Extracellular matrix modulates depot-specific adipogenic capacity in adipose tissue of dairy cattle

Adipose tissue (AT) expands through both hyperplasia and hypertrophy. During adipogenesis, adipose stromal and progenitor cells (ASPC) proliferate and then accumulate lipids, influenced by the local AT microenvironment. Increased adipogenic capacity is desirable as it relates to metabolic health, especially in transition dairy cows where excess free fatty acids in circulation can compromise metabolic and immune health. Our aim was to elucidate the depot-specific adipogenic capacity and extracellular matrix (EMX) properties of subcutaneous (SAT) and visceral (VAT) AT of dairy cows and define how the EMX affects adipogenesis. Flank SAT and omental VAT samples were collected from dairy cows in a local abattoir. Tissue samples were used for transcriptome analysis, targeted real-time quantitative PCR (RT-qPCR) for adipogenic markers, adipocyte sizing, assessment of viscoelastic properties and collagen accumulation, and then decellularized for native EMX isolation. For in vitro analyses, SAT and VAT samples were digested via collagenase, and ASPC cultured for metabolic analysis. Adipogenic capacity was assessed by adipocyte size, quantification of ASPC in stromal vascular fraction (SVF) via flow cytometry, and gene expression of adipogenic markers. In addition, functional assays including lipolysis and glucose uptake were performed to further characterize SAT and VAT adipocyte metabolic function. Data were analyzed using SAS (version 9.4; SAS Institute Inc., Cary, NC) and GraphPad Prism 9. Subcutaneous AT adipogenic capacity was greater than VAT's, as indicated by increased ASPC abundance, increased magnitude of adipocyte ADIPOQ and FASN expression during differentiation, and higher adipocyte lipid accumulation as shown by an increased proportion of larger adipocytes and abundance of lipid droplets. Rheologic analysis revealed that VAT is stiffer than SAT, which led us to hypothesize that differences between SAT and VAT adipogenic capacity were partly mediated by depot-specific EMX microenvironment. Thus, we studied depot-specific EMX-adipocyte crosstalk using a 3-dimensional model with native EMX (decellularized AT). Subcutaneous AT and VAT ASPC were cultured and differentiated into adipocytes within depot-matched and mismatched EMX for 14 d, followed by ADIPOQ expression analysis. Visceral AT EMX impaired ADIPOQ expression in SAT cells. Our results demonstrate that SAT is more adipogenic than VAT and suggest that divergences between SAT and VAT adipogenesis are partially mediated by the depot-specific EMX microenvironment.