Inhibition of miR-29c promotes proliferation, and inhibits apoptosis and differentiation in P19 embryonic carcinoma cells

Chen,Bin; Song,Guixian; Liu,Ming; Qian,Lingmei; Wang,Lihua; Gu,Haitao; Shen,Yahui

doi:10.3892/mmr.2016.4832

Inhibition of miR-29c promotes proliferation, and inhibits apoptosis and differentiation in P19 embryonic carcinoma cells

Authors:
- Bin Chen
- Guixian Song
- Ming Liu
- Lingmei Qian
- Lihua Wang
- Haitao Gu
- Yahui Shen
View Affiliations

Affiliations: Department of Cardiothoracic Surgery, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China, Department of Cardiology and Respiratory Medicine, Taizhou People's Hospital, Taizhou, Jiangsu 225300, P.R. China, Department of Cardiology, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China, Department of Neonatology, Nanjing Children's Hospital Affiliated to Nanjing Medical University, Nanjing, Jiangsu 210029, P.R. China
Published online on: January 29, 2016 https://doi.org/10.3892/mmr.2016.4832
Pages: 2527-2535
Copyright: © Chen et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

In our previous study, the upregulation of microRNA (miR)-29c was identified in the mother of a fetus with a congenital heart defect. However, the functional and regulatory mechanisms of miR‑29c in the development of the heart remain to be elucidated. In the present study, the role and mechanism of miR‑29c inhibition in heart development were investigated in an embryonic carcinoma cell model. Inhibition of miR‑29c promoted proliferation, and suppressed the apoptosis and differentiation of P19 cells. It was also demonstrated that Wingless‑related MMTV integration site 4 (Wnt4) was a target of miR‑29c, determined using bioinformatic analysis combined with luciferase assays. The inhibition of miR‑29c stimulated the WNT4/β‑catenin pathway, promoting proliferation of the P19 cells, but suppressing their differentiation into cardiomyocytes. Furthermore, the inhibition of miR‑29c promoted the expression of B cell lymphoma‑2 and inhibited cell apoptosis. These results demonstrate the significance of miR‑29c in the process of cardiac development and suggest that miR-29c dysregulation may be associated with the occurrence of CHD. Thus, miR-29c may have therapeutic potential in the future.

View Figures

View References

1	Khoshnood B, Lelong N, Houyel L, Thieulin AC, Jouannic JM, Magnier S, Delezoide AL, Magny JF, Rambaud C, Bonnet D, et al: Prevalence, timing of diagnosis and mortality of newborns with congenital heart defects: A population-based study. Heart. 98:1667–1673. 2012. View Article : Google Scholar : PubMed/NCBI
2	Hoffman JIe: The global burden of congenital heart disease. Cardiovasc J Afr. 24:141–145. 2013. View Article : Google Scholar : PubMed/NCBI
3	Lin CJ, Lin CY, Chen CH, Zhou B and Chang CP: Partitioning the heart: Mechanisms of cardiac septation and valve development. Development. 139:3277–3299. 2012. View Article : Google Scholar : PubMed/NCBI
4	Van der Bom T, Zomer AC, Zwinderman AH, Meijboom FJ, Bouma BJ and Mulder BJ: The changing epidemiology of congenital heart disease. Nat Rev Cardiol. 8:50–60. 2011. View Article : Google Scholar
5	Brennan P and Young ID: Congenital heart malformations: Aetiology and associations. Semin Neonatol. 6:17–25. 2001. View Article : Google Scholar : PubMed/NCBI
6	Bartel DP: MicroRNAs: Genomics, biogenesis, mechanism and function. Cell. 116:281–297. 2004. View Article : Google Scholar : PubMed/NCBI
7	Xiang R, Lei H, Chen M, Li Q, Sun H, Ai J, Chen T, Wang H, Fang Y and Zhou Q: The miR-17-92 cluster regulates FOG-2 expression and inhibits proliferation of mouse embryonic cardiomyocytes. Braz J Med Biol Res. 45:131–138. 2012. View Article : Google Scholar : PubMed/NCBI
8	Sluijter JP, van Mil A, van Vliet P, Metz CH, Liu J, Doevendans PA and Goumans MJ: MicroRNA-1 and -499 regulate differentiation and proliferation in human-derived cardiomyocyte progenitor cells. Arterioscler Thromb Vasc Biol. 30:859–868. 2010. View Article : Google Scholar : PubMed/NCBI
9	Bartel DP: MicroRNAs: Target recognition and regulatory functions. Cell. 136:215–233. 2009. View Article : Google Scholar : PubMed/NCBI
10	Li Y and Kowdley KV: MicroRNAs in common human diseases. Genomics Proteomics Bioinformatics. 10:246–253. 2012. View Article : Google Scholar : PubMed/NCBI
11	Chen J and Wang DZ: microRNAs in cardiovascular development. J Mol Cell Cardiol. 52:949–957. 2012. View Article : Google Scholar : PubMed/NCBI
12	Zhao Y, Ransom JF, Li A, Vedantham V, von Drehle M, Muth AN, Tsuchihashi T, McManus MT, Schwartz RJ and Srivastava D: Dysregulation of cardiogenesis, cardiac conduction and cell cycle in mice lacking miRNA-1-2. Cell. 129:303–317. 2007. View Article : Google Scholar : PubMed/NCBI
13	Liu and Olson EN: MicroRNA regulatory networks in cardiovascular development. Dev Cell. 18:510–525. 2010. View Article : Google Scholar : PubMed/NCBI
14	Zhao Y, Samal E and Srivastava D: Serum response factor regulates a muscle-specific microRNA that targets Hand2 during cardiogenesis. Nature. 436:214–220. 2005. View Article : Google Scholar : PubMed/NCBI
15	Liu N, Bezprozvannaya S, Williams AH, Qi X, Richardson JA, Bassel-Duby R and Olson EN: microRNA-133a regulates cardiomyocyte proliferation and suppresses smooth muscle gene expression in the heart. Genes Dev. 22:3242–3254. 2008. View Article : Google Scholar : PubMed/NCBI
16	Meder B, Katus HA and Rottbauer W: Right into the heart of microRNA-133a. Genes Dev. 22:3227–3231. 2008. View Article : Google Scholar : PubMed/NCBI
17	Zhu S, Cao L, Zhu J, Kong L, Jin J, Qian L, Zhu C, Hu X, Li M, Guo X, et al: Identification of maternal serum microRNAs as novel non-invasive biomarkers for prenatal detection of fetal congenital heart defects. Clin Chim Acta. 424:66–72. 2013. View Article : Google Scholar : PubMed/NCBI
18	Nguyen T, Kuo C, Nicholl MB, et al: Low miR-29c levels associated with TRG hypermethylation and melanoma progression. RNA Biology. 8:3572011.
19	Wang CM, Wang Y, Fan CG, Xu FF, Sun WS, Liu YG and Jia JH: miR-29c targets TNFAIP3, inhibits cell proliferation and induces apoptosis in hepatitis B virus-related hepatocellular carcinoma. Biochem Biophys Res Commun. 411:586–592. 2011. View Article : Google Scholar : PubMed/NCBI
20	Zhou K, Yu Z, Yi S, Li Z, An G, Zou D, Qi J, Zhao Y and Qiu L: miR-29c down-regulation is associated with disease aggressiveness and poor survival in Chinese patients with chronic lymphocytic leukemia. Leuk Lymphoma. 55:1544–1550. 2014. View Article : Google Scholar
21	Wang Y, Li Y, Sun J, Wang Q, Sun C, Yan Y, Yu L, Cheng D, An T, Shi C, et al: Tumor-suppressive effects of miR-29c on gliomas. Neuroreport. 24:637–645. 2013. View Article : Google Scholar : PubMed/NCBI
22	Xu F, Zhang Q, Cheng W, Zhang Z, Wang J and Ge J: Effect of miR-29b-1^* and miR-29c knockdown on cell growth of the bladder cancer cell line T24. J Int Med Res. 41:1803–1810. 2013. View Article : Google Scholar : PubMed/NCBI
23	Zeng X, Xiang J, Wu M, Xiong W, Tang H, Deng M, Li X, Liao Q, Su B and Luo Z: Circulating miR-17, miR-20a, miR-29c and miR-223 combined as non-invasive biomarkers in nasopharyngeal carcinoma. Plos One. 7:e463672012. View Article : Google Scholar
24	Matsuo M, Nakada C, Tsukamoto Y, Noguchi T, Uchida T, Hijiya N, Matsuura K and Moriyama M: MiR-29c is down-regulated in gastric carcinomas and regulates cell proliferation by targeting RCC2. Mol Cancer. 12:152013. View Article : Google Scholar
25	Kluiver J, Slezak-Prochazka I, Smigielska-Czepiel K, Halsema N, Kroesen BJ and van den Berg A: Generation of miRNA sponge constructs. Methods. 58:113–117. 2012. View Article : Google Scholar : PubMed/NCBI
26	Ebert MS, Neilson JR and Sharp PA: MicroRNA sponges: Competitive inhibitors of small RNAs in mammalian cells. Nat Methods. 4:721–726. 2007. View Article : Google Scholar : PubMed/NCBI
27	Pfaffl MW: A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res. 29:e452001. View Article : Google Scholar : PubMed/NCBI
28	Van der Heyden MA and Defize LH: Twenty one years of P19 cells: What an embryonal carcinoma cell line taught us about cardiomyocyte differentiation. Cardiovasc Res. 58:292–302. 2003. View Article : Google Scholar : PubMed/NCBI
29	Mukherji S, Ebert MS, Zheng GX, Tsang JS, Sharp PA and van Oudenaarden A: MicroRNAs can generate thresholds in target gene expression. Nat Genet. 43:854–859. 2011. View Article : Google Scholar : PubMed/NCBI
30	Tay FC, Lim JK, Zhu H, Hin LC and Wang S: Using artificial microRNA sponges to achieve microRNA loss-of-function in cancer cells. Adv Drug Deliv Rev. 81:117–127. 2015. View Article : Google Scholar
31	Ebert MS and Sharp PA: MicroRNA sponges: Progress and possibilities. RNA. 16:2043–2050. 2010. View Article : Google Scholar : PubMed/NCBI
32	Marvin MJ, Di Rocco G, Gardiner A, Bush SM and Lassar AB: Inhibition of Wnt activity induces heart formation from posterior mesoderm. Genes Dev. 15:316–327. 2001. View Article : Google Scholar : PubMed/NCBI
33	Eisenberg LM and Eisenberg CA: Wnt signal transduction and the formation of the myocardium. Dev Biol. 293:305–315. 2006. View Article : Google Scholar : PubMed/NCBI
34	Ueno S, Weidinger G, Osugi T, Kohn AD, Golob JL, Pabon L, Reinecke H, Moon RT and Murry CE: Biphasic role for Wnt/beta-catenin signaling in cardiac specification in zebrafish and embryonic stem cells. Proc Natl Acad Sci USA. 104:9685–9690. 2007. View Article : Google Scholar : PubMed/NCBI
35	Qyang Y, Martin-Puig S, Chiravuri M, Chen S, Xu H, Bu L, Jiang X, Lin L, Granger A, Moretti A, et al: The renewal and differentiation of Isl1+ cardiovascular progenitors are controlled by a Wnt/beta-catenin pathway. Cell Stem Cell. 1:165–179. 2007. View Article : Google Scholar
36	de la Pompa JL and Epstein JA: Coordinating tissue interactions: Notch signaling in cardiac development and disease. Dev Cell. 22:244–254. 2012. View Article : Google Scholar : PubMed/NCBI
37	Caliceti C, Nigro P, Rizzo P and Ferrari R: ROS, Notch and Wnt signaling pathways: Crosstalk between three major regulators of cardiovascular biology. Biomed Res Int. 2014:3187142014. View Article : Google Scholar
38	Zeng YA and Nusse R: Wnt proteins are self-renewal factors for mammary stem cells and promote their long-term expansion in culture. Cell Stem Cell. 6:568–577. 2010. View Article : Google Scholar : PubMed/NCBI
39	Gittenberger-de Groot AC, Bartelings MM, Poelmann RE, Haak MC and Jongbloed MR: Embryology of the heart and its impact on understanding fetal and neonatal heart disease. Semin Fetal Neonatal Med. 18:237–244. 2013. View Article : Google Scholar : PubMed/NCBI
40	Biala AK and Kirshenbaum LA: The interplay between cell death signaling pathways in the heart. Trends Cardiovasc Med. 24:325–331. 2014. View Article : Google Scholar : PubMed/NCBI