Biomedical subjects
J Silva
Publications and source records attributed to J Silva.
[Clinical results of double versus single mammary artery myocardiac revascularization: 15 years of follow-up].
INTRODUCTION: Use of the left internal mammary artery to bypass the left anterior descending coronary artery reduces cardiac events and increases survival. However, there is some controversy as to the benefits of using both mammary arteries. OBJECTIVES: To assess the long-term outcome of the use of both mammary arteries in comparison with the use of only one. PATIENTS AND METHOD: A retrospective cohort study with a mean follow-up of 9.0 +/- 4.2 years was performed including 108 patients consecutively revascularized using both mammary arteries (II) and 108 patients randomly chosen in whom one mammary artery (I) was used for this purpose. RESULTS: Both groups were similar. There were no differences between the groups in operative morbidity or mortality. The survival at 10 years was similar (II: 84.61 +/- 4%; I: 85.18 +/- 3.8%), whereas recurrence of angina (II: 29.63 +/- 5.3%; I: 47.55 +/- 5.6%) (p = 0.012), the requirement for percutaneous angioplasty (II: 3.98 +/- 2%; I: 12.99 +/- 4.1%) (p = 0.009) and cardiologic events (II: 33.48 +/- 5.5%; I: 48.48 +/- 5.5%)(p = 0.022) were all lower in the group in which both mammary arteries were used. In the multivariate analysis, the use of both mammary arteries was an independent protective factor against angina recurrence (RR = 0.55), angioplasty (RR = 0.18) and cardiologic event (RR = 0.60). CONCLUSIONS: The use of both mammary arteries for revascularization does not increase operative morbidity. Since this procedure acts as an independent factor against angina recurrence, angioplasty and cardiologic event
Isolation of Clostridium innocuum from cases of recurrent diarrhea in patients with prior Clostridium difficile associated diarrhea.
Clostridium innocuum isolates resistant to vancomycin (MIC values of 16-24 microg/mL) were isolated from three patients with recurrent Clostridium difficile -associated diarrhea (CDAD). We discuss the clinical significance and problems associated with the identification and differentiation of these two clostridial species, which may result in misdiagnosis of patients.
Wild type p73 overexpression and high-grade malignancy in breast cancer.
The overexpression of wild type p73 is the most frequent alteration of p73 in malignancies. We investigated, in 70 breast carcinomas, p73 mRNA expression and its relationship to p53 mutations, determined by an immunohistochemical method, and loss heterozygosity (LOH) status of the 1p36 region, together with its possible implication in the pathogenesis of breast carcinomas. LOH, amplifying DNA by PCR using 5 markers, of 1p36 region (one intragenic to p73 gene) was found in 17% of cases but no significant correlation was observed with p73 overexpression. p53 positive immunostaining was present in 33% of breast carcinomas, and these exhibited a statistically significant relation with p73 overexpressed tumors. Overexpression of p73 mRNA was observed in 19 tumors (27%). The analysis of cases with p73 overexpression and cases with normal mRNA expression, in terms of age and pathologic characteristics of the tumors showed a significant association of p73 overexpression and tumors with lymph node metastases, vascular invasion and higher pathologic stage. These results suggest that p73 overexpression is a molecular alteration that could be implicated in the tumorigenesis of breast carcinomas and, eventually, in a poor clinical behavior.
Molecular epidemiology of a multiresistant Pseudomonas aeruginosa outbreak in a paediatric intensive care unit.
After isolation of multiresistant (MR) Pseudomonas aeruginosa from 3 hospitalized patients in a paediatric intensive care unit (PICU), a prospective surveillance programme was established to detect infected and/or colonized patients in the hospital. Isolates were examined by means of outer membrane protein (OMP) profiles, serotyping and DNA genomic analysis using pulsed-field gel electrophoresis (PFGE). Fifty-five P. aeruginosa strains were isolated from 23 hospitalized patients during September and October 1997. The median hospital stay before isolation of P. aeruginosa was 8 d. PFGE demonstrated that the same clone infected 14 patients, 4 of whom were not hospitalized in the PICU. Susceptibility patterns and OMP profiles correlated with PFGE results in 37.8% and 36.4% of cases, respectively. Serotype O11 correlated with pattern A in 77% of cases and serotype O4 correlated with unrelated strains in 75% of cases but did not discriminate between outbreak and unrelated isolates. Extensive investigation of cultures failed to identify a reservoir of P. aeruginosa. PFGE was superior to OMP analysis and serotyping for discriminating between strains. The possible mode of acquisition for most of the patients infected with the same clone was cross-contamination.
In vitro activity of sitafloxacin against Clostridium difficile.
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Gastric intramucosal pH and intraluminal PCO2 during weaning from mechanical ventilation.
OBJECTIVE: To study the value of gastric intramucosal pH and gastric intraluminal PCO2 measurements to predict weaning outcome from mechanical ventilation. DESIGN: Prospective clinical study. SETTING: Intensive care medicine department of a university hospital. PATIENTS: Nineteen adult critically ill patients who were mechanically ventilated because of acute respiratory failure and were considered ready to be weaned. INTERVENTIONS: The patients were weaned with: synchronized intermittent mandatory ventilation plus positive end-expiratory pressure (SIMV+PEEP) or continuous positive airway pressure with pressure support ventilation (CPAP+PSV). A gastric tonometer was placed in all the patients. Tonometric, respiratory, and hemodynamic variables were measured during the weaning process. MEASUREMENTS: Hemodynamic variables, respiratory mechanics, pulmonary gas exchange, respiratory muscle force, spontaneous pattern of breathing, and the central control of breathing were recorded. Simultaneously, the intramucosal pH and gastric intraluminal PCO2 were measured. MAIN RESULTS: Eleven patients were successfully extubated and eight failed. The patients who failed showed higher values of mouth occlusion pressure, respiratory rate, and effective inspiratory impedance (mouth occlusion pressure/mean inspiratory flow). The intramucosal pH was initially 7.19 +/- 0.22 and decreased to 7.10 +/- 0.16 during the weaning process in patients who failed (p < .05). At the same time, the intramucosal pH showed a nonsignificant change from 7.36 +/- 0.07 to 7.32 +/- 0.07 in the patients who were successfully extubated. The intramucosal pH was statistically different when both groups were compared during the initial and the final evaluations (p < .05). For the initial evaluation, the sensitivity and specificity to predict weaning failure when the intramucosal pH was < or =7.30 were 0.88 (95% confidence interval [CI], 0.66-1) and 0.82 (95% CI, 0.59-1), respectively. The gastric intraluminal PCO2 was higher in patients who failed (p < .05). When gastric intraluminal PCO2 was . or =40 torr during the initial evaluation, weaning failure occurred with a sensitivity of 1 (95% CI, 0.31-1) and a specificity of 0.55 (95% CI, 0.26-0.84). CONCLUSIONS: Weaning failure was associated with gastric intramucosal acidosis. The intramucosal pH and gastric intraluminal PCO2 may be helpful to predict weaning outcome. Further controlled clinical trials in a larger group of patients are needed.
Resistance to moxifloxacin in toxigenic Clostridium difficile isolates is associated with mutations in gyrA.
Clostridium difficile is the etiological agent of antibiotic-associated colitis and the most common cause of hospital-acquired infectious diarrhea. Fluoroquinolones such as ciprofloxacin are associated with lower risks of C. difficile-associated diarrhea. In this study, we have analyzed 72 C. difficile isolates obtained from patients with different clinical courses of disease, such as toxic megacolon and relapses; the hospital environment; public places; and horses. They were investigated for their susceptibilities to moxifloxacin (MXF), metronidazole (MEO), and vancomycin (VAN). Mutants highly resistant to fluoroquinolones were selected in vitro by stepwise exposure to increasing concentrations of MXF. The resulting mutants were analyzed for the presence of mutations in the quinolone resistance-determining regions of DNA gyrase (gyrA), the production of toxins A and B, and the epidemiological relationship of these isolates. These factors were also investigated using PCR-based methods. All strains tested were susceptible to MEO and VAN. Twenty-six percent of the clinical isolates (19 of 72) were highly resistant to MXF (MIC > or = 16 microg/ml). Fourteen of these 19 strains contained nucleotide changes resulting in amino acid substitutions at position 83 in the gyrA protein. Resistant strains selected in vitro did not contain mutations at that position. These findings indicate that resistance to MXF in a majority of cases may be due to amino acid substitution in the gyrA gene.
Outbreak of infection with extended-spectrum beta-lactamase-producing Klebsiella pneumoniae in a Mexican hospital.
Thirty-one strains of Klebsiella pneumoniae (including 10 duplicates) from 21 septicemic pediatric patients (age, <2 months) were studied during a 4-month period (June to October 1996) in which the fatality rate was 62% (13 of 21). These isolates identified by the API 20E system yielded the same biotype. Pulsed-field gel electrophoresis experiments revealed the same clone in 31 strains. The isolates were multidrug-resistant but were still susceptible to ciprofloxacin, imipenem, and cefoxitin. A 135-kb plasmid was harbored in all of the isolates. No transconjugants were obtained that were resistant to ampicillin, cefotaxime, tetracycline, or gentamicin. Isoelectric focusing for beta-lactamases was performed on all strains, and three bands with pIs of 5.4, 7.6, and 8.2 were obtained. Of these, the pI 8.2 beta-lactamase had an extended-spectrum beta-lactamase phenotype. PCR amplification of both TEM- and SHV-type genes was obtained. The sequence analysis of the SHV PCR product indicated a mutation corresponding to the SHV-5 beta-lactamase.
Molecular typing methods for the epidemiological identification of Clostridium difficile strains.
Toxigenic Clostridium difficile is the etiologic agent of C. difficile-associated diarrhea (CDAD), the most common cause of nosocomial diarrhea. Cross-infection between patients and transmission through the environment and medical personnel are important factors in the acquisition of CDAD. In order to understand differences in epidemiology and pathogenesis, a number of typing schemes have been developed. We will review the typing methods used to study the epidemiology of C. difficile infections and how they have evolved from a phenotypic identification to state of the art molecular methods, detecting genetic polymorphisms among strains. These molecular methods include PCR-based methods (arbitrarily primed-PCR [AP-PCR] and PCR ribotyping), restriction endonuclease analysis (REA) and pulse field gel electrophoresis (PFGE). The application, usefulness and feasibility of these methods are compared and discussed. Finally, the role of genomics as a tool to investigate CDAD is introduced.
Broad band spectral measurements of EEG during emotional tasks.
Broad band spectral measurements were calculated on EEG recordings of 40 healthy male university students during emotional tasks using the emotion induction technique. Emotional stimuli were sentences with positive valence (joy of love), with negative valence (frustration) and neutral situation. Broad band measurements showed that delta and beta bands reflected better the differences by emotional state. Hemispheric differences associated to emotional states were obtained only for mean frequency data. Higher delta frequencies at left frontopolar, right frontal, parietal and temporal and higher beta frequencies at left frontotemporal and temporal were related to love. Meanwhile higher delta frequencies especially at left frontopolar and temporal regions and lower beta frequencies at midline frontal (Fz) and central (Cz) leads were associated to frustration. These findings suggest that narrow band measurements may reflect more efficiently EEG changes during emotional tasks.
In vitro metabolism of the COX-2 inhibitor DFU, including a novel glutathione adduct rearomatization.
The metabolic profile of DFU [5,5-dimethyl-3-(3-fluorophenyl)-4-(4-methylsulphonyl)phenyl-2(5H)-furanone], a potent and selective COX-2 inhibitor, was characterized using in vitro microsomal and hepatocyte incubations. A single product, corresponding to p-hydroxylation, p-OH-DFU [(5,5-dimethyl-3-(3-fluoro-4-hydroxyphenyl)-4-(4-methylsulphonyl)phenyl-2(5H)-furanone)], was produced in rat microsomal incubations of DFU. In contrast, three metabolites were produced in incubations using suspensions of freshly isolated rat hepatocytes. Microsomal production of the p-O-glucuronide metabolite of DFU from synthetic p-OH-DFU was shown to have chromatographic and mass spectrometric properties identical to the earliest eluting hepatocyte metabolite (M1). The molecular weights of the other two hepatocyte metabolites were readily obtained using capillary high-performance liquid chromatography continuous-flow liquid secondary ion mass spectrometry (HPLC/CF-LSIMS); however, the elemental composition of these metabolites was not. Unlike typical metabolic products, which produce readily identified increments in molecular weight, metabolites M2 and M3 produced molecular ions in positive- and negative-ion CF-LSIMS that were consistent with oxidation of DFU (+16 Da), followed by addition of glutathione (+306 Da) and subsequent loss of 20 and 18 Da, respectively. Capillary HPLC/high-resolution CF-LSIMS was used to generate accurate mass data for M2 and M3 that provided evidence that the losses of 20 and 18 Da, respectively, corresponded to a rearomatization through loss of HF or H(2)O. Isolation and NMR characterization provided the definitive structural proof for these metabolites. Overall, the metabolism of DFU in rat hepatocytes is proposed to proceed through an epoxide intermediate, which then either rearranges to the p-OH-DFU and is conjugated with glucuronic acid, or is trapped with glutathione, followed by rearomatization with loss of HF (M2) or H(2)O (M3).
[Central pontine myelinolysis and hyponatremia. Clinical case].
There is a controversy whether central pontine myelinolysis can complicate either hyponatremia or its rapid correction. We report a 69 years old woman with a history of one month of vertigo, nausea, vomiting and diarrhea. She was admitted dehydrated ad stuporous, and initial laboratory values showed a serum sodium of 96 mEq/L. She was treated with dextrose 5% and 3% NaCl. Serum sodium raised to 120 mEq/L at the next day and the level of consciousness improved. At the 4th day of admission, the patient was again stuporous and with spastic quadriplegia. A magnetic resonance imaging showed a central and symmetrical pontine lesion on T1 and T2 weighed images. Thereafter, the patient experienced a progressive improvement of her neurological condition and was discharged three months later, moving her lower limbs. Nine month later she was able to walk.
Detection of epithelial messenger RNA in the plasma of breast cancer patients is associated with poor prognosis tumor characteristics.
PURPOSE: Free plasma RNA has been scarcely studied in patients with cancer. Here we examine the presence of RNA from epithelial tumors in plasma from a series of breast cancer patients and its correlation with tumor characteristics and circulating tumor cells. EXPERIMENTAL DESIGN: beta-actin mRNA was analyzed to check the viability of plasma RNA in samples from 45 patients with breast cancer and 25 controls. Nested primers were used to detect the presence of cytokeratin 19 (CK19) and Mammaglobin in the same samples. Eleven clinicopathological parameters were studied and correlated with molecular parameters. Additionally, we looked for circulating tumor cells in 16 of these patients and in 10 of the controls. RESULTS: All samples showed detectable quantities of beta-actin RNA. In controls, 3 cases (12%) were positive for Mammaglobin, and 5 (20%) were positive for CK19 RNA; of the 45 patients, 27 cases (60%) were positive for Mammaglobin, and 22 (49%) were positive for CK19. These differences were statistically significant (P = 0.001). Tumor size (P = 0.01) and proliferative index (P = 0.02) were associated with the presence of Mammaglobin, CK19, or both RNAs in plasma. Pathological stage (P = 0.06) was close to significance. Although a statistical relationship was not demonstrated, 9 of the 10 patients with circulating tumor cells showed epithelial mRNAs in plasma. CONCLUSIONS: We conclude that epithelial tumor RNA is detectable in plasma from breast cancer patients and that this finding is associated with a probable poor prognosis and circulating tumor cells.
[Enterotoxin-producing Bacteroides fragilis strains isolated from horses].
Seven Bacteroides fragilis strains were cultured from samples collected from horses. From all the tested strains, as well as from the reference B. fragilis strains: enterotoxigenic NCTC 11925 and nonenterotoxigenic IPL 323 strain, DNA was isolated using Genomic DNA PREP PLUS isolation kit manufactured by A&A Biotechnology (Poland). To detect the enterotoxin (fragilysin) gene, polymerase chain reaction (PCR) was applied, using the following starters: 404 (GAG CCG AAG ACG GTG TAT GTG ATT TGT) and 407 (TGC TCA GCG CCC AGT ATA TGA CCT AGT). DNA obtained from bacterial cells was amplified in a thermocycler (Techne). The temperature profile was as follows: 1 cycle (4 min. 94 degrees C), 40 cycles (1 min. 94 degrees C, 1 min. 52 degrees C, 1 min. 74 degrees C). Amplification products were detected by electrophoresis in agarose gel (1%) with ethidium bromide added. The presence of the fragilysin gene was detected in two strains. Among the strains isolated from horses enterotoxin gene-possessing Bacteroides fragilis strains (ETBF) can be detected.
Cross-tool communication: from protocol authoring to eligibility determination.
To be effective, informatics tools for clinical trial protocols must inter-operate and share knowledge. We demonstrate a simple XML-based communication of eligibility criteria information between two independently-developed informatics tools. Using a shared DTD model of criteria, an authoring tool (developed within the Protégé environment) can send a list of eligibility criteria to a commercial system for automatic eligibility determination (the "iKnowChart" system by iKnowMed). The criteria model, developed as a Protégé ontology, includes both the terminology and the logic needed to compute eligibility for a given patient. As a demonstration of cross-tool communication, we have encoded criteria from an active clinical trial protocol (E1199), and shown how use of the authoring tool can effectively update the eligibility knowledge and the behavior of the commercial iKnowChart system. As part of the cross-tool knowledge sharing, we use Common Data Elements, an oncology terminology developed by the National Cancer Institute.
DNA damage after chemotherapy correlates with tumor response and survival in small cell lung cancer patients.
To explore the induction of chemotherapy (CT) DNA damage and its correlation with tumor response and patient survival, we undertook the present study in 20 small cell lung cancer (SCLC) patients. All patients underwent the same treatment based on CT courses of carboplatin and etoposide. Blood samples were taken before and immediately after CT and every 12 weeks during follow-up. Nuclear DNA damage was determined through the variations in three mitochondrial pseudogene mutations in DNA of peripheral blood mononuclear cells. They were detected by mutation-specific PCR and assessed by a semiquantitative method. The relative level of mutation rose after chemotherapy in all cases. Among the 11 patients (55%) with higher relative levels of mutations, 9 (82%) of them achieved a complete response. In contrast, of the 9 patients (45%) with lower relative levels of mutations, only 2 (18%) achieved a complete response, displaying a statistically significant difference (P=0.02). The overall survival for patients with marked genomic damage was 18 months (range 10-24), and for patients with low degree of DNA damage, it was 12 months (range 5-15) (P=0.002). Genomic damage detected after chemotherapy treatment correlates positively with tumor response and patient survival.
Absence of point mutations at codon 17 of the mdm2 gene (serine 17) in human primary tumors.
Mdm2 is a phosphoprotein that interacts with protein p53, inhibiting its activity. A serine located in position 17 of Mdm2, has been implicated in its phosphorylation process. We hypothesize that point mutations at serine 17 could block its phosphorylation and thereby increase the p53-Mdm2 interaction. This mechanism could increase the p53 degradation and cause a loss of the protective effect of p53 against tumorigenesis. This hypothesis was based on recent studies in vitro, demonstrating that when serine 17 is mutated, the DNA-dependent protein kinase, activated by genomic damage, is unable to phosphorylate it. Thus, we investigated whether structural point mutations at exon 3 of the Mdm2 gene, affecting codon 17, were present in 162 human primary tumors, 70 breast carcinomas, 14 bladder tumors, 18 colon adenocarcinomas and 60 testicular tumors. Direct sequencing of a fragment (204 bp) of exon 3 of the Mdm2 gene that contains the codon 17 showed no mutations at this position, independently of the presence or absence of p53 gene mutations in the same tumors. These results do not support the hypothesis that mutations in the Mdm2 gene at this level are involved in the tumorigenic process of human cancers.