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Biomedical subjects

A Greco

Publications and source records attributed to A Greco.

At least 37 records · Page 2Linked to original sources
PubMed2001

Detection of haemoparasites in cattle by reverse line blot hybridisation with a note on the distribution of ticks in Sicily.

A reverse line blot hybridisation (RLB) of 21 oligonucleotides with polymerase chain reaction (PCR) amplified regions of 16S rRNA (Ehrlichia/Anaplasma group) or 18S rRNA (Babesia/Theileria group) genes of haemoparasites detected Theileria annulata, T. buffeli/orientalis, Babesia bovis, B. bigemina, B. divergens, Ehrlichia bovis, Anaplasma marginale, A. centrale and unknown species within the Rickettsia tribe.A very high prevalence of mixed infections was detected, which indicated that animals infected with Babesia spp. were also infected with Theileria spp. and/or Anaplasma spp. The tick distribution appeared to be seasonal with Hyalomma marginatum as the most frequently observed tick and Boophilus annulatus and Ixodes ricinus as the least frequently observed ticks. Other species identified in the 818 ticks collected during the five sampling periods between April 1998 and November 1999 included H. lusitanicum, Rhipicephalus sanguineus group, R. bursa, Dermacentor marginatus, Haemaphysalis punctata, B. annulatus and I. ricinus.

Anaplasma↗
PubMed2001

Deregulated FGFR3 mutants in multiple myeloma cell lines with t(4;14): comparative analysis of Y373C, K650E and the novel G384D mutations.

The t(4;14)(p16.3;q32) chromosomal translocation occurs in approximately 20% of multiple myelomas (MM) and leads to the apparent deregulation of two genes located on 4p16.3: the fibroblast growth factor receptor 3 (FGFR3) and the putative transcription factor WHSC1/MMSET. Interestingly, FGFR3 mutations known to be associated with autosomal dominant human skeletal disorders have also been found in some MM cell lines with t(4;14) but their pathogenetic role in MM is still controversial. Since cell lines may represent useful models for investigating the effects of deregulated FGFR3 mutants in MM, we analysed the expression, activation, signaling pathways and oncogenic potential of three mutants identified so far: the Y373C and K650E in the KMS-11 and OPM-2 cell lines respectively, and the novel G384D mutation here identified in the KMS-18 cell line. All of the cell lines present a heterozygous FGFR3 gene mutation and transcribe the mutated allele; unlike KMS-11 and OPM-2 (which express the IIIc isoform), the KMS-18 cell line expresses prevalently the isoform IIIb. We demonstrated that, under serum-starved conditions, KMS-11 and OPM-2 cells express appreciable levels of phosphorylated FGFR3 mutants indicating a constitutive activation of the Y373C and K650E receptors; the addition of the aFGF ligand further increased the level of receptor phosphorylation. Conversely, the FGFR3 mutant in KMS-18 does not seem to be constitutively activated since it was phosphorylated only in the presence of the ligand. In all three MM cell lines, ligand-stimulated FGFR3 mutants activated the MAP kinase signaling pathway but did not apparently involve either the STAT1 or STAT3 cascades. However, when transfected in 293T cells, G384D, like Y373C and K650E, was capable of activating MAPK, STAT1 and STAT3 under serum-starved condition. Finally, a focus formation assay of NIH3T3 cells transfected with FGFR3-expressing plasmid vectors showed that Y373C and K650E (albeit at different levels) but not G384D or the wild-type receptor, can induce transformed foci. Overall, our results support the idea that FGFR3 mutations are graded in terms of their activation capability, thus suggesting that they may play a critical role in the tumor progression of MM patients with t(4;14).

3T3 Cells↗
PubMed2001

Growth-inhibitory effect of STI571 on cells transformed by the COL1A1/PDGFB rearrangement.

Dermatofibrosarcoma protuberans (DP) is a skin tumor of intermediate malignancy characterized by high recurrence rates, for which surgical excision is the main therapy. All DP cases carry a specific t(17;22) translocation, resulting in a COL1A1/PDGFB rearrangement. The subsequently deregulated production of PDGFB generates autocrine stimulation of PDGFrbeta, leading to malignant transformation. Using NIH-3T3 cells transformed by the COL1A1/PDGFB rearrangement (5A cell line), we explored the possibility of blocking the PDGFB autocrine loop, both in vitro and in vivo, using STI571, an inhibitor of the PDGF receptor and of ABL kinase activity. The presence of small amounts of serum in the culture medium was required for the in vitro growth and morphological transformation of 5A cells. In the presence of STI571, the growth rate was reduced and the associated transformed phenotype changed to a flattened one. This effect could be reversed on removal of the inhibitor. The growth rate of tumors induced by 5A cells in nude mice was reduced by STI571 administration. Interestingly, this effect was also evident on pre-existing tumors, but no tumor eradication was observed. This is consistent with the reversible effects of the inhibitor observed in vitro but differs from the eradication effect of STI571 on BCR-ABL-induced tumors. Our data indicate that STI571 might be a candidate compound for the pharmacological treatment of DP and demonstrate that the same compound may act in different ways (cytotoxic vs. cytostatic), according to the specificity of the inhibited tyrosine kinase, namely, ABL or PDGFrbeta.

3T3 Cells↗
PubMed2001

IRS-1 and IRS-2 are recruited by TrkA receptor and oncogenic TRK-T1.

TRK-T1 oncogene is generated by the rearrangement of the NGF receptor TrkA with TPR. This gives rise to the constitutive tyrosine autophosphorylation and activation of the kinase. To study TRK-T1 oncogenic signaling and compare it to that induced by the genuine receptor TrkA, we investigated the involvement of IRS-1, a docking protein implicated in mitogenic signaling induced by several growth factors, in TRK-T1 and TrkA signaling. Here, we show that IRS-1 and IRS-2 are phosphorylated on tyrosine in presence of both TRK-T1 and the activated TrkA receptor. These tyrosine phosphorylations lead to IRS-1- and IRS-2-induced recruitment of p85PI3K, SHP-2, and Grb2 and increase in PI 3-kinase activity associated with IRS-1. Furthermore, we found that TRK-T1 is able to activate c-fos serum responsive element in cooperation with IRS-1 and IRS-2. We observed that TRK-T1 stimulates DNA synthesis in wild-type fibroblasts but not in IRS-1(-/-) mouse embryo fibroblasts. Yeast two-hybrid system experiments showed the occurrence of direct interaction between TRK and IRS molecules, which suggests involvement of different modes of interactions. On the whole, our results suggest that IRS-1 and IRS-2 could be substrates of TRK-T1 and TrkA, and hence could participate in their signal generation.

Adaptor Proteins, Signal Transducing↗
PubMed2001

Identification of ribosome-associated viral and cellular basic proteins during the course of infection with herpes simplex virus type 1.

Herpes simplex virus type 1 (HSV-1) infection induces severe alterations of the translational apparatus, including the phosphorylation of a few ribosomal proteins, and the progressive association of several nonribosomal proteins to ribosomes. Therefore, we hypothesized that ribosomes themselves could contribute to the HSV-1-induced translational control of host and viral gene expression. As a prerequisite to test this hypothesis, we undertook the identification of the nonribosomal proteins associated to the ribosomes during the course of HSV-1 infection. After separation by two-dimensional polyacrylamide gel electrophoresis of basic proteins extracted from the ribosomal fraction, the identification of unknown protein spots was carried out by N-terminal sequencing and peptide mass determination by mass spectrometry. This allowed us to identify HSV-1 VP19C and VP26 that associated to ribosomes with different kinetics. Another nonribosomal protein turned out to be the poly(A)-binding protein 1 (PAB1P). Newly synthesized PAB1P continued to associate to ribosomes all along infection.

Gene Expression Regulation, Viral↗
PubMed2001

Phase III, randomized, double-blind, cross-over comparison of gadoteridol and gadopentetate dimeglumine in magnetic resonance imaging of patients with intracranial lesions.

This study compares the efficacy and safety of gadoteridol with that of gadopentetate dimeglumine for enhanced MRI in subjects with intracranial lesions. A total of 92 subjects at three European centres underwent one MRI study enhanced with 0.1 mmol/kg gadoteridol and another with 0.1 mmol/kg gadopentetate dimeglumine. Contrast agents were assigned in random order, separated by 3-7 days. Eighty subjects were evaluated for efficacy. The presence of pathology, degree of enhancement, location and number of lesions, as well as additional information gained, were compared for each subject's unenhanced and enhanced scans for both the gadoteridol and gadopentetate dimeglumine examination. Safety was evaluated in all treated subjects by means of pre- and post-dose vital signs, laboratory tests and by monitoring for adverse events. There was no significant difference in the number of lesions visualized pre- and post-contrast for the two contrast agents. A high degree of correlation was noted between the two blinded readers. When post-contrast image sets were compared between contrast agents, there was no significant difference in superiority of one agent over the other for any of the evaluators (P > 0.05). No significant differences for any safety parameter were noted between the two agents. Gadoteridol and gadopentetate dimeglumine are effective and well tolerated for use in contrast-enhanced MRI of the CNS at a dose of 0.1 mmol/kg.

Adolescent↗
PubMed2001

[Early diastolic sound in a patient with hepatocarcinoma].

On physical examination an early diastolic sound is usually associated with mitral stenosis, prosthetic mitral valve replacement and chronic constrictive pericarditis. In case of an atrial myxoma, an early diastolic sound can be usually heard due to movement of the tumor towards the tricuspid valve (tumor plop). The following case report shows an example in which an early diastolic sound was heard in a patient presenting with a hepatocellular carcinoma. This sound was due to the presence of a thrombus that originated from the inferior vena cava and invaded the right atrium up to the tricuspid valve. It was thus similar to an atrial myxoma and produced a tumor plop.

Carcinoma, Hepatocellular↗
PubMed2001

[Isoprostanes and oxidative stress in brain damage in newborns].

Isoprostanes are a family of biologically active molecules recently characterized, which is emerging as a new class of specific and reliable markers of in vivo and ex vivo lipid peroxidation and oxidative damage. These molecules are stable, relatively abundant and easily detectable by sensitive and specific analytical methods. In the last years, the measurement of their levels in tissue homogenates or biological fluids has significantly improved our knowledge on the involvement of oxidative stress in several neurological diseases. Here we present evidence indicating that isoprostanes can be successfully used also to study the mechanisms involved in free radical brain damage following hypoxic-ischaemic or inflammatory conditions in newborns and preterm infants.

Brain Damage, Chronic↗
PubMed2000

The TRK-T1 fusion protein induces neoplastic transformation of thyroid epithelium.

Genetic analysis of human papillary thyroid carcinomas (PTC) has revealed unique chromosomal translocations that form oncogenic fusion proteins and promote thyroid tumorigenesis in up to 60% of tumors examined. Although, the majority of thyroid specific translocations involve the growth factor receptor c-RET, variant rearrangements of the receptor for nerve growth factor, NTRK1 have also been described. One such translocation, TRK-T1, forms a fusion protein composed of the carboxyl terminal tyrosine kinase domain of NTRK1 and the amino terminal portion of TPR (Translocated Promoter Region). To determine if TRK-T1 expression can cause thyroid cancer in vivo, we developed transgenic mice that express the human TRK-T1 fusion protein in the thyroid. Immunohistochemical analysis of TRK-T1 transgenic mouse thyroids revealed TRK-T1 staining within the thyroid follicular epithelium. In contrast to nontransgenic littermates, 54% of transgenic mice developed thyroid abnormalities that included follicular hyperplasia and papillary carcinoma. Furthermore, all transgenic mice examined greater than 7 months of age developed thyroid hyperplasia and/or carcinoma. These data support the conclusion that TRK-T1 is oncogenic in vivo and contributes to the neoplastic transformation of the thyroid.

Animals↗
PubMed2000

Translation initiation of the insulin-like growth factor I receptor mRNA is mediated by an internal ribosome entry site.

The insulin-like growth factor I receptor (IGF-IR) is a heterotetrameric receptor mediating the effects of insulin-like growth I and other growth factors. This receptor is encoded by an mRNA containing an unusually long, G-C-rich, and highly structured 5' untranslated region. Using bicistronic constructs, we demonstrated here that the 5' untranslated region of the IGF-IR allows translation initiation by internal ribosome entry and therefore constitutes an internal ribosome entry site. In vitro cross-linking revealed that this internal ribosome entry site binds a protein of 57 kDa. Immunoprecipitation of UV cross-linked proteins proved that this protein was the polypyrimidine tract-binding protein, a well known regulator of picornavirus mRNA translation. The efficiency of translation of the endogenous IGF-IR mRNA is not affected by rapamycin, which is a potent inhibitor of cap-dependent translation. This result provides evidence that the endogenous IGF-IR mRNA is translated, at least in part, through a cap-independent mechanism. This is the first report of a growth factor receptor containing sequence elements that allow translation initiation to occur by internal initiation. Because the IGF-IR has a pivotal function in the cell cycle, this mechanism of translation regulation could play a crucial role in the control of cell proliferation and differentiation.

5' Untranslated Regions↗
PubMed2000

The Gly571Arg mutation, associated with the autonomic and sensory disorder congenital insensitivity to pain with anhidrosis, causes the inactivation of the NTRK1/nerve growth factor receptor.

Point mutations affecting the NTRK1/TRKA gene, encoding one of the receptors for the nerve growth factor (NGF), have been detected in congenital insensitivity to pain with anhidrosis (CIPA), a human hereditary sensory neuropathy characterized by absence of reaction to noxious stimuli and anhidrosis. To define the defect of NTRK1 in CIPA patients, we have introduced one of the previously reported mutations (Gly571Arg) into both the NTRK1 and the TRK-T3 oncogene cDNAs. The expression of the mutated constructs into COS1 cells revealed that the introduced mutation, while not affecting its correct membrane localization, rendered the NTRK1 protein unable to undergo activation upon stimulation with NGF. Similarly, the mutation abolished the constitutive activation of the TRK-T3 oncogene. Transfection into NIH3T3 and PC12 cells showed the loss of transforming and differentiating activity by the mutated constructs. Our results demonstrate clearly that the CIPA mutations cause the inactivation of the NTRK1 receptor, thus exerting a loss of function effect, and provide an experimental approach to distinguish functional mutations from genetic polymorphisms.

Amino Acid Substitution↗
PubMed2000

Development of the caudal neurosecretory system of the nile tilapia Oreochromis niloticus: an immunohistochemical and electron microscopic study.

The development of the caudal neurosecretory system (CNSS) of the Nile tilapia, Oreochromis niloticus, has been investigated by means of UI/oCRF (urotensin I/ovine corticotropin-releasing factor) immunohistochemistry and transmission electron microscopy. UI-like immunoreactive perikarya and fibers are first detected in the caudal spinal cord of larval fish about 4 days after hatching (stage 21). In the region of the future urophysis two bundles of strongly immunoreactive neurosecretory fibers are observed. At this stage, neurosecretory axons terminate on the meninx sheath of the spinal cord with immature neurosecretory terminals. The histogenesis of the urophysis begins at stage 24. The future neurohemal organ consists of a small ventral swelling of the spinal cord, which is associated with dilated vessels. At this stage, neurosecretory axons terminate on the basal lamina of the ingrowing blood vessels. Further development occurs by means of progressive branching of vessels and the concomitant increase in the number of neurosecretory terminals. In the caudal spinal cord, immunoreactive neurons also increase in number and progressively differentiate morphologically. Typical features of the mature CNSS are recognizable in 4-month-old juveniles. Data suggest that in tilapia both the synthesis and the release of urophysial hormones begin before morphogenesis of the neurohemal organ takes place.

Aging↗
PubMed2000

Characterization by two-dimensional gel electrophoresis of host proteins whose synthesis is sustained or stimulated during the course of herpes simplex virus type 1 infection.

Herpes simplex virus type 1 (HSV-1) gene expression is concomitant with a selective shutoff of host protein synthesis. While the synthesis of the vast majority of cellular proteins is inhibited immediately after infection, several cellular proteins continue to be synthesized, even during the late phase of infection. Because these cellular proteins may intervene in the life cycle of the virus, we undertook two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) analyses to evaluate the proportion of cellular proteins that is represented by these particular proteins. Human cells were infected with HSV-1. At different times after infection, proteins were labeled with 35S just prior to harvesting. The rate of synthesis of a set of 183 acidic host proteins, as well as that of ribosomal proteins, was measured during the course of infection, after separation by 2-D PAGE. As expected, HSV-1 induces a strong inhibition of host protein synthesis immediately after infection. However, the synthesis of basic ribosomal proteins and that of an unexpected high proportion of the sub-set of cellular proteins analyzed is sustained or stimulated during HSV-1 infection. A 2-D PAGE analysis outlining the expression patterns of these proteins at different times of infection is presented.

Electrophoresis, Gel, Two-Dimensional↗
PubMed2000

Isoprostanes, novel markers of oxidative injury, help understanding the pathogenesis of neurodegenerative diseases.

Isoprostanes are prostaglandin-like compounds which are formed by free radical catalysed peroxidation of arachidonic acid esterified in membrane phospholipids. They are emerging as a new class of sensitive, specific and reliable markers of in vivo lipid peroxidation and oxidative damage. Since their initial description of in 1990, the rapid development of analytical methods for isoprostane measurement has allowed to overcome some of the pitfalls of the previous and most widely used methods of assessing free radical injury. Here, we summarise the current knowledge on these novel class lipid peroxidation products and the advantages of monitoring their formation to better define the involvement of oxidative stress in neurological diseases. Although the literature data are still not abundant, they indicate that in vivo or post mortem cerebrospinal fluid and brain tissue levels of isoprostane are increased in some diseases such as multiple sclerosis, Alzheimer's disease, Huntington's disease, and Creutzfeldt-Jakob disease.

Alzheimer Disease↗
PubMed2000

Increased brain synthesis of prostaglandin E2 and F2-isoprostane in human and experimental transmissible spongiform encephalopathies.

The levels of 2 arachidonic acid metabolites formed either by enzymatic activity of cyclooxygenase, i.e. prostaglandin E2 (PGE2), or by free radical-catalyzed peroxidation, i.e. F2-isoprostane 8-epi-prostaglandin F2alpha (8-epi-PGF2alpha), were measured in the CSF of subjects with sporadic and familial Creutzfeldt-Jakob disease (CJD) and in brain homogenates of scrapie-infected mice. The CSF levels of both metabolites were increased in sporadic CJD (n = 52) and familial CJD (n = 10) patients when compared with a group of patients with noninflammatory disorders. Similarly, PGE2 and 8-epi-PGF2alpha levels were higher in brain homogenates obtained from C57BL/6J mice infected with the ME7 scrapie strain than in brain homogenates from control animals. As PGE2 is 1 of the most abundant prostaglandins released during inflammation and 8-epi-PGF2alpha is a quantitative marker of lipid peroxidation, our results provide in vivo biochemical evidence for the occurrence of inflammation and oxidative stress in human and experimental transmissible spongiform encephalopathies (TSEs), a concept so far based mainly on histopathological and in vitro evidence. Interestingly, in sporadic CJD patients, high CSF levels of PGE2, but not 8-epi-PGF2alpha, correlated with short survival time, suggesting that the inflammatory response correlates with the clinical duration of disease.

Adult↗
PubMed2000

BK virus infection in AIDS.

The BK virus (BKV) belongs to the family of the polyoma group, which contains three species: JC, which is responsible for progressive multifocal leukoencephalopathy in acquired immunodeficiency syndrome (AIDS); simian virus 40 (SV40), which is a simian virus of little pathologic significance in humans; and BKV, which is usually not pathogenic and is found in the urine of asymptomatic individuals. Recently BKV has been reported to cause symptomatic infection in renal transplant patients. The authors report a rare case of a 14-year-old boy with AIDS who developed a BKV infection of the lung and kidney that progressed to diffuse alveolar damage and death. The infected type II pneumocytes in the lung and the tubular epithelial cells in the kidney showed large, homogenous purple intranuclear inclusions. The absence of necrosis and destruction made it possible to distinguish BKV infection from herpes simplex. The size of the infected cells and the lack of a halo around the nuclear inclusion helped rule out cytomegalovirus as the cause of infection. Electron microscopy detected the presence of 40-nm intranuclear viral particles compatible with BKV, and in situ hybridization established the diagnosis.

AIDS-Related Opportunistic Infections↗
PubMed2000

Focal liver lesions: evaluation of the efficacy of gadobenate dimeglumine in MR imaging--a multicenter phase III clinical study.

PURPOSE: To evaluate gadobenate dimeglumine (Gd-BOPTA) for dynamic and delayed magnetic resonance (MR) imaging of focal liver lesions. MATERIALS AND METHODS: In 126 of 214 patients, MR imaging was performed before Gd-BOPTA administration, immediately after bolus administration of a 0.05- mmol/kg dose of Gd-BOPTA, and 60-120 minutes after an additional intravenously infused 0.05-mmol/kg dose. In 88 patients, imaging was performed before and 60-120 minutes after a single, intravenously infused 0.1-mmol/kg dose. T1- and T2-weighted spin-echo and T1-weighted gradient-echo images were acquired. On-site and blinded off-site reviewers prospectively evaluated all images. Intraoperative ultrasonography, computed tomography (CT) during arterial portography, and/or CT with iodized oil served as the reference methods in 110 patients. RESULTS: Significantly more lesions were detected on combined pre- and postcontrast images compared with on precontrast images alone (P <. 01). All reviewers reported a decreased mean size of the smallest detected lesion and improved lesion conspicuity on postcontrast images. All on-site reviewers and two off-site reviewers reported increased overall diagnostic confidence (P <.01). Additional lesion characterization information was provided on up to 109 (59%) of 184 delayed images and for up to 50 (42%) of 118 patients in whom dynamic images were assessed. Gd-BOPTA would have helped change the diagnosis in 99 (47%) of 209 cases and affected patient treatment in 408 (23%) of 209 cases. CONCLUSION: Gd-BOPTA increases liver lesion conspicuity and detectability and aids in the characterization of lesions.

Adult↗