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Generating Hypertrophic Adipocytes from 3T3 L1 Cell Line and Characterizing the Phenotypic Transition
Author(s) -
Yuan Xinxu,
Xu Xiaoyang,
Li Ningjun,
Dewey William L.,
Li PinLan,
Zhang Fan
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.701.7
Subject(s) - adipogenesis , 3t3 l1 , medicine , adipocyte , lipid droplet , endocrinology , muscle hypertrophy , cell culture , phenotype , biology , adipose tissue , microbiology and biotechnology , gene , biochemistry , genetics
3T3 L1 is a classical preadipocyte cell line that has been extensively used for the study of adipogenesis, a physiological process giving rise to the adipocytes. Adipocytic hypertrophy, a pathological enlargement of adipocytes, underscores the occurrences of obesity‐associated metabolic syndrome. Currently, it is lack of a reliable cell model to simulate the hypertrophic transition of adipocytes. This study aims to develop adipocytic hypertrophy from 3T3 L1 and characterize the related phenotypic transition by examining the genes key to the pathophysiology of adipocytes as well as their functional outcomes. To generate hypertrophic adipocytes, we grew the post‐differentiated 3T3 L1 cells in DMEM/F12 medium (the starting date designated as day 0), supplemented with 10% FBS and 0.1% insulin, to their maturation approximately at day 10; and then continued loading lipids to the matured adipocytes by culturing these cells in the same media. The hypertrophic phenotype appeared around day 24. Morphologically, the further enlargement of lipid droplets during the hypertrophy was confirmed by Bodipy‐stained fluorescence images. Quantitative RT‐PCR analysis showed the expression of PPARγ, an adipogenesis‐promoting nuclear factor, and adiponectin, an adipogenetic indicator, were first progressively increased from day 0 to reach their maximum at the day 10, and then regressively declined to day 32, an end point in experimental observations. Consistent with the decline in adipogenesis, the mRNA levels of pro‐inflammatory factor MCP‐1 were markedly elevated in the hypertrophic adipocytes after day 24, as well, their MCP‐1‐containing culture media displayed significant macrophage‐attracting activity in cell migration assays. But the expression and activity of cytochrome C oxidase, a critical enzyme for energy generation, were reduced during the hypertrophy. Functionally, the viability of adipocytes, as determined by colorimetrically reading the formation of formazan from the metabolized tetrazolium, was gradually decreased after day 16. In situ TUNEL (Terminal Deoxynucleotidyl Transferase dUTP Nick End Labeling) analysis showed the adipocyte apoptosis was significantly increased during the hypertrophy. These results for the first time demonstrate that hypertrophic adipocytes can be generated from 3T3 L1 cell line. And this hypertrophic transition may be able to serve as a model to develop adipocytic hypertrophy‐targeting remedy in the treatment of obesity‐associated metabolic syndrome. Support or Funding Information Supported by NIH R01HL115068