Combination use of suboptimal dose of FK 506 and cyclosporine in canine lung transplantation.

The immunosuppressive potency and the side effects of combination therapy with FK 506 and cyclosporine A were studied in dogs that had undergone lung transplantation. The animals were divided into four groups: group A (one third optimal FK 506 dose: FK 506, 0.03 mg/kg intramuscularly) (n = 5), group B (one third optimal cyclosporine dose: cyclosporine 6 mg/kg orally) (n = 5), group C (one third FK 506 and one third cyclosporine optimal doses): FK 506, 0.03 mg/kg intramuscularly plus cyclosporine 6 mg/kg orally) (n = 5), and group D (half FK 506 plus half cyclosporine optimal doses: FK 506, 0.05 mg/kg intramuscularly, plus cyclosporine, 10 mg/kg orally) (n = 10). Assessments including chest x-ray film, fiberoptic bronchoscopy, hematologic and biochemical tests, FK 506 and cyclosporine blood trough level measurement, right pulmonary artery occlusion test, and histopathologic observations were performed. In group A two of five dogs survived 28 days and three died on postoperative days 7, 14, and 21. In group B one dog survived 28 days and four died on postoperative days 9 (two dogs), 14, and 21. Histologic examination showed severe rejection in both group A and group B. In group C all five dogs survived 28 days but showed mild rejection. In group D one dog died of intestinal bleeding on postoperative day 7 and nine survived 28 days. No pathologic changes were observed except in one case of mild rejection. The ventilation function of the transplanted lung was poor in groups A, B, and C but good in group D. No abnormal rise of FK 506 and cyclosporine trough levels was observed. There were no significant side effects and abnormal hematologic and biochemical data except in one dog in group D. We concluded (1) the combination of FK 506, 0.03 mg/kg, and cyclosporine, 6 mg/kg, is much more effective than either drug used singly, (2) the combination of FK 506, 0.05 mg/kg, and cyclosporine, 10 mg/kg, prevents rejection with tolerable side effects, and (3) no worse side effects are caused by combination therapy with FK 506 and cyclosporine than by either one used singly.     

Drug delivery system using microspheres that contain tacrolimus in porcine small bowel transplantation

Abstract Rejection remains a major barrier to successful bowel transplantation, in spite of improved immunosuppressive techniques. Therefore, new, more effective, immunosuppressants, with fewer side effects, are needed. Biodegradable microspheres containing tacrolimus (FK506) were used in an experimental porcine small bowel transplantation. Twenty pigs underwent transplantation and were divided into four groups according to the immunosuppressive regimen. Group A (n = 5): no immunosuppression; group B (n = 6): 0.2 mg/kg per day of FK506; group C (n = 3): 1.0 mg/kg per day of FK506; group D (n = 6): 0.04 mg/kg per day of FK506 contained in biodegradable microspheres. Rejection was diagnosed macroscopically by endoscopic examination and histologically by biopsy specimen analysis. The mean survival time and standard deviation (SD) were 8.8±3.5, 11.0±1.4, 9.7±2.5 and 28.6 ±22.5 days for groups A, B, C, and D, respectively, with a statistically significant difference found between group D, on the one hand, and groups A, B and C, on the other. The mean trough blood concentration of FK506 was 10.5 ±2.2, 27.9 ±6.0 and 10.5 ±3.5 ng/ml in groups B, C and D, respectively. In groups A and B, all pigs died of rejection, without infection. In group C, all died of infection, without rejection. In contrast, none of the pigs in group D developed rejection or infection. Our results clearly show that the drug delivery system using biodegradable micro-spheres that contain FK506 is effective for controlling rejection with fewer side effects in the porcine small bowel transplantation.     

Removal of cartilage rings prevents graft stenosis in extended tracheal allotransplantation with omentopexy and immunosuppression: an experimental study.

BACKGROUND: One of the serious problems in longer size tracheal transplantation is severe stenosis of the graft, probably caused by an inadequate blood supply. We have previously reported that removal of some cartilage rings of the graft and omentopexy helps to provide sufficient blood flow to the graft mucosal tissue and results in satisfactory survival and non-significant graft stenosis in extended tracheal autotransplantation. However, it is unclear whether this method can be applied to extended tracheal allotransplantation that requires immunosuppression. In this report, we describe midterm results of extended tracheal allotransplantation with the technique. METHODS: Twenty-four adult mongrel dogs were used. In 18 dogs, a nine-cartilage-ring length of the trachea was allotransplanted when five cartilage rings of the graft were removed, leaving two rings intact at both ends of the graft for simple fixing to the recipient. Two artificial tracheal rings outside the graft and a stent inside the graft were used for maintaining the lumen width. Omentopexy was done for sufficient blood supply to the graft. FK 506 (0.1 mg/kg) was given on each day after the operation in Group A (n = 10), but was not given at all in Group B (n = 8). In Group C (n = 6), a nine-cartilage-ring length of the trachea, without removal of any cartilage ring, was transplanted into the recipient dog and covered with an omental pedicle flap. The same dose of FK 506 as that used in Group A dogs was given to Group C dogs. RESULTS: In Group A, 2 dogs died of graft stenosis within 9 weeks after surgery and 1 died of emaciation without tracheal stenosis. Seven dogs (70%) survived until time of killing. Among the 8 dogs in Group B, 6 died of graft stenosis within 9 weeks after surgery, with 1 dying of pneumonia and only 1 (13%) surviving for >1 year until killing. In Group C, all 6 dogs died of graft stenosis within 6 weeks after surgery. Survival at 16 weeks after surgery was 70% in Group A, 13% in Group B and 0% in Group C (p < 0.01, A vs B and C). No significant graft stenosis was found in 6 dogs and mild stenosis was found in 2 dogs at the time of death or killing in Group A (80%), whereas mild stenosis was found in only 2 dogs in Group B (25%) (p < 0.05). Mucosal blood flow of the graft in Group A was higher than that in Group C and was the same as that in Group B within 4 weeks after surgery; however, it remained unchanged to ultimately be higher than in Group B at 6 and 8 weeks after surgery. CONCLUSIONS: Removal of some cartilage rings, omentopexy and immunosuppression improved blood supply to the graft and resulted in good survival and non-significant tracheal stenosis in extended tracheal allotransplantation.     

Feasibility of cryopreserved tracheal xenotransplants with the use of short-course immunosuppression

OBJECTIVE: We evaluated the feasibility of discordant xenotransplantation of the cryopreserved trachea with intermittent immunosuppression to help solve the shortage of donor tracheas. METHODS: Two experiments were performed with heterotopic transplantation models in 14 guinea pigs and 85 rats. So that the minimal dose of FK506 for viable fresh xenografts could be determined, FK506 was given in escalating doses (0, 1.5, 2.5, and 3.5 mg/kg) for recipient animals after xenogeneic transplantation. With the goal of obtaining a long-term survival of the xenografts, the effect of cryopreservation on xenografts was assessed and thereafter different cycles of immunosuppression every third week were evaluated in fresh or cryopreserved xenografts in the second experiment. RESULTS: An FK506 dosage of more than 2.5 mg/kg per day was much more effective than smaller dosages, as demonstrated by morphologic assessment. A higher dosage of FK506 potentially delayed the rejection of xenografts and can thus maintain tracheal xenograft viability for less than 4 weeks in rat recipients. In experiment 2, the cryopreserved xenografts showed less histologic viability than fresh xenografts but greater patency of the lumen. The patency of cryopreserved xenografts was favorably maintained for a longer period than that of fresh xenografts with either the same number or more cycles of immunosuppression. CONCLUSIONS: We conclude that the synergistic effect of cryopreservation and adequate intermittent immunosuppression may enable tracheal xenografts to remain viable over longer periods.     

Co-administration of ketoconazole and tacrolimus therapy: a transplanted rat model

Background/aim The aim of this work is to study the effect of addition of ketoconazole to experimental kidney transplanted rat treated with tacrolimus and precludes the percentage of tacrolimus dose reduction. Material and methods The material of this work included 60 male Sprague Dawely rats subjected to renal allotransplantation. They were equally divided into five groups: Group I: served as control group, Group II: received FK506 3.2 mg/kg/bw, Group III: received FK506 2 mg/kg/bw, Group IV: received FK506 1 mg/kg/bw, Group V: received FK506 1 mg/kg/bw plus ketoconazole 20 mg/kg/day. FK506 trough level and laboratory investigations were determined at 0, 3, 7, 10, 14, and 27 days post-transplantation. Results In all groups loss of body weight was observed at day 27 after treatment compared to that before transplantation. Serum creatinine significantly increased at day 27 compared to the basal level in groups treated with 1.0 mg and 3.2 mg FK506 (1.80 ± 0.50 vs. 0.39 ± 0.06 P = 0.001) and (1.03 ± 0.26 vs. 0.50 ± 0.07 P = 0.001) respectively. While for 2.0 mg or 1.0 mg plus keto groups, no significant differences in serum creatinine levels over time (0.56 ± 0.22 vs. 0.44 ± 0.10 P = 0.106) and (0.55 ± 0.30 vs. 0.42 ± 0.08 P = 0.160) were observed. Conclusion Concomitant administration of ketoconazole and FK506 is safe and results in increase blood trough level concentration of FK506 with 50% dose reduction in transplanted rat model.     

Efficacy of transient treatment with FK506 in the early phase on cyclophosphamide-induced bone marrow chimerism and transplant tolerance across MHC barriers

BACKGROUND: The use of mixed allogeneic bone marrow chimerism to induce donor-specific transplantation tolerance has been extensively demonstrated. In the present study, we assessed the effect of combined use of a short course of FK506 and a single-dose cyclophosphamide (CYP) on the induction of tolerance and development of GVHD after allogeneic BMT. MATERIALS AND METHODS: Lewis rat (RT1(l)) recipients received BMT from Brown Norway (RT1(n)) donors on the next day after injection of CYP at a dose of 200 mg/kg. The recipients were further treated with no FK506 (n = 8), 0.3 mg/kg/day FK506 on days 10-16 (n = 6), or the same dose of FK506 on days 0-6 (n = 6). In a subgroup of animals, heterotopic heart transplantation was performed to investigate transplantation tolerance. RESULTS: Six of eight recipient rats that did not receive FK506 died of severe GVHD, while high levels of chimerism were induced. Recipients of FK506 in the later phase developed mild transient GVHD around 2 to 3 weeks after BMT and recovered thereafter; however, the level of chimerism was significantly decreased (2.8 +/- 2.3% on day 100). Treatment with FK506 in the early phase completely prevented the development of GVHD and induced stable allogeneic chimerism in the long-term (13.8 +/- 8.3% on day 100). These recipients with stable chimerism accepted subsequent BN heart allografts indefinitely (>200 days x 5), while rejecting third-party (BUF) heart allografts by day 12. CONCLUSIONS: Early transient FK506 promotes the induction of stable bone marrow chimerism without GVHD after BMT with CYP pretreatment. The timing of treatment with FK506 is critical with a view to preventing GVHD and inducing stable long-lasting chimerism.     

Effects of inhaled FK 506 on the suppression of acute rejection after lung transplantation: use of a rat orthotopic lung transplantation model.

BACKGROUND: FK 506 inhalant was recently developed for localized administration. We investigated its effects on acute lung allograft rejection and compared its efficacy with that of intramuscular administration of FK 506. METHODS: Rats (n = 123) with orthotopic left lung transplantation were divided into 9 groups. Six groups inhaled FK 506 (5 puffs, 10 puffs or 20 puffs per day), or were given intramuscular administration of FK 506 (0.05, 0.1 or 1.0 mg/kg/day). The other groups included rats receiving an isograft, rats with an untreated allograft, and a placebo group. All groups (n = 6 each) were monitored for 14 days post-operatively as an end-point and graft survival time was determined. The remaining animals were killed 4 days after transplantation. The histologic grade of rejection was determined for all groups (n = 6 each). With both (n = 3 each) inhalation therapy and intramuscular administration of FK 506, which showed similar degrees of effectiveness, both blood FK 506 concentration and cytokine expression in the graft and spleen were evaluated. RESULTS: FK 506 inhalation therapy extended allograft survival time and reduced histologic rejection on Day 4 in all groups. Graft survival time and histologic rejection scores at a dose of 10 puffs/day were comparable to those with 0.1 mg/kg/day of intramuscular FK 506. Trough concentrations of FK 506 in blood were detectable with 0.1 mg/kg/day of intramuscular FK 506, but not with 10 puffs/day. The messenger RNA expression levels of interferon-gamma in the lung allograft was suppressed significantly at a dose of 10 puffs/day. CONCLUSIONS: FK 506 inhalant enhances acute lung allograft survival with lower blood concentrations than when using comparable intramuscular administration.     

Transplantation of infantile bladder in rats: an alternative procedure for bladder augmentation

BACKGROUND: Our purpose was to evaluate whether bladder transplantation (BTx) can be used for bladder augmentation (BA). METHODS: Bladders from infantile Brown-Norway rats (less than 21 days old) were excised and each transplanted into a pouch created in the distal omentum of a 6-week-old Lewis rat (fully allogeneic BTx). No immunosuppressant was used in group I (n=12). Intramuscular FK506 was used daily from the day of BTx in group II (n=16; 0.2 mg/kg), group III (n=22; 0.6 mg/kg), and group IV (n=16; 1.2 mg/kg) until harvesting 3, 4, 5, or 6 weeks after BTx. FK506 was used for only 2 weeks in group V (n=12; 0.6 mg/kg/day) and group VI (n=12; 1.2 mg/kg/day). Syngeneic bladder transplants acted as controls (n=16). Hematoxylin and eosin staining was used to examine all grafts. In six rats from group III, BA was performed by anastomosing the graft to the recipient bladder 10 days after BTx. RESULTS: Each successfully transplanted graft appeared macroscopically as a thin-walled cyst. Rejection was seen in all grafts from groups I, II, V, and VI, and was minimal or absent in groups III and IV. On medium to long-term follow-up the only side effect of FK506 observed was reduced weight gain. Graft survival in the control group was 100%. BA was successful in all six cases, and the mucosa was normal throughout each augmented bladder. CONCLUSIONS: This is the first report of the successful transplantation of infantile tissue without vascular anastomosis. Because of the efficient, safe immunosuppression possible with FK506, our BTx technique could find clinical application for creating viable vesical tissue that could be used for BA.     

Co-administration of Ketoconazole and Tacrolimus Therapy: A Transplanted Rat Model

Background/Aim: The aim of this work is to study the safety and the effect of addition of ketoconazole to experimental kidney transplanted rat treated with tacrolimus and predicts the percentage of tacrolimus dose reduction. Material and Methods: The material of this work included 60 male Sprague Dawely rats subjected to renal allotransplantation. They were equally divided into five groups: Group I: served as control group, Group II: received FK506 3.2 mg/kg/bw, Group III: received FK506 2 mg/kg/bw, Group IV: received FK506 1 mg/kg/bw, Group V: received FK506 1 mg/kg/bw plus Ketoconazole 20 mg/kg/day. FK506 trough level and laboratory investigations were determined at 0, 3, 7, 10, 14, and 27 days post-transplantation. Results: In all groups loss of body weight was observed at day 27 after treatment compared to that before transplantation. Serum creatinine significantly increased at day 27 compared to the basal level in groups treated with 1.0 and 3.2 mg FK506 (1.80 ± 0.50 versus 0.39 ± 0.06 P = 0.001) and (1.03 ± 0.26 versus 0.50 ± 0.07 P = 0.001) respectively, while for 2.0 mg or 1.0 mg plus keto groups, no significant differences in serum creatinine levels over time (0.56 ± 0.22 versus 0.44 ± 0.10 P = 0.106) and (0.55 ± 0.30 versus 0.42 ± 0.08 P=0.160) were observed. Conclusion: Concomitant administration of Ketoconazole and FK506 in transplanted rat model is safe and results in increase of blood trough level concentration of FK506 with 50% reduction of its dose.     

Donor leukocytes combine with immunosuppressive drug therapy to prolong limb allograft survival

Donor leukocytes administered at the time of transplantation may prolong organ allograft survival. This study examined the effectiveness of donor leukocyte injection combined with immunosuppression for limb transplantation across the strong histocompatibility barrier of a Brown Norway donor to a Lewis recipient. Eight animals received 6 x 10(7) donor leukocytes injected on the day of transplantation. From day 1, FK506 (2 mg/kg/d), mycophenolate mofetil (MMF) (15 mg/kg/d), and prednisone (0.5 mg/kg/d) were administered for 2 weeks. After week 2, prednisone and MMF were both tapered by 20% of the initial dosage per week. After week 7, the animals received only FK506 (2 mg/kg/d). From week 8, FK506 was tapered to the maintenance dose of 0.8 mg/kg/d at week 10 and was stopped on week 24. A control group of 8 animals underwent identical treatment except that the leukocyte injection was omitted. Rejection was observed in both groups during FK506 monotherapy; however, the onset of early rejection episodes was significantly later, the period for reversal of the first rejection was significantly shorter, and the dosage of FK506 at the time of rejection was significantly lower among leukocyte-treated recipients. After completion of immunosuppression, survival was modestly prolonged in the leukocyte-treated group. One animal is surviving without immunosuppression on day 234. This trial of donor leukocyte injection combined with immunosuppression in limb transplantation showed a modest, but significant, improvement in outcome.     

Comparative study of allograft survival of heterotopic and orthotopic small bowel transplantation in rat

BACKGROUND: Small bowel allografts can either be placed heterotopically or orthotopically. In heterotopic small bowel transplantation (H-SBT), the host small intestine containing a substantial amount of gut-associated lymphoid tissue is removed, whereas in conventional orthotopic small bowel transplantation (O-SBT) it is retained. This study compared the allograft survival of H-SBT and O-SBT and evaluated the effect of retaining the host intestine in O-SBT in an altered O-SBT (AO-SBT) model. METHODS: SBT was performed in a high-responder rat stain combination (blood group D Agouti --> Lewis). Immunosuppressive treatment consisting of a short course of FK506 (2 mg/kg/day IM for 3 days before transplantation and 0.3 mg/kg/day for 14 days after transplantation) was used. RESULTS: Survival (mean +/- SD) of H-, O-, and AO-SBT untreated animals was 6.25+/-0.58 days, 6.5+/-0.58 days, and 6.7+/-0.25 days, respectively. With FK506 immunosuppression, survival of H-SBT animals was 49.3+/-13.17 days, whereas 75% (12/16) and 80% (4/5), respectively, of O-SBT and AO-SBT animals achieved indefinite survival (>120 days) with functioning grafts. CONCLUSION: Our data suggest that heterotopic placement of intestinal allografts results in a more severe graft rejection than orthotopic placement. The indefinite survival of O-SBT is not due to of the removal of host intestine carrying a heavy load of gut-associated lymphoid tissue.     

Drug delivery system using microspheres that contain tacrolimus in porcine small bowel transplantation

Rejection remains a major barrier to successful bowel transplantation, in spite of improved immunosuppressive techniques. Therefore, new, more effective, immunosuppressants, with fewer side effects, are needed. Biodegradable microspheres containing tacrolimus (FK506) were used in an experimental porcine small bowel transplantation. Twenty pigs underwent transplantation and were divided into four groups according to the immunosuppressive regimen. Group A (n=5): no immunosuppression; group B (n=6): 0.2 mg/kg per day of FK506; group C (n=3): 1.0 mg/kg per day of FK506; group D (n=6): 0.04 mg/kg per day of FK506 contained in biodegradable microspheres. Rejection was diagnosed macroscopically by endoscopic examination and histologically by biopsy specimen analysis. The mean survival time and standard deviation (SD) were 8.8±3.5, 11.0±1.4, 9.7±2.5 and 28.6±22.5 days for groups A, B, C, and D, respectively, with a statistically significant difference found between group D, on the one hand, and groups A, B and C, on the other. The mean trough blood concentration of FK506 was 10.5±2.2, 27.9±6.0 and 10.5±3.5 ng/ml in groups B, C and D, respectively. In groups A and B, all pigs died of rejection, without infection. In group C, all died of infection, without rejection. In contrast, none of the pigs in group D developed rejection or infection. Our results clearly show that the drug delivery system using biodegradable microspheres that contain FK506 is effective for controlling rejection with fewer side effects in the porcine small bowel transplantation.     

Synergistic effect of vincristine with tacrolimus or sirolimus in prevention of acute heart allograft rejection in the rat

The application of multiple immunosuppressive therapy for organ transplantation could enhance therapeutic efficacy, while minimizing the toxicity of individual drugs used in the regimen. In this study, the effect of the combined therapy of vincristine (VCR) with tacrolimus (FK506) or sirolimus (rapamycin, RAPA) was tested in prevention of acute heart allograft rejection in the rat. A Brown Norway (BN, RT 1(n)) to Lewis (LEW, RT 1(1)) rat combination was used in a heart allografting model. VCR was administered intraperitoneally once daily, while FK506 and RAPA were given by gavage once daily for 14 days after transplantation. There were dose-related prolongations of mean survival time (MST) to monotherapy of VCR, FK506, or RAPA. The MST in combination therapy indicated that a synergistic interaction was produced when compared with the respective monotherapies: VCR 0.05 mg/kg/day + FK506 0.5 mg/kg/day (16.00 +/- 1.79 days, P = 0.001; combination index (CI) = 0.557); VCR 0.05 mg/kg/day + FK506 1.0 mg/kg/day (29.00 +/- 10.54 days, P = 0.001; CI = 0.598); VCR 0.05 mg/kg/day + RAPA 0.2 mg/kg/day (17.33 +/- 1.97 days, P = 0.001; CI = 0.500); and VCR 0.05 mg/kg/day + RAPA 0.4 mg/kg/day (21.17 +/- 3.19 days, P = 0.001; CI = 0.838). Combination therapy of VCR and FK506 or RAPA produced synergistic effects in prevention of acute heart allograft rejection in the rat.     

FK506 as the sole immunosuppressive agent for prolongation of islet allograft survival in the rat.

The purpose of the present study was to determine immunosuppressive effects of a new immunosuppressive agent, FK506, on rat islet allografts and also whether FK is toxic to the islet grafts since the diabetogenic effects of FK is controversial. Hand-picked clean fresh islets (WKA/Qdj:RT1u) were transplanted either beneath the renal capsule or into the liver via the portal vein of the diabetic (STZ, 60 mg/kg) rats (Lewis:RT1(1)). FK506 was administered s.c. for 7 days after transplantation. The mean survival times (MST)* of the renal subcapsular grafts receiving 0 (control), 0.32 or 1.0 mg/kg FK were 7.2 +/- 1.1 (mean +/- SD, n = 5), 13.8 +/- 4.8 (n = 4), and 20.2 +/- 8.0 days (n = 5), respectively. The MST of the intrahepatic grafts receiving 0, 0.1, 0.32, or 1.0 mg/kg FK were 4.4 +/- 1.1 (n = 5), 7.2 +/- 0.8 (n = 5), greater than 45.3 +/- 23.1 (n = 6) or greater than 54.4 +/- 8.8 days (n = 5), respectively. Histologically, islets were found easily in the liver of normoglycemic recipients for more than 60 days after transplantation and appeared intact, with well-granulated beta cells. Foci of mononuclear cells were occasionally seen adjacent to the islet cells. The plasma glucose of the recipients with 1.0 mg/kg FK fluctuated between 150 and 350 mg/dl without rejection. In the recipients treated with 3.2 mg/kg FK the plasma glucose of all the recipients (n = 3) returned to pretransplant levels by 21 days after transplantation. However, islet cells were present in the liver of all these recipients without mononuclear cell infiltration. Immunohistochemically islet grafts stained weakly for insulin, but to the same extent as the controls for glucagon and somatostatin. These findings clearly demonstrate the immunosuppressive effect of FK506 on islet allografts and the importance of the transplant site for prolongation of graft survival by FK, and also suggest that FK has toxic effects on the islet grafts (B cells) when used in high dosages.     

Donor leukocytes combined with delayed immunosupressive drug therapy prolong limb allograft survival

Donor leukocytes administered at the time of transplantation may prolong organ allograft survival. Delayed administration of calcineurin inhibitors, such as FK506 or cyclosporine, may enhance their efficacy. Herein the effectiveness of this strategy to promote limb transplant survival was investigated in the strong histocompatibility barrier of Brown-Norway donor to Lewis recipients. Donor leukocytes (6 x 10(7) intravenously) were injected on the day of transplantation followed on day 1 to 14 with mycophenolate mofetil (MMF; 15 mg/kg/d) and prednisone, (0.5 mg/kg/d) which were then tapered by 20% each week and stopped at week 7. Administration of of FK506 (2 mg/kg/d) was started on day 4 and continued for 8 weeks, then tapered for 4 weeks to a maintenance dose of 0.8 mg/kg/d, which was continued for 12 weeks (group A; n = 8). A control group (n = 8) underwent identical treatment save for donor leukocyte injection but rather commencement of FK506 on day 1. Rejection was common during FK506 tapering in both groups. However group A showed a significantly later onset, a shorter period for reversal of the first rejection, and a significantly lower dosage of FK506 at the time of rejection. After the completion of immunosuppression, rejection occurred significantly later in group A than the control group with one animal surviving without immunosuppression on day 344. This is the first trial of a donor leukocyte injection combined with delayed FK506 administration in limb transplantation, which suggested that it could produce a modest but significant improvement in outcome.     

Recipient FK506 pretreatment regimens in rat small bowel transplantation: allograft survival, function, and systemic infection

PURPOSE: Successful small bowel transplantation requires effective immunosuppression that preserves intestinal function but avoids opportunistic infection. This study aims to evaluate FK506 as a single immunosuppressant in different pretreatment regimens in a rat high responder strain combination. METHODS: Lewis --> DA rat heterotopic small bowel transplantation was performed. Studied groups were (1) untreated control, n = 12; (2) FK-1, n = 8; (3) FK-3, n = 8. FK506 (2 mg/kg/d, intramuscularly) was given to the recipients for 1 day (FK-1) and 3 days (FK-3) before small bowel transplantation, followed by 2 weeks of subtherapeutic treatment (0.3 mg/kg/d, intramuscularly) after small bowel transplantation. Syngeneic small bowel transplantation also was performed (n = 8). FK blood levels, maltose absorption test, histology, and bacteriology were performed at different postoperative days. RESULTS: Allograft survival was prolonged significantly with FK pretreatment, being more so in FK-3 group (FK-1, 22.2 +/- 1.5 d; FK-3, 40.7 +/- 14.1 d; control, 6.6 +/- 0.8 d; P< .01). In the first postoperative week, FK blood level was significantly higher in FK-3 group (19.8 +/- 1.5 ng/mL) than in FK-1 group (5.0 +/- 0.4 ng/mL; P < .05). There was no evidence of systemic infection in either FK-treated group. For maltose absorption, control allograft was abnormal on day 7 correlating to severely damaged intestinal architecture. In contrast, FK-treated allografts showed well-protected intestinal structure and normal absorption on days 7 and 21. CONCLUSION: High FK506 blood levels in the first postoperative week, achieved with FK pretreatment, prolonged intestinal allograft survival and preserved intestinal structure and function without allowing systemic infection.     

FK506和供体特异性输血在大鼠异位心脏移植中的作用

目的探讨FK506和供体特异性输血在大鼠异位心脏移植中的作用。方法利用大鼠异位心脏移植模型以了解在移植当天或移植术后第4日进行供者特异性输注（DST）的基础上,应用FK506能否延长移植物的存活。结果在移植当天行DST或单独用FK5061mg／kg连续10天,可将移植心中位存活时间从对照组的5天分别有效延长至7天和11天,而FK506与DST联合应用时并不产生增强效应。结论FK506和DST单独应用时虽均能延长大鼠同种移植心存活,但是它们没有协同作用。     

Biologics and donor bone marrow cells for targeted immunomodulation in vascularized composite allotransplantation: a translational trial in swine

Introduction

Bone marrow (BM) infusion following organ transplantation is a prerequisite for potential donor-antigen-specific tolerance induction. We developed a preclinical swine model to determine the optimal dose of BM cells to achieve microchimerism. Furthermore, induction therapy was optimized by augmenting the BM infusion with biologics in the form of costimulatory blockade: cytotoxic T-lymphocyte antigen 4 immunoglobulin (CTLA4-Ig).

Materials and Methods

Yucatan miniature swine (n = 12) underwent total body and thymic irradiation for cytodepletion. Animal groups received 15, 30, or 60 million cells per kilogram of whole unmodified BM. The optimal dose of BM cell infusion (BMT) was then applied to subsequent experiments evaluating the addition of CTLA4lg. Group 1 (control) received no treatment. Group 2 received FK506 only; group 3 received irradiation, BMT, and FK506; group 4 received FK506 and CTLA4-lg.

Results

Microchimerism was established in all animals after BM cell infusion; at postoperative day 9, it was significantly increased for 60 million cells per kilogram (P = .0001). Transplanted animals in group 1 rejected the allograft 5 to 8 days after transplantation. Group 2 rejected the allograft (skin and muscle) 30 to 32 days after transplantation (2 days after cessation of immunosuppression). Group 3 rejected the skin portion of the allograft at 50, 52, and 53 days posttransplant. Remaining allograft components (muscle, bone, nerve, vessel) survived indefinitely. Group 4 animals demonstrated significantly prolonged muscle survival beyond 150 days posttransplant; the skin component survived past 150 days in two of three animals. Skin and muscle histology in all long-term surviving animals were normal.

Conclusions

BM cell infusion with 60 million cells per kilogram results in stable levels of microchimerism. The addition of costimulatory blockade (CTLA4lg) prolonged allograft skin survival and overall graft survival. Such targeted immunomodulatory protocols might facilitate immune tolerance and eliminate the need for multidrug immunosuppression to maintain graft survival after vascularized composite allotransplantation.     

Longer survival of rat limb allograft: Combined immunosuppression of FK-506 and 15-deoxyspergualin

We studied the individual and synergistic effect of 3 immunosuppressive drugs, FK-506 (1 mg/kg/day), 15-deoxyspergualin (2.5mg/kg/day) and cyclospo-rine (15 mg/kg/day) in a DA/Lewis rat limb allotransplantation model. 74 right hindlimb transplantations were performed. The median time for onset of rejection was 4 days in animals without immunosuppression, 37 days in animals receiving cyclosporine immunosuppression for 30 days, 61 days in animals receiving FK-506 for 30 days, 36 days in animals receiving a 30-day course of cyclosporine and, in the first 15 days, a course of 15-deoxyspergualin, and 76 days in animals receiving a 30-day course of FK-506 and 15-deoxyspergualin in the first 15 days. The combination of cyclosporine with 15-deoxyspergualin did not prolong graft survival and no synergistic effect was evident. In contrast, survival time in rat limb allografts receiving FK-506 and 15-deoxyspergualin was longer than in those receiving single FK-506 therapy. Our findings suggest a positive synergistic immunosuppressive effect with FK-506 and 15-deoxyspergualin in limb allotransplantation.     

The immunosuppressive effect of FK 506 on canine lung transplantation.

The immunosuppressive potency of FK 506 was studied after left lung transplantation in adult mongrel dogs in comparison with cyclosporine. Fiberoptic bronchoscopy, bronchial mucosal blood flow measurement with laser Doppler velocimetry, and chest x-ray and pathologic examinations were performed. Group A had no immunosuppression (n = 5); group B received FK 506 (0.10 mg/kg/day intramuscularly (n = 5); group C received cyclosporine (20 mg/kg/day orally) (n = 5). In group A four dogs died of rejection on the seventh to the twenty-first postoperative days. Another one was killed on the fourteenth postoperative day because bronchial dehiscence occurred at the anastomosis. In group B one died of alveolar rejection on the seventh postoperative day. The remaining four survived 28 days and were put to death. In group C all five dogs survived 28 days and were put to death. In group A bronchial stenosis or dehiscence at the anastomosis was found in every one during the early postoperative period. In group B stenosis did not develop in any of the dogs, including the one that died on the seventh postoperative day. In group C slight stenosis was seen in one dog, severe narrowing in another, and good healing in the remaining three. The transplanted lungs were almost normal histologically in four animals of group B, and one showed alveolar phase rejection. In all animals of group A severe rejection was observed, and in group C two of five animals showed vascular phase rejection, two latent phase, and one fibrosis. Histologic examination of the bronchial anastomosis in group B showed almost normal bronchial epithelium and slight submucosal infiltration of mononuclear cells. In group A there was desquamation of epithelium and mild to moderate mononuclear cell infiltration. In group C hyperplasia of the epithelium was observed in two animals, an abscess at the site of anastomosis in one, and mild to moderate mononuclear cell infiltration in all five. With use of laser Doppler velocimetry, bronchial blood flow in group B was found to be the same as in group C. Laser Doppler velocimetry values reached preoperative levels by the twenty-eighth postoperative day in both groups. Although diarrhea developed in two dogs of group B, no other significant side effect of FK 506 was seen.