Biomedical subjects
Publications and source records attributed to P Fumagalli.
There is still a major debate about the pathogenicity of Lactobacillus spp. and some reports emphasize that these microorganisms are never isolated from endovascular devices. In this report we present a case of catheter-related bacteremia due to L. rhamnosus in a patient who underwent a single-lung transplant.
Methicillin-resistant Staphylococcus aureus (MRSA) is frequently isolated in nosocomial outbreaks. In our study, we analysed the occurrence of colonisation and infection in an Intensive Care Unit of our hospital during a 12-month period. We also evaluated the possibility of using automated ribotyping as a molecular method in order to type the isolates. Twice a week a nasal swab and a rectal swab were performed on all patients; from ventilator-assisted patients, a sputum culture was also taken. All the MRSA isolated were identified by using commonly phenotypic procedures and on all isolates susceptibility tests were performed. An automated ribotyping using EcoRI was also done. Out of 292 patients enrolled in the study, 205 were never colonised (group N); among the other 87 who were colonised by MRSA (29.8%), 40 patients (group A) were MRSA carriers at the time of admission, while 47 (group B) were colonised in the ICU. Twenty-seven patients (11 from group A, 15 from group B and 1 from group N) developed 31 infections due to MRSA. Patients from group A exhibited, as a rule, worse clinical conditions than those from the other two groups. For the former group, MRSA infection was frequently systemic (sepsis), while in group B pneumonia was the predominant infection. The prevalence of colonisations in our study was 30%, which is a value comparable to those presented by other authors in similar cases. MRSA colonisation is a necessary condition for subsequent infections in almost all cases, with an average lag of 7 days. Susceptibility tests were non-discriminating among the isolates: all the strains were susceptible to glycopeptides; nearly all of them were resistant to erythromycin, clindamycin, ciprofloxacin and gentamicin. Automated ribotyping allowed us to distinguish 12 different ribogroups, the most frequent of which was composed of 146 isolates. In our study, this molecular method was able to define a possible endemic clone that should be better investigated by using methods with a higher discriminatory power, such as RAPD or PFGE. The method that we employed is highly reliable, easy to perform and not time-consuming. In our opinion, it could be the method of choice in the first screening of high numbers of isolates.
We applied cDNA selection methods to a genomic clone (YAC 761B5) from chromosome 21 located in the so-called 'Down critical region' in 21q22.3. Starting from human fetal heart and brain mRNAs we obtained and sequenced several cDNA clones. One of these clones (Down region aspartic protease (DRAP), named also BACE2 according to the gene nomenclature) revealed a striking nucleotide and amino acid sequence identity with several motifs present in members of the aspartic protease family. In particular the amino acid sequences comprising the two catalytic sites found in all mammalian aspartic proteases are perfectly conserved. Interestingly, the predicted protein shows a typical membrane spanning region; this is at variance with most other known aspartic proteases, which are soluble molecules. We present preliminary evidence, on the basis of in vitro translation studies and cell transfection, that this gene encodes a glycosylated protein which localizes mainly intracellularly but to some extent also to the plasma membrane. Furthermore DRAP/BACE2 shares a high homology with a newly described beta-secretase enzyme (BACE-1) which is a transmembrane aspartic protease. The implications of this finding for Down syndrome are discussed.
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We report the cloning of the human homologue of the rat metalloprotease N-arginine dibasic convertase (NRD convertase). This endopeptidase is responsible for the processing, at the Arg-Lys dibasic site on the N-terminal side of the arginine residue, of propeptides and proproteins. Comparisons of the human and rat full-length cDNAs show similarity and identity of 94 and 91%, respectively. In humans NRD convertase is predominantly expressed in heart, skeletal muscle, and testis. We have also studied the expression of this gene in mouse at various developmental stages and found that the neural tissue is the almost exclusive site of expression in early development (between E 10.5 and E 16.5). To gain information about the possibility that defects in this gene are linked to inherited neuromuscular disorders, we determined the chromosomal location of the human NRD convertase gene by FISH analysis, showing that the gene resides at 1p32.2.
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The identification and functional characterization of genes on chromosome 21 is a necessary step to understand the pathogenesis of the various phenotypic anomalies that affect Down syndrome patients. Using direct cDNA selection we have identified a new gene, SH3BGR, that maps to 21q22.3, proximal to HMG14, and is differentially expressed in heart and skeletal muscle. SH3BGR encodes a novel protein that is characterized by the presence of a proline-rich region containing the consensus sequence for a SH3-binding domain and by an acidic carboxyl-terminal region containing a glutamic acid-rich domain predicted to assume a coiled coil. The presence of two functional domains involved in protein-protein interactions suggests that SH3BGR could be part of a multimeric complex. Its overexpression might alter specific functions of muscular tissue and therefore take part in the pathophysiology of muscular hypotonia in Down syndrome.
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The present findings provide experimental evidence for the hypothesis that an impairment of mitochondrial function may be involved in manganese neurotoxicity. Specifically, the treatment of dopaminergic neuronal-derived cell line (PC12) with MnCl2 produced a significant inhibition of some mitochondrial complexes of the respiratory chain, while in the glial-derived cell line (C6) this effect was not observed. In PC12 the decrease in complex I activity was more pronounce than in other mitochondrial complexes. However treatment of cells with ZnSO4 exerted no significant variations in enzymatic activities. A direct exposure of mitochondrial fraction to MnCl2 reduced enzymatic activities of mitochondria in both cell lines adding further support to the proposed theory that the different sensitivity of the cells to manganese may be explained by a difference in uptake or intracellular storage. These data indicate that manganese neurotoxicity could be the result of a direct effect just on complex I activity or due to a secondary effect of oxidative stress induced by an excess of this transition metal.
This report describes one complication related to retrograde positioning of a catheter in the jugular vein in a patient in a coma resulting from subarachnoid haemorrhage. The catheter was found in the cervical subarachnoid space, as confirmed by radiography with contrast medium. Attention is focused on the fact that this technique, usually performed easily and safely, may occasionally present potentially severe complications.
Azospirillum brasilense glutamate synthase, a complex iron-sulfur flavoprotein, was subjected to limited proteolysis using trypsin and chymotrypsin, in the absence or presence of its substrates or their analogs. Time-dependent degradation of glutamate synthase alpha and beta subunits, to yield several fragments of different stability, was observed, the alpha subunit being more sensitive than the beta to proteolytic attack. The main sites of proteolytic cleavage were determined by densitometric analysis of the electrophoretic patterns obtained under denaturing conditions and by N-terminal sequencing of the major proteolytic products. These analyses showed that most of the peptide bonds sensitive to the proteases are clustered in two regions of the alpha subunit, outside the proposed substrate and cofactor binding regions of glutamate synthase [Pelanda, R., Vanoni, M. A., Perego, M., Piubelli, L., Galizzi, A., Curti, B. & Zanetti, G. (1993) J. Biol. Chem. 268, 3099-3106]. Therefore, these protease-sensitive sites can be identified as flexible loops, exposed to solvent, connecting adjacent domains of the protein. The presence of the enzyme substrates or their analogs caused significant changes in the proteolytic patterns. NADP+ protected the C-terminal region of glutamate synthase beta subunit from tryptic cleavage, supporting the proposal that it contains the pyridine-nucleotide-binding site. Furthermore, NADP+, and to a lesser extent the glutamine analog L-methionine sulfone, which binds presumably to the N-terminal region of the alpha subunit, altered the sensitivity to proteolysis of the sites of the alpha subunit proposed to be part of links between domains of glutamate synthase. These results show that long-range conformational changes of glutamate synthase occur on binding of its substrates. The study of several NADPH-dependent diaphorase activities of glutamate synthase was also undertaken in order to test if proteolytic fragments of the enzyme retained their ability to transfer electrons from NADPH to synthetic electron acceptors. Although proteolysis yielded partial loss of all enzyme NADPH-dependent reactions, the kinetic analysis showed that the rates of reduction of iodonitrotetrazolium, ferricyanide and dichlorophenolindophenol were at least twofold faster than the rate of the physiological glutamate synthase reaction. These results indicate that enzyme reduction and intramolecular electron transfer are not rate limiting during catalysis of the physiological glutamate synthase reaction.
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A patient's admission to an ICU should benefit the patient (either short or long term). Consequently patients admitted to the ICU should be only those neurological-neurosurgical ones with one or more organ failure who need immediate or prolonged therapy with adequate monitoring and technical support not available in other wards. Normally admission depends on may other factors, the most important ones seem to be medico-legal responsibilities of the possible refusal of admission even if due to clinical reasons, organizational problems such as equipment and bed availability. There is a need for pre- and post-surgical monitoring for high risk patients. We routinely admit patients to our ICU according to the following directions: respiratory and or cardio-circulatory failure connected to the neurological pathology, impaired consciousness, neurological signs and symptoms indicating deterioration, loss of airway protective reflexes, seizured in neurological-neurosurgical patients, severe electrolyte, metabolic or nutritional disturbances which may affect the CNS; neurosurgical postoperative control, performance of complex techniques (central venous and arterial catheterization, ICP monitoring), management of multiorgan donors suitable for therapeutic transplant. The groups of pathologies observed in our ICU during 1991 where the follow-up: 442 neurological-neurosurgical patients--head trauma 141 (32%)--cerebral tumours 88 (19.9%)--vasculopathies 135 (30.6%)--others 77 (17.5%). 228 (51.5%) patients stayed for a period less than three days. According to our experience we can affirm that causes for patient's admission to neuro-ICUs are various, the legal organizational reasons have a great importance compared to the clinical ones.
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