做了上百个单细胞转录组项目，绝大部分都是肿瘤研究，在教程 CNS图表复现08—肿瘤单细胞数据第一次分群通用规则，这个第一次分群规则是 ：

• immune (CD45+,PTPRC),
• epithelial/cancer (EpCAM+,EPCAM),
• stromal (CD10+,MME,fibo or CD31+,PECAM1,endo)

的表达量来划分即可，这就是肿瘤免疫微环境的话题了！

肿瘤微环境这个热点应该不仅仅是集中在免疫细胞，其实还有基质细胞，其中热度最高的基质细胞应该是Cancer-associated fibroblasts (CAFs) ，但是目前呢，学界对CAFs的来源本来就是并不那么清晰，理论上不可能存的单一的标记基因来区分出来CAFs。通常CAFs有4种来源：

• The primary source is normal local fibroblasts, which are activated by stimuli from the tumor microenvironment.
• Mesenchymal stem cells (MSCs) and other mesenchymal precursor cells are other sources.
• Endothelial cells and epithelial cells do not belong to the fibroblast lineage, but they could transdifferentiate into CAFs-state cells.
• Finally, a self-renewable CAFs-stem cell population might exist in the hierarchical organization, and these cells share similar characteristics as MSCs.

如果要筛选CAFs，首先要去除4个基因表达量为阳性的细胞亚群 ：

• CD31 (an endothelial marker)
• CD45 (a hematopoietic cell marker),
• desmin (a smooth muscle cell marker),
• EPCAM (epithelial cell adhesion molecule, an epithelial cell marker).

然后各种文献整理一下，发现它的阳性标记基因可能是有：α-SMA, Collegen1A1, FAP, FSP1, PDGFRα and PDGFRβ, Podoplanin, and vimentin. 这么多，但是呢，都会有例外：

• α-SMA is unable to identify all CAFs in the TME, and it is expressed in smooth muscle in normal gastrointestinal and vascular
• FAP combined with CD45 is used to label a subgroup of cancer-associated macrophages
• FSP1 also corresponds to epithelial cells experiencing epithelial-to-mesenchymal transition (EMT;
• In a liver fibrosis model, FSP1 distinguishes inflammatory macrophage subpopulation

在人类疾病领域，可以区分成为：CAFs pro-tumorigenic function (pCAFs) or CAFs tumor-restain (rCAFs)

以上资料整理来源于新鲜出炉的综述：Front. Cell Dev. Biol., 04 February 2021 | https://doi.org/10.3389/fcell.2021.613534 标题是：《Cancer-Associated Fibroblasts Suppress Cancer Development: The Other Side of the Coin》

在小鼠模型里面，比如胰腺癌的小鼠模型里面，通常是3种不一样的，myofibroblast (myCAF), inflammatory fibroblast (iCAF), and antigen-presenting fibroblast (apCAF).  ：

• myCAF produces the majority of αSMA and collagen I transcripts
• S100a4 (Fsp1) expression in apCAFs
• iCAF 当然也是有自己的标记基因啦！

这3个分群，来源于文章：《Type I collagen deletion in αSMA+ myofibroblasts augments immune suppression and accelerates progression of pancreatic cancer》，于 March 2021发表于 cancer cell杂志，链接是：https://www.sciencedirect.com/science/article/pii/S1535610821001094

最后借用2018的一篇CELL文章《Structural Remodeling of the Human Colonic Mesenchyme in Inflammatory Bowel Disease》的 CyTOF panel 表格，信息如下：

可以看到，细胞发育谱系，细胞类型，细胞状态，其实还是水蛮深的！