Asunto(s)

Insectos Vectores , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Thysanoptera , Tospovirus , Animales , Insectos Vectores/virología , Enfermedades de las Plantas/virología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/normas , Thysanoptera/virología , Tospovirus/genética

RESUMEN

Since the discovery of opioid receptor dimers their possible roles in opioid actions were intensively investigated. Here we suggest a mechanism that may involve the µ-δ opioid heterodimers. The exact role of δ opioid receptors in antinociception and in the development of opioid tolerance is still unclear. While receptor up-regulation can be observed during the development of opioid tolerance no µ receptor down-regulation could be detected within five days. In our present work we investigated how the selective δ opioid receptor agonists and antagonists influence the antinociceptive effect of the selective µ receptor agonist DAMGO in naïve and morphine-tolerant mice. We treated male NMRI mice with 200 µmol/kg subcutaneous (s.c.) morphine twice daily for three days. On the fourth day we measured the antinociceptive effect of DAMGO alone and combined with delta ligands: DPDPE, deltorphin II (agonists), TIPP and TICPψ (antagonists), respectively, administered intrathecally (i.t.) in mouse tail-flick test. In naive control mice none of the δ ligands caused significant changes in the antinociceptive action of DAMGO. The treatment with s.c. morphine resulted in approximately four-fold tolerance to i.t. DAMGO, i.e. the ED50 value of DAMGO was four times as high as in naive mice. 500 and 1000 pmol/mouse of the δ1 selective agonist DPDPE enhanced the tolerance to DAMGO while 1000 pmol/mouse of the δ2 selective agonist deltorphin II did not influence the degree of tolerance. However, both δ antagonists TIPP and TICPψ potentiated the antinociceptive effect of i.t. DAMGO, thus they restored the potency of DAMGO to the control level. The inhibitory action of DPDPE against the antinociceptive effect of DAMGO could be antagonized by TIPP and TICPψ. We hypothesize that during the development of morphine tolerance the formation of µÎ´ heterodimers may contribute to the spinal opioid tolerance. δ ligands may affect the dimer formation differently. Those, like DPDPE may facilitate the dimer formation hence inhibit the antinociceptive effect of DAMGO by causing virtual µ receptor down-regulation. Ligands that do not affect the dimer formation do not influence antinociception either but ligands with the presumed capability of disconnecting the dimers may decrease the spinal tolerance to DAMGO.

Asunto(s)

Analgésicos Opioides/farmacología , Tolerancia a Medicamentos/fisiología , Encefalina Ala(2)-MeFe(4)-Gli(5)/farmacología , Morfina/farmacología , Receptores Opioides delta/metabolismo , Médula Espinal/metabolismo , Animales , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Vías de Administración de Medicamentos , Esquema de Medicación , Interacciones Farmacológicas , Ligandos , Masculino , Ratones , Dimensión del Dolor/efectos de los fármacos , Somatostatina/análogos & derivados , Somatostatina/farmacología , Médula Espinal/efectos de los fármacos , Conducto Deferente/efectos de los fármacos , Conducto Deferente/patología

RESUMEN

Previous studies showed that opioid drugs-oxycodone-6-oxime and 14-methoxy-5-methyl-dihydromorphinone (14-methoxymetopon)-produced less respiratory depressive effect and slower rate of tolerance and dependence, respectively. It was also reported that morphine decreased the prodynorphin gene expression in the rat hippocampus, striatum and hypothalamus. In this study, we determined the prodynorphin gene expression and dynorphin levels in selected brain regions of opioid tolerant rats. We found that in the striatum morphine decreased, while oxycodone-6-oxime increased and 14-methoxymetopon did not alter the prodynorphin gene expression. In the nucleus accumbens, morphine and oxycodone-6-oxime did not change, while 14-methoxymetopon increased the prodynorphin gene expression. In the hippocampus both oxycodone-6-oxime and 14-methoxymetopon enhanced, whereas morphine did not alter the prodynorphin gene expression. In the rat striatum only oxycodone-6-oxime increased dynorphin levels significantly in accordance with the prodynorphin mRNA changes. In the hippocampus both opioid agonists increased the dynorphin levels significantly similarly to the augmented prodynorphin gene expression. In ventral tegmental area only 14-methoxymetopon increased dynorphin levels significantly. In nucleus accumbens and the temporal-parietal cortex the changes in the prodynorphin gene expression and the dynorphin levels did not correlate. Since the endogenous prodynorphin system may play a modulatory role in the development of opioid tolerance, the elevated supraspinal dynorphin levels appear to be partly responsible for the reduced degree of tolerance induced by the investigated opioids.

Asunto(s)

Encéfalo/efectos de los fármacos , Dinorfinas/efectos de los fármacos , Encefalinas/efectos de los fármacos , Derivados de la Morfina/administración & dosificación , Narcóticos/administración & dosificación , Oxicodona/administración & dosificación , Precursores de Proteínas/efectos de los fármacos , Animales , Northern Blotting , Tolerancia a Medicamentos/fisiología , Dinorfinas/biosíntesis , Encefalinas/biosíntesis , Encefalinas/genética , Expresión Génica/efectos de los fármacos , Masculino , Precursores de Proteínas/biosíntesis , Precursores de Proteínas/genética , ARN Mensajero/análisis , Radioinmunoensayo , Ratas , Ratas Sprague-Dawley , Factores de Tiempo

RESUMEN

UNLABELLED: This study was designed to create a reproducible model for experimental burn wound research in pigs. Previously, the thicker paraspinal skin has been used. We used the more human-like ventral skin to create burns of different depths. Contact burns were created to 11 pigs using a brass plate heated to 100 degrees C in boiling water. Different contact times were used to create burns of different depths. In pigs 1-6, the follow-up time was 72 h and in pigs 7-11 24 h. Burn depth was determined by histology. Histologically, samples were classified into five anatomical layers: epidermis, upper one-third of the dermis, middle third of the dermis, deepest third of the dermis and subcutaneous fat. The location of both thromboses and burn marks were evaluated, respectively. The 1 s contact time lead to a superficial thermal injury, 3 s to a partial thickness and 9 s to a full thickness injury. A progression of burn depth was found until 48 h post-injury. The intra-observer correlation after repeated histological analyses of burn depths by the same histopathologist and the repeatability of burn depth creation yielded kappa coefficients 0.83 and 0.92, respectively. CONCLUSION: a reproducible burn model for further research purposes was obtained.

Asunto(s)

Quemaduras/patología , Modelos Animales de Enfermedad , Animales , Superficie Corporal , Temperatura Corporal , Peso Corporal , Quemaduras/etiología , Quemaduras/fisiopatología , Progresión de la Enfermedad , Femenino , Hemodinámica , Piel/patología , Porcinos

RESUMEN

OBJECTIVE: To evaluate the effects of intra-articular (IA) injections of bufexamac in horses, focusing particularly on the effects of bufexamac on articular cartilage. ANIMALS: 20 Standardbreds. PROCEDURE: Horses were randomly allocated into 4 groups consisting of 5 horses each, and 20, 60, or 100 mg of bufexamac or 1 ml of sterile saline (0.9% NaCl) solution (control) was injected into 1 intercarpal joint at weekly intervals for 6 treatments (days 0, 7, 14, 21, 28, and 35). Clinical signs and results of hematologic, serum biochemical, and synovial fluid (SF) analyses and radiography were used to evaluate treatment effects. On day 49, all horses were euthanatized; gross necropsy and histologic examinations of internal organs and articular tissues were performed. Glycosaminoglycan concentration of the articular cartilage was evaluated in safranin O-stained sections by use of a semiquantitative microspectrophotometric method. RESULTS: No systemic signs were observed. Temporary mild to moderate heat and effusion were the only clinical signs observed in a number of joints after IA injections and more often only in the 100 mg group, compared with controls. The 100 mg dose resulted in significant increases in SF WBC counts, with relative neutrophilia and SF total protein concentration 24 hours after injection (day 1). No lesions suggestive of toxic effects were detected at necropsy or on histologic examination. No changes in articular cartilage glycosaminoglycan concentration were detected. CONCLUSIONS AND CLINICAL RELEVANCE: Six injections of 20, 60, or 100 mg of bufexamac at weekly intervals did not cause any untoward systemic or local effects. These data suggest that bufexamac is a safe nonsteroidal anti-inflammatory drug for IA administration in horses.

Asunto(s)

Antiinflamatorios no Esteroideos/farmacología , Bufexamac/farmacología , Cartílago Articular/efectos de los fármacos , Caballos/metabolismo , Animales , Antiinflamatorios no Esteroideos/administración & dosificación , Bufexamac/administración & dosificación , Cartílago Articular/diagnóstico por imagen , Cartílago Articular/patología , Relación Dosis-Respuesta a Droga , Femenino , Miembro Anterior/diagnóstico por imagen , Miembro Anterior/efectos de los fármacos , Glicosaminoglicanos/análisis , Histocitoquímica/veterinaria , Inyecciones Intraarticulares , Masculino , Radiografía , Distribución Aleatoria , Suspensiones , Líquido Sinovial/química

RESUMEN

OBJECTIVE: To compare responses of the collagen network and glycosaminoglycans (GAGs) of articular cartilage to physiological type of joint loading in young growing and adult mature guinea-pigs. DESIGN: 10- and 44-week-old guinea-pigs were accustomed to treadmill running for 3 weeks. Thereafter the animals ran 2500 m/day, 5 days a week, for 15 weeks. Articular cartilage specimens from knee joints were collected at 28 and 62 weeks. Osteoarthritis (OA) prevalence and severity was evaluated by aid of light microscopy. The degree of collagen fibril network organization and content was analyzed with quantitative polarized light microscopy. The local concentration of GAGs was determined from cartilage sections with digital densitometry after safranin-O staining. RESULTS: In the young guinea-pigs, running increased up to 24% the optical retardation of polarized light by collagen in the superficial articular cartilage of femur, indicating either a higher degree of fibril assembly and organization or increased amount of collagen, or both. In contrast, in the adult mature animals the optical retardation decreased almost 50% after joint loading (P< 0.01-0.001). Running did not increase cartilage fibrillation. Significant changes in GAG content of cartilage were not found either in the young or adult mature runners. CONCLUSIONS: Increased birefringence of the superficial articular cartilage after joint loading in young guinea-pigs can be interpreted to be a sign of improved and decreased birefringence in older animals a sign of worsened property of the collagen network. It can be suggested therefore that joint loading strengthened the collagen network in the young runners. It can be hypothesized further that with time the inferior property of the collagen network predisposes the older runners to earlier OA than in controls.

Asunto(s)

Envejecimiento/fisiología , Cartílago Articular/fisiología , Colágeno/fisiología , Articulación de la Rodilla/fisiología , Animales , Birrefringencia , Distribución de Chi-Cuadrado , Densitometría , Femenino , Glicosaminoglicanos/fisiología , Cobayas , Microscopía de Polarización , Osteoartritis de la Rodilla/patología , Estadísticas no Paramétricas , Soporte de Peso/fisiología

RESUMEN

CD44 was detected with an antibody recognizing all forms of CD44 (CD44 standard) and others specific for its v3 and v6 variant isoforms; their prognostic value was evaluated in 213 patients with differentiated thyroid carcinoma (DTC). The staining patterns of CD44 standard (s) and CD44v6 in tumour tissue were quite similar, 176 cases (83%) being highly positive for CD44s and 153 cases (72%) for CD44v6. Only 18 (9%) tumours showed high expression of CD44v3. Papillary carcinomas were significantly more often high expressors of CD44s and CD44v6 than follicular carcinomas (p<0.001 for both). Age older than 60 years, distant metastases, and advanced pTNM stage were related to loss of expression of CD44s (p<0.001, p=0.021, and p=0.003, respectively). Tumour recurrence and cancer-related mortality were related to the reduced level of CD44s (p=0.049 and p=0.042). CD44v3 did not associate with any of the clinicopathological factors. In univariate analysis, CD44s was the only significant prognostic factor for disease-free survival (p=0.0488). In multivariate analysis, CD44s and thyroglobulin level were significant prognostic factors for disease-free survival (p=0.040 and p<0.001, respectively). The reduced level of CD44s in DTC patients seems to be an independent prognostic factor for unfavourable disease outcome.

Asunto(s)

Carcinoma Papilar Folicular/inmunología , Receptores de Hialuranos/inmunología , Recurrencia Local de Neoplasia/inmunología , Neoplasias de la Tiroides/inmunología , Adolescente , Adulto , Factores de Edad , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor , Carcinoma Papilar Folicular/secundario , Niño , Supervivencia sin Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Análisis Multivariante , Metástasis de la Neoplasia/inmunología , Pronóstico , Tiroglobulina/análisis , Neoplasias de la Tiroides/patología , Neoplasias de la Tiroides/cirugía , Tiroidectomía/métodos

RESUMEN

Catenins (alpha, ss, and gamma) are a group of intracellular cell adhesion molecules that unite cytoskeleton with extracellular adhesion system. Abnormal expression of these molecules may have prognostic relevance in various carcinomas, including differentiated thyroid carcinoma (DTC). We have, therefore, evaluated the prognostic value of alpha-, ss-, and gamma-catenins along with traditional risk factors in 206 consecutive DTC patients by immunohistochemistry. Papillary carcinomas showed normal staining pattern for alpha-, ss-, and gamma-catenins in 124 (60%), 136 (67%), and 94 (46%) cases, respectively. Follicular carcinomas expressed alpha-, ss-, and gamma-catenins normally in 16 (48%), 18 (55%), and 8 (32%) cases, respectively. Follicular type of tumor showed more often reduced staining for all catenins than papillary carcinoma (P: = 0.009, P: = 0.004, and P: = 0.002, respectively). Age (>60 yr) and pTNM-stage were related to reduced alpha- and ss-catenin expression levels (P: = 0.027 and P: = 0.026, respectively) and larger size of the tumor to reduced ss- and gamma-catenin expressions (P: = 0.039 and P: = 0.007, respectively). Nodal metastases at the time of primary treatment related to reduced alpha-catenin expression and distal metastases to reduced ss- and gamma-catenin staining signals (P: = 0.022, P: = 0.014, and P: = 0.039, respectively). Reduced alpha-catenin associated with tumor recurrence (P: = 0.002) and reduced ss-catenin with cancer-related mortality (P: = 0.005). The multivariate analysis for recurrence-free survival showed that alpha-catenin and serum thyroglobulin level 1 yr after primary treatment were prognostic of recurrent disease (hazards ratio, 3.42, P: = 0.022; and hazards ratio, 10.03, P: = 0.0001). In addition, alpha-catenin retained its prognostic significance in low-stage patients (P: = 0.0151). We propose that the evaluation of alpha-catenin expression by immunohistochemistry in DTC patients has prognostic value in addition to that obtained by traditional prognostic factors.

Asunto(s)

Cadherinas/metabolismo , Carcinoma Papilar/metabolismo , Proteínas del Citoesqueleto/metabolismo , Neoplasias de la Tiroides/metabolismo , Transactivadores , Anciano , Carcinoma Papilar/patología , Desmoplaquinas , Femenino , Estudios de Seguimiento , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Pronóstico , Tiroglobulina/metabolismo , Neoplasias de la Tiroides/patología , Tiroidectomía , Tirotropina/sangre , alfa Catenina , beta Catenina , gamma Catenina

RESUMEN

Results obtained by the indirect zonal isotropic uniform random (IUR) estimation were compared with those obtained by the direct point and interception counting methods on vertical (VS) or IUR sections in a stereological study of bovine articular cartilage collagen fibrils at the ultrastructural level. Besides comparisons between the direct and indirect estimations (direct IUR vs indirect IUR estimations) and between different sampling methods (VS vs IUR sampling), simultaneous comparison of the 2 issues took place (direct VS vs indirect IUR estimation). Using the direct VS method, articular cartilage superficial zone collagen volume fraction (Vv 41%) was 67% and fibril surface density (S(v) 0.030 nm2/nm3) 15% higher (P < 0.05) than values obtained by the indirect IUR method (V(v) 25 % and Sv 0.026 nm2/nm3). The same was observed when the direct IUR method was used: collagen volume fraction (Vv 40 %) was 63 % and fibril surface density (Sv 0.032 nm2/nm3) 21 % higher (P < 0.05) than those obtained by the indirect IUR technique. Similarly, in the deep zone of articular cartilage direct VS and direct IUR methods gave 50 and 55% higher (P < 0.05) collagen fibril volume fractions (Vv 43 and 44% vs 29%) and the direct IUR method 25% higher (P < 0.05) fibril surface density values (Sv) 0.025 vs 0.020 nm2/nm3) than the indirect IUR estimation. On theoretical grounds, scrutiny calculations, as well as earlier reports, it is concluded that the direct VS and direct IUR methods systematically overestimated the Vv and Sv of collagen fibrils. This bias was due to the overprojection which derives from the high section thickness in relation to collagen fibril diameter. On the other hand, factors that during estimation tend to underestimate Vv and Sv, such as profile overlapping and truncation ('fuzzy' profiles), seemed to cause less bias. As length density Lv and collagen fibril diameter are minimally biased by the high relative section thickness, the indirect IUR method, based on utilisation of these estimates, is here regarded as representing a 'gold standard'. The sensitivity of these 3 methods was also tested with cartilage from an in vitro loading experiment which caused tissue compression. In the superficial zone of articular cartilage Vv and Sv of collagen fibrils increased (P < 0.05). This difference in the stereological estimates was only detected by the indirect IUR estimation but not by the direct VS or direct IUR methods. This indicated that the indirect IUR estimation was more sensitive than the direct VS or direct IUR estimations. On the basis of these observations, the indirect zonal IUR estimation can be regarded as the technique of choice in the electron microscopic stereology of cartilage collagen.

Asunto(s)

Cartílago Articular/ultraestructura , Colágeno/ultraestructura , Microfibrillas/ultraestructura , Animales , Fenómenos Biomecánicos , Bovinos , Articulación de la Rodilla , Microscopía Electrónica , Estadísticas no Paramétricas

RESUMEN

INTRODUCTION: The articular cartilage collagen network and proteoglycans are subject to changes in deteriorating joint diseases. In this study, we exposed articular cartilage plugs to cyclic loading and investigated the properties of collagen network and proteoglycans in different zones of the articular cartilage. METHODS: Articular cartilage full-depth plugs were exposed in vitro to 4.1 MPa cyclic (0.5 Hz) loading for 1 to 20 hr and investigated using quantitative microscopic methods (i.e., polarized light microscopy, microspectrophotometry, and autoradiography). RESULTS: The loading caused packing or condensation of the tissue. In histological sections, the height of uncalcified articular cartilage decreased by an average of 12.8% (range, 4 to 19.7%). Loading increased the birefringence of collagen in the superficial cartilage (P < 0.05), with thickening of the zone up to 41.4% at 20 hr. The thickness of the intermediate zone increased also (22% at 1 hr and 434% at 20 hr). Concomitantly, the birefringence (P < 0.05) and the thickness of the deep zone decreased (18.5 to 27.8%). Loading for 4 hr increased the 35S-sulphate incorporation of the cartilage explants by an average of 67% (P < 0.05). The increase was most significant in the deep cartilage. A simultaneous increase was observed in the proteoglycan concentration of the cartilage; the staining intensity with safranin-O increased by 8.8% (P < 0.05). After 8 hr loading, this stimulation decreased; at 20 hr, loading caused a clear inhibitory effect on proteoglycan synthesis in the superficial zone. DISCUSSION: According to these results, the chosen loading regimen increased the thickness and collagen orientation in the superficial zone. In contrast, the thickness and birefringence in the deep cartilage were reduced. The proteoglycan metabolism of chondrocytes was first stimulated deep in the cartilage, but as the loading continued, the effect proved to be inhibitory (especially in the superficial part of uncalcified cartilage).

Asunto(s)

Cartílago Articular/metabolismo , Colágeno/metabolismo , Proteoglicanos/biosíntesis , Soporte de Peso/fisiología , Animales , Autorradiografía , Birrefringencia , Cartílago Articular/química , Bovinos , Colágeno/química , Glicosaminoglicanos/análisis , Placa de Crecimiento/anatomía & histología , Placa de Crecimiento/fisiología , Técnicas In Vitro , Microscopía de Polarización , Microespectrofotometría

RESUMEN

A great many studies in the literature describe the variety of ways to provide physical training to small laboratory animals. Forced running on a treadmill or swimming training require considerable effort from the researcher, but permit the investigator to control the amount of exercise. Another option is to provide animals with access to running wheels, which they can voluntarily operate. Wheels can be used to investigate the running behavior of the animals. Wheel motion usually has been detected with magnets and microswitches and has been recorded using strip chart recorders or electromechanical counters. Computers also have been used in recording, but the measured parameters usually have been able to define only the total distance run in a fixed period. We designed a system, using running wheels, that can be used for long periods, up to years, for training a large number of laboratory mice simultaneously (maximal n = 96). The running parameters estimated by our system include running distance, speed, and time. Cumulative estimates of the running parameters can be produced for any period, as short as 1 sec. It is also easy to perform statistical analyses on the data. Using the system, we investigated the running behavior of 21 young C57BL/6 male mice. After the fast growth period, until 8 weeks of age, the mice ran 4 to 5 km/day at an average speed of 23 m/min, and spent 3 h running each day. This took place during the hours of darkness.

Asunto(s)

Animales de Laboratorio/fisiología , Computadores , Condicionamiento Físico Animal , Esfuerzo Físico/fisiología , Animales , Conducta Animal , Masculino , Ratones , Ratones Endogámicos C57BL , Carrera

RESUMEN

To establish an optimal method for analysis of the collagen structures from unstained tissue sections, a computerized image analysis system using a charge coupled device camera coupled to a polarizing light microscope was used. Retardation values of birefringence, which are proportional to the content and fibril orientation of collagen in the extracellular matrix of articular cartilage, were determined from sections prepared in different ways. In the superficial zone of articular cartilage, the highest retardation values were recorded from sections cut parallel to the so-called split lines indicating the anisotropic arrangement of collagen. Complete digestion of glycosaminoglycans reduced the retardation value by approximately 6.0%, suggesting a minor, but not insignificant, contribution of glycosaminoglycans to the birefringence of the matrix. The use of a mounting medium with a refractive index close to that of the collagen (e.g. DPX) increased the specificity of the method, since the optical anisotropy of collagen derives predominantly from the intrinsic (structural) birefringence. In conclusion, analysis of unstained sections after careful removal of paraffin and glycosaminoglycans from the tissues provides a sensitive and rapid quantitative assessment of oriented collagen structures in articular cartilage.

Asunto(s)

Cartílago Articular/química , Colágeno/química , Técnicas de Preparación Histocitológica , Animales , Birrefringencia , Bovinos , Miembro Posterior , Procesamiento de Imagen Asistido por Computador , Articulaciones , Microscopía de Polarización

RESUMEN

Selected commonly used cationic dyes, viz. Thionin, Safranin O, Toluidine Blue O, Dimethylmethylene Blue, Cuprolinic Blue, Cupromeronic Blue, N,N'-Diethylpseudoisocyanine, and a modified PAS-method, and staining methods with a variety of alternative procedures, e.g., variation of pH, use of the critical electrolyte concentration method, and blocking reactions (methylation-saponification, carboxymethylation), were tested to select optimal staining procedures for the semiquantitative histochemical estimation of glycosaminoglycans by microspectrophotometry in sections of articular cartilage. The methods were carried out on 3 microns-thick paraffin and 1 microns-thick glycolmethacrylate sections of bovine articular cartilage. The staining intensity of the sections was measured from spots 25 microns apart using a Leitz MPV 3 microspectrophotometer, starting at the surface of the cartilage and ending up at the tidemark. The result was compared with the fixed-charge density graph determined from the adjacent articular cartilage. Of the dyes tested, Thionin and Safranin O proved to be excellent cationic dyes for the histochemical quantification of cartilage matrix proteoglycans, since the staining intensity curves showed a linear correlation (r = 0.900-0.995) with the fixed charge density curves from the adjacent cartilage. Also, the stain distribution was consistently uniform across the sections. In 1 microns-thick glycolmethacrylate sections, the Safranin O staining gradient showed almost perfect identity with the fixed-charge density curve. Cuprolinic Blue and Cupromeronic Blue combined with the critical electrolyte concentration technique were also useful for the microspectrophotometric assays of glycosaminoglycans, but the presence of metachromasia should be checked prior to the measurements. The reliability of blocking procedures for quantitative histochemical work was not convincing.

Asunto(s)

Cartílago Articular/metabolismo , Cationes , Colorantes/química , Glicosaminoglicanos/análisis , Microespectrofotometría/métodos , Animales , Bovinos , Electrólitos/química , Concentración de Iones de Hidrógeno , Cloruro de Magnesio/química , Coloración y Etiquetado

RESUMEN

The ability of Safranin O, added to fixation and decalcification solutions, to prevent the escape of glycosaminoglycans (GAGs) from small cartilage tissue blocks during histological processing of cartilage has been studied. GAGs in the fixatives and decalcifying solutions used and those remaining in the 1 mm3 cubes of cartilage were assayed biochemically. The quantity of GAGs remaining in the cartilage cubes were determined from Safranin O-stained sections using videomicroscopy or microspectrophotometry. A quantity (10.6%) of GAGs were lost during a conventional 4% buffered formaldehyde fixation (48 h) and a subsequent decalcification in 10% EDTA (12 days) at 4 degrees C. Roughly one-quarter of the total GAG loss occurred during the 48 h fixation, and three-quarters during the 12 days of decalcification. Inclusion of 4% formaldehyde in the decalcification fluid decreased the loss of GAGs to 6.2%. The presence of 0.5% Safranin O in the fixative reduced this loss to 3.4%. When 0.5% Safranin O was included in the fixative and 4% formaldehyde in the decalcification solution, Safranin O staining of the histological sections increased on average by 13.5%. After fixation in the presence of 0.5% Safranin O, there was no difference in the staining intensities when decalcification was carried out in the presence of either Safranin O or formaldehyde, or both. It took 24 h for Safranin O to penetrate into the deep zone of articular cartilage, warranting a fixation period of at least this long. In conclusion, the addition of Safranin O to the fixative and either Safranin O or formaldehyde in the following decalcification fluid, markedly reduces the loss of GAGs from small articular cartilage explants during histological processing. However, for immunohistochemical studies, Safranin O cannot be included in the processing solutions, because it may interfere.

Asunto(s)

Cartílago Articular/efectos de los fármacos , Colorantes/farmacología , Glicosaminoglicanos/análisis , Fenazinas/farmacología , Animales , Cartílago Articular/metabolismo , Cartílago Articular/patología , Bovinos , Ácido Edético/farmacología , Fijadores/farmacología , Formaldehído/farmacología , Rodilla/patología

RESUMEN

Homologous recombination in embryonic stem cells was used to prepare transgenic mice with an inactivated Col2a1 gene for collagen II, the major protein component of the extracellular matrix of cartilage. Heterozygous mice had a minimal phenotype. Homozygous mice developed into fetuses that were delivered vaginally but died either just before or shortly after birth. The cartilage in the mice consisted of highly disorganized chondrocytes with a complete lack of extracellular fibrils discernible by electron microscopy. There was no endochondrial bone or epiphyseal growth plate in long bones. However, many skeletal structures such as the cranium and ribs were normally developed and mineralized. The results demonstrate that a well-organized cartilage matrix is required as a primary tissue for development of some components of the vertebrate skeleton, but it is not essential for others.

Asunto(s)

Huesos/fisiopatología , Colágeno/genética , Osteogénesis/fisiología , Animales , Animales Recién Nacidos , Secuencia de Bases , Colágeno/metabolismo , Cartilla de ADN , Femenino , Expresión Génica , Marcación de Gen , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Datos de Secuencia Molecular , Fenotipo

RESUMEN

BACKGROUND: Inbred C57BL male mice express a high incidence of spontaneous osteoarthritis of the knee joint at the age of 18 months. We used this strain of mice to find out the effects of life-long, moderate running exercise on the health of articular cartilage and the incidence of osteoarthritis. METHODS: Male mice (294) were divided into controls and runners. The runners were trained daily between 2 and 18 months of age. The speed was 13.3 m/min and the distance on a flatbelt treadmill was 1,000 m/day. The mice were sacrificed at the ages of 2, 6, 10, 14, and 18 months. The knee joints were sectioned in frontal direction and the osteoarthritic changes were graded using a conventional light microscope. The reproducibility of the grading method was tested by calculating the extended kappa-coefficient for the results of six researchers. RESULTS: The incidence of osteoarthritis at the age of 18 months increased from 72% in controls to 88% in runners in the medial tibial condyles (P < 0.05), and from 80 to 96% in the lateral tibial condyles (P < 0.001). The incidence of the most severe osteoarthritic changes rose from 16% in controls to 36% in runners in the medial tibial condyles, and from 4 to 36% in the lateral tibial condyles. CONCLUSION: According to our results, the moderate, long-lasting running exercise accelerates the development of osteoarthritis in the knee joints of C57BL mice.

Asunto(s)

Articulación de la Rodilla , Osteoartritis/etiología , Condicionamiento Físico Animal/efectos adversos , Animales , Cartílago Articular/patología , Modelos Animales de Enfermedad , Articulación de la Rodilla/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Osteoartritis/patología , Esfuerzo Físico , Carrera

RESUMEN

Studies were carried out on a line of transgenic mice that expressed an internally deleted COL2A1 gene and developed a phenotype resembling human chondrodysplasias (Vandenberg et al. 1991. Proc. Natl. Acad. Sci. USA. 88:7640-7644. Marked differences in phenotype were observed with propagation of the mutated gene in an inbred strain of mice in that approximately 15% of the transgenic mice had a cleft palate and a lethal phenotype, whereas the remaining mice were difficult to distinguish from normal littermates. 1-d- and 3-mo-old transgenic mice that were viable showed microscopic signs of chondrodysplasia with reduced amounts of collagen fibrils in the cartilage matrix, dilatation of the rough surfaced endoplasmic reticulum in the chondrocytes, and decrease of optical path difference in polarized light microscopy. The transgenic mice also showed signs of disturbed growth as evidenced by lower body weight, lower length and weight of the femur, decreased bone collagen, decreased bone mineral, and decreased resistance of bone to breakage. Comparisons of mice ranging in age from 1 d to 15 mo demonstrated that there was decreasing evidence of a chondrodysplasia as the mice grew older. Instead, the most striking feature in the 15-mo-old mice were degenerative changes of articular cartilage similar to osteoarthritis.

Asunto(s)

Cartílago/ultraestructura , Eliminación de Gen , Procolágeno/genética , Envejecimiento/fisiología , Animales , Secuencia de Bases , Peso Corporal , Desarrollo Óseo , Huesos/metabolismo , Cartílago/crecimiento & desarrollo , Fisura del Paladar/genética , Colágeno/biosíntesis , Colágeno/metabolismo , Cósmidos , Exones , Matriz Extracelular/ultraestructura , Femenino , Genes Letales , Placa de Crecimiento/ultraestructura , Humanos , Masculino , Ratones , Ratones Endogámicos , Ratones Transgénicos , Microscopía Electrónica , Datos de Secuencia Molecular , Oligodesoxirribonucleótidos , Linaje , Reacción en Cadena de la Polimerasa , Valores de Referencia , Mapeo Restrictivo , Factores Sexuales

RESUMEN

A salmonella panama outbreak was observed in the months May through July, 1979, in the newborn and intensive wards of a paediatric hospital. The isolates proved to be multiresistant to antibiotics. To clarify the presumed R-plasmid nature of the multiresistance, the transfer of the resistance to five antibiotics, viz. ampicillin, chloramphenicol, streptomycin, kanamycin and tetracycline, was studied. Escherichia cole K12 Nalr, the strain used as recipient, acquired resistance simultaneously to four antibiotics, viz. ampicillin, chloramphenicol, streptomycin and kanamycin. The minimum inhibitory concentration for the transconjugants agreed well with that of the original S. panama strain. Resistant and multiresistant E. coli. strains not belonging to serogroups associated with infantile enteritis were isolated from the intestinal flora of several patients and symptomless carriers. Elimination of the transmissible multiresistance was observed in 0.15-1.1% of the strains.

Asunto(s)

Enfermedades del Recién Nacido/microbiología , Factores R , Infecciones por Salmonella/microbiología , Salmonella/efectos de los fármacos , Antibacterianos/farmacología , Conjugación Genética , Brotes de Enfermedades , Escherichia coli/genética , Humanos , Hungría , Recién Nacido , Salmonella/genética , Salmonella/aislamiento & purificación , Infecciones por Salmonella/epidemiología

Asunto(s)

Antifúngicos/uso terapéutico , Administración Tópica , Candidiasis Cutánea/tratamiento farmacológico , Combinación de Medicamentos , Evaluación de Medicamentos , Humanos , Miconazol/uso terapéutico , Pomadas , Prednisolona/uso terapéutico , Pruebas Cutáneas

Asunto(s)

Imidazoles/uso terapéutico , Miconazol/uso terapéutico , Prednisolona/análogos & derivados , Prednisolona/uso terapéutico , Adulto , Ensayos Clínicos como Asunto , Dermatomicosis/tratamiento farmacológico , Dermatomicosis/microbiología , Método Doble Ciego , Combinación de Medicamentos , Eccema/tratamiento farmacológico , Eccema/microbiología , Humanos , Intertrigo/tratamiento farmacológico , Intertrigo/microbiología , Pomadas