Isolation method and characterization of adipocytes as a tool for equine obesity research - In vitro study.

Abstract

Adipose tissue functions as an endocrine organ; however, excessive lipid accumulation can lead to obesity and metabolic disorders, such as Equine Metabolic Syndrome (EMS), characterized by insulin resistance, fat deposition, and increased inflammation. Despite the growing prevalence of obesity in horses, knowledge of equine adipocytes and their metabolic functions remains limited. The main objective of the study was to develop and optimize a method for isolating equine adipocytes and to characterize their metabolic activity. Using slaughterhouse-derived horse visceral adipose tissue, we developed a protocol to isolate mature adipocytes. Metabolic activity of cells was assessed by examining their sensitivity to lipolytic factors: isoproterenol (0.001-10&#x202f;&#xb5;M), epinephrine (0.001-1&#x202f;&#xb5;M), and forskolin (0.001-1&#x202f;&#xb5;M)-and lipogenesis intensity after stimulation with insulin. We obtained mature equine adipocytes with diameters ranging from 50 to 160&#x202f;&#xb5;m. These cells demonstrated full metabolic functionality, responding to lipolytic factors such as isoproterenol (all doses: p&#x202f;<&#x202f;0.001), epinephrine (0.01&#x202f;&#xb5;M: p&#x202f;<&#x202f;0.05; 0.1-1&#x202f;&#xb5;M: p&#x202f;<&#x202f;0.0001), and forskolin (0.001&#x202f;&#xb5;M: p&#x202f;<&#x202f;0.0001). The adipocytes also responded to insulin from all tested species, with effects being dose- and time-dependent (after 2&#x202f;h human insulin 10&#x202f;nM, p&#x202f;<&#x202f;0.05; bovine 10, 100&#x202f;nM p&#x202f;<&#x202f;0.05 and after 8&#x202f;h all doses p&#x202f;<&#x202f;0.05). The presented method for isolating mature equine adipocytes is effective, yielding metabolically functional cells, which can serve as a valuable in vitro model for studying the effects of various factors on adipocyte function, contributing to a better understanding of equine adipose tissue dysfunction, particularly in the context of metabolic disorders.