高　煒

(北京市仁和醫(yī)院， 北京　102600)

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哮喘患者CD4+CD25+調(diào)節(jié)性T細胞比例與炎癥因子的相關(guān)性*

高煒

(北京市仁和醫(yī)院， 北京102600)

[摘要]目的： 探討哮喘患者外周血CD4+CD25+調(diào)節(jié)性T細胞比例與血清中炎癥因子的相關(guān)性。方法： 110例哮喘患者均分為哮喘急性發(fā)作組和哮喘緩解組， 55例健康體檢者作為對照組；用流式細胞儀測定哮喘患者外周血中CD4+CD25+調(diào)節(jié)性T細胞的比例，酶聯(lián)免疫法(ELISA)檢測被檢者血清中白細胞介素18(IL-18)、IL-4及轉(zhuǎn)化生長因子β(TGF-β)含量；用Spearman統(tǒng)計方法分析哮喘患者外周血中CD4+CD25+調(diào)節(jié)性T細胞的比例與血清中IL-18、IL-4和TGF-β含量的相關(guān)性。結(jié)果： 與對照組比較，哮喘緩解組、哮喘急性發(fā)作組患者全血細胞中CD4+CD25+ T細胞比例降低，血清中IL-18、IL-4及TGF-β含量升高(P<0.05)；與哮喘緩解組比較，哮喘急性發(fā)作組外周血中CD4+CD25+ T細胞的比例降低而血清中IL-18、IL-4及TGF-β含量升高(P<0.05)；Spearman相關(guān)分析結(jié)果顯示，哮喘患者外周血CD4+CD25+ T細胞比例均與血清IL-18、IL-4和TGF-β含量呈顯著負相關(guān)關(guān)系(r=-0.712、-0.629、-0.593，P=0.006、0.008、0.005)。結(jié)論： 外周血CD4+CD25+ T細胞的比例與哮喘的發(fā)生和發(fā)展密切相關(guān)，其機制可能與IL-18、IL-4和TGF-β升高有關(guān)。

[關(guān)鍵詞]哮喘； 白細胞介素類； 轉(zhuǎn)化生長因子β； CD4+CD25+調(diào)節(jié)性T細胞； 因素分析,統(tǒng)計學

哮喘是一種與氣道高反應性密切相關(guān)的慢性炎癥疾病，調(diào)節(jié)性T細胞(regulatory T cells)分化異常與哮喘的發(fā)病機制中的炎癥細胞及炎癥介質(zhì)有關(guān)[1]。尤其是CD4+CD25+T細胞以其獨特的作用方式和功能特征，在支氣管哮喘的發(fā)生發(fā)展中發(fā)揮著重要作用[2-4]。本研究通過檢測不同哮喘患者外周血CD4+CD25+T細胞的變化及其與炎癥因子水平的關(guān)系，探討CD4+CD25+T細胞在哮喘中的作用機制及其與炎癥反應的相關(guān)性。

1資料與方法

1.1一般資料

110例哮喘患者分為哮喘急性發(fā)作組和哮喘緩解組，每組55例。所有患者均符合中華醫(yī)學會呼吸病學分會制定的支氣管哮喘防治指南對哮喘的診斷標準及分級標準[5]，近1月內(nèi)未使用過其它哮喘治療類的藥物。排除合并心腦血管疾病、慢性支氣管炎、支氣管擴張、肺結(jié)核等其他肺部疾病的患者，排除近1月內(nèi)有呼吸道感染者，或近期口服或吸入糖皮質(zhì)激素的患者。哮喘急性發(fā)作組男31例，女24例，年齡41～72歲，平均(53.2±8.3)歲；哮喘緩解組男34例，女21例，年齡39～76歲，平均(51.9±6.8)歲。本研究得到醫(yī)院倫理委員會的批準，全部受試者均知情同意。另選取同期門診健康體檢者55例作為對照組，男34例，女21例，年齡39～76歲，平均(51.9±6.8)歲。對照組均無各器官系統(tǒng)急慢性疾病及過敏性疾病史。

1.2方法

全部受檢者均于清晨采集空腹靜脈血 5 mL，其中2.5 mL 3 000 r/min離心3 min分離血清，置于-20 ℃保存，用于檢測白細胞介素18(IL-18)、IL-4及轉(zhuǎn)化生長因子-β(transforming growth factor-β，TGF-β)含量，采用酶聯(lián)免疫法(ELISA)檢測，試劑盒購自Biosource 公司；另2.5 mL血液抗凝后，采用流式細胞儀(Beckman-CoulterXL100)測定CD4+CD25+T細胞數(shù)，計算外周血中CD4+CD25+T細胞比例。

1.3統(tǒng)計學方法

2結(jié)果

2.1CD4+CD25+調(diào)節(jié)性T細胞比例

與對照組比較，哮喘急性發(fā)作組、哮喘緩解組患者外周血中CD4+CD25+T細胞比例降低，哮喘急性發(fā)作組最低，差異有統(tǒng)計學意義(P<0.05)。見表1。

表1　3組受檢者外周血中CD4+CD25+調(diào)節(jié)性T細胞比例

(1)與對照組比較，P<0.05；(2)與哮喘緩解組比較，P<0.05

2.2IL-18、IL-4及TGF-β含量

哮喘急性發(fā)作組和哮喘緩解組血清中IL-18、IL-4及TGF-β含量高于對照組，差異有統(tǒng)計學意義(P<0.05)；哮喘急性發(fā)作組的IL-18、IL-4和TGF-β含量高于哮喘緩解組，差異有統(tǒng)計學意義(P<0.05)。見表2。

2.3CD4+CD25+T 細胞比例與血清炎癥因子含量相關(guān)性分析

Spearman相關(guān)分析結(jié)果顯示，哮喘患者外周血CD4+CD25+T細胞比例均與哮喘患者血清IL-18、IL-4和TGF-β含量呈顯著負相關(guān)關(guān)系(r=-0.712、-0.629、-0.593，P=0.006、0.008、0.005)。

表2　3組受檢者血清IL-18、IL-4及TGF-β水平

(1)與對照組比較，P<0.05；(2)與哮喘緩解組比較，P<0.05

3討論

哮喘是一種由多種細胞及細胞因子參與的慢性變態(tài)反應性氣道炎癥疾病，患者可因炎癥反應導致可逆性氣流受限，氣道縮窄及氣道重塑。1995 年首次報道CD4+CD25+T細胞以來，近年來CD4+CD25+T細胞在哮喘機制中的作用逐漸引起業(yè)內(nèi)學者的關(guān)注[6-7]。調(diào)節(jié)性T細胞可強烈抑制效應性T細胞，屬于可能抑制其它細胞功能的T細胞群體，CD4+CD25+T細胞可通過細胞毒T淋巴細胞抗原-4和糖皮質(zhì)激素誘導的腫瘤壞死因子受體(GITR)與靶細胞相應受體的結(jié)合，從而抑制效應性T細胞的增殖，還可通過與樹突狀細胞等抗原提呈細胞作用來調(diào)節(jié)機體免疫[8-9]。近年來多項研究表明，炎癥狀態(tài)下多種細胞如T細胞、巨噬細胞等炎性細胞均能分泌IL-18，使其水平顯著升高，而升高的IL-18能刺激 NK 細胞和 T 細胞產(chǎn)生更多Th1型細胞因子和Th2型細胞因子，促進IL-4分泌增加，加重炎癥反應[10-11]。TGF-β對維持外周CD4+CD 25+T細胞的數(shù)量、功能至關(guān)重要，研究證實TGF-β能使CD4+CD25-T細胞轉(zhuǎn)變?yōu)榫哂姓{(diào)節(jié)功能的CD4+CD25+T細胞，減低Th1型和Th2型細胞因子的產(chǎn)生，在哮喘慢性氣道炎癥中發(fā)揮重要作用[12-13]。

本研究結(jié)果顯示哮喘急性發(fā)作組、哮喘緩解組患者全血細胞中CD4+CD25+T細胞比例低于對照組，且哮喘急性發(fā)作組最明顯(P<0.05)；推測CD4+CD25+調(diào)節(jié)性T細胞參與介導機體對過敏原耐受的形成過程，通過抑制氣道的嗜酸性粒細胞炎癥反應，控制哮喘惡化，哮喘患者外周血 CD4+CD25+T細胞比例降低是哮喘的發(fā)病機制之一。本研究還發(fā)現(xiàn)，哮喘急性發(fā)作組、哮喘緩解組患者血清IL-18、IL-4和TGF-β含量高于對照組，哮喘急性發(fā)作組最高(P<0.05)。提示哮喘患者處于緩解期時，雖無任何臨床癥狀，但支氣管仍存在持續(xù)炎癥狀態(tài)，因此致使體內(nèi)血清炎癥因子的表達增加。此外，本研究將哮喘患者CD4+CD25+T細胞比例與患者外周血IL-18、IL-4和TGF-β含量進行相關(guān)性分析，結(jié)果發(fā)現(xiàn)哮喘患者外周血CD4+CD25+T細胞比例與患者血清IL-18、IL-4和TGF-β含量呈顯著負相關(guān)，提示隨著哮喘患者CD4+CD25+調(diào)節(jié)性T細胞比例降低，哮喘患者免疫抑制功能下降，減弱了對Th2細胞的抑制作用，使Th2型免疫反應增強，造成患者血清中IL-18、IL-4及TGF-β等細胞炎癥因子的生成增加，患者機體全身和局部免疫功能紊亂，加重了哮喘氣道炎癥癥狀[14-15]。

綜上所述， IL-18、IL-4和TGF-β炎性細胞因子參與哮喘的發(fā)病機制，且在炎癥發(fā)生時和調(diào)節(jié)性T細胞共同發(fā)揮抗炎致炎雙重作用，調(diào)控哮喘病情進展。

4參考文獻

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(2016-05-15收稿，2016-06-20修回)

中文編輯： 吳昌學； 英文編輯： 劉華

[中圖分類號]R562.25

[文獻標識碼]A

[文章編號]1000-2707(2016)07-0851-04

DOI:10.19367/j.cnki.1000-2707.2016.07.027

Correlation between the CD4+CD25+T Regulatory Cells Proportion and Serum Inflammatory Factors in Peripheral Blood of Asthmatic Patients

GAO Wei

(BeijingRenheHospital,Beijing102600,China)

[Abstract]Objective: To investigate the relationship between the CD4+ CD25+ T regulatory cells proportion and serum inflammatory factors in peripheral blood of asthmatic patients. Methods: 110 cases of patients with asthma were selected as the research subjects, and divided into asthma acute attack group (n=55) and asthma remission group(n=55). Another 55 healthy people were selected as control group. Enzyme linked immunosorbent assay (ELISA) was used to detect the content of IL-18, IL-4 and TGF-β, and flow cytometry was used to detect the proportion of CD4+ CD25+ T regulatory cells in peripheral venous blood. Spearman statistical method was adopted to analyze the correlation between the CD4+ CD25+ T regulatory cells proportion and serum inflammatory factors in peripheral blood of asthmatic patients. Results: Compared with control group, CD4+ CD25+ T regulatory cells proportion of blood cell in asthma acute attack group and asthma remission group was significantly decreased while the content of IL-18, IL-4 and TGF-β significantly was increased(P<0.05). Compared with asthma remission group, in asthma acute attack group the CD4+ CD25+ T regulatory cells proportion of blood cell was significantly decreased while the content of IL-18, IL-4 and TGF-β significantly was increased(P<0.05). The correlation analysis showed that the contents of IL-18, IL-4 and TGF-β in asthma patients were negatively correlated with the proportion of CD4+ CD25+ T regulatory cells (r=-0.712, -0.629, -0.593, P=0.006, 0.008, 0.005). Conclusions: The proportion of CD4+ CD25+ regulatory T cells is closely related to the occurrence and development of asthma, and its mechanism may be related to content increase of IL-18, IL-4 and TGF-β.

[Key words]asthma; interleukins; transforming growth factor β; CD4+CD25+ T regulatory cells; factors analysis, statistics

*網(wǎng)絡(luò)出版時間：2016-07-17網(wǎng)絡(luò)出版地址：http://www.cnki.net/kcms/detail/52.5012.R.20160717.1318.028.html