Disruption of Quorum Sensing and Virulence in Burkholderia cenocepacia by a Structural Analogue of the cis-2-Dodecenoic Acid Signal.

Quorum sensing (QS) signals are widely used by bacterial pathogens to control biological functions and virulence in response to changes in cell population densities. Burkholderia cenocepacia employs a molecular mechanism in which the cis-2-dodecenoic acid (named Burkholderiadiffusible signal factor [BDSF]) QS system regulates N-acyl homoserine lactone (AHL) signal production and virulence by modulating intracellular levels of cyclic diguanosine monophosphate (c-di-GMP). Thus, inhibition of BDSF signaling may offer a non-antibiotic-based therapeutic strategy against BDSF-regulated bacterial infections. In this study, we report the synthesis of small-molecule mimics of the BDSF signal and evaluate their ability to inhibit BDSF QS signaling in B. cenocepacia A novel structural analogue of BDSF, 14-Me-C16:Δ2 (cis-14-methylpentadec-2-enoic acid), was observed to inhibit BDSF production and impair BDSF-regulated phenotypes in B. cenocepacia, including motility, biofilm formation, and virulence, while it did not inhibit the growth rate of this pathogen. 14-Me-C16:Δ2 also reduced AHL signal production. Genetic and biochemical analyses showed that 14-Me-C16:Δ2 inhibited the production of the BDSF and AHL signals by decreasing the expression of their synthase-encoding genes. Notably, 14-Me-C16:Δ2 attenuated BDSF-regulated phenotypes in various Burkholderia species. These findings suggest that 14-Me-C16:Δ2 could potentially be developed as a new therapeutic agent against pathogenic Burkholderia species by interfering with their QS signaling.IMPORTANCEBurkholderia cenocepacia is an important opportunistic pathogen which can cause life-threatening infections in susceptible individuals, particularly in cystic fibrosis and immunocompromised patients. It usually employs two types of quorum sensing (QS) systems, including the cis-2-dodecenoic acid (BDSF) system and N-acyl homoserine lactone (AHL) system, to regulate virulence. In this study, we have designed and identified an unsaturated fatty acid compound (cis-14-methylpentadec-2-enoic acid [14-Me-C16:Δ2]) that is capable of interfering with B. cenocepacia QS signaling and virulence. We demonstrate that 14-Me-C16:Δ2 reduced BDSF and AHL signal production in B. cenocepacia It also impaired QS-regulated phenotypes in various Burkholderia species. These results suggest that 14-Me-C16:Δ2 could interfere with QS signaling in many Burkholderia species and might be developed as a new antibacterial agent.

Chemotherapy diminishes lipid storage capacity of adipose tissue in a preclinical model of colon cancer.

BACKGROUND: Accelerated loss of adipose tissue in cancer is associated with shorter survival, and reduced quality of life. Evidence is emerging suggesting tumour association with alterations in adipose tissue, but much less is known about drug-related mechanisms contributing to adipose atrophy. Identification of mechanisms by which tumour and cancer treatments, such as chemotherapy, affect adipose tissue are required to develop appropriate therapeutic interventions to prevent fat depletion in cancer. This pre-clinical study aimed to assess alterations in adipose tissue during the clinical course of cancer. METHODS: Fischer 344 rats bearing the Ward colorectal tumour were euthanized before chemotherapy, after 1- cycle, or 2-cycles of a combination chemotherapy consisting of Irinotecan (CPT-11) combined with 5-fluorouracil (5-FU), which recapitulates first line treatment for human colorectal cancer. Periuterine adipose tissue was isolated. Healthy rats served as a reference group. Histological analysis (hematoxylin and eosin), Real-time PCR (TaqMan) and proteomic analysis (LC-MS/MS) were performed. RESULTS: Larger adipocytes (3993.7 ± 52.6 µm2) in tumour-bearing animals compared to the reference group (3227.7 ± 36.7 µm2; p < 0.001) was associated with reduced expression of proteins involved in mitochondrial fatty acid oxidation. The presence of a tumour has a significant effect on phospholipid but not triglyceride fatty acid composition. There were greater proportions of saturated fatty acids concurrent with lower monounsaturated fatty acids within the PL fraction of adipocytes in tumour-bearing animals. Chemotherapy treatment decreased the size of adipocytes (2243.9 ± 30.4 µm2; p < 0.001) and led to depletion of n-3 polyunsaturated fatty acids in adipose tissue triglyceride. Evaluation of the proteome profile revealed decreased expression of proteins involved in ATP generation, ß-oxidation, and lipogenesis. Overall, adipose tissue may not be able to efficiently oxidize fatty acids to provide energy to maintain energy demanding pathways like lipogenesis inside the tissue. CONCLUSIONS: In conclusion, metabolic adaptations to mitochondrial impairment may contribute to diminished lipid storage capacity of adipose tissue following chemotherapy delivery.

Marigold flower-powder exhibits significant potential to inhibit lipid oxidation in rice bran tea.

We supplemented marigold flower-powder (MFP) in rice bran tea at different proportions as a source of natural antioxidant compounds. Changes of phenolic compounds, antioxidant activity, fatty acid composition and lipid oxidation in the rice bran tea with MFP after 30 days of storage were investigated, comparing results with the initial data. Adding MFP in rice bran tea resulted in an increased content and composition of phenolics and flavonoids along with enhanced antioxidant activities, which were increased in a dose-dependent manner. As a result, MFP supplementation of rice bran tea was able to retard the lipid oxidation as determined by the peroxide value (PV), due to the protection of essential fatty acids during 30 days of storage. The PVs were strongly negatively correlated (p < 0.01) with phenolic compounds, total phenolic content (TPC) and total flavonoid content (TFC), but were positively correlated with tocopherols and γ-oryzanol contents. We also found that the PV was positively correlated with the PUFA (poly unsaturated fatty acid) content but adverse results were found for SFA (saturated fatty acid) and MUFA (mono unsaturated fatty acid) contents. These findings suggest that MFP could be used as a natural antioxidant in foods for preventing lipid oxidation as well as extending the shelf-life of food products.

Fluvastatin inhibits the expression of fibronectin in human peritoneal mesothelial cells induced by high-glucose peritoneal dialysis solution via SGK1 pathway.

BACKGROUND: Previous studies showed that statins may have protective effects on peritoneal mesothelial cells (PMC) cultured in high glucose. However, the mechanisms are not clear yet. Several studies demonstrated that serum- and glucocorticoid-inducible kinase 1 (SGK1) is implicated in tissue fibrosis of liver, lung and kidney by regulating the expression of many profibrogenic cytokines and extracellular matrix (e.g., fibronectin). However, few available reports elucidated whether the SGK1 is involved in the pathogenesis of peritoneal fibrosis (PF) in patients with peritoneal dialysis (PD). So far, there is no study about the interaction between the statins and SGK1 in PMC. The purpose of this study was to identify whether fluvastatin may decrease the expression of fibronectin (FN) in human peritoneal mesothelial cells (HPMC) cultured with high-glucose peritoneal dialysis solution (HGPDS) by affecting SGK1 signal pathway. METHODS: Cultured HPMC were divided into groups of control, high-glucose peritoneal dialysis solution (HGPDS), HGPDS with fluvastatin (10(-8) mol/L ~ 10(-6) mol/L) or GSK650394 10(-5) mol/L (the competitive inhibitor of SGK1), fluvastatin 10(-6) mol/L or GSK650394 10(-5) mol/L alone. The expression of SGK1 and FN was detected by RT-PCR, western immunoblotting or ELISA. RESULTS: Compared with the control, the mRNA and protein expression of SGK1 and FN increased significantly in HPMC treated with HGPDS (p < 0.05). GSK650394 significantly decreased the upregulated mRNA and protein expression of SGK1 and FN induced by HGPDS (p < 0.05), and fluvastatin had the same effects as GSK650394 in a dose-dependent manner (p < 0.05). CONCLUSIONS: Expression of SGK1 and FN increased in HPMC induced by HGPDS. Treated with fluvastatin and the SGK1-inhibitor GSK650394, abnormalities of SGK1 and FN could be corrected partially, which suggested that the SGK1 pathway was implicated in the pathogenesis of PF, and that fluvastatin might decrease the expression of SGK1 so as to meliorate the progression of PF.

Oleic acid attenuates trans-10,cis-12 conjugated linoleic acid-mediated inflammatory gene expression in human adipocytes.

The weight loss supplement conjugated linoleic acid (CLA) consists of an equal mixture of trans-10,cis-12 (10,12) and cis-9,trans-11 (9,11) isomers. However, high levels of mixed CLA isomers, or the 10,12 isomer, causes chronic inflammation, lipodystrophy, or insulin resistance. We previously demonstrated that 10,12 CLA decreases de novo lipid synthesis along with the abundance and activity of stearoyl-CoA desaturase (SCD)-1, a δ-9 desaturase essential for the synthesis of monounsaturated fatty acids (MUFA). Thus, we hypothesized that the 10,12 CLA-mediated decrease in SCD-1, with the subsequent decrease in MUFA, was responsible for the observed effects. To test this hypothesis, 10,12 CLA-treated human adipocytes were supplemented with oleic acid for 12 h to 7 days, and inflammatory gene expression, insulin-stimulated glucose uptake, and lipid content were measured. Oleic acid reduced inflammatory gene expression in a dose-dependent manner, and restored the lipid content of 10,12 CLA-treated adipocytes without improving insulin-stimulated glucose uptake. In contrast, supplementation with stearic acid, a substrate for SCD-1, or 9,11 CLA did not prevent inflammatory gene expression by 10,12 CLA. Notably, 10,12 CLA impacted the expression of several G-protein coupled receptors that was attenuated by oleic acid. Collectively, these data show that oleic acid attenuates 10,12 CLA-induced inflammatory gene expression and lipid content, possibly by alleviating cell stress caused by the inhibition of MUFA needed for phospholipid and neutral lipid synthesis.

Highly purified eicosapentaenoic acid prevents the progression of hepatic steatosis by repressing monounsaturated fatty acid synthesis in high-fat/high-sucrose diet-fed mice.

Eicosapentaenoic acid (EPA) is a member of the family of n-3 polyunsaturated fatty acids (PUFAs) that are clinically used to treat hypertriglyceridemia. The triglyceride (TG) lowering effect is likely due to an alteration in lipid metabolism in the liver, but details have not been fully elucidated. To assess the effects of EPA on hepatic TG metabolism, mice were fed a high-fat and high-sucrose diet (HFHSD) for 2 weeks and were given highly purified EPA ethyl ester (EPA-E) daily by gavage. The HFHSD diet increased the hepatic TG content and the composition of monounsaturated fatty acids (MUFAs). EPA significantly suppressed the hepatic TG content that was increased by the HFHSD diet. EPA also altered the composition of fatty acids by lowering the MUFAs C16:1 and C18:1 and increasing n-3 PUFAs, including EPA and docosahexaenoic acid (DHA). Linear regression analysis revealed that hepatic TG content was significantly correlated with the ratios of C16:1/C16:0, C18:1/C18:0, and MUFA/n-3 PUFA, but was not correlated with the n-6/n-3 PUFA ratio. EPA also decreased the hepatic mRNA expression and nuclear protein level of sterol regulatory element binding protein-1c (SREBP-1c). This was reflected in the levels of lipogenic genes, such as acetyl-CoA carboxylase alpha (ACCalpha), fatty acid synthase, stearoyl-CoA desaturase 1 (SCD1), and glycerol-3-phosphate acyltransferase (GPAT), which are regulated by SREBP-1c. In conclusion, oral administration of EPA-E ameliorates hepatic fat accumulation by suppressing TG synthesis enzymes regulated by SREBP-1 and decreases hepatic MUFAs accumulation by SCD1.

4,8-Dimethyldecanal, the aggregation pheromone of Tribolium castaneum, is biosynthesized through the fatty acid pathway.

4,8-Dimethyldecanal (4,8-DMD) is the aggregation pheromone produced by male red flour beetles (RFB), Tribolium castaneum. To elucidate the biosynthetic origin of 4,8-DMD, the following studies were performed: (1) effects of juvenile hormone (JH) III, and pathway inhibitors mevastatin, an inhibitor of the mevalonate pathway, and 2-octynoic acid, an inhibitor of the fatty acid pathway, were tested to determine whether 4,8-DMD is derived from the fatty acid pathway or the mevalonate pathway; (2) incorporation of 13C-labeled acetate, propionate, and mevalonolactone into 4,8-DMD was measured to directly determine the biosynthetic origin of 4,8-DMD; and (3) incorporation of deuterium-labeled precursors, including 2-methylbutanoate (C5D), 4-methylhexanoate (C7D), 2,6-dimethyloctanoate (C10D), and 4,8-dimethyldecanoate (C12D) was tested to determine whether 4,8-DMD is biosynthesized in the sequence Ac-Pr-Ac-Pr-Ac (Ac; acetate, Pr; propionate). JH III was topically applied to males at various doses. Inhibitors and isotopically labeled substrates were administered orally by feeding the beetles flour treated with the substrates of interest, after which volatiles were collected from both sexes of RFBs. The amount of 4,8-DMD produced was significantly reduced with increasing doses of JH III. Also, 2-octynoic acid inhibited the production of 4,8-DMD, but mevastatin did not. Exposure of RFBs to [1-(13)C]acetate and [1-(13)C]propionate, but not [2-(13)C]mevalonolactone, resulted in incorporation of the labeled compounds into 4,8-DMD. RFBs fed flour treated with deuterium-labeled C5D, C10D, and C12D, but not C7D, incorporated these compounds into 4,8-DMD. The findings that the production of 4,8-DMD was inhibited by 2-octynoic acid but unaffected by mevastatin, combined with the high incorporation of [1-(13)C]acetate and [1-(13)C]propionate into 4,8-DMD and the incorporation of deuterated precursors, unambiguously demonstrated that 4,8-DMD is of fatty acid rather than terpene biosynthetic origin, and that the biosynthesis of 4,8-DMD proceeds in the sequence Ac-Pr-Ac-Pr-Ac.

Fluvastatin prevents vascular hyperplasia by inhibiting phenotype modulation and proliferation through extracellular signal-regulated kinase 1 and 2 and p38 mitogen-activated protein kinase inactivation in organ-cultured artery.

OBJECTIVE: We examined the inhibitory mechanisms of fluvastatin on FBS-induced vascular hypertrophy assessed by organ-cultured rat tail artery. METHODS AND RESULTS: After 5 days of culture with 10% FBS, hyperplastic morphological changes in the media layer were induced. Treatment with 1 mumol/L fluvastatin significantly inhibited these changes. In the FBS-cultured arteries, the protein expression ratio of alpha-actin/beta-actin was significantly decreased, indicating the change to synthetic phenotype. Fluvastatin restored the decreased expression ratio, and the addition of mevalonate (100 mumol/L) suppressed this recovery. In accordance with the synthetic morphological changes, the absolute force of contractions induced by stimuli was decreased. Fluvastatin treatment also restored the decreased contractility, and the addition of mevalonate suppressed this recovery. In the arteries cultured with FBS, extracellular signal-regulated kinase 1 and 2 (ERK1/2) and p38 mitogen-activated protein kinase (p38MAPK) phosphorylation were significantly increased. Fluvastatin inhibited these phosphorylations, and mevalonate prevented the action of fluvastatin. CONCLUSIONS: These results suggest that fluvastatin inhibits vascular smooth muscle phenotype modulation to synthetic phenotype and proliferation by inhibiting the local metabolic pathway of cholesterol in smooth muscle cells, which inhibits hyperplastic changes in the vascular wall. The antihyperplastic actions by statins may be induced by inhibiting the ERK1/2 and p38MAPK activities, possibly through inhibition of prenylated Ras. We examined the inhibitory mechanisms of fluvastatin on FBS-induced vascular hypertrophy assessed by organ-cultured artery. Results suggest that fluvastatin inhibits vascular smooth muscle phenotype modulation and proliferation by inhibiting the ERK1/2 and p38MAPK activities through depletion of mevalonate in smooth muscle cells, resulting in inhibiting vascular hyperplastic changes.

Fluvastatin reduces renal fibroblast proliferation and production of type III collagen: therapeutic implications for tubulointerstitial fibrosis.

BACKGROUND: Accumulating evidence suggests that hydroxymethylglutaryl-CoA reductase inhibitors have many biological effects beyond reducing cholesterol synthesis. In a mouse model of renal interstitial fibrosis induced by unilateral ureteral obstruction, fluvastatin, one of the lipophilic hydroxymethylglutaryl-CoA reductase inhibitors, was shown to ameliorate fibrosis. METHODS: In the present study, we examined the direct effects of fluvastatin on proliferation, matrix and growth factor production by rat kidney fibroblasts (NRK-49F cells). RESULTS: Treatment with fluvastatin reduced proliferation of NRK-49F cells in a dose-dependent manner. The addition of mevalonate or geranylgeranyl pyrophosphate but not farnesyl pyrophosphate to the culture medium almost completely abolished the effect of fluvastatin. Moreover, fluvastatin treatment decreased the expression of activated Rho in NRK-49F cells suggesting that fluvastatin may decrease cell growth through blocking the activation of Rho. The majority of fluvastatin-treated cells were arrested at the G1 phase, associated with down-regulation of cyclin A and up-regulation of cyclin-dependent kinase inhibitor p27kip1, indicating that cell cycle modulation is an important mechanism. Fluvastatin significantly decreased messenger RNA expression of type III collagen and connective tissue growth factor. CONCLUSIONS: Taken together, it is suggested that fluvastatin may prevent tubulointerstitial fibrosis in a variety of progressive renal diseases by inhibiting proliferation of interstitial fibroblasts and their matrix synthesis.

Mono-unsaturated fatty acids protect against beta-cell apoptosis induced by saturated fatty acids, serum withdrawal or cytokine exposure.

Long-chain saturated fatty acids are cytotoxic to pancreatic beta-cells while shorter-chain saturated and long-chain unsaturated molecules are better tolerated. Mono-unsaturated fatty acids are not, however, inert since they inhibit the pro-apoptotic effects of saturated molecules. In the present work we show that the mono-unsaturates palmitoleate (C16:1) or oleate (C18:1) also cause marked inhibition of apoptosis induced by exposure of clonal BRIN-BD11 beta-cells to serum withdrawal or a combination of interleukin-1beta plus interferon-gamma. This response was dose-dependent and not accompanied by changes in NO formation. Taken together, the results suggest that mono-unsaturated fatty acids regulate a distal step common to several apoptotic pathways in pancreatic beta-cells.

Statins induce suppressor of cytokine signaling-3 in macrophages.

Our previous study has shown that lipophilic 3-hydroxy-3-methyl-glutaryl coenzyme A reductase inhibitors of statins can inhibit interferon-gamma-induced inducible nitric oxide synthase gene expression in RAW264.7 macrophages. In this study, we showed that lovastatin and fluvastatin are able to upregulate the mRNA expression of the suppressor of cytokine signaling-3 (SOCS-3) gene. This effect is specific for SOCS-3 and could be blocked by mevalonate, farnesyl pyrophosphate and geranylgeranyl pyrophosphate, while it was not affected by inhibitors of protein kinase C and A, mitogen-activated protein/extracellular signal-regulated kinase kinase, p38 mitogen-activated protein kinase, c-Jun N-terminal kinase, Src, Raf and Rho kinase. SOCS-3 expression results in the inhibition of interferon-gamma-, interleukin-6- and macrophage colony-stimulating factor-elicited signal transducer and activator of transcription phosphorylation, suggesting a novel anti-inflammatory mechanism of statins to down-modulate the functions of interferon-gamma-activated macrophages.

Methyl palmitate: a novel product of the Medfly (Ceratitis capitata) corpus allatum.

The corpus allatum (CA) of adult female Ceratitis capitata produces methyl palmitate (MP) in vitro, in addition to JHB(3) and JH III. Biosynthesized MP migrates on TLC and co-elutes from RP-18 HPLC with synthetic MP. Its identity is verified herein by GCMS. MP production is up-regulated twofold by mevastatin, an inhibitor of mevalonic acid-dependent isoprene biosynthesis. Fosmidomycin, an inhibitor of mevalonic acid-independent isoprene synthesis in graminaceous plants, up-regulates MP synthesis by about fourfold. However, it does not depress JHB(3) biosynthesis concurrently. This suggests that the initial enzyme(s) in the conversion of 1-deoxy-xylulose 5-phosphate to isoprene is presumably present in C. capitata, but is inhibited by fosmidomycin, and this inhibition diverts precursors to MP synthesis. Phytol, an acyclic diterpene, might be suppressing isoprene biosynthesis by CA, thereby resulting in a fourfold increase in the MP biosynthesis. Linolenic acid is an end-product and its presence in incubation media up-regulates MP biosynthesis by twofold, presumably due to the feedback diversion to biosynthesis of C(16:0) and its methyl ester. Biosynthesis of MP is markedly depressed after mating, while otherwise maintained at significantly higher levels in virgin females. MP biosynthesis is significantly reduced in virgin females by direct axonal control but is less consistent after mating.

A role for the de novo sphingolipids in apoptosis of photosensitized cells.

Sphingolipids have been implicated in apoptosis after various stress inducers. To assess the involvement of the de novo sphingolipid pathway in apoptosis, photodynamic therapy (PDT) with the photosensitizer Pc 4 was used as a novel stress inducer. Here we provide biochemical and genetic evidence of the role of the de novo sphingolipids in apoptosis post-Pc 4-PDT. In Jurkat cells PDT-induced intracellular sphinganine accumulation, DEVDase activation, PARP cleavage, and apoptosis were suppressed by the de novo sphingolipid synthesis inhibitor ISP-1 (Myriocin). Coincubation with sphinganine, sphingosine, or C16-ceramide specifically reversed the antiapoptotic actions of ISP-1 or the singlet oxygen scavenger L-histidine. PDT-induced cytochrome c release from mitochondria into the cytosol was inhibited by L-histidine, but not by ISP-1. Cotreatment with sphinganine did not reverse the inhibitory effect of L-histidine. In addition, PDT-induced sphinganine accumulation and apoptosis were ISP-1-sensitive in A431 human epidermoid and HT29 human carcinoma cells. Furthermore, in LY-B cells, CHO-derived mutants deficient in the de novo sphingolipid synthesis enzyme serine palmitoyltransferase (SPT) activity, DEVDase activation and apoptosis were delayed and suppressed post-PDT. Hence, the data are consistent with the partial involvement of the de novo sphingolipid pathway in apoptosis via DEVDase activation downstream of mitochondrial cytochrome c release post-Pc 4-PDT.

Fluvastatin inhibits basal and stimulated plasminogen activator inhibitor 1, but induces tissue type plasminogen activator in cultured human endothelial cells.

The effects of fluvastatin, a synthetic hydroxymethylglutaryl coenzyme A (HMG-CoA) inhibitor, on the biosynthesis of tissue plasminogen activator (t-PA) and of its major physiological inhibitor (plasminogen activator inhibitor type 1, PAI-1) were investigated in cultured human umbilical vein endothelial cells (HUVEC). Fluvastatin (0.1 to 2.5 microM), concentration-dependently reduced the release of PAI-1 antigen by unstimulated HUVEC, subsequent to a reduction in PAI-1 steady-state mRNA levels and de novo protein synthesis. In contrast, it increased t-PA secretion. The drug also reduced PAI-1 antigen secreted in response to 10 microg/ml bacterial lipopolysaccharide (LPS), 100 U/ml tumour necrosis factor alpha (TNFalpha) or 0.1 microM phorbol myristate acetate (PMA). Mevalonate (100 microM), a precursor of isoprenoids, added to cells simultaneously with fluvastatin, suppressed the effect of the drug on PAI-1 both in unstimulated and stimulated cells as well as on t-PA antigen. Among intermediates of the isoprenoid pathway, all-transgeranylgeraniol (5 microM) but not farnesol (10 microM) prevented the effect of 2.5 microM fluvastatin on PAI-1 antigen, which suggests that the former intermediate of the isoprenoid synthesis is responsible for the observed effects.

Induction of the mitochondrial permeability transition in vitro by short-chain carboxylic acids.

We recently reported that acrylic acid (AA) induces the MPT in vitro, which we suggested might be a critical event in the acute inflammatory and hyperplastic response of the olfactory epithelium. The purpose of the present investigation was to determine if induction of the MPT is a general response to short-chain carboxylic acids or if there are critical physical chemical parameters for this response. Freshly isolated rat liver mitochondria were incubated in the presence of varying concentrations of selected carboxylic acids. All of the acids that we tested caused a concentration-dependent induction of the MPT, which was blocked by cyclosporine A. Although the C4 carboxylic acids were slightly more potent than the C5 acids, there was no correlation with the degree of saturation, the octanol/water coefficient (log P), or the dissociation constant (pK(a)) of the acids that we tested. We conclude that induction of the MPT in vitro is a general response to short-chain carboxylic acids having a pK(a) of 4 to 5.

Prevention of fluvastatin-induced toxicity, mortality, and cardiac myopathy in pregnant rats by mevalonic acid supplementation.

Mevalonic acid is a product of the enzyme HMG-CoA reductase which is essential for cholesterol biosynthesis. Fluvastatin (Sandoz compound XU 62-320) is a potent inhibitor of this enzyme and, hence, mevalonic acid production. In three separate studies, oral administration of fluvastatin at 12 and 24 mg/kg/day to mated rats from day 15 of gestation through weaning resulted in unanticipated maternal mortality at the time of parturition and during lactation. Microscopic evaluations performed in two studies revealed significant cardiac myopathy in the dying animals. Drug-related clinical signs, significant maternal body weight loss, and an increase in stillborn pups and neonatal mortality were also noted at one or both dose levels. Supplementation of fluvastatin administration with 500 mg/kg b.i.d. of mevalonic acid completely blocked and/or ameliorated the mortality, cardiac myopathy, and other adverse effects. These studies indicate that the adverse maternal effects observed with fluvastatin before or following parturition resulted from exaggerated pharmacologic activity at the dose levels administered, i.e., inhibition of the enzyme HMG-CoA reductase, its immediate product mevalonic acid, and cholesterol biosynthesis.

Reversal of the adverse chronic effects of the unsaturated derivative of valproic acid--2-n-propyl-4-pentenoic acid--on ketogenesis and liver coenzyme A metabolism by a single injection of pantothenate, carnitine, and acetylcysteine in developing mice.

Like treatment with the parent compound valproic acid (VPA), acute and/or chronic treatment with the unsaturated derivative, 2-n-propyl-4-pentenoic acid (4-en-VPA), decreased ketogenesis and lowered free CoA, acetyl CoA, and free carnitine levels in the livers of normal developing mice. Concomitantly, there were manifold increases in the content of medium-chain acyl CoA esters (4-en-VPA CoA and 4-en-VPA CoA metabolites). Acute cotreatment of 4-en-VPA-treated animals with pantothenate, carnitine, and acetylcysteine caused significant amelioration of these metabolic aberrations. In animals chronically treated with 4-en-VPA, a single injection of pantothenate, carnitine, and acetylcysteine returned the 4-en-VPA-depressed levels of beta-hydroxybutyrate in plasma and free CoA and acetyl CoA in liver to normal. These findings support the hypothesis that VPA- and 4-en-VPA-induced hepatic dysfunction is produced by CoA sequestration rather than by irreversible inhibition by alkylation of the enzymes of fatty acid beta-oxidation by reactive intermediates. The findings also support the important but little-known role of carnitine in CoA metabolism--carnitine relieves the inhibition of pantothenate kinase, the rate-controlling first enzyme in the pathway of CoA synthesis by its product, free CoA, and by CoA esters.

Fumaraldehydic acid-induced inactivation of aspartase.

Fumaraldehydic acid (FAA) induced a time-dependent inactivation of aspartase (L-aspartate ammonia-lyase, EC 4.3.1.1) from Escherichia coli at 30 degrees C and pH 7.4 following pseudo-first order kinetics. The rate of inactivation increased in proportion to the FAA concentration. In addition, the rate of inactivation increased, as the pH was increased. Determination of sulfhydryl groups showed that approximately one among 11 sulfhydryl groups was modified by FAA concomitant with the inactivation. L-Aspartate and fumarate protected the enzyme against FAA-inactivation, when Mg2+ ions were present. Unlike E. coli aspartase, P. fluorescens aspartase was not inactivated by FAA.