Combined Exposure to High-Cholesterol Diet and PM2.5: Brain Injury, Behavioral Alteration and Regulatory Mechanism of HIF-1α in Female Mice.

Both high-cholesterol diet (HCD) and fine particulate matter (PM2.5) exposure are associated with increased stroke risk; however, their combined effects-particularly in females-remain poorly understood. This study aimed to investigate the synergistic effects of HCD and PM2.5 exposure on brain injury, behavior, and underlying molecular mechanisms in female mice. Female mice were exposed to HCD, PM2.5 inhalation, or both for 3 months (3M) or 6 months (6M), with control groups for comparison. We assessed brain-to-body weight ratio, behavioral performance, histopathological changes, inflammatory markers (IL-6, TNF-α), and expression of stroke- and blood-brain barrier (BBB)-related proteins. Additionally, we analyzed the epigenetic regulation of hypoxia-inducible factor-1α (HIF-1α) via chromatin immunoprecipitation (ChIP) assays. Co-exposure to HCD and PM2.5 led to greater alterations in the brain-to-body weight ratio, exacerbated neuropathology, behavioral deficits, and increased neuroinflammation compared to exposure to either factor alone. Moreover, HCD+PM2.5 significantly altered the expression of HIF-1α and genes involved in its downstream signaling pathway in the brain. ChIP assays revealed that HIF-1α was directly regulated by histone deacetylase 4 (HDAC4) and histone H3 lysine 9 acetylation (H3K9ac), indicating epigenetic modulation in response to dual exposure. HCD and PM2.5 synergistically promote brain injury in female mice, potentially through HDAC4/H3K9ac-mediated epigenetic activation of the HIF-1α pathway. These findings highlight a novel molecular mechanism that may contribute to increased stroke risk in females exposed to both environmental and metabolic stressors.