蓝萼甲素激活巨噬细胞M2极化促进腰椎间盘突出重吸收OA

Objective To investigate the effect of glaucocalyxin A(GLA)on the resorption of lumbar disc herniation(LDH)and its molecular mechanisms.Methods The human monocytic leukemia cells THP-1 were divided into control group,PMA induction group,5 μmol/L GLA group,10 μmol/L GLA group and 20 μmol/L GLA group.The polarization levels of M1/M2 in cells were detected by immunofluorescence staining.Cells in each group were co-cultured with human nucleus pulposus cells in Transwell experiments.The proliferation activity of nucleus pulposus cells was detected by CCK-8 assay,and the apoptosis of nucleus pulposus cells was detected by TUNEL staining.The LDH rat model was established by autologous nucleus transplantation.The LDH rats were divided into LDH group,GLA(5 mg/kg)group and activator group(5 mg/kg GLA+5 mg/kg PMA),with 10 rats in each group,and ten unmodeled rats were set as control group.One day after the last administration,the mechanical pain threshold was determined by the Von Frey filament method,the levels of TNF-α,IL-1β,IL-6 and IL-10 in serum were detected by ELISA,the pathological and morphological changes of rat intervertebral disc tissues were detected by HE and Masson staining,the polarization of macrophages M1/M2 was detected by immuno-fluorescence staining,and the protein expression levels of MMP-3,TIMP-1,VEGF,COL Ⅱ,p-NF-κB p65,NF-κB p65,p-IκBα and IκBα were detected by Western blot.Results Compared with control group,the apoptosis rate of nucleus pulposus cells,the positive rate of iNOS,and the ratio of iNOS+/Arg1+were increased in PMA induction group(P<0.05),while the proliferation activity of nucleus pulposus cells and the positive rate of Arg1 were decreased(P<0.05).Compared with PMA induction group,the apoptosis rate of nucleus pulposus cells,the positive rate of iNOS,and the ratio of iNOS+/Arg1+in 5,10,20 μmol/L GLA groups were all decreased(P<0.05),and the proliferation activity of nucleus pulposus cells and the positive rate of Arg1 were all increased(P<0.05).Compared with LDH group,the damage of intervertebral disc structure was alleviated,the tissue pathological score was decreased in GLA group(P<0.05),the resorption of intervertebral disc tissue,mechanical pain threshold,IL-10 level,protein levels of COLⅡ and TIMP-1,and Arg1 positive rate were all increased(P<0.05),and the levels of TNF-α,IL-1β and IL-6,protein levels of MMP-3 and VEGF,iNOS positive rate,and the ratios of MMP-3/TIMP-1,iNOS+/Arg1+,p-NF-κB p65/NF-κB p65 and p-IκBα/IκBα were all decreased(P<0.05).The activator PMA reversed the improvement effect of GLA on the resorption of intervertebral disc protrusion.Conclusion GLA may promote the resorption of herniated intervertebral disc tissues by inhibiting the activation of NF-κB signaling pathway,activating M2-type polarization of macrophages,inhibiting the secretion of inflammatory factors and the apoptosis of nucleus pulposus cells,and promoting the synthesis of extracellular matrix of nucleus pulposus cells.