Biomedical subjects

G Vecchio

Publications and source records attributed to G Vecchio.

At least 91 records · Page 5Linked to original sources

PubMed1993

The TRK and RET tyrosine kinase oncogenes cooperate with ras in the neoplastic transformation of a rat thyroid epithelial cell line.

We have recently reported that about 50% of papillary thyroid carcinomas harbor an activated TRK or RET oncogene. Two retroviral vectors containing the activated TRK or RET/PTC oncogene have been used to infect a differentiated rat thyroid epithelial cell line, namely the PC Clone 3 cell line. Upon infection with the TRK virus, the PC Clone 3 cells lost only the ability to trap iodide and to express the thyroperoxidase gene. Conversely, when infected with the PTC virus, the PC Clone 3 cells completely lost all of their differentiated functions. However, both the PC-TRK and PC-PTC cell lines were unable to grow in soft agar, and they were not tumorigenic when injected into nude mice. A completely undifferentiated and malignant phenotype was obtained by the cooperation between the TRK or RET and the viral Ha-ras or Ki-ras oncogenes.

Animals↗

PubMed1992

In vivo and in vitro growth-inhibitory effect of bovine seminal ribonuclease on a system of rat thyroid epithelial transformed cells and tumors.

We investigated the antitumoral effect of bovine seminal RNase (BS-RNase) in vivo and in vitro on a model system of epithelial tumor- and metastasis-derived cells as well as on epithelial tumors derived from the same system. We found that while BS-RNase significantly inhibited the growth in vitro of the epithelial tumor-derived cells, its inhibitory effect was even more dramatic on the growth of metastasis-derived cells. BS-RNase exerted no appreciable growth inhibition on normal thyroid epithelial cells. When administered in vivo to rats bearing solid carcinomas, having the same thyroid origin, BS-RNase induced a drastic reduction in the tumor weight, with no detectable toxic effects on the treated animals. These data show, for the first time on a system of neoplastically transformed epithelial cells, that BS-RNase has a potent specific antitumoral activity.

Animals↗

PubMed1992

Conformation for a beta-cyclodextrin monosubstituted with a cyclic dipeptide.

The structural characterization of a beta-cyclodextrin monosubstituted with the peptide cyclo(L-His-L-Leu) is reported. This work provides an x-ray example of a covalently bound group that folds in such a way that the terminal apolar side chain is retained in the hydrophobic interior of the cone-shaped cyclodextrin cavity. 6-Deoxy-6-cyclo(L-histidyl-L-leucyl)-beta-cyclodextrin crystallizes in the space group P1 with cell dimensions a = 14.728(8) A, b = 15.084(7) A, c = 18.182(10) A, alpha = 94.36(6) degrees, beta = 95.81(5) degrees, gamma = 116.08(9) degrees; overall isotropic agreement R = 10.6% for 5703 observed reflections (Fo greater than 3 sigma). The molecular structure consists of two independent molecules with the formula C54H86N4O36.7.25H2O. Each molecule assumes a "sleeping swan"-like overall shape with the hydrophobic leucine side chain inserted inside the cavity of the macrocycle. The two independent units give rise to a head-to-tail dimer linked by hydrogen bonds occurring between primary and secondary hydroxyl groups of the two monomers. The packing of the dimers produces cavities containing water molecules. There are infinite hydrophilic channels running in the crystal, which is similar to what is found in the structures of cyclic peptides.

Carbohydrate Conformation↗

PubMed1992

Evidence that a salt bridge in the light chain contributes to the physical stability difference between heavy and light human ferritins.

Human ferritin, a multimeric iron storage protein, is composed by various proportions of two subunit types: the H- and L-chains. The biological functions of these two genic products have not been clarified, although differences in reactivity with iron have been shown. Starting from the hypothesis that the high stability typical of ferritin is an important property which may be relevant for its iron storage function, we studied ferritin homopolymers of H- and L-chains in different denaturing conditions. In addition we analyzed 13 H-chain variants with alterations in regions conserved within mammalian H-chains. In all the denaturation experiments H-chain ferritin showed lower stability than L-chain ferritin. The difference was greater in guanidine HCl denaturation experiments, where the end products are fully unfolded peptides, than in acidic denaturation experiments, where the end products are peptides with properties analogous to "molten globule." The study on H-chain variants showed: (i) ferritin stability was not affected by alterations of regions exposed to the inner or outer surface of the shell and not involved in intra- or inter-chain interactions; (ii) stability was reduced by alterations of sequences involved in inter-subunit interactions such as the deletion of the N-terminal extension or substitutions along the hydrophobic and hydrophilic channels; (iii) stability was increased by the substitution of 2 amino acids inside the four-helix bundle with those of the homologous L-chain. One of the residues is involved in a salt bridge in the L-chain, and we concluded that the stability difference between H- and L-ferritins is to a large extent due to the stabilizing effect of this salt bridge on the L-subunit fold.

Amino Acid Sequence↗

PubMed1992

The legumin precursor from white lupin seed. Identity of the subunits, assembly and proteolysis.

The precursors of the legumin-like storage protein from developing white lupin seeds (35 days after flowering) are trimers composed of protomers of M(r) 72,000 or 67,000. Some subunits of these oligomers contain processed precursor polypeptides, namely alpha polypeptides of either 52,000 or 44,000 linked through disulphide bonds to a beta polypeptide of 21,000, typical of the mature legumin. The prolegumin is glycosylated. Legumin oligomers purified from the same seeds are both trimers and hexamers; some of their subunits are still made of precursor polypeptides. The hexamer contains less precursor polypeptide than the trimer. A low level or absence of precursor appears to be a condition of hexamer assembly. The heterogenous prolegumin and legumin oligomers represent intermediates in the processing of the prolegumin to mature legumin. Hydrophobic-interaction chromatography on TSK-phenyl-5PW and titration with the hydrophobic probe 8-anilino-1-naphthalenesulphonate indicate that the legumin is less hydrophobic than the prolegumin. This is attributed to structural rearrangements at processing of the propolypeptide, made evident by the behaviour in CD and by the second-derivative ultraviolet spectra of the two proteins. The total protein extract of developing cotyledons at 40 days after flowering contains endopeptidases, similar to those existing in the resting seeds, which cause a limited cascade degradation of the prolegumin and legumin.

Blotting, Western↗

PubMed1992

[Inter-electrode distance and atrial sensing. Our experience with atrial-guided pacing (VDD) in the biennium 1990-1991].

We implanted 47 Phymos VDD pacemakers (53.7% for complete atrioventricular block-42.57% for symptomatic lower heart block) in the 1990-91 period. In this series of cases with EDE 830-830S leads by means Holter technique we evaluated the atrial trigger. A high percentage of atrial sensing (97.8%) was found with 11 cm leads. The right atrial motion study is realized to explain the atrial sensing with floating leads.

Aged↗

PubMed1991

Preferential expression of the dbl protooncogene in some tumors of neuroectodermal origin.

We have investigated the expression of the dbl protooncogene in a wide variety of human tumors of different embryological derivation. We found that proto-dbl mRNA could be detected preferentially in a few neoplastic histiotypes of neuroectodermal origin. The common, normal 5-kilobase size of the proto-dbl transcript detected indicated that the proto-dbl in these tumors was not rearranged. These data, in agreement with our previous reports, suggest that the dbl protooncogene is expressed in a highly tissue-specific manner and indicate that this gene may be involved in the growth and differentiation of some cells of neuroectodermal origin.

Animals↗

PubMed1991

Circular dichroism study on the conformational stability of the dimerization domain of transcription factor LFB1.

LFB1, a dimeric DNA binding protein, is a major determinant of hepatocyte-specific transcription. The thermal and chemical equilibrium unfolding of a 32-residue alpha-helical peptide comprising its dimerization domain (B1-Dim) was monitored by circular dichroism spectroscopy. The conformational stability of this peptide is shown to be concentration dependent, and the unfolding reaction is described as a two-state transition between folded dimers and unfolded monomers. The thermodynamic parameters associated with the unfolding reaction were determined under the two-state assumption by the van't Hoff procedure. The enthalpy of unfolding increases linearly with temperature, and the corresponding value of delta Cp, the difference in heat capacity between the unfolded and the folded forms of the peptide, is estimated to be ca. 0.7 kcal mol-1 K-1. The dimeric folded structure of the peptide is stabilized, at 25 degrees C, by a delta G of about 11.5 kcal mol-1, which is equivalent to a dimerization constant greater than 10(8) mol-1. These results indicate that the dimerization domain of LFB1 can fold and dimerize independently of the rest of the protein, with a thermodynamic stability comparable to that of a small globular protein.

Amino Acid Sequence↗

PubMed1991

Constitutive expression of transforming growth factor alpha does not transform rat thyroid epithelial cells.

To evaluate the role of transforming growth factor alpha (TGF alpha) in transformed rat thyroid epithelial cells, expression and production of TGF alpha were measured in a normal rat thyroid epithelial cell line, FRTL-5, and in the same cells transformed by the Ki-ras oncogene. FRTL-5 cells transformed with Ki-ras exhibit a 5- to 7-fold increase in the levels of TGF alpha-specific mRNA and a 3- to 4-fold enhancement of the amounts of TGF alpha protein in the conditioned medium, as compared with the normal thyroid cells. Although conditioned medium from the Ki-ras transformed FRTL-5 cells or authentic epidermal growth factor or TGF alpha are able to stimulate the anchorage-dependent growth of nontransformed FRTL-5 cells, neither conditioned medium nor the growth factors are able to induce the anchorage-independent growth of these cells in soft agar. FRTL-5 cells were then transfected with an expression vector plasmid containing the human TGF alpha cDNA to directly ascertain if over-expression of this growth factor is able to induce transformation in these cells. The TGF alpha-transfected FRTL-5 clones constitutively produced high amounts of TGF alpha at a level equivalent to or greater than the level found in the conditioned medium from the Ki-ras transformed FRTL-5 clones. Moreover, in contrast to the Ki-ras transformed cells, the TGF alpha transfectants were unable to grow in soft agar, did not form tumors in nude mice, and showed no reduction in the secretion of thyroglobulin. These data demonstrate that unlike Ki-ras, the constitutive expression of biologically active TGF alpha is not entirely sufficient to elicit a transformed phenotype in these cells.

Animals↗

PubMed1991

Refined localization to contiguous regions on chromosome 10q of the two genes (H4 and RET) that form the oncogenic sequence PTC.

In this report we confirm the localization of the human RET proto-oncogene to chromosome 10q11.2, both by Southern blot analysis of a panel of human-rodent somatic cell hybrids and by in situ hybridization on human metaphase chromosomes. Previously, we had assigned to the same chromosome region the gene termed H4. In about 25% of papillary thyroid carcinomas, this gene was shown to rearrange with RET to give rise to the transforming sequence PTC. The analysis of different cell hybrids containing subfragments of chromosome 10, in conjunction with pulse field gel electrophoresis, established that H4 is mapped distally to RET at a distance not less than 280 kb. These findings suggest that intrachromosomal rearrangements are responsible for PTC activation in papillary thyroid carcinomas.

Carcinoma, Papillary↗

PubMed1990

Thyrotropin receptor gene expression in oncogene-transfected rat thyroid cells: correlation between transformation, loss of thyrotropin-dependent growth, and loss of thyrotropin receptor gene expression.

Rat FRTL-5 and PC-Cl-3 thyroid cells are continuously cultured, clonal lines which require thyrotropin to grow and function. Both can be efficiently transformed when infected with RNA or DNA viruses carrying oncogenes or when directly transfected with activated oncogenes. Transformation, assayed by the appearance of cell growth in agar and by tumorigenicity in syngeneic rats or nude mice, is associated with the loss of thyrotropin-dependent cell division and thyrotropin-regulated functions such as thyroglobulin synthesis. In 16 clones of FRTL-5 or PC-Cl-3 cells transformed with different oncogenes, we show that loss of thyrotropin-dependent growth and function correlates with the loss of thyrotropin receptor gene expression, measured with a rat thyrotropin receptor cDNA probe.

Animals↗

PubMed1990

PTC is a novel rearranged form of the ret proto-oncogene and is frequently detected in vivo in human thyroid papillary carcinomas.

We recently detected a novel activated oncogene by transfection analysis on NIH 3T3 cells in five out of 20 primary human thyroid papillary carcinomas and in the available lymph node metastases. We designated this transforming gene PTC (for papillary thyroid carcinoma). Here we describe the molecular cloning and sequencing of the gene. The new oncogene resulted from the rearrangement of an unknown amino-terminal sequence to the tyrosine kinase domain of the ret proto-oncogene. This gene rearrangement was detected in all of the transfectants and in all of the original tumor DNAs, but not in normal DNA of the same patients, thus indicating that this genetic lesion occurred in vivo and is specific to somatic tumors. Moreover, the transcript coded for by the fused gene was detected in an additional PTC-positive human papillary carcinoma for which mRNA was available.

Amino Acid Sequence↗

PubMed1990

Opioid peptides in micellar systems: conformational analysis by CD and by one-dimensional and two-dimensional 1H-NMR spectroscopy.

beta-Endorphin has been studied in SDS micelles by one- and two-dimensional nmr spectroscopy (1D and 2D nmr), and to explore the influence of peptide length and composition on the polypeptide structure, the investigation was extended to a number of fragments. The nmr results are compared with those obtained from CD experiments and discussed in terms of a secondary structure that involves the central region of beta-endorphin.

Amino Acid Sequence↗

PubMed1989

CD and small-angle x-ray scattering of silk fibroin in solution.

We investigated the structure of silk fibroin dissolved in water and in water-organic solvent mixtures by CD and small-angle x-ray scattering (SAXS). CD spectra indicated a disordered secondary structure in water and a beta-sheet conformation in aqueous organic solvents, such as methanol, dioxane, and trifluoroethanol (in trifluoroethanol a transient form evolving toward beta-sheet conformation was seen just after dissolution). The SAXS technique indicated the presence of fibroin particles of lamellar shape. The molecular weight was 188,000 daltons in water and 302,000 daltons in aqueous methanol.

Animals↗

PubMed1989

Denaturation by urea and renaturation of 20 beta-hydroxysteroid dehydrogenase studied by high-performance size exclusion chromatography.

The denaturation by urea and renaturation of 20 beta-hydroxysteroid dehydrogenase, a tetrameric enzyme consisting of four identical subunits, were followed by high-performance size exclusion chromatography to detect intermediates in the processes. During the denaturation process no intermediate form (structured monomers or dimers) between the tetramer and the denatured monomer was observed. During the renaturation process, carried out either with or without NADH, high molecular weight aggregates, native tetramers, and low molecular weight intermediates were evidenced and quantified. The contemporaneous measurement of recovery of activity unambiguously demonstrated that the tetrameric structure is essential for enzymatic activity.

20-Hydroxysteroid Dehydrogenases↗

PubMed1989

The Kirsten murine sarcoma virus induces rat thyroid carcinomas in vivo.

The injection of a retrovirus carrying the v-ras-Ki oncogene into the thyroid gland of adult Fischer rats induces thyroid carcinomas when associated with a treatment of the animals with a goitrogenic agent. More than one hundred adult Fischer rats have been treated with the goitrogen agent propylthiouracil in order to induce thyroid hyperplasia. Twenty days after treatment, rat thyroid glands, surgically prepared, were injected with the Kirsten murine sarcoma virus (KiMSV). Within three months more than 90% of the animals developed thyroid tumors. Histologically the tumors had the appearance of well differentiated carcinomas. Thirty animals had lung metastases in addition to the thyroid carcinoma. The presence of KiMSV specific transcripts and the specific transforming protein (p21) in thyroid carcinomas and in the metastases was detected by Northern blot analysis and immunoprecipitation, respectively. Only three rats, among thirty that had not received the goitrogen treatment, but only the injection with KiMSV, developed thyroid carcinomas of very small size and with a very long latency period (almost one year). The results described represent the first instance of thyroid carcinoma induction by retroviruses. This system may be regarded as a useful model to investigate the process of thyroid carcinogenesis in vivo. These results suggest that this model may also be useful for investigating the interaction between hormones and cells harboring the activated oncogene in the development of thyroid carcinoma since activated ras oncogenes have been implicated in human thyroid carcinoma.

Animals↗

PubMed1989

The oncogene associated with human papillary thyroid carcinoma (PTC) is assigned to chromosome 10 q11-q12 in the same region as multiple endocrine neoplasia type 2A (MEN2A).

In this report we assigned to chromosome 10q the human oncogene PTC frequently associated with the papillary type of thyroid carcinoma. Using an informative panel of human-mouse somatic cell hybrids and 'in situ' hybridization to human metaphase chromosomes, we localized the PTC gene at bands q11-q12 of chromosome 10. These bands belong to one of the two regions on chromosome 10 linked to the cancer syndrome multiple endocrine neoplasia type 2A (MEN2A). Therefore, it is suggested that genes clustered in certain regions of chromosome 10 could be involved in the developmental regulation of the thyroid gland.

Carcinoma, Papillary↗

PubMed1989

Expression of the dbl proto-oncogene in Ewing's sarcomas.

We have investigated the expression of the dbl proto-oncogene in childhood tumors of known or suspected neuroectodermal origin. We found that while the dbl gene is consistently found expressed in Ewing's sarcoma as a single mRNA species, of approximately 5.0 kb, it is generally absent in two seemingly related categories of tumors, neuroblastoma and neuroepithelioma. The specificity of expression of the dbl proto-oncogene in Ewing's sarcoma supports the concept that Ewing's sarcoma may be differentiated from two closely related tumors, neuroblastoma and neuroepithelioma, on the basis of the presence of specific molecular markers.

Guanine Nucleotide Exchange Factors↗