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Biomedical subjects

M Innocenti

Publications and source records attributed to M Innocenti.

At least 19 recordsLinked to original sources
PubMed2002

Helicobacter pylori-induced activation of human endothelial cells.

Helicobacter pylori infection causes active chronic inflammation with a continuous recruitment of neutrophils to the inflamed gastric mucosa. To evaluate the role of endothelial cells in this process, we have examined adhesion molecule expression and chemokine and cytokine production from human umbilical vein endothelial cells stimulated with well-characterized H. pylori strains as well as purified proteins. Our results indicate that endothelial cells actively contribute to neutrophil recruitment, since stimulation with H. pylori bacteria induced upregulation of the adhesion molecules VCAM-1, ICAM-1, and E-selectin as well as the chemokines interleukin 8 (IL-8) and growth-related oncogene alpha (GRO-alpha) and the cytokine IL-6. However, there were large variations in the ability of the different H. pylori strains to stimulate endothelial cells. These interstrain variations were seen irrespective of whether the strains had been isolated from patients with duodenal ulcer disease or asymptomatic carriers and were not solely related to the expression of known virulence factors, such as the cytotoxin-associated gene pathogenicity island, vacuolating toxin A, and Lewis blood group antigens. In addition, one or several unidentified proteins which act via NF-kappaB activation seem to induce endothelial cell activation. In conclusion, human endothelial cells produce neutrophil-recruiting factors and show increased adhesion molecule expression after stimulation with certain H. pylori strains. These effects probably contribute to the continuous recruitment of neutrophils to H. pylori-infected gastric mucosa and may also contribute to tissue damage and ulcer formation.

Adult↗
PubMed2001

An effector region in Eps8 is responsible for the activation of the Rac-specific GEF activity of Sos-1 and for the proper localization of the Rac-based actin-polymerizing machine.

Genetic and biochemical evidence demonstrated that Eps8 is involved in the routing of signals from Ras to Rac. This is achieved through the formation of a tricomplex consisting of Eps8-E3b1-Sos-1, which is endowed with Rac guanine nucleotide exchange activity. The catalytic subunit of this complex is represented by Sos-1, a bifunctional molecule capable of catalyzing guanine nucleotide exchange on Ras and Rac. The mechanism by which Sos-1 activity is specifically directed toward Rac remains to be established. Here, by performing a structure-function analysis we show that the Eps8 output function resides in an effector region located within its COOH terminus. This effector region, when separated from the holoprotein, activates Rac and acts as a potent inducer of actin polymerization. In addition, it binds to Sos-1 and is able to induce Rac-specific, Sos-1-dependent guanine nucleotide exchange activity. Finally, the Eps8 effector region mediates a direct interaction of Eps8 with F-actin, dictating Eps8 cellular localization. We propose a model whereby the engagement of Eps8 in a tricomplex with E3b1 and Sos-1 facilitates the interaction of Eps8 with Sos-1 and the consequent activation of an Sos-1 Rac-specific catalytic ability. In this complex, determinants of Eps8 are responsible for the proper localization of the Rac-activating machine to sites of actin remodeling.

Actins↗
PubMed2001

Cloning and characterization of mouse UBPy, a deubiquitinating enzyme that interacts with the ras guanine nucleotide exchange factor CDC25(Mm)/Ras-GRF1.

We used yeast "two-hybrid" screening to isolate cDNA-encoding proteins interacting with the N-terminal domain of the Ras nucleotide exchange factor CDC25(Mm). Three independent overlapping clones were isolated from a mouse embryo cDNA library. The full-length cDNA was cloned by RACE-polymerase chain reaction. It encodes a large protein (1080 amino acids) highly homologous to the human deubiquitinating enzyme hUBPy and contains a well conserved domain typical of ubiquitin isopeptidases. Therefore we called this new protein mouse UBPy (mUBPy). Northern blot analysis revealed a 4-kilobase mRNA present in several mouse tissues and highly expressed in testis; a good level of expression was also found in brain, where CDC25(Mm) is exclusively expressed. Using a glutathione S-transferase fusion protein, we demonstrated an "in vitro" interaction between mUBPy and the N-terminal half (amino acids 1-625) of CDC25(Mm). In addition "in vivo" interaction was demonstrated after cotransfection in mammalian cells. We also showed that CDC25(Mm), expressed in HEK293 cells, is ubiquitinated and that the coexpression of mUBPy decreases its ubiquitination. In addition the half-life of CDC25Mm protein was considerably increased in the presence of mUBPy. The specific function of the human homolog hUBPy is not defined, although its expression was correlated with cell proliferation. Our results suggest that mUBPy may play a role in controlling degradation of CDC25(Mm), thus regulating the level of this Ras-guanine nucleotide exchange factor.

3T3 Cells↗
PubMed2001

Orally administered bovine lactoferrin systemically inhibits VEGF(165)-mediated angiogenesis in the rat.

Lactoferrin (Lf) systemically suppresses tumor growth and metastasis by unknown mechanisms. We have studied the effect of orally administered iron-unsaturated bovine Lf on angiogenesis induced by VEGF(165) and IL-1-alpha in adult rats using the mesenteric-window angiogenesis assay. VEGF(165) is a major angiogenic factor in most, if not all, tumors and other angiogenesis diseases of clinical relevance. A number of objective angiogenesis variables were analyzed using microscopic morphometry and image analysis. Lf treatment significantly inhibited the VEGF(165)-mediated response in terms of microvessel spatial extension, overall vascularity and incidence of crossover. The response to IL-1-alpha decreased significantly only in terms of microvessel crossover. In vitro, Lf exerted an antiproliferative effect on endothelial cells. To our knowledge, Lf is the first endogenous protein that has been shown to be antiangiogenic following oral administration. The oral administration of Lf thus appears to be of potential interest as an antiangiogenesis treatment modality in the clinical setting. Since tumor growth is angiogenesis dependent, the extensive therapeutic potential warrants further study to elucidate the mechanisms responsible for the angiostatic effect of Lf.

Administration, Oral↗
PubMed2001

Skeletal reconstruction with a free vascularized fibula graft associated to bone allograft after resection of malignant bone tumor of limbs.

Over the last twenty years, progress in diagnosis and in adjuvant therapy in the field of malignant bone tumor treatment has allowed for development of limb-saving surgical techniques after oncological excision. In this context, the use of vascularized fibula for transplantation represents an important instrument in the reconstruction of bone, either with or without allografts.Moreover, in pediatric cases, the vascularized transplant of the proximal fibula with its open physis allows for an adequate reconstruction of the bone loss and the possibility of conserving the growth potential of the segment. The purpose of this article is to illustrate the various reconstructive possibilities that the use of the combined graft technique (VFT plus allograft) offers in the treatment of large-scale bone loss. In our department from 1988 to 2000, 142 vascularized fibula transplants were performed in oncological cases. Surgical reconstruction was carried out on the tibia in 70 cases, on the femur in 40, on the humerus in 26 and on the radius in 6. Combined graft intercalary reconstructions were 92. In 22 pediatric cases the fibula was transplanted, including the proximal growing epiphysis in the graft; in two of these cases massive allograft was associated to the VFG. Because of its biological properties, the grafted vascularized fibula allowed for fast bone fusion at the level of the osteotomy. It has also demonstrated a tendency of progressive hypertrophy and osteointegration with the allograft, when used. In 22 pediatric cases, the fibula graft with the proximal epiphysis maintained its ability to grow. Unsuccessful outcomes caused by vascular, mechanical, or septic failure were equal to 8.2 %. The fibula graft in the reconstruction of bone loss secondary to oncological excision is a trustworthy and versatile technique.

Adolescent↗
PubMed2001

Helicobacter pylori lipopolysaccharides preferentially induce CXC chemokine production in human monocytes.

Helicobacter pylori infection can cause duodenal ulcers and may also induce gastric adenocarcinoma. The bacteria colonize the gastric mucosa and areas of gastric metaplasia in the duodenum for decades, resulting in active chronic inflammation in the infected areas. A characteristic feature of the infection is the ongoing recruitment of neutrophils to the infected sites. To evaluate the role of H. pylori lipopolysaccharides (LPS) in the recruitment of leukocytes to the gastric mucosa, we have examined the cytokine and chemokine production from human monocytes stimulated with LPS isolated from different H. pylori strains, as well as from several other gram-negative bacteria. Our results show that H. pylori LPS induce a large production of neutrophil-recruiting CXC chemokines (interleukin-8 and growth-related oncogene alpha) from purified human monocytes, to almost the same extent as Escherichia coli LPS. However, and in agreement with previous studies, H. pylori LPS was much less potent in inducing production of proinflammatory cytokines by purified human monocytes and was also a weak inducer of the CC chemokine RANTES. There was no difference between LPS preparations from different H. pylori strains in their ability to induce cytokines and chemokines. The preferential production of CXC chemokines after stimulation with H. pylori LPS indicates an important contribution of this molecule in maintaining neutrophil recruitment during the infection, irrespective of the infecting strain.

Chemokines, CXC↗
PubMed2000

Signaling from Ras to Rac and beyond: not just a matter of GEFs.

Members of a family of intracellular molecular switches, the small GTPases, sense modifications of the extracellular environment and transduce them into a variety of homeostatic signals. Among small GTPases, Ras and the Rho family of proteins hierarchically and/or coordinately regulate signaling pathways leading to phenotypes as important as proliferation, differentiation and apoptosis. Ras and Rho-GTPases are organized in a complex network of functional interactions, whose molecular mechanisms are being elucidated. Starting from the simple concept of linear cascades of events (GTPase-->activator--> GTPase), the work of several laboratories is uncovering an increasingly complex scenario in which upstream regulators of GTPases also function as downstream effectors and influence the precise biological outcome. Furthermore, small GTPases assemble into macromolecular machineries that include upstream activators, downstream effectors, regulators and perhaps even final biochemical targets. We are starting to understand how these macromolecular complexes work and how they are regulated and targeted to their proper subcellular localization. Ultimately, the acquisition of a cogent picture of the various levels of integration and regulation in small GTPase-mediated signaling should define the physiology of early signal transduction events and the pathological implication of its subversion.

Animals↗
PubMed1999

CDC25(Mm)/Ras-GRF1 regulates both Ras and Rac signaling pathways.

The Ras-GRF1 exchange factor molecule contains in addition to the catalytic domain two pleckstrin homology (PH1 and PH2), one IQ and one Dbl homology (DH) domains. In this study we investigated the role of such additional domains. We found that a Ras-GRF1 mutant lacking PH1 and IQ domains is sufficient to activate c-fos promoter in response to lysophosphatidic acid (LPA). The same mutant did not increase external stimuli-regulated kinase (ERK) activity, suggesting an additional mechanism for the induction of gene transcription. Isolated DH-PH2 module activates c-Jun NH(2)-terminal kinase and the c-fos promoter in response to LPA, providing the basis for an ERK-independent mechanism. These results provide evidence that Ras-GRF1 acts as a bifunctional molecule on both ERK-dependent and independent pathways.

3T3 Cells↗
PubMed1999

Confocal and scanning electron microscopic study of teeth restored with fiber posts, metal posts, and composite resins.

Forty-two single-rooted lower premolars, extracted for periodontal reasons, were endodontically treated and divided into 7 groups of 6 teeth each. In five of the groups, three different types of carbon fiber posts, quartz fiber posts, and titanium posts were used in combination with All Bond 2 dental adhesive. In two groups, two types of carbon fiber posts were also cemented with Panavia 21 dental adhesive. After 3 wk storage in saline, the teeth were longitudinally sectioned; one half was observed using confocal microscopy and the other by scanning electron microscopy. The specimens were evaluated for the presence of a resin dentin interdiffusion zone for the presence of voids at post-resin-dentin interfaces and for the determination of the fiber posts' structure. Upon examination with the confocal microscope, the interfaces of teeth restored with All Bond 2 showed a higher percentage (p < 0.05) of resin dentin interdiffusion zone than those treated with panavia. The fiber size and the post structure were similar in all the fiber posts observed. Some voids were present inside the fiber post structure.

Bicuspid↗
PubMed1999

Low risk of Lyme borreliosis in a protected area on the Tyrrhenian coast, in central Italy.

A comprehensive Lyme borreliosis risk assessment process was applied in S. Rossore Estate, on the Tyrrhenian coast, near Pisa, Italy. Host-seeking Ixodes ricinus nymphs peaked in May in oak-dominated deciduous wood (median, Q1-Q3, number of nymphs/50 m dragging = 4.5, 2.5-8), whereas host-seeking larvae peaked in August in the same habitat type (6.0, 4-17/50 m dragging). Prevalence of I. ricinus infestation was 88.9% in wild rodents (n = 11), 64.3% in fallow deer (n = 28) and 0.0% in wild boars (n = 5). Borrelia burgdorferi sensu lato was not isolated from rodents' organs, and from 80 I. ricinus nymphs and 50 adults. Moreover, PCR for B. burgdorferi sl carried out on 110 nymphs and 12 adult ticks also gave negative results. Forest workers were at higher risk of tick bite than other Estate employees (relative risk (RR): 1.7, p = 0.02). In spite of high levels of tick exposure, B. burgdorferi sl specific antibodies were not detected in sera from Estate personnel (n = 30) and sentinel animals (dogs, n = 23, fallow deer, n = 61).

Animals↗
PubMed1999

Dye leakage and SEM study of roots obturated with Thermafill and dentin bonding agent.

The apical seal of roots obturated with a dentin bonding agent and Thermafill with and without the use of sodium hypochlorite as an irrigating solution was compared by a dye leakage test. Roots obturated with Thermafill and a zinc oxide eugenol sealer were used as controls. Thirty-eight roots were prepared chemomechanically and divided into three experimental groups. The teeth of group 1 were filled with Thermafill and the dentin bonding agent using sodium hypochlorite as the irrigant. The teeth of group 2 were filled in the same way, but saline solution was used as the irrigant. Before the root canal was filled the smear layer was removed from the root canal walls of both groups by rinsing the root canal with a 17% EDTA solution. The teeth of group 3 were filled with Thermafill and a zinc oxide eugenol sealer. The teeth were immersed in 2% methylene blue solution. The root fillings of groups 1 and 2 leaked significantly more than those of group 3. The resin-dentin-guttapercha interface of group 1 was observed by scanning electron microscopy and showed a typical hybrid layer. An intimate contact between resin and dentin was present in group 2, but a resin-dentin interdiffusion zone was only occasionally observed. The use of dental adhesives and the hybrid layer formation did not improve the seal of Thermafill root canal fillings.

Coloring Agents↗
PubMed1999

Intraepiphyseal resection of the proximal tibia and its impact on lower limb growth.

From 1989 through 1996, 10 children affected by high grade bone tumors of the proximal tibia underwent an intraepiphyseal intercalary resection. The residual epiphyseal bone segment measured less than 2 cm in thickness in all cases and reconstruction always was performed using the combination of a vascularized fibular autograft and a massive bone allograft. The proximal epiphyseal osteosynthesis was fixed by small fragment screws. The aim of this study was to report the growth pattern of the residual proximal tibial epiphysis and to evaluate any possible lower limb discrepancy and/or deformity after the end of skeletal maturity. At current followup six patients were available for the final evaluation. Radiographic documentation included computed tomography scan of both knees before surgery, a panoramic radiographic view and a computed tomography scan of both lower limbs after the end of skeletal growth. The length of both femurs and tibias, the size of the tibial plateau and of the opposite distal femur, and any possible deformity of femur or tibia were measured and compared with the preoperative data. No patient had a limb length discrepancy greater than 3.5 cm. In all cases the ipsilateral femur had a valgus deformity of the hip develop. In two patients this deformity was associated with an elongation of the femur, partially compensating for the shortening of the tibia. The tibial plateau close to reconstruction grew less than the contralateral one (range 2%-8%) but maintained its normal relationship with the distal femur. None of these patients reported any restriction in recreational activities. They could walk, run, and jump. Their functional result according the International Society of Limb Salvage functional grading system was satisfactory in all cases.

Bone Neoplasms↗
PubMed1998

Stereomicroscopic and scanning electron microscopic study of roots obturated with vertically condensed gutta-percha, epoxy resin cement, and dentin bonding agent.

Root canal fillings performed with dentin bonding agent, gutta-percha, and epoxy resin sealer were observed by stereomicroscope. Fourteen palatal roots of extracted maxillary molars were used. Seven roots (group 1) were filled with gutta-percha, epoxy resin sealer, and dentin bonding agent. Seven (group 2) were filled with gutta-percha and epoxy resin sealer alone. The presence of these materials (gutta-percha, adhesive, cement, and a combination of these materials) and the number of voids present were evaluated in the apical, middle, and coronal thirds of the canals. In group 1, a combination of the three materials used was found to be the statistically most frequent case only in the apical third. No statistically significant difference was found between the overall number of voids present in the two groups. The resin-dentin-gutta-percha interface of roots from group 1 was observed by scanning electron microscopy: no statistically significant difference was found between apical and coronal sections regarding the presence of hybrid layer, penetration of resin into the dentinal tubules, or the presence of gaps in either the dentin-resin interface or the resin-gutta-percha interface.

Bismuth↗
PubMed1998

Free vascularized growth-plate transfer after bone tumor resection in children.

Limb-salvage surgery is the standard care for most malignant tumors affecting the extremities, and a vascularized fibula transfer is probably the most popular microsurgical option to reconstruct long-bone defects. Skeletal reconstruction after bone-tumor resection involving the metepiphysis of a growing child can be successfully achieved with a vascularized fibula graft incorporating the proximal physis and active growth plate. Such a procedure has been utilized in 12 children under the age of 10 years who had malignant bone tumors located in the upper limb (3 in the distal radius, 9 in the proximal humerus). The follow-up ranged between 4 years and 3 months. Ten grafts were supplied by the anterior tibial artery, and two by the peroneal artery. The average growth rate of the grafts based on the former artery has been more than 1 cm per year, ranging between 0.75 and 1.33 cm. The authors describe a modified operative technique and discuss the clinical results of the procedure which offers a satisfactory skeletal reconstruction and prevents future limb-size discrepancy.

Bone Neoplasms↗
PubMed1997

Modeling arterial hypotension during hemodialysis.

A mathematical model of the hemodynamic response to hemodialysis is presented. This model includes the dynamics of sodium, urea, and potassium in the intracellular and extracellular pools; fluid balance equations for the intracellular, interstitial, and plasma volumes; systemic and pulmonary hemodynamics; and the action of several short-term arterial pressure control mechanisms. The control mechanisms are triggered by information coming from both arterial and cardiopulmonary pressoreceptors, and they work on systemic arterial resistance, heart rate, and systemic venous unstressed volume. Moreover, the model hypothesizes that decreasing left atrial pressure below a given threshold causes a paradoxical withdrawal of the sympathetic drive and a consequent vasodepressor syncope. The model is used to simulate the pattern of the main hemodynamic quantities (systemic arterial pressure, heart rate, total systemic resistance, and cardiac output) during hemodialysis in several groups of patients (both hypotension resistant and hypotension prone) whose data were drawn from the clinical literature. The simulation results point out that the model is able to reproduce a variety of different conditions, including no hypotension, moderate hypotension, and severe hypotension with ultimate vasodepressor syncope, by adjusting a few parameters with clear physiological meanings. Hypotension is principally imputed to a loss of the sympathetic mechanisms working on systemic resistance and to an impairment of vascular refilling at the capillary wall. The results suggest that hypotension during hemodialysis is a complex phenomenon that depends on the superimposition of several concomitant factors working together that can lead to a variety of distinct individual patterns.

Baroreflex↗
PubMed1997

Mathematical investigation of some physiological factors involved in hemodialysis hypotension.

A previously developed mathematical model is used to investigate the role of some hemodynamic, regulatory, and osmotic factors in the development of symptomatic hypotension during hemodialysis. Sensitivity analysis of the model parameters suggests that a decrease in atrial pressure, with a consequent fall in cardiac output (Frank-Starling mechanism), is the primary hemodynamic perturbation induced by ultrafiltration. Also, during the first hours of a hemodialysis session, the sympathetic mechanism working on systemic resistance is the main factor responsible for arterial pressure maintenance, and the physiological response is probably characterized by a prevalence of the cardiopulmonary over the arterial baroreflex control. During this period, a decrease in plasma osmolarity, caused mainly by urea removal, may contribute to the reduction of vascular refilling. During the last hours of the session, the arterial pressure level is also significantly affected by other factors that influence vascular refilling and mean circulatory filling pressure (systemic compliance; action of feedback mechanisms working on venous unstressed volume; plasma oncotic pressure; and, especially, capillary wall permeability and interstitial space elastance). Simulation of hemodialysis with different modalities emphasizes the importance of avoiding high ultrafiltration rates and of maintaining the sodium concentration in the dialysate close to the sodium concentration of the extracellular fluid to limit the risk of symptomatic hypotension. Higher values of sodium in the dialysate are, however, associated with poor sodium removal from the extracellular pool with risks of interdialytic morbidity. In the future, the model may be used to optimize the ultrafiltration rate and sodium profile in the dialysate according to individual patient prescriptions.

Baroreflex↗