Since a bromine compound with REM-sleep-inducing and anti-choline esterase activities have been isolated from human cerebrospinal fluid, and was identified as 1-methylheptyl gamma-bromoacetoacetate, the compound was chemically synthesized. It was found that this compound was composed with three forms, i.e., a keto-form, an enol-form that changed gradually from keto-form by tautomerism, and a stable six-membered ring form (= cyclic r-Br) converted from enol form, when it was chemically synthesized. In addition, it was found that the six-membered ring form of this bromine compound was present in the human blood. However, in this case, the keto-form and the enol-form were not detected. When 14C-butyrate was injected to rats, it was incorporated into the bromine compound in the blood of the animal and the bromine compound formed was found to be present mainly as the six-membered ring form. From these results, the mechanism for the formation of bromine compounds in human and animal blood were deduced. (+info)
Lactic acidosis complicating treatment of ketosis of labour.
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Hypertonic glucose, fructose, and sorbitol solutions were given intravenously to women in the first stage of labour who had ketonuria and ketonaemia as evidenced by a raised blood acetoacetate and 3-hydrosybutyrate. There was no difference in the antiketogenic action of these, which was rapid and effective, but when compared with a control group who were given normal saline they had a high incidence of hyperlactataemia, and nine out of 28 patients developed lactic acidosis after the infusions. The "lactatogenic" effect was shared by all three substrates, and when they are used in the treatment of ketosis of labour, and the mother develops lactic acidosis, they might exacerbate pre-existing lactic acidosis and precipitate fetal distress. (+info)
Compartmentation of acetyl-coA in rat-liver mitochondria.
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The ratio of the specific radioactivities of 3-hydroxybutyrate: citrate was determined in rat liver mitochondria which were incubated in the presence of [1-14C]palmitate, pyruvate, bicarbonate, ATP, phosphate and malonate. Without compartmentation this ratio would maximally be 2, however, under our conditions values of 2.5-3.7 were observed. In further experiments with mitochondria, the sensitivity of pyruvate carboxylase for acetyl-CoA produced from various precursors was tested. It was found that acetyl-CoA produced from L-acetylcarnitine or by oxidation from either pyruvate, octanoate or palmitylcarnitine but not from leucine led to a stimulation of pyruvate carboxylation. These results demonstrate a compartmentation of acetyl-CoA in liver mitochondria. The further finding that different mitochondrial fractions showed varying ratios of specific radioactivities of 3-hydroxybutyrate:citrate indicates that the observed compartmentation may be explained by the existence of different types of mitochondria with varying enzyme patterns and acetyl-CoA pools. (+info)
Maleylacetoacetate isomerase (MAAI/GSTZ)-deficient mice reveal a glutathione-dependent nonenzymatic bypass in tyrosine catabolism.
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In mammals, the catabolic pathway of phenylalanine and tyrosine is found in liver (hepatocytes) and kidney (proximal tubular cells). There are well-described human diseases associated with deficiencies of all enzymes in this pathway except for maleylacetoacetate isomerase (MAAI), which converts maleylacetoacetate (MAA) to fumarylacetoacetate (FAA). MAAI is also known as glutathione transferase zeta (GSTZ1). Here, we describe the phenotype of mice with a targeted deletion of the MAAI (GSTZ1) gene. MAAI-deficient mice accumulated FAA and succinylacetone in urine but appeared otherwise healthy. This observation suggested that either accumulating MAA is not toxic or an alternate pathway for MAA metabolism exists. A complete redundancy of MAAI could be ruled out because substrate overload of the tyrosine catabolic pathway (administration of homogentisic acid, phenylalanine, or tyrosine) resulted in renal and hepatic damage. However, evidence for a partial bypass of MAAI activity was also found. Mice doubly mutant for MAAI and fumarylacetoacetate hydrolase (FAH) died rapidly on a normal diet, indicating that MAA could be isomerized to FAA in the absence of MAAI. Double mutants showed predominant renal injury, indicating that this organ is the primary target for the accumulated compound(s) resulting from MAAI deficiency. A glutathione-mediated isomerization of MAA to FAA independent of MAAI enzyme was demonstrated in vitro. This nonenzymatic bypass is likely responsible for the lack of a phenotype in nonstressed MAAI mutant mice. (+info)
Breath acetone is a reliable indicator of ketosis in adults consuming ketogenic meals.
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BACKGROUND: Ketogenic diets are used therapeutically to treat intractable seizures. Clinically, it appears that the maintenance of ketosis is crucial to the efficacy of the diet in ameliorating seizures. To understand how ketosis and seizure protection are related, a reliable, noninvasive measure of ketosis that can be performed frequently with minimal discomfort is needed. OBJECTIVE: The objective was to determine which index, breath acetone or urinary acetoacetate, is more strongly related to the plasma ketones acetoacetate and beta-hydroxybutyrate. DESIGN: After fasting overnight for 12 h, 12 healthy adults consumed 4 ketogenic meals over 12 h. Blood, breath, and urine samples were collected hourly. Blood was analyzed for plasma acetoacetate and beta-hydroxybutyrate, breath for acetone, and urine for acetoacetate. RESULTS: By the end of the 12-h dietary treatment, plasma acetoacetate, plasma beta-hydroxybutyrate, and breath acetone had increased 3.5-fold, whereas urinary acetoacetate increased 13-fold when measured enzymatically and 25-fold when measured with urinary ketone dipsticks. Plasma acetoacetate was best predicted by breath acetone (R(2) = 0.70, P < 0.0001). Plasma beta-hydroxybutyrate was equally predicted by breath acetone and urinary acetoacetate (R(2) = 0.54, P = 0.0040). CONCLUSIONS: Breath acetone is as good a predictor of ketosis as is urinary acetoacetate. Breath acetone analysis is noninvasive and can be performed frequently with minimal discomfort to patients. As an indicator of ketosis in epilepsy patients consuming a ketogenic diet, breath acetone may be useful for understanding the mechanism of the diet, elucidating the importance of ketosis in seizure protection, and ultimately, enhancing the efficacy of the diet by improving patient monitoring. (+info)
Hyperketonemia increases tumor necrosis factor-alpha secretion in cultured U937 monocytes and Type 1 diabetic patients and is apparently mediated by oxidative stress and cAMP deficiency.
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An elevated blood level of tumor necrosis factor (TNF)-alpha is a validated marker of vascular inflammation, which can result in the development of vascular disease and atherosclerosis. This study examined the hypothesis that ketosis increases the TNF-alpha secretion, both in a cell culture model using U937 monocytes and in type 1 diabetic patients in vivo. U937 cells were cultured with ketone bodies (acetoacetate [AA] and beta-hydroxybutyrate [BHB]) in the presence or absence of high levels of glucose in medium at 37 degrees C for 24 h. This study demonstrates the following points. First, hyperketonemic diabetic patients have significantly higher levels of TNF-alpha than normoketonemic diabetic patients (P < 0.01) and normal control subjects (P < 0.01). There was a significant correlation (r = 0.36, P < 0.05; n = 34) between ketosis and oxidative stress as well as between oxidative stress and TNF-alpha levels (r = 0.47, P < 0.02; n = 34) in the blood of diabetic patients. Second, ketone body AA treatment increases TNF-alpha secretion, increases oxygen radicals production, and lowers cAMP levels in U937 cells. However, BHB did not have any effect on TNF-alpha secretion or oxygen radicals production in U937 cells. Third, exogenous addition of dibutyryl cAMP, endogenous stimulation of cAMP production by forskolin, and antioxidant N-acetylcysteine (NAC) prevented stimulation of TNF-alpha secretion caused by AA alone or with high glucose. Similarly, NAC prevented the elevation of TNF-alpha secretion and lowering of cAMP levels in H(2)O(2)-treated U937 cells. Fourth, the effect of AA on TNF-alpha secretion was inhibited by specific inhibitors of protein kinase A (H89), p38-mitogen-activated protein kinase (SB203580), and nuclear transcription factor (NF)kappaB (NFkappaB-SN50). This study demonstrates that hyperketonemia increases TNF-alpha secretion in cultured U937 monocytic cells and TNF-alpha levels in the blood of type 1 diabetic patients and is apparently mediated by AA-induced cellular oxidative stress and cAMP deficiency. (+info)
Complex I-mediated reactive oxygen species generation: modulation by cytochrome c and NAD(P)+ oxidation-reduction state.
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Several lines of evidence indicate that mitochondrial reactive oxygen species (ROS) generation is the major source of oxidative stress in the cell. It has been shown that ROS production accompanies cytochrome c release in different apoptotic paradigms, but the site(s) of ROS production remain obscure. In the current study, we demonstrate that loss of cytochrome c by mitochondria oxidizing NAD(+)-linked substrates results in a dramatic increase of ROS production and respiratory inhibition. This increased ROS production can be mimicked by rotenone, a complex I inhibitor, as well as other chemical inhibitors of electron flow that act further downstream in the electron transport chain. The effects of cytochrome c depletion from mitoplasts on ROS production and respiration are reversible upon addition of exogenous cytochrome c. Thus in these models of mitochondrial injury, a primary site of ROS generation in both brain and heart mitochondria is proximal to the rotenone inhibitory site, rather than in complex III. ROS production at complex I is critically dependent upon a highly reduced state of the mitochondrial NAD(P)(+) pool and is achieved upon nearly complete inhibition of the respiratory chain. Redox clamp experiments using the acetoacetate/L-beta-hydroxybutyrate couple in the presence of a maximally inhibitory rotenone concentration suggest that the site is approx. 50 mV more electronegative than the NADH/NAD(+) couple. In the absence of inhibitors, this highly reduced state of mitochondria can be induced by reverse electron flow from succinate to NAD(+), accounting for profound ROS production in the presence of succinate. These results lead us to propose a model of thermodynamic control of mitochondrial ROS production which suggests that the ROS-generating site of complex I is the Fe-S centre N-1a. (+info)
Inhibition of human immunodeficiency virus type 1 integration by diketo derivatives.
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A series of diketo derivatives was found to inhibit human immunodeficiency virus type 1 (HIV-1) integrase activity. Only L-708,906 inhibited the replication of HIV-1(III(B)) (50% effective concentration, 12 micro M), HIV-1 clinical strains, HIV-1 strains resistant to reverse transcriptase or fusion inhibitors, HIV-2 (ROD strain) and simian immunodeficiency virus (MAC(251)). The combinations of L-708,906 with zidovudine, nevirapine, or nelfinavir proved to be subsynergistic. In cell culture, addition of L-708,906 could be postponed for 7 h after infection, a moment coinciding with HIV integration. Inhibition of integration in cell culture was confirmed by quantitative Alu-PCR. (+info)