發(fā)布時(shí)間：2018-06-18 02:48

  本文選題：慢性阻塞性肺疾病 + 肺動(dòng)脈高壓��； 參考：《廣州醫(yī)學(xué)院》2012年碩士論文

【摘要】：研究背景 慢性阻塞性肺疾病(COPD)是一種重要的慢性呼吸系統(tǒng)疾病，患病人數(shù)多，病死率高。由于其緩慢進(jìn)行性發(fā)展，嚴(yán)重影響患者的勞動(dòng)能力和生活質(zhì)量。COPD患者在急性發(fā)作期過后，臨床癥狀雖有所緩解，但其肺功能仍在繼續(xù)惡化，并且由于自身防御和免疫功能的降低以及外界各種有害因素的影響，經(jīng)常反復(fù)發(fā)作，而逐漸產(chǎn)生各種心肺并發(fā)癥。世界衛(wèi)生組織認(rèn)為，慢性阻塞性肺疾病可以預(yù)防，但目前無法治愈。治療有助于減緩病情發(fā)展，但這一疾病通常在一段時(shí)間后逐漸惡化。氣道炎癥是COPD的起始階段，氣道重塑是COPD氣流阻塞的病理基礎(chǔ)，是COPD病變持續(xù)發(fā)展的關(guān)鍵因素。因此，，研究其病理改變及發(fā)生機(jī)制對改善其防治效果及患者預(yù)后具有重要意義。慢性阻塞性肺疾病的一個(gè)重要的合并癥就是肺動(dòng)脈高壓，慢性阻塞性肺疾病合并肺動(dòng)脈高壓是逐漸發(fā)生和緩慢進(jìn)展的，肺動(dòng)脈高壓是慢性阻塞性肺疾病發(fā)展至肺源性心臟病的重要病理生理過程，如果肺動(dòng)脈壓持續(xù)升高，可導(dǎo)致右心負(fù)荷增加，最終導(dǎo)致右心衰竭。 研究表明Rho/Rho激酶信號通路是機(jī)體各組織細(xì)胞普遍存在的一條信號轉(zhuǎn)導(dǎo)通路，其通路的關(guān)鍵信號分子包括：Rho蛋白、Rho激酶和肌球蛋白磷酸酶。其通過調(diào)節(jié)細(xì)胞肌動(dòng)蛋白骨架的聚合狀態(tài)，參與調(diào)控了細(xì)胞形態(tài)維持、細(xì)胞粘附與遷移、細(xì)胞增殖與凋亡、基因轉(zhuǎn)錄、平滑肌收縮等多種生物學(xué)行為，介導(dǎo)了多種平滑肌與非平滑肌功能異常相關(guān)疾病的發(fā)生機(jī)制。在多種疾病動(dòng)物模型平滑肌細(xì)胞中均存在Rho激酶的過多表達(dá)。Rho/Rho激酶信號通路參與了COPD的發(fā)病過程，Rho激酶抑制劑可抑制炎癥細(xì)胞的遷移和趨化、拮抗炎性因子分泌，阻斷炎性因子作用，從而發(fā)揮抗炎作用，改善氣道重塑。 本研究主要研究Rho/Rho激酶信號通路在慢性阻塞性肺疾病(COPD)及COPD伴發(fā)肺動(dòng)脈高壓（PAH）患者血清Rho激酶1（ROCK1）及外周血單核細(xì)胞ROCK1的表達(dá)變化,為臨床防治COPD提供依據(jù),為COPD及伴發(fā)肺動(dòng)脈高壓提供新的治療靶點(diǎn)。 目的 通過酶聯(lián)免疫吸附法(ELISA)及細(xì)胞流式儀技術(shù)等檢測方法，檢測正常人群、慢性阻塞性肺疾病(COPD)伴發(fā)或不伴發(fā)肺動(dòng)脈高壓（PAH）患者的血清和外周血單核細(xì)胞中的Rho關(guān)聯(lián)含卷曲螺旋蛋白1（ROCK1）的表達(dá)變化，探討Rho/Rho激酶信號通路在COPD伴發(fā)或不伴發(fā)PAH患者發(fā)病中的作用。 方法 1.研究對象 收集2010年1月~2011年1月在廣州市第一人民醫(yī)院鶴洞分院收治的COPD患者40例，其中COPD患者20例，COPD伴發(fā)肺動(dòng)脈高壓(PAH)患者20例，其中COPD患者組符合中華醫(yī)學(xué)會(huì)呼吸分會(huì)2007年制定的慢性阻塞性肺疾病診治指南的診斷標(biāo)準(zhǔn)；COPD伴發(fā)肺動(dòng)脈高壓患者組同時(shí)符合中華醫(yī)學(xué)會(huì)呼吸分會(huì)2007年制定的慢性阻塞性肺疾病診治指南的診斷標(biāo)準(zhǔn)及ESC2009年制定的肺動(dòng)脈高壓診斷與治療指南的診斷標(biāo)準(zhǔn)。另取在本院體檢中心健康體檢者20例作為對照組。三組在年齡、性別、體重上盡量匹配。 2.肺功能測定 所有研究對象均需進(jìn)行肺功能檢測，主要的檢測指標(biāo)包括用力肺活量（FVC）、第一秒用力呼氣容積（FEV1）、FEV1/FVC及FEV1占預(yù)計(jì)值的百分比（FEV1％）、吸入支氣管擴(kuò)張劑（沙丁胺醇400ug）后的FEV1/FCV。 3.肺動(dòng)脈平均壓的測定 所有研究對象均通過心臟彩色多普勒超聲檢查，測得安靜狀態(tài)下的肺動(dòng)脈平均壓（mPAP）。 4.血清ROCK1的表達(dá)水平檢測 4.1標(biāo)本采集：抽取研究對象空腹靜脈血3ml，離心(3000R/15min.20℃)，留取上層血清1ml,于低溫－80℃冰箱中保存待測。 4.2測定方法：血清ROCK1的表達(dá)水平檢測方法為酶聯(lián)免疫吸附法（ELISA），其中標(biāo)本處理、測定方法和樣本濃度計(jì)算均按照相關(guān)試劑盒說明書進(jìn)行。 5.外周血單核細(xì)胞ROCK1的表達(dá)水平檢測 5.1.標(biāo)本采集：抽取研究對象空腹靜脈血20ml，以改進(jìn)的Ficoll-Hypaque密度分離法和塑料吸附法分離出單核細(xì)胞，于低溫－80℃冰箱中保存待測。 5.2測定方法：采用流式細(xì)胞儀檢測研究對象的外周血單核細(xì)胞ROCK1的表達(dá)水平。其中標(biāo)本處理、測定方法和樣本濃度計(jì)算均按照相關(guān)試劑盒說明書進(jìn)行。 結(jié)果 1.血清ROCK1表達(dá)水平在對照組、COPD組、COPD伴發(fā)PAH組分別為13.10±2.67、23.55±4.97和36.72±7.18pmol/L，COPD組、COPD伴發(fā)PAH組明顯高于對照組。三組之間存在顯著性差異（P0.01）。 2.外周血單核細(xì)胞ROCK1表達(dá)水平在對照組、COPD組、COPD伴發(fā)PAH組分別為3.61±2.44%、10.56±3.72%和31.21±8.83%，統(tǒng)計(jì)學(xué)分析顯示三組之間存在顯著性差異（P0.01）。 3.相關(guān)性分析顯示受試者血清ROCK1水平與肺動(dòng)脈平均壓呈正相關(guān)（r=0.654，P0.05），外周血單核細(xì)胞ROCK1表達(dá)水平與肺動(dòng)脈平均壓亦呈正相關(guān)（r=0.664，P0.05），血清ROCK1水平與外周血單核細(xì)胞ROCK1表達(dá)水平呈顯著正相關(guān)（r=0.864，P0.01）。 結(jié)論 Rho/Rho激酶信號通路的啟動(dòng)參與了COPD疾病肺動(dòng)脈高壓的形成和發(fā)展，血清中的ROCK1表達(dá)升高可能與ROCK1在單核細(xì)胞中的高表達(dá)有關(guān)。
[Abstract]:Research background
Chronic obstructive pulmonary disease (COPD) is an important chronic respiratory disease, the number of patients and the high mortality. Because of its slow progressive development, the patients' working ability and quality of life seriously affect the patient's working ability and quality of life (.COPD) after the acute attack period, although the clinical symptoms are slowly solved, but the lung function is still deteriorating, and because of its own The decline in defense and immunity, as well as the influence of various harmful factors on the outside world, often recurs and gradually produces various kinds of cardiopulmonary complications. The WHO believes that chronic obstructive pulmonary disease can be prevented but can not be cured at present. Treatment helps to slow the development of the disease, but the disease usually worsens after a period of time. Airway inflammation is the starting stage of COPD. Airway remodeling is the pathological basis of COPD airflow obstruction. It is the key factor for the continuous development of COPD lesions. Therefore, it is of great significance to study its pathological changes and mechanism to improve the effect of prevention and treatment and the prognosis of the patients. Pulmonary hypertension is a gradual and slow progression of pulmonary hypertension. Pulmonary arterial hypertension is an important pathophysiological process for the development of chronic obstructive pulmonary disease to pulmonary heart disease. If the pulmonary arterial pressure continues to rise, it can lead to an increase in the right heart load and eventually lead to right heart failure.
The Rho/Rho kinase signaling pathway is a common signal transduction pathway in the tissues and cells of the body. The key signaling molecules of the pathway include: Rho protein, Rho kinase and myosin phosphatase, which regulate cell morphology maintenance, cell adhesion and migration by regulating the polymerization of actin cytoskeleton. Multiple biological behaviors, such as cell proliferation and apoptosis, gene transcription and smooth muscle contraction, mediate the pathogenesis of a variety of smooth muscle and smooth muscle dysfunction related diseases. There is an overexpression of Rho kinase signaling pathway in the pathogenesis of COPD kinase in the smooth muscle cells of various disease animal models, and the inhibition of Rho kinase inhibition. It can inhibit the migration and chemotaxis of inflammatory cells, inhibit secretion of inflammatory factors and block inflammatory factors, thereby playing an anti-inflammatory role and improving airway remodeling.
This study mainly studies the changes of the expression of Rho/Rho kinase signaling pathway in serum Rho kinase 1 (ROCK1) and peripheral blood mononuclear cells (ROCK1) in patients with chronic obstructive pulmonary disease (COPD) and COPD associated pulmonary hypertension (PAH), providing a basis for clinical prevention and treatment of COPD and providing new therapeutic targets for COPD and associated pulmonary hypertension.
objective
The expression of Rho in serum and peripheral blood mononuclear cells in patients with chronic obstructive pulmonary disease (COPD) with or without pulmonary hypertension (PAH) was detected by enzyme linked immunosorbent assay (ELISA) and cell flow cytometry, and the Rho/Rho kinase signaling pathway was discussed in COPD The role of the patients with or without PAH.
Method
1. research objects
40 patients with COPD were admitted to Guangzhou No.1 People's Hospital in January 2010 ~2011, including 20 cases of COPD patients and 20 cases of COPD associated with pulmonary hypertension (PAH), of which the COPD patients were in accordance with the diagnostic criteria for the diagnosis and treatment of chronic obstructive pulmonary disease in 2007; COPD accompanied by lung movement. Patients with pulse hypertension conformed to the diagnostic criteria of the guidelines for the diagnosis and treatment of chronic obstructive pulmonary disease in 2007, and the diagnostic criteria for the diagnosis and treatment of pulmonary hypertension in ESC2009, which were formulated in the year of the Chinese Medical Association in 2007. In addition, 20 patients in the physical examination center of the medical center of our hospital were taken as the group. The three groups were in the age, sex, and weight as much as possible. Match.
2. pulmonary function measurement
All the subjects were required to perform lung function tests, including the forced expiratory volume (FVC), the first second forced expiratory volume (FEV1), the percentage of FEV1/FVC and FEV1 (FEV1%), and the FEV1/ FCV. after inhalation of bronchodilator (salbutamol 400ug).
3. pulmonary artery mean pressure measurement
The mean pulmonary arterial pressure (mPAP) was measured by color Doppler echocardiography in all subjects.
Detection of expression level of 4. serum ROCK1
4.1 specimen collection: extract the fasting venous blood 3ml of the subjects, centrifugation (3000R/15min.20 C), leave the upper serum 1ml, and store it in the low temperature 80 degree refrigerator.
4.2 determination method: the expression level of serum ROCK1 was detected by enzyme linked immunosorbent assay (ELISA), in which the sample treatment, determination method and sample concentration were calculated according to the instructions of the related kit.
Detection of expression level of ROCK1 in 5. peripheral blood mononuclear cells
Sample collection of 5.1.: extracting 20ml from fasting venous blood, and separating mononuclear cells by improved Ficoll-Hypaque density separation and plastic adsorption, and preserved in the refrigerator at low temperature to 80 degrees C.
5.2 determination method: flow cytometry was used to detect the expression level of ROCK1 in peripheral blood mononuclear cells of the study subjects. The sample treatment, determination method and sample concentration were calculated according to the instructions of the related kit.
Result
1. the level of serum ROCK1 expression in the control group, the COPD group and the group of COPD with PAH were 13.10 + 2.67,23.55 + 4.97 and 36.72 + 7.18pmol/L respectively. The COPD group and the COPD accompanying PAH group were significantly higher than those in the control group. There was a significant difference between the three groups (P0.01).
2. the expression level of ROCK1 in peripheral blood mononuclear cells was 3.61 + 2.44%, 10.56 + 3.72% and 31.21 + 8.83% in the control group, COPD group and COPD group with PAH, respectively. Statistical analysis showed significant difference between the three groups (P0.01).
3. correlation analysis showed that the serum ROCK1 level was positively correlated with the average pressure of pulmonary artery (r=0.654, P0.05), and the level of ROCK1 expression in peripheral blood mononuclear cells was also positively correlated with the average pressure of pulmonary artery (r=0.664, P0.05). The serum ROCK1 level was significantly positively correlated with the level of ROCK1 table of peripheral blood mononuclear cells (r=0.864, P0.01).
conclusion
The initiation of Rho/Rho kinase signaling pathway is involved in the formation and development of pulmonary hypertension in COPD disease. The increase of ROCK1 expression in serum may be related to the high expression of ROCK1 in mononuclear cells.
【學(xué)位授予單位】：廣州醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R563.9

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相關(guān)期刊論文 前1條

1 孫興珍;田向陽;;Rho激酶在肺動(dòng)脈高壓大鼠肺組織中的表達(dá)[J];河北醫(yī)藥;2008年11期

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