Essential Role of Extracellular Type V Collagen for Porcine Adipocyte Development

Abstract

The aim of this study was to examine the role of collagens in the differentiation of porcine subcutaneous preadipocytes (PSPA) using a specific post-transcriptional inhibitor of collagen synthesis, ethyl-3,4-dihydroxybenzoate (EDHB). Exponentially growing fibroblast-like PSPA cells were found to produce types I, III, IV, V and VI collagens by indirect immunofluorescence staining. When confluent preadipocyte cultures were stimulated with differentiation medium, various types of collagen matrices were assembled in the extracellular space of adipocytes. Since preadipocytes scarcely interconnect collagens despite the steady-state level expression of procollagen mRNAs, it appears that increasing the cell-associated form of collagens is an event unique to adipocytes. PSPA cells exposed to 40μM EDHB severely damaged the development of collagen fibrillar networks in adipocytes without affecting their transcription levels, and their triglyceride (TG) accumulation levels were reduced to 30% those of normally differentiated adipocytes. RT-PCR analysis also demonstrated a decrease in the expressions of some adipogenic genes including the master regulator of adipocyte gene transcription, PPARγ2, by EDHB-treated adipocytes, indicating the requirement for collagen during adipose conversion. The exogenous collagen types I, III, IV, V and VI were then individually supplied under EDHB conditions to determine whether or not they could make up for the lack of their own endogenous products. Of the five types of collagen, cells cultured on type V collagen were the only ones to improve the inhibitory effect of EDHB so as to recover up to 63.8% of TG content accompanied by the up-regulated expressions of PPARγ2 and adiponectin. These findings suggest that the extracellular signal from type V collagen is one of the major contributing factors in the differentiation pathway of porcine adipocytes.