miR-363-3p表达对人子宫内膜间质细胞生物学表型的影响
|
摘要
目的研究微RNA-363-3p(miR-363-3p)在子宫内膜异位症(endometriosis,EMs)患者在位内膜(eutopic endometria,Eu)及异位内膜(ectopic endometria,Ec)中的表达并探究miR-363-3p对人子宫内膜间质细胞(endometrial stroma cells,EnSCs)生物学表型的影响。方法选择2017年10月-2018年10月在本中心妇产科行腹腔镜手术且术后经病理检查确诊为EMs的患者30例,年龄24-48岁。采用qRT-PCR法检测Eu(n=30)、Ec(n=37)组织中miR-363-3p的表达。运用脂质体转染的方法分别将miR-363-3p类似物核酸序列(简称mimics)及阴性对照序列(简称NC)转染EnSCs,检测细胞中凋亡、增殖及黏附能力相关指标Caspase3、Ki-67、MMP-2、MMP-9的变化并利用CCK8和流式细胞术检测miR-363-3p表达变化后对细胞增殖及凋亡能力的影响。结果与Eu组相比,Ec组miR-363-3p表达下降(P<0.001)。在EnSCs中过表达miR-363-3p后,Caspase3表达升高(P=0.01),Ki-67表达下降(P<0.001),MMP-2、MMP-9表达无明显变化。过表达miR-363-3p能显著增强EnSCs的凋亡率(P<0.001),削弱其增殖能力(P<0.001)。结论在子宫内膜异位症患者异位内膜中miR-363-3p表达下调可能诱使异位内膜细胞凋亡能力减弱,促进EMs的发病。
Objective To explore the expression of miR-363-3 p in eutopic endometria(Eu)and ectopic endometria(Ec),and explore their impacts on the biological behaviors of endometrial stroma cells(EnSCs).Methods The eutopic(n=30)and ectopic(n=37)endometria from 30 women with endometriosis who underwent laparoscopic surgical procedures in our hospital from October 2017 to October 2018 with age ranging from 24 to 48 years were collected.Real time quantitative PCR was used to detect the expression of miR-363-3 p in these tissues.EnSCs were transfected with the miR-363-3 p mimics and negative control.After transfection,apoptosis marker caspase3,proliferation,Ki-67 and invasive makers MMP-2,MMP-9 were measured and CCK8,flow cytometry assay were used to evaluate cell proliferation and apoptosis.Results Compared to Eu,the levels of miR-363-3 p was significantly lower in Ec(P<0.001).Compared with the negative control,overexpression of miR-363-3 p was seen in EnSCs.After overexpression of miR-363-3 p in EnSCs,Caspase3 increased(P=0.01),Ki-67 decreased(P<0.001),while MMP-2,MMP-9 did not remarkably change(P>0.05).However,the apoptosis rate significantly increased(P<0.001)and proliferation rate reduced(P<0.001).Conclusion Downregulation of miR-363-3 p in Ec lesions increased of endomereiosis may promote cell proliferation and result in the occurrence of endometriosis.
Objective To explore the expression of miR-363-3 p in eutopic endometria(Eu)and ectopic endometria(Ec),and explore their impacts on the biological behaviors of endometrial stroma cells(EnSCs).Methods The eutopic(n=30)and ectopic(n=37)endometria from 30 women with endometriosis who underwent laparoscopic surgical procedures in our hospital from October 2017 to October 2018 with age ranging from 24 to 48 years were collected.Real time quantitative PCR was used to detect the expression of miR-363-3 p in these tissues.EnSCs were transfected with the miR-363-3 p mimics and negative control.After transfection,apoptosis marker caspase3,proliferation,Ki-67 and invasive makers MMP-2,MMP-9 were measured and CCK8,flow cytometry assay were used to evaluate cell proliferation and apoptosis.Results Compared to Eu,the levels of miR-363-3 p was significantly lower in Ec(P<0.001).Compared with the negative control,overexpression of miR-363-3 p was seen in EnSCs.After overexpression of miR-363-3 p in EnSCs,Caspase3 increased(P=0.01),Ki-67 decreased(P<0.001),while MMP-2,MMP-9 did not remarkably change(P>0.05).However,the apoptosis rate significantly increased(P<0.001)and proliferation rate reduced(P<0.001).Conclusion Downregulation of miR-363-3 p in Ec lesions increased of endomereiosis may promote cell proliferation and result in the occurrence of endometriosis.
引文
1 Kim MK,Lee SK,Park JH,et al.Ginsenoside Rg3 Decreases Fibrotic and Invasive Nature of Endometriosis by Modulating miRNA-27b:In Vitro and In Vivo Studies[J].Sci Rep,2017,7(1):17670.
2 Graham A,Falcone T,Nothnick WB.The expression of microRNA-451 in human endometriotic lesions is inversely related to that of macrophage migration inhibitory factor(MIF)and regulates MIFexpression and modulation of epithelial cell survival[J].Hum Reprod,2015,30(3):642-652.
3 Zhou M,Fu J,Xiao L,et al.miR-196a overexpression activates the MEK/ERK signal and represses the progesterone receptor and decidualization in eutopic endometrium from women with endometriosis[J].Hum Reprod,2016,31(11):2598-2608.
4 Wang Y,Chen T,Huang H,et al.miR-363-3p inhibits tumor growth by targeting PCNA in lung adenocarcinoma[J].Oncotarget,2017,8(12):20133-20144.
5 Khuu C,Sehic A,Eide L,et al.Anti-proliferative Properties of miR-20b and miR-363 from the miR-106a-363 Cluster on Human Carcinoma Cells[J].Microrna,2016,5(1):19-35.
6 Li M,Zhou Y,Xia T,et al.Circulating microRNAs from the miR-106a-363 cluster on chromosome X as novel diagnostic biomarkers for breast cancer[J].Breast Cancer Res Treat,2018,170(2):257-270.
7 Ghazal S,McKinnon B,Zhou J,et al.H19 lncRNA alters stromal cell growth via IGF signaling in the endometrium of women with endometriosis[J].EMBO Mol Med,2015,7(8):996-1003.
8 Giudice LC,Kao LC.Endometriosis[J].Lancet,2004,364(9447):1789-1799.
9 Vetvicka V,Lagana AS,Salmeri FM,et al.Regulation of apoptotic pathways during endometriosis:from the molecular basis to the future perspectives[J].Arch Gynecol Obstet,2016,294(5):897-904.
10 Cho S,Mutlu L,Zhou Y,et al.Aromatase inhibitor regulates let-7expression and let-7f-induced cell migration in endometrial cells from women with endometriosis[J].Fertil Steril,2016,106(3):673-680.
11 Abe W,Nasu K,Nakada C,et al.miR-196b targets c-myc and Bcl-2 expression,inhibits proliferation and induces apoptosis in endometriotic stromal cells[J].Hum Reprod,2013,28(3):750-761.
12 Nyholt DR,Low SK,Anderson CA,et al.Genome-wide association meta-analysis identifies new endometriosis risk loci[J].Nat Genet,2012,44(12):1355-1359.
13 Chen J,Gu L,Ni J,et al.MiR-183 Regulates ITGB1P Expression and Promotes Invasion of Endometrial Stromal Cells[J/OL].https://dx.doi.org/10.1155/2015/340218.
14 Hu F,Min J,Cao X,et al.MiR-363-3p inhibits the epithelial-tomesenchymal transition and suppresses metastasis in colorectal cancer by targeting Sox4[J].Biochem Biophys Res Commun,2016,474(1):35-42.
15 Liu J,Li Q,Li R,et al.MicroRNA-363-3p inhibits papillary thyroid carcinoma progression by targeting PIK3CA[J].Am JCancer Res,2017,7(1):148-158.
16 Zhao L,Gu C,Ye M,et al.Integration analysis of microRNA and mRNA paired expression profiling identifies deregulated microRNA-transcription factor-gene regulatory networks in ovarian endometriosis[J].Reprod Biol Endocrinol,2018,16(1):4.
17 Shi XY,Gu L,Chen J,et al.Downregulation of miR-183 inhibits apoptosis and enhances the invasive potential of endometrial stromal cells in endometriosis[J].Int J Mol Med,2014,33(1):59-67.
18 Szymanowski K,Mikolajczyk M,Wirstlein P,et al.Matrix metalloproteinase-2(MMP-2),MMP-9,tissue inhibitor of matrix metalloproteinases(TIMP-1)and transforming growth factor-beta2(TGF-beta2)expression in eutopic endometrium of women with peritoneal endometriosis[J].Ann Agric Environ Med,2016,23(4):649-653.
2 Graham A,Falcone T,Nothnick WB.The expression of microRNA-451 in human endometriotic lesions is inversely related to that of macrophage migration inhibitory factor(MIF)and regulates MIFexpression and modulation of epithelial cell survival[J].Hum Reprod,2015,30(3):642-652.
3 Zhou M,Fu J,Xiao L,et al.miR-196a overexpression activates the MEK/ERK signal and represses the progesterone receptor and decidualization in eutopic endometrium from women with endometriosis[J].Hum Reprod,2016,31(11):2598-2608.
4 Wang Y,Chen T,Huang H,et al.miR-363-3p inhibits tumor growth by targeting PCNA in lung adenocarcinoma[J].Oncotarget,2017,8(12):20133-20144.
5 Khuu C,Sehic A,Eide L,et al.Anti-proliferative Properties of miR-20b and miR-363 from the miR-106a-363 Cluster on Human Carcinoma Cells[J].Microrna,2016,5(1):19-35.
6 Li M,Zhou Y,Xia T,et al.Circulating microRNAs from the miR-106a-363 cluster on chromosome X as novel diagnostic biomarkers for breast cancer[J].Breast Cancer Res Treat,2018,170(2):257-270.
7 Ghazal S,McKinnon B,Zhou J,et al.H19 lncRNA alters stromal cell growth via IGF signaling in the endometrium of women with endometriosis[J].EMBO Mol Med,2015,7(8):996-1003.
8 Giudice LC,Kao LC.Endometriosis[J].Lancet,2004,364(9447):1789-1799.
9 Vetvicka V,Lagana AS,Salmeri FM,et al.Regulation of apoptotic pathways during endometriosis:from the molecular basis to the future perspectives[J].Arch Gynecol Obstet,2016,294(5):897-904.
10 Cho S,Mutlu L,Zhou Y,et al.Aromatase inhibitor regulates let-7expression and let-7f-induced cell migration in endometrial cells from women with endometriosis[J].Fertil Steril,2016,106(3):673-680.
11 Abe W,Nasu K,Nakada C,et al.miR-196b targets c-myc and Bcl-2 expression,inhibits proliferation and induces apoptosis in endometriotic stromal cells[J].Hum Reprod,2013,28(3):750-761.
12 Nyholt DR,Low SK,Anderson CA,et al.Genome-wide association meta-analysis identifies new endometriosis risk loci[J].Nat Genet,2012,44(12):1355-1359.
13 Chen J,Gu L,Ni J,et al.MiR-183 Regulates ITGB1P Expression and Promotes Invasion of Endometrial Stromal Cells[J/OL].https://dx.doi.org/10.1155/2015/340218.
14 Hu F,Min J,Cao X,et al.MiR-363-3p inhibits the epithelial-tomesenchymal transition and suppresses metastasis in colorectal cancer by targeting Sox4[J].Biochem Biophys Res Commun,2016,474(1):35-42.
15 Liu J,Li Q,Li R,et al.MicroRNA-363-3p inhibits papillary thyroid carcinoma progression by targeting PIK3CA[J].Am JCancer Res,2017,7(1):148-158.
16 Zhao L,Gu C,Ye M,et al.Integration analysis of microRNA and mRNA paired expression profiling identifies deregulated microRNA-transcription factor-gene regulatory networks in ovarian endometriosis[J].Reprod Biol Endocrinol,2018,16(1):4.
17 Shi XY,Gu L,Chen J,et al.Downregulation of miR-183 inhibits apoptosis and enhances the invasive potential of endometrial stromal cells in endometriosis[J].Int J Mol Med,2014,33(1):59-67.
18 Szymanowski K,Mikolajczyk M,Wirstlein P,et al.Matrix metalloproteinase-2(MMP-2),MMP-9,tissue inhibitor of matrix metalloproteinases(TIMP-1)and transforming growth factor-beta2(TGF-beta2)expression in eutopic endometrium of women with peritoneal endometriosis[J].Ann Agric Environ Med,2016,23(4):649-653.