Interfering PTBP1 expression inducing M2 macrophage polarization via miR-124/MAPK pathway

Background Modulating the polarization phenotype of macrophages from M1 to M2 type appears to offer significant potential for treating CNS diseases, such as spinal cord injury and stroke. Polypyrimidine tract-binding protein 1 (PTBP1) is an indispensable RNA-binding protein that participates in nearly all stages of RNA maturation, with particular importance in pre-mRNA splicing. However, the role of PTBP1 in macrophage polarization remains much less explored. In this study, we investigated the impact of PTBP1 on macrophage polarization and unveiled its potential molecular mechanism. Methods Firstly, we employed immunostaining techniques to evaluate the expression of PTBP1 in macrophages infiltrating the spinal cord injury area and cerebral ischemic zone. RAW264.7 cells were induced to M1 macrophages by lipopolysaccharide (LPS). Immunofluorescence staining and western blotting were used to detect the expression and localization of PTBP1 before and after induction. Additionally, PTBP1siRNA was utilized in LPS-induced RAW264.7 cells to specifically knock down PTBP1 expression. We subsequently examined the impact of PTBP1 inhibition on the phenotypic and metabolic alterations in macrophage polarization. Furthermore, RNA-seq analysis was employed to screen differentially expressed genes in LPS-induced RAW264.7 cells before and after PTBP1 inhibition, enabling us to identify significant signaling pathways related to PTBP1. Finally, qRT-PCR and western blot were conducted to validate the molecular mechanism by which PTBP1 regulates the polarization of M1/M2 macrophages. Results PTBP1 was highly expressed in M1 macrophages infiltrating the spinal cord injury area and cerebral ischemic zone. In LPS-induced RAW264.7 cells, PTBP1 tended to be located in the nucleus. Inhibiting PTBP1 resulted in downregulation of M1-like genes (inducible nitric oxide synthase [iNOS, tumor necrosis factor [TNF]-α, and interleukin [IL]-6) and upregulation of M2-like genes (Arginase [ARG]-1, IL-10, and tumor necrosis factor [TGF]-β1). In addition, there was a shift in macrophage metabolism from M1-type glycolysis to M2-type aerobic oxidation. PTBP1 inhibition enhanced the expression of mature miR-124, thereby suppressing the activation of the downstream MAPK pathway. Conclusion PTBP1 shifts the polarization phenotype of macrophages via regulating microRNA-124 (miR-124)/mitogen-activated protein kinases (MAPK) signaling pathway and glucose metabolism. PTBP1 may be a novel therapeutic target for macrophages polarization related-CNS injuries.