RESUMEN
BACKGROUND: Wnt signaling plays an important role in development and maintenance of many organs and tissues. The most-studied secreted Wnt inhibitors are sclerostin (SOST), Dickkopf-related protein 1 (DKK-1), and secreted frizzled related protein 1 (SFRP-1) which play important roles in bone turnover. The present study investigated the relationship between serum Wnt inhibitors and diseases with excessive ossification structures, such as ossification of posterior longitudinal ligament (OPLL), ankylosing spondylitis (AS), diffuse idiopathic skeletal hyperostosis (DISH), and ossification of yellow ligament (OYL). METHODS: Twenty-five patients with AS, DISH, OPLL, or OYL were recruited in this study. Fasting peripheral blood samples were collected from all patients and nine controls. Various biomarkers of bone turnover including osteocalcin (OSC), osteoprotegerin (OPG), SFRP-1, DKK-1, and SOST were investigated. RESULTS: Our data showed that serum levels of OSC were higher, but Dkk-1 levels were lower in AS, DISH, OPLL, and OYL patients than those in the controls. Serum levels of SFRP-1 were significantly higher in DISH patients than those in the controls. Serum levels of SOST were significantly higher in DISH and OPLL patients than both levels in the controls. Serum levels of OPG were lower in AS patients than those in the controls. Serum levels of OSC were higher in the OPLL patients than those in the AS patients. Serum levels of DKK-1, SFRP-1, SOST, and OPG were not significantly different between the different disease groups. CONCLUSIONS: In this exploratory study, both OSC and DKK-1 levels are correlated with the clinical conditions associated with excessive ossification, indicating that blood OSC and DKK-1 levels may serve as diagnostic biomarkers for AS, DISH, OPLL, and OYL. These findings may also help discover potential drug therapies for management of these diseases in the future.
Asunto(s)
Remodelación Ósea , Hiperostosis Esquelética Difusa Idiopática/sangre , Osificación del Ligamento Longitudinal Posterior/sangre , Espondilitis Anquilosante/sangre , Proteínas Wnt/antagonistas & inhibidores , Proteínas Adaptadoras Transductoras de Señales , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/sangre , Densidad Ósea , Proteínas Morfogenéticas Óseas/sangre , Femenino , Marcadores Genéticos , Humanos , Péptidos y Proteínas de Señalización Intercelular/sangre , Péptidos y Proteínas de Señalización Intracelular , Masculino , Proteínas de la Membrana/sangre , Persona de Mediana Edad , Osteocalcina/sangre , Osteoprotegerina/sangre , Proteínas , Proteínas Wnt/metabolismo , Vía de Señalización WntRESUMEN
BACKGROUND: Hyperbaric oxygenation was shown to increase bone healing in a rabbit model. However, little is known about the regulatory factors and molecular mechanism involved.We hypothesized that the effect of hyperbaric oxygen (HBO) on bone formation is mediated via increases in the osteogenic differentiation of mesenchymal stem cells (MSCs) which are regulated by Wnt signaling. METHODS: The phenotypic characterization of the MSCs was analyzed by flow cytometric analysis. To investigate the effects of HBO on Wnt signaling and osteogenic differentiation of MSCs, mRNA and protein levels of Wnt3a, beta-catenin, GSK-3beta, Runx 2, as well as alkaline phosphatase activity, calcium deposition, and the intensity of von Kossa staining were analyzed after HBO treatment. To investigate the effects of HBO on Wnt processing and secretion, the expression of Wntless and vacuolar ATPases were quantified after HBO treatment. RESULTS: Cells expressed MSC markers such as CD105, CD146, and STRO-1. The mRNA and protein levels of Wnt3a, ß-catenin, and Runx 2 were up-regulated, while GSK-3ß was down-regulated after HBO treatment. Western blot analysis showed an increased ß-catenin translocation with a subsequent stimulation of the expression of target genes after HBO treatment. The above observation was confirmed by small interfering (si)RNA treatment. HBO significantly increased alkaline phosphatase activity, calcium deposition, and the intensity of von Kossa staining of osteogenically differentiated MSCs. We further showed that HBO treatment increased the expression of Wntless, a retromer trafficking protein, and vacuolar ATPases to stimulate Wnt processing and secretion, and the effect was confirmed by siRNA treatment. CONCLUSIONS: HBO treatment increased osteogenic differentiation of MSCs via regulating Wnt processing, secretion, and signaling.
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Células de la Médula Ósea/efectos de los fármacos , Células Madre Mesenquimatosas/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Oxígeno/farmacología , Vía de Señalización Wnt/efectos de los fármacos , Adulto , Anciano , Biomarcadores , Células de la Médula Ósea/metabolismo , Células Cultivadas , Femenino , Humanos , Oxigenoterapia Hiperbárica , Péptidos y Proteínas de Señalización Intracelular/biosíntesis , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Células Madre Mesenquimatosas/metabolismo , Persona de Mediana Edad , Interferencia de ARN , ARN Mensajero/biosíntesis , ARN Mensajero/genética , ARN Interferente Pequeño/farmacología , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores Acoplados a Proteínas G/biosíntesis , Receptores Acoplados a Proteínas G/genética , Regulación hacia Arriba , ATPasas de Translocación de Protón Vacuolares/biosíntesis , ATPasas de Translocación de Protón Vacuolares/genética , Vía de Señalización Wnt/fisiologíaRESUMEN
MicroRNA (miRNA) 107 expression is downregulated but Wnt3a protein and ß-catenin are upregulated in degenerated intervertebral disc (IVD). We investigated mir-107/Wnt3a-ß-catenin signaling in vitro and in vivo following hyperbaric oxygen (HBO) intervention. Our results showed 96 miRNAs were upregulated and 66 downregulated in degenerated nucleus pulposus cells (NPCs) following HBO treatment. The 3' untranslated region (UTR) of the Wnt3a mRNA contained the "seed-matched-sequence" for miR-107. MiR-107 was upregulated and a marked suppression of Wnt3a was observed simultaneously in degenerated NPCs following HBO intervention. Knockdown of miR-107 upregulated Wnt3a expression in hyperoxic cells. HBO downregulated the protein expression of Wnt3a, phosphorylated LRP6, and cyclin D1. There was decreased TOP flash activity following HBO intervention, whereas the FOP flash activity was not affected. HBO decreased the nuclear translocation of ß-catenin and decreased the secretion of MMP-3 and -9 in degenerated NPCs. Moreover, rabbit serum KS levels and the stained area for Wnt3a and ß-catenin in repaired cartilage tended to be lower in the HBO group. We observed that HBO inhibits Wnt3a/ß-catenin signaling-related pathways by upregulating miR-107 expression in degenerated NPCs. HBO may play a protective role against IVD degeneration and could be used as a future therapeutic treatment.
Asunto(s)
Oxigenoterapia Hiperbárica , MicroARNs , Núcleo Pulposo , Animales , Conejos , beta Catenina , Oxígeno , Modelos Animales , Regiones no Traducidas 3' , MicroARNs/genéticaRESUMEN
The authors investigated poly(lactide-co-glycolide) (PLGA) capsule and collagen composite system for antibiotics and bone cells delivery to treat infected bone defects. Poly(lactide-co-glycolide) was mixed with vancomycin and hot compressing molded to form an antibiotic capsule. Rabbit mesenchymal stem cells (MSCs) were entrapped in collagen gel phase and dispersed throughout the void volume of capsule. In vitro study, the composite systems were cultured in complete or osteogenic medium for 21 days. The profiles of vancomycin released from the systems were evaluated using the high-performance liquid chromatography method. Relative activity of vancomycin against Staphylococcus aureus was determined by an antibiotic disk diffusion method. The expression of osteogenic gene was determined by reverse-transcription polymerase chain reaction. The alkaline phosphatase activity and calcium level of the MSCs were assessed. Analytical results demonstrated that the concentrations of vancomycin eluted from the composite system were above the minimal inhibitory concentration for 21 days. Sample inhibition zone was 10 to 24 mm, and the relative activity was 17.6% to 100%. mRNA of Cbfa1 and osteocalcin were detected, and increased alkaline phosphatase activity and calcium levels were noted. In in vivo investigation, the PKH 26-labeled MSCs and composite systems were implanted in the distal femoral cavities of four rabbits. The local concentration of vancomycin was above the minimal inhibitory concentration for 56 days. Sample inhibition zone was 9 mm to 24 mm, and the relative activity was 11.8% to 100%. Implanted PKH 26-labeled MSCs were identified in the newly formed bony trabeculae in specimens at 2 and 4 months after implantation. The results offer a potential approach to meet clinical requirements in the treatment of infected bone defects.
Asunto(s)
Huesos/citología , Colágeno/farmacología , Sistemas de Liberación de Medicamentos/métodos , Ácido Láctico/farmacología , Trasplante de Células Madre Mesenquimatosas/métodos , Osteomielitis/tratamiento farmacológico , Ácido Poliglicólico/farmacología , Vancomicina/farmacología , Fosfatasa Alcalina/análisis , Animales , Materiales Biocompatibles/química , Calcio/análisis , Cápsulas , Células Cultivadas , Cromatografía Líquida de Alta Presión , Portadores de Fármacos , Fémur/cirugía , Técnicas In Vitro , Ácido Láctico/administración & dosificación , Células Madre Mesenquimatosas/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Microscopía Fluorescente , Osteogénesis , Ácido Poliglicólico/administración & dosificación , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Conejos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Coloración y Etiquetado , Vancomicina/administración & dosificaciónRESUMEN
BACKGROUND: Posterolateral spinal fusion is used to treat patients with degenerative spinal disorders. In this study, we investigated the effectiveness of a mesenchymal stem cell (MSC)/hydroxyapatite/type I collagen hybrid graft for posterolateral spinal fusion in a rabbit model. METHODS: In vitro study, the hybrid graft was cultured in complete or osteogenic medium for 7 days and 14 days and examined by scanning electron microscopy. The alkaline phosphatase activity of the MSCs was assessed and the expression of osteogenic gene was determined by reverse transcription polymerase chain reaction. In vivo investigation, spinal fusion was examined using radiography, manual palpation, computed tomography, torsional loading tests, and histologic analysis. Furthermore, using a PKH fluorescence labeling system, we examined whether the newly formed bone was derived from the transplanted MSCs. RESULTS: Our data suggested that the MSCs differentiated into osteoblasts and produced extracellular matrix in the hybrid graft. Increased alkaline phosphatase activity was noted and mRNA of Cbfa-1 and osteopontin were detected. Radiographs and computed tomography images showed a continuous bone bridge and a satisfactory fusion mass incorporated into the transverse processes. The results of manual palpation and biomechanical data did not significantly differ between the two groups. Histologic examination of both groups revealed the presence of cartilage and endochondral ossification in the gaps between the grafted fragments. In situ tracing of the PKH 67-labeled MSCs indicated that the transplanted MSCs were partly responsible for the new bone formation. CONCLUSION: The hybrid graft could be effectively used to achieve posterolateral spinal fusion.
Asunto(s)
Colágeno/farmacología , Durapatita/farmacología , Ácido Láctico/farmacología , Trasplante de Células Madre Mesenquimatosas , Ácido Poliglicólico/farmacología , Fusión Vertebral/métodos , Fosfatasa Alcalina/análisis , Animales , Trasplante de Médula Ósea , Sustitutos de Huesos , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Fluorescencia , Curación de Fractura/efectos de los fármacos , Vértebras Lumbares/diagnóstico por imagen , Vértebras Lumbares/efectos de los fármacos , Vértebras Lumbares/patología , Vértebras Lumbares/cirugía , Microscopía Electrónica de Rastreo , Compuestos Orgánicos/farmacología , Oseointegración/efectos de los fármacos , Osteogénesis/efectos de los fármacos , Palpación , Copolímero de Ácido Poliláctico-Ácido Poliglicólico , Conejos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Estrés Mecánico , Tomografía Computarizada por Rayos X , Soporte de PesoRESUMEN
BACKGROUND: MicroRNA (miRNA) plays a vital role in the intervertebral disc (IVD) degeneration. The expression level of miR-573 was downregulated whereas Bax was upregulated notably in human degenerative nucleus pulposus cells. In this study, we aimed to investigate the role of miR-573 in human degenerative nucleus pulposus (NP) cells following hyperbaric oxygen (HBO) treatment. METHODS: NP cells were separated from human degenerated IVD tissues. The control cells were maintained in 5% CO2/95% air and the hyperoxic cells were exposed to 100% O2 at 2.5 atmospheres absolute. MiRNA expression profiling was performed via microarray and confirmed by real-time PCR, and miRNA target genes were identified using bioinformatics and luciferase reporter assays. The mRNA and protein levels of Bax were measured. The proliferation of NPCs was detected using MTT assay. The protein expression levels of Bax, cleaved caspase 9, cleaved caspase 3, pro-caspase 9, and pro-caspase 3 were examined. RESULTS: Bioinformatics analysis indicated that the 3' untranslated region (UTR) of the Bax mRNA contained the "seed-matched-sequence" for hsa-miR-573, which was validated via reporter assays. MiR-573 was induced by HBO and simultaneous suppression of Bax was observed in NP cells. Knockdown of miR-573 resulted in upregulation of Bax expression in HBO-treated cells. In addition, overexpression of miR-573 by HBO increased cell proliferation and coupled with inhibition of cell apoptosis. The cleavage of pro-caspase 9 and pro-caspase 3 was suppressed while the levels of cleaved caspase 9 and caspase 3 were decreased in HBO-treated cells. Transfection with anti-miR-573 partly suppressed the effects of HBO. CONCLUSION: Mir-573 regulates cell proliferation and apoptosis by targeting Bax in human degenerative NP cells following HBO treatment.
Asunto(s)
Apoptosis/genética , Proliferación Celular/genética , Oxigenoterapia Hiperbárica , MicroARNs/fisiología , Núcleo Pulposo/citología , Proteína X Asociada a bcl-2/metabolismo , Anciano , Células Cultivadas , Femenino , Expresión Génica/genética , Humanos , Degeneración del Disco Intervertebral/metabolismo , Degeneración del Disco Intervertebral/patología , Masculino , Persona de Mediana Edad , Núcleo Pulposo/metabolismo , Proteína X Asociada a bcl-2/genéticaRESUMEN
PURPOSE: A retrospective study was conducted on arthroscopic ganglionectomy in wrists using a novel intrafocal cystic portal. The safety and efficacy of this technique were assessed by treatment of 15 wrists in 15 patients. METHODS: Arthroscopic ganglionectomy was performed by the same surgeon with the patient under general anesthesia or regional block. Preoperative complaints, intraoperative findings, and postoperative results of all the patients were reported. The mean follow-up was 15.3 months. Functional assessment by use of modified Mayo wrist scores, patient satisfaction, and recurrence were included in the follow-up evaluation. RESULTS: Two thirds of the patients acquired good to excellent results, whereas the results for the remaining third were fair. Complications included 1 recurrence and 1 case of transient paresthesia sensation. The most common arthroscopic findings were capsular and ligament lesions, rather than ganglionic stalks. CONCLUSIONS: Arthroscopic ganglionectomy through an intrafocal cystic portal is a safe and efficacious option for the treatment of painful wrist ganglia. LEVEL OF EVIDENCE: Level IV, therapeutic case series.
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Artroscopios , Artroscopía/métodos , Quistes Óseos/cirugía , Ganglionectomía/métodos , Articulación de la Muñeca/cirugía , Adulto , Anciano , Anciano de 80 o más Años , Artralgia/diagnóstico , Artralgia/etiología , Quistes Óseos/complicaciones , Diseño de Equipo , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Dimensión del Dolor , Estudios Retrospectivos , Resultado del Tratamiento , Adulto JovenRESUMEN
Spontaneous cell fusion can occur in cocultured stem cells. We examined whether telomerase activity change and cell fusion occurred in mesenchymal stem cell (MSC) and nucleus pulposus cell (NPC) coculture. MSCs and NPCs were labeled with PKH26 and PKH67 dyes and cocultured at a 50:50 ratio. An equal number of MSCs or NPCs were used as the control. After 14 days, cells were evaluated by cell growth, telomerase activity, quantitative reverse transcriptase-polymerase chain reaction (RT-PCR), immunohistochemistry, and histologic observation. Cell fusion was confirmed by microscopic observation and fluorescence-activated cell sorter (FACS) analysis. The results suggested cell growth rate and telomerase activity were higher in cocultured cells than in NPCs cultured alone. The mRNA expression levels of the Type II collagen and aggrecan were elevated in cocultured cells. Immunohistochemical analysis revealed positive staining for Type II collagen and keratan sulfate in NPCs cultured alone and in a proportion of cocultured cells. Histologic observation revealed binucleated cocultured cells expressed both PKH dyes in the same location and slide focus. The FACS analysis revealed 42% of cocultured cells were double-stained. Cocultured cells partially maintained the NPC phenotype. The partially maintained phenotype of the NPCs may be attributable to spontaneous cell fusion in association with increased telomerase activity.
Asunto(s)
Células de la Médula Ósea/fisiología , Comunicación Celular , Disco Intervertebral/fisiología , Células Madre Mesenquimatosas/fisiología , Agrecanos/genética , Animales , Biomarcadores/metabolismo , Células de la Médula Ósea/enzimología , Fusión Celular , Proliferación Celular , Separación Celular , Células Cultivadas , Técnicas de Cocultivo , Colágeno Tipo II/genética , Colágeno Tipo II/metabolismo , Citometría de Flujo , Inmunohistoquímica , Disco Intervertebral/citología , Disco Intervertebral/enzimología , Sulfato de Queratano/metabolismo , Células Madre Mesenquimatosas/enzimología , Microscopía Fluorescente , Fenotipo , ARN Mensajero/metabolismo , Conejos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Telomerasa/metabolismo , Factores de TiempoRESUMEN
BACKGROUND: The expression of both high-mobility group box 1 (HMGB1) and receptor for advanced glycation end-products (RAGE) is upregulated in degenerated discs. HMGB1 is known to function as a coupling factor between hypoxia and inflammation in arthritis, and this inflammatory response is modulated by microRNAs (miRNAs), with miR-107 expression downregulated during hypoxia. In this study, we investigated the regulation of the miR-107/HMGB1/RAGE pathway in degenerated nucleus pulposus cells (NPCs) after hyperbaric oxygen (HBO) treatment. METHODS: NPCs were separated from human degenerated intervertebral disc tissues. The control cells were maintained in 5% CO2/95% air, and the hyperoxic cells were exposed to 100% O2 at 2.5 atmospheres absolute. MiRNA expression profiling was performed via microarray and confirmed by real-time PCR, and miRNA target genes were identified using bioinformatics and luciferase reporter assays. The cellular protein and mRNA levels of HMGB1, RAGE, and inducible nitric oxide synthase (iNOS) were assessed, and the phosphorylation of MAPK (p38MAPK, ERK, and JNK) was evaluated. Additionally, cytosolic and nuclear fractions of the IκBα and NF-κB p65 proteins were analyzed, and secreted HMGB1 and metalloprotease (MMP) levels in the conditioned media were quantified. RESULTS: Using microarray analyses, 96 miRNAs were identified as upregulated and 66 downregulated following HBO treatment. Based on these results, miR-107 was selected for further investigation. Bioinformatics analyses indicated that the 3' untranslated region of the HMGB1 mRNA contained the "seed-matched-sequence" for hsa-miR-107, which was validated via dual-luciferase reporter assays. MiR-107 was markedly induced by HBO, and simultaneous suppression of HMGB1 was observed in NPCs. Knockdown of miR-107 resulted in upregulation of HMGB1 expression in HBO-treated cells, and HBO treatment downregulated the mRNA and protein levels of HMGB1, RAGE, and iNOS and the secretion of HMGB1. In addition, HBO treatment upregulated the protein levels of cytosolic IκBα and decreased the nuclear translocation of NF-κB in NPCs. Moreover, HBO treatment downregulated the phosphorylation of p38MAPK, ERK, and JNK and significantly decreased the secretion of MMP-3, MMP-9, and MMP-13. CONCLUSIONS: HBO inhibits pathways related to HMGB1/RAGE signaling via upregulation of miR-107 expression in degenerated human NPCs.
Asunto(s)
Proteína HMGB1/genética , Oxigenoterapia Hiperbárica/métodos , Degeneración del Disco Intervertebral/terapia , MicroARNs/genética , Receptor para Productos Finales de Glicación Avanzada/efectos de los fármacos , Adulto , Anciano , Anciano de 80 o más Años , Células Cultivadas , Femenino , Perfilación de la Expresión Génica/métodos , Proteína HMGB1/metabolismo , Humanos , Degeneración del Disco Intervertebral/genética , Degeneración del Disco Intervertebral/metabolismo , Masculino , Persona de Mediana Edad , Núcleo Pulposo/efectos de los fármacos , Núcleo Pulposo/metabolismo , Núcleo Pulposo/patología , Oxígeno/farmacología , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Transducción de Señal/genética , Regulación hacia ArribaRESUMEN
Despite moderate success in clinical applications, outcome of tendon grafts employed for anterior cruciate ligament (ACL) reconstruction remains unsatisfactory. This study investigated the effects of hyperbaric oxygen (HBO) on neovascularization at the tendon-bone junction, collagen fibers of the tendon graft, and the tendon graft-bony interface incorporated into the osseous tunnel in rabbits. Forty rabbits were assigned to two groups. The HBO group was exposed to 100% oxygen at 2.5 atmospheres pressure for 2 h daily, 5 consecutive days in a week. The control group was maintained in cages exposed to normal air. Histological studies of 12 rabbits were performed postoperatively at 6, 12, and 18 weeks. Biomechanical studies of 24 rabbits were conducted postoperatively at 12 and 18 weeks. Electron microscopy (EM) analyses of four rabbits were performed postoperatively at 18 weeks. Experimental results demonstrated that a higher number of Sharpey's fibers bridged the newly formed fibrocartilage and graft in the HBO group than in the control group. In addition, HBO treatment increased neovascularization and enhanced the incorporation of the progressive interface between tendon graft and bone. Biomechanical analysis showed that the HBO group achieved higher maximal pullout strength than the control group. Examination by EM showed that HBO treatment resulted in regenerated collagen fibers with increased compaction and regularity. Based on experimental results, HBO treatment is a treatment modality that potentially improves outcome following ACL reconstruction.
Asunto(s)
Supervivencia de Injerto , Oxigenoterapia Hiperbárica , Neovascularización Fisiológica , Tendones/trasplante , Cicatrización de Heridas , Animales , Huesos/irrigación sanguínea , Huesos/patología , Colágeno/metabolismo , Fibroblastos/patología , Fibroblastos/ultraestructura , Microscopía Electrónica , Complicaciones Posoperatorias/fisiopatología , Complicaciones Posoperatorias/psicología , Conejos , Tendones/irrigación sanguínea , Tendones/patología , Resistencia a la TracciónRESUMEN
PURPOSE: We prospectively assessed 22 consecutively treated patients to determine the effectiveness and safety of arthroscopically assisted posterior cruciate ligament (PCL) reconstruction by using a quadriceps tendon autograft. METHODS: Twenty-two patients with isolated PCL injury who underwent PCL reconstruction with a quadriceps tendon autograft were enrolled in the prospective study. The average follow-up period was 66 months (range, 60-76). Follow-up included Lysholm knee scores, Tegner activity scores, International Knee Documentation Committee (IKDC) score, thigh muscle assessment, and radiographic assessment. RESULTS: The mean preoperative Lysholm score for 22 knees was 67 (range, 50 to 75), and the mean postoperative Lysholm score was 89 (range, 75 to 98). Nineteen of 22 patients (86%) displayed good or excellent results in the final assessment. The mean preoperative Tegner score for 22 knees was 3 (range, 2 to 5), whereas the mean postoperative Tegner score was 6 (range, 3 to 9). There were statistically significant improvements in Lysholm score (P = .009), Tegner score (P = .039), postoperative KT-1000 arthrometer (MEDmetric, San Diego, CA) scores (P = .006), final IKDC rating (P = .035), and thigh atrophy and muscle strength (P < .05) when compared with preoperative data. Regarding IKDC final rating, 82% of the patients (18 of 22) were assessed as normal or nearly normal (grade A or B). CONCLUSIONS: After follow-up for more than 60 months, the analytical results showed patients achieved satisfactory function after PCL reconstruction by using a quadriceps tendon-patellar bone autograft. This study suggests that a quadriceps tendon autograft is sufficiently large and strong and can achieve good ligament function after reconstruction. LEVEL OF EVIDENCE: Level IV, therapeutic study.
Asunto(s)
Artroscopía/métodos , Ligamento Cruzado Posterior/cirugía , Músculo Cuádriceps , Rango del Movimiento Articular/fisiología , Tendones/trasplante , Adolescente , Adulto , Femenino , Estudios de Seguimiento , Humanos , Inestabilidad de la Articulación/prevención & control , Traumatismos de la Rodilla/diagnóstico , Traumatismos de la Rodilla/cirugía , Masculino , Persona de Mediana Edad , Ligamento Cruzado Posterior/fisiopatología , Probabilidad , Estudios Prospectivos , Procedimientos de Cirugía Plástica/métodos , Recuperación de la Función , Factores de Riesgo , Factores de Tiempo , Trasplante Autólogo , Resultado del TratamientoRESUMEN
PURPOSE: This study prospectively evaluated 20 patients treated consecutively to determine patient outcomes, efficacy, and complication potential of arthroscopically assisted posterior cruciate ligament (PCL) reconstruction performed with hamstring tendon grafts. METHODS: Twenty patients (15 men and 5 women), each with an isolated PCL injury, underwent PCL reconstruction with hamstring tendon autograft and were enrolled in this prospective study. Average age at time of surgery was 29 years (range, 20 to 57 years). Average time from injury to surgery was 4 months (range, 3 to 12 months). Average follow-up period was 40 months (range, 36 to 50 months). Patients underwent regular follow-up after clinical and radiographic preoperative and postoperative evaluation. Follow-up examinations comprised the Lysholm Knee Score, the Tegner Activity Score, the International Knee Documentation Committee (IKDC) score, thigh muscle assessment, and radiographic evaluation. RESULTS: Mean preoperative Lysholm score for 20 knees was 63 +/- 10 (range, 48-73); mean postoperative Lysholm score was 93 +/- 9 (range, 77-100). Eighteen of 20 patients (90%) showed good or excellent results at final assessment. Mean preinjury and preoperative Tegner scores were 7 +/- 1.5 (range, 5-9) and 3 +/- 1.9 (range, 2-5), respectively; mean postoperative Tegner score for 20 knees was 6.3 +/- 2.4 (range, 4-9). In final IKDC ratings, 85% of patients (17 of 20) were assessed as normal or near normal (grade A or B). A statistically significant improvement was seen in thigh girth difference, extensor strength ratio, and flexor strength ratio before and after reconstruction at a minimum of 3 years of follow-up. CONCLUSIONS: After follow-up for longer than 36 months, analytical results showed satisfactory function after PCL reconstruction with the use of hamstring tendon autografts. We suggest that the hamstring tendon autograft is a safe, effective, and acceptable choice for PCL reconstruction, and that it affords good ligament reconstruction. LEVEL OF EVIDENCE: IV, therapeutic case series.
Asunto(s)
Artroscopía , Procedimientos de Cirugía Plástica , Ligamento Cruzado Posterior/lesiones , Ligamento Cruzado Posterior/cirugía , Tendones/trasplante , Adulto , Femenino , Estudios de Seguimiento , Humanos , Articulación de la Rodilla/fisiopatología , Masculino , Persona de Mediana Edad , Músculo Esquelético/patología , Músculo Esquelético/fisiopatología , Atrofia Muscular/patología , Ligamento Cruzado Posterior/fisiopatología , Periodo Posoperatorio , Estudios Prospectivos , Rango del Movimiento Articular , Procedimientos de Cirugía Plástica/efectos adversos , Recuperación de la Función , Muslo , Trasplante Autólogo , Índices de Gravedad del Trauma , Resultado del TratamientoRESUMEN
BACKGROUND: Clinical experience and animal studies have suggested that positron emission tomography (PET) using fluorine-18-labeled fluorodeoxyglucose ((18)F-FDG) may be promising for imaging of bone infections. In this study, we aimed to establish the accuracy of (18)F-FDG PET scanning for monitoring the response to poly(lactide-co-glycolide) (PLGA) vancomycin beads for treatment of bone infection. METHODS: PLGA was mixed with vancomycin and hot-compress molded to form antibiotic beads. In vitro, elution assays and bacterial inhibition tests were employed to characterize the released antibiotics. In vivo, cylindrical cavities were made in six adult male New Zealand white rabbits, and Staphylococcus aureus or saline was injected into the cavity to create a bone infection. After 2 weeks, the infection was confirmed by bacterial cultures, and the defect was filled with PLGA vancomycin beads. The treatment response was monitored by (18)F-FDG PET. RESULTS: The biodegradable beads released high concentrations of vancomycin (well above the breakpoint sensitivity concentration) for treatment of bone infection. In bacterial inhibition tests, the diameter of the sample inhibition zone ranged from 6.5 to 10 mm, which was equivalent to 12.5-100 % relative activity. (18)F-FDG PET results showed that uncomplicated bone healing was associated with a temporary increase in (18)F-FDG uptake at 2 weeks, with return to near baseline at 6 weeks. In the infected animals, localized infection resulted in intense continuous uptake of (18)F-FDG, which was higher than that in uncomplicated healing bones. Bone infection was confirmed with positive bacterial cultures. In vancomycin-treated animals, data showed rapidly decreasing amounts of (18)F-FDG uptake after treatment. CONCLUSIONS: In vitro and in vivo analyses showed that the use of biodegradable PLGA vancomycin beads successfully eradicated S. aureus infection in damaged bone.
Asunto(s)
Antibacterianos/administración & dosificación , Osteomielitis/tratamiento farmacológico , Infecciones Estafilocócicas/tratamiento farmacológico , Staphylococcus aureus/efectos de los fármacos , Vancomicina/administración & dosificación , Implantes Absorbibles , Animales , Antibacterianos/farmacología , Modelos Animales de Enfermedad , Sistemas de Liberación de Medicamentos , Evaluación Preclínica de Medicamentos/métodos , Implantes de Medicamentos , Fluorodesoxiglucosa F18 , Masculino , Pruebas de Sensibilidad Microbiana , Osteomielitis/diagnóstico por imagen , Osteomielitis/microbiología , Poliglactina 910 , Tomografía Computarizada por Tomografía de Emisión de Positrones , Tomografía de Emisión de Positrones , Conejos , Infecciones Estafilocócicas/diagnóstico por imagen , Vancomicina/farmacologíaRESUMEN
BACKGROUND: The use of mesenchymal stem cells (MSCs) and coralline hydroxyapatite (HA) or biphasic calcium phosphate (BCP) as a bone substitute for posterolateral spinal fusion has been reported. However, the genes and molecular signals by which MSCs interact with their surrounding environment require further elucidation. METHODS: MSCs were harvested from bone grafting patients and identified by flow cytometry. A composite scaffold was developed using poly(lactide-co-glycolide) (PLGA) copolymer, coralline HA, BCP, and collagen as a carrier matrix for MSCs. The gene expression profiles of MSCs cultured in the scaffolds were measured by microarrays. The alkaline phosphatase (ALP) activity of the MSCs was assessed, and the expression of osteogenic genes and proteins was determined by quantitative polymerase chain reaction (Q-PCR) and Western blotting. Furthermore, we cultured rabbit MSCs in BCP or coralline HA hybrid scaffolds and transplanted these mixtures into rabbits for spinal fusion. We investigated the differences between BCP and coralline HA hybrid scaffolds by dual-energy X-ray absorptiometry (DEXA) and computed tomography (CT). RESULTS: Tested in vitro, the cells were negative for hematopoietic cell markers and positive for MSC markers. There was higher expression of 80 genes and lower of 101 genes of MSCs cultured in BCP hybrid scaffolds. Some of these genes have been shown to play a role in osteogenesis of MSCs. In addition, MSCs cultured in BCP hybrid scaffolds produced more messenger RNA (mRNA) for osteopontin, osteocalcin, Runx2, and leptin receptor (leptin-R) than those cultured in coralline HA hybrid scaffolds. Western blotting showed more Runx2 and leptin-R protein expression in BCP hybrid scaffolds. For in vivo results, 3D reconstructed CT images showed continuous bone bridges and fusion mass incorporated with the transverse processes. Bone mineral content (BMC) values were higher in the BCP hybrid scaffold group than in the coralline HA hybrid scaffold group. CONCLUSIONS: The BCP hybrid scaffold for osteogenesis of MSCs is better than the coralline HA hybrid scaffold by upregulating expression of leptin-R. This was consistent with in vivo data, which indicated that BCP hybrid scaffolds induced more bone formation in a spinal fusion model.
Asunto(s)
Diferenciación Celular/fisiología , Hidroxiapatitas/administración & dosificación , Células Madre Mesenquimatosas/metabolismo , Osteogénesis/fisiología , Receptores de Leptina/biosíntesis , Andamios del Tejido , Animales , Sustitutos de Huesos/administración & dosificación , Diferenciación Celular/efectos de los fármacos , Células Cultivadas , Regulación de la Expresión Génica , Humanos , Trasplante de Células Madre Mesenquimatosas/métodos , Osteogénesis/efectos de los fármacos , Prótesis e Implantes , Conejos , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiologíaRESUMEN
The authors investigated the lyophilized poly-L-lysine-coated alginate antibiotic delivery system in vivo for the treatment of musculoskeletal infections. The sodium alginate was mixed with vancomycin, coated with poly-L-lysine and lyophilized to form 3 mm in diameter biodegradable antibiotic beads. The antibiotic beads were implanted in the distal femoral cavities of rabbits for in vivo investigation. The local concentration of vancomycin was well above the minimal inhibitory concentration of Staphylococcus aureus for 21 days. The release was most marked during the first two days. The diameters of sample inhibition zone ranged from 8 to 16 mm, the relative activity of vancomycin ranged from 12.5% to 100%. The blood level of vancomycin reached its peak (46.0 mg/l) two days after implantation and fell to 3.2 mg/l at two weeks. It was undetectable after three weeks. There was no increase in the concentration of blood urea nitrogen and serum creatinine after the implantation. Histological observations showed that bead materials were biodegradable, resorbed slowly and only cause mild host reaction. This study offers a biodegradable delivery system of antibiotics to treat musculoskeletal infections.
Asunto(s)
Alginatos/administración & dosificación , Antibacterianos/administración & dosificación , Sistemas de Liberación de Medicamentos , Implantes de Medicamentos , Polilisina/análogos & derivados , Polilisina/administración & dosificación , Vancomicina/administración & dosificación , Animales , Enfermedades Óseas/tratamiento farmacológico , Masculino , Enfermedades Musculares/tratamiento farmacológico , Conejos , Vancomicina/sangre , Vancomicina/farmacologíaRESUMEN
The authors investigated poly (DL-lactide-co-glycolide) beads as an antibiotic delivery system in vivo for the treatment of various surgical infections. In this study, the copolymer 50:50 poly (DL-lactide):co-glycolide was mixed with vancomycin powder and hot compressing molded at 55 degrees C to form 8 mm in diameter biodegradable antibiotic beads. The antibiotic beads were implanted in the distal femoral cavities of rabbits for in vivo investigation. The local concentration of vancomycin was well above the breakpoint sensitivity concentration (the antibiotic concentration at the transition point between bacterial killing and resistance to the antibiotic) for 56 days. The release was most marked during the first day. The diameters of the sample inhibition zone ranged from 8 to 18 mm, and the relative activity of vancomycin ranged from 9.1% to 100%. Only low systemic blood levels of vancomycin were measured after beads implantation. There was no increase in the concentration of blood urea nitrogen and serum creatinine after the implantation. Histological observations showed that the bead materials were biodegradable, resorbed slowly, and did not cause a significant host reaction. This study offers a biodegradable delivery system of antibiotics to treat various surgical infections.
Asunto(s)
Antibacterianos/administración & dosificación , Poliglactina 910/administración & dosificación , Vancomicina/administración & dosificación , Animales , Bacterias/efectos de los fármacos , Nitrógeno de la Urea Sanguínea , Sistemas de Liberación de Medicamentos , Implantes de Medicamentos , Masculino , Complicaciones Posoperatorias/tratamiento farmacológico , Conejos , Vancomicina/farmacocinéticaRESUMEN
Proinflammatory cytokine, nitric oxide (NO) and localized hypoxia-induced apoptosis and proteoglycan (PG) degradation are thought to be correlated to the degree of cartilage injury. This study evaluated hyperbaric oxygen (HBO)-induced changes in joint cavity oxygen tension, antigenickeratan sulfate (KS) content, inducible nitric oxide synthase (iNOS) expression, PG synthesis, and cell apoptosis in full-thickness defects of rabbit cartilage. The HBO group was exposed to 100% oxygen at 2.5 atm for 2 h daily, 5 days per week. Meanwhile, the control group was kept in housing cages with normal air. The joint cavity oxygen tension was determined with an oxygen sensor. Blood serum KS was quantified by competitive indirect enzyme-linked immunosorbent assay (ELISA). After sacrifice, specimen sections were sent for histological and histochemical examination with a standardized scoring system. In situ analysis of iNOs expression and apoptosis detection were performed using immunostaining and TUNEL staining, respectively and quantified by a computerized imagine analysis system. This study demonstrated that HBO treatment increased joint cavity oxygen tension but decreased blood KS content. Histological and histochemical score results showed that HBO treatment significantly increased the cartilage repair. Moreover, immunostaining and TUNEL staining showed that HBO treatment suppressed the iNOs expression and apoptosis of chondrocytes, respectively. Accordingly, HBO offers a potential treatment method for cartilage injury.
Asunto(s)
Apoptosis , Cartílago Articular/metabolismo , Condrocitos/citología , Oxigenoterapia Hiperbárica , Óxido Nítrico/biosíntesis , Proteoglicanos/biosíntesis , Animales , Óxido Nítrico Sintasa/análisis , Óxido Nítrico Sintasa de Tipo II , ConejosRESUMEN
PURPOSE: Our aim was to determine the outcome of arthroscopic-assisted reduction with bilateral buttress plate fixation for the treatment of closed complex tibial plateau fractures. TYPE OF STUDY: Case series. METHODS: 18 consecutive patients (12 men, 6 women) with complex tibial plateau fractures were enrolled in this prospective study. All patients underwent arthroscopic-assisted bilateral buttress plate fixation of closed complex tibial plateau fractures. The average age at operation was 35 years (range, 23 to 45 years). The follow-up period ranged from 39 to 69 months, with an average of 48 months. Using the Schatzker classification, there were 11 type V and 7 type VI fractures. The clinical and radiological outcomes were determined according to Rasmussen's system. RESULTS: All of the 18 fractures united. Overall, 4 (22%) patients were rated as excellent, 12 (67%) good, and 2 (11%) fair. Secondary osteoarthritis appeared in 3 injured knees (16.7%). One patient had a wound dehiscence (3 cm long) of the medial incision. Condylar joint surface depression was noted in 3 patients without functional instability. Two patients had valgus alignment between 10 degrees and 15 degrees. Two patients had the paresthesia over the lateral calf. There were no complications directly associated with arthroscopy in any of the 18 patients. No deep vein thrombosis, infection, or knee stiffness was found at final follow-up. CONCLUSIONS: Arthroscopic-assisted reduction with bilateral buttress plate fixation for complex tibial plateau fractures allows accurate fracture reduction, diagnosis, and treatment of associated intra-articular lesions, and less dissection than open reduction internal fixation.
Asunto(s)
Placas Óseas , Fijación Interna de Fracturas/métodos , Fracturas Cerradas/cirugía , Fijadores Internos , Fracturas de la Tibia/cirugía , Adulto , Femenino , Fijación Interna de Fracturas/instrumentación , Humanos , Ligamentos Articulares/lesiones , Masculino , Persona de Mediana Edad , Traumatismo Múltiple/cirugía , Osteoartritis/etiología , Complicaciones Posoperatorias/etiología , Estudios Prospectivos , Rotura/cirugía , Lesiones de Menisco Tibial , Resultado del TratamientoRESUMEN
BACKGROUND: An in vivo animal study and a prospective clinical study have indicated that bone marrow aspirate (BMA) augments spinal arthrodesis. However, there is no quantified data to explain why fusion rate can be augmented by BMA in lumbar posterolateral fusion. METHODS: To analyze the proportion of mesenchymal stem cells (MSCs) and osteogenic factors in human BMA and peripheral blood (PB) of the same patient. Autologous BMA and PB from the patients were analyzed by flow cytometry (FACS) using cell markers for MSCs. The osteogenic potential of MSCs was determined by alkaline phosphatase (ALP) activity and calcium level quantification. Proteomics were used for the qualitative and quantitative mapping of the whole proteome from BMA and PB plasma. The mass-to-charge ratio was calculated by time-of-flight mass spectrometry (TOF-MS). The overexpression of protein was confirmed using Western blot analysis. RESULTS: The proportion of MSCs (CD34-/CD29+/CD105+) was higher in the BMA than that in the PB. Colony-forming cell (CFC) assays suggested that fewer colonies were formed in PB cultures than in BMA culture. There was no significant difference in the osteogenic potential of the MSCs between the PB and BMA. Proteomic mass spectrometry assays suggested that the levels of catalase (osteoclast inhibitor) and glutathione peroxidase 3 (osteogenic biomarker) were higher in the BMA than those in the PB, and this was confirmed by Western blot analysis. CONCLUSIONS: The proportions of MSCs and osteogenic factors were higher in the BMA than in the PB. This may explain why fusion rate can be augmented by BMA in lumbar posterolateral fusion.
Asunto(s)
Médula Ósea/fisiología , Citometría de Flujo/métodos , Células Madre Mesenquimatosas/fisiología , Osteogénesis/fisiología , Proteómica/métodos , Fusión Vertebral/métodos , Anciano , Secuencia de Aminoácidos , Femenino , Humanos , Leucocitos Mononucleares/fisiología , Masculino , Persona de Mediana Edad , Datos de Secuencia MolecularRESUMEN
We hypothesized that the effect of hyperbaric oxygen (HBO) on bone formation is increased via osteogenic differentiation of bone marrow stromal cells (BMSCs), which is regulated by Wnt3a/ß-catenin signaling. Our in vitro data showed that HBO increased cell proliferation, Wnt3a production, LRP6 phosphorylation, and cyclin D1 expression in osteogenically differentiated BMSCs. The mRNA and protein levels of Wnt3a, ß-catenin, and Runx2 were upregulated while those of GSK-3ß were downregulated after HBO treatment. The relative density ratio (phospho-protein/protein) of Akt and GSK-3ß was both up-regulated while that of ß-catenin was down-regulated after HBO treatment. We next investigated whether HBO affects the accumulation of ß-catenin. Our Western blot analysis showed increased levels of translocated ß-catenin that stimulated the expression of target genes after HBO treatment. HBO increased TCF-dependent transcription, Runx2 promoter/Luc gene activity, and the expression of osteogenic markers of BMSCs, such as alkaline phosphatase activity, type I collagen, osteocalcin, calcium, and the intensity of Alizarin Red staining. HBO dose dependently increased the bone morphogenetic protein (BMP2) and osterix production. We further demonstrated that HBO increased the expression of vacuolar-ATPases, which stimulated Wnt3a secretion from BMSCs. Finally, we showed that the beneficial effects of HBO on bone formation were related to Wnt3a/ß-catenin signaling in a rabbit model by histology, mechanical testing, and immunohistochemical assays. Accordingly, we concluded that HBO increased the osteogenic differentiation of BMSCs by regulating Wnt3a secretion and signaling.