Biomedical subjects

Bruno Dallapiccola

Publications and source records attributed to Bruno Dallapiccola.

131 records · Page 8Linked to original sources

PubMed2002

Polymorphic changes in the 5' flanking region of factor VII have a combined effect on promoter strength.

Polymorphic differences in the 5' flanking region of the gene encoding procoagulant protein Factor VII (FVII) are associated with variations in FVII coagulant activity (FVII:C) and FVII antigen (FVII:Ag) levels. A decanucleotide insert polymorphism (CCTATATCCT) at 323 bp upstream of the start site of translation correlates with a decrease of approximately 20% FVII: C levels per allele containing this insert. However, linkage disequilibrium of the decanucleotide polymorphism with two single nucleotide polymorphisms (SNPs) at -122 and -401 have made it difficult to pinpoint the functional role, if any, of these genetic changes in lowering FVII levels. In vitro reporter gene studies in HepG2 cells analyzing the 8 possible combinations of polymorphic sites at -401, -323, and -122 reveal the necessity of the presence of the three concurrent polymorphic changes to maximally decrease promoter strength. In addition, these in vitro results are supported by in vivo studies in 89 individuals of African heritage, 34% of whom display a new haplotype that shows the polymorphic changes at -323 and -401 but lacks the change at -122.

5' Flanking Region↗

PubMed2002

Analysis of clinically relevant single-nucleotide polymorphisms by use of microelectronic array technology.

BACKGROUND: Microelectronic DNA chip devices represent an emerging technology for genotyping. We developed methods for detection of single-nucleotide polymorphisms (SNPs) in clinically relevant genes. METHODS: Primer pairs, with one containing a 5'-biotin group, were used to PCR-amplify the region encompassing the SNP to be interrogated. After denaturation, the biotinylated strand was electronically targeted to discrete sites on streptavidin-coated gel pads surfaces by use of a Nanogen Molecular Workstation. Allele-specific dye-labeled oligonucleotide reporters were used for detection of wild-type and variant sequences. Methods were developed for SNPs in genes, including factor VII, beta-globin, and the RET protooncogene. We genotyped 331 samples for five DNA variations in the factor VII gene, >600 samples from patients with beta-thalassemia, and 15 samples for mutations within the RET protooncogene. All samples were previously typed by various methods, including DNA sequence analysis, allele-specific PCR, and/or restriction enzyme digestion of PCR products. RESULTS: Analysis of amplified DNA required 4-6 h. After mismatched DNA was removed, signal-to-noise ratios were >5. More than 940 samples were typed with the microelectronic array platform, and results were totally concordant with results obtained previously by other genotyping methods. CONCLUSIONS: The described protocols detect SNPs of clinical interest with results comparable to those of other genotyping methods.

Factor VII↗

PubMed2002

A peptidase gene in chromosome 8q is disrupted by a balanced translocation in a duane syndrome patient.

PURPOSE: To identify the gene disrupted by a de novo reciprocal balanced translocation t(6;8)(q26;q13) in a patient with Duane retraction syndrome (DURS). The break point in chromosome arm 8q is positioned within the DURS1 critical region. METHODS: Fluorescence in situ hybridization (FISH) analysis using cosmid and BAC clones covering the DURS1 locus was performed to define the break point position and its relationship with expressed sequence tags (ESTs) in the region. Once the interrupted gene was identified, the full-length cDNA was sequenced and the genomic organization defined. Eighteen patients with sporadic DURS without cytogenetic abnormalities involving the DURS1 region were screened for point mutations in the candidate DURS1 gene. RESULTS: A carboxypeptidase gene (CPAH) was directly interrupted between the first and second exons in a patient with DURS who carried a de novo reciprocal balanced translocation t(6;8)(q26;q13) involving the DURS1 region on chromosome arm 8q13. The gene was transcribed in at least two alternative mRNA forms, with different start and stop codons. CONCLUSIONS: The CPAH gene was interrupted in a patient with DURS carrying a translocation break point in the DURS1 region on chromosome 8q13. CPAH is therefore a likely candidate for this abnormality, even if the possibility that other genes are involved, either by direct effects on transcription units present in the first CPAH intron or by position effects, cannot be ruled out. Functional studies of the influence of this gene on the morphogenesis of eye muscles and their innervation may clarify this question.

Adult↗

PubMed2001

Single primer pair for PCR identification of Candida parapsilosis group I isolates.

A DNA fragment, isolated from a genomic DNA mini-library of Candida parapsilosis group I reference strain ATCC 22019, was sequenced and characterised. The fragment was first probed by Southern blotting against a pool of DNA from several yeasts. The hybridisation tests revealed that the probe was specific for strain ATCC 22019 and 49 (90.74%) of 54 C. parapsilosis clinical and soil isolates that were attributed to C. parapsilosis group I by the electrophoretic images of their restriction fragment length polymorphisms (RFLPs) and electrophoretic karyotype (EK). The remaining five negative isolates, according to the same criteria, were attributed to group II (one isolate) and III (four isolates). Two primers were selected from the probe DNA sequence and a PCR-based test was developed for specifically detecting C. parapsilosis group I isolates, which represent the majority of the common clinical isolates. The PCR assay confirmed the Southern hybridisation results. This PCR assay could be a simple and reproducible tool for the rapid, sensitive and species-specific identification of C. parapsilosis major group I isolates.

Base Sequence↗

PubMedDate not supplied

Low doses of dexamethasone constantly delivered by autologous erythrocytes slow the progression of lung disease in cystic fibrosis patients.

OBJECTIVE: To evaluate the safety and efficacy of the administration of low doses of glucocorticoids in patients with cystic fibrosis (CF) by using autologous erythrocytes loaded with dexamethasone 21-phosphate. STUDY DESIGN: Nine consecutive CF patients (patients nos. 1-9) received autologous erythrocytes loaded with increasing amounts of dexamethasone 21-phosphate to obtain a slow delivery of dexamethasone in circulation. The appearance of possible adverse effects, the reproducibility of the procedure, and the dexamethasone pharmacokinetics were evaluated. Subsequently, patient no. 9 and eight additional patients (patient nos. 10-17) received dexamethasone 21-phosphate-loaded erythrocytes at 1-month intervals to evaluate the efficacy of continuous release in circulation of low doses of dexamethasone. RESULTS: Erythrocytes from CF patients can be processed to be loaded with increasing dexamethasone 21-P concentrations. Once reinfused in respective donors, a slow and prolonged delivery of dexamethasone in the blood stream was measured up to 28 days. Repeated administrations of drug-loaded erythrocytes at 4-week intervals for 15 months showed that very low doses of glucocorticoids provide significant improvement in FEV1 values and significant reduction of infective relapses due to Pseudomonas aeruginosa without adverse effects. CONCLUSIONS: The administration of very low doses of glucocorticoids using autologous erythrocytes is possible, with benefits for patients and without side effects. This method is likely to be extended to other chronic diseases.

Adolescent↗