穿孔素和颗粒酶B在急性期登革病毒患者体内的表达水平及调控机制初探

Regulation mechanisms and expression levels of perforin and granzyme B in acute dengue virus-infected patients

目的登革病毒(Dengue virus,DENV)感染引起的登革热和登革出血热已给世界带来极大危害,明确其感染引起的免疫病理损伤机制尤为关键。方法以DENV感染急性期患者样本为研究对象,抽取静脉血,利用密度梯度法提取外周单个核细胞(PBMCs),流式细胞术检测PBMCs各亚群表达穿孔素和颗粒酶B的水平,进而探讨DENV急性期患者T细胞亚群和NK细胞亚群免疫功能的差异。结果DENV患者CD8^+T细胞和NK细胞内穿孔素和颗粒酶B的表达水平发生显著上调,细胞脱颗粒水平也显著增加;同时表达穿孔素或颗粒酶B的NK细胞比例显著高于CD8^+T细胞。另外,本研究还发现三个重要miRNAs(miR-27a*、miR-30e和miR-378)在DENV患者体内的表达量明显降低,已有的研究表明其可能靶向调控穿孔素或/和颗粒酶B的表达。结论DENV感染可诱导CD8+T细胞和NK细胞发挥效应功能,并依赖穿孔素和颗粒酶途径杀伤被病毒感染的靶细胞;DENV患者体内PRF1和GrzB的表达很可能受到miR-27a*、miR-30e和miR-378的靶向调控。通过上述研究为阐明T细胞和NK细胞在DENV感染过程中所发挥的免疫效应功能提供更多的理论基础。

Objective Dengue fever and dengue hemorrhagic fever caused by Dengue virus infection have brought great harm to the world.The mechanism of immunopathological damage caused by Dengue virus infection remains to be further studied.Methods In this study,peripheral mononuclear cells(PBMCs)were extracted from venous blood of patients with acute DENV infection by density gradient method.The expression of perforin and granzyme B in PBMCs was detected by flow cytometry,and then the difference of immune function between T cell subsets and NK cell subsets in DENV patients in acute phase was analyzed.Results The levels of perforin and granzyme B in CD8+T cells and NK cells of DENV patients were significantly up-regulated,and the degranulation level of these toxic killer cells was also significantly increased.At the same time,the percentage of NK cells expressing perforin or granzymeB was significantly higher than that of CD8+T cells.In addition,the expression of three microRNAs(miR-27 a*,miR-30 e and miR-378)were significantly reduced,which potentially target and regulate perforin or/and granzyme B expression.Conclusion These results showed that CD8+T cells and NK cells can be activated to function effectively during DENV infection,and kill target cells infected by the virus may depend on perforin and granzyme pathways.Moreover,the expression of PRF1 and GrzB in DENV patients may be regulated by the targeted molecules,such as miR-27 a*,miR-30 e and miR-378.These results provide more theoretical basis for clarifying the immune function of T cells and NK cells during DENV infection.