Quantitative analyses and transcriptomic profiling of circulating messenger RNAs as biomarkers of rat liver injury.
Serum aminotransferases have been the clinical standard for evaluating liver injury for the past 50-60 years. These tissue enzymes lack specificity, also tracking injury to other tissues. New technologies assessing tissue-specific messenger RNA (mRNA) release into blood should provide greater specificity and permit indirect assessment of gene expression status of injured tissue. To evaluate the potential of circulating mRNAs as biomarkers of liver injury, rats were treated either with hepatotoxic doses of D-(+)-galactosamine (DGAL) or acetaminophen (APAP) or a myotoxic dose of bupivacaine HCl (BPVC). Plasma, serum, and liver samples were obtained from each rat. Serum alanine aminotransferase and aspartate aminotransferase were increased by all three compounds, whereas circulating liver-specific mRNAs were only increased by the hepatotoxicants. With APAP, liver-specific mRNAs were significantly increased in plasma at doses that had no effect on serum aminotransferases or liver histopathology. Characterization of the circulating mRNAs by sucrose density gradient centrifugation revealed that the liver-specific mRNAs were associated with both necrotic debris and microvesicles. DGAL treatment also induced a shift in the size of plasma microvesicles, consistent with active release of microvesicles following liver injury. Finally, gene expression microarray analysis of the plasma following DGAL and APAP treatment revealed chemical-specific profiles.


CONCLUSIONS
The comparative analysis of circulating liver mRNAs with traditional serum transaminases and histopathology indicated that the circulating liver mRNAs were more specific and more sensitive biomarkers of liver injury. Further, the possibility of identifying chemical-specific transcriptional profiles from circulating mRNAs could open a range of possibilities for identifying the etiology of drug/chemical-induced liver injury.
The Hamner Institutes for Health Sciences, Research Triangle Park, North Carolina 27709-2137, USA.
Barbara A
BA
Wetmore
Dominique J
DJ
Brees
Reetu
R
Singh
Paul B
PB
Watkins
Melvin E
ME
Andersen
James
J
Loy
Russell S
RS
Thomas
D000082:Acetaminophen; D000410:Alanine Transaminase / Q000097:blood; D000418:Albumins / Q000378:metabolism; D018712:Analgesics, Non-Narcotic; D000779:Anesthetics, Local; D000818:Animals; D001219:Aspartate Aminotransferases / Q000097:blood; D015415:Biomarkers / Q000097:blood*; D002045:Bupivacaine; D002499:Centrifugation, Density Gradient; D056486:Chemical and Drug Induced Liver Injury / Q000097:blood* / Q000209:etiology / Q000473:pathology; D004305:Dose-Response Relationship, Drug; D005340:Fibrinogen / Q000378:metabolism; D005688:Galactosamine; D020869:Gene Expression Profiling; D006242:Haptoglobins / Q000378:metabolism; D017403:In Situ Hybridization; D008099:Liver / Q000473:pathology; D008297:Male; D009336:Necrosis; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000097:blood*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012680:Sensitivity and Specificity; D013997:Time Factors
D000418:Albumins; D018712:Analgesics, Non-Narcotic; D000779:Anesthetics, Local; C115499:BBeta fibrinogen; D015415:Biomarkers; D006242:Haptoglobins; D012333:RNA, Messenger; D000082:Acetaminophen; D005688:Galactosamine; D005340:Fibrinogen; D001219:Aspartate Aminotransferases; D000410:Alanine Transaminase; D002045:Bupivacaine