Postcholecystectomy bile duct injuries (BDI) produce hepatic cholestasis and cause infection of the biliary tract. The biliary cells participate in secreting cytokines and in expression of immune response receptors. Toll-like receptors (TLRs) conduct signalling and activate the innate and adaptive inflammatory response. The objective was to determine the serum levels of TLR-2 and the expression of TLR-1 and TLR-2 and β-defensin in liver biopsies of postcholecystectomy BDI patients. A transverse, analytical study with 2 groups was done. One group included healthy volunteers (control group) and other included 25 postcholecystectomy BDI patients with complete biliary obstruction. Using the Enzyme-linked Immunosorbent Assay (ELISA) technique, serum levels of TLR-2 were determined, and with immunofluorescence the morphologic analysis of TLR-1 and TLR-2 and β-defensin in liver biopsies of postcholecystectomy BDI patients was performed. The average TLR-2 serum level in the control group was 0.0 pg/mL and in the BDI group, 0.023 ± 0.0045 pg/mL (P < 0.0001, bilateral Mann Whitney U). Immunofluorescence was used to determine the expression in liver biopsies, blood vessels, bile ducts, and hepatic parenchyma where 12 hepatic biopsies were positive for TLR-1 with average of 3213057.74 ± 1071019.25 μm(2); and 7 biopsies were positive for β-defensin with an average of 730364.33 ± 210838.02 μm(2); and 6 biopsies positive for TLR-2, obtaining an average of 3354364.24 ± 838591.06 μm(2). In conclusion, TLR-1 and TLR-2 and β-defensin play an important role in the innate antimicrobial defense of the hepatobiliary system.