Low-intensity Pulsed Ultrasound Inhibits VEGFA Expression In Chondrocytes And Protects Against Cartilage Degeneration In Experimental Osteoarthritis

Objective: Osteoarthritis(OA)is the most common joint degenerative disease.As the population ages,the prevalence rate of OA increasing year by year.The patients manifested as pain and limited joint activity,and even disabled,which seriously affects the patient’s quality of life.The basic pathological features of osteoarthritis include progressive articular cartilage degeneration,synovial inflammation,subchondral bone sclerosis and osteophyte formation.Articular cartilage is a special kind of tissue without blood vessels,and there is currently no effective treatment.Chondrocytes are the most important cell in it,which maintain the metabolic balance of extracellular matrix by secreting and degrading the cartilage matrix.The state is maintained and regulated by various growth factors,such as cytokines and mechanical stimulation.Low-intensity pulsed ultrasound(LIPUS)has been used clinically for more than 20 years.It is a non-invasive and safe physical factor rehabilitation treatment method.Its functions include fracture healing and soft tissue injury.At present,more and more studies have shown thatLIPUS can relieve the pain of OA patients and improve their daily life ability,but the related mechanism is still not very clear.However,vascular endothelium growth factor(VEGFA)plays an important role in the development of arthritis.In normal articular cartilage,seldom VEGFA expressed,and it can even be considered as a promoting factor of OA.The expression of VEGFA in articular cartilage,synovium,and synovial fluid in OA patients is considered to be correlated with the severity of OA and pain.Therefore,reducing the VEGFA level of articular cartilage may be a target for OA treatment.Accordingly,this study explored the role of LIPUS in regulating the expression of VEGFA of chondrocytes in alleviating experimental OA,and its possible molecular mechanism.Methods:(1)We established OA model,using IL-1β-induced chondrocytes in vitro,and LIPUS intervened: a.qPCR was used to detect the VEGFA and cartilage matrix degradation key molecules such as MMP-13 mRNA levels;b.ELISA was used to detect VEGFA protein levels in chondrocyte culture medium of each group;c.Western blot was used to detect main molecules of p38 MAPK and JNK signaling pathways in each group.(2)We also used the right knee joints of 10-week-old C57 BL / 6 male mouse to establish a destabilization of the medial meniscus(DMM)traumatic arthritis model,further divided the mice after surgery into DMM group and DMM + LIPUS group randomly,the left knee joints wereconsidered as Sham operation group.a.Immunohistochemical staining of paraffin section specimens of knee joints to detect the expression changes of VEGFA and cartilage matrix related factors such as MMP-13,Agrrecan,Col-(47)in articular cartilage of each group;b.Articular paraffin sections were stained with Safranin-O/Fast Green,and the stained articular cartilage was scored by OARSI.Results:(1)LIPUS can directly reduce the mRNA levels of VEGFA,MMP-13 and the marker of hypertrophy Col-X in OA chondrocytes(p<0.05),while relieving IL-1β reduced down-regulation of Col-(47)(p <0.001).(2)LIPUS can down-regulate the protein lever of VEGFA secreted by OA chondrocytes(p<0.05).(3)LIPUS inhibits the activation of p38 MAPK signaling pathway in OA chondrocytes.Blocked this pathway,its down-regulation effect on VEGFA of chondrocyte is markedly weakened.(4)LIPUS can reduce the expression of VEGFA in articular cartilage after DMM(p <0.001)and alleviate the loss of cartilage matrix(p <0.05).(5)After LIPUS treatment,the OARSI score of DMM-operated mice was significantly reduced(p <0.01).Conclusion: LIPUS can directly reduce the expression of VEGFA in osteoarthritis chondrocytes,and delay the progress of experimental arthritis in mice.This effect depends on p38 MAPK signaling pathway.