Abstract:

Objective: To investigate the effects of Lmo7 gene in the process of proliferation, migration and osteogenic differentiation of MC3T3-E1 cells. Methods: The preosteoblasts MC3T3-E1 cells were cultured in vitro with osteogenic induction. The relative expression of Lmo7 was detected by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting at 0, 3, 7 and 14 d respectively. At the same time, MC3T3-E1 cells were transfected with pLV-shLmo7 and pLV-shControl to construct the stably interfering cell line. Cell proliferation and cell migration were detected by CCK-8 assay, EdU assay, wound healing assay and Transwell migration assay. While alkaline phosphatase (ALP) staining was used to detect the expression activity of ALP, and RT-qPCR was used to detect the expression of osteogenesis-related genes such as Runt-related transcription factor 2 (Runx2), Osterix, alkaline phosphatase (ALP) and osteocalcin (OCN). Results: Lmo7 gene expression level was up-regulated during the osteogenic differentiation of MC3T3-E1 cells in vitro. The stable MC3T3-E1 cell line with Lmo7 interference was successfully constructed, and its proliferation activity was significantly improved, while its migration ability was inhibited. After osteogenic induction, compared with the control group, the ALP staining of the Lmo7 interfering cells was lighter, and the expression levels of osteogenesis-related genes such as Runx2, Osterix, ALP, and OCN were significantly down-regulated (P<0.05). Conclusion: Lmo7 interference promoted the cell proliferation but inhibited the cell migration. Furthermore, the osteogenic differentiation ability of cells was suppressed.

Key words: Lmo7, MC3T3-E1 cells, RNA interference

摘要：

目的：研究Lmo7基因对小鼠胚胎成骨细胞前体细胞(mouse embryo osteoblast precursor cells，MC3T3-E1 cells)增殖、迁移及成骨能力的影响。方法：体外培养MC3T3-E1细胞并对其进行成骨诱导，在0、3、7、14 d时收样，通过实时定量聚合酶链反应（real-time quantitative polymerase chain reaction，RT-qPCR）与蛋白质印迹法（Western blotting）验证Lmo7的表达变化。利用慢病毒转染pLV-shLmo7获得稳定干扰Lmo7 表达的细胞株，并通过CCK-8实验、EdU细胞增殖实验、细胞划痕实验、Transwell实验检测Lmo7对细胞增殖、迁移能力的影响。通过碱性磷酸酶（alkaline phosphatase，ALP）染色法检测ALP的表达活性，通过RT-qPCR检测成骨相关基因Runt相关转录因子2（Runt-related transcription factor 2，Runx2）、Osterix、ALP、骨钙素（osteocalcin，OCN）的表达。结果：Lmo7基因在成骨诱导的过程中表达上调。成功构建了Lmo7干扰的稳转细胞株，其增殖活性显著提高，而迁移能力受到抑制。对其进行成骨诱导后发现，与对照组相比，Lmo7干扰稳转株的ALP染色较浅，成骨相关基因Runx2、Osterix、ALP、OCN表达水平明显下调，且差异具有统计学意义（P<0.05）。结论：Lmo7低表达后可促进细胞增殖，抑制细胞迁移，下调Runx2、Osterix、ALP、OCN的表达能够抑制MC3T3-E1细胞体外成骨分化。

关键词: Lmo7, MC3T3-E1细胞, RNA干扰

CLC Number: 

• R782