Nuclear and chromatin rearrangement associate to epigenome and gene expression changes in a model of in vitro adipogenesis and hypertrophy

Abstract

Hypertrophy of adipocytes represents the main cause of obesity. We investigated in vitro the changes associated with adipocyte differentiation and hypertrophy focusing on the nuclear morphometry and chromatin epigenetic remodelling. The 3 T3-L1 pre-adipocytes were firstly differentiated into mature adipocytes, then cultured with long-chain fatty acids to induce hypertrophy. Confocal and super-resolution stimulation emission depletion (STED) microscopy combined with ELISA assays allowed us to explore nuclear architecture, chromatin distribution and epigenetic modifications. In each condition, we quantified the triglyceride accumulation, the mRNA expression of adipogenesis and dysfunction markers, the release of five pro-inflammatory cytokines. Confocal microscopy revealed larger volume and less elongated shape of the nuclei in both mature and hypertrophic cells respect to pre-adipocytes, and a trend toward reduced chromatin compaction. Compared to mature adipocytes, the hypertrophic phenotype showed larger triglyceride content, increased PPARγ expression reduced IL-1a release, and up-regulation of a pool of genes markers for adipose tissue dysfunction. Moreover, a remodelling of both epigenome and chromatin organization was observed in hypertrophic adipocytes, with an increase in the average fluorescence of H3K9 acetylated domains in parallel with the increase in KAT2A expression, and a global hypomethylation of DNA. These findings making light on the nuclear changes during adipocyte differentiation and hypertrophy might help the strategies for treating obesity and metabolic complications.