RESUMEN
The FHIT gene, a member of the histidine triad gene family, is a tumor suppressor gene exhibiting deletions in the majority of human cancers. Aberrant transcripts of this gene have been found in about 50% of esophageal, stomach and colon carcinomas. Little is known about the molecular mechanisms involved in malignant transformation of the lining cells of the anus. In this study FHIT gene expression was investigated in this particular kind of human cancer. FHIT expression was comparatively analyzed at the mRNA level, by RT-PCR, in squamous anal cancers, normal anal tissue and peripheral blood samples. cDNA analyses showed variability in FHIT transcripts, without apparent effects on the predicted amino acid sequence. These different FHIT mRNAs could represent transcripts from an alternative splicing event. Our data indicate that the FHIT mRNA detected in anal cancers and in normal samples is heterogeneous. Immunohistochemical data suggest that the Fhit protein is expressed only in a fraction of the tumor cells, while it is strongly expressed in the epithelial cells of glands of the normal anal mucosa. The absence or poor expression of the Fhit protein in anal cancers suggests a role for this tumor suppressor gene product, as a risk factor, in the onset of this human cancer, as reported before for other human gastrointestinal tumors.
Asunto(s)
Ácido Anhídrido Hidrolasas/biosíntesis , Neoplasias del Ano/metabolismo , Proteínas de Neoplasias/biosíntesis , Neoplasias del Ano/genética , Secuencia de Bases , Perfilación de la Expresión Génica , Humanos , Inmunohistoquímica , Datos de Secuencia Molecular , ARN Mensajero/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The aim of this study was to characterize some phenotypic expressions of fibroblasts from the human oral mucosa. Gingival and lower forearm fibroblasts from young (20-30 years) and elderly (> 60 years) subjects were analyzed. Gingival fibroblasts were taken from donors with (P) and without (NP) periodontal disease, while skin biopsies were taken from healthy subjects. Cell proliferation was assessed by evaluating the cell multiplication coefficient (C.M.C.). The proliferation potential of gingival fibroblasts from elderly individuals with and without periodontopathy did not differ from that of young subjects in the same condition but differed significantly in the skin samples. Enzyme neutral endopeptidase (EC 3.4.24.11) (NEP) activity, studied as a possible marker of cell ageing, showed an age-related increase in human skin fibroblasts but not consistently in gingival fibroblasts from individuals with or without periodontal disease. Cell area and substrate adhesion were evaluated by morphometric analysis. There were no significant differences between elderly P and NP subjects, while significant differences were observed between young and elderly P subjects. In conclusion, proliferative capacity and NEP activity in gingival fibroblasts did not appear to be age-related, probably because their microenvironment is continually moistened by saliva, which continues to contain growth factors, notably EGF, even into senescence. Tissue reaction and repair are important clinical and therapeutic implications.
Asunto(s)
Envejecimiento/fisiología , Fibroblastos/fisiología , Encía/fisiología , Periodontitis/patología , Adulto , Anciano , Humanos , Técnicas In VitroRESUMEN
To investigate whether ESE-1 gene abnormalities are involved in alterations of epithelial cell differentiation in squamous anal cancer ESE-1 expression and structure were screened in six patients by reverse transcriptase-polymerase chain reaction (RT-PCR) and automated sequence analysis. The complete cDNA of isoform ESE-1b was always expressed and correctly spliced, with single nucleotide polymorphism being observed in two cases. Presence of ESE-1b point mutations was excluded. Expression of SPRR2A and ENDOA/CK8, two epithelium-specific ESE-1 target genes, were revealed by RT-PCR in all cases. This first report of expression of ESE-1, and of SPRR2A and ENDOA/CK8 (both related to terminal differentiation in different types of epithelia lining) in anal cancer excludes the hypothesis that these genes influenced carcinogenesis in our patients. Despite selecting of patients without clinical evidence of HPV infection, PCR consistently revealed HPV-16 DNA, highlighting the importance of HPV infection in anal cancer.
Asunto(s)
Neoplasias del Ano/genética , Proteínas de Unión al ADN , Neoplasias de Células Escamosas/genética , Proteínas Proto-Oncogénicas , Transactivadores/genética , Factores de Transcripción , Adulto , Anciano , Anciano de 80 o más Años , Quimiocinas CC/genética , Proteínas Ricas en Prolina del Estrato Córneo , Femenino , Humanos , Masculino , Proteínas de la Membrana/genética , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Precursores de Proteínas/genética , Proteínas Proto-Oncogénicas c-etsRESUMEN
Anal cancer originates from a peculiar histological region and provides a useful model for investigating alterations in proliferation and/or differentiation of neoplastic keratinocytes. Epidermal differentiation complex (EDC) genes, which form one of the major gene clusters in the human genome, are involved in the terminal differentiation of epithelial cells and in many instances have been implicated in epithelial tumours. We constructed a DNA macroarray capable of characterising the expression profiles of the entire EDC gene complex in normal mucosa and anal cancer biopsies of seven unrelated patients. Brain tissue and cultured keratinocytes were used as controls. All anal cancer samples showed expression profiles in which none of the EDC genes was silent, as evaluated by phosphor-imager analysis. Variance analysis showed significantly lower expression of SPRR2 with respect to SPRR1 or SPRR3, and significantly higher expression of S100A8 than of other S100A subfamily members. At hierarchical clustering analysis, the four basaloid anal cancer cases conglomerated in the top five positions. The macroarray method used by us provides the first demonstration of the expression profile of the EDC gene family in anal cancer, and is capable of producing significant information on the subgrouping of epithelial tumours such as anal cancer.
Asunto(s)
Neoplasias del Ano/genética , Membrana Mucosa/metabolismo , Proteínas de Neoplasias/genética , Adulto , Anciano , Neoplasias del Ano/metabolismo , Diferenciación Celular , Cartilla de ADN/química , ADN de Neoplasias/análisis , Células Epiteliales/metabolismo , Femenino , Expresión Génica , Perfilación de la Expresión Génica/métodos , Humanos , Procesamiento de Imagen Asistido por Computador , Masculino , Persona de Mediana Edad , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Bone defects were surgically produced in the tibiae of rabbits and medicated with freeze-dried methylpyrrolidinone chitosan. Histological observations 60 d after surgery showed a considerable presence of neoformed bone tissue, as opposed to controls, originating from the pre-existing bone as well as from the periosteum. The cationic nature and the chelating ability of the methylpyrrolidinone chitosan apparently favoured mineralization. Endosteal-periosteal and bone marrow osteoblast-like precursors, stimulated by growth factors entrapped in the coagulum-polysaccharide mixture, gave rise to intramembranous bone formation. The ultrastructural examination evidenced that bone osteoid was followed by mineralization of the tissue.
Asunto(s)
Conducción Ósea/efectos de los fármacos , Quitina/análogos & derivados , Quitosano , Modelos Animales de Enfermedad , Osteoblastos/efectos de los fármacos , Pirrolidinonas/farmacología , Animales , Desarrollo Óseo/efectos de los fármacos , Enfermedades Óseas/tratamiento farmacológico , Regeneración Ósea/efectos de los fármacos , Quitina/farmacología , Femenino , Luz , Microscopía , Microscopía Electrónica , Conejos , Tibia/cirugíaRESUMEN
In patients undergoing plastic surgery, donor sites were treated with soft pads of freeze-dried N-carboxybutyl chitosan to promote ordered tissue regeneration. Compared to control donor sites, better histoarchitectural order, better vascularization and the absence of inflammatory cells were observed at the dermal level, whilst fewer aspects of proliferation of the malpighian layer were reported at the epidermal level. Accordingly, N-carboxybutyl chitosan leads to formation of regularly organized cutaneous tissue and reduces anomalous healing.
Asunto(s)
Apósitos Biológicos , Quitina/análogos & derivados , Quitosano , Trasplante de Piel/métodos , Piel/ultraestructura , Cirugía Plástica/métodos , Cicatrización de Heridas/fisiología , Adulto , Colágeno/ultraestructura , Femenino , Fibroblastos/ultraestructura , Humanos , Masculino , Microscopía Electrónica , Persona de Mediana Edad , Trasplante de Piel/fisiología , MusloRESUMEN
OBJECTIVE: A positive balance in bone remodelling is an important goal of bone metabolism both in the presence of the osteoporotic processes characteristic of ageing and, especially, of prosthetic implants. The aim of the present work was to obtain new information about the initial steps of osteoblastic growth in an in vitro osteoblastic model in the presence of two bisphosphonates. METHODS: Experiments were performed with Alendronate and Neridronate, two molecules used in the therapy of osteoporosis. Since differentiating features into osteoblastic cells are known to parallel the presence in the cytoplasm of alkaline phosphatase and osteocalcin, we also carried out immunohistochemical typing. RESULTS: Good differentiation and osteoblastic activity were generally observed in the cells in contact with these compounds, except for 10(-4) Neridronate, where biochemical data clearly indicated its toxic effect on the cells. CONCLUSION: The detection of osteoblastic markers associated with an ultrastructural picture of correct organellar morphology in our cultures further supports the hypothesis of a metabolically positive action of these molecules on osteoblasts.
Asunto(s)
Difosfonatos/farmacología , Osteoblastos , Alendronato/farmacología , Fosfatasa Alcalina/metabolismo , Animales , Biomarcadores/análisis , Células Cultivadas , Inmunohistoquímica , Ratones , Microscopía Electrónica de Rastreo , Osteoblastos/efectos de los fármacos , Osteoblastos/fisiología , Osteoblastos/ultraestructuraRESUMEN
UNLABELLED: To evaluate orthopedic devices in pathological bone, an experimental study was performed by implanting Titanium (Ti) and Hydroxyapatite (HA) rods in normal and osteopenic bone. Twenty-four rats were used: 12 were left intact ( CONTROL: C) while the other 12 were ovariectomized (OVX). After 4 months all the animals were submitted to the implant of Ti or HA in the left femoral condyle (Ti-C, HA-C, Ti-OVX, HA-OVX). Two months later the animals were sacrificed for histomorphometric, ultrastructural and microanalytic studies. Our results show a significant difference between the Affinity Index (A.I.) of HA-C and Ti-C (77.0 +/- 7.4 vs 61.2 +/- 9.7) (p < 0.05). No significant differences were observed between the osteointegration of Ti-C and Ti-OVX (61.2 +/- 9.7 vs 48.2 +/- 6.7). Significant differences also exist between the osteointegration of HA-C and HA-OVX (77.0 +/- 7.4 vs 57.6 +/- 11.5) (p < 0.01). Microanalysis shows some modifications in Sulphur (S) concentration at the bone/biomaterial interface of the Ti-OVX group. Therefore our results confirmed the importance of biomaterials characteristics and of bone quality in osteointegration processes.
Asunto(s)
Materiales Biocompatibles , Fémur/cirugía , Osteoporosis Posmenopáusica/cirugía , Prótesis e Implantes , Animales , Enfermedades Óseas Metabólicas/patología , Enfermedades Óseas Metabólicas/cirugía , Durapatita , Espectroscopía de Resonancia por Spin del Electrón , Femenino , Fémur/patología , Humanos , Microscopía Electrónica de Rastreo , Oseointegración , Osteoporosis Posmenopáusica/patología , Ovariectomía , Ratas , Ratas Sprague-Dawley , TitanioRESUMEN
The aim of this study was to evaluate several chemical pretreatments on the shear bond strength of three dentin bonding agents (DBAs) and to try to correlate dentin morphology with bond strength using scanning electron microscopy (SEM). The DBAs tested were: Gluma Primer in association with Scotchbond DC, Scotchbond DC alone and Clearfil New Bond. Bond strengths of Scotchbond DC and Clearfil New Bond were not significantly modified or reduced by acidic dentin pretreatments. Gluma/Scotchbond adhesion was increased by several treatments but only maleic acid treatment produced shear bond strengths significantly higher than EDTA treatment. SEM evaluation of dentin treatments revealed a wide variety of morphological changes in the dentin surface with partial and complete removal of smear layer by acidic solutions, and only minor modification by amino acid solutions. Only maleic acid was capable of the complete removal of the smear layer and smear plugs coupled with extensive exposure of dentin collagen fibrils.
Asunto(s)
Recubrimiento Dental Adhesivo , Dentina/efectos de los fármacos , Ácido Edético , Maleatos , Grabado Ácido Dental , Adhesivos , Adulto , Humanos , Microscopía Electrónica de RastreoRESUMEN
Shear bond strength of seven dentin bonding systems (DBS) was evaluated. After shear bond testing, the specimens were analyzed under scanning electron microscope (SEM). Buccal dentin just below the dentin-enamel junction of erupted third molars of young subjects were used. A statistical difference was observed among the material bond values. SEM examination of both sides of the sheared bonds frequently showed an adhesive failure between dentin and bonding agents in the specimens where resin penetrated into the opened dentinal tubules. A complex cohesive fracture of both resin and dentin was observed for bonding agents that required the treatment of the smear layer, while a cohesive fracture in the smear layer thickness was noted when dentin was not altered by primer solutions. The study demonstrated that the DBS that require the removal of the smear layer showed a significantly higher shear bond strength and that the bond value reflects the mode of material fracture and the presence of smear layer.
Asunto(s)
Resinas Compuestas , Recubrimiento Dental Adhesivo , Dentina/ultraestructura , Metacrilatos , Ácido Nítrico , Poliuretanos , Cementos de Resina , Cementos Dentales , Glutaral , Humanos , Microscopía Electrónica de Rastreo , Diente Molar , Nitratos , Oxalatos , Ácidos Polimetacrílicos , Resistencia a la TracciónRESUMEN
The interdental papilla is the anatomic site most susceptible to periodontal disease. The gingival epithelium plays an important role as a barrier to protect the underlying connective tissue from exogenous noxious agents. A study was undertaken to evaluate the papillary epithelial response to reconstructive stresses conducted by gingival brushing (oral therapy) and scaling and root planing, intended to induce a keratinization associated with reduction or resolution of connective tissue inflammation. Results showed that patients who prolonged the interproximal hygiene practices for more than 30 minutes a day achieved a satisfactory keratinization of the papillary gingival epithelium.
Asunto(s)
Placa Dental/prevención & control , Encía/ultraestructura , Gingivitis/terapia , Cepillado Dental/métodos , Humanos , Higiene Bucal/métodos , Enfermedades Periodontales/terapiaRESUMEN
CD99, a transmembrane protein encoded by MIC2 gene is involved in multiple cellular events including cell adhesion and migration, apoptosis, cell differentiation and regulation of protein trafficking either in physiological or pathological conditions. In osteosarcoma, CD99 is expressed at low levels and functions as a tumour suppressor. The full-length protein (CD99wt) and the short-form harbouring a deletion in the intracytoplasmic domain (CD99sh) have been associated with distinct functional outcomes with respect to tumour malignancy. In this study, we especially evaluated modulation of cell-cell contacts, reorganisation of the actin cytoskeleton and modulation of signalling pathways by comparing osteosarcoma cells characterised by different metastasis capabilities and CD99 expression, to identify molecular mechanisms responsible for metastasis. Our data indicate that forced expression of CD99wt induces recruitment of N-cadherin and ß-catenin to adherens junctions. In addition, transfection of CD99wt inhibits the expression of several molecules crucial to the remodelling of the actin cytoskeleton, such as ACTR2, ARPC1A, Rho-associated, coiled-coil containing protein kinase 2 (ROCK2) as well as ezrin, an ezrin/radixin/moesin family member that has been clearly associated with tumour progression and metastatic spread in osteosarcoma. Functional studies point to ROCK2 as a crucial intracellular mediator regulating osteosarcoma migration. By maintaining c-Src in an inactive conformation, CD99wt inhibits ROCK2 signalling and this leads to ezrin decrease at cell membrane while N-cadherin and ß-catenin translocate to the plasma membrane and function as main molecular bridges for actin cytoskeleton. Taken together, we propose that the re-expression of CD99wt, which is generally present in osteoblasts but lost in osteosarcoma, through inhibition of c-Src and ROCK2 activity, manages to increase contact strength and reactivate stop-migration signals that counteract the otherwise dominant promigratory action of ezrin in osteosarcoma cells.
Asunto(s)
Antígenos CD/metabolismo , Moléculas de Adhesión Celular/metabolismo , Movimiento Celular , Invasividad Neoplásica/genética , Osteosarcoma/genética , Osteosarcoma/metabolismo , Quinasas Asociadas a rho/metabolismo , Antígeno 12E7 , Citoesqueleto de Actina/metabolismo , Antígenos CD/genética , Western Blotting , Cadherinas/genética , Cadherinas/metabolismo , Adhesión Celular/genética , Moléculas de Adhesión Celular/genética , Línea Celular Tumoral , Movimiento Celular/genética , Proteínas del Citoesqueleto , Técnica del Anticuerpo Fluorescente , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Inmunohistoquímica , Inmunoprecipitación , Microscopía Electrónica de Transmisión , Análisis de Secuencia por Matrices de Oligonucleótidos , Osteosarcoma/patología , ARN Interferente Pequeño , Reacción en Cadena en Tiempo Real de la Polimerasa , Transfección , Quinasas Asociadas a rho/genéticaAsunto(s)
Cesárea , Hospitales Especializados , Adulto , Femenino , Humanos , Mortalidad Infantil , Recién Nacido , Italia , Edad Materna , Mortalidad Materna , Paridad , Complicaciones Posoperatorias , Embarazo , PronósticoRESUMEN
Toothbrushing technique may represent an important tool to improve gingival keratinization. Our experience evidenced a close relationship between this endoral therapy and interdental epithelial recovery of gingiva, after two months of treatment. Mechanical or microenvironmental stimuli and genetically determined potentialities are the main factors involved in this clinical-therapeutical recovery to modulate structural epithelial behaviour.
Asunto(s)
Encía/ultraestructura , Enfermedades Periodontales/terapia , Cepillado Dental , Enfermedad Crónica , Epitelio/fisiología , HumanosRESUMEN
The post-ovariectomy osteoporotic rat model is widely used to mimic post menopausal human osteoporosis and to test the efficacy of the therapies used in its treatment An experimental study was performed in order to improve the reliability of bone mineral density (BMD) measurements by correcting also the relocation error. In addition, a BMD map of the whole femur was carried out to detect the areas of uniform density, where the reliability of measuring was improved. Right femurs were taken from 5 Sprague Dawley female rats, 10 months old, and tested every 2 mm from the supracondylar line to the peritrochanteric line (6 scans for each site). Repositioning error was tested for each site of measurement, and measurements done by 3 different operators in a double blind test were compared. At site 4 and 24 (where the differences are high) and site 10 (where the differences of BMD are low) the coefficient of variability (CV) was calculated repositioning the bone after each measurement scan for a total of 6 scans. The CV at sites 4 and 24 were significantly higher than at site 10 (p < 0.05). In our opinion, the bone segment between 8 and 14 mm from the supracondylar line is that which presents the highest homogeneity and where it is preferable to perform the measurements in order to obtain the maximum effect of the method (improvement of precision of about 30%) in comparison with the other femur sites tested.
Asunto(s)
Absorciometría de Fotón , Densidad Ósea , Fémur/química , Ratas/anatomía & histología , Absorciometría de Fotón/normas , Animales , Huesos/química , Femenino , Fémur/anatomía & histología , Humanos , Minerales/análisis , Osteoporosis Posmenopáusica , Ratas Sprague-Dawley , Estándares de Referencia , Valores de Referencia , Reproducibilidad de los ResultadosRESUMEN
We have investigated structural and functional properties of skeletal muscle mitochondria obtained from biopsies from young and old individuals. The morphometric analysis of muscle sections revealed a tendency to an increase of total area, numerical density and volume density of mitochondria in the aged. The enzymatic activities of NADH-Coenzyme Q reductase, succinate cytochrome c reductase, ubiquinol-cytochrome c reductase exhibited a high variability of specific activities without any correlation with age. Expression of the values as enzyme turnovers reduced the variability but was unable to reveal any age-dependent modification.
Asunto(s)
Complejo III de Transporte de Electrones/análisis , Mitocondrias Musculares/enzimología , Músculo Esquelético/enzimología , NADH NADPH Oxidorreductasas/análisis , Succinato Citocromo c Oxidorreductasa/análisis , Adolescente , Adulto , Factores de Edad , Anciano , Anciano de 80 o más Años , Complejo I de Transporte de Electrón , Humanos , Microscopía Electrónica , Músculo Esquelético/ultraestructuraRESUMEN
One characteristic feature of the aged central nervous system (CNS) is neuron loss. Programmed cell death (PCD) has been implicated in neuronal death during development and may be involved in a number of age-related neurodegenerative diseases of the CNS. Cell death in the aging cerebral cortex was investigated in the present morphometric and immunohistochemical study of rat frontal cortex by detection of bcl-2 as the factor preventing PCD. The results were interpreted in the light of the bioenergetic features of aged motoneuron cells. Our results showed that 1) bcl-2 does not influence neuronal survival, and ii) the presence in aging frontal cortex of minor cellular morphometric and bioenergetic modifications, confirming the difference between normal aging and neurodegenerative disease.
Asunto(s)
Envejecimiento , Corteza Cerebral/anatomía & histología , Envejecimiento/metabolismo , Animales , Muerte Celular , Senescencia Celular , Corteza Cerebral/irrigación sanguínea , Corteza Cerebral/metabolismo , Técnica del Anticuerpo Fluorescente , Inmunohistoquímica , Masculino , Microscopía Electrónica , Mitocondrias/ultraestructura , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Ratas , Ratas WistarRESUMEN
The application of bioactive ceramic coatings to prostheses confers strength to a material (ceramic or biological glass) that exerts beneficial effects on bone-tissue growth but that itself lacks the toughness and stability required of an implant device. The rate of bioactivity is related to the chemical reactivity of the material and causes interface dissolution, precipitation and ion-exchange reactions. Ceramics may differ in sintering temperature and thus exhibit differences in their in vitro dissolution features and in vivo performance. To test these effects, in vitro and in vivo studies were carried out on two biocompatible biological glasses and a ceramic of proven bioactivity in view of their potential utilization as covering materials. In addition, a modified chitosan was adsorbed on the surface of a series of hydroxyapatite (HA) samples. Human fibroblasts and/or osteoblasts were used for the in vitro tests, and normal (INT) and osteoporotic (OVX) rats, normal rabbits and sheep for the in vivo studies. Similar chemical changes were observed in both glasses, suggesting that these materials underwent modifications directly dependent on their biological environment. The in vivo tests point to the possibility of improving the bioactivity of ceramic substrates with chitosan. However, the different behaviour of the materials in vitro and in vivo suggests that these tests should be conducted in parallel.