GSE196054
Dnase1l3 enhances antitumor immunity and suppresses tumor progression [scRNAseq]
For the chronic inflammation-related colon cancer model (AOM/DSS-colon cancer model), 3-month-old Dnase1l3 WT and KO mice were injected with AOM (8 mg/kg, body weight). One week later, mice were challenged with 2.5% DSS water for 7 days followed by a 14-day recovery with regular drinking water for three cycles. Body weight, rectal bleeding and diarrhea were monitored during the entire experiment. Mice with more than 25% weight loss were removed during the experiment. For the AOM model, 2-month-old Dnase1l3 WT and KO mice were injected intraperitoneally with AOM (8 mg/kg, body weight). Colon tissues were isolated 9 hours, or five days after injection. All animal experiments were conducted in accordance with guidelines of NIEHS/NIH Animal Care and Use Committee. Overall design: Tumor associated DCs and total leukocytes were isolated and enriched. Cell surface antigens were stained with following fluorochrome-conjugated Abs together with eFluor780-conjugated Live/Dead dye (ThermoFisher Scientific). PerCP-Cy5.5-anti-mouse CD11c (N418, eBio 45-0114-82), APC-anti-mouse CD26 (H194-112, BL 137807), BV711-anti-mouse CD45.2 (104, BL 137807), PE-anti-mouse CD88 (20/70, BL 135806), eFluor450-anti-mouse MHC class-II I-Ab (AF6-120.1, eBio 48-5320-82), BV510-anti-mouse Ly-6G (1A8, BL 127633). Among stained cells, cDCs (CD11c+ CD26+ CD45.2+ CD88– I-A+ Ly-6G– Live/Dead–) or leukocytes (CD45.2+ Live/Dead–) were purified using a cell sorter FACS ARIA-II (BD Biosciences). The cells were counted and examined for viability using a TC-20 cell counter (Bio-Rad). About 8000-9900 live cells at 2×10^5 cells/mL concentration with 70% or higher viability were loaded into the Single Cell Chip followed by forming single cell emulsion in Chromium Single Cell Controller (10x Genomics, Chromium Single Cell 3' Library & Gel Bead Kit v3.1). The mRNA reverse transcription, cDNA generation and amplification, and single cell gene expression library construction were carried out according to the protocols provided by the manufacture. The libraries were then sequenced by the NIEHS Epigenomics and DNA Sequencing Core Laboratory on NovaSeq 6000 (Illumina) with paired-end sequencing (Read 1: 28; Read 2: 90). A total of 8.7×10^8 reads were obtained for the four samples