| Objective: To explore whether LSM4 gene would be expressed in endometrium and the relationship between LSM4 gene expression and endometrial decidualization in order to find a new entrance to understanding the mechanism of decidualization.Methods: 1. Characterization and localization of LSM4-m RNA and LSM4 protein in mice endometrium respectively and to see their relationship with uterine decidualization using in situ hybridization(ISH) and immunohistochemistry(IHC). 2. Appling In-Vitro decidualization model and Realtime-PCR technique to detect dynamic expression of LSM4-m RNA of stroma cells during decidualization. 3. LSM4-sh RNA–lentivirus infection in primary cultivated stroma cells for gene knock-down(LSM4-KD) and then induction for decidualization. Detection decidualization markers: DRRP and AP of LSM4-KD cells, in order to check whether the down regulation of LSM4 would inhibit decidualization of stroma cells.Results: 1. The accordant results of ISH and IHC showed that both m RNA and protein of LSM4 were expressed in endometrial luminal/glandular epithelium in D1; In D4(the implantation window stage), LSM4 expressed stronger in epithelium and expressed in some stromas cells nearby cavity laying epithelium; After D5, LSM4 expression reduced in epithelium but expressed significantly in endometrial stroma cells synchronously with decidualization; While the expression of LSM4 went higher in endometrial stroma cells and spreaded out to the whole endometrium, the amount of LSM4-cells reduced in shallow-layer stroma surround implanted embryos when decidulization moved on. In D8, the decidualization developed to its maximum, the signals of LSM4-m RNA in stroma cells reduced significantly yet the protein signals still high both in nucleus and cytoplasm. Inside the decidual cells, LSM4 protein showed reticular/particle aggregation and attached to the cell’s border.2. In-vitro decidualization experiments showed that the relative copy number of LSM4-m RNA in stroma cells reduced gradually from 0h~96h(P<0.05); Negative control groups in the LSM4-KD assay showed that LSM4-m RNA also reduced which was different from DRRP during decidualization progressing. 3. Compared with control group, LSM4-sh RNA infected stroma cells showed its reduction of LSM4-m RNA extremely significant(P<0.01), which proved the fact of LSM4-KD. The relative transcript level of DRRP-m RNA in LSM4-KD cells reduced apparently than control(p<0.05) at 48 h and more significant at 96h(p<0.01) during decidualization. The linear-fit analysis showed the quantitys of LSM4/DRRP m RNA in the same kind of cells showed that, at the same stage of decidualization, there was a certain correlation between LSM4/DRRP, while LSM4-KD led both m RNA/DRRP quantitys very low. 4. At the time of 96 h in decidualization, the activity of AP in LSM4-KD cells generally reduced apparently and cell volume was also small.Conclusion: 1. It is the first time to find the expression of LSM4 gene in mice endometrium in early stage of gestation. The expression pattern of LSM4 showed spatio-temporal specificity. There was a temporal expression wave surrounding the implantation and initiation for decidualization and also a spatio wave from uterine cavity to the deepest bottom of endometrium. For individual stroma the histological spatio wave was still temporal in fact: surrounding the initial point of decidualization. 2. It is the first time to show that LSM4-KD significantly inhibited the expression of DPRP and AP affirming the key role of LSM4 in decidualization of stromal cells; 3. We suggest that LSM4 may be one of the initial factors for decidualization of stroma cells. And also, LSM4 might be one of the family to form endometrial receptive stage... |
