本文選題：同種異體牙移植 + 樹突狀細胞�。� 參考：《重慶醫(yī)科大學》2009年碩士論文

【摘要】： 本研究旨在運用未成熟樹突狀細胞(immature dendritic cells, imDC)誘導特異性免疫耐受的特性,研究其對小鼠同種異體牙移植后排斥反應的抑制作用。 在該研究中建立了小鼠同種異體牙及自體牙異位移植模型,分別通過尾靜脈和移植局部注射imDC的方式,將供體小鼠骨髓來源的imDC回輸入受體小鼠體內,并與自體牙移植組和未經處理的同種異體牙移植組比較,HE染色和免疫組化法觀察不同回輸方式的imDC對小鼠同種異體牙移植后排斥反應的抑制效果。研究共分為兩個部分,內容如下: 第一部分:不同濃度粒巨噬細胞集落刺激因子在未成熟樹突狀細胞培養(yǎng)中的效果研究 目的:探討不同濃度細胞因子在培養(yǎng)小鼠骨髓來源未imDC中的作用,尋找培養(yǎng)所需最佳細胞因子濃度。方法:分別用低、中、高劑量粒細胞-巨噬細胞集落刺激因子(granulocyte-macrophage colony-stimulating factor,GM-CSF)聯(lián)合或不聯(lián)合白細胞介素4(Interleukin-4, IL-4)培養(yǎng)小鼠骨髓源性imDC,對其進行形態(tài)學觀察和細胞表型檢測。結果:培養(yǎng)第7 d,低劑量組DC數(shù)量最少、突起細小,CD80、CD86及MHCII(major histocompatibility complex)類分子表達低,聯(lián)合IL-4培養(yǎng)可增加DC數(shù)量,CD80、CD86及MHCII類分子的表達亦相應升高;中等劑量組與高劑量組DC數(shù)量增多,突起較低劑量組更明顯,CD80、CD86及MHCII類分子表達也較前者提高,但二組無明顯差異,在中等劑量組和高劑量組中,不論是否聯(lián)合IL-4培養(yǎng),所誘導出的DC在數(shù)量和成熟度上的差異亦不顯著。結論:GM-CSF低劑量時,可誘導出純度較高的imDC,但DC的數(shù)量較少,GM-CSF中等劑量和高劑量時,可使DC數(shù)量增多,但所誘導出的DC為成熟與未成熟DC的混合物。IL-4僅在GM-CSF低劑量時表現(xiàn)出一定程度增加DC產量和促進DC成熟的作用,當GM-CSF中等劑量和高劑量時,無論是否聯(lián)合誘導IL-4培養(yǎng),誘導imDC的產量及純度無明顯差異。 第二部分:未成熟樹突狀細胞作用于小鼠同種異體牙移植排斥反應的效果研究 目的:研究采用不同輸注方式的供體小鼠骨髓來源imDC對受體小鼠同種異體牙異位移植后排斥反應的抑制效果。方法:建立小鼠自體牙及同種異體牙異位移植模型,運用50U/mlGM-CSF誘導出小鼠骨髓來源imDC,分別于同種異體牙移植術前7d以尾靜脈注射方式或同種異體牙移植術后第0、3d以局部注射的方式2*106/只回輸入受者體內,并與自體牙移植組和未經處理的同種異體牙移植組比較,組織病理切片和免疫組化觀察imDC對小鼠同種異體牙移植后排斥反應的抑制效果。結果:自體牙移植組未見明顯排斥反應;未經處理的同種異體牙移植組出現(xiàn)相對明顯的排斥反應,表現(xiàn)為移植部位皮膚呈紫紅色,耳廓腫脹,皮溫較高,組織包裹移植牙松弛,鏡下可見大量淋巴細胞浸潤,移植牙周圍組織血管擴張充血出血,可見大量紅細胞;尾靜脈回輸imDC組排斥反應程度較輕,表現(xiàn)為移植部位充血、可見紅色結痂及少量稀薄滲出物,皮溫較對側稍高,組織包裹移植牙不甚緊密,鏡下觀中性粒細胞及部分淋巴細胞浸潤,移植牙周圍組織血管可見擴張、充血,出現(xiàn)小范圍細胞變性和單核細胞浸潤;局部注射imDC組表現(xiàn)與未經處理的同種異體牙移植組類似,出現(xiàn)較明顯的排斥反應。對免疫組化結果進行半定量分析表明,自體牙移植組CD4+T細胞和CD8+T細胞的陽性率在所有實驗組中為最低;經過尾靜脈回輸imDC處理的實驗組,CD4+T細胞和CD8+T細胞陽性率較局部注射imDC組和異體牙移植組明顯降低;但局部注射imDC組和異體牙移植組之間CD4+T細胞和CD8+T細胞陽性率無顯著差異。結論:供體骨髓來源的imDC于術前7d經尾靜脈回輸入受者體內后可減輕受者同種異體牙移植后的排斥反應,但不能完全消除排斥反應,而局部注射供者骨髓來源的imDC在本研究中發(fā)現(xiàn)對受體同種異體牙移植后的排斥反應沒有明顯抑制作用。
[Abstract]:The purpose of this study was to use the characteristics of immature dendritic cells (imDC) to induce specific immune tolerance, and to study the inhibition effect on rejection after allograft tooth transplantation in mice.
In this study, a model of heterotopic transplantation of allogenic teeth and autologous teeth in mice was established. By the way of the tail vein and the local injection of imDC, the imDC of the bone marrow from the donor mice was returned to the recipient mice. Compared with the autologous tooth transplantation group and the untreated allograft group, the HE staining and immunohistochemistry were observed. The inhibitory effect of imDC with different reinfusion methods on rejection after allograft tooth transplantation in mice is divided into two parts.
Part I: effect of granulocyte macrophage colony-stimulating factor on immature dendritic cells cultured in vitro
Objective: To explore the effect of different concentrations of cytokines on the culture of mouse bone marrow, and to find the best cytokine concentration needed for culture. Methods: granulocyte-macrophage colony-stimulating factor (GM-CSF) combined with or not combined with or without interleukin 4 (Interle) 4 (Interle). Ukin-4, IL-4) cultured mouse bone marrow derived imDC. Morphological observation and cell phenotype detection. Results: Seventh D, low dose group DC, CD80, CD86 and MHCII (major histocompatibility complex) molecules are low. The increase in the number of DC in the medium and high dose group, the lower dose group is more obvious, the expression of CD80, CD86 and MHCII is also higher than the former, but there is no significant difference between the two groups. In the medium dose group and the high dose group, no matter whether or not the IL-4 culture is combined, the differences in the quantity and maturity of the induced DC are not significant. Conclusion: GM-C At low doses of SF, a higher purity of imDC can be induced, but the number of DC is less, the number of DC can be increased when the GM-CSF is medium and high, but the induced DC is a mixture of mature and immature DC,.IL-4 only increases DC production and promotes DC maturation at a low dose of GM-CSF, when GM-CSF medium and high doses are used. No significant difference was found in the yield and purity of imDC induced by IL-4 culture.
The second part: the effect of immature dendritic cells on allograft rejection in mice.
Objective: To study the inhibitory effect of bone marrow derived imDC from donor mice with different infusion methods on rejection of allogenic tooth allograft in recipient mice. Methods: to establish autologous teeth and allograft model of allogenic teeth in mice, and to induce imDC in mouse bone marrow by 50U/mlGM-CSF, and 7d before allograft. 2*106/ only returned to the recipient after the tail vein injection or the local injection of 0,3d, and compared with the autologous tooth transplantation group and the untreated allograft group. Histopathology and immunohistochemistry were used to observe the inhibitory effect of imDC on the rejection after allograft tooth transplantation. There was no obvious rejection in the autologous tooth transplantation group, and the untreated allograft group had a relatively obvious rejection reaction, which showed that the skin of the transplanted site was purple red, the auricle swelled, the skin temperature was high, the tissue parcel was relaxed, a large number of lymphocytes were soaked in the microscope, and the blood vessels around the transplanted teeth were dilated and bleed. A large number of red blood cells; the rejection in the imDC group of the tail vein transfusion was light, showing the hyperemia of the transplant site, the red scab and a small amount of exudate, the skin temperature was slightly higher than the opposite side, the tissue wrapped in the transplanted teeth was not very close, the neutrophils and some lymphocytes infiltrated under the microscope, and the tissue vessels around the transplanted teeth were dilated, congested and small. The local injection of imDC was similar to the untreated group of allograft teeth, and there was a significant rejection. Semi quantitative analysis of the immunohistochemical results showed that the positive rate of CD4+T and CD8+T cells in the autologous tooth transplantation group was the lowest in all the experimental groups; through the caudal vein. The positive rate of CD4+T cells and CD8+T cells in the experimental group treated with imDC was significantly lower than that in the local imDC group and the allograft group, but there was no significant difference in the positive rate of CD4+T and CD8+T cells between the local injection of imDC and the allograft teeth. Conclusion: the imDC of the donor bone marrow derived from the tail vein of the donor bone marrow was entered into the recipient's body before the operation. The rejection reaction after the allograft tooth transplantation was alleviated, but the rejection reaction was not completely eliminated, and the local injection of donor bone marrow imDC in this study found no significant inhibitory effect on rejection after the allograft tooth allograft.
【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2009
【分類號】：R392

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