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目的:探究重楼皂苷Ⅱ(polyphyllin,PPⅡ)与反式肉桂醛(trans-cinnamaldehyde,TCA)通过调控ROCK/LIMK/cofilin通路抑制子宫腺肌病(adenomyosis,AM)进展的机制。方法:提取人子宫腺肌病异位内膜细胞(adenomyosis-derived cells,AMDC)进行培养,采用CCK-8法检测梯度浓度的重楼皂苷Ⅰ、Ⅱ、Ⅶ(PPⅠ、PPⅡ、PPⅦ)及TCA对AMDC活力的影响;采用正交试验设计筛选PPⅡ与TCA(简称PT)的最佳配比;采用Transwell实验和免疫荧光实验检测PT抑制AMDC侵袭、迁移的能力;采用Western blot检测ROCK1、LIMK、p-LIMK、cofilin、p-cofilin的蛋白表达水平;对ICR小鼠腹腔注射梯度浓度的PT 30 d,采用苏木素-伊红染色法观察小鼠肝、脾和肾组织的形态学改变,采用酶联免疫吸附法检测血清中丙氨酸氨基转移酶(alanine aminotransferase,ALT)、谷草转氨酶(aspartate aminotransferase,AST)、碱性磷酸酶(alkaline phosphatase,ALP)和血尿素氮(blood urea nitrogen,BUN)活性。他莫昔芬滴喂新生ICR雌鼠建立AM模型,设置空白对照组、模型组、PT低剂量组、PT高剂量组和阳性药物复方米非司酮组,药物干预30 d,采用H&E染色检测小鼠子宫的形态学改变;采用免疫组化评估子宫组织内ROCK1、p-LIMK、p-cofilin以及KI67的表达水平改变。结果:体外实验结果显示,PPⅡ和TCA抑制AMDC活力效果较好;PT抑制AMDC侵袭、迁移效果最显著(P<0.001),下调ROCK1、LIMK、p-LIMK、cofilin、p-cofilin蛋白表达(P<0.05),抑制AMDC骨架重排(P<0.001)。体内实验结果表明,PT药物未损伤小鼠肝、脾和肾组织形态,不影响血清ALP、BUN活性和AST/ALT比值,能够减轻AM模型小鼠子宫内膜腺体向肌层侵袭的数量,降低AM小鼠子宫组织中ROCK1、p-LIMK、p-cofilin和KI67的表达水平(P<0.05)。结论:PT可通过调控ROCK/LIMK/cofilin通路抑制AM腺体向肌层的侵袭和迁移,延缓AM进展。
Abstract：Objective:To investigate the mechanism of polyphyllin(PP Ⅱ) and trans-cinnamaldehyde(TCA) inhibiting the progression of adenomyosis(AM) through modulating the ROCK/LIMK/cofilin pathway.Methods:Human adenomyosis-derived cells(AMDC) were extracted and cultured,and the effects of gradient concentrations of polyphyllin Ⅰ,Ⅱ,Ⅶ(PP Ⅰ,PPⅡ,PP Ⅶ) and TCA on the viability of AMDC were detected using CCK-8 assay;the optimal ratio of PP Ⅱ to TCA(PT for short) was screened by orthogonal experiment;Transwell assay and immunofluorescence assay were used to detect the ability of PT to inhibit AMDC invasion and migration;Western blot was used to detect the protein expression levels of ROCK1,LIMK,pLIMK,cofilin,and p-cofilin;graded concentrations of PT were injected intrap erit one ally into ICR mice for 30 d.Hematoxylin-eosin staining(H&E) was used to observe the morphological changes in the liver,spleen,and kidney tissues of the mice,and enzyme-linked immunosorbent assay(ELISA) was used to measure the activities of alanine aminotransferase(ALT),aspartate aminotransferase(AST),alanine aminotransferase(ALP),and blood urea nitrogen(BUN) in serum.Tamoxifen was fed to the neonatal ICR female mice to establish the AM model,as well as the blank control group,the model group,the low-dose group of PT,the high-dose group of PT,and the group of the positive compounded mifepristone group were set up,and administered drugintervention for 30 d.Morphological changes in the mouse uterus were detected by H&E staining,and the expression levels of ROCK1,p-LIMK,p-cofilin,and KI67 in uterine tissues were detected by immunohistochemistry.Results:The results of in vitro experiments showed that PP Ⅱ and TCA were more effective in inhibiting AMDC viability;PT most significantly inhibited AMDC invasion and migration(P<0.001),downregulated the protein expression of ROCK1,LIMK,p-LIMK,cofilin,and p-cofilin(P<0.05),and inhibited the cytoskeletal rearrangement of AMDC(P<0.001).The results of in vivo experiments showed that PT drug did not damage the liver,spleen,and kidney tissue morphology;did not affect serum ALP,BUN activity,and AST/ALT ratio;attenuated the number of endometrial glands invading the myometrium in AM model mice,and reduced the expression levels of ROCK1,p-LIMK,p-cofilin,and KI67 in uterine tissues of AM mice(P<0.05).Conclusion:PT can reduce the invasion of glands into the myometrium and delay the progression of AM by inhibiting the expression of ROCK/LIMK/cofilin pathway.
1 Tsikouras P,Kritsotaki N,Nikolettos K,et al.The impact of adenomyosis on pregnancy[J].Biomedicines,2024,12(8):1925.
2 Liu Z,Sun Z,Liu H,et al.Single-cell transcriptomic analysis of eutopic endometrium and ectopic lesions of adenomyosis[J].Cell Biosci,2021,11(1):51.
3 Jensen KK,Pyle C,Foster BR,et al.Adenomyosis in pregnancy:Diagnostic pearls and pitfalls[J].Radiographics,2021,41(3):929-944.
4 Jensen JT,Lukkari-Lax E,Schulze A,et al.Contraceptive efficacy and safety of the 52-mg levonorgestrel intrauterine system for up to 8 years:Findings from the mirena extension trial[J].Am J Obstet Gynecol,2022,227(6):873.e871-873.e812.
5中国医师协会妇产科医师分会子宫内膜异位症专业委员会,中国医学科学院北京协和医院妇产科.子宫腺肌病诊治中国专家共识[J].中华妇产科杂志,2020,55(6):376-383.Endometriosis Professional Committee of the Obstetricians and Gynaecologists Branch of the ChinesePhysicians Association,Department of Obstetrics and Gynecology,Peking Union Medical College Hospital,Chinese Academy of Medical Sciences.Chinese expert consensus on the diagnosis and treatment of adenomyosis[J].Chin J Obstet Gynecol,2020,55(6):376-383.
6王领弟,刘洋,石雅馨,等.通脉化癥合剂治疗阳虚血瘀型子宫腺肌病的疗效评价[J].中国中西医结合影像学杂志,2024,22(1):51-54,95.Wang LD,Liu Y,Shi YX,et al.Efficacy evaluation of Tongmai Huazheng Mixture on adenomyosis of yang deficiency and blood stasis syndrome[J].Chin Imag J Integr Tradit West Med,2024,22(1):51-54,95.
7 《中成约治疗优势病种临床应用指南》标淮化项目组,中成药治疗痛经临床应用指南(2021年)[J].中国中西医结合杂志,2021,41(12):1413-1425.Standardization Project Team of"Guidelines for the Clinical Application of Proprietary Chinese Medicine in the Treatment of Dominant Diseases".Guidelines for the clinical application of proprietary Chinese medicines in the treatment of dysmenorrhea(2021)[J].Chin J Integr Tradit West Med,2021,41(12):1413-1425.
8 Wang H,Zhang J,Zhu Q,et al.Integrating network pharmacology and experimental validation deciphers the mechanism of Guizhi Fuling Wan against adenomyosis[J].Evid Based Complement Alternat Med,2021,2021:6034147.
9陈蓉,贺丰杰,李长忠,等.宫血宁胶囊妇产科临床应用指导建议[J].中国实用妇科与产科杂志,2017,33(4):383-384.Chen R,He FJ,Li CZ,et al.Advice on the clinical application of Gongxuening Capsules in obstetrics and gynecology[J].Chin J Pract Gynecol Obstet,2017,33(4):383-384.
10张一诺,王子璐,石雅馨,等.桂楼复方对子宫腺肌病异位内膜细胞迁移和侵袭的影响研究[J].中国全科医学,2024,27(24):3007-3014.Zhang YN,Wang ZL,Shi YX,et al.Effect of Guilou Compound on migration and invasion of adenomyosis derived cells[J].Chin Gen Pract,2024,27(24):3007-3014.
11喻梦蝶,王信,刘洪云,等.桂枝挥发油对子宫腺肌病异位内膜细胞侵袭、迁移的研究[J].中药药理与临床,2023,39(3):52-58.Yu MD,Wang X,Liu HY,et al.Effects of Cinnamomi ramulus volatile oil on the invasion and migration of adenomyosis derived cells[J].Pharmacol Clin Chin Mater Med,2023,39(3):52-58.
12石雅馨,王子璐,曹涵,等.重楼皂苷Ⅶ通过调控上皮间充质转化抑制子宫腺肌病异位内膜迁移和侵袭的分子机制[J].中华中医药杂志,2024,39(5):2534-2541.Shi YX,Wang ZL,Cao H,et al.Molecular mechanismof polyphyllinⅦinhibiting ectopic endometrial migration and invasion of ad eno my os is by regulating epithelialmesenchymal transition[J].Chin J Tradit ChinMed Parm,2024,39(5):2534-2541.
13 Shi Y,Xu L,Wang X,et al.Paris polyphylla ethanol extract and polyphyllin I ameliorate ad eno my os is by inhibiting epithelial-mesenchymal transition[J].Phytomedicine,2024,127:155461.
14张紫叶,任晓亮,王萌.组分中药生物药剂分类学研究进展[J].中成药,2024,46(6):1914-1921.Zhang ZY,Ren XL,Wang M.Progress in the taxonomy of biopharmaceuticals of component Chinese medicines[J].Chin Tradit Patent Med,2024,46(6):1914-1921.
15 Yang D,Zhu Z,Yao Q,et al.ccTCM:A quantitative component and compound platform for promoting the research of traditional Chinese medicine[J].Comput Struct Biotechnol J,2023,21:5807-5817.
16赵伟,孙国志.不同种实验动物间用药量换算[J].畜牧兽医科技信息,2010,26(5):52-53.Zhao W,Sun GZ.Conversion of drug dosage between different species of experimental animals[J].Chin J Anim Husb Vet Med,2010,26(5):52-53.
17林益,李咏倩,齐聪,等.米非司酮致胚胎着床障碍动物模型的再评价[J].生殖医学杂志,2020,29(5):650-656.Lin Y,Li YQ,Qi C,et al.Reassessment of animal model of embryo implantation failure induced by mifepristone[J].J Reprod Med,2020,29(5):650-656.
18 Guo S,Li Z,Yan L,et al.GnRH agonist improves pregnancy outcome in mice with induced adenomyosis by restoring endometrial receptivity[J].Drug Des Devel Ther,2018,12:1621-1631.
19 Li M,Li T,Jin T,et al.Abnormal activation of the Wnt3a/β-catenin signaling pathway promotes the expression of T-box transcription factor 3(TBX3)and the epithelial-mesenchymal transition pathway to mediate the occurrence of adenomyosis[J].Mol Biol Rep,2023,50(12):9935-9950.
20王楚矗,王信,于敏敏,等.基于PI3K/Akt/HIF-1α通路探讨桂枝茯苓胶囊-散结镇痛胶囊-宫血宁胶囊联用抑制子宫腺肌病进展的作用机制[J].中草药,2024,55(3):842-852.Wang CC,Wang X,Yu MM,et al.Mechanism of Guizhi F uling Capsules-Sanjie Zhentong CapsulesGongxue Ning Capsules combination in inhibiting progression of adenomyosis based on PI3K/Akt/HIF-1αpathway[J].Chin Tradit Herb Drugs,2024,55(3):842-852.
21武红莉,荆志伟,王忠.中药复方优化设计方法研究进展[J].中国实验方剂学杂志,2010,16(13):224-226.Wu HL,Jing ZZ,Wang Z.Development of research method of further optimization of prescription[J].Chin J Exp Tradit Med Form,2010,16(13):224-226.
22 Piriyev E,Namazov A,Mahalov I,et al.Clinical and surgical characteristics of abdominal wall endometriosis:A multicenter case series of 80 women[J].In Vivo,2023,37(2):756-762.
23郭诗韵,邓志灏,李艳,等.中药复方配伍科学内涵阐释研究的新理论[J].中国实验方剂学杂志,2024,30(23):338-344.Guo SY,Deng ZH,Li Y,et al.A review of theoretical research on interpretation of scientific connotation of compatibility of traditional Chinese medicine compounds[J].Chin J Exp Tradit Med Form,2024,30(23):338-344.
24张宏,李奇璋,刘爱芬,等.基于成分配伍的中药复方研究策略[J].中国科学:生命科学,2024,54(5):911-924.Zhang H,Li QZ,Liu AF,et al.Chinese medicine formula research strategy based on compound compatibility[J].Sci Sin(Vitae),2024,54(5):911-924.
25 Peng J,Song X,Yu W,et al.The role and mechanism of cinnamaldehyde in cancer[J].J Food Drug Anal,2024,32(2):140-154.
26刁翰林,王雁飞,张毅然,等.桂枝-重楼-三七宫内缓释系统对子宫腺肌病模型大鼠子宫内膜细胞增殖及侵袭能力的影响[J].中华中医药杂志,2021,36(10):6059-6063.Diao HL,Wang YF,Zhang YR,et al.Research of Guizhi-Chonglou-Sanqi intrauterine sustained release system on proliferation and invasion of endometrial[J].Chin J Tradit Chin Med,2021,36(10):6059-6063.
27 Zhang K,Zhou Z,Wang C,et al.Mechanism study of Cinnamomi ramulus and Paris polyphylla Sm.drug pair in the treatment of adenomyosis by network pharmacology and experimental validation[J].Evid Based Complement Alternat Med,2022,2022:2624434.
28梁杰,柳维林,韩平,等.补肾壮筋汤对兔膝骨关节炎软骨形态学和骨架蛋白ROCK、Cofilin、Phospho-Cofilin、LIMK1和Phospho-LIMK1的影响[J].中华中医药杂志,2015,30(10):3732-3735.Liang J,Liu WL,Han P,et al.Influence of Bushen Zhuangjin Decoction on cartilage morphology and skeleton proteins of ROCK,Cofilin,Phospho-Cofilin,LIMK1,and Phospho-LIMK1 in rabbits with knee osteoarthritis[J].Chin J Tradit Chin Med Pharm,2015,30(10):3732-3735.
29时俊宇,徐晓黎,高红艳,等.川楝素对卵巢癌细胞恶性生物学行为的影响[J].中国临床药理学杂志,2021,37(7):847-850.Shi JY,Xu XL,Gao HY,et al.Effect of ligustrazine on malignant biological behavior of ovarian cancer cells[J].Chin J Clin Pharmacol,2021,37(7):847-850.
30国家药典委员会.中华人民共和国药典:一部[M].北京:中国医药科技出版社,2020:1088.Chinese Pharmacopoeia Commission.Pharmacopoeia of the People's Republic of China:PartⅠ[M].Beijing:China Medical Science Press,2020:1088.
31 Li J,Sun H,Jiang X,et al.PolyphyllinⅡinduces protective autophagy and apoptosis via inhibiting PI3K/AKT/mTOR and STAT3 signaling in colorectal cancer cells[J].Int J Mol Sci,2022,23(19):11890.
32 Miao W,Wang Z,Gao J,et al.PolyphyllinⅡinhibits breast cancer cell proliferation via the PI3K/Akt signaling pathway[J].Mol Med Rep,2024,30(6):224.
33 Yan S,Bao S,Chen T,et al.Cinnamaldehyde alleviates aspirin-induced gastric mucosal injury by regulating PI3K/AKT pathway-mediated apoptosis,autophagy,and ferroptosis[J].Phytomedicine,2024,132:155791.
34 Nile A,Shin J,Shin J,et al.Cinnamaldehyde-rich cinnamon extract induces cell death in colon cancer cell lines HCT 116 and HT-29[J].Int J Mol Sci,2023,24(9):8191.
35 Lee MH,Kundu JK,Chae JI,et al.Targeting ROCK/LIMK/cofilin signaling pathway in cancer[J].Arch Pharm Res,2019,42(6):481-491.
36 Quadri R,Rotondo G,Sertic S,et al.A Haspin-ARHGAP11A axis regulates epithelial morphogenesis through Rho-ROCK dependent modulation of LIMK1-Cofilin[J].iScience,2023,26(10):108011.
37 Guan G,Cannon RD,Coates DE,et al.Effect of the Rho-Kinase/ROCK signaling pathway on cytoskeleton components[J].Genes(Basel),2023,14(2):272.
38 Tavakoli Pirzaman A,Mansoori R,Hosseini SM,et al.The effect of melatonin on capecitabine-induced hepatic and renal toxicity in rats[J].Hum Exp Toxicol,2024,43:9603271231223506.
39 Raeeszadeh M,Moradi M,Ayar P,et al.The antioxidant effect of Medicago sativa L.(Alfalfa)ethanolic extract against mercury chloride(HgCl_2)toxicity in rat liver and kidney:An in vitro and in vivo study[J].Evid Based Complement Alternat Med,2021,2021:8388002.
40 Jiao X,Wang B,Yang L,et al.FMNL2 suppresses cell migration and invasion of breast cancer:A reduction of cytoplasmic p27 via RhoA/LIMK/Cofilin pathway[J].Cell Death Discov,2022,8(1):155.
基本信息:
DOI：10.13210/j.cnki.jhmu.20241217.002
中图分类号:R285
引用信息:
[1]曹涵,王子璐,王楚矗,等.重楼皂苷Ⅱ与反式肉桂醛调控ROCK/LIMK/cofilin通路抑制子宫腺肌病异位内膜细胞侵袭迁移的机制研究[J].海南医科大学学报,2025,31(19):1477-1487.DOI:10.13210/j.cnki.jhmu.20241217.002.
基金信息:
国家自然科学基金资助项目（82474559）; 山东省自然科学创新发展联合基金资助项目（ZR2023LZY024）~~
2024-12-17
2024-12-17
2024-12-17