[J]. Endocrinology, 2023, 164(6)：57－93.

王猛，马浩天，关思宇，等．螺旋藻多糖对子宫内膜异位大鼠血清指标及PI3K/Akt/mTOR信号通路基因表达的影响[J].天然产物研究与开发，2021，33(1)：16－22.

曹立花，李丽娟，胡晓丹，等. 姜黄素对子宫内膜异位症大鼠PI3K/AKT/GSK－3β通路及肥大细胞活化的影响[J]. 中国免疫学杂志，2021, 37(4): 480－485.

毛海燕，陈元欢，武权生，等．加味少腹逐瘀汤对子宫内膜异位症寒凝血瘀证盆腔疼痛神经血管生成的影响[J].中国实验方剂学杂志，2022，28(12)：141－147.

毛海燕，陈元欢，吉秀家，等．加味少腹逐瘀汤抑制NOD1/RIP2/NF－κB信号通路改善子宫内膜异位症痛经大鼠异位内膜及炎性微环境[J].时珍国医国药，2022，33(10)：2334－2338.

黄灿灿，吉秀家，张作良，等．寒湿瘀结证子宫内膜异位症纤维化小鼠腹腔微环境NLRP3炎性小体活化相关蛋白及Th17/Treg的表达特点[J].时珍国医国药，2023，34(8)：2016－2021.

卢建军，戴晓怡，李响，等. 萝卜硫素对子宫内膜异位症模型大鼠在位内膜增殖、血管生成及JAK2/STAT3信号通路的影响[J]. 中国免疫学杂志，2021, 37(11): 1297－1301.

赵瑞华．中医药治疗子宫内膜异位症的研究述评[J].北京中医药大学学报，2023，46(9)：1185－1194.

黄灿灿，毛海燕，吉秀家，等. 基于TLR2/MyD88/NF－κB信号通路探讨加味少腹逐瘀汤对寒湿瘀结证子宫内膜异位症痛经小鼠腹腔炎症微环境干预作用[J].中国药理学通报，2024, 40(4): 784－791.

MIZUSHIMA N, KOMATSU M. Autophagy: renovation of cells and tissues[J]. Cell, 2011，147(4)：728－741.

SAMARE-NAJAF M, NEISY A, SAMAREH A, et al. The constructive and destructive impact of autophagy on both genders′ reproducibility, a comprehensive review[J]. Autophagy, 2023, 19(12): 3033－3061.

PATEL BG, RUDNICKI M, YU J, et al. Progesterone resistance in endometriosis: origins, consequences and interventions[J].Acta Obstet Gynecol Scand，2017，96(6)：623－632.

SHEN HH, ZHANG T, YANG HL, et al. Ovarian hormones-autophagy-immunity axis in menstruation and endometriosis[J]. Theranostics, 2021，11(7)：3512－3526.

黄毓菲. MST1调控子宫内膜异位症相关腹腔巨噬细胞功能影响异位内膜细胞自噬的作用机制研究[D]. 济南：山东大学，2023.

HUANG JY, CHEN X, LV YC. HMGB1 mediated inflammation and autophagy contribute to endometriosis[J]. Front Endocrinol, 2021, 12: 616696.

HE R, LIU X, ZHANG J, et al. NLRC5 inhibits inflammation of secretory phase ectopic endometrial stromal cells by up-regulating autophagy in ovarian endometriosis[J]. Front Pharmacol, 2020, 11: 1281.

MATSUZAKI S, GREMEAU AS, POULY JL. Impaired pathogen-induced autophagy and increased IL-1β and TNFα release in response to pathogenic triggers in secretory phase endometrial stromal cells of endometriosis patients[J]. Reprod Biomed Online, 2020, 41(5): 767－781.

CHOI J, JO M, LEE E, et al. Inhibition of the NLRP3 inflammasome by progesterone is attenuated by abnormal autophagy induction in endometriotic cyst stromal cells: implications for endometriosis[J]. Mol Hum Reprod, 2022, 28(4): gaac007.

KHASHCHENKO EP, VYSOKIKH MY, MAREY MV, et al. Altered glycolysis, mitochondrial biogenesis, autophagy and apoptosis in peritoneal endometriosis in adolescents[J]. Int J Mol Sci, 2024, 25(8): 4238.

HUNG SW, ZHANG R, TAN Z, et al. Pharmaceuticals targeting signaling pathways of endometriosis as potential new medical treatment: a review[J]. Med Res Rev, 2021, 41(4): 2489－2564.

张艳青，周燕飞，龙玲，等．EGFR和Annexin－Ⅰ在子宫内膜异位症异位内膜的表达及意义[J].实用妇产科杂志，2012，28(11)：946－949.

D′AMICO R, IMPELLIZZERI D, CORDARO M, et al. Complex interplay between autophagy and oxidative stress in the development of endometriosis[J]. Antioxidants, 2022, 11(12): 2484.

浏览量

下载量

CSCD

文章被引用时，请邮件提醒。

提交

工具集

关联资源

活血消异方对子宫内膜异位症大鼠卵巢功能及妊娠结局的影响研究

大建中汤调控小胶质细胞自噬治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制

基于AMPK/mTOR/ULK1通路探讨解毒通络保肾方改善糖尿病肾脏疾病的作用机制

抗纤益心方通过AMPK/mTOR通路改善扩张型心肌病大鼠心功能的作用机制

左归降糖解郁方对糖尿病并发抑郁症海马NVU神经元的调控作用及其机制