RESUMEN
Steroidinduced avascular necrosis of the femoral head (SANFH) is a common orthopaedic disease that is difficult to treat. The present study investigated the effects of total flavonoids of Rhizoma drynariae (TFRD) on SANFH and explored its underlying mechanisms. The SANFH rat model was induced by intramuscular injection of lipopolysaccharides and methylprednisolone. Osteoblasts were isolated from the calvariae of neonatal rats and then cultured with dexamethasone (Dex). TFRD was used in vitro and in vivo, respectively. Haematoxylin and eosin staining was used to assess the pathological changes in the femoral head. Terminal deoxynucleotidyl transferasemediated deoxyuridine triphosphate nick end labelling assay and flow cytometry were conducted to detect apoptosis of osteoblasts. The 2',7'dichlorofluoresceindiacetate staining method was used to detect reactive oxygen species (ROS) levels in osteoblasts and the 3(4,5dimethylthiazol2yl)2,5diphenyltetrazolium bromide assay was used to detect osteoblast proliferation. The expression of caspase3, Bax, Bcl2, VEGF, runtrelated transcription factor 2 (RUNX2), osteoprotegerin (OPG), osteocalcin (OCN), receptor activator of nuclear factor κB ligand (RANKL) and phosphoinositide 3kinase (PI3K)/AKT pathway relatedproteins were detected via western blotting. It was found that TFRD reduced the pathological changes, inhibited apoptosis, increased the expression of VEGF, RUNX2, OPG and OCN, decreased RANKL expression and activated the PI3K/AKT pathway in SANFH rats. TFRD promoted proliferation, inhibited apoptosis and reduced ROS levels by activating the PI3K/AKT pathway in osteoblasts. In conclusion, TFRD protected against SANFH in a rat model. In addition, TFRD protected osteoblasts from Dexinduced damage through the PI3K/AKT pathway. The findings of the present study may contribute to find an effective treatment for the management of SANFH.
Asunto(s)
Flavonoides/farmacología , Osteonecrosis/tratamiento farmacológico , Extractos Vegetales/farmacología , Polypodiaceae/química , Animales , Proliferación Celular/efectos de los fármacos , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Modelos Animales de Enfermedad , Cabeza Femoral/patología , Flavonoides/química , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Masculino , Osteoblastos/efectos de los fármacos , Osteogénesis por Distracción/métodos , Osteonecrosis/inducido químicamente , Osteonecrosis/patología , Osteoprotegerina/genética , Fosfatidilinositol 3-Quinasas/genética , Extractos Vegetales/química , Proteínas Proto-Oncogénicas c-akt/genética , Ligando RANK/genética , Ratas , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Esteroides/efectos adversosRESUMEN
This study reports the use of diethylenetriamine as background electrolyte for the simultaneous separation of hyaluronan acid, chondroitin sulfate, dermatan sulfate and heparin. The analytes were baseline separated by using an uncoated fused silica capillary at 37°C with a run time of 23min. The migration order, with hyaluronan acid at first and heparin at last, was related to the sulfation degree. The effect of salt concentration on resolution and migration order was also investigated. The developed method was applied to the simultaneous determination of hyaluronan acid and chondroitin sulfate in mouse plasma. Interferences in plasma were removed by protein precipitation and glycosaminoglycans were further purified by ethanol precipitation. The method was validated over the concentration range from 50 to 600µg/mL for hyaluronan acid and 500 to 6000µg/mL for chondroitin sulfate in mouse plasma. Results from assay validations showed that the method was selective and robust.