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Biomedical subjects

G Micheli

Publications and source records attributed to G Micheli.

At least 19 recordsLinked to original sources
PubMed2001

The looped domain organization of the nucleoid in histone-like protein defective Escherichia coli strains.

We have investigated the major Escherichia coli histone-like proteins (H-NS, HU, FIS, and IHF) as putative factors involved in the maintenance of the overall DNA looped arrangement of the bacterial nucleoid. The long-range architecture of the chromosome has been studied by means of an assay based on in vivo genomic fragmentation mediated by endogenous DNA gyrase in the presence of oxolinic acid. The fragmentation products were analysed by CHEF electrophoresis. The results indicate that in vivo a large fraction of the bacterial chromatin constitutes an adequate substrate for the enzyme. DNA fragments released upon oxo-treatment span a size range from about 1000 kb to a limit-size of about 50 kb. The latter value is in excellent agreement with the average size reported for bacterial chromosomal domains. The DNA gyrase-mediated fragmentation does not appear to be significantly altered in strains depleted in histone-like proteins as compared to an E. coli wild type strain. This suggests that these proteins may not represent critical determinants for the maintenance of the supercoiled loop organisation of the E. coli chromosome.

Amino Acid Motifs↗
PubMed2000

Prognostic value of pulmonary venous flow Doppler signal in left ventricular dysfunction: contribution of the difference in duration of pulmonary venous and mitral flow at atrial contraction.

OBJECTIVES: We assessed the contribution of difference in duration of pulmonary venous and mitral flow at atrial contraction (ARd-Ad) for prognostic stratification of patients with left ventricular (LV) systolic dysfunction. BACKGROUND: Although pulmonary venous flow (PVF) variables may supplement mitral flow patterns in evaluating left ventricular (LV) diastolic function, their value to the prognostic stratification of patients has not been investigated. METHODS: Pulsed wave Doppler mitral and PVF velocity curves were recorded in 145 patients (mean age: 70 years) with LV systolic dysfunction secondary to ischemic or nonischemic cardiomyopathy who were followed for 15 +/- 8 months. In 38% of patients, PVF signal was enhanced by the intravenous (IV) administration of a galactose-based echo-contrast agent. Based on E-wave deceleration time < or = or >130 ms and ARd-Ad, patients were grouped into restrictive (group 1, n = 40), nonrestrictive with ARd-Ad > or =30 ms (group 2, n = 55) and nonrestrictive with ARd-Ad <30 ms (group 3, n = 50). RESULTS: During follow-up, 29 patients died from cardiac causes and 28 were hospitalized for worsening heart failure (HF). On multivariate Cox model, ARd-Ad > or =30 ms provided important prognostic information with regard to cardiac mortality and emerged as the single best predictor of cardiac events (cardiac mortality, hospitalization). The 24-month cardiac event-free survival was best (86.3%) for group 3; it was intermediate (37.9%) for group 2; and it was worst (22.9%) for group 1 (p < 0.0002 group 1 vs. 3; p < 0.0005 group 2 vs. 3; p < 0.0003 group 1 vs. group 2). CONCLUSIONS: Assessment of ARd-Ad exhibited an independent value in the prognostic evaluation of patients with LV systolic dysfunction. Moreover, it contributed to identify patients at low, intermediate and high risk of cardiac events.

Adult↗
PubMed2000

[Is a totally non-invasive assessment of the hemodynamic profile possible in patients with chronic heart failure?].

Relevant hemodynamic information can be obtained by a comprehensive Doppler echocardiographic examination in patients with various cardiac diseases. The assessment of left heart hemodynamics by Doppler echocardiography has been addressed by several investigators. The feasibility and the accuracy of methods for the estimation of left ventricular filling pressure and cardiac output have been validated by comparative right heart catheterization. Studies have shown that Doppler echocardiography can allow the measurement of pulmonary artery pressures from the pressure gradients across the tricuspid and pulmonary valves. The possibility of completely characterizing cardiac hemodynamics noninvasively has recently been documented: in patients with acute myocardial infarction, automated cardiac output measurement along with the assessment of left ventricular filling by Doppler echocardiography may be used for the identification of hemodynamic subsets. Although Doppler echocardiography can provide noninvasive measures of hemodynamic indices, its value has been disputed since the technique is patient-dependent, time-consuming and requires meticulous acquisition and interpretation by skilled operators. The use of contrast agents may improve the accessibility of both right-sided and left-sided Doppler signals, potentially increasing the number of patients to whom the noninvasive hemodynamic assessment could be applied.

Chronic Disease↗
PubMed1998

Thermoregulation of Shigella and Escherichia coli EIEC pathogenicity. A temperature-dependent structural transition of DNA modulates accessibility of virF promoter to transcriptional repressor H-NS.

The expression of plasmid-borne virF of Shigella encoding a transcriptional regulator of the AraC family, is required to initiate a cascade of events resulting in activation of several operons encoding invasion functions. H-NS, one of the main nucleoid-associated proteins, controls the temperature-dependent expression of the virulence genes by repressing the in vivo transcription of virF only below a critical temperature (approximately 32 degrees C). This temperature-dependent transcriptional regulation has been reproduced in vitro and the targets of H-NS on the virF promoter were identified as two sites centred around -250 and -1 separated by an intrinsic DNA curvature. H-NS bound cooperatively to these two sites below 32 degrees C, but not at 37 degrees C. DNA supercoiling within the virF promoter region did not influence H-NS binding but was necessary for the H-NS-mediated transcriptional repression. Electrophoretic analysis between 4 and 60 degrees C showed that the virF promoter fragment, comprising the two H-NS sites, undergoes a specific and temperature-dependent conformational transition at approximately 32 degrees C. Our results suggest that this modification of the DNA target may modulate a cooperative interaction between H-NS molecules bound at two distant sites in the virF promoter region and thus represents the physical basis for the H-NS-dependent thermoregulation of virulence gene expression.

Bacterial Proteins↗
PubMed1998

A role for H-NS in the regulation of the virF gene of Shigella and enteroinvasive Escherichia coli.

We have investigated the role of H-NS, one of the major components of the bacterial nucleoid, in the expression of the virF gene present on the large virulence plasmid of Shigella and enteroinvasive Escherichia coli in response to different environmental conditions. VirF is an AraC-like protein which activates at least two promoters, virB and virG, both repressed by H-NS. Band shift experiments reveal that the affinity of H-NS for the virF and virB promoters is comparable, while the affinity for the virG promoter is higher. Polyacrylamide gel electrophoresis of three DNA fragments containing the virF, the virB and the VirG promoters demonstrates, in agreement with computer predictions, that they have an intrinsically curved structure, confirming the preference of H-NS for bent DNA. In vivo transcriptional analysis of virF mRNA shows that H-NS negatively controls the expression of virF at 30 degrees C. The expression of a virF-lacZ translational fusion in E.coli wild type and in an hns-defective derivative grown at 30 degrees or 37 degrees C and at pH 6.0 or 7.0 indicates that, in the absence of H-NS, virF expression becomes insensitive to temperature and to limited pH changes. Our results strongly suggest that H-NS controls virF expression by binding to the virF promoter and by repressing its expression at low temperature and at low pH.

Bacterial Proteins↗
PubMed1998

Left ventricular size and function in patients with noninsulin-dependent diabetes and postinfarction total or subtotal coronary occlusions.

The aim of this study was to establish the effects of postinfarction total or subtotal coronary occlusion on left ventricular remodeling in patients with noninsulin-dependent diabetes (NIDD) compared with the effects in postinfarct nondiabetic patients. The authors selected 100 patients submitted to coronary angiography between 1 and 5 weeks after acute myocardial infarction (T0: 20.5+/-15.4 days) and classified into three groups: G1: NIDD with coronary occlusion/subocclusion (n=24), G2: controls with coronary occlusion/subocclusion (n=43), G3: controls without coronary occlusion/subocclusion (n=33). At time zero (T0) the following parameters were evaluated: end-systolic and end-diastolic volume indexes (ESVi, EDVi), ejection fraction (EF), echocardiographic wall motion score index (WMI), presence of left ventricular aneurysm, and triple-vessel coronary disease. The frequencies of major cardiovascular events were recorded during follow-up. Significantly greater ESVi and EDVi were noted in G2 compared with G3 (P<0.0001), while no significant differences were observed between NIDD patients and controls. Although left ventricular global and segmental dysfunctions were increased in diabetics, controls with coronary occlusion/subocclusion presented more pronounced EF reduction (P<0.0001 G2 vs G3) and higher elevation in WMI (P<0.005 G2 vs G3). Cardiac events during follow-up were elevated in G1 and G2, particularly as regards the occurrence of congestive heart failure. The authors conclude that NIDD seems to influence in a positive way left ventricular remodeling associated with postinfarct total or subtotal coronary occlusion.

Coronary Angiography↗
PubMed1997

[Dipyridamole-echocardiography test in the diagnosis of vasomotor angina].

Dipyridamole-atropine echocardiography testing is used extensively for the diagnosis of coronary artery disease and it is highly effective in diagnosing "organic" coronary artery disease by inducing myocardial ischemia via three different mechanisms: maximal coronary artery vasodilatation with phoenomena of flow-maldistribution caused by dipyridamole, increase in myocardial oxygen consumption and reduction of the oxygen supply to the myocardium caused by atropine. Moreover, the abrupt withdrawal of the coronary artery vasodilatation caused by aminophylline, which is routinely infused at the end of the test, may trigger coronary artery spasms in patients with variant angina, thus enhancing the diagnostic power of the test. We report two clinical cases of patients with rest angina and angiographically normal coronary arteries, in whom coronary artery spasm was induced by administering aminophylline during the stress test.

Coronary Vasospasm↗
PubMed1994

Monoclonal antibodies that recognise filamentous phage: tools for phage display technology.

We generated six hybridoma cell lines that secrete monoclonal antibodies (mAb) which specifically bind filamentous phage coat proteins. Two of these mAb recognise epitopes that include the N terminus of the coat protein III (pIII), while two others are specific for the N terminus of the major coat protein VIII (pVIII). These mAb are valuable tools to study phage assembly and structure. Furthermore, we describe two examples of how these mAb can be exploited in the construction and screening of peptide libraries displayed by the filamentous phase major coat protein. We have used one of these mAb to develop a sensitive ELISA with crude phage supernatants. This assay allows rapid screening of large numbers of clones from random peptide phage libraries. Some of the anti-phage mAb described here can interfere with wild-type phage propagation, while phage carrying modifications in their coat proteins are insensitive to growth inhibition. We have exploited this observation as a tool to favour the growth of phage displaying peptides fused to pVIII, with respect to vector phage.

Amino Acid Sequence↗
PubMed1994

Copper-dependent metabolism of Cu,Zn-superoxide dismutase in human K562 cells. Lack of specific transcriptional activation and accumulation of a partially inactivated enzyme.

The regulation of Cu,Zn-superoxide dismutase by copper was investigated in human K562 cells. Copper ions caused a dose- and time-dependent increase, up to 3-fold, of the steady-state level of Cu,Zu-superoxide dismutase mRNA. A comparable increase was also observed for actin and ribosomal protein L32 mRNAs, but not for metallothionein mRNA which was augmented more than 50-fold and showed a different induction pattern. The copper-induced mRNAs were actively translated as judged from their enhanced loading on polysomes, the concomitantly increased cellular protein levels and an augmented incorporation of [3H]lysine into acid-precipitable material. Cu,Zn-superoxide dismutase protein followed this general trend, as demonstrated by dose- and time-dependent increases in immunoreactive and enzymically active protein. However, a specific accumulation of Cu,Zn-superoxide dismutase was noticed in cells grown in the presence of copper, that was not detectable for other proteins. Purification of the enzyme demonstrated that Cu,Zn-superoxide dismutase was present as a reconstitutable, copper-deficient protein with high specific activity (kcat./Cu = 0.89 x 10(9) M-1.s-1) in untreated K562 cells and as a fully metallated protein with low specific activity (kcat./Cu = 0.54 x 10(9) M-1.s-1) in copper-treated cells. Pulse-chase experiments using [3H]lysine indicated that turnover rates of Cu,Zn-superoxide dismutase in K562 cells were not affected by growth in copper-enriched medium, whereas turnover of total protein was significantly enhanced as a function of metal supplementation. From these results we conclude that: (i) unlike in yeast [Carrì, Galiazzo, Ciriolo and Rotilio (1991) FEBS Lett. 278, 263-266] Cu,Zn-superoxide dismutase is not specifically regulated by copper at the transcriptional level in human K562 cells, suggesting that this type of regulation has not been conserved during the evolution of higher eukaryotes; (ii) copper ions cause an inactivation of the enzyme in intact K562 cells; and (iii) the metabolic stability of Cu,Zn-superoxide dismutase results in its relative accumulation under conditions that lead to increased protein turnover.

Animals↗
PubMed1994

A novel illegitimate recombination event: precise excision and reintegration with the Mu gem mutant prophage.

The bacteriophage Mu is known to insert its DNA more or less randomly within the Escherichia coli chromosome, as do transposable elements, but unlike the latter, precise excision of the prophage, thereby restoring the original sequence, is not observed with wild-type Mu, although it has been reported with certain defective mutants. We show here that the mutant prophage Mu gem2ts can excise precisely from at least three separate loci -- malT, lac and thyA (selected as Mal+, Lac+ and Thy+, respectively). This excision occurs under permissive conditions for phage development, is observed in fully immune (c+) lysogens, and is independent of RecA and of Mu transposase. Mu gemts2 excision is invariably accompanied by reintegration of a Mu gem2ts prophage elsewhere in the chromosome. In the case of Mal+ revertants, this prophage is systematically located at 94 min on the E. coli chromosome. Mu gem2ts excision therefore sheds some light on the long-standing paradox of the lack of precise Mu excision.

Bacterial Proteins↗
PubMed1993

Chromosome length and DNA loop size during early embryonic development of Xenopus laevis.

The looped organization of the eukaryotic genome mediated by a skeletal framework of non-histone proteins is conserved throughout the cell cycle. The radial loop/scaffold model envisages that the higher order architecture of metaphase chromosomes relies on an axial structure around which looped DNA domains are radially arranged through stable attachment sites. In this light we investigated the relationship between the looped organization and overall morphology of chromosomes. In developing Xenopus laevis embryos at gastrulation, the bulk of the loops associated with histone-depleted nuclei exhibit a significant size increase, as visualized by fluorescence microscopy of the fully extended DNA halo surrounding high salt treated, ethidium bromide stained nuclei. This implies a reduction in the number of looped domains anchored to the supporting nucleoskeletal structure. The cytological analysis of metaphase plates from acetic acid fixed whole embryos, carried out in the absence of drugs inducing chromosome condensation, reveals a progressive thickening and shortening of metaphase chromosomes during development. We interpret these findings as a strong indication that the size and number of DNA loops influence the thickness and length of the chromosomes, respectively. The quantitative analysis of chromosome length distributions at different developmental stages suggests that the shortening is timed differently in different embryonic cells.

Animals↗
PubMed1993

[A case of an aneurysm of the main pulmonary artery and of its 2 branches].

A case of a 76-year-old female with a large aneurysm of the main pulmonary artery and initial tract of its branches is reported. Literature is reviewed as to possible etiopathogenesis. Based on past medical history, clinical findings and investigative procedures, we hypothesize a possible congenital origin of the anomaly; its gradual aggravation might be due to the development of pulmonary hypertension.

Aged↗
PubMed1989

[Supraventricular tachyarrhythmia of recent onset in the absence of documented cardiac pathology: different therapeutic approaches].

The number of supraventricular tachyarrhythmias recently arisen among patients admitted to the Coronary Care Unit in Piombino, and the results of therapeutical interventions, have been estimated by analysing, retrospectively, the cases pertinent to the period February '84- October '87. Out of 1451 hospitalized patients, the supraventricular tachyarrhythmias were 145; 85 of them (46 F and 39 M, average age 63 +/- 14) with an assured cardiopathy and 60 of them (34 F and 26 M, average age 58 +/- 13) with no assured cardiopathy. The employed drugs, as a first choice, were verapamil, amiodarone or flecainide on the ground of study protocols in order to value their compared effectiveness. When amiodarone was not employed, as a first choice, in conformity with a specific protocol, it was utilized as a second choice after verapamil or flecainide. A DC shock was carried out in the case of pharmacological failure. Among the patients with an assured cardiopathy, the pharmacologically cardioversed ones turn out 55.3%; the spontaneously ones 9.4%; the ones treated by DC shock 14.1%; the non cardioversed ones 21.2%. Among the patients with no assured cardiopathy, the pharmacologically cardioversed ones turn out 78.3%; the spontaneously ones 5%; the treated by DC shock ones 11.7%; the non-cardioversed ones 5%. Therefore, the supraventricular tachyarrhythmias of recent onset make up nearly 10% of the hospitalizations into a Coronary Care Unit; the percentage of cardioversions in a sinusal rhythm with a pharmacological treatment reaches 95% in those patients with no assured cardiopathy.(ABSTRACT TRUNCATED AT 250 WORDS)

Aged↗
PubMed1988

The Salmonella wien virulence plasmid pZM3 carries Tn1935, a multiresistance transposon containing a composite IS1936-kanamycin resistance element.

Tn1935, a 23.5-kb transposon mediating resistance to ampicillin, kanamycin, mercury, spectinomycin, and sulfonamide was isolated from pZM3, an IncFIme virulence plasmid from Salmonella wien. Tn1935 possesses the entire sequence of Tn21 and contains two additional DNA segments of 0.95 and 2.7 kb carrying the ampicillin and kanamycin resistance genes, respectively. The latter is part of a composite element since it is flanked by two IS15-like insertion sequences (IS1936) in direct orientation. IS1936 is about 800 bp long and is closely related to IS15 delta, IS26, IS46, IS140, and IS176. Functional analysis of IS1936-mediated cointegrates shows that both insertion sequences are active and able to form cointegrates at the same frequency. Resolution of the cointegrates requires the presence of the host Rec system. The presence of the composite IS1936-element within Tn1935 supports the hypothesis that multidrug resistance transposons evolved by insertion of antibiotic determinants which are themselves transposable.

Anti-Bacterial Agents↗
PubMed1986

The relationship between chromosomal origins of replication and the nuclear matrix during the cell cycle.

A cytological investigation into the dynamic behaviour of the origins of replication with respect to the nuclear matrix has been carried out on Xenopus laevis cultured cells. In order to preferentially label origins or 'non-origin' regions along DNA fibres, 5-fluoro-2'-deoxyuridine (FUdR)-treated cells were pulsed with [3H]deoxyadenosine in early or late S phase. Samples were then allowed to proceed through the cell cycle for increasing times. The DNA loops were induced in situ to completely uncoil around the nuclear matrix. The autoradiographic analysis shows that, under the experimental conditions used, 'non-origin' regions behave as expected from previous studies, i.e., they associate with the nuclear matrix only when they become part of a replication fork, whereas active origins of replication remain associated with the matrix throughout the cell cycle.

Animals↗
PubMed1985

Tadpole Xenopus laevis hemoglobin. Correlation between structure and functional properties.

Perutz & Brunori (1982) proposed that the COOH-terminal His and Ser F9 of the beta-chains of fish and amphibian hemoglobins are responsible for their Root effect and part of their alkaline Bohr effect. Analysis of the kinetics of carbon monoxide binding by hemoglobin from the tadpole of Xenopus laevis supports that model and suggests an explanation for the absence of an alkaline Bohr effect in many aquatic Anura and Urodela.

Amino Acid Sequence↗