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C Fernandez

Publications and source records attributed to C Fernandez.

At least 109 records · Page 6Linked to original sources
PubMed1998

The value of histological examination in the audit of hospital autopsies: a quantitative approach.

The aims of this study were to compare the clinical with autopsy diagnoses, to evaluate the role of histological examination in the pathological diagnoses and to assess the new pathological diagnoses uncovered by autopsy. We aimed to obtain quantitative assessment of the sensitivity, specificity and accuracy of clinical diagnoses. The guidelines for postmortem reports by the Royal College of Pathologists (1993) were implemented for reports used in this study. These guidelines are similar in intent to those of the College of American Pathologists. Complete macroscopic and histological studies of 108 (53 females) autopsies were analysed. The mean age was 78.0+/-9.0 (SD) years (range 54-94 years). The interquartile range (25%ile 75%ile) was 72-84 years, with a median of 79.5 years. Seventy per cent of all causes of death were confirmed by macroscopical and histological examination. Sixty-one clinical diagnoses were inconsistent with the pathological findings. Histological examination contributed significantly to the final diagnosis in major (5%) and minor (6%) clinicopathological as well as new pathological findings (23%). The most common causes of death not suspected clinically were pulmonary embolism (23%), bronchopneumonia (22%), ischemic heart disease (13%) and malignancies (10%). The clinical sensitivity of antemortem diagnoses was 25% for peritonitis and 24% for pulmonary embolism. The overall clinical sensitivity was 54% and specificity 92%. The accuracy of positive diagnosis was 69% and accuracy of negative diagnosis 88%. Unexpected causes of death represented a third of all causes of death reported. Histological examination is an important tool in hospital autopsy audit. A quantitative approach can be used to assess the accuracy of postmortem clinical diagnoses, to identify the possible source of clinical diagnostic weakness, and provide data that may be of use for diagnostic precision in the more difficult clinical subjects.

Aged↗
PubMed1998

Germinal center formation following immunization with the polysaccharide dextran B512 is substantially increased by cholera toxin.

We have compared the splenic responses following immunization with the T cell-independent (TI)-2 antigen native dextran B512 and with a thymus-dependent (TD) protein-dextran conjugate. Interestingly, primary immunization with native dextran induced germinal center (GC) formation in the spleen to the same extent as the protein-dextran conjugate. The GC were antigen-specific as characterized by the presence of peanut agglutinin (PNA)-positive areas that were also binding FITC-conjugated dextran. Dextran-binding B cells were also detected outside the GC as sites of antibody-producing cells. The secondary splenic response to native dextran was suppressed compared to the primary response, with almost no dextran-specific GC or extra-follicular sites with dextran-specific B cells present in the sections. This suppression could be reverted by using cholera toxin (CT) as an adjuvant for the dextran immunizations. Following native dextran immunization with CT adjuvant a secondary splenic GC response similar to a TD secondary splenic GC response was generated, with almost all the dextran-specific B cells located in the GC. Collectively, this indicates that the difference between TI and TD antigen responses is not due to different abilities in inducing GC development, rather the GC reaction is less productive for a TI antigen than for a TD antigen. CT can both increase secondary GC formation in particular for the TI form of dextran and ameliorate the GC reaction, as reflected by increased anti-dextran antibody levels.

Adjuvants, Immunologic↗
PubMed1998

High-performance liquid chromatography with ultraviolet and fluorimetric detection for the simultaneous determination of tacrine, nimodipine, and their respective metabolites in the plasma of patients with Alzheimer disease.

A new high-performance liquid chromatography (HPLC) assay was developed for the simultaneous determination of tacrine (THA), nimodipine, and their three metabolites (MI, MII, and MIII) using a 1-ml plasma sample volume. A liquid-liquid extraction procedure was coupled with a reverse-phase HPLC separation. Quantification was performed by fluorometric detection for THA and metabolites and by ultraviolet detection for nimodipine and metabolites. Peak-height ratios were linear across the ranges 0.5 to 100 micro/l for THA and its three metabolites; 2 to 500 microg/l for nimodipine, MII, and MIII; and 4 to 500 microg/l for MI. Correlation coefficients were better than 0.998 for all compounds. Accuracy and precision were less than 12% for the entire concentration range for each substance. This method is sensitive and selective. Analysis of plasma samples collected from patients with Alzheimer disease demonstrated that the assay is suitable for clinical and pharmacokinetic trials including drug-drug interactions studies.

Alzheimer Disease↗
PubMed1998

Denoising of the uterine EHG by an undecimated wavelet transform.

We propose two original methods of denoising of the uterine electrohysterography (EHG) signal by wavelets. This external electrophysiological signal is corrupted by electronic, electromagnetic noises and by the remaining electrocardiogram of the mother. The interfering signals have overlapping spectra. Therefore, a classical filtering is unusable. Wavelets should be a very well-suited denoising tool. The first proposed method uses the algorithm "à trou" with nonsymmetrical filters. The computation is rapid and the results are satisfying compared to the classical denoising techniques. The second algorithm is an improvement of the first method. It uses orthogonal wavelets and the result of the thresholding corresponds to the average of all circulant shifts denoised by a decimated wavelet transform. Results are compared to traditional denoising algorithms by wavelet (orthogonal, maximally decimated). The proposed algorithms are more efficient on simulated signals as well as on uterine EHG.

Algorithms↗
PubMed1998

Physicochemical characteristics of pentamidine-loaded polymethacrylate nanoparticles: implication in the intracellular drug release in Leishmania major infected mice.

This work describes the preparation, the physicochemical properties, the tolerance and the intracellular trafficking of pentamidine loaded nanoparticles. Pentamidine was bound to the polymer by ionic interaction. This interaction involved the carboxylic acid functions of methacrylic acid (10% of the polymer) and the amine groups of the drug. Pentamidine fixation and release were pH dependent. An acidic pH led to a decrease of fixation or a release. At pH 5, which is the pH value of lysosomes and parasitophorous vacuoles, the release reached up to 50%. At this pH value, pentamidine is ionized and therefore can not traverse the biological membranes. Unloaded nanoparticles and pentamidine-loaded nanoparticles were tested in vitro on U937 cells and no cytotoxicity was observed. In vivo, in Leishmania infected mice, no significant weight loss was found. Ultrastructural studies showed the different steps of drug loaded nanoparticles trafficking inside Leismania-infected Küpffer cells. The nanoparticle uptake by macrophagic cells led to the location of nanoparticles inside phagocytosis vacuoles which fused with primary lysosomes to form secondary lysosomes. Ultimate fusion of secondary lysosomes containing nanoparticles with parasitophorous vacuoles was also observed. Nanoparticles were identified close to amastigotes but internalization by the parasite was not observed.

Animals↗
PubMed1997

Minimally invasive management of low-grade and benign gastric tumors.

BACKGROUND: Benign gastric tumors and tumors of low-grade malignancy can be safely removed laparoscopically. METHODS: Seven patients were considered candidates for laparoscopic resection of gastric tumors. Inclusion criteria included small tumor size (less than 6 cm), exophytic or endophytic tumor morphology, and benign characteristics. Indications for surgical intervention included bleeding, weight loss, and need for tissue diagnosis. Patients ranged in age from 38 to 70. There were five female and two male patients. All patients underwent preoperative upper GI endoscopy. The procedures were performed using a four- or five-port technique. An Endo-GIA (US Surgical Company, Norwalk, Connecticut) was used to amputate those tumors located on the serosal surface of the stomach. Tumors on the mucosal surface were exposed via a gastrotomy, then likewise amputated using an Endo-GIA. The gastrotomy closure was then either hand sewn or stapled. Operating time ranged from 95 to 225 min. RESULTS: Final pathologic diagnoses included lipoma, lymphoma, leiomyoma, and leiomyosarcoma. There was a 28% conversion rate. There were no complications. Length of postoperative stay ranged from 4 to 7 days. There have been no tumor recurrences in 6-38-month follow-up. CONCLUSIONS: Minimally invasive management of benign and low-grade gastric tumors can be performed safely with excellent short- and long-term results.

Adult↗
PubMed1997

Quantitative study of the short range order in B2O3 and B2S3 by MAS and two-dimensional triple-quantum MAS 11B NMR.

Two-dimensional multiple-quantum magic angle spinning (MQMAS) NMR and MAS NMR of 11B at various magnetic fields, were applied to elucidate the structure of vitreous (glassy) boron trioxide (v-B2O3), vitreous boron trisulfide (v-B2S3) and crystalline boron trisulfide (c-B2S3). These techniques, when combined with computer simulations of the resulting spectra, provide the isotropic chemical shifts and the quadrupole parameters, as well as a quantitative measure of the intensities of various boron resonances. The MAS NMR of v-B2O3 produced overlapping anisotropic lineshapes corresponding to the -1/2<-->1/2 transition in two distinct types of BO3 units with 3(+/-0.08):] intensity ratio. A combination of MAS and the multiple-quantum method resulted in a better resolved, isotropic 11B spectrum of v-B2O3. A remarkable enhancement of resolution of the MQMAS NMR proved instrumental in finding and identifying various impurities present in v-B2S3 and c-B2S3. In addition to the resonances from boron in two types of BS3 groups, four other structural units, BOS2, BO2S, BO3 and BS4, were elucidated from the spectra of vitreous and crystalline samples. The effects of various experimental parameters, such as the magnitude of the B0 and B1 fields, on the resolution of the MAS and MQMAS techniques are also shown.

Algorithms↗
PubMed1997

Multiple-quantum magic-angle spinning NMR with cross-polarization: spectral editing of high-resolution spectra of quadrupolar nuclei.

An experiment is presented that combines the multiple-quantum magic-angle spinning (MQMAS) technique with cross-polarization (CP). As a preliminary test of this new method, we measured and compared the 27Al 3QMAS and 19F-->27Al CP 3QMAS spectra of a fluorinated AlPO4 aluminophosphate. Complete discrimination between the fluorinated and nonfluorinated Al sites was easily achieved, which demonstrates the usefulness of CP MQMAS for spectral editing. Future applications of this experiment will include other spin pairs and heteronuclear correlation NMR spectroscopy.

Magnetic Resonance Spectroscopy↗
PubMed1997

Solid-state NMR studies of interstitial phosphorus atoms in rhodium carbonyl clusters.

Pure samples and as-prepared mixtures of Rh9 and Rh10 carbonyl clusters with interstitial P atoms have been studied quantitatively by 31P MAS and 1H-31P CP/MAS NMR. Information on the 31P chemical shift tensor of the Rh9 and Rh10 clusters has been derived from spinning sideband simulations. The chemical shift anisotropy is slightly larger in the Rh10 clusters (340-400 ppm) than in the Rh9 clusters (230-300 ppm), while the asymmetry parameters are similar (eta = 0.1-0.4). The results contribute to the understanding of the relationship between the shielding anisotropy and the structure of the cluster cavity.

Anisotropy↗
PubMed1997

Immunogenicity of bacterial carbohydrates: cholera toxin modulates the immune response against dextran B512.

Native dextran B512 is a T-cell-independent (TI) antigen. By conjugating low molecular weight (MW) dextran to protein, a T-cell-dependent (TD) response against dextran can be obtained. We have previously reported the effects of native dextran and two different protein-dextran conjugates on the immune system. While one type of conjugate induced an optimal TD response, the other conjugate ('suboptimal') evoked a response more similar to that induced by native dextran, i.e. with little immunoglobulin class switch and with a secondary response of similar magnitude to the primary response. In order to investigate if it was possible to augment the anti-dextran response we examined the effects of cholera toxin (CT) in our dextran model system. CT is a potent mucosal, as well as systemic, adjuvant with effects on T cells, B cells and antigen-presenting cells. We show that CT is a very efficient adjuvant for both the TD and TI forms of dextran. A major increase in IgM and IgG anti-dextran antibody production was detected after administration of CT together with the conjugates compared with a conventional alum adjuvant. The effect was most pronounced for the suboptimal TD conjugate. CT was also able partially to abrogate the unresponsiveness to dextran in the TI type 2 (TI-2) non-responder strain CBA/N. CT was also found to be a very potent adjuvant for native dextran, secondary IgM levels were enhanced eightfold by the co-administration of CT. Furthermore CTB-Dx, which is a conjugate of the non-toxic part of CT and low MW, non-immunogenic dextran, elicited an anti-dextran response in nude mice. Collectively, our data show that it is possible to improve the immunogenicity of both TD and TI forms of a carbohydrate by co-administration of CT. This is indicative of two components of the adjuvant effect, one could enhance antigen presentation and costimulation of T cells and the other could have a direct stimulatory effect on B cells.

Adjuvants, Immunologic↗
PubMed1997

Pathways of protein degradation in L6 myotubes.

To assess the importance of different proteolytic pathways in anabolic and catabolic states, L6 myotubes were labeled with 3H-phenylalanine for 24 hr, then exposed for another 24 hr to control medium, alone or with one of the following: insulin (1 microgram/ml), insulin-like growth factor-I (IGF-I, 100 ng/ml), clenbuterol (100 microM), or dexamethasone (1 microM). Release of acid-soluble radioactivity and 3-methylhistidine were used to determine rates of total and myofibrillar protein degradation, respectively. Within each hormone treatment, cells were also exposed to none, one, or both of the following protease inhibitors: chloroquine (25 microM); E-64d (100 microM). Chloroquine inhibits lysosomal proteases, and E-64d is a cell-penetrating inhibitor of cysteine proteases (including the calpains). Total protein degradation was reduced by insulin (-33%) and IGF-I (-40%), increased by clenbuterol (+20%), and was unaffected by dexamethasone. Myofibrillar protein degradation was reduced by insulin (-35%), IGF-I (-55%), and clenbuterol (-22%), and was unaffected by dexamethasone. Therefore, total (mainly sarcoplasmic) and myofibrillar protein degradation were regulated independently of one another. The influence of the inhibitors was not consistent across hormone treatments. Chloroquine inhibited total but not myofibrillar proteolysis, indicating that lysosomes may be rate limiting in the former but not the latter. In contrast, E-64d had no effect on total protein degradation, but increased myofibrillar proteolysis. Interactions between anabolic and catabolic hormones and these two protease inhibitors indicate that both lysosomal and nonlysosomal proteolytic pathways are involved in regulation of total protein degradation in myotubes. In contrast, lysosomes are probably not involved in regulating the rate of myofibrillar protein degradation, the initial step likely being due to calpain activity.

Animals↗
PubMed1996

Critical issues in chiral drug analysis in biological fluids by high-performance liquid chromatography.

This review article focuses on the specificities of chiral liquid chromatography, with particular emphasis on stability, stereoconversion, enantiomeric separation, recovery and drug concentration determinations. In addition, the paper presents an overview of the different steps which have to be followed for a chiral method to be validated. Sensitivity, selectivity, linearity, precision and accuracy all have to be ensured for three chemical entities, the two enantiomers and the racemate. Only accurate and precise concentrations of the parent drug and its metabolites will lead to the reliable description of their in vitro stability and in vivo body disposition.

Chromatography, High Pressure Liquid↗
PubMed1996

Ultrastructural analysis of mineralized matrix from human osteoblastic cells: effect of tumor necrosis factor-alpha.

Recent work by a number of investigators has demonstrated that the process of bone matrix formation and mineralization is under the influence of growth factors and cytokines present in the local environment. Utilizing primary and established osteoblast cell culture systems, these studies have examined the regulation of bone matrix protein synthesis and deposition into the extracellular matrix (ECM) and subsequent mineralization. In previous studies, we have utilized the human osteoblastic cell line, HOS TE85, to study the effects of Tumor Necrosis Factor-alpha (TNF-alpha) on the regulation of matrix proteins and proteolytic function in monolayer cultures as well as during the development and calcification of ECM formed by HOS TE85 cells during extended culture. Our studies demonstrate that TNF-alpha inhibited formation and mineralization of nodules. In the study reported here, we evaluated the ultrastructural morphology of the cell-matrix complex formed by HOS TE85 cells in the presence and absence of TNF-alpha at selected time points during the matrix development process utilizing both transmission electron microscopy and light microscopy. In the presence of TNF-alpha, the cell-matrix complex does not develop normally, with a lack of organization and mineralization, when compared to untreated cells. The lack of mineralization appears to result from the lack of normal collagen fibril deposition and formation of an appropriate ECM essential for the mineralization process. These results support our previous observations that TNF-alpha inhibits HOS TE85 cells from forming a mineralizing ECM by inhibiting incorporation of collagen into the ECM and inducing the synthesis of proteolytic enzymes capable of degrading collagen in the ECM.

Calcification, Physiologic↗
PubMed1996

Separation of testosterone metabolites in microsomal incubates using a new capillary electrophoresis assay.

A capillary electrophoresis method has been developed to separate the products of liver microsomal testosterone metabolism. The microsomal mixture undergoes liquid-liquid extraction and pre-concentration, and then electrophoretic analysis takes less than 25 min including capillary conditioning steps. The development of the complex background electrolyte (Tris-HCl and borate buffers, sodium dodecyl sulfate, beta-cyclodextrin and ethanol) necessary for this separation is described. A z-type capillary flow cell is used to obtain adequate detection sensitivity. The proportion in which the metabolites are produced as determined by this method allows assignment of the relative activity of cytochrome P-450 enzymes in the microsomes. The technique is useful for comparison of activity in normal and abnormal hepatic microsomes.

Animals↗
PubMed1996

Expression and folding of an interleukin-2-proinsulin fusion protein and its conversion into insulin by a single step enzymatic removal of the C-peptide and the N-terminal fused sequence.

We report the expression in E. coli of a proinsulin fusion protein carrying a modified interleukin-2 N-terminal peptide linked to the N-terminus of proinsulin by a lysine residue. The key aspects investigated were: (a) the expression of the fused IL2-PI gene, (b) the folding efficiency of the insulin precursor when still carrying the N-fused peptide and (c) the selectivity of the enzymatic cleavage reaction with trypsin in order to remove simultaneously the C-peptide and the N-terminal extension. It was found that this construction expresses the chimeric proinsulin at high level (20%) as inclusion bodies; the fused protein was refolded at 100-200 micrograms/ml to yield about 80% of correctly folded proinsulin and then it was converted into insulin by prolonged reaction (5 h) with trypsin and carboxypeptidase B at a low enzyme/substrate rate (1:600). This approach is based on a single enzymatic reaction for the removal of both the N-terminal fused peptide and the C-peptide and avoids the use of toxic cyanogen bromide.

Amino Acid Sequence↗
PubMed1996

Triple-quantum MAS-NMR of quadrupolar nuclei.

From two-dimensional multiquantum NMR spectra of quadrupolar nuclei, it is now possible to obtain much greater resolution than in a classical single-quantum magic-angle spinning or variable-angle spinning spectrum. We describe here a very simple pulse scheme which efficiently excites the desired multiquantum NMR coherence and a new acquisition procedure which yields to pure-absorption mode 2D spectra. Experimental spectra for 87Rb in polycrystalline rubidium nitrate illustrate the method.

Magnetic Resonance Spectroscopy↗