Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, No.725 South Wanping Road, Shanghai, 200032, China.
School of Public Health, Shanghai University of Traditional Chinese Medicine, Shanghai, 201203, China.
BMC Complement Altern Med. 2019 Jul 3;19(1):156. doi: 10.1186/s12906-019-2577-6.
The traditional Chinese medicine prescription, Qianggan formula have been confirmed to be effective on non-alcoholic steatohepatitis (NASH), however, the underlying molecular mechanisms remain obscure.
Thirty-six male C57BL/6 mice were randomly divided into three groups: normal chow diet group; methionine-and-choline-deficient diet (MCD) group, and Qianggan extract (QG) intervention group (0.4 g/kg daily) that fed with MCD. The efficacy of QG was biochemically and histologically evaluated. The expression profiles of messenger ribonucleic acids (mRNAs), long non-coding RNAs (lncRNAs) and circular RNAs (circRNAs) were examined using microarray and verified by RT-qPCR.
QG significantly improved the phenotypic characteristics of NASH, as serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), and lactate dehydrogenase (LDH) levels and liver inflammatory cytokines were significantly decreased. By the cutoff of a 1.5-fold change and P < 0.05, 6193 mRNAs, 5692 lncRNAs and 4843 circRNAs were identified as differentially expressed between the MCD and normal groups, and 514 mRNAs, 1182 lncRNAs and 443 circRNAs were identified as differentially expressed between the QG and MCD groups. The intersections (244 mRNAs, 259 lncRNAs and 98 circRNAs) among the three groups were chosen for analysis. Gene Ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment revealed that most overlapping mRNAs were related to immune functions such as natural-killer-cell-mediated cytotoxicity, intestinal immune network for IgA production, and T cell receptor signaling pathway. Pathway interactions, protein-protein interactions and molecular complex detection (MCODE) analysis identified numerous immune-related hub genes e.g. natural cytotoxicity triggering receptor 1(Ncr1), C-X-C motif chemokine ligand 9 (Cxcl9), Klra1, and Cd28. Finally, two lncRNAs (Sngh1 and Slc36a3os) and four circRNAs (circ_0009029, circ_0004572, circ_0009212 and circ_0009453) in competing endogenous RNA (ceRNA) networks were constructed by Cytoscape, and immune-related mRNAs (e.g., Cd28, Cd8a, Il15, and Klrk1) were involved in the ceRNA networks.
LncRNA and circRNA-associated immune ceRNA networks might be the targets of QG in alleviating NASH, and our work may provide valuable clues for exploring the mechanisms underlying the effect of QG.
中药方剂强肝方已被证实对非酒精性脂肪性肝炎（NASH）有效，但潜在的分子机制仍不清楚。
36 只雄性 C57BL/6 小鼠随机分为三组：正常饮食组；蛋氨酸和胆碱缺乏饮食（MCD）组，以及强肝提取物（QG）干预组（每天 0.4 g/kg），喂食 MCD。通过生化和组织学评估 QG 的疗效。使用微阵列检测信使核糖核酸（mRNA）、长非编码 RNA（lncRNA）和环状 RNA（circRNA）的表达谱，并通过 RT-qPCR 进行验证。
QG 显著改善了 NASH 的表型特征，血清丙氨酸氨基转移酶（ALT）、天冬氨酸氨基转移酶（AST）、碱性磷酸酶（ALP）和乳酸脱氢酶（LDH）水平以及肝炎症细胞因子明显降低。通过 1.5 倍变化和 P < 0.05 的截止值，在 MCD 和正常组之间鉴定出 6193 个 mRNA、5692 个 lncRNA 和 4843 个 circRNA 差异表达，在 QG 和 MCD 组之间鉴定出 514 个 mRNA、1182 个 lncRNA 和 443 个 circRNA 差异表达。三个组之间的交集（244 个 mRNA、259 个 lncRNA 和 98 个 circRNA）被选择用于分析。基因本体论（GO）术语和京都基因与基因组百科全书（KEGG）途径富集显示，大多数重叠的 mRNA 与免疫功能有关，如自然杀伤细胞介导的细胞毒性、肠道免疫网络中 IgA 的产生以及 T 细胞受体信号通路。途径相互作用、蛋白质-蛋白质相互作用和分子复合物检测（MCODE）分析鉴定了许多免疫相关的枢纽基因，如自然细胞毒性触发受体 1（Ncr1）、C-X-C 基序趋化因子配体 9（Cxcl9）、Klra1 和 Cd28。最后，通过 Cytoscape 构建了竞争性内源性 RNA（ceRNA）网络中的两个 lncRNA（Sngh1 和 Slc36a3os）和四个 circRNA（circ_0009029、circ_0004572、circ_0009212 和 circ_0009453），并涉及免疫相关的 mRNA（如 Cd28、Cd8a、Il15 和 Klrk1）ceRNA 网络。
lncRNA 和 circRNA 相关的免疫 ceRNA 网络可能是 QG 缓解 NASH 的靶点，我们的工作可能为探索 QG 作用机制提供有价值的线索。
