本文選題：熱缺血 + 家兔�。� 參考：《南昌大學(xué)》2014年碩士論文

【摘要】：目的： 建立家兔腎臟在體機(jī)械灌注與冷保存模型;低溫機(jī)械灌注對比傳統(tǒng)冷保存在邊緣供腎保存中的優(yōu)勢并探討其機(jī)理。試驗分為兩部分 第一部分：建立家兔DCD(Donation After Cardiac Death, DCD)腎臟在體機(jī)械灌注與冷保存模型。 方法：健康成年家兔10只，雌雄不限，體重（3.0±0.3）Kg，隨機(jī)分為低溫機(jī)械灌注組（MP組）與靜態(tài)冷保存組（CS組），每組5只。MP、CS組均開腹，游離腎動、靜脈，夾閉腎動、靜脈35分鐘，開放血流60分鐘后腎動脈穿刺置管行冷灌洗并將腎臟移至體外，MP組灌洗后接Lifeport（美國OrganRecovery Systems公司）,選擇20mmHg灌注壓，4小時低溫機(jī)械灌注；CS組灌洗后將腎臟放入器官保存液中并低溫保存4小時；每半小時測量一次兩組保存液的溫度。4小時后移除MP、CS組動靜脈所置導(dǎo)管并縫合血管，開放血流，左腎復(fù)灌、復(fù)溫，切除右腎，膀胱造瘺留置尿管。術(shù)前、術(shù)中2小時（B2）、術(shù)畢（B3）檢測三次血氣并據(jù)此調(diào)整水、電解質(zhì)與酸堿平衡。夾閉腎動、靜脈期間每5分鐘記錄一次體溫。MP組器械灌注期間每小時記錄一次灌注流量。術(shù)后記錄24小時尿量與生存率。 第二部分：機(jī)械灌注與冷保存對免腎保存效果比較研究 方法：健康成年家兔15只，雌雄不限，體重（3.0±0.3）Kg，隨機(jī)分為假手術(shù)組(SO組)、低溫機(jī)械灌注組（MP組）及冷保存組（CS組），每組5只。SO組開腹游離腎動、靜脈； MP組復(fù)制機(jī)械灌注模型，CS組復(fù)制冷保存模型。24小時后摘取腎臟標(biāo)本，心臟取血液標(biāo)本。檢測血清指標(biāo)：肌酐(CRE)、尿素氮(BUN)（自動生化儀檢測）、L-6（酶聯(lián)免疫吸附測定法）。腎臟組織檢測指標(biāo)：ICAM-1mRNA轉(zhuǎn)錄水平（實時熒光定量PCR）。NF-κB p65蛋白的表達(dá)情況（免疫組織化學(xué)），腎組織病理學(xué)改變（HE染色，，光鏡觀察）。 結(jié)果： 第一部分：1、體溫：腎動、靜脈夾閉期間各組家兔體溫較為恒定，組間、組內(nèi)無明顯差異(P＞0.05)。 2、灌注流量：60min及此后的時間點流量顯著高于0點，具有統(tǒng)計學(xué)意義（P0.001）；其他各時間點灌注流量也高于0點，具有統(tǒng)計學(xué)意義（P0.05）。 3、24小時生存率：MP、CS組24小時生存率均為100%，無統(tǒng)計學(xué)差異(P＞0.05)。 第二部分：1、尿量: SO組、MP術(shù)后24小時尿量顯著高于CS組（P＜0.05）；MP組顯著高于SO組; 2、血清CRE、BUN：N、MP組CRE、BUN顯著低于CS組（P＜0.05＝；MP組中CRE、BUN水平顯著高于SO組（P＜0.05＝；3組組內(nèi)無顯著差異(P0.05)。 3、血清IL-6水平: IL-6表達(dá)CS組高于MP組與SO組，存在統(tǒng)計學(xué)差異(P0.05)；MP組顯著高于SO組(P0.05);3組組內(nèi)無顯著差異(P0.05)。 4、腎臟組織NF-κB p65蛋白表達(dá)免疫組化法檢測,采用Image-Pro Plus6.0圖像定量分析軟件，測量平均光密度值并對結(jié)果進(jìn)行定量分析。腎臟組織NF-κBp65蛋白表達(dá)CS組高于SO組、MP組，(P0.05)；差異有顯著性意義。SO組與MP組之間無顯著差異(P0.05)。 5、ICAM-1mRNA：RTFQ-PCR檢測，ICAM-1mRNA：轉(zhuǎn)錄水平CS組高于MP組與SO組(P0.05)，MP組高于SO組(P0.05)，均有顯著性意義。 6、腎臟組織學(xué)檢查：HE染色光鏡觀測，SO組腎臟組織結(jié)構(gòu)正常。MP組中腎小管上皮細(xì)胞輕度水腫，腎小管管徑增大，尤其近曲小管為甚。CS組中腎小管上皮細(xì)胞輕度水腫，近曲小管與遠(yuǎn)曲小管的管徑均變小。雖然MP、CS組腎小管上皮細(xì)胞均為輕度水腫但是CS組對比MP組則更為嚴(yán)重。 結(jié)論： 1、本實驗建立的家兔腎臟在體機(jī)械灌注與冷保存模型穩(wěn)定成功率高，實驗條件易于控制，制作方法簡單巧妙，適合家免在體腎灌注與冷保存系列研究。 2、低溫機(jī)械灌注可降低移植術(shù)后血清中CRE、BUN、IL-6水平，減少NF-κB與ICAM-1蛋白的表達(dá)，提示機(jī)械灌注相比靜態(tài)冷保存能夠更好保護(hù)腎臟功能。 3、機(jī)械灌注可能通過其對腎臟的微循環(huán)機(jī)械疏通作用，而改善移植腎臟的濾過功能。 4、機(jī)械灌注可能通過去除腎臟低溫保存時所產(chǎn)生的代謝物從而減輕移植術(shù)后的非特異性炎性損傷。
[Abstract]:Purpose :

To establish a model of rabbit kidney perfusion and cold preservation in vivo ; compare the advantages of low temperature mechanical perfusion with the traditional cold storage in the preservation of the marginal donor kidney and discuss its mechanism . The test is divided into two parts .

The first part : To establish the model of rabbit ' s DCD ( DCD ) kidney in vivo mechanical perfusion and cold storage .

Methods : 10 healthy adult rabbits were randomly divided into two groups : low temperature mechanical perfusion group ( MP group ) and static cold storage group ( CS group ) . Each group was divided into low temperature mechanical perfusion group ( MP group ) and static cold storage group ( CS group ) .
the kidney is put into an organ preservation solution after the CS group is filled and washed , and the kidney is stored at low temperature for 4 hours ;
The temperature of the two groups of preservation solution was measured every half an hour . After 4 hours removal of MP and CS group , the catheter was removed and the blood vessel , open blood flow , left renal complex irrigation , rewarming , right kidney and cystostomy indwelling catheter were removed . Before operation , three blood gases were recorded and the water and electrolyte were left in balance with acid and alkali . After operation , the temperature was recorded every 5 minutes during operation . The perfusion flow rate was recorded every hour during the infusion of MP group . The urine volume and survival rate were recorded 24 hours after operation .

The second part : comparative study on the effect of mechanical perfusion and cold preservation on kidney - free preservation

Methods : Fifteen healthy adult rabbits were randomly divided into sham operation group ( SO group ) , low temperature mechanical perfusion group ( MP group ) and cold storage group ( CS group ) .
In MP group , mechanical perfusion model and CS group were used to copy cold storage model . After 24 hours , kidney specimens were taken and blood samples were collected . Serum parameters were measured : creatinine ( CRE ) , urea nitrogen ( BUN ) ( automatic biochemistry analyzer ) , L - 6 ( enzyme linked immunosorbent assay ) . The expression of NF - 魏B P65 protein ( immunohistochemistry ) , pathological changes of renal tissue ( HE staining , light microscope observation ) were detected .

Results :

The first part : 1 , body temperature : the temperature of the rabbits in each group was relatively constant during the clamping and closing of the vein , and there was no significant difference between the groups and the group ( P > 0.05 ) .

2 . The flow rate of perfusion : 60min and later time point flow was significantly higher than 0 point , which had statistical significance ( P0.001 ) ;
The perfusion flow at other time points was also higher than 0 point , which had statistical significance ( P0.05 ) .

The 24 - hour survival rate of MP and CS group was 100 % and no statistical difference ( P > 0.05 ) .

The second part : 1 , urine volume : 24 hours after MP , the urine volume was significantly higher than that in CS group ( P < 0.05 ) ;
MP group was significantly higher than that in SO group .

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本文編號：1731205