摘要
目的观察银连含漱液对脂多糖(LPS)诱导人牙龈成纤维细胞(HGFs)炎症反应、氧化应激及细胞凋亡的影响。方法收集临床患者的健康牙龈组织,采用组织块法培养HGFs并采用免疫荧光法进行细胞鉴定。采用CCK-8法检测不同体积分数银连含漱液(2.5%、5%、10%、20%、40%)、不同浓度绿原酸(0.01、0.1、0.5、1、10mg·mL^(-1))及LPS(1、10、20、35、50μg·mL^(-1))分别与HGFs共培养24、48、72h后的增殖变化情况。依据CCK-8检测结果,将HGFs分为4组:DMEM对照组、1μg·mL^(-1) LPS组、1μg·mL^(-1) LPS+20%银连含漱液组、1μg·mL^(-1) LPS+1 mg·mL^(-1)绿原酸组。干预24 h后,采用流式细胞术检测HGFs的凋亡水平;WST-1法检测细胞超氧化物歧化酶(SOD)活性;ELISA法检测细胞上清液中炎性因子肿瘤坏死因子α(TNF-α)、白细胞介素6(IL-6)的含量。结果选择对HGFs无明显毒性作用的1 mg·mL^(-1)绿原酸、20%银连含漱液和1μg·mL^(-1) LPS作用24 h。与对照组比较,LPS组的HGFs凋亡率显著升高(P<0.001),SOD活力显著降低(P<0.001),上清液中的IL-6、TNF-α水平显著升高(P<0.001)。与LPS组比较,LPS+20%银连含漱液组及LPS+1 mg·mL^(-1)绿原酸组的HGFs凋亡率显著降低(P<0.001),SOD活力显著升高(P<0.01,P<0.001),上清液中的IL-6、TNF-α水平显著降低(P<0.001)。与LPS+20%银连含漱液组比较,LPS+1 mg·mL^(-1)绿原酸组的HGFs凋亡率显著降低(P<0.001),SOD活力显著升高(P<0.01),上清液中的IL-6、TNF-α水平显著降低(P<0.05,P<0.01)。结论银连含漱液可抑制LPS诱导的HGFs细胞凋亡,提高其抗氧化能力,降低炎症反应,该作用可能与活性成分绿原酸密切相关。
Objective To study the effects of Yinlian Gargle on inflammation,oxidative stress response,and apoptosis of human gingival fibroblasts(HGFs)stimulated by lipopolysaccharide(LPS).Methods Healthy gingival samples were collected from the donors.HGFs were cultured by tissue-explant technique and identified by immunofluorescence assay.CCK-8 method was used to observe the effects of different volume fractions(2.5%,5%,10%,20%,40%)of Yinlian Gargle,different concentrations of chlorogenic acid(0.01,0.1,0.5,1,10 mg·mL^(-1))and different concentrations of LPS(1,10,20,35,50μg·mL^(-1))on cell proliferation of HGFs at different time duration(24,48,72 h).According to CCK-8 results,HGFs were divided into four groups:DMEM control group,1μg·mL^(-1) LPS group,1μg·mL^(-1) LPS+20%Yinlian Gargle group,and 1μg·mL^(-1) LPS+1 mg·mL^(-1)chlorogenic acid group.After 24 h of intervention,flow cytometry,WST-1 assay and enzyme-linked immunosorbent assay(ELISA)were used to test apoptosis of HGFs,superoxide dismutase(SOD)activity,TNF-αand IL-6 levels.Results No significant toxic effect on HGFs was observed by 24 h treatment of 20%of Yinlian Gargle,1 mg·mL^(-1) of chlorogenic acid and 1μg·mL^(-1) of LPS,which were chosen for subsequent experiments.Compared to the control group,the apoptosis rate of HGFs in LPS group significantly increased(P<0.001),SOD activity significantly decreased(P<0.001),while the levels of IL-6 and TNF-αin the supernates significantly increased(P<0.001).Compared to LPS group,the apoptosis rate of 1μg·mL^(-1) LPS+20%Yinlian Gargle group,and 1μg·mL^(-1) LPS+1 mg·mL^(-1)chlorogenic acid group significantly decreased(P<0.001),SOD activities significantly increased(P<0.01,P<0.001),while the levels of IL-6 and TNF-αin the supernates significantly decreased(P<0.001).Compared to 1μg·mL^(-1) LPS+20%Yinlian Gargle group,the apoptosis rate of HGFs in 1μg·mL^(-1) LPS+1 mg·mL^(-1)chlorogenic acid group significantly decreased(P<0.001),SOD activity significantly increased(P<0.01),while the levels of IL-6 and TNF-αin the supernates significantly decreased(P<0.05,P<0.01).Conclusion Yinlian Gargle could inhibit LPS-induced apoptosis of HGFs,enhance antioxidant capacity,reduce inflammatory response,which may be closely related to chlorogenic acid.
作者
齐刘英
洪楠锐
刘伟南
QI Liuying;HONG Nanrui;LIU Weinan(Department of Stomatology,the First Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangdong Clinical Research Academy of Chinese Medicine,Guangzhou 510405 Guangdong,China)
出处
《中药新药与临床药理》
北大核心
2025年第1期17-23,共7页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
广东省中医药局科研项目(20231168)。
关键词
银连含漱液
绿原酸
人牙龈成纤维细胞
脂多糖
细胞凋亡
炎症反应
氧化应激
Yinlian Gargle
chlorogenic acid
human gingival fibroblasts
lipopolysaccharide
cell apoptosis
inflammatory response
oxidative stress
