Abstract Body: Background: Isolevuglandins (isoLGs) are products of lipid peroxidation that are formed in the presence of reactive oxygen species. IsoLGs are increased in circulating CD14⁺ monocytes in patients with hypertension and contribute to inflammatory activation. When adducted to histones, isoLGs prevent nucleosome assembly. Moreover, isoLGs induce disassembly of pre-formed nucleosomes. IsoLGs can be induced in cell culture with tert-butyl hydroperoxide (tBHP) and are efficiently scavenged by ethyl-2-hydroxybenzylamine (Et-2HOBA). We hypothesized that isoLGs regulate chromatin accessibility in hypertensive (HTN) monocytes.

Methods: Human CD14⁺ monocytes were isolated from 3 normotensive (NT) and 5 HTN subjects. HTN monocytes were treated with vehicle. NT monocytes were treated with vehicle, tBHP, or co-treated with tBHP and Et-2HOBA. Cells were cultured for 24 hours and submitted for bulk ATAC sequencing to assess regions of open chromatin across the genome. Data were analyzed using Partek Flow software. Reads were aligned to the GRCh38 human genome. Differences in chromatin accessibility were determined by DESeq2 analysis and were considered significant at an adjusted P-value of 0.05 and fold change of 1.5. Reads were filtered to focus on accessible chromatin at transcription start sites (TSS). First, chromatin accessibility of HTN monocytes was compared to NT monocytes. Second, to determine chromatin regions with isoLG-regulated accessibility that are associated with hypertension, we compared control and tBHP+Et2HOBA treated cells to a combined HTN and tBHP treated group.

Results: HTN monocytes exhibited increased chromatin accessibility at 131 TSS and decreased accessibility at 56 TSS compared to NT monocytes. Chromatin accessibility was increased at 205 TSS and decreased at 183 TSS in response to both hypertension and isoLGs. Gene set enrichment revealed a significant increase in chromatin accessibility in genes regulating chemokine signaling, RAS signaling, lysosomal activity, and focal adhesion.

Conclusions: We determined that significant differences in chromatin accessibility are present in HTN monocytes compared to NT monocytes. IsoLGs regulate chromatin accessibility in HTN monocytes. Moreover, isoLGs regulate accessibility of the TSS of genes involved in inflammation and cell migration. These findings implicate isoLGs as novel post-translational histone modifications that regulate chromatin accessibility and contribute to inflammation in hypertension.