Sulcotrione soil metabolism in summer corn crops.
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Belgium↗
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Store operated Ca2+ influx by selective depletion of ryanodine sensitive Ca2+ pools in primary human skeletal muscle cells.
The contraction and relaxation of skeletal muscle is driven by release of Ca2+ from sarcoplasmic reticulum through the ryanodine receptor type 1 and extruding the ion from the cytosol by Ca2+ ATPases. Efficient refilling of the empty Ca2+ stores is essential for repetitive cycles of muscle contraction and relaxation, but not investigated in human skeletal muscle cells. Here we show that under conditions of selective depletion of the ryanodine-sensitive Ca2+ pool Ca2+ influx occurs in differentiated human skeletal muscle cells using the Ca2+ imaging technique. This Ca2+ influx is not due to permeation through the L-type Ca2+ channel and not observed under conditions of inhibited Ca2+ ATPase. The Ca2+ influx was visualised by quenching the intracellular fura2 signal with Mn2+ on single cell level and also using fluorescence photometry of cell suspensions. The Mn2+ influx was inhibited by the Ca2+ channel blockers La(3+) and SKF96356. The delineation of the signalling cascade leading to Ca2+ influx evoked by selective depletion of ryanodine sensitive Ca2+ stores showed that phospholipase C or protein kinase C were not involved. Interestingly, a Mn2+ influx was triggered by the cell-permeant analogue of diacylglycerol and further augmented by the application of RHC80267, a diacylglycerol lipase inhibitor. This signalling pathway could be attributed to the participation of a protein kinase C activity. However, Mn2+ influx evoked by selective depletion of ryanodine sensitive Ca2+ stores was not altered by RHC80267 or protein kinase C inhibitors. Using RT-PCR, correctly spliced mRNA fragments were detected corresponding to human transient receptor potential (TRPC) Ca2+ channels type 1, 3, 4 and 6. These data show that in skeletal muscle at least two independent mechanisms of Ca2+ influx exist. For Ca2+ influx triggered by the selective depletion of ryanodine sensitive Ca2+ stores we propose a phospholipase C independent coupling of ryanodine receptors to voltage insensitive Ca2+ channels.
Caffeine↗
In situ product recovery (ISPR) by crystallization: basic principles, design, and potential applications in whole-cell biocatalysis.
The removal of inhibiting or degrading product from a bioreactor as soon as the product is formed is an important issue in industrial bioprocess development. In this review, the potential of crystallization as an in situ product removal (ISPR) technique for the biocatalytic production of crystalline compounds is discussed. The emphasis of this review is on the current status of crystalline product formation by metabolically active cells for application in fine-chemicals production. Examples of relevant biocatalytic conversions are summarized, and some basic process options are discussed. Furthermore, a case study is presented in which two conceptual process designs are compared. In one process, product formation and crystallization are integrated by applying ISPR, whereas a second, nonintegrated process is based on a known conventional process equivalent for the production of 6R-dihydro-oxoisophorone. The comparison indicates that employing ISPR leads to significant advantages over the nonintegrated case in terms of increased productivity and yield with a corresponding decrease in the number of downstream processing steps, as well as in the quantity of waste streams. This leads to an economically more interesting process alternative. Finally, a general outlook on the various research aspects of ISPR by crystallization is given.
Biological Products↗
New developments in ochronosis: review of the literature.
Ochronosis commonly affects all connective tissue. Recognition of changes secondary to the deposition of ochronotic pigments has increased with advances in diagnostic technology, allowing both improved imaging and early biochemical and genetics-based diagnosis of alkaptonuria, the cause of ochronosis. Successful symptomatic treatment of ochronotic arthropathy with joint replacement has been documented, and a new pharmacotherapeutic agent, nitisinone, is currently under investigation for both prevention and treatment of ochronosis. This review of the literature highlights recently recognized complications, new diagnostic techniques, and treatment options.
Alkaptonuria↗
The determination of firing distance applying a microscopic quantitative method and confocal laser scanning microscopy for detection of gunshot residue particles.
In this study, we applied a microscopic quantitative method based on the use of sodium rhodizonate to verify the presence of residues and their distribution on the cutis of gunshot wounds. A total of 250 skin samples were selected from cases in which the manner of death (accidental, suicide, and homicide) and the shooting distance could be reliably determined. The samples were examined under a light microscope, in transmitted bright field illumination and phase contrast mode, and with confocal laser scanning microscopy. In all skin specimens the area of each histological section was directly measured by an image analysis system. Both the number and the size of powder particles were measured. The distribution of gunshot residues (GSR) in the epidermal and subepidermal layers was also analyzed. The evaluation of the microscopic entrance wounds demonstrated different findings related to the range of fire. The data derived from the evaluation of both macroscopic and microscopic features demonstrated that the amount and the spatial distribution of GSR deposits in the skin surrounding entrance wounds strictly correlate with shooting distance.
Case-Control Studies↗
Determination of firing distance using the rhodizonate staining technique.
The histological staining technique using rhodizonate is also effective for the determination of the firing distance by examining the distribution and intensity of the staining reaction. The differentiation between absolute close-range shots and long-range shots is generally possible without any doubt. The method is not recommended for routine examinations but it is very useful for cases lacking the possibility to investigate smoke and powder deposits in a criminalistic manner, i.e. surgical skin biopsies of hospitalised victims and skin highly altered by the effects of fire, water or by post-mortem decomposition.
Coloring Agents↗
Effect of chronic hypoxia on voltage-independent calcium influx activated by 5-HT in rat intrapulmonary arteries.
5-Hydroxytryptamine (5-HT) is a potent pulmonary vasoconstrictor and mitogenic agent whose concentration increases in pulmonary hypertensive patients. Chronic hypoxia induces selective pulmonary arterial hypertension; therefore, we investigated chronic hypoxia effect on the calcium and contractile responses to 5-HT focusing on voltage-independent calcium influx in rat intrapulmonary arteries. Chronic hypoxia, induced by introducing rats in a hypobaric chamber for 3 weeks, potentiated the contraction to 5-HT and this effect was insensitive to nitrendipine. Calcium signal to 5-HT was characterized by a transient followed by a sustained phase in both normoxia and chronic hypoxia. The sustained phase was dependent on extracellular calcium and inhibited by lanthanum. RHC 80267, a specific inhibitor of diacylglycerol lipase, reduced the 5-HT-induced calcium influx in chronic hypoxia but not in normoxia. Furthermore, unlike gadolinium, RHC 80267 inhibited more the contraction to 5-HT in chronic hypoxia. Despite the apparent role of voltage-independent calcium channels in chronic hypoxia, Western blot and flow cytometry analyses demonstrated no variations in TRPC(6) expression. This study shows for the first time that the 5-HT-induced calcium and contractile signals in chronic hypoxia are more dependent on a voltage-independent, RHC 80267-sensitive calcium influx and the hyperreactivity to 5-HT may thus be explained by this influx.
Animals↗
Formation of benzoquinol moiety in cornoside by salidroside mono-oxygenase, a cytochrome P450 enzyme, from Abeliophyllum distichumcell suspension cultures.
A microsomal fraction prepared from Abeliophyllum distichumNakai (Oleaceae) cell suspension cultures oxidized salidroside, a glucoside of 4-hydroxyphenylethyl alcohol, to cornoside possessing a unique benzoquinol ring. The enzyme named salidroside mono-oxygenase required NADPH as the only cofactor, and molecular oxygen. The reaction was strongly inhibited by CO as well as several cytochrome P450 inhibitors, such as cytochrome c and miconazole, indicating the involvement of a cytochrome P450 enzyme. Salidroside mono-oxygenase accepted salidroside as the only substrate, but did not oxidize 4-hydroxyphenylethyl alcohol, the salidroside aglucone, and 4-hydroxybenzoic acid. The optimum pH of the reaction was 7.5, and apparent K(m) values for salidroside and NADPH were 44 micro M and 33 micro M, respectively. The benzoquinol ring formation mechanism is discussed in comparison to the mechanism for ipso substitution of 4-hydroxybenzoate by active oxygen species followed by elimination leading to hydroquinone.
Benzoquinones↗
An isoleucine-leucine substitution in chloroplastic acetyl-CoA carboxylase from green foxtail (Setaria viridis L. Beauv.) is responsible for resistance to the cyclohexanedione herbicide sethoxydim.
The cDNAs encoding chloroplastic acetyl-CoA carboxylase (ACCase, EC 6.4.1.2) from three lines of Setaria viridis (L. Beauv.) resistant or sensitive to sethoxydim, and from one sethoxydim-sensitive line of Setaria italica (L. Beauv.) were cloned and sequenced. Sequence comparison revealed that a single isoleucine-leucine substitution discriminated ACCases from sensitive and resistant lines. Using near-isogenic lines of S. italica derived from interspecific hybridisation, we demonstrated that the transfer of the S. viridis mutant ACCase allele into a sethoxydim-sensitive S. italica line conferred resistance to this herbicide. We confirmed this result using allele-specific polymerase chain reaction and showed that a single copy of the mutant allele is sufficient to confer resistance to sethoxydim. We conclude that a mutant allele of chloroplastic ACCase encoding a leucine residue instead of an isoleucine residue at position 1780 is a major gene of resistance to sethoxydim.
Acetyl-CoA Carboxylase↗
Cutaneous porphyria in a neonate with tyrosinaemia type 1.
UNLABELLED: A term infant born to consanguineous parents presented at birth with hypoglycaemia, thrombocytopenia, coagulopathy and hyperbilirubinaemia associated with polycythaemia due to delayed cord clamping. Despite phototherapy and correction of polycythaemia by partial exchange transfusion, coagulopathy, hypoglycaemia and conjugated hyperbilirubinaemia persisted, suggesting hepatic failure. Metabolic work-up led to the diagnosis of tyrosinaemia type 1 on day 4. Two--(2-nitro-4-trifluoromethylbenzoyl)--1,3 cyclohexanedione (NTBC) treatment, started on day 5, resulted in progressive clinical improvement and unambiguous biochemical response. Severe skin purpuric lesions occurred in areas exposed to phototherapy. These resolved slowly after its discontinuation. Urine analysis sampled just before and 6 days after starting NTBC treatment showed high levels of type 1 coproporphyrin isomers. Such findings do not seem directly related to tyrosinaemia type 1 where succinylacetone inhibits delta-aminolevulinic acid (delta-ALA) dehydratase and where the accumulation of delta-ALA results in neurotoxicity without photosensitivity. CONCLUSION: We describe a cutaneous form of porphyria in a neonate presenting with severe liver failure due to tyrosinaemia type 1. This porphyria is tentatively attributed to a secondary accumulation of coproporphyrins due to cholestasis, as reported in the bronze baby syndrome and recently described in neonates with purpuric phototherapy-induced eruption, rather than to a primary defect of porphyrin metabolism. The hypothesis of a direct effect of tyrosinaemia type 1 on porphyrin excretion is also discussed.
Combined Modality Therapy↗
Kinetics of chlorination of monochlorodimedone by myeloperoxidase.
The phagocyte-derived enzyme myeloperoxidase has been recently implicated in the pathogenesis of atherosclerosis, because it catalyzes the reaction of hydrogen peroxide with chloride ions to give the highly toxic oxidant hypochlorous acid. The aim of this study was to determine the dependence of this reaction on the concentration of hydrogen peroxide and of the enzyme by means of the photometric monochlorodimedone assay. The initial rate of hypochlorous acid formation increased less than proportionally with increasing myeloperoxidase concentrations. Variation of the concentration of hydrogen peroxide had a biphasic effect, with an optimal concentration of hydrogen peroxide. Above this concentration enzyme destruction is apparently predominant. The progress curves of hypochlorous acid formation showed two distinct maxima. It was concluded that hypochlorous acid not only reacts with monochlorodimedone but also with the amino groups of myeloperoxidase to form intermediary chloramines that may further chlorinate monochlorodimedone. This was supported by the kinetics in the presence of the amino compound glycine, a competitive substrate for chlorination by hypochlorous acid. In the presence of high concentrations of glycine the progress curve rises continuously, yielding a greatly increased concentration of chlorinating species, either hypochlorous acid or chloramines. We concluded that glycine protects myeloperoxidase against hypochlorous acid-induced self-destruction.
Binding, Competitive↗
A novel therapeutic trial of homogentisic aciduria in a murine model of alkaptonuria.
Alkaptonuria is a rare autosomal recessive disorder characterized by homogentisic aciduria, ochronosis, and arthritis. Although a deficiency of homogentisic acid 1,2-dioxygenase has recently been confirmed at the molecular level, no effective treatment regimen has yet been developed for this disorder. In the present study, 2(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione (NTBC), a potent inhibitor of p-hydroxyphenylpyruvate dioxygenase (which catalyzes the formation of homogentisic acid from p-hydroxyphenylpyruvic acid) was adopted as a possible therapeutic agent for alkaptonuria. NTBC dose-dependently reduced the urinary output of homogentisic acid in a murine model of alkaptonuria that had been created with ethylnitrosourea. These findings suggest that NTBC may be the first potent pharmacotherapeutic agent for alkaptonuria.
4-Hydroxyphenylpyruvate Dioxygenase↗
Characterization of the lipolytic pathways that mediate free fatty acid release during Fas/CD95-induced apoptosis.
We have undertaken a study to characterize the lipolytic pathway responsible for the generation of free fatty acids (FFA) during Fas/CD95-induced apoptosis in Jurkat cells. It was initially shown that the cellular lipid fraction that suffered the major quantitative decrease during Fas-induced apoptosis was that of phosphatidylcholine (PC). In addition, the secretion of palmitic acid-derived FFA was largely prevented by D609, an inhibitor of PC-specific phospholipase C (PC-PLC) and also by the diacylglycerol lipase (DAGL) inhibitor RHC-80267, suggesting that the secretion of these FFA during Fas-induced apoptosis is mediated by the generation of DAG by a PC-PLC activity and, sequentially, by a 1-DAGL activity which generates the FFA from its sn-1 position. The endocannabinoid 2-arachidonoyl glycerol (2-AG) should be generated as a sub-product of this pathway, but it did not accumulate inside the cells nor was secreted into the supernatant. Interestingly, the complete inhibition of free AA secretion during Fas-induced apoptosis was only achieved by using the AA trifluoromethylketone, which not only inhibits all types of phospholipase-A(2) (PLA(2)) activities, but also the described lytic activities on 2-AG. Using a combination of RHC-80267 and the iPLA(2)-specific inhibitor bromoenol lactone, it was shown that the DAGL pathway also cooperates with iPLA(2) in the generation of free arachidonate.
Apoptosis↗
Biotransformation of racemic diisophorone by Cephalosporium aphidicola and Neurospora crassa.
Racemic diisophorone (500 mg) was converted by Cephalosporium aphidicola and Neurospora crassa over 10 days at 25 degrees C to 8beta-hydroxydiisophorone in yields of 10% (52 mg) and 20% (103 mg), respectively. The structure was established by IR, specific rotation, mass spectral, 1D and 2D-NMR studies.
Acremonium↗
Successful treatment of severe cardiomyopathy with NTBC in a child with tyrosinaemia type I.
We report the case of a child who developed severe obstructive hypertrophic cardiomyopathy revealing hereditary tyrosinaemia type I, who was successfully treated with NTBC. The mechanisms underlying the association are discussed.
Amino Acid Metabolism, Inborn Errors↗
Cardiomyopathy in tyrosinaemia type I is common but usually benign.
UNLABELLED: Tyrosinaemia type I (TTI) is an inherited multisystemic disorder of tyrosine metabolism. In addition to hepatic and renal involvement, cardiomyopathy is an important clinical manifestation. OBJECTIVE: To evaluate the incidence and outcome of cardiomyopathy in TTI. SUBJECTS AND METHODS: A retrospective study was performed of 20 consecutive children with TTI (12 male, 8 female) referred to a single centre between 1986 and 2002. All were initially treated with standard dietary therapy and, since 1992, with nitisinone. The indications for orthotopic liver transplantation (LT) changed during the study. Serial echocardiography was undertaken in all subjects. RESULTS: 9/20 (45%) children had an acute hepatic presentation. Five (25%) received dietary treatment followed by LT, and 14 (70%) were treated with nitisinone at presentation. 6/20 (30%) had cardiomyopathy at initial assessment, with interventricular septal hypertrophy being the commonest finding (5/6). Cardiomyopathy was significantly less common in those treated initially with nitisinone. After a median follow-up of 3.6 (0.45-13.5) years, 5/6 (83%) had complete resolution of cardiomyopathy and 1/6 showed significant improvement. No child with a normal initial echocardiography subsequently developed cardiomyopathy. CONCLUSION: Cardiomyopathy is a common manifestation of TTI and it has a favourable long-term outcome. Children initially treated with nitisinone are less likely to develop this complication.
Adolescent↗
Chemical response of Picea glehnii seed-epiphytic Penicillium species to Pythium vexans under in vitro competitive conditions for mycelial growth.
The potential protection of Picea glehnii seedlings from damping-off by seed-epiphytic Penicillium species was investigated. We studied the chemical response of seed-epiphytic Penicillium species (Pen. cyaneum, Pen. damascenum, and Pen. implicatum) to Pythium vexans, a damping-off fungus, in vitro. Penicillium species were cultured singly or cocultured with Pyt. vexans for 14 or 18 d, and mycelial growth, pH of culture filtrate, antifungal activity of the culture filtrate against Pyt. vexans, and the amount of antifungal compound produced by each Penicillium species, were examined. The filtrate of both the single culture of Penicillium and the coculture of Penicillium and Pyt. vexans showed antifungal activity against Pyt. vexans. In a coculture with Pyt. vexans, Pen. cyaneum produced an antifungal compound (patulin) as in the single culture. Pen. damascenum cocultured with Pyt. vexans produced an antifungal compound (citrinin), as it did in the single culture and in larger amounts on day 10. Pen. implicatum produced two antifungal compounds, frequentin and palitantin, and the ratio of frequentin (with higher antifungal activity than palitantin) to palitantin was higher in the coculture with Pyt. vexans than in the single culture. Our results indicate that these Penicillium species have the ability to produce antifungal compounds and to keep anti-fungal activity under competitive condition with Pyt. vexans. The chemical response of these Penicillium species to Pyt. vexans may contribute to protect P. glehnii seedlings from damage by Pyt. vexans.
Antifungal Agents↗
Fatty acid synthetase assay employing bicyclic diones as substrates.
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Animals↗