Isoaspartate, carbamoyl phosphate synthase-1, and carbonic anhydrase-III as biomarkers of liver injury

Abstract

We had previously shown that alcohol consumption can induce cellular isoaspartate protein damage via an impairment of the activity of protein isoaspartyl methyltransferase (PIMT), an enzyme that triggers repair of isoaspartate protein damage. To further investigate the mechanism of isoaspartate accumulation, hepatocytes cultured from control or 4-week ethanol-fed rats were incubated in vitro with tubercidin or adenosine. Both these agents, known to elevate intracellular S-adenosylhomocysteine levels, increased cellular isoaspartate damage over that recorded following ethanol consumption in vivo. Increased isoaspartate damage was attenuated by treatment with betaine. To characterize isoaspartate-damaged proteins that accumulate after ethanol administration, rat liver cytosolic proteins were methylated using exogenous PIMT and 3H-S-adenosylmethionine and proteins resolved by gel electrophoresis. Three major protein bands of ~75-80 kDa, ~95-100 kDa, and ~155-160 kDa were identified by autoradiography. Column chromatography used to enrich isoaspartate-damaged proteins indicated that damaged proteins from ethanol-fed rats were similar to those that accrued in the livers of PIMT knockout (KO) mice. Carbamoyl phosphate synthase-1 (CPS-1) was partially purified and identified as the ~160 kDa protein target of PIMT in ethanol-fed rats and in PIMT KO mice. Analysis of the liver proteome of 4-week ethanol-fed rats and PIMT KO mice demonstrated elevated cytosolic CPS-1 and betaine homocysteine S-methyltransferase-1 when compared to their respective controls, and a significant reduction of carbonic anhydrase-III (CA-III) evident only in ethanol-fed rats. Ethanol feeding of rats for 8 weeks resulted in a larger (~2.3-fold) increase in CPS-1 levels compared to 4-week ethanol feeding indicating that CPS-1 accumulation correlated with the duration of ethanol consumption. Collectively, our results suggest that elevated isoaspartate and CPS-1, and reduced CA-III levels could serve as biomarkers of hepatocellular injury.

Identification of consensus biomarkers for predicting non-genotoxic hepatocarcinogens

“…The downregulation of Ca3 levels by the treatment of ethanol and a hepatotoxicant of carbon tetrachloride was also reported124849. The depletion of Ca3 is also a useful biomarker for liver injury50. The usefulness of downregulated Ca3 levels for identifying NGHCs has also been demonstrated in this study.…”

Early urinary candidate biomarker discovery in a rat thioacetamide-induced liver fibrosis model

“…Carbonic anhydrase 3 promotes transformation and 343 invasion capability in hepatoma cells through FAK signaling pathway [60] and is a major 344 participant in the liver response to oxidative stress [61] . Carbonic anhydrase 3 is also a 345 biomarker of liver injury [62] . Thioredoxin has a potential to attenuate liver fibrosis via 346 suppressing oxidative stress and inhibiting proliferation of stellate cells [63] and plays 347 important roles in the pathophysiology of liver diseases [64] .…”

“…To minimize the affecting 62 factors, animal models can be used for urinary biomarker discovery of liver fibrosis [34] .…”

Carbamoyl phosphate synthetase 1 (CPS1) as a prognostic marker in chronic hepatitis C infection

“…In this light, an experimental study done by Carter et al. revealed that the levels of cytosolic CPS1 increased in conjunction with the duration of alcohol consumption in ethanol‐fed rats. This implied its potential use as a measure for the extent or length of exposure of the liver to stress.…”