Analysis of infiltrating leukocytes in the tumor microenvironment by single cell transcriptome
Hashimoto, S.1,*, Yurino, H.1, Kaneko S.1, Torigoe, T.2, Matsushima, K.3
1Graduate School of Medicine, Kanazawa University, Kanazawa; 2School of Medicine, Sapporo Medical University, Sapporo, Japan; 3Graduate School of Medicine, University of Tokyo, Japan
The phenotypic heterogeneity of cancer cells and their surrounding normal cells was observed in cancer tissue. In addition, cancer development is influenced by differential microenvironmental conditions such as hypoxia, immune infiltration and drug diffusion. Single-cell gene-expression profiling allows us to identify and characterize various cell types and subtypes in a cancer environment. Here, we have developed a novel strategy (Nx1-seq) of single-cell transcriptome analysis for thousands of single cells. In our approach, single cells are deposited in a high-density microwell plate and lysed in situ. mRNA is then captured on barcoding microbeads and reverse transcribed. The diversity of barcode on the beads is made by emulsion PCR using randomly synthesized barcode oligo DNA. The pooling of Nx1-seq data of single cells in two homogeneous cell populations can provide us rich and highly reproducible transcriptional profiles. Gene expression patterns among libraries resulting in a large number of sequenced reads were close (r=0.92). In this study, we applied Nx1-seq to characterize complex heterogeneous samples in the human endometrioid adenocarcinoma tissue. Endometrial cancer is the commonest pelvic gynaecological cancer in the world. These results showed the distinction of the cancer cell state with tumorigenicity, the EMT and infiltrated leukocytes. In addition, it was clarified that the phenotypic heterogeneity of the infiltrated immune cells from various cancer tissues such as hepatocellular carcinoma and sarcoma. Finally, Single cell transcriptome analysis is a powerful approach for characterizing and understanding cellular diversity in cancer tissues.
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