GSE201026
Fluorescent tracking identifies tumor migratory dendritic cells as critical for cross-presentation in the lymph node following radiation therapy induced maturation
To understand the early effect of radiation therapy on tumor-infiltrating immune cells in two different tumor types, we established MC38 or Panc02-SIY tumors in C57BL/6 mice. scRNASeq was performed on CD45+ cells sorted by flow cytometry from single cell suspensions of tumors 1 day following 12Gy radiation therapy (RT), or control untreated (NT) tumors. Overall design: Tumors were implanted subcutaneously into the right flank as follows; 2 x 10^5 MC38, 5 x 10^6 Panc02-SIY. When tumors were approximately 5mm in average diameter, mice were randomized to receive treatment with CT-guided radiation using the Small Animal Radiation Research Platform from XStrahl. Dosimetry was performed using Murislice software from XStrahl. The SARRP delivered a single dose of 12Gy to an isocenter within the tumor using a 10mm x 10mm collimator and a 45 degree beam angle to minimize dose delivery to normal tissues. Tumors were harvested 24 hours post treatment (n=4 animals/group), processed into a single cell suspension as described above and magnetically enriched using CD45+ TIL MicroBeads (Miltenyi Biotec). Enriched cells were labeled with viability dye and CD45-APC. Live CD45+ cells were sorted using a 100uM nozzle on a BD Biosciences Aria cell sorter and cells were processed according to the manufacturers protocol for the Chromium Single Cell 3' Reagent kit (v3.0) from 10X Genomics. Libraries were sequenced using an Illumina NovaSeq 6000 using the NovaSeq 6000 S2 reagent kit (v1.0).