造血干細(xì)胞誘導(dǎo)大鼠腎臟移植免疫耐受的研究
發(fā)布時(shí)間：2019-01-20 17:28
【摘要】： 目的: 如何誘導(dǎo)供體特異性移植耐受成為解決器官移植排斥反應(yīng)的最佳手段,也成為臨床器官移植免疫學(xué)研究的熱點(diǎn)領(lǐng)域及主要目標(biāo)之一。本研究通過大鼠腎臟移植受體在接受低劑量放射線全身照射預(yù)處理后,輸注供體造血干細(xì)胞(HSC)對(duì)腎臟移植受體及移植腎的影響,探討造血干細(xì)胞能否誘導(dǎo)出針對(duì)供者特異性的免疫耐受。 方法: 1、建立支架管置入并固定于腎靜脈代替靜脈吻合大鼠腎臟移植模型:將供腎動(dòng)脈開口端與受體左腎動(dòng)脈作端端吻合,取1cm長度輸尿管導(dǎo)管作為支架管,置入并固定供受體腎靜脈,輸尿管膀胱瓣與受體膀胱壁吻合。 2、造血干細(xì)胞誘導(dǎo)大鼠腎臟移植免疫耐受:(1)實(shí)驗(yàn)分組:32只受體Wistar大鼠隨機(jī)分為4組,各組8只。A:對(duì)照組;B:造血干細(xì)胞(HSC)+腎移植;C:全身照射(TBI)+腎移植;D:HSC+TBI+腎移植。(2)受體Wistar大鼠術(shù)前接受低劑量放射線全身照射,提取供者造血干細(xì)胞,手術(shù)當(dāng)日經(jīng)尾靜脈推注SD大鼠的造血干細(xì)胞,并當(dāng)天完成腎移植,受體實(shí)驗(yàn)前1天,實(shí)驗(yàn)當(dāng)天,及實(shí)驗(yàn)后1、3、4、5天按10mg/kg口服環(huán)孢素A。(3)觀察各組大鼠的存活時(shí)間,監(jiān)測(cè)血清肌酐濃度,并進(jìn)行移植腎彩色多普勒超聲。移植物病理學(xué)檢查。采用體外混合淋巴細(xì)胞反應(yīng)(MLR)法檢測(cè)長期存活大鼠的免疫耐受狀態(tài)。 結(jié)果: 1、建立模型:共施行大鼠。腎移植30例,5例動(dòng)脈吻合處有少量出血,棉球壓迫5min后2例出血停止,3例因吻合處出血于術(shù)后1-3h死亡(尸體解剖證實(shí)),2例因術(shù)中動(dòng)脈吻合口狹窄死亡,2例因麻醉過量呼吸抑制于術(shù)中死亡,成功率為76.7%。 2、存活時(shí)間:D組平均存活時(shí)間為58.9d,與A組比較,差異有統(tǒng)計(jì)學(xué)意義(P＜0.01)。D組與B組及C組比較,差異有統(tǒng)計(jì)學(xué)意義(P＜0.05)。 3、移植腎功能測(cè)定:A組術(shù)后3d切除自體腎臟后,血清肌酐濃度迅速上升;B組及C組血清肌酐濃度呈現(xiàn)緩慢上升趨勢(shì),D組的移植腎功能基本保持穩(wěn)定。 4、病理學(xué)結(jié)果:對(duì)照組移植腎腎間質(zhì)水腫,并有淋巴細(xì)胞浸潤,多見于間質(zhì)小血管周圍,腎小管上皮細(xì)胞變性壞死、脫落,血管內(nèi)皮細(xì)胞腫脹,有纖維素樣壞死并可見微血栓形成。B組和C組未見急性排斥反應(yīng),腎間質(zhì)水腫不明顯,見少量淋巴細(xì)胞浸潤。腎小管上皮細(xì)胞變性壞死、脫落,血管內(nèi)皮細(xì)胞腫脹不明顯。同期D組未見淋巴細(xì)胞浸潤,腎小管上皮細(xì)胞未見變性壞死、脫落。無微血栓形成。 5、移植腎超聲檢測(cè):A組移植腎血供差,阻力指數(shù)0.7。D組移植腎血供豐富,阻力指數(shù)介于0.3～0.6之間,A組與B組差別不明顯(P=0.06),A組與C組差別明顯(P＜0.05),A組與D組差別明顯(P＜0.05)。 6、混合淋巴細(xì)胞反應(yīng)(MLR):A、B組大鼠脾細(xì)胞對(duì)供體SD大鼠或無關(guān)品系Lewis大鼠脾細(xì)胞的CPM值與空白對(duì)照組相比無顯著性差異(P＞0.05)。而對(duì)SD大鼠脾細(xì)胞,D組的CPM值較空白對(duì)照組明顯降低(P＜0.01),對(duì)Lewis大鼠脾細(xì)胞,上述三組的CPM與對(duì)照組比較未見顯著性差異(P＞0.05)。 結(jié)論: 1、建立大鼠腎臟移植模型:供腎動(dòng)脈端端吻合,輸尿管導(dǎo)管作為支架,將其置入供受體腎靜脈斷端,行供受體腎靜脈端端吻合,行輸尿管膀胱瓣和受體膀胱壁吻合。此方法簡便、易操作,手術(shù)成功率高。一般實(shí)驗(yàn)室均可開展,對(duì)腎移植實(shí)驗(yàn)研究有應(yīng)用價(jià)值。 2、通過大鼠腎臟移植受體在接受全身照射預(yù)處理后,輸注供體HSC可延長受者存活時(shí)間,移植腎功能基本保持穩(wěn)定,短期內(nèi)未見排斥反應(yīng),HSC可誘導(dǎo)針對(duì)供者特異性的免疫耐受。
[Abstract]:Purpose: How to induce donor-specific transplantation tolerance is the best way to solve the rejection of organ transplantation, and also become the hot spot and main purpose of the research of clinical organ transplantation immunology The effect of the donor hematopoietic stem cell (HSC) on the renal transplantation receptor and the transplanted kidney was studied by the donor hematopoietic stem cell (HSC) after the low-dose radiation. epidemic tolerance The method comprises the following steps of: 1, establishing a stent tube and fixing the renal vein in place of the renal vein in place of the kidney transplantation model of the vein anastomosis rat: the open end of the renal artery is matched with the end of the left renal artery of the receptor, and the 1cm length of the ureter catheter is taken as the stent tube; is placed and fixed for recipient renal vein, ureter, Bladder flap and receptor bladder wall anastomosis. 2. Hematopoietic stem cell induced kidney transplantation tolerance in rats: (1) experimental group: 32 receptor Wist ar rats were randomly divided into 4 groups, each group of 8. A: control group; B: hematopoietic stem cell (HSC) + kidney transplantation; C: whole body irradiation (TBI) + kidney removal Plant; D: HSC + TBI + kidney transplantation. (2) The recipient Wistar rats were irradiated with low-dose radiation before the operation, and the donor hematopoietic stem cells were extracted. The hematopoietic stem cells of SD rats were injected by tail vein on the same day, and the kidney was completed on the same day. The survival time of each group was observed at 10 mg/ kg in 1, 3, 4 and 5 days prior to the experiment and 1, 3, 4 and 5 days after the experiment. degree, and graft the kidney color Color Doppler ultrasound. Graft pathology examination. In vitro mixed lymphocyte reaction (MLR) method Length of test The immune tolerance status of the surviving rats. Results: 1. The model was established: a total of 30 cases of kidney transplantation, 5 cases of arterial anastomosis had a small amount of bleeding, 2 cases of hemorrhage were stopped after the cotton ball was compressed for 5min, 3 cases died from 1 to 3 hours after the operation of the anastomosis, 2 cases were due to intra-operative arterial anastomosis. stenosis of the mouth and death of 2 cases The total survival time of D group was 5. 7%. 2. The survival time: the average survival time of D group was 5. 8. 9d, compared with group A, the difference was statistically significant (P <0.01). In group D, compared with group B and group C, the difference was statistically significant (P <0.05). 3. The determination of renal function: after 3 days after the operation of group A, the concentration of serum myoglobin increased rapidly; group B and group B were in group B and group B. In group C, the concentration of serum myoglobin decreased slowly, and the transplanted renal function in group D was stable. 4. Pathological results: the control group transplanted renal interstitial edema and the infiltration of lymphocytes, which was found in the surrounding of the interstitial small vessels and the renal tubular. Skin cell degeneration and necrosis, shedding, swelling of vascular endothelial cells, fibrinoid necrosis and visible microthrombi The group B and group C did not meet the acute rejection, and the edema of the renal interstitial edema was not obvious. See Small number of lymphocyte infiltration. Renal tubular epithelial cell degeneration and necrosis, shedding, vascular endothelial cells The swelling is not obvious. It's the same. There was no infiltration of lymphocytes in the D group. The renal tubular epithelial cells were not found to have degeneration and necrosis. There was no microthrombosis. 5. Transplanted renal ultrasound: A group of renal blood was transplanted in group A, and the resistance index was 0. 3-0.6. The difference between group A and group B was not significant (P = 0). The difference between group A and group C was significant (P <0.05), and the difference between group A and group D was significant (P <0.05). 6. Mixed lymphocyte reaction (MLR): A and B rats spleen cells were large for donor SD. Compared with the blank control group, the CPM value of the spleen cells of the mouse or the unrelated strain Lewis rats was no significant difference (P> 0.05), and the CPM value of the spleen cells and the D group in the SD rats was significantly lower than that of the blank control group (P <0.01).), for Conclusion: 1. The rat kidney transplantation model is established: the end of the renal artery is in line with the end of the renal artery. and the ureteral catheter is used as a support, and the ureter catheter is placed for receiving and receiving the stent, The end of the renal vein of the body is connected with the end of the renal vein of the recipient, and the ureter is connected with the end of the vein. the method is simple, easy to operate and high in operation success rate, and the method is simple, easy to operate and high in operation success rate.
【學(xué)位授予單位】：中國人民解放軍軍醫(yī)進(jìn)修學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2009
【分類號(hào)】：R392;R699.2
本文編號(hào)：2412232
[Abstract]:Purpose: How to induce donor-specific transplantation tolerance is the best way to solve the rejection of organ transplantation, and also become the hot spot and main purpose of the research of clinical organ transplantation immunology The effect of the donor hematopoietic stem cell (HSC) on the renal transplantation receptor and the transplanted kidney was studied by the donor hematopoietic stem cell (HSC) after the low-dose radiation. epidemic tolerance The method comprises the following steps of: 1, establishing a stent tube and fixing the renal vein in place of the renal vein in place of the kidney transplantation model of the vein anastomosis rat: the open end of the renal artery is matched with the end of the left renal artery of the receptor, and the 1cm length of the ureter catheter is taken as the stent tube; is placed and fixed for recipient renal vein, ureter, Bladder flap and receptor bladder wall anastomosis. 2. Hematopoietic stem cell induced kidney transplantation tolerance in rats: (1) experimental group: 32 receptor Wist ar rats were randomly divided into 4 groups, each group of 8. A: control group; B: hematopoietic stem cell (HSC) + kidney transplantation; C: whole body irradiation (TBI) + kidney removal Plant; D: HSC + TBI + kidney transplantation. (2) The recipient Wistar rats were irradiated with low-dose radiation before the operation, and the donor hematopoietic stem cells were extracted. The hematopoietic stem cells of SD rats were injected by tail vein on the same day, and the kidney was completed on the same day. The survival time of each group was observed at 10 mg/ kg in 1, 3, 4 and 5 days prior to the experiment and 1, 3, 4 and 5 days after the experiment. degree, and graft the kidney color Color Doppler ultrasound. Graft pathology examination. In vitro mixed lymphocyte reaction (MLR) method Length of test The immune tolerance status of the surviving rats. Results: 1. The model was established: a total of 30 cases of kidney transplantation, 5 cases of arterial anastomosis had a small amount of bleeding, 2 cases of hemorrhage were stopped after the cotton ball was compressed for 5min, 3 cases died from 1 to 3 hours after the operation of the anastomosis, 2 cases were due to intra-operative arterial anastomosis. stenosis of the mouth and death of 2 cases The total survival time of D group was 5. 7%. 2. The survival time: the average survival time of D group was 5. 8. 9d, compared with group A, the difference was statistically significant (P <0.01). In group D, compared with group B and group C, the difference was statistically significant (P <0.05). 3. The determination of renal function: after 3 days after the operation of group A, the concentration of serum myoglobin increased rapidly; group B and group B were in group B and group B. In group C, the concentration of serum myoglobin decreased slowly, and the transplanted renal function in group D was stable. 4. Pathological results: the control group transplanted renal interstitial edema and the infiltration of lymphocytes, which was found in the surrounding of the interstitial small vessels and the renal tubular. Skin cell degeneration and necrosis, shedding, swelling of vascular endothelial cells, fibrinoid necrosis and visible microthrombi The group B and group C did not meet the acute rejection, and the edema of the renal interstitial edema was not obvious. See Small number of lymphocyte infiltration. Renal tubular epithelial cell degeneration and necrosis, shedding, vascular endothelial cells The swelling is not obvious. It's the same. There was no infiltration of lymphocytes in the D group. The renal tubular epithelial cells were not found to have degeneration and necrosis. There was no microthrombosis. 5. Transplanted renal ultrasound: A group of renal blood was transplanted in group A, and the resistance index was 0. 3-0.6. The difference between group A and group B was not significant (P = 0). The difference between group A and group C was significant (P <0.05), and the difference between group A and group D was significant (P <0.05). 6. Mixed lymphocyte reaction (MLR): A and B rats spleen cells were large for donor SD. Compared with the blank control group, the CPM value of the spleen cells of the mouse or the unrelated strain Lewis rats was no significant difference (P> 0.05), and the CPM value of the spleen cells and the D group in the SD rats was significantly lower than that of the blank control group (P <0.01).), for Conclusion: 1. The rat kidney transplantation model is established: the end of the renal artery is in line with the end of the renal artery. and the ureteral catheter is used as a support, and the ureter catheter is placed for receiving and receiving the stent, The end of the renal vein of the body is connected with the end of the renal vein of the recipient, and the ureter is connected with the end of the vein. the method is simple, easy to operate and high in operation success rate, and the method is simple, easy to operate and high in operation success rate.
【學(xué)位授予單位】：中國人民解放軍軍醫(yī)進(jìn)修學(xué)院
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2009
【分類號(hào)】：R392;R699.2
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