Nutrient deprivation increases CD3 expression in RAW cells and augments the CD3-induced proinflammatory profile,...

Myeloid cells can express lymphoid markers, including components of the CD3-TCR complex. CD3+ macrophages have been identified in both infectious and non-infectious pathologies; however, their origin and signaling mechanisms remain poorly studied, primarily due to their low prevalence in human samples. Utilizing the RAW murine macrophage cell line as a model, this study aimed to ascertain whether nutrient deprivation induces CD3 expression and to identify the signaling molecules involved in the activation of a CD3-dependent proinflammatory profile.

Firstly, the impact of environmental stress, specifically nutrient deprivation, on CD3 expression over a period was assessed. Subsequently, RAW cells were stimulated with anti-CD3 and IgG2a, and the profile of proinflammatory cytokines was evaluated alongside the expression of signaling proteins, including NFAT, c-Jun, and IKK. Furthermore, the transcriptional regulation of IRF-1, GATA-3, and MAFB was examined. Lastly, the functional capacity of CD3+ RAW cells was determined through a phagocytosis assay.

RAW cells were found to express molecules classically associated with T cells, including CD3, TCR, and CD4. Notably, CD3 expression was significantly upregulated at both the protein and transcriptional levels under nutrient deprivation, although it did not reach the levels observed in T cells. Functionally, CD3+ RAW cells exhibited enhanced phagocytic activity toward latex beads. Furthermore, stimulation with anti-CD3 plus IgG2a induced a robust proinflammatory cytokine response, characterized by the secretion of IFN-γ, TNF, and IL-6 as early as 5 hours post-stimulation. This response was associated with activation of signaling pathways involving NFAT, c-Jun, and IKK, along with increased IRF-1 expression and downregulation of MAFB, supporting the establishment of a CD3-dependent proinflammatory profile in RAW cells.

Our findings establish RAW macrophages as a model to study CD3+ signaling machinery in myeloid cells. We demonstrate for the first time that nutrient deprivation induces CD3 expression and that the resulting CD3-driven proinflammatory program is associated, at least in part, with NFAT and IRF-1. These conclusions provide new insights into the origin and functional significance of CD3+ macrophages and offer a valuable platform for investigating the impact of this pathway on innate immune responses across various pathological contexts.