The use of the Dhcr7 knockout mouse to accurately determine the origin of fetal sterols

Mice with a targeted mutation of 3beta-hydroxysterol Delta(7)-reductase (Dhcr7) that cannot convert 7-dehydrocholesterol to cholesterol were used to identify the origin of fetal sterols. Because their heterozygous mothers synthesize cholesterol normally, virtually all sterols found in a Dhcr7 knockout fetus having a Delta(7) or a Delta(8) double bond must have been synthesized by the fetus itself but any cholesterol had to have come from the mother. Early in gestation, most fetal sterols were of maternal origin, but at approximately E13-14, in situ synthesis became increasingly important, and by birth, 55-60% of liver and lung sterols had been made by the fetus. In contrast, at E10-11, upon formation of the blood-brain barrier, the brain rapidly became the source of almost all of its own sterols (90% at birth). New, rapid, de novo sterol synthesis in brain was confirmed by the observation that concentrations of C24,25-unsaturated sterols were low in the brains of all very young fetuses but increased rapidly beginning at approximately E11-12. Reduced activity of sterol C24,25-reductase (Dhcr24) in brain, suggested by the abundance of C24,25-unsaturated compounds, seems to be the result of suppressed Dhcr24 expression. The early fetal brain also appears to conserve cholesterol by keeping cholesterol 24-hydroxylase expression low until approximately E18.     

Effects of Sulpiride on Prolactin and mRNA Levels of Steroid 5α-reductase Isozymes in Adult Rat Brain

Prolactin (PRL) promotes maternal behavior (MB), a complex pattern of behavior aimed at maximizing offspring survival. 3α,5α-reduced neurosteroids may also regulate MB. Indeed, PRL, 3α,5α-reduced neurosteroids, and 5α-reductase (5α-R), the key enzyme in the biosynthesis of these neuroactive steroids, are all increased in stress situations These facts led us to hypothesize a possible interrelation between PRL levels and 5α-R. In the present study we quantified mRNA levels of both 5α-R isozymes in prefrontal cortex of male and female rats after administration of sulpiride, an inductor of PRL secretion. Our results demonstrated that mRNA levels of both 5α-R isozymes were significantly increased in male and female rats by sulpiride, directly or via sulpiride-induced hyperprolactinemia. Since 3α,5α-reduced neurosteroids and PRL exert anxiolytic effects in response to stress, these molecules and 5α-R may possibly participate in a common pathway of significant adaptation to stress situations.     

The Potential Use of Mechanism-Based Enzyme Inactivators in Medicine

Abstract

A new three dimensional representation of enzyme inhibition, applied to Lineweaver-Burk, Hanes and Eadie-Hofstee plots is presented. This type of representation has advantages for enzyme inhibition diagnosis, showing graphic characteristics that pass unnoticed in linear plots.     

Evaluation of metabolic alteration in transgenic rice overexpressing dihydroflavonol-4-reductase

BACKGROUND AND AIMS: Previous studies have shown that transgenic rice plants overexpressing YK1, which possesses dihydroflavonol-4-reductase (DFR) activity, showed biotic and abiotic stress tolerance. High throughput profiles of metabolites have also been shown in such transgenic plants by Fourier transform ion cyclotron mass spectrometry. In this study, capillary electrophoresis mass spectrometry analysis (CE/MS) was employed to identify precise metabolites such as organic acids, amino acids and sugars. METHODS: Using CE/MS, we analysed several metabolites of glycolysis, the tricarboxylic acid (TCA) cycle and the pentose phosphate pathway. In addition, the concentrations of sugars and ion were quantified. KEY RESULTS: In YK1 (DFR)-overexpressing plants, the concentrations of cis-aconitate, isocitrate and 2-oxoglutarate were higher in leaves, whereas those of fructose-1,6-bisphosphate and glyceraldehyde-3-phosphate were lower in roots. In seeds, the amounts of free amino acids and metals were altered, whereas sugars in seeds were kept constant. In YK1 calli, an approx. 3-fold increase in glutathione was observed, whereas the activities of glutathione peroxidase and glutathione reductase were concomitantly increased. CONCLUSIONS: The overexpression of YK1 (DFR) was associated with slight changes in the amounts of several metabolites analysed in whole plants, whilst glutathione derivatives were substantially increased in suspension-cultured cells.