Diagnostic area

URIFIB

IDENTIFICATION OF FIBROSIS BIOMARKERS IN HUMAN URINE SAMPLES

Alterations or fibrotic diseases are a major cause of morbidity and mortality and its chronic character impact on a patients and society with considerable financial burden. In particular, liver fibrosis is the main complication of chronic liver damage and leads to long-term progression to cirrhosis. The most common causes of hepatic fibrosis are alcohol intake, infections with hepatitis C virus and nonalcoholic steatohepatitis (NASH). Moreover, it significantly contributes to the development of liver failure and portal hypertension. Consequently, the evaluation of the presence and severity of hepatic fibrosis is an important parameter in clinical practice and is a valuable indicator of risk of progression to cirrhosis. Although there are methods that detect extracellular matrix accumulation in various tissues, none is ideal, because on the one hand they are invasive methods or, on the other hand their relative effectiveness to accurately determine the degree of fibrosis, mainly in the early stages. The identification of biomarkers to effectively define the stage of a disease is one of the major areas of interest in biomedicine. On the other hand, the availability of appropriate methods for assessing the presence and severity of fibrosis is a crucial tool in research of new molecules potentially anti-fibrosing.

There are evidences that many disease processes are associated with quantitative and functional changes in the molecular constituents of body fluids. As the urine body fluid is readily available, would be desirable and advantageous to develop a method of evaluating liver fibrosis based on the identification and quantification of analytes fibrosis indicators, such as proteins, in urine samples. Our goal is therefore to develop a method for assessing the presence and severity of hepatic fibrosis, and fibrosis in other vital organs too, through the identification and quantification of different analytes detectable indicators in a urine sample.

Previous studies allowed detection of 4 potential markers of fibrosis in urine using two dimensional electrophoresis analyses. We are currently extending our proteomic study using the most common and successful techniques in biomarkers research:

1.- DIGE

2.- SELDI-TOF

3.- MicroHPLC (n-dimensional) combined with MALDI TOF MS and ESI-MS/MS to analyze the low molecular weight proteome.

Thus, fractionation of complex protein samples by chromatographic retention on chips and subsequent analysis of these by mass spectrometry, SELDI-TOF (Surface Enhanced Laser Desorption/Ionization-Time of Flight), is a fast and extremely useful analysis platform for comparative analysis of protein expression profiles. These studies have provided 20 potential markers that are under evaluation, which probably help to create the first URIFIB test prototype. Furthermore, the use of the MicroHPLC (n-dimensional) technique combined with MALDI TOF MS and ESI-MS/MS, it gives us the analysis of the low molecular weight proteome. Since urine comes from the filtration of blood, it would be logical to think that the low molecular weight proteins and peptides can significantly enrich the information obtained from proteomics and increase the possibilities of the design of diagnostic devices for clinical applications.