Identification of HLA-B*4905 in a Caucasian family.
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Alleles↗
Biomedical subjects
L Castellani
Publications and source records attributed to L Castellani.
Feedback inhibition by RALT controls signal output by the ErbB network.
The ErbB-2 interacting protein receptor-associated late transducer (RALT) was previously identified as a feedback inhibitor of ErbB-2 mitogenic signals. We now report that RALT binds to ligand-activated epidermal growth factor receptor (EGFR), ErbB-4 and ErbB-2.ErbB-3 dimers. When ectopically expressed in 32D cells reconstituted with the above ErbB receptor tyrosine kinases (RTKs) RALT behaved as a pan-ErbB inhibitor. Importantly, when tested in either cell proliferation assays or biochemical experiments measuring activation of ERK and AKT, RALT affected the signalling activity of distinct ErbB dimers with different relative potencies. RALT deltaEBR, a mutant unable to bind to ErbB RTKs, did not inhibit ErbB-dependent activation of ERK and AKT, consistent with RALT exerting its suppressive activity towards these pathways at a receptor-proximal level. Remarkably, RALT deltaEBR retained the ability to suppress largely the proliferative activity of ErbB-2.ErbB-3 dimers over a wide range of ligand concentrations, indicating that RALT can intercept ErbB-2.ErbB-3 mitogenic signals also at a receptor-distal level. A suppressive function of RALT deltaEBR towards the mitogenic activity of EGFR and ErbB-4 was detected at low levels of receptor occupancy, but was completely overcome by saturating concentrations of ligand. We propose that quantitative and qualitative aspects of RALT signalling concur in defining identity, strength and duration of signals generated by the ErbB network.
3T3 Cells↗
In which ways have the endovascular techniques modified the management of polyvascular patients?
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Aged↗
An immunogenetic map of Lombardy (Northern Italy).
For this study we consulted the Bone Marrow Donors' Registry of Lombardy (Italy) and analyzed 43937 HLA-A,B phenotypes and 13922 HLA-A,B,DR phenotypes. We estimated the HLA-A,B and HLA-A,B,DR haplotype frequencies via the maximum-likelihood method. We analyzed the genetic structure of the 11 provinces of Lombardy by means of Principal Component Analysis and Correspondence Analysis, and estimated the variety of the different haplotypes at provincial level and the percentage of unique phenotypes at village level. We found 11189 different HLA-A,B phenotypes, 661 different HLA-A,B haplotypes and more than 4000 different HLA-A,B,DR haplotypes. We identified 20 villages, in Western Lombardy, very rich in unique/rare phenotypes. Here we report a formula which allows the identification of a putative donor matched for two haplotypes with a recipient. This result may be of great importance for the genetic study of the population of Lombardy and, even more, for bone marrow transplantation programs.
Alleles↗
A feasibility study of a single event spectrometer based on semiconductor devices.
The electronics employed around particle accelerators can be disturbed or damaged because of single event effects (SEE). The most likely effect is the single event upset (SEU) which may affect all memory devices. In the case of high energy accelerators, SEUs are mostly produced by secondary charged particles generated by neutron interactions. The measurement of the energy and the lineal energy distribution of these neutron-induced charged particles was proposed. As a first approach, a commercial p-i-n photodiode was employed. This device was irradiated with thermal and monoenergetic fast neutrons. Some effects limiting the use of such a detector as a SEE spectrometer were observed, giving guidelines for the design of an application specific integrated circuit (ASIC). The possibility of creating a solid state microdosemeter by coupling the ASIC with a tissue-equivalent radiator is discussed. Moreover, the p-i-n photodiode covered with a hydrogenated plastic radiator may be employed as a proton-recoil spectrometer.
Dose-Response Relationship, Radiation↗
Aortic aneurysm: construction of a life-size model by rapid prototyping.
Development of new endovascular techniques for repair of abdominal aortic aneurysm (AAA) requires the use of experimental models. Stereolithography is a rapid prototyping technique used in industry to prototype parts during the design phase. A stereolithography apparatus (STL) employs laser technology to build a digital model layer by layer with photopolymer resin. The purpose of this study was to use this technology to produce a life-size AAA model. Data were acquired by CT scan and stored in DICOM 3 format. Specifically designed software was used for 3-D imaging and conversion of data to a standard STL format. Two replicas were made: one to scale and the other 3 mm larger. The final model was made by pouring silicone rubber or polyurethane into the mold over the life-size model so as to obtain a sturdy, life-size, soft, transparent plastic casting. Arterial models made for living subjects with these rapid prototyping techniques can be used to simulate surgical procedures, calibrate imaging modalities, and design new stent grafts.
Aortic Aneurysm, Abdominal↗
Intracellular localization and isoform expression of the voltage-dependent anion channel (VDAC) in normal and dystrophic skeletal muscle.
Voltage-dependent anion channels (VDACs) are a family of pore-forming proteins encoded by different genes, with at least three protein products expressed in mammalian tissues. The major recognized functional role of VDACs is to permit the almost free permeability of the outer mitochondrial membrane (OMM). Although VDAC1 is the best known among VDAC isoforms, its exclusively mitochondrial location is still debated. Therefore, we have measured its co-localization with markers of cellular organelles or compartments in skeletal muscle fibers by single or double immunofluorescence and traditional as well as confocal microscopy. Our results show that VDAC1 immunoreactivity corresponds to mitochondria and sarcoplasmic reticulum, while sarcolemmal reactivity, previously reported, was not observed. Since VDAC1 has been suggested to be involved in the control of oxidative phosphorylation, we sought for possible gene regulation of VDAC1, VDAC2 and VDAC3 in skeletal muscle of the dystrophin-deficient mdx mouse, which suffers of an impaired control of energy metabolism. Our results show that, while VDAC1 mRNA and protein and VDAC2 mRNA are normally expressed. VDAC3 mRNA is markedly down-regulated in mdx mouse muscle at different ages (before, during and after the outburst of myofiber necrosis). This finding suggests a possible involvement of VDAC3 expression in the early pathogenic events of the mdx muscular dystrophy.
Animals↗
SDF-1alpha-mediated modulation of synaptic transmission in rat cerebellum.
The functional expression of the seven-transmembrane domain G protein-coupled chemokine receptor CXCR-4/fusin in rat nerve cell was demonstrated by staining with a polyclonal anti-CXCR-4 Ab, and by evaluating the calcium responses to the physiological agonist stromal-derived cell factor-1alpha (SDF-1alpha) in both cerebellar granule cells in culture and Purkinje neurons (PNs) in cerebellar slices. Cerebellar glial, granule and Purkinje cells showed a pronounced staining for CXCR-4. Furthermore, cultured granule cells exhibited Ca2+ transients elicited by the application of SDF-1alpha, both in cell bodies and in neuronal processes. Whole-cell patch-clamped PNs in cerebellar slices responded to SDF-1alpha application by a slow inward current followed by an increase of both intracellular Ca2+ level and spontaneous synaptic activity. In particular, the SDF-1alpha-induced slow inward current was considerably reduced by ionotropic glutamate receptor blockers, but developed fully in a medium in which synaptic transmission was inhibited, indicating that this current might be, at least in part, mediated by extrasynaptic glutamate, possibly released from the surrounding glial and/or nerve cells. Taken together, these findings indicate a functional involvement of CXCR-4 in the modulation of synaptic transmission, adding another member to the repertoire of the chemokine receptors exerting a neuromodulatory role in the cerebellum.
6-Cyano-7-nitroquinoxaline-2,3-dione↗
Inhibition of ErbB-2 mitogenic and transforming activity by RALT, a mitogen-induced signal transducer which binds to the ErbB-2 kinase domain.
The product of rat gene 33 was identified as an ErbB-2-interacting protein in a two-hybrid screen employing the ErbB-2 juxtamembrane and kinase domains as bait. This interaction was reproduced in vitro with a glutathione S-transferase fusion protein spanning positions 282 to 395 of the 459-residue gene 33 protein. Activation of ErbB-2 catalytic function was required for ErbB-2-gene 33 physical interaction in living cells, whereas ErbB-2 autophosphorylation was dispensable. Expression of gene 33 protein was absent in growth-arrested NIH 3T3 fibroblasts but was induced within 60 to 90 min of serum stimulation or activation of the ErbB-2 kinase and decreased sharply upon entry into S phase. New differentiation factor stimulation of mitogen-deprived mammary epithelial cells also caused accumulation of gene 33 protein, which could be found in a complex with ErbB-2. Overexpression of gene 33 protein in mouse fibroblasts inhibited (i) cell proliferation driven by ErbB-2 but not by serum, (ii) cell transformation induced by ErbB-2 but not by Ras or Src, and (iii) sustained activation of ERK 1 and 2 by ErbB-2 but not by serum. The gene 33 protein may convey inhibitory signals downstream to ErbB-2 by virtue of its association with SH3-containing proteins, including GRB-2, which was found to associate with gene 33 protein in living cells. These data indicate that the gene 33 protein is a feedback inhibitor of ErbB-2 mitogenic function and a suppressor of ErbB-2 oncogenic activity. We propose that the gene 33 protein be renamed with the acronym RALT (receptor-associated late transducer).
3T3 Cells↗
Semiclosed endarterectomy for long superficial femoral artery atherosclerotic occlusive disease. One-year angiographic results.
BACKGROUND: The anatomic results of endarterectomy of long occlusive lesions in the superficial femoral artery (SFA) are required for reconsidering this procedure and comparing the results with those of newly developed endovascular techniques. We designed a prospective study to determine the arteriographic findings one year after a successful semiclosed endarterectomy of long occlusive lesions in the SFA. METHODS: From January 1995 until July 1996, an endarterectomy in the SFA was attempted in 12 successive patients and was successfully performed in 10 patients (6 men and 4 women), with an average age of 73 years (range 66 to 90 years). Indications for successfully performed procedures were claudication in 6, non healing ulcer in 2, local gangrene in 2 patients. The mean length of the occlusion was 17 cm (12-33). Six patients had poor run-off with 0 to 1 patent tibial artery. The mean length of the endarterectomised segment was 31 cm (27-39). An angioscopy and an angiography were performed in all procedures. There were no postoperative complications. All patients had an angiography at 12 months or before because of ipsilateral disease. Primary angiographic patency was defined as patency of the treated artery with stenosis of less than 30%. A short stenosis was defined as <5 cm. These lesions were an indication of percutaneous balloon angioplasty (PTA) and the final results were listed as secondary patency. RESULTS: Angiography revealed 3 patent arteries, 5 short stenosis, 2 long stenosis and no occlusions. The primary patency rate was 30% at 12 months. The secondary patency was 80% with a mean follow-up of 19 months (ranging from 13 to 25 months). The mortality and amputation rate at 12 months was nil. CONCLUSIONS: Despite a modern intraoperative control, there was a high incidence of restenosis after semiclosed endarterectomy performed for long occlusive lesions of the SFA. As a result of a close surveillance and PTA, the secondary patency at one year was good. Before a widespread use of newly developed endovascular techniques, comparative patency studies with the mere endarterectomy should be performed. Endarterectomy followed by a PTA in cases of restenosis, is an alternative to bypass when the vein is not available.
Aged↗
[Intravascular ultrasound: limitation and prospects].
The aims of intravascular ultrasound (IVUS) are visualization of the arterial wall, characterization of the arterial plaque, measurement of vessel diameter and endoluminal procedure assessment. The main sources of errors are: limitation of resolution, distortion of images due to non uniform angular velocity of mechanical type transducers, overestimation or underestimation of cross sectional area secondary to eccentric position or non coaxial orientation of the catheter. Awareness of these problems may prevent misinterpretations. Combined balloon imaging catheters, forward-looking sector scanners and ultrasound imaging guide wires may be achieve by miniaturization of transducers. Three dimensional imaging, blood flow quantification, local elasticity of tissues may be achieve by new softwares.
Arteries↗
Distinct effects of Rac1 on differentiation of primary avian myoblasts.
Rho family GTPases have been implicated in the regulation of the actin cytoskeleton in response to extracellular cues and in the transduction of signals from the membrane to the nucleus. Their role in development and cell differentiation, however, is little understood. Here we show that the transient expression of constitutively active Rac1 and Cdc42 in unestablished avian myoblasts is sufficient to cause inhibition of myogenin expression and block of the transition to the myocyte compartment, whereas activated RhoA affects myogenic differentiation only marginally. Activation of c-Jun N-terminal kinase (JNK) appears not to be essential for block of differentiation because, although Rac1 and Cdc42 GTPases modestly activate JNK in quail myoblasts, a Rac1 mutant defective for JNK activation can still inhibit myogenic differentiation. Stable expression of active Rac1, attained by infection with a recombinant retrovirus, is permissive for terminal differentiation, but the resulting myotubes accumulate severely reduced levels of muscle-specific proteins. This inhibition is the consequence of posttranscriptional events and suggests the presence of a novel level of regulation of myogenesis. We also show that myotubes expressing constitutively active Rac1 fail to assemble ordered sarcomeres. Conversely, a dominant-negative Rac1 variant accelerates sarcomere maturation and inhibits v-Src-induced selective disassembly of I-Z-I complexes. Collectively, our findings provide a role for Rac1 during skeletal muscle differentiation and strongly suggest that Rac1 is required downstream of v-Src in the signaling pathways responsible for the dismantling of tissue-specific supramolecular structures.
Animals↗
Percutaneous stenting of the internal carotid artery: the European CAST I Study. Carotid Artery Stent Trial.
PURPOSE: To report the results of a multicenter safety trial of percutaneous carotid stenting performed by vascular surgeons. METHODS: Symptomatic or asymptomatic patients > or = 65 years of age with internal carotid artery (ICA) stenoses > or = 70% and < or = 2-cm long were eligible for enrollment. The procedures were performed in an operating room with the choice of anesthesia and the percutaneous access site at the discretion of the surgeon. Only Palmaz stents were used. RESULTS: From January 1, 1996 to December 31, 1997, 99 patients (74 men, mean age 70 years, range 51 to 94) were enrolled in the study. More than half (57 of 99 patients) were asymptomatic. The direct cervical approach was used predominantly (97%). Three (3%) cases were converted to surgery for inability to access the artery or deploy the stent (technical success 97%). No perioperative death or myocardial infarction was reported. Six (6%) procedural complications included 1 reversible arterial spasm, 2 dissections, 1 cervical hematoma, and 2 residual stenoses. One neurological event reversed within 7 days (1% minor stroke rate) and 4 (4%) transient ischemic attacks resolved within 24 hours. One (1%) asymptomatic early occlusion occurred 2 days postoperatively. No neurological event was observed in the 1- to 24-month follow-up (mean 13 months). Two (2%) patients died of nonprocedurally related causes. No stent compression was seen, but 1 asymptomatic occlusion and 3 asymptomatic, non-flow-limiting restenoses (2 < 40%, 1 at 60%) were found within 1 year (3% restenosis rate on an intention-to-treat basis). Patency was 98% at 1 year. CONCLUSIONS: The results of this trial support the contention that carotid stenting of short ICA lesions can be performed with a low neurological complication rate.
Aged↗
Endovascular management of axillary artery trauma.
A 17-year-old man was seen with an expanding false aneurysm of the right axillary artery. This was treated by an intraluminal covered-stent introduced through the brachial artery via an 11F sheath. The covered-stent was constructed from a segment of great saphenous vein anchored in the axillary artery by a 29 mm Palmaz stent. Postoperative arteriography and duplex scanning confirmed normal flow through the axillary artery with complete exclusion of the aneurysm. Postoperative recovery was uneventful.
Accidents, Traffic↗
Eps8, a tyrosine kinase substrate, is recruited to the cell cortex and dynamic F-actin upon cytoskeleton remodeling.
Eps8 is a recently identified substrate of receptor and nonreceptor tyrosine kinases implicated in the control of cell proliferation. To investigate potential functions of Eps8, its intracellular localization has been examined in several cell types. In cycling fibroblasts immunolabeling with antibodies to Eps8 reveals a punctate pattern within the perinuclear region and staining of motile peripheral cell extensions and cell-cell contact regions. Stimulation of quiescent Swiss 3T3 fibroblasts with serum induces a striking reorganization of the actin cytoskeleton which is accompanied by the enrichment of Eps8 and cortactin in membrane ruffles and lamellipodia. A similar accumulation of Eps8 to membrane ruffles is observed in cells treated with phorbol esters, which also induce marked changes of the F-actin cytoskeleton. The localization of Eps8 at the cell cortex is largely independent from the binding of Eps8 to an EGFR/ErbB-2 chimeric receptor. Moreover, fractionation studies reveal that a portion of the Eps8 molecules present in the cell periphery, unlike cortactin and the receptor, is resistant to mild extraction with detergent. Upon cellular transformation by the tyrosine kinase v-Src, a pool of Eps8 is recruited to newly formed specialized regions of the cytoskeleton, such as actin bodies in terminally differentiated myotubes and podosomes in fibroblasts, where cortactin and a variety of cytoskeletal proteins are also found. Extraction with Triton X-100 preserves the association of Eps8 to podosomes and leaves the majority of the v-Src tyrosine-phosphorylated Eps8 in the detergent-resistant fraction. The observed recruitment of Eps8 to highly dynamic cytoskeletal structures of normal and transformed cells suggests that Eps8 may play a role in the reorganization of the cytoskeleton, perhaps acting as a docking site for other signaling molecules.
3T3 Cells↗
Increased amyloidogenic secretion in cerebellar granule cells undergoing apoptosis.
Some clues suggest that neuronal damage induces a secondary change of amyloid beta protein (Abeta) metabolism. We investigated this possibility by analyzing the secretion of Abeta and processing of its precursor protein (amyloid precursor protein, APP) in an in vitro model of neuronal apoptosis. Primary cultures of rat cerebellar granule neurons were metabolically labeled with [35S]methionine. Apoptosis was induced by shifting extracellular KCl concentration from 25 mM to 5 mM for 6 h. Control and apoptotic neurons were then subjected to depolarization-stimulated secretion. Constitutive and stimulated secretion media and cell lysates were immunoprecipitated with antibodies recognizing regions of Abeta, full-length APP, alpha- and beta-APP secreted forms. Immunoprecipitated proteins were separated by SDS/PAGE and quantitated with a PhosphorImager densitometer. Although intracellular full-length APP was not significantly changed after apoptosis, the monomeric and oligomeric forms of 4-kDa Abeta were 3-fold higher in depolarization-stimulated secretion compared with control neurons. Such increments were paralleled by a corresponding increase of the beta-APPs/alpha-APPs ratio in apoptotic secretion. Immunofluorescence studies performed with an antibody recognizing an epitope located in the Abeta sequence showed that the Abeta signal observed in the cytoplasm and in the Golgi apparatus of control neurons is uniformly redistributed in the condensed cytoplasm of apoptotic cells. These studies indicate that neuronal apoptosis is associated with a significant increase of metabolic products derived from beta-secretase cleavage and suggest that an overproduction of Abeta may be the consequence of neuronal damage from various causes.
Amyloid beta-Peptides↗
Characterization of the ryanodine receptor/channel of invertebrate muscle.
Electron-microscopic analysis was used to show that invertebrate muscle has feetlike structures on the sarcoplasmic reticulum (SR) displaying the typical four-subunit appearance of the calcium (Ca2+) release channel/ryanodine receptor (RyR) observed in vertebrate skeletal muscle (K. E. Loesser, L. Castellani, and C. Franzini-Armstrong. J. Muscle Res. Cell Motil. 13: 161-173, 1992). SR vesicles from invertebrate muscle exhibited specific ryanodine binding and single channel currents that were activated by Ca2+, caffeine, and ATP and inhibited by ruthenium red. The single channel conductance of this invertebrate RyR was lower than that of the vertebrate RyR (49 and 102 pS, respectively). Activation of lobster and scallop SR Ca2+ release channel, in response to cytoplasmic Ca2+ (1 nM-10 mM), reflected a bell-shaped curve, as is found with the mammalian RyR. In contrast to a previous report (J.-H. Seok, L. Xu, N. R. Kramarcy, R. Sealock, and G. Meissner, J. Biol. Chem. 267: 15893-15901, 1992), our results show that regulation of the invertebrate and vertebrate RyRs is quite similar and suggest remarkably similar paths in these diverse organisms.
Adenosine Triphosphate↗
Regulation of the tyrosine kinase substrate Eps8 expression by growth factors, v-Src and terminal differentiation.
SH3-containing proteins are involved in signal transduction by a number of growth factor receptors and in the organization of the cytoskeleton. The recently identified Eps8 protein, which contains an SH3 domain, is coupled functionally and physically to the EGFR and is tyrosine phosphorylated by this receptor and other receptors as well. Here, we examined the regulation of eps8 expression in response to mitogenic or differentiative signals. We show that Eps8 is expressed at low levels in resting fibroblasts, but its expression is strongly induced during activation by serum, phorbol esters and the v-src oncogene. Conversely, expression of Eps8, but not of other EGFR substrates such as Shc or Eps15, is virtually extinguished in non-proliferating, terminally differentiated murine myogenic cells. The putative role of Eps8 protein as a v-Src substrate was analysed in murine fibroblasts and in quail myogenic cells expressing a temperature-sensitive variant of the tyrosine kinase. Tyrosine phosphorylation of Eps8 was detected only at the permissive temperature. A non-myristylated, transformation-defective mutant of v-Src did not phosphorylate Eps8, whereas it phosphorylated Shc. Together, these findings indicate that Eps8 may be a critical substrate of v-Src. They further establish Eps8 as an example of a signal transducer whose expression senses the balance between growth and differentiation and might, therefore, be involved in the determination of the phenotype.
3T3 Cells↗