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Biomedical subjects

L Guida

Publications and source records attributed to L Guida.

At least 37 records · Page 2Linked to original sources
PubMed1999

Clinic-daytime blood pressure difference and cardiovascular damage.

OBJECTIVE: To investigate whether the clinic-daytime blood pressure difference can provide information on vascular reactivity to stress comparable to that of simple noninvasive stimuli such as a cold pressor test and isometric exercise, and whether there is any relationship between this blood pressure difference and noninvasive measurements of the left ventricular mass and carotid arterial wall. DESIGN: Patients with newly discovered, never-treated, sustained hypertension were included in the study after a 1 month run-in, during which time their blood pressure was measured three times at 2 week intervals. METHODS: Blood pressure was measured by a noninvasive procedure at rest and during a cold pressor test and an isometric exercise. The difference was calculated for systolic, diastolic and mean blood pressure as resting minus daytime ambulatory blood pressure. Parameters of the posterior wall and septal thickness of the left ventricle, aortic root and left atrium were studied by M-mode echocardiography. Carotid wall thickness and diameter were measured using ultrasound. RESULTS: The 90 patients enrolled in the study were divided into tertiles of clinic-daytime blood pressure difference. The composition of the groups differed in sex, since the majority of women were in the highest tertile, but was comparable for age, body mass index, renin-aldosterone axis and lipid and carbohydrate metabolism. Blood pressure responses to cold and isometric exercise were more pronounced in patients in the lowest tertile of blood pressure difference. No intergroup differences were detected in echocardiographic parameters of ventricular (left ventricular mass, tertiles I-III: 46.5 +/- 10, 42.3 +/- 8, 44.8 +/- 13 g/m2.7, respectively) and carotid (intima-media thickness, tertiles I-III 0.58 +/- 0.1, 0.54 +/- 0.1, 0.62 +/- 0.1 mm, respectively) structure. CONCLUSIONS: The present study indicates that the clinic-daytime blood pressure difference provides different information on cardiovascular reactivity compared with that obtained from the cold pressor test and isometric exercise. Moreover, it does not seem to have any relationship with ventricular hypertrophy and/or carotid wall thickening.

Adult↗
PubMed1998

Dimeric and tetrameric forms of catalytically active transmembrane CD38 in transfected HeLa cells.

CD38, a type II transmembrane glycoprotein, behaves as a catalytically active transporter responsible for ectocellular generation of cyclic ADP-ribose (cADPR) from NAD+ and for subsequent influx of cADPR across membranes [Franco, L., Guida, L., Bruzzone, S., Zocchi, E., Usai, C. and De Flora, A. (1998) FASEB J. in press]. cADPR regulates intracellular calcium homeostasis by releasing calcium from responsive stores. The cADPR-transporting function of CD38 requires channel-generating oligomeric forms of the protein rather than the 46 kDa monomers that have been described so far in CD38+ cells. Here we demonstrate that CD38, both in reconstituted proteoliposomes and in CD38-transfected HeLa cells, is a mixture of catalytically active monomers, homodimers and homotetramers. A soluble recombinant form of CD38 corresponding to its ectocellular region proved to be monomeric. Thus, association of native CD38 with either artificial or natural membranes seems to result in a reversible juxtaposition of monomers suitable to cADPR-transporting activity.

ADP-ribosyl Cyclase↗
PubMed1998

Expression of CD38 increases intracellular calcium concentration and reduces doubling time in HeLa and 3T3 cells.

CD38 is a bifunctional ectoenzyme, predominantly expressed on hematopoietic cells during differentiation, that catalyzes the synthesis (cyclase) and the degradation (hydrolase) of cyclic ADP-ribose (cADPR), a powerful calcium mobilizer from intracellular stores. Due to the well established role of calcium levels in the regulation of apoptosis, proliferation, and differentiation, the CD38/cADPR system seems to be a likely candidate involved in the control of these fundamental processes. The ectocellular localization of the cyclase activity, however, contrasts with the intracellular site of action of cADPR. Here we demonstrate that ectocellular expression of human CD38 in CD38(-) HeLa and 3T3 cells results in intracellular CD38 substrate (NAD+ + NADH) consumption and product (cADPR) accumulation. Furthermore, a causal relationship is established between presence of intracellular cADPR, partial depletion of thapsigargin-sensitive calcium stores, increase in basal free cytoplasmic calcium concentration, and decrease of cell doubling time. The significant shortening of the S phase in CD38(+) HeLa cells, as compared with controls, demonstrates an effect of intracellular cADPR on the mammalian cell cycle.

3T3 Cells↗
PubMed1998

Ectocellular CD38-catalyzed synthesis and intracellular Ca(2+)-mobilizing activity of cyclic ADP-ribose.

CD38 is a type-II transmembrane glycoprotein occurring in several hematopoietic and mature blood cells as well as in other cell types, including neurons. Although classified as an orphan receptor, CD38 is also a bifunctional ectoenzyme that catalyzes both the conversion of NAD+ to nicotinamide and cyclic ADP-ribose (cADPR), via an ADP-ribosyl cyclase reaction, and also the hydrolysis of cADPR to ADP-ribose (hydrolase). Major unresolved questions concern the correlation between receptor and catalytic properties of CD38, and also the apparent contradiction between ectocellular generation and intracellular Ca(2+)-mobilizing activity of cADPR. Results are presented that provide some explanations to this topological paradox in two different cell types. In cultured rat cerebellar granule neurons, extracellular cADPR (either generated by CD38 or directly added) elicited an enhanced intracellular Ca(2+)-response to KCl-induced depolarization, a process that can be qualified as a Ca(2+)-induced Ca2+ release (CICR) mechanism. On the other hand, in the CD38+ human Namalwa B lymphoid cells, NAD+ (and thiol compounds as well) induced a two-step process of self-aggregation followed by endocytosis of CD38, which resulted in a shift of cADPR metabolism from the cell surface to the cytosol. Both distinctive types of cellular responses to extracellular NAD+ seem to be suitable to elicit changes in the intracellular Ca2+ homeostasis.

ADP-ribosyl Cyclase↗
PubMed1998

Cardiovascular abnormalities in never-treated hypertensives according to nondipper status.

Ambulatory blood pressure monitoring allows a better understanding of blood pressure fluctuations over 24 h than simple clinic measurements. In this way the diagnosis of "white coat" versus "sustained" hypertension and that of "dipper" (patient with blood pressure fall during nighttime > 10% of daytime levels) versus "nondipper" status were made possible. This pilot study has been undertaken to investigate whether patients with recently discovered, never-treated, mild, sustained hypertension have cardiovascular abnormalities according to their dipper/nondipper status. Patients with long-standing (n = 123) and newly discovered (n = 56) sustained hypertension were classified according to their nighttime blood pressure fall, and compared with normotensive controls. Ambulatory blood pressure monitoring was performed noninvasively. Parameters of left ventricular structure, cardiac systolic and diastolic function, and carotid anatomy were determined noninvasively by echographic methods. Significant increases in parameters of cardiac structure as well as abnormalities in diastolic function were observed in patients with long-standing hypertension, regardless of their dipper status. In the group with newly discovered hypertension, left atrium (3.4+/-0.3, 3.7+/-0.5, 3.2+/-0.4 cm in dippers, nondippers, and controls, respectively), end-diastolic diameter index (2.9+/-0.3, 3.0+/-0.2, 2.8+/-0.2 cm/m), and atrial filling fraction (0.50+/-0.07, 0.52+/-0.05, 0.42+/-0.04) were significantly altered only in the nondipper subgroup, in comparison with controls. Significant changes in cardiac structure and diastolic function were observed in nondipper patients with recently discovered hypertension, who, at variance with dippers, show changes similar to those in patients with long-standing hypertension. Hypertensives with the observed abnormalities may benefit from active antihypertensive treatment, which appears, therefore, justified even in an early phase of mild hypertension, in terms of potential reduction of end-organ complications as well as cost-effectiveness.

Adult↗
PubMed1998

The transmembrane glycoprotein CD38 is a catalytically active transporter responsible for generation and influx of the second messenger cyclic ADP-ribose across membranes.

CD38 is a type II transmembrane glycoprotein expressed in many vertebrate cells. It is a bifunctional ectoenzyme that catalyzes both the synthesis of Cyclic ADP-ribose (cADPR) from NAD+ and the degradation of cADPR to ADP-ribose by means of its ADP-ribosyl cyclase and cADPR-hydrolase activities, respectively. The cyclase also converts NGD+ to cyclic GDP-ribose (cGDPR), which is refractory to cADPR-hydrolase. cADPR, but not cGDPR, is a potent calcium mobilizer from intracellular stores. It has been demonstrated to be a new second messenger involved in the regulation of calcium homeostasis in many cell types, from plants to mammals. The number of physiological processes shown to be regulated by cADPR is steadily increasing. A topological paradox exists because ectocellularly generated cADPR acts intracellularly. Here we demonstrate that the catalytic functioning of CD38 is accompanied by a cADPR (cGDPR) -transporting activity across natural and artificial membranes. In resealed membranes from CD38(+) human erythrocytes, transport of catalytically generated cADPR or cGDPR was saturation dependent and occurred against a concentration gradient. Likewise, CD38-reconstituted proteoliposomes were active in concentrating NAD+ (NGD+) -derived cADPR (cGDPR) inside the vesicle compartment. Moreover, the cADPR-transporting activity in CD38 proteoliposomes prevented the hydrolase-catalyzed degradation to ADPR that occurs conversely with detergent-solubilized CD38, resulting in selective influx of cADPR. In the CD38 proteoliposomes, catalytically active CD38 exhibited monomeric, dimeric, and tetrameric structures. In CD38 sense- but not in antisense-transfected HeLa cells, externally added NAD+ resulted in significant, transient increases in cytosolic calcium. These data suggest that transmembrane juxtaposition of two or four CD38 monomers can generate a catalytically active channel for selective formation and influx of cADPR (cGDPR) to reach cADPR-responsive intracellular calcium stores.

ADP-ribosyl Cyclase↗
PubMed1997

CD38 and ADP-ribosyl cyclase catalyze the synthesis of a dimeric ADP-ribose that potentiates the calcium-mobilizing activity of cyclic ADP-ribose.

CD38, a lymphocyte differentiation antigen, is also a bifunctional enzyme catalyzing the synthesis of cyclic ADP-ribose (cADPR) from NAD+ and its hydrolysis to ADP-ribose (ADPR). An additional enzymatic activity of CD38 shared by monofunctional ADP-ribosyl cyclase from Aplysia californica is the exchange of the base group of NAD+ (nicotinamide) with various nucleophiles. Both human CD38 (either recombinant or purified from erythrocyte membranes) and Aplysia cyclase were found to catalyze the exchange of ADPR with the nicotinamide group of NAD+ leading to the formation of a dimeric ADPR ((ADPR)2). The dimeric structure of the enzymatic product, which was generated by recombinant CD38 and by CD38(+) Namalwa cells from as low as 10 microM NAD+, was demonstrated using specific enzyme treatments (dinucleotide pyrophosphatase and 5'-nucleotidase) and mass spectrometry analyses of the resulting products. The linkage between the two ADPR units of (ADPR)2 was identified as that between the N1 of the adenine nucleus of one ADPR unit and the anomeric carbon of the terminal ribose of the second ADPR molecule by enzymatic analyses and by comparison with patterns of cADPR cleavage with Me2SO:tert-butoxide. Although (ADPR)2 itself did not release Ca2+ from sea urchin egg microsomal vesicles, it specifically potentiated the Ca2+-releasing activity of subthreshold concentrations of cADPR. Therefore, (ADPR)2 is a new product of CD38 that amplifies the Ca2+-mobilizing activity of cADPR.

ADP-ribosyl Cyclase↗
PubMed1997

The CD38/cyclic ADP-ribose system: a topological paradox.

CD38 was first identified as a lymphocyte differentiation antigen that showed typical properties of an orphan receptor involved in many programs of cell proliferation and activation. However, CD38 proved also to be a bifunctional ectoenzyme that catalyzes the transient formation of cyclic ADP-ribose (cADPR) in a variety of cell types. This property raises many intriguing and so far unanswered questions, since cADPR is a new second messenger molecule directly involved in the control of calcium homeostasis by means of receptor-mediated release of calcium from ryanodine-sensitive intracellular stores. The relationship between receptor-like and enzymatic properties of CD38 is still unknown. The apparent topological paradox of ectocellular synthesis and intracellular activity of cADPR might be explained by: (a) influx of cADPR across the plasma membrane to reach its target stores, as suggested by experiments on cerebellar granule cells; and (b) NAD(+)-induced internalization, following membrane oligomerization, of CD38 with consequent partial import of cADPR metabolism to an intracellular compartment, as recently observed in lymphoid B cells. These two distinct mechanisms and other potential ones (e.g. binding of ectocellularly formed cADPR to cell surface receptors and initiation of signal-transducing pathways across the plasmamembrane) seem to be paradigmatic of processes affecting different types of cells. Although in some biological systems, such as Aplysia and sea urchin egg, cADPR metabolism is restricted to the intracellular environment, in mammalian cells the CD38/cADPR system provides new challenges in terms of subcellular compartmentation and qualifies as an unusual example of "ectobiochemistry" with potential, still unrecognized, properties of cellular regulation.

ADP-ribosyl Cyclase↗
PubMed1997

Isolated office hypertension and end-organ damage.

BACKGROUND: Patients with elevated blood pressure levels in the doctor's office but normal blood pressures at other times have recently been described as having 'isolated office hypertension' (IOH). There is debate concerning whether this condition is really benign and thus not in need of treatment. Most of the previous studies on this topic included patients who had already been administered antihypertensive treatment, which unavoidably alters their cardiovascular profile. OBJECTIVE: To evaluate whether recently discovered and never-treated patients with isolated office hypertension have structural or functional abnormalities in comparison with normotensive controls. METHODS: Patients included in the study underwent 24 h ambulatory blood pressure monitoring, M-mode echocardiography and high-resolution echography of carotid arteries. Parameters of lipid and carbohydrate metabolism were also determined. RESULTS: We investigated 76 patients (20 with IOH and 56 with sustained hypertension) who had recently been diagnosed hypertensive but never been administered antihypertensive treatment and 32 matched controls. No changes were detected in left ventricular mass (LVM h2.7, 41.5 +/- 11, 44.5 +/- 10 and 41.5 +/- 10 g/cm2.7 in IOH, sustained hypertension and controls, respectively) and in intimal-medial thickness (IMT, 0.54 +/- 0.13, 0.59 +/- 0.14 and 0.55 +/- 0.16 mm, respectively). However, the left ventricular diastolic function was significantly different (E/A = 1.08 +/- 0.3, 1.04 +/- 0.3 and 1.43 +/- 0.3, respectively, P = 0.02) and the carotid diameter significantly lower than that expected from the pressure-diameter relationship for normotensives. CONCLUSIONS: These results, at variance with those of others, suggest that IOH affects the cardiovascular system even during the early phases of the disease and indicate the need for prospective clinical trials to evaluate the benefit from early treatment of IOH patients.

Adult↗
PubMed1996

Ectocellular in vitro and in vivo metabolism of cADP-ribose in cerebellum.

CD38, a type II transmembrane glycoprotein predominantly expressed in blood cells, is a bifunctional ectoenzyme directly involved in the metabolism of cADP-ribose (cADPR). This is a potent Ca2+ mobilizer in several types of cells. The relationship between the ectocellular site of cADPR production and its intracellular calcium-related functions is poorly understood. Cultured rat cerebellar granule cells showed both enzymic activities of CD38, ADP-ribosyl cyclase and cADPR hydrolase, at a ratio of 16 to 1 respectively, and were immunostained by the anti-(human CD38) monoclonal antibody IB4. In these cells externally added cADPR and beta-NAD+ (the precursor of cADPR), but not alpha-NAD+ or ADP-ribose, enhanced the peak of the depolarization-induced rise in intracellular Ca2+ concentration. This effect was inhibited by 1 microM ryanodine, suggesting a potentiation of calcium-induced calcium release by cADPR. CD38 ectoenzyme activities, ADP-ribosyl cyclase and cADPR hydrolase, were also demonstrated in vivo by microdialysis of adult rat cerebellum, where IB4 bound to granule neurons selectively. Trace amounts (11.5 +/- 3.8 nM) of NAD+ were detected by microdialysis sampling and sensitive assays in the basal interstitial fluid of the cerebellum. These results provide a link between ectocellular cADPR turnover and intracellular calcium mobilization in cerebellum.

ADP-ribosyl Cyclase↗
PubMed1996

NAD+-dependent internalization of the transmembrane glycoprotein CD38 in human Namalwa B cells.

CD38 is a transmembrane glycoprotein involved as an orphan receptor in many physiological processes of lymphocytes. It is also a bifunctional enzyme that catalyzes at its ectocellular domain the synthesis from NAD+ (cyclase) and the hydrolysis (hydrolase) of the calcium-mobilizing metabolite cyclic ADP-ribose (cADPR). A still unexplained paradox concerns the relationship between ectocellular localization of CD38 and intracellular calcium-releasing activity of its intermediate product cADPR. Incubation of CD38+ human Namalwa B cells with external NAD+ elicited extensive membrane down-regulation of CD38 and its internalization in non-clathrin-coated vesicles. Since the internalized CD38 was demonstrated to be enzymatically active, this NAD+-dependent process is a hitherto unrecognized means for shifting cADPR metabolism from the cell surface to the intracellular environment.

ADP-ribosyl Cyclase↗
PubMed1995

Neutrophils activated by granulocyte-macrophage colony-stimulating factor express receptors for interleukin-3 which mediate class II expression.

Freshly isolated peripheral blood neutrophils, unlike monocytes and eosinophils, do not bind interleukin-3 (IL-3) or respond to IL-3). We show that neutrophils cultured for 24 hours in granulocyte-macrophage colony-stimulating factor (GM-CSF) express mRNA for the IL-3 receptor (R) alpha subunit, as shown by RNase protection assays, and IL-3R alpha chain protein, as shown by cytometric analysis using two different specific monoclonal antibodies. This effect was selective for GM-CSF, because granulocyte colony-stimulating factor, tumor necrosis factor-alpha, interferon-gamma, and IL-1 failed to induce the IL-3 receptor. Saturation binding curves with 125I-IL-3 and Scatchard transformation showed the presence of about 100 high-affinity and 4,000 low-affinity receptors. Because neutrophils have been shown to express human leukocyte antigen (HLA)-DR in response to GM-CSF, we examined the possibility that IL-3 could augment HLA-DR expression on GM-CSF-treated cells. We found that neutrophils incubated with 30 ng/mL IL-3 as well as 0.1 ng/mL GM-CSF expressed a mean of 2.1-fold higher levels of HLA-DR than with GM-CSF alone (P < .005), confirming the signaling competence of the newly expressed IL-3R. This increase was seen even at maximal concentrations of GM-CSF and IL-3 can have an additive effect on mature human cells. The augmentation of HLA-DR by IL-3 was specific because it could be inhibited by a blocking anti-IL-3R antibody. Expression of class II molecules by neutrophils under these conditions may have significance for antigen presentation. These results provide further evidence for the role of GM-CSF as an amplification factor in inflammation by inducing neutrophil responsiveness to IL-3 produced by T cells or mast cells.

Cells, Cultured↗
PubMed1995

Structural role of disulfide bridges in the cyclic ADP-ribose related bifunctional ectoenzyme CD38.

Human CD38, a type II cell surface glycoprotein, is a bifunctional ectoenzyme catalyzing both ADP-ribosyl cyclase and cyclic ADP-ribose (cADPR) hydrolase reactions. It shares a high degree of sequence homology with the cyclase from Aplysia species and studies of site-directed mutagenesis have recently demonstrated the importance, but not elucidated the role, of several cysteine residues highly conserved between these proteins. N-Ethylmaleimide, iodoacetamide and thiol-oxidizing reagents failed to affect either the cyclase or the weaker hydrolase activity of the Aplysia californica protein. Likewise, these reagents did not impair the two activities of CD38 purified from human erythrocytes. beta-mercaptoethanol had no effect on the Aplysia enzyme activities, while it inactivated both the cyclase and the cADPR hydrolase of CD38 by inducing its extensive oligomerization. In intact erythrocytes the beta-mercaptoethanol-dependent enzyme inactivation was completely prevented by prior cross-linking of the membrane proteins with glutaraldehyde. These data demonstrate that none of the cysteine residues plays any direct catalytic role in CD38 and Aplysia proteins, and that disulfide bridges are essential for maintaining the monomeric, catalytically active structure of CD38.

ADP-ribosyl Cyclase↗
PubMed1995

Self-aggregation of purified and membrane-bound erythrocyte CD38 induces extensive decrease of its ADP-ribosyl cyclase activity.

The transmembrane glycoprotein CD38 is a bifunctional enzyme that catalyzes at its ectocellular domain both the synthesis and the hydrolysis of cyclic ADP-ribose (cADPR). The complete reaction, converting NAD+ to nicotinamide and ADP-ribose, reproduces an NAD+glycohydrolase (NADase) reaction. CD38 purified from human erythrocyte membranes has been recently shown to undergo stable oligomerization induced by either NAD+ or beta-mercaptoethanol. We demonstrate that oligomerization is also triggered by reduced glutathione (GSH) and that the GSH-induced self-aggregation of purified CD38 is accompanied by extensive and comparable decrease of its ADP-ribosyl cyclase and NADase activities. GSH-induced oligomerization of CD38 and strong enzyme inactivation take place also in situ on erythrocyte membranes.

ADP-ribosyl Cyclase↗