基于CB2受体活化探讨CD4+T细胞分化缓解中性粒细胞大鼠哮喘模型的作用机制  

Study on the mechanism of CD4+T cell differentiation in alleviating neutrophil asthma rat model based on CB2 receptor activation

作　　者：莫濡冰[1] 黄琳惠[1] 蒙冲[1] 蔡兴俊[1] Mo Rubing;Huang Linhui;Meng Chong(Dept of Respiratory, Hainan Provincial People′s Hospital,Haikou 570311)

机构地区：[1]海南省人民医院呼吸科,海口570311

出　　处：《安徽医科大学学报》2021年第7期1057-1063,共7页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81860007)。

摘　　要：目的分析JTE-907(大麻素类CB2受体反向激动剂)对中性粒细胞大鼠哮喘模型的治疗效果,并通过观察其对CD4+幼稚T细胞分化影响来确定CB2受体活化的潜在治疗机制。方法采用大鼠脾脏分离的T淋巴细胞Th系特异性分化系统分析JTE907对T细胞亚型分化的影响。通过qRT-PCR检测Tbx21、Gata3、白介素(IL)-9、维甲酸受体相关孤儿受体(ROR)-C、FoxP3、CNR2 mRNA表达。将40只动物随机分为4组(n=10):正常对照组、模型组、JTE-907低剂量组(JTE-907-L,腹腔注射5 mg/kg)和JTE-907高剂量组(JTE-907-H,20 mg/kg)。除正常对照组外,其他组的大鼠建立中性粒细胞性哮喘模型。采用ELISA试剂盒检测血清中OVA特异性IgE和BALF中细胞因子水平,流式细胞术检测脾脏中Th17和Treg细胞百分比,Western blot检测肺组织中RORγt、Foxp3和STAT5蛋白的表达。结果qRT-PCR结果显示,JTE907在所有测试浓度下诱导FoxP3和CB2受体的表达。OVA致敏和LPS刺激后,模型组大鼠血清中OVA-sIgE水平上调(P<0.05),并且BALF中中性粒细胞数和干扰素(IFN)-γ、肿瘤坏死因子(TNF)-α、IL-17和IL-5水平增加(P<0.05)。JTE-907治疗逆转了上述变化(P<0.05)。此外,与模型组相比,JTE-907治疗组大鼠脾脏中Th17/Treg比例降低(P<0.05)以及肺组织中RORγt蛋白水平降低(P<0.05),而Foxp3蛋白和STAT5的磷酸化水平增加(P<0.05)。结论CB2受体的激活对Th-17介导的中性粒细胞哮喘大鼠有一定的保护作用,其作用机制包括改善Th17/Treg平衡以及调节其各自的细胞因子水平。Objective To evaluate the therapeutic value of JTE-907(cannabinoid CB2 receptor reverse agonists)in a model of neutrophilic asthma,and to determine the potential therapeutic mechanism of CB2 receptor activation by observing its effect on the differentiation of CD4^(+) naive T cells.Methods In order to study the effect of JTE907 on the differentiation of T cell subtypes,used an in vitro system of Th lineage-specific differentiation of naive CD4^(+) T lymphocytes isolated from the rat spleen.The expression of Tbx21,Gata3,interleukin(IL)-9,ROR-C,FoxP3,CNR2 mRNA was detected by qRT-PCR.40 rats were divided randomly into four experimental groups with 10 animals each:normal control group,model group,JTE-907 low-dose group(JTE-907-L,intraperitoneal injection of 5 mg/kg)and JTE-907 high Dose group(JTE-907-H,20 mg/kg).Except for the normal control group,rats at other three groups were established neutrophil asthma models.The concentration of serum OVA-specific IgE and BALF cytokine was detected by ELISA.The percentage of Th17 and Treg cells in spleen was analyzed by flow cytometry,and the expression of RORγt,Foxp3 and STAT5 protein in lung tissue was detected by Western blot.Results qRT-PCR analysis showed that JTE907 induced the expression of FoxP3 and CB2 receptors at all the concentrations tested.After OVA sensitization and LPS stimulation,the level of OVA-sIgE in the model group increased(P<0.05),and the number of neutrophils and the levels of interferon(IFN)-γ,tumor necrosis factors(TNF)-α,IL-17 and IL-5 in BALF significant increased(P<0.05).JTE-907 treatment significantly reversed the above changes(P<0.05).In addition,compared with the model group,the ratio of Th17/Treg in spleen of JTE-907 treatment group significantly decreased(P<0.05),and the level of RORγt protein in lung tissue significantly decreased(P<0.05),while the levels of Foxp3 and p-STAT5 significantly increased(P<0.05).Conclusion The activation of CB2 receptor has a protective effect in rat with Th-17 mediated neutrophilic asthma.The mechanism of J

关 键 词：CB2受体 中性粒细胞 大鼠 TH17/TREG 哮喘 

分 类 号：R562[医药卫生—呼吸系统]