title: Tight association between macrophages and adipocytes in obesity: implications for adipocyte preparation.
abstract: OBJECTIVE
  To determine the cellular architecture of the inflammatory infiltrate in adipose tissue from obese mice, and identify the source of inflammatory cytokines in adipose tissue at a single cell level.
  
  
  METHODS
  Adipose tissue from diet-induced obese mice was digested by collagenase treatment and fractionated by density centrifugation to obtain an adipocyte floating layer and a pellet of stromal vascular cells. The cellular architecture of the adipocyte-macrophage interaction in both intact white adipose tissue (WAT) and the separated density gradient floating layer fraction was analyzed by confocal immunohistochemistry. Cytokine expression was detected by semi-quantitative real time PCR and immunohistochemical analysis.
  
  
  RESULTS
  Three dimensional image analysis of WAT and the separated "adipocyte" floating layer revealed lipid-engorged macrophages, macrophages in contact with lipid droplets and sheath-like assemblies of macrophages surrounding adipocytes. The macrophages immunostained for TNFα and to a lesser extent for the immunoregulatory cytokine IL-10. TNFα staining was associated only with macrophages indicating that macrophages and not adipocytes are the source of TNFα expression in the adipocyte floating layer.
  
  
  CONCLUSIONS
  Macrophages form assemblies that tightly adhere to and cover adipocytes and lipid droplets. TNFα found in low density adipocyte preparations is due to contamination with macrophages.
authors.affiliation: Department of Medicinal and Biological Chemistry, College of Pharmacy and Pharmaceutical Sciences, University of Toledo, Toledo, Ohio, USA; Cole Eye Institute, Cleveland Clinic, Cleveland, Ohio, USA.
authors.forename: Lindsey A
authors.initials: LA
authors.lastname: Ebke
authors.forename: Andrea L
authors.initials: AL
authors.lastname: Nestor-Kalinoski
authors.forename: Brandon D
authors.initials: BD
authors.lastname: Slotterbeck
authors.forename: Ali G
authors.initials: AG
authors.lastname: Al-Dieri
authors.forename: Sumona
authors.initials: S
authors.lastname: Ghosh-Lester
authors.forename: Lucia
authors.initials: L
authors.lastname: Russo
authors.forename: Sonia M
authors.initials: SM
authors.lastname: Najjar
authors.forename: Hermann
authors.initials: H
authors.lastname: von Grafenstein
authors.forename: Marcia F
authors.initials: MF
authors.lastname: McInerney
mesh_terms: D017667:Adipocytes / Q000166:cytology / Q000648:ultrastructure*; D052436:Adipose Tissue, White / Q000166:cytology*; D000818:Animals; D002469:Cell Separation; D007249:Inflammation; D016753:Interleukin-10 / Q000378:metabolism; D008264:Macrophages / Q000166:cytology / Q000648:ultrastructure*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008820:Mice, Obese; D018613:Microscopy, Confocal; D009765:Obesity; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism
chemical_list: D014409:Tumor Necrosis Factor-alpha; D016753:Interleukin-10