RESUMEN
Detecting and locating prey are key to predatory success within trophic chains. Predators use various signals through specialized visual, olfactory, auditory or tactile sensory systems to pinpoint their prey. Snakes chemically sense their prey through a highly developed auxiliary olfactory sense organ, the vomeronasal organ (VNO). In natricine snakes that are able to feed on land and water, the VNO plays a critical role in predatory behavior by detecting cues, known as vomodors, which are produced by their potential prey. However, the chemical nature of these cues remains unclear. Recently, we demonstrated that specific proteins-parvalbumins-present in the cutaneous mucus of the common frog (Rana temporaria) may be natural chemoattractive proteins for these snakes. Here, we show that parvalbumins and parvalbumin-like proteins, which are mainly intracellular, are physiologically present in the epidermal mucous cells and mucus of several frog and fish genera from both fresh and salt water. These proteins are located in many tissues and function as Ca(2+) buffers. In addition, we clarified the intrinsic role of parvalbumins present in the cutaneous mucus of amphibians and fishes. We demonstrate that these Ca(2+)-binding proteins participate in innate bacterial defense mechanisms by means of calcium chelation. We show that these parvalbumins are chemoattractive for three different thamnophiine snakes, suggesting that these chemicals play a key role in their prey-recognition mechanism. Therefore, we suggest that recognition of parvalbumin-like proteins or other calcium-binding proteins by the VNO could be a generalized prey-recognition process in snakes. Detecting innate prey defense mechanism compounds may have driven the evolution of this predator-prey interaction.
Asunto(s)
Parvalbúminas/fisiología , Conducta Predatoria/fisiología , Olfato/fisiología , Serpientes/fisiología , Animales , Evolución Biológica , Reconocimiento en Psicología/fisiologíaRESUMEN
Superparamagnetic iron oxide nanoparticles (SPION) are very promising contrast media, especially for molecular imaging, due to their superior NMR efficacy. They even have wider biomedical applications such as in drug and gene delivery, tissue engineering and bioseparation, or as sensitive biological nanosensors. By coupling them to affinity ligands, SPION can bind to drugs, proteins, enzymes, antibodies or nucleotides. For in vitro biomedical applications, the detection of molecular interaction is possible by using a diversity of systems capable of sensing the magnetic properties of these materials. The goal of the present work was to develop and validate various in vitro biomedical applications of ultrasmall superparamagnetic particles of iron oxide (USPIO), including some that are not related to their magnetic properties. USPIO coated with dextran, starch or bisphosphonate exposing carboxylate groups were synthesized and some of them were functionalized by conjugating various biomolecules, such as biotin, streptavidin and apoptosis, or VCAM-1 specific peptides. The in vitro biomedical applications assessed in the present work included: (1) the relaxometric measurement of antibody concentration, cell receptor expression, molecular interaction, and enzymatic activity in aqueous suspensions; (2) MRI visualization of cells and detection of molecular interaction in an ELISA system; (3) ELISA applications of USPIO derivatives; and (4) detection of specific biomolecules by histochemistry. Our results confirm that rapid and simple in vitro detection of a diversity of functionalized SPION with relevance in medicine is possible by the existing NMR techniques and by chemical staining reactions. The protocols can be applied to minimally prepared biological samples (e.g. whole blood, blood plasma or serum, cell suspensions, biopsies, histological preparations, etc.), and often do not need complicated systems of signal amplification. The use of SPION labeled compounds could furthermore contribute to cost reductions in the diagnosis and in patient care.
Asunto(s)
Medios de Contraste/química , Compuestos Férricos/química , Nanopartículas/química , Apoptosis , Complejo CD3/metabolismo , Línea Celular , Ensayo de Inmunoadsorción Enzimática , Humanos , Inmunohistoquímica , Imagen por Resonancia MagnéticaRESUMEN
AIMS: We used a rat model of renal ischemia (35 min) to test the potential involvement of platelet/endothelial cell adhesion molecule 1 (PECAM-1/CD31) in the process of S3 tubule regeneration. METHODS: A monoclonal antibody specific for murine PECAM-1 was injected i.p. immediately after kidney reperfusion or 48 h post-ischemia. One day before ischemia, each animal received an i.p. injection of 80 mg/kg 5-bromo-2'-deoxyuridine (BrdU). Experimental animals were sacrificed 1, 2, 3, 7 and 14 days post-ischemia. Renal sections were processed to characterize the histopathological alterations and the distribution of BrdU-immunopositive cells. RESULTS: Our observations showed that anti-PECAM-1 administration was associated with an inhibition of S3 tubule regeneration along with a progressive cystic dilatation of renal tubules that was particularly prominent 2 weeks post-ischemia. Interestingly, injection of anti-PECAM-1 48 h post-ischemia failed to block renal regeneration and was followed by a normal re-epithelialization of S3 tubules. CONCLUSION: Our data showed that the blockade of PECAM-1 immediately after kidney reperfusion inhibits tubular regeneration. These observations suggest that transendothelial migration of extrarenal cells could be a precocious and pivotal step in kidney reparation, but also suggest that these extrarenal cells could be essential to the process of tubular regeneration.
Asunto(s)
Anticuerpos Monoclonales de Origen Murino/farmacología , Túbulos Renales/fisiología , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/inmunología , Daño por Reperfusión/inmunología , Animales , Necrosis Tubular Aguda/inmunología , Necrosis Tubular Aguda/patología , Masculino , Ratones , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/fisiología , Ratas , Ratas Wistar , Regeneración/efectos de los fármacos , Regeneración/inmunología , Daño por Reperfusión/tratamiento farmacológico , Migración Transendotelial y TransepitelialRESUMEN
Endogenous lectins act as effectors of cellular activities such as growth regulation, migration and adhesion. In this study, we report the histochemical detection of galectins and their binding sites in rat kidneys after ischemic injury (35 min) with regard to renal regeneration. In this context, we have shown in a previous publication (Vansthertem et al., 2008) that extrarenal cells (CD44+, vimentin +) could be involved in this process of tubular restoration. In controls, galectin-1 is expressed by fusiform-shaped cells within cortical and medullar interstitium. Two days after ischemia, the number of positive interstitial cells increased temporarily within OSOM in the vicinity of altered tubules to later reach control level. After ischemia, we identified a population of galectin-3 (+), CD44 (+), and vimentin (+) interstitial round cells located in the outer stripe of outer medulla (OSOM) in the vicinity of necrotic tubules, but also in the lumen of adjacent blood vessels. The immunocytochemical characteristics of theses cells, along with their distribution within OSOM, suggest the involvement of a unique cell population during kidney regeneration. On the other hand, the distribution and density of binding sites for galectins within OSOM were not modified after ischemia and remained similar to controls. Altogether, our observations suggest that galectin-3 may be involved in the complex process of kidney regeneration following ischemia/reperfusion injury.
Asunto(s)
Galectina 1/metabolismo , Galectina 3/metabolismo , Túbulos Renales/metabolismo , Daño por Reperfusión/metabolismo , Animales , Sitios de Unión , Inmunohistoquímica , Masculino , Ratas , Ratas Wistar , Regeneración , Daño por Reperfusión/fisiopatologíaRESUMEN
This work explores the distribution of various markers expressed by interstitial cells in rat kidneys after ischemic injury (35 minutes) during regeneration of S3 tubules of outer stripe of outer medulla (OSOM). Groups of experimental animals (n = 4) were sacrificed every two hours during the first 24 hours post-ischemia as well as 2, 3, 7, 14 days post-ischemia. The occurrence of lineage markers was analyzed on kidney sections by immunohistochemistry and morphometry during the process of tubular regeneration. In postischemic kidneys, interstitial cell proliferation, assessed by 5-bromo-2'-deoxyuridine (BrdU) and Proliferating Cell Nuclear Antigen (PCNA) labeling, was prominent in outer medulla and reach a maximum between 24 and 72 hours after reperfusion. This population was characterized by the coexpression of vimentin and nestin. The density of -Neural Cell Adhesion Molecule (NCAM) positive interstitial cells increased transiently (18-72 hours) in the vicinity of altered tubules. We have also localized a small population of alpha-Smooth Muscle Actin (SMA)-positive cells confined to chronically altered areas and characterized by a small proliferative index. In conclusion, we observed in the postischemic kidney a marked proliferation of interstitial cells that underwent transient phenotypical modifications. These interstitial cells could be implicated in processes leading to renal fibrosis.
Asunto(s)
Proteínas de Filamentos Intermediarios/metabolismo , Isquemia/metabolismo , Túbulos Renales Proximales , Proteínas del Tejido Nervioso/metabolismo , Moléculas de Adhesión de Célula Nerviosa/metabolismo , Vimentina/metabolismo , Actinas/metabolismo , Animales , Bromodesoxiuridina , Proliferación Celular , Inmunohistoquímica , Túbulos Renales Proximales/citología , Túbulos Renales Proximales/metabolismo , Masculino , Microscopía Fluorescente , Nestina , Antígeno Nuclear de Célula en Proliferación/metabolismo , Ratas , Ratas Wistar , Fase SRESUMEN
Amyloid plaques (AP) represent one of the main molecular hallmarks of Alzheimer's disease (AD). In order to develop new AP-specific contrast agents for AD molecular imaging, the phage display technology was used to identify peptides specific to amyloid-beta (A beta(42)). A random disulfide constrained heptapeptide phage display library was screened against A beta(42). After biopanning, 72 phage clones were isolated and their binding affinity to A beta(42) was evaluated by enzyme-linked immunosorbent assay (ELISA). The final library was enriched in two peptide sequences. The K(d) of candidate phage clones for binding to A beta(42) are in the picomolar range. The binding affinity for A beta(42) of two selected peptides was confirmed by ELISA, and the specific interaction with AP was validated by immunohistochemistry on brain sections. The preliminary MRI in vivo study, which was performed with a peptide functionalized contrast agent on AD transgenic mouse, showed encouraging results. To conclude, low molecular weight peptides presenting a specific affinity for A beta(42) were identified by phage display. As specific carriers, they have a real potential for molecular imaging of AD thanks to AP binding.
Asunto(s)
Enfermedad de Alzheimer/diagnóstico , Encéfalo/patología , Medios de Contraste , Fragmentos de Péptidos , Placa Amiloide/patología , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Medios de Contraste/química , Masculino , Ratones , Ratones Transgénicos , Datos de Secuencia Molecular , Fragmentos de Péptidos/química , Biblioteca de Péptidos , Ratas , Ratas WistarRESUMEN
The vascular cell adhesion molecule-1 (VCAM-1) has distinct roles in inflammatory cell recruitment to the damaged vessel wall. In the present work, a cyclic heptapeptide phage displayed library was screened in vitro during four rounds of biopanning. On the basis of Kd and IC50 values, a peptide (encoded as R832) was selected for in vitro and in vivo validation. After conjugation to Gd-DOTA, VCAM-1 imaging was assessed by MRI on a model of T cell mediated hepatitis, induced in mice by concanavalin A. On histological samples, the location of biotinylated R832 (R832-Bt) around liver veins in hepatitis resembles the pattern of MRI enhancement. Gd-DOTA-R832 was then assessed on ApoE(-/-) mice and produced an important signal enhancement of the aortic wall, while R832-Bt interacted with morphologic structures comparable to those marked by anti-VCAM-1 antibody. In conclusion, the in vitro and in vivo evaluation of peptide R832 suggests a specific interaction with the targeted biomolecule. Its conjugation to imaging reporters could assist the diagnosis of inflammatory diseases.
Asunto(s)
Hepatitis/metabolismo , Imagen por Resonancia Magnética/métodos , Biblioteca de Péptidos , Molécula 1 de Adhesión Celular Vascular/análisis , Secuencia de Aminoácidos , Animales , Apolipoproteínas E/fisiología , Células Cultivadas , Compuestos Heterocíclicos , Humanos , Integrina alfa4beta1/metabolismo , Integrina beta1/metabolismo , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Compuestos OrganometálicosRESUMEN
OBJECTIVES/HYPOTHESIS: To examine the level of expression of galectin-3 in relation to neoplastic progression of hypopharyngeal squamous cell carcinomas (HSCCs) and laryngeal squamous cell carcinomas (LSCCs). STUDY DESIGN: Retrospective study. METHODS: Using a polyclonal antibody against galectin-3 without cross-reactivity to other galectins, we analyzed the presence of galectin-3 using quantitative immunohistochemistry in i) a series of 79 HSCCs compared with 16 normal epithelia, 20 low-grade dysplasia (Low_D) and 25 high-grade dysplasia (High_D) and in ii) a series of 58 LSCCs compared with 34 normal epithelia, 12 Low_D, and 18 High_D. In parallel, galectin-3 expression was studied using Western blotting on a series of 19 fresh biopsies from patients presenting a head and neck tumor. RESULTS: Western blotting excluded a notable degree of proteolytic truncation of galectin-3 in situ. Immunohistochemical galectin-3 positivity expressed as percentage of cells was significantly higher in LSCCs and HSCCs than in Low_D (P = .01) or High_D (P = .0002), respectively. Increased expression of galectin-3 in HSCCs was accompanied by a shift from the cytoplasmic compartment to the nucleus (P = .007). In intertumor-type comparison, laryngeal carcinomas presented nuclear presence of galectin-3 only rarely (1 of 58 cases in laryngeal cancer vs. 27 of 79 cases in hypopharyngeal cancer, P = .00006) and a comparatively low labeling index (P < 10(-6)). CONCLUSIONS: Our data reveal an association between level of presence of galectin-3 and neoplastic progression of HSCCs and LSCCs.
Asunto(s)
Carcinoma de Células Escamosas/metabolismo , Galectina 3/biosíntesis , Neoplasias Hipofaríngeas/metabolismo , Neoplasias Laríngeas/metabolismo , Regulación hacia Arriba , Adulto , Anciano , Anticuerpos Antineoplásicos/inmunología , Biomarcadores de Tumor/biosíntesis , Western Blotting , Carcinoma de Células Escamosas/patología , Progresión de la Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Estudios de Seguimiento , Galectina 3/inmunología , Neoplasias de Cabeza y Cuello/metabolismo , Neoplasias de Cabeza y Cuello/patología , Humanos , Neoplasias Hipofaríngeas/patología , Inmunohistoquímica , Neoplasias Laríngeas/patología , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Estudios RetrospectivosRESUMEN
BACKGROUND: In this study, we have examined rat kidneys after ischaemic injury (35 min) with regard to the dynamics of S3 tubule regeneration. METHODS: One day before ischaemia, each rat received four successive i.p. injections of BrdU (5-bromo-2'-deoxyuridine: 80 mg/kg) at 2 h intervals. Groups of experimental animals (n = 4) were killed every 2 h during the first 24 h post-ischaemia as well as 2, 3, 7 and 14 days post-ischaemia. Renal sections were processed to characterize by immunohistochemistry the distribution and phenotype of BrdU-positive cells. RESULTS: Renal regeneration after ischaemia was associated with a typical sequence of transient events: (1) absence of immunostaining during the first 8 h after reperfusion; (2) between 8 and 16 h, detection of a small population of BrdU-positive cells (CD44(+), vimentin(+), CD45(-)) restricted to the lumen of blood vessels characterized by the endothelial expression of selectin E; (3) between 16 and 24 h, progressive decrease of labelled cells in renal capillaries and a concomitant increase in the interstitial compartment; (4) after 1 day, labelled cells disappeared progressively from peritubular interstitium and were mainly observed in regenerating S3 tubules, and (5) after 3 days numerous positive cells were only present in regenerated tubules. CONCLUSIONS: Our data suggest that positive cells (BrdU(+), CD44(+), vimentin(+) and CD45(-)) observed in kidney tubules after ischaemia could originate from an extrarenal source and reach the renal parenchyma via blood vessels. We postulate that these immature cells migrate to injured tubules, proliferate and finally differentiate into mature epithelial cells leading to the replacement of a majority (>80%) of altered S3 cells.
Asunto(s)
Riñón/irrigación sanguínea , Riñón/lesiones , Actinas/metabolismo , Lesión Renal Aguda/etiología , Lesión Renal Aguda/patología , Lesión Renal Aguda/fisiopatología , Animales , Antígenos CD/metabolismo , Antígenos de Diferenciación Mielomonocítica/metabolismo , Bromodesoxiuridina/metabolismo , Movimiento Celular , Selectina E/metabolismo , Receptores de Hialuranos/metabolismo , Ácido Hialurónico/metabolismo , Inmunohistoquímica , Riñón/fisiopatología , Necrosis Tubular Aguda/etiología , Necrosis Tubular Aguda/patología , Necrosis Tubular Aguda/fisiopatología , Túbulos Renales/patología , Túbulos Renales/fisiopatología , Antígenos Comunes de Leucocito/metabolismo , Masculino , Cadenas Pesadas de Miosina/metabolismo , Ratas , Ratas Wistar , Regeneración , Daño por Reperfusión/complicaciones , Daño por Reperfusión/patología , Daño por Reperfusión/fisiopatología , Vimentina/metabolismoRESUMEN
AIMS: The integrin alpha v beta3 is highly expressed in atherosclerotic plaques by medial and intimal smooth muscle cells and by endothelial cells of angiogenic microvessels. In this study, we have assessed non-invasive molecular magnetic resonance imaging (MRI) of plaque-associated alpha v beta3 integrin expression on transgenic ApoE-/- mice with a low molecular weight peptidomimetic of Arg-Gly-Asp (mimRGD) grafted to gadolinium diethylenetriaminepentaacetate (Gd-DTPA-g-mimRGD). The analogous compound Eu-DTPA-g-mimRGD was employed for an in vivo competition experiment and to confirm the molecular targeting. The specific interaction of mimRGD conjugated to Gd-DTPA or to 99mTc-DTPA with alpha v beta3 integrin was furthermore confirmed on Jurkat T lymphocytes. METHODS AND RESULTS: The mimRGD was synthesized and conjugated to DTPA. DTPA-g-mimRGD was complexed with GdCl3.6H2O, EuCl3.6H2O, or with [99mTc(CO)3(H2O)3]+. MRI evaluation was performed on a 4.7 T Bruker imaging system. Blood pharmacokinetics of Gd-DTPA-g-mimRGD were assessed in Wistar rats and in c57bl/6j mice. The presence of angiogenic blood vessels and the expression of alpha v beta3 integrin were confirmed in aorta specimens by immunohistochemistry. Gd-DTPA-g-mimRGD produced a strong enhancement of the external structures of the aortic wall and of the more profound layers (possibly tunica media and intima). The aortic lumen seemed to be restrained and distorted. Pre-injection of Eu-DTPA-g-mimRGD diminished the Gd-DTPA-g-mimRGD binding to atherosclerotic plaque and confirmed the specific molecular targeting. A slower blood clearance was observed for Gd-DTPA-g-mimRGD, as indicated by a prolonged elimination half-life and a diminished total clearance. CONCLUSION: The new compound is potentially useful for the diagnosis of vulnerable atherosclerotic plaques and of other pathologies characterized by alpha v beta3 integrin expression, such as cancer and inflammation. The delayed blood clearance, the significant enhancement of the signal-to-noise ratio, and the low immunogenicity of the mimetic molecule highlight its potential for an industrial and clinical implementation.
Asunto(s)
Aorta/metabolismo , Aterosclerosis/metabolismo , Medios de Contraste/metabolismo , Integrina alfaVbeta3/metabolismo , Angiografía por Resonancia Magnética/métodos , Compuestos Organometálicos/metabolismo , Ácido Pentético/análogos & derivados , Animales , Aorta/patología , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Aterosclerosis/patología , Unión Competitiva , Medios de Contraste/farmacocinética , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunohistoquímica , Células Jurkat , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Compuestos Organometálicos/farmacocinética , Ácido Pentético/metabolismo , Ácido Pentético/farmacocinética , Radiofármacos/metabolismo , Ratas , Ratas Wistar , Procesamiento de Señales Asistido por Computador , Pentetato de Tecnecio Tc 99m/metabolismoRESUMEN
To evaluate galectin-1, -3 and -7 serum levels as diagnostic and/or prognostic markers for head and neck squamous cell carcinomas (HNSCCs). ELISA was employed to test sera from 102 patients with HNSCCs and from 38 healthy control volunteers for galectin-1, -3 and -7 serum levels. Serum galectin levels were assayed by ELISA and the levels of galectin expression in HNSCCs were determined by means of immunohistochemistry. HNSCCs display significant immunohistochemical amounts of galectin-7, but this galectin cannot be detected in the blood of HNSCC patients. Galectin-3 levels differ significantly (p=0.03) in healthy volunteers and HNSCC patients. Using a threshold value of 4.3 ng/ml, galectin-3 serum level enabled a significant level of discrimination (p=0.03) to be established between the cancer patients and the healthy volunteers, with 90% level of specificity and 36% level of sensitivity. The discrimination was even better when using a threshold value of 13.5 ng/ml for galectin-1 (p=0.001), with 100% level of specificity and 22% level of sensitivity. A subgroup of stage IV HNSCC patients displayed significantly reduced levels of circulating galectin-1 (p=0.003) and galectin-3 (p=0.001) after treatment as opposed to before. Galectin-3 concentrations in sera from the patients with a metastatic disease were significantly (p=0.01) higher than in sera from the patients with localized tumors. The determination of circulating levels of galectin-1 and -3 could be used to monitor the progression of their disease or their response to therapy.
Asunto(s)
Biomarcadores de Tumor/sangre , Carcinoma de Células Escamosas/diagnóstico , Galectina 1/sangre , Galectina 3/sangre , Neoplasias de Cabeza y Cuello/diagnóstico , Adulto , Anciano , Anciano de 80 o más Años , Ensayo de Inmunoadsorción Enzimática , Estudios de Factibilidad , Femenino , Galectinas/sangre , Humanos , Masculino , Persona de Mediana Edad , Sensibilidad y EspecificidadRESUMEN
Monitoring of gene-expression profiles is assumed to refine tumor characterization of laryngeal squamous cell carcinomas (LSCCs) with a therapeutic perspective. This is especially expected for adhesion/growth-regulatory effectors such as galectins, a class of endogenous lectins. Using computer-assisted microscopy, we investigated the prognostic value contributed by the quantitative determination of the immunohistochemical levels of expression of galectin-1, -3 and -7 in a series of 62 LSCCs including 42 low- and 20 high-stage LSCCs. As galectin-1 may have a key role leading to a tumor escape from immune surveillance, we also investigated whether or not the level of galectin-1 expression correlated with lymphocyte infiltration in LSCCs. The immunohistochemical determination of expression of galectin-1 is of prognostic value in human squamous laryngeal cancers. LSCCs that display high levels of galectin-1 have worse prognoses than laryngeal cancers with low levels of galectin-1 expression. Elevation of galectin-1 levels in laryngeal cancers can contribute to the process of tumor immune escape by killing the activated T-cells and other protumoral activities such as promoting motility or activity of oncogenic H-Ras proteins. The quantitative determination of galectin-1 in LSCCs is an independent prognostic marker when opposed to TNM staging. It has the potential to identify patients unlikely to benefit from T-cell-mediated immunotherapy, although the definitive effector function from its pro- and antitumoral activity profile has not been delineated.
Asunto(s)
Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Galectina 1/biosíntesis , Regulación Neoplásica de la Expresión Génica , Neoplasias Laríngeas/metabolismo , Neoplasias Laríngeas/patología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Linfocitos/metabolismo , Masculino , Persona de Mediana Edad , Pronóstico , Recurrencia , Estudios RetrospectivosRESUMEN
Head and neck squamous cell carcinomas (HNSCCs) remain a significant cause of morbidity worldwide. Biological therapies able to induce and/or upregulate antitumor immune responses could represent a complementary approach to conventional treatments for patients with HNSCC because, despite advances in surgery, radiotherapy, and chemotherapy, the overall survival rates for these patients have not changed over recent decades. Galectins are involved in the control of cell proliferation, cell death, and cell migration and in the modulation of various functions of the immune system. In this context, galectin-1 is known to protect HNSCCs from the immune system. The present review details the involvement of galectins in HNSCC biology and suggests a number of approaches to reduce the levels of expression of galectin-1 in HNSCCs, with the aim of improving the efficiency of HNSCC immunotherapy.
Asunto(s)
Carcinoma de Células Escamosas/fisiopatología , Galectina 1/fisiología , Galectinas/fisiología , Neoplasias de Cabeza y Cuello/fisiopatología , Carcinoma de Células Escamosas/epidemiología , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/inmunología , Carcinoma de Células Escamosas/cirugía , Movimiento Celular/fisiología , Progresión de la Enfermedad , Galectina 1/metabolismo , Galectina 3/metabolismo , Galectinas/metabolismo , Neoplasias de Cabeza y Cuello/epidemiología , Neoplasias de Cabeza y Cuello/genética , Neoplasias de Cabeza y Cuello/inmunología , Humanos , Inmunohistoquímica , Pronóstico , Linfocitos T/fisiologíaRESUMEN
The vomeronasal organ is a chemosensory organ present in most vertebrates and involved in chemical communication. In the last decade, the deciphering of the signal transduction process of this organ has progressed. However, less is known about the vomeronasal organ ligands and their structure-function relationships. Snakes possess a highly developed vomeronasal system that is used in various behaviors such as mating, predator detection, or prey selection, making this group a suitable model for study of the vomeronasal chemoreception. In this work, we used a proteomics approach to identify and characterize proteins from frog cutaneous mucus proteome involved in prey recognition by snakes of the genus Thamnophis. Herein we report the purification and characterization of two proteins isolated from the frog skin secretome that elicit the vomeronasal organ-mediated predatory behavior of Thamnophis marcianus. These proteins are members of the parvalbumin family, which are calcium-binding proteins generally associated to muscular and nervous tissues. This is the first report that demonstrates parvalbumins are not strictly restricted to intracellular compartments and can also be isolated from exocrine secretions. Purified parvalbumins from frog muscle and mucus revealed identical chemoattractive properties for T. marcianus. Snake bioassay revealed the Ca(2+)/Mg(2+) dependence of the bioactivity of parvalbumins. So parvalbumins appear to be new candidate ligands of the vomeronasal organ.
Asunto(s)
Anuros/metabolismo , Factores Quimiotácticos/aislamiento & purificación , Colubridae/fisiología , Parvalbúminas/aislamiento & purificación , Proteoma/química , Piel/química , Secuencia de Aminoácidos , Animales , Bioensayo , Calcio/farmacología , Cationes Bivalentes/química , Factores Quimiotácticos/análisis , Factores Quimiotácticos/metabolismo , Inmunohistoquímica , Magnesio/farmacología , Datos de Secuencia Molecular , Moco/química , Parvalbúminas/análisis , Parvalbúminas/metabolismo , Proteoma/metabolismo , Órgano Vomeronasal/fisiologíaRESUMEN
BACKGROUND: Eighty percent of hypopharyngeal squamous cell carcinoma patients have advanced stages (III and IV) of the disease, and biological markers are required to predict high-risk head and neck squamous cell carcinoma patients in need of highly aggressive treatments after surgery to improve the survival rate. We analyzed the potential prognostic value of galectin 7 in a series of 81 stage IV hypopharyngeal SCCs because galectin 7 is an emerging marker involved in the epidermal development of pluristratified epithelia and in epidermal cell migration. METHODS: The immunohistochemical expression of galectin 7 was determined on a series of 81 stage IV hypopharyngeal SCCs and was compared with that of galectins 1 and 3. RESULTS: High levels of galectin 7 expression were associated with rapid recurrence rates and dismal prognoses in these 81 stage IV hypopharyngeal SCCs, a feature not observed with galectin 3 and one observed weakly, if at all, with galectin 1. CONCLUSIONS: These data suggest that the immunohistochemical determination of galectin 7 expression in the case of high-risk hypopharyngeal cancers is a meaningful tool to identify patients who should benefit from aggressive postsurgical adjuvant therapy after surgery, including not only radiotherapy, but also chemotherapy.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Galectinas/metabolismo , Neoplasias Hipofaríngeas/metabolismo , Neoplasias Hipofaríngeas/mortalidad , Recurrencia Local de Neoplasia/patología , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidad , Adenocarcinoma/patología , Adulto , Anciano , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/mortalidad , Carcinoma de Células Escamosas/patología , Femenino , Galectina 3/metabolismo , Humanos , Neoplasias Hipofaríngeas/patología , Técnicas para Inmunoenzimas , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Tasa de SupervivenciaRESUMEN
Ischemia/reperfusion (I/R) injury in the kidney involves hemodynamic and cellular dysfunctions as well as leukocyte infiltration. Functional recovery occurs via cell proliferation and/or migration. To determine the roles of hyaluronan (HA) and its main receptor CD44 in renal postischemic processes, we compared their localization and expression with that of neutrophils, macrophages, and PCNA-positive (regenerative) cells as characterized by immunohistochemistry, up to 28 days after I/R in uninephrectomized rats. Observations covered all kidney zones, i.e. cortex (C), outer and inner stripes of outer medulla (OSOM, ISOM), and inner medulla (IM). In controls, HA was localized to the interstitium of IM and ISOM, and CD44 was mostly present on the basolateral membranes of collecting ducts in ISOM, the thin descending limb of Henle's loop and macula densa cells. After I/R, HA and CD44 staining appeared in C and OSOM at 12 h and persisted throughout the regenerative period, i.e. until day 7. Thereafter, they regressed but remained associated with remodeling areas. CD44 expression was found de novo on the apical pole of regenerating, not fully differentiated tubular cells and on some interstitial cells. It was prominent on all infiltrating neutrophils, as soon as 2 h post-I/R, and on 30% of the macrophages, including those in late HA-rich inflammatory granulomas. CD44 is probably involved in early leukocyte infiltration, in tubular regeneration, and in macrophage activity, while HA modifies the physico-chemical environment of interstitial and migrating cells. Based on its presence in remodeling areas, the HA-CD44 pair may be implicated in persistent postichemic inflammation as observed in chronic allograft nephropathy.
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Receptores de Hialuranos/análisis , Ácido Hialurónico/análisis , Daño por Reperfusión/metabolismo , Animales , Acuaporina 3/análisis , Modelos Animales de Enfermedad , Ectodisplasinas , Inmunohistoquímica , Riñón/irrigación sanguínea , Riñón/química , Riñón/patología , Corteza Renal/química , Corteza Renal/patología , Enfermedades Renales/metabolismo , Enfermedades Renales/patología , Médula Renal/química , Médula Renal/patología , Túbulos Renales/química , Túbulos Renales/patología , Masculino , Proteínas de la Membrana/análisis , Neutrófilos/química , Neutrófilos/patología , Ratas , Ratas Wistar , Daño por Reperfusión/patología , Factores de Tiempo , Factores de Necrosis Tumoral/análisisRESUMEN
BACKGROUND: The Helicase-Like Transcription Factor (HLTF/SMARCA3) belongs to the family of SWI/SNF proteins that use the energy of ATP hydrolysis to remodel chromatin in a variety of cellular processes. Several SWI/SNF genes are disrupted in cancer, suggesting a role of tumor suppressor. Similarly, the HLTF gene was recently found to be inactivated by hypermethylation in a number of advanced colon and gastric tumors. However, other evidences indicated a 20-fold HLTF overexpression in cell lines derived from various neoplasms (ovary, breast, cervix, kidney...). RESULTS: In the present study, we investigated HLTF expression by immunohistochemistry in a model of kidney tumors induced by continuous administration of diethylstilbestrol to male Syrian golden hamsters. A strong labeling was already detected in small tumor buds, making HLTF an early cancer marker in this model. Although every cell stained for HLTF at this early stage, the number of HLTF-positive cells decreased to 10% with cancer progression, and these positive cells were dispersed in the tumor mass. HLTF expression was conserved in the HKT-1097 cell line established from kidney tumors, but again only 10% of positive cells were found in xenografts produced by HKT-1097 cells in nude mice. CONCLUSION: In conclusion, our data suggest that HLTF gene activation is linked to initial steps of carcinogenesis in this model and should be investigated in early stages of other neoplasms.
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ADN Helicasas/metabolismo , Proteínas de Unión al ADN/metabolismo , Estrógenos/farmacología , Neoplasias Renales/genética , Factores de Transcripción/metabolismo , Animales , Secuencia de Bases , Cricetinae , Proteínas de Unión al ADN/análisis , Proteínas de Unión al ADN/genética , Dietilestilbestrol/farmacología , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Inmunohistoquímica , Riñón/efectos de los fármacos , Riñón/metabolismo , Riñón/patología , Neoplasias Renales/inducido químicamente , Neoplasias Renales/metabolismo , Masculino , Mesocricetus , Ratones , Ratones Desnudos , Datos de Secuencia Molecular , Neoplasias Experimentales/genética , Neoplasias Experimentales/metabolismo , Neoplasias Experimentales/patología , Homología de Secuencia de Ácido Nucleico , Factores de Transcripción/análisis , Factores de Transcripción/genética , Activación Transcripcional , Trasplante Heterólogo , Células Tumorales CultivadasRESUMEN
Endogenous lectins act as effectors of cellular activities such as growth regulation, migration, and adhesion. Following their immunohistochemical localization in our previous study (Saussez et al. in Histochem Cell Biol 123:29-41, 2005) we purified several galectins and used them as tools for monitoring accessible binding sites. Herein, we report the use of galectin histochemistry for the analysis of diethylstilbestrol (DES)-induced renal tumors in male Syrian hamster kidney (SHKT). Sections of normal kidney and DES-treated kidney were analyzed with biotinylated galectins-1, -3 (full-length and truncated), and -7. Accessible binding sites were detected, localization was predominantly extracellular and confined to medium-sized and large tumors. Monitoring the SHKT-derived HKT-1097 line, processed in vitro or as xenograft material, cytoplasmic and nuclear staining for galectins-1, -3, and -3tr could be observed. Adaptation of SHKT cells to long-term growth in culture is thus associated with emergence of this signal. Our data set illustrates the feasibility to complement immunohistochemical data by application of the tissue lectins as probes, and to detect regulation of galectin reactivity with differential characteristics within tumor progression in vivo and unique features of the tumor cell line in vitro and in vivo.
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Carcinógenos , Dietilestilbestrol , Galectinas/metabolismo , Neoplasias Renales/inducido químicamente , Neoplasias Renales/metabolismo , Animales , Sitios de Unión , Biotina/metabolismo , Línea Celular , Línea Celular Tumoral , Cricetinae , Técnica del Anticuerpo Fluorescente , Galectina 1/metabolismo , Galectina 3/metabolismo , Procesamiento de Imagen Asistido por Computador , Inmunohistoquímica , Neoplasias Renales/patología , Lectinas , Masculino , Mesocricetus , Ratones , Ratones Desnudos , Trasplante de Neoplasias , Trasplante HeterólogoRESUMEN
1. The renin-angiotensin system may be involved in the compensatory adaptations occurring after the reduction of renal mass and during the consecutive changes leading to chronic renal failure. We therefore investigated the regulation of angiotensin II receptors in two models of renal hypertrophy in the rat: hypertrophy following uninephrectomy (UNx) or subtotal nephrectomy (STNx). The level of angiotensin type 1 (AT1A-R and AT1B-R) and type 2 (AT2-R) receptor mRNA was quantified by competitive reverse transcription-polymerase chain reaction (RT-PCR) in specific renal zones and the intrarenal distribution of angiotensin II receptors was analysed by immunohistochemistry. 2. In the UNx rats, AT1-R mRNA expression was not modified in the cortex or in the inner stripe of the outer medulla of the residual kidney at any time after the surgery (1, 4 and 12 weeks). In contrast, AT1-R mRNA expression was significantly reduced in these zones in STNx rats (-33% and -40%, respectively). This downregulation was organ-specific, as AT1-R mRNA levels were not modified in the liver. The proportions of AT1-R subtype (AT1A and AT1B) mRNA were unchanged by UNx or STNx. Very low levels of AT2-R mRNA were found in the cortex of all groups. Immunostaining revealed a similar localization of AT1-R in mesangial cells, proximal tubule, basolateral membrane of thick ascending limb, in both models of hypertrophy. AT1-R labelling was also detected in the apical membrane of intercalated cells of cortical collecting ducts. 3. This differential mRNA expression of angiotensin II receptors during compensatory hypertrophy and renal injury suggests that the development of renal hypertrophy is independent of AT1-R and AT2-R gene expression levels.
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Regulación hacia Abajo/genética , Riñón/lesiones , Receptores de Angiotensina/genética , Animales , Peso Corporal , Creatina/sangre , Creatina/orina , Ingestión de Alimentos , Hipertrofia/genética , Hipertrofia/patología , Hipertrofia/fisiopatología , Inmunohistoquímica , Riñón/metabolismo , Riñón/patología , Corteza Renal/química , Corteza Renal/metabolismo , Corteza Renal/patología , Médula Renal/química , Médula Renal/metabolismo , Médula Renal/patología , Hígado/química , Hígado/metabolismo , Masculino , Nefrectomía , Tamaño de los Órganos , Ratas , Ratas Wistar , Receptor de Angiotensina Tipo 1/análisis , Receptor de Angiotensina Tipo 1/genética , Receptor de Angiotensina Tipo 2/análisis , Receptor de Angiotensina Tipo 2/genética , Receptores de Angiotensina/análisis , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Orina/químicaRESUMEN
E-selectin-targeted contrast enhancement of blood vessels in inflamed tissues was investigated with a new contrast agent, Gd-DTPA-B(sLe(x))A, which was recently obtained by grafting a synthetic mimetic of sialyl-Lewis(x), an E-selectin ligand, onto Gd-DTPA. The pharmacokinetics, biodistribution, and potential to image inflammation by MRI of this E-selectin-targeted contrast agent were evaluated. The inhibition (by 15-34%) produced by Gd-DTPA-B(sLe(x))A on Sialyl Le(x)-PAA-biotin binding to E-selectin confirmed the specific interaction of the new contrast agent with this adhesion molecule. Gd-DTPA-B(sLe(x))A was tested at a dose of 0.1 mmol/kg b.w. on mice and rats in a fulminant hepatitis model induced by the co-administration of D-galactosamine and E. coli lipopolysaccharide. A significant and prolonged contrast enhancement between blood vessels and liver parenchyma was obtained in pathological conditions, which attests to the specificity of the agent for E-selectin. The prolonged vascular residence (48.9 min in hepatitis vs. 29.8 min in healthy animals), as evidenced by the pharmacokinetic characterization, suggests that Gd-DTPA-B(sLe(x))A interacts with the specific receptors expressed during inflammation. The biodistribution of the compound indicates its retention in inflamed liver by both specific mechanisms and nonspecific accumulation due to the necrotic lesions. The same mechanisms are invoked to account for its retention in the spleen.