Regenerating gene 4 promotes chemoresistance of colorectal cancer by affecting lipid droplet synthesis and assembly

Figure 1 Effects of full-length regenerating gene 4 and nonsignal peptide regenerating gene 4 on phenotypes of DLD-1 cells. A: DLD-1 cells were incubated with increasing doses of recombinant human regenerating gene 4 (REG4) (rhREG4) (0-500 nmol/L) for 48 h and subjected to cell proliferation assay by tetrazolium salt (MTT) assay; B: Treatment with anti-REG4 antibody produced a dose-dependent decrease in cell number of full-length (FL)-REG4-overexpressing DLD-1 cells; C: MTT assay was used to detect the proliferation of DLD-1 cells transfected with FL-REG4 or nonsignal peptide (NSP)-REG4, and treated with rhREG4 or REG4 antibody; D: Apoptosis of DLD-1 cells transfected with FL-REG4 or NSP-REG4, and treated with rhREG4 or REG4 antibody detected by flow cytometry; E: Wound healing assay was used to detect migration of DLD-1 cells transfected with FL-REG4 or NSP-REG4, and treated with rhREG4 or REG4 antibody; F: Transwell assay was used to detect migration and invasion of DLD-1 cells transfected with FL-REG4 or NSP-REG4, and treated with rhREG4 or REG4 antibody; G: Compared with DLD-1 cells, western blotting showed that transfection with FL-REG4 and treatment with rhREG4 increased expression of epidermal growth factor receptor (EGFR)-Tyr992, Tyr1068, Tyr1148, Tyr1173, Akt, p-Akt, phosphorylated phosphoinositide 3-kinase (p-PI3K), nuclear factor (NF)-κB, p-NF-KB, Bcl-2 and Bcl-X/L. REG4 antibody inhibited the effect of FL-REG4 transfection; H: Western blotting showed that compared with parental cells, DLD-1 cells transfected with NSP-REG4 had no change in expression of EGFR-Tyr992, Tyr1068, Tyr1148, Tyr1173, AKT, p-AKT, p-PI3K, NF-KB, p-NF-KB, BCL-2, or BCL-XL. ^aP < 0.001; ^bP < 0.01; ^dNo significance. Ab: Anti-REG4 antibody; REG4: Regenerating gene 4; rhREG4: Recombinant human regenerating gene 4; FL: Full-length; NSP: Nonsignal peptide; EGFR: Epidermal growth factor receptor; p-PI3K: Phosphorylated phosphoinositide 3-kinase; NF: Nuclear factor.