造血干细胞诱导大鼠肾脏移植免疫耐受的研究
发布时间：2019-01-20 17:28
【摘要】： 目的: 如何诱导供体特异性移植耐受成为解决器官移植排斥反应的最佳手段,也成为临床器官移植免疫学研究的热点领域及主要目标之一。本研究通过大鼠肾脏移植受体在接受低剂量放射线全身照射预处理后,输注供体造血干细胞(HSC)对肾脏移植受体及移植肾的影响,探讨造血干细胞能否诱导出针对供者特异性的免疫耐受。 方法: 1、建立支架管置入并固定于肾静脉代替静脉吻合大鼠肾脏移植模型:将供肾动脉开口端与受体左肾动脉作端端吻合,取1cm长度输尿管导管作为支架管,置入并固定供受体肾静脉,输尿管膀胱瓣与受体膀胱壁吻合。 2、造血干细胞诱导大鼠肾脏移植免疫耐受:(1)实验分组:32只受体Wistar大鼠随机分为4组,各组8只。A:对照组;B:造血干细胞(HSC)+肾移植;C:全身照射(TBI)+肾移植;D:HSC+TBI+肾移植。(2)受体Wistar大鼠术前接受低剂量放射线全身照射,提取供者造血干细胞,手术当日经尾静脉推注SD大鼠的造血干细胞,并当天完成肾移植,受体实验前1天,实验当天,及实验后1、3、4、5天按10mg/kg口服环孢素A。(3)观察各组大鼠的存活时间,监测血清肌酐浓度,并进行移植肾彩色多普勒超声。移植物病理学检查。采用体外混合淋巴细胞反应(MLR)法检测长期存活大鼠的免疫耐受状态。 结果: 1、建立模型:共施行大鼠。肾移植30例,5例动脉吻合处有少量出血,棉球压迫5min后2例出血停止,3例因吻合处出血于术后1-3h死亡(尸体解剖证实),2例因术中动脉吻合口狭窄死亡,2例因麻醉过量呼吸抑制于术中死亡,成功率为76.7%。 2、存活时间:D组平均存活时间为58.9d,与A组比较,差异有统计学意义(P＜0.01)。D组与B组及C组比较,差异有统计学意义(P＜0.05)。 3、移植肾功能测定:A组术后3d切除自体肾脏后,血清肌酐浓度迅速上升;B组及C组血清肌酐浓度呈现缓慢上升趋势,D组的移植肾功能基本保持稳定。 4、病理学结果:对照组移植肾肾间质水肿,并有淋巴细胞浸润,多见于间质小血管周围,肾小管上皮细胞变性坏死、脱落,血管内皮细胞肿胀,有纤维素样坏死并可见微血栓形成。B组和C组未见急性排斥反应,肾间质水肿不明显,见少量淋巴细胞浸润。肾小管上皮细胞变性坏死、脱落,血管内皮细胞肿胀不明显。同期D组未见淋巴细胞浸润,肾小管上皮细胞未见变性坏死、脱落。无微血栓形成。 5、移植肾超声检测:A组移植肾血供差,阻力指数0.7。D组移植肾血供丰富,阻力指数介于0.3～0.6之间,A组与B组差别不明显(P=0.06),A组与C组差别明显(P＜0.05),A组与D组差别明显(P＜0.05)。 6、混合淋巴细胞反应(MLR):A、B组大鼠脾细胞对供体SD大鼠或无关品系Lewis大鼠脾细胞的CPM值与空白对照组相比无显著性差异(P＞0.05)。而对SD大鼠脾细胞,D组的CPM值较空白对照组明显降低(P＜0.01),对Lewis大鼠脾细胞,上述三组的CPM与对照组比较未见显著性差异(P＞0.05)。 结论: 1、建立大鼠肾脏移植模型:供肾动脉端端吻合,输尿管导管作为支架,将其置入供受体肾静脉断端,行供受体肾静脉端端吻合,行输尿管膀胱瓣和受体膀胱壁吻合。此方法简便、易操作,手术成功率高。一般实验室均可开展,对肾移植实验研究有应用价值。 2、通过大鼠肾脏移植受体在接受全身照射预处理后,输注供体HSC可延长受者存活时间,移植肾功能基本保持稳定,短期内未见排斥反应,HSC可诱导针对供者特异性的免疫耐受。
[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.
【学位授予单位】：中国人民解放军军医进修学院
【学位级别】：博士
【学位授予年份】：2009
【分类号】：R392;R699.2
本文编号：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.
【学位授予单位】：中国人民解放军军医进修学院
【学位级别】：博士
【学位授予年份】：2009
【分类号】：R392;R699.2
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