Direct Effect Of Cyclosporin A Research Articles

Cyclosporin in cell therapy for cardiac regeneration.

Stem cell therapy is a promising strategy in promoting cardiac repair in the setting of ischemic heart disease. Clinical and preclinical studies have shown that cell therapy improves cardiac function. Whether autologous or allogeneic cells should be used, and the need for immunosuppression in non-autologous settings, is a matter of debate. Cyclosporin A (CsA) is frequently used in preclinical trials to reduce cell rejection after non-autologous cell therapy. The direct effect of CsA on the function and survival of stem cells is unclear. Furthermore, the appropriate daily dosage of CsA in animal models has not been established. In this review, we discuss the pros and cons of the use of CsA on an array of stem cells both in vitro and in vivo. Furthermore, we present a small collection of data put forth by our group supporting the efficacy and safety of a specific daily CsA dosage in a pig model.

Cyclosporin A-treated dendritic cells may affect the outcome of organ transplantation by decreasing CD4+CD25+ regulatory T cell proliferation.

One of the mechanisms for generation of tolerance involves immature dendritic cells (DCs) and a subpopulation of regulatory CD4+ CD25+ T lymphocytes (T REG). The purpose of this work was to analyze how Cyclosporine A (CsA), a widely used immunosuppressive drug, may affect T REG proliferation. Purified and activated murine DCs obtained from bone marrow precursors differentiated with rGMCSF were co-cultured with purified CFSE-labeled T REG from OTII mice, and their phenotype and proliferation analyzed by flow cytometry. Our data indicate that DCs differentiated in the presence of CsA show an altered phenotype, with a lower expression of MHC-II and a lower activating capacity. Additionally, these CsA-treated DCs show decreased production of IL-2 and IL-12 and increased IL-10 secretion when stimulated with LPS, indicating an effect on the polarization of the immune response. Interestingly, CsA-treated DCs show an anti-tolerogenic effect since they reduce the proliferation of T REG cells from 72 to 47%. Further inhibition to a 24% of T REG proliferation was obtained as a direct effect of CsA on T REG. In conclusion, the anti-tolerogenic effect of CsA should be considered in the planning of immunosuppression in the context of clinical transplantation.

Cyclosporin A-treated Dendritic Cells may affect the outcome of organ transplantation by decreasing CD4+CD25+ regulatory T cell proliferation

One of the mechanisms for generation of tolerance involves immature dendritic cells (DCs) and a subpopulation of regulatory CD4+ CD25+ T lymphocytes (T REG). The purpose of this work was to analyze how Cyclosporine A (CsA), a widely used immunosuppressive drug, may affect T REG proliferation. Purified and activated murine DCs obtained from bone marrow precursors differentiated with rGMCSF were co-cultured with purified CFSE-labeled T REG from OTII mice, and their phenotype and proliferation analyzed by flow cytometry. Our data indicate that DCs differentiated in the presence of CsA show an altered phenotype, with a lower expression of MHC-II and a lower activating capacity. Additionally, these CsA-treated DCs show decreased production of IL-2 and IL-12 and increased IL-10 secretion when stimulated with LPS, indicating an effect on the polarization of the immune response. Interestingly, CsA-treated DCs show an anti-tolerogenic effect since they reduce the proliferation of T REG cells from 72 to 47%. Further inhibition to a 24% of T REG proliferation was obtained as a direct effect of CsA on T REG. In conclusion, the anti-tolerogenic effect of CsA should be considered in the planning of immunosuppression in the context of clinical transplantation.

Cyclosporin A Inhibits Hypoxia-induced Pulmonary Hypertension and Right Ventricle Hypertrophy

Hypoxia-induced pulmonary hypertension involves hypoxia-inducible factor-1alpha (HIF-1alpha) activation as well as elevated resting calcium levels. Cyclosporin A (CsA) inhibits calcium-induced calcineurin activation and blocks the stabilization of HIF-1alpha in cultured cells. We hypothesized that treatment of rats with CsA would prevent HIF-1-dependent gene transcription, lower specific responses to acute hypoxia, and prevent pulmonary hypertension and right ventricle hypertrophy resulting from prolonged exposure to hypoxia. Acute and chronic responses to hypoxia were studied in rats treated or not treated with CsA (25 mg x kg(-1) x d(-1)). Transcript levels of genes encoding the serotonin transporter or four HIF-1 target genes, in rats exposed for 6 h to ambient hypoxia, treated or not by CsA, were measured. In vivo hemodynamics, hematocrit, and heart morphologic characteristics were assessed in rats subjected to hypoxia for 3 wk, treated or not treated with CsA. Changes in mRNA levels of the modulatory calcineurin-interacting protein-1 (MCIP-1) were used as a sensitive indicator of calcineurin activity in lung and heart. Acute exposure to hypoxia led to a marked increase in mRNA levels of serotonin transporter, modulatory calcineurin-interacting protein-1, and HIF-1 target genes, which was blunted by CsA treatment. Prolonged exposure to hypoxia raised right ventricle pressure, induced right ventricle hypertrophy, and activated cardiac calcineurin, effects that were fully prevented by CsA treatment. These results suggest that CsA prevents hypoxia-induced pulmonary hypertension and right ventricle hypertrophy, either by inhibiting HIF-1 transcriptional activity in lung, by decreasing calcineurin activity in lung and heart, by direct effects of CsA, or by a combination of these factors.

Effect of Cyclosporin A on Cell Adhesion Molecules and Leukocyte-Endothelial Cell Interactions in Experimental Colitis

Cyclosporin A (CsA) is an immunosuppressive agent that is believed to act primarily through effects on T-helper lymphocyte function and proliferation. The aim of this study was to investigate whether modulation of leukocyte recruitment and expression of cell adhesion molecules contribute to the therapeutic efficacy of CsA in a model of experimental colitis. The therapeutic effects of CsA were assessed in mice with dextran sulfate sodium-induced colitis. Leukocyte-endothelial cell interactions were determined in colonic venules by intravital microscopy. The expression of cell adhesion molecules intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and mucosal addressin cell adhesion molecule 1 (MAd-CAM-1) was measured by the radiolabeled antibody technique. Treatment with CsA (4 mg/kg/day) significantly improved the clinical course of colitis, decreasing weight loss, diarrhea, rectal bleeding, disease activity index, colon weight, and colonic shortening. Microscopic damage score, myeloperoxidase activity, tumor necrosis factor alpha (TNF-alpha), and interleukin-6 in colonic tissue were significantly diminished by CsA. CsA also significantly reduced ICAM-1 and VCAM-1, but not MAdCAM-1, expression in colitic mice. TNF-alpha-induced ICAM-1 and VCAM-1 expression in primary cultures of human umbilical vein endothelial cells was reduced by co-incubation with CsA. The reduction in adhesion molecule expression was followed by a marked decrease in leukocyte adhesion in colonic venules of colitic mice. CsA ameliorates experimental colitis in mice. Reduced adhesion molecule expression resulting from diminished pro-inflammatory cytokine production and from a direct effect of CsA in endothelial cells decreases leukocyte recruitment into the inflamed intestine, contributing to this protective effect.

Enhancement by cyclosporine A and tacrolimus of serotonin-induced formation of small platelet aggregation.

Cyclosporine A (CsA) may increase the incidence of thrombotic events, but whether tacrolimus (Tc) has such effects is still unclear. The serotonergic system has been linked to the thrombotic effects of CsA, but a direct effect of CsA on serotonin-induced platelet aggregation has not been demonstrated because of methodological difficulties. We measured the effects of CsA and Tc on serotonin-induced platelet aggregate formation by particle counting using light scattering. CsA and Tc both enhanced serotonin-induced formation of small platelet aggregates, however, neither CsA nor Tc affected aggregation induced by high or low concentrations of ADP, with or without addition of a serotonin receptor antagonist. Both CsA and Tc enhance platelet aggregation induced via the serotonin pathway.

Enhanced Ca2+Channel Currents in Cardiac Hypertrophy Induced by Activation of Calcineurin-dependent Pathway

Cardiac-specific expression of an activated calcineurin protein in the hearts of transgenic (CLN) mice produces a profound hypertrophy that rapidly progresses to heart failure. While calcineurin is regulated by Ca2+, the potential effects of calcineurin on cardiac myocyte Ca2+handling has not been evaluated. To this end, we examined L-type Ca2+currents (ICa) in left ventricular myocytes. CLN myocytes had larger (≊80%) cell capacitance and enhanced ICadensity (≊20%) compared with non-transgenic (NTG) littermates, but no change in the current–voltage relationship, single-channel conductance or protein levels ofα 1 or β 2 subunit of L-type Ca2+channels. Interestingly, the kinetics of ICainactivation was faster (≊two-fold) in CLN myocytes compared with NTG myocytes. Ryanodine application slowed the rate of ICainactivation in both groups and abolished the kinetic difference, suggesting that Ca2+-dependent inactivation is increased in CLN myocytes due to altered SR Ca2+release. Treatment of CLN mice with Cyclosporine A (CsA), a calcineurin inhibitor, prevented myocyte hypertrophy and changes in ICaactivity and inactivation kinetics. However, there was no direct effect of CsA on ICain either NTG or CLN myocytes, suggesting that endogenous calcineurin activity does not directly regulate Ca2+channel activity. This interpretation is consistent with the observation that ICadensity, inactivation kinetics and regulation by isoproterenol were normal in cardiac-specific transgenic mice expressing calcineurin inhibitory protein domains from either Cain or AKAP79. Taken together these data suggest that chronic activation of calcineurin is associated with myocyte hypertrophy and a secondary enhancement of intracellular Ca2+handling that is tied to the hypertrophy response itself.

Upregulation of vasopressin V1A receptor mRNA and protein in vascular smooth muscle cells following cyclosporin A treatment.

1. The major side effects of the immunosuppressive drug cyclosporin A (CsA) are hypertension and nephrotoxicity. It is likely that both are caused by local vasoconstriction. 2. We have shown previously that 20 h treatment of rat vascular smooth muscle cells (VSMC) with therapeutically relevant CsA concentrations increased the cellular response to [Arg8]vasopressin (AVP) by increasing about 2 fold the number of vasopressin receptors. 3. Displacement experiments using a specific antagonist of the vasopressin V1A receptor (V1AR) showed that the vasopressin binding sites present in VSMC were exclusively receptors of the V1A subtype. 4. Receptor internalization studies revealed that CsA (10(-6) M) did not significantly alter AVP receptor trafficking. 5. V1AR mRNA was increased by CsA, as measured by quantitative polymerase chain reaction. Time-course studies indicated that the increase in mRNA preceded cell surface expression of the receptor, as measured by hormone binding. 6. A direct effect of CsA on the V1AR promoter was investigated using VSMC transfected with a V1AR promoter-luciferase reporter construct. Surprisingly, CsA did not increase, but rather slightly reduced V1AR promoter activity. This effect was independent of the cyclophilin-calcineurin pathway. 7. Measurement of V1AR mRNA decay in the presence of the transcription inhibitor actinomycin D revealed that CsA increased the half-life of V1AR mRNA about 2 fold. 8. In conclusion, CsA increased the response of VSMC to AVP by upregulating V1AR expression through stabilization of its mRNA. This could be a key mechanism in enhanced vascular responsiveness induced by CsA, causing both hypertension and, via renal vasoconstriction, reduced glomerular filtration.

Cyclosporin-A-induced effects on the free Ca2+ concentration in LLC-PK1-cells and their mechanisms.

Here we have examined the effects of Cyclosporin A (CyA) on the free intracellular Ca2+ concentration ([Ca2+]i) of LLC-PK1/PKE20 cells to evaluate mechanisms of CyA nephrotoxicity using Fura-2 microspectrofluorometry or digital fluorescence video imaging. The CyA-associated changes were compared to the effects of tacrolimus (Tac), a structurally unrelated immunosuppressant with similar cellular pathways which also causes nephrotoxicity. CyA (EC50(: 1 nmol/l, n=16) and Tac (EC50: 1 nmol/l, n=5) caused a concentration-dependent increase of [Ca2+]i which was substantially attenuated by reducing the external Ca2+ concentration (10(-6) mol/l). Similarly Cyclosporin H, a non-immunosuppressive analogue of CyA, stimulated a Ca2+ influx. Nicardipine (10(-6) mol/l) reduced the CyA- and the Tac-induced Ca2+ influx to 52+/-16% (n=10) and 13+/-10% (n=13) of control respectively. Diltiazem and verapamil (10(-6) mol/l) were also effective, but flufenamate (10(-4) mol/l), Gd3+ (10(-5) mol/l) and La3+ (10(-5) mol/l) were not. In the absence of extracellular Ca2+ CyA led to a small but significant [Ca2+]i increase, indicating additional release from internal stores. Depletion of inositol-1,4,5-trisphosphate-(InsP3-) sensitive Ca2+ stores by extracellular ATP (10(4) mol/l) in low-Ca2+ solution completely suppressed the CyA-induced [Ca2+]i rise. CyA had no effect on the cellular InsP3 concentration. Furthermore, inhibition of phospholipase-Cbeta (PLCbeta) by U73122 (2x10(-5) mol/l) did not alter the CyA-stimulated [Ca2+]i rise. A direct effect of CyA on InsP3-sensitive Ca2+ stores, the InsP3 receptor, the Ca2+ content of the stores or involvement of additional stores is assumed. Incubation with CyA for 1, 12 and 24 h enhanced the rise in [Ca2+]i peak induced by ATP, arginine vasopressin (AVP) and angiotensin II. In summary, CyA stimulated a [Ca2+]i increase in LLC-PK1 cells through Ca2+ release from InsP3-sensitive stores and Ca2+ influx via a nicardipine-sensitive pathway. The CyA-mediated [Ca2+]i increase is independent of PLCbeta activity and InsP3 metabolism. CyA caused long-term enhancement of the agonist-induced rise in [Ca2+]i. The effects of CyA on Ca2+ signaling appear to be independent of its immunosuppressive action.

Cyclosporin A and brain excitability studied in vitro.

Seizures are frequently observed after organ transplantations. This has been attributed to a direct effect of cyclosporin A (CsA) on the brain, although other mechanisms may also be of importance. The aim of this study was to investigate possible acute and direct effects of CsA on neuronal excitability. Female rat hippocampal slices were perfused with CsA solutions containing 400 (n = 4), 1,000 (n = 4), 2,000 (n = 6), 8,000 (n = 8) microg/L CsA or control (n = 8) for 30 min, or penicillin, 2,000 IE/ml (n = 7). Actual concentrations of CsA were measured in the perfusate drawn from the slice chamber. To study CsA accumulation in the slices, uptake of radioactive CsA was measured in 12 living and 11 dead slices. Despite a significant accumulation of CsA in the living neuronal slices, no effects were observed on prevolley, field excitatory postsynaptic potential (fEPSP), or population spike amplitude. Penicillin, however, led to epileptiform activity within 10 min in all cases. Concentrations of CsA in the perfusate from the slice chamber were about half the calculated levels, demonstrating that the slices had been exposed to actual CsA concentrations in the range of approximately 200-4,000 microg/L CsA. Our results demonstrate a lack of acute effects of CsA on neuronal excitability within clinically relevant concentrations despite an active accumulation of the drug in the slices. Long-term effects on brain tissue, indirect metabolic effects, or synergistic effects may be responsible for the neurotoxicity of the drug.

Effects of cyclosporin and cremophor on working rat heart and incidence of myocardial lipid peroxidation.

Cyclosporin A (CsA) is widely used as the immunosuppressant of choice for preventing graft rejection. However, its clinical use is hampered by certain side effects, especially its nephrotoxicity and other cardiovascular side effects. CsA for intravenous infusion contains cremophor (Cre) and this vehicle has significant adverse effects on endothelial function and vascular muscle. The present study was aimed at investigating the direct effects of CsA and Cre on isolated and perfused rat hearts in the dosage that closely approximates the peak level achieved for the prevention of graft rejection in the rat. Transplantation is a clinical setting in which the myocardium may be exposed to transient ischemia. In this study, we have shown that the vehicle of CsA, namely Cre, has significant adverse effects on cardiac function. We observed a reduction in coronary flow and aortic output. Addition of CsA appeared to induce a further reduction of aortic flow. We have also shown that a significant increase of thiobarbituric acid reactive substances, considered as an index of lipid peroxidation, occurred in the reperfused heart in the presence of Cre+CsA. Our experimental study shows that Cre turned out to be toxic to myocardium by itself. In the heart, potential Cre-CsA interactions possibly potentiating CsA toxicity could not be excluded. The increase of lipid peroxidation in the heart perfused with CsA suggests that reactive oxygen species may be involved in the detrimental effects of this substance on the heart.

Effect of cyclosporin A on interleukin-6 and soluble interleukin-2 receptor in patients with rheumatoid arthritis.

To investigate the effect of cyclosporin A (CyA) therapy on circulating concentrations of interleukin-6 (IL-6) and soluble interleukin-2 receptor (sIL-2R) in patients with rheumatoid arthritis (RA). Twenty four RA patients with active disease were studied. Plasma was collected before and after 16 weeks of CyA treatment. IL-6 was measured by B9 bioassay and sIL-2R by enzyme linked immunosorbent assay (ELISA). The initial median IL-6 concentration of 165 IU/ml decreased significantly to 71 IU/ml after 16 weeks (p < 0.05). Similarly, the initial median plasma sIL-2R value of 665 U/ml decreased significantly to 570 U/ml (p < 0.05). This decrease was accompanied by an improvement in clinical parameters of disease activity. Some association between sIL-2R, IL-6, haemoglobin, and platelets was also observed. This study has demonstrated that, in vivo, CyA therapy in RA can significantly reduce circulating concentrations of IL-6 and sIL-2R. Modulation of both T and non-T cell derived cytokines may be one mechanism by which CyA improves rheumatoid disease. Whether this is a direct effect of CyA on the cells within the rheumatoid joint producing these cytokines or an indirect effect mediated by other cytokines which can influence IL-6 and Il-2R values remains to be determined.

Effects of cyclosporin A on rat osteoblasts (ROS 17/2.8 cells) in vitro.

The effects of the immunosuppressive drug cyclosporin A (CsA) were evaluated on ROS 17/2.8 cells in vitro. ROS cells were treated with CsA (0, 0.5, 1.0, 5.0 micrograms/ml) for 3 days with and without bovine parathyroid hormone (bPTH) (1-34) 10 nM. CsA at 0.5, 1.0, 5.0 micrograms/ml without PTH and at 5.0 micrograms/ml in the presence of PTH significantly inhibited proliferation, as determined by a tetrazolium colorimetric assay. In addition, ROS cell number was significantly reduced at 3 and 4 days with CsA (5.0 micrograms/ml) without affecting cell viability. Incorporation of [3H]-thymidine into DNA was significantly reduced by 3.0 and 5.0 micrograms/ml CsA after 12 and 24 hours exposure. Basal and 1,25-dihydroxyvitamin D3-stimulated alkaline phosphatase levels in confluent ROS cells were reduced (P less than 0.05) with CsA (1.0 and 3.0 microgramS/ml). Pretreatment of ROS 17/2.8 cells with CsA did not alter PTH-stimulated cAMP levels or [125I]-PTHrP binding to ROS cells. CsA treatment of ROS 17/2.8 cells induced a spindle-shaped appearance with loss of attachment in confluent cultures. When ROS cells were cultured in CsA-containing media, cellular attachment at 6 and 12 hours was reduced (P less than 0.05) compared with untreated ROS cells. These findings indicate that CsA was capable of inhibiting proliferation, cell number, mitogenesis, alkaline phosphatase levels, and cell attachment of ROS cells without affecting PTH binding or cAMP levels. This direct effect of CsA on osteoblasts may be important in changes of bone remodeling observed in CsA-treated humans and animals.

Influence of cyclosporine A on growth and extracellular matrix synthesis of human fibroblasts.

Cyclosporine A (CyA) is a powerful nonsteroidal immunosuppressive agent used to prevent graft rejection of organ and bone marrow transplants. A major side effect observed can be attributed to the fibroblast and its functions: proliferation of fibroblasts and formation of fibrotic tissue in the gingiva (fibrous hyperplasia) and in the kidney are induced. The mechanism of both is still obscure. In order to elucidate whether these side effects are due to the drug acting on human fibroblasts itself or whether they are indirect ones mediated by factors released by lymphocytes, cultures of human gingiva fibroblasts were exposed to CyA under defined in vitro conditions. Incubation with CyA for 72 hours resulted in a dose-dependent stimulation of DNA synthesis, whereas glycosaminoglycan (GAG) synthesis was slightly suppressed. Long-term incubation (6 weeks) with 1 micrograms/ml CyA resulted again in stimulation of growth parameters: compared to the drug-free control, cell number increased to 168%, incorporation of 3H-thymidine into DNA to 143%, and overall protein content to 159%. Collagen and GAG synthesis were elevated to approximately 120%. When corrected for cell number or cell protein content, this represents a decline in matrix synthesis, comparable to short-term incubations. These results indicate that a direct effect of CyA on proliferation of human gingival fibroblasts is responsible for some of the observed hyperplasia.

Cyclosporin A renders target cells resistant to immune cytolysis

Exposure of cytolytically susceptible human target cells with therapeutic concentrations of the immunosuppressive drug cyclosporin A renders these cells highly resistant to T cell-mediated, natural killer (NK) cell-mediated, and complement-mediated cytolysis. The resistance is dose dependent, time dependent and reversible. The resistance is accompanied by target cell growth inhibition as measured by thymidine uptake. Surprisingly, target cell growth inhibition induced by serum depletion is associated with cell-mediated cytolytic resistance. These data suggest that cyclosporin A (CsA) may block some target cell biochemical pathway(s) important in the suicidal cytolytic process which is (are) linked to some G0/G1 cell cycle events. In addition, these results suggest that the increased risk of Epstein-Barr virus (EBV)-associated lymphoproliferative disease in human organ transplant recipients may be contributed to by CsA-induced resistance of EBV-transformed B lymphocytes to immune cytolysis. In the post-transplant setting, CsA probably blocks T cell-dependent responses to EBV-transformed B lymphocytes (Bird, A.G., McLachlan, S.M. and Britton, S., Nature 1981, 289: 300) yet leaving the NK cell and antibody-dependent responses intact (Shao-Hsien, C. et al. Transplantation 1983. 35: 127). However, given the direct effect of CsA upon EBV-transformed B lymphocytes, these cells would be rendered resistant to nearly all forms of cytolytic immune control (cytotoxic T lymphocyte, natural killer, antibody-dependent cell-mediated cytotoxicity, complement). Unregulated EBV-transformed B lymphocytes may then proliferate in the CsA-treated host thus leading to a polyclonal B cell hyperplasia. Our data would suggest that this early pre-malignant process is likely to be reversible following CsA dose reduction. Indeed, EBV-dependent polyclonal B cell hyperplasia is seen in early post-transplant lymphoproliferative disorders (Hanto, D.W., et al., Transplantation 1989, 47: 458). Furthermore, in some cases CsA dose reduction does lead to disease regression (Starzl, T., et al., Lancet 1984. i: 583). However, further progression of the disease probably occurs following chromosomal changes leading to oncogene activation and might be resistant to CsA dose reduction.

In Vitro Effects of Cyclosporine on Glomerular Function

Cyclosporine A (CsA) causes vasoconstriction and decrease in glomerular filtration rate. Experiments were conducted in isolated glomeruli to study effects of CsA of glomerular perfusion and independent of systemic or renal factors. CsA caused a dose dependent decrease in glomerular volume consistent with mesangial contraction. The ultrafiltration coefficient Kf was significantly lower after incubation in 4 X 10(-4) M CsA when compared to control (2.81 +/- 0.2 vs 5.19 +/- 0.5 nl/min X mm Hg; p less than 0.001). Hydraulic conductivity Lp was diminished by CsA from 2.69 +/- 0.11 to 1.41 +/- 0.05 microliter/min X mm Hg X cm2 (p less than 0.001). These findings demonstrate a direct effect of CsA on the glomerulus which must be considered in addition to drug-induced changes in perfusion and tubular function.

Effects of cyclosporin A on the cells responsible for the anticryptococcal cell-mediated immune response and its regulation

Cyclosporin A (CsA), a potent immunosuppressive drug, was used to explore further the induction, expression, and regulation of lymphoid cells involved in the delayed-type hypersensitivity (DTH) response to cryptococcal antigen(s). We found that the induction of the cells responsible for DTH (TDH cells) was not affected by CsA, but their expression was inhibited in CsA-treated mice. The inhibition of expression of the TDH cells could not be attributed to the Cryptococcus neoformans-specific suppressor T (Ts) cells, even though the Ts cells were induced in CsA-treated mice. Instead, the suppressed expression of the TDH cells in CsA-treated mice was a direct effect of CsA or its products. Our studies with CsA also resulted in the first identification of a population of cells that significantly amplify the anticryptococcal DTH response. The amplifier cells were induced in mice that were given a primary immunizing dose of cryptococcal antigen in complete Freund adjuvant, and they amplified the anticryptococcal DTH response in recipient mice when they were transferred at the time of immunization of the recipient. The amplifier cell population was distinct from the TDH cells in that CsA inhibited the production of the amplifying cells but did not affect the induction of TDH cells. Amplification of the DTH response was a cell-mediated event, since cells but not serum from immunized mice mediated the amplified response in recipient mice. Thus, CsA enabled us to characterize anticryptococcal TDH and Ts cells further and to add to the immune cell circuit of the cryptococcal system a distinct population of cells that amplifies the anticryptococcal DTH response.

Direct effects of cyclosporin A on human pancreatic beta-cells.

Cyclosporin A (CyA) may induce clinical remission in newly diagnosed insulin-dependent diabetes mellitus patients. Recently, however, adverse effects of high doses of CyA on rodent islets have been reported in vivo and in vitro. The possible direct effects of CyA on the human endocrine pancreas were therefore evaluated. Islets isolated from eight necrokidney donors were cultured in the presence of a therapeutically relevant dose of CyA, i.e., 100 ng/ml. During a 5-day culture period the release of insulin was reduced by 36% (range 7-61%), whereas the islets' content of insulin was increased by 59% (range 3-268%). The glucagon content was not affected. Cyclosporin G inhibited the insulin release, whereas dihydrocyclosporin D had no consistent effects. Glucose-stimulated insulin release from perifused islets was markedly depressed in CyA-treated islets. This effect was not fully reversed 48 h after removal of the drug. We concluded that CyA has a direct inhibitory effect on insulin release from human pancreatic islets with a concomitant increase in the residual insulin content. If applicable to the in vivo condition, CyA may therefore, in addition to its immunosuppressive effect, have direct effects on the endocrine pancreas, which may be relevant for clinical application of the drug.

Protection against schistosomiasis produced by cyclosporin A.

Mice infected with Schistosoma mansoni at day 0 were given cyclosporin A (Cy-A) from day 1 to day 3 and reinfected at day 45. They were perfused to recover worms at day 66, at which time mature and immature worms, established from infection and reinfection, respectively, could easily be distinguished. A high degree of protection (90-100%) was obtained against both primary and secondary infection. Elimination of the parasites was an early event since only 10-30% of schistosomula, as compared to those in control mice, were recovered at day 6 by pulmonary perfusion. Our data indicate that a direct effect of Cy-A cannot be demonstrated. Nevertheless, Cy-A did evoke a high level of protection against schistosome infection.