基于HDAC6调控NLRP3转运的生骨再造丸对BMSCs焦亡水平影响的探讨  

Exploring the effect of Shenggu Zaizao Pill on the pyroptosis level of BMSCs through regulating NLRP3 transport based on HDAC6

作　　者：尚征亚 曹林忠[1,2] SHANG Zheng-ya;CAO Lin-zhong(Gansu University of Chinese Medicine,Lanzhou 730000,China;Affiliated Hospital of Gansu University of Chinese Medicine,Lanzhou 730000,China)

机构地区：[1]甘肃中医药大学,甘肃兰州730000 [2]甘肃中医药大学附属医院,甘肃兰州730000

出　　处：《时珍国医国药》2025年第1期9-16,共8页Lishizhen Medicine and Materia Medica Research

基　　金：国家自然科学基金(82160915,81860859)。

摘　　要：目的从组蛋白去乙酰化酶6(HDAC6)调控NOD样受体热蛋白结构域相关蛋白3(NLRP3)转运的角度,探讨生骨再造丸对骨髓间充质干细胞(BMSCs)焦亡水平的影响。方法购买原代兔BMSCs,制备焦亡BMSCs模型,采用HDAC6激动剂(Theophylline)与抑制剂(Trichostain A)对焦亡的BMSCs进行干预;制备生骨再造丸含药血清,对焦亡的BMSCs进行干预。将上述细胞分为空白组、模型组、模型组+HDAC6激动剂组(模型组+Theophylline)、模型组+HDAC6抑制剂组(模型组+Trichostain A)、模型组+生骨再造丸含药血清组。采用CCK-8法检测BMSCs活力及含药血清对BMSCs的最佳干预时间;采用PCR法检测各组BMSCs中膜孔蛋白D(GSDMD)的基因水平;采用WB法检测各组BMSCs中天冬氨酸特异性半胱氨酸蛋白酶-1(Caspase-1)、GSDMD、NLRP3蛋白表达;采用ELISA法检测各组BMSCs中HDAC6、白细胞介素1β(IL-1β)、白细胞介素18(IL-18)含量。结果与空白组相比,模型组细胞GSDMD mRNA水平上升(P<0.05),NLRP3、Caspase-1、GSDMD蛋白水平显著升高(P<0.05),HDAC6含量明显升高(P<0.05),IL-18、IL-1β水平上升(P<0.05)。与模型组相比,模型组+HDAC6激动剂组中细胞GSDMDmRNA水平上升(P<0.05),NLRP3、Caspase-1、GSDMD蛋白水平明显升高(P<0.05),HDAC6含量明显升高(P<0.05),IL-18、IL-1β水平上升(P<0.05);模型组+HDAC6抑制剂组中细胞GSDMD mRNA水平下降(P<0.05),NLRP3、Caspase-1、GSDMD蛋白水平降低(P<0.05),HDAC6含量降低(P<0.05),IL-18、IL-1β水平降低(P<0.05);模型组+生骨再造丸含药血清组中细胞GSDMDmRNA水平明显下降(P<0.05),NLRP3、Caspase-1、GSDMD蛋白水平明显降低(P<0.05),HDAC6含量明显降低(P<0.05),IL-18、IL-1β水平明显降低(P<0.05)。结论生骨再造丸通过抑制HDAC6的活性,抑制NLRP3炎症小体的激活,可以有效改善BMSCs焦亡水平。Objective To explore the effect of Shenggu Zaizao Pill(SZP)on the pyroptosis level of bone mesenchymal stem cells(BMSCs)from the perspective of histone deacetylase 6(HDAC6)in regulating the transport of NOD-like receptor thermal protein domain associated protein 3(NLRP3).Methods Primary rabbit BMSCs were purchased to prepare the pyroptosis model of BMSCs.HDAC6 agonist(Theophylline)and inhibitor(Trichostain A)were used to intervene the pyrogenic BMSCs.SZP containing serum was prepared to intervene the pyrogenic BMSCs.These cells were divided into the blank group,the model group,the model group+HDAC6 agonist group(model group+Theophylline),the model group+HDAC6 inhibitor group(model group+Trichostain A),and model group+SZP containing serum group.The CCK-8 method was used to detect the viability of BMSCs and the optimal intervention time of drug-containing serum on BMSCs.The PCR method was exploited to measure the gene level of Gasdermin D(GSDMD)in BMSCs.Western blot(WB)was adopted to detect the protein expression of cysteine-containing aspartate-specific proteases-1(Caspase-1),GSDMD,and NLRP3 in BMSCs of different groups.The ELISA was utilized to detect the levels of HDAC6,interleukin-1β(IL-1β),and interleukin-18(IL-18)in BMSCs.Results Compared with the blank group,the level of GSDMD mRNA in the model group was increased(P<0.05),and the protein levels of NLRP3,Caspase-1 and GSDMD were significantly increased(P<0.05).Additionally,the content of HDAC6 was significantly increased(P<0.05),and the levels of IL-18 and IL-1β were elevated(P<0.05).Compared with model group,the level of GSDMD mRNA in model group+HDAC6 agonist group was increased(P<0.05),and the protein levels of NLRP3,Caspase-1 and GSDMD and the content of HDAC6 were significantly increased(P<0.05).The levels of IL-18 and IL-1β were increased(P<0.05).In model group+HDAC6 inhibitor group,the level of GSDMD mRNA,the protein levels of NL⁃RP3,Caspase-1 and GSDMD,the content of HDAC6,and the levels of IL-18 and IL-1β were decreased(P<0.05).The level of GS

关 键 词：生骨再造丸 SANFH 焦亡 HDAC6 NLRP3 

分 类 号：R285.5[医药卫生—中药学]