Ni Chen, Hua Rulin, Yang Yuanyuan, Liang Jialu, Liu Wentao, Wang Linlin, Yao Xiaohan, Li Anqi, Yu Long, Feng Ruo, Lv Dekang, Qin Zhihai, Zhai Wenlong
Department of Hepatobiliary and Pancreatic Surgery, Medical Research Center, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450052, China.
Henan China-Germany International Joint Laboratory of Tumor Immune Microenvironment and Disease, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, 450052, China.
BMC Gastroenterol. 2025 Apr 9;25(1):235. doi: 10.1186/s12876-025-03829-8.
Extrahepatic cholangiocarcinoma (eCCA) is a rare but refractory cancer with dense desmoplasia. Prognosis-associated stromal cells in eCCA remain poorly characterized. Here, we profiled the tumor cellular composition and identified prognosis-related stromal signatures by single-cell RNA sequencing (scRNA-seq) in eCCA. ECCA patients were further stratified into different categories based on identified stromal signatures.
Using scRNA-seq, we profiled the transcriptomes of 37,498 individual cells from eight eCCA biopsies, including five tumor tissues and three paired adjacent normal tissues. Bulk RNA sequencing (bRNA-seq) was also performed on 43 eCCA tumor tissues. Stromal cell composition and heterogeneity were examined through differential gene expression and gene set enrichment analyses. By assessing the expression levels of marker genes in bRNA-seq data, the correlation of stromal cell clusters with survival was explored. The GSVA scores of the cell-specific signature genes of the prognosis-related stromal cell subtypes were calculated and used to stratify eCCA patients.
The results revealed that tumor stroma in eCCA were composed of hematopoietic progenitor-like cells (HPLCs), fibroblasts (Fb), Schwann cells (Sch), endothelial cells and immune cells. Prognosis-associated stromal cell subpopulations included MKI67 + HPLC, TMEM158 + C3-Fb, FOXP3 + regulatory T cells (Treg), SLIT2 + Sch, TPSD1 + C2-mast cells (MC) and CTSG + C3-MC. Based on these stromal signatures, the eCCA tumors were categorized into three classes: proliferative Group 1 with enrichment of MKI67 + HPLC, inflammatory and fibrotic Group 2 with enrichment of TPSD1 + C2- MC, FOXP3 + Treg and TMEM158 + C3-Fb, and neuronal Group 3 with enrichment of SLIT2 + Sch and CTSG + C3-MC. ECCA patients in Group 3 had a better prognosis when compared to Group 1 and 2, reflecting different impact of stromal subtypes on tumor progression.
Single-cell transcriptomic analysis reveals prognosis-related stromal signatures that potentiate the stratification of eCCA into proliferative, inflammatory and fibrotic, and neuronal phenotypes, which has important implications on molecular classification and exploring therapeutic targets in eCCA.
肝外胆管癌（eCCA）是一种罕见但难治的癌症，伴有致密的促结缔组织增生。eCCA中与预后相关的基质细胞仍未得到充分表征。在此，我们通过单细胞RNA测序（scRNA-seq）分析了eCCA的肿瘤细胞组成，并确定了与预后相关的基质特征。基于所确定的基质特征，将eCCA患者进一步分为不同类别。
我们使用scRNA-seq分析了来自8例eCCA活检组织的37498个单个细胞的转录组，包括5个肿瘤组织和3个配对的相邻正常组织。还对43例eCCA肿瘤组织进行了批量RNA测序（bRNA-seq）。通过差异基因表达和基因集富集分析来检查基质细胞组成和异质性。通过评估bRNA-seq数据中标记基因的表达水平，探讨基质细胞簇与生存的相关性。计算预后相关基质细胞亚型的细胞特异性特征基因的GSVA评分，并用于对eCCA患者进行分层。
结果显示，eCCA中的肿瘤基质由造血祖细胞样细胞（HPLCs）、成纤维细胞（Fb）、雪旺细胞（Sch）、内皮细胞和免疫细胞组成。与预后相关的基质细胞亚群包括MKI67 + HPLC、TMEM158 + C3-Fb、FOXP3 + 调节性T细胞（Treg）、SLIT2 + Sch、TPSD1 + C2-肥大细胞（MC）和CTSG + C3-MC。基于这些基质特征，将eCCA肿瘤分为三类：富含MKI67 + HPLC的增殖性第1组、富含TPSD1 + C2-MC、FOXP3 + Treg和TMEM158 + C3-Fb的炎症和纤维化第2组，以及富含SLIT2 + Sch和CTSG + C3-MC的神经元第3组。与第1组和第2组相比，第3组的eCCA患者预后更好，这反映了基质亚型对肿瘤进展的不同影响。
单细胞转录组分析揭示了与预后相关的基质特征，可以将eCCA分为增殖性、炎症和纤维化以及神经元表型，这对eCCA的分子分类和探索治疗靶点具有重要意义。
