摘要
研究广叶绣球菌多糖(polysaccharides,SLPs)对小鼠CD8^(+)T细胞活化、增殖分化以及效应阶段的影响,探索广叶绣球菌多糖对CD8^(+)T细胞的免疫应答调控机制。通过腹腔注射环磷酰胺溶液(80mg/kg)构建免疫低下小鼠模型,灌胃广叶绣球菌多糖(低、中和高剂量分别为100、200和400 mg/kg),连续饲养20 d后处死,取脾脏和胸腺。血细胞分析仪测定血常规;MTT检测脾T淋巴细胞的扩增能力;流式细胞术分析脾脏CD8^(+)T细胞表面CD28、CTLA-4和PD1以及树突状细胞表面MHCⅠ、CD80和PD-L1的表达量;ELISA测定外周血中TNF-α、IFN-γ、IL-2、IL-10、穿孔素及颗粒酶B的含量;RT-PCR测定脾脏Psmb9、Psmd9、TAP1、TAP2、IFN-γ、IL-2、JAK1、JAK2、STAT1、STAT3、SOCS-3和Blimp-1的mRNA表达量;Westernblot测定pJAK2/JAK2、pSTAT3/STAT3和Blimp-1的蛋白表达量。试验表明广叶绣球菌多糖高剂量组脾脏指数显著升高,外周血中红细胞、白细胞、淋巴细胞、单核细胞和血小板的含量显著升高(P<0.05或P<0.01);多糖中、高剂量组脾T淋巴细胞的扩增能力显著增强(P<0.05);中、高剂量组CD8^(+)T细胞含量显著降低,CD8^(+)T细胞表面PD-1分子含量显著降低,树突状细胞表面PD-L1分子含量显著降低、MHCⅠ细胞含量显著升高(P<0.05或P<0.01);多糖高剂量组TNF-α、IFN-γ、IL-2、穿孔素和颗粒酶B的含量显著升高(P<0.05或P<0.01),IL-10的含量显著降低(P<0.05);中、高剂量组Psmb9、Pmsd9、TAP1、TAP2和Blimp-1的mRNA含量显著升高(P<0.05或P<0.01),JAK1、JAK2、STAT1和STAT3的mRNA和蛋白表达量显著降低(P<0.05或P<0.01)。多糖中高剂量组Blimp-1蛋白表达量显著升高(P<0.05),多糖组与阳性对照组趋势一致。因此,认为广叶绣球菌多糖能够调节MHCⅠ类抗原加工呈递分子,并通过JAK2/STAT3信号通路促进CD8^(+)T细胞的活化、增殖分化和效应杀伤过程,从而调控CD8^(+)T细胞的免疫应答,起到改善小鼠免疫低下的作用。
This study was designed to investigate the effects of Sparassis latifolia polysaccharides(SLPs)on activation,proliferation and differentiation,and effect stage of CD8^(+)T cells in mice in order to explore the regulatory mechanism of SLPs in response to immune mediated by CD8^(+)T cells.The immunosuppressive mouse model was established by intraperitoneal injection of cyclophosphamide solution(80 mg/kg).SLPs(100,200 and 400 mg/kg as low,medium and high doses,respectively)was administered by gavage to alleviate the immunosuppression of mice.After feeding for 20 days,the mice were sacrificed,and the spleen and thymus were taken for further use.Blood routine was measured by hematology analyzer.MTT method was used to detect the expansion ability of splenic T lymphocytes.Flow cytometry was used to analyze the CD8^(+)T cells,the expression levels of CD28,CTLA-4 and PD1 on CD8^(+)T cells and MHCⅠ,CD80 and PD-L1 on dendritic cells.The content of TNF-α,IFN-γ,IL-2,IL-10,perforin and granase B in peripheral blood was determined by ELISA.The mRNA expression levels of Psmb9,Psmd9,TAP1,TAP2,IFN-γ,IL-2,JAK1,JAK2,STAT1,STAT3,SOCS-3 and Blimp-1 in spleen were determined by qRT-PCR.The protein expression levels of pJAK2/JAK2,pSTAT3/STAT3 and Blimp-1 were determined by Western blot.The results showed that the splenic index,and the content of red blood cells(RBC),white blood cells(WBC),lymphocytes,monocytes and platelets in peripheral blood were significantly increased in the SLPs high dose group(P<0.05 or P<0.01).The amplification ability of splenic T lymphocytes was significantly increased in SLPs medium and high dose groups(P<0.05).The content of CD8^(+)T cells,PD-1 on CD8^(+)T cells and PD-L1 on dendritic cells was significantly decreased in SLPs medium and high dose groups(P<0.05 or P<0.01),but the content of MHCⅠon dendritic cells was significantly increased(P<0.05).The content of TNF-α,IFN-γ,IL-2,perforin and granase B in SLPs high dose group was significantly increased(P<0.05 or P<0.01),while the IL-10 content was si
作者
贺楷雄
程艳芬
云少君
程菲儿
曹谨玲
冯翠萍
HE Kaixiong;CHENG Yanfen;YUN Shaojun;CHENG Feier;CAO Jinling;FENG Cuiping(College of Food Science and Engineering,Shanxi Agricultural University,Taigu 030801,Shanxi,China)
出处
《菌物学报》
CAS
CSCD
北大核心
2023年第5期1163-1174,共12页
Mycosystema
基金
山西省农谷建设研发计划项目(SXNGJSKYZX201903)
食用菌山西省科技创新重点团队(201805D131009)
山西省自然科学基金(202103021224126)。
