Genesis of epicardial adipocytes and its association with progenitor markers, muscarinic receptor type 3 and b-blockers intake in patients with cardiovascular disease

Abstract

Abstract Introduction Epicardial fat thickness or volume was found to be associated with cardiovascular disease (CVD). Our aim was to study the epicardial adipocyte-progenitors' markers and its association with cholinergic or adrenergic activity in patients with cardiovascular disease. Materials and methods We have included epicardial adipose tissue (EAT) biopsies from 29 patients underwent open-heart surgery. From 10 patients (69±5 years old, 31±8 kg/ m2, 40% CAD, 40% HF, 60% AF, 0% T2DM) stromal cells from epicardial and subcutaneous fat were isolated after collagenase activity and cultured for 14 days and then submitted to adipogenesis for next 14 days. Samples from 19 patients (60±9 years old, 29±4 kg/m2, 42% CAD, 37% HF, 32% AF, 32% T2DM, 53% β-blockers) were used for “ex vivo assays”. Explants were split into equal pieces (100 mg), treated with or without acetylcholine (ACh) for 30 min. Afterwards RNA was isolated and cDNA was amplified by real time PCR. We selected adipocytes progenitors (CD36, PREF1, COL1A1), adipocytes markers (ADIPO, FABP4), muscarinic (muscarinic receptor type 2 (CHRM2) and 3 (CHRM2)) and β-adrenergic receptors (ABRD1, ABRD2 and ABRD3). Gene expression was represented regarding ACTB as 2HK/GEN. Results The stromal vascular cells (SVC) from subcutaneous fat (SAT) had higher expression levels of CD36, PREF1 and COL1A1 than SVC from epicardial fat (EAT). It explains the higher adipocytes markers after adipogenesis induction in SAT than EAT cells. However, an upregulation of fibroblasts markers was detected on EAT. The levels of CD36 and PREF1 in SVC were associated with higher adipogenesis. Although CHRM2 was higher in EAT than SAT SVC, the adipogenesis induction upregulated only CHRM3 (1.48±0.065 vs 1.42±0.036 a.u.) in EAT cells. Thus, this receptor was associated with adipocytes markers in epicardial fat (r=0.777 for CD36 and r=0.746 for FABP4) and incremented in epicardial fat biopsies from patients who were taken β-blockers (1.61±0.011 n=10 vs 1.54±0.097 a.u. n=9; p=0.05) and modulated by ACh treatment (p=0.05). Conclusions Our results showed that CD36 and PREF1 in epicardial SVC are adipocytes progenitors. The higher presence of adipocytes markers is associated with higher levels of muscarinic receptor (CHRM3), which are upregulated in epicardial fat from patients who were taken β-blockers and modulated by cholinergic activity. Because a metabolic and lipolytic dysfunction was associated with CHRM3, the sympathetic modulation might play a role in the epicardial adipocytes genesis. Further studies are needed to understand if this mechanism might improve or not future cardiovascular events. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): ISCIII (PFIS2020)