LPS RECEPTOR CD14 PARTICIPATES IN RELEASE OF TNF-ALPHA IN RAW-264.7 AND PERITONEAL-CELLS BUT NOT IN KUPFFER CELLS

Lipopolysaccharide (LPS) is a bacterial polymer that stimulates macrop hages to release tumor necrosis factor-alpha (TNF-alpha). In macrophag es (RAW 264.7 and peritoneal cells), LPS binds to the CD14 surface rec eptor as the first step toward signaling. Liver macrophages, Kupffer c ells, are the most numerous fixed-tissue macrophage in the body. The p resence of CD14 on Kupffer cells and its role in LPS stimulation of TN F-alpha were examined. TNF-alpha release by Kupffer cells after LPS st imulation was the same in the presence and absence of serum. RAW 264.7 and peritoneal cells, which utilize the CD14 receptor, released signi ficantly less TNF-alpha after LPS stimulation in the absence of serum because of the absence of LPS-binding protein. Phosphatidyl-inositol-p hospholipase C treatment, which cleaves the CD14 receptor, decreased L PS-stimulated TNF-alpha release by RAW 264.7 cells but not by Kupffer cells. Deacylated LPS (dLPS) competes with LPS at the CD14 receptor wh en incubated in a ratio of 100:1 (dLPS/LPS). Such competition blocked LPS-stimulated TNF-alpha release from RAW 264.7 cells but not from Kup ffer cells. Western and fluorescence-activated cell sorter analysis di rectly demonstrated the presence of CD14 on RAW 264.7 cells and murine peritoneal cells but showed only minimal amounts of CD14 in murine Ku pffer cells. LPS stimulation did not increase the amount of CD14 detec table on mouse Kupffer cells. CD14 expression is very low in Kupffer c ells, and LPS-stimulated TNF-alpha release is independent of CD14 in t hese cells.