FK506 Group Research Articles

Promotion of Nerve Regeneration in Peripheral Nerve by Short-Course FK506 after End-to-Side Neurorrhaphy

To discuss the feasibility of peripheral nerve injury treated by end-to-side neurorrhaphy in clinic and to evaluate the effect of short-course FK506 on promoting nerve regeneration after end-to-side neurorrhaphy. Thirty adult male Sprague Dawley rats were randomly divided into 3 groups: Group A, 10 rats received end-to-end anastomosis; Group B, 10 rats received end-to-side neurorrhaphy; Group C, 10 rats received the same operation as Group B. After operation, rats in Groups B and C received muscle injection with saline water (1 mg/kg x d(-1)) and FK506 (1 mg/kg x d(-1)), respectively, both for 4 wk. Histological and morphological examinations were performed 12 wk after the operation. In the 2nd, 4th, 6th, 8th, and 12th wk after operation, function recovery analysis was performed. The results of histological and immunochemistry study (the total number of Schwann cells and the axon numbers at the distal stump of the peroneal nerve, wet weight of extensor digitorum longus muscle) suggested that there were significant differences between Group B (saline water group) and Group C (FK506 group) (P < 0.05), also between Group A (end-to-end group) and Group C (P < 0.05). There were statistically significant differences in function recovery (peroneal functional index and sciatic functional index) between Groups B and C (P < 0.05) and also between Groups A and C (P < 0.05). End-to-side repair combined with FK506 has a potential for application in selected cases of peripheral nerve injury in clinic.

Skin Allograft Survival and Analysis of Renal Parameters After FTY720 + Tacrolimus Treatment in Mice

Calcineurin inhibitors such as cyclosporine (CsA) and tacrolimus (FK506) show similar efficacy to prevent rejection within the first year after organ transplantation. However, their use is limited by side effects, such as kidney damage, hypertension, new-onset diabetes, and hyperlipidemia. The consensus opinion suggests that compared with CsA, FK506 has fewer negative effects on blood pressure, serum lipids, and renal function. Nevertheless, FK506 use is associated with a higher incidence of posttransplantation diabetes mellitus. FTY720 is a new compound that has shown beneficial effects in animal models of rejection in transplantation, ischemia/reperfusion injury, autoimmune diseases, and tumor development. Our aim was to investigate whether FTY720 + tacrolimus association could provide additional immunosuppression without causing renal toxicity. FTY720 as a monotherapy or in association with FK506 was administered to C57BL/6 mice for 21 days to prevent skin graft rejection and to evaluate renal function and structure. Increased skin allograft survival in the FTY720 + FK506 group was associated with decreased cell numbers in the spleen, blood, and axillary lymph nodes. Changes in major histocompatibility complex (MHC) class II and intercellular adhesion molecule-1 (ICAM-1) expressions in splenocytes were also found in this group. The major effects already described for FK506 (diabetes) or FTY720 (lymphopenia) were observed after 21 days administration even when the drugs were associated. FTY720 associated with FK506 caused fewer changes in kidney structure, and blood glucose levels were lower than in FK506 monotherapy.

The Role of Graft and Host Accommodation in a Hamster-to-Rat Cardiac Transplantation Model

We evaluated the importance and mechanism of graft and host accommodation in hamster-to-rat cardiac xenotransplantation models. To evaluate graft accommodation, accommodated hamster grafts (Group 2) were transplanted to naïve host rats treated with FK506, and compared with naïve hamster grafts (Group 1). To evaluate host accommodation, three groups were evaluated: naive hamster hearts were transplanted to naïve hosts treated with FK506 (Group 3: 0.5 mg/kg, Group 4: 1.0 mg/kg) and splenectomy, and compared with accommodating hosts (Group 5) with FK506 0.5 mg/kg and splenectomy. We examined graft survival, histopathology, antihamster antibodies and B-1 cells in blood. Graft survival in Group 2 (3.4+/-0.9 days) was not significantly different from that in Group 1 (2.8+/-0.4 days). Graft survival in Groups 4 and 5 (>30 days) was significantly prolonged compared with that in Group 3 (6.0+/-0.7 days). Histopathology of Groups 1-3 showed humoral rejection, whereas Groups 4 and 5 showed normal histology and expression of protective genes. In Groups 1-3, antihamster immunoglobulin (Ig) M and B-1 cells increased significantly compared to Groups 4 and 5, where IgM and B-1 cells remained low or were reduced. Host accommodation was more important than graft accommodation. Accommodating grafts expressing protective genes were rejected with an elevation of both IgM and B-1 cells. In accommodated hosts, both IgM and B-1 cells decreased, suggesting that B-1 cells may be responsible for the production of antihamster antibodies. These results suggest that sufficient suppression of B-1 cells, resulting in decreased titers of antihamster antibodies, may play an important role in host accommodation.

Impact of Calcineurin Inhibitors on Bone Metabolism in Primary Kidney Transplant Patients

Objective Posttransplant bone disease and bone metabolism markers were investigated in primary kidney transplant recipients receiving calcineurin inhibitor (CNI) based triple immunosuppression. We examined the safety profile and independent potential of CNIs on bone formation and bone resorption. The study also attempted to correct for modifiable and nonmodifiable factors that impact on posttransplantation bone metabolism, such as age, renal function, rejection, steroid dosage, and secondary hyperparathyroidism. Materials and methods Serum alkaline phosphatase and osteocalcin were used as indices of bone formation and urinary deoxypyridinoline as a marker for bone resorption. Bone mineral density (BMD) data were assessed in all patients. Osteocalcin and deoxypyridinoline data were correlated with BMD scores to predict the clinical utility and sensitivity of these tests. Sixty-six patients among 300 kidney transplant recipients were enrolled as eligible candidates based upon more than 12 months’ posttransplantation follow-up excellent graft function (GFR values >60 mL/min), and intact parathormone levels <100 pg/mL. Results Mean follow-up was 1395.3 ± 179.3 days and 1488.9 ± 225.1 days for cyclosporine (CsA) and FK506 groups, respectively. Mean values for alkaline phosphatase and osteocalcin were 108.8 ± 6.0 versus 98.4 ± 9.7 U/L and 10.1 ± 1.2 versus 9.8 ± 1.5 ng/mL for the CsA and FK506 groups, respectively. Both CsA and FK506 caused mild osteoblastic proliferation and matrix mineralization activity, as reflected by increased osteocalcin and alkaline phosphatase levels in 22.6% and 12.5% of patients, respectively. This bone formation activity was counterbalanced by a three-fold increase in urine deoxypyridinoline levels in both groups. Mean deoxypyridinoline levels were, respectively, 13.8 ± 4.4 versus 11.3 ± 2.1 nM/mMCr in the CsA and FK506 groups. Thirty-four (68%) patients in the CsA and 10 (62.5%) in the FK506 groups had elevated deoxypyridinoline levels. A strong correlation existed between deoxypyridinoline levels and BMD scores for the CsA group ( P < .0001). Despite the presence of relatively greater elevations in deoxypyridinoline and BMD values among CsA-treated patients, there was no significant difference in terms of bone resorption potential of both groups. No correlation existed between iPTH values (<65 pg/mL or among 65 to 98.2 pg/mL) at any time versus osteocalcin, alkaline phosphatase, deoxypyridinoline, or BMD levels. The symptomatic bone disease and fracture rates were 0% in this series. Conclusion Calcineurin inhibitor-based immunosuppression with low maintenance doses of glucocorticoids induces slight bone formation but relatively potent, clinically relevant bone resorption.

Clinical Outcome by Acute Graft-Versus Host Disease (GVHD) Prophylaxis in Patients Underwent Allogeneic Bone Marrow Transplantation from Unrelated Donors: Nationwide Survey in Japan.

Acute GVHD is one of the most serious complications and the main cause of morbidity and mortality for patients undergoing allogeneic bone marrow transplantation (BMT). So choice of GVHD prophylaxis has great impact on transplant outcome. We performed large-scale retrospective study for acute GVHD by using the data on patients who underwent BMT from unrelated donors (UR-BMT) facilitated by Japan Marrow Donor Program (JMDP). From January 1993 to September 2004, 4,999 patients were registered in the JMDP. The study population consisted of 3,410 out of 4,999 patients who received UR-BMT from HLA A,B,DR genotyping matched or HLA DR DNA allele mismatched donors(2,626 and 784, respectively). We evaluated the effect of age, HLA compatibility, disease status, GVHD prophylaxis (Cyclosporine:CsA+Methotrexate:MTX or Tacrolimus:FK+MTX), severity of acute GVHD and other factors. The Kaplan-Meier estimates of over all survival(OS) were higher for the FK group than for the CsA group in the case of HLA A,B,DR genotyping matched BMT with 5-year survival rates of 54 and 51.4% (P<0.05) and also in the case of HLA DR DNA allele mismatched BMT, 53.2 and 43.4% (P<0.05), respectively. Those of OS were higher for the FK group than for the CsA group in HLA A,B,DR genotyping matched patients with standard-risk leukemia with 5-year survival rates of 65.9 and 60.8% (P<0.05) but in those patients with high-risk leukemia were not different significantly between FK and CsA group (27.4 and 31%: p=0.61). Those of OS in HLA DR DNA allele mismatched patients with standard-risk leukemia were higher for the FK group than for the CsA group with 5-year survival rates of 64.1 and 52.2%(P<0.05) and also in those patients with high-risk leukemia were higher for the FK group than for the CsA group with 5-year survival rates of 42.4 and 27%(P<0.05). Those of OS in patients under 16 years were higher for the FK group than for the CsA group with 5-year survival rates of 78% and 62.2%(P<0.005) and in patients of 16 and over were comparable for the FK and CsA groups with 5-year survival rates of 48.3% and 45.2%(p=0.179). During the first 100 days post-transplant the incidence of grade I- IV acute GVHD was lower in the FK group compared with CsA group (65.8% and 72.7%, respectively: P<0.005), but the incidence of grade II- IV acute GVHD was similar between FK and CsA groups (40.3% and 41.5%, respectively). The incidence of grade I- IV acute GVHD was lower in the HLA A,B,DR genotyping matched group than in the HLA DR DNA allele mismatched group (68.5% and 74.7%, respectively: P<0.001). Multivariate analysis demonstrated that OS was higher for the younger age group (<16 y/o) than for the older age group (≥16 y/o)(HR:1.86, 95%CI:1.58–2.18) and for the standard-risk leukemia group than for the high-risk leukemia group(HR:2.27, 95%CI:1.58–2.59). These findings indicate that the use of FK for acute GVHD prophylaxis is beneficial for patients undergoing UR-BMT, especially for patients under 16 years and with standard-risk leukemia.

Effect of FK506 on Functional Recovery After Facial Nerve Injury in the Rat

To examine the effect of the immunosuppressive agent FK506 on the rate of functional recovery of the rat facial nerve after crush injury. Forty rats underwent facial nerve crush injury and were randomly assigned to 4 experimental groups: isotonic sodium chloride solution control, FK binding protein 52 (FKBP-52) antibody control, FK506, and FK506 and FKBP-52 antibody. Rats underwent daily recovery testing from postoperative day 9 until postoperative day 21 by videotaping 3 validated variables in this model: blink reflex return, vibrissial fibrillation loss, and return of vibrissial sweeping symmetry. FK506-treated animals demonstrated improved recovery in all 3 variables compared with control animals. The FK506 and FKBP-52 antibody group demonstrated improved recovery of only the return of the blink reflex. FK506 accelerated functional recovery of facial nerve function after crush injury. Neuroregeneration was inhibited by FKBP-52 antibody in the rat midface but not the upper face. FK506 may be a viable adjuvant treatment for facial nerve neurapraxic injury.

Prevention of diltiazem in tacrolimus-induced nephrotoxicity: experiment with rats

To study the nephrotoxicity induced by first oral administration of tacrolimus (FK506) and the prevention of diltiazem (Dil). 24 Sprague-Dawley male rats were randomly divided into 4 equal groups: control (n = 6), cyclosporine A (CsA) group (receiving CsA 25 mg.kg(-1).d(-1) so as to develop CsA-induced nephropathy model), FK506 group (receiving FK506 0.8 mg.kg(-1).d(-1), the common renal transplantation therapeutic dose, so as to develop FK506-induced nephropathy model), FK506 + Dil group (receiving CsA 0.8 mg.kg(-1).d(-1) and Dil 8 mg.kg(-1).d(-1)), and control group. Four weeks later body weight was measured, blood samples were collected to examine the creatinine, urea nitrogen, and uric acid, and urine samples were collected to examine the 24 h urine protein, uric acid, and creatinine. Then the rats were killed with their kidneys taken out to undergo histopathological examination. The urine creatinine levels of the CsA and FK506 groups were significantly lower than that of the control group (both P < 0.05), however, there was no significant difference in urine creatinine between the FK506 + Dil group and control group. The blood creatinine levels of both CsA and FK506 groups were significantly higher than those of the FK506 + Dil group and control group (all P < 0.05), however, there was no significant difference in blood creatinine between the FK506 + Dil group and control group. The urea nitrogen level of the CsA group was significantly higher than those of the other 3 groups (all P < 0.05). The creatinine clearance rates of the CsA and FK506 groups were both significantly lower than that of the control group (both P < 0.05), and the creatinine clearance rate of the FK506 + Dil group was between those of the FK506 group and control group, however, with significant differences with both of them. Histopathology examination showed cloudy swelling and vacuolization of the renal tubular epithelial cells in the CsA and FK506 groups. However, the pathological changes of the FK506 + Dil group were remarkably milder in comparison with these 2 groups. FK506 and CsA at the renal transplantation therapeutic dose induce nephrotoxicity. Diltiazem prevents FK506-induced nephrotoxicity.

FK506 and Sildenafil Promote Erectile Function Recovery after Cavernous Nerve Injury Through Antioxidative Mechanisms

Immunophilin ligands and phosphodiesterase type 5 (PDE5) inhibitors are touted to promote erectile function recovery after cavernous nerve (CN) injury. However, the mechanisms for their effects remain unclear. To compare the erection recovery effects of the immunophilin ligand FK506 and the PDE5 inhibitor sildenafil after CN injury and determine whether they involve antioxidative and/or antiapoptotic mechanisms. Initial experiments established conditions of our CN injury model in adult male Sprague-Dawley rats. Subsequently, we evaluated treatment effects 14 days after: (i) unilateral CN injury (UNI) + saline (vehicle control); (ii) UNI + FK506 (5 mg/kg once daily, subcutaneous x 5 days); (iii) UNI + sildenafil (20 mg/kg every 8 hours, subcutaneous x 7 days); (iv) UNI + FK506/sildenafil; and (v) sham surgery. Intracavernous pressure (ICP) measurement after CN electrical stimulation to assess erectile function and Western blot analysis of expressions of glutathione peroxidase (GPX; antioxidant enzyme), nitrotyrosine (NT; oxidative stress marker), and phosphorylated and total Akt (antiapoptotic factor) in penes. In the UNI model, GPX expression was increased at Days 1 and 7, while p-Akt expression decreased at Day 1 and returned to baseline at Day 7. GPX expression was significantly higher in the UNI + FK506 group compared with the saline-treated group (P < 0.05). ICP increased in all treatment groups compared with that of the saline-treated group (P < 0.05). NT levels were increased after saline treatment (P < 0.05) but not after FK506 and sildenafil treatment, alone or in combination. GPX was localized to nerves coursing through the penis and to smooth muscle and endothelium of the dorsal vein and arteries. Both FK506 and sildenafil protect erectile function after CN injury by decreasing oxidative stress-associated tissue damage. FK506 may act through increased GPX activity. Further research is required to elucidate mechanisms associated with the beneficial effect of sildenafil.

The blockade of the renin-angiotensin system reverses tacrolimus related cardiovascular toxicity at the histopathological level

In this study, we investigate the toxic effects of tacrolimus (FK506) on the cardiovascular system at the histopathological level in a rat model and whether these effects can be reversed by the blockade of the renin-angiotensin system (RAS) by either an angiotensin-converting enzyme inhibitor (ACE-inhibitors) or an angiotensin receptor antagonist (ARB). Thirty-one Wistar rats were divided into four groups. FK506 group was treated with FK506 intraperitoneally (i.p.), FK506+ACE-inhibitors and FK506+ARB groups were treated with either quinapril or valsartan orally in addition to FK506. Control group was treated with saline i.p. Histological and immunohistochemical staining of cardiovascular tissue in the FK506 group showed increased vacuolar degeneration (11.2 vs. 5.8, p=0.008), arterial hyalinosis (10.7 vs. 6.3, p=0.036), transforming growth factor-beta (TGF-beta) (12.2 vs. 4.8, p=0.001) and vascular endothelial growth factor expression (VEGF) (10.7 vs. 6.3, p=0.036), elastic van Gieson (11.5 vs. 5.5, p=0.004), and periodic acid Schiff stain scores (12.5 vs. 4.5, p<0.001) compared to the control group. Immunohistochemical scores showed that expression of TGF-beta is up-regulated, and bone morphogenic protein (BMP-7) is down-regulated with FK506 toxicity. Adding RAS blockade with either an ACE-inhibitor or an ARB could reverse FK506 induced changes. Both FK506+ACE-inhibitors and FK506+ARB groups demonstrated decrease in arterial hyalinosis (22.1 vs. 14.4 (FK506+ACE-inhibitor) and 13.6 (FK506+ARB), p=0.09) and vacuolar degeneration (23.1 vs. 16.1 (FK506+ACE-inhibitor) and 12.4 (FK506+ARB), p=0.006) scores compared to the FK506 group. Blockade of RAS could reverse the histopathological signs of FK506 induced cardiac toxicity in a rat model.

Effects of a new immunosuppressive agent, β-SQAG9, in swine kidney transplantation

We previously reported that 1,2-di- O-acyl-3- O-(- d-sulfoquinovosyl)-glyceride with two stearic acids (β-SQAG9) bound to L-selectin on the cell surface of the CD62L + T-cell subset and inhibited T-cell migration into lymph nodes in a rat skin allograft model. The aim of this study was to verify the efficacy of β-SQAG9 for kidney allograft survival in miniature swine. Recipient swine underwent bilateral nephrectomy and then received renal allograft transplantation from a swine leukocyte antigen-mismatched donor. Swine were divided into 4 experimental groups. The control ( n = 2), 25-SQ ( n = 3), FK ( n = 3) and 10-SQ/FK ( n = 2) groups were treated with no immunosuppressant, 25 mg/kg β-SQAG9, 0.1 mg/kg FK506, and a combination of 10 mg/kg β-SQAG9 and 0.1 mg/kg FK506, respectively, for 14 days. All recipients were autopsied on the day of death to evaluate the cause of death histopathologically. In the control group, the grafts survived for 12 and 15 days. By comparison with the control, β-SQAG9 alone did not contribute to prolongation of graft survival (9, 10 and 24 days), whereas the FK group had significantly longer graft survival (19, 20 and 68 days, p = 0.0289). The 10-SQ/FK pigs died of lethal visceral hemorrhage, although the grafts were still functioning. In conclusion, our results suggest that β-SQAG9 possesses an insufficient immunosuppressive effect for kidney allografts in miniature swine, and may affect blood coagulation and fibrinolysis. In addition, the combination of β-SQAG9 and FK506 can potentially cause severe hemorrhagic complications.

Limited Sampling Strategy for Mycophenolic Acid in Japanese Heart Transplant Recipients

The purpose of the study was to characterize the pharmacokinetics of mycophenolic acid (MPA) in Japanese heart transplant recipients and to find the time point that has the best correlation with the MPA area under the plasma concentration curve (AUC). Twenty-two Japanese recipients treated with mycophenolate mofetil were evaluated in the study. Approximately 9 months after transplantation, the area under the MPA serum concentration-time curve from 0 to 12 h (AUC(0-12 h)) was evaluated. The MPA AUC(0-12 h) h values in the cyclosporine (CsA) and tacrolimus (FK) groups ranged from 13.11 to 50.98 mug . h/ml and from 39.19 to 93.18 mug . h/ml, respectively. Fourteen models were developed and analyzed for their ability to estimate the MPA AUC(0-12 h) based on a limited number of samples in the CsA group. Sixteen models were developed in the FK group. The best model for predicting the full MPA AUC(0-12 h) in the CsA group was a 3-time-point model that included C(0 h), C(1 h) and C(2 h) (r(2), 0.96; mean prediction error, 0.15+/-7.85%); a 2-time-point model that included C(0 h), and C(2 h) (r(2), 0.94; mean prediction error, 0.495+/-10.35%) was also reliable. In the FK group, a 3-time-point model that included C(1 h), C(2 h) and C(4h) (r(2), 0.73; mean prediction error, 2.73+/-17.09%) was the best model for predicting the full MPA AUC(0-12 h), but it was not reliable in clinical practice. A 3-(C(0 h), C(1 h) and C(2 h)) and a 2-time-point model (C(0 h) and C(2 h)) are useful for predicting the full MPA AUC(0-12 h) in Japanese heart transplant recipients treated with CsA but not with FK.

Induction of long-term liver allograft survival by delayed immunosuppression is dependent on interleukin-10

This study aims to investigate the potential role of endogenous interleukin (IL)-10 in long-term liver allograft survival induced by delayed immunosuppression (FK506 days 2-7). Liver transplantation was performed by using Dark Agouti and Lewis rats as donors and recipients, respectively. The delayed immunosuppression protocol induced indefinite allograft survival. A transient upregulation of plasma IL-10 levels was detected in the nontreatment and FK506 treatment groups. Macrophages were found to be one of the major sources of IL-10 produced from the liver allografts. Administration of IL-10-neutralizing antibody shortened the long-term isograft survival and FK506-induced indefinite allograft survival, particularly in the FK506 group. Damaged liver graft histology and increase of plasma alanine aminotransferase levels were detected in the groups with IL-10 antibody treatment. In an ex vivo setting, IL-10 recombinant protein augmented the expression of Foxp3, downregulated the expression of IL-2 and interferon gamma, and induced the generation of CD4(+)CD25(+)Foxp3(+) and CD8(+)CD25(+)Foxp3(+) cells, but this effect was blocked by the administration of IL-10 antibody. Finally, administration of IL-10 recombinant protein after the decline of endogenous IL-10 levels improved allograft survival, and a 100% long-term allograft survival was achieved by the combination of IL-10 with low-dose FK506. In conclusion, the delayed immunosuppression could induce long-term liver allograft survival in the presence of endogenous IL-10 produced by the tissue macrophages. Supplementary exogenous IL-10 administration combined with low-dose immunosuppressive drug may be a useful strategy to induce long-term liver allograft survival.

Small Dense Low-Density Lipoprotein in Renal Transplant Recipients: A Potential Target for Prevention of Cardiovascular Complications?

Background Immunosuppressive therapy is frequently associated with dyslipidemia, which is involved in cardiovascular morbidity and mortality in transplant patients. Beyond classical factors, such as low-density lipoprotein (LDL) cholesterol (LDL-C), qualitative abnormalities of lipoproteins, such as presence of the atherogenic factor, small dense LDL, may be of interest for a cardiovascular risk assessment. This study was designed to explore LDL size in renal transplant recipients in relation to quantitative lipid parameters and apolipoprotein (apo) CIII polymorphism. Methods Total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), LDL-C, apoA1, apoB, apoCIII, and LDL size were measured in 62 patients of mean age 45 ± 13 years including 71% men at 2 ± 0.5 years after renal transplantation. Thirty two patients received cyclosporine (CsA), while 30 received tacrolimus (FK). ApoCIII Sstl genotype was determined by restriction fragment length polymorphism. Results The CsA group exhibited higher TC ( P = .001), LDL-C ( P = .004), non-HDL-C ( P = .009), HDL-C ( P = .03), apoB ( P = .008), and apoCIII ( P = .002) levels than the FK group. However, LDL-C (CsA: 3.7 ± 1.2, FK: 3.0 ± 0.6 mmol/L) and triglyceride levels (CsA: 1.55 mmo/L, FK: 1.37 mmol/L) were near the normal range in both groups. Allelic frequency of the sparse A2 allele associated with hypertriglyceridemia was 6%, similar to the general population. LDL size, which was comparable in the CsA and FK groups (25.87 ± 0.89 vs 25.75 ± 0.62 nm, respectively), inversely correlated with TG/HDL ratio ( P = 10 −4). Prevalence of small dense LDL (defined as <25.5 nm) was 26% in the CsA group and 33% in the FK group. Conclusion After LDL-C goal has been achieved, LDL size modulation may be taken into account in order to prevent cardiovascular complications.

Comparison of Rapamycin Versus FK506 on Expression of Cytoprotective Genes in the Rat Kidney Allografts Undergoing Chronic Allograft Nephropathy

Aims It is increasingly recognized that expression of cytoprotective genes in grafts can affect the progress of chronic allograft nephropathy (CAN). Little is known about the influence of different immunosuppressive regimens on expression of cytoprotective genes in allografts undergoing CAN. We investigate whether there is difference between rapamycin (Rapa) and FK506 in the expression of cytoprotective genes in rat kidney allografts undergoing CAN. Methods Sprague-Dawley rat renal grafts were orthotopically transplanted into Wistar rats following the procedure of Kamada with our modification. The recipients were divided into three oral treatment groups: group 1: vehicle group (cyclosporine [CsA] 10 mg/kg · d × 10 days followed by vehicle); group 2: Rapa group (CsA 10 mg/kg · d × 10 d followed by Rapa 0.8 mg/kg · d); group 3: FK506 group (CsA 10 mg/kg · d × 10 d followed by FK506 0.15 mg/kg · d). At the same times after transplantation, the rats were sacrificed to harvest the renal allografts. The expression of four cytoprotective genes, A20, heme oxygenase (HO)-1, Bcl-2, and Bcl-X/L were analyzed in these grafted kidneys by quantitative reverse transcriptase polymerase chain reaction and immunohistochemistry. Results Four cytoprotective genes were all detected in rat kidney allografts undergoing CAN. The expression of A20 in the Rapa group was significantly higher than that in the FK506 or the vehicle group ( P < .05). There was no significant difference between the Rapa group and FK506 group in the expressions of HO-1, Bcl-2, and Bcl-X/L. Conclusions We demonstrate that various immunosuppressive agents have different effects on the expression of cytoprotective genes.

Comparison of Expression of TGF-β1, its Receptors TGFβ1R-I and TGFβ1R-II in Rat Kidneys During Chronic Nephropathy Induced by Cyclosporine and Tacrolimus

Objective Chronic rejection is a major cause of graft dysfunction after kidney transplantation. Long-term treatment with cyclosporine (CsA) or tacrolimus (FK506) results in chronic nephrotoxicity. Transforming growth factor-beta1 (TGFβ1) and its receptors type I (TR-I) and type II (TR-II) have been known to contribute to this side effect. The expression of TGF-β1, TR-I, and TR-II in rat kidneys has not been compared during chronic nephropathy induced by CsA or FK506. Methods Rat models of chronic CsA- or FK506-induced nephropathy were established using Sandimun Neoral or Prograf administration. The kidneys were dissected and TGF-β1, TR-I, and TR-II proteins and TR-I and TR-II mRNAs measured by immunohistochemistry and in situ hybridization, respectively, to compare the results of the two groups. Results The functional and morphologic studies showed that in the rats the nephrotoxic effects of FK506 were not as significant as those of CsA. The results of immunohistochemistry and in situ hybridization showed that the expression of renal TGFβ1, TR I, TR-II proteins and TR and TR II mRNA in the FK506 group were lower than those in the CsA groups. Conclusion These results showed that both FK506 and CSA display nephrotoxicity, but that the nephrotoxicity of FK506 was less than that of CsA in chronic nephropathy.

The Effects of Different Immunosuppressants on Chronic Allograft Nephropathy by Affecting the Transforming Growth Factor-β and Smads Signal Pathways

Objective This study investigated the effects of various immunosuppressants on chronic allograft nephropathy (CAN) by affecting transforming growth factor-β (TGF-β) and Smads signal pathway. Methods Vascular smooth muscle cells (VMSC) from rat aorta were incubated for 6 or 12 hours with various immunosuppressants. Cyclosporine (CsA) (3 μg/mL), FK506 (1 μg/mL), mycophenolate mofetil (MMF) (0.3 μg/mL), rapamycine (Rapa) (10 μg/mL), CsA (1 μg/mL/MMF 0.3 μg/mL). We used the Sprague-Dawly Wistar rat accelerated kidney sclerosis model. Before transplantation, the kidney was preserved 1 hour in 0°C to 4°C heparin sodium chloride solution to reinforce the cold ischemia injury. The rats were divided into eight groups (each group n = 8): group A, pseudo-OP; group B, isotransplantation; group C, CsA 6 mg/kg · d; group D, FK506 0.15 mg/kg · d; group E, MMF 20 mg/kg · d; group F, Rapa 0.8 mg/kg · d; group G, CsA 3 mg/kg · d + MMF 20 mg/kg · d. The serum creatinine levels and pathological changes, according to the Banff scheme, were observed at 2, 4, 6, 8 and 12 weeks posttransplantation. Immunohistochemistry and quantitative fluorescence polymerase chain reactions were used to end localize and quantitate the expression of TGF-β1 and Smad 2, 3, 7 in VMSC and in the transplanted kidney. Results CsA and FK506 stimulated gene expression and protein production of TGF-β1, smad2, and smad3, but inhibited expression of smad7 both in VSMC and in the transplanted kidney. In contrast, MMF and Rapa down-regulated gene expression and protein production of TGF-β1, smad2, 3 while up-regulating expression of smad7. There was no significant difference between the CsA group and the FK506 group, as well as the MMF group and the Rapa group. The group treated with CsA + MMF was similar to the MMF and the Rapa groups. Conclusion Our study suggested that various immunosuppresants affected differentially TGF-β1 and Smads signal pathways in rat VSMC and kidney grafts. CsA and FK506 can cause CAN, owing to up-regulated expression of smad2 and smad3, and down-regulation of smad7 expression. MMF and Rapa can prevent the CAN progression, because of down-regulation of the expression of smad2 and smad3, with increased smad7 production.

Neuroregenerative Effects of Preinjury FK-506 Administration

FK-506 is used in organ transplantation because it promotes neurite outgrowth in vitro and enhances neuroregeneration in peripheral nerve injury transection models. Immunosuppressive mechanisms of FK-506 are well defined, with demonstration of decreased neuroregenerative effects with delayed administration. The purpose of this study was to describe the effects of preinjury administration of FK-506 in rats with tibial nerve transection injury. Eight inbred male Lewis rats per group in three separate groups underwent tibial nerve transection with primary repair. Group I received placebo, group II received FK-506 treatment at 1 day before surgery, and group III received FK-506 preloading 3 days before surgery. Histologic and histomorphometric results demonstrated the preload FK-506 group had superior results compared with the immediate FK-506 group. Both FK-506 groups were superior to the placebo group. The preload FK-506 demonstrated superior regeneration in mean total nerve fiber counts (p < 0.05), greater percentage neural tissue (p < 0.05), greater mean nerve fiber density (p < 0.05), and lower percentage of debris (p > 0.05). Mean nerve fiber widths were similar in the preload and immediate FK-506 groups but superior to the placebo group. These data suggest that enhancement of FK-506's neuroregenerative effect is enhanced when administered before nerve injury such as when performing elective surgery.

Steroid withdrawal in living donor renal transplant recipients using tacrolimus and cyclosporine: a randomized prospective study*

Steroids have been a mainstay of immunosuppressive regimens in renal transplantation despite their adverse effects. The introduction of new immunosuppressant has improved the survival rates and prompted trials of steroid withdrawal. We conducted a randomized prospective study to compare steroid withdrawal at 6 months post-transplant between tacrolimus + mycophenolate mofetil (MMF) (FK group) versus cyclosporine A + MMF (CsA group). Steroid was withdrawn at 6 months post-transplant under the condition of no rejection episode proven by biopsy and maintenance of serum creatinine level <2.0 mg/dl. Fourteen recipients were excluded because of acute rejection within 6 months or protocol violation. Steroid could be tapered off in 62 in FK group and 55 in CsA. Three cases in FK group and five in CsA had acute rejection within another 6 months after steroid withdrawal (P > 0.05). At 12 months, the incidence of post-transplant diabetes was 18.6% vs. 8.0% in FK and CsA group. And hypercholesterolemia was presented in 8.5% vs. 2.0%, hypertension in 47.5% vs. 56.0%, and serum creatinine level 1.18 +/- 0.24 mg/dl vs. 1.18 +/- 0.20 mg/dl, respectively (P > 0.05). Steroid withdrawal may be carried out successfully using both FK and CsA with MMF, but long-term follow-up is necessary.

Liver Pathology and Cell Proliferation After Calcineurin Inhibitors and Antiproliferative Drugs Following Partial Hepatectomy in Rats

Immunosuppressants are the cornerstones of treatment after solid organ transplantation. This study investigated the pathology and cell proliferation following partial hepatectomy (PH) in rats undergoing immunosuppressive treatment. After 1 day, all rats were subjected to 70% PH. Groups A and B (n = 10) received calcineurin inhibitors subcutaneously: either FK506 or cyclosporine (CyA). Groups C and D (n = 10) received antiproliferative drugs: either mycophenolate mofetil (MMF) or sirolimus (SRL) by gavage. A control group (n = 5) received 1 mL of tap water daily. On postoperative day 2, all rats were sacrificed to obtain liver tissue for pathologic examination. Using immunohistochemistry we separately examined the hepatectomy surface and the liver parenchyma. In the parenchyma, the Ki-67 indices were higher in the CyA and FK506 groups and lower in the SRL and MMF groups compared with controls ( P < .01). CyA had the highest and MMF the lowest values. On the hepatectomy surface, Ki-67 indices and TGF-alpha expressions were higher in the CyA group and lower in the SRL and MMF groups compared with the control group ( P < .01). Slightly higher values in the FK506 group were not significantly different compared with the control group ( P > .05). All groups other than FK506 showed prominent cholangiolar epithelial phenotypes compared with the control group. In the CyA and SRL groups, the number of cholangiolar cells was higher ( P < .01), and in the MMF group lower than in the control group ( P < .01). Among all groups, SRL had the highest values.

The Effect of Combined Fentanyl and Ketamine Administration in the Monitored Anesthetic Care of Ambulatory Strabismus Surgery

Background: Monitored anesthetic care can provide patient safety and optimal surgical conditions. However, propofol and fentanyl decrease the hypoxic ventilatory response and heart rate. Ketamine has less respiratory depression and blocks bradycardia. Furthermore, ketamine can reduce the amount of opioid use but it delays discharge in the outpatient anesthesia. Therefore, this study retrospectively examined the effects of combined fentanyl and ketamine administered during monitored anesthetic care on the use of opioids, cardiorespiratory side effects, and patient discharge. Methods: The anesthetic room, the recovery room and day surgery center records of ambulatory strabismus surgery with monitored anesthetic care was reviewed by anesthesiologists from Oct. 2004 to July 2005. The patients were classified as those receiving either fentanyl (F group: n = 32) or fentanyl and ketamine (FK group: n = 19) with a propofol infusion. The fentanyl dose used, the need for airway support, anticholinergics and naloxone were compared. The stay in the recovery room and the day surgery center was also examined. Results: The FK group used significantly less fentanyl than the F group (P ＜ 0.05). Although airway support was needed, there was less anticholinergics and naloxone used in the FK group, but this difference was not significant. The stay in recovery room and the day surgery center were similar. Conclusions: Co-administered ketamine reduces the amount of fentanyl-needed, but it does not reduce the need for airway support and anticholinergics. In addition, co-administered ketamine does not affect the number of days in the recovery room and day surgery center.