Dynamics of protein secretion during adipocyte differentiation.

Koichi Ojima,Susumu Muroya,Ikuyo Nakajima,Takanori Nishimura,Mika Oe

doi:10.1002/2211-5463.12091

Koichi Ojima, Susumu Muroya + Show 3 more

Open Access

https://doi.org/10.1002/2211-5463.12091

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Abstract

The major functions of adipocytes include both lipid storage and the production of secretory factors. However, the type of proteins released from mouse 3T3‐L1 cells during adipocyte differentiation remains poorly understood. We examined the dynamics of secreted proteins during adipocyte differentiation using mass spectrometry (MS) combined with an iTRAQ ® labeling method that enables the simultaneous analysis of relative protein expression levels. A total of 215 proteins were identified and quantified from approximately 10 000 MS/MS spectra. Of these, approximately 38% were categorized as secreted proteins based on gene ontology classification. Adipokine secretion levels were increased with the progression of differentiation. By contrast, levels of fibril collagen components, such as subunits of type I and III collagens, were decreased during differentiation. Basement membrane components attained their peak levels at day 4 when small lipid droplets accumulated in differentiated 3T3‐L1 cells. Simultaneously, peak levels of collagen microfibril components that comprise type V and VI collagen subunits were also observed. Our data demonstrated that extracellular matrix components were predominantly released during the early and middle stages of adipocyte differentiation, with a subsequent increase in the secretion of adipokines. This suggests that 3T3‐L1 cells secrete adipokines after their ECM is constructed during adipocyte differentiation.

Highlights

The major functions of adipocytes include both lipid storage and the production of secretory factors
Each adipocyte is surrounded by the basement membrane that includes type IV collagen, laminin, and Abbreviations CM, conditioned medium; DM, differentiation medium; DMEM, Dulbecco’s Modified Eagle Medium; ECM, extracellular matrix; FBS, fetal bovine serum; GM, growth medium; iTRAQâ, isobaric tags for relative and absolute quantitation; ITSTM premix media (ITS), insulin, transferrin, and selenous acid; MS, mass spectrometry
DM1 or DM2 were shifted to ITSTM premix media (ITS) that contained insulin, transferrin, and sodium selenite in DMEM, and cells were subsequently cultured for 2 days

Summary

Introduction

The major functions of adipocytes include both lipid storage and the production of secretory factors. Our data demonstrated that extracellular matrix components were predominantly released during the early and middle stages of adipocyte differentiation, with a subsequent increase in the secretion of adipokines. Each adipocyte is surrounded by the basement membrane that includes type IV collagen, laminin, and Abbreviations CM, conditioned medium; DM, differentiation medium; DMEM, Dulbecco’s Modified Eagle Medium; ECM, extracellular matrix; FBS, fetal bovine serum; GM, growth medium; iTRAQâ, isobaric tags for relative and absolute quantitation; ITS, insulin, transferrin, and selenous acid; MS, mass spectrometry. Collagen type V and type VI are secreted from cultured adipocytes to facilitate triglyceride accumulation during differentiation in vitro [9] These studies show that adipocytes produce various types of ECM components as releasing factors to provide mechanical support to adipocytes and to regulate adipogenesis

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