FK 506 reverses acute graft-versus-host disease after allogeneic bone marrow transplantation in rats.

Severe graft-versus-host disease was induced by transplantation of ACI rat bone marrow and spleen cells into irradiated Lewis rat recipients. Treatment with FK 506 or cyclosporine A (CsA) was started after clinical and histologic evidence of acute GVHD was present. A 14-day course of FK 506 at 1.0 mg/kg/day could rescue 100% of the animals suffering from GVHD. In contrast only one half of the animals treated with CsA at a high dose of 25 mg/kg/day recovered. After cessation of immunosuppressive therapy, FK 506-treated animals displayed a marked prolonged disease-free interval as compared to CsA-treated bone marrow recipients. Recurrence of the disease in these animals could be prevented when FK 506 treatment was continued after the induction period with a low maintenance dose of 0.1 mg/kg/day every other day.     

Structural skeletal impairment induced by immunosuppressive therapy in rats: cyclosporine A vs tacrolimus

The exact role of immunosuppressive drugs in the development of osteoporosis and pathologic fractures frequently reported in patients following organ transplantation is still not known. In two experiments, the effects of immunosuppressive drugs were studied on growing rats allocated randomly into five groups of eight rats each which received either FK506 (1.5 mg/kg or 3 mg/kg) or cyclosporine A (15 mg/kg or 30 mg/kg) for 28 days by daily oral gavage. In experiment I ( n=40), bone mineral content (BMC) of the femur by dual energy X-ray absorptiometry (DXA), and bone ash weight were measured. In experiment II ( n=40), stereologic measurements of decalcified tibiae were carried out. The BMC and ash weight values of the whole femur were significantly lower both in the low- and high-dose FK506 groups as well as in the high-dose CsA group. Decalcified sections showed lower volume density of trabecular bone of the tibial metaphysis in both CsA-treated groups and in the high-dose FK506 group. Furthermore, the volume density of the hypertrophic zone volume of the growth plate was higher in high-dose CsA-treated rats. Our data demonstrate that both CsA and FK506 have adverse effects on bone and that high doses of CsA or FK506 alter both cortical and trabecular bone with subsequent osteopenia. In addition, CsA-treated groups showed histological changes in some aspect resembling rickets/osteomalacia.     

Lesser reduction in bone mineral density by the immunosuppressant, FK506, compared with cyclosporine in rats

BACKGROUND: Posttransplantation osteopenia leading to osteoporosis induced commonly by treatment with immunosuppressants including cyclosporine (CsA) is a severe complication and results in lowering the quality of life in patients receiving organ transplantation. FK506 is a newly developed immunosuppressant and is currently being used for the prevention of rejection after organ transplantation. In this study, to investigate whether FK506 as well as CsA would cause osteopenia or not, we evaluated the effect of FK506 on bone mineral density and several parameters relevant to bone metabolism in comparison with that of CsA using normal rats. METHODS: Ten-week-old male Sprague-Dawley rats were treated with FK506 (vehicle, 1 mg/kg, and 3.2 mg/kg) or CsA (vehicle, 10 mg/kg, and 32 mg/kg) by daily oral gavage for 28 days. Bone mineral density of the femur, plasma insulin-like growth factor I (IGF-I), and urinary deoxypyridinoline were determined by peripheral quantitative computerized tomography, radioimmunoassay, and enzyme-linked immunosorbent assay, respectively. RESULTS: The reduction in bone mineral density of the femur was observed in both FK506- and CsA-treated rats. The reduction in CsA-treated rats, however, was statistically significant and strikingly severe, whereas that in FK506-treated rats was much less severe than CsA. Plasma IGF-I levels were significantly elevated in FK506-treated rats but not in CsAtreated rats. Urinary deoxypyridinoline levels were unchanged in FK506-treated rats but elevated in CsA-treated rats. CONCLUSIONS: Compared with CsA, FK506 does not appear to induce severe osteopenia by high-turnover bone metabolism in the rat by mediating via IGF-I induction in part. The results suggest that FK506 may exert favorable effects on bone metabolism in patients with organ transplantation compared with CsA. To assess this idea, further clinical investigations focused on bone metabolism will be required.     

Use of mixed lymphocyte reaction to identify subimmunosuppressive FK-506 levels in mice

The immunosuppressive agent FK-506 has a well-described neuroregenerative effect that is mediated by a mechanism independent of calcineurin inhibition. FK-506 levels that fall below the threshold for immunosuppression could therefore potentially enhance nerve regeneration while minimizing toxicity. The purpose of this study was to characterize the dose-dependent effects of FK506 on T-cell proliferation, and establish a subimmunosuppressive dosing regimen for FK-506 in mice. Forty BALB/cJ mice were randomized to four groups corresponding to 0, 0.25, 0.5, or 1.0 mg/kg/day doses of FK-506. Ten days postoperatively, animals were sacrificed, and mixed lymphocyte reaction assays were performed to quantify the immune response to nerve allografts. Mice receiving 0.25 and 0.5 mg/kg/day of FK-506 exhibited a robust T-cell proliferation response, with stimulation indices approaching those of untreated animals. Mice treated with 1.0 mg/kg/day of FK-506 demonstrated significantly decreased T-cell proliferation. These results establish 0.5 mg/kg/day as an upper limit for subimmunosuppressive FK-506 administration.     

Allogeneic hepatocyte transplantation using fk 506

Hepatocyte transplantation is an intriguing alternative to orthotopic liver transplantation. While engraftment of syngeneic hepatocytes can be achieved with relative ease, engraftment of allogeneic hepatocytes has been far more complicated. We used FK 506 (Tacrolimus), a novel and highly efficient immunosuppressant, which has been reported to augment liver regeneration in rats. Recipients of isolated syngeneic (LEW) and allogeneic (Wistar F.) rat hepatocytes (major histocompatibility barrier) recieved different immunosuppressive regiments with FK 506 or Cyclosporine A (CsA). Mature syngeneic hepatocytes could be retrieved up to post op day 300 with the lowest number of hepatocytes on post op day 20. Following allogeneic transplantation, no mature hepatocytes could be identified after post op day 10, though ductular like structures within the spleen were found in FK 506 but not CsA-treated animals. The epithelial cells of ductular like structures exhibit cytological features of CK-19 positive cells. Our results suggest that under CsA or FK 506 immunosuppression long-term survival of mature allogeneic hepatocytes within the spleen cannot be achieved across a major histocompatibility barrier though FK 506 allows engraftment of allogeneic donor type ductular cells.     

Efficacy of rapamycin and FK 506 in prolonging rat hind limb allograft survival.

OBJECTIVE: Graft rejection and the toxicity of current immunosuppressive regimens preclude the application of microsurgical advances to transplantation of limbs or other nonessential parts. If limb transplantation is to become a clinical reality, newer, safer, more effective immunosuppressive agents are needed. SUMMARY BACKGROUND DATA: Rapamycin (RPM) and FK 506 are fungal macrolide antibiotics with effective immunosuppressive properties demonstrated in several animal models. RPM is more potent and effective than is FK 506 in rat cardiac allografts and has demonstrated synergy with cyclosporine (CsA) in limb allograft models. METHODS: An orthotopic rat hind limb allograft model (Brown-Norway [RT-1n] to Lewis [RT-1(1)] rats was used. RPM (doses, 3.0, 4.5, and 6.0 mg/kg/day) was administered intraperitoneally on postoperative days 1 to 14. FK 506 (6 mg/kg/day) was administered orally on postoperative 1 to 14 and 1 to 90 and at rejection onset (10 mg/kg/day for salvage). CsA with RPM (postoperative days 1 to 14) was used to assess synergy, with CsA alone serving as the control. Other controls included untreated and placebo-treated allografted animals. The permutation test and Mann-Whitney test were applied to the data. RESULTS: The mean survival times were assessed as follows: (1) control (placebo, untreated), 5 days; (2) RPM groups, 9.5, 10.6, and 8.7 days; (3) 14-day FK 506, 28 days; (4) 90-day FK 506, > 90 days; (5) CsA, 17.3 days; and (6) CsA with RPM, 19.3 days. FK 506 significantly prolonged graft survival compared with RPM (Permutation Test, p < 0.001 and Mann-Whitney Test, p < 0.05). FK 506 salvage reversed early rejection. High-dose RPM produced significant toxicity. Synergy between CsA and RPM was not demonstrated. CONCLUSIONS: FK 506 prolongs allograft survival, reverses early rejection, and prevents rejection without clinical toxicity when given continually. RPM does not prevent rejection in this model and produces significant toxicity at high doses. FK 506 may be a first step in making limb transplantation a clinical reality in reconstructive surgery.     

环孢素A及FK506对人肝癌细胞株增殖的影响

目的:探讨免疫抑制剂环孢素(CsA)及他克莫司(FK506)对肝癌细胞增殖的影响及可能机制。方法:采用噻唑蓝(MTT)比色法观察CsA及FK506对QGY鄄7701、QGY鄄7703、SMMC鄄7721和BEL鄄7402等4株肝癌细胞增殖的影响。还应用Hoechst33258荧光染料涂片,在荧光显微镜下观察用药组细胞的形态学改变。结果:随着作用时间延长,10μM的CsA对4株肝癌细胞均呈现明显的生长抑制效应(P<0.01),而0.5μM的FK506与对照组相比,其生长抑制作用不明显(P>0.05)。荧光显微镜下CsA用药组肝癌细胞呈现凋亡改变;而FK506用药组与对照组无显著差异。结论:免疫抑制剂CsA非但未促进体外肝癌细胞生长,相反呈明显的抑制效应。FK506尽管未呈现对肝癌细胞增殖的抑制效应,也未显示有生长促进作用。CsA对肝癌细胞的增殖抑制作用与诱导癌细胞发生凋亡有关。     

Accommodation after lung xenografting from hamster to rat

BACKGROUND: Long-term xenograft survival can be achieved in hamster hearts transplanted into rats treated with cobra venom factor (CVF) and cyclosporine A (CsA). This phenomenon of "accommodation" is associated with expression of protective genes such as bcl-2, bcl-X(L), and heme-oxygenase-1. We examined whether accommodation could be induced in hamster-to-rat lung xenografts and whether the pattern of protective genes is similar to cardiac xenografts. METHODS: We used hamster-to-rat cardiac and lung xenotransplantation models. Cardiac xenotransplants were treated with CVF+CsA and compared with untreated controls. Lung xenotransplants were treated with either CVF+CsA or FK506 and cyclophosphamide (Cp) and compared with untreated controls. All recipients were killed by 21 days after transplantation. We examined graft survival and protein expression of protective genes, and we performed histologic and immunohistologic analyses. RESULTS: Rejection occurred rapidly in untreated rats. CVF+CsA or FK506+Cp treatment significantly influenced graft survival. Eight of 12 CVF+CsA-treated heart transplants survived 21 days. Seven of 16 CVF+CsA-treated lung grafts and five of 12 FK506+Cp-treated lung xenografts survived 21 days. We observed significant protein expression of bcl-2, bcl-X(L), and heme-oxygenase-1 in cardiac xenografts treated with CVF+CsA at 2, 14, and 21 days after transplantation, compared with normal hamster hearts. We also observed significant expression of these proteins in lung xenografts treated with either CVF+CsA or FK506+Cp at 21 days after transplantation, compared with normal lungs. CONCLUSIONS: Accommodation may be a general phenomenon for all organs, mediated through protective genes. Induction of accommodation does not require disruption of the complement system.     

FK506--its influence on anti-class 1 MHC alloantibody responses to blood transfusions.

The influence of FK506 on in vivo alloantibody responses to major histocompatability class 1 antigens was investigated in inbred rat strains, and compared with the effect of cyclosporine. AO rats received transfusions of DA blood on days 0 and 7. From days 0 to 14 the rats also received, daily, either FK506 0.3 mg/kg suspended in saline or dissolved in olive oil, or CsA 10 mg/kg. The administration of FK506 suspended in saline at the time of blood transfusion completely abrogated the development of anti-MHC class 1 alloantibodies as detected by indirect hemagglutination (IHA)* and 51Cr release complement dependent cytotoxicity assays (CDC). Isotyping studies showed that FK506 suspended in saline suppressed IgM production and inhibited the switch to IgG production. Similar responses were seen in CsA-treated animals. In contrast, rats treated with FK506 dissolved in olive oil developed high titers of anti-class 1 alloantibodies. On days 49 and 56 the rats were challenged with further DA blood transfusions given without immunosuppression. In the groups given FK506 suspended in saline or CsA, cytotoxic antibodies did not develop; low titer antibodies were, however, detected by IHA in the animals that had previously received FK506 suspended in saline. The results indicate that FK506, in common with CsA, inhibits anti-class 1 MHC alloantibody production, and at the same time enables the development of tolerance. The vehicle in which FK506 is administered is, however, critical to its efficacy at the low doses used. These results may be of relevance to clinical transplantation as similar antibodies mediate hyperacute renal allograft rejection in man.     

Gemcitabine with cyclosporine or with tacrolimus exerts a synergistic effect and induces tolerance in the rat

BACKGROUND: This study investigated the effect of the antineoplastic agent gemcitabine (dFdC) in combination with cyclosporine (CsA) or with FK506 on acute heart allograft rejection in a rat model. METHODS: Transplantations were performed in the fully allogeneic Lewis-to-Brown Norway strain combination. dFdC, CsA, and FK506 single-drug therapy and combinations of dFdC with CsA and FK506 were administered at various dosages starting on day 1 to prevent and on day 4 to treat acute rejection until day 20. Animals who did not reject their graft were intraperitoneally injected with 108 splenic donor-type lymphocytes. In addition, Lewis and third-party skin grafts were transplanted to these animals. RESULTS: Mean graft survival times under CsA, FK506, and dFdC monotherapy were 18.3/63.7 days (1 mg/5 mg per kg), 41.7 days, and 24.7/38.7 days (100 microg/150 microg per kg), respectively. CsA and FK506 in combination with dFdC prolonged graft survival to more than 100 days (CsA) and more than 95.2 days (FK506). Graft survival after treatment of an ongoing rejection was 21.5/38.3 days for CsA (1 mg/5 mg per kg) and 17.7/59.2 days for dFdC (100 microg/150 microg per kg). The combination of CsA+dFdC prompted indefinite survival of five of six hearts. Lymphocyte inoculation did not induce graft rejection. Notably, none of the Lewis, but all third-party, skin grafts were rejected immediately. Histomorphologic analysis of grafted hearts, however, demonstrated typical features of chronic rejection. CONCLUSIONS: The combination of CsA and FK506 with low-dose dFdC exerts a synergistic effect in the prevention and treatment of acute allograft rejection in this model. Although chronic rejection could not be prevented, strain-specific tolerance was achieved. Therefore, combining standard immunosuppressants with dFdC is a novel, promising strategy for prevention and treatment of acute allograft rejection.     

Nephrotoxicity after orthotopic liver transplantation in cyclosporin A and FK 506-treated patients

Abstract Nephrotoxicity represents a serious side-effect of immunosuppression following orthotopic liver transplantation. In order to preserve the therapeutic potential of cyclosporin (CsA) and FK 506 in human liver transplantation and to differentiate the nephrotoxic action of either drug in a clinical setting, we evaluated the incidence of early and late nephrotoxicity in 121 patients, 60 randomly assigned to CsA- and 61 to FK 506-based immunosuppression. Early postoperative renal insufficiency (between PODO and 30; SCr 1.5-3 mg/dl) was observed to a similar extent in patients treated with CsA (36.7%) and FK 506 (42.6%). Early postoperative acute renal failure (ARF; SCr > 3 mg/dl) occurred in 18.3%, regardless of the immunosuppressive management. Approximately 50% of patients with ARF required hemodialysis (CsA: 11.7%; and FK 506: 8.3%). Mean onset of hemodialysis in CsA-treated patients was POD1 and in FK 506-treated patients, POD 6, which demonstrated a different time course of drug-pecific nephrotoxicity of CsA and FK 506 in early ARF. All patients with early postoperative ARF requiring hemodialysis survived more than 1 year (100% survival). New onset of late ARF (between POD 30 and 365), however, occurred in 6.5% under FK 506 and in 1.7% under CsA immunosuppression due to severe infections with the multiple organ failure syndrome. This observation was consistent with the assumption of overimmunosuppression rather than a primary nephrotoxic effect. Mortality of patients with late ARF requiring hemodialysis was 100%. Late renal insufficiency appeared in 23.3% of CsA- and in 29.4% of FK 506-treated patients, and represented a slowly progressing form of drug-pecific nephrotoxicity. These preliminary results demonstrated a similar outcome in terms of early and late nephrotoxicity, but longer follow-up will delineate its overall efficacy and toxicity in humans.     

Kidney function and morphology after short-term combination therapy with cyclosporine A, tacrolimus and sirolimus in the rat.

BACKGROUND: Sirolimus (SRL) may supplement calcineurin inhibitors in clinical organ transplantation. These are nephrotoxic, but SRL seems to act differently displaying only minor nephrotoxic effects, although this question is still open. In a number of treatment protocols where SRL was combined with a calcineurin inhibitor indications of a synergistic nephrotoxic effect were described. The aim of this study was to examine further the renal function, including morphological analysis of the kidneys of male Sprague-Dawley rats treated with either cyclosporine A (CsA), tacrolimus (FK506) or SRL as monotherapies or in different combinations. METHODS: For a period of 2 weeks, CsA 15 mg/kg/day (given orally), FK506 3.0 mg/kg/day (given orally) or SRL 0.4 mg/kg/day (given intraperitoneally) was administered once a day as these doses have earlier been found to achieve a significant immunosuppressive effect in Sprague-Dawley rats. In the 'conscious catheterized rat' model, the glomerular filtration rate (GFR) was measured as the clearance of Cr(EDTA). The morphological analysis of the kidneys included a semi-quantitative scoring system analysing the degree of striped fibrosis, subcapsular fibrosis and the number of basophilic tubules, plus an additional stereological analysis of the total grade of fibrosis in the cortex stained with Sirius Red. RESULTS: CsA, FK506 and SRL all significantly decreased the GFR. A further deterioration was seen when CsA was combined with either FK506 or SRL, whereas the GFR remained unchanged in the group treated with FK506 plus SRL when compared with treatment with any of the single substances. The morphological changes presented a similar pattern. The semi-quantitative scoring was significantly worst in the group treated with CsA plus SRL (P<0.001 compared with controls) and the analysis of the total grade of fibrosis also showed the highest proportion in the same group and was significantly different from controls (P<0.02). The FK506 plus SRL combination showed only a marginally higher degree of fibrosis as compared with controls (P=0.05). CONCLUSION: This rat study demonstrated a synergistic nephrotoxic effect of CsA plus SRL, whereas FK506 plus SRL was better tolerated.     

Superior T-cell suppression by rapamycin and FK506 over rapamycin and cyclosporine A because of abrogated cytotoxic T-lymphocyte induction,impaired memory responses,and persistent apoptosis

Immunosuppressive therapy is best achieved with a combination of agents targeting multiple activation steps of T cells. In transplantation, cyclosporine A (CsA) or tacrolimus (FK506) are successfully combined with rapamycin (Rap). Rap and CsA were first considered for combination therapy because FK506 and Rap target the same intracellular protein and thus may act in an antagonistic way. However, in clinical studies, FK506+Rap proved to be effective. To date, there is no in vitro data supporting these in vivo findings, and it is unclear whether the observed effects are T-cell mediated. In a human polyclonal allogeneic in vitro model, we found that although combined drug treatment markedly reduced expansion of naive T cells, T-cell activation occurred irrespective of the drug combination used. The induction of cytotoxic effector T cells was reduced by CsA+Rap but completely abolished by FK506+Rap. Importantly, combined immunosuppression allowed generation of memory CD4+ and CD8+ T cells and hence did not result in T-cell anergy. However, FK506+Rap treatment resulted in a reduced number of allospecific memory T cells showing a decreased cell-cycle turnover and cytokine producing capacity. In contrast, CsA+Rap treatment led to increased memory T-cell numbers responding with elevated kinetics. The ability of Rap to promote apoptosis, which contributes to T-cell suppression, remained unaffected upon combination with FK506 or CsA. These data support the combined use of FK506+Rap over CsA+Rap for immunosuppressive therapy.     

Evidence that the immunosuppressive effects of FK506 and cyclosporine are identical.

The fungal metabolite FK506 was discovered because it shared an important property, the ability to inhibit production of IL-2, with another well-known immunosuppressive fungal metabolite, CsA. FK506 has, since its isolation, also been shown to share other immunosuppressive effects with CsA. This study was performed to further investigate the in vitro immunological properties of FK506, in comparison with and in combination with CsA, to evaluate the plausibility that their mechanisms of action were identical or similar, in spite of their different molecular structures. The ability to inhibit several responses of human peripheral blood lymphocytes to mitogenic and alloantigenic stimulation was explored. Synergistic effects of the two drugs were extensively studied, since this would provide additional information regarding their mechanisms of action. Also, similar immunosuppressive properties would enable the use of the two drugs in combination. We found that FK506 and CsA had very similar mechanisms of action and additive effects were recorded.     

The effects of maintenance doses of FK506 versus cyclosporin A on glucose and lipid metabolism after orthotopic liver transplantation

BACKGROUND: Posttransplant diabetes mellitus (PTDM) has gained widespread attention due to the micro and macro-vascular complications that increase the morbidity and mortality of patients receiving solid organs. The higher incidence of PTDM has been mainly attributed to the immunosuppressive therapy. Therefore, this study compares the metabolic side effects of low dose maintenance therapy of FK-506 and Cyclosporin A (CsA) in 14 patients 1 year after orthotopic liver transplant and analyzes possible factors that contribute to the development of PTDM. METHODS: Two groups (n=7) differing in their immunosuppressive regimen (FK506 or CsA) were matched to eight control subjects and compared to each other. The effects of in vivo insulin action were assessed by means of the euglycemic hyperinsulinemic clamp technique. Arginine stimulation tests at normo- (5.5 mM) and hyperglycemic (15 mM) levels were performed and the acute insulin, C-peptide, and glucagon response (2-5 min) to arginine were determined. RESULTS: Insulin sensitivity (total glucose disposal) was statistically lower in patients treated with FK-506 and CsA (5.05+/-0.47 and 5.05+/-0.42 mg/kg/min) as compared to controls (6.62+/-0.38 mg/kg/min) (P<0.02), with a significantly higher nonoxidative glucose disposal for the control group (P<0.01), and lower free fatty acid levels (P<0.05). Absolute values for acute insulin response were higher but not significantly different for the transplanted groups. The lower percentage of increase of insulin release after arginine stimulation observed in the FK-506 and CsA groups as compared with controls (754%+/-100, 644%+/-102 vs. 1191%+/-174) (P<0.03 and 0.02, respectively), suggests a reduced beta cell secretory reserve in both treated groups. Also, the acute glucagon response to arginine during hyperglycemia declined less in the FK-506 (28%) and CsA groups (29%) compared with controls (48%) (P<0.05) indicating a defect in the pancreatic beta cell-alpha cell axis. CONCLUSIONS: There are no major metabolic differences on low maintenance doses between FK-506 and CsA. Both immunosuppressant agents contribute to the development of PTDM at different levels.     

Effects of FK506 on aminonucleoside-induced nephrotic rats]

We examined the effect of immunosuppressive agent, FK506 (Fujisawa, Co.), on puromycin aminonucleoside (PAN)-induced nephrosis. Single i.p. injection of PAN in a dose of 100 mg/kg was introduced into Munich-Wistar rats weighing about 200 g. Those rats were divided into four groups. PAN-induced nephrosis rats in group 1 (PAN-FK0.1, n = 5), group 2 (PAN-FK0.3, n = 5), group 3 (PAN-FK1.0, n = 5) were treated with i.m. injection of FK506 for 10 days in a dose of 0.1 mg/kg, 0.3 mg/kg, 1.0 mg/kg, respectively, and rats in group 4 were treated with FK-placebo (PAN-PL, n = 5). The rats in group 5 with Saline+placebo were served as a control (NS-PL, n = 5). Among 5 groups, urinary protein, anionic sites (AS) in GBM, and subsets of peripheral lymphocytes through FACS were compared. After 9 days of PAN injection, the rats in PAN-FK0.1 (160.0 +/- 38.4), PAN-FK0.3 (118.0 +/- 34.4) & PAN-FK1.0 (89.2 +/- 40.0) given FK-506 had significantly less proteinuria in a PAN dose dependent manner, compared to those in NS-PL (349 +/- 86.8 mg/day). The numbers of AS/1000 mmGBM were more attenuated in FK506-treated PAN rats (PAN-FK1.0; 16.2 +/- 3.9) than those in PAN-PL (11.7 +/- 4.4). In related to subset of lymphocytes, increased W3/25 in PAN-PL was regressed in PAN-FK0.1, PAN-FK0.3 & PAN-1.0 after 10 days of PAN-injection. W3/25/OX-8 was significantly higher in PAN-PL (3.6) than those in NS-PL (2.4), but not between PAN-1.0 & NS-PL. These data indicate that the mechanism for therapeutic effect of FK506 on PAN-induced nephrosis includes a revision of abnormal cellular immunity, which attenuates the decrease of AS structure and as a result decrease proteinuria.     

Strategies to develop chimerism in vascularized skin allografts across MHC barrier

In this study, we investigated the effects of 7-day-protocols of alphabeta-T-cell receptor monoclonal antibody (alphabeta-TCRmAb), cyclosporine A (CsA), and tacrolimus (FK-506) immunosuppressive monotherapies, and their combinations on the survival of vascularized skin allografts (VSA). Forty-two transplantations of VSA across a strong MHC barrier were performed between ACI (RT1a) donors and Lewis (RT1(l)) recipients in seven groups. Isograft and allograft rejection controls received no treatment. Treatment groups received a 7-day protocol of alphabeta-TCRmAb, CsA, or FK-506 monotherapy, or a combination of alphabeta-TCRmAb/CsA and alphabeta-TCRmAb/FK-506. VSA transplants were evaluated on a daily basis. Donor-specific chimerism was determined by flow cytometry (FC). The combined protocols of alphabeta-TCRmAb/FK-506 and alphabeta-TCRmAb/CsA significantly prolonged VSA survivals compared to monotherapy groups ( P < 0.005). FC analysis revealed 15.82% of donor-specific chimerism on day 7 under the alphabeta-TCRmAb/CsA protocol and a gradual chimerism decline on day 63 posttransplant. The significant extension of VSA survival achieved under 7-day protocols of combined therapies was directly associated with the presence of donor-specific chimerism.     

Pharmacologic preconditioning of random-pattern skin flap in rats using local cyclosporine and FK-506: interaction with nitric oxide system

It has been suggested that immunophilin ligands such as cyclosporine and FK-506 (tacrolimus) affect the survival of ischemic tissues. Our objective was to show an acute effect of local cyclosporin-A (CsA) and FK-506 on ischemic protection in a random-pattern skin-flap model in rats and investigate the effect of nitric oxide (NO) pathways as a modulator of protection of these agents.Ninety male Sprague-Dawley rats were randomly assigned to treatment groups. Bipedicled dorsal flaps (2 x 8 cm) were elevated at midline. Prior to cutting the cranial pedicle to induce permanent ischemia, pharmacologic preconditioning groups received local injection of CsA (0.3, 1, or 3 nmol/flap) or FK-506 (0.01, 0.03, or 0.1 pmol/flap), and the ischemic preconditioning (IPC) group underwent temporary clamping of the cranial pedicle. At the seventh day postoperatively, the survival of the flaps was measured. In other groups, nitric oxide synthase inhibitor N omega-nitro-l-arginine methyl ester hydrochloride (L-NAME) was administered with effective CsA and FK-506, and ischemic preconditioning. Nitric oxide precursor L-arginine doses were also studied, and a systemic subeffective dose (100 mg/kg) was coadministered with subeffective CsA and FK-506.Significant increase in flap survival was obtained with CsA (1 nmol/flap), FK-506 (0.1 pmol/flap), and IPC. These protections were abolished by systemic administration of L-NAME (10 mg/kg). Coadministration of subeffective doses of CsA (0.3 nmol/flap) and FK-506 (0.03 pmol/flap), with subeffective systemic l-arginine, significantly improved flap survival.Pharmacologic preconditioning with local, single, low doses of CsA or FK-506 is shown to be even more effective than IPC. Administration of the NOS substrate l-arginine potentiates these effects.     

Differential Effect of Diarrhea on FK506 Versus Cyclosporine A Trough Levels and Resultant Prevention of Allograft Rejection in Renal Transplant Recipients

Diarrhea is the most frequently reported adverse event in patients treated with mycophenolate mofetil. Twenty-six renal transplant patients on a mycophenolate mofetil-based immunosuppressive regime with persistent afebrile diarrhea were examined. Diarrhea caused a significant rise in FK-506 trough levels despite intake of stable doses, necessitating FK-506 dose reductions of 30% to obtain pre-diarrhea trough levels. In contrast, trough levels of cyclosporine A remained stable without dose adjustments. This suggests that absorption and/or metabolism is differentially altered for FK506 compared with cyclosporine A in patients with diarrhea. In nine patients mycophenolate mofetil was reduced or stopped because of persistent diarrhea without identifiable cause. This resulted in end-stage renal disease because of chronic rejection in two patients, and in acute rejection in two patients, all taking FK506 and steroids. Therefore, dose adjustments of FK506 in patients with diarrhea must be carefully monitored, especially when doses of mycophenolate mofetil are also reduced.     

Soluble antigen and cyclosporine-induced specific unresponsiveness in rats. Frequency of alloantigen-specific T cytotoxic cells in normal, sensitized, and unresponsive rats

The cellular mechanisms of unresponsiveness induced with a combined KCl-extracted donor antigen (HAg) and CsA regimen were dissected by limiting-dilution (LD) assay. While untreated Wistar-Furth (WFu, RT1u) rats reject Buffalo (BUF, RT1b) heart allografts within a mean survival time of 6.6 +/- 0.5 days, recipients treated with 3 cycles of CsA alone (-1,0,1; 7,8,9; 15,16,17) maintained BUF heart allografts up to an MST of 22.5 +/- 8.9 days. When CsA was combined with BUF HAg (-1), BUF heart survival was further prolonged up to an MST of 34.2 +/- 6.6 days, while third-party BN HAg was ineffective (MST of 21.5 +/- 2.1 days). On day 10 postgrafting, the frequency of T cytotoxic cells (fTc) within the splenic pan-T-cell population was 1:1437 +/- 301 in CsA and 1:1087 +/- 438 in CsA/HAg treated recipients. In contrast, on day 30 postgrafting, both CsA and CsA/HAg treated WFu rats bearing functional BUF hearts showed within their splenic pan-T-cell populations a profound decrease in fTc to 1:2966 +/- 824 with CsA alone and to 1:4946 +/- 938 with CsA/HAg treatment. In contrast, both untreated WFu rats who rejected BUF heart allografts and CsA-treated WFu recipients who had rejected their BUF heart allografts on day 20 displayed an increased fTc to 696 +/- 243 and to 1:1169, respectively, when examined at day 30 postgrafting. Additionally, both the W3/25+ and OX8+ T cell subsets specifically suppressed the proliferative response of normal WFu T cells against BUF and, to a lesser degree, third-party BN irradiated stimulators. Thus, CsA-treated animals develop a potent specific-suppressor mechanism that is augmented by pretreatment with donor soluble antigen. This suppressor activity may decrease the frequency of alloantigen-specific Tc cells and thereby prolong the survival of BUF heart allografts.