Biomedical subjects

G Zambetti

Publications and source records attributed to G Zambetti.

At least 19 recordsLinked to original sources

PubMed2005

Assessment of Cottle's areas through the application of a mathematical model deriving from acoustic rhinometry and rhinomanometric data.

OBJECTIVES: Each nasal area, as defined by Cottle, has a different influence on the nasal airflow. The longitudinal distribution of resistances in nasal cavities was calculated by the anterior rhinomanometry and acoustic rhinometry data. DESIGN: Dynamic study of Cottle's areas in normal subjects was carried out by rhinomanometry and acoustic rhinometry. SETTING: Study by the Department of Otolaryngology of the University of Rome-La Sapienza. PARTICIPANTS: Twenty-seven Caucasian adults in local and general healthy conditions took part and completed this study, with a total of 54 nasal cavities included because of negativity at ENT-examination and clinical history, with normal respiratory parameters at the rhinomanometry and acoustic rhinometry. MAIN OUTCOME MEASURES: We determined nasal and acoustic resistances, nasal volumes and cross-sectional surface areas, as defined by Cottle, using nasal endoscopy. The longitudinal distribution of nasal resistances was obtained by integrating experimental surface areas using a novel mathematical model. The estimation of the longitudinal nasal resistance variations as a result of a theoretical reduction of the surface areas. RESULTS: The reduction of the 2-3-1 areas (in this order of importance) showed the greatest influence on the nasal resistances with coefficients of determinations greater than 0.98, this being quite different from that of the areas 4 and 5 for quite smaller area reduction percentages. CONCLUSIONS: The areas 2-3-1 control the overall nasal resistance so the surgical procedures on these areas greatly influence the dynamics of nasal airflow. The mathematical model developed here gives useful information to nasal functional surgery and may be applied to other schemes of nasal cavity.

Acoustics↗

PubMed2001

Study and application of a mathematical model for the provisional assessment of areas and nasal resistance, obtained using acoustic rhinometry and active anterior rhinomanometry.

Nasal resistance (NR) depends on the geometrical features and tortuosity of the nasal airway and on the air flow. Knowing the longitudinal distribution of cross-sectional areas (CSAs) in the nasal cavity (which can be obtained using acoustic rhinometry) and the laminar nasal resistance (obtainable by processing the rhinomanometric results), it is possible to calculate, utilizing a mathematical model elaborated on the basis of fluid dynamics, the differential nasal resistance (NRdiff) and the cumulative nasal resistance (NRcum), thus localizing the position at which the highest resistance is concentrated and the related longitudinal distribution. Using a mathematical model, we integrated the sigmoid curves DeltaP/Q of rhinomanometry with the cross-sectional areas obtained using acoustic rhinometry, thus obtaining the normal distribution of differential and cumulative nasal resistances. Afterwards, we empirically reduced the cross-sectional areas corresponding to the head, body, tail and the whole inferior turbinate, recalculating the differential and cumulative nasal resistance distribution curves. The results show that reduction of up to 50% of cross-sectional areas does not substantially affect the resistivity role of the nasal valve, while greater reductions move the highest resistivity point to an area at the junction of the body and the head of the inferior turbinate. The study of the differential nasal resistance trend curves as a function of the reduction of cross-sectional areas shows that the resistance variation of the body and the whole inferior turbinate prevail with reductions of up to 40%, while the variation of cross-sectional areas of the body bordering the inferior turbinate head is predominant with higher reductions. The cross-sectional areas of the nasal airway cavity with highest resistivity are mainly located in an anterior position, where the differential nasal resistances are higher, but there are substantial variations produced by reducing the cross-sectional area of the posterior nasal airway. A similar model can produce provisional values for the results obtainable with functional nasal surgery.

Acoustics↗

PubMed2000

Amplification of 9q34 in childhood adrenocortical tumors: a specific feature unrelated to ethnic origin or living conditions.

Adrenocortical tumors (ACT) in children under 15 years of age exhibit some clinical and biological features distinct from ACT in adults. Cell proliferation, hypertrophy and cell death in adrenal cortex during the last months of gestation and the immediate postnatal period seem to be critical for the origin of ACT in children. Studies with large numbers of patients with childhood ACT have indicated a median age at diagnosis of about 4 years. In our institution, the median age was 3 years and 5 months, while the median age for first signs and symptoms was 2 years and 5 months (N = 72). Using the comparative genomic hybridization technique, we have reported a high frequency of 9q34 amplification in adenomas and carcinomas. This finding has been confirmed more recently by investigators in England. The lower socioeconomic status, the distinctive ethnic groups and all the regional differences in Southern Brazil in relation to patients in England indicate that these differences are not important to determine 9q34 amplification. Candidate amplified genes mapped to this locus are currently being investigated and Southern blot results obtained so far have discarded amplification of the abl oncogene. Amplification of 9q34 has not been found to be related to tumor size, staging, or malignant histopathological features, nor does it seem to be responsible for the higher incidence of ACT observed in Southern Brazil, but could be related to an ACT from embryonic origin.

Adenoma↗

PubMed1999

Non-specific nasal provocation test with histamine. Analysis of the dose-response curve.

Non-Specific Nasal Hyper-reactivity (NSNH) is described as a clinical condition characterized by the presence of rhinitic symptoms that are a consequence of non-specific stimulations. Because of its effects on vascular, epithelial, and glandular receptors, NSNP Test (NSNPT) with histamine allows the study of NSNH. The aims of this study are 1. to analyze the behavior of NSNH both in non-allergic chronic vasomotory patients and in healthy control subjects 2. to correlate total nasal resistance(TNR) to each dosage of histamine to derive the dose/response curves and 3. to study these curves to analyze and possibly define different stages according to the intensities of response of NSNH. We have studied 26 subjects affected by non-allergic vasomotor rhinitis and 10 healthy control subjects. We sprayed a NSNPT with histamine-phosphate (0.2-0.3-0.4-0.5-0.6-0.8 mg) in different sessions to avoid accumulation phenomena. Five minutes before and five minutes after each challenge, TNR was determined by active anterior rhinomanometry. TNR was correlated to the doses of histamine by an empirical equation. The most important results of this study are as follows: a) the variation of TNR follows a model of exponential curve, b) it is possible to classify NSNH, as a function of the regression b coefficient belonging to the empirical equation used, in reactivity classes, c) from one reactivity class to another, post-stimulation TNRs double; 0.5 mg of histamine of the NSNPT is the optimal dose, d) there is an overlap between the responses of some normal subjects and rhinopathic patients that will be the subject of a further study. Finally, our data suggest that, in a future perspective, it is possible to use the NSNPT with histamine for diagnostic, prognostic and therapeutic control purposes.

Adult↗

PubMed1998

Functional and physical interactions of the ARF tumor suppressor with p53 and Mdm2.

The INK4a-ARF locus encodes two proteins, p16(INK4a) and p19(ARF), that restrain cell growth by affecting the functions of the retinoblastoma protein and p53, respectively. Disruption of this locus by deletions or point mutations is a common event in human cancer, perhaps second only to the loss of p53. Using insect cells infected with baculovirus vectors and NIH 3T3 fibroblasts infected with ARF retrovirus, we determined that mouse p19(ARF) can interact directly with p53, as well as with the p53 regulator mdm2. ARF can bind p53-DNA complexes, and it depends upon functional p53 to transcriptionally induce mdm2 and the cyclin-dependent kinase inhibitor p21(Cip1), and to arrest cell proliferation. Binding of p19(ARF) to p53 requires the ARF N-terminal domain (amino acids 1-62) that is necessary and sufficient to induce cell cycle arrest. Overexpression of p19(ARF) in wild type or ARF-null mouse embryo fibroblasts increases the half-life of p53 from 15 to approximately 75 min, correlating with an increased p53-dependent transcriptional response and growth arrest. Surprisingly, when overexpressed at supra-physiologic levels after introduction into ARF-null NIH 3T3 cells or mouse embryo fibroblasts, the p53 protein is handicapped in inducing this checkpoint response. In this setting, reintroduction of p19(ARF) restores p53's ability to induce p21(Cip1) and mdm2, implying that, in addition to stabilizing p53, ARF modulates p53-dependent function through an additional mechanism.

3T3 Cells↗

PubMed1997

E2F-1 cooperates with topoisomerase II inhibition and DNA damage to selectively augment p53-independent apoptosis.

Mutations in the retinoblastoma (pRb) tumor suppressor pathway including its cyclin-cdk regulatory kinases, or cdk inhibitors, are a hallmark of most cancers and allow unrestrained E2F-1 transcription factor activity, which leads to unregulated G1-to-S-phase cell cycle progression. Moderate levels of E2F-1 overexpression are tolerated in interleukin 3 (IL-3)-dependent 32D.3 myeloid progenitor cells, yet this induces apoptosis when these cells are deprived of IL-3. However, when E2F activity is augmented by coexpression of its heterodimeric partner, DP-1, the effects of survival factors are abrogated. To determine whether enforced E2F-1 expression selectively sensitizes cells to cytotoxic agents, we examined the effects of chemotherapeutic agents and radiation used in cancer therapy. E2F-1 overexpression in the myeloid cells preferentially sensitized cells to apoptosis when they were treated with the topoisomerase II inhibitor etoposide. Although E2F-1 alone induces moderate levels of p53 and treatment with drugs markedly increased p53, the deleterious effects of etoposide in E2F-1-overexpressing cells were independent of p53 accumulation. Coexpression of Bcl-2 and E2F-1 in 32D.3 cells protected them from etoposide-mediated apoptosis. However, Bcl-2 also prevented apoptosis of these cells upon exposure to 5-fluorouracil and doxorubicin, which were also cytotoxic for control cells. Pretreating E2F-1-expressing cells with ICRF-193, a second topoisomerase II inhibitor that does not damage DNA, protected the cells from etoposide-induced apoptosis. However, ICRF-193 cooperated with DNA-damaging agents to induce apoptosis. Therefore, topoisomerase II inhibition and DNA damage can cooperate to selectively induce p53-independent apoptosis in cells that have unregulated E2F-1 activity resulting from mutations in the pRb pathway.

Antineoplastic Agents↗

PubMed1996

Cytoplasmic sequestration of wild-type p53 protein impairs the G1 checkpoint after DNA damage.

Wild-type p53 protein is abnormally sequestered in the cytoplasm of a subset of primary human tumors including neuroblastomas (NB) (U. M. Moll, M. LaQuaglia, J. Benard, and G. Riou, Proc. Natl. Acad. Sci. USA 92:4407-4411, 1995; U. M. Moll, G. Riou, and A. J. Levine, Proc. Natl. Acad. Sci.USA 89:7262-7266, 1992). This may represent a nonmutational mechanism for abrogating p53 tumor suppressor function. To test this hypothesis, we established the first available in vitro model that accurately reflects the wild-type p53 sequestration found in NB tumors. We characterized a series of human NB cell lines that overexpress wild-type p53 and show that p53 is preferentially localized to discrete cytoplasmic structures, with no detectable nuclear p53. These cell lines, when challenged with a variety of DNA strand-breaking agents, all exhibit impaired p53-mediated G1 arrest. Induction analysis of p53 and p53-responsive genes show that this impairment is due to suppression of nuclear p53 accumulation. Thus, this naturally occurring translocation defect compromises the suppressor function of p53 and likely plays a role in the tumorigenesis of these tumors previously thought to be unaffected by p53 alterations.

Cytoplasm↗

PubMed1995

E2F-1:DP-1 induces p53 and overrides survival factors to trigger apoptosis.

The E2F DNA binding activity consists of a heterodimer between E2F and DP family proteins, and these interactions are required for association of E2F proteins with pRb and the pRb-related proteins p107 and p130, which modulate E2F transcriptional activities. E2F-1 expression is sufficient to release fibroblasts from G0 and induce entry into S phase, yet it also initiates apoptosis. To investigate the mechanisms of E2F-induced apoptosis, we utilized interleukin-3 (IL-3)-dependent 32D.3 myeloid cells, a model of hematopoietic progenitor programmed cell death. In the absence of IL-3, E2F-1 alone was sufficient to induce apoptosis, and p53 levels were diminished. DP-1 alone was not sufficient to induce cell cycle progression or alter rates of death following IL-3 withdrawal. However, overexpression of both E2F-1 and DP-1 led to the rapid death of cells even in the presence of survival factors. In the presence of IL-3, levels of endogenous wild-type p53 increased in response to E2F-1, and coexpression of DP-1 further augmented p53 levels. These results provide evidence that E2F is a functional link between the tumor suppressors p53 and pRb. However, induction of p53 alone was not sufficient to trigger apoptosis, suggesting that the ability of E2F to override survival factors involves additional effectors.

Animals↗

PubMed1995

Intranasal immunotherapy with Dermatophagoides extract: in vivo and in vitro results of a double-blind placebo-controlled trial.

Intranasal immunotherapy (IT) has been proposed as a means to induce an effective immunity of the nasal mucosa in patients with allergic rhinitis, avoiding systemic side effects. In the present study 20 individuals with chronic allergic rhinitis, and skin prick test reactive to Dermatophagoides pteronyssinus (DP) only, were randomized and subjected to a three months' double-blind placebo-controlled trial of intranasal IT with DP extract. All patients received also sodium cromoglycate as pre-medication. Before and at the end of the treatment the patients performed specific nasal provocation tests, and samples of serum and nasal secretions were collected to measure total and specific IgE, levels of eosinophil cationic protein (ECP), and mast-cell-derived tryptase. A clinical score was computed by the symptoms indicated by the patients. The clinical score did not change in the two groups after the treatment, whereas a decrease in nasal reactivity was observed. Total IgE increased only in secretions from placebo-treated patients, but were not modified in sera. IgE to DP in sera and nasal secretions did not change significantly. Tryptase levels in nasal secretions decreased in both groups, while ECP was unchanged after IT. Serum ECP levels decreased more in actively treated patients than in the placebo group. The data suggest that changes of IgE and inflammatory mediators may be affected by the use of sodium cromoglycate in both groups, but some parameters change early in different directions in IT- and placebo-treated groups.

Administration, Intranasal↗

PubMed1993

Position and orientation-selective silencer in protein-coding sequences of the rat osteocalcin gene.

Osteocalcin (OC) is a bone-specific protein which is expressed postproliferatively by osteoblasts during late stages of differentiation. We have found that a silencer element is present within the rat OC gene (between nt +39 and +104), overlapping the OC signal prepropeptide-coding sequence. The presence of this sequence in OC promoter-CAT reporter constructs suppresses promoter activity in transiently transfected proliferating osteoblasts, which do not express OC, by up to 50-fold. This is the first demonstration of contribution from protein-coding sequences to silencing of animal genes. The element appears to be bipartite; silencer activity requires both the protein-coding sequence +39 to +63 and the +93 to +104 exon 1/intron 1 border region. Both of these domains contain sequences highly similar to silencer motifs in several other genes, including chicken lysozyme as well as rat collagen type II, insulin, and growth hormone. OC silencer activity is fully retained when the element is placed outside the RNA-coding region, 3' but not 5' of the OC-CAT fusion gene. Repression activity is orientation independent in the native position but requires the native orientation when located in 3' extragenic positions. The silencer does not inhibit the activity of the heterologous SV40 early promoter. These results suggest interaction between the transcribed silencer and specific OC promoter element(s) residing farther upstream. The OC transcribed silencer may contribute to developmental control of OC expression.

Animals↗

PubMed1993

Gain of function mutations in p53.

We report that the expression of murine or human mutant p53 proteins in cells with no endogenous p53 proteins confers new or additional phenotypes upon these cells. Mutant p53 proteins expressed in cell lines lacking p53 resulted in either enhanced tumorigenic potential in nude mice ((10)3 cells) or enhanced plating efficiency in agar cell culture (human SAOS-2 cells). Also, mutant human p53 alleles, unlike the wild-type p53 protein, could also enhance the expression of a test gene regulated by the multi-drug resistance enhancer-promoter element. These data demonstrate a gain of function associated with p53 mutations in addition to the loss of function shown previously to be associated with mutations in this tumour suppressor gene.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1991

Disruption of the cytoskeleton with cytochalasin D induces c-fos gene expression.

The treatment of exponentially growing HeLa cells and quiescent WI-38 cells with cytochalasin D, which disrupts the cytoskeletal microfilaments, results in a rapid and marked increase in the transcription of the c-fos protooncogene with a concomitant increase in c-fos mRNA steady-state levels. Transcription of rRNA, HLA-B7, and H4 histone genes was not significantly affected by the drug treatment. These results suggest that the nucleus can respond to signals related to the structural organization of the cytoskeleton and selectively adjust the regulation of gene expression.

Cytochalasin D↗

PubMed1990

Multiple types of mRNA-cytoskeleton interactions.

Nearly all actively translated mRNAs are associated with the cytoskeleton in HeLa cells and the nature of this association is poorly understood. To gain insight into this association, we have examined and compared the cytoskeleton-mRNA interactions of a signal peptide-histone fusion mRNA (membrane-bound polysomal mRNA) to those of endogenous histone mRNA (nonmembrane-bound polysomal mRNA). We report here the detection of a cytoskeleton attachment site within the signal peptide-histone fusion mRNP/mRNA nucleotide sequence that is not present in wild-type histone mRNA or in HLA-B7 and chorionic gonadotropin-alpha membrane-bound polysomal mRNAs. These results support the possibility that there are multiple mechanisms for the attachment of specific classes of mRNAs to the cytoskeleton.

Base Sequence↗

PubMed1990

Role of messenger RNA subcellular localization in the posttranscriptional regulation of human histone gene expression.

Histone mRNAs are naturally localized on non-membrane-bound polysomes and selectively destabilized during inhibition of DNA replication. Targeting histone mRNA to membrane-bound polysomes, by incorporating sequences coding for a signal peptide into the message, results in the stabilization of the histone fusion mRNA when DNA synthesis is interrupted (Zambetti et al.: Proceedings of the National Academy of Sciences of the United States of America 84:2683-2687, 1987). A single nucleotide substitution that abolishes the synthesis of the signal peptide results in the localization of the histone fusion mRNA on non-membrane-bound polysomes to the same extent as endogenous histone mRNA and fully restores the coupling of histone fusion mRNA stability to DNA replication. Signal peptide-histone fusion mRNAs containing two point mutations that result in the incorporation of two positively charged amino acids into the hydrophobic domain of the signal peptide are partially retained on non-membrane-bound polysomes and are partially destabilized during inhibition of DNA synthesis. These data indicate that the degree to which the signal peptide-histone fusion mRNAs are associated with non-membrane-bound polysomes is correlated with the extent to which the mRNAs are degraded during inhibition of DNA synthesis. These results suggest that the subcellular location of histone mRNA plays an important role in the posttranscriptional regulation of histone gene expression.

Amino Acid Sequence↗

PubMed1990

Differential association of membrane-bound and non-membrane-bound polysomes with the cytoskeleton.

We report here a differential release of specific mRNAs from the cytoskeleton by cytochalasin D treatment. Non-membrane-bound polysomal mRNAs, such as histone mRNA and c-fos mRNA, are readily released from the cytoskeleton of HeLa cells during cytochalasin D treatment. Over 90% of H3 and H4 histone mRNA is associated with the cytoskeleton in control cells and only 25% in cells treated with cytochalasin D (40 micrograms/ml). In contrast, the membrane-bound polysomal mRNAs for HLA-B7 and chorionic gonadotropin-alpha are inefficiently released from the cytoskeletal framework by cytochalasin D alone; approximately 98% of the HLA-B7 mRNA in control cells is associated with the cytoskeleton, whereas approximately 65% of the HLA-B7 mRNA is retained on the cytoskeleton in cells treated with cytochalasin D (40 micrograms/ml). Disruption of polysome structure with puromycin during cytochalasin D treatment results in the efficient release of HLA-B7 mRNA from the cytoskeleton. Under these conditions, only 25% of the HLA-B7 mRNA remains associated with the cytoskeletal framework. Thus, membrane-bound polysomes appear to be attached to the cytoskeleton through a cytochalasin D-sensitive site as well as through association with the nascent polypeptide and/or ribosome. These results demonstrate a complex association of polysomes with the cytoskeleton and elements of the endoplasmic reticulum.

Actin Cytoskeleton↗

PubMed1989

Structure of the rat osteocalcin gene and regulation of vitamin D-dependent expression.

The osteocalcin gene encodes a 6-kDa polypeptide, which represents one of the most abundant noncollagenous bone proteins, and the present studies establish that osteocalcin mRNA is detected only in bone tissue. An osteocalcin gene was isolated from a rat genomic DNA library, and sequence analysis indicated that the mRNA is represented in a 953-nucleotide segment of DNA consisting of four exons and three introns. A modular organization of the 5' flanking sequences of the gene is reflected by the presence of at least three classes of regulatory elements, which include the following: (i) RNA polymerase II canonical sequences; (ii) a series of consensus sequences for hormone receptor binding sites and cyclic nucleotide responsive elements consistent with physiologic expression of the osteocalcin gene; and (iii) a 24-nucleotide sequence in the proximal promoter region with a CAAT motif as a central element. We have designated this highly conserved sequence as an "osteocalcin box" since only 2 nucleotide substitutions are found in the rat and human osteocalcin genes. We have demonstrated two factors regulating osteocalcin gene expression. First, a 200-fold increase occurs in normal fetal calvaria osteoblasts producing a mineralizing matrix, compared to confluent osteoblasts in a nonmineralizing matrix. Second, contained within the 600 nucleotides immediately upstream from the transcription start site are sequences that support a 10-fold stimulated transcription of the gene by 1,25-dihydroxyvitamin D.

Amino Acid Sequence↗

PubMed1987

Histone gene expression remains coupled to DNA synthesis during in vitro cellular senescence.

Despite a decrease in the extent to which confluent monolayers of late compared to early passage CF3 human diploid fibroblasts can be stimulated to proliferate, the time course of DNA synthesis onset is similar regardless of the in vitro age of the cells. A parallel and stoichiometric relationship is maintained between the rate of DNA synthesis and the cellular levels of histone mRNA independent of the age of the cell cultures. Furthermore, DNA synthesis and cellular histone mRNA levels decline in a coordinate manner after inhibition of DNA replication by hydroxyurea treatment. These results indicate that while the proliferative activity of human diploid fibroblasts decreases with passage in culture, those cells that retain the ability to proliferate continue to exhibit a tight coupling of DNA replication and histone gene expression.

Cell Survival↗

PubMed1987

Targeting of a chimeric human histone fusion mRNA to membrane-bound polysomes in HeLa cells.

The subcellular location of histone mRNA-containing polysomes may play a key role in the posttranscriptional events that mediate histone mRNA turnover following inhibition of DNA synthesis. Previously, it has been shown that histone mRNA is found primarily on free polysomes that are associated with the cytoskeleton. We report here the construction of an Escherichia coli pBR322 beta-lactamase signal peptide-human H3 histone fusion gene. The fusion transcript is targeted to membrane-bound polysomes and remains stable following interruption of DNA replication. Relocating mRNA within the cell may provide a procedure for studying the posttranscriptional regulation of gene expression.

Cell Membrane↗