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Donor dendritic cells-mediated recipient lymphocyte reaction after living related kidney transplantation

Chibing HUANG ; Xiaoting XU ; Jiayu FENG ; Mingqi FAN ; Genfu ZHANG ; Pingxian WANG ; Ya XIAO

Chinese Journal of Organ Transplantation.2011;32(1):39-42. doi:10.3760/cma.j.issn.0254-1785.2011.01.011

Objective To explore the feasibility of mediating recipient lymphocyte reaction with donor dendritic cells (DCs) in renal allograft recipients. Methods Donor bone marrow monocytes (BMMCs) were isolated and cryopreserved in liquid nitrogen before kidney transplantation. At 0 day, 1month,3 month, 6 month and 9 month post-operation, CD34+ cells which were isolated from frozen BMMCs and cultured into DCs as well as the peripheral blood lymphocytes (PBLs) of donors were used as the stimulating cells to the PBLs of recipients and healthy volunteers. The number of viable DCs from frozen- and room temperature-preserved BMMCs was counted and the reactions of recipients'and healthy volunteers' lymphocytes to DCs and donor PBLs were measured. Results 6. 8 × 107BMMCs were isolated from each 10 ml of donor bone marrow on average while (4. 10 ± 0. 58) × 105CD34+ cells were isolated by magnetic active cell sorting (MACS). There was no significant difference in the isolating rate of recovered CD34+ cells at each observation point postoperatively. The percentage of viable BMMCs and CD34+ was decreased significantly at 1 month after surgery, then, decreased slowly and progressively. The decreasing rate of BMMCs was higher than CD34+. The rate of viable DCs was maintained stable (93. 2%-94. 8% ) in each group. The reactions of recipients' and healthy volunteers' lymphocytes to DCs were stronger than those to donor PBLs (P＜0. 05). The reactions of healthy volunteers' lymphocytes to DCs were maintained stable while those of recipients' were fluctuating. Conclusion Bone marrow-derived DCs are superior to PBLs in mediating long-term lymphocyte reaction after kidney transplantation due to their stable viability and stimulating ability to lymphocytes. Only once collection of a small quality of bone marrow of donors is needed to meet the demand of immune monitoring at any time after transplantation.

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A novel bone marrow transplantation strategy for donor-specific tolerance induction after heart transplantation

Kequan GUO ; Xu MENG ; Yuanlong YU ; Jie HAN ; Haiming JIANG ; Xiaojun XU ; Xiaojun LU ; Yixin JIA ; Junmeng ZHENG ; Haibo ZHANG ; Yan LI ; Tie ZHENG ; Chunlei XU ; Wen ZENG ; Jiangang WANG ; Yongqiang CUI ; Tiange LUO ; Jun WANG ; Susumu IKEHARA

Chinese Journal of Organ Transplantation.2011;32(1):32-35. doi:10.3760/cma.j.issn.0254-1785.2011.01.009

Objective To investigate a new strategy of bone marrow transplantation (BMT) for donor-specific tolerance induction after heart transplantation. Methods Donor bone marrow cells (BMCs)were harvested simultaneously with donor cardiac graft using modified perfusion method (PM) ,then stored in a -80 ℃ refrigerator after filtration and centrifugation. Whole BMCs (IBM-BMT) (monocytes 1.2 ×107/kg,CD34+ cells 2.38× 105/kg) in host iliac bones were injected into the bone marrow cavity 40 days after heart transplantation. Preconditoning regimens that consisted of fludarabine, antithymoctye globin and total lymphoid irradiation were performed 3 days before BMT. Tacrolimus (Tac) was administrated intravenously after BMT or orally in conjunction with mycophenolate mofetil (MMF) 3 weeks later.Cyclosporine and MMF were orally administrated 6 weeks later. Donor chimerism was detected using short tandem repeats-polymerase chain reaction in monocytes from peripheral blood at the 2nd,4th, 8th or 12th week after BMT or BMCs at the 4th, 8th or 12th week after BMT. Intramyocardium electrocardiography examination or endomyocardial biopsy was performed weekly or monthly respectively. Mixed lymphocyte reactions (MLR) were performed 3 months after BMT. Results Donor chimerism in monocytes in peripheral blood or BMCs in iliac bones measured at the 1 st,2nd and 3rd month after BMT was 26.3%, 19.1%,4.8% ,and 46.3%, 24.4%, 7.6%, respectively. After 3-month follow-up, there was no rejection confirmed by endomyocardial biopsy or intramyocardium electrocardiography. Echocardiography revealed that the diastolic and systolic function of the cardiac graft was maintained well 3 months after BMT. MLR revealed donor-specific hyporesponsiveness while immunocompetence was preserved to third-party antigens. Conclusion These findings indicate that the two-stage BMT strategy is a safe and feasible method for the induction of donor-specific tolerance via stable mixed chimerism and needs to be further confirmed after a long-term observation.

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Perioperative application of extracorporeal membrane oxygenation (ECMO) in lung transplantation

Mingfeng ZHENG ; Jingyu CHEN ; Xingfeng ZHU ; Yijun HE ; Ji ZHANG ; Shugao YE ; Feng LIU ; Ruo CHEN ; Dong WEI

Chinese Journal of Organ Transplantation.2011;32(1):28-31. doi:10.3760/cma.j.issn.0254-1785.2011.01.008

Objective To explore the perioperative application of extracorporeal membrane oxygenation (ECMO) in lung transplantation. Methods Thirty patients with primary and end-stage pulmonary disease accompanied by pulmonary hypertension were subjected to operation under the accessory of ECMO. Eighteen patients received single-lung transplantation and 12 patients bilateral sequential lung transplantation without sternal division in our hospital from November 2005 to July 2009. In 2 patients ECMO was given before operation and maintained for 19 days and 6 days respectively. In the remaining patients, ECMO pipeline was placed after anesthesia. After lung trarnsplantation,ECMO was removed after the recipients' oxygen saturation and hemodynamics were stable. Results In all recipients lung transplantation was successfully done. ECOM was removed in 27recipients after operation, and the rest 3 recipients were supported by ECMO after operation: the ECMO was removed at 36th h and 7th day after lung transplantation in two patients respectively,and another one was supported by ECMO for 5 days after operation and suffered acute kidney failure, and died of multiple organ failure 2 weeks post-transplantation. Two recipients were infected in thigh arteriovenous cut and one suffered femoral artery thrombosis, but all of them got better and discharged from hospital after treatment. Conclusion ECMO can be used for lung transplantation on patients with primary and secondary pulmonary hypertension. The complications may be associated with patients'serious condition and unstable hemodynamics. Early detection and active and effective treatment can improve patient's prognosis.

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Influence of the lentiviral vectors-mediated mouse genetic engineering Treg on GVHD and GVL effect after allogeneic bone marrow transplantation in mice

Jiang CAO ; Li LI ; Chong CHEN ; Lingyu ZENG ; Zhenyu LI ; Hai CHENG ; Kailin XU

Chinese Journal of Organ Transplantation.2010;31(12):715-719. doi:10.3760/cma.j.issn.0254-1785.2010.12.002

Objective To explore the influence of the lentiviral vectors-mediated mouse genetic engineering regulatory T cells (Treg) infused after allogeneic bone marrow transplantation (alloBMT) on graft-versus-host disease (GVHD) and graft-versus-leukemia (GVL) effect in mice.Methods Lentivirus-mediated expression of Forkhead box P3 (Foxp3) transformed CD4 + CD25- T cells from Balb/c mice into engineered Tregs in vitro. An allo-BMT model of Balb/c→C57BL/6 mice was established. The recipients were given lethal X-ray total body irradiation before transplantation.Mice were randomly assigned into five groups and each group contained 10 recipients: (1) The recipients in radiation group were injected with 0.2 ml RPMI 1640; (2) The recipients in leukemia control group were injected with 5 × 106 donor bone marrow cells and 500 mouse T-cell leukemia/lymphoma cells (EL4 cells); (3) The recipients in transplantation control group were injected with 5 × 106 donor bone marrow cells and 5 × 106 splenocytes plus 500 EL4 cells; (4) The recipients in engineering Treg group were injected with 5 × 106 donor bone marrow cells, 5 × 106 splenocytes and 500 EL4 cells plus 5 × 106 genetic engineering Treg; (5) The recipients in empty vector control group were injected with 5 × 106 donor bone marrow cells, 5 × 106 splenocytes and 500 EL4 cells plus 5 × 106 empty vector-transduced CD4+ CD25- T cells. Survival time, clinical GVHD score or histopathological analysis (skin, liver and small intestine) were observed after allo-BMT. Chimerism of bone marrow cells from recipients survived for 60 days after transplantation was measured. Results The mean survival time in radiation group, leukemia control group, transplantation control group,engineering Treg group and empty vector control group was ( 10. 3 ± 1.5), (20. 7 ± 1.9), (26. 0 ±4.3), (49. 0 ± 17. 7) and (24. 4 ± 4. 1 ) days respectively. The survival time in engineering Treg group was significantly prolonged as compared with other groups as judged by the log-rank test (P＜0. 05).Histopathological analysis in several target organs (skin, liver and small intestine) confirmed the presence of severe GVHD in transplantation control group and empty vector control group. No histological signs of GVHD or leukemia were observed in recipients in engineering Treg group and clinical GVHD scores in this group were significantly decreased as compared with transplantation control group and empty vector control group. Conclusion Co-injection of genetic engineering Treg can efficiently prevent recipients from lethal GVHD without affecting GVL activity during allo-BMT in mice.

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Application of flow cytometry to detect natural specific antibody levels in human ABO blood group

Chi ZHANG ; Xiaowei GUO ; Ying XIANG ; Qing WEI ; Shi CHEN ; Gang CHEN

Chinese Journal of Organ Transplantation.2011;32(1):24-27. doi:10.3760/cma.j.issn.0254-1785.2011.01.007

Objective To establish and validate a standard method using flow cytometry (FCM)in human ABO blood group antibody (Ab) detection. Methods Sera samples from 52 blood donors were incubated with standardize red blood cells (RBCs), and anti-A/B antibody (IgM/IgG) was measured by indirect flow cytometry after adding secondary isotype-specific fluoresce labeled Ab. The results were compared with those by using ELISA reader. Results The changes of anti-A/B IgM measured by both methods correlated well and an overall correlation coefficiency of 0.730 for IgM was obtained by Spearman's rank testing (P＜0.05). FCM showed better specificity, sensitivity and reproducibility over ELISA reader. In blood group A samples (n = 16) or B samples (n = 16) ,the antirespectively. IgG blood group Ab could only be detected in blood group O sera. Conclusion Compared with traditional methods,FCM is a more objective method to offer accurate detection of natural ABO blood group Ab (IgM/IgG) and allowed semi-quantitative measurement of antibody levels.

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Infusion of donor-specific regulatory T cells prolonged islet allograft survival in mice

Ming LI ; Dong CHEN ; Yonghai LI ; Sheng CHANG ; Ying XIANG ; Hui GUO ; Weijie ZHANG

Chinese Journal of Organ Transplantation.2011;32(1):20-23. doi:10.3760/cma.j.issn.0254-1785.2011.01.006

Objective To investigate the effects of donor-specific regulatory T cells (Treg) transfusion on islet allograft survival. Methods Allogeneic fresh islets from Balb/c mice were transplanted to streptozotocin-induced diabetic C57 mice. The survival of islet allografts was observed. The experiment was divided into 3 groups: control group, nothing had been done to the recipients; simple islet transplantation group, the recipients received the islet transplantation only; experimental group, the recipients were given 1 ×106 Treg, then received islet transplantation. Results Blood glucose (BG) was above 16. 7 mmol/L after islet transplantation in control group; In simple islet transplantation group,BG level returned to normal level 1 to 2 days after transplantation, and hyperglycemia appeared 7 to 11 days after transplantation and maintained as the same as that before transplantation; In experimental group, BG level returned to normal level 2 days after transplantation and maintained at a low level,and at the 21st day after transplantation BG level was over 16. 7mmol/L in some recipients. Islet allograft survival in experimental group was significantly prolonged as compared with simple islet transplantation group. Conclusion Donor-specific Treg transfusion could prolong the islet allograft survival,and maybe have positive effect on tolerance induction of islet transplantation.

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Treatment of jejunum perforation after living-related liver transplantation for infants with biliary atresia a report of 4 cases

Mingman ZHANG ; Chonglun PU ; Chunbao GUO ; Yingcun LI ; Xiaoke DAI ; Qiang XIONG ; Quan KANG ; Zhimei REN ; Yuhua DENG ; Xiang LAN

Chinese Journal of Organ Transplantation.2011;32(1):47-49. doi:10.3760/cma.j.issn.0254-1785.2011.01.013

Objective To investigate the cause of jejunum perforation after infantile livingrelated liver transplantation (ILRLT) and summarize the experience of treatment. Methods The clinical data of 28 infants with biliary atresia who underwent ILRLT were analyzed and 4 of 28 infantile recipients (14. 3%) developed jejunum perforation after ILDLT. Results Four patients had 7 episodes of jejunum perforation after transplantation among 28 infantile recipients who underwent ILRLT because of biliary atresia. The median time between transplantation and perforation was 11 days.Perforation occurred at the point of silk in jejunum stoma (n = 3) and the Roux-en-Y limb (n = 1 ).None had a history of prior operation including Kasai in 4 patients. Clinical manifestation included fever, increased heart rate, abdominal distention, leukocytosis, and no free air on abdominal roentgenograrns. A simple repair was performed in three infants with silk: two developed recurrent perforation (67%) and underwent a re-exploration,and another had a third perforation and underwent a third repair because of re-perforation. Another child underwent a simple repair with prolene, and there was no recurrence. None died from the perforation in our study. Conclusion The occurrence and location of jejunum perforation after ILDLT suggests that the cause of the perforation is related to the jejunal anastomosis with silk, and the jejunum perforation may be avoided in the jejunal anastomosis with prolene. Early diagnosis and exploration may ensure better survival.

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Applications of magnetic resonance cholangiopancreatography after fat meal in the preoperative evaluation of the biliary system of the donors for living liver transplantation

Peng LI ; Wen SHEN ; Hongyan NI ; Jianzhong YIN ; Miaomiao LONG ; Shuang XIA ; Qian JI ; Minghui CUI ; Tie LIU ; Ji QI

Chinese Journal of Organ Transplantation.2011;32(1):43-46. doi:10.3760/cma.j.issn.0254-1785.2011.01.012

Objective To evaluate the applications of magnetic resonance cholangiopancreatography (MRCP) after fat meal in the preoperative evaluation of biliary anatomy of living liver donors.Methods Fifty cases of the preoperative donors for living liver transplantation were included and all had the corresponding intraoperative cholangiography (IOC) information. The MRCP of the donors for living liver transplantation was performed before and after fat meal (two fried eggs). The visualization and diameter of the secondary bile duct were analyzed before and after the fat meal. The results of the biliary branching pattern by MRCP after fat meal were compared with the corresponding IOC results. The accuracy, sensitivity,specificity, positive predictive value and negative predictive value of MRCP after the fat meal in distinguishing normal and any type of variant biliary anatomy were calculated. Results In all cases,82% of the 50 cases in MRCP before the fat meal could meet the diagnosis needs of the preoperative evaluation,and 100% of the 50 cases in MRCP after the fat meal could meet the diagnosis needs. There was significant difference in the demonstration quality and diameter of the secondary bile duct in MRCP before and after the fat meal (P＜0. 05). MRCP showed accurate anatomy of the biliary system, using IOC as the reference standard, in 49(98%) subjects. The sensitivity, specificity, positive predictive value and negative predictive value of MRC in distinguishing normal and any type of variant biliary anatomy were 98%,94. 7%, 100%, 10% and 96. 9%,respectively. Conclusion The MRCP after fat meal can clearly demonstrate the secondary bile duct and perfectly meet the needs of the preoperative evaluation of the living liver transplantation. The MRCP after fat meal and routine MRCP should be considered complementary to one another in order to avoid complications in living liver transplantation donors.

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Clinical research of Hangzhou domestic tacrolimus in kidney transplantation

Bingyi SHI ; Lixin YU ; Wujun XUE ; Jianming TAN ; Zhishui CHEN ; Liming WANG ; Xiaodong ZHANG ; Chibing HUANG ; Zhenli GAO

Chinese Journal of Organ Transplantation.2011;32(1):36-38. doi:10.3760/cma.j.issn.0254-1785.2011.01.010

Objective To demonstrate the efficacy and safety of Hangzhou tacrolimus capsule(Saishi Tac capsule, Hangzhou Zhongmei Huadong Pharmaceutical Co. Ltd, China) in Chinese kidney transplant recipients. Methods Multicenter, randomized open-labeled, prospective controlled clinical trial was performed in de novo Chinese kidney transplant recipients. According to including and excluding criterions, 65 kidney recipients from 9 transplant centers were enrolled. The mean age of recipients was (36.53 ± 5.71 ) years, and 8 received living donor transplantion. The time of cold ischemia and warm ischemia was (4.08 ± 5.43) h and (3.90 ± 2.15) min respectively. The number of mismatched HLA was (2.1 ± 0.8). The recipients accepted Saishi Tac capsule + mycophenolate steroid 60 days, followed by 5-10 μg/L until the terminal observation time point (12 weeks after transplantation). The efficacy and safety were estimated during the period. The primary efficacy endpoint of the study was the incidence of biopsy-confirmed acute rejection. Graft survival and renal function (evaluated by serum creatinine) were the secondary endpoints. Safety was assessed by monitoring laboratory parameters and adverse events reported over the course of the study, such as infection, hepatic damage, hypertension, hyperlipema, diabetes mellitus and other adverse affairs.Results The dose of Tac at 1 st, 2nd, 4th and 8th week postoperation was (6.54 ± 1.69), (6.39 ±1.45),(6.73± 1.25), (6.25 ± 1.02) and (6.03 ± 1.16) mg, corresponding values to the C0 were (8.24±2.09),(9.39± 1.35),(9.93± 1.87),(7.23± 1.16) and (6.43± 1.26) μg/L. During 12weeks of follow-up, the incidence of biopsy-confirmed acute rejection was 12.3% (8/65), among which 6 cases were reversed by implosive therapy. The survival rate of graft kidney was 96.9% (63/65). The incidence of hypertension and hepatic damage was both 7.7% and morbidity of lung infection was 7.6%. There were 3 patients (4.6%) complicated with hyperlipema and diabetes mellitus respectively. Conclusion During the first 3 months of treatment Saishi Tac capsule was safe and effective to Chinese kidney transplant recipients.

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Effects of mesenchymal stem cells derived from bone marrow of patients with chronic myelogenous leukemia on function of monocyte-derived dendritic cells

Zhigang ZHAO ; Li SUN ; Xiaofang WANG ; Yizhuo ZHANG ; Yong YU ; Ping ZOU

Chinese Journal of Organ Transplantation.2011;32(1):16-19. doi:10.3760/cma.j.issn.0254-1785.2011.01.005

Objective To study the effects and mechanisms of mesenthymal stem cells (MSCs)derived bone marrow of patients with chronic myelogenous leukemia (CML) on function of monocytederived dendritic cells in vitro. Methods Bone marrow mononuclear cells from CML patients were obtained and cultured. Peripheral blood mononuclear cells (PBMCs) derived from normal volunteers were isolated and cultured in DC differentiational condition. Moreover, PBMCs were co-cultured with CML bone marrow-derived MSCs (CML-MSC) or normal volunteers' bone marrow-derived MSC (normal-MSC) in DC differentiational condition. Immunophenotype and the endocytosis of monocytederived DCs were investigated by FACS. The level of IL-12 was evaluated by enzyme linked immunosorbant assay (ELISA). The immunoregulatory ability was detected by mixed lymphocyte culture assay. Results CML-MSCs or normal-MSC inhibited the up-regulation of CD1a,CD40,CD80,CD86,and HLA-DR during DC differentiation and reduced CD40,CD86,and CD83 expression during DC maturation. CML-MSCs inhibited the endocytosis of DCs and decreased their capacity to secret IL-12. CML-MSC could significantly suppress the function of DCs stimulating proliferation of T lymphocytes. Conclusion CML-derived MSCs harbored effect on the differentiation and maturation of DCs in vitro ; CML-MSC could inhibit the immunregulation of DCs.

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