目的 探究空气负离子(negative air ions，NAIs)对原发性高血压大鼠(spontaneous hypertension rat，SHR)血压、氧化应激及炎症状态的影响及可能机制。方法 选择60只SHR(雌雄各半)，根据干预时间随机分为1个月组和3个月组，每组30只;各组大鼠根据每日干预时间再进一步随机分为SHR对照组、2 h NAIs-SHR组和6 h NAIs-SHR组，每组10只。另将20只SHR同系正常血压成年大鼠(Wistar-Kyoto，WKY)作为正常血压对照组，根据干预时间随机分为1个月WKY组和3个月WKY组，每组10只。2 h NAIs-SHR组和6 h NAIs-SHR组每天分别置于NAIs浓度为4。5×104～5×104个/cm3的环境中2 h和6 h，WKY组和SHR对照组每天置于正常空气环境。各组大鼠每3天测一次血压，每周测一次体重，于饲养第1个月和第3个月后处死，称量脏器湿重;采用酶联免疫吸附法检测血清8-羟脱氧鸟苷(8-hydroxylated deoxyguanosine，8-OHdG)、白细胞介素-6(interleukin-6，IL-6)、白细胞介素-8(interleukin-8，IL-8)、肿瘤坏死因子 α(tumor necrosis factor-α，TNF-α)、一氧化氮(nitric oxide，NO)和内皮素-1(endothelin-1，ET-1)水平;活性氧自由基(reactive oxygen species，ROS)检测试剂 盒测 定血清ROS水平;比色法检测血清丙二醛(malondialdehyde，MDA)、超氧化物歧化酶(superoxide dismutase，SOD)、还原型谷胱甘肽(glutathione，GSH)和氧化型谷胱甘肽(glutathione disulfide，GSSG)水平。HE染色观察各组大鼠胸主动脉组织病理形态，Western blot检测各组大鼠胸主动脉p38 丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase，p38 MAPK)、细胞外信号调节激酶(extracellular signal-regulated kinases，ERK)、c-Jun 氨基末端激酶(c-Jun N-terminal kinase，JNK)、c-fos、c-jun蛋白及其磷酸化蛋白的表达水平。结果 同周龄雄鼠WKY组体重较SHR对照组高，其他各组间体重差异无统计学意义。SHR组心脏脏器系数(4。66±0。48)较WKY组(3。73±0。15)高(P＜0。05)，其余大脑、肾脏、肝脏和脾脏脏器系数各组间差异无统计学意义。同周龄 WKY组血压均较SHR对照组低，第2～5、8～11周SHR对照组血压均较2 h NAIs-SHR组和6 h NAIs-SHR组高(P＜0。05)。HE染色发现2 h NAIs-SHR组和6 h NAIs-SHR组大鼠胸主动脉均较SHR对照组内膜结构紊乱、中膜增厚、外膜破裂等病理改变有不同程度的改善。氧化还原水平方面，与SHR对照组相比，6 h NAIs-SHR组ROS[0。66％±0。17％、0。49％±0。32％]、8-OHdG[(48。29±8。00)ng/mL、(33。13±14。67)ng/mL]水平均较低(P＜0。05)，1 个月 6 h NAIs-SHR 组 GSH/GSSG(10。08±4。93)较高;与 2 h NAIs-SHR 组相比，6 h NAIs-SHR 组 ROS 水平[0。99％±0。19％]较低(P＜0。05)。炎性因子水平方面，与SHR对照组相比，6 h NAIs-SHR组IL-8水平[(160。44±56。54)ng/L、(145。77±38。39)ng/L]均较低(P＜0。05)，1 个月 WKY 组 ET-1[(249。55± 16。98)ng/L]较高，各组间NO水平差异无统计学意义。1个月SHR对照组大鼠胸主动脉p-p38蛋白相对表达量较WKY组低(P＜0。05)，3个月SHR对照组大鼠胸主动脉p-p38和p-c-fos蛋白相对表达量均较2 h NAIs-SHR组和6 h NAIs-SHR组高(P＜0。05)。结论 4。5×104～5×104 个/cm3 浓度的 NAIs 干预可能通过 ROS/MAPK/AP1信号通路调节SHR大鼠部分氧化及炎症状态，进而降低其血压水平。
Effects and mechanisms of negative air ions on blood pressure,oxidative stress and inflammation levels in SHR rats
OBJECTIVE To investigate the effects and possible mechanisms of negative air ions(NAIs)on blood pressure,oxidative stress,and inflammatory status in spontaneous hypertension rats(SHR).METHODS A total of 60 SHR(half male and half female)were randomly divided into one-month and three-month groups,30 rats per groups,based on the duration of the intervention.Each group was further randomized into three groups based on the daily intervention time:SHR control group,2 h NAIs-SHR group,and 6 h NAIs-SHR group,10 rats per groups.In addition,20 Wistar Kyoto(WKY)(half male and half female),were randomized into one-month WKY group and three-month WKY group,10 rats per groups,based on the intervention time.The 2 h NAIs-SHR group and 6 h NAIs-SHR group were exposed to an environment with NAIs concentrations of 4.5×104-5×104 cm3 per day for 2 h and 6 h.The WKY group and SHR group were exposed to normal air on a daily basis.Blood pressure of rats in each group was measured every three days,while weight was measured once a week.After sacrificing the rats in the first month and the third month of rearing,wet weight of the organs was weighed.The enzyme linked immunosorbent assay(ELISA)was used to detect 8-hydroxylated deoxyguanosine(8-OHdG),interleukin-6(IL-6),interleukin-8(IL-8),tumor necrosis factor-α(TNF-α),nitric oxide(NO)and endothelin-1(ET-1)levels.Reactive oxygen species(ROS)detection kit was used to detect ROS level.Malondialdehyde(MDA)and superoxide dismutase(SOD),glutathione(GSH)and glutathione disulfide(GSSG)were measured by colorimetric analysis.HE staining was conducted to observe the histopathological morphological changes of the thoracic aorta in each group,and Western blot was conducted to detect the thoracic aortap38 mitogen-activated protein kinase(p38 MAPK),extracellular signal-regulated kinases(ERK),c-Jun n-terminal kinase(JNK),c-fos proteins,c-jun proteins and their phosphorylated proteins level.RESULTS The weight of WKY male mice in the same week age group was higher than that of SHR control group,and there was no significant difference in the weight between the other groups.The coefficient of heart in SHR control group(4.66± 0.48)was higher than that in WKY group(3.73±0.15)(P＜0.05),while there were no significant differences in the coefficients of brain,kidney,liver and spleen among the groups.Blood pressure in WKY group at the same age was lower than that in SHR group,and blood pressure in SHR control group at 2-5 and 8-11 weeks was higher than that in 2 h NAIs-SHR and 6 h NAIs-SHR groups(P＜0.05).HE staining showed that the internal,middle and external membranes of thoracic aorta in 2 h NAIs-SHR group and 6 h NAIs-SHR group were improved to varying degrees compared with those in SHR control group,including disordered internal membrane structure,thickened middle membrane and broken external membrane.In terms of oxidative stress levels,compared with the SHR control group,the ROS(0.66％±0.17％,0.49％±0.32％)and 8-OHdG((48.29± 8.00)ng/mL,(33.13±14.67)ng/mL)levels were lower in the 6 h NAIs-SHR group(P＜0.05),while the GSH/GSSG ratio was higher in the one-month 6 h NAIs-SHR group(10.08±4.93).Compared with the 2 h NAIs-SHR group,the ROS level(0.99％± 0.19％)was lower in the 6 h NAIs-SHR group(P＜0.05).In terms of inflammatory factor levels,compared with the SHR control group,the IL-8 levels((160.44±56.54)ng/L,(145.77±38.39)ng/L)were lower in the 6 h NAIs-SHR group(P＜0.05),while the ET-1 level((249.55±16.98)ng/L)was higher in the one-month WKY group.There was no significant difference in NO levels among the groups.The relative expression of p-p38 protein in the thoracic aorta of rats in the one-month SHR control group was lower than that in the WKY group(P＜0.05).The relative expression of p-p38 and p-c-fos proteins in the thoracic aorta of rats at three-months was higher in the SHR control group than in the 2 h NAIs-SHR and 6 h NAIs-SHR groups(P＜0.05).CONCLUSION The intervention of NAIs at a concentration of 4.5× 104-5× 104/cm3 may regulate the partial oxidation and inflammatory state of SHR rats through the ROS/MAPK/AP1 signaling pathway,thereby reducing their blood pressure level.
negative air ionsspontaneous hypertension ratreactive oxygen speciesmitogen-activated protein kinasestranscription factor activator protein 1
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