Background <p>Sepsis is characterized by dysregulated immunologic reactions to infectious insults, representing a main cause of death for seriously ill patients. Studies have demonstrated correlations between rapidly declined high-density lipoprotein (HDL) levels during initial sepsis phases with increased mortality and adverse clinical outcomes. Furthermore, HDL subcomponents, specifically HDL2b and HDL3, undergo significant alterations in composition in sepsis patients. This article was aimed at investigating the effects of alterations in the ratio of HDL2b to HDL3 (HDL2b/3) on the function of human umbilical vein endothelial cells (HUVECs) and monocyte-macrophages (THP-1 and RAW264.7 cells).</p> Methods <p>Plasma samples were collected from 10 sepsis patients and 10 healthy controls. High-density lipoprotein (HDL) subclasses (HDL2b and HDL3) were separated and quantified using fast protein liquid chromatography (FPLC) and ELISA. Human monocytic leukemia cells (THP-1) and human umbilical vein endothelial cells (HUVECs) were co-incubated with different reconstituted ratios of HDL2b/3 under lipopolysaccharide (LPS) stimulation to evaluate cell viability, inflammatory cytokine expression, angiogenesis, and cell migration.</p> Results <p>Our results showcase evidently reduced HDL2b/3 ratios for the sepsis group compared with their non-sepsis counterparts. Notably, HDL2b/3 from both groups attenuated the gene expression of lipopolysaccharide-stimulated inflammatory factors (interleukin-1β and tumor necrosis factor-α) within THP-1 cells, RAW264.7 cells, and HUVECs. Additionally, HDL2b/3 mitigated the detrimental effects of LPS on THP-1 and RAW264.7 cells.</p> Conclusion <p>This research highlights that altered HDL subcomponent ratios can potentially modulate inflammation and cellular responses during sepsis.</p>