基于转录组测序技术探讨黄精芡实汤治疗糖尿病前期模型小鼠的机制研究

Mechanism of Huangjing Qianshi Decoction in treatment of prediabetic mice based on transcriptome sequencing

基于转录组测序技术对黄精芡实汤治疗的糖尿病前期模型小鼠进行测序,探究治疗糖尿病前期可能的机制。首先对BKS-DB小鼠正常组、糖尿病前期模型组(模型组)和黄精芡实汤治疗组(治疗组)进行转录组测序,获取小鼠骨骼肌标本中的差异表达基因,并对各组进行血清生化指标检测,筛选得到黄精芡实汤干预糖尿病前期的核心基因。运用基因本体论(Gene Ontology,GO)数据库和京都基因与基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)数据库对差异表达基因进行GO和KEGG信号通路富集分析,最后进行RT-qPCR验证。结果发现模型小鼠经黄精芡实汤治疗后,空腹血糖(fasting blood glucose,FBG)、空腹胰岛素(fasting insulin,FINS)、胰岛素抵抗指数(HOMA-IR)、血清总胆固醇(serum total cholesterol,TC)、甘油三脂(triglyceride,TG)、低密度脂蛋白胆固醇(low-density lipoprotein cholesterol,LDL-C)水平明显降低。差异基因筛选结果中,模型组与正常组相比共有1666个差异表达基因。治疗组与模型组相比共有971个差异表达基因。其中与胰岛素抵抗功能调节密切相关的IL-6、NR3C2基因在模型组和正常组之间显著上调,VEGFA基因在模型组和正常组之间显著下调,而在治疗组和模型组之间IL-6、NR3C2和VEGFA基因表达结果相反。GO功能富集分析发现,生物学过程注释中以细胞合成、周期和代谢过程为主,在细胞组分注释中以细胞内过程控制为主,在分子功能注释中以绑定分子功能为主。KEGG通路富集分析发现,涉及PTK6通路、CD28依赖的PI3K/AKT通路、p53途径等。因此,黄精芡实汤对糖尿病前期状态有改善作用,机制可能与IL-6、NR3C2和VEGFA调节的细胞周期、凋亡、PI3K/AKT通路、p53途径等生物学途径有关。

Based on transcriptome sequencing technology, the mouse model of prediabetes treated with Huangjing Qianshi Decoction was sequenced to explore the possible mechanism of treating prediabetes. First of all, transcriptome sequencing was performed on the normal BKS-DB mouse group, the prediabetic model group, and the Huangjing Qianshi Decoction treatment group(treatment group) to obtain differentially expressed genes in the skeletal muscle samples of mice. The serum biochemical indexes were detected in each group to screen out the core genes of Huangjing Qianshi Decoction in prediabetes. Gene Ontology(GO) database and Kyoto Encyclopedia of Genes and Genomes(KEGG) database were used to conduct signaling pathway enrichment analysis of differentially expressed genes, and real-time quantitative polymerase chain reaction(RT-qPCR) was used to verify them. The results showed that the levels of fasting blood glucose(FBG), fasting insulin(FINS), insulin resistance index(HOMA-IR), total cholesterol(TC), triglycerides(TG), and low-density lipoprotein cholesterol(LDL-C) in the mouse model were significantly decreased after treatment with Huangjing Qianshi Decoction. In the results of differential gene screening, there were 1 666 differentially expressed genes in the model group as compared with the normal group, and there were 971 differentially expressed genes in the treatment group as compared with the model group. Among them, interleukin-6(IL-6) and NR3C2 genes, which were closely related to the regulation of insulin resis-tance function, were significantly up-regulated between the model group and the normal group, and vascular endothelial growth factor A(VEGFA) genes were significantly down-regulated between the model group and the normal group. However, the expression results of IL-6, NR3C2, and VEGFA genes were adverse between the treatment group and the model group. GO functional enrichment analysis found that the biological process annotation mainly focused on cell synthesis, cycle, and metabolism;cell component annotation mainly focused on organelles and internal components;and molecular function annotation mainly focused on binding molecular functions. KEGG pathway enrichment analysis found that it involved the protein tyrosine kinase 6(PTK6) pathway, CD28-dependent phosphoinositide 3-kinase/protein kinase B(PI3K/AKT) pathway, p53 pathway, etc. Therefore, Huangjing Qianshi Decoction can improve the state of prediabetes, and the mechanism may be related to cell cycle and apoptosis, PI3K/AKT pathway, p53 pathway, and other biological pathways regulated by IL-6, NR3C2, and VEGFA.