Gram-negative bacteria and endotoxins promote hepatocellular carcinoma progression through the recruitment of myeloid-derived suppressor cells

Figure 5 Gram-negative bacteria and endotoxins mediate monocytic myeloid-derived suppressor cells aggregation through lipopolysaccharide/TLR4/CCL2. A: The mRNA expression levels of TLR4 and CCL2 in human HepG2, MHCC-97H and LX-2 cells were detected by phosphate-buffered saline (PBS), lipopolysaccharide (LPS), and lipoteichoic acid (LTA) after stimulation, respectively (n = 3 for each group); B: After the intervention with PBS, LPS, and LTA, the mRNA expression level of CCL2 was detected by reverse transcription-quantitative PCR (RT-qPCR); C: The aggregation of monocytic myeloid-derived suppressor cells (MMDSCs) was detected by flow cytometry (n = 4 for each group); D: Weekly in vivo imaging was performed to monitor tumor growth in control (Ctrl), LPS, and LTA groups; E: The changes in tumor size and liver weight in Ctrl, LPS, and LTA groups were assessed in animals that received different interventions; F: Volcano map of differential gene expression in Ctrl and LPS groups (n = 3); G: Kyoto Encyclopedia of Genes and Genomes enrichment pathway; H: Gene Set Enrichment Analyses enrichment pathway; I: Transcriptome sequencing showed the expression levels of interest genes. The mRNA expression levels of CCR2 and Ly6C in Ctrl and LPS group were detected by RT-qPCR (n = 4 for each group); J: The expression of CCL2 mRNA was detected by RT-qPCR in Ctrl and TAK-242 groups; K: The aggregation of MMDSCs was detected by flow cytometry in Ctrl and TAK-242 groups; L: Weekly in vivo imaging was performed to monitor tumor growth in Ctrl and TAK-242 groups; M: The changes in tumor size in Ctrl and TAK-242 groups were assessed in animals that received different interventions. ^aP < 0.05, ^bP < 0.01; ^cP < 0.001; ^dP < 0.0001; NS: Not significant. LPS: Lipopolysaccharide; LTA: Lipoteichoic acid; MMDSC: Monocytic myeloid-derived suppressor cell; GSEA: Gene Set Enrichment Analyses; Ctrl: Control.