Monocytic ontogeny of regenerated macrophages characterizes the mesotheliomagenic responses to carbon nanotubes

Macrophages not only derive from circulating blood monocytes, but also originate from embryonic progenitors that can proliferate. This dichotomic origin has an impact on macrophage functions in steady state and pathological conditions. Long needle-like carbon nanotubes (CNT) damage serous membranes and induce mesothelioma like asbestos fibers. Macrophages predominantly infiltrate CNT-induced injured tissue and contribute to cancer development. Here, we revealed the exact ontogeny of macrophage populations after CNT exposure by comparing the response to needle-like mesotheliomagenic CNT-7 with tangled-like non-mesotheliomagenic CNT-T. In a rat peritoneal cavity model, both CNT induced a rapid and complete depletion of MHCIIlow resident macrophages generating an empty niche available for macrophage repopulation. Macrophage disappearance after mesotheliomagenic CNT-7 was followed by a substantial inflammatory reaction, and macrophage replenishment completed after 7 days. Fifteen days after non-mesotheliomagenic CNT-T, macrophage repopulation was incomplete and accompanied by a limited inflammatory reaction. Cell depletion experiments, flow cytometry and RNA-seq analysis demonstrated that, after CNT-7, resident macrophages were mainly replaced by an influx of monocytes, which differentiated locally into MHCIIhigh inflammatory macrophages. In contrast, the low inflammatory response induced by CNT-T was linked to the activation of self-renewing MHCIIlow macrophages. In conclusion, the mesotheliomagenic response to CNT specifically relies on macrophage niche recolonization by monocytic progenitors. The homeostasis returns after exposure to non mesotheliomagenic CNT-T involves, in contrast, a macrophage maintenance by proliferation. Macrophage depletion and repopulation are thus decisive events characterizing the carcinogenic activity of fibrous particles.巨噬细胞不仅源自循环血液单核细胞，还可来源于具有增殖能力的胚胎祖细胞。这种二分起源对稳态及病理状态下的巨噬细胞功能具有重要影响。长针状碳纳米管（carbon nanotubes, CNT）可损伤浆膜，并如同石棉纤维一样诱发间皮瘤。巨噬细胞会大量浸润碳纳米管诱导的损伤组织，并促进癌症发生发展。 本研究通过对比针状致间皮瘤型CNT-7与缠结状非致间皮瘤型CNT-T诱导的应答，阐明了碳纳米管暴露后巨噬细胞群的确切起源发育过程。在大鼠腹膜腔模型中，两种碳纳米管均可快速、完全清除MHCII低表达驻留巨噬细胞（MHCIIlow），形成可供巨噬细胞重新定植的空白生态位。暴露于致间皮瘤型CNT-7后，巨噬细胞消失伴随显著的炎症反应，且巨噬细胞的补充在7天后完成。暴露于非致间皮瘤型CNT-T 15天后，巨噬细胞重新定植不完全，且伴随轻度炎症反应。 细胞清除实验、流式细胞术（flow cytometry）及RNA测序（RNA-seq）分析结果显示，暴露于CNT-7后，驻留巨噬细胞主要被涌入的单核细胞替代，后者在局部分化为MHCII高表达炎性巨噬细胞（MHCIIhigh）。与之相反，CNT-T诱导的轻度炎症反应与自我更新型MHCII低表达巨噬细胞的激活相关。 综上，碳纳米管诱导的致间皮瘤应答特异性依赖于单核细胞祖细胞对巨噬细胞生态位的重新定植。相比之下，暴露于非致间皮瘤型CNT-T后体内稳态的恢复则依赖于巨噬细胞的增殖以维持种群数量。因此，巨噬细胞的清除与重新定植是表征纤维颗粒致癌活性的决定性事件。