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At least 757 records · Page 42Linked to original sources
PubMed2000

Antioxidants protect against dopamine-induced metallothionein-III (GIF) mRNA expression in mouse glial cell line (VR-2g).

Metallothionein (MT)-III, originally discovered as a growth inhibitory factor (GIF), is a brain specific isomer of MTs and is markedly reduced in the brain of patients with Alzheimer's disease (AD) or other neurodegenerative diseases. We analyzed the level and regulation of mRNA expression of MT-III in immortalized fetal mouse brain glial cells (VR-2g) by reverse transcriptase-polymerase chain reaction (RT-PCR). We have recently reported that dopamine (DA) increases the expression of MT-III mRNA in vitro. In this study, we investigated the mechanism of such increase by examining the effects of DA agonists (SKF38393 or bromocriptine) and DA antagonists (SCH23390 or sulpiride) on the expression of MT-III mRNA. MT-III mRNA did not change by either agonist and DA-increased MT-III mRNA was not inhibited by either antagonist. These results suggested that the induction of MT-III mRNA by DA was not mediated by stimulation of DA receptors. On the other hand, DA-induced MT-III mRNA expression was strongly inhibited by the addition of antioxidants (glutathione, vitamin E or ascorbic acid), indicating that DA-enhanced MT-III mRNA was mediated by reactive oxygen species. Our results suggest that oxidative stress may be one of the principle factors that modulate MT-III mRNA expression.

Adrenergic Agents↗
PubMed1997

Effect of antispermatogenic compound CDRI-84/35 on marker enzymes of rat testis cells. A study on site of action.

Marker enzymes of Sertoli and germ cells were estimated to study the mechanism of action of antispermatogenic compound CDRI 84/35 in adult male rat testis. Animals were killed after 22, 41, and 64 days of treatment with antispermatogenic dose of CDRI 84/35 in order to evaluate the effect of the compound on spermatid, spermatocyte, and spermatogonial stages, respectively. Studies were also extended to a recovery period of 90 days. Results indicate a direction action of the compound on germ cells, with no apparent effect on Sertoli cells. Studies also show a massive depletion of postmeiotic germ cells after the treatment, with some damage to premeiotic germ cells as well. Reversibility of the compound was partial, with the marker enzymes of pre- and postmeiotic germ cells not being restored to control levels after withdrawal of treatment.

Animals↗
PubMed2003

Commitment of 3T3-F442A cells to adipocyte differentiation takes place during the first 24-36 h after adipogenic stimulation: TNF-alpha inhibits commitment.

We studied the commitment of 3T3-F442A cells during stimulation with adipogenic serum or growth hormone. Confluent 3T3-F442A preadipocytes were incubated with adipogenic medium for increasing times; the number of adipose clusters, GPDH activity, and lipid accumulation were evaluated. Results show that cell commitment took place during the first 24-36 h after stimulation under adipogenic conditions. Then, cultures underwent a 2-fold increase in total cell number through selective multiplication of committed cells, followed by a dramatic decrease in colony-forming ability and 300- to 1000-fold raise in GPDH activity. Cell commitment was not modulated by insulin, but this hormone stimulated clonal expansion of committed cells and lipogenesis. Commitment was inhibited by TNF-alpha at concentrations as low as 5 ng/ml, and by retinoic acid. The results show that TNF-alpha inhibits adipose conversion at two different levels: at concentrations as low as 5 ng/ml, it blocks commitment, and at concentrations of 100 ng/ml or higher the cytokine seems to block mitotic expansion and other steps of differentiation after cell commitment. The identification of a specific time for cell commitment would allow the study of the early genes that might regulate cell reprogramming into adipocytes.

3T3 Cells↗
PubMed1997

Population genetic structure of Ixodes ricinus in Switzerland from allozymic data: no evidence of divergence between nearby sites.

Ixodes ricinus is a vector and reservoir of numerous infectious agents, especially Borrelia burgdorferi, the agent of Lyme disease. In Switzerland, its ecology and physiology have been well studied. Moreover, the foci of some infectious agents transmitted by this tick are identified. They can display relatively to extremely small geographical size depending on the diseases considered. In order to understand how the gene flows occur and to characterise the genetic structure of Ixodes ricinus populations, we used an indirect method based on genetic markers: allozymes. The sampling was carried out in 5 localities. Eighteen loci were analysed and 2 appeared polymorphic. This shows the low allozymic variability displayed by Ixodes ricinus. Based on these 2 loci, the populations appeared panmictic in Switzerland. This may be explained by the wide range of vertebrate species this tick can infest, especially birds. However, the result is surprising if we consider the extreme localisation of the foci of some infectious agents. We conclude that more powerful genetic markers could be used in order to better understand the epidemiology of tick-borne diseases in Switzerland.

Alleles↗
PubMed1985

The adult human female urethra. Enzyme-histochemical study.

In the female urethra, the activity and distribution of 15 enzymes was determined by using both conventional and special histochemical methods. The enzymatic equipment differed according to the type of epithelial lining whose variation is characteristic for the female urethra. In the stratified squamous epithelium of the urethra, alkaline phosphatase, beta-glucuronidase, acetyl-beta-D-glucosaminidase, thiamine pyrophosphatase, and glucose-6-phosphatase exhibited but minimal or no activity, yet the other 10 enzymes studied displayed activity particularly in basally situated cells. Nearer to the lumen of the urethra, the activity in the epithelium kept decreasing and was mostly absent in superficial and desquamated cells. In the pseudostratified columnar and in the transitional epithelium of the urethra, the majority of enzymes showed an evenly distributed activity at all epithelial levels. In the apical parts of the most superficially situated cells bounding the lumen of the urethra, a distinct narrow zone of higher activity was observed. It was seen not only on determining the majority of dehydrogenases but also on examining acid phosphatase and naphthyl esterase. The endocrine cells occurring in the uroepithelial lining of the female urethra displayed, yet always with the exception of squamous epithelium (Zaviacic et al. 1983), distinct activity of acid hydrolytic enzymes, and of the enzymes studied it was particularly acid phosphatase. The majority of the demonstrated enzymes, of the dehydrogenases priority, is to be given to succinate dehydrogenase, enabled to differentiate readily between the highly active striated muscle fibers located in the most peripheral parts of the excisions along the urethral circumference and the smooth musculature of the urethral wall with a lower or only minimal activity.

Acetylglucosaminidase↗
PubMed1996

Changes in eosinophil and leukocyte infiltration and expression of IL-6 and IL-7 messenger RNA in mite allergen patch test reactions in atopic dermatitis.

Patch testing with crude dust mite extracts after removal of homy layers was performed on normal-appearing skin of 11 adult patients with atopic dermatitis and high mite-specific IgE antibody scores. Positive skin reactions were observed in 9 subjects. Skin biopsy specimens were obtained from positive reaction sites at 2, 6, 12, 24, and 48 hours after allergen challenge and subjected to histologic studies and extraction of messenger RNA (mRNA). Perivascular infiltration of small mononuclear cells began at 2 hours and was followed by eosinophilic infiltration at 6 hours, and the number of eosinophils continued to increase at 24 and 48 hours. In addition to the increased expression of IL-4, IL-5, and tumor necrosis factor-alpha mRNA during the time course detected by reverse-transcription polymerase chain reaction, mRNA of IL-6 and IL-7 was also up-regulated. After the removal of test patches with mite allergen, the number of eosinophils started to decrease in a time-dependent manner. Histopathologic findings at 48 hours after removal showed lymphocyte-dominant infiltration intermingled with occasional eosinophils. These mite allergen patch test reactions may provide a useful model for studying the pathogenesis of atopic eczema, especially with regard to the initiation of eosinophil infiltration and the alternative increase in lymphocytes.

Adult↗
PubMed1998

Nucleotide sequence and expression of the sn-glycerol-3-phosphate dehydrogenase gene in Locusta migratoria.

The sn-glycerol-3-phosphate dehydrogenase gene (Gpdh) in the locust (Locusta migratoria) encodes three mature transcripts and a number of isozymes. Gpdh expression is tissue- and developmentally regulated such that two transcripts are unique to flight muscle. Identical proteins are encoded by two transcripts which are generated by alternative splicing downstream of the stop codon in the penultimate exon.

Animals↗
PubMed2003

ACTH and alpha-MSH inhibit leptin expression and secretion in 3T3-L1 adipocytes: model for a central-peripheral melanocortin-leptin pathway.

Leptin is the 167 amino-acid protein product of the Lep (obese) gene that is released predominantly from adipose tissue and circulates at levels related to the amount of fat. Leptin expression is hormonally regulated: insulin and glucocorticoids are stimulators, while inhibitors include beta-adrenergic agonists and testosterone. Recently, adenylate cyclase-coupled melanocortin receptors have been identified in murine adipose tissue, the 3T3-L1 adipocyte cell line, and in human fat tissue. These studies prompted us to evaluate the effects of pro-opiomelanocortin (POMC)-derived peptides on leptin production and expression in 3T3-L1 adipocytes in culture. 3T3-L1 pre-adipocytes differentiated by the insulin/indomethacin (I/I) method produced leptin at levels that were two times higher than those obtained in cells differentiated by the more traditional insulin/dexamethasone/isobutylmethylxanthine (I/D/M) method. By RT-PCR studies, 3T3-L1 cells expressed both the melanocortin 2 receptors (MC2-R) and melanocortin 5 receptors (MC5-R) isoforms of the melanocortin receptor at an early stage of differentiation. When I/I differentiated 3T3-L1 adipocytes were incubated with different concentrations of dibutyryl cAMP (db-cAMP) or POMC-derived peptides (ACTH and alpha-MSH), ACTH and alpha-MSH stimulated cAMP production after 30 min (2-fold increase) associated with a dose-dependent inhibition of leptin secretion (ACTHz.Gt;alpha-MSH; IC(50)=3.2+/-0.4 SE and 36+/-5 nM, respectively), maximal after 3 h of incubation (30% inhibition). In addition, 100 nM ACTH and alpha-MSH induced a 60% reduction in leptin expression by RT-PCR. Incubation of cells with 0.5 mM db-cAMP led to a more prominent inhibition of leptin expression and secretion (up to 80% at 1 and 24 h, respectively). The ACTH and alpha-MSH inhibitory effects on leptin secretion were mediated by activation of the MC2-R and MC5-R and were reversed by the MC-R antagonists ACTH(11-24) and ACTH(7-38). In summary, we have shown that POMC-peptides are potent inhibitors of leptin expression and production in 3T3-L1 adipocytes. The finding of ACTH/alpha-MSH receptor-induced inhibition of leptin production and expression in adipocytes support the possibility that there is a control mechanism for modulation of adipose tissue function via a melanocortin-leptin axis.

3T3-L1 Cells↗
PubMed2001

Histamine N-methyltransferase regulates histamine-induced phosphoinositide hydrolysis in guinea pig cerebellum.

We report here that the dose-response curve of the histamine-stimulated phosphoinositide hydrolysis in the guinea pig cerebellar slices was shifted to the left when the slices were pretreated with SKF 91488 (100 microM), a specific inhibitor of histamine N-methyltransferase (HMT). In contrast, the pretreatment of the cerebellar slices with aminoguanidine (100 microM - 1 mM), an inhibitor of diamine oxidase, had no effect on histamine-induced phosphoinositide hydrolysis. HMT mRNA was expressed abundantly in cerebellum, especially in Purkinje cells. These observations suggest that HMT regulates histaminergic neurotransmission in guinea pig cerebellum more predominantly than diamine oxidase in histamine degradation.

Animals↗
PubMed2003

Red fluorescent protein (DsRed2) as a novel reporter in Fusarium oxysporum f. sp. lycopersici.

pAn-DsRed2 vector was constructed for constitutive cytoplasmic expression of the red fluorescent protein (DsRed2) under control of the glyceraldehyde-3-phosphate dehydrogenase gene promoter from Aspergillus nidulans. DsRed2-transformation of two Fusarium oxysporum f. sp. lycopersici strains pathogenic against tomato host resulted in bright red cytoplamic fluorescence of the fungus. The transformants were screened based on the hygromycin B resistance, brightness, stability and rate of appearance of the DsRed2 fluorescence. The transormed fungi were growing normally and their pathogenicity did not change after transformation procedure. The function of novel DsRed2 marker was verified by fluorescence microscopy of the infected tomato seedlings. The results indicate that DsRed2 can be used as a efficient novel reporter gene for monitoring of the F. oxysporum within the host tissues.

Aspergillus nidulans↗
PubMed2002

Changes in gene expression in rat thymocytes identified by cDNA array support the occurrence of oxidative stress in early magnesium deficiency.

Magnesium deficiency in experimental animals leads to inflammation, exacerbated immune stress response and a decrease of specific immune response. It also results in a significant increase in free radical species and subsequent tissue injury. An accelerated thymus involution was observed in Mg-deficient rats in relation to enhanced apoptosis and enhanced susceptibility to oxidative stress. To examine the stress-inducing effects of low Mg status on thymocytes, cDNA arrays were used to evaluate changes in gene expression in weaning rats submitted to Mg deficiency of short duration (2 days). Several genes exhibited changes in their expression caused by Mg deficiency before any perceptible modification in cell integrity and functions. The up-regulated genes included cytochrome c oxidase, glutathione transferase, CuZn superoxide dismutase, genes associated with the stress response (HSP70 and HSP84) and a gene involved in DNA synthesis and repair (GADD45). The down-regulated genes included Na/P cotransporter 1. These findings are consistent with altered cell growth, modifications of ion fluxes and oxidative stress described during Mg deficiency. The observation of induction of genes involved in protection and repair in cells from Mg-deficient animals provides additional evidence of the role of oxidative stress in the pathobiology of this deficiency.

Animals↗
PubMed2000

A potential target enzyme for trypanocidal drugs revealed by the crystal structure of NAD-dependent glycerol-3-phosphate dehydrogenase from Leishmania mexicana.

BACKGROUND: NAD-dependent glycerol-3-phosphate dehydrogenase (GPDH) catalyzes the interconversion of dihydroxyacetone phosphate and L-glycerol-3-phosphate. Although the enzyme has been characterized and cloned from a number of sources, until now no three-dimensional structure has been determined for this enzyme. Although the utility of this enzyme as a drug target against Leishmania mexicana is yet to be established, the critical role played by GPDH in the long slender bloodstream form of the related kinetoplastid Trypanosoma brucei makes it a viable drug target against sleeping sickness. RESULTS: The 1.75 A crystal structure of apo GPDH from L. mexicana was determined by multiwavelength anomalous diffraction (MAD) techniques, and used to solve the 2.8 A holo structure in complex with NADH. Each 39 kDa subunit of the dimeric enzyme contains a 189-residue N-terminal NAD-binding domain and a 156-residue C-terminal substrate-binding domain. Significant parts of both domains share structural similarity with plant acetohydroxyacid isomeroreductase. The discovery of extra, fatty-acid like, density buried inside the C-terminal domain indicates a possible post-translational modification with an associated biological function. CONCLUSIONS: The crystal structure of GPDH from L. mexicana is the first structure of this enzyme from any source and, in view of the sequence identity of 63%, serves as a valid model for the T. brucei enzyme. The differences between the human and trypanosomal enzymes are extensive, with only 29% sequence identity between the parasite and host enzyme, and support the feasibility of exploiting the NADH-binding site to develop selective inhibitors against trypanosomal GPDH. The structure also offers a plausible explanation for the observed inhibition of the T. brucei enzyme by melarsen oxide, the active form of the trypanocidal drugs melarsoprol and cymelarsan.

Animals↗
PubMed2003

Outlines for the definition of halotolerance/halophily in yeasts: Candida versatilis (halophila) CBS4019 as the archetype?

Candida versatilis (halophila) CBS4019 was chosen to study the physiological reactions of long-term exposure to extremely high salt concentrations. In general, our results show a significant increase in enzyme expression during growth under stress conditions. Although glycerol and mannitol pathways are not under glucose repression, they were found to be metabolically regulated. Glycerol-3P-dehydrogenase used either of its cofactors NADPH or NADH, being in favor of NADPH during growth with high salt concentrations. This ability of interchanging cofactors, an increased fermentation rate, and the observed mannitol pathway activity are suggested to contribute to the yeasts' redox stability. Enzymes per se were not salt-tolerant in vitro. Consistently, intracellular sodium was low and intracellular potassium, a requirement for growth, was high. The concept of halophily and its applicability to yeasts is discussed.

Alcohol Dehydrogenase↗
PubMed2001

Survey of normal appearing mouse strain which lacks malic enzyme and Nad+-linked glycerol phosphate dehydrogenase: normal pancreatic beta cell function, but abnormal metabolite pattern in skeletal muscle.

We studied a mouse doubly homozygous for mutations in the genes encoding malic enzyme (EC 1.1.1.40) and cytosolic glycerol phosphate dehydrogenase (EC 1.1.1.8) (cGPD). This mouse, which we call the mmgg mouse and which is the product of intercrosses between the Mod-1 mouse and the BALB/cHeA mouse, lacks activity of both enzymes. Like both parental strains the mmgg mouse is completely normal in appearance. cGPD is one of the two enzymes that catalyze the reactions of the glycerol phosphate shuttle. The activity of the other enzyme of the glycerol phosphate shuttle, mitochondrial glycerol phosphate dehydrogenase (EC 1.1.99.5) (mGPD), is abundant in tissues, such as brain, skeletal muscle and the pancreatic islet, suggesting that the glycerol phosphate shuttle is important in these tissues which rapidly metabolize glucose. Cytosolic malic enzyme activity is important for shuttles which transport NADPH equivalents from mitochondria to the cytosol. The major finding of the study was a highly abnormal metabolite pattern in tissues of the mmgg mouse suggesting a block in the glycerol phosphate shuttle due to cGPD deficiency. The metabolite pattern did not suggest that malic enzyme deficiency caused an abnormality. Tissue levels of glycerol phosphate (low) and dihydroxyacetone phosphate (high) were only abnormal in skeletal muscle. Glycolytic intermediates, situated at or before the triose phosphates in the pathway, such as fructose bisphosphate and glyceraldehyde phosphate were increased depending on the tissue. Taken together with previous extensive data on the mouse deficient only in cGPD, this suggests a block in glycolysis at the step catalyzed by glyceraldehyde phosphate dehydrogenase caused by an abnormally low NAD/NADH ratio resulting from a nonfunctional glycerol phosphate shuttle. Consistent with this idea the lactate/pyruvate ratio was high in skeletal muscle signifying a low cytosolic NAD/NADH ratio. The mmgg mouse was normal in all other factors studied including blood glucose and serum insulin levels, pancreatic islet mass, insulin release from isolated pancreatic islets, as well as the activities of five metabolic enzymes, including mGPD, in liver, kidney, skeletal muscle and pancreatic islets. cGPD enzyme activity was undetectable in pancreatic islets, 0.5% of normal in liver, and 2.1% of normal in kidney and skeletal muscle. Malic enzyme activity was undetectable in these same tissues.

Animals↗
PubMedDate not supplied

Organization and regulation of the cytosolic NADH metabolism in the yeast Saccharomyces cerevisiae.

Keeping a cytosolic redox balance is a prerequisite for living cells in order to maintain a metabolic activity and enable growth. During growth of Saccharomyces cerevisiae, an excess of NADH is generated in the cytosol. Aerobically, it has been shown that the external NADH dehydrogenase, Nde1p and Nde2p, as well as the glycerol-3-phosphate dehydrogenase shuttle, comprising the cytoplasmic glycerol-3-phosphate dehydrogenase, Gpdlp, and the mitochondrial glycerol-3-phosphate dehydrogenase, Gut2p, are the most important mechanisms for mitochondrial oxidation of cytosolic NADH. In this review we summarize the recent results showing (i) the contribution of each of the mechanisms involved in mitochondrial oxidation of the cytosolic NADH, under different physiological situations; (ii) the kinetic and structural properties of these metabolic pathways in order to channel NADH from cytosolic dehydrogenases to the inner mitochondrial membrane and (iii) the organization in supramolecular complexes and, the peculiar ensuing kinetic regulation of some of the enzymes (i.e. Gut2p inhibition by external NADH dehydrogenase activity) leading to a highly integrated functioning of enzymes having a similar physiological function. The cell physiological consequences of such an organized and regulated network are discussed.

Cytosol↗