连翘酯苷A对内毒素致鸡脾淋巴细胞白介素-6的影响

Effect of Forsythiaside A on IL-6 Level in Lipopolysaccharide-treated Spleen Lymphocyte of Chickens

试验旨在研究内毒素和连翘酯苷A对鸡脾淋巴细胞中炎症因子白介素-6(IL-6)水平的影响,将1日龄雏鸡正常饲养至50日龄时,心脏采血处死,表面消毒后,剖检分离鸡脾脏,用脾淋巴细胞分离液分离脾淋巴细胞,将脾淋巴细胞共分为20个组,具体为4个剂量组×5个时间点=20个组。4个剂量组分别为:对照组、LPS组(5μg/m L)、连翘酯苷A低剂量预防组(5μg/m L LPS+200μg/m L连翘酯苷A)、连翘酯苷A高剂量预防组(5μg/m L LPS+400μg/m L连翘酯苷A)。5个时间点分别为:0、6、12、24、48 h,经内毒素和连翘酯苷A处理不同剂量和不同时间后,收集脾淋巴细胞,通过采用酶联免疫吸附试验(ELISA)和实时荧光定量PCR(q RT-PCR)方法检测脾淋巴细胞中IL-6水平。ELISA结果显示:与对照组相比,LPS组脾淋巴细胞中炎症因子IL-6蛋白含量升高;与LPS组相比,连翘酯苷A不同剂量预防组脾淋巴细胞中IL-6蛋白含量均下降,表明脾脏中IL-6蛋白含量的变化与连翘酯苷A的剂量呈正相关性,通过抑制LPS诱导的鸡脾脏中IL-6蛋白含量的升高,可减轻炎症反应。q RT-PCR结果显示:与对照组相比,LPS组脾淋巴细胞中IL-6 mRNA表达量升高;与LPS组相比,连翘酯苷A不同剂量预防组脾淋巴细胞中IL-6 mRNA表达量均下降,IL-6 mRNA表达量与连翘酯苷A的剂量呈负相关性,试验表明连翘酯苷A可以通过转录水平抑制LPS诱导的鸡脾淋巴细胞中IL-6 mRNA表达量的升高,减轻炎症反应。结果表明,连翘酯苷A抑制鸡脾淋巴细胞中IL-6水平,对免疫功能有一定免疫影响,具有抗炎功能。

This study was aimed to assess the effect of forsythiaside A on inflammatory cytokine interleukin-6（IL-6） level in lipopolysaccharide-treated spleen lymphocyte of chickens. Chicks were reared from the age of 1 to 50 days at normal management condition; at the age of 50 days, chickens were sacrificed and the heart blood samples were collected; the skin of the sacrificed chickens was disinfected and the spleen samples were collected; the spleen lymphocytes were subsequently isolated by lymphocyte separating medium; four treatment groups were set in this study, including the control group, LPS treatment group（5μg/m L）, low dosage forsythiaside A prevention group（5 μg/m L LPS＋200 μg/m L forsythiaside A）, and high dosage forsythiaside A prevention group（5 μg/m L LPS ＋400 μg/m L forsythiaside A）; the protein and m RNA expression level of IL-6 in spleen lymphocytes of different treatment groups were determined by ELISA and q RT-PCR assay respectively during a 48-hour time period at 0, 6, 12, 24 and 48 h. The ELISA assay results revealed that the level of IL-6 protein was found increased in the LPS group compared to the control group; both the low dosage and high dosage forsythiaside A prevention groups showed lower IL-6protein level compared to the LPS group, indicating that the change of the IL-6 protein level in chicken spleen was positively correlated with the dosage of forsythiaside A and forsythiaside A alleviated the inflammation in chicken spleen lymphocytes induced by LPS via inhibiting the increase of IL-6 protein level. The q RT-PCR results showed that LPS group had higher IL-6m RNA expression level compared to the control group; the level of IL-6 m RNA expression was lower in both the low dosage and high dosage forsythiaside A prevention groups compared to the LPS group, and the change of the IL-6m RNA expression level was negatively correlated with the dosage of forsythiaside A, which further demonstrated that forsythiaside A relieved the inflammation in chicken spleen lymphocytes induced by LPS through suppressing the increase of IL-6 m RNA expression in transcriptional level.The combined data suggest that forsythiaside A exerts anti-inflammation in lipopolysaccharide-treated spleen lymphocyte of chickens by inhibit the expression of IL-6 at transcriptional and translational level.