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Biomedical subjects

G Vecchio

Publications and source records attributed to G Vecchio.

At least 37 records · Page 2Linked to original sources
PubMed2000

[Duodenal and pancreatic injuries].

BACKGROUND: Pancreatic and duodenal injuries occur rather infrequently and the incidence ranges between 1% and 12% of all abdominal injuries. The high rate of mortality and morbidity (10-40%) depends on associated complication rate of all intra-abdominal organs (90%). METHODS: Twenty-five cases of pancreatic and duodenal injuries observed between 1987 and 1997, with an incidence of 0.7% of all abdominal injuries, are reported. In 16 cases the cause was penetrating injury (gunshot) and in 9 cases it was blunt abdominal trauma. Only two patients presented an isolated pancreatic lesion, all the others had at least an associated lesion. In all the cases the patients were male and they were submitted to emergency laparotomy. RESULTS: The mortality rate was 20%, the morbidity was 24%. CONCLUSIONS: The relatively low incidence of these injuries and the high rate of associated lesions cause a difficult diagnostic and therapeutic approach, the absence of a unified method to follow and the unsatisfactory results observed.

Adolescent↗
PubMed1999

Fourier-transform infrared spectroscopy study of dehydrated lipases from candida antarctica B and pseudomonas cepacia

Fourier-transform infrared (FT-IR) spectroscopy was employed to investigate potential lyophilization-induced changes in the secondary structure of lipases from Candida antarctica B and Pseudomonas cepacia. The secondary structure elements were determined by curve fitting of the amide III bands of the two lipases in the lyophilized state in KBr pellets and in solution. It was found that lyophilization decreased the alpha-helix and increased the beta-sheet content. However, FT-IR analysis of crosslinked enzyme crystals of Pseudomonas cepacia lipase also indicated an increase in the beta-sheet content, which appears despite the fact that the enzyme, being in the crystallized state, should possess native conformation. This result partially questions the suitability of FT-IR for analysis of the structure of solid proteins, at least as far as the beta-sheet content is concerned, because it is possible that the method overestimates the beta-sheets by measuring other hydrogen-bonded nonperiodic intermolecular structures. No significant modification was observed when lipase from Pseudomonas cepacia was lyophilized in the presence of methoxypoly(ethylene glycol). Copyright 1999 John Wiley & Sons, Inc.

Journal Article↗
PubMed1999

Spectroscopic investigation of lipase from pseudomonas cepacia solubilized in 1,4-dioxane by non-covalent complexation with methoxypoly(ethylene glycol)

Lipase from Pseudomonas cepacia was made soluble in 1,4-dioxane by lyophilization of the enzyme from aqueous solutions containing methoxypoly(ethylene glycol) (PEG). The solubility of the enzyme-PEG complex depended both on protein concentration and PEG protein ratio. Intrinsic protein fluorescence and far- and near-UV circular dichroism revealed that not only did the enzyme not unfold in the organic solvent, but rather became more compact. This was seen by the slight quenching of fluorescence intensity and by the enhancement of the near-UV circular dichroism negative signals, which are indicative of stronger interactions of tryptophanyl and/or tyrosyl residues among themselves or with other parts of the enzyme molecule. The specific activity of the lipase-PEG complex in the organic solvent was at least 2 orders of magnitude higher than that of the enzyme powder. This can be attributed both to the maintenance of native conformation and to enzyme dissolution in the reaction medium which should minimize possible limitations to enzyme-substrate interactions. &copy 1999 John Wiley & Sons, Inc.,

Journal Article↗
PubMed1999

Ret-mediated mitogenesis requires Src kinase activity.

The proto-oncogene RET encodes a transmembrane growth neurotrophic receptor with tyrosine kinase (TK) activity. RET mutations are associated with several human neoplastic and nonneoplastic diseases, including thyroid papillary carcinoma, multiple endocrine neoplasia type 2 syndromes, and Hirschsprung's disease. Activation of receptor TKs results in the binding and activation of downstream signaling proteins, among which are nonreceptor TKs of the Src family. To test the involvement of c-Src in Ret-mediated signaling, we measured the levels of c-Src activity in NIH3T3 cells coexpressing Ret and the accessory GFR alpha-1 receptor or an epidermal growth factor receptor/Ret chimeric receptor when the cells were stimulated by glial cell line-derived neurotrophic factor or epidermal growth factor, respectively. Ret stimulation resulted in the activation of c-Src. We also measured the levels of Src kinase activity in cell lines expressing isoforms of the Ret receptor activated by different mutations. These cells showed higher Src kinase activity than the normal counterpart. Furthermore, we show that Ret is able to associate with the SH2 domain of Src in a phosphotyrosine-dependent fashion. Microinjection of a kinase inactive mutant of c-Src blocked Ret-mediated mitogenic effect. These experiments demonstrate that activated Ret is able to bind and stimulate c-Src kinase and that Src activation is essential for the mitogenic activity of Ret.

3T3 Cells↗
PubMed1999

Different mutations of the RET gene cause different human tumoral diseases.

The RET gene encodes a tyrosine kinase receptor for neurotrophic molecules. RET is a conceptually valuable example of how different mutations of a single gene may cause different diseases. Gene rearrangements activate the oncogenic potential of RET in human thyroid papillary carcinomas. On the other side, different point mutations activate RET in familial multiple endocrine neoplasia syndromes. Finally, inactivating mutations of RET can be present in Hirschsprung's disease patients. The detailed knowledge of the specific RET mutations responsible for human tumors provides relevant tools for the clinical management of these diseases. Moreover, the recent discovery of the growth factors which in vivo stimulate its signaling may shed new light on the role played by RET in the development and differentiation of the central and peripheral nervous system.

Carcinoma, Papillary↗
PubMed1999

High prevalence of RET/PTC rearrangements in Ukrainian and Belarussian post-Chernobyl thyroid papillary carcinomas: a strong correlation between RET/PTC3 and the solid-follicular variant.

A sharp increase in the incidence of pediatric thyroid papillary cancer was documented after the Chernobyl power plant explosion. An increased prevalence of rearrangements of the RET protooncogene (RET/PTC rearrangements) has been reported in Belarussian post-Chernobyl papillary carcinomas arising between 1990 and 1995. We analyzed 67 post-Chernobyl pediatric papillary carcinomas arising in 1995-1997 for RET/PTC activation: 28 were from Ukraine and 39 were from Belarus. The study, conducted by a combined immunohistochemistry and RT-PCR approach, demonstrated a high frequency (60.7% of the Ukrainian and 51.3% of the Belarussian cases) of RET/PTC activation. A strong correlation was observed between the solid-follicular subtype of papillary carcinoma and the RET/PTC3 isoform: 19 of the 24 RET/PTC-positive solid-follicular carcinomas harbored a RET/PTC3 rearrangement, whereas only 5 had a RET/PTC1 rearrangement. Taken together these results support the concept that RET/PTC activation plays a central role in the pathogenesis of thyroid papillary carcinomas in both Ukraine and Belarus after the Chernobyl accident.

Adolescent↗
PubMed1999

[Thoracic drainage in trauma emergencies].

A group of 191 cases of emergency tube thoracostomy for acute trauma reviewed retrospectively from March 1993 to March 1998 is reported. Of this group 169 were men and 22 were women. Their ages ranged from 16 to 73 years. The causes were as follows: 89 cases (46%) road accident; 33 cases (17%) accidental trauma; 33 cases (17%) someone else violence (assault, gunshot or stab wound); 15 cases (8%) work accident; 11 cases (6%) domestic accident and 5 cases (3%) iatrogenic trauma. In 32 patients a diagnosis of pneumothorax was made (2 tension, 11 for penetrating chest injuries, 19 after blunt trauma). In 2 cases of tension pneumothorax and in 3 cases of open pneumothorax a chest tube (24-28 Fr) in the third space in the mid-clavicular line was introduced. In the other patients it was decided to place a chest tube in the mid-axillary line in the fifth intercostal space to drain pneumothorax. Only in 7 cases suction was necessary. Fifty-four hemothorax (3 bilateral) were treated in 11 cases using thoracentesis, while the remaining cases were treated using the insertion of multiple drainage holes in the intercostal space (fifth in the mid-axillary line directed inferiorly and posteriorly). One hundred and three were the cases of hemopneumothorax: 24 of them received 2 chest tubes, the first (20-26 Fr) apically in the second intercostal space in the mid-clavicular line, the second (32-38 Fr) in the fifth intercostal space in the mid-axillary line. All the other cases were treated using a single thoracostomy. In 14 cases suction was applied. Two cases of chylothorax resolved by a large tube positioned in the chest (fifth intercostal space in the mid-axillary line) with a constant negative pressure were also observed. Duration of tube drainage ranged from 4 and 18 days, with an average of 11 days. Five infections of thoracostomy site and 1 empyema resolved by rethoracotomy were observed. Moreover, there were 3 complications: 2 subcutaneous placements and 1 little laceration of the lung. Thirty-one drained patients were operated: in 5 cases thoracotomy and laparotomy (2 exitus in tabula); only thoracotomy in 8 cases; 19 laparotomy and thoracostomy (1 exitus in tabula).

Accidents, Home↗
PubMed1999

Human N-ras, TRK-T1, and RET/PTC3 oncogenes, driven by a thyroglobulin promoter, differently affect the expression of differentiation markers and the proliferation of thyroid epithelial cells.

The ras family members and the tyrosine kinases RET and TRK are frequently activated in human tumors of the thyroid gland. To ascertain the effects of these oncogenes in cultured thyroid cells we have generated expression vectors containing activated versions of the three genes under the control of the thyroid-specific thyroglobulin gene promoter. Here we show that the expression of the three oncogenes differently affects thyroid differentiation. While the TRK-T1 oncogene interferes with the capability of thyroid cells of trapping iodide and only marginally affects thyroglobulin gene expression, both RET/PTC3 and N-ras(Gln61-Lys) induce a dramatic reduction of thyroglobulin mRNA and alleviate TSH dependency for cellular growth. However, none of the three oncogenes is able to induce the appearance of neoplastic transformation markers, such as growth in semisolid medium and tumorigenicity in athymic mice. This indicates that genetic events additional to TRK, RET, or N-ras activation are required for fully malignant transformation of thyroid cells.

Animals↗
PubMed1998

Electrophoresis of proteins across a transverse sodium dodecyl sulfate gradient.

We describe a new protocol for denaturant gradient gel electrophoresis, namely the migration of proteins across transverse sodium dodecyl sulfate (SDS)-gradients. We show how such gradients may be reproducibly cast, and demonstrate their stability with time once an appropriate SDS reservoir is arranged at the cathode. SDS affects both size and surface charge of the molecules, and influences the secondary and tertiary structure to a variable extent, even in opposite directions. Hence, distinct, sometimes complex, denaturation patterns may be observed for different proteins.

Circular Dichroism↗
PubMed1998

Molecular biology of the MEN2 gene.

Cancer is a genetic disease caused by 'gain of function' mutations of oncogenes and 'loss of function' mutations of tumour suppressors and of genes involved in DNA repair mechanisms. The RET gene encodes a tyrosine kinase receptor for molecules belonging to the glial cell line-derived neurotrophic factor (GDNF) family. RET is a paradigmatic example of how different mutations of a single gene can lead to different neoplastic phenotypes. Indeed, gene rearrangements, often caused by chromosomal inversions, activate the oncogenic potential of RET in a fraction of human thyroid papillary carcinomas. On the other hand, different point mutations activate RET in familial multiple endocrine neoplasia syndromes familial medullary thyroid carcinoma (FMTC), MEN-2A and MEN-2B. Little information is so far available on the biochemical mechanisms by which the potent transforming and mitogenic signals of RET are delivered to the nucleus. However, recent data indicate coupling to the Shc-Ras-MAPK pathway as a necessary step in RET signal transduction.

Carcinoma, Papillary↗
PubMed1998

Glial cell line-derived neurotrophic factor differentially stimulates ret mutants associated with the multiple endocrine neoplasia type 2 syndromes and Hirschsprung's disease.

Ret is a receptor tyrosine kinase involved in several neoplastic and developmental diseases affecting the thyroid gland and tissues of neuroectodermal origin. Different ret mutations are associated with different disease phenotypes. Gain-of-function of ret is caused by gene rearrangements in thyroid papillary carcinomas and by point mutations in multiple endocrine neoplasia (MEN) type 2A syndrome (MEN2A), in familial medullary thyroid carcinoma (FMTC), and in the more severe MEN2B syndrome. Conversely, Hirschsprung's disease (HSCR) is associated with loss of function of ret. Recently, it has been shown that glial cell line-derived neurotrophic factor (GDNF), by binding to the accessory molecule GDNFR-alpha, acts as a functional ligand of Ret and stimulates its tyrosine kinase and biological activity. To ascertain whether the biological effects of ret mutations are modulated by GDNF, we have investigated the responsiveness to GDNF of ret mutants in cell lines coexpressing GDNFR-alpha and MEN2A-, MEN2B-, FMTC-, or HSCR-associated ret mutants. Here, we show that triggering of GDNF affected only ret/MEN2B, i.e. it stimulated ret/MEN2B mitogenic and kinase activities, as well as its ability to phosphorylate Shc, a bona fide Ret substrate. In contrast, ret mutants associated with MEN2A or FMTC (carrying Cys634 or Cys620 mutations) were unresponsive to GDNF. HSCR mutations, by affecting either the extracellular or the intracellular Ret domain, impaired responsiveness to GDNF. These data suggest that the phenotype of human diseases caused by ret mutations can be differentially influenced by GDNF.

3T3 Cells↗
PubMed1998

TPA induces a block of differentiation and increases the susceptibility to neoplastic transformation of a rat thyroid epithelial cell line.

The PC Cl 3 cell line is a well-characterized epithelial cell line of rat thyroid origin. This cell line retains in vitro the typical markers of thyroid differentiation: thyroglobulin (TG) synthesis and secretion, iodide uptake, thyroperoxidase (TPO) expression, and dependency on TSH for growth. Although the differentiated phenotype of thyroid cells has been relatively well described, the molecular mechanisms that regulate both differentiation and neoplastic transformation of thyroid cells still need to be investigated in detail. Protein kinase C (PKC), the target of tetradecanoylphorbol acetate (TPA), regulates growth and differentiation of several cell types. Here we show that treatment of PC Cl 3 cells with TPA induces an acute block of thyroid differentiation. TPA-treated PC Cl 3 cells are unable to trap iodide and the expression levels of thyroglobulin, TSH receptor, and TPO genes are drastically reduced by TPA treatment. This differentiation block is not caused by a reduced expression of one of the master genes of thyroid differentiation, the thyroid transcription factor 1 (TTF-1). TPA-treated PC Cl 3 cells display an increased growth rate indicating that, in addition to the differentiation block, TPA also significantly affects the growth regulation of thyroid cells. Finally, TPA treatment dramatically increases the number of transformation foci induced in PC Cl 3 cells by retroviruses carrying v-Ki-ras, v-Ha-ras, and v-mos oncogenes. These findings support the notion that the PKC pathway can influence proliferation, differentiation, and neoplastic transformation of thyroid cells in culture.

Animals↗
PubMed1998

[A rare case of bilateral spigelian hernia].

A rare case of bilateral spigelian hernia is presented and the anatomy and clinical problems related to this disease are discussed. The role of TC scan in the diagnosis of cases with difficult clinical evaluations is stressed.

Abdominal Muscles↗
PubMed1997

Reconstituted high-density lipoproteins with a disulfide-linked apolipoprotein A-I dimer: evidence for restricted particle size heterogeneity.

The apolipoprotein A-IMilano (apoA-IM) is a molecular variant of apoA-I characterized by the Arg173-->Cys substitution, resulting in the formation of homodimers (A-IM/A-IM) and heterodimers with apoA-II. In order to examine the effects of the introduction of an interchain disulfide bridge on the lipid-binding properties of apoA-I, the present studies compare the kinetics of association of A-IM/A-IM and apoA-I with dimyristoylphosphatidylcholine (DMPC), and the structure and properties of reconstituted HDL containing palmitoyloleoylphosphatidylcholine (POPC) and either A-IM/A-IM or apoA-I. The results show that apoA-I dimerization does not affect the rate of association with DMPC. Apolipoprotein-POPC complexes instead, when analyzed by nondenaturing gradient gel electrophoresis, demonstrate that, differently from apoA-I, A-IM/A-IM forms only two species of rHDL particles despite a wide range of initial lipid to protein ratios. These two rHDL species contain one or two A-IM/A-IM molecules and have a diameter of 7.8 nm and 12.5 nm. Investigations of the A-IM/A-IM structure in these two rHDL, by circular dichroism, fluorescence, and second-derivative UV spectroscopy, suggest that the secondary and tertiary structures of A-IM/A-IM are remarkably similar in both small and large particles. These results suggest that the introduction of an interchain disulfide bridge does not affect the association of apoA-I with lipids but restricts HDL particle size heterogeneity, thus possibly affecting HDL function in lipid metabolism and atherosclerosis protection.

Apolipoprotein A-I↗
PubMed1997

Pro108 is important for folding and stabilization of adrenal ferredoxin, but does not influence the functional properties of the protein.

The truncated mutant Met-adrenodoxin-(4-107)-peptide of bovine adrenal ferredoxin was expressed as apoprotein in Escherichia coli BL21 and could be reconstituted to the holoform by chemical or enzymatic methods. The reconstituted protein had spectroscopic, functional and redox properties similar to the Met-adrenodoxin-(4-108)-peptide of adrenal ferredoxin, into which the cluster was inserted upon expression in the same Escherichia coli strain. Rate of in vitro cluster insertion into the Met-adrenodoxin-(4-107) apoprotein was much lower than for the Met-adrenodoxin-(4-108) apoprotein under identical conditions. Comparative thermodynamic studies with the Met-adrenodoxin-(4-108)-peptide indicated that removal of Pro108 resulted in an extensive decrease of the overall stability of the protein in either oxidation state. The Met-adrenodoxin-(4-107)-peptide showed a higher sensitivity to urea denaturation and had a sensibly lower denaturation temperature, 44.8 degrees C, compared with 51.7 degrees C for mutant Met-adrenodoxin-(4-108). The stability of the reduced state of both mutants is slightly lower than that of the oxidized state indicating that this protein region does not undergo major structural changes upon reduction.

Adrenal Glands↗
PubMed1997

Expression of the RET receptor tyrosine kinase and GDNFR-alpha in normal and leukemic human hematopoietic cells and stromal cells of the bone marrow microenvironment.

The RET proto-oncogene product is a receptor tyrosine kinase representing the signal-transducing molecule of a multisubunit surface receptor complex for the glial cell line-derived neurotrophic factor (GDNF), in which a novel glycosyl-phosphatidylinositol (PI)-linked protein (termed GDNFR-alpha) acts as the ligand-binding component. We have analyzed expression of RET and GDNFR-alpha in purified normal hematolymphopoietic cells, leukemia/lymphoma cell lines, and 154 primary samples from patients with hematopoietic malignancies encompassing different lineages and differentiation stages. Relatively low amounts of RET mRNA were found in early CD34+ hematopoietic progenitors, but RET transcripts appeared to increase after myelomonocytic maturation. No expression of RET was found in peripheral blood and tissue B and T lymphocytes. Analysis of human myelomonocytic cell lines was overall consistent with results obtained on purified normal cells. Accordingly, RET expression was mainly confined to acute myeloid leukemias (AMLs) displaying either monocytic (French-American-British M4 and M5) or intermediate-mature myeloid (M2 and M3) phenotypes, being less frequently detected in early myeloid (M0 and M1) AMLs. In contrast, RET mRNA was sporadically detected in B-cell tumors, whereas, among T-cell malignancies, RET transcripts were mainly detected in cells of postthymic and mature T-cell phenotype. RET broad detection in primary tumors was not paralleled by the mutual expression of GDNFR-alpha, which was detected only in 2 isolated primary samples and in 3 leukemia/lymphoma cell lines. However, GDNFR-alpha transcripts, in the absence of RET mRNA, were found in normal bone marrow stromal cells (BMSC), in BM fibroblasts, and in two osteoblast cell lines previously described to support normal hematopoiesis. In the presence of GDNF-receptors derived from BMSC by PI-specific phospholipase C cleavage, GDNF efficiently bound RET-expressing AML blasts and was functionally active by reducing their clonogenic growth and triggering the monocytic maturation of leukemic cells.

Adipose Tissue↗