不同浓度白介素8对破骨细胞MMP-9表达的影响
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摘要
目的:观察不同浓度的白介素8(IL-8)对体外培养的破骨细胞中基质金属蛋白酶9(MMP-9)表达水平的影响。方法:体外培养小鼠单核巨噬细胞RAW264.7后,用浓度为50ng/L的RANKL将其诱导成为破骨细胞。在诱导后第5天时,进行细胞形态观察,以及抗酒石酸酸性磷酸酶(TRAP)染色,以鉴定破骨细胞诱导成功。然后分为a、b、c、d、e组,其中a组为空白对照组,不加IL-8; b、c、d、e组分别加入0.1、1.0、10、100ng/ml的IL-8,分别作用24h、48h、72h后,通过酶联免疫吸附(ELISA)法及实时定量PCR(q PCR)检测细胞中MMP-9的表达水平。结果:TRAP染色结果:可见蓝紫色胞核,胞浆内有棕红色沉淀。ELISA检测结果显示:48h组和72h组中,与a组(对照组)相比,b、c、d、e组MMP-9表达水平均升高,组间差异有统计学意义(P<0.05)。q PCR检测结果显示:在同一时间下,与对照组相比,b、c、d、e组中的MMP-9mRNA表达水平均升高,组间差异具有统计学意义(P<0.05),同一浓度下,表达水平72h组>48h组>24h组,且差异具有统计学意义(P<0.05)。结论:IL-8可以上调破骨细胞中MMP-9的表达,并具有浓度依赖性,为研究IL-8在正畸力致牙根吸收过程中的作用提供参考。
Objective: To observe the effect of different concentrations of interleukin 8( IL-8) on the expression of MMP-9 protein in osteoclasts cultured in vitro.Methods: After culturing the mononuclear macrophage RAW264.7 of mice,the RANKL was induced into osteoclasts by the concentration of 50 ng/L. At fifth days after induction,cell morphology and tartrate resistant acid phosphatase( TRAP) staining were performed to identify osteoclast induction success. Then divided into a,b,c,d and e groups,a group was the control group,without IL-8; b,c,d,e groups were added 0.1,1,10 and 100 ng/ml of IL-8,and after the action of 24 h,48 h and 72 h,the expression level of MMP-9 in cell culture fluid was detected by enzyme-linked immunosorbent assay( ELISA) and q PCR. Results: TRAP staining showed that the nucleus of blue purple cells was red brown.The results of ELISA showed that the expression of MMP-9 in b,c,d and e groups was significantly higher in the 48 h group and the 72 h group than in the a group( control group). The difference between the two groups was statistically significant( P<0.05). The results of q PCR showed that at the same time,the expression of MMP-9 m RNA in the b,c,d,e groups was significantly higher than that in the a group( control group). The difference between the two groups was statistically significant( P<0.05). At the same concentration,the expression level was 72 h group>48 h group> 24 h group,and the difference was statistically significant( P<0.05).Conclusions: IL-8 can upregulate the expression of MMP-9 in osteoclasts,and is a concentration dependent manner,providing a reference for studying the role of IL-8 in orthodontic force induced root resorption.
Objective: To observe the effect of different concentrations of interleukin 8( IL-8) on the expression of MMP-9 protein in osteoclasts cultured in vitro.Methods: After culturing the mononuclear macrophage RAW264.7 of mice,the RANKL was induced into osteoclasts by the concentration of 50 ng/L. At fifth days after induction,cell morphology and tartrate resistant acid phosphatase( TRAP) staining were performed to identify osteoclast induction success. Then divided into a,b,c,d and e groups,a group was the control group,without IL-8; b,c,d,e groups were added 0.1,1,10 and 100 ng/ml of IL-8,and after the action of 24 h,48 h and 72 h,the expression level of MMP-9 in cell culture fluid was detected by enzyme-linked immunosorbent assay( ELISA) and q PCR. Results: TRAP staining showed that the nucleus of blue purple cells was red brown.The results of ELISA showed that the expression of MMP-9 in b,c,d and e groups was significantly higher in the 48 h group and the 72 h group than in the a group( control group). The difference between the two groups was statistically significant( P<0.05). The results of q PCR showed that at the same time,the expression of MMP-9 m RNA in the b,c,d,e groups was significantly higher than that in the a group( control group). The difference between the two groups was statistically significant( P<0.05). At the same concentration,the expression level was 72 h group>48 h group> 24 h group,and the difference was statistically significant( P<0.05).Conclusions: IL-8 can upregulate the expression of MMP-9 in osteoclasts,and is a concentration dependent manner,providing a reference for studying the role of IL-8 in orthodontic force induced root resorption.
引文
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[2]程旭锋,张新峰,刘琦,等.药对“附子-白术”对裸鼠乳腺癌骨转移NF-κB通路的影响[J].中华中医药学刊,2017,35(11):2773-2776.
[3]孙志新,张云涛.IL-1β对人牙龈成纤维细胞在不同钛板表面表达炎性因子的影响[J].上海口腔医学,2015,24(02):151-156.
[4]胡鹏,盖兰珍.正畸治疗中牙根吸收的相关因素分析[J].现代医药卫生,2012,28(8):1205-1207.
[5]鲍艳举.消癥止痛外用方调控破骨细胞活化及PAR2/TRPV1信号通路治疗骨癌痛的作用机制研究[D].中国中医科学院,2015.
[6]李志厂,徐国成.白介素-8在儿童过敏性紫癜中的研究进展[J].齐齐哈尔医学院学报,2015,36(27):4155-4157.
[7]陈雪.氧化性低密度脂蛋白对破骨细胞增殖及分化的影响[D].泰山医学院,2014.
[8]艾承冲,蒋佳,陈世益.髋关节假体周围骨质溶解的生物学机制[J].中华骨科杂志,2017,37(7):441-448.
[9]张媛媛,何嘉承,林煜,张文明,黄云梅,吴银生,陈翔,贾晓康,林燕萍.健骨颗粒含药血清对体外破骨细胞CAⅡ、CK、MMP-9mRNA表达的影响[J].中国骨质疏松杂志,2015,21(12):-收稿日期:--1436-1440.