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Biomedical subjects

Mario Cazzola

Publications and source records attributed to Mario Cazzola.

At least 91 records · Page 5Linked to original sources
PubMed2003

Screening hepcidin for mutations in juvenile hemochromatosis: identification of a new mutation (C70R).

Juvenile or type 2 hemochromatosis (JH) is a genetic disease caused by increased intestinal iron absorption that leads to early massive iron overload. The main form of the disease is caused by mutations in a still unknown gene on chromosome 1q. Recently, we recognized a second type of JH with clinical features identical to the 1q-linked form, caused by mutations in the gene encoding hepcidin (HEPC). Hepcidin is a hepatic antimicrobial-like peptide whose role in iron homeostasis was first defined in animal models; deficiency of hepcidin in mice leads to iron overload, whereas its hepatic overexpression in transgenic animals causes iron deficiency. To define the prevalence of HEPC mutations in JH we screened the HEPC gene for mutation in 21 unrelated JH subjects. We identified a new mutation (C70R), which affects 1 of the 8 conserved cysteines that form the disulfide bonds and are critical for the stability of the polypeptide.

Amino Acid Sequence↗
PubMed2003

Denaturing HPLC analysis of DNA deletions and insertions.

Denaturing HPLC (DHPLC) is a useful technique for the fast screening of known and unknown heterozygous gene mutations. Most DNA mutations causing genetic disorders consist of nucleotide substitutions, but insertions and deletions occur, albeit less frequently. The heteroduplexes with insertions/deletions have gaps that may affect molecular stability differently from the mismatches caused by substitutions. Therefore, gaps and mismatches may be distinguished by DHPLC analysis, which is based on the differential thermal stability of amplicons with different characteristics. To verify this hypothesis, we examined 12 DNA samples containing insertions and deletions of different sizes (one to 29 residues) from four different genes (ABCA4, CFTR, FTL, and SLC11A3). We found that all of them were detected by DHPLC runs at 50 degrees C, which is considered a non-denaturing temperature, as well as by runs at the temperature optimized for mismatch recognition. The finding confirms that gaps reduce heteroduplex stability more than mismatches, and indicates that DHPLC analysis at low temperature may be applied to distinguish DNA deletions/insertions from substitutions.

5' Untranslated Regions↗
PubMed2003

Novel approaches to the treatment of pneumonia.

As the prevalence of resistance to multiple antibiotics increases it is progressively more difficult to treat pneumonia in hospitalized patients. Therefore, anti-infectious agents that have new modes of action are needed urgently. Recent advances in DNA sequencing technology make it possible to elucidate the sequences of the entire genomes of pathogenic bacteria. This allows many novel, non-traditional targets for therapeutic intervention to be identified, such as those involved in disease pathogenesis, and in adaptation and growth at sites of infection. In the past few years, inhibitors of new bacterial targets have been developed, including inhibitors of genes that are required for either virulence or pathogenesis. The challenge is to optimize and develop these agents to provide novel approaches to the treatment of pneumonia in hospitalized patients.

Adjuvants, Immunologic↗
PubMed2003

Bronchodilator response to formoterol Turbuhaler in patients with COPD under regular treatment with formoterol Turbuhaler.

Formoterol Turbuhaler has been suggested for as-needed use in asthmatic patients. We investigated whether regular treatment with formoterol would modify the dose-response curves to formoterol in patients with partially reversible COPD. In this randomised, double-blind, cross-over study taking place over four non-consecutive days 16 outpatients with moderate to severe COPD, who were under regular treatment with formoterol Turbuhaler (18 microg in two daily doses) from at least 4 months, inhaled a conventional dose of formoterol Turbuhaler 9 microg or placebo. Two hours later, a FEV(1) value was established, following which a dose-response curve to formoterol (4.5 microg/inhalation) or placebo was constructed using four inhalations (1+1+2)--total cumulative delivered dose of 18 microg formoterol--with the following sequences: (1) formoterol pre-treatment + formoterol 18 microg, (2) formoterol pre-treatment + placebo, (3) placebo pre-treatment + formoterol 18 microg, (4) placebo pre-treatment + placebo. Formoterol 9 microg induced significant (P < 0.0001) bronchodilation at 2 h after inhalation (best mean increase in FEV(1): 0.170 L). Afterwards, dose-dependent increases in FEV(1) occurred with formoterol (maximum mean increase from 2-h value with formoterol: 0.072 after formoterol pre-treatment, and 0.201 L after placebo pre-treatment). Both maximum values of bronchodilation after the last inhalation of formoterol were statistically different (P < 0.001) from 2-h levels. These results show that dose-dependent bronchodilatation of formoterol is maintained despite regular treatment.

Aged↗
PubMed2003

Novelties in the field of antimicrobial compounds for the treatment of lower respiratory tract infections.

Emergence of antimicrobial resistance is a growing problem and a public health threat. New drugs must be designed with emerging needs in mind: specific resistant and hard-to-treat organisms. But the difficulty to find real new drugs is a major problem. Only the oxazolidinones, the cationic peptides and the lipopeptide antibiotics can be truly regarded as structurally novel drugs, although the peptide deformylase inhibitors and, possibly, the pleuromutilins can be considered a potential advancement in the field. Obviously, these antibiotics must be reserved only to cases of documented ineffectiveness of the common antimicrobial agents.

Acetamides↗
PubMed2003

Scanning mutations of the 5'UTR regulatory sequence of L-ferritin by denaturing high-performance liquid chromatography: identification of new mutations.

Hereditary hyperferritinaemia cataract syndrome is an autosomal dominant disorder caused by heterogeneous mutations of the iron regulatory element (IRE) in the ferritin l-chain mRNA. The mutations are rare and fast DNA scanning would facilitate diagnosis. The aim of the study was to compare the analytical performances of two fast DNA scanning techniques: denaturing high-performance liquid chromatography (DHPLC) and double-gradient denaturing gradient gel electrophoresis (DG-DGGE). We analysed the sequence encoding the 5' untranslated flanking region of ferritin l-chain mRNA, which includes an IRE stem loop structure. The two systems unambiguously identified all the 12 accessible mutations in a single run, including the difficult C-G transversions. DHPLC and DG-DGGE identified seven abnormal patterns in DNA samples from 47 subjects with unexplained hyperferritinaemia; all had mutations in the IRE sequence, including two not reported before: C36G and A37G. The scanning of 250 DNA samples from subjects genotyped for HFE led to the identification of four new mutations, all outside the IRE structure: C10T, C16T, C90T and del-T156. We conclude that DHPLC, similar to DG-DGGE, detects all the mutations in the l-ferritin 5'UTR sequence in a single run, and that various mutations occur outside the IRE structure.

5' Untranslated Regions↗
PubMed2003

Once-weekly epoetin beta is highly effective in treating anaemic patients with lymphoproliferative malignancy and defective endogenous erythropoietin production.

Epoetin beta, three-times weekly (t.i.w.), is effective in reversing anaemia in lymphoproliferative disorders. The current study investigated whether an epoetin beta dose of 30,000 IU given subcutaneously once weekly (q.w.) was at least as effective as 10,000 t.i.w. administration in anaemic patients with lymphoproliferative malignancy and defective endogenous erythropoietin (Epo) production. Overall, 241 anaemic patients with multiple myeloma, low-grade non-Hodgkin's lymphoma or chronic lymphocytic leukaemia, all with serum Epo values </= 100 mU/ml, were randomized to receive the q.w. (n = 119) or t.i.w. (n = 122) regimen for 16 weeks. The primary efficacy criterion, i.e. the time-adjusted area under the haemoglobin-time curve from weeks 5-16, was comparable between the q.w. and t.i.w. groups [difference = - 0.20 g/dl (90% confidence interval - 0.52-0.11)]. Moreover, response rates were high and similar in both arms (72%vs 75%, q.w. and t.i.w. groups respectively). Baseline serum Epo was predictive of response: the lower serum Epo, the higher the likelihood of response (P = 0.002). Thus, epoetin beta administered q.w. is an effective and convenient treatment for anaemia in patients with lymphoproliferative disorders. Tailoring this treatment modality to subjects with defective endogenous Epo production represents a rational use of epoetin from both a medical and a community perspective.

Adult↗
PubMed2003

Therapeutic potential of the new quinolones in the treatment of lower respiratory tract infections.

The impact of respiratory infections on public health is increasing and lower respiratory tract infections are a major cause of morbidity and mortality. We are also facing a worldwide burst of antibiotic bacterial resistance. The new fluoroquinolones have an excellent spectrum covering the most important respiratory pathogens, including atypical and 'typical' pathogens. Pharmacokinetic and dynamic properties of the new fluoroquinolones have a significant impact on their clinical and bacteriological efficacy. They cause a concentration-dependent killing with a sustained postantibiotic effect. Fluoroquinolones combine exceptional efficacy with cost-effectiveness. Not surprisingly, different guidelines have inserted these agents among the drugs of choice in the empirical therapy of community-acquired pneumonia. This review discusses the more recent data on bacteriological and clinical activity and critically analyses the risks of a potential overuse of this valuable new class of drugs.

4-Quinolones↗
PubMed2003

Hematologic passport for athletes competing in endurance sports: a feasibility study.

BACKGROUND AND OBJECTIVES: Strategies based on the use of upper thresholds of hemoglobin or hematocrit to detect blood doping in endurance sports have essentially failed to deter this malpractice. With the aim of establishing a more effective strategy, we analyzed the biological variations of hematologic parameters in professional athletes and investigated the possibility of defining subject-specific reference ranges that could distinguish between physiologic and abnormal variability. DESIGN AND METHODS: Hemoglobin concentration, hematocrit, reticulocyte count, serum ferritin and soluble transferrin receptor levels were sequentially evaluated in 923 professional football players. Using the analysis of variance we tested the effect of age, ethnicity, exercise modalities and training phases on hematologic parameters and then estimated components of variation. The significance of the difference between two measures was obtained from the distribution of the within-subject variance (the so-called reference change). Subject-specific reference ranges were centered around the individual mean value with dispersion based on the 95th percentile of the coefficient of variation distribution. RESULTS: A total of 2,506 hematologic determinations were made. Exercise modalities were found to have important effects on hematologic parameters. Hemoglobin and hematocrit values were higher at the beginning of the competition season, and then declined in well-trained athletes. Aerobic exercise was clearly associated with lower values, suggesting that marginally low hemoglobin and hematocrit values should physiologically be found in endurance sports. At least five determinations were required to define subject-specific reference ranges reliably. Considering athletes showing normal indices of red cell production (i.e., reticulocyte count and soluble transferrin receptor), the 95th percentile of the coefficient of variation distribution was lower than 5% for both hemoglobin and hematocrit. Increases exceeding 10% in these latter parameters should to be considered abnormal. Score systems capable of efficiently detecting non-physiologic increases in red cell production were developed. INTERPRETATION AND CONCLUSIONS: Using proper sequential determinations of hematologic variables subject-specific reference ranges can be defined for hemoglobin and hematocrit. Thus, the hematologic passport is feasible and might be employed to exclude athletes with non-physiologic increases in hemoglobin and hematocrit from competitions. The hematologic passport should be used within a global strategy to deter blood doping.

Adult↗
PubMed2003

Clinical utility of the absolute number of circulating CD34-positive cells in patients with chronic myeloproliferative disorders.

BACKGROUND AND OBJECTIVES: Flow cytometry enumeration of peripheral blood CD34-positive cells provides reliable measurements of circulating hematopoietic progenitors in humans. Since the absolute number of circulating CD34-positive cells has been previously found to be elevated in myelofibrosis with myeloid metaplasia (MMM), we prospectively studied the clinical utility of this parameter in the work-up of patients with chronic myeloproliferative disorders. DESIGN AND METHODS: Of the 248 consecutive patients enrolled in this study, 106 had polycythemia vera (PV), 90 essential thrombocythemia (ET), and 52 myelofibrosis with myeloid metaplasia (MMM). The study population included both newly diagnosed and established cases, and of these latter some patients were on cytoreductive treatment while others were chemotherapy naive. Both cross-sectional and longitudinal investigations were carried out. Flow cytometry enumeration of CD34-positive cells was performed using a single-platform assay. RESULTS: Median numbers and ranges of circulating CD34-positive cells were 2.3x10(6)/L (0-5) in 20 control subjects, 2.2x10(6)/L (0-14) in those with PV, 2.4x10(6)/L (0-14) in those with ET, and 114x10(6)/L (6-2,520) in MMM patients. Analysis of variance demonstrated that values were markedly higher in MMM patients than in the remaining groups, and counts did not appear to fluctuate over short periods of follow-up. In both cross-sectional and longitudinal investigations on patients at clinical onset and/or out of cytoreductive treatment, a CD34-positive count of > or = 15x10(6)/L was always associated with MMM, clearly indicating a disease-related specificity. INTERPRETATION AND CONCLUSIONS: The absolute number of circulating CD34-positive cells is normal or anyhow lower than 15x10(6)/L in patients with uncomplicated PV or ET, whereas it is equal or above this cut-off in those with MMM, likely reflecting abnormal hematopoietic progenitor cell trafficking. Thus, enumeration of circulating CD34-positive cells may be useful in the work-up of patients with myeloproliferative disorders.

Adolescent↗
PubMed2002

Mitochondrial ferritin expression in erythroid cells from patients with sideroblastic anemia.

The sideroblastic anemias are characterized by ring sideroblasts, that is, red cell precursors with mitochondrial iron accumulation. We therefore studied the expression of mitochondrial ferritin (MtF) in these conditions. Erythroid cells from 13 patients with refractory anemia with ring sideroblasts (RARS) and 3 patients with X-linked sideroblastic anemia (XLSA) were analyzed for the distribution of cytoplasmic H ferritin (HF) and MtF using immunocytochemical methods. We also studied 11 healthy controls, 5 patients with refractory anemia without ring sideroblasts (RA), and 7 patients with RA with excess of blasts (RAEB). About one fourth of normal immature red cells, mostly proerythroblasts and basophilic erythroblasts, showed diffuse cytoplasmic positivity for HF, but very few were positive for MtF (0%-10%). Similar patterns were found in anemic patients without ring sideroblasts. In contrast, many erythroblasts from patients with sideroblastic anemia (82%-90% in XLSA and 36%-84% in RARS) were positive for MtF, which regularly appeared as granules ringing the nucleus. Double immunocytochemical staining confirmed the different cellular distribution of HF and MtF. There was a highly significant relationship between the percentage of MtF(+) erythroblasts and that of ring sideroblasts (Spearman R = 0.90; P <.0001). Reverse transcription-polymerase chain reaction studies demonstrated the presence of MtF mRNA in circulating reticulocytes of 2 patients with XLSA but not in controls. These findings suggest that most of the iron deposited in perinuclear mitochondria of ring sideroblasts is present in the form of MtF and that this latter might be a specific marker of sideroblastic anemia.

5-Aminolevulinate Synthetase↗
PubMed2002

Absent phenotypic expression of X-linked sideroblastic anemia in one of 2 brothers with a novel ALAS2 mutation.

X-linked sideroblastic anemia (XLSA) is caused by mutations in the erythroid-specific 5-aminolevulinic acid synthase (ALAS2) gene. Hemizygous males have microcytic anemia and iron overload. A 38-year-old male presented with this phenotype (hemoglobin [Hb] 7.6 g/dL, mean corpuscular volume [MCV] 64 fL, serum ferritin 859 microg/L), and molecular analysis of ALAS2 showed a mutation 1731G>A predicting an Arg560His amino acid change. A 36-year-old brother was hemizygous for this mutation and expressed the mutated ALAS2 mRNA in his reticulocytes, but showed almost no phenotypic expression. All 5 heterozygous females from this family, including the 3 daughters of the nonanemic hemizygous male, showed marginally increased red-cell distribution width (RDW). Although variable penetrance for XLSA in males has been previously described, this is the first report showing that phenotypic expression can be absent in hemizygous males. This observation is relevant to genetic counseling, emphasizing the importance of gene-based diagnosis.

5-Aminolevulinate Synthetase↗
PubMed2002

Delivering antibacterials to the lungs: considerations for optimizing outcomes.

An important determinant of clinical outcome of a lower respiratory tract infection may be sterilization of the infected lung, which is also dependent on sustained antibacterial concentrations achieved in the lung. For this reason, recently there has been increased interest in measuring the concentration of antimicrobial agents at different potential sites of infection in the lung. Levels of antibacterials are now measured in bronchial mucosa, epithelial lining fluid (ELF) and alveolar macrophages, as well as in sputum. Penicillins and cephalosporins reach only marginal concentrations in bronchial secretions, whereas fluoroquinolones and macrolides have been shown to achieve high concentrations. The extent of penetration of different antibacterials into the bronchial mucosa is relatively high. This is also true for beta-lactams, although their tissue concentrations never reach blood concentrations. Antibacterials penetrate less into the ELF than into the bronchial mucosa, but fluoroquinolones appear to concentrate more into alveolar lavage than into bronchial mucosa. Pulmonary pharmacokinetics is a very useful tool for describing how drugs behave in the human lung, but it does not promote an understanding of the pharmacological effects of a drug. More important, instead, is the correlation between pulmonary disposition of the drug and its minimum inhibitory concentration (MIC) values for the infectious agent. The addition of bacteriological characteristics to in vivo pharmacokinetic studies has triggered a 'pharmacodynamic approach'. Pharmacodynamic parameters integrate the microbiological activity and pharmacokinetics of an anti-infective drug by focusing on its biological effects, particularly growth inhibition and killing of pathogens. Drugs that penetrate well and remain for long periods at the pulmonary site of infection often induce therapeutic responses greater than expected on the basis of in vitro data. However, although the determination of antibacterial concentrations at the site of infection in the lung has been suggested to be important in predicting the therapeutic efficacy of antimicrobial treatment during bacterial infections of the lower respiratory tract, some studies have demonstrated that pulmonary bacterial clearance is correlated more closely to concentrations in the serum than to those in the lung homogenates, probably because they better reflect antibacterial concentration in the interstitial fluid.

Anti-Infective Agents↗
PubMed2002

Hereditary hyperferritinaemia/ cataract syndrome.

In mammalian cells, cellular iron homeostasis is maintained by the co-ordinated regulation of transferrin receptor and ferritin synthesis that occurs at the translational level. This regulation is mediated by iron-responsive elements (IREs) that are found within the untranslated regions (UTRs) of mRNA and by cytoplasmic mRNA-binding proteins, known as iron regulatory proteins (IRPs). When cellular iron is scarce, IRPs are available for binding the 5' IRE of ferritin mRNA, initiation of translation is prevented and ferritin synthesis is inhibited. By contrast, the presence of abundant intracellular iron prevents binding of the IRPs to the 5' IRE and allows efficient mRNA translation to proceed. Hereditary hyperferritinaemia/cataract syndrome (HHCS) arises as a result of various point mutations or deletions within a protein binding sequence in the 5'-UTR of the L-ferritin mRNA, which results in increased efficiency of L-ferritin translation. Each unique mutation confers a characteristic degree of hyperferritinaemia and severity of cataract in affected individuals. This exemplifies a new paradigm in which mutations in mRNA cis-acting elements may be responsible for phenotypic variability in disease states.

Cataract↗
PubMed2002

A novel deletion of the L-ferritin iron-responsive element responsible for severe hereditary hyperferritinaemia-cataract syndrome.

In the last few years, mutations that cause disease through increased efficiency of mRNA translation have been discovered. Hereditary hyperferritinaemia-cataract syndrome (HHCS) arises from various point mutations or deletions within the iron-responsive element (IRE) in the 5'-UTR of the L-ferritin mRNA. Each unique mutation confers a characteristic degree of hyperferritinaemia and severity of cataract in affected individuals. We report a novel six-nucleotide deletion identified in an Italian family presenting with elevated serum ferritin and early onset bilateral cataract. This deletion involves a sequence with a TCT repetition and may have occurred through a mechanism of slippage mispairing. Because of the above repetition, the observed mutation can be interpreted as deletion 22-27, 23-28, 24-29 or 25-30. Structural modelling predicted an IRE stem modification that is expected to markedly reduce the binding to iron-regulatory proteins. A double-gradient denaturing gradient gel electrophoresis (DG-DGGE) method easily detected the above deletion.

Adult↗