摘要
目的研究生物活性肽Epi-1在脂多糖(LPS)诱导下的炎症微环境中对人牙髓干细胞(hDPSC)表达炎症因子的影响和可能机制。方法通过组织块酶消化法分离、培养hDPSC,并通过流式细胞术鉴定。利用CCK-8试剂检测Epi-1对hDPSC细胞活性的影响,筛选出适宜的浓度。实验分为4组,分别为空白对照组(不含LPS和Epi-1)、LPS组(1.0μg/mL LPS)、2.5μg/mL Epi-1组(1.0μg/mL LPS和2.5μg/mL Epi-1)和5.0μg/mL Epi-1组(1.0μg/mL LPS和5.0μg/mL Epi-1),通过实时荧光定量聚合酶链式反应检测Epi-1对hDPSC白介素(IL)-6、IL-1β、IL-8基因表达的影响,进一步通过蛋白质免疫印迹法检测核因子κB信号通路关键蛋白p65、p-p65的表达情况,使用荧光探针检测活性氧生成情况。采用SPSS 22.0分析数据。结果在2.5、5.0μg/mL的质量浓度下,Epi-1无明显细胞毒性;Epi-1可显著降低LPS诱导后hDPSC中IL-6、IL-1β、IL-8的mRNA表达(P<0.01),减少p-p65的表达(P<0.05)和活性氧的生成(P<0.001)。结论Epi-1可能通过抑制核因子κB信号通路的激活和减轻氧化应激反应,从而降低LPS诱导的hDPSC的炎症反应。
Objective To explore the effect of Epi-1 on the expression of inflammatory cytokines in human dental pulp stem cell(hDPSC)induced by lipopolysaccharide(LPS),and to explore the underlying mechanism.Methods hDPSC were cultured by tissue block enzyme digestion method and identified by flow cytometry.CCK-8 assay were used to determine the cell viability of hDPSC to screen out the appropriate concentration of Epi-1.hDPSC were randomly divided into 4 groups:blank group(without LPS and Epi-1),LPS group(1.0μg/mL LPS),2.5μg/mL Epi-1 group(1.0μg/mL LPS+2.5μg/mL Epi-1),5.0μg/mL Epi-1 group(1.0μg/mL LPS+5.0μg/mL Epi-1).The gene levels of IL-1β,IL-6 and IL-8 in hDPSC were detected by real time qPCR.The change of nuclear factor kappa-B signaling pathways were detected by Western blotting and ROS generation was detected by fluorescence probe.SPSS 22.0 software was applied for statistical analysis.Results Epi-1 with concentrations of 2.5μg/mL and 5.0μg/mL had no significant cytotoxicity on hDPSC.Compared with LPS group,2.5μg/mL and 5.0μg/mL Epi-1 group were significantly reduced LPS-induced expression of IL-1β,IL-6 and IL-8 at gene levels(P<0.01),inhibited the expression of p-p65 protein(P<0.05)and reduced the production of ROS(P<0.001).Conclusion Epi-1 could alleviate the inflammatory response of hDPSC induced by LPS by inhibiting the activation of nuclear factorκB pathway and reducing the oxidative stress reaction.
作者
罗宇仪
赵望泓
梁悦娥
Luo Yuyi;Zhao Wanghong;Liang Yuee(Department of Stomatology,Nanfang Hospital,Southern Medical University,Guangzhou 510515,China;Department of Pediatric Dentistry,Foshan Stomatological Hospital,Foshan 528099,Guangdong,China)
出处
《兰州大学学报（医学版）》
2024年第10期13-19,35,共8页
Journal of Lanzhou University（Medical Sciences）
基金
南方医科大学南方医院院长基金资助项目(2020C028)
南方医科大学南方医院“临床研究专项”资助项目(2020CR029)
广州市科技计划重点资助项目(2023B0J1253)
南方医科大学“临床研究启动计划”重点资助项目(LC2019ZD023)。
关键词
人牙髓干细胞
炎症
Epi-1
核因子ΚB
牙髓炎
生物活性肽
human dental pulp stem cell
inflammation
Epi-1
nuclear factor kappa-B
pulpitis
bioactive peptides
作者简介
通信作者:梁悦娥,女,主治医师,研究方向为牙体牙髓疾病防治,e-mail:yueeliang734@sina.com。
