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Henoch-Schonlein purpura (HSP) is the most common form of IgA-mediated systemic vasculitis in children. HSP mainly affects the skin, joints, gastrointestinal tract and kidney. The overall prognosis of the disorder is favorable, but the long-term prognosis depends on the degree of renal involvement. An older age at onset, purpura persistent for more than >4 weeks, severe abdominal symptoms, decreased factor XIII activity and the occurrence of relapses are associated with renal involvement in children with HSP. Although the pathogenesis of HSP is not fully understood, genetic predisposition, abnormal glycosylation of IgA1, complement activation, various cytokines, autoantibodies and abnormal coagulation are involved. HSP nephritis is a mesangial proliferative glomerulonephritis characterized by IgA deposition with varying degrees of mesangial hypercellularity, segmental sclerosis and crescents. Corticosteroid therapy does not prevent the development of nephritis in HSP. Treatment of severe HSP nephritis remains controversial due to a paucity of randomized controlled trials, but various immunosuppressive interventions including methylprednisolone pulse therapy, azathioprine, cyclosporin A and plasmapheresis have been suggested to be efficacious. The necessity of early treatment in severe HSP nephritis has been emphasized.

Cyclosporin A (CsA) and tacrolimus have greatly improved graft survival since their introduction in the 1980 and 1990s, respectively. While the drugs are structurally unrelated, their mechanism of immunosuppression is remarkably similar. The dramatic immunosuppressive and nephrotoxic effects of CsA and tacrolimus are largely explained by their calcineurin inhibition. The pathology of CsA and tacrolimus toxicity is pathologically indistinguishable.

Introduction: Modern immunosuppressives have markedly improved graft survival after solid organ transplantation. However, these substances are associated with negative effects on transplant function attributing to chronic graft dysfunction. It has been speculated that recurrent microcirculatory graft dysregulation caused by peak drug levels could play an important role in this scenario. Therefore, this study examines renal microcirculation, microcirculation of oral mucosae and peripheral vasoreactivity after application of clinically relevant doses of immusuppressive drugs in healthy subjects and patients after kidney transplantation. Methods: Healthy volunteers (n = 18) after single-dose oral application of Cyclosporin A (CyA), Mycophenolat moffetil (MMF), or Tacrolimus (FK 506) (n = 6/group) as well as patients after allogenic kidney transplantation with CyA-, MMF- or FK 506- based immunosuppression (n = 5/group) were studied according to the following protocol: Microcirculation was examined pre-dose as well as 60 min., 120 min., 180 min. and 240 min post-dose. At each time point, venous blood samples were drawn and macrohemodynamic parameters recorded. Perfusion of oral mucosa was quantified by Orthogonal Polarization Spectral imaging. Furthermore, hemoglobin saturation, flow and velocity were examined quantitatively as well as peripheral vasoreactivity after 2 min. of forearm ischemia using laser-doppler-flowmetry. In addition, intrarenal resistive indices (RI) and arterio-capsular distances (ACD) as parameters of organ perfusion were measured by duplex ultrasound in healthy kidneys (healthy volunteers) and kidney grafts (patients). Results: While patient recruitment is still ongoing, serum drug concentrations measured in healthy volunteers are comparable with therapeutic levels targeted in transplanted patients. MMF application tended to reduce RI in grafts, whereas an increase in intrarenal vascular resistance was observed under CyA-treatment. Laserdoppler-flowmetry and power-doppler ultrasound demonstrated a homogenous study collective with low interindividual variances at baseline measurements, and highly reproducible measurements of peripheral vasoreactivity. While tissue oxygen saturation did not increase during reactive hyperemia in volunteers after CyA administration as opposed to MMF, the increase in venular flow after reperfusion remained unchanged after CyA treatment. Conclusion: The methods used in our study facilitate a parallel, non-invasive, reproducible and easy-to-tolerate assessment of peripheral and central microcirculation in healthy volunteers and patients after kidney transplantation. A preliminary data analysis shows a differential regulation of graft perfusion by CNI-inhibitors and MMF. After final evaluation of all results, further studies will have to show, if microcirculatory measurements can help to reduce drug side effects on transplant function.

Systems consisting of cyclosporin A (CsA), a cyclic peptide which is an effective immunosuppressive agent for transplanted organs, and dipalmitoylphosphatidylcholine (DPPC), a liposomal phospholipid, obey the molecular area additivity rule when spread as mixed monolayers at the air/water interface. This behaviour is ascribed to the immiscibility of the system components, which orientate themselves differently on the surface. The insertion of a pyrene residue in the DPPC molecule, whether in the non-polar chain to obtain 1-hexa-decanoyl-2-(1-pyrenedecanoyl)-sn-glycero-3-phosphocholine (H-361) or in the polar group to form N-(1-pyrenesulfonyl)-l,2-phospho-ethanolamine triethylammonium (P-58), has no effect on the surface behaviour of the cyclosporin-phos-pholipid system, which remains additive in its molecular areas and hence immiscible at the interface.

The study of bioactive microbial metabolites was initially focused on the search for antimicrobial agents which various bacteria and fungi were believed to produce to give them a competitive growth advantage by inhibiting other organisms. The most famous member of this group is penicillin, isolated from the bread mould Penicillium notatum by Fleming in 1929. Another well-known antibiotic, streptomycin, was isolated from Streptomyces griseus by Waksman in 1943. The significance of Streptomyces will be addressed later in this chapter. Many antibiotics, in addition to other antimicrobials, enzymes, enzyme inhibitors and immunomodulatory agents have also been isolated from microbial fermentation products. For example, cyclosporin A is a fungal metabolite, isolated from Tolypocladium inflatum and initially envisaged as an antifungal antibiotic before its immunomodulatory properties became clear.

Calcineurin, also known as PP2B and PPP3, is a phospho-serine/threonine specific protein phosphatase that is active only after binding Ca2+ and Ca2+/calmodulin in response to physiological Ca2+ signals. It is expressed to varying degrees in all tissues and highly conserved among most eukaryotes. Calcineurin has received much attention over the last twelve years since the discovery that the widely used immunosuppressive drugs Cyclosporin A and FK506 specifically bind and inhibit calcineurin after recruiting the cellular chaperones cyclophilin A and FKBP-12. Calcineurin is therefore unique among protein phosphatases both in its dramatic responsiveness to a second messenger and in the clinical utility of its inhibitors in a growing number of therapeutic situations. Calcineurin is widely conserved in nature and recently determined to be a major virulence factor in many pathogenic fungi. This review focuses on the progress towards understanding the broader roles of calcineurin in diverse eukaryotic species, which may offer useful insights into calcineurin function in humans.

Introduction The chemokines IFN-γ-inducible protein-10 (IP-10, CXCL10), monokine induced by IFN-a (MIG, CLCL9) and IFN-inducible T cell α chemoattractant (I-TAC, CXCL11) represent potent chemoattractants for antigen-primed T cells, involved in inflammation and allograft rejection. In this study we tested the influence of Cyclosporin A (CsA) on intragraft expression of IP-10, MIG and I-TAC mRNA in a murine cardiac transplant model.

Background: Allogeneic transplantation of parathyroid tissue could be a possible curative treatment for patients with permanent hypoparathyroidism. Various methods to overcome rejection in this experimental setting have been published, but longtime acceptance has not yet been achieved. The aim of this study was to compare the effect of a shorttime immunosuppression on organ function after rat parathyroid allotransplantation, with those after rat small bowel allotransplantation. Methods: Heterotopic parathyroid transplantation (donor: Wistar-Furth RT1u, recipient: Lewis RT11) to the paravertebral muscles was performed in 5 groups: 1. syngeneic control LEW-LEW (n — 5), 2. allogeneic control WF-LEW (n = 5), 3. allogeneic Tx plus cyclosporin A (CyA) 20 mg/kg/day from day 0-13(n = 3),4. allogeneic Tx plus CyA 10 mg/kg/day from day 0-13(n = 3),5. allogeneic Tx plus CyA 10 mg/kg/day from day - 7 to day 7. In addition orthotopic small bowel transplantation was performed in two groups: 1. WF-LEW plus CyA 10 mg/kg/day from day 0 – 13 (n = 3), 2. WF-LEW plus CyA 20 mg/kg/day from day 0 – 13 (n = 3). In case of rejection organs were explanted for histological examination. Results: Recipients of syngeneic transplants showed organ function up to day 100, while allogeneic transplants were rejected at 13 ± 1 days (median). Recipients treated with CyA showed a cellular organ rejection after 28 ± 2 days dosage independentely. In contrast organ function after small bowel transplantation lasted 88 ± 19 days in group 1, and 100 days (80%) in group 2. In those animals transplantspecific tolerance was seen at day 300 after transplantation. Discussion: While organ function is dependent on the amount of immunosuppression in small bowel transplantation, this could not be observed for parathyroid allotransplantation. Missing vascularisation of transplants, or the heterotopic location of implantation could be possible reasons, and will therefore be the target of further investigations.

Refractory acute myeloid leukemia (R-AML), defined as primary resistance to induction chemotherapy, early relapse (within 6-12 months from remission), and second or subsequent relapse, is typically a multidrugresistant illness, for which effective treatment is presently lacking. We have treated R-AML patients with a combination of intermediatedose ara-C, carboplatin and either mitoxantrone or idarubicin in a cross-over design, obtaining an encouraging response rate (46%) in a small group of 13 R-AML patients who were given an amended, low-toxicity regimen [1]. Five more patients were treated with a similar carboplatin-containing schedule delivered in a sequential manner, but only one remission was achieved, to an overall response rate of 33% (6/18). This result prompted a further change in retreatment strategy. In R-AML, clinical multidrug resistance is often associated with functional overexpression of the MDR1 drug efflux machinery and/or other mechanisms such as LRP (lung-related protein) and MRP (multidrug resistance-associated protein) [2-11]. Theoretically, in view of the high incidence of MDR1 activation in R-AML and its adverse clinical significance, functional MDR1 inhibition with cyclosporin A (CsA), verapamil, tamoxifen, PSC 833, quinine [12-19] or the use of drugs scarcely (Idarubicin=IDR) or not (high-dose ara-C=HiDAC, carboplatin) MDR1-sensitive could underlie therapeutic improvement. In preclinical in vitrostudies on a MDR1+ human leukemic cell line [20,21], we found that both cellular uptake and pro-apoptotic effects of IDR used at therapeutic concentration (50-100 ng/ml, corresponding to the ranges of peak plasma level obtainable after rapid intravenous administration of IDR 10-12 mg/m2) were significantly increased by co-incubation with CsA 1500 ng/ml. Interestingly, CsA enhanced IDR-related cytotoxicity only in the early phase of IDR-blast cell interaction while, contrary to daunorubicin, 12-hour IDR retention by blast cells was not modified by CsA. The study conclusion was that both cellular uptake of and related biologic effects from IDR can be upregulated by CsA in MDR1+ cells but, partly because IDR is highly lypophilic and partly because it is less vulnerable to MDR1, this effect does not require a prolonged CsA exposure. On this basis we initiated a new clinical study, using CsA and aiming to determine the upper IDR concentration tolerated by patients with R-AML. This new sequential regimen included HiDAC 3 g/m2 every 12 hours for two consecutive days followed by IDR 12.5 mg/m2 as starting dose level (both drugs were given for two times on days 1-3 and 8-10) together with CsA 6 mg/kg over 1 hour and then 7.5 mg/kg over 11 hours according to the List schedule [22]. IDR was delivered 4 hours after starting CsA. The three-day sequence was repeated on days 8-10. Recombinanat human granulocyte colony-stimulating factor (G-CSF) was administered from day 11 until response evaluation. We report the therapeutic results obtained in the first 10 patients, who received IDR from 12.5 to 17.5 mg/m2/dose.

The damage incurred by neurons and glial cells after ischemia and reperfusion is caused by events occurring during three periods: the period of ischemia, the maturation period, and the sequence of events leading to the final demise of cells. The ischemic period sets the stage for the subsequent steps since it is associated with bioenergetic failure and loss of ion homeostasis, the latter leading to uncontrolled influx of calcium into cells. Presynaptic release of glutamate and influx of calcium trigger events that can give rise to immediate or delayed neuronal (and glial cell) damage. These events encompass activation of kinases and phosphatases which change the expression of many transcription factors, and of genes that modulate the cell response to the initial ischemic insult. Some of these events may be instrumental in causing delayed tissue damage, such as the expression of inducible nitric oxide synthetase and cyclo-oxygenase-2, both of which would promote an inflammatory and immunological reaction. Several lines of evidence incriminate mitochondrial dysfunction in the pathogenesis of delayed ischemic brain damage. Mitochondria may be damaged by being exposed to a constant (post-ischemic) load of calcium and/or to oxidative stress. This may cause peroxidation of lipids, constituting the backbone of the mitochondrial membranes, oxidation of proteins in respiratory complexes, or the assembly of a mitochondrial permeability transition (MPT) pore. The anti-ischemic effects of the immunosuppressants cyclosporin A (CsA) and FK506 suggest that the rotamase activity of the immunophilins, or the effect of ischemia on calcineurin, play important pathogenetic roles. However, since CsA has effects over and above those of FK506, an MPT pore assembly may play a contributory and, under certain circumstances, dominant role.

Recent clinical and experimental data have shown that a variety of donor-associated risk factors may influence adversely the short- and long-term outcome after transplantation (Tx). In our study, we focused on systemic changes occurring after donor brain death (BD). Donor BD leads in the experimental situation to accelerated acute kidney allograft rejection in unmodified hosts and intensifies their later functional and morphological deterioration. As shown previously, sPSGL-Ig inhibits selectin activity and subsequent inflammatory events in organs damaged by ischemia/reperfusion injury. To define its effects on late graft changes, we compared kidney allo- and isografts from BD and living donors over time in a chronic rejection model in the rat. Gradual onset BD was induced in F344 donors via inflation of an intracranially inserted Fogarty catheter. Animals were mechanically ventilated and hemodynamically monitored for 6 h. The left kidney was then grafted into LEW recipients. Kidneys from living donors (LD, group I) and normotensive BD donors (MAP > 80 mmHg, group II) served as controls. To determine the effect of sPSGL-Ig, donors were treated i.v. (50 µg) 3 h after induction of BD. A second dose was given to the recipient immediately after Tx (n = 8, group III). All recipients received low dose cyclosporin A (1.5 mg/kg) for 10 days post Tx. Proteinuria (24-h urine) was measured monthly up to 6 months after Tx (n = 8/group). Grafts were examined by semiquantitative RT-PCR for mRNA of cell products. Urinary protein levels from animals in group I increased progressively after 12 weeks post Tx (168 ± 55.5 mg at 6 months), but was more pronounced in group II animals (273 ± 87.2 mg). However, minimal protein loss occurred over time in animals from group III (25.6 ± 10.6 mg). RT-PCR analysis of the contralateral kidney at the initial time point, i.e. 6 h after induction of BD, showed higher transcription of representative cytokines and chemokines in untreated BD controls compared with those from sPSGL-Ig treated animals. These results suggest that treatment with sPSGL-Ig inhibits donor BD-associated early inflammatory processes in transplanted kidneys. This new form of intervention of early antigen-independent events within the graft appears to influence both short- and long-term survival of renal allografts and therefore may be of special interest for those grafts of potentially diminished quality.