基于运脾法拆方研究异功散治疗慢性病贫血的机制

Study on the mechanism of Yigong Powder in the treatment of anemia of chronic disease by means of formula components analysis based on activating spleen method

目的观察陈皮在异功散改善慢性病贫血铁代谢、促进红细胞生成中的作用。方法将异功散进行拆方分为陈皮单药和陈皮缺味方,采用2×2析因设计方法,将32只酵母糖诱导的全身炎症反应模型小鼠按随机数字表法分为模型组、陈皮组、陈皮缺味方组和异功散全方组,每组8只。陈皮组(3.083 g/kg)、陈皮缺味方组(12.33 g/kg)和异功散全方组(15.413 g/kg)小鼠分别灌胃相应药物,模型组小鼠灌胃等量生理盐水,连续7 d。给药结束后,记录各组小鼠体质量,采用全自动细胞计数仪检测各组小鼠血红蛋白含量,比色法检测血清铁含量,酶联免疫吸附测定(ELISA)法检测血清铁蛋白含量,计算脾指数,Luminex法检测肝脏组织炎症因子白细胞介素-1β(IL-1β)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、γ干扰素(IFN-γ)、白细胞介素-4(IL-4)和白细胞介素-10(IL-10)水平,实时荧光PCR法检测肝脏组织铁调素基因(HAMP)和膜铁转运蛋白(Fpn)的mRNA表达。结果陈皮组和陈皮缺味方组在调节血红蛋白、血清铁、血清铁蛋白含量,改善脾指数,调节HAMP、Fpn mRNA及IL-1β、IFN-γ表达方面均呈现交互作用(均P<0.05)。与模型组比较,陈皮组血红蛋白、血清铁含量及Fpn mRNA表达水平升高,铁蛋白含量、脾指数、HAMP mRNA及IL-1β、IL-6、TNF-α、IFN-γ水平降低(均P<0.05);陈皮缺味方组血清铁含量、Fpn mRNA表达水平升高,脾指数、铁蛋白含量、HAMP mRNA及IL-1β、IFN-γ、IL-4水平降低(均P<0.05)。结论陈皮对促炎症因子IL-6、TNF-α的抑制作用及对Fpn的上调作用可能在异功散全方改善慢性病贫血及铁代谢中发挥了关键作用。

Objective To observe the role of dried tangerine peel in Yigong Powder improves iron metabolism and promotes red blood cell generation in anemia of chronic disease(ACD).Methods With a two-by-two factorial design,the Yigong Powder was divided into dried tangerine peel and Chenpi absent Decoction.According to the random number table method,32 zymosan-induced generalized inflammation(ZIGI)mice were randomly divided into the model group,the dried tangerine peel group,the Chenpi absent Decoction group,and the Yigong Powder group.The dried tangerine peel group,Chenpi absent Decoction group and the Yigong Powder group were given dried tangerine peel(3.083 g/kg),Chenpi absent Decoction(12.33 g/kg),and Yigong Powder(15.413 g/kg)by gavage to the corresponding group of mice.The model group was given an equal amount of physiological saline by gavage,and treated continuously for 7 days.After the completion of administration,the body weight of each group of mice was recorded.The hemoglobin content of each group of mice was detected using a fully automatic cell counter,the serum iron content was detected using colorimetry,the serum ferritin content was detected using enzyme-linked immunosorbent assay(ELISA),and the spleen index was calculated.The liver tissue inflammatory factors interleukin-1β(IL-1β),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),interferon-γ(IFN-γ),interleukin-4(IL-4),and interleukin-10(IL-10)levels were detected using Luminex method.The mRNA expressions of liver tissue hepcidin gene(HAMP)and membrane iron transporter(Fpn)were detected using real-time fluorescence PCR method.Results Dried tangerine peel and Chenpi absent Decoction both showed interactive effects in regulating hemoglobin,serum iron,serum ferritin content,improving spleen index,and regulating the mRNA expressions of HAMP,Fpn,as well as IL-1βand IFN-γ(P<0.05).Compared with the model group,dried tangerine peel significantly increased hemoglobin,serum iron content,and Fpn mRNA expression in ZIGI model mice,while decreasing ferritin content,spleen index,HAMP mRNA expression,and the levels of IL-1β,IL-6,TNF-α,and IFN-γ(P<0.05).Chenpi absent Decoction significantly increased serum iron content and Fpn mRNA expression in ZIGI model mice,while reducing spleen index,ferritin content,HAMP mRNA expression,and the levels of IL-1βand IFN-γ、IL-4(P<0.05).Conclusion The effects of dried tangerine peel on inflammatory factors(IL-6 and TNF-α)and Fpn may play a key role in the improvement effects of Yigong Powder on ACD and iron metabolism.