壳聚糖/甘油磷酸钠/海藻酸钠/益母草碱水凝胶的抗炎与促成骨作用  

Anti-inflammatory and osteogenic effects of chitosan/sodium glycerophosphate/sodium alginate/leonurine hydrogel

作　　者：赵增波[1] 李晨曦[2] 窦晨雷 马娜 周冠军[2] Zhao Zengbo;Li Chenxi;Dou Chenlei;Ma Na;Zhou Guanjun(Stomatological Hospital of Hebei Medical University,Shijiazhuang 050000,Hebei Province,China;Second Hospital,Hebei Medical University,Shijiazhuang 050000,Hebei Province,China)

机构地区：[1]河北医科大学口腔医院,河北省石家庄市050000 [2]河北医科大学第二医院,河北省石家庄市050000

出　　处：《中国组织工程研究》2025年第4期678-685,共8页Chinese Journal of Tissue Engineering Research

基　　金：河北省医学科学研究课题计划项目(20230621),项目负责人;周冠军。

摘　　要：背景:益母草碱具有改善微循环、抗氧化、抗细胞凋亡、清除自由基、抗炎、抗纤维化等多种生物活性作用,并且可促进骨髓间充质干细胞的成骨分化,具有应用于牙周炎治疗的潜能。目的:探讨益母草碱负载于壳聚糖/甘油磷酸钠/海藻酸钠水凝胶的抗炎与促成骨作用。方法:①分别制备壳聚糖/甘油磷酸钠/海藻酸钠水凝胶(空白水凝胶)与壳聚糖/甘油磷酸钠/海藻酸钠/益母草碱水凝胶(载药水凝胶),将两种水凝胶分别培养RAW 264.7细胞、MC3T3-E1细胞,利用CCK-8实验与活/死细胞染色检测水凝胶的细胞毒性。②将RAW 264.7细胞分5组培养:空白组常规培养24 h,脂多糖组加入脂多糖,单独水凝胶组加入脂多糖与空白水凝胶,载药水凝胶组加入脂多糖与载药水凝胶,抑制剂组加入脂多糖、载药水凝胶与PI3K抑制剂LY294002,处理24 h后,利用qRT-PCR法检测炎症相关因子mRNA表达,Western Blotting检测炎症相关因子与PI3K/AKT信号通路的蛋白表达。③将MC3T3-E1细胞分4组培养:空白组不加入任何材料,单独水凝胶组加入空白水凝胶,载药水凝胶组加入载药水凝胶,抑制剂组加入载药水凝胶与PI3K抑制剂LY294002,处理7 d后,进行碱性磷酸酶染色,利用qRT-PCR检测成骨相关因子mRNA表达水平,Western Blotting检测PI3K/AKT信号通路的蛋白表达。结果与结论:①CCK-8实验与活/死细胞染色结果显示,两种水凝胶无细胞毒性作用,具有良好的细胞相容性;②与空白组相比,脂多糖组白细胞介素6、肿瘤坏死因子α、白细胞介素1β的mRNA与蛋白表达升高(P<0.05),p-AKT、p-PI3K、p-p65、p-IκBα的蛋白表达升高(P<0.05);与脂多糖组比较,载药水凝胶组上述指标的mRNA与蛋白表达均降低(P<0.05);与载药水凝胶组相比,抑制剂组上述指标的mRNA与蛋白表达均降低(P<0.05);③载药水凝胶组碱性磷酸酶活性高于空白组、单独水凝胶组、抑制剂组(P<0.05);与空白组相比BACKGROUND:Leonurine has many biological activities such as improving microcirculation,anti-oxidation,anti-apoptosis,scavenging free radicals,antiinflammation,and anti-fibrosis,and can promote osteogenic differentiation of bone marrow mesenchymal stem cells,which has the potential to be applied in the treatment of periodontitis.OBJECTIVE:To explore the anti-inflammatory and osteogenic effects of leonurine loading into chitosan/sodium glycerophosphate/sodium alginate hydrogel.METHODS:(1)Chitosan/sodium glycerophosphate/sodium alginate hydrogel(blank hydrogel)and chitosan/sodium glycerophosphate/sodium alginate/leonurus alkali hydrogel were prepared respectively.RAW 264.7 and MC3T3-E1 cells were inoculated with the two kinds of hydrogel.The cytotoxicity of hydrogels was detected by CCK-8 assay and live/dead cell staining.(2)RAW 264.7 cells were cultured in five groups.The blank group was cultured for 24 hours routinely.The lipopolysaccharide group was treated with lipopolysaccharide.The simple hydrogel group was treated with lipopolysaccharide and blank hydrogel.The drugloaded hydrogel group was treated with lipopolysaccharide and drug-loaded hydrogel.The inhibitor group was treated with lippolysaccharide,drug-loaded hydrogel,and PI3K inhibitor LY294002.24 hours later,mRNA expression of inflammation-related factors was detected by qRT-PCR.Western blot assay was utilized to detect the protein expression of inflammation-related factors and PI3K/AKT signaling pathway.(3)MC3T3-E1 cells were inoculated in four groups.The blank group was cultured without any material.The simple hydrogel group was treated with blank hydrogel.The drug-loaded hydrogel group was treated with drug-loaded hydrogel.The inhibitor group was treated with drug-loaded hydrogel and PI3K inhibitor LY294002 for 7 days.Alkaline phosphatase staining was performed.mRNA expression levels of osteogenic factors were detected by qRT-PCR.The protein expression levels of the PI3K/AKT signaling pathway were detected by western blot assay.RESULTS AND CONCLUSION:(

关 键 词：牙周炎 水凝胶 壳聚糖 海藻酸钠 益母草碱 炎症因子 成骨作用 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学] R781.4