A Lipid Nanoparticle-Formulated GP5-mRNA Vaccine Induces PRRSV-Specific Humoral and Cellular Immune Responses in...

Porcine reproductive and respiratory syndrome virus (PRRSV) remains a major threat to the global swine industry, and the continuous emergence of genetically diverse strains poses challenges to vaccine development. In this study, a lipid nanoparticle-formulated mRNA vaccine encoding the PRRSV GP5 protein was constructed, and its preliminary immunogenicity was evaluated in BALB/c mice. The full-length ORF5 gene was amplified and cloned into the pIVT5 vector to generate the recombinant plasmid pIVT5-GP5. GP5-mRNA was synthesized via in vitro transcription and transfected into Marc-145 cells to verify antigen expression. Indirect immunofluorescence assay showed specific fluorescence signals in GP5-mRNA-transfected cells, and Western blotting further confirmed the expression of GP5 protein. The GP5-mRNA was subsequently encapsulated into lipid nanoparticles to prepare the GP5 mRNA-LNP vaccine formulation. BALB/c mice were immunized intramuscularly to evaluate humoral and cellular immune responses. The GP5 mRNA-LNP vaccine induced a GP5-specific IgG response, and sera collected on days 28, 35, and 42 after the initial immunization showed detectable neutralizing activity against PRRSV. Flow cytometric analysis showed an increased proportion of CD3⁺CD4⁺ and CD3⁺CD8⁺ T-cell subsets in the spleens of immunized mice. In addition, splenocytes from GP5 mRNA-LNP-immunized mice produced higher levels of IL-2, IFN-γ, and IL-4 after PRRSV antigen stimulation. Because mice are not the natural host of PRRSV, these results indicate that the GP5 mRNA-LNP vaccine candidate induced both humoral and cellular immune responses in BALB/c mice, providing a preliminary immunological basis for further optimization and evaluation in pigs.