Biomedical subjects

Jennie L Walgren

Publications and source records attributed to Jennie L Walgren.

8 recordsLinked to original sources

PubMed2006

Identification of novel peptide safety markers for exocrine pancreatic toxicity induced by cyanohydroxybutene.

Historically, serum amylase and lipase levels have been used to indicate pancreas injury; however, these enzyme levels are often not predictive of pathology. In an effort to discover novel biomarkers of pancreatic acinar cell injury, we analyzed serum and pancreas tissue from cyanohydroxybutene (CHB)-treated male IGS rats using proteomics methods. CHB produces an "edematous pancreatitis," characterized by depletion of zymogen granules, acinar cell apoptosis, and mild to moderate inflammation. Secondary necrosis occurs at higher doses. Rats were treated with 150 mg/kg of CHB and samples were collected at 4, 8, and 24 h. Analyses of serum tryptic digests by surface-enhanced laser desorption-ionization mass spectrometry revealed two novel peptide biomarkers (RA1609 and RT2864) that were predictive of pancreatic damage. Levels of RA1609 decreased, while levels of RT2864 increased by 8 h following CHB treatment. The changes in RA1609 and RT2864 were detected in media from CHB-treated primary rat acini, demonstrating that these peptides are either of pancreatic cell origin or are produced by proteases released from acinar cells. Sequencing revealed that RA1609 is a fragment of rat albumin (accession number P02770, residues 348-360) and RT2864 is a portion of either rat trypsin III (accession number P08426, residues 39-65) or bovine trypsin (accession number P00760, residues 35-61). These two peptides, and possibly other fragments of serum proteins that are digested by pancreatic proteases, may be useful as safety markers for exocrine pancreatic toxicity during drug development or as biomarkers for the diagnosis and/or grading of severity of pancreatic disease.

Acute Disease↗

PubMed2006

Evaluation of two novel peptide safety markers for exocrine pancreatic toxicity.

Current markers of exocrine pancreatic toxicity have historically been poor indicators for both early diagnosis of disease and prediction of disease severity. Recently we identified two peptide markers (RA1609 and RT2864) of pancreatic toxicity that are target organ specific. In order to evaluate sensitivity of these markers versus current standard tests for pancreatic damage (i.e., lipase), we measured amylase and lipase, as well as RA1609 and RT2864 marker levels, in serum from rats treated with four doses (50-200 mg/kg) of the model pancreatic toxicant cyanohydroxybutene (CHB). In addition, to determine whether these peptide markers could detect pancreatic injury induced by different toxicants and in different species, we measured RA1609 and RT2864 marker levels in rats treated with the pancreatic toxicant caerulein, and in mice treated with CHB. RA1609 and RT2864 peptide markers proved to be more sensitive than amylase or lipase in detecting pancreatic damage, especially at an early time point (8 h) following CHB administration. The peptide markers also accurately predicted pancreatic injury induced by caerulein in rats. These markers were sensitive in detecting very mild pancreatic damage following CHB administration in mice, which are less susceptible to CHB-induced pancreatic toxicity. In addition, a species comparison of the RA1609 albumin fragment sequence indicated that cleavage of albumin from pancreatic proteases produces a similar fragment marker in several species, including humans. To determine whether the comparable human albumin fragment could be detected in sera from pancreatitis patients, we analyzed sera from normal individuals and from patients with diabetes, vasculitis, pancreatic cancer, and pancreatitis. It was found that markers corresponding to the fragments found in rat serum (RA1609 and RT2864) were present in human serum, and changes in these were indicative of and specific to pancreatitis. In conclusion, the RA1609 and RT2864 peptides are sensitive indicators of exocrine pancreatic damage that may be useful as safety markers for general pancreatic toxicity in multiple species.

Acute Disease↗

PubMed2006

Proteomics: recent applications and new technologies.

Interest in proteomics as a tool for drug development and a myriad of other applications continues to expand at a rapid rate. Proteomic analyses have recently been conducted on tissues, biofluids, subcellular components and enzymatic pathways as well as various disease and toxicological states, in both animal models and man. In addition, several recent studies have attempted to integrate proteomics data with genomics and/or metabonomics data in a systems biology approach. The translation of proteomic technology and bioinformatics tools to clinical samples, such as in the areas of disease and toxicity biomarkers, represents one of the major opportunities and challenges facing this field. An ongoing challenge in proteomics continues to be the analysis of the serum proteome due to the vast number and complexity of proteins estimated to be present in this biofluid. Aside from the removal of the most abundant proteins, a number of interesting approaches have recently been suggested that may help reduce the overall complexity of serum analysis. In keeping with the increasing interest in applications of proteomics, the tools available for proteomic analyses continue to improve and expand. For example, enhanced tools (such as software and labeling procedures) continue to be developed for the analysis of 2D gels and protein quantification. In addition, activity-based probes are now being used to tag, enrich and isolate distinct sets of proteins based on enzymatic activity. One of the most active areas of development involves microarrays. Antibody-based microarrays have recently been released as commercial products while numerous additional capture agents (e.g. aptamers) and many additional types of microarrays are being explored.

Animals↗

PubMed2005

Lack of direct mitogenic activity of dichloroacetate and trichloroacetate in cultured rat hepatocytes.

Dichloroacetate (DCA) and trichloroacetate (TCA) are hepatocarcinogenic metabolites of the common groundwater contaminant, 1,1,2-trichloroethylene. DCA and TCA have been shown to induce hepatocyte proliferation in vivo, but it is not known if this response is the result of direct mitogenic activity or whether cell replication occurs indirectly in response to tissue injury or inflammation. In this study we used primary cultures of rat hepatocytes, a species susceptible to DCA- but not TCA-induced hepatocarcinogenesis, to determine whether DCA and TCA are direct hepatocyte mitogens. Rat hepatocytes, cultured in growth factor-free medium, were treated with 0.01-1.0 mM DCA or TCA for 10-40 h; cell replication was then assessed by measuring incorporation of 3H-thymidine into DNA and by cell counts. DCA or TCA treatment did not alter 3H-thymidine incorporation in the cultured hepatocytes. Although an increase in cell number was not observed, DCA treatment significantly abrogated the normal background cell loss, suggesting an ability to inhibit apoptotic cell death in primary hepatocyte cultures. Furthermore, treatment with DCA synergistically enhanced the mitogenic response to epidermal growth factor. The data indicate that DCA and TCA are not direct mitogens in hepatocyte cultures, which is of interest in view of their ability to stimulate hepatocyte replication in vivo. Nevertheless, the synergistic enhancement of epidermal growth factor-induced hepatocyte replication by DCA is of particular interest and warrants further study.

Animals↗

PubMed2004

Induction of peroxisome proliferation in cultured hepatocytes by a series of halogenated acetates.

Trichloroacetate (TCA) and dichloroacetate (DCA) are hepatocarcinogenic metabolites of the environmental pollutant trichloroethylene (TCE) and are common water contaminants. Induction of peroxisome proliferation via activation of the peroxisome proliferator-activated receptor alpha (PPARalpha) has been proposed as a mechanism for their hepatocarcinogenic action. However, it is unclear whether these compounds are direct ligands of PPARalpha or whether activation occurs by a ligand-independent process. The present studies were undertaken to determine whether a primary rat hepatocyte model system could be used to examine structure-activity relationships of haloacetates for the induction of peroxisomal palmitoyl-CoA oxidation. The haloacetates tested differed in both type (iodo, bromo, chloro and fluoro) and extent (mono, di and tri) substitution. Significant differences were observed in both potency and efficacy. Potency varied over about two orders of magnitude, in the order of mono > di = tri. Within the monohalo-substituted series, the order of potency was iodo > bromo > chloro, with the fluoro analog being essentially inactive. The monoiodo- and monobromo-derivatives showed significant induction at 50 and 100 microM, respectively, but cytotoxicity precluded obtaining full concentration-response curves. The dihalo- and trihalo-acetates had generally similar potency, and, with the exception of the diflouro- and dibromoacetates, showed a maximal induction of two- to three-fold. Difluoroacetate and dibromoacetate induced palmitoyl-CoA oxidation by nine- and six-fold, respectively, approaching the effectiveness of Wy-14,643 (50 microM) in this system. Of interest, the slopes of the concentration-dependence lines of the difluoro- and dibromo-acetates were markedly dissimilar from the other di- and tri-haloacetates, suggesting either a marked difference in the way they activate the PPARalpha receptor or a substantial difference in the way they are metabolized or transported by the hepatocytes.

Acetates↗

PubMed2004

Application of proteomic technologies in the drug development process.

Proteins are the principal targets of drug discovery. Most large pharmaceutical companies now have a proteomics-oriented biotech or academic partner or have started their own proteomics division. Common applications of proteomics in the drug industry include target identification and validation, identification of efficacy and toxicity biomarkers from readily accessible biological fluids, and investigations into mechanisms of drug action or toxicity. Target identification and validation involves identifying proteins whose expression levels or activities change in disease states. These proteins may serve as potential therapeutic targets or may be used to classify patients for clinical trials. Proteomics technologies may also help identify protein-protein interactions that influence either the disease state or the proposed therapy. Efficacy biomarkers are used to assess whether target modulation has occurred. They are used for the characterization of disease models and to assess the effects and mechanism of action of lead candidates in animal models. Toxicity (safety) biomarkers are used to screen compounds in pre-clinical studies for target organ toxicities as well as later on in development during clinical trials. Complementary approaches such as metabolomics and genomics can be used in conjunction with proteomics throughout the drug development process to create more of a unified, systems biology approach.

Animals↗

PubMed2004

Effect of R(+)alpha-lipoic acid on pyruvate metabolism and fatty acid oxidation in rat hepatocytes.

R-(+)-alpha-lipoic acid (R-LA) is the naturally occurring enantiomer of LA. It is a strong antioxidant and cofactor of key metabolic enzyme complexes catalyzing the decarboxylation of alpha-keto acids. Racemic LA (rac-LA) has shown promise in treating diabetic polyneuropathy, and some studies suggest that it improves glucose homeostasis in patients with type 2 diabetes. We examined the effects of R-LA on pyruvate metabolism and free fatty acid (FFA) oxidation in primary cultured hepatocytes isolated from 24-hour fasted rats. After overnight culture in serum-free medium, cells were pre-exposed to R-LA for 3 hours before assays. R-LA (25 to 200 micromol/L) significantly increased pyruvate oxidation ( approximately 2-fold at the highest dose tested) measured as (14)CO(2) production from [1-(14)C]pyruvate by the cells over 1 hour post-treatment. These effects correlated with proportional, significant increases in the activation state of the pyruvate dehydrogenase (PDH) complex. R-LA treatment inhibited glucose production from pyruvate by approximately 50% at 50 micromol/L R-LA and approximately 90% at 200 micromol/L. Palmitate oxidation was measured in hepatocytes cultured in the presence of albumin and physiological (0.1 mmol/L) or high (1.5 mmol/L) concentrations of FFA. The latter markedly enhanced FFA oxidation. R-LA treatment significantly inhibited FFA oxidation in both media, but was more effective in high FFA, where it reduced FFA oxidation by 48% to 82% at 25 to 200 micromol/L, respectively. Identical doses of R-LA did not affect FFA oxidation by L6 myotubes (a cell culture model for skeletal muscle) in either high or low FFA medium, but enhanced pyruvate oxidation. In conclusion, 3-hour exposure of primary cultured rat hepatocytes to R-LA at therapeutically relevant concentrations increased pyruvate oxidation, apparently by activation of the PDH complex, and decreased gluconeogenesis and FFA oxidation. These features may prove useful in the control of type 2 diabetes.

Animals↗

PubMedDate not supplied

Role of metabolism in drug-induced idiosyncratic hepatotoxicity.

Rare adverse reactions to drugs that are of unknown etiology, or idiosyncratic reactions, can produce severe medical complications or even death in patients. Current hypotheses suggest that metabolic activation of a drug to a reactive intermediate is a necessary, yet insufficient, step in the generation of an idiosyncratic reaction. We review evidence for this hypothesis with drugs that are associated with hepatotoxicity, one of the most common types of idiosyncratic reactions in humans. We identified 21 drugs that have either been withdrawn from the U.S. market due to hepatotoxicity or have a black box warning for hepatotoxicity. Evidence for the formation of reactive metabolites was found for 5 out of 6 drugs that were withdrawn, and 8 out of 15 drugs that have black box warnings. For the other drugs, either evidence was not available or suitable studies have not been carried out. We also review evidence for reactive intermediate formation from a number of additional drugs that have been associated with idiosyncratic hepatotoxicity but do not have black box warnings. Finally, we consider the potential role that high dosages may play in these adverse reactions.

Animals↗