Analysis of the expression profile of skin macrophages FACS-sorted from mice overexpressing activin and/or oncogenes of human papilloma virus 8 in keratinocytes.

We have shown that activin promoted skin tumorigenesis in mice induced by the human papilloma virus 8 oncogenes. Activin attracted blood monocytes to the skin as revealed by depletion of CCR2-positive monocytes. To determine if activin also altered the gene expression profile of these cells, we performed RNA-Sequencing of macrophages FACS-sorted from the pre-cancerous ear skin. We have found that activin induces a pro-migratiory, pro-angiogenic and pro-tumorigenic genes in skin macrophages in vivo. This largely contributes to the pro-tumorigenic function of activin, since macrophage depletion delayed spontaneous tumorigenesis in HPV8-transgenic mice by reducing keratinocyte proliferation and angiogenesis. Overall design: F4/80+CD11b+CD45+ cells were FACS-sorted from the pre-cancerous ear skin of wt/wt, HPV8/wt, wt/Act and HPV8/Act mice and their expression profile was analysed by RNA-Sequencing. Experiment was performed in triplicates, for each replicate ear skin of 3-6 mice of corresponding genotype was pooled.

本研究证实，激活素（activin）可促进人乳头瘤病毒8型（human papilloma virus 8, HPV8）致癌基因诱导的小鼠皮肤肿瘤发生。通过CC趋化因子受体2（CCR2）阳性单核细胞耗竭实验证实，激活素可招募血液单核细胞向皮肤浸润。为明确激活素是否可改变此类细胞的基因表达谱，我们对从癌前病变耳皮肤中经荧光激活细胞分选（FACS）得到的巨噬细胞进行了RNA测序（RNA-Sequencing）。本研究发现，激活素可在体内诱导皮肤巨噬细胞表达促迁移、促血管生成及促肿瘤发生相关基因。该效应是激活素发挥促肿瘤发生功能的核心机制之一：巨噬细胞耗竭可通过降低角质形成细胞增殖与血管生成，延缓HPV8转基因小鼠的自发性皮肤肿瘤发生。实验设计：从野生型纯合（wt/wt）、HPV8杂合转基因（HPV8/wt）、激活素过表达野生型（wt/Act）及HPV8+激活素过表达（HPV8/Act）小鼠的癌前病变耳皮肤中，经FACS分选得到F4/80+CD11b+CD45+细胞，通过RNA测序分析其基因表达谱。本实验设置三次生物学重复，每个重复混合3~6只同基因型小鼠的耳皮肤组织。