艾灸对慢性萎缩性胃炎大鼠外周血基因表达谱的影响

Effects of Moxibustion on Gene Expression Profile in Peripheral Blood of Rats with Chronic Atrophic Gastritis

目的探讨隔药饼灸对慢性萎缩性胃炎大鼠外周血基因表达谱的影响,为后续研究提供研究方向。方法将雄性Wistar大鼠随机分成正常组、模型组、隔药饼灸组、电针组和西药组,均采用MNNG诱导CAG大鼠模型,取穴中脘、气海,隔药饼灸每次每穴1壮,每日1次,共4周;电针频率100 Hz,强度1-3 mA,20 min,每日1次,共4周;西药采用叶酸悬浊液1.0 mL/100 g灌胃,每天1次,持续4周。采用Trizol法提取各组CAG大鼠的外周血总RNA,质控后建立文库,采用RNA-seq高通量测序,结合生物信息学的方法深入分析隔药饼灸逆转的差异表达基因。结果与正常组比较,CAG组大鼠外周血有1542个差异表达基因,涉及的通路主要有癌症、免疫疾病、能量代谢、消化系统、免疫系统通路。隔药饼灸干预CAG大鼠的外周血差异表达基因2956个,涉及的通路与分子相互作用、信号转导、癌症、免疫疾病、氨基酸代谢、能量代谢、脂质代谢、维生素代谢、核苷酸代谢、消化系统、免疫系统、神经内分泌系统等相关;电针干预CAG外周血的差异表达基因3136个,涉及的通路与癌症、免疫疾病、氨基酸代谢、能量代谢、脂质代谢、维生素代谢、核苷酸代谢、消化系统、免疫系统、神经内分泌系统等相关;西药干预CAG外周血差异表达基因1308个,涉及的通路与蛋白翻译、信号转导、癌症、免疫系统、消化系统等相关。经RT-qPCR验证,各组外周血Foxo3、Uba52、S100a1、Nod2的结果和RNA-seq测序结果一致。结论隔药饼灸、电针和西药干预CAG的外周血差异表达基因的数量不同,西药较针灸改变CAG的差异基因数目少。3种干预方法均能影响CAG大鼠外周血免疫相关、肿瘤相关的基因表达谱,隔药饼灸和电针干预的差异基因还涉及能量、氨基酸、脂质、维生素代谢。

Objective To explore the effect of herb-partitioned moxibustion on gene expression profile in peripheral blood of rats with chronic atrophic gastritis, to provide direction for follow-up research. Methods Male Wistar rats were divided into normal group, model group, herb-partitioned moxibustion group, electro-acupuncture group and western medicine group randomly, CAG rat model induced by MNNG, acupoint selected by Zhongwan(CV12) and Qihai(CV17),one cone every time at each point once daily about 4 weeks for the herb-partitioned moxibustion group;frequency by100 Hz, current intensity by 1-3 mA, 20 min, once daily about 4 weeks for the electro-acupuncture group;1.0 ml/100 g folic acid suspension solution by gavage administration, once daily about 4 weeks for the western medicine group.Extraction of total RNA from peripheral blood of CAG rats by Trizol method, establishment of library after quality control, RNA-seq high throughput sequencing was combined with bioinformatics to analyze the differentially expressed genes reversed by herb-partitioned moxibustion. Results Compared with the normal group, there were 1542 differentially expressed genes in peripheral blood of rats in CAG group, KEGG analysis included cancer, immune diseases, energy metabolism, digestive system and immune system pathways. There were 2956 differentially expressed genes in peripheral blood of CAG rats treated with herb-partitioned moxibustion, pathways involved in molecular interactions,signal transduction, cancer, immune diseases, amino acid metabolism, energy metabolism, lipid metabolism, vitamin metabolism, nucleotide metabolism, digestive system, immune system, neuroendocrine system, etc.;there were 3136 differentially expressed genes in peripheral blood of CAG rats treated with electro-acupuncture, pathways involved in cancer, immune diseases, amino acid metabolism, energy metabolism, lipid metabolism, vitamin metabolism, nucleotide metabolism, digestive system, immune system, neuroendocrine system, etc.;there were 1308 differentially expressed genes in peripheral blood of CAG rats treated with western medicine, pathways involved in protein translation, signal transduction, cancer, immune system and digestive system. Verification by RT-q PCR, results of Foxo3, Uba52, S100 a1 and Nod2 in peripheral blood of each group were consistent with the RNA-seq sequencing. Conclusion There were differences in numbers of differentially expressed genes in peripheral blood of CAG about herb-partitioned moxibustion,electro-acupuncture and western medicine, the number of differentially expressed genes of CAG in western medicine is less than that acupuncture and moxibustion. Three intervention methods could affect the expression profiles of immunerelated and tumor-related genes in peripheral blood of CAG rats, differential genes also involve metabolism of energy,amino acids, lipids and vitamins by herb-partitioned moxibustion and electro-acupuncture.