tacrolimus and Necrosis

Antibody-mediated rejection (AMR) frequently causes refractory graft dysfunction. This randomized controlled trial was designed to evaluate whether immunoadsorption (IA) is effective in the treatment of severe C4d-positive AMR. Ten out of 756 kidney allograft recipients were included. Patients were randomly assigned to IA with protein A (N = 5) or no such treatment (N = 5) with the option of IA rescue after 3 weeks. Enrolled recipients were subjected to tacrolimus conversion and, if indicated, 'anti-cellular' treatment. All IA-treated patients responded to treatment. One death unrelated to IA occurred after successful reversal of rejection. Four control subjects remained dialysis-dependent. With the exception of one patient who developed graft necrosis, non-responders were subjected to rescue IA, however, without success. Because of a high graft loss rate in the control group the study was terminated after a first interim analysis. Even though limited by small patient numbers, this trial suggests efficiency of IA in reversing severe AMR.

Topics: Adult; Aged; Complement C4b; Graft Rejection; Humans; Immunotherapy; Kidney Transplantation; Middle Aged; Necrosis; Peptide Fragments; Renal Dialysis; Staphylococcal Protein A; Tacrolimus; Treatment Outcome

Topics: Biopsy; Fibrosis; Graft Rejection; Humans; Kidney Glomerulus; Kidney Transplantation; Necrosis; Tacrolimus; Time Factors; Transplantation, Homologous

Topics: Arterioles; Biopsy; Creatinine; Follow-Up Studies; Graft Rejection; Humans; Kidney Glomerulus; Kidney Transplantation; Kidney Tubules; Microscopy, Electron; Necrosis; Tacrolimus; Vacuoles

Renal graft cortical necrosis (GCN) is a catastrophic cause of graft failure. We evaluated the incidence, causes, management, and outcome of GCN across two decades from our center.. This is a retrospective analysis of transplant patients who had biopsy-proven GCN transplanted between 2000 and 2020. The clinical details, immunological workup, induction, maintenance regimen, causes of cortical necrosis, and the outcomes were compared between the first period 2000-2012, and the second period 2013-2020, when Flow cytometric and Luminex based crossmatch were included in the workup plan.. Among 2333 live ABO-compatible renal transplants, 37 (0.015%) patients (36 patients between 2000 and 2012 and 1 between 2013 and 2020) developed GCN (60% had diffuse and 40% patchy GCN) at a median of 8 days after transplantation.Twenty-six (60%) received ATG, 4 received plasmapheresis and ATG (10.8%) as antirejection therapy. The cyclosporine-based regimen was associated with a higher risk of GCN (RR 2.54; 95% CI 1.26 to 5.12, p = 0.009), whereas tacrolimus-based therapy had a lower risk (RR 0.39; 95% CI 0.19 to 0.79, p = 0.009). The introduction of flow cytometry and DSA assay has significantly decreased the incidence of acute rejection and GCN. Only one patient had GCN during the 2013-2020 period because of graft's mucormycosis. Twenty-five (67.56%) patients had no recovery, and 12 (32.43%) had partial recovery of graft function.. GCN is mainly associated with rejection, and cyclosporin-based maintenance regimen had a higher incidence. The remarkable decrease in GCN after 2012 onwards could be attributed to the use of Flowcytometry, Luminex-based DSA assays, and tacrolimus-based regimens.

Topics: Allografts; Cyclosporine; Graft Rejection; Graft Survival; Humans; Immunosuppressive Agents; Kidney Transplantation; Living Donors; Necrosis; Retrospective Studies; Tacrolimus

Our previous rodent studies demonstrated significantly decreased full-thickness necrosis in pedicled dorsal skin flaps with topical tacrolimus as compared with petroleum jelly. Histologically, we found that topical tacrolimus was correlated with increased vascular growth in areas more susceptible to ischemic damage. The purpose of this study was to investigate the potential benefits of pretreatment with tacrolimus. By applying tacrolimus in advance of raising the dorsal skin flaps, we hoped to increase vascularity and thus increase the overall viability of the flaps.. Twenty Sprague-Dawley rats were initially randomized to 4 groups based on timing of tacrolimus treatment (presurgical/postsurgical treatment): control/control (C/C), control/tacrolimus (C/T), tacrolimus/control (T/C), and tacrolimus/tacrolimus (T/T). Treatments consisted of 0.2 g of the control (topical petroleum jelly) and 0.1% topical tacrolimus to the rat dorsum twice per day. After 7 days of presurgical treatment, a cranially based dorsal skin flap measuring 3 × 10 cm was created. Two rats perished during surgery and were excluded for further analysis. Each rat was treated for a further 7 days and sacrificed. Two blinded reviewers marked the total skin flap area as well as areas of viable tissue, reversible ischemia, and full-thickness necrosis. Percentage areas were calculated using Fiji/ImageJ, and statistical analysis was performed in R.. The average viable areas for C/C, C/T, T/C, and T/T were 31.4%, 31.9%, 35.6%, and 22.6%, respectively. The average reversible ischemic area for C/C, C/T, T/C, and T/T was 53.1%, 54.0%, 54.1%, and 71.5%, respectively. The average necrotic area for C/C, C/T, T/C, and T/T was 15.4%, 14.0%, 10.2%, and 5.9%, respectively. For areas of reversible ischemia, T/T arm had higher areas compared with C/T (P = 0.004) and T/C (P = 0.044). There was no significance between treatment arms for areas of viable and necrotic tissue.. We observed higher areas of reversible ischemia for continuous tacrolimus treatment compared with only pre-tacrolimus application or post-tacrolimus application. This suggests that tacrolimus application before and after surgical insult may be associated with improved ischemic survival of the skin. Although we did not observe decreased areas of necrosis for tacrolimus treatment compared with control, this was likely due to the limited number of rats available in each arm to reach significance. Further study is needed to fully elucidate the encouraging trends that were observed.

Topics: Animals; Graft Survival; Ischemia; Necrosis; Petrolatum; Pilot Projects; Rats; Rats, Sprague-Dawley; Soft Tissue Injuries; Surgical Flaps; Tacrolimus

A 62-year-old Japanese man with swollen fingers and walking difficulty due to myalgia and muscle weakness in proximal limb muscles was admitted to our hospital. Serum creatine kinase was remarkably increased (7,380 U/l) and rapidly progressing interstitial pneumonia developed. Muscle biopsy showed necrotic and regenerating fibers without mononuclear infiltration and fibrosis. Anti-Th/To antibodies were detected in the serum, and anti-Th/To antibody-positive systemic sclerosis was diagnosed. Anti-Th/To antibody-positive sclerosis-associated myopathy has not yet been reported in the literature. The present case suggests that anti-Th/To antibody-positive systemic sclerosis can be accompanied by immune-mediated necrotizing myopathy and be effectively treated with immunotherapy comprising corticosteroids, tacrolimus and immunoglobulin.

Topics: Adrenal Cortex Hormones; Autoantibodies; Autoantigens; Humans; Immunoglobulins; Immunotherapy; Lung Diseases, Interstitial; Male; Middle Aged; Muscle, Skeletal; Muscular Diseases; Necrosis; Scleroderma, Systemic; Skin; Tacrolimus

Our previous rodent study demonstrated significantly decreased full-thickness necrosis in pedicled dorsal skin flaps with topical tacrolimus as compared with petroleum jelly. The pathophysiology of tissue necrosis involves lymphatic congestion, followed by venous congestion and ultimately arterial insufficiency. Topical tacrolimus has been shown to increase growth of lymphatic collateral vessels and decrease lymphedema, potentially obviating one contributor to necrosis. The purpose of this study was to investigate the vascular and histological differences between these 2 groups to identify the etiology of our research findings.. A 3 × 10-cm cranially based dorsal skin flap was raised and reinset on 22 Sprague Dawley rats. They were randomized to receive 0.2 g of either topical petroleum jelly or topical 0.1% tacrolimus ointment daily to the flaps. The rats were killed 7 days postoperatively. Two blinded reviewers marked the total flap area as well as areas of viable tissue, reversible ischemia, and necrotic tissue. Full-thickness biopsies of each area were taken from 2 randomly chosen animals in each group. Paraffin-embedded tissue was sectioned to generate hematoxylin and eosin-stained slides. Representative images of each area of the flap were taken less than 40× magnification using light microscopy. Arteries, veins, and lymphatics in the dermal layer were quantified under blinded conditions by a trained pathologist and calculated per cross-sectional area using Fiji software.. The average area of the dorsal flaps in the control and tacrolimus groups was 22.5 and 23.9 cm2, respectively. Total necrotic area was significantly lower in rats receiving topical tacrolimus as compared with controls (P = 0.015). In the control cohort, average total number of vessels was 12.5, 6, and 0, in the areas of viable tissue, reversible ischemia, and necrosis, respectively. In the tacrolimus cohort, average total number of vessels increased was 20, 11.5, and 5.4, in the areas of viable tissue, reversible ischemia, and necrosis, respectively.. On a histological level, topical tacrolimus is correlated with increased vascular growth in areas most susceptible for ischemic damage as compared with topical control. Future work is needed to investigate vascular biomarkers and increase the power of our study.

Topics: Animals; Graft Survival; Necrosis; Rats; Rats, Sprague-Dawley; Skin Transplantation; Surgical Flaps; Tacrolimus

Skin necrosis is a known postoperative complication of mastectomies. The pathophysiology of tissue necrosis involves lymphatic congestion, followed by venous congestion and ultimately arterial insufficiency. Recent mouse model studies have shown topical tacrolimus to increase growth of lymphatic collateral vessels and decrease lymphedema, potentially obviating the cycle of necrosis and increasing skin survival. The purpose of this study was to investigate the effect of topical tacrolimus on skin flap necrosis in a rat model.. A cranially based dorsal skin flap measuring 3 × 10 cm was raised and reinset on 22 Sprague-Dawley rats. They were then randomized to either the control (topical petroleum jelly) or the treatment (topical 0.1% tacrolimus) arm. In addition, 0.2 g of either ointment was spread over the flap and then covered with an occlusive dressing. Dressings were changed daily with reapplication of both the topical ointment and occlusive dressing. The rats were sacrificed 7 days postoperatively; areas of viable tissue, reversible ischemia, and full thickness necrosis were measured with Fiji software, and comparative analysis was performed with GraphPad statistical software.. The average area of the dorsal flaps in the control and tacrolimus groups was 22.5 and 23.9 cm, respectively. In the control cohort, the average viable area was 42.4%, the average reversible ischemia area was 43.6%, and the average necrotic area was 13.9%. In the tacrolimus cohort, the average viable area was 31.5%, the average reversible ischemia area was 59.3%, and the average necrotic area was 9.2%. Total necrotic area was significantly lower in rats receiving topical tacrolimus as compared with controls (P = 0.015). Furthermore, the ratios of necrotic to reversible ischemia and necrotic to viable tissue were significantly lower in the tacrolimus group as compared with controls (P = 0.003, P = 0.015). There was one incidence of wound dehiscence secondary to rodent self-removal of dressings and suture that required reoperation and reinset.. Topical tacrolimus was associated with significantly less full thickness necrosis as compared with topical.

Topics: Animals; Graft Survival; Ischemia; Mice; Necrosis; Rats; Rats, Sprague-Dawley; Skin Transplantation; Surgical Flaps; Tacrolimus

Topics: Administration, Topical; Antiphospholipid Syndrome; Humans; Necrosis; Tacrolimus; Warfarin

Topics: Age of Onset; Autoantibodies; Biopsy; Child; Drug Administration Schedule; Drug Therapy, Combination; Glucocorticoids; Humans; Male; Methotrexate; Muscle, Skeletal; Muscular Diseases; Necrosis; Signal Recognition Particle; Tacrolimus; Treatment Outcome

To study the clinical and histopathologic features of post-transplant kidney biopsy tissues from pediatric C-III donors.. The clinical and pathologic features of 20 cases (22 case-times) of renal transplant biopsies from pediatric cadaveric donors were analyzed by light microscopy and immunohistochemistry according to the Banff system of working classification of renal allograft pathology. Biopsies were compared to those from adult C-III donors and adult cadaveric donors.. Sixteen cases (72.7%) showed renal allograft drug toxicity damage by Tacrolimus, seven cases (31.8%) showed degeneration and necrosis of renal tubular epithelial cells, four cases (18.2%) showed T cell-mediated acute rejection and six cases (27.3%) showed renal interstitial inflammation. There were two cases (9.1%) of renal dysplasia and one case (4.5%) of renal infarction. There was insufficient evidence for diagnosis of renal allograft nephropathy. Compared to post-transplant kidney from adult C-III donors, the proportion of drug toxicity damage was higher (P<0.05). Compared to post-transplant kidney from adult cadavers, the proportions of drug toxicity damage, degeneration and necrosis of renal tubular epithelial cells were higher (P<0.05) while the proportion of acute rejection was lower (P<0.05).. The pathologic changes in the post-transplant kidneys from pediatric donors are different from those from adult donors. Optimal long-term outcome can be accomplished by effective treatment based on timely or procedural biopsy.

Topics: Adult; Age Factors; Biopsy; Cadaver; Child; Graft Rejection; Humans; Immunohistochemistry; Immunosuppressive Agents; Infarction; Kidney; Kidney Transplantation; Kidney Tubules; Necrosis; Tacrolimus; Transplantation, Homologous; Treatment Outcome

Radiocontrast agents are required for radiographic procedures, but these agents can injure tissues by unknown mechanisms. We investigated whether exposure of pancreatic tissues to radiocontrast agents during endoscopic retrograde cholangiopancreatography (ERCP) causes pancreatic inflammation, and studied the effects of these agents on human cell lines and in mice.. We exposed mouse and human acinar cells to the radiocontrast agent iohexol (Omnipaque; GE Healthcare, Princeton, NJ) and measured intracellular release of Ca(2+), calcineurin activation (using a luciferase reporter), activation of nuclear factor-κB (NF-κB, using a luciferase reporter), and cell necrosis (via propidium iodide uptake). We infused the radiocontrast agent into the pancreatic ducts of wild-type mice (C57BL/6) to create a mouse model of post-ERCP pancreatitis; some mice were given intraperitoneal injections of the calcineurin inhibitor FK506 before and after infusion of the radiocontrast agent. CnAβ(-/-) mice also were used. This experiment also was performed in mice given infusions of adeno-associated virus 6-NF-κB-luciferase, to assess activation of this transcription factor in vivo.. Incubation of mouse and human acinar cells, but not HEK293 or COS7 cells, with iohexol led to a peak and then plateau in Ca(2+) signaling, along with activation of the transcription factors NF-κB and nuclear factor of activated T cells. Suppressing Ca(2+) signaling or calcineurin with BAPTA, cyclosporine A, or FK506 prevented activation of NF-κB and acinar cell injury. Calcineurin Aβ-deficient mice were protected against induction of pancreatic inflammation by iohexol. The calcineurin inhibitor FK506 prevented contrast-induced activation of NF-κB in pancreata of mice, this was observed by live imaging of mice given infusions of adeno-associated virus 6-NF-κB-luciferase.. Radiocontrast agents cause pancreatic inflammation in mice, via activation of NF-κB, Ca(2+) signaling, and calcineurin. Calcineurin inhibitors might be developed to prevent post-ERCP pancreatitis in patients.

Topics: Animals; Calcineurin; Calcineurin Inhibitors; Calcium Signaling; Chlorocebus aethiops; Contrast Media; COS Cells; Disease Models, Animal; Gene Expression Regulation; HEK293 Cells; Humans; Iohexol; Male; Mice, Inbred C57BL; Mice, Knockout; Necrosis; NF-kappa B; Pancreas, Exocrine; Pancreatitis; Tacrolimus; Time Factors

Proliferative and necrotising otitis externa is a rare and recently described disease affecting the ear canals and concave pinnae of kittens. This article describes a case of proliferative and necrotising otits externa in a young adult cat. In this case, the lesions did not affected the pinnae, but both ear canals were severely involved. Video-otoscopy revealed a digitally proliferative lesion, growing at 360° all around the ear canals for their entire length, without involvement of the middle ear. Histopathological examination confirmed the diagnosis, and the cat responded completely to a once-daily application of 0.1% tacrolimus ointment diluted in mineral oil in the ear canals. Video-otoscopy findings, not described previously, were very peculiar and may help clinicians to diagnose this rare disease.

Topics: Animals; Cat Diseases; Cats; Ear; Immunosuppressive Agents; Male; Necrosis; Otitis Externa; Tacrolimus; Treatment Outcome

To evaluate the effects of the immunosuppressant tacrolimus on rejection of a transplanted uterus and on uterine expression of markers of inflammation and implantation.. Experimental study.. University laboratory.. Female rats.. Uteri from brown Norway rats were transplanted to Lewis rats, receiving either tacrolimus or no treatment. Sham groups underwent either hemihysterectomy or tacrolimus treatment.. Gross morphology, histology, density of T-lymphocytes by immunohistochemistry, and mRNA levels of interleukin (IL)-1α, leukemia inhibitory factor (LIF), galectin-1, CD200, IL-15, interferon-inducible protein-10 (IP-10), and nuclear factor-κB (NF-κB) at 14 days' post-transplantation.. Nontreated uterine grafts showed rejection with necrosis. Sham groups and the tacrolimus-treated transplanted group exhibited normal uterine morphology with low numbers of T-lymphocytes in all uteri except in two out of seven uteri of the tacrolimus-treated transplant group. Uteri of the nontreated transplanted group showed elevated mRNA expression of IL-1α and IP-10 and reduced galectin-1, compared with the tacrolimus-treated transplanted group. There was no difference between any groups concerning uterine expression of LIF, NF-κB, IL-15, and CD200.. Tacrolimus monotherapy suppresses rejection of an allotransplanted uterus and normalizes the expression of IL-1α and IP-10 and prevents T-lymphocyte infiltration.

Topics: Animals; Biomarkers; Chemokine CXCL10; Female; Galectin 1; Graft Rejection; Hysterectomy; Immunosuppressive Agents; Inflammation; Interleukin-1alpha; Necrosis; Organ Transplantation; Rats; Rats, Inbred BN; Rats, Inbred Lew; Tacrolimus; Transplantation, Homologous; Uterus

Liver cell transplantation (LCT) is considered a new therapeutic strategy for the treatment of acute liver failure and inborn metabolic defects of the liver. Although minimally invasive, known safety risks of the method include portal vein thrombosis and pulmonary embolism. Since no systematic data on these potential side effects exist, we investigated the toxicological profile of repeated intraportal infusion of allogeneic liver cells in 30 rabbits under GLP conditions. Rabbit liver cells were administered once daily for 6 consecutive days at 3 different dose levels, followed by a 2-week recovery period. No test item-related mortality was observed. During cell infusion, clinical findings such as signs of apathy and hyperventilation, moderate elevations of liver enzymes ALT and AST and a slight decrease in AP were observed, all fully reversible. Cell therapy-related macroscopic and histological findings, especially in liver and lungs, were observed in animals of all dose groups. In conclusion, the liver and lungs were identified as potential toxicological target organs of intraportal allogeneic liver cell infusion. A NOAEL (no observed adverse effect level) was not defined because of findings observed also in the low-dose group. No unexpected reactions became apparent in this GLP study. Overall, LCT at total doses up to 12 % (2 % daily over 6 days) of the total liver cell count were tolerated in rabbits. Observed adverse effects are not considered critical for treatment in the intended patient populations provided that a thorough monitoring of safety relevant parameters is in place during the application procedure.

Topics: Animals; Cell Transplantation; Central Venous Catheters; Embolism; Female; Germany; Hepatocytes; Immunosuppressive Agents; Liver; Liver Transplantation; Lung; Male; Necrosis; Portal Vein; Pulmonary Embolism; Rabbits; Risk Assessment; Tacrolimus; Thrombosis; Transplantation, Homologous

To detect the mechanisms of death in allogeneic myofibers rejected by the immune system, myoblasts were allotransplanted in muscles of macaques immunosuppressed with tacrolimus. Immunosuppression was stopped 1month later to induce a massive rejection of allogeneic myofibers. Grafted sites were biopsied at 2-week intervals and analyzed by histology. The loss of allogeneic myofibers was rapid and concomitant with an intense infiltration of CD8+ lymphocytes. Several necrotic myofibers were observed in the lymphocyte accumulations by intracellular complement immunodetection. Dystrophin and spectrin immunodetection showed sarcolemmal damage in myofibers surrounded and invaded by CD8+ lymphocytes. Active caspase-3 was immunodetected in some myofibers surrounded by CD8+ lymphocytes. This is the first evidence that the collapse of myofibers attacked by T lymphocytes occurs by necrosis possibly due to damage of the sarcolemma. Caspase 3 is activated at least in some myofibers, but there was no evidence of a complete classical process of apoptosis.

Topics: Animals; Apoptosis; Caspase 3; CD8-Positive T-Lymphocytes; Immunosuppression Therapy; Immunosuppressive Agents; Macaca; Muscle, Skeletal; Necrosis; Sarcolemma; Tacrolimus

Topics: Animals; Cat Diseases; Cats; Ear; Immunosuppressive Agents; Male; Necrosis; Otitis Externa; Tacrolimus; Treatment Outcome

The protective potential of immunosuppressants has been reported in many experimental models of ischemia both in vivo and in vitro, suggesting a novel therapeutic application of these drugs. Because high-mobility group box 1 (HMGB1) protein has recently been reported to be involved in ischemic brain injury, the purpose of the present study was to determine whether treatment with immunosuppressants could decrease the expression and release of HMGB1 in astrocytes exposed to simulated ischemic conditions (combined oxygen-glucose deprivation, OGD). We also investigated whether immunosuppressive drugs could attenuate necrosis in astrocyte cultures exposed to OGD. Finally, we studied the influence of immunosuppressants on the expression of NFκB, inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2). Cells were treated with cyclosporine A, FK506 and rapamycin (all drugs at concentrations of 0.1, 1 and 10 μM). Our study provides evidence that immunosuppressants decrease the expression and release of HMGB1 in ischemic astrocytes. Our data suggest that HMGB1 release may be partly an active process triggered by oxidative stress because the antioxidant N-acetylcysteine (NAC) clearly attenuated HMGB1 expression and release. Furthermore, we show that the immunosuppressants, at the same concentrations that significantly suppressed HMGB1 expression and release, were also able to prevent the necrosis of ischemic astrocytes and inhibit the expression of inflammatory mediators (NFκ, iNOS and COX-2). These results provide further information about the cytoprotective mechanisms of immunosuppressants on ischemic astrocytes, especially in relation to the pathophysiology of ischemic brain injury. It appears that the protective effects of immunosuppressants can be mediated in part by the suppressing the expression and release of HMGB1 in astrocytes, which leads to the attenuation of ischemia-induced necrosis and neuroinflammation.

Topics: Animals; Astrocytes; Cell Hypoxia; Cells, Cultured; Cyclosporine; Dose-Response Relationship, Drug; Glucose; HMGB1 Protein; Immunosuppressive Agents; Inflammation; Inflammation Mediators; Ischemia; Necrosis; Oxidative Stress; Rats; Rats, Wistar; Sirolimus; Tacrolimus

Huntington's disease (HD) is a genetic neurodegenerative disorder characterized by striatal neurodegeneration, involving apoptosis. FK506, an inhibitor of calcineurin (or protein phosphatase 3, formerly known as protein phosphatase 2B), has shown neuroprotective effects in several cellular and animal models of HD. In the present study, we show the protective effects of FK506 in two striatal HD models, primary rat striatal neurons treated with 3-nitropropionic acid (3-NP) and immortalized striatal STHdh cells derived from HD knock-in mice expressing normal (STHdh(7/7)) or full-length mutant huntingtin (FL-mHtt) with 111 glutamines (STHdh(111/111)), under basal conditions and after exposure to 3-NP or staurosporine (STS). In rat striatal neurons, FK506 abolished 3-NP-induced increase in caspase-3 activation, DNA fragmentation/condensation and necrosis. Nevertheless, in STHdh(111/111) cells under basal conditions, FK506 did not prevent, in a significant manner, the release of cytochrome c and apoptosis inducing factor (AIF) from mitochondria, or alter Bax/Bcl-2 ratio, but significantly reverted caspase-3 activation. In STHdh(111/111) cells treated with 0.3mM 3-NP or 25 nM STS, linked to high necrosis, exposure to FK506 exerted no significant effects on caspase-3 activation. However, treatment of STHdh(111/111) cells exposed to 10nM STS with FK506 effectively prevented cell death by apoptosis and moderate necrosis. The results suggest that FK506 may be neuroprotective against apoptosis and necrosis under mild cell death stimulus in the presence of FLmHtt.

Topics: Animals; Apoptosis; Blotting, Western; Caspase 3; Cell Death; Cell Line; Corpus Striatum; Cytosol; DNA Fragmentation; Humans; Huntingtin Protein; Huntington Disease; Immunosuppressive Agents; Mice; Mice, Transgenic; Mitochondria; Necrosis; Nerve Tissue Proteins; Neurons; Neuroprotective Agents; Nitro Compounds; Nuclear Proteins; Propionates; Staurosporine; Subcellular Fractions; Tacrolimus

Otitis externa in cats is relatively uncommon. This report describes a case of a rare, visually distinctive, proliferative and necrotising otitis in a three-month-old Persian kitten. The cat had proliferative, erythematous and necrotic tissue covering most of the proximal pinnae and vertical ear canals. On histopathological examination, the most striking feature was the existence of scattered apoptotic-appearing keratinocytes within severely hyperplastic epithelium. For the first time, immunohistochemistry was used to show a closed association between CD3(+) T cells and caspase-3 stained keratinocytes, consistent with a keratinocyte apoptosis by epidermal-infiltrating T cells. Treatment was initiated using topical tacrolimus twice daily and an ear cleanser once daily. A marked improvement was observed after 10 days of treatment and the lesions completely resolved over a period of three weeks. The origin of T cells directed against keratinocytes is currently unknown.

Topics: Animals; Apoptosis; Caspase 3; Cat Diseases; Cats; CD3 Complex; Female; Immunohistochemistry; Immunosuppressive Agents; Keratinocytes; Necrosis; Otitis Externa; T-Lymphocytes; Tacrolimus

Ischemia-reperfusion injury (IRI) in the early posttransplant period affects immediate graft function and late allograft dysfunction. This study determines the influence of pharmacologic preconditioning with a calcineurin inhibitor on IRI in a syngeneic F344 rat kidney transplant model. Donor rats were pretreated with one dose of cyclosporine (10 mg/kg) or tacrolimus (1 mg/kg) administered at 24 hr or 7 days before being subjected to 2 hr of cold ischemia and then transplanted. Pharmacologic preconditioning significantly improved renal function, as assessed by serum creatinine and inulin clearance, and histologic score versus vehicle-treated rats. There were no differences between cyclosporine and tacrolimus in the measured outcomes. This renoprotective effect, although not complete, was seen with only one dose of calcineurin inhibitor, and the effect was sustained for at least 7 days before IRI. This approach may represent a viable pharmacologic intervention to decrease IRI at the time of organ transplantation.

Topics: Animals; Creatinine; Cyclosporine; Immunosuppressive Agents; Inflammation; Kidney Transplantation; Kidney Tubules; Models, Animal; Necrosis; Rats; Rats, Inbred F344; Tacrolimus; Transplantation Conditioning; Transplantation, Homologous; Transplantation, Isogeneic; Treatment Outcome

Previously described neurologic damage induced by immunosuppressive treatments includes transient or reversible central nervous system involvement. We describe a 57-year-old man who underwent liver transplantation and was started on immunosuppressive therapy with tacrolimus (FK506). Six months later, he started complaining of a progressive motor and sensory impairment of the left side, together with cognitive impairment. Brain MRI showed an enlarging lesion of the white matter with peripheral contrast enhancement. PET study indicated severe hypometabolism in the right hemisphere and spectroscopic MRI showed a peak of choline and relative reduction of other metabolites. Findings of CSF examinations and cultures, serology, and molecular techniques were normal. Tacrolimus treatment was stopped. A cerebral biopsy of the lesion showed a sub acute necrotizing process. In the following months, cognitive status of the patient tended to improve although he remained hemiplegic, while serial MRI confirmed the tendency to the recovery of the lesion that was still present 1 year after. The present observation describes a progressive encephalopathy associated with immune suppression with an unusual feature and permanent brain damage.

Topics: Brain; Brain Diseases; Disease Progression; Follow-Up Studies; Hepatitis, Chronic; Humans; Immunosuppressive Agents; Liver Transplantation; Magnetic Resonance Imaging; Magnetic Resonance Spectroscopy; Male; Middle Aged; Necrosis; Positron-Emission Tomography; Tacrolimus; White People

Necrolytic acral erythema is a relatively recently described psoriasis-like skin eruption seen in people infected with hepatitis C virus. Hepatitis C virus infection is endemic in many parts of the world with a steady increase of incidence in Pakistan. Recognition of this disorder is crucial to an early treatment of the underlying liver disease. Herein, we report the first case of necrolytic acral erythema from Asia and also describe good therapeutic response to topical tacrolimus ointment.

Topics: Adolescent; Hepatitis C, Chronic; Humans; Immunosuppressive Agents; Male; Necrosis; Ointments; Skin Diseases, Papulosquamous; Tacrolimus

Drug-eluting stents are widely used to prevent restenosis but are associated with late endothelial damage. To understand the basis for this effect, we have studied the consequences of a prolonged incubation with rapamycin on the viability and functions of endothelial cells.. Human umbilical vein or aorta endothelial cells were exposed to rapamycin in the absence or in the presence of tumour necrosis factor alpha (TNFalpha). After a 24 h-incubation, rapamycin (100 nM) caused a significant cell loss associated with the increase of both apoptosis and necrosis, as quantified by propidium iodide staining, caspase 3 activity, and lactate dehydrogenase release. Rapamycin also impaired cell mobility, as assessed by a wound test, and promoted the formation of actin stress fibres, as determined with confocal microscopy. Moreover, the inhibitor prolonged TNFalpha-dependent E-selectin induction, inhibited endothelial nitric oxide synthase expression at both mRNA (quantitative real-time polymerase chain reaction) and protein level (enzyme-linked immunosorbent assay and western blot), and lowered bioactive nitric oxide output (RFL-6 reporter cell assay). Under the conditions adopted, rapamycin inhibited both mammalian target-of-rapamycin complexes (mTORC1 and mTORC2), as indicated by the reduced amount of raptor and rictor bound to mTOR in immunoprecipitates and by the marked hypophosphorylation of protein S6 kinase I (p70S6K) and Akt, determined by western blotting. The selective inhibition of mTORC1 by AICAR did not affect endothelial viability.. A prolonged treatment with rapamycin impairs endothelial function and hinders cell viability. Endothelial damage seems dependent on mTORC2 inhibition.

Topics: Adaptor Proteins, Signal Transducing; Apoptosis; Blotting, Western; Cardiovascular Agents; Carrier Proteins; Caspase 3; Cell Movement; Cell Survival; Dose-Response Relationship, Drug; E-Selectin; Endothelial Cells; Enzyme-Linked Immunosorbent Assay; Humans; Immunoprecipitation; L-Lactate Dehydrogenase; Mechanistic Target of Rapamycin Complex 1; Microscopy, Confocal; Multiprotein Complexes; Necrosis; Nitric Oxide; Nitric Oxide Synthase Type III; Polymerase Chain Reaction; Protein Kinases; Proteins; Rapamycin-Insensitive Companion of mTOR Protein; Regulatory-Associated Protein of mTOR; Ribosomal Protein S6 Kinases, 70-kDa; Sirolimus; Stress Fibers; Tacrolimus; Theophylline; Time Factors; TOR Serine-Threonine Kinases; Transcription Factors; Tumor Necrosis Factor-alpha; Up-Regulation

Proliferative and necrotizing feline otitis externa is a rare disorder of unknown aetiology. This condition was diagnosed by skin biopsy in three adult domestic shorthair cats (3-5 years old) and one kitten (6 months old). The affected cats had large tan to dark brown-black coalescing plaques covering the concave surface of the pinnae and external ear canals. Friable material from the plaques and a thick exudate occluded the ear canals. The cats had a secondary bacterial and/or yeast otitis. Prior to the histopathological diagnosis, all cats received numerous otic preparations as well as oral antibiotics and corticosteroids without resolution. Histologically, all cases had strikingly similar changes; acanthosis with pronounced hair follicle outer root sheath hyperplasia and neutrophilic luminal folliculitis, follicular keratosis and individually necrotic keratinocytes in the outer root sheath of hair follicles. One case was documented via skin biopsy to have persisted for 4 years. The adult cats were treated with topical 0.1% tacrolimus and all showed marked improvement although one cat was lost to follow up. The lesions completely resolved with topical tacrolimus alone in one cat and topical tacrolimus in addition to oral prednisolone in another cat.

Topics: Administration, Cutaneous; Animals; Cat Diseases; Cats; Diagnosis, Differential; Female; Immunosuppressive Agents; Male; Necrosis; Otitis Externa; Severity of Illness Index; Tacrolimus

Ischemia-reperfusion (I-R) injury is poorly tolerated by fatty livers, most probably secondary to reduced cellular adenosine triphosphate (ATP) levels. We investigated the effectiveness of tacrolimus pretreatment on fatty liver I-R injury in obese Zucker rats. Tacrolimus (0.3 mg/kg, intravenously) was injected 24 hours before a 75-minute ischemic period and rats were sacrificed 6 hours later. Tacrolimus modified the response to I-R observed in obese Zucker rats, when compared to nontreated obese rats: a significant reduction in hepatocyte necrosis was associated with a significant increase in hepatocyte apoptosis. In addition, cell necrosis and apoptosis were significantly and inversely correlated in lean nontreated and treated obese Zucker rats following I-R. Tacrolimus also significantly increased the hepatic ATP levels, reduced in nontreated obese rats, toward values found in lean Zucker rat livers. This protective effect of tacrolimus was further confirmed in vivo by a significantly improved survival following pretreatment with tacrolimus, 24 hours prior to ischemia. In conclusion, in obese Zucker rat livers, tacrolimus pretreatment reversed the I-R injury toward the one found in lean Zucker rats. The correlations between ATP levels and the opposite changes in necrosis and apoptotic pathways strongly suggest a cause-effect relationship between tacrolimus and changes in ATP levels.

Topics: Animals; Biopsy, Needle; Body Weight; Disease Models, Animal; Fatty Liver; Immunohistochemistry; Liver Circulation; Liver Function Tests; Liver Regeneration; Male; Necrosis; Probability; Random Allocation; Rats; Rats, Zucker; Reference Values; Reperfusion Injury; Statistics, Nonparametric; Survival Rate; Tacrolimus; Time Factors

Acute pancreatitis is associated with substantial alterations of the immunologic host response which has been claimed to promote remote organ dysfunction, septic complications, and mortality. Treatment with immunomodulating substances has been subject of few experimental studies with still conflicting results.. We used the taurocholate-induced model of severe acute pancreatitis (SAP) in rats which were assigned to different treatment regimen: isotonic saline (SAP-S) for nontreated controls, recombinant rat interferon-gamma for immunostimulation (SAP-IFN-gamma), and FK506 for immunosuppression (SAP-FK506). Animals were killed after 3, 6, and 24 hours as well as 3 and 7 days, and parameters of local and systemic severity were assessed.. Treatment with IFN-gamma and FK506 attenuated the progression of intrapancreatic necrosis within the first 24 hours after pancreatitis induction along with a substantial reduction of subsequent neutrophil tissue infiltration as shown by decreased myeloperoxidase activity. Enhanced cell death by apoptosis during the postacute course was reduced in FK506-treated animals only. Pancreatic interleukin (IL) 1beta messenger RNA up-regulation occurred early and was slightly suppressed in both treatment groups; tumor necrosis factor alpha (TNF-alpha) and IL-2 messenger RNA expression paralleled the onset of apoptosis and was markedly decreased in IFN-gamma- and FK506-treated rats. The 2 therapeutic regimens had similar effects on intrapancreatic and systemic IL-1beta and TNF-alpha protein release; however, the profiles of both cytokines were differently influenced. Whereas IFN-gamma and FK506 treatment lead to an enhanced intrapancreatic and systemic TNF-alpha protein release during the early course, IL-1beta concentrations were significantly reduced within the late intervals. Seven-day mortality was 44% in saline-, 29% in IFN-gamma-, and 25% in FK506-treated rats (P = not significant).. Severe acute pancreatitis is associated with early alterations of the immune response comprising overt T-cell activation and impaired monocyte/macrophage function alike. Targeting either immunologic derangement improves local pancreatic damage and systemic severity. However, because mortality was not improved, a therapeutic benefit of immunomodulating substances in clinical SAP remains to be defined.

Topics: Adjuvants, Immunologic; Animals; Apoptosis; Disease Models, Animal; Gene Expression Regulation; Immunosuppressive Agents; Interferon-gamma; Interleukin-1beta; Interleukin-2; Male; Necrosis; Pancreas; Pancreatitis, Acute Necrotizing; Rats; Rats, Wistar; RNA, Messenger; Tacrolimus; Taurocholic Acid; Time Factors; Tumor Necrosis Factor-alpha

Ischemia/reperfusion (I/R) carries significant injury to endothelial cells in transplanted organs and is an important factor in chronic rejection. Immunosuppressive drugs, notably cyclosporin A (CyA) and FK506, can potentially augment this injury. Here, our goal was to determine the combined effects of I/R and CyA or FK506 on endothelial cells. Transformed mouse endothelial cells (SVEC 4-10) were subjected to ischemia or I/R for 2-24 hours by incubating cells in 100 per cent N2 (ischemia) followed by 5 per cent CO2 and 95 per cent O2 (reperfusion) for 24 hours. In separate experiments, CyA or FK506 was added to cells subjected to ischemia or I/R. Nonviable cells were determined by Trypan blue exclusion assay. All experiments (done in triplicate) were analyzed by Student's t test. Increasing ischemia times resulted in a greater number of nonviable cells (2% nonviable cells at 0 hours and 57% at 24 hours of I/R). Addition of CyA significantly increased the number of nonviable cells when compared with the control (I/R only) group (P = 0.014). Interestingly, FK506 did not increase the percentage of nonviable cells compared with the control group (P = 0.2). Unlike FK506, CyA augments I/R injury to endothelial cells in vitro. These findings could be relevant in chronic rejection and transplantation.

Topics: Analysis of Variance; Animals; Apoptosis; Calcineurin Inhibitors; Cell Count; Cell Line, Transformed; Cell Survival; Cells, Cultured; Coloring Agents; Cyclosporine; Endothelial Cells; Graft Rejection; Immunosuppressive Agents; Ischemia; Mice; Necrosis; Reperfusion Injury; Tacrolimus; Time Factors; Trypan Blue

The aim of this study was to demonstrate that tacrolimus (FK506) has a hepatoprotective effect by reducing ischemia-reperfusion-induced apoptosis and necrosis, both of which lead to post-surgical liver dysfunction. An ischemia-reperfusion model and primary cultured rat hepatocytes subjected to hypoxic and reoxygenation phases, mimicking the surgical process, were used. c-Jun N-terminal kinase 1/stress-activated protein kinase 1 (JNK1/SAPK1) activation leads to caspase 3 activation, a trigger of apoptosis. The activation status of JNK1/SAPK1 was evaluated by immunoprecipitation or Western-blotting experiments. Apoptosis was assessed by measuring caspase activation and by TUNEL (terminal deoxynucleotidyltransferase-mediated deoxyuridine triphosphate-biotin nick-end labeling) reaction. Necrosis was assessed histologically. Tacrolimus improved the survival rate of rats subjected to ischemia-reperfusion. After FK506 pretreatment, the liver necrosis rate was reduced, and ischemia-reperfusion-induced JNK1/SAPK1 activation and apoptosis were significantly decreased. In hypoxia-reoxygenation-subjected hepatocytes, tacrolimus reduced JNK1/SAPK1 and caspase 3 activation. In the liver, tacrolimus prevented ischemia-reperfusion-induced apoptosis and necrosis.

Topics: Animals; Apoptosis; Blotting, Western; Caspases; Cells, Cultured; Hepatocytes; Immunosuppressive Agents; In Situ Nick-End Labeling; Ischemia; JNK Mitogen-Activated Protein Kinases; Liver; Male; Mitogen-Activated Protein Kinases; Necrosis; Rats; Rats, Wistar; Reperfusion Injury; Tacrolimus

To investigate the effect of combined CsA and FK506 with 5-FU on hepatocellular carcinoma rats.. A syngeneic rat model of hepatocellular carcinoma was used. Control group (A) underwent 4 ml 5% GS. Treatment group was divided into 3 groups namely, group B: only 5-FU and 5% GS; group C: 5-FU, CsA and 5% GS; group D: 5-FU, FK506 and 5%GS. Cell cycle, apoptosis, necrosis and mitochondrial transmembrane potential were measured by flow cytometry, laser scanning confocal microscopy, and electron transmission microscopy. Statistical analysis was performed by SPSS 10.0 for Windows software. Statistical comparisons were made with ANOVA followed by Dunnett's T3 or LSD test.. Compared to the control group, the percentage of apoptotic cells including trifle necrotic cells was significantly higher, and among the treatment group, group D was the highest, and group C was higher than group B. In the treatment group, cell cycle of hepatoma cells was mainly arrested at S phase, but in group D, G0/G1 phase cells were significantly decreased and S phase cells significantly increased. Compared to the control group, mitochondrial transmembrane potential was significantly decreased in the treatment group, among with, group B was the lowest, group C was higher than group D. Morphological changes demonstrated by electron microscopy included dispersed nuclear chromatin, loss of nucleoli, membrane bleeding, cell shrinkage, typical apoptotic bodies and marked swelling of mitochondria in the treatment group. In the control group, however, they were characterized by normal cell ultrastructure.. The present study reveals that 5-FU combined with CsA or FK506 demonstrated a synergistic effect on hepatocellular carcinoma rats. For FK506, the powerful mutual effect is related to the increase of tumor cell's quantity in S phase. Both CsA and FK506 can provide protection on mitochondrial transmembrane potential reduction against hepatoma cells damage from 5-FU.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinoma, Hepatocellular; Cyclosporine; Fluorouracil; Liver Neoplasms; Male; Membrane Potentials; Mitochondria; Necrosis; Rats; Rats, Wistar; Tacrolimus

Topics: Biopsy; Calcineurin Inhibitors; Cyclosporine; Humans; Immunosuppressive Agents; Liver Transplantation; Necrosis; Postoperative Complications; Retrospective Studies; Tacrolimus

Apoptosis, necrosis, and cell proliferation induced by S-(1,2-dichlorovinyl)-L-cysteine (DCVC), the cysteine conjugate of the environmental and occupational contaminant trichloroethylene, were studied in primary cultures of human proximal tubular (hPT) cells. Cells from male and female donors were incubated with a range of concentrations of DCVC (10 to 1000 microM) for up to 48 h, and assessments of cellular morphology (phase-contrast microscopy), necrosis (lactate dehydrogenase (LDH) release), apoptosis(cell cycle analysis, annexin V staining, and caspase activation), and proliferation (cell cycle analysis and DNA synthesis) were made. Time- and concentration-dependent changes in cellular morphology, including elongation of cell shape, formation of intracellular vesicles, and formation of apoptotic bodies, were observed. Significant increases in LDH release occurred in hPT cells incubated with < or =100 microM DCVC for at least 24 h. hPT cells from males were modestly more sensitive to DCVC than those from females, with maximal LDH release of 78 and 65% in cells from males and females, respectively. Flow cytometry analysis of propidium iodide-stained and DCVC-treated hPT cells showed that apoptosis occurred at markedly lower concentrations (10 microM) and at much earlier incubation times (2 h) than necrosis. A small increase was also noted in the percentage of cells in S-phase after a 4-h treatment with as little as 10 microM DCVC, suggesting that cell proliferation was stimulated. This was supported further by increased DNA synthesis. These results show that DCVC causes apoptosis and enhances cell proliferation in hPT cells at environmentally relevant doses and at earlier time points and lower concentrations than necrosis.

Topics: Adult; Aged; Annexin A5; Apoptosis; Caspases; Cell Cycle; Cell Division; Cell Survival; Cells, Cultured; Cysteine; DNA; Dose-Response Relationship, Drug; Female; Flow Cytometry; Humans; Kidney Tubules, Proximal; L-Lactate Dehydrogenase; Male; Methimazole; Microscopy, Confocal; Middle Aged; Necrosis; Tacrolimus; Time Factors

We report three patients in whom neurologic symptoms and cortical laminar necrosis developed after immunosuppressive treatment (cyclosporin A and FK 506) and polychemotherapy (vincristine and methotrexate). Initial neuroradiologic studies showed cortical and white matter involvement. Follow-up studies showed cortical hyper-intense lesions on T1-weighted MR images, consistent with cortical laminar necrosis. The clinical and radiologic data indicate that a transient hypoxic-ischemic process could have been responsible for the encephalic lesions in these three patients.

Topics: Adolescent; Adult; Antineoplastic Agents; Brain Diseases; Cerebral Cortex; Cyclosporine; Female; Humans; Immunosuppressive Agents; Magnetic Resonance Imaging; Male; Methotrexate; Middle Aged; Necrosis; Tacrolimus; Tomography, X-Ray Computed; Vincristine

Ceramide is a key mediator of apoptosis during the cellular stress response which is also involved in stroke-induced death. Transient occlusion of the middle cerebral artery (MCA) in rats led to a strong generation of ceramide as measured in thalamus and entorhinal cortex of the ischemic brain tissue. Enhanced levels of ceramide may be involved in apoptosis signaling following stroke since exogenously added synthetic C2-ceramide increased expression of c-jun and the death-inducing ligands (DILs) CD95-L, TRAIL and TNF-alpha in neuroblastoma cells. DILs in turn mediated death via binding to their respective receptors as concluded from diminished apoptosis upon blocking of the common pathway by dominant negative FADD. C2-ceramide induced both necrosis and apoptosis in a concentration-dependent manner corresponding to the situation present in the ischemic brain. The immunosuppressant FK506 inhibited the release of ceramide, expression of CD95-L and apoptosis in an in vitro and in vivo model for ischemia/reperfusion. These data suggest that ceramide is a crucial initiator of death, e.g., by induction of DILs following stroke.

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Brain Chemistry; Cerebrovascular Disorders; Enzyme Inhibitors; Fas Ligand Protein; Gene Expression; Humans; Immunosuppressive Agents; Ischemic Attack, Transient; Male; Membrane Glycoproteins; Necrosis; Neuroblastoma; Neurons; Proto-Oncogene Proteins c-jun; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Sphingosine; Tacrolimus; TNF-Related Apoptosis-Inducing Ligand; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha; Up-Regulation

Programmed cell death in animals is usually associated with apoptotic morphology and requires caspase activation. Necrosis and caspase-independent cell death have been reported, but mostly in experimental conditions that lead some to question their existence it in vivo. Loss of interdigital cells in the mouse embryo, a paradigm of cell death during development [1], is known to include an apoptotic [2] and caspase-dependent [3] [4] mechanism. Here, we report that, when caspase activity was inhibited using drugs or when apoptosis was prevented genetically (using Hammertoe mutant mice, or mice homozygous for a mutation in the gene encoding APAF-1, a caspase-activating adaptor protein), interdigital cell death still occurred. This cell death was negative for the terminal-deoxynucleotidyl-mediated dUTP nick end-labelling (TUNEL) assay and there was no overall cell condensation. At the electron microscopy level, peculiar 'mottled' chromatin alterations and marked mitochondrial and membrane lesions, suggestive of classical necrotic cell death, were observed with no detectable phagocytosis and no local inflammatory response. Thus, in this developmental context, although caspase activity confers cell death with an apoptotic morphotype, in the absence of caspase activity an underlying mechanism independent of known caspases can also confer cell death, but with a necrotic morphotype. This cell death can go undetected when using apoptosis-specific methodology, and cannot be blocked by agents that act on caspases.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptotic Protease-Activating Factor 1; Bone Morphogenetic Proteins; Caspase Inhibitors; Caspases; Chromatin; Cysteine Proteinase Inhibitors; Embryonic and Fetal Development; Fetal Proteins; Hindlimb; In Situ Nick-End Labeling; Mice; Mice, Knockout; Mice, Mutant Strains; Morphogenesis; Necrosis; Organelles; Proteins; Receptors, Growth Factor; Signal Transduction; Tacrolimus

Myoblast transplantation is a potential treatment for Duchenne muscular dystrophy. One of the problems possibly responsible for the limited success of clinical trials is the rapid death of the myoblasts after transplantation. To investigate this problem, myoblasts expressing beta-galactosidase were injected in the tibialis anterior muscles of mice. Beta-galactosidase activity was reduced by 74.7% after 3 days. Myoblast death observed at 3 days was reduced to 57.2% when the hosts were irradiated. This result suggested that host cells were contributing to this phenomenon. Transplantation in SCID and FK506-treated mice did not reduce cell death, indicating that mortality was not due to an acute specific reaction. In contrast, administration of the anti-LFA-1 (TIB-213) mAb markedly reduced myoblast death at 3 days without altering leukocyte tissue infiltration. We postulated that neutrophils were mediating myoblast mortality by an LFA-1-dependent mechanism. To test this hypothesis, IL-1beta-activated myoblasts were loaded with 6-carboxy-2',7'-dichlorodihydrofluorescein diacetate, di(acetoxymethylester) (DCFH), a marker for oxidative stress. Addition of neutrophils and zymosan-activated serum resulted in a time-dependent DCFH fluorescence; this neutrophil-induced oxidation was considerably inhibited by TIB-213. These results indicate that an effective control of the inflammatory reaction will be necessary for any new clinical trials of myoblast transplantation and suggest that neutrophil-mediated myoblast injury occurs by an LFA-1-dependent pathway.

Topics: Animals; Anti-Inflammatory Agents; Anti-Inflammatory Agents, Non-Steroidal; Antibodies, Monoclonal; Apoptosis; beta-Galactosidase; Biomarkers; Cell Adhesion; Cell Death; Cell Line, Transformed; Fluoresceins; Genes, Reporter; Graft Rejection; Immunosuppressive Agents; Inflammation; Interleukin-1; Lymphocyte Function-Associated Antigen-1; Macrophage-1 Antigen; Methylprednisolone; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred mdx; Mice, SCID; Muscle, Skeletal; Muscular Dystrophy, Animal; Naproxen; Necrosis; Neutrophils; Oxidative Stress; Peroxidase; Piroxicam; Radiation Chimera; Stem Cell Transplantation; Tacrolimus; Whole-Body Irradiation; Zymosan

One-hundred and fourteen limb transplantations have been performed across a major histoincompatibility barrier between donor ACI (RT1a) and recipient Lewis (RT1l) rats immunosuppressed with various dosages of FK-506 and cyclosporine. Three-hundred and thirty biopsy specimens from 64 animals have been evaluated histologically for signs of rejection. A new histologic grading system is introduced to classify the process of rejection in the component tissues (skin, muscle, bone, and articular cartilage) of a limb allograft. The results indicate that FK-506 is a more potent immunosuppressive agent than cyclosporine in preventing the rejection of the skin component of a limb transplant. With twice-weekly intermittent immunosuppression with FK-506, the rejection of muscle, bone, and cartilage can be prevented for an indefinite time, although all long-term surviving animals died at around 300 days, probably of graft-versus-host disease. Based on the histologic stages of rejection in the different tissues at the same point in time, it is evident that each component tissue of a limb transplant rejects over a different time period. This probably reflects a hierarchy of antigenicity, with skin being most antigenic, muscle being intermediate in antigenicity, and bone and cartilage being least antigenic. Although this grading system is not the ultimate solution, it may allow a more objective comparison of experimental limb transplantation in the future.

Topics: Analysis of Variance; Animals; Biopsy; Bone and Bones; Cartilage, Articular; Cyclosporine; Drug Administration Schedule; Epidermis; Fibrosis; Graft Rejection; Hindlimb; Immunosuppression Therapy; Muscle, Skeletal; Necrosis; Pneumonia, Bacterial; Rats; Rats, Inbred ACI; Rats, Inbred Lew; Tacrolimus

Topics: Animals; Animals, Newborn; Cyclosporine; Edema; Female; Graft Rejection; Graft Survival; Guanidines; Heart Transplantation; Hemorrhage; Immunosuppressive Agents; Male; Methotrexate; Necrosis; Papio; Splenectomy; Swine; Tacrolimus; Time Factors; Transplantation, Heterologous

We have previously reported that glutamate can trigger a succession of necrosis and apoptosis in cerebellar granule cells (CGC). Since specific blockers of the N-methyl-D-aspartate (NMDA) receptor channel prevented both types of cell death, the role of Ca2+-dependent processes in the initiation of glutamate toxicity was further investigated. We examined the possible involvement of mitochondria and the role of the Ca2+/calmodulin-regulated protein phosphatase, calcineurin, in the development of either type of cell death. Cyclosporin A and the more selective calcineurin inhibitor, FK-506, prevented the development of both early necrosis and delayed apoptosis. In addition, cyclosporin A prevented the collapse of mitochondrial membrane potential observed during the exposure to glutamate and the concomitant necrotic phase. When CsA was added immediately after glutamate removal, it also prevented delayed apoptosis of neurons that had survived the necrotic phase. Altogether, these results suggest the involvement of calcineurin and a role for mitochondrial deenergization as early signals in neuronal apoptosis induced by glutamate.

Topics: Animals; Apoptosis; Calcineurin; Calmodulin-Binding Proteins; Cells, Cultured; Cerebellum; Cyclosporine; Glutamic Acid; Membrane Potentials; Mitochondria; Necrosis; Neurons; Phosphoprotein Phosphatases; Rats; Tacrolimus

To use mice to examine the effects of cyclosporine and tacrolimus (FK 506) on two forms of acute pancreatitis often seen after clinical organ transplantation.. In the first experiment, male CD-1 mice received cyclosporine (10 mg/kg), tacrolimus (0.32 mg/kg), or saline solution (control) subcutaneously once a day for 10 days. On the 11th day, acute edematous pancreatitis was induced by ceruletide (cerulein). In the second experiment, female ICR mice were fed with a choline-deficient, ethionine-supplemented (CDE) diet for 72 hours to induce necrotizing pancreatitis. After 30 hours on the CDE diet, the mice received cyclosporine (10 mg/kg), tacrolimus (0.32 mg/kg), or saline solution (control) subcutaneously twice daily for 3 days.. The pancreatic dry-to-wet weight ratios after ceruletide injections significantly decreased in mice treated with cyclosporine but did not with tacrolimus. Cyclosporine also significantly increased serum amylase levels, but tacrolimus did not. Cyclosporine or tacrolimus alone did not produce pancreatitis. In the CDE diet groups there was a significant difference in survival among the cyclosporine-treated, the tacrolimus-treated, and the control groups.. Cyclosporine or tacrolimus given alone does not induce acute pancreatitis. In contrast, cyclosporine can adversely affect the course of acute edematous pancreatitis, and both immunosuppressants may worsen the survival of mice with acute hemorrhagic necrotizing pancreatitis. This study also demonstrated that the deteriorating effect of tacrolimus is less potent than that of cyclosporine.

Topics: Acute Disease; Animals; Ceruletide; Choline Deficiency; Cyclosporine; Edema; Ethionine; Female; Male; Mice; Necrosis; Pancreatitis; Tacrolimus

Topics: Animals; Dose-Response Relationship, Drug; Heart; Heart Ventricles; Injections, Intravenous; Male; Myocardium; Necrosis; Rabbits; Tacrolimus

Topics: Animals; Digestive System; Dogs; Female; Heart; Hypertrophy; Immunosuppressive Agents; Kidney Transplantation; Mycophenolic Acid; Myocardium; Necrosis; Polyenes; Sirolimus; Tacrolimus; Transplantation, Homologous

The authors established a new experimental model of fulminant hepatic failure (FHF) with prolonged hepatocellular necrosis and impaired liver regeneration, and evaluated the immunological mechanisms related to the impaired liver regeneration in this model. A novel lipid A analogue, FS-112, was injected intravenously into male Balb/c mice, followed by a 70% partial hepatectomy 2 days later. Serum levels of T.Bil. and ALT rose 7 days after the partial hepatectomy, as compared with controls. In mice pretreated with FS-112, labeling indices of both BrdU and PCNA 36 hrs after the partial hepatectomy were significantly lower than those in the controls. Splenic lymphocytes harvested from the FHF mice 1-5 days after the partial hepatectomy showed a cytotoxic activity against regenerating hepatocytes with a peak effect on day 5. Cytotoxic activity against YAC-1 cells was also found up to 5 days after the partial hepatectomy, and resembled that directed against the regenerating hepatocytes. On the 5th day of FS-112 administration, there was a marked rise in the production of IFN-gamma from splenocytes. When FK-506, an immunosuppressive agent, was given intracutaneously daily for 7 days, serum levels of T.Bil. and ALT significantly decreased, as compared with controls. Furthermore, the PCNA-labeling index 36 hrs after the partial hepatectomy was enhanced by the administration with FK-506 in the FHF mice. These results strongly suggest that the NK cells activated by IFN-gamma may be involved in killing the regenerating liver cells, and thus play a role in the pathogenesis of the impaired liver regeneration in FHF.2+ recovery from the impaired liver regeneration in FHF.

Topics: Animals; Cytotoxicity, Immunologic; Hepatic Encephalopathy; Interferon-gamma; Killer Cells, Natural; Liver; Liver Regeneration; Male; Mice; Mice, Inbred BALB C; Necrosis; Spleen; Tacrolimus

FK506, a newly developed immunosuppressive agent, has recently been used for human liver and kidney transplantation. The present study was carried out to assess the functional and morphological changes by acute or chronic administration of FK506 in heminephrectomized rats. FK506 was given intravenously at the dose of 0.384 mg/kg/hr for 90 min in acute experiment. FK506 was administered by gastric gavage at doses of 1, 2.5, 5 mg/kg for 21 days. Blood and urinary biochemistry were monitored every week. Inulin and PAH clearance studies were conducted during the infusion of FK506 for acute study, and at day 21 for chronic study. Some of the rats were treated with diltiazem (Dilt), captopril or prazosine for 21 days to prevent FK506 nephrotoxicity. Acute infusion of FK506 did not change renal and systemic hemodynamics. Creatinine clearance showed a dose dependent decrease by 10-20% in the rats with chronic administration of FK506. Inulin/PAH clearance indicated a decrease in glomerular filtration rate and renal plasma flow with an increase in renal vascular resistance. The renal histology showed vacuolization in proximal tubuli and media of smooth muscle cells of arterioles. The administration of Dilt functionally and morphologically improved renal impairment induced by FK506. In conclusion, FK506 induces a dose dependent decrease in renal function with significant histological changes in tubuli and arterioles in rat kidney. Intracellular calcium deregulation seems to be involved in FK506-induced nephrotoxicity.

Topics: Animals; Calcium; Captopril; Diltiazem; Hemodynamics; Kidney; Male; Necrosis; Prazosin; Rats; Rats, Inbred Strains; Tacrolimus

Topics: Animals; Anti-Bacterial Agents; Cyclosporins; Cytotoxicity, Immunologic; Graft Rejection; Graft Survival; Hindlimb; Immunosuppressive Agents; Lymphocyte Culture Test, Mixed; Male; Necrosis; Rats; Rats, Inbred Lew; Rats, Inbred Strains; Skin; Tacrolimus; Transplantation, Homologous

The effect of FK 506 on regeneration of the liver was studied in rats after a two-thirds partial hepatectomy after 60 min of ischemia of the unresected liver. The animals were divided into three distinct groups of 10 rats each. Group 1 (controls) received 0.5 ml saline solution intravenously 30 min after the induction of ischemia. Groups 2 and 3 were injected with FK 506 (0.3 mg/kg) intravenously 30 min after and 24 min before the induction of hepatic ischemia, respectively. The hepatic content of ATP and serum levels of ALT and lactate dehydrogenase were determined on each animal. In addition, the histological appearance and mitotic activity of the remnant liver was determined at regular 24-hr intervals after hepatic ischemia. All 10 control animals died within 72 hr. Treatment with FK 506 resulted in improved survival in groups 2 and 3 (30% and 80%, respectively). The improved survival seen in the FK 506-treated animals was reflected by a restoration of hepatic ATP content, a reduction in the serum levels of ALT and lactate dehydrogenase, an amelioration of hepatic necrosis and neutrophilic infiltration and an increase in the mitotic activity of the liver. These results suggest that FK 506 ameliorates the hepatic injury associated with ischemia/reperfusion and has a potent stimulatory effect on liver cell regeneration that may make it valuable as a hepatoprotective agent when administered to organ donors before graft harvesting.

Topics: Adenosine Triphosphate; Alanine Transaminase; Animals; Anti-Bacterial Agents; Immunosuppressive Agents; L-Lactate Dehydrogenase; Liver; Liver Diseases; Liver Regeneration; Male; Mitosis; Necrosis; Rats; Rats, Inbred Lew; Reperfusion Injury; Tacrolimus

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Galactosamine; Hepatitis, Animal; Liver; Liver Function Tests; Male; Necrosis; Rats; Rats, Inbred Strains; Tacrolimus; Time Factors

Topics: Child, Preschool; Graft Rejection; Humans; Liver Transplantation; Necrosis; Pilot Projects; Tacrolimus; Transplantation, Homologous; Treatment Outcome

Topics: Adult; Humans; Male; Necrosis; Pyoderma; Skin Ulcer; Tacrolimus