ObjectiveTo investigate the effect of Yanghetang （YHT） on the treatment of osteoarthritis （OA） and its mechanism.MethodsIn vivo experiments： After 1 week of acclimatization feeding， 30 C57BL/6J mice were randomly divided into normal group， model group， low and high dose groups of YHT （8.3， 16.6 g·kg^-1）， and celecoxib group （18 mg·kg^-1）， with six mice in each group. Except for the normal group， the remaining groups were constructed with the OA model by right-sided joint meniscus destruction surgery. After successful modeling， the normal group and the model group were given an equal amount of saline gavage daily， and each treatment group was given the corresponding dose of the drug， respectively. The intervention lasted for 4 weeks. Micro-computed tomography （Micro-CT） was used to scan the right knee of the sampled mice. Alisin blue （ABH） and hematoxylin-eosin （HE） staining were used to observe the pathologic morphology of synovium and cartilage in mice. Enzyme-linked immunosorbent assay （ELISA） was used to detect the effect of YHT on the serum levels of interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α） inflammatory factors in mice， and immunohistochemistry was employed to observe the relevant pathways' expression. In vitro experiments： RAW264.7 cells were induced with lipopolysaccharides （LPSs） to establish an M1-type inflammation model， and 1%， 3%， and 5% YHT were added for intervention， and the above macrophage supernatant （CM） was collected and added to the chondrocyte culture medium for intervention， respectively. High performance liquid chromatography （HPLC） was used to determine the content of YHT for compositional analysis. Cell counting kit-8 （CCK-8） was used to detect the effect of YHT on cell viability. Immunofluorescence and flow cytometry were adopted to detect the degree of macrophage M1/M2 type polarization， and Annexin V apoptosis detection kit was used to detect the degree of apoptosis of chondrocytes. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western bolt were utilized to determine the expression of relevant proteins and mRNAs in RAW264.7 cells and chondrocytes.ResultsCompared with those of the normal group， the bone-related parameters such as BV/TV， Tb.N， and Tb.Th were significantly down-regulated （P<0.01）， and Tb.Sp was significantly elevated （P<0.01） in mice of the model group， with significant pathological changes in synovium and cartilage， and the levels of inflammatory factors in serum including IL-6 and TNF-α， were significantly elevated （P<0.01）. The viability of chondrocytes was significantly reduced， while the apoptosis rate was significantly higher （P<0.01）， and the average fluorescence intensity of CD86 in RAW264.7 cells was significantly improved （P<0.01）. Real-time PCR results showed that the levels of iNOS， IL-6， and IL-1β mRNA in RAW264.7 cells and chondrocytes were significantly higher （P<0.05， P<0.01）， and Arg-1， IL-10， and CD206 mRNA levels were significantly decreased （P<0.01）. Western blot assay revealed that the protein phosphorylation levels of PI3K， Akt， and NF-κB were significantly up-regulated in the LPS group （P<0.01）， and the MMP-13 expression was up-regulated， but the Col-2 protein expression was down-regulated （P<0.05， P<0.01）. Compared with the model group， YHT groups significantly improved bone-related parameters in mice （P<0.01）， significantly reduced synovial cell thickness and cartilage surface abrasion， attenuated serum levels of IL-6 and TNF-α in mice （P<0.01）， increased chondrocyte survival， attenuated chondrocyte apoptosis （P<0.01）， and significantly attenuated M1-type RAW264.7 cell polarization （P<0.01）. Real-time PCR results showed that iNOS， IL-6， and IL-1β mRNA levels in RAW264.7 cells and chondrocytes were significantly decreased （P<0.05， P<0.01）， and Arg-1， IL-10， and CD206 mRNA levels were significantly up-regulated （P<0.05， P<0.01）. Western blot assay revealed that YHT groups significantly down-regulated the protein phosphorylation levels of PI3K， Akt， and NF-κB （P<0.05， P<0.01）， simultaneously inhibited the expression of MMP-13， and up-regulated Col-2 protein expression （P<0.05， P<0.01）.ConclusionYHT significantly improved osteoarthritis caused by synovial macrophage inflammation. The pathway may be related to the inhibition of the PI3K/Akt/NF-κB signaling pathway.

soupe de soleil et d'air; arthrose; macrophages; chondrocytes; étude de mécanisme