Identification of Oxidative Stress-Related Proteins for Predictive Screening of Hepatotoxicity Using a Proteomic Approach

Abstract

-We investigated the effects of three hepatotoxicants, acetaminophen (APAP), amiodarone (AD) and tetracycline (TC), on protein expression in primary cultured rat hepatocytes with toxicoproteomic approach, which is two-dimensional gel electrophoresis (2DE) and mass spectrometry. The objectives of this study were to search for alternative toxicity biomarkers which could be detected with high sensitivity prior to the appearance of morphological changes or alterations of analytical conventional biomarkers. The related proteins in the process of cell degeneration/necrosis such as cell death, lipid metabolism and lipid/carbohydrate metabolism were mainly affected under exposure to APAP, AD and TC, respectively. Among the differentially expressed proteins, several oxidative stress-related proteins were clearly identified after 24-hr exposure, even though they were not affected for 6-hr exposure. They were glutathione peroxidase (GPX) as a down-regulated protein as well as peroxiredoxin 1 (PRX1) and peroxiredoxin 2 (PRX2) as up-regulated proteins, which are known to serve as antioxidative enzymes in cells. These findings suggested that the focused proteins, GPX and PRXs, could be utilized as biomarkers of hepatotoxicity, and they were useful for setting high throughput screening methods to assess hepatotoxicity in the early stage of drug discovery.

Genomic Cluster and Network Analysis for Predictive Screening for Hepatotoxicity

“…APAP, in overdose amount, depletes hepatic glutathione and leads to hepatocellular necrosis (Mitchell et al, 1973;Pumford et al, 1997). In an in vitro model of rat hepatocytes in primary culture or in a proteomic analysis of hepatocytes, APAP-mediated oxidative stress was observed (Kikkawa et al, 2004;Yamamoto et al, 2005). Furthermore, Nrf2 is known to regulate antioxidants, detoxification enzymes, and protective proteins (Rangasamy et al, 2004); and was reported to be one of the important transcription factors for oxidative stress or detoxification, based on the results of a study on Nrf2 knockout Table 5.…”

“…The omic technologies are more helpful than traditional safety evaluation for investigation of these matters. In several studies using omic technologies, the toxicological mechanism or potential biomarkers were revealed (Fukushima et al, 2005;Yamamoto et al, 2005;Vickers et al, 2004;Mortishire-Smith et al, 2004;Sawada et al, 2005). Investigations using genomic approaches such as DNA microarray have especially increased in number.…”

In-depth Identification of Pathways Related to Cisplatin-induced Hepatotoxicity through an Integrative Method Based on an Informatics-assisted Label-free Protein Quantitation and Microarray Gene Expression Approach

“…PRDX 1 and 5 participate in the signaling transduction cascades and modulation of cell proliferation/ differentiation and cancer development or progression (39,40). Based on proteomic analysis, acetaminophen, aminoda- rone, and tetracyclin induced hepatotoxicity and increased PRDX1 expression (41). These proteins were considered to be potential biomarkers of hepatotoxicity that would be more sensitive, reliable, and mechanism-based (41,42).…”

“…Based on proteomic analysis, acetaminophen, aminoda- rone, and tetracyclin induced hepatotoxicity and increased PRDX1 expression (41). These proteins were considered to be potential biomarkers of hepatotoxicity that would be more sensitive, reliable, and mechanism-based (41,42). Therefore, PRDX1 and PRDX 5 might be important biomarkers in cisplatin-induced hepatocyte damage.…”

“…A single oral dose of cyclosporine A in rats has been reported to induce marked hepatotoxicity, as well as antioxidant enzymes, including CAT and SOD2 (43). In addition, many studies have reported that drug toxicity is modulated by antioxidant enzymes (41,42). Oxidative stress-related proteins are known to be controlled in redox regulation within hepatocytes.…”