Research Of Protective Effects And Mechanism Of Achyranthes Bidentata Bl. Saponins On Chondrocytes

Osteoarthritis (OA) is a chronic and degenerative joint disease. Conservativetreatment for osteoarthritis is based on nonsteroial anti-inflammatory drugs. Thesedrugs not only alleviate early symptoms, relief pain and improve the joint function,but also can induce gastrointestinal adverse effects and chondrocytes destruction.Achyranthes bidentata Bl., which belongs to the amaranthaceous plant family, haslong been used in Traditional Chinese Medicine to nourish liver and kiney, strengthenthe bones and muscles, purse blood stasis and stimulate the menstrual flow, ensureproper downward flow of the blood. Accumulative evidence has demonstrated thatmany plants from Achyranthes bidentata Bl. species possess anti-inflammatory andanalgesic activities, and triterpenoid saponins might be the main active components.The aim of the present study was to investigate the effect of Achyranthes bidentata Bl.saponins (ABS) on rat OA induced by monosodium iodoacetate (MIA) andinterleukin-1β (IL-1β). Furthermore, the protective effects of ABS on chondrocyteswere examined in an attempt to exploit the mechanisms of action of ABS.Achyranthes bidentata Bl. slices were extracted with75%ethanol under refluxand N-butanol, evaporated in vacuo, the residue was diluted with distilled water andpurified with D101macroporous absorptive resin. The50%ethanol eluted fractionwas collected to yield a yellowish-brown powder. Results from the effects of differentchromogenic agents on determination of content of ABS by UV spectrophotometrysuggested5%vanillin-glacial acetic acid: perchloric acid:glacial acetic acid (2:8:50)was the best chromogenic method. The maximum absorption wavelength of themethod was at548±1nm, peak pattern and sample recovery rare were better thanothers. The total content of saponin was52.45%.Chondrocytes were isolated from the knee joints of100±20g Sprague-Dawleyrats by enzymatic digestion.10ng/mL IL-1β suppressed chondrocytes viability,3、10、30μg/mL ABS up-regulated cell viability in a dose-dependent manner. Hoechst33258staining and Annexin V-FITC/PI dual staining demonstrated ABS could protectIL-1β-induced chondrocytes injury. Western blotting assays presented ABS couldsuppress IL-1β-induced apoptosis by inhibiting levels of Bax and Bad, decreasing p53phosphorylation and promoting expression of Bcl-xLand PCNA. Simultaneously,ABS inhibited caspase3activity. IL-1β-induced inflammation and matrix degrationwere also alleviated by ABS with down-regulating expression of MMP3, MMP9andCOX-2. Moreover, ABS inhibited NF-κB p65phosphorylation, IκBα degradation andphosphorylation induced by IL-1β in rat chondrocytes.The rats were divided randomly into six groups: normal control, model control,(50,100,200mg/kg) three concentrations of ABS,(75mg/mL) glucosamine sulfateas positive control. The joint cavity of the rats received a single injection of1mg MIAexcept normal control instead of equal volume physiological saline. ABS (50,100,200mg/kg) and glucosamine sulfate (75mg/kg) dministered orally in a volume of10mL/kg once a day for28days from the next day of arthritis induction. After28days,the rats were decapitated and picked up blood. Then cartilage samples were separated.The results showed that (100,200mg/kg) ABS inhibited acute swelling of knee joint,degradation of proteoglycan and collagen, serum IL-1β and NO level, in ratsexperimental OA model.The findings demonstrate that ABS is effective in the treatment of OA in rat.The mechanisms involve with inhibiting PG and collagen degradation in vivo,preventing chondrocyte from apoptosis and inflammation by inhibiting NF-κB signalpathway in vitro.