Abstract：
BACKGROUND: Synovial inflammation plays a crucial role in osteoarthritis (OA). Gastrodin (GAS), an active ingredient derived from the Gastrodia elata Blume rhizome, possesses antioxidant and anti-inflammatory pharmacological effects. This research aimed to evaluate the function and molecular mechanism of GAS on human fibroblast-like synoviocytes of osteoarthritis (HFLS-OA) induced by interleukin (IL)-1β.
METHODS: The impact of GAS on the viability of IL-1β-treated HFLS-OA cells was assessed using the cell counting kit-8 (CCK-8). Quantitative real-time reverse transcription PCR (qRT-PCR) was employed to detect changes in IL-8, IL-6, monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor (TNF)-α, and Gremlin-1 mRNA expression in each group. Corresponding kits were utilized to measure the catalase (CAT) and superoxide dismutase (SOD) activities, as well as the nitric oxide (NO) level. Western blot analysis was conducted to examine the expression of extracellular matrix degradation-associated proteins and nuclear factor kappa-B (NF-κB) pathway-correlated proteins in each group.
RESULTS: GAS significantly promoted the proliferation of IL-1β-induced HFLS-OA cells and concurrently down-regulated Gremlin-1 mRNA expression (p < 0.05). Through the down-regulation of Gremlin-1 expression, GAS exhibited the following effects: decreased IL-8, IL-6, and TNF-α mRNA expression, as well as NO levels (p < 0.05); increased SOD and CAT activities (p < 0.05); down-regulated matrix metallopeptidase 13 (MMP-13) and MMP-1 protein expression levels (p < 0.01); and up-regulated collagen II protein expression level (p < 0.01) in IL-1β-treated HFLS-OA cells. Additionally, GAS decreased phospho-inhibitory kappa B (p-IκB)/IκB, phospho-inhibitory kappa B kinase (p-IKK)/IKK, and p-p65/p65 ratios in IL-1β-induced HFLS-OA cells by inhibiting Gremlin-1 expression (p < 0.01).
CONCLUSIONS: GAS demonstrates a positive impact on inflammation, oxidative stress, and extracellular matrix degradation in IL-1β-mediated HFLS-OA cells. This effect is achieved by suppressing Gremlin-1 expression and reducing NF-κB pathway activity.
METHODS: The impact of GAS on the viability of IL-1β-treated HFLS-OA cells was assessed using the cell counting kit-8 (CCK-8). Quantitative real-time reverse transcription PCR (qRT-PCR) was employed to detect changes in IL-8, IL-6, monocyte chemotactic protein-1 (MCP-1), tumor necrosis factor (TNF)-α, and Gremlin-1 mRNA expression in each group. Corresponding kits were utilized to measure the catalase (CAT) and superoxide dismutase (SOD) activities, as well as the nitric oxide (NO) level. Western blot analysis was conducted to examine the expression of extracellular matrix degradation-associated proteins and nuclear factor kappa-B (NF-κB) pathway-correlated proteins in each group.
RESULTS: GAS significantly promoted the proliferation of IL-1β-induced HFLS-OA cells and concurrently down-regulated Gremlin-1 mRNA expression (p < 0.05). Through the down-regulation of Gremlin-1 expression, GAS exhibited the following effects: decreased IL-8, IL-6, and TNF-α mRNA expression, as well as NO levels (p < 0.05); increased SOD and CAT activities (p < 0.05); down-regulated matrix metallopeptidase 13 (MMP-13) and MMP-1 protein expression levels (p < 0.01); and up-regulated collagen II protein expression level (p < 0.01) in IL-1β-treated HFLS-OA cells. Additionally, GAS decreased phospho-inhibitory kappa B (p-IκB)/IκB, phospho-inhibitory kappa B kinase (p-IKK)/IKK, and p-p65/p65 ratios in IL-1β-induced HFLS-OA cells by inhibiting Gremlin-1 expression (p < 0.01).
CONCLUSIONS: GAS demonstrates a positive impact on inflammation, oxidative stress, and extracellular matrix degradation in IL-1β-mediated HFLS-OA cells. This effect is achieved by suppressing Gremlin-1 expression and reducing NF-κB pathway activity.
摘要：
背景：滑膜炎症在骨关节炎（OA）中起着至关重要的作用。天麻素(GAS),一种来自天麻根茎的活性成分，具有抗氧化和抗炎的药理作用。本研究旨在探讨GAS对白细胞介素(IL)-1β诱导的人成纤维样骨关节炎滑膜细胞(HFLS-OA)的作用及分子机制。
方法：使用细胞计数试剂盒-8（CCK-8）评估GAS对IL-1β处理的HFLS-OA细胞活力的影响。定量实时逆转录PCR（qRT-PCR）检测IL-8，IL-6，单核细胞趋化蛋白-1（MCP-1）的变化，肿瘤坏死因子（TNF）-α，各组Gremlin-1mRNA表达。使用相应的试剂盒来测量过氧化氢酶（CAT）和超氧化物歧化酶（SOD）的活性，以及一氧化氮（NO）水平。Westernblot检测各组细胞外基质降解相关蛋白和核因子κB(NF-κB)通路相关蛋白的表达。
结果：GAS显著促进IL-1β诱导的HFLS-OA细胞的增殖，同时下调Gremlin-1mRNA的表达（p<0.05）。通过下调Gremlin-1的表达,GAS表现出以下作用：IL-8，IL-6和TNF-αmRNA表达降低，以及NO水平（p<0.05）；增加SOD和CAT活性（p<0.05）；下调IL-1β处理的HFLS-OA细胞中基质金属肽酶13（MMP-13）和MMP-1蛋白表达水平（p<0.01）；上调胶原II蛋白表达水平（p<0.01）。此外,GAS降低磷酸化抑制性κB（p-IκB）/IκB，磷酸化抑制性κB激酶(p-IKK)/IKK,通过抑制Gremlin-1表达在IL-1β诱导的HFLS-OA细胞中的p-p65/p65比值(p<0.01)。
结论：GAS显示出对炎症的积极影响，氧化应激，IL-1β介导的HFLS-OA细胞的细胞外基质降解。这种作用是通过抑制Gremlin-1表达和降低NF-κB途径活性来实现的。
方法：使用细胞计数试剂盒-8（CCK-8）评估GAS对IL-1β处理的HFLS-OA细胞活力的影响。定量实时逆转录PCR（qRT-PCR）检测IL-8，IL-6，单核细胞趋化蛋白-1（MCP-1）的变化，肿瘤坏死因子（TNF）-α，各组Gremlin-1mRNA表达。使用相应的试剂盒来测量过氧化氢酶（CAT）和超氧化物歧化酶（SOD）的活性，以及一氧化氮（NO）水平。Westernblot检测各组细胞外基质降解相关蛋白和核因子κB(NF-κB)通路相关蛋白的表达。
结果：GAS显著促进IL-1β诱导的HFLS-OA细胞的增殖，同时下调Gremlin-1mRNA的表达（p<0.05）。通过下调Gremlin-1的表达,GAS表现出以下作用：IL-8，IL-6和TNF-αmRNA表达降低，以及NO水平（p<0.05）；增加SOD和CAT活性（p<0.05）；下调IL-1β处理的HFLS-OA细胞中基质金属肽酶13（MMP-13）和MMP-1蛋白表达水平（p<0.01）；上调胶原II蛋白表达水平（p<0.01）。此外,GAS降低磷酸化抑制性κB（p-IκB）/IκB，磷酸化抑制性κB激酶(p-IKK)/IKK,通过抑制Gremlin-1表达在IL-1β诱导的HFLS-OA细胞中的p-p65/p65比值(p<0.01)。
结论：GAS显示出对炎症的积极影响，氧化应激，IL-1β介导的HFLS-OA细胞的细胞外基质降解。这种作用是通过抑制Gremlin-1表达和降低NF-κB途径活性来实现的。
