本文選題：慢性胰腺炎 + SD大鼠　； 參考：《中南大學》2008年博士論文

【摘要】： 第一部分DBTC誘發(fā)慢性胰腺炎動物模型的建立 目的 應(yīng)用DBTC誘導建立SD大鼠CP模型,并加以兩種作用點不同的藥物干預,研究CP發(fā)生發(fā)展過程中病理學變化、纖維化程度、血請AMS變化及浸潤炎性細胞的作用。 方法 實驗組大鼠一次性尾靜脈注入0.8 mg/kg DBTC的80%乙醇溶液,2天后再隨機分為A組30只,不用任何藥物;B組26只,按6mg/kg.d行每天一次腹腔內(nèi)注射己酮可可堿注射液(PTX);C組37只,每周腹腔內(nèi)注射曲古霉素A(TSA)1ml(1μg/ml);對照組尾靜脈注入等量80%乙醇溶液。實驗組鼠分別于第14天、28天和56天處死,對照組鼠于第56天處死。處死鼠均行大體肉眼觀察及胰腺、肝、肺、腎鏡下病理學觀察及胰腺病理學評分;血清AMS含量由全自動生化分析儀測定,膠原纖維染色為VG染色法,肥大細胞為甲苯胺藍染色法;巨噬細胞、CD_4~+和CD_8~+染色均為EnVision免疫組化法。 結(jié)果 (1)大體觀察:A、B和C三組胰腺在14天、28天和56天處死鼠均出現(xiàn)不同程度的水腫、包膜張力增加、局部胰腺壞死出血及與周圍組織有粘連,28天和56天鼠部分胰腺出現(xiàn)局限性硬結(jié)節(jié)形成,以上表現(xiàn)B組和C組比A組輕。對照組胰腺均正常。實驗組胰腺外部分肝、肺出現(xiàn)出血、壞死及膿腫形成。 (2)鏡下觀察及評分:對照組胰腺組織學結(jié)構(gòu)正常。實驗組中56天處死鼠病理學評分、纖維化評分及CP發(fā)生率均明顯高于14天處死鼠(P＜0.05或P＜0.01)。實驗組之間CP發(fā)生率無明顯差異,但A組28天和56天處死鼠中至重度CP發(fā)生率明顯高于B和C組(P＜0.05)。實驗組胰腺外肝、肺鏡下表現(xiàn)相似于肉眼表現(xiàn)。 (3)血清AMS:各實驗組血清AMS含量均明顯高于對照組(P＜0.01),但各組之間無明顯差異(P＞0.05)。 (4)炎性細胞計數(shù):各實驗組56天處死鼠巨噬細胞和肥大細胞計數(shù)明顯高于14天處死鼠(P＜0.05)和對照組(P＜0.01);B和C組56天處死鼠CD_4~+計數(shù)明顯低于14天處死鼠(P＜0.01),但CD_8~+計數(shù)則相反(P＜0.01);實驗組之間CD_4~+/CD_8~+比值無明顯差異,但均明顯低于對照組(P＜0.05或P＜0.01)。 (5)相關(guān)性分析:實驗組病理學評分、纖維化評分、巨噬細胞計數(shù)和肥大細胞計數(shù)之間均呈密切正相關(guān)(P＜0.01);A組和C組病理學評分與CD_4~+計數(shù)均呈密切負相關(guān)(P＜0.05或P＜0.01);A組纖維化評分與CD_8~+計數(shù)呈密切負相關(guān)(P＜0.01)。 結(jié)論 (1) DBTC一次性尾靜脈注射能成功地建立SD大鼠CP模型,該模型具有操作簡單、建模時間短、CP發(fā)病率高及費用低等優(yōu)點。但DBTC對肝、肺等主要臟器有一定非致死性毒副作用。(2)該模型較符合人類CP病理形態(tài)學特征及其血清AMS含量改變。(3)胰腺組織內(nèi)浸潤的巨噬細胞、肥大細胞及CD_8~+細胞在該模型CP發(fā)生發(fā)展過程中起了重要作用。 第二部分大鼠慢性胰腺炎纖維化發(fā)生機制研究 目的 探討大鼠CP發(fā)生發(fā)展與炎性細胞、PSC及細胞生長因子的關(guān)系以及PTX、TSA干預大鼠CP的作用。 方法 實驗組和對照組大鼠胰腺組織經(jīng)4%甲醛固定后制作石蠟包埋切片。α-SMA和desmin染色為EnVision二步法,PDGF-BmRNA、TGF-β1mRNA、CTGFmRNA染色為原位雜交法。 結(jié)果 (1)炎性細胞計數(shù)見第一部分;(2)各實驗組不同時間點活化-PSC(a-PSC)計數(shù)均明顯高于對照組(P＜0.01);A組28天和56天處死鼠a-PSC計數(shù)明顯高于14天處死鼠(P＜0.05),B和C組56天處死鼠a-PSC計數(shù)明顯高于14天處死鼠(P＜0.05)。(3)實驗組各時間點靜止-PSC(s-PSC)計數(shù)均明顯低于對照組(P＜0.05),但三組各時間點之間均無明顯差異(P＞0.05)。(4) B、C兩組28天和56天處死鼠CTGFmRNA和PDGF-BmRNA表達陽性率及其評分均明顯高于對照組(P＜0.01);A組56天處死鼠TGF-β1mRNA表達陽性率及其評分均明顯高于對照組(P＜0.05或P＜0.01);A組和B組56天處死鼠TGF-β1mRNA表達陽性率明顯高于14天處死鼠(P＜0.05);A組28天和56天處死鼠CTGFmRNA和PDGF-BmRNA表達陽性率及其評分均明顯高于14天處死鼠(P＜0.05或P＜0.01)。(5)實驗組a-PSC、s-PSC計數(shù)及PDGF-BmRNA、TGF-β1mRNA、CTGFmRNA表達陽性率和評分之間均無明顯差異(P＞0.05)。(6)輕、中和重度CP a-PSC計數(shù)及TGF-β1mRNA、CTGFmRNA、PDGF-BmRNA表達評分和計數(shù)均高于正常組(P＜0.01);中度和重度CP a-PSC計數(shù)及CTGFmRNA、PDGF-BmRNA表達評分和計數(shù)均明顯高于輕度CP(P＜0.05或P＜0.01);輕度和中度CP s-PSC計數(shù)明顯低于正常組織(P＜0.01)。(7)實驗組中a-PSC計數(shù)及TGF-β1mRNA、CTGFmRNA、PDGF-BmRNA表達評分均與病理評分、纖維化評分呈密切正相關(guān)(P＜0.05或P＜0.01);實驗組中a-PSC計數(shù)與TGF-β1mRNA、CTGFmRNA、PDGF-BmRNA表達評分均呈密切正相關(guān)(P＜0.01)。 結(jié)論 (1)a-PSC參與了CP的纖維化過程,PDGF-BmRNA、TGF-β1mRNA、CTGFmRNA三種細胞因子能活化PSC及與胰腺纖維化關(guān)系密切。(2)肥大細胞和巨噬細胞與活化PSC有較密切關(guān)系。(3) CP發(fā)病機制復雜,單獨作用于某個環(huán)節(jié)的藥物難以對其發(fā)病進行控制,如PTX和TSA。
[Abstract]:Establishment of Animal Model of Chronic Pancreatitis Induced by the First Part of DBTC



Purpose



The CP model of SD rats was induced by DBTC , and two different drug interventions were used to study the changes of pathology , degree of fibrosis , AMS changes and infiltration inflammatory cells in the development of CP .



method



The rats in the experimental group were injected with 0.8 mg / kg DBTC 80 % ethanol solution , 2 days later , 30 rats were randomly divided into group A . Twenty - seven rats were given intraperitoneal injection of pentoxifylline injection ( PTX ) once a day .



Results



( 1 ) The rats of A , B and C showed different degrees of edema on 14 days , 28 days and 56 days . The results showed that there were localized hard nodules on the pancreas in both group A , B and C .



In the experimental group , the incidence of CP was significantly higher than that in control group ( P < 0.05 or P < 0.01 ) , but the incidence of CP in the experimental group was significantly higher than that in group B and C ( P < 0.05 ) .



( 3 ) AMS : AMS in the experimental group was significantly higher than that in the control group ( P < 0.01 ) , but there was no significant difference between the groups ( P > 0.05 ) .



( 4 ) Inflammatory cell count : The counts of mouse macrophage and mast cells were significantly higher than those in control group ( P < 0 . 05 ) and control group ( P < 0 . 01 ) , but CD _ 4 ~ + / CD _ 8 ~ + ratio was not significantly different between experimental group and control group ( P < 0 . 05 or P < 0 . 01 ) .



( 5 ) Correlation analysis : There was a close correlation between pathological score , fibrosis score , macrophage count and mast cell count in experimental group ( P < 0 . 01 ) ; the pathological score of group A and C was negatively correlated with the count of CD _ 4 ~ + ( P < 0 . 05 or P < 0 . 01 ) ; the fibrosis score of group A was negatively correlated with the count of CD _ 8 ~ + ( P < 0.01 ) .



Conclusion



( 1 ) The CP model of SD rats can be successfully established by DBTC single - use tail vein injection , and the model has the advantages of simple operation , short modeling time , high CP incidence and low cost .



Study on the mechanism of fibrosis in rats with chronic pancreatitis in the second part



Purpose



To investigate the relationship between the development of CP and inflammatory cell , PSC and cell growth factor in rats and the effect of PTX and TSA on CP in rats .



method



The paraffin - embedded sections were prepared by 4 % formaldehyde fixation in the experimental group and control group . 偽 - SMA and Desin were stained with EnVision two - step method , PDGF - B mRNA , TGF - 尾1 mRNA and CTGFmRNA were stained by in situ hybridization .



Results



( 4 ) The positive rate of expression of TGF - 尾1 mRNA and PDGF - BmRNA in group A and group B were significantly higher than those in control group ( P < 0.05 or P < 0.01 ) .



Conclusion



( 1 ) a - PSC is involved in the fibrotic process of CP , PDGF - BmRNA , TGF - 尾1 mRNA and CTGFmRNA can activate PSC and closely related to pancreatic fibrosis .

【學位授予單位】：中南大學
【學位級別】：博士
【學位授予年份】：2008
【分類號】：R657.5;R-332

【引證文獻】

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本文編號：1756058