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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. Cyclosporine A enhances platelet aggregation.

    PubMed

    Grace, A A; Barradas, M A; Mikhailidis, D P; Jeremy, J Y; Moorhead, J F; Sweny, P; Dandona, P

    1987-12-01

    In view of the reported increase in thromboembolic episodes following cyclosporine A (CyA) therapy, the effect of this drug on platelet aggregation and thromboxane A2 release was investigated. The addition of CyA, at therapeutic concentrations to platelet rich plasma from normal subjects in vitro was found to increase aggregation in response to adrenaline, collagen and ADP. Ingestion of CyA by healthy volunteers was also associated with enhanced platelet aggregation. The CyA-mediated enhancement of aggregation was further enhanced by the addition in vitro of therapeutic concentrations of heparin. Platelets from renal allograft recipients treated with CyA also showed hyperaggregability and increased thromboxane A2 release, which were most marked at "peak" plasma CyA concentration and less so at "trough" concentrations. Platelet hyperaggregability in renal allograft patients on long-term CyA therapy tended to revert towards normal following the replacement of CyA with azathioprine. Hypertensive patients with renal allografts on nifedipine therapy had normal platelet function and thromboxane release in spite of CyA therapy. These observations suggest that CyA-mediated platelet activation may contribute to the pathogenesis of the thromboembolic phenomena associated with the use of this drug. The increased release of thromboxane A2 (a vasoconstrictor) may also play a role in mediating CyA-related nephrotoxicity.

  3. Modeling conformational changes in cyclosporin A.

    PubMed Central

    O'Donohue, M. F.; Burgess, A. W.; Walkinshaw, M. D.; Treutlein, H. R.

    1995-01-01

    NMR and X-ray structures for the immunosuppressant cyclosporin A (CsA) reveal a remarkable difference between the unbound (free) conformation in organic solvents and the conformation bound to cyclophilin. We have performed computer simulations of the molecular dynamics of CsA under a variety of conditions and confirmed the stability of these two conformations at room temperature in water and in vacuum. However, when the free conformation was modeled in vacuum at 600 K, a transition pathway leading to the bound conformation was observed. This involved a change in the cis MeLeu-9 peptide bond to a trans conformation and the movement of the side chains forming the dominant hydrophobic cluster (residues MeBmt-1, MeLeu-4, MeLeu-6, and MeLeu-10) to the opposite side of the plane formed by the backbone atoms in the molecular ring. The final conformation had a backbone RMS deviation from the bound conformation of 0.53 A and was as stable in dynamics simulations as the bound conformation. Our calculations allowed us to make a detailed analysis of a transition pathway between the free and the bound conformations of CsA and to identify two distinct regions of coordinated movement in CsA, both of which underwent transitions independently. PMID:8535256

  4. Importance of endogenous prostaglandins for the toxicity of cyclosporin A to rat endocrine and exocrine pancreas?

    PubMed Central

    Rünzi, M; Peskar, B M; von Schönfeld, J; Müller, M K

    1992-01-01

    Previous work has shown that cyclosporin A is toxic to the endocrine and exocrine pancreas. The aim of this study was to examine whether endogenous eicosanoids play a role in controlling cyclosporin A induced toxicity. Rats were treated for eight days with indomethacin (2 mg/kg, twice daily) in addition to cyclosporin A (5 or 10 mg/kg daily). Effects of drug treatments on exocrine (as assessed by amylase and protein secretion into the pancreatic juice) and endocrine (as assessed by the glucose dependent insulin release) pancreatic functions, and pancreatic formation of prostaglandins and thromboxane were evaluated. Treatment with cyclosporin A in the doses used did not inhibit eicosanoid formation by the pancreatic tissue ex vivo. Indomethacin caused significant inhibition of pancreatic formation of prostaglandin E2, 6k prostaglandin F1 alpha and thromboxane B2. Combined treatment with indomethacin and cyclosporin A (5 or 10 mg/kg) augmented cyclosporin A induced pancreatic toxicity with further impairment of insulin release, amylase secretion, and pancreatic juice protein content, but did not result in more pronounced inhibition of pancreatic eicosanoid formation. The increased toxicity of the combined treatment was, however, associated with raised cyclosporin A whole blood concentrations. The data suggest that the potentiation of pancreatic toxicity of cyclosporin A observed during coadministration of indomethacin is not the result of suppression of endogenous pancreatic eicosanoid biosynthesis, but more likely results from altered cyclosporin A pharmacokinetic which may be caused by an interference of indomethacin with the hepatic cytochrome P-450 dependent monooxygenase involved in cyclosporin A metabolism. The possibility that coadministration of non-steroidal antiinflammatory drugs aggravates toxic effects in cyclosporin A treated patients should be considered. PMID:1280611

  5. Cyclosporin A inhibits the replication of diverse coronaviruses.

    PubMed

    de Wilde, Adriaan H; Zevenhoven-Dobbe, Jessika C; van der Meer, Yvonne; Thiel, Volker; Narayanan, Krishna; Makino, Shinji; Snijder, Eric J; van Hemert, Martijn J

    2011-11-01

    Low micromolar, non-cytotoxic concentrations of cyclosporin A (CsA) strongly affected the replication of severe acute respiratory syndrome coronavirus (SARS-CoV), human coronavirus 229E and mouse hepatitis virus in cell culture, as was evident from the strong inhibition of GFP reporter gene expression and a reduction of up to 4 logs in progeny titres. Upon high-multiplicity infection, CsA treatment rendered SARS-CoV RNA and protein synthesis almost undetectable, suggesting an early block in replication. siRNA-mediated knockdown of the expression of the prominent CsA targets cyclophilin A and B did not affect SARS-CoV replication, suggesting either that these specific cyclophilin family members are dispensable or that the reduced expression levels suffice to support replication. PMID:21752960

  6. 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.

  7. Parasite cyclophilins and antiparasite activity of cyclosporin A.

    PubMed

    Page, A P; Kumar, S; Carlow, C K

    1995-10-01

    Cyclosporin A (CsA) was initially developed as an immunosuppressive drug. In the past several years, it has been shown to possess antiparasite activity independent of the immune system. It is not known how the drug exerts these antiparasite effects, or why it is stage and/or species specific. The answers may lie in the enzymatic function of cyclophilins. The cyclophilins are a growing family of proteins that exhibit peptidyl-prolyl cis-trans isomerase (PPiase) activity and bid CsA to varying degrees. PPiases have been shown to play a role in the folding of many essential proteins. Antony Page, Sanjay Kumar and Clotilde Carlow here review parasite cyclophilins and their association with CsA. The possible biological function of parasite cyclophilins and their potential role in future drug discovery are also discussed.

  8. L-arginine prevents bone loss and bone collagen breakdown in cyclosporin A-treated rats.

    PubMed

    Fiore, C E; Pennisi, P; Cutuli, V M; Prato, A; Messina, R; Clementi, G

    2000-11-24

    Cyclosporin A is implicated in the pathogenesis of post-transplantation bone disease. Because of recent evidence that cyclosporin A may cause renal and cardiovascular toxicity by inhibiting nitric oxide (NO) activity, and that NO slows bone remodeling and bone loss in animal and human studies, we investigated a possible link between NO production and beneficial effects on bone health in cyclosporin A-treated rats. Thirty-six 10-week-old male rats were assigned to six groups of six animals each, and treated for 4 weeks with: vehicle; cyclosporin A; L-arginine; N(G)-nitro-L-arginine methylester (L-NAME, a general inhibitor of NO synthase activity); a combination of cyclosporin A+L-arginine; and a combination of cyclosporin A+L-NAME. Whole body and regional (spine and pelvis) bone mineral content of rats were measured under basal conditions and at the end of the treatment period by dual-energy X-ray absorptiometry (DXA) scanning. Femur weights and serum concentrations of pyridinoline, a reliable marker of bone resorption, were measured at the end of the study period. Cyclosporin A-, L-NAME-, and cyclosporin A+L-NAME-treated rats had significantly lower bone mineral content and femur weights, and significantly higher pyridinoline levels than did control animals. The administration of L-arginine appeared to prevent bone loss caused by cyclosporin A, suggesting that this amino acid, which can be converted to produce NO, might prove useful in preventing disturbed bone modeling and inhibition of bone growth associated with cyclosporin A therapy. PMID:11090650

  9. Cyclosporine A stimulated hair growth from mouse vibrissae follicles in an organ culture model

    PubMed Central

    Xu, Wenrong; Fan, Weixin; Yao, Kun

    2012-01-01

    Hypertrichosis is one of the most common side effects of systemic cyclosporine A therapy. It has been previously shown that cyclosporine A induces anagen and inhibits catagen development in mice. In the present study, to explore the mechanisms of cyclosporine A, we investigated the effects of cyclosporine A on hair shaft elongation, hair follicle cell proliferation, apoptosis, and mRNA expression of selected growth factors using an organ culture model of mouse vibrissae. In this model, cyclosporine A stimulated hair growth of normal mouse vibrissae follicles by inhibiting catagen-like development and promoting matrix cell proliferation. In addition, cyclosporine A caused an increase in the expression of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and nerve growth factor (NGF), and inhibited follistatin expression. Our findings provide an explanation for the clinically observed effects of cyclosporine A on hair growth. The mouse vibrissae organ culture offers an attractive model for identifying factors involved in the modulation of hair growth. PMID:23554774

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

    PubMed

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

    2001-03-15

    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.

  11. Cyclosporine A-loaded lipid nanoparticles in inflammatory bowel disease.

    PubMed

    Guada, Melissa; Beloqui, Ana; Alhouayek, Mireille; Muccioli, Giulio G; Dios-Viéitez, Maria del Carmen; Préat, Véronique; Blanco-Prieto, Maria J

    2016-04-30

    Cyclosporine A (CsA) is a well-known immunosuppressive agent used as rescue therapy in severe steroid-refractory ulcerative colitis (UC). However, toxicity issues associated with CsA when administered in its commercially available formulations have been reported in clinical practice. Since nanotechnology has been proposed as a promising strategy to improve safety and efficacy in the treatment of inflammatory bowel disease (IBD), the main purpose of this study was to evaluate the effect of oral administration of CsA-loaded lipid nanoparticles (LN) in the dextran sodium sulfate (DSS)-induced colitis mouse model using Sandimmune Neoral(®) as reference. The results showed that the formulations used did not decrease colon inflammation in terms of myeloperoxidase activity (MPO), tumor necrosis factor (TNF)-α expression, or histological scoring in the acute stage of the disease. However, further studies are needed in order to corroborate the efficacy of these formulations in the chronic phase of the disease. PMID:26972380

  12. Suppression of feline coronavirus replication in vitro by cyclosporin A.

    PubMed

    Tanaka, Yoshikazu; Sato, Yuka; Osawa, Shuichi; Inoue, Mai; Tanaka, Satoka; Sasaki, Takashi

    2012-04-30

    The feline infectious peritonitis virus (FIPV) is a member of the feline coronavirus family that causes FIP, which is incurable and fatal in cats. Cyclosporin A (CsA), an immunosuppressive agent that targets the nuclear factor pathway of activated T-cells (NF-AT) to bind cellular cyclophilins (CyP), dose-dependently inhibited FIPV replication in vitro. FK506 (an immunosuppressor of the pathway that binds cellular FK506-binding protein (FKBP) but not CyP) did not affect FIPV replication. Neither cell growth nor viability changed in the presence of either CsA or FK506, and these factors did not affect the NF-AT pathway in fcwf-4 cells. Therefore, CsA does not seem to exert inhibitory effects via the NF-AT pathway. In conclusion, CsA inhibited FIPV replication in vitro and further studies are needed to verify the practical value of CsA as an anti-FIPV treatment in vivo.

  13. Cyclosporine A protects podocytes by regulating WAVE1 phosphorylation

    PubMed Central

    Li, Xuejuan; Ding, Fangrui; Wang, Suxia; Li, Baihong; Ding, Jie

    2015-01-01

    Accumulating evidence suggests that podocytes are direct targets of many classic antiproteinuric drugs. The immunosuppressive drug cyclosporine A (CsA), which is a calcineurin inhibitor, is used to treat proteinuric kidney diseases. One novel mechanism by which CsA reduces proteinuria is by directly stabilizing the podocyte cytoskeleton. Previous studies showed that calcineurin can directly regulate WAVE1 within mouse striatal slices. In this study, WAVE1 was expressed in podocytes and was localized in the podocyte cell bodies and foot processes (FPs). WAVE1 expression increased in both in vivo and in vitro models of puromycin aminonucleoside (PAN)-induced podocyte injury. CsA restored WAVE1 expression and also partially rescued the disordered F-actin arrangement after PAN injury. Co-immunoprecipitation assays showed that calcineurin directly interacted with WAVE1 and regulated WAVE1 phosphorylation in podocytes. Synaptopodin is a well-characterized target of CsA. WAVE1 overexpression and synaptopodin knockdown experiments directly demonstrated that WAVE1 expression is not dependent on synaptopodin expression, and vice versa. Overexpression of WAVE1 using a WAVE1 plasmid disrupted F-actin structure and promoted podocyte migration compared with the empty vector group. Therefore, WAVE1 may be a novel molecular target for the maintenance of podocyte FPs and for antiproteinuric treatment in the future. PMID:26634693

  14. Supersaturated polymeric micelles for oral cyclosporine A delivery.

    PubMed

    Yu, Hongzhen; Xia, Dengning; Zhu, Quanlei; Zhu, Chunliu; Chen, Dan; Gan, Yong

    2013-11-01

    Polymeric micelles provide a promising platform for improving oral absorption of poorly soluble drugs. However, improved understanding of how drug retention within the hydrophobic micelle core can reduce drug absorption is required. We designed supersaturated polymeric micelles (Super-PMs) to increase molecularly dissolved drug concentration and gain an insight into the effect of the degree of supersaturation on oral absorption of cyclosporine A (CsA) in rats. The drug release from Super-PMs increased with an increase in initial supersaturation degrees in micelles. The cellular uptake of coumarin-6 was reduced by the retention of drug in polymer micelles. The transport flux of CsA across Caco-2 monolayer was increased with initial supersaturation degrees of 0.81-3.53 (p < 0.05). However, increase in supersaturation to 5.64 actually resulted in decreased CsA transport. The same trend was observed in a rat in vivo absorption study, in which the highest bioavailability of 134.6 ± 24.7% (relative to a commercial product, Sandimmun Neoral®, p<0.01) was achieved when the supersaturation degree was 3.53. These results demonstrated that Super-PMs were a promising drug delivery system for compounds with low aqueous solubility. This study also provided an experimental proof for the hypothesis that moderately supersaturated formulations are valuable alternative to high supersaturation formulations, resulting in optimal in vivo performance, and the degree of supersaturation should be carefully controlled to optimize drug absorption.

  15. Skeletal effects of cyclosporin A are gender related in rats.

    PubMed

    Erben, Reinhold G; Brunner, Katrin S; Breig, Bianca; Eberle, Johannes; Goldberg, Michel; Hofbauer, Lorenz C

    2003-01-01

    The immunosuppressive drug cyclosporin A (CsA) is thought to be involved in the pathogenesis of posttransplantation osteoporosis. To evaluate further the skeletal effects of CsA, we treated aged male and female sham-operated and gonadectomized rats with low doses of CsA for 4 months. Here, we show that CsA is antiresorptive and bone-sparing in aged female rats but increases bone resorption and reduces bone mass in aged male rats. However, even in male rats, CsA treatment, at clinically relevant doses, increased bone resorption only transiently and did not result in pronounced long-term cancellous bone loss. The gender-specific skeletal effects of CsA were not modulated by sex hormones or gonadectomy. CsA did not influence sex steroid metabolism in male or female rats. However, endogenous estradiol in sham-operated female rats (and especially, exogenous administration of 17beta-estradiol in ovariectomized rats) markedly diminished blood levels of CsA, probably by increasing hepatic CsA metabolism. Although the mechanism for the gender-specific skeletal effects of CsA is still obscure, our findings may have important implications for clinical therapy with CsA.

  16. Chronic pyelonephritis: Modulation of host defenses by cyclosporin A

    SciTech Connect

    Findon, G.; Miller, T.E. )

    1989-08-01

    Chronic experimental pyelonephritis is characterized by a stable level of infection, which persists for many months. Administration of cyclosporin A (CsA) reactivated previously healed renal lesions and caused a marked increase in bacterial numbers in the kidney. Studies were then carried out to compare the effects of CsA, and the nonselective cytodepletive agents irradiation and cyclophosphamide, on both host defenses and the bacteriologic status of chronically infected kidneys. Two different responses were observed. In animals treated with CsA, bacterial numbers increased markedly, although circulating neutrophil numbers were relatively unaffected. This observation was in contrast to the severe ablation of leukocyte numbers and competence needed to achieve an equivalent effect when irradiation and cyclophosphamide were used. One possible explanation for the adverse effect of CsA on the host-parasite balance in chronic pyelonephritis is that CsA affects mediators that control the inflammatory response or induces a qualitative change in a critical cellular defense compartment.

  17. Kinetics of acid-catalyzed degradation of cyclosporin A and its analogs in aqueous solution.

    PubMed

    Oliyai, R; Safadi, M; Meier, P G; Hu, M K; Rich, D H; Stella, V J

    1994-03-01

    The kinetics and mechanism of the degradation of cyclosporin A have been studied under aqueous acidic conditions. The rate of degradation was found to be specific acid-catalyzed over the pH range studied (1-4), with isocyclosporin A as the predominant degradation product. Selective reduction of the olefinic bond of the amino acid 2-N-methyl-(R)-((E)-2-butenyl)-4-methyl-L-threonine (MeBmt) did not affect the overall degradation kinetics and product distribution of cyclosporin A. These observations indicate that the alternative degradation pathway involving intramolecular alkoxy addition to the olefinic bond of amino acid MeBmt apparently does not significantly contribute to the overall degradation kinetics of cyclosporin A in the pH range 1-4. The chemical reactivity of O-acetyl-cyclosporin A was examined to probe the governing mechanism for the isomerization of cyclosporin A. Under identical conditions, O-acetyl-cyclosporin A showed a much greater chemical stability than cyclosporin A, consistent with a mechanism involving the hydroxyoxazolidine intermediate. The chemical stability of cyclosporin C, which contains two beta-hydroxyl groups, was also examined. The rate and product distribution for the degradation of cyclosporin C suggest that under aqueous acidic conditions it undergoes N,O-acyl migration solely at the amino acid residue MeBmt. Additionally, the impact of side-chain bulkiness of amino acid MeBmt was examined by studying the degradation kinetics of a series of cyclosporin A analogs.(ABSTRACT TRUNCATED AT 250 WORDS)

  18. Development of cyclosporine A microemulsion for parenteral delivery.

    PubMed

    Yuan, Yue; Che, Xin; Zhao, Mingyi; Wang, Yan; Liu, Yajun; Schwendeman, Anna; Li, Sanming

    2015-01-01

    The goal of this study was to develop a parenteral microemulsion formulation of cyclosporine A (CyA). The CyA solubility in caprylic capric triglyceride (GTCC), ethyl oleate and soybean oil were determined. The pseudo-ternary diagrams of oil (GTCC), surfactant (Solutol® HS-15), cosurfactants (ethanol/polyethylene glycol 400 [PEG 400] mixture) and water were constructed to identify boundaries for microemulsion existence. The CyA was added at 3, 6 and 9% w/w to the optimal microemulsion composition. Microemulsion particle size, solution viscosity and conductivity were examined. The microemulsion stability and haemolytic potential were examined after dilution in 5% dextrose solution for injection to 1 mg/mL CyA. Microemulsion stability was examined after a three-month storage at 4 and 25 °C. The GTCC was selected as an oil phase for CyA microemulsion based on solubility results. The optimum CyA microemulsion formulation consisted of 2.5% CyA, 9% GTCC, 24% Solutol® HS 15, 8% PEG 400, 4% ethanol and 52.5% water based on weight percent. The average particle sizes of the optimized blank and drug-loaded microemulsions were 68.7 nm and 71.6 nm, respectively and remained unchanged upon 25-fold dextrose dilution. The results of microemulsion physical and CyA chemical were confirmed by a three-month stability study at 4 and 25 °C. In vitro haemolysis studies indicated that CyA microemulsions were well tolerated by erythrocytes. The novel microemulsion formulation of CyA was developed that is suitable for parenteral administration. This new formulation could potentially have less vehicle-associated side effects that current commercial formulation of CyA based on Cremophor® EL and ethanol solution. PMID:25761521

  19. Cyclosporin A-induced gingival hyperplasia in psoriasis: review of the literature and case reports.

    PubMed

    V'lckova-Laskoska, Marija T

    2005-01-01

    Cyslosporin A (CyA) treatment of psoriasis is warranted in severe cases where other, conventional antipsoriatic approaches have failed. Gingival hyperplasia is a rare side effect of cyclosporin A treatment in psoriasis patients. Previous studies in cyclosporin A-treated patients (mostly transplant recipients) have demonstrated correlation between cyclosporin A serum levels and oral hygiene status on the one hand, and the prevalence and severity of this overgrowth on the other hand. Severe cases of gingival overgrowth may call for radical treatments such as periodontal surgery. Our aim was to present a severe form of cyclosporin A-induced hyperplasia in two female patients and to give an overview of the current literature on the issue. High serum levels of cyclosporin A were observed in both patients. Moreover, high initial plaque accumulation was noted in both patients. Upon cessation of drug administration and a combined periodontal treatment, virtually complete reduction of the gingival enlargement and inflammation was observed. Consequently, early diagnosis and an all-inclusive treatment of cyclosporin A-induced hyperplasia can result in virtually complete remission of the symptoms and eliminate the need of aggressive treatments such as periodontal surgery.

  20. Early cyclosporin A treatment retards axonal degeneration in an experimental peripheral nerve injection injury model

    PubMed Central

    Erkutlu, Ibrahim; Alptekin, Mehmet; Geyik, Sirma; Geyik, Abidin Murat; Gezgin, Inan; Gök, Abdulvahap

    2015-01-01

    Injury to peripheral nerves during injections of therapeutic agents such as penicillin G potassium is common in developing countries. It has been shown that cyclosporin A, a powerful immunosuppressive agent, can retard Wallerian degeneration after peripheral nerve crush injury. However, few studies are reported on the effects of cyclosporin A on peripheral nerve drug injection injury. This study aimed to assess the time-dependent efficacy of cyclosporine-A as an immunosuppressant therapy in an experimental rat nerve injection injury model established by penicillin G potassium injection. The rats were randomly divided into three groups based on the length of time after nerve injury induced by cyclosporine-A administration (30 minutes, 8 or 24 hours). The compound muscle action potentials were recorded pre-injury, early post-injury (within 1 hour) and 4 weeks after injury and compared statistically. Tissue samples were taken from each animal for histological analysis. Compared to the control group, a significant improvement of the compound muscle action potential amplitude value was observed only when cyclosporine-A was administered within 30 minutes of the injection injury (P < 0.05); at 8 or 24 hours after cyclosporine-A administration, compound muscle action potential amplitude was not changed compared with the control group. Thus, early immunosuppressant drug therapy may be a good alternative neuroprotective therapy option in experimental nerve injection injury induced by penicillin G potassium injection. PMID:25883626

  1. 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}.

  2. Effect of cyclosporin A on the allergen-induced late asthmatic reaction

    PubMed Central

    Sihra, B. S.; Kon, O. M.; Durham, S. R.; Walker, S.; Barnes, N. C.; Kay, A. B.

    1997-01-01

    BACKGROUND: The allergen-induced late asthmatic reaction (LAR) is associated with mucosal inflammation involving several cell types including activated T lymphocytes and eosinophils. In contrast, the early asthmatic reaction (EAR) is considered to results from rapid allergen-induced release of bronchoconstrictor mediators from IgE sensitised mast cells. Cyclosporin A has efficacy in chronic severe corticosteroid-dependent asthma and is believed to act principally by inhibiting cytokine mRNA transcription in T lymphocytes. However, it has effects on other cell types in vitro, including the inhibition of exocytosis/degranulation events in mast cells. It was therefore hypothesised that cyclosporin A would attenuate both the EAR and LAR in subjects with mild asthma. METHODS: Twelve sensitised atopic asthmatic subjects with documented dual asthmatic responses were studied in a double blind, placebo controlled, crossover trial. On two separate study visits subjects received two oral doses of either cyclosporin A or matched placebo before inhaled allergen challenges. The forced expiratory volume in one second (FEV1) was measured half hourly for eight hours and blood eosinophil counts were analysed three, six, and 24 hours after the challenge. Treatment effects on blood eosinophil counts as well as the EAR and LAR, respectively defined as the areas under the curve (AUC) of FEV1 changes from baseline between 0-1 and 4-8 hours after challenge, were compared by non-parametric crossover analysis. RESULTS: Cyclosporin A reduced both the LAR (median AUC -41.9 1.h (interquartile range -82.7 to -12.4) for cyclosporin A and -84.5 1.h (-248.9 to -39.1) for placebo; p = 0.007) and the late increase in blood eosinophils (median 0.2 x 10(9)/1 (0.15 to 0.4) for cyclosporin A and 0.4 x 10(9)/1 (0.25 to 0.55) for placebo; p = 0.024) but had no effect on the EAR. The reduction of the LAR by cyclosporin A correlated significantly with prechallenge blood concentrations of cyclosporin A (r

  3. 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.

  4. Tissue and subcellular localizations of 3H-cyclosporine A in mice.

    PubMed

    Bäckman, L; Brandt, I; Appelkvist, E L; Dallner, G

    1988-02-01

    The tissue and subcellular localizations of 3H-cyclosporine A after administration to mice were determined with whole-body autoradiography and scintillation counting of lipid extracts of tissues and subcellular fractions. The radioactivity was widely distributed in the body and the pattern of distribution after oral or parenteral administration was the same, except that tissue levels were generally lower after oral administration. Pretreatment of the animals with a diet containing cyclosporine A for 30 days before the injection of radioactive cyclosporine A did not change the pattern of distribution substantially. No significant radioactivity was found in the central nervous system, except for the choroidal plexus and the area postrema region of the brain. In pregnant mice no passage of radioactivity from the placentas to fetuses was observed after a single injection. 3H-cyclosporine A and/or its metabolites showed a high affinity for the lympho-myeloid tissues, with a marked long-term retention in bone marrow and lymph nodes. There was massive excretion in the intestinal tract after parenteral administration, and the liver, bile, pancreas and salivary glands contained high levels of radioactivity. In the kidney radioactivity was confined to the outer zone of the outer kidney medulla. In liver homogenates no quantitatively significant binding of 3H-cyclosporine A and/or its metabolites to cellular molecules such as proteins, DNA, phospho- or neutral lipids was found. After lipid extraction with organic solvents, almost all radioactivity was recovered in the organic phase.

  5. Beneficial effects of nilotinib, tyrosine kinase inhibitor on cyclosporine-A induced renal damage in rats.

    PubMed

    Nader, Manar A; Attia, Ghalia M

    2016-04-01

    Nilotinib is a known tyrosine kinase inhibitor that has been approved for treatment of leukemia. The possible protective effect of nilotinib on cyclosporine A-induced nephropathy was investigated in this study and the possible underlying mechanism was explored. Nilotinib (25mg/kg, orally) and cyclosporine A (15 mg/kg/day, subcutaneous) were given to male SD rats for 28 days. Cyclosporine A alone was found to significantly increase serum creatinine, blood urea nitrogen, lactate dehydrogenase, urinary micrototal protein, renal thiobarbituric acid reactive substance, Bax, cytosol cytochrome c release and nuclear factor kappa B activation. Moreover, cyclosporine A significantly reduced serum albumin, creatinine clearance, urinary total antioxidant, superoxide dismutase, glutathione and Bcl2 protein levels. Pathological results showed that in the model group; there was an obvious shrinkage and congestion of the glomeruli and widening of urinary spaces of renal corpuscles, in addition to marked renal tubular injury and fibrosis, while in the group pretreated with nilotinib all measured serum, renal and pathological changes were significantly reduced. This protective effect of nilotinib is linked to the enhanced antioxidant status and reduced inflammation and apoptosis induced by cyclosporine A.

  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. The effect of systemic cyclosporin A on a hairless mouse model of photoaging.

    PubMed

    Moloney, S J; Learn, D B

    1992-10-01

    The mechanisms that cause skin wrinkling in response to chronic exposure to sunlight are unknown. We investigated the possibility that wrinkling of Skh-1 hairless mice is associated with an ultraviolet (UV) radiation-induced immunologic alteration. Exposing Skh-1 hairless mice to a regimen of nonerythemal UV-B (290-320 nm) radiation induced skin wrinkles after 6-7 weeks. Concomitant treatment with cyclosporin A decreased the time to the onset of wrinkles to approximately 4 weeks. Exposing HRS/J hairless mice or athymic nude mice to a similar nonerythemal UV-B radiation regimen for 10 weeks failed to induce skin wrinkles. Concomitant administration of cyclosporin A and UV-B radiation for 7 weeks to HRS/J hairless mice induced no skin wrinkles. Ultraviolet-B or UV-B plus cyclosporin A exposure caused increased immunohistochemical staining for Ia and F4/80 antigens in the upper dermis of tissue from Skh-1 mice, as compared to controls. Treating Skh-1 mice with UV-B radiation plus cyclosporin A was also associated with a large increase in the number of CD3+ cells in the dermis. These staining patterns were absent in similarly treated HRS/J hairless mice. Dermal mast cell numbers in Skh-1 mice were 2-3-fold higher than in HRS/J, athymic nude or NSA mice. Treatment with cyclosporin A increased Skh-1 dermal mast cell numbers approximately 2-fold but had no effect on the dermal mast cell numbers in HRS/J or NSA mice. Based on these findings we postulate that UV-B light and cyclosporin A exacerbate an immunological condition in Skh-1 mice, one consequence of which is manifested as skin wrinkles. Thus, the induction of skin wrinkles in this mouse strain may have no relevance to the wrinkles observed in human skin after chronic exposure to sunlight. PMID:1454879

  8. Timing of cyclosporin-A therapy for abrogation of HVG and GVH responses in rats.

    PubMed

    Markwick, J R; Chambers, J D; Hobbs, J R; Pegrum, G D

    1979-11-17

    Treatment with cyclosporin A was most effective in abrogating popliteal-lymph-node enlargement induced by host-versus-graft and graft-versus-host reactivity in rats when started before injection of donor-strain lymphocytes. Popliteal lymph-node enlargement was never completely abolished, and splenic lymphocytes from recipients treated with cyclosporin A showed no significant reduction in their response to donor-strain lymphocytes in mixed lymphocyte cultures, suggesting that clonal deletion had not taken place. Mixed lymphocyte cultures also indicated that cyclosporin treatment had not reduced the antigenicity of recipient lymphocytes towards donor strain.

  9. Cyclosporine A attenuates 3-nitropropionic acid-induced Huntington-like symptoms in rats: possible nitric oxide mechanism.

    PubMed

    Kumar, Puneet; Kalonia, Harikesh; Kumar, Anil

    2010-01-01

    Cyclosporine A is a well-known immunosuppressant drug that is currently used for prevention of allograft rejection. The current study was conducted to explore the therapeutic potential of cyclosporine A against 3-nitropropionic acid (3-NP)-induced neurotoxicity, an animal model of Huntington disease (HD). Systemic administration of 3-NP (10 mg/kg) for 14 days significantly impaired body weight, motor activity, biochemical parameters (raised lipid peroxidation, nitrite concentration, depletion of superoxide dismutase [SOD] and catalase), and mitochondrial enzymes. Cyclosporine A (2.5, 5, and 10 mg/kg) treatment significantly attenuated behavioral, biochemical, and cellular alterations. Furthermore, L-arginine pretreatment with cyclosporine A (5 mg/kg) significantly reversed the protective effect of cyclosporine A. However, L-nitro-arginine methyl ester (L-NAME; 10 mg/kg) pretreatment potentiated the protective effect of cyclosporine A (5 mg/kg). Study highlights the therapeutic potential of cyclosporine A in the treatment of HP. Study suggests that nitric oxide (NO) modulation is involved in the neuroprotective effect of cyclosporine A against 3-NP neurotoxicity. PMID:20448265

  10. Effect of cyclosporin A particles of varying diameters on gastric cancer cell apoptosis.

    PubMed

    Xing, X L; Lu, Y; Qiu, H L

    2016-01-01

    Human health is significantly threatened by gastric cancer, which is the most common malignant tumor; although drastic, surgery is currently the only way to cure it. However, high recurrence rates and low survival rates are associated with the disease. Therefore, to improve the effectiveness of gastric cancer treatment and to increase the clinical cure rate, we investigated the effect of cyclosporin A particles of varying diameter on gastric cancer cell apoptosis. Flow cytometry was used to detect apoptosis induced by Annexin V-fluorescein isothiocyanate/propidium iodide-double labeling. We also determined the content of reactive oxygen species and the expression level of P-glycoprotein in cells after treatment with cyclosporin A. The results indicated that increases in the concentration and action time of cyclosporin A were associated with statistically significant increases in the apoptosis rate of gastric cancer cells when the experimental and control groups were compared (P < 0.05 and P < 0.01, respectively). In conclusion, during a certain action time and concentration range, cyclosporin A inhibits the proliferation of human gastric cancer cells and can induce their apoptosis. PMID:27173251

  11. CYCLOSPORIN A AFFECTS THE PROLIFERATION PROCESS IN NORMAL HUMAN DERMAL FIBROBLASTS.

    PubMed

    Janikowska, Grazyna; Janikowsk, Tomasz; Pyka, Alina; Wilczok, Adam; Mazurek, Urszula

    2016-01-01

    Cyclosporin A is an immunosuppressant drug that is used not only in solid transplant rejection, but also in moderate and severe forms of psoriasis, pyoderma, lupus or arthritis. Serious side effects of the drug such as skin cancer or gingival hyperplasia probably start with the latent proliferation process. Little is known about the influence of cyclosporin A on molecular signaling in epidermal tissue. Thus, the aim of this study was to estimate the influence of cyclosporin A on the process of proliferation in normal human dermal fibroblasts. Fibroblasts were cultured in a liquid growth medium in standard conditions. Cyclosporin A was added to the culture after the confluence state. Survival and proliferation tests on human dermal fibroblast cells were performed. Total RNA was extracted from fibroblasts, based on which cDNA and cRNA were synthesized. The obtained cRNA was hybridized with the expression microarray HGU-133A_2.0. Statistical analysis of 2734 mRNAs was performed by the use of GeneSpring 13.0 software and only results with p < 0.05 were accepted. Analysis of variance with Tukey post hoc test with Benjamini-Hochberg correction for all three (8, 24, 48 h) culture stages (with and without cyclosporin A) was performed to lower the number of statistically significant results from 679 to 66, and less. Between statistically and biologically significant mRNAs down-regulated were EGRJ, BUBIB, MKI67, CDK1, TTK, E2F8, TPX2, however, the INSIG1, FOSL1, HMOX1 were up-regulated. The experiment data revealed that cyclosporin A up-regulated FOSL1 in the first 24 h, afterwards down-regulating its expression. The HMOX1 gene was up-regulated in the first stage of the experiment (CsA 8 h), however, after the next 16 h of culture time its expression was down-regulated (CsA 24 h), to finally increased in the later time period. The results indicate that cyclosporin A had a significant effect on proliferation in normal human dermal fibroblasts through the changes in the

  12. 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

  13. Cyclosporine a-nanosuspension: formulation, characterization and in vivo comparison with a marketed formulation.

    PubMed

    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 2â8ÂC. 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

  14. 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.

  15. 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

  16. 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.

  17. Cyclosporin A inhibits programmed cell death and cytochrome c release induced by fusicoccin in sycamore cells.

    PubMed

    Contran, N; Cerana, R; Crosti, P; Malerba, M

    2007-01-01

    Programmed cell death plays a vital role in normal plant development, response to environmental stresses, and defense against pathogen attack. Different types of programmed cell death occur in plants and the involvement of mitochondria is still under investigation. In sycamore (Acer pseudoplatanus L.) cultured cells, the phytotoxin fusicoccin induces cell death that shows apoptotic features, including chromatin condensation, DNA fragmentation, and release of cytochrome c from mitochondria. In this work, we show that cyclosporin A, an inhibitor of the permeability transition pore of animal mitochondria, inhibits the cell death, DNA fragmentation, and cytochrome c release induced by fusicoccin. In addition, we show that fusicoccin induces a change in the shape of mitochondria which is not prevented by cyclosporin A. These results suggest that the release of cytochrome c induced by fusicoccin occurs through a cyclosporin A-sensitive system that is similar to the permeability transition pore of animal mitochondria and they make it tempting to speculate that this release may be involved in the phytotoxin-induced programmed cell death of sycamore cells.

  18. Full factorial design, physicochemical characterisation and biological assessment of cyclosporine A loaded cationic nanoparticles.

    PubMed

    Hermans, Kris; Van den Plas, Dave; Everaert, Arnout; Weyenberg, Wim; Ludwig, Annick

    2012-09-01

    Cyclosporine A loaded poly(lactide-co-glycolide) nanoparticles coated with chitosan were prepared using the o/w emulsification solvent evaporation method. A 2(3) full factorial design was used to investigate the effect of 3 preparation parameters on the particle size, polydispersity index, zeta potential and drug release. In vitro experiments were performed in order to evaluate the cytotoxicity and anti-inflammatory activity of the developed nanoparticles. Particle sizes varied from 156 nm to 314 nm, and polydispersity index values of 0.07-0.56 were obtained depending on the different preparation parameters. All nanoparticles showed positive zeta potential values. Nanoparticles prepared with the highest concentration chitosan retained a positive zeta potential after dispersion in simulated lachrymal fluid, which supports the possibility of an electrostatic interaction between these particles and the negatively charged mucus layer at the eye. The in vitro release profile of cyclosporine A from the chitosan-coated nanoparticles was strongly dependent on the release medium used. None of the cationic nanoparticle formulations showed significant cytotoxicity compared to the negative control using human epithelial cells (HaCaT). Cyclosporine A encapsulated in the various nanoparticle formulations remained anti-inflammatory active as significant suppression of interleukine-2 secretion in concanavalin A stimulated Jurkat T cells was observed.

  19. Expressional time phase of leukocyte molecules induced by allogenic cardiac antigen and cyclosporin A in rats’ in vitro model

    PubMed Central

    Xu, Xiu-Fang; Xin, Yi; Zhang, Ying; Huang, Yi-Min; Gu, Yun; Li, Na; Li, Wen-Bin; Zhou, Yu-Jie; Zhang, Zhao-Guang

    2013-01-01

    The immunosuppressive agent cyclosporin A has been proven to reduce the rejection rate and prolong the survival time of transplanted hearts. But some reports showed that cyclosporine A did not completely suppress the rejection. We performed in vitro studies to model a time course to observe the effect of cyclosporin A. Methods: The experiment was divided into a control group (group I), an antigen group (group II), a cyclosporin A group (group III) and an antigen + cyclosporin A group (group IV). After transplantation, at 2 h, 6 h, 12 h, 24 h, 48 h and 72 h, leukocyte molecules were monitored. Results: The expression of IL-2R peaked at 12 h in group II and at 6 h in group III. There was a gradual decline in the expression of the P59 gene in group I, positive expression at 2 h and between 12 h and 24 h in group II, in group IV, there was a decrease at 48 h. The expression of the CD4 gene was lowest at 2 h in group I and at 6 h in group II. CD4 expression then quickly increased to a maximum at 48 h in group III, at 2 h in group IV. There was a minimal expression was reached at 12 h in group I and IV and at 6 h in group III in the expression of the CD8 gene. Conclusions: Alloantigen induced lymphocytes to release IL-2R and P59 and stimulated the induction of the CD4 gene’ transcription for 6 h. Cyclosporin A stimulated the release of IL-2R for 2 h. These results provide an in vitro basis for describing the time phases of rejection inhibited by cyclosporin A. PMID:23724161

  20. Orally co-administrated oleo-gum resin of Commiphora myrrha decreases the bioavailability of cyclosporine A in rats.

    PubMed

    Al-Jenoobi, F I; Alam, M A; Al-Mohizea, A M; Ahad, A; Raish, M

    2015-08-01

    Cyclosporine A is a narrow therapeutic indexed immunosuppressant used after organ transplantation. Several herbs have been reported to alter its pharmacokinetics. Myrrh, dried oleogum resin obtained from Commiphora myrrha (Burseraceae) has been used for many common ailments. The present study was carried out to investigate the effect of myrrh on the pharmacokinetics of cyclosporine A. The rats of the control group received 60 mg/kg, p.o. cyclosporine A, and blood samples were collected at predetermined time intervals. Rats of the test group were treated with an aqueous suspension of myrrh (380 mg/kg p.o.) for eight days and on 8th day a single dose of cyclosporine A was administered to the treated group after 1 h of myrrh administration. Blood samples were drawn at predetermined time points and the drug was analyzed in whole blood by using H-Class UPLC-TQD. Pharmacokinetic profiles of control and test group were compared. Statistically significant differences were observed between the pharmacokinetic parameters of control and treated groups. In the myrrh treated group, the AUC(0-t) and C(max) of cyclosporine A was decreased by about 45% and 48%, respectively. The time to reach maximum concentration (T(max)) remained almost unchanged in both groups. Results indicated that the bioavailability of cyclosporine A was reduced by about 45% when co-administered with myrrh. This observation suggests that concurrent consumption of myrrh and cyclosporine A should be avoided. To confirm the clinical relevance of these findings, P-gp and CYP3A based molecular investigations can be performed along with a well-planned clinical study.

  1. 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. PMID:25775512

  2. A novel calcineurin-independent activity of cyclosporin A in Saccharomyces cerevisiae.

    PubMed

    Singh-Babak, Sheena D; Shekhar, Tanvi; Smith, Andrew M; Giaever, Guri; Nislow, Corey; Cowen, Leah E

    2012-10-01

    Fungi rely on regulatory networks to coordinate sensing of environmental stress with initiation of responses crucial for survival. Antifungal drugs are a specific type of environmental stress with broad clinical relevance. Small molecules with antifungal activity are ubiquitous in the environment, and are produced by a myriad of microbes in competitive natural communities. The echinocandins are fungal fermentation products and the most recently developed class of antifungals, with those in clinical use being semisynthetic derivatives that target the fungal cell wall by inhibiting 1,3-β-D-glucan synthase. Recent studies implicate the protein phosphatase calcineurin as a key regulator of cellular stress responses required for fungal survival of echinocandin-induced cell wall stress. Pharmacological inhibition of calcineurin can be achieved using the natural product and immunosuppressive drug cyclosporin A, which inhibits calcineurin by binding to the immunophilin Cpr1. This drug-protein complex inhibits the interaction between the regulatory and catalytic subunits of calcineurin, an interaction necessary for calcineurin function. Here, we report on potent activity of cyclosporin A when combined with the echinocandin micafungin against the model yeast Saccharomyces cerevisiae that is independent of its known mechanism of action of calcineurin inhibition. This calcineurin-independent synergy does not involve any of the 12 immunophilins known in yeast, individually or in combination, and is not mediated by any of the multidrug transporters encoded or controlled by YOR1, SNQ2, PDR5, PDR10, PDR11, YCF1, PDR15, ADP1, VMR1, NFT1, BPT1, YBT1, YNR070w, YOL075c, AUS1, PDR12, PDR1 and/or PDR3. Genome-wide haploinsufficiency profiling (HIP) and homozygous deletion profiling (HOP) strongly implicate the cell wall biosynthesis and integrity pathways as being central to the calcineurin-independent activity of cyclosporin A. Thus, systems level chemical genomic approaches implicate

  3. Conversion from Tacrolimus to Cyclosporine A Improves Glucose Tolerance in HCV-Positive Renal Transplant Recipients

    PubMed Central

    Handisurya, Ammon; Kerscher, Corinna; Tura, Andrea; Herkner, Harald; Payer, Berit Anna; Mandorfer, Mattias; Werzowa, Johannes; Winnicki, Wolfgang; Reiberger, Thomas; Kautzky-Willer, Alexandra; Pacini, Giovanni; Säemann, Marcus; Schmidt, Alice

    2016-01-01

    Background Calcineurin-inhibitors and hepatitis C virus (HCV) infection increase the risk of post-transplant diabetes mellitus. Chronic HCV infection promotes insulin resistance rather than beta-cell dysfunction. The objective was to elucidate whether a conversion from tacrolimus to cyclosporine A affects fasting and/or dynamic insulin sensitivity, insulin secretion or all in HCV-positive renal transplant recipients. Methods In this prospective, single-center study 10 HCV-positive renal transplant recipients underwent 2h-75g-oral glucose tolerance tests before and three months after the conversion of immunosuppression from tacrolimus to cyclosporine A. Established oral glucose tolerance test-based parameters of fasting and dynamic insulin sensitivity and insulin secretion were calculated. Data are expressed as median (IQR). Results After conversion, both fasting and challenged glucose levels decreased significantly. This was mainly attributable to a significant amelioration of post-prandial dynamic glucose sensitivity as measured by the oral glucose sensitivity-index OGIS [422.17 (370.82–441.92) vs. 468.80 (414.27–488.57) mL/min/m2, p = 0.005), which also resulted in significant improvements of the disposition index (p = 0.017) and adaptation index (p = 0.017) as markers of overall glucose tolerance and beta-cell function. Fasting insulin sensitivity (p = 0.721), insulinogenic index as marker of first-phase insulin secretion [0.064 (0.032–0.106) vs. 0.083 (0.054–0.144) nmol/mmol, p = 0.093) and hepatic insulin extraction (p = 0.646) remained unaltered. No changes of plasma HCV-RNA levels (p = 0.285) or liver stiffness (hepatic fibrosis and necroinflammation, p = 0.463) were observed after the conversion of immunosuppression. Conclusions HCV-positive renal transplant recipients show significantly improved glucose-stimulated insulin sensitivity and overall glucose tolerance after conversion from tacrolimus to cyclosporine A. Considering the HCV

  4. Interaction between castanospermine an immunosuppressant and cyclosporin A in rat cardiac transplantation

    PubMed Central

    Hibberd, Adrian D; Clark, David A; Trevillian, Paul R; Mcelduff, Patrick

    2016-01-01

    AIM: To investigate the interaction between castanospermine and cyclosporin A (CsA) and to provide an explanation for it. METHODS: The alkaloid castanospermine was prepared from the seeds of Castanospermum austral consistently achieving purity. Rat heterotopic cardiac transplantation and mixed lymphocyte reactivity were done using genetically inbred strains of PVG (donor) and DA (recipient). For the mixed lymphocyte reaction stimulator cells were irradiated with 3000 rads using a linear accelerator. Cyclosporin A was administered by gavage and venous blood collected 2 h later (C2). The blood levels of CsA (Neoral) were measured by immunoassay which consisted of a homogeneous enzyme assay (EMIT) on Cobas Mira. Statistical analyses of interactions were done by an accelerated failure time model with Weibull distribution for allograft survival and logistic regression for the mixed lymphocyte reactivity. RESULTS: Castanospermine prolonged transplant survival times as a function of dose even at relatively low doses. Cyclosporin A also prolonged transplant survival times as a function of dose particularly at doses above 2 mg/kg. There were synergistic interactions between castanospermine and CsA in the prolongation of cardiac allograft survival for dose ranges of CsA by castanospermine of (0 to 2) mg/kg by (0 to 200) mg/kg (HR = 0.986; 95%CI: 0.981-0.992; P < 0.001) and (0 to 3) mg/kg by (0 to 100) mg/kg (HR = 0.986; 95%CI: 0.981-0.992; P < 0.001) respectively. The addition of castanospermine did not significantly increase the levels of cyclosporin A on day 3 or day 6 for all doses of CsA. On the contrary, cessation of castanospermine in the presence of CsA at 2 mg/kg significantly increased the CsA level (P = 0.002). Castanospermine inhibited mixed lymphocyte reactivity in a dose dependent manner but without synergistic interaction. CONCLUSION: There is synergistic interaction between castanospermine and CsA in rat cardiac transplantation. Neither the mixed lymphocyte

  5. 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.

  6. 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

  7. Longstanding obliterative panarteritis in Kawasaki disease: lack of cyclosporin A effect.

    PubMed

    Kuijpers, Taco W; Biezeveld, Maarten; Achterhuis, Annemiek; Kuipers, Irene; Lam, Jan; Hack, C E; Becker, Anton E; van der Wal, Allard C

    2003-10-01

    Kawasaki disease is a childhood vasculitis of medium-sized vessels, affecting the coronary arteries in particular. We have treated a therapy-resistant child who met all diagnostic criteria for Kawasaki disease. After the boy was given intravenous immunoglobulins and salicylates, as well as several courses of pulsed methylprednisolone, disease recurred and coronary artery lesions became progressively detectable. Cyclosporin A was started and seemed clinically effective. In contrast to the positive effect on inflammatory parameters, ie, C-reactive protein and white blood cell counts, a novel plasma marker for cytotoxicity (granzyme B) remained elevated. Coronary disease progressed to fatal obstruction and myocardial infarction. Echocardiography, electrocardiograms, and myocardial creatine phosphokinase did not predict impending death. At autopsy an obliterative panarteritis was observed resulting from massive fibrointimal proliferation, affecting the aorta and several large and medium-sized arteries. Immunophenotypic analysis of the inflammatory infiltrates in arteries revealed mainly granzyme-positive cytotoxic T cells and macrophages in the intima and media, as well as nodular aggregates of T cells, B cells, and plasma cells in the adventitia of affected arteries. These findings further endorse the role of specific cellular and humoral immunity in Kawasaki disease. Unremitting coronary arteritis and excessive smooth muscle hyperplasia resulted in coronary occlusion despite the use of cyclosporin A.

  8. Minimization of maintenance immunosuppressive therapy after renal transplantation comparing cyclosporine A/azathioprine or cyclosporine A/mycophenolate mofetil bitherapy to cyclosporine A monotherapy: a 10-year postrandomization follow-up study.

    PubMed

    Thierry, Antoine; Lemeur, Yann; Ecotière, Laure; Abou-Ayache, Ramzi; Etienne, Isabelle; Laurent, Charlotte; Vuiblet, Vincent; Colosio, Charlotte; Bouvier, Nicolas; Aldigier, Jean-Claude; Rerolle, Jean-Philippe; Javaugue, Vincent; Gand, Elise; Bridoux, Frank; Essig, Marie; Hurault de Ligny, Bruno; Touchard, Guy

    2016-01-01

    Long-term outcomes in renal transplant recipients withdrawn from steroid and submitted to further minimization of immunosuppressive regimen after 1 year are lacking. In this multicenter study, 204 low immunological risk kidney transplant recipients were randomized 14.2 ± 3.7 months post-transplantation to receive either cyclosporine A (CsA) + azathioprine (AZA; n = 53), CsA + mycophenolate mofetil (MMF; n = 53), or CsA monotherapy (n = 98). At 3 years postrandomization, the occurrence of biopsy for graft dysfunction was similar in bitherapy and monotherapy groups (21/106 vs. 26/98; P = 0.25). At 10 years postrandomization, patients' survival was 100%, 94.2%, and 95.8% (P = 0.25), and death-censored graft survival was 94.9%, 94.7%, and 95.2% (P = 0.34) in AZA, MMF, and CsA groups, respectively. Mean estimated glomerular filtration rate was 70.4 ± 31.1, 60.1 ± 22.2, and 60.1 ± 19.0 ml/min/1.73 m(2), respectively (P = 0.16). The incidence of biopsy-proven acute rejection was 1.4%/year in the whole cohort. None of the patients developed polyomavirus-associated nephropathy. The main cause of graft loss (n = 12) was chronic antibody-mediated rejection (n = 6). De novo donor-specific antibodies were detected in 13% of AZA-, 21% of MMF-, and 14% of CsA-treated patients (P = 0.29). CsA monotherapy after 1 year is safe and associated with prolonged graft survival in well-selected renal transplant recipient (ClinicalTrials.gov number: 980654).

  9. Safety of cyclosporin A in HCV-infected patients: experience with cyclosporin A in patients affected by rheumatological disorders and concomitant HCV infection.

    PubMed

    Galeazzi, Mauro; Bellisai, Francesca; Giannitti, Chiara; Manganelli, Stefania; Morozzi, Gabriella; Sebastiani, Gian Domenico

    2007-09-01

    Because of the relatively high prevalence of both hepatitis C virus (HCV) infection and autoimmune disorders (ADs), it is not rare to encounter in daily clinical practice patients with ADs also carrying HCV. Corticosteroids and/or immunosuppressant drugs are needed to treat ADs, but they place HCV-infected patients at risk of worsening the infection. So, rheumatologists have often refrained from using corticosteroids or immunosuppressants in AD when HCV-RNA is also present. Cyclosporin A (CsA) is an immunosuppressive agent used to treat a wide range of ADs, but there is a large evidences in the literature, both in vitro and in vivo, suggesting that CsA also exerts an inhibitory effect on HCV replication at standard therapeutic dose. Therefore, this evidence has opened new ways to improve the therapy and the prognosis in patients with HCV-related liver diseases, including those with transplants. Recent reports, although limited in number, also suggest the safety of CsA in the treatment of patients with AD and concomitant HCV infection. In this review we also report our personal experience on the combination treatment with CsA and anti-TNF-alpha agents in rheumatoid arthritis.

  10. Inhibition of rhodamine 123 secretion by cyclosporin A as a model of P-glycoprotein mediated transport in liver.

    PubMed

    Stapf, V; Thalhammer, T; Huber-Huber, R; Felberbauer, F; Gajdzik, L; Graf, J

    1994-01-01

    The interaction between P-glycoprotein modulators and P-glycoprotein mediated transport was investigated using rhodamine 123 in the isolated perfused rat liver of a mutant (TR-) rat strain. TR- rats, deficient in the canalicular multispecific anion transport system, are unable to extrude organic anions (glucuronides) and therefore excrete solely unconjugated rhodamine 123 via P-glycoprotein. Cyclosporin A, a modulator of multidrug resistance in tumor cells, inhibited the biliary secretion of rhodamine 123 dose dependently in a non-competitive manner. Both cyclosporin A and rhodamine inhibited photoaffinity labeling of immunoprecipitated P-glycoprotein with azidopine, indicating binding to hepatic P-glycoprotein. Our results indicate that monitoring the biliary rhodamine 123 secretion in the isolated perfused liver of TR- rats offers a new system for testing modulators of P-glycoprotein like cyclosporin A.

  11. 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

  12. Some optimal culture conditions for production of cyclosporin a by Fusarium roseum

    PubMed Central

    Ismaiel, Ahmed A.; El-Sayed, A.; Mahmoud, Asmaa A.

    2010-01-01

    A suitable chemically defined culture medium was selected and some optimal conditions for the production of the highly immunosuppressive compound, cyclosporin A (Cyc A) are reported. Medium of the following composition was favorable for the production of Cyc A by Fusarium roseum: glucose, 30; NaNO3, 2; KH2PO4, 1; MgSO4.7H2O, 0.5 and KCL, 0.5 (g/l). Maximum productivity of Cyc A was achieved at pH 6.0 when 50 ml of the fermentation medium/250 ml flask, inoculated with five fungal agar discs (6 mm, diameter) of 7-days old F. roseum culture after incubation at 30 ºC at 120 rpm for 7 days. PMID:24031594

  13. Metastatic Listeria monocytogenes infection of the peritoneum in mice with cyclosporine a-induced peritonitis.

    PubMed

    Prats, N; López, S; Domingo, M; Briones, V; Domínguez, L; Marco, A J

    2002-01-01

    Inoculation of mice with Listeria monocytogenes intragastrically or by parenteral routes has not been reported to cause peritonitis. In this study, however, severe listerial peritonitis was induced in mice infected subcutaneously and treated intraperitoneally with cyclosporin A (Cs A) in an oil carrier. In both uninfected and listeria-infected mice, intraperitoneal administration of Cs A consistently produced overexpression of P-selectin in the peritoneal microvasculature and pyogranulomatous inflammation of the peritoneum, suggesting that Cs A causes endothelial damage. We suggest that in listeria-infected mice the non-specific irritant peritonitis induced by the intraperitoneal administration of Cs A results in transfer of listeria-infected phagocytes from the liver and spleen to the peritoneal microvasculature, producing metastatic infection.

  14. Heteropterys tomentosa (A. Juss.) infusion counteracts Cyclosporin a side effects on the ventral prostate

    PubMed Central

    2013-01-01

    Background Cyclosporin A (CsA) is an immunosuppressive drug widely used in treatment of auto-immune diseases or after organ transplants. However, several side effects are commonly associated with CsA long term intake, some regarding to loss of reproductive organ function due to oxidative damage. Considering that phytotherapy is an important tool often used against oxidative stress, we would like to describe the beneficial effects of Heteropterys tomentosa intake to minimize the damage caused by CsA to the ventral prostate tissue of Wistar rats under laboratorial conditions. Methods Thirty adult Wistar rats (Rattus norvegicus albinus) were divided into: control group (water); CsA group (Cyclosporin A); Ht group (H. tomentosa infusion) and CsA + Ht group (CsA and H. tomentosa infusion). Plasmic levels of hepatotoxicity markers, triglycerides, cholesterol and glucose were quantified. The ventral prostate tissue was analyzed under light microscopy, using stereological, morphometrical and immunohistochemical techniques. Results H. tomentosa did not cause any alterations either of the plasmic parameters or of the ventral prostate structure. CsA caused alterations of GOT, total and indirect bilirubin, cholesterol, triglycerides and glucose levels in the plasma; CsA-treated rats showed alterations of the ventral prostate tissue. There were no alterations regarding the plasma levels of GOT, triglycerides and glucose of CsA + Ht animals. The same group also showed normalization of most of the parameters analyzed on the ventral prostate tissue when compared to the CsA group. The treatments did not alter the pattern of AR expression or the apoptotic index of the ventral prostate epithelium. Conclusions The results suggest a protective action of the H. tomentosa infusion against the side effects of CsA on the ventral prostate tissue, which could also be observed with plasmic biochemical parameters. PMID:23406403

  15. Immunosuppressive therapy of cyclosporin A for severe benzene-induced haematopoietic disorders and a 6-month follow-up.

    PubMed

    Song, Yuguo; Du, Xuqin; Hao, Fentong; Gu, Xiaoxin; Zhang, Zhenghua; Zhang, Songquan; Li, Chunsheng; Li, Huiling; Ma, Jing

    2010-06-01

    Long-term exposure to benzene can potentially result in severe haematotoxicities, including pancytopaenia, aplastic anaemia and myelodysplastic syndrome, which are often accompanied by life-threatening symptoms and high mortality. Previous studies demonstrate that benzene-induced haematotoxicities are immune-mediated and that cyclosporin A is a prominent treatment in acquired aplastic anaemia. This study aims to evaluate the potential role of cyclosporin A immunosuppressive therapy for severe benzene-induced haematotoxicity. Between January 2002 and December 2008, 41 patients with severe benzene-induced haematopoietic disorders from five hospitals were enrolled in the study, 22 patients received cyclosporin A, supportive treatments and/or oral testosterone undecanoate, 19 patients were treated with supportive treatments and/or oral testosterone undecanoate as the control group, and a 6-month follow-up was conducted. The results showed that in the cyclosporin A group, 19 of 22 patients (86.36%) had responded to the treatments completely or partially with increased platelets, white blood cells and hemoglobulin counts by the fourth week (P=0.005), the sixth week (P=0.001) and the third month post-treatment (P=0.034), respectively. However, in the control group treated by supportive methods, only 5 of 19 patients (26.32%) responded to the treatments partially (P<0.001). Cyclosporin A in conjunction with supportive treatments may be an effective treatment modality for patients with severe benzene-induced haematopoietic disorders, which in turn implies that these haematotoxicities are immune-mediated. PMID:20381478

  16. Effect of cyclosporin A and trifluoperazine on rat liver mitochondria swelling and lipid peroxidation.

    PubMed

    Nepomuceno, M F; Pereira-da-Silva, L

    1993-10-01

    The effect of cyclosporin A (CsA) or trifluoperazine (TFP) on lipid peroxidation and mitochondrial swelling was determined using liver mitochondria incubated with 30 microM Ca2+ and 250 microM t-butylhydroperoxide or 5 mM inorganic phosphate (P(i)). Lipid peroxidation was not modified by either 1 microM CsA or 40 microM TFP. These compounds presented a distinct effect on mitochondrial permeability. Under oxidative conditions, CsA only showed a transient protective effect whereas TFP completely inhibited mitochondrial swelling. Conversely, CsA was very efficient when Ca2+ and P(i) were used, a condition under which TFP was unable to prevent the swelling. These data are consistent with our previous results (M.F. Nepomuceno, D.V. Macedo and L. Pereira-da-Silva (1991). Brazilian Journal of Medical and Biological Research, 24: 833-836) showing that lipid peroxidation is one among other different components of the permeabilization process. The data suggest that lipid peroxidation is independent of swelling, occurring later than swelling, presumably when the mitochondrial reductant systems are depleted. The differential effects of CsA and TFP suggest that these compounds can be used as specific probes in the elucidation of the two distinct mechanisms responsible for mitochondrial swelling.

  17. Inhibition by cyclosporin A of rodent malaria in vivo and human malaria in vitro.

    PubMed Central

    Nickell, S P; Scheibel, L W; Cole, G A

    1982-01-01

    The development and course of normally lethal parasitemias in mice inoculated intraperitoneally with erythrocytic stages of Plasmodium yoelii or Plasmodium berghei were markedly affected by treatment with the antilymphoid drug cyclosporin A (CS-A). When the first of four daily subcutaneous 25-mg/kg doses of CS-A was given at the time of parasite inoculation, patent infections failed to develop. If begun up to 5 days earlier, this same treatment regimen prolonged the prepatent period, attenuated parasitemia, and reduced mortality. In mice with patient infections, two consecutive daily 25-mg/kg doses of CS-A were sufficient to terminate parasitemias which, after several days, reappeared but were self-limiting. This pattern of apparent cure followed by transient recrudescence remained unaltered even when daily treatment with the same drug dose was continued for 3 weeks. Recrudescence was associated with the emergence of parasite populations that were relatively resistant to CS-A and, in the case of P. yoelii, of reduced virulence. In more limited experiments, CS-A was found to be active in vitro against erythrocytic stages of the human malarial parasite palsmodium falciparum. Depending on the concentration of drug in the culture medium, parasite growth was either prevented or inhibited. PMID:6752020

  18. Effects of cyclosporine A on biomembranes. Vibrational spectroscopic, calorimetric and hemolysis studies.

    PubMed

    O'Leary, T J; Ross, P D; Lieber, M R; Levin, I W

    1986-04-01

    Cyclosporine A (CSA)-dipalmitoylphosphatidylcholine (DPPC) interactions were investigated using scanning calorimetry, infrared spectroscopy, and Raman spectroscopy. CSA reduced both the temperature and the maximum heat capacity of the lipid bilayer gel-to-liquid crystalline phase transition; the relationship between the shift in transition temperature and CSA concentration indicates that the peptide does not partition ideally between DPPC gel and liquid crystalline phases. This nonideality can be accounted for by excluded volume interactions between peptide molecules. CSA exhibited a similar but much more pronounced effect on the pretransition; at concentrations of 1 mol % CSA the amplitude of the pretransition was less than 20% of its value in the pure lipid. Raman spectroscopy confirmed that the effects of CSA on the phase transitions are not accompanied by major structural alterations in either the lipid headgroup or acyl chain regions at temperatures away from the phase changes. Both infrared and Raman spectroscopic results demonstrated that CSA in the lipid bilayer exists largely in a beta-turn conformation, as expected from single crystal x-ray data; the lipid phase transition does not induce structural alterations in CSA. Although the polypeptide significantly affects DPPC model membrane bilayers, CSA neither inhibited hypotonic hemolysis nor caused erythrocyte hemolysis, in contrast to many chemical agents that are believed to act through membrane-mediated pathways. Thus, agents, such as CSA, that perturb phospholipid phase transitions do not necessarily cause functional changes in cell membranes.

  19. 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.

  20. Formulation Strategy for the Delivery of Cyclosporine A: Comparison of Two Polymeric Nanospheres.

    PubMed

    Goyal, Ritu; Macri, Lauren; Kohn, Joachim

    2015-01-01

    A wide range of nanoparticles has been explored for the delivery of highly hydrophobic drugs, but very few publications provide comparative data of the performance of different nanoparticles. To address this need, this publication compares poly(lactic-co-glycolic acid) (PLGA) nanoparticles and nanospheres made from tyrosine-derived tri-block copolymers (termed TyroSpheres) for their respective performance as carriers for cyclosporine A (CSA). Using previously reported data on PLGA, we followed similar experimental protocols to evaluate the in vitro characteristics of TyroSpheres. Although there are some similarities between the two particle systems for the delivery of CSA, such as effective encapsulation and epidermal skin penetration, several differences were notable. First, the methods of preparation were different, i.e., self-assembly and emulsion-diffusion-evaporation process for TyroSpheres and PLGA, respectively. Second, TyroSpheres provided 7-day diffusion-controlled release, whereas PLGA nanoparticles provided >21-day erosion-controlled release. Third, the size of TyroSpheres was measured to be ~60-70 nm irrespective of drug loading, whereas the size of PLGA nanoparticles (~100-250 nm) was dependent on drug loading and the method of preparation. Overall, this publication provides a direct comparison between two different types of nanoparticles and illuminates the respective advantages and disadvantages, using CSA as a model for the release of highly hydrophobic drugs. PMID:26268451

  1. 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.

  2. Lycopene protects against cyclosporine A-induced testicular toxicity in rats.

    PubMed

    Türk, Gaffari; Ateşşahin, Ahmet; Sönmez, Mustafa; Yüce, Abdurrauf; Ceribaşi, Ali Osman

    2007-03-01

    Cyclosporine A (CsA)-induced direct failures in hypothalamic-pituitary-gonadal axis and Sertoli cell phagocytic function have been considered for testicular toxicity so far. It has clearly been reported that oxidative stress leads to damage in sperm functions and structure of the testis. Therefore, this study was conducted to demonstrate whether CsA causes testicular and spermatozoal toxicity associated with the oxidative stress, and to investigate the possible protective effect of lycopene against CsA-induced damages in all reproductive organs and sperm characteristics in male rats. While the daily administration of CsA at the dose 15 mg/kg for 21 days significantly decreased the seminal vesicles weight, epididymal sperm concentration, motility, testicular tissue glutathione (GSH), glutathione peroxidase (GSH-Px) and catalase (CAT), diameter of seminiferous tubules and germinal cell thickness, it increased malondialdehyde (MDA) level and abnormal sperm rates along with degeneration, necrosis, desquamative germ cells in testicular tissue. However, the CsA along with simultaneous administration of lycopene at the dose of 10mg/kg markedly ameliorated the CsA-induced all the negative changes observed in the testicular tissue, sperm parameters and oxidant/antioxidant balance. In conclusion, CsA-induced oxidative stress leads to the structural and functional damages in the testicular tissue and sperm quality of rats and, lycopene has a potential protective effect on these damages.

  3. 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. PMID:20705124

  4. An Overview on Dry Eye Treatment: Approaches for Cyclosporin A Delivery

    PubMed Central

    Yavuz, Burçin; Bozdağ Pehlivan, Sibel; Ünlü, Nurşen

    2012-01-01

    Dry eye syndrome (DES, Keratoconjunctivitis sicca) is a common disorder of the tear film caused by decreased tear production or increased evaporation. Changes in tear composition also promote inflammation on the ocular surface by various mechanisms. Artificial tear drops, tear retention treatment, stimulation of tear secretion, or anti-inflammatory drugs may be used for dry eye treatment according to the severity of the disease. For untreated patients, the risk of ocular infection increases at considerable level and clinical course of the disease may proceed up to infection, corneal ulcer, and blindness. Artificial tears and/or punctual occlusions are used for tear replacement or preservation. New treatment approaches are designed to modify the underlying disease process. For the treatment of severe dry eye disease, cyclosporin A (CsA), the first one of the new generation immunomodulatory drugs, which has an anti-inflammatory effect, is frequently used. CsA has immunosuppressive effects following systemic application. Following local administration of CsA, it is expected to obtain effective drug concentration at the target area and to avoid the various side effects associated with systemic delivery. Microspheres, implants, and liposomes have been developed for administration of CsA subconjunctivally in order to enhance its efficiency. PMID:22619624

  5. Formulation Strategy for the Delivery of Cyclosporine A: Comparison of Two Polymeric Nanospheres

    PubMed Central

    Goyal, Ritu; Macri, Lauren; Kohn, Joachim

    2015-01-01

    A wide range of nanoparticles has been explored for the delivery of highly hydrophobic drugs, but very few publications provide comparative data of the performance of different nanoparticles. To address this need, this publication compares poly(lactic-co-glycolic acid) (PLGA) nanoparticles and nanospheres made from tyrosine-derived tri-block copolymers (termed TyroSpheres) for their respective performance as carriers for cyclosporine A (CSA). Using previously reported data on PLGA, we followed similar experimental protocols to evaluate the in vitro characteristics of TyroSpheres. Although there are some similarities between the two particle systems for the delivery of CSA, such as effective encapsulation and epidermal skin penetration, several differences were notable. First, the methods of preparation were different, i.e., self-assembly and emulsion-diffusion-evaporation process for TyroSpheres and PLGA, respectively. Second, TyroSpheres provided 7-day diffusion-controlled release, whereas PLGA nanoparticles provided >21-day erosion-controlled release. Third, the size of TyroSpheres was measured to be ~60–70 nm irrespective of drug loading, whereas the size of PLGA nanoparticles (~100–250 nm) was dependent on drug loading and the method of preparation. Overall, this publication provides a direct comparison between two different types of nanoparticles and illuminates the respective advantages and disadvantages, using CSA as a model for the release of highly hydrophobic drugs. PMID:26268451

  6. The effects of cyclosporin A and Heteropterys tomentosa on the rat liver.

    PubMed

    de Freitas, Karine M; Almeida, Jacqueline M; Monteiro, Juliana C; Diamante, Maria Aparecida S; Vale, Jéssica S F do; Camargo, Camila; Jorge, Marçal H A; Dolder, Heidi

    2015-03-01

    Cyclosporin A (CsA) is a widely employed immunosuppressive drug that is associated with several side effects, among then hepatotoxicity. Heteropterys tomentosa is a Brazilian plant efficient in reducing damage caused by CsA on the rat testis and prostate. The aim of this study was to evaluate the effect of CsA and H. tomentosa (administered isolated or simultaneously) on the liver of Wistar rats. The animals were treated daily with water (control), CsA (15 mg/kg/day), H. tomentosa infusion or CsA+H. tomentosa, for 21 or 56 days. The treatments did not alter liver morphology or cause fibrosis. H. tomentosa administered for 21 days increased the number of hepatocyte nuclei and Kupffer cell volumetric proportion. After 56 days of treatment, H. tomentosa administration did not alter the parameters analyzed. Biochemical plasma dosages and liver stereology showed impairment caused by CsA-treatment after 21 days; these results were not observed after 56 days of treatment. The simultaneous treatment with CsA and H. tomentosa for 21 or 56 days did not alleviate nor accentuate CsA hepatic effects. The present study showed that the 21 days treatment with CsA caused more alteration to the liver than the 56 days treatment; this could be related to hepatic recovery after the long term treatment.

  7. The effect of nifedipine on renal function in normotensive cyclosporin-A-treated renal allograft recipients.

    PubMed

    McNally, P G; Walls, J; Feehally, J

    1990-01-01

    Intrarenal vasoconstriction is a characteristic feature of CsA nephrotoxicity. The influence of nifedipine, a dihydropyridine calcium channel blocker and potent renal vasodilator, on renal haemodynamics was investigated in 11 cyclosporin A (CsA)- and 9 azathioprine (Aza)-treated normotensive long-term renal allograft recipients. Baseline Cr51-EDTA clearance and effective renal plasma flow (ERPF) were similar in both groups. Nifedipine 20 mg twice daily for 28 days significantly increased Cr51-EDTA clearance (+14.8%) in the CsA group; however, ERPF, renal vascular resistance (RVR), and filtration fraction did not change. Nifedipine did not influence renal haemodynamics in the azathioprine group. The increase in Cr51-EDTA clearance in the CsA group did not correlate with baseline renal function, CsA dose or whole blood levels, donor age, duration of graft, or renal functional reserve capacity. This study suggests that nifedipine confers a beneficial effect on renal haemodynamics in long-term CsA-treated renal allograft recipients and appears to improve renal function by a non-haemodynamic mechanism.

  8. The calcineurin inhibitor cyclosporin A exhibits synergism with antifungals against Candida parapsilosis species complex.

    PubMed

    Cordeiro, Rossana de Aguiar; Macedo, Ramila de Brito; Teixeira, Carlos Eduardo Cordeiro; Marques, Francisca Jakelyne de Farias; Bandeira, Tereza de Jesus Pinheiro Gomes; Moreira, José Luciano Bezerra; Brilhante, Raimunda Sâmia Nogueira; Rocha, Marcos Fábio Gadelha; Sidrim, José Júlio Costa

    2014-07-01

    Candida parapsilosis complex comprises three closely related species, C. parapsilosis sensu stricto, Candida metapsilosis and Candida orthopsilosis. In the last decade, antifungal resistance to azoles and caspofungin among C. parapsilosis sensu lato strains has been considered a matter of concern worldwide. In the present study, we evaluated the synergistic potential of antifungals and the calcineurin inhibitor cyclosporin A (Cys) against planktonic and biofilms of C. parapsilosis complex from clinical sources. Susceptibility assays with amphotericin, fluconazole, voriconazole, caspofungin and Cys were performed by microdilution in accordance with Clinical and Laboratory Standards Institute guidelines. Synergy testing against planktonic cells of C. parapsilosis sensu lato strains was assessed by the chequerboard method. Combinations formed by antifungals with Cys were evaluated against mature biofilms in microtitre plates. No differences in the antifungal susceptibility pattern among species were observed, but C. parapsilosis sensu stricto strains were more susceptible to Cys than C. orthopsilosis and C. metapsilosis. Synergism between antifungals and Cys was observed in C. parapsilosis sensu lato strains. Combinations formed by antifungals and Cys were able to prevent biofilm formation and showed an inhibitory effect against mature biofilms of C. parapsilosis sensu stricto, C. metapsilosis and C. orthopsilosis. These results strengthen the potential of calcineurin inhibition as a promising approach to enhance the efficiency of antifungal drugs. PMID:24722799

  9. Enhanced oral bioavailability of cyclosporine A by liposomes containing a bile salt

    PubMed Central

    Guan, Peipei; Lu, Yi; Qi, Jianping; Niu, Mengmeng; Lian, Ruyue; Hu, Fuqiang; Wu, Wei

    2011-01-01

    The main purpose of this study was to evaluate liposomes containing a bile salt, sodium deoxycholate (SDC), as oral drug delivery systems to enhance the oral bioavailability of the poorly water-soluble and poorly permeable drug, cyclosporine A (CyA). Liposomes composed of soybean phosphatidylcholine (SPC) and SDC were prepared by a thin-film dispersion method followed by homogenization. Several properties of the liposomes including particle size, polydispersity index, and entrapment efficiency were characterized. The in vitro release of CyA from these liposomes was less than 5% at 12 hours as measured by a dynamic dialysis method. The pharmacokinetic results in rats showed improved absorption of CyA in SPC/SDC liposomes, compared with CyA-loaded conventional SPC/cholesterol (Chol) liposomes and microemulsion-based Sandimmune Neoral®. The relative oral bioavailability of CyA-loaded SPC/SDC and SPC/Chol liposomes was 120.3% and 98.6%, respectively, with Sandimmun Neoral as the reference. The enhanced bioavailability of CyA was probably due to facilitated absorption by the liposomes containing SDC rather than improved release rate. PMID:21720508

  10. 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

  11. Treatment of primary HIV-1 infection with cyclosporin A coupled with highly active antiretroviral therapy

    PubMed Central

    Rizzardi, G. Paolo; Harari, Alexandre; Capiluppi, Brunella; Tambussi, Giuseppe; Ellefsen, Kim; Ciuffreda, Donatella; Champagne, Patrick; Bart, Pierre-Alexandre; Chave, Jean-Philippe; Lazzarin, Adriano; Pantaleo, Giuseppe

    2002-01-01

    Primary HIV-1 infection causes extensive immune activation, during which CD4+ T cell activation supports massive HIV-1 production. We tested the safety and the immune-modulating effects of combining cyclosporin A (CsA) treatment with highly active antiretroviral therapy (HAART) during primary HIV-1 infection. Nine adults with primary HIV-1 infection were treated with CsA along with HAART. At week 8, all patients discontinued CsA but maintained HAART. Viral replication was suppressed to a comparable extent in the CsA + HAART cohort and in 29 control patients whose primary infection was treated with HAART alone. CsA restored normal CD4+ T cell levels, both in terms of percentage and absolute numbers. The increase in CD4+ T cells was apparent within a week and persisted throughout the study period. CsA was not detrimental to virus-specific CD8+ or CD4+ T cell responses. At week 48, the proportion of IFN-γ–secreting CD4+ and CD4+CCR7– T cells was significantly higher in the CsA + HAART cohort than in the HAART-alone cohort. In conclusion, rapid shutdown of T cell activation in the early phases of primary HIV-1 infection can have long-term beneficial effects and establish a more favorable immunologic set-point. Appropriate, immune-based therapeutic interventions may represent a valuable complement to HAART for treating HIV infection. PMID:11877476

  12. Effect of periodontal treatment on gingival overgrowth among cyclosporine A-treated renal transplant recipients.

    PubMed

    Pernu, H E; Pernu, L M; Knuuttila, M L

    1993-11-01

    No data exist on any association between combined cyclosporine A (Cy A) and dihydropyridine (DHP) medication and the effect of periodontal treatment on the occurrence of gingival overgrowth (GO) among renal transplant recipients. Clinical data on 27 renal transplant recipients treated with Cy A are presented here, including determinations of serum creatinine, whole blood Cy A concentration, existence of DHP treatment, and periodontal status. GO was classified into four categories according to the clinical changes: score 0 = no GO; score 1 = mild GO; score 2 = moderate GO; and score 3 = severe GO. All participants received hygiene phase periodontal treatment and gingivectomies were performed on 10 who originally had score 2 or 3 GO and pocketing. Fourteen (14) of the recipients had no overgrown gingiva or less than at the initial examination, and none of them had GO score 2 or 3 at the time of re-examination (group A). Thirteen (13) participants had more overgrown gingiva than initially or developed score 2 GO after gingivectomies (group B). Group B included significantly more DHP-medicated recipients than group A (6/13 and 1/14 respectively; P < 0.03). The concomitant administration of Cy A and DHP resulted in a significantly increased percentage of score 2 overgrown gingival units as compared with Cy A alone (P < 0.03). It is concluded that combined treatment with Cy A and DHP is a significant risk factor for progression or recurrence of GO after periodontal treatment among susceptible patients.

  13. The effects of Cyclosporine A and azathioprine on human T cells activated by different costimulatory signals

    PubMed Central

    Leitner, Judith; Drobits, Karin; Pickl, Winfried F.; Majdic, Otto; Zlabinger, Gerhard; Steinberger, Peter

    2011-01-01

    Immunosuppression is an important treatment modality in transplantation and human diseases that are associated with aberrant T cell activation. There are considerable differences regarding the cellular processes targeted by the immunosuppressive drugs that are in clinical use. Drugs like azathioprine (Aza) mainly act by halting proliferation of fast dividing cells, whereas others like cyclosporine A (CsA) specifically target signaling pathways in T cells. Since the outcome of T cell responses critically depends on the quality and strength of costimulatory signals, this study has addressed the interplay between costimulation and the immunosuppressive agents CsA and Aza during the in vitro activation of human T cells. We used an experimental system that allows analyzing T cells activated in the presence of selected costimulatory ligands to study T cells stimulated via CD28, CD2, LFA-1, ICOS or 4-1BB. The mean inhibitory concentrations (IC50) for Aza and CsA were determined for the proliferation of T cells receiving different costimulatory signals as well as for T cells activated in the absence of costimulation. CD28 signals but not costimulation via CD2, 4-1BB, ICOS or LFA-1 greatly increased the IC50 for CsA. By contrast, the inhibitory effects of Aza were not influenced by T cell costimulatory signals. Our results might have implications for combining standard immunosuppressive drugs with CTLA-4Ig fusion proteins, which act by blocking CD28 costimulation. PMID:21756939

  14. Rats taste-aversive learning with cyclosporine a is not affected by contextual changes.

    PubMed

    Tuerkmen, Akin; Bösche, Katharina; Lückemann, Laura; Engler, Harald; Schedlowski, Manfred; Hadamitzky, Martin

    2016-10-01

    In conditioned taste aversion (CTA) rats associate a novel taste (conditioned stimulus; CS) with a treatment (unconditioned stimulus; US) that induces symptoms of malaise. During retrieval, animals learn that the CS no longer predicts the US, with the consequence that the behavior elicited by the CS extinguishes. Importantly, CTA data with lithium chloride (LiCl) as US indicate that extinction learning is affected by changing the physical context. However, if this is also the case in different taste-aversion paradigms employing compounds other than LiCL as US is unknown. Against this background the present study investigated in a CTA paradigm with saccharin as CS and the immunosuppressant cyclosporine A (CsA) as US the influence of contextual changes on CTA extinction. Our results show, that extinction of a learned CS-US association with CsA is not prone to contextual changes. Due to the direct effects of CsA on CNS functioning, CTA with this immunosuppressant apparently operates under different mechanisms compared to other drugs, such as LiCl. These data indicate that taste aversive learning and its extinction are not necessarily specific to the context in which it is learned but also depends, at least in part, on the physiological and neuropharmacological effects of the drug employed as US. PMID:27316343

  15. Influence of cyclosporine A on molecular interactions in lyotropic reverse hexagonal liquid crystals.

    PubMed

    Ben Ishai, Paul; Libster, Dima; Aserin, Abraham; Garti, Nissim; Feldman, Yuri

    2010-10-14

    We present a dielectric study of H(II) mesophases (H(II)) based on a GMO/tricaprylin/phosphatidylcholine/water system seeded with the peptide Cyclosporine A (CSA). The study covers a frequency range 0.01 Hz to 1 MHz and a temperature range of 293 to 319 K, with a 3 K temperature step. Three dielectric relaxation processes are observed and discussed. This picture is further elucidated by comparison with a dielectric study of the empty H(II) mesophase system, previously published, where the same three processes were involved. A complex picture emerges whereby the CSA is intercalated between the surfactant tails yet protrudes into the interface as well. Whereas the CSA remains hydrophobic, it still influences the relaxation behavior of the GMO head and counterion movement along the interface in a nontrivial manner. The third dipolar species, the tricaprylin molecule, is also influenced by the presence of CSA. A critical temperature T(0) = 307 K is recognized and identified as the dehydration temperature of the surfactant heads. This induces a conformal transition in the CSA, drastically changing its effect on the three dielectric processes evident in the raw data. The implications of this behavior are discussed in detail.

  16. Molecular interactions in reverse hexagonal mesophase in the presence of Cyclosporin A.

    PubMed

    Libster, Dima; Ishai, Paul Ben; Aserin, Abraham; Shoham, Gil; Garti, Nissim

    2009-02-01

    The present work investigates the detailed molecular structure of the H(II) mesophase of GMO/tricaprylin/phosphatidylcholine/water system in the presence of hydrophobic model peptide Cyclosporin A (CSA) via ATR-FTIR analysis. The conformation of the peptide in the hexagonal mesophase, as well as its location and specific interactions with the components of the carrier, were studied. Incorporation of phosphatidylcholine to the ternary GMO/tricaprylin/water system caused competition for water binding between the hydroxyl groups of GMO and the phosphate groups of the phosphatidylcholine (PC) leading to dehydration of the GMO hydroxyls in favor of phospholipid hydration. Analysis of CSA solubilization effect on the H(II) mesophase revealed a significant increase in the strength of hydrogen bonding with surfactant hydrogen-bonded carbonyls, indicating interaction of the peptide with the CO groups of the surfactants. The peptide probably caused partial replacement of the intramolecular hydrogen bonds of the mesophase carbonyl groups with intermolecular hydrogen bonds of these carbonyl groups with the peptide. Furthermore, analysis of the Amide I' peak in the FTIR spectra of the peptide demonstrated that two pairs of its internal hydrogen bonds are disrupted when it is incorporated. The partial disruption of the internal hydrogen bonds seems to cause an outward rotation of the peptide amide groups involved, resulting in more efficient intermolecular hydrogen-bonding ability. Apparently, this conformational change increased the hydrophilic properties of CSA, even making it susceptible to a weak interaction with the GMO hydroxyl groups in the interfacial region.

  17. 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.

  18. Inhibition of human immunodeficiency virus and growth of infected T cells by the immunosuppressive drugs cyclosporin A and FK 506.

    PubMed Central

    Karpas, A; Lowdell, M; Jacobson, S K; Hill, F

    1992-01-01

    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 approximately 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. Images PMID:1381509

  19. 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.

  20. In vitro and in vivo characterization on amorphous solid dispersion of cyclosporine A for inhalation therapy.

    PubMed

    Onoue, Satomi; Sato, Hideyuki; Kawabata, Yohei; Mizumoto, Takahiro; Hashimoto, Naofumi; Yamada, Shizuo

    2009-08-19

    Cyclosporine A (CsA) has been clinically used as immunosuppressant, and new application for airway inflammation was also proposed. However, the clinical use of CsA was limited due to severe adverse effects after systemic exposure and the poor solubility. In the present investigation, novel respirable powder (RP) of CsA was developed for pulmonary administration with use of solid dispersion of wet-milled CsA (WM/CsA), and the physicochemical and pharmacological properties of the WM/CsA and its RP formulation were characterized. CsA in the solid dispersion was found to be amorphous by X-ray powder diffraction and differential scanning calorimetry. It exhibited the improved dissolution behavior as compared to active pharmaceutical ingredients. Laser diffraction and cascade impactor analysis of newly developed WM/CsA-RP, consisting of jet-milled WM/CsA and lactose carriers, suggested high dispersion and deposition in the respiratory organs with the emitted dose and the fine particle fraction of 96 and 54%, respectively. Intratracheal administration of WM/CsA-RP (100 microg CsA) in experimental inflammatory rats led to 71 and 85% reduction of granulocyte recruitment in bronchoalveolar lavage fluids and lung tissues, respectively, with showing ca 10(2)-fold reduced AUC and C(max) values of plasma CsA as compared to the oral dosage form of CsA at toxic concentration (10 mg/kg). Upon these findings, WM/CsA-RP would be efficacious dosage form for clinical treatment of airway inflammations with minimal systemic side effects. PMID:19376169

  1. Synergistic inhibition of endothelial cell proliferation, tube formation, and sprouting by cyclosporin A and itraconazole.

    PubMed

    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 IC(50) 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.

  2. Efficacy and safety of topical cyclosporine A 0.05% in vernal keratoconjunctivitis

    PubMed Central

    Yücel, Ozlem Eski; Ulus, Nihal Demir

    2016-01-01

    INTRODUCTION While corticosteroids are an effective choice of treatment for severe vernal keratoconjunctivitis (VKC), their long-term use is restricted due to side effects. This study was conducted to evaluate the efficacy and safety of topical cyclosporine A (CsA) 0.05% in the treatment of VKC. METHODS A total of 30 patients with VKC that was resistant to topical corticosteroids, antihistamines and mast cell stabilisers were treated with topical CsA 0.05%. Patients were evaluated at Weeks 4, 8 and 12 after the initiation of therapy. Symptoms and signs observed before and after treatment were recorded and scores were assigned. Scores for symptoms and signs, the need for topical corticosteroids and ocular side effects were evaluated. RESULTS At baseline, the median values of the symptom and sign scores were 10.0 (range 5.0–18.0) and 6.0 (range 2.0–13.0), respectively. At Week 4 of treatment with topical CsA 0.05%, the median values of the symptom and sign scores were 3.0 (range 0–14.0) and 3.0 (range 0–8.0), respectively. The reductions in the symptom and sign scores were statistically significant. The reduction in the need for corticosteroid was statistically significant by Week 12 of therapy. No significant side effects were reported. CONCLUSION Topical CsA 0.05%, which can help to reduce corticosteroid usage, is an effective and safe alternative for the treatment of resistant VKC. Further studies are needed to determine the optimal duration of therapy and possibility of recurrence. PMID:26768065

  3. Cyclosporin A Preserves Mitochondrial Function after Traumatic Brain Injury in the Immature Rat and Piglet

    PubMed Central

    Kilbaugh, Todd J.; Bhandare, Sunita; Lorom, David H.; Saraswati, Manda; Robertson, Courtney L.

    2011-01-01

    Abstract Cyclosporin A (CsA) has been shown to be neuroprotective in mature animal models of traumatic brain injury (TBI), but its effects on immature animal models of TBI are unknown. In mature animal models, CsA inhibits the opening of the mitochondrial permeability transition pore (MPTP), thereby maintaining mitochondrial homeostasis following injury by inhibiting calcium influx and preserving mitochondrial membrane potential. The aim of the present study was to evaluate CsA's ability to preserve mitochondrial bioenergetic function following TBI (as measured by mitochondrial respiration and cerebral microdialysis), in two immature models (focal and diffuse), and in two different species (rat and piglet). Three groups were studied: injured+CsA, injured+saline vehicle, and uninjured shams. In addition, we evaluated CsA's effects on cerebral hemodynamics as measured by a novel thermal diffusion probe. The results demonstrate that post-injury administration of CsA ameliorates mitochondrial dysfunction, preserves cerebral blood flow (CBF), and limits neuropathology in immature animals 24 h post-TBI. Mitochondria were isolated 24 h after controlled cortical impact (CCI) in rats and rapid non-impact rotational injury (RNR) in piglets, and CsA ameliorated cerebral bioenergetic crisis with preservation of the respiratory control ratio (RCR) to sham levels. Results were more dramatic in RNR piglets than in CCI rats. In piglets, CsA also preserved lactate pyruvate ratios (LPR), as measured by cerebral microdialysis and CBF at sham levels 24 h after injury, in contrast to the significant alterations seen in injured piglets compared to shams (p<0.01). The administration of CsA to piglets following RNR promoted a 42% decrease in injured brain volume (p<0.01). We conclude that CsA exhibits significant neuroprotective activity in immature models of focal and diffuse TBI, and has exciting translational potential as a therapeutic agent for neuroprotection in children. PMID

  4. [Assessment of the peripheral circulation in children with nephrotic syndrome treated with cyclosporin A].

    PubMed

    Czupryniak, Aneta; Kałuzyńska, Anna; Tkaczyk, Marcin; Półtorak-Krawczyk, Anna; Ostrowski, Bartosz; Wiecek, Bogusław; Nowicki, Michał

    2006-01-01

    Cyclosporin A (CsA) is an immunosuppressive agent used in children for the treatment of steroid-dependent idiopathic nephrotic syndrome (INS). Despite its benefitial effect on a course of the disease CsA may exert nephrotoxic effects because of its vasoconstrictive properties. CsA-dependent disorders of the peripheral flow (Raynaud phenomenon--RP) have been recently described. The aim of the study was to assess the effect of CsA on the peripheral circulation. The study group comprised 16 children (12 male, 4 female; mean age 9.8 +/- 4.5 years) treated with CsA for at least 6 months due to INS (mean treatment time 39 +/- 27 months). Thirteen age- and sex-matched individuals served as controls. Peripheral circulation disorders were evaluated by means of a cold stress test (both hands were held in lukewarm water (20 degrees C) for 1 minute and thereafter the changes in the hand temperature were recorded with thermographic camera (Inframetrics SC1000). RP assessment was performed according to the method described by Ammer and Ring. The temperature gradient of 4 degrees C or greater maintained between metacarpal and peripheral areas of a hand after 10 minutes was considered diagnostic for RP. According to these criteria RP was confirmed in only 3 patients from the study group and in 2 controls. However, the time of the temperature increase in the first 5 minutes after cooling was considerably shorter in the children with INS (0.26 +/- 0.26 degrees C/min vs 0.51 +/- 0.29 degrees C/min, p=0.02). No correlation between CsA serum concentration, CsA dose and impairment of the hand temperature increase was found. The study confirmed that in children suffering from INS treated with CsA peripheral blood flow disorders can be seen. It seems that impaired vessel reactivity may result from the vasoconstrictive effect of CsA.

  5. [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.

  6. Topical Cyclosporine A for Treatment of Dry Eye Due to Chronic Mustard Gas Injury

    PubMed Central

    Jadidi, Khosrow; Panahi, Yunes; Ebrahimi, Ali; Mafi, Mostafa; Nejat, Farhad; Sahebkar, Amirhossein

    2014-01-01

    Purpose: To evaluate the efficacy of topical cyclosporine A (tCsA) for treatment of dry eye disease in patients suffering from chronic ocular complications of mustard gas (MG) injury. Methods: This interventional case series included patients with MG injury suffering from severe dry eye despite receiving artificial tears and punctal plugs. Patients were administered tCsA 0.05% twice daily for 3 months. Severity of the condition was evaluated by measuring tear osmolarity, ocular surface disease index (OSDI), tear break-up time (TBUT), and Schirmer's test at baseline and at the end of study. Results: A total of 34 patients with chronic MG injury and mean age of 47.1 ± 6.5 years were studied. Compared to baseline values, tear osmolarity (301.7 ± 11.5 vs. 286.3 ± 7.9 mOsmol/L, P < 0.001) and OSDI (47.5 ± 7.2 vs. 42.7 ± 7.1, P < 0.001) were significantly improved. Likewise, Schirmer's test (4.6 ± 1.3 vs. 5 ± 1.3 mm, P < 0.001) and TBUT (1.9 ± 1.4 vs. 2.7 ± 1.5 s, P < 0.001) also significantly recovered at the end of the study. Conclusion: TCsA 0.05% reduces tear osmolarity and improves dry eye symptoms and can serve as an efficacious treatment for ocular complications in patients with chronic MG injury. PMID:25709764

  7. 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.

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

    PubMed

    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

  9. 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

  10. Mechanism of enhanced vasoconstrictor hormone action in vascular smooth muscle cells by cyclosporin A

    PubMed Central

    Lo Russo, Alexandre; Passaquin, Anne-Catherine; Rüegg, Urs T

    1997-01-01

    The use of the immunosuppressive drug cyclosporin A (CsA) is limited by two major side effects, nephrotoxicity and hypertension, which are caused by drug-induced local vasoconstriction. We have recently shown that CsA potentiates the contraction of isolated resistance arteries to vasoconstrictor hormones and increases the calcium response to these agents in vascular smooth muscle cells (VSMC). The goal of the present study was to investigate further the molecular mechanism(s) involved in these effects. Stimulation of VSMC with [Arg]8vasopressin (AVP) induced a concentration-dependent increase in total inositol phosphates (InsP) and cellular calcium response (as measured by 45Ca2+ efflux). Preincubation of VSMC with CsA increased both InsP formation and 45Ca2+ efflux. The potentiating effect of CsA on AVP-elicited InsP formation and 45Ca2+ efflux was inhibited by co-incubation with the protein synthesis inhibitors actinomycin D and cycloheximide, indicating that CsA acted on gene expression. Binding experiments with [3H]-AVP on VSMC showed that CsA increased the number of AVP receptors by about two fold without affecting receptor affinity. Actinomycin D completely blocked this increase. These results demonstrate for the first time that incubation of VSMC with CsA increases the expression of AVP receptors, resulting in a potentiation of InsP formation and calcium response upon stimulation with AVP. This effect of CsA is likely to occur with other vasoconstrictor hormone receptors as well and could be a key mechanism in the induction of vasoconstriction, and subsequent drug-induced nephrotoxicity and hypertension. PMID:9154334

  11. The effect of nifedipine on graft function in renal allograft recipients treated with cyclosporin A.

    PubMed

    Propper, D J; Whiting, P H; Power, D A; Edward, N; Catto, G R

    1989-08-01

    The effect of the calcium channel antagonist nifedipine on renal allograft function was assessed in two groups of renal transplant recipients at least one year after transplantation. Group 1 comprised 10 patients receiving low-dose prednisolone and cyclosporin A, and Group 2 comprised 9 patients receiving low-dose prednisolone and azathioprine. Before commencing nifedipine, creatinine and sodium clearance rates and the fractional excretion of sodium were similar in both two groups. Lithium clearance rates and the fractional excretion of lithium were, however, significantly lower (p less than 0.01) in Group 1 than in Group 2. The absolute reabsorption of sodium from the distal nephron (p less than 0.01), the absolute reabsorption of water from the distal nephron segment (p less than 0.01) and the fractional reabsorption of sodium from the distal tubule relative to the delivery of sodium from the proximal tubule (p less than 0.05) were also lower in Group 1. After seven days of nifedipine treatment (10 mg/8 h) there was a significant fall in sodium clearance (p less than 0.01) and fractional sodium excretion (p less than 0.05), and an increase in the fractional distal reabsorption of sodium relative to the delivery of sodium from the proximal tubule (p less than 0.01), and the fractional distal reabsorption of water relative to the delivery of water from the proximal tubule (p less than 0.02), in Group 1 but not Group 2. The only alterations observed in Group 2 were an increase in fractional lithium excretion (p less than 0.05), and a significant fall in the absolute proximal tubular reabsorption of iso-osmotic fluids (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

  12. Gingival hyperplasia in renal allograft recipients receiving cyclosporin-A and calcium antagonists.

    PubMed

    King, G N; Fullinfaw, R; Higgins, T J; Walker, R G; Francis, D M; Wiesenfeld, D

    1993-04-01

    Although it is established that the immunosuppressant cyclosporin-A (CsA) and calcium antagonists [Nifedipine (Nif) and Diltiazem (Dz)] can independently induce gingival enlargement, little has been documented on the significance of the salivary CsA levels and the combined effect of CsA and a calcium antagonist upon gingival tissues. In the present cross-sectional investigation, clinical periodontal parameters and the pharmacologic profiles of CsA, Nif, and Dz were determined for 66 renal transplant recipients. Subjects were divided into the following groups: Group (Gp) 1: CsA [n = 18]; Gp 2: CsA + Nif [n = 15]; Gp 3: CsA + Dz [n = 12] and a negative Control Gp 4: azathioprine [n = 21]. A gingival enlargement score was assessed for each patient from study models using a hyperplastic index (HI). Pharmacologic profiles included CsA whole blood and whole saliva levels as measured by fluorescence polarization immunoassay. The HI scores between Gp 1, 2 and 3 were not significantly different. However, when compared with controls (Gp 4), there was a significant difference in HI and all individual groups (Gp 1, 2, 3) (p < 0.05). Gingival hyperplasia was only weakly related to plaque and calculus but was unrelated to CsA dose (mg/kg/day), duration of CsA therapy (months), CsA blood or saliva levels (ng/ml), or the concurrent administration of a Nif or Dz. Gingival enlargement was found to occur in 49% of subjects who were either on CsA or CsA and a calcium antagonist. It is concluded that CsA alone or in combination with a calcium antagonist caused a significant increase in gingival enlargement compared with controls.(ABSTRACT TRUNCATED AT 250 WORDS)

  13. 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

  14. Cyclosporin A treatment in children with minimal change nephrotic syndrome and focal segmental glomerulosclerosis.

    PubMed

    Brodehl, J; Brandis, M; Helmchen, U; Hoyer, P F; Burghard, R; Ehrich, J H; Zimmerhackl, R B; Klein, W; Wonigeit, K

    1988-11-15

    In a pilot study 23 children with nephrotic syndrome were treated with cyclosporin A (Cs) for 6-45 months. 8 children suffered from steroid dependent minimal change nephrotic syndrome (MCNS) and had experienced at least one course with cytotoxic drugs, but had relapsed thereafter. 2 children had diabetes mellitus type I with nephrotic syndrome and 13 children had steroid resistant focal segmental glomerulosclerosis (FSGS). Cs was started with 100 mg/m2/day in two doses and increased stepwise to obtain a Cs whole blood trough level of 200-400 ng/ml. In steroid dependent MCNS treatment with Cs reduced relapse rate significantly, and prednisone therapy could be stopped completely. After discontinuation of Cs, relapses reoccurred as frequently as before. Renal function remained unimpaired despite repeated Cs treatment courses up to 38 months. In cases of nephrotic syndrome with diabetes type I Cs treatment led to complete remission without changing the insulin requirement. However, after discontinuation of Cs relapses reoccurred. In steroid resistant FSGS 6 children benefited from Cs treatment: 4 went into complete remission, 2 into partial remission. The 2 children with complete remission relapsed but remained Cs responsive. The remaining 7 children with FSGS did not respond to Cs but continued the course of their disease, with two patients rapidly progressing to terminal renal failure. Side-effects of Cs treatment were mild. It is concluded that Cs is an effective agent in steroid dependent MCNS and can be used as an alternative drug in specific cases like steroid toxicity or diabetes mellitus. In steroid resistant FSGS a trial with Cs seems to be warranted since some cases do respond favorably. To avoid nephrotoxicity treatment with Cs should always be monitored closely by determination of blood levels and renal function.

  15. 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. PMID:27085046

  16. Green tea extract attenuates cyclosporine A-induced oxidative stress in rats.

    PubMed

    Mohamadin, A M; El-Beshbishy, H A; El-Mahdy, M A

    2005-01-01

    Cyclosporine A (CsA) nephrotoxicity underweighs the therapeutic benefits of such a powerful immunosuppressant. Whether oxidative stress plays a role in such toxicity is not well delineated. We investigated the potential of green tea extract (GTE) to attenuate CsA-induced renal dysfunction in rats. Three main groups of Sprague-Dawley rats were used: CsA, GTE, and GTE plus CsA-receiving animals. Corresponding control groups were also used. CsA was administered in a dose of 20mg kg(-1) day(-1), i.p., for 21 days. In the GTE/CsA groups, the rats received different concentrations of GTE (0.5, 1.0 and 1.5%), as their sole source of drinking water, 4 days before and 21 days concurrently with CsA. The GTE group was treated with 1.5% concentration of GTE only for 25 days. A concomitant administration of GTE, to CsA receiving rats, markedly prevented the generation of thiobarbituric acid-reacting substances (TBARS) and significantly attenuated CsA-induced renal dysfunction as assessed by estimating serum creatinine, blood urea nitrogen, uric acid and urinary excretion of glucose. A considerable improvement in terms of reduced glutathione content and activity of antioxidant enzymes in the kidney homogenate of the GTE/CsA-receiving rats was observed. The activity of lysosomal enzymes, N-acetyl-beta-glucosaminidase, beta-glucuronidase and acid phosphatase was significantly inhibited following GTE co-administration. Our data prove the role of oxidative stress in the pathogenesis of CsA-induced kidney dysfunction. Supplementation of GTE could be useful in reducing CsA nephrotoxicity in rats. However, clinical studies are warranted to investigate such an effect in human subjects.

  17. 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

  18. PEGylated single-walled carbon nanotubes as nanocarriers for cyclosporin A delivery.

    PubMed

    Hadidi, Naghmeh; Kobarfard, Farzad; Nafissi-Varcheh, Nastaran; Aboofazeli, Reza

    2013-06-01

    Single-walled carbon nanotubes (SWCNTs) have attracted the attention of many researchers due to their remarkable physicochemical features and have been found to be a new family of nanovectors for the delivery of therapeutic molecules. The ability of these nanostructures to load large amounts of drug molecules on their outer surface has been considered as the main advantage by many investigators. Here, we report the development of a PEGylated SWCNT-mediated delivery system for cyclosporin A (CsA) as a potent immunosuppressive agent. The available OH group in the CsA structure was first linked to a bi-functional linker (i.e., succinic anhydride) in order to provide a COOH terminal group. CsA succinylation process was optimized by using the modified simplex method. The resulting compound, CsA-CO-(CH(2))(2)-COOH, was then grafted onto the exterior surface of SWCNTs, previously PEGylated with phospholipid-PEG(5000)-NH(2) conjugates, through the formation of an amide bond with the free amine group of PEGylated SWCNTs. Drug loading, stability of the PEGylated SWCNT-CsA complex, and in vitro release of the drug were evaluated. Loading efficiencies of almost 72% and 68% were achieved by UV spectrophotometry and elemental analysis methods, respectively. It was observed that 57.3% of cyclosporine was released from CsA-Pl-PEG(5000)-SWCNTs after 3 days. In this investigation, we conjugated CsA to an amine-terminated phospholipid-polyethylene glycol chain attached on SWCNTs via a cleavable ester bond and demonstrated the possible potential of PEGylated SWCNT-based systems for CsA delivery.

  19. 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.

  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. Effectiveness and Optical Quality of Topical 3.0% Diquafosol versus 0.05% Cyclosporine A in Dry Eye Patients following Cataract Surgery

    PubMed Central

    Lee, Jang Hoon; Song, In Seok; Kim, Kyoung Lae; Yoon, Sam Young

    2016-01-01

    Purpose. To evaluate the effectiveness and optical quality of 3.0% topical diquafosol versus 0.05% cyclosporine A in dry eye patients following cataract surgery. Methods. In total, 40 eyes of 40 patients newly diagnosed with dry eye syndrome 1 week after cataract surgery were randomized to receive either 3.0% diquafosol ophthalmic solution six times daily or 0.05% cyclosporine A twice daily for 3 months. Outcome measures were tear film break-up time (TBUT), results on Schirmer 1 test, ocular surface staining score, the ocular surface disease index (OSDI) score, and higher-order aberrations (HOAs). Measurements were taken at baseline and at 1, 2, and 3 months. Results. In the diquafosol group, TBUT showed higher outcomes than the cyclosporine A group at 1 and 3 months. Both groups showed increased scores on Schirmer 1 test. The ocular surface staining score decreased in all periods in both groups. Vertical coma and total HOAs decreased more in the cyclosporine A group than in the diquafosol group at 3 months. Conclusion. Both 3.0% diquafosol and 0.05% cyclosporine A were effective in treating dry eye after cataract surgery. Diquafosol was more effective in increasing the tear secretion, but cyclosporine A was more effective in improving optical aberrations. PMID:26989503

  2. Endothelium-dependent relaxation in the isolated rat kidney: impairment by cyclosporine A.

    PubMed

    Stephan, D; Billing, A; Krieger, J P; Grima, M; Fabre, M; Hofner, M; Imbs, J L; Barthelmebs, M

    1995-12-01

    The therapeutical use of cyclosporine A (CsA) is hampered by the development of nephrotoxicity characterized by a marked increase in renal vascular resistance (RVR). We investigated vascular functions in kidneys of rats treated with CsA. The ex vivo vascular reactivity of kidneys from control rats and animals treated subacutely with CsA [50 mg/kg/day subcutaneously (s.c.) for 16-21 days] or an olive oil vehicle (1 ml/kg) was analyzed in male Wistar rats. The right kidney was isolated and perfused with Tyrode's or Krebs solution in an open circuit. The effects of acetylcholine (Ach), fenoldopam (FEN), and sodium nitroprusside (SNP) on norepinephrine (NE) preconstricted kidneys were studied. In control kidneys (untreated or vehicle-treated), Ach induced a relaxation (EC50 = 0.56 +/- 0.05 x 10(-9)M; Emax = 88.2 +/- 2.1% decrease in the vascular tone restored by NE) which was endothelium-dependent [near-complete abolition after treatment with a detergent, 3-[(3-cholamidopropyl)-dimethyl-ammonio]-1-propane-sulfonate (CHAPS) treatment] but only partially inhibited by indomethacin (EC50 = 1.71 +/- 0.39 x 10(-9)M, p < 0.05; Emax = 87.1 +/- 4.9%, NS) or indomethacin with NG-nitro-L-arginine methyl ester (L-NAME: EC50 = 1.04 +/- 0.38 x 10(-9)M, NS; Emax = 63.8 +/- 2.5%, p < 0.01). CsA treatment induced a marked decrease in creatinine clearance and natriuresis measured in vivo but had no effect on systolic blood pressure (SBP). In CsA-treated rats, Ach-induced renal relaxation was partially blunted (EC50 = 1.88 +/- 0.34 x 10(-9)M, p < 0.01; Emax = 82.8 +/- 4.6, NS), with both a defect in prostaglandin (PG) and nitric oxide (NO)-related responses. CsA treatment had no effect on endothelium-independent relaxations induced by FEN and SNP. These results show that subacute CsA treatment selectively impairs renal endothelium-dependent relaxation related to PGs and NO release.

  3. Cyclosporin A affects the bioavailability of ginkgolic acids via inhibition of P-gp and BCRP.

    PubMed

    Li, Li; Yao, Qing-Qing; Xu, Si-Yun; Hu, Hai-Hong; Shen, Qi; Tian, Ye; Pan, Lan-Ying; Zhou, Hui; Jiang, Hui-di; Lu, Chuang; Yu, Lu-Shan; Zeng, Su

    2014-11-01

    Ginkgolic acids (GAs) in natural product Ginkgobiloba L. are the pharmacological active but also toxic components. Two compounds, GA (C15:1) and GA (C17:1) are the most abundant GAs. In this study, several in vitro and in vivo models were applied to investigate transport mechanism of GAs. A rapid and sensitive LC-MS/MS method for the simultaneous determination of GA (C15:1) and GA (C17:1) was applied to analyze the biological specimens. The Papp(AP→BL) values of GA (C15:1) and GA (C17:1) were 1.66-2.13×10(-)(6)cm/s and 1.34-1.85×10(-)(6)cm/s determined using MDCK and MDCK-MDR1 cell monolayers, respectively. The Papp(BL→AP) were remarkably greater in the MDCK-MDR1 cell line, which were 6.77-11.2×10(-)(6)cm/s for GA (C15:1) and 4.73-5.15×10(-)(6)cm/s for GA (C17:1). Similar results were obtained in LLC-PK1 and LLC-PK1-BCRP cell monolayers. The net efflux ratio of GA (C15:1) and GA (C17:1) in both cell models was greater than 2 and markedly reduced by the presence of Cyclosporin A (CsA) or GF120918, inhibitors of P-gp and BCRP, suggesting that GAs are P-gp and BCRP substrates. The results from a rat bioavailability study also showed that co-administrating CsA intravenously (20mg/kg) could significantly increase GA (C15:1) and GA (C17:1) AUC0-t by 1.46-fold and 1.53-fold and brain concentration levels of 1.43-fold and 1.51-fold, respectively, due to the inhibition of P-gp and BCRP efflux transporters by CsA.

  4. 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

  5. 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

  6. Effect of cyclosporin A on the survival and ultrastructure of Echinococcus granulosus protoscoleces in vitro.

    PubMed

    Colebrook, A L; Jenkins, D J; Jones, M K; Tatarczuch, L; Lightowlers, M W

    2004-10-01

    Surgical treatment of human hydatidsosis involves the use of various scolicidal agents to kill infective Echinococcus granulosus protoscoleces that may disseminate into the peritoneal cavity during surgery and potentially re-infect the patient. Currently, no scolicidal agent is completely effective in killing intracystic protoscoleces in humans. Cyclosporin A (CsA) has previously been found to be lethal for E. granulosus protoscoleces in vitro. In this study, we further assessed the effectiveness of CsA as a scolicidal agent by testing the toxic effect of CsA at higher doses over various time-periods. Experiments were performed on activated and unactivated protoscoleces cultured in nutrient medium or sheep hydatid cyst fluid. All activated protoscoleces were killed following culture in 100 microg/ml of CsA for 3 days and 50 or 20 microg/ml for 5 days. The lethal effect of CsA on unactivated protoscoleces varied but reached 100% over 15 days in culture with 100 or 50 microg/ml of CsA. Pulse treatment of protoscoleces with 50, 20 or 10 microg/ml of CsA for 5 min or 72 h killed all parasites by day 10 and day 5 respectively. Untreated protoscoleces remained greater than 95 % viable for the duration of experiments. Changes in protoscolex ultrastructure induced by treatment with 10 microg/ml of CsA over 10 days in in vitro culture was assessed by TEM. Protoscolex alterations observed in treated parasites included an increase in cellular vacuolization, swelling of mitochondria, rounding of cells, damage to the tegument, decrease in glycogen, a breakdown of the extracellular matrix and an increase in lipid globules. The untreated protoscoleces, by comparison, had few changes during the 10-day culture period with the exception of large amounts of extracellular glycogen observed in the protoscoleces at culture days 7 and 10. From these results, CsA is clearly an effective scolicidal agent in vitro that may have potential application as a new therapeutic agent in the

  7. Conversion to generic cyclosporine A in stable chronic patients after heart transplantation

    PubMed Central

    Kraeuter, Maximilian; Helmschrott, Matthias; Erbel, Christian; Gleissner, Christian A; Frankenstein, Lutz; Schmack, Bastian; Ruhparwar, Arjang; Ehlermann, Philipp; Katus, Hugo A; Doesch, Andreas O

    2013-01-01

    Background Cyclosporine A (CSA) is a narrow therapeutic index drug. Available CSA products differ in the constitution of their emulsion. To compare intra-individual differences after a conversion to a generic CSA, a retrospective single-center study was initiated. Methods Twenty adult stable chronic (>24 months post heart transplant) recipients were included in the present retrospective study. These patients were previously switched from Sandimmune Neoral® to the generic CSA (Equoral®) according to the patients’ preference during the clinical routine. Dose-normalized trough levels (DNL) and trough levels (C0) at 8 months, 4 months, and 2 weeks before the switch were retrospectively compared with the corresponding values at 2 weeks, 4 months, and 8 months after the switch to the generic CSA. Additionally, changes in the routine laboratory parameters, the number of treated rejection episodes, and the adherence to the CSA target levels were compared. Results The mean DNL (adapted to the daily CSA dose in mg) was 0.71±0.26 (ng/mL)/mg on Neoral therapy; on Equoral it was 0.68±0.23 (ng/mL)/mg, (P=0.38). In comparison to the CSA daily dose prior to the conversion, at postconversion, no significant changes of CSA daily dose were observed (Neoral 140.67±39.81 mg versus Equoral 134.58±41.61 mg; P=0.13). No rejection episodes requiring therapy occurred prior to or postconversion (P=0.99). Additionally, no statistically significant changes of routine laboratory parameters regarding the Modification of Diet in Renal Disease or hematological parameters were seen (all P=not significant). No adverse events after the conversion were observed. Conclusion This study in chronic and stable HTx patients demonstrated no statistically significant differences in the CSA DNL after a conversion to generic CSA (Equoral). The generic CSA was generally well-tolerated. We concluded that a conversion from Neoral to Equoral is safe and clinically feasible in this distinct patient

  8. 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

  9. Cyclosporine A and MnTMPyP Alleviate α-Synuclein Expression and Aggregation in Cypermethrin-Induced Parkinsonism.

    PubMed

    Agrawal, Sonal; Dixit, Anubhuti; Singh, Ashish; Tripathi, Pratibha; Singh, Dhirendra; Patel, Devendra Kumar; Singh, Mahendra Pratap

    2015-12-01

    Cypermethrin induces the mitochondrial dysfunction and oxidative damage to the nigrostriatal dopaminergic neurons leading to Parkinsonism in rats. Despite α-synuclein aggregation is reported to be critical in Parkinson's disease, its role and alliance with the mitochondrial dysfunction and oxidative damage leading to cypermethrin-induced Parkinsonism have not yet been deciphered. The present study aimed to examine the effect of cypermethrin on the expression and aggregation of α-synuclein and its subsequent connection with oxidative damage and mitochondrial dysfunction leading to the nigrostriatal dopaminergic neurodegeneration in the presence or absence of a mitochondrial membrane transition pore opening inhibitor, cyclosporine A and a superoxide dismutase/catalase mimetic, manganese (III) tetrakis (1-methyl-4-pyridyl) porphyrin pentachloride (MnTMPyP). The expression of α-synuclein, 3-nitrotyrosine (3-NT), 4-hydroxynonenal (4-HNE)-modified proteins, mitochondrial dysfunction-dependent apoptotic proteins, nitrite content, lipid peroxidation (LPO) and number of tyrosine hydroxylase (TH)-positive neurons were estimated in the substantia nigra and dopamine content in the striatum of control and treated rats employing standard procedures. Cypermethrin augmented the expression of α-synuclein, 3-NT, 4-HNE-modified proteins, caspase-3, mitochondrial Bax and cytosolic cytochrome-c along with nitrite and LPO and reduced the expression of cytosolic Bax, mitochondrial cytochrome-c, dopamine and number of TH-positive neurons. Cyclosporine A or MnTMPyP alleviated the expression and aggregation of α-synuclein along with indicators of the mitochondrial dysfunction, oxidative damage and dopaminergic neurodegeneration. The results demonstrate that cypermethrin induces α-synuclein expression and aggregation while cyclosporine A or MnTMPyP rescues from α-synuclein over-expression and aggregation along with the mitochondrial dysfunction and oxidative damage leading to

  10. Integrative cross-omics analysis in primary mouse hepatocytes unravels mechanisms of cyclosporin A-induced hepatotoxicity.

    PubMed

    Van den Hof, Wim F P M; Van Summeren, Anke; Lommen, Arjen; Coonen, Maarten L J; Brauers, Karen; van Herwijnen, Marcel; Wodzig, Will K W H; Kleinjans, Jos C S

    2014-10-01

    The liver is responsible for drug metabolism and drug-induced hepatotoxicity is the most frequent reason for drug withdrawal, indicating that better pre-clinical toxicity tests are needed. In order to bypass animal models for toxicity screening, we exposed primary mouse hepatocytes for exploring the prototypical hepatotoxicant cyclosporin A. To elucidate the mechanisms underlying cyclosporin A-induced hepatotoxicity, we analyzed expression levels of proteins, mRNAs, microRNAs and metabolites. Integrative analysis of transcriptomics and proteomics showed that protein disulfide isomerase family A, member 4 was up-regulated on both the protein level and mRNA level. This protein is involved in protein folding and secretion in the endoplasmic reticulum. Furthermore, the microRNA mmu-miR-182-5p which is predicted to interact with the mRNA of this protein, was also differentially expressed, further emphasizing endoplasmic reticulum stress as important event in drug-induced toxicity. To further investigate the interaction between the significantly expressed proteins, a network was created including genes and microRNAs known to interact with these proteins and this network was used to visualize the experimental data. In total 6 clusters could be distinguished which appeared to be involved in several toxicity related processes, including alteration of protein folding and secretion in the endoplasmic reticulum. Metabonomic analyses resulted in 5 differentially expressed metabolites, indicative of an altered glucose, lipid and cholesterol homeostasis which can be related to cholestasis. Single and integrative analyses of transcriptomics, proteomics and metabonomics reveal mechanisms underlying cyclosporin A-induced cholestasis demonstrating that endoplasmic reticulum stress and the unfolded protein response are important processes in drug-induced liver toxicity.

  11. 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.

  12. Proliferative and inductive effects of Cyclosporine a on gingival fibroblast of child and adult

    PubMed Central

    Salman, Bahareh Nazemi; Vahabi, Surena; Movaghar, Sepideh Ebrahimi; Mahjour, Faranak

    2013-01-01

    Background: Gingival overgrowth is a serious side-effect that accompanies the use of Cyclosporin A (CsA). Up to 97% of the transplant recipient children, who were submitted to CsA therapy, have been reported to suffer from this side-effect. Several conflicting theories have been proposed to explain the fibroblast's function in CsA-induced gingival overgrowth. The aim of this study is to assess the proliferation of gingival fibroblasts and levels of released cytokines after being exposed to CsA, in both adults and pediatric groups, and to make a comparison between the results of the two groups. Materials and Methods: The adult fibroblast samples were derived from four healthy adults, aged 35 to 42 years and pediatric samples were obtained from four healthy children, age between four and eleven years. Tissue samples were plated in Dulbecco's Modified Eagle Medium (DMEM) containing 10% fetal bovine serum (FBS), Streptomycin and Penicillin. The samples were cultured in 25 cm2 plates containing 5% CO2, and incubated at 37°C. The cells used for all the experiments were at the fourth passage. The concentration of PGE2, IL-1β, IL-6, IL-8, TNF-α, and TGF-β1 was determined by the enzyme-linked immunosorbent assay (ELISA) and the proliferation rate was assessed by the MTT assay. Alpha error levels were set as 0.05. Results: CsA stimulated significantly higher levels of IL-6, IL-8 and TGF-β1 in adult gingival fibroblasts than it did in the control group; whereas, the expression of IL-1β and PGE2 in the fibroblasts exposed to CsA was significantly weaker (P < 0.05). The fibroblasts in the two groups did not reveal any noticeable difference in the production of TNF-α. Furthermore, cell proliferation in the CsA group was not significantly higher than that in the control group. No significant differences in cytokines TNF-α and IL-1β were noted between the two groups. The results indicated that CsA stimulated cell proliferation in the pediatric fibroblast cell line

  13. 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

  14. Screening for Drug-Induced Hepatotoxicity in Primary Mouse Hepatocytes Using Acetaminophen, Amiodarone, and Cyclosporin A as Model Compounds: An Omics-Guided Approach

    PubMed Central

    Van Summeren, Anke; Renes, Johan; Lizarraga, Daneida; Bouwman, Freek G.; Noben, Jean-Paul; van Delft, Joost H. M.; Kleinjans, Jos C. S.

    2013-01-01

    Abstract Drug-induced hepatotoxicity is a leading cause of attrition for candidate pharmaceuticals in development. New preclinical screening methods are crucial to predict drug toxicity prior to human studies. Of all in vitro hepatotoxicity models, primary human hepatocytes are considered as ‘the gold standard.’ However, their use is hindered by limited availability and inter-individual variation. These barriers may be overcome by using primary mouse hepatocytes. We used differential in gel electrophoresis (DIGE) to study large-scale protein expression of primary mouse hepatocytes. These hepatocytes were exposed to three well-defined hepatotoxicants: acetaminophen, amiodarone, and cyclosporin A. Each hepatotoxicant induces a different hepatotoxic phenotype. Based on the DIGE results, the mRNA expression levels of deregulated proteins from cyclosporin A-treated cells were also analyzed. We were able to distinguish cyclosporin A from controls, as well as acetaminophen and amiodarone-treated samples. Cyclosporin A induced endoplasmic reticulum (ER) stress and altered the ER-Golgi transport. Moreover, liver carboxylesterase and bile salt sulfotransferase were differentially expressed. These proteins were associated with a protective adaptive response against cyclosporin A-induced cholestasis. The results of this study are comparable with effects in HepG2 cells. Therefore, we suggest both models can be used to analyze the cholestatic properties of cyclosporin A. Furthermore, this study showed a conserved response between primary mouse hepatocytes and HepG2 cells. These findings collectively lend support for use of omics strategies in preclinical toxicology, and might inform future efforts to better link preclinical and clinical research in rational drug development. PMID:23308384

  15. 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

  16. Screening for drug-induced hepatotoxicity in primary mouse hepatocytes using acetaminophen, amiodarone, and cyclosporin a as model compounds: an omics-guided approach.

    PubMed

    Van Summeren, Anke; Renes, Johan; Lizarraga, Daneida; Bouwman, Freek G; Noben, Jean-Paul; van Delft, Joost H M; Kleinjans, Jos C S; Mariman, Edwin C M

    2013-02-01

    Drug-induced hepatotoxicity is a leading cause of attrition for candidate pharmaceuticals in development. New preclinical screening methods are crucial to predict drug toxicity prior to human studies. Of all in vitro hepatotoxicity models, primary human hepatocytes are considered as 'the gold standard.' However, their use is hindered by limited availability and inter-individual variation. These barriers may be overcome by using primary mouse hepatocytes. We used differential in gel electrophoresis (DIGE) to study large-scale protein expression of primary mouse hepatocytes. These hepatocytes were exposed to three well-defined hepatotoxicants: acetaminophen, amiodarone, and cyclosporin A. Each hepatotoxicant induces a different hepatotoxic phenotype. Based on the DIGE results, the mRNA expression levels of deregulated proteins from cyclosporin A-treated cells were also analyzed. We were able to distinguish cyclosporin A from controls, as well as acetaminophen and amiodarone-treated samples. Cyclosporin A induced endoplasmic reticulum (ER) stress and altered the ER-Golgi transport. Moreover, liver carboxylesterase and bile salt sulfotransferase were differentially expressed. These proteins were associated with a protective adaptive response against cyclosporin A-induced cholestasis. The results of this study are comparable with effects in HepG2 cells. Therefore, we suggest both models can be used to analyze the cholestatic properties of cyclosporin A. Furthermore, this study showed a conserved response between primary mouse hepatocytes and HepG2 cells. These findings collectively lend support for use of omics strategies in preclinical toxicology, and might inform future efforts to better link preclinical and clinical research in rational drug development.

  17. 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

    2010-01-01

    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. PMID:20671795

  18. Preparation and Evaluation of Poly(Ethylene Glycol)-Poly(Lactide) Micelles as Nanocarriers for Oral Delivery of Cyclosporine A

    NASA Astrophysics Data System (ADS)

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

    2010-06-01

    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.

  19. 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

  20. Role of sulphated polysaccharides from Sargassum Wightii in Cyclosporine A-induced oxidative liver injury in rats

    PubMed Central

    Josephine, Anthony; Nithya, Kalaiselvam; Amudha, Ganapathy; Veena, Coothan Kandaswamy; Preetha, Sreenivasan P; Varalakshmi, Palaninathan

    2008-01-01

    Background Seaweeds or marine algae have long been made up a key part of the Asian diet, and as an antioxidant, sulphated polysaccharides have piqued the interest of many researchers as one of the ocean's greatest treasures. The present investigation suggests the therapeutic potential of sulphated polysaccharides from marine brown algae "Sargassum wightii" in Cyclosporine A (CsA)- induced liver injury. CsA is a potent immunosuppressive agent used in the field of organ transplantations and various autoimmune disorders. However, hepatotoxicity due to CsA remains to be one of the major clinical challenges. Methods The effect of sulphated polysaccharides on CsA-induced hepatotoxicity was studied in adult male albino rats of Wistar strain, and the animals were randomized into four groups with six rats in each. Group I served as vehicle control. Group II rats were given CsA at a dosage of 25 mg/kg body weight, orally for 21 days. Group III rats were given sulphated polysaccharides at a dosage of 5 mg/kg body weight, subcutaneously for 21 days. Group IV rats were given sulphated polysaccharides simultaneously along with CsA, as mentioned in Group II for 21 days. Results CsA provoked hepatotoxicity was evident from the decreased activities of hepatic marker enzymes. A significant rise in the level of oxidants, along with a striking decline in both the enzymic and non-enzymic antioxidants, marks the severity of oxidative stress in CsA-induced rats. This in turn led to enhanced levels of lipid peroxidation, 8-hydroxy-2-deoxy guanosine and protein carbonyls, along with a decrease in ATPase activities and alterations in lipid profile. Histopathological changes also strongly support the above aberrations. However, concomitant treatment with sulphated polysaccharides restored the above deformities to near control and prevented the morphological alterations significantly. Conclusion Thus, the present study highlights that sulphated polysaccharides can act therapeutically against

  1. Cyclosporin A and methotrexate in canine marrow transplantation: engraftment, graft-versus-host disease, and induction of intolerance

    SciTech Connect

    Deeg, H.J.; Storb, R.; Weiden, P.L.; Raff, R.F.; Sale, G.E.; Atkinson, K.; Graham, T.C.; Thomas, E.D.

    1982-07-01

    We examined the effect of methotrexate (MTX) and cyclosporin A (Cy A) on engraftment, graft-versus-host disease (GVHD), and the induction of tolerance in dogs prepared for marrow transplantation by 9 Gy of total body irradiation and grafted with bone marrow and buffy coat cells. Nineteen dogs were given grafts from DLA-identical littermates followed by immunosuppression with Cy A for 25 or 100 days. All had sustained engraftment, and 12 became healthy long-term chimeras. Sixty dogs were given grafts from DLA-nonidentical unrelated donors. Among nine given MTX only postgrafting, one rejected the graft nd eight died with GVHD. Among 18 dogs given Cy A only postgrafting, eight failed to achieve engraftment, seven died of various causes, and three died with GVHD. Thirty-four dogs were given both MTX and Cy A in various regimens postgrafting. The only long-term survivors were 4 of 10 dogs given MTX on days 1, 3, 6, and 11 and Cy A from days 0 through 100. Two have chronic GVHD. We conclude that Cy A can induce graft-host tolerance across minor, but not major, histocompatibility differences. The combination of MTX early after transplantation with Cy A prevents failure of engraftment of histoincompatible marrow and some recipients become long-term survivors.

  2. Effects of dietary sodium and magnesium on cyclosporin A-induced hypertension and nephrotoxicity in spontaneously hypertensive rats.

    PubMed

    Mervaala, E M; Pere, A K; Lindgren, L; Laakso, J; Teräväinen, T L; Karjala, K; Vapaatalo, H; Ahonen, J; Karppanen, H

    1997-03-01

    Arterial hypertension, nephrotoxicity, and magnesium loss are common side effects of the immunosuppressive agent cyclosporin A (CsA). In the present study, the effects of dietary sodium and magnesium on CsA toxicity were examined in spontaneously hypertensive rats. A 6-week treatment with CsA during a moderately low-sodium diet (Na 0.3%, Mg 0.2% of the dry weight of the chow) raised blood pressure only slightly, without evidence of nephrotoxicity. By contrast, CsA during a high-sodium diet (Na 2.6%) produced a pronounced rise in blood pressure as well as marked nephrotoxicity, comprising decreased creatinine clearance, increased levels of serum creatinine and urea, and increased urinary protein excretion. During the high-sodium diet, CsA decreased myocardial and bone magnesium concentration and increased myocardial and renal calcium concentration. Magnesium supplementation (Mg 0.6%) protected against the CsA-induced hypertension and nephrotoxicity during the high-sodium diet. Magnesium supplementation also completely prevented the CsA-induced myocardial magnesium depletion and calcium accumulation in the heart and kidney during the high-sodium diet. Our findings indicate a detrimental interaction between increased sodium intake and CsA treatment and a marked protection by concomitant oral magnesium supplementation.

  3. Calcineurin inhibitors cyclosporin A and tacrolimus protect against podocyte injury induced by puromycin aminonucleoside in rodent models

    PubMed Central

    Shen, Xiujin; Jiang, Hong; Ying, Meike; Xie, Zhoutao; Li, Xiayu; Wang, Haibing; Zhao, Jie; Lin, Chuan; Wang, Yucheng; Feng, Shi; Shen, Jia; Weng, Chunhua; Lin, Weiqiang; Wang, Huiping; Zhou, Qin; Bi, Yan; Li, Meng; Wang, Lingyan; Zhu, Tongyu; Huang, Xiaoru; Lan, Hui-Yao; Zhou, Jing; Chen, Jianghua

    2016-01-01

    Podocyte injury and the appearance of proteinuria are features of minimal-change disease (MCD). Cyclosporin A (CsA) and tacrolimus (FK506) has been reported to reduce proteinuria in patients with nephrotic syndrome, but mechanisms remain unknown. We, therefore, investigated the protective mechanisms of CsA and FK506 on proteinuria in a rat model of MCD induced by puromycin aminonucleoside (PAN) and in vitro cultured mouse podocytes. Our results showed that CsA and FK506 treatment decreased proteinuria via a mechanism associated to a reduction in the foot-process fusion and desmin, and a recovery of synaptopodin and podocin. In PAN-treated mouse podocytes, pre-incubation with CsA and FK506 restored the distribution of the actin cytoskeleton, increased the expression of synaptopodin and podocin, improved podocyte viability, and reduced the migrating activities of podocytes. Treatment with CsA and FK506 also inhibited PAN-induced podocytes apoptosis, which was associated with the induction of Bcl-xL and inhibition of Bax, cleaved caspase 3, and cleaved PARP expression. Further studies revealed that CsA and FK506 inhibited PAN-induced p38 and JNK signaling, thereby protecting podocytes from PAN-induced injury. In conclusion, CsA and FK506 inhibit proteinuria by protecting against PAN-induced podocyte injury, which may be associated with inhibition of the MAPK signaling pathway. PMID:27580845

  4. Liver-targeted cyclosporine A-encapsulated poly (lactic-co-glycolic) acid nanoparticles inhibit hepatitis C virus replication.

    PubMed

    Jyothi, K R; Beloor, Jagadish; Jo, Ara; Nguyen, Minh Nam; Choi, Tae Gyu; Kim, Jin-Hwan; Akter, Salima; Lee, Sang-Kyung; Maeng, Chi Hoon; Baik, Hyung Hwan; Kang, Insug; Ha, Joohun; Kim, Sung Soo

    2015-01-01

    Therapeutic options for hepatitis C virus (HCV) infection have been limited by drug resistance and adverse side effects. Targeting the host factor cyclophilin A (CypA), which is essential for HCV replication, offers a promising strategy for antiviral therapy. However, due to its immunosuppressive activity and severe side effects, clinical application of cyclosporine A (CsA) has been limited as an antiviral agent. To overcome these drawbacks, we have successfully developed a liver-specific, sustained drug delivery system by conjugating the liver-targeting peptide (LTP) to PEGylated CsA-encapsulated poly (lactic-co-glycolic) acid (PLGA) nanoparticles. Furthermore, our delivery system exhibited high specificity to liver, thus contributing to the reduced immunosuppressive effect and toxicity profile of CsA. Finally, targeted nanoparticles were able to effectively inhibit viral replication in vitro and in an HCV mouse model. As a proof of principle, we herein show that our delivery system is able to negate the adverse effects of CsA and produce therapeutic effects in an HCV mouse model.

  5. 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

  6. 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. PMID:17853428

  7. Calcineurin inhibitors cyclosporin A and tacrolimus protect against podocyte injury induced by puromycin aminonucleoside in rodent models.

    PubMed

    Shen, Xiujin; Jiang, Hong; Ying, Meike; Xie, Zhoutao; Li, Xiayu; Wang, Haibing; Zhao, Jie; Lin, Chuan; Wang, Yucheng; Feng, Shi; Shen, Jia; Weng, Chunhua; Lin, Weiqiang; Wang, Huiping; Zhou, Qin; Bi, Yan; Li, Meng; Wang, Lingyan; Zhu, Tongyu; Huang, Xiaoru; Lan, Hui-Yao; Zhou, Jing; Chen, Jianghua

    2016-01-01

    Podocyte injury and the appearance of proteinuria are features of minimal-change disease (MCD). Cyclosporin A (CsA) and tacrolimus (FK506) has been reported to reduce proteinuria in patients with nephrotic syndrome, but mechanisms remain unknown. We, therefore, investigated the protective mechanisms of CsA and FK506 on proteinuria in a rat model of MCD induced by puromycin aminonucleoside (PAN) and in vitro cultured mouse podocytes. Our results showed that CsA and FK506 treatment decreased proteinuria via a mechanism associated to a reduction in the foot-process fusion and desmin, and a recovery of synaptopodin and podocin. In PAN-treated mouse podocytes, pre-incubation with CsA and FK506 restored the distribution of the actin cytoskeleton, increased the expression of synaptopodin and podocin, improved podocyte viability, and reduced the migrating activities of podocytes. Treatment with CsA and FK506 also inhibited PAN-induced podocytes apoptosis, which was associated with the induction of Bcl-xL and inhibition of Bax, cleaved caspase 3, and cleaved PARP expression. Further studies revealed that CsA and FK506 inhibited PAN-induced p38 and JNK signaling, thereby protecting podocytes from PAN-induced injury. In conclusion, CsA and FK506 inhibit proteinuria by protecting against PAN-induced podocyte injury, which may be associated with inhibition of the MAPK signaling pathway. PMID:27580845

  8. 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

  9. Treatment effect of cyclosporine A in patients with painful bladder syndrome/interstitial cystitis: A systematic review

    PubMed Central

    WANG, ZHIKUI; ZHANG, LEI

    2016-01-01

    Cyclosporine A (CyA) is emerging as a potential therapeutic strategy for painful bladder syndrome/interstitial cystitis (PBS/IC), which is currently an incurable disease. The purpose of this systematic review was to evaluate the treatment effects of CyA in PBS/IC. Electronic and manual retrieval procedures were carried out to identify eligible references for the systematic review. The entire contents of the included articles were assessed, from study design to reported results. Eight studies, comprising three randomized controlled trials (RCTs), four prospective studies and one retrospective cohort study, were included, involving a total of 298 subjects. Meta-analysis was not implemented due to heterogeneity of the manner of reporting the outcome parameters. All studies reported an improvement in symptoms following treatment with CyA. The results of the three RCTs implied that the treatment effects of CyA were better than those of pentosan polysulfate sodium. Some adverse events, for example, elevation of serum creatinine levels and an increase in blood pressure, were noted in five studies. In conclusion, the evidence from the studies implied that treatment of CyA can result in a long-term benefit in patients of PBS/IC; however, further evidence is required to verify this. PMID:27347076

  10. Cross-Linking the TCR Complex Induces Apoptosis in CD4+8+ Thymocytes in the Presence of Cyclosporin A

    PubMed Central

    Poetschke, Heather L.; Klug, David B.; Walker, Dawn

    1996-01-01

    Although it is generally agreed that TCR ligation is a minimal requirement for negative selection in the CD+8+ double-positive (DP) thymocyte subset, the costimulatory requirements and specific signaling events necessary to induce apoptosis are not well defined. We have explored the consequences of cross-linking CD3/TCR complexes on thymocytes from H-Y TCR transgenic (Tg) mice. In agreement with previous reports, we demonstrate that culturing DP thymocytes with plate-bound anti-TCR antibody induces downregulation of CD4 and CD8 and upregulation of CD69 expression. Nevertheless, the activated cells did not undergo apoptosis, as determined by viable cell recoveries and by quantitation of DNA fragmentation using the TUNEL assay. However, specific depletion of the DP subset occurred within 24 hr when thymocytes were incubated in the presence of both anti-TCR and the immunosuppressant cyclosporin A (CsA). CsA also induced depletion of anti-CD3 stimulated normal DP thymocytes. Using mice homozygous for the lpr or gld mutation, we also have shown that Fas/Fas ligand interactions are not involved in the CsA-induced death of TCR-stimulated DP thymocytes. These data verify that TCR cross-linking alone is insufficient to induce apoptosis of DP thymocytes and further suggest that TCR stimulation activates a CsA-sensitive protective pathway that interferes with signaling events leading to apoptosis in DP thymocytes. PMID:8828007

  11. A hydrogel composite system for sustained epi-cortical delivery of Cyclosporin A to the brain for treatment of stroke.

    PubMed

    Caicco, Matthew J; Cooke, Michael J; Wang, Yuanfei; Tuladhar, Anup; Morshead, Cindi M; Shoichet, Molly S

    2013-03-28

    Stimulation of endogenous neural stem/progenitor cells (NSPCs) with therapeutic factors holds potential for the treatment of stroke. Cyclosporin A (CsA) is a particularly promising candidate molecule because it has been shown to act as a survival factor for these cells over a period of weeks both in vitro and in vivo; however, systemically-delivered CsA compromises the entire immune system, necessitating sustained localized delivery. Herein we describe a local delivery strategy for CsA using an epi-cortical hydrogel of hyaluronan-methylcellulose (HAMC) as the drug reservoir. Three methods of incorporating the drug into the hydrogel (solubilized, particulate, and poly(lactic-co-glycolic) acid (PLGA) microsphere-encapsulated) resulted in tunable release, spanning a period of hours to weeks. Importantly, PLGA-encapsulated CsA released from the hydrogel had equivalent bioactivity to fresh drug as measured by the neurosphere assay. Moreover, when CsA was released from the PLGA/HAMC composite that was injected on the cortex of adult mice, CsA was detected in the NSPC niche at a constant concentration for at least 24days post-implant. Thus this hydrogel composite system may be promising for the treatment of stroke. PMID:23306024

  12. Alpha-difluoromethylornithine, an inhibitor of polyamine biosynthesis, augments cyclosporin A inhibition of cytolytic T lymphocyte induction.

    PubMed Central

    Bowlin, T L; Rosenberger, A L; McKown, B J

    1989-01-01

    The objective of the present investigation was to examine the effect of alpha-difluoromethylornithine (DFMO), an irreversible inhibitor of ornithine decarboxylase, in combination with the immunosuppressant cyclosporin A (CsA) on cytolytic T lymphocytes (CTL) induction in vitro and in vivo. Treatment with DFMO (0.2 mg/ml) or CsA (10 ng/ml) alone in vitro inhibited mitogen-induced CTL generation by 56% and 51%, respectively. Similarly, DFMO or CsA treatment alone inhibited alloantigen-induced CTL generation by 50% and 62%, respectively. Combination treatment with DFMO and CsA reduced mitogen- and alloantigen-mediated CTL induction by 79% and 90%, respectively. In vivo, DFMO treatment alone did not inhibit alloantigen induced CTL generation. However, DFMO potentiated the immunosuppressive effects of CsA in vivo on CTL induction. DFMO treatment reduced activated lymphocyte putrescine and spermidine levels by 81% and 91%, respectively. Combination treatment with DFMO and CsA, at concentrations that effectively inhibited CTL induction, did not further deplete polyamine levels beyond those levels observed with DFMO alone. CsA treatment with or without DFMO did reduce detectable levels of interleukin 2 (IL-2) activity. DFMO treatment alone did not impair IL-2 production. These results indicate that CsA and DFMO may inhibit different processes required for CTL induction, IL-2 production and polyamine biosynthesis. Therefore, inhibitors of polyamine biosynthesis may be useful in lowering the doses of CsA required to inhibit CTL induction. PMID:2504519

  13. alpha-Melanocyte stimulating hormone (MSH) decreases cyclosporine a induced apoptosis in cultured human proximal tubular cells.

    PubMed Central

    Jo, S. K.; Lee, S. Y.; Han, S. Y.; Cha, D. R.; Cho, W. Y.; Kim, H. K.; Won, N. H.

    2001-01-01

    The pathogenesis of chronic cyclosporine A (CsA) nephrotoxicity has not been elucidated, but apoptosis is thought to play an important role in CsA induced tubular atrophy. Recently Fas-Fas ligand system mediated apoptosis has been frequently reported in many epithelial cells as well as in T lymphocytes. We investigated the ability of CsA to induce apoptosis in cultured human proximal tubular epithelial cells and also the effect of alpha-MSH on them. Fas, Fas ligand, and an intracellular adaptor protein, Fas-associating protein with death domain (FADD) expression, and poly-ADP ribose polymerase (PARP) cleavage were also studied. CsA induced apoptosis in cultured tubular epithelial cells demonstrated by increased number of TUNEL positive cells and it was accompanied by a significant increase in Fas mRNA and Fas ligand protein expressions. FADD and the cleavage product of PARP also increased, indicating the activation of caspase. In alpha-MSH co-treated cells, apoptosis markedly decreased with downregulation of Fas, Fas ligand and FADD expressions and also the cleavage product of PARP. In conclusion, these data suggest that tubular cell apoptosis mediated by Fas system may play a role in tubular atrophy in chronic CsA nephrotoxicity and pretreatment of alpha-MSH may have a some inhibitory effect on CsA induced tubular cell apoptosis. PMID:11641530

  14. Successful combination therapy with corticosteroids, biweekly intravenous pulse cyclophosphamide and cyclosporin A for acute interstitial pneumonia in patients with dermatomyositis : report of three cases.

    PubMed

    Suzuki, Akitake; Shoji, Norikazu; Kikuchi, Eigo; Uekubo, Kazuaki; Aoki, Naoko; Sonoda, Yui; Torigai, Hideyuki; Yamashita, Hiroyuki; Fujita, Kentaro; Okai, Takahiro

    2013-01-01

    We report three patients with dermatomyositis (DM) complicated with acute interstitial pneumonia (AIP). All of them complained of fever and acutely worsening dyspnea and were treated immediately by combination therapies with pulse therapy with methylprednisone (mPSL) followed by corticosteroids, biweekly intravenous pulse cyclophosphamide (IVCY) and cyclosporine A (CSA). They recovered rapidly soon after an initiation of this combination regimen. Early intervention with aggressive combination therapy is life-saving for the treatment of AIP in patients with DM.

  15. Development of a cyclosporin-A-induced immune tolerant rat model to test marrow allograft cell type effects on bone repair.

    PubMed

    Espitalier, Florent; Durand, Nicolas; Rémy, Séverine; Corre, Pierre; Sourice, Sophie; Pilet, Paul; Weiss, Pierre; Guicheux, Jérôme; Malard, Olivier

    2015-05-01

    Bone repair is an important concept in tissue engineering, and the ability to repair bone in hypotrophic conditions such as that of irradiated bone, represents a challenge for this field. Previous studies have shown that a combination of bone marrow and (BCP) was effective to repair irradiated bone. However, the origin and role played by each cell type in bone healing still remains unclear. In order to track the grafted cells, the development of an animal model that is immunotolerant to an allograft of bone marrow would be useful. Furthermore, because the immune system interacts with bone turnover, it is of critical importance to demonstrate that immunosuppressive drugs do not interfere with bone repair. After a preliminary study of immunotolerance, cyclosporin-A was chosen to be used in immunosuppressive therapy. Ten rats were included to observe qualitative and quantitative bone repair 8 days and 6 weeks after the creation of bone defects. The defects were filled with an allograft of bone marrow alone or in association with BCP under immunosuppressive treatment (cyclosporin-A). The results showed that there was no significant interaction of cyclosporin-A with osseous regeneration. The use of this new immunotolerant rat model of bone marrow allograft in future studies will provide insight on how the cells within the bone marrow graft contribute to bone healing, especially in irradiated conditions.

  16. Short-term effects of topical cyclosporine A 0.05% (Restasis) in long-standing prosthetic eye wearers: a pilot study

    PubMed Central

    Han, J W; Yoon, J S; Jang, S Y

    2014-01-01

    Purpose Long-standing prosthetic eye wearing induces ocular surface inflammation. We investigated the short-term effects of topical cyclosporine A 0.05% (Restasis) in patients with ocular discomfort resulting from long-standing prosthetic eye wearing. Methods This was a prospective, interventional case series. Patients who were unilateral prosthetic eye wearers over a period of 5 years were enrolled at a single institution from March to July 2013. The subjects were instructed to instill topical cyclosporine A 0.05% twice per day. Measurements were made pre-treatment and after 1 and 3 months of treatment. Outcome measures were the ocular symptom score, the lid margin abnormality score, the Schirmer test, and the tear meniscus amount, using Fourier-domain optical coherence tomography. Results In total, 20 consecutive patients (mean age: 60.1 years, 8 males, 12 females) were included. Ocular symptoms were improved after treatment for 1 month in all patients (ocular symptom score pre-treatment 76.83 vs 46.75 after treatment; P<0.001). There was no statistically significant difference in lid margin abnormality score or tear meniscus amount. The Schirmer test results were improved after treatment for 3 months (pre- and after treatment, 6.70 vs 11.40; P<0.001). Conclusions Topical cyclosporine A 0.05% showed a satisfactory effect in long-standing prosthetic eye wearers. Ocular symptoms were markedly relieved in all subjects after treatment for 1 month. PMID:25081289

  17. 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.

  18. Alterations in cyclosporin A pharmacokinetics and metabolism during treatment with St John's wort in renal transplant patients

    PubMed Central

    Bauer, Steffen; Störmer, Elke; Johne, Andreas; Krüger, Hagen; Budde, Klemens; Neumayer, Hans-Hellmut; Roots, Ivar; Mai, Ingrid

    2003-01-01

    Aim This study investigated the effects of St John's wort extract (SJW) on the pharmacokinetics and metabolism of the immunosuppressant cyclosporin A (CSA). Methods In an open-label study, 11 renal transplant patients received 600 mg SJW extract daily for 14 days in addition to their regular regimen of CSA. Blood concentrations of CSA and its metabolites AM1, AM1C, AM9, AM19, and AM4N were measured by HPLC. Results After 2 weeks of SJW coadministration, dose-corrected AUC0–12, Cmax and Ctrough values for CSA decreased significantly by 46%[geometric mean ratio baseline/SJW (95% CI): 1.83 (1.63–2.05)], 42%[1.72 (1.42–2.08)], and 41%[1.70 (1.17–2.47)], respectively. CSA doses were increased from a median of 2.7 mg day−1 kg−1 at baseline to 4.2 mg day−1 kg−1 at day 15, with the first dose adjustment required only 3 days after initiation of SJW treatment. Additionally, the metabolite pattern of CSA was substantially altered during SJW treatment. Whereas dose-corrected AUC values for AM1, AM1c and AM4N significantly decreased by 59%, 61%, and 23% compared with baseline, AUC values for AM9 and AM19 were unchanged. Following the increase in CSA dose, observed AUC and Cmax values for AM9, AM19, and AM4N increased by 20–51% and 43–90%, respectively. Conclusion Administration of SJW extract to patients receiving CSA treatment resulted in a rapid and significant reduction of plasma CSA concentrations. Additionally, the substantial alterations in CSA metabolite kinetics observed may affect the toxicity profile of the drug. PMID:12580993

  19. Pharmacokinetics and enhanced oral bioavailability in beagle dogs of cyclosporine A encapsulated in glyceryl monooleate/poloxamer 407 cubic nanoparticles.

    PubMed

    Lai, Jie; Lu, Yi; Yin, Zongning; Hu, Fuqiang; Wu, Wei

    2010-01-01

    Efforts to improve the oral bioavailability of cyclosporine A (CyA) remains a challenge in the field of drug delivery. In this study, glyceryl monooleate (GMO)/poloxamer 407 cubic nanoparticles were evaluated as potential vehicles to improve the oral bioavailability of CyA. Cubic nanoparticles were prepared via the fragmentation of a bulk GMO/poloxamer 407 cubic phase gel by sonication and homogenization. The cubic inner structure formed was verified using Cryo-TEM. The mean diameters of the nanoparticles were about 180 nm, and the entrapment efficiency of these particles for CyA was over 85%. The in vitro release of CyA from these nanoparticles was less than 5% at 12 h. The results of a pharmacokinetic study in beagle dogs showed improved absorption of CyA from cubic nanoparticles as compared to microemulsion-based Neoral((R)); higher C(max) (1371.18 +/- 37.34 vs 969.68 +/- 176.3 ng mL(-1)), higher AUC(0-t) (7757.21 +/- 1093.64 vs 4739.52 +/- 806.30 ng h mL(-1)) and AUC(0-infinity) (9004.77 +/- 1090.38 vs 5462.31 +/- 930.76 ng h mL(-1)). The relative oral bioavailability of CyA cubic nanoparticles calculated on the basis of AUC(0-infinity) was about 178% as compared to Neoral((R)). The enhanced bioavailability of CyA is likely due to facilitated absorption by cubic nanoparticles rather than improved release.

  20. The effects of cyclosporin-A on axonal conduction deficits following traumatic brain injury in adult rats

    PubMed Central

    Colley, Beverly S.; Phillips, Linda L.; Reeves, Thomas M.

    2010-01-01

    Immunophilin ligands, including cyclosporin-A (CsA), have been shown to be neuroprotective in experimental models of traumatic brain injury (TBI), and to attenuate the severity of traumatic axonal injury. Prior studies have documented CsA treatment to reduce essential components of posttraumatic axonal pathology, including impaired axoplasmic transport, spectrin proteolysis, and axonal swelling. However, the effects of CsA administration on axonal function, following TBI, have not been evaluated. The present study assessed the effects of CsA treatment on compound action potentials (CAPs) evoked in corpus callosum of adult rats following midline fluid percussion injury. Rats received a 20 mg/Kg bolus of CsA, or cremaphor vehicle, at either 15 m or 1 h postinjury, and at 24 h postinjury CAP recording was conducted in coronal brain slices. To elucidate how injury and CsA treatments affect specific populations of axons, CAP waveforms generated largely by myelinated axons (N1) were analyzed separately from the CAP signal which predominantly reflects activity in unmyelinated axons (N2). CsA administration at 15 m postinjury resulted in significant protection of CAP area, and this effect was more pronounced in N1, than in the N2, CAP component. This treatment also significantly protected against TBI-induced reductions in high frequency responding of the N1 CAP signal. In contrast, CsA treatment at 1 h did not significantly protect CAPs, but was associated with atypical waveforms in N1 CAPs, including decreased CAP duration and reduced refractoriness. The present findings also support growing evidence that myelinated and unmyelinated axons respond differentially to injury and neuroprotective compounds. PMID:20362574

  1. Protective effect of ellagic acid against cyclosporine A-induced histopathological, ultrastructural changes, oxidative stress, and cytogenotoxicity in albino rats.

    PubMed

    Abdul-Hamid, Manal; Abdella, Ehab M; Galaly, Sanaa R; Ahmed, Rania H

    2016-01-01

    Cyclosporine A (CsA) is an immunosuppressor agent, which is most frequently used in transplant surgeries and in the treatment of autoimmune diseases. This study was undertaken to investigate the protective effects of ellagic acid (EA) against CsA-induced testicular histopathology and ultrastructure changes, oxidative stress, and cytogenotoxicity in male albino rats. Rats were divided into six groups; the first group was used as a control, the second group received a subcutaneous injection of slightly alkaline solution, the third group received olive oil orally, the fourth group was injected subcutaneously with EA at a dose of 10 mg/kg b. wt./day, the fifth group was treated with CsA as oral solution at a dose of 15 mg/kg b. wt for 30 days, and the sixth group was treated with CsA simultaneously with EA. Treatment with EA simultaneously with CsA resulted in significant protection. The positive control animals taking CsA alone showed marked histopathological, ultrastructure, and genetic manifestations accompanied by an elevated content of lipid peroxidation and marked reduction of catalase (CAT), peroxidase (Px) activity, and glutathione concentration in the homogenate of testis tissues. The toxic side effects in testis and bone marrow tissues were greatly ablated with a significant reduction in lipid peroxidation level and elevation in CAT and Px activities and glutathione concentration when using EA. Thus, EA may be used in combination with CsA to improve the histopathological, oxidative stress, and cytogenotoxicity parameters of testicular toxicity induced by CsA due to its antioxidant effects. PMID:27430433

  2. 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.

  3. 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.

  4. 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

  5. Nifedipine improves immediate, and 6- and 12-month graft function in cyclosporin A (CyA) treated renal allograft recipients.

    PubMed

    Harper, S J; Moorhouse, J; Veitch, P S; Horsburgh, T; Walls, J; Bell, P R; Donnelly, P K; Feehally, J

    1992-01-01

    To investigate the effect of oral nifedipine, a calcium channel blocker known not to modify cyclosporin A (CyA) pharmacokinetics, on immediate transplant function and CyA nephrotoxicity, 68 adult renal transplant recipients were pre-operatively randomized to one of three regimes: A (high-dose CyA, initial dose 17 mg/kg per day, maintenance dose 7 mg/kg per day); B (regime A plus oral nifedipine); C low-dose CyA, initial dose 10 mg/kg per day, maintenance 4 mg/kg per day plus azathioprine 1 mg/kg per day). All three groups received identical steroid regimes. Calcium channel blockers of all types were avoided in groups A and C. Delayed graft function (dialysis dependence by day 4) was seen least frequently in group B (P < 0.02). Group B had improved graft function at 6 months compared with group A, identified by differences in serum creatinine (P < 0.05), GFR (P < 0.01) and ERPF (P < 0.05). Similar differences in serum creatinine (P < 0.05) and GFR (P < 0.05) were also identified at 12 months. Group C also had better 6- and 12-month GFR values than group A (P < 0.05 each). The three groups did not differ in donor or recipient age, HLA matching, ischaemic or anastomosis times, frequency of early rejection or whole-blood CyA levels. These results indicate that nifedipine significantly improves immediate and medium-term graft function.

  6. Relationship between P-glycoprotein expression and cyclosporin A in kidney. An immunohistological and cell culture study.

    PubMed Central

    García del Moral, R.; O'Valle, F.; Andújar, M.; Aguilar, M.; Lucena, M. A.; López-Hidalgo, J.; Ramírez, C.; Medina-Cano, M. T.; Aguilar, D.; Gómez-Morales, M.

    1995-01-01

    P-glycoprotein (P-gp), encoded in humans by the mdr-1 gene, acts physiologically as an efflux pump to expel hydrophobic substances from cells. This glycoprotein is closely related to multidrug resistance in tumor cells and can be modulated by cyclosporin A (CsA). We investigated the relationship between CsA and P-gp in 52 renal allograft biopsies and in cultures of Madin-Darby canine kidney (MDCK) renal tubule cells to determine whether the intrarenal accumulation of CsA or chronic stimulation with the drug modified the expression of P-gp. Expression of P-gp and CsA was analyzed by immunohistochemistry. Immunostaining was evaluated semiquantitatively. Modulation of P-gp in MDCK cells after chronic stimulation with CsA for 7, 30, and 60 days was analyzed by flow cytometry. P-gp and CsA immunostaining in renal post-transplant biopsies showed considerable overlap in all cases (Spearman's test, r = 0.577, P < 0.001). After 7 days in vitro, the number of cells expressing P-gp increased progressively; a further increase in mean fluorescence was found after 60 days (P < 0.001, Student's t-test). Our findings suggest that in non-neoplastic cells, CsA may stimulate P-gp as a mechanism of detoxification. Individual differences in the adaptive responses to glycoprotein may be responsible for the appearance of nephrotoxicity or a CsA-resistant rejection reaction in cases of overexpression on lymphocytes and macrophages. Images Figure 1 PMID:7856751

  7. Anti-parasitic effect of cyclosporin A on Echinococcus granulosus and characterization of the associated cyclophilin protein.

    PubMed

    Colebrook, A L; Jenkins, D D; Lightowlers, M W

    2002-11-01

    Cyclophilins are a family of proteins found ubiquitously in eukaryotes, many of which bind to the immunosuppressive drug cyclosporin A (CsA). CsA has been found to have anti-parasitic effects against a variety of helminth and protozoan parasites and this activity could be mediated via cyclophilin. In this study we characterize a full length cyclophilin gene from Echinococcus granulosus, the associated natural gene and expression pattern, and investigate the functional properties of the recombinant E. granulosus cyclophilin protein. In addition, the effects of CsA were investigated on E. granulosus protoscoleces in in vitro culture. The full length E. granulosus cyclophilin cDNA encodes a protein of 20 kDa and is encoded by a single gene (EGCyP-1) comprising 2 exons separated by a 31 bp intron. The gene is expressed constitutively in all E. granulosus life-cycle stages examined. Recombinant E. granulosus cyclophilin (egCyP-l) exhibited functional enzyme activity as an isomerase. Treatment of in vitro cultures of E. granulosus protoscoleces with CsA was found to be lethal to the parasites. No protoscoleces survived treatment with 10 microg/ml of CsA over 7 culture days, as determined by observing motility and the uptake of toluidine blue dye. Untreated protoscoleces remained viable for the duration of experiments. The survival of protoscoleces was CsA dose dependent. A concentration of 10 microg/ml CsA was 100% lethal while doses of 8 microg/ml and 5 microg/ml resulted in 82% and 32% killing, respectively, after 7 days in culture. The anti-parasitic activity of CsA may have the potential to be developed as a new therapeutic agent for treatment of cystic hydatidosis in humans.

  8. Advantageous effects of immunosuppression with tacrolimus in comparison with cyclosporine A regarding renal function in patients after heart transplantation

    PubMed Central

    Helmschrott, Matthias; Rivinius, Rasmus; Ruhparwar, Arjang; Schmack, Bastian; Erbel, Christian; Gleissner, Christian A; Akhavanpoor, Mohammadreza; Frankenstein, Lutz; Ehlermann, Philipp; Bruckner, Tom; Katus, Hugo A; Doesch, Andreas O

    2015-01-01

    Background Nephrotoxicity is a serious adverse effect of calcineurin inhibitor therapy in patients after heart transplantation (HTX). Aim In this retrospective registry study, renal function within the first 2 years after HTX in patients receiving de novo calcineurin inhibitor treatment, that is, cyclosporine A (CSA) or tacrolimus (TAC), was analyzed. In a consecutive subgroup analysis, renal function in patients receiving conventional tacrolimus (CTAC) was compared with that of patients receiving extended-release tacrolimus (ETAC). Methods Data from 150 HTX patients at Heidelberg Heart Transplantation Center were retrospectively analyzed. All patients were continuously receiving the primarily applied calcineurin inhibitor during the first 2 years after HTX and received follow-up care according to center practice. Results Within the first 2 years after HTX, serum creatinine increased significantly in patients receiving CSA (P<0.0001), whereas in patients receiving TAC, change of serum creatinine was not statistically significant (P=not statistically significant [ns]). McNemar’s test detected a significant accumulation of patients with deterioration of renal function in the first half year after HTX among patients receiving CSA (P=0.0004). In patients receiving TAC, no significant accumulation of patients with deterioration of renal function during the first 2 years after HTX was detectable (all P=ns). Direct comparison of patients receiving CTAC versus those receiving ETAC detected no significant differences regarding renal function between patients primarily receiving CTAC or ETAC treatment during study period (all P=ns). Conclusion CSA is associated with a more pronounced deterioration of renal function, especially in the first 6 months after HTX, in comparison with patients receiving TAC as baseline immunosuppressive therapy. PMID:25759566

  9. Preparation and in vitro/in vivo evaluation of cyclosporin A-loaded nanodecorated ocular implants for subconjunctival application.

    PubMed

    Pehlivan, Sibel Bozdağ; Yavuz, Burçin; Çalamak, Semih; Ulubayram, Kezban; Kaffashi, Abbas; Vural, İmran; Çakmak, Hasan Basri; Durgun, Meltem Ezgi; Denkbaş, Emir Baki; Ünlü, Nurşen

    2015-05-01

    In terms of ocular drug delivery, biodegradable implant systems have several advantages including the ability to provide constant drug concentration at the target site, no necessity for surgical removal, and minimum systemic side effects. Cyclosporin A (CsA) is a neutral, hydrophobic, cyclic peptide of amino acids that frequently used for dry eye disease treatment. The aim of this study was to develop a nanoparticle-loaded implant system for sustained-release CsA delivery following subconjunctival implantation. Poly(lactide-co-glycolide) (85:15) or poly-ε-caprolactone (PCL) were used to prepare two different nanoparticle formulations. These nanoparticles loaded into PCL or poly(lactide-co-caprolactone) implant formulations were prepared by two different methods, which were molding and electrospinning. Size and zeta potential of nanoparticles were determined and the morphology of the formulations were investigated by scanning electron microscopy. CsA-loading efficiencies were calculated and the in vitro degradation and in vitro release studies were performed. MTT test was also performed using L929 fibroblast cells to evaluate the cytotoxicity of the formulations. PCL-PCL-NP-I formulation was implanted to Swiss Albino mice with induced dry eye syndrome to evaluate the efficacy. In vitro release studies showed that the release from the formulations continues between 30 and 60 days, and the cell viability was found to be 77.4%-99.0%. In vivo studies showed that healing is significantly faster in the presence of the selected implant formulation. Results indicated that nanodecorated implants are promising ocular carriers for controlled-release CsA application.

  10. 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.

  11. 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

  12. Cyclosporin A differentially inhibits multiple steps in VEGF induced angiogenesis in human microvascular endothelial cells through altered intracellular signaling

    PubMed Central

    Rafiee, Parvaneh; Heidemann, Jan; Ogawa, Hitoshi; Johnson, Nathan A; Fisher, Pamela J; Li, Mona S; Otterson, Mary F; Johnson, Christopher P; Binion, David G

    2004-01-01

    The immunosuppressive agent cyclosporin A (CsA), a calcineurin inhibitor which blocks T cell activation has provided the pharmacologic foundation for organ transplantation. CsA exerts additional effects on non-immune cell populations and may adversely effect microvascular endothelial cells, contributing to chronic rejection, a long-term clinical complication and significant cause of mortality in solid-organ transplants, including patients with small bowel allografts. Growth of new blood vessels, or angiogenesis, is a critical homeostatic mechanism in organs and tissues, and regulates vascular populations in response to physiologic requirements. We hypothesized that CsA would inhibit the angiogenic capacity of human gut microvessels. Primary cultures of human intestinal microvascular endothelial cells (HIMEC) were used to evaluate CsA's effect on four in vitro measures of angiogenesis, including endothelial stress fiber assembly, migration, proliferation and tube formation, in response to the endothelial growth factor VEGF. We characterized the effect of CsA on intracellular signaling mechanisms following VEGF stimulation. CsA affected all VEGF induced angiogenic events assessed in HIMEC. CsA differentially inhibited signaling pathways which mediated distinct steps of the angiogenic process. CsA blocked VEGF induced nuclear translocation of the transcription factor NFAT, activation of p44/42 MAPK, and partially inhibited JNK and p38 MAPK. CsA differentially affected signaling cascades in a dose dependent fashion and completely blocked expression of COX-2, which was integrally linked to HIMEC angiogenesis. These data suggest that CsA inhibits the ability of microvascular endothelial cells to undergo angiogenesis, impairing vascular homeostatic mechanisms and contributing to the vasculopathy associated with chronic rejection. PMID:15175101

  13. Direct and endothelial cell-mediated effect of cyclosporin A on the proliferation of rat smooth muscle cells in vitro.

    PubMed Central

    Leszczynski, D.; Zhao, Y.; Yeagley, T. J.; Foegh, M. L.

    1993-01-01

    Cyclosporin A (CsA) has been suggested to potentiate graft vascular disease by stimulation of smooth muscle cell (SMC) proliferation. Both the in vitro and in vivo data are discordant, showing both stimulatory and inhibitory effects of CsA on vascular SMC proliferation. The direct and endothelial cell-mediated effects of CsA on vascular SMC proliferation were examined in vitro using the incorporation of [3H]thymidine. All experiments were done in serum-free conditions. The exposure of SMC to CsA (0.0001 to 0.1 micrograms/ml) had no effect on proliferation. High doses of CsA (0.5 to 10.0 micrograms/ml) were toxic to the SMC and endothelial cells; 90% of SMC population died within 3 to 6 days of exposure to 10.0 micrograms/ml CsA. In the studies on the endothelial cell-mediated effect of CsA, the endothelial cell-conditioned medium (ECCM) significantly increased SMC proliferation. This stimulatory effect was significantly attenuated when the ECCM was obtained from endothelial cells exposed to CsA. Endothelin (ET) is suggested to be an endothelial-cell-derived growth factor for SMC, and implicated as a possible cause of the uncontrolled proliferation of SMC during development of graft vascular disease. Exposure of SMC to levels of recombinant ET similar to the levels found in the ECCM (0.19 + 0.01 pg/ml) significantly increased SMC proliferation. CsA increased fivefold ET concentration in the ECCM. However, despite this rise in ET levels, there was a 45% decrease in SMC proliferation. In conclusion, CsA does not exert a direct modulatory effect on SMC proliferation in vitro, but may inhibit SMC proliferation indirectly via endothelial cell-derived factors. These unidentified factor(s) inhibit SMC proliferation and abolish the mitogenic effect of ET on SMC. PMID:8424452

  14. Cyclosporin A in Membrane Lipids Environment: Implications for Antimalarial Activity of the Drug--The Langmuir Monolayer Studies.

    PubMed

    Dynarowicz-Łątka, Patrycja; Wnętrzak, Anita; Makyła-Juzak, Katarzyna

    2015-12-01

    Cyclosporin A (CsA), a hydrophobic cyclic peptide produced by the fungus Tolypocladium inflatum, is well known for its high efficiency as an immunosuppressor for transplanted organs and anti-inflammatory properties; however, it is also active as antiparasitic (antimalarial) drug. Antimalarial mechanism of CsA action lacks a detailed understanding at molecular level. Due to a high lipophilicity of CsA, it is able to interact with lipids of cellular membrane; however, molecular targets of this drug are still unknown. To get a deeper insight into the mode of antimalarial activity of CsA, it is of utmost importance to examine its interactions with membrane components. To reach this goal, the Langmuir monolayer technique, which serves as a very useful, easy to handle and controllable model of biomembranes, has been employed. In this work, the interactions between CsA and main membrane lipids, i.e., cholesterol (Chol), 2-oleoyl-1-palmitoyl-3-phosphocholine (POPC), and sphingomyelin (SM), have been investigated. Attractive interactions are observed only for CsA mixtures with SM, while repulsive forces occur in systems containing remaining membrane lipids. Taking into consideration mutual interactions between membrane lipids (Chol-SM; Chol-POPC and SM-POPC), the behavior of CsA in model erythrocyte membrane of normal and infected cells has been analyzed. Our results prove strong affinity of CsA to SM in membrane environment. Since normal and parasitized erythrocytes differ significantly in the level of SM, this phospholipid may be considered as a molecular target for antimalarial activity of CsA.

  15. 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.

  16. 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

  17. 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

  18. Cyclosporin A Treatment of Leishmania donovani Reveals Stage-Specific Functions of Cyclophilins in Parasite Proliferation and Viability

    PubMed Central

    Yau, Wai-Lok; Blisnick, Thierry; Taly, Jean-François; Helmer-Citterich, Manuela; Schiene-Fischer, Cordelia; Leclercq, Olivier; Li, Jing; Schmidt-Arras, Dirk; Morales, Miguel A.; Notredame, Cedric; Romo, Daniel; Bastin, Philippe; Späth, Gerald F.

    2010-01-01

    Background Cyclosporin A (CsA) has important anti-microbial activity against parasites of the genus Leishmania, suggesting CsA-binding cyclophilins (CyPs) as potential drug targets. However, no information is available on the genetic diversity of this important protein family, and the mechanisms underlying the cytotoxic effects of CsA on intracellular amastigotes are only poorly understood. Here, we performed a first genome-wide analysis of Leishmania CyPs and investigated the effects of CsA on host-free L. donovani amastigotes in order to elucidate the relevance of these parasite proteins for drug development. Methodology/Principal Findings Multiple sequence alignment and cluster analysis identified 17 Leishmania CyPs with significant sequence differences to human CyPs, but with highly conserved functional residues implicated in PPIase function and CsA binding. CsA treatment of promastigotes resulted in a dose-dependent inhibition of cell growth with an IC50 between 15 and 20 µM as demonstrated by proliferation assay and cell cycle analysis. Scanning electron microscopy revealed striking morphological changes in CsA treated promastigotes reminiscent to developing amastigotes, suggesting a role for parasite CyPs in Leishmania differentiation. In contrast to promastigotes, CsA was highly toxic to amastigotes with an IC50 between 5 and 10 µM, revealing for the first time a direct lethal effect of CsA on the pathogenic mammalian stage linked to parasite thermotolerance, independent from host CyPs. Structural modeling, enrichment of CsA-binding proteins from parasite extracts by FPLC, and PPIase activity assays revealed direct interaction of the inhibitor with LmaCyP40, a bifunctional cyclophilin with potential co-chaperone function. Conclusions/Significance The evolutionary expansion of the Leishmania CyP protein family and the toxicity of CsA on host-free amastigotes suggest important roles of PPIases in parasite biology and implicate Leishmania CyPs in key

  19. 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

  20. Adjuvant treatment with cyclosporin A increases the toxicity of chemotherapy for remission induction in acute non-lymphocytic leukemia.

    PubMed

    Damiani, D; Michieli, M; Ermacora, A; Russo, D; Fanin, R; Zaja, F; Baraldo, M; Pea, F; Furlanut, M; Baccarani, M

    1998-08-01

    P-glycoprotein (Pgp)-related multidrug resistance (MDR) is frequently observed in acute non-lymphocytic leukemia (ANLL) and is associated with a poor response to standard chemotherapy. Cyclosporin A (CsA) is an effective downmodulator of Pgp-related MDR in vitro and has already been tested for that purpose in vivo also. Since Pgp is expressed in several normal cells and tissues, the modulation of Pgp can also modify total body exposure to antileukemic drugs and can alter and increase the toxicity of the antileukemic treatment. We report here the results of a study where 46 consecutive adult patients with ANLL were assigned to receive the same standard chemotherapy regimen of arabinosyl cytosine and idarubicin (IDA) for remission induction or consolidation, without or with CsA. Twenty-eight patients received 36 courses of chemotherapy without CsA and 18 patients received 32 courses of chemotherapy with CsA. CsA dose was 10-12.5 mg/kg/day and was given as a continuous i.v. infusion for 72 h. Whole blood CsA steady-state concentration ranged between 0.61 and 1.14 microM. The IDA area-under-the-curve was about twice as high in the cases that received CsA than in the other cases. CsA had no detectable effects on renal function and fluid balance, but significantly increased systemic blood diastolic pressure and conjugated bilirubine concentration. Furthermore, CsA-treated patients had greater, and more severe, oral and intestinal mucosal toxicity, with more severe adverse events, including more cases of gram-negative bacteremia, and with a delayed hemopoietic recovery. In conclusion, this study showed that an attempt at an effective downmodulation of Pgp-mediated MDR would substantially increase the hemopoietic and mucosal toxicity of antileukemic treatment and that the increase is accounted for, at least in part, by an increase of total body exposure to IDA.

  1. 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

  2. Modulation of metabolism in HepG2 cells upon treatment with cyclosporin A and Nva2-cyclosporin.

    PubMed

    Bäckman, L; Appelkvist, E L; Sundberg, A; Teclebrhan, H; Brunk, U

    1991-06-01

    HepG2 cells were cultured in the presence of different concentrations of cyclosporin A (CsA) or Nva2-cyclosporin (Nva2-Cs) for up to 20 days. At a low concentration (2 micrograms/ml) of CsA or Nva2-Cs, the [3H]thymidine incorporation into DNA and the rate of incorporation of [3H]leucine into total protein decreased by 20-25%. Concentrations of 10 micrograms/ml resulted in a 70% reduction of the [3H]thymidine incorporation in comparison with controls. Low concentrations of CsA resulted in mitochondria in the condensed state together with autophagosomes, large vacuoles, and elevated numbers of coated vesicles, as shown by electron microscopy. Low concentrations of Nva2-Cs resulted in swollen mitochondria, increased autophagocytosis, and increased numbers of intermediate filaments and microtubules. Higher doses of these substances (5 micrograms/ml) caused disarrangement of mitochondrial cristae, vesiculation of the endoplasmic reticulum, an elevated number of free polysomes, and accelerated autophagocytosis. Labeling of phospholipids and triglycerides with [3H]glycerol and of cholesterol and dolichol with [3H]acetate was decreased after exposure of HepG2 cells to CsA, or, in particular, Nva2-Cs. Phospholipids secreted from the cells into the medium exhibited an increased level of labeling, but the specific radioactivity of the neutral lipids in the medium was significantly decreased. Treatment of HepG2 cells with either CsA or Nva2-Cs doubled the mitochondrial cytochrome oxidase and carnitine acetyl-transferase, as well as microsomal NADPH-cytochrome c reductase activities. Such treatment also increased the cyanide-insensitive beta-oxidation of fatty acids in peroxisomes, as well as cytoplasmic DT-diaphorase and glutathione transferase activities. Prolonged treatment of the cells with CsA did not result in any cumulative effect. HepG2 cells appear to be suitable for studying the effects of cyclosporins on cellular structure and metabolism and in this system the two

  3. 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

  4. The effect of different fatty acids on the intestinal lymphatic absorption of cyclosporin-A after oral administration in the rat

    SciTech Connect

    Jensen, B.K.

    1988-01-01

    Four studies were conducted in male Sprague-Dawley rats to evaluate the effect of saturated fatty acids (FA) of varying chain lengths on cyclosporin-A (CSA) intestinal lymphatic absorption. {sup 3}H-CSA was given to thoracic duct-ligated and sham rats in a nonlipid-(NL) or busyric (BA), octanoic (OA), lauric (LA), palmitic (PA), or stearic (SA) acid dosage form ({sup 14}C-FA) in an oral absorption study. The dosage forms were given to thoracic duct cannulated (TDC) rats to assess CSA intestinal lymphatic absorption. CSA blood-to-lymph transfer was assessed by intravenous {sup 3}H-CSA in TDC rats. Colchicine pretreated TDC rats received CSA in the NL and PA dosage forms. CSA and FA concentrations in blood and lymph were measured radiometrically. CSA and FA in the chylomicron and aqueous fractions were determined from ultracentrifugation of pooled lymph samples.

  5. Fast LC-MS/MS analysis of tacrolimus, sirolimus, everolimus and cyclosporin A in dried blood spots and the influence of the hematocrit and immunosuppressant concentration on recovery.

    PubMed

    Koster, Remco A; Alffenaar, Jan-Willem C; Greijdanus, Ben; Uges, Donald R A

    2013-10-15

    We developed a method for the analysis of four immunosuppressants in dried blood spot (DBS) samples to facilitate therapeutic drug monitoring for transplant patients outside the hospital. An 8mm disc from the central part of the DBS was punched, extracted and followed by LC-MS/MS analysis. The method was validated with ranges from 1.00-50.0 µg/L for tacrolimus, sirolimus and everolimus, and from 20.0-2000 µg/L for cyclosporin A. The validation showed a maximum overall bias of 13.0% for the sirolimus LLOQ, while the maximum overall CV was 15.7% for the everolimus LLOQ. All four immunosuppressants showed to be stable in DBS for at least 7 days at 22°C. The volume of the blood spot showed to have minor effect on measured concentrations. A cross-validation test between the 31 ET CHR paper and the Whatman FTA DMPK-C cards showed no significant difference between the two types of paper. During validation the hematocrit (HT) showed to have significant influence on the analytical results. When the measured concentrations were corrected for the effect of the HT, biases improved significantly. Additional recovery tests proved that the combination of especially low HT and high concentration does not only affect the spot size but can also affect the extraction recoveries of sirolimus and especially everolimus. Although the tested parameters like HT and concentrations are extreme and unlikely for routine analysis of outpatients, the fundamental effect of the combination of these parameters on extraction recoveries are proven with this research. The protein binding in the blood and hydrogen binding to the cellulose of the paper is suggested to influence extractions and gives new insights in the extraction methodology of DBS samples. The observed HT effect during the validation appeared to be negligible during the correlation study as no concentration corrections for the HT values were needed. Nevertheless, results from DBS samples with extremely high concentrations combined

  6. Comparison of atmospheric pressure chemical ionization, electrospray ionization, and atmospheric pressure photoionization for the determination of cyclosporin A in rat plasma.

    PubMed

    Wang, Ganfeng; Hsieh, Yunsheng; Korfmacher, Walter A

    2005-01-15

    Atmospheric pressure chemical ionization was compared with electrospray ionization and atmospheric pressure photoionization (APPI) as an interface of high-performance liquid chromatography (HPLC)-tandem mass spectrometry (MS/MS) for the determination of cyclosporin A (CsA) in biological fluids in support of in vivo pharmacodynamic studies. These ion sources were investigated in terms of their suitability and sensitivity for the detection of CsA. The effects of the eluent flow rate and composition as well as the nebulizer temperatures on the photoionization efficiency of CsA in the positive ion mode under normal-phase HPLC conditions were explored. The ionization mechanism in the APPI environment with and without the use of the dopant was studied using two test compounds and a few solvent systems employed for normal-phase chromatography. The test compounds were observed to be ionized mainly by proton transfer with the self-protonated solvent molecules produced through photon irradiation. Furthermore, ion suppression due to sample matrix interference in the normal-phase HPLC-APPI-MS/MS system was monitored by the postcolumn infusion technique. The applicability of these proposed HPLC-API-MS/MS approaches for the determination of CsA at low nanogram per milliliter levels in rat plasma was examined. These proposed methods were then compared with respect to specificity, linearity, detection limit, and accuracy.

  7. 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.

  8. Ocular delivery of cyclosporine A based on glyceryl monooleate/poloxamer 407 liquid crystalline nanoparticles: preparation, characterization, in vitro corneal penetration and ocular irritation.

    PubMed

    Chen, Yan; Lu, Yi; Zhong, Yanqiang; Wang, Qingping; Wu, Wei; Gao, Shen

    2012-12-01

    The purpose of this study was to develop an ophthalmic drug delivery system for cyclosporine A (CsA) based on glyceryl monooleate (GMO)/poloxamer 407 liquid crystalline nanoparticles with reduced ocular irritancy and improved corneal penetration. CsA-loaded liquid crystalline nanoparticles were prepared via fragmentation of a bulk GMO/poloxamer 407 cubic phase gel by high-pressure homogenization and characterized. Corneal permeation and retention was evaluated using modified Franz diffusing cells. Intra-corneal transportation was investigated with fluorescein isothiocyanate (FITC)-labeled liquid crystalline nanoparticles. Ocular irritation was then evaluated using the Draize method. The mean particle size of liquid crystalline nanoparticles was 193.5 nm and the entrapment efficiency was 95.11 ± 0.67%. A bicontinuous cubic phase of cubic P-type was determined using cryo-transmission electron microscopy (cryo-TEM) observation and small angle X-ray diffraction (SAXD) analysis. A 1.52-fold increase in J(s) and a 2.2-fold increase in corneal retention was achieved by liquid crystalline nanoparticles compared with oil solution. In vitro corneal permeation investigated with FITC-labeled liquid crystalline nanoparticles revealed that CsA penetrated across the cornea under the transportation of liquid crystalline nanoparticles. Liquid crystalline nanoparticles exhibited excellent ocular tolerance in the ocular irritation test. This low-irritant vehicle based on liquid crystalline nanoparticles might be a promising system for effective ocular CsA delivery.

  9. Characterization of Long-Lasting Oatp Inhibition by Typical Inhibitor Cyclosporine A and In Vitro-In Vivo Discrepancy in Its Drug Interaction Potential in Rats.

    PubMed

    Taguchi, Takayuki; Masuo, Yusuke; Kogi, Tatsuya; Nakamichi, Noritaka; Kato, Yukio

    2016-07-01

    Quantitative assessment of potential drug-drug interactions (DDIs) is one of the major focuses in drug development. The aim of the present study was to quantitatively evaluate in vitro-in vivo discrepancy of DDI potential for prototypical organic anion transporting polypeptide (Oatp) inhibitor cyclosporine A (CsA) using rats. Plasma concentration of pravastatin, prototypical Oatp substrate, after oral administration was increased by CsA intravenously administered at 1 d before the pravastatin administration. The ratio of the area under the curve of pravastatin to the control was much higher than the R-values calculated using the plasma unbound concentrations of CsA and the inhibition constant (Ki) assessed in isolated hepatocytes, indicating in vitro-in vivo discrepancy. This interaction with pravastatin persisted for 3 d after CsA administration, demonstrating long-lasting inhibition in vivo. The Ki value for unbound CsA in the presence of serum was comparable with that in its absence. M1, the major metabolite of CsA inhibited pravastatin uptake at much higher concentration compared with its plasma unbound concentration. Thus, the DDI potential of CsA-mediated hepatic Oatp inhibition cannot be extrapolated from in vitro data, and this could be due to the long-lasting Oatp inhibition by CsA, but not the effect of plasma protein or metabolites.

  10. 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.

  11. Molecular mechanisms underlying the effects of cyclosporin A and sirolimus on glucose and lipid metabolism in liver, skeletal muscle and adipose tissue in an in vivo rat model.

    PubMed

    Fuhrmann, A; Lopes, Pc; Sereno, J; Pedro, J; Espinoza, D O; Pereira, M J; Reis, F; Eriksson, J W; Carvalho, E

    2014-03-15

    Cyclosporin A (CsA) and sirolimus (SRL) are immunosuppressive agents (IAs) associated with dyslipidemia, insulin resistance and new onset diabetes after transplantation (NODAT). However, the molecular mechanisms involved are not fully understood. We investigated the effects of six-week treatment of either CsA or SRL on glucose and lipid metabolism in Wistar rats. The results show that, compared with vehicle-treated rats, SRL-treated rats were significantly lighter starting at week 5. CsA or SRL caused glucose intolerance, increased storage of lipids in the liver and skeletal muscle, and decreased the insulin-stimulated glucose uptake in isolated adipocytes. Furthermore, these agents significantly decreased genes involved in insulin action and glucose uptake, such as, IRS-1, Glut4 and Glut1, and increased genes and/or proteins involved in hepatic lipogenesis and gluconeogenesis, while decreasing them in adipose tissue. After either treatment PGC1α gene expression was down regulated in skeletal muscle, an important player in fatty acid oxidation. Moreover, there was an increase in IL-6 gene expression in adipose tissue in the SRL-treated rats, suggesting stimulation of lipolysis. The results of the present study suggest that CsA and SRL lead to metabolic alterations in liver, muscle and adipose tissue, which may contribute to the development of dyslipidemia and insulin resistance associated with immunosuppressive therapy. PMID:24462915

  12. 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.

  13. Nanoparticle-Mediated Targeting of Cyclosporine A Enhances Cardioprotection Against Ischemia-Reperfusion Injury Through Inhibition of Mitochondrial Permeability Transition Pore Opening

    PubMed Central

    Ikeda, Gentaro; Matoba, Tetsuya; Nakano, Yasuhiro; Nagaoka, Kazuhiro; Ishikita, Ayako; Nakano, Kaku; Funamoto, Daiki; Sunagawa, Kenji; Egashira, Kensuke

    2016-01-01

    Myocardial ischemia-reperfusion (IR) injury limits the therapeutic effects of early reperfusion therapy for acute myocardial infarction (MI), in which mitochondrial permeability transition pore (mPTP) opening plays a critical role. Our aim was to determine whether poly-lactic/glycolic acid (PLGA) nanoparticle-mediated mitochondrial targeting of a molecule that inhibits mPTP opening, cyclosporine A (CsA), enhances CsA-induced cardioprotection. In an in vivo murine IR model, intravenously injected PLGA nanoparticles were located at the IR myocardium mitochondria. Treatment with nanoparticles incorporated with CsA (CsA-NP) at the onset of reperfusion enhanced cardioprotection against IR injury by CsA alone (as indicated by the reduced MI size at a lower CsA concentration) through the inhibition of mPTP opening. Left ventricular remodeling was ameliorated 28 days after IR, but the treatment did not affect inflammatory monocyte recruitment to the IR heart. In cultured rat cardiomyocytes in vitro, mitochondrial PLGA nanoparticle-targeting was observed after the addition of hydrogen peroxide, which represents oxidative stress during IR, and was prevented by CsA. CsA-NP can be developed as an effective mPTP opening inhibitor and may protect organs from IR injury. PMID:26861678

  14. TMBIM6 (transmembrane BAX inhibitor motif containing 6) enhances autophagy and reduces renal dysfunction in a cyclosporine A-induced nephrotoxicity model.

    PubMed

    Yadav, Raj Kumar; Lee, Geum-Hwa; Lee, Hwa-Young; Li, Bo; Jung, Han-Eul; Rashid, Harun-Or; Choi, Min Kyung; Yadav, Binod Kumar; Kim, Woo-Ho; Kim, Kyung-Woon; Park, Byung-Hyun; Kim, Won; Lee, Yong-Chul; Kim, Hyung-Ryong; Chae, Han-Jung

    2015-01-01

    Cyclosporine A (CsA) is widely used as an immunosuppressor in transplantation. Previous studies reported that CsA induces autophagy and that chronic treatment with CsA results in accumulation of autophagosomes and reduced autophagic clearance. Autophagy is a prosurvival process that promotes recovery from acute kidney injury by degrading misfolded proteins produced in the kidney. In the present study, we used TMBIM6-expressing HK-2, human kidney tubular cells (TMBIM6 cells) and Tmbim6 knockout (tmbim6(-/-)) mice. When exposed to CsA, the TMBIM6 cells maintained autophagy activity by preventing autophagosome accumulation. With regard to signaling, PRKKA/AMPK phosphorylation and mechanistic target of rapamycin (serine/threonine kinase) complex 1 (MTORC1) expression and its downstream target TFEB (transcription factor EB), a lysosome biogenesis factor, were regulated in the TMBIM6 cells. Lysosomal activity was highly increased or stably maintained in the presence of TMBIM6. In addition, treatment of tmbim6(-/-) mice with CsA resulted in increased autophagosome formation and decreased lysosome formation and activity. We also found that tmbim6(-/-) mice were susceptible to CsA-induced kidney injury. Taken together, these results indicate that TMBIM6 protects against CsA-induced nephrotoxicity both in vitro and in vivo by inducing autophagy and activating lysosomes. PMID:26305401

  15. 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

  16. Cyclosporin A-sensitive induction of the Epstein-Barr virus lytic switch is mediated via a novel pathway involving a MEF2 family member.

    PubMed Central

    Liu, S; Liu, P; Borras, A; Chatila, T; Speck, S H

    1997-01-01

    Induction of the Epstein-Barr virus (EBV) lytic cycle by crosslinking surface immunoglobulin is inhibited by the immunosuppressants cyclosporin A (CsA) and FK506. This correlates with the ability of CsA to inhibit Ca2+-dependent transcription of the lytic cycle switch gene BZLF1. It is shown here that CsA sensitivity maps to three sites (ZIA, ZIB and ZID) that bind the serum response factor-related protein MEF2D. A synthetic promoter containing multiple copies of a MEF2D site from Zp, in conjunction with a CREB/AP-1 site (ZII) from Zp, exhibits CsA-sensitive inducibility. Furthermore, the Zp MEF2D sites were functionally interchangeable with MEF2 sites derived from heterologous promoters. While no evidence of a NFAT family member binding to either the MEF2 or CREB/AP-1 sites was obtained, it could be demonstrated that CsA-sensitive induction of Zp was mediated by calcineurin and NFATc2 in synergy with either phorbol ester or especially with the EBV-induced Ca2+/calmodulin-dependent kinase type IV/Gr. These studies identify Zp as prototypic of a novel class of CsA-sensitive and NFAT-dependent promoters defined by the presence of MEF2 sites. PMID:9009275

  17. Transfer of mesenchymal stem cells and cyclosporine A on alkali-injured rabbit cornea using nanofiber scaffolds strongly reduces corneal neovascularization and scar formation.

    PubMed

    Cejka, Cestmir; Cejkova, Jitka; Trosan, Peter; Zajicova, Alena; Sykova, Eva; Holan, Vladimir

    2016-09-01

    The aim of this study was to examine whether nanofiber scaffolds seeded with rabbit bone marrow mesenchymal stem cells (MSCs nanofibers) transferred onto the damaged corneal surface and covered with cyclosporine A (CsA)-loaded nanofiber scaffolds (CsA nanofibers) enable healing of the rabbit cornea injured with 1N NaOH. The healing of damaged corneas was examined morphologically, immunohistochemically and biochemically on day 24 after the injury. Compared to untreated injured corneas, where corneal ulceration or large corneal thinning or even perforation were developed, injured corneas treated with drug free nanofibers healed without profound disturbances in a majority of cases, although with fibrosis and scar formation. In injured corneas treated with CsA nanofibers or MSCs nanofibers, the development of scar formation was reduced. Best healing results were obtained with a combination of MSCs and CsA nanofibers (MSCs-CsA nanofibers). Corneas healed with highly restored transparency. Neovascularization highly expressed in untreated injured corneas and reduced in corneas treated with CsA nanofibers or MSCs nanofibers, was suppressed in corneas treated with MSCs-CsA nanofibers. The levels of matrix metalloproteinase 9, inducible nitric oxide synthase, interleukin 6, α-smooth muscle actin, tumor growth factor β and vascular endothelial growth factor were significantly decreased in these corneas as compared to untreated corneas, where the levels of the above mentioned markers were high. In conclusion, MSCs-CsA nanofibers were effective in the treatment of severe alkali-induced corneal injury. PMID:26797822

  18. Interleukin-1beta partially alleviates cyclosporin A-induced suppression of IgG1 isotype response to thyroglobulin in BALB/c mice in vivo.

    PubMed Central

    Dalai, S K; Miriyala, B; Kar, S K

    1998-01-01

    Cyclosporin A (CsA) at 120 mg/kg body weight when injected subcutaneously into BALB/c mice along with thyroglobulin emulsified in incomplete Freund's adjuvant (IFA) was found to suppress antigen-specific IgG titre by 86%. Isotyping revealed that both IgG1 and IgG2a titres were suppressed by 87% and 57%, respectively. But under identical conditions when complete Freund's adjuvant (CFA) was used, the suppression of antigen-specific IgG, IgG1 and IgG2a titres was 50%, 51% and 55%, respectively. Injection of anti-IL-1beta-neutralizing hamster monoclonal antibodies along with thyroglobulin and CsA emulsified in CFA increased the suppression of antigen-specific IgG titre. Under such conditions the IgG1 titre was suppressed more than the IgG2a titre. Recombinant human interleukin-1 receptor antagonist (rhuIL-1ra) also enhanced the suppression caused by CsA in the presence of CFA but control hamster immunoglobulin had no such effect. Recombinant human IL-1beta, when administered along with thyroglobulin and CsA emulsified in IFA, alleviated the suppression of antigen-specific IgG titre and the IgG1 titre was alleviated more than the IgG2a titre. Under identical conditions, rhuIL-1ra did not alleviate CsA-induced suppression. Lymphocytes from the lymph nodes of thyroglobulin-sensitized BALB/c mice when stimulated in vitro by thyroglobulin in the presence of CsA, secreted very little interferon-gamma (IFN-gamma) and IL-4, but on addition of an optimal dose of rhuIL-1beta, IFN-gamma and IL-4 secretion was partially restored. PMID:9767461

  19. 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

  20. The Crystal Structure of PPIL1 Bound to Cyclosporine A Suggests a Binding Mode for a Linear Epitope of the SKIP Protein

    PubMed Central

    Stegmann, Christian M.; Lührmann, Reinhard; Wahl, Markus C.

    2010-01-01

    Background The removal of introns from pre-mRNA is carried out by a large macromolecular machine called the spliceosome. The peptidyl-prolyl cis/trans isomerase PPIL1 is a component of the human spliceosome and binds to the spliceosomal SKIP protein via a binding site distinct from its active site. Principal Findings Here, we have studied the PPIL1 protein and its interaction with SKIP biochemically and by X-ray crystallography. A minimal linear binding epitope derived from the SKIP protein could be determined using a peptide array. A 36-residue region of SKIP centred on an eight-residue epitope suffices to bind PPIL1 in pull-down experiments. The crystal structure of PPIL1 in complex with the inhibitor cyclosporine A (CsA) was obtained at a resolution of 1.15 Å and exhibited two bound Cd2+ ions that enabled SAD phasing. PPIL1 residues that have previously been implicated in binding of SKIP are involved in the coordination of Cd2+ ions in the present crystal structure. Employing the present crystal structure, the determined minimal binding epitope and previously published NMR data [1], a molecular docking study was performed. In the docked model of the PPIL1·SKIP interaction, a proline residue of SKIP is buried in a hydrophobic pocket of PPIL1. This hydrophobic contact is encircled by several hydrogen bonds between the SKIP peptide and PPIL1. Conclusion We characterized a short, linear epitope of SKIP that is sufficient to bind the PPIL1 protein. Our data indicate that this SKIP peptide could function in recruiting PPIL1 into the core of the spliceosome. We present a molecular model for the binding mode of SKIP to PPIL1 which emphasizes the versatility of cyclophilin-type PPIases to engage in additional interactions with other proteins apart from active site contacts despite their limited surface area. PMID:20368803

  1. Suppression of adult T cell leukemia-derived factor/human thioredoxin induction by FK506 and cyclosporin A: a new mechanism of immune modulation via redox control.

    PubMed

    Furuke, K; Nakamura, H; Hori, T; Iwata, S; Maekawa, N; Inamoto, T; Yamaoka, Y; Yodoi, J

    1995-06-01

    Adult T cell leukemia-derived factor (ADF), which is identical to a disulfide reducing enzyme human thioredoxin (TRX), is produced and released by activated or virus-infected lymphocytes. Here we report that, in peripheral blood mononuclear cells (PBMC) stimulated with phytohemagglutinin (PHA), ADF/TRX mRNA was induced within 8 h after stimulation as detected by in situ hybridization study. To analyze the mechanism of ADF/TRX induction during T cell activation, the effects of immunosuppressants including FK506, rapamycin (Rap) and cyclosporin A (CsA) on ADF/TRX expression were investigated by immunoblot analysis. ADF/TRX induction in PBMC by PHA, Con A or OKT3 mAb was almost completely suppressed by FK506. Whereas CsA also inhibited ADF/TRX expression in OKT3 mAb-stimulated PBMC, Rap failed to affect it in spite of exhibiting growth inhibition. In addition, exogenous IL-2 could not increase ADF/TRX production in FK506-treated PBMC or in PHA blasts. These results indicate that ADF/TRX induction in T cell activation depends on calcineurin-dependent events in the early phase and that IL-2 production is not directly involved in ADF/TRX induction. Furthermore, when recombinant ADF (rADF) was added to a culture of PBMC 1 h before the addition of PHA and FK506, the action of FK506 was partially reversed as determined by [3H]thymidine incorporation and viable cell counts. These results suggest that ADF/TRX produced and released from PBMC may be a crucial event in lymphocyte activation, and that FK506 and CsA may exert the immune suppression partly through inhibiting the induction of the endogenous reducing factor ADF/TRX. PMID:7577807

  2. Discovery of cyclosporine A and its analogs as broad-spectrum anti-influenza drugs with a high in vitro genetic barrier of drug resistance.

    PubMed

    Ma, Chunlong; Li, Fang; Musharrafieh, Rami Ghassan; Wang, Jun

    2016-09-01

    As the number of drug-resistant influenza viruses continues to increase, antivirals with novel mechanisms of action are urgently needed. Among the two classes of FDA-approved antiviral drugs, neuraminidase (NA) inhibitors, oseltamivir, zanamivir, and peramivir, are currently the only choice for the prevention and treatment of influenza virus infection. Due to the antigenic drift and antigenic shift, it will only be a matter of time before influenza viruses become completely resistant to these NA inhibitors. In pursuing the next generation of antiviral drugs with complementary mechanisms of action to those of the NA inhibitors, we have identified a natural product, cyclosporine A (CsA) (1), as a desired drug candidate. In this study, we discovered that CsA (1) and its analogs have broad-spectrum antiviral activity against multiple influenza A and B strains, including strains that are resistant to either NA or M2 inhibitors or both. Moreover, CsA (1) displays a high in vitro genetic barrier of drug resistance than oseltamivir carboxylate Mechanistic studies revealed that CsA (1) acts at the intermediate step of viral replication post viral fusion. Its antiviral mechanism is independent of inhibiting the isomerase activity of cyclophilin A (CypA), and CsA (1) has no effect on the viral polymerase activity The potent antiviral efficacy of CsA (1), coupled with the high in vitro genetic barrier of drug resistance and novel mechanism of action, renders CsA (1) a promising anti-influenza drug candidate for further development. PMID:27478032

  3. PEG-PE-based micelles co-loaded with paclitaxel and cyclosporine A or loaded with paclitaxel and targeted by anticancer antibody overcome drug resistance in cancer cells.

    PubMed

    Sarisozen, Can; Vural, Imran; Levchenko, Tatyana; Hincal, A Atilla; Torchilin, Vladimir P

    2012-05-01

    The over-expression of the P-glycoprotein (P-gp) in cancer cells is one of the main reasons of the acquired Multidrug Resistance (MDR). Combined treatment of MDR cancer cells with P-gp inhibitors and chemotherapeutic agents could result in reversal of resistance in P-gp-expressing cells. In this study, paclitaxel (PTX) was co-encapsulated in actively targeted (anticancer mAb 2C5-modified) polymeric lipid-core PEG-PE-based micelles with Cyclosporine A (CycA), which is one of the most effective first generation P-gp inhibitors. Cell culture studies performed using MDCKII (parental and MDR1) cell lines to investigate the potential MDR reversal effect of the formulations. The average size of both empty and loaded PEG₂₀₀₀-PE/Vitamin E mixed micelles was found between 10 and 25 nm. Zeta potentials of the formulations were found between -7 and -35 mV. The percentage of PTX in the micelles was found higher than 3% for both formulations and cumulative PTX release of about 70% was demonstrated. P-gp inhibition with CycA caused an increase in the cytotoxicity of PTX. Dual-loaded micelles demonstrated significantly higher cytotoxicity in the resistant MDCKII-MDR1 cells than micelles loaded with PTX alone. Micelle modification with mAb 2C5 results in the highest cytotoxicity against resistant cells, with or without P-gp modulator, probably because of better internalization bypassing the P-gp mechanism. Our results suggest that micelles delivering a combination of P-gp modulator and anticancer drug or micelles loaded with only PTX, but targeted with mAb 2C5 represent a promising approach to overcome drug resistance in cancer cells. PMID:22506922

  4. 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

  5. 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

  6. The effect of cyclosporin-A on peri-operative myocardial injury in adult patients undergoing coronary artery bypass graft surgery: a randomised controlled clinical trial

    PubMed Central

    Hausenloy, DJ; Kunst, G; Boston-Griffiths, E; Kolvekar, S; Chaubey, S; John, L; Desai, J; Yellon, DM

    2014-01-01

    Objective Cyclosporin-A (CsA) has been reported to reduce myocardial infarct size in both the experimental and clinical settings. This protective effect is dependent on its ability to prevent the opening of the mitochondrial permeability transition pore, a critical determinant of cell death in the setting of acute ischaemia-reperfusion injury. Whether CsA can reduce the extent of peri-operative myocardial injury (PMI) in patients undergoing coronary artery bypass graft (CABG) surgery is unknown, and is investigated in this randomised controlled clinical trial. Methods 78 adult patients undergoing elective CABG surgery were randomised to receive either an intravenous bolus of CsA (2.5 mg/kg) or placebo administered after induction of anaesthesia and prior to sternotomy. PMI was assessed by measuring serum cardiac enzymes, troponin T (cTnT) and CK-MB at 0, 6, 12, 24, 48 and 72 h after surgery. Results There was no significant difference in mean peak cTnT levels between control (n=43) and CsA treatment (n=40) patients (0.56±0.06 ng/mL with control vs 0.35±0.05 ng/mL with CsA; p=0.07). However, in higher-risk patients with longer cardiopulmonary bypass times, there was a significant reduction in PMI with CsA therapy (p=0.049), with a reduced postoperative cTnT rise by 0.03 ng/mL for every 10 min, when compared with control. Conclusions In patients with longer cardiopulmonary bypass times, a single intravenous bolus of CsA administered prior to CABG surgery reduced the extent of PMI. PMID:24488610

  7. Association of CD14-260 Polymorphisms, Red-complex Periodontopathogens and Gingival Crevicular Fluid Cytokine Levels with Cyclosporine A-induced Gingival Overgrowth in Renal Transplant Patients

    PubMed Central

    Gong, Yiming; Bi, Wei; Cao, Lingfeng; Yang, Yi; Chen, Jake; Yu, Youcheng

    2012-01-01

    Backgroud and Objective Genetic factors may influence the colonization of pathogenic bacteria, therefore increasing the risk for the initiation and development of periodontal disease. The present study was carried out to investigate the association of CD14-260 polymorphisms, subgingival microbiota, and gingival crevicular fluid (GCF) cytokine levels with cyclosporine A (CsA)-induced gingival overgrowth (GO) in renal transplant patients. Material and Methods 204 patients were dichotomized into two groups: 124 with GO and 80 without GO. The CD14-260 polymorphisms were measured using an allele-specific PCR method. The levels of periodontal pathogens were determined by real-time PCR of subgingival samples. GCF levels of IL-1β and sCD14 were detected by ELISA. Results The frequency of CD14-260 genotype CT + TT was found to be similar in both groups. Patients with GO presented increased prevalence of Pg, Td, and Tf (red complex) and significantly higher levels of IL-1β than those without GO. GO patients carrying CT+TT genotypes were found to have higher frequencies of Pg, Td, and Tf than those carrying CC genotype. Furthermore, in the presence of red complex, CT+TT genotypes were associated with higher IL-1β levels and severe GO. Multiple logistic regression analysis demonstrated that the severity of GO is not dependent on age, gender and pharmacological variables, being only associated with CD14-260 genotype and red complex periodontopathogens. Conclusions No association between CD14-260 polymorphisms and the prevalence of GO was revealed in renal transplant patients administered CsA. However, CD14-260 CT+TT genotypes are associated with the prevalence of red complex periodontopathogens in GO patients, and may thus play some role in the development of severe CsA-induced GO. PMID:22934794

  8. Endoplasmic reticulum vacuolation and unfolded protein response leading to paraptosis like cell death in cyclosporine A treated cancer cervix cells is mediated by cyclophilin B inhibition.

    PubMed

    Ram, Babul Moni; Ramakrishna, Gayatri

    2014-11-01

    Cyclosporine A (CsA), a widely used immunosuppressant shows cytotoxic effects by either inducing apoptosis or redirecting the cell towards non-apoptotic cell death. However, there still remains a lacuna in understanding the mechanism of CsA induced non-apoptotic cell death. In the present study we investigated calcineurin dependent or independent cytotoxic effects of CsA, a calcineurin inhibitor, in cervical cancerous SiHa cells. Decreased cell viability and massive cytoplasmic vacuolations were observed in CsA treated SiHa cells, having increased calcineurin activity. Endoplasmic reticulum (ER) stress and unfolded protein response (UPR), accompanied by a decrease in cyclophilin B (ER resident PPIase), preceded the formation of the vacuoles. These vacuoles stained positive for many ER resident markers confirming their ER origin; but the absence of autophagosomal marker, LC3II, ruled out autophagy. Extensively vacuolated cells eventually undergo cell death which lacked the typical apoptotic features, but showed significant decrease in AIP (ALG2 interacting protein) as seen in paraptosis. ER-vacuolation was prevented by cycloheximide and salubrinal thereby indicating requirement of active protein synthesis. Inhibiting calcineurin activity by either Tacrolimus (FK506) or by knockdown of calcineurin B subunit did not result in either ER-stress or cellular vacuolation. However, knockdown of cyclophilin B by siRNA resulted in increased expression of Bip and IRE1α, together with cytoplasmic vacuolation. In conclusion, we report that persistent ER stress due to cyclophilin B inhibition in CsA treated cervical cancer cells caused cellular vacuolation which culminated in a non-apoptotic cell death response similar to paraptosis. Additionally, the paraptotic effects of CsA are independent of calcineurin inhibition. PMID:25003316

  9. 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.

  10. In vivo microdialysis with LC-MS for analysis of spinosin and its interaction with cyclosporin A in rat brain, blood and bile.

    PubMed

    Ma, Rong-Hua; Yang, Jie; Qi, Lian-Wen; Xin, Gui-Zhong; Wang, Chong-Zhi; Yuan, Chun-Su; Wen, Xiao-Dong; Li, Ping

    2012-03-01

    Spinosin, a major bioactive herbal ingredient isolated from Semen Ziziphi Spinosae, plays an important role in sedation and hypnosis. However, the pharmacokinetic behavior of spinosin in special sites has not been reported. Microdialysis (MD) technique, as a continuous, realtime monitoring sampling technique, is very suitable for the evaluation of the disposition of diverse drugs. To obtain more useful information on spinosin, an in vivo microdialysis sampling technique with High Performance Liquid Chromatography-mass spectrograph (HPLC-MS) method was developed to investigate the pharmacokinetics of spinosin and its interaction with cyclosporin A (CsA) in the brain, blood and bile of rats. The method was validated in terms of selectivity, linearity and sensitivity, and showed advantages in monitoring the pharmacokinetic behavior of drugs. The results revealed that CsA has obvious effects on the pharmacokinetic process of spinosin. When co-administered, the area under the curve (AUC) of spinosin in blood, bile and brain increased from 205.70 to 673.51 mg min/L, 7.77 × 10(4) to 1.25 × 10(5) mg min/L, and 2.09 to 5.58 mg min/L, respectively. The t(1/2) values of spinosin in blood, bile and brain also changed from 48.07 to 95.04 min, from 97.20 to 152.21 and from 42.18 to 73.83 min, respectively. These results demonstrated that the CsA decreased the efflux of spinosin through the inhibition of P-glycoprotein (P-gp) efflux transporter and it might be used as a group of P-gp substrate. Other transporters or pathways may also be involved in the metabolism of spinosin. PMID:22169469

  11. Sirolimus and cyclosporine A alter barrier function in renal proximal tubular cells through stimulation of ERK1/2 signaling and claudin-1 expression.

    PubMed

    Martin-Martin, Natalia; Ryan, Gavin; McMorrow, Tara; Ryan, Michael P

    2010-03-01

    Alteration of the tight junction complex in renal epithelial cells can affect renal barrier function and perturb normal kidney homeostasis. The immunosuppressant drugs cyclosporine A (CsA) and sirolimus (SRL) used in combination demonstrated beneficial effects in organ transplantation but this combination can also result in increased adverse effects. We previously showed that CsA treatment alone caused an alteration of the tight junction complex, resulting in changes in transepithelial permeability in Madin-Darby canine kidney distal tubular/collecting duct cells. The potential effect of SRL on transepithelial permeability in kidney cells is unknown. In this study, subcytotoxic doses of SRL or CsA were found to decrease the paracellular permeability of the porcine proximal tubular epithelial cells, LLC-PK1 cell monolayers, which was detected as an increase in transepithelial electrical resistance (TER). The cotreatment with SRL and CsA was found to increase TER in a synergistic manner. CsA treatment increased total cellular expression and membrane localization of the tight junction protein claudin-1 and this further increased with the combination of SRL/CsA. SRL and CsA treatment alone or in combination stimulated the phosphorylation of ERK1/2. The MEK-ERK1/2 pathway inhibitor, U0126, reduced the SRL, CsA, and CsA/SRL-induced increase in TER. U0126 also reduced the CsA and CsA/SRL-induced increase in the membrane localization of claudin-1. Alterations in claudin-2 and claudin-4 were also detected. However, the results suggest that the modulation in expression and localization of claudin-1 appears to be pivotal in the SRL- and CsA-induced modulation of the epithelial barrier function and that modulation is regulated by ERK1/2 signaling pathway. PMID:19955189

  12. A step-wise expansion of intestinal intraepithelial T lymphocytes in association with microbial colonization is defined by sensitivity to cyclosporin A.

    PubMed Central

    Kawaguchi-Miyashita, M; Nanno, M; Shimada, S; Nagaoka, N; Okada, Y; Matsumoto, S; Umesaki, Y; Matsuoka, Y; Ohwaki, M

    1997-01-01

    Murine intestinal intraepithelial lymphocytes (IELs) consist of T cells bearing alpha beta-antigen receptor (alpha beta-IELs) and those bearing gamma delta-IELs). Although gamma delta-IELs outnumber alpha beta-IELs in germ-free (GF) mice, oral inoculation of fecal suspension from conventional (CV) mice into GF mice induced the increase in number of alpha beta-IELs, leaving the number of gamma delta-IELs unchanged, and the number of alpha beta-IELs reached the level of CV mice by 3 weeks after conventionalization. Expansion of alpha beta-IELs and increase in their CD44+ subset in conventionalized mice were not affected until 2 weeks after beginning of daily injection of cyclosporin A (CsA). However, further expansion of alpha beta-IELs during 2-3 weeks after conventionalization was blocked by injection of CsA. Although the relative constitution of CD4- 8-, CD4+ 8-, CD4- 8 alpha alpha+, CD4- 8 alpha beta+ and CD4+ 8+ subsets among alpha beta-IELs was comparable between control and CsA-treated groups, CsA injection resulted in the decrease in ratio of high-density fraction cells to low density fraction cells in IELs. CsA completely abrogated the expansion of T cells in peripheral lymph nodes stimulated by alloantigens in vivo, and proliferation of IELs from GF mice induced by immobilized anti-alpha beta-T-cell receptor (TCR) monoclonal antibodies (mAb) in vitro was also eliminated by CsA. These results indicate that microbial colonization-induced expansion of alpha beta-IELs is subdivided into two steps: the early phase of expansion takes place via TCR-non-mediated pathway and the late phase of expansion requires TCR-mediated signal transduction. Images Figure 4 PMID:9378505

  13. 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

  14. Inter-laboratory comparison of human renal proximal tubule (HK-2) transcriptome alterations due to Cyclosporine A exposure and medium exhaustion.

    PubMed

    Jennings, Paul; Aydin, Sonia; Bennett, Jason; McBride, Rachael; Weiland, Christina; Tuite, Niamh; Gruber, Leonhard N; Perco, Paul; Gaora, Peadar O; Ellinger-Ziegelbauer, Heidrun; Ahr, Hans-Juergen; Kooten, Cees Van; Daha, Mohamed R; Prieto, Pilar; Ryan, Michael P; Pfaller, Walter; McMorrow, Tara

    2009-04-01

    There is an acknowledged need to promote and further develop in vitro techniques in order to achieve the goal of improved risk assessment of chemicals and pharmaceuticals to humans. The EU 6th framework project "PREDICTOMICS" was established in order to contribute to the further development of in vitro toxicology, with a particular focus on emerging techniques including toxicogenomics. DNA microarray technology is being used more frequently in the in vitro field, however, only very few studies have assessed the reproducibility of this technique with respect to in vitro toxicology. To this end we conducted an interlaboratory comparison to test the reproducibility of transcriptomic changes induced by the immunosuppressive agent, Cyclosporine A (CsA) on the human renal proximal tubular cell line, HK-2 cell. Four European laboratories took part in this study. Under standardised conditions, each laboratory treated HK-2 cells with 5microM CsA for 12 and 48h. RNA was isolated and hybridised to Affymetrix HGU-133 plus two arrays at three different sites. Analysis of the transcription profiles demonstrated that one laboratory clustered away from the other laboratories, potentially due to an inclusion of a trypsinisation step by this laboratory. Once the genes responsible for this separate clustering were removed all laboratories showed similar expression profiles. There was a major impact of time since feed, due to medium exhaustion in the 48h arrays compared to the 12h arrays, regardless of CsA treatment. Biological processes including general vesicle transport, amino acid metabolism, amino acid transport and amino acid biosynthesis were over-represented due to time since feed, while cell cycle, DNA replication, mitosis and DNA metabolism were under-represented. CsA responsive genes were involved in cell cycle, the p53 pathway and Wnt signaling. Additionally there was an overlap of differentially expressed genes due to CsA and medium exhaustion which is most likely due to

  15. Addition of cyclosporin-A to chemotherapy in secondary (post-MDS) AML in the elderly. A multicenter randomized trial of the Leukemia Working Group of the Hellenic Society of Hematology.

    PubMed

    Matsouka, P; Pagoni, M; Zikos, P; Giannakoulas, N; Apostolidis, I; Asprogeraka, T; Arvanitopoulou, E; Spanoudakis, E; Kotsianidis, I; Tsatalas, K; Papaioannou, M; Marinakis, T; Skandali, A; Viniou, N; Yataganas, X; Bakiri, M

    2006-04-01

    In elderly patients with secondary leukemia, poor therapeutic response and low overall survival have been attributed mainly to age and to the primary resistance of leukemic cells to chemotherapy. Modulation of resistance has been attempted in different studies, but the results have been contradictory. We conducted an open, randomized multicenter clinical trial involving patients more than 60 years old with secondary leukemia preceded by a myelodysplastic syndrome. The induction chemotherapy regimen included idarubicin, cytarabine, and etoposide (group A); randomization involved simultaneous administration of cyclosporin-A per os (group B). Fifty-five patients were evaluated, 26 in group A and 29 in group B. Overall complete remission was achieved in 40% of the patients, 27% vs 52% in groups A and B, respectively (p=0.01). Leukemia-free survival was more favorable in patients who received cyclosporin-A, 12 vs 7 months for groups B and A, respectively (p=0.03). In a follow up period of 30 months, 7 out of 55 patients (13%) were alive, 4 of whom were in complete remission. Five out of the 7 alive patients were randomized in group B and had received cyclosporin-A. Treatment failure was higher in group A [19 of 26 patients (73%)] than in group B with CsA [14 of 29 patients (48%)] (p<0.0001). Treatment-related toxicity/mortality was 13%. Modulation of drug resistance by CsA in elderly people suffering from secondary acute leukemia may improve the outcome of chemotherapy without increasing drug toxicity and treatment-related mortality.

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

    PubMed Central

    Debouzy, Jean-Claude; 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 1H-NMR in solution and its membrane interactions were studied by 1H-NMR in small unilamellar vesicles and by 31P 2H NMR in phospholipidic dispersions of DMPC (dimyristoylphosphatidylcholine) in comparison with those of POLYA and CYSP alone. 1H-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 (31P). 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. PMID:24883210

  17. Cyclosporine A prevents the generation of single positive (Lyt2+ L3T4-, Lyt2- L3T4+) mature T cells, but not single positive (Lyt2+ T3-) Immature thymocytes, in newborn mice.

    PubMed

    Heeg, K; Bendigs, S; Wagner, H

    1989-12-01

    The influence of cyclosporine A (CsA) on T-cell maturation was investigated in newborn mice. CsA treatment during the pre- and postnatal periods resulted in a hypoplasia of peripheral lymphatic organs, and absence of mature T3+ T cells in lymph nodes and spleens; no functional T-cell reactivity was observed. In thymuses of CsA-treated mice, no T3+ single positive Lyt2+ or T3+L3T4+ thymocytes could be found, but double positive (DP) cells were readily detected. A thymocyte subset with the phenotype Lyt2+L3T4-T3- was still discernible; this population was non-functional in vitro. The data show that the maturation of single positive (SP) T cells is critically influenced by CsA; under the conditions used here we found no evidence that 'leaky' autoreactive SP T cells develop in CsA-treated newborn mice.

  18. Downregulation of CYP3A and P-glycoprotein in the secondary inflammatory response of mice with dextran sulfate sodium-induced colitis and its contribution to cyclosporine A blood concentrations.

    PubMed

    Kawauchi, Shoji; Nakamura, Tsutomu; Miki, Ikuya; Inoue, Jun; Hamaguchi, Tsuneo; Tanahashi, Toshihito; Mizuno, Shigeto

    2014-01-01

    CYP3A and P-glycoprotein (P-gp) play important roles in drug metabolism and excretion; however, their functions in pathological conditions remain unclear. Hepatobiliary abnormalities have been described in patients with ulcerative colitis, which may affect drug metabolism and excretion in the liver and small intestine. We examined the functions of CYP3A and P-gp in the liver and small intestine of mice with dextran sodium sulfate (DSS)-induced colitis. Up to day 7, inflammatory markers were significantly increased in the livers of DSS-treated mice, accompanied by decreased CYP3A. Additionally hepatobiliary transporters and Pregnane X receptor, which regulates the transcriptional activation of CYP3A, were reduced. Both CYP3A and P-gp were significantly decreased in the upper small intestine of DSS-treated mice on day 7. This was associated with the increased expression of inducible nitric oxide synthase, but not changes in nuclear receptor expression. On day 7 of DSS treatment, the concentrations of cyclosporine A (CsA), a substrate of both CYP3A and P-gp, were significantly higher than controls. These results indicated the existence of a second inflammatory response in the liver and upper small intestine of mice with DSS-induced colitis, and bioavailability of CsA was increased by the dysfunction of CYP3A and P-gp in these organs.

  19. 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

  20. 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.

  1. Study protocol for a phase III multicentre, randomised, open-label, blinded-end point trial to evaluate the efficacy and safety of immunoglobulin plus cyclosporin A in patients with severe Kawasaki disease (KAICA Trial)

    PubMed Central

    Aoyagi, Reiko; Hamada, Hiromichi; Sato, Yasunori; Suzuki, Hiroyuki; Onouchi, Yoshihiro; Ebata, Ryota; Terauchi, Moe; Terai, Masaru; Hanaoka, Hideki; Hata, Akira

    2015-01-01

    Introduction Kawasaki disease (KD) is an acute, self-limited vasculitis of unknown aetiology that predominantly affects infants and young children. We hypothesise that cyclosporin A (CsA) may be effective in treating KD by regulating the Ca2+/NFAT signalling pathway. This trial compares the current standard therapy of intravenous immunoglobulin (IVIG) and the combined IVIG+CsA therapy in paediatric patients with severe KD. Methods and analysis This trial is a phase III, multicentre, randomised, open-label, blinded-end point trial that evaluates the efficacy and safety of IVIG+CsA therapy. Patients with severe KD who satisfy the eligibility criteria are randomised (1:1) to receive either CsA (5 mg/kg/day for 5 days; Neoral) plus high-dose IVIG (2 g/kg for 24 h and aspirin 30 mg/kg/day), or high-dose IVIG alone (2 g/kg for 24 h and aspirin 30 mg/kg/day). The primary end point is the frequency of occurrence of coronary artery abnormalities during the trial period. An independent end point review committee will be in charge of the trial assessment. Ethics and dissemination The protocol was approved by the Institutional Review Board of each institution. The trial was notified and registered at the Pharmaceutical and Medical Devices Agency, in Japan. The trial is currently on-going and is scheduled to finish in April 2017. The findings will be disseminated through peer-reviewed publications and conference presentations. Trial registration number JMA-IIA00174; Pre-results. PMID:26628527

  2. CNS involvement occurs more frequently in patients with Behçet's disease under cyclosporin A (CSA) than under other medications--results of a retrospective analysis of 117 cases.

    PubMed

    Kötter, Ina; Günaydin, Ilhan; Batra, Marion; Vonthein, Reinhard; Stübiger, Nicole; Fierlbeck, Gerhard; Melms, Arthur

    2006-07-01

    The aim of this study was to evaluate the incidence of neurological manifestations of Behçet's disease (BD) in patients on cyclosporin A (CSA) compared with those on other medications. The records of 117 patients with BD who visited our hospital between 1990 and 2003 were reviewed with respect to symptoms and medication. All episodes of constant therapy prior to central nervous system (CNS) involvement were counted, and then the associations were analysed by the exact Fisher-Freeman-Halton test and adjusted for multiple tests by the Bonferroni-Holm method. We observed ten new cases of CNS manifestations in our patients with BD being regularly seen and treated in our tertiary care centre. The overall prevalence of neuro-BD in our patient group was 8.5%. In a retrospective analysis, the incidence of new-onset neurological disease (neuro-BD) in all patients with BD who regularly visited our hospital was significantly higher in patients on CSA than in those on other medications (6 of 21 vs 0 of 175 episodes, P<0.0001). This contrasts the obvious efficacy of CSA on extracerebral manifestations of BD, such as severe ocular disease, mucocutaneous lesions or arthritis. CSA exerts differential efficacy on various manifestations of BD. It is very effective for severe ocular and other moderate to severe manifestations of BD, but its efficacy for the prevention of neuro-BD seems to be inferior to that of other medications used in BD, such as azathioprine or interferon-alpha. The reasons for this are unclear, but the potential toxic effects of CSA on the CNS may be a predisposing factor for CNS vasculitis in BD.

  3. Safety and effects on the lipid and C-reactive protein plasma concentration of the association of ezetimibe plus atorvastatin in renal transplant patients treated by cyclosporine-A: a pilot study.

    PubMed

    Panichi, V; Manca-Rizza, G; Paoletti, S; Taccola, D; Consani, C; Sbragia, G; Mantuano, E; Marchetti, V; Carpi, A; Barsotti, G

    2006-06-01

    Ezetimibe (E) is a new cholesterol adsorption inhibitor which prevents the adsorption of dietary and biliary cholesterol by binding to a recently described cholesterol transporter. This pilot study was performed to evaluate the safety and the low-density lipoprotein (LDL)-C and C-reactive protein lowering efficacy of atorvastatin (A) and of the association of A plus E in five renal transplant patients with hypercholesterolemia and mild renal functional impairment receiving cyclosporine-A (CsA). Patients received for three periods, each of 3 weeks, A at a dose of 20 mg/day; A at a dose of 10 mg/day and finally, A 10 mg plus E 10 mg daily. The medications were well-tolerated and no important clinical or laboratory (muscle enzyme, creatinine clearance and CsA concentration) abnormalities were observed throughout the study period. A alone lead to target LDL-C values only in two of five patients and did not significantly reduce the mean CRP values. The combination of E plus A produced the lowest lipid levels and significantly reduced CRP mean values and allowed all patients to attain target levels of LDL-C: total cholesterol decreased from 240 +/- 42 (mean +/- S.D.) to 171 +/- 34 mg/dl, LDL-C from 129 +/- 32 to 87 +/- 21 mg/dl, plasma triglycerides from 330 +/- 54 to 194 +/- 71 mg/dl and CRP from 6.2 +/- 1.9 to 3.9 +/- 2.4 mg/l (P < 0.05 for all). This pilot study suggests that the co-administration of E and A at 10 mg/day in renal transplant patients receiving CsA is well-tolerated and effective in reducing important cardiovascular risk factors.

  4. Bax-mediated mitochondrial outer membrane permeabilization (MOMP), distinct from the mitochondrial permeability transition, is a key mechanism in diclofenac-induced hepatocyte injury: Multiple protective roles of cyclosporin A

    SciTech Connect

    Siu, W.P.; Pun, Pamela Boon Li; Latchoumycandane, Calivarathan; Boelsterli, Urs A.

    2008-03-15

    Diclofenac, a widely used nonsteroidal anti-inflammatory drug, has been associated with rare but severe cases of clinical hepatotoxicity. Diclofenac causes concentration-dependent cell death in human hepatocytes (after 24-48 h) by mitochondrial permeabilization via poorly defined mechanisms. To explore whether the cyclophilin D (CyD)-dependent mitochondrial permeability transition (mPT) and/or the mitochondrial outer membrane permeabilization (MOMP) was primarily involved in mediating cell death, we exposed immortalized human hepatocytes (HC-04) to apoptogenic concentrations of diclofenac (> 500 {mu}M) in the presence or absence of inhibitors of upstream mediators. The CyD inhibitor, cyclosporin A (CsA, 2 {mu}M) fully inhibited diclofenac-induced cell injury, suggesting that mPT was involved. However, CyD gene silencing using siRNA left the cells susceptible to diclofenac toxicity, and CsA still protected the CyD-negative cells from lethal injury. Diclofenac induced early (9 h) activation of Bax and Bak and caused mitochondrial translocation of Bax, indicating that MOMP was involved in cell death. Inhibition of Bax protein expression by using siRNA significantly protected HC-04 from diclofenac-induced cell injury. Diclofenac also induced early Bid activation (tBid formation, 6 h), which is an upstream mechanism that initiates Bax activation and mitochondrial translocation. Bid activation was sensitive to the Ca{sup 2+} chelator, BAPTA. In conclusion, we found that Bax/Bak-mediated MOMP is a key mechanism of diclofenac-induced lethal cell injury in human hepatocytes, and that CsA can prevent MOMP through inhibition of Bax activation. These data support our concept that the Ca{sup 2+}-Bid-Bax-MOMP axis is a critical pathway in diclofenac (metabolite)-induced hepatocyte injury.

  5. 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

    2012-09-01

    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.

  6. 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

  7. 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

    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

  8. Imaging the impact of cyclosporin A and dipyridamole on P-glycoprotein (ABCB1) function at the blood-brain barrier: A [(11)C]-N-desmethyl-loperamide PET study in nonhuman primates.

    PubMed

    Damont, Annelaure; Goutal, Sébastien; Auvity, Sylvain; Valette, Héric; Kuhnast, Bertrand; Saba, Wadad; Tournier, Nicolas

    2016-08-25

    Cyclosporin A (CsA) and dipyridamole (DPy) are potent inhibitors of the P-glycoprotein (P-gp; ABCB1) in vitro. Their efficacy at inhibiting P-gp at the blood-brain barrier (BBB) is difficult to predict. Efficient and readily available (i.e. marketed) P-gp inhibitors are needed as probes to investigate the role of P-gp at the human BBB. In this study, the P-gp inhibition potency at the BBB of therapeutic doses of CsA or DPy was evaluated in baboons using Positron Emission Tomography (PET) imaging with [(11)C]-N-desmethyl-loperamide ([(11)C]dLop), a radiolabeled P-gp substrate. The preparation of dLop as authentic standard and [(11)C]dLop as radiotracer were revisited so as to improve their production yields. [(11)C]dLop PET imaging was performed in the absence (n=3, baseline condition) and the presence of CsA (15mg/kg/h i.v., n=3). Three animals were injected with i.v. DPy at either 0.56 or 0.96 or 2mg/kg (n=1), corresponding to the usual, maximal and twice the maximal dose in patients, respectively, administered immediately before PET. [(11)C]dLop brain kinetics as well as [(11)C]dLop kinetics and radiometabolites in arterial plasma were measured to calculate [(11)C]dLop area-under the time-activity curve from 10 to 30min in the brain (AUCbrain) and in plasma (AUCplasma). [(11)C]dLop brain uptake was described by AUCR=AUCbrain/AUCplasma. CsA as well as DPy did not measurably influence [(11)C]dLop plasma kinetics and metabolism. Baseline AUCR (0.85±0.29) was significantly enhanced in the presence of CsA (AUCR=10.8±3.6). Injection of pharmacologic dose of DPy did not enhance [(11)C]dLop brain distribution with AUCR being 1.2, 0.9 and 1.1 after administration of 0.56, 0.96 and 2mg/kg DPy doses, respectively. We used [(11)C]dLop PET imaging in baboons, a relevant in vivo model of P-gp function at the BBB, to show the P-gp inhibition potency of therapeutic dose CsA. Despite in vitro P-gp inhibition potency, usual doses DPy are not likely to inhibit P-gp function at

  9. Prevalence of acute and chronic viral seropositivity and characteristics of disease in patients with psoriatic arthritis treated with cyclosporine: a post hoc analysis from a sex point of view on the observational study of infectious events in psoriasis complicated by active psoriatic arthritis

    PubMed Central

    Colombo, Delia; Chimenti, Sergio; Grossi, Paolo Antonio; Marchesoni, Antonio; Bardazzi, Federico; Ayala, Fabio; Simoni, Lucia; Vassellatti, Donatella; Bellia, Gilberto

    2016-01-01

    Background Sex medicine studies have shown that there are sex differences with regard to disease characteristics in immune-mediated inflammatory diseases, including psoriasis, in immune response and susceptibility to viral infections. We performed a post hoc analysis of the Observational Study of infectious events in psoriasis complicated by active psoriatic arthritis (SYNERGY) study in patients with psoriatic arthritis (PsA) treated with immunosuppressive regimens including cyclosporine, in order to evaluate potential between-sex differences in severity of disease and prevalence of viral infections. Methods SYNERGY was an observational study conducted in 24 Italian dermatology clinics, which included 238 consecutively enrolled patients with PsA, under treatment with immunosuppressant regimens including cyclosporin A. In this post hoc analysis, patients’ demographical data and clinical characteristics of psoriasis, severity and activity of PsA, prevalence of seropositivity for at least one viral infection, and treatments administered for PsA and infections were compared between sexes. Results A total of 225 patients were evaluated in this post hoc analysis, and 121 (54%) were males. Demographic characteristics and concomitant diseases were comparable between sexes. Statistically significant sex differences were observed at baseline in Psoriasis Area and Severity Index score (higher in males), mean number of painful joints, Bath Ankylosing Spondylitis Disease Activity Index, and the global activity of disease assessed by patients (all higher in females). The percentage of patients with at least one seropositivity detected at baseline, indicative of concomitant or former viral infection, was significantly higher among women than among men. No between-sex differences were detected in other measures, at other time points, and in treatments. Patients developed no hepatitis B virus or hepatitis C virus reactivation during cyclosporine treatment. Conclusion Our post hoc

  10. Selective lymphoid irradiation and cyclosporin A in rat heart allografts

    SciTech Connect

    Kuromoto, N.; Hardy, M.A.; Fawwaz, R.; Reemtsma, K.; Nowygrod, R.

    1984-05-01

    Short-term peritransplant treatment utilizing 2-dose ALG and 1-dose Palladium-109-hematoporphyrin (PD-H) for selective lymphoid irradiation (SLI) leads to donor-specific permanent acceptance of heart allografts in the Fisher to Lewis rat model. The same treatment significantly prolongs survival of hearts transplanted to strongly histoincompatable , presensitized, and xenogeneic recipients. The purpose of this study was to evaluate synergistic effects of short-term, low-dose cyclosporin treatment and SLI in an attempt to develop a nontoxic protocol utilizing peritransplant treatment for immune preconditioning with minimal subsequent immunosuppression. Single-agent treatment alone with cyclosporin, ALG, or Pd-H resulted in a maximal mean graft survival time (MST) of 33 days. Immunosuppression with 1-dose Pd-H, 2-dose ALG, and low-dose cyclosporin (5 mg/kg) for 14 days doubled the MST to 78 days. Use of therapeutic-dose cyclosporin (20 mg/kg), given for just 3 days, was also quite effective, MST of 57 days with SLI and 43 days with ALG, but toxic; 3 of 12 recipients died of infection with functioning grafts. These results demonstrate that the use of low-dose cyclosporin over a short interval, when combined with peritransplant SLI, is a highly effective and safe method for prolonging heart allograft survival.

  11. Cyclosporin A monotherapy versus cyclosporin A and methotrexate combination therapy in patients with early rheumatoid arthritis: a double blind randomised placebo controlled trial

    PubMed Central

    Gerards, A; Landewe, R; Prins, A; Bruijn, G; Goei, T; Laan, R; Dijkmans, B

    2003-01-01

    Patients and methods: 120 patients with active RA, rheumatoid factor positive and/or erosive, were randomly allocated to receive CsA with MTX (n=60) or CsA with placebo (n=60). Treatment with CsA was started in all patients at 2.5 mg/kg/day and increased to a maximum of 5 mg/kg/day in 16 weeks. MTX was started at 7.5 mg/week and increased to a maximal dose of 15 mg/week at week 16. Primary outcomes were clinical remission (Pinals criteria) and radiological damage (Larsen score), at week 48. Results: Treatment was discontinued prematurely in 27 patients in the monotherapy group (21 because of inefficacy, and six because of toxicity) and in 26 patients in the combination therapy group (14 and 12, respectively). At week 48, clinical remission was achieved in four patients in the monotherapy group and in six patients in the combination therapy group (p=0.5). The median Larsen score increased to 10 (25th, 75th centiles: 3.5; 13.3) points in the monotherapy group and to 4 (1.0; 10.5) points in the combination therapy group (p=0.004). 28/60 (47%) of patients in the monotherapy group v 34/60 (57%) of patients in the combination therapy group had reached an American college of Rheumatology 20% (ACR20) response (p=0.36) at week 48; 15/60 (25%) v 29/60 (48%) of patients had reached an ACR50 response (p=0.013); and 7 (12%) v 12 (20%) of patients had reached an ACR70 response (p=0.11).Their was a tendency towards more toxicity in the combination therapy group. Conclusions: In patients with early RA, neither CsA plus MTX combination therapy nor CsA monotherapy is very effective in inducing clinical remission. Combination therapy is probably better at improving clinical disease activity, and definitely better at slowing radiological progression. Combination therapy should still be compared with methotrexate monotherapy. PMID:12634224

  12. 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

  13. 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.

  14. Insights into cyclosporine A-induced atherosclerotic risk in transplant recipients: macrophage scavenger receptor regulation.

    PubMed

    Jin, Song; Mathis, A Scott; Rosenblatt, Joseph; Minko, Tamara; Friedman, Gary S; Gioia, Kevin; Serur, David S; Knipp, Gregory T

    2004-02-27

    Clinical monitoring of organ-transplant recipients suggests that administration of cyclosporine (CsA) may increase the risk of atherosclerosis when compared with the general population. The purpose of this work is to demonstrate the utility of the in vitro Tamm-Horsfall protein (THP)-1 human monocyte cell culture model for determining drug-related atherosclerotic potential in macrophages. The effect of CsA on the mRNA expression of macrophage scavenger receptor genes including CD36, CD68, scavenger receptor (SR)-A, SR-BII, and lectin-like oxidized low-density lipoprotein receptor (LOX-1); the nuclear hormone receptors, including peroxisome-proliferator activated receptor (PPAR)gamma and liver-X-receptor (LXR)alpha; and the cholesterol efflux pump ABCA1 were investigated as markers of atherosclerotic progression. The THP-1 cells were cultured and differentiated into macrophages. The macrophages were then treated with CsA to assess gene expression. Time- (1, 2, 4, 8, and 24 hours) and dose- (concentrations [mg/L] corresponding to the trough [0.5], peak [1.25] and 4x peak [5]) dependency of CsA was assessed. The treated macrophage mRNA gene expression of CD36, CD68, and PPARgamma were up-regulated in the presence of CsA. Interestingly, SR-A, SR-BII, LOX-1, and LXRalpha expression appeared to be slightly down-regulated, and ABCA1 was relatively unchanged. Immunoblotting studies demonstrated that the protein expression of CD36 was unchanged or increased, PPARgamma was unchanged, and ABCA1 was unchanged or decreased at 4 and 8 hours. The results document CsA-induced mRNA and protein changes in receptors relevant to lipid-laden foam cell formation and demonstrate the utility of THP-1 macrophages for screening of atherosclerotic risk potential. PMID:15084924

  15. Influence of nifedipine on interstitial fibrosis in renal transplant allografts treated with cyclosporin A.

    PubMed Central

    McCulloch, T A; Harper, S J; Donnelly, P K; Moorhouse, J; Bell, P R; Walls, J; Feehally, J; Furness, P N

    1994-01-01

    AIMS--To compare the degree of interstitial fibrosis in renal transplant biopsy specimens from immunosuppressed patients using conventional doses of cyclosporin with and without calcium channel blockade with a combination of low dose cyclosporin and azathioprine; to correlate the degree of interstitial fibrosis with the glomerular filtration rate. METHODS--A single blind histomorphometric assessment was done of cortical interstitial volume fraction from biopsy specimens taken intraoperatively and at one, six, and 12 months after transplantation from three prospectively randomised groups of patients: (A) conventional dose cyclosporin; (B) conventional dose cyclosporin plus nifedipine; (C) low dose cyclosporin plus azathioprine. RESULTS--Interstitial volume increased with time in all groups. No differences in interstitial volume were present at operation or at one month, but at six months interstitial volume was significantly less in group B than group A (p < 0.001) or group C (p < 0.05). More grafts failed in group A than group B leaving only small numbers for comparison at 12 months. At 12 months the differences persisted but did not reach significance. These results strongly reflected the clinical findings, where glomerular filtration rate was significantly lower in group A than groups B or C at six and 12 months; no differences in glomerular filtration rate were found at one month. In a direct comparison glomerular filtration rate showed a significant negative correlation with interstitial volume fraction. CONCLUSIONS--These findings suggest that calcium channel blockade with nifedipine slows the development of interstitial fibrosis in renal transplant recipients treated with cyclosporin. When clinical data are considered, it is suggested that calcium channel blockade may have a mitigating effect on the long term nephrotoxic effects of cyclosporin and should be considered as adjunctive treatment in patients requiring this immunosuppressant following renal transplantation. Images PMID:7962654

  16. 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.

  17. Insights into cyclosporine A-induced atherosclerotic risk in transplant recipients: macrophage scavenger receptor regulation.

    PubMed

    Jin, Song; Mathis, A Scott; Rosenblatt, Joseph; Minko, Tamara; Friedman, Gary S; Gioia, Kevin; Serur, David S; Knipp, Gregory T

    2004-02-27

    Clinical monitoring of organ-transplant recipients suggests that administration of cyclosporine (CsA) may increase the risk of atherosclerosis when compared with the general population. The purpose of this work is to demonstrate the utility of the in vitro Tamm-Horsfall protein (THP)-1 human monocyte cell culture model for determining drug-related atherosclerotic potential in macrophages. The effect of CsA on the mRNA expression of macrophage scavenger receptor genes including CD36, CD68, scavenger receptor (SR)-A, SR-BII, and lectin-like oxidized low-density lipoprotein receptor (LOX-1); the nuclear hormone receptors, including peroxisome-proliferator activated receptor (PPAR)gamma and liver-X-receptor (LXR)alpha; and the cholesterol efflux pump ABCA1 were investigated as markers of atherosclerotic progression. The THP-1 cells were cultured and differentiated into macrophages. The macrophages were then treated with CsA to assess gene expression. Time- (1, 2, 4, 8, and 24 hours) and dose- (concentrations [mg/L] corresponding to the trough [0.5], peak [1.25] and 4x peak [5]) dependency of CsA was assessed. The treated macrophage mRNA gene expression of CD36, CD68, and PPARgamma were up-regulated in the presence of CsA. Interestingly, SR-A, SR-BII, LOX-1, and LXRalpha expression appeared to be slightly down-regulated, and ABCA1 was relatively unchanged. Immunoblotting studies demonstrated that the protein expression of CD36 was unchanged or increased, PPARgamma was unchanged, and ABCA1 was unchanged or decreased at 4 and 8 hours. The results document CsA-induced mRNA and protein changes in receptors relevant to lipid-laden foam cell formation and demonstrate the utility of THP-1 macrophages for screening of atherosclerotic risk potential.

  18. Effect of HLA matching and cyclosporine A in renal transplantation. ANZDATA Registry.

    PubMed

    Disney, A P

    1986-01-01

    1. Graft survival rates increased significantly in primary and second cadaver transplants as well as in one-haplotype matched living related donor renal allografts treated with CsA. 2. HLA mismatching did not seem to influence survival of primary cadaver grafts in transfused recipients. 3. No pertinent analysis was available owing to the small size of the group of nontransfused patients. 4. HLA mismatching did not appear to influence survival of second cadaver grafts in CsA-treated patients; the number of patients was small. 5. Graft survival of living donor grafts was not influenced by haplotype identity.

  19. Accelerated atherosclerosis in hyperlipidemic C57BL/6 mice treated with cyclosporin A.

    PubMed Central

    Emeson, E. E.; Shen, M. L.

    1993-01-01

    We and others have demonstrated that T lymphocytes are prominent components of atherosclerotic lesions. We hypothesized that if T cells were necessary for the development of atherosclerosis it would be possible to demonstrate its prevention or retardation in T-cell-suppressed mice. To test this hypothesis, CyA, a potent suppressor of T-cell activation, was used to treat C57BL/6 mice undergoing lipid hyperalimentation. Mice receiving normal mouse chow were completely free of atherosclerotic lesions. In mice receiving the atherogenic diet plus control oil injections, lesions of the aorta and coronary arteries were observed at 135 days and increased progressively in area until 310 days. Somewhat surprisingly, mice given the atherogenic diet plus CyA injections displayed even larger lesions at all three observed time intervals. Although CyA did suppress T-cell reactivity sufficiently to obtain the expected prolongation of skin allografts, it did not suppress the development or progression of atherosclerotic lesions. Images Figure 4 Figure 5 Figure 6 PMID:8506958

  20. Cyclosporine A-induced gingival hyperplasia pemphigus vulgaris: literature review and report of a case.

    PubMed

    Oettinger-Barak, O; Machtei, E E; Peled, M; Barak, S; L-Naaj, I A; Laufer, D

    2000-04-01

    Gingival hyperplasia appears in 8% to 85% of patients treated with cyclosporine. Most studies show an association between oral hygiene status and the prevalence and severity of this gingival overgrowth. Thus, besides attempting to substitute this drug with another whenever possible, treatment usually involves maintenance of strict oral hygiene coupled with scaling and root planing and removal of iatrogenic factors. Sometimes a second treatment phase involving periodontal surgery is necessary. Cyclosporine-induced gingival overgrowth has been mainly described in post-organ transplant patients. The present case describes, for the first time, a severe form of cyclosporine-induced gingival overgrowth arising in a 15 year-old male with pemphigus vulgaris. Periodontal treatment included oral hygiene and scaling and root planing under local anesthesia. There was a significant reduction in gingival enlargement, as well as a reduction in plaque levels and inflammation. Cessation of drug administration, combined with continuous periodontal treatment, brought further improvement. This successful conservative treatment of cyclosporine-induced gingival overgrowth in a pemphigus vulgaris patient suggests that early diagnosis and comprehensive treatment of these lesions may yield good response and reduce the need for periodontal surgery.

  1. 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

  2. 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

  3. 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.

  4. Purification of a modified cyclosporine A by co-current centrifugal partition chromatography: process development and intensification.

    PubMed

    Amarouche, Nassima; Boudesocque, Leslie; Sayagh, Charlotte; Giraud, Matthieu; McGarrity, John; Butte, Alessandro; Marchal, Luc; Foucault, Alain; Renault, Jean-Hugues

    2013-10-11

    Synthetic hydrophobic non-ionizable peptides are not soluble in most common solvents and are thus difficult to purify by preparative reversed-phase HPLC, normally used for industrial production. The challenge exists to develop alternative purification chromatographic processes using suitable solvents and providing good yields, high purity and sufficient productivity. A 11mer hydrophobic synthetic modified cyclosporine, showing an anti-HIV activity, was successfully purified by centrifugal partition chromatography using the biphasic solvent system heptane/ethyl acetate/acetone/methanol/water (1:2:2:1:2, v/v). A 5% co-current elution - made possible by the liquid nature of the two phases - has been used in order to avoid hydrodynamic instabilities mainly due to the physico-chemical properties of the target peptide. This original solution was developed after the study of the effect of the peptide on the hydrodynamic behavior of the two phases during the separation, and the visualization of the flow patterns using the Visual-CPC device. Critical impurities were efficiently eliminated and the peptide was recovered in high yield and high productivity achieving the specifications requirements.

  5. Effects of glycerides on the intestinal absorption of cyclosporine a using the in-situ mesenteric vein cannulated rat model.

    PubMed

    Ghorab, Mamdouh M; Abdel-Salam, Heba M; Abdel-Moate, Mohamed M

    2005-07-01

    The purpose of this study was to evaluate and compare the effect of glycerides with different fatty acid distributions (e.g. Arlacel 186, Capmul GMO and Captex 350) on Cyclosporine absorption in rat ileum segment using the modified single-pass intestinal perfusion with mesenteric vein cannulation. Drug concentration in the perfusate and blood plasma was analyzed by HPLC; and permeability coefficients were calculated from drug appearance in blood (P(blood)) and disappearance from perfusate (P(lumen)). Particle size was measured using Malvern Zetasaizer 1000HSA. Rheologic properties were measured using Brookfield viscometer. The results show that the average particle sizes after dilution (100 folds) of formulae containing Arlacel 186, Capmul GMO and Captex 350 and containing 0.8 mM CsA were 260+/-35.8, 130+/-11.4 and 37.5+/-6.0 nm, respectively. The polydispersity index was 0.6, 0.7 and 0.108 for formulations with Arlacel 186, Capmul GMO and Captex 350, respectively. CsA permeability coefficients (P(blood)) calculated from drug appearance in the blood in presence of Arlacel 186, Capmul GMO and Captex 350 were 0.3x10(-6), 1.0x10(-6) and 1.7x10(-6) cm2/sec, respectively. Phenol red was used as a water marker to determine net water absorption and secretion. Its constant concentration suggested that formulation did not alter intestinal water flux. From the results we can conclude that degree of glyceride esterification has a potential impact on the average particle size distribution and polydispersity of the formed micelles on dilution, which on turn contribute to the interaction between membrane and drug.

  6. 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

  7. 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

  8. Efficacy of combination therapy with anti-thymocyte globulin and cyclosporine A as a first-line treatment in adult patients with aplastic anemia: a comparison of rabbit and horse formulations.

    PubMed

    Suzuki, Takahiro; Kobayashi, Hiroyuki; Kawasaki, Yasufumi; Okazuka, Kiyoshi; Hatano, Kaoru; Fujiwara, Shin-Ichiro; Oh, Iekuni; Ohmine, Ken; Kanda, Yoshinobu

    2016-10-01

    Anti-thymocyte globulin (ATG) is a key drug in immunosuppressive therapy for patients with aplastic anemia. The mainstay of ATG therapy had been a horse ATG (hATG) formulation, Lymphoglobulin or ATGAM, but Lymphoglobulin was recently discontinued, and Thymoglobulin, a rabbit ATG (rATG) formulation, is currently used as the first-line drug in many countries, including Japan. However, a recent randomized clinical trial reported significantly unfavorable outcomes associated with the use of rATG regimens. We retrospectively analyzed clinical outcomes of adult patients with moderate to severe aplastic anemia who were treated with 3.5 mg/kg of Thymoglobulin (n = 22) or 15 mg/kg of Lymphoglobulin (n = 25) in our facility. The estimated overall response rates in the rATG and hATG groups were 64.6 versus 56.0 % at 6 months, and 76.4 versus 69.2 % at 12 months, respectively; and there was no statistical difference between the two groups (P = 0.32). Overall survival at 24 months was not significantly different: rATG 89.8 % versus hATG 96.0 % (P = 0.39). Early phase infection was observed in 37.5 % of cases in the rATG and 14.8 % in the hATG group, but the frequency was not statistically different (P = 0.107). Our data indicate that Thymoglobulin at a dose of 3.5 mg/kg is a viable alternative when hATG is not available.

  9. Inflammatory Myopathies

    MedlinePlus

    ... the symptoms can be treated. Options include medication, physical therapy, exercise, heat therapy (including microwave and ultrasound), orthotics ... and polymyositis include cyclosporine A, cyclophosphamide, and tacrolimus. Physical therapy is usually recommended to prevent muscle atrophy and ...

  10. 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

  11. Calcineurin Regulates Cyclin D1 Accumulation in Growth-stimulated Fibroblasts

    PubMed Central

    Kahl, Christina R.; Means, Anthony R.

    2004-01-01

    Calcium (Ca2+) and calmodulin (CaM) are required for progression of mammalian cells from quiescence into S phase. In multiple cell types, cyclosporin A causes a G1 cell cycle arrest, implicating the serine/threonine phosphatase calcineurin as one Ca2+/CaM-dependent enzyme required for G1 transit. Here, we show, in diploid human fibroblasts, that cyclosporin A arrested cells in G1 before cyclin D/cdk4 complex activation and retinoblastoma hyperphosphorylation. This arrest occurred in early G1 with low levels of cyclin D1 protein. Because cyclin D1 mRNA was induced normally in the cyclosporin A-treated cells, we analyzed the half-life of cyclin D1 in the presence of cyclosporin A and found no difference from control cells. However, cyclosporin A treatment dramatically reduced cyclin D1 protein synthesis. Although these pharmacological experiments suggested that calcineurin regulates cyclin D1 synthesis, we evaluated the effects of overexpression of activated calcineurin on cyclin D1 synthesis. In contrast to the reduction of cyclin D1 with cyclosporin A, ectopic expression of calcium/calmodulin-independent calcineurin promoted synthesis of cyclin D1 during G1 progression. Therefore, calcineurin is a Ca2+/CaM-dependent target that regulates cyclin D1 accumulation in G1. PMID:14767060

  12. Evaluation of traditional medicines III: the mechanism of immune modulation by PHELA.

    PubMed

    Lekhooa, Makhotso; Walubo, Andrew; Du Plessis, Jan J B; Matsabisa, Motlalepula G

    2012-01-01

    PHELA is a herbal traditional medicine that is under development for use as an immune booster in immune compromised individuals. Therefore, the aim of this study was to determine PHELA's mechanism of action by observing for changes in cytokine profiles. Four groups of Sprague Dawley rats (n = 8) were treated daily and separately with normal-saline, cyclosporine-A, PHELA-only and PHELA+ cyclosporine-A. Thereafter, 4 animals from each group were sacrificed after 7 and 14 days of treatment. Serum Th1 cytokines (IL-2, IFN-γ and TNF-α) and Th2 cytokines (IL-4 and IL-10) were measured by ELISA. The concentrations of Th1 cytokines in the PHELA-only treated group were similar to the control group on days 7 and 14. However, the Th1 cytokines were higher in the PHELA+cyclosporine-A treated group compared to cyclosporine-A group, and cyclosporine-A concentrations were similar in both groups. These results show that PHELA did not stimulate Th1 cytokines of a normal immune system but stimulated them when the immune system was suppressed by cyclosporine-A. In conclusion, PHELA is an immune-stimulant to a compromised immune system.

  13. Plasma endothelin in psoriasis: possible relations to therapy and toxicity.

    PubMed

    Zachariae, H; Heickendorff, L; Bjerring, P

    1996-11-01

    Plasma endothelin levels were studied in 71 patients suffering from severe psoriasis. The psoriatics were treated either with topical therapy alone (n = 18) or with cyclosporin A (n = 26), methotrexate (n = 21), or with hydroxyurea, acitretin or ranitidin (n = 6) with or without topical therapy. The psoriatics had a significantly higher average plasma endothelin than 40 healthy controls. The patients treated with cyclosporin A had the highest values and these were in contrast to patients on methotrexate and other systemic therapy higher than patients treated with topical therapy alone. There was not significant difference between endothelin levels in patients treated with methotrexate compared to those in patients only receiving topical treatment. Whether the increased endothelin levels in plasma are derived from keratinocytes or enlarged vessels need to be investigated. An increased plasma endothelin level could be related to therapy and for patients on cyclosporin A be of importance for toxicity.

  14. Multiple cotton wool spots following bone marrow transplantation for treatment of acute lymphatic leukaemia.

    PubMed Central

    Gloor, B; Gratwohl, A; Hahn, H; Kretzschmar, S; Robert, Y; Speck, B; Daicker, B

    1985-01-01

    Three patients with acute lymphatic leukaemia developed visual impairment due to occlusion of small retinal vessels with multiple cotton wool spots after treatment which included whole body and skull irradiation followed by bone marrow transplantation and cyclosporin A. Withdrawal of cyclosporin A and treatment with corticosteroids was followed by recovery of visual acuity. This retinopathy and the retinal changes seen in the immunodeficiency syndrome are thought to be closely related. The possible role of cyclosporin A is discussed, though cotton wool spots and retinal haemorrhages have never been described in renal transplant patients during treatment with this drug. Withdrawal of cyclosporin A, which is highly effective in preventing graft-versus-host disease, can be fatal. Irradiation of the skull prior to bone marrow transplantation and intrathecal administration of methotrexate may be the most important factors causing the retinal ischaemic signs described here. The inclusion of an ophthalmologist in the team monitoring transplant patients would lead to increased documentation and a better understanding of this disease. Images PMID:3888252

  15. Bile canalicular cationic dye secretion as a model for P-glycoprotein mediated transport.

    PubMed

    Thalhammer, T; Stapf, V; Gajdzik, L; Graf, J

    1994-04-01

    This study explores properties of P-glycoprotein dependent membrane transport in rat liver with the use of acridine orange as the substrate. We studied the biliary secretion of the dye, its binding to canalicular membrane P-glycoprotein, and effects of the inhibitor cyclosporin A: acridine orange is excreted into bile together with less hydrophobic and glucuronidated metabolites. Cyclosporin A inhibited both the secretion of acridine orange and of its metabolites. In TR- animals, a rat strain that is deficient of the canalicular multi-specific organic anion transport system, non-metabolized acridine orange is the predominant species in bile and its secretion is also inhibited by cyclosporin A. Binding of acridine orange to liver P-glycoprotein was analyzed by photoaffinity labeling with azidopine, a substrate of P-glycoprotein dependent transport in multi-drug resistant tumor cells. Labeling of the immunoprecipitated P-glycoprotein was inhibited by acridine orange, verapamil, and by cyclosporin A. The results show that biliary secretion of acridine orange is highly analogous to P-glycoprotein mediated membrane drug transport in tumor cells that exhibit multi-drug resistance.

  16. Identification of cyclophilin as a proinflammatory secretory product of lipopolysaccharide-activated macrophages.

    PubMed Central

    Sherry, B; Yarlett, N; Strupp, A; Cerami, A

    1992-01-01

    We have isolated an 18-kDa peptide (designated sp18, for 18-kDa secreted protein) from the conditioned medium of lipopolysaccharide-stimulated RAW 264.7 murine macrophages. Purified sp18 had in vivo inflammatory activity and in vitro chemotactic activity for human peripheral blood polymorphonuclear leukocytes and monocytes. Surprisingly, N-terminal sequencing and tryptic mapping studies revealed that sp18 and cyclophilin, an intracellular protein that binds the immunosuppressive drug cyclosporin A, are highly homologous. The in vitro chemotactic activity of sp18 on monocytes was blocked by cyclosporin A but not by cyclosporin H, a structural analog of cyclosporin A that does not bind cyclophilin. Like purified porcine cyclophilin, mouse sp18 exhibited peptidyl-prolyl cis-trans isomerase activity. Medium conditioned by lipopolysaccharide-stimulated resident peritoneal exudate macrophages isolated from C57BL/6 mice contained substantially higher levels of sp18/cyclophilin than medium conditioned by nonstimulated macrophages. The observation that sp18/cyclophilin exhibits proinflammatory activity and is secreted by macrophages in response to endotoxin suggests that this protein may function as a cytokine, and invites the hypothesis that the immunosuppressive action of cyclosporin A results in part from interaction with an extracellular form of cyclophilin released as a mediator of immune and inflammatory functions. Images PMID:1565646

  17. Rgg protein structure-function and inhibition by cyclic peptide compounds.

    PubMed

    Parashar, Vijay; Aggarwal, Chaitanya; Federle, Michael J; Neiditch, Matthew B

    2015-04-21

    Peptide pheromone cell-cell signaling (quorum sensing) regulates the expression of diverse developmental phenotypes (including virulence) in Firmicutes, which includes common human pathogens, e.g., Streptococcus pyogenes and Streptococcus pneumoniae. Cytoplasmic transcription factors known as "Rgg proteins" are peptide pheromone receptors ubiquitous in Firmicutes. Here we present X-ray crystal structures of a Streptococcus Rgg protein alone and in complex with a tight-binding signaling antagonist, the cyclic undecapeptide cyclosporin A. To our knowledge, these represent the first Rgg protein X-ray crystal structures. Based on the results of extensive structure-function analysis, we reveal the peptide pheromone-binding site and the mechanism by which cyclosporin A inhibits activation of the peptide pheromone receptor. Guided by the Rgg-cyclosporin A complex structure, we predicted that the nonimmunosuppressive cyclosporin A analog valspodar would inhibit Rgg activation. Indeed, we found that, like cyclosporin A, valspodar inhibits peptide pheromone activation of conserved Rgg proteins in medically relevant Streptococcus species. Finally, the crystal structures presented here revealed that the Rgg protein DNA-binding domains are covalently linked across their dimerization interface by a disulfide bond formed by a highly conserved cysteine. The DNA-binding domain dimerization interface observed in our structures is essentially identical to the interfaces previously described for other members of the XRE DNA-binding domain family, but the presence of an intermolecular disulfide bond buried in this interface appears to be unique. We hypothesize that this disulfide bond may, under the right conditions, affect Rgg monomer-dimer equilibrium, stabilize Rgg conformation, or serve as a redox-sensitive switch.

  18. Rgg protein structure–function and inhibition by cyclic peptide compounds

    PubMed Central

    Parashar, Vijay; Aggarwal, Chaitanya; Federle, Michael J.; Neiditch, Matthew B.

    2015-01-01

    Peptide pheromone cell–cell signaling (quorum sensing) regulates the expression of diverse developmental phenotypes (including virulence) in Firmicutes, which includes common human pathogens, e.g., Streptococcus pyogenes and Streptococcus pneumoniae. Cytoplasmic transcription factors known as “Rgg proteins” are peptide pheromone receptors ubiquitous in Firmicutes. Here we present X-ray crystal structures of a Streptococcus Rgg protein alone and in complex with a tight-binding signaling antagonist, the cyclic undecapeptide cyclosporin A. To our knowledge, these represent the first Rgg protein X-ray crystal structures. Based on the results of extensive structure–function analysis, we reveal the peptide pheromone-binding site and the mechanism by which cyclosporin A inhibits activation of the peptide pheromone receptor. Guided by the Rgg–cyclosporin A complex structure, we predicted that the nonimmunosuppressive cyclosporin A analog valspodar would inhibit Rgg activation. Indeed, we found that, like cyclosporin A, valspodar inhibits peptide pheromone activation of conserved Rgg proteins in medically relevant Streptococcus species. Finally, the crystal structures presented here revealed that the Rgg protein DNA-binding domains are covalently linked across their dimerization interface by a disulfide bond formed by a highly conserved cysteine. The DNA-binding domain dimerization interface observed in our structures is essentially identical to the interfaces previously described for other members of the XRE DNA-binding domain family, but the presence of an intermolecular disulfide bond buried in this interface appears to be unique. We hypothesize that this disulfide bond may, under the right conditions, affect Rgg monomer–dimer equilibrium, stabilize Rgg conformation, or serve as a redox-sensitive switch. PMID:25847993

  19. Intranigral grafts of fetal ventral mesencephalic tissue in adult 6-hydroxydopamine-lesioned rats can induce behavioral recovery.

    PubMed

    Johnston, R E; Becker, J B

    1997-01-01

    Intrastriatal grafts of fetal ventral mesencephalon in rats with unilateral 6-hydroxydopamine lesions can reduce and even reverse rotational behavior in response to direct and indirect dopamine agonists. These grafts can ameliorate deficits on simple spontaneous behaviors, but do not improve complex behaviors that require the skilled integration of the use of both paws. We report here that rats with grafts into the DA-depleted substantia nigra, that receive cyclosporine A, can experience recovery on spontaneous behaviors that mimic those observed in Parkinson's disease. Specific cyclosporine A treatment conditions can differentially affect whether intranigral grafts normalize paw use during initiation or termination of a movement sequence. These findings may have important implications for the treatment of Parkinson's disease. PMID:9171159

  20. Cyclophilins of a novel subfamily interact with SNW/SKIP coregulator in Dictyostelium discoideum and Schizosaccharomyces pombe.

    PubMed

    Skruzný, M; Ambrozková, M; Fuková, I; Martínková, K; Blahůsková, A; Hamplová, L; Půta, F; Folk, P

    2001-10-31

    We screened the Dictyostelium discoideum two-hybrid cDNA library with the SNW/SKIP transcription coregulator SnwA and identified a novel cyclophilin CypE. Independently, the Schizosaccharomyces pombe cDNA library was screened with the SnwA ortholog Snw1 and the ortholog of CypE (named Cyp2) was found. Both cyclophilins bind the respective SNW protein in their autologous systems. The interaction was localized to the N-terminal part of SnwA as well as of Snw1. CypE was confirmed in vitro to be a cyclosporin A-sensitive peptidyl-prolyl cis-trans isomerase. Remarkably, both SNW proteins bind the cyclophilins in a cyclosporin A independent manner, possibly serving as adaptors for these novel isomerases. These results are the first characterization of the members of a novel cyclophilin subfamily, which includes the human CGI-124/PPIL1 protein. PMID:11690648

  1. The cyclophilins

    PubMed Central

    Wang, Ping; Heitman, Joseph

    2005-01-01

    Summary: Cyclophilins (Enzyme Commission (EC) number 5.1.2.8) belong to a group of proteins that have peptidyl-prolyl cis-trans isomerase activity; such proteins are collectively known as immunophilins and also include the FK-506-binding proteins and the parvulins. Cyclophilins are found in all cells of all organisms studied, in both prokaryotes and eukaryotes; humans have a total of 16 cyclophilin proteins, Arabidopsis up to 29 and Saccharomyces 8. The first member of the cyclophilins to be identified in mammals, cyclophilin A, is the major cellular target for, and thus mediates the actions of, the immunosuppressive drug cyclosporin A. Cyclophilin A forms a ternary complex with cyclosporin A and the calcium-calmodulin-activated serine/threonine-specific protein phosphatase calcineurin; formation of this complex prevents calcineurin from regulating cytokine gene transcription. Recent studies have implicated a diverse array of additional cellular functions for cyclophilins, including roles as chaperones and in cell signaling. PMID:15998457

  2. c-Jun N-terminal kinase negatively regulates epidermal growth factor-induced cyclooxygenase-2 expression in oral squamous cell carcinoma cell lines.

    PubMed

    Husvik, Camilla; Bryne, Magne; Halstensen, Trond S

    2009-12-01

    Epidermal growth factor (EGF)-induced cyclooxygenase-2 (COX-2) expression in squamous cell carcinomas is mediated through the extracellular signal-regulated kinase 1/2 and p38 pathways. Examination of a basaloid and a conventional oral squamous cell carcinoma cell line revealed that inhibition of c-Jun N-terminal kinase (JNK) with SP600125 increased EGF-induced (but not basal) COX-2 transcription 1.5-1.9-fold in extracellular signal-regulated kinase 1/2 and p38 pathway-dependent manners. Although JNK may phosphorylate the cyclosporine A-sensitive transcription factor, nuclear factor of activated T cells c3, it was seemingly not involved because cyclosporine A did not reduce EGF-induced COX-2 expression. Thus, JNK negatively regulated EGF-induced extracellular signal-regulated kinase 1/2 and/or p38-mediated COX-2 transcription, presumably through activating an unidentified phosphatase. PMID:20121928

  3. Immunosuppression in pancreas transplantation: the Euro SPK trials and beyond.

    PubMed

    Malaise, J; De Roover, A; Squifflet, J P; Land, W; Neuhaus, P; Pratschke, J; Kahl, A; Pascher, A; Boas-Knoop, S; Arbogast, H; Hoffmann, J; Illner, W D; Seissler; Schlamp; Viebahn; Wunsch; Hajt; Klar, E; Scharek, W; Hopt; Pisarski, P; Drognitz, O; Thurow, C; Dette, K; Bechstein, W O; Woeste, G; Klempnauer, J; Becker, T; Lück; Neipp; Königsrainer, A; Steurer, W; Margreiter, R; Mark; Bonatti; Saudek, F; Boucek, P; Adamec, M; Havrdova, T; Koznarova, R; Vanrenterghem, Y; Pirenne, J; Maes, B; Kuypers, D; Coosemans, W; Evenepoel, P; van Ophem, D; Marcelis, V; van Vlem; Peeters; de Hemptinne; de Roose; Fernandez-Cruz, L; Ricart, M J; Nakache, R; Morel, P; Berney, T; Demuylder, S

    2008-01-01

    The Immunosuppression in Pancreas Transplantation was historically based on the fact that the pancreas is an extremely immunogenic organ. Quadruple drug therapy with polyclonal or monoclonal antibodies induction was the mainstay therapy since the introduction of Cyclosporine A. In the modern era of Immunosuppression, Mycophenolate Mofetil replaced Azathioprine while Tacrolimus-another potent calcineurin inhibitor-had-and still has-a difficult challenge to replaced Cyclosporine A, due to its potential diabetogenic effect. Thanks to the first two EuroSPK studies which prospectively tried to answer several questions in that field. But, the future challenge will be in understanding the impact of innate immunity and ischemic reperfusion injuries on the long-term graft function. Hopefully, new drugs will be available and tested to block unspecific deleterious reactions to attenuate the proinflammatory response. It will be the aim of the third Euro SPK Study.

  4. 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

  5. A CD2AP Mutation Associated with Focal Segmental Glomerulosclerosis in Young Adulthood

    PubMed Central

    Tsvetkov, Dmitry; Hohmann, Michael; Anistan, Yoland Marie; Mannaa, Marwan; Harteneck, Christian; Rudolph, Birgit; Gollasch, Maik

    2016-01-01

    Mutations in CD2-associated protein (CD2AP) have been identified in patients with focal segmental glomerulosclerosis (FSGS); however, reports of CD2AP mutations remain scarce. We performed Sanger sequencing in a patient with steroid-resistant FSGS and identified a heterozygous CD2AP mutation (p.T374A, c.1120 A > G). Our patient displayed mild cognitive decline, a phenotypic characteristic not previously associated with CD2AP-associated FSGS. His proteinuria was remarkably reduced by treatment with cyclosporine A. Our findings expand the genetic spectrum of CD2AP-associated disorders and broaden the associated phenotype with the co-occurrence of cognitive decline. Our case shows that cyclosporin A is a treatment option for CD2AP-associated nephropathy. PMID:26997877

  6. Vasopressin attenuates ischemia-reperfusion injury via reduction of oxidative stress and inhibition of mitochondrial permeability transition pore opening in rat hearts.

    PubMed

    Nazari, Afshin; Sadr, Seyed Shahabeddin; Faghihi, Mahdieh; Azizi, Yaser; Hosseini, Mir-Jamal; Mobarra, Naser; Tavakoli, Asadollah; Imani, AliReza

    2015-08-01

    Aim of this study was to investigate the involvement of the mitochondrial permeability transition pore (MPTP) and oxidative stress in the cardioprotective effect of vasopressin (AVP) on ischemia/reperfusion (I/R) injury. Anesthetized male wistar rats were subjected to regional 30 min ischemia and 120 min reperfusion and randomly divided into nine groups: (1) Control; saline was administered intravenously before ischemia, (2) vasopressin was administrated 10 min prior to ischemia, (3, 4) Atractyloside as MPTP opener, was injected 5 min prior to reperfusion without and with vasopressin, (5, 6) Cyclosporine A as a MPTP closer, was injected 5 min prior to reperfusion without and with vasopressin, (7) mitochondria were isolated from control group and CaCl2 was added as MPTP opener and swelling inducer, (8) isolated mitochondria from Control hearts was incubated with Cyclosporine A before adding the CaCl2 (9) CaCl2 was added to isolated mitochondria from vasopressin group. Infusion of vasopressin decreased infarct size (18.6±1.7% vs. control group 37.6±2.4%), biochemical parameters [LDH (Lactate Dehydrogenase), CK-MB (Creatine Kinase-MB) and MDA (Malondialdehyde) plasma levels, PAB (Prooxidant-antioxidant balance)] compared to control group. Atactyloside suppressed the cardioprotective effect of vasopressin (32.5±1.9% vs. 18.6±1.7%) but administration of the Cyclosporine A without and with vasopressin significantly reduced infarct size to 17.7±4% (P<0.001) and 22.7±3% (P<0.01) respectively, vs. 37.6±2.4% in control group. Also, vasopressin, similar to Cyclosporine A, led to decrease in CaCl2-induced swelling. It seems that vasopressin through antioxidant effect and MPTP inhibition has created a cardioprotection against ischemia/reperfusion injuries.

  7. 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. PMID:26613512

  8. The Effect of TNF-α Blocker HL036337 and Its Best Concentration to Inhibit Dry Eye Inflammation

    PubMed Central

    Choi, Wungrak; Noh, Hyemi; Yeo, Areum; Jang, Hanmil; Ahn, Hyea Kyung; Song, Yeon Jung

    2016-01-01

    Purpose Dry eye syndrome is commonly thought of as an inflammatory disease, and we have previously presented data showing the effectiveness of topical TNF-α blocker agents for the treatment of this condition. The purpose of this study was to investigate the effectiveness of the TNF-α blocking agent HL036337 compared to cyclosporine A for the treatment of dry eye induced inflammation in order to establish whether HL036337 represents a more effective method for suppressing inflammation. The efficacy of HL036337 and cyclosporine A was determined using an experimental murine dry eye model. Methods The TNF-α blocker HL036337 is a modified form of TNF receptor I. Using dry eye induced C57BL/6 mice (n = 45), corneal erosion was measured at day 4 and 7 after topical treatment with cyclosporine A or HL036337. To determine the effective treatment dose, 0.25, 0.5, 1, 2.5, and 5 mg/mL of HL036337 were topically administered twice per day to dry eye induced murine corneas for 1 week. Results The optimal concentration of the TNF-α blocker HL036337 for treatment of dry eye induced corneal erosion was determined to be 1 mg/mL. Dry eye induced corneal erosion was improved after 1 week with topically applied cyclosporine A and HL036337 at 1 mg/mL. Conclusions HL036337 administered topically at 1 mg/mL effectively improved corneal erosion induced by dry eye. This finding may also suggest that inhibition of TNF-α can improve dry eye syndrome. PMID:27478358

  9. Molecular Evidence and Functional Expression of a Novel Drug Efflux pump (ABCC2) in Human Corneal Epithelium and Rabbit Cornea and its role in Ocular drug efflux

    PubMed Central

    Karla, Pradeep K.; Pal, Dhananjay; Quinn, Tim; Mitra, Ashim K.

    2007-01-01

    Cornea is considered as a major barrier for ocular drug delivery. Low ocular bioavailability of drugs has been attributed primarily to low permeability across corneal epithelium thus leading to sub-therapeutic concentrations of drug in the eye and treatment failure. The role of drug efflux proteins, particularly the Pglycoprotein in ocular drug bioavailability has been reported. The objective of this research was to determine whether human corneal epithelium expresses multi drug resistance associated proteins contributing to drug efflux by employing both cultured corneal cells and freshly excised rabbit cornea. SV40 HCEC and rPCEC were selected for in-vitro testing. SV40-HCEC and freshly excised rabbit corneas were utilized for transport studies. [3H]-cyclosporine-A and [14C]-erythromycin which are known substrates for ABCC2 and MK-571, a specific inhibitor for MRP were applied in this study. RT-PCR indicated a unique and distinct band at ∼272 bp corresponding to ABCC2 in HCEC, SV40-HCEC, rabbit cornea, rPCEC and MDCKII-MRP2 cells. Also RT-PCR indicated a unique band ∼181 bp for HCEC and SV40-HCEC. Immunoprecipitation followed by Western Blot analysis revealed a specific band at ∼190-kDa in membrane fraction of SV40-HCEC, MDCKII-MRP2 and no band with isotype control. Uptake of [3H]-cyclosporine-A and [14C]-erythromycin in the presence of MK-571 was significantly enhanced than control in both SV40 HCEC and rPCEC. Similarly a significant elevation in (A→B) permeability of [3H]-cyclosporine-A and [14C]-erythromycin was observed in the presence of MK-571 in SV40-HCEC. A→B transport of [3H]-cyclosporine-A was elevated in the presence of MK-571 in freshly excised rabbit cornea indicating potential role of this efflux transporter and high clinical significance of this finding. PMID:17156953

  10. Therapeutic perspectives in atopic dermatitis.

    PubMed

    Misery, Laurent

    2011-12-01

    Therapy of atopic dermatitis should comprise emollients, topical glucocorticosteroids, or calcineurin inhibitors, phototherapies, immunosuppressants like cyclosporin A, and other treatments. All these treatments should be improved, thanks to research. But new therapeutic perspectives should be given by topical anti-inflammatory substances, selective glucocorticoid receptor agonists, probiotics, interferon γ, TNFα inhibitors, inhibition of T cells or B cells, inhibition of IgE binding, and many other possibilities.

  11. Eosinophilic Fasciitis: A Rare Skin Sclerosis

    PubMed Central

    Servy, Amandine; Clérici, Thierry; Malines, Caroline; Le Parc, Jean-Marie; Côté, Jean-François

    2011-01-01

    Eosinophilic fasciitis (Schulman's syndrome) is a rare disease with specific clinical symptoms such as the groove sign which facilitate diagnosis. We report a typical case of eosinophilic fasciitis in an otherwise healthy 49-year-old man who presented with “prayer and groove signs”. Histological analysis showed sclerosis and eosinophilic infiltration of the fascia. The patient was successfully treated with systemic corticotherapy and Cyclosporine. A short review of the clinicopathological features of the lesions is presented. PMID:21151540

  12. Slitlamp biomicroscopy of the tear film of patients using topical Restasis and Refresh Endura.

    PubMed

    Solomon, Renée; Perry, Henry D; Donnenfeld, Eric D; Greenman, Herb E

    2005-04-01

    We describe a technique in which slitlamp biomicroscopy is performed to evaluate eyes in patients using a topical dry-eye agent; ie, topical cyclosporine A ophthalmic emulsion 0.05% (Restasis) or glycerin 1%-polysorbate 80 1% (Refresh Endura). Both are delivered in an oil-emulsion formulation. We also describe a previously unreported physical finding in the tear film of these patients. The examination technique can help confirm patient compliance and the tear-film stability of the 2 agents.

  13. 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.

  14. 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

  15. 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.

  16. Immunosuppressive macrolides of the type FK 506: a novel class of topical agents for treatment of skin diseases?

    PubMed

    Meingassner, J G; Stütz, A

    1992-06-01

    The immunosuppressive macrolide antibiotics FK 506 and rapamycin were tested for topical activity in experimental allergic contact dermatitis of farm pigs. This species was used because pig skin, in comparison to rodent skin, resembles human skin more closely. For comparison, cyclosporine A (CyA), which is orally but not topically active in patients with skin disease, dexamethasone, and clobetasol propionate were used. Treatment was performed twice, 30 min and 6 h after elicitation of challenge reaction. Topical application of 0.4 to 0.04% FK 506 caused a pronounced inhibition of inflammatory skin reactions of hypersensitivity to dinitrofluorobenzene. The treatment response was similar to the activity of 0.13% clobetasole. Dexamethasone (1.2%) was less active than clobetasol. In contrast, rapamycin and CyA were inactive at concentrations of 1.2 and 10%, respectively. Because the pig data on corticosteroids and cyclosporine A are in agreement with clinical findings, these studies indicate that immunosuppressive macrolides of the type FK 506 may be useful drugs for the topical treatment of human skin diseases that respond to local corticosteroids and oral treatment with cyclosporine A.

  17. Enhanced dissolution of inhalable cyclosporine nano-matrix particles with mannitol as matrix former.

    PubMed

    Yamasaki, Keishi; Kwok, Philip Chi Lip; Fukushige, Kaori; Prud'homme, Robert K; Chan, Hak-Kim

    2011-11-25

    This study aims to improve the dissolution of inhalable cyclosporine A nanoparticles by formulating the drug with mannitol as a hydrophilic nano-matrix former. The effect of mannitol content on the aerosol performance of the nano-matrix particles was also examined. Cyclosporine A nanosuspensions were produced by anti-solvent precipitation using a multi-inlet vortex mixer. Various amounts of mannitol were dissolved into the suspensions before spray drying to obtain micron-sized aggregates (nano-matrix powders). Dissolution properties of the powders in an aqueous medium, with the drug content, aggregate size distribution, surface roughness, physicochemical properties and aerosol performance were determined. The powders contained amorphous cyclosporine A and α-crystalline mannitol, with drug content being very close to the theoretical doses. Inclusion of mannitol enhanced the dissolution rate of the drug, without significantly affecting the aggregate size distribution, surface roughness and aerosol performance. This formulation approach may be applicable to improving the dissolution rate and bioavailability of hydrophobic drugs. PMID:21864662

  18. 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. PMID:22319884

  19. 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

  20. 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.

  1. Physiological stimuli evoke two forms of endocytosis in bovine chromaffin cells

    PubMed Central

    Chan, Shyue-An; Smith, Corey

    2001-01-01

    Exocytosis and endocytosis were measured following single, or trains of, simulated action potentials (sAP) in bovine adrenal chromaffin cells. Catecholamine secretion was measured by oxidative amperometry and cell membrane turnover was measured by voltage clamp cell capacitance measurements. The sAPs evoked inward Na+ and Ca2+ currents that were statistically identical to those evoked by native action potential waveforms. On average, a single secretory granule underwent fusion following sAP stimulation. An equivalent amount of membrane was then quickly internalised (τ = 560 ms). Stimulation with sAP trains revealed a biphasic relationship between cell firing rate and endocytic activity. At basal stimulus frequencies (single to 0.5 Hz) cells exhibited a robust membrane internalisation that then diminished as firing increased to intermediate levels (1.9 and 6 Hz). However at the higher stimulation rates (10 and 16 Hz) endocytic activity rebounded and was again able to effectively maintain cell surface near pre-stimulus levels. Treatment with cyclosporin A and FK506, inhibitors of the phosphatase calcineurin, left endocytosis characteristics unaltered at the lower basal stimulus levels, but blocked the resurgence in endocytosis seen in control cells at higher sAP frequencies. Based on these findings we propose that, under physiological electrical stimulation, chromaffin cells internalise membrane via two distinct pathways that are separable. One is prevalent at basal stimulus frequencies, is lessened with increased firing, and is insensitive to cyclosporin A and FK506. A second endocytic form is activated by increased firing frequencies, and is selectively blocked by cyclosporin A and FK506. PMID:11744761

  2. 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

  3. Decreased mineralocorticoid receptor expression in blood cells of kidney transplant recipients undergoing immunosuppressive treatment: cost efficient determination by quantitative PCR

    PubMed Central

    Heering, P J; Klein-Vehne, N; Fehsel, K

    2004-01-01

    Aims: Electrolyte imbalances caused by impaired ion transport are a frequent side effect of immunosuppressive treatment in renal transplant recipients. Clinical symptoms resemble features of hypoaldosteronism, although concentrations of aldosterone are in the normal range. Because immunosuppression might affect the hormone receptor status of cells, mineralocorticoid receptor (hMR) expression by peripheral blood leucocytes (PBL) was studied in these patients. Methods: Twenty one renal transplant recipients being treated with cyclosporine A and 19 healthy controls were tested. hMR expression was quantified by means of competitive reverse transcription polymerase chain reaction (cRT-PCR) and compared with receptor binding studies with subsequent Scatchard plot analysis carried out previously on 20 renal transplant recipients and 25 controls. Advantages of PCR were summarised and compared with Scatchard plot analysis. Results: Cyclosporine A caused a 37% decrease in hMR molecules on PBL in 75% of renal transplant recipients, and this effect was attributable to the downregulation of hMR transcription. PCR was 99% specific for the detection of hMR in PBL and highly reproducible. Conclusions: Decreases in hMR protein and RNA in PBL of transplant recipients revealed an inhibitory effect of cyclosporine A on hMR transcription. Because hMR acts as a transcription factor, the expression of several genes involved in electrolyte homeostasis is affected, leading to signs of nephrotoxicity that require therapeutic adjustments. Because of the small volume of blood, the assay can be repeated during treatment and is therefore useful for measuring treatment outcomes. Lower costs and the absence of radioactive challenge are further advantages of the PCR method. PMID:14693832

  4. [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. PMID:17505584

  5. FR901459, a novel immunosuppressant isolated from Stachybotrys chartarum No. 19392. Taxonomy of the producing organism, fermentation, isolation, physico-chemical properties and biological activities.

    PubMed

    Sakamoto, K; Tsujii, E; Miyauchi, M; Nakanishi, T; Yamashita, M; Shigematsu, N; Tada, T; Izumi, S; Okuhara, M

    1993-12-01

    FR901459, a novel immunosuppressant, has been isolated from the fermentation broth of Stachybotrys chartarum No. 19392. The molecular formula of FR901459 was determined as C62H111N11O13. FR901459 was found to be a member of the cyclosporin family. However, it is structurally distinct from any other cyclosporins discovered so far, in that Leu is present at position 5 instead of Val. FR901459 was capable of prolonging the survival time of skin allografts in rats with one third the potency of cyclosporin A. PMID:8294235

  6. [Conservative treatment of dry eye].

    PubMed

    Hefner, J; Reinshagen, H

    2014-11-01

    The use of topic anti-inflammatory drugs has become very important in the treatment of dry eye disease. Besides the basic therapy including tear replacement, use of serum eye drops and mucolytic eye drops, the topical application of corticosteroids and cyclosporin A is more commonly used in moderate to severe forms of dry eye disease. The consistent treatment of Meibomian gland dysfunction as a frequent reason for evaporative dry eye is also of particular importance. Understanding the chronicity of the disease and long-term compliance are the essential for successful therapy of this widespread disease.

  7. 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

  8. Angiotensin II Activates the Calcineurin/NFAT Signaling Pathway and Induces Cyclooxygenase-2 Expression in Rat Endometrial Stromal Cells

    PubMed Central

    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 Ca2+ concentration by targeting phospholypase C-gamma in endometrial stromal cells (ESC). A key element of the cellular response to Ca2+ signals is the activity of the Ca2+- 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 cells of

  9. 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.

  10. [Long-term course of immunosuppressive therapy of Vogt-Koyanagi-Harada syndrome].

    PubMed

    Wand, K; Abraham, S; Loos, D; Stumpfe, S; Lohmann, C; Maier, M; Feucht, N

    2015-10-01

    This article describes the case of a 22-year old female patient, who first presented with holocephalic headaches and bilateral loss in vision. After diagnosis of a complete Vogt-Koyanagi-Harada syndrome, high-dose corticosteroid therapy was initiated. Due to recurrent headaches 6 weeks later, immunosuppressive therapy was initiated with cyclosporine A. Because of an adverse effect (hirsutism) treatment was changed to azathioprine. In a long-term follow-up over 2 years the patient showed stable clinical findings with good visual acuity.

  11. Horse anti-thymocyte globulin and eculizumab as concomitant therapeutic approach in an aplastic paroxysmal nocturnal hemoglobinuria patient: go or no-go?

    PubMed

    Alashkar, Ferras; Dührsen, Ulrich; Röth, Alexander

    2016-10-01

    We present the case of a 64-yr-old female paroxysmal nocturnal hemoglobinuria (PNH) patient with subsequent transition to severe aplastic anemia in whom a concomitant successful immunosuppressive therapy (horse anti-thymocyte globulin (hATG) and cyclosporin A) in addition to an ongoing long-term treatment with the terminal complement inhibitor eculizumab was initiated. No reduced ATG efficacy or severe adverse events were observed in the presence of eculizumab. In addition, the possible reoccurrence of PNH symptoms (thromboembolic events and/or intravascular hemolysis) was prevented, while T-cell depletion was similar to non-eculizumab-treated patients with partial remission being evident by day 83.

  12. 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

  13. In vitro and in vivo replication of seal gammaherpesviruses in cells of multiple species.

    PubMed

    Martina, Byron E; Verjans, Georges M G M; Kuiken, Thijs; van Amerongen, Geert; Osterhaus, Albert D M E

    2007-01-01

    Phocid herpesvirus virus type 2 (PhHV-2), a putative gammaherpesvirus of seals, has been isolated from harbor seals (Phoca vitulina) and grey seals (Halichoerus grypus). In the present study, different PhHV-2 isolates were shown to have a broad in vitro tropism for various cell types from several mammalian species. Inbred mice and two species of non-human primates proved to be susceptible to experimental infection with PhHV-2. The development of myoepitheliomas and spleen hyperplasia upon cyclosporin A treatment in some of the PhHV-2-infected animals warrants further investigation of the oncogenic and zoonotic potential of this virus. PMID:17194610

  14. [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.

  15. Glycyrrhetinic acid-induced permeability transition in rat liver mitochondria.

    PubMed

    Salvi, Mauro; Fiore, Cristina; Armanini, Decio; Toninello, Antonio

    2003-12-15

    Glycyrrhetinic acid, a hydrolysis product of one of the main constituents of licorice, the triterpene glycoside of glycyrrhizic acid, when added to rat liver mitochondria at micromolar concentrations induces swelling, loss of membrane potential, pyridine nucleotide oxidation, and release of cytochrome c and apoptosis inducing factor. These changes are Ca(2+) dependent and are prevented by cyclosporin A, bongkrekic acid, and N-ethylmaleimide. All these observations indicate that glycyrrhetinic acid is a potent inducer of mitochondrial permeability transition and can trigger the pro-apoptotic pathway. PMID:14637195

  16. [Conservative treatment of dry eye].

    PubMed

    Hefner, J; Reinshagen, H

    2014-11-01

    The use of topic anti-inflammatory drugs has become very important in the treatment of dry eye disease. Besides the basic therapy including tear replacement, use of serum eye drops and mucolytic eye drops, the topical application of corticosteroids and cyclosporin A is more commonly used in moderate to severe forms of dry eye disease. The consistent treatment of Meibomian gland dysfunction as a frequent reason for evaporative dry eye is also of particular importance. Understanding the chronicity of the disease and long-term compliance are the essential for successful therapy of this widespread disease. PMID:25275793

  17. 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.

  18. 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. PMID:21370423

  19. [Treatment of periodontitis associated with cyclosporin-induced severe gingival hyperplasia with regular mechanical plaque removal in a patient with renal transplantation (Case report)].

    PubMed

    Keglevich, Tibor; Windisch, Péter; Gera, István

    2002-02-01

    A clinical case of a middle-aged kidney transplanted woman is presented. The woman has been taking Cyclosporin-A for over 13 years and has had advanced periodontitis and severe gingival swellings and gingival inflammation. The kidney transplant patient was treated and followed up for approx. four years. The treatment protocol included very thorough mechanical scaling and root planing, oral hygienic instructions and the regular professional maintenance program resulted in complete remission of the gingival overgrowth and stabilization of the periodontal condition. The gingival and periodontal conditions showed a continuing improvement over the time despite of the continuous CSA administration.

  20. [Behaviour of gingival elastic constituents in periodontal disease].

    PubMed

    Bonnaure-Mallet, M; Apiou, J; Chambon, Y

    1991-01-01

    Disorganization of gingival elastic constituents, including oxytalan fibers, elaunin fibers, and mature elastic fibers, is a feature of periodontal disease. In early stages where inflammation is moderate, this disorganization involves only oxytalan fibers. As the disease progresses, fragmentation and lamination of elaunin fibers and mature elastic fibers is seen. In cyclosporin A-induced gingival hyperplasia, there is a substantial increase in the amount of elastic fibers as compared with healthy gingiva. In gingival fragments from edentulous elderly individuals the elastic constituents form dense granules.

  1. Peptidyl prolyl cis-trans-isomerase activity associated with the lumen of the endoplasmic reticulum.

    PubMed Central

    Bose, S; Freedman, R B

    1994-01-01

    Peptidyl prolyl cis-trans-isomerase (PPI) activity was detected in microsomal fractions from bovine and rat liver. Extensive washing, proteinase and sonication treatments indicated that although some of this activity was due to adsorbed cytosolic enzymes, there was also an active but latent microsomal PPI activity. Density-gradient subfractionation indicated that activity was associated with vesicles derived from both the rough and the smooth endoplasmic reticulum (ER), suggesting that the activity was located within the ER lumen. The luminal PPI activity was inhibited by cyclosporin A and was active towards an unfolded protein substrate as well as towards the standard peptide substrate. PMID:8010971

  2. 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

  3. New approaches for immunosuppression

    SciTech Connect

    Eiseman, B.; Hansbrough, J.; Weil, R.

    1980-01-01

    New approaches for experimental immunosuppression have been reviewed. These include the following: (1) cyclosporin A, a metabolite from fungus that suppresses multiplying but not resting T and B lymphocytes and can be used in pulsed manner with interspersed drug-free periods; (2) total lymphoid irradiation (transplantation tolerance in rats has been achieved by pretransplant radiation); (3) thoracic duct drainage, which is being revived following its demonstrated effectiveness in the treatment of some autoimmune diseases; (4) hyperbaric oxygen (HBOX). We have found that HBOX 2 1/2 ATA for five hours daily depresses cell-mediated immunity in mice and that this can be reversed by intravenous administration of autologous macrophages.

  4. 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.

  5. Clinical pharmacokinetics and drug-drug interactions of endothelin receptor antagonists in pulmonary arterial hypertension.

    PubMed

    Venitz, Jürgen; Zack, Julia; Gillies, Hunter; Allard, Martine; Regnault, Jean; Dufton, Christopher

    2012-12-01

    The authors review the basic pharmacology and potential for adverse drug-drug interactions (DDIs) of bosentan and ambrisentan, the 2 endothelin receptor antagonists currently approved for pulmonary arterial hypertension (PAH) treatment. Bosentan, an endothelin (ET) receptor-type ET(A) and ET(B) antagonist, is metabolized to active metabolites by and an inducer of cytochrome P450 (CYP)2C9 and CYP3A. Ambrisentan, a selective ET(A) receptor antagonist, is metabolized primarily by uridine 5'diphosphate glucuronosyltransferases (UGTs) 1A9S, 2B7S, and 1A3S and, to a lesser extent, by CYP3A and CYP2C19. Drug interactions observed with bosentan DDI studies have demonstrated a potential for significant clinical implications during PAH management: bosentan is contraindicated with cyclosporine A and glyburide, and additional monitoring/dose adjustments are required when coadministered with hormonal contraceptives, simvastatin, lopinavir/ritonavir, and rifampicin. As bosentan carries a boxed warning regarding risks of liver injury and showed dose-dependant increases in serum aminotransferase abnormalities, drug interactions that increase bosentan exposure are of particular clinical concern. Ambrisentan DDI studies performed to date have shown only one clinically relevant DDI, an interaction with cyclosporine A that requires ambrisentan dose reduction. As the treatment of PAH moves toward multimodal combination therapy, scrutiny should be placed on ensuring that drug combinations achieve maximal clinical benefit while minimizing side effects.

  6. 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

  7. Secreted Cyclophilin A, a Peptidylprolyl cis-trans Isomerase, Mediates Matrix Assembly of Hensin, a Protein Implicated in Epithelial Differentiation*S⃞

    PubMed Central

    Peng, Hu; Vijayakumar, Soundarapandian; Schiene-Fischer, Cordelia; Li, Hui; Purkerson, Jeffrey M.; Malesevic, Miroslav; Liebscher, Jürgen; Al-Awqati, Qais; Schwartz, George J.

    2009-01-01

    Hensin is a rabbit ortholog of DMBT1, a multifunctional, multidomain protein implicated in the regulation of epithelial differentiation, innate immunity, and tumorigenesis. Hensin in the extracellular matrix (ECM) induced morphological changes characteristic of terminal differentiation in a clonal cell line (clone C) of rabbit kidney intercalated cells. Although hensin is secreted in monomeric and various oligomeric forms, only the polymerized ECM form is able to induce these phenotypic changes. Here we report that hensin secretion and matrix assembly were inhibited by the peptidylprolyl cis-trans isomerase (PPIase) inhibitors cyclosporin A (CsA) and a derivative of cyclosporin A with modifications in the d-Ser side chain (Cs9) but not by the calcineurin pathway inhibitor FK506. PPIase inhibition led to failure of hensin polymerization in the medium and ECM, plus the loss of apical cytoskeleton, apical microvilli, and the columnar epithelial shape of clone C cells. Cyclophilin A was produced and secreted into the media to a much greater extent than cyclophilins B and C. Our results also identified the direct CsA-sensitive interaction of cyclophilin A with hensin, suggesting that cyclophilin A is the PPIase that mediates the polymerization and matrix assembly of hensin. These results are significant because this is the first time a direct role of peptidylprolyl cis-trans isomerase activity has been implicated in the process of epithelial differentiation. PMID:19112104

  8. Effects of immunomodulatory drugs on TNF-α and IL-12 production by purified epidermal langerhans cells and peritoneal macrophages

    PubMed Central

    2011-01-01

    Background Langerhans cells constitute a special subset of immature dendritic cells localized in the epidermis that play a key role in the skin's immune response. The production of cytokines is a key event in both the initiation and the regulation of immune responses, and different drugs can be used to remove or modify their production by DC and, therefore, alter immune responses in a broad spectrum of diseases, mainly in human inflammatory and autoimmune diseases. In the present study, we examined the effects of prednisone, thalidomide, cyclosporine A, and amitriptyline, drugs used in a variety of clinical conditions, on the production of TNF-α, IL-10, and IL-12 by purified epidermal Langerhans cells and peritoneal macrophages in BALB/c mice. Findings All drugs inhibited TNF-α production by Langerhans cells after 36 hours of treatment at two different concentrations, while prednisone and thalidomide decreased IL-12 secretion significantly, amitriptyline caused a less pronounced reduction and cyclosporine A had no effect. Additionally, TNF-α and IL-12 production by macrophages decreased, but IL-10 levels were unchanged after all treatments. Conclusions Our results demonstrate that these drugs modulate the immune response by regulating pro-inflammatory cytokine production by purified epidermal Langerhans cells and peritoneal macrophages, indicating that these cells are important targets for immunosuppression in various clinical settings. PMID:21276247

  9. OsCYP21-4, a novel Golgi-resident cyclophilin, increases oxidative stress tolerance in rice.

    PubMed

    Lee, Sang S; Park, Hyun J; Jung, Won Y; Lee, Areum; Yoon, Dae H; You, Young N; Kim, Hyun-Soon; Kim, Beom-Gi; Ahn, Jun C; Cho, Hye S

    2015-01-01

    OsCYP21-4 is a rice cyclophilin protein that binds to cyclosporine A, an immunosuppressant drug. CYP21-4s in Arabidopsis and rice were previously shown to function as mitochondrial cyclophilins, as determined by TargetP analysis. In the current study, we found that OsCYP21-4-GFP localized to the Golgi, rather than mitochondria, in Nicotiana benthamiana leaves, which was confirmed based on its co-localization with cis Golgi α-ManI-mCherry protein. OsCYP21-4 transcript levels increased in response to treatments with various abiotic stresses and the phytohormone abscisic acid, revealing its stress-responsiveness. CYP21-4 homologs do not possess key peptidyl prolyl cis/trans isomerase (PPIase) activity/cyclosporine A (CsA) binding residues, and recombinant OsCYP21-4 protein did not convert the synthetic substrate Suc-AAPF-pNA via cis- trans- isomerization in vitro. In addition, transgenic plants overexpressing OsCYP21-4 exhibited increased tolerance to salinity and hydrogen peroxide treatment, along with increased peroxidase activity. These results demonstrate that OsCYP21-4 is a novel Golgi-localized cyclophilin that plays a role in oxidative stress tolerance, possibly by regulating peroxidase activity. PMID:26483814

  10. 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

  11. 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. PMID:27551545

  12. 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.

  13. The pro-oxidative drug WF-10 inhibits serial killing by primary human cytotoxic T-cells

    PubMed Central

    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. PMID:27551545

  14. 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

  15. A novel cyclophilin from parasitic and free-living nematodes with a unique substrate- and drug-binding domain.

    PubMed

    Ma, Dong; Nelson, Laura S; LeCoz, Krystel; Poole, Catherine; Carlow, Clotilde K S

    2002-04-26

    A highly diversified member of the cyclophilin family of peptidyl-prolyl cis-trans isomerases has been isolated from the human parasite Onchocerca volvulus (OvCYP-16). This 25-kDa cyclophilin shares 43-46% similarity to other filarial cyclophilins but does not belong to any of the groups previously defined in invertebrates or vertebrates. A homolog was also isolated from Caenorhabditis elegans (CeCYP-16). Both recombinant O. volvulus and C. elegans cyclophilins were found to possess an enzyme activity with similar substrate preference and insensitivity to cyclosporin A. They represent novel cyclophilins with important differences in the composition of the drug-binding site in particular, namely, a Glu(124) (C. elegans) or Asp(123) (O. volvulus) residue present in a critical position. Site-directed mutagenesis studies and kinetic characterization demonstrated that the single residue dictates the degree of binding to substrate and cyclosporin A. CeCYP-16::GFP-expressing lines were generated with expression in the anterior and posterior distal portions of the intestine, in all larval stages and adults. An exception was found in the dauer stage, where fluorescence was observed in both the cell bodies and processes of the ventral chord motor neurons but was absent from the intestine. These studies highlight the extensive diversification of cyclophilins in an important human parasite and a closely related model organism.

  16. Influence of immunomodulation on the development of Listeria monocytogenes infection in aged guinea pigs.

    PubMed

    Pang, Hoan-Jen E; Lo, Chih-Yu; Matthews, Karl R

    2007-04-01

    We investigated the impact of immunomodulation on the development of listeriosis within an aged population of guinea pigs after an intragastric challenge with Listeria monocytogenes. Supplementation with vitamin E for 35 days significantly increased the level of cytotoxic T cells (CD8(+)), while treatment with cyclosporin A resulted in a 25% decrease of CD8(+) T cells. In the animals receiving the low dose (10(2) CFU) of L. monocytogenes, 50% of the control-group animals became infected. Only 22% of animals receiving the orthomolecular dose of vitamin E became infected, whereas animals that were immunosuppressed had an infection rate of 89%. In the immunosuppressed group three animals (16%) developed listerial infection with a quantifiable bacterial level of 0.3-3 log CFU g(-1) of organ in the spleen and liver. In the high-dose study, the population of L. monocytogenes was consistently 1 log CFU g(-1) lower in the spleen or liver of the vitamin E-supplemented group, compared with the control and cyclosporin A-treated animals. At day 4, a significant increase in the levels of CD8(+) during listerial infection occurred in vitamin E-supplemented animals, suggesting an increased ability to produce CD8(+) T cells. The results suggest that immunomodulation of the host can influence listerial infection within an aged population of guinea pigs. PMID:17378902

  17. Calcineurin is associated with the cytoskeleton of cultured neurons and has a role in the acquisition of polarity.

    PubMed Central

    Ferreira, A; Kincaid, R; Kosik, K S

    1993-01-01

    Calcineurin is a calmodulin-dependent serine-threonine phosphatase found in many cell types but most abundant in neurons. To determine its localization in developing neurons, dissociated cultures from embryonic day 15 rat cerebellum were analyzed immunocytochemically after treatment with cytoskeletal-disrupting drugs. During the initial outgrowth of neurites, calcineurin is enriched in growth cones where its localization depends upon the integrity of both microtubules and actin filaments. Treatment with cytochalasin shifts calcineurin from the growth cone to the neurite shaft, and with nocadozole calcineurin translocates to the cell body. Therefore calcineurin is well positioned to mediate interactions between cytoskeletal systems during neurite elongation. By 14 d in culture, when the neurons have developed extensive neuronal contacts and synapses are present, calcineurin is predominantly in the neurite shaft. Incubation of cultured cells with Cyclosporin A or a specific peptide, both of which selectively inhibit calcineurin's phosphatase activity, prevented axonal elongation. Because the microtubule-associated protein tau appears to play a key role in asymmetric neurite elongation, we examined modifications in its phosphorylation state resulting from calcineurin inhibition. In contrast to the normal development of cerebellar macroneurons in which reactivity with the phosphorylation-dependent antibody, tau-1, progressively increases, there was a persistent inhibition of tau-1 reactivity in cells exposed to Cyclosporin A. These findings suggest a role for calcineurin in regulating tau phosphorylation and possibly modulating other steps required for the determination of polarity. Images PMID:8167406

  18. Renal hemodynamics in hypertensive renal allograft recipients: effects of calcium antagonists and ACE inhibitors.

    PubMed

    Grekas, D; Dioudis, C; Kalevrosoglou, I; Alivanis, P; Derveniotis, V; Tourkantonis, A

    1996-06-01

    Hypertension present in more than 50% of successfully renal transplanted patients and its prevalence has slightly increased since the introduction of cyclosporine A. Twenty patients, 9 women and 11 men aged from 30 to 58 years, with stable cadaveric renal allograft function and moderate to severe hypertension, were included in the study. Renal artery graft stenosis causing hypertension were excluded. All patients were given triple drug immunosuppressive treatment with methylprednisolone, azathioprine and cyclosporine A (CsA) and their hypertension was treated with a nifedipine dose of 20 mg twice daily. To evaluate the effect of ACE inhibitors on renal hemodynamics and hypertension, a 4 mg/daily dose of perindopril was added to the above regimen for two months. Effective renal plasma flow (ERPF) decreased from 208 +/- 54 to 168 +/- 61 ml/min and renal vascular resistance (RVR) increased from 75 +/- 12 to 88 +/- 17 mm Hg/ml/min (P < 0.05 and P < 0.01, respectively). Mean blood pressure was significantly (P < 0.001) reduced by the combination of both agents in comparison to the blood pressure control by monotherapy with nifedipine. It is suggested that the combination of both antihypertensive agents was more effective than monotherapy with nifedipine in controlling blood pressure, but less favorable on the renal hemodynamic response in hypertensive renal transplant patients who were maintained on CsA.

  19. Respiratory complex I dysfunction due to mitochondrial DNA mutations shifts the voltage threshold for opening of the permeability transition pore toward resting levels.

    PubMed

    Porcelli, Anna Maria; Angelin, Alessia; Ghelli, Anna; Mariani, Elisa; Martinuzzi, Andrea; Carelli, Valerio; Petronilli, Valeria; Bernardi, Paolo; Rugolo, Michela

    2009-01-23

    We have studied mitochondrial bioenergetics in HL180 cells (a cybrid line harboring the T14484C/ND6 and G14279A/ND6 mtDNA mutations of Leber hereditary optic neuropathy, leading to an approximately 50% decrease of ATP synthesis) and XTC.UC1 cells (derived from a thyroid oncocytoma bearing a disruptive frameshift mutation in MT-ND1, which impairs complex I assembly). The addition of rotenone to HL180 cells and of antimycin A to XTC.UC1 cells caused fast mitochondrial membrane depolarization that was prevented by treatment with cyclosporin A, intracellular Ca2+ chelators, and antioxidant. Both cell lines also displayed an anomalous response to oligomycin, with rapid onset of depolarization that was prevented by cyclosporin A and by overexpression of Bcl-2. These findings indicate that depolarization by respiratory chain inhibitors and oligomycin was due to opening of the mitochondrial permeability transition pore (PTP). A shift of the threshold voltage for PTP opening close to the resting potential may therefore be the underlying cause facilitating cell death in diseases affecting complex I activity. This study provides a unifying reading frame for previous observations on mitochondrial dysfunction, bioenergetic defects, and Ca2+ deregulation in mitochondrial diseases. Therapeutic strategies aimed at normalizing the PTP voltage threshold may be instrumental in ameliorating the course of complex I-dependent mitochondrial diseases.

  20. 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.

  1. RCAN1 regulates vesicle recycling and quantal release kinetics via effects on calcineurin activity.

    PubMed

    Zanin, Mark P; Mackenzie, Kimberly D; Peiris, Heshan; Pritchard, Melanie A; Keating, Damien J

    2013-02-01

    We have previously shown that Regulator of Calcineurin 1 (RCAN1) regulates multiple stages of vesicle exocytosis. However, the mechanisms by which RCAN1 affects secretory vesicle exocytosis and quantal release kinetics remain unknown. Here, we use carbon fibre amperometry to detect exocytosis from chromaffin cells and identify these underlying mechanisms. We observe reduced exocytosis with repeated stimulations in chromaffin cells over-expressing RCAN1 (RCAN1(ox)), but not in wild-type (WT) cells, indicating a negative effect of RCAN1 on vesicle recycling and endocytosis. Acute exposure to calcineurin inhibitors, cyclosporine A and FK-506, replicates this effect in WT cells but has no additional effect in RCAN1(ox) cells. When we chronically expose WT cells to cyclosporine A and FK-506 we find that catecholamine release per vesicle and pre-spike foot (PSF) signal parameters are decreased, similar to that in RCAN1(ox) cells. Inhibiting calcineurin activity in RCAN1(ox) cells has no additional effect on the amount of catecholamine release per vesicle but further reduces PSF signal parameters. Although electron microscopy studies indicate these changes are not because of altered vesicle number or distribution in RCAN1(ox) cells, the smaller vesicle and dense core size we observe in RCAN1(ox) cells may underlie the reduced quantal release in these cells. Thus, our results indicate that RCAN1 most likely affects vesicle recycling and quantal release kinetics via the inhibition of calcineurin activity.

  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. 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

    2016-01-01

    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

  4. 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

  5. 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

  6. A screen for apoptotic synergism between clinical relevant nephrotoxicant and the cytokine TNF-α.

    PubMed

    Benedetti, Giulia; Ramaiahgaris, Sreenivasa; Herpers, Bram; van de Water, Bob; Price, Leo S; de Graauw, Marjo

    2013-12-01

    Nephrotoxicity remains one of the main reasons for post-market drug withdrawal. Tumour necrosis factor α (TNF-α) secretion has been shown to underlie the nephrotoxicity induced by some of these drugs. Yet, there is currently no reliable and sensitive in vitro assay available to screen for nephrotoxicants of which toxicity largely depends on TNF-α secretion. Therefore, we developed and applied a sensitive fluorescence-based in vitro assay for TNF-α-mediated nephrotoxicity screening using mouse immortalized proximal tubular epithelial cells (IM-PTECs). Our assay allows rapid evaluation of TNF-α-mediated toxicant-induced apoptosis and necrosis using fixed endpoint and live cell measurements. To evaluate our assay, sixteen nephrotoxicants and two control non-nephrotoxicants were used. Out of the sixteen nephrotoxicants, eight induced cell death, of which five induced apoptosis as well as necrosis. Moreover, TNF-α significantly enhanced apoptotic cell death induced by cisplatin, cyclosporine A, tacrolimus and azidothymidine. These nephrotoxicants are known to induce inflammation in vivo which has been linked to an enhancement of nephrotoxicity for cisplatin, cyclosporine A and tacrolimus, confirming the functionality of our assay. Overall, our assay allows rapid and sensitive measurement of apoptosis and necrosis induced by a combination of nephrotoxicants and inflammatory components such as TNF-α and can be used as an alternative assay for nephrotoxicity prediction in vitro.

  7. Signaling pathway involved in the immunomodulatory effect of Ganoderma atrum polysaccharide in spleen lymphocytes.

    PubMed

    Yu, Qiang; Nie, Shao-Ping; Wang, Jun-Qiao; Huang, Dan-Fei; Li, Wen-Juan; Xie, Ming-Yong

    2015-03-18

    The aim of this study was to investigate the molecular mechanism underlying the immunomodulatory effect of Ganoderma atrum polysaccharide (PSG-1) in spleen lymphocytes. Our results showed that PSG-1 increased the intracellular Ca2+ concentration and calcineurin (CaN) activity. Moreover, PSG-1 was found to elevate nuclear factor of activated T cells (NFAT) activity, but this effect could be diminished by the treatment of CaN inhibitors (cyclosporin A and FK506). PSG-1-induced interleukin (IL)-2 production was also inhibited by cyclosporin A and FK506. In addition, PSG-1 was found to significantly enhance protein kinase C (PKC) activity. PKC was involved in induction of NFAT activity by PSG-1, as evidenced by abrogation of NFAT activity by PKC inhibitor calphostin C, which significantly decreased PSG-1-induced IL-2 production. On the basis of these results, we concluded that PSG-1 may induce activation of spleen lymphocytes at least in part via the Ca2+/CaN/NFAT/IL-2 signaling pathway and the PKC/NFAT/IL-2 signaling pathway cooperatively regulated PSG-1-induced activation of spleen lymphocytes.

  8. Minocycline, a possible neuroprotective agent in Leber's hereditary optic neuropathy (LHON): studies of cybrid cells bearing 11,778 mutation.

    PubMed

    Haroon, Mohammad Fahad; Fatima, Ambrin; Schöler, Susanne; Gieseler, Anne; Horn, Thomas F W; Kirches, Elmar; Wolf, Gerald; Kreutzmann, Peter

    2007-12-01

    Leber's hereditary optic neuropathy (LHON) is a retinal neurodegenerative disorder caused by mitochondrial DNA point mutations. Complex I of the respiratory chain affected by the mutation results in a decrease in ATP and an increase of reactive oxygen species production. Evaluating the efficacy of minocycline in LHON, the drug increased the survival of cybrid cells in contrast to the parental cells after thapsigargin-induced calcium overload. Similar protection was observed by treatment with cyclosporine A, a blocker of the mitochondrial permeability transition pore (mPTP). Ratiometric Ca(2+) imaging reveals that acetylcholine/thapsigargin triggered elevation of the cytosolic calcium concentration is alleviated by minocycline and cyclosporine A. The mitochondrial membrane potential of LHON cybrids was significantly conserved and the active-caspase-3/procaspase-3 ratio was decreased in both treatments. Our observations show that minocycline inhibits permeability transition induced by thapsigargin in addition to its antioxidant effects. In relation with its high safety profile, these results would suggest minocycline as a promising neuroprotective agent in LHON.

  9. Novel Molecular Imaging Approach for Subclinical Detection of Iritis and Evaluation of Therapeutic Success

    PubMed Central

    Xie, Fang; Sun, Dawei; Schering, Alexander; Nakao, Shintaro; Zandi, Souska; Liu, Ping; Hafezi-Moghadam, Ali

    2010-01-01

    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. PMID:20581051

  10. Differential diagnosis of acute rejection and chronic cyclosporine nephropathy after rat renal transplantation by detection of endothelial microparticles (EMP).

    PubMed

    Cui, Jiewei; Yang, Jing; Cao, Weike; Sun, Yi

    2010-12-01

    Endothelial microparticles (EMP) are small vesicles smaller than 1.0μm, released from endothelial cells (EC) during their activation and (or) apoptosis. The assay of the level of elevated EMP is a new approach to evaluate the dysfunction of endothelial cell. EMP can be classified into several types according to their membrane molecular, and the levels of various types of EMP may be different. As the most cost-effective immunodepressant, cyclosporine A (CsA) has been used widely in organ transplantation. But its dose is hard to control, under-medication may cause the acute rejection (AR) and overdose may cause chronic cyclosporine nephropathy (CCN). The cyclosporine A (CsA) caused CCN and the AR caused renal injury after renal transplantation are both vascular diseases related with endothelial dysfunction, and up to now, there is still no effective method to distinguish the two kinds of diseases. Owing to distinct pathogenesis of the two kinds of vascular diseases, the level of each type of EMP originated from vascular endothelial cells may be different. We hypothesize that maybe we can distinguish them by detecting the different levels of some types of EMP which is also related with vascular disease, and we propose to prove our hypothesis through animal experiment. If our hypothesis is proved, it will be more helpful for clinicians to adjust the dose of CsA promptly according to the differential diagnosis of the two kinds of diseases.

  11. 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.

  12. 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

    2015-01-01

    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.

  13. Sperm calcineurin inhibition prevents mouse fertility with implications for male contraceptive.

    PubMed

    Miyata, Haruhiko; Satouh, Yuhkoh; Mashiko, Daisuke; Muto, Masanaga; Nozawa, Kaori; Shiba, Kogiku; Fujihara, Yoshitaka; Isotani, Ayako; Inaba, Kazuo; Ikawa, Masahito

    2015-10-23

    Calcineurin inhibitors, such as cyclosporine A and FK506, are used as immunosuppressant drugs, but their adverse effects on male reproductive function remain unclear. The testis expresses somatic calcineurin and a sperm-specific isoform that contains a catalytic subunit (PPP3CC) and a regulatory subunit (PPP3R2). We demonstrate herein that male mice lacking Ppp3cc or Ppp3r2 genes (knockout mice) are infertile, with reduced sperm motility owing to an inflexible midpiece. Treatment of mice with cyclosporine A or FK506 creates phenocopies of the sperm motility and morphological defects. These defects appear within 4 to 5 days of treatment, which indicates that sperm-specific calcineurin confers midpiece flexibility during epididymal transit. Male mouse fertility recovered a week after we discontinued treatment. Because human spermatozoa contain PPP3CC and PPP3R2 as a form of calcineurin, inhibition of this sperm-specific calcineurin may lead to the development of a reversible male contraceptive that would target spermatozoa in the epididymis.

  14. [Resistance to cytostatic agents in patients with refractory multiple myeloma and possible ways to control it].

    PubMed

    Adam, Z; Vorlícek, J

    1993-04-01

    At present several mechanisms responsible for the resistance of tumour cells to cytostatics are known. Most explored is the resistance conditioned by glycoprotein-P. Under laboratory conditions it proved possible to inhibit the resistance caused by glycoprotein-P by many substances which, however, on clinical administration have serious adverse side-effects. In practice verapamil was used, its effect was most marked in haematological malignant diseases. Side-effects of verapamil, however, do not permit to attain the necessary concentration. Obviously better results will be achieved by cyclosporin A which can attain a concentration in the organism which is necessary to inhibit glycoprotein-P without serious undesirable consequences. In the submitted paper the author reviews clinical studies concerned with possible inhibition of glycoprotein-P in patients with refractory myeloma.

  15. Cardioprotection in the clinical setting.

    PubMed

    Ivanes, Fabrice; Mewton, Nathan; Rioufol, Gilles; Piot, Christophe; Elbaz, Meyer; Revel, Didier; Croisille, Pierre; Ovize, Michel

    2010-06-01

    Reperfusion therapy is the primary treatment of acute myocardial infarction and must be applied as soon as possible to limit the ischemic insult. Unfortunately, reperfusion is responsible for additional myocardial damage likely involving opening of the mitochondrial permeability transition pore. Ischemic postconditioning is a powerful intervention that dramatically reduces lethal reperfusion injury. Several clinical studies using angioplasty postconditioning now support its protective effects in patients with an acute myocardial infarction. Alternatively, pharmacological postconditioning could afford comparable protection and be applied to a much larger number of patients. Indeed, the mitochondrial permeability transition pore inhibitor cyclosporine A has been shown to generate a similar protection in acute myocardial infarction patients. Future large-scale trials are needed to determine whether angioplasty or pharmacological postconditioning may improve clinical outcome in STEMI patients. PMID:20589423

  16. Cytomegalovirus keratitis after penetrating keratoplasty.

    PubMed

    Wehrly, S R; Manning, F J; Proia, A D; Burchette, J L; Foulks, G N

    1995-11-01

    We report the development of cytomegalovirus (CMV) keratitis in the penetrating keratoplasty of a 59-year-old human immunodeficiency virus-negative woman after uncomplicated corneal transplantation. Immunosuppression with topical cyclosporine A 2% in corn oil and topical prednisolone acetate 1% suspension was used postoperatively. The 15-month postoperative course was complicated by multiple episodes of endothelial rejection, medically controlled elevated intraocular pressure, polymicrobial bacterial (coagulase-negative staphlococcus and alpha-hemolytic streptococcus) keratitis, and endothelial plaque formation with associated hypopyon and epithelial defect. The graft failed and penetrating keratoplasty was repeated. Cytomegalovirus infection of superficial keratocytes in a region of scarring was identified in histological sections stained with hematoxylin and eosin and confirmed using mouse monoclonal anti-cytomegalovirus antibodies. Excision of the diseased corneal button with no additional treatment appears to have been curative. Low-grade keratitis was the only manifestation of the CMV infection, and it has not recurred 6 months postoperatively.

  17. Treatment of rheumatic diseases in patients with HCV and HIV infection.

    PubMed

    Galeazzi, Mauro; Giannitti, Chiara; Manganelli, Stefania; Benucci, Maurizio; Scarpato, Salvatore; Bazzani, Chiara; Caporali, Roberto; Sebastiani, Gian Domenico

    2008-12-01

    A wide variety of rheumatic diseases has been documented in the presence of hepatitis C virus (HCV) infection and in human immunodeficiency virus (HIV) infection. In this conditions, physicians are refrained from using corticosteroids and/or immunosuppressants agents because of the risk of favouring viral replication and the progression of the underlying viral disease. In the present review we have focused our attention on the possible role of cyclosporine A (CsA), anti-Tumour Necrosis Factor (TNF) alpha agents in the treatment of HIV or HCV infected autoimmune patients. The results drown from the literature and from our personal experience confirm the safety of CsA and anti-TNF alpha agents, in terms of viral load and liver toxicity. A limited experience also suggest that both therapies can be given in combination in rheumatoid arthritis patients without increasing the risk of adverse events.

  18. Acquired pure red cell aplasia: updated review of treatment

    PubMed Central

    Sawada, Kenichi; Fujishima, Naohito; Hirokawa, Makoto

    2008-01-01

    Pure red cell aplasia (PRCA) is a syndrome characterized by a severe normocytic anaemia, reticulocytopenia, and absence of erythroblasts from an otherwise normal bone marrow. Primary PRCA, or secondary PRCA which has not responded to treatment of the underlying disease, is treated as an immunologically-mediated disease. Although vigorous immunosuppressive treatments induce and maintain remissions in a majority of patients, they carry an increased risk of serious complications. Corticosteroids were used in the treatment of PRCA and this has been considered the treatment of first choice although relapse is not uncommon. Cyclosporine A (CsA) has become established as one of the leading drugs for treatment of PRCA. However, common concerns have been the number of patients treated with CsA who achieve sustained remissions and the number that relapse. This article reviews the current status of CsA therapy and compares it to other treatments for diverse PRCAs. PMID:18510682

  19. Immune Pathways in Atopic Dermatitis, and Definition of Biomarkers through Broad and Targeted Therapeutics.

    PubMed

    Mansouri, Yasaman; Guttman-Yassky, Emma

    2015-04-29

    Atopic dermatitis (AD) is the most common inflammatory skin disease. Recent research findings have provided an insight into the complex pathogenic mechanisms involved in this disease. Despite a rising prevalence, effective and safe therapeutics for patients with moderate-to-severe AD are still lacking. Biomarkers of lesional, nonlesional skin, and blood have been developed for baseline as well as after treatment with broad and specific treatments (i.e., cyclosporine A and dupilumab). These biomarkers will help with the development of novel targeted therapeutics and assessment of disease reversal, with the promise of a more personalized treatment approach. Since AD involves more than one subtype (i.e., intrinsic/extrinsic, pediatric/adult, etc.), these molecular fingerprints needs to be validated in all subpopulations with AD.

  20. Pharmacogenomics and personalized medicine: a review focused on their application in the Chinese population.

    PubMed

    Shu, Wen-ying; Li, Jia-li; Wang, Xue-ding; Huang, Min

    2015-05-01

    The field of pharmacogenomics was initiated in the 1950s and began to thrive after the completion of the human genome project 10 years ago. Thus far, more than 100 drug labels and clinical guidelines referring to pharmacogenomic biomarkers have been published, and several key pharmacogenomic markers for either drug safety or efficacy have been identified and subsequently adopted in clinical practice as pre-treatment genetic tests. However, a tremendous variation of genetic backgrounds exists between different ethnic groups. The application of pharmacogenomics in the Chinese population is still a long way off, since the published guidelines issued by the organizations such as US Food and Drug Administration require further confirmation in the Chinese population. This review highlights important pharmacogenomic discoveries in the Chinese population and compares the Chinese population with other nations regarding the pharmacogenomics of five most commonly used drugs, ie, tacrolimus, cyclosporine A, warfarin, cyclophosphamide and azathioprine.

  1. 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

  2. Calcineurin Undergoes a Conformational Switch Evoked via Peptidyl-Prolyl Isomerization

    PubMed Central

    Guasch, Alicia; Aranguren-Ibáñez, Álvaro; Pérez-Luque, Rosa; Aparicio, David; Martínez-Høyer, Sergio; Mulero, M. Carmen; Serrano-Candelas, Eva

    2015-01-01

    A limited repertoire of PPP family of serine/threonine phosphatases with a highly conserved catalytic domain acts on thousands of protein targets to orchestrate myriad central biological roles. A major structural reorganization of human calcineurin, a ubiquitous Ser/Thr PPP regulated by calcium and calmodulin and targeted by immunosuppressant drugs cyclosporin A and FK506, is unveiled here. The new conformation involves trans- to cis- isomerization of proline in the SAPNY sequence, highly conserved across PPPs, and remodels the main regulatory site where NFATc transcription factors bind. Transitions between cis- and trans- conformations may involve peptidyl prolyl isomerases such as cyclophilin A and FKBP12, which are known to physically interact with and modulate calcineurin even in the absence of immunosuppressant drugs. Alternative conformations in PPPs provide a new perspective on interactions with substrates and other protein partners and may foster development of more specific inhibitors as drug candidates. PMID:26248042

  3. Tracing fungi secondary metabolites in Brazil nuts using LC-MS/MS.

    PubMed

    Freitas-Silva, Otniel; de Lourdes, Maria; de Souza, Mendes; Venãncio, Armando

    2011-08-01

    This screening aimed to evaluate quantitatively the occurrence of fungal metabolites in Brazil nuts. Nuts were collected from Agroforest production areas in Amazon basin region. A total of 235 mycotoxins (including the most prominent ones) was screened by a multi-mycotoxin method based on HPLC-MS/MS. The recovery of metabolites by the method was between 56 and 136%. Fifteen mycotoxins were detected and quantified, in at least one sample; namely, aflatoxins (AFB(1), AFB(2), AFG(1), and AFM(1)), sterigmatocystin, methyl-sterigmatocystin, kojic acid, citrinin, cyclosporin A, cyclosporin C, cyclosporin D, cyclosporin H, rugulosin, alternariol-methylether and emodin. This is the first study dealing with the detection of the latter nine metabolites in Brazil nuts. Alternariol-methylether (from 0.75 to 3.2 µg/kg) was the only metabolite detected in all analyzed samples. PMID:21722090

  4. Treatment of myelodysplasia with oral cyclosporin.

    PubMed

    Atoyebi, W; Bywater, L; Rawlings, L; Brunskill, S; Littlewood, T J

    2002-08-01

    Recent studies have shown a good response to immunosuppressive treatment with cyclosporin A (CSA) in patients with the myelodysplastic syndrome (MDS). We have treated six transfusion-dependent MDS patients with CSA for a minimum of 3 months. None of these patients showed a significant response, while the drug was withdrawn in 3/6 patients because of intolerable side-effects. Two reasons for the failure of this treatment in our patients can be advanced. Firstly, the hypoplastic variant of MDS predominated in previous studies in contrast to ours. Secondly, the concomitant use of other immunosuppressive agents in previous studies might have enhanced the effect of CSA. We suggest further therapeutic trials of CSA in MDS, selecting patients on the basis of in vitro studies that predict an immunological basis for their disease, to assess its efficacy in prolonging survival. PMID:12181023

  5. [Immunosuppressives to prevent rejection reactions after allogeneic corneal transplantation].

    PubMed

    Lapp, T; Maier, P; Birnbaum, F; Schlunck, G; Reinhard, T

    2014-03-01

    In order to prevent rejection of an allogeneic corneal transplant after perforating (high risk) keratoplasty, active agents from different classes of pharmacological substances are used, as with solid organ transplantation. In addition to glucocorticoids, antiproliferative agents, small molecule inhibitors and antibodies, those belonging to the group of macrolides with their many derivatives represent an interesting class of substances in this context. As a supplement to cyclosporin A (CSA) the most successful macrolide in transplantation medicine, animal experiments are currently being carried out to test newer macrolide derivatives, such as sanglifehrin A (SFA). This overview describes the classes of drugs and modes of action of currently administered standard medications in the clinical routine and new developments are presented.

  6. A new model of corneal transplantation in the miniature pig: efficacy of immunosuppressive treatment.

    PubMed

    Tavandzi, Urania; Procházka, Radek; Usvald, Dusan; Hlucílová, Jana; Vitásková, Martina; Motlík, Jan; Vítová, Andrea; Filipec, Martin; Forrester, John V; Holán, Vladimír

    2007-05-27

    Corneal allograft rejection is frequently studied in small rodent or rabbit models. To study mechanisms of rejection in a model that more closely mimics transplantation in humans, we performed orthotopic corneal transplantation in the miniature pig using a 7-mm diameter donor graft. Four groups of recipients were studied: 1) untreated naive, 2) untreated vascularized (high risk), 3) high-risk grafts treated by topical application of prednisolone, or 4) high-risk grafts treated with a combined systemic immunosuppression regime of oral prednisone, cyclosporine A, and mycophenolate mofetil. Both the clinical features and histological assessment of corneal graft rejection showed close similarities to graft rejection in humans. Interestingly, preliminary results indicated that topical steroid treatment was superior to systemic immunosuppression in significantly promoting graft survival. Thus, corneal transplantation in the pig represents an animal model most closely resembling corneal grafting in humans, and offers possibilities for testing various clinically applicable immunosuppressive treatments.

  7. Activation of the Ca{sup 2+}/calcineurin/NFAT{sub 2} pathway controls smooth muscle cell differentiation

    SciTech Connect

    Larrieu, Daniel; Thiebaud, Pierre; Duplaa, Cecile; Sibon, Igor; Theze, Nadine; Lamaziere, Jean-Marie Daniel . E-mail: jean-marie.d-lamaziere@bordeaux.inserm.fr

    2005-10-15

    Cellular mechanisms controlling smooth muscle cells (SMCs) phenotypic modulation are largely unknown. Intracellular Ca{sup 2+} movements are essential to ensure SMC functions; one of the roles of Ca{sup 2+} is to regulate calcineurin, which in turn induces nuclear localization of the nuclear factor of activated T-cell (NFAT). In order to investigate, during phenotypic differentiation of SMCs, the effect of calcineurin inhibition on NFAT{sub 2} nuclear translocation, we used a culture model of SMC differentiation in serum-free conditions. We show that the treatment of cultured SMC with the calcineurin inhibitor cyclosporine A induced their dedifferentiation while preventing their differentiation. These findings suggest that nuclear translocation of NFAT{sub 2} is dependent of calcineurin activity during the in vitro SMC differentiation kinetic and that the nuclear presence of NFAT{sub 2} is critical in the acquisition and maintenance of SMC differentiation.

  8. Macrocyclic drugs and clinical candidates: what can medicinal chemists learn from their properties?

    PubMed

    Giordanetto, Fabrizio; Kihlberg, Jan

    2014-01-23

    Macrocycles are ideal in efforts to tackle "difficult" targets, but our understanding of what makes them cell permeable and orally bioavailable is limited. Analysis of approximately 100 macrocyclic drugs and clinical candidates revealed that macrocycles are predominantly used for infectious disease and in oncology and that most belong to the macrolide or cyclic peptide class. A significant number (N = 34) of these macrocycles are administered orally, revealing that oral bioavailability can be obtained at molecular weights up to and above 1 kDa and polar surface areas ranging toward 250 Å(2). Moreover, insight from a group of "de novo designed" oral macrocycles in clinical studies and understanding of how cyclosporin A and model cyclic hexapeptides cross cell membranes may unlock wider opportunities in drug discovery. However, the number of oral macrocycles is still low and it remains to be seen if they are outliers or if macrocycles will open up novel oral druggable space.

  9. [Effect of blockade of opiate receptors by naloxone on lutropin (LH), follitropin (FSH) and testosterone secretion in patients after kidney transplantation].

    PubMed

    Grzeszczak, W; Zukowska-Szczechowska, E; Kokot, F

    1992-01-01

    The influence of opioid receptors blockade by naloxone on lutropin (LH), follitropin (FSH) and testosterone secretion induced by LH-RH was assessed in 12 male kidney transplant patients with stable graft function (KTP) treated by cyclosporine A and prednisone and in 15 healthy subjects. In KTP normal plasma levels of LH in spite of significantly reduced testosteronemia and a reduced response of LH to LH-RH was observed. After blockade of opioid receptors by naloxone normalization of the response of LH secretion to LH-RH was found and a higher increase of the plasma testosterone level was observed in KTP than in healthy subjects. Results obtained in this study suggest abnormal functioning of the hypothalamic-pituitary--gonadal axis in KTP and participation of opioid receptors in its pathogenesis.

  10. 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

  11. Metabolism of tacrolimus (FK506) and recent topics in clinical pharmacokinetics.

    PubMed

    Iwasaki, Kazuhide

    2007-10-01

    Tacrolimus (FK506), an immunosuppressive drug, is co-medicated with multiple drugs under clinical conditions. Tacrolimus is highly lipophilic and is excreted from the body after receiving extensive metabolism. Due to its narrow therapeutic window following organ transplantation, tacrolimus requires therapeutic drug monitoring by an enzyme immunoassay using the monoclonal antibody raised against tacrolimus. Therefore, metabolism studies including drug-drug interaction and metabolite identification studies are essential for the efficient development and clinically optimal usage of this drug. Tacrolimus was metabolized by the cytochrome P450 (CYP) 3A subfamily. Metabolic drug-drug interaction studies were conducted to provide information regarding the optimal usage of tacrolimus, and its metabolism was inhibited by known CYP3A inhibitors such as ketoconazole, cyclosporine A, and nifedipine. Recent reports on clinical pharmacokinetics indicate that dose levels of tacrolimus need to be adjusted in transplant patients with CYP3A5 polymorphism. PMID:17965516

  12. 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

  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. Dry eye: diagnosis and current treatment strategies.

    PubMed

    O'Brien, Paul D; Collum, Louis M T

    2004-07-01

    One in four patients attending ophthalmic clinics report symptoms of dry eye, making it one of the most common complaints seen by ophthalmologists. Aqueous-layer deficiency is the most common form of dry eye and is frequently caused by decreased secretion of tears by the lacrimal glands. Evaporative dry eye is often secondary to meibomian gland disease and results in a defective lipid layer. Tear replacement or preservation using artificial tears and/or punctal occlusion are the mainstay of treatment. Newer forms of therapy were designed to modify the underlying disease process. These include the use of topical cyclosporin A, autologous serum, and sodium hyaluronate drops, which suppress underlying inflammation, provide growth factors, and prevent the onset of squamous metaplasia in ocular surface epithelium. Hormonal therapy might have a role in the future of dry eye therapy.

  15. Safety and Efficacy of Cyclosporine in the Treatment of Chronic Dry Eye

    PubMed Central

    Schultz, Clyde

    2014-01-01

    Dry-eye syndrome (DES) is a multifactorial disease affecting millions of individuals worldwide. Various factors, including age, hormonal status, genetics, sex, immune status, innervation status, nutrition, pathogens, and environmental stress, can alter the cellular and molecular structure or function of components of the ocular surface system. The resulting imbalance increases susceptibility to desiccation and epithelial damage, leading to a vicious circle in which inflammation amplifies and sustains further damage by chronic deregulation of the system. Lubricating agents and steroids have been used as treatment options. However, as the causes of the disease become better elucidated, the more chemically complex cyclosporine A has become an increasingly useful treatment option and in the United States is currently the only Food and Drug Administration (FDA)-approved prescription drug for the treatment of dry eye. The safety and efficacy of cyclosporine have been shown in numerous studies. PMID:25002818

  16. 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.

  17. Peripheral benzodiazepine receptor regulates vascular endothelial activations via suppression of the voltage-dependent anion channel-1.

    PubMed

    Joo, Hee Kyoung; Lee, Yu Ran; Lim, Sun Young; Lee, Eun Ji; Choi, Sunga; Cho, Eun Jung; Park, Myoung Soo; Ryoo, Sungwoo; Park, Jin Bong; Jeon, Byeong Hwa

    2012-05-01

    Peripheral benzodiazepine receptor (PBR) is a multifunctional protein mainly found on the outer mitochondrial membrane. PBR expression is increased by tumor necrosis factor-α (TNF-α) in endothelial cells. Adenoviral overexpression of PBR inhibits monocyte adhesion, VCAM-1, and ICAM-1 expression in TNF-α-activated endothelial cells. Rotenone, cyclosporine A, and bongkrekic acid suppress TNF-α-induced VCAM-1 expression. Overexpression of PBR inhibits voltage-dependent anion channel-1 (VDAC-1) expression and the silencing of PBR increases VDAC-1 expression in endothelial cells. Moreover, TNF-α-induced VCAM-1 expression is suppressed by VDAC-1 gene silencing. PBR overexpression significantly decreases TNF-α-induced mitochondrial reactive oxygen species and MnSOD expression. These results suggest that PBR can inhibit endothelial activation and this action is related to the inhibition of mitochondrial ROS and/or VDAC-1 expression in endothelial cells.

  18. 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.

  19. Ledipasvir and sofosbuvir for treatment of post- renal transplant hepatitis C infection: A case report with review of literature.

    PubMed

    Jha, R; Fatima, R; Lakhtakia, S; Jha, A; Srikant, P; Narayan, G

    2016-01-01

    Liver disease due to hepatitis C infection in renal transplant recipients is difficult to treat and often associated with reduced patient survival. A 43-year-old male, a renal allograft recipient, presented at 6 years follow-up with significant weight loss over 3 months. He was detected to have new onset diabetes mellitus together with hepatitis C virus (HCV) infection (genotype 1). His HCV load remained high despite the change of immuno-suppression from tacrolimus to cyclosporine. A decision to treat with a new anti-viral combination of ledipasvir and sofosbuvir for 12 weeks was taken. Within 3 weeks, his raised serum transaminases levels normalized and viral load became undetectable. At the end of 16 weeks, he continues to do well with normal renal function, has sustained remission from hepatitis C infection and resolution of diabetes.

  20. 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.

  1. 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

  2. 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.

  3. 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.

  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. 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. 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.

  7. Ledipasvir and sofosbuvir for treatment of post- renal transplant hepatitis C infection: A case report with review of literature

    PubMed Central

    Jha, R.; Fatima, R.; Lakhtakia, S.; Jha, A.; Srikant, P.; Narayan, G.

    2016-01-01

    Liver disease due to hepatitis C infection in renal transplant recipients is difficult to treat and often associated with reduced patient survival. A 43-year-old male, a renal allograft recipient, presented at 6 years follow-up with significant weight loss over 3 months. He was detected to have new onset diabetes mellitus together with hepatitis C virus (HCV) infection (genotype 1). His HCV load remained high despite the change of immuno-suppression from tacrolimus to cyclosporine. A decision to treat with a new anti-viral combination of ledipasvir and sofosbuvir for 12 weeks was taken. Within 3 weeks, his raised serum transaminases levels normalized and viral load became undetectable. At the end of 16 weeks, he continues to do well with normal renal function, has sustained remission from hepatitis C infection and resolution of diabetes. PMID:27194839

  8. 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

  9. New-onset diabetes mellitus after pediatric liver transplantation.

    PubMed

    Regelmann, Molly O; Goldis, Marina; Arnon, Ronen

    2015-08-01

    In the first five yr after liver transplant, approximately one in 10 pediatric recipients will develop NODAT. Factors associated with higher risk for NODAT have been difficult to identify due to lack of uniformity in reporting and data collection. Limited studies have reported higher risk in those who are at an older age at transplant, those with high-risk ethnic backgrounds, and in those with particular underlying conditions, such as CF and primary sclerosing cholangitis. Immunosuppressive medications, including tacrolimus, cyclosporine A, GC, and sirolimus, have been implicated as contributing to NODAT, to varying degrees. Identifying those at highest risk, appropriately screening, and diagnosing NODAT is critical to initiating timely treatment and avoiding potential complications. In the pediatric population, treatment is limited primarily to insulin, with some consideration for metformin. Children with NODAT should be monitored carefully for complications of DM, including microalbuminuria, hypertension, hyperlipidemia, and retinopathy.

  10. The mitochondrial permeability transition from yeast to mammals

    PubMed Central

    Azzolin, Luca; von Stockum, Sophia; Basso, Emy; Petronilli, Valeria; Forte, Michael A.; Bernardi, Paolo

    2010-01-01

    Regulated permeability changes have been detected in mitochondria across species. We review here their key features, with the goal of assessing whether a “permeability transition” similar to that observed in higher eukaryotes is present in other species. The recent discoveries (i) that treatment with cyclosporin A unmasks an inhibitory site for Pi [Basso et al. (2008) J. Biol Chem. 283, 26307–26311], the classical inhibitor of the permeability transition of yeast; and (ii) that under proper experimental conditions a matrix Ca2+-dependence can be demonstrated in yeast as well [Yamada et al. (2009) Biochim. Biophys. Acta 1787, 1486–1491] suggest that the mitochondrial permeability transition has been conserved during evolution. PMID:20398660

  11. Itch in Atopic Dermatitis Management.

    PubMed

    Kamata, Yayoi; Tominaga, Mitsutoshi; Takamori, Kenji

    2016-01-01

    Patients with atopic dermatitis (AD) suffer from chronic inflammatory dermatitis and antihistamine-resistant itch. The management of intractable pruritus in AD is important, requiring the development of new therapeutic approaches. At present, the standard treatments for AD include topical anti-inflammatory drugs such as calcineurin inhibitors and corticosteroids. Topical emollient treatment is recommended to moisten the skin and to restore and maintain barrier function. Phototherapy is also effective in reducing the number of epidermal nerve fibers, normalizing imbalances in the levels of expression of axon guidance molecules, and inhibiting pruritus. Systemic treatments such as cyclosporine A and aprepitant are used to treat severe and intractable pruritus in AD. Clinical trials of dupilumab and CIM331 have displayed a significant reduction of pruritus in patients with AD. New antipruritic approaches are targeted to the central nervous system such as spinal interneurons and glial cells. This chapter describes therapeutic approaches for attenuating intractable itch in AD. PMID:27578076

  12. 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.

  13. [Dry eye. An update on epidemiology, diagnosis, therapy and new concepts].

    PubMed

    Schirra, F; Ruprecht, K W

    2004-01-01

    Recent epidemiologic studies quote a prevalence of 5,2% to 63% for dry eye depending on definition. Many risk factors have been identified, among other things the female gender. Dry eye interferes significantly with quality of live. Measurement of the change in temperature and humidity during blinking turned out to be a reliable diagnostic tool. Videokeratoscopy explains well-known visual impairments related to dry eye and, along with lipid film interferometry, provides insight into tear film dynamics. The importance of tear film proteins is underestimated. Among therapeutics for symptomatic relief hyaluronic acid proved to be particularly useful but also the sequence of lid hygiene, warm compresses and lid massage is fundamentally important. The effectiveness of punctum plugs is ascribed considerably to a more efficient impact of essential tear film components. Topical cyclosporine A, INS365, 15(S)-HETE as well as topical androgens represent a whole new class of drugs for causal therapy of dry eye.

  14. 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.

  15. Discovery of a potent peptidic cyclophilin A inhibitor Trp-Gly-Pro.

    PubMed

    Pang, Xiaodong; Zhang, Mingjun; Zhou, Linxiang; Xie, Fang; Lu, Hong; He, Wu; Jiang, Shibo; Yu, Long; Zhang, Xinyi

    2011-05-01

    Through virtual screening of a rationally built database consisting of 40 peptides, we identified three short peptides. After testing these three synthetic peptides, we found that the peptide Trp-Gly-Pro (WGP) showed comparable inhibitory ability as positive control cyclosporine A (CsA) on CypA-mediated PPIase activity with IC50 values of 33.11 nM and 10.25 nM, respectively. The peptide WGP had same order of CypA-binding affinity as CsA with dissociation equilibrium constant KD of 3.41×10(-6) and 6.42×10(-6) M, respectively. This peptide could also inhibit HIV-1IIIB infection. This study provides a novel strategy for rational design and development of peptidic drugs.

  16. 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.

  17. Immune Pathways in Atopic Dermatitis, and Definition of Biomarkers through Broad and Targeted Therapeutics

    PubMed Central

    Mansouri, Yasaman; Guttman-Yassky, Emma

    2015-01-01

    Atopic dermatitis (AD) is the most common inflammatory skin disease. Recent research findings have provided an insight into the complex pathogenic mechanisms involved in this disease. Despite a rising prevalence, effective and safe therapeutics for patients with moderate-to-severe AD are still lacking. Biomarkers of lesional, nonlesional skin, and blood have been developed for baseline as well as after treatment with broad and specific treatments (i.e., cyclosporine A and dupilumab). These biomarkers will help with the development of novel targeted therapeutics and assessment of disease reversal, with the promise of a more personalized treatment approach. Since AD involves more than one subtype (i.e., intrinsic/extrinsic, pediatric/adult, etc.), these molecular fingerprints needs to be validated in all subpopulations with AD. PMID:26239452

  18. Validation of quinidine as a probe substrate for the in vitro P-gp inhibition assay in Caco-2 cell monolayer.

    PubMed

    Patil, Anand G; D'Souza, Russell; Dixit, Neeta; Damre, Anagha

    2011-09-01

    Although quinidine has been recommended as a probe substrate for the P-gp inhibition assay using Caco-2 cell monolayer, it has not been studied widely in the in vitro system. In the present investigation, in vitro permeability studies using Caco-2 cell monolayer were carried out in order to optimize and validate quinidine as a P-gp probe substrate. In bi-directional Caco-2 assay across different passages, a good efflux ratio of more than ten was consistently obtained at 100 nM donor concentration of quinidine. Quinidine was found to have a good mass balance in the Caco-2 system. The inhibitory potencies of known P-gp inhibitors viz verapamil, ketoconazole, tacrolimus and cyclosporine A, determined over a wide concentration range, showed low apparent IC(50) values. Overall, quinidine was found to be a good probe substrate for routine use to assess the in vitro inhibitory potency of NCEs on Pgp-mediated transport.

  19. [Present situation and new challenges for medical treatment of Kawasaki disease].

    PubMed

    Ikeda, Kazuyuki; Hamaoka, Kenji

    2014-09-01

    To predict resistance to intravenous immunoglobulin (MIG) treatment in patients with severe Kawasaki disease (KD), Gunma, Kurume, and Osaka risk scores were established. Using these scores, prospective randomised trials were performed, and addition of predni- solone or intravenous methylprednisolone -pulse to the standard regimen of IVIG improves coronary artery outcomes in severe KD. Adding the evidences of new and existing therapies, such as infliximab, cyclosporin A, ulinastatin, and plasma exchange, the clinical guideline for medical treatment of the acute phase of KD was revised in 2012. Additional large cohort studies are needed to clarify the applicability of intensive initial treatment. Challenges for the future are: 1) clarification of etiology and establishment of specific therapy, 2) establishment of personalized therapy using genetic markers related to the severity of KD.

  20. Synthesis of a new cyclosporine-based stationary phase and separation behaviors toward aromatic positional isomers by high-performance liquid chromatography.

    PubMed

    Liu, Wen-na; Fan, Jun; Lin, Chun; Zheng, Sheng-run; Zhang, Wei-guang

    2015-04-01

    A new cyclosporine-bonded stationary phase has been synthesized through the Staudinger reaction between azido cyclosporine A (CsA) and aminopropyl silica gel and applied in separations of six disubstituted aromatic analytes by high-performance liquid chromatography. For dimethyl phthalate, nitroaniline and chloronitrobenzene, their positional isomers were well-separated on this CsA stationary phase, in which the separation factor for m-/o-dimethyl phthalates was the biggest. For nitrotoluene, dichlorobenzene and benzenediol, the m-/o-isomers were co-eluted. Then, effects of chromatographic conditions (such as types and content of alcoholic modifiers) on separation of nitroaniline positional isomers have been investigated. Retention behaviors of nitroaniline isomers on the column exhibited the strengthening trend along with increasing carbon number of alcohols, from ethanol to 1-propanol, and to 1-butanol. A similar phenomenon was observed by lowering the content of alcohol.

  1. Role of romiplostim in splenectomized and nonsplenectomized patients with immune thrombocytopenia.

    PubMed

    Perdomo, Jose

    2016-01-01

    Romiplostim is a thrombopoietin receptor agonist (TPO-RA) used for the treatment of adult primary immune thrombocytopenia (ITP). ITP is an autoimmune condition characterized by low platelet counts due to increased destruction and reduced platelet production. First-line interventions include corticosteroids, anti-D, and intravenous immunoglobulins, while second-line therapies comprise splenectomy, rituximab, cyclosporine A, and TPO-RAs. The recognition that compromised platelet production is a critical part of the pathogenesis of ITP prompted the development of therapeutic strategies based on the stimulation of the TPO receptor. TPO-RAs enhance megakaryocyte proliferation, increase platelet production, and lead to a reduction in bleeding episodes in ITP patients. This review will summarize current data on the TPO-RA romiplostim, with a particular focus on its relation to splenectomy. PMID:27529057

  2. [Orthotopic liver transplantation: surgery research center's experience].

    PubMed

    Got'e, S V; Eramishantsev, A K; Tsirul'nikova, O M; Skipenko, O G; Filin, A V; Lokshin, L S; Kozhevnikov, V A; Eremenko, A A; Zhidkov, I L

    1997-01-01

    From 1990 to 1996, 178 patients with severe diffuse and advanced focal liver diseases with grave prognosis were observed for indications to orthotopic liver transplantation (OLT). According to the revealed indications and contraindications, 95 patients were enrolled for the procedure. Eighteen patients underwent 19 OLTs, including 1 urgent retransplantation and 3 living related graftings. Surgical techniques were conventional; with these, intraoperative mortality and biliary complications could be avoided. Long-term survival (6 to 68 months) was achieved in 8 recipients, including in those after liver retransplantation. Double or triple suppressive regimens were used. These included corticosteroids, cyclosporin A, azathioprine, antithymocytic globulin. The life quality of recipients promoted restoration of their working capacities. Two recipients gave birth to healthy babies.

  3. T-cell specific immunosuppression by prodigiosin isolated from Serratia marcescens.

    PubMed

    Han, S B; Kim, H M; Kim, Y H; Lee, C W; Jang, E S; Son, K H; Kim, S U; Kim, Y K

    1998-01-01

    Prodigiosin was isolated from the culture broth of Serratia marcescens B-1231. This compound inhibited the T-cell mediated immune responses such as concanavalin-A induced proliferation, mixed lymphocyte response, local graft vs host reaction and T-dependent antibody response at non-toxic concentrations. However, prodigiosin did not affect B-cell mediated immune functions such as lipopolysaccharide-induced proliferation and-activated polyclonal antibody production at the same concentrations. Prodigiosin did not cause death in vitro to lymphocytes at effective concentrations (< 100 nM) and also did not show toxicity in vivo to lymphoid organs at effective dosages (10 and 30 mg/kg). The pharmacological potencies were comparable to the activities of other T-cell specific immunosuppressants such as cyclosporin A and FK-506. In conclusion, it might be suggested that prodigiosin could be used as an immunosuppressant in clinical and immunological studies.

  4. Nodular malignant melanoma and multiple cutaneous neoplasms under immunosuppression with azathioprine.

    PubMed

    Guenova, Emmanuella; Lichte, Verena; Hoetzenecker, Wolfram; Woelbing, Florian; Moehrle, Matthias; Roecken, Martin; Schaller, Martin

    2009-08-01

    Immunosuppressed patients are at increased risk of skin cancer. A 67-year-old renal transplant recipient developed a nodular malignant melanoma after 30 years of immunosuppression with azathioprine and prednisolone. The patient died of metastatic disease 3 months after the diagnosis was made. The function of the renal graft was not affected at all. Renal transplant recipients are at high risk of developing nonmelanocytic skin tumors when on immunosuppressive therapy with cyclosporine A. Less common is the development of skin cancer during immunosuppression with azathioprine. Latest reports show the increased incidence of malignant melanoma in immunosuppressed patients. Our case illustrates the necessity of close dermatological surveillance of allograft recipients, to assure an early recognition of any malignant skin tumor and to reduce the risk of systemic metastatic disease. PMID:19550360

  5. [Dry eye. An update on epidemiology, diagnosis, therapy and new concepts].

    PubMed

    Schirra, F; Ruprecht, K W

    2004-01-01

    Recent epidemiologic studies quote a prevalence of 5,2% to 63% for dry eye depending on definition. Many risk factors have been identified, among other things the female gender. Dry eye interferes significantly with quality of live. Measurement of the change in temperature and humidity during blinking turned out to be a reliable diagnostic tool. Videokeratoscopy explains well-known visual impairments related to dry eye and, along with lipid film interferometry, provides insight into tear film dynamics. The importance of tear film proteins is underestimated. Among therapeutics for symptomatic relief hyaluronic acid proved to be particularly useful but also the sequence of lid hygiene, warm compresses and lid massage is fundamentally important. The effectiveness of punctum plugs is ascribed considerably to a more efficient impact of essential tear film components. Topical cyclosporine A, INS365, 15(S)-HETE as well as topical androgens represent a whole new class of drugs for causal therapy of dry eye. PMID:14872262

  6. Campylobacter jejuni bacteremia and Guillain-Barré syndrome in a renal transplant recipient.

    PubMed

    Maccario, M; Tarantino, A; Nobile-Orazio, E; Ponticelli, C

    1998-01-01

    In patients who have not undergone transplantation, Guillain-Barré syndrome (GBS) is typically preceded by an acute infection often sustained by Campylobacter jejuni. Thus far, in renal transplant recipients, only eight cases of GBS have been reported. In seven patients GBS was attributed to cytomegalovirus infection and in the eighth patient to cyclosporin A neurotoxicity. We report here the case of a GBS in a renal transplant recipient following C. jejuni bacteremia. The infection quickly disappeared after erythromycin and methronidazole therapy. GBS progressively evolved into a paraparesis within 1 week. After reaching a plateau phase, the clinical status improved and the patient was able to walk unassisted after 3 weeks. At this last check-up, 54 months later, the patient was doing well with a functioning graft and only minimal weakness of the lower limbs.

  7. Nucleotide sequences of fic and fic-1 genes involved in cell filamentation induced by cyclic AMP in Escherichia coli.

    PubMed Central

    Kawamukai, M; Matsuda, H; Fujii, W; Utsumi, R; Komano, T

    1989-01-01

    The nucleotide sequences of fic-1 involved in the cell filamentation induced by cyclic AMP in Escherichia coli and its normal counterpart fic were analyzed. The open reading frame of both fic-1 and fic coded for 200 amino acids. The Gly at position 55 in the Fic protein was changed to Arg in the Fic-1 protein. The promoter activity of fic was confirmed by fusing fic and lacZ. The gene downstream from fic was found to be pabA (p-aminobenzoate). There is an open reading frame (ORF190) coding for 190 amino acids upstream from the fic gene. Computer-assisted analysis showed that Fic has sequence similarity with part of CDC28 of Saccharomyces cerevisiae, CDC2 of Schizosaccharomyces pombe, and FtsA of E. coli. In addition, ORF190 has sequence similarity with the cyclosporin A-binding protein cyclophilin. PMID:2546924

  8. On the Mechanism(s) of Membrane Permeability Transition in Liver Mitochondria of Lamprey, Lampetra fluviatilis L.: Insights from Cadmium

    PubMed Central

    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 Cd2+ and Ca2+ plus Pi as the pore inducers. We found that Ca2+ 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 Cd2+ did not produce the mitochondrial swelling in these media. Low Cd2+ 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 Cd2+ or Ca2+ 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 Cd2+ 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. PMID:24995321

  9. Acetaminophen hepatotoxicity and HIF-1{alpha} induction in acetaminophen toxicity in mice occurs without hypoxia

    SciTech Connect

    Chaudhuri, Shubhra; McCullough, Sandra S.; Hennings, Leah; Letzig, Lynda; Simpson, Pippa M.; Hinson, Jack A.; James, Laura P.

    2011-05-01

    HIF-1{alpha} 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-1{alpha}. 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-1{alpha} 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-1{alpha} in APAP toxicity in the mouse. Time course studies using the hypoxia marker pimonidazole showed no staining for pimonidazole at 1 or 2 h in B6C3F1 mice treated with APAP. Staining for pimonidazole was present in the midzonal to periportal regions at 4, 8, 24 and 48 h 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; 10 mg/kg) reduced HIF-1{alpha} induction in APAP treated mice at 1 and 4 h and did not inhibit the metabolism of APAP (depletion of hepatic non-protein sulfhydryls and hepatic protein adduct levels). The data suggest that HIF-1{alpha} 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.

  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. The Isomerase Active Site of Cyclophilin A Is Critical for Hepatitis C Virus Replication*

    PubMed Central

    Chatterji, Udayan; Bobardt, Michael; Selvarajah, Suganya; Yang, Feng; Tang, Hengli; Sakamoto, Noayo; Vuagniaux, Gregoire; Parkinson, Tanya; Gallay, Philippe

    2009-01-01

    Cyclosporine A and nonimmunosuppressive cyclophilin (Cyp) inhibitors such as Debio 025, NIM811, and SCY-635 block hepatitis C virus (HCV) replication in vitro. This effect was recently confirmed in HCV-infected patients where Debio 025 treatment dramatically decreased HCV viral load, suggesting that Cyps inhibitors represent a novel class of anti-HCV agents. However, it remains unclear how these compounds control HCV replication. Recent studies suggest that Cyps are important for HCV replication. However, a profound disagreement currently exists as to the respective roles of Cyp members in HCV replication. In this study, we analyzed the respective contribution of Cyp members to HCV replication by specifically knocking down their expression by both transient and stable small RNA interference. Only the CypA knockdown drastically decreased HCV replication. The re-expression of an exogenous CypA escape protein, which contains escape mutations at the small RNA interference recognition site, restored HCV replication, demonstrating the specificity for the CypA requirement. We then mutated residues that reside in the hydrophobic pocket of CypA where proline-containing peptide substrates and cyclosporine A bind and that are vital for the enzymatic or the hydrophobic pocket binding activity of CypA. Remarkably, these CypA mutants fail to restore HCV replication, suggesting for the first time that HCV exploits either the isomerase or the chaperone activity of CypA to replicate in hepatocytes and that CypA is the principal mediator of the Cyp inhibitor anti-HCV activity. Moreover, we demonstrated that the HCV NS5B polymerase associates with CypA via its enzymatic pocket. The study of the roles of Cyps in HCV replication should lead to the identification of new targets for the development of alternate anti-HCV therapies. PMID:19380579

  12. Interaction of Poly(ethylenimine)-DNA Polyplexes with Mitochondria; Implications for a Mechanism of Cytotoxicity

    PubMed Central

    Grandinetti, Giovanna; Ingle, Nilesh P.; Reineke, Theresa M.

    2011-01-01

    Poly(ethylenimine) (PEI) and PEI-based systems have been widely studied for use as nucleic acid delivery vehicles. However, many of these vehicles display high cytotoxicity, rendering them unfit for therapeutic use. By exploring the mechanisms that cause cytotoxicity, and through understanding structure-function relationships between polymers and intracellular interactions, nucleic acid delivery vehicles with precise intracellular properties can be tailored for specific function. Previous research has shown that PEI is able to depolarize mitochondria, but the exact mechanism as to how depolarization is induced remains elusive and therefore is the focus of the current study. Potential mechanisms for mitochondrial depolarization include direct mitochondrial membrane permeabilization by PEI or PEI polyplexes, activation of the mitochondrial permeability transition pore, and interference with mitochondrial membrane proton pumps, specifically Complex I of the electron transport chain and F0F1-ATPase. Herein, confocal microscopy and live cell imaging showed that PEI polyplexes do colocalize to some degree with mitochondria early in transfection, and the degree of colocalization increases over time. Cyclosporine a was used to prevent activation of the mitochondrial membrane permeability transition pore, and it was found that early in transfection cyclosporine a was unable to prevent the loss of mitochondrial membrane potential. Further studies done using rotenone and oligomycin to inhibit Complex I of the electron transport chain and F0F1-ATPase, respectively, indicate that both of these mitochondrial proton pumps are functioning during PEI transfection. Overall, we conclude that direct interaction between polyplexes and mitochondria may be the reason why mitochondrial function is impaired during PEI transfection. PMID:21699201

  13. [Analysis of clinical characteristics in 45 cases of Diamond-Blackfan anemia].

    PubMed

    Chen, Yu-Mei; Ruan, Min; Wang, Ya-Qin; Zou, Yao; Zhang, Li; Liu, Tian-Feng; Zhu, Xiao-Fan

    2012-06-01

    In order to explore the diagnosis and therapy of Diamond Blackfan anemia (DBA), the clinical data of 45 cases of DBA admitted in our hospital from February 1994 to July 2011 were analyzed retrospectively. The clinical characteristics, results of laboratory examination, treatment reaction and outcome of disease were investigated. The results indicated that out of 45 children diagnosed as DBA, 14 cases (31.1%) had short stature and physical malformation. All patients had anemia with reticulocytopenia. Thirty-four patients (75.6%) had mean corpuscular volume. Eleven patients (24.4%) had macrocytic anemia. Bone marrow examination showed a marked erythroid hypoplasia in all patients. Out of 29 cases tested for fetal hemoglobin (HbF), 13 cases (44.8%) had high level of HbF. Erythroid colony-forming unit of bone marrow was tested in 25 patients, among them 12 patients (48%) showed normal plasia, 13 (52%) showed hypoplasia. The erythropoietin (EPO) levels of 17 patients were elevated. Karyotypes were examined in 28 patients, and showed all normal. The treatment was based on corticosteroids and Cyclosporine A. Thirty patients had good response to corticosteroid therapy, and 10 of them obtained a sustained corticosteroid-induced remission. Twenty cases discontinued corticosteroid therapy after remission, as a result, 15 cases (75%) relapsed, moreover all the relapsed cases still had good response to corticosteroid. Two relapsed patients suffered from aplastic anemia, one of them died of therapy failure. Six patients were unresponsive to corticosteroid, 1 of which achieved remission with cyclosporine A and the others continued to receive regular transfusions. 3 patients received iron chelation therapy. It is concluded that the clinical characteristics, complete blood count, bone marrow smear, HbF level and EPO level are useful to make a diagnosis of DBA. Most patients have a good response to corticosteroid therapy, but relapse rate is high when drug was discontinued. Patients

  14. 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.

  15. Expression of lung resistance protein in epithelioid sarcoma in vitro and in vivo.

    PubMed

    Kusakabe, H; Iwasaki, H; Sano, K; Kiyokane, K

    2000-06-01

    The incidence of epithelioid sarcoma among patients with malignant soft tissue tumors is small, but the rates of recurrence and metastasis of this type of sarcoma are high. To date, effective chemotherapy for advanced epithelioid sarcoma has not been established and, furthermore, epithelioid sarcoma is known to exhibit multidrug resistance (MDR). The chemosensitivities to anticancer agents of two cell lines established from epithelioid sarcoma were examined in this study. The results showed that the ES-OMC-MN and SFT-8606 cell lines were resistant to vincristine (IC50 1190 nM and 872 nM, respectively) and Adriamycin (IC50 921 nM and 650 nM, respectively), but sensitive to actinomycin D (IC50 < 10 nM). P-glycoprotein (p-Gp) and MDR-associated protein (MRP) were not expressed in these cell lines, but a high expression level of lung resistance protein (LRP) was observed. The original tumor tissues from which the two cell lines were established were also found to be LRP-positive but not to express p-Gp or MRP. Their chemosensitivities to Adriamycin were not significantly altered in the presence of 2.5 microg/ml anti-LRP antibody (LRP-56), but the IC50 of vincristine was much less (IC50 128 nM and 27 nM, respectively) than that for an untreated cell line. It is thus suggested that the vincristine resistance in the two cell lines is LRP-mediated. Since cyclosporin A, known to be a modifier of p-Gp, also induced reversal of vincristine resistance in the ES-OMC-MN and SFT-8606 cell lines (IC50 6.2 nM and 17 nM, respectively), it is suggested that cyclosporin A acts as a modifier of MDR mediated by LRP.

  16. 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.

  17. Reactive nitrogen species in acetaminophen-induced mitochondrial damage and toxicity in mouse hepatocytes.

    PubMed

    Burke, Angela S; MacMillan-Crow, Lee Ann; Hinson, Jack A

    2010-07-19

    Acetaminophen (APAP) toxicity in primary mouse hepatocytes occurs in two phases. The initial phase (0-2 h) occurs with metabolism to N-acetyl-p-benzoquinoneimine which depletes glutathione, and covalently binds to proteins, but little toxicity is observed. Subsequent washing of hepatocytes to remove APAP and reincubating in media alone (2-5 h) results in toxicity. We previously reported that the reincubation phase occurs with mitochondrial permeability transition (MPT) and increased oxidative stress (dichlorodihydrofluorescein fluorescence) (DCFH(2)). Since DCFH(2) may be oxidized by multiple oxidative mechanisms, we investigated the role of reactive nitrogen species (RNS) leading to 3-nitrotyrosine in proteins by ELISA and by immunoblots. Incubation of APAP with hepatocytes for 2 h did not result in toxicity or protein nitration; however, washing hepatocytes and reincubating in media alone (2-5 h) resulted in protein nitration which correlated with toxicity. Inclusion of the MPT inhibitor, cyclosporine A, in the reincubation media eliminated toxicity and protein nitration. The general nitric oxide synthase (NOS) inhibitor L-NMMA and the neuronal NOS (NOS1) inhibitor, 7-nitroindazole, added in the reincubation media decreased toxicity and protein nitration; however, neither the inducible NOS (NOS2) inhibitors L-NIL (N6-(1-iminoethyl)-L-lysine) nor SAIT (S-(2-aminoethyl)isothiourea) decreased protein nitration or toxicity. The RNS scavengers, N-acetylcysteine, and high concentrations of APAP, added in the reincubation phase decreased toxicity and protein nitration. 7-Nitroindazole and cyclosporine A inhibited the APAP-induced loss of mitochondrial membrane potential when added in the reincubation phase. The data indicate a role for RNS in APAP induced toxicity.

  18. 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.

  19. Na+/H+ exchanger-1 inhibitors decrease myocardial superoxide production via direct mitochondrial action.

    PubMed

    Garciarena, Carolina D; Caldiz, Claudia I; Correa, María V; Schinella, Guillermo R; Mosca, Susana M; Chiappe de Cingolani, Gladys E; Cingolani, Horacio E; Ennis, Irene L

    2008-12-01

    The possibility of a direct mitochondrial action of Na(+)/H(+) exchanger-1 (NHE-1) inhibitors decreasing reactive oxygen species (ROS) production was assessed in cat myocardium. Angiotensin II and endothelin-1 induced an NADPH oxidase (NOX)-dependent increase in anion superoxide (O(2)(-)) production detected by chemiluminescence. Three different NHE-1 inhibitors [cariporide, BIIB-723, and EMD-87580] with no ROS scavenger activity prevented this increase. The mitochondria appeared to be the source of the NOX-dependent ROS released by the "ROS-induced ROS release mechanism" that was blunted by the mitochondrial ATP-sensitive potassium channel blockers 5-hydroxydecanoate and glibenclamide, inhibition of complex I of the electron transport chain with rotenone, and inhibition of the permeability transition pore (MPTP) by cyclosporin A. Cariporide also prevented O(2)(-) production induced by the opening of mK(ATP) with diazoxide. Ca(2+)-induced swelling was evaluated in isolated mitochondria as an indicator of MPTP formation. Cariporide decreased mitochondrial swelling to the same extent as cyclosporin A and bongkrekic acid, confirming its direct mitochondrial action. Increased O(2)(-) production, as expected, stimulated ERK1/2 and p90 ribosomal S6 kinase phosphorylation. This was also prevented by cariporide, giving additional support to the existence of a direct mitochondrial action of NHE-1 inhibitors in preventing ROS release. In conclusion, we report a mitochondrial action of NHE-1 inhibitors that should lead us to revisit or reinterpret previous landmark observations about their beneficial effect in several cardiac diseases, such as ischemia-reperfusion injury and cardiac hypertrophy and failure. Further studies are needed to clarify the precise mechanism and site of action of these drugs in blunting MPTP formation and ROS release. PMID:18801963

  20. Analysis of engraftment, graft-versus-host disease, and immune recovery following unrelated donor cord blood transplantation.

    PubMed

    Thomson, B G; Robertson, K A; Gowan, D; Heilman, D; Broxmeyer, H E; Emanuel, D; Kotylo, P; Brahmi, Z; Smith, F O

    2000-10-15

    Unrelated cord blood (UCB) is being used as a source of alternative hematopoietic stem cells for transplantation with increasing frequency. From November 1994 to February 1999, 30 UCB transplant procedures were performed for both malignant and nonmalignant diseases in 27 children, aged 0.4 to 17.1 years. Patients received either HLA-matched (n = 3) or 1- or 2-antigen-mismatched (n = 27) UCB following 1 of 2 standardized preparative and graft-versus-host disease regimens (hyperfractionated total body irradiation, cyclophosphamide, and antithymocyte globulin [ATG] with cyclosporine A and methotrexate; or busulfan, melphalan, and ATG with cyclosporine A and prednisone). The median time to neutrophil and platelet engraftment was 27 days (12-60 days) and 75 days (33-158 days) posttransplantation, respectively. No correlation was noted between neutrophil and platelet engraftment and nucleated cells per kilogram, CD34(+) cells per kilogram infused, or cytomegalovirus status of recipient. The cumulative probability of acute grade 2 or greater graft-versus-host disease (GVHD) was 37.2%, and of grade 3 or greater GVHD was 8.8%. No patients developed chronic GVHD. CD4, CD19, and natural killer cell recovery was achieved at a median of 12, 6, and 2 months, respectively. CD8 recovery was delayed at a median of 9 months. Normal mitogen response was achieved at 6 to 9 months. The probability of survival, disease-free survival, and event-free survival at 1 year was 52.3% (34.1%-70.5%), 54.7% (34.5%-74.9 %) and 49.6% (29.9%-69.4%), respectively. This series of 30 UCB transplants suggests that although CD8 cell recovery is delayed, the pattern of immune reconstitution with UCB is similar to that reported for other stem cell sources. (Blood. 2000;96:2703-2711)

  1. 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

  2. Role of TASK2 in the control of apoptotic volume decrease in proximal kidney cells.

    PubMed

    L'Hoste, Sébastien; Poet, Mallorie; Duranton, Christophe; Belfodil, Radia; é Barriere, Herv; Rubera, Isabelle; Tauc, Michel; Poujeol, Chantal; Barhanin, Jacques; Poujeol, Phillipe

    2007-12-14

    Apoptotic volume decrease (AVD) is prerequisite to apoptotic events that lead to cell death. In a previous study, we demonstrated in kidney proximal cells that the TASK2 channel was involved in the K+ efflux that occurred during regulatory volume decrease. The aim of the present study was to determine the role of the TASK2 channel in the regulation of AVD and apoptosis phenomenon. For this purpose renal cells were immortalized from primary cultures of proximal convoluted tubules (PCT) from wild type and TASK2 knock-out mice (task2-/-). Apoptosis was induced by staurosporine, cyclosporin A, or tumor necrosis factor alpha. Cell volume, K+ conductance, caspase-3, and intracellular reactive oxygen species (ROS) levels were monitored during AVD. In wild type PCT cells the K+ conductance activated during AVD exhibited characteristics of TASK2 currents. In task2-/- PCT cells, AVD and caspase activation were reduced by 59%. Whole cell recordings indicated that large conductance calcium-activated K+ currents inhibited by iberiotoxin (BK channels) partially compensated for the deletion of TASK2 K+ currents in the task2-/- PCT cells. This result explained the residual AVD measured in these cells. In both cell lines, apoptosis was mediated via intracellular ROS increase. Moreover AVD, K+ conductances, and caspase-3 were strongly impaired by ROS scavenger N-acetylcysteine. In conclusion, the main K+ channels involved in staurosporine, cyclosporin A, and tumor necrosis factor-alpha-induced AVD are TASK2 K+ channels in proximal wild type cells and iberiotoxin-sensitive BK channels in proximal task2-/- cells. Both K+ channels could be activated by ROS production.

  3. A systematic evaluation of solubility enhancing excipients to enable the generation of permeability data for poorly soluble compounds in Caco-2 model.

    PubMed

    Shah, Devang; Paruchury, Sundeep; Matta, Muralikrishna; Chowan, Gajendra; Subramanian, Murali; Saxena, Ajay; Soars, Matthew G; Herbst, John; Haskell, Roy; Marathe, Punit; Mandlekar, Sandhya

    2014-01-01

    The study presented here identified and utilized a panel of solubility enhancing excipients to enable the generation of flux data in the Human colon carcinoma (Caco-2) system for compounds with poor solubility. Solubility enhancing excipients Dimethyl acetamide (DMA) 1 % v/v, polyethylene glycol (PEG) 400 1% v/v, povidone 1% w/v, poloxamer 188 2.5% w/v and bovine serum albumin (BSA) 4% w/v did not compromise Caco-2 monolayer integrity as assessed by trans-epithelial resistance measurement (TEER) and Lucifer yellow (LY) permeation. Further, these excipients did not affect P-glycoprotein (P-gp) mediated bidirectional transport of digoxin, permeabilities of high (propranolol) or low permeability (atenolol) compounds, and were found to be inert to Breast cancer resistant protein (BCRP) mediated transport of cladribine. This approach was validated further using poorly soluble tool compounds, atazanavir (poloxamer 188 2.5% w/v) and cyclosporine A (BSA 4% w/v) and also applied to new chemical entity (NCE) BMS-A in BSA 4% w/v, for which Caco-2 data could not be generated using the traditional methodology due to poor solubility (<1 µM) in conventional Hanks balanced salt solution (HBSS). Poloxamer 188 2.5% w/v increased solubility of atazanavir by >8 fold whereas BSA 4% w/v increased the solubility of cyclosporine A and BMS-A by >2-4 fold thereby enabling permeability as well as efflux liability estimation in the Caco-2 model with reasonable recovery values. To conclude, addition of excipients such as poloxamer 188 2.5% w/v and BSA 4% w/v to HBSS leads to a significant improvement in the solubility of the poorly soluble compounds resulting in enhanced recoveries without modulating transporter-mediated efflux, expanding the applicability of Caco-2 assays to poorly soluble compounds.

  4. Expression of a constitutively active calcineurin encoded by an intron-retaining mRNA in follicular keratinocytes.

    PubMed

    Fujimura, Atsushi; Michiue, Hiroyuki; Nishiki, Tei-ichi; Ohmori, Iori; Wei, Fan-Yan; Matsui, Hideki; Tomizawa, Kazuhito

    2011-03-14

    Hair growth is a highly regulated cyclical process. Immunosuppressive immunophilin ligands such as cyclosporin A (CsA) and FK506 are known as potent hair growth modulatory agents in rodents and humans that induce active hair growth and inhibit hair follicle regression. The immunosuppressive effectiveness of these drugs has been generally attributed to inhibition of T cell activation through well-characterized pathways. Specifically, CsA and FK506 bind to intracellular proteins, principally cyclophilin A and FKBP12, respectively, and thereby inhibit the phosphatase calcineurin (Cn). The calcineurin (Cn)/NFAT pathway has an important, but poorly understood, role in the regulation of hair follicle development. Here we show that a novel-splicing variant of calcineurin Aß CnAß-FK, which is encoded by an intron-retaining mRNA and is deficient in the autoinhibitory domain, is predominantly expressed in mature follicular keratinocytes but not in the proliferating keratinocytes of rodents. CnAß-FK was weakly sensitive to Ca(2+) and dephosphorylated NFATc2 under low Ca(2+) levels in keratinocytes. Inhibition of Cn/NFAT induced hair growth in nude mice. Cyclin G2 was identified as a novel target of the Cn/NFATc2 pathway and its expression in follicular keratinocytes was reduced by inhibition of Cn/NFAT. Overexpression of cyclin G2 arrested the cell cycle in follicular keratinocytes in vitro and the Cn inhibitor, cyclosporin A, inhibited nuclear localization of NFATc2, resulting in decreased cyclin G2 expression in follicular keratinocytes of rats in vivo. We therefore suggest that the calcineurin/NFAT pathway has a unique regulatory role in hair follicle development.

  5. Expression of a Constitutively Active Calcineurin Encoded by an Intron-Retaining mRNA in Follicular Keratinocytes

    PubMed Central

    Fujimura, Atsushi; Michiue, Hiroyuki; Nishiki, Tei-ichi; Ohmori, Iori; Wei, Fan-Yan; Matsui, Hideki; Tomizawa, Kazuhito

    2011-01-01

    Hair growth is a highly regulated cyclical process. Immunosuppressive immunophilin ligands such as cyclosporin A (CsA) and FK506 are known as potent hair growth modulatory agents in rodents and humans that induce active hair growth and inhibit hair follicle regression. The immunosuppressive effectiveness of these drugs has been generally attributed to inhibition of T cell activation through well-characterized pathways. Specifically, CsA and FK506 bind to intracellular proteins, principally cyclophilin A and FKBP12, respectively, and thereby inhibit the phosphatase calcineurin (Cn). The calcineurin (Cn)/NFAT pathway has an important, but poorly understood, role in the regulation of hair follicle development. Here we show that a novel-splicing variant of calcineurin Aß CnAß-FK, which is encoded by an intron-retaining mRNA and is deficient in the autoinhibitory domain, is predominantly expressed in mature follicular keratinocytes but not in the proliferating keratinocytes of rodents. CnAß-FK was weakly sensitive to Ca2+ and dephosphorylated NFATc2 under low Ca2+ levels in keratinocytes. Inhibition of Cn/NFAT induced hair growth in nude mice. Cyclin G2 was identified as a novel target of the Cn/NFATc2 pathway and its expression in follicular keratinocytes was reduced by inhibition of Cn/NFAT. Overexpression of cyclin G2 arrested the cell cycle in follicular keratinocytes in vitro and the Cn inhibitor, cyclosporin A, inhibited nuclear localization of NFATc2, resulting in decreased cyclin G2 expression in follicular keratinocytes of rats in vivo. We therefore suggest that the calcineurin/NFAT pathway has a unique regulatory role in hair follicle development. PMID:21423799

  6. 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.

  7. Hypertension and ace gene insertion/deletion polymorphism in pediatric renal transplant patients.

    PubMed

    Serdaroglu, Erkin; Mir, Sevgi; Berdeli, Afig

    2005-10-01

    The objective of the present study was to define the risk factors for hypertension and to analyze the influence of insertion/deletion (I/D) polymorphism of the angiotensin-converting enzyme (ACE) on hypertension in pediatric renal transplant recipients. Twenty-six pediatric renal transplant recipients with stable renal function and treated with the same immunosuppression protocol were included in the study. Their mean age was 12.5 +/- 3.3 yr and mean time after transplantation was 38.5 +/- 39.8 month. Twenty-four hour ambulatory blood pressure monitoring (ABPM) was performed by SpaceLabs (90207) device. The I/D polymorphism of the ACE was determined by PCR and ACE serum level was analyzed by colorimetric method. Hypertension was present in 15 patients (57.7%) by causal blood pressure measurements and 19 patients (73.1%) by ABPM. Twenty-two patients (84.6%) were found to be non-dipper and eight of them had reverse dipping. Only time after transplantation (38 +/- 31 vs. 79 +/-49 month, p = 0.016) and cyclosporin A trough plasma levels (206 +/-78 vs. 119 +/- 83 ng/mL, p = 0.020) influenced the presence of hypertension by multiple logistic regression analysis. The distribution of genotypes were II = 2 (7.7%), ID = 8 (30.8%), DD = 16 (61.5%). There was no effect of ACE gene I/D polymorphism or serum ACE levels on hypertension prevalence and circadian variability of blood pressures. Hypertension was related to the time after transplantation and cyclosporin A levels. The ACE gene I/D polymorphism and serum ACE levels did not influence the blood pressure values or circadian variability of blood pressure among pediatric renal transplant patients. PMID:16176418

  8. 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.

  9. Decreased calcineurin immunoreactivity in the postmortem brain of a patient with schizophrenia who had been prescribed the calcineurin inhibitor, tacrolimus, for leukemia

    PubMed Central

    Wada, Akira; Kunii, Yasuto; Matsumoto, Jyunya; Hino, Mizuki; Nagaoka, Atsuko; Niwa, Shin-ichi; Yabe, Hirooki

    2016-01-01

    Background The calcineurin (CaN) inhibitor, tacrolimus, is widely used in patients undergoing allogeneic organ transplantation and in those with certain allergic diseases. Recently, several reports have suggested that CaN is also associated with schizophrenia. However, little data are currently available on the direct effect of tacrolimus on the human brain. Case A 23-year-old Japanese female experienced severe delusion of persecution, delusional mood, suspiciousness, aggression, and excitement. She visited our hospital and was diagnosed with schizophrenia. When she was 27 years old, she had severe general fatigue, persistent fever, systemic joint pain, gingival bleeding, and breathlessness and was diagnosed with acute myelomonocytic leukemia. Later she underwent bone marrow transplantation (BMT), she was administered methotrexate and cyclosporin A to prevent graft versus host disease (GVHD). Three weeks after BMT, she showed initial symptoms of GVHD and was prescribed tacrolimus instead of cyclosporin A. Seven months after BMT at the age of 31 years, she died of progression of GVHD. Pathological anatomy was examined after her death, including immunohistochemical analysis of her brain using anti-CaN antibodies. For comparison, we used our previous data from both a schizophrenia group and a healthy control group. No significant differences were observed in the percentage of CaN-immunoreactive neurons among the schizophrenia group, healthy control group, and the tacrolimus case (all P>0.5, analysis of covariance). Compared with the healthy control group and schizophrenia group, the percentages of CaN-immunoreactive neurons in layers III–VI of the BA46 and the putamen tended to be lower in the tacrolimus case. Conclusion Tacrolimus may decrease CaN immunoreactivity in some regions of the human brain. Thus, tacrolimus may introduce side effects such as cognitive dysfunction and extrapyramidal symptoms. In addition, we also found that the effect of tacrolimus on Ca

  10. 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.

  11. Mitochondrial aquaporin-8 knockdown in human hepatoma HepG2 cells causes ROS-induced mitochondrial depolarization and loss of viability.

    PubMed

    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(2)O(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(2)O(2) release, which may lead to organelle dysfunction and cell death. We confirmed AQP8 expression in HepG2 inner mitochondrial membranes and found that 72h 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(2)O(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(2)O(2) release and that its defective expression causes ROS-induced mitochondrial depolarization via the mitochondrial permeability transition mechanism, and cell death. PMID:22910329

  12. Presence of Cyclophilin A in Synovial Fluids of Patients with Rheumatoid Arthritis

    PubMed Central

    Billich, Andreas; Winkler, Gottfried; Aschauer, Heinrich; Rot, Antal; Peichl, Peter

    1997-01-01

    Cyclophilins have been suggested to act as leukocyte chemotactic factors produced in the course of inflammation. Therefore we looked for the presence of cyclophilins in the synovial fluids (SF) from patients with rheumatoid arthritis (RA). Peptidyl prolyl cis–trans isomerase activity (PPIase) was measured in SF from knee punctures of 26 patients with RA and five patients with knee osteoarthritis (OA). PPIase was detected in SF from RA patients, but not in samples from OA patients. Enzyme activity was sensitive to inhibition by cyclosporin A (IC50 = 28–50 nM). Estimated concentrations of the SF-derived cyclophilin based on the enzyme activity were in the range of 11 to 705 nM. The presence of cyclophilin in the SF showed disease correlation; its concentration correlated with the number of cells in the SF (r   = 0.91, P <0.0001) and with the percentage of neutrophils in the cellular infiltrate and was higher in more acute cases of joint swelling. In immunoblots of partially purified preparations of SF from RA patients, an ∼18-kD protein band reacted with polyclonal antibodies that recognize cyclophilin A and B, but not with antibodies specific for cyclophilin B. Sequencing of this protein revealed identity of the NH2-terminal amino acids with those of human cyclophilin A. The finding is unexpected since cyclophilin B rather than A is generally regarded as the secreted isoform, the presence of cyclophilin A being confined to the cytoplasm. Our data support the hypothesis that cyclophilins may contribute to the pathogenesis of inflammatory diseases, possibly by acting as cytokines. This may offer a possible explanation of the effectiveness of cyclosporin A in RA, in addition to the known immunosuppressive effects of the drug. PMID:9120404

  13. 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

  14. Evaluation of potential chemoprotectants against microcystin-LR hepatotoxicity in mice.

    PubMed

    Hermansky, S J; Stohs, S J; Eldeen, Z M; Roche, V F; Mereish, K A

    1991-02-01

    Microcystin-LR (MCLR) is a potent cyclic heptapeptide hepatotoxin produced by the blue-green algae, Microcystis aeruginosa. Toxic blooms of this cyanobacteria have been reported throughout the temperate world. In spite of the potential economic loss and health hazard posed by this toxin, few studies on the development of an antidote have been conducted. Thus, a number of biologically active compounds were tested in mice for effectiveness in preventing the toxicity of a lethal dose of MCLR (100 micrograms kg-1). Efficacy was evaluated based upon the percentage of surviving mice, time to death and serum lactate dehydrogenase activity 45 min after treatment with the toxin. The biologically active compounds were separated into groups based upon proposed mechanisms of action. Enzyme induction by phenobarbital but not by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) resulted in partial protection against toxicity. Calcium channel blockers, free-radical scavengers and water-soluble antioxidants produced little protection against toxicity. The membrane-active antioxidants vitamin E and silymarin, as well as glutathione and the monoethyl ester of glutathione, produced significant protection from lethality. Rifampin and cyclosporin-A, both immunosuppressive and membrane-active agents, which also block the bile acid uptake system of hepatocytes, produced complete protection from the toxicity of MCLR. Thus, lipophilic antioxidants provide partial protection against MCLR toxicity while cyclosporin-A and rifampin are highly effective and potentially useful antidotes. The toxicity of MCLR may depend upon stimulation of the immune system and may be mediated by membrane alterations. PMID:1902496

  15. 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

  16. 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

  17. 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

  18. Mitochondrial Dysfunction Induced by Different Organochalchogens Is Mediated by Thiol Oxidation and Is Not Dependent of the Classical Mitochondrial Permeability Transition Pore Opening

    PubMed Central

    Puntel, Robson L.; Roos, Daniel H.; Folmer, Vanderlei; Nogueira, Cristina W.; Galina, Antonio; Aschner, Michael; Rocha, João B. T.

    2010-01-01

    Ebselen (Ebs) and diphenyl diselenide [(PhSe)2] readily oxidize thiol groups. Here we studied mitochondrial swelling changes in mitochondrial potential (Δψm), NAD(P)H oxidation, reactive oxygen species production, protein aggregate formation, and oxygen consumption as ending points of their in vitro toxicity. Specifically, we tested the hypothesis that organochalchogens toxicity could be associated with mitochondrial dysfunction via oxidation of vicinal thiol groups that are known to be involved in the regulation of mitochondrial permeability (Petronilli et al. J. Biol. Chem., 269; 16638; 1994). Furthermore, we investigated the possible mechanism(s) by which these organochalchogens could disrupt liver mitochondrial function. Ebs and (PhSe)2 caused mitochondrial depolarization and swelling in a concentration-dependent manner. Furthermore, both organochalchogens caused rapid oxidation of the mitochondrial pyridine nucleotides (NAD(P)H) pool, likely reflecting the consequence and not the cause of increased mitochondrial permeability (Costantini, P., Chernyak, B. V., Petronilli, V., and Bernardi, P. (1996). Modulation of the mitochondrial permeability transition pore (PTP) by pyridine nucleotides and dithiol oxidation at two separate sites. J. Biol. Chem. 271, 6746–6751). The organochalchogens-induced mitochondrial dysfunction was prevented by the reducing agent dithiothreitol (DTT). Ebs- and (PhSe)2-induced mitochondrial depolarization and swelling were unchanged by ruthenium red (4μM), butylated hydroxytoluene (2.5μM), or cyclosporine A (1μM). N-ethylmaleimide enhanced the organochalchogens-induced mitochondrial depolarization, without affecting the magnitude of the swelling response. In contrast, iodoacetic acid did not modify the effects of Ebs or (PhSe)2 on the mitochondria. Additionally, Ebs and (PhSe)2 decreased the basal 2' 7' dichlorofluorescin diacetate (H2-DCFDA) oxidation and oxygen consumption rate in state 3 and increased it during the state 4 of

  19. Interleukin 1 stimulates proliferation of a nontransformed T lymphocyte line in the absence of a co-mitogen.

    PubMed

    Lacey, D L; Chappel, J C; Teitelbaum, S L

    1987-10-15

    Although interleukin (IL) 2-responsive T cell lines provide an opportunity to study the cellular effects of this lymphokine on homogeneous T lymphocyte populations, T cell clones which proliferate in response to IL-1 alone have not been available. We have isolated from cultures of the nontransformed murine T helper cell line, D10 . G4 . 1, a variant (MD10 cells) which proliferates (no lectin or antigen needed) in response to IL-1 alone. The MD10 cells are markedly sensitive to either murine or human recombinant IL-alpha (HrIL-1 alpha) with half-maximal responses observed at monokine concentrations as low as 0.4 X 10(-12) M or 0.8 U/ml, respectively. MD10 cells show the maximal IL-1 effect at 72 hr where the response exceeds the base line by 100-fold (approximately 3,000----300,000 cpm of [3H]thymidine). Whereas both HrIL-2 and purified murine B cell-stimulatory factor 1 (MpBSF-1) induce MD10 proliferation, the maximal response to either is much lower (HrIL-2: 50X baseline; MpBSF-1: less than 20X base line) than to IL-1. Conditioned media from control, concanavalin A-, or IL-1-treated MD10 cells fail to stimulate CTLL or HT-2 cell proliferation alone or inhibit CTLL mitogenesis in the presence of added HrIL-2. Furthermore, monoclonal antibodies to BSF-1 fail to inhibit IL-1-stimulated MD10 replication, and neither HT-2 nor CTLL cells proliferate despite direct cell-to-cell contact with IL-1-treated MD10 cells. When combined, IL-1 (10(-13), 10(-12) M) and IL-2 (10(-13) to 10(-10) M) act synergistically in their MD10 cell growth-promoting effects. MD10 proliferation induced by either IL-1 or IL-2 is relatively resistant to cyclosporine A, with the ID50 of cyclosporine for both IL-1- and IL-2-exposed MD10 cells (ID50 5000 ng/ml) exceeding that for concanavalin A-activated splenocytes (ID50 20 ng/ml) by 2 to 3 orders of magnitude. Finally, MD10 cells bear the L3T4 antigen, IL-2 receptors, and the same clonotypic antigen receptor as the parent clone as recognized by

  20. 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

  1. 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

  2. Inward transport of [3H]-1-methyl-4-phenylpyridinium in rat isolated hepatocytes: putative involvement of a P-glycoprotein transporter.

    PubMed Central

    Martel, F.; Martins, M. J.; Hipólito-Reis, C.; Azevedo, I.

    1996-01-01

    1. The liver has an important role in the detoxification of organic cations from the circulation. [3H]-1-methyl-4-phenylpyridinium ([3H]-MPP+), a low molecular weight organic cation, is efficiently taken up and accumulated by rat hepatocytes through mechanisms partially unknown. 2. The aim of the present work was to characterize further the uptake of MPP+ by rat isolated hepatocytes. The putative interactions of a wide range of drugs, including inhibitors/substrates of P-glycoprotein, were studied. 3. The uptake of MPP+ was investigated in rat freshly isolated hepatocytes (incubated in Krebs-Henseleit medium with 200 nM [3H]-MPP+ for 5 min) and in the rat liver in situ (perfused with Krebs-Henseleit/BSA medium with 200 nM [3H]-MPP+ for 30 min). [3H]-MPP+ accumulation in the cells and in tissue was determined by liquid scintillation counting. 4. Verapamil (100 microM), quinidine (100 microM), amiloride (1 mM), (+)-tubocurarine (100 microM), vecuronium (45 microM), bilirubin (200 microM), progesterone (200 microM), daunomycin (100 microM), vinblastine (100 microM), cyclosporin A (100 microM) and cimetidine (100 microM) had a significant inhibitory effect on the accumulation of [3H]-MPP+ in isolated hepatocytes. Tetraethylammonium (100 microM) had no effect. 5. In the rat perfused liver, both cyclosporin A (100 microM) and verapamil (100 microM) had much less marked inhibitory effects as compared to their effects on isolated hepatocytes (0% against 35% and 45% against 96% of inhibition, respectively). 6. Inhibition of alkaline phosphatase activity by increasing or decreasing the pH of the incubation medium or by the presence of vanadate (1 mM) or homoarginine (500 microM) led to a significant increase in the accumulation of [3H]-MPP+ in isolated hepatocytes. 7. It was concluded that, in addition to the type I organic cation hepatic transporter, [3H]-MPP+ is taken up by rat hepatocytes through P-glycoprotein, a canalicular transport system that usually excretes

  3. 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

  4. 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

  5. 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

  6. Inhibition of cyclophilin A suppresses H2O2-enhanced replication of HCMV through the p38 MAPK signaling pathway.

    PubMed

    Xiao, Jun; Song, Xin; Deng, Jiang; Lv, Liping; Ma, Ping; Gao, Bo; Zhou, Xipeng; Zhang, Yanyu; Xu, Jinbo

    2016-09-01

    Human cytomegalovirus (HCMV) infection can be accelerated by intracellular and extracellular hydrogen peroxide (H2O2) stimulation, mediated by the activation of the p38 mitogen-activated protein kinase (MAPK) pathway. However, it remains unknown whether host gene expression is involved in H2O2-upregulated HCMV replication. Here, we show that the expression of the host gene, cyclophilin A (CyPA), could be facilitated by treatment with H2O2 in a dose-dependent manner. Experiments with CyPA-specific siRNA, or with cyclosporine A, an inhibitor of CyPA, confirmed that H2O2-mediated upregulation of HCMV replication is specifically mediated by upregulation of CyPA expression. Furthermore, depletion or inhibition of CyPA reduced H2O2-induced p38 activation, consistent with that of H2O2-upregulated HCMV lytic replication. These results show that H2O2 is capable of activating ROS-CyPA-p38 MAPK interactions to enhance HCMV replication. PMID:27642560

  7. Critical Diamond-Blackfan anemia due to ribosomal protein S19 missense mutation.

    PubMed

    Ozono, Shuichi; Mitsuo, Miho; Noguchi, Maiko; Nakagawa, Shin-Ichiro; Ueda, Koichiro; Inada, Hiroko; Ohga, Shouichi; Ito, Etsuro

    2016-09-01

    Diamond-Blackfan anemia (DBA) is a rare congenital disorder characterized by pure erythrocyte aplasia, and approximately 70% of patients carry mutations in the genes encoding ribosomal proteins (RP). Here, we report the case of a male infant with DBA who presented with anemic crisis (hemoglobin [Hb] concentration 1.5 g/dL) at 58 days after birth. On admission, the infant was pale and had tachypnea, but recovered with intensive care, including red blood cell transfusions, and prednisolone. Based on the clinical diagnosis of DBA, the father of the infant had cyclosporine-A-dependent anemia. On analysis of RP genes when the infant was 6 months old, both the infant and the father, but not the mother, were found to harbor a mutation of RPS19 (c.167G > C, p. R56P). Therefore, genetic background search and early neonatal health check-ups are recommended for families with a history of inherited bone marrow failure syndromes. PMID:27601194

  8. Phlyctenular keratoconjunctivitis – an atypically severe case treated with systemic biologic immunosuppressive therapy

    PubMed Central

    Valério Sequeira Valadares, Joana; Bastos-Carvalho, Ana; Pedroso Franco, José Manuel; Mourão, Ana Filipa; Monteiro-Grillo, Manuel

    2014-01-01

    Purpose: To report an atypically severe and refractory phlyctenular keratoconjunctivitis case treated successfully with systemic biologic immunosuppressive therapy. Methods: A 10-year-old female was followed in the ophthalmology clinic for three years for a severe form of bilateral PKC. The patient was treated for blepharitis and intestinal parasitosis, and underwent topical corticosteroid therapy, followed by subconjunctival injections and systemic corticosteroids with no clinical improvement. An association of topical cyclosporine A and oral methotrexate had no clinical response either. Phlyctenae of the cornea remained evident with neovascularization, progressive peripheral corneal thinning and occasional anterior chamber reaction. Results: The patient was treated with a combination of infliximab and methotrexate and corticosteroid therapy was tapered, with a fast and sustained resolution of the symptoms and corneal signs. Eleven months past initiation of the treatment, the patient remains asymptomatic and without any recurrence of the disease. Conclusion: Phlyctenular keratoconjunctivitis may present with a broad spectrum of symptoms and signs, and its severity varies significantly. In cases of severe PKC, which are refractory to conventional therapy, systemic biologic immunosuppressive therapy may be a valuable alternative.

  9. Thermodynamic Analysis of Protein-Ligand Binding Interactions in Complex Biological Mixtures using the Stability of Proteins from Rates of Oxidation (SPROX) Method

    PubMed Central

    Strickland, Erin C.; Geer, M. Ariel; Tran, Duc T.; Adhikari, Jagat; West, Graham M.; DeArmond, Patrick D.; Xu, Ying; Fitzgerald, Michael C.

    2013-01-01

    The detection and quantitation of protein-ligand binding interactions is critical in a number of different areas of biochemical research from fundamental studies of biological processes to drug discovery efforts. Described here is a protocol that can be used to identify the protein targets of biologically relevant ligands (e.g. drugs like tamoxifen or cyclosporin A) in complex protein mixtures such as cell lysates. The protocol utilizes quantitative, bottom-up, shotgun proteomics technologies (iTRAQ) with a covalent labeling technique, termed Stability of Proteins from Rates of Oxidation (SPROX). In SPROX, the thermodynamic properties of proteins and protein-ligand complexes are assessed using the hydrogen peroxide-mediated oxidation of methionine residues as a function of the chemical denaturant (e.g. guanidine Hydrochloride or urea) concentration. The proteome-wide SPROX experiments described here enable the ligand binding properties of hundreds of proteins to be simultaneously assayed in the context of complex biological samples. The proteomic capabilities of the protocol render it amenable to detection of both the on- and off-target effects of ligand binding. PMID:23257983

  10. Management of chronic urticaria in Asia: 2010 AADV consensus guidelines.

    PubMed

    Chow, Steven K W

    2012-04-01

    This guideline is a result of a consensus reached during the 19th Asian-Australasian Regional Conference of Dermatology by the Asian Academy of Dermatology and Venereology Study Group in collaboration with the League of Asian Dermatological Societies in 2010. Urticaria has a profound impact on the quality of life in Asia and the need for effective treatment is required. In line with the EAACI/GA(2)LEN/EDF/WAO guideline for the management of urticaria the recommended first-line treatment is new generation, non-sedating H1-antihistamines. If standard dosing is ineffective, increasing the dosage up to four-fold is recommended. For patients who do not respond to a four-fold increase in dosage of non-sedating H1-antihistamines, it is recommended that therapies such as H2-antihistamine, leukotriene antagonist, and cyclosporine A should be added to the antihistamine treatment. In the choice of second-line treatment, both their costs and risk/benefit profiles are the most important considerations.

  11. Bone morphogenic protein-4 expression in vascular lesions of calciphylaxis.

    PubMed

    Griethe, Wanja; Schmitt, Roland; Jurgensen, Jan Steffen; Bachmann, Sebastian; Eckardt, Kai-Uwe; Schindler, Ralf

    2003-01-01

    Calciphylaxis is characterized by an extensive media-calcification of cutaneous and subcutaneous arterioles and capillaries. Recent studies have provided evidence that vascular calcification is a process with similarities to bone metabolism. Bone morphogenic protein-4 (BMP-4) is physiologically involved in bone development and repair. The presence of BMP-4 in atherosclerosis and in sclerotic heart valves led us to suggest that BMP-4 is also involved in calciphylaxis. A 47-year-old male patient developed end-stage renal failure due to chronic glomerulonephritis. He has had two kidney transplants with an immunosuppressive regimen consisting of cyclosporine A and steroids. He was admitted to our hospital because of an increase in serum creatinine (Cr) and he subsequently developed progressive dermal ulcerations. A skin biopsy led to the diagnosis of calciphylaxis. Immunohistochemistry for BMP-4 of a skin specimen from our patient showed strong cytoplasmic immunoreactivity of intradermal cells with clear spatial association to arterioles and hair follicles. Whereas there are identified inhibitors and promoters of vascular calcification, the presence of BMP-4 has not been demonstrated in calcific uremic arteriolopathy. In contrast to atherosclerosis, BMP-4 in calciphylaxis cannot be found in vascular media, but in intradermal cells at the border of arterioles and hair follicles. Therefore, in calciphylaxis BMP-4 can play the role of a cytokine, a growth factor or a media-calcification promoter. PMID:14733421

  12. Malonate induces cell death via mitochondrial potential collapse and delayed swelling through an ROS-dependent pathway

    PubMed Central

    Fernandez-Gomez, Francisco J; Galindo, Maria F; Gómez-Lázaro, Maria; Yuste, Victor J; Comella, Joan X; Aguirre, Norberto; Jordán, Joaquín

    2005-01-01

    Herein we study the effects of the mitochondrial complex II inhibitor malonate on its primary target, the mitochondrion. Malonate induces mitochondrial potential collapse, mitochondrial swelling, cytochrome c (Cyt c) release and depletes glutathione (GSH) and nicotinamide adenine dinucleotide coenzyme (NAD(P)H) stores in brain-isolated mitochondria. Although, mitochondrial potential collapse was almost immediate after malonate addition, mitochondrial swelling was not evident before 15 min of drug presence. This latter effect was blocked by cyclosporin A (CSA), Ruthenium Red (RR), magnesium, catalase, GSH and vitamin E. Malonate added to SH-SY5Y cell cultures produced a marked loss of cell viability together with the release of Cyt c and depletion of GSH and NAD(P)H concentrations. All these effects were not apparent in SH-SY5Y cells overexpressing Bcl-xL. When GSH concentrations were lowered with buthionine sulphoximine, cytoprotection afforded by Bcl-xL overexpression was not evident anymore. Taken together, all these data suggest that malonate causes a rapid mitochondrial potential collapse and reactive oxygen species production that overwhelms mitochondrial antioxidant capacity and leads to mitochondrial swelling. Further permeability transition pore opening and the subsequent release of proapoptotic factors such as Cyt c could therefore be, at least in part, responsible for malonate-induced toxicity. PMID:15655518

  13. Role of CXCR2/CXCR2 ligands in vascular remodeling during bronchiolitis obliterans syndrome

    PubMed Central

    Belperio, John A.; Keane, Michael P.; Burdick, Marie D.; Gomperts, Brigitte; Xue, Ying Ying; Hong, Kurt; Mestas, Javier; Ardehali, Abbas; Mehrad, Borna; Saggar, Rajan; Lynch, Joseph P.; Ross, David J.; Strieter, Robert M.

    2005-01-01

    Angiogenesis and vascular remodeling support fibroproliferative processes; however, no study has addressed the importance of angiogenesis during fibro-obliteration of the allograft airway during bronchiolitis obliterans syndrome (BOS) that occurs after lung transplantation. The ELR+ CXC chemokines both mediate neutrophil recruitment and promote angiogenesis. Their shared endothelial cell receptor is the G-coupled protein receptor CXC chemokine receptor 2 (CXCR2). We found that elevated levels of multiple ELR+ CXC chemokines correlated with the presence of BOS. Proof-of-concept studies using a murine model of BOS not only demonstrated an early neutrophil infiltration but also marked vascular remodeling in the tracheal allografts. In addition, tracheal allograft ELR+ CXC chemokines were persistently expressed even in the absence of significant neutrophil infiltration and were temporally associated with vascular remodeling during fibro-obliteration of the tracheal allograft. Furthermore, in neutralizing studies, treatment with anti-CXCR2 Abs inhibited early neutrophil infiltration and later vascular remodeling, which resulted in the attenuation of murine BOS. A more profound attenuation of fibro-obliteration was seen when CXCR2–/– mice received cyclosporin A. This supports the notion that the CXCR2/CXCR2 ligand biological axis has a bimodal function during the course of BOS: early, it is important for neutrophil recruitment and later, during fibro-obliteration, it is important for vascular remodeling independent of neutrophil recruitment. PMID:15864347

  14. Immunosuppressive drugs affect high-mannose/hybrid N-glycans on human allostimulated leukocytes.

    PubMed

    Pocheć, Ewa; Bocian, Katarzyna; Ząbczyńska, Marta; Korczak-Kowalska, Grażyna; Lityńska, Anna

    2015-01-01

    N-glycosylation plays an important role in the majority of physiological and pathological processes occurring in the immune system. Alteration of the type and abundance of glycans is an element of lymphocyte differentiation; it is also common in the development of immune-mediated inflammatory diseases. The N-glycosylation process is very sensitive to different environmental agents, among them the pharmacological environment of immunosuppressive drugs. Some results show that high-mannose oligosaccharides have the ability to suppress different stages of the immune response. We evaluated the effects of cyclosporin A (CsA) and rapamycin (Rapa) on high-mannose/hybrid-type glycosylation in human leukocytes activated in a two-way mixed leukocyte reaction (MLR). CsA significantly reduced the number of leukocytes covered by high-mannose/hybrid N-glycans, and the synergistic action of CsA and Rapa led to an increase of these structures on the remaining leukocytes. This is the first study indicating that β1 and β3 integrins bearing high-mannose/hybrid structures are affected by Rapa and CsA. Rapa taken separately and together with CsA changed the expression of β1 and β3 integrins and, by regulating the protein amount, increased the oligomannose/hybrid-type N-glycosylation on the leukocyte surface. We suggest that the changes in the glycosylation profile of leukocytes may promote the development of tolerance in transplantation.

  15. The Next Generation Non-competitive Active Polyester Nanosystems for Transferrin Receptor-mediated Peroral Transport Utilizing Gambogic Acid as a Ligand.

    PubMed

    Saini, P; Ganugula, R; Arora, M; Kumar, M N V Ravi

    2016-01-01

    The current methods for targeted drug delivery utilize ligands that must out-compete endogenous ligands in order to bind to the active site facilitating the transport. To address this limitation, we present a non-competitive active transport strategy to overcome intestinal barriers in the form of tunable nanosystems (NS) for transferrin receptor (TfR) utilizing gambogic acid (GA), a xanthanoid, as its ligand. The NS made using GA conjugated poly(lactide-co-glycolide) (PLGA) have shown non-competitive affinity to TfR evaluated in cell/cell-free systems. The fluorescent PLGA-GA NS exhibited significant intestinal transport and altered distribution profile compared to PLGA NS in vivo. The PLGA-GA NS loaded with cyclosporine A (CsA), a model peptide, upon peroral dosing to rodents led to maximum plasma concentration of CsA at 6 h as opposed to 24 h with PLGA-NS with at least 2-fold higher levels in brain at 72 h. The proposed approach offers new prospects for peroral drug delivery and beyond.

  16. Cyclophilin and the regulation of symbiosis in Aiptasia pallida.

    PubMed

    Perez, S; Weis, V

    2008-08-01

    The sea anemone Aiptasia pallida, symbiotic with intracellular dinoflagellates, expresses a peptydyl-prolyl cis-trans isomerase (PPIase) belonging to the conserved family of cytosolic cyclophilins (ApCypA). Protein extracts from A. pallida exhibited PPIase activity. Given the high degree of conservation of ApCypA and its known function in the cellular stress response, we hypothesized that it plays a similar role in the cnidarian-dinoflagellate symbiosis. To explore its role, we inhibited the activity of cyclophilin with cyclosporin A (CsA). CsA effectively inhibited the PPIase activity of protein extracts from symbiotic A. pallida. CsA also induced the dose-dependent release of symbiotic algae from host tissues (bleaching). Laser scanning confocal microscopy using superoxide and nitric oxide-sensitive fluorescent dyes on live specimens of A. pallida revealed that CsA strongly induced the production of these known mediators of bleaching. We tested whether the CsA-sensitive isomerase activity is important for maintaining the activity of the antioxidant enzyme superoxide dismutase (SOD). SOD activity of protein extracts was not affected by pre-incubation with CsA in vitro. PMID:18723638

  17. Regression of Adjuvant-Induced Arthritis in Rats Following Bone Marrow Transplantation

    NASA Astrophysics Data System (ADS)

    van Bekkum, Dirk W.; Bohre, Els P. M.; Houben, Paul F. J.; Knaan-Shanzer, Shoshan

    1989-12-01

    Total body irradiation followed by bone marrow transplantation was found to be an effective treatment for adjuvant arthritis induced in rats. This treatment is most effective when applied shortly after the clinical manifestation of arthritis--i.e., 4-7 weeks after administration of Mycobacterium tuberculosis. Transplantation of bone marrow at a later stage results in a limited recovery, in that the inflammatory reaction regresses but the newly formed excessive bone is not eliminated. Local irradiation of the affected joints had no effect on the disease. It could also be excluded that the recovery of arthritis following marrow transplantation is due to lack of available antigen. Transplantation of syngeneic bone marrow is as effective as that of allogeneic bone marrow from a rat strain that is not susceptible to induction of adjuvant arthritis. The beneficial effect of this treatment cannot be ascribed to the immunosuppressive effect of total body irradiation, since treatment with the highly immunosuppressive drug Cyclosporin A resulted in a regression of the joint swelling but relapse occurred shortly after discontinuation of the treatment.

  18. HIV-1 Capsid Stabilization Assay.

    PubMed

    Fricke, Thomas; Diaz-Griffero, Felipe

    2016-01-01

    The stability of the HIV-1 core in the cytoplasm is crucial for productive HIV-1 infection. Mutations that stabilize or destabilize the core showed defects in HIV-1 reverse transcription and infection. We developed a novel and simple assay to measure stability of in vitro-assembled HIV-1 CA-NC complexes. This assay allowed us to demonstrate that cytosolic extracts strongly stabilize the HIV-1 core (Fricke et al., J Virol 87:10587-10597, 2013). By using our novel assay, one can measure the ability of different drugs to modulate the stability of in vitro-assembled HIV-1 CA-NC complexes, such as PF74, CAP-1, IXN-053, cyclosporine A, Bi2, and the peptide CAI. We also found that purified CPSF6 (1-321) protein stabilizes in vitro-assembled HIV-1 CA-NC complexes (Fricke et al., J Virol 87:10587-10597, 2013). Here we describe in detail the use of this capsid stability assay. We believe that our assay can be a powerful tool to assess HIV-1 capsid stability in vitro.

  19. Correlation between Radiological and Pathological Findings in Patients with Mycoplasma pneumoniae Pneumonia

    PubMed Central

    Tanaka, Hiroshi

    2016-01-01

    Studies focused on the pathological–radiological correlation of human Mycoplasma (M) pneumoniae pneumonia have rarely been reported. Therefore, we extensively reviewed the literature regarding pathological and radiological studies of Mycoplasma pneumonia, and compared findings between open lung biopsy specimen and computed tomography (CT). Major three correlations were summarized. (1) Peribronchial and perivascular cuffing characterized by mononuclear cells infiltration was correlated with bronchovascular bundles thickening on CT, which was the most common finding of this pneumonia. (2) Cellular bronchitis in the small airways accompanied with exudates or granulation tissue in the lumen revealed as centrilobular nodules on CT. (3) Neutrophils and exudates in the alveolar lumen radiologically demonstrated as air-space consolidation or ground-glass opacities. In M. pulmonis-infected mice model, pathologic patterns are strikingly different according to host cell-mediated immunity (CMI) levels; treatment with interleukin-2 lead to marked cellular bronchitis in the small airways and treatment with prednisolone or cyclosporin-A lead to neutrophils and exudates in the alveolar lumen. Patients with centrilobular nodules predominant radiologic pattern have a high level of CMI, measuring by tuberculin skin test. From these findings, up-regulation of host CMI could change radiological pattern to centrilobular nodules predominant, on the other hand down-regulation of host CMI would change radiological pattern to ground-glass opacity and consolidation. It was suggested the pathological features of M. pneumoniae pneumonia may be altered by the level of host CMI. PMID:27242720

  20. Allosteric modulation of the human P-glycoprotein involves conformational changes mimicking catalytic transition intermediates.

    PubMed

    Ghosh, Pratiti; Moitra, Karobi; Maki, Nazli; Dey, Saibal

    2006-06-01

    The drug transport function of human P-glycoprotein (Pgp, ABCB1) can be inhibited by a number of pharmacological agents collectively referred to as modulators or reversing agents. In this study, we demonstrate that certain thioxanthene-based Pgp modulators with an allosteric mode of action induce a distinct conformational change in the cytosolic domain of Pgp, which alters susceptibility to proteolytic digestion. Both cis and trans-isomers of the Pgp modulator flupentixol confer considerable protection of an 80 kDa Pgp fragment against trypsin digestion, that is recognized by a polyclonal antibody specific for the NH(2)-terminal half to Pgp. The protection by flupentixol is abolished in the Pgp F983A mutant that is impaired in modulation by flupentixols, indicating involvement of the allosteric site in generating the conformational change. A similar protection to an 80 kDa fragment is conferred by ATP, its nonhydrolyzable analog ATPgammaS, and by trapping of ADP-vanadate at the catalytic domain, but not by transport substrate vinblastine or by the competitive modulator cyclosporin A, suggesting different outcomes from modulator interaction at the allosteric site and at the substrate site. In summary, we demonstrate that allosteric interaction of flupentixols with Pgp generates conformational changes that mimic catalytic transition intermediates induced by nucleotide binding and hydrolysis, which may play a crucial role in allosteric inhibition of Pgp-mediated drug transport.

  1. Salinomycin overcomes ABC transporter-mediated multidrug and apoptosis resistance in human leukemia stem cell-like KG-1a cells

    SciTech Connect

    Fuchs, Dominik; Daniel, Volker; Sadeghi, Mahmoud; Opelz, Gerhard; Naujokat, Cord

    2010-04-16

    Leukemia stem cells are known to exhibit multidrug resistance by expression of ATP-binding cassette (ABC) transporters which constitute transmembrane proteins capable of exporting a wide variety of chemotherapeutic drugs from the cytosol. We show here that human promyeloblastic leukemia KG-1a cells exposed to the histone deacetylase inhibitor phenylbutyrate resemble many characteristics of leukemia stem cells, including expression of functional ABC transporters such as P-glycoprotein, BCRP and MRP8. Consequently, KG-1a cells display resistance to the induction of apoptosis by various chemotherapeutic drugs. Resistance to apoptosis induction by chemotherapeutic drugs can be reversed by cyclosporine A, which effectively inhibits the activity of P-glycoprotein and BCRP, thus demonstrating ABC transporter-mediated drug resistance in KG-1a cells. However, KG-1a are highly sensitive to apoptosis induction by salinomycin, a polyether ionophore antibiotic that has recently been shown to kill human breast cancer stem cell-like cells and to induce apoptosis in human cancer cells displaying multiple mechanisms of drug and apoptosis resistance. Whereas KG-1a cells can be adapted to proliferate in the presence of apoptosis-inducing concentrations of bortezomib and doxorubicin, salinomycin does not permit long-term adaptation of the cells to apoptosis-inducing concentrations. Thus, salinomycin should be regarded as a novel and effective agent for the elimination of leukemia stem cells and other tumor cells exhibiting ABC transporter-mediated multidrug resistance.

  2. The Candida albicans plasma membrane protein Rch1p, a member of the vertebrate SLC10 carrier family, is a novel regulator of cytosolic Ca2+ homoeostasis.

    PubMed

    Jiang, Linghuo; Alber, Joerg; Wang, Jihong; Du, Wei; Yang, Xuexue; Li, Xichuan; Sanglard, Dominique; Geyer, Joachim

    2012-06-15

    Candida albicans RCH1 (regulator of Ca(2+) homoeostasis 1) encodes a protein of ten TM (transmembrane) domains, homologous with human SLC10A7 (solute carrier family 10 member 7), and Rch1p localizes in the plasma membrane. Deletion of RCH1 confers hypersensitivity to high concentrations of extracellular Ca(2+) and tolerance to azoles and Li(+), which phenocopies the deletion of CaPMC1 (C. albicans PMC1) encoding the vacuolar Ca(2+) pump. Additive to CaPMC1 mutation, lack of RCH1 alone shows an increase in Ca(2+) sensitivity, Ca(2+) uptake and cytosolic Ca(2+) level. The Ca(2+) hypersensitivity is abolished by cyclosporin A and magnesium. In addition, deletion of RCH1 elevates the expression of CaUTR2 (C. albicans UTR2), a downstream target of the Ca(2+)/calcineurin signalling. Mutational and functional analysis indicates that the Rch1p TM8 domain, but not the TM9 and TM10 domains, are required for its protein stability, cellular functions and subcellular localization. Therefore Rch1p is a novel regulator of cytosolic Ca(2+) homoeostasis, which expands the functional spectrum of the vertebrate SLC10 family.

  3. [Topical cyclosporine in ophthalmology: Pharmacology and clinical indications].

    PubMed

    Levy, O; Labbé, A; Borderie, V; Laroche, L; Bouheraoua, N

    2016-03-01

    Cyclosporine A (CsA) is a cyclic undecapeptide, which is an immunosuppressive drug in the calcineurin inhibitor class. CsA was initially used as a systemic immunosuppressant to minimize rejection of solid organ transplants. In ophthalmology, topically applied CsA was first used to inhibit corneal allograft rejection in the 1980s and later in various inflammatory ocular surface disorders (OSD). Currently, topical ophthalmic CsA is available as a licensed commercial emulsion or is prepared by hospital pharmacies with concentration ranging from 0.05 to 2%. Many of its pharmacological effects on the ocular surface are direct consequences of its ability to inhibit T ciclosporine activation and apoptosis. Topical CsA differs from topical steroids in its favourable local and systemic tolerability at the concentrations used. Most clinical studies have evaluated topical CsA in moderate to severe dry eye disease (DED) and demonstrated its efficacy for improvement of signs and symptoms, thus providing the sole indication for market approval and treatment protocols. For the other indications - corneal graft rejection, blepharitis, allergic or viral keratitis, and ocular surface disease due to graft versus host disease or post-operative DED - evidence-based medicine remains unclear due to the lack of major randomized controlled trials. Despite the lack of standardized protocols or market approval for these conditions, numerous studies suggest clinical efficacy.

  4. A Prospective Study of an Alemtuzumab Containing Reduced-intensity Allogeneic Stem Cell Transplantation Program in Patients with Poor-Risk and Advanced Lymphoid Malignancies

    PubMed Central

    Sauter, Craig S.; Chou, Joanne F.; Papadopoulos, Esperanza B.; Perales, Miguel-Angel; Jakubowski, Ann A.; Young, James W.; Scordo, Michael; Giralt, Sergio; Castro-Malaspina, Hugo

    2015-01-01

    Reduced-intensity conditioning (RIC) regimens for allogeneic stem cell transplantation (allo-SCT) have used alemtuzumab to abrogate the risk of graft-versus-host disease (GVHD). Thirty-eight advanced lymphoma patients underwent a prospective phase II study of melphalan, fludarabine and alemtuzumab containing RIC allo-SCT from 20 matched related and 18 unrelated donors with cyclosporin-A as GVHD prophylaxis. The cumulative incidence of grade II-IV acute GVHD at 3 months was 10.5% and three evaluable patients experienced chronic GVHD. Progression-free (PFS) and overall (OS) survival at 5 years is 25% (95% CI: 13-40) and 44% (95% CI: 28-59%) respectively. Previous high-dose therapy and autologous stem cell transplantation (HDT-ASCT) and elevated LDH at the time of allo-SCT resulted in inferior OS. Within this cohort of high-risk lymphoma patients, alemtuzumab containing RIC resulted in a low risk of GVHD and a high incidence of POD, especially in those with poor-risk features defined by elevated LDH pre-allo-SCT and previous HDT-ASCT. PMID:24528216

  5. Characteristics of Cardiac Allograft Vasculopathy Induced by Immunomodulation in the Miniature Swine

    PubMed Central

    Akashima, Tomohiro; Terasaki, Takamitsu; Wada, Yuko; Ito-Amano, Midori; Suzuki, Jun-ichi; Isobe, Mitsuaki

    2014-01-01

    Purpose: We aimed to develop swine cardiac transplantation model for study of cardiac allograft vasculopathy (CAV) and to characterize the mechanisms of its formation. Methods: Heterotropic cardiac transplantation was performed in swine leukocyte antigen mismatched miniature swine, and CAV was induced by immunomodulation by cyclosporine A (CyA). Histology and immunohistochemistry were performed to identify cellular components of CAV. Fluorescence in situ hybridization (FISH) was developed for detection of 1 and Y-chromosome for identification of cell origin in the female donor to the male recipient heart transplantation model. Results: CAV was successfully developed by immunomodulation of CyA. Severity of CAV revealed more prominent in the distal epicardial coronary arteries than proximal coronary arteries. Phenotype of the SMCs proliferated in the intimal thickening of CAV were mostly embryonal/secretory type. Our new chromosome specific probes for FISH method were useful for discrimination of sex of each cell, and proliferated SMCs were revealed to be mainly donor origin. Conclusion: CAV mimicking human heart transplantation can be developed by appropriate immunomodulation in the swine. In swine CAV, proliferated SMCs seen in the intimal thickening were demonstrated to be from the donor origin. PMID:24747545

  6. Calcineurin inhibition in splenocytes induced by pavlovian conditioning.

    PubMed

    Pacheco-López, Gustavo; Riether, Carsten; Doenlen, Raphael; Engler, Harald; Niemi, Maj-Britt; Engler, Andrea; Kavelaars, Annemieke; Heijnen, Cobi J; Schedlowski, Manfred

    2009-04-01

    Pavlovian conditioning is one of the major neurobiological mechanisms of placebo effects, potentially influencing the course of specific diseases and the response to a pharmacological therapy, such as immunosuppression. In our study with behaviorally conditioned rats, a relevant taste (0.2% saccharin) preceded the application of the immunosuppressive drug cyclosporin A (CsA), a specific calcineurin (CaN) inhibitor. Our results demonstrate that through pavlovian conditioning the particular pharmacological properties of CsA can be transferred to a neutral taste, i.e., CaN activity was inhibited in splenocytes from conditioned rats after reexposure to the gustatory stimulus. Concomitant immune consequences were observed on ex vivo mitogenic challenge (anti-CD3). Particularly, Th1-cytokine, but not Th2-cytokine, production and cell proliferation were impeded. Appropriate pharmacological and behavioral controls certify that all these changes in T-lymphocyte reactivity are attributable to mere taste reexposure. Furthermore, the underlying sympathetic-lymphocyte interaction was revealed modeling the conditioned response in vitro. CaN activity in CD4(+) T lymphocytes is reduced by beta-adrenergic stimulation (terbutaline), with these effects antagonized by the beta-adrenoreceptor antagonist nadolol. In summary, CaN was identified as the intracellular target for inducing conditioned immunosuppression by CsA, contributing to our understanding of the intracellular mechanisms behind "learned placebo effects."

  7. Mitochondrial Thioredoxin System as a Modulator of Cyclophilin D Redox State

    NASA Astrophysics Data System (ADS)

    Folda, Alessandra; Citta, Anna; Scalcon, Valeria; Calì, Tito; Zonta, Francesco; Scutari, Guido; Bindoli, Alberto; Rigobello, Maria Pia

    2016-03-01

    The mitochondrial thioredoxin system (NADPH, thioredoxin reductase, thioredoxin) is a major redox regulator. Here we have investigated the redox correlation between this system and the mitochondrial enzyme cyclophilin D. The peptidyl prolyl cis-trans isomerase activity of cyclophilin D was stimulated by the thioredoxin system, while it was decreased by cyclosporin A and the thioredoxin reductase inhibitor auranofin. The redox state of cyclophilin D, thioredoxin 1 and 2 and peroxiredoxin 3 was measured in isolated rat heart mitochondria and in tumor cell lines (CEM-R and HeLa) by redox Western blot analysis upon inhibition of thioredoxin reductase with auranofin, arsenic trioxide, 1-chloro-2,4-dinitrobenzene or after treatment with hydrogen peroxide. A concomitant oxidation of thioredoxin, peroxiredoxin and cyclophilin D was observed, suggesting a redox communication between the thioredoxin system and cyclophilin. This correlation was further confirmed by i) co-immunoprecipitation assay of cyclophilin D with thioredoxin 2 and peroxiredoxin 3, ii) molecular modeling and iii) depleting thioredoxin reductase by siRNA. We conclude that the mitochondrial thioredoxin system controls the redox state of cyclophilin D which, in turn, may act as a regulator of several processes including ROS production and pro-apoptotic factors release.

  8. Critical Diamond-Blackfan anemia due to ribosomal protein S19 missense mutation.

    PubMed

    Ozono, Shuichi; Mitsuo, Miho; Noguchi, Maiko; Nakagawa, Shin-Ichiro; Ueda, Koichiro; Inada, Hiroko; Ohga, Shouichi; Ito, Etsuro

    2016-09-01

    Diamond-Blackfan anemia (DBA) is a rare congenital disorder characterized by pure erythrocyte aplasia, and approximately 70% of patients carry mutations in the genes encoding ribosomal proteins (RP). Here, we report the case of a male infant with DBA who presented with anemic crisis (hemoglobin [Hb] concentration 1.5 g/dL) at 58 days after birth. On admission, the infant was pale and had tachypnea, but recovered with intensive care, including red blood cell transfusions, and prednisolone. Based on the clinical diagnosis of DBA, the father of the infant had cyclosporine-A-dependent anemia. On analysis of RP genes when the infant was 6 months old, both the infant and the father, but not the mother, were found to harbor a mutation of RPS19 (c.167G > C, p. R56P). Therefore, genetic background search and early neonatal health check-ups are recommended for families with a history of inherited bone marrow failure syndromes.

  9. Clinical features, mutations and treatment of 104 patients of Diamond-Blackfan anemia in China: a single-center retrospective study.

    PubMed

    Wan, Yang; Chen, Xiaojuan; An, Wenbin; Ruan, Min; Zhang, Jingliao; Chang, Lixian; Zhang, Ranran; Zhu, Shuai; Zhang, Yingchi; Yang, Wenyu; Guo, Ye; Yuan, Weiping; Zou, Yao; Chen, Yumei; Zhu, Xiaofan

    2016-10-01

    Diamond-Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by a paucity of erythroid progenitors. We summarized the clinical and genetic features of 104 DBA patients in a single-center retrospective study in China. Data of DBA patients who received consultations at our center from 2003 to 2015 were analyzed retrospectively. Genes encoding 10 ribosomal proteins (RPs) and GATA1 were sequenced for mutation detection. Our cohort was composed of 65 males and 39 females. Congenital malformations were observed in 19 patients. Mutations of the RP genes were detected in 58.3 % patients. Twenty different mutations were first reported. Thirty-four patients received prednisone combined with CsA therapy, and improvement was observed in 20 cases. During follow-up for a median 39 months, 33.7 % of the patients achieved remission, 41.3 % of the patients were persistently transfusion independent, 21.7 % of the patients were transfusion dependent, and three patients died. The patient group with detected mutations had a younger age of disease onset, a higher malformation rate, and tended to have a lower remission rate and a higher transfusion-dependence rate. Prednisone in combination with cyclosporine A can be a second-line choice for DBA patients. Differences were detected between DBA patients with and without detectable mutations in the genes studied.

  10. Emerging treatment options for meibomian gland dysfunction

    PubMed Central

    Qiao, Jing; Yan, Xiaoming

    2013-01-01

    Meibomian gland dysfunction (MGD) is one of the most common diseases observed in clinics; it influences a great number of people, and is the leading cause of evaporative dry eye. Given the increased recognition of the importance of MGD, a great amount of attention has been paid to therapies targeting this condition. The traditional treatments of MGD consist of warm compresses and lid hygiene for removing an obstructed meibum, as well as antibiotics and anti-inflammatory agents to improve the quality of the meibum. However, each of these treatments has a different shortcoming and the treatment of MGD remains challenging. Despite the numerous possible treatment options for MGD, it is still difficult to obtain complete relief of signs and symptoms. This review focuses on current emerging treatment options for MGD including intraductal meibomian gland probing, emulsion eye drops containing lipids, the LipiFlow® thermal pulsation system, N-acetyl-cysteine, azithromycin, oral supplementation with omega-3 essential fatty acids, and cyclosporine A. PMID:24043929

  11. Methylglyoxal induces mitochondrial dysfunction and cell death in liver.

    PubMed

    Seo, Kyuhwa; Ki, Sung Hwan; Shin, Sang Mi

    2014-09-01

    Degradation of glucose is aberrantly increased in hyperglycemia, which causes various harmful effects on the liver. Methylglyoxal is produced during glucose degradation and the levels of methylglyoxal are increased in diabetes patients. In this study we investigated whether methylglyoxal induces mitochondrial impairment and apoptosis in HepG2 cells and induces liver toxicity in vivo. Methylglyoxal caused apoptotic cell death in HepG2 cells. Moreover, methylglyoxal significantly promoted the production of reactive oxygen species (ROS) and depleted glutathione (GSH) content. Pretreatment with antioxidants caused a marked decrease in methylglyoxal-induced apoptosis, indicating that oxidant species are involved in the apoptotic process. Methylglyoxal treatment induced mitochondrial permeability transition, which represents mitochondrial impairment. However, pretreatment with cyclosporin A, an inhibitor of the formation of the permeability transition pore, partially inhibited methylglyoxal-induced cell death. Furthermore, acute treatment of mice with methylglyoxal increased the plasma levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), indicating liver toxicity. Collectively, our results showed that methylglyoxal increases cell death and induces liver toxicity, which results from ROS-mediated mitochondrial dysfunction and oxidative stress. PMID:25343013

  12. [Interstitial cystitis/bladder pain syndrome (IC/BPS)].

    PubMed

    Scheiner, David A; Perucchini, Daniele; Fink, Daniel; Betschart, Cornelia

    2015-08-19

    Interstitial cystitis/bladder pain syndrome (BPS) is still an etiologically poorly understood chronic pain syndrome. BPS is a clinical diagnosis. The current treatment modalities are aimed at symptom relief because no cure is possible. Analgesics may be used at any point in treatment but preferably for short-term relief for flares or bladder pain. AUA has issued clinical practice guidelines with a stepwise approach. The first-line therapy begins with self-care and behavior modification. Physical therapy and oral medications such as amitriptyline, PPS, or antihistamines belong to the second-line therapy. Third-line therapy requires cystoscopy and hydrodistension, treatment of Hunner lesions, or intravesical use of e.g. DMSO. Neuromodulation is considered a fourth-line therapy in patients who have failed third-line treatments. Fifth-line therapies consist of intravesical injection of BoNT or oral cyclosporin A. Cystectomy is the sixth-line therapy and the treatment of last resort. PMID:26286495

  13. Neutron scattering in Australia

    SciTech Connect

    Knott, R.B.

    1994-12-31

    Neutron scattering techniques have been part of the Australian scientific research community for the past three decades. The High Flux Australian Reactor (HIFAR) is a multi-use facility of modest performance that provides the only neutron source in the country suitable for neutron scattering. The limitations of HIFAR have been recognized and recently a Government initiated inquiry sought to evaluate the future needs of a neutron source. In essence, the inquiry suggested that a delay of several years would enable a number of key issues to be resolved, and therefore a more appropriate decision made. In the meantime, use of the present source is being optimized, and where necessary research is being undertaken at major overseas neutron facilities either on a formal or informal basis. Australia has, at present, a formal agreement with the Rutherford Appleton Laboratory (UK) for access to the spallation source ISIS. Various aspects of neutron scattering have been implemented on HIFAR, including investigations of the structure of biological relevant molecules. One aspect of these investigations will be presented. Preliminary results from a study of the interaction of the immunosuppressant drug, cyclosporin-A, with reconstituted membranes suggest that the hydrophobic drug interdigitated with lipid chains.

  14. A novel manganese complex selectively induces malignant glioma cell death by targeting mitochondria

    PubMed Central

    Geng, Ji; Li, Jing; Huang, Tao; Zhao, Kaidi; Chen, Qiuyun; Guo, Wenjie; Gao, Jing

    2016-01-01

    Despite advances in treatment, malignant glioma commonly exhibits recurrence, subsequently leading to a poor prognosis. As manganese (Mn) compounds can be transported by the transferrin-transferrin receptor system, the present study synthesized and examined the potential use of Adpa-Mn as a novel antitumor agent. Adpa-Mn time and dose-dependently inhibited U251 and C6 cell proliferation; however, it had little effect on normal astrocytes. Apoptosis was significantly elevated following treatment with Adpa-Mn, as detected by chromatin condensation, Annexin V/propidium iodide staining, cytochrome c release from mitochondria to the cytoplasm, and the activation of caspases-9, -7 and -3 and poly (ADP-ribose) polymerase. In addition, Adpa-Mn enhanced fluorescence intensity of monodansylcadaverine and elevated the expression levels of the autophagy-related protein microtubule-associated protein 1 light chain 3. Pretreatment with the autophagy inhibitors 3-methyladenine and chloroquine enhanced Adpa-Mn-induced cell inhibition, thus indicating that autophagy has an essential role in this process. Furthermore, evidence of mitochondrial dysfunction was detected in the Adpa-Mn-treated group, including disrupted membrane potential, elevated levels of reactive oxygen species (ROS) and depleted adenosine triphosphate. Conversely, treatment with the mitochondrial permeability transition inhibitor cyclosporin A reversed Adpa-Mn-induced ROS production, mitochondrial damage and cell apoptosis, thus suggesting that Adpa-Mn may target the mitochondria. Taken together, these data suggested that Adpa-Mn may be considered for use as a novel anti-glioma therapeutic option. PMID:27432745

  15. Suppression of Coronavirus Replication by Cyclophilin Inhibitors

    PubMed Central

    Tanaka, Yoshikazu; Sato, Yuka; Sasaki, Takashi

    2013-01-01

    Coronaviruses infect a variety of mammalian and avian species and cause serious diseases in humans, cats, mice, and birds in the form of severe acute respiratory syndrome (SARS), feline infectious peritonitis (FIP), mouse hepatitis, and avian infectious bronchitis, respectively. No effective vaccine or treatment has been developed for SARS-coronavirus or FIP virus, both of which cause lethal diseases. It has been reported that a cyclophilin inhibitor, cyclosporin A (CsA), could inhibit the replication of coronaviruses. CsA is a well-known immunosuppressive drug that binds to cellular cyclophilins to inhibit calcineurin, a calcium-calmodulin-activated serine/threonine-specific phosphatase. The inhibition of calcineurin blocks the translocation of nuclear factor of activated T cells from the cytosol into the nucleus, thus preventing the transcription of genes encoding cytokines such as interleukin-2. Cyclophilins are peptidyl-prolyl isomerases with physiological functions that have been described for many years to include chaperone and foldase activities. Also, many viruses require cyclophilins for replication; these include human immunodeficiency virus, vesicular stomatitis virus, and hepatitis C virus. However, the molecular mechanisms leading to the suppression of viral replication differ for different viruses. This review describes the suppressive effects of CsA on coronavirus replication. PMID:23698397

  16. Cyclophilin A as a New Therapeutic Target for Hepatitis C Virus-induced Hepatocellular Carcinoma

    PubMed Central

    2013-01-01

    Hepatocellular carcinoma (HCC) related to hepatitis B virus (HBV) and hepatitis C virus (HCV) infections is thought to account for more than 80% of primary liver cancers. Both HBV and HCV can establish chronic liver inflammatory infections, altering hepatocyte and liver physiology with potential liver disease progression and HCC development. Cyclophilin A (CypA) has been identified as an essential host factor for the HCV replication by physically interacting with the HCV non structural protein NS5A that in turn interacts with RNA-dependent RNA polymerase NS5B. CypA, a cytosolic binding protein of the immunosuppressive drug cyclosporine A, is overexpressed in many cancer types and often associated with malignant transformation. Therefore, CypA can be a good target for molecular cancer therapy. Because of antiviral activity, the CypA inhibitors have been tested for the treatment of chronic hepatitis C. Nonimmunosuppressive Cyp inhibitors such as NIM811, SCY-635, and Alisporivir have attracted more interests for appropriating CypA for antiviral chemotherapeutic target on HCV infection. This review describes CypA inhibitors as a potential HCC treatment tool that is contrived by their obstructing chronic HCV infection and summarizes roles of CypA in cancer development. PMID:24227937

  17. Drug biokinetic and toxicity assessments in rat and human primary hepatocytes and HepaRG cells within the EU-funded Predict-IV project.

    PubMed

    Mueller, Stefan O; Guillouzo, André; Hewitt, Philip G; Richert, Lysiane

    2015-12-25

    The overall aim of Predict-IV (EU-funded collaborative project #202222) was to develop improved testing strategies for drug safety in the late discovery phase. One major focus was the prediction of hepatotoxicity as liver remains one of the major organ leading to failure in drug development, drug withdrawal and has a poor predictivity from animal experiments. In this overview we describe the use and applicability of the three cell models employed, i.e., primary rat hepatocytes, primary human hepatocytes and the human HepaRG cell line, using four model compounds, chlorpromazine, ibuprofen, cyclosporine A and amiodarone. This overview described the data generated on mode of action of liver toxicity after long-term repeat-dosing. Moreover we have quantified parent compound and its distribution in various in vitro compartments, which allowed us to develop biokinetic models where we could derive real exposure concentrations in vitro. In conclusion, the complex data set enables quantitative measurements that proved the concept that we can define human relevant free and toxic exposure levels in vitro. Further compounds have to be analyzed in a broader concentration range to fully exploit these promising results for improved prediction of hepatotoxicity and hazard assessment for humans. PMID:25952325

  18. Neurologic complications in adult living donor liver transplant patients: an underestimated factor?

    PubMed

    Saner, Fuat Hakan; Gensicke, Julia; Olde Damink, Steven W M; Pavlaković, Goran; Treckmann, Juergen; Dammann, Marc; Kaiser, Gernot M; Sotiropoulos, Georgios C; Radtke, Arnold; Koeppen, Susanne; Beckebaum, Susanne; Cicinnati, Vito; Nadalin, Silvio; Malagó, Massimo; Paul, Andreas; Broelsch, Christoph E

    2010-02-01

    Liver transplantation is the only curative treatment in patients with end-stage liver disease. Neurological complications (NC) are increasingly reported to occur in patients after cadaveric liver transplantation. This retrospective cohort study aims to evaluate the incidence and causes of NC in living donor liver transplant (LDLT) patients in our transplant center. Between August 1998 and December 2005, 121 adult LDLT patients were recruited into our study. 17% of patients experienced NC, and it occurred significantly more frequently in patients with alcoholic cirrhosis (42%) and autoimmune hepatitis (43%) as compared with patients with hepatitis B or C (9/10%, P = 0.013). The most common NC was encephalopathy (47.6%) followed by seizures (9.5%). The choice of immunosuppression by calcineurin inhibitor (Tacrolimus or Cyclosporin A) showed no significant difference in the incidence of NC (19 vs. 17%). The occurrence of NC did not influence the clinical outcome, since mortality rate, median ICU stay and length of hospital stay were similar between the two groups. Most patients who survived showed a nearly complete recovery of their NC. NCs occur in approximately 1 in 6 patients after LDLT and seem to be predominantly transient in nature, without major impact on clinical outcome. PMID:19727899

  19. IGF-1 induces skeletal myocyte hypertrophy through calcineurin in association with GATA-2 and NF-ATc1

    NASA Technical Reports Server (NTRS)

    Musaro, A.; McCullagh, K. J.; Naya, F. J.; Olson, E. N.; Rosenthal, N.

    1999-01-01

    Localized synthesis of insulin-like growth factors (IGFs) has been broadly implicated in skeletal muscle growth, hypertrophy and regeneration. Virally delivered IGF-1 genes induce local skeletal muscle hypertrophy and attenuate age-related skeletal muscle atrophy, restoring and improving muscle mass and strength in mice. Here we show that the molecular pathways underlying the hypertrophic action of IGF-1 in skeletal muscle are similar to those responsible for cardiac hypertrophy. Transfected IGF-1 gene expression in postmitotic skeletal myocytes activates calcineurin-mediated calcium signalling by inducing calcineurin transcripts and nuclear localization of calcineurin protein. Expression of activated calcineurin mimics the effects of IGF-1, whereas expression of a dominant-negative calcineurin mutant or addition of cyclosporin, a calcineurin inhibitor, represses myocyte differentiation and hypertrophy. Either IGF-1 or activated calcineurin induces expression of the transcription factor GATA-2, which accumulates in a subset of myocyte nuclei, where it associates with calcineurin and a specific dephosphorylated isoform of the transcription factor NF-ATc1. Thus, IGF-1 induces calcineurin-mediated signalling and activation of GATA-2, a marker of skeletal muscle hypertrophy, which cooperates with selected NF-ATc isoforms to activate gene expression programs.

  20. A prospective study of an alemtuzumab containing reduced-intensity allogeneic stem cell transplant program in patients with poor-risk and advanced lymphoid malignancies.

    PubMed

    Sauter, Craig S; Chou, Joanne F; Papadopoulos, Esperanza B; Perales, Miguel-Angel; Jakubowski, Ann A; Young, James W; Scordo, Michael; Giralt, Sergio; Castro-Malaspina, Hugo

    2014-12-01

    Reduced-intensity conditioning (RIC) regimens for allogeneic stem cell transplant (allo-SCT) have used alemtuzumab to abrogate the risk of graft-versus-host disease (GVHD). Thirty-eight patients with advanced lymphoma underwent a prospective phase II study of melphalan, fludarabine and alemtuzumab containing RIC allo-SCT from 20 matched related and 18 unrelated donors with cyclosporine-A as GVHD prophylaxis. The cumulative incidence of grade II-IV acute GVHD at 3 months was 10.5% and three evaluable patients experienced chronic GVHD. Progression-free (PFS) and overall (OS) survival at 5 years was 25% (95% confidence interval [CI]: 13-40%) and 44% (95% CI: 28-59%), respectively. Previous high-dose therapy and autologous stem cell transplant (HDT-ASCT) and elevated lactate dehydrogenase (LDH) at the time of allo-SCT resulted in inferior OS. Within this cohort of patients with high-risk lymphoma, alemtuzumab containing RIC resulted in a low risk of GVHD and a high incidence of progression of disease, especially in those with poor-risk features defined by elevated LDH pre-allo-SCT and previous HDT-ASCT.

  1. Updosing of Nonsedating Anti-histamines in Recalcitrant Chronic Urticaria

    PubMed Central

    Godse, Kiran; Bhattar, Prachi; Patil, Sharmila; Nadkarni, Nitin; Gautam, Manjyot

    2016-01-01

    Chronic urticaria (CU) is a persistent, debiliating condition that causes severe impairment on the quality of life (QoL) of patient by interrupting work productivity. Current guidelines recommend second-generation (nonsedating) anti-histamines for the treatment for all forms of urticaria. In patients who do not respond adequately to conventional doses of anti-histamines, it is recommended to increase the dose to up to four times to obtain control. But there are only few controlled studies that have assessed the efficacy and safety of nonsedating anti-histamines. Though sedating histamines are frequently used as an add-on therapy in severe cases, they have a negative impact on QoL by compromising sleep and performance. The use of other suggested therapeutic options (omalizumab, cyclosporine A, montelukast and dapsone) is also limited by paucity of data on their efficacy and adverse effect profile. Second-generation anti-histamines which are relatively safer require more proven data to support their judicious use to improve disease in patients with CU. PMID:27293247

  2. Transverse relaxation of scalar-coupled protons.

    PubMed

    Segawa, Takuya F; Baishya, Bikash; Bodenhausen, Geoffrey

    2010-10-25

    In a preliminary communication (B. Baishya, T. F. Segawa, G. Bodenhausen, J. Am. Chem. Soc. 2009, 131, 17538-17539), we recently demonstrated that it is possible to obtain clean echo decays of protons in biomolecules despite the presence of homonuclear scalar couplings. These unmodulated decays allow one to determine apparent transverse relaxation rates R(2) (app) of individual protons. Herein, we report the observation of R(2) (app) for three methyl protons, four amide H(N) protons, and all 11 backbone H(α) protons in cyclosporin A. If the proton resonances overlap, their R(2) (app) rates can be measured by transferring their magnetization to neighboring (13)C nuclei, which are less prone to overlap. The R(2) (app) rates of protons attached to (13)C are faster than those attached to (12)C because of (13)C-(1)H dipolar interactions. The differences of these rates allow the determination of local correlation functions. Backbone H(N) and H(α) protons that have fast decay rates R(2) (app) also feature fast longitudinal relaxation rates R(1) and intense NOESY cross peaks that are typical of crowded environments. Variations of R(2) (app) rates of backbone H(α) protons in similar amino acids reflect differences in local environments.

  3. Therapy of inflammatory bowel diseases in pregnancy and lactation.

    PubMed

    Cassina, Matteo; Fabris, Luca; Okolicsanyi, Lajos; Gervasi, Maria Teresa; Memmo, Alessia; Tiboni, Gian Mario; Di Gianantonio, Elena; Clementi, Maurizio

    2009-11-01

    Inflammatory bowel diseases (IBDs) are a group of disorders characterised by chronic or relapsing inflammation within the gastrointestinal tract of variable severity. A chronic medication is often needed and management of fertile women is a crucial point because of the possible adverse effects associated with the administered drugs and the disease itself. The risk of pregnancy-related complications and the disease behaviour during pregnancy depends mainly on disease activity at time of conception. So, it is very important to plan the pregnancy and reach and maintain a clinical remission of the disease before conception. Drugs usually used in IBD treatment include 5- aminosalicylic acid compounds, corticosteroids, azathioprine and 6-mercaptopurine, cyclosporine A, mesalazine, and antibiotics such as metronidazole and ciprofloxacin. Management of IBD in pregnancy at present is not standardised or supported by strong evidence. In this report, we summarise the available data, mainly derived from retrospective and case-control studies, about IBD management in pregnancy, focusing mostly on the safety of drugs during gestation and peripartum.

  4. Hydration-Induced Phase Separation in Amphiphilic Polymer Matrices and its Influence on Voclosporin Release

    SciTech Connect

    Khan, I. John; Murthy, N. Sanjeeva; Kohn, Joachim

    2015-10-30

    Voclosporin is a highly potent, new cyclosporine -- a derivative that is currently in Phase 3 clinical trials in the USA as a potential treatment for inflammatory diseases of the eye. Voclosporin represents a number of very sparingly soluble drugs that are difficult to administer. It was selected as a model drug that is dispersed within amphiphilic polymer matrices, and investigated the changing morphology of the matrices using neutron and x-ray scattering during voclosporin release and polymer resorption. The hydrophobic segments of the amphiphilic polymer chain are comprised of desaminotyrosyl-tyrosine ethyl ester (DTE) and desaminotyrosyl-tyrosine (DT), and the hydrophilic component is poly(ethylene glycol) (PEG). Water uptake in these matrices resulted in the phase separation of hydrophobic and hydrophilic domains that are a few hundred Angstroms apart. These water-driven morphological changes influenced the release profile of voclosporin and facilitated a burst-free release from the polymer. No such morphological reorganization was observed in poly(lactide-co-glycolide) (PLGA), which exhibits an extended lag period, followed by a burst-like release of voclosporin when the polymer was degraded. An understanding of the effect of polymer composition on the hydration behavior is central to understanding and controlling the phase behavior and resorption characteristics of the matrix for achieving long-term controlled release of hydrophobic drugs such as voclosporin.

  5. Beneficial effect of shallot (Allium ascalonicum L.) extract on cyclosporine nephrotoxicity in rats.

    PubMed

    Wongmekiat, O; Leelarugrayub, N; Thamprasert, K

    2008-05-01

    The clinical use of an immunosuppressive cyclosporine A (CsA) is limited by its serious nephrotoxic effect. Evidences have suggested the role of oxidative stress in its pathogenesis. Shallot (Allium ascalonicum L.) has recently been shown to possess antioxidative and free radical scavenging abilities. The present study was undertaken to investigate the possible beneficial effect of shallot extract on renal injury caused by CsA. Male Wistar rats were treated orally with vehicle, CsA (25 mg/kg), shallot extract (1 g/kg), and CsA plus shallot extract for 21 days. Renal function, histopathology, tissue malondialdehyde (MDA) and glutathione (GSH) levels were evaluated 24 h after the last treatment. CsA-induced nephrotoxicity was evidenced by increased blood urea nitrogen and serum creatinine, but decreased urea and creatinine clearance. The kidney of CsA treated rats exhibited severe vacuolations and tubular necrosis. CsA also induced oxidative stress, as indicated by increased renal MDA and reduced GSH concentrations. Administration of shallot extract along with CsA counteracted the deleterious effects of CsA on renal dysfunction, oxidative stress markers, and morphological changes. These data indicate the protective potential of shallot extract against CsA nephrotoxicity and suggest a significant contribution of its antioxidant property to this beneficial effect.

  6. Increased Susceptibility of Gracilinanus microtarsus Liver Mitochondria to Ca2+-Induced Permeability Transition Is Associated with a More Oxidized State of NAD(P)

    PubMed Central

    Ronchi, Juliana A.; Henning, Barbara; Ravagnani, Felipe G.; Figueira, Tiago R.; Castilho, Roger F.; dos Reis, Sergio F.; Vercesi, Anibal E.

    2015-01-01

    In addition to be the cell's powerhouse, mitochondria also contain a cell death machinery that includes highly regulated processes such as the membrane permeability transition pore (PTP) and reactive oxygen species (ROS) production. In this context, the results presented here provide evidence that liver mitochondria isolated from Gracilinanus microtarsus, a small and short life span (one year) marsupial, when compared to mice, are much more susceptible to PTP opening in association with a poor NADPH dependent antioxidant capacity. Liver mitochondria isolated from the marsupial are well coupled and take up Ca2+ but exhibited a much lower Ca2+ retention capacity than mouse mitochondria. Although the known PTP inhibitors cyclosporin A, ADP, and ATP significantly increased the marsupial mitochondria capacity to retain Ca2+, their effects were much larger in mice than in marsupial mitochondria. Both fluorescence and HPLC analysis of mitochondrial nicotinamide nucleotides showed that both content and state of reduction (mainly of NADPH) were lower in the marsupial mitochondria than in mice mitochondria despite the similarity in the activity of the glutathione peroxidase/reductase system. Overall, these data suggest that PTP opening is an important event in processes of Ca2+ signalling to cell death mediated by mitochondrial redox imbalance in G. microtarsus. PMID:26583063

  7. Pre-exposure to the unconditioned or conditioned stimulus does not affect learned immunosuppression in rats.

    PubMed

    Lueckemann, Laura; Bösche, Katharina; Engler, Harald; Schwitalla, Jan-Claudius; Hadamitzky, Martin; Schedlowski, Manfred

    2016-01-01

    In order to analyze the effects of pre-exposure to either the unconditioned (US) or conditioned stimulus (CS) on learned immunosuppression, we employed an established conditioned taste aversion (CTA) paradigm in rats. In our model, a sweet-tasting drinking solution (saccharin) serves as CS and injection of the immunosuppressive drug cyclosporine A (CsA) is used as US. The conditioned response is reflected by a pronounced CTA and diminished cytokine production by anti-CD3 stimulated splenic T cells. In the present study, experimental animals were exposed either to the US or the CS three times prior to the acquisition phase. On the behavioral level, we found a significantly diminished CTA when animals were pre-exposed to the US or the CS before acquisition. In contrast, US or CS pre-exposure did not affect the behaviorally conditioned suppression of interleukin (IL)-2 production. From the clinical perspective, our data may suggest that conditioning paradigms could be systemically integrated as supportive therapeutic interventions in patients that are already on immunosuppressive therapy or have had previous contact to the gustatory stimulus. Such supportive therapies to pharmacological regimens could not only help to reduce the amount of medication needed and, thus, unwanted toxic side effects, but may also maximize the therapeutic outcome.

  8. Is the control of disease progression within our grasp? Review of the GRISAR study. (Gruppo Reumatologi Italiani Studio Artrite Reumatoide).

    PubMed

    Ferraccioli, G F; Della Casa-Alberighi, O; Marubini, E; Priolo, F; Mathieu, A; Fantini, F; Cutolo, M; Pasero, G

    1996-09-01

    A prospective, open, multicentre, randomized study with a blinded radiological end-point was started in 1991. The aim of the study was to assess whether cyclosporin A (CyA) controls ongoing anatomical damage in active early rheumatoid arthritis (RA) better than conventional disease-modifying anti-rheumatic drugs (DMARDs) as used in everyday clinical practice. A total of 340 consenting patients with early RA (mean duration 1.4 yr) were recruited; 167 were randomized to CyA 3mg/kg per day and 173 to DMARDs. Hand, wrist and foot X-rays were blindly scored by a central committee of three radiologists using the Larsen-Dale method. Any side-effects were carefully recorded. The control of clinical symptoms was similar in both groups. Radiological evaluation of 284 patients (141 on CyA; 143 on DMARDs) after 12 months showed a significant decrease in the mean progression in the eroded joint count (1.3 +/- 3.1 vs 2.4 +/- 3.0, P < 0.001). There was also better maintenance on treatment with CyA than in the group treated with DMARDs (89.2 vs 77.5%, respectively; P = 0.002). CyA seems to offer greater control of ongoing anatomical joint damage in early RA than conventional DMARDs after 12 months.

  9. [Therapy of myositis].

    PubMed

    Keck, A D; Walker, U A

    2013-04-01

    Idiopathic inflammatory myopathy consists of dermatomyositis (DM), polymyositis (PM), inclusion body myositis (IBM) and necrotizing autoimmune myopathy (NAM). At all stages of myositis, physiotherapy is effective in improving muscle strength, endurance and in maintaining joint motion. In DM and PM the therapy is initiated with glucocorticosteroids. Steroid-sparing agents (azathioprine, methotrexate and cyclosporin A) are added to prevent Cushing's syndrome or an unsatisfactory response. Therapy can also be escalated with intravenous immunoglobulins. Tacrolimus and mycophenolate mofetil (MMF) were effective in small case series. Cyclophosphamide is restricted to patients not responding to previous agents. For treatment intensification immunoglobulins can also be combined with MMF. There is not enough evidence to routinely recommend rituximab. The results with TNF-alpha inhibitors and plasmapheresis were negative or inconsistent. In DM skin involvement responds to sun blockers, antimalarials, topical corticosteroids or calcineurin inhibitors. In NAM statins should be discontinued and treatment with prednisone and immunosuppressants initiated. In IBM a therapeutic trial with prednisone, methotrexate or azathioprine may be warranted, especially in cases in which the serum creatine kinase (CK) is elevated or an inflammatory infiltrate is present in the muscle biopsy.

  10. Au-ACRAMTU-PEt3 Alters Redox Balance To Inhibit T Cell Proliferation and Function#

    PubMed Central

    Langston, P. Kent; Yang, Mu; Bierbach, Ulrich; Parsonage, Derek; Poole, Leslie B.; Price, Madeline J.; Grayson, Jason M.

    2015-01-01

    Although T cells play a critical role in protection from viruses, bacteria and tumors, they also cause autoimmune diseases such as systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and multiple sclerosis (MS). Unwanted T cell responses during organ transplant, graft versus host disease (GVHD), and allergies are also major clinical problems. While drugs are available to suppress unwanted immune responses they have limited efficacy with serious side effects. Thus new therapeutics limiting T cell activation, proliferation and function can make an immediate clinical impact. To identify new suppressors of lymphocyte activation, proliferation and function, we examined the immunosuppressive activity of gold(I) analogues of platinum-acridine antitumor agents. We found that the gold complex, Au-ACRAMTU-PEt3 is a potent suppressor of murine and human T cell activation. Preincubation with Au-ACRAMTU-PEt3 suppresses the proliferation of CD4+ and CD8+ T cells at a similar concentration as pharmaceutical grade cyclosporine A. Au-ACRAMTU-PEt3 pretreatment decreases the production of IFNγ, TNFα, IL-2, and IL-17 by human and murine CD4+ and CD8+ T cells. When mice were treated with Au-ACRAMTU-PEt3 during viral infection the expansion of virus-specific CD8+ T cells was decreased 10-fold and viral load was elevated. Taken together these results demonstrate that Au-ACRAMTU-PEt3 has potent immunosuppressive activity that could be used to suppress immune responses during transplantation and autoimmunity. PMID:26209624

  11. Characterization of human lymphoblastoid cell lines as a novel in vitro test system to predict the immunotoxicity of xenobiotics.

    PubMed

    Markovič, Tijana; Gobec, Martina; Gurwitz, David; Mlinarič-Raščan, Irena

    2015-02-17

    Evaluating immunomodulatory effects of xenobiotics is an important component of the toxicity studies. Herein we report on the establishment of a novel invitro test system for the immunotoxicity screening of xenobiotics based on human lymphoblastoid cell lines (LCLs). Four immunotoxic compounds; tributyltin chloride, cyclosporine A, benzo(a)pyrene and verapamil hydrochloride, as well as three immune-inert compounds; urethane, furosemide and mannitol were selected for characterization. The treatment of LCLs with immunosuppressive compounds resulted in reduced viability. The IC50 values determined in human LCLs were in agreement with the data obtained for human peripheral mononuclear cells. Since cytokine production reflects lymphocytes responses to external stimuli, we evaluated the functional responses of LCLs by monitoring their pro-inflammatory and immunoregulatory cytokine production. Our findings prove that LCLs allowed for reliable differentiation between immunomodulatory and immune-inert compounds. Hence, pre-treatment with immunomodulatory compounds led to a decrease in the production of pro-inflammatory TNFα, IL-6 and immunoregulatory IL-2, IL-4, IL-10 and IFNγ cytokines, when compared to untreated ionomycin/PMA stimulated cells. Moreover, testing a panel of ten LCLs derived from unrelated healthy individuals reflects inter-individual variability in response to immunomodulatory xenobiotics. In conclusion, LCLs provide a novel alternative method for the testing of the immunotoxic effects of xenobiotics.

  12. [Quality of life following renal transplantation in childhood].

    PubMed

    Wingen, A M; Feldhoff, C

    1999-01-01

    The goal of renal transplantation is to achieve the best possible quality of life in patients with terminal renal failure. To evaluate some aspects of quality of life in patients with renal grafts during childhood of our center, data on medical, educational and professional rehabilitation were collected retrospectively. Between 1972 and 1997 135 renal transplantations had been performed in 123 patients below the age of 18 years. 12-year graft survival of patients transplanted before 1983 figured at 21% and rose to 62% during the following years, after introduction of cyclosporine A into the immunosuppressive regimen. The proportion of patients in the respective age group attending a secondary school (16%) was lower and of those attending elementary school (71%) or a school for disabled and handicapped children (11%) was higher than usual in the German population. But, 83% of all patients reached a school degree. After school 78% proceeded with a vocational training or university. 89% of patients completing professional training were employed at last observation as compared to only 60% of those who never finished a professional training. Renal replacement therapy starting already during the early phase of education is difficult to coordinate with normal schooling. Considering these health- and time-related obstacles, the degree of educational and professional rehabilitation of the patients is good. But, there is a need for special support accompanying educational and professional training. PMID:10209838

  13. Mitochondrial permeability transition pore (MPTP) desensitization increases sea urchin spermatozoa fertilization rate.

    PubMed

    Torrezan-Nitao, Elis; Boni, Raianna; Marques-Santos, Luis Fernando

    2016-10-01

    Mitochondrial permeability transition pore (MPTP) is a protein complex whose opening promotes an abrupt increase in mitochondrial inner membrane permeability. Calcium signaling pathways are described in gametes and are involved in the fertilization process. Although mitochondria may act as Ca(2+) store and have a fast calcium-releasing mechanism through MPTP, its contribution to fertilization remains unclear. The work aimed to investigate the MPTP phenomenon in sea urchin spermatozoa and its role on the fertilization. Several pharmacological tools were used to evaluate the MPTP's physiology. Our results demonstrated that MPTP occurs in male gametes in a Ca(2+) - and voltage-dependent manner and it is sensitive to cyclosporine A. Additionally, our data show that MPTP opening does not alter ROS generation in sperm cells. Inhibition of MPTP in spermatozoa strongly improved the fertilization rate, which may involve mechanisms that increase the spermatozoa lifespan. The present work is the first report of the presence of a voltage- and Ca(2+) -dependent MPTP in gametes of invertebrates and indicates MPTP opening as another evolutionary feature shared by sea urchins and mammals. Studies about MPTP in sea urchin male gametes may contribute to the elucidation of several mechanisms involved in sperm infertility.

  14. [Treatment of Crohn's disease].

    PubMed

    Kocián, J; Kociánová, J

    1999-10-20

    Dietetic treatment of Crohn's disease has the objective to calm down the gut either by a polymeric or elemental diet. In superacute conditions also total parenteral nutrition can be used. In the quiescent stage the patient must have an adequate energy intake and a low-residue (low-fibre) diet. As to medication, sulfasalazine which has some side-effects is abandoned and 5-ASA (5-aminosalicylic acid) preparations are used, either by the oral route or in enemas, while 4-ASA is little used in this country and is known as PAS. The administration of corticoids which also have side-effects will be abandoned in favour of so-called rapidly metabolized corticoids (Tixocortol pivalate, beclomethasone, budenoside and fluticasone) by the oral route and in enemas and foams. As antimicrobial treatment ciprofloxacine is used combined with metronidazole. As to immunosuppressive drugs azathioprine, 6-mercaptopurine, cyclosporin A, cyclophosphamide and methotrexate are tested. A hope for the future is so-called biological treatment (anticytokines, monoclonal antibodies against cytokines, against CD4+ TNF, interleukines IL-10 and 11, immunoglobulin; plasmapheresis is also tested). Treatment must be individual and surgery as late as possible, only in case of complications, and should be very sparing, stenoses should be treated by plastic operations of strictures.

  15. Inhibition of human immunodeficiency virus type 1 replication by SDZ NIM 811, a nonimmunosuppressive cyclosporine analog.

    PubMed Central

    Rosenwirth, B; Billich, A; Datema, R; Donatsch, P; Hammerschmid, F; Harrison, R; Hiestand, P; Jaksche, H; Mayer, P; Peichl, P

    1994-01-01

    (Me-Ile-4)cyclosporin (SDZ NIM 811) is a 4-substituted cyclosporin which is devoid of immunosuppressive activity but retains full capacity for binding to cyclophilin and exhibits potent anti-human immunodeficiency virus type 1 (HIV-1) activity. SDZ NIM 811 selectively inhibits HIV-1 replication in T4 lymphocyte cell lines, in a monocytic cell line, and in HeLa T4 cells. Furthermore, its antiviral activity against laboratory strains and against clinical isolates from geographically distinct regions in primary T4 lymphocytes and in primary monocytes (50% inhibitory concentration = 0.011 to 0.057 micrograms/ml) was demonstrated. SDZ NIM 811 does not inhibit proviral gene expression or virus-specific enzyme functions, either free or bound to cyclophilin. The compound does not influence CD4 expression or inhibit fusion between virus-infected and uninfected cells. SDZ NIM 811 was, however, found to block formation of infectious particles from chronically infected cells. Oral administration to mice, rats, dogs, and monkeys resulted in levels in blood considerably exceeding the drug concentration, which completely blocked virus replication in primary cells. SDZ NIM 811 caused changes of toxicity parameters in rats to a smaller degree than cyclosporine (formerly cyclosporin A). Thus, the potent and selective anti-HIV-1 activity of SDZ NIM 811 and its favorable pharmacokinetic behavior together with its lower nephrotoxicity than that of cyclosporine make this compound a promising candidate for development as an anti-HIV drug. PMID:7527198

  16. Endurance training increases stimulation of uncoupling of skeletal muscle mitochondria in humans by non-esterified fatty acids: an uncoupling-protein-mediated effect?

    PubMed Central

    Tonkonogi, M; Krook, A; Walsh, B; Sahlin, K

    2000-01-01

    Uncoupled respiration (UCR) is an essential property of muscle mitochondria and has several functions in the cell. We hypothesized that endurance training may alter the magnitude and properties of UCR in human muscle. Isolated mitochondria from muscle biopsies taken before and after 6 weeks of endurance exercise training (n=8) were analysed for UCR. To investigate the role of uncoupling protein 2 (UCP2) and UCP3 in UCR, the sensitivity of UCR to UCP-regulating ligands (non-esterified fatty acids and purine nucleotides) and UCP2 and UCP3 mRNA expression in muscle were examined. Oleate increased the mitochondrial oxygen consumption rate, an effect that was not attenuated by GDP and/or cyclosporin A. The effect of oleate was significantly greater after compared with before training. Training had no effect on UCP2 or UCP3 mRNA levels, but after training the relative increase in respiration rate induced by oleate was positively correlated with the UCP2 mRNA level. In conclusion, we show that the sensitivity of UCR to non-esterified fatty acids is up-regulated by endurance training. This suggests that endurance training causes intrinsic changes in mitochondrial function, which may enhance the potential for regulation of aerobic energy production, prevent excess free radical generation and contribute to a higher basal metabolic rate. PMID:11042137

  17. Natural products as promising drug candidates for the treatment of hepatitis B and C.

    PubMed

    Wohlfarth, Carolin; Efferth, Thomas

    2009-01-01

    Hepatitis B virus (HBV) or hepatitis C virus (HCV) infections are a major threat worldwide. Combination therapy of interferon-alpha and ribavirin is currently the treatment of choice for HCV-infected patients. However, this regimen is only effective in approximately 50% of patients and provokes severe side-effects. Numerous natural alternatives for treating HCV have been suggested. Deoxynojirimycin and its derivatives are iminosugars which exert anti-HCV activity by inhibiting alpha-glucosidases. A non-immunosuppressive derivate of cyclosporine A, NIM811, exerts anti-HCV activity by binding to cyclophilin. Other natural products with promising anti-HCV activity are 2-arylbenzofuran derivatives, Mellein, and pseudoguaianolides. For HBV treatment, several drugs are available, specifically targeting the virus polymerase (lamivudine, entecavir, telbivudine, and adefovir dipivoxil). The efficacy of these drugs is hampered by the development of resistance due to point mutations in the HBV polymerase. Due to drug resistance and adverse side-effects, the search for novel drugs is mandatory. Wogonin, ellagic acid, artemisinin and artesunate, chrysophanol 8-O-beta-D-glucoside, saikosaponin C, and protostane triterpenes are active against HBV. Natural products need to be investigated in more detail to explore their potential as novel adjuncts to established HBV or HCV therapy.

  18. Natural products as promising drug candidates for the treatment of hepatitis B and C

    PubMed Central

    Wohlfarth, Carolin; Efferth, Thomas

    2009-01-01

    Hepatitis B virus (HBV) or hepatitis C virus (HCV) infections are a major threat worldwide. Combination therapy of interferon-α and ribavirin is currently the treatment of choice for HCV-infected patients. However, this regimen is only effective in approximately 50% of patients and provokes severe side-effects. Numerous natural alternatives for treating HCV have been suggested. Deoxynojirimycin and its derivatives are iminosugars which exert anti-HCV activity by inhibiting α-glucosidases. A non-immunosuppressive derivate of cyclosporine A, NIM811, exerts anti-HCV activity by binding to cyclophilin. Other natural products with promising anti-HCV activity are 2-arylbenzofuran derivatives, Mellein, and pseudoguaianolides. For HBV treatment, several drugs are available, specifically targeting the virus polymerase (lamivudine, entecavir, telbivudine, and adefovir dipivoxil). The efficacy of these drugs is hampered by the development of resistance due to point mutations in the HBV polymerase. Due to drug resistance and adverse side-effects, the search for novel drugs is mandatory. Wogonin, ellagic acid, artemisinin and artesunate, chrysophanol 8-O-β-D-glucoside, saikosaponin C, and protostane triterpenes are active against HBV. Natural products need to be investigated in more detail to explore their potential as novel adjuncts to established HBV or HCV therapy. PMID:19060918

  19. The mitochondrial permeability transition pore and its role in myocardial ischemia reperfusion injury.

    PubMed

    Ong, Sang-Bing; Samangouei, Parisa; Kalkhoran, Siavash Beikoghli; Hausenloy, Derek J

    2015-01-01

    Ischemic heart disease (IHD) remains the leading cause of death and disability worldwide. For patients presenting with an acute myocardial infarction, the most effective treatment for limiting myocardial infarct (MI) size is timely reperfusion. However, in addition to the injury incurred during acute myocardial ischemia, the process of reperfusion can itself induce myocardial injury and cardiomyocyte death, termed 'myocardial reperfusion injury', the combination of which can be referred to as acute ischemia-reperfusion injury (IRI). Crucially, there is currently no effective therapy for preventing this form of injury, and novel cardioprotective therapies are therefore required to protect the heart against acute IRI in order to limit MI size and preserve cardiac function. The opening of the mitochondrial permeability transition pore (MPTP) in the first few minutes of reperfusion is known to be a critical determinant of IRI, contributing up to 50% of the final MI size. Importantly, preventing its opening at this time using MPTP inhibitors, such as cyclosporin-A, has been reported in experimental and clinical studies to reduce MI size and preserve cardiac function. However, more specific and novel MPTP inhibitors are required to translate MPTP inhibition as a cardioprotective strategy into clinical practice. In this article, we review the role of the MPTP as a mediator of acute myocardial IRI and as a therapeutic target for cardioprotection. This article is part of a Special Issue entitled "Mitochondria: From Basic Mitochondrial Biology to Cardiovascular Disease". PMID:25446182

  20. Synergistic and complete reversal of the multidrug resistance of mitoxantrone hydrochloride by three-in-one multifunctional lipid-sodium glycocholate nanocarriers based on simultaneous BCRP and Bcl-2 inhibition

    PubMed Central

    Ling, Guixia; Zhang, Tianhong; Zhang, Peng; Sun, Jin; He, Zhonggui

    2016-01-01

    Multidrug resistance (MDR) is a severe obstacle to successful chemotherapy due to its complicated nature that involves multiple mechanisms, such as drug efflux by transporters (P-glycoprotein and breast cancer resistance protein, BCRP) and anti-apoptotic defense (B-cell lymphoma, Bcl-2). To synergistically and completely reverse MDR by simultaneous inhibition of pump and non-pump cellular resistance, three-in-one multifunctional lipid-sodium glycocholate (GcNa) nanocarriers (TMLGNs) have been designed for controlled co-delivery of water-soluble cationic mitoxantrone hydrochloride (MTO), cyclosporine A (CsA – BCRP inhibitor), and GcNa (Bcl-2 inhibitor). GcNa and dextran sulfate were incorporated as anionic compounds to enhance the encapsulation efficiency of MTO (up to 97.8%±1.9%) and sustain the release of cationic MTO by electrostatic interaction. The results of a series of in vitro and in vivo investigations indicated that the TMLGNs were taken up by the resistant cancer cells by an endocytosis pathway that escaped the efflux induced by BCRP, and the simultaneous release of CsA with MTO further efficiently inhibited the efflux of the released MTO by BCRP; meanwhile GcNa induced the apoptosis process, and an associated synergistic antitumor activity and reversion of MDR were achieved because the reversal index was almost 1.0. PMID:27601896

  1. Rho-kinase/myosin light chain kinase pathway plays a key role in the impairment of bile canaliculi dynamics induced by cholestatic drugs

    PubMed Central

    Sharanek, Ahmad; Burban, Audrey; Burbank, Matthew; Le Guevel, Rémy; Li, Ruoya; Guillouzo, André; Guguen-Guillouzo, Christiane

    2016-01-01

    Intrahepatic cholestasis represents a frequent manifestation of drug-induced liver injury; however, the mechanisms underlying such injuries are poorly understood. In this study of human HepaRG and primary hepatocytes, we found that bile canaliculi (BC) underwent spontaneous contractions, which are essential for bile acid (BA) efflux and require alternations in myosin light chain (MLC2) phosphorylation/dephosphorylation. Short exposure to 6 cholestatic compounds revealed that BC constriction and dilation were associated with disruptions in the ROCK/MLCK/myosin pathway. At the studied concentrations, cyclosporine A and chlorpromazine induced early ROCK activity, resulting in permanent MLC2 phosphorylation and BC constriction. However, fasudil reduced ROCK activity and caused rapid, substantial and permanent MLC2 dephosphorylation, leading to BC dilation. The remaining compounds (1-naphthyl isothiocyanate, deoxycholic acid and bosentan) caused BC dilation without modulating ROCK activity, although they were associated with a steady decrease in MLC2 phosphorylation via MLCK. These changes were associated with a common loss of BC contractions and failure of BA clearance. These results provide the first demonstration that cholestatic drugs alter BC dynamics by targeting the ROCK/MLCK pathway; in addition, they highlight new insights into the mechanisms underlying bile flow failure and can be used to identify new predictive biomarkers of drug-induced cholestasis. PMID:27169750

  2. Calcineurin orchestrates dimorphic transitions, antifungal drug responses, and host-pathogen interactions of the pathogenic mucoralean fungus Mucor circinelloides

    PubMed Central

    Lee, Soo Chan; Li, Alicia; Calo, Silvia; Inoue, Makoto; Tonthat, Nam K.; Bain, Judith M.; Louw, Johanna; Shinohara, Mari L.; Erwig, Lars P.; Schumacher, Maria A.; Ko, Dennis C.; Heitman, Joseph

    2015-01-01

    Summary Calcineurin plays essential roles in virulence and growth of pathogenic fungi and is a target of the natural products FK506 and Cyclosporine A. In the pathogenic mucoralean fungus Mucor circinelloides, calcineurin mutation or inhibition confers a yeast-locked phenotype indicating that calcineurin governs the dimorphic transition. Genetic analysis in this study reveals that two calcineurin A catalytic subunits (out of three) are functionally diverged. Homology modeling illustrates modes of resistance resulting from amino substitutions in the interface between each calcineurin subunit and the inhibitory drugs. In addition, we show how the dimorphic transition orchestrated by calcineurin programs different outcomes during host-pathogen interactions. For example, when macrophages phagocytose Mucor yeast, subsequent phagosomal maturation occurs, indicating host cells respond appropriately to control the pathogen. On the other hand, upon phagocytosis of spores, macrophages fail to form mature phagosomes. Cytokine production from immune cells differs following exposure to yeast vs. spores (which germinate into hyphae). Thus, the morphogenic transition can be targeted as an efficient treatment option against Mucor infection. In addition, genetic analysis (including gene disruption and mutational studies) further strengthens the understanding of calcineurin and provides a foundation to develop antifungal agents targeting calcineurin to deploy against Mucor and other pathogenic fungi. PMID:26010100

  3. Induction of the nuclear factor HIF-1{alpha} in acetaminophen toxicity: Evidence for oxidative stress

    SciTech Connect

    James, Laura P. . E-mail: jameslaurap@uams.edu; Donahower, Brian; Burke, Angela S.; McCullough, Sandra; Hinson, Jack A.

    2006-04-28

    Hypoxia inducible factor (HIF) controls the transcription of genes involved in angiogenesis, erythropoiesis, glycolysis, and cell survival. HIF-1{alpha} levels are a critical determinant of HIF activity. The induction of HIF-1{alpha} was examined in the livers of mice treated with a toxic dose of APAP (300 mg/kg IP) and sacrificed at 1, 2, 4, 8, and 12 h. HIF-1{alpha} was induced at 1-12 h and induction occurred prior to the onset of toxicity. Pre-treatment of mice with N-acetylcysteine (1200 mg/kg IP) prevented toxicity and HIF-1{alpha} induction. In further studies, hepatocyte suspensions were incubated with APAP (1 mM) in the presence of an oxygen atmosphere. HIF-1{alpha} was induced at 1 h, prior to the onset of toxicity. Inclusion of cyclosporine A (10 {mu}M), an inhibitor of mitochondrial permeability transition, oxidative stress, and toxicity, prevented the induction of HIF-1{alpha}. Thus, HIF-1{alpha} is induced before APAP toxicity and can occur under non-hypoxic conditions. The data suggest a role for oxidative stress in the induction of HIF-1{alpha} in APAP toxicity.

  4. Agrobacterium VirD2 protein interacts with plant host cyclophilins.

    PubMed

    Deng, W; Chen, L; Wood, D W; Metcalfe, T; Liang, X; Gordon, M P; Comai, L; Nester, E W

    1998-06-01

    Agrobacterium tumefaciens induces crown gall tumors on plants by transferring a nucleoprotein complex, the T-complex, from the bacterium to the plant cell. The T-complex consists of T-DNA, a single-stranded DNA segment of the tumor-inducing plasmid, VirD2, an endonuclease covalently bound to the 5' end of the T-DNA, and perhaps VirE2, a single-stranded DNA binding protein. The yeast two-hybrid system was used to screen for proteins interacting with VirD2 and VirE2 to identify components in Arabidopsis thaliana that interact with the T-complex. Three VirD2- and two VirE2-interacting proteins were identified. Here we characterize the interactions of VirD2 with two isoforms of Arabidopsis cyclophilins identified by using this analysis. The VirD2 domain interacting with the cyclophilins is distinct from the endonuclease, omega, and the nuclear localization signal domains. The VirD2-cyclophilin interaction is disrupted in vitro by cyclosporin A, which also inhibits Agrobacterium-mediated transformation of Arabidopsis and tobacco. These data strongly suggest that host cyclophilins play a role in T-DNA transfer. PMID:9618535

  5. Debate around infection-dependent hemophagocytic syndrome in paediatrics

    PubMed Central

    2013-01-01

    Background Hemophagocytic syndrome (HPS) is clinically defined as a combination of fever, liver dysfunction, coagulation abnormalities, pancytopenia, progressive macrophage proliferation throughout the reticuloendothelial system, and cytokine over-production, and may be primary or secondary to infectious, auto-immune, and tumoral diseases. The most consistent association is with viral infections but, as it is still debated whether any micro-organisms are involved in its pathogenesis, we critically appraised the literature concerning HPS and its relationship with infections. Discussion Infection-dependent HPS has been widely observed, but there are no data concerning its incidence in children. A better understanding of the pathophysiology of HPS may clarify the interactions between the immune system and the variously implicated potential infectious agents. Epstein-Barr virus (EBV) infection has been prominently associated with HPS, with clonal proliferation and the hyperactivation of EBV-infected T cells. However, a number of other viral, bacterial, fungal, and parasitic infections have been reported in association with HPS. In the case of low-risk HPS, corticosteroids and/or intravenous immunoglobulin or cyclosporine A may be sufficient to control the biological process, but etoposide is recommended as a means of reversing infection-dependent lymphohistiocytic dysregulation in high-risk cases. Summary HPS is a potential complication of various infections. A polymerase chain reaction search for infectious agents including EBV, cytomegalovirus and Leishmania is recommended in clinical settings characterised by non-remitting fever, organomegaly, cytopenia and hyperferritinemia. PMID:23324497

  6. Ellagic acid, a polyphenolic compound, selectively induces ROS-mediated apoptosis in cancerous B-lymphocytes of CLL patients by directly targeting mitochondria.

    PubMed

    Salimi, Ahmad; Roudkenar, Mehryar Habibi; Sadeghi, Leila; Mohseni, Alireza; Seydi, Enayatollah; Pirahmadi, Nahal; Pourahmad, Jalal

    2015-12-01

    To investigate the effects ofellagic acid (EA) on the cytotoxicity, B-lymphocytes isolated from CLL patients and healthy individuals. Flow cytometric assay was used to measure the percentage of apoptosis versus necrosis, intracellular active oxygen radicals (ROS), mitochondrial membrane potential (MMP) and the caspase-3 activity and then mitochondria were isolated from both groups B-lymphocytes and parameters of mitochondrial toxicity was investigated. Based on our results EA decreased the percentage of viable cells and induced apoptosis. EA increased ROS formation, mitochondria swelling, MMP decrease and cytochrome c release in mitochondria isolated from CLL BUT NOT healthy B-lymphocytes while pre-treatment with cyclosporine A and Butylated hydroxyl toluene (BHT) prevented these effects. Our results suggest that EA can act as an anti cancer candidate by directly and selectively targeting mitochondria could induce apoptosis through mitochondria pathway with increasing ROS production which finally ends in cytochrome c release, caspase 3 activation and apoptosis in cancerous B-lymphocytes isolated from CLL patients.

  7. Novel quinazoline-urea analogues as modulators for Aβ-induced mitochondrial dysfunction: design, synthesis, and molecular docking study.

    PubMed

    Elkamhawy, Ahmed; Lee, Jiyoun; Park, Beoung-Geon; Park, Insun; Pae, Ae Nim; Roh, Eun Joo

    2014-09-12

    A novel series of twenty-six quinazoline-urea derivatives was designed and synthesized. Their blocking activities against β-amyloid peptide (Aβ) induced mitochondrial permeability transition pore (mPTP) opening were evaluated by JC-1 assay which measured the change of mitochondrial membrane potential. Seven compounds showed better inhibitory activities than the standard Cyclosporin A (CsA). The most active analogues were tested by MTT assay to evaluate their toxicity on the cellular survival; they revealed excellent cellular viability. To explain the difference in inhibitory activity, molecular docking study using (GOLD) program was performed for selected sets of the most active and inactive compounds on cyclophilin D (CypD) receptor as a major component of mPTP. Moreover, ADME profiling, in silico toxicity, drug-likeness, and drug-score studies were discussed. From these results, we report compound 31 as the most active nonpeptidyl mPTP blocker possessing quinazoline-urea scaffold; 2 folds of CsA activity, which would constitute a new direction for the design of novel mPTP modulators.

  8. Synergistic and complete reversal of the multidrug resistance of mitoxantrone hydrochloride by three-in-one multifunctional lipid-sodium glycocholate nanocarriers based on simultaneous BCRP and Bcl-2 inhibition

    PubMed Central

    Ling, Guixia; Zhang, Tianhong; Zhang, Peng; Sun, Jin; He, Zhonggui

    2016-01-01

    Multidrug resistance (MDR) is a severe obstacle to successful chemotherapy due to its complicated nature that involves multiple mechanisms, such as drug efflux by transporters (P-glycoprotein and breast cancer resistance protein, BCRP) and anti-apoptotic defense (B-cell lymphoma, Bcl-2). To synergistically and completely reverse MDR by simultaneous inhibition of pump and non-pump cellular resistance, three-in-one multifunctional lipid-sodium glycocholate (GcNa) nanocarriers (TMLGNs) have been designed for controlled co-delivery of water-soluble cationic mitoxantrone hydrochloride (MTO), cyclosporine A (CsA – BCRP inhibitor), and GcNa (Bcl-2 inhibitor). GcNa and dextran sulfate were incorporated as anionic compounds to enhance the encapsulation efficiency of MTO (up to 97.8%±1.9%) and sustain the release of cationic MTO by electrostatic interaction. The results of a series of in vitro and in vivo investigations indicated that the TMLGNs were taken up by the resistant cancer cells by an endocytosis pathway that escaped the efflux induced by BCRP, and the simultaneous release of CsA with MTO further efficiently inhibited the efflux of the released MTO by BCRP; meanwhile GcNa induced the apoptosis process, and an associated synergistic antitumor activity and reversion of MDR were achieved because the reversal index was almost 1.0.

  9. The calcineurin-NFAT pathway negatively regulates megakaryopoiesis.

    PubMed

    Zaslavsky, Alexander; Chou, Stella T; Schadler, Keri; Lieberman, Allyson; Pimkin, Maxim; Kim, Yeo Jung; Baek, Kwan-Hyuck; Aird, William C; Weiss, Mitchell J; Ryeom, Sandra

    2013-04-18

    The calcium regulated calcineurin-nuclear factor of activated T cells (NFAT) pathway modulates the physiology of numerous cell types, including hematopoietic. Upon activation, calcineurin dephosphorylates NFAT family transcription factors, triggering their nuclear entry and activation or repression of target genes. NFATc1 and c2 isoforms are expressed in megakaryocytes. Moreover, human chromosome 21 (Hsa21) encodes several negative regulators of calcineurin-NFAT, candidates in the pathogenesis of Down syndrome (trisomy 21)-associated transient myeloproliferative disorder and acute megakaryoblastic leukemia. To investigate the role of calcineurin-NFAT in megakaryopoiesis, we examined wild-type mice treated with the calcineurin inhibitor cyclosporin A and transgenic mice expressing a targeted single extra copy of Dscr1, an Hsa21-encoded calcineurin inhibitor. Both murine models exhibited thrombocytosis with increased megakaryocytes and megakaryocyte progenitors. Pharmacological or genetic inhibition of calcineurin in mice caused accumulation of megakaryocytes exhibiting enhanced 5-bromo-2'-deoxyuridine uptake and increased expression of messenger RNAs encoding CDK4 and G1 cyclins, which promote cell division. Additionally, human megakaryocytes with trisomy 21 show increased proliferation and decreased NFAT activation compared with euploid controls. Our data indicate that inhibition of calcineurin-NFAT drives proliferation of megakaryocyte precursors by de-repressing genes that drive cell division, providing insights into mechanisms of normal megakaryopoiesis and megakaryocytic abnormalities that accompany Down syndrome.

  10. Isoflurane induced cognitive impairment in aged rats through hippocampal calcineurin/NFAT signaling.

    PubMed

    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.

  11. Calcineurin/NFAT signalling inhibits myeloid haematopoiesis.

    PubMed

    Fric, Jan; Lim, Clarice X F; Koh, Esther G L; Hofmann, Benjamin; Chen, Jinmiao; Tay, Hock Soon; Mohammad Isa, Siti Aminah Bte; Mortellaro, Alessandra; Ruedl, Christiane; Ricciardi-Castagnoli, Paola

    2012-04-01

    Nuclear factor of activated T cells (NFAT) comprises a family of transcription factors that regulate T cell development, activation and differentiation. NFAT signalling can also mediate granulocyte and dendritic cell (DC) activation, but it is unknown whether NFAT influences their development from progenitors. Here, we report a novel role for calcineurin/NFAT signalling as a negative regulator of myeloid haematopoiesis. Reconstituting lethally irradiated mice with haematopoietic stem cells expressing an NFAT-inhibitory peptide resulted in enhanced development of the myeloid compartment. Culturing bone marrow cells in media supplemented with Flt3-L in the presence of the cal