甲硫脑啡肽通过阿片受体促进脂多糖作用下奶牛子宫内膜基质细胞的增殖

doi:10.11843/j.issn.0366-6964.2023.03.034

畜牧兽医学报 ›› 2023, Vol. 54 ›› Issue (3): 1229-1239.doi: 10.11843/j.issn.0366-6964.2023.03.034

甲硫脑啡肽通过阿片受体促进脂多糖作用下奶牛子宫内膜基质细胞的增殖

孙雯叶^1,2, 李建基^1,2, 王亨^1,2, 董俊升^1,2, 李俊^1,2, 崔璐莹^1,2*

1. 扬州大学兽医学院江苏省重要疫病与人兽共患病防控协同创新中心, 扬州 225009;
2. 教育部农业与农产品安全国际合作联合实验室, 扬州 225009

收稿日期:2022-05-31 出版日期:2023-03-23 发布日期:2023-03-21
通讯作者: 崔璐莹,主要从事反刍动物围产期疾病发病机理与防控技术的教学与研究,E-mail:dwyxcly@126.com
作者简介:孙雯叶(1997-)，女，江苏常州人，硕士生，主要从事反刍动物子宫内膜炎发病机制的研究，E-mail：itfwfliu@163.com
基金资助:
国家自然科学基金项目(32072937；31802253；32102735)；中国博士后科学基金项目(2018M632398)；江苏现代农业产业技术体系建设专项(JATS[2018]315)；江苏省333高层次人才培养工程资助项目；江苏省基础研究计划青年基金项目(BK20210808)；扬州大学优秀青年骨干教师资助项目；江苏高校优势学科建设工程资助项目(PAPD)；江苏高校品牌专业建设工程资助项目(TAPP)

Effect of Met-enkephalin on Lipopolysaccharide-stimulated Proliferation of Bovine Endometrial Stromal Cells

SUN Wenye^1,2, LI Jianji^1,2, WANG Heng^1,2, DONG Junsheng^1,2, LI Jun^1,2, CUI Luying^1,2*

1. Jiangsu Co-innovation Center for the Prevention and Control of Important Animal Infectious Diseases and Zoonoses/College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China;
2. Joint International Research Laboratory of Agriculture and Agri-product Safety of the Ministry of Education, Yangzhou 225009, China

Received:2022-05-31 Online:2023-03-23 Published:2023-03-21

摘要/Abstract

摘要： 为明确甲硫脑啡肽(Met-enkephalin，MENK)调控原代奶牛子宫内膜基质细胞(bovine endometrial stromal cell，BESC)增殖的作用机制，采用脂多糖(lipopolysaccharide，LPS)、MENK和阿片受体拮抗剂共处理细胞进行了本试验。本研究使用的阿片受体拮抗剂包括非选择性阿片受体拮抗剂纳洛酮(naloxone，Nx)、δ阿片受体拮抗剂ICI 154129和μ阿片受体拮抗剂CTAP，分组情况：空白对照组，LPS处理组，LPS与MENK(10^-10mol·L^-1)共处理组(LM组)，LPS、MENK和阿片受体拮抗剂(Nx、CTAP和ICI 154129，均为10^-10mol·L^-1)共处理组以及LPS与阿片受体拮抗剂共处理组。检测细胞活性、细胞周期、细胞增殖相关因子基因表达以及Wnt/β-catenin和PI3K/AKT相关通路变化。结果表明：阿片受体拮抗剂单独或与LPS共处理细胞24 h，对细胞活力无影响(P > 0.05)。与LM组相比，LPS、MENK和Nx共处理组(LMN组)以及LPS、MENK和ICI 154129共处理组(LMI组)的G1期相对细胞数升高(P<0.05)，LMN组以及LPS、MENK和CTAP共处理组(LMC组)S期相对细胞数降低(P<0.05)。与LM组相比，LMN组和LMI组细胞CCN2、TGFB1和TGFB3基因表达下调(P<0.01)，LMC组细胞CCN2基因表达下调(P < 0.01)。与LM组相比，LMN组cyclinD1和GSK-3β蛋白表达上调(P < 0.05)，LMI组细胞β-catenin和GSK-3β蛋白表达上调(P < 0.01)，LMC组细胞cyclinD1蛋白表达上调(P < 0.05)；与LM组相比，LMN组、LMI组和LMC组AKT磷酸化水平升高(P < 0.05)。以上结果说明，μ阿片受体和δ阿片受体均参与MENK对BESC的促增殖作用。

关键词: 甲硫脑啡肽, 奶牛子宫内膜基质细胞, 阿片受体, 阿片受体拮抗剂, 细胞增殖

Abstract: To clarify the mechanism of Met-enkephalin (MENK) on the proliferation of bovine endometrial stromal cell (BESC), the BESC were co-treated with lipopolysaccharide (LPS), opioid receptor antagonists and MENK. The opioid receptor antagonists used in this study included the nonselective opioid receptor antagonist naloxone (Nx), the δ opioid receptor antagonist ICI 154129 were the μ opioid receptor antagonist CTAP, and the groups were as follows: the blank control group, the LPS treatment group, the LPS and MENK (10^-10 mol·L^-1) co-treatment group (LM group), the LPS, MENK and opioid receptor antagonists (Nx, CTAP and ICI 154129 were all 10^-10 mol·L^-1) co-treatment group and the LPS and opioid receptor antagonist co-treatment groups. Cell viability, cell cycle, the expression of cell proliferation-related genes and the expression of Wnt/β-catenin and PI3K/AK pathway proteins were detected. The results showed that the treatment of opioid receptor antagonist alone in treatment concentration or co-treatment of LPS for 24 h had no effect on cell viability (P>0.05). Compared with the LM group, the cell numbers of G1 phase increased (P<0.05) in the co-treatment groups of LPS, MENK and Nx (LMN) and LPS, MENK and ICI 154129 (LMI), the cell numbers of S phase decreased (P<0.05) in LMN group and the co-treatment group of LPS, MENK and CTAP (LMC). Compared with LM group, the gene expressions of CCN2, TGFB1 and TGFB3 decreased (P<0.01) in the groups of LMN and LMI; the CCN2 expression decreased (P<0.01) in LMC group. Compared with LM group, there were increases in the protein expressions of cyclinD1 and GSK-3β in LMN group (P<0.05), the β-catenin and GSK-3β in LMC group (P<0.01) , and the cyclinD1 in LMC group (P<0.05) . Compared with LM group, the phosphorylation level of AKT increased (P<0.05) in LMN, LMI, and LMC groups. In conclusion, both δ opioid receptor and μ opioid receptor are involved in the proliferation of MENK on BESC.

Key words: Met-enkephalin, bovine endometrial stromal cell, opioid receptors, opioid receptor antagonist, proliferation

中图分类号:

S857.21

孙雯叶, 李建基, 王亨, 董俊升, 李俊, 崔璐莹. 甲硫脑啡肽通过阿片受体促进脂多糖作用下奶牛子宫内膜基质细胞的增殖[J]. 畜牧兽医学报, 2023, 54(3): 1229-1239.

SUN Wenye, LI Jianji, WANG Heng, DONG Junsheng, LI Jun, CUI Luying. Effect of Met-enkephalin on Lipopolysaccharide-stimulated Proliferation of Bovine Endometrial Stromal Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(3): 1229-1239.

参考文献 38

[1]	STOJKOV J, VON KEYSERLINGK M A G, MARCHANT-FORDE J N, et al. Assessment of visceral pain associated with metritis in dairy cows[J]. J Dairy Sci, 2015, 98(8):5352-5361.
[2]	RITTNER H L, BRACK A, STEIN C. Pain and the immune system[J]. Br J Anaesth, 2008, 101(1):40-44.
[3]	BALL R. Opioid peptides and psychiatric illness[J]. Br J Hosp Med, 1987, 37(1):49-50, 52.
[4]	SHAN F P, XIA Y J, WANG N, et al. Functional modulation of the pathway between dendritic cells (DCs) and CD4+T cells by the neuropeptide:methionine enkephalin (MENK)[J]. Peptides, 2011, 32(5):929-937.
[5]	ZAGON I S, VERDERAME M F, MCLAUGHLIN P J. The biology of the opioid growth factor receptor (OGFr)[J]. Brain Res Rev, 2002, 38(3):351-376.
[6]	TUO Y L, TIAN C, LU L L, et al. The paradoxical role of methionine enkephalin in tumor responses[J]. Eur J Pharmacol, 2020, 882:173253.
[7]	KAROUSSIOTIS C, MARTI-SOLANO M, STEPNIEWSKI T M, et al. A highly conserved δ-opioid receptor region determines RGS4 interaction[J]. FEBS J, 2020, 287(4):736-748.
[8]	ZHANG H, ZHOU D, GU J H, et al. Targeting the mu-opioid receptor for cancer treatment[J]. Curr Oncol Rep, 2021, 23(10):111.
[9]	MILAN-LOBO L, ENQUIST J, VAN RIJN R M, et al. Anti-analgesic effect of the mu/delta opioid receptor heteromer revealed by ligand-biased antagonism[J]. PLoS One, 2013, 8(3):e58362.
[10]	STEIN E A. Effects of intracranial self-stimulation on brain opioid peptides[J]. Peptides, 1985, 6(1):67-73.
[11]	SHEN D F, CAO X G, ZHAO L, et al. Naloxone ameliorates retinal lesions in Ccl2/Cx3cr1 double-deficient mice via modulation of microglia[J]. Investig Ophthalmol Vis Sci, 2011, 52(6):2897-2904.
[12]	LIN H Y, CHANG Y Y, KAO M C, et al. Naloxone inhibits nod-like receptor protein 3 inflammasome[J]. J Surg Res, 2017, 219:72-77.
[13]	LIANG L N, CHEN J L, LI Y, et al. Morphine and naloxone facilitate neural stem cells proliferation via a TET1-dependent and receptor-independent pathway[J]. Cell Rep, 2020, 30(11):3625-3631.e6.
[14]	JIANG Y L, HE X F, SHEN Y F, et al. Analgesic roles of peripheral intrinsic met-enkephalin and dynorphin A in long-lasting inflammatory pain induced by complete Freund's adjuvant in rats[J]. Exp Ther Med, 2015, 9(6):2344-2348.
[15]	ILLES P, PFEIFFER N, VON KVGELGEN I, et al. Presynaptic opioid receptor subtypes in the rabbit ear artery[J]. J Pharmacol Exp Ther, 1985, 232(2):526-533.
[16]	焦雪.蛋氨酸脑啡肽通过阿片受体调节CD8+T细胞活性机理的研究[D].沈阳:中国医科大学, 2019.JIAO X. Mechanism of regulation on CD8+T cells mediated by methionine enkephalin via binding opioid receptor[D]. Shenyang:China Medical University, 2019.(in Chinese)
[17]	孙雯叶,李建基,王亨,等.甲硫脑啡肽对脂多糖作用下奶牛子宫内膜基质细胞增殖的影响[J].中国兽医科学, 2022, 52(3):387-395.SUN W Y, LI J J, WANG H, et al. Effect of met-enkephalin on lipopolysaccharide-stimulated proliferation of bovine endometrial stromal cell[J]. Chinese Veterinary Science, 2022, 52(3):387-395.(in Chinese)
[18]	MARTIN-KLEINER I, BALOG T, GABRILOVAC J. Signal transduction induced by opioids in immune cells:a review[J]. Neuroimmunomodulation, 2006, 13(1):1-7.
[19]	MAO X F, AHSAN M Z, APRYANI E, et al. Dual μ-opioid receptor and norepinephrine reuptake mechanisms contribute to dezocine-and tapentadol-induced mechanical antiallodynia in cancer pain[J]. Eur J Pharmacol, 2020, 876:173062.
[20]	GADDIS R R, DIXON W R. Modulation of peripheral adrenergic neurotransmission by methionine-enkephalin[J]. J Pharmacol Exp Ther, 1982, 221(2):282-288.
[21]	LI Y, VAN DEN POL A N. μ-opioid receptor-mediated depression of the hypothalamic hypocretin/orexin arousal system[J]. J Neurosci, 2008, 28(11):2814-2819.
[22]	KENNEDY C, KRIER J.[Met5]enkephalin acts via δ-opioid receptors to inhibit pelvic nerve-evoked contractions of cat distal colon[J]. Br J Pharmacol, 1987, 92(2):291-298.
[23]	LAW P Y, WONG Y H, LOH H H. Molecular mechanisms and regulation of opioid receptor signaling[J]. Annu Rev Pharmacol Toxicol, 2000, 40:389-430.
[24]	SHELDON I M, CRONIN J, GOETZE L, et al. Defining postpartum uterine disease and the mechanisms of infection and immunity in the female reproductive tract in cattle[J]. Biol Reprod, 2009, 81(6):1025-1032.
[25]	SHELDON I M, CRONIN J G, HEALEY G D, et al. Innate immunity and inflammation of the bovine female reproductive tract in health and disease[J]. Reproduction, 2014, 148(3):R41-R51.
[26]	MATEUS L, DA COSTA L L, BERNARDO F, et al. Influence of puerperal uterine infection on uterine involution and postpartum ovarian activity in dairy cows[J]. Reprod Domest Anim, 2002, 37(1):31-35.
[27]	WANG Y Y, HANIFI-MOGHADDAM P, HANEKAMP E E, et al. Progesterone inhibition of Wnt/β-catenin signaling in normal endometrium and endometrial cancer[J]. Clin Cancer Res, 2009, 15(18):5784-5793.
[28]	DE BOER J, WANG H J, VAN BLITTERSWIJK C. Effects of Wnt signaling on proliferation and differentiation of human mesenchymal stem cells[J]. Tissue Eng, 2004, 10(3-4):393-401.
[29]	CHENG L, ZHAO Y, QI D, et al. Wnt/β-catenin pathway promotes acute lung injury induced by LPS through driving the Th17 response in mice[J]. Biochem Biophys Res Commun, 2018, 495(2):1890-1895.
[30]	刘新宁,曹滨滨,刘杰,等.阿魏酸对脂多糖诱导的OA软骨细胞Wnt/β-catenin信号通路的影响[J].河北医药, 2020, 42(11):1610-1614.LIU X N, CAO B B, LIU J, et al. Effects of ferulic acid on Wnt/β catenin signaling pathway of the OA chondrocytes induced by lipopolysaccharide[J]. Hebei Medical Journal, 2020, 42(11):1610-1614.(in Chinese)
[31]	SHI X, RAN L, LIU Y, et al. Knockdown of hnRNP A2/B1 inhibits cell proliferation, invasion and cell cycle triggering apoptosis in cervical cancer via PI3K/AKT signaling pathway[J]. Oncol Rep, 2018, 39(3):939-950.
[32]	邵帅,洪乐鹏,邓雪华,等.抑制PI3K/AKt信号通路对帕金森病炎症细胞模型反应的影响[J].中国老年学杂志, 2019, 39(9):2200-2203.SHAO S, HONG L P, DENG X H, et al. Effect of inhibition of PI3K/AKt signaling pathway on response in the inflammatory cell model of Parkinson's disease[J]. Chinese Journal of Gerontology, 2019, 39(9):2200-2203.(in Chinese)
[33]	李思翰,练东银,张广平,等.基于PI3K/Akt信号通路探讨金骨莲提取物抑制脂多糖诱导RAW264. 7细胞炎症反应的作用及机制[J].中国实验方剂学杂志, 2021, 27(14):29-35.LI S H, LIAN D Y, ZHANG G P, et al. Inhibitory effect and mechanism of Jingulian extract on LPS-induced RAW 264. 7 cell inflammatory response based on PI3K/AKT signaling pathway[J]. Chinese Journal of Experimental Traditional Medical Formulae, 2021, 27(14):29-35.(in Chinese)
[34]	BLALOCK J E. The immune system as the sixth sense[J]. J Intern Med, 2005, 257(2):126-138.
[35]	MALENDOWICZ L K, REBUFFAT P, TORTORELLA C, et al. Effects of met-enkephalin on cell proliferation in different models of adrenocortical-cell growth[J]. Int J Mol Med, 2005, 15(5):841-845.
[36]	HUCKLEBRIDGE F H, HUDSPITH B N, MUHAMED J, et al. Methionine-enkephalin stimulates in vitro proliferation of human peripheral lymphocytes via δ-opioid receptors[J]. Brain Behav Immun, 1989, 3(2):183-189.
[37]	TANG Z J, SHAO X, WU J, et al. Naloxone protects against lipopolysaccharide-induced neuroinflammation and microglial activation via inhibiting ATP-sensitive potassium channel[J]. Comput Math Methods Med, 2021, 2021:7731528.
[38]	JIAO X, WANG X N, WANG R Z, et al. Rules to activate CD8+T cells through regulating subunits of opioid receptors by methionine enkephalin (MENK)[J]. Int Immunopharmacol, 2018, 65:76-83.

甲硫脑啡肽通过阿片受体促进脂多糖作用下奶牛子宫内膜基质细胞的增殖

Effect of Met-enkephalin on Lipopolysaccharide-stimulated Proliferation of Bovine Endometrial Stromal Cells

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