Biomedical subjects

A Lombardo

Publications and source records attributed to A Lombardo.

At least 37 records · Page 2Linked to original sources

PubMed1998

Expression and functions of FGF-3 in Xenopus development.

We have analyzed the expression pattern of the Xenopus FGF-3 gene during early development and examined its biological activity in three different bioassays using Xenopus embryos. We show that from the early gastrula stage there is a domain of expression around the blastopore which becomes a posterior domain as the blastopore closes. An anterior ectodermal domain becomes detectable from mid-gastrula stages in the prospective hind-brain, and there are several later domains of expression: the midbrain-hindbrain junction, the otocyst, the pharyngeal pouches and the tailbud region. By using double whole-mount in situ hybridizations we show that the XFGF-3 expression in the brain is dynamically regulated both in time and space during development. The anterior domain of early neurula stage embryos corresponds to the prospective rhombomeres 3-5. By the time the neural tube is closed, XFGF-3 expression is restricted to r4 and later a new domain of expression is established at the midbrain/hindbrain junction. In addition, we show that, despite its difference in receptor specificity, XFGF-3 can induce the formation of mesoderm from animal caps similarly to other FGFs. It also displays a posteriorizing activity on whole embryos similar to other FGFs. Although the absence of maternal expression makes it unlikely that XFGF-3 is involved in mesoderm induction in vivo, its posterior domain of expression during gastrulation and its posteriorizing activity suggests that it participates in the maintenance of mesodermal gene expression and in the FGF mediated patterning of the anteroposterior axis during gastrulation.

Animals↗

PubMed1997

Effects of lead and manganese on the release of lysosomal enzymes in vitro and in vivo.

In this study we evaluated the effects of two heavy metals, lead and manganese, on the release of some glycohydrolases of lysosomal origin. N-acetyl-beta-D-glucosaminidase and its major isoenzymes, beta-D-glucuronidase and alpha-D-galactosidase. We have studied release of these enzymes in vitro from peripheral mitogen-activated lymphocytes from healthy subjects after addition of Pb or Mn to the medium and their plasma levels in individuals exposed at work to Pb (31 subjects) or to manganese (36 subjects), versus matched controls. We also determined the plasma levels in a general population (417 subjects). The enzymatic activities were assayed fluorimetrically with 4-methylumbelliferyl-glycosides as substrates. Particular attention was given to some technical aspects: enzymatic activity was preserved by addition of ethylene glycol and stable liquid material was employed for calibration purposes. N-acetyl-beta-D-glucosaminidase isoenzymes were separated by a routine chromatofocusing procedure on PBE 94. The addition of both metals to lymphocytes inhibits lysosomal enzyme release. These data were supported by the plasma levels for the exposed subjects, in which enzyme levels were significantly decreased after either type of exposure. In the general population of subjects not professionally exposed, the effect of lead appears to be masked by concomitant effects of alcohol consumption. Undoubtedly, some heavy metals can alter distribution of glycohydrolases of lysosomal origin between the intra- and extracellular environment, probably interfering with membrane mechanisms. Lysosomal enzymes seem to behave as sensitive biomarkers for early subclinical changes that might later lead to clinical disease.

Acetylglucosaminidase↗

PubMed1997

Inhibition of eFGF expression in Xenopus embryos by antisense mRNA.

We studied the effects in Xenopus embryos of overexpression of antisense RNA complementary to the messenger RNA of eFGF. We show that the expression of sense RNA can be severely depressed in the presence of an excess of antisense RNA. This occurs by both partial destruction of the message and by a depression of translation of the residual message. The diminution of inducing activity of eFGF, measured in animal cap assays either by activation of the Brachyury gene or by morphology, parallels the reduction of translation. Endogenous eFGF expression is reduced to a similar extent, again by a combination of mRNA destruction and inhibition of translation. This shows that the overexpression of antisense RNA is, contrary to general opinion, a potentially useful technique for studying gene function in Xenopus embryos. However, in the case of eFGF, there is little or no overall phenotypic effect on whole embryos. This is probably because of the presence of several other FGFs with overlapping expression domains in the early embryo.

Animals↗

PubMed1997

Contractile reserve of dysfunctional myocardium after revascularization: a dobutamine stress echocardiography study.

OBJECTIVES: We sought to investigate the effects of revascularization on the contractile reserve of dysfunctional myocardium. BACKGROUND: The improvement in dysfunctional but viable myocardium after revascularization is frequently less than expected from the amount of contractile reserve detected on dobutamine stress echocardiography. The fate of the contractile reserve, when it does not result in an adequate contractile recovery, is unknown. METHODS: Basal contraction and contractile reserve of infarct zones were assessed by dobutamine stress echocardiography in 21 postinfarction male patients before and > 3 months after revascularization (30 infarct zones; mean +/- SD left ventricular ejection fraction 35 +/- 8%). An infarct zone wall motion score index (WMSI) was calculated. RESULTS: Before revascularization, contractile reserve was present in 14 infarct zones (12 patients) and absent in 16 (9 patients). After revascularization, ejection fraction increased by 5 +/- 4% (p < 0.01) in patients classified as positive for contractile reserve and remained unchanged in those classified as negative. New York Heart Association classification improved in 58.3% and 22.2% of patients, respectively. Basal contraction improved in eight zones with previous contractile reserve (57.1%) and in one zone without (6.3%) (p < 0.01). Contractile reserve was still evident in 13 zones with previous contractile reserve (93%; 8 with contractile recovery), and it developed in 6 zones without (38%; none with contractile recovery). WMSI values after revascularization were decreased from values before revascularization during low dose dobutamine in zones with and without previous contractile reserve (p < 0.01 and < 0.05, respectively). CONCLUSIONS: After revascularization, contractile reserve is maintained or even increases in viable infarct zones that do not recover as expected. It may also develop in some infarct zones judged not to be viable before revascularization. This increased contractile reserve may play a role in the functional improvement of patients after revascularization.

Aged↗

PubMed1997

Detection of hibernated myocardium using intracoronary technetium-99m-sestamibi.

A possible limitation of intravenously administered perfusion tracers in detecting hibernating myocardium is their poor availability in low-flow areas. We have hypothesized that intracoronary administration might overcome such a limitation. In two patients with a previous anterior myocardial infarction and a severe residual stenosis of the left anterior descending coronary artery, technetium-99m-sestamibi (74 MBq) was selectively injected into the infarct-related coronary artery before successful PTCA. SPECT imaging was carried out 1 hour after the PTCA. In patient 1, evident tracer uptake was detected in the majority of the antero-apical dys-synergic area, previously characterized by a severe irreversible defect (less than 50% of peak activity) at rest-redistribution thallium SPECT ("mismatched pattern"); in patient 2, no uptake of sestamibi was detected in the dys-synergic area showing a severe irreversible defect (less than 50% of peak activity) at rest-redistribution thallium SPECT, ("matched pattern"). Forty days after PTCA, contractile recovery as well as normalization of thallium uptake occurred in patient 1, confirming that most of the dys-synergic area was viable (hibernating), while neither wall motion nor thallium uptake improvement occurred in patient 2, confirming that most of the dys-synergic area was not viable (scar). We conclude that sestamibi retention in the dys-synergic area after selective intracoronary administration correctly identified viable and scarred myocardium. The potential diagnostic value of intracoronary sestamibi at the time of coronary angiography deserves to be validated as a possible alternative to other current procedures for detecting hibernating myocardium.

Angioplasty, Balloon, Coronary↗

PubMed1996

Low incidence of serotonin-induced occlusive coronary artery spasm in patients with recent myocardial infarction.

Intracoronary infusion of serotonin has been reported to induce varying degrees of coronary vasoconstriction in different coronary syndromes, but it has never been studied in patients after myocardial infarction. In patients with recent myocardial infarction, we found a low incidence (11%) of serotonin-induced occlusive spasm only in the infarct-related artery (IRA), and a significantly higher vasoconstriction in the distal segment of the IRA than in the same segment of the non-IRA.

Cardiac Catheterization↗

PubMed1996

Plasma lysosomal glycohydrolases in a general population.

In this study we evaluated the differences in plasma levels of some glycohydrolases of lysosomal origin that appear to be the most interesting for possible usefulness for diagnosis (N-acetyl-beta-D-glucosaminidase, beta-D-glucuronidase, alpha-D-galactosidase, beta-D-galactosidase, alpha-L-fucosidase and alpha-D-mannosidase) in a general population of 417 subjects, as related to age and sex and also to body mass and to some habits, such as smoking and consumption of alcohol. The enzymatic activities were assayed by fluorimetric techniques with 4-methylumbelliferyl-glycosides as substrates. Particular attention was given to some technical aspects. Enzymatic activity was preserved by addition of ethylene glycol and stable liquid material was employed for calibration purposes. Blood was sampled rigorously at the same time of day and all the samples were obtained within a short period of time to exclude effects of the circadian and circannual rhythms. beta-Glucuronidase levels were the most affected by sex and body mass. beta-D-Galactosidase was not affected by differences in age, sex, body mass or by smoking, but appeared to be the most sensitive to modification by alcohol consumption. The data in this report emphasize that, whenever changes or differences in the levels of lysosomal enzymes in body fluids are studied, it is essential to have a reference population rigorously correlated with the study population. When possible, repetitive measurements in the same subject could better indicate a clinical trend.

Adult↗

PubMed1996

Significance of transient ST-T segment changes during dobutamine testing in Q wave myocardial infarction.

OBJECTIVES: We evaluated dobutamine stress electrocardiography for detecting potentially reversible contractile dysfunction or residual ischemia in the infarct-related area. BACKGROUND: ST-T segment changes in pathologic Q wave leads during stress testing may reflect contractile reserve, inducible ischemia or passive mechanical stretching. Dobutamine echocardiography allows detection of contractile reserve at low doses and inducible ischemia at high doses. METHODS: We used low (5 to 10 microg/kg body weight per min) and high doses (20 to 40 microg/kg per min) of dobutamine in 49 patients with a previous Q wave myocardial infarction and analyzed the relation between ST-T segment changes in pathologic Q wave leads and regional contraction. RESULTS: At low dose dobutamine, regional contraction improved in the infarct-related area in 23 patients. New or further ST segment elevation and pseudonormalization of negative T waves developed at low doses more frequently in patients with than without contractile reserve (both p < 0.001), giving a sensitivity of 43.5% and 60.9% and a specificity of 100% and 96.2%, respectively. At high dose dobutamine (43 patients), new or further ST segment elevation and pseudonormalization of negative T waves, occurring beyond those observed at low doses, had a low predictive accuracy for contractile reserve (sensitivity of 9.5% and 14.3% and specificity of 68.2% and 81.8%, respectively). Pseudonormalization of negative T waves at high dose dobutamine was 100% specific (but only 25% sensitive) for homozonal ischemia. CONCLUSIONS: ST segment elevation or pseudonormalization of negative T waves, or both, is indicative of contractile reserve in the infarct-related area when either develops at low dose dobutamine, but may be associated with worsening or no change in contractile function at high doses.

Aged↗

PubMed1996

Variable response of the peripheral circulation to acetylcholine in patients with coronary artery disease.

The purpose of this study was to assess endothelium-mediated vasodilation in the peripheral circulation of patients with coronary artery disease who are free from hypertension, hypercholesterolemia, diabetes mellitus, and congestive heart failure. The vascular response of the superficial femoral artery to an endothelium-dependent (i.e., acetylcholine 10-7, 10-6, and 10-5 mol/L) and to an endothelium-independent (i.e., nitroglycerin 10-8 and 10-6 mol/L) dilator was compared in 13 patients with angiographically documented coronary artery disease and in 7 patients with normal coronary angiograms. Vascular response was assessed by Doppler ultrasonography. Whereas the vascular responses to nitroglycerin in patients with abnormal and normal findings on coronary angiograms were similar, the responses to acetylcholine were clearly different. The ratio of mean blood flow velocity (+/-SD) measured during administration of acetylcholine 10-6 mol/L and mannitol was significantly lower in patients with abnormal versus normal results of coronary angiography (1.15 +/- 0.35 vs 2.20 +/- 1.06; p < 0.05). The vascular response to acetylcholine 10-5 mol/L in patients with an abnormal finding on their coronary angiogram was highly variable when compared with that in patients with normal results. Thus, in patients with angiographically proven coronary artery disease, the response of the peripheral circulation to acetylcholine is characterized by a great variability and a reduced sensitivity, when compared with that in patients with normal findings on coronary angiography.

Acetylcholine↗

PubMed1996

The organizer formation: two molecules are better than one.

A recent paper by Watabe et al.(1) describes the identification, in the promoter region of the organizer-specific gene goosecoid, of two distinct growth factor responsive elements that are required for the regulation of this gene both in vitro and in vivo. This is an important finding since it provides molecular evidence in support of the recently proposed model in which the production of dorsal type mesoderm is thought to require a synergistic interaction between a general mesoderm inducing factor and a dorsalizing competence modifier(2,6-8). Furthermore, the authors provide an elegant example of how a response element can become a useful tool to map the presence of a biologically available ligand within the embryo.

Animals↗

PubMed1996

Conformational flexibility and crystallization of tandemly linked type III modules of human fibronectin.

Fibronectin is a large cell adhesion molecule that is composed of several functional domains. The cell-binding domain that binds to cell surface integrins consists of repeated homologous type III modules. In this study, recombinant fragments from the cell-binding domain of human fibronectin that participate in a newly characterized fibronectin-fibronectin interaction with FNIII1 were crystallized. In each case, the crystals had more than one fibronectin fragment in the asymmetric unit. Crystals of FNIII10-11 grew in the space group C2 with a = 117.1 A, b = 38.6 A, c = 80.6 A, beta = 97.2 degrees, and two molecules in the asymmetric unit. These crystals diffracted to 2.5 A resolution. Fragment FNIII8-11 and a shorter fragment, FNIII8-10, crystallized in hexagonal space groups with large unit cells and two to four molecules per asymmetric unit. Even very large crystals of these fragments did not diffract beyond 4 A. The crystal packing for this collection of fibronectin fragments suggests conformational flexibility between linked type III modules. The functional relevance of this flexibility for elongated versus compact models of the cell-binding domain of fibronectin is discussed.

Ammonium Sulfate↗

PubMed1996

Regional wall motion analysis by dobutamine stess echocardiography to distinguish between ischemic and nonischemic dilated cardiomyopathy.

To distinguish between ischemic and nonischemic dilated cardiomyopathy (DCM), we studied 43 patients with left ventricular dysfunction (15 ischemic and 28 nonischemic detected by coronary angiography) by dobutamine stress echocardiography. At rest, there were more normal segments (p<0.001) and a trend toward more akinetic segments (p, not significant) per ischemic than per nonischemic DCM patient. However, either at rest or with low-dose dobutamine, individual data largely overlapped. At peak dose, in ischemic DCM, regional contraction worsened in many normal or dys-synergic regions at rest (in the latter case after improvement with low-dose dobutamine); in contrast, in nonischemic DCM, further mild improvement was observed in a variable number of left ventricular areas. Thus with peak-dose dobutamine, more akinetic and less normal segments were present per ischemic than per nonischemic DCM patient (both, p<0.001). A value of six or more akinetic segments was 80% sensitive and 96% specific for ischemic DCM. Our data show that analysis of regional contraction by dobutamine stress echocardiography can distinguish between ischemic and nonischemic DCM.

Aged↗

PubMed1996

Trisomy 20 in a papillary urothelial carcinoma of the ureter.

To contribute to the knowledge on tumorigenesis and the evolution of urothelial carcinoma of the ureter, we analyzed the clinical, histological, and cytogenetic aspects of a case. Primary cell cultures obtained from tumor specimens showed a trisomy of chromosome 20 where the c-src proto-oncogene, already described in literature as having an important role in the etiology and progression of some tumors, is located. In our case trisomy 20 is the only present marker and for this reason we think that it could play a role in the tumorigenesis of the urothelial carcinoma of the ureter.

Carcinoma, Papillary↗

PubMed1995

Co-crystallization of an ETS domain (PU.1) in complex with DNA. Engineering the length of both protein and oligonucleotide.

The PU.1 transcription factor is a member of the ets gene family of regulatory proteins. These molecules play a role in normal development and also have been implicated in malignant processes such as the development of erythroid leukemia. The Ets proteins share a conserved DNA-binding domain (the ETS domain) that recognizes a purine-rich sequence with the core sequence: 5'-C/AGGAA/T-3'. This domain binds to DNA as a monomer, unlike many other DNA-binding proteins. The ETS domain of the PU.1 transcription factor has been crystallized in complex with a 16-base pair oligonucleotide that contains the recognition sequence. The crystals formed in the space group C2 with a = 89.1, b = 101.9, c = 55.6 A, and beta = 111.2 degrees and diffract to at least 2.3 A. There are two complexes in the asymmetric unit. Production of large usable crystals was dependent on the length of both protein and DNA components, the use of oligonucleotides with unpaired A and T bases at the termini, and the presence of polyethylene glycol and zinc acetate in the crystallization solutions. This is the first ETS domain to be crystallized, and the strategy used to crystallize this complex may be useful for other members of the ets family.

Amino Acid Sequence↗

PubMed1995

Correlation of retinoid binding affinity to retinoic acid receptor alpha with retinoid inhibition of growth of estrogen receptor-positive MCF-7 mammary carcinoma cells.

Both anchorage-dependent growth and anchorage-independent growth of the estrogen receptor-positive mammary carcinoma cell line MCF-7 are inhibited by all-trans-retinoic acid. This cell line has nuclear retinoic acid receptors (RARs) alpha and gamma. The natural retinoids all-trans-retinoic acid and 9-cis-retinoic acid and a series of 12 conformationally restricted retinoids, which showed a range of binding selectivities for these receptors and had either agonist or antagonist activity for gene transcriptional activation by the RARs, were evaluated for their abilities to inhibit anchorage-dependent (adherent) and anchorage-independent (clonal) growth of MCF-7 cells. Correlation analyses were performed to relate growth inhibition by these retinoids with their binding affinity to RAR alpha or RAR gamma. Inhibition of anchorage-dependent growth in culture after 7 days of retinoid treatment correlated with binding to RAR alpha (n = 14; P < or = 0.001) and not to RAR gamma (n = 14; P > 0.1). Both the RAR alpha-selective retinoid agonists and the two RAR antagonists that were evaluated inhibited adherent cell growth. The RAR gamma-selective agonists had very low growth inhibitory activity (< 10%) at concentrations as high as 12.5 microM. These results suggest that RAR alpha is the retinoid receptor involved in the inhibition of adherent cell growth by retinoids and that transcriptional activation by this receptor on a RAR response element does not appear to be required for this process to occur. For this series of retinoids, inhibition of anchorage-independent growth after 21 days of retinoid treatment only correlated (n = 12; P < or = 0.005) with binding affinity to RAR alpha for the retinoid agonists, although the RAR gamma-selective retinoids displayed weak activity. The RAR antagonists were very poor inhibitors of growth. These results suggest that activation of gene transcription by RAR alpha appears to be required for inhibition of anchorage-independent growth by retinoids in this estrogen receptor-positive mammary carcinoma cell line.

Animals↗

PubMed1995

A developmentally regulated nerve growth factor-induced gene, VGF, is expressed in geniculocortical afferents during synaptogenesis.

The expression of the nerve growth factor-inducible gene VGF has been examined by in situ hybridization. Western blot and immunohistochemical studies in the developing and adult rat central nervous system, with particular emphasis on the visual system. Both the messenger RNA and the protein are particularly abundant in the developing dorsal lateral geniculate nucleus, appearing, respectively, at embryonal day 16 and 18. After its onset at E16, VGF messenger RNA expression increases progressively in the dorsal lateral geniculate nucleus and remains high during the first two post-natal weeks; afterwards, it gradually decreases and, at the offset of the plasticity period, it reaches very low levels maintained in adulthood. A similar time course has been observed for VGF protein in the dorsal lateral geniculate nucleus area, by semi-quantitative Western blots. In addition to the presence of the protein in the geniculate neurons, a strong, transient immunoreactivity has been found at the embryonic cortical subplate at E18, reflecting the presence of the antigen in axonal terminals originating from thalamic neurons. Interestingly, we found that the blockade of afferent electrical activity by intraocular injection of tetrodotoxin strongly reduces the level of VGF messenger RNA in the dorsal lateral geniculate nucleus. Although the function of the VGF protein is not known, it had been previously proposed that VGF could be a precursor for neuropeptide/s. The spatiotemporal expression of VGF, together with the observation of a regulation by electrical activity, suggest that this protein may be relevant in the process of synaptogenesis and/or synaptic stabilization in the developing geniculocortical connections.

Afferent Pathways↗

PubMed1995

Automated fluorimetric assay procedure for glucohydrolases using a routine centrifugal analyser assay of enzymes of lysosomal origin in plasma, II.

The manual fluorimetric procedure, considered as a reference method for the determination of N-acetyl-beta-D-glucosaminidase, beta-D-glucuronidase and beta-D-galactosidase in human plasma, was automated as a routine method, using the IL Monarch centrifugal analyser. Using a liquid standard with a known enzyme content, the automated assay correlated fairly well with the reference manual method (r values very close to 1). Its analytical imprecision was much lower than that of the manual method. The automated assay of N-acetyl-beta-D-glucosaminidase, beta-D-glucuronidase and beta-D-galactosidase gave coefficients of variation of 5.7-6.9, 3.6-5.0 and 3.8-4.2%, respectively, detection limits of 4, 2 and 1 mU/l plasma respectively, and linear responses of up to 73, 8.4 and 0.9 U/l of plasma respectively. Furthermore, the method required only small volumes of undiluted plasma (4-10 microliters). This method appears to be reliable, sensitive, simple enough for routine analyses and as cost effective as the most common routine serum enzyme assays.

Acetylglucosaminidase↗

PubMed1994

Recombinant human retinoic acid receptor beta. Binding of synthetic retinoids and transcriptional activation.

All-trans-retinoic acid mediates cell growth and differentiation by binding to and then activating nuclear retinoid receptor proteins that regulate gene transcription. Recombinant human retinoic acid receptor beta was cloned and expressed in Escherichia coli as a fusion protein rMBP-RAR beta with maltose-binding protein to facilitate purification. After isolation from bacterial lysates, rMBP-RAR beta was used for binding with selected retinoids. Scatchard analysis with [11,12-3H2]all-trans-retinoic acid gave a Kd of 0.34 nM. Competitive binding studies with a series of conformationally restricted aromatic retinoids indicated that the Ki values for binding to rMBP-RAR beta correlated with the logs of the EC50 values for gene transcriptional activation (p < or = 0.05) and with those for the relative activation compared to that of all-trans-retinoic acid (p < or = 0.01). Inspection of binding-activation correlation diagrams indicates candidate structures for improved retinoid agonists or antagonists.

Amino Acid Sequence↗