目的 观察夏朴消瘿方(XPXYF)含药血清对碘化钠(NaI)诱导的人甲状腺上皮细胞(Nthy ori 3-1)焦亡的影响.方法 将Nthy ori 3-1细胞分为3组,空白组不予任何干预措施,模型组用NaI诱导细胞焦亡模型,实验组在造模的基础上加入含药血清培养.CCK-8法筛选含药血清最佳浓度,流式细胞术检测Nthy ori 3-1细胞焦亡,LDH法检测细胞毒性,细胞电镜观察Nthy ori 3-1细胞形态变化,流式细胞术检测氧化应激指标[活性氧(ROS)],Western blotting法检测细胞焦亡相关蛋白[焦亡蛋白D(GSDMD)、核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、Caspase-1、核因子κB(NF-κB)]表达.结果 与空白对照组比较,模型组细胞活力低(P<0.01);与模型组比较,实验组细胞活力高(P<0.01),实验组10%含药血清干预48 h,Nthy ori 3-1细胞活力最高.因此将10%含药血清用于后续研究.与空白对照组比较,模型组FLICA(+)-PI(+)双阳性比例高,焦亡率高(P<0.01).与模型组比较,实验组FLICA(+)-PI(+)双阳性比例低,焦亡率低(P<0.05).与空白对照组比较,模型组Nthy ori 3-1细胞LDH释放量多(P<0.01);与模型组比较,实验组细胞毒性少(P<0.01).实验组甲状腺滤泡细胞形态较规则,细胞核变小,胞质内空泡形成,空泡较模型组少,线粒体呈椭圆形,线粒体结构比模型组较完整.与空白对照组比较,模型组ROS水平高(P<0.01);与模型组比较,实验组ROS水平低(P<0.01).与空白对照组比较,模型组GSDMD、NF-κB p56蛋白表达高(P<0.05);与模型组比较,实验组GSDMD、Caspase-1、NF-κB p56、NLRP3蛋白表达低(P<0.05).结论 XPXYF含药血清可改善高碘诱导的Nthy ori 3-1细胞焦亡,可能与抑制氧化应激及激活GSDMD/NF-κB信号通路有关.
Effects of serum containing Xiapu Xiaoying Formula on pyroptosis of human thyroid epithelial cells induced by high iodine
Objective To observe the effect of serum containing Xiapu Xiaoying Formula(XPXYF)on sodium iodide(NaI)-induced pyroptosis in human thyroid epithelial cells(Nthy ori 3-1).Methods Nthy ori 3-1 cells were divided into three groups.The blank control group received no intervention,the model group was treated with NaI to establish a cell pyroptosis model,and the experimental group was cultured with drug-containing serum based on the established model.The optimal concentration of drug-containing serum was determined using the CCK-8 assay.Pyroptosis of Nthy ori 3-1 cells was detected using flow cytometry.Cytotoxicity was assessed,and morphological changes in Nthy ori 3-1 cells were observed under an electron microscope.Oxidative stress indicators[reactive oxygen species(ROS)]were measured by flow cytometry,and pyroptosis-related proteins[gasdermin D(GSDMD),nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3),Caspase-1,and nuclear factor κB(NF-κB)]were analyzed using Western blotting.Results Compared with the blank control group,cell viability in the model group was significantly reduced(P<0.01).Compared with the model group,cell viability in the experimental group was significantly increased(P<0.01),with the highest viability observed after 48 hours of treatment with 10%drug-containing serum.Therefore,10%drug-containing serum was used for subsequent studies.Compared with the blank control group,the proportion of FLICA(+)-PI(+)double-positive cells and the pyroptosis rate were significantly higher in the model group(P<0.01).Compared with the model group,the experimental group showed a significantly lower proportion of FLICA(+)-PI(+)double-positive cells and pyroptosis rate(P<0.05).Compared with the blank control group,LDH release in the model group was significantly higher(P<0.01),while cytotoxicity in the experimental group was significantly lower than that in the model group(P<0.01).Morphological observation revealed that thyroid follicular cells in the experimental group were more regular in shape,with smaller nuclei,fewer vacuoles in the cytoplasm compared to the model group,oval mitochondria,and a more intact mitochondrial structure.Compared with the blank control group,ROS levels in the model group were significantly elevated(P<0.01),whereas ROS levels in the experimental group were significantly reduced compared to the model group(P<0.01).Compared with the blank control group,the expression levels of GSDMD and NF-κB p56 proteins were significantly increased in the model group(P<0.05).Compared with the model group,the experimental group showed significantly reduced expression levels of GSDMD,Caspase-1,NF-κB p56,and NLRP3 proteins(P<0.05).Conclusion XPXYF-containing serum can alleviate NaI-induced pyroptosis in Nthy ori 3-1 cells,potentially through inhibition of oxidative stress and suppression of the GSDMD/NF-κB signaling pathway activation.
万方数据
维普
