AGPAT2 deficiency impairs adipogenic differentiation in primary cultured preadipocytes in a non-autophagy or apoptosis dependent mechanism

M Fernández-Galilea, P Tapia, K Cautivo… - Biochemical and …, 2015 - Elsevier
M Fernández-Galilea, P Tapia, K Cautivo, E Morselli, VA Cortés
Biochemical and biophysical research communications, 2015Elsevier
Abstract Aims Mutations in 1-acylglycerol-3-phosphate O-acyltransferase 2 (AGPAT2) result
in lipodystrophy, insulin resistance and diabetes. Autophagy is required for normal
adipogenesis and adipose tissue development. The aim of this study was to determine
whether impaired autophagy or excessive cell death underlie the adipogenic inability of
Agpat2−/− mice preadipocytes. Methods Preadipocytes were isolated from interscapular
brown adipose tissue (BAT) of Agpat2−/− and Agpat2+/+ newborn mice and …
Aims
Mutations in 1-acylglycerol-3-phosphate O-acyltransferase 2 (AGPAT2) result in lipodystrophy, insulin resistance and diabetes. Autophagy is required for normal adipogenesis and adipose tissue development. The aim of this study was to determine whether impaired autophagy or excessive cell death underlie the adipogenic inability of Agpat2−/− mice preadipocytes.
Methods
Preadipocytes were isolated from interscapular brown adipose tissue (BAT) of Agpat2−/− and Agpat2+/+ newborn mice and cultured/differentiated in vitro. Intracellular lipids were quantified by oil red O staining. Cell death was assessed by lactate dehydrogenase (LDH) activity. Apoptosis and autophagy regulatory factors were determined at the mRNA and protein level with Real-time PCR, immunoblot and immunofluorescence.
Results
Adipogenically induced Agpat2−/− preadipocytes had fewer lipid-loaded cells and lower levels of adipocyte markers than wild type preadipocytes. Before adipogenic differentiation, autophagy-related proteins (ATGs) ATG3, ATG5–ATG12 complex, ATG7 and LC3II were increased but autophagic flux was reduced, as suggested by increased p62 levels, in Agpat2−/− preadipocytes. Adipogenic induction increased LDH levels in the culture media in Agpat2−/− preadipocytes but no differences were observed in the activation of Caspase 3 or in markers of autophagic flux.
Conclusions
AGPAT2 is required for in vitro adipogenesis of mouse preadipocytes. Autophagy defects or apoptosis are not involved in the adipogenic failure of Agpat2−/− preadipocytes.
Elsevier