Generation of mature fat pads in vitro and in vivo utilizing 3-D long-term culture of 3T3-L1 preadipocytes

Exp Cell Res. 2004 Oct 15;300(1):54-64. doi: 10.1016/j.yexcr.2004.05.036.

Abstract

Tissue-inherent factors such as cell-cell and cell-extracellular matrix interactions are regarded to exert a potentially large impact on adipogenesis as well as on secretory functions of adipose tissue. However, an appropriate 3-D adipogenesis model useful for addressing such interactions is still lacking. In this study, using tissue-engineering techniques, we demonstrate for the first time the development of coherent fat pads consisting of unilocular signet-ring cells in vitro. The constructs were generated by differentiating 3T3-L1 preadipocytes on 3-D polymeric scaffolds for either 9, 21, or 35 days in vitro. Only long-term culture yielded uniform tissues histologically comparable to native fat. Light and scanning electron microscopy provided direct evidence of 3-D tissue coherence and cell-cell contact in a tissue context, which was in strong contrast to conventional 2-D monolayer culture. Further differences between the two culture systems included enhanced secretion of leptin in 3-D tissue culture and differences in laminin expression (mRNA and protein level). Increase of triglyceride content over culture time and mRNA expression of other adipocyte genes, such as PPARgamma and Glut-4, were found to be similar. Implantation of long-term differentiated tissue constructs in nude mice resulted in further development and maintenance of fat pads. The presented model system is suggested to contribute to a better understanding of adipose tissue development and function facilitating studies on tissue-inherent interactions in vitro and in vivo.

MeSH terms

  • 3T3-L1 Cells
  • Adipocytes / metabolism*
  • Adipocytes / ultrastructure
  • Adipose Tissue / metabolism*
  • Adipose Tissue / ultrastructure
  • Animals
  • Cell Adhesion / physiology
  • Cell Communication / physiology*
  • Cell Culture Techniques / methods
  • Extracellular Matrix / metabolism
  • Glucose Transporter Type 4
  • Laminin / metabolism
  • Leptin / metabolism
  • Mice
  • Mice, Nude
  • Microscopy, Electron, Scanning
  • Monosaccharide Transport Proteins / genetics
  • Muscle Proteins / genetics
  • Organ Culture Techniques / methods
  • PPAR gamma / genetics
  • RNA, Messenger / metabolism
  • Time Factors
  • Tissue Engineering / methods*
  • Triglycerides / metabolism
  • Up-Regulation / physiology

Substances

  • Glucose Transporter Type 4
  • Laminin
  • Leptin
  • Monosaccharide Transport Proteins
  • Muscle Proteins
  • PPAR gamma
  • RNA, Messenger
  • Slc2a4 protein, mouse
  • Triglycerides