REVIEW Sub-proteome approach to the knowledge of liver Juan M. Falco ´n-Pe ´rez 1 , Shelly C. Lu 2 and Jose ´ M. Mato 1 1 Metabolomics Unit, CICbioGUNE, CIBERehd, Bizkaia Technology Park, Derio, Bizkaia, Spain 2 Division of Gastrointestinal and Liver Diseases, Keck School of Medicine, University Southern California, Los Angeles, CA, USA Received: June 13, 2009 Accepted: August 12, 2009 In the recent years, global proteomics approaches have been widely used to characterize a number of tissue proteomes including plasma and liver; however, the elevated complexity of these samples in combination with the high abundance of some specific proteins make the study of the lowest abundant proteins difficult. This review is focused on different strategies that have been developed to extend the proteome focused on these two tissues, as, for example, the analysis of sub-cellular proteomes. In this regard, two special kind of extra- cellular vesicles – exosomes and membrane plasma shedding vesicles – are emerging as excellent biological source both to extend the liver and plasma proteomes and to be applied in the discovery of non-invasive liver-specific disease biomarkers. Keywords: Exosomes / Microparticles / Microvesicles / Plasma / Sub-cellular 1 Liver disease The liver is a multifunctional organ involved in important metabolic functions, detoxification of xenobiotics, and synth- esis of blood plasma components, among many other roles. Liver injury ranging from mild infection to life-threatening liver failure is a serious worldwide health issue that our society is currently facing. One of the most common liver affections is hepatitis, an inflammation of the liver that can be caused by many factors such us exposure to certain drugs and chemicals, by autoimmune diseases, and by bacterial and viral infections. Approximately 350 million people are infected with hepatitis B virus worldwide, and the World Health Organization esti- mates that approximately 170 million people are infected with hepatitis C virus. In developed countries, non-alcoholic fatty liver disease is emerging as a major global cause of liver disease in the background of an increasing prevalence of obesity and type 2 diabetes [1]. Non-alcoholic fatty liver disease encompasses a spectrum of diseases from simple steatosis through steatohepatitis, to fibrosis and ultimately cirrhosis and hepatocellular carcinoma, which is the fifth most common cancer worldwide and the third most common cause of cancer mortality [2]. Many studies have highlighted the heterogeneity of each of these pathologic processes itself emphasizing the complex nature of the establishment and progression of hepatic diseases. A major goal in liver pathol- ogy is the identification of molecular markers for early detection – before clinical manifestations are produced – for the prognosis of the different liver alterations and for moni- toring treatment efficacy. Currently, differentiation of these liver affections depends mainly on histological examination of liver biopsies. However, liver biopsy is invasive and is limited by sampling error and diagnostic accuracy, hazard to the patient [3, 4], and, remarkably, in many cases it is just infor- mative when the lesions are already produced. In addition, the numbers of patients with liver injury mean that use of liver biopsy in their investigation to reach a reliable diagnosis is both practically and financially impractical. These arguments urge the search for non-invasive alternatives such as the identification in the blood of molecular markers for each of the different clinical manifestations –steatosis, hepatitis, fibrosis, apoptosis, necrosis, proliferation – associated with liver injury. To unravel these specific biomarkers, the use of well-characterized in vitro [5, 6] and in vivo [7, 8] experimental models is necessary to control variables such as age, gender, and genetic and life-style heterogeneity, accelerating in this way the biomarker discovery process. 2 Tissue/organ proteomics Also, to accelerate this process, recent advances in proteo- mics have allowed the applications of high-throughput Abbreviations: MP, microparticle; MVB, multivesicular bodies Correspondence: Dr. Juan Manuel Falco ´ n-Pe ´ rez, Metabolomics Unit, CIC bioGUNE, CIBERehd, Bizkaia Technology Park, Bldg.801-A, Derio 48160, Bizkaia, Spain E-mail: jfalcon@cicbiogune.es Fax: 134-944-061301 & 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.clinical.proteomics-journal.com Proteomics Clin. Appl. 2010, 4, 407–415 407 DOI 10.1002/prca.200900123