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目的以日本血吸虫感染小鼠为模型,研究p47GTP酶家族成员IGTP及IRG-47缺失对巨噬细胞发育成熟及功能的影响。方法常规方法建立日本血吸虫感染小鼠模型,通过实时荧光定量PCR观察日本血吸虫不同感染时期igtp或irg47单一基因缺失的巨噬细胞igtp、lrg47和irg47的mRNA水平;流式细胞术检测巨噬细胞表面分子MHC-Ⅱ类(I-A)、CD16/32、CD40和CD80的表达;应用硝酸还原酶法检测感染急性期巨噬细胞产生炎症因子NO的能力。结果巨噬细胞igtp、irg47及lrg47基因在感染血吸虫后3周表达快速上调,感染后6周上调表达的幅度相差各异,IRG-47-/-小鼠igtp和lrg47转录明显上调。感染前和感染后3周IGTP-/-组的巨噬细胞表面CD16/32和CD40表达均较IRG-47-/-组和WT组高;感染后6周表达差异均无统计学意义(P>0.05)。在感染后6周,无有丝分裂原刺激情况下IGTP-/-组的巨噬细胞较IRG-47-/-组和WT组产生更高水平的NO;LPS刺激后,3组巨噬细胞产生的NO水平差异无统计学意义(P>0.05)。结论IGTP及IRG-47缺失不影响巨噬细胞发育成熟,也不影响巨噬细胞产生炎症因子NO的潜能,提示p47GTP酶可能具有独特的、不依赖NO的作用方式。
OBJECTIVE: To study the effects of IGTP and IRG-47 deletion in p47GTPase family on the maturation and function of macrophages in mice infected with Schistosoma japonicum. Methods The mouse model of Schistosoma japonicum infection was established by routine methods. The mRNA levels of igtp, lrg47 and irg47 were detected by real-time fluorescence quantitative PCR in igtp or irg47 single-gene-deficient macrophages at different infection stages. Flow cytometry was used to detect the expression of macrophages The expression of MHC class Ⅱ (IA), CD16 / 32, CD40 and CD80 were detected by enzyme-linked immunosorbent assay (ELISA). Nitric acid reductase was used to detect the production of inflammatory cytokines NO in acute macrophages. Results The expression of igtp, irg47 and lrg47 genes in macrophages were rapidly upregulated 3 weeks after infection with Schistosoma japonicum. The amplitude of up - regulation was different at 6 weeks after infection, and the transcription of igtp and lrg47 in IRG-47 - / - mice was significantly upregulated. The expression of CD16 / 32 and CD40 on macrophages before and 3 weeks after infection were higher than those in IRG-47 - / - and WT groups, but there was no significant difference at 6 weeks after infection (P > 0.05). At 6 weeks after infection, macrophages of IGTP - / - group produced higher levels of NO than IRG-47 - / - group and WT group without mitogen stimulation. After LPS stimulation, macrophages of 3 groups There was no significant difference in NO level (P> 0.05). Conclusions The deletion of IGTP and IRG-47 did not affect the maturation of macrophages, nor the potential of macrophage to produce inflammatory cytokines NO, suggesting that p47GTPase may have a unique and non-dependent NO mode of action.