Physiological significance of placental leucine aminopeptidase /oxytocinase at the interface between mother and fetus

我们首次证明了绒毛膜癌滋养层细胞的-297到-94区域包含四个足迹位点(fp1到fp4)，对于转录是重要的。AP-2α、AP-2γ和Ikaros与Fp3(-214~-183)的结合对于这些细胞的高启动子活性是重要的。功能分析表明，AP-2对人P-LAP基因的表达至关重要，并且Ikaros与AP-2协同诱导最大的启动子活性。这显然是最初被描述为淋巴限制转录因子的Ikaros在滋养层细胞中表达的第一个结果。P-LAP启动子区域包含细胞因子诱导的转录因子的假定结合部位。因此，我们推测炎性细胞因子抑制滋养层细胞中P-LAP的表达。白介素1β(IL-1β)可增加P-LAP活性和蛋白表达。半定量逆转录聚合酶链式反应和Southern blotting结果显示，IL-1β还能增加P-LAP基因的表达，而这一作用可被放线菌素预先作用于放线菌酮所抑制。荧光素酶检测没有发现任何可以解释IL-1β诱导的P-LAP基因积聚的调控区域。绒毛膜羊膜炎患者胎盘的免疫组织化学分析显示，P-LAP染色显示在大量炎性细胞浸润的部位。这些发现表明，长期暴露于IL-1β可诱导胎盘中的P-LAP，可能是通过其他从头合成蛋白质来实现的。透射免疫电子显微镜显示P-LAP表达于合体滋养层细胞顶端微绒毛表面。我们观察到，P-LAP存在于母亲和胎儿之间相互作用的微绒毛上，这表明这些酶可能参与了分解胎儿和母亲的多肽激素，以调节生物活性。

We firstly demonstrated that the region from-297 to-94, which contains four footprint sites (FP1 to FP4), is important for transcription in choriocarcinoma trophoblastic cells. AP-2α, AP-2γ, and Ikaros binding to FP3 (-214 to -183) was important for high promoter activity in these cells. Functional analysis showed that AP-2 is critical for human P-LAP gene expression and Ikaros cooperates with AP-2 to induce maximal promoter activity. This is apparently the first result that Ikaros, initially characterized as a lymphoid-restricted transcriptional factor, is expressed in trophoblasts. P-LAP promoter region contains putative binding sites for cytokine-induced transcription factors. We therefore postulated that inflammatory cytokines suppress P-LAP expression in trophoblasts. Interleukin-1beta (IL-1β) increased P-LAP activity and proteins. Semi-quantitative RT-PCR and Southern blotting showed that IL-1β also increased P-LAP mRNA, which was abrogated by prior exposure to cycloheximide. Luciferase assays did not reveal any regulatory regions that could explain IL-1β-induced P-LAP mRNA accumulation. Immunohistochemical analysis of the human placenta with chorioamnionitis demonstrated prominent P-LAP staining at sites of abundant inflammatory cell infiltration. These findings indicated that prolonged exposure to IL-1β induces P-LAP in the placenta, possibly via other de novo protein synthesis. Transmission immunoelectron microscopy revealed that P-LAP was expressed on the surface of apical microvilli of syncytiotrophoblast cells. Our observation that P-LAP is present on the microvilli, which is a site of interaction between the mother and fetus, suggests possible involvement of these enzymes in cleaving peptide hormones from the fetus and mother in order to regulate bioactivity.