The COVID-19 pandemic underscored the critical role of the SARS-CoV-2 spike (S) protein in facilitating viral entry and modulating immune responses. As mRNA-based vaccines utilize the S protein to elicit immunity, understanding its direct effects on immune cells is essential. In this study, we investigated the impact of an S protein-expressing mRNA vaccine on macrophage differentiation, cytokine production, and phagocytic activity. THP-1 cells were treated with mRNA encoding the S protein, or a scrambled mRNA control, and changes in the cellular transcriptome, cytokine profiles, and phagocytosis were assessed. Proteome changes were also analyzed using Olink proximity extension assay technology. Parallel experiments in primary macrophages from healthy donors were also conducted. Our results demonstrate that S protein expression significantly alters macrophage differentiation, including cytokine secretion and phagocytic capacity, suggesting potential implications for immune modulation by mRNA vaccines. These findings highlight the need to further elucidate the effects of mRNA vaccines on immune cells, particularly macrophages, to optimize vaccine design and mitigate the risks of immune dysregulation.