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It is known that exogenous RNA molecules can be taken up by eukaryotic cells and can exert a variety of biological effects both in vitro and in vivo. The modulation of human lymphocytes by exogenous RNAs has medical implications. The exogenous RNA used in this study was obtained from lymphoid organs of animals immunized with the synthetic peptide p12 of HIV-1 and was referred to as p12-RNA. Human lymphocytes were transfected with the p12-RNA and the transfer of immunoreactivity of p12 was assessed by the lymphocyte proliferation and the leukocyte adherence inhibition assays. Our results indicate that the transfer of cellular immune response to the p12 occurred in 9 donors (60%) who were named responsive individuals whereas 6 donors (40%) were non-responsives. We also found that the calcium phosphate-mediated RNA uptake method is effective in converting non-responsive into responsive donors. The calcium phosphate-mediated RNA uptake may also be used to increase the efficiency of RNA transfection in other models with medical implications and to contribute to a better understanding of the molecular events involved in the uptake of RNA. Our findings give support for the use of exogenous RNAs obtained from lymphoid organs of immunized animals with synthetic peptides of HIV-1 in the immune reconstitution of individuals infected with HIV-1.
It is known that exogenous RNA molecules can be taken up by eukaryotic cells and can exert a variety of biological effects both in vitro and in vivo. The modulation of human lymphocytes by exogenous RNAs has medical implications. The exogenous RNA used in this study was obtained from lymphoid organs of animals immunized with the synthetic peptide p12 of HIV-1 and was referred to as p12-RNA. Human lymphocytes were transfected with the p12-RNA and the transfer of immunoreactivity of p12 was assessed by the lymphocyte proliferation and the leukocyte adherence inhibition assays. Our results indicate that the transfer of cellular immune response to the p12 occurred in 9 donors (60%) who were named responsive individuals whereas 6 donors (40%) were non-responsives. We also found that the calcium phosphate-mediated RNA uptake method is effective in converting non-responsive into responsive donors. The calcium phosphate-mediated RNA uptake may also be used to increase the efficiency of RNA transfection in other models with medical implications and to contribute to a better understanding of the molecular events involved in the uptake of RNA. Our findings give support for the use of exogenous RNAs obtained from lymphoid organs of immunized animals with synthetic peptides of HIV-1 in the immune reconstitution of individuals infected with HIV-1.
Exogenous RNA molecules can be incorporated into eukaryotic cells and can exert a variety of biological effects. We have previously showed that exogenous RNAs obtained from lymphoid organs of animals immunized with synthetic peptides of HIV-1 are able to induce cell-mediated immune responses. In this study, animals were immunized with a synthetic peptide (pol: 476-484) of HIV-1, referred to as p9, which is a cytotoxic T lymphocyte (CTL) epitope. The RNA extracted from the lymphoid organs of animals immunized with p9 was termed p9-RNA. We have demonstrated that p9-RNA is active in inducing human CTL. The p9-RNA was also able to activate the RNA-dependent protein kinase (PKR) of human lymphocytes. The polyA(+) p9-RNA was the fraction responsible for the activation of this protein kinase. We also found that p9-RNA activates the transcription factor nuclear kappa B (NF-kappaB) by inducing the degradation of its inhibitor I-kappaB. Thus, these findings suggest that p9-RNA may act as a regulatory RNA and that the induction of CTL activity by p9-RNA could be mediated by PKR through NF-kappaB activation. It is known that CTL activity plays an important role in host defense against HIV-1 infection. Elucidating the molecular mechanism of p9-RNA could contribute to determining the basis for the use of p9-RNA as an immunomodulator in HIV-infected patients.
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