| Background and objectives A large clinic information show that the prevention and treatment of transplantation rejection can influence the result of organ transplantation. The most effective measure is to induce recipient to produce donor specific immune tolerance. Previous experiments indicate that HLA-derived peptides have significant immunomodulatory activity. Some peptides can even induce and maintain immune tolerance, which provides a novel immunotherapeutic strategy. RDP1258, the analogue of HLA-B2702. 75-84 peptide, designed by computer, has significant immunosuppression, low decomposing speed and enhanced efficacy. Perioperative administration of RDP1258 in combine with subtherapeutic dose of CsA can greatly prolong survival of transplantated organ. RDP1258 will possibly become a novel anti-rejection drug, which has greatly clinical employment potential. Currently, the study of RDP1258 is in the primary stage, and there is no domestic reports on this field. In this study, RDP1258 was synthesized, purified and validated. The effect of it on lymphocyte function was tested in vitro. Then, we observed the influence of RDP1258 on survival of transplantated rat heart, examined the histology and ultrastructure change in grafts at acute rejection period, measured the expression of TNFαmRNA, IFNγmRNA and intragraft cell apoptosis in order to discuss the possible mechanisms of transplantation immune tolerance induced by RDP1258.Materials and Methods 1) RDP1258 was synthesized using Fmoc chemistry with the peptide synthesizer ABI 431A and purified by reversed phase HPLC method. The molecular weight was validated by mass-spectrum. 2) Rat spleen lymphocytes and human peripheral blood mononuclear cells(PBMCS) were isolated by Ficoll-paque density gradient centrifugation. Then, rat T lymphocyte proliferation to ConA, rat mixed lymphocyte reaction(MLR) and human cytotoxic T lymphocyte(CTL) function were assayed by MTT method. RDP1258 and control peptide were administrated respectively in every experiment. 3) Healthy cleaning SD rats were used as heart donors and Wistar ratswere used as recipients. The model of rat heart abdominal transplantation was established. Group1(n=6) received no immunosuppression, Group2(n=7) received CsA alone, Group3(n=8) received RDP1258 alone, Group4(n=8) received RDP1258+ subtherapeutic CsA. CsA in all cases was gavaged at a dose of 5mg/kg per day from days 0 to 5. RDP1258 was administrated intraperitoneally at a dose of 5mg/kg per day on days -7,-1 and 0 until day 5. Transplantated hearts were monitored daily by direct palpation, and rejection was defined as termination of palpable cardiac contractility. The transplanted hearts were harvested for light and electron microscopic examination at day 7 post-transplantation. On the other hand, the expression of TNFαmRNA , IFNγmRNA and intragraft cell apoptosis were also tested respectively by RT-PCR and TUNEL technology. Results The purity of synthesized RDP1258 was above 95% . The molecular weight was 1229.9Da in accord with theoretical values. RDP1258 could inhibit proliferation response of rat T lymphocyte to ConA in a dose -dependent manner. The inhibitive effect of RDP1258 reached maximum with the concentration of 50μg/ ml. Addition of RDP1258 to rat mixed lymphocyte reactions also inhibited this proliferation. Moreover, RDP1258 could significantly inhibit the lytic function of human CTL. There was slight or no rejection with 0 or 1 pathological grade in RDP1258 group and RDP1258+ CsA group, and the allograft structure was normal at day 7 post-transplantation. We observed that RDP1258 combined CsA therapy significantly prolonged allograft survival with two animals surviving more than 100 days. The mRNA expression of TNFαin control group was higher than other groups. It gradually declined in CsA group, RDP1258 group and RDP1258+ CsA group with the lowest expression in RDP1258+CsA group. However, IFNγmRNA expression did not differ significantly among four groups. The intragraft cell apoptosis index was the highest in contr...
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