血源性传播是结直肠癌（CRC）转移的一种普遍途径。然而，作为血管门卫的肿瘤周细胞（TPCs）在血源性转移中的作用尚不清楚。因此，我们旨在研究TPCs的异质性及其对CRC转移的影响。我们从CRC患者中分离出TPCs，并通过单细胞RNA测序（scRNA-seq）进行分析，其中包括有或没有肝转移的患者。采集了临床CRC标本，以分析TPCs的分子特征与CRC转移之间的关联。进行了RNA测序、染色质免疫共沉淀-测序和亚硫酸-测序，以研究TCF21调控的基因和机制，以及TCF21 DNA高甲基化下的整合蛋白α5的作用。构建了压脉细胞特异性Tcf21敲除小鼠，以研究TCF21在TPCs中对CRC转移的影响。采用Masson染色、原子力显微镜、二次谐波和双光子荧光显微镜来观察周血管外基质（ECM）重塑。通过scRNA-seq识别了13个TPC亚群。一种新的TCF21高TPCs子集，被称为“基质压脉细胞”，与CRC患者的肝转移相关。TPCs中的TCF21增加了周血管ECM的硬度、胶原重组和基底膜降解，建立了周血管转移微环境，促进了结直肠癌肝转移（CRCLM）。TPCs中的Tcf21消除减轻了周血管ECM重塑和CRCLM，而TCF21高TPCs和CRC细胞的联合注射显着促进了CRCLM。在机制上，α5整合蛋白的缺失抑制了FAK/PI3K/AKT/DNMT1轴，在TCF21高TPCs中破坏了TCF21 DNA高甲基化。本研究揭示了TPCs在血源性转移中以前未知的作用，并为CRC转移提供了潜在的诊断标志物和治疗靶点。© 作者（或其雇主）2022年的著作权。在CC BY-NC下允许再利用。不允许商业再利用。BMJ出版。

Haematogenous dissemination is a prevalent route of colorectal cancer (CRC) metastasis. However, as the gatekeeper of vessels, the role of tumour pericytes (TPCs) in haematogenous metastasis remains largely unknown. Here, we aimed to investigate the heterogeneity of TPCs and their effects on CRC metastasis.TPCs were isolated from patients with CRC with or without liver metastases and analysed by single-cell RNA sequencing (scRNA-seq). Clinical CRC specimens were collected to analyse the association between the molecular profiling of TPCs and CRC metastasis. RNA-sequencing, chromatin immunoprecipitation-sequencing and bisulfite-sequencing were performed to investigate the TCF21-regulated genes and mechanisms underlying integrin α5 on TCF21 DNA hypermethylation. Pericyte-conditional Tcf21-knockout mice were constructed to investigate the effects of TCF21 in TPCs on CRC metastasis. Masson staining, atomic force microscopy, second-harmonic generation and two-photon fluorescence microscopy were employed to observe perivascular extracellular matrix (ECM) remodelling.Thirteen TPC subpopulations were identified by scRNA-seq. A novel subset of TCF21high TPCs, termed 'matrix-pericytes', was associated with liver metastasis in patients with CRC. TCF21 in TPCs increased perivascular ECM stiffness, collagen rearrangement and basement membrane degradation, establishing a perivascular metastatic microenvironment to instigate colorectal cancer liver metastasis (CRCLM). Tcf21 depletion in TPCs mitigated perivascular ECM remodelling and CRCLM, whereas the coinjection of TCF21high TPCs and CRC cells markedly promoted CRCLM. Mechanistically, loss of integrin α5 inhibited the FAK/PI3K/AKT/DNMT1 axis to impair TCF21 DNA hypermethylation in TCF21high TPCs.This study uncovers a previously unidentified role of TPCs in haematogenous metastasis and provides a potential diagnostic marker and therapeutic target for CRC metastasis.© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.