miR-3074-5p Promotes the Apoptosis but Inhibits the Invasiveness of Human Extravillous Trophoblast

Abstract

Objective:

The objective of this study was to observe the effects of the overexpression of miR-3074-5p in human trophoblast cells in vitro.

Design:

Experimental in vitro study in HTR8/SVneo cells.

Methods:

HTR8/SVneo cells were transfected with miR-3074-5p mimic. The cell apoptosis and invasion were measured via flow cytometry and transwell assay, respectively. The expression levels of P53, Cyclin Dependent Kinase Inhibitor 1B (P27), BCL-2, BCL2 associated X (BAX), and BCL2 like 14 (BCL-G) in HTR8/SVneo cells were determined by Western blot. The alterations in gene expression profile of HTR8/SVneo cells were evaluated by complementary DNA microarray assay, and the differential expressions of dihydrolipoamide S-succinyltransferase (DLST), growth-associated protein 43 (GAP43), runt-related transcription factor 2 (RUNX2), and C-C type chemokine receptor 3 (CCR3) were validated by Western blot. Biofunctions of these differentially expressed genes were enriched by Gene Ontology analysis.

Results:

The overexpression of miR-3074-5p in HTR8/SVneo cells promoted cell apoptosis but inhibited cell invasion, being accompanied by the significantly elevated expressions of P27, BCL-2, and BCL-G. Meanwhile, an increased expression of P27 and P57 was also detected in a small sample size of placental villi of recurrent miscarriage (RM) patients. Totally, 411 genes and 397 genes were screened out, respectively, to be downregulated or upregulated at least by 2-folds in miR-3074-5p overexpressed HTR8/SVneo cells. These differentially expressed genes were involved in several important functions related to pregnancy. Subsequently, the reduced expressions of DLST and GAP43 proteins, as well as the increased expressions of CCR3 and RUNX2 proteins, were validated in miR-3074-5p overexpressed HTR8/SVneo cells.

Conclusion:

These data suggested a potential contribution of miR-3074-5p in the pathogenesis of RM by disturbing the normal activities of trophoblast cells.

Keywords

miR-3074-5p trophoblast cells apoptosis invasion p27

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References

Brezina

Kutteh

. Classic and cutting-edge strategies for the management of early pregnancy loss. Obstet Gynecol Clin North Am. 2014;41(1):1–18.

Rai

Regan

. Recurrent miscarriage. Lancet. 2006;368(9535):601–611.

Lunghi

Ferretti

Medici

. Control of human trophoblast function. Reprod Biol Endocrinol. 2007;5(1):6.

Winship

Correia

Krishnan

. Blocking endogenous leukemia inhibitory factor during placental development in mice leads to abnormal placentation and pregnancy loss. Sci Rep. 2015;5(1):13237.

Wightman

Ruvkun

. Posttranscriptional regulation of the heterochronic gene lin-14 by lin-4 mediates temporal pattern formation in C. elegans . Cell. 1993;75(5):855–862.

Bartel

. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 2004;116(2):281–297.

Harun

Ruban

Matin

. Cytotrophoblast stem cell lines derived from human embryonic stem cells and their capacity to mimic invasive implantation events. Hum Reprod. 2006;21(6):1349–1358.

Galliano

Pellicer

. MicroRNA and implantation. Fertil Steril. 2014;101(6):1531–1544.

Hull

Nisenblat

. Tissue and circulating microRNA influence reproductive function in endometrial disease. Reprod Biomed Online. 2013;27(5):515–529.

10.

Dong

Zhang

Xia

. Genome-wide miRNA profiling of villus and decidua of recurrent spontaneous abortion patients. Reproduction. 2014;148(1):33–41.

11.

Qin

Tang

Yang

. Potential role of circulating microRNAs as a biomarker for unexplained recurrent spontaneous abortion. Fertil Steril. 2016;105(5):1247–1254 e1243.

12.

Tang

Gao

. Expression profile of micro-RNAs and functional annotation analysis of their targets in human chorionic villi from early recurrent miscarriage. Gene. 2016;576(1 pt 2):366–371.

13.

Zhang

Yang

. Aberrant placental villus expression of miR-486-3p and miR-3074-5p in recurrent miscarriage patients and uterine expression of these micrornas during early pregnancy in mice. Gynecol Obstet Invest. 2015;1(1):1.

14.

Abou-Kheir

Barrak

Hadadeh

. HTR-8/SVneo cell line contains a mixed population of cells. Placenta. 2017;50(1):1–7.

15.

Yang

Chen

. miR-15b-AGO2 play a critical role in HTR8/SVneo invasion and in a model of angiogenesis defects related to inflammation. Placenta. 2016;41(1):62–73.

16.

Ding

Chen

Wang

. MicroRNA-128a-induced apoptosis in HTR-8/SVneo trophoblast cells contributes to pre-eclampsia. Biomed Pharmacother. 2016;81(1):63–70.

17.

Graham

Hawley

. Establishment and characterization of first trimester human trophoblast cells with extended lifespan. Exp Cell Res. 1993;206(2):204–211.

18.

Salic

Mitchison

. A chemical method for fast and sensitive detection of DNA synthesis in vivo. Proc Natl Acad Sci U S A. 2008;105(7):2415–2420.

19.

Szklarczyk

Franceschini

Wyder

. STRING v10: protein-protein interaction networks, integrated over the tree of life. Nucleic Acids Res. 2015;43(database issue):D447–D452.

20.

Unek

Ozmen

Kipmen-Korgun

. Immunolocalization of PCNA, Ki67, p27 and p57 in normal and dexamethasone-induced intrauterine growth restriction placental development in rat. Acta Histochemica. 2012;114(1):31–40.

21.

Dior

Kogan

Chill

. Emerging roles of microRNA in the embryo-endometrium cross talk. Semin Reprod Med. 2014;32(5):402–409.

22.

. Expression profiles and clinical significance of MicroRNAs in papillary renal cell carcinoma: a STROBE-compliant observational study. Medicine. 2015;94(16):e767.

23.

Korkmaz

Sakinci

Akyol

. Location of proliferating cell nuclear antigen and p53 protein in human first trimester and term placenta. Anal Quant Cytopathol Histpathol. 2013;35(6):335–343.

24.

Nadeem

Brkic

Chen

. Cytoplasmic mislocalization of p27 and CDK2 mediates the anti-migratory and anti-proliferative effects of Nodal in human trophoblast cells. J Cell Sci. 2013;126(pt 2):445–453.

25.

Zhang

. Deficiency of monoclonal non-specific suppressor factor beta (MNSFB) promotes pregnancy loss in mice. Mol Reprod Dev. 2015;82(6):475–488.

26.

Suzuki

Young

Sharp

. Super-enhancer-mediated RNA processing revealed by integrative MicroRNA network analysis. Cell. 2017;168(6):1000–1014 e1015.

27.

Borriello

Bencivenga

Criscuolo

. Targeting p27Kip1 protein: its relevance in the therapy of human cancer. Expert Opin Ther Targets. 2011;15(6):677–693.

28.

Podmirseg

Jakel

Ranches

. Caspases uncouple p27(Kip1) from cell cycle regulated degradation and abolish its ability to stimulate cell migration and invasion. Oncogene. 2016;35(35):4580–4590.

29.

Wei

Jin

Zhang

. Expression of Bcl-2 and p53 at the fetal-maternal interface of rhesus monkey. Reprod Biol Endocrinol. 2005;3(1):4.

30.

Gross

. BCL-2 family proteins as regulators of mitochondria metabolism. Biochim Biophys Acta. 2016;1857(8):1243–1246.

31.

Bai

Wang

Han

. BCL2L10 inhibits growth and metastasis of hepatocellular carcinoma both in vitro and in vivo. Mol Carcinog. 2016;1(1):1.

32.

Unek

Ozmen

Mendilcioglu

. The expression of cell cycle related proteins PCNA, Ki67, p27 and p57 in normal and preeclamptic human placentas. Tissue & cell. 2014;46(3):198–205.

33.

Gundogan

Elwood

Mark

. Ethanol-induced oxidative stress and mitochondrial dysfunction in rat placenta: relevance to pregnancy loss. Alcohol Clin Exp Res. 2010;34(3):415–423.

34.

Wang

Zhang

. Deep-sequencing identification of differentially expressed miRNAs in decidua and villus of recurrent miscarriage patients. Arch Gynecol Obstet. 2016;293(5):1125–1135.

35.

Miled

Pontoglio

Garbay

. A genomic map of p53 binding sites identifies novel p53 targets involved in an apoptotic network. Cancer Res. 2005;65(12):5096–5104.

36.

Zhang

Qian

. Down-regulation of type I Runx2 mediated by dexamethasone is required for 3T3-L1 adipogenesis. Mol Endocrinol. 2012;26(5):798–808.

37.

Henriquez

Hepp

Merino

. C/EBPbeta binds the P1 promoter of the Runx2 gene and up-regulates Runx2 transcription in osteoblastic cells. J Cell Physiol. 2011;226(11):3043–3052.

38.

Anderson

Peng

. The TCA cycle transferase DLST is important for MYC-mediated leukemogenesis. Leukemia. 2016;30(6):1365–1374.

39.

Diaz-Munoz

Bell

Fairfax

. The RNA-binding protein HuR is essential for the B cell antibody response. Nat Immunol. 2015;16(4):415–425.

40.

Zhang

Kong

. Nerve growth factor improves functional recovery by inhibiting endoplasmic reticulum stress-induced neuronal apoptosis in rats with spinal cord injury. J Transl Med. 2014;12(1):130.

41.

Luo

Shen

Liu

. CART peptide induces neuroregeneration in stroke rats. J Cereb Blood Flow Metab. 2013;33(2):300–310.

42.

Hannan

Jones

White

. The chemokines, CX3CL1, CCL14, and CCL4, promote human trophoblast migration at the feto-maternal interface. Biol Reprod. 2006;74(5):896–904.

43.

Athanassakis

Papadimitriou

Vassiliadis

. Murine ectoplacental cone-derived trophoblast cells express chemokine receptors. J Reprod Immunol. 2001;50(2):105–119.

44.

Chang

Tsai

Liou

. Effect of butyrate on collagen expression, cell viability, cell cycle progression and related proteins expression of MG-63 osteoblastic cells. PloS one. 2016;11(11):e0165438.

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miR-3074-5p Promotes the Apoptosis but Inhibits the Invasiveness of Human Extravillous Trophoblast–Derived HTR8/SVneo Cells In Vitro

Abstract

Objective:

Design:

Methods:

Results:

Conclusion:

Keywords

Get full access to this article

References

Supplementary Material