Novel Biomarker Discovery in Dogs with Chronic Hepatitis

Chronic hepatitis can present a diagnostic challenge, with different types of chronic hepatic disease being associated with similar clinical and laboratory findings. Clinical signs of chronic hepatitis are often non-specific; therefore, this disease is frequently diagnosed in an advanced stage that makes successful intervention less likely. Differentiating dogs with chronic hepatitis from those with acquired hepatopathies is often difficult requiring invasive diagnostic procedures such as laparoscopic liver biopsy. There is also a need for biological markers that are able to stage chronic hepatitis non-invasively. Currently, the only way to establish a definitive diagnosis of chronic hepatitis in dogs is through the histopathological assessment of a liver biopsy specimen. The acquisition of a liver biopsy specimen, although considered the gold standard, is invasive and has limitations and risks. The identification of biomarkers that are differentially expressed in dogs with chronic hepatitis could contribute to the development of novel diagnostic tests for this disease and provide insight into its pathogenesis. The objective of these studies was to identify candidate biomarkers that are differentially expressed in the liver, serum, or urine of dogs with chronic hepatitis. The hepatic proteome, serum and urine metabolome, and hepatic microRNA transcriptome were analyzed by 2-dimensional fluorescence difference gel electrophoresis coupled to nanoflow liquid chromatography tandem mass spectrometry, gas chromatography – quadrupole time of flight mass spectrometry, and high-throughput small RNA Illumina sequencing, respectively. Differential hepatic protein expression of cytokeratin 18 and annexin 5 was demonstrated in dogs with chronic hepatitis. Untargeted metabolite profiling documented a decreased ratio of branched chain amino acids to aromatic amino acids in the serum of dogs with a congenital portosystemic shunt. The assessment of serum trans-4-hydroxyl-proline confirmed a decreased concentration in dogs with chronic hepatitis. Untargeted metabolomic profiling of urine in dogs with chronic liver disease documented alterations in metabolites involved in glutathione, arginine, proline, and nitrogen metabolism in addition to metabolites involved in fatty acid biosynthesis. Differential microRNA expression of transcripts implicated in hepatic fibrosis and apoptosis was demonstrated in dogs with chronic hepatitis. Further targeted assessment of the novel biomarkers identified in these studies is needed to determine their utility