<em>PWAR6</em> interacts with miR‑106a‑5p to regulate the osteogenic differentiation of human periodontal ligament stem cells

Xiang,Juan; Bian,Ying

doi:10.3892/mmr.2021.11907

<em>PWAR6</em> interacts with miR‑106a‑5p to regulate the osteogenic differentiation of human periodontal ligament stem cells

Authors:
- Juan Xiang
- Ying Bian
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Affiliations: Department of Oral and Maxillofacial Surgery, Jingmen No. 1 People's Hospital, Jingmen, Hubei 448000, P.R. China
Published online on: February 8, 2021 https://doi.org/10.3892/mmr.2021.11907
Article Number: 268
Copyright: © Xiang et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Human periodontal ligament stem cells (hPDLSCs) associated with bone regeneration serve an important role in the treatment of periodontal disease. Long non‑coding RNAs are involved in the osteogenesis of multiple stem cells and can act as a sponge of microRNAs (miRs). The present study aimed to investigate the interaction between Prader Willi/Angelman region RNA 6 (PWAR6) and miR‑106a‑5p, as well as their influences on the osteogenic differentiation of hPDLSCs. hPDLSCs were isolated and cultured in osteogenic medium (OM) or growth medium (GM) for 7 days prior to transfection with PWAR6 overexpression vector, short hairpin RNA PWAR6 or miR‑106a‑5p mimic. The expression levels of runt‑related transcription factor 2, osteocalcin and bone morphogenetic protein 2 (BMP2) were detected by western blotting and reverse transcription‑quantitative PCR (RT‑qPCR), and the expression levels of PWAR6, miR‑106a‑5p and alkaline phosphatase (ALP) were determined by RT‑qPCR. ALP activity assays and Alizarin red staining were performed to detect osteogenesis and mineralization, respectively. Luciferase activities of wild‑type and mutant PWAR6 and BMP2 were assessed by conducting a dual‑luciferase reporter assay. The results indicated that PWAR6 expression was upregulated in OM‑incubated hPDLSCs compared with GM‑incubated hPDLSCs, and PWAR6 overexpression increased the osteogenic differentiation and mineralization of hPDLSCs compared with the corresponding control group. By contrast, miR‑106a‑5p expression was downregulated in OM‑incubated hPDLSCs compared with GM‑incubated hPDLSCs. PWAR6 acted as a sponge of miR‑106a‑5p and PWAR6 overexpression promoted the osteogenesis of miR‑106a‑5p mimic‑transfected hPDLSCs. BMP2 was predicted as a target gene of miR‑106a‑5p. Collectively, the results indicated that PWAR6 displayed a positive influence on the osteogenic differentiation of hPDLSCs. The results of the present study demonstrated that the PWAR6/miR‑106a‑5p interaction network may serve as a potential regulatory mechanism underlying hPDLSCs osteogenesis.

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1	Liu Y, Zeng X, Miao J, Liu C, Wei F, Liu D, Zheng Z, Ting K, Wang C and Guo J: Upregulation of long noncoding RNA MEG3 inhibits the osteogenic differentiation of periodontal ligament cells. J Cell Physiol. 234:4617–4626. 2019. View Article : Google Scholar : PubMed/NCBI
2	Pihlstrom BL, Michalowicz BS and Johnson NW: Periodontal diseases. Lancet. 366:1809–1820. 2005. View Article : Google Scholar : PubMed/NCBI
3	Xue P, Li B, An Y, Sun J, He X, Hou R, Dong G, Fei D, Jin F, Wang Q and Jin Y: Decreased MORF leads to prolonged endoplasmic reticulum stress in periodontitis-associated chronic inflammation. Cell Death Differ. 23:1862–1872. 2016. View Article : Google Scholar : PubMed/NCBI
4	Zhang H and Zhang D: Effects of periodontal ligament cells on alveolar bone metabolism under the action of force and inflammatory factors and its molecular mechanisms. Zhongguo Yi Xue Ke Xue Yuan Xue Bao. 39:432–437. 2017.PubMed/NCBI
5	Nagata M, Iwasaki K, Akazawa K, Komaki M, Yokoyama N, Izumi Y and Morita I: Conditioned medium from periodontal ligament stem cells enhances periodontal regeneration. Tissue Eng Part A. 23:367–377. 2017. View Article : Google Scholar : PubMed/NCBI
6	Liu N, Shi S, Deng M, Tang L, Zhang G, Liu N, Ding B, Liu W, Liu Y, Shi H, et al: High levels of β-catenin signaling reduce osteogenic differentiation of stem cells in inflammatory microenvironments through inhibition of the noncanonical Wnt pathway. J Bone Miner Res. 26:2082–2095. 2011. View Article : Google Scholar : PubMed/NCBI
7	Zhang J, Li ZG, Si YM, Chen B and Meng J: The difference on the osteogenic differentiation between periodontal ligament stem cells and bone marrow mesenchymal stem cells under inflammatory microenviroments. Differentiation. 88:97–105. 2014. View Article : Google Scholar : PubMed/NCBI
8	Jia B, Qiu X, Chen J, Sun X, Zheng X, Zhao J, Li Q and Wang Z: A feed-forward regulatory network lncPCAT1/miR-106a-5p/E2F5 regulates the osteogenic differentiation of periodontal ligament stem cells. J Cell Physiol. 234:19523–19538. 2019. View Article : Google Scholar : PubMed/NCBI
9	Wei F, Yang S, Guo Q, Zhang X, Ren D, Lv T and Xu X: MicroRNA-21 regulates osteogenic differentiation of periodontal ligament stem cells by targeting Smad5. Sci Rep. 7:166082017. View Article : Google Scholar : PubMed/NCBI
10	Yan GQ, Wang X, Yang F, Yang ML, Zhang GR, Wang GK and Zhou Q: MicroRNA-22 promoted osteogenic differentiation of human periodontal ligament stem cells by targeting HDAC6. J Cell Biochem. 118:1653–1658. 2017. View Article : Google Scholar : PubMed/NCBI
11	Gu X, Li M, Jin Y, Liu D and Wei F: Identification and integrated analysis of differentially expressed lncRNAs and circRNAs reveal the potential ceRNA networks during PDLSC osteogenic differentiation. BMC Genet. 18:1002017. View Article : Google Scholar : PubMed/NCBI
12	Ugawa Y, Yamamoto T, Kawamura M, Yamashiro K, Shimoe M, Tomikawa K, Hongo S, Maeda H and Takashiba S: Rho-kinase regulates extracellular matrix-mediated osteogenic differentiation of periodontal ligament cells. Cell Biol Int. 41:651–658. 2017. View Article : Google Scholar : PubMed/NCBI
13	Kang W, Liang Q, Du L, Shang L, Wang T and Ge S: Sequential application of bFGF and BMP-2 facilitates osteogenic differentiation of human periodontal ligament stem cells. J Periodontal Res. 54:424–434. 2019. View Article : Google Scholar : PubMed/NCBI
14	Lee JS, Lee JC and Heo JS: Polydopamine-assisted BMP-2 immobilization on titanium surface enhances the osteogenic potential of periodontal ligament stem cells via integrin-mediated cell-matrix adhesion. J Cell Commun Signal. 12:661–672. 2018. View Article : Google Scholar : PubMed/NCBI
15	Cao F, Zhan J, Chen X, Zhang K, Lai R and Feng Z: miR-214 promotes periodontal ligament stem cell osteoblastic differentiation by modulating Wnt/β-catenin signaling. Mol Med Rep. 16:9301–9308. 2017. View Article : Google Scholar : PubMed/NCBI
16	Li S, Shao J, Zhou Y, Friis T, Yao J, Shi B and Xiao Y: The impact of Wnt signalling and hypoxia on osteogenic and cementogenic differentiation in human periodontal ligament cells. Mol Med Rep. 14:4975–4982. 2016. View Article : Google Scholar : PubMed/NCBI
17	Chen LJ, Hu BB, Shi XL, Ren MM, Yu WB, Cen SD, Hu RD and Deng H: Baicalein enhances the osteogenic differentiation of human periodontal ligament cells by activating the Wnt/β-catenin signaling pathway. Arch Oral Biol. 78:100–108. 2017. View Article : Google Scholar : PubMed/NCBI
18	Wang L, Wu F, Song Y, Li X, Wu Q, Duan Y and Jin Z: Long noncoding RNA related to periodontitis interacts with miR-182 to upregulate osteogenic differentiation in periodontal mesenchymal stem cells of periodontitis patients. Cell Death Dis. 7:e23272016. View Article : Google Scholar : PubMed/NCBI
19	Zhang J, Wang P, Wan L, Xu S and Pang D: The emergence of noncoding RNAs as Heracles in autophagy. Autophagy. 13:1004–1024. 2017. View Article : Google Scholar : PubMed/NCBI
20	Cech TR and Steitz JA: The noncoding RNA revolution-trashing old rules to forge new ones. Cell. 157:77–94. 2014. View Article : Google Scholar : PubMed/NCBI
21	Sharp PA: The centrality of RNA. Cell. 136:577–580. 2009. View Article : Google Scholar : PubMed/NCBI
22	Zhuang W, Ge X, Yang S, Huang M, Zhuang W, Chen P, Zhang X, Fu J, Qu J and Li B: Upregulation of lncRNA MEG3 promotes osteogenic differentiation of mesenchymal stem cells from multiple myeloma patients by targeting BMP4 transcription. Stem Cells. 33:1985–1997. 2015. View Article : Google Scholar : PubMed/NCBI
23	Zhang W, Dong R, Diao S, Du J, Fan Z and Wang F: Differential long noncoding RNA/mRNA expression profiling and functional network analysis during osteogenic differentiation of human bone marrow mesenchymal stem cells. Stem Cell Res Ther. 8:302017. View Article : Google Scholar : PubMed/NCBI
24	Liao J, Yu X, Hu X, Fan J, Wang J, Zhang Z, Zhao C, Zeng Z, Shu Y, Zhang R, et al: lncRNA H19 mediates BMP9-induced osteogenic differentiation of mesenchymal stem cells (MSCs) through Notch signaling. Oncotarget. 8:53581–53601. 2017. View Article : Google Scholar : PubMed/NCBI
25	Memczak S, Jens M, Elefsinioti A, Torti F, Krueger J, Rybak A, Maier L, Mackowiak SD, Gregersen LH, Munschauer M, et al: Circular RNAs are a large class of animal RNAs with regulatory potency. Nature. 495:333–338. 2013. View Article : Google Scholar : PubMed/NCBI
26	Hansen TB, Jensen TI, Clausen BH, Bramsen JB, Finsen B, Damgaard CK and Kjems J: Natural RNA circles function as efficient microRNA sponges. Nature. 495:384–388. 2013. View Article : Google Scholar : PubMed/NCBI
27	Beermann J, Piccoli MT, Viereck J and Thum T: Non-coding RNAs in development and disease: Background, mechanisms, and therapeutic approaches. Physiol Rev. 96:1297–1325. 2016. View Article : Google Scholar : PubMed/NCBI
28	Mozaffari SV, Stein MM, Magnaye KM, Nicolae DL and Ober C: Parent of origin gene expression in a founder population identifies two new candidate imprinted genes at known imprinted regions. PLoS One. 13:e02039062018. View Article : Google Scholar : PubMed/NCBI
29	Lei M, Mitsuhashi S, Miyake N, Ohta T, Liang D, Wu L and Matsumoto N: Translocation breakpoint disrupting the host SNHG14 gene but not coding genes or snoRNAs in typical Prader-Willi syndrome. J Hum Genet. 64:647–652. 2019. View Article : Google Scholar : PubMed/NCBI
30	Lin X, Jiang T, Bai J, Li J, Wang T, Xiao J, Tian Y, Jin X, Shao T, Xu J, et al: Characterization of transcriptome transition associates long noncoding RNAs with glioma progression. Mol Ther Nucleic Acids. 13:620–632. 2018. View Article : Google Scholar : PubMed/NCBI
31	Du L, Yang P and Ge S: Isolation and characterization of human gingiva-derived mesenchymal stem cells using limiting dilution method. J Dent Sci. 11:304–314. 2016. View Article : Google Scholar : PubMed/NCBI
32	Wang T, Kang W, Du L and Ge S: Rho-kinase inhibitor Y-27632 facilitates the proliferation, migration and pluripotency of human periodontal ligament stem cells. J Cell Mol Med. 21:3100–3112. 2017. View Article : Google Scholar : PubMed/NCBI
33	Li JH, Liu S, Zhou H, Qu LH and Yang JH: starBase v2.0: Decoding miRNA-ceRNA, miRNA-ncRNA and protein-RNA interaction networks from large-scale CLIP-Seq data. Nucleic Acids Res. 42((Database Issue)): D92–D97. 2014. View Article : Google Scholar : PubMed/NCBI
34	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
35	Bae WJ, Shin MR, Kang SK, Zhang-Jun, Kim JY, Lee SC and Kim EC: HIF-2 inhibition supresses inflammatory responses and osteoclastic differentiation in human periodontal ligament cells. J Cell Biochem. 116:1241–1255. 2015. View Article : Google Scholar : PubMed/NCBI
36	Wu XS, Wang F, Li HF, Hu YP, Jiang L, Zhang F, Li ML, Wang XA, Jin YP, Zhang YJ, et al: LncRNA-PAGBC acts as a microRNA sponge and promotes gallbladder tumorigenesis. EMBO Rep. 18:1837–1853. 2017. View Article : Google Scholar : PubMed/NCBI
37	Ballantyne MD, McDonald RA and Baker AH: lncRNA/MicroRNA interactions in the vasculature. Clin Pharmacol Ther. 99:494–501. 2016. View Article : Google Scholar : PubMed/NCBI
38	Wang K, Jin W, Song Y and Fei X: LncRNA RP11-436H11.5, functioning as a competitive endogenous RNA, upregulates BCL-W expression by sponging miR-335-5p and promotes proliferation and invasion in renal cell carcinoma. Mol Cancer. 16:1662017. View Article : Google Scholar : PubMed/NCBI
39	Kallen AN, Zhou XB, Xu J, Qiao C, Ma J, Yan L, Lu L, Liu C, Yi JS, Zhang H, et al: The imprinted H19 lncRNA antagonizes let-7 microRNAs. Mol Cell. 52:101–112. 2013. View Article : Google Scholar : PubMed/NCBI
40	Yu C, Li L, Xie F, Guo S, Liu F, Dong N and Wang Y: LncRNA TUG1 sponges miR-204-5p to promote osteoblast differentiation through upregulating Runx2 in aortic valve calcification. Cardiovasc Res. 114:168–179. 2018. View Article : Google Scholar : PubMed/NCBI
41	Kim J, Abdelmohsen K, Yang X, De S, Grammatikakis I, Noh JH and Gorospe M: LncRNA OIP5-AS1/cyrano sponges RNA-binding protein HuR. Nucleic Acids Res. 44:2378–2392. 2016. View Article : Google Scholar : PubMed/NCBI
42	Guo L, Zhao RC and Wu Y: The role of microRNAs in self-renewal and differentiation of mesenchymal stem cells. Exp Hematol. 39:608–616. 2011. View Article : Google Scholar : PubMed/NCBI
43	Zhang Z, Liu J, Zeng Z, Fan J, Huang S, Zhang L, Zhang B, Wang X, Feng Y, Ye Z, et al: lncRNA Rmst acts as an important mediator of BMP9-induced osteogenic differentiation of mesenchymal stem cells (MSCs) by antagonizing Notch-targeting microRNAs. Aging (Albany NY). 11:12476–12496. 2019. View Article : Google Scholar : PubMed/NCBI
44	Seenprachawong K, Nuchnoi P, Nantasenamat C, Prachayasittikul V and Supokawej A: Computational identification of miRNAs that modulate the differentiation of mesenchymal stem cells to osteoblasts. PeerJ. 4:e19762016. View Article : Google Scholar : PubMed/NCBI
45	Vimalraj S and Selvamurugan N: MicroRNAs expression and their regulatory networks during mesenchymal stem cells differentiation toward osteoblasts. Int J Biol Macromol. 66:194–202. 2014. View Article : Google Scholar : PubMed/NCBI
46	Pan YJ, Wei LL, Wu XJ, Huo FC, Mou J and Pei DS: MiR-106a-5p inhibits the cell migration and invasion of renal cell carcinoma through targeting PAK5. Cell Death Dis. 8:e31552017. View Article : Google Scholar : PubMed/NCBI
47	Hai B, Ma Y, Pan X, Yong L, Liang C, He G, Yang C, Zhu B and Liu X: Melatonin benefits to the growth of human annulus fibrosus cells through inhibiting miR-106a-5p/ATG7 signaling pathway. Clin Interv Aging. 14:621–630. 2019. View Article : Google Scholar : PubMed/NCBI
48	Hao W, Liu H, Zhou L, Sun Y, Su H, Ni J, He T, Shi P and Wang X: MiR-145 regulates osteogenic differentiation of human adipose-derived mesenchymal stem cells through targeting FoxO1. Exp Biol Med (Maywood). 243:386–393. 2018. View Article : Google Scholar : PubMed/NCBI
49	Zhu S, Peng W, Li X, Weng J, Zhang X, Guo J, Huang D, Rong Q and Chen S: miR-1827 inhibits osteogenic differentiation by targeting IGF1 in MSMSCs. Sci Rep. 7:461362017. View Article : Google Scholar : PubMed/NCBI
50	Wang H, Cui Y, Luan J, Zhou X, Li C, Li H, Shi L and Han J: MiR-5100 promotes osteogenic differentiation by targeting Tob2. J Bone Miner Metab. 35:608–615. 2017. View Article : Google Scholar : PubMed/NCBI
51	Zhou N, Li Q, Lin X, Hu N, Liao JY, Lin LB, Zhao C, Hu ZM, Liang X, Xu W, et al: BMP2 induces chondrogenic differentiation, osteogenic differentiation and endochondral ossification in stem cells. Cell Tissue Res. 366:101–111. 2016. View Article : Google Scholar : PubMed/NCBI
52	Sun MH, Wang WJ, Li Q, Yuan T and Weng WJ: Autologous oxygen release nano bionic scaffold composite miR-106a induced BMSCs enhances osteoblast conversion and promotes bone repair through regulating BMP-2. Eur Rev Med Pharmacol Sci. 22:7148–7155. 2018.PubMed/NCBI
53	Park SH, Kwon JS, Lee BS, Park JH, Lee BK, Yun JH, Lee BY, Kim JH, Min BH, Yoo TH and Kim MS: BMP2-modified injectable hydrogel for osteogenic differentiation of human periodontal ligament stem cells. Sci Rep. 7:66032017. View Article : Google Scholar : PubMed/NCBI
54	Li H, Li T, Wang S, Wei J, Fan J, Li J, Han Q, Liao L, Shao C and Zhao RC: miR-17-5p and miR-106a are involved in the balance between osteogenic and adipogenic differentiation of adipose-derived mesenchymal stem cells. Stem Cell Res. 10:313–324. 2013. View Article : Google Scholar : PubMed/NCBI
55	Oliveira OR, Martins SP, Lima WG and Gomes MM: The use of bone morphogenetic proteins (BMP) and pseudarthrosis, a literature review. Rev Bras Ortop. 52:124–140. 2016. View Article : Google Scholar : PubMed/NCBI
56	Bais MV, Wigner N, Young M, Toholka R, Graves DT, Morgan EF, Gerstenfeld LC and Einhorn TA: BMP2 is essential for post natal osteogenesis but not for recruitment of osteogenic stem cells. Bone. 45:254–266. 2009. View Article : Google Scholar : PubMed/NCBI