Visceral fat accumulation in Japanese high school students and related atherosclerotic risk factors.

AIM: To investigate the factors that influence visceral fat accumulation in adolescence, we performed a medical examination of high school students and assessed abdominal fat thickness and fatty change of the liver. METHODS: A cohort of 374 Japanese high school students aged 15-16 years (193 boys and 181 girls) in public high schools in Chiba prefecture were enrolled. Anthropometric parameters, blood cell count, blood chemistry and adipocytokine levels were measured. Preperitoneal fat thickness (PFT) and echoic contrast of the liver were measured by ultrasonography. RESULTS: Anthropometric parameters, systolic blood pressure, blood cell count, ALT, AST, FBS, gamma-GTP, HDL-C, LpL, UA, adiponectin, resistin and leptin levels differed between sexes. Multivariate regression analysis revealed that leptin was the most appropriate marker for PFT in both sexes (p<0.0001). Visceral obesity, categorized as PFT exceeding 8 mm, was observed in 9.6% of all students. Boys with visceral obesity showed apparent liver dysfunction, hyperlipidemia, hyperinsulinemia, and high leptin and low adiponectin levels. Overall, 16.6% of boys and 30.4% of girls showed hepatorenal echo contrast positivity. Boys with visceral obesity and fatty liver had more risk factors for atherosclerosis. CONCLUSIONS: Physical examination of high school students is important for early detection of atherosclerosis.

NAD(H)-dependent glutamate dehydrogenase is essential for the survival of Arabidopsis thaliana during dark-induced carbon starvation.

Interconversion between glutamate and 2-oxoglutarate, which can be catalysed by glutamate dehydrogenase (GDH), is a key reaction in plant carbon (C) and nitrogen (N) metabolism. However, the physiological role of plant GDH has been a controversial issue for several decades. To elucidate the function of GDH, the expression of GDH in various tissues of Arabidopsis thaliana was studied. Results suggested that the expression of two Arabidopsis GDH genes was differently regulated depending on the organ/tissue types and cellular C availability. Moreover, Arabidopsis mutants defective in GDH genes were identified and characterized. The two isolated mutants, gdh1-2 and gdh2-1, were crossed to make a double knockout mutant, gdh1-2/gdh2-1, which contained negligible levels of NAD(H)-dependent GDH activity. Phenotypic analysis on these mutants revealed an increased susceptibility of gdh1-2/gdh2-1 plants to C-deficient conditions. This conditional phenotype of the double knockout mutant supports the catabolic role of GDH and its role in fuelling the TCA cycle during C starvation. The reduced rate of glutamate catabolism in the gdh2-1 and gdh1-2/gdh2-1 plants was also evident by the growth retardation of these mutants when glutamate was supplied as the alternative N source. Furthermore, amino acid profiles during prolonged dark conditions were significantly different between WT and the gdh mutant plants. For instance, glutamate levels increased in WT plants but decreased in gdh1-2/gdh2-1 plants, and aberrant accumulation of several amino acids was detected in the gdh1-2/gdh2-1 plants. These results suggest that GDH plays a central role in amino acid breakdown under C-deficient conditions.

Contribution of the GABA shunt to hypoxia-induced alanine accumulation in roots of Arabidopsis thaliana.

When subjected to low oxygen stress, plants accumulate alanine and gamma-aminobutyric acid (GABA). To investigate the function of GABA metabolism under hypoxia and its contribution to alanine accumulation, we studied the genes that encode the two key enzymes of the GABA shunt, glutamate decarboxylase (GAD) and GABA transaminase (GABA-T). Among the five homologous GAD genes found in Arabidopsis thaliana, GAD1 expression was predominantly found in roots, while GAD2 expression was evident in all organs. Expression of the other three GAD genes was generally weak. In response to hypoxia, transcriptional induction was observed for GAD4 only. For GABA-T1, its expression was detected in all organs, but there was no significant transcriptional change under hypoxic conditions. Moreover, we have isolated and characterized Arabidopsis mutants defective in GAD1 and GABA-T1. In gad1 mutants, GAD activity was significantly reduced in roots but was not affected in shoots. In the gaba-t1 mutant, GABA-T activity was decreased to negligible levels in both shoots and roots. These mutants were phenotypically normal under normal growth conditions except for the reduced seed production of the pop2 mutants as described previously. However, metabolite analysis revealed significant changes in GABA content in gad1 and gaba-t1 mutants. The levels of alanine under hypoxic conditions were also affected in the roots of gad1 and gaba-t1 mutants. The partial inhibition of the hypoxia-induced alanine accumulation in roots of these mutants suggests that the GABA shunt is, in part, responsible for the alanine accumulation under hypoxia.

Glutamate deamination by glutamate dehydrogenase plays a central role in amino acid catabolism in plants.

Glutamate is of central importance in plant N metabolism since the biosynthesis of all other amino acids requires this compound. Glutamate dehydrogenase (GDH; EC 1.4.1.2), which catalyzes in vitro reversible reductive amination of 2-oxoglutatre to form glutamate, is a key player in the metabolism of glutamate. While most previous studies have indicated that the oxidative deamination is the in vivo direction of the GDH reaction, its physiological role has remained ambiguous for decades. We have recently isolated mutants for the two known Arabidopsis GDH genes and created a gdh double mutant. Our recent work revealed an increased susceptibility of the gdh double mutant to dark-induced C starvation, the first phenotype associated with the loss of GDH activity in plants. Monitoring the amino acid breakdown during the dark treatment also suggested that the deamination of glutamate catalyzed by GDH is central to the catabolism of many other amino acids.

Functional analysis of lactate dehydrogenase during hypoxic stress in Arabidopsis.

During waterlogging conditions plants switch from aerobic respiration to anaerobic fermentation to cope with the lack of available oxygen. Plants have two main fermentation pathways: ethanol and lactic acid fermentation. In this paper we carry out a functional analysis of the Arabidopsis lactate dehydrogenase gene, LDH1. Our results indicate that LDH1, like some other anaerobic genes, is expressed in a root-specific manner and is affected by a variety of abiotic stresses (hypoxia, drought, cold) and mechanical wounding. Functional analysis of LDH1 was carried out using transgenic Arabidopsis overexpressing the gene (35S promoter) and a T-DNA knockout line. Overexpression of LDH1 resulted in improved survival of low oxygen stress conditions in roots but not in shoots. Increased lactic acid fermentation also resulted in significantly higher activities of pyruvate decarboxylase (PDC). Knockout mutants of LDH1 showed reduced survival under low oxygen conditions and PDC activity levels were not changed compared with the wild type. Our data suggest that there is an interdependency between the lactic and ethanol fermentation pathways and that lactic acid fermentation may play a role in stimulating ethanol fermentation and improving plant survival. We show also that Arabidopsis plants are able to exude lactate efficiently into the medium, preventing it accumulating to toxic levels in the cells.

Alanine aminotransferase catalyses the breakdown of alanine after hypoxia in Arabidopsis thaliana.

Alanine aminotransferase (AlaAT) catalyses the reversible transfer of an amino group from glutamate to pyruvate to form 2-oxoglutarate and alanine. The regulation of AlaAT in several plant species has been studied in response to low-oxygen stress, light and nitrogen application. In this study, induction of Arabidopsis AlaAT1 and AlaAT2 during hypoxia was observed at the transcriptional level, and an increase in enzyme activity was detected in hypoxically treated roots. In addition, the tissue-specific expression of AlaAT1 and AlaAT2 was analysed using promoter:GUS fusions. The GUS staining patterns indicated that both AlaAT genes are expressed predominantly in vascular tissues. We manipulated AlaAT expression to determine the relative importance of this enzyme in low-oxygen stress tolerance and nitrogen metabolism. This was done by analysing T-DNA mutants and over-expressing barley AlaAT in Arabidopsis. The AlaAT1 knockout mutant (alaat1-1) showed a dramatic reduction in AlaAT activity, suggesting that AlaAT1 is the major AlaAT isozyme in Arabidopsis. Over-expression of barley AlaAT significantly increased the AlaAT activity in the transgenic plants. These plants were analysed for metabolic changes over a period of hypoxic stress and during their subsequent recovery. The results showed that alaat1-1 plants accumulate more alanine than wild-type plants during the early phase of hypoxia, and the decline in accumulated alanine was delayed in the alaat1-1 line during the post-hypoxia recovery period. When alanine was supplied as the nitrogen source, alaat1-1 plants utilized alanine less efficiently than wild-type plants did. These results indicate that the primary role of AlaAT1 is to break down alanine when it is in excess. Therefore, AlaAT appears to be crucial for the rapid conversion of alanine to pyruvate during recovery from low-oxygen stress.