10.6084/M9.FIGSHARE.21546089
Lintao Li
Lintao
Li
Beiyue Wang
Beiyue
Wang
Xing Zhou
Xing
Zhou
Hao Ding
Hao
Ding
Chang Sun
Chang
Sun
Yicun Wang
Yicun
Wang
Fan Zhang
Fan
Zhang
Jianning Zhao
Jianning
Zhao
METTL3-mediated long non-coding RNA MIR99AHG methylation targets miR-4660 to promote bone marrow mesenchymal stem cell osteogenic differentiation
<p>Whether long non-coding RNA Mir-99a-Let-7c Cluster Host Gene (LncRNA MIR99AHG) is involved in osteoporosis (OP) remains vague, so we hereby center on its implication. Old C57BL/6J mice were injected with the silencing lentivirus of MIR99AHG and subjected to microCT analysis and immunohistochemistry on osteogenic cells. The osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) with or without transfection was determined by alkaline phosphatase (ALP) and Alizarin Red S staining. Total N(6)-methyladenosine (m<sup>6</sup>A) on the bone marrow mesenchymal stem cells (BMSCs) was quantified. The potential methylation site and the complementary binding sites with candidate microRNA (miR) were predicted via bioinformatic analyses, with the latter being confirmed via dual-luciferase reporter, RNA immunoprecipitation and RNA pull-down assays. Quantitative real-time PCR and Western blot were used for quantification assays. MIR99AHG was decreased during the osteogenic differentiation of BMSCs, where increased Osterix (OSX), Collagen, Type I, Alpha 1 (Col1A1), Osteocalcin (OCN) and RUNX Family Transcription Factor 2 (RUNX2) as well as more color-stained areas were found. Also, silencing MIR99AHG relieved the OP in mice and reduced the loss of osteogenic cells. M<sup>6</sup>A methylation in undifferentiated BMSCs was low and MIR99AHG overexpression abolished the effects of overexpressed METTL3 on promoting osteogenic differentiation. MiR-4660, which was downregulated in BMSCs without differentiation but increased during osteogenic differentiation, could bind with MIR99AHG. Furthermore, miR-4660 promoted osteogenic differentiation and reversed the effects of overexpressed MIR99AHG. The present study demonstrated that METTL3-mediated LncRNA MIR99AHG methylation enhanced the osteogenic differentiation of BMSCs via targeting miR-4660.</p>
Medicine
Cell Biology
Genetics
Molecular Biology
Physiology
Ecology
Immunology
Biological Sciences not elsewhere classified
Developmental Biology
Taylor & Francis
2022
2022-11-12
2023-05-31
Dataset
12069297 Bytes
10.1080/15384101.2022.2125751
CC BY 4.0