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Protective mechanism of sirolimus pretreatment against liver ischemia-reperfusion injury in rats

Jianhua RAO ; Ling LU ; Feng CHENG ; Liyong PU ; Xiaofeng QIAN ; Xuehao WANG ; Feng ZHANG

Chinese Journal of Organ Transplantation.2009;30(12):749-752. doi:10.3760/cma.j.issn.0254-1785.2009.12.011

Objective To investigate the protective effect of sirolimus pretreatment against liver ischemia-reperfusion(I/R)injury in rat model and the possible mechanism.Methods Forty-eight male SD rats were randomized into four groups (12/group):A:sham group with saline,B:sham group with sirolimus,C:saline-operated group,D:sirolimus-operated group.The rats were pretreated with either saline or sirolimus (2 mg·kg~(-1)·d~(-1))by oral gavage for two weeks．The rat partial liver model of I/R injury was established,and the samples were collected at the 24th h after the I/R The serum ALT and AST levels were determined,the histologic changes were observed by HE staining under the light microscopy,the frequency of CD4~+ CD25~+ T cells among mononuclear cells in liver tissue was analyzed by using flow cytometry,the expression of Foxp3 mRNA was detected in liver tissue by real-time PCR,and the serum TGF-β,IL-10 levels were measured by enzyme-linked immunosorbent assay (ELISA).Results Serum ALT and AST levels were significantly decreased and the histological damage was significantly alleviated in the sirolimus-operated group as compared with saline-operated group(P<0.05).The percentage of CD4~+ CD25~+ T cells among mononuclear cells in groups A,B,C,and D was(6.12±1.87)%,(22.36±6.75)%,(4.53±1.02)% and(13.29±3.16)% respectively in liver tissue The expression levels of the Foxp3 mRNA were significantly higher in sirolimus group than in saline group(P<0.05).The ELISA showed that sirolimus could significantly increase the levels of TGF-β and IL-H)(P<0.05).Conclusion Pretreatment of sirolimus can effectively protect against liver ischemia-reperfusion injury in rats,which may be related to induction of CD4~+ CD25~+ Foxp3~+ T regulator cells by sirolimus,and the increase of TGF-β and IL-10 secretion to inhibit the imflammatory response.

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Clinical analysis of 2520 renal transplantations in one center

Rending WANG ; Qiang HE ; Jianyong WU ; Xuanmin WANG ; Zhangfei SHOU ; Hongfeng HUANG ; Jianguo ZHANG ; Suya WANG ; Jianghua CHEN

Chinese Journal of Organ Transplantation.2009;30(12):725-728. doi:10.3760/cma.j.issn.0254-1785.2009.12.005

Objective To evaluate the effects of different strategies on short-and long-term clinical outcomes of renal transplantation in Chinese subjects.Methods 2520 renal transplantations were retrospectively evaluated,including 2490 first renal transplantations and 30 second renal transplantations.Triple-immunosuppressant including cyclosporine A,azathioprine or myeophenolate mofetil(MMF)and prednisone(Pred)was adopted.Patients receiving kidney transplantation were given low dose immunosuppressants since 2000.Immunosuppressants including tacrolimus,MMF and Pred were adopted in some patients since 2000.Risk factors leading to graft loss and patients'death were analyzed.Results Until the cut date of June 30,2009,135 patients lost follow-up,and the follow-up rate was 94.6%.Incidence of acute(within 6 months post-transplantation) rejection was 18% among 2520 patients.Incidence of acute rejection (within 6 months post-transplantation) was 25.7% in panel reactive antibody (PRA) positive patients,significantly higher than 17.0% in PRA negative patients(P<0.05).Incidence of acute rejection within 6 months post-transplantation was 16.9% in HLA mismatches<4 patients,significantly lower than 23.7% in HLA≥4 patients (P<0.01).Total patient/death censored graft 1-,3-,5- and 1O-year survivals were 94.5%/96.0%,91.6%/93.1%,88.5%/90.1% and 81.7%/80.6%,respectively.Acute rejection and immunosuppressant regimen were independent risks for allograft loss.1mmunosuppressant regiment,pulmonary infection,cardio-brain-vessel accident, hepatic failure and tumor were independent risks for patients' death.Conclusion Renal allograft and patient survival appeared to be improved by optimal immunosuppressant regimen,strict HLA match and efficient post-transplant complication prophylaxis.

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Cellular transplant applied with rat bone marrow stromal cells preconditioned with stromal-derived factor 1 to treat acute myocardial infarction

Jun CHEN ; Kailun ZHANG ; Xinling DU

Chinese Journal of Organ Transplantation.2009;30(6):362-365. doi:10.3760/cma.j.issn.0254-1785.2009.06.012

Objective To investigate the effects of preconditioning (PC) with stromal-derived factor 1 alpha (SDF-1) on the levels of apoptosis of bone marrow stromal cells (BMSC) treated with hypoxia plus serum deprivation, and observe the therapeutic efficacy of cellular transplant with BMSC preconditioned with SDF-1 in rats with acute myocardial infarction. Methods BMSC were cultured with the whole marrow-adherence way. RT-PCR and immunohistochemistry were used to determine the expression of CXCR4. BMSC were incubated in medium for 24 h with 10 and 100 μg/L SDF-1 respectively, then treated with hypoxia plus serum deprivation for 6 h. The levels of apoptosis were detected by flow cytometry and TUNEL method. Acute myocardial infarction (AMI) model was established in SD rats, and BMSC preconditioned or non-preconditioned with SDF-1 were transplanted into border zone around infarct area, then heart function was measured after two weeks by ultrasonography. Results BMSC exhibited the CXCR4 expression. The number of apoptotic cells was significantly reduced in SDF-1 PC group than in control group (P<0.05), and 100μg/L SDF-1 PC group had the lowest level of apoptosis. AMI model was established successfully. Two weeks after BMSC transplant, significant improvement in cardiac function was observed in 100 μg/L SDF-1 PC group as compared with the non-PC group (P<0.05). Conclusions PC with the chemokine SDF-1 suppresses the apoptosis of BMSC treated with hypoxia plus serum deprivation. SDF-1 PC is a novel approach for enhancing therapeutic efficacy of cellular transplant in rats with AML

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Significance of histopathologic analysis vs cytomegalovirus (CMV) detection in the diagnosis of CMV-infected hepatitis post liver transplantation

Long CHENG ; Shuguang WANG ; Zhanyu YANG ; Ke ZHANG ; Xiaochu YAN

Chinese Journal of Organ Transplantation.2009;30(6):348-350. doi:10.3760/cma.j.issn.0254-1785.2009.06.008

Objective To evaluate the sensitivity and specificity of histopathologie analysis vs cytomegalovirus (CMV) detection for the diagnosis of CMV-infected hepatitis post liver transplantation. Methods Twenty-five biopsies with CMV infection and twenty-five without CMV infection were collected. Histopathologic observation, immunohistochemical staining and virus detection were performed on both groups to evaluate the sensitivity and specificity of these examinations for the diagnosis of CMV-infected hepatitis. Results The detection rate of microabscess, aggregation of monocyte and rnacrophage, and cytomegalic change in CMV infection group was higher than that in the group without CMV infection (P<0.05), but there was no significant difference in intranuclear inclusion and eosinophilic body between the two groups (P>0.05). The sensitivity and specificity of IHC and PCR for CMV detection were 20% and 100%, 72% and 84%, respectively. Conclosions CMV detection with PCR combined with histological observation is the most effective diagnostic scheme for CMV disease of liver.

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Expression and significance of IFN-γ and IL-10 in the process of graft rejection following liver transplantation in rats

Jie CHEN ; Yong CHEN ; Tenggang DENG ; Jianping GONG

Chinese Journal of Organ Transplantation.2009;30(10):601-603. doi:10.3760/cma.j.issn.0254-1785.2009.10.006

Objective To explore the expression of IFN-γ and IL-10 in lymphocytes in the process of the graft rejection process following liver transplantation in rats and their biological significances.Methods Orthotopic liver transplantation (OLT) models were established by modified Kamada two-cuff technique.Recipients were divided into three groups.The allograft group underwent OLT from male Wistar-to-Sprague-Dawley (SD) rats.The isograft group of SD-to-SD liver transplantation was complemented with a control group of normal male SD rats that had sham operations.The histological changes were examined,as well as serum and Mrna expression levels of IFN-γand IL-10.Results Serious morphological changes were observed in the allografted hepatic tissue.Many necrotic hepatocytes were seen and vacuolar degeneration of kytoplasm,karyopycnosis even nuclear fragmentation was observed in biliary duct endothelial cells.Many inflammatory cells infiltrated the portal areas,and the architecture of the liver lobules was destroyed.In contrast there were mild morphological changes in isograft hepatic tissue .A few inflammatory cells infiltrated the portal area;the hepatic lobule structure remained and biliary duct endothelial cells were nearly normal.Serum IFN-γ levels (pg/ml)determined by ELISA in the allograft group and isograft group were 386.7±14.4 and 159.8±16.5 respectively(P<0.05);for IL-10,they were 126.3±13.1 and 288.3±17.1,respectively(P<0.05).The semiquantitative reverse-transcriptase polymerase chain reaction assay showed that expression levels of IFN-γand IL-10 Mrna were similar to those in serum.Conclusion The expression of IFN-γ was obviously advanced in graft rejection following liver transplantation in rats and the expression of IL-10 was depressed. The dynamic equilibrium of T_H1/T_H2 cytokines is very important in the process of graft rejection following liver transplantation in rats.

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The effects of cryopreservation on the immunogenicity of HUVEC

Weifu CHEN ; Weihua YAN ; Boli LI ; Min ZHU ; Baoguo CHEN ; Qin ZHAO ; Zhenghua HONG ; Zhong ZHU ; Haixiao CHEN

Chinese Journal of Organ Transplantation.2009;30(10):594-597. doi:10.3760/cma.j.issn.0254-1785.2009.10.004

Objective To study the effects of cryopreservation on the immunogenicity of human umbilical vein endothelial cells(HUVEC).Methods HUVEC were isolated ex vivo and cryopreserved.Lymphocyte stimulation index(SI)was analyzed by MTT in lymphoeyte-endothelial cell co-culture.Both HLA-ABC and HLA-DR antigen expression on fresh or cryopreserved HUVEC,and the effects of IFN-γ treatment on HLA antigen expression in both groups were determined by using flow cytometry.Results No difference in SI was observed between fresh prepared and cryopreserved HUVEC(1.716±0.181 vs 1.686±0.145,P>0.05).The percentage of HLA-ABC expression was(96.6±1.9)%and(96.0±1.4)%in fresh and cryopreserved HUVEC(P>0.05),and the mean intensity for HLA-ABC expression was 84.1±5.7 and 82.4±4.8 in fresh and cryopreserved HUVEC(P>0.05),respectively.However,no HLA-DR expression was observed in both groups.When treated with IFN-γ,HLA-ABC expression was significantly up-regulated,and HLA-DR expression was induced in a dose-dependent manner.No significant difference was found in the HLA-ABC expression between fresh and cryopreserved HUVEC(P>0.05),while the HLA-DR expression in cryopreserved HUVEC was remarkably lower than in fresh HUVEC with the increase of IFN-γ(P<0.01).Conclusion The immunogenicity of HUVEC remains stable by cryopreservation without IFN-γtreatment or treated with low concentration of IFN-γ(≤50 U/ml).However,the HLA-DR expression in HUVEC was remarkably reduced in eryopreserved cells treated with a high concentration of IFN-γ(≥100 U/ml).These data indieated that the effects of cryopreservation on immunogenicity of HUVEC may result from the decreased responses of HLA-DR expression by the stimulation of IFN-γ treatment.

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Allograft survival in diabetic mice transplanted with Sil-1RI gene-modified islet cells

Yi TANG ; Shaohua SONG ; Fang LIU ; Quanxing WANG

Chinese Journal of Organ Transplantation.2009;30(10):581-584. doi:10.3760/cma.j.issn.0254-1785.2009.10.001

Objective To discuss the effect of sIL-1RI on allograft survival after islet transplantation.Methods Islets were isolated and transfected with Ad-sIL-1RI-Ig.Mice were treated with STZ to induce insulin-dependent diabetes mellitus(IDDM) model.Islet transplantation was carried out to IDDM mice with sIL-1RI-Ig gene-modified islet cells.Then the survival time of grafts was tested by daily observing blood glucose and insulin levels.The expression of cytokines was detected in graft after transplantation by using RT-PCR. Pathological changes of the graft were also observed by chromoscopy with HE after transplantation.Results The survival time of the grafts in sIL-1RI-Ig-islet group (39±3 days) was prolonged significantly (P<0.01) as compared with controls.A down-regulation of cytokines expression was detected in grafts after transplantation.Less damage and infiltration of lymphocytes were found in sIL-1RI-Ig gene-transfected group.Conclusion The effects of islet cells modified with sIL-1RI-Ig before transplantation on the rejection of murine islet allograft were investigated.The results verified that sIL-1RI-Ig-modified islet allograft could prolong the survival of grafts significantly,and demonstrated it was possible that sIL-1RI-Ig preventedallograft rejection via reducing the expression of cytokines(TNF-α,IFN-γ,RANTES,etc.).

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Clinical report of 18 cases of preventing hematuria by tube flushing in donor duodenum after combine pancreas-kidney transplantation

Wenli SONG ; Yingxin FU ; Gang FENG ; Zhongyang SHEN

Chinese Journal of Organ Transplantation.2009;30(9):559-561. doi:10.3760/cma.j.issn.0254-1785.2009.09.013

Objective To investigate the clinical effects of tube flushing in donor duodenum to prevent the hematuria post combined pancreas-kidney transplantation(SPK)with pancreatic fluid drainage through bladder.Methods 18 cases of diabetic patients associated with end-stage renal disease were subjected to combined pancreas-kidney transplantation with pancreatic fluid drainage through bladder,within which 12 cases were pre-placed douche tube in donor duodenum,while the other six were not.As for the cases in group with the tube,T tube of No.10 was put in the donor duodenum through the abdominal wall and then bladder.After that the tube was fixed using 5-0 absorbable suture,then the bladder sutured if the tube was smooth confirmed by flushing with saline.After the operation,flushing was maintained using saline consecutively with the speed of 500 ml/h through the douche tubes of these 12 patients.Then the speed was changed to 250 ml/h 3 days later if the flushing fluid was limpid.One week later,changed to rinse intermittently and prolonged the flushing interval gradually.Till 14 days post the operation,flushing was ceased.After 2 days' survey,the urethral catheter was removed.As for the other 6 cases without douche tube,the urethral catheter was removed during 7-10 days after the operation if hematuria didn't occur.Results In the 12 cases with douche tube,there was only one patient(8.3 %,1/12)having slight hematuria on the 7th day after the cessation of the bladder washout.Through strengthening the flushing,the hematuria disappeared.The urethral catheter was removed on the 14th day after the operation and the hematuria never happened again.In the group without the douche tube,4 cases(66.7 %,4/6)had serious hematuria complicated with bladder obturation.The incidence of that was obviously higher than in the group with the douche tube(P＜0.05).Only one patient(1/12,8.3%)in the group of regular insertion of douche tube had urinary system infection,but in the group without the tube,the incidence of urinary system infection was 66.7 %(4/6)(P＜0.05).Conclusion The tube flushing in donor duodenum can significantly reduce the occurrence of hematuria after combined pancreas-kidney transplantation with pancreatic fluid drainage through bladder.

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Efficacy and safety of routine intraoperative insertion of double-J stents in kidney transplantation;a systematic review

Xianding WANG ; Shaoxiang MA ; Jing ZHUANG ; Ke WU ; Guobiao LIANG ; Xiaoke HUANG ; Yiping LU

Chinese Journal of Organ Transplantation.2009;30(9):540-544. doi:10.3760/cma.j.issn.0254-1785.2009.09.008

Objective To systematically evaluate the effect of routine insertion of double-J stents to prevent major urological complications(MUCs)in kidney transplant recipients.Methods Medline,Embase,Cochrane Library,and Chinese Biomedicine database were searched to locate relevant randomized controlled trials(RCT).Data extraction and assessment of methodologic quality were performed independently by two reviewers.Meta-analysis was performed by Revman 5.0 software.Results Ten RCT(including 1616 patients)were identified.By comparing the routine stent group with the no stent group,the meta-analysis showed:(1)incidence of urine leak,urinary obstruction and UTI was 4 times lower,6 times lower,increased by 52 % respectively(P＜0.0001);(2)Patient and graft survival,rate of acute rejection,delayed graft function and hematuria were of no significant difference.In subgroup analysis,it was found:(1)Compared with the no stent group,the group in which stent duration was≤ 4 weeks had a lower incidence of MUCs and a higher incidence of UTI;meanwhile,the group in which stent duration was ＞ 4 weeks had a much lower incidence of MUCs and the rate of UTI was increased without significant difference;(2)In the RCT of which urethral catheter duration was ＜ 5 days,there were no significant differences between the two groups in MUCs and UTI.In the RCT of which urethral catheter duration was ≥5 days,the stent group had a lower incidence of MUCs and a higher incidence of UTI.Conclusion Routine stenting reduces the incidence of MUCs.Although the double-J stent increases the risk of UTI,it seems that UTI doesn't affect the outcome of transplantation.The stent duration should be within 4 weeks.For the stent recipients,the longer duration of urethral catheter,the lower incidence of MUCs,the higher incidence of UTI;thus,it is up to clinicians to decide the optimal duration of urethral catheter.Long term prescription of 480 mg cotrimoxazole once daily,from the operation day till after stent removal,effectively reduces the risk of UTI associated with stent placement.

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Immunologic and pathologic features of an accelerated rejection model of renal allotransplantation in rhesus monkeys

Song CHEN ; Junhua LI ; Ying XIANG ; Shan ZHONG ; Hui GUO ; Hong WANG ; Ying WU ; Jianguo ZHU ; Shi CHENG ; Gang CHENG

Chinese Journal of Organ Transplantation.2009;30(9):515-518. doi:10.3760/cma.j.issn.0254-1785.2009.09.001

Objective To study the immunologic and pathologic features of an accelerated rejection model of renal allotransplantation in presensitized monkeys.Methods The accelerated rejection model of renal allotransplantation was established in presensitized monkeys,which received donor skin transplantation in advance(n=3).The changes of donor specific antibody(DSA)levels in the recipient monkeys before/after skin and kidney transplantation were measured.The kidney grafts were examined for routine pathology,antibody and complement depositions,various lymphocyte subsets infiltration by HE staining,immunofluorescence,or immunohistochemistry.Results All renal allografts in 3 presensitized monkeys developed accelerated rejection within 4 days.In 2 presentized monkeys,the levels of DSA and their mediated complement-dependent cytotoxicity(CDC)were significantly increased after skin transplantation,and further markedly elevated at the time of kidney graft rejection.In the rejected renal grafts,massive C3,C4,C5b-9 and IgG deposits with few lymphocytes infiltration were found.Typical pathologic changes included severe arterionecrosis,thrombosis,interstitial hemorrhage,and infiltration of neutrophils.In the rest one presentized monkey,the levels of DSA and CDC were only marginally increased,and the pathological changes of the rejected renal graft were characterized mainly by the injury of renal tubules.Conclusion Presensitization by donor skin transplantation could elevate the levels of DSA and CDC in recipient monkeys,which resulted in severe antibody-mediated acute humoral rejection in most of the following renal transplants.

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