Endocrine Abstracts (2006) 11 P334

Regulation of hexose-6-phosphate dehydrogenase (H6PDH) in human fetal liver WRL-68 cells

A Swali, I Bujalska, PM Stewart & EA Walker

The University of Birmingham, Birmingham, United Kingdom.

Excessive glucocorticoid exposure has been implicated in the pathogenesis of obesity and the metabolic syndrome. The in vivo conversion of inactive to active glucocorticoids is catalysed by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), requiring NADPH as a cofactor. Hexose-6-phosphate dehydrogenase (H6PDH) is co-localised with 11β-HSD1 in the lumen of the endoplasmic reticulum and controls local NADPH availability. Thus H6PDH plays an important role in the directionality of 11β-HSD1 and the activation of glucocorticoids. While various regulators of 11β-HSD1 have been identified, little is currently known about the regulation of H6PDH. This study aimed to assess changes in H6PDH gene expression after the administration of a variety of treatments, using a human fetal liver cell line (WRL-68). Following 24 h serum starvation, confluent cells were treated with a range of concentrations of dexamethasone (1–1000 nM), testosterone (0.1–10 μg/ml) or estradiol (0.1–10 μg/ml) for 8 or 24 h. RNA was extracted from the cells to measure H6PDH mRNA expression by real-time PCR, and Western Blots and H6PDH activity assays were utilised to assess protein expression levels. Increasing concentrations of dexamethasone (>250 nM) resulted in a dose-dependent increase in H6PDH mRNA (fold change 36.9±14.1, P<0.006, n=4). This effect was seen at 8 and 24 hours of culture (fold change: 59.5±3.2 (8 h) vs 11.0±0.2 (24 h); P<0.004). H6PDH protein expression also increased in a dose-dependent manner. Upregulation of H6PDH mRNA also occurred following treatment with testosterone (fold change: control 1.0±0.0, 0.1 μg/ml 1.1±0.03, 1.0 μg/ml 2.3±0.12, 10 μg/ml 3.5±0.03; P<0.001, n=2) and estradiol (fold change: control 1.0±0.0, 0.1μg/ml 0.6±0.08, 1.0 μg/ml 1.2±0.05, 10 μg/ml 5.1±0.49; P<0.05, n=2). Glucocorticoids and sex hormones appear to play a role in the regulation of H6PDH within liver. This regulation of H6PDH could have implications for the pathology of the metabolic syndrome.

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