模擬高原低壓艙治療哮喘緩解期豚鼠對血清皮質(zhì)醇及嗜酸性粒細(xì)胞的影響

1、模擬高原低壓艙治療哮喘緩解期豚鼠對血清皮質(zhì)醇及嗜酸性粒細(xì)胞的影響    提要目的：探討模擬高原缺氧條件治療哮喘的機(jī)制，為臨床應(yīng)用模擬高原低壓艙治療兒童哮喘提供部分理論依據(jù)。方法：采用10%卵蛋白(OA)致敏和5次1%OA誘發(fā)復(fù)制哮喘豚鼠動物模型，設(shè)立正常對照組、哮喘對照組和低壓艙治療組3組。利用模擬4000m海拔高原條件的低壓艙治療干預(yù)，結(jié)束后3組均作外周血和BALF中EOS計(jì)數(shù)以及血清皮質(zhì)醇檢測，并進(jìn)行統(tǒng)計(jì)學(xué)分析。結(jié)果：哮喘對照組和低壓艙治療組外周血EOS、BALF中EOS絕對值均非常顯著高于正常對照組，低壓艙組非常顯著低于哮喘對照組(P<0.0

2、1)；哮喘對照組和低壓艙治療組的BALF中HEOS百分率均非常顯著高于正常對照組，低壓艙治療組非常顯著低于哮喘對照組(P<0.01)；正常對照組和低壓艙治療組的血清皮質(zhì)醇含量均非常顯著地高于哮喘對照組，但低壓艙治療組與正常對照組差異不顯著(P>0.05)。結(jié)論：模擬高原低壓艙治療能使哮喘緩解期豚鼠血清皮質(zhì)醇升高，進(jìn)而使其外周血EOS和BALF中EOS減少，有利于預(yù)防哮喘復(fù)發(fā)或減輕哮喘發(fā)作癥狀，可作為臨床預(yù)防和治療哮喘的一種新方法。關(guān)鍵詞低壓艙；豚鼠；哮喘；皮質(zhì)醇；嗜酸性粒細(xì)胞中圖法分類號R562.25;R845.22Effects of hypobaric chamber ther

3、apy on the content of serum cortisol and number of eosinophils in treatment of asthma in the remission stage in guinea pigsAbstractObjective: To explore the mechanism of the therapeutic effects of hypobaric chamber on asthma to provide theoretical basis for clinical practice. Methods: Guinea pigs we

4、re randomized into the normal control group (NCG), asthmatic group (AG) and hypobaric chamber-treated group (HCTG). The model of asthma was established in the animals in the latter two groups through sensitization and induction with 10% ovalbumin (OA) and 1% OA respectively. The guinea pigs in HCTG

5、were treated with hypobaric chamber simulating 4000 m altitude. The content of serum cortisol and the number of eosinophils (EOS) in peripheral blood (PB) and in bronchoalveolar lavage fluid (BALF) were determined. Results: The number of EOS in PB and BALF were significantly higher in AG and HCTG th

6、an in NCG, but significantly lower in HCTG than in AG (P<0.01). The percentage of HEOS in BALF was significantly higher in AG and HCTG than in NCG, but markedly lower in HCTG than in AG (P<0.01). The content of serum cortisol was remarkably higher in NCG and HCTG than in AG. However, no signif

7、icant difference was found between HCTG and NCG (P>0.05). Conclusion: After the treatment of asthma with hypobaric chamber, the content of serum cortisol in asthmatic guinea pigs was significantly increased to result in marked decrease of the number of EOS and the percentage of HEOS in PB and BAL

8、F. All these may be beneficial to prevent the relapse of asthma and alleviate asthmatic syndrome. Hypobaric chamber can be used as a new way for prevention and treatment of asthma.Key wordshypobaric chamber; asthma; cortisol; eosinophil; guinea pig哮喘是兒童較為常見的一種阻塞性肺疾病，嚴(yán)重影響身心健康。目前臨床上仍以皮質(zhì)激素作為控制哮喘癥狀的首選藥物

9、，而國內(nèi)外早有報(bào)道應(yīng)用高原氣候條件治療哮喘，取得良好療效。本實(shí)驗(yàn)利用模擬高原環(huán)境的低壓艙干預(yù)哮喘緩解期豚鼠，通過測定血清皮質(zhì)醇、外周血和支氣管肺泡灌洗液中EOS計(jì)數(shù)等，來探討低壓艙治療哮喘的機(jī)制，現(xiàn)報(bào)告如下。1材料和方法1.1動物分組和哮喘動物模型的復(fù)制1.2低壓艙治療低壓艙治療組豚鼠在5次哮喘誘發(fā)成功后，次日在模擬海拔4000m高原條件的動物低壓艙治療10d，2h/d。1.3檢測方法表1血清皮質(zhì)醇水平的測定方法Tab 1The procedure of determination of serum cortisol Type of tubes No.of tubes Diluent

10、 Standard Sample 125I-Cortisol Antibody ISR NSB 12 150 - - 100 - 1000 B0 34 50 - - 100 100 1000 S1S6 516 - 50 - 100 100 1000 U 17 - - 50 100 100 1000ISR：Immunological separating reagent搖勻，4放置15min，任取3管測總放射性計(jì)算均值(T)，3500r/min離心20min，棄掉上清液，采用放射免疫分析儀測定標(biāo)本放射性計(jì)數(shù)，并經(jīng)計(jì)算機(jī)處理，將結(jié)果打印出來。1.4統(tǒng)計(jì)學(xué)方法采用第三軍醫(yī)大學(xué)數(shù)學(xué)教研室提供的醫(yī)用統(tǒng)計(jì)

11、程序包SPMR中單因素方差分析(PDA-2)，實(shí)驗(yàn)數(shù)據(jù)用±s表示，應(yīng)用計(jì)算機(jī)作統(tǒng)計(jì)學(xué)處理。2結(jié)果外周血EOS計(jì)數(shù)(單位：×106/L)、BALF中EOS絕對值計(jì)數(shù)(單位：×106)和HEOS百分比以及血清皮質(zhì)醇含量(單位：g/L)比較見表2。表2外周血、BALF中EOS計(jì)數(shù)和HEOS百分比及血清皮質(zhì)醇水平Tab 2EOS count of PB and BALF, percentage of HEOS and content of serum cortisol NCG AG HCTG EOS count of PB 28.60±10.63

12、60;288.20±59.98? 138.60±27.54*# EOS count of BALF 2.08±0.41 6.42±2.01* 3.11±0.31*# Percentage of HEOS 0.28±0.11 0.68±0.07* 0.58±0.07*# Content of serum cortisol 390.57±86.39 118.81±58.81* 383.01±98.01#AG：Asthmatic group;HCTG:Hypobaric

13、chamber-treated group;NCG:Normal control group;?:P<0.01 vs NCG;#:P<0.01 vs AG3討論哮喘是一種以氣道高反應(yīng)性為主要特征的阻塞性肺疾病。這種氣道高反應(yīng)性與慢性變態(tài)反應(yīng)性氣道炎癥有關(guān)，且嗜酸性粒細(xì)胞(EOS)是重要的效應(yīng)細(xì)胞?，F(xiàn)有資料顯示，哮喘機(jī)體內(nèi)源性皮質(zhì)醇水平偏低或其受體功能缺陷而造成體內(nèi)糖皮質(zhì)激素相對不足是哮喘發(fā)作和惡化的一個(gè)原因，也是臨床上首選激素類藥物來控制哮喘發(fā)作的依據(jù)。然而長期使用激素治療哮喘也有其弊端，國外早有人利用高原環(huán)境治療哮喘，取得較好療效，并證實(shí)高原氣候條件能提高患者內(nèi)源性皮質(zhì)醇水平，降

14、低其外周血及BALF中EOS數(shù)，并降低氣道的高反應(yīng)性4，與國內(nèi)繆琪蕾5報(bào)道的結(jié)果基本一致。而且，中科院上海生理研究所和上海第六人民醫(yī)院合作利用模擬高原條件的低壓艙治療來預(yù)防哮喘復(fù)發(fā)取得較好療效，但治療機(jī)制未闡述。本實(shí)驗(yàn)通過OA致敏和誘發(fā)哮喘，成功地復(fù)制了豚鼠哮喘模型，其外周血EOS及BALF中EOS顯著增多。利用模擬高原條件的低壓艙治療哮喘緩解期豚鼠10d，能較為有效地提高血清皮質(zhì)醇水平，并降低外周血和BALF中的嗜酸性粒細(xì)胞數(shù)量，同時(shí)亦使HEOS不同程度下降，取得與國內(nèi)外高原環(huán)境治療哮喘基本相同的結(jié)果。推測其機(jī)理可能是通過低氧這一良性應(yīng)激原，刺激機(jī)體下丘腦-垂體-腎上腺軸等興奮而產(chǎn)生應(yīng)激反應(yīng)

15、，從而提高機(jī)體內(nèi)源性血清皮質(zhì)醇水平，進(jìn)而通過：抑制T淋巴細(xì)胞尤其是CD4+的T淋巴細(xì)胞活性，進(jìn)而降低全身和局部的IL-5、GM-CSF等水平；促進(jìn)淋巴細(xì)胞高表達(dá)Fas抗原，通過Fas/Fas L誘導(dǎo)EOS等炎癥細(xì)胞的凋亡6，7，共同抑制EOS活化、縮短其生存時(shí)間，促進(jìn)EOS凋亡。因此，活化的EOS(HEOS或EG2+)減少，可使其合成和釋放白三烯(LTs)、血小板活化因子(PAF)、主要堿性蛋白(MBP)、嗜酸性陽離子蛋白(ECP)、神經(jīng)毒蛋白(ENP)、過氧化酶(EPO)和氧自由基等毒性顆粒相應(yīng)減少8，同時(shí)，其表達(dá)IL-5和VLA-4等亦減少9，10，進(jìn)一步減輕了EOS在炎癥局部的粘附、聚集

16、、趨化和活化。此外糖皮質(zhì)激素可與細(xì)胞內(nèi)其受體結(jié)合，誘導(dǎo)一種分子量約(4045)×103u的蛋白質(zhì)-巨皮質(zhì)素(又稱調(diào)脂蛋白)合成，抑制磷脂酶A2活性而減少花生四烯酸的合成和釋放，從而減少了前列腺素(PGs)、白三烯(LTs)及血栓素(Tx)等炎性介質(zhì)的生成、釋放和激活，能較為有效地抑制變態(tài)反應(yīng)的強(qiáng)度11。三者共同減輕哮喘患者氣道變態(tài)反應(yīng)性炎癥，進(jìn)而降低氣道高反應(yīng)性，最終使哮喘病人減輕癥狀、減少復(fù)發(fā)。參考文獻(xiàn)1劉朝暉，徐軍，鐘南山.內(nèi)皮素、腫瘤壞死因子在哮喘發(fā)病中的作用.中華結(jié)核和呼吸雜志，1995，18(6)：3663Day R P.Eosinophil cell separation

17、 from human peripheral blood.Immunology,1970,18(2):9554De Bie T J, Hessel E M, Van Ark, et al. Effect of dexamethasone and endogenous corticosterone on airway hyperresponsiveness and eosinophilia in the mouse. Br J Pharmacology,1996,119(7):14845繆琪蕾.高原氣候降低居室塵埃螨過敏性哮喘小兒的周圍血T淋巴細(xì)胞活性、嗜曙紅細(xì)胞增多和支氣管阻塞.高原醫(yī)學(xué)雜志，

18、1995，5(3)：356Adachi T, Motojima S, Hirata A, et al. Eosinophil apoptosis caused by theophylline, glucocorticoids, and macrolides after stimulation with IL-5. J Allergy Clin Immunol,1996,98(6 pt2):S2077Matsumoto K, Schleimer R P, Saito H, et al. Induction of apoptosis in human eosinophils by anti Fas antibody treatment in vitro. Blood, 1995,86(2):14378Ackerm