LncRNA MALAT1调节miR-383-5p/SOCS3轴对人牙周膜干细胞的影响

doi:10.13591/j.cnki.kqyx.2022.11.002

摘要/Abstract

摘要： 目的探究长链非编码RNA(long non-coding RNA,LncRNA)肺腺癌转移相关转录本1(metastasis-associated lung adenocarcinoma transcript 1,MALAT1)调控微小RNA-383-5p(microRNA-383-5p,miR-383-5p)/细胞因子信号转导负调控因子3(suppressor of cytokine signaling 3,SOCS3)轴对脂多糖(lipopolysaccharide,LPS)刺激的人牙周膜干细胞(human periodontal ligament stem cells,hPDLSCs)增殖、凋亡及成骨分化的影响。方法 LPS处理hPDLSCs后用靶向MALAT1的小干扰RNA(small interfering RNA-MALAT1,si-MALAT1)或miR-383-5p抑制物(miR-383-5p inhibitor,in-miR-383-5p)或SOCS3过表达质粒(SOCS3)转染,ELISA检测炎症因子水平;RT-qPCR和Western blot检测转染效率;CCK-8和TUNEL检测细胞增殖、凋亡;使用碱性磷酸酶(alkaline phosphatase,ALP)染色和茜素红S(alizarin red S,ARS)染色分析成骨分化;Western blot对增殖、凋亡以及成骨分化相关蛋白进行检测。生物信息学与双荧光素酶实验分析miR-383-5p与MALAT1/SOCS3之间的关系。结果 LPS可诱导hPDLSCs细胞炎症和凋亡,降低了细胞增殖能力和成骨分化程度;同时上调MALAT1和SOCS3表达,下调miR-383-5p表达(P<0.05)。敲低MALAT1可明显降低LPS诱导的hPDLSCs细胞炎症反应、凋亡能力,细胞增殖能力和成骨分化程度增加;同时还可上调miR-383-5p表达,下调SOCS3表达(P<0.05)。此外,回复实验表明,抑制miR-383-5p表达或上调SOCS3表达可部分逆转MALAT1敲低后对LPS诱导的hPDLSCs细胞的影响(P<0.05)。进一步分析验证,MALAT1海绵化miR-383-5p靶向调控SOCS3表达(P<0.05)。结论敲低MALAT1可通过调节miR-383-5p/SOCS3轴改善炎症状态下hPDLSCs的增殖、凋亡和成骨分化。

关键词: 长链非编码RNA肺腺癌转移相关转录本1, 微小RNA-383-5p, 细胞因子信号转导负调控因子3, 人牙周膜干细胞

Abstract: Objective To investigate the impacts of long non-coding RNA (LncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) on the proliferation, apoptosis and osteogenic differentiation of lipopolysaccharide (LPS)-stimulated human periodontal ligament stem cells (hPDLSCs) by regulating the microRNA-383-5p (miR-383-5p)/suppressor of cytokine signaling 3 (SOCS3) axis. Methods hPDLSCs were treated with LPS and transfected with small interfering RNA targeting MALAT1 (si-MALAT1) or miR-383-5p inhibitor (in-miR-383-5p) or SOCS3 overexpression plasmid (SOCS3). ELISA was implemented to detect inflammatory factor levels; RT-qPCR and Western blot were used to check transfection efficiency; CCK-8 and TUNEL were implemented to detect cell proliferation and apoptosis;alkaline phosphatase (ALP) staining and alizarin red S (ARS) staining were performed to analyze osteogenic differentiation; Western blot was implemented to measure the proliferation, apoptosis and osteogenic differentiation-related proteins. Bioinformatics and dual-luciferase experiments were performed to analyze the relationship between miR-383-5p and MALAT1/SOCS3. Results LPS induced the inflammation and apoptosis of hPDLSCs, and down-regulated cell proliferation and osteogenic differentiation. In the meantime, the expressions of MALAT1 and SOCS3 were up-regulated, and the expression of miR-383-5p was down-regulated (P<0.05).Knockdown of MALAT1 could greatly reduce LPS-induced inflammatory response and apoptosis of hPDLSCs, and increase cell proliferation and osteogenic differentiation. Meanwhile, it also up-regulated the expression of miR-383-5p and down-regulated the expression of SOCS3 (P<0.05). In addition, reverse experiments showed that inhibiting miR-383-5p expression or up-regulating SOCS3 expression could partially reverse the effects of MALAT1 knockdown on LPS-induced hPDLSCs (P<0.05). Further analysis confirmed that MALAT1 targeted the regulation of SOCS3 expression by sponging miR-383-5p (P<0.05). Conclusion Knockdown of MALAT1 can modulate the miR-383-5p/SOCS3 axis to improve the proliferation, apoptosis and osteogenic differentiation of inflammatory hPDLSCs.

Key words: long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1, microRNA-383-5p, suppressor of cytokine signaling 3, human periodontal ligament stem cells

中图分类号:

R781.4

顾婷立, 刘亚华, 钱靓, 章茜. LncRNA MALAT1调节miR-383-5p/SOCS3轴对人牙周膜干细胞的影响[J]. 口腔医学, 2022, 42(11): 966-973.

GU Tingli, LIU Yahua, QIAN Liang, ZHANG Qian. Impacts of LncRNA MALAT1 on human periodontal ligament stem cells by regulating miR-383-5p/SOCS3 axis[J]. Stomatology, 2022, 42(11): 966-973.

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