Quantitative and qualitative evaluation of proteinuria in non human primates recipients of porcine xenografts

Introduction: Immunological and histopathological features in pig-to-primate renal xenotransplantation are widely studied. However, only limited data have been reported on clinical-pathological findings in primate recipients of life supporting renal xenografts. In human medicine, proteinuria represents a common complication in kidney transplantation and is associated with impaired patient and graft survival. The detection of low molecular weight proteins of tubular origin such as \u3b11-microglobulin, \u3b22-microglobulin, retinol-binding protein, lysozime, is considered an early marker for predicting potential graft rejection. In the present study the presence and significance of quantitative and qualitative proteinuria were evaluated in xenotransplanted non-human primates in which kidney function is supported only by the transplanted organ. Material and methods: Eight captive-bred, bilaterally nephrectomized cynomolgus monkeys (Macaca fascicularis), recipient of a life-supporting transgenic porcine kidney, were included in the present study. Quantitative and qualitative analysis of proteinuria, evaluated with urinary protein to creatinine ratio (UPC ratio) and sodium dodecyl sulphate-agarose gel electrophoresis (SDSAGE) respectively, have been performed. Results: In urine samples, the measurement of UPC ratio was low before transplantation and increased after transplantation. Similarly, SDS-AGE was negative before transplantation but evidenced bands consistent with mixed (i.e. tubular and glomerular) proteinuria in all the samples collected posttransplantation. Proteinuria and presence of LMW proteins was consistently found in urine after transplantation, independently of the fluctuations of creatinine values and/or of the status of renal functions. Conclusions: The evaluation of UPC ratio and the use of SDS-AGE technique in urine samples of cynomolgus monkey recipients of a life-supporting renal xenograft, may be considered a valid, cheap and rapid technique to monitor proteinuria in the post-transplanted period

Refinement of a macaque transplantation model: application of a subcutaneous port as a means for long-term enteral drug administration and nutritional supplementation.

A new application of a device enabling the long-term enteral administration of drugs or nutritional supplementation was developed for implementing in research entailing the use of macaques (Macaca fascicularis). After implanting a subcutaneous port, a surgically-placed gastrostomy (SPG) was completed to afford access to the gastric lumen and enable the administration of substances. In this study, the device was left in place for a period ranging between two and 12 months in macaques (n = 16). In five cases, the SPG was used successfully for 8-12 months, until the experimental endpoint was reached. In six cases, the SPG had to be removed earlier due to local infection at the implant site, which promptly regressed after the SPG was removed and antibiotic treatment was administered. One SPG-implanted macaque was euthanized for reasons unrelated to the SPG or the xenotransplantation procedure. In four cases, the SPG was implanted without any complications but has yet to be used to administer substances to the animals. From an ethical standpoint, the SPG device described here minimizes the forced handling of macaques otherwise needed for the oral administration of viscous or unpalatable substances by gavage. The device thus represents an effective refinement that fully complies with the tenet of the '3 Rs' that should be considered by primate centres exposing non-human primates to the long-term daily administration of substances by oral gavage

In vitro and in vivoimmunomodulatory effects of cobalt protoporphyrin administered in combinationwith immunosuppressive drugs.

BACKGROUND: Immunosuppressive strategies are designed to take advantage of potential synergies between drugs to possibly decrease the risk of side-effects. In the present study, the ability of Cobalt protoporphyrin (CoPP) to potentiate the effect of the immunosuppressive drugs mycophenolate sodium (MPS) or cyclosporin A (CsA) was explored in vitro and in vivo. METHODS: In vitro analyses of proliferation and apoptosis were performed on primate T cell cultures, following incubation with the immunosuppressive drugs MPS or CsA, alone or in combination with CoPP. In vivo the effect of CoPP and CsA combination therapy was assessed in a rat heterotopic cardiac allotransplantation model. RESULTS: In vitro results suggest that co-administration of CoPP with CsA or MPS increases immunosuppressive effects of these drugs when combined with CoPP. In particular, the co-administration of CoPP with CsA resulted in the synergistic induction of lymphocyte apoptosis. In vivo, animals immunosuppressed with CsA (1.5 mg/kg) or CoPP (20 mg/kg) alone, had a median survival of 7 or 8 days, respectively. In contrast, animals immunosuppressed with CsA (1.5 mg/kg) combined with CoPP (20 mg/kg) had significantly prolonged median survival (12 days), compared to recipients treated with CsA or CoPP alone (p<0.05). CONCLUSION: Our in vitro and in vivo studies demonstrate that CoPP can potentiate the immunomodulatory effects of CsA, ultimately extending allograft survival