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Biomedical subjects

L Guida

Publications and source records attributed to L Guida.

At least 19 recordsLinked to original sources
PubMed2006

Effects of Emdogain on osteoblast gene expression.

OBJECTIVE: Emdogain (EMD) is a protein extract purified from porcine enamel and has been introduced in clinical practice to obtain periodontal regeneration. EMD is composed mainly of amelogenins (90%), while the remaining 10% is composed of non-amelogenin enamel matrix proteins such as enamelins, tuftelin, amelin and ameloblastin. Enamel matrix proteins seem to be involved in root formation. EMD has been reported to promote proliferation, migration, adhesion and differentiation of cells associated with healing periodontal tissues in vivo. DESIGN: How this protein acts on osteoblasts is poorly understood. We therefore attempted to address this question by using a microarray technique to identify genes that are differently regulated in osteoblasts exposed to enamel matrix proteins. RESULTS: By using DNA microarrays containing 20,000 genes, we identified several upregulated and downregulated genes in the osteoblast-like cell line (MG-63) cultured with enamel matrix proteins (Emd). The differentially expressed genes cover a broad range of functional activities: (i) signaling transduction, (ii) transcription, (iii) translation, (iv) cell cycle regulation, proliferation and apoptosis, (v) immune system, (vi) vesicular transport and lysosome activity, and (vii) cytoskeleton, cell adhesion and extracellular matrix production. CONCLUSIONS: The data reported are the first genome-wide scan of the effect of enamel matrix proteins on osteoblast-like cells. These results can contribute to our understanding of the molecular mechanisms of bone regeneration and as a model for comparing other materials with similar clinical effects.

Animals↗
PubMed2005

In vitro cell-type specific biological response of human periodontally related cells to platelet-rich plasma.

OBJECTIVES: Platelet-rich plasma is a blood-derived fraction containing high concentrations of platelets and growth factors. Applied in the form of a gel on surgical wounds, it is able to stimulate hard and soft tissue repair and has been proposed for use in the field of periodontal regeneration. However to date, little is known about the biological interactions between platelet-rich plasma and periodontally related cells. In this study, we investigated the effects between platelet-rich plasma and cell populations involved in periodontal regeneration, namely primary human periodontal ligament cells, gingival fibroblasts and keratinocytes. MATERIAL AND METHODS: The proliferation of human periodontal ligament cells, gingival fibroblasts and keratinocytes by [3H]thymidine incorporation was assessed. The alkaline phosphatase activity and type I collagen levels of human periodontal ligament cells were also evaluated by a spectrophotometric assay and western blot analysis, respectively. RESULTS: Incubation of human periodontal ligament cells with platelet-rich plasma resulted in time-dependent growth stimulation (up to fourfold of control at 72 h). Likewise, an increase in the specific activity of alkaline phosphatase (fourfold at 6 days) and collagen (twofold at 7 days) was observed. Platelet-rich plasma also enhanced human gingival fibroblasts proliferation by twofold, whereas it inhibited human keratinocytes growth by 40%, with respect to their own controls at 72 h. CONCLUSION: Cell populations related to periodontal tissue were differently affected by platelet-rich plasma. In fact, a strong stimulation of human periodontal ligament cells proliferation, a minor increase in the growth rate of human gingival fibroblasts and a marked decrease of human keratinocytes proliferation were evident. In addition, in human periodontal ligament cells increased collagen and alkaline phosphatase activity levels were observed. These findings appear interesting in view of platelet-rich plasma utilization in periodontal regeneration.

Alkaline Phosphatase↗
PubMed2002

Serum cholesterol affects blood pressure regulation.

A close relationship between abnormalities of the lipid metabolism and arterial hypertension has been observed in several epidemiological studies. The aim of the present study was to investigate whether serum cholesterol might affect blood pressure (BP) levels at rest, during ambulatory monitoring or during sympathetic stimulation-independently of other variables such as body weight or serum insulin-thus influencing the outcome of hypertensive complications. Seventy-three patients with sustained newly-discovered and never-treated hypertension were divided into tertiles according to their serum cholesterol levels and their resting BP, 24-h BP and BP during isometric exercise (handgrip) were compared. Cardiac mass and carotid wall thickness were measured by echographic technique. The results were that tertiles were similar for body weight, blood glucose and serum insulin, but different for serum cholesterol and triglycerides. BP at rest and during 24-h monitoring was similar in the three groups, whilst a significant difference was detected during sympathetic stimulation by handgrip, with systolic and diastolic BP increasing by 16/12, 28/19 and 30/23 mm Hg (P < 0.01) in lower, medium and higher tertiles, respectively. Intima-media layer of the carotid arteries was also significantly thickened in the groups with higher cholesterol levels (0.54 +/- 0.07, 0.67 +/- 0.14, 0.68 +/- 0.15, P < 0.05). These data support the conclusion that even in patients with recently discovered hypertension, cholesterol levels may influence the BP response to adrenergic stimulation as well as the outcome of target organ disease.

Adolescent↗
PubMed2002

Clinic and ambulatory pulse pressure segregate a cluster of cardiovascular risk factors.

Office pulse pressure (PP) has been found to be independently associated with cardiovascular morbidity and mortality. While there is evidence that 24 h blood pressure (BP) is a better marker of the cardiovascular complications of hypertension than office BP, this information is still lacking in regard to PP. The aim of the present study was, therefore, to evaluate possible differences between office and 24 h PP in their relationship with cardiovascular risk profile. This cross-sectional study was performed in a group of 175 (104 M, 71 F) hypertensives (43 never treated before the study) free of clinical evidence of target organ disease. BP was measured at rest and during 24 h monitoring; cardiac structure and function was studied by ultrasounds; biochemical analyses were performed to evaluate some parameters of lipid and carbohydrate metabolism. Patients were divided into tertiles of office PP and of 24 h PP. Those in the highest tertile of PP had a significantly higher office and 24 h systolic BP along with a reduction in peripheral insulin sensitivity. Regarding cardiac structure and function, a significantly higher prevalence of concentric left ventricular hypertrophy (23 vs 55%; P=0.05) and an initial impairment of diastolic function with increase of the A wave was detected in hypertensives with higher PP. No difference between office and 24 h PP in detecting patients at higher cardiovascular risk was observed. In conclusion, office and 24 h pulse pressures were both able to segregate patients with a cluster of cardiovascular risk factors.

Adult↗
PubMed2001

The temperature-signaling cascade in sponges involves a heat-gated cation channel, abscisic acid, and cyclic ADP-ribose.

Sponges (phylum Porifera) are the phylogenetically oldest metazoan animals, their evolution dating back to 600 million years ago. Here we demonstrate that sponges express ADP-ribosyl cyclase activity, which converts NAD(+) into cyclic ADP-ribose, a potent and universal intracellular Ca(2+) mobilizer. In Axinella polypoides (Demospongiae, Axinellidae), ADP-ribosyl cyclase was activated by temperature increases by means of an abscisic acid-induced, protein kinase A-dependent mechanism. The thermosensor triggering this signaling cascade was a heat-activated cation channel. Elucidation of the complete thermosensing pathway in sponges highlights a number of features conserved in higher organisms: (i) the cation channel thermoreceptor, sensitive to heat, mechanical stress, phosphorylation, and anesthetics, shares all of the functional characteristics of the mammalian heat-activated background K(+) channel responsible for central and peripheral thermosensing; (ii) involvement of the phytohormone abscisic acid and cyclic ADP-ribose as its second messenger is reminiscent of the drought stress signaling pathway in plants. These results suggest an ancient evolutionary origin of this stress-signaling cascade in a common precursor of modern Metazoa and Metaphyta.

ADP-ribosyl Cyclase↗
PubMed2001

A self-restricted CD38-connexin 43 cross-talk affects NAD+ and cyclic ADP-ribose metabolism and regulates intracellular calcium in 3T3 fibroblasts.

Connexin 43 (Cx43) hexameric hemichannels, recently demonstrated to mediate NAD(+) transport, functionally interact in the plasma membrane of several cells with the ectoenzyme CD38 that converts NAD(+) to the universal calcium mobilizer cyclic ADP-ribose (cADPR). Here we demonstrate that functional uncoupling between CD38 and Cx43 in CD38-transfected 3T3 murine fibroblasts is paralleled by decreased [Ca(2+)](i) levels as a result of reduced intracellular conversion of NAD(+) to cADPR. A sharp inverse correlation emerged between [Ca(2+)](i) levels and NAD(+) transport (measured as influx into cells and as efflux therefrom), both in the CD38(+) cells (high [Ca(2+)](i), low transport) and in the CD38(-) fibroblasts (low [Ca(2+)](i), high transport). These differences were correlated with distinctive extents of Cx43 phosphorylation in the two cell populations, a lower phosphorylation with high NAD(+) transport (CD38(-) cells) and vice versa (CD38(+) cells). Conversion of NAD(+)-permeable Cx43 to the phosphorylated, NAD(+)-impermeable form occurs via Ca(2+)-stimulated protein kinase C (PKC). Thus, a self-regulatory loop emerged in CD38(+) fibroblasts whereby high [Ca(2+)](i) restricts further Ca(2+) mobilization by cADPR via PKC-mediated disruption of the Cx43-CD38 cross-talk. This mechanism may avoid: (i) leakage of NAD(+) from cells; (ii) depletion of intracellular NAD(+) by CD38; (iii) overproduction of intracellular cADPR resulting in potentially cytotoxic [Ca(2+)](i).

3T3 Cells↗
PubMed2001

Paracrine roles of NAD+ and cyclic ADP-ribose in increasing intracellular calcium and enhancing cell proliferation of 3T3 fibroblasts.

CD38 is a bifunctional ectoenzyme synthesizing from NAD(+) (ADP-ribosyl cyclase) and degrading (hydrolase) cyclic ADP-ribose (cADPR), a powerful universal calcium mobilizer from intracellular stores. Recently, hexameric connexin 43 (Cx43) hemichannels have been shown to release cytosolic NAD(+) from isolated murine fibroblasts (Bruzzone, S., Guida, L., Zocchi, E., Franco, L. and De Flora, A. (2001) FASEB J. 15, 10-12), making this dinucleotide available to the ectocellular active site of CD38. Here we investigated transwell co-cultures of CD38(+) (transfected) and CD38(-) 3T3 cells in order to establish the role of extracellular NAD(+) and cADPR on [Ca(2+)](i) levels and on proliferation of the CD38(-) target cells. CD38(+), but not CD38(-), feeder cells induced a [Ca(2+)](i) increase in the CD38(-) target cells which was comparable to that observed with extracellular cADPR alone and inhibitable by NAD(+)-glycohydrolase or by the cADPR antagonist 8-NH(2)-cADPR. Addition of recombinant ADP-ribosyl cyclase to the medium of CD38(-) feeders induced sustained [Ca(2+)](i) increases in CD38(-) target cells. Co-culture on CD38(+) feeders enhanced the proliferation of CD38(-) target cells over control values and significantly shortened the S phase of cell cycle. These results demonstrate a paracrine process based on Cx43-mediated release of NAD(+), its CD38-catalyzed conversion to extracellular cADPR, and influx of this nucleotide into responsive cells to increase [Ca(2+)](i) and stimulate cell proliferation.

3T3 Cells↗
PubMed2001

Nicotine effects on polymorphonuclear cell apoptosis and lipopolysaccharide-induced monocyte functions. A possible role in periodontal disease?

Apoptosis provides a mechanism for clearance of unwanted cells in a variety of situations in which programmed or physiological cell death occurs; but the premature death of defensive cells could promote infection, inflammation and concomitant disease. We detected high values of apoptosis in polymorphonuclear cells (PMN) elicited from crevicular sulci of smokers affected by adult periodontitis. To learn more about the effects of nicotine on the periodontal environment, we studied its ability to modulate the apoptosis of two phagocytic lines, PMN and mononuclear cells, which are continuously recruited from gingival vessels to prevent or control plaque extension. Brief exposure of PMN to nicotine concentrations ranging from 0.01 to 0.3% shortened, in a dose-dependent relationship, the lag culture time required to observe at fluorescent microscopy the morphological traces of apoptosis. These observations were confirmed by specific tools of apoptosis: DNA fragmentation on gel electrophoresis and expression of the apoptosis-signaling receptor Fas/Apo-1. The apoptotic effect excited by nicotine on these first line defensive cells may be an important feature of the pathogenesis of periodontal disease. As for mononuclear leukocytes, nicotine was unable to induce apoptotic modifications on cells observed up to 72 h culture time, but the drug inhibited IL-1beta release and procoagulant activity (PCA) expression. The conflicting role played by these lipopolysaccharide (LPS)-induced monocyte functions in the inflammatory process is a further intrigue in the mechanism by which nicotine compromises the oral health.

Apoptosis↗
PubMed2001

Extracellular cyclic ADP-ribose potentiates ACh-induced contraction in bovine tracheal smooth muscle.

Cyclic ADP-ribose (cADPR), a universal calcium releaser, is generated from NAD(+) by an ADP-ribosyl cyclase and is degraded to ADP-ribose by a cADPR hydrolase. In mammals, both activities are expressed as ectoenzymes by the transmembrane glycoprotein CD38. CD38 was identified in both epithelial cells and smooth myocytes isolated from bovine trachea. Intact tracheal smooth myocytes (TSMs) responded to extracellular cADPR (100 microM) with an increase in intracellular calcium concentration ([Ca(2+)](i)) both at baseline and after acetylcholine (ACh) stimulation. The nonhydrolyzable analog 3-deaza-cADPR (10 nM) elicited the same effects as cADPR, whereas the cADPR antagonist 8-NH(2)-cADPR (10 microM) inhibited both basal and ACh-stimulated [Ca(2+)](i) levels. Extracellular cADPR or 3-deaza-cADPR caused a significant increase of ACh-induced contraction in tracheal smooth muscle strips, whereas 8-NH(2)-cADPR decreased it. Tracheal mucosa strips, by releasing NAD(+), enhanced [Ca(2+)](i) in isolated TSMs, and this increase was abrogated by either NAD(+)-ase or 8-NH(2)-cADPR. These data suggest the existence of a paracrine mechanism whereby mucosa-released extracellular NAD(+) plays a hormonelike function and cADPR behaves as second messenger regulating calcium-related contractility in TSMs.

ADP-ribosyl Cyclase↗
PubMed2001

Insulin resistance, ventricular mass and function in normoglycaemic hypertensives.

BACKGROUND AND AIM: To investigate the possible relationship between insulin sensitivity and the main non-invasively determined parameters of left ventricular structure and function in an early phase of hypertensive disease. METHODS AND RESULTS: We studied a group of 60 patients with previously untreated primary arterial hypertension of recent onset. Insulin sensitivity was evaluated by means of homeostasis model assessment (HOMA), and cardiac structure and function by means of ultrasonography. The patients were subsequently divided on the basis of HOMA tertiles and the between-group differences were statistically evaluated using one-way analysis of variance. No difference was found in the main demographic parameters, but high-density lipoprotein (HDL)-cholesterol was significantly higher (1.43 +/- 0.4, 1.16 +/- 0.2, 1.16 +/- 0.2 mmol/L; p < 0.05) and total triglycerides significantly lower (0.94 +/- 0.3, 1.41 +/- 0.8, 1.54 +/- 0.7 mmol/L; p < 0.05) in the first tertile. In relation to the non-invasive parameters of diastolic function, the E wave was significantly reduced in the third tertile (0.64 +/- 0.1, 0.63 +/- 0.1, 0.52 +/- 0.1 m/s; p < 0.05), with a concomitant significant increase in AFF (0.51 +/- 0.03, 0.51 +/- 0.06, 0.56 +/- 0.03; p < 0.05). The E wave (r = 0.326, p < 0.05), E/A wave ratio (r = -0.277, p < 0.05) and AFF (r = 0.432, p < 0.001) were significantly related to HOMA, regardless of age, body mass index, heart rate, left ventricular mass and blood pressure. CONCLUSIONS: Our results indicate that hypertensive patients in the third HOMA tertile (i.e. those with reduced insulin sensitivity) have early diastolic left ventricular abnormalities.

Adolescent↗
PubMed2000

Connexin 43 hemi channels mediate Ca2+-regulated transmembrane NAD+ fluxes in intact cells.

A previously unrecognized passive transport for pyridine dinucleotides has been described recently in the plasmamembrane of several mammalian cells. Despite elucidation of some functional and kinetic properties of this transport system, it is still undefined at the molecular level. Therefore, we have addressed the molecular characterization of the NAD+ transporter and identified it as connexin 43 (Cx43). This is a structural component of hexameric hemichannels that, when juxtaposed on adjacent cells, builds up intercellular gap junctions and mediates exchange of molecules between cells. However, the role of connexin hemichannels as potential pores in individual, noncoupled cells remains elusive. Bidirectional NAD+ transport in isolated Cx43-expressing mur ine 3T3 fibroblasts was affected by known modulators of connexin-mediated intercellular coupling and was completely inhibited by treatment of the cells with a Cx43-antisense oligonucleotide. NAD+ transport in proteoliposomes reconstituted with 3T3 membrane proteins was inhibited in the presence of a monoclonal anti-Cx43 antibody. Finally, Cx43 immunopurified to homogeneity was reconstituted in unilamellar proteoliposomes, which displayed full NAD+-transporting activity. This finding is the first evidence that connexin hemichannels can mediate transmembrane fluxes of a nucleotide in whole cells: The pleiotropy of NAD+-dependent cellular events, including redox reactions, signaling, and DNA repair, implicates Cx43 hemichannels in intercellular NAD+ trafficking, which suggests new paracrine functions of NAD.

3T3 Cells↗
PubMed2000

Olive oil and reduced need for antihypertensive medications.

BACKGROUND: The blood pressure (BP) effects of changing the total fat intake and saturated-unsaturated fat ratio are still controversial, despite evidence that saturated fat-enriched diets are associated with higher BP levels. This double-blind, randomized crossover study evaluated a possible difference between antihypertensive effects of monounsaturated (MUFA) (extra-virgin olive oil) and polyunsaturated fatty acids (PUFA) (sunflower oil). METHODS: Twenty-three hypertensive patients were assigned randomly to MUFA or PUFA diet for 6 months and then crossed over to the other diet; effects were evaluated on the basis of daily antihypertensives needed. RESULTS: Diets high in MUFA and PUFA differed from the habitual diet for reduced total and saturated fats, whereas they differed from each other for MUFA (17.2% vs 10.5%) and PUFA content (3.8% vs 10.5%). Resting BP was significantly lower (P = .05 for systolic BP; P = .01 for diastolic BP) at the end of the MUFA diet compared with the PUFA diet. Blood pressure responses during sympathetic stimulation with the cold pressor test and isometric exercise were similar. Daily drug dosage was significantly reduced during the MUFA but not the PUFA diet (-48% vs - 4%, P<.005). All patients receiving the PUFA diet required antihypertensive treatment, whereas 8 of those receiving the MUFA diet needed no drug therapy. CONCLUSIONS: A slight reduction in saturated fat intake, along with the use of extra-virgin olive oil, markedly lowers daily antihypertensive dosage requirement, possibly through enhanced nitric oxide levels stimulated by polyphenols.

Adult↗
PubMed2000

Extracellular cyclic ADP-ribose increases intracellular free calcium concentration and stimulates proliferation of human hemopoietic progenitors.

Cyclic ADP-ribose (cADPR) is a universal second messenger that regulates many calcium-related cellular events by releasing calcium from intracellular stores. Since these events include enhanced cell proliferation and since the bone marrow harbors both ectoenzymes that generate cADPR from NAD(+) (CD38 and BST-1), we investigated the effects of extracellular cADPR on human hemopoietic progenitors (HP). Exposure of HP to 100 microM cADPR for 24 h induced a significant increase in colony output (P<0.01) and colony size (P<0.003). A horizontal expansion of HP, as demonstrated by a markedly increased replating efficiency in semisolid medium (up to 700 times compared to controls), was also observed, indicating that cADPR priming can affect cell growth for multiple generations over several weeks after exposure. Influx of extracellular cADPR into the cells was demonstrated, and a causal relationship between the functional effects and the increase of intracellular free calcium concentration induced by cADPR on HP was established through the use of specific antagonists. Similar effects on HP were produced by nanomolar concentrations of the nonhydrolyzable cADPR analog 3-deaza-cADPR. These data demonstrate that extracellular cADPR behaves as a cytokine enhancing the proliferation of human HP, a finding that may have biomedical applications for the ex vivo expansion of hemopoietic cells.

Adenosine Diphosphate Ribose↗
PubMed2000

Topology of CD38.

Explore the source record for details and available documents.

ADP-ribosyl Cyclase↗
PubMed1999

Sex-related influence of body size and sexual maturation on blood pressure in adolescents.

OBJECTIVE: This study is aimed at investigating the influence of body size, body fat and sexual maturation on blood pressure (BP) in adolescents. DESIGN: A cross-sectional study. SETTING: A suburban student population of Southern Italy. SUBJECTS: One hundred ninety students attending the first and second year of a secondary school. Five were excluded because they were affected by major diseases. The remaining were 98 M and 87 F (mean age for either group = 12.0+/-0.8 y). METHODS: Blood pressure was measured by a mercury sphygmomanometer, body weight by a platform beamscale, other measurements included height, biceps, triceps, subscapular and suprailiac skinfolds by a caliper; sexual maturation was evaluated according to Tanner. RESULTS: Body size was greater than in Tanner's population: in particular body weight (but not height) in our sample markedly exceeded that of the children of the same age in Tanner's population. Boys had higher systolic blood pressure (SBP) than girls (BP = 109/64+/-12/10 vs. 103/63+/-11/8 mm Hg, P<0.02 for SBP), while heart rate and waist/hip ratio were lower. During puberty evaluated on the basis of pubic hair growth BP in girls was higher than in the prepubertal phase (107/66+/-9/7 vs. 99/61+/-10/7, P<0.01). Pubertal boys showed a reduced percent of body fat (calculated from four skinfold measurements) in comparison to prepubertal ones (21.0%+/-4.5 vs. 24.5%+/-7.1, P<0.01). In linear correlation analysis, height, BW, BMI and lean body mass were found to be significantly associated with SBP in both sexes and to diastolic blood pressure (DBP) in girls. Percent body fat was correlated with SBP in boys, while sexual maturation was associated to SBP and DBP in girls only. Multiple regression analysis indicated a significant contribution of body size to BP variability, particularly in the girls. Sexual maturation was excluded from the final regression equations when height, BW or lean body mass were present. CONCLUSIONS: These data indicate that body weight in these adolescents is greater that in Tanner's population of the same age and sex. Body size appears to be a major determinant of BP, whereas sexual maturation seems to influence BP levels mainly through body growth. The influence of percent body fat on BP setting seems to be of limited importance.

Adolescent↗
PubMed1999

Ligand-induced internalization of CD38 results in intracellular Ca2+ mobilization: role of NAD+ transport across cell membranes.

CD38, a transmembrane glycoprotein widely expressed in vertebrate cells, is a bifunctional ectoenzyme catalyzing the synthesis and hydrolysis of cyclic ADP-ribose (cADPR). cADPR is a universal second messenger that releases calcium from intracellular stores. Since cADPR is generated by CD38 at the outer surface of many cells, where it acts intracellularly, increasing attention is paid to addressing this topological paradox. Recently, we demonstrated that CD38 is a catalytically active, unidirectional transmembrane transporter of cADPR, which then reaches its receptor-operated intracellular calcium stores. Moreover, CD38 was reported to undergo a selective and extensive internalization through non clathrin-coated endocytotic vesicles upon incubating CD38(+) cells with either NAD+ or thiol compounds: these endocytotic vesicles can convert cytosolic NAD into cADPR despite an asymmetric unfavorable orientation that makes the active site of CD38 intravesicular. Here we demonstrate that the cADPR-generating activity of the endocytotic vesicles results in remarkable and sustained increases of intracellular free calcium concentration in different cells exposed to either NAD+, or GSH, or N-acetylcysteine. This effect of CD38-internalizing ligands on intracellular calcium levels was found to involve a two-step mechanism: 1) influx of cytosolic NAD+ into the endocytotic vesicles, mediated by a hitherto unrecognized dinucleotide transport system that is saturable, bidirectional, inhibitable by 8-N3-NAD+, and characterized by poor dinucleotide specificity, low affinity, and high efficiency; 2) intravesicular CD38-catalyzed conversion of NAD+ to cADPR, followed by outpumping of the cyclic nucleotide into the cytosol and subsequent release of calcium from thapsigargin-sensitive stores. This unknown intracellular trafficking of NAD+ and cADPR based on two distinctive and specific transmembrane carriers for either nucleotide can affect the intracellular calcium homeostasis in CD38(+) cells.

3T3 Cells↗