Skeletal muscle aldolase an overexpression in endotoxemic rats and inhibited by GSNO via potential role for S-nitrosylation in vitro.

ABSTRACT

BACKGROUND: Hepatic aldolase (ALD) A mRNA transcription and ALD B S-nitrosylation have been confirmed in endotoxemic rats and mice, respectively. In the present study we investigated whether the skeletal muscle ALD A shared potential for S-nitrosylation to act as a hypoxia-related signaling mechanism in lipopolysaccharide (LPS) challenged rats. MATERIALS AND METHODS: Male Sprague Dawley rats were treated (i.p.) as follows, control group (n = 6) with 0.9% NaCl, tested group (n = 6) with a single dose of 2 mg/kg LPS. Protein S-nitrosylation was determined by biotin switch and dot blotting analysis. ALD A, hypoxia-inducible factor 1α and vascular endothelial growth factor were determined by western blotting. ALD A catalytic activity treated with S-nitrosoglutathione (GSNO), an exogenous NO-donor, was examined in vitro. RESULTS: There were several S-nitrosylated proteins under basal conditions. ALD A was over-expressed in a hypoxia-related way in the skeletal muscle of LPS challenged rats. Importantly, treatment of ALD A with GSNO at concentration 50 µmol/L ∼ 1000 µmol/L that inhibited catalytic activity, increased the number of S-nitrosylated bands and led to hyper-nitrosylation of basally S-nitrosylated proteins of ALD A. Quantization of enzyme S-nitrosothiol showed that a maximal of four cysteines per subunit was modified by S-nitrosylation in the presence of GSNO. CONCLUSIONS: These findings suggested that S-nitrosylation of ALD A might serve as a novel mechanism for controlling ALD A activity at the post-translational level in endotoxemic rats.