Department of Anesthesiology & Center for Brain Science, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, 710061, Shaanxi, China.
Shaanxi University of Chinese Medicine, Xianyang, Shaanxi, China.
Eur J Med Res. 2023 Sep 12;28(1):340. doi: 10.1186/s40001-023-01321-1.
The exact mechanisms of type 2 diabetes mellitus (T2DM) remain largely unknown. We intended to authenticate critical genes linked to T2DM progression by tandem single-cell sequencing and general transcriptome sequencing data.
T2DM single-cell RNA-sequencing data were submitted by the Gene Expression Omnibus (GEO) database and ArrayExpress (EBI), from which gene expression matrices were retrieved. The common cell clusters and representative marker genes were ascertained by principal component analysis (PCA), t-distributed stochastic neighbor embedding (t-SNE), CellMarker, and FindMarkers in two datasets (GSE86469 and GSE81608). T2DM-related differentially expressed marker genes were defined by intersection analysis of marker genes and GSE86468-differentially expressed genes. Receiver operating characteristic (ROC) curves were utilized to assign representative marker genes with diagnostic values by GSE86468, GSE29226 and external validation GSE29221, and their prospective target compounds were forecasted by PubChem. Besides, the R package clusterProfiler-based functional annotation was designed to unveil the intrinsic mechanisms of the target genes. At last, western blot was used to validate the alternation of CDKN1C and DLK1 expression in primary pancreatic islet cells cultured with or without 30mM glucose.
Three common cell clusters were authenticated in two independent T2DM single-cell sequencing data, covering neurons, epithelial cells, and smooth muscle cells. Functional ensemble analysis disclosed an intimate association of these cell clusters with peptide/insulin secretion and pancreatic development. Pseudo-temporal trajectory analysis indicated that almost all epithelial and smooth muscle cells were of neuron origin. We characterized CDKN1C and DLK1, which were notably upregulated in T2DM samples, with satisfactory availability in recognizing three representative marker genes in non-diabetic and T2DM samples, and they were also robustly interlinked with the clinical characteristics of patients. Western blot also demonstrated that, compared with control group, the expression of CDKN1C and DLK1 were increased in primary pancreatic islet cells cultured with 30 mM glucose for 48 h. Additionally, PubChem projected 11 and 21 potential compounds for CDKN1C and DLK1, respectively.
It is desirable that the emergence of the 2 critical genes indicated (CDKN1C and DLK1) could be catalysts for the investigation of the mechanisms of T2DM progression and the exploitation of innovative therapies.
2 型糖尿病（T2DM）的确切发病机制尚不清楚。我们拟通过串联单细胞测序和全转录组测序数据，鉴定与 T2DM 进展相关的关键基因。
从基因表达综合数据库（GEO）和 ArrayExpress（EBI）中提交 T2DM 单细胞 RNA 测序数据，从中获取基因表达矩阵。通过主成分分析（PCA）、t 分布随机邻域嵌入（t-SNE）、CellMarker 和 FindMarkers 在两个数据集（GSE86469 和 GSE81608）中确定常见的细胞簇和代表性标记基因。通过交集分析标记基因和 GSE86468 差异表达基因，定义 T2DM 相关差异表达标记基因。利用 GSE86468、GSE29226 和外部验证 GSE29221 的 ROC 曲线为 GSE86468 分配具有诊断价值的代表性标记基因，并预测其潜在的靶化合物PubChem。此外，基于 R 包 clusterProfiler 的功能注释设计，以揭示靶基因的内在机制。最后，采用 Western blot 验证在 30mM 葡萄糖培养的原代胰岛细胞中 CDKN1C 和 DLK1 的表达变化。
在两个独立的 T2DM 单细胞测序数据中鉴定了 3 个常见的细胞簇，包括神经元、上皮细胞和平滑肌细胞。功能集分析揭示了这些细胞簇与肽/胰岛素分泌和胰腺发育密切相关。拟时序轨迹分析表明，几乎所有的上皮细胞和平滑肌细胞都起源于神经元。我们对 CDKN1C 和 DLK1 进行了特征描述，这两个基因在 T2DM 样本中显著上调，在非糖尿病和 T2DM 样本中识别 3 个代表性标记基因具有良好的可用性，并且它们与患者的临床特征也具有很强的关联性。Western blot 也表明，与对照组相比，在 30mM 葡萄糖培养 48 小时后，原代胰岛细胞中 CDKN1C 和 DLK1 的表达增加。此外，PubChem 分别为 CDKN1C 和 DLK1 预测了 11 种和 21 种潜在化合物。
出现的 2 个关键基因（CDKN1C 和 DLK1）可能是研究 T2DM 进展机制和开发创新疗法的催化剂。
