MicroRNA-136靶向CD163抑制CD68+CD163+M2型巨噬细胞极化的作用研究

doi:10.12092/j.issn.1009-2501.2019.01.001

中国临床药理学与治疗学 ›› 2019, Vol. 24 ›› Issue (1): 1-8.doi: 10.12092/j.issn.1009-2501.2019.01.001

• 基础研究 • 下一篇

MicroRNA-136靶向CD163抑制CD68⁺CD163⁺M2型巨噬细胞极化的作用研究

韩晨阳，杨毅，李文燕，王瑾，郭丽

嘉兴市第二医院，嘉兴 314001，浙江

收稿日期:2018-08-15 修回日期:2018-09-12 出版日期:2019-01-26 发布日期:2019-01-25
通讯作者: 郭丽，通信作者，女，硕士，主管技师，研究方向：肿瘤生物学。 E-mail：taishanglg@126.com
作者简介:韩晨阳，男，硕士，药师，研究方向：肿瘤药理学。 E-mail：691513770@qq.com
基金资助:
浙江省科技厅实验动物项目（2017C37174）；浙江省卫生厅面上项目（2018KY804）；嘉兴市科技计划项目（2018AY32007）

Inhibitory effect of MicroRNA-136 targeting CD163 on the polarization of CD68+CD163+M2 macrophages

HAN Chenyang, YANG Yi, LI Wenyang, WANG Jin, GUO Li

The Second Hospital of Jiaxing, Jiaxing 314001, Zhejiang, China

Received:2018-08-15 Revised:2018-09-12 Online:2019-01-26 Published:2019-01-25

摘要/Abstract

摘要：

目的: 研究微小RNA（miR-136）通过靶向CD163抑制CD68+M0巨噬细胞向CD68+CD163+M2巨噬细胞极化的作用机制。方法: 采用流式细胞术检测20例肝细胞肝癌患者肿瘤组织及癌旁组织中CD68+CD163+M2巨噬细胞的比例，实时荧光定量PCR（RT-qPCR）检测肿瘤及癌旁组织中miR-136的水平。分离人脾脏中CD68+M0巨噬细胞，将细胞分为对照组（Control），miRNA模拟物组（miRNA mimic）和miRNA抑制物组（miRNA inhibitor）。IL-4和IL-13 10 ng/mL诱导巨噬细胞M2型极化。采用流式细胞术检测CD68+CD163+M2巨噬细胞比例，Western blot检测细胞中CD163、TNF-α、诱导型一氧化氮合酶（iNOS）的表达以及M2巨噬细胞标志物精氨酸酶（Arg1）的表达，机制研究中检测JAK/STAT信号中蛋白酪氨酸激酶1（JAK1）、信号转导子与转录激活子1（STAT1）和STAT6的水平，PI3K/AKT信号中磷脂酰肌醇-3激酶（PI3K）、丝氨酸-苏氨酸蛋白激酶1（AKT1）的表达。荧光素酶报告基因法确定miR-136与CD163的靶向关系。结果: 肝癌组织中M2型巨噬细胞比例显著高于癌旁组织，而miR-136的表达显著低于癌旁组织，两者表达具有负相关。荧光素酶报告基因结果显示CD163是miR-136的靶基因。miRNA mimic组中CD68+CD163+M2巨噬细胞比例显著低于miRNA inhibitor组，与Control组比较无统计学差异。机制检测中发现miRNA mimic中JAK1、STAT6、PI3K、AKT1低表达，说明miR-136可以间接抑制JAK1-STAT6以及PI3K-AKT1的表达，抑制M2巨噬细胞极化。结论: miR-136可以靶向CD163抑制M2型巨噬细胞极化，其作用机制与JAK1-STAT6以及PI3K-AKT1抑制有关，这是M2巨噬细胞在肝癌免疫中的调节机制之一。

关键词: 微小RNA-136, CD163, M2型巨噬细胞, 荧光素酶报告基因

Abstract:

AIM: To study the mechanism of MicroRNA (miR-136) inhibiting the polarization of CD68+M0 macrophages to CD68+CD163+M2 macrophages by targeting CD163. METHODS: The ratio of CD68+CD163+M2 macrophages in the tumor tissues and adjacent tissues of 20 patients with hepatocellular carcinoma was detected by flow cytometry, and the level of miR-136 in the tumor and adjacent tissues was detected by real-time fluorescence quantitative PCR (RT-qPCR). CD68+M0 macrophages were separated from the spleen, and the cells were divided into the control group (Control), the miRNA analogue group (miRNA mimic) and the miRNA inhibitor group (miRNA inhibitor). IL-4 and IL-13 10 ng/mL were used to induce M2 polarization. Western blot was used to detect the expression of CD163, TNF-α, iNOS and Arg1 in M2 macrophage. In the mechanism study, the levels of JAK1, STAT1 and STAT6 in the JAK/STAT signal were detected, and the expressions of PI3K and AKT1 in the PI3K/AKT signal were detected also. Luciferase reporter gene method was used to determine the target relationship between miR-136 and CD163. RESULTS: The proportion of M2 macrophages in HCC tissues was significantly higher than that of adjacent tissues, while the expression of miR-136 was significantly lower than that of adjacent tissues, and the expression of the two was negatively correlated. Luciferase reporter gene results showed that CD163 was the target gene of miR-136. The percentage of CD68+CD163+M2 macrophages in miRNA mimic was significantly lower than that in miRNA inhibitor, and there was no significant difference compared with control group. The low expression of JAK1, STAT6, PI3K, AKT1 in miRNA mimic showed that miR-136 could inhibit the expression of JAK1-STAT6 and PI3K-AKT1 to inhibit the polarization of M2 macrophage. CONCLUSION: miR-136 can target CD163 to inhibit the M2 polarization of macrophage, and its mechanism is related to the inhibition of JAK1-STAT6 and PI3K-AKT1, which is one of the regulatory mechanisms of M2 macrophages in the immunization of liver cancer.

Key words: MicroRNA-136, CD163, M2 macrophages, luciferase reporter genes

中图分类号:

R965.2
R730

韩晨阳，杨毅，李文燕，王瑾，郭丽. MicroRNA-136靶向CD163抑制CD68⁺CD163⁺M2型巨噬细胞极化的作用研究[J]. 中国临床药理学与治疗学, 2019, 24(1): 1-8.

HAN Chenyang, YANG Yi, LI Wenyang, WANG Jin, GUO Li. Inhibitory effect of MicroRNA-136 targeting CD163 on the polarization of CD68+CD163+M2 macrophages[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2019, 24(1): 1-8.

参考文献

［1］Ravelli A, Minoia F, Davì S, et al. Development and initial validation of classification criteria for macrophage activation syndrome complicating systemic juvenile idiopathic arthritis［J］. Arth Rheumatol, 2015, 75(3): 481-495.
［2］Sindrilaru A, Peters T, Wieschalka S, et al. An unrestrained proinflammatory M1 macrophage population induced by iron impairs wound healing in humans and mice［J］. J Clin Invest, 2011, 121(3): 985-997.
［3］Qin H, Holdbrooks AT, Liu Y, et al. SOCS3 deficiency promotes M1 macrophage polarization and inflammation［J］. J Immunol, 2016, 189(7): 3439-3448.
［4］Cai X, Yin Y, Li N, et al. Re-polarization of tumor-associated macrophages to pro-inflammatory M1 macrophages by microRNA-155［J］. J Mol Cell Biol, 2012, 4(5): 341-343.
［5］Ren F, Fan M, Mei J, et al. Interferon-γ and celecoxib inhibit lung-tumor growth through modulating M2/M1 macrophage ratio in the tumor microenvironment［J］. Drug Design Devel Ther, 2014, 8(default):1527.
［6］Wang Y, Lin YX, Qiao SL, et al. Polymeric nanoparticles promote macrophage reversal from M2 to M1 phenotypes in the tumor microenvironment［J］. Biomaterials, 2017, 112: 153-163.
［7］Shabo I, Stal O, Olsson H, et al. Breast cancer expression of CD163, a macrophage scavenger receptor, is related to early distant recurrence and reduced patient survival［J］. Int J Cancer, 2008 123(4):780-786.
［8］Fujii N, Shomori K, Shiomi T, et al. Cancer-associated fibroblasts and CD163-positive macrophages in oral squamous cell carcinoma: their clinicopathological and prognostic significance［J］. J Oral Pathol Med, 2012, 41(6): 444-451.
［9］Capano S, Maione F, Casanovas O, et al. Zoledronic acid normalizes tumor vessels and blocks metastasis formation by skewing macrophages from a M2-to an M1-like phenotype in a mouse model of spontaneous cervical cancer［J］. Angiogenesis, 2014, 17(3): 723-724.
［10］Li R, Xing H, Lee TA, et al. Abstract A49: Tumor-associated interstitial flow promotes macrophage migration and pro-metastatic M2 phenotype in 3D ECM［J］. Cancer Res, 2017, 77(2 Supplement): A49-A49.
［11］Zheng J, Yang M, Shao J, et al. Chemokine receptor CX3CR1 contributes to macrophage survival in tumor metastasis.［J］. Mol Cancer, 2013, 12(1): 1-11.
［12］Yang M, Liu J, Piao C, et al. ICAM-1 suppresses tumor metastasis by inhibiting macrophage M2 polarization through blockade of efferocytosis［J］. Cell Death Dis, 2015, 6(6): e1780.
［13］Chen W, Wang J, Jia L, et al. Attenuation of the programmed cell death-1 pathway increases the M1 polarization of macrophages induced by zymosan［J］. Cell Death Dis, 2016, 7(2): e2115.
［14］Chen FY, Zhou J, Guo N, et al. Curcumin retunes cholesterol transport homeostasis and inflammation response in M1 macrophage to prevent atherosclerosis［J］. Biochem Biophysical Res Commun, 2015, 467(4): 872-878.
［15］Furudate S, Fujimura T, Kambayashi Y, et al. Comparison of CD163+CD206+M2 macrophages in the lesional skin of bullous pemphigoid and pemphigus vulgaris: the possible pathogenesis of bullous pemphigoid［J］. Dermatology, 2014, 229(4): 369-378.
［16］Fujimura T, Kambayashi Y, Furudate S, et al. Immunomodulatory effect of bisphosphonate risedronate sodium on CD163+ Arginase 1+ M2 macrophages: the development of a possible supportive therapy for angiosarcoma［J］. Clin Develop Immunol, 2014, 2013(1): 325412.
［17］魏续福, 蒲俊良, 郭振, 等. 肿瘤相关巨噬细胞促进索拉非尼耐药肝癌细胞的增殖及侵袭和迁移［J］. 细胞与分子免疫学杂志, 2017, 33(5): 617-622.
［18］O'Connell RM, Taganov KD, Boldin MP, et al. MicroRNA-155 is induced during the macrophage inflammatory response［J］. Proc Natl Acad Sci U S A, 2007, 104(5): 1604-1609.
［19］Chaudhuri AA, So AY, Sinha N, et al. MicroRNA-125b potentiates macrophage activation［J］. J Immunol, 2011, 187(10): 5062-5068.
［20］赵小彬，余陈欢，夏爱晓，等. 纳米载体特异性靶向TAMs治疗肿瘤的研究进展［J］. 中国临床药理学与治疗学, 2018, 23(7): 830-835.
［21］邢欣，李丽萍，王鹿，等. 丙型病毒性肝炎中肝脏巨噬细胞的变化及作用［J］. 中国临床药理学与治疗学, 2017, 22(9): 1072-1076.
［22］Liu H, Xie X, Yang X, et al. Enhanced inflammatory damage by microRNA-136 targeting Klotho expression in HK-2 cells by modulating JAK/STAT pathway［J］. Die Pharmazie, 2017, 72(5): 265-272.

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MicroRNA-136靶向CD163抑制CD68⁺CD163⁺M2型巨噬细胞极化的作用研究

Inhibitory effect of MicroRNA-136 targeting CD163 on the polarization of CD68+CD163+M2 macrophages

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