Lentiviral vector mediated steroidogenic factor-1 silencing regulating decidualization and autophagy of endometrial stromal cells / 解剖学报

ABSTRACT

Objective To explore the influence and mechanism of lentiviral vector-mediated steroidogenic factor-1(SF-1)silence on decidual process of human endometrial stromal cells(hESCs).Methods The endometrial tissues of patients with endometriosis(EM)and normal pregnant women were collected,and the differential expression of SF-1 was detected by Real-time PCR and immunohistochemical staining.hESCs were isolated from the endometrial tissue of normal pregnancy and identified,hESCs were divided into control group,estradiol(E2)+progesterone(P4)group,short hairpin RAN(shRNA,sh)-normal control(NC)+E2+P4 group,sh-SF-1+E2+P4 group,after the corresponding processing,the morphological changes of hESCs were observed under an inverted microscope.Real-time PCR and Western blotting were used to detect the mRNA expression levels and protein levels of human insulin-like growth factor-binding protein-1(IGFBP-1)and prolactin(PRL)in cells,respectively,flow cytometry was used to determine the cycle distribution of the cells,immunofluorescence staining was used to observe the expression of the intracellular auto-bid microtubule-associated protein light chain 3(LC3),Western blotting was used to determine the protein level of intracellular autophagy-related proteins LC3-Ⅱ,LC3-Ⅰ and Beclin-1.Results The relative expression of SF-1 mRNA and the positive rate of SF-1 protein in endometrial tissue of EM patients were higher than those of normal pregnancy endometrial tissue(P<0.05).Compared with the sh-NC+E2+P4 group,the cells in the sh-SF-1+E2+P4 group were mostly long-spindle,and there was no obvious decidualization.The relative mRNA expression and protein expression of IGFBP1 and PRL were significantly down-regulated(P<0.05),the proportion of cells in G0/G1 phase was significantly decreased,the proportion of cells in S phase was significantly increased(P<0.05),the ratio of LC3-Ⅱ/LC3-Ⅰwas significantly increased,and the relative expression of Beclin-1 protein was also significantly up-regulated(P<0.05).Conclusion The expression of SF-1 in the endometrial tissue of EM patients is increased,and SF-1 silencing mediated by lentiviral vector can inhibit the decidualization process of hESCs,which may be related to regulating the level of autophagy.