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Sample records for cyclosporine-a

  1. Cyclosporine A-Induced Renal Fibrosis

    PubMed Central

    Slattery, Craig; Campbell, Eric; McMorrow, Tara; Ryan, Michael P.

    2005-01-01

    Cyclosporine A, which has been the foremost immunosuppressive agent since the early 1980’s, significantly improves the success of organ transplantation. However, common complications of cyclosporine A therapy, such as severe renal tubulointerstitial fibrosis, limit the drug’s clinical use. Although the exact mechanisms driving cyclosporine A-induced tubulointerstitial fibrosis remain elusive, we hypothesized that epithelial-mesenchymal transition (EMT) may play a major role. We investigated this in vitro by treating human proximal tubular cells with cyclosporine A. Morphological changes were observed after cyclosporine A treatment, including cell elongation (with a large degree of detachment), cytoskeletal rearrangement, and junctional disruption. In addition, expression of the myofibroblast-specific marker α-smooth muscle actin was detected in treated cells. These observations are consistent with events described during EMT. Using Affymetrix gene microarrays, we identified 128 genes that were differentially regulated in renal tubular cells after cyclosporine A treatment, including known profibrotic factors, oncogenes, and transcriptional regulators. Cyclosporine A induced a dose-dependent increase in transforming growth factor-β secretion from proximal tubular cells. Subsequent functional studies revealed that protein kinase C-β isoforms play a key role in cyclosporine A-induced effects. These findings provide novel insights into cyclosporine A-induced renal fibrosis and the molecular mechanisms underlying EMT, events that may be relevant in other disease states. PMID:16049326

  2. Cyclosporin A in cadaveric organ transplantation.

    PubMed Central

    Calne, R Y; White, D J; Evans, D B; Thiru, S; Henderson, R G; Hamilton, D V; Rolles, K; McMaster, P; Duffy, T J; MacDougall, B R; Williams, R

    1981-01-01

    The use of cyclosporin A (CyA) with a protocol designed to avoid the effects of nephrotoxicity resulted in a one-year survival of 86% in recipients of renal allografts from unmatched cadaveric donors. The drug also controlled rejection of liver and pancreatic allografts. It was possible to change patients initially treated with CyA to azathioprine and corticosteroids and vice versa, thus enlarging the potential value of CyA in organ allografting. Of 34 recipients of renal allografts, 29 were currently receiving only CyA as immunosuppressive treatment. Twelve patients never required any adjuvant steroid treatment. These results suggest that CyA is an effective immunosuppressant, and if used with care side effects need not be severe. PMID:6781658

  3. Cyclosporin A inhibits DNA synthesis by epidermal Langerhans cells.

    PubMed

    Haftek, M; Urabe, A; Kanitakis, J; Dusserre, N; Thivolet, J

    Cyclosporin A, a potent immunosuppressive drug currently used in organ transplant recipients, has been shown to exert in vitro a direct antiproliferative effect on a number of cell types present in the skin, including keratinocytes, fibroblasts, and endothelial cells. Although in vitro studies suggest that cyclosporin A may interfere with the functional capacities of epidermal Langerhans cells, there is no evidence that the treatment influences the distribution or number of Langerhans cells in vivo. We used a model of normal human skin graft to "nude" mice, which is free of the human systemic control mechanisms, for studies on the DNA synthesis of human Langerhans cells under the influence of cyclosporin A. The grafted animals were given daily subcutaneous (50 mg/kg) or intraperitoneal (5, 12.5, and 25 mg/kg) drug injections during three weeks, which resulted in mean blood levels comparable to those observed in treated patients with organ transplants or psoriasis, respectively. BrdU administered during the last week of the experiment was incorporated by all cells synthesizing DNA, including those passing through S-phase. Langerhans cells were detected on deparaffinized or frozen tissue sections of xenografts with anti-CD1a and anti-HLA DR monoclonal antibodies, and the number of BrdU-positive cells was determined by double labeling. Our results indicate that the Langerhans cell DNA synthesis is impaired by therapeutic levels of cyclosporin A.

  4. [Pyoderma gangrenosum--positive effect of cyclosporin A therapy ].

    PubMed

    Krauze, Ewa; Lis, Anna; Kamińska-Budzińska, Grazyna; Wygledowska-Kania, Mariola; Pierzchała, Ewa; Brzezińska-Wcisło, Ligia

    2002-10-01

    Although pyoderma gangrenosum is a disorder known since over 70 years, it still remains a diagnostic and therapeutic problem. We describe three subjects with pyoderma gangrenosum; two were females, one was male, one case was associated with colitis ulcerosa, two were without any related disorders. Histopathologic examinations supported the diagnosis in all cases. In spite of intensive topical and systemic treatment with corticosteroids, Dapsone, Clofazimine, no sufficient effects were achieved. Cyclosporin A introduced in the dose of 5 mg/kg/d resulted in dramatic response and complete remission. Serum CyA levels, biochemical parameters of liver and kidney function, blood pressure were monitored during the therapy. No adverse events due to Cyclosporin A were observed.

  5. Chitosan functionalized nanocochleates for enhanced oral absorption of cyclosporine A

    PubMed Central

    Liu, Min; Zhong, Xiaoming; Yang, Zhiwen

    2017-01-01

    It remains a significant challenge to overcome the poor permeability of cyclosporine A and enhance its oral absorption. In this study, we have identified a positively charged chitosan that is able to induce coiling up of anionic lipids to form nanocochleates with an average size of 114.2 ± 0.8 nm, without the need for calcium ions. These functional chitosan-induced nanocochleates enhanced gastrointestinal absorption of cyclosporine A, up to a 3-fold increase in oral bioavailability. A fluorescence-labeling study confirmed that absorption mainly occurred in the duodenum and jejunum. Transport studies indicated that uptake of chitosan-induced nanocochleates by Caco-2 cells was by clathrin- and caveolae-mediated endocytosis, but not by macropinocytosis. Furthermore, three cellular tight junction proteins, ZO-1, F-actin and claudin-4, were significantly down-regulated, suggesting that chitobiose-induced nanocochleates are able to reconstruct and open tight junctions in intestinal epithelial cells to enhance drug absorption. In summary, these novel bifunctional chitosan-induced nanocochleates appear to have potential to facilitate oral delivery of cyclosporine A. PMID:28112262

  6. Cyclosporin A and multiple fibroadenomas of the breast.

    PubMed

    Baildam, A D; Higgins, R M; Hurley, E; Furlong, A; Walls, J; Venning, M C; Ackrill, P; Mansel, R E

    1996-12-01

    Multiple bilateral fibroadenomas are uncommon. This finding in four women who had received renal transplants prompted further inquiry. A prospective study was performed on 39 women under the age of 55 years who had received a renal transplant at least 1 year earlier. Clinical examination and breast ultrasonography were performed. Factors considered included immunosuppressive therapy, concurrent medication and renal function. Blood was taken for estimation of oestradiol, prolactin, follicle-stimulating hormone (FSH) and sex hormone binding globulin levels. Fibroadenomas were found in 13 of 29 women who had received cyclosporin A: multiple in ten and bilateral in five. No abnormal breast findings were seen in 10 patients immunosuppressed with steroids and azathioprine alone (chi 2 = 7.30, 1 d.f., P < 0.01). Serum oestradiol concentration was raised in women with fibroadenomas compared with that in those with normal breasts (P < 0.05) and the level of FSH was lower (P < 0.01). Cyclosporin A may act on breast fibroblasts by humoral mechanisms and direct action.

  7. Cutaneous malignant melanomas occurring under cyclosporin A therapy: a report of two cases.

    PubMed

    Mérot, Y; Miescher, P A; Balsiger, F; Magnenat, P; Frenk, E

    1990-08-01

    Two patients are reported with cutaneous malignant melanoma who had been on treatment with cyclosporin A. The first case was a 44-year-old man with systemic sclerosis and the second a 52-year-old woman who had a renal transplant. In both cases cyclosporin A was administered with a low dose of prednisone.

  8. Formulation and evaluation of Cyclosporin A emulgel for ocular delivery.

    PubMed

    Shen, Yan; Ling, Xiang; Jiang, Weiwei; Du, Shuang; Lu, Yang; Tu, Jiasheng

    2015-01-01

    Emulgels have been extensively covered as a promising drug delivery system for the administration of lipophilic drugs. This work was conducted to develop an emulgel formulation for Cyclosporin A (CsA) employing polycarbophil as the gelling agent for ocular delivery. The prepared emulgels were evaluated for their physical appearance, rheological behavior, drug release, stability, precorneal clearance and irritation. Results showed that CsA emulgel formulations prepared with polycarbophil exhibited acceptable physical properties and drug release, which remained consistent after storage for 3 months. A prolonged retention time was also observed on the ocular surface with improved ocular bioavailability and no irritation. Therefore, the polycarbophil-based emulgel could be exploited as a potential hydrophobic drug carrier for topical ocular drug delivery.

  9. Conformational Heterogeneity of Cyclosporin A in Cyclophilin 18 Binding

    PubMed Central

    Lin, Weilin; Quintero, Andres; Zhang, Yixin

    2016-01-01

    The immunosuppressive drug cyclosporin A (CsA) binds to its receptor protein cyclophilin 18 (Cyp18) in two distinct kinetic phases, while the mechanism remains elusive. Stopped-flow measurements coupled with titration and competition experiments were used to investigate the puzzling two-phase process of CsA and Cyp18 interaction. This study leads to the dissection of different conformational fractions of either direct fast binding or slow binding with rate-limiting conformational inter-conversion and the real-time measurement of kon value (8.34 ± 0.22 x106 M-1s-1) in solution. Furthermore, our study indicates that the structure of CsA during dissociation from the protein possesses a distribution of conformations different from those in solution under equilibrium condition. PMID:27082870

  10. Cyclosporin A acute encephalopathy and seizure syndrome in childhood: clinical features and risk of seizure recurrence.

    PubMed

    Gleeson, J G; duPlessis, A J; Barnes, P D; Riviello, J J

    1998-07-01

    Cyclosporin A is associated with an acute encephalopathy including seizures and alterations in mental status, herein referred to as cyclosporin A acute encephalopathy and seizure syndrome. The clinical history, electroencephalogram (EEG), and neuroimaging findings in 19 children with cyclosporin A acute encephalopathy and seizure syndrome over a 10-year period were reviewed in order to delineate clinical characteristics, imaging features, and to determine the risk of seizure recurrence in this population. All 19 had motor seizures associated with other features of cortical and subcortical dysfunction. The acute mean cyclosporin A level was 342 microg/L, but was within the "therapeutic" range in five cases. Brain imaging by computed tomography (CT) or magnetic resonance imaging (MRI) in the acute or subacute phase revealed lesions characteristic of cyclosporin A toxicity in 14 cases. Acute EEG abnormalities were present in all and included epileptiform discharges or focal slowing. Patients were followed for a median of 49 months (1-9 years). Follow-up imaging (n = 10) showed lesion resolution or improvement in the majority while EEG (n = 10) had normalized in only three. Seizures recurred in six patients and only in those with persistent EEG or imaging abnormalities. No patient had a second episode of cyclosporin A associated neurotoxicity or seizure. It appears that a significant risk of seizure recurrence exists following cyclosporin A acute encephalopathy and seizure syndrome and primarily in those children with persistent EEG or imaging abnormalities.

  11. Investigation into the potential of low-frequency ultrasound facilitated topical delivery of Cyclosporin A.

    PubMed

    Liu, Hongzhuo; Li, Sanming; Pan, Weisan; Wang, Yongjun; Han, Fei; Yao, Huimin

    2006-12-01

    The potential for low-frequency ultrasound facilitated topical transport of Cyclosporin A was investigated using rat skin. Studies of intensity and exposure time acting on the deposition of Cyclosporin A into deeper skin of in vitro sonophoresis were performed. Low-frequency ultrasound increased the amount of Cyclosporin A retained in the skin only seven times than the passive diffusion. Furthermore, we also tested the synergistic effect of ultrasound and other approaches such as chemical enhancers and electroporation on topical drug delivery of Cyclosporin A. We found that the efficacy of low-frequency ultrasound in enhancing topical delivery could be further increased by pretreatment of skin with chemical enhancers, such as laurocapram (Azone) and sodium lauryl sulfate (SLS). Meanwhile only a small amount was seen to across the full skin into the receiver compartment. Trimodality treatment comprising of pretreatment with Azone+ultrasound in combination followed by electroporation was not effective in enhancing the topical delivery of Cyclosporin A. However, this combination strategy increased the penetration of Cyclosporin A through rat skin by order of 15. The histopathological findings revealed that there was almost no change observed in the structure of skin after ultrasound or combination with ultrasound and enhancers as compared with the control group. In general, the enhanced skin accumulation of Cyclosporin A by the combination of low-frequency ultrasound and chemical enhancers could help significantly to optimize the targeting of the drug without of a concomitant increase of the systemic side effects.

  12. Cyclosporin A reduces canalicular membrane fluidity and regulates transporter function in rats.

    PubMed Central

    Yasumiba, S; Tazuma, S; Ochi, H; Chayama, K; Kajiyama, G

    2001-01-01

    Changes of the biliary canalicular membrane lipid content can affect membrane fluidity and biliary lipid secretion in rats. The immunosuppressant cyclosporin A is known to cause intrahepatic cholestasis. This study investigated whether cyclosporin A influenced canalicular membrane fluidity by altering membrane phospholipids or transporter expression. In male Sprague-Dawley rats, a bile-duct cannula was inserted to collect bile, and sodium taurocholate was infused (100 nmol/min per 100 g) for 60 min. During steady-state taurocholate infusion, cyclosporin A (20 mg/kg) or vehicle was injected intravenously and then bile was collected for 80 min. After killing the rats, canalicular membrane vesicles were prepared. Expression of canalicular membrane transporters was assessed by Western blotting and canalicular membrane vesicle fluidity was estimated by fluorescence polarization. Cyclosporin A reduced biliary lipid secretion along with a disproportionate reduction of lipids relative to bile acids. Cyclosporin A significantly decreased canalicular membrane fluidity along with an increase of the cholesterol/phospholipid molar ratio. Only expression of the transporter P-glycoprotein was increased by cyclosporin A. Because canalicular membrane transporter expression was largely unchanged by cyclosporin A despite a marked decrease of biliary lipid secretion, transporter activity may partly depend upon canalicular membrane fluidity. PMID:11237863

  13. Ocular penetration of cyclosporin A. III: The human eye.

    PubMed Central

    BenEzra, D; Maftzir, G; de Courten, C; Timonen, P

    1990-01-01

    The distribution of cyclosporin A (CsA) in the blood, saliva, tears, aqueous humour, vitreous, and cerebrospinal fluid has been studied after oral treatment with 5 mg/kg/day of CsA or application of 2% CsA eye drops in olive oil solution. After oral treatment all patients had high CsA levels in blood. Measurable levels of CsA were also found in the saliva and tears. Patients without any intraocular inflammation or patients with mild uveitis did not have any detectable CsA in the aqueous humour. However, patients with severe uveitis had significant levels of CsA in the aqueous humour and in the vitreous. No CsA was found in the cerebrospinal fluid of two patients with central nervous system manifestations of Behçet's disease. After local treatment with 2% CsA eye drops no detectable levels of CsA were found in the blood, the saliva, the aqueous humour, or the vitreous even in patients with severe uveitis. PMID:2378841

  14. Cyclosporin a. Inhibition of experimental autoimmune uveitis in Lewis rats.

    PubMed Central

    Nussenblatt, R B; Rodrigues, M M; Wacker, W B; Cevario, S J; Salinas-Carmona, M C; Gery, I

    1981-01-01

    Cyclosporin A (CS-A), a selective inhibitor of T lymphocytes, is reported here to prevent S antigen (S-Ag) induced uveitis in Lewis rats. The S-Ag, found in all mammalian retinas, is uveitogenic under experimental conditions and patients with certain uveitic entities demonstrate cell mediated responses to this antigen. Daily treatment with CS-A (10 mg/kg) begun on the same day as S-Ag immunization totally inhibited the development of the uveitis in this experimental autoimmune model. Moreover a greater CS-A dose (40 mg/kg) efficiently prevented the disease process when therapy was started 7 d after S-Ag immunization. Anti-S-Ag antibody titers were observed to be similar in rats either protected or not protected with CS-A. Our data support strongly the need for T cell participation in this disease model. Since ocular inflammatory disease is an important cause of visual impairment, the data further suggest that CS-A may be useful in the treatment of patients with intractable uveitis. Images PMID:7204576

  15. Paradoxical effects of cyclosporin A on collagen arthritis in rats

    PubMed Central

    1983-01-01

    The effect of the immunosuppressive agent cyclosporin A (CS-A) on collagen arthritis in Sprague-Dawley rats is investigated. A 14-d course of CS-A treatment at doses of 15 mg/kg per day or more, begun on the same day as type II collagen immunization, suppressed the development of arthritis as well as humoral and delayed-type hypersensitivity (DTH) skin test responses to type II collagen, possibly by interfering with helper T cells. Additional studies demonstrated that CS-A treatment only during the induction phase of immunity proved to be successful. When CS-A treatment was started only during the immediately preclinical phase of arthritis or after the disease onset, a significant enhancement of the disease was obtained in a dose-dependent manner. This enhancement was accompanied by an augmentation of DTH skin reactions, while antibody responses were either suppressed or unaffected. These results appear to be attributable at least in part to a suppressive effect of CS-A on a population of suppressor T cells, thus resulting in a T cell-mediated helper effect. It is therefore reasonable to assume that the paradoxical effects of CS-A on collagen arthritis in rats might be caused by an altering of the sensitive balance of the two regulatory subpopulations of T cells. It is also possible that cell-mediated immune responses may play an important role in influencing the course of the disease. PMID:6644238

  16. The use of cyclosporin A in clinical organ grafting.

    PubMed Central

    Calne, R Y; White, D J

    1982-01-01

    Experiments in animals with organ allografts showed that Cyclosporin A (CyA) was an extremely powerful immunosuppressant with a good therapeutic index. A pilot study of the drug in human recipients of renal allografts revealed an unexpected side effect, nephrotoxicity, which made care of patients difficult. Following a policy of deliberate hydration of patients in the perioperative phase and withholding CyA until diuresis was occurring in the graft, excellent results have been obtained in clinical practice. An 82% actuarial functional survival at both one and two years has been obtained in the 59 patients treated with this protocol. A multicenter trial is now in progress in eight centers in Europe, comparing CyA used in the manner described above with conventional azathioprine and steroids. CyA has also been used in 17 recipients of liver allografts, ten of whom are still alive and 11 recipients of segmental pancreatic allografts, one of whom remains off insulin after two and a quarter years. Sudden graft failure occurred between three months and two years in three patients whose pancreatic duct had been occluded. The authors' most recent segmental pancreas graft has been drained into a long roux loop without complications. The main objective in the use of this drug is to obtain consistent immunosuppression without nephrotoxicity. It is possible that maintaining blood level between defined limits would improve results. PMID:7051997

  17. Pharmacokinetics of oral cyclosporin A (Sandimmun) in healthy subjects.

    PubMed

    Grevel, J; Nüesch, E; Abisch, E; Kutz, K

    1986-01-01

    Extensive pharmacokinetic (PK) profiles after oral dosing of 300 mg cyclosporin A (CsA) were determined in whole blood by radioimmunoassay (RIA) in 14 healthy male volunteers, using two-compartment models with either first order (M1) or zero order (M0) absorption. According to zero order absorption the mean of the following PK parameters was determined: terminal half-life = 12.1 +/- 5.0 h, apparent volume of distribution at steady-state = 5.6 +/- 2.11 X kg-1, apparent clearance = 0.51 +/- 0.11 l X h-1 X kg-1. The time lag between drug ingestion and first blood level was short, 0.38 +/- 0.11 h. Drug absorption lasted for 2.8 +/- 1.6 h. The end of absorption was indicated in each individual by a sharp drop in blood levels. The observations support the assumption that CsA is absorbed in the upper part of the small intestine with a clear-cut termination (absorption window). This assumption may explain the high degree of variability in the bioavailability of CsA.

  18. Ca(2+)-loading modulates potencies of cyclosporin A, Mg2+ and ADP to recouple permeabilized rat liver mitochondria.

    PubMed

    Andreyev AYu; Mikhaylova, L M; Starkov, A A; Kushnareva YuE

    1994-09-01

    We studied the relative potencies of cyclosporin A and endogenous effectors (Mg2+ and ADP) to recouple rat liver mitochondria permeabilized by different Ca(2+)-loading in a P(i)-containing medium. Recoupling efficiency of cyclosporin A dramatically decreased at high Ca(2+)-loading (approx. 100 nM of Ca2+/mg protein and more). Mitochondria permeabilized by high Ca2+ were recoupled with approximately equal efficiency by higher cyclosporin A concentrations or by adding 1-5 mM Mg2+ together with low concentrations of cyclosporin A while potentiating effect of ADP on the cyclosporin A recoupling potency was insignificant. Mg2+ ions at concentrations of 3 mM and higher also prevented the carboxyatractylate-induced reversion of cyclosporin A recoupling effect. The data point to competitive relationships between cyclosporin A and/or Mg2+ ions and Ca2+ ions for the site(s) regulating permeability state of the pore.

  19. Immunosuppressive and antiparasitic effects of cyclosporin A on Hymenolepis nana infection in mice.

    PubMed

    Matsuzawa, K; Nakamura, F; Abe, M; Okamoto, K

    1998-04-01

    The effect of cyclosporin A, which is known to act both as immunosuppressant and as an antiparasitic drug in many host-parasite systems, was examined in a mouse-Hymenolepis nana system. When BDF1 mice were injected s.c. with cyclosporin A (100 mg kg-1 day-1) every 48 h from 11 days p.i. with eggs, expulsion of the adult worms from the intestines of mice was prevented completely until at least 30 days p.i. Worm burden, dry weight and the number of gravid proglottids were not significantly reduced. By contrast, in untreated mice most of the worms were eliminated by 19 days p.i. The drug also completely abolished acquired resistance to a challenge infection with eggs when mice were injected s.c. with cyclosporin A (100 mg kg-1 day-1) around the time of challenge infection (Days -2, -1, 0, 1 and 2 relative to challenge). Such immunosuppressive effects of cyclosporin A on worm expulsion and protective immunity to reinfection were similar to those of another immunosuppressant, cyclophosphamide. As for the antiparasitic action of cyclosporin A against H. nana, a smaller number of cysticercoids developed from eggs in mice given cyclosporin A (100 mg kg-1 day-1) for 5 days beginning 1 day before infection, than in untreated controls.

  20. The NMR structure of cyclosporin A bound to cyclophilin in aqueous solution

    SciTech Connect

    Weber, C.; Wilder, G.; von Freyberg, B.; Braun, W.; Wuethrich, K. ); Traber, R.; Widmer, H. )

    1991-07-02

    Cyclosporin A bound to the presumed receptor protein cyclophilin was studied in aqueous solution at pH 6.0 by nuclear magnetic resonance spectroscopy using uniform {sup 15}N- or {sup 13}C-labeling of cyclosporin A and heteronuclear spectral editing techniques. With an input of 108 intramolecular NOEs and four vicinal {sup 3}J{sub HN{alpha}} coupling constants, the three-dimensional structure of cyclosporin A bound to cyclophilin was calculated with the distance geometry program DISMAN, and the structures resulting from 181 converged calculations were energy refined with the program FANTOM. A group of 120 conformers was selected on the basis of the residual constraint violations and energy criteria to represent the solution structure. The average of the pairwise root-mean-square distances calculated for the backbone atoms of the 120 structures was 0.58 {angstrom}. The structure represents a novel conformation of cyclosporin A, for which the backbone conformation is significantly different from the previously reported structures in single crystals and in chloroform solution. The structure has all peptide bonds in the trans form, contains no elements of regular secondary structure and no intramolecular hydrogen bonds, and exposes nearly all polar groups to its environment. The root-mean-square distance between the backbone atoms of the crystal structure of cyclosporin A and the mean of the 120 conformers representing the NMR structure of cyclosporin A bound to cyclophilin is 2.5 {angstrom}.

  1. Enzyme and combination therapy with cyclosporin A in the rat developing adjuvant arthritis.

    PubMed

    Rovenská, E; Svík, K; Stancíková, M; Rovenský, J

    1999-01-01

    Recent knowledge of the pathophysiology of rheumatoid arthritis and the mechanism of drug effects have enabled the use of new drugs and drug combinations in rheumatoid arthritis therapy. This study investigates the efficacy of both enzyme therapy and combined therapy with cyclosporin in rats with adjuvant arthritis. Rats with adjuvant-induced arthritis were administered either cyclosporin A (2.5 or 5.0 mg/kg/day per os), a mixture of enzymes (Phlogenzym (PHL); 45 mg/kg twice daily intrarectally), or a combination of 2.5 mg cyclosporin A and 90 mg PHL for a period of 40 days from the adjuvant application. Levels of serum albumin, changes in hind paw swelling and bone erosions were measured in rats as variables of inflammation and arthritis-associated destructive changes. Treatment with 5 mg of cyclosporin A, as well as with the combination therapy with cyclosporin A plus PHL, significantly inhibited both the inflammation and destructive arthritis-associated changes. However, 2.5 mg of cyclosporin A and PHL alone inhibited these disease markers, although to a lesser extent and at a later stage of arthritis development. The results show the inhibitory effect of enzyme therapy on rat adjuvant arthritis, as well as the efficacy of a low dose of cyclosporin A given in combination with enzyme therapy, which may be useful in the treatment of rheumatoid arthritis.

  2. Identification of Spongionella compounds as cyclosporine A mimics.

    PubMed

    Sánchez, Jon Andoni; Alfonso, Amparo; Leirós, Marta; Alonso, Eva; Rateb, Mostafa E; Jaspars, Marcel; Houssen, Wael E; Ebel, Rainer; Tabudravu, J; Botana, Luís M

    2016-05-01

    Marine sponges are found to be a wide source of bioactive compounds with different effects such as anti-inflammatory or anticancer actions among others. Cyclophilin A (Cyp A) is a target protein implicated in the mechanism of action of immunosuppressive compounds such as Cyclosporine A (CsA). In the present paper we studied the binding between 4 Spongionella compounds (Gracilins H, A, L and Tetrahydroaplysulphurin-1) and Cyp A immobilized over a CM5 sensor chip. Thus, we found that Spongionella compounds showed to have similar binding affinities than CsA with dissociation equilibrium constant in the range. Next, the effect of these Spongionella isolated compounds was tested over calcineurin phosphatase activity. The same than CsA, Gracilin H, A and Tetrahydroaplysulphurin-1 were able to inhibit phosphatase activity once the complex between Cyp A-CsA/Spongionella compounds was formed. The ability to avoid the dephosphorylation of NFATc1 was also checked in human T cells isolated from peripheral blood. First, cells were pre-treated with Spongionella compounds or CsA following by Concanavalin A (Con A) stimulation. In these conditions nuclear NFATc1 levels were diminished either by CsA or Gracilin A, L, and Tetrahydroaplysulphurin-1 treatment. Moreover, as happens with CsA due to the inhibition of NFATc1, Interleukine-2 (IL-2) released to the culture medium was significantly decreased with all Spongionella compounds. Results conclude that, Spongionella derivatives preserve T lymphocytes from activation modulating the same pathway than CsA. Thus, this mechanism of action suggests that these compounds could be interesting candidates in drug development as immunosuppressive or anti-inflammatory drugs.

  3. Bioequivalence of a new cyclosporine a formulation to Neoral.

    PubMed

    David-Neto, Elias; Kakehashi, Erica; Alves, Cristiane Feres; Pereira, Lilian M; de Castro, Maria Cristina R; de Mattos, Renata Maciel; Sumita, Nairo Massakazu; Romano, Paschoalina; Mendes, Maria Elizabete; Nahas, William Carlos; Ianhez, Luiz Estevam

    2004-02-01

    New cyclosporine A (CsA) formulations must prove their bioequivalence to Neoral, the reference CsA formulation, to allow free prescription for the patients. The aim of this study was to compare the pharmacokinetics (PK) of a new CsA formulation (Zinograf-ME), produced by Strides-Arcolab, to Neoral and to demonstrate their interchangeability in stable renal transplant recipients. Twelve-hour PK studies were obtained from 18 (13 M/5 F) adult patients (mean age 44.7 +/- 12 years). They received their renal allografts from 13 cadaver and 5 living donors. Before enrollment, all patients were receiving a third generic CsA for a mean of 48 months. Nine patients were also under azathioprine and 9 under mycophenolate mofetil; 17 received prednisone. A single oral dose of either Zinograf or Neoral was administered. The first PK study was performed with one formulation, and 1 week later, a second PK was done with the other formulation. During the washout period, patients continued taking the third CsA formulation. The drug substitution was done milligram-for-milligram. The CsA whole-blood level was measured by TDx immunoassay. Mean +/- SD of area under the curve (AUC), maximum concentration (C(max)), and concentration at the second hour (C2) of Zinograf were not statistically different from those with Neoral (4019 +/- 1466 vs 3971 +/- 1325 ng x h/mL, 998 +/- 376 vs 1021 +/- 356 ng/mL, and 707 +/- 254 vs 734 +/- 229 ng/mL, respectively). In the same way, the Zinograf 90% confidence interval for either C(max) (-123, +77 ng/mL) or AUC (-214, +311 ng.mL/h) were within the Neoral bioequivalence interval for the same parameters (+/-204 ng/mL and +/-794 ng x mL/h, respectively). These data demonstrate that the ZinografME CsA formulation is bioequivalent to Neoral.

  4. Cyclosporin A Disrupts Notch Signaling and Vascular Lumen Maintenance

    PubMed Central

    Pandey, Raghav; Botros, Mark A.; Nacev, Benjamin A.; Albig, Allan R.

    2015-01-01

    Cyclosporin A (CSA) suppresses immune function by blocking the cyclophilin A and calcineurin/NFAT signaling pathways. In addition to immunosuppression, CSA has also been shown to have a wide range of effects in the cardiovascular system including disruption of heart valve development, smooth muscle cell proliferation, and angiogenesis inhibition. Circumstantial evidence has suggested that CSA might control Notch signaling which is also a potent regulator of cardiovascular function. Therefore, the goal of this project was to determine if CSA controls Notch and to dissect the molecular mechanism(s) by which CSA impacts cardiovascular homeostasis. We found that CSA blocked JAG1, but not Dll4 mediated Notch1 NICD cleavage in transfected 293T cells and decreased Notch signaling in zebrafish embryos. CSA suppression of Notch was linked to cyclophilin A but not calcineurin/NFAT inhibition since N-MeVal-4-CsA but not FK506 decreased Notch1 NICD cleavage. To examine the effect of CSA on vascular development and function, double transgenic Fli1-GFP/Gata1-RFP zebrafish embryos were treated with CSA and monitored for vasculogenesis, angiogenesis, and overall cardiovascular function. Vascular patterning was not obviously impacted by CSA treatment and contrary to the anti-angiogenic activity ascribed to CSA, angiogenic sprouting of ISV vessels was normal in CSA treated embryos. Most strikingly, CSA treated embryos exhibited a progressive decline in blood flow that was associated with eventual collapse of vascular luminal structures. Vascular collapse in zebrafish embryos was partially rescued by global Notch inhibition with DAPT suggesting that disruption of normal Notch signaling by CSA may be linked to vascular collapse. However, multiple signaling pathways likely cause the vascular collapse phenotype since both cyclophilin A and calcineurin/NFAT were required for normal vascular function. Collectively, these results show that CSA is a novel inhibitor of Notch signaling and

  5. An underlying role for hepatobiliary dysfunction in cyclosporine A nephrotoxicity

    SciTech Connect

    Aleo, Michael D.

    2008-07-01

    Renal-derived cysteinyl leukotrienes (cysLT), such as leukotrienes C{sub 4} (LTC{sub 4}) and D{sub 4} (LTD{sub 4}) are thought to mediate acute and chronic cyclosporine A (CSA) nephrotoxicity. However, whole-body cysLT elimination is regulated primarily by hepatobiliary excretion. Since CSA is known to alter hepatobiliary function, the effects of CSA on whole-body cysLT elimination were investigated in vivo, with respect to hepatobiliary and renal function. Male rats were anesthetized and cannulated (jugular vein, bile duct, and urinary bladder). A tracer dose of tritiated LTC{sub 4} ({sup 3}H-LTC{sub 4}) was administered systemically (i.v.) immediately following vehicle and then 90 min later after vehicle or CSA. In vehicle/vehicle controls, hepatobiliary {sup 3}H-cysLT elimination predominated over renal elimination without altering glomerular filtration rate (GFR), bile flow, and urine production. {sup 3}H-cysLT elimination kinetics were comparable between each 90 min collection period. In vehicle/CSA-treated rats, an acutely nephrotoxic dose of CSA (20 mg/kg, i.v.) reduced urine flow 74 {+-} 9% and caused a transient reduction in GFR, while total bile flow decreased 40 {+-} 13%. Hepatobiliary and renal {sup 3}H-cysLT elimination was also impaired 59 {+-} 5 and 61 {+-} 18%, respectively. In contrast, a non-nephrotoxic dose (2 mg/kg i.v.) increased renal {sup 3}H-cysLT elimination due to impaired hepatobiliary elimination without affecting GFR, bile flow or urine production. Both doses caused {sup 3}H-cysLT retention in hepatic and renal tissue. These findings demonstrate that CSA alters whole-body handling of cysLT by disrupting hepatobiliary cysLT elimination. This disruption leads to increased renal exposure to systemically derived cysLT and renal cysLT tissue retention. Renal exposure to and accumulation of systemically derived cysLT products may be underlying factors in CSA nephrotoxicity.

  6. Modification of c and n sources for enhanced production of cyclosporin 'a' by Aspergillus Terreus.

    PubMed

    Tanseer, Sundas; Anjum, Tehmina

    2011-10-01

    Most of the studies regarding cyclosporin 'A' production through fungi concentrate around Tolypocladium inflatum. This is mainly due to lower reported production of this drug in other fungi. The present study was therefore conducted to explore indigenous isolates of Aspergillus terreus for synthesis of this drug and defining a production medium for obtaining high yield of cyclosporin 'A'. For this purpose carbon and nitrogen sources were optimized for the selected best strain of A. terreus. Overall results depicted that the best cyclosporin 'A' yield from selected Aspergillus terreus (FCBP58) could be obtained by using production medium containing glucose 10% as carbon source and peptone 0.5% as nitrogen source. This modification in production medium enhanced drug synthesis by selected fungi significantly. The production capabilities when compared with biomass of fungi there was found no relationship between the two confirming that the medium modification increased overall drug synthesis powers of the fungi.

  7. Conversion from tacrolimus to cyclosporine--a based immunosuppression following liver transplantation.

    PubMed

    Doria, Cataldo; Jain, Ashok Kumar B; Scott, Victor L; Gruttadauria, Salvatore; Marino, Ignazio R; Doyle, Howard R; Fung, John J

    2003-06-01

    We examined the frequency, reasons and outcome after conversion from Tacrolimus to Cyclosporine A. From August 1989 to December 1992, 1000 consecutive liver transplantation patients were studied, which included 834 adults (age>18 yr.) and 166 children with mean follow-up of 77 months (range 56 to 96). A prospectively populated electronic database was queried to identify patients that underwent conversion, the clinical indication and outcomes. Thirty-seven out of 834 adult recipients (4.43%), mean age of 48.4+/-12.9 years, 19 male (51.35%) and 18 females (48.64%) required conversion from Tacrolimus to Cyclosporine A baseline immunosuppressive therapy. No pediatric patient required conversion. The mean time interval from liver transplantation to Cyclosporine A conversion was 443.45+/-441.44 days (range 22 to 1641). The clinical indications for conversion included: 20 neurological (54%), 6 gastrointestinal (16%), 5 hematological (14%), and 6 other (16%) scenarios. Seven of the 37 patients (18.9%) died. The causes of death were multi-organ failure (2), sepsis (2), pancreatitis (1), hepatic failure due to relapse of ethanol abuse (1), and unknown cause (1). Nine out of 37 patients (24.32%) had to be reconverted to Tacrolimus (mean 282.22+/-499.79 days; range 15 to 1583 day with a median of 135) after institution of Cyclosporine A; none showed recurrence of the original symptoms. The reasons for these re-conversions were acute cellular rejection (44%, n=4), chronic rejection (11%, n=1), increased hepatic enzymes (33%, n=3) and progressively worsening neurological symptoms (11%, n=1). The frequency of conversion from Tacrolimus to Cyclosporine A was 4.43%. Conversion is safe and efficacious if done in a controlled setting. Additionally, re-conversion to Tacrolimus for lack of efficacy of Cyclosporine A did not appear to be associated with a recurrence of the condition that caused the initial switch.

  8. Cyclosporine A or intravenous cyclophosphamide for lupus nephritis: the Cyclofa-Lune study.

    PubMed

    Zavada, J; Pesickova, Ss; Rysava, R; Olejarova, M; Horák, P; Hrncír, Z; Rychlík, I; Havrda, M; Vítova, J; Lukác, J; Rovensky, J; Tegzova, D; Böhmova, J; Zadrazil, J; Hána, J; Dostál, C; Tesar, V

    2010-10-01

    Intravenous cyclophosphamide is considered to be the standard of care for the treatment of proliferative lupus nephritis. However, its use is limited by potentially severe toxic effects. Cyclosporine A has been suggested to be an efficient and safe treatment alternative to cyclophosphamide. Forty patients with clinically active proliferative lupus nephritis were randomly assigned to one of two sequential induction and maintenance treatment regimens based either on cyclophosphamide or Cyclosporine A. The primary outcomes were remission (defined as normal urinary sediment, proteinuria <0.3 g/24 h, and stable s-creatinine) and response to therapy (defined as stable s-creatinine, 50% reduction in proteinuria, and either normalization of urinary sediment or significant improvement in C3) at the end of induction and maintenance phase. Secondary outcomes were incidence of adverse events, and relapse-free survival. At the end of the induction phase, 24% of the 21 patients treated by cyclophosphamide achieved remission, and 52% achieved response, as compared with 26% and 43%, respectively of the 19 patients treated by the Cyclosporine A. At the end of the maintenance phase, 14% of patients in cyclophosphamide group, and 37% in Cyclosporine A group had remission, and 38% and 58% respectively response. Treatment with Cyclosporine A was associated with transient increase in blood pressure and reversible decrease in glomerular filtration rate. There was no significant difference in median relapse-free survival. In conclusion, Cyclosporine A was as effective as cyclophosphamide in the trial of sequential induction and maintenance treatment in patients with proliferative lupus nephritis and preserved renal function.(ClinicalTrials.gov identifier: NCT00976300)

  9. Cyclosporine A-Nanosuspension: Formulation, Characterization and In Vivo Comparison with a Marketed Formulation

    PubMed Central

    Nakarani, Mahendra; Patel, Priyal; Patel, Jayvadan; Patel, Pankaj; Murthy, Rayasa S. R.; Vaghani, Subhash S.

    2010-01-01

    Cyclosporine A-nanosuspensions were prepared using zirconium oxide beads as a milling media, Poloxamer 407 as a stabilizer and distilled water as an aqueous medium using the Pearl Milling technique. The optimized formulation was characterized in terms of particle size distribution, surface morphology, drug-surfactant interaction, drug content, saturation solubility, osmolarity, and stability. The nanoparticles consisting of Poloxamer-bound cyclosporin A with a mean diameter of 213 nm revealed a spherical shape and 5.69 fold increased saturation solubility as compared to the parent drug. The formulation was found to be iso-osmolar with blood and stable up to 3 months at 2–8°C. In-vivo studies were carried out in albino rats and the pharmacokinetic parameters were compared with a marketed formulation, which indicated better results of the prepared formulation than the marketed one. PMID:21179351

  10. The influence of cyclosporin A on experimental autoimmune thyroid disease in the rat

    SciTech Connect

    McGregor, A.M.; Rennie, D.P.; Weetman, A.P.; Hassman, R.A.; Foord, S.M.; Dieguez, C.; Hall, R.

    1983-01-01

    Female PVG/c rats, thymectomised on weaning and given 4 courses of whole body irradiation to a total dose of 1000 rads, developed experimental autoimmune thyroid disease (EAITD) as assessed by histological evidence of thyroiditis and circulating levels of antithyroglobulin antibodies. Hypothyroidism resulted. Induction of the disease was associated with a highly significant fall in T lymphocyte numbers. Eight weeks after their last dose of irradiation the animals commenced treatment with cyclosporin A (10 mg/kg rat/day, intragastrically) and were treated for varying time intervals thereafter. The reversal of the T lymphocyte helper: suppressor ratio on cyclosporin A therapy was associated with a significant improvement in the disease process. The alterations in the T cell subsets and in the disease lasted only as long as the drug was administered and thereafter reverted towards that seen in the control groups of animals receiving no treatment.

  11. Surgical treatment of cyclosporine A- and nifedipine-induced gingival enlargement: gingivectomy versus periodontal flap.

    PubMed

    Pilloni, A; Camargo, P M; Carere, M; Carranza, F A

    1998-07-01

    The purpose of this study was to compare probing depth resolution achieved by gingivectomy and periodontal flap techniques in the treatment of cyclosporine A- and nifedipine-induced gingival enlargement. Ten kidney transplant patients who were receiving cyclosporine A and nifedipine for at least 6 months participated in the study. Five patients were randomly assigned to the gingivectomy group and 5 patients to the periodontal flap group. Only anterior segments of the oral cavity (canine to canine) were surgically treated. Clinical measurements, including probing depths, plaque index, and gingival sulcus index, were taken at baseline, 6 weeks, 6 months, and 1 year. Results showed that probing depths, while similar for both groups in the first 6 weeks of the study, were significantly shallower for the periodontal flap group when compared to the gingivectomy group at 6 months (2.48 +/- 0.34 mm versus 4.87 +/- 0.79 mm, respectively) and 1 year (322 +/- 0.65 mm versus 6.40 +/- 1.02 mm, respectively). Within its limitations, this study suggests that the pocket reduction achieved by the periodontal flap may be sustained for longer periods of time than by the gingivectomy technique in the treatment of cyclosporine A- and nifedipine-induced gingival enlargement.

  12. Effect of Cyclosporin A on the Uptake of D3-Selective PET Radiotracers in Rat Brain

    PubMed Central

    Tu, Zhude; Li, Shihong; Xu, Jinbin; Chu, Wenhua; Jones, Lynne A.; Luedtke, Robert R.; Mach, Robert H.

    2011-01-01

    Introduction Four benzamide analogs having a high affinity and selectivity for D3 versus D2 receptors were radiolabeled with 11C or 18F for in vivo evaluation. Methods Precursors were synthesized and the four D3 selective benzamide analogs were radiolabeled. The tissue distribution and brain uptake of the four compounds were evaluated in control rats and rats pretreated with cyclosporin A, a modulator of P-glycoprotein and an inhibitor of other ABC efflux transporters that contribute to the blood brain barrier. MicroPET imaging was carried out for [11C]6 in a control and a cyclosporin A pre-treated rat. Results All four compounds showed low brain uptake in control rats at 5 and 30 min post-injection; despite recently reported rat behavioral studies conducted on analogs 6 (WC-10) and 7 (WC-44). Following administration of cyclosporin A, increased brain uptake was observed with all four PET radiotracers at both 5 and 30 min post-i.v. injection. An increase in brain uptake following modulation/inhibition of the ABC transporters was also observed in the microPET study. Conclusions These data suggest that D3 selective conformationally-flexible benzamide analogs which contain a N-2-methoxyphenylpiperazine moiety are substrates for P-glycoprotein or other ABC transporters expressed at the blood-brain barrier, and that PET radiotracers containing this pharmacophore may display low brain uptake in rodents due to the action of these efflux transporters. PMID:21718948

  13. Management of Toxic Epidermal Necrolysis with Plasmapheresis and Cyclosporine A: Our 10 Years’ Experience

    PubMed Central

    Giudice, Giuseppe; Maggio, Giulio; Bufano, Loredana; Memeo, Giuseppe

    2017-01-01

    Background: The management of toxic epidermal necrolysis (TEN) is controversial and there is no uniform strategy. Objective: To share our 10 years’ experience in treating severe TEN with a novel protocol based on the association of cyclosporine A and plasmapheresis. Methods: In this case series, we retrospectively collected and assessed the 12 cases of severe TEN treated from 2005 to 2015 at the Burn Unit of the University of Bari Policlinico hospital. Results: Average body surface area was 77; average SCORETEN was 4.3. The 12 patients had been treated with culprit drug withdrawal, systemic corticosteroids, and/or cyclosporine A with no response. The protocol was successfully administered in all 12 cases. Average time to response from protocol start was 4.9 days. Average time to remission from protocol start was 22 days; average hospital stay at our unit was 24.8 days. Four patients developed severe complications; 1 patient died. No complications linked to the protocol therapeutic measures were observed. The relatively small number of cases given the rarity of the condition is a limitation of this report. Conclusion: Our protocol based on the association of cyclosporine A and plasmapheresis is safe and efficacious in treating severe TEN. PMID:28280663

  14. Cyclosporin A corrects mitochondrial dysfunction and muscle apoptosis in patients with collagen VI myopathies.

    PubMed

    Merlini, Luciano; Angelin, Alessia; Tiepolo, Tania; Braghetta, Paola; Sabatelli, Patrizia; Zamparelli, Alessandra; Ferlini, Alessandra; Maraldi, Nadir M; Bonaldo, Paolo; Bernardi, Paolo

    2008-04-01

    Ullrich congenital muscular dystrophy and Bethlem myopathy are skeletal muscle diseases that are due to mutations in the genes encoding collagen VI, an extracellular matrix protein forming a microfibrillar network that is particularly prominent in the endomysium of skeletal muscle. Myoblasts from patients affected by Ullrich congenital muscular dystrophy display functional and ultrastructural mitochondrial alterations and increased apoptosis due to inappropriate opening of the permeability transition pore, a mitochondrial inner membrane channel. These alterations could be normalized by treatment with cyclosporin A, a widely used immunosuppressant that desensitizes the permeability transition pore independently of calcineurin inhibition. Here, we report the results of an open pilot trial with cyclosporin A in five patients with collagen VI myopathies. Before treatment, all patients displayed mitochondrial dysfunction and increased frequency of apoptosis, as determined in muscle biopsies. Both of these pathologic signs were largely normalized after 1 month of oral cyclosporin A administration, which also increased muscle regeneration. These findings demonstrate that collagen VI myopathies can be effectively treated with drugs acting on the pathogenic mechanism downstream of the genetic lesion, and they represent an important proof of principle for the potential therapy of genetic diseases.

  15. Dose-response curve and optimal dosing regimen of cyclosporin A after traumatic brain injury in rats.

    PubMed

    Sullivan, P G; Rabchevsky, A G; Hicks, R R; Gibson, T R; Fletcher-Turner, A; Scheff, S W

    2000-01-01

    Acute neuropathology following experimental traumatic brain injury results in the rapid necrosis of cortical tissue at the site of injury. This primary injury is exacerbated in the ensuing hours and days via the progression of secondary injury mechanism(s) leading to significant neurological dysfunction. Recent evidence from our laboratory demonstrates that the immunosuppressant cyclosporin A significantly ameliorates cortical damage following traumatic brain injury. The present study extends the previous findings utilizing a unilateral controlled cortical impact model of traumatic brain injury in order to establish a dose-response curve and optimal dosing regimen of cyclosporin A. Following injury to adult rats, cyclosporin A was administrated at various dosages and the therapy was initiated at different times post-injury. In addition to examining the effect of cyclosporin A on the acute disruption of the blood-brain barrier following controlled cortical impact, we also assessed the efficacy of cyclosporin A to reduce tissue damage utilizing the fluid percussion model of traumatic brain injury. The findings demonstrate that the neuroprotection afforded by cyclosporin A is dose-dependent and that a therapeutic window exists up to 24h post-injury. Furthermore, the optimal cyclosporin dosage and regimen markedly reduces disruption of the blood-brain barrier acutely following a cortical contusion injury, and similarly affords significant neuroprotection following fluid percussion injury. These findings clearly suggest that the mechanisms responsible for tissue necrosis following traumatic brain injury are amenable to pharmacological intervention.

  16. Glucocorticoid-resistant Th17 cells are selectively attenuated by cyclosporine A.

    PubMed

    Schewitz-Bowers, Lauren P; Lait, Philippa J P; Copland, David A; Chen, Ping; Wu, Wenting; Dhanda, Ashwin D; Vistica, Barbara P; Williams, Emily L; Liu, Baoying; Jawad, Shayma; Li, Zhiyu; Tucker, William; Hirani, Sima; Wakabayashi, Yoshiyuki; Zhu, Jun; Sen, Nida; Conway-Campbell, Becky L; Gery, Igal; Dick, Andrew D; Wei, Lai; Nussenblatt, Robert B; Lee, Richard W J

    2015-03-31

    Glucocorticoids remain the cornerstone of treatment for inflammatory conditions, but their utility is limited by a plethora of side effects. One of the key goals of immunotherapy across medical disciplines is to minimize patients' glucocorticoid use. Increasing evidence suggests that variations in the adaptive immune response play a critical role in defining the dose of glucocorticoids required to control an individual's disease, and Th17 cells are strong candidate drivers for nonresponsiveness [also called steroid resistance (SR)]. Here we use gene-expression profiling to further characterize the SR phenotype in T cells and show that Th17 cells generated from both SR and steroid-sensitive individuals exhibit restricted genome-wide responses to glucocorticoids in vitro, and that this is independent of glucocorticoid receptor translocation or isoform expression. In addition, we demonstrate, both in transgenic murine T cells in vitro and in an in vivo murine model of autoimmunity, that Th17 cells are reciprocally sensitive to suppression with the calcineurin inhibitor, cyclosporine A. This result was replicated in human Th17 cells in vitro, which were found to have a conversely large genome-wide shift in response to cyclosporine A. These observations suggest that the clinical efficacy of cyclosporine A in the treatment of SR diseases may be because of its selective attenuation of Th17 cells, and also that novel therapeutics, which target either Th17 cells themselves or the effector memory T-helper cell population from which they are derived, would be strong candidates for drug development in the context of SR inflammation.

  17. Oral administration of cyclosporin A for recipients of allogeneic marrow transplants: implications of clinical gut dysfunction.

    PubMed

    Atkinson, K; Biggs, J C; Britton, K; Short, R; Mrongovius, R; Concannon, A; Dodds, A

    1984-02-01

    Cyclosporin A (CyA) was used to minimize graft-versus-host disease (GVHD) in 28 recipients of allogeneic marrow transplants. When given orally, the absorption of CyA was markedly dependent on normal gut function. Patients without gut dysfunction showed normal serum concentration-time curves while those with diarrhoea from any cause (chemo-radiation enteritis, acute GVHD of the gut, infectious enteritis) showed minimal absorption of the drug. These data indicate the desirability of the intravenous administration of CyA during periods of gut dysfunction in marrow transplant recipients.

  18. Cyclosporin A in the treatment of CLL associated PRCA and bone marrow hypoplasia.

    PubMed

    Tura, S; Finelli, C; Bandini, G; Cavo, M; Gobbi, M

    1988-01-01

    Three patients (1 PRCA-T-CLL, 1 PRCA-B-CLL, 1 B-CLL aplasia) were treated with cyclosporin A (CS-A). Patient no 1 had relapsed during steroid therapy and the remaining two patients had been resistant to conventional immunosuppression. CS-A produced in all cases a prompt remission (within 1-4 weeks) of bone marrow failure. Mild reversible renal toxicity was the only side-effect noted. CS-A might be tried in every case of CLL-associated bone marrow failure.

  19. The effect of cyclosporin A on peripheral blood T cell subpopulations in renal allografts.

    PubMed Central

    Sweny, P; Tidman, N

    1982-01-01

    Treatment with cyclosporin A (CyA) produces a reversal of the normal ratio of OKT4+ (inducer type) to OKT84 (suppressor-cytotoxic type) cells so that renal allograft recipients on CyA alone develop a four-fold increase in the absolute number of circulating OKT8 positive cells. Conventional immunosuppression with azathioprine and prednisolone reduces both populations of T cells without altering the ratio of OKT4+ to OKT8+ cells. This effect of CyA may help to explain its action as an immunosuppressive agent. PMID:6210475

  20. Cyclosporine A decreases the fluconazole minimum inhibitory concentration of Candida albicans clinical isolates but not biofilm formation and cell growth.

    PubMed

    Wibawa, T; Nurrokhman; Baly, I; Daeli, P R; Kartasasmita, G; Wijayanti, N

    2015-03-01

    Among the genus Candida, Candida albicans is the most abundant species in humans. One of the virulent factors of C. albicans is its ability to develop biofilm. Biofilm forming microbes are characterized by decreasing of its susceptibility to antibiotics and antifungal. The fungicidal effect of fluconazole may be enhanced by cyclosporine A in laboratory engineered C. albicans strains. The aim of this work is to analyze the synergistic effect of cyclosporine A with fluconazole in C. albicans clinical isolates and the effect of cycolsporine A alone in the biofilm formation. Six fluconazole resistant and six sensitive C. albicans clinical isolates were analyzed for its minimum inhibitory concentration (MICs), biofilm formation, and cell growths. A semi-quantitative XTT [2,3-bis(2-methoxy-4-nitro-5- sulfo-phenyl)-2H-tetrazolium-5-carboxanilide] reduction assay was conducted to measure the biofilm formation. Cyclosporine A has synergistic effect with fluconazole that was shown by decreasing MICs of both fluconazole resistant and sensitive C. albicans clinical isolates. However, cyclosporine A alone did not influence the biofilm formation and cell growth of both fluconazole resistant and sensitive C. albicans clinical isolates. These results indicated that cyclosporine A might be a promising candidate of adjuvant therapy for fluconazole against both fluconazole resistant and sensitive C. albicans clinical isolates.

  1. Pure red cell aplasia associated with autoimmune hepatitis successfully treated with cyclosporine A.

    PubMed

    Sato, Akira; Sano, Fumiaki; Ishii, Toshiya; Adachi, Kayo; Negishi, Ryujirou; Matsumoto, Nobuyuki; Okuse, Chiaki

    2014-02-01

    A 47-year-old female with a 17-year history of autoimmune hepatitis had been treated with prednisolone, azathioprine, and ursodeoxycholic acid. Although her alanine aminotransferase level occasionally showed mild abnormality, the prednisolone dose could not be increased because she had developed cataract during the course of her illness. In May 2012, she developed severe normochromic normocytic anemia without hemorrhage, and azathioprine was discontinued because it was suspected of being the cause. However, anemia recurred frequently even after discontinuation, necessitating repeated blood transfusions. Bone marrow analysis revealed selective erythroblastopenia, thus leading to a diagnosis of pure red cell aplasia. Cyclosporine A was administered, which led to a dramatic recovery from anemia, and stabilized her alanine aminotransferase levels. Furthermore, the prednisolone dose could be gradually tapered. Pure red cell aplasia associated with autoimmune hepatitis is extremely rare. The present case shows that patients with autoimmune hepatitis refractory to the standard treatment regimen and those with concomitant pure red cell aplasia may be treated with cyclosporine A.

  2. Oxidative Stress and Liver Morphology in Experimental Cyclosporine A-Induced Hepatotoxicity

    PubMed Central

    Czechowska, Grażyna; Irla-Miduch, Joanna

    2016-01-01

    Cyclosporine A is an immunosuppressive drug used after organ's transplantation. The adverse effects on such organs as kidney or liver may limit its use. Oxidative stress is proposed as one of the mechanisms of organs injury. The study was designed to elucidate CsA-induced changes in liver function, morphology, oxidative stress parameters, and mitochondria in rat's hepatocytes. Male Wistar rats were used: group A (control) receiving physiological saline, group B cyclosporine A in a dose of 15 mg/kg/day subcutaneously, and group C the CsA-vehicle (olive oil). On the 28th day rats were anesthetized. The following biochemical changes were observed in CsA-treated animals: increased levels of ALT, AST, and bilirubin in the serum, statistically significant changes in oxidative stress parameters, and lipid peroxidation products in the liver supernatants: MDA+4HAE, GSH, GSSG, caspase 3 activity, and ADP/ATP, NAD+/NADH, and NADP+/NADPH ratios. Microscopy of the liver revealed congestion, sinusoidal dilatation, and focal hepatocytes necrosis with mononuclear cell infiltration. Electron microscope revealed marked mitochondrial damage. Biochemical studies indicated that CsA treatment impairs liver function and triggers oxidative stress and redox imbalance in rats hepatocytes. Changes of oxidative stress markers parallel with mitochondrial damage suggest that these mechanisms play a crucial role in the course of CsA hepatotoxicity. PMID:27298826

  3. Cyclosporin A inhibits HTLV-I tax expression and shows anti-tumor effects in combination with VP-16.

    PubMed

    Ozaki, Atsuo; Arima, Naomichi; Matsushita, Kakushi; Uozumi, Kimiharu; Akimoto, Masaki; Hamada, Heiichiro; Kawada, Hideaki; Horai, Sawako; Tanaka, Yuetsu; Tei, Chuwa

    2007-12-01

    Adult T cell leukemia (ATL) is one of the most refractory malignant hematological diseases. Our previous studies demonstrated HTLV-1Tax protein involvement in clinical manifestation of the aggressive type of ATL and suggested the potential application of agents to inhibit Tax expression for ATL treatment. In the present study, we first examined Tax involvement in the resistance to VP-16-induced apoptosis using four HTLV-1 infected T cell clones and cTax DNA-transfected cells. Next, we examined whether cyclosporin A reduced expression of Tax and its related transfer factors on Western blot and CAT assay. We further investigated whether cyclosporin A in combination with VP-16 can induce apoptosis in HTLV-1 infected T cells. Tax-producing T cells, K3T and F6T, were resistant to VP-16 induced growth inhibition compared with that of the nonproducing cells, S1T and Su9T01. Experiments using S1T and Tax-expressing cDNA-transfected S1T demonstrated Tax-induced resistance to VP-16 induction of apoptosis by DNA ladder formation. Cyclosporin A reduced Tax expression in K3T by Western blot analysis and on CAT assay, showing maximal reduction of 61% and 60% compared to control culture using LTR CAT transfected Jurkat cells and K3T cells, respectively. Cyclosporin A also reduced the nuclear expression of two Tax-related transfer factors, ATF-1 and ATF-2 on Western blot. Cyclosporin A alone did not show any cytotoxicity by itself, but sensitized cells to VP-16 when combined with VP-16. Cyclosporin A may be a useful anti-ATL agent when combined with other anti-cancer agents possibly related to Tax inhibition.

  4. Safety of Eplerenone for Kidney-Transplant Recipients with Impaired Renal Function and Receiving Cyclosporine A

    PubMed Central

    Barbe, Coralie; Lavaud, Sylvie; Toupance, Olivier; Nazeyrollas, Pierre; Jaisser, Frederic; Rieu, Philippe

    2016-01-01

    Background Animal studies have highlighted the role of vascular mineralocorticoid receptor during Cyclosporine A-induced nephrotoxicity. Mineralocorticoid receptor antagonists could improve kidney survival but are not commonly used during renal impairment and in association with several immunosuppressive drugs due to a supposed higher risk of adverse events. We tested the tolerance of eplerenone according to its expected adverse events: hyperkalemia, metabolic acidosis, hypotension, acute kidney failure, or any other adverse event. Methods We conducted a single-center, prospective, open-label study in 31 kidney-transplant recipients with impaired renal function (30 and 50 mL/min/1.73m2) and receiving cyclosporine A. All patients received eplerenone 25 mg/d for 8 weeks. Serum potassium, renal function and expected adverse events were closely monitored. Results Eight patients experienced mild hyperkalemia (>5 mmol/L), one moderate hyperkalemia (>5.5 mmol/L) and had to receive potassium-exchange resin. No severe hyperkalemia (>6 mmol/L) occurred. One acute kidney failure was observed, secondary to diarrhea. Basal serum potassium and bicarbonate were independently associated with a higher risk of developing mild hyperkalemia (>5 mmol/L) under treatment (OR 6.5, p = 0.003 and 0.7, p = 0.007, respectively). A cut-off value of 4.35 mmol/L for basal serum potassium was the best factor to predict the risk of developing mild hyperkalemia (>5 mmol/L). Conclusions Until eGFR falls to 30 mL/min/1.73m2, eplerenone could be safely given to kidney-transplant recipients receiving cyclosporine A, if kalemia is closely monitored. When renal function is impaired and if basal kalemia is >4.35 mmol/L, then clinicians should properly balance risk and benefit of eplerenone use and offer dietary advice. An adequately powered prospective randomized study is now needed to test its efficiency (and safety) in this population. Trial Registration ClinicalTrials.gov NCT01834768 PMID:27088859

  5. Structural characterization of cyclosporin A, C and microbial bio-transformed cyclosporin A analog AM6 using HPLC-ESI-ion trap-mass spectrometry.

    PubMed

    Ahn, Eun Young; Shrestha, Anil; Hoang, Nguyen Huu; Huong, Nguyen Lan; Yoon, Yeo Joon; Park, Je Won

    2014-06-01

    Cyclosporin A (CyA), a cyclic undecapeptide produced by a number of fungi, contains 11 unusual amino acids, and has been one of the most commonly prescribed immunosuppressive drugs. To date, there are over sixty different analogs reported as congeners and analogs resulting from precursor-directed biosynthesis, human CYP-mediated metabolites, or microbial bio-transformed analogs. However, there is still a need for more structurally diverse CyA analogs in order to discover new biological potentials and/or improve the physicochemical properties of the existing cyclosporins. As a result of the complexity of the resulting mass spectrometric (MS) data caused by its unusual amino acid composition and its cyclic nature, structural characterization of these cyclic peptides based on fragmentation patterns using multiple tandem MS analyses is challenging task. Here, we describe, an efficient HPLC-ESI-ion trap MS(n) (up to MS(8)) was developed for the identification of CyA and CyC, a (Thr(2))CyA congener in which L-aminobutyric acid (Abu) is replaced by L-threonine (Thr). In addition, we examined the fragmentation patterns of a CyA analog obtained from the cultivation of a recombinant Streptomyces venezuelae strain fed with CyA, assigning this analog as (γ-hydroxy-MeLeu(6))CyA (otherwise, known as an human CYP metabolite AM6). This is the first report on both the MS(n)-aided identification of CyC and the structural characterization of a CyA analog by employing HPLC-ESI-ion trap MS(n) analysis.

  6. Cyclosporin a aerosol improves the anticancer effect of Paclitaxel aerosol in mice.

    PubMed Central

    Knight, Vernon; Koshkina, N. V.; Golunski, E.; Roberts, L. E.; Gilbert, B. E.

    2004-01-01

    Paclitaxel (PTX) is a lipophilic agent with broad anticancer activity. In the present study we examined the antitumor effect and toxicity of co-administration of cyclosporine A (CsA) and PTX in liposomal aerosol using the Renca lung metastases mouse model. The untreated and PTX-only groups exhibited cancer growth while CsA aerosol plus PTX had more favorable effects on tumor growth. Weight loss was seen in mice treated with CsA/PTX+CsA by day 9 to 22. Histopathological examination showed no toxicity following treatment. The findings offer evidence that a combination of CsA and PTX may be suitable for aerosol treatment of lung cancer if it is possible to control toxicity of the therapy. Images Fig. 1 PMID:17060982

  7. Prevention of diabetes mellitus in the BB/W rat with Cyclosporin-A.

    PubMed Central

    Like, A. A.; Dirodi, V.; Thomas, S.; Guberski, D. L.; Rossini, A. A.

    1984-01-01

    Autoimmune diabetes mellitus occurs spontaneously in 40-60% of a colony of BioBreeding/Worcester rats. Pretreatment of susceptible animals for 10-day intervals prior to 70 days of age with Cyclosporin-A (CSA) significantly reduced the frequency and delayed the onset of diabetes. The relatively narrow time frame of successful treatment suggests that effector cells responsible for beta cell destruction in this model of Type I diabetes may be activated during this period of time prior to the onset of overt hyperglycemia. CSA administration did not protect against the occurrence of lymphocytic thyroiditis or autoantibodies directed against smooth muscle or thyroid colloid, suggesting that these BB immunologic phenomena may be controlled by a distinct series of immunologic events. PMID:6385729

  8. Hypoplastic myelodysplastic syndrome transformed in acute myeloid leukemia after androgens and cyclosporin. A treatment.

    PubMed

    Gologan, R; Ostroveanu, Daniela; Dobrea, Camelia; Gioadă, Liliana

    2003-01-01

    The apparent contradiction between clonal expansion and marrow failure encountered in myelodysplastic syndromes (MDS) is more evident in hypocellular forms at presentation. Hypoplastic MDS (hMDS) appears to be a distinct clinicopathologic entity, accounting for about 15% from all MDS. The pathogeny is supposed to result from immunosupressive mechanisms and some observations on successful treatment with Cyclosporine A (CsA) are reported. The case of a young female patient diagnosed by bone marrow core biopsy with hMDS - refractory anemia (FAB and WHO classification) with normal karyotype and scarce CD34(+) cells by immunohistophenotyping is presented. She was treated with androgens followed by CsA for a few months and shortly after she developed an acute myeloid leukemia (M4) which responded to low-doses of daily oral melphalan. This is one of the first few reports on such an event during the immunosuppressive therapy in MDS and the possible explanations for this unusual evolution are discussed.

  9. Characterization of Folding Cores in the Cyclophilin A-Cyclosporin A Complex

    PubMed Central

    Heal, Jack W.; Wells, Stephen A.; Blindauer, Claudia A.; Freedman, Robert B.; Römer, Rudolf A.

    2015-01-01

    Determining the folding core of a protein yields information about its folding process and dynamics. The experimental procedures for identifying the amino acids that make up the folding core include hydrogen-deuterium exchange and Φ-value analysis and can be expensive and time consuming. Because of this, there is a desire to improve upon existing methods for determining protein folding cores theoretically. We have obtained HDX data for the complex of cyclophilin A with the immunosuppressant cyclosporin A. We compare these data, as well as literature values for uncomplexed cyclophilin A, to theoretical predictions using a combination of rigidity analysis and coarse-grained simulations of protein motion. We find that in this case, the most specific prediction of folding cores comes from a combined approach that models the rigidity of the protein using the first software suite and the dynamics of the protein using the froda tool. PMID:25863065

  10. Cyclosporin A significantly improves preeclampsia signs and suppresses inflammation in a rat model.

    PubMed

    Hu, Bihui; Yang, Jinying; Huang, Qian; Bao, Junjie; Brennecke, Shaun Patrick; Liu, Huishu

    2016-05-01

    Preeclampsia is associated with an increased inflammatory response. Immune suppression might be an effective treatment. The aim of this study was to examine whether Cyclosporin A (CsA), an immunosuppressant, improves clinical characteristics of preeclampsia and suppresses inflammation in a lipopolysaccharide (LPS) induced preeclampsia rat model. Pregnant rats were randomly divided into 4 groups: group 1 (PE) rats each received LPS via tail vein on gestational day (GD) 14; group 2 (PE+CsA5) rats were pretreated with LPS (1.0 μg/kg) on GD 14 and were then treated with CsA (5mg/kg, ip) on GDs 16, 17 and 18; group 3 (PE+CsA10) rats were pretreated with LPS (1.0 μg/kg) on GD 14 and were then treated with CsA (10mg/kg, ip) on GDs 16, 17 and 18; group 4 (pregnant control, PC) rats were treated with the vehicle (saline) used for groups 1, 2 and 3. Systolic blood pressure, urinary albumin, biometric parameters and the levels of serum cytokines were measured on day 20. CsA treatment significantly reduced LPS-induced systolic blood pressure and the mean 24-h urinary albumin excretion. Pro-inflammatory cytokines IL-6, IL-17, IFN-γ and TNF-α were increased in the LPS treatment group but were reduced in (LPS+CsA) group (P<0.05). Anti-inflammatory cytokine IL-4 was decreased in the LPS group but was increased in (LPS+CsA) group (P<0.05). Cyclosporine A improved preeclampsia signs and attenuated inflammatory responses in the LPS induced preeclampsia rat model which suggests that immunosuppressant might be an alternative management option for preeclampsia.

  11. The protective effect of erdosteine against cyclosporine A-induced cardiotoxicity in rats.

    PubMed

    Selcoki, Yusuf; Uz, Ebru; Bayrak, Reyhan; Sahin, Semsettin; Kaya, Arif; Uz, Burak; Karanfil, Aydin; Ozkara, Adem; Akcay, Ali

    2007-09-24

    Cyclosporine A (CsA) is a frequently used immunosuppressive agent in transplant medicine to prevent rejection and in the treatment of autoimmune diseases. However, CsA generates reactive oxygen species, which causes nephrotoxicity, hepatotoxicity and cardiotoxicity. The use of antioxidants reduces the adverse effects of CsA. The aim of this study is to determine the protective effects of erdosteine on CsA-induced heart injury through tissue oxidant/antioxidant parameters and light microscopic evaluation in rats. CsA cardiotoxicity was induced by administrating an oral dose of 15mg/kg CsA daily for 21 days. The rats were divided into four groups: control group (n=4), CsA administrated group (15mg/kg, n=5), CsA+erdosteine administrated group (10mg/kg day orally erdosteine, n=4) and only erdosteine administrated group (10mg/kg day orally n=5). CsA treated rats showed increase in the number of infiltrated cells and disorganization of myocardial fibers with interstitial fibrosis. The number of infiltrated cells, disorganization of myocardial fibers and interstitial fibrosis was diminished in the hearts of CsA-treated rats given erdosteine. The malondialdehyde, the protein carbonyl content and nitric oxide levels were increased in the cyclosporine A group in comparison with the control and CsA plus erdosteine groups. The activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) were higher in CsA plus erdosteine group than CsA group. However, the CAT, GSH-Px and SOD activities were significantly lower in CsA group than in control group and erdosteine group. These results suggest that erdosteine has protective effect against CsA-induced cardiotoxicity.

  12. Ultra fast liquid chromatography-tandem mass spectrometry routine method for simultaneous determination of cyclosporin A, tacrolimus, sirolimus, and everolimus in whole blood using deuterated internal standards for cyclosporin A and everolimus.

    PubMed

    Meinitzer, Andreas; Gartner, Gabriele; Pilz, Stefan; Stettin, Mariana

    2010-02-01

    Specific chromatographic methods for the measurement of cyclosporin A, tacrolimus, sirolimus, and everolimus blood levels in patients with organ transplants are time consuming when large numbers of samples must be processed. The authors developed a robust and fast (1 minute) online solid-phase extraction liquid chromatography/tandem mass spectrometry method for the simultaneous quantification of cyclosporin A, tacrolimus, sirolimus, and everolimus. After protein precipitation of the whole blood with zinc sulphate and methanol, the supernatant was loaded on a wide pore reversed-phase column and cleansed of potential interferences with high flow for 20 seconds. After column switching, the analytes were transferred within 20 seconds in the back-flush mode to a short phenyl-hexyl column. The valve was then returned to its initial position and the chromatographic separation performed within 20 seconds. In the meantime, the loading column was prepared for the next injection. Ammoniated adducts of protonated molecules were used as precursor ions for all analytes. Multiple-reaction mode transitions for each immunosuppressant and the internal standards were used for quantification. The working range of the method was 10-1500 microg/L for cyclosporin A, 1.0-44 microg/L for tacrolimus, 1.0-48 microg/L for sirolimus, and 1.2-48 microg/L for everolimus. Within and between-run assay coefficients of variation ranged from 1.8% to 13.0%. The described liquid chromatography/tandem mass spectrometry method shows best performance using the internal standards cyclosporin A-d4 for cyclosporin A, everolimus-d4 for everolimus and ascomycin for tacrolimus and sirolimus. In conclusion, the authors present a very fast, robust, and economical analytical method for therapeutic monitoring of multiple immunosuppressants in daily clinical practice.

  13. Cyclosporin A pharmacokinetics in liver transplant recipients in relation to biliary T-tube clamping and liver dysfunction.

    PubMed Central

    Naoumov, N V; Tredger, J M; Steward, C M; O'Grady, J G; Grevel, J; Niven, A; Whiting, B; Williams, R

    1989-01-01

    Cyclosporin A pharmacokinetics were studied after oral (4-14 mg/kg body weight) and intravenous dosing (1.5-3.5 mg/kg) in 13 orthotopic liver transplant recipients before and after permanent clamping of the biliary T-tube. After T-tube clamping, cyclosporin A absorption was faster and more complete with the mean time of peak concentration, tmax, reduced to around three hours from around six hours and mean bioavailability rising from only 16.6% (n = 13) to 30% in the entire group (n = 11 after clamping) or to 35% after excluding two patients who developed severe cholestasis after the preclamping study. Bioavailability in these two patients fell below 8% and to around 1% in a further patient with severe graft dysfunction. Clamping reduced the metabolic clearance of cyclosporin A by only 25% from a mean before clamping of 2.9 ml/min/kg to 2.3 ml/min/kg (n = 11). Oral cyclosporin A becomes a reliable means of maintaining therapeutic drug concentrations only after bioavailability increases in association with T-tube clamping and in the absence of severe liver dysfunction or cholestasis. PMID:2651227

  14. Cyclosporin A pharmacokinetics in liver transplant recipients in relation to biliary T-tube clamping and liver dysfunction.

    PubMed

    Naoumov, N V; Tredger, J M; Steward, C M; O'Grady, J G; Grevel, J; Niven, A; Whiting, B; Williams, R

    1989-03-01

    Cyclosporin A pharmacokinetics were studied after oral (4-14 mg/kg body weight) and intravenous dosing (1.5-3.5 mg/kg) in 13 orthotopic liver transplant recipients before and after permanent clamping of the biliary T-tube. After T-tube clamping, cyclosporin A absorption was faster and more complete with the mean time of peak concentration, tmax, reduced to around three hours from around six hours and mean bioavailability rising from only 16.6% (n = 13) to 30% in the entire group (n = 11 after clamping) or to 35% after excluding two patients who developed severe cholestasis after the preclamping study. Bioavailability in these two patients fell below 8% and to around 1% in a further patient with severe graft dysfunction. Clamping reduced the metabolic clearance of cyclosporin A by only 25% from a mean before clamping of 2.9 ml/min/kg to 2.3 ml/min/kg (n = 11). Oral cyclosporin A becomes a reliable means of maintaining therapeutic drug concentrations only after bioavailability increases in association with T-tube clamping and in the absence of severe liver dysfunction or cholestasis.

  15. [Treatment of transfusion-dependent nonsevere aplastic anemia with cyclosporine A plus ATG/ALG versus cyclosporine A plus androgens: a retrospective single center study].

    PubMed

    Song, L; Peng, G X; Wu, Z J; Zhang, L; Jing, L P; Zhou, K; Li, Y; Li, Y; Ye, L; Li, J P; Fan, H H; Zhao, X; Yang, W R; Yang, Y; Zhang, F K

    2016-11-14

    Objective: To determine whether cyclosporine A (CsA) plus androgens was as effective as the current standard immunosuppressive therapy (IST) for transfusion-dependent nonsevere aplastic anemia (TD-NSAA). Methods: The records of 125 consecutive TD-NSAA patients who were treated between Aug. 2007 and Sept. 2014, with either CsA plus androgen or ALG/ATG plus CsA regimen were reviewed. The 3-month and 6-month hematologic responses and survival were evaluated. Results: There were 125 TD-NSAA patients (70 were male and 55 female, 1.25∶1). Median age was 27 (6-66) years. There was no significant difference in early mortality between 48 treated by ATG/ALG plus CsA and 77 by CsA plus androgen patients (1/48 vs 0/77, P=0.384). Both the total hematologic response and the better hematological response rates at 3-month (70.8% vs 45.5%, P=0.006 and 27.1% vs 10.4%, P=0.015, respectively) and 6-month (75.0% vs 55.8%, P=0.031 and 41.7% vs 22.1% P =0.020, respectively) after treatment were much higher in the standard IST group than that in CsA plus androgen group. The median time to transfusion independent of 36.5 (0-149) days in the standard IST group was significantly shorter than 98 (14-180) days in CsA plus androgen group (P<0.001). Survival was comparable between the two groups (97.9% vs 100.0%, P=0.227). It was superior (71.2% vs 59.5%) but not significantly (P=0.227) in event-free survival in standard IST group. Conclusions: CsA plus androgen was inferior to the standard IST of ATG/ALG and CsA regimen in treating TD-NSAA in terms of the hematologic response and the quality of response, despite of comparable short-term survival.

  16. Cyclosporin A drug interactions. Screening for inducers and inhibitors of cytochrome P-450 (cyclosporin A oxidase) in primary cultures of human hepatocytes and in liver microsomes.

    PubMed

    Pichard, L; Fabre, I; Fabre, G; Domergue, J; Saint Aubert, B; Mourad, G; Maurel, P

    1990-01-01

    In previous papers we demonstrated that cyclosporin A (CsA) was specifically oxidized in rabbit and human liver by cytochrome P-450IIIA. We therefore anticipated that any drug that is an inducer or an inhibitor of this cytochrome should lead to interaction with CsA when given in association with it. In order to confirm this hypothesis, primary cultures of human hepatocytes and human liver microsomes were used to "reproduce" in vitro clinically significant interactions observed between CsA and drugs known either as specific inducers (i.e., rifampicin) or as specific inhibitors (i.e., erythromycin) of P-450IIIA. Our results were in close agreement with the clinical reports. Human hepatocytes maintained in primary cultures for 72 hr in the presence of 50 microM rifampicin exhibited increased levels of P-450IIIA, determined by Western blot using specific antibodies, and concomitant increase in CsA oxidase activity, determined by HPLC analysis of extra and intracellular media. Conversely, these cultures exhibited erythromycin concentration-dependent decreases in CsA oxidase activity when incubated in the presence of 5, 20, and 100 microM erythromycin. In addition, a Lineweaver-Burk analysis of the erythromycin-mediated inhibition of CsA oxidase activity in human liver microsomes revealed competitive inhibition (with Ki of 75 microM) as expected, this macrolide being a specific substrate of P-450IIIA. Using this experimental approach, 59 molecules representative of 17 different therapeutic classes were screened for inducers and inhibitors of CsA oxidase activity. Our results allowed us to elucidate the molecular mechanism of previously observed, but unexplained, drug interactions involving CsA, and to detect drugs that should interfere with CsA metabolism as inducers or inhibitors. Drugs detected as potential inducers of CsA oxidase included: rifampicin, sulfadimidine, phenobarbital, phenytoin, phenylbutazone, dexamethasone, sulfinpyrazone, and carbamazepine. Drugs

  17. Calcineurin phosphatase activity in T lymphocytes is inhibited by FK 506 and cyclosporin A.

    PubMed Central

    Fruman, D A; Klee, C B; Bierer, B E; Burakoff, S J

    1992-01-01

    The immunosuppressive agents cyclosporin A (CsA) and FK 506 bind to distinct families of intracellular proteins (immunophilins) termed cyclophilins and FK 506-binding proteins (FKBPs). Recently, it has been shown that, in vitro, the complexes of CsA-cyclophilin and FK 506-FKBP-12 bind to and inhibit the activity of calcineurin, a calcium-dependent serine/threonine phosphatase. We have investigated the effects of drug treatment on phosphatase activity in T lymphocytes. Calcineurin is expressed in T cells, and its activity can be measured in cell lysates. Both CsA and FK 506 specifically inhibit cellular calcineurin at drug concentrations that inhibit interleukin 2 production in activated T cells. Rapamycin, which binds to FKBPs but exhibits different biological activities than FK 506, has no effect on calcineurin activity. Furthermore, excess concentrations of rapamycin prevent the effects of FK 506, apparently by displacing FK 506 from FKBPs. These results show that calcineurin is a target of drug-immunophilin complexes in vivo and establish a physiological role for calcineurin in T-cell activation. Images PMID:1373887

  18. Improved dissolution and pharmacokinetic behavior of cyclosporine A using high-energy amorphous solid dispersion approach.

    PubMed

    Onoue, Satomi; Sato, Hideyuki; Ogawa, Kumiko; Kawabata, Yohei; Mizumoto, Takahiro; Yuminoki, Kayo; Hashimoto, Naofumi; Yamada, Shizuo

    2010-10-31

    The aim of the present investigation is to develop solid dispersion (SD) formulations of cyclosporine A (CsA) for improving the oral bioavailability of CsA. Amorphous SDs of CsA with eight hydrophilic polymers were prepared with wet-mill employing zirconia beads. The physicochemical properties were characterized with a focus on morphology, crystallinity, thermal behavior, dissolution, and interaction of CsA with co-existing polymer. Although CsA molecules were found to be amorphous in all wet-milled formulations, some SD formulations failed to improve the dissolution. Of all CsA formulations, SD using polymer with HPC(SSL) exhibited the largest improvement in dissolution behavior. Pharmacokinetic profiling of orally dosed CsA in rats was carried out using UPLC/ESI-MS. After the oral administration of HPC(SSL)-based SD, enhanced CsA exposure was observed with increases in C(max) and AUC of ca. 5-fold, and the variation in AUC was ca. 40% less than that of amorphous CsA. Infrared spectroscopic studies suggested an interaction between CsA and HPC(SSL), as evidenced by the conformational transition of CsA. From the improved dissolution and pharmacokinetic data, the amorphous SD approach using wet-milling technology should lead to consistent and enhanced bioavailability, leading to an improved therapeutic potential of CsA.

  19. Prevention of murine cerebral malaria by low-dose cyclosporin A.

    PubMed Central

    Grau, G E; Gretener, D; Lambert, P H

    1987-01-01

    The effects of cyclosporin A (CsA) were investigated in an experimental model of cerebral malaria. In this model, Plasmodium berghei ANKA-infected CBA/Ca mice develop a clinically and histologically characterized neurological syndrome which is considered to be the result of immunopathological reactions mediated by L3T4+ T cells. It was shown that CsA displayed a strong protective effect on neurological complications when given at a dose 1 mg/kg/day for 5 consecutive days (Days 4-8), which had no effect on the parasite. Paradoxically, this protection against neurological complications was not seen when parasiticidal doses were used during this limited 5-day period. A similar protective effect was observed with two CsA derivatives, C5-34 and H7-94. The mechanisms by which CsA and the two derivatives could prevent murine cerebral malaria are unknown but can be related to exquisite effects on some lymphocyte functions. In view of these results, it might be conceivable to investigate the benefits of using low doses of CsA in man, in conjunction with the classical antiparasite therapy, for the management of cerebral malaria. PMID:3327806

  20. Long-term salvage therapy with cyclosporin A in refractory idiopathic thrombocytopenic purpura.

    PubMed

    Emilia, Giovanni; Morselli, Monica; Luppi, Mario; Longo, Giuseppe; Marasca, Roberto; Gandini, Giovanna; Ferrara, Leonardo; D'Apollo, Nicola; Potenza, Leonardo; Bertesi, Marcello; Torelli, Giuseppe

    2002-02-15

    Treatment of severe, chronic idiopathic thrombocytopenic purpura (ITP) refractory to most usual therapies is a difficult challenge. Little information exists on the clinical use of cyclosporin A (CyA) in the treatment of ITP. This report describes long-term treatment with CyA (median, 40 months) and follow-up (median, 36.8 months) in 12 adult patients with resistant ITP. CyA used in relatively low doses (2.5-3 mg/kg of body weight per day) led to a clinical improvement in 10 patients (83.3%). Five had a complete response (41.1%), 4 a complete response to maintenance therapy (33.3%), and one a partial response (8.3%). Two patients had no response. Most patients with a response (60%) had a long-term remission (mean, 28.6 months) after discontinuation of CyA. One patient had a relapse of ITP 4 years after CyA therapy was stopped. Side effects were moderate and transient, even in patients dependent on continued CyA treatment. CyA seems to represent reasonable salvage treatment in severe, potentially life-threatening, refractory ITP.

  1. Suppression of delayed-type hypersensitivity reactions and lymphokine production by cyclosporin A in the mouse.

    PubMed Central

    Thomson, A W; Moon, D K; Nelson, D S

    1983-01-01

    Two consecutive daily i.m. injections of cyclosporin A (Cs A) (greater than 50 mg/kg) inhibited delayed type hypersensitivity (DTH) responses in mice immunized with SRBC. Maximal suppression was observed when Cs A was administered 24 and 48 h after sensitization. Culture of spleen cells from these animals with antigen, insoluble concanavalin A (iCon A) or PHA revealed inhibition of the production of two lymphokines: that inducing macrophage procoagulant activity (MPCA) and macrophage chemotactic factor (LDCF). The inhibitory effect on lymphokine production was not due to depletion of T cells. In vitro, 25 ng/ml Cs A suppressed T cell proliferative responses to antigen and mitogen but much higher doses were required to impair the response to LPS. Similar doses of Cs A also suppressed lymphokine production, but the responses of macrophages to these lymphokines was unaffected, even at doses which totally inhibited lymphokine production. Production of interleukin 1 by LPS stimulated macrophages was inhibited by Cs A only at concentrations much greater than those required to suppress lymphokine production. PMID:6872317

  2. Cyclosporine A-induced hyperactivity in rats: is it mediated by immunosuppression, neurotrophism, or both?

    PubMed

    Borlongan, C V; Stahl, C E; Fujisaki, T; Sanberg, P R; Watanabe, S

    1999-01-01

    Cyclosporine A (CsA) immunosuppressive treatment has become an adjunctive therapy in neural transplantation of dopamine-secreting cells for treatment of Parkinson's disease (PD). Recently, CsA and its analogues have been shown to promote trophic effects against neurodegenerative disorders, and therefore CsA may have direct beneficial effects on dopaminergic neurons and dopamine-mediated behaviors. The present study examined the interaction between the reported CsA-induced hyperactivity and the possible alterations in nigral tyrosine hydroxylase (TH)-immunoreactive neurons in rats with damaged blood-brain barrier. CsA was administered at a therapeutic dose (10 mg/kg/day, IP, for 9 days) used in neural transplantation protocol for PD animal models. CsA-treated animals displayed significantly higher general spontaneous locomotor activity than control animals at drug injection days 7 and 9. Histological assays at day 9 revealed that there was a significant increase in TH-immunoreactive neurons in the nigra of CsA-treated rats compared to that of the vehicle-treated rats. The nigral TH elevation was accompanied by suppressed calcium-phosphotase calcineurin activity, indicating an inhibition of host immune response. This is the first report of CsA exerting simultaneous immunosuppressive and neurotrophic effects, as well as increasing general spontaneous locomotor behavior. These results support the utility of CsA as a therapeutic agent for PD and other movement disorders.

  3. Cyclosporin A inhibits calcineurin (phosphatase 2B) and P-glycoprotein activity in Entamoeba histolytica.

    PubMed

    Carrero, Julio C; Lugo, Haydee; Pérez, D Guillermo; Ortiz-Martínez, César; Laclette, Juan P

    2004-08-01

    Cyclosporin A (CsA) inhibits the proliferation of several protozoan parasites through blocking the activity of calcineurin (Cn) or P-glycoproteins (Pgp). We report here, that inhibition of the proliferation of Entamoeba histolytica trophozoites, the causal agent of human amebiasis, is due to interference of the phosphatase activity of Cn, in a similar fashion to the effect of this immunosuppressive drug on T lymphocytes. The non-immunosuppressive CsA analog PSC-833, which binds Pgp without interfering the function of Cn, did not inhibit the proliferation of HM1:IMSS trophozoites. Moreover, phosphatase activity of amebic Cn, detected using the phosphopeptide RII, was drastically affected by incubation with CsA, but not with PSC-833. On the other hand, both drugs were also tested on clone C2 trophozoites, which grow in the presence of emetine due to over-expression of Pgp. The effect of CsA was similar to that observed on HM1:IMSS trophozoites, whereas PSC-833 only affected the proliferation and viability of clone C2 when the trophozoites were grown in the presence of 40 microM of emetine, suggesting an interference of the Pgp activity. This suggestion was confirmed by results from experiments of Pgp-dependent effux of rhodamine from pre-loaded trophozoites, in the presence of either of these drugs. Therefore, CsA inhibition of E. histolytica trophozoite proliferation is more likely due to Cn than Pgp activity inhibition.

  4. Safety of the concomitant use of caspofungin and cyclosporin A in patients with invasive fungal infections.

    PubMed

    Sanz-Rodriguez, C; Lopez-Duarte, M; Jurado, M; Lopez, J; Arranz, R; Cisneros, J-M; Martino, M L; Garcia-Sanchez, P J; Morales, P; Olivé, T; Rovira, M; Solano, C

    2004-07-01

    Caspofungin, an echinocandin antifungal agent, is active against invasive Aspergillus and Candida infections. In a phase I study in healthy volunteers, mild transient increases in serum aminotransferases were observed with the concomitant administration of caspofungin and cyclosporin A (CsA). As a result, it is recommended that the concomitant use of the two drugs be limited to those settings with appropriate risk-benefit balance. We retrospectively assessed safety data in 14 patients with refractory invasive mycoses who were treated concomitantly with CsA and caspofungin before the drug was licensed in Spain. In all, 13 patients were adults (median age, 31.5 years; range, 14-67 years). The average duration of concomitant therapy was 15 days (range, 2-43 days). No clinically significant elevations of serum aminotransferases were observed, and no patient had concomitant therapy discontinued or interrupted due to a drug-related adverse event. In this study of a limited number of patients, the coadministration of caspofungin and CsA was generally well tolerated.

  5. Preparation, characterization and in silico modeling of biodegradable nanoparticles containing cyclosporine A and coenzyme Q10

    NASA Astrophysics Data System (ADS)

    Ankola, D. D.; Durbin, E. W.; Buxton, G. A.; Schäfer, J.; Bakowsky, U.; Kumar, M. N. V. Ravi

    2010-02-01

    Combination therapy will soon become a reality, particularly for those patients requiring poly-therapy to treat co-existing disease states. This becomes all the more important with the increasing cost, time and complexity of the drug discovery process prompting one to look at new delivery systems to increase the efficacy, safety and patient compliance of existing drugs. Along this line, we attempted to design nano-scale systems for simultaneous encapsulation of cyclosporine A (CsA) and coenzyme Q10 (CoQ10) and model their encapsulation and release kinetics. The in vitro characterization of the co-encapsulated nanoparticles revealed that the surfactant nature, concentration, external phase volume, droplet size reduction method and drug loading concentration can all influence the overall performance of the nanoparticles. The semi-quantitative solubility study indicates the strong influence of CoQ10 on CsA entrapment which was thought to be due to an increase in the lipophilicity of the overall system. The in vitro dissolution profile indicates the influence of CoQ10 on CsA release (64%) to that of individual particles of CsA, where the release is faster and higher (86%) on 18th day. The attempts to model the encapsulation and release kinetics were successful, offering a possibility to use such models leading to high throughput screening of drugs and their nature, alone or in combination for a particular polymer, if chi-parameters are understood.

  6. Spore inoculum optimization to maximize cyclosporin A production in Tolypocladium niveum.

    PubMed

    Lee, Mi-Jin; Lee, Han-Na; Han, Kyubeom; Kim, Eung-Soo

    2008-05-01

    The cyclic undecapeptide, cyclosporin A (CyA), is one of the most commonly prescribed immunosuppressive drugs. It is generated nonribosomally from a multifunctional cyclosporin synthetase enzyme complex by the filamentous fungus Tolypocladium niveum. In order to maximize the production of CyA by wild-type T. niveum (ATCC 34921), each of three culture stages (sporulation culture, growth culture, and production culture) were sequentially optimized. Among the three potential sporulation media, the SSMA medium generated the highest numbers of T. niveum spores. The SSM and SM media were then selected as the optimal growth and production culture media, respectively. The addition of valine and fructose to the SM production medium was also determined to be crucial for CyA biosynthesis. In this optimized three-stage culture system, 3% of the spore inoculum generated the highest level of CyA productivity in a 15-day T. niveum production culture, thereby implying that the determination of an appropriate size of T. niveum spore inoculum plays a critical role in the maximization of CyA production.

  7. Experimental immunologically mediated aplastic anemia (AA) in mice: cyclosporin A fails to protect against AA

    SciTech Connect

    Knospe, W.H.; Steinberg, D.; Gratwohl, A.; Speck, B.

    1984-07-01

    Immunologically mediated aplastic anemia (AA) in mice was induced by the i.v. injection of 10(7) lymph node cells (LNC) from H-2k identical but Mls mismatched CBA/J donor mice into previously irradiated (600 rad total body gamma) C3H/HeJ mice. Cyclosporin A (CsA), 25 mg/kg, was administered subcutaneously from day -1 to day 30. Control mice included C3H/HeJ mice which received 600 rad alone, C3H/HeJ mice which received 600 rad plus CsA as above, and C3H/HeJ mice which received 600 rad total body irradiation followed by 10(7) LNC from CBA/J donors. CsA failed to prevent lethal AA. These results suggest that the pathogenetic mechanisms operating in immunologically mediated AA differ from the mechanisms operating in rodents transplanted with allogeneically mismatched marrow or spleen cells which develop graft-versus-host disease. The results are consistent with a non-T cell-dependent mechanism causing the AA.

  8. Comparison of the effects of FK-506, cyclosporin A and rapamycin on IL-2 production.

    PubMed Central

    Henderson, D J; Naya, I; Bundick, R V; Smith, G M; Schmidt, J A

    1991-01-01

    The immunosuppressive compounds FK-506, cyclosporin A (CsA) and rapamycin inhibit both the human and mouse mixed lymphocyte reactions (MLR) with IC50s of 2-5 x 10(-10) M for FK-506 and rapamycin and 10(-8) M for CsA. FK-506 and CsA were also potent inhibitors of A23187/PMA-stimulated IL-2 production by Jurkat and HuT-78 cells but had no effect on the response of mouse CTLL cells to IL-2. IC50 values for inhibition of IL-2 production closely matched those for inhibition of the MLR and both drugs were active only during the first 4-6 hr following stimulation. In contrast, rapamycin was a poor inhibitor of IL-2 production, although it inhibited cellular responses to IL-2. The IC50 values for these two activities indicated that neither alone accounted for rapamycin inhibition of the MLR. FK-506 and CsA affected IL-2 gene transcription in Jurkat cells by the same mechanism. Both inhibited the appearance of the transcription factor, NFAT, whereas rapamycin did not. The appearance of another transcription factor, NFK beta, was unaffected by all three drugs. The effects of FK-506 and CsA on IL-2 gene expression, therefore, are similar even though the two drugs act through distinct cytosolic receptors. Images Figure 4 PMID:1715317

  9. Statistical evaluation and experimental design of a psoriasis xenograft transplantation model treated with cyclosporin A.

    PubMed

    Stenderup, Karin; Rosada, Cecilia; Alifrangis, Lene; Andersen, Søren; Dam, Tomas Norman

    2011-05-01

    Psoriasis xenograft transplantation models where human skin is transplanted onto immune-deficient mice are generally accepted in psoriasis research. Over the last decade, they have been widely employed to screen for new therapeutics with a potential anti-psoriatic effect. However, experimental designs differ in several parameters. Especially, the number of donors and grafts per experimental design varies greatly; numbers that are directly related to the probability of detecting statistically significant drug effects. In this study, we performed a statistical evaluation of the effect of cyclosporine A, a recognized anti-psoriatic drug, to generate a statistical model employable to simulate different scenarios of experimental designs and to calculate the associated statistical study power, defined as the probability of detecting a statistically significant anti-psoriatic drug treatment effect. Results showed that to achieve a study power of 0.8, at least 20 grafts per treatment group and a minimum of five donors should be included in the chosen experimental setting. To our knowledge, this is the first time that study power calculations have been performed to evaluate treatment effects in a psoriasis xenograft transplantation model. This study was based on a defined experimental protocol, thus other parameters such as drug potency, treatment protocol, mouse strain and graft size should, also, be taken into account when designing an experiment. We propose that the results obtained in this study may lend a more quantitative support to the validity of results obtained when exploring new potential anti-psoriatic drug effects.

  10. Cyclosporin-A-induced prion protein aggresomes are dynamic quality-control cellular compartments.

    PubMed

    Ben-Gedalya, Tziona; Lyakhovetsky, Roman; Yedidia, Yifat; Bejerano-Sagie, Michal; Kogan, Natalya M; Karpuj, Marcela Viviana; Kaganovich, Daniel; Cohen, Ehud

    2011-06-01

    Despite the activity of cellular quality-control mechanisms, subsets of mature and newly synthesized polypeptides fail to fold properly and form insoluble aggregates. In some cases, protein aggregation leads to the development of human neurodegenerative maladies, including Alzheimer's and prion diseases. Aggregates of misfolded prion protein (PrP), which appear in cells after exposure to the drug cyclosporin A (CsA), and disease-linked PrP mutants have been found to accumulate in juxtanuclear deposition sites termed 'aggresomes'. Recently, it was shown that cells can contain at least two types of deposition sites for misfolded proteins: a dynamic quality-control compartment, which was termed 'JUNQ', and a site for terminally aggregated proteins called 'IPOD'. Here, we show that CsA-induced PrP aggresomes are dynamic structures that form despite intact proteasome activity, recruit chaperones and dynamically exchange PrP molecules with the cytosol. These findings define the CsA-PrP aggresome as a JUNQ-like dynamic quality-control compartment that mediates the refolding or degradation of misfolded proteins. Together, our data suggest that the formation of PrP aggresomes protects cells from proteotoxic stress.

  11. Cyclosporin A in the treatment of refractory immune thrombocytopenia purpura in children.

    PubMed

    Gesundheit, B; Cividalli, G; Freeman, A; Yatziv, S; Koren, G; Baruchel, S

    2001-05-01

    Patients with refractory autoimmune thrombocytopenia do not respond to standard therapy with high-dose corticosteroids, intravenous immunoglobulin, and splenectomy. We describe the cases of two patients with refractory autoimmune thrombocytopenia treated with oral cyclosporin A (CsA) to evaluate the efficacy of this alternative therapy. Blood pressure and hepatic and renal function were in the normal range before initiation of treatment. Induction therapy with pulses of high-dose methylprednisolone was used for 3 consecutive days to improve the initial immune suppression. Gradual dose reduction of CsA, according the platelet count, minimized the long-term adverse effects of CsA. Oral CsA with pulses of high-dose methylprednisolone induced remission of the thrombocytopenia. Gradual weaning of CsA over months, according the platelet count, produced no observable adverse effects of the CsA. Rapid dose reduction caused thrombocytopenia, which resolved with higher dosages of CsA. Our cases show the efficacy of CsA for refractory immune thrombocytopenia. This therapeutic option with oral CsA as an additional salvage option may avoid splenectomy and the adverse effects of long-term corticosteroids. Larger clinical investigations are necessary to establish the indications and therapeutic regimen for CsA in immune thrombocytopenia.

  12. Calcineurin-independent inhibition of mitochondrial Ca2+ uptake by cyclosporin A

    PubMed Central

    Montero, M; Lobatón, C D; Gutierrez-Fernández, S; Moreno, A; Alvarez, J

    2003-01-01

    Cyclosporin A (CsA) is a widely used compound because of its potent immunosupressive properties, derived mainly from the inhibition of calcineurin, and also because of its ability to block the mitochondrial permeability transition pore (PTP). This second effect has been involved in the protection against apoptosis mediated by release of mitochondrial factors. We show here that CsA (1–10μM) has an additional effect on Ca2+ homeostasis in mitochondria that cannot be attributed to inhibition of PTP. By measuring specifically mitochondrial [Ca2+] with targeted aequorin, we show that CsA inhibited Ca2+ entry into mitochondria both in intact and in permeabilized cells, and this effect was stronger when Ca2+ entry was triggered by low cytosolic [Ca2+], below 5 μM. Inhibition of mitochondrial Ca2+ uptake required micromolar concentrations of CsA and was not mimicked by other inhibitors of calcineurin such as FK-506 or cypermethrin, nor by a different inhibitor of the PTP, bongkrekic acid. CsA blocked the increase in mitochondrial Ca2+ uptake rate induced by the mitochondrial Ca2+ uniporter activator SB202190. Our results suggest that CsA inhibits Ca2+ entry through the Ca2+ uniporter by a mechanism independent of the inhibition of PTP or calcineurin. This effect may contribute to reduce depolarization and Ca2+ overloading in mitochondria after cell stimulation, and thus cooperate with the direct inhibition of PTP to prevent apoptosis. PMID:14691054

  13. Calcineurin-independent inhibition of mitochondrial Ca2+ uptake by cyclosporin A.

    PubMed

    Montero, M; Lobatón, C D; Gutierrez-Fernández, S; Moreno, A; Alvarez, J

    2004-01-01

    1. Cyclosporin A (CsA) is a widely used compound because of its potent immunosupressive properties, derived mainly from the inhibition of calcineurin, and also because of its ability to block the mitochondrial permeability transition pore (PTP). This second effect has been involved in the protection against apoptosis mediated by release of mitochondrial factors. We show here that CsA (1-10 microm) has an additional effect on Ca(2+) homeostasis in mitochondria that cannot be attributed to inhibition of PTP. 2. By measuring specifically mitochondrial [Ca(2+)] with targeted aequorin, we show that CsA inhibited Ca(2+) entry into mitochondria both in intact and in permeabilized cells, and this effect was stronger when Ca(2+) entry was triggered by low cytosolic [Ca(2+)], below 5 microm. 3. Inhibition of mitochondrial Ca(2+) uptake required micromolar concentrations of CsA and was not mimicked by other inhibitors of calcineurin such as FK-506 or cypermethrin, nor by a different inhibitor of the PTP, bongkrekic acid. 4. CsA blocked the increase in mitochondrial Ca(2+) uptake rate induced by the mitochondrial Ca(2+) uniporter activator SB202190. 5. Our results suggest that CsA inhibits Ca(2+) entry through the Ca(2+) uniporter by a mechanism independent of the inhibition of PTP or calcineurin. This effect may contribute to reduce depolarization and Ca(2+) overloading in mitochondria after cell stimulation, and thus cooperate with the direct inhibition of PTP to prevent apoptosis.

  14. Prolactin and prolactin secretagogues reverse immunosuppression in mice treated with cysteamine, glucocorticoids, or cyclosporin-A.

    PubMed

    Bernton, E; Bryant, H; Holaday, J; Dave, J

    1992-12-01

    Suppression of prolactin (PRL) secretion with the dopamine agonist, bromocriptine, has been shown in rodents to diminish a variety of immunologic responses, including delayed type hypersensitivity, primary antibody response, T-cell dependent macrophage activation, and ex vivo T- and B-lymphocyte proliferation in response to mitogens. These same responses can be suppressed by endogenous or exogenous glucocorticosteroids and, in large measure, the immunosuppressant peptide cyclosporin A. The sulfhydryl reducing agent cysteamine (2-aminoethanethiol) is known to reduce pituitary and plasma prolactin levels. Treatment of mice with cysteamine at doses which suppressed circulating PRL levels resulted in suppression of ex vivo blastogenic responses of lymphocytes from treated mice. The T-cell-dependent primary IgM response to immunization with sheep red blood cells was also suppressed by cysteamine treatment. Treatment of mice with drugs stimulating the release of endogenous PRL, or with exogenous ovine PRL, was found to antagonize the suppression of lymphocyte proliferative responses to mitogens induced in mice by glucocorticoid or cyclosporin treatment. These data suggest that many drugs in common clinical use could have potential immunomodulatory actions due to suppression or stimulation of pituitary PRL secretion. Furthermore, lactogenic hormones appear to exert counterregulatory actions which may modify glucocorticosteroid actions on immune and other target issues.

  15. Effect of Cyclosporin A and Angiotensin II on cytosolic calcium levels in primary human gingival fibroblasts

    PubMed Central

    Supraja, Ajitkumar; Dinesh, Murugan Girija; Rajasekaran, Subbarayan; Balaji, Thodur Madapusi; Rao, Suresh Ranga

    2016-01-01

    Background: To evaluate the effect of Cyclosporin A (CsA) and angiotensin II (Ang II) on cytosolic calcium levels in cultured human gingival fibroblasts (HGFs). Materials and Methods: Healthy gingival samples from six volunteers were obtained, and primary HGFs were cultured. Cell viability and proliferation assay were performed to identify the ideal concentrations of CsA and Ang II. Cytosolic calcium levels in cultured gingival fibroblasts treated with CsA and Ang II were studied using colorimetric assay, confocal and fluorescence imaging. Statistical analyses were done using SPSS software and GraphPad Prism. Results: Higher levels of cytosolic levels were evident in cells treated with CsA and Ang II when compared to control group and was statistically significant (P < 0.05) in both colorimetric assay and confocal imaging. Fluorescent images of the cultured HGFs revealed the same. Conclusion: Thus calcium being a key player in major cellular functions, plays a major role in the pathogenesis of drug-induced gingival overgrowth. PMID:27857765

  16. Identification of novel indicators of cyclosporine A nephrotoxicity in a CD-1 mouse model

    SciTech Connect

    O'Connell, Sein; Slattery, Craig; Ryan, Michael P.; McMorrow, Tara

    2011-04-15

    The calcineurin inhibitor cyclosporine A (CsA) is a widely used immunosuppressive agent. However, nephrotoxicity is a serious side effect observed in patients which limits clinical use of CsA. CsA nephrotoxicity is associated with tubulointerstitial injury progressing to nephropathy. This is typically diagnosed by invasive renal biopsy and is often only detected when the disease process is well advanced. Therefore identification of novel, early indicators of CsA nephrotoxicity could be clinically advantageous. This study aimed to establish a murine model of CsA nephrotoxicity and to identify urinary proteins that may indicate the onset of CsA-induced nephropathy using 2-D gel electrophoresis. CsA nephrotoxicity was induced in CD-1 mice by daily CsA administration for 4 weeks. By week 4, elevated serum creatinine and proteinuria were observed after CsA treatment indicating significant renal dysfunction. Decreased cadherin-1, increased {alpha}-smooth muscle actin and fibroblast specific protein 1 in kidney tissue indicated disruption of normal tubular architecture. Alterations in podocin and uromodulin were also observed which may indicate damage to other segments of the nephron. Proteomic analysis of urine identified a number of differentially regulated proteins that may be involved in early CsA nephropathy including cadherin 1, superoxide dismutase and vinculin. These findings suggest novel mechanisms of CsA nephrotoxicity and identify novel potential markers of the disease.

  17. Effects of erdosteine on cyclosporine-A-induced hepatotoxicity in rats.

    PubMed

    Erarslan, Elife; Ekiz, Fuat; Uz, Burak; Koca, Cemile; Turkcu, Ummuhani Ozel; Bayrak, Reyhan; Delibasi, Tuncay

    2011-01-01

    Cyclosporine A (CsA) is a potent immunosuppressive agent used for organ transplantations and various autoimmune disorders. However, hepatotoxicity due to CsA remains one of the major side effects. The use of antioxidants reduces the adverse effects of CsA. The aim of this study was to determine the protective effects of erdosteine on CsA-induced liver injury through tissue oxidant/antioxidant parameters and to evaluate light microscopic alterations in rat-liver tissues. Rats were randomly divided into four experimental groups: The control group received sunflower oil (2 mL/kg/day, per orally; p.o.), while the other groups were treated with CsA (25 mg/kg/day, p.o.) or erdosteine (10 mg/kg/day, p.o.) or CsA+erdosteine, respectively. Serum aspartate aminotransferase and alanine aminotransferase levels, tissue malondialdehyde and nitric oxide levels, and superoxide dismutase, glutathione peroxidase and catalase enzyme activities were measured. Histological examination was performed. CsA caused a significant deterioration in the hepatic function tests, morphology, and gave rise to severe oxidative stress in the liver. Erdostein significantly improved the functional and histological parameters and attenuated the oxidative stresss induced by CsA. Erdostein protects liver tissue against oxygen free radicals and prevents hepatic dysfunction and morphological abnormalities associated with chronic CsA administration.

  18. A case of fatal systemic toxoplasmosis in a cat being treated with cyclosporin A for feline atopy.

    PubMed

    Last, Robert D; Suzuki, Yasuhiro; Manning, Thomas; Lindsay, David; Galipeau, Laura; Whitbread, Trevor J

    2004-06-01

    Acute systemic toxoplasmosis was diagnosed in a 4-5-year-old, male, Domestic Short Hair cat, which had been on cyclosporine A immunomodulatory therapy for feline atopy, over an 8-month period. Cyclosporin A (CsA) has shown promising results as a immunosuppressive agent in the cat for the treatment of eosinophilic plaque and granulomas, allergic cervico-facial pruritus, feline atopy and other immune-mediated dermatoses. However, inhibition of T-lymphocyte function by CsA is believed to have predisposed this cat to the development of a newly acquired, acute Toxoplasma gondii infection, as characterized by severe hepatic and pancreatic pathology in conjunction with the heavy parasite load demonstrated on immunohistochemical (IHC) stains for T. gondii. Cats on CsA therapy appear to be at risk of developing fatal systemic toxoplasmosis.

  19. Inhibition of Human Immunodeficiency Virus and Growth of Infected T Cells by the Immunosuppressive Drugs Cyclosporin A and FK 506

    NASA Astrophysics Data System (ADS)

    Karpas, Abraham; Lowdell, Mark; Jacobson, S. Kim; Hill, Fergal

    1992-09-01

    The effects of the immunosuppressive drugs cyclosporin A and FK 506 were studied on cells chronically infected with human immunodeficiency virus type 1 (HIV-1) as well as on uninfected and newly infected cells. When cells chronically infected with HIV-1 or with HIV-2 were cocultivated with uninfected cells in the presence of cyclosporin A or FK 506 there was a delay in the formation of syncytia and of cytopathic effects. This inhibitory effect was not due to decreased membrane expression of CD4. In addition, there was an ≈100-fold reduction in the yield of infectious HIV-1 when the infected cells were grown in the presence of these drugs, a finding consistent with other evidence of decreased HIV expression. Both drugs were found to inhibit the growth of chronically infected cells at concentrations that did not inhibit the growth of the uninfected cells. These results, demonstrating that cyclosporin A and FK 506 interfere with HIV production and selectively inhibit the growth of infected cells, suggest that they may be useful in the treatment of this infection and indicate further cellular targets for antiviral agents.

  20. Oral Cyclosporin A Inhibits CD4 T cell P-glycoprotein Activity in HIV-Infected Adults Initiating Treatment with Nucleoside Reverse Transcriptase Inhibitors

    PubMed Central

    Hulgan, Todd; Donahue, John P.; Smeaton, Laura; Pu, Minya; Wang, Hongying; Lederman, Michael M.; Smith, Kimberly; Valdez, Hernan; Pilcher, Christopher; Haas, David W.

    2010-01-01

    Purpose P-glycoprotein limits tissue penetration of many antiretroviral drugs. We characterized effects of the P-glycoprotein substrate cyclosporin A on T cell P-glycoprotein activity in HIV-infected AIDS Clinical Trials Group study A5138 participants. Methods We studied P-glycoprotein activity on CD4 and CD8 T cells in 16 participants randomized to receive oral cyclosporin A (n=9) or not (n=7) during initiation antiretroviral therapy (ART) that did not include protease or non-nucleoside reverse transcriptase inhibitors. Results CD4 T cell P-glycoprotein activity decreased by a median of 8 percentage points with cyclosporin A/ART (difference between cyclosporin A/ART versus ART only P=0.001). Plasma trough cyclosporin A concentrations correlated with change in P-glycoprotein activity in several T cell subsets. Conclusions Oral cyclosporin A can inhibit peripheral blood CD4 T cell P-glycoprotein activity. Targeted P-glycoprotein inhibition might enhance delivery of ART to T cells. PMID:19779705

  1. Cyclosporin A-induced free radical generation is not mediated by cytochrome P-450

    PubMed Central

    Krauskopf, Alexandra; Buetler, Timo M; Nguyen, Nathalie S D; Macé, Katherine; Ruegg, Urs T

    2002-01-01

    Reactive oxygen species (ROS) have been proposed to play a role in the side effects of the immunosuppressive drug cyclosporin A (CsA). The aim of this study was to investigate whether cytochrome P-450 (CYP) dependent metabolism of CsA could be responsible for ROS generation since it has been suggested that CsA may influence the CYP system to produce ROS. We show that CsA (1 – 10 μM) generated antioxidant-inhibitable ROS in rat aortic smooth muscle cells (RASMC) using the fluorescent probe 2,7-dichlorofluorescin diacetate. Using cytochrome c as substrate, we show that CsA (10 μM) did not inhibit NADPH cytochrome P-450 reductase in microsomes prepared from rat liver, kidney or RASMC. CsA (10 μM) did not uncouple the electron flow from NADPH via NADPH cytochrome P-450 reductase to the CYP enzymes because CsA did not inhibit the metabolism of substrates selective for several CYP enzymes that do not metabolize CsA in rat liver microsomes. CsA (10 μM) did not generate more radicals in CYP 3A4 expressing immortalized human liver epithelial cells (T5-3A4 cells) than in control cells that do not express CYP 3A4. Neither diphenylene iodonium nor the CYP 3A inhibitor ketoconazole were able to block ROS formation in rat aortic smooth muscle or T5-3A4 cells. These results demonstrate that CYP enzymes do not contribute to CsA-induced ROS formation and that CsA neither inhibits NADPH cytochrome P-450 reductase nor the electron transfer to the CYP enzymes. PMID:11861326

  2. In vitro models to study mechanisms involved in cyclosporine A-mediated glomerular contraction.

    PubMed

    L'Azou, B; Medina, J; Frieauff, W; Cordier, A; Cambar, J; Wolf, A

    1999-08-01

    The immunosuppressive drug, cyclosporin A (CsA), which is successfully used to prevent rejection in organ transplantation, induces renal side-effects as shown by a decrease in glomerular filtration rate and ultrafiltration coefficient regulated by the tone of mesangial cells.The aim of the present study was to investigate the effect of CsA on isolated glomeruli and mesangial cells, which constitute appropriate in vitro models for renal vasoreactivity studies. The roles of different intracellular and extracellular mediators such as calcium, endothelin-1 (ET-1), prostaglandins (TXA(2 )and PGI(2)) and reactive oxygen intermediates (ROIs) were analysed. CsA caused a concentration- and time-dependent decrease in the planar cross-sectional areas of isolated glomeruli and mesangial cells as determined by image analysis. Intracytosolic free calcium concentration determined by fluorimetric analysis was significantly increased after 30 min CsA (10 microM) incubation. In the contraction experiment, the calcium antagonist verapamil inhibited the CsA response. ET-1, TXB(2) and keto-PGF(1alpha) were determined directly, however no changes were found statistically significantly different from respective controls. In contrast to these results, the ET-1 specific antibody was able to reduce CsA-mediated cell contraction. In the presence of a prostacyclin agonist iloprost, CsA-induced contraction was also modified. The role of ROIs using a 2'7'-dichlorofluorescein diacetate (DCFdAc) fluorimetric method was directly determined by observing, with 10 microM CsA, a significant production of hydrogen peroxide (H(2)O(2)), which was able alone to induce mesangial cell contraction. Coincubation with the antioxidants led to a significant inhibition of mesangial cell contraction. These results suggest that CsA caused an imbalance in the normal level of all investigated vasoconstrictive and vasodilator mediators, which shifted towards the advantage of vasoconstrictive action.

  3. Amorphous solid dispersion of cyclosporine A prepared with fine droplet drying process: Physicochemical and pharmacokinetic characterization.

    PubMed

    Suzuki, Hiroki; Moritani, Tatsuru; Morinaga, Tadahiko; Seto, Yoshiki; Sato, Hideyuki; Onoue, Satomi

    2017-03-15

    The present study aimed to develop an amorphous solid dispersion (ASD) of cyclosporine A (CsA) by a fine droplet drying (FDD) process for improvement in oral absorption of CsA. CsA and hydroxypropyl cellulose-SSL were dissolved in 1,4-dioxane, and the solution was powdered by the FDD process to obtain the ASD formulation of CsA (ASD/CsA). The ASD/CsA was characterized in terms of morphology, particle size distribution, crystallinity, dissolution behavior, physicochemical stability, and pharmacokinetic behavior in rats. The ASD/CsA was obtained in the form of uniform spherical particles, and the span factor was calculated to be ca. 0.4. CsA in the formulation existed in an amorphous state. The ASD/CsA exhibited a higher dissolution behavior of CsA than amorphous CsA, whereas storage of the ASD/CsA under accelerated conditions led to impairment in the dissolution behavior. The constant release of CsA from non-aged ASD/CsA was observed during dissolution testing. After oral administration of CsA samples (10mg-CsA/kg) in rats, the ASD/CsA showed a high and sustained plasma concentration of CsA as evidenced by a 18-fold increase in the oral bioavailability of CsA compared with amorphous CsA. From these findings, the FDD process might be an efficacious option for the ASD formulation of CsA with enhanced biopharmaceutics properties.

  4. Induction of synapse associated protein 102 expression in cyclosporin A-stimulated hair growth.

    PubMed

    Kim, Chang Deok; Lee, Min-Ho; Sohn, Kyung-Cheol; Kim, Jin-Man; Li, Sheng Jin; Rang, Moon-Jeong; Roh, Seok-Seon; Oh, Young-Seon; Yoon, Tae-Jin; Im, Myung; Seo, Young-Joon; Lee, Jeung-Hoon; Park, Jang-Kyu

    2008-08-01

    Cyclosporin A (CsA) has been used as a potent immunosuppressive agent for inhibiting the graft rejection after organ transplantation. However, CsA provokes lots of side effects including hirsutism, the phenomenon of abnormal hair growth in the body. In the present study, we investigated the hair growth stimulating effect of CsA using in vivo and in vitro test models. When topically applied on the back skin of mice, CsA induced fast telogen to anagen transition. In contrast, CsA had no effect on the growth of human hair follicle tissues cultured in vitro, indicating that it might not have the mitogenic effect on hair follicles. To identify the genes related with CsA-induced hair growth, we performed differential display RT-PCR. Among the genes obtained, the expression of synapse associated protein 102 (SAP102) was verified using competitive RT-PCR. The result showed that the expression of SAP102 was significantly induced by CsA treatment in the back skin of C57BL/6 mice. However, the increase of SAP102 mRNA was also seen in spontaneous anagen mice, suggesting that induction of SAP102 is one event of the anagen hair growth response regardless of how the growth state was induced. SAP102 was not expressed in cultured human hair outer root sheath and dermal papilla cells. Immunohistochemistry analysis showed that CsA induced the expression of SAP102 in perifollicular region of mouse anagen hair. Together, these results suggest that SAP102 is one of hair-cycle-dependent genes, whose expression is related with the anagen progression.

  5. Attenuation of cyclosporine A toxicity by sublethal heat shock. Role of catalase.

    PubMed

    Andrés, David; Bautista, Mirandeli; Cascales, María

    2005-02-01

    Cyclosporine A (CsA) is the immunosuppressor most frequently used in transplant surgery and in the treatment of autoimmune diseases because of its specific inhibiting effect on signal transduction pathways of cell T receptor. It has been shown that CsA is able to generate reactive oxygen species and lipid peroxidation, which are directly involved in the CsA hepatotoxicity. In the present study, we investigated the effect of a sublethal heat pre-treatment (43 degrees C for 30 min) on the hepatoma cell line HepG2 exposed to cytotoxic concentrations of CsA (10 and 25 microM) for 3 and 24 h. Parameters of cytotoxicity were assayed by measuring LDH (lactate dehydrogenase) leakage into the medium. Peroxide concentration was tested by flow cytometry by measuring the fluorescence intensity of DCF (dichlorofluorescein). Gene expression of catalase was detected by measuring the respective mRNA and proteins, as well as protein level of HSP70. The enzymatic activity of catalase was also determined. Heat pre-treatment significantly reduced CsA cytotoxicity as well as the level of peroxide generation. The protective effect of the previous heat treatment (corroborated by the irreversible catalase inhibitor 3-aminotriazole) against the CsA cytotoxicity was due to an increased expression and activity of catalase that was significantly reduced by the effect of CsA. We conclude that heat pre-treatment strongly protects against CsA injury, and the mechanism of this protection is by means of inducing not only the expression of HSP70 but also the expression and activity of catalase, the main enzyme system involved in H(2)O(2) elimination.

  6. A Multidisciplinary Evaluation of the Effectiveness of Cyclosporine A in Dystrophic Mdx Mice

    PubMed Central

    De Luca, Annamaria; Nico, Beatrice; Liantonio, Antonella; Paola Didonna, Maria; Fraysse, Bodvael; Pierno, Sabata; Burdi, Rosa; Mangieri, Domenica; Rolland, Jean-François; Camerino, Claudia; Zallone, Alberta; Confalonieri, Paolo; Andreetta, Francesca; Arnoldi, Elisa; Courdier-Fruh, Isabelle; Magyar, Josef P.; Frigeri, Antonio; Pisoni, Michela; Svelto, Maria; Conte-Camerino, Diana

    2005-01-01

    Chronic inflammation is a secondary reaction of Duchenne muscular dystrophy and may contribute to disease progression. To examine whether immunosuppressant therapies could benefit dystrophic patients, we analyzed the effects of cyclosporine A (CsA) on a dystrophic mouse model. Mdx mice were treated with 10 mg/kg of CsA for 4 to 8 weeks throughout a period of exercise on treadmill, a protocol that worsens the dystrophic condition. The CsA treatment fully prevented the 60% drop of forelimb strength induced by exercise. A significant amelioration (P < 0.05) was observed in histological profile of CsA-treated gastrocnemius muscle with reductions of nonmuscle area (20%), centronucleated fibers (12%), and degenerating area (50%) compared to untreated exercised mdx mice. Consequently, the percentage of normal fibers increased from 26 to 35% in CsA-treated mice. Decreases in creatine kinase and markers of fibrosis were also observed. By electrophysiological recordings ex vivo, we found that CsA counteracted the decrease in chloride conductance (gCl), a functional index of degeneration in diaphragm and extensor digitorum longus muscle fibers. However, electrophysiology and fura-2 calcium imaging did not show any amelioration of calcium homeostasis in extensor digitorum longus muscle fibers. No significant effect was observed on utrophin levels in diaphragm muscle. Our data show that the CsA treatment significantly normalized many functional, histological, and biochemical endpoints by acting on events that are independent or downstream of calcium homeostasis. The beneficial effect of CsA may involve different targets, reinforcing the usefulness of immunosuppressant drugs in muscular dystrophy. PMID:15681831

  7. Antioxidant properties of repaglinide and its protections against cyclosporine A-induced renal tubular injury

    PubMed Central

    Li, Dao; Li, Jin; Li, Hui; Wu, Qiong; Li, Qi-Xiong

    2016-01-01

    Objective(s): Repaglinide (RG) is an antihyperglycemic agent used for the treatment of non-insulin-dependent diabetes mellitus. It has a good safety and efficacy profile in diabetic patients with complications in renal impairment and is an appropriate treatment choice, even for individuals with more severe degrees of renal malfunctions. The aim of the present study was to examine the protective effect of RG on cyclosporine A (CsA)-induced rat renal impairment and to evaluate the antioxidant mechanisms by which RG exerts its protective actions. Materials and Methods: Fifty male Sprague-Dawley rats weighing 250–300 g were randomly divided into five groups: administrations of olive oil (control, PO), RG (0.4 mg/kg, PO), CsA (30 mg/kg in olive oil, SC), RG (0.2 or 0.4 mg/kg, PO) plus CsA (30 mg/kg in olive oil SC) every day for 15 days. Results: SC administration of CsA (30 mg/kg) to rats produced marked elevations in the levels of renal impairment parameters such as urinary protein, N-acetyl-beta-D-glucosaminidase (NAG), serum creatinine (SCr), and blood urea nitrogen (BUN). It also caused histologic injury to the kidneys. Oral administration of RG (0.2 and 0.4 mg/kg) markedly decreased all the aforementioned changes. In addition, CsA caused increases in the levels of malondialdehyde (MDA) and decreases in superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), glutathione reductase (GSR), glutathione-S-transferase (GST), and glutathione in kidney homogenate, which were reversed significantly by both doses of RG. Conclusion: The findings of our study indicate that RG may play an important role in protecting the kidney from oxidative insult. PMID:27635199

  8. Cyclosporine A and palmitic acid treatment synergistically induce cytotoxicity in HepG2 cells.

    PubMed

    Luo, Yi; Rana, Payal; Will, Yvonne

    2012-06-01

    Immunosuppressant cyclosporine A (CsA) treatment can cause severe side effects. Patients taking immunosuppressant after organ transplantation often display hyperlipidemia and obesity. Elevated levels of free fatty acids have been linked to the etiology of metabolic syndromes, nonalcoholic fatty liver and steatohepatitis. The contribution of free fatty acids to CsA-induced toxicity is not known. In this study we explored the effect of palmitic acid on CsA-induced toxicity in HepG2 cells. CsA by itself at therapeutic exposure levels did not induce detectible cytotoxicity in HepG2 cells. Co-treatment of palmitic acid and CsA resulted in a dose dependent increase in cytotoxicity, suggesting that fatty acid could sensitize cells to CsA-induced cytotoxicity at the therapeutic doses of CsA. A synergized induction of caspase-3/7 activity was also observed, indicating that apoptosis may contribute to the cytotoxicity. We demonstrated that CsA reduced cellular oxygen consumption which was further exacerbated by palmitic acid, implicating that impaired mitochondrial respiration might be an underlying mechanism for the enhanced toxicity. Inhibition of c-Jun N-terminal kinase (JNK) attenuated palmitic acid and CsA induced toxicity, suggesting that JNK activation plays an important role in mediating the enhanced palmitic acid/CsA-induced toxicity. Our data suggest that elevated FFA levels, especially saturated FFA such as palmitic acid, may be predisposing factors for CsA toxicity, and patients with underlying diseases that would elevate free fatty acids may be susceptible to CsA-induced toxicity. Furthermore, hyperlipidemia/obesity resulting from immunosuppressive therapy may aggravate CsA-induced toxicity and worsen the outcome in transplant patients.

  9. Protective effect of schisandrin B against cyclosporine A-induced nephrotoxicity in vitro and in vivo.

    PubMed

    Zhu, Shaohua; Wang, Yan; Chen, Meiwan; Jin, Jing; Qiu, Yuwen; Huang, Min; Huang, Zhiying

    2012-01-01

    Schisandrin B (Sch B) is an active ingredient of the fruit of Schisandra chinensis. It has many therapeutic effects arising from its tonic, sedative, antitussive and antiaging activities and is also used in the treatment of viral and chemical hepatitis. The aim of this study was to investigate the protective effects of Sch B on cyclosporine A (CsA)-induced nephrotoxicity in mice and HK-2 cells (a human proximal tubular epithelial cell line). After gavage with Sch B (20 mg/kg) or olive oil (vehicle), mice received CsA (30 mg/kg) by subcutaneous injection once daily for four weeks. Renal function, histopathology, and tissue glutathione (GSH) and malondialdehyde (MDA) levels were evaluated after the last treatment. The effects of Sch B on CsA-induced oxidative damage in HK-2 cells were investigated by measuring cell viability, the release of lactate dehydrogenase (LDH), the level of reactive oxygen species (ROS), and the cellular GSH and ATP concentrations. Cellular apoptosis was assessed by flow cytometry. Treatment with Sch B in CsA-treated mice significantly suppressed the elevation of blood urea nitrogen (BUN) and serum creatinine levels and attenuated the histopathological changes. Additionally, Sch B also decreased renal MDA levels and increased GSH levels in CsA-treated mice. Using an in vitro model, Sch B (2.5, 5 and 10 μM) significantly increased the cell viability and reduced LDH release and apoptosis induced by CsA (10 μM) in HK-2 cells. Furthermore, Sch B increased the intracellular GSH and ATP levels and attenuated CsA-induced ROS generation. In conclusion, Sch B appears to protect against CsA-induced nephrotoxicity by decreasing oxidative stress and cell death.

  10. Precipitation of experimental autoallergic uveoretinitis by cyclosporin A withdrawal: an experimental model of uveitis relapse.

    PubMed Central

    Atkinson, E G; Dinning, W J; Kasp, E; Graham, E M; Dumonde, D C

    1989-01-01

    This study set out to determine whether withdrawal of cyclosporin A (CyA) in Lewis rats sensitized to retinal S antigen would precipitate experimental autoallergic uveoretinitis (EAU), and whether challenge of such animals with S antigen or an unrelated stimulus would accelerate EAU onset after drug withdrawal. Rats were sensitized with 50 micrograms S antigen in Freund's complete adjuvant (FCA) and EAU onset was suppressed by 18 days of treatment with CyA at doses ranging from 3 to 10 mg/kg daily. Without challenge, seven out of 11 animals developed EAU with a median onset of 78 days. This was reduced to 68 days in rats challenged on day 32 with FCA alone, to 48 days with 10 micrograms S antigen in FCA, and to 41 days with 50 micrograms S antigen in FCA. The incidence, onset and severity of anterior uveitis and extent of photoreceptor destruction were related to both CyA dose and nature of challenge. The extent of photoreceptor destruction ran parallel with severity of anterior uveitis; and delayed-type hypersensitivity reactivity on day 43 was related to both severity of anterior uveitis (P less than 0.001) and photoreceptor damage (P less than 0.002). At the highest dose, CyA also delayed the appearance of antibody to S antigen; however, subsequent antibody levels were unrelated to EAU severity or to nature of challenge. The results indicate that CyA-induced suppression of the immunological response to S antigen can recover spontaneously after drug withdrawal, that challenge with either S antigen or FCA alone can accelerate the subsequent onset of EAU, and that these phenomena may provide a basis for investigating mechanisms underlying relapse of human uveoretinitis. Images Fig. 5 PMID:2805414

  11. Metabolism of cyclosporin A. I. Study in freshly isolated rabbit hepatocytes

    SciTech Connect

    Fabre, G.; Bertault-Peres, P.; Fabre, I.; Maurel, P.; Just, S.; Cano, J.P.

    1987-05-01

    The metabolism of cyclosporin A (CsA), a widely used immunosuppressive agent, was evaluated in freshly isolated rabbit hepatocytes by HPLC which separated CsA from its major group of derivatives, e.g. first generation metabolites (monohydroxylated and N-demethylated) and second generation derivatives (dihydroxylated and dihydroxy-N-demethylated). After exposure of hepatocytes to radiolabeled CsA (0.5 mg/liter), CsA was rapidly accumulated inside the cells and metabolized. The dihydroxylated metabolites represent the major intracellular forms after 1 hr. CsA metabolites synthesized inside the cells are then rapidly detected in the extracellular compartment. Unchanged drug and the various metabolites are concentrated inside the cells with transmembrane chemical gradients ranging between 20:1 and 40:1. Transport and metabolic processes for CsA have been evaluated over the following CsA extracellular concentration range, 0.1-10 mg/liter. Metabolism appears to be the rate-limiting step. The apparent affinity constant of CsA for the enzyme system involved in its metabolism is approximately 15 microM. Besides the lipophilicity of the molecule, which is responsible for the retention of CsA and its metabolites in the intracellular compartment, the presence of a binding component(s) in the hepatocytes was also demonstrated. CsA and its metabolites seem to have similar affinities for this binding site. These studies demonstrate that CsA is rapidly transformed inside the hepatocytes to various metabolites which may play an important role in the pharmacological activity of the drug and/or in its clinical toxicity.

  12. Effect of Wuzhi tablet (Schisandra sphenanthera extract) on the pharmacokinetics of cyclosporin A in rats.

    PubMed

    Xue, Xin-ping; Qin, Xiao-ling; Xu, Chenshu; Zhong, Guo-ping; Wang, Ying; Huang, Min; Bi, Hui-chang

    2013-08-01

    In our previous reports, Wuzhi tablet (an herbal preparation of ethanol extract of Wuweizi (Schisandra sphenanthera)) can significantly increase the blood concentration of tacrolimus and paclitaxel in rats by inhibiting the CYP3A-mediated metabolism and the P-gp-mediated efflux. Cyclosporin A (CsA), a well-known immunosuppressant agent, is also a substrate of CYP3A and P-gp. Therefore, this study aimed to investigate whether and how WZ affects pharmacokinetics of CsA in rats. The AUC0-48 h and Cmax of CsA were increased by 40.1% and 13.1%, respectively, with a single oral co-administration of WZ and high dose of CsA (37.8 mg/kg). Interestingly, after a single oral co-administration of WZ and low dose of CsA (1.89 mg/kg), the AUC0-36 h and Cmax of CsA were dramatically increased by 293.1% (from 1103.2 ± 293.0 to 4336.5 ± 1728.3 ng.h/mL; p < 0.05) and 84.1% (from 208.5 ± 67.9 to 383.1 ± 92.5 ng/mL; p < 0.05), respectively. The CL/F was decreased from 1.7 L/h/kg to 0.5 L/h/kg. Thus, the effect of WZ on high dose of CsA was not significant, but pharmacokinetic parameters of CsA at low dose were significantly influenced by co-administration of WZ. The herb-drug interaction should be taken into consideration at this situation.

  13. Cyclosporine a augments P-glycoprotein expression in the regenerating rat liver.

    PubMed

    Daoudaki, Maria; Fouzas, Ioannis; Stapf, Verena; Ekmekcioglu, Cem; Imvrios, George; Andoniadis, Antonios; Demetriadou, Aphrodite; Thalhammer, Theresia

    2003-03-01

    In the liver, the multidrug resistance (MDR) protein P-glycoprotein (P-gp) is physiologically expressed at the bile canalicular membrane, where it participates in the biliary excretion of various lipophilic drugs and xenobiotics. Previous studies showed that the immunosuppressive agent cyclosporine A (CsA) modulates P-gp and exerts a hepatotrophic influence in the regenerating liver. Hepatocytes isolated from regenerating rat liver, after 2/3 partial hepatectomy (PH 2/3), were used as an in vivo experimental model of cells with high proliferating activity in order to investigate whether CsA influences cellular levels of P-gp in those cells. Male Wistar rats were treated with CsA (20 mg/kg body weight) for 4 d preoperatively and 1 d postoperatively, and regenerating hepatocytes were isolated by collagenase perfusion 12, 24 and 48 h after PH 2/3. Flow cytometry and Western blotting studies with the monoclonal antibodies C494 and C219 showed that after PH 2/3, cellular levels of P-gp were initially suppressed, 12 h after PH 2/3, by 23%, but were significantly elevated thereafter, 24 and 48 h after PH 2/3 by 28% and 73%, respectively. In CsA pretreated animals, P-gp levels were increased even in normal hepatocytes by 34%, and an additional augmentation was seen in hepatocytes from 24 and 48 h regenerating livers (60% and 56%, respectively). In summary, we demonstrate for the first time that CsA has an additive effect on the expression of P-glycoprotein during liver regeneration in the rat. Therefore, induction of P-gp might also be considered in patients receiving CsA after liver transplantation for hepatocellular carcinoma and chemotherapy as an adjuvant treatment for the prevention of tumor recurrence.

  14. Effects of dietary conjugated linoleic acids on cellular immune response of piglets after cyclosporin A injection.

    PubMed

    Liu, Y X; Zhu, K Y; Liu, Y L; Jiang, D F

    2016-10-01

    The present study investigated the effects of dietary conjugated linoleic acid (CLA) on the cellular immune response of piglets after cyclosporin A (CsA) treatment. The experimental study had a 2×2 factorial design, and the main factors consisted of diets (0% or 2% CLA) and immunosuppression treatments (CsA or saline injection). CsA injection significantly increased feed : gain (F : G) of piglets (P<0.05); however, dietary CLA significantly decreased F : G of piglets (P<0.05). Dietary CLA partly ameliorated the deterioration of the feed conversion rate caused by CsA treatment (P<0.01). CsA treatment significantly decreased the percentages of CD4+ and CD8+ T lymphocytes in the thymus (P<0.01). Dietary CLA increased the percentages of CD4+ CD8+ double-positive and CD8+ single-positive T lymphocytes in the thymus (P<0.05), and had the trend to inhibit the decrease of CD4+ T lymphocytes in the thymus after CsA injection (P=0.07). CsA treatment significantly depleted the peripheral blood CD3+, CD4+ and CD8+ T lymphocytes (P<0.01). Dietary CLA significantly increased the number of peripheral blood CD8+ T lymphocytes and interleukin-2 (IL-2) production (P<0.05), and inhibited the decreases of peripheral blood CD3+, CD4+ and CD8+ T lymphocytes counts (P<0.01) as well as IL-2 production (P<0.05) after CsA treatment. Dietary CLA partly rescued the decrease of lymphocyte proliferation after CsA injection (P<0.05). In summary, dietary CLA effectively ameliorated CsA-induced cellular immunosuppression in piglets.

  15. Cyclosporin A inhibits colon cancer cell growth independently of the calcineurin pathway

    PubMed Central

    Werneck, Miriam B.F.; Hottz, Eugênio; Bozza, Patrícia T.; Viola, João P.B.

    2012-01-01

    Chronic inflammation is a risk factor for the development of colon cancer, providing genotoxic insults, growth and pro-angiogenic factors that can promote tumorigenesis and tumor growth. Immunomodulatory agents can interfere with the inflammation that feeds cancer, but their impact on the transformed cell is poorly understood. The calcium/calcineurin signaling pathway, through activation of NFAT, is essential for effective immune responses, and its inhibitors cyclosporin A (CsA) and FK506 are used in the clinics to suppress immunity. Moreover, the kinases GSK3β and mTOR, modulated by PI-3K/Akt, can inhibit NFAT activity, suggesting a cross-talk between the calcium and growth factor signaling pathways. Both NFAT and mTOR activity have been associated with tumorigenesis. We therefore investigated the impact of calcineurin and PI-3K/mTOR inhibition in growth of human colon carcinoma cells. We show that despite the efficient inhibition of NFAT1 activity, FK506 promotes tumor growth, whereas CsA inhibits it due to a delay in cell cycle progression and induction of necroptosis. We found NFκB activation and mTORC1 activity not to be altered by CsA or FK506. Similarly, changes to mitochondrial homeostasis were equivalent upon treatment with these drugs. We further show that, in our model, NFAT1 activation is not modulated by PI3K/mTOR. We conclude that CsA slows cell cycle progression and induces necroptosis of human carcinoma cell lines in a TGFβ-, NFAT-, NFκB- and PI3K/mTOR-independent fashion. Nevertheless, our data suggest that CsA, in addition to its anti-inflammatory capacity, may target transformed colon and esophagus carcinoma cells without affecting non-transformed cells, promoting beneficial tumoristatic effects. PMID:22992618

  16. Effects of cyclosporine A on plasma cells in experimental gingivitis in dogs.

    PubMed

    Berglundh, T; Lindhe, J; Tarkowski, A

    1996-06-01

    The aim of the present investigation was to evaluate the effect of Cyclosporine A (CsA) on the inflammatory lesion formed in the gingival tissues during de novo plaque formation. 5 beagle dogs were used. On day 0, all teeth of the 5 dogs were scaled and polished. A 6-week period of plaque control including daily tooth cleaning with toothbrush and dentifrice was initiated. A clinical examination regarding plaque and gingivitis was performed, and the plaque control measures were abandoned on the right side of mandible. 3 weeks later, the clinical examination was repeated, samples of subgingival plaque harvested and biopsies obtained from the 3rd and 4th right mandibular premolar regions. The tooth cleaning measures on the left side of the mandible were terminated at this interval. During the following 3 weeks, the animals formed plaque in the lower left premolar regions, and received, 1 x daily, a subcutaneous injection of CsA. At the end of this 2nd plaque formation period (test), the clinical examination was repeated, subgingival plaque was sampled and biopsies from the 3rd and 4th left mandibular premolar regions harvested. The biopsies were prepared for histometric and morphometric analyses. The clinical and histological examinations demonstrated that plaque formation resulted in a gingival lesion (ICT) which, in the 2 periods, had similar size and apical extension. The ICT formed during the CsA administration period, however, harbored an increased number of plasma cells and a reduced macrophage density than the control lesion. It is suggested that CsA administration may result in a Th-2 (T-helper 2-cell) dependent activation of B-lymphocytes.

  17. Enhancement of oral bioavailability of cyclosporine A: comparison of various nanoscale drug-delivery systems

    PubMed Central

    Wang, Kai; Qi, Jianping; Weng, Tengfei; Tian, Zhiqiang; Lu, Yi; Hu, Kaili; Yin, Zongning; Wu, Wei

    2014-01-01

    A variety of nanoscale delivery systems have been shown to enhance the oral absorption of poorly water-soluble and poorly permeable drugs. However, the performance of these systems has seldom been evaluated simultaneously. The aim of this study was to compare the bioavailability enhancement effect of lipid-based nanocarriers with poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) to highlight the importance of the lipid composition, with cyclosporine A (CyA) as a model drug. CyA-loaded PLGA NPs, nanostructured lipid carriers (NLCs), and self-microemulsifying drug-delivery systems (SMEDDS) were prepared. The particle size of PLGA NPs (182.2±12.8 nm) was larger than that of NLCs (89.7±9.0 nm) and SMEDDS (26.9±1.9 nm). All vehicles are charged negatively. The entrapment efficiency of PLGA NPs and NLCs was 87.6%±1.6% and 80.3%±0.6%, respectively. In vitro release tests indicated that the cumulative release of CyA was lower than 4% from all vehicles, including Sandimmun Neoral®, according to the dialysis method. Both NLCs and SMEDDS showed high relative oral bioavailability, 111.8% and 73.6%, respectively, after oral gavage administration to beagle dogs, which was not statistically different from commercial Sandimmun Neoral®. However, PLGA NPs failed to achieve efficient absorption, with relative bioavailability of about 22.7%. It is concluded that lipid-based nanoscale drug-delivery systems are superior to polymeric NPs in enhancing oral bioavailability of poorly water-soluble and poorly permeable drugs. PMID:25378925

  18. [Cyclosporin A causes oxidative stress and mitochondrial dysfunction in renal tubular cells].

    PubMed

    Pérez de Hornedo, J; de Arriba, G; Calvino, M; Benito, S; Parra, T

    2007-01-01

    Reactive oxygen species (ROS) have been implicated in cyclosporin A (CsA) nephrotoxicity. As mitochondria are one of the main sources of ROS in cells, we evaluated the role of CsA in mitochondrial structure and function in LLC-PK1 cells. We incubated cells with CsA 1 microM for 24 hours and studies were performed with flow citometry and confocal microscopy. We studied mitochondrial NAD(P)H content, superoxide anion (O2.-) production (MitoSOX Red), oxidation of cardiolipin of inner mitochondrial membrane (NAO) and mitochondrial membrane potential (DIOC2(3)). Also we analyzed the intracellular ROS synthesis (H2DCF-DA) and reduced glutation (GSH) of cells. Our results showed that CsA decreased NAD(P)H and membrane potential, and increased O2.- in mitochondria. CsA also provoked oxidation of cardiolipin. Furthermore, CsA increased intracellular ROS production and decreased GSH content. These results suggest that CsA has crucial effects in mitochondria. CsA modified mitochondrial physiology through the decrease of antioxidant mitochondrial compounds as NAD(P)H and the dissipation of mitochondrial membrane potential and increase of oxidants as O2.-. Also, CsA alters lipidic structure of inner mitochondrial membrane through the oxidation of cardiolipin. These effects trigger a chain of events that favour intracellular synthesis of ROS and depletion of GSH that can compromise cellular viability. Nephrotoxic cellular effects of CsA can be explained, at least in part, through its influence on mitochondrial functionalism.

  19. Readily restoring freeze-dried probilosomes as potential nanocarriers for enhancing oral delivery of cyclosporine A.

    PubMed

    Guan, Peipei; Lu, Yi; Qi, Jianping; Wu, Wei

    2016-08-01

    Formulating vesicular nanocarriers into dried precursors so as to overcome the drawbacks associated with liquid formulations is challengeable due to low efficiency of restoration. In this study, bilosomes interiorly thickened with gelatin (G-BLs) was evaluated for the ability to withstand freeze-drying stress and enhanced oral bioavailability of a model drug, cyclosporine A (CyA). The restoration efficiency of freeze-dried pro-G-BLs is investigated by comparing the particle size distribution, entrapment efficiency and morphology of the bilosomes before and after freeze-drying. Particle size and polydispersity index (PI) of pro-G-BLs after restoration was similar to that before freeze-drying, whereas freeze-dried bilosomes without gelatin thickening (pro-BLs) show irreversible damage and aggregation along with significantly increased particle size and PI after restoration. Entrapment efficiency of pro-G-BLs remains as high as 83.7%, in sharp contrast with 66.7% for pro-BLs. Pharmacokinetics in beagle dogs show improved absorption of CyA in pro-G-BLs as compared to pro-BLs, G-BLs and microemulsion-based Sandimmun Neoral(®). The relative oral bioavailability of CyA-loaded pro-G-BLs, pro-BLs and G-BLs was 165.2%, 123.5% and 130.1%, respectively, with Neoral(®) as the reference. It is concluded that interior thickening with gelatin significantly enhanced the stability against freeze-drying stress, which as a result improves the restoring efficiency and oral bioavailability.

  20. Ubiquitous protective effects of cyclosporine A in preventing cardiac arrest-induced multiple organ failure.

    PubMed

    Cour, Martin; Abrial, Maryline; Jahandiez, Vincent; Loufouat, Joseph; Belaïdi, Elise; Gharib, Abdallah; Varennes, Annie; Monneret, Guillaume; Thibault, Hélène; Ovize, Michel; Argaud, Laurent

    2014-10-15

    Opening of the mitochondrial permeability transition pore (mPTP) appears to be a pivotal event in myocardial ischemia-reperfusion (I/R) injury. Resuscitated cardiac arrest (CA) leads to the post-CA syndrome that encompasses, not only myocardial dysfunction, but also brain injury, failure of other organs (kidney, liver, or lung), and systemic response to I/R. We aimed to determine whether cyclosporine A (CsA) might prevent multiple organ failure following CA through a ubiquitous mPTP inhibition in each distant vital organ. Anesthetized New Zealand White rabbits were subjected to 15 min of CA and 120 min of reperfusion. At the onset of resuscitation, the rabbits received CsA, its non-immunosuppressive derivative NIM811, or vehicle (controls). Survival, hemodynamics, brain damage, organ injuries, and systemic I/R response were analyzed. Fresh mitochondria were isolated from the brain, heart, kidney, liver, and lung to assess both oxidative phosphorylation and permeability transition. CsA analogs significantly improved short-term survival and prevented multiple organ failure, including brain damage and myocardial dysfunction (P < 0.05 vs. controls). Susceptibility of mPTP opening was significantly increased in heart, brain, kidney, and liver mitochondria isolated from controls, while mitochondrial respiration was impaired (P < 0.05 vs. sham). CsA analogs prevented these mitochondrial dysfunctions (P < 0.05 vs. controls). These results suggest that CsA and NIM811 can prevent the post-CA syndrome through a ubiquitous mitochondrial protective effect at the level of each major distant organ.

  1. Cyclosporin A markedly enhances superantigen-induced peripheral T cell deletion and inhibits anergy induction

    PubMed Central

    1992-01-01

    Cyclosporin A (CsA) is a well-known immunosuppressive agent that modulates immune tolerance in many ways. CsA can give rise to a state of long-term nonimmunosuppressed transplantation tolerance, but it can also aggravate autoimmune diseases, and provoke specific forms of autoimmunity. These effects, which are often paradoxical, remain largely unexplained. In this study, we investigated the effects of CsA on superantigen (superAg)-reactive peripheral T cells. The intravenous injection of either staphylococcal enterotoxin B (SEB), or Mls-1a cells into Mls-1b recipients, causes long-term in vitro nonresponsiveness (anergy) and partial elimination of the peripheral T cell receptor (TCR) V beta 8+/CD4+ and -V beta 6+/CD4+ T cell subsets, respectively. We report that CsA markedly enhances the peripheral elimination of SEB- and Mls-1a-reactive T cells such that up to 90% of the targeted CD4+/V beta subpopulations are deleted. The degree of deletion depends on the dose and the schedule of CsA administration, and the number of superAg injections. In situations where the extent of deletion is only moderate, we find that the remaining superAg-reactive T cells fail to develop anergy, unlike the T cells of control superAg-immunized mice. Higher doses of CsA are required to enhance T cell deletion (greater than or equal to 25 mg/kg/d, i.p.) than to impair anergy induction (greater than or equal to 6.25 mg/kg/d, i.p.). In view of these results, it appears that the degree of tolerance in CsA/superAg-treated mice depends on the balance between these opposing effects, i.e., enhancement of peripheral elimination versus the abrogation of anergy. The possibility of enhancing or preventing immune tolerance with a drug may have important clinical implications. PMID:1613464

  2. Synergistic Inhibition of Endothelial Cell Proliferation, Tube Formation, and Sprouting by Cyclosporin A and Itraconazole

    PubMed Central

    Nacev, Benjamin A.; Liu, Jun O.

    2011-01-01

    Pathological angiogenesis contributes to a number of diseases including cancer and macular degeneration. Although angiogenesis inhibitors are available in the clinic, their efficacy against most cancers is modest due in part to the existence of alternative and compensatory signaling pathways. Given that angiogenesis is dependent on multiple growth factors and a broad signaling network in vivo, we sought to explore the potential of multidrug cocktails for angiogenesis inhibition. We have screened 741 clinical drug combinations for the synergistic inhibition of endothelial cell proliferation. We focused specifically on existing clinical drugs since the re-purposing of clinical drugs allows for a more rapid and cost effective transition to clinical studies when compared to new drug entities. Our screen identified cyclosporin A (CsA), an immunosuppressant, and itraconazole, an antifungal drug, as a synergistic pair of inhibitors of endothelial cell proliferation. In combination, the IC50 dose of each drug is reduced by 3 to 9 fold. We also tested the ability of the combination to inhibit endothelial cell tube formation and sprouting, which are dependent on two essential processes in angiogenesis, endothelial cell migration and differentiation. We found that CsA and itraconazole synergistically inhibit tube network size and sprout formation. Lastly, we tested the combination on human foreskin fibroblast viability as well as Jurkat T cell and HeLa cell proliferation, and found that endothelial cells are selectively targeted. Thus, it is possible to combine existing clinical drugs to synergistically inhibit in vitro models of angiogenesis. This strategy may be useful in pursuing the next generation of antiangiogenesis therapy. PMID:21969860

  3. Effect of cyclosporin A intervention on the immunological mechanisms of coronary heart disease and restenosis

    PubMed Central

    Wang, Xuan; Hu, Yue-Cheng; Zhang, Ru-Yan; Jin, Dong-Xia; Jiang, Yuan; Zhang, He-Nan; Cong, Hong-Liang

    2016-01-01

    The present study aimed to investigate the effect of cyclosporin A (CSA) intervention on the immunological mechanisms underlying coronary heart disease (CHD) and restenosis (RS) in rabbits. A total of 48 rabbits were randomly divided into normal control (N), N + CSA, CHD model, CHD + CSA, RS model and RS + CSA groups. Rabbits in the respective groups received different treatments prior to sacrifice at the end of week 12. Iliac arteries were harvested from the rabbits for morphological analysis and to determine the mRNA and protein expression levels of cluster of differentiation (CD) 40/CD40 ligand (CD40L), CD134/CD134 ligand (CD134L) and inflammatory factors, including matrix metalloproteinase (MMP)-1, MMP-9, vascular cell adhesion protein (VCAM)-1, interleukin (IL)-6 and tumor necrosis factor (TNF)-α, by reverse transcription-quantitative polymerase chain reaction and immunohistochemical staining. As compared with the N group, the mRNA expression levels of MMP-9, VCAM-1 and TNF-α were significantly increased in the CHD and RS groups (P<0.05), but were significantly decreased in the groups with CSA intervention, as compared with those without CSA intervention (P<0.05). Conversely, there were no significant differences in the expression levels of MMP-1 and IL-6 among the six groups, although a decreasing trend of IL-6 expression was observed following intervention with CSA. Furthermore, there were significant differences in the mRNA and protein expression levels of CD40/CD40L and CD134/CD134L among the N, CHD and RS groups (P<0.05), and between the groups with and without CSA intervention. The present study demonstrated that CSA intervention exerted beneficial effects on CHD and RS, and further studies are required to investigate the mechanisms underlying the effects of CSA on CHD. PMID:27882144

  4. The role of inflammation and apoptosis in cyclosporine A – induced gingival overgrowth

    PubMed Central

    Mitic, Kristina; Popovska, Mirjana; Pandilova, Maja; Jovanovic, Rubens; Spasovski, Goce; Nikolov, Vladimir

    2013-01-01

    Cyclosporin A(CsA) - induced gingival overgrowth(GO) is a current problem of tissue-specific mechanism which is still incompletely explained. The apoptotic process has been of particular interest like a new concept in the etiology of this unwanted effect. The aim of our study was to detect the level of apoptosis, expression bcl-2 and p53, associated with the different dosis of CsA. in gingival stroma. A cohort of 84 kidney transplant recipients was divided into four subgroups based on average daily dose of therapeutically applied CsA (Ne-oral®), (100 mg, 125 mg, 150 mg and 175 mg). The control group consisted of 21 patients, clinically diagnosed with periodontitis, who were not subjected to any medicamentous treatment causing gingival overgrowth. The following indexes were analyzed: plaque index (PI), index of gingival inflammation (GI) according to Loe-Silnes, and gingival overgrowth index (GOI) according to MacGaw et al. The tissue samples were subjected to a semiquantitative analysis to detect apoptotical cells and imunohistochemically stained to detect the expression of the bcl-2 and p53 proteins. The difference in percentage of apoptotic cells between the group taking 175mg and other subgroups, as well as the control group was statistically significant (p<0.05). There was a significant difference in percentage of expression bcl-2 between the 175 mg group compared to the other three subgroups and the control (p=0.001). However, a statistically significant positive correlation between the medicament dose, p53, apoptosis, and bcl-2 was registered (p<0.05). Inflammation plays the most important role in the induction of apoptosis and proliferation in gingival tissues. PMID:23448605

  5. Cyclosporin A reduces expression of adhesion molecules in the kidney of rats with chronic serum sickness

    PubMed Central

    Rincón, J; Parra, G; Quiroz, Y; Benatuil, L; Rodríguez-Iturbe, B

    2000-01-01

    Treatment with cyclosporin A (CsA) improves proteinuria and reduces renal cellular infiltration in chronic serum sickness (CSS). We examined if these effects were associated with a reduced renal expression of CD54 and its ligands, interferon-gamma (IFN-γ), tumour necrosis factor-alpha (TNF-α) and MHC class II molecules. We studied two groups of rats in which CSS was induced by daily injections of ovalbumin (OVA): a group treated with CsA (OVA.CsA group, n = 11) and a group that received no treatment (OVA.CSS group, n = 11). An additional group of five rats (control group) received only phosphate buffer. Immunostaining techniques were used to follow CSS and to study the expression of CD54, CD18, CD11b/c, IFN-γ, TNF-α and MHC class molecules. Proteinuria (mg/24 h) was reduced from 248·2 ± 73·1 (OVA.CCS group) to 14·5 ± 13·1 with CsA treatment (P < 0·0001). The renal expression of CD54 and its ligands (CD18 and CD11b/c) was reduced by 50% to 75%. Correspondingly, there was a 60% to 85% reduction in the number of infiltrating leucocytes. The number of cells expressing TNF-α, IFN-γ and MHC II molecules was also reduced. CsA reduces expression of CD54 and its ligands. This effect is associated with a reduction of cellular infiltration, IFN-γ, TNF-α-producing cells and with MHC II expression in the kidney. These findings suggest that expression of adhesion molecules plays a critical role in CSS and underline the importance of cellular immunity in this experimental model. PMID:10931158

  6. Evidence for a functional receptor for cyclosporin A on the surface of lymphocytes.

    PubMed Central

    Cacalano, N A; Chen, B X; Cleveland, W L; Erlanger, B F

    1992-01-01

    Cyclosporin A (CsA) is an immunosuppressive agent that inhibits the synthesis of lymphokines by T lymphocytes at the level of transcription. A cytoplasmic protein, cyclophilin, is the most thoroughly studied CsA-binding protein, but its ubiquitous presence in cells of all types raises questions about its role in immunosuppression. In an attempt to ascertain the presence of a cell surface receptor, we synthesized two polyvalent macromolecular CsA derivatives, CsA-BBa-ovalbumin and CsA-BBa-aminodextran (CBD), from the product of the photochemical reaction of CsA and 4-benzoylbenzoic acid (CsA-BBa). (i) They inhibited the peptidylprolyl cis-trans isomerase activity of cyclophilin and the synthesis of interleukin 2 by phorbol ester-activated EL-4 cells. (ii) CBD also inhibited interleukin 2 secretion by Con A-activated T-cell-enriched mouse splenocytes. 4-Benzoylbenzoic acid (BBa)-aminodextran and aminodextran were inactive. (iii) Direct binding and competition studies with [3H]CsA indicated that CBD does not enter EL-4 cells (i.e., it acted at the surface). (iv) CBD caused agglutination of EL-4 cells, murine B and T lymphocytes, human thymocytes, and two T-cell hybridomas. Agglutination was inhibited by a monoclonal antibody to CsA and by CsA and CsA-BBa, but not by BBa. No agglutination was seen with BBa-aminodextran or aminodextran. HeLa cells, Vero (monkey kidney) cells, a mouse plasmacytoma, COS cells, and a poorly differentiated B-cell lymphoma were not agglutinated. (v) EL-4 cells failed to be agglutinated after treatment with trypsin or chymotrypsin. Specific agglutination was again possible after incubation for 5 h at 37 degrees C in the absence of enzyme. (vi) CBD covalently linked to crosslinked agarose beads inhibited interleukin 2 production by phorbol ester-stimulated EL-4 cells. No activity was seen if cell-to-bead contact was prevented by a 0.02-microns microporous filter that did not interfere with the passage of CBD. Our findings support the presence

  7. Mechanisms of hepatic transport of cyclosporin A: an explanation for its cholestatic action?

    PubMed Central

    Fricker, G.; Fahr, A.

    1997-01-01

    The hepatic transport of the immunosuppressive Cyclosporin A (CyA) was studied using liposomal phospholipid membranes, freshly isolated rat hepatocytes and bile canalicular plasma membrane vesicles from rat liver. The Na(+)-dependent, saturable uptake of the bile acid 3H-taurocholate into isolated rat liver cells was apparently competitively inhibited by CyA. However, the uptake of CyA into the cells was neither saturable, nor temperature-dependent nor Na(+)-dependent, nor could it be inhibited by bile salts or CyA-derivatives, indicating passive diffusion. In steady state depolarization fluorescence studies, CyA caused a concentration-dependent decrease of anisotropy, indicating a membrane fluidizing effect. Ion flux experiments demonstrated that CyA dramatically increases the permeability of Na+ and Ca2+ across phospholipid membranes in a dose- and time-dependent manner, suggesting a iontophoretic activity that might have a direct impact on cellular ion homeostasis and regulation of bile acid uptake. Photoaffinity labeling with a [3H]-labeled photolabile CyA-derivative resulted in the predominant incorporation of radioactivity into a membrane polypeptide with an apparent molecular weight of 160,000 and a minor labeling of polypeptides with molecular weights of 85,000-90,000. In contrast, use of a photolabile bile acid resulted in the labeling of a membrane polypeptide with an apparent molecular weight of 110,000, representing the bile canalicular bile acid carrier. The photoaffinity labeling as well as CyA transport by canalicular membrane vesicles were inhibited by CyA and the p-glycoprotein substrates daunomycin and PSC-833, but not by taurocholate, indicating that CyA is excreted by p-glycoprotein. CyA uptake by bile canalicular membrane vesicles was ATP-dependent and could not be inhibited by taurocholate. CyA caused a decrease in the maximum amount of bile salt accumulated by the vesicles with time. However, initial rates of [3H]-taurocholate uptake within

  8. Lipid nanoparticles for cyclosporine A administration: development, characterization, and in vitro evaluation of their immunosuppression activity

    PubMed Central

    Guada, Melissa; Sebastián, Victor; Irusta, Silvia; Feijoó, Esperanza; Dios-Viéitez, María del Carmen; Blanco-Prieto, María José

    2015-01-01

    Cyclosporine A (CsA) is an immunosuppressant commonly used in transplantation for prevention of organ rejection as well as in the treatment of several autoimmune disorders. Although commercial formulations are available, they have some stability, bioavailability, and toxicity related problems. Some of these issues are associated with the drug or excipients and others with the dosage forms. With the aim of overcoming these drawbacks, lipid nanoparticles (LN) have been proposed as an alternative, since excipients are biocompatible and also a large amount of surfactants and organic solvents can be avoided. CsA was successfully incorporated into LN using the method of hot homogenization followed by ultrasonication. Three different formulations were optimized for CsA oral administration, using different surfactants: Tween® 80, phosphatidylcholine, taurocholate and Pluronic® F127 (either alone or mixtures). Freshly prepared Precirol nanoparticles showed mean sizes with a narrow size distribution ranging from 121 to 202 nm, and after freeze-drying were between 163 and 270 nm, depending on the stabilizer used. Surface charge was negative in all LN developed. High CsA entrapment efficiency of approximately 100% was achieved. Transmission electron microscopy was used to study the morphology of the optimized LN. Also, the crystallinity of the nanoparticles was studied by X-ray powder diffraction and differential scanning calorimetry. The presence of the drug in LN surfaces was confirmed by X-ray photoelectron spectroscopy. The CsA LN developed preserved their physicochemical properties for 3 months when stored at 4°C. Moreover, when the stabilizer system was composed of two surfactants, the LN formulations were also stable at room temperature. Finally, the new CsA formulations showed in vitro dose-dependent immuno-suppressive effects caused by the inhibition of IL-2 levels secreted from stimulated Jurkat cells. The findings obtained in this paper suggest that new lipid

  9. Cyclosporine A and palmitic acid treatment synergistically induce cytotoxicity in HepG2 cells

    SciTech Connect

    Luo, Yi Rana, Payal; Will, Yvonne

    2012-06-01

    Immunosuppressant cyclosporine A (CsA) treatment can cause severe side effects. Patients taking immunosuppressant after organ transplantation often display hyperlipidemia and obesity. Elevated levels of free fatty acids have been linked to the etiology of metabolic syndromes, nonalcoholic fatty liver and steatohepatitis. The contribution of free fatty acids to CsA-induced toxicity is not known. In this study we explored the effect of palmitic acid on CsA-induced toxicity in HepG2 cells. CsA by itself at therapeutic exposure levels did not induce detectible cytotoxicity in HepG2 cells. Co-treatment of palmitic acid and CsA resulted in a dose dependent increase in cytotoxicity, suggesting that fatty acid could sensitize cells to CsA-induced cytotoxicity at the therapeutic doses of CsA. A synergized induction of caspase-3/7 activity was also observed, indicating that apoptosis may contribute to the cytotoxicity. We demonstrated that CsA reduced cellular oxygen consumption which was further exacerbated by palmitic acid, implicating that impaired mitochondrial respiration might be an underlying mechanism for the enhanced toxicity. Inhibition of c-Jun N-terminal kinase (JNK) attenuated palmitic acid and CsA induced toxicity, suggesting that JNK activation plays an important role in mediating the enhanced palmitic acid/CsA-induced toxicity. Our data suggest that elevated FFA levels, especially saturated FFA such as palmitic acid, may be predisposing factors for CsA toxicity, and patients with underlying diseases that would elevate free fatty acids may be susceptible to CsA-induced toxicity. Furthermore, hyperlipidemia/obesity resulting from immunosuppressive therapy may aggravate CsA-induced toxicity and worsen the outcome in transplant patients. -- Highlights: ► Palmitic acid and cyclosporine (CsA) synergistically increased cytotoxicity. ► The impairment of mitochondrial functions may contribute to the enhanced toxicity. ► Inhibition of JNK activity attenuated

  10. Total lymphoid irradiation in rat heart albgrafts: dose, fractionation, and combination with cyclosporin-A. [X-ray

    SciTech Connect

    Rynasiewicz, J.J.; Sutherland, D.E.R.; Kawahara, K.; Kim, T.; Najarian, J.S.

    1981-03-01

    The survival or organ allografts is prolonged in mice and rats treated with fractionated, high-dose total lymphoid irradiation (TLI). We have studied the effect of TLI, alone or in combination with donor bone marrow or pharmacologic immunosuppression (cyclosporin-A: CY-A), on the survival of heterotopic rat heart allografts. Specifically, we evaluated the generalized immunosuppressive effect of TLI as a function of accumulated dose and fractionation schedule. In addition, TLI and CY-A were used individually in schedules that by themselves gave only moderate graft prolongation and then subsequently in sequential combination.

  11. Development and Validation of a HPLC Method for the Determination of Cyclosporine A in New Bioadhesive Nanoparticles for Oral Administration

    PubMed Central

    Pecchio, M.; Salman, H.; Irache, J. M.; Renedo, M. J.; Dios-Viéitez, M. C.

    2014-01-01

    A simple and reliable high performance liquid chromatography method was developed and validated for the rapid determination of cyclosporine A in new pharmaceutical dosage forms based on the use of poly (methylvinylether-co-maleic anhydride) nanoparticles. The chromatographic separation was achieved using Ultrabase C18 column (250×4.6 mm, 5 μm), which was kept at 75°. The gradient mobile phase consisted of acetonitrile and water with a flow rate of 1 ml/min. The effluent was monitored at 205 nm using diode array detector. The method exhibited linearity over the assayed concentration range (22-250 μg/ml) and demonstrated good intraday and interday precision and accuracy (relative standard deviations were less than 6.5% and the deviation from theoretical values is below 5.5%). The detection limit was 1.36 μg/ml. This method was also applied for quantitative analysis of cyclosporine A released from poly (methylvinylether-co-maleic anhydride) nanoparticles. PMID:24843186

  12. Cyclosporine a 0.05% eye drops for the treatment of subepithelial infiltrates after epidemic keratoconjunctivitis

    PubMed Central

    2012-01-01

    Background To evaluate the treatment with topical 0.05% cyclosporine A (CsA) in patients with subepithelial corneal infiltrates (SEI). Methods We reviewed 16 patients (22 eyes) before and after the treatment with 0.05% CsA eye drops. All patients had been treated previously with topical corticosteroids without any improvement and also they had to stop the medication secondary to intraocular pressure elevation. The objective data recorded included best-corrected visual acuity (BCVA), evaluation of corneal subepithelial infiltrate scores (CSIS), intraocular pressure (IOP) prior to treatment and the last follow-up visit. Results Six males (37.5%) and 10 females (62.5%), mean age of 35.2 ± 16.6 years, were included. The patients’ average topical CsA use duration was 5.1 ± 3.5 months (1 – 13 months). The average follow up time of the patients was 9.2 ± 4.7 months (4 – 22 months). One patient, although he didn’t have a 0 scale of SCIS, did not show up for follow up examinations after six months. The mean BCVA (logarithm of the minimum angle of resolution) before and after the treatment were 0.15 ± 0.15 and 0.07 ± 0.07 respectively, CSIS 1.68 ± 0.89 and 0.23 ± 0.53 respectively, IOP 18.50 ± 3.82 and 16.86 ± 2.76 mmHg respectively. There were statistically significant improvements in BCVA (p = 0.002), reduction of CSIS (p = 0.002) and reduction of IOP (p < 0.001) prior to treatment and the last follow-up visit. 18 eyes (81.9%) showed clinical improvement and 4 (18.1%) had decreased SEI which did not fully disappear during the treatment period. The eyes which reached CSIS score 0 (18 eyes) were treated with CsA for 1 – 13 months; while the eyes which had clinical improvement but had not CSIS score 0 (4 eyes) were decided to discontinue of CsA treatment in last follow-up visit. There were recurrences in 2 eyes 3 months after the treatment. Patients reported reduction in the severity of symptoms after the

  13. Effects of rifampin, cyclosporine A, and probenecid on the pharmacokinetic profile of canagliflozin, a sodium glucose co-transporter 2 inhibitor, in healthy participants

    PubMed Central

    Devineni, Damayanthi; Vaccaro, Nicole; Murphy, Joe; Curtin, Christopher; Mamidi, Rao N.V.S.; Weiner, Sveta; Wang, Shean-Sheng; Ariyawansa, Jay; Stieltjes, Hans; Wajs, Ewa; Di Prospero, Nicholas A.; Rothenberg, Paul

    2015-01-01

    Objective: Canagliflozin, a sodium-glucose co-transporter 2 inhibitor, approved for the treatment of type-2 diabetes mellitus (T2DM), is metabolized by uridine diphosphate-glucuronosyltransferases (UGT) 1A9 and UGT2B4, and is a substrate of P-glycoprotein (P-gp). Canagliflozin exposures may be affected by coadministration of drugs that induce (e.g., rifampin for UGT) or inhibit (e.g. probenecid for UGT; cyclosporine A for P-gp) these pathways. The primary objective of these three independent studies (single-center, open-label, fixed-sequence) was to evaluate the effects of rifampin (study 1), probenecid (study 2), and cyclosporine A (study 3) on the pharmacokinetics of canagliflozin in healthy participants. Methods: Participants received; in study 1: canagliflozin 300 mg (days 1 and 10), rifampin 600 mg (days 4 – 12); study 2: canagliflozin 300 mg (days 1 – 17), probenecid 500 mg twice daily (days 15 – 17); and study 3: canagliflozin 300 mg (days 1 – 8), cyclosporine A 400 mg (day 8). Pharmacokinetics were assessed at pre-specified intervals on days 1 and 10 (study 1); on days 14 and 17 (study 2), and on days 2 – 8 (study 3). Results: Rifampin decreased the maximum plasma canagliflozin concentration (Cmax) by 28% and its area under the curve (AUC) by 51%. Probenecid increased the Cmax by 13% and the AUC by 21%. Cyclosporine A increased the AUC by 23% but did not affect the Cmax. Conclusion: Coadministration of canagliflozin with rifampin, probenecid, and cyclosporine A was well-tolerated. No clinically meaningful interactions were observed for probenecid or cyclosporine A, while rifampin coadministration modestly reduced canagliflozin plasma concentrations and could necessitate an appropriate monitoring of glycemic control. PMID:25407255

  14. Comparison of Efficacy of Two Different Topical 0.05% Cyclosporine A Formulations in the Treatment of Adenoviral Keratoconjunctivitis-Related Subepithelial Infiltrates

    PubMed Central

    Bayraktutar, Betül N.; Uçakhan, Ömur Ö.

    2016-01-01

    Subepithelial infiltrates secondary to adenoviral keratoconjunctivitis may persist for years and cause blurred vision, halos, glare, and photophobia. These infiltrates arise from immune reaction against the virus, and few studies have reported topical cyclosporine A to be effective in the treatment of subepithelial infiltrates. Herein, we describe a patient with adenoviral keratoconjunctivitis-related subepithelial infiltrates who did not respond to treatment with a new topical cyclosporine A emulsion prepared with castor oil (Depores 0.05%; Deva İlaç, Kocaeli, Turkey), while the FDA-approved nanoemulsion formulation provided improvement in symptoms and reduced the inflammatory reaction (Restasis 0.05%; Allergan, Irvine, Calif., USA). PMID:27065851

  15. Reversible fibroadenomatous mammary hyperplasia in male and female New Zealand white rabbits associated with cyclosporine A administration.

    PubMed

    Krimer, P M; Harvey, S B; Blas-Machado, U; Lauderdale, J D; Moore, P A

    2009-11-01

    All male and female New Zealand white rabbits in a limbal cell graft study developed marked generalized mammary gland hypertrophy. Postprocedural medications included ophthalmic 0.1% dexamethasone, ophthalmic 0.5% cyclosporine, and subcutaneous cyclosporine A. Cytologic examination revealed epithelial clusters with minimal malignant criteria. On histologic evaluation, there was diffuse glandular hyperplasia with mild cellular atypia and ductal ectasia separated by abundant hypercellular fibrous stroma, consistent with fibroadenomatous mammary gland hyperplasia. The hyperplasia resolved within 2 weeks of cessation of cyclosporine, and at necropsy identifiable mammary masses were not found. Very little has been reported about the use of cyclosporine in laboratory rabbits and its association with development of mammary gland hyperplasia. This is the first report in which administration of cyclosporine to male and female rabbits at a dose as low as 5 mg/kg/day induced benign fibroadenomatous mammary gland hyperplasia. This change regressed after cessation of the drug.

  16. Preparation and evaluation of poly(ethylene glycol)-poly(lactide) micelles as nanocarriers for oral delivery of cyclosporine a.

    PubMed

    Zhang, Yanhui; Li, Xinru; Zhou, Yanxia; Wang, Xiaoning; Fan, Yating; Huang, Yanqing; Liu, Yan

    2010-03-27

    A series of monomethoxy poly(ethylene glycol)-poly(lactide) (mPEG-PLA) diblock copolymers were designed according to polymer-drug compatibility and synthesized, and mPEG-PLA micelle was fabricated and used as a nanocarrier for solubilization and oral delivery of Cyclosporine A (CyA). CyA was efficiently encapsulated into the micelles with nanoscaled diameter ranged from 60 to 96 nm with a narrow size distribution. The favorable stabilities of CyA-loaded polymeric micelles were observed in simulated gastric and intestinal fluids. The in vitro drug release investigation demonstrated that drug release was retarded by polymeric micelles. The enhanced intestinal absorption of CyA-loaded polymeric micelles, which was comparable to the commercial formulation of CyA (Sandimmun Neoral®), was found. These suggested that polymeric micelles might be an effective nanocarrier for solubilization of poorly soluble CyA and further improving oral absorption of the drug.

  17. Cyclosporine A treatment for Ullrich congenital muscular dystrophy: a cellular study of mitochondrial dysfunction and its rescue.

    PubMed

    Hicks, D; Lampe, A K; Laval, S H; Allamand, V; Jimenez-Mallebrera, C; Walter, M C; Muntoni, F; Quijano-Roy, S; Richard, P; Straub, V; Lochmüller, H; Bushby, K M D

    2009-01-01

    Mutations in COL6A1, COL6A2 and COL6A3, the genes which encode the extra-cellular matrix component collagen VI, lead to Bethlem myopathy and Ullrich congenital muscular dystrophy (UCMD). Although the Col6a1(-/-) null mouse has an extremely mild neuromuscular phenotype, a mitochondrial defect has been demonstrated, linked to dysregulation of the mitochondrial permeability transition pore (PTP) opening. This finding has been replicated in UCMD muscle cells in culture, providing justification for a clinical trial using cyclosporine A, an inhibitor of PTP opening. We investigated whether PTP dysregulation could be detected in UCMD fibroblasts (the predominant source of muscle collagen VI), in myoblast cells from patients with other diseases and its response to rescue agents other than collagen VI. Although we confirm the presence of PTP dysregulation in muscle-derived cultures from two UCMD patients, fibroblasts from the same patients and the majority of fibroblasts from other well-characterized UCMD patients behave normally. PTP dysregulation is found in limb girdle muscular dystrophy (LGMD) type 2B myoblasts but not in myoblasts from patients with Bethlem myopathy, merosin-deficient congenital muscular dystrophy, LGMD2A, Duchenne muscular dystrophy and Leigh syndrome. In addition to rescue by cyclosporine A and collagen VI, this cellular phenotype was also rescued by other extra-cellular matrix constituents (laminin and collagen I). As the muscle derived cultures demonstrating PTP dysregulation shared poor growth in culture and lack of desmin labelling, we believe that PTP dysregulation may be a particular characteristic of the state of these cells in culture and is not specific to the collagen VI defect, and can in any case be rescued by a range of extra-cellular matrix components. Further work is needed on the relationship of PTP dysregulation with UCMD pathology.

  18. [Closure of Ca2+-dependent pores by cyclosporin A: the role of magnesium ions, adenine nucleotides, and conformation status of the ADP/ATP antiporter].

    PubMed

    Andreev, A Iu; Mikhaĭlova, L M; Starkov, A A

    1994-10-01

    Effects of ADP and Mg2+ on the ability of cyclosporin A to "reseal" mitochondria permeabilized by Ca2+ and P(i) have been studied. Cyclosporin A was completely ineffective, when ADP and Mg2+ were not included into the incubation medium. Both ADP and Mg2+ used at high concentrations potentiated the effect of cyclosporin A and prevented it reversal by carboxyatractylate. Data on the influence of different concentrations of ADP and Mg2+ on the resealing efficiency of cyclosporin A suggest that the true effector modulating the state of the Ca(2+)-dependent pore is the ADP-Mg2+ complex, but not ADP or Mg2+ used separately. The ability of non-hydrolyzable analogs of adenine nucleotides, ADP-S and ATP-S, to potentiate the resealing action of cyclosporin on mitochondria permeabilized by loading of different Ca2+ concentrations to that of ADP was compared. ATP-S was ineffective when the pore was induced by high concentrations of Ca2+. The results obtained are discussed in terms of hypothesis on the direct involvement of the ADP/ATP antiporter in regulation of the inner mitochondrial membrane Ca(2+)-dependent pore state.

  19. Molecular dynamics simulations of cyclosporin A: The crystal structure and dynamic modelling of a structure in apolar solution based on NMR data

    NASA Astrophysics Data System (ADS)

    Lautz, J.; Kessler, H.; Kaptein, R.; van Gunsteren, W. F.

    1987-10-01

    The conformation of the immunosuppressive drug cyclosporin A (CPA), both in apolar solution and in crystalline state, has been studied by computer simulation techniques. Three molecular dynamics (MD) simulations have been performed: one modelling the crystal structure and two modelling the structure in apolar solution, using a restrained MD approach in which data from nuclear magnetic resonance (NMR) and infrared (IR) spectroscopy are taken into account. The simulation of the crystalline state (MDC) concerns a system of 4 unit cells containing 16 cyclosporin A molecules and 22 water molecules, which is simulated using crystalline periodic boundary conditions. The simulations modelling the apolar solvent conformation (MDS) concern one isolated cyclosporin A molecule. In these simulations an extra term in the interatomic potential function is used, which forces the molecule to satisfy a set of 57 atom-atom distance constraints originating from nuclear Overhauser effects (NOEs) obtained from NMR spectroscopy and one distance constraint deduced from IR spectroscopy. From a comparison of the results of the crystal simulation to those of the X-ray experiment in terms of structure, atomic fluctuations, hydrogen bond pattern, etc., it is concluded that the force field that is used yields an adequate representation of crystalline cyclosporin A. Secondly, it is shown that the dynamic modelling technique that is used to obtain a structure in a polar solution from NMR distance information works well. Starting from initial conformations which have a root mean square difference of 0.14 nm both distance restrained MD simulations converge to the same final solution structure. A comparison of the crystal structure of cyclosporin A and the one in apolar solution shows that there are significant differences. The overall difference in atomic positions is 0.09 nm for the Cx atoms and 0.17 nm for all atoms. In apolar solution, the molecule is slightly more bent and the side chains of 1

  20. Relevance of p-glycoprotein for the enteral absorption of cyclosporin A: in vitro-in vivo correlation.

    PubMed Central

    Fricker, G.; Drewe, J.; Huwyler, J.; Gutmann, H.; Beglinger, C.

    1996-01-01

    1. The interaction of cyclosporin A (CyA) with p-glycoprotein during intestinal uptake was investigated by a combination of in vitro experiments with human Caco-2 cells and an intubation study in healthy volunteers. 2. CyA uptake into the cells was not saturable and exhibited only a low temperature sensitivity, suggesting passive diffusion. When the permeation of CyA across Caco-2 monolayers from the apical to the basolateral side was determined, overall transport had an apparently saturable component up to a concentration of 1 microM. At higher concentrations permeation increased over-proportionally. Calculation of the kinetic parameters of apical to basolateral permeation suggested a diffusional process with a KD of 0.5 microliter min-1 per filter, which was overlayed by an active system in basolateral to apical direction with a KM of 3.8 microM and a Jmax of 6.5 picomol min-1 per filter. 3. CyA permeation was significantly higher when the drug was given from the basolateral side as compared to the permeation from the apical side. Apical to basolateral transport of CyA was increased in the presence of vinblastine, daunomycin and a non-immunosuppressive CyA-derivative. All compounds inhibit p-glycoprotein-mediated transport processes. Basolateral to apical permeation of CyA showed a dose-dependent decrease in the presence of vinblastine. Permeation of daunomycin across Caco-2 cell monolayers was also higher from the basolateral to the apical side than vice versa. Basolateral to apical permeation was decreased in the presence of SDZ PSC 833 and cyclosporin A. 4. Western blot analysis of Caco-2 cells with the monoclonal antibody C219 confirmed the presence of p-glycoprotein in the used cell system. 5. When the absorption of CyA in the gastrointestinal (GI)-tract of healthy volunteers was determined, a remarkable decrease of the plasma AUC could be observed dependent on the location of absorption in the rank order stomach > jejunum/ileum > colon. The decrease in

  1. Cyclosporine A-induced increase in glomerular cyclic GMP in rats and the involvement of the endothelinB receptor

    PubMed Central

    Tack, Ivan; Marin-Castano, Encarna; Bascands, Jean-Loup; Pecher, Christiane; Ader, Jean-Louis; Girolami, Jean-Pierre

    1997-01-01

    A transient two fold increase in the cyclic GMP content was observed in rat freshly isolated glomeruli 6 to 9 h after a single subcutaneous injection of 20 mg kg−1 cyclosporine A (CsA) in conscious animals. In vitro stimulation with endothelin 3 (ET-3) of isolated glomeruli obtained from CsA-untreated rats resulted in a dose-dependent increase in cyclic GMP content. The increase observed with 10 nM ET-3 was similar to that observed in glomeruli isolated 9 h after in vivo CsA administration. The rise in glomerular cyclic GMP content after in vivo CsA injection was prevented by in vivo treatment with L-NAME (10 mg kg−1) or by in vitro calcium deprivation of the incubation medium. The stimulating effects of CsA on glomerular cyclic GMP content were inhibited by in vivo administration of the ETB receptor antagonist BQ-788 (2 mg kg−1) but not by the ETA receptor antagonist BQ-123 (2 mg kg−1). The maximum increase in glomerular cyclic GMP content induced in vitro by acetylcholine (100 μM) and by ET-3 (100 nM) was slightly lower (approximately by 20–25%, P<0.05) in glomeruli from CsA-treated rats than in glomeruli from untreated rats. In contrast, the maximum increase achieved with 1 μM sodium nitroprusside was similar in both groups. A single subcutaneous injection of CsA did not significantly alter the glomerular mRNA expression of constitutive endothelial NO synthase (eNOS), as evaluated by RT–PCR, whereas the mRNA expression of the inducible NO synthase (iNOS), which follows pretreatment with lipopolysaccharide, was prevented. These results indicate that in vivo administration of a single dose of cyclosporine A transiently increases the cyclic GMP content of freshly isolated glomeruli, and that activation of ETB receptors and stimulation of the NO pathway are involved in this process. Furthermore, a single administration of CsA does not impair eNOS mRNA expression and only slightly reduces NO-dependent glomerular cyclic GMP

  2. Hyaluronic acid coated poly-epsilon-caprolactone nanospheres deliver high concentrations of cyclosporine A into the cornea.

    PubMed

    Yenice, Irem; Mocan, Mehmet C; Palaska, Erhan; Bochot, Amélie; Bilensoy, Erem; Vural, Imran; Irkeç, Murat; Hincal, A Atilla

    2008-09-01

    The objective of this study was to determine cyclosporine A (Cy A) levels in ocular tissues and fluids after topical administration of poly-epsilon-caprolactone (PCL)/benzalkonium chloride (BKC) nanospheres and hyaluronic acid (HA) coated PCL/BKC nanospheres onto healthy rabbit corneas. Nanospheres were prepared by nanoprecipitation and purified by gradient-rate centrifugation. Cy A (0.1%) in either castor oil solution (group 1), PCL/BKC nanosphere formulation (group 2) or HA coated PCL/BKC nanosphere formulation (group 3) was instilled onto rabbit corneas. Tear samples were adsorbed onto Schirmer tear strips. Cy A concentrations of fluid (blood, aqueous humor, tear) and specimen extracts (cornea, conjunctiva, iris/ciliary body) were determined by high performance liquid chromatography-mass spectrometry (LC-MS). The mean corneal Cy A concentration obtained at 0.5, 1, 2, 4, 8 and 24h following instillation of the formulations ranged between 0.12 and 1.2 ng/mg tissue for group 1, 5.9-15.5 ng/mg tissue for group 2 and 11.4-23.0 ng/mg for group 3 (one-way analysis of variance (ANOVA) and pairwise tests (SNK (Student-Newman-Keuls) and Tukey); p<0.05). Conjunctival Cy A levels of group 2 and 3 were not significantly different at any of the time points tested. However, there was a significant difference between Cy A concentration of castor oil formulation and that of PCL/BKC nanosphere formulation at 1 and 8h (p<0.05). The mean iris/ciliary body concentrations obtained with the three formulations were not significantly different at any time point with the exception of group 2 levels being higher than those of groups 1 and 3 at 1h (p<0.05). The lowest ocular tear Cy A concentrations (16-114 ng/ml) were found following the instillation of HA coated PCL/BKC nanoparticles (group 3) during the time period tested. Cy A loaded PCL/BKC and HA coated PCL/BKC nanospheres are able to achieve high levels of Cy A in the cornea that is 10-15-fold higher than that is achieved with Cy A

  3. Solution structures of cyclosporin a and its complex with dysprosium(III) in SDS micelles: NMR and molecular dynamics studies.

    PubMed

    Bernardi, Francesca; D'Amelio, Nicola; Gaggelli, Elena; Molteni, Elena; Valensin, Gianni

    2008-01-24

    Cyclosporin A (CsA) is a cyclic naturally occurring peptide used to prevent graft rejection in organ transplantations. Its immunosuppressive activity is due to the formation of a complex with cyclophilin A (Cyp), in which the cis 9MeLeu-10MeLeu amide bond of CsA assumes a trans conformation. The mechanism of the conformational inversion has not been delineated, but it has been postulated that metal ions binding induces a conformational change that enables CsA to bind Cyp. In this work, we solved the structures of CsA in sodium dodecyl sulfate (SDS) micelles (which enhance its solubility and mimic the hydrophobic environment clinically used for drug delivery) and its complex with Dy(III) ion, whose coordination chemistry is frequently used to reproduce the effect of Ca(II). The paramagnetic properties of Dy(III) allowed us to build up a structure using proton relaxation enhancements, which remains stable in a MD simulation in the micelle environment.

  4. The effects of cyclosporin-A on functional outcome and axonal regrowth following spinal cord injury in adult rats.

    PubMed

    Roozbehi, Amrollah; Joghataie, Mohammad Taghi; Mehdizadeh, Mehdi; Mirzaei, Ali; Delaviz, Hamdollah

    2012-01-01

    It has been shown that the immunophilin ligands have the special advantage in spinal cord repair. In this study, the effects of cyclosporine A (CsA) on functional recovery and histological outcome were evaluated following spinal cord injury in rats. After spinal cord hemisection in thirty six adult female Sprague-Dawley rats (200- 250 g), treatment groups received CsA (2.5 mg/kg i.p.) at 15min and 24h after lesion (CsA 15min group and CsA 24h group) daily, for 8 weeks. Control and sham groups received normal saline and in sham operated animals the spinal cord was exposed in the same manner as treatment groups, but was not hemisected. Hindlimb motor function was assessed in 1, 3, 5 and 7 weeks after lesion, using locomotive rating scale developed by Basso, Bresnahan and Beattie (BBB). Motor neurons were counted within the lamina IX of ventral horn and lesion size was measured in 5 mm of spinal lumbar segment with the epicenter of the lesion site. The mean number of motor neurons and the mean BBB scale in 3, 5 and 7 weeks in CsA 15min groups significantly increased compared to the control group. Although, the lesion size reduced in rats with CsA treatment compared to the control group, no significant difference was observed. Thus, it can be concluded that CsA can improve locomotor function and histological outcome in the partial spinal cord injury.

  5. Fatal Epstein-Barr Virus Reactivation in an Acquired Aplastic Anemia Patient Treated with Rabbit Antithymocyte Globulin and Cyclosporine A

    PubMed Central

    Takahashi, Tohru; Maruyama, Yumiko; Saitoh, Mayuko; Itoh, Hideto; Yoshimoto, Mitsuru; Tsujisaki, Masayuki

    2015-01-01

    Epstein-Barr virus (EBV) associated lymphoproliferative disorder (LPD) after immunosuppressive therapy for aplastic anemia (AA) is extremely rare in a nontransplant setting and has not been well described. This report describes a severe AA patient in whom fatal EBV-LPD developed after being treated with rabbit antithymocyte globulins (ATG) and cyclosporine A (CsA). An 81-year-old man was diagnosed as having severe AA. He was started on CsA followed by administration of ATG for five consecutive days. One month after the start of ATG, persistent fever which was not responsive to antibiotics or antifungal agents developed and atypical lymphocytes emerged in peripheral blood. Repeated blood cultures were negative. An extremely high level of EBV virus in his peripheral blood plasma was detected by means of a quantitative real-time PCR assay. Even after the cessation of CsA, the fever persisted and the peripheral atypical lymphocytes proliferated rapidly. The patient suffered from respiratory failure, liver dysfunction, and metabolic acidosis. Rituximab was administered without success and he died. PMID:26425376

  6. EAU in the guinea pig: inhibition of cell-mediated immunity and Ia antigen expression by cyclosporin A.

    PubMed Central

    Liversidge, J; Thomson, A W; Sewell, H F; Forrester, J V

    1987-01-01

    Guinea pigs were immunized subcutaneously with highly purified bovine retinal S antigen (SAg) in complete Freund's adjuvant and treated from day 0 with cyclosporin A (CsA; 25 mg/kg by mouth) or drug vehicle. Skin tests carried out at 7 and 13 days showed maximal reactions to SAg at 24 h; at 13 days, however, strong, early, 'Arthus'-like responses to SAg were also recorded. CsA profoundly reduced DTH skin reactions to SAg and PPD, and prevented vitreal inflammation assessed at 17 days and retinal damage. Lymphocytes from the draining lymph nodes but not spleens of immunized guinea pigs showed a proliferative response to SAg which was suppressed by CsA administration. Responses to PHA, Con A or LPS were not so affected. Immunohistochemical staining (alkaline phosphatase-anti-alkaline phosphatase; APAAP) of the eye with newly available monoclonal antibodies to guinea pig T lymphocytes revealed a predominantly T cytotoxic/suppressor cell (Tc/s) infiltrate of the choroid and retina. CsA administration did not affect choroidal infiltration of Tc/s cells but markedly inhibited Ia antigen expression. Images Fig. 3 Fig. 4 Fig. 5 PMID:3478162

  7. Establishing a Clinically Relevant Large Animal Model Platform for TBI Therapy Development: Using Cyclosporin A as a Case Study

    PubMed Central

    Margulies, Susan S.; Kilbaugh, Todd; Sullivan, Sarah; Smith, Colin; Propert, Kathleen; Byro, Melissa; Saliga, Kristen; Costine, Beth A.; Duhaime, Ann-Christine

    2015-01-01

    We have developed the first immature large animal translational treatment trial of a pharmacologic intervention for traumatic brain injury (TBI) in children. The preclinical trial design includes multiple doses of the intervention in two different injury types (focal and diffuse) to bracket the range seen in clinical injury and uses two post-TBI delays to drug administration. Cyclosporin A (CsA) was used as a case study in our first implementation of the platform because of its success in multiple preclinical adult rodent TBI models and its current use in children for other indications. Tier 1 of the therapy development platform assessed the short-term treatment efficacy after 24 h of agent administration. Positive responses to treatment were compared with injured controls using an objective effect threshold established prior to the study. Effective CsA doses were identified to study in Tier 2. In the Tier 2 paradigm, agent is administered in a porcine intensive care unit utilizing neurological monitoring and clinically relevant management strategies, and intervention efficacy is defined as improvement in longer term behavioral endpoints above untreated injured animals. In summary, this innovative large animal preclinical study design can be applied to future evaluations of other agents that promote recovery or repair after TBI. PMID:25904045

  8. Buparvaquone but not cyclosporin A prevents Theileria annulata-infected bovine lymphoblastoid cells from stimulating uninfected lymphocytes.

    PubMed

    Rintelen, M; Schein, E; Ahmed, J S

    1990-06-01

    The influence of Buparvaquone on the morphology, proliferation, and stimulation with T and B cell mitogens of Theileria annulata-infected cells was studied. In addition, the stimulatory capacity of the infected cells before and after treatment with Buparvaquone or cyclosporin A (CsA) was also examined and compared to that of ConA-stimulated bovine peripheral blood cells (PBL). After incubation of the cells for 4 days with Buparvaquone only few schizonts were detectable in the cells. Prolongation of the incubation time to 8, 12, or 14 days eliminated completely the parasites. Despite the elimination of the parasites, the cells were still unable to undergo a proliferative response to Con A or PWM. However, the drug did not interfere with the response of normal PBL to these mitogens. Furthermore, Buparvaquone but not CsA inhibits the generation of mixed lymphocyte reaction (MLR). None of the drugs could prevent ConA-blasts from stimulating autologous PBL. These results suggest that the antigen expressed by the infected cells and recognised by the responder PBL was induced by the schizonts.

  9. [Comparison of the determination of cyclosporin-A in blood samples collected on filter paper and by the ordinary technique].

    PubMed

    Azevedo, L S; Manrique, R; Sabbaga, E

    1995-01-01

    Monitoring cyclosporin-A (CsA) blood levels is of utmost importance for the rational use of this drug. Although many centers perform transplants, in Brazil there are few laboratories able to measure CsA blood levels. Therefore making blood samples reach the laboratory emerged as a problem. Collection of blood on filter paper has been a technique used for a long time in special cases. PURPOSE--To confirm the usefulness of measuring CsA blood levels in blood samples collected on filter paper and in the usual way. METHOD--We studied twenty renal cadaver kidney recipients who were receiving CsA, azathioprine and prednisone. Ninety five blood samples were collected and divided into two aliquots. One of them was sent routinely to one laboratory to perform whole blood CsA measurements. From the other aliquot, 20 microliters were pipetted on filter paper. When dried they were mailed to the other laboratory, where, after elution, CsA was measured. In both cases radioimmunoassay with polyclonal antibody was used. RESULTS--Linear correlation between both measurements revealed r = 0.81 with no statistical difference. CONCLUSION--The technique showed to be useful in clinical practice. In countries with continental size, as Brazil, it may be very helpful.

  10. Intracellular Ca2+ elevation and cyclosporin A synergistically induce TGF-beta 1-mediated apoptosis in lymphocytes.

    PubMed

    Andjelíc, S; Khanna, A; Suthanthiran, M; Nikolić-Zugić, J

    1997-03-15

    Apoptosis plays an essential role in the development and homeostasis of the immune system. During lymphocyte development, potentially autoreactive cells are eliminated via the activation of a tightly regulated cell death program(s). Similar processes operate in mature lymphocytes, to control the magnitude of the normal immune response by eliminating activated lymphocytes. However, differences in susceptibility to signal-induced apoptosis between immature and mature lymphocytes are numerous. One well-characterized example occurs in response to Ca2+ elevation: peripheral T lymphocytes are resistant, while immature thymocytes are highly susceptible, to Ca2+-mediated cell death (CMCD). In this study, we show that the immunosuppressant cyclosporin A (CsA) primes splenic lymphocytes to undergo CMCD upon ionomycin stimulation. This CsA-induced CMCD affected both T and B lymphocytes. CsA-plug Ca2+-mediated apoptosis was dissected into a two-step process: first, CsA and Ca2+ synergized to induce TGF-beta 1 secretion by B cells; and then TGF-beta 1 and Ca2+ synergistically triggered T and B lymphocyte apoptosis. Together, our results suggest that lymphocyte apoptosis may play a role in CsA-induced immunosuppression via a TGF-beta-dependent mechanism.

  11. Cyclosporin A Impairs the Secretion and Activity of ADAMTS13 (A Disintegrin and Metalloprotease with Thrombospondin Type 1 Repeat)*

    PubMed Central

    Hershko, Klilah; Simhadri, Vijaya L.; Blaisdell, Adam; Hunt, Ryan C.; Newell, Jordan; Tseng, Sandra C.; Hershko, Alon Y.; Choi, Jae Won; Sauna, Zuben E.; Wu, Andrew; Bram, Richard J.; Komar, Anton A.; Kimchi-Sarfaty, Chava

    2012-01-01

    The protease ADAMTS13 (a disintegrin and metalloprotease with thrombospondin type 1 repeat) cleaves multimers of von Willebrand factor, thus regulating platelet aggregation. ADAMTS13 deficiency leads to the fatal disorder thrombotic thrombocytopenic purpura (TTP). It has been observed that cyclosporin A (CsA) treatment, particularly in transplant patients, may sometimes be linked to the development of TTP. Until now, the reason for such a link was unclear. Here we provide evidence demonstrating that cyclophilin B (CypB) activity plays an important role in the secretion of active ADAMTS13. We found that CsA, an inhibitor of CypB, reduces the secretion of ADAMTS13 and leads to conformational changes in the protein resulting in diminished ADAMTS13 proteolytic activity. A direct, functional interaction between CypB (which possesses peptidyl-prolyl cis-trans isomerase (PPIase) and chaperone functions) and ADAMTS13 is demonstrated using immunoprecipitation and siRNA knockdown of CypB. Finally, CypB knock-out mice were found to have reduced ADAMTS13 levels. Taken together, our findings indicate that cyclophilin-mediated activity is an important factor affecting secretion and activity of ADAMTS13. The large number of proline residues in ADAMTS13 is consistent with the important role of cis-trans isomerization in the proper folding of this protein. These results altogether provide a novel mechanistic explanation for CsA-induced TTP in transplant patients. PMID:23144461

  12. The efficacy of cyclosporine A in cats with presumed atopic dermatitis: a double blind, randomised prednisolone-controlled study.

    PubMed

    Wisselink, Marinus A; Willemse, Ton

    2009-04-01

    The objective of this study was to compare the efficacy of cyclosporine A (CsA) and prednisolone in feline atopic dermatitis (AD) in a randomised, controlled double blind study. Twenty-nine cats with feline AD were randomly allocated to two groups. Eleven cats were treated orally with prednisolone (1mg/kg SID) and 18 were treated with CsA (5mg/kg/day) for 4 weeks. At day 0 (D0) and D28, skin lesions were graded by means of the canine atopic dermatitis extent and severity index (CADESI). Skin biopsies and intradermal allergy tests were performed at D0 and blood samples for haematology and serum biochemistry were collected at D0 and D28. During the trial the cat owners were asked to evaluate the intensity of the pruritus once weekly on a linear analog scale and to record side effects. Based on the CADESI there was no significant difference between the two groups in the amount of remission (P=0.0562) or in the number of cats that improved by >25% (P=0.0571). The effect of CsA and prednisolone on pruritus as evaluated by the owners was not significantly different (P=0.41) between the two groups. No serious side effects were observed. The conclusion was that CsA is an effective alternative to prednisolone therapy in cats with presumed atopic dermatitis.

  13. Enhanced and sustained topical ocular delivery of cyclosporine A in thermosensitive hyaluronic acid-based in situ forming microgels

    PubMed Central

    Wu, Yijun; Yao, Jing; Zhou, Jianping; Dahmani, Fatima Zohra

    2013-01-01

    For nearly a decade, thermoresponsive ophthalmic in situ gels have been recognized as an interesting and promising ocular topical delivery vehicle for lipophilic drugs. In this study, a series of thermosensitive copolymers, hyaluronic acid-g-poly(N-isopropylacrylamide) (HA-g-PNIPAAm), was synthesized, by coupling carboxylic end-capped PNIPAAm to aminated hyaluronic acid through amide bond linkages, and was used as a potential carrier for the topical ocular administration of cyclosporine A (CyA). The lower critical solution temperature of HA-g-PNIPAAm59 in aqueous solutions was measured as 32.7°C, which was not significantly affected by the polymer concentration. Moreover, HA-g-PNIPAAm59 microgels showed a high drug loading efficiency (73.92%) and a controlled release profile that are necessary for biomedical application. Transmission electron microscopy (TEM) and atomic force microscopy (AFM) observations showed that HA-g-PNIPAAm microgels were spherical in shape with homogeneous size. Based on the result of the eye irritation test, the HA-g-PNIPAAm microgels formulation was shown to be safe and nonirritant for rabbit eyes. In addition, HA-g-PNIPAAm microgels achieved significantly higher CyA concentration levels in rabbit corneas (1455.8 ng/g of tissue) than both castor oil formulation and commercial CyA eye drops. Therefore, these newly described thermoresponsive HA-g-PNIPAAm microgels demonstrated attractive properties to serve as pharmaceutical delivery vehicles for a variety of ophthalmic applications. PMID:24092975

  14. Cyclosporin A inhibits caspase-independent death of NGF-deprived sympathetic neurons: a potential role for mitochondrial permeability transition.

    PubMed

    Chang, Louis K; Johnson, Eugene M

    2002-05-27

    Opening of the permeability transition pore (PTP) has been implicated as an important mitochondrial event that occurs during apoptosis. We examined the role of the PTP in the well-characterized cell death of rat sympathetic neurons deprived of nerve growth factor (NGF) in vitro. Removal of NGF causes these neurons to undergo either a classic apoptotic cell death or, when treated with a broad-spectrum caspase inhibitor such as boc-aspartyl(OMe)-fluoromethylketone (BAF), a delayed, nonapoptotic cell death. The PTP inhibitor, cyclosporin A (CsA), blocked commitment-to-die in the presence of BAF, as defined by the ability of NGF readdition to rescue cells, but had little effect on commitment-to-die in the absence of BAF. CsA did not have trophic effects on BAF-saved cells, but did block the decrease in mitochondrial membrane potential. These data suggest that PTP opening is a critical event in caspase-independent, nonapoptotic (but not caspase-dependent, apoptotic) death of NGF-deprived rat sympathetic neurons.

  15. A Cell-Impermeable Cyclosporine A Derivative Reduces Pathology in a Mouse Model of Allergic Lung Inflammation

    PubMed Central

    Balsley, Molly A.; Malesevic, Miroslav; Stemmy, Erik J.; Gigley, Jason; Jurjus, Rosalyn A.; Herzog, Dallen; Bukrinsky, Michael I.; Fischer, Gunter; Constant, Stephanie L.

    2013-01-01

    Although the main regulators of leukocyte trafficking are chemokines, another family of chemotactic agents is cyclophilins. Intracellular cyclophilins function as peptidyl-protyl cis-trans isomerases and are targets of the immunosuppressive drug, cyclosporine A (CsA). Cyclophilins can also be secreted in response to stress factors, with elevated levels of extracellular cyclophilins detected in several inflammatory diseases. Extracellular cyclophilins are known to have potent chemotactic properties, suggesting they might contribute to inflammatory responses by recruiting leukocytes into tissues. The objective of the current study was to determine the impact of blocking cyclophilin activity using a cell-impermeable derivative of CsA, MM218, to specifically target extracellular pools of cyclophilins. We show that treatment with this compound in a mouse model of allergic lung inflammation: 1) demonstrates up to 80% reduction in inflammation, 2) directly inhibits the recruitment of antigen-specific CD4+ T cells, and 3) works equally well when delivered at 100-fold lower doses to the airways. Our findings suggest that cell-impermeable analogs of CsA can effectively reduce inflammatory responses by targeting leukocyte recruitment mediated by extracellular cyclophilins. Specifically blocking the extracellular function(s) of cyclophilins may provide a novel approach for inhibiting the recruitment of one of the principal immune regulators of allergic lung inflammation, antigen-specific CD4+ T cells, into inflamed airways and lungs. PMID:21057089

  16. Using a modified shepards method for optimization of a nanoparticulate cyclosporine a formulation prepared by a static mixer technique.

    PubMed

    Douroumis, Dionysios; Scheler, Stefan; Fahr, Alfred

    2008-02-01

    An innovative methodology has been used for the formulation development of Cyclosporine A (CyA) nanoparticles. In the present study the static mixer technique, which is a novel method for producing nanoparticles, was employed. The formulation optimum was calculated by the modified Shepard's method (MSM), an advanced data analysis technique not adopted so far in pharmaceutical applications. Controlled precipitation was achieved injecting the organic CyA solution rapidly into an aqueous protective solution by means of a static mixer. Furthermore the computer based MSM was implemented for data analysis, visualization, and application development. For the optimization studies, the gelatin/lipoid S75 amounts and the organic/aqueous phase were selected as independent variables while the obtained particle size as a dependent variable. The optimum predicted formulation was characterized by cryo-TEM microscopy, particle size measurements, stability, and in vitro release. The produced nanoparticles contain drug in amorphous state and decreased amounts of stabilizing agents. The dissolution rate of the lyophilized powder was significantly enhanced in the first 2 h. MSM was proved capable to interpret in detail and to predict with high accuracy the optimum formulation. The mixer technique was proved capable to develop CyA nanoparticulate formulations.

  17. Treatment of premalignancy: prevention of lymphoma in radiation leukemia virus-inoculated mice by cyclosporin A and immunotoxin.

    PubMed Central

    Yefenof, E; Abboud, G; Epszteyn, S; Vitetta, E S

    1992-01-01

    Radiation leukemia virus (RadLV)-induced preleukemic (PL) latency is characterized by the appearance of virus-infected PL cells in the thymus. The survival of these PL cells is dependent upon autostimulation with interleukin 4 (IL-4). We have intervened prophylactically in RadLV-induced preleukemia by using cyclosporin-A (CSA), which inhibits IL-4 production, and an immunotoxin (ITx) that kills PL cells. CSA efficiently inhibited IL-4 secretion from RadLV-induced PL and leukemic cells, and its administration to PL mice caused a significant delay in their death. An ITx consisting of anti-RadLV glycoprotein-70 (gp70) antibody coupled to ricin A chain efficiently inhibited protein synthesis in virus-infected cells in vitro and, when injected into PL mice, also delayed their death. Combined treatment with CSA and ITx prevented 75% of the treated PL mice from developing lymphoma. These results show that the development of malignancy from a premalignant state can be averted by a combination of therapeutic modalities that decrease the size and growth rate of the premalignant cell population. PMID:1731346

  18. Nanoliposome containing cyclosporine A reduced neuroinflammation responses and improved neurological activities in cerebral ischemia/reperfusion in rat.

    PubMed

    Partoazar, Alireza; Nasoohi, Sanaz; Rezayat, Sayed M; Gilani, Kambiz; Mehr, Shahram E; Amani, Amir; Rahimi, Nastaran; Dehpour, Ahmad R

    2017-04-01

    Cyclosporine A (CsA) is known as a neuroprotective agent against cerebral ischemia/reperfusion (I/R) in animal models. However, the significant therapeutic effects of CsA have been observed in high systemic doses or manipulating the blood-brain barrier, resulting in systemic side effects and toxicity. As the liposome nanocarriers have been developed for efficient delivery of peptide and proteins, liposomal CsA (Lipo-CsA) could improve cerebral (I/R) injuries. In this study, the liposomal CsA formulation (CsA at dose of 2.5 mg/kg) was prepared to assess the brain injury outcomes in 90 min middle cerebral artery occlusion (MCAO) stroke model followed by 48 h reperfusion in treating rats. Five minutes after induction of cerebral ischemia in rats, intravenous (iv) administration of Lipo-CsA significantly (P < 0.001) recovered the infarct size, the brain edema, and the neurological activities compared to corresponding control groups following 48 h I/R. In addition, after 48 h cerebral I/R, Lipo-CsA potentially (P < 0.001) inhibited the inflammation responses including MPO activity and tumor necrosis factor-alpha level in comparison to other groups. In conclusion, the results indicate that the low dose of CsA in liposomal formulation is more effective compared to higher dose of free form of CsA in treatment of ischemic brain in rats.

  19. Outcome of anti-thymocyte immunoglobulin plus cyclosporine A for severe aplastic anaemia with chronic hepatitis B virus infection.

    PubMed

    Chen, Miao; Zhuang, Junling; Zhou, Daobin; Xu, Ying; Zhao, Yongqiang; Wang, Shujie; Zhang, Wei; Duan, Minghui; Zhu, Tienan; Li, Jian; Cai, Huacong; Cao, Xinxin; Han, Bing

    2017-04-01

    The influence of chronic hepatitis B virus (HBV) infection on the efficacy of intensive immunosuppressive treatment (IST) of severe aplastic anaemia (SAA) patients remains unclear. Previous reports on this topic have been mostly case reports or have had a relatively short follow-up. Eight SAA patients carrying chronic HBV infection and 24 matched patients without HBV at a ratio of 1:3 were included in this retrospective analysis. The patients were treated with anti-thymocyte globulin (ATG) and cyclosporine A. Entecavir was or was not administered throughout the IST course to patients with positive or negative HBV-DNA results, respectively. No evident HBV reactivation developed. The overall response was 87.5% by 12 months, and the recurrence rate was 12.5%. There were no significant differences in overall response, overall survival and event-free survival between groups. Entecavir can effectively prevent reactivation of HBV in SAA patients with positive HBV-DNA who received intensive IST. Regular surveillance may be sufficient for HBV-DNA negative patients who should receive antiviral drugs immediately when their HBV-DNA status changes from negative to positive. The prognosis of SAA patients with chronic HBV infection after intensive IST treatment is not worse than those without HBV infection.

  20. Generation of oxygen free radicals during the metabolism of cyclosporine A: a cause-effect relationship with metabolism inhibition.

    PubMed

    Serino, F; Grevel, J; Napoli, K L; Kahan, B D; Strobel, H W

    1993-05-26

    A better understanding of the mechanism of lipid peroxidation during the metabolism of cyclosporine A (CsA) might help explain the toxicities of this immunosuppressive drug on various organs. Our in vitro work used microsomes prepared from livers of phenobarbital-induced male rats. The incubations (total volume 1ml) also contained a NADPH regenerating system and substrate (i.e., CsA, carbon tetrachloride, or aminopyrine) dissolved in ethanol. Lipid peroxidation was inferred from the presence of malondialdehyde (MDA) which was detected by the thiobarbituric acid assay. The formation of CsA hydroxylated metabolites (AM9 and AM1) was monitored by liquid chromatography. The activity of the microsomal incubation was confirmed by measurements of MDA and formaldehyde production caused by increasing concentrations of CsA, carbon tetrachloride, and aminopyrine. The occurrence of hydroxylated metabolites was not coupled to the production of MDA. Aminopyrine could inhibit MDA production by CsA, but CsA could not reduce the formation of formaldehyde by aminopyrine. Erythromycin, a competitor for the binding site of CsA on cytochrome P450, reduced MDA production by CsA, and CsA inhibited formaldehyde production by erythromycin. Interaction studies with SKF 525A, ketoconazole, superoxide dismutase, catalase, alpha-tocopherol, and reduced glutathione confirmed the role of cytochrome P450 and the presence of activated oxygen species as a source of microsomal peroxidation which in return may explain the inhibitory effect of CsA on cytochrome P450 itself.

  1. Establishing a Clinically Relevant Large Animal Model Platform for TBI Therapy Development: Using Cyclosporin A as a Case Study.

    PubMed

    Margulies, Susan S; Kilbaugh, Todd; Sullivan, Sarah; Smith, Colin; Propert, Kathleen; Byro, Melissa; Saliga, Kristen; Costine, Beth A; Duhaime, Ann-Christine

    2015-05-01

    We have developed the first immature large animal translational treatment trial of a pharmacologic intervention for traumatic brain injury (TBI) in children. The preclinical trial design includes multiple doses of the intervention in two different injury types (focal and diffuse) to bracket the range seen in clinical injury and uses two post-TBI delays to drug administration. Cyclosporin A (CsA) was used as a case study in our first implementation of the platform because of its success in multiple preclinical adult rodent TBI models and its current use in children for other indications. Tier 1 of the therapy development platform assessed the short-term treatment efficacy after 24 h of agent administration. Positive responses to treatment were compared with injured controls using an objective effect threshold established prior to the study. Effective CsA doses were identified to study in Tier 2. In the Tier 2 paradigm, agent is administered in a porcine intensive care unit utilizing neurological monitoring and clinically relevant management strategies, and intervention efficacy is defined as improvement in longer term behavioral endpoints above untreated injured animals. In summary, this innovative large animal preclinical study design can be applied to future evaluations of other agents that promote recovery or repair after TBI.

  2. Cyclosporin A-associated changes in red blood cell membrane composition, deformability, blood and plasma viscosity in rats.

    PubMed

    Ademoglu, Evin; Tamer, Sule; Albeniz, Isil; Gokkusu, Cahide; Tanrikulu, Sevda

    2004-01-01

    Most of the studies concerning the effects of cyclosporin A (Cs A) on red blood cell (RBC) rheology were carried out in human transplant recipients who may still have residual insufficiency and concomitant administration of other immunosuppressive and antihypertensive drugs. The aim of this study is to evaluate the effects of Cs A on red cell rheology and membrane composition in nontransplant healthy rats. Female Wistar albino rats were divided into two groups of 10 animals each. Rats received 10 mg/kg Cs A, i.p. or saline for 4 weeks. Cs A administration significantly increased the RBC deformability, and plasma and blood viscosity (p < 0.001, p < 0.01 and p < 0.01, respectively). Cs A administration to the rats increased RBC membrane cholesterol (CHO) levels and the CHO/phospholipid (PL) ratio significantly (p < 0.01 and p < 0.05, respectively) but did not change RBC membrane proteins and membrane PL levels. These results suggest that Cs A changes the rheological functions of RBC and lipid content of RBC membrane in healthy rats and thereby it may play an important role in the regulation of microcirculation.

  3. Cyclosporin A does not protect the disruption of the inner mitochondrial membrane potential induced by potassium ionophores in intact K562 cells.

    PubMed

    Marques-Santos, Luis F; Coqueiro, Vivian M; Rumjanek, Vivian M

    2006-03-01

    Mitochondrial dysfunction has been widely associated with programmed cell death. Studies of intact cells are important for the understanding of the process of cell death and its relation to mitochondrial physiology. Using cytofluorometric approaches we studied the mitochondrial behavior in an erythroleukemic cell line. The effects of protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP), potassium exchanger (nigericin), potassium ionophore (valinomycin), Na+K+-ATPase inhibitor (ouabain) and mitochondrial permeability transition pore inhibitor (cyclosporin A) were evaluated. Cyclosporin A (CSA) was very effective in attenuating the disruption of inner mitochondrial membrane potential induced by CCCP. However, CSA failed to protect the loss of inner mitochondrial membrane potential induced by potassium intracellular flux manipulation. Our findings suggest that mitochondrial cyclophilin is not involved in the cell events mediated by deregulation of potassium flux, underlining the need for further studies in intact tumor cells for a better understanding of the involvement of mitochondria physiology in cell death events.

  4. Indomethacin and cyclosporin a inhibit in vitro ischemia-induced expression of ICAM-1 and chemokines in human brain endothelial cells.

    PubMed

    Zhang, W; Smith, C; Monette, R; Hutchison, J; Stanimirovic, D B

    2000-01-01

    Brain inflammation has been implicated in the development of brain edema and secondary brain damage in ischemia and trauma. Mechanisms involved in leukocyte infiltration across the blood-brain barrier are still unknown. In this study, we show that human cere-bromicrovascular endothelial cells (HCEC) subjected to a 4 h in vitro ischemia (hypoxia + glucose deprivation) followed by a 4-24 h recovery express elevated levels of ICAM-1, IL-8, and MCP-1 mRNAs (semi-quantitative RT-PCR) and secrete increased amounts of the immunoreactive chemokines IL-8 and MCP-1 (ELISA). The ischemia-induced expression of ICAM-1 in HCEC, and the expression/release of IL-8 and MCP-1 in HCEC were abolished by the non-steroid anti-inflammatory drug, indomethacin (100-300 microM). The immunosuppressant cyclosporin A (50 microM) partially reduced the ischemia-stimulated IL-8 and MCP-1 secretion by HCEC. Both indomethacin and cyclosporin A also inhibited the ischemia-induced neutrophil chemotaxis elicited by HCEC media. The study indicates that in vitro ischemia augments the expression of adhesion molecules and leukocyte chemoattractants at the site of the BBB. This ischemic pro-inflammatory activation of HCEC may constitute a key event in initiating post-ischemic inflammation, and it can be suppressed by the anti-inflammatory drugs, indomethacin and cyclosporin A.

  5. Enhancing effect of negative polypropylene electret on in vitro transdermal delivery of cyclosporine A solution and its synergistic effect with ethyl oleate

    NASA Astrophysics Data System (ADS)

    Cui, L. L.; Ma, L.; Liang, Y. Y.; Liu, H. Y.; Guo, X.; Jiang, J.

    2013-03-01

    In this study, the corona charged electrets at voltages of -500 V, -1000 V and -2000 V were made from polypropylene (PP) film. The cyclosporine A (CsA) and 10% ethyl oleate were chosen as the model drug and chemical enhancer, respectively. The charge storage stability of the electrets and the in vitro transdermal behaviour of the model drug in solution under different conditions were studied. The results indicate that the external electrostatic field of the negative PP electrets could penetrate through the rat skin and enhance the transdermal delivery of cyclosporine A. A synergistic effect on enhancing the transdermal delivery of cyclosporine A was observed by combining different surface potential negative PP electrets with 10% ethyl oleate, and the amount of transdermal delivery of CsA was greatly increased comparing with only application of electrets. Therefore, the combination application of electret and chemical enhancer could be a feasible strategy in enhancing transdermal delivery of small peptide drugs or some large molecular drugs.

  6. Kimura's disease: case report of an Italian young male and response to oral cyclosporine A in an 8 years follow-up.

    PubMed

    Beccastrini, Enrico; Emmi, Giacomo; Chiodi, Michela; Di Paolo, Camilla; Benedetta Silvestri, Elena; Massi, Daniela; Maggi, Enrico; Liotta, Francesco; Emmi, Lorenzo

    2013-03-01

    Kimura's disease is a benign chronic inflammatory disease, common in Asian males and rare in Western people. Clinically, Kimura's disease is characterized by subcutaneous nodular lesions, usually localised in head and neck, often associated with regional lymphadenopathy. Peripheral blood eosinophilia and elevated serum IgE are often observed. We report a case of a 40-year-old Italian patient presenting with nodular subcutaneous lesions and peripheral eosinophilia. Based on clinical, histopathological and laboratory findings, a diagnosis of Kimura's disease was made. The patient was treated with very low doses of cyclosporine A with no evidence of disease recurrence over the following 8 years. However, the discontinuation of cyclosporine A determined a relapse of the disease. The relevance of this case is due to the rarity of the disease in Italy, to its peculiar clinical presentation and, moreover, it is the first case in literature that has a good response to treatment with low doses of cyclosporine A, documented in an 8-year follow-up.

  7. Physicochemical characterization and aerosol dispersion performance of organic solution advanced spray-dried cyclosporine A multifunctional particles for dry powder inhalation aerosol delivery.

    PubMed

    Wu, Xiao; Zhang, Weifen; Hayes, Don; Mansour, Heidi M

    2013-01-01

    In this systematic and comprehensive study, inhalation powders of the polypeptide immunosuppressant drug - cyclosporine A - for lung delivery as dry powder inhalers (DPIs) were successfully designed, developed, and optimized. Several spray drying pump rates were rationally chosen. Comprehensive physicochemical characterization and imaging was carried out using scanning electron microscopy, hot-stage microscopy, differential scanning calorimetry, powder X-ray diffraction, Karl Fischer titration, laser size diffraction, and gravimetric vapor sorption. Aerosol dispersion performance was conducted using a next generation impactor with a Food and Drug Administration-approved DPI device. These DPIs displayed excellent aerosol dispersion performance with high values in emitted dose, respirable fraction, and fine particle fraction. In addition, novel multifunctional inhalation aerosol powder formulations of cyclosporine A with lung surfactant-mimic phospholipids were also successfully designed and developed by advanced organic solution cospray drying in closed mode. The lung surfactantmimic phospholipids were 1,2-dipalmitoyl-sn-glycero-3-phosphocholine and 1,2-dipalmitoyl-snglycero- 3-(phosphor-rac-1-glycerol). These cyclosporine A lung surfactant-mimic aerosol powder formulations were comprehensively characterized. Powder X-ray diffraction and differential scanning calorimetry confirmed that the phospholipid bilayer structure in the solid state was preserved following advanced organic solution spray drying in closed mode. These novel multifunctional inhalation powders were optimized for DPI delivery with excellent aerosol dispersion performance and high aerosol performance parameters.

  8. Effect of FK506 and cyclosporine A on the expression of BDNF, tyrosine kinase B and p75 neurotrophin receptors in astrocytes exposed to simulated ischemia in vitro.

    PubMed

    Gabryel, Bozena; Bernacki, Jacek

    2009-07-01

    We investigated whether the immunosuppressive drugs, FK506 and cyclosporine A, increase BDNF protein and/or mRNA expression in ischemic astrocytes and if an increase could be related to changes in the nuclear expression of p-CREB, p-Erk1/2 and p-Akt. The influence of these immunosuppressants on protein and mRNA levels of TrkB and p75(NTR) receptors was also examined. On day 21, cultures of rat astrocytes were subjected to ischemic conditions simulated in vitro (combined oxygen glucose deprivation, OGD) for 8h and exposed to FK506 (10-1000nM) and cyclosporine A (0.25-10microM). FK506 and cyclosporine A (at 1000nM and 0.25microM, respectively) stimulated the expression and release of BDNF in cultured rat cerebral cortical astrocytes exposed to OGD. The immunosuppressants at these doses simultaneously increased p-CREB and p-Erk1/2 expression in the nuclear fraction of astrocytes. The results RT-PCR and Western blot analysis provided further evidence of a modulating influence of the drugs on the expression of trkB and p75(NTR) genes and their protein products in ischemic astrocytes.

  9. Physicochemical characterization and aerosol dispersion performance of organic solution advanced spray-dried cyclosporine A multifunctional particles for dry powder inhalation aerosol delivery

    PubMed Central

    Wu, Xiao; Zhang, Weifen; Hayes, Don; Mansour, Heidi M

    2013-01-01

    In this systematic and comprehensive study, inhalation powders of the polypeptide immunosuppressant drug – cyclosporine A – for lung delivery as dry powder inhalers (DPIs) were successfully designed, developed, and optimized. Several spray drying pump rates were rationally chosen. Comprehensive physicochemical characterization and imaging was carried out using scanning electron microscopy, hot-stage microscopy, differential scanning calorimetry, powder X-ray diffraction, Karl Fischer titration, laser size diffraction, and gravimetric vapor sorption. Aerosol dispersion performance was conducted using a next generation impactor with a Food and Drug Administration-approved DPI device. These DPIs displayed excellent aerosol dispersion performance with high values in emitted dose, respirable fraction, and fine particle fraction. In addition, novel multifunctional inhalation aerosol powder formulations of cyclosporine A with lung surfactant-mimic phospholipids were also successfully designed and developed by advanced organic solution cospray drying in closed mode. The lung surfactantmimic phospholipids were 1,2-dipalmitoyl-sn-glycero-3-phosphocholine and 1,2-dipalmitoyl-snglycero- 3-(phosphor-rac-1-glycerol). These cyclosporine A lung surfactant-mimic aerosol powder formulations were comprehensively characterized. Powder X-ray diffraction and differential scanning calorimetry confirmed that the phospholipid bilayer structure in the solid state was preserved following advanced organic solution spray drying in closed mode. These novel multifunctional inhalation powders were optimized for DPI delivery with excellent aerosol dispersion performance and high aerosol performance parameters. PMID:23569375

  10. Concentrations of cyclosporin A and FK506 that inhibit IL-2 induction in human T cells do not affect TGF-beta1 biosynthesis, whereas higher doses of cyclosporin A trigger apoptosis and release of preformed TGF-beta1.

    PubMed

    Minguillón, Jordi; Morancho, Beatriz; Kim, Seong-Jin; López-Botet, Miguel; Aramburu, José

    2005-05-01

    Cyclosporin A (CsA) and FK506 suppress T cell activation by inhibiting calcineurin and the calcineurin-dependent transcription factors nuclear factor of activated T cells (NFATc), which are central regulators of T cell function. It was reported that CsA up-regulated the transcription of transforming growth factor-beta1 (TGF-beta1) in lymphocytes and other cells and activated its promoter in A549 lung carcinoma cells, but the mechanisms involved are poorly understood, and it is unclear whether calcineurin plays any role. We have studied the regulation of TGF-beta1 in normal human lymphocytes and cell lines. In Jurkat T cells, the TGF-beta1 promoter was activated by calcineurin and NFATc and inhibited by CsA and FK506. However, the promoter was insensitive to both drugs in A549 cells. In human T cells preactivated with phytohemagglutinin, biosynthesis of TGF-beta1, induced by the T cell receptor (TCR) or the TGF-beta receptor, was not substantially affected by CsA and FK506 concentrations (< or = 1 microM) that effectively inhibited interleukin-2 production. However, pretreatment of fresh lymphocytes with CsA or FK506 during primary TCR stimulation reduced their production of TGF-beta1 during secondary TCR activation. Finally, high concentrations of CsA (10 microM), in the range attained in vivo in experiments in rodents, caused apoptosis in human T cells and the release of preformed, bioactive TGF-beta1. These effects are unlikely to owe to calcineurin inhibition, as they were not observed with FK506. Our results indicate that CsA and FK506 are not general inducers of TGF-beta1 biosynthesis but can cause different effects on TGF-beta1 depending on the cell type and concentrations used.

  11. Concurrent short-term use of prednisolone with cyclosporine A accelerates pruritus reduction and improvement in clinical scoring in dogs with atopic dermatitis

    PubMed Central

    2013-01-01

    Background A randomized, unmasked, multicenter study was conducted to evaluate the rate of pruritus reduction and improvement in clinical scoring by cyclosporine A (5 mg/kg orally, once daily for 28 days) either alone (n = 25 dogs) or with concurrent prednisolone (1 mg/kg once daily for 7 days, followed by alternate dosing for 14 days; n = 23 dogs) for the treatment of atopic dermatitis in dogs. Dogs were included in the study after exclusion of other causes of pruritic dermatitis, and were assessed by dermatologists on days 0, 14 ± 1 and 28 ± 2. Assessments included: general physical examination, CADESI-03 lesion scoring, overall clinical response, evaluation of adverse events (AEs), body weight and clinical pathology (hematology, clinical chemistry and urinalysis). Owner assessments, including pruritus (visual analogue scale, VAS) and overall assessment of response were conducted every 3–4 days, either during visits to the clinic or at home. Owners reported AEs to the investigator throughout the study. Results By day 28 ± 2 both treatment groups resulted in a significant improvement of the atopic dermatitis. Both investigators and owners agreed that concurrent therapy resulted in a quicker improvement of the dogs ‘overall’ skin condition and of pruritus (significant reduction of pruritus by day 3–4, 72.8% improvement by day 14 ± 1), when compared to cyclosporine A alone (significant reduction of pruritus by day 7–8, 24.7% improvement by day 14 ± 1). CADESI-03 scores significantly improved in both groups by day 14 ± 1 onwards, and there were no significant differences in the scores between treatment groups at any time points. A total of 56 AEs (cyclosporine A alone = 34; concurrent therapy = 22) were reported in 33 dogs. No dogs died or stopped treatment due to an AE. The most commonly reported AEs in the cyclosporine A group were associated with the digestive tract, whilst systemic disorders were

  12. The P-glycoprotein inhibitor cyclosporin A differentially influences behavioural and neurochemical responses to the antidepressant escitalopram.

    PubMed

    O'Brien, Fionn E; O'Connor, Richard M; Clarke, Gerard; Donovan, Maria D; Dinan, Timothy G; Griffin, Brendan T; Cryan, John F

    2014-03-15

    Recent studies have raised the possibility that P-glycoprotein (P-gp) inhibition may represent a putative augmentation strategy for treatment with certain antidepressants. Indeed, we have previously shown that administration of the P-gp inhibitor verapamil increased the brain distribution and behavioural effects of the antidepressant escitalopram. The aim of the current study was to investigate if similar effects occur with another P-gp inhibitor, cyclosporin A (CsA). CsA pre-treatment exacerbated the severity of behaviours in an escitalopram-induced mouse model of serotonin syndrome, a potentially life-threatening adverse drug reaction associated with serotonergic drugs. P-gp inhibition by CsA enhanced the brain distribution of escitalopram by 70-80%. Serotonin (5-HT) turnover in the prefrontal cortex was reduced by escitalopram, and this effect was augmented by CsA. However, CsA pre-treatment did not augment the effect of escitalopram in the tail suspension test (TST) of antidepressant-like activity. Microdialysis experiments revealed that pre-treatment with CsA failed to augment, but blunted, the increase in extracellular 5-HT in response to escitalopram administration. This blunting effect may contribute to the lack of augmentation in the TST. Taken together, the present studies demonstrate that co-administration of CsA and escitalopram produces differential effects depending on the behavioural and neurochemical assays employed. Thus, the results highlight the need for further studies involving more selective pharmacological tools to specifically evaluate the impact of P-gp inhibition on behavioural responses to antidepressants which are subject to efflux by P-gp.

  13. Nanomicelle formulation for topical delivery of cyclosporine A into the cornea: in vitro mechanism and in vivo permeation evaluation

    PubMed Central

    Guo, Chuanlong; Zhang, Yan; Yang, Zhao; Li, Mengshuang; Li, Fengjie; Cui, Fenghua; Liu, Ting; Shi, Weiyun; Wu, Xianggen

    2015-01-01

    A stable topical ophthalmic cyclosporine A (CsA) formulation with good tolerance and high efficacy is still a desire in pharmaceutics and clinics. This article describes the preparation of CsA containing nanomicelles using a polyvinyl caprolactam-polyvinyl acetate-polyethylene glycol (PVCL-PVA-PEG) graft copolymer. Both the polymer itself and the CsA nanomicelles were evaluated for cytotoxicity and ocular irritation. The in vitro uptake and intracellular fate of nanomicelles were characterized. In vivo cornea permeation test performed with 0.5 mg/mL CsA containing nanomicelles, and compared with a commercially available CsA (10 mg/mL) oil-based ophthalmic solution. The CsA nanomicelle ophthalmic solution was simple to prepare and remained storage stable. PVCL-PVA-PEG had no cytotoxicity as its monomer solution, and as its micelle solution (IC50(48 h) = 14.02 mg/mL). CsA nanomicelles also had excellent ocular tolerance in rabbits. The use of nanomicelles significantly improved in vitro cellular uptake, apparently by an energy dependent intracellular endocytosis pathway that involved early endosomes, late endosomes, lysosomes, and ER. In vivo permeation showed that 0.5 mg/mL CsA nanomicelles delivered high levels of CsA into the cornea, when compared to the oil-based 10 mg/mL CsA ophthalmic solution. These findings indicated PVCL-PVA-PEG nanomicelles could be a promising topical delivery system for ocular administration of CsA.

  14. Different Dose-Dependent Mechanisms Are Involved in Early Cyclosporine A-Induced Cholestatic Effects in HepaRG Cells

    PubMed Central

    Sharanek, Ahmad; Azzi, Pamela Bachour-El; Al-Attrache, Houssein; Savary, Camille C.; Humbert, Lydie; Rainteau, Dominique; Guguen-Guillouzo, Christiane; Guillouzo, André

    2014-01-01

    Mechanisms involved in drug-induced cholestasis in humans remain poorly understood. Although cyclosporine A (CsA) and tacrolimus (FK506) share similar immunosuppressive properties, only CsA is known to cause dose-dependent cholestasis. Here, we have investigated the mechanisms implicated in early cholestatic effects of CsA using the differentiated human HepaRG cell line. Inhibition of efflux and uptake of taurocholate was evidenced as early as 15 min and 1 h respectively after addition of 10μM CsA; it peaked at around 2 h and was reversible. These early effects were associated with generation of oxidative stress and deregulation of cPKC pathway. At higher CsA concentrations (≥50μM) alterations of efflux and uptake activities were enhanced and became irreversible, pericanalicular F-actin microfilaments were disorganized and bile canaliculi were constricted. These changes were associated with induction of endoplasmic reticulum stress that preceded generation of oxidative stress. Concentration-dependent changes were observed on total bile acid disposition, which were characterized by an increase and a decrease in culture medium and cells, respectively, after a 24-h treatment with CsA. Accordingly, genes encoding hepatobiliary transporters and bile acid synthesis enzymes were differently deregulated depending on CsA concentration. By contrast, FK506 induced limited effects only at 25–50μM and did not alter bile canaliculi. Our data demonstrate involvement of different concentration-dependent mechanisms in CsA-induced cholestasis and point out a critical role of endoplasmic reticulum stress in the occurrence of the major cholestatic features. PMID:24973091

  15. Combined inhibition of p38 and Akt signaling pathways abrogates cyclosporine A-mediated pathogenesis of aggressive skin SCCs

    SciTech Connect

    Arumugam, Aadithya; Walsh, Stephanie B.; Xu, Jianmin; Afaq, Farrukh; Elmets, Craig A.; Athar, Mohammad

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer p38 and Akt are the crucial molecular targets in the pathogenesis of SCCs in OTRs. Black-Right-Pointing-Pointer Combined inhibition of these targets diminished tumor growth by 90%. Black-Right-Pointing-Pointer Inhibition of these targets act through downregulating mTOR signaling pathway. -- Abstract: Non-melanoma skin cancers (NMSCs) are the most common neoplasm in organ transplant recipients (OTRs). These cancers are more invasive and metastatic as compared to those developed in normal cohorts. Previously, we have shown that immunosuppressive drug, cyclosporine A (CsA) directly alters tumor phenotype of cutaneous squamous cell carcinomas (SCCs) by activating TGF-{beta} and TAK1/TAB1 signaling pathways. Here, we identified novel molecular targets for the therapeutic intervention of these SCCs. We observed that combined blockade of Akt and p38 kinases-dependent signaling pathways in CsA-promoted human epidermoid carcinoma A431 xenograft tumors abrogated their growth by more than 90%. This diminution in tumor growth was accompanied by a significant decrease in proliferation and an increase in apoptosis. The residual tumors following the combined treatment with Akt inhibitor triciribine and p38 inhibitors SB-203580 showed significantly diminished expression of phosphorylated Akt and p38 and these tumors were less invasive and highly differentiated. Diminished tumor invasiveness was associated with the reduced epithelial-mesenchymal transition as ascertained by the enhanced E-cadherin and reduced vimentin and N-cadherin expression. Consistently, these tumors also manifested reduced MMP-2/9. The decreased p-Akt expression was accompanied by a significant reduction in p-mTOR. These data provide first important combinatorial pharmacological approach to block the pathogenesis of CsA-induced highly aggressive cutaneous neoplasm in OTRs.

  16. Effect of cyclosporin A on human bone marrow granulocyte-macrophage progenitors with anti-cancer agents.

    PubMed

    Ishida, Y; Matsuda, H; Kida, K

    1995-10-01

    Cyclosporin A (CyA) overcomes P-glycoprotein (P-gp) associated multidrug resistance (MDR). P-gp expression is frequently observed among, not only various cancer cells, but also several normal tissues including bone marrow progenitor cells. These findings lead us to examine whether CyA enhances the myelotoxicity of anti-cancer agents. Bone marrow mononuclear cells were incubated with anti-cancer agents (vincristine, VCR; doxorubicin, ADM; etoposide, VP-16; cytarabine, Ara-C; methotrexate, MTX) and a concentration of CyA (0.5, 5.0 micrograms/mL). The methylcellulose assay for granulocyte-macrophage progenitors (CFU-GM) was conducted using the post-treated cells. There was no significant toxicity for marrow CFU-GM formation after 72 h incubation with CyA (84-108% of control). The inhibitory concentration that reduced colonies by 50% (IC50) was 12 nmol/L for VCR, 6 nmol/L for ADM, 220 nmol/L for VP-16, 15 nmol/L for Ara-C and 35 nmol/L for MTX, respectively. For VCR, ADM and VP-16, the number of CFU-GM was unchanged with the addition of CyA at 0.5 microgram/mL concentration. In contrast at 5 micrograms/mL CyA, the number of CFU-GM (% of control) was reduced significantly (P < 0.05 or P < 0.01). With MTX and Ara-C, the number of CFU-GM was unchanged after addition of CyA, even at 5 micrograms/mL concentration. We conclude CyA may therefore enhance cytotoxic drug sensitivity in MDR tumor cells at a clinically achievable concentration (0.5 microgram/mL) without marrow toxicity.

  17. On the Performance of Trimetazidine and Vitamin E as Pharmacoprotection Agents in Cyclosporin A-Induced Toxicity

    PubMed Central

    Cristina, De la Cruz Rodríguez Lilia; del Rosario, Rey María; Carmen Rosa, Araujo; Ana Veronica, Oldano

    2013-01-01

    The immunosuppressant drug cyclosporin A (CyA) has been used in diseases with immunological basis and in transplant patients. Nephrotoxicity and hepatotoxicity are the main adverse effects of this drug. To find a protective drug against those effects we assayed the cardioprotector Trimetazidine (TMZ) and vitamin E, used as nutritional supplements to alleviate oxidative stress. Six groups of eight male Wistar rats each were prepared (groups A–F): A, control; B, vitamin E (10 mg/Kg/day); C, TMZ (20 mg/Kg/day); D, 25 mg/Kg/day CyA; E, CyA and vitamin E (25 mg/Kg/day CyA + 10 mg/Kg/day Vit E); F, TMZ for 20 days (20 mg/kg/day); and then CyA (25 mg/kg/day) and TMZ (20 mg/Kg/day). The experiment lasted 120 days. The exposure of rats to CyA promoted nephrotoxicity and hepatotoxicity with an increase in serum urea, creatinine, and glutamate dehydrogenase (GLDH). Structural and ultrastructural studies of liver and kidney were performed. Group D showed adverse effects induced by CyA since statistically significant differences were found with respect to the control group (A). Vitamin E (E) showed no protective effect. Pretreatment with TMZ (F) attenuated the adverse effects of CyA. We conclude that CyA-induced nephrotoxicity and hepatotoxicity are attenuated by the cytoprotective effect of TMZ. TMZ inhibits the reabsorption and, consequently, the accumulation of CyA in the cell. The antioxidant capacity of vitamin E did not improve the effect of CyA. PMID:23691353

  18. Mechanisms of rapid induction of interleukin-22 in activated T cells and its modulation by cyclosporin a.

    PubMed

    Rudloff, Ina; Bachmann, Malte; Pfeilschifter, Josef; Mühl, Heiko

    2012-02-10

    IL-22 is an immunoregulatory cytokine displaying pathological functions in models of autoimmunity like experimental psoriasis. Understanding molecular mechanisms driving IL-22, together with knowledge on the capacity of current immunosuppressive drugs to target this process, may open an avenue to novel therapeutic options. Here, we sought to characterize regulation of human IL22 gene expression with focus on the established model of Jurkat T cells. Moreover, effects of the prototypic immunosuppressant cyclosporin A (CsA) were investigated. We report that IL-22 induction by TPA/A23187 (T/A) or αCD3 is inhibited by CsA or related FK506. Similar data were obtained with peripheral blood mononuclear cells or purified CD3(+) T cells. IL22 promoter analysis (-1074 to +156 bp) revealed a role of an NF-AT (-95/-91 nt) and a CREB (-194/-190 nt) binding site for gene induction. Indeed, binding of CREB and NF-ATc2, but not c-Rel, under the influence of T/A to those elements could be proven by ChIP. Because CsA has the capability to impair IκB kinase (IKK) complex activation, the IKKα/β inhibitor IKKVII was evaluated. IKKVII likewise reduced IL-22 induction in Jurkat cells and peripheral blood mononuclear cells. Interestingly, transfection of Jurkat cells with siRNA directed against IKKα impaired IL22 gene expression. Data presented suggest that NF-AT, CREB, and IKKα contribute to rapid IL22 gene induction. In particular the crucial role of NF-AT detected herein may form the basis of direct action of CsA on IL-22 expression by T cells, which may contribute to therapeutic efficacy of the drug in autoimmunity.

  19. Development of cyclosporine A-loaded dry-emulsion formulation using highly purified glycerol monooleate for safe inhalation therapy.

    PubMed

    Sato, Hideyuki; Ogawa, Kumiko; Kojo, Yoshiki; Kawabata, Yohei; Mizumoto, Takahiro; Yamada, Shizuo; Onoue, Satomi

    2013-05-01

    The main objective of this study was to improve the safety and oxidative stability of glycerol monooleate (GMO)-based dry-emulsion (DE) formulation containing cyclosporine A (CsA) for inhalation therapy. GMO or highly purified GMO (hpGMO) was used as surfactant for the DE formulations (GMO/DE or hpGMO/DE), the toxicological and physicochemical properties of which were characterized with a focus on oxidative stability, in vitro/in vivo toxicity, and dissolution property. Incubation of GMO at oxidation accelerating conditions for 10 days at 60°C resulted in the formation of lipid peroxides as evidenced by increased malondialdehyde (111 μmol/mg); however, hpGMO samples exhibited increase of only 20.7 μmol/mg in malondialdehyde level. No significant acute cytotoxicity was observed in rat alveolar L2 cells exposed to hpGMO (0.28mM), and intratracheal administration of hpGMO powder in rats did not cause an increase of the plasma LDH level. The hpGMO/DE exhibited marked improvement in dissolution behavior of CsA, and stable fine micelles with a mean diameter of 320 nm were formed when suspended in water. A respirable powder formulation of hpGMO/DE (hpGMO/DE-RP) was newly prepared, and its in vitro inhalation property and in vivo efficacy were also evaluated. The hpGMO/DE-RP exhibited high dispersibility in laser diffraction analysis and significantly improved potency to attenuate recruitment of inflammatory cells into airway and thickening of airway wall in an animal model. Thus, the strategic use of hpGMO would improve oxidative stability and local toxicity compared with a GMO-based DE formulation, and its application to RP formulation could be a promising approach for effective inhalation therapy.

  20. Synaptic Mitochondria Sustain More Damage than Non-Synaptic Mitochondria after Traumatic Brain Injury and Are Protected by Cyclosporine A.

    PubMed

    Kulbe, Jacqueline R; Hill, Rachel L; Singh, Indrapal N; Wang, Juan A; Hall, Edward D

    2016-10-13

    Currently, there are no Food and Drug Administration (FDA)-approved pharmacotherapies for the treatment of those with traumatic brain injury (TBI). As central mediators of the secondary injury cascade, mitochondria are promising therapeutic targets for prevention of cellular death and dysfunction after TBI. One of the most promising and extensively studied mitochondrial targeted TBI therapies is inhibition of the mitochondrial permeability transition pore (mPTP) by the FDA-approved drug, cyclosporine A (CsA). A number of studies have evaluated the effects of CsA on total brain mitochondria after TBI; however, no study has investigated the effects of CsA on isolated synaptic and non-synaptic mitochondria. Synaptic mitochondria are considered essential for proper neurotransmission and synaptic plasticity, and their dysfunction has been implicated in neurodegeneration. Synaptic and non-synaptic mitochondria have heterogeneous characteristics, but their heterogeneity can be masked in total mitochondrial (synaptic and non-synaptic) preparations. Therefore, it is essential that mitochondria targeted pharmacotherapies, such as CsA, be evaluated in both populations. This is the first study to examine the effects of CsA on isolated synaptic and non-synaptic mitochondria after experimental TBI. We conclude that synaptic mitochondria sustain more damage than non-synaptic mitochondria 24 h after severe controlled cortical impact injury (CCI), and that intraperitoneal administration of CsA (20 mg/kg) 15 min after injury improves synaptic and non-synaptic respiration, with a significant improvement being seen in the more severely impaired synaptic population. As such, CsA remains a promising neuroprotective candidate for the treatment of those with TBI.

  1. Procarcinogenic effects of cyclosporine A are mediated through the activation of TAK1/TAB1 signaling pathway

    SciTech Connect

    Xu, Jianmin; Walsh, Stephanie B.; Verney, Zoe M.; Kopelovich, Levy; Elmets, Craig A.; Athar, Mohammad

    2011-05-13

    Research highlights: {yields} Organ transplant recipients are highly susceptible to early skin cancer development. {yields} CsA-mediated TGFB1-dependent TAK1/TAB1 signaling augments invasive tumor growth. {yields} CsA enhances accumulation of upstream kinases, ZMP, AMPK and IRAK to activate TAK1. {yields} TAK1 mediates enhanced proliferation and reduced apoptosis via CsA-dependent NF{kappa}B. -- Abstract: Cyclosporine A (CsA) is an immunosuppressive drug commonly used for maintaining chronic immune suppression in organ transplant recipients. It is known that patients receiving CsA manifest increased growth of aggressive non-melanoma skin cancers. However, the underlying mechanism by which CsA augments tumor growth is not fully understood. Here, we show that CsA augments the growth of A431 epidermoid carcinoma xenograft tumors by activating tumor growth factor {beta}-activated kinase1 (TAK1). The activation of TAK1 by CsA occurs at multiple levels by kinases ZMP, AMPK and IRAK. TAK1 forms heterodimeric complexes with TAK binding protein 1 and 2 (TAB1/TAB2) which in term activate nuclear factor {kappa}B (NF{kappa}B) and p38 MAP kinase. Transcriptional activation of NF{kappa}B is evidenced by IKK{beta}-mediated phosphorylation-dependent degradation of I{kappa}B and consequent nuclear translocation of p65. This also leads to enhancement in the expression of its transcriptional target genes cyclin D1, Bcl2 and COX-2. Similarly, activation of p38 leads to enhanced inflammation-related signaling shown by increased phosphorylation of MAPKAPK2 and which in turn phosphorylates its substrate HSP27. Activation of both NF{kappa}B and p38 MAP kinase provide mitogenic stimuli to augment the growth of SCCs.

  2. Effect of azithromycin on gingival overgrowth induced by cyclosporine A + nifedipine combination therapy: A morphometric analysis in rats

    PubMed Central

    Ratre, Madhu Singh; Mehta, Dhoom Singh

    2016-01-01

    Background: Drug-induced gingival overgrowth (DIGO) is a well-known adverse effect of cyclosporine A (CsA) and nifedipine (Nf) therapy. The aim of the present morphometric study was to evaluate the effect of azithromycin (Azi) on the combined GO in rats induced by CsA + Nf combination. Materials and Methods: Thirty Sprague-Dawley male rats were randomly divided equally into three groups. Group 1 (control) received olive oil only; Group 2 received a combination of CsA and Nf in olive oil throughout the study period; Group 3 received CsA + Nf combination therapy, and Azi was added for 1 week in the 5th week. All the drugs were delivered by oral route. Impressions of the mandibular central incisal regions were taken, and study models were prepared at baseline and biweekly up to the 8 weeks. Statistical analysis was done by one-way analysis of variance and intergroup comparisons were made using Tukey's post hoc analysis. Results: Significant GO was evident in Group 2 and Group 3 rats when compared to Group 1. However, in Group 3 (Azi), GO was observed up to the 4th week, but a significant decrease in GO was noticed during 6–8th week after the administration of Azi in 5th week. Conclusion: Azi is an effective drug in the remission of DIGO induced by combined therapy of CsA + Nf and thereby can be considered as a useful therapeutic regimen in minimizing the DIGO in transplant patients. PMID:28298821

  3. Vitamin E protects against the mitochondrial damage caused by cyclosporin A in LLC-PK1 cells

    SciTech Connect

    Arriba, G. de Perez de Hornedo, J.; Ramirez Rubio, S.; Calvino Fernandez, M.; Benito Martinez, S.; Maiques Camarero, M.; Parra Cid, T.

    2009-09-15

    Cyclosporin A (CsA) has nephrotoxic effects known to involve reactive oxygen species (ROS), since antioxidants prevent the kidney damage induced by this drug. Given that mitochondria are among the main sources of intracellular ROS, the aims of our study were to examine the mitochondrial effects of CsA in the porcine renal endothelial cell line LLC-PK1 and the influence of the antioxidant Vitamin E (Vit E). Following the treatment of LLC-PK1 cells with CsA, we assessed the mitochondrial synthesis of superoxide anion, permeability transition pore opening, mitochondrial membrane potential, cardiolipin peroxidation, cytochrome c release and cellular apoptosis, using flow cytometry and confocal microscopy procedures. Similar experiments were done after Vit E preincubation of cells. CsA treatment increased superoxide anion in a dose-dependent way. CsA opened the permeability transition pores, caused Bax migration to mitochondria, and decreased mitochondrial membrane potential and cardiolipin content. Also CsA released cytochrome c into cytosol and provoked cellular apoptosis. Vit E pretreatment inhibited the effects that CsA induced on mitochondrial structure and function in LLC-PK1 cells and avoided apoptosis. CsA modifies mitochondrial LLC-PK1 cell physiology with loss of negative electrochemical gradient across the inner mitochondrial membrane and increased lipid peroxidation. These features are related to apoptosis and can explain the cellular damage that CsA induces. As Vit E inhibited these effects, our results suggest that they were mediated by an increase in ROS production by mitochondria.

  4. The NRF2-heme oxygenase-1 system modulates cyclosporine A-induced epithelial-mesenchymal transition and renal fibrosis

    PubMed Central

    Shin, Dong-ha; Park, Hyun-Min; Jung, Kyeong-Ah; Choi, Han-Gon; Kim, Jung-Ae; Kim, Dae-Duk; Kim, Sang Geon; Kang, Keon Wook; Ku, Sae Kwang; Kensler, Thomas W.; Kwak, Mi-Kyoung

    2013-01-01

    Epithelial-mesenchymal transition (EMT) is an underlying mechanism of tissue fibrosis by generating myofibroblasts, which serve as the primary source of extracellular matrix production from tissue epithelial cells. Recently, it has been suggested that EMT is implicated in immunosuppressive cyclosporine A (CsA)-induced renal fibrosis. In the present study, the potential role of NRF2, which is the master regulator of genes associated with the cellular antioxidant defense system, in CsA-induced EMT-renal fibrosis has been investigated. Pre-treatment of rat tubular epithelial NRK-52E cells with sulforaphane, an activator of NRF2, could prevent EMT gene changes such as the loss of E-cadherin and the increase of α-smooth muscle actin (α-SMA) expression. Conversely, genetic inhibition of NRF2 in these cells aggravated changes in CsA-induced EMT markers. These in vitro observations could be confirmed in vivo: CsA-treatment developed severe renal damage and fibrosis with increased expression of α-SMA in NRF2-deficient mice compared to wild-type mice. NRF2-mediated amelioration of CsA-EMT changes could be accounted in part by the regulation of heme oxygenase-1 (HO-1). CsA treatment increased HO-1 expression in an NRF2-dependent manner in NRK cells as well as murine fibroblasts. Induction of HO-1 by CsA appears to be advantageous by counteracting EMT gene changes: specific increase of HO-1 expression by cobalt protoporphyrin prevented CsA-mediated α-SMA induction, while genetic inhibition of HO-1 by siRNA substantially enhanced α-SMA induction compared to control cells. Collectively, our current results suggest that the NRF2-HO-1 system plays a protective role against CsA-induced renal fibrosis by modulating EMT gene changes. PMID:20096777

  5. Pharmacokinetics of baicalin in rats and its interactions with cyclosporin A, quinidine and SKF-525A: a microdialysis study.

    PubMed

    Tsai, Pi-Lo; Tsai, Tung-Hu

    2004-11-01

    Baicalin, a flavone glucuronide derived mainly from the root of Scutellaria baicalensis, has been used in traditional Chinese medicine as an anti-inflammatory and anti-viral agent. To explore whether the disposition of baicalin is related to multidrug resistance P-glycoprotein (P-gp), baicalin (3, 10 and 30 mg kg(-1); i. v.) was injected to rats for a pharmacokinetic study using microdialysis coupled with HPLC. The results indicate that baicalin goes through hepatobiliary excretion against a concentration gradient based on the blood-to-bile distribution ratio (AUCbile/AUCblood), but that AUCblood or AUCbile did not show any dose-related increase in the range from 3 to 30 mg kg(-1). Coadministration of cyclosporin A (CsA) or quinidine (both are P-gp inhibitors) was used to delineate the role of P-gp on baicalin disposition, while SKF-525A (a cytochrome P450 inhibitor) could specifically inhibit the cytochrome P450 catalysis of baicalin without crossing with P-gp function. Both CsA and quinidine promoted the active transport of baicalin into bile and reduced its level in blood, and this result was the same as that obtained by treating with SKF-525A. Hence, the association of the involvement of P-gp in active baicalin efflux into bile seems to be excluded since CsA and quinidine are also cytochrome P450 inhibitors. In addition, baicalin was not detected in the brain striatum after treating with baicalin alone in the present study. Also, neither CsA nor quinidine co-administered with baicalin is able to induce measurable levels of baicalin in rat brain, which suggests that baicalin might not be able to pass through the blood-brain barrier (BBB).

  6. Active site mutants of human cyclophilin A separate peptidyl-prolyl isomerase activity from cyclosporin A binding and calcineurin inhibition.

    PubMed Central

    Zydowsky, L. D.; Etzkorn, F. A.; Chang, H. Y.; Ferguson, S. B.; Stolz, L. A.; Ho, S. I.; Walsh, C. T.

    1992-01-01

    Based on recent X-ray structural information, six site-directed mutants of human cyclophilin A (hCyPA) involving residues in the putative active site--H54, R55, F60, Q111, F113, and H126--have been constructed, overexpressed, and purified from Escherichia coli to homogeneity. The proteins W121A (Liu, J., Chen, C.-M., & Walsh, C.T., 1991a, Biochemistry 30, 2306-2310), H54Q, R55A, F60A, Q111A, F113A, and H126Q were assayed for cis-trans peptidyl-prolyl isomerase (PPIase) activity, their ability to bind the immunosuppressive drug cyclosporin A (CsA), and protein phosphatase 2B (calcineurin) inhibition in the presence of CsA. Results indicate that H54Q, Q111A, F113A, and W121A retain 3-15% of the catalytic efficiency (kcat/Km) of wild-type recombinant hCyPA. The remaining three mutants (R55A, F60A, and H126Q) each retain less than 1% of the wild-type catalytic efficiency, indicating participation by these residues in PPIase catalysis. Each of the mutants bound to a CsA affinity matrix. The mutants R55A, F60A, F113A, and H126Q inhibited calcineurin in the presence of CsA, whereas W121A did not. Although CsA is a competitive inhibitor of PPIase activity, it can complex with enzymatically inactive cyclophilins and inhibit the phosphatase activity of calcineurin. PMID:1338979

  7. Cyclosporine A Treatment Inhibits Abcc6-Dependent Cardiac Necrosis and Calcification following Coxsackievirus B3 Infection in Mice

    PubMed Central

    Marton, Jennifer; Albert, Danica; Wiltshire, Sean A.; Park, Robin; Bergen, Arthur; Qureshi, Salman; Malo, Danielle; Burelle, Yan; Vidal, Silvia M.

    2015-01-01

    Coxsackievirus type B3 (CVB3) is a cardiotropic enterovirus. Infection causes cardiomyocyte necrosis and myocardial inflammation. The damaged tissue that results is replaced with fibrotic or calcified tissue, which can lead to permanently altered cardiac function. The extent of pathogenesis among individuals exposed to CVB3 is dictated by a combination of host genetics, viral virulence, and the environment. Here, we aimed to identify genes that modulate cardiopathology following CVB3 infection. 129S1 mice infected with CVB3 developed increased cardiac pathology compared to 129X1 substrain mice despite no difference in viral burden. Linkage analysis identified a major locus on chromosome 7 (LOD: 8.307, P<0.0001) that controlled the severity of cardiac calcification and necrosis following infection. Sub-phenotyping and genetic complementation assays identified Abcc6 as the underlying gene. Microarray expression profiling identified genotype-dependent regulation of genes associated with mitochondria. Electron microscopy examination showed elevated deposition of hydroxyapatite-like material in the mitochondrial matrices of infected Abcc6 knockout (Abcc6-/-) mice but not in wildtype littermates. Cyclosporine A (CsA) inhibits mitochondrial permeability transition pore opening by inhibiting cyclophilin D (CypD). Treatment of Abcc6 -/- mice with CsA reduced cardiac necrosis and calcification by more than half. Furthermore, CsA had no effect on the CVB3-induced phenotype of doubly deficient CypD-/-Abcc6-/- mice. Altogether, our work demonstrates that mutations in Abcc6 render mice more susceptible to cardiac calcification following CVB3 infection. Moreover, we implicate CypD in the control of cardiac necrosis and calcification in Abcc6-deficient mice, whereby CypD inhibition is required for cardioprotection. PMID:26375467

  8. PEG-b-PPS diblock copolymer aggregates for hydrophobic drug solubilization and release: cyclosporin A as an example.

    PubMed

    Velluto, Diana; Demurtas, Davide; Hubbell, Jeffrey A

    2008-01-01

    Micelles formed from amphiphilic block copolymers have been explored in recent years as carriers for hydrophobic drugs. In an aqueous environment, the hydrophobic blocks form the core of the micelle, which can host lipophilic drugs, while the hydrophilic blocks form the corona or outer shell and stabilize the interface between the hydrophobic core and the external medium. In the present work, mesophase behavior and drug encapsulation were explored in the AB block copolymeric amphiphile composed of poly(ethylene glycol) (PEG) as a hydrophile and poly(propylene sulfide) PPS as a hydrophobe, using the immunosuppressive drug cyclosporin A (CsA) as an example of a highly hydrophobic drug. Block copolymers with a degree of polymerization of 44 on the PEG and of 10, 20 and 40 on the PPS respectively (abbreviated as PEG44-b-PPS10, PEG44-b-PPS20, PEG44-b-PPS40) were synthesized and characterized. Drug-loaded polymeric micelles were obtained by the cosolvent displacement method as well as the remarkably simple method of dispersing the warm polymer melt, with drug dissolved therein, in warm water. Effective drug solubility up to 2 mg/mL in aqueous media was facilitated by the PEG- b-PPS micelles, with loading levels up to 19% w/w being achieved. Release was burst-free and sustained over periods of 9-12 days. These micelles demonstrate interesting solubilization characteristics, due to the low glass transition temperature, highly hydrophobic nature, and good solvent properties of the PPS block.

  9. Adrenochrome reaction implicates oxygen radicals in metabolism of cyclosporine A and FK-506 in rat and human liver microsomes.

    PubMed

    Ahmed, S S; Strobel, H W; Napoli, K L; Grevel, J

    1993-06-01

    The role of oxygen radicals in the metabolism of cyclosporine A (CyA), FR900506 (FK-506) and carbon tetrachloride (CCl4) catalyzed by the cytochrome P450 system was investigated in vitro in rat and human microsomal preparations. Varying concentrations of CyA, FK-506 and CCl4 (100 microM-1.0 mM) were added to microsomal preparations, and lipid peroxidation was measured by malondialdehyde (MDA) formation as detected by the thiobarbituric acid assay. The effects of oxygen radical scavengers [superoxide dismutase (SOD) and catalase (CAT)] and an antioxidant [glutathione (GLUT)] were tested on various incubations of CyA, FK-506 and CCl4 to assess the role of oxygen radicals in lipid peroxidation. CyA-dependent MDA formation was moderately inhibited by SOD in the rat model and increased by SOD in the human model. In both models, CAT slightly inhibited CyA-dependent MDA formation and GLUT significantly inhibited MDA formation. FK-506-dependent MDA formation, studied only in the rat model, paralleled CyA-induced MDA formation but showed greater inhibition with CAT and less inhibition with SOD or GLUT. In both models, CCl4-dependent MDA formation was significantly inhibited by GLUT and showed no sensitivity to SOD or CAT. In addition, the adrenochrome reaction, which measures the oxidation of epinephrine to adrenochrome, was used to measure the increased oxygen radical-flux resulting from the metabolism of CyA, FK-506 and CCl4. CyA with epinephrine showed the highest oxidative activity, followed by FK-506 and then CCl4, which showed the least formation of adrenochrome. These results indicated a role for oxygen radicals in CyA and FK-506 metabolism.

  10. Inhibition of the mitochondrial permeability transition by cyclosporin A prevents pyrazole plus lipopolysaccharide-induced liver injury in mice.

    PubMed

    Zhuge, Jian; Cederbaum, Arthur I

    2009-02-01

    Previous results showed that pyrazole potentiates lipopolysaccharide (LPS)-induced liver injury in mice. Mechanisms involved the overexpression of cytochrome P450 2E1 (CYP2E1), oxidative stress, and activation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK). The current study was carried out to test the hypothesis that the mitochondria permeability transition (MPT) plays a role in this pyrazole plus LPS toxicity. Mice were injected intraperitoneally with pyrazole for 2 days, followed by a challenge with LPS with or without treatment with cyclosporin A (CsA), an inhibitor of the MPT. Serum alanine aminotransferase and aspartate aminotransferase were increased by pyrazole plus LPS treatment, and CsA treatment could attenuate these increases. CsA also prevented pyrazole plus LPS-induced hepatocyte necrosis. Formation of 4-hydroxynonenal protein adducts and 3-nitrotyrosine protein adducts in liver tissue was increased by the pyrazole plus LPS treatment, and CsA treatment blunted these increases. Swelling, cytochrome c release from mitochondria to the cytosol, and lipid peroxidation were increased in mitochondria isolated from the pyrazole plus LPS-treated mice, and CsA treatment prevented these changes. CsA did not prevent the increased levels of inducible nitric oxide synthase (iNOS), tumor necrosis factor-alpha (TNF-alpha), pp38 MAPK, and p-JNK2. In conclusion, although CsA does not prevent elevations in upstream mediators of the pyrazole plus LPS toxicity (iNOS, TNF-alpha, CYP2E1, MAPK), it does protect mice from the pyrazole plus LPS-induced liver toxicity by preventing the MPT and release of cytochrome c and decreasing mitochondrial oxidative stress. These results indicate that mitochondria are the critical targets of pyrazole plus LPS in mediating liver injury.

  11. Systems biology modeling of omics data: effect of cyclosporine a on the Nrf2 pathway in human renal cells

    PubMed Central

    2014-01-01

    Background Incorporation of omic data streams for building improved systems biology models has great potential for improving their predictions of biological outcomes. We have recently shown that cyclosporine A (CsA) strongly activates the nuclear factor (erythroid-derived 2)-like 2 pathway (Nrf2) in renal proximal tubular epithelial cells (RPTECs) exposed in vitro. We present here a quantitative calibration of a differential equation model of the Nrf2 pathway with a subset of the omics data we collected. Results In vitro pharmacokinetic data on CsA exchange between cells, culture medium and vial walls, and data on the time course of omics markers in response to CsA exposure were reasonably well fitted with a coupled PK-systems biology model. Posterior statistical distributions of the model parameter values were obtained by Markov chain Monte Carlo sampling in a Bayesian framework. A complex cyclic pattern of ROS production and control emerged at 5 μM CsA repeated exposure. Plateau responses were found at 15 μM exposures. Shortly above those exposure levels, the model predicts a disproportionate increase in cellular ROS quantity which is consistent with an in vitro EC50 of about 40 μM for CsA in RPTECs. Conclusions The model proposed can be used to analyze and predict cellular response to oxidative stress, provided sufficient data to set its parameters to cell-specific values. Omics data can be used to that effect in a Bayesian statistical framework which retains prior information about the likely parameter values. PMID:24964791

  12. Functional and morphological analysis of the subretinal injection of human retinal progenitor cells under Cyclosporin A treatment

    PubMed Central

    Huang, Rui; Baranov, Petr; Lai, Kunbei; Zhang, Xinmei; Ge,, Jian

    2014-01-01

    Purpose The purpose of this study is to evaluate the functional and morphological changes in subretinal xenografts of human retinal progenitor cells (hRPCs) in B6 mice treated with Cyclosporin A (CsA; 210 mg/l in drinking water). Methods The hRPCs from human fetal eyes were isolated and expanded for transplantation. These cells, with green fluorescent protein (GFP) at 11 passages, were transplanted into the subretinal space in B6 mice. A combination of invasive and noninvasive approaches was used to analyze the structural and functional consequences of the subretinal injection of the hRPCs. The process of change was monitored using spectral domain optical coherence tomography (SDOCT), histology, and electroretinography (ERG) at 3 days, 1 week, and 3 weeks after transplantation. Cell counts were used to evaluate the survival rate with a confocal microscope. ERGs were performed to evaluate the physiologic changes, and the structural changes were evaluated using SDOCT and histological examination. Results The results of the histological examination showed that the hRPCs gained a better survival rate in the mice treated with CsA. The SDOCT showed that the bleb size of the retinal detachment was significantly decreased, and the retinal reattachment was nearly complete by 3 weeks. The ERG response amplitudes in the CsA group were less decreased after the injection, when compared with the control group, in the dark-adapted and light-adapted conditions. However, the cone-mediated function in both groups was less affected by the transplantation after 3 weeks than the rod-mediated function. Conclusions Although significant functional and structural recovery was observed after the subretinal injection of the hRPCs, the effectiveness of CsA in xenotransplantation may be a novel and potential approach for increasing retinal progenitor cell survival. PMID:25352736

  13. Acute, subacute and chronic effect of cyclosporin-A on mean arterial pressure of rats with severe spinal cord contusion.

    PubMed

    Romero, Samanta E; Bravo, Guadalupe; Hong, Enrique; Rojas, Guillermo; Ibarra, Antonio

    2008-11-07

    Cyclosporin-A (CsA) protects and regenerates the neural tissue after spinal cord (SC) injury. These beneficial effects are achieved when CsA is administered at a dose of 2.5mg/kg/12h during the first 2 days after lesion. In view of these observations, it is realistic to envision that, CsA could be tested in SC-clinical trials. Since CsA is a drug strongly related to hypertension, results imperative to evaluate experimentally the effect of the above CsA-dose regimen on blood pressure. For this purpose, one hundred and twenty adult rats were subjected (10 groups) or not (10 groups) to SC-injury. Five injured and five Sham-operated groups received CsA. The remaining groups received only vehicle. Mean arterial pressure (MAP) was recorded from these animals at acute (6 and 24h post surgery; p.s.), subacute (96h), or chronic (30 days) stages of injury. In the latter, the therapy (CsA or vehicle) was administered only during the first 2 days p.s. or daily during 30 days of follow-up. The results of this study showed that SC-injury by itself induces a significant decrease of MAP during the acute and subacute phases of injury. CsA therapy was able to reestablish MAP parameters to control values in these phases. Regardless the therapy, a reestablishment of MAP was observed in chronic stages. Only the daily administration of CsA induced a significant increase in MAP, however; such variation remained into the normal ranges of MAP for rats. The potential benefits offered by CsA support its usefulness after SC-injury.

  14. HMG-CoA reductase inhibitor-induced myopathy in the rat: cyclosporine A interaction and mechanism studies.

    PubMed

    Smith, P F; Eydelloth, R S; Grossman, S J; Stubbs, R J; Schwartz, M S; Germershausen, J I; Vyas, K P; Kari, P H; MacDonald, J S

    1991-06-01

    Recent clinical evidence indicates a potential for skeletal muscle toxicity after therapy with HMG-CoA reductase inhibitors (HMGRIs) in man. Although the incidence of drug-induced skeletal muscle toxicity is very low (0.1-0.2%) with monotherapy, it may increase following concomitant drug therapy with the immunosuppressant, cyclosporine A (CsA), and possibly with certain other hypolipidemic agents. In the Sprague-Dawley rat, very high, pharmacologically comparable dosages (150-1200 mg/kg/day) of structurally similar HMGRIs (lovastatin, simvastatin, pravastatin and L-647, 318) produced dose-related increases in the incidence and severity of skeletal muscle degeneration. Physical signs included inappetence, decreased activity, loss of body weight, localized alopecia and mortality. To evaluate the interaction between HMGRIs and CsA, a rat model of CsA-induced cholestasis was developed. In this 2-week model, the skeletal muscle toxicity of the HMGRIs was clearly potentiated by CsA (10 mg/kg/day). Doses of HMGRIs which did not produce skeletal muscle toxicity when given alone caused between 75 and 100% incidence of myopathy (very slight to marked skeletal muscle degeneration) when CsA was coadministered. Typical light microscopic changes included myofiber necrosis with interstitial edema and inflammatory infiltration in areas of acute injury. Histochemical characterization of the muscle lesion indicated that type 2B fibers (primarily glycolytic white fibers) were most sensitive to this toxicity but that, with prolonged administration, all fiber types were ultimately affected. Results of pharmacokinetic studies in rats treated with various HMGRIs +/- CsA indicated that coadministration of CsA alters the disposition of these compounds, resulting in increased systemic exposure (e.g., increased area under the plasma drug concentration vs. time curve-AUC) and consequent (up to 13-fold) increases in skeletal muscle drug levels. Evaluation of the potential interaction between

  15. Everolimus in combination with cyclosporin a as pre- and posttransplantation immunosuppressive therapy in nonmyeloablative allogeneic hematopoietic stem cell transplantation.

    PubMed

    Junghanss, Christian; Rathsack, Susanne; Wacke, Rainer; Weirich, Volker; Vogel, Heike; Drewelow, Bernd; Mueller, Sabrina; Altmann, Simone; Freund, Mathias; Lange, Sandra

    2012-07-01

    Everolimus (RAD001) is an mTOR inhibitor that has been successfully used as an immunosuppressant in solid-organ transplantation. Data in allogeneic hematopoietic stem cell transplantation (HSCT) is limited. This study aimed to investigate pharmacokinetics, safety, and efficacy of RAD001 in a canine allogeneic HSCT model. First, pharmacokinetics of RAD001 were performed in healthy dogs in order to determine the appropriate dosing. Doses of 0.25 mg RAD001 twice daily in combination with 15 mg/kg cyclosporin A (CsA) twice daily were identified as appropriate starting doses to achieve the targeted range of RAD001 (3-8 μg/L) when orally administered. Subsequently, 10 dogs were transplanted using 2 Gy total body irradiation (TBI) for conditioning and 0.25 mg RAD001 twice daily plus 15 mg/kg CsA twice daily for pre- and posttransplantation immunosuppression. Seven of the 10 transplanted dogs were maintained at the starting RAD001 dose throughout the study. For the remaining 3 dogs, dose adjustments were necessary. RAD001 accumulation over time did not occur. All dogs initially engrafted. Five dogs eventually rejected the graft (weeks 10, 10, 13, 27, and 56). Two dogs died of pneumonia (weeks 8 and 72) but were chimeric until then. Total cholesterol rose from median 4.1 mmol/L (3.5-5.7 mmol/L) before HSCT to 6.0 mmol/l (5.0-8.5 mmol/l) at day 21 after HSCT, but remained always within normal range. Changes in creatinine and triglyceride values were not observed. Long-term engraftment rates were inferior to sirolimus/CsA and mycophenolate mofetil (MMF)/CsA regimen, respectively. RAD001/CsA caused a more pronounced reduction of platelet counts to median 2 × 10(9)/L (range: 0-21 × 10(9)/L) and longer time to platelet recovery of 21 days (range: 14-24 days) compared with MMF/CsA. CsA c(2h) levels were significantly enhanced in the RAD001/CsA regimen, but c(0h) and area under the curve from 0 to 12 hours (AUC(0-12h)) values did not differ compared with an MMF

  16. Pharmacokinetic and nephroprotective benefits of using Schisandra chinensis extracts in a cyclosporine A-based immune-suppressive regime.

    PubMed

    Lai, Qiao; Wei, Jiabao; Mahmoodurrahman, Mohammed; Zhang, Chenxue; Quan, Shijian; Li, Tongming; Yu, Yang

    2015-01-01

    Cyclosporine A (CsA) is a powerful immunosuppressive drug. However, nephrotoxicity resulting from its long-term usage has hampered its prolonged therapeutic usage. Schisandra chinensis extracts (SCE) have previously been used in traditional Chinese medicine and more recently coadministered with Western medicine for the treatment of CsA-induced side effects in the People's Republic of China. This study aimed to investigate the possible effects of SCE on the pharmacokinetics of CsA in rats and elucidate the potential mechanisms by which it hinders the development of CsA-induced nephrotoxicity. A liquid chromatography/tandem mass spectrometry method was developed and validated for determining the effect of SCE on the pharmacokinetics of CsA. Male Sprague Dawley rats, which were administered with CsA (25 mg/kg/d) alone or in combination with SCE (54 mg/kg/d and 108 mg/kg/d) for 28 days, were used to evaluate the nephroprotective effects of SCE. Our study showed that SCE increased the mean blood concentration of CsA. Furthermore, we found that the concomitant administration of SCE alongside CsA prevented the disruption of catalase activity and reduction in creatinine, urea, renal malondialdehyde, and glutathione peroxidase levels that would have otherwise occurred in the absence of SCE administration. SCE treatment markedly suppressed the expression of 4-hydroxynonenal, Bcl-2-associated X protein, cleaved caspase 3, and autophagy-related protein LC3 A/B. On the other hand, the expression of heme oxygenase-1, nuclear factor erythroid 2-related factor 2 (Nrf2), and P-glycoprotein was enhanced by the very same addition of SCE. SCE was also able to increase the systemic exposure of CsA in rats. The renoprotective effects of SCE were thought to be mediated by its antiapoptotic and antioxidant abilities, which caused the attenuation of CsA-induced autophagic cell death. All in all, these findings suggest the prospective use of SCE as an effective adjunct in a Cs

  17. Pharmacokinetic and nephroprotective benefits of using Schisandra chinensis extracts in a cyclosporine A-based immune-suppressive regime

    PubMed Central

    Lai, Qiao; Wei, Jiabao; Mahmoodurrahman, Mohammed; Zhang, Chenxue; Quan, Shijian; Li, Tongming; Yu, Yang

    2015-01-01

    Cyclosporine A (CsA) is a powerful immunosuppressive drug. However, nephrotoxicity resulting from its long-term usage has hampered its prolonged therapeutic usage. Schisandra chinensis extracts (SCE) have previously been used in traditional Chinese medicine and more recently coadministered with Western medicine for the treatment of CsA-induced side effects in the People’s Republic of China. This study aimed to investigate the possible effects of SCE on the pharmacokinetics of CsA in rats and elucidate the potential mechanisms by which it hinders the development of CsA-induced nephrotoxicity. A liquid chromatography/tandem mass spectrometry method was developed and validated for determining the effect of SCE on the pharmacokinetics of CsA. Male Sprague Dawley rats, which were administered with CsA (25 mg/kg/d) alone or in combination with SCE (54 mg/kg/d and 108 mg/kg/d) for 28 days, were used to evaluate the nephroprotective effects of SCE. Our study showed that SCE increased the mean blood concentration of CsA. Furthermore, we found that the concomitant administration of SCE alongside CsA prevented the disruption of catalase activity and reduction in creatinine, urea, renal malondialdehyde, and glutathione peroxidase levels that would have otherwise occurred in the absence of SCE administration. SCE treatment markedly suppressed the expression of 4-hydroxynonenal, Bcl-2-associated X protein, cleaved caspase 3, and autophagy-related protein LC3 A/B. On the other hand, the expression of heme oxygenase-1, nuclear factor erythroid 2-related factor 2 (Nrf2), and P-glycoprotein was enhanced by the very same addition of SCE. SCE was also able to increase the systemic exposure of CsA in rats. The renoprotective effects of SCE were thought to be mediated by its antiapoptotic and antioxidant abilities, which caused the attenuation of CsA-induced autophagic cell death. All in all, these findings suggest the prospective use of SCE as an effective adjunct in a Cs

  18. A pharmacologically-based array to identify targets of cyclosporine A-induced toxicity in cultured renal proximal tubule cells

    SciTech Connect

    Sarró, Eduard; Jacobs-Cachá, Conxita; Itarte, Emilio; Meseguer, Anna

    2012-01-15

    Mechanisms of cyclosporine A (CsA)-induced nephrotoxicity were generally thought to be hemodynamic in origin; however, there is now accumulating evidence of a direct tubular effect. Although genomic and proteomic experiments by our group and others provided overall information on genes and proteins up- or down-regulated by CsA in proximal tubule cells (PTC), a comprehensive view of events occurring after CsA exposure remains to be described. For this purpose, we applied a pharmacologic approach based on the use of known activities of a large panel of potentially protective compounds and evaluated their efficacy in preventing CsA toxicity in cultured mouse PTC. Our results show that compounds that blocked protein synthesis and apoptosis, together with the CK2 inhibitor DMAT and the PI3K inhibitor apigenin, were the most efficient in preventing CsA toxicity. We also identified GSK3, MMPs and PKC pathways as potential targets to prevent CsA damage. Additionally, heparinase-I and MAPK inhibitors afforded partial but significant protection. Interestingly, antioxidants and calcium metabolism-related compounds were unable to ameliorate CsA-induced cytotoxicity. Subsequent experiments allowed us to clarify the hierarchical relationship of targeted pathways after CsA treatment, with ER stress identified as an early effector of CsA toxicity, which leads to ROS generation, phenotypical changes and cell death. In summary, this work presents a novel experimental approach to characterizing cellular responses to cytotoxics while pointing to new targets to prevent CsA-induced toxicity in proximal tubule cells. Highlights: ► We used a novel pharmacological approach to elucidate cyclosporine (CsA) toxicity. ► The ability of a broad range of compounds to prevent CsA toxicity was evaluated. ► CsA toxicity was monitored using LDH release assay and PARP cleavage. ► Protein synthesis, PI3K, GSK3, MMP, PKC and caspase inhibitors prevented CsA toxicity. ► We also identified ER

  19. Green tea polyphenols stimulate mitochondrial biogenesis and improve renal function after chronic cyclosporin a treatment in rats.

    PubMed

    Rehman, Hasibur; Krishnasamy, Yasodha; Haque, Khujista; Thurman, Ronald G; Lemasters, John J; Schnellmann, Rick G; Zhong, Zhi

    2014-01-01

    Our previous studies showed that an extract from Camellia sinenesis (green tea), which contains several polyphenols, attenuates nephrotoxicity caused by cyclosporine A (CsA). Since polyphenols are stimulators of mitochondrial biogenesis (MB), this study investigated whether stimulation of MB plays a role in green tea polyphenol protection against CsA renal toxicity. Rats were fed a powdered diet containing green tea polyphenolic extract (0.1%) starting 3 days prior to CsA treatment (25 mg/kg, i.g. daily for 3 weeks). CsA alone decreased renal nuclear DNA-encoded oxidative phosphorylation (OXPHOS) protein ATP synthase-β (AS-β) by 42%, mitochondrial DNA (mtDNA)-encoded OXPHOS protein NADH dehydrogenase-3 (ND3) by 87% and their associated mRNAs. Mitochondrial DNA copy number was also decreased by 78% by CsA. Immunohistochemical analysis showed decreased cytochrome c oxidase subunit IV (COX-IV), an OXPHOS protein, in tubular cells. Peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α, the master regulator of MB, and mitochondrial transcription factor-A (Tfam), the transcription factor that regulates mtDNA replication and transcription, were 42% and 90% lower, respectively, in the kidneys of CsA-treated than in untreated rats. These results indicate suppression of MB by chronic CsA treatment. Green tea polyphenols alone and following CsA increased AS-β, ND3, COX-IV, mtDNA copy number, PGC-1α mRNA and protein, decreased acetylated PGC-1α, and increased Tfam mRNA and protein. In association with suppressed MB, CsA increased serum creatinine, caused loss of brush border and dilatation of proximal tubules, tubular atrophy, vacuolization, apoptosis, calcification, and increased neutrophil gelatinase-associated lipocalin expression, leukocyte infiltration, and renal fibrosis. Green tea polyphenols markedly attenuated CsA-induced renal injury and improved renal function. Together, these results demonstrate that green tea polyphenols attenuate Cs

  20. Effects of immunosuppressants, FK506 and cyclosporin A, on the osteogenic differentiation of rat mesenchymal stem cells

    PubMed Central

    Byun, Yu-Kyung; Kim, Kyoung-Hwa; Kim, Su-Hwan; Kim, Young-Sung; Koo, Ki-Tae; Kim, Tai-Il; Seol, Yang-Jo; Ku, Young; Rhyu, In-Chul

    2012-01-01

    Purpose The purpose of this study was to investigate the effects of the immunosuppressants FK506 and cyclosporin A (CsA) on the osteogenic differentiation of rat mesenchymal stem cells (MSCs). Methods The effect of FK506 and CsA on rat MSCs was assessed in vitro. The MTT assay was used to determine the deleterious effect of immunosuppressants on stem cell proliferation at 1, 3, and 7 days. Alkaline phosphatase (ALP) activity was analyzed on days 3, 7, and 14. Alizarin red S staining was done on day 21 to check mineralization nodule formation. Real-time polymerase chain reaction (RT-PCR) was also performed to detect the expressions of bone tissue-specific genes on days 1 and 7. Results Cell proliferation was promoted more in the FK506 groups than the control or CsA groups on days 3 and 7. The FK506 groups showed increased ALP activity compared to the other groups during the experimental period. The ALP activity of the CsA groups did not differ from the control group in any of the assessments. Mineralization nodule formation was most prominent in the FK506 groups at 21 days. RT-PCR results of the FK506 groups showed that several bone-related genes-osteopontin, osteonectin, and type I collagen (Col-I)-were expressed more than the control in the beginning, but the intensity of expression decreased over time. Runx2 and Dlx5 gene expression were up-regulated on day 7. The effects of 50 nM CsA on osteonectin and Col-I were similar to those of the FK506 groups, but in the 500 nM CsA group, most of the genes were less expressed compared to the control. Conclusions These results suggest that FK506 enhances the osteoblastic differentiation of rat MSCs. Therefore, FK506 might have a beneficial effect on bone regeneration when immunosuppressants are needed in xenogenic or allogenic stem cell transplantation to treat bone defects. PMID:22803008

  1. Cyclosporin A inhibits flow-mediated activation of endothelial nitric-oxide synthase by altering cholesterol content in caveolae.

    PubMed

    Lungu, Andreea O; Jin, Zheng-Gen; Yamawaki, Hideyuki; Tanimoto, Tatsuo; Wong, Chelsea; Berk, Bradford C

    2004-11-19

    Fluid shear stress generated by blood flowing over the endothelium is a major determinant of arterial tone, vascular remodeling, and atherogenesis. Nitric oxide (NO) produced by endothelial NO synthase (eNOS) plays an essential role in regulation of vascular function and structure by blood flow. Although cyclosporin A (CsA), an inhibitory ligand of cyclophilin A, is a widely used immunosuppressive drug, it causes arterial hypertension in part by impairing eNOS-dependent vasodilation. Here we show that CsA inhibits fluid shear stress-mediated eNOS activation in endothelial cells via decreasing cholesterol content in caveolae. Exposure of cultured bovine aortic endothelial cells to 1 mum CsA for 1 h significantly inhibited NO production and eNOS phosphorylation at Ser-1179 induced by flow (shear stress=dynes/cm2). The effect of CsA was not related to inhibition of two known eNOS kinases, protein kinase B (Akt) and protein kinase A, because CsA did not affect Akt or protein kinase A activation. In rabbit aorta perfused ex vivo, CsA also significantly inhibited flow-induced eNOS phosphorylation at Ser-1179 but had no effect on Akt measured by phosphorylation at Ser-473. However, CsA treatment decreased cholesterol content in caveolae and displaced eNOS from caveolae, which may be caused by CsA disrupting the association of caveolin-1 and cyclophilin A. The magnitude of the cholesterol depleting effect was similar to that of beta-cyclodextrin, a cholesterol-binding molecule, and beta-cyclodextrin had a similar inhibitory effect on flow-mediated eNOS activation. Treating bovine aortic endothelial cells for 24 h with 30 mug/ml cholesterol blocked the CsA effect and restored eNOS phosphorylation in response to flow. These data suggest that decreasing cholesterol content in caveolae by CsA is a potentially important pathogenic mechanism for CsA-induced endothelial dysfunction and hypertension.

  2. Cyclosporin A and doxorubicin-ifosfamide in resistant solid tumours: a phase I and an immunological study.

    PubMed Central

    González-Manzano, R.; Cid, J.; Brugarolas, A.; Piasecki, C. C.

    1995-01-01

    In order to test whether circumvention of clinical resistance can be obtained in common solid tumours by targeting different drug resistance mechanisms, a phase I clinical and immunological study was designed. The purpose of the study was to determine the dose of cyclosporin A (CsA), in combination with doxorubicin (DOX) and ifosfamide (IFX), needed to achieve steady-state whole-blood levels of 2000 ng ml-1 and the associated toxicity of this combination. Treatment consisted of CsA 5 mg kg-1 as a 2 h loading infusion, followed by a CsA 3 day continuous infusion (c.i.) (days 1-3) at doses that were escalated from 10 to 18 mg kg-1 day-1. Chemotherapy consisted of DOX 55 mg m-2 by i.v. 24 h c.i. (day 2) and IFX 2 g m-2 i.v. over 1 h on days 1 and 3. Treatments were repeated every 4 weeks. Eighteen patients with previously treated resistant solid tumours received 39 cycles. Mean steady-state CsA levels > or = 2000 ng ml-1 were reached at 5 mg kg-1 loading dose followed by a 3 day c.i. of 16 mg kg-1 day-1 or greater. Haematological toxicity was greater than expected for the same chemotherapy alone. One patient died of intracranial haemorrhage due to severe thrombopenia. Other observed toxicities were: asymptomatic hyperbilirubinaemia (46% cycles), mild nephrotoxicity (20% cycles), hypomagnesaemia (72% cycles), mild increase in body weight (100% cycles), hypertension (15% cycles) and headache (15% cycles). Overall the toxicity was acceptable and manageable. No alterations in absolute lymphocyte number, the lymphocyte subsets studied (CD3, CD4, CD8, CD19) or CD4/CD8 ratio were observed in patients receiving more than one treatment cycle, although there were significant and non-uniform variations in the values of the different lymphocyte subsets studied when pre- and post-treatment values were compared. There was also a significant increase in the CD4/CD8 ratio. Tumour regressions were observed in two patients (epidermoid carcinoma of the cervix and Ewing's sarcoma). The

  3. The hepatitis B virus X protein activates nuclear factor of activated T cells (NF-AT) by a cyclosporin A-sensitive pathway.

    PubMed Central

    Lara-Pezzi, E; Armesilla, A L; Majano, P L; Redondo, J M; López-Cabrera, M

    1998-01-01

    The X gene product of the human hepatitis B virus (HBx) is a transcriptional activator of various viral and cellular genes. We recently have determined that the production of tumor necrosis factor-alpha (TNF-alpha) by HBV-infected hepatocytes is transcriptionally up-regulated by HBx, involving nuclear factor of activated T cells (NF-AT)-dependent activation of the TNF-alpha gene promoter. Here we show that HBx activates NF-AT by a cyclosporin A-sensitive mechanism involving dephosphorylation and nuclear translocation of the transcription factor. Luciferase gene expression assays demonstrated that HBx transactivates transcription through NF-AT-binding sites and activates a Gal4-NF-AT chimeric protein. DNA-protein interaction assays revealed that HBx induces the formation of NF-AT-containing DNA-binding complexes. Immunofluorescence analysis demonstrated that HBx induces the nuclear translocation of NF-AT, which can be blocked by the immunosuppressive drug cyclosporin A. Furthermore, immunoblot analysis showed that the HBx-induced activation and translocation of NF-AT are associated with its dephosphorylation. Thus, HBx may play a relevant role in the intrahepatic inflammatory processes by inducing locally the expression of cytokines that are regulated by NF-AT. PMID:9843511

  4. Transforming growth factor beta and cyclosporin A inhibit the inducible activity of the interleukin-2 gene in T cells through a noncanonical octamer-binding site.

    PubMed Central

    Brabletz, T; Pfeuffer, I; Schorr, E; Siebelt, F; Wirth, T; Serfling, E

    1993-01-01

    Transforming growth factor beta (TGF-beta) has a growth-inhibitory effect on numerous different cell types of the immune system, including T lymphocytes. We show in this study that the inhibitory action of TGF-beta on T lymphocytes is accompanied by a block of interleukin 2 (IL-2) gene expression which is mediated, at least in part, by inhibition of IL-2 promoter/enhancer activity. The functional analysis of cis-regulatory (proto-enhancer) elements of the IL-2 enhancer/promoter region showed that the most TGF-beta-responsive element maps to its so-called upstream promoter site. The proto-enhancer activity of the upstream promoter site element is also inhibited by cyclosporin A. The upstream promoter site DNA harbors two noncanonical, closely linked binding sequences for octamer and AP-1-like factors. Both sites are involved in the establishment of IL-2 enhancer activity. Since the activity of genuine octamer sites but not that of AP-1-binding sites is also impaired by TGF-beta and cyclosporin A in El4 T lymphoma cells, we conclude that both immunosuppressives interfere with the activity but not the DNA binding of octamer factors in T lymphocytes. Images PMID:8423782

  5. Novel micelle carriers for cyclosporin A topical ocular delivery: in vivo cornea penetration, ocular distribution and efficacy studies.

    PubMed

    Di Tommaso, Claudia; Bourges, Jean-Louis; Valamanesh, Fatemeh; Trubitsyn, Gregory; Torriglia, Alicia; Jeanny, Jean-Claude; Behar-Cohen, Francine; Gurny, Robert; Möller, Michael

    2012-06-01

    Cornea transplantation is one of the most performed graft procedures worldwide with an impressive success rate of 90%. However, for "high-risk" patients with particular ocular diseases in addition to the required surgery, the success rate is drastically reduced to 50%. In these cases, cyclosporin A (CsA) is frequently used to prevent the cornea rejection by a systemic treatment with possible systemic side effects for the patients. To overcome these problems, it is a challenge to prepare well-tolerated topical CsA formulations. Normally high amounts of oils or surfactants are needed for the solubilization of the very hydrophobic CsA. Furthermore, it is in general difficult to obtain ocular therapeutic drug levels with topical instillations due to the corneal barriers that efficiently protect the intraocular structures from foreign substances thus also from drugs. The aim of this study was to investigate in vivo the effects of a novel CsA topical aqueous formulation. This formulation was based on nanosized polymeric micelles as drug carriers. An established rat model for the prevention of cornea graft rejection after a keratoplasty procedure was used. After instillation of the novel formulation with fluorescent labeled micelles, confocal analysis of flat-mounted corneas clearly showed that the nanosized carriers were able to penetrate into all corneal layers. The efficacy of a 0.5% CsA micelle formulation was tested and compared to a physiological saline solution and to a systemic administration of CsA. In our studies, the topical CsA treatment was carried out for 14 days, and the three parameters (a) cornea transparency, (b) edema, and (c) neovascularization were evaluated by clinical observation and scoring. Compared to the control group, the treated group showed a significant higher cornea transparency and significant lower edema after 7 and 13 days of the surgery. At the end point of the study, the neovascularization was reduced by 50% in the CsA-micelle treated

  6. Cyclosporine A inhibits colorectal cancer proliferation probably by regulating expression levels of c-Myc, p21(WAF1/CIP1) and proliferating cell nuclear antigen.

    PubMed

    Masuo, Takashige; Okamura, Shinichi; Zhang, Yajing; Mori, Masatomo

    2009-11-18

    The present study investigated the role of calcineurin (CaN) in the proliferation of human colorectal cancers. CaN activity and protein expression were increased in human colorectal cancers. Nuclear transcription factor NFAT, a physiological substrate for CaN, was activated in human colon cancer specimen as well as in the human colon cancer cell lines. CaN inhibitor cyclosporine A (CsA) reduced cell growth in these cell lines. CsA decreased the expressions of c-Myc and the proliferating cell nuclear antigen (PCNA) but also increased p21(WAF1/CIP1) expression. Our results suggest that CaN promotes colorectal cancer proliferation probably by regulating levels of c-Myc, p21(WAF1/CIP1), and PCNA.

  7. Renal and segmental pancreatic grafting with draining of exocrine secretion and initial continuous intravenous cyclosporin A in a patient with insulin-dependent diabetes and renal failure

    PubMed Central

    Calne, R Y; White, D J G; Rolles, K; Duffy, T J; Kass, T

    1982-01-01

    A patient with renal failure and insulin-dependent diabetes received renal and segmental pancreatic allografts from the same donor, with exocrine drainage of the pancreas being directed into the bowel. An attempt was made to maintain the serum concentrations of cyclosporin A between 300 and 1000 μg/l to avoid serious nephrotoxicity and rejection. Considerable difficulty was experienced in controlling the serum concentrations even with continuous intravenous infusion. When the concentrations were maintained between 300 and 1000 μg/l function in both allografts was satisfactory. At seven months the patient required no insulin and had good renal function. He was not receiving corticosteroids. ImagesFIG 1 PMID:6809184

  8. An in vivo microdialysis measurement of harpagoside in rat blood and bile for predicting hepatobiliary excretion and its interaction with cyclosporin A and verapamil.

    PubMed

    Wu, Qian; Wen, Xiao-Dong; Qi, Lian-Wen; Wang, Wei; Yi, Ling; Bi, Zhi-Ming; Li, Ping

    2009-03-15

    Harpagoside, a major bioactive iridoid glucoside in genus Scrophularia, has been widely used in clinical practice for the treatment of pain in the joints and lower back for its neuroprotective and anti-inflammation activities. To investigate the pharmacokinetics and hepatobiliary excretion, an in vivo microdialysis method coupled with high performance liquid chromatography was developed to monitor the concentration of harpagoside in blood and bile. The harpagoside bile-to-blood distribution ratio (AUC(bile)/AUC(blood)) up to 986.28+/-78.46 significantly decreased to 6.41+/-0.56 or 221.20+/-18.92 after co-administration of cyclosporin A or verapamil. The results indicated that harpagoside went through concentrative elimination from the bile which was probably regulated by P-glucoprotein, providing possible clinical trials of co-administration of transporter inhibitors to decrease drug efflux, thus to enhance the curative effects.

  9. The immunosuppressives FK 506 and cyclosporin A inhibit the generation of protein factors binding to the two purine boxes of the interleukin 2 enhancer.

    PubMed Central

    Brabletz, T; Pietrowski, I; Serfling, E

    1991-01-01

    Like Cyclosporin A (CsA), the macrolide FK 506 is a potent immunosuppressive that inhibits early steps of T cell activation, including the synthesis of Interleukin 2 (II-2) and numerous other lymphokines. The block of II-2 synthesis occurs at the transcriptional level. At concentrations that block T cell activation, FK 506 and CsA inhibit the proto-enhancer activity of Purine boxes of the II-2 promoter and the generation of lymphocyte-specific factors binding to the Purine boxes. Under the same conditions, the DNA binding of other II-2 enhancer factors remains unaffected by both compounds. These results support the view that FK 506 and CsA, which both inhibit the activity of peptidylprolyl cis/trans isomerases, suppress T cell activation by a similar, if not identical mechanism. Images PMID:1707162

  10. [Cyclosporin A: experience of the Renal Transplant Unit of the Clinical Hospital of the Medical College of the University of Sao Paulo].

    PubMed

    Ianhez, L E; Chocair, P R; Fonseca, J A; Azevedo, L S; de Paula, F J; David Neto, E; Romão Júnior, J E; Galvão, M M; de Castro, M C; Arap, S

    1991-01-01

    The authors report their experience using cyclosporine-A (CsA) in renal transplant patients. When compared with azathioprine/prednisone, CsA contributed significantly to a better graft and patient survival, either if used associated with prednisone of with azathioprine plus prednisone. CsA was also used in substitution to azathioprine in patients with hepatopathy attributed to azathioprine toxicity. The initial results are promising. The association of CsA and azathioprine with corticosteroids withdrawal was used as an attempt to allow normal growth in children. This seems to be the best choice of treatment for children. Careful monitoring of CsA blood levels avoids, or at least, minimizes nephrotoxicity. To achieve therapeutic CsA levels, patients with liver damage need lower, while children need higher oral CsA doses. To summarise: when CsA in carefully used, it is an excellent immunosuppressive drug.

  11. In vitro study of drug-eluting stent coatings based on poly(L-lactide) incorporating cyclosporine A - drug release, polymer degradation and mechanical integrity.

    PubMed

    Sternberg, Katrin; Kramer, Sven; Nischan, Claudia; Grabow, Niels; Langer, Thomas; Hennighausen, Gerhard; Schmitz, Klaus-Peter

    2007-07-01

    In this study, absorbable polymer stent coatings for localized drug delivery based on poly(L-lactide) (PLLA) and cyclosporine A (CsA) were developed and tested in vitro. Metallic stents were coated with different compositions of PLLA/CsA (70/30, 60/40, 50/50% w/w) and beta-sterilized. The specimens were used to assess the drug release kinetics with HPLC. Sterilization influenced polymer degradation was measured with GPC. Mechanical integrity of the stent coatings was studied with SEM. The interconnection of the coated stents with a balloon-catheter was characterized by the measurement of stent dislodgment force. A migration assay was used to determine the inhibitory effect of the model drug CsA on smooth muscle cell (SMC) migration. The release of CsA was established over time periods up to 24 days in sodium chloride solution and in porcine blood plasma. An inhibition of SMC migration (max. 26-33%) was found for CsA concentrations of 4 x 10(-5) to 4 x 10(-7) mol/l. Marked molecular weight reduction (70-80%) of the PLLA matrix occurred after beta-sterilization. We also observed a substantial decrease of in vitro degradation time. The maintenance of the mechanical integrity of the polymer coating during crimping and dilation of the specimens could be verified, and a sufficient stent dislodgment force of 0.8-0.9 N was measured.

  12. Stimulatory effects of Cuminum cyminum and flavonoid glycoside on Cyclosporine-A and restraint stress induced immune-suppression in Swiss albino mice.

    PubMed

    Chauhan, Prashant Singh; Satti, Naresh Kumar; Suri, Krishan Avtar; Amina, Musarat; Bani, Sarang

    2010-04-15

    Many herbs and spices are known to modulate the immune system and have been shown to restore the immunity in immuno-compromised individuals. Spices generally used to increase the taste and flavor of food also has the history of usage as an ayurvedic medicine. Therefore to explore the health modulating effects of Cuminum cyminum and to identify the active compound, immunomodulatory properties were evaluated using flowcytometry and ELISA in normal and immune-suppressed animals. C. cyminum and compound 1 stimulated the T cells and Th1 cytokines expression in normal animals. Swiss albino mice subjected to Cyclosporine-A induced immune-suppression were dosed orally with C. cyminum (25, 50, 100 and 200 mg/kg) on consecutive days. The results showed that administration significantly increased T cells (CD4 and CD8) count and Th1 predominant immune response in a dose dependent manner thereby suggesting immunomodulatory activity through modulation of T lymphocytes expression. In restraint stress induced immune-suppressed animals, compound 1 countered the depleted T lymphocytes, decreased the elevated corticosterone levels and size of adrenal glands and increased the weight of thymus and spleen. Based on the data we may conclude that C. cyminum is a potent immunomodulator and may develop as a lead to recover the immunity of immuno-compromised individuals.

  13. Sirolimus reduces the incidence and progression of UVB-induced skin cancer in SKH mice even with co-administration of cyclosporine A.

    PubMed

    Wulff, Brian C; Kusewitt, Donna F; VanBuskirk, Anne M; Thomas-Ahner, Jennifer M; Duncan, F Jason; Oberyszyn, Tatiana M

    2008-10-01

    Transplant immunosuppressants have been implicated in the increased incidence of non-melanoma skin cancer in transplant recipients, most of whom harbor considerable UVB-induced DNA damage in their skin prior to transplantation. This study was designed to evaluate the effects of two commonly used immunosuppressive drugs, cyclosporine A (CsA) and sirolimus (SRL), on the development and progression of UVB-induced non-melanoma skin cancer. SKH-1 hairless mice were exposed to UVB alone for 15 weeks, and then were treated with CsA, SRL, or CsA+SRL for 9 weeks following cessation of UVB treatment. Compared with vehicle, CsA treatment resulted in enhanced tumor size and progression. In contrast, mice treated with SRL or CsA+SRL had decreased tumor multiplicity, size, and progression compared with vehicle-treated mice. CsA, but not SRL or combined treatment, increased dermal mast cell numbers and TGF-beta1 levels in the skin. These findings demonstrate that specific immunosuppressive agents differentially alter the cutaneous tumor microenvironment, which in turn may contribute to enhanced development of UVB-induced skin cancer in transplant recipients. Furthermore, these results suggest that CsA alone causes enhanced growth and progression of skin cancer, whereas co-administration of SRL with CsA causes the opposite effect. JID JOURNAL CLUB ARTICLE: For questions, answers, and open discussion about this article please go to http://network.nature.com/group/jidclub

  14. The immunosuppressive agents rapamycin, cyclosporin A and tacrolimus increase lipolysis, inhibit lipid storage and alter expression of genes involved in lipid metabolism in human adipose tissue.

    PubMed

    Pereira, Maria J; Palming, Jenny; Rizell, Magnus; Aureliano, Manuel; Carvalho, Eugénia; Svensson, Maria K; Eriksson, Jan W

    2013-01-30

    Cyclosporin A (CsA), tacrolimus and rapamycin are immunosuppressive agents (IAs) associated with insulin resistance and dyslipidemia, although their molecular effects on lipid metabolism in adipose tissue are unknown. We explored IAs effects on lipolysis, lipid storage and expression of genes involved on lipid metabolism in isolated human adipocytes and/or adipose tissue obtained via subcutaneous and omental fat biopsies. CsA, tacrolimus and rapamycin increased isoproterenol-stimulated lipolysis and inhibited lipid storage by 20-35% and enhanced isoproterenol-stimulated hormone-sensitive lipase Ser552 phosphorylation. Rapamycin also increased basal lipolysis (~20%) and impaired insulin's antilipolytic effect. Rapamycin, down-regulated the gene expression of perilipin, sterol regulatory element-binding protein 1 (SREBP1) and lipin 1, while tacrolimus down-regulated CD36 and aP2 gene expression. All three IAs increased IL-6 gene expression and secretion, but not expression and secretion of TNF-α or adiponectin. These findings suggest that CsA, tacrolimus and rapamycin enhance lipolysis, inhibit lipid storage and expression of lipogenic genes in adipose tissue, which may contribute to the development of dyslipidemia and insulin resistance associated with immunosuppressive therapy.

  15. Reversion of resistance to immunosuppressive agents in three patients with psoriatic arthritis by cyclosporine A: modulation of P-glycoprotein function.

    PubMed

    Diamanti, Andrea Picchianti; Rosado, Manuela; Germano, Valentina; Scarsella, Marco; Giorda, Ezio; Podestà, Edoardo; D'Amelio, Raffaele; Carsetti, Rita; Laganà, Bruno

    2011-01-01

    Secondary resistance may be a major problem in the management of autoimmune diseases. P-glycoprotein (P-gp) over-function has been described as a mechanism of drug resistance in autoimmune patients. P-gp function can in vitro be inhibited by cyclosporine A (CSA) and verapamil; moreover, P-gp reduction by CSA in systemic lupus erythematosus and rheumatoid arthritis has been demonstrated. Here, P-gp function before and after CSA administration in three psoriatic arthritis (PsA) patients, who developed a resistance to MTX/SSA, has been evaluated. P-gp function on patient cells was analyzed by measuring the changes in rhodamine-123 (Rh-123) fluorescence after verapamil incubation. CSA treatment resulted in good clinical outcome that was related with a significant P-gp function reduction at CD3+ and CD8+ levels. In addition to its immunosuppressive activity, CSA results may also be related to MTX/SSA effect restoration through P-gp inhibition. This is the first time that CSA has been demonstrated as being able to revert MTX/SSA resistance in PsA.

  16. Nanoparticles made of multi-block copolymer of lactic acid and ethylene glycol containing periodic side-chain carboxyl groups for oral delivery of cyclosporine A

    PubMed Central

    Ankola, D. D.; Battisti, A.; Solaro, R.; Kumar, M. N. V. Ravi

    2010-01-01

    The purpose of this study was to evaluate the potential of new carboxylated multi-block copolymer of lactic acid and ethylene glycol (EL14) for nanoparticle (NP) formation and their ability to deliver high molecular weight hydrophobic drug—cyclosporine A (CsA). CsA-loaded EL14 NPs were compared with traditional poly(lactide-co-glycolide) (PLGA) NPs, both prepared by emulsion–diffusion–evaporation process. On the one hand, the increase in drug payload from 10 to 30 per cent for EL14 NPs showed no difference in particle size, however the entrapment efficiency tends to decrease from 50 to 43 per cent; on the other hand, the more hydrophobic PLGA showed an increasing trend in entrapment efficiency from 20 to 62 per cent with increasing particle size. Over 90 per cent of CsA was released in vitro from both the nanoparticulates; however, the release was much slower in the case of more hydrophobic PLGA. On in vivo evaluation in rats, the NPs made of EL14 showed a higher Cmax, a faster Tmax and enhanced tissue levels to that of PLGA that are crucial for CsA's activity and toxicity; however, the overall bioavailability of the nanoparticulates was similar and higher than Neoral. Together these data demonstrate the feasibility of NPs made of low molecular weight, hydrophilic polymer EL14 for efficient delivery of CsA. PMID:20504806

  17. Cyclosporin A inhibits nucleotide excision repair via downregulation of the xeroderma pigmentosum group A and G proteins, which is mediated by calcineurin inhibition.

    PubMed

    Kuschal, Christiane; Thoms, Kai-Martin; Boeckmann, Lars; Laspe, Petra; Apel, Antje; Schön, Michael P; Emmert, Steffen

    2011-10-01

    Cyclosporin A (CsA) inhibits nucleotide excision repair (NER) in human cells, a process that contributes to the skin cancer proneness in organ transplant patients. We investigated the mechanisms of CsA-induced NER reduction by assessing all xeroderma pigmentosum (XP) genes (XPA-XPG). Western blot analyses revealed that XPA and XPG protein expression was reduced in normal human GM00637 fibroblasts exposed to 0.1 and 0.5 μm CsA. Interestingly, the CsA treatment reduced XPG, but not XPA, mRNA expression. Calcineurin knockdown in GM00637 fibroblasts using RNAi led to similar results suggesting that calcineurin-dependent signalling is involved in XPA and XPG protein regulation. CsA-induced reduction in NER could be complemented by the overexpression of either XPA or XPG protein. Likewise, XPA-deficient fibroblasts with stable overexpression of XPA (XP2OS-pCAH19WS) did not show the inhibitory effect of CsA on NER. In contrast, XPC-deficient fibroblasts overexpressing XPC showed CsA-reduced NER. Our data indicate that the CsA-induced inhibition of NER is a result of downregulation of XPA and XPG protein in a calcineurin-dependent manner.

  18. Increased Advanced Oxidation Protein Products Generation by Cyclosporine-A and Angiotensin II in Human Gingival Fibroblasts – Ex-vivo Study

    PubMed Central

    Subbarayan, Rajasekaran; Ajitkumar, Supraja; Murugan Girija, Dinesh

    2017-01-01

    Introduction Cyclosporin-A (CsA), an immunosuppressant, induces renal fibrosis and Renin Angiotensin System (RAS) is known to play a major role. CsA has the potential to increase the oxidative stress; specifically through the Advanced Oxidation Protein Products (AOPP) which could possibly stimulate fibrosis. A similar type of pathology occurs even in the gingiva known as CsA Induced Gingival Overgrowth (CIGO). Aim This study was undertaken to estimate the AOPP generation by Human Gingival Fibroblasts (HGF) under the influence of CsA and Angiotensin II (Ang II). Materials and Methods Six healthy gingival tissue samples were obtained during crown lengthening procedure and primary HGF were cultured using enzymatic digestion method. The ideal non-cytotoxic concentrations of CsA and Ang II were identified using cytotoxicity assay. Later, HGF were incubated with CsA and Ang II for 12 hours and AOPP assay was performed at zero and one hour interval. Results There was a statistically significant increase in AOPP production in both the CsA and Ang II when compared to the control group with a p value<0.05. Conclusion CsA can induce oxidative stress and preventing/controlling it may be necessary to prevent untoward effect of the drug. PMID:28274044

  19. Cyclosporin A improves murine pregnancy outcome in abortion-prone matings: involvement of CD80/86 and CD28/CTLA-4.

    PubMed

    Zhou, Wen-Hui; Dong, Lin; Du, Mei-Rong; Zhu, Xiao-Yong; Li, Da-Jin

    2008-03-01

    Immune regulation during pregnancy is complex, and thus an optimal therapy for pregnancy complications is always a big challenge to reproductive medicine. Cyclosporin A (CsA), a potent immunosuppressant, prevents rejection of allografts by hosts, but little is known about the modulating effect of CsA on the materno-fetal relationship. Here, pregnant CBA/J females mated with DBA/2 males as an abortion-prone model were administered with CsA on day 4.5 of gestation, and the pregnant CBA/J females mated with BALB/c males were established as successful pregnancy control. It was demonstrated that administration of CsA at the window of implantation significantly up-regulated the expression of CTLA-4, while down-regulating the levels of CD80, CD86, and CD28 at the materno-fetal interface in the CBA/J x DBA/2 abortion-prone matings, and the embryo resorption rate of the abortion-prone matings reduced significantly after CsA treatment, implying that modulation of costimulatory molecule expression by CsA might contribute to preventing the fetus from maternal immune attack. In addition, treatment with CsA induced enhanced growth and reduced cell apoptosis of the murine trophoblast cells. Together, these findings indicate that CsA has a beneficial effect on the materno-fetal interface in abortion-prone matings, leading to a pregnancy outcome improvement, which might provide new therapeutics for spontaneous pregnancy wastage.

  20. Antithymocyte globulin combined with cyclosporine A down-regulates T helper 1 cells by modulating T cell immune response cDNA 7 in aplastic anemia.

    PubMed

    Zhu, Feng; Qiao, Jianlin; Zhong, Xiao-min; Wu, Qing-yun; Chen, Wei; Yao, Yao; Niu, Ming-shan; Fu, Chun-ling; Zeng, Ling-yu; Li, Zhen-yu; Xu, Kai-lin

    2015-07-01

    Antithymocyte globulin (ATG) combined with cyclosporine A (CsA) has been widely used as a standard regimen in the treatment of aplastic anemia (AA), especially in severe aplastic anemia (SAA). Abnormally activated T cells might be the immune pathogenesis of AA. T cell immune response cDNA 7 (TIRC7) has been demonstrated its essential role in T cell activation; however, little is known about the role of TIRC7 in AA. In this study, we documented that TIRC7 levels in CsA group were higher than that in ATG + CsA (AC) group only in the follow-up phase (P < 0.05; P < 0.05); nevertheless, TIRC7 levels in SAA group were elevated than non severe aplastic anemia group not only in the treatment phase (P < 0.05; P < 0.05) but also in the follow-up phase (P < 0.05; P < 0.01). The trend of changes of T helper (Th) 1, Th17 and Th22 levels before and after treatment was similar to the changes of TIRC7 levels in either AC group or CsA group. Thus, TIRC7 might be involved in the pathogenesis of AA and AC might down-regulate Th1 cells by modulating the expression of TIRC7 in AA.

  1. Identification of the immunophilins capable of mediating inhibition of signal transduction by cyclosporin A and FK506: roles of calcineurin binding and cellular location.

    PubMed Central

    Bram, R J; Hung, D T; Martin, P K; Schreiber, S L; Crabtree, G R

    1993-01-01

    The immunosuppressants cyclosporin A (CsA) and FK506 appear to block T-cell function by inhibiting the calcium-regulated phosphatase calcineurin. While multiple distinct intracellular receptors for these drugs (cyclophilins and FKBPs, collectively immunophilins) have been characterized, the functionally active ones have not been discerned. We found that overexpression of cyclophilin A or B or FKBP12 increased T-cell sensitivity to CsA or FK506, respectively, demonstrating that they are able to mediate the inhibitory effects of their respective immunosuppressants in vivo. In contrast, cyclophilin C, FKBP13, and FKBP25 had no effect. Direct comparison of the Ki of each drug-immunophilin complex for calcineurin in vitro revealed that although calcineurin binding was clearly necessary, it was not sufficient to explain the in vivo activity of the immunophilin. Subcellular localization was shown also to play a role, since gene deletions of cyclophilins B and C which changed their intracellular locations altered their activities significantly. Cyclophilin B has been shown previously to be located within calcium-containing intracellular vesicles; its ability to mediate CsA inhibition implies that certain components of the signal transduction machinery are also spatially restricted within the cell. Images PMID:7687744

  2. Cyclosporin A suppresses the expression of the interleukin 2 gene by inhibiting the binding of lymphocyte-specific factors to the IL-2 enhancer.

    PubMed Central

    Randak, C; Brabletz, T; Hergenröther, M; Sobotta, I; Serfling, E

    1990-01-01

    Cyclosporin A (CsA), a powerful immunosuppressive drug, inhibits the synthesis of lymphokines in T lymphocytes at the level of gene transcription. Using protein extracts from El4 lymphoma cells we show that the binding of lymphocyte-specific factors interacting with the two so-called purine boxes (Pu-boxes) of the interleukin 2 (IL-2) enhancer are missing in CsA-treated cells. The CsA-sensitive factors are newly synthesized upon induction. The most prominent factor consists of 45 kd polypeptides and contacts both Pu-boxes at the two central G residues within the identical core sequence AAGAGGAAAA. The CsA-mediated suppression of factor binding to the Pu-boxes correlates well with functional studies in which the inducible, T cell-restricted proto-enhancer activity of Pu-boxes was selectively repressed by CsA. These observations support the conclusion that the suppression of factor binding to the Pu-boxes by CsA impairs the activity of IL-2 and of further lymphokine genes, thereby inhibiting the synthesis of lymphokines in T lymphocytes. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 5. Fig. 6. PMID:2369902

  3. Cyclosporin A suppresses the expression of the interleukin 2 gene by inhibiting the binding of lymphocyte-specific factors to the IL-2 enhancer.

    PubMed

    Randak, C; Brabletz, T; Hergenröther, M; Sobotta, I; Serfling, E

    1990-08-01

    Cyclosporin A (CsA), a powerful immunosuppressive drug, inhibits the synthesis of lymphokines in T lymphocytes at the level of gene transcription. Using protein extracts from El4 lymphoma cells we show that the binding of lymphocyte-specific factors interacting with the two so-called purine boxes (Pu-boxes) of the interleukin 2 (IL-2) enhancer are missing in CsA-treated cells. The CsA-sensitive factors are newly synthesized upon induction. The most prominent factor consists of 45 kd polypeptides and contacts both Pu-boxes at the two central G residues within the identical core sequence AAGAGGAAAA. The CsA-mediated suppression of factor binding to the Pu-boxes correlates well with functional studies in which the inducible, T cell-restricted proto-enhancer activity of Pu-boxes was selectively repressed by CsA. These observations support the conclusion that the suppression of factor binding to the Pu-boxes by CsA impairs the activity of IL-2 and of further lymphokine genes, thereby inhibiting the synthesis of lymphokines in T lymphocytes.

  4. T cell activation by concanavalin A in the presence of cyclosporin A: immunosuppressor withdrawal induces NFATp translocation and interleukin-2 gene transcription.

    PubMed

    Bemer, V; Truffa-Bachi, P

    1996-07-01

    Cyclosporin A (CSA), an immunosuppressive agent used in organ transplantation and to treat some autoimmune diseases, blocks the Ca2+-dependent steps involved in T cell receptor triggering leading to interleukin (IL)-2 production. Considering that the early steps of T cell activation are insensitive to CSA, we asked whether the initial activation achieved in presence of this immunosuppressor could affect the capacity of the T cell to respond to a mitogenic restimulation. We found that T cells activated by concanavalin A (ConA) for 48 h in the presence of CSA retain the capacity to proliferate in response to ConA once the immunosuppressor is removed. These cells are able to transcribe anew the IL-2 gene, without the requirement of new protein synthesis, and to up-regulate the alpha chain of the IL-2 receptor. Furthermore, we present the first direct evidence that the nuclear factor AP-1 is present in the nucleus of the T cells primed for 48 h in presence of CSA and that withdrawal of the immunosuppressor leads to the translocation of NFATp from the cytoplasm to the nucleus.

  5. A randomized trial of basiliximab with three different patterns of cyclosporin A initiation in renal transplant from expanded criteria donors and at high risk of delayed graft function.

    PubMed

    Andrés, Amado; Marcén, Roberto; Valdés, Francisco; Plumed, Jaime Sánchez; Solà, Ricard; Errasti, Pedro; Lauzurica, Ricardo; Pallardó, Luis; Bustamante, Jesús; Amenábar, Juan José; Plaza, Juan José; Gómez, Ernesto; Grinyó, Josep Maria; Rengel, Manuel; Puig, Josep Maria; Sanz, Aurelio; Asensio, Concepción; Andrés, Inés

    2009-01-01

    This study assays therapy with basiliximab and different patterns of cyclosporin A (CsA) initiation in renal transplant (RT) recipients from expanded criteria donors (ECD) and at high risk of delayed graft function (DGF). A multicentre six-month open-label randomized trial with three parallel groups treated with basiliximab plus steroids, mycophenolate mofetil and different patterns of CsA initiation: early within 24 h post-RT at 3 mg/kg/d (Group 1; n = 38), and at 5 mg/kg/d (Group 2; n = 40), or delayed after 7-10 d at 5 mg/kg/d (Group 3; n = 36). There were no differences among groups in six months GFR (43.1 +/- 12, 48.0 +/- 14 and 47.2 +/- 17 mL/min, respectively), DGF (Group 1: 31%, Group 2: 37%, Group 3: 42%), nor biopsy-proven acute rejection, although clinically treated and biopsy-proven acute rejection was significantly higher in Group 3 (25%) vs. Group 1 (5.3%, p < 0.05). At six months no differences were observed in death-censored graft survival or patient survival. Induction therapy with basiliximab and three CsA-ME initiation patterns in RT recipients from ECD and at high risk of DGF presented good renal function and graft survival at six months. Late onset group did not achieve improvement in DGF rate and showed a higher incidence of clinically treated and biopsy-proven acute rejection.

  6. Brugia pahangi in nude mice: protective immunity to infective larvae is Thy 1.2+ cell dependent and cyclosporin A resistant.

    PubMed

    Vickery, A C; Nayar, J K

    1987-03-01

    Mechanisms of protective immunity to larvae of Brugia pahangi were studied in congenitally athymic nude C3H/HeN mice and their syngeneic heterozygous littermates. An average 11% of subcutaneous larval inocula was recovered from control nudes 28 days after inoculation. No worms were recovered from nude recipients of viable splenic Thy 1.2+ T lymphocytes from heterozygotes which had killed a priming dose of B. pahangi larvae. Primed T lymphocytes, depleted of either Lyt 1.1+ or Lyt 2.1+ cells or incubated with anti-Thy 1.2 monoclonal antibody and complement, failed to protect nude mice against a larval challenge. Nor were primed B lymphocytes depleted by Thy 1.2+ T cell contaminants protective. Treatment with cyclosporin A (CsA) did not increase the numbers of worms recovered from heterozygotes nor did CsA treatment of heterozygous cell donors abolish the ability of primed Thy 1.2+ T lymphocytes to transfer protection to nude mice. IgG but not IgM antibody titres to B. pahangi antigens were depressed in all CsA-treated mice. CsA treatment of nude mice had no direct effect upon development of B. pahangi larvae. These results show that protective immunity to larvae of B. pahangi in mice depends upon small numbers of Thy 1.2+ T cells which are CsA-resistant.

  7. Cyclosporin-A inhibits constitutive nitric oxide synthase activity and neuronal and endothelial nitric oxide synthase expressions after spinal cord injury in rats.

    PubMed

    Diaz-Ruiz, Araceli; Vergara, Paula; Perez-Severiano, Francisca; Segovia, Jose; Guizar-Sahagún, Gabriel; Ibarra, Antonio; Ríos, Camilo

    2005-02-01

    Nitric oxide (NO) plays a role in the pathophysiology of spinal cord injury (SCI). NO is produced by three types of nitric oxide synthase (NOS) enzymes: The constitutive Ca2+/calmodulin-dependent neuronal NOS (nNOS) and endothelial NOS (eNOS) isoforms, and the inducible calcium-independent isoform (iNOS). During the early stages of SCI, nNOS and eNOS produce significant amounts of NO, therefore, the regulation of their activity and expression may participate in the damage after SCI. In the present study, we used Cyclosporin-A (CsA) to further substantiate the role of Ca-dependent NOS in neural responses associated to SCI. Female Wistar rats were subjected to SCI by contusion, and killed 4 h after lesion. Results showed an increase in the activity of constitutive NOS (cNOS) after lesion, inhibited by CsA (2.5 mg/kg i.p.). Western blot assays showed an increased expression of both nNOS and eNOS after trauma, also antagonized by CsA administration.

  8. Analysis of cyclosporin A and a set of analogs as inhibitors of a T. cruzi cyclophilin by docking and molecular dynamics.

    PubMed

    Carraro, Roberto; Iribarne, Federico; Paulino, Margot

    2016-01-01

    Cyclophilins (CyPs) are enzymes involved in protein folding. In Trypanosoma cruzi (T. cruzi), the most abundantly expressed CyP is the isoform TcCyP19. It has been shown that TcCyP19 is inhibited by the immunosuppressive drug cyclosporin A (CsA) and analogs, which also proved to have potent trypanosomicidal activity in vitro. In this work, we continue and expand a previous study on the molecular interactions of CsA, and a set of analogs modeled in complexes with TcCyP19. The modeled complexes were used to evaluate binding free energies by molecular dynamics (MD), applying the Linear Interaction Energy (LIE) method. In addition, putative binding sites were identified by molecular docking. In our analysis, the binding free energy calculations did not correlate with experimental data. The heterogeneity of the non-bonded energies and the variation in the pattern of hydrogen bonds suggest that the systems may not be suitable for the application of the LIE method. Further, the docking calculations identified two other putative binding sites with comparable scoring energies to the active site, a fact that may also explain the lack of correlation found. Kinetic experiments are needed to confirm or reject the multiple binding sites hypothesis. In the meantime, MD simulations at the alternative sites, employing other methods to compute binding free energies, might be successful at finding good correlations with the experimental data.

  9. Increased incidence of acute graft-versus-host disease with the continuous infusion of cyclosporine A compared to twice-daily infusion.

    PubMed

    Ogawa, N; Kanda, Y; Matsubara, M; Asano, Y; Nakagawa, M; Sakata-Yanagimoto, M; Kandabashi, K; Izutsu, K; Imai, Y; Hangaishi, A; Kurokawa, M; Tsujino, S; Ogawa, S; Aoki, K; Chiba, S; Motokura, T; Hirai, H

    2004-03-01

    We retrospectively compared the incidence of acute graft-versus-host disease (GVHD) before and after September 1999, when we changed the mode of cyclosporine A (CsA) administration from twice-daily infusions (TD) (n=58) to continuous infusion (CIF) (n=71). The incidence of grade II-IV acute GVHD in the CIF group (56%) was significantly higher than that in the TD group (27%, P=0.00022). Multivariate analysis identified only two independent significant risk factors for the development of grade II-IV acute GVHD; CIF of CsA (relative risk 2.59, 95% CI 1.46-4.60, P=0.0011) and the presence of HLA mismatch (2.01, 95% CI 1.15-3.53, P=0.014). The incidence of relapse was significantly lower in the CIF group when adjusted for disease status before transplantation (0.41, 95% CI 0.18-0.95, P=0.038), which resulted in better disease-free survival in high-risk patients (43 vs 16% at 2 years, P=0.039), but not in standard-risk patients (72 vs 80%, P=0.45). CIF of CsA with a target level of 250-400 ng/ml may not be appropriate for GVHD prophylaxis in standard-risk patients.

  10. Cyclosporine A enhances Th2 bias at the maternal-fetal interface in early human pregnancy with aid of the interaction between maternal and fetal cells.

    PubMed

    Piao, Hai-Lan; Wang, Song-Cun; Tao, Yu; Zhu, Rui; Sun, Chan; Fu, Qiang; Du, Mei-Rong; Li, Da-Jin

    2012-01-01

    Our previous study has demonstrated that cyclosporine A (CsA) administration in vivo induces Th2 bias at the maternal-fetal interface, leading to improved murine pregnancy outcomes. Here, we investigated how CsA treatment in vitro induced Th2 bias at the human maternal-fetal interface in early pregnancy. The cell co-culture in vitro in different combination of component cells at the maternal-fetal interface was established to investigate the regulation of CsA on cytokine production from the interaction of these cells. It was found that interferon (IFN)-γ was produced only by decidual immune cells (DICs), and not by trophoblasts or decidual stromal cells (DSCs); all these cells secreted interleukin (IL)-4, IL-10, and tumor necrosis factor (TNF)-α. Treatment with CsA completely blocked IFN-γ production in DICs and inhibited TNF-α production in all examined cells. CsA increased IL-10 and IL-4 production in trophoblasts co-cultured with DSCs and DICs although CsA treatment did not affect IL-10 or IL-4 production in any of the cells when cultured alone. These results suggest that CsA promotes Th2 bias at the maternal-fetal interface by increasing Th2-type cytokine production in trophoblasts with the aid of DSCs and DICs, while inhibiting Th1-type cytokine production in DICs and TNF-α production in all investigated cells. Our study might be useful in clinical therapeutics for spontaneous pregnancy wastage and other pregnancy complications.

  11. Efficacy of a new topical cationic emulsion of cyclosporine A on dry eye clinical signs in an experimental mouse model of dry eye.

    PubMed

    Daull, Philippe; Feraille, Laurence; Barabino, Stefano; Cimbolini, Nicolas; Antonelli, Sophie; Mauro, Virgine; Garrigue, Jean-Sébastien

    2016-12-01

    Dry eye disease (DED) is a complex, multifactorial pathology characterized by corneal epithelium lesions and inflammation. The aim of the present study was to evaluate the efficacy of a cationic emulsion of cyclosporine A (CsA) in a mouse model that mimics severe dry eye. Eight to 12-week-old female C57BL/6N mice with tail patches of scopolamine were housed in controlled environment chambers to induce dry eye. At day three, following dry eye confirmation by corneal fluorescein staining (CFS, score 0-15) and phenol red thread (PRT) lacrimation test, the mice (n = 10/gp) were either treated 3 times a day in both eyes with drug-free cationic emulsion, a 0.1% CsA cationic emulsion, or 1% methylprednisolone (positive control), or non-treated. Aqueous tear production and CFS scores were evaluated at baseline and throughout the treatment period. The lacrimation test confirmed the scopolamine-induced decrease in aqueous production by the lacrimal gland. A reduction of 59% in induced-CFS was observed following topical treatment with 0.1% CsA. The beneficial effect of the cationic emulsion vehicle itself on keratitis was also clearly evidenced by its better performance over 1% methylprednisolone, -36%, vs. -28% on the CFS scores, respectively. This study indicates that the cationic emulsion of CsA (0.1%) was a very effective formulation for the management of corneal epithelium lesions in a severe DED mouse model. In addition, it performed better than a potent glucocorticosteroid (1% methylprednisolone). This cationic emulsion of CsA (0.1%), combining CsA and a tear film oriented therapy (TFOT), i.e. with vehicle properties that mechanically stabilize the tear film, represents a promising new treatment strategy for the management of the signs of dry eye.

  12. Cyclodextrins and chitosan derivatives in sublingual delivery of low solubility peptides: A study using cyclosporin A, alpha-cyclodextrin and quaternary chitosan N-betainate.

    PubMed

    Mannila, Janne; Järvinen, Kristiina; Holappa, Jukka; Matilainen, Laura; Auriola, Seppo; Jarho, Pekka

    2009-10-20

    Systemic drug delivery through intraoral membranes may offer a promising administration route for lipophilic peptide drugs. The aim of the present study was to investigate the effect of alpha-cyclodextrin (alpha-CD) and a novel chitosan derivative, chitosan N-betainate (CH), on sublingual absorption of a hydrophobic model peptide cyclosporin A (CsA), and the effect of temperature on the complexation of CsA with alpha-CD. Complexation of CsA with alpha-CD was studied using the phase-solubility method. Sublingual absorption of CsA was studied by administration of solid CsA/alpha-CD complex (with and without CH solution), solid CsA/alpha-CD/CH formulation and solid plain CsA to rabbits. The solubility of CsA in aqueous alpha-CD solution (14%) increased with decreasing temperature; the solubility of CsA at room temperature, +5 and +1 degrees C was 1.2, 12 and 19mg/ml, respectively. The bioavailability of CsA after administration of plain CsA, solid CsA/alpha-CD and solid CsA/alpha-CD/CH (0.6+/-0.5, 1.4+/-0.7 and 1.7+/-0.8%, respectively; mean+/-S.D.) was further increased when solid CsA/alpha-CD was administered together with CH solution (3.2+/-2.2%). The present study shows that decreased temperature can be effectively utilized to produce CsA/alpha-CD complexes. It was also shown that alpha-CD and CH may be advantageous in sublingual delivery of lipophilic peptides, although the absolute bioavailability remains low.

  13. The Tumorigenicity of Multipotent Adult Germline Stem Cells Transplanted into the Heart Is Affected by Natural Killer Cells and by Cyclosporine A Independent of Its Immunosuppressive Effects

    PubMed Central

    Hübscher, Daniela; Kaiser, Diana; Elsner, Leslie; Monecke, Sebastian; Dressel, Ralf; Guan, Kaomei

    2017-01-01

    Transplantation of stem cells represents an upcoming therapy for many degenerative diseases. For clinical use, transplantation of pluripotent stem cell-derived cells should lead to integration of functional grafts without immune rejection or teratoma formation. Our previous studies showed that the risk of teratoma formation is highly influenced by the immune system of the recipients. In this study, we have observed a higher teratoma formation rate when undifferentiated so-called multipotent adult germline stem cells (maGSCs) were transplanted into the heart of T, B, and natural killer (NK) cell-deficient RAG2−/−γc−/− mice than in RAG2−/− mice, which still have NK cells. Notably, in both strains, the teratoma formation rate was significantly reduced by the immunosuppressive drug cyclosporine A (CsA). Thus, CsA had a profound effect on teratoma formation independent of its immunosuppressive effects. The transplantation into RAG2−/− mice led to an activation of NK cells, which reached the maximum 14 days after transplantation and was not affected by CsA. The in vivo-activated NK cells efficiently killed YAC-1 and also maGSC target cells. This NK cell activation was confirmed in C57BL/6 wild-type mice whether treated with CsA or not. Sham operations in wild-type mice indicated that the inflammatory response to open heart surgery rather than the transplantation of maGSCs activated the NK cell system. An activation of NK cells during the transplantation of stem cell-derived in vitro differentiated grafts might be clinically beneficial by reducing the risk of teratoma formation by residual pluripotent cells. PMID:28220117

  14. Tumour necrosis factor-α expression and cell recruitment in Sephadex particle-induced lung inflammation: effects of dexamethasone and cyclosporin A

    PubMed Central

    Williams, Cara M M; Smith, Lance; Flanagan, Brian F; Steve Clegg, L; Coleman, John W

    1997-01-01

    Tumour necrosis factor-α (TNF-α) is a cytokine with diverse properties consistent with a possible role in inflammatory disease. We investigated whether TNF-α is induced during the progression of lung inflammation elicited by a particulate non-antigenic stimulus, and whether pharmacological control of TNF-α expression influences recruitment of specific inflammatory cell types. A single intravenous injection of Sephadex particles into rats led to extensive granulomatous inflammation in lung alveolar and bronchial tissue that peaked in intensity after 24–72 h. Mononuclear cells were the principal component of granulomas, but neutrophils and eosinophils were also abundant. Numbers of mononuclear cells, neutrophils and eosinophils recovered by bronchoalveolar lavage (BAL) peaked at 72 h, 48 h and 72 h, respectively. Messenger RNA encoding TNF-α was induced in lung epithelial cells, lung granulomas and BAL cells 6 h after Sephadex administration and remained elevated for 72 h before declining to baseline by 7 days. In BAL cell populations TNF-α protein was localized to mononuclear cells at all times points pre- and post-Sephadex administration. Treatment of rats with dexamethasone significantly reduced the Sephadex-induced recruitment of mononuclear cells, neutrophils and eosinophils into the bronchoalveolar cavity, and significantly reduced TNF-α mRNA expression by BAL cells. Treatment of rats with cyclosporin A was without effect on Sephadex-induced elevations of mononuclear cell numbers and expression of TNF-α, but did reduce significantly recruitment of neutrophils and eosinophils to BAL cell populations. These results show that a sequential asthma-like recruitment of neutrophils, eosinophils and mononuclear cells into lung tissue can be induced by single exposure to a non-antigenic stimulus. Pharmacological and histological studies reveal that mononuclear cell mobilization relates closely to induced TNF-α expression, whereas mobilization of

  15. Do Cyclosporine A, an IL-1 Receptor Antagonist, Uridine Triphosphate, Rebamipide, and/or Bimatoprost Regulate Human Meibomian Gland Epithelial Cells?

    PubMed Central

    Kam, Wendy R.; Liu, Yang; Ding, Juan; Sullivan, David A.

    2016-01-01

    Purpose Researchers have hypothesized that treatment with cyclosporine A (CyA), interleukin-1 receptor antagonists (IL-1RA; e.g., anakinra), P2Y2 receptor agonists (e.g., uridine triphosphate; UTP), and rebamipide may alleviate human meibomian gland dysfunction (MGD) and/or dry eye disease. Investigators have also proposed that prostaglandin analogues (e.g., bimatoprost) may induce MGD. Our goal was to determine whether these compounds directly influence human meibomian gland epithelial cell (HMGEC) function. Methods Multiple concentrations of each compound were tested for effects on immortalized (I) HMGEC morphology and survival. Nontoxic dosages were used for our studies. Immortalized HMGEC were cultured in the presence of vehicle, CyA, IL-1RA, UTP, rebamipide, or bimatoprost for up to 6 days in various media. Experiments included positive controls for proliferation (epidermal growth factor and bovine pituitary extract), differentiation (azithromycin), and signaling pathway activation (insulin-like growth factor 1). Cells were analyzed for neutral lipid staining, lysosome accumulation, lipid composition, and phosphatidylinositol-3-kinase/Akt (AKT), phosphorylation. Results Our findings demonstrate that CyA, IL-1RA, UTP, rebamipide, and bimatoprost had no effect on the proliferation; neutral lipid content; lysosome number; or levels of free cholesterol, triglycerides, or phospholipids in IHMGECs. Cylosporine A, IL-1RA, rebamipide, and bimatoprost significantly reduced the phosphorylation of AKT, as compared to control. Of interest, tested doses of CyA above 8 nM killed the IHMGECs. Conclusions Our results show that CyA, IL-1RA, UTP, rebamipide, and bimatoprost do not influence the proliferation or differentiation of IHMGEC. However, with the exception of UTP, these compounds do decrease the activity of the AKT signaling pathway, which is known to promote cell survival. PMID:27552406

  16. Inhibition of programmed cell death by cyclosporin A; preferential blocking of cell death induced by signals via TCR/CD3 complex and its mode of action.

    PubMed Central

    Yasutomi, D; Odaka, C; Saito, S; Niizeki, H; Kizaki, H; Tadakuma, T

    1992-01-01

    Cyclosporin A (CsA) is reported to inhibit programmed cell death. We confirmed this by using T-cell hybridomas which are inducible to programmed cell death by activation with immobilized anti-CD3 antibody or with anti-Thy 1.2 antibody. Cell death and DNA fragmentation, characteristic features of programmed cell death, were almost completely blocked by CsA or FK506. To investigate whether CsA inhibits only the cell death through the signals via the TCR/CD3 complex or all of the programmed cell death induced by various reagents, we further established CD4+8+ thymic lymphomas which result in programmed cell death after activation with calcium ionophore, dexamethasone, cyclic AMP or anti-CD3 antibody. It was revealed that CsA could block only the cell death mediated by the TCR/CD3 complex. For the clarification of the site of action of CsA, Ca2+ influx and endocytosis of receptors after stimulation with anti-CD3 antibody were monitored in the presence of CsA, and no significant effects of CsA were observed. Furthermore, prevention of cell death was examined by adding CsA at various periods of time after initiation of culture. CsA was found to exert its effect even when added after 4 h of cultivation, and the kinetic pattern of suppression was similar to that of the suppressive effect on IL-2 production. These observations indicate that in the events of programmed cell death, the major site of action of CsA will not be the inhibition of the immediate membrane events after activation of the TCR/CD3 complex but rather the interference in the function of molecules that transmit signals between membrane events and the activation of genes in the nucleus. Images Figure 2 Figure 3 PMID:1383138

  17. Effects of Gallic Acid and Cyclosporine A on Antioxidant Capacity and Cardiac Markers of Rat Isolated Heart After Ischemia/Reperfusion

    PubMed Central

    Badavi, Mohammad; Sadeghi, Najmeh; Dianat, Mahin; Samarbafzadeh, Alireza

    2014-01-01

    Background: Myocardial infarction is one of the important causes of death during old ages. Gallic acid as an antioxidant or cyclosporine A (CsA) as inhibitor of mitochondrial permeability transition pore (mPTP) alone could prevent these complications to some extent, but their combination effect has not been investigated. Objectives: The aim of this study was to determine the combined effect of gallic acid and CsA on antioxidant capacity of isolated heart tissues during ischemia reperfusion. Materials and Methods: Eighty male Wistar rats were randomly assigned to different groups: sham, control (Ca, received saline, 1 mL/kg); 3 groups were pretreated with gallic acid (G1a: 7.5, G2a: 15, G3a: 30 mg/kg) for 10 days, and the other 3 groups were pretreated with gallic acid and received CsA (0.2 µM) for 10 minutes before induction of ischemia and during the first 10 minutes of reperfusion (G1b, G2b and G3b) and the last group received CsA alone (Cb). After 10 days of pretreatment, the heart was isolated and transferred to the Langendorff apparatus and exposed to 30 minutes ischemia followed by 60 minutes of reperfusion. After that cardiac markers and antioxidant enzymes were assessed in cardiac tissues. Results: Lactate dehydrogenase (LDH), Superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) activity increased and malondialdehyde (MDA) decreased in animals pretreated with gallic acid significantly. However, pretreatment with gallic acid followed by CsA during reperfusion improved the antioxidant capacity and cardiac marker enzymes and restored the lipid peroxidation more effective than gallic acid or CsA alone. Nevertheless, CsA did not change the cardiac marker enzymes significantly. Conclusions: Gallic acid and CsA combination improved antioxidant capacity and cell membrane integrity more than each one alone. Therefore, it can be a therapeutic approach to reduce the I/R injury. PMID:25068044

  18. Improved oral bioavailability of cyclosporin A in male Wistar rats. Comparison of a Solutol HS 15 containing self-dispersing formulation and a microsuspension.

    PubMed

    Bravo González, Roberto Carlos; Huwyler, Jörg; Walter, Isabelle; Mountfield, Richard; Bittner, Beate

    2002-10-01

    Oral bioavailability of the highly lipophilic and poorly water-soluble immunosuppressive agent cyclosporin A (CyA) in two different formulations was investigated in male Wistar rats. An aqueous microsuspension and a self-dispersing formulation composed of the surface-active ingredients Solutol HS 15:Labrafil M2125CS:oleic acid=7:2:1 (v/v/v) were administered to the animals at a dose level of 20 mg/kg. In order to calculate the absolute oral bioavailability, CyA was additionally administered intravenously at 10 mg/kg as microsuspension. It was found that the oral bioavailability of CyA in the Solutol HS 15-based formulation was twofold higher as compared to the microsuspension (69.9+/-2.8 vs. 35.7+/-3.3%, P=0.001). By contrast, the time to reach maximum plasma concentration (t(max)) and the terminal half-life (t(1/2)) did not differ significantly with the different formulations (t(max): 7.0+/-1.0 vs. 6.3+/-1.7 h; t(1/2): 20.5+/-2.9 vs. 16.7+/-4.7 h). In vitro solubility experiments demonstrated a marked increase in the aqueous solubility of CyA in the presence of the self-dispersing formulation as compared to the micronized powder alone (solubility after 120 min at 37 degrees C: 136 vs. 23.2 microg/ml in human gastric juice; 133 vs. 10.8 microg/ml in simulated intestinal juice). Most likely, the enhanced systemic exposure of CyA in the self-dispersing formulation was caused by improved solubility of CyA in the gastrointestinal fluids in the presence of the surface-active ingredients. Additional factors that may have contributed to increased oral bioavailability are inhibition of metabolism and/or transport processes as well as permeability enhancement by the co-administered excipients.

  19. Characterization and stability studies of a novel liposomal cyclosporin A prepared using the supercritical fluid method: comparison with the modified conventional Bangham method

    PubMed Central

    Karn, Pankaj Ranjan; Cho, Wonkyung; Park, Hee-Jun; Park, Jeong-Sook; Hwang, Sung-Joo

    2013-01-01

    A novel method to prepare cyclosporin A encapsulated liposomes was introduced using supercritical fluid of carbon dioxide (SCF-CO2) as an antisolvent. To investigate the strength of the newly developed SCF-CO2 method compared with the modified conventional Bangham method, particle size, zeta potential, and polydispersity index (PDI) of both liposomal formulations were characterized and compared. In addition, entrapment efficiency (EE) and drug loading (DL) characteristics were analyzed by reversed-phase high-performance liquid chromatography. Significantly larger particle size and PDI were revealed from the conventional method, while EE (%) and DL (%) did not exhibit any significant differences. The SCF-CO2 liposomes were found to be relatively smaller, multilamellar, and spherical with a smoother surface as determined by transmission electron microscopy. SCF-CO2 liposomes showed no significant differences in their particle size and PDI after more than 3 months, whereas conventional liposomes exhibited significant changes in their particle size. The initial yield (%), EE (%), and DL (%) of SCF-CO2 liposomes and conventional liposomes were 90.98 ± 2.94, 92.20 ± 1.36, 20.99 ± 0.84 and 90.72 ± 2.83, 90.24 ± 1.37, 20.47 ± 0.94, respectively, which changed after 14 weeks to 86.65 ± 0.30, 87.63 ± 0.72, 18.98 ± 0.22 and 75.04 ± 8.80, 84.59 ± 5.13, 15.94 ± 2.80, respectively. Therefore, the newly developed SCF-CO2 method could be a better alternative compared with the conventional method and may provide a promising approach for large-scale production of liposomes. PMID:23378759

  20. The Immunophilin Ligands Cyclosporin A and FK506 Suppress Prostate Cancer Cell Growth by Androgen Receptor-Dependent and -Independent Mechanisms

    PubMed Central

    PERIYASAMY, SUMUDRA; WARRIER, MANYA; TILLEKERATNE, MANORANJANI P. M.; SHOU, WEINIAN; SANCHEZ, EDWIN R.

    2008-01-01

    Androgen receptor (AR) contributes to growth of prostate cancer even under conditions of androgen ablation. Thus, new strategies to target AR activity are needed. AR interacts with the immunophilin FK506-binding protein 52 (FKBP52), and studies in the FKBP52 KO mouse have shown this protein is essential to AR activity in the prostate. We therefore tested whether the immunophilin ligand FK506 affected AR activity in prostate cancer cell lines. We also tested the hypothesis that AR interacts with another immunophilin Cyp40 and is regulated by its cognate ligand cyclosporin A (CsA). We show that levels of FKBP52, FKBP51, Cyp40 and a related co-chaperone PP5 were much higher in prostate cancer cells lines (LNCaP, PC-3 and DU145) compared to primary prostate cells, and that the AR of LNCaP cells can interact with Cyp40. In the absence of androgen, CsA caused inhibition of cell growth in the AR-positive LNCaP and AR-negative PC-3 and DU145 cell lines. Interestingly, FK506 only inhibited LNCaP cells, suggesting a dependence on AR for this effect. Both CsA and FK506 inhibited growth without inducing apoptosis. In LNCaP cells, CsA completely blocked androgen-stimulated growth, whereas FK506 was partially effective. Further studies in LNCaP cells revealed that CsA and FK506 were able to block or attenuate several stages of AR signaling, including hormone binding, nuclear translocation and activity at several AR-responsive reporter and endogenous genes. These findings provide the first evidence that CsA and FK506 can negatively modulate proliferation of prostate cells in vitro. Immunophilins may now serve as new targets to disrupt AR-mediated prostate cancer growth. PMID:17615153

  1. Inhibition of Pterygium Fibroblast Migration and Outgrowth by Bevacizumab and Cyclosporine A Involves Down-Regulation of Matrix Metalloproteinases-3 and -13

    PubMed Central

    Gum, Sang Il; Park, Su-Bin; Ma, Jin Yeul; Kim, Yong Il; Lee, Kyoo Won

    2017-01-01

    We examined the connection between matrix metalloproteinase (MMP) expression/activity and pterygium fibroblast migration, and how these were affected by bevacizumab and/or cyclosporine A (CsA). Fibroblasts were obtained from 20 pterygia and 6 normal conjunctival specimens. Expression levels of MMP-3 and MMP-13 were examined after bevacizumab administration. Immunofluorescence staining was used to examine expression of both MMPs in fibroblasts migrating out from explanted pterygium tissues. Rates of cell migration from explant-cultured pterygia tissues and scratch-wounded confluent pterygium fibroblasts were examined in the presence of MMP-3 or MMP-13 inhibitors, as well as bevacizumab and/or CsA. A scratch wound healing migration assay was performed to determine the effects of bevacizumab and/or CsA. Protein expression of both MMPs in pterygium tissues and in cells migrating from organ-cultured pterygium tissues was greater than that observed in normal cells. Inhibition of the activities of both MMPs decreased their expression levels; these were also significantly reduced in bevacizumab-injected pterygium tissues. Bevacizumab significantly reduced the expression of both MMPs and cell migration. Pretreatment with CsA prior to bevacizumab exposure markedly inhibited cell migration and the expression of both MMPs. CsA enhanced the inhibitory effects of bevacizumab on pterygium fibroblast migration in vitro, possibly by inhibiting expression of both MMPs. These findings suggest that combined CsA and bevacizumab treatment may provide a potential therapeutic strategy for reducing the rate of pterygium recurrence. PMID:28068383

  2. The Cratylia mollis seed lectin induces membrane permeability transition in isolated rat liver mitochondria and a cyclosporine a-insensitive permeability transition in Trypanosoma cruzi mitochondria.

    PubMed

    Fernandes, Mariana P; Leite, Ana C R; Araújo, Flavia F B; Saad, Sara T O; Baratti, M O; Correia, M T S; Coelho, Luana C B B; Gadelha, Fernanda R; Vercesi, Anibal E

    2014-01-01

    Previous results provided evidence that Cratylia mollis seed lectin (Cramoll 1,4) promotes Trypanosoma cruzi epimastigotes death by necrosis via a mechanism involving plasma membrane permeabilization to Ca(2+) and mitochondrial dysfunction due to matrix Ca(2+) overload. In order to investigate the mechanism of Ca(2+) -induced mitochondrial impairment, experiments were performed analyzing the effects of this lectin on T. cruzi mitochondrial fraction and in isolated rat liver mitochondria (RLM), as a control. Confocal microscopy of T. cruzi whole cell revealed that Cramoll 1,4 binding to the plasma membrane glycoconjugates is followed by its internalization and binding to the mitochondrion. Electrical membrane potential (∆Ψm ) of T. cruzi mitochondrial fraction suspended in a reaction medium containing 10 μM Ca(2+) was significantly decreased by 50 μg/ml Cramoll 1,4 via a mechanism insensitive to cyclosporine A (CsA, membrane permeability transition (MPT) inhibitor), but sensitive to catalase or 125 mM glucose. In RLM suspended in a medium containing 10 μM Ca(2+) this lectin, at 50 μg/ml, induced increase in the rate of hydrogen peroxide release, mitochondrial swelling, and ∆Ψm disruption. All these mitochondrial alterations were sensitive to CsA, catalase, and EGTA. These results indicate that Cramoll 1, 4 leads to inner mitochondrial membrane permeabilization through Ca(2+) dependent mechanisms in both mitochondria. The sensitivity to CsA in RLM characterizes this lectin as a MPT inducer and the lack of CsA effect identifies a CsA-insensitive MPT in T. cruzi mitochondria.

  3. cps1+, a Schizosaccharomyces pombe gene homolog of Saccharomyces cerevisiae FKS genes whose mutation confers hypersensitivity to cyclosporin A and papulacandin B.

    PubMed Central

    Ishiguro, J; Saitou, A; Durán, A; Ribas, J C

    1997-01-01

    The Schizosaccharomyces pombe cps1-12 (for chlorpropham supersensitive) mutant strain was originally isolated as hypersensitive to the spindle poison isopropyl N-3-chlorophenyl carbamate (chlorpropham) (J. Ishiguro and Y. Uhara, Jpn. J. Genet. 67:97-109, 1992). We have found that the cps1-12 mutation also confers (i) hypersensitivity to the immunosuppressant cyclosporin A (CsA), (ii) hypersensitivity to the drug papulacandin B, which specifically inhibits 1,3-beta-D-glucan synthesis both in vivo and in vitro, and (iii) thermosensitive growth at 37 degrees C. Under any of these restrictive treatments, cells swell up and finally lyse. With an osmotic stabilizer, cells do not lyse, but at 37 degrees C they become multiseptated and multibranched. The cps1-12 mutant, grown at a restrictive temperature, showed an increase in sensitivity to lysis by enzymatic cell wall degradation, in in vitro 1,3-beta-D-glucan synthase activity (173% in the absence of GTP in the reaction), and in cell wall biosynthesis (130% of the wild-type amount). Addition of Ca2+ suppresses hypersensitivity to papulacandin B and septation and branching phenotypes. All of these data suggest a relationship between the cps1+ gene and cell wall synthesis. A DNA fragment containing the cps1+ gene was cloned, and sequence analysis indicated that it encodes a predicted membrane protein of 1,729 amino acids with 15 to 16 transmembrane domains. S. pombe cps1p has overall 55% sequence identity with Fks1p or Fks2p, proposed to be catalytic or associated subunits of Saccharomyces cerevisiae 1,3-beta-D-glucan synthase. Thus, the cps1+ product might be a catalytic or an associated copurifying subunit of the fission yeast 1,3-beta-D-glucan synthase that plays an essential role in cell wall synthesis. PMID:9401022

  4. Cyclosporin-A associated malignancy

    PubMed Central

    Durnian, Jonathan M; Stewart, Rosalind MK; Tatham, Richard; Batterbury, Mark; Kaye, Stephen B

    2007-01-01

    The use of cyclosporin is well established within the ophthalmology community, especially against sight threatening intraocular inflammation. It is well known however, that immunosuppression in general is a risk factor for the development of malignancy and numerous studies point to the risk imposed by cyclosporin. This article analyses and reviews all relevant studies with regard to the development of malignancy associated with the use of cyclosporin and extrapolates this into the ophthalmic setting. This is to enable clinicians to assess the risks in individual patients and to present a monitoring regime which can be used in patients undergoing cyclosporin treatment. The review is solely concerned with the risk of the development of malignancy following cyclosporin immunosuppression and not with any other adverse effect. PMID:19668519

  5. Evaluation of Cyclosporine A with β-TCP in the Treatment of Human Infra bony Defects – A Randomized Controlled Pilot Study

    PubMed Central

    Reddy, Krishnajaneya; Avula, Haritha; Mishra, Ashank; Kalakonda, Butchibabu; Pandey, Ruchi

    2017-01-01

    Abstract Introduction Cyclosporine A (CsA), an immunosuppressant, is considered a life saver drug in organ transplant cases. It has also been tested in animal and human studies for periodontal applications as it selectively inhibits T lymphocyte proliferation, Interleukin-2 (IL-2) and other cytokine production, without any effect on T suppressor cells, thereby suppressing the cell mediated immunity and suppressing the inflammation. Inflammatory and immunological responses have been found to be decreased and bone formation is found to be increased in immunosuppressed animals. CsA is also supposed to potentiate osseous regeneration due to increase in the bone alkaline phosphatase levels and a direct activating effect on osteoblasts. Aim The present study was aimed at evaluating locally administered low dose of CsA which is potent immunosuppressant along with β-Tricalcium phosphate (β-TCP) in comparison with β TCP alone, in the treatment of human infrabony defects, over a period of six months. Materials and Methods Thirty two systemically healthy chronic periodontitis patients with infrabony defects were included in the randomized, controlled, parallel arm study and were allocated into either Group A (n =16), patients treated with β-TCP + CsA (2 mg) or Group B (n =16), patients treated with β-TCP. Clinical parameters [Relative Attachment Level (RAL), Probing Depth (PD), Gingival Recession (GR)] and radiographic parameters were measured at baseline and six months postoperatively. Statistical analysis was done using SPSS version 16 software. Student’s paired and independent t-test were used for intra and inter-group analysis. Results Both Group A and Group B showed statistically significant improvements in clinical and radiographic parameters from base line to six months post-operatively. The Clinical Attachment Level (CAL) gain, Linear Bone Growth (LBG) and Percentage Bone Fill (% BF) were 2.38±1.12 mm, 1.90±1.48 mm and 49.83±29.23 mm in Group A and 2.57±1

  6. Distinct cyclosporin a doses are required to enhance bone formation induced by cyclic and rest-inserted loading in the senescent skeleton.

    PubMed

    Srinivasan, Sundar; Threet, Dewayne; Worton, Leah E; Ausk, Brandon J; Bain, Steven D; Gardiner, Edith M; Kwon, Ronald Y; Gross, Ted S

    2014-01-01

    Age-related decline in periosteal adaptation negatively impacts the ability to utilize exercise to enhance bone mass and strength in the elderly. We recently observed that in senescent animals subject to cyclically applied loading, supplementation with Cyclosporin A (CsA) substantially enhanced the periosteal bone formation rates to levels observed in young animals. We therefore speculated that if the CsA supplement could enhance bone response to a variety of types of mechanical stimuli, this approach could readily provide the means to expand the range of mild stimuli that are robustly osteogenic at senescence. Here, we specifically hypothesized that a given CsA supplement would enhance bone formation induced in the senescent skeleton by both cyclic (1-Hz) and rest-inserted loading (wherein a 10-s unloaded rest interval is inserted between each load cycle). To examine this hypothesis, the right tibiae of senescent female C57BL/6 mice (22 Mo) were subjected to cyclic or rest-inserted loading supplemented with CsA at 3.0 mg/kg. As previously, we initially found that while the periosteal bone formation rate (p.BFR) induced by cyclic loading was enhanced when supplemented with 3.0 mg/kg CsA (by 140%), the response to rest-inserted loading was not augmented at this CsA dosage. In follow-up experiments, we observed that while a 30-fold lower CsA dosage (0.1 mg/kg) significantly enhanced p.BFR induced by rest-inserted loading (by 102%), it was ineffective as a supplement with cyclic loading. Additional experiments and statistical analysis confirmed that the dose-response relations were significantly different for cyclic versus rest-inserted loading, only because the two stimuli required distinct CsA dosages for efficacy. While not anticipated a priori, clarifying the complexity underlying the observed interaction between CsA dosage and loading type holds potential for insight into how bone response to a broad range of mechanical stimuli may be substantially enhanced in

  7. Preparation and evaluation of cyclosporin A-containing proliposomes: a comparison of the supercritical antisolvent process with the conventional film method

    PubMed Central

    Karn, Pankaj Ranjan; Jin, Su-Eon; Lee, Benjamin Joon; Sun, Bo Kyung; Kim, Min-Soo; Sung, Jong-Hyuk; Hwang, Sung-Joo

    2014-01-01

    Objectives The objectives of this study were to prepare cyclosporin A (CsA)-containing proliposomes using the supercritical antisolvent (SAS) process and the conventional thin film method for the comparative study of proliposomal formulations and to evaluate the physicochemical properties of these proliposomes. Methods CsA-containing proliposomes were prepared by the SAS process and the conventional film method, composed of natural and synthetic phospholipids. We investigated particle size, polydispersity index, and zeta potential of CsA-containing proliposomes. In addition, both production yield and entrapment efficiency of CsA in different proliposomes were analyzed. Physicochemical properties of CsA-containing proliposomes were also evaluated, using differential scanning calorimetry and X-ray diffraction. The morphology and size of CsA-containing proliposomes were confirmed, using scanning electron microscopy. We checked the in vitro release of CsA from CsA-containing proliposomes prepared by different preparation methods, comparing them with Restasis® as a positive control and the stability of SAS-mediated proliposomes was also studied. Results CsA-containing proliposomes formed by the SAS process had a relatively smaller particle size, with a narrow size distribution and spherical particles compared with those of conventionally prepared proliposomes. The yield and entrapment efficiency of CsA in all proliposomes varied from 85% to 92% and from 86% to 89%, respectively. Differential scanning calorimetry and X-ray diffraction studies revealed that the anhydrous lactose powder used in this formulation retained its crystalline form and that CsA was present in an amorphous form. Proliposome powders were rapidly converted to liposomes on contact with water. The in vitro release study of proliposomal formulations demonstrated a similar pattern to Restasis®. The SAS-mediated CsA-containing proliposomes were stable on storage, with no significant changes in particle

  8. Long-chain α,ω-dioic acids as inducers of cyclosporin A-insensitive nonspecific permeability of the inner membrane of liver mitochondria loaded with calcium or strontium ions.

    PubMed

    Dubinin, M V; Adakeeva, S I; Samartsev, V N

    2013-04-01

    Long-chain saturated monocarboxylic fatty acids can induce nonspecific permeability of the inner membrane (open pores) of liver mitochondria loaded with Ca2+ or Sr(2+) by the mechanism insensitive to cyclosporin A. In this work we investigated the effect of their metabolites - α,ω-dioic (dicarboxylic) acids - as potential inducers of pore opening by a similar mechanism. It was established that the addition of α,ω-hexadecanedioic acid (HDA) at a concentration of 10-30 µM to liver mitochondria loaded with Ca2+ or Sr(2+) leads to swelling of the organelles and release of these ions from the matrix. The maximum effect of HDA is observed at 50 µM Ca2+ concentration. Cyclosporin A at a concentration of 1 µM, previously added to the mitochondria, did not inhibit the observed processes. The calcium uniporter inhibitor ruthenium red, which blocks influx of Ca2+ and Sr(2+) to the matrix of mitochondria, prevented HDA-induced swelling. The effect of HDA as inducer of swelling of mitochondria was compared with similar effects of α,ω-tetradecanedioic and α,ω-dodecanedioic acids whose acyl chains are two and four carbon atoms shorter than HDA, respectively. It was found that the efficiency of these α,ω-dioic acids decreases with reducing number of carbon atoms in their acyl chains. It was concluded that in the presence of Ca2+ or Sr(2+) long-chain saturated α,ω-dioic acids can induce a cyclosporin A-insensitive permeability of the inner membrane (open pores) of liver mitochondria as well as their monocarboxylic analogs.

  9. Anti-CD11a ameliorates disease in the human psoriatic skin-SCID mouse transplant model: comparison of antibody to CD11a with Cyclosporin A and clobetasol propionate.

    PubMed

    Zeigler, M; Chi, Y; Tumas, D B; Bodary, S; Tang, H; Varani, J

    2001-09-01

    The present study assesses the applicability of human skin-SCID (severe combined immunodeficiency) mouse chimeras in testing antipsoriatic therapeutics. Three agents were examined: (1) a monoclonal antibody to the alpha subunit of leukocyte function associated antigen-1 integrin (CD11a); (2) Cyclosporin A; and (3) clobetasol propionate (Temovate), a potent topical corticosteroid used clinically in the treatment of psoriasis. Skin transplanted to SCID mice from normal human volunteers or from psoriatic lesional skin was allowed to heal for 3 to 5 weeks before application of test reagents. During this period, psoriatic skin, which was 3.8-fold thicker than the corresponding normal skin before transplantation, maintained its phenotype (ie, increased epidermal thickness, rete ridges with blunted ends, and intralesional presence of T lymphocytes). Transplanted normal human skin, however, underwent a hyperplastic response during this period, resulting in a 2.4-fold increase in epidermal thickness. After the healing period, animals transplanted with normal or psoriatic skin were treated for 14 days by daily intraperitoneal injection of either Cyclosporin A or a monoclonal antibody to human CD11a, or by topical application of clobetasol propionate. At the end of the treatment period, the mice were killed and the tissue evaluated morphometrically for changes in epidermal thickness and immunohistologically for the presence of T lymphocytes. Both Cyclosporin A and anti-CD11a reduced the epidermal thickness of transplanted psoriatic skin, whereas neither reagent significantly reduced the thickness of transplanted normal skin. T lymphocytes were detected in the skin from treated animals; there did not seem to be any reduction in the number of T lymphocytes. Clobetasol propionate reduced the epidermal thickness of both normal and psoriatic skin. These data indicate that, in this model, therapies directed against pathophysiologic mechanisms that contribute to psoriasis can be

  10. Interaction Study of an Amorphous Solid Dispersion of Cyclosporin A in Poly-Alpha-Cyclodextrin with Model Membranes by (1)H-, (2)H-, (31)P-NMR and Electron Spin Resonance.

    PubMed

    Debouzy, Jean-Claude; Crouzier, David; Bourbon, Fréderic; Lahiani-Skiba, Malika; Skiba, Mohamed

    2014-01-01

    The properties of an amorphous solid dispersion of cyclosporine A (ASD) prepared with the copolymer alpha cyclodextrin (POLYA) and cyclosporine A (CYSP) were investigated by (1)H-NMR in solution and its membrane interactions were studied by (1)H-NMR in small unilamellar vesicles and by (31)P (2)H NMR in phospholipidic dispersions of DMPC (dimyristoylphosphatidylcholine) in comparison with those of POLYA and CYSP alone. (1)H-NMR chemical shift variations showed that CYSP really interacts with POLYA, with possible adduct formation, dispersion in the solid matrix of the POLYA, and also complex formation. A coarse approach to the latter mechanism was tested using the continuous variations method, indicating an apparent 1 : 1 stoichiometry. Calculations gave an apparent association constant of log Ka = 4.5. A study of the interactions with phospholipidic dispersions of DMPC showed that only limited interactions occurred at the polar head group level ((31)P). Conversely, by comparison with the expected chain rigidification induced by CYSP, POLYA induced an increase in the fluidity of the layer while ASD formation led to these effects almost being overcome at 298 K. At higher temperature, while the effect of CYSP seems to vanish, a resulting global increase in chain fluidity was found in the presence of ASD.

  11. Effect of the six-mer synthetic peptide (AT1002) fragment of zonula occludens toxin on the intestinal absorption of cyclosporin A.

    PubMed

    Song, Keon-Hyoung; Fasano, Alessio; Eddington, Natalie D

    2008-03-03

    Zonula occludens toxin (Zot) and its biologically active fragment, delta G, have been shown to reversibly open tight junctions (TJ) in endothelial and epithelial cells. Recently, a six-mer synthetic peptide H-FCIGRL-OH (AT1002) was identified and synthesized that retains the Zot permeating effect on intercellular TJ. The objective of this study was to evaluate the biological activity of AT1002 on enhancing the oral administration of cyclosporin A (CsA). The intestinal permeability enhancing effect of AT1002 on the transport of CsA across Caco-2 cell monolayers was examined after the following treatments, i.e., CsA, CsA/protease inhibitors (PI), CsA/PI/benzalkonium chloride (BC), CsA/AT1002, CsA/PI/AT1002, and CsA/PI/BC/AT1002 (CsA 0.5 microCi/ml, PI (bestatin 15 mM and E-64 5mM), BC 0.005 w/v%, and AT1002 5mM, respectively). Apparent permeability coefficients (P app) were calculated for each treatment. In addition, four treatments, i.e., CsA, CsA/PI/BC, CsA/AT1002, and CsA/PI/BC/AT1002 (CsA 120 microCi/kg, PI (bestatin 30 mg/kg and E-64 10mg/kg), BC 0.1 w/v%, and AT1002 doses of 5, 10 or 40 mg/kg, respectively) were prepared and administered intraduodenally to male Sprague-Dawley rats (230-280 g, n=4-5). Blood samples were collected at 0, 20, 60, and 120 min post-dosing and CsA plasma concentrations were determined subsequently using a Beckman Liquid Scintillation Counter. No significant increases in CsA transport were observed in the Caco-2 cell culture experiments following pre-treatment with AT1002 (5mM). Even though, AT1002 appeared to increase the P app of CsA in each treatment (CsA/AT1002, 1.54+/-0.13 x 10(-6)cm/s and CsA/PI/AT1002, 1.76+/-0.05 x 10(-6)cm/s) compared to each control (CsA and CsA/PI), respectively. The plasma concentration of CsA was significantly increased over a range of 1.55-2.50 at 10 and 40 mg/kg dose of AT1002. Also, AUC 0-120 min of CsA over a range of 1.64-2.14 and the Cmax of CsA over a range of 1.77-2.56 was statistically and

  12. Suppression of local and systemic responses in streptococcal cell wall-induced acute inflammation of the air pouch by cyclosporine A. Comparison with the effects of two anti-inflammatory bis-benzimidazoles.

    PubMed Central

    Dieter Geratz, J.; Pryzwansky, K. B.; Schwab, J. H.; Anderle, S. K.; Tidwell, R. R.

    1993-01-01

    Injection of streptococcus group A cell wall-derived peptidoglycan polysaccharide into a subcutaneous air pouch causes local outpouring of neutrophils and macrophages and distant hemopoietic proliferation in spleen and bone marrow. Cyclosporine A (CyA) suppressed neutrophil accumulation and all cell lines of hemopoiesis. trans-1,2-Bis(5-amidino-2-benzimidazolyl)ethene (BBE) also interfered with neutrophil exudation, yet reduced only the erythroid component of the hemopoietic process. The ethane analogue of BBE, on the other hand, did not prevent neutrophil emigration, but held down splenic erythropoiesis and myelopoiesis. All three compounds stimulated streptococcus group A cell wall-derived peptidoglycan polysaccharide uptake by pouch macrophages. CyA being the least active, BBE and its ethane analogue also produced a shift of wear-and-tear pigment from large numbers of small splenic macro-phages into small numbers of large macrophages. The pouch model is very useful in the study of anti-inflammatory compounds and has furnished the first evidence of CyA interference with massive neutrophilic infiltration and with hemopoietic signals. Images Figure 3 Figure 4 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 PMID:8475995

  13. Extended follow-up of the CYCLOFA-LUNE trial comparing two sequential induction and maintenance treatment regimens for proliferative lupus nephritis based either on cyclophosphamide or on cyclosporine A.

    PubMed

    Závada, J; Sinikka Pesicková, S; Rysavá, R; Horák, P; Hrncír, Z; Lukác, J; Rovensky, J; Vítová, J; Havrda, M; Rychlík, I; Böhmova, J; Vlasáková, V; Slatinská, J; Zadrazil, J; Olejárová, M; Tegzova, D; Tesar, V

    2014-01-01

    Objective To evaluate the extended follow-up of the CYCLOFA-LUNE trial, a randomized prospective trial comparing two sequential induction and maintenance treatment regimens for proliferative lupus nephritis based either on cyclophosphamide (CPH) or cyclosporine A (CyA). Patients and methods Data for kidney function and adverse events were collected by a cross-sectional survey for 38 of 40 patients initially randomized in the CYCLOFA-LUNE trial. Results The median follow-up time was 7.7 years (range 5.0-10.3). Rates of renal impairment and end-stage renal disease, adverse events (death, cardiovascular event, tumor, premature menopause) did not differ between the CPH and CyA group, nor did mean serum creatinine, 24 h proteinuria and SLICC damage score at last follow-up. Most patients in both groups were still treated with glucocorticoids, other immunosuppressant agents and blood pressure lowering drugs. Conclusion An immunosuppressive regimen based on CyA achieved similar clinical results to that based on CPH in the very long term.

  14. Cyclosporin A inhibits CD11a/CD18 adhesion molecules due to inhibition of TNFα and IL-1β levels in the mouse model of pleurisy induced by carrageenan

    PubMed Central

    Dalmarco, Eduardo Monguilhott; Medeiros, Yara Santos

    2008-01-01

    The mouse model of pleurisy induced by carrageenan is characterized by a significant enhancement of cell migration due to neutrophils 4 h after pleurisy induction. Forty-eight hours after pleurisy induction, a significant increase in cell migration due to mononuclear cells occurs. Recently, studies in our laboratory have demonstrated that cyclosporine A (CsA) inhibits leukocyte migration in the pleural cavity and lungs in the mouse model of pleurisy induced by carrageenan. In the present work we evaluated whether CsA was able to downregulate CD11a/CD18 adhesion molecule in the lungs, as well as TNFα and IL-1β levels in the fluid leakage of the pleural cavity in this model. Our results showed that CsA significantly decreased CD11a/CD18 in the lungs, as well as TNFα and IL-1β levels in the fluid leakage of the pleural cavity 4 h and 48 h after pleurisy induction. It is our hypothesis that the inhibitory effect elicited by CsA upon these adhesion molecules may be also be attributed to the downregulation of TNFα and IL-1β cytokines. PMID:19262158

  15. 1,25(OH)2D3 and VDR Signaling Pathways Regulate the Inhibition of Dectin-1 Caused by Cyclosporine A in Response to Aspergillus Fumigatus in Human Corneal Epithelial Cells

    PubMed Central

    Xia, Yiping; Zhao, Guiqiu; Lin, Jing; Li, Cui; Cong, Lin; Jiang, Nan; Xu, Qiang; Wang, Qian

    2016-01-01

    Background The objective of this study is to observe whether cyclosporine A (CsA) inhibits the expression of dectin-1 in human corneal epithelial cells infected with Aspergillus fumigatus (A. fumigatus) and to investigate the molecular mechanisms of the inhibition. Methods Immortalized human corneal epithelial cells (HCECs) were pretreated with 1,25(OH)2D3 and VDR inhibitor for 1 h, and then they were pretreated with CsA for 12h. After these pretreatments, the HCECs were stimulated with A. fumigatus and curdlan respectively, and the expression of dectin-1 and proinflammatory cytokines (IL-1β and TNF-α) were detected by RT-PCR, western blot and ELISA. Results Dectin-1 mRNA and dectin-1 protein expression increased when HCECs were stimulated with A. fumigatus or curdlan, and CsA inhibited the dectin-1 expression both in mRNA and protein levels specifically. Dectin-1 and proinflammatory cytokine expression levels were higher when HCECs were pretreated with VDR inhibitor and CsA compared to pretreatment with CsA alone, while dectin-1 and proinflammatory cytokine levels were lower when HCECs were pretreated with 1,25(OH)2D3 and CsA compared to pretreatment with CsA alone. Conclusions These data provide evidence that CsA can inhibit the expression of dectin-1 and proinflammatory cytokines through dectin-1 when HCECs are stimulated by A. fumigatus or curdlan. The active form of vitamin D, 1,25(OH)2D3, and VDR signaling pathway regulate the inhibition of CsA. The inhibition is enhanced by 1,25(OH)2D3, and the VDR inhibitor suppresses the inhibition. PMID:27755569

  16. Cyclosporin A is superior to cyclophosphamide in children with steroid-resistant nephrotic syndrome-a randomized controlled multicentre trial by the Arbeitsgemeinschaft für Pädiatrische Nephrologie.

    PubMed

    Plank, Christian; Kalb, Veronica; Hinkes, Bernward; Hildebrandt, Friedhelm; Gefeller, Olaf; Rascher, Wolfgang

    2008-09-01

    First line immunosuppressive treatment in steroid-resistant nephrotic syndrome in children is still open to discussion. We conducted a controlled multicentre randomized open label trial to test the efficacy and safety of cyclosporin A (CSA) versus cyclophosphamide pulses (CPH) in the initial therapy of children with newly diagnosed primary steroid-resistant nephrotic syndrome and histologically proven minimal change disease, focal segmental glomerulosclerosis or mesangial hypercellularity. Patients in the CSA group (n = 15) were initially treated with 150 mg/m(2) CSA orally to achieve trough levels of 120-180 ng/ml, while patients in the CPH group (n = 17) received CPH pulses (500 mg/m(2) per month intravenous). All patients were on alternate prednisone therapy. Patients with proteinuria >40 mg/m(2) per hour at 12 weeks of therapy were allocated to a non-responder protocol with high-dose CSA therapy or methylprednisolone pulses. At week 12, nine of the 15 (60%) CSA patients showed at least partial remission, evidences by a reduction of proteinuria <40 mg/h per m(2). In contrast, three of the 17 (17%) CPH patients responded (p < 0.05, intention-to-treat). Given these results, the study was stopped, in accordance with the protocol. After 24 weeks, complete remission was reached by two of the 15 (13%) CSA and one of the 17 (5%) CPH patients (p = n.s.). Partial remission was achieved by seven of the 15 (46%) CSA and two of the 15 (11%) CPH patients (p <0.05). Five patients in the CSA group and 14 patients in the CPH group were withdrawn from the study, most of them during the non-responder protocol. The number of adverse events was comparable between both groups. We conclude that CSA is more effective than CPH in inducing at least partial remission in steroid-resistant nephrotic syndrome in children.

  17. Cyclosporin A promotes proliferating cell nuclear antigen expression and migration of human cytotrophoblast cells via the mitgen-activated protein kinase-3/1-mediated nuclear factor-κB signaling pathways.

    PubMed

    Wang, Song-Cun; Yu, Min; Li, Yan-Hong; Piao, Hai-Lan; Tang, Chuan-Lin; Sun, Chan; Zhu, Rui; Li, Ming Qing; Jin, Li-Ping; Li, Da-Jin; Du, Mei-Rong

    2013-01-01

    Our previous studies have demonstrated that cyclosporin A (CsA) promotes the proliferation and migration of human trophoblasts via the mitgen-activated protein kinase-3/1 (MAPK3/1) pathway. In the present study, we further investigated the role of nuclear factor (NF)-κB in the CsA-induced trophoblast proliferating cell nuclear antigen (PCNA) expression and migration, and its relationship to MAPK3/1 signal. Flow cytometry was used to analyze the expression of PCNA in trophoblasts. The migration of human primary trophoblasts was determined by wound-healing assay and transwell migration assay. Western blot analysis was performed to evaluate the activation of NF-κB p65 and NF-κB inhibitory protein I-κB in human trophoblasts. We found that treatment with CsA promotes PCNA expression and migration of human trophoblast in a dose-associated manner. Blocking of the MAPK3/1 signal abrogated the enhanced PCNA expression and migration in trophoblasts by CsA. In addition, CsA increased the phosphorylation of NF-κB p65 and the inhibitor I-κB in human trophoblasts in a time-related manner. Pretreatment with MAPK3/1 inhibitor U0126 abrogated the phosphorylation of NF-κB p65 and I-κB. Accordingly, the CsA-induced enhancement of PCNA expression and migration in trophoblasts was also decreased. This CsA-induced enhancement in the expression and migration of trophoblasts was abolished by pretreatment with pyrrolidine dithiocarbamate, a specific NF-κB inhibitor. Thus, our results suggest that CsA promotes PCNA expression and migration of human trophoblasts via MAPK-mediated NF-κB activation.

  18. Cinnamaldehyde-induced apoptosis in human hepatoma PLC/PRF/5 cells involves the mitochondrial death pathway and is sensitive to inhibition by cyclosporin A and z-VAD-fmk.

    PubMed

    Lin, Liang-Tzung; Tai, Chen-Jei; Chang, Shun-Pang; Chen, Jin-Liang; Wu, Shu-Jing; Lin, Chun-Ching

    2013-12-01

    Cinnamaldehyde (CIN) has been shown to exert chemopreventive activity against several types of human cancer cells. We previously reported that CIN induced apoptosis of human hepatoma PLC/PRF/5 cells and this effect was associated with activation of the pro-apoptotic Bcl-2 family of proteins and the MAPK cascade. To further clarify the underlying mechanism of CIN-induced apoptosis, we examined in this study its relationship with the mitochondrial death pathway using the mitochondrial permeability transition (MPT) inhibitor, cyclosporin A (CsA), and the general caspase inhibitor, z-VAD-fmk. Results indicated that CIN-induced apoptosis involved enhanced ROS generation, disruption of mitochondrial potential, and the mitochondrial release of cytochrome c and Smac/DIABLO into the cytosol, which in turn promoted caspase-3 to its active form and the subsequent cleavage of PARP. Treatment with CIN also downregulated protein levels of the anti-apoptotic factors XIAP and Bcl-2 with concomitant accumulation of the pro-apoptotic Bax in a timedependent manner. These mitochondria-related apoptotic effects induced by CIN were however blocked by CsA and z-VAD-fmk pretreatments, which prevented cells from undergoing programmed cell death triggered by CIN. Furthermore, the increase of Bax and decrease of Bcl-2 and XIAP protein expression due to CIN treatment were also reversely modulated by the two inhibitors. Taken together, these results suggested that CIN is an apoptotic inducer that acts on the mitochondrial death pathway in PLC/PRF/5 cells and its effect could be blocked by CsA and z-VAD-fmk.

  19. An antioxidant Trolox restores decreased oral absorption of cyclosporine A after liver ischemia-reperfusion through distinct mechanisms between CYP3A and P-glycoprotein in the small intestine.

    PubMed

    Ikemura, Kenji; Inoue, Koichi; Mizutani, Hideki; Oka, Hisao; Iwamoto, Takuya; Okuda, Masahiro

    2012-09-05

    Oxidative stress is a critical mediator of various injuries following ischemia-reperfusion (I/R) associated with organ transplantation. Although oral bioavailability of cyclosporine A (CsA) was decreased by increased first-pass metabolism through CYP3A and P-glycoprotein (P-gp) specifically in the upper small intestine after liver I/R, the mechanism responsible for them remained to be clarified. In the present study, the effect of Trolox (an α-tocopherol analogue) on the decreased oral absorption of CsA through elevated intestinal CYP3A and P-gp after liver I/R and their regulations were investigated. Rats were subjected to 60 min of liver ischemia followed by 12h of reperfusion. Trolox was administered intravenously 5 min before reperfusion. Trolox diminished the increased malondialdehyde and total glutathione levels in plasma by liver I/R and concomitantly prevented the decreased area under the blood concentration-time curve of orally administered CsA as well as initial absorption rate of CsA from upper small intestine. The elevated CYP3A mRNA and activity in the upper small intestine as well as expression levels of P-gp in upper, middle, and lower small intestines after liver I/R were attenuated by Trolox administration. The elevations of CYP3A levels specifically in the upper small intestine of I/R rats were correlated with the lithocholic acid levels in the bile. These results demonstrate that Trolox ameliorates the decreased oral absorption of CsA through elevated intestinal CYP3A and P-gp by preventing oxidative stress, where the biliary lithocholic acid may be responsible for the elevated transcription of CYP3A specifically in the upper small intestine after liver I/R.

  20. Comparative evaluation of the antiproliferative effect of cyclosporin A and gamma-interferon on normal and HPV-transformed keratinocytes by cell counting, MTT assay and tritiated thymidine incorporation.

    PubMed

    Marionnet, A V; Lizard, G; Chardonnet, Y; Schmitt, D

    1997-02-01

    We compared three techniques, the MTT tetrazolium assay, cell counting, and tritiated thymidine ([3H]TdR) incorporation assay to measure the antiproliferative effect of cyclosporin A (CsA) and interferon-gamma (IFN-gamma) on normal human skin keratinocyte cultures (NHK) used at the second passage and human papilomavirus type 16- and 18-transformed cell lines (EK16 and EK18) exposed continuously to the drugs for 3 days. The three techniques showed that under CsA (0.5 and 8 micrograms/ml) and IFN-gamma (5 and 160 U/ml) treatments the cells remained viable and that the growth of keratinocytes was inhibited. For IFN-gamma, the MTT colorimetric assay consistently underestimated its growth inhibitory activity as compared to cell counting or [3H]TdR incorporation, whatever the cells used. For high doses of CsA, MTT and cell counting gave similar percentages, of inhibitory activity whatever the cells; MTT underestimated this activity as compared to [3H]TdR incorporation only in NHK and EK18 cells, whereas similar results were obtained with EK16 cells. In conclusion, this investigations shows that MTT sensitivity differed with the drug and also according to the keratinocyte cultures. The MTT test is clearly not appropriate for study of IFN-gamma treatment whatever the keratinocytes used. Such discrepancies indicate that the MTT test should be done with care on cultures to measure the effects of drugs on cell growth; the growth inhibition should be carefully considered and it would be best if two different methods were used.

  1. Absorption of cyclosporine A after oral dosing.

    PubMed

    Grevel, J

    1986-12-01

    Variability in the absorption of CsA seems to contribute to the observed lack of correlation between the size of the oral dose and the trough concentration at steady state. Absorption is probably improved by thorough dispersion of the oral solution of CsA in the drink the patient prefers. Evidence for GI metabolism of CsA has only been gathered in animal experiments. The importance of bile for absorption of CsA into the portal blood is established. The bioavailability of CsA does not seem to be determined by the metabolism during the first passage through the liver. Enterohepatic recycling is likely for CsA metabolites and unlikely for unchanged CsA. A pharmacokinetic model that assumes zero-order absorption of CsA describes human data better than a model with first-order absorption. According to the zero-order model, CsA is absorbed only in the upper part of the small intestine by a mechanism that operates under saturation. Two independent findings in transplantation patients support this model. First, it was shown that small doses of CsA produce disproportionally high blood concentrations, probably due to a better bioavailability. Second, accelerated transit times in the intestine (diarrhea) lead to unexpectedly low blood concentrations, probably due to poor bioavailability. Further factors have been identified that cause low absorption of CsA: liver dysfunction and external bile drainage after liver transplantation. The influence of food on the absorption of CsA is still not determined conclusively, but it seems that giving CsA together with a standard breakfast results in higher blood concentrations. The observed increase in the bioavailability of CsA with time after transplantation could be caused by the attempt to steadily lower the dose.

  2. Dose Adjustment Strategy of Cyclosporine A in Renal Transplant Patients: Evaluation of Anthropometric Parameters for Dose Adjustment and C0 vs. C2 Monitoring in Japan, 2001-2010

    PubMed Central

    Kokuhu, Takatoshi; Fukushima, Keizo; Ushigome, Hidetaka; Yoshimura, Norio; Sugioka, Nobuyuki

    2013-01-01

    The optimal use and monitoring of cyclosporine A (CyA) have remained unclear and the current strategy of CyA treatment requires frequent dose adjustment following an empirical initial dosage adjusted for total body weight (TBW). The primary aim of this study was to evaluate age and anthropometric parameters as predictors for dose adjustment of CyA; and the secondary aim was to compare the usefulness of the concentration at predose (C0) and 2-hour postdose (C2) monitoring. An open-label, non-randomized, retrospective study was performed in 81 renal transplant patients in Japan during 2001-2010. The relationships between the area under the blood concentration-time curve (AUC0-9) of CyA and its C0 or C2 level were assessed with a linear regression analysis model. In addition to age, 7 anthropometric parameters were tested as predictors for AUC0-9 of CyA: TBW, height (HT), body mass index (BMI), body surface area (BSA), ideal body weight (IBW), lean body weight (LBW), and fat free mass (FFM). Correlations between AUC0-9 of CyA and these parameters were also analyzed with a linear regression model. The rank order of the correlation coefficient was C0 > C2 (C0; r=0.6273, C2; r=0.5562). The linear regression analyses between AUC0-9 of CyA and candidate parameters indicated their potential usefulness from the following rank order: IBW > FFM > HT > BSA > LBW > TBW > BMI > Age. In conclusion, after oral administration, C2 monitoring has a large variation and could be at high risk for overdosing. Therefore, after oral dosing of CyA, it was not considered to be a useful approach for single monitoring, but should rather be used with C0 monitoring. The regression analyses between AUC0-9 of CyA and anthropometric parameters indicated that IBW was potentially the superior predictor for dose adjustment of CyA in an empiric strategy using TBW (IBW; r=0.5181, TBW; r=0.3192); however, this finding seems to lack the pharmacokinetic rationale and thus warrants further basic and clinical

  3. Cremophor EL releases cyclosporin A adsorbed on blood cells and blood vessels, and increases apparent plasma concentration of cyclosporin A.

    PubMed

    Jin, Mingji; Shimada, Tsutomu; Yokogawa, Koichi; Nomura, Masaaki; Mizuhara, Yasuharu; Furukawa, Hiroyuki; Ishizaki, Junko; Miyamoto, Ken-Ichi

    2005-04-11

    We examined the influence of cremophor EL (crEL) on the disposition kinetics of CyA in rats. A dose of 10mg/kg of CyA in a volume of 750 microL containing 4.3, 16 or 30% concentration of crEL was intravenously administered over 1 min to rats. The values of distribution volume at the steady-state (Vd(ss)) and total clearance (CL(tot)) of CyA in the presence of increasing amounts of crEL were decreased to about 1/3-1/5 of those with 4.3% crEL, in a crEL concentration-dependent manner. The values of blood to plasma concentration ratio (RBP) and the apparent tissue to plasma concentration ratio (K(p,app)) of CyA with 30% crEL were both only about 1/2 of those of CyA with 4.3% crEL. Next, rats were intravenously given 30% crEL solution at 30 min after an intravenous administration of CyA (10 mg/kg) with 4.3% crEL. Subsequently, the blood and plasma concentrations of CyA rose significantly to 2.4 and 4.7 times those seen when i.v. 30% crEL was not given, respectively. In an in vitro study, we found that the uptake of CyA by red blood cells is inhibited by crEL, and that CyA adsorbed on the inner surface of blood vessels after the administration of CyA is released by crEL. The disposition kinetics of CyA is altered by i.v. administration in combination with the surfactant vehicle crEL, in a crEL concentration-dependent manner.

  4. Segmental pancreatic allograft survival in baboons treated with combined irradiation and cyclosporine: a preliminary report

    SciTech Connect

    du Toit, D.F.; Heydenrych, J.J.; Smit, B.; Louw, G.; Zuurmond, T.; Laker, L.; Els, D.; Weideman, A.; Wolfe-Coote, S.; van der Merwe, E.A.

    1985-04-01

    The present study was undertaken to evaluate the effectiveness of cyclosporine (CS) alone, total lymphoid irradiation (TLI) alone, and CS in combination with total body irradiation (TBI) in suppressing segmental pancreatic allograft rejection in totally pancreatectomized outbred chacma baboons. The administration of CS 25 mg/kg/day and 50 mg/ kg/day resulted in mean graft survival of 21.5 days and 24.5 days, respectively. CS 85 mg/kg/day resulted in median graft survival of 9 days. There was a wide daily fluctuation of CS serum trough levels exhibited between primates receiving the same oral dose. TBI in excess of 300 rads resulted in irreversible bone marrow suppression. Modest results were achieved in recipients of TBI-76 rads (38 x 2 rads), with median graft survival of 21 days, results not different from recipients treated with CS. TLI recipients of 600 rads (150 x 4 rads) resulted in median pancreatic graft survival of 16 days. TBI together with oral CS administration exhibited no synergistic or additive effect and a single peroperative donor-specific blood transfusion did not enhance pancreatic allograft survival in this model. However, of 10 primates receiving TBI 100 rads (50 x 2 rads) and CS 25 mg/kg/day administered orally indefinitely, four remained normoglycemic for more than 60 days. TBI 100 rads (50 x 2 rads) together with oral and parenteral CS resulted in necrotizing enterocolitis in four of six recipients.

  5. Administration of cyclosporine a (CyA) to rats from birth: increased mortality and NK activity

    SciTech Connect

    Clancy, J. Jr.; Tseng, G.; Kodali, S.; Love, S.

    1986-03-01

    Neonatal DA and LEW rats received 15, 7.5, and 3.75 mg/Kg of CyA or saline subcutaneously 3x each week for 1-12 weeks. In animals receiving 15 and 7.5 mg/Kg a significant (p<0.05-0.01) decrease in body weight was observed by 1 and 2 weeks, respectively. Most animals given 15 mg/Kg died by 4 weeks. Rats receiving 7.5 and 3.75 mg/Kg survived but weighed less (p<0.05) than controls at 2 and 5 weeks, respectively. Morphologically, rats receiving CyA exhibited decreased cellularity in their thymic cortex as well as medulla and spleen white pulp. In addition, there was also a significant decrease in total number of cells harvested from each organ. Rats receiving the 7.5 and 3.75 mg/Kg dose had 1.5-2x more LGLs in their peripheral blood (PBL) and spleen (SPL) then controls after 6-12 weeks. In addition, their PBL and spleen cells were 2-3x more effective than controls in causing /sup 51/Cr release from YAC-1 target cells. Also, SPL cells stained with propidium iodide from the 3.75 mg/Kg group demonstrated a 1.5-2x increase in cells within the S phase of their cell cycle by flow cytometry. Thus, prolonged administration of CyA may have selective enhancing effects on certain lymphoid compartments and subpopulations of neonatal rats as well as a selective toxic effects on neonatal rat development.

  6. The protective effect of vildagliptin in chronic experimental cyclosporine A-induced hepatotoxicity.

    PubMed

    El-Sherbeeny, Nagla A; Nader, Manar A

    2016-03-01

    The study examined the effect of dipeptidyl peptidase-4 (DPP-4) inhibitor, vildagliptin, in cyclosporine (CsA)-induced hepatotoxicity. Rats were divided into 4 groups treated for 28 days: control (vehicle), vildagliptin (10 mg/kg, orally), CsA (20 mg/kg, s.c.), and CsA-vildagliptin group. Liver function was assessed by measuring serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma glutamyltransferase (γGT), lactate dehydrogenase (LDH), and albumin, and histopathological changes of liver were examined. Oxidative stress markers were evaluated. Assessment of nuclear factor-kappa B (NF-κB) activity in hepatic nuclear extract, serum DPP-4, and expression of Bax and Bcl2 were also done. CsA-induced hepatotoxicity was evidenced by increase in serum levels of AST, ALT, and γGT; a decrease in serum albumin; and a significant alteration in hepatic architecture. Also, significant increase in thiobarbituric acid reactive substance (TBARS) and decrease in superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione (GSH) levels, increased expression Bax proteins with deceased expression of Bcl2, and increased hepatic activity of NF-κB and serum DPP-4 level were observed upon CsA treatment. Vildagliptin significantly improved all altered parameters induced by CsA administration. Vildagliptin has the potential to protect the liver against CsA-induced hepatotoxicity by reducing oxidative stress, DPP-4 activity, apoptosis, and inflammation.

  7. Cyclosporine A immunosuppression drives catastrophic squamous cell carcinoma through IL-22

    PubMed Central

    Mitsui, Hiroshi; Roudiani, Nazanin; Ovits, Channa; Bryan, Teddy; Oberyszyn, Tatiana M.; Tober, Kathleen L.; Gonzalez, Juana; Krueger, James G.; Felsen, Diane; Carucci, John A.

    2016-01-01

    Immune-suppressed organ transplant recipients (OTRs) can develop catastrophic squamous cell carcinoma (SCC), characterized by multiple primary tumors, extensive body surface area involvement, or metastases. There are currently no curative systemic therapies available. We previously showed that IL-22 enhances SCC proliferation. Herein, we examined links between cyclosporine (CSA), IL-22, and SCC in patients, cell lines, and mice with UV light–induced SCC. Eighteen of 114 OTRs developed catastrophic SCC, which was strongly associated with CSA treatment. We found that CSA drives T cell polarization toward IL-22–producing T22 cells, and CSA treatment increased IL-22 receptor in SCC cells. SCC tissue from OTRs showed increased expression of IL-22RA1. CSA potentiated rescue by IL-22 of serum-starved SCC cells; treatment of SCC cells with IL-22 and CSA increased both their migratory and invasive capacity. In a UV-induced model of SCC in SKH-1 immunocompetent mice, treatment with anti–IL-22 antibody reduced tumor number and tumor burden. We found that catastrophic SCC in OTRs is associated with CSA use, which may be acting by favoring T22 polarization. Since anti–IL-22 antibody administration decreased tumor number and tumor burden in vivo, blockade of the IL-22 axis may be developed as a viable therapeutic option for catastrophic SCC. PMID:27699266

  8. Calcineurin inhibitors cyclosporine A and tacrolimus induce vascular inflammation and endothelial activation through TLR4 signaling

    PubMed Central

    Rodrigues-Diez, Raquel; González-Guerrero, Cristian; Ocaña-Salceda, Carlos; Rodrigues-Diez, Raúl R.; Egido, Jesús; Ortiz, Alberto; Ruiz-Ortega, Marta; Ramos, Adrián M.

    2016-01-01

    The introduction of the calcineurin inhibitors (CNIs) cyclosporine and tacrolimus greatly reduced the rate of allograft rejection, although their chronic use is marred by a range of side effects, among them vascular toxicity. In transplant patients, it is proved that innate immunity promotes vascular injury triggered by ischemia-reperfusion damage, atherosclerosis and hypertension. We hypothesized that activation of the innate immunity and inflammation may contribute to CNI toxicity, therefore we investigated whether TLR4 mediates toxic responses of CNIs in the vasculature. Cyclosporine and tacrolimus increased the production of proinflammatory cytokines and endothelial activation markers in cultured murine endothelial and vascular smooth muscle cells as well as in ex vivo cultures of murine aortas. CNI-induced proinflammatory events were prevented by pharmacological inhibition of TLR4. Moreover, CNIs were unable to induce inflammation and endothelial activation in aortas from TLR4−/− mice. CNI-induced cytokine and adhesion molecules synthesis in endothelial cells occurred even in the absence of calcineurin, although its expression was required for maximal effect through upregulation of TLR4 signaling. CNI-induced TLR4 activity increased O2−/ROS production and NF-κB-regulated synthesis of proinflammatory factors in cultured as well as aortic endothelial and VSMCs. These data provide new insight into the mechanisms associated with CNI vascular inflammation. PMID:27295076

  9. Sirolimus and tacrolimus rather than cyclosporine A cause bone loss in healthy adult male rats.

    PubMed

    Rubert, Mercedes; Montero, Mercedes; Guede, David; Caeiro, Jose-Ramón; Martín-Fernández, Marta; Díaz-Curiel, Manuel; de la Piedra, Concepción

    2015-06-01

    The aim of this work was to study the effects of cyclosporine (CsA), tacrolimus (FK-506), and rapamycin (RAPA) on bone mass, femoral microstructure, femoral biomechanical properties, and bone remodeling in healthy adult male rats. Forty-eight 5-month-old male Wistar rats were used. CsA (2 mg/kg/day), FK-506 (3 mg/kg/day), RAPA (1.25 mg/kg/day), or water (0.5 ml/rat/day, control group) were administered orally for 3 months. After sacrifice, mean values of immunosuppressants in blood were: CsA (670.4 ng/ml), FK-506 (19.2 ng/ml), and RAPA (4.8 ng/ml). Levels of biochemical parameters were normal in all groups. Femoral BMD was decreased in FK-506 and RAPA groups and lumbar BMD in FK-506 group. Trabecular volume fraction (BV/TV) decreased only in FK-506 group. RAPA and CsA affected femoral cortical structure, but FK-506 did not. FK-506 produced an increase in bone remodeling, and CsA a decrease. FK-506 group showed a decrease in biomechanical parameters relative to all groups. RAPA group showed a decrease in ultimate stress vs control group, and CsA group presented an increase in biomechanical parameters versus control group. We found that administration of both RAPA and FK-506 as monotherapy for healthy rats produced osteopenia. CsA treatment only produces slight damages in the cortical zone of the femur.

  10. Cyclosporine: A Historical Perspective on Its Role in the Treatment of Noninfectious Uveitis.

    PubMed

    Smith, Wendy M

    2017-03-13

    The history of cyclosporine and uveitis is intertwined with the development of experimental autoimmune uveitis (EAU) animal models and the understanding that T lymphocytes play a major role in the pathogenesis of uveitis. The early studies of CsA in uveitis also demonstrated the power of collaborative efforts in translational research. Dr. Robert Nussenblatt and his colleagues were the first to show that CsA can inhibit EAU. Over many years after the initial CsA experiments, Dr. Nussenblatt's group as well as others continued to study CsA under experimental conditions as well as in clinical trials with human patients. The data and observations from these studies significantly advanced our knowledge of uveitis pathophysiology and demonstrated the value of well-designed masked, controlled treatment trials in uveitis. Dr. Nussenblatt and his collaborators delved into the most significant adverse effect of CsA, renal toxicity, and helped elucidate the pathophysiology of renal injury. They explored adjunctive treatments to improve the efficacy and decrease the toxicity of CsA and also studied other members of the cyclosporine family. Among the immunosuppressives used to treat ocular inflammation, CsA was the first, and remains the only medication comprehensively studied under both experimental and clinical conditions.

  11. Cyclosporine A: Novel concepts in its role in drug-induced gingival overgrowth

    PubMed Central

    Ponnaiyan, Deepa; Jegadeesan, Visakan

    2015-01-01

    Cyclosporine is a selective immunosuppressant that has a variety of applications in medical practice. Like phenytoin and the calcium channel blockers, the drug is associated with gingival overgrowth. This review considers the pharmacokinetics, pharmacodynamics, and unwanted effects of cyclosporine, in particular the action of the drug on the gingival tissues. In addition, elucidates the current concepts in mechanisms of cyclosporine-induced gingival overgrowth. Clinical and cell culture studies suggest that the mechanism of gingival overgrowth is a result of the interaction between the drug and its metabolites with susceptible gingival fibroblasts. Plaque-induced gingival inflammation appears to enhance this interaction. However, understanding of the pathogenesis of gingival overgrowth is incomplete at best. Hence, it would be pertinent to identify and explore possible risk factors relating to both prevalence and severity of drug-induced gingival overgrowth. Newer molecular approaches are needed to clearly establish the pathogenesis of gingival overgrowth and to provide novel information for the design of future preventive and therapeutic modalities. PMID:26759584

  12. Beyond cyclosporine: a systematic review of limited sampling strategies for other immunosuppressants.

    PubMed

    Ting, Lillian S L; Villeneuve, Eric; Ensom, Mary H H

    2006-06-01

    Therapeutic drug monitoring has gained much attention in the management of immunosuppressive therapy. Area under the plasma drug concentration-time curve (AUC) is the pharmacokinetic (PK) parameter most commonly used to assess total exposure to a drug. However, estimation of AUC requires multiple blood samples throughout the dosing period, which is often inconvenient and expensive. Limited sampling strategies (LSSs) are therefore developed to estimate AUC and other PK parameters accurately and precisely while minimizing the number of blood samples needed. This greatly reduces costs, labor and inconvenience for both patients and clinical staff. In the therapeutic management of solid organ transplantation, LSSs for cyclosporine are commonplace and have been extensively reviewed. Thus, this systematic review paper focuses on other immunosuppressive agents and categorizes the 24 pertinent citations according to the U.S. Preventive Services Task Force rating scale. Thirteen articles (3 level I, 1 level II-1, 2 level II-2, and 7 level III) involved LSSs for mycophenolate, 7 citations (1 level I and 6 level III) for tacrolimus (TAC), and 3 citations (all level III) for other drugs (sirolimus) or multiple drugs. The 2 main approaches to establishing LSSs, multiple regression and Bayesian analyses, are also reviewed. Important elements to consider for future LSS studies, including proper validation of LSSs, convenient sampling times, and application of LSSs to the appropriate patient population and drug formulation are discussed. Limited sampling strategies are a useful tool to help clinicians make decisions on drug therapy. However, patients' pathophysiology, environmental and genetic factors, and pharmacologic response to therapy, in conjunction with PK profiling tools such as LSSs, should be considered collectively for optimal therapy management.

  13. Immunohistochemical Localization of Epithelial Mesenchymal Transition Markers in Cyclosporine A Induced Gingival Overgrowth

    PubMed Central

    Arora, Hitesh; Madapusi, Balaji Thodur; Ramamurti, Anjana; Narasimhan, Malathi; Periasamy, Soundararajan

    2016-01-01

    Introduction Cyclosporine, an immunosuppressive agent used in the management of renal transplant patients is known to produce Drug Induced Gingival Overgrowth (DIGO) as a side effect. Several mechanisms have been elucidated to understand the pathogenesis of DIGO. Recently, epithelial mesenchymal transition has been proposed as a mechanism underlying fibrosis of various organs. Aim The aim of the study was to investigate if Epithelial Mesenchymal Transition (EMT) operates in Cyclosporine induced gingival overgrowth. Materials and Methods The study involved obtaining gingival tissue samples from healthy individuals (n=17) and subjects who exhibited cyclosporine induced gingival overgrowth (n=18). Presence and distribution of E-Cadherin, S100 A4 and alpha smooth muscle actin (α-SMA) was assessed using immunohistochemistry and cell types involved in their expression were determined. The number of α– SMA positive fibroblasts were counted in the samples. Results In control group, there was no loss of E-Cadherin and a pronounced staining was seen in the all layers of the epithelium in all the samples analysed (100%). S100 A4 staining was noted in langerhans cells, fibroblasts, endothelial cells and endothelial lined blood capillaries in Connective Tissue (CT) of all the samples (100%) while α - SMA staining was seen only on the endothelial lined blood capillaries in all the samples (100%). However in DIGO, there was positive staining of E-Cadherin only in the basal and suprabasal layers of the epithelium in all the samples (100%). Moreover there was focal loss of E-Cadherin in the epithelium in eight out of 18 samples (44%). A break in the continuity of the basement membrane was noted in three out of 18 samples (16%) on H & E staining. Conclusion Based on the analysis of differential staining of the markers, it can be concluded that EMT could be one of the mechanistic pathways underlying the pathogenesis of DIGO. PMID:27656563

  14. Effect of cyclosporin A treatment on the production of antibody in insulin-dependent (type I) diabetic patients.

    PubMed Central

    Boitard, C; Feutren, G; Castano, L; Debray-Sachs, M; Assan, R; Hors, J; Bach, J F

    1987-01-01

    Anti-islet cell and anti-insulin antibody production was studies over a 12-mo period in 82 recently diagnosed diabetics randomly receiving either cyclosporin or placebo. Cyclosporin had only minimal effects on the production of anti-islet cell antibodies whether directed to islet cytoplasmic (immunofluorescence) or membrane (cytotoxicity assay) antigens even in patients undergoing remission. These data suggest that these antibodies do not play a major role in the pathogenesis of the disease particularly since their (irregular) presence is not predictive of the clinical response to cyclosporin. Conversely, cyclosporin completely suppressed the synthesis of antibodies elicited by exogenous insulin irrespective of the insulin doses received, and decreased the autoantibody production against thyroid antigens, indicating that cyclosporin has variable effects on antibody production against various antigens. PMID:3316278

  15. Synchrotron radiation small- and wide- angle scattering study of dispergation of Equoral, a novel drug delivery system with cyclosporine A.

    PubMed

    Uhríková, D; Andrýsek, T; Funari, S S; Balgavý, P

    2004-08-01

    Equoral oral solution is a novel drug delivery system for cyclosporine consisting mainly of non-ionic surfactants, polyglycerol esters and polyoxyethylated fatty acids aggregates, and gives microdispersions in the aqueous enviroment. To simulate dispergation, Equoral was mixed with varying amounts of water. Changes in the structure of the prepared aggregates were studied using synchrotron x-ray small- and wide-angle scattering. A lamellar phase is the most probable structure, arising spontaneously after dispergation of Equoral in the region of 30-70 wt% H2O.

  16. Polyomavirus BK Replication in De Novo Kidney Transplant Patients Receiving Tacrolimus or Cyclosporine: A Prospective, Randomized, Multicenter Study

    PubMed Central

    Hirsch, H H; Vincenti, F; Friman, S; Tuncer, M; Citterio, F; Wiecek, A; Scheuermann, E H; Klinger, M; Russ, G; Pescovitz, M D; Prestele, H

    2013-01-01

    Polyomavirus BK (BKV)-associated nephropathy causes premature kidney transplant (KT) failure. BKV viruria and viremia are biomarkers of disease progression, but associated risk factors are controversial. A total of 682 KT patients receiving basiliximab, mycophenolic acid (MPA), corticosteroids were randomized 1:1 to cyclosporine (CsA) or tacrolimus (Tac). Risk factors were analyzed in 629 (92.2%) patients having at least 2 BKV measurements until month 12 posttransplant. Univariate analysis associated CsA-MPA with lower rates of viremia than Tac-MPA at month 6 (10.6% vs. 16.3%, p = 0.048) and 12 (4.8% vs. 12.1%, p = 0.004) and lower plasma BKV loads at month 12 (3.9 vs. 5.1 log10 copies/mL; p = 0.028). In multivariate models, CsA-MPA remained associated with less viremia than Tac-MPA at month 6 (OR 0.60; 95% CI 0.36–0.99) and month 12 (OR 0.33; 95% CI 0.16–0.68). Viremia at month 6 was also independently associated with higher steroid exposure until month 3 (OR 1.19 per 1 g), and with male gender (OR 2.49) and recipient age (OR 1.14 per 10 years) at month 12. The data suggest a dynamic risk factor evolution of BKV viremia consisting of higher corticosteroids until month 3, Tac-MPA compared to CsA-MPA at month 6 and Tac-MPA, older age, male gender at month 12 posttransplant. PMID:23137180

  17. Estimation of Abbreviated Cyclosporine A Area under the Concentration-Time Curve in Allogenic Stem Cell Transplantation after Oral Administration.

    PubMed

    Eljebari, Hanene; Ben Fradj, Nadia; Salouage, Issam; Gaies, Emna; Trabelsi, Sameh; Jebabli, Nadia; Lakhal, Mohamed; Ben Othman, Tarek; Kouz, Anis

    2012-01-01

    Measurements of Cyclosporine (CsA) systemic exposure permit its dose adjustment in allogenic stem cell transplantation recipients to prevent graft-versus-host disease. CsA LSSs were developed and validated from 60 ASCT patients via multiple linear regressions. All whole-blood samples were analyzed by fluorescence polarization immunoassay (FPIA-Axym). The 10 models that have used CsA concentrations at a single time point did not have a good fit with AUC(0-12) (R(2) < 0.90). C(2) and C(4) were the time points that correlated best with AUC(0-12 h), R(2) were respectively 0.848, and 0.897. The LSS equation with the best predictive performance (bias, precision and number of samples) utilized three sampling concentrations was AUC(0-12 h) = 0.607 + 1.569 × C(0.5) + 2.098 × C(2) + 3.603 × C(4) (R(2) = 0.943). Optimal LSSs equations which limited to those utilizing three timed concentrations taken within 4 hours post-dose developed from ASCT recipient's patients yielded a low bias <5% ranged from 1.27% to 2.68% and good precision <15% ranged from 9.60% and 11.02%. We propose an LSS model with equation AUC(0-12 h) = 0.82 + 2.766 × C(2) + 3.409 × C(4) for a practical reason. Bias and precision for this model are respectively 2.68% and 11.02%.

  18. Estimation of Abbreviated Cyclosporine A Area under the Concentration-Time Curve in Allogenic Stem Cell Transplantation after Oral Administration

    PubMed Central

    ELjebari, Hanene; Ben Fradj, Nadia; Salouage, Issam; Gaies, Emna; Trabelsi, Sameh; Jebabli, Nadia; Lakhal, Mohamed; Ben Othman, Tarek; Kouz, Anis

    2012-01-01

    Measurements of Cyclosporine (CsA) systemic exposure permit its dose adjustment in allogenic stem cell transplantation recipients to prevent graft-versus-host disease. CsA LSSs were developed and validated from 60 ASCT patients via multiple linear regressions. All whole-blood samples were analyzed by fluorescence polarization immunoassay (FPIA-Axym). The 10 models that have used CsA concentrations at a single time point did not have a good fit with AUC0–12 (R2 < 0.90). C2 and C4 were the time points that correlated best with AUC0–12 h, R2 were respectively 0.848, and 0.897. The LSS equation with the best predictive performance (bias, precision and number of samples) utilized three sampling concentrations was AUC0–12 h = 0.607 + 1.569 × C0.5 + 2.098 × C2 + 3.603 × C4 (R2 = 0.943). Optimal LSSs equations which limited to those utilizing three timed concentrations taken within 4 hours post-dose developed from ASCT recipient's patients yielded a low bias <5% ranged from 1.27% to 2.68% and good precision <15% ranged from 9.60% and 11.02%. We propose an LSS model with equation AUC0–12 h = 0.82 + 2.766 × C2 + 3.409 × C4 for a practical reason. Bias and precision for this model are respectively 2.68% and 11.02%. PMID:22132303

  19. Combination of glucosamine improved therapeutic effect of low-dose cyclosporin A in patients with atopic dermatitis: a pilot study.

    PubMed

    Kwon, Hyok-Bu; Ahn, Byung-Jin; Choi, Yunseok; Jin, Sang Y; Cheong, Kyung A; Lee, Joongyub; Lee, Ai-Young

    2013-03-01

    Both glucosamine and cyclosporin have been reported to show immunomodulatory effect with inhibition of each different key transcription factor for cytokine gene expression and T-cell function. The overall purpose of this pilot study was to assess the feasibility of the combination of cyclosporin with glucosamine for the treatment of patients with atopic dermatitis. Twelve patients more than 12 years old who required systemic cyclosporin were included in the study. Two of them dropped out due to violation of medication schedule. The single (S) and combination (C) regimens were crossed over every 2 weeks without a washout period between the cross-over for 6 months. Five patients were randomly assigned to the S regimen first (SC sequence), whereas the other five were given the C first (CS sequence). The change of SCORAD index was analyzed as the primary efficacy end-point by general linear model and piecewise linear mixed model. The SCORAD index was reduced with both SC and CS sequence regimens. In particular, index reduction with the C was more than that associated with S regimen; this difference increased as time lapsed. The glucosamine combination was predicted to cause an additive decrease in the mean percent change of the SCORAD index (~6%), with decreasing interleukin (IL)-4 and IL-5 cytokine levels but without increasing treatment-related adverse events. This study suggests that the C would produce better clinical outcomes than the S regimen in patients with atopic dermatitis, although confirmatory clinical trials are warranted to determine the effect of combination.

  20. Oxidative capacities of cardiac and skeletal muscles of heart transplant recipients: mitochondrial effects of cyclosporin-A and its vehicle Cremophor-EL.

    PubMed

    N' Guessan, Benoit Banga; Sanchez, Hervé; Zoll, Joffrey; Ribera, Florence; Dufour, Stéphane; Lampert, Eliane; Kindo, Michel; Geny, Bernard; Ventura-Clapier, Renée; Mettauer, Bertrand

    2014-04-01

    Chronic immunosuppressive treatment was suspected to alter maximal muscle oxidative capacity (Vmax ) of heart transplant recipients, leading to a limitation of their exercise tolerance. It remains undefined whether the mitochondrial respiratory chain (MRC) of right ventricle (RV) and vastus lateralis (VL) muscles were altered by immunosuppressants and/or their vehicles. Vmax was measured polarographically in saponin-skinned fibres of RV and VL biopsies of patients and compared with Vmax of healthy VL and myocardium. Effects of increasing concentrations (1-10-100 μM) of Sandimmune(®) , its vehicle, Cyclosporine (CsA) in ethanol (EtOH), or EtOH alone were tested. The vehicle's effects on MRC complexes were investigated using specific substrates and inhibitors. Ten months after grafting, Vmax of RV and VL of immunosuppressed patients were similar to their Vmax at time of transplantation and to that of control tissues. In Vitro, Sandimmune(®) significantly decreased Vmax while CsA in EtOH or EtOH exerted small and similar effects. Effects of the vehicle were higher than (RV) or identical to (VL) those of Sandimmune(®) . The sites of action of the vehicle on MRC were located on complexes I and IV. While unchanged under chronic immunosuppressive therapy, Vmax of RV and VL muscles was depressed by acute exposure to intravenous Sandimmune(®) in vitro, an effect attributed to its vehicle by inhibition of complexes I and IV of the MRC. This work provides an in vitro proof of a toxic effect on the mitochondria respiratory chain of the vehicle used in the intravenous formulation of Sandimmune(®) but with no clinical consequences in chronically immunosuppressed patients.

  1. Controlled release of cyclosporine A self-nanoemulsifying systems from osmotic pump tablets: near zero-order release and pharmacokinetics in dogs.

    PubMed

    Zhang, Xi; Yi, Yueneng; Qi, Jianping; Lu, Yi; Tian, Zhiqiang; Xie, Yunchang; Yuan, Hailong; Wu, Wei

    2013-08-16

    It is very important to enhance the absorption simultaneously while designing controlled release delivery systems for poorly water-soluble and poorly permeable drugs (BCS IV). In this study, controlled release of cyclosporine (CyA) was achieved by the osmotic release strategy taking advantage of the absorption-enhancing capacity of self-nanoemulsifying drug delivery systems (SNEDDSs). The liquid SNEDDS consisting of Labrafil M 1944CS, Transcutol P and Cremophor EL was absorbed by the osmotic tablet core excipients (sucrose, lactose monohydrate, polyethylene oxide, and partly pregelatinized starch) and then transformed into osmotic tablets. Near zero-order release could be achieved for CyA-loaded nanoemulsions reconstituted from the SNEDDS. In general, the influencing factor study indicated that the release rate increased with increase of inner osmotic pressure, ratio of osmotic agent to suspending agent, content of pore-forming agent, and size of release orifice, whereas the thickness of the membrane impeded the release of CyA nanoemulsion. Pharmacokinetic study showed steady blood CyA profiles with prolonged Tmax and MRT, and significantly reduced Cmax for self-nanoemulsifying osmotic pump tablet (SNEOPT) in comparison with highly fluctuating profiles of the core tablet and Sandimmune Neoral(®). However, similar oral bioavailability was observed for either controlled release or non-controlled release formulations. It was concluded that simultaneous controlling on CyA release and absorption-enhancing had been achieved by a combination of osmotic tablet and SNEDDS.

  2. The Usefulness of Determining Neutrophil Gelatinase-Associated Lipocalin Concentration Excreted in the Urine in the Evaluation of Cyclosporine A Nephrotoxicity in Children with Nephrotic Syndrome

    PubMed Central

    2016-01-01

    Introduction. The use of cyclosporine (CsA) in the treatment of nephrotic syndrome (NS) contributed to a significant reduction in the amount of corticosteroids used in therapy and its cumulative side effects. One of the major drawbacks of CsA therapy is its nephrotoxicity. Prolonged CsA treatment protocols require sensitive, easily available, and simple to measure biomarkers of nephrotoxicity. NGAL is an antibacterial peptide, excreted by cells of renal tubules in response to their toxic or inflammatory damage. Aim of the Study. The aim of this study was to assess the suitability of the NGAL concentration in the urine as a potential biomarker of the CsA nephrotoxicity. Material and Methods. The study was performed on a group of 31 children with NS treated with CsA. The control group consisted of 23 children diagnosed with monosyptomatic enuresis. The relationship between NGAL excreted in urine and the time of CsA treatment, concentration of CsA in blood serum, and other biochemical parameters was assessed. Results. The study showed a statistically significant positive correlation between urine NGAL concentration and serum triglycerides concentration and no correlation between C0 CsA concentration and other observed parameters of NS. The duration of treatment had a statistically significant influence on the NGAL to creatinine ratio. Conclusions. NGAL cannot be used alone as a simple CsA nephrotoxicity marker during NS therapy. Statistically significant correlation between NGAL urine concentration and the time of CsA therapy indicates potential benefits of using this biomarker in the monitoring of nephrotoxicity in case of prolonged CsA therapy. PMID:28115789

  3. Synergistic effect of deoxyspergualin (DSP) and cyclosporin A (CsA) in the prevention of spontaneous autoimmune diabetes in BB rats

    PubMed Central

    DI MARCO, R; ZACCONE, P; MAGRO, G; GRASSO, S; LUNETTA, M; BARCELLINI, W; NICOLOSI, V M; MERONI, P L; NICOLETTI, F

    1996-01-01

    Dose-dependent side effects are frequently observed with immunosuppressive drugs of potential relevance for the immunotherapy of insulin-dependent diabetes mellitus (IDDM), such as CsA and DSP. If CsA and DSP acted synergistically in vivo, their combined use would allow using each compound at lower doses than those required when each drug is given in monotherapy. Consequently, dose-dependent side effects could be reduced and the therapeutic activity maintained or even enforced. Toward this end we studied the effects of combined treatment with CsA and DSP on the course of IDDM in the diabetes-prone (DP)-BB rat. The results show that two ‘low’ doses of CsA (2mg/kg) and DSP (1mg/kg) that are clinically ineffective in suppressing IDDM development in BB rats when administered alone under a prolonged prophylactic regimen (30–105 days old), may successfully prevent, but not cure, the disease when given contemporaneously under the same experimental conditions. The combined treatment was well tolerated, and no side effects were noticed. These data suggest that the combined use of CsA and DSP may deserve consideration for its possible application in the prevention/treatment of human IDDM and other autoimmune diseases. PMID:8706343

  4. Preliminary results of Tc-99m Sulfur Colloid (SC) and glucoheptonate (GHP) surveillance of renal allografts (AG) on cyclosporin a (CSA) maintenance

    SciTech Connect

    George, F.A.; Brown, W.W.; Malone, J.D.; Naidu, R.; Carney, K.; Garvin, P.J.

    1985-05-01

    The potential nephro-toxicity of CSA complicates the therapeutic management of AG with unexplained reduced function. Seven living related donor (LRD) and 23 cadaver (C) AG maintained, at least initially, on CSA and low dose prednisone and functioning for at least 2 months were studied during a 10 month period; mean serum creatinine (Scr) was 1.5 mg/dl for LRD and 2.35 mg/dl for C AG recipients. A total of 12 biopsies were performed in 10 recipients (2 LRD, 10 C AG) for unexplained SCr levels above 3.0 mg/dl. SC-GHP surveillance was carried out in each recipient between 4 and 15 times. SC exam was analyzed by temporal quantitation (as previously reported) and GHP exam was evaluated visually for AG perfusion, uptake and excretion of GHP. Biopsies were examined with H and E and immunofluorescent staining. Results were compared to therapeutic outcome, subsequent biopsy or AG nephrectomy. In summary, the comparison of SC-GHP exam and biopsy diagnosis in 12 instances for the differential diagnosis of CSA toxicity versus AR demonstrated correct assessment by the SC-GHP exam 12/12. AG biopsy diagnosis was correct in 8/12 instances.

  5. The Advantage of Cyclosporine A and Methotrexate Rotational Therapy in Long-Term Systemic Treatment for Chronic Plaque Psoriasis in a Real World Practice

    PubMed Central

    Choi, Chong Won; Kim, Bo Ri; Ohn, Jungyoon

    2017-01-01

    Background Psoriasis is a chronic inflammatory disease. In the treatment of psoriasis, cyclosporine is commonly prescribed systemic agents. However, long-term use of cyclosporine is not recommended because of side effects such as nephrotoxicity or hypertension. Objective To ascertain the improved safety of rotational therapy using cyclosporine and methotrexate, we investigated the frequency of abnormal results in laboratory test after long term rotational therapy using cyclosporine and methotrexate. Methods From January 2009 to June 2014, patients who were treated with cyclosporine or methotrexate were enrolled. The clinical data and usage of medications were reviewed. Laboratory tests were conducted before starting the treatment and regularly follow-up. The occurrences of any laboratory abnormalities during the treatments were investigated. Results A total of 21 psoriatic patients were enrolled. The mean of medication period and cumulative dose of cyclosporine and methotrexate were 497.81±512.06 days and 115.68±184.34 g in cyclosporine and 264.19±264.71 days and 448.71±448.63 mg in methotrexate. Laboratory abnormalities were found in total two patients after rotational therapy: two patients (9.5%) in aspartate aminotransferase/alanine aminotransferase and one patient (4.8%) in uric acid. No laboratory abnormalities were found in renal function test. Conclusion We found that the rotational approaches using cyclosporine and methotrexate reduced the possibility of the development of nephrotoxicity. In addition to other advantage such as quick switching from one agent to another, the rotational therapy using cyclosporine and methotrexate can minimize the adverse events during the systemic treatment of chronic plaque psoriasis. PMID:28223747

  6. Cyclosporin A sensitivity of the NF-kappa B site of the IL2R alpha promoter in untransformed murine T cells.

    PubMed Central

    McCaffrey, P G; Kim, P K; Valge-Archer, V E; Sen, R; Rao, A

    1994-01-01

    We have investigated the characteristics of IL2R alpha gene induction in untransformed murine T cells. Induction of IL2R alpha mRNA by TCR/CD3 ligands in a murine T cell clone and in short-term splenic T cell cultures was inhibited by protein synthesis inhibitors and by CsA. This result was contrary to previous observations in JURKAT T leukemia cells and human peripheral blood T cells, suggesting a difference in the mechanisms of IL2R alpha gene induction in these different cell types. The CsA sensitivity of IL2R alpha mRNA induction represented a direct effect on the TCR/CD3 response, and was not due to CsA-sensitive release of the lymphokines IL2 or tumour necrosis factor alpha (TNF alpha) and consequent lymphokine-mediated induction of IL2R alpha mRNA. The NF-kappa B site of the IL2R alpha promoter was essential for gene induction through the TCR/CD3 complex, and the induction of reporter plasmids containing multimers of this site was significantly inhibited by CsA. Northern blotting analysis indicated that while the p65 subunit of NF-kappa B was constitutively expressed and not appreciably induced upon T cell activation, mRNA for the p105 precursor of p50 NF-kappa B was induced in response to TCR/CD3 stimulation and this induction was sensitive to CsA. Electrophoretic mobility shift assays and antiserum against the p50 subunit of NF-kappa B indicated that p50 was a component of the inducible nuclear complex that bound to the IL2R alpha kappa B site. Appearance of the kB-binding proteins was insensitive to CsA at early times after activation (approximately 15 min), but was partially sensitive to CsA at later times. Based on these results, we propose that the NF-kappa B site of the IL2R alpha promoter mediates at least part of the CsA sensitivity of IL2R alpha gene induction in untransformed T cells, possibly because de novo synthesis of p105 NF-kappa B is required for sustained IL2R alpha expression. Images PMID:8029023

  7. Role of reactive oxygen species in the signalling cascade of cyclosporine A-mediated up-regulation of eNOS in vascular endothelial cells

    PubMed Central

    López-Ongil, S; Hernández-Perera, O; Navarro-Antolín, J; Pérez de Lema, G; Rodríguez-Puyol, M; Lamas, S; Rodríguez-Puyol, D

    1998-01-01

    Cyclosporine A (CsA) increases eNOS mRNA expression in bovine cultured aortic endothelial cells (BAEC). As some effects of CsA may be mediated by reactive oxygen species (ROS), present experiments were devoted to test the hypothesis that the CsA-induced eNOS up-regulation could be dependent on an increased synthesis of ROS.CsA induced a dose-dependent increase of ROS synthesis, with the two fluorescent probes used, DHR123 (CsA 1 μM: 305±7% over control) and H2DCFDA (CsA 1 μM: 178±6% over control).Two ROS generating systems, xanthine plus xanthine oxidase (XXO) and glucose oxidase (GO), increased the expression of eNOS mRNA in BAEC, an effect which was maximal after 8 h of incubation (XXO: 168±21% of control values. GO: 208±18% of control values). The ROS-dependent increased eNOS mRNA expression was followed by an increase in eNOS activity.The effect of CsA on eNOS mRNA expression was abrogated by catalase, and superoxide dismutase (SOD). In contrast, the antioxidant PDTC augmented eNOS mRNA expression, both in basal conditions and in the presence of CsA.The potential participation of the transcription factor AP-1 was explored. Electrophoretic mobility shift assays were consistent with an increase in AP-1 DNA-binding activity in BAEC treated with CsA or glucose oxidase.The present results support a role for ROS, particularly superoxide anion and hydrogen peroxide, as mediators of the CsA-induced eNOS mRNA up-regulation. Furthermore, they situate ROS as potential regulators of gene expression in endothelial cells, both in physiological and pathophysiological situations. PMID:9647467

  8. Human coronavirus NL63 replication is cyclophilin A-dependent and inhibited by non-immunosuppressive cyclosporine A-derivatives including Alisporivir.

    PubMed

    Carbajo-Lozoya, Javier; Ma-Lauer, Yue; Malešević, Miroslav; Theuerkorn, Martin; Kahlert, Viktoria; Prell, Erik; von Brunn, Brigitte; Muth, Doreen; Baumert, Thomas F; Drosten, Christian; Fischer, Gunter; von Brunn, Albrecht

    2014-05-12

    Until recently, there were no effective drugs available blocking coronavirus (CoV) infection in humans and animals. We have shown before that CsA and FK506 inhibit coronavirus replication (Carbajo-Lozoya, J., Müller, M.A., Kallies, S., Thiel, V., Drosten, C., von Brunn, A. Replication of human coronaviruses SARS-CoV, HCoV-NL63 and HCoV-229E is inhibited by the drug FK506. Virus Res. 2012; Pfefferle, S., Schöpf, J., Kögl, M., Friedel, C., Müller, M.A., Stellberger, T., von Dall'Armi, E., Herzog, P., Kallies, S., Niemeyer, D., Ditt, V., Kuri, T., Züst, R., Schwarz, F., Zimmer, R., Steffen, I., Weber, F., Thiel, V., Herrler, G., Thiel, H.-J., Schwegmann-Weßels, C., Pöhlmann, S., Haas, J., Drosten, C. and von Brunn, A. The SARS-Coronavirus-host interactome: identification of cyclophilins as target for pan-Coronavirus inhibitors. PLoS Pathog., 2011). Here we demonstrate that CsD Alisporivir, NIM811 as well as novel non-immunosuppressive derivatives of CsA and FK506 strongly inhibit the growth of human coronavirus HCoV-NL63 at low micromolar, non-cytotoxic concentrations in cell culture. We show by qPCR analysis that virus replication is diminished up to four orders of magnitude to background levels. Knockdown of the cellular Cyclophilin A (CypA/PPIA) gene in Caco-2 cells prevents replication of HCoV-NL63, suggesting that CypA is required for virus replication. Collectively, our results uncover Cyclophilin A as a host target for CoV infection and provide new strategies for urgently needed therapeutic approaches.

  9. The effect of prostaglandin E1 analog misoprostol on chronic cyclosporin nephrotoxicity.

    PubMed

    John, E G; Fornell, L C; Radhakrishnan, J; Anutrakulchai, S; Jonasson, O

    1993-11-01

    Cyclosporin A has markedly improved graft survival in transplant patients but its side effects, such as renal toxicity and hypertension, pose management problems in transplant recipients. This toxicity has been attributed to prostaglandin inhibition. Concurrent administration of misoprostol (a prostaglandin E1 analog) prevents chronic cyclosporin A-induced nephrotoxicity but not hypertension in rats.

  10. An endoplasmic reticulum-specific cyclophilin.

    PubMed Central

    Hasel, K W; Glass, J R; Godbout, M; Sutcliffe, J G

    1991-01-01

    Cyclophilin is a ubiquitously expressed cytosolic peptidyl-prolyl cis-trans isomerase that is inhibited by the immunosuppressive drug cyclosporin A. A degenerate oligonucleotide based on a conserved cyclophilin sequence was used to isolate cDNA clones representing a ubiquitously expressed mRNA from mice and humans. This mRNA encodes a novel 20-kDa protein, CPH2, that shares 64% sequence identity with cyclophilin. Bacterially expressed CPH2 binds cyclosporin A and is a cyclosporin A-inhibitable peptidyl-prolyl cis-trans isomerase. Cell fractionation of rat liver followed by Western blot (immunoblot) analysis indicated that CPH2 is not cytosolic but rather is located exclusively in the endoplasmic reticulum. These results suggest that cyclosporin A mediates its effect on cells through more than one cyclophilin and that cyclosporin A-induced misfolding of T-cell membrane proteins normally mediated by CPH2 plays a role in immunosuppression. Images PMID:1710767

  11. Pharmacokinetic role of P-glycoprotein in oral bioavailability and intestinal secretion of grepafloxacin in vivo.

    PubMed

    Yamaguchi, Hiroaki; Yano, Ikuko; Saito, Hideyuki; Inui, Ken-ichi

    2002-03-01

    The purpose of this study was to clarify the contribution of P-glycoprotein to the bioavailability and intestinal secretion of grepafloxacin and levofloxacin in vivo. Plasma concentrations of grepafloxacin and levofloxacin after intravenous and intraintestinal administration were increased by cyclosporin A, a P-glycoprotein inhibitor, in rats. The total body clearance and volume of distribution at steady state of grepafloxacin were significantly decreased to 60 and 63% of the corresponding control values by cyclosporin A. The apparent oral clearance of grepafloxacin was decreased to 33% of the control, and the bioavailability of grepafloxacin was increased to 95% by cyclosporin A from 53% in the controls. Intestinal clearance of grepafloxacin and levofloxacin were decreased to one-half and one-third of the control, respectively, and biliary clearance of grepafloxacin was also decreased to one-third with cyclosporin A in rats. Intestinal secretion of grepafloxacin in mdr1a/1b (-/-) mice, which lack mdr1-type P-glycoproteins, was significantly decreased compared with wild-type mice, although the biliary secretion was similar. Intestinal secretion of grepafloxacin in wild-type mice treated with cyclosporin A was comparable to those in mdr1a/1b (-/-) mice with or without cyclosporin A, indicating that cyclosporin A completely inhibited P-glycoprotein-mediated intestinal transport of grepafloxacin. In conclusion, our results indicated that P-glycoprotein mediated the intestinal secretion of grepafloxacin and limited the bioavailability of this drug in vivo.

  12. Morphometric and ultrastructural analysis of the effect of bromocriptine and cyclosporine on the vasospastic femoral artery of rats

    PubMed Central

    Tokmak, Mehmet; Başocak, Kahan; Canaz, Hüseyin; Canaz, Gökhan; İplikçioğlu, Celal

    2015-01-01

    Vasospasm is the main causes of mortality and morbidity in patiens with subarachnoid hemorrhage (SAH). The arterial narrowing mechanism that develops after SAH is not yet fully understood but many studies showed that hypotension, neurogenic reflexes, clots in the subarachnoidal space, spasmogenic agents, humoral and celluler immunity play a role in the etiology. In this study we investigate the effects of Bromocriptine and Cyclosporine A in vasospasm secondary to SAH on rat femoral artery from ultrastructural and morphometric perspectives. 120 male Sprague-Dawley rats divided into 12 groups: Vasospasm (V), control (K), surgical control (CK) groups, vasospasm+Bromocriptine and/or Cyclosporine-A groups (VCyA, VBr, VBr+CyA), Bromocriptine and/or Cyclosporine-A control groups (CK, BK, Br+CyAK), Bromocriptine and/or Cyclosporine-A surgical control groups (BCK, CyCK, Br+CyACK). In order to create SAH model, 0, 1 cm3 blood injected into silastic sheath wrapped rat femoral artery. Bromocriptine (2 mg/kg/d) and Cyclosporine A (10 mg/kg/d) combinations applied to control, surgical control and vasospastic models. Light microscopy, transmission electron microscopy and scanning electron microscopy used during this study. Statistical evaluation of the morphometric measurement data concerning vascular wall thickness and luminal cross-sectional areas of all groups were performed using Mann-Whitney U, Wilcoxon-signed rank, and Student-t tests. Cyclosporine A, whose effects in the prevention of vasospasm have been demonstrated in previous studies. In this study we discovered that Bromocriptine demonstrated strong effects similar to Cyclosporine-A. Bromocriptine and Cyclosporine A markedly prevent the development of chronic morphologic vasospasm following SAH. The combined use of both drugs does not change this preventive effect. PMID:26770311

  13. Long-term responses and outcomes following immunosuppressive therapy in large granular lymphocyte leukemia-associated pure red cell aplasia: a Nationwide Cohort Study in Japan for the PRCA Collaborative Study Group.

    PubMed

    Fujishima, Naohito; Sawada, Ken-ichi; Hirokawa, Makoto; Oshimi, Kazuo; Sugimoto, Koichi; Matsuda, Akira; Teramura, Masanao; Karasawa, Masamitsu; Arai, Ayako; Yonemura, Yuji; Nakao, Shinji; Urabe, Akio; Omine, Mitsuhiro; Ozawa, Keiya

    2008-10-01

    Large granular lymphocyte leukemia-associated pure red cell aplasia accounts for a significant portion of secondary pure red cell aplasia cases. However, because of its rarity, long-term responses and relapse rates after immunosuppressive therapy are largely unknown. We conducted a nationwide survey in Japan and collected 185 evaluable patients. Fourteen patients with large granular lymphocyte leukemia-associated pure red cell aplasia were evaluated. Cyclophosphamide, cyclosporine A and prednisolone produced remissions in 6/8, 1/4 and 0/2 patients respectively. Seven and 5 patients were maintained on cyclophosphamide or cyclosporine A respectively. Two patients relapsed after stopping cyclophosphamide, and 2 patients relapsed during maintenance therapy with cyclosporine A. The median relapse-free survival in the cyclophosphamide - and the cyclosporine A groups was 53 and 123 months respectively. Large granular lymphocyte leukemia-associated pure red cell aplasia showed a good response to either cyclophosphamide or cyclosporine A. Most patients continued to receive maintenance therapy and it remains uncertain whether cyclophosphamide or cyclosporine A can induce a maintenance-free hematologic response in large granular lymphocyte leukemia-associated pure red cell aplasia.

  14. Secretory mechanisms of grepafloxacin and levofloxacin in the human intestinal cell line caco-2.

    PubMed

    Yamaguchi, H; Yano, I; Hashimoto, Y; Inui, K I

    2000-10-01

    Grepafloxacin and levofloxacin transport by Caco-2 cell monolayers was examined to characterize the intestinal behavior of these quinolones. The levels of transcellular transport of [(14)C]grepafloxacin and [(14)C]levofloxacin from the basolateral to the apical side were greater than those in the opposite direction. The unidirectional transport was inhibited by the presence of excess unlabeled quinolones, accompanied by increased accumulation. The inhibitory effects of cyclosporin A plus grepafloxacin on basolateral-to-apical transcellular transport and cellular accumulation of [(14)C]grepafloxacin were comparable to those of cyclosporin A alone, indicating that the transport of grepafloxacin across the apical membrane was mainly mediated by P-glycoprotein. On the other hand, basolateral-to-apical transcellular transport of [(14)C]levofloxacin in the presence of cyclosporin A was decreased by unlabeled levofloxacin, grepafloxacin, and enoxacin, accompanied by significantly increased cellular accumulation. The organic cation cimetidine, organic anion p-aminohippurate, and the multidrug resistance-related protein (MRP) modulator probenecid did not affect the transcellular transport of [(14)C]grepafloxacin or [(14)C]levofloxacin in the presence of cyclosporin A. The basolateral-to-apical transcellular transport of levofloxacin in the presence of cyclosporin A showed concentration-dependent saturation with an apparent Michaelis constant of 5.6 mM. In conclusion, these results suggested that basolateral-to-apical flux of quinolones was mediated by P-glycoprotein and a specific transport system distinct from organic cation and anion transporters and MRP.

  15. Tamoxifen inhibits mitochondrial oxidative stress damage induced by copper orthophenanthroline.

    PubMed

    Buelna-Chontal, Mabel; Hernández-Esquivel, Luz; Correa, Francisco; Díaz-Ruiz, Jorge Luis; Chávez, Edmundo

    2016-12-01

    In this work, we studied the effect of tamoxifen and cyclosporin A on mitochondrial permeability transition caused by addition of the thiol-oxidizing pair Cu(2+) -orthophenanthroline. The findings indicate that tamoxifen and cyclosporin A circumvent the oxidative membrane damage manifested by matrix Ca(2+) release, mitochondrial swelling, and transmembrane electrical gradient collapse. Furthermore, it was found that tamoxifen and cyclosporin A prevent the generation of TBARs promoted by Cu(2+) -orthophenanthroline, as well as the inactivation of the mitochondrial enzyme aconitase and disruption of mDNA. Electrophoretic analysis was unable to demonstrate a cross-linking reaction between membrane proteins. Yet, it was found that Cu(2+) -orthophenanthroline induced the generation of reactive oxygen species. It is thus plausible that membrane leakiness is due to an oxidative stress injury.

  16. Successful treatment of severe myasthenia gravis developed after allogeneic hematopoietic stem cell transplantation with plasma exchange and rituximab.

    PubMed

    Unal, Sule; Sag, Erdal; Kuskonmaz, Baris; Kesici, Selman; Bayrakci, Benan; Ayvaz, Deniz C; Tezcan, Ilhan; Yalnızoglu, Dilek; Uckan, Duygu

    2014-05-01

    Myasthenia gravis is among the rare complications after allogeneic hematopoietic stem cell transplantation and is usually associated with chronic GVHD. Herein, we report a 2-year and 10 months of age female with Griscelli syndrome, who developed severe myasthenia gravis at post-transplant +22nd month and required respiratory support with mechanical ventilation. She was unresponsive to cyclosporine A, methylprednisolone, intravenous immunoglobulin, and mycophenolate mofetil and the symptoms could only be controlled after plasma exchange and subsequent use of rituximab, in addition to cyclosporine A and mycophenolate mofetil maintenance. She is currently asymptomatic on the 6th month of follow-up.

  17. Interaction of Cyclic Peptides and Depsipeptides with Calmodulin

    DTIC Science & Technology

    1990-04-10

    by block number) FIELD GROUP SUB-GROUP cairodulin, cyclosporin A, gramicidin S, valinomycin, enniatin-B, microcystin -LR, phosphodiesterase 19...investigated the ability of other cyclic peptides: microcystin -LR (MLR) and depsipeptides, valinomycin (VLM) and enniatin-B (ENB), to bind dansylated CaM

  18. 21 CFR 520.522 - Cyclosporine.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Cyclosporine. 520.522 Section 520.522 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS ORAL DOSAGE FORM NEW ANIMAL DRUGS § 520.522 Cyclosporine. (a)...

  19. 21 CFR 520.522 - Cyclosporine.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Cyclosporine. 520.522 Section 520.522 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS ORAL DOSAGE FORM NEW ANIMAL DRUGS § 520.522 Cyclosporine. (a)...

  20. 21 CFR 520.522 - Cyclosporine.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Cyclosporine. 520.522 Section 520.522 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS ORAL DOSAGE FORM NEW ANIMAL DRUGS § 520.522 Cyclosporine. (a)...

  1. 21 CFR 520.522 - Cyclosporine.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Cyclosporine. 520.522 Section 520.522 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS ORAL DOSAGE FORM NEW ANIMAL DRUGS § 520.522 Cyclosporine. (a)...

  2. Cyclosporine treatment of perianal gland adenoma concurrent with benign prostatic hyperplasia in a dog

    PubMed Central

    Park, Chul; Yoo, Jong-Hyun; Kim, Ha-Jung; Lim, Chae-Young; Kim, Ju-Won; Lee, So-Young; Kim, Jung-Hyun; Jang, Jae-Im; Park, Hee-Myung

    2010-01-01

    A 13-year-old, intact male, mixed-breed dog was evaluated for multiple intradermal nodules around the anus. The nodules were diagnosed as perianal gland adenoma based on histopathologic examination. After therapy with cyclosporin A for 5 wk, the perianal masses were moderately shrunken. The dog’s condition has remained stable over 6 mo. PMID:21286331

  3. A Fascinating Journey into History: Exploration of the World of Isonitriles En Route to Complex Amides

    PubMed Central

    Wilson, Rebecca M.; Stockdill, Jennifer L.; Wu, Xiangyang; Li, Xuechen; Vadola, Paul A.; Park, Peter K.; Danishefsky, Samuel J.

    2012-01-01

    We describe herein our recent explorations in the field of isonitrile chemistry. An array of broadly useful coupling methodologies has been developed for the formation of peptidyl and glycopeptidyl amide bonds. We further describe the application of these methods to the syntheses of complex systems, including the cyclic peptide cyclosporine A, constrained peptide systems, and heterocycles. PMID:22368033

  4. [Membranous aspects of kidney transplantation and reasons of transplant antioxidant defense].

    PubMed

    Berdichevskiĭ, B A; Tsvettskikh, V E; Zhmurov, V A; Kononov, S L; Berdichevskaia, E B; Zhuravleva, T D; Kiianiuk, N S; Razumiak, T V; Nedorezoniuk, S V

    2000-01-01

    Special membranological studies of the role of antioxidant emoxipine in combined pharmacological support of stable function of the transplanted kidney in 30 patients have shown that this drug restores phospholipid pool of red cell membranes and improves their physical properties. No significant fluctuations of cyclosporin A blood concentrations were registered. Emoxipine addition to combined therapy of patients with transplanted kidney is recommended.

  5. Reoxygenation-induced mitochondrial damage is caused by the Ca2+-dependent mitochondrial inner membrane permeability transition.

    PubMed

    Tanaka, T; Hakoda, S; Takeyama, N

    1998-07-01

    Anoxia/reoxygenation injury of isolated rat liver mitochondria was investigated. During anoxia of up to 60 min, the membrane potential was largely preserved and mitochondrial swelling was not observed. Reoxygenation of anoxic mitochondria rapidly caused swelling, cyclosporin A-sensitive Ca2+ efflux, [14C]sucrose trapping, and loss of the membrane potential along with increased generation of reactive oxygen intermediates (ROI). Although pretreatment with catalase and superoxide dismutase completely abolished reoxygenation-induced generation of ROI, mitochondrial damage was not prevented, as indicated by swelling, loss of the membrane potential, a decrease of the ATP content, and cyclosporin A-sensitive Ca2+ efflux. However, addition of the immunosuppressant cyclosporin A or addition of ADP completely prevented the mitochondrial damage induced by reoxygenation. The same protective effect was noted when Ca2+ cycling was prevented, either by chelating Ca2+ with EGTA or by inhibiting Ca2+ reuptake with ruthenium red. These findings indicate that mitochondrial anoxia/reoxygenation injury is caused by the cyclosporin A-sensitive and Ca2+-dependent membrane permeability transition. In contrast, reoxygenation injury does not appear to be triggered by the enhanced production of ROI.

  6. Effect of CIPC and intervention of Ca(2+)-regulated factors on CaN, cbl-b and p-AKT expression in neurons.

    PubMed

    Jiang, Yong-Mei; Zhang, Hui; Yin, Lin

    2011-05-10

    CaN induces the apoptosis in neurons, but the influence of CIPC and the intervention of pretreament with Ca(2+)-regulated factors, such as nimodipine, MK801 and cyclosporine A, on CaN expression is not clear. We also do not know whether cbl-b takes part in the induction of ischemia or induces an expression change of cbl-b in CIPC. So we will discuss the effect of CIPC, pretreatment with nimodipine, MK801 and cyclosporine A on the expression of the CaN, cbl-b and p-AKT in the hippocampus neurons. In our study, we established rat models including sham, ischemia, CIPC, nimodipine, MK801 and cyclosporine A. The neurological deficit scores were processed. The right hippocampus was removed and stained with TTC, and the volume of cerebral infarction was calculated. The apoptotic neurons were detected by TUNEL staining. The expressions of CaN, cbl-b and p-AKT at the protein level were examined by Western blotting, and the transcription of cbl-b by RT-PCR, respectively. The results showed that the neurological deficit scores, the volume of the cerebral infarction, the numbers of the apoptotic neurons, the protein expression of CaN, cbl-b and the transcription of cbl-b were the highest in the ischemia and MK801 groups, there were no difference between the two groups(P>0.05); these factors in CIPC group were all lower than those in the ischemia group(P<0.05); and much lower in the nimodipine and cyclosporine A group than those in the CIPC group (among them, the volume of the cerebral infarction in the nimodipine and cyclosporine A groups P<0.01, the expression of CaN in nimodipine group P<0.01, others were P<0.05), but no significant difference existed between the nimodipine and cyclosporine A groups(P>0.05). The expression of p-AKT was the lowest in the ischemia and MK801 groups, and there was no difference between the two groups (P>0.05), This factor was higher in CIPC group than that in the ischemia group (P<0.05); it was the highest in the nimodipine and cyclosporine A

  7. Cyclosporine therapy during pregnancy in a patient with β-thalassemia major and autoimmune haemolytic anemia: a case report and review of the literature

    PubMed Central

    Agapidou, A; Vlachaki, E; Theodoridis, T; Economou, M; Perifanis, V

    2013-01-01

    Recent advances in the management of hemoglobinopathies offer an improved potential for safe pregnancy with favourable outcome in patients with β-thalassemia major. Autoimmune diseases that are common in women at reproductive age might be fulminant and hardly manageable in pregnant women with thalassemia. Thus immunosuppressant drugs like cyclosporine A could be necessary in order to maintain good maternal and foetal health. We present a case report of a 35-year-old woman with β-thalassemia major, splenectomy, autoimmune hemolytic anemia and insulin treated diabetes mellitus who was treated with cyclosporine A during her pregnancy, and delivered a healthy male infant. First line therapy with steroids was ineffective, due to deregulation of diabetes mellitus. PMID:23935353

  8. Predicting the Toxicity of Adjuvant Breast Cancer Drug Combination Therapy

    DTIC Science & Technology

    2012-09-01

    hepatic metabo- lism based on the ratio of total liver:intestinal CYP3A, the major cytochrome P450 enzyme sub-family responsible for lapatinib...combination with cyclosporin A. Alternatively, the increase in exposure was likely more resultant of competitive inhibition of cytochrome P450 enzymes by...sex on the clearance of cytochrome P450 3A substrates. Clin Pharmacokinet 44(1):33–60 27. Bischoff KB, Dedrick RL, Zaharko DS (1970) Preliminary

  9. Acquired generalized anhidrosis: review of the literature and report of a case with lymphocytic hidradenitis and sialadenitis successfully treated with cyclosporine.

    PubMed

    Fujita, Kumi; Hatta, Kazuhiro

    2013-01-01

    We report a case of acquired generalized anhidrosis successfully treated with cyclosporine. A skin biopsy showed T cell infiltration around the sweat glands and labial biopsy revealed lymphoplasmacytic infiltration around the minor salivary gland, suggesting an underlying autoimmune disease such as Sjögren's syndrome. Administration of cyclosporine markedly improved the patient's condition and sympathetic skin response; thus cyclosporine may be effective for treating anhidrosis in patients with autoimmune disorders.

  10. Effects of mTOR and calcineurin inhibitors combined therapy in Epstein-Barr virus positive and negative Burkitt lymphoma cells.

    PubMed

    Wowro, Sylvia J; Schmitt, Katharina R L; Tong, Giang; Berger, Felix; Schubert, Stephan

    2016-01-01

    Post-transplant lymphoproliferative disorder is a severe complication in solid organ transplant recipients, which is highly associated with Epstein-Barr virus infection in pediatric patients and occasionally presents as Burkitt- or Burkitt-like lymphoma. The mammalian target of rapamycin (mTOR) pathway has been described as a possible antitumor target whose inhibition may influence lymphoma development and proliferation after pediatric transplantation. We treated Epstein-Barr virus positive (Raji and Daudi) and negative (Ramos) human Burkitt lymphoma derived cells with mTOR inhibitor everolimus alone and in combination with clinically relevant immunosuppressive calcineurin inhibitors (tacrolimus or cyclosporin A). Cell proliferation, toxicity, and mitochondrial metabolic activity were analyzed. The effect on mTOR Complex 1 downstream targets p70 S6 kinase, eukaryotic initiation factor 4G, and S6 ribosomal protein activation was also investigated. We observed that treatment with everolimus alone significantly decreased Burkitt lymphoma cell proliferation and mitochondrial metabolic activity. Everolimus in combination with cyclosporin A had a stronger suppressive effect in Epstein-Barr virus negative but not in Epstein-Barr virus positive cells. In contrast, tacrolimus completely abolished the everolimus-mediated suppressive effects. Moreover, we showed a significant decrease in activation of mTOR Complex 1 downstream targets after treatment with everolimus that was attenuated when combined with tacrolimus, but not with cyclosporin A. For the first time we showed the competitive effect between everolimus and tacrolimus when used as combination therapy on Burkitt lymphoma derived cells. Thus, according to our in vitro data, the combination of calcineurin inhibitor cyclosporin A with everolimus is preferred to the combination of tacrolimus and everolimus.

  11. Adoptive Immunotherapy Combined with Hematopoietic Cell Transplantation as a Therapeutic Treatment of Prostate Cancer

    DTIC Science & Technology

    2009-07-01

    irradiation (TBI) followed by postgrafting immunosuppression with mycophenolate mofetil ( MMF ) and cyclosporin A (CSP) for 28 and 35 days respectively...immunosuppression of MMF and CSP (11). We now believe this problem was due to our replacing an aged 4 meV linear accelerator (Linac) with a newer 6 meV...lymphocyte reaction MMF mycophenolate mofetil PBL peripheral blood lymphocytes PBMC peripheral blood mononuclear cells PBS phosphate buffered

  12. Examination of serum class I antigen in liver-transplanted rats.

    PubMed Central

    Sumimoto, R; Shinomiya, T

    1991-01-01

    We examined the appearance of donor (DA) type class I antigen in the serum of rats that had received isogeneic (DA----DA) or allogeneic (DA----PVG, DA----BN, DA----LEW) liver transplants with or without cyclosporin A treatment, using two-site enzyme immunoassay. We also tested the serum titre of class I antigen in the normal DA rats with either 70% hepatectomy or cyclosporin A treatment, in order to clarify the relationship between the fluctuation in the serum titre of class I antigen in the recipient and the outcome of the transplanted liver graft. The suppression of liver graft rejection by cyclosporin A treatment significantly lowered the serum level of donor liver-derived class I antigen as compared with that of the recipient without cyclosporin A for a certain period. However, there was almost no correlation between the intensity of rejection of the liver graft and the serum level type class I among these allogeneic rejection and non-rejection liver transplantation combinations. The amount of donor-type class I antigen in the recipient's serum is dependent on whether the grafted liver is severely damaged following partial hepatectomy, whether the liver has associated biliary complications or ischaemic damage, or whether the liver has had absolute residual parenchymal cell volume or function following liver rejection. Our results suggest that the appearance of donor type class I antigen following liver transplantation is dependent on many factors, and therefore the titre of serum class 1 antigen may not always be a decisive indicator of liver graft rejection. PMID:2070555

  13. A Model for the Interfacial Kinetics of Phospholipase D Activity on Long-Chain Lipids

    DTIC Science & Technology

    2013-07-01

    activity. MATERIALS AND METHODS Materials We purchased cesium acetate, cyclosporin A, and 1-octadecanethiol from Sigma Aldrich (St. Louis, MO... cesium chloride (CsCl) from International Biotechnologies (New Haven, CT); and calcium chloride (CaCl2), pentane, and hexadecane from Fluka. gA was...sn- glycero-3-phosphate ( sodium salt) (DiPhyPA). PLD from cabbage (EC 3.1.4.4) was obtained from Sigma Aldrich. Storage and final concentration of PLD

  14. Inhibition of the immune response to experimental fresh osteoarticular allografts

    SciTech Connect

    Rodrigo, J.J.; Schnaser, A.M.; Reynolds, H.M. Jr.; Biggart, J.M. 3d.; Leathers, M.W.; Chism, S.E.; Thorson, E.; Grotz, T.; Yang, Q.M. )

    1989-06-01

    The immune response to osteoarticular allografts is capable of destroying the cartilage--a tissue that has antigens on its cells identical to those on the bone and marrow cells. Osteoarticular allografts of the distal femur were performed in rats using various methods to attempt to temporarily inhibit the antibody response. The temporary systemic immunosuppressant regimens investigated were cyclophosphamide, azathioprine and prednisolone, cyclosporine A, and total lymphoid irradiation. The most successful appeared to be cyclosporine A, but significant side effects were observed. To specifically inhibit the immune response in the allograft antigens without systemically inhibiting the entire immune system, passive enhancement and preadministration of donor blood were tried. Neither was as effective as coating the donor bone with biodegradable cements, a method previously found to be successful. Cyclosporine A was investigated in dogs in a preliminary study of medial compartmental knee allografts and was found to be successful in inhibiting the antibody response and in producing a more successful graft; however, some significant side effects were similarly observed.

  15. In vivo association of immunophenotyped macrophages expressing CD163 with PDGF-B in gingival overgrowth-induced by three different categories of medications

    PubMed Central

    Almahrog, Amina J.; Radwan, Lobna R.S.; El-Zehery, Rehab R.; Mourad, Mohamed I.; Grawish, Mohammed E.

    2016-01-01

    Aims This study was carried out to identify and outline the degree of relationship between immunophenotyped macrophages expressing CD163 and PDGF-B in cyclosporine-A, phenytoin, and nifedipine-induced gingival overgrowth. Methods Eighty adult male albino rats were selected and divided into four equal groups. Group I received no treatment. Rats of groups II, III, and IV were administered cyclosporine-A, phenytoin, and nifedipine, respectively. Routine tissue processing was carried out for staining with CD163 and PDGF-B. The results of this study were analyzed statistically. Results Group I exhibited score 0 gingival overgrowth while group II yielded score 3 with blunt and bulbous gingival crests. Rats of group III showed score 2 with knife edge and group IV revealed less pronounced gingival overgrowth and mostly the gingival crest was knife edge. Group II had the highest mean value for CD163 while group I showed the lowest value. In addition, group II had the highest mean value for PDGF-B while group I showed the lowest value. Statistically, there was an overall significant difference between the studied groups as well as between each two groups. Conclusion Strong association exists between immunophenotyped macrophages expressing CD163 and PDGF-B in GO induced by these medications. In addition, CD163 and PDGF-B upregulated in cyclosporine-A-induced GO compared to phenytoin and nifedipine medications. PMID:26937363

  16. Possible involvement of P-glycoprotein in the biliary excretion of grepafloxacin.

    PubMed

    Zhao, Ying Lan; Cai, Shao Hui; Wang, Li; Kitaichi, Kiyoyuki; Tatsumi, Yasuaki; Nadai, Masayuki; Yoshizumi, Hideo; Takagi, Kenji; Takagi, Kenzo; Hasegawa, Takaaki

    2002-03-01

    1. In the present study, we have examined the effects of the quinolones norfloxacin (NFLX), enoxacin (ENX), ofloxacin (OFLX), tosufloxacin (TFLX), lomefloxacin (LFLX), sparfloxacin (SPFX) and grepafloxacin (GPFX) on the efflux of doxorubicin from mouse leukaemia P388/ADR cells expressing P-glycoprotein. The relationship between their partition coefficients (hydrophobicity) and effluxing potencies was also elucidated. 2. Both TFLX and SPFX strongly increased the intracellular accumulation of doxorubicin (5 micromol/L) in P388/ADR cells, but had no effect on P388/S cells not expressing P-glycoprotein. The rank of order of the potency of the quinolones (TFLX > SPFX > GPFX > NFLX) was not related directly to their hydrophobicity. These results suggest that some quinolones can reverse anticancer drug resistance. 3. Because GPFX is more highly excreted into the bile than other known quinolones, the effects of doxorubicin (10 mg/kg) or the well-known inhibitors of P-glycoprotein, namely cyclosporine A (10 mg/kg) and erythromycin (100 mg/kg), on the biliary excretion of GPFX at steady state was studied in rats. 4. Doxorubicin, cyclosporine A and erythromycin significantly decreased the biliary clearance of GPFX. Cyclosporine A and erythromycin had a much stronger inhibitory effect on the biliary excretion of GPFX than doxorubicin. These results suggest the possibility that GPFX is, at least in part, excreted into the bile by a P-glycoprotein-mediated transport mechanism.

  17. Active pharmaceutical ingredient (api) from an estuarine fungus, Microdochium nivale (Fr.).

    PubMed

    Bhosale, S H; Patil, K B; Parameswaran, P S; Naik, C G; Jagtap, T G

    2011-09-01

    Various marine habitats sustain variety of bio-sources of ecological and biotech potentials. Pharmaceutical potential compound Cyclosporine A was reported from marine fungus Microdochium nivale associated with Porteresia coarctata, a marine salt marsh grass from mangrove environment distributed along the Central West Coast (CWC) of India. This study involves association of M. nivale with P. coarctata plant, fermentation conditions, purification of Cyclosporine A, chemical characterization etc. Its antifungal inhibition and MIC (Minimum inhibitory concentration) against Aspergillus strains (A. niger, A. japonicus, A. fresenii), yeasts and dermatophytes (Candida sp., Cryptococcus neoformans, Trichophyton mentagrophytes, T. tonsurans, T. violaceum, Microsporium gypsum and Fusarium sp.) were evaluated. However, the MIC against A. japonicus, C. neoformans, Candida sp. and T. tonsurans were confirmed to be as low as 12.5-25 mg disc(-1). The antifungal properties of Cyclosporine A against Aspergillus species, yeast and dermatophytes revealed that CyclosporineAwould be a potential compound for life threatening diseases caused by above fungi in both human and animals. Furthermore, we have reported herewith another source of Cyclosporin Aderived from filamentous fungus, M. nivale. occurring in marine environment.

  18. The treatment of peripheral nerve injuries using irradiated allografts and temporary host immunosuppression (in a rat model)

    SciTech Connect

    Easterling, K.J.; Trumble, T.E. )

    1990-10-01

    Irradiation of allografts prior to transplantation and host immunosuppression with cyclosporin-A were studied separately and in combination as means of lessening the rejection of transplanted peripheral nerve tissue. Lewis and Brown Norway rats were used in the animal model, as they differ at both major and minor histocompatibility loci. Sciatic nerve grafts (2.5 cm) were used and the animals were followed for 16 weeks after nerve grafting. The outcome was studied by functional measurements (sensory testing, gait analysis, joint flexion contracture, and muscle weight), as well as by measurements of biochemical and histologic parameters (hydroxyproline concentration and axon counts, respectively). Sensory testing was not reliable because of crossover innervation by the saphenous nerve. Evaluation by standard gait-testing techniques was found to be unsatisfactory. However, the allografted animals receiving cyclosporin-A had significantly smaller flexion contractures, compared to the allografted animals without immunosuppression (17 degrees +/- 12 degrees vs. 44 degrees +/- 13 degrees and 51 degrees +/- 13 degrees, p less than 0.005). Allografted animals receiving short-term cyclosporin-A had contractures that were not significantly different from those seen in isografted control animals (17 degrees +/- 12 degrees vs. 22 degrees +/- 15 degrees, NS). Muscle hydroxyproline concentration analysis revealed a lower hydroxyproline concentration among the allografted groups that received irradiated allografts, compared to groups receiving nonirradiated allogeneic grafts. The studies of muscle hydroxyproline concentration and muscle weight both showed substantial reinnervation, even in allografted animals without pretreatment of the grafts or immunosuppression of the recipient animal.

  19. Ethanol immunosuppression in vitro

    SciTech Connect

    Kaplan, D.R.

    1986-03-01

    Ethanol in concentrations equivalent to levels achieved by the ingestion of moderate to large amounts of alcoholic beverages has been shown to inhibit mitogen and anti-CD3 stimulated human T lymphocyte proliferation. This inhibition was monophasic suggesting that ethanol affected a single limiting component of T cell proliferation. In experiments designed to test the effect of ethanol on various aspects of proliferation, it was demonstrated that ethanol inhibited the capacity of exogenously supplied interleukin 2 to stimulate proliferation of T cells that had previously acquired interleukin 2 receptors in a monophasic, dose-dependent manner. Moreover, there was no suppression of interleukin 2 production or interleukin 2 receptor acquisition. Thus, ethanol was shown to mediate immunosuppression by a mechanism specific to one component of proliferation. Additive inhibition of T cell proliferation was seen with ethanol plus cyclosporin A which inhibits interleukin 2 production. The level of inhibition with 250 ng/ml cyclosporin A alone was equivalent to the level seen with 62 ng/ml cyclosporin A plus 20 mM (94 mg%) ethanol. Ethanol also suppressed an immune effector mechanism. NK cytotoxicity was depressed in a monophasic, dose-dependent manner. Thus, ethanol might be considered as a possible adjunct in immunosuppressive therapy.

  20. Cyclosporine-loaded solid lipid nanoparticles (SLN): drug-lipid physicochemical interactions and characterization of drug incorporation.

    PubMed

    Müller, R H; Runge, S A; Ravelli, V; Thünemann, A F; Mehnert, W; Souto, E B

    2008-03-01

    Solid lipid nanoparticles (SLN) were produced loaded with cyclosporine A in order to develop an improved oral formulation. In this study, the particles were characterized with regard to the structure of the lipid particle matrix, being a determining factor for mode of drug incorporation and drug release. Differential scanning calorimetry (DSC) and wide-angle X-ray scattering (WAXS) measurements were employed for the analysis of the polymorphic modifications and mode of drug incorporation. Particles were produced using Imwitor 900 as lipid matrix (the suspension consisted of 10% particles, 8% Imwitor 900, 2% cyclosporine A), 2.5% Tagat S, 0.5% sodium cholate and 87% water. DSC and WAXS were used to analyse bulk lipid, bulk drug, drug incorporated in the bulk and unloaded and drug-loaded SLN dispersions. The processing of the bulk lipid into nanoparticles was accompanied by a polymorphic transformation from the beta to the alpha-modification. After production, the drug-free SLN dispersions converted back to beta-modification, while the drug-loaded SLN stayed primarily in alpha-modification. After incorporation of cyclosporine A into SLN, the peptide lost its crystalline character. Based on WAXS data, it could be concluded that cyclosporine is molecularly dispersed in between the fatty acid chains of the liquid-crystalline alpha-modification fraction of the loaded SLN.

  1. Cyclophilin A is required for the replication of group M human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus SIV(CPZ)GAB but not group O HIV-1 or other primate immunodeficiency viruses.

    PubMed Central

    Braaten, D; Franke, E K; Luban, J

    1996-01-01

    The human immunodeficiency virus type 1 (HIV-1) Gag polyprotein binds to cyclophilin A and incorporates this cellular peptidyl prolyl-isomerase into virions. Disruption of cyclophilin A incorporation, either by gag mutations or by cyclosporine A, inhibits virion infectivity, indicating that cyclophilin A plays an essential role in the HIV-1 life cycle. Using assays for packaging of cyclophilin A into virions and for viral replication sensitivity to cyclosporine A, as well as information gleaned from the alignment of Gag residues encoded by representative viral isolates, we demonstrate that of the five lineages of primate immunodeficiency viruses, only HIV-1 requires cyclophilin A for replication. Cloned viral isolates from clades A, B, and D of HIV-1 group M, as well as a phylogenetically related isolate from chimpanzee, all require cyclophilin A for replication. In contrast, the replication of two outlier (group O) HIV-1 isolates is unaffected by concentrations of cyclosporine A which disrupt cyclophilin A incorporation into virions, indicating that these viruses are capable of replicating independently of cyclophilin A. These studies identify the first phenotypic difference between HIV-1 group M and group O and are consistent with phylogenetic studies suggesting that the two HIV-1 groups were introduced into human populations via separate zoonotic transmission events. PMID:8676442

  2. Involvement of mitochondrial permeability transition pore opening in 7-xylosyl-10-deacetylpaclitaxel-induced apoptosis.

    PubMed

    Jiang, Shougang; Zu, Yuangang; Wang, Zhuo; Zhang, Yu; Fu, Yuejie

    2011-07-01

    7-Xylosyl-10-deacetylpaclitaxel is a natural hydrophilic paclitaxel derivative. It has long been used in Chinese clinics to treat cancer. In order to further explore the underlying intracellular target of 7-xylosyl-10-deacetylpaclitaxel towards the PC-3 cell line, the ultra-structural morphology of mitochondria, the intracellular Ca (2+), the intracellular ATP, the intracellular hydrogen peroxide and pro-apoptotic Bax and Bcl-2 protein expression were measured. Additionally, the changes of mitochondrial morphology and membrane potential ( ΔΨm) were analyzed by atomic force microscopy (AFM) and flow cytometry, respectively. Our results suggest that the intracellular target of 7-xylosyl-10-deacetylpaclitaxel may be the mitochondrial permeability transition pore (mPTP). To further evaluate this hypothesis, we assessed the effect of a specific mPTP inhibitor (cyclosporine A) on the toxic action of 7-xylosyl-10-deacetylpaclitaxel. The 7-xylosyl-10-deacetylpaclitaxel-induced decrease in mitochondrial inner transmembrane potential (ΔΨm) was abolished by the addition of cyclosporine A (CsA) in PC-3 cells, indicating that 7-xylosyl-10-deacetylpaclitaxel may target mPTP. Furthermore, treatment with 7-xylosyl-10-deacetylpaclitaxel increased ROS levels in PC-3 cells. This effect was counteracted by 10 µM cyclosporine A. These data indicate that oxidative damage is involved in mPTP.

  3. Targeting glucosylceramide synthase induction of cell surface globotriaosylceramide (Gb3) in acquired cisplatin-resistance of lung cancer and malignant pleural mesothelioma cells

    SciTech Connect

    Tyler, Andreas; Johansson, Anders; Karlsson, Terese; Gudey, Shyam Kumar; Brännström, Thomas; Grankvist, Kjell; Behnam-Motlagh, Parviz

    2015-08-01

    Background: Acquired resistance to cisplatin treatment is a caveat when treating patients with non-small cell lung cancer (NSCLC) and malignant pleural mesothelioma (MPM). Ceramide increases in response to chemotherapy, leading to proliferation arrest and apoptosis. However, a tumour stress activation of glucosylceramide synthase (GCS) follows to eliminate ceramide by formation of glycosphingolipids (GSLs) such as globotriaosylceramide (Gb3), the functional receptor of verotoxin-1. Ceramide elimination enhances cell proliferation and apoptosis blockade, thus stimulating tumor progression. GSLs transactivate multidrug resistance 1/P-glycoprotein (MDR1) and multidrug resistance-associated protein 1 (MRP1) expression which further prevents ceramide accumulation and stimulates drug efflux. We investigated the expression of Gb3, MDR1 and MRP1 in NSCLC and MPM cells with acquired cisplatin resistance, and if GCS activity or MDR1 pump inhibitors would reduce their expression and reverse cisplatin-resistance. Methods: Cell surface expression of Gb3, MDR1 and MRP1 and intracellular expression of MDR1 and MRP1 was analyzed by flow cytometry and confocal microscopy on P31 MPM and H1299 NSCLC cells and subline cells with acquired cisplatin resistance. The effect of GCS inhibitor PPMP and MDR1 pump inhibitor cyclosporin A for 72 h on expression and cisplatin cytotoxicity was tested. Results: The cisplatin-resistant cells expressed increased cell surface Gb3. Cell surface Gb3 expression of resistant cells was annihilated by PPMP whereas cyclosporin A decreased Gb3 and MDR1 expression in H1299 cells. No decrease of MDR1 by PPMP was noted in using flow cytometry, whereas a decrease of MDR1 in H1299 and H1299res was indicated with confocal microscopy. No certain co-localization of Gb3 and MDR1 was noted. PPMP, but not cyclosporin A, potentiated cisplatin cytotoxicity in all cells. Conclusions: Cell surface Gb3 expression is a likely tumour biomarker for acquired cisplatin

  4. Stable Isotope Labeling Strategy for Protein-Ligand Binding Analysis in Multi-Component Protein Mixtures

    NASA Astrophysics Data System (ADS)

    DeArmond, Patrick D.; West, Graham M.; Huang, Hai-Tsang; Fitzgerald, Michael C.

    2011-03-01

    Described here is a stable isotope labeling protocol that can be used with a chemical modification- and mass spectrometry-based protein-ligand binding assay for detecting and quantifying both the direct and indirect binding events that result from protein-ligand binding interactions. The protocol utilizes an H{2/16}O2 and H{2/18}O2 labeling strategy to evaluate the chemical denaturant dependence of methionine oxidation in proteins both in the presence and absence of a target ligand. The differential denaturant dependence to the oxidation reactions performed in the presence and absence of ligand provides a measure of the protein stability changes that occur as a result of direct interactions of proteins with the target ligand and/or as a result of indirect interactions involving other protein-ligand interactions that are either induced or disrupted by the ligand. The described protocol utilizes the 18O/16O ratio in the oxidized protein samples to quantify the ligand-induced protein stability changes. The ratio is determined using the isotopic distributions observed for the methionine-containing peptides used for protein identification in the LC-MS-based proteomics readout. The strategy is applied to a multi-component protein mixture in this proof-of-principle experiment, which was designed to evaluate the technique's ability to detect and quantify the direct binding interaction between cyclosporin A and cyclophilin A and to detect the indirect binding interaction between cyclosporin A and calcineurin (i.e., the protein-protein interaction between cyclophilin A and calcineurin that is induced by cyclosporin A binding to cyclophilin A).

  5. The effect of the mitochondrial permeability transition pore on apoptosis in Eimeria tenella host cells.

    PubMed

    Xu, Zhi-Yong; Zheng, Ming-Xue; Zhang, Yan; Cui, Xiao-Zhen; Yang, Sha-Sha; Liu, Rui-Li; Li, Shan; Lv, Qiang-Hua; Zhao, Wen-Long; Bai, Rui

    2016-10-01

    Although the mitochondrial permeability transition pore (MPTP) is associated with cellular apoptosis and necrosis, its effect in host response to Eimeria infections is not well understood. In an effort to better understand the effect of MPTP on apoptosis in Eimeria tenella host cells, an MPTP inhibitor (cyclosporin A) was used to inhibit MPTP opening in vitro. Cecal epithelial cells from chick embryos, which were either treated or non-treated with cyclosporin A, were used as Eimeria tenella host cells. In addition, primary chick embryo cecum epithelial cell culture techniques and flow cytometry were used to detect the dynamic changes in MPTP opening, mitochondrial transmembrane potential, and cell apoptosis rate of Eimeria tenella host cells. Compared with the control group, cytometric techniques showed that untreated host cells exhibited a significantly higher (P < 0.01) degree of MPTP opening but lower (P < 0.01 or P < 0.05) mitochondrial transmembrane potential. Moreover, untreated group cells had less apoptosis (P < 0.01) at 4 h and more apoptosis (P < 0.05 or P < 0.01) at 24 to 120 h as compared with control group cells. After the application of cyclosporin A, the degree of MPTP opening in the treated group was significantly lower (P < 0.01) at 4 to 120 h compared to the untreated group, whereas the treated group had higher (P < 0.05 or P < 0.01) mitochondrial transmembrane potentials at 24 to 120 h. Flow cytometry assays also showed that there was less (P < 0.05 or P < 0.01) apoptosis after 24 h in the treated group than in the untreated group. Taken together, these observations indicate that MPTP is a key node that plays a predominant role in the mitochondrial apoptosis pathway in the host cell induced by Eimeria tenella.

  6. Regulatory T cell levels and cytokine production in active non-infectious uveitis: in-vitro effects of pharmacological treatment

    PubMed Central

    Molins, B; Mesquida, M; Lee, R W J; Llorenç, V; Pelegrín, L; Adán, A

    2015-01-01

    The aim of this study was to quantify the proportion of regulatory T cells (Treg) and cytokine expression by peripheral blood mononuclear cells (PBMCs) in patients with active non-infectious uveitis, and to evaluate the effect of in-vitro treatment with infliximab, dexamethasone and cyclosporin A on Treg levels and cytokine production in PBMCs from uveitis patients and healthy subjects. We included a group of 21 patients with active non-infectious uveitis and 18 age-matched healthy subjects. The proportion of forkhead box protein 3 (FoxP3)+ Treg cells and intracellular tumour necrosis factor (TNF)-α expression in CD4+ T cells was determined by flow cytometry. PBMCs were also either rested or activated with anti-CD3/anti-CD28 and cultured in the presence or absence of dexamethasone, cyclosporin A and infliximab. Supernatants of cultured PBMCs were collected and TNF-α, interleukin (IL)-10, IL-17 and interferon (IFN)-γ levels were measured by enzyme-linked immunosorbent assay (ELISA). No significant differences were observed in nTreg levels between uveitis patients and healthy subjects. However, PBMCs from uveitis patients produced significantly higher amounts of TNF-α and lower amounts of IL-10. Dexamethasone treatment in vitro significantly reduced FoxP3+ Treg levels in PBMCs from both healthy subjects and uveitis patients, and all tested drugs significantly reduced TNF-α production in PBMCs. Dexamethasone and cyclosporin A significantly reduced IL-17 and IFN-γ production in PBMCs and dexamethasone up-regulated IL-10 production in activated PBMCs from healthy subjects. Our results suggest that PBMCs from patients with uveitis express more TNF-α and less IL-10 than healthy subjects, and this is independent of FoxP3+ Treg levels. Treatment with infliximab, dexamethasone and cyclosporin A in vitro modulates cytokine production, but does not increase the proportion of FoxP3+ Treg cells. PMID:25354724

  7. Regulatory T cell levels and cytokine production in active non-infectious uveitis: in-vitro effects of pharmacological treatment.

    PubMed

    Molins, B; Mesquida, M; Lee, R W J; Llorenç, V; Pelegrín, L; Adán, A

    2015-03-01

    The aim of this study was to quantify the proportion of regulatory T cells (Treg ) and cytokine expression by peripheral blood mononuclear cells (PBMCs) in patients with active non-infectious uveitis, and to evaluate the effect of in-vitro treatment with infliximab, dexamethasone and cyclosporin A on Treg levels and cytokine production in PBMCs from uveitis patients and healthy subjects. We included a group of 21 patients with active non-infectious uveitis and 18 age-matched healthy subjects. The proportion of forkhead box protein 3 (FoxP3)(+) Treg cells and intracellular tumour necrosis factor (TNF)-α expression in CD4(+) T cells was determined by flow cytometry. PBMCs were also either rested or activated with anti-CD3/anti-CD28 and cultured in the presence or absence of dexamethasone, cyclosporin A and infliximab. Supernatants of cultured PBMCs were collected and TNF-α, interleukin (IL)-10, IL-17 and interferon (IFN)-γ levels were measured by enzyme-linked immunosorbent assay (ELISA). No significant differences were observed in nTreg levels between uveitis patients and healthy subjects. However, PBMCs from uveitis patients produced significantly higher amounts of TNF-α and lower amounts of IL-10. Dexamethasone treatment in vitro significantly reduced FoxP3(+) Treg levels in PBMCs from both healthy subjects and uveitis patients, and all tested drugs significantly reduced TNF-α production in PBMCs. Dexamethasone and cyclosporin A significantly reduced IL-17 and IFN-γ production in PBMCs and dexamethasone up-regulated IL-10 production in activated PBMCs from healthy subjects. Our results suggest that PBMCs from patients with uveitis express more TNF-α and less IL-10 than healthy subjects, and this is independent of FoxP3(+) Treg levels. Treatment with infliximab, dexamethasone and cyclosporin A in vitro modulates cytokine production, but does not increase the proportion of FoxP3(+) Treg cells.

  8. Immunomodulation Stimulates the Innervation of Engineered Tooth Organ

    PubMed Central

    Kökten, Tunay; Bécavin, Thibault; Keller, Laetitia; Weickert, Jean-Luc; Kuchler-Bopp, Sabine; Lesot, Hervé

    2014-01-01

    The sensory innervation of the dental mesenchyme is essential for tooth function and protection. Sensory innervation of the dental pulp is mediated by axons originating from the trigeminal ganglia and is strictly regulated in time. Teeth can develop from cultured re-associations between dissociated dental epithelial and mesenchymal cells from Embryonic Day 14 mouse molars, after implantation under the skin of adult ICR mice. In these conditions however, the innervation of the dental mesenchyme did not occur spontaneously. In order to go further with this question, complementary experimental approaches were designed. Cultured cell re-associations were implanted together with trigeminal ganglia for one or two weeks. Although axonal growth was regularly observed extending from the trigeminal ganglia to all around the forming teeth, the presence of axons in the dental mesenchyme was detected in less than 2.5% of samples after two weeks, demonstrating a specific impairment of their entering the dental mesenchyme. In clinical context, immunosuppressive therapy using cyclosporin A was found to accelerate the innervation of transplanted tissues. Indeed, when cultured cell re-associations and trigeminal ganglia were co-implanted in cyclosporin A-treated ICR mice, nerve fibers were detected in the dental pulp, even reaching odontoblasts after one week. However, cyclosporin A shows multiple effects, including direct ones on nerve growth. To test whether there may be a direct functional relationship between immunomodulation and innervation, cell re-associations and trigeminal ganglia were co-implanted in immunocompromised Nude mice. In these conditions as well, the innervation of the dental mesenchyme was observed already after one week of implantation, but axons reached the odontoblast layer after two weeks only. This study demonstrated that immunodepression per se does stimulate the innervation of the dental mesenchyme. PMID:24465840

  9. Evidence for a role of claudin 2 as a proximal tubular stress responsive paracellular water channel

    SciTech Connect

    Wilmes, Anja Aschauer, Lydia; Limonciel, Alice; Pfaller, Walter; Jennings, Paul

    2014-09-01

    Claudins are the major proteins of the tight junctions and the composition of claudin subtypes is decisive for the selective permeability of the paracellular route and thus tissue specific function. Their regulation is complex and subject to interference by several factors, including oxidative stress. Here we show that exposure of cultured human proximal tubule cells (RPTEC/TERT1) to the immunosuppressive drug cyclosporine A (CsA) induces an increase in transepithelial electrical resistance (TEER), a decrease in dome formation (on solid growth supports) and a decrease in water transport (on microporous growth supports). In addition, CsA induced a dramatic decrease in the mRNA for the pore forming claudins -2 and -10, and the main subunits of the Na{sup +}/K{sup +} ATPase. Knock down of claudin 2 by shRNA had no discernable effect on TEER or dome formation but severely attenuated apical to basolateral water reabsorption when cultured on microporous filters. Generation of an osmotic gradient in the basolateral compartment rescued water transport in claudin 2 knock down cells. Inhibition of Na{sup +}/K{sup +} ATPase with ouabain prevented dome formation in both cell types. Taken together these results provide strong evidence that dome formation is primarily due to transcellular water transport following a solute osmotic gradient. However, in RPTEC/TERT1 cells cultured on filters under iso-osmotic conditions, water transport is primarily paracellular, most likely due to local increases in osmolarity in the intercellular space. In conclusion, this study provides strong evidence that claudin 2 is involved in paracellular water transport and that claudin 2 expression is sensitive to compound induced cellular stress. - Highlights: • Cyclosporine A increased TEER and decreased water transport in RPTEC/TERT1 cells. • Claudins 2 and 10 were decreased in response to cyclosporine A. • Knock down of claudin 2 inhibited water transport in proximal tubular cells. • We

  10. Isoflurane induced cognitive impairment in aged rats through hippocampal calcineurin/NFAT signaling

    SciTech Connect

    Ni, Cheng; Li, Zhengqian; Qian, Min; Zhou, Yang; Wang, Jun; Guo, Xiangyang

    2015-05-15

    Calcineurin (CaN) over-activation constrains synaptic plasticity and memory formation. Upon CaN activation, NFAT imports into the nucleus and guides its downstream genes, which also affect neuronal and synaptic function. Aberrant CaN/NFAT signaling involves in neurotoxicity and cognitive impairment in neurological disorders such as Alzheimer's disease, but its role in postoperative cognitive dysfunction (POCD) remains uninvestigated. Inhaled anesthetic isoflurane facilitates the development of POCD, and the present study investigated the role of CaN/NFAT signaling in isoflurane induced cognitive impairment of aged rats, and the therapeutic effects of CaN inhibitor cyclosporine A (CsA). The results indicated that hippocampal CaN activity increased and peaked at 6 h after isoflurane exposure, and NFAT, especially NFATc4, imported into the nucleus following CaN activation. Furthermore, phamacological inhibition of CaN by CsA markedly attenuated isoflurane induced aberrant CaN/NFATc4 signaling in the hippocampus, and rescued relevant spatial learning and memory impairment of aged rats. Overall, the study suggests hippocampal CaN/NFAT signaling as the upstream mechanism of isoflurane induced cognitive impairment, and provides potential therapeutic target and possible treatment methods for POCD. - Highlights: • Isoflurane induces hippocampal calcineurin activation. • Isoflurane induces hippocampal NFAT, especially NFATc4, nuclear import. • Cyclosporine A attenuates isoflurane induced aberrant calcineurin/NFAT signaling. • Cyclosporine A rescues isoflurane induced cognitive impairment. • Calcineurin/NFAT signaling is the upstream mechanism of isoflurane induced synaptic dysfunction and cognitive impairment.

  11. Natural history and etiology of hyperuricemia following pediatric renal transplantation.

    PubMed

    Edvardsson, V O; Kaiser, B A; Polinsky, M S; Palmer, J A; Quien, R; Baluarte, H J

    1995-02-01

    A retrospective review was conducted to determine the incidence, etiology, natural history and complications of hyperuricemia after pediatric renal transplantation. Of 81 active transplant recipients aged 10.1 +/- 4.8 (mean +/- SD) years being followed by St. Christopher's Hospital for Children, 57 (70%) were males and 59 (73%) Caucasian. Their immunosuppression consisted of azathioprine, cyclosporine A and prednisone. Mean serum uric acid concentrations peaked at 6 months post transplantation (6.2 +/- 2.6 mg/dl), when 39% of the patients had hyperuricemia and 60% were receiving diuretics, and decreased thereafter. At 30 months, 23% of the patients had hyperuricemia and 17% required diuretics. When we compared 42 normouricemic (group A) with 24 hyperuricemic (group B) patients at 18 months post transplantation, we found that patients in group B were older (11.6 +/- 4.2 vs. 8.6 +/- 5.2 years, P = 0.01), had worse renal function (77 +/- 25 vs. 96 +/- 36 ml/min per 1.73 m2, P = 0.03) and required diuretics more frequently (63% vs. 21%, P = 0.001), but had identical blood levels of cyclosporine A (82 +/- 28 vs. 84 +/- 35 ng/ml, P = 0.78). A family history of gout did not affect the prevalence of hyperuricemia after transplantation. Asymptomatic hyperuricemia is common following pediatric renal transplantation and is more likely attributable to reduced renal function and diuretic therapy than to the known hyperuricemic effect of cyclosporine A. Of these variables, only diuretic therapy is readily controllable and should be closely regulated following pediatric renal transplantation.

  12. Modulation of Human Immune Response by Fungal Biocontrol Agents

    PubMed Central

    Konstantinovas, Cibele; de Oliveira Mendes, Tiago A.; Vannier-Santos, Marcos A.; Lima-Santos, Jane

    2017-01-01

    Although the vast majority of biological control agents is generally regarded as safe for humans and environment, the increased exposure of agriculture workers, and consumer population to fungal substances may affect the immune system. Those compounds may be associated with both intense stimulation, resulting in IgE-mediated allergy and immune downmodulation induced by molecules such as cyclosporin A and mycotoxins. This review discusses the potential effects of biocontrol fungal components on human immune responses, possibly associated to infectious, inflammatory diseases, and defective defenses. PMID:28217107

  13. [Rapamycin: a new immunosuppressive agent capable of inhibiting chronic rejection?].

    PubMed

    Viklický, O; Matl, I

    2001-01-19

    Chronic rejection represents the most common cause of transplanted graft loss in the long term. Rapamycin (sirolimus), and it's derivate RAD, are new and potent, immunosuppressive drugs. They inhibit cell proliferation driven by various growth factors. These drugs were successfully tested in some experimental models of the chronic rejection. Results of the first clinical trials have defined rapamycin pharmacokinetics and proved immunosuppressive efficacy. Rapamycin acts synergistically with cyclosporin A. The side effects are a dose-dependent thrombocytopenia and leukopenia but the most frequent is hyperlipidemia. The question, if rapamycin and RAD inhibit development of chronic rejection in man, will be solved by the prospective clinical trials over years.

  14. Radionuclide evaluation of renal transplants

    SciTech Connect

    Dubovsky, E.V.; Russell, C.D.

    1988-07-01

    In this review article, the following topics are treated: the radiopharmaceuticals /sup 99m/Tc-diethylenetriaminepentaacetic acid (DTPA), /sup 131/I-orthoiodohippurate (OIH), /sup 99m/Tc-mercaptoacetyltriglycine (MAG3), /sup 67/Ga-citrate, radioiodinated fibrinogen, /sup 99m/Tc-sulfur colloid, 111In-labelled white cells and platelets; gamma camera methods based on images, on first pass and on tubular transit; blood clearance methods; and the diagnosis of surgical complications, acute rejection (AR), acute tubular necrosis (ATN), chronic rejection (CR), and cyclosporine-A (CYA) toxicity. 94 references.

  15. The Postconditioning Effects Of Diazoxide in the Brain Following Hemorrhagic Shock and Cerebral Hypoperfusion

    DTIC Science & Technology

    2009-10-13

    battlefield casualty should be a 250 ml bolus of 7.5% hypertonic saline delivered by a rapid- infusion system [12]. Recently, it has been suggested...dose of 7.5 mg/kg is required to elicit hypotensive effects but infusions of 3.5 mg/min do not create hypotension [54, 55]. DZ is a mKATP and plasma...inhibited the mPTP by infusion of cyclosporin A 15 min before reperfusion. It was discovered that both measures significantly increased functional

  16. Roles of mucosal mast cells in intestinal cell-mediated immunity

    SciTech Connect

    Ferguson, A.; Cummins, A.G.; Munro, G.H.; Gibson, S.; Miller, H.R.

    1987-11-01

    Mucosal mast cells in rats with GvHR have been studied by cell counts, tissue levels of the specific protease RMCPII, and, as an index of MMC activation, serum RMCPII. In semi-allogeneic GvHR without host irradiation, GvHR produced modest increases in these three indices. In contrast, irradiation profoundly depleted MMC even though enteropathy was more severe than in non-irradiated hosts. We suggest that enteropathy is not dependent on the presence of MMC. In rats given cyclosporin A, lesions of GvHR were mild and numbers of MMC were low.

  17. Derivatives of cyclosporin compatible with antibody-based assays. I. The generation of (/sup 125/I)-labeled cyclosporin

    SciTech Connect

    Mahoney, W.C.; Orf, J.W.

    1985-03-01

    The immunosuppressive drug cyclosporin A, has been successfully iodinated to a specific activity of 300 Ci per gram. /sup 125/I-labeled cyclosporin and (/sup 3/H)cyclosporin are nearly equivalent as tracers in a radioimmunoassay in producing standard lines (suppression by unlabeled cyclosporin) and in assigning values to clinical samples. In addition, the (/sup 125/I)-labeled cyclosporin has greater than twice the sensitivity, and it is stable to long-term storage. Use of a (/sup 125/I)-labeled cyclosporin tracer is more convenient, more reproducible, more precise, and easier than the tritiated-cyclosporin alternative in radioimmunoassay of this compound.

  18. Structural changes of the heart during severe sepsis or septic shock.

    PubMed

    Smeding, Lonneke; Plötz, Frans B; Groeneveld, A B Johan; Kneyber, Martin C J

    2012-05-01

    Cardiovascular dysfunction is common in severe sepsis or septic shock. Although functional alterations are often described, the elevated serum levels of cardiac proteins and autopsy findings of myocardial immune cell infiltration, edema, and damaged mitochondria suggest that structural changes to the heart during severe sepsis and septic shock may occur and may contribute to cardiac dysfunction. We explored the available literature on structural (versus functional) cardiac alterations during experimental and human endotoxemia and/or sepsis. Limited data suggest that the structural changes could be prevented, and myocardial function improved by (pre-)treatment with platelet-activating factor, cyclosporin A, glutamine, caffeine, simvastatin, or caspase inhibitors.

  19. [Cyclosporine-induced gingival hyperplasia: report of one case].

    PubMed

    Bahamondes, Carlos; Godoy, Jorge

    2007-03-01

    Gingival enlargement can be an adverse effect of cyclosporine A and nifedipine use. It has a high relapse rate if the drugs are not discontinued. There is a genetic predisposition to the development of this condition and dental biofilm can also play a role. We report a 64 years old male who received a renal allograft and was treated with cyclosporine and nifedipine. He required six surgical interventions for generalized gingival enlargement. After the sixth relapse, the patient was subjected to a periodontal treatment to eliminate the dental biofilm, which decreased the rate of recurrence of gingival enlargement.

  20. Angiotensin II activates the calcineurin/NFAT signaling pathway and induces cyclooxygenase-2 expression in rat endometrial stromal cells.

    PubMed

    Abraham, Florencia; Sacerdoti, Flavia; De León, Romina; Gentile, Teresa; Canellada, Andrea

    2012-01-01

    Cyclooxygenase (COX)-2, the inducible isoform of cyclooxygenase, plays a role in the process of uterine decidualization and blastocyst attachment. On the other hand, overexpression of COX-2 is involved in the proliferation of the endometrial tissue during endometriosis. Deregulation of the renin-angiotensin-system plays a role in the pathophysiology of endometriosis and pre-eclampsia. Angiotensin II increases intracellular Ca(2+) concentration by targeting phospholypase C-gamma in endometrial stromal cells (ESC). A key element of the cellular response to Ca(2+) signals is the activity of the Ca(2+)- and calmodulin-dependent phosphatase calcineurin. Our first aim was to study whether angiotensin II stimulated Cox-2 gene expression in rat ESC and to analyze whether calcineurin activity was involved. In cells isolated from non-pregnant uteri, COX-2 expression--both mRNA and protein--was induced by co-stimulation with phorbol ester and calcium ionophore (PIo), as well as by angiotensin II. Pretreatment with the calcineurin inhibitor cyclosporin A inhibited this induction. We further analyzed the role of the calcineurin/NFAT signaling pathway in the induction of Cox-2 gene expression in non-pregnant rat ESC. Cyclosporin A abolished NFATc1 dephosphorylation and translocation to the nucleus. Cyclosporin A also inhibited the transcriptional activity driven by the Cox-2 promoter. Exogenous expression of the peptide VIVIT -specific inhibitor of calcineurin/NFAT binding- blocked the activation of Cox-2 promoter and the up-regulation of COX-2 protein in these cells. Finally we analyzed Cox-2 gene expression in ESC of early-pregnant rats. COX-2 expression--both mRNA and protein--was induced by stimulation with PIo as well as by angiotensin II. This induction appears to be calcineurin independent, since it was not abrogated by cyclosporin A. In conclusion, angiotensin II induced Cox-2 gene expression by activating the calcineurin/NFAT signaling pathway in endometrial stromal

  1. Clinical Lessons to Be Learned from Patients Developing Chronic Myeloid Leukemia While on Immunosuppressive Therapy after Solid Organ Transplantation: Yet Another Case after Orthotopic Heart Transplantation

    PubMed Central

    Oberender, Christian; Kleeberg, Lorenz; Nienhues, Nicola; Dörken, Bernd; Riess, Hanno

    2014-01-01

    Chronic myeloid leukemia developing after transplantation of solid organs and concomitant immunosuppression is a rare but still significant clinical phenomenon. We here describe an additional case of a 62-year-old male patient developing CML after orthotopic heart transplantation and medication with cyclosporine A, mofetil-mycophenolate, and steroids. Initial antileukemic therapy was imatinib at a standard dose and within 15 months of therapy a complete cytogenetic response was noted. In this report we discuss the clinical implications of these rare but biologically important cases. PMID:25478254

  2. Possible interaction between cyclosporine and glibenclamide in posttransplant diabetic patients.

    PubMed

    Islam, S I; Masuda, Q N; Bolaji, O O; Shaheen, F M; Sheikh, I A

    1996-10-01

    The possible occurrence of a kinetic interaction between cyclosporine A and glibenclamide was assessed by reviewing data of six posttransplant diabetic patients who received the two drugs concurrently. Coadministration of the two drugs resulted in a 57% increase in the steady-state plasma cyclosporine levels despite normal hepatic and renal functions in the patients. This elevation in cyclosporine level is possibly due to an interaction between the two drugs resulting from an inhibition of CYP3A4-mediated metabolism of cyclosporine by glibenclamide. This observation calls for a closer monitoring of cyclosporine plasma levels during concomitant administration of these two drugs in this group of patients.

  3. Successful treatment of dwarfism secondary to long-term steroid therapy in steroid-dependent nephrotic syndrome.

    PubMed

    Sun, Linlin; Chen, Dongping; Zhao, Xuezhi; Xu, Chenggang; Mei, Changlin

    2010-01-01

    Prolonged steroid therapy is generally used for steroid-dependent nephrotic syndrome in pediatric patients. However, dwarfism secondary to a long-term regimen and its successful reverse is rarely reported. The underlying mechanism of dwarfism is still poorly understood, as both long-term steroid use and nephrotic syndrome may interact or independently interfere with the process of growth. Here, we present a 17-year-old patient with dwarfism and steroid-dependent nephrotic syndrome and the successful treatment by recombinant human growth factor and cyclosporine A with withdrawal of steroid. We also briefly review the current understanding and the management of dwarfism in pediatric patients with nephrotic syndrome.

  4. Immunomodulation on the ocular surface: a review

    PubMed Central

    Szaflik, Jerzy; Szaflik, Jacek P.; Ambroziak, Maciej; Witkiewicz, Jan; Skopiński, Piotr

    2016-01-01

    The increasing understanding of immune mechanisms changed our perception of the ocular surface, which is now considered a compartment of the common mucosal immune system. It offered the possibility to alter the physiological immune response on the ocular surface and effectively combat inflammation, which impairs stability of the tear film and causes tear hyperosmolarity, causing symptoms of dry eye disease. The paper provides an overview of ocular surface anatomy and physiology, explains the underlying mechanisms of dry eye disease and discusses novel and promising treatment modalities, such as cyclosporine A, biological therapies using autologous serum and various growth factors as well as experimental treatment methods which are currently being investigated. PMID:27536206

  5. The Design and Use of Animal Models for Translational Research in Bone Tissue Engineering and Regenerative Medicine

    DTIC Science & Technology

    2010-01-07

    Stevenson, S., Davy, D.T., Field, G.A., Klein, L., Li, X.Q., Zika , J.M., and Goldberg, V.M. Cyclosporin A and tissue antigen matching in bone trans...plantation. Fibular allografts studied in the dog. Acta Or thop Scand 61, 517, 1990. 200. Bos, G.D., Goldberg, V.M., Powell, A.E., Heiple, K.G., and Zika , J.M...K.G., Zika , J.M., and Powell, A.E. Improved acceptance of frozen bone allografts in genetically mismatched dogs by immunosuppression. J Bone Joint

  6. Clinical efforts to reduce myocardial infarct size--the next step.

    PubMed

    Braunwald, Eugene

    2011-01-01

    Prompt myocardial reperfusion reduces infarct size in patients experiencing coronary occlusion. However, its clinical value is limited because reperfusion also causes ischemic myocardial reperfusion injury (IMRI). Considerable research to reduce IMRI has been conducted. Three interventions appear to be promising: 1) myocardial conditioning, which consists of repetitive occlusions of coronary or other arteries prior to or at the time of myocardial reperfusion; 2) the administration of cyclosporine A; and 3) the administration of adenosine. A plan for the testing of these interventions in patients with acute myocardial infarction is described.

  7. Modulation of Human Immune Response by Fungal Biocontrol Agents.

    PubMed

    Konstantinovas, Cibele; de Oliveira Mendes, Tiago A; Vannier-Santos, Marcos A; Lima-Santos, Jane

    2017-01-01

    Although the vast majority of biological control agents is generally regarded as safe for humans and environment, the increased exposure of agriculture workers, and consumer population to fungal substances may affect the immune system. Those compounds may be associated with both intense stimulation, resulting in IgE-mediated allergy and immune downmodulation induced by molecules such as cyclosporin A and mycotoxins. This review discusses the potential effects of biocontrol fungal components on human immune responses, possibly associated to infectious, inflammatory diseases, and defective defenses.

  8. Label-free imaging to study phenotypic behavioural traits of cells in complex co-cultures

    NASA Astrophysics Data System (ADS)

    Suman, Rakesh; Smith, Gabrielle; Hazel, Kathryn E. A.; Kasprowicz, Richard; Coles, Mark; O’Toole, Peter; Chawla, Sangeeta

    2016-02-01

    Time-lapse imaging is a fundamental tool for studying cellular behaviours, however studies of primary cells in complex co-culture environments often requires fluorescent labelling and significant light exposure that can perturb their natural function over time. Here, we describe ptychographic phase imaging that permits prolonged label-free time-lapse imaging of microglia in the presence of neurons and astrocytes, which better resembles in vivo microenvironments. We demonstrate the use of ptychography as an assay to study the phenotypic behaviour of microglial cells in primary neuronal co-cultures through the addition of cyclosporine A, a potent immune-modulator.

  9. Treatment of Refractory Chronic Urticaria

    PubMed Central

    Mehta, Aayushi; Godse, Kiran; Patil, Sharmila; Nadkarni, Nitin; Gautam, Manjyot

    2015-01-01

    Chronic spontaneous urticaria is a distressing disease encountered frequently in clinical practice. The current mainstay of therapy is the use of second-generation, non-sedating antihistamines. However, in patients who do not respond satisfactorily to these agents, a variety of other drugs are used. This article examines the available literature for frequently used agents including systemic corticosteroids, leukotriene receptor antagonists, dapsone, sulfasalazine, hydroxychloroquine, H2 antagonists, methotrexate, cyclosporine A, omalizumab, autologous serum therapy, and mycophenolate mofetil, with an additional focus on publications in Indian literature. PMID:26120147

  10. Treatment-refractory multi-lineage autoimmune cytopenia after unrelated cord blood transplantation: remission after combined bortezomib and vincristine treatment.

    PubMed

    Waespe, Nicolas; Zeilhofer, Ulrike; Güngör, Tayfun

    2014-11-01

    Autoimmune cytopenias (AC) after allogeneic hematopoietic stem cell transplantation (HSCT) are associated with a dismal prognosis. We describe a 1-year-old female with multi-lineage AC occurring on day +43 after HSCT. Multi-agent treatment with high-dose prednisolone, intravenous immunoglobulins, cyclosporine A, mycophenolate mofetil, sirolimus, and rituximab was unsuccessful. Combined treatment with bortezomib and vincristine in addition to ongoing immunosuppressive therapy was started on day +414 with transfusion-independence after day +444. Immunosuppressants were tapered until day +638. On day +1,121 the patient remained in remission. Bortezomib with vincristine may be a promising treatment modality for refractory AC after HSCT that requires further study.

  11. Chromoblastomycosis Caused by Cladophialophora bantiana in a Renal Transplant Recipient From Delhi, India.

    PubMed

    Verma, Prashant; Karmakar, Somenath; Pandhi, Deepika; Singal, Archana; Yadav, Pravesh; Khare, Shashi

    2015-01-01

    A 45-year-old accountant residing in Delhi, India, presented to our dermatology clinic with a small asymptomatic plaque on the little finger of his left hand of 3 months' duration. The onset of the lesion was insidious and gradually progressed to 4 cm across at the time of his first visit. The patient had undergone renal transplantation twice (the first procedure 3 months prior and the second 18 months prior). Since then, he had been receiving cyclosporine A (400 mg daily) and prednisolone (40 mg) daily in immunosuppessive doses. The patient denied any kind of cutaneous injury prior to the onset of the lesion and any similar lesions in the past.

  12. Chlamydia pneumoniae infection-related hemophagocytic lymphohistiocytosis and acute encephalitis and poliomyelitis-like flaccid paralysis.

    PubMed

    Yagi, Kanae; Kano, Gen; Shibata, Mayumi; Sakamoto, Izumi; Matsui, Hirofumi; Imashuku, Shinsaku

    2011-05-01

    A 3-year-old male presented with Chlamydia pneumoniae infection-related hemophagocytic lymphohistiocytosis (HLH). The patient developed an episode of HLH with severe skin eruption following C. pneumoniae pneumonia. Symptoms responded to steroid/cyclosporine A therapy, but the patient slowly lost consciousness and developed systemic flaccid paralysis. He was diagnosed with encephalitis/myelitis by brain and spinal MRI. Neurological symptoms and signs gradually resolved. We thought that the immune response to C. pneumoniae infection triggered the development of HLH, associated with unusual neurological complications. This report describes a novel case of C. pneumoniae-associated HLH and with poliomyelitis like flaccid paralysis.

  13. Fluorescence characterization of the hydrophobic pocket of cyclophilin B.

    PubMed

    Albani, J R; Carpentier, M; Lansiaux, C

    2008-01-01

    Human cyclophilin B is a monomeric protein that contains two tryptophan residues, Trp104 and 128. Trp128-residue belongs to the binding site of cyclosporin A and is the homologous of Trp 121 in CyPA, while Trp104 residue belongs to the hydrophobic pocket. In the present work, we studied the dynamics of Trp residue(s) of cyclophilin B and of the CyPB(w128A) mutant and of TNS-mutant complex. Our results showed that Trp-104 and TNS show restricted motions within their environments and that energy transfer between the two fluorophores is occurring.

  14. Rituximab-based immunosuppression for autoimmune haemolytic anaemia in infants.

    PubMed

    Svahn, Johanna; Fioredda, Francesca; Calvillo, Michaela; Molinari, Angelo C; Micalizzi, Concetta; Banov, Laura; Schmidt, Madalina; Caprino, Daniela; Marinelli, Doretta; Gallisai, Domenico; Dufour, Carlo

    2009-04-01

    We report a case series of four infants with severe autoimmune haemolytic anaemia (AIHA) who responded to treatment with rituximab and cyclosporine after having failed first line therapy with high-dose steroid (prednisolone 4-8 mg/kg/d). Rituximab was started at 11-90 d from onset due to continued haemolysis; three infants also received cyclosporine A. Three of four infants reached complete response, defined as normal haemoglobin, reticulocytes and negative indices of haemolysis, at 7-21 months from diagnosis. In long-term follow-up two infants remained disease-free with normal immunology, one had undefined immunodeficiency and one had autoimmune lymphoproliferative syndrome.

  15. In vitro endothelial cell susceptibility to xenobiotics: comparison of three cell types.

    PubMed

    L'Azou, B; Fernandez, P; Bareille, R; Beneteau, M; Bourget, C; Cambar, J; Bordenave, L

    2005-03-01

    In three different endothelial cell (EC) cultures (primary human umbilical cord vein, so-called HUVEC; and immortalized cell lines HBMEC and EA-hy-926), the effects of different xenobiotics were studied in order to standardize vascular EC models for in vitro pharmacotoxicological studies. Cell characteristics were first investigated by the production and the mRNA levels of known endothelial markers in the three EC culture models. EC secretory products, tissue plasminogen activator (tPA) and von Willebrand factor (vWF), were present in the supernatant of the immortalized cell lines. The mRNA levels of vWF, tPA, platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31), and beta -integrin subunit, which are involved in the control of platelet function, coagulation, and fibrinolysis as well as in cell-matrix interactions, were investigated in all EC types. For at least three parameters, cultured cells provided marked characteristics of EC phenotype, in HUVEC and in immortalized cell lines, regardless of their origin from the macro- or microcirculation. Toxicity experiments were assessed after 24 h exposure to cadmium, cyclosporin A and cisplatin by MTT assay. These experiments show nonsignificant difference in susceptibility to cyclosporin A and cadmium on HUVEC, HBMEC, and EA-hy-926. However, HBMEC, seems to be highly susceptible to cisplatin compared to HUVEC, the latter being more sensitive than EA-hy-926. For experiments conducted with cyclosporin and cadmium, cell lines could constitute an alternative material for routine cytotoxicity studies.

  16. Phentolamine inhibits exocytosis of glucagon by Gi2 protein-dependent activation of calcineurin in rat pancreatic alpha -cells.

    PubMed

    Høy, M; Bokvist, K; Xiao-Gang, W; Hansen, J; Juhl, K; Berggren, P O; Buschard, K; Gromada, J

    2001-01-12

    Capacitance measurements were used to investigate the molecular mechanisms by which imidazoline compounds inhibit glucagon release in rat pancreatic alpha-cells. The imidazoline compound phentolamine reversibly decreased depolarization-evoked exocytosis >80% without affecting the whole-cell Ca(2+) current. During intracellular application through the recording pipette, phentolamine produced a concentration-dependent decrease in the rate of exocytosis (IC(50) = 9.7 microm). Another imidazoline compound, RX871024, exhibited similar effects on exocytosis (IC(50) = 13 microm). These actions were dependent on activation of pertussis toxin-sensitive G(i2) proteins but were not associated with stimulation of ATP-sensitive K(+) channels or adenylate cyclase activity. The inhibitory effect of phentolamine on exocytosis resulted from activation of the protein phosphatase calcineurin and was abolished by cyclosporin A and deltamethrin. Exocytosis was not affected by intracellular application of specific alpha(2), I(1), and I(2) ligands. Phentolamine reduced glucagon release (IC(50) = 1.2 microm) from intact islets by 40%, an effect abolished by pertussis toxin, cyclosporin A, and deltamethrin. These data suggest that imidazoline compounds inhibit glucagon secretion via G(i2)-dependent activation of calcineurin in the pancreatic alpha-cell. The imidazoline binding site is likely to be localized intracellularly and probably closely associated with the secretory granules.

  17. Antitubercular Nanocarrier Combination Therapy: Formulation Strategies and in Vitro Efficacy for Rifampicin and SQ641.

    PubMed

    D'Addio, Suzanne M; Reddy, Venkata M; Liu, Ying; Sinko, Patrick J; Einck, Leo; Prud'homme, Robert K

    2015-05-04

    Tuberculosis (TB) remains a major global health concern, and new therapies are needed to overcome the problems associated with dosing frequency, patient compliance, and drug resistance. To reduce side effects associated with systemic drug distribution and improve drug concentration at the target site, stable therapeutic nanocarriers (NCs) were prepared and evaluated for efficacy in vitro in Mycobacterium tuberculosis-infected macrophages. Rifampicin (RIF), a current, broad-spectrum antibiotic used in TB therapy, was conjugated by degradable ester bonds to form hydrophobic prodrugs. NCs encapsulating various ratios of nonconjugated RIF and the prodrugs showed the potential ability to rapidly deliver and knockdown intracellular M. tuberculosis by nonconjugated RIF and to obtain sustained release of RIF by hydrolysis of the RIF prodrug. NCs of the novel antibiotic SQ641 and a combination NC with cyclosporine A were formed by flash nanoprecipitation. Delivery of SQ641 in NC form resulted in significantly improved activity compared to that of the free drug against intracellular M. tuberculosis. A NC formulation with a three-compound combination of SQ641, cyclosporine A, and vitamin E inhibited intracellular replication of M. tuberculosis significantly better than SQ641 alone or isoniazid, a current first-line anti-TB drug.

  18. Antitubercular Nanocarrier Combination Therapy: Formulation Strategies and in Vitro Efficacy for Rifampicin and SQ641

    PubMed Central

    2015-01-01

    Tuberculosis (TB) remains a major global health concern, and new therapies are needed to overcome the problems associated with dosing frequency, patient compliance, and drug resistance. To reduce side effects associated with systemic drug distribution and improve drug concentration at the target site, stable therapeutic nanocarriers (NCs) were prepared and evaluated for efficacy in vitro in Mycobacterium tuberculosis-infected macrophages. Rifampicin (RIF), a current, broad-spectrum antibiotic used in TB therapy, was conjugated by degradable ester bonds to form hydrophobic prodrugs. NCs encapsulating various ratios of nonconjugated RIF and the prodrugs showed the potential ability to rapidly deliver and knockdown intracellular M. tuberculosis by nonconjugated RIF and to obtain sustained release of RIF by hydrolysis of the RIF prodrug. NCs of the novel antibiotic SQ641 and a combination NC with cyclosporine A were formed by flash nanoprecipitation. Delivery of SQ641 in NC form resulted in significantly improved activity compared to that of the free drug against intracellular M. tuberculosis. A NC formulation with a three-compound combination of SQ641, cyclosporine A, and vitamin E inhibited intracellular replication of M. tuberculosis significantly better than SQ641 alone or isoniazid, a current first-line anti-TB drug. PMID:25811733

  19. Metabolic bioactivation and toxicity of ethyl 4-hydroxybenzoate in human SK-MEL-28 melanoma cells.

    PubMed

    Vad, Nikhil M; Shaik, Imam H; Mehvar, Reza; Moridani, Majid Y

    2008-05-01

    The metabolism and toxicity of ethyl 4-hydroxybenzoate (4-HEB) were investigated in vitro using tyrosinase enzyme, a melanoma molecular target, and CYP2E1 induced rat liver microsomes, and in human SK-MEL-28 melanoma cells. The results were compared to 4-hydroxyanisole (4-HA). At 90 min, 4-HEB was metabolized 48% by tyrosinase and 26% by liver microsomes while the extent of 4-HA metabolism was 196% and 88%, respectively. The IC50 (day 2) of 4-HEB and 4-HA towards SK-MEL-28 cells were 75 and 50 microM, respectively. Dicoumarol, a diaphorase inhibitor, and 1-bromoheptane, a GSH depleting agent, increased 4-HEB toxicity towards SK-MEL-28 cells indicating o-quinone formation played an important role in 4-HEB induced cell toxicity. Addition of ascorbic acid and GSH to the media was effective in preventing 4-HEB cell toxicity. Cyclosporin A and trifluoperazine, inhibitors of permeability transition pore in mitochondria, were significantly potent in inhibiting 4-HEB cell toxicity. 4-HEB caused time-dependent decline in intracellular GSH concentration which preceded cell death. 4-HEB also led to reactive oxygen species (ROS) formation in melanoma cells which exacerbated by dicoumarol and 1-bromoheptane whereas cyclosporin A and trifluoperazine prevented it. Our findings suggest that the mechanisms of 4-HEB toxicity in SK-MEL-28 were o-quinone formation, intracellular GSH depletion, ROS formation and mitochondrial toxicity.

  20. Design, synthesis, and biological evaluation of chalcone oxime derivatives as potential immunosuppressive agents.

    PubMed

    Luo, Yin; Song, Ran; Li, Yao; Zhang, Shuai; Liu, Zhi-Jun; Fu, Jie; Zhu, Hai-Liang

    2012-05-01

    A series of deoxybenzoin oximes were recently reported as potent immunosuppressive agents by our group. In order to continue the original research for potential immunosuppressive agents with high efficacy and low toxicity, we synthesized a series of new chalcone oximes and evaluated them for their cytotoxicities and immunosuppressive activities. Among the synthesized compounds, chalcone oximes 25 and 27 exhibited lower cytotoxicities and higher inhibitory activities on anti-CD3/anti-CD28 co-stimulated lymph node cells than other compounds. Specially, compound 27 displayed 200-fold lower cytotoxicity (CC(50)=2174.39 μM) than cyclosporin A (CC(50)=10.10 μM) and showed SI value (SI=176.69) close to cyclosporin A (SI=154.13). Besides, the preliminary mechanism of inhibition effect of compounds 25 and 27 was also detected by flow cytometry, and the compounds exerted immunosuppressive activities via inducing the apoptosis of activated lymph node cells in a dose dependent manner. Also, the deep mechanism of apoptosis was detected by Western blot analysis.

  1. Intravenous immunoglobulin therapy for refractory interstitial lung disease associated with polymyositis/dermatomyositis.

    PubMed

    Suzuki, Yuzo; Hayakawa, Hiroshi; Miwa, Seiichi; Shirai, Masahiro; Fujii, Masato; Gemma, Hitoshi; Suda, Takafumi; Chida, Kingo

    2009-01-01

    Interstitial lung disease (ILD) associated with polymyositis/dermatomyositis (ILD-PM/DM), including amyopathic dermatomyositis (ADM), is recognized as an important condition because it frequently causes death, despite intensive therapy with high-dose corticosteroid and immunosuppressive agents, such as cyclosporine A and cyclophosphamide. Intravenous immunoglobulin therapy (IVIG) has shown efficacy for myopathy associated with PM/DM, but its usefulness for ILD-PM/DM is unclear. This study was designed to investigate the efficacy of IVIG for refractory ILD-PM/DM. A review was made of medical charts of five patients (2 men and 3 women) who were treated with IVIG for refractory ILD-PM/DM resistant to high-dose corticosteroid and cyclosporine A and/or cyclophosphamide. One patient had acute ILD-PM and four patients had acute ILD-ADM. Of the five patients, one patient with ILD-PM and one patient with ILD-ADM survived. No adverse reactions were seen due to IVIG treatment. There were no critical differences in the clinical parameters and clinical courses between survivors and nonsurvivors. IVIG treatment is safe and could be an effective salvage therapy for refractory ILD-PM/DM in certain cases, suggesting that further controlled trials are worthwhile.

  2. Effect of Immunosuppressive Therapy on Proteinogram in Rats

    PubMed Central

    Kędzierska, Karolina; Sindrewicz, Krzysztof; Sporniak-Tutak, Katarzyna; Bober, Joanna; Stańczyk-Dunaj, Małgorzata; Dołęgowska, Barbara; Kaliszczak, Robert; Sieńko, Jerzy; Kabat-Koperska, Joanna; Gołembiewska, Edyta; Ciechanowski, Kazimierz

    2016-01-01

    Background It has been observed that the use of immunosuppressive drugs in patients after transplantation of vascularized organs may be associated with changes in the concentration of certain fractions of plasma proteins. The concentration of these proteins was correlated with an increased risk of occurrence of stage 3 chronic kidney disease (CKD). This article examines the effect of the most commonly used immunosuppressive drugs on the concentration of plasma proteins in Wistar rats. Material/methods The study involved 36 rats grouped according to the immunosuppressive regimen used (tacrolimus, mycophenolate mofetil, cyclosporine A, rapamycin, and prednisone). The rats in all study groups were treated with a 3-drug protocol for 6 months. The treatment dose was adjusted based on available data in the literature. No drugs were administered to the control group. The rats were sacrificed and blood samples collected to determine the concentration of plasma proteins using electrophoresis technique. Results Statistically significant differences were observed between protein concentrations within the studied groups. The differences related to the proteins with masses of 195 kDa, 170 kDa, 103 kDa, and 58 kDa. Conclusions (1) Immunosuppressive drugs caused changes in the proteinogram of plasma proteins. (2) The strongest effect on rat plasma proteins was exerted by a regimen based on rapamycin. Intermediate, weak, and weakest effects were observed in regimens based on cyclosporine A, tacrolimus, and mycophenolate mofetil, respectively. PMID:27288069

  3. 1,25(OH)2D3 Promotes the Efficacy of CD28 Costimulation Blockade by Abatacept

    PubMed Central

    Gardner, David H.; Jeffery, Louisa E.; Soskic, Blagoje; Briggs, Zoe; Hou, Tie Zheng; Raza, Karim

    2015-01-01

    Inhibition of the CD28:CD80/CD86 T cell costimulatory pathway has emerged as an effective strategy for the treatment of T cell–mediated inflammatory diseases. However, patient responses to CD28-ligand blockade by abatacept (CTLA-4-Ig) in conditions such as rheumatoid arthritis are variable and often suboptimal. In this study, we show that the extent to which abatacept suppresses T cell activation is influenced by the strength of TCR stimulation, with high-strength TCR stimulation being associated with relative abatacept insensitivity. Accordingly, cyclosporin A, an inhibitor of T cell stimulation via the TCR, synergized with abatacept to inhibit T cell activation. We also observed that 1,25-dihydroxyvitamin D3 enhanced the inhibition of T cell activation by abatacept, strongly inhibiting T cell activation driven by cross-linked anti-CD3, but with no effect upon anti-CD28 driven stimulation. Thus, like cyclosporin A, 1,25-dihydroxyvitamin D3 inhibits TCR-driven activation, thereby promoting abatacept sensitivity. Vitamin D3 supplementation may therefore be a useful adjunct for the treatment of conditions such as rheumatoid arthritis in combination with abatacept to promote the efficacy of treatment. PMID:26276872

  4. TRESK channel as a potential target to treat T-cell mediated immune dysfunction

    SciTech Connect

    Han, Jaehee; Kang, Dawon

    2009-12-25

    In this review, we propose that TRESK background K{sup +} channel could serve as a potential therapeutic target for T-cell mediated immune dysfunction. TRESK has many immune function-related properties. TRESK is abundantly expressed in the thymus, the spleen, and human leukemic T-lymphocytes. TRESK is highly activated by Ca{sup 2+}, calcineurin, acetylcholine, and histamine which induce hypertrophy, whereas TRESK is inhibited by immunosuppressants, such as cyclosporin A and FK506. Cyclosporine A and FK506 target the binding site of nuclear factor of activated T-cells (NFAT) to inhibit calcineurin. Interestingly, TRESK possesses an NFAT-like docking site that is present at its intracellular loop. Calcineurin has been found to interact with TRESK via specific NFAT-like docking site. When the T-cell is activated, calcineurin can bind to the NFAT-docking site of TRESK. The activation of both TRESK and NFAT via Ca{sup 2+}-calcineurin-NFAT/TRESK pathway could modulate the transcription of new genes in addition to regulating several aspects of T-cell function.

  5. Phenylephrine promotes cardiac fibroblast proliferation through calcineurin-NFAT pathway.

    PubMed

    Wang, Jing; Wang, Yibing; Zhang, Wei; Zhao, Xi; Chen, Xiangfan; Xiao, Wenyan; Zhang, Lingling; Chen, Yunxuan; Zhu, Weizhong

    2016-01-01

    Ca(2+)/calmodulin-dependent calcineurin (CaN) plays an important role in various Ca(+2) signaling pathways, among which are those involved in cardiac diseases. It has also been shown that a heightened sympathetic tone accelerates the development of heart failure. The present study investigates whether the CaN-mediated nuclear factor of activated T-cells (NFAT) pathway is involved in cultured neonatal rat cardiac fibroblast proliferation induced by phenylephrine. CF proliferation was assessed by a cell survival assay and cell counts. Green fluorescent protein-tagged NFAT3 was used to determine the cellular location of NFAT3. CaN activity and protein levels were also determined by an activity assay kit and Western blotting, respectively. Results showed that phenylephrine promoted CF proliferation, which was abolished by α1-adrenergic receptor antagonist (prazosin), a blocker of Ca(+2) influx (nifedipine), an intracellular Ca(2+) buffer (BAPTA-AM), CaN inhibitors (cyclosporin A and FK506), and over-expression of dominant negative CaN. Phenylephrine activated CaN and evoked NFAT3 nuclear translocation, both of which were blocked by cyclosporine A (CsA) or over-expression of dominant negative CaN. These results suggest that the Ca(2+)/CaN/NFAT pathway mediates PE-induced CF proliferation, and this pathway might be a possible therapeutic target in cardiac fibrosis.

  6. Battery of behavioral tests in mice to study postoperative delirium

    PubMed Central

    Peng, Mian; Zhang, Ce; Dong, Yuanlin; Zhang, Yiying; Nakazawa, Harumasa; Kaneki, Masao; Zheng, Hui; Shen, Yuan; Marcantonio, Edward R.; Xie, Zhongcong

    2016-01-01

    Postoperative delirium is associated with increased morbidity, mortality and cost. However, its neuropathogenesis remains largely unknown, partially owing to lack of animal model(s). We therefore set out to employ a battery of behavior tests, including natural and learned behavior, in mice to determine the effects of laparotomy under isoflurane anesthesia (Anesthesia/Surgery) on these behaviors. The mice were tested at 24 hours before and at 6, 9 and 24 hours after the Anesthesia/Surgery. Composite Z scores were calculated. Cyclosporine A, an inhibitor of mitochondria permeability transient pore, was used to determine potential mitochondria-associated mechanisms of these behavioral changes. Anesthesia/Surgery selectively impaired behaviors, including latency to eat food in buried food test, freezing time and time spent in the center in open field test, and entries and duration in the novel arm of Y maze test, with acute onset and various timecourse. The composite Z scores quantitatively demonstrated the Anesthesia/Surgery-induced behavior impairment in mice. Cyclosporine A selectively ameliorated the Anesthesia/Surgery-induced reduction in ATP levels, the increases in latency to eat food, and the decreases in entries in the novel arm. These findings suggest that we could use a battery of behavior tests to establish a mouse model to study postoperative delirium. PMID:27435513

  7. The influence of intrauterine exposure to immunosuppressive treatment on changes in the immune system in juvenile Wistar rats

    PubMed Central

    Kabat-Koperska, Joanna; Kolasa-Wołosiuk, Agnieszka; Wojciuk, Bartosz; Wojciechowska-Koszko, Iwona; Roszkowska, Paulina; Krasnodębska-Szponder, Barbara; Paczkowska, Edyta; Safranow, Krzysztof; Gołembiewska, Edyta; Machaliński, Bogusław; Ciechanowski, Kazimierz

    2016-01-01

    Background In our study, we assessed the impact of immunosuppressive drug combinations on changes in the immune system of juvenile Wistar rats exposed to these drugs during pregnancy. We primarily concentrated on changes in two organs of the immune system – the thymus and the spleen. Methods The study was conducted on 40 (32+8) female Wistar rats administered full and half dose of drugs, respectively, subjected to regimens commonly used in therapy of human kidney transplant recipients ([1] cyclosporine A, mycophenolate mofetil, and prednisone; [2] tacrolimus, mycophenolate mofetil, and prednisone; [3] cyclosporine A, everolimus, and prednisone). The animals received drugs by oral gavage 2 weeks before pregnancy and during 3 weeks of pregnancy. Results There were no statistically significant differences in the weight of the thymus and spleen, but changes were found in the results of blood hematology, cytometry from the spleen, and a histologic examination of the examined immune organs of juvenile Wistar rats. In the cytokine assay, changes in the level of interleukine 17 (IL-17) after increasing amounts of concanavaline A were dose-dependent; the increase of IL-17 was blocked after administration of higher doses of immunosuppressive drugs. However, after a reduction of doses, its increase resumed. Conclusion Qualitative, quantitative, and morphological changes in the immune system of infant rats born to pharmacologically immunosuppressed females were observed. Thymus structure, spleen composition, and splenocyte IL-17 production were mostly affected in a drug regimen–dependent manner. PMID:27471376

  8. Extracellular Cyclophilins Contribute to the Regulation of Inflammatory Responses1

    PubMed Central

    Arora, Kamalpreet; Gwinn, William M.; Bower, Molly A.; Watson, Alan; Okwumabua, Ifeanyi; MacDonald, H. Robson; Bukrinsky, Michael I.; Constant, Stephanie L.

    2010-01-01

    The main regulators of leukocyte trafficking during inflammatory responses are chemokines. However, another class of recently identified chemotactic agents is extracellular cyclophilins, the proteins mostly known as receptors for the immunosuppressive drug, cyclosporine A. Cyclophilins can induce leukocyte chemotaxis in vitro and have been detected at elevated levels in inflamed tissues, suggesting that they might contribute to inflammatory responses. We recently identified CD147 as the main signaling receptor for cyclophilin A. In the current study we examined the contribution of cyclophilin-CD147 interactions to inflammatory responses in vivo using a mouse model of acute lung injury. Blocking cyclophilin-CD147 interactions by targeting CD147 (using anti-CD147 Ab) or cyclophilin (using nonimmunosuppressive cyclosporine A analog) reduced tissue neutrophilia by up to 50%, with a concurrent decrease in tissue pathology. These findings are the first to demonstrate the significant contribution of cyclophilins to inflammatory responses and provide a potentially novel approach for reducing inflammation-mediated diseases. PMID:15972687

  9. The pro-oxidative drug WF-10 inhibits serial killing by primary human cytotoxic T-cells.

    PubMed

    Wabnitz, G H; Balta, E; Schindler, S; Kirchgessner, H; Jahraus, B; Meuer, S; Samstag, Y

    2016-01-01

    Cytotoxic T-cells (CTLs) play an important role in many immune-mediated inflammatory diseases. Targeting cytotoxicity of CTLs would allow to interfere with immune-mediated tissue destruction. Here we demonstrate that WF-10, a pro-oxidative compound, inhibits CTL-mediated cytotoxicity. WF-10 did not influence early steps of target-cell killing, but impaired the ability of CTLs to detach from the initial target cell and to move to a second target cell. This reduced serial killing was accompanied by stronger enrichment of the adhesion molecule LFA-1 in the cytolytic immune synapse. LFA-1 clustering requires activation of the actin-bundling protein L-plastin and was accordingly diminished in L-plastin knockdown cells. Interestingly, WF-10 likely acts through regulating L-plastin: (I) It induced L-plastin activation through phosphorylation leading to enhanced LFA-1-mediated cell adhesion, and, importantly, (II) WF-10 lost its influence on target-cell killing in L-plastin knockdown cells. Finally, we demonstrate that WF-10 can improve immunosuppression by conventional drugs. Thus, while cyclosporine A alone had no significant effect on cytotoxicity of CTLs, a combination of cyclosporine A and WF-10 blocked target-cell killing synergistically. Together, our findings suggest that WF-10 - either alone or in combination with conventional immunosuppressive drugs - may be efficient to control progression of diseases, in which CTLs are crucially involved.

  10. Ascorbate and low concentrations of FeSO4 induce Ca2+-dependent pore in rat liver mitochondria.

    PubMed

    Brailovskaya, I V; Starkov, A A; Mokhova, E N

    2001-08-01

    Oxidative stress is one of the most frequent causes of tissue and cell injury in various pathologies. The molecular mechanism of mitochondrial damage under conditions of oxidative stress induced in vitro with low concentrations of FeSO4 and ascorbate (vitamin C) was studied. FeSO4 (1-4 microM) added to rat liver mitochondria that were incubated in the presence of 2.3 mM ascorbate induced (with a certain delay) a decrease in membrane potential and high-amplitude swelling. It also significantly decreased the ability of mitochondria to accumulate exogenous Ca2+. All the effects of FeSO4 + ascorbate were essentially prevented by cyclosporin A, a specific inhibitor of the mitochondrial Ca2+-dependent pore (also known as the mitochondrial permeability transition). EGTA restored the membrane potential of mitochondria de-energized with FeSO4 + ascorbate. We hypothesize that oxidative stress induced in vitro with FeSO4 and millimolar concentrations of ascorbate damages mitochondria by inducing the cyclosporin A-sensitive Ca2+-dependent pore in the inner mitochondrial membrane.

  11. The influence of immunosuppressive drugs on neural stem/progenitor cell fate in vitro

    SciTech Connect

    Skardelly, Marco; Glien, Anja; Groba, Claudia; Schlichting, Nadine; Kamprad, Manja; Meixensberger, Juergen; Milosevic, Javorina

    2013-12-10

    In allogenic and xenogenic transplantation, adequate immunosuppression plays a major role in graft survival, especially over the long term. The effect of immunosuppressive drugs on neural stem/progenitor cell fate has not been sufficiently explored. The focus of this study is to systematically investigate the effects of the following four different immunotherapeutic strategies on human neural progenitor cell survival/death, proliferation, metabolic activity, differentiation and migration in vitro: (1) cyclosporine A (CsA), a calcineurin inhibitor; (2) everolimus (RAD001), an mTOR-inhibitor; (3) mycophenolic acid (MPA, mycophenolate), an inhibitor of inosine monophosphate dehydrogenase and (4) prednisolone, a steroid. At the minimum effective concentration (MEC), we found a prominent decrease in hNPCs' proliferative capacity (BrdU incorporation), especially for CsA and MPA, and an alteration of the NAD(P)H-dependent metabolic activity. Cell death rate, neurogenesis, gliogenesis and cell migration remained mostly unaffected under these conditions for all four immunosuppressants, except for apoptotic cell death, which was significantly increased by MPA treatment. - Highlights: • Four immunosuppresants (ISs) were tested in human neural progenitor cells in vitro. • Cyclosporine A and mycophenolic acid showed a prominent anti-proliferative activity • Mycophenolic acid exhibited a significant pro-apoptotic effect. • NAD(P)H-dependent metabolic activity was occasionally induced by ISs. • Neuronal differentiation and migration potential remained unaffected by ISs treatment.

  12. Subcutaneous Panniculitis-Like T Cell Lymphoma Mimicking Early-Onset Nodular Panniculitis

    PubMed Central

    Shen, Guifen; Dong, Lingli; Zhang, Shengtao

    2016-01-01

    Patient: Male, 24 Final Diagnosis: Subcutaneous panniculitis-like T-cell lymphoma Symptoms: Fever • skin nodules Medication: — Clinical Procedure: Skin biopsy • PET-CT Specialty: Hematology Objective: Rare disease Background: Subcutaneous panniculitis-like T cell lymphoma is a very uncommon subtype of cutaneous T cell lymphoma. The manifestations of this rare disease are atypical at onset, and may mimic some rheumatic or dermatologic diseases, which causes the delay of diagnosis and treatment. Case Report: We report a 24-year-old man suffering from intermittent fever and skin nodules on the left anterior chest wall, who was initially misdiagnosed with nodular panniculitis and finally diagnosed with subcutaneous panniculitis-like T cell lymphoma through repeat examination of biopsy of the skin nodule. Positron emission tomography revealed extracutaneous adipose tissue involvement. Subsequently, hemophagocytic syndrome occurred while under a conventional dose of glucocorticoid, but remission was induced by treatment with cyclosporine A and high doses of dexamethasone. Conclusions: In order to avoid the delay diagnosis and inappropriate treatment of subcutaneous panniculitis-like T cell lymphoma, in addition to a thorough physical examination, PET-CT and disease-specific pathologic, immunophenotypic, and T cell receptor tests of the skin biopsy should be performed. Extracutaneous involvement, especially hemophagocytic syndrome, indicated worse prognosis. Even so, cyclosporine A plus high-dose corticosteroid could be an option of treatment. PMID:27342380

  13. Multilayer Spheroids To Quantify Drug Uptake and Diffusion in 3D

    PubMed Central

    2015-01-01

    There is a need for new quantitative in vitro models of drug uptake and diffusion to help assess drug toxicity/efficacy as well as new more predictive models for drug discovery. We report a three-dimensional (3D) multilayer spheroid model and a new algorithm to quantitatively study uptake and inward diffusion of fluorescent calcein via gap junction intercellular communication (GJIC). When incubated with calcein-AM, a substrate of the efflux transporter P-glycoprotein (Pgp), spheroids from a variety of cell types accumulated calcein over time. Accumulation decreased in spheroids overexpressing Pgp (HEK-MDR) and was increased in the presence of Pgp inhibitors (verapamil, loperamide, cyclosporin A). Inward diffusion of calcein was negligible in spheroids that lacked GJIC (OVCAR-3, SK-OV-3) and was reduced in the presence of an inhibitor of GJIC (carbenoxolone). In addition to inhibiting Pgp, verapamil and loperamide, but not cyclosporin A, inhibited inward diffusion of calcein, suggesting that they also inhibit GJIC. The dose response curves of verapamil’s inhibition of Pgp and GJIC were similar (IC50: 8 μM). The method is amenable to many different cell types and may serve as a quantitative 3D model that more accurately replicates in vivo barriers to drug uptake and diffusion. PMID:24641346

  14. Coexistence of translocated cytochrome c and nitrated protein in neurons of the rat cerebral cortex after oxygen and glucose deprivation.

    PubMed

    Alonso, D; Encinas, J M; Uttenthal, L O; Boscá, L; Serrano, J; Fernández, A P; Castro-Blanco, S; Santacana, M; Bentura, M L; Richart, A; Fernández-Vizarra, P; Rodrigo, J

    2002-01-01

    Changes in the distribution of immunoreactive cytochrome c and protein nitration were studied in the rat cerebral cortex after oxygen and glucose deprivation by bright field, confocal and electron microscopy. In control cerebral cortex, nitrotyrosine immunoreactivity indicating protein nitration was found mostly in the neuronal nuclear region, with only a small amount distributed in the cytosol, whereas cytochrome c immunoreactivity was found at the inner membrane and in the intermembrane space of the mitochondria. During the recovery phase after oxygen and glucose deprivation, cytochrome c immunoreactivity was released from the intermembrane space of swollen mitochondria into the surrounding cytosol. The cytosol now also displayed nitrotyrosine immunoreactivity, which had diminished in the nuclear region. Both immunoreactivities were dispersed throughout the soma and processes of the cortical neurons. These changes were largely prevented by the administration of cyclosporin A, which inhibits both the mitochondrial permeability transition and the neuronal isoform of nitric oxide synthase while blocking the induction of the inducible isoform. Ischemia/reperfusion injury increases the production of nitric oxide, reactive oxygen species and intracellular factors that damage the mitochondria and liberate apoptotic factors. We suggest that translocation of cytochrome c from the mitochondria to the cytosol, which has been shown to precede the mitochondrial permeability transition, could result from peroxynitrite-mediated nitration. This phenomenon is attenuated by cyclosporin A administration, suggesting a neuroprotective role for this agent.

  15. Xenon treatment attenuates early renal allograft injury associated with prolonged hypothermic storage in rats.

    PubMed

    Zhao, Hailin; Yoshida, Akira; Xiao, Wei; Ologunde, Rele; O'Dea, Kieran P; Takata, Masao; Tralau-Stewart, Catherine; George, Andrew J T; Ma, Daqing

    2013-10-01

    Prolonged hypothermic storage elicits severe ischemia-reperfusion injury (IRI) to renal grafts, contributing to delayed graft function (DGF) and episodes of acute immune rejection and shortened graft survival. Organoprotective strategies are therefore needed for improving long-term transplant outcome. The aim of this study is to investigate the renoprotective effect of xenon on early allograft injury associated with prolonged hypothermic storage. Xenon exposure enhanced the expression of heat-shock protein 70 (HSP-70) and heme oxygenase 1 (HO-1) and promoted cell survival after hypothermia-hypoxia insult in human proximal tubular (HK-2) cells, which was abolished by HSP-70 or HO-1 siRNA. In the brown Norway to Lewis rat renal transplantation, xenon administered to donor or recipient decreased the renal tubular cell death, inflammation, and MHC II expression, while delayed graft function (DGF) was therefore reduced. Pathological changes associated with acute rejection, including T-cell, macrophage, and fibroblast infiltration, were also decreased with xenon treatment. Donors or recipients treated with xenon in combination with cyclosporin A had prolonged renal allograft survival. Xenon protects allografts against delayed graft function, attenuates acute immune rejection, and enhances graft survival after prolonged hypothermic storage. Furthermore, xenon works additively with cyclosporin A to preserve post-transplant renal function.

  16. Battery of behavioral tests in mice to study postoperative delirium.

    PubMed

    Peng, Mian; Zhang, Ce; Dong, Yuanlin; Zhang, Yiying; Nakazawa, Harumasa; Kaneki, Masao; Zheng, Hui; Shen, Yuan; Marcantonio, Edward R; Xie, Zhongcong

    2016-07-20

    Postoperative delirium is associated with increased morbidity, mortality and cost. However, its neuropathogenesis remains largely unknown, partially owing to lack of animal model(s). We therefore set out to employ a battery of behavior tests, including natural and learned behavior, in mice to determine the effects of laparotomy under isoflurane anesthesia (Anesthesia/Surgery) on these behaviors. The mice were tested at 24 hours before and at 6, 9 and 24 hours after the Anesthesia/Surgery. Composite Z scores were calculated. Cyclosporine A, an inhibitor of mitochondria permeability transient pore, was used to determine potential mitochondria-associated mechanisms of these behavioral changes. Anesthesia/Surgery selectively impaired behaviors, including latency to eat food in buried food test, freezing time and time spent in the center in open field test, and entries and duration in the novel arm of Y maze test, with acute onset and various timecourse. The composite Z scores quantitatively demonstrated the Anesthesia/Surgery-induced behavior impairment in mice. Cyclosporine A selectively ameliorated the Anesthesia/Surgery-induced reduction in ATP levels, the increases in latency to eat food, and the decreases in entries in the novel arm. These findings suggest that we could use a battery of behavior tests to establish a mouse model to study postoperative delirium.

  17. Expression and functional activity of P-glycoprotein in cultured cerebral capillary endothelial cells.

    PubMed

    Hegmann, E J; Bauer, H C; Kerbel, R S

    1992-12-15

    Analysis of a panel of endothelial cells passaged between 5 and 25 times and derived from various organs and species demonstrated that murine and porcine cerebral capillary endothelial cells actively excluded the fluorescent dye rhodamine 123, a substrate of P-glycoprotein. In addition, rhodamine 123 accumulation could be enhanced by the multidrug resistance chemosensitizer verapamil, known to reduce P-glycoprotein-mediated drug efflux. Cloned murine and porcine cerebral capillary endothelial cells were immunoreactive with the C219 monoclonal antibody to P-glycoprotein, and a C219 epitope-specific blocking peptide could abolish staining. The antiproliferative and cytotoxic effects of vincristine, but not cis-platinum(II) diamminedichloride, were increased by the addition of either verapamil or cyclosporin A to brain endothelial cell cultures in a 72-h assay, as determined by [3H]thymidine incorporation and total protein measurement. Cyclosporin A was a more effective reversal agent than verapamil. Thus, a P-glycoprotein isoform may be constitutively expressed in brain endothelial cells in vitro and supports the available data on in situ immunohistochemical staining of P-glycoprotein at the blood-brain barrier. In addition, these findings may indicate that one function of P-glycoprotein in vivo at the blood-brain barrier is the exclusion of xenobiotics from central nervous system tissues.

  18. Anabolic steroids activate calcineurin-NFAT signaling and thereby increase myotube size and reduce denervation atrophy.

    PubMed

    Qin, Weiping; Pan, Jiangping; Wu, Yong; Bauman, William A; Cardozo, Christopher

    2015-01-05

    Anabolic androgens have been shown to reduce muscle loss due to immobilization, paralysis and many other medical conditions, but the molecular basis for these actions is poorly understood. We have recently demonstrated that nandrolone, a synthetic androgen, slows muscle atrophy after nerve transection associated with down-regulation of regulator of calcineurin 2 (RCAN2), a calcineurin inhibitor, suggesting a possible role of calcineurin-NFAT signaling. To test this possibility, rat gastrocnemius muscle was analyzed at 56 days after denervation. In denervated muscle, calcineurin activity declined and NFATc4 was excluded from the nucleus and these effects were reversed by nandrolone. Similarly, nandrolone increased calcineurin activity and nuclear NFATc4 levels in cultured L6 myotubes. Nandrolone also induced cell hypertrophy that was blocked by cyclosporin A or overexpression of RCAN2. Finally protection against denervation atrophy by nandrolone in rats was blocked by cyclosporin A. These results demonstrate for the first time that nandrolone activates calcineurin-NFAT signaling, and that such signaling is important in nandrolone-induced cell hypertrophy and protection against paralysis-induced muscle atrophy.

  19. Chicken FK506-binding protein, FKBP65, a member of the FKBP family of peptidylprolyl cis-trans isomerases, is only partially inhibited by FK506.

    PubMed Central

    Zeng, B; MacDonald, J R; Bann, J G; Beck, K; Gambee, J E; Boswell, B A; Bächinger, H P

    1998-01-01

    The chicken FK506-binding protein FKBP65, a peptidylprolyl cis-trans isomerase, is a rough endoplasmic reticulum protein that contains four domains homologous to FKBP13, another rough endoplasmic reticulum PPIase. Analytical ultracentrifugation suggests that in FKBP65 these four domains are arranged in a linear extended structure with a length of about 26 nm and a diameter of about 3 nm. All four domains are therefore expected to be accessible to substrates. The specificity of FKBP65 towards a number of peptide substrates was determined. The specific activity of FKBP65 is generally lower than that of FKBP12 when expressed as a per domain activity. The substrate specificity of FKBP65 also differs from that of FKBP12. Inhibition studies show that only one of the four domains can be inhibited by FK506, a powerful inhibitor of all other known FKBPs. Furthermore, the same domain seems to be susceptible to inhibition by cyclosporin A. No other FKBPs were shown to be inhibited by cyclosporin A. It is also shown that FKBP65 can catalyse the re-folding of type III collagen in vitro with a kcat/Km = 4.3 x 10(3) M-1.s-1. PMID:9461498

  20. Immunophilins interact with calcineurin in the absence of exogenous immunosuppressive ligands.

    PubMed Central

    Cardenas, M E; Hemenway, C; Muir, R S; Ye, R; Fiorentino, D; Heitman, J

    1994-01-01

    The peptidyl-prolyl isomerases FKBP12 and cyclophilin A (immunophilins) form complexes with the immunosuppressants FK506 and cyclosporin A that inhibit the phosphatase calcineurin. With the yeast two hybrid system, we detect complexes between FKBP12 and the calcineurin A catalytic subunit in both the presence and absence of FK506. Mutations in FKBP12 surface residues or the absence of the calcineurin B regulatory subunit perturb the FK506-dependent, but not the ligand-independent, FKBP12-calcineurin complex. By affinity chromatography, both FKBP12 and cyclophilin A bind calcineurin A in the absence of ligand, and FK506 and cyclosporin A respectively potentiate these interactions. Both in vivo and in vitro, the peptidyl-prolyl isomerase active sites are dispensable for ligand-independent immunophilin-calcineurin complexes. Lastly, by genetic analyses we demonstrate that FKBP12 modulates calcineurin functions in vivo. These findings reveal that immunophilins interact with calcineurin in the absence of exogenous ligands and suggest that immunosuppressants may take advantage of the inherent ability of immunophilins to interact with calcineurin. Images PMID:7529175

  1. Novel molecular imaging approach for subclinical detection of iritis and evaluation of therapeutic success.

    PubMed

    Xie, Fang; Sun, Dawei; Schering, Alexander; Nakao, Shintaro; Zandi, Souska; Liu, Ping; Hafezi-Moghadam, Ali

    2010-07-01

    There is an urgent need for early diagnosis in medicine, whereupon effective treatments could prevent irreversible tissue damage. Acute anterior chamber inflammation is the most common form of uveitis and a major cause of vision loss. The proximity of the iris vasculature to the light-permeable cornea and its involvement in ocular inflammation make it an ideal target for noninvasive molecular imaging. To accomplish this, carboxylated fluorescent microspheres (MSs) were conjugated with recombinant P-selectin glycoprotein ligand-1 and systemically injected in endotoxin-induced uveitic animals. MS adhesion in the microcirculation of the anterior and posterior chamber was visualized by intravital microscopy and scanning laser ophthalmoscopy. In iritic animals, significantly higher numbers of recombinant P-selectin glycoprotein ligand-1-conjugated MSs adhered to the endothelium (P = 0.03) matching the increase in leukocyte adhesion. Conjugated MSs specifically interacted with firmly adhering leukocytes, allowing quantification of the endogenous immune response. Topical eye drop treatment with dexamethasone (P < 0.01) or cyclosporine A (P < 0.01) significantly lowered MS adhesion in iris vessels. Surprisingly, topical dexamethasone significantly reduced MS interaction in the fundus vessels (P < 0.01), while cyclosporine A did not. In vivo MS accumulation preceded clinical signs of anterior uveitis and leukocyte adhesion in iris vasculature. This work introduces noninvasive subclinical detection of endothelial injury in the iris vasculature, providing a unique opportunity for quantifying vascular injury and immune response in vivo.

  2. An alternative model of vascularized bone marrow transplant: partial femur transplantation.

    PubMed

    Chen, Jian-Wu; Chen, Chen; Su, Ying-Jun; Yan, Lun; Wang, Shi-Ping; Guo, Shu-Zhong

    2014-12-01

    The vascularized whole femur transplantation model is one of the commonly used vascularized bone marrow transplant models. It involves technical complexity and morbidities. To optimize this model, we took 2/3 femur as the carrier of bone marrow cells, and developed a vascularized partial femur model. Four experimental groups were carried out, namely, the syngeneic partial femur transplantation, allogeneic partial femur transplantation with or without cyclosporine A, and allogeneic whole femur transplantation with cyclosporine A. The results showed that the partial femur model was technically simpler and shortened the operative and ischemia time compared to the whole femur model. Gross and histologic appearance confirmed the viability of femur, and its bone marrow inside the bone could also maintain normal morphologically at 60-day posttransplant. Besides, donor multilineage chimerism could be continuously detected in immunosuppressed allogeneic partial femur recipients at 1-, 2-, 3-, 4-, and 8-week posttransplant, and it showed no significant differences when compared with whole femur transplantation. Meanwhile, long-term engraftment of donor-origin cells was also confirmed in recipients' bone marrow, lymph nodes, and spleen, but not in thymus. Therefore, the vascularized partial femur can serve as a continuous resource of bone morrow cells and may provide a useful tool for the study of immune tolerance in vascularized composite allotransplantation.

  3. Examination of metabolic pathways and identification of human liver cytochrome P450 isozymes responsible for the metabolism of barnidipine, a calcium channel blocker.

    PubMed

    Teramura, T; Fukunaga, Y; Van Hoogdalem, E J; Watanabe, T; Higuchi, S

    1997-09-01

    1. In a human liver microsomal system, barnidipine was converted into three primary metabolites, an N-debenzylated product (M-1), a hydrolyzed product of the benzyl-pyrrolidine ester (M-3) and an oxidized product of the dihydropyridine ring (M-8). 2. Involvement of CYP3A in the three primary metabolic pathways was revealed by the following studies: (a) inhibition of CYP3A, (b) a correlation study using 10 individual human liver microsomes and (c) cDNA-expression studies. The secondary metabolites, M-2 and M-4 (pyridine forms of M-1 and M-3), were most likely generated from M-8 but were unlikely from M-1 or M-3. Involvement of CYP3A in the secondary pathways of metabolism is also suggested. 3. The possibility of interactions between barnidipine and coadministered drugs was examined in vitro. The formation rate of the primary metabolites was little affected by warfarin, theophylline, phenytoin, diclofenac and amitriptyline at concentrations of 200 microM, but was inhibited by glibenclamide, simvastatin and cyclosporin A. IC50 for the latter drugs was estimated to be > 200, 200 and 20 microM respectively, which was roughly > 200, 6000 and 50 times higher than their respective therapeutic plasma levels, suggesting that interactions with cyclosporin A, a CYP3A inhibitor, are of possible clinical relevance.

  4. Effect of recipient's race on pediatric renal allograft survival: a single-center study.

    PubMed

    Schulman, S L; Palmer, J; Dunn, S; Kaiser, B A; Polinsky, M S; Baluarte, H J

    1992-06-01

    One hundred twenty-seven children (83 males, 44 females, 86 white, 41 nonwhite; mean age 12.1 years) who received 160 renal transplants between 1980 and 1989 were retrospectively studied. Variables such as age, sex, primary diagnosis, type, HLA-DR mismatching, and repeated transplants were compared between races and found not to be significant. However, HLA-A and -B cadaveric-graft mismatching, which was equivalent between whites and nonwhites prior to 1985 (pre-cyclosporine A era), has significantly favored whites (49% with 0 to 2 HLA-A and -B mismatch vs 16% in nonwhites) since 1985 (P less than .05), and a significantly higher proportion of nonwhite patients (59%) were receiving medical assistance (P less than .0001). Graft survival was evaluated with significantly poorer results in nonwhites as compared to whites (P less than .05). Although no difference was found between white and nonwhite cadaveric-graft survival before 1985, nonwhites had significantly worse graft survival since 1985 (72% vs 59% for 1 year and 61% vs 24% for 3 years in whites and nonwhites, respectively; P less than .05). Subpopulations such as nonwhite adolescents, nonwhite females, nonwhites with repeated transplants, and all low socioeconomic patients were identified as high-risk children with poor long-term survival. It is concluded that secondary to poorer matching since 1985 there has been decreased graft survival in nonwhites despite cyclosporine A. Attempts to improve matching and attention to high-risk groups are needed for equivalent survival.

  5. FKBP51, a novel T-cell-specific immunophilin capable of calcineurin inhibition.

    PubMed Central

    Baughman, G; Wiederrecht, G J; Campbell, N F; Martin, M M; Bourgeois, S

    1995-01-01

    The immunosuppressive drugs FK506 and cyclosporin A block T-lymphocyte proliferation by inhibiting calcineurin, a critical signaling molecule for activation. Multiple intracellular receptors (immunophilins) for these drugs that specifically bind either FK506 and rapamycin (FK506-binding proteins [FKBPs]) or cyclosporin A (cyclophilins) have been identified. We report the cloning and characterization of a new 51-kDa member of the FKBP family from murine T cells. The novel immunophilin, FKBP51, is distinct from the previously isolated and sequenced 52-kDa murine FKBP, demonstrating 53% identity overall. Importantly, Western blot (immunoblot) analysis showed that unlike all other FKBPs characterized to date, FKBP51 expression was largely restricted to T cells. Drug binding to recombinant FKBP51 was demonstrated by inhibition of peptidyl prolyl isomerase activity. As judged from peptidyl prolyl isomerase activity, FKBP51 had a slightly higher affinity for rapamycin than for FK520, an FK506 analog. FKBP51, when complexed with FK520, was capable of inhibiting calcineurin phosphatase activity in an in vitro assay system. Inhibition of calcineurin phosphatase activity has been implicated both in the mechanism of immunosuppression and in the observed toxic side effects of FK506 in nonlymphoid cells. Identification of a new FKBP that can mediate calcineurin inhibition and is restricted in its expression to T cells suggests that new immunosuppressive drugs may be identified that, by virtue of their specific interaction with FKBP51, would be targeted in their site of action. PMID:7542743

  6. Juvenile idiopathic arthritis complicated by amyloidosis with secondary nephrotic syndrome - effective treatment with tocilizumab.

    PubMed

    Kwiatkowska, Małgorzata; Jednacz, Ewa; Rutkowska-Sak, Lidia

    2015-01-01

    A case report of a boy with juvenile idiopathic arthritis since the age of 2 years, generalized onset, complicated by nephrotic syndrome due to secondary type A amyloidosis is presented. In the patient the disease had an especially severe course, complicated by frequent infections, making routine treatment difficult. Amyloidosis was diagnosed in the 5(th) year of the disease based on a rectal biopsy. Since the disease onset the boy has been taking prednisolone and sequentially cyclosporine A, methotrexate, chlorambucil, etanercept, and cyclophosphamide. Clinical and laboratory remission was observed after treatment with tocilizumab. After 42 months of treatment with tocilizumab the boy's condition is good. There is no pain or joint edema, and no signs of nephrotic syndrome.

  7. [Protective effect of verapamil and dopamine against cyclosporine-induced vasoconstriction in isolated glomeruli in rats].

    PubMed

    L'Azou, B; Lagroye, I; Plande, J; Lakhdar, B; Cambar, J

    1992-12-02

    Cyclosporin A (CsA)-induced nephrotoxicity is characterized by dramatic changes in glomerular filtration rate and renal plasma flow, largely limiting the clinical use of this drug. The vasoconstrictive response of CsA could explain, in part, these hemodynamic alterations. The present study compares the area changes in rat-isolated glomeruli incubated with CsA alone or after pre-treatment with verapamil and dopamine. In verapamil-pretreated CsA-intoxicated glomeruli, size decrease was reduced (-1.5 percent at T10, -3.1 percent at T20 and -4.8 percent at T30), when compared with CsA alone (-4.7 percent at T10, -10.1 percent at T20 and -12 percent at T30). The results obtained with dopamine were similar. In conclusion, verapamil and dopamine can be regarded as fair protective agents against CsA-induced vasoconstriction in rat-isolated glomeruli.

  8. Alisporivir inhibits MERS- and SARS-coronavirus replication in cell culture, but not SARS-coronavirus infection in a mouse model.

    PubMed

    de Wilde, Adriaan H; Falzarano, Darryl; Zevenhoven-Dobbe, Jessika C; Beugeling, Corrine; Fett, Craig; Martellaro, Cynthia; Posthuma, Clara C; Feldmann, Heinz; Perlman, Stanley; Snijder, Eric J

    2017-01-15

    Currently, there is no registered treatment for infections with emerging zoonotic coronaviruses like SARS- and MERS-coronavirus. We here report that in cultured cells low-micromolar concentrations of alisporivir, a non-immunosuppressive cyclosporin A-analog, inhibit the replication of four different coronaviruses, including MERS- and SARS-coronavirus. Ribavirin was found to further potentiate the antiviral effect of alisporivir in these cell culture-based infection models, but this combination treatment was unable to improve the outcome of SARS-CoV infection in a mouse model. Nevertheless, our data provide a basis to further explore the potential of Cyp inhibitors as host-directed, broad-spectrum inhibitors of coronavirus replication.

  9. Jatrophane diterpenes as modulators of multidrug resistance. Advances of structure-activity relationships and discovery of the potent lead pepluanin A.

    PubMed

    Corea, Gabriella; Fattorusso, Ernesto; Lanzotti, Virginia; Motti, Riccardo; Simon, Pierre-Noël; Dumontet, Charles; Di Pietro, Attilio

    2004-02-12

    From the whole plant of Euphorbia peplus L., five new diterpenes based on a jatrophane skeleton (pepluanins A-E, 1-5) were isolated, together with two known analogues (6 and 7), which served to divulge in detail the structure-activity relationships within this class of P-glycoprotein inhibitors. The results revealed the importance of substitutions on the medium-sized ring (carbons 8, 9, 14, and 15). In particular, the activity is collapsed by the presence of a free hydroxyl at C-8, while it increases with a carbonyl at C-14, an acetoxyl at C-9, and a free hydroxyl at C-15. The most potent compound of the series, pepluanin A, showed a very high activity for a jatrophane diterpene, outperforming cyclosporin A by a factor of at least 2 in the inhibition of Pgp-mediated daunomycin transport.

  10. Cyclic peptide oral bioavailability: Lessons from the past.

    PubMed

    Wang, Conan K; Craik, David J

    2016-11-01

    Achieving high oral bioavailability for drugs is a key design objective in drug development. It is not surprising then that with the growing expectation of peptides as future drugs, there has also been an increasing interest in developing oral peptide therapeutics. Brought to the fore are questions such as what makes peptides orally bioavailable and how this can be achieved; questions which have inspired research into the area for decades. Early research in the area focused on linear peptides with more recent literature focusing on cyclic peptides, motivated in part by cyclic peptides like cyclosporine A that have demonstrated drug-like oral bioavailability. In this review, we take a look at research on the oral bioavailability of peptides, focusing on factors that affect passive permeability. © 2016 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 106: 901-909, 2016.

  11. Complexity of the primary genetic response to mitogenic activation of human T cells

    SciTech Connect

    Zipfel, P.F.; Siebenlist, U. ); Irving, S.G.; Kelly, K. )

    1989-03-01

    The authors describe the isolation and characterization of more than 60 novel cDNA clones that constitute part of the immediate genetic response to resting human peripheral blood T cells after mitogen activation. This primary response was highly complex, both in the absolute number of inducible genes and in the diversity of regulation. Although most of the genes expressed in activated T cells were shared with the activation response of normal human fibroblasts, a significant number were more restricted in tissue specificity and thus likely encode or effect the differentiated functions of activated T cells. The activatable genes could be further differentiated on the basis of kinetics of induction, response to cycloheximide, and sensitivity to the immunosuppressive drug cylcosporin A. It is of note that cyclosporin A inhibited the expression of more than 10 inducible genes, which suggests that this drug has a broad genetic mechanism of action.

  12. Excessive naked megakaryocyte nuclei in myelodysplastic syndrome mimicking idiopathic thrombocytopenic purpura: a complicated pre- and post-transplantation course.

    PubMed

    Olcay, Lale; Tuncer, A Murat; Okur, Hamza; Erdemli, Esra; Uysal, Zumrut; Cetin, Mualla; Duru, Feride; Cetinkaya, Duygu Uckan

    2009-09-01

    A boy 3 years 7 months old with thrombocytopenia and history of intracranial hemorrhage who underwent bone marrow transplantation is presented. He was refractory to steroids, immunoglobulin G, vincristine, azathioprine, cyclosporine A, interleukin-11, chemotherapy, and splenectomy. Idiopathic thrombocytopenic purpura was excluded by light /electron microscopic and flow cytometric findings; the diagnosis of refractory cytopenia, a subgroup of pediatric myelodysplastic syndrome, was made. Naked megakaryocyte nuclei were 55.38 +/- 28.2% vs. 31.67 +/- 23.22% of all megakaryocytes in the patient and the control group of 9 patients with idiopathic thrombocytopenic purpura, respectively (p = .016). The posttransplatation course was complicated by delayed platelet engraftment, bronchiolitis obliterans associated with pneumocystis carinii pneumonia, which resolved completely.

  13. Use of Cyclosporine in Uterine Transplantation

    PubMed Central

    Saso, Srdjan; Logan, Karl; Abdallah, Yazan; Louis, Louay S.; Ghaem-Maghami, Sadaf; Smith, J. Richard; Del Priore, Giuseppe

    2012-01-01

    Uterine transplantation has been proposed as a possible solution to absolute uterine factor infertility untreatable by any other option. Since the first human attempt in 2000, various teams have tried to clarify which immunosuppressant would be most suitable for protecting the allogeneic uterine graft while posing a minimal risk to the fetus. Cyclosporine A (CsA) is an immunosuppressant widely used by transplant recipients. It is currently being tested as a potential immunosuppressant to be used during UTn. Its effect on the mother and fetus and its influence upon the graft during pregnancy have been of major concern. We review the role of CsA in UTn and its effect on pregnant transplant recipients and their offspring. PMID:22132302

  14. Blood-brain barrier permeability of ginkgolide: Comparison of the behavior of PET probes 7α-[(18)F]fluoro- and 10-O-p-[(11)C]methylbenzyl ginkgolide B in monkey and rat brains.

    PubMed

    Doi, Hisashi; Sato, Kengo; Shindou, Hideo; Sumi, Kengo; Koyama, Hiroko; Hosoya, Takamitsu; Watanabe, Yasuyoshi; Ishii, Satoshi; Tsukada, Hideo; Nakanishi, Koji; Suzuki, Masaaki

    2016-11-01

    The blood-brain barrier permeability of ginkgolide B was examined using positron emission tomography (PET) probes of a (18)F-incorporated ginkgolide B ([(18)F]-2) and a (11)C-incorporated methylbenzyl-substituted ginkgolide B ([(11)C]-3). PET studies in monkeys showed low uptake of [(18)F]-2 into the brain, but small amounts of [(11)C]-3 were accumulated in the parenchyma. Furthermore, when cyclosporine A was preadministered to rats, the accumulation of [(18)F]-2 in the rat brain did not significantly change, however, the accumulation of [(11)C]-3 was five times higher than that in the control rat. These results provide effective approaches for investigating the drug potential of ginkgolides.

  15. Novel inhibitors of the calcineurin/NFATc hub - alternatives to CsA and FK506?

    PubMed Central

    Sieber, Matthias; Baumgrass, Ria

    2009-01-01

    The drugs cyclosporine A (CsA) and tacrolimus (FK506) revolutionized organ transplantation. Both compounds are still widely used in the clinic as well as for basic research, even though they have dramatic side effects and modulate other pathways than calcineurin-NFATc, too. To answer the major open question - whether the adverse side effects are secondary to the actions of the drugs on the calcineurin-NFATc pathway - alternative inhibitors were developed. Ideal inhibitors should discriminate between the inhibition of (i) calcineurin and peptidyl-prolyl cis-trans isomerases (PPIases; the matchmaker proteins of CsA and FK506), (ii) calcineurin and the other Ser/Thr protein phosphatases, and (iii) NFATc and other transcription factors. In this review we summarize the current knowledge about novel inhibitors, synthesized or identified in the last decades, and focus on their mode of action, specificity, and biological effects. PMID:19860902

  16. Discovery of 7-N-piperazinylthiazolo[5,4-d]pyrimidine analogues as a novel class of immunosuppressive agents with in vivo biological activity.

    PubMed

    Jang, Mi-Yeon; Lin, Yuan; De Jonghe, Steven; Gao, Ling-Jie; Vanderhoydonck, Bart; Froeyen, Mathy; Rozenski, Jef; Herman, Jean; Louat, Thierry; Van Belle, Kristien; Waer, Mark; Herdewijn, Piet

    2011-01-27

    Herein we describe the synthesis and in vitro and in vivo activity of thiazolo[5,4-d]pyrimidines as a novel class of immunosuppressive agents, useful for preventing graft rejection after organ transplantation. This research resulted in the discovery of a series of compounds with potent activity in the mixed lymphocyte reaction (MLR) assay, which is well-known as the in vitro model for in vivo rejection after organ transplantation. The most potent congeners displayed IC(50) values of less than 50 nM in this MLR assay and hence are equipotent to cyclosporin A, a clinically used immunosuppressive drug. One representative of this series was further evaluated in a preclinical animal model of organ transplantation and showed excellent in vivo efficacy. It validates these compounds as new promising immunosuppressive drugs.

  17. Synthesis, biological evaluation of chrysin derivatives as potential immunosuppressive agents.

    PubMed

    Lv, Peng-Cheng; Cai, Tian-Tian; Qian, Yong; Sun, Juan; Zhu, Hai-Liang

    2011-01-01

    A series of novel chrysin derivatives was firstly synthesized and evaluated on their immunosuppressive activity in the search for potential immunosuppressive agents. Among them, compounds 5c displayed the most potent immunosuppressive inhibitory activity with IC(50) of 0.78 μM, which was comparable to that of cyclosporin A (IC(50) = 0.06 μM). The preliminary mechanism of compound 5c inhibition effects was also detected by flow cytometry (FCM), and the compound exerted immunosuppressive activity via inducing the apoptosis of activated lymph node cells in a dose dependent manner. Furthermore, the estimated LD(50) (in mg/kg) in vivo of compound 5c is 738.2, which indicated that compound 5c was low toxic.

  18. Inhibition of the MDR1 transporter by new phenothiazine derivatives.

    PubMed

    Kónya, Attila; Andor, Attila; Sátorhelyi, Péter; Németh, Klára; Kurucz, István

    2006-07-21

    The MDR1 transporter mediated efflux of different xenobiotics out of the cells serves as the most important mechanisms of the multidrug resistance in cancer cells, thus inhibition of the MDR1 transporter may increase the efficiency of anticancer drugs in the therapy. Here we describe some new phenothiazine derivatives, which possess strong in vitro MDR1 inhibitory activity. The effectiveness of the compounds on the MDR1 mediated calcein-AM efflux, ATPase activity, and colchicine resistance was proven by microplate assays and flow cytometry using recombinant and control cell lines. Some of these derivatives were more active than verapamil and one of them was at least as active as cyclosporin A. According to our results the new structural elements built in these phenothiazine type compounds increased their MDR1 inhibitory activity, which may serve as a basis of the development of an effective MDR1 inhibitor drug.

  19. Generation of PDGFRα+ Cardioblasts from Pluripotent Stem Cells

    PubMed Central

    Hong, Seon Pyo; Song, Sukhyun; Cho, Sung Woo; Lee, Seungjoo; Koh, Bong Ihn; Bae, Hosung; Kim, Kyun Hoo; Park, Jin-Sung; Do, Hyo-Sang; Im, Ilkyun; Heo, Hye Jin; Ko, Tae Hee; Park, Jae-Hyeong; Youm, Jae Boum; Kim, Seong-Jin; Kim, Injune; Han, Jin; Han, Yong-Mahn; Koh, Gou Young

    2017-01-01

    Isolating actively proliferating cardioblasts is the first crucial step for cardiac regeneration through cell implantation. However, the origin and identity of putative cardioblasts are still unclear. Here, we uncover a novel class of cardiac lineage cells, PDGFRα+Flk1− cardioblasts (PCBs), from mouse and human pluripotent stem cells induced using CsAYTE, a combination of the small molecules Cyclosporin A, the rho-associated coiled-coil kinase inhibitor Y27632, the antioxidant Trolox, and the ALK5 inhibitor EW7197. This novel population of actively proliferating cells is cardiac lineage–committed but in a morphologically and functionally immature state compared to mature cardiomyocytes. Most important, most of CsAYTE-induced PCBs spontaneously differentiated into functional αMHC+ cardiomyocytes (M+CMs) and could be a potential cellular resource for cardiac regeneration. PMID:28165490

  20. Calcium ions affect the hepatitis B virus core assembly

    SciTech Connect

    Choi, Yongwook; Gyoo Park, Sung; Yoo, Jun-hi; Jung, Guhung . E-mail: drjung@snu.ac.kr

    2005-02-05

    Previous report showed that cytosolic Ca{sup 2+} induced by hepatitis B virus X protein (HBx) promotes HBV replication. In this study, in vitro experiments showed that (i) HBV core assembly in vitro was promoted by Ca{sup 2+} through the sucrose density gradient and the analytical ultracentrifuge analysis. Also (ii) transmission electron microscope analysis demonstrated these assembled HBV core particles were the capsids. Ex vivo experiments showed that the treatment of BAPTA-AM and cyclosporine A (CsA) reduced HBV capsids in the transfected HepG2 cells. In addition to that, the treatment of Thapsigargin (TG) increased HBV capsids in the transfected HepG2 cells. Furthermore, we investigated the increased HBV core assembly by HBx. The results show that the increased cytosolic calcium ions by HBx promote the HBV core assembly.

  1. Reversible posterior leucoencephalopathy syndrome associated with bone marrow transplantation.

    PubMed

    Teive, H A; Brandi, I V; Camargo, C H; Bittencourt, M A; Bonfim, C M; Friedrich, M L; de Medeiros, C R; Werneck, L C; Pasquini, R

    2001-09-01

    Reversible posterior leucoencephalopathy syndrome (RPLS) has previously been described in patients who have renal insufficiency, eclampsia, hypertensive encephalopathy and patients receiving immunosuppressive therapy. The mechanism by which immunosuppressive agents can cause this syndrome is not clear, but it is probably related with cytotoxic effects of these agents on the vascular endothelium. We report eight patients who received cyclosporine A (CSA) after allogeneic bone marrow transplantation or as treatment for severe aplastic anemia (SSA) who developed posterior leucoencephalopathy. The most common signs and symptoms were seizures and headache. Neurological dysfunction occurred preceded by or concomitant with high blood pressure and some degree of acute renal failure in six patients. Computerized tomography studies showed low-density white matter lesions involving the posterior areas of cerebral hemispheres. Symptoms and neuroimaging abnormalities were reversible and improvement occurred in all patients when given lower doses of CSA or when the drug was withdrawn. RPLS may be considered an expression of CSA neurotoxicity.

  2. [Subcutaneous phaeohyphomycose due to Exophiala jeanselmir. Report of 3 cases in patients with a kidney transplant].

    PubMed

    Sabbaga, E; Tedesco-Marchesi, L M; Lacaz, C da S; Cucé, L C; Salebian, A; Heins-Vaccari, E M; Sotto, M N; Valente, N Y; Porto, E; Levy Neto, M

    1994-01-01

    We report three cases of subcutaneous phaeohyphomycosis due to Exophiala jeanselmei (Langeron) McGinnis et Padhye 1977, in kidney transplant patients. Exophiala jeanselmei is a dematious fungus having also ability to rarely procedure eumycetoma (black grains). According to KWON-CHUNG & BENNETT (1992) such fungus is antigenically very heterogeneous, since so far three serotypes have been identified; each serotype including subgroups. Subcutaneous phaeohyphomycosis is becoming more and more frequent in kidney transplant patients submitted to an immunosuppressive treatment. As Exophiala jeanselmei has already been isolated from the environment it becomes difficult to explain the pathogenicity of these cases by a reactivation of quiescent processes. The authors suggest an occasional fungistatic action of cyclosporine A upon Exophiala jeanselmei.

  3. Cost Effectiveness Analysis of Disease-Modifying Antirheumatic Drugs in Rheumatoid Arthritis. A Systematic Review Literature

    PubMed Central

    Benucci, Maurizio; Saviola, Gianantonio; Manfredi, Mariangela; Sarzi-Puttini, Piercarlo; Atzeni, Fabiola

    2011-01-01

    The cost effectiveness of treatments that have changed the “natural history” of a chronic progressive disease needs to be evaluated over the long term. Disease-modifying antirheumatic drugs (DMARDs) are the standard treatment of rheumatoid arthritis (RA) and should be started as early as possible. A number of studies have shown that they are effective in improving disease activity and function, and in joint damage. Our review was focused on revision and critical evaluation of the studies including the literature on cost effectiveness of DMARDs (cyclosporine A, sulphasalazine, leflunomide, and methotrexate). The European League Against Rheumatism (EULAR) recommendations showed that traditional DMARDs are cost effective at the time of disease onset. They are less expensive than biological DMARDs and can be useful in controlling disease activity in early RA. PMID:22162693

  4. Multifocal osteonecrosis in a patient with anamnestic ulcerative colitis. Is there a relationship with the disease and the use of glucocorticoids twenty years before? A brief review of the literature.

    PubMed

    Saviola, G; Abdi-Ali, L; Sacco, S; Dalle Carbonare, L G

    2016-01-01

    In 2013 a 40 year old man came to visit in our Rheumatology Unit because of a recent bilateral shoulder and hip pain. He had been treated from 1990 to 2000 with Cyclosporin A and Sulfasalazyn because of an ulcerative colitis which was completely in remission from 2000 . Glucocorticoids at the mean daily dose of 50 mg were administered only in the first period (1990-92). X-plain rays showed a suspicious multifocal osteonecrosis of both femoral and humeral heads. Magnetic Resonance confirmed the diagnosis (stage III and IV following Ficat and Arlet's criteria). The patient was treated with a cycle of hyperbaric oxygen therapy, with two cycles of intravenous clodronate and with a 2-month cycle of teriparatide. The treatment was able to save a sufficient function for both shoulders, while for both hips arthroplasty the surgery was required. The risks of osteonecrosis linked to inflammatory bowel diseases or to its therapy are discussed.

  5. Bioprinting of 3D Convoluted Renal Proximal Tubules on Perfusable Chips

    PubMed Central

    Homan, Kimberly A.; Kolesky, David B.; Skylar-Scott, Mark A.; Herrmann, Jessica; Obuobi, Humphrey; Moisan, Annie; Lewis, Jennifer A.

    2016-01-01

    Three-dimensional models of kidney tissue that recapitulate human responses are needed for drug screening, disease modeling, and, ultimately, kidney organ engineering. Here, we report a bioprinting method for creating 3D human renal proximal tubules in vitro that are fully embedded within an extracellular matrix and housed in perfusable tissue chips, allowing them to be maintained for greater than two months. Their convoluted tubular architecture is circumscribed by proximal tubule epithelial cells and actively perfused through the open lumen. These engineered 3D proximal tubules on chip exhibit significantly enhanced epithelial morphology and functional properties relative to the same cells grown on 2D controls with or without perfusion. Upon introducing the nephrotoxin, Cyclosporine A, the epithelial barrier is disrupted in a dose-dependent manner. Our bioprinting method provides a new route for programmably fabricating advanced human kidney tissue models on demand. PMID:27725720

  6. Rule of five in 2015 and beyond: Target and ligand structural limitations, ligand chemistry structure and drug discovery project decisions.

    PubMed

    Lipinski, Christopher A

    2016-06-01

    The rule of five (Ro5), based on physicochemical profiles of phase II drugs, is consistent with structural limitations in protein targets and the drug target ligands. Three of four parameters in Ro5 are fundamental to the structure of both target and drug binding sites. The chemical structure of the drug ligand depends on the ligand chemistry and design philosophy. Two extremes of chemical structure and design philosophy exist; ligands constructed in the medicinal chemistry synthesis laboratory without input from natural selection and natural product (NP) metabolites biosynthesized based on evolutionary selection. Exceptions to Ro5 are found mostly among NPs. Chemistry chameleon-like behavior of some NPs due to intra-molecular hydrogen bonding as exemplified by cyclosporine A is a strong contributor to NP Ro5 outliers. The fragment derived, drug Navitoclax is an example of the extensive expertise, resources, time and key decisions required for the rare discovery of a non-NP Ro5 outlier.

  7. Successful Treatment of Ascites using a DenverⓇ Peritoneovenous Shunt in a Patient with Paroxysmal Nocturnal Hemoglobinuria and Budd-Chiari syndrome

    PubMed Central

    Kogiso, Tomomi; Hashimoto, Etsuko; Ito, Taito; Hara, Toshifumi; Ikarashi, Yuichi; Kodama, Kazuhisa; Taniai, Makiko; Torii, Nobuyuki; Yoshinaga, Kentaro; Morita, Satoru; Takahashi, Yutaka; Tanaka, Junji; Sakai, Shuji; Yamamoto, Masakazu; Tokushige, Katsutoshi

    2016-01-01

    A 56-year-old man was diagnosed with aplastic anemia and paroxysmal nocturnal hemoglobinuria at 43 years of age and treatment with cyclosporin A was started. Liver cirrhosis, ascites, and thrombus in the hepatic veins were found at 56 years of age and Budd-Chiari syndrome (BCS) was diagnosed according to angiography findings. He was treated with diuretics and paracentesis was performed several times, but with limited efficacy. A DenverⓇ peritoneovenous shunt (PVS) was inserted into the right jugular vein; his ascites and renal function improved immediately and his general condition has remained good for 12 months since starting the above treatment regimen. A PVS is a treatment option for ascites due to BCS. PMID:27746432

  8. Newer treatments for inflammatory bowel disease.

    PubMed

    Stotland, B R; Lichtenstein, G R

    1998-02-01

    Inflammatory bowel disease represents chronic idiopathic disorders which involve either the colon exclusively (ulcerative colitis) of any part of the gastrointestinal tract (Crohn's disease). The course of these entities is typified by periods of symptomatic exacerbation interspersed with clinical remissions. Management is based upon regimens which decrease mucosal inflammation. Colonic disease distal to the splenic flexure may be treated with topical therapy, but other regions generally necessitate oral therapy. Currently used medications include the aminosalicylates, glucocorticoids, antibiotics and immunomodulators. The immunomodulator class of medications includes azathioprine, 6-mercaptopurine, cyclosporine A and methotrexate. Newer agents include short-chain fatty acids, omega-3 fatty acids and antibodies directed to tumor necrosis factor. Medical management also occasionally involves optimizing nutritional status with the addition of elemental diets or total parenteral nutrition. Management of specific clinical presentations is discussed.

  9. Pharmacokinetics of PSC 833 (valspodar) in its Cremophor EL formulation in rat.

    PubMed

    Binkhathlan, Z; Hamdy, D A; Brocks, D R; Lavasanifar, A

    2010-01-01

    Valspodar is a P-glycoprotein inhibitor widely used in preclinical and clinical studies for overcoming multidrug resistance. Despite this, the pharmacokinetics of valspodar in rat, a commonly used animal model, have not been reported. Here, we report on the pharmacokinetics of valspodar in Sprague-Dawley rats following intravenous and oral administration of its Cremophor EL formulation, which has been used for humans in clinical trials. After intravenous doses, valspodar displayed properties of slow clearance and a large volume of distribution. Its plasma unbound fraction was around 15% in the Cremophor EL formulation used in the study. After 10 mg kg(-1) orally it was rapidly absorbed with an average maximal plasma concentration of 1.48 mg l(-1) within approximately 2 h. The mean bioavailability of valspodar was 42.8%. In rat, valspodar showed properties of low hepatic extraction and wide distribution, similar to that of its structural analogue cyclosporine A.

  10. Taste-immunosuppression engram: reinforcement and extinction.

    PubMed

    Niemi, Maj-Britt; Härting, Margarete; Kou, Wei; Del Rey, Adriana; Besedovsky, Hugo O; Schedlowski, Manfred; Pacheco-López, Gustavo

    2007-08-01

    Several Pavlovian conditioning paradigms have documented the brain's abilities to sense immune-derived signals or immune status, associate them with concurrently relevant extereoceptive stimuli, and reinstate such immune responses on demand. Specifically, the naturalistic relation of food ingestion with its possible immune consequences facilitates taste-immune associations. Here we demonstrate that the saccharin taste can be associated with the immunosuppressive agent cyclosporine A, and that such taste-immune associative learning is subject to reinforcement. Furthermore, once consolidated, this saccharin-immunosuppression engram is resistant to extinction when avoidance behavior is assessed. More importantly, the more this engram is activated, either at association or extinction phases, the more pronounced is the conditioned immunosuppression.

  11. [Chronic myeloid leukemia after renal transplantation: report of a new case and review of the bibliography].

    PubMed

    Sanz, L; Cervantes, F; Esteve, J; Vilardell, J; Marín, P; Rozman, C; Montserrat, E

    1996-10-01

    The increase in cancer incidence in renal transplant recipients is a well recognized fact, which has been related to post-transplant immunosuppressive therapy. Solid tumors, skin cancer and non-Hodgkin's lymphomas account for most of the neoplasms in these patients, whereas chronic myeloproliferative disorders are infrequent. A patient is reported who developed chronic myelogenous leukemia (CML) six years after renal transplantation, which was followed by immunosuppressive with azathioprine, and the published data on such an association are reviewed. In all 10 cases reported azathioprine had been administered after transplantation. The amount and type of post-transplant immunosuppressive therapy seems to be the most important risk factor for the development of secondary CML in these patients, since no cases of CML in patients receiving cyclosporine A have been reported.

  12. Interaction of calcineurin with substrates and targeting proteins

    PubMed Central

    Li, Huiming; Rao, Anjana; Hogan, Patrick G.

    2011-01-01

    Calcineurin is a calcium-activated protein phosphatase with a major role in calcium signaling in diverse cells and organs, and importance in clinical practice as the target of the immunosuppressive drugs cyclosporin A and FK506. Cell biological studies have focused mainlyy on the role of calcineurin in transcriptional signaling. Calcium entry in response to extracellular stimuli results in calcineurin activation and signal transmission from the cytosol into the nucleus through dephosphorylation and nuclear translocation of the transcription factor NFAT. This initiates a cascade of transcriptional events involved in physiological and developmental processes. Molecular analyses of the calcineurin/NFAT interaction have been extended recently to encompass the interaction of calcineurin with other substrates, targeting proteins and regulators of calcineurin activity. These studies have increased our understanding of how this essential calcium-activated enzyme orchestrates intracellular events in cooperation with other signaling pathways, and suggested a link between altered calcineurin signaling and the developmental anomalies of Down syndrome. PMID:21115349

  13. Drugs to cure avian influenza infection – multiple ways to prevent cell death

    PubMed Central

    Yuan, S

    2013-01-01

    New treatments and new drugs for avian influenza virus (AIV) infection are developed continually, but there are still high mortality rates. The main reason may be that not all cell death pathways induced by AIV were blocked by the current therapies. In this review, drugs for AIV and associated acute respiratory distress syndrome (ARDS) are summarized. The roles of antioxidant (vitamin C) and multiple immunomodulators (such as Celecoxib, Mesalazine and Eritoran) are discussed. The clinical care of ARDS may result in ischemia reperfusion injury to poorly ventilated alveolar cells. Cyclosporin A should effectively inhibit this kind of damages and, therefore, may be the key drug for the survival of patients with virus-induced ARDS. Treatment with protease inhibitor Ulinastatin could also protect lysosome integrity after the infection. Through these analyses, a large drug combination is proposed, which may hypothetically greatly reduce the mortality rate. PMID:24091678

  14. Intravenous immunoglobulin therapy for resistant ocular Behçet's disease

    PubMed Central

    Seider, N.; Beiran, I.; Scharf, J.; Miller, B.

    2001-01-01

    AIMS—The present report was aimed at finding out whether gammaglobulin could have a role in treating ocular Behçet's disease (BD) refractory to accepted medical therapy.
METHODS—Six eyes of four patients with ocular BD refractory to steroids and cyclosporin A were treated with a course of intravenous gammaglobulin and followed up for their response to treatment.
RESULTS—All six eyes of all four patients showed good response to gammaglobulin therapy.
CONCLUSION—Gamma globulin may have a role in treating refractory ocular BD. A wide range of controlled studies with longer follow up is needed to substantiate this impression.

 PMID:11673289

  15. Lifitegrast: A novel drug for treatment of dry eye disease

    PubMed Central

    Abidi, Afroz; Shukla, Pooja; Ahmad, Ali

    2016-01-01

    Dry eye disease (DED) is an inflammatory disorder of ocular surfaces leading to severe disability, especially in the elderly age group. The mainstay of therapy includes artificial tears, punctual plugs, topical anti-inflammatory agents, and corticosteroids. In the past few years, only cyclosporine-A emulsions have been added to the existing therapy, but it is discontinued by most patients as it causes burning sensation in the eye. Hence, progress in new research for a better therapeutic option led to the discovery of lymphocyte function-associated antigen intercellular adhesion molecule 1 antagonist, lifitegrast. It hinders the T-cell activation, release of inflammatory mediators, and consequently inhibits the inflammatory pathways in DED. It was approved by the US Food and Drug Administration in July 2016 for the treatment of DED. This review highlights the development process and approval of lifitegrast. PMID:28163544

  16. Fragment-based discovery of a new family of non-peptidic small-molecule cyclophilin inhibitors with potent antiviral activities

    PubMed Central

    Ahmed-Belkacem, Abdelhakim; Colliandre, Lionel; Ahnou, Nazim; Nevers, Quentin; Gelin, Muriel; Bessin, Yannick; Brillet, Rozenn; Cala, Olivier; Douguet, Dominique; Bourguet, William; Krimm, Isabelle; Pawlotsky, Jean-Michel; Guichou, Jean- François

    2016-01-01

    Cyclophilins are peptidyl-prolyl cis/trans isomerases (PPIase) that catalyse the interconversion of the peptide bond at proline residues. Several cyclophilins play a pivotal role in the life cycle of a number of viruses. The existing cyclophilin inhibitors, all derived from cyclosporine A or sanglifehrin A, have disadvantages, including their size, potential for side effects unrelated to cyclophilin inhibition and drug–drug interactions, unclear antiviral spectrum and manufacturing issues. Here we use a fragment-based drug discovery approach using nucleic magnetic resonance, X-ray crystallography and structure-based compound optimization to generate a new family of non-peptidic, small-molecule cyclophilin inhibitors with potent in vitro PPIase inhibitory activity and antiviral activity against hepatitis C virus, human immunodeficiency virus and coronaviruses. This family of compounds has the potential for broad-spectrum, high-barrier-to-resistance treatment of viral infections. PMID:27652979

  17. [Endovascular thrombolysis for massive cerebral venous thrombosis in a teenager with nephrotic syndrome].

    PubMed

    Costa, Paula; Biscoito, Luísa; Vieira, Marisa; Marçal, Mónica; Camilo, Cristina; Neto, Lia; Abecasis, Francisco; Almeida, Margarida; Correia, Manuela

    2010-01-01

    Cerebral venous thrombosis is a rare but potentially severe condition in children. We present the case of a teenager with corticodependent nephrotic syndrome diagnosed at five months of age and treated with cyclosporine A. In the context of recurrence of nephrotic syndrome he presented with headache, vomiting and severe intracranial hypertension. While the raised intracranial pressure and the status epilepticus were controlled, the brain imaging revealed venous thrombosis of all venous sinus, with absence of venous drainage. He was submitted to local thrombolysis with recombinant tissue plasminogen activator, with recanalization of the venous sinuses. The outcome was favourable, without neurological deficits. In this case, the early radiologic intervention was crucial, enabling a full neurological recovery, in a teenager whose initial prognosis was very poor.

  18. Antidotal Effects of Curcumin Against Agents-Induced Cardiovascular Toxicity.

    PubMed

    Farkhondeh, Tahereh; Samarghandian, Saeed

    Curcumin, the major phenolic compound in turmeric, shows preventive effects in various diseases. Curcumin is commonly found in rhizome of the Curcuma species and traditionally used in herbal medicine. Numeros studies has indicated that curcumin posses protective effects against toxic agents in various systems including cardiovascular. This study found that curcumin may be effective in cardiovascular diseases induced by toxic agents including Streptozotocin, Doxorubicin, Cyclosporin A, Methotrexate, Isoproterenol, Cadmium, Diesel exhaust particle, Nicotine, Hydrogen peroxide, and tert- Butyl hydroperoxide. However, due to the lake of information on human, further studies are needed to determine the efficacy of curcumin as an antidote agent. The present study aimed to critically review the recent literature data from that regarding the protective effects of curcumin against agents-induced cardiovascular toxicity.

  19. Quantitative analysis of biomolecules by time-of-flight secondary-ion mass spectrometry: Fundamental considerations

    SciTech Connect

    Muddiman, D.C.; Nicola, A.J.; Proctor, A.

    1995-12-31

    Static Time-of-Flight Secondary-Ion Mass Spectrometry (TOF-SIMS) has been applied to investigate an extensive assortment of analytical systems; from semiconductors to DNA sequencing. Recently, the TOF-SIMS method has been successfully applied to real biological systems. This report focuses on some important aspects that must be taken into consideration when conducting measurements on biomaterials in order to observe the potential the TOF-SIMS method affords. The current data are presented using Cyclosporin A (CsA, 1202 Da) and cocaine (303 Da) as model compounds. CsA is observed in the TOF-SIMS mass spectrum predominately as a Ag-cationized species and cocaine as a protonated species; thus, they are complementary probe molecules.

  20. Hematopoietic Stem Cell Transplantation Nephropathy Associated with Chronic Graft-versus-Host Disease without Extrarenal Involvement

    PubMed Central

    Ishida, Ryo; Shimizu, Akira; Kitani, Takashi; Nakata, Mayumi; Ota, Noriyoshi; Kado, Hiroshi; Shiotsu, Yayoi; Ishida, Mami; Tamagaki, Keiichi

    2016-01-01

    A 30-year-old woman with myelodysplastic syndrome underwent allogeneic hematopoietic stem cell transplantation (HSCT) derived from her HLA-matched sister six years previously. She received preconditioning total body irradiation with renal shielding and was subsequently administered cyclosporin A (CyA) as prophylaxis against graft-versus-host disease (GVHD). Four months after HSCT, asymptomatic proteinuria and glomerular hematuria developed during CyA tapering without obvious extrarenal involvements of GVHD, and persisted for six years. A renal biopsy revealed endothelial injury in the glomeruli, and the deposition of C4d was detected diffusely on glomerular capillaries and focally on peritubular capillaries, suggesting that nephropathy involved antibody- or complement-associated immune reactions. PMID:27725545

  1. Peripheral Ulcerative Keratitis with Pyoderma Gangrenosum

    PubMed Central

    Imbernón-Moya, Adrián; Vargas-Laguna, Elena; Aguilar, Antonio; Gallego, Miguel Ángel; Vergara, Claudia; Nistal, María Fernanda

    2015-01-01

    Pyoderma gangrenosum is an unusual necrotizing noninfective and ulcerative skin disease whose cause is unknown. Ophthalmic involvement in pyoderma gangrenosum is an unusual event. Only a few cases have been reported, from which we can highlight scleral, corneal, and orbital cases. Peripheral ulcerative keratitis is a process which destroys the peripheral cornea. Its cause is still unknown although it is often associated with autoimmune conditions. Pyoderma gangrenosum should be included in the differential diagnosis of peripheral ulcerative keratitis. Early recognition of these manifestations can vary the prognosis by applying the appropriate treatment. We introduce a 70-year-old woman who suffered pyoderma gangrenosum associated with peripheral ulcerative keratitis in her left eye. The patient's skin lesions and peripheral keratitis responded successfully to systemic steroids and cyclosporine A. PMID:26527531

  2. Induction of bilateral ligneous conjunctivitis with the use of a prosthetic eye.

    PubMed

    Yazıcı, Bülent; Yıldız, Meral; Irfan, Tayfun

    2011-02-01

    The purpose of this article to report a case of ligneous conjunctivitis in an anophthalmic socket, in respect of a 20-year-old woman. The subject woman had a history of left enucleation surgery presented with bilateral palpebral ligneous conjunctivitis and ligneous gingivitis. The hematologic study revealed a severe plasma plasminogen deficiency. The eyelid lesions were successfully treated with surgical excision, topical heparin and corticosteroid eyedrops. However, the ligneous lesions recurred bilaterally after she was fitted with a prosthetic eye and were refractory to intensive topical treatment with heparin and cyclosporin A eye drops. This case shows that the use of a prosthetic eye may induce ligneous conjunctivitis in an anophthalmic socket and normal eye which is refractory to topical treatment.

  3. Drug interaction in a renal transplant patient: cyclosporin-Neoral and orlistat.

    PubMed

    Evans, Sally; Michael, Robson; Wells, Hayley; Maclean, Douglas; Gordon, Isabel; Taylor, John; Goldsmith, David

    2003-02-01

    An overweight 56-year-old type II diabetic on peritoneal dialysis (body mass index 35 kg/m(2)) was taking Orlistat for some months up until live-unrelated renal transplantation. Despite oral cyclosporin A (CyA) for 48 hours pretransplantation, it was very difficult to achieve adequate CyA blood levels for the first week postengraftment despite the use of much larger oral CyA doses. After opening his bowels on day 7, and the use of 3 days intravenous CyA, good CyA blood levels were achieved then maintained with conventional oral doses. The authors believe that this case shows important interactions between CyA and Orlistat.

  4. PROTECTIVE EFFECTS OF POTASSIUM TRANSPORT IN MITOCHONDRIA FROM RAT MYOMETRIUM UNDER ACTIVATION OF MITOCHONDRIAL PERMEABILITY TRANSITION PORE.

    PubMed

    Vadzyuk, O B

    2015-01-01

    We demonstrated using PBFI K(+)-sensitive fluorescent probe an enhancement of both components of K(+)-cycle--the ATP-sensitive K(+)-uptake and quinine-sensitive K+/H(+)-exchange--under the Ca2+ induced opening of mitochondrial permeability transition pore (MPTP) in rat myometrium mitochondria. Addition of CaCl2 (100 μM to K(+)-free medium results in the enhancement of reactive oxygen species (ROS) production, which was eliminated by cyclosporine A. Addition of CaCl2 to K(+)-rich medium did not increase the rate of ROS production, but blocking of mitoK+(ATP)-channels with glybenclamide (10 mcM increased production of ROS. We conclude that K(+)-cycle exerts a protective influence in mitochondria from rat myometrium by regulation of matrix volume and rate of ROS production under the condition of Ca(2+)-induced MPTP.

  5. NFATc1 balances quiescence and proliferation of skin stem cells

    PubMed Central

    Horsley, Valerie; Aliprantis, Antonios O.; Polak, Lisa; Glimcher, Laurie H.; Fuchs, Elaine

    2008-01-01

    Quiescent adult stem cells reside in specialized niches where they become activated to proliferate and differentiate during tissue homeostasis and injury. How stem cell quiescence is governed is poorly understood. We report here that NFATc1 is preferentially expressed by hair follicle stem cells in their niche, where it's expression is activated by BMP signaling upstream and it acts downstream to transcriptionally repress CDK4 and maintain stem cell quiescence. As stem cells become activated during hair growth, NFATc1 is downregulated, relieving CDK4 repression and activating proliferation. When calcineurin/NFATc1 signaling is suppressed, pharmacologically or via complete or conditional NFATc1 gene ablation, stem cells are activated prematurely, resulting in precocious follicular growth. Our findings may explain why patients receiving cyclosporine A for immunosuppressive therapy display excessive hair growth, and unveil a functional role for calcium-NFATc1-CDK4 circuitry in governing stem cell quiescence. PMID:18243104

  6. Inhibitory effects of Cnidium monnieri fruit extract on pulmonary inflammation in mice induced by cigarette smoke condensate and lipopolysaccharide.

    PubMed

    Kwak, Ho-Geun; Lim, Heung-Bin

    2014-09-01

    The aim of this study was to investigate the inhibitory effect of Cnidium monnieri fruit (CM) extracts on pulmonary inflammation induced in mice by cigarette smoke condensate (CSC) and lipopolysaccharide (LPS). Pulmonary inflammation was induced by intratracheal instillation of LPS and CSC five times within 12 days. CM extract was administered orally at a dose of 50 or 200 mg·kg(-1). The number of inflammatory cells in the bronchoalveolar lavage fluid was counted using a fluorescence activated cell sorter. Inflammatory mediator levels were determined by enzyme-linked immunosorbent assay. The administration of LPS and CSC exacerbated airway hyper-responsiveness (AHR) and induced an accumulation of inflammatory cells and mediators, and led to histological changes. However, these responses are modulated by treatment with CM, and the treatment with CM extract produces similar or more extensive results than the treatment with cyclosporin A (CSA). CM extract may have an inhibitory effect on pulmonary inflammation related with chronic obstructive pulmonary disease.

  7. Bioprinting of 3D Convoluted Renal Proximal Tubules on Perfusable Chips

    NASA Astrophysics Data System (ADS)

    Homan, Kimberly A.; Kolesky, David B.; Skylar-Scott, Mark A.; Herrmann, Jessica; Obuobi, Humphrey; Moisan, Annie; Lewis, Jennifer A.

    2016-10-01

    Three-dimensional models of kidney tissue that recapitulate human responses are needed for drug screening, disease modeling, and, ultimately, kidney organ engineering. Here, we report a bioprinting method for creating 3D human renal proximal tubules in vitro that are fully embedded within an extracellular matrix and housed in perfusable tissue chips, allowing them to be maintained for greater than two months. Their convoluted tubular architecture is circumscribed by proximal tubule epithelial cells and actively perfused through the open lumen. These engineered 3D proximal tubules on chip exhibit significantly enhanced epithelial morphology and functional properties relative to the same cells grown on 2D controls with or without perfusion. Upon introducing the nephrotoxin, Cyclosporine A, the epithelial barrier is disrupted in a dose-dependent manner. Our bioprinting method provides a new route for programmably fabricating advanced human kidney tissue models on demand.

  8. An atypical presentation of adult-onset Still’s disease complicated by pulmonary hypertension and macrophage activation syndrome treated with immunosuppression: a case-based review of the literature

    PubMed Central

    Manson, Daniel K.; Horn, Evelyn M.; Haythe, Jennifer

    2016-01-01

    Abstract Pulmonary arterial hypertension (PAH) is a known complication of rheumatologic diseases, but it is only rarely associated with adult-onset Still’s disease (AOSD). We describe the case of a 30-year-old woman who presented in a pulmonary hypertension crisis and was found to have underlying AOSD with PAH and nonspecific interstitial pneumonia (NSIP) with a course complicated by macrophage activation syndrome (MAS). She dramatically improved with steroids, cyclosporine A, and anakinra, with total resolution of the MAS and significant improvement of her pulmonary arterial pressures. While there are only select case reports of AOSD associated with PAH, this is the first reported case of (1) AOSD complicated by both PAH and MAS and (2) AOSD complicated by biopsy-proven NSIP. Clinically, this case highlights the efficacy of immunosuppressive agents in the treatment of PAH and MAS from underlying AOSD and supports their use in this setting. PMID:27162622

  9. Differential modulation of apoptosis and necrosis by antioxidants in immunosuppressed human lymphocytes.

    PubMed

    Rojas, Mauricio; Rugeles, María Teresa; Gil, Diana Patricia; Patiño, Pablo

    2002-04-15

    In the present study, we explored whether mitogenic stimulation of dexamethasone (DXM)- and cyclosporine A (CsA)-immunosuppressed peripheral blood lymphocytes (PBML) induced apoptosis or necrosis and their relation with the production of reactive oxygen intermediates. Our results indicate that both phenomena can occur in these cells and that antioxidants such as N-acetyl cysteine (NAC) and ascorbic acid (AA) can modulate them. However, DXM-induced apoptosis was only partially inhibited by NAC and AA, suggesting that DXM-treated PBMC had an additional apoptotic pathway independent of ROIs. Furthermore, we observed that the inhibition of apoptosis by antioxidants correlated with an increased cell proliferation, suggesting that the immunomodulation of both DXM and CsA may be related to induction of apoptosis. The ability to differentially modulate apoptosis and necrosis by antioxidants opens new possibilities in the management of immunosuppressive therapy, since the inhibition of necrosis may avoid inflammation and the tissue damage associated with immunosuppressors.

  10. Autoimmune hemolytic anemia in patients with β-thalassemia major.

    PubMed

    Xu, Lu-Hong; Fang, Jian-Pei; Weng, Wen-Jun; Huang, Ke; Zhang, Ya-Ting

    2012-04-01

    Hemolysis is a common feature in patients with β-thalassemia major. As a result, autoimmune hemolytic anemia complicating β-thalassemia is easily overlooked. Here, the authors described the clinical features and management of 7 patients with β-thalassemia major and autoimmune hemolytic anemia. These patients had fever, cough, and tea-colored urine on admission. The laboratory investigations showed a significant drop in hemoglobin and increased serum bilirubin. Coombs' tests revealed that anti-immunoglobulin G (IgG) and anti-C3 was positive in 7 and 5 cases, respectively, whereas anti-Rh E alloantibody was positive in 3 cases. All the patients received corticosteroids treatments and blood transfusions. Patients with anti-Rh E alloantibodies also received immunoglobulin treatments. Six of the patients responded well to the management, but 1 patient developed recurrent autoimmune hemolytic anemia that required cyclosporin A treatment. All the patients remained well by following up for more than 6 months.

  11. Influence of SDZ RAD vs. MMF on gastric emptying in renal transplant recipients.

    PubMed

    Maes, Bart D; Evenepoel, Pieter; Kuypers, Dirk; Geypens, Benny; Ghoos, Yvo; Vanrenterghem, Yves

    2003-06-01

    SDZ RAD and mycophenolate mofetil (MMF) are increasingly used in the prevention of renal allograft rejection. SDZ RAD, having a macrolide structure, and MMF, known with gastrointestinal side-effects, may have gastric motility modifying properties. Gastric emptying was examined 1 yr after renal transplantation in eight patients taking corticosteroids (CS), cyclosporin A (CsA) and SDZ RAD and six patients treated with CS, CsA and MMF. Comparing the two groups, no significant differences in gastric emptying of solids and liquids were noted. Compared with normal volunteers, solid gastric emptying was faster in the SDZ RAD group and similar in the MMF group. It is concluded that in stable renal transplant recipients treated with MMF, gastric emptying was normal. Because of the impact on drug absorption and gastrointestinal symptoms, further studies are indicated to corroborate the potential prokinetic properties of SDZ RAD.

  12. MMF and eye disease.

    PubMed

    Zierhut, M; Stübiger, N; Siepmann, K; Deuter, C M E

    2005-01-01

    Immunosuppressive treatment has shown to be effective in various ocular inflammatory disorders. Factors limiting their use are the individual response and the rate of side effects. This report summarizes our knowledge about the use of mycophenolate mofetil (MMF) in the treatment of ocular cicatricial pemphigoid (OCP), uveitis, atopic keratoconjunctivitis (AKC), prevention of graft rejection after penetrating keratoplasty (PK) and scleritis. Controlled studies have been performed for prevention of graft rejection after PK, showing MMF as effective in the prevention of graft rejection as cyclosporine A. In experimental uveitis, MMF has been demonstrated to be highly effective in prevention of retinal destruction. A number of studies have now shown that MMF also seems effective in uveitis. There are also studies with smaller patient groups which point out the effectiveness of MMF in OCP, AKC, and scleritis. In most of the studies, the spectrum of side effects was small, compared to other immunosuppressives.

  13. The mitochondrial permeability transition pore provides a key to the diagnosis and treatment of traumatic brain injury

    PubMed Central

    Veech, Richard L.; Valeri, C. Robert; VanItallie, Theodore B.

    2012-01-01

    Summary The pathological consequences of traumatic head injury result largely from the opening of the mitochondrial permeability transition pore, mPTP. The mPTP opens due to a decrease in brain phosphorylation energy resulting in a further decrease in brain ATP production and a measurable increase in brain heat generation and temperature. The increase in brain temperature can be measured transcranially by near infrared spectroscopy which can be used to diagnoses TBI and to monitor treatment. Effective therapy of TBI can be achieved by closure of the mPTP by administration of cyclosporine A or by oral administration of ketone body esters. While ketosis has previously been known to prevent damage from TBI, the availability of oral ketone esters presents the first practical modality of achieving therapeutic levels of ketone bodies. PMID:22241645

  14. Study on Biological Effects of La(3+) on Rat Liver Mitochondria by Microcalorimetric and Spectroscopic Methods.

    PubMed

    Wu, Man; Gao, Jia-Ling; Feng, Zhi-Jiang; Liu, Wen; Zhang, Ye-Zhong; Liu, Yi; Dai, Jie

    2015-09-01

    The effects of lanthanum on heat production of mitochondria isolated from Wistar rat liver were investigated with microcalorimetry; simultaneously, the effects on mitochondrial swelling and membrane potential (Δψ) were determined by spectroscopic methods. La(3+) showed only inhibitory action on mitochondrial energy turnover with IC50 being 55.8 μmol L(-1). In the spectroscopic experiments, La(3+), like Ca(2+), induced rat liver mitochondrial swelling and decreased membrane potential (Δψ), which was inhibited by the specific permeability transition inhibitor, cyclosporine A (CsA). The induction ability of La(3+) was stronger than that of Ca(2+). These results demonstrated that La(3+) had some biotoxicity effect on mitochondria; the effects of La(3+) and Ca(2+) on rat liver mitochondrial membrane permeability transition (MPT) are different, and La represents toxic action rather than Ca analogy.

  15. Treatment of pure red-cell aplasia with cyclosporine in a renal transplant patient.

    PubMed

    Yildirim, Rahsan; Bilen, Yusuf; Keles, Mustafa; Uyanik, Abdullah; Gokbulut, Puren; Aydinli, Bulent

    2013-02-01

    Acquired pure red-cell aplasia is a rare disorder that can be either idiopathic or associated with certain autoimmune diseases, pregnancy, lymphoproliferative disorders, nutritional deficiencies, or medicines. We present a deceased-donor renal transplant patient who developed pure red-cell aplasia associated with mycophenolate mofetil or tacrolimus and was treated with cyclosporine. A 20-year-old woman was transplanted from a deceased donor 1 month earlier and presented to us with symptoms of fatigue, prostration, and palpitation. The results of a laboratory examination revealed anemia. A diagnostic work-up resulted in a diagnosis of pure red-cell aplasia. Mycophenolate mofetil was discontinued. Tacrolimus also was replaced with cyclosporine 2 months after mycophenolate mofetil was halted because of a lack of improvement in anemia. Three months later, her anemia improved with cyclosporine. Starting cyclosporine instead of tacrolimus or mycophenolate mofetil showed good improvement in our patient within 6 months of therapy.

  16. An analysis of the expression of cyclophilin C reveals tissue restriction and an intriguing pattern in the mouse kidney.

    PubMed Central

    Friedman, J.; Weissman, I.; Friedman, J.; Alpert, S.

    1994-01-01

    Cyclophilin C (cyp C) is a cyclosporin A (CsA) binding protein originally isolated from a mouse bone marrow stromal cell line. We have compared the expression patterns of the mammalian cyclophilins A, B, and C in mouse tissues using in situ hybridization. These studies reveal that cyp C is expressed in a restricted subset of tissues including mouse ovary, testis, bone marrow, and kidney. Within the kidney, cyp C is highly expressed in a narrow zone in the outer medulla. Using monoclonal antibodies reactive against cyp C, we find that the kidney cells expressing cyp C correspond to the S3 segment of the nephron. The S3 segment has been shown to sustain histopathological damage from high dosages of CsA, raising the possibility that cyp C may be involved in mediating this damage. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:8203464

  17. Mitochondrial localization of P-glycoprotein and peptide transporters in corneal epithelial cells--novel strategies for intracellular drug targeting.

    PubMed

    Barot, Megha; Gokulgandhi, Mitan R; Pal, Dhananjay; Mitra, Ashim K

    2013-01-01

    This study was designed to investigate functional localization of both efflux (P-glycoprotein, P-gp) and influx (peptide) transporters in the mitochondrial membrane of cultured rabbit primary corneal epithelial cells (rPCECs). Isolation and purification of mitochondria was performed by optimized cell fractionation method. Mitochondrial integrity was measured by JC-1 uptake experiment. The efflux activity of P-gp was assessed by performing in vitro uptake studies on isolated mitochondria with Rhodamine 123 (Rho-123) alone and in the presence of P-gp inhibitors (quinidine and cyclosporine A) using fluorimetry and flow cytometry analysis. Functional activity of peptide transporter was assessed by performing in vitro uptake studies of [3H] Gly-sar on isolated mitochondria in the presence or absence of peptide transporter substrate (Val-Val). Molecular characterization of P-gp and peptide transporter was assessed by western blot and confocal analysis. Enhanced JC-1 accumulation in the isolated fraction confirmed mitochondrial membrane integrity. Significantly higher uptake of Rho-123 on isolated mitochondria was observed in the presence of quinidine (75 and 100 μM) and cyclosporine A (10 μM). Significantly lower uptake of [3H] Gly-sar was observed in the presence of val-val due to competitive inhibition of peptide transporter on isolated mitochondria. Western blot and confocal analysis further confirmed the presence of P-gp and peptide transporter on the mitochondrial membrane of rPCECs. The present study demonstrates the functional and molecular characterization of P-gp and peptide transporters in the mitochondrial membranes of rPCECs. This knowledge of mitochondrial existence of P-gp and peptide transporter will aid in the development of subcellular ocular drug delivery strategies.

  18. 3-Nitropropionic acid induces autophagy by forming mitochondrial permeability transition pores rather than activatiing the mitochondrial fission pathway

    PubMed Central

    Solesio, Maria E; Saez-Atienzar, Sara; Jordan, Joaquin; Galindo, Maria F

    2013-01-01

    BACKGROUND AND PURPOSE Huntington's disease is a neurodegenerative process associated with mitochondrial alterations. Inhibitors of the electron–transport channel complex II, such as 3-nitropropionic acid (3NP), are used to study the molecular and cellular pathways involved in this disease. We studied the effect of 3NP on mitochondrial morphology and its involvement in macrophagy. EXPERIMENTAL APPROACH Pharmacological and biochemical methods were used to characterize the effects of 3NP on autophagy and mitochondrial morphology. SH-SY5Y cells were transfected with GFP-LC3, GFP-Drp1 or GFP-Bax to ascertain their role and intracellular localization after 3NP treatment using confocal microscopy. KEY RESULTS Untreated SH-SY5Y cells presented a long, tubular and filamentous net of mitochondria. After 3NP (5 mM) treatment, mitochondria became shorter and rounder. 3NP induced formation of mitochondrial permeability transition pores, both in cell cultures and in isolated liver mitochondria, and this process was inhibited by cyclosporin A. Participation of the mitochondrial fission pathway was excluded because 3NP did not induce translocation of the dynamin-related protein 1 (Drp1) to the mitochondria. The Drp1 inhibitor Mdivi-1 did not affect the observed changes in mitochondrial morphology. Finally, scavengers of reactive oxygen species failed to prevent mitochondrial alterations, while cyclosporin A, but not Mdivi-1, prevented the generation of ROS. CONCLUSIONS AND IMPLICATIONS There was a direct correlation between formation of mitochondrial permeability transition pores and autophagy induced by 3NP treatment. Activation of autophagy preceded the apoptotic process and was mediated, at least partly, by formation of reactive oxygen species and mitochondrial permeability transition pores. LINKED ARTICLE This article is commented on by González-Polo et al., pp. 60–62 of this issue. To view this commentary visit http://dx.doi.org/10.1111/j.1476-5381.2012.02203.x PMID

  19. Depletion of Cyclophilins B and C Leads to Dysregulation of Endoplasmic Reticulum Redox Homeostasis*

    PubMed Central

    Stocki, Pawel; Chapman, Daniel C.; Beach, Lori A.; Williams, David B.

    2014-01-01

    Protein folding within the endoplasmic reticulum is assisted by molecular chaperones and folding catalysts that include members of the protein-disulfide isomerase and peptidyl-prolyl isomerase families. In this report, we examined the contributions of the cyclophilin subset of peptidyl-prolyl isomerases to protein folding and identified cyclophilin C as an endoplasmic reticulum (ER) cyclophilin in addition to cyclophilin B. Using albumin and transferrin as models of cis-proline-containing proteins in human hepatoma cells, we found that combined knockdown of cyclophilins B and C delayed transferrin secretion but surprisingly resulted in more efficient oxidative folding and secretion of albumin. Examination of the oxidation status of ER protein-disulfide isomerase family members revealed a shift to a more oxidized state. This was accompanied by a >5-fold elevation in the ratio of oxidized to total glutathione. This “hyperoxidation” phenotype could be duplicated by incubating cells with the cyclophilin inhibitor cyclosporine A, a treatment that triggered efficient ER depletion of cyclophilins B and C by inducing their secretion to the medium. To identify the pathway responsible for ER hyperoxidation, we individually depleted several enzymes that are known or suspected to deliver oxidizing equivalents to the ER: Ero1αβ, VKOR, PRDX4, or QSOX1. Remarkably, none of these enzymes contributed to the elevated oxidized to total glutathione ratio induced by cyclosporine A treatment. These findings establish cyclophilin C as an ER cyclophilin, demonstrate the novel involvement of cyclophilins B and C in ER redox homeostasis, and suggest the existence of an additional ER oxidative pathway that is modulated by ER cyclophilins. PMID:24990953

  20. Angiotensin Type-2 (AT-2)-Receptor activation reduces renal fibrosis in cyclosporine nephropathy: Evidence for blood-pressure independent effect.

    PubMed

    Castoldi, Giovanna; di Gioia, Cira R T; Carletti, Raffaella; Roma, Francesca; Zerbini, Gianpaolo; Stella, Andrea

    2016-09-27

    Compound 21 (C21), selective agonist of AT2 receptors, shows antinflammatory effects in hypertension and nephroprotection in diabetes. The aim of this study was to evaluate the effects of C21 in cyclosporine nephropathy, which is characterized mainly by tubulo-interstitial fibrosis. Ten days before and during the experimental periods, low-salt diet was administered to Sprague Dawley rats. Cyclosporine-A (15mg/kg/day, i.p.) and cyclosporine-A plus C21 (0.3 mg/kg /day, i.p) were administered for 1 and 4 weeks. Control groups was left without any treatment. Blood pressure (plethysmographic method) and 24 hour albuminuria were measured once a week. At the end of the experiments, the kidneys were excised for histomorphometric analysis of renal fibrosis and for immunohistochemical evaluation of inflammatory infiltrates and type I and IV collagen expression.
    After 1 and 4 weeks, the rats treated with cyclosporine showed a significant increase (p <0.01) in blood pressure, no significant changes in albuminuria, a significant increase (p <0.01) in glomerular and tubulo-interstitial fibrosis and inflammatory infiltrates as compared to the control rats. Treatment with C21 did not modify the cyclosporine dependent increase of blood pressure, which was higher than in control rats, but after 4 weeks of treatment significantly reduced (p <0.01) glomerular and tubulo-interstitial fibrosis, type 1 collagen expression and macrophage infiltration, as compared to rats treated with cyclosporine.The administration of C21 showed a protective effect on cyclosporine nephropathy, decreasing renal fibrosis and macrophage infiltration. These data suggest that C21 may counteract tubulo-interstitial fibrosis, the most potent predictor of the progression of renal diseases.

  1. Calcium regulates caveolin-1 expression at the transcriptional level

    SciTech Connect

    Yang, Xiao-Yan; Huang, Cheng-Cheng; Kan, Qi-Ming; Li, Yan; Liu, Dan; Zhang, Xue-Cheng; Sato, Toshinori; Yamagata, Sadako; Yamagata, Tatsuya

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer Caveolin-1 expression is regulated by calcium signaling at the transcriptional level. Black-Right-Pointing-Pointer An inhibitor of or siRNA to L-type calcium channel suppressed caveolin-1 expression. Black-Right-Pointing-Pointer Cyclosporine A or an NFAT inhibitor markedly reduced caveolin-1 expression. Black-Right-Pointing-Pointer Caveolin-1 regulation by calcium signaling is observed in several mouse cell lines. -- Abstract: Caveolin-1, an indispensable component of caveolae serving as a transformation suppressor protein, is highly expressed in poorly metastatic mouse osteosarcoma FBJ-S1 cells while highly metastatic FBJ-LL cells express low levels of caveolin-1. Calcium concentration is higher in FBJ-S1 cells than in FBJ-LL cells; therefore, we investigated the possibility that calcium signaling positively regulates caveolin-1 in mouse FBJ-S1 cells. When cells were treated with the calcium channel blocker nifedipine, cyclosporin A (a calcineurin inhibitor), or INCA-6 (a nuclear factor of activated T-cells [NFAT] inhibitor), caveolin-1 expression at the mRNA and protein levels decreased. RNA silencing of voltage-dependent L-type calcium channel subunit alpha-1C resulted in suppression of caveolin-1 expression. This novel caveolin-1 regulation pathway was also identified in mouse NIH 3T3 cells and Lewis lung carcinoma cells. These results indicate that caveolin-1 is positively regulated at the transcriptional level through a novel calcium signaling pathway mediated by L-type calcium channel/Ca{sup 2+}/calcineurin/NFAT.

  2. Haem uptake is essential for egg production in the haematophagous blood fluke of humans, Schistosoma mansoni.

    PubMed

    Toh, Shu Qin; Gobert, Geoffrey N; Malagón Martínez, David; Jones, Malcolm K

    2015-09-01

    Schistosomes ingest host erythrocytes, liberating large quantities of haem. Despite its toxicity, haem is an essential factor for numerous biological reactions, and may be an important iron source for these helminths. We used a fluorescence haem analogue, palladium mesoporphyrin, to investigate pathways of haem acquisition, and showed that palladium mesoporphyrin accumulates in the vitellaria (eggshell precursor glands) and ovary of female Schistosoma mansoni. Furthermore, incubation of adult females in 10-100 μm cyclosporin A (IC50 = 2.3 μm) inhibits the uptake of palladium mesoporphyrin to these tissues, with tenfold reductions in fluorescence intensity of the ovary. In vitro exposure to cyclosporin A resulted in significant perturbation of egg production, reducing egg output from 34 eggs per female to 5.7 eggs per female over the incubation period, and retardation of egg development. We characterized a S. mansoni homologue of the haem-responsive genes of Caenorhabditis elegans. The gene (Smhrg-1) encodes a protein with a molecular weight of approximately 17 kDa. SmHRG-1 was able to rescue growth in haem transport-deficient HEM1Δ yeast. Transcriptional suppression of Smhrg-1 in adult S. mansoni worms resulted in significant delay in egg maturation, with 47% of eggs from transcriptionally suppressed worms being identified as immature compared with only 27% of eggs laid by control worms treated with firefly luciferase. Our findings indicate the presence of transmembrane haem transporters in schistosomes, with a high abundance of these molecules being present in tissues involved in oogenesis.

  3. Posaconazole plasma concentration in pediatric patients receiving antifungal prophylaxis after allogeneic hematopoietic stem cell transplantation.

    PubMed

    Heinz, Werner J; Cabanillas Stanchi, Karin M; Klinker, Hartwig; Blume, Olivia; Feucht, Judith; Hartmann, Ulrike; Feuchtinger, Tobias; Lang, Peter; Handgretinger, Rupert; Döring, Michaela

    2016-02-01

    Posaconazole has been proven to be effective for antifungal prophylaxis in adults after hematopoietic stem cell transplantation (HSCT). Due to low gastrointestinal resorption of posaconazole suspension, bioavailability is impaired. Fatty food improves the uptake of posaconazole, but insufficient data on the pharmacokinetics of posaconazole in pediatric patients are available so far. The single-center analysis investigated 161 posaconazole serum concentrations in 27 pediatric patients after HSCT receiving 12 mg·kg BW(-1)·d(-1) posaconazole suspension depending on age, gender, and intestinal graft-versus-host (iGvHD) disease, and the influence of posaconazole on cyclosporine A plasma concentrations. To improve the uptake of posaconazole, one patient cohort received higher fat nutrition with the drug administration. A comparison of the regular nutrition and higher-fat nutrition groups revealed the following values: 31 (27.4%) versus 8 (16.7%) < 500 ng/ml; 12 (10.6%) versus 7 (14.6%) 500-700 ng/ml; 8 (7.1%) versus 6 (12.5%) 700-1000 ng/ml; 51 (45.1%) versus 21 (43.8%) 1000-2000 ng/ml; and 11 (9.7%) versus 6 (12.5%) > 2000 ng/ml. The mean posaconazole concentrations in patients with regular nutrition was 1123 ± 811 ng/ml and with higher-fat nutrition was 1191 ± 673 ng/ml. Posaconazole levels in patients with iGvHD were significantly lower (P = 0.0003) than in patients without GvHD. The majority of samples showed a sufficient posaconazole concentration above 700 ng/ml. Posaconazole levels were slightly higher in patients with higher-fat nutrition and significantly lower in patients with iGvHD. Cyclosporine A levels were not significantly higher during posaconazole administration.

  4. Novel mode of action of the calcium antagonist mibefradil (Ro 40-5967): potent immunosuppression by inhibition of T-cell infiltration through allogeneic endothelium.

    PubMed Central

    Blaheta, R A; Hailer, N P; Brude, N; Wittig, B; Oppermann, E; Leckel, K; Harder, S; Scholz, M; Weber, S; Encke, A; Markus, B H

    1998-01-01

    Cyclosporin A reduces the mitotic activity of allosensitized lymphocytes, but fails to limit emigration of these cells into the donor organ. However, the modulation of both lymphocyte proliferation and infiltration are desirable characteristics of immunosuppressive therapy. The calcium-channel blocker, verapamil, has recently been shown to effectively prevent the transmigration of CD4+ and CD8+ T cells through allogeneic endothelium. Mibefradil (Ro 40-5967) represents a new generation of calcium antagonists with high potency and long-term activity. To evaluate the immunosuppressive potential of this drug, the influence of mibefradil on lymphocyte adhesion to, horizontal locomotion along, and penetration through allogeneic endothelium (HUVEC) was performed. When lymphocytes were prestimulated for 24 hr with mibefradil, adhesion and penetration were dose-dependently reduced. The adhesion ID50 values were 3.4 microM (CD4+ T cells) versus 9.2 microM (CD8+ T cells) and 2.1 microM (CD4+ T cells) versus 3.9 microM (CD8+ T cells) with regard to penetration. Mibefradil also effectively blocked horizontal locomotion. Specific down-regulation of T-cell binding to the P-selection receptor (ID50: CD4+ T cells, 0.8 microM: CD8+ T cells, 1.2 microM) and to the intracellular adhesion molecule-1 (ICAM-1) receptor (ID50: CD4+ T cells, 1.9 microM; CD8+ T cells, 1.5 microM) by mibefradil seems to be responsible for the decreased adhesion and penetration rates. Reduction of intracellular F-actin in T lymphocytes could diminish cell locomotion. In conclusion, the potent suppressive properties of mibefradil support its use as a co-medication in cyclosporin A-based immunosuppressive therapy. PMID:9741343

  5. Modulation of F0F1-ATP synthase activity by cyclophilin D regulates matrix adenine nucleotide levels

    PubMed Central

    Chinopoulos, Christos; Konràd, Csaba; Kiss, Gergely; Metelkin, Eugeniy; Töröcsik, Beata; Zhang, Steven F.; Starkov, Anatoly A.

    2011-01-01

    Cyclophilin D was recently shown to bind to and decrease the activity of F0F1-ATP synthase in submitochondrial particles and permeabilized mitochondria (Giorgio et al. 2009, J Biol Chem, 284:33982). Cyclophilin D binding decreased both the ATP synthesis and hydrolysis rates. Here, we reaffirm these findings by demonstrating that in intact mouse liver mitochondria energized by ATP, absence of cyclophilin D or presence of cyclosporin A led to a decrease in the extent of uncoupler-induced depolarization. Accordingly, in substrate-energized mitochondria an increase in F0F1-ATP synthase activity mediated by a relief of inhibition by cyclophilin D was evident as slightly increased respiration rates during arsenolysis. However, the modulation of F0F1-ATP synthase by cyclophilin D did not increase the ANT-mediated ATP efflux rate in energized mitochondria or the ATP influx rate in de-energized mitochondria. The lack of effect of cyclophilin D on the ANT-mediated adenine nucleotide exchange rate was attributed to the ~2.2 times lower flux control coefficient of the F0F1-ATP synthase than that of ANT, deduced from measurements of adenine nucleotide flux rates in intact mitochondria. These findings were further supported by a recent kinetic model of the mitochondrial phosphorylation system, suggesting that a ~30% change in F0F1-ATP synthase activity in fully energized or fully deenergized mitochondria affects ADP-ATP exchange rate mediated by the ANT in the range of 1.38-1.7%. We conclude that in mitochondria exhibiting intact inner membranes, the absence of cyclophilin D or inhibition of its binding to F0F1-ATP synthase by cyclosporin A will affect only matrix adenine nucleotides levels. PMID:21281446

  6. K restriction inhibits protein phosphatase 2B (PP2B) and suppression of PP2B decreases ROMK channel activity in the CCD

    PubMed Central

    Zhang, Yan; Lin, Dao-Hong; Wang, Zhi-Jian; Jin, Yan; Yang, Baofeng; Wang, Wen-Hui

    2009-01-01

    We used Western blot analysis to examine the effect of dietary K intake on the expression of serine/threonine protein phosphatase in the kidney. K restriction significantly decreased the expression of catalytic subunit of protein phosphatase (PP)2B but increased the expression of PP2B regulatory subunit in both rat and mouse kidney. However, K depletion did not affect the expression of PP1 and PP2A. Treatment of M-1 cells, mouse cortical collecting duct (CCD) cells, or 293T cells with glucose oxidase (GO), which generates superoxide anions through glucose metabolism, mimicked the effect of K restriction on PP2B expression and significantly decreased expression of PP2B catalytic subunits. However, GO treatment increased expression of regulatory subunit of PP2B and had no effect on expression of PP1, PP2A, and protein tyrosine phosphatase 1D. Moreover, deletion of gp91-containing NADPH oxidase abolished the effect of K depletion on PP2B. Thus superoxide anions or related products may mediate the inhibitory effect of K restriction on the expression of PP2B catalytic subunit. We also used patch-clamp technique to study the effect of inhibiting PP2B on renal outer medullary K (ROMK) channels in the CCD. Application of cyclosporin A or FK506, inhibitors of PP2B, significantly decreased ROMK channels, and the effect of PP2B inhibitors was abolished by blocking p38 mitogen-activated protein kinase (MAPK) and ERK. Furthermore, Western blot demonstrated that inhibition of PP2B with cyclosporin A or small interfering RNA increased the phosphorylation of ERK and p38 MAPK. We conclude that K restriction suppresses the expression of PP2B catalytic subunits and that inhibition of PP2B decreases ROMK channel activity through stimulation of MAPK in the CCD. PMID:18184875

  7. On the mechanism(s) of membrane permeability transition in liver mitochondria of lamprey, Lampetra fluviatilis L.: insights from cadmium.

    PubMed

    Belyaeva, Elena A; Emelyanova, Larisa V; Korotkov, Sergey M; Brailovskaya, Irina V; Savina, Margarita V

    2014-01-01

    Previously we have shown that opening of the mitochondrial permeability transition pore in its low conductance state is the case in hepatocytes of the Baltic lamprey (Lampetra fluviatilis L.) during reversible metabolic depression taking place in the period of its prespawning migration when the exogenous feeding is switched off. The depression is observed in the last year of the lamprey life cycle and is conditioned by reversible mitochondrial dysfunction (mitochondrial uncoupling in winter and coupling in spring). To further elucidate the mechanism(s) of induction of the mitochondrial permeability transition pore in the lamprey liver, we used Cd(2+) and Ca(2+) plus Pi as the pore inducers. We found that Ca(2+) plus Pi induced the high-amplitude swelling of the isolated "winter" mitochondria both in isotonic sucrose and ammonium nitrate medium while both low and high Cd(2+) did not produce the mitochondrial swelling in these media. Low Cd(2+) enhanced the inhibition of basal respiration rate of the "winter" mitochondria energized by NAD-dependent substrates whereas the same concentrations of the heavy metal evoked its partial stimulation on FAD-dependent substrates. The above changes produced by Cd(2+) or Ca(2+) plus Pi in the "winter" mitochondria were only weakly (if so) sensitive to cyclosporine A (a potent pharmacological desensitizer of the nonselective pore) added alone and they were not sensitive to dithiothreitol (a dithiol reducing agent). Under monitoring of the transmembrane potential of the "spring" lamprey liver mitochondria, we revealed that Cd(2+) produced its decrease on both types of the respiratory substrates used that was strongly hampered by cyclosporine A, and the membrane potential was partially restored by dithiothreitol. The effects of different membrane permeability modulators on the lamprey liver mitochondria function and the seasonal changes in their action are discussed.

  8. Antineoplastic copper coordinated complexes (Casiopeinas) uncouple oxidative phosphorylation and induce mitochondrial permeability transition in cardiac mitochondria and cardiomyocytes.

    PubMed

    Silva-Platas, Christian; Guerrero-Beltrán, Carlos Enrique; Carrancá, Mariana; Castillo, Elena Cristina; Bernal-Ramírez, Judith; Oropeza-Almazán, Yuriana; González, Lorena N; Rojo, Rocío; Martínez, Luis Enrique; Valiente-Banuet, Juan; Ruiz-Azuara, Lena; Bravo-Gómez, María Elena; García, Noemí; Carvajal, Karla; García-Rivas, Gerardo

    2016-02-01

    Copper-based drugs, Casiopeinas (Cas), exhibit antiproliferative and antineoplastic activities in vitro and in vivo, respectively. Unfortunately, the clinical use of these novel chemotherapeutics could be limited by the development of dose-dependent cardiotoxicity. In addition, the molecular mechanisms underlying Cas cardiotoxicity and anticancer activity are not completely understood. Here, we explore the potential impact of Cas on the cardiac mitochondria energetics as the molecular mechanisms underlying Cas-induced cardiotoxicity. To explore the properties on mitochondrial metabolism, we determined Cas effects on respiration, membrane potential, membrane permeability, and redox state in isolated cardiac mitochondria. The effect of Cas on the mitochondrial membrane potential (Δψm) was also evaluated in isolated cardiomyocytes by confocal microscopy and flow cytometry. Cas IIIEa, IIgly, and IIIia predominately inhibited maximal NADH- and succinate-linked mitochondrial respiration, increased the state-4 respiration rate and reduced membrane potential, suggesting that Cas also act as mitochondrial uncouplers. Interestingly, cyclosporine A inhibited Cas-induced mitochondrial depolarization, suggesting the involvement of mitochondrial permeability transition pore (mPTP). Similarly to isolated mitochondria, in isolated cardiomyocytes, Cas treatment decreased the Δψm and cyclosporine A treatment prevented mitochondrial depolarization. The production of H2O2 increased in Cas-treated mitochondria, which might also increase the oxidation of mitochondrial proteins such as adenine nucleotide translocase. In accordance, an antioxidant scavenger (Tiron) significantly diminished Cas IIIia mitochondrial depolarization. Cas induces a prominent loss of membrane potential, associated with alterations in redox state, which increases mPTP opening, potentially due to thiol-dependent modifications of the pore, suggesting that direct or indirect inhibition of mPTP opening might

  9. Quercetin-3-O-(2″-galloyl)-α-l-rhamnopyranoside inhibits TNF-α-activated NF-κB-induced inflammatory mediator production by suppressing ERK activation.

    PubMed

    Lee, Chung Soo; Jeong, Eun Byul; Kim, Yun Jeong; Lee, Min Sung; Seo, Seong Jun; Park, Kwan Hee; Lee, Min Won

    2013-08-01

    Quercetin and its derivatives have anti-inflammatory and anti-oxidant effects. However, the effect of quercetin-3-O-(2″-galloyl)-α-l-rhamnopyranoside (QGR), a new quercetin derivative, on the tumor necrosis factor (TNF)-α-stimulated production of inflammatory mediators in keratinocytes is unclear. In addition, the effect of QGR on the ERK and NF-κB-mediated inflammatory process has not been studied. In human keratinocyte HaCat cells, we investigated the effect of QGR on the TNF-α-stimulated production of inflammatory mediators in relation to the nuclear factor (NF)-κB, which regulates the transcription genes involved in immune and inflammatory responses. QGR inhibited the TNF-α-stimulated production of cytokines and chemokines in HaCaT cells. QGR, dexamethasone, cyclosporine A, Bay 11-7085 (an inhibitor of NF-κB activation) and cell signaling ERK inhibitor attenuated the TNF-α-induced formation of inflammatory mediators and activation of the NF-κB and ERK. Unlike other compounds, dexamethasone and cyclosporine A did not reduce formation of reactive oxygen species. The results show that QGR may attenuate TNF-α-stimulated inflammatory mediator production in HaCaT cells by suppressing the activation of the ERK-mediated NF-κB pathway that is mediated by reactive oxygen species. Additionally, QGR may exhibit a preventive effect against the proinflammatory mediator-induced skin diseases by inhibiting the activation of the ERK and NF-κB pathways.

  10. Intermedilysin induces EGR-1 expression through calcineurin/NFAT pathway in human cholangiocellular carcinoma cells

    SciTech Connect

    Susilowati, Heni; Okamura, Hirohiko; Hirota, Katsuhiko; Shono, Masayuki; Yoshida, Kaya; Murakami, Keiji; Tabata, Atsushi; Nagamune, Hideaki; Haneji, Tatsuji; Miyake, Yoichiro

    2011-01-07

    Research highlights: {yields} ILY leads to the accumulation of [Ca{sup 2+}]i in the nucleus in HuCCT1 cells. {yields} ILY induced activation of NFAT1 through a calcineurin-dependent pathway. {yields} Calcineuri/NFAT pathway is involved in EGR-1 expression in response to ILY treatment. -- Abstract: Intermedilysin (ILY) is a cholesterol-dependent cytolysin produced by Streptococcus intermedius, which is associated with human brain and liver abscesses. Although intrahepatic bile duct cells play a valuable role in the pathogenesis of liver abscess, the molecular mechanism of ILY-treated intrahepatic bile duct cells remains unknown. In this study, we report that ILY induced a nuclear accumulation of intracellular calcium ([Ca{sup 2+}]i) in human cholangiocellular cells HuCCT1. We also demonstrate that 10 ng/ml ILY induced NFAT1 dephosphorylation and its nuclear translocation in HuCCT1 cells. In contrast to the result that ILY induced NF-{kappa}B translocation in human hepatic HepG2 cells, ILY did not affect NF-{kappa}B localization in HuCCT1 cells. Dephosphorylation and nuclear translocation of NFAT1 caused by ILY were prevented by [Ca{sup 2+}]i calcium chelator, BAPTA/AM, and calcineurin inhibitors, cyclosporine A and tacrolimus. ILY induced early growth response-1 (EGR-1) expression and it was inhibited by the pre-treatment with cyclosporine A, indicating that the calcineurin/NFAT pathway was involved in EGR-1 expression in response to ILY. ILY-induced calcineurin/NFAT1 activation and sequential EGR-1 expression might be related to the pathogenesis of S. intermedius in human bile duct cells.

  11. Seasonal influence on mitogen and cyclosporin responses of peripheral blood lymphocytes.

    PubMed

    Michelis, Fotios V; Delitheos, Andreas K; Tiligada, Ekaterini

    2013-06-01

    The immune response and lymphocyte activation in particular are affected by environmental factors. In vivo and in vitro experiments demonstrate variability in lymphocyte activation according to seasonal changes. This study focused on the effects of season on the ex vivo mitogen-induced activation of lymphocytes from peripheral blood of healthy humans living in a temperate climate, as well as the ex vivo lymphocyte activation of rabbits living under constant laboratory conditions. The possible impact of season on the action of the immunosuppressant drug cyclosporin A (CsA) on lymphocyte activation was investigated in both species. Cultured peripheral blood lymphocytes from human donors (n=13, 22-63years of age) and from animals housed under 12:12hour light:dark cycle were stimulated with phytohemagglutinin (PHA) in the absence or presence of 10 and 25μg/mL CsA. Lymphocyte activation was assessed by morphometric analysis under a light microscope. Percentages of unactivated lymphocytes, activated lymphoblasts and aberrant cells reflecting cytotoxicity were determined. Human lymphocytes demonstrated a significant decrease in response to PHA during the winter months, in comparison to the rest of the year. In contrast, the peripheral blood lymphocytes of rabbits housed under constant conditions did not demonstrate similar variations in response to PHA stimulation. The immunosuppressive action of cyclosporin A on this experimental model was unaffected by the observed seasonal variation in mitogen response in humans. These findings may guide research towards the identification of factors associated with the seasonality of the immune response and its potential influence on therapeutic interventions.

  12. BK and JC polyomavirus infections in Tunisian renal transplant recipients.

    PubMed

    Boukoum, Hanen; Nahdi, Imen; Sahtout, Wissal; Skiri, Habib; Aloui, Sabra; Achour, Abdelatif; Segondy, Michel; Aouni, Mahjoub

    2015-10-01

    The aim of this prospective study was to investigate the rate of BK (BKPyV) and JC (JCPyV) polyomavirus infections and their influence on allograft function in Tunisian renal transplant recipients. A total of 72 renal transplant recipients were studied. BKPyV and JCPyV were detected and quantified by real-time PCR in urine and plasma. Demographic and laboratory characteristics were collected for each patient. Polyomavirus DNAuria was detected in 54 (75%) of renal transplant recipients: 26 (36%) had BKPyV DNAuria, 20 (28%) had JCPyV DNAuria, and 8 (11%) had a dual BKPyV/JCPyV DNAuria. BKPyV DNAemia was detected in four (5.5%) patients, whereas no patient had JCPyV viremia. More than 70% of BKPyV and JCPyV infections started within the first 3 months post-transplant. The risk for positive DNAemia was observed in patients with DNAuria level >10(7) copies/ml. BK Polyomavirus-associated nephropathy (BKPyVAN) was observed in two patients. This study highlights the high frequency of BKPyV and JCPyV viruria during the first year post-transplant with the highest incidence observed in the third month. We identified several risk factors that were associated with BKV DNAuria including age, sex of patients, and the use of tacrolimus instead of cyclosporine A at month 3. The use of cyclosporine A instead of tacrolimus was identified as risk factor for JCV viruria in month 3. No statistical difference in the allograft function was found between BKPyV and/or JCPyV infected and uninfected patients.

  13. Herpesviral G protein-coupled receptors activate NFAT to induce tumor formation via inhibiting the SERCA calcium ATPase.

    PubMed

    Zhang, Junjie; He, Shanping; Wang, Yi; Brulois, Kevin; Lan, Ke; Jung, Jae U; Feng, Pinghui

    2015-03-01

    G protein-coupled receptors (GPCRs) constitute the largest family of proteins that transmit signal to regulate an array of fundamental biological processes. Viruses deploy diverse tactics to hijack and harness intracellular signaling events induced by GPCR. Herpesviruses encode multiple GPCR homologues that are implicated in viral pathogenesis. Cellular GPCRs are primarily regulated by their cognate ligands, while herpesviral GPCRs constitutively activate downstream signaling cascades, including the nuclear factor of activated T cells (NFAT) pathway. However, the roles of NFAT activation and mechanism thereof in viral GPCR tumorigenesis remain unknown. Here we report that GPCRs of human Kaposi's sarcoma-associated herpesvirus (kGPCR) and cytomegalovirus (US28) shortcut NFAT activation by inhibiting the sarcoplasmic reticulum calcium ATPase (SERCA), which is necessary for viral GPCR tumorigenesis. Biochemical approaches, entailing pharmacological inhibitors and protein purification, demonstrate that viral GPCRs target SERCA2 to increase cytosolic calcium concentration. As such, NFAT activation induced by vGPCRs was exceedingly sensitive to cyclosporine A that targets calcineurin, but resistant to inhibition upstream of ER calcium release. Gene expression profiling identified a signature of NFAT activation in endothelial cells expressing viral GPCRs. The expression of NFAT-dependent genes was up-regulated in tumors derived from tva-kGPCR mouse and human KS. Employing recombinant kGPCR-deficient KSHV, we showed that kGPCR was critical for NFAT-dependent gene expression in KSHV lytic replication. Finally, cyclosporine A treatment diminished NFAT-dependent gene expression and tumor formation induced by viral GPCRs. These findings reveal essential roles of NFAT activation in viral GPCR tumorigenesis and a mechanism of "constitutive" NFAT activation by viral GPCRs.

  14. c-Jun N-terminal kinase (JNK)-mediated modulation of brain mitochondria function: new target proteins for JNK signalling in mitochondrion-dependent apoptosis.

    PubMed Central

    Schroeter, Hagen; Boyd, Clinton S; Ahmed, Ruhi; Spencer, Jeremy P E; Duncan, Roger F; Rice-Evans, Catherine; Cadenas, Enrique

    2003-01-01

    The molecular mechanisms underlying the initiation and control of the release of cytochrome c during mitochondrion-dependent apoptosis are thought to involve the phosphorylation of mitochondrial Bcl-2 and Bcl-x(L). Although the c-Jun N-terminal kinase (JNK) has been proposed to mediate the phosphorylation of Bcl-2/Bcl-x(L) the mechanisms linking the modification of these proteins and the release of cytochrome c remain to be elucidated. This study was aimed at establishing interdependency between JNK signalling and mitochondrial apoptosis. Using an experimental model consisting of isolated, bioenergetically competent rat brain mitochondria, these studies show that (i) JNK catalysed the phosphorylation of Bcl-2 and Bcl-x(L) as well as other mitochondrial proteins, as shown by two-dimensional isoelectric focusing/SDS/PAGE; (ii) JNK-induced cytochrome c release, in a process independent of the permeability transition of the inner mitochondrial membrane (imPT) and insensitive to cyclosporin A; (iii) JNK mediated a partial collapse of the mitochondrial inner-membrane potential (Deltapsim) in an imPT- and cyclosporin A-independent manner; and (iv) JNK was unable to induce imPT/swelling and did not act as a co-inducer, but as an inhibitor of Ca-induced imPT. The results are discussed with regard to the functional link between the Deltapsim and factors influencing the permeability transition of the inner and outer mitochondrial membranes. Taken together, JNK-dependent phosphorylation of mitochondrial proteins including, but not limited to, Bcl-2/Bcl-x(L) may represent a potential of the modulation of mitochondrial function during apoptosis. PMID:12614194

  15. Look Different: Effect of Radiation Hormesis on the Survival Rate of Immunosuppressed Mice

    PubMed Central

    Alavi, M.; Taeb, S.; Okhovat, M.A.; Atefi, M.; Negahdari, F.

    2016-01-01

    Background: Hormesis is defined as the bio-positive response of something which is bio-negative in high doses. In the present study, the effect of radiation hormesis was evaluated on the survival rate of immunosuppressed BALB/c mice by Cyclosporine A. Material and Methods: We used 75 consanguine, male, BALB/c mice in this experiment. The first group received Technetium-99m and the second group was placed on a sample radioactive soil of Ramsar region (800Bq) for 20 days. The third group was exposed to X-rays and the fourth group was placed on the radioactive soil and then injected Technetium-99m. The last group was the sham irradiated control group. Finally, 30mg Cyclosporine A as the immunosuppressive agent was orally administered to all mice 48 hours after receiving X-rays and Technetium-99m. The mean survival rate of mice in each group was estimated during time. Results: A log rank test was run to determine if there were differences in the survival distribution for different groups and related treatments. According to the results, the survival rate of all pre-irradiated groups was more than the sham irradiated control group (p < .05). The highest survival time was related to the mice which were placed on the radioactive soil of Ramsar region for 20 days and then injected Technetium-99m. Conclusion: This study confirmed the presence of hormetic models and the enhancement of survival rate in immunosuppressed BALB/c mice as a consequence of low-dose irradiation. It is also revealed the positive synergetic radioadaptive response on survival rate of immunosuppressed animals. PMID:27853721

  16. Micromolar changes in lysophosphatidylcholine concentration cause minor effects on mitochondrial permeability but major alterations in function.

    PubMed

    Hollie, Norris I; Cash, James G; Matlib, M Abdul; Wortman, Matthew; Basford, Joshua E; Abplanalp, William; Hui, David Y

    2014-06-01

    Mice deficient in group 1b phospholipase A2 have decreased plasma lysophosphatidylcholine and increased hepatic oxidation that is inhibited by intraperitoneal lysophosphatidylcholine injection. This study sought to identify a mechanism for lysophosphatidylcholine-mediated inhibition of hepatic oxidative function. Results showed that in vitro incubation of isolated mitochondria with 40-200μM lysophosphatidylcholine caused cyclosporine A-resistant swelling in a concentration-dependent manner. However, when mitochondria were challenged with 220μM CaCl2, cyclosporine A protected against permeability transition induced by 40μM, but not 80μM lysophosphatidylcholine. Incubation with 40-120μM lysophosphatidylcholine also increased mitochondrial permeability to 75μM CaCl2 in a concentration-dependent manner. Interestingly, despite incubation with 80μM lysophosphatidylcholine, the mitochondrial membrane potential was steady in the presence of succinate, and oxidation rates and respiratory control indices were similar to controls in the presence of succinate, glutamate/malate, and palmitoyl-carnitine. However, mitochondrial oxidation rates were inhibited by 30-50% at 100μM lysophosphatidylcholine. Finally, while 40μM lysophosphatidylcholine has no effect on fatty acid oxidation and mitochondria remained impermeable in intact hepatocytes, 100μM lysophosphatidylcholine inhibited fatty acid stimulated oxidation and caused intracellular mitochondrial permeability. Taken together, these present data demonstrated that LPC concentration dependently modulates mitochondrial microenvironment, with low micromolar concentrations of lysophosphatidylcholine sufficient to change hepatic oxidation rate whereas higher concentrations are required to disrupt mitochondrial integrity.

  17. Acetaminophen hepatotoxicity and HIF-1α induction in acetaminophen toxicity in mice occurs without hypoxia.

    PubMed

    Chaudhuri, Shubhra; McCullough, Sandra S; Hennings, Leah; Letzig, Lynda; Simpson, Pippa M; Hinson, Jack A; James, Laura P

    2011-05-01

    HIF-1α is a nuclear factor important in the transcription of genes controlling angiogenesis including vascular endothelial growth factor (VEGF). Both hypoxia and oxidative stress are known mechanisms for the induction of HIF-1α. Oxidative stress and mitochondrial permeability transition (MPT) are mechanistically important in acetaminophen (APAP) toxicity in the mouse. MPT may occur as a result of oxidative stress and leads to a large increase in oxidative stress. We previously reported the induction of HIF-1α in mice with APAP toxicity and have shown that VEGF is important in hepatocyte regeneration following APAP toxicity. The following study was performed to examine the relative contribution of hypoxia versus oxidative stress to the induction of HIF-1α in APAP toxicity in the mouse. Time course studies using the hypoxia marker pimonidazole showed no staining for pimonidazole at 1 or 2h in B6C3F1 mice treated with APAP. Staining for pimonidazole was present in the midzonal to periportal regions at 4, 8, 24 and 48h and no staining was observed in centrilobular hepatocytes, the sites of the toxicity. Subsequent studies with the MPT inhibitor cyclosporine A showed that cyclosporine A (CYC; 10mg/kg) reduced HIF-1α induction in APAP treated mice at 1 and 4h and did not inhibit the metabolism of APAP (depletion of hepatic non-protein sulfhydryls and hepatic protein adduct levels). The data suggest that HIF-1α induction in the early stages of APAP toxicity is secondary to oxidative stress via a mechanism involving MPT. In addition, APAP toxicity is not mediated by a hypoxia mechanism.

  18. Calcineurin-mediated Bad translocation regulates cyanide-induced neuronal apoptosis.

    PubMed Central

    Shou, Yan; Li, Li; Prabhakaran, Krishnan; Borowitz, Joseph L; Isom, Gary E

    2004-01-01

    In cyanide-induced apoptosis, an increase in cytosolic free Ca2+ and generation of reactive oxygen species are initiation stimuli for apoptotic cell death. Previous studies have shown that cyanide-stimulated translocation of Bax (Bcl-associated X protein) to mitochondria is linked with release of cytochrome c and subsequent activation of a caspase cascade [Shou, Li, Prabhakaran, Borowitz and Isom (2003) Toxicol. Sci. 75, 99-107]. In the present study, the relationship of the cyanide-induced increase in cytosolic free Ca2+ to activation of Bad ( Bcl-2/Bcl-X(L)- antagonist, causing cell death) was determined in cortical cells. Bad is a Ca2+-sensitive pro-apoptotic Bcl-2 protein, which on activation translocates from cytosol to mitochondria to initiate cytochrome c release. In cultured primary cortical cells, cyanide produced a concentration- and time-dependent translocation of Bad from cytosol to mitochondria. Translocation occurred early in the apoptotic response, since mitochondrial Bad was detected within 1 h of cyanide treatment. Mitochondrial levels of the protein continued to increase up to 12 h post-cyanide exposure. Concurrent with Bad translocation, a Ca2+-sensitive increase in cellular calcineurin activity was observed. Increased cytosolic Ca2+ and calcineurin activation stimulated Bad translocation since BAPTA [bis-(o-aminophenoxy)ethane-N, N, N', N'-tetra-acetic acid], an intracellular Ca2+ chelator, and cyclosporin A, a calcineurin inhibitor, significantly reduced translocation. BAPTA also blocked release of cytochrome c from mitochondria as well as apoptosis. Furthermore, treatment of cells with the calcineurin inhibitors cyclosporin A or FK506 blocked the apoptotic response, linking calcineurin activation and the subsequent translocation of Bad to cell death. These observations show that by inducing a rapid increase in cytosolic free Ca2+, cyanide can partially initiate the apoptotic cascade through a calcineurin-mediated translocation of Bad to

  19. Fluorescence assay for mitochondrial permeability transition in cardiomyocytes cultured in a microtiter plate.

    PubMed

    Christensen, Marie Louise Muff; Braunstein, Thomas Hartig; Treiman, Marek

    2008-07-01

    Mitochondrial permeability transition pore (MPTP) is a voltage-dependent, large-conductance channel of the inner mitochondrial membrane with an important role in a range of pathophysiological conditions. To facilitate studies of pharmacological pore modulation, we describe an assay in a model using neonatal cardiomyocytes in a 96-well microtiter plate format. In the presence of mitochondrial membrane potential Delta Psi m, accumulation of rhodamine-123 in mitochondria (40,000 cells/well, 2.6 microM rhodamine-123) caused fluorescence signal quenching. Following substitution of dye-free buffer, dequenching occurred on the distribution of rhodamine-123 into the extracellular volume. The addition of a small buffer volume containing digitonin (final concentration 10 microg/ml) and Ca(2+) (final concentrations up to 100 microM free Ca(2+)) caused dequenching (Delta F) due to Delta Psi m dissipation by MPTP, as evidenced by inhibition in the presence of cyclosporin A (0.2-2 microM) and facilitation by pH 6.2. Delta F due to Delta Psi m-dissipating agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) or alamethicin (10 microM) was insensitive to either pH or cyclosporin A. Inhibition of Ca(2+)-induced (but not of FCCP- or alamethicin-induced) Delta F by glycogen synthase kinase 3beta (GSK3 beta) antagonist SB216763 and adenosine, acting at the level of intracellular signaling and plasma membrane receptors, respectively, is shown to illustrate potential applications of this assay. Limitation of the assay to cells with energized mitochondria is stressed.

  20. Ca2+ acting at the external side of the inner mitochondrial membrane can stimulate mitochondrial permeability transition induced by phenylarsine oxide.

    PubMed

    Kowaltowski, A J; Castilho, R F

    1997-12-15

    Mitochondrial permeability transition (MPT) induced by the thiol cross-linker phenylarsine oxide (PhAsO) in Ca(2+)-depleted mitochondria incubated in the presence of ruthenium red, an inhibitor of the Ca2+ uniporter, is stimulated by the addition of extramitochondrial Ca2+. The presence of extramitochondrial Ca2+ stimulates the reaction of mitochondrial membrane protein thiol groups with PhAsO. Both Ca(2+)-induced increase in mitochondrial membrane permeabilization and protein thiol group reaction with PhAsO are dependent on time (5-10 min to be complete) and the concentration of Ca2+ (1-25 microM). Mitochondrial permeabilization induced by PhAsO (15 microM) and extramitochondrial Ca2+ is inhibited by ADP, cyclosporin A, dibucaine and Mg2+, while mitochondrial permeabilization induced by high concentrations of PhAsO (60 microM) in the absence of Ca2+ is inhibited only by ADP and cyclosporin A. These results suggest that dibucaine and Mg2+ can inhibit mitochondrial permeabilization by antagonizing the effect of Ca2+ on the mitochondrial membrane. Once mitochondrial permeabilization induced by 15 microM PhAsO and extramitochondrial Ca2+ has already occurred, the addition of the Ca2+ chelator EGTA restores mitochondrial membrane potential (MPT pore closure), suggesting that the presence of Ca2+ is essential for the maintenance of the permeability of the mitochondrial membrane to protons (MPT pore opening). In conclusion, the results presented indicate that low Ca2+ concentrations acting at the external side of the inner mitochondrial membrane can stimulate mitochondrial permeability transition induced by PhAsO, due to increased accessibility of protein thiol groups to the reaction with PhAsO and increased probability of MPT pore opening.

  1. Not all mitochondrial carrier proteins support permeability transition pore formation: no involvement of uncoupling protein 1.

    PubMed

    Crichton, Paul G; Parker, Nadeene; Vidal-Puig, Antonio J; Brand, Martin D

    2009-12-15

    The mPTP (mitochondrial permeability transition pore) is a non-specific channel that is formed in the mitochondrial inner membrane in response to several stimuli, including elevated levels of matrix calcium. The pore is proposed to be composed of the ANT (adenine nucleotide translocase), voltage-dependent anion channel and cyclophilin D. Knockout studies, however, have demonstrated that ANT is not essential for permeability transition, which has led to the proposal that other members of the mitochondrial carrier protein family may be able to play a similar function to ANT in pore formation. To investigate this possibility, we have studied the permeability transition properties of BAT (brown adipose tissue) mitochondria in which levels of the mitochondrial carrier protein, UCP1 (uncoupling protein 1), can exceed those of ANT. Using an improved spectroscopic assay, we have quantified mPTP formation in de-energized mitochondria from wild-type and Ucp1KO (Ucp1-knockout) mice and assessed the dependence of pore formation on UCP1. When correctly normalized for differences in mitochondrial morphology, we find that calcium-induced mPTP activity is the same in both types of mitochondria, with similar sensitivity to GDP (approximately 50% inhibited), although the portion sensitive to cyclosporin A is higher in mitochondria lacking UCP1 (approximately 80% inhibited, compared with approximately 60% in mitochondria containing UCP1). We conclude that UCP1 is not a component of the cyclosporin A-sensitive mPTP in BAT and that playing a role in mPTP formation is not a general characteristic of the mitochondrial carrier protein family but is, more likely, restricted to specific members including ANT.

  2. Cobalt induces oxidative stress in isolated liver mitochondria responsible for permeability transition and intrinsic apoptosis in hepatocyte primary cultures.

    PubMed

    Battaglia, Valentina; Compagnone, Alessandra; Bandino, Andrea; Bragadin, Marcantonio; Rossi, Carlo Alberto; Zanetti, Filippo; Colombatto, Sebastiano; Grillo, Maria Angelica; Toninello, Antonio

    2009-03-01

    It is well established that cobalt mediates the occurrence of oxidative stress which contributes to cell toxicity and death. However, the mechanisms of these effects are not fully understood. This investigation aimed at establishing if cobalt acts as an inducer of mitochondrial-mediated apoptosis and at clarifying the mechanism of this process. Cobalt, in the ionized species Co(2+), is able to induce the phenomenon of mitochondrial permeability transition (MPT) in rat liver mitochondria (RLM) with the opening of the transition pore. In fact, Co(2+) induces mitochondrial swelling, which is prevented by cyclosporin A and other typical MPT inhibitors such as Ca(2+) transport inhibitors and bongkrekic acid, as well as anti-oxidant agents. In parallel with mitochondrial swelling, Co(2+) also induces the collapse of electrical membrane potential. However in this case, cyclosporine A and the other MPT inhibitors (except ruthenium red and EGTA) only partially prevent DeltaPsi drop, suggesting that Co(2+) also has a proton leakage effect on the inner mitochondrial membrane. MPT induction is due to oxidative stress, as a result of generation by Co(2+) of the highly damaging hydroxyl radical, with the oxidation of sulfhydryl groups, glutathione and pyridine nucleotides. Co(2+) also induces the release of the pro-apoptotic factors, cytochrome c and AIF. Incubation of rat hepatocyte primary cultures with Co(2+) results in apoptosis induction with caspase activation and increased level of expression of HIF-1alpha. All these observations allow us to state that, in the presence of calcium, Co(2+) is an inducer of apoptosis triggered by mitochondrial oxidative stress.

  3. [Therapy of terminal heart failure using heart transplantation].

    PubMed

    Hummel, M; Warnecke, H; Schüler, S; Hempel, B; Spiegelsberger, S; Hetzer, R

    1991-08-16

    Heart transplantation (HTx) has now become an accepted treatment modality for end-stage heart disease. The limited supply of suitable donor organs imposes constraints upon the decision of who should be selected for transplantation. Usually patients are candidates for HTx, who remain NYHA functional class III or IV despite maximal medical therapy. Further criteria are low left ventricular ejection fraction (less than 20%) with heart rhythm disturbances class IIIA-V (LOWN), which are associated with poor prognosis. Additionally, the suffering of the patient and also the course of heart failure are essential for judging the urgency of HTx. Contraindications are absolute in patients with untreated infections, fixed pulmonary vascular resistance (PVR) above 8 WOOD-degrees, severe irreversible kidney and liver disease, active ventricular or duodenal ulcers and acute, psychiatric illness. HTx is relatively contraindicated in patients with diabetes mellitus, age over 60 years, PVR above 6 WOOD-degrees and an unstable psychosocial situation. To prevent rejection of the transplant heart, live-long immunosuppressive therapy is needed. Most immunosuppressive regimes consist of Cyclosporine A and Azathioprine (double drug therapy) or in combination (tripple drug therapy) with Prednisolone. For monitoring of this therapy, control of hole blood cyclosporine A level and white blood count is needed. Rejection episodes can be suspected if there is a greater than 20 mmHg decrease of systolic blood pressure, elevated body temperature, malaise, tachycardia or heart rhythm disturbance. The diagnosis of cardiac rejection can be established by endomyocardial biopsy. Measurement of the voltage of either the surface or intramyocardial ECG, echocardiography with special consideration to early left ventricular filling time as well as immunological methods are additionally used tools. Graft sclerosis as the main risk factor of the late transplant period remains an unsolved problem.

  4. Morphine-Induced Preconditioning: Involvement of Protein Kinase A and Mitochondrial Permeability Transition Pore

    PubMed Central

    Dorsch, Marianne; Behmenburg, Friederike; Raible, Miriam; Blase, Dominic; Grievink, Hilbert; Hollmann, Markus W.; Heinen, André; Huhn, Ragnar

    2016-01-01

    Background Morphine induces myocardial preconditioning (M-PC) via activation of mitochondrial large conductance Ca2+-sensitive potassium (mKCa) channels. An upstream regulator of mKCa channels is protein kinase A (PKA). Furthermore, mKCa channel activation regulates mitochondrial bioenergetics and thereby prevents opening of the mitochondrial permeability transition pore (mPTP). Here, we investigated in the rat heart in vivo whether 1) M-PC is mediated by activation of PKA, and 2) pharmacological opening of the mPTP abolishes the cardioprotective effect of M-PC and 3) M-PC is critically dependent on STAT3 activation, which is located upstream of mPTP within the signalling pathway. Methods Male Wistar rats were randomised to six groups (each n = 6). All animals underwent 25 minutes of regional myocardial ischemia and 120 minutes of reperfusion. Control animals (Con) were not further treated. Morphine preconditioning was initiated by intravenous administration of 0.3 mg/kg morphine (M-PC). The PKA blocker H-89 (10 μg/kg) was investigated with and without morphine (H-89+M-PC, H-89). We determined the effect of mPTP opening with atractyloside (5 mg/kg) with and without morphine (Atr+M-PC, Atr). Furthermore, the effect of morphine on PKA activity was tested in isolated adult rat cardiomyocytes. In further experiments in isolated hearts we tested the protective properties of morphine in the presence of STAT3 inhibition, and whether pharmacological prevention of the mPTP-opening by cyclosporine A (CsA) is cardioprotective in the presence of STAT3 inhibition. Results Morphine reduced infarct size from 64±5% to 39±9% (P<0.05 vs. Con). H-89 completely blocked preconditioning by morphine (64±9%; P<0.05 vs. M-PC), but H-89 itself had not effect on infarct size (61±10%; P>0.05 vs. Con). Also, atractyloside abolished infarct size reduction of morphine completely (65±9%; P<0.05 vs. M-PC) but had no influence on infarct size itself (64±5%; P>0.05 vs. Con). In isolated

  5. Drug binding in human P-glycoprotein causes conformational changes in both nucleotide-binding domains.

    PubMed

    Loo, Tip W; Bartlett, M Claire; Clarke, David M

    2003-01-17

    The human multidrug resistance P-glycoprotein (P-gp, ABCB1) uses ATP to transport many structurally diverse compounds out of the cell. It is an ABC transporter with two nucleotide-binding domains (NBDs) and two transmembrane domains (TMDs). Recently, we showed that the "LSGGQ" motif in one NBD ((531)LSGGQ(535) in NBD1; (1176)LSGGQ(1180) in NBD2) is adjacent to the "Walker A" sequence ((1070)GSSGCGKS(1077) in NBD2; (427)GNSGCGKS(434) in NBD1) in the other NBD (Loo, T. W., Bartlett, M. C., and Clarke, D. M. (2002) J. Biol. Chem. 277, 41303-41306). Drug substrates can stimulate or inhibit the ATPase activity of P-gp. Here, we report the effect of drug binding on cross-linking between the LSGGQ signature and Walker A sites (Cys(431)(NBD1)/C1176C(NBD2) and Cys(1074)(NBD2)/L531C(NBD1), respectively). Seven drug substrates (calcein-AM, demecolcine, cis(Z)-flupentixol, verapamil, cyclosporin A, Hoechst 33342, and trans(E)-flupentixol) were tested for their effect on oxidative cross-linking. Substrates that stimulated the ATPase activity of P-gp (calcein-AM, demecolcine, cis(Z)-flupentixol, and verapamil) increased the rate of cross-linking between Cys(431)(NBD1-Walker A)/C1176C(NBD2-LSGGQ) and between Cys(1074)(NBD2-Walker A)/L531C(NBD1-LSGGQ) when compared with cross-linking in the absence of drug substrate. By contrast, substrates that inhibited ATPase activity (cyclosporin A, Hoechst 33342, and trans(E)-flupentixol) decreased the rate of cross-linking. These results indicate that interaction between the LSGGQ motifs and Walker A sites must be essential for coupling drug binding to ATP hydrolysis. Drug binding in the transmembrane domains can induce long range conformational changes in the NBDs, such that compounds that stimulate or inhibit ATPase activity must decrease and increase, respectively, the distance between the Walker A and LSGGQ sequences.

  6. Modulation of NFAT-5, an outlying member of the NFAT family, in human keratinocytes and skin

    PubMed Central

    Al-Daraji, Wael I; Afolayan, John; Zelger, Bettina G; Abdellaoui, Adel; Zelger, Bernhard

    2009-01-01

    Background Cyclosporin A (CsA) and tacrolimus block T cell activation by inhibiting the phosphatase calcineurin and preventing translocation from the cytoplasm to the nucleus of the transcription factor Nuclear Factor of Activated T cells (NFAT). NFAT compose a family of transcription factors that are turned on during T cell activation. Aims To study the expression of NFAT-5 mRNA and protein in normal human keratinocytes and to investigate the cellular and subcellular pattern of expression of NFAT-5 in normal human skin and psoriasis, and analyze effects of different agonists and ultraviolet radiation on NFAT-5 in normal human skin. Methods Tissue cultures, Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR), Western analysis, immunostaining, confocal microscopy. Results Sequencing of RT-PCR products confirmed the identity of the product that showed 100 % homology with the predicted NFAT-5 sequence. anti-NFAT-5 mainly detected a single band in cultured keratinocytes and dermal fibroblasts using Western analysis. Immunohistochemistry showed that epidermal keratinocytes and dermal fibroblasts in normal human and psoriatic skin express NFAT-5. NFAT-5 showed predominantly nuclear localization in epidermal keratinocytes and dermal fibroblasts within five normal adult skin biopsies. Our data also suggest that UV irradiation reduces NFAT-5 nuclear localization within the epidermis. Unlike NFAT 1-4, NFAT-5/TonEBP was localized to both nucleus and cytoplasm of cultured keratinocytes. Cyclosporin A induces nuclear membrane translocation of NFAT-5 in cultured keratinocytes and raffinose (a hypertonicity inducing agent) induces more nuclear localization of NFAT-5 compared to untreated cells. In addition, differentiation-promoting agonists that induce sustained rise in intracellular calcium did not result in changes in NFAT-5 localization in cultured keratinocytes. Conclusion These studies provide the first observation of expression of NFAT-5/TonEBP mRNA protein in

  7. P-glycoprotein is responsible for the poor intestinal absorption and low toxicity of oral aconitine: In vitro, in situ, in vivo and in silico studies

    SciTech Connect

    Yang, Cuiping Zhang, Tianhong Li, Zheng Xu, Liang Liu, Fei Ruan, Jinxiu Liu, Keliang Zhang, Zhenqing

    2013-12-15

    Aconitine (AC) is a highly toxic alkaloid from bioactive plants of the genus Aconitum, some of which have been widely used as medicinal herbs for thousands of years. In this study, we systematically evaluated the potential role of P-glycoprotein (P-gp) in the mechanisms underlying the low and variable bioavailability of oral AC. First, the bidirectional transport of AC across Caco-2 and MDCKII-MDR1 cells was investigated. The efflux of AC across monolayers of these two cell lines was greater than its influx. Additionally, the P-gp inhibitors, verapamil and cyclosporin A, significantly decreased the efflux of AC. An in situ intestinal perfusion study in rats showed that verapamil co-perfusion caused a significant increase in the intestinal permeability of AC, from 0.22 × 10{sup −5} to 2.85 × 10{sup −5} cm/s. Then, the pharmacokinetic profile of orally administered AC with or without pre-treatment with verapamil was determined in rats. With pre-treatment of verapamil, the maximum plasma concentration (C{sub max}) of AC increased sharply, from 39.43 to 1490.7 ng/ml. Accordingly, a 6.7-fold increase in the area under the plasma concentration–time curve (AUC{sub 0–12} {sub h}) of AC was observed when co-administered with verapamil. In silico docking analyses suggested that AC and verapamil possess similar P-gp recognition mechanisms. This work demonstrated that P-gp is involved in limiting the intestinal absorption of AC and attenuating its toxicity to humans. Our data indicate that potential P-gp-mediated drug–drug interactions should be considered carefully in the clinical application of aconite and formulations containing AC. - Highlights: • Verapamil and cyclosporin A decreased the efflux of aconitine across Caco-2 cells. • Both inhibitors decreased the efflux of aconitine across MDCKII-MDR1 cells. • Co-perfusion with verapamil increased the intestinal permeability of aconitine. • Co-administration with verapamil sharply increased the C{sub max

  8. Immune reconstitution following allogeneic peripheral blood progenitor cell transplantation: comparison of recipients of positive CD34+ selected grafts with recipients of unmanipulated grafts.

    PubMed

    Martínez, C; Urbano-Ispizua, A; Rozman, C; Marín, P; Rovira, M; Sierra, J; Montfort, N; Carreras, E; Montserrat, E

    1999-03-01

    We compared the kinetic recovery of lymphocytes and their subsets in two groups of patients submitted to allogeneic peripheral blood progenitor cell transplantation (allo-PBT): those receiving lymphocyte-depleted leukaphereses by positive selection of CD34+ cells (group 1, n = 18) and those receiving unmanipulated leukaphereses (group 2, n = 15). Patients were conditioned with cyclophosphamide (120 mg/kg) and fractioned total body irradiation (13 Gy, group 1; 12 Gy, group 2). The mean number (x 10(6)/kg) of CD34+ and CD3+ cells infused was 4.0 and 0.67, respectively, in group 1 patients, and 4.7 and 274, respectively, for group 2 patients. Graft-versus-host disease prophylaxis consisted of cyclosporin A + methylprednisolone for group 1 and cyclosporin A + methotrexate for group 2. Median follow-up was 7 months (range 2-8 months) for both groups. During the first 6 months post-transplant, CD4+ cell counts were lower in group 1 as compared with group 2 (p = 0.014, 0.010, 0.011, 0.0003, and 0.052 at 0.5, 1, 2, 3, and 6 months, respectively), whereas there was no difference at 8 months. The number of CD4+CD45RA+ cells was very low throughout the study in both groups, being lower in group 1 than in group 2, especially during the first 3 months post-transplant (p = 0.007 and 0.0006 at 1 and 3 months). Normal levels of CD8+ cells were reached by 1 month post-transplant in both groups. TCR gamma delta + cell counts were lower in group 1 than in group 2 during the first 4 months post-transplant (p = 0.001, 0.004, and 0.04 at 1, 3, and 4 months). A normal number of natural killer cells (CD3-CD56+) was achieved 1 month post-transplant in both groups. B lymphocytes (CD19+) showed low or undetectable counts throughout the first 4 months in both groups, achieving the normal range at 8 months. These results show that, during the first 6 months following allo-PBT with CD34+ selected grafts, the number of CD4+, CD4+CD45RA+, and TCR gamma delta + cells is significantly lower than

  9. Bilirubin and amyloid-beta peptide induce cytochrome c release through mitochondrial membrane permeabilization.

    PubMed Central

    Rodrigues, C. M.; Solá, S.; Silva, R.; Brites, D.

    2000-01-01

    BACKGROUND: The pathogenesis of bilirubin encephalopathy and Alzheimer's disease appears to result from accumulation of unconjugated bilirubin (UCB) and amyloid-beta (Abeta) peptide, respectively, which may cause apoptosis. Permeabilization of the mitochondrial membrane, with release of intermembrane proteins, has been strongly implicated in cell death. Inhibition of the mitochondrial permeability is one pathway by which ursodeoxycholate (UDC) and tauroursodeoxycholate (TUDC) protect against apoptosis in hepatic and nonhepatic cells. In this study, we further characterize UCB- and Abeta-induced cytotoxicty in isolated neural cells, and investigate membrane perturbation during incubation of isolated mitochondria with both agents. In addition, we evaluate whether the anti-apoptotic drugs UDC and TUDC prevent any changes from occurring. MATERIALS AND METHODS: Primary rat neuron and astrocyte cultures were incubated with UCB or Abeta peptide, either alone or in the presence of UDC. Apoptosis was assessed by DNA fragmentation and nuclear morphological changes. Isolated mitochondria were treated with each toxic, either alone or in combination with UDC, TUDC, or cyclosporine A. Mitochondrial swelling was measured spectrophotometrically and cytochrome c protein levels determined by Western blot. RESULTS: Incubation of neural cells with both UCB and Abeta induced apoptosis (p < 0.01). Coincubation with UDC reduced apoptosis by > 50% (p < 0.05). Both toxins caused membrane permeabilization in isolated mitochondria (p < 0.001); whereas, pretreatment with UDC was protective (p < 0.05). TUDC was even more effective at preventing matrix swelling mediated by Abeta (p < 0.01). UDC and TUDC markedly reduced cytochrome c release associated with mitochondrial permeabilization induced by UCB and Abeta, respectively (p < 0.05). Moreover, cyclosporine A significantly inhibited mitochondrial swelling and cytochrome c efflux mediated by UCB (p < 0.05). CONCLUSION: UCB and Abeta peptide

  10. The NFAT-1 DNA binding complex in activated T cells contains Fra-1 and JunB.

    PubMed Central

    Boise, L H; Petryniak, B; Mao, X; June, C H; Wang, C Y; Lindsten, T; Bravo, R; Kovary, K; Leiden, J M; Thompson, C B

    1993-01-01

    Activation of T cells induces transcription of the interleukin-2 (IL-2) gene. IL-2 expression is regulated through the binding of transcription factors to multiple sites within the IL-2 enhancer. One such cis-acting element within the IL-2 enhancer is the NFAT-1 (nuclear factor of activated T cells) binding site. NFAT-1 binding activity is absent in resting cells but is induced upon T-cell activation. The induction of NFAT-1 binding activity can be inhibited by cyclosporin A, potentially accounting for the ability of cyclosporin A to inhibit IL-2 production by T cells. We have previously reported that the NFAT-1 binding complex is composed of at least two proteins and that the 5' portion of the NFAT-1 sequence acts as a binding site for one or more proteins from the Ets family of transcription factors. We now report that the 3' portion of the NFAT-1 sequence contains a variant AP-1 binding site. NFAT-1 binding can be specifically inhibited by oligonucleotides containing a consensus AP-1 site. Moreover, mutation of the AP-1 site at the 3' end of the NFAT-1 sequence inhibits both NFAT-1 binding and the ability of the NFAT-1 binding site to activate expression from a reporter plasmid upon T-cell activation. Since AP-1 sites bind dimeric protein complexes composed of individual members of the Fos and Jun families of transcription factors, we used antibodies specific for individual Fos and Jun family members to determine whether they are present in the NFAT-1 binding complex. These experiments demonstrated that the NFAT-1 binding complex contains JunB and Fra-1 proteins. Northern (RNA) blot analyses demonstrate that both fra-1 and junB mRNAs are induced upon T-cell activation, although fra-1 mRNA is present even in quiescent T cells. Of interest, junB is not expressed in quiescent T cells, and it is induced with kinetics that are similar to those for the induction of IL-2 mRNA expression. Taken together, these results suggested that the JunB-Fra-1 heterodimer is the

  11. Interleukin-2 transcription is regulated in vivo at the level of coordinated binding of both constitutive and regulated factors.

    PubMed Central

    Garrity, P A; Chen, D; Rothenberg, E V; Wold, B J

    1994-01-01

    Interleukin-2 (IL-2) transcription is developmentally restricted to T cells and physiologically dependent on specific stimuli such as antigen recognition. Prior studies have shown that this stringent two-tiered regulation is mediated through a transcriptional promoter/enhancer DNA segment which is composed of diverse recognition elements. Factors binding to some of these elements are present constitutively in many cell types, while others are signal dependent, T cell specific, or both. This raises several questions about the molecular mechanism by which IL-2 expression is regulated. Is the developmental commitment of T cells reflected molecularly by stable interaction between available factors and the IL-2 enhancer prior to signal-dependent induction? At which level, factor binding to DNA or factor activity once bound, are individual regulatory elements within the native enhancer regulated? By what mechanism is developmental and physiological specificity enforced, given the participation of many relatively nonspecific elements? To answer these questions, we have used in vivo footprinting to determine and compare patterns of protein-DNA interactions at the native IL-2 locus in cell environments, including EL4 T-lymphoma cells and 32D clone 5 premast cells, which express differing subsets of IL-2 DNA-binding factors. We also used the immunosuppressant cyclosporin A as a pharmacological agent to further dissect the roles played by cyclosporin A-sensitive factors in the assembly and maintenance of protein-DNA complexes. Occupancy of all site types was observed exclusively in T cells and then only upon excitation of signal transduction pathways. This was true even though partially overlapping subsets of IL-2-binding activities were shown to be present in 32D clone 5 premast cells. This observation was especially striking in 32D cells because, upon signal stimulation, they mobilized a substantial set of IL-2 DNA-binding activities, as measured by in vitro assays using

  12. Recombinant interleukin 2 regulates levels of c-myc mRNA in a cloned murine T lymphocyte.

    PubMed Central

    Reed, J C; Sabath, D E; Hoover, R G; Prystowsky, M B

    1985-01-01

    The cellular oncogene c-myc has been implicated in the regulation of growth of normal and neoplastic cells. Recently, it was suggested that c-myc gene expression may control the G0----G1-phase transition in normal lymphocytes that were stimulated to enter the cell cycle by the lectin concanavalin A (ConA). Here we describe the effects of purified recombinant interleukin 2 (rIL2) and of ConA on levels of c-myc mRNA in the noncytolytic murine T-cell clone L2. In contrast to resting (G0) primary cultures of lymphocytes, quiescent L2 cells have a higher RNA content than resting splenocytes and express receptors for interleukin 2 (IL2). Resting L2 cells are therefore best regarded as early G1-phase cells. Purified rIL2 was found to stimulate the rapid accumulation of c-myc mRNA in L2 cells. Levels of c-myc mRNA became maximal within 1 h and declined gradually thereafter. In contrast, ConA induced slower accumulation of c-myc mRNA in L2 cells, with increased levels of c-myc mRNA becoming detectable 4 to 8 h after stimulation. Experiments with the protein synthesis inhibitor cycloheximide demonstrated that the increase in levels of c-myc mRNA that were induced by ConA was a direct effect of this lectin and not secondary to IL2 production. Cyclosporin A, an immunosuppressive agent, markedly reduced the accumulation of c-myc mRNA that was induced by ConA but only slightly diminished the accumulation of c-myc mRNA that was induced by rIL2. Taken together, these data provide evidence that (i) c-myc gene expression can be regulated by at least two distinct pathways in T lymphocytes, only one of which is sensitive to cyclosporine A, and (ii) the accumulation of c-myc mRNA can be induced in T cells by IL2 during the G1 phase of the cell cycle. Images PMID:3879814

  13. Mitochondrial aquaporin-8 knockdown in human hepatoma HepG2 cells causes ROS-induced mitochondrial depolarization and loss of viability

    SciTech Connect

    Marchissio, Maria Julia; Francés, Daniel Eleazar Antonio; Carnovale, Cristina Ester; Marinelli, Raúl Alberto

    2012-10-15

    Human aquaporin-8 (AQP8) channels facilitate the diffusional transport of H{sub 2}O{sub 2} across membranes. Since AQP8 is expressed in hepatic inner mitochondrial membranes, we studied whether mitochondrial AQP8 (mtAQP8) knockdown in human hepatoma HepG2 cells impairs mitochondrial H{sub 2}O{sub 2} release, which may lead to organelle dysfunction and cell death. We confirmed AQP8 expression in HepG2 inner mitochondrial membranes and found that 72 h after cell transfection with siRNAs targeting two different regions of the human AQP8 molecule, mtAQP8 protein specifically decreased by around 60% (p < 0.05). Studies in isolated mtAQP8-knockdown mitochondria showed that H{sub 2}O{sub 2} release, assessed by Amplex Red, was reduced by about 45% (p < 0.05), an effect not observed in digitonin-permeabilized mitochondria. mtAQP8-knockdown cells showed an increase in mitochondrial ROS, assessed by dichlorodihydrofluorescein diacetate (+ 120%, p < 0.05) and loss of mitochondrial membrane potential (− 80%, p < 0.05), assessed by tetramethylrhodamine-coupled quantitative fluorescence microscopy. The mitochondria-targeted antioxidant MitoTempol prevented ROS accumulation and dissipation of mitochondrial membrane potential. Cyclosporin A, a mitochondrial permeability transition pore blocker, also abolished the mtAQP8 knockdown-induced mitochondrial depolarization. Besides, the loss of viability in mtAQP8 knockdown cells verified by MTT assay, LDH leakage, and trypan blue exclusion test could be prevented by cyclosporin A. Our data on human hepatoma HepG2 cells suggest that mtAQP8 facilitates mitochondrial H{sub 2}O{sub 2} release and that its defective expression causes ROS-induced mitochondrial depolarization via the mitochondrial permeability transition mechanism, and cell death. -- Highlights: ► Aquaporin-8 is expressed in mitochondria of human hepatoma HepG2 cells. ► Aquaporin-8 knockdown impairs mitochondrial H{sub 2}O{sub 2} release and increases ROS. ► Aquaporin

  14. Protein kinase C-α interaction with iHSP70 in mitochondria promotes recovery of mitochondrial function after injury in renal proximal tubular cells.

    PubMed

    Nowak, Grazyna; Soundararajan, Sridharan; Mestril, Ruben

    2013-09-01

    This study determined the role of PKC-α and associated inducible heat shock protein 70 (iHSP70) in the repair of mitochondrial function in renal proximal tubular cells (RPTCs) after oxidant injury. Wild-type PKC-α (wtPKC-α) and an inactive PKC-α [dominant negative dn; PKC-α] mutant were overexpressed in primary cultures of RPTCs, and iHSP70 levels and RPTC regeneration were assessed after treatment with the oxidant tert-butylhydroperoxide (TBHP). TBHP exposure increased ROS production and induced RPTC death, which was prevented by ferrostatin and necrostatin-1 but not by cyclosporin A. Overexpression of wtPKC-α maintained mitochondrial levels of active PKC-α, reduced cell death, and accelerated proliferation without altering ROS production in TBHP-injured RPTCs. In contrast, dnPKC-α blocked proliferation and monolayer regeneration. Coimmunoprecipitation and proteomic analysis demonstrated an association between inactive, but not active, PKC-α and iHSP70 in mitochondria. Mitochondrial iHSP70 levels increased as levels of active PKC-α decreased after injury. Overexpression of dnPKC-α augmented, whereas overexpression of wtPKC-α abrogated, oxidant-induced increases in mitochondrial iHSP70 levels. iHSP70 overexpression (1) maintained mitochondrial levels of phosphorylated PKC-α, (2) improved the recovery of state 3 respiration and ATP content, (3) decreased RPTC death (an effect abrogated by cyclosporine A), and (4) accelerated proliferation after oxidant injury. In contrast, iHSP70 inhibition blocked the recovery of ATP content and exacerbated RPTC death. Inhibition of PKC-α in RPTC overexpressing iHSP70 blocked the protective effects of iHSP70. We conclude that active PKC-α maintains mitochondrial function and decreases cell death after oxidant injury. iHSP70 is recruited to mitochondria in response to PKC-α dephosphorylation and associates with and reactivates inactive PKC-α, which promotes the recovery of mitochondrial function, decreases RPTC

  15. Sequential reduction of mitochondrial transmembrane potential and generation of reactive oxygen species in early programmed cell death

    PubMed Central

    1995-01-01

    Programmed cell death (PCD) is a physiological process commonly defined by alterations in nuclear morphology (apoptosis) and/or characteristic stepwise degradation of chromosomal DNA occurring before cytolysis. However, determined characteristics of PCD such as loss in mitochondrial reductase activity or cytolysis can be induced in enucleated cells, indicating cytoplasmic PCD control. Here we report a sequential disregulation of mitochondrial function that precedes cell shrinkage and nuclear fragmentation. A first cyclosporin A-inhibitable step of ongoing PCD is characterized by a reduction of mitochondrial transmembrane potential, as determined by specific fluorochromes (5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine++ + iodide; 3,3'dihexyloxacarbocyanine iodide). Cytofluorometrically purified cells with reduced mitochondrial transmembrane potential are initially incapable of oxidizing hydroethidine (HE) into ethidium. Upon short-term in vitro culture, such cells acquire the capacity of HE oxidation, thus revealing a second step of PCD marked by mitochondrial generation of reactive oxygen species (ROS). This step can be selectively inhibited by rotenone and ruthenium red yet is not affected by cyclosporin A. Finally, cells reduce their volume, a step that is delayed by radical scavengers, indicating the implication of ROS in the apoptotic process. This sequence of alterations accompanying early PCD is found in very different models of apoptosis induction: glucocorticoid-induced death of lymphocytes, activation-induced PCD of T cell hybridomas, and tumor necrosis factor-induced death of U937 cells. Transfection with the antiapoptotic protooncogene Bcl-2 simultaneously inhibits mitochondrial alterations and apoptotic cell death triggered by steroids or ceramide. In vivo injection of fluorochromes such as 5,5',6,6'-tetrachloro-1,1',3,3'- tetraethylbenzimidazolcarbocyanine iodide; 3,3'dihexyloxacarbocyanine iodide; or HE allows for the detection of

  16. Haploidentical T Cell-Replete Transplantation with Post-Transplantation Cyclophosphamide for Patients in or above the Sixth Decade of Age Compared with Allogeneic Hematopoietic Stem Cell Transplantation from an Human Leukocyte Antigen-Matched Related or Unrelated Donor.

    PubMed

    Blaise, Didier; Fürst, Sabine; Crocchiolo, Roberto; El-Cheikh, Jean; Granata, Angela; Harbi, Samia; Bouabdallah, Reda; Devillier, Raynier; Bramanti, Stephania; Lemarie, Claude; Picard, Christophe; Chabannon, Christian; Weiller, Pierre-Jean; Faucher, Catherine; Mohty, Bilal; Vey, Norbert; Castagna, Luca

    2016-01-01

    It has recently been shown that a T cell-replete allogeneic (allo) hematopoietic stem cell transplantation (HSCT) from a haploidentical donor (haplo-ID) could be a valid treatment for hematological malignancies. However, little data exist concerning older populations. We provided transplantation to 31 patients over the age of 55 years from a haplo-ID and compared their outcomes with patients of the same ages who underwent transplantation from a matched related (MRD) or an unrelated donor (UD). All 3 groups were comparable, except for their conditioning. Patients in haplo-ID group received 2 days of post-transplantation high-dose cyclophosphamide followed by cyclosporine A and mycophenolate mofetil, whereas patients in other groups received pretransplantation antithymocyte globulin, cyclosporine A, and additional mycophenolate mofetil in case of 1-antigen mismatch. All patients but 1 in the haplo-ID group engrafted. The incidence of grades 2 to 4 acute graft-versus-host disease (GVHD) was not statistically different between recipients from haplo-ID (cumulative incidence, 23%) and MRD (cumulative incidence, 21%) transplantations but it was lower than after UD HSCT (cumulative incidence, 44%). No patient in the haplo-ID group developed severe chronic GVHD, compared with cumulative incidences of 16% and 14% after MRD (P = .02) and UD (P = .03) grafts, respectively. The cumulative incidences of relapse were similar in the 3 groups, whereas nonrelapse mortality after UD HSCT was 3-fold higher than after haplo-ID or MRD HSCT. Overall, 2-year overall survival (70%), progression-free survival (67%), and progression and severe chronic GVHD-free survival (67%) probabilities after haplo-ID did not statistically differ from MRD transplantation (78%, 64%, and 51%, respectively), although they were higher than after UD transplantation (51% [P = .08], 38% [P = .02], and 31% [P = .007]). We conclude that T cell-replete haplo-ID HSCT followed by post-transplantation high

  17. NFAT2 mediates high glucose-induced glomerular podocyte apoptosis through increased Bax expression

    SciTech Connect

    Li, Ruizhao; Zhang, Li; Shi, Wei; Zhang, Bin; Liang, Xinling; Liu, Shuangxin; Wang, Wenjian

    2013-04-15

    Background: Hyperglycemia promotes podocyte apoptosis and plays a key role in the pathogenesis of diabetic nephropathy. However, the mechanisms that mediate hyperglycemia-induced podocyte apoptosis is still far from being fully understood. Recent studies reported that high glucose activate nuclear factor of activated T cells (NFAT) in vascular smooth muscle or pancreatic β-cells. Here, we sought to determine if hyperglycemia activates NFAT2 in cultured podocyte and whether this leads to podocyte apoptosis. Meanwhile, we also further explore the mechanisms of NFAT2 activation and NFAT2 mediates high glucose-induced podocyte apoptosis. Methods: Immortalized mouse podocytes were cultured in media containing normal glucose (NG), or high glucose (HG) or HG plus cyclosporine A (a pharmacological inhibitor of calcinerin) or 11R-VIVIT (a special inhibitor of NFAT2). The activation of NFAT2 in podocytes was detected by western blotting and immunofluorescence assay. The role of NFAT2 in hyperglycemia-induced podocyte apoptosis was further evaluated by observing the inhibition of NFAT2 activation by 11R-VIVIT using flow cytometer. Intracellular Ca{sup 2+} was monitored in HG-treated podcocytes using Fluo-3/AM. The mRNA and protein expression of apoptosis gene Bax were measured by real time-qPCR and western blotting. Results: HG stimulation activated NFAT2 in a time- and dose-dependent manner in cultured podocytes. Pretreatment with cyclosporine A (500 nM) or 11R-VIVIT (100 nM) completely blocked NFAT2 nuclear accumulation. Meanwhile, the apoptosis effects induced by HG were also abrogated by concomitant treatment with 11R-VIVIT in cultured podocytes. We further found that HG also increased [Ca{sup 2+}]i, leading to activation of calcineurin, and subsequent increased nuclear accumulation of NFAT2 and Bax expression in cultured podocytes. Conclusion: Our results identify a new finding that HG-induced podocyte apoptosis is mediated by calcineurin/NFAT2/Bax signaling pathway

  18. Noninvasive Imaging of Myocardial Inflammation in Myocarditis using 68Ga-tagged Mannosylated Human Serum Albumin Positron Emission Tomography

    PubMed Central

    Lee, Seung-Pyo; Im, Hyung-Jun; Kang, Shinae; Chung, Seock-Jin; Cho, Ye Seul; Kang, Hyejeong; Park, Ho Seon; Hwang, Do-Won; Park, Jun-Bean; Paeng, Jin-Chul; Cheon, Gi-Jeong; Lee, Yun-Sang; Jeong, Jae Min; Kim, Yong-Jin

    2017-01-01

    The diagnosis of myocarditis traditionally relies on invasive endomyocardial biopsy but none of the imaging studies so far are specific for infiltration of the inflammatory cells itself. We synthesized 68Ga-2-(p-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid (NOTA) mannosylated human serum albumin (MSA) by conjugating human serum albumin with mannose, followed by conjugation with NOTA and labeling it with 68Ga. The efficacy of 68Ga-NOTA-MSA positron emission tomography (PET) for imaging myocardial inflammation was tested in a rat myocarditis model. A significant number of mannose receptor-positive inflammatory cells infiltrated the myocardium in both human and rat myocarditis tissue. 68Ga-NOTA-MSA uptake was upregulated in organs of macrophage accumulation, such as liver, spleen, bone marrow and myocardium (0.32 (0.31~0.33) for normal versus 1.02 (0.86~1.06) for myocarditis (median (range), SUV); n=4~6 per group, p-value=0.01). 68Ga-NOTA-MSA uptake in the left ventricle was upregulated in myocarditis compared with normal rats (2.29 (1.42~3.40) for normal versus 4.18 (3.43~6.15) for myocarditis (median (range), average standard uptake value ratio against paraspinal muscle); n=6 per group, p-value<0.01), which was downregulated in rats with cyclosporine-A treated myocarditis (3.69 (2.59~3.86) for myocarditis versus 2.28 (1.76~2.60) for cyclosporine-A treated myocarditis; n=6 per group, p-value<0.01). The specificity of the tracer was verified by administration of excess non-labeled MSA. 68Ga-NOTA-MSA uptake was significantly enhanced earlier in the evolution of myocarditis before any signs of inflammation could be seen on echocardiography. These results demonstrate the potential utility of visualizing infiltration of mannose receptor-positive macrophages with 68Ga-NOTA-MSA PET in the early diagnosis of as well as in the monitoring of treatment response of myocarditis. PMID:28042344

  19. An Integrated Model of Atopic Dermatitis Biomarkers Highlights the Systemic Nature of the Disease.

    PubMed

    Ungar, Benjamin; Garcet, Sandra; Gonzalez, Juana; Dhingra, Nikhil; Correa da Rosa, Joel; Shemer, Avner; Krueger, James G; Suarez-Farinas, Mayte; Guttman-Yassky, Emma

    2017-03-01

    Current atopic dermatitis (AD) models link epidermal abnormalities in lesional skin to cytokine activation. However, there is evolving evidence of systemic immune activation and detectable abnormalities in nonlesional skin. Because some of the best single correlations with severity (Scoring of AD, or SCORAD) are detected not only in lesional but also nonlesional skin and blood, more complex biomarker models of AD are needed. We thus performed extensive biomarker measures in these compartments using univariate and multivariate approaches to correlate disease biomarkers with SCORAD and with a combined hyperplasia score [thickness and keratin 16 (K16) mRNA] at baseline and after cyclosporine A treatment in 25 moderate to severe AD patients. Increases in serum cytokines and chemokines (IL-13, IL-22, CCL17) were found in AD versus healthy individuals and were reduced with treatment. SCORAD correlated with immune (IL-13, IL-22) and epidermal (thickness, K16) measures in lesional and, even more strongly, in nonlesional AD. Serum cytokines also had higher correlations with nonlesional markers at baseline and with treatment. Multivariate U statistics improved baseline and treatment-response SCORAD correlations. Nonlesional models showed the strongest correlations, with further improvement upon integration of serum markers. Even better correlations were obtained between biomarkers and the hyperplasia score. Larger cohorts are needed to confirm these preliminary data.

  20. Identification of small-molecule inhibitors of calcineurin-NFATc signaling that mimic the PxIxIT motif of calcineurin binding partners.

    PubMed

    Matsoukas, Minos-Timotheos; Aranguren-Ibáñez, Álvaro; Lozano, Teresa; Nunes, Virginia;