Abstract
The oxythiamine (OXY) is antivitamin of thiamine. The finding that OXY increases the cytoplasmic concentration of pyruvate, known to enhance collagen biosynthesis, led us to investigate the mechanism of this antivitamin action on collagen biosynthesis in cultured human skin fibroblasts. Confluent fibroblasts were treated with micromolar concentrations (30–1,000 µM) of OXY for 24 and 48 h. It was found that OXY-dependent increase in collagen biosynthesis was accompanied by parallel increase in prolidase activity and level, compared to untreated cells. Since phosphoenolpyruvate (PEP) is known as an inhibitor of prolidase—the enzyme that plays important role in collagen biosynthesis, the mechanism of pyruvate interconversion was considered as a regulatory switch in collagen biosynthesis. In fact, 3-MPA, specific inhibitor of phosphoenolpyruvate carboxykinase (PEPCK), contributed to up-regulation of prolidase activity, suggesting that down-regulation of PEP formation is an underlying mechanism. Since collagen biosynthesis and prolidase activity are regulated by signal induced by activated α2β1 integrin receptor as well as insulin-like growth factor-I receptor (IGF-IR), the expression of these receptors was measured by Western immunoblot analysis. The exposure of the cells to OXY contributed to decrease in IGF-IR, α2β1 integrin receptor, pERK1/2, and NF-κB p65 expressions. It was accompanied by increase in total ERK1/2 expression and induction of phosphorylation of Akt protein. The data suggest that OXY-dependent increase of collagen biosynthesis in cultured human skin fibroblasts results from activation of prolidase activity and level, induction in pAkt expression and down-regulation of pERK1/2 and NF-κB p65, the known inhibitor of collagen gene expression.
Highlights
Since collagen biosynthesis and prolidase activity are regulated by signal induced by activated a2b1 integrin receptor as well as insulin-like growth factor-I receptor (IGF-IR), the expression of these receptors was measured by Western immunoblot analysis
Studies were performed on confluent fibroblasts, since collagen synthesis, prolidase activity as well as IGF-IR expression depend on cell density [32, 33]
The increase in collagen biosynthesis was correlated with an increase in prolidase activity and the enzyme protein level
Summary
It interferes with the enzymes of thiamine pyrophosphate-dependent pathways and can inhibit conversion of glucose into pentose phosphate that is necessary for the synthesis of nucleotides and metabolism of amino acids [1]. OXY increases the amount of pyruvate by inhibition of pyruvate decarboxylase [2]. Pyruvate has a key position in several metabolic pathways such as glycolysis and gluconeogenesis. Inhibition of pyruvate decarboxylase, which is part of the pyruvate dehydrogenase complex may interfere with the course of the oxidative decarboxylation of pyruvate. The result of this phenomenon is accumulation of pyruvate in the cell
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