0145-6008/96/2009-1564$03.00/0 zyxwvutsrqpo ALCOHOLISM: CLINICAL AND EXPERIMENTAL RESEARCH Vol. 20, No. 9 December 1996 Distribution of Peripheral Blood Lymphoid Subsets in Alcoholic Liver Cirrhosis: Influence of Ethanol Intake z Francisco Javier Laso, zyxwvuts Jose lgnacio Madruga, Antonio Lopez, Juana Ciudad, Melchor Alvarez-Mon, Jesus San Miguel, and Albert0 Orfao The aim of the present study was to investigatethe effect of chronic ethanol (EtOH) consumption on the immune system in patients with alcoholic liver cirrhosis (ALC), as analyzed by the distribution of pe- ripheral blood (PB-) T, B, and NK lymphoid subsets using multiple stainings with monoclonal antibodies and flow cytometry. For that purpose, we have analyzed a group of patients with ALC and active EtOH intake (ALCET group) which were re-evaluated3 months after alcohol withdrawal. As controls, both ALC patients with at least 1 year of alcoholwithdrawal (ALCAW group) and healthysubjectswere used. Regarding the alcohol intake period, the most relevantfindings were a significant activation of the PB T-cell compartment, and spe- cifically of the TCR alpha beta+ subset, as reflected by an increased expression of both the HLA DR and C D l l c antigens as well as a significant increase of both the PB NK cells (CD3-/CD56+) and the cytotoxic T cells coexpressing the CD3 and CD56 molecules. In ad- dition, a decrease of both the numbers of total B cells and their CD5+/CD19+ subset were observed. After a relatively short with- drawal period (3 months), the abnormalities of T, P, and NK cells disappeared. These findings suggest the existence of a close rela- tionship between EtOH consumption and the abnormalities of the immune system observed during active alcoholism. Nevertheless, ALCAW individualsdisplayed marked alterationson the immunophe- notypic profile, as reflected by a significantly decreased number of total T cells, due to reduced levels of the CD3+/TCR alpha beta+, CD4+, CD8+, and CD4+/CD45RA+ T-cell subsets. In addition, a significantly decreased number of total PB B cells was observed in this group of patients. Our results show that in patients suffering from ALC, the abnormalities of the immune system due to a direct effect of EtOH intake (or its metabolites) should be distinguished from the immunologicalalterations related to the liver disease itself. Key Words: Alcoholism, Ethanol, Alcohol Withdrawal, Lymphoid Subsets, Liver Cirrhosis. EVERAL PIECES of evidence have shown that immu- S nological reactions are present in patients with alco- holic liver disease (ALD).l The advent of the automated flow cytometry systems has stimulated investigations into whether specific alterations can be detected in the periph- eral blood (PB) immunological cell compartment, but zyxwvu From the Servicio de Medicina Intema zyxwvutsrqpo II, Hospital Universitario (F.J.L., J.I.M.),Servicio de Hematologia, Hospital Universitario (J.S.M.)and Servicio General de Citometria,. Universidad de Salamanca (J.C., A.L., A.O.), Salamanca, Spain; Departamento de Medicina, Universidad de Alcala de Henares (M.A.M.),Alcala de Henares, Madrid Spain. Received for publication February 12, 1996; accepted August 19, 1996. This study was supported by Institutional Grant FIS 91l190201 from the Fondo de Investigaciones Sanitarias. Reprint requests: Francisco Javier Laso, M.D., Servicio de Medicina Intema II, Hospital Universitario, Paseo de San Mcente 58-182, 37007 Salamanca, Spain. Copyright zyxwvutsrqpon 0 I996 by The Research Society on Alcoholism. 1564 unfortunately the results obtained are frequently contra- dictory.2-6 Discrepant results about immunological abnor- malities in alcoholism may be related to several factors including: (1) the heterogeneity of the lymphoid subsets evaluated; (2) the timing that the study was performed (active alcoholism versus withdrawal period); (3) factors associated with alcoholism and/or ALD, such as malnutri- ti~n,~ or chronic infection by hepatitis B and C vir~ses,'~~ and; (4) wide range of methodological approaches used to analyze PB lymphoid subsets, most of them lacking on the simultaneous assessment of more than one antigen, which is necessary for their correct evaluation. Moreover, at present, an open debate still exists on whether immunolog- ical alterations represent a consequence or a cause of the liver injury, and little is known about the direct role played by ethanol (EtOH) in these immunological changes. The aim of the present study was to investigate the effect of chronic EtOH consumption on the immune system in patients with alcoholic liver cirrhosis zyx (ALC), as analyzed by the distribution of PB T-, B-, and NK-lymphoid subsets using multiple stainings with monoclonal antibodies and flow cytometry. For that purpose, we have analyzed a group of 20 patients with ALC and active EtOH intake (ALCET group). After this evaluation, the patients were subse- quently studied 3 months after alcohol consumption was discontinued. As reference controls, we have used both a group of 15 patients with ALC and at least 1 year of alcohol withdrawal (ALCAW group) before our laboratory study and a group of 25 healthy subjects. Our results indicate that in ALC, EtOH either directly or through its metabolites induces marked alterations of the immunophenotypic pro- file, irrespective of liver damage. MATERIALS AND METHODS Patients A total of 35 patients with ALC (20 patients with ALCET and 15 with ALCAW) were included in the study. All patients were referred to the Alcoholism Unit of the University Hospital of Salamanca, and they had consumed at least 90 g of EtOH daily for more than 10 years. Patients within the ALCET group displayed an active alcohol intake at the moment of entering the study, whereas those from the ALCAW group reported at least 1 year of alcohol withdrawal. In 24 cases, the liver histopathological examination showed a micronodular cirrhosis; none of the liver biopsy specimens showed signs of alcoholic hepatitis. In 11 patients, liver biopsy was not performed because of blood coagulation abnormalities, the diag- nosis of ALC was established on the basis of the presence of physical Alcohol Clin Exp Res, Vol20, No 9, 1996: pp 1564-1568