本文選題：封閉負壓引流技術　切入點：燒傷　出處：《第四軍醫(yī)大學》2014年碩士論文　論文類型：學位論文

【摘要】：熱能可引起燒傷部位即刻而不可逆性的壞死，鄰近該部位未直接燒傷部位也可產(chǎn)生損傷，而且損傷程度進行性加重，不斷加深擴大，最終導致組織壞死。這種現(xiàn)象的發(fā)生不僅加重了患者的病情，而且增加燒傷的致殘率及致死率，是燒傷治療面臨的問題之一。封閉負壓引流(vacuum-assisted closure，VAC)療法是利用負壓吸引裝置與特殊敷料結合后，在創(chuàng)面處產(chǎn)生低于大氣壓的壓力，促進創(chuàng)面愈合的技術，已廣泛應用于各類創(chuàng)面的治療。有研究證明封閉負壓引流治療可逆轉燒傷瘀滯區(qū)，，減少燒傷創(chuàng)面損傷深度及壞死面積，但缺乏進一步的機制研究。在燒傷瘀滯區(qū)進行性壞死過程中，血管內皮細胞既是損傷的主要靶細胞，也是病理進展的效應細胞。有效的保護血管內皮細胞，減輕其損傷及功能失常是逆轉燒傷瘀滯區(qū)的關鍵。自噬是指細胞消化自身多余、受損、衰老的蛋白質及細胞器，以維持內環(huán)境穩(wěn)定的細胞機制。本課題以大鼠梳狀燒傷模型應用封閉負壓引流技術進行治療，從內皮細胞功能和自噬水平的變化，初步探索VAC技術治療燒傷瘀滯區(qū)的效果和作用機制，為臨床治療提供理論依據(jù)。實驗共分以下四個部分： 1.建立大鼠銅梳燒傷模型。銅梳梳齒間未直接燒傷的間隙區(qū)即為瘀滯區(qū)。實驗動物隨機分為VAC組和對照組。對瘀滯區(qū)攝像，用圖像分析軟件，測量燒傷瘀滯區(qū)壞死面積的變化。HE染色觀察瘀滯區(qū)壞死深度。結果顯示：燒傷6小時后VAC組燒傷瘀滯區(qū)壞死面積及深度明顯小于對照組。 2. NO和ET-1含量檢測。我們用硝酸還原酶法檢測瘀滯區(qū)組織的一氧化氮（NO）的含量，用ELISA法檢測內皮素-1(ET-1)水平，對比VAC組和對照組之間兩者比值的差異，分析VAC對血管內皮細胞分泌功能的影響。結果顯示：燒傷后VAC組瘀滯區(qū)組織NO含量明顯多于傷前對照組，傷后12小時達到峰值，72小時仍高于正常值。對照組中NO含量低于VAC組，傷后6小時達到峰值，12小時即降低到正常含量以下，并持續(xù)下降。兩組傷后6小時及其后各時間點差異有統(tǒng)計學差異；燒傷后早期VAC組燒傷區(qū)瘀滯組織中ET-1含量較傷前對照組明顯上升，傷后24小時達到峰值，48小時后低于正常值。對照組燒傷瘀滯區(qū)組織中ET-1含量也上升，但上升程度低于VAC組，傷后6小時達到峰值，傷后12小時即降低到正常值以下。兩組傷后6、12、24小時差異有統(tǒng)計學意義（P＜0.05）。傷后早期VAC組ET-1/NO比值較低，12小時到達最低值后逐步回升，接近正常值。而對照組ET-1/NO比值持續(xù)升高，在72小時仍很高。兩組傷后6小時及其后各時間點差異均有明顯差異（P＜0.05）。 3. TM（血栓調節(jié)蛋白）和TM mRNA檢測。用免疫組化染色和實時熒光PCR檢測內皮細胞TM和TM mRNA表達，分析VAC治療對血管內皮細胞抗凝血、抗炎功能的影響。結果顯示：傷后各時間點VAC組瘀滯區(qū)組織均可見TM陽性表達。對照組傷后12小時可見TM陽性表達，但傷后24小時及其后時間點均為陰性。傷后各時間點，兩組TM mRNA表達均呈下降趨勢，但VAC組表達量高于對照組。傷后6小時及其后各時間點差異明顯（P＜0.05）。 4. LC3和Beclin-1表達檢測。免疫組化染色觀察燒傷瘀滯區(qū)LC3及Beclin-1的表達，對比VAC組與對照組之間的差異，分析VAC治療對燒傷瘀滯區(qū)細胞自噬的影響。結果顯示: VAC組瘀滯區(qū)LC3和Beclin-1表達率傷后6至24小時低于對照組，傷后48至72小時高于對照組，半定量分析顯示兩組在傷后6小時及其后各時間點差異有統(tǒng)計學意義（P＜0.05）。
[Abstract]:Heat can cause burns immediate and irreversible necrosis, adjacent to the site without direct burn can also cause damage, and the damage degree of progressive deepening, expanding, resulting in tissue necrosis. This phenomenon not only aggravate the patient's condition, but also increase the burn rate of morbidity and mortality, is one of the burn treatment problems. VSD (vacuum-assisted closure, VAC) therapy is the use of negative pressure suction device with special dressings combined, produce subatmospheric pressure on the wound, promote wound healing technique has been widely used in the treatment of various wounds. Studies have shown that vacuum sealing drainage treatment can reverse burn stasis stagnation zone, reduce the damage of deep burn wound and necrosis area, but the lack of further studies. In the process of burn necrosis of Stasis Zone, both vascular endothelial cells Is the main target of cell damage, is the pathological progress of effector cells. Vascular endothelial cell protection effectively, reduce the injury and dysfunction is the key to reverse the burn Stasis Zone. Autophagy is refers to the cell to digest their excess, damaged proteins and organelles of aging, to maintain a stable environment within the cellular mechanism of this subject. By using the rat comb burn model of vacuum sealing drainage for treatment, from the changes of endothelial cell function and autophagy, explore the effect and mechanism of VAC technique in the treatment of burn Stasis Zone, provide a theoretical basis for clinical treatment. The experiment is divided into the following four parts:
1. to establish a rat model of burn. Copper comb gap between copper comb comb is not directly burn the Stasis Zone. The experimental animal were randomly divided into VAC group and control group. The camera of the Stasis Zone, using image analysis software to observe the Stasis Zone of necrosis depth changes of.HE measuring burn Stasis Zone of necrosis area staining. The results showed that 6 hours after burn VAC group burn the Stasis Zone of necrosis area and depth significantly less than the control group.
2. the detection of NO and ET-1 content of nitric oxide. We detected by nitrate reductase method the Stasis Zone tissue (NO) were detected by ELISA et -1 (ET-1) level, the difference of the ratio between the two comparison between VAC group and control group, to analyze the effect of VAC on the secretion of vascular endothelial cells. The results showed that burn the content of NO in group VAC was significantly more than the Stasis Zone tissue injury control group before and 12 hours after injury and reached the peak, 72 hours is still higher than normal. NO content of control group was lower than that of VAC group, 6 hours after injury and reached the peak, 12 hours reduced to normal levels, and continued to decline. There were significant differences in 6 hours the two groups after injury and after each time point of difference; early lag in VAC group after burn in burn area and the content of ET-1 is the pre injury control group increased significantly, and reached the peak at 24 hours after injury, 48 hours less than normal. The control group also ET-1 content in blood stasis stagnation area burn The rise, but the rise is less than the VAC group, and reached the peak at 6 hours after injury, 12 hours after injury is reduced to below normal. Two groups of 6,12,24 hours after injury was statistically significant (P < 0.05). Early after injury VAC group ET-1/NO ratio is low, 12 hours to reach the lowest value gradually rise, close to normal value. While the control group, the ratio of ET-1/NO increased in 72 hours is still very high. 6 hours after injury and two groups had significant difference at each time point after the difference (P < 0.05).
3. TM (thrombomodulin). The expression of mRNA and TM detected by immunohistochemical staining and real-time fluorescence PCR detection of endothelial cells TM and TM mRNA analysis of VAC treatment on vascular endothelial cell function of anticoagulant, anti-inflammatory effects. The results showed that the positive expression of TM in the VAC group the Stasis Zone tissue could be seen after injury 12 hours after injury. The control group showed positive expression of TM, but 24 hours after injury and later time points were negative. At each time point, the expression of mRNA TM in two groups were decreased, but the expression of VAC group was higher than the control group. 6 hours after injury and after each time point difference (P < 0.05).
4. LC3 and Beclin-1 expression was detected by immunohistochemical staining. The expression of LC3 and Beclin-1 to observe the burn of the Stasis Zone, the difference between the VAC group and the control group comparison, analysis of effect of VAC treatment on burn cell Stasis Zone of autophagy. The results showed that VAC LC3 and Beclin-1 group the Stasis Zone expression rate after 6 to 24 hours compared with the control group, 48 hours after injury to 72 higher than the control group, semi quantitative analysis showed that the two groups in 6 hours after injury and after each time point had significant difference (P < 0.05).

【學位授予單位】：第四軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R644

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