本文關(guān)鍵詞：大容量肺灌洗對(duì)家兔肺泡上皮claudin-4和E-cadherin表達(dá)的影響　出處：《西南醫(yī)科大學(xué)》2016年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 大容量肺灌洗 肺損傷 肺泡上皮屏障 緊密連接蛋白-4 E-鈣粘蛋白

【摘要】：目的:觀察大容量肺灌洗(whole lung lavage,WLL)對(duì)家兔肺泡上皮緊密連接蛋白(claudin-4)和E-鈣粘蛋白(E-cadherin)表達(dá)的影響,探討WLL相關(guān)性肺損傷的修復(fù)機(jī)制。方法:將15只家兔隨機(jī)分為對(duì)照組、大容量肺灌洗后2小時(shí)組(肺灌洗2h組)、大容量肺灌洗后72小時(shí)組(肺灌洗72h組),每組5只。行WLL前各組家兔均禁食12h,禁飲6h。肺灌洗2h組和肺灌洗72h組家兔耳緣靜脈注入3%戊巴比妥(1mL/kg),待進(jìn)入全麻狀態(tài)后行氣管切開,插入氣管插管導(dǎo)管,確保左、右肺分隔良好,左肺給予氧氣吸入,氧流量控制在160~200mL/min,生理鹽水(溫度約37℃)反復(fù)灌洗右肺,灌洗總量按25~30mL/kg計(jì)算,每次灌入約10mL,生理鹽水沿灌洗導(dǎo)管注入(速度約5mL/min),每次注入完畢后將家兔處于頭低足高位,讓灌洗液自然引流干凈,重復(fù)上述操作,灌洗液回收率均在90%以上,后拔出氣管插管,逐層縫合,術(shù)后常規(guī)處理,肺灌洗72h組家兔清醒后可逐漸恢復(fù)正常飲食。對(duì)照組家兔進(jìn)入全麻狀態(tài)后行氣管切開,插入氣管插管導(dǎo)管,確保左、右肺分隔良好,左肺給予氧氣吸入,氧流量控制在160~200mL/min,右肺不予以灌洗。對(duì)照組家兔經(jīng)以上操作后2小時(shí)行肺部CT檢查,后股動(dòng)脈放血處死家兔;肺灌洗2h組和肺灌洗72h組分別于設(shè)定時(shí)間點(diǎn)前15min再次麻醉家兔,行肺部CT檢查后股動(dòng)脈放血處死家兔。測(cè)量右肺濕/干重比,取部份右肺組織行石蠟切片,he染色觀察肺組織的病理改變并進(jìn)行評(píng)分;電鏡觀察肺泡上皮的超微結(jié)構(gòu);免疫組化檢測(cè)肺組織claudin-4及e-cadherin的表達(dá),其相對(duì)表達(dá)量用平均光密度值(od值)表示。結(jié)果:(1)家兔表現(xiàn):肺灌洗2h組及肺灌洗72h組家兔大容量肺灌洗時(shí)出現(xiàn)呼吸急促,唇鼻部皮膚發(fā)紺明顯,偶有嗆咳等不適反應(yīng),術(shù)畢后上述癥狀逐漸消失,肺灌洗72h組術(shù)后清醒后飲食、活動(dòng)無明顯異常;對(duì)照組家兔無上述反應(yīng)。(2)家兔肺部ct改變:肺灌洗2h組家兔左側(cè)肺組織未見明顯異常,但右肺組織見不均勻磨玻璃樣浸潤(rùn)影;對(duì)照組和肺灌洗72h組家兔雙肺組織肺部ct檢查未見明顯異常改變。(3)肺組織的大體表現(xiàn):肺灌洗2h組家兔右肺組織體積增大,彈性減退,部份色澤暗紅,表面可見散在出血點(diǎn),切面可見少量泡沫狀液體流出,左側(cè)肺組織未見明顯異常;肺灌洗72h組及對(duì)照組兩側(cè)肺組織表面光滑,彈性好,無明顯出血點(diǎn)及水腫。(4)肺濕/干重比(w/d):與對(duì)照組肺w/d值(4.34±0.06)比較,肺灌洗2h組的肺w/d值(4.72±0.09)明顯增高,差異有統(tǒng)計(jì)學(xué)意義(p0.05);與肺灌洗2h組比較,肺灌洗72h組的肺w/d值(4.36±0.07)明顯降低,差異有統(tǒng)計(jì)學(xué)意義(p0.05);肺灌洗72h組w/d值較對(duì)照組高,但差異無統(tǒng)計(jì)學(xué)意義(p0.05)。(5)he染色:肺灌洗2h組家兔肺泡壁毛細(xì)血管擴(kuò)張充血,可見少量炎癥細(xì)胞浸潤(rùn),肺泡間隔增厚、局灶性肺水腫,肺泡腔內(nèi)可見淡紅色液體,但肺組織結(jié)構(gòu)完整;肺灌洗72h組家兔肺組織病理改變較肺灌洗2h組減輕,肺泡結(jié)構(gòu)完整,肺泡腔內(nèi)無滲液,但仍有少許炎癥細(xì)胞浸潤(rùn),對(duì)照組家兔he染色未見明顯異常改變。肺灌洗2h組的病理評(píng)分(5.0±0.7)明顯高于對(duì)照組(0.2±0.4)和肺灌洗72h組(1.2±0.4),差異有統(tǒng)計(jì)學(xué)意義(P均0.05);肺灌洗72h組病理評(píng)分較對(duì)照組高,差異有統(tǒng)計(jì)學(xué)意義(P0.05)。(6)電鏡顯示:對(duì)照組肺泡上皮細(xì)胞間緊密連接結(jié)構(gòu)完整;肺灌洗2h組細(xì)胞間緊密連接有破壞,細(xì)胞間隙增寬,部分I型肺泡上皮細(xì)胞斷裂;肺灌洗72h組上述改變減輕。(7)免疫組織化學(xué):對(duì)照組家兔claudin-4及E-cadherin主要在肺泡上皮細(xì)胞包膜呈連續(xù)的陽性表達(dá),包漿有少許表達(dá);肺灌洗2h組肺泡上皮細(xì)胞claudin-4及E-cadherin的表達(dá)均明顯低于對(duì)照組,為不連續(xù)表達(dá);肺灌洗72h組claudin-4及E-cadherin表達(dá)較肺灌洗2h組增強(qiáng),但仍低于對(duì)照組。肺灌洗2h組claudin-4及E-cadherin表達(dá)的OD值分別為(0.93±0.12)、(1.12±0.09),較對(duì)照組claudin-4及E-cadherin表達(dá)的OD值(1.44±0.05)、(1.74±0.05)及肺灌洗72h組claudin-4及E-cadherin表達(dá)的OD值(1.32±0.04)、(1.62±0.07)明顯降低,差異均有統(tǒng)計(jì)學(xué)意義(P0.05);與對(duì)照組比較,肺灌洗72h組claudin-4及E-cadherin表達(dá)的OD值仍有降低,差異有統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論:(1)大容量肺灌洗可導(dǎo)致家兔肺泡上皮屏障一定程度的損傷。(2)大容量肺灌洗后家兔肺泡上皮claudin-4及E-cadherin表達(dá)降低。(3)大容量肺灌洗相關(guān)性肺損傷可在一定時(shí)間內(nèi)自行修復(fù)。(4)Claudin-4及E-cadherin可能參與了肺灌洗液的清除及肺損傷的修復(fù)。
[Abstract]:Objective: To observe the effects of large volume lung lavage (whole lung, lavage, WLL) on rabbit alveolar epithelial tight junction protein (claudin-4) and E- cadherin (E-cadherin) expression, explore the repair mechanism of WLL induced lung injury. Methods: 15 rabbits were randomly divided into control group, large volume lung lavage group (2 hours after lung lavage group 2H), large volume lung lavage group (72 hours after lung lavage 72h group), 5 rats in each group. All rabbits were fasted before WLL 12h, 6h. 2H drink ban lung lavage group and lung lavage group 72h rabbit ear vein injection of 3% pentobarbital (1mL/kg), to enter the state of general anesthesia endotracheal open, insert the endotracheal tube, to ensure that the left and right lung left lung separated well received oxygen inhalation and oxygen flow control in 160~200mL/min, normal saline (temperature of about 37 DEG C) repeated lavage right lung lavage volume calculated according to 25~30mL/kg, every time pour about 10mL, saline lavage catheter implantation along (speed of about 5mL/min), after each injection in rabbits in the Trendelenburg position, let the natural drainage lavage fluid clean, repeat the above operation, lavage fluid recovery was above 90% after extubation, sutured, routine postoperative treatment, lung lavage in 72h group awake after gradually returned to normal diet. The rabbits in control group were in the state of general anesthesia after tracheotomy, endotracheal intubation catheter insertion, ensure that the left and right lung left lung separated well received oxygen inhalation and oxygen flow control in 160~200mL/min, not to be the right lung lavage. Control group rabbits after more than 2 hours after operation for lung CT examination after femoral artery bleeding of rabbits; lung lavage in 2H group and 72h group respectively in lung lavage set time before 15min again for lung CT anesthetized rabbits, rabbits were killed by exsanguination after measuring. The right lung wet / dry weight ratio, lung tissue of right part of paraffin section, he staining The pathological changes of lung tissues and score the ultrastructure of alveolar epithelium; electron microscope; immunohistochemistry the expression of claudin-4 and E-cadherin in lung tissue were determined, the relative expression for the average optical density (OD). Results: (1) in rabbits: shortness of breath appeared lung lavage in 2H group and 72h group lung lavage rabbit mwll, lip nasal skin cyanosis, occasional cough and other symptoms, the symptoms disappeared after surgery, lung lavage diet group 72h after awake, no obvious abnormal activity; control group rabbits without the above reaction. (2) the rabbit lung CT change: lung lavage in 2H group the left lung tissue had no obvious abnormalities, but the right lung tissue is not homogeneous ground glass opacity; control group and lung lavage in 72h group lung tissue of lung CT examination showed no obvious abnormal change. (3) the gross appearance of lung: lung lavage group 2H rabbits right lung tissue 縐澶，

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