骨碎补总黄酮调控Notch1/Hes1信号轴促进软骨细胞自噬并抑制凋亡的机制研究OA
Regulation of Notch1/Hes1 signaling axis by total flavonoids of Drynariae Rhizoma for promoting chondrocyte autophagy and inhibiting apoptosis:a mechanistic study
目的 探究骨碎补总黄酮(TFRD)调控Notch1/发状分裂相关增强子1(Hes1)信号轴对脂多糖(LPS)诱导的软骨细胞自噬及凋亡的影响.方法 采用5 μg/mL LPS处理人软骨细胞系C28/I2细胞构建体外炎症损伤模型,将细胞分为正常对照组、模型组、TFRD组(200 μg/mL)、TFRD+过氧化物氧化还原酶1(Prdx1)小干扰RNA(si-Prdx1)组和TFRD+si-Prdx1阴性对照(si-NC)组,每组设6个复孔.细胞以si-Prdx1或si-NC转染24 h、TFRD预处理2 h,再以LPS处理,总计培养48 h.检测细胞凋亡率、凋亡细胞占比、单丹磺酰尸胺(MDC)荧光强度和基质金属蛋白酶13(MMP-13)、含Ⅰ型血小板结合蛋白基序的解聚蛋白样金属蛋白酶5(ADAMTS5)、软骨寡聚基质蛋白(COMP)含量以及X连锁凋亡抑制蛋白(XIAP)、多腺苷二磷酸核糖聚合1(PARP1)、Beclin-1、微管相关蛋白1轻链3Ⅱ/Ⅰ(LC3-Ⅱ/Ⅰ)、PTEN诱导激酶1(PINK1)、Notch1、Hes1、Prdx1蛋白表达情况.结果 与模型组比较,TFRD组细胞凋亡率、凋亡细胞占比和MMP-13、ADAMTS5含量以及PARP1蛋白表达水平均显著降低,MDC荧光强度、COMP含量和XIAP、Beclin-1、LC3-Ⅱ/Ⅰ、PINK1、Notch1、Hes1、Prdx1蛋白表达水平均升高(P＜0.05).与TFRD+si-NC组比较,TFRD+si-Prdx1组细胞中除Notch1、Hes1以外上述指标的变化均被显著逆转(P＜0.05).结论 TFRD可能通过激活Notch1/Hes1信号轴上调下游靶分子Prdx1的表达,进而抑制LPS诱导的软骨细胞凋亡、促进其保护性自噬,从而改善软骨代谢稳态.
OBJECTIVE To investigate the effects of total flavonoids of Drynariae Rhizoma(TFRD)on autophagy and apoptosis in LPS-induced chondrocytes via the regulation of the Notch1/hairy and enhancer of split 1(Notch1/Hes1)signaling axis.METHODS Human chondrocyte cell line C28/I2 cells were cultured with 5 μg/mL LPS to establish in vitro inflammatory injury model.The cells were separated into normal control group,model group,TFRD group(200 μg/mL),TFRD+peroxiredoxin 1(Prdx1)small interfering RNA(si-Prdx1)group and TFRD+si-Prdx1 negative control(si-NC)group,with 6 replicate wells in each group.Cells were transfected with si-Prdx1 or si-NC for 24 hours,pretreated with TFRD for 2 hours,and then exposed to LPS,with a total culture duration of 48 hours.Apoptotic rate,the proportion of apoptotic cells,monodansylcadaverine(MDC)fluorescence intensity,as well as the contents of matrix metalloproteinase-13(MMP-13),a disintegrin and metalloproteinase with thrombospondin motifs 5(ADAMTS5),and cartilage oligomeric matrix protein(COMP)were measured.Additionally,the protein expression levels of X-linked inhibitor of apoptosis protein(XIAP),poly(ADP-ribose)polymerase 1(PARP1),Beclin-1,microtubule-associated protein 1 light chain 3 Ⅱ/Ⅰ(LC3-Ⅱ/Ⅰ),PTEN-induced putative kinase 1(PINK1),Notch1,Hes1,and Prdx1 were assessed.RESULTS Compared with model group,the apoptotic rate,the proportion of apoptotic cells,the contents of MMP-13 and ADAMTS5 as well as protein expressions of PARP1 were significantly decreased,while MDC fluorescence intensity,COMP content,protein expressions of XIAP,Beclin-1,LC3-Ⅱ/Ⅰ,PINK1,Notch1,Hes1 and Prdx1 were significantly increased(P<0.05).Compared with TFRD+si-NC group,the changes in the aforementioned indicators(except for Notch1 and Hes1)in the cells of the TFRD+si-Prdx1 group were significantly reversed(P<0.05).CONCLUSIONS TFRD may activate the Notch1/Hes1 signaling axis,and up-regulate the expression of the downstream target molecule Prdx1,thereby inhibiting LPS-induced chondrocyte apoptosis,promoting protective autophagy,and consequently improving cartilage metabolic homeostasis.
鲁林;方虹
武汉市中医医院(湖北中医药大学附属国医医院)骨伤科,武汉 430006湖北省中医院(湖北中医药大学附属医院)妇科,武汉 430006
医药卫生
骨碎补总黄酮软骨细胞Notch受体1/Hes1信号轴自噬凋亡
total flavonoids of Drynariae RhizomachondrocyteNotch1/Hes1 signaling axisautophagyapoptosis
《中国药房》 2026 (8)
1027-1032,6
湖北省中医药管理局中医药科研项目(No.ZY2025L063)
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