ular carcinoma or to animal models that have less BChE than AChE activity. Using the rat as an animal model with low serum BChE activity, we have reported that AChE is significantly altered during liver cirrhosis. However, the Peretinoin biological activity contribution of AChE and BChE and their different R-AChE in Liver Cirrhosis molecular forms varies between rats and humans, and to date, there is no data on potential alterations of AChE expression in the cirrhotic human liver. Classical studies on cholinesterase 
usually focus on enzymatic activity and molecular forms of the enzyme. AChE occurs as both active and inactive subunits. Different AChE species are derived from alternative RNA splicing, generating different polypeptide transcripts with the same catalytic domain, but with distinct C-termini. It would be useful to perform further western blot analyses using different anti-AChE antibodies raised against different C-terminal peptides. In this study, we have measured AChE activity in non-diseased and cirrhotic human liver and plasma after BChE removal by immunoprecipitation. We have compared the different molecular forms and subunit banding pattern of AChE by SDS-PAGE under reducing conditions followed by Western blotting. AChE expression was assessed by quantitative RT-PCR analysis of the different AChE mRNAs, and the distribution of AChE protein was investigated in normal and cirrhotic human liver by immunohistochemistry. BChE resin ensured that the majority of the BChE activity in the samples was removed. Plasma and liver samples immunodepleted of BChE were then used for AChE determination. Enzyme assays and protein determination AChE and BChE activity were determined by a modified microassay method of Ellman. AChE was assayed with 1 mM acetylthiocholine and 50 mM tetraisopropyl pyrophosphoramide, a specific inhibitor of BChE; while BChE was measured with 1 mM butyrylthiocholine and 10 mM BW284c51, a specific inhibitor of AChE. One milliunit of AChE or BChE activity was defined as the number of nmoles of acetylthiocholine or butyrylthiocholine hydrolysed per min at 22uC. Protein concentrations were determined using the bicinchoninic acid method, with bovine serum albumin as standard. Sedimentation analysis Molecular forms of AChE and BChE were separated according to their sedimentation coefficients by centrifugation on 520% sucrose gradients containing 0.5% Triton X-100, as previously described. Approximately 40 fractions were collected from the bottom of each tube and assayed for cholinesterase activities. The cholinesterases present in liver and plasma are mainly tetramers and light globular species. Patients and Methods Patients For this study we obtained ethics approval from the ethics committee at our institutions and obtained written informed consent from all involved participants. The study was carried out in accordance with the Declaration of Helsinki. Plasma samples from patients with liver cirrhosis and agematched controls were provided by the Hospital General Universitario de Alicante, as previously described. Causes of cirrhosis were alcoholism, Hepatitis C virus infection and both alcoholism and HCV. Plasma was separated from whole blood by centrifugation, aliquoted and frozen at 280uC until use. Liver samples were obtained from the Hospital Clinic of Barcelona and collected as described in a previous study. Fragments of normal liver adjacent to colon cancer metastasis and liver specimens obtained immediately after laparotomy and before vascul
