Chronotherapy involving rosiglitazone regulates the phenotypic switch of vascular smooth muscle cells by shifting the phase of TNF-α rhythm through triglyceride accumulation in macrophages

Objectives: Vascular diseases are often caused by the interaction between macrophages and vascular smooth muscle cells (VSMCs). This study aims to elucidate whether chronotherapy with rosiglitazone (RSG) can regulate the secretion rhythm of macrophages, thereby controlling the phenotypic switch of VSMCs and clarifying the potential molecular mechanisms, providing a chronotherapeutic approach for the treatment of vascular diseases. Methods: RAW264.7 cells and A7r5 cells were synchronized via a 50 % FBS treatment. M1-type macrophages were induced through Lipopolysaccharide (LPS) exposure. Additionally, siRNA and plasmids targeting PPAR gamma were transfected into macrophages. The assessment encompassed cell viability, migration, inflammatory factor levels, lipid metabolites, clock gene expression, and relative protein expression. Results: We revealed that, in alignment with core clock genes Bmal1 and CLOCK, RSG administration at ZT2 advanced the phase of TNF-alpha release rhythm, while ZT12 administration shifted it backward. Incubation with TNF-alpha at ZT2 significantly promoted the phenotype switch of VSMCs. This effect diminished when incubated at ZT12, implicating the involvement of the clock-MAPK pathway in VSMCs. Furthermore, RSG administration at ZT2 advanced the phases of PPAR gamma and Bmal1 genes, whereas ZT12 administration shifted them backward. Additionally, PPAR gamma overexpression significantly induced triglyceride (TG) accumulation in macrophages. Exogenous TG upregulated Bmal1 and CLOCK gene expression in macrophages and significantly increased TNF-alpha release. Conclusion: Chronotherapy involving RSG induces TG accumulation within macrophages, resulting in alterations in circadian gene rhythms. These changes, in turn, modulate the phase of rhythmic TNF-alpha release and play a regulatory role in VSMCs phenotype switch. Our study establishes a theoretical foundation for chronotherapy of PPAR gamma agonists.