Activation of macrophages by endotoxin is assumed responsible for increased circulating tumor necrosis factor α (TNF-α) and soluble TNF receptor (sTNFR) levels in cirrhosis. Relevant to this is expression of Toll-like receptor (TLR) 4 and TLR2, which is critically involved in production of TNF-α in response to endotoxin and Gram-positive microbial stimuli, respectively. The first studies on this in cirrhosis are reported here. In 36 cirrhotic patients and 32 controls, we measured (1) circulating endotoxin, TNF-α, and sTNFR levels; (2) peripheral blood mononuclear cell (PBMC) expression of TLR4 and TLR2, and (3) in vitro TNF-α production by PBMCs stimulated with endotoxin or Staphylococcus aureus enterotoxin B (SEB). PBMC expression of TLR2, circulating TNF-α levels, and in vitro TNF-α production were reassessed after supplementation with a synbiotic regimen known to increase intestinal levels of Gram-positive bacteria. Endotoxin, TNF-α, and sTNFR levels were significantly increased in cirrhosis. Endotoxin levels did not correlate significantly with other parameters. PBMC expression of TLR2 but not TLR4 was significantly up-regulated in cirrhosis and correlated significantly with serum TNF-α and sTNFR levels. In vitro TNF-α production by PBMCs stimulated by SEB was significantly blunted. Supplementation with the synbiotic regimen resulted in significant up-regulation of PBMC expression of TLR2. Serum TNF-α levels were further increased and in vitro TNF-α production further reduced in most patients. In conclusion, up-regulation of PBMC expression of TLR2 but not TLR4 occurs in cirrhosis, which implies, contrary to previous assumptions, an important stimulatory role for Gram-positive microbial components but not endotoxin. TLR2 likely contributes to increased circulating TNF-α and sTNFR levels in cirrhosis. H epatology 2003;37:1154-1164.)