Function And Regulatory Mechanisms Of Hmgn During Uterine Decidualization In Mice

Uterine decidualization involves extensive stromal cells proliferation and differentiation, which is essential for normal embryo implantation and pregnancy maintenance. Genome-wide microarray analyses have identified a number of genes that are up-regulated or down-regulated during decidualization. To date, accumulating data have shown that gene expression involves a change of chromatin structure which can be regulated by structural proteins such as the high mobility group nucleosomal binding(Hmgn) family. The Hmgn family contains five members named Hmgn1-5, which bind specifically to nucleosome core particle, reduce the compaction of the chromatin fiber, alter the structure of chromatin and thereby affect a variety of DNA-dependent activities such as transcription, replication, recombination and DNA repair. Although some studies have demonstrated that Hmgn is involved in the regulation of gene expression, cellular differentiation and development, the function and regulatory mechanisms of Hmgn during uterine decidualization are still unknown. The present study was to investigate the expression, function and regulation of the Hmgn family in mouse uterus during the peri-implantation period by in situ hybridization, real-time PCR, gene overexpression, RNA interference, etc.The results found that Hmgn1 m RNA was highly expressed in the stromal cells around implanting blasocyst on day 5 of pregnancy, and the expression then spreaded to the decidual region on days 6-8 of pregnancy. Simultaneously, increased expression of Hmgn1 was also observed in the artificial and in vitro induced decidualization models. Hmgn1 induced the proliferation of uterine stromal cells and expression of Ccna1, Ccnb1, Ccnb2 and Cdk1 in the absence of estrogen and progesterone. Overexpression of Hmgn1 could enhance the expression of Prl8a2 and Prl3c1 which were two well-known differentiation markers for decidualization, whereas inhibition of Hmgn1 with specific si RNA could reduce their expression. Further studies found that Hmgn1 could mediate the effects of C/EBPβ on the two differentiation markers during in vitro decidualization. In the uterine stromal cells, c AMP could stimulate the expression of Hmgn1 via C/EBPβ. si RNA-mediated down-regulation of Hmgn1 could attenuate the effects of c AMP on expression of the differentiation markers in uterine stromal cells. During in vitro decidualization, Hmgn1 might act downstream of C/EBPβ to regulate the expression of Cox-2, m PGES-1 and Vegf. Progesterone could up-regulate the expression of Hmgn1 in the ovariectomized mouse uterus, uterine epithelial cells and stromal cells. Knockdown of C/EBPβ with si RNA alleviated the up-regulation of progesterone on Hmgn1 expression.Hmgn2 m RNA was detected in the luminal epithelium, glandular epithelium and stromal cells on days 2-5 of pregnancy, and then widely expressed in the decidual region on days 6-8 of pregnancy. Furthermore, observations of remarkable induction of Hmgn2 m RNA in decidualizing stromal cells under artificially induced decidualization in vivo. Hmgn2 might have the vital role on the differentiation of uterine stromal cells, but not proliferation. In the uterine stromal cells,, progesterone could significantly induce Hmgn2 m RNA expression. Hmgn2 could mediate the effects of progesterone on the differentiation markers Prl8a2 and Prl3c1. Hmgn2 might regulate the expression of decidualization-related gene Hand2. Inhibition of Hmgn2 could impede the effect of progesterone on the expression of Hand2. Estrogen could up-regulate the expression of Hmgn2 in uterine epithelial cells, but the up-regulation was abrogated by a pretreatment with estrogen receptor antagonist ICI 182,780.High levels of Hmgn3 m RNA were detected in the decidua and decidualizing stromal cells. Hmgn3 overexpression enhanced the expression of decidualization markers Prl8a2 and Prl3c1, whereas inhibition of Hmgn3 reduced their expression. Hmgn3 could mediate the effects of Hoxa10 and c AMP on the expression of Prl8a2 and Prl3c1. Further study found that Hmgn3 directed the process of decidualization through influencing the expression of Hand2. Progesterone could induce the expression of Hmgn3 in the ovariectomized mouse uterus, uterine epithelial cells and stromal cells. Knockdown of Hoxa10 with si RNA alleviated the induction of progesterone and c AMP on Hmgn3 expression. si RNA-mediated down-regulation of Hmgn3 in the uterine stromal cells could attenuate the effects of progesterone, c AMP and Hoxa10 on the expression of Hand2.Hmgn5 m RNA was strongly expressed in the stromal cells around implanting blasocyst on day 5 of pregnancy, and the signal then specially spreaded to the mesometrial decidua on days 6-8 of pregnancy and artificially induced decidualization in vivo. Meanwhile, high levels of Hmgn5 m RNA were also detected in vitro induced decidualization models. During in vitro decidualization, Hmgn5 could affect the proliferation of uterine stromal cells through Ccnd3 and Cdk4. Hmgn5 could regulate the expression of decidualization markers Prl8a2 and Prl3c1. Hmgn5 mediated the effects of Hoxa10 on the expression of Prl8a2 and Prl3c1. c AMP could regulate Hmgn5 expression in uterine stromal cells through Hoxa10, whereas loss of Hmgn5 impaired the effects of c AMP on the differentiation of uterine stromal cells. Hmgn5 might act downstream of Hoxa10 to regulate the expression of Cox-2, Vegf and Mmp2 during in vitro decidualization. Progesterone could induce Hmgn5 expression in uterine stromal cells and epithelial cells. Moreover, Hoxa10 mediated the effects of progesterone on Hmgn5 expression in this study.In conclusion, these results suggest that the Hmgn family may play an important role during uterine decidualization in mice.