Acetylation State of Histone Core Defines Macrophage Dynamics in Diabetic Wounds

Dysregulation of macrophage populations at the wound site is responsible for the non-healing state of chronic wounds. The molecular mechanisms underlying macrophage dysfunction and its control in diabetes are largely unexplored on an epigenetic level. Here, we report that acetyl histone-H3 (Lys27), an epigenetic mark regulating the macrophage transcriptome, is lost in the hostile tissue microenvironment in diabetes. The diabetic microenvironment, profoundly suppresses the acetylation of histone by activating HDACs-dependent deacetylation pathways. This, in consequence, suppress the STAT1 signaling in macrophages maintained in diabetic conditions. Interestingly, the HDAC inhibitor butyrate - via restoring the acetyl histone-H3 (Lys27)-dependent transcriptome - effectively rescues macrophage functions in a diabetic microenvironment. Butyrate reinstalls the STAT1 mediated transcription program and consequently macrophage activity depicting a unique fingerprint of tissue regeneration and inflammation control even in a hostile diabetic microenvironment. Most interesting, butyrate breaks the vicious cycle of inflammation in diabetic wounds. Our study offers novel pathogenic insight and the unique opportunity to reverse perturbed macrophage function thus holding promise to successfully treat diabetic and other chronic wounds and conditions of unrestrained inflammation. For RNA-Seq analysis, 1-5 µg of total RNA from total skin was first rRNA depleted using either Ribominus Eukaryotic system v2 kit (ThermoFisher Scientific). rRNA depleted RNA was subjected to preparation of Illumina compatible RNA-seq libraries using NEBNext Ultra II Directional RNA Library Preparation kit (NEB). RNA-seq libraries were quality controlled through Qiaxcel advanced system (Qiagen) and Bioanalyzer (Agilent) and finally measured by Qubit fluorimeter (ThermoFisher Scientific). Validated libraries were sequenced on Illumia NEXTSEQ 500 and NOVASEQ 6000 system. Single cell RNA sequencing was performed using Evercode WT v3 single cell whole transcriptome kit (Parse biosciences, USA), as per manufacturer instructions. In brief, single cell suspensions (~50,000 cells from each sample) from unwounded (Day 0) and post-wound (Day 5 and Day 10) mouse skin, were prepared and used for scRNAseq.