摘要
目的基于细胞焦亡NOD样受体热蛋白结构域相关蛋白3(NLRP3)/含半胱氨酸的天冬氨酸蛋白水解酶1(Caspase-1)/消皮素D(GSDMD)信号通路,以实验性自身免疫性甲状腺炎(Experimental autoimmune thyroiditis,EAT)模型大鼠为研究对象,探究逍遥散加味治疗桥本氏甲状腺炎(Hashimoto’s thyroiditis,HT)的作用机制。方法40只SD大鼠随机分配空白组10只、造模组30只。造模组经皮下多点注射猪甲状腺球蛋白佐剂免疫乳结合饮用高碘水建立EAT大鼠模型。成模后随机分为模型组10只、逍遥散加味组10只及硒酵母组10只。逍遥散加味组予7.65 g/(kg·d)逍遥散加味溶液,硒酵母组予36μg/(kg·d)硒酵母片水溶液,余两组均予蒸馏水等体积灌胃。8周后,HE染色观察EAT大鼠甲状腺组织形态结构;透射电子显微镜下观察EAT大鼠甲状腺滤泡上皮细胞超微结构;酶联免疫吸附测定法(Enzym-linkedimmunosorbent assay,ELISA)检测EAT大鼠血清甲状腺过氧化物酶抗体(Thyroid peroxidase antibody,TPOAb)、甲状腺球蛋白抗体(Thyroglobulin antibody,TgAb)水平、白细胞介素-1β(Interleukin-1β,IL-1β)、白细胞介素-18(Interleukin-18,IL-18)水平;Western Blot检测EAT大鼠甲状腺组织细胞焦亡相关蛋白表达。结果(1)HE染色及透射电镜结果示:模型组大鼠甲状腺组织存在明显的病理损伤,甲状腺滤泡上皮细胞膜结构模糊、破损,线粒体明显肿胀,膜结构模糊、溶解,粗面内质网扩张;各治疗组的大鼠甲状腺病理损伤程度均减轻,线粒体与内质网损伤程度均有减轻,其中逍遥散加味组减轻程度最为明显。(2)ELISA检测可见:模型组大鼠血清TgAb、TPOAb抗体及血清IL-1β、IL-18水平较空白组呈显著升高趋势(P<0.01),各治疗组均能降低大鼠血清TPOAb、TgAb抗体及血清IL-1β、IL-18水平(P<0.01,P<0.05),逍遥散加味组下降尤甚。(3)WB检测各组焦亡相关蛋白表达水平所示:模型组大鼠甲状腺组织NLRP3、Cleaved-Caspase-1、GSDMD-N、凋亡相关的斑点样蛋白(ASC)蛋白的表达水平均呈显著升高趋势(P<0.01),余治疗组均显著下降(P<0.01,P<0.05),以逍遥散加味组降低程度最为明显。结论逍遥散加味减轻甲状腺组织的病理损伤,降低TPOAb、TgAb抗体水平,抑制细胞焦亡状态,其作用机制可能与调控NLRP3/Caspase-1/GSDMD信号通路相关。
Objective Based on the NOD like receptor heat protein domain associated protein 3(NLRP3)/cysteine containing aspartic protease 1(Caspase-1)/deskininoid D(GSDMD)signaling pathway,experimental autoimmune thyroiditis(EAT)model rats were used as the research subjects to explore the mechanism of action of Modified Xiaoyao Powder(逍遥散加味)in the treat-ment of Hashimoto's thyroiditis(HT).Methods Forty SD rats were randomly assigned to a blank group(10 rats)and a model group(30 rats).The EAT rat model was established by subcutaneous multi-point injection of pig thyroglobulin adjuvant immu-noemulsion combined with drinking high iodine water.After modeling,the rats were randomly divided into a model group(10 rats),a Modified Xiaoyao Powder group(10 rats),and a selenium yeast group(10 rats).The Modified Xiaoyao Powder group was given 7.65 g/(kg·d)Modified Xiaoyao Powder solution,the selenium yeast group was given 36μg/(kg·d)selenium yeast tab-let aqueous solution,and the remaining two groups were given distilled water by gavage of equal volume.After 8 weeks,HE stai-ning was used to observe the morphological structure of thyroid tissue in EAT rats.The ultrastructure of thyroid follicular epitheli-al cells in EAT rats was observed under transmission electron microscopy.Enzym-linkedimmunosorbent assay(ELISA)was used to detect the levels of serum thyroid peroxidase antibody(TPO Ab),thyroid globulin antibody(TgAb),interleukin-1β(IL-1β),and interleukin-18(IL-18)in EAT rats.Western Blot was used to detect the expressions of apoptosis related pro-teins in thyroid tissue cells of EAT rats.Results①The results of HE staining and transmission electron microscopy showed that there was obvious pathological damage in the thyroid tissue,the membrane structure of thyroid follicular epithelial cells was blurred and damaged,the mitochondria were obviously swollen,the membrane structure was blurred and dissolved,and the rough endoplasmic reticulum was dilated.The degree of thyroid pathological damage,mitochondrial and endoplasmic reticulum damage in all treatment groups was reduced,and the degree of reduction was the most obvious in the Modified Xiaoyao Powder group.②ELISA showed that the serum levels of TgAb,TPOAb antibody,and serum levels of IL-1βand IL-18 in the model group were significantly higher than those in the blank group(P<0.01),and the serum levels of TPOAb,TgAb antibody,IL-1βand IL-18 in each treatment group were reduced(P<0.01,P<0.05),especially in the Modified Xiaoyao Powder group.③As shown by the expression level of pyroptosis-related proteins in each group by WB,the expression levels of NLRP3,GSDMD,Caspase-1 and apoptosis-associated speck-like protein containing a CARD(ASC)protein in the thyroid tissue of rats increased significant-ly(P<0.01),and the remaining treatment groups decreased significantly(P<0.01,P<0.05),with the decrease being the most obvious in the Modified Xiaoyao Powder group.Conclusion Modified Xiaoyao Powder alleviates the pathological damage of thyroid tissue,reduces the levels of TPOAb and TgAb antibodies,and inhibits the pyroptosis state,which may be related to the regulation of NLRP3/Caspase-1/GSDMD signaling pathway.
作者
赵培伯
戚晓楠
贾春艳
张兰
ZHAO Peibo;QI Xiaonan;JIA Chunyan;ZHANG Lan(Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning,China;Affiliated Hospital of Liaoning University of Traditional Chinese Medicine,Shenyang 110032,Liaoning,China)
出处
《中华中医药学刊》
北大核心
2025年第11期55-59,I0008,共6页
Chinese Archives of Traditional Chinese Medicine
基金
国家自然科学基金青年科学基金项目(82305275)
国家中医药管理局全国名老中医药专家传承工作室项目(国中医药人教函[2022]75号)。
作者简介
赵培伯(1991-),女,辽宁盖县人,主治医师,博士在读,研究方向:中西医结合内分泌;通讯作者:张兰(1962-),女,辽宁沈阳人,教授,博士研究生导师,博士,研究方向:中西医结合内分泌。E-mail:zhanglanzhu@163.com。
