本文選題：右美托嘧啶 + COPD大鼠　； 參考：《鄭州大學》2014年碩士論文

【摘要】：背景與目的 慢性阻塞性肺疾�。–hronic obstructive pulmonary disease,COPD）是一種以氣流受限為特征、呈不完全可逆并伴有肺功能逐漸下降的慢性呼吸系統(tǒng)疾病。流行病學調查顯示，全球40歲以上的男性中有10%患有COPD。由于環(huán)境污染的加劇和吸煙等因素的影響，其發(fā)病人數(shù)仍在劇增，在發(fā)展中國家其發(fā)病率和死亡率增長速度較發(fā)達國家增加迅速。目前研究認為其發(fā)病機制主要為炎癥反應、氧化應激和蛋白酶-抗蛋白酶失衡等作用導致肺實質和氣道的病變[1]。 機械通氣是COPD患者急性加重期和全身麻醉過程中必不可少的輔助通氣措施，在保證機體氧供的同時，亦可加重患者的肺損傷。目前在機械通氣肺保護的研究中，主要集中于不同模式的機械通氣策略，以及糖皮質激素等藥物的應用，其目的是減少外界刺激損傷，阻斷炎癥反應和氧化應激的影響。 鹽酸右美托咪定是目前廣泛應用于ICU和麻醉科的一種新型的高選擇性α2-腎上腺素受體激動劑，具有鎮(zhèn)靜、鎮(zhèn)痛、催眠、抗交感和抑制血漿中兒茶酚胺釋放等作用。除此之外，近年的多項研究表明右美托咪定在神經(jīng)保護[2-4]、心肌保護[5，6]、腎臟保護[7]等重要器官的保護中有重要的作用。 目前，對于右美托咪定對肺臟的保護作用研究較少。本研究采用氣管內滴加脂多糖聯(lián)合被動香煙吸入法建立COPD大鼠模型，然后給予2h常規(guī)潮氣量機械通氣，通過不同劑量右美托咪定的干預處理，觀察右美托咪定對機械通氣COPD大鼠氧化應激的影響，探討鹽酸右美托咪定對COPD大鼠機械通氣時肺臟的作用機制。 材料與方法 雄性清潔級成年Wistar大鼠48只，平均體重為180±20g，購自河南省實驗動物中心。將所購大鼠隨機分為6組：正常組：D組、D1組，模型組：C組、C1組、C2組、C3組。模型組C組、C1組、C2組、C3組參照宋一平[8]方法復制COPD大鼠模型28d，正常組D組和D1組單純飼養(yǎng)28d。行大鼠肺功能檢測，評價模型復制成功與否。除D組和C組以外，每組大鼠行機械通氣2h，潮氣量設置為8ml/kg。C2組和C3組在通氣過程中分別給予1.0μg/kg/h和5.0μg/kg/h的右美托咪定稀釋液。D1組和C1組注射等量生理鹽水。通氣結束后各組大鼠取腹主動脈血3ml，靜止30min后離心取上清液，待做ELISA測定IL-β、TNF-α含量；取肺組織：左肺測定濕重后70℃恒溫箱放置48h至恒重，測干濕比W/D，右肺上葉和中葉液氮冷凍30min后置于-70℃冰箱保存，右肺上葉待做ELISA測定8-iso-PGF2α，右肺下葉石蠟包埋后切片待做免疫組化和HE染色。統(tǒng)計學分析 采用SPSS17.0統(tǒng)計分析軟件進行處理。符合正態(tài)分布的計量資料采用均數(shù)±標準差（X±S）表示，兩組間比較采用t檢驗，多組間比較采用單因素方差分析，兩兩比較方差齊者采用LSD法，不齊者采用Dunnett’s t-test。顯著性檢驗水準取α=0.05。 結果 1肺組織W/D變化 與正常組D組比較，模型組C組大鼠W/D增高，差異有統(tǒng)計學意義（P＜0.05）。COPD模型組C1組、C2組、C3組與正常組D1組大鼠比較，W/D的水平明顯升高，差異有統(tǒng)計學意義（P＜0.05）。C1組、C2組、C3組與正常組C組大鼠比較，W/D的水平明顯升高，差異有統(tǒng)計學意義（P＜0.05）。C1組與D1組相比，W/D明顯增加，差異有統(tǒng)計學意義（P＜0.05）。C2組與C1組相比，W/D明顯下降，差異有統(tǒng)計學意義（P＜0.05）。C3組與C2組相比，W/D明顯下降，差異有統(tǒng)計學意義（P＜0.05）。 2血清中TNF-α和IL-1β的變化 與正常組D組比較，模型組C組大鼠血清TNF-α和IL-1β明顯增高。D1組與D組相比，兩者表達量有所增加，但差異無統(tǒng)計學意義（P＞0.05）。COPD模型組C1組、C2組、C3組與正常組D1組大鼠比較，血清中IL-1β、TNF-α的水平明顯升高，差異有統(tǒng)計學意義（P＜0.05）。C1組、C2組、C3組與C組大鼠比較，血清中IL-β、TNF-α的水平明顯升高，差異有統(tǒng)計學意義（P＜0.05）。C2組與C1組相比，兩者表達量明顯下降，差異有統(tǒng)計學意義（P＜0.05）。C3組與C2組相比，兩者含量明顯下降，差異有統(tǒng)計學意義（P＜0.05）。 3肺組織8-異前列腺素變化 正常組大鼠D1組與D組相比，其表達量明顯增加，差異有統(tǒng)計學意義（P＜0.05）。COPD模型組C1組、C2組、C3組與正常組D1組大鼠比較，肺組織勻漿中8-iso-PGF2α的水平明顯升高，差異有統(tǒng)計學意義（P＜0.05）。C1組、C2組、C3組與正常組C組大鼠比較，肺組織勻漿中8-iso-PGF2α的水平明顯升高，差異有統(tǒng)計學意義（P＜0.05）。C2組與C1組相比，其表達量下降，但差異無統(tǒng)計學意義（P＞0.05）。C3組與C2組相比，其含量明顯下降，差異有統(tǒng)計學意義（P＜0.05）。 4肺組織結構變化 HE染色顯示結果為D組和D1組大鼠肺組織結構基本正常，肺紋理清晰，無肺大泡，但D1組大鼠肺組織有少許炎癥細胞浸潤。C組、C1組、C2組、和C3組肺紋理紊亂，，有明顯肺大泡生成，且有大量炎癥細胞浸潤。 5肺組織Nrf2和HO-1mRNA表達 與D組比較，C組大鼠肺組織Nrf2和HO-1mRNA表達水平增高，差異有統(tǒng)計學意義。機械通氣2h后，C1組、C2組、C3組大鼠肺組織Nrf2和HO-1mRNA表達水平顯著高于C組，差異有統(tǒng)計學意義。與C1組比較，C2組大鼠肺組織Nrf2和HO-1mRNA表達水平增加，差異無統(tǒng)計學意義。與C2組比較，C3組大鼠肺組織Nrf2和HO-1mRNA表達水平增加，差異有統(tǒng)計學意義。 6肺組織Nrf2和HO-1蛋白表達 與D組比較，C組大鼠肺組織中Nrf2和HO-1蛋白表達增高，差異有顯著性。與C組比較，C1組，C2組，C3組大鼠肺組織中Nrf2和HO-1蛋白表達顯著增強，差異具有顯著性。與C1組比較，C2組大鼠肺組織中Nrf2和HO-1蛋白表達增加，差異無顯著性。與C2組比較，C3組大鼠肺組織Nrf2和HO-1蛋白表達增加，有明顯差異。 結論 1、機械通氣對正常組大鼠肺臟氧化應激和炎癥反應無明顯影響，但可加重COPD大鼠肺臟的氧化應激反應和炎癥反應，加重肺損傷。 2、機械通氣過程中，右美托咪定可以通過降低炎癥因子表達和增強抗氧化應激作用減輕COPD大鼠肺損傷；且持續(xù)輸注5.0μg/kg/h的鹽酸右美托咪定較1.0μg/kg/h對COPD大鼠肺保護作用強。
[Abstract]:Background and purpose
Chronic obstructive pulmonary disease (Chronic obstructive pulmonary disease, COPD) is a chronic respiratory disease characterized by airflow limitation, which is not completely reversible with a gradual decline in lung function. Epidemiological investigation shows that 10% of men over 40 years of age suffer from the aggravation of environmental pollution and smoking and other factors of COPD.. The incidence of morbidity and mortality in developing countries is increasing rapidly in developing countries. It is believed that the main pathogenesis is inflammatory reaction, oxidative stress and protease anti protease imbalance in the lung parenchyma and airway disease [1].
Mechanical ventilation is an essential auxiliary ventilation measure in the acute exacerbation and general anesthesia of COPD patients. It can also aggravate the lung injury of the patients while ensuring the oxygen supply of the body. In the study of mechanical ventilation protection, the main focus is on the different modes of mechanical ventilation strategy and the application of glucocorticoid. It is to reduce external irritation injury, block inflammatory reaction and oxidative stress.
Dexmeimidine hydrochloride is a new type of high selective alpha 2- adrenergic agonist, which is widely used in ICU and anesthesiology. It has the effects of sedative, analgesic, hypnotic, anti sympathetic and inhibiting the release of catecholamine in plasma. In addition, a number of studies in recent years have shown that dexmedetomidin in neuroprotective [2-4] and myocardial protection of [5,6], Kidney plays an important role in protecting [7] and other important organs.
At present, there are few studies on the protective effect of dexmedetomidine on the lung. The COPD rat model was established by intratracheal drops plus lipopolysaccharide combined with passive cigarette inhalation, and then 2H conventional tidal volume mechanical ventilation was given, and right metomomidine was treated by different doses of dexmedetomidine to observe the oxidation of COPD rats with mechanical ventilation. To explore the mechanism of dexmedetomidine hydrochloride on the lungs of COPD rats during mechanical ventilation.
Materials and methods
48 adult male Wistar rats, with an average weight of 180 20g, were purchased from the experimental animal center of Henan province. The rats were randomly divided into 6 groups: normal group, group D, group D1, model group, group C, C1 group, C2 group, C3 group. The model group C group, C1 group, C2 group, C3 group refer to Song Yiping method, normal group and simple group simple 28d. rats were fed with lung function test to evaluate the success of model replication. Except for group D and group C, the rats in each group underwent mechanical ventilation 2H. The tidal volume was set in 8ml/kg.C2 group and C3 group in group 8ml/kg.C2 and 5 micron g/kg/h in.D1 group and C1 group, respectively. The rat aorta blood 3ml was taken from the abdominal aorta, and the supernatant was centrifuged after static 30min. The content of IL- beta and TNF- alpha was measured by ELISA. The lung tissue: the left lung was measured at 70 C after wet weight and placed 48h to constant weight, the dry and wet ratio W/D, the right upper lobe and the middle lobe liquid nitrogen frozen 30min were kept in the -70 centigrade refrigerator, and the right lung lobe was waiting for ELISA to determine 8-iso-PGF2 A and right lung. The paraffin embedded sections of lower lobe were stained with immunohistochemistry and HE staining.
SPSS17.0 statistical analysis software was used to deal with it. The measurement data conforming to normal distribution were represented by mean number + standard deviation (X + S). The two groups were compared with t test, the multiple groups were compared with single factor analysis of variance, and 22 were compared with the LSD method, and the non homogeneous people adopted the Dunnett 's t-test. significance test to take the alpha =0.05..
Result
1 W/D changes in lung tissue
Compared with the normal group D group, the model group C group W/D increased, the difference was statistically significant (P < 0.05).COPD model group C1 group, C2 group, C3 group and normal group D1 group compared with the normal group, the level of W/D significantly increased, the difference was statistically significant (P < 0.05).C1 group, compared with the normal group rats, the level was significantly higher, the difference was statistically significant. P < 0.05).C1 group compared with group D1, W/D was significantly increased, the difference was statistically significant (P < 0.05) the W/D decreased significantly in.C2 group and C1 group, and the difference was statistically significant (P < 0.05). The W/D significantly decreased in.C3 group and C2 group, and the difference was statistically significant (0.05).
Changes of serum TNF- A and IL-1 beta in 2 serum
Compared with the normal group D group, the serum TNF- A and IL-1 beta in the C group of the model group were significantly higher than those in the D group. The difference was not statistically significant (P > 0.05).COPD model group C1 group, C2 group, C3 group compared with the normal group rats, the level of serum beta and alpha was significantly higher, the difference was statistically significant (0.05). In group.C1, group C2, group C3 and group C, the level of IL- beta, TNF- alpha in serum was significantly higher, the difference was statistically significant (P < 0.05) compared with the C1 group, the expression of the two groups decreased significantly (P < 0.05) the difference was significantly lower than that in the C2 group, and the difference was statistically significant (0.05).
Changes of 8- isoprostaglandin in 3 lung tissues
The expression of D1 in the normal group was significantly higher than that in the D group, the difference was statistically significant (P < 0.05).COPD model group C1, C2, C3 group and normal group D1 group, the level of 8-iso-PGF2 alpha in lung homogenate was significantly higher, the difference was statistically significant (P < 0.05).C1 group, C2 group and normal group rats, lung tissue homogenization The level of 8-iso-PGF2 alpha in the pulp increased significantly (P < 0.05) (P < 0.05) the expression of.C2 group was lower than that of the C1 group, but the difference was not statistically significant (P > 0.05), the content of.C3 group was significantly lower than that of the C2 group, and the difference was statistically significant (P < 0.05).
4 changes in the structure of lung tissue
HE staining showed that the lung tissue structure was normal in group D and group D1, with clear lung texture and no pulmonary bullae, but there were a few inflammatory cells infiltrating.C group in group D1, C1 group, C2 group, and C3 group with lung texture disorder, obvious pulmonary bullae formation and large number of inflammatory cells infiltrating.
Expression of Nrf2 and HO-1mRNA in 5 lung tissues
Compared with group D, the expression level of Nrf2 and HO-1mRNA in lung tissue of rats in group C was higher, and the difference was statistically significant. After 2h of mechanical ventilation, the expression level of Nrf2 and HO-1mRNA in lung tissue of C1 group, C2 group and C3 group was significantly higher than that in C group, and the difference was statistically significant. Significance. Compared with group C2, the expression level of Nrf2 and HO-1mRNA in lung tissue of C3 group increased, the difference was statistically significant.
Expression of Nrf2 and HO-1 protein in 6 lung tissues
Compared with group D, the expression of Nrf2 and HO-1 protein in lung tissue of group C rats increased significantly. Compared with group C, the expression of Nrf2 and HO-1 proteins in lung tissues of group C1, C2 and C3 group increased significantly. The expression of Nrf2 and HO-1 protein in rat lung tissue increased significantly.
conclusion
1, mechanical ventilation has no obvious effect on the oxidative stress and inflammatory response in the lungs of normal rats, but can aggravate the oxidative stress reaction and inflammatory response in the lungs of COPD rats and aggravate the lung injury.
2, in the process of mechanical ventilation, dexmedetomidine can reduce the lung injury in COPD rats by reducing the expression of inflammatory factors and enhancing the anti oxidative stress, and the protective effect of right metomomidine on the lung of COPD rats is stronger than that of 1 u g/kg/h.

【學位授予單位】：鄭州大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R614

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