Search PubMed⌕ Search

Biomedical subjects

M Marra

Publications and source records attributed to M Marra.

At least 73 records · Page 4Linked to original sources
PubMed1999

Up-regulated EGF receptors undergo to rapid internalization and ubiquitin-dependent degradation in human cancer cells exposed to 8-Cl-cAMP.

8-Cl-cAMP, a cAMP analogue that antagonizes type I cAMP-dependent protein kinase, is a novel anti-tumor agent presently under investigation in clinical trials. Herein we report the effects of this agent on epidermal growth factor receptor expression and degradation in human KB cancer cells. Exposure to 10 microM 8-Cl-cAMP for 48 h induced a 65% increase in epidermal growth factor receptor surface expression while the receptor synthesis was 22-fold enhanced. Analysis of epidermal growth factor-dependent receptor internalization in 8-Cl-cAMP-treated cells showed a higher endocytosis rate as well as an accelerated degradation which occurred together with an increased receptor ubiquitination. The enhanced degradation of epidermal growth factor receptor correlated with the lack of epidermal growth factor-induced proliferation and mitogen-activated protein kinase stimulation. The disregulation of epidermal growth factor receptor internalization and ubiquitin-dependent degradation could underlay a new mechanism of the anti-tumor activity of 8-Cl-cAMP suggesting its combination with agents that disrupt epidermal growth factor receptor signalling.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1999

Meal-induced oxidative stress and low-density lipoprotein oxidation in diabetes: the possible role of hyperglycemia.

Oxidative stress and its contribution to low-density lipoprotein (LDL) oxidation have been implicated in the pathogenesis of vascular diabetic complications. However, the relationship between hyperglycemia, hyperinsulinemia, hyperlipidemia, and oxidative stress is still debated. If plasma glucose and/or insulin and/or lipid are some of the most important determinants of oxidative stress in diabetes, then their typical postprandial elevations in diabetes would be expected to favor oxidative stress and LDL oxidation. To test this hypothesis, in type 2 diabetic patients, we evaluated the effects of two different standard meals designed to produce different levels of postprandial hyperglycemia on the plasma oxidative status and LDL oxidation. The meals were administered in randomized order to each of 10 type 2 diabetic patients. Blood samples were collected at baseline and 60 and 120 minutes after the meals. In every sample, plasma levels of glucose, insulin, cholesterol, triglycerides, nonesterified fatty acids (NEFAs), malondialdehyde (MDA), and the total radical-trapping antioxidant parameter (TRAP) were measured. LDL susceptibility to oxidation was evaluated at baseline and after 120 minutes. Plasma glucose, insulin, triglycerides, and MDA increased and NEFAs and TRAP significantly decreased after either meal. The variations in plasma glucose, MDA, and TRAP were significantly greater and LDL was more susceptible to oxidation after the meal that produced a significantly higher degree of hyperglycemia. These results suggest that postprandial hyperglycemia may contribute to oxidative stress in diabetic patients, providing a mechanistic link between hyperglycemia and diabetic vascular disease.

Cross-Over Studies↗
PubMed1999

zA map for sequence analysis of the Arabidopsis thaliana genome.

Arabidopsis thaliana has emerged as a model system for studies of plant genetics and development, and its genome has been targeted for sequencing by an international consortium (the Arabidopsis Genome Initiative; http://genome-www. stanford.edu/Arabidopsis/agi.html). To support the genome-sequencing effort, we fingerprinted more than 20,000 BACs (ref. 2) from two high-quality publicly available libraries, generating an estimated 17-fold redundant coverage of the genome, and used the fingerprints to nucleate assembly of the data by computer. Subsequent manual revision of the assemblies resulted in the incorporation of 19,661 fingerprinted BACs into 169 ordered sets of overlapping clones ('contigs'), each containing at least 3 clones. These contigs are ideal for parallel selection of BACs for large-scale sequencing and have supported the generation of more than 5.8 Mb of finished genome sequence submitted to GenBank; analysis of the sequence has confirmed the integrity of contigs constructed using this fingerprint data. Placement of contigs onto chromosomes can now be performed, and is being pursued by groups involved in both sequencing and positional cloning studies. To our knowledge, these data provide the first example of whole-genome random BAC fingerprint analysis of a eucaryote, and have provided a model essential to efforts aimed at generating similar databases of fingerprint contigs to support sequencing of other complex genomes, including that of human.

Arabidopsis↗
PubMed1999

A complete BAC-based physical map of the Arabidopsis thaliana genome.

Arabidopsis thaliana is a small flowering plant that serves as the major model system in plant molecular genetics. The efforts of many scientists have produced genetic maps that provide extensive coverage of the genome (http://genome-www. stanford.edu/Arabidopsis/maps.html). Recently, detailed YAC, BAC, P1 and cosmid-based physical maps (that is, representations of genomic regions as sets of overlapping clones of corresponding libraries) have been established that extend over wide genomic areas ranging from several hundreds of kilobases to entire chromosomes. These maps provide an entry to gain deeper insight into the A. thaliana genome structure. A. thaliana has been chosen as the subject of the first large-scale project intended to determine the full genome sequence of a plant. This sequencing project, together with the increasing interest in map-based gene cloning, has highlighted the requirement for a complete and accurate physical map of this plant species. To supply the scientific community with a high-quality resource, we present here a complete physical map of A. thaliana using essentially the IGF BAC library. The map consists of 27 contigs that cover the entire genome, except for the presumptive centromeric regions, nucleolar organization regions (NOR) and telomeric areas. This is the first reported map of a complex organism based entirely on BAC clones and it represents the most homogeneous and complete physical map established to date for any plant genome. Furthermore, the analysis performed here serves as a model for an efficient physical mapping procedure using BAC clones that can be applied to other complex genomes.

Arabidopsis↗
PubMed1999

An encyclopedia of mouse genes.

The laboratory mouse is the premier model system for studies of mammalian development due to the powerful classical genetic analysis possible (see also the Jackson Laboratory web site, http://www.jax.org/) and the ever-expanding collection of molecular tools. To enhance the utility of the mouse system, we initiated a program to generate a large database of expressed sequence tags (ESTs) that can provide rapid access to genes. Of particular significance was the possibility that cDNA libraries could be prepared from very early stages of development, a situation unrealized in human EST projects. We report here the development of a comprehensive database of ESTs for the mouse. The project, initiated in March 1996, has focused on 5' end sequences from directionally cloned, oligo-dT primed cDNA libraries. As of 23 October 1998, 352,040 sequences had been generated, annotated and deposited in dbEST, where they comprised 93% of the total ESTs available for mouse. EST data are versatile and have been applied to gene identification, comparative sequence analysis, comparative gene mapping and candidate disease gene identification, genome sequence annotation, microarray development and the development of gene-based map resources.

Animals↗
PubMed1999

Changes in bioimpedance analysis after stable refeeding of undernourished anorexic patients.

OBJECTIVE: To evaluate bioimpedance analysis (BIA) parameters in patients with anorexia nervosa when undernourished and then after stable refeeding. DESIGN: Follow-up study. SUBJECTS: Thirteen patients with anorexia nervosa who were studied when undernourished (body weight (Wt): 36.9+/-5.6kg, body mass index (BMI): 14.8+/-1.8kg/m2) and after stable refeeding (Wt: 52.9+/-7.1kg, BMI: 21.2+/-2.3 kg/m2) compared to 25 well-nourished control women (Wt: 53.7+/-4.9 kg, BMI: 21.1+/-1.3 kg/m2). MEASUREMENTS: Impedance and phase angle were determined for the whole body (13 patients) and separately for arms and legs (10 patients). RESULTS: Bioimpedance index (height2/impedance) and phase angle were lower in the undernourished group. Whole-body impedance declined (median, min-max) by 45, 1-151 Ohm after refeeding; the variations of bioimpedance index (BI-Index) were weakly correlated (P < 0.10) with BMI changes. Limb phase angles increased with refeeding, but only changes in whole-body phase angle were correlated (P < 0.025) with the corresponding variations of Wt or BMI. CONCLUSION: Whole-body phase angle increased after weight recovery of anorexic patients, suggesting the occurence of modifications in the extracellular-to-intracellular water ratio. These changes were proportional to the increase in BMI and Wt.

Adolescent↗
PubMed1999

Interspecies differences in time course of pulmonary toxicity following repeated exposure to ozone.

To compare the extent and time course of pulmonary injury and repair in 3 rodent species, rats, mice and guinea pigs were continuously exposed for 3, 7, 28, and 56 days to 400 and 800 microg O3/m(3) (0.2 and 0.4 ppm). Recovery from 28 days of exposure was studied at 3, 7, and 28 days after exposure. Pulmonary injury and repair was studied at various time points by histology, electron microscopy, morphometry, and biochemistry. In all 3 species a concentration-related centriacinar inflammation occurred, with a maximum after 3 days of exposure. The number of alveolar macrophages and the pulmonary cell density in the centriacinar region increased progressively until 56 days of exposure, with the guinea pig the most sensitive species. Only the mouse displayed a concentration and exposure-time dependent hypertrophy of bronchiolar epithelium. After 56 days of exposure to 800 microg O3/m(3) in the rat and the guinea pig, giant lamellar bodies in type II cells were present. Exposures for 3 and 7 days at near ambient ozone concentrations (400 microg O3/m(3)) resulted in significantly elevated lung enzyme activities in the mouse, and in significant histological and morphometric changes in all 3 species. In rat and guinea pigs exposures for 56 days resulted in alveolar duct fibrosis. The highest biochemical response and the slowest recovery from ozone exposure were seen in the mouse. Histology, morphometry, and biochemistry revealed a total recovery from a 28-day exposure period in rats after 28 days, while in guinea pigs the ductular septa were still thickened and in mice all enzyme activities were still elevated in comparison with control values. In conclusion, the response of mice to ozone was evaluated as most severe, followed by those of guinea pigs and least in rats.

Administration, Inhalation↗
PubMed1999

Homologs of the Caenorhabditis elegans masculinizing gene her-1 in C. briggsae and the filarial parasite Brugia malayi.

The masculinizing gene her-1 in Caenorhabditis elegans (Ce-her-1) encodes a novel protein, HER-1A, which is required for male development. To identify conserved elements in her-1 we have cloned and characterized two homologous nematode genes: one by synteny from the closely related free-living species C. briggsae (Cb-her-1) and the other, starting with a fortuitously identified expressed sequence tag, from the distantly related parasite Brugia malayi (Bm-her-1). The overall sequence identities of the predicted gene products with Ce-HER-1A are only 57% for Cb-HER-1, which is considerably lower than has been found for most homologous briggsae genes, and 35% for Bm-HER-1. However, conserved residues are found throughout both proteins, and like Ce-HER-1A, both have putative N-terminal signal sequences. Ce-her-1 produces a larger masculinizing transcript (her-1a) and a smaller transcript of unknown function (her-1b); both are present essentially only in males. By contrast, Cb-her-1 appears to produce only one transcript, corresponding to her-1a; it is enriched in males but present also in hermaphrodites. Injection of dsRNA transcribed from Cb-her-1 into C. briggsae hermaphrodites (RNA interference) caused XO animals to develop into partially fertile hermaphrodites. Introducing a Cb-her-1 construct as a transgene under control of the C. elegans unc-54 myosin heavy chain promoter caused strong masculinization of both C. briggsae and C. elegans hermaphrodites. Introduction of a similar Bm-her-1 construct into C. elegans caused only very weak, if any, masculinization. We conclude that in spite of considerable divergence the Cb gene is likely to be a functional ortholog of Ce-her-1, while the function of the distantly related Bm gene remains uncertain.

Amino Acid Sequence↗
PubMed1999

A strong inverse relationship between PAI-1 and Lp(a) in hypertensive Type 2 diabetic patients.

Thrombophilia with a contemporary reduction of fibrinolytic activity has been observed both in diabetes mellitus and hypertension. Previously, we found a relationship between plasminogen activator inhibitor Type 1 (PAI-1) and lipoprotein(a) [Lp(a)] in Type 2 diabetes mellitus patients without complications. We hypothesised that this relationship could be due to a compensatory mechanism able to lower the risk of hypofibrinolysis as found in Type 2 diabetes mellitus. The present work was aimed at investigating the influence of concurrent hypertension and diabetes mellitus on the plasma levels of these two fibrinolytic inhibitors. In addition, other risk factors, known to influence the fibrinolytic parameters, were taken into account. Forty-nine Type 2 nonhypertensive diabetic patients without complications, 47 Type 2 hypertensive diabetic patients without complications, 54 non-diabetic hypertensive subjects without complications as well as 87 control subjects were studied. Plasma concentrations of Lp(a), PAI-1 antigen and activity, and the main parameters of oxidative, lipo- and glycometabolic balance were determined. Significant statistical differences between diabetic and non-diabetic subjects were found concerning triglycerides and antioxidant defence (p<0.01). Analysis of variance showed the F test statistically significant in evaluating the Log PAI-1/Lp(a) (p = 0.02). Correlation analysis between Log PAI-1 antigen and Lp(a) was significant in non-hypertensive diabetic patients, as expected (r = -0.38, p<0.01), and even stronger in hypertensive diabetic patients (r = -O.72,p<0.01). These results allow to hypothesise that the relationship between PAI-1/Lp(a) could be determinant in avoiding vascular complications due to diabetes mellitus and hypertension.

Aged↗
PubMed1998

Fusicoccin effect on the in vitro interaction between plant 14-3-3 proteins and plasma membrane H+-ATPase.

A 17-amino acid peptide was selectively cleaved from the highly variant C terminus of the 33-kDa 14-3-3 isoform occurring in fusicoccin receptor preparations from maize and was sequenced. The determined C-terminal sequence was identical to that of the already known maize 14-3-3 homolog GF14-6, thus prompting the use of recombinant GF14-6 in an in vitro protein-protein interaction study. The cDNA of GF14-6 was expressed in Escherichia coli as a 32P-phosphorylatable glutathione S-transferase fusion protein and was used as a probe in overlay experiments with H+-ATPase partially purified from maize roots. The results demonstrated that the recombinant protein specifically bound to H+-ATPase. The binding was dependent on Mg2+ and was strongly increased by fusicoccin. Controlled trypsin digestion of H+-ATPase abolished the association with GF14-6, a finding that was suggestive of an interaction with the C terminus of the enzyme. To confirm this result, the C-terminal domain of H+-ATPase was expressed as a glutathione S-transferase fusion peptide and was used in overlay experiments. GF14-6 was also able to bind to the isolated C terminus, but only in the presence of fusicoccin.

14-3-3 Proteins↗
PubMed1998

Studies on the life prolonging effect of food restriction: glutathione levels and glyoxalase enzymes in rat liver.

Cytosolic and mitochondrial levels of glutathione (GSH) as well as the activities of glyoxalase I (GI) and glyoxalase II (GII), GSH-dependent enzymes involved in the detoxification of 2-ketoaldehydes, were investigated in the liver of ad libitum (AL) fed and food restricted (FR) rat during aging. Both cytosolic and mitochondrial GSH level was lower in old than in adult AL fed rats. Food restriction did not prevent this decrease, but its extent was attenuated considering the cytosolic GSH. As regards the mitochondrial GSH, its content was higher in adult FR animals than in the age-matched AL fed ones. Thus, the subsequent age-dependent decrease of GSH, occurring also in FR animals, resulted in a thiol concentration not different from that observed in young and adult AL fed animals. Considering the enzymatic activities, cytosolic GI decreased in old rats irrespective of diet, whereas GII activity remained constant in all the experimental groups. The higher glutathione content found in both cellular compartments of old FR rats as compared to the old AL fed ones, could help to explain the life prolonging effect of FR treatment. Moreover, the observation that the activity of glyoxalases was not influenced by food restriction does not necessarily mean that the cells of diet-conditioned animals are scarcely protected against the toxic effect of methylglyoxal. Indeed, the production of this compound should be lower in FR animals as compared to AL fed ones, due to the lower level serum glucose concentration during the life span of the former with respect to the latter group.

Aging↗
PubMed1998

A significant relationship between plasminogen activator inhibitor type-1 and lipoprotein(a) in non-insulin-dependent diabetes mellitus without complications.

We previously found a relationship between plasminogen activator inhibitor type-1 and lipoprotein(a) in non-insulin-dependent diabetes mellitus and hypothesized that this could be due to a compensatory mechanism able to lower the risk of hypofibrinolysis found in type II diabetes mellitus. The aims of the present study were: (1) to confirm the association between plasminogen activator inhibitor type-1 and lipoprotein(a) in a different group of non-insulin-dependent diabetes mellitus patients and (2) to investigate whether the association could be related to diabetic complications. Other vascular risk factors able to influence fibrinolytic parameters such as glycemia, obesity, hypertension, dyslipidemia, and oxidative stress were also considered. Sixty-six non-insulin-dependent diabetes mellitus patients without diabetic complications (48 men, 18 women), 45 non-insulin-dependent diabetes mellitus patients with complications (21 men, 24 women), and 31 control subjects (17 men, 14 women) were studied. Plasma concentrations of lipoprotein(a), plasminogen activator inhibitor type-1 antigen and activity, and the main parameters of lipo- and glycometabolic balance were determined. Antioxidant defense was assayed as oxygen radical absorbance capacity of serum. Statistically significant differences among controls and the two diabetic groups were found for fasting glucose, cholesterol, triglycerides, and oxygen radical absorbance capacity of serum, while no statistically significant differences were evident for plasminogen activator inhibitor type-1 antigen and activity and lipoprotein(a). Regression analysis of log plasminogen activator inhibitor type-1/lipoprotein(a) showed a significant correlation only in diabetic patients without complications (r = -0.57, P < 0.001). These results show that a relationship between plasminogen activator inhibitor type-1 and lipoprotein(a) is characteristic of a diabetic population without complications, supporting the suggestion that this relationship could be a compensatory mechanism of the fibrinolytic system to limit the risks of hypofibrinolysis. A lack or a loss of capacity to balance lipoprotein(a) and plasminogen activator inhibitor type-1 could contribute to the pathogenesis of the diabetic complications.

Adult↗
PubMed1998

Fasting respiratory quotient as a predictor of weight changes in non-obese women.

OBJECTIVE: To detect predictive variables of weight changes in non-obese women. DESIGN: Three year follow-up study. SUBJECTS: Fifty-eight women (age 40.0+/-12.8 y, height 159+/-7 cm, weight 62.5+/-9.6 kg and body mass index ((BMI) 24.7+/-3.7 kg/m2). MEASUREMENTS: At baseline, basal metabolic rate (BMR) and respiratory quotient (RQ) by indirect calorimetry. Weight and BMI at baseline and after 3 y. RESULTS: In both univariate and multivariate analyses, age and RQ at baseline were significant predictors (P<0.05) of subsequent changes in weight and BMI. CONCLUSION: In non-obese women, RQ and age are independent predictors of subsequent weight changes.

Adult↗
PubMed1998

Gene discovery by EST sequencing in Toxoplasma gondii reveals sequences restricted to the Apicomplexa.

To accelerate gene discovery and facilitate genetic mapping in the protozoan parasite Toxoplasma gondii, we have generated >7000 new ESTs from the 5' ends of randomly selected tachyzoite cDNAs. Comparison of the ESTs with the existing gene databases identified possible functions for more than 500 new T. gondii genes by virtue of sequence motifs shared with conserved protein families, including factors involved in transcription, translation, protein secretion, signal transduction, cytoskeleton organization, and metabolism. Despite this success in identifying new genes, more than 50% of the ESTs correspond to genes of unknown function, reflecting the divergent evolutionary status of this parasite. A newly recognized class of genes was identified based on its similarity to sequences known only from other members of the same phylum, therefore identifying sequences that are apparently restricted to the Apicomplexa. Such genes may underlie pathways common to this group of medically important parasites, therefore identifying potential targets for intervention.

Animals↗
PubMed1998

Melatonin regulates the respiratory burst of human neutrophils and their depolarization.

The effect of different doses of melatonin on the respiratory burst as well as on the membrane potential changes of human neutrophils stimulated with phorbol 12-myristate 13-acetate (PMA) was investigated. The intracellular production of reactive oxygen species (ROS) in stimulated neutrophils was quantified in individual cells by flow cytometry, measuring the oxidation of nonfluorescent dihydrorhodamine 123 to the green fluorescent rhodamine 123. The transmembrane potential change was measured using the fluorescent probe oxonol. Preincubating the cells with micromolar concentrations of the indole resulted in an increase of the response to PMA. In two of six subjects investigated, the respiratory burst was also increased by a 10 nM concentration of the indole, but when the melatonin concentration was increased to 2 mM the respiratory burst was inhibited. The change in the transmembrane potential of neutrophils paralleled the respiratory burst. Indeed, the treatment of the cells with doses of melatonin up to 0.5 mM increased the depolarization occurring subsequent to PMA stimulation, whereas 2 mM melatonin concentration decreased the extent of depolarization. To investigate whether melatonin could directly affect the transmembrane potential changes of neutrophils, the extent of depolarization, induced by increasing the extracellular potassium concentration, was measured in cells preincubated with 2 mM melatonin. This treatment resulted in a decrease of the extent of depolarization, which suggests that melatonin can directly alter membrane ion conductance in human neutrophils.

Antioxidants↗
PubMed1998

Expressed sequence tag analysis of the bradyzoite stage of Toxoplasma gondii: identification of developmentally regulated genes.

Toxoplasma gondii is a protozoan parasite responsible for widespread infections in humans and animals. Two major asexual forms are produced during the life cycle of this parasite: the rapidly dividing tachyzoite and the more slowly dividing, encysted bradyzoite. To further study the differentiation between these two forms, we have generated a large number of expressed sequence tags (ESTs) from both asexual stages. Previously, we obtained data on approximately 7,400 ESTs from tachyzoites (J. Ajioka et al., Genome Res. 8:18-28, 1998). Here, we report the results from analysis of approximately 2,500 ESTs from bradyzoites purified from the cysts of infected mice. We also report the results from analysis of 760 ESTs from parasites induced to differentiate from tachyzoites to bradyzoites in vitro. Comparison of the data sets from bradyzoites and tachyzoites reveals many previously uncharacterized sequence clusters which are largely or completely specific to one or other developmental stage. This class includes a bradyzoite-specific form of enolase. Combined with the previously identified bradyzoite-specific form of lactate dehydrogenase, this finding suggests significant differences in flux through the lower end of the glycolytic pathway in this stage. Thus, the generation of this data set provides valuable insights into the metabolism and growth of the parasite in the encysted form and represents a substantial body of information for further study of development in Toxoplasma.

Amino Acid Sequence↗