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| Raji-MG63细胞共培养体系中心肌营养蛋白样细胞因子1（CLCF1）对RANKL/OPG
比值及成骨细胞分化的影响 |
| The effect of cardiotrophin-like cytokine 1 (CLCF1) on the RANKL/OPG ratio and osteoblast differentiation in Raji-MG63 cell co-culture system |
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| DOI：10.3969/j.issn.1006-7108.2021.02.001 |
| 中文关键词: CLCF1 RANKL/OPG JAK2/STAT3通路 成骨分化 骨免疫 |
| 英文关键词:CLCF1 RANKL/OPG JAK2/STAT3 osteogenic differentiation bone immunity |
| 基金项目:福建省自然科学基金（2017J01535）；福建省卫生计生科研人才培养项目（2018-ZQN-72）；国家自然科学基金（81674007） |
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| 中文摘要: |
| 目的 探讨Raji-MG63细胞共培养体系中，心肌营养蛋白样细胞因子1（cardiotrophin-like cytokine
factor 1，CLCF1）基因对骨保护素（osteoprotegerin，OPG）、核因子κB受体活化因子配体(receptor activator
of NF-κB ligand，RANKL）和成骨细胞分化的影响。方法 通过CRISPR/Cas9基因编辑技术构建CLCF1
基因敲除Raji细胞系，实验分2组：对照组（Control组）和CLCF1敲除组（CLCF1-KO组）。应用Transwell
技术建立Raji-MG63细胞共培养体系，通过蛋白质印迹法（western blot，WB）检测CLCF1、OPG、RANKL
及JAK2/STAT3蛋白的变化。运用碱性磷酸酶（ALP）活性检测和茜素红染色观察成骨细胞分化能力。结
果 WB结果显示，与对照组比较，CLCF1敲除组OPG（P<0.01）蛋白明显下调，RANKL/OPG
比值升高（P<0.001），磷酸化JAK2（P<0.001）和STAT3（P<0.01）蛋白明显下调，差异具有统计学意
义；ALP活性检测和茜素红染色结果显示，Raji-MG63细胞共培养体系中，CLCF1敲除组MG63细胞ALP
活性和矿化形成能力降低，与对照组比较差异具有统计学意义（P<0.001）。结论 CLCF1基因能通过调控
RANKL/OPG比值和JAK2/STAT3通路影响成骨细胞分化。 |
| 英文摘要: |
| Objective To explore the effect of inhibiting the expression of Cardiotrophin-Like Cytokine Factor 1 (CLCF1) gene in Raji-MG63 cell co-culture system on osteoprotegerin (OPG) and nuclear factor-κB receptor activator factor ligand (receptor activator of NF-κB ligand, RANKL) and osteoblast differentiation. Methods The CLCF1 knockout Raji cell line was constructed by CRISPR/Cas9 gene editing technology. The experiment was divided into 2 groups: control group (Ctrl group) and CLCF1 knockout group (CLCF1-KO group). Transwell technology was used to establish a Raji-MG63 cell co-culture system, and Western blot (WB) was used to detect the changes of CLCF1, OPG, RANKL and JAK2/STAT3 proteins. Alkaline phosphatase (ALP) activity detection and alizarin red staining were used to observe the differentiation ability of osteoblasts. Results Western blot results showed that compared with the control group, the OPG (P<0.01) protein in the CLCF1 knockout group was significantly down-regulated, the RANKL/OPG ratio increased (P<0.001), and JAK2 (P<0.001) and STAT3 (P<0.01) phosphorylated The protein was significantly down-regulated, and the difference was statistically significant; the results of ALP activity detection and Alizarin Red staining showed that the Raji-MG63 cell co-culture system, the ALP activity and mineralization ability of MG63 cells in the CLCF1 knockout group were reduced, which was different from the control Statistically significant (P<0.001). Conclusion CLCF1 gene can affect osteoblast differentiation by regulating RANKL/OPG ratio and JAK2/STAT3 pathway. |
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