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AbtsractThe emergence and rapid dissemination worldwide of a novel Coronavirus (SARS-CoV-2) results in decrease of swabs availability for clinical samples collection, as well as, reagents for RT-qPCR diagnostic kits considered a confirmatory test for COVID-19 infection. This scenario, showed the requirement of improve de diagnostic capacity, so the aim of this study were to verify the possibility of reducing the reaction volume of RT-qPCR and to test cotton swabs as alternative for sample collection. RT-qPCR volumes and RNA sample concentration were optimized without affecting the sensitivity of assays, using both probe-based and intercalation dyes methods. Although rayon swabs showed better performance, cotton swabs could be used as alternative type for clinical sample collection. COVID-19 laboratory diagnosis is important to isolate and restrict the dissemination of virus, so seek for alternatives to decrease the coast of assays improve the control of disease.
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OBJECTIVE: To evaluate the biomechanical properties of single 3.5 mm broad dynamic compression plate (DCP) and double 3.5 mm String-of-Pearls (SOP) plate constructs in single-cycle bending and torsion. We hypothesized that the double SOP construct would outperform the broad DCP in both bending and torsional testing. METHODS: Broad DCP plates and double 3.5 mm SOP plates were secured to a previously validated bone model in an effort to simulate bridging osteosynthesis. Constructs were tested in both four-point bending and torsional testing. RESULTS: The double SOP constructs had significantly greater bending stiffness, bending strength, bending structural stiffness, and torsional stiffness when compared to the broad DCP constructs. The single broad DCP constructs had significantly higher yield torque and yield angles during torsional testing. CLINICAL RELEVANCE: Although the in vitro mechanical performance of the double SOP construct was significantly greater than the single broad DCP constructs under bending loads, the actual differences were small. Various patient, fracture, and implant factors must be considered when choosing an appropriate implant for fracture fixation.
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Placas Óseas , Fenómenos Biomecánicos , Placas Óseas/normas , Tornillos Óseos , Fracturas Óseas/cirugía , Humanos , Docilidad , Falla de Prótesis , Torsión MecánicaRESUMEN
OBJECTIVE: To compare the in vitro biomechanical performance of two String-of-Pearls (SOP) plate constructs in a canine distal humeral metaphyseal gap model. METHODS: Seven pairs of canine cadaveric humeri, including the elbow joints, were prepared. One group consisted of a unilateral medially placed SOP plate with bicortical screws (UNI). The second group consisted of bilateral caudo-medial and caudo-lateral SOP plates applied with monocortical screws (BI). A 2 cm ostectomy was performed immediately proximal to the supratrochlear foramen. Constructs were tested in torsion and axial compression. RESULTS: The UNI constructs had significantly lower stiffness in torsion and axial compression than the BI group. However, UNI constructs had a significantly higher ultimate strength than BI constructs. All UNI constructs failed by bending of the transcondylar screw and SOP plate. All BI constructs failed by axial pullout of the distal most screws. CLINICAL SIGNIFICANCE: In stabilizing canine supracondylar humeral fractures as modeled here, both the UNI model and the BI model demonstrated biomechanical advantages. The incorporation of a transcondylar screw through the medial plate appears to be beneficial to construct strength.
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Placas Óseas/veterinaria , Perros , Miembro Anterior , Ensayo de Materiales/veterinaria , Animales , Fenómenos Biomecánicos , CadáverRESUMEN
REASONS FOR PERFORMING STUDY: Hereditary equine regional dermal asthenia (HERDA) is an autosomal recessive disorder of Quarter Horses characterised by skin fragility. Horses with HERDA have a missense mutation in peptidyl-prolyl cis-trans isomerase B (PPIB), which encodes cyclophilin B and alters folding and post translational modifications of fibrillar collagen. OBJECTIVES: The study aimed to test the hypothesis that tendons, ligaments and great vessels, which, like skin, are rich in fibrillar collagen, will also have abnormal biomechanical properties in horses with HERDA. STUDY DESIGN: Ex vivo biomechanical study comparing horses with and without a diagnosis of HERDA. METHODS: Forelimb suspensory ligament, superficial and deep digital flexor tendons; withers, forelimb and abdominal skin; the main pulmonary artery and the aortic arch were harvested from 6 horses with HERDA and 6 control horses without the HERDA allele. Tissues were distracted to failure. Tensile strength (TS), elastic modulus (EM) and energy to failure (ETF) were compared. RESULTS: Horses with HERDA had significantly lower TS and EM in tendinoligamentous tissues and great vessels, respectively. The TS, EM and ETF were significantly lower in skin from horses with HERDA. Differences in TS and ETF were more extreme at the withers than at the forelimb or abdomen. CONCLUSIONS: Tendinoligamentous tissue, great vessels and skin are significantly weaker in horses with HERDA than in horses lacking the PPIB mutation, substantiating that diverse tissues with high fibrillar collagen content are abnormal in HERDA and that the HERDA phenotype is not limited to the integument.
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Enfermedades de los Caballos/fisiopatología , Anomalías Cutáneas/veterinaria , Enfermedades Cutáneas Genéticas/veterinaria , Piel/patología , Animales , Fenómenos Biomecánicos , Enfermedades de los Caballos/genética , Caballos , Anomalías Cutáneas/genética , Anomalías Cutáneas/fisiopatología , Enfermedades Cutáneas Genéticas/genética , Resistencia a la TracciónRESUMEN
The in vitro biomechanical properties of three methods for internal fixation of femoral neck fractures were evaluated. Fifty cadaveric femura from Beagle dogs were used. Ten intact femora served as controls. In 40 femura, an osteotomy of the femoral neck was performed to simulate a transverse fracture. With the remaining 30 femura, three repair methods (two medium Orthofix pins, a 2.7 mm cortical bone screw placed in lag fashion and an anti- rotational Kirschner wire, or three divergent 1.1 mm Kirschner wires) were used to stabilize the osteotomies, and 10 osteotomies were stabilised per repair method. These 30 femura where then subject to monotonic loading to failure. Construct stiffness and load to failure were measured. In the remaining 10 femura, pressure sensitive film was placed at the osteotomy site prior to stabilization with either two Orthofix pins (n = 5) or a screw placed in lag fashion (n = 5) to determine the compressive pressure (MPa), compressive force (KN) and area of compression (cm²). There was no significant difference in the stiffness or load to failure for the three repair methods evaluated. There was no significant difference in the compressive pressure, compressive force or area of compression in osteotomies stabilized with Orthofix pins and 2.7 mm bone screws.
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Perros/lesiones , Fracturas del Cuello Femoral/veterinaria , Fijación Interna de Fracturas/veterinaria , Fijadores Internos/veterinaria , Osteotomía/veterinaria , Animales , Fenómenos Biomecánicos , Cadáver , Perros/fisiología , Perros/cirugía , Fracturas del Cuello Femoral/cirugía , Fijación Interna de Fracturas/métodos , Osteotomía/métodos , Distribución AleatoriaRESUMEN
OBJECTIVES: To evaluate the effect of tibial tuberosity advancement (TTA) and meniscal release on cranial-caudal and axial rotational displacement during early, middle and late stance phases in the canine cranial cruciate ligament- (CCL) deficient stifle. STUDY DESIGN: In vitro biomechanical study. METHODS: Eighteen pelvic limbs were evaluated for the effects of TTA on cranial-caudal displacement and axial rotation under a load equivalent to 30% bodyweight, and under the following treatment conditions: normal (intact CCL), CCL deficient, TTA-treated (CCL deficient + TTA), and meniscal release (TTA treated + meniscal release). The limbs were evaluated in the early, middle, and late stance phases using electromagnetic tracking sensors to determine cranial tibial displacement and tibial rotation relative to the femur. RESULTS: Transection of the CCL resulted in significant cranial tibial displacement during early, middle, and late stance (p < 0.0001) and significant internal rotation during early (p = 0.049) and middle stance (p = 0.0006). Performance of TTA successfully eliminated cranial tibial displacement in early, middle, and late stance (p <0.0001); however, the TTA was unsuccessful in normalizing axial rotation in middle stance (p = 0.030). Meniscal release had no effect on cranial-caudal or rotational displacement when performed in conjunction with the TTA. CLINICAL SIGNIFICANCE: Tibial tuberosity advancement effectively eliminates cranial tibial displacement during early, middle and late stance; however, TTA failed to provide rotational stability in mid-stance.
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Lesiones del Ligamento Cruzado Anterior , Perros , Osteotomía/veterinaria , Tibia/cirugía , Animales , Ligamento Cruzado Anterior/patología , Fenómenos Biomecánicos , Cadáver , Osteotomía/métodos , Rodilla de Cuadrúpedos/fisiología , Rodilla de Cuadrúpedos/cirugíaRESUMEN
OBJECTIVE: To describe the use of a cryogenic clamp of novel design for tensile strength testing of tendinous and ligamentous tissues with inherently high tensile strength. METHODS: Inexpensive, easily machined steel clamps were manufactured to facilitate rapid insertion into a standard wedge-screw grip apparatus installed on a testing system with a control system attached. The deep digital flexor tendon (DDFT) of six horses was trimmed to a uniform dumbbell shape and secured in clamps using partial submersion in liquid nitrogen for approximately 45 seconds and immediately tested. Approximate time between removal from liquid nitrogen and failure of tendon was four minutes. RESULTS: Failure was achieved in all tendons tested in a region approximating a midpoint between the clamps. Ultimate failure loads of up to 6745 N were achieved without slippage of the tissue from the grips. The ultimate tensile strength of the normal equine DDFT determined in this study was 111.82 ± 11.53 N/mm2, and the stress versus grip-to-grip elongation plots for our equine DDFT were representative of a standard non-linear elastic curve obtained in similar studies. CLINICAL SIGNIFICANCE: We present a low cost device for quantifying physical properties of specimens with high connective tissue concentrations and inherent high tensile strength. Results of this study indicate that this device provides a practical alternative to other more costly methods of adequately securing larger tendons and ligaments for tensile strength testing.
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Ligamentos/fisiología , Manejo de Especímenes/instrumentación , Tendones/fisiología , Resistencia a la Tracción/fisiología , Animales , Fenómenos Biomecánicos , CongelaciónRESUMEN
3.5 locking compression plate (LCP) fixation was compared to 3.5 limited contact dynamic compression plate (LC-DCP) fixation in a canine cadaveric, distal humeral metaphyseal gap model. Thirty paired humeri from adult, large breed dogs were separated into equal groups based on testing: static compression, cyclic compression, and cyclic torsion. Humeral constructs stabilised with LCP were significantly stiffer than those plated with LC-DCP when loaded in static axial compression (P = 0.0004). When cyclically loaded in axial compression, the LCP constructs were significantly less stiff than the LC-DCP constructs (P = 0.0029). Constructs plated with LCP were significantly less resistant to torsion over 500 cycles than those plated with LC-DCP (P<0.0001). The increased stiffness of LCP constructs in monotonic loading compared to constructs stabilised with non-locking plates may be attributed to the stability afforded by the plate-screw interface of locking plates. The LCP constructs demonstrated less stiffness in dynamic testing in this model, likely due to plate-bone offset secondary to non-anatomic contouring and occasional incomplete seating of the locking screws when using the torque-limiting screw driver. Resolution of these aspects of LCP application may help improve the stiffness of fixation in fractures modeled by the experimental set-up of this investigation.
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Placas Óseas/veterinaria , Huesos/fisiología , Fijación Interna de Fracturas/veterinaria , Animales , Fenómenos Biomecánicos , Cadáver , Perros , Fijación Interna de Fracturas/instrumentación , Estrés MecánicoRESUMEN
The healing process in guinea pig skin following surgical incisions was evaluated at the molecular level, in vivo, by the use of Raman spectroscopy. After the incisions were closed either by suturing or by laser tissue welding (LTW), differences in the respective Raman spectra were identified. The study determined that the ratio of the Raman peaks of the amide III (1247 cm(-1)) band to a peak at 1326 cm(-1) (the superposition of elastin and keratin bands) can be used to evaluate the progression of wound healing. Conformational changes in the amide I band (1633-1682 cm(-1)) and spectrum changes in the range of 1450-1520 cm(-1) were observed in LTW and sutured skin. The stages of the healing process of the guinea pig skin following LTW and suturing were evaluated by Raman spectroscopy, using histopathology as the gold standard. LTW skin demonstrated better healing than sutured skin, exhibiting minimal hyperkeratosis, minimal collagen deposition, near-normal surface contour, and minimal loss of dermal appendages. A wavelet decomposition-reconstruction baseline correction algorithm was employed to remove the fluorescence wing from the Raman spectra.
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Terapia por Láser/métodos , Fenómenos Fisiológicos de la Piel , Espectrometría Raman/métodos , Cicatrización de Heridas/efectos de la radiación , Amidas/química , Animales , Colágeno/análisis , Procedimientos Quirúrgicos Dermatologicos , Elastina/química , Cobayas , Queratinas/química , Piel/patología , Técnicas de SuturaRESUMEN
OBJECTIVE: To determine the short- and long-term changes in the biomechanical properties and metabolic activity of articular cartilage following the remote application of bipolar radiofrequency (bRF) and monopolar radiofrequency (mRF) energy within the rabbit stifle joint. METHODS: The rabbits were randomly assigned to either Group-1 (normal rabbit food), or they were assigned to Group-2 (2% Cosequin in the diet). Each rabbit underwent bilateral stifle arthroscopy with either bRF or mRF applied to the infrapatellar fat pad for 45 seconds. Cartilage samples were collected at zero, four, and 14 weeks after surgery. Data were analyzed with a mixed model analysis of variance (ANOVA) for chondrocyte death, amount of GAG synthesis, and the equilibrium compressive modulus. RESULTS: A significant increase in histological damage was noted at weeks four and 14 compared to week zero. Most of the chondrocyte death noted with confocal laser microscopy (49 of 56 samples) was noted in the superficial region (outer 25%) of the articular cartilage. GAG synthesis was not significantly different between groups or devices at any time point. A significant difference was not noted in equilibrium compressive modulus throughout the study. CONCLUSIONS: Remote application of bRF and mRF energy lead to immediate chondrocyte death. Most of the damage was superficial hence the metabolic activity and biomechanical properties of the extracellular matrix were maintained throughout this study. Treatment with Cosequin did not prevent superficial chondrocyte death caused by the application of radiofrequency (RF) energy with in the joint.
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Cartílago Articular/patología , Cartílago Articular/efectos de la radiación , Ablación por Catéter/métodos , Condrocitos , Traumatismos Experimentales por Radiación/patología , Análisis de Varianza , Alimentación Animal , Animales , Artroscopía , Cartílago Articular/citología , Cartílago Articular/metabolismo , Supervivencia Celular/efectos de la radiación , Condrocitos/citología , Condrocitos/metabolismo , Condrocitos/efectos de la radiación , Sulfatos de Condroitina/metabolismo , Modelos Animales de Enfermedad , Glicosaminoglicanos/metabolismo , Conejos , Traumatismos Experimentales por Radiación/metabolismo , Distribución Aleatoria , Rodilla de CuadrúpedosRESUMEN
The objective of this study was to evaluate the effect of screw omission on plate strain during axial load to failure and cycling using a Delrin rod gap model. In addition, the differences between the 3.5 mm limited-contact dynamic compression plate (LC-DCP) and the 3.5 mm dynamic compression plate (DCP) were evaluated. Six, 12-hole LC-DCP and DCP plates were applied to Delrin rods with a 1 cm gap located within the central portion of the plate. Six screw configurations were tested with the following variations: three proximal and three distal (six open), four proximal and three distal (five open), four proximal and four distal (four open), five proximal and four distal (three open), five proximal and five distal (two open), six proximal and five distal (one open). Three strain gauges were mounted on each plate within the gap (gauge three) and extended proximally. Additionally, three constructs (six, three, and one open hole) were tested to failure in cyclic loading. The strain measured within the gap (gauge three) was significantly greater than the strain at other gauges for each screw configuration. Strain within the gap did not significantly change with any screw configuration, but did significantly increase at other locations as screws were omitted. Overall, the DCP withstood significantly more cycles than the LC-DCP. Differences were noted within the DCP group with the 6/5 screw configuration lasting for significantly more cycles than the 5/4 and 3/3 constructs. Although overall strain at the gap did not significantly increase with screw omission, the clinical significance remains to be determined.
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Placas Óseas/veterinaria , Tornillos Óseos/veterinaria , Fijación Interna de Fracturas/veterinaria , Ensayo de Materiales/veterinaria , Animales , Fenómenos Biomecánicos , Fuerza Compresiva , Fijación Interna de Fracturas/instrumentación , Fijación Interna de Fracturas/métodos , Fracturas Óseas/cirugía , Fracturas Óseas/veterinaria , Ensayo de Materiales/métodos , Estrés Mecánico , Resistencia a la TracciónRESUMEN
OBJECTIVES: To design a device for the conversion of linear to rotational motion and to use it in order to determine torsional properties of canine tibiae in a universal tension/compression testing machine. METHODS: A fixture incorporating a cable and pulley at one end and a guide block and rail assembly at the other was used to test 10 whole canine tibiae to failure in torsion at 1 degrees/s. RESULTS: The device produced spiral fractures in all tibiae. Torsional strength and stiffness were 13.9 +/- 1.4 N m and 0.53 +/- 0.07 N m/degree, respectively. CLINICAL SIGNIFICANCE: This inexpensive device can be used to evaluate the mechanical properties of long bones after various interventions to improve fracture healing. The device can be adapted for use with any single-axis linear motion testing machine. Assessment of fracture healing often includes loading to failure in torsion (1-5). Torsional testing is typically performed using a biaxial servohydraulic testing machine or similar sophisticated electromechanical equipment. Due to its high cost, many researchers do not have such equipment at their disposal. However, uniaxial tension-compression testing machines are widely available. Therefore, a device that can be used to perform torsional testing in a tension-compression machine would be beneficial. In this study we present a custom fixture for converting the vertical linear motion of a universal testing machine to rotational motion and validate its performance in torsional testing of canine tibiae.
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Biofisica , Curación de Fractura/fisiología , Ensayo de Materiales/veterinaria , Tibia/fisiología , Fracturas de la Tibia/veterinaria , Anomalía Torsional/veterinaria , Animales , Fenómenos Biomecánicos , Biofisica/economía , Biofisica/instrumentación , Análisis Costo-Beneficio , Perros , Diseño de Equipo , Ensayo de Materiales/instrumentación , Ensayo de Materiales/métodos , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Estrés Mecánico , Fracturas de la Tibia/etiología , Anomalía Torsional/complicacionesRESUMEN
An in vitro experimental cadaveric mechanical testing study was performed using 20 radiographically mature dogs, weighing between 18-33 kg. The aim of the study was to compare the axial pull-out strength of 3.5 mm cortical and 4.0 mm cancellous bone screws inserted in the canine proximal tibia using manual and power tapping techniques. 3.5 cortical and 4.0 cancellous bone screws were inserted in canine cadaver proximal tibiae using a manual or power tapping technique. The screws were extracted using a servohydraulic materials testing machine in order to measure axial pull-out strength. Axial pull-out strength was recorded relative to the total bone width and total cortical width of each tibia. The mean axial pull-out strength for all constructs was 717.8+/-56.5 N without any statistically significant difference among groups (p=0.4183). The groups were equal in animal body weight, cortical width and total bone width (p=0.2808). The axial pull-out strength in proportion to cortical and total bone width was not significantly different among groups (p=0.5318). Axial pull-out strengths of 3.5 mm cortical and 4.0 mm cancellous bone screws inserted in the proximal tibial metaphysis were not significantly different. Axial pull-out strength was not affected by the use of power tapping in either screw type.
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Tornillos Óseos/veterinaria , Cadáver , Perros/cirugía , Ensayo de Materiales/veterinaria , Tibia/cirugía , Animales , Fenómenos Biomecánicos , Densidad Ósea , Fijadores Internos , Ensayo de Materiales/instrumentación , Ensayo de Materiales/métodos , Distribución Aleatoria , Resistencia a la TracciónRESUMEN
Chitosan has been researched for implant and wound healing applications. However, there are inconsistencies in reports on the tissue and fibroblast responses to chitosan materials. These inconsistencies may be due to variations in chitosan material characteristics. The aim of this study was to correlate fibroblast responses with known chitosan material characteristics. To achieve this aim, chitosan was characterized for degree of deacetylation (DDA), molecular weight (MW), residual protein and ash contents, and then solution cast into films and characterized for hydrophilicity by water contact angle. The films were seeded with normal human dermal fibroblasts and the number of attached cells was evaluated for after 30 min. Cell proliferation was evaluated over 5 days. This study found no relationship between DDA, contact angle, cell attachment, and or proliferation. General trends were observed for increasing proliferation with increasing residual ash content and decreasing residual protein. These data indicate that chitosan characteristics other than DDA may be important to their biological performance.
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Materiales Biocompatibles/química , Proliferación Celular , Quitosano/química , Fibroblastos/fisiología , Acetilación , Adulto , Animales , Materiales Biocompatibles/metabolismo , Braquiuros , Adhesión Celular/fisiología , Células Cultivadas , Quitosano/metabolismo , Fibroblastos/metabolismo , HumanosRESUMEN
The objective of this study was to evaluate the effect of negative intra-articular pressure on shoulder joint stability in canine cadavers. Cadaver forelimbs from 12 mature dogs were used. The forelimbs were placed in a testing frame and axially preloaded with 4 kg of weight. Shoulder joint stability was tested in flexion, extension, and neutral position before and after venting of the joint capsule. Humeral translation relative to the glenoid was induced by applying a 3 kg load in three different directions (cranial, lateral, and medial) and quantitatively measured by use of an electromagnetic motion tracking system. Peak translational data were compared in each joint position before and after venting of the joint capsule. After venting the shoulder joint capsule, a significant increase in translation was observed in the cranial direction with the joint in neutral position and in the medial direction with the joint in extension. The horizontal translations measured after venting of the joint capsule were likely not clinically relevant. Negative intra-articular pressure is not a major contributor to shoulder stability in dogs during weight-bearing.
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Articulación del Hombro/fisiología , Animales , Fenómenos Biomecánicos , Cadáver , Perros/fisiología , PresiónRESUMEN
In February and in June 1998, two people developed acute hepatitis B following in-patient care in a district general hospital. Initial enquiries indicated their infections were not attributable to staff undertaking exposure-prone procedures (EPPs). We report the findings and implications of the subsequent investigation: a multi-disciplinary, multi-agency investigation, including molecular epidemiological analysis. Occupational Health records showed that staff involved in EPPs with the patients were HBsAg negative. No contact between the patients was identified nor were there failures in sterilization. The patients' HBV strains were identical, indicating a common source. A total of 231 out of 232 staff who might have treated either patient were tested for HBsAg; the remaining doctor, working abroad, was HBsAg- and HBeAg-positive and had the same HBV strain as the patients. On two occasions the doctor's hand had been cut while breaking glass vials, but there was no documentation linking these events to the two patients. The doctor had been vaccinated in 1993 and tested for anti-HBs prior to commencing work in 1997. The doctor was recalled to Occupational Health but did not attend and was not followed up. In total, 4948 patients potentially treated by the doctor received an explanatory letter and 3150 were tested for HBsAg. Only one was positive, and HBV sequencing showed no link to the doctor. Occasionally transmission of HBV from heath-care workers can occur in a non-EPP setting and the implications of this require examination by those setting national policy. Occupational Health Services should investigate clinical heath-care workers who do not respond to vaccination. They should ensure HBV carriers are identified and offer them appropriate advice to prevent transmission to patients.
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Infección Hospitalaria , Hepatitis B/transmisión , Transmisión de Enfermedad Infecciosa de Profesional a Paciente , Exposición Profesional , Adulto , Anciano , ADN Viral/análisis , Femenino , Vacunas contra Hepatitis B , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/patogenicidad , Humanos , Control de Infecciones , Masculino , Personal de Hospital , Factores de RiesgoRESUMEN
The goal of this study was to demonstrate whether cyclically imposed hydrostatic pressure, compressive in nature, could induce fibrocartilaginous metaplasia in a purely tendinous cell source in vitro. The effect of short-duration cyclic hydrostatic pressure on tendon fibroblasts (tenocytes) expanded from rat Achilles tendon was studied. Total RNA was isolated either immediately after loading or 24 h later. The mRNA expression of tendon and cartilage specific markers - Collagen types I and II, Sox9, and Aggrecan was quantified by real-time reverse transcription polymerase chain reaction over multiple biological samples (n=6). For immediately isolated RNA samples, there were statistically significant increases in mRNA expression of Aggrecan and Collagen type II, while Collagen type I significantly decreased. Noticeably, for RNA samples isolated 24 h later, there were further increases in mRNA expression of Aggrecan and Collagen type II, whereas Collagen type I increased roughly three-fold relative to the non-loaded control. These findings support the hypothesis that cyclic hydrostatic pressurization can induce fibrocartilaginous metaplasia in tenocytes by upregulation of cartilaginous gene expression. Also, it was demonstrated that changes in mRNA expression as a result of single 2 h pressurization persist even up to 24 h.
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Tendón Calcáneo/citología , Tendón Calcáneo/patología , Fibroblastos/patología , Fibroblastos/fisiología , Agrecanos/metabolismo , Animales , Animales Recién Nacidos , Biomarcadores/metabolismo , Técnicas de Cultivo de Célula , Células Cultivadas , Colágeno Tipo I/metabolismo , Colágeno Tipo II/metabolismo , Proteínas del Grupo de Alta Movilidad/metabolismo , Presión Hidrostática , Masculino , Metaplasia , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Factor de Transcripción SOX9 , Factores de Tiempo , Factores de Transcripción/metabolismoRESUMEN
Living bone cells are responsive to mechanical loading. Consequently, numerous in vitro models have been developed to examine the application of loading to cells. However, not all systems are suitable for the fibrous and porous three-dimensional materials, which are preferable for tissue repair purposes, or for the production of tissue engineering scaffolds. For three-dimensional applications, mechanical loading of cells with either fluid flow systems or hydrodynamic pressure systems has to be considered. Here, we aimed to evaluate the response of osteoblast-like cells to hydrodynamic compression, while growing in a three-dimensional titanium fiber mesh scaffolding material. For this purpose, a custom hydrodynamic compression chamber was built. Bone marrow cells were obtained from the femora of young (12-day-old) or old (1-year-old) rats, and precultured in the presence of dexamethasone and beta-glycerophosphate to achieve an osteoblast-like phenotype. Subsequently, cells were seeded onto the titanium mesh scaffolds, and subjected to hydrodynamic pressure, alternating between 0.3 to 5.0 MPa at 1 Hz, at 15-min intervals for a total of 60 min per day for up to 3 days. After pressurization, cell viability was checked. Afterward, DNA levels, alkaline phosphatase (ALP) activity, and extracellular calcium content were measured. Finally, all specimens were observed with scanning electron microscopy. Cell viability studies showed that the applied pressure was not harmful to the cells. Furthermore, we found that cells were able to detect the compression forces, because we did see evident effects on the cell numbers of the cells derived from old animals. However, there were no other changes in the cells under pressure. Finally, it was also noticeable that cells from old animals did not express ALP activity, but did show similar calcified extracellular matrix formation to the cells from young animals. In conclusion, the difference in DNA levels as reaction toward pressure, and the difference in ALP levels, suggest that the osteogenic properties of bone marrow-derived osteoblast-like cells are different with respect to the age of the donor.
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Materiales Biocompatibles , Osteoblastos/citología , Titanio , Envejecimiento/patología , Fosfatasa Alcalina/metabolismo , Animales , Calcio/metabolismo , Supervivencia Celular , Células Cultivadas , ADN/metabolismo , Presión Hidrostática , Ensayo de Materiales , Microscopía Electrónica de Rastreo , Osteoblastos/metabolismo , Ratas , Estrés Mecánico , Ingeniería de TejidosRESUMEN
While the potential for intermittent hydrostatic pressure to promote cartilaginous matrix synthesis is well established, its potential to influence chondroinduction remains poorly understood. This study examined the effects of relatively short- and long-duration cyclic hydrostatic compression on the chondroinduction of C3H/10T1/2 murine embryonic fibroblasts by recombinant human bone morphogenetic protein-2 (rhBMP-2). Cells were seeded at high density into round bottom wells of a 96-well plate and supplemented with 25 ng/ml rhBMP-2. Experimental cultures were subjected to either 1,800 cycles/day or 7,200 cycles/day of 1 Hz sinusoidal hydrostatic compression to 5 MPa (applied 10 min on/10 min off) for 3 days. Non-pressurized control and experimental cultures were maintained in static culture for an additional 5 days. Cultures were then analyzed for alcian blue staining intensity, DNA and sulfated glycosaminoglycan (sGAG) content, and for the rate of collagen synthesis. Whereas cultures subjected to 1,800 pressure cycles exhibited no significant differences (statistical or qualitative) compared to controls, those subjected to 7,200 cycles stained more intensely with alcian blue, contained nearly twice as much sGAG, and displayed twice the rate of collagen synthesis as non-pressurized controls. This study demonstrates the potential for cyclic hydrostatic compression to stimulate chondrogenic differentiation of the C3H/10T1/2 cell line in a duration-dependent manner.
Asunto(s)
Proteínas Morfogenéticas Óseas/farmacología , Condrocitos/citología , Condrocitos/fisiología , Condrogénesis/fisiología , Fibroblastos/citología , Fibroblastos/fisiología , Mecanotransducción Celular/fisiología , Factor de Crecimiento Transformador beta/farmacología , Animales , Proteína Morfogenética Ósea 2 , Diferenciación Celular/efectos de los fármacos , Diferenciación Celular/fisiología , Células Cultivadas , Condrocitos/efectos de los fármacos , Condrogénesis/efectos de los fármacos , Fuerza Compresiva/fisiología , Fibroblastos/efectos de los fármacos , Mecanotransducción Celular/efectos de los fármacos , Ratones , Ratones Endogámicos C3H , Presión , Ingeniería de Tejidos/métodosRESUMEN
Chitosan, a derivative of the bio-polysaccharide chitin, has shown promise as a bioactive material for implant, tissue engineering and drug-delivery applications. The aim of this study was to evaluate the contact angle, protein adsorption and osteoblast precursor cell attachment to chitosan coatings bonded to titanium. Rough ground titanium (Ti) coupons were solution cast and bonded to 91.2% de-acetylated chitosan (1 wt% chitosan in 0.2% acetic acid) coatings via silane reactions. Non-coated Ti was used as controls. Samples were sterilized by ethylene oxide gas prior to experiments. Contact angles on all surfaces were measured using water. 5 x 10(4) cells/ml of ATCC CRL 1486 human embryonic palatal mesenchyme (HEPM) cells, an osteoblast precursor cell line, were used for the cell attachment study. SEM evaluations were performed on cells attached to all surfaces. Contact angles and cell attachment on all surfaces were statistically analyzed using ANOVA. The chitosan-coated surfaces (76.4 +/- 5.1 degrees) exhibited a significantly greater contact angle compared to control Ti surfaces (32.2 +/- 6.1 degrees). Similarly, chitosan-coated surfaces exhibited significantly greater (P < 0.001) albumin adsorption, fibronectin adsorption and cell attachment, as compared to the control Ti surfaces. Coating chitosan on Ti surfaces decreased the wettability of the Ti, but increased protein adsorption and cell attachment. Increased protein absorption and cell attachment on the chitosan-coated Ti may be of benefit in enhancing osseointegration of implant devices.