Tumor-infiltrating CCR2+ inflammatory monocytes counteract specific immunotherapy

We used Single-cell RNA-sequencing to analyze the composition of the tumor microenvironment of MC38mOVA tumors upon transcutaneous immunization with our immunization method DIVA. Overall design: MC38mOVA tumor cells were inoculated s.c. into WT mice. Mice were treated by transcutaneous immunization at day 6/7 and 13/14 after inoculation or left untreated. At day 16, respectively day 20, single cell suspensions were prepared from excised whole tumors, and CD45+ cells were enriched by magnetic bead separation. Afterwards Single-cell RNA-sequencing of these CD45+ cells was performed.

本研究采用单细胞RNA测序（Single-cell RNA-sequencing），分析采用本团队开发的DIVA免疫方法实施经皮免疫（transcutaneous immunization）后，MC38mOVA肿瘤的肿瘤微环境（tumor microenvironment）组成。整体实验设计：将MC38mOVA肿瘤细胞经皮下（s.c.，subcutaneous）接种至野生型（Wild Type, WT）小鼠体内。于接种后第6/7天及第13/14天，对部分小鼠施以经皮免疫处理，其余小鼠不予任何处理作为空白对照。分别于接种后第16天及第20天，从切除的完整肿瘤组织中制备单细胞悬液，并通过磁珠分选（magnetic bead separation）富集CD45阳性（CD45+）细胞。随后对上述富集得到的CD45+细胞开展单细胞RNA测序。