The cancer stem cells characteristics analysis of LGR5 + cells that influence lung cancer risk by using single cell RNA-seq analysis.

Lung adenocarcinoma (LUAD) is the most popular lung cancer type with highly mortality. We performed a single cell RNA-seq analysis to explore characteristic of cancer stem cells in LUAD. We downloaded the single cell RNA-seq data (GSE149655) from the GEO database, the scRNA-seq analysis was performed by using the "Seurat" and "harmony" R package. The FindMarkers function and "ClusterProlifer" package was used for differentially expressed genes (DEGs) and function enrichment analysis. The protein-protein interaction and transcriptional regulatory network were performed by STRING and ChIPBase database. Immunohistochemistry tests to be used to observe differences in the expression of specific genes in LUAD and paracancerous tissue samples. BEAS-2B and A549 cells was used for vitro assay and the qRT-PCR, western blotting, wound healing, trans-well assays, EdU tests, and flow cytometry were performed. A total of 9 cell clusters were obtained after scRNA-seq analysis, in which the cancer stem cells had higher proportion in LUAD samples. Subsequently, function enrichment analysis revealed that the amino sugar and nucleotide sugar metabolism and DNA replication pathways were activated in cancer stem cells (CSCs), which were further sub-divided into 3 subtypes, the LGR5 + stem cell is a major contributor to cancer progression, its hub genes, such as HLA-DPB1, CD74, CTSH and HLA-DRB5 mediated the unique transcriptional state. In addition, the marker genes of three CSCs were also overexpressed in LUAD cells and the CXCL3 played an important role in mediating cell proliferation, apoptosis, migration and invasion of tumor. We performed a scRNA-seq analysis and identified the LGR5 + stem cell as a major contributor in LUAD progression, our findings are expected to provide new insights into the pathogenesis of LUAD.