Ultrastructural immunogold localization of subcellular sites of TNF‐α in colonic Crohn's disease

Tumor necrosis factor‐α, a proinflammatory cytokine, might have an important role(s) in initiating, modifying, and/or sustaining chronic inflammatory processes such as those that characterize Crohn's disease, an inflammatory bowel disease of unknown etiology. We used an immunogold ultrastructural morphometric approach to localize tumor necrosis factor‐α in colonic Crohn's disease biopsies. Tumor necrosis factor‐α was present in seven cell types (fibroblasts, eosinophils, mast cells, macrophages, colonic epithelial absorptive cells, Paneth cells, neutrophils). Tumor necrosis factor‐α‐containing subcellular organelles included lipid bodies (fibroblasts, eosinophils, macrophages, mast cells, colonic epithelial cells, neutrophils), secretory granules (eosinophils, Paneth cells), phagolysosomes (macrophages, colonic epithelial cells), and Golgi structures and vesicle membranes (neutrophils). A gradient of extracellular tumor necrosis factor‐α immunoreactivity surrounded eosinophils, mast cells, and macrophages. P values of gold counts/μm 2 were significant for all cells, organelles, and extracellular spaces measured, and all positive structures significantly exceeded the background labeling density/μm 2 . Specificity controls (normal rabbit serum, tumor necrosis factor‐α‐absorbed primary antibody) either failed to label these sites or gave markedly reduced specific tumor necrosis factor‐α labeling, respectively. These findings represent the first ultrastructural localization of the subcellular sites of TNF‐α in vivo in seven cell lineages in human colonic tissues.