支气管哮喘合并焦虑小鼠大脑中神经肽S的含量及其与炎性因子的关系

The levels of neuropeptide S in the brain of asthmatic mice with anxiety and the relationship with the inflammatory mediators

目的 探究支气管哮喘（哮喘）合并焦虑小鼠模型大脑中神经肽S含量的变化,并在体外实验研究炎性因子对神经元分泌神经肽S的影响.方法 将40只BALB/C小鼠按随机数字表法随机分为对照组、哮喘组、焦虑组以及哮喘合并焦虑组,实时荧光定量PCR法检测各组动物模型脑组织中神经肽S mRNA相对表达量;体外原代培养大鼠大脑皮层神经元细胞,分为磷酸盐缓冲液（PBS）对照组、IL-1β组、IL-6组和肿瘤坏死因子（TNF）-α组,用炎性因子刺激神经元后,QRT-PCR法检测各组神经肽S mRNA表达水平,ELISA法检各组细胞培养液中神经肽S的含量.结果 各组动物模型大脑中神经肽S mRNA的相对表达量,焦虑组（0.88 ±0.05）和哮喘合并焦虑组（0.79 ±0.03）明显低于对照组（1.01 ±0.05）和哮喘组（0.96±0.06）,并且哮喘合并焦虑组明显低于其他3组,差异均有统计学意义（均P ＜0.05）.IL-6组和TNF-α组神经肽S mRNA相对表达量分别为0.88±0.07和0.86±0.07,均明显低于PBS对照组（1.00±0.06）和IL-1β组（0.94 ±0.08）,差异均有统计学意义（均P＜0.05）;IL-1β组神经肽S mRNA相对表达量与PBS对照组相比,差异无统计学意义（t=-1.81,P=0.079）.IL-6组和TNF-α组培养液中神经肽S含量分别为（45.4±1.2） ng/L和（46.0±1.0） ng/L,均明显低于PBS对照组的（50.6±1.8） ng/L和IL-1β组的（49.5±1.0） ng/L,差异均有统计学意义（均P＜0.05）;IL-1β组培养液中神经肽S含量与PBS对照组相比,差异无统计学意义（t=-1.89.P=0.067）.结论 哮喘血清中升高的IL-6、TNF-α可抑制神经元细胞分泌神经肽S,具有抗焦虑作用的神经肽S含量下降,易导致焦虑的发生,这可能是哮喘与焦虑共病的机制之一.

Objective To investigate the levels of neuropeptide S in the brain of asthmatic mice with anxiety and the effects of inflammatory mediatores on changes of neuropeptide S in in vitro experiments.Methods According to the random number table method,40 BALB/C mice were randomly divided into 4 groups：the control group,the asthma group,the anxiety group and the asthma and anxiety group.The relative expressions of neuropeptidc S mRNA in the brain tissue of each group were detected by quantitative real-time polymerase chain reaction （QRT-PCR）.Rat cortex neurons obtained by primary culture were divided into 4 groups：the PBS control group,the interleukin-1 beta group,the interleukin-6 group and the tumor necrosis factor-alpha group.After stimulation with inflammatory cytokines the mRNA expressions of neuropeptide S were measured by QRT-PCR and neuropeptide S levels in the cell culture supernatants were measured by emzyme linked immunosorbent assay （ELISA）.Results The relative expressions of neuropeptide S mRNA were decreased in the anxiety group（0.87 ± 0.05） and the asthma and anxiety group （0.79 ± 0.03） compared with the control group （1.00 ± 0.05） and the asthma group （0.96 ± 0.06）,most notably in the asthma and anxiety group （all P 〈 0.05）.Compared to the PBS control group [（1.00 ±0.06）,（50.6 ± 1.8） ng/L] and the interleukin-1 beta group [（0.94 ± 0.08）,（49.5 ± 1.0） ng/L],the levels of neuropeptide S mRNA and neuropeptide S were decreased in the interleukin-6 group [（0.88 ± 0.07）,（45.4 ± 1.2） ng/L] and the tumor necrosis factor-alpha group [（0.86 ± 0.07）,（46.0 ± 1.0） ng/L] （all P 〈 0.05）.There were no significant differences between the interleukin-1 beta group and the PBS control group（all P 〉 0.05）.Conclusions Up-regulated interleukin-6 and tumor necrosis factor-alpha in asthma can inhibit the secretion of neuropeptide S in neuronal cells.The decline of brain neuropeptide S,which has anti-anxiety effect,may lead to the occurrence of anxiety,which may be a potential mechanism of comorbidity of asthma and anxiety.