Upregulating the Expression of LncRNA ANRIL Promotes Osteogenesis via the miR-7-5p/IGF-1R Axis in the Inflamed Periodontal Ligament Stem Cells.
Upregulating the Expression of LncRNA ANRIL Promotes Osteogenesis via the miR-7-5p/IGF-1R Axis in the Inflamed Periodontal Ligament Stem Cells.
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上调 LncRNA ANRIL 的表达通过 miR-7-5p/IGF-1R 轴促进发炎牙周膜干细胞的成骨
DOI：
10.3389/fcell.2021.604400
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发表时间：
2021
影响因子：
5.5
通讯作者：
Yu J
中科院分区：
文献类型：
--
作者：
Bian M;Yu Y;Li Y;Zhou Z;Wu X;Ye X;Yu J
INK 4基因座上的长非编码RNA（Long non-coding RNA，lncRNA）反义非编码RNA（ANRIL）是一种长约3.8kb的lncRNA，在细胞增殖、迁移和衰老等生物学功能中发挥重要作用。本研究确定了lncRNA ANRIL在炎症牙周膜干细胞（iPDLSC）衰老和成骨分化中的作用。方法分别从健康牙周膜组织和炎症牙周膜组织中分离健康牙周膜干细胞（hPDLSCs）和iPDLSCs。采用CCK-8法、EdU法和流式细胞术（FCM）检测细胞增殖能力。采用Western blot、实时荧光定量PCR、茜素红染色、碱性磷酸酶（ALP）染色、ALP活性检测和免疫荧光染色等方法检测lncRNA ANRIL对iPDLSCs的生物学效应。进行衰老相关（SA）-β-半乳糖苷酶（gal）染色、蛋白质印迹分析和qRT-PCR以确定细胞衰老。进行双荧光素酶报告基因测定以确认lncRNA ANRIL和miR-7-5-p以及miR-7- 5 p和胰岛素样生长因子受体（IGF-1 R）的结合。结果成功分离培养了HPDLSCs和iPDLSCs。与hPDLSC相比，iPDLSC中LncRNA ANRIL和IGF-1 R下降，而miR-7- 5 p上调。ANRIL的过表达增强了OSX、RUNX 2、ALP等成骨蛋白的表达，并下调了p16、p21、p53等衰老蛋白的表达。LncRNA ANRIL可通过海绵状表达miR-7- 5 p促进iPDLSCs定向分化。上调miR-7- 5 p抑制iPDLSC的成骨分化。进一步的分析确定IGF-1 R是miR-7- 5 p的直接靶点。通过双荧光素酶报告基因分析证实了lncRNA ANRIL与miR-7- 5 p、miR-7- 5 p和IGF-1 R的3′-UTR的直接结合。此外，拯救实验表明，miR-7- 5 p的敲低逆转了lncRNA ANRIL缺陷对iPDLSC成骨的抑制作用。结论lncRNA ANRIL通过调控miR-7- 5 p/IGF-1 R轴促进iPDLSCs向成骨细胞分化。
Background Long non-coding RNA (lncRNA) antisense non-coding RNA in the INK4 locus (ANRIL) is a base length of about 3.8 kb lncRNA, which plays an important role in several biological functions including cell proliferation, migration, and senescence. This study ascertained the role of lncRNA ANRIL in the senescence and osteogenic differentiation of inflamed periodontal ligament stem cells (iPDLSCs). Methods Healthy periodontal ligament stem cells (hPDLSCs) and iPDLSCs were isolated from healthy/inflamed periodontal ligament tissues, respectively. The proliferation abilities were determined by CCK-8, EdU assay, and flow cytometry (FCM). The methods of Western blot assay (WB), quantitative real-time polymerase chain reaction (qRT-PCR), alizarin red staining, alkaline phosphatase (ALP) staining, ALP activity detection, and immunofluorescence staining were described to determine the biological influences of lncRNA ANRIL on iPDLSCs. Senescence-associated (SA)-β-galactosidase (gal) staining, Western blot analysis, and qRT-PCR were performed to determine cell senescence. Dual-luciferase reporter assays were conducted to confirm the binding of lncRNA ANRIL and miR-7-5-p, as well as miR-7-5p and insulin-like growth factor receptor (IGF-1R). Results HPDLSCs and iPDLSCs were isolated and cultured successfully. LncRNA ANRIL and IGF-1R were declined, while miR-7-5p was upregulated in iPDLSCs compared with hPDLSCs. Overexpression of ANRIL enhanced the osteogenic protein expressions of OSX, RUNX2, ALP, and knocked down the aging protein expressions of p16, p21, p53. LncRNA ANRIL could promote the committed differentiation of iPDLSCs by sponging miR-7-5p. Upregulating miR-7-5p inhibited the osteogenic differentiation of iPDLSCs. Further analysis identified IGF-1R as a direct target of miR-7-5p. The direct binding of lncRNA ANRIL and miR-7-5p, miR-7-5p and the 3′-UTR of IGF-1R were verified by dual-luciferase reporter assay. Besides, rescue experiments showed that knockdown of miR-7-5p reversed the inhibitory effect of lncRNA ANRIL deficiency on osteogenesis of iPDLSCs. Conclusion This study disclosed that lncRNA ANRIL promotes osteogenic differentiation of iPDLSCs by regulating the miR-7-5p/IGF-1R axis.
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影响因子：
12.8
作者：
Liu GX;Ma S;Li Y;Yu Y;Zhou YX;Lu YD;Jin L;Wang ZL;Yu JH
通讯作者：
Yu JH
DOI：
10.1016/j.bbrc.2018.07.166
发表时间：
2018-09-10
影响因子：
3.1
作者：
Li, Jun;Qiu, Mingxing;Gong, Chunyu
通讯作者：
Gong, Chunyu
LncRNA H19通过miR-141/SPAG9通路促进根尖乳头干细胞定向分化
影响因子：
9
作者：
Li, Zehan;Yan, Ming;Yu, Jinhua
通讯作者：
Yu, Jinhua
影响因子：
3
作者：
Goetz, Werner;Heinen, Michael;Jaeger, Andreas
通讯作者：
Jaeger, Andreas
IGF-1/IGF-1R/hsa-let-7c轴调节根尖乳头干细胞的定向分化
影响因子：
4.6
作者：
Ma S;Liu G;Jin L;Pang X;Wang Y;Wang Z;Yu Y;Yu J
通讯作者：
Yu J