Prolongation of rat skin and cardiac allograft survival by low molecular weight heparin.

BACKGROUND: We have previously reported that very low doses of low molecular weight heparin compounds (LMWH) inhibit a variety of T-cell-mediated reactions by down-regulation of TNF-alpha production. This study tested the efficacy of LMWH in organ transplantation. METHODS: Skin and heterotopic heart transplantations were performed between recipient Wistar rats and donor BN rats. Two doses of LMWH were given sc, 1 and 20 micrograms, each in three protocols, with day of grafting as Day 0: (A) Daily: -1, 0, 1 ellipsis, (B) Late Weekly: -1, 6, 13 ellipsis, and (C) Early Weekly: -7, 0, 7 ellipsis. Doses and schedules were selected based on efficacy in autoimmune models. Skin graft rejection was defined by complete separation of the graft, and heart transplant rejection was defined as cessation of heartbeat. RESULTS: Treatment with 1 microgram (26.8 +/- 2.0 days) and 20 micrograms (24.5 +/- 2.3 days) of LMWH using the Early Weekly protocol significantly prolonged skin allograft survival compared to controls (17.8 +/- 4.4 days), P < 0.001 for both, whereas other protocols did not. Compared to controls (8.3 +/- 1.4 days), treatment with both 1 and 20 micrograms of LMWH using all three protocols significantly prolonged cardiac allograft survival. The efficacy, however, varied considerably. Increase in graft survival ranged from 18% (1 microgram, Daily, 9.8 +/- 0.7 days, P = 0.02) to more than twofold (20 micrograms, Early Weekly, 20.8 +/- 5.5 days, P < 0.001) according to the dose and schedule of LMWH. CONCLUSIONS: Treatment with very low doses of nonanticoagulant LMWH preparations having anti-TNF-alpha activity significantly prolongs rat skin and cardiac allograft survival in a dose- and schedule-dependent manner.     

Prolongation of renal allograft survival in the rat treated with amniotic fluid.

To assess the role of amniotic fluid (AMF) in the maintenance of pregnancy, immunosuppressive effects of AMF were studied in vivo, and the mechanisms of suppressor activity were analyzed immunologically in vitro in the rat. Female Lewis (LEW, RT-1l) rats mated with Brown-Norway (BN, RT-1n) rats for 14 days were sacrificed and cell-free AMF was obtained. AMF was diafiltered with PBS (PH 7.2) and reconstituted to 2 OD units measured at 280 nm. Untreated LEW hosts rejected BN renal grafts at 7.8 +/- 0.2 days (n = 10). Five days of intravenous inoculation of AMF into LEW hosts remarkably enhanced BN graft survivals (MST = 20.3 +/- 4.4 days, n = 12) compared with controls (P less than 0.01), and slightly prolonged third-party DA (RT-1a) graft survivals (MST = 9.4 +/- 0.8 days, n = 7) compared with control LEW hosts engrafted with a DA kidney (MST = 7.6 +/- 0.2 days, n = 6). Five days of intravenous inoculation of pregnant sera into LEW hosts had no effect on BN graft survival. The AMF suppressed the proliferative response of LEW lymphocytes against not only irradiated BN stimulator cells but also irradiated third-party DA stimulators. The AMF also suppressed allokiller T cell generation of normal LEW lymphocytes against BN cells by 70.1% and 51.3%, and against DA cells by 64.9% and 38.9% at concentrations of 25% and 12.5%, respectively (P less than 0.01). To dissect the immunosuppressive activity of AMF, the effect of AMF on cytokine production and interleukin 2 (IL-2) receptor expression of concanavalin A-stimulated lymphocytes were investigated. AMF suppressed interferon and IL-2 production. Interestingly, however, AMF did not suppress interleukin 3 (IL-3) and interleukin 6 (IL-6) production, as well as IL-2 receptor expression. These results demonstrated that rat AMF displayed a strong immunosuppression in vivo as well as in vitro, and that AMF might play an important role in the maintenance of pregnancy.     

Prolongation of rat heart allograft survival with K(+)ATP-dependent channel modulators

Controversies exist regarding the immunoregulatory properties of K(+) ATP channel modulators. We investigated the effects of aprikalim, a K(+) ATP-dependent channels activator, and glibenclamide and gliclazide, two inhibitors of K(+) ATP-dependent channels, on the prolongation of heart allograft survival in the rat. Nine groups (n >/= 5) were involved in this study with the Brown-Norway to Lewis rat combination treated with aprikalim, glibenclamide, gliclazide, and/or cyclosporine. The results indicate that modulators of K(+) ATP-dependent channels can improve the survival of rat heart allograft without interfering with the immunosuppressive properties of cyclosporine.     

Prolonged survival of rat cardiac allograft with proinflammatory cytokine inhibitor.

BACKGROUND: Proinflammatory cytokines, such as tumor necrosis factor (TNF-alpha) and interleukin-1 (IL-1), play important roles in acute allograft rejection. FR167653 is an inhibitor of these cytokines that acts through inhibition of the mitogen-activated protein kinase p38 pathway. We examined the effect of FR167653 on allograft rejection. METHODS: We used Brown-Norway and Lewis rats as donors and recipients, respectively. We performed heterotopic cardiac transplantation. The control group consisted of untreated rats. In the experimental groups, recipients were intraperitoneally injected with FR167653 just after operation, followed by daily injection of the drug from Day 1 to 10. We divided 20 rats into 5 groups, which received varying doses of FR167653, ranging from 75 to 300 mg/kg/day. RESULTS: In the control group, the mean graft survival was 6.8 +/- 0.3 days. FR167653 at 150 mg/kg/day significantly prolonged the survival period (up to 12.1 +/- 1.5 days, p = 0.002). Histologically, FR167653 markedly suppressed cellular infiltration on Day 5 post-transplantation. The serum level of TNF-alpha in the control group was persistently elevated from 9.3 +/- 3.9 pg/ml to 11.3 +/- 3.8 pg/ml, whereas FR167653 significantly suppressed the level to <1.4 +/- 1.4 pg/ml. CONCLUSIONS: FR167653 prolonged rat cardiac allograft survival by suppressing the action of proinflammatory cytokines.     

Th2细胞因子在大鼠心脏移植排斥反应中的变化

目的：探讨Th2细胞因子在移植排斥反应中的作用及意义。方法：用酶联免疫法（ELISA）检测大鼠异位心脏移植术后不同时间外周血中白细胞介素4（IL－4）及白细胞介素10（IL－10）的变化。结果：术后IL－4及IL－10的变化与排斥反应的进程有关,两者的峰值均出现于术后第5d,其中IL－10升高则更为明显,此时移植物排斥反应表现为2－3级。当排斥反应加重,出现明显组织坏死时（4级）,血清IL－4及IL－10水平下降。应用环孢素（A（CsA)后,延援了排斥反应的发生及IL－4、IL－10水平的升高。结论：IL－4及IL－10在排斥反应的较早阶段可能发挥重要作用,动态监测外周血IL－4及IL－10可能有助于对临床移植排斥反应 状态的评价。     

Prolongation of cardiac allograft survival by inhibition of ERK1/2 signaling in a mouse model

BACKGROUND: It has been demonstrated that in vitro the presence of extracellular signal-regulated kinase 1 and 2 (ERK1/2) signaling inhibitor suppresses T cell activation and Th1 development. However, pharmacological interference of ERK1/2 signaling by administration of its small molecule inhibitor has not been tested as a therapeutic target in the prevention of allograft rejection. METHODS: The immunosuppressive effect of targeting ERK1/2 signaling was tested on cardiac allograft survival in C57BL/6 (H-2b) to Balb/c (H-2d) murine model using PD98059 inhibitor. Phosphorylation/activation of ERK 1/2 and STAT6 proteins were assessed by Western blot. RESULTS: Blockade of ERK1/2 using PD98059 had significant immunosuppressive effect and prolonged survival of mouse cardiac allografts from 8.3+/-0.5 days (vehicle) to 12.6+/-1.3 days (100 mg/kg PD98059; P<0.0001). Combination therapy of PD98059 (100 mg/kg) with cyclosporine (CsA, 15 mg/kg for 20 days) additionally enhanced graft survival (34.4+/-1.2 days) compared to CsA (14.9+/-1.1 days; P<0.0001) or PD98059 monotherapy (P<0.0001). Attenuation of graft rejection by PD98059 correlated to reduction of intragraft ERK phosphorylation and leukocyte infiltration, and to increase in interleukin (IL)-4 or decrease in interferon-gamma production within the grafts. In vitro inhibition of ERK1/2 by PD98059 promoted Th2 differentiation by upregulation IL-4 production but not altering IL-4 stimulating STAT6 pathway. CONCLUSION: Targeting ERK1/2 signaling results in suppression of alloimmune responses by an unique mechanism that involves Th1/Th2 skewing, suggesting a therapeutic potential of inhibition of ERK1/2 signaling for transplant rejection, particularly in combination with CsA.     

Prolongation of rat cardiac allograft survival by treatment with prostacyclin or aspirin during acute rejection

Hearts taken from DA (RT1a) rats were transplanted heterotopically to PVG (RT1c) rats of the same sex (day 0). On day 1 or on day 5 rats were treated with prostacyclin (PGI2), 250 ng/kg/min, by continuous infusion of alkaline solution into the inferior vena cava until the time of rejection. Controls received glycine buffer infusion alone, from day 1 or day 5. Cessation of palpable graft beat was taken as the end point of rejection. When PGI2 was infused from day 5 median graft survival time was prolonged from a control of 7.8 days to 9.3 days (P less than 0.05). When PGI2 was infused from day 1, graft survival time was prolonged from a control of 7.4 days to 8.6 days (P less than 0.05). Other groups of rats were treated from day 1 or from day 5 with aspirin (acetylsalicylic acid), 200 mg/kg/day, by 8-hourly subcutaneous injection in saline. Control groups received saline alone. When aspirin was given from day 5, graft survival time was prolonged from a control of 7.3 days to 9.5 days (P less than 0.05). When aspirin was given from day 1 graft survival time was prolonged from a control of 7.2 days to 14.9 days (P less than 0.01), and in two cases this led to very prolonged survival. Histological examination at the time of rejection showed lymphocyte and neutrophil infiltration to be much more prominent than vessel occlusion in all groups. These results imply that PGI2 and aspirin may be beneficial to graft survival in acute rejection, but this is not due to reduced occlusion of blood vessels by platelets.     

Prolongation of skin allograft survival in rats by a novel immunosuppressive agent, FK506

FK506, an immunosuppresant, was isolated from Streptomyces tsukubaensis. Intramuscular administration of FK506 (0.32 mg/kg or more) 5 days a week for two weeks after grafting prolonged the acceptance time of F344 skin allograft to WKA rats. Similar results were obtained with cyclosporine at 32 mg/kg or more, but other immunosuppressives (i.e., prednisolone, azathioprine, and bredinin) gave only a marginal prolongation. The prolonging effect of FK506 was obtained in various donor-recipient combinations across a major or minor histocompatibility barrier. The agent also prolonged the acceptance time of mouse skin xenografts to rats. Furthermore, maintenance doses of 3.2 or 0.32 mg/kg twice a week after an initial 14-day treatment with the agent at 3.2 mg/kg gave graft survival as long as the treatment was continued for more than 120 days. Our findings show that FK506 has a potent immunosuppressive effect in rats and suggest that the agent merits further investigation.     

供者骨髓基质干细胞输注延长大鼠移植心存活时间的探讨

目的 探讨供者骨髓基质干细胞(MSC)输注在大鼠同种心脏移植术后的免疫调节及延长移植心存活时间的作用.方法 供者为近交系Wistar大鼠,受者为Fisher 344大鼠.处死供者后抽取其股骨和胫骨中骨髓,分离和培养MSC.通过混合淋巴细胞试验观察不同密度的MSC对异源性T淋巴细胞增殖反应的抑制作用.建立大鼠异位心脏移植模型,根据处理方式的不同,将受者分为MSC输注组和对照组,每组8只.Msc输注组:将含有2×106个MSC的林格氏液分别于术前1周、术中及术后连续3 d经尾静脉注入受者体内;对照组:用与MSC输注组相同的方法在相同时间点注入不含MSC的林格氏液.术后第5天,采用实时逆转录聚合酶链反应检测移植心组织中细胞因子的表达情况.结果 供者MSC可明显抑制异源性T淋巴细胞的增殖反应,且MsC密度越高抑制作用越强.MSC输注组Th1类细胞因子白细胞介素(IL)-1β和γ干扰素的表达要显著低于对照组;MSC输注组Th2类细胞冈子IL-4和IL-10呈高表达,而对照组基本不表达.MSC输注组移植心平均存活时间为(12.4±5.3)d,与对照组的(6.4±2.0)d比较,差异有统计学意义(P＜0.01).结论 输注供者MSC可通过改变Th1/Th2类细胞因子的平衡向Th2偏移诱导受者产生免疫调节作用,从而延长移植心存活时间.     

Donor G protein beta3 subunit 825TT genotype is associated with reduced kidney allograft survival.

Recent studies have identified a novel polymorphism (C825T) of the gene encoding the beta3 subunit of heterotrimeric G proteins (Gbeta3), associated with enhanced activation of G proteins, which appears to be more common in hypertensive patients. In the present study, the relationship between this genetic variant and kidney allograft survival was examined over the first 3 yr after transplantation, in 320 consecutive Caucasian patients recruited from the Berlin-Steglitz transplantation center between 1988 and 1993. Clinical parameters, transplantation data, and details of graft survival were retrieved from clinical records. After multivariate adjustment for covariates (Cox hazard regression), the Gbeta3 825TT donor-genotype was associated with a significantly decreased graft survival representing a relative risk of graft loss of 2.2 (95% confidence interval, 1.1 to 4.8) compared to TC and CC grafts within the observation period. This association between donor TT genotype and graft survival remained stable even after stepwise exclusion of covariates from the multivariate model. In contrast, there was no significant relationship between recipient genotype and allograft function. These findings indicate that individuals receiving renal allografts from donors homozygous for the Gbeta3-825T allele may have an increased risk of developing allograft failure. Additional studies on the role of this genetic marker as well as the role of pertussis toxin-sensitive G proteins in the development of chronic rejection appear warranted.