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31 December 2012

Modulation of Porcine Endogenous Retroviruses (PERVs) expression in vitro by shRNA in the presence of cyclosporine A and dexamethasone

Daniel SypniewskiABCDEF, Ilona BednarekABCDEFG, Daria MatczyńskaABCDEF, Sabina GałkaABCDEF, Tomasz LochBCD, Dagna SołtysikBCD, Grzegorz MachnikABC, Ewa NowakBCDF

DOI: 10.12659/AOT.883699

Ann Transplant 2012; 17(4): 92-107

Abstract

Background: Despite the fact that the risk of Porcine Endogenous Retroviruses (PERV) infection and propagation in human recipients is extremely low, such an event cannot be completely ruled out, especially in immunosuppressed patients. Therefore, the aim of this study was to analyze the expression of PERVs in vitro in the presence of immunosuppression agents: cyclosporine A (CsA), and dexamethasone (DEX). We investigated the possible interactions between immunosuppression drugs, CsA and DEX, and the efficiency of anti-PERV RNAi.
Material/Methods: Plasmid-based vectors expressing shRNAs against all PERV genes were constructed and analyzed. PERVs expression in cultures transfected with anti-PERV RNAi constructions and treated with CsA or DEX was analyzed by Real-Time RT-PCR, Western blot, and by the measurement of RT activity.
Results: Both CsA and DEX inhibited PERVs expression in cell cultures in vitro. RNAi constructions efficiently knocked down PERV expression in Circe, and de novo PERV-infected HeLa and HEK-293 cell cultures. Pretreatment of Circe cultures with CsA or DEX increased PERVs knockdown by RNAi, but no specific interaction between the drugs and transfection efficiency was observed.
Conclusions: Our results demonstrate that cyclosporine A and dexamethasone decrease expression of PERVs in vitro. We also proved that these drugs did not synergize or antagonize RNAi-mediated knockdown of PERVs. These observations may be beneficial in immunosuppressed xenograft recipients; however, due to the controversial literature data concerning influence of immune suppression on graft recipients, our results should be further analyzed.

Keywords: RNA interference (RNAi), porcine endogenous retrovirus (PERV), Xenotransplantation, Immunosuppression, Cyclosporine A, Dexamethasone

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Annals of Transplantation eISSN: 2329-0358
Annals of Transplantation eISSN: 2329-0358