不明原因习惯性流产与血浆激肽释放酶表达量及妊娠免疫耐受的相关性

Correlation of unexplained recurrent pregnancy loss with plasma kallikrein expression and pregnancy immune tolerance

[目的]探讨不明原因习惯性流产(RPL)与血浆激肽释放酶(PK)表达量及妊娠免疫耐受的相关性.[方法]选择30例不明原因RPL患者作为观察组,另选择30名健康女性作为对照组,检测两组血清中PK的活性;采用Real-time PCR与Western Blot法检测两组外周血单核细胞(PBMC)中PK mRNA和蛋白表达水平;分别将MOCK、pcDNA3-6myc-PK、pcDNA3-6myc-PK S578A及MOCK、SiControl、SiPK转染入JEG-3细胞,比较Th1/Th2细胞因子的分泌量;利用Transwell小室法和CCK-8实验检测干扰及过表达PK基因对滋养层细胞的侵袭及免疫细胞增殖的影响.[结果]观察组血清中PK活性显著高于对照组,且观察组患者PBMC中PK mRNA和蛋白表达水平均明显高于对照组(P<0.05);过表达和RNA干扰实验结果表明,观察组促炎性细胞因子的表达水平明显上调(P<0.05);PK基因siRNA沉默可明显增加滋养层细胞的侵袭,而siPK组对免疫细胞增殖有抑制作用(P<0.05).[结论] PK在观察组患者血清与PBMC中呈高表达,且PK可增强炎性反应及免疫细胞增殖,且对滋养层细胞侵袭有抑制作用.

OBJECTIVE To investigate the correlation of unexplained recurrent pregnancy loss(RPL) with plasma kallikrein(PK) and pregnancy immune tolerance. METHODS Thirty patients with unexplained RPL were selected as the observation group, and 30 healthy women were selected as the control group. The PK activity in serum of the two groups were analyzed. Real-time PCR and Western Blot were used to detect the expression of PK mRNA and protein in peripheral blood mononuclear cells(PBMC) in the two groups. MOCK, pcDNA3-6 myc-PK, pcDNA3-6 myc-PK S578 A and MOCK, SiControl, and SiPK were transfected into JEG-3 cells, respectively, and the secretion of Th1/Th2 cytokines was compared. Transwell chamber assay and CCK-8 assay were used to detect the effects of interference and overexpression of PK gene on the invasion of trophoblast cells and proliferation of immune cells. RESULTS The activity of PK in serum of the observation group was significantly higher than that of the control group, and the expressions of PK mRNA and protein in PBMC of the observation group were significantly higher than that of the control group(P<0.05). The results of overexpression and RNA interference experiments showed that the expressions of proinflammatory cytokines in the observation group was significantly up-regulated(P<0.05). SiRNA silences of PK gene could significantly increase the invasion of trophoblast cells, while siPK group could inhibit the proliferation of immune cells(P<0.05). CONCLUSION PK showed high expressions in serum and PBMC of the observation group, and PK could significantly enhance the inflammatory response and immune cell proliferation, and had inhibitory effect on trophoblast cell invasion.