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目的:通过体外活化Toll样受体3(TLR3),探索将Poly I:C(免疫佐剂)作为TLR3配体提高HBV宫内感染者TLR3蛋白含量,进而通过改变HBV宫内感染者DCs细胞及B细胞百分比,提供改善机体免疫状态的可能性。方法:采用面对面问卷调查方法连续收集152对HBsAg阳性孕妇及其新生儿流行病学资料,并采集新生儿脐血及股静脉血。分别采用酶联免疫吸附试验(ELISA)及实时荧光定量PCR检测新生儿出生24 h内外周血乙肝病毒标志物及HBV DNA含量,密度梯度离心法分离脐血单个核细胞(CBMCs)以进行体外培养,经HBV与Poly I:C刺激后,流式细胞术(FCM)检测新生儿CBMCs的TLR3蛋白含量及DCs细胞、B细胞的百分比。结果:HBV非宫内感染的TLR3蛋白含量在经HBV以及PolyI:C+HBV后呈现下降趋势。而HBV宫内感染组呈现先下降后升高的趋势。HBV非宫内感染组与宫内感染组CBMCs经PolyI:C+HBV刺激活化后,两组TLR3蛋白含量、mDC、pDC比例差异均无统计学意义(t=0.11,P=0.91;t=0.46,P=0.64;t=0.21,P=0.23);而两组B细胞百分比差异有统计学意义(t=2.36,P=0.02)。HBV宫内感染组经Poly I:C+HBV刺激后与非宫内感染组经HBV刺激后的B细胞百分比比较差异亦有统计学意义(t=2.45,P=0.02)。结论:Poly I:C作为免疫佐剂一定程度上提高HBV宫内感染者的TLR3蛋白水平,使宫内感染组的TLR3蛋白水平逐渐接近于非宫内感染组,进而改变CBMCs中DCs细胞及B细胞百分比含量,为改善HBV宫内感染新生儿的免疫状况使其接近非宫内感染者水平提供可能。
OBJECTIVE: To investigate the effect of poly I: C (TLR3) ligand on the expression of TLR3 protein in HBV intrauterine infection by in vitro activation of Toll-like receptor 3 (TLR3) B cell percentage, to provide the possibility of improving the body’s immune status. Methods: The epidemiological data of 152 pairs of HBsAg positive pregnant women and their newborns were collected continuously by face-to-face questionnaire and cord blood and femoral vein blood were collected. Serum levels of hepatitis B virus (HBV) and HBV DNA were detected by enzyme-linked immunosorbent assay (ELISA) and real-time fluorescence quantitative PCR (PCR) respectively within 24 h after birth, and cord blood mononuclear cells (CBMCs) were isolated by density gradient centrifugation . After stimulated by HBV and Poly I: C, the content of TLR3 and the percentage of DCs and B cells in neonatal CBMCs were detected by flow cytometry (FCM). Results: The content of TLR3 protein in non-intrauterine HBV infection showed a decreasing trend after HBV and PolyI: C + HBV. HBV intrauterine infection showed the trend of first decreasing and then increasing. There was no significant difference in TLR3 protein content, mDC and pDC between the two groups (t = 0.11, P = 0.91; t = 0.46) after stimulation with Polyl: C + HBV in non-HBV intrauterine infection group and intrauterine infection group , P = 0.64; t = 0.21, P = 0.23). The percentage of B cells in both groups had statistical significance (t = 2.36, P = 0.02). HBV intrauterine infection by Poly I: C + HBV stimulation and non-intrauterine infection after HBV stimulation of the percentage of B cells was also statistically significant difference (t = 2.45, P = 0.02). CONCLUSION: Poly I: C as an immunological adjuvant can increase TLR3 protein level in HBV intrauterine infection to a certain extent, and make TLR3 protein level in intrauterine infection group gradually close to non-intrauterine infection group, and then change the expression of DCs cells and B The percentage of cells, to improve the immune status of neonates with intrauterine infection of HBV to make it close to the level of non-intrauterine infection may provide.