IL-17对破骨细胞中MMP-9表达水平的影响及其意义

doi:10.13481/j.1671-587x.20160309

摘要/Abstract

摘要：

目的: 探讨白细胞介素17(IL-17)对破骨细胞中基质金属蛋白酶9(MMP-9)表达水平的影响,为研究IL-17在正畸力致牙根吸收过程中的作用提供依据。方法: 培养小鼠单核/巨噬系细胞RAW264.7,将细胞分为实验组和对照组,实验组细胞培养液中加入梯度浓度0.1、1.0、10.0和100.0μg·L^-1IL-17,作为0.1、1.0、10.0和100.0μg·L^-1IL-17组,对照组细胞培养液中不加入IL-17。对各组RAW264.7细胞进行破骨细胞诱导,诱导第5天时采用抗酒石酸酸性磷酸酶(TRAP)染色鉴定破骨细胞诱导成功。采用酶联免疫吸附(ELISA)法检测破骨细胞诱导第5天时各组RAW264.7细胞培养液中MMP-9表达水平;采用逆转录-聚合酶链式反应(RT-PCR)法检测破骨细胞诱导第5天时各组RAW264.7细胞培养液中MMP-9mRNA表达水平。结果: TRAP染色,RAW264.7细胞诱导第5天时可见TRAP染色阳性细胞,体积大,多核(细胞核≥3个)。ELISA和PT-PCR法检测,经破骨细胞诱导第5天时,与对照组比较,1.0、10.0和100.0μg·L^-1IL-17组RAW264.7细胞培养液中MMP-9和MMP-9mRNA表达水平升高(P<0.05);与1.0μg·L^-1IL-17组比较;10.0和100.0μg·L^-1IL-17组RAW264.7细胞培养液中MMP-9和MMP-9mRNA表达水平升高(P<0.05);与10.0μg·L^-1IL-17组比较,100μg·L^-1IL-17组RAW264.7细胞培养液中MMP-9和MMP-9mRNA表达水平升高(P<0.05)。结论: IL-17可以促进破骨细胞中MMP-9表达,且其促进作用随着IL-17浓度的升高而增强。

关键词: 白介素细胞17, 基质金属蛋白酶9, RAW264.7, 破骨细胞

Abstract:

Objective: To explore the effects of interleukin-17 (IL-17) on the expression levels of matrix metalloproteinase-9 (MMP-9) in the osteoclasts, and to provide the references for the study on the effect of IL-17 on the orthodontic force-induced root resorption. Methods: The mouse monocytes/macrophages RAW264.7 were cultured and divided into experiment group and control group; the culture medium in experiment group were added with the gradient concentrations of 0.1, 1.0, 10.0, and 100.0 μg·L^-1 IL-17, and set as 0.1,1.0,10.0, and 100.0 μg·L^-1 IL-17 groups,and the culture medium in control group was IL-17-free.The RAW264.7 cells in various groups were induced to form osteoclasts and identified by TRAP staining on the 5th day after induction. ELISA method was used to detect the expression levels of MMP-9 in the culture medium of RAW264.7 cells in various groups when the osteoclasts were induced for 5 d; RT-PCR method was used to detect the expression levels of MMP-9 mRNA in the culture medium of RAW264.7 cells in various groups when the osteoclasts were induced for 5 d. Results: The TRAP staining results showed TRAP staining positive cells were found, with large volume and large number of nucleus (nucleus≥3) on the 5th day after induction. The ELISA and RT-PCR results showed the expression levels of MMP-9 and MMP-9 mRNA in the culture medium of RAW264.7 cells in 1.0,10.0 and 100.0 μg·L^-1IL-17 groups were increased compared with control group(P<0.05); the expression levels of MMP-9 and MMP-9 mRNA in the culture medium of RAW264.7 cells in 10.0 and 100.0 μg·L^-1 IL-17 groups were increased compared with 1.0 μg·L^-1 IL-17 group(P<0.05); the expression levels of MMP-9 and MMP-9 mRNA in the culture medium of RAW264.7 cells in 100.0 μg·L^-1 IL-17 group were increased compared with 10.0 μg·L^-1 IL-17 group(P<0.05). Conclusion: IL-17 can promot the expressions of MMP-9 in the osteoclasts, and the promoting effect is enhanced with the increasing of IL-17 concentration.

Key words: interleukin-17, matrix metalloproteinase-9, RAW264.7, osteoclast

中图分类号:

R329.28

姚金丹, 韩光红, 胡敏. IL-17对破骨细胞中MMP-9表达水平的影响及其意义[J]. 吉林大学学报(医学版), 2016, 42(03): 462-466.

YAO Jindan, HAN Guanghong, HU Min. Effects of IL-17 on expressions of MMP-9 in osteoclasts and their significances[J]. Journal of Jilin University Medicine Edition, 2016, 42(03): 462-466.

参考文献

[1] Albaker BK, Wong RW. Diagnosis and management of root resorption by erupting canines using cone -beam computed tomography and fixed palatal appliance: a case report [J].J Med Case Rep, 2010, 4(1):399.

[2] 范瑞贤, 李晓智. 白细胞介素-17与正畸牙根吸收相关性研究进展[J]. 现代口腔医学杂志, 2013, 27 (3):167-169.

[3] Xu S, Cao X. Interleukin-17 and its expanding biological functions [J]. Cell Mol Immunol, 2010, 7(3):164-174.

[4] Hayashi N, Yamaguchi M, Nakajima R, et al. T-helper 17 cells mediate the osteo/odontoclastogenesis induced by excessive orthodontic forces [J]. Oral Dis, 2012, 18(4):375-388.

[5] Ota M, Yanagisawa M, Tachibana H, et al. A significant induction of neutrophilic chemoattractants but not RANKL in synoviocytes stimulated with interleukin 17[J]. J Bone Miner Metab, 2014, 33(1):40-47.

[6] Wucherpfennig AL, Li YP, Stetler-Stevenson WG, et al. Expression of 92 kD type IV collagenase/gelatinase B in human osteoclasts [J]. J Bone Miner Res, 2009, 9(4): 549- 556.

[7] Colnot C, Thompson Z, Miclau T, et al. Altered fracture repair in the absence of MMP9 [J]. Development, 2003, 130(17):4123-4133.

[8] 韩光红. 不同正畸力值对牙髓活力、Fos和MMP-9蛋白表达变化作用的实验研究[D].长春: 吉林大学, 2012.

[9] Cheng G, Wei L, Xiurong W, et al. IL-17 stimulates migration of carotid artery vascular smooth muscle cells in an MMP-9 dependent manner via p38 MAPK and ERK1/2-dependent NF-κB and AP-1 activation [J]. Cell Mol Neurobiol, 2009, 29(8):1161-1168.

[10] 张祎. 牙龈卟啉单胞菌脂多糖对成骨细胞-破骨细胞间EphB4-ephrinB2双向传导信号的影响[D]. 长春: 吉林大学, 2013.

[11] Haug SR, Brudvik P, Fristad I, et al. Sympathectomy causes increased root resorption after orthodontic tooth movement in rats: immunohistochemical study [J]. Cell Tissue Res, 2003, 313(2):167-175.

[12] Shimizu M, Yamaguchi M, Fujita S, et al. Interleukin-17/T-helper 17 cells in an atopic dermatitis mouse model aggravate orthodontic root resorption in dental pulp [J]. Eur J Oral Sci, 2013, 121(2): 101-110.

[13] Takahashi K, Azuma T, Motohira H, et al. The potential role of interleukin-17 in the immunopathology of periodontal disease [J]. J Clin Periodontal, 2005, 32(4): 369-374.

[14] Xiong H, Wei L, Peng B. Immunohistochemical localization of IL-17 in induced rat periapical lesions [J]. J Endod, 2009, 35(2):216-220.

[15] Yang S, Zhu L, Xiao L, et al. Imbalance of interleukin-17+ T-cell and Foxp3+ regulatory T-cell dynamics in rat periapical lesions [J]. J Endod, 2014, 40(1):52-62.

[16] Zhu L, Wu Y, IL-17R activation of human periodontal ligament fibroblasts induces IL-23 p19 production: Differential involvement of NF-κB versus JNK/AP-1 pathways [J]. Mol Immunol, 2011, 48(4):647-656.

[17] Ohba Y, Ohba T, Terai K, et al. Expression of cathepsin K mRNA during experimental tooth movement in rat as revealed by in situ hybridization [J]. Arch Oral Biol, 2000, 45(1):63-69.

[18] Oshiro T, Shibasaki Y, Martin TJ, et al. Immunolocalization of vacuolar-type H+-ATPase, cathepsin K, matrix metalloproteinase-9, and receptor activator of NFkappaB ligand in odontoclasts during physiological root resorption of human deciduous teeth [J]. Anat Rec, 2001, 264(3):305-311.

[19] Abreu RD, Penalva LO, Marcotte EM, et al. Global signatures of protein and mRNA expression levels [J]. Mol Biosyst, 2009, 5(12):1512-1526.