| Background:The incidence of cutaneous squamous cell carcinoma（Cutaneous squamous cell carcinoma,c SCC）is secondary to basal cell carcinoma in non-melanomatous skin cancers,and keep increasing year by year.The prognosis of c SCC is usually poor if distant metastatic occur due to the highly invasiveness of c SCC cells.Surgical treatment is often the first choice for this cancer.For patients with advanced and metastatic c SCC,who lose the operative indication,or those in high risk of progression and need preventing measures,drug therapy may be a better method.Many studies have reported the possible effect and mechanism of retinoic acid in the treatment of cancer,but their mechanism for the treatment of c SCC remains unclear.Objective:The aim of this study is to explore the expression and regulatory mechanism of related genes via which acitretin affects c SCC cells from the level of cell transcriptomic analysis.This research provides a new theoretical basis for the clinical treatment of c SCC with retinoic acid.Methods:SCL-1 cells were divided into two groups in triplicate.Cells in the treatment group were incubated with RPMI-1640 medium containing acitretin at a concentration of 1.6×10-2mg/ml for 24h,while cells in the control group treated with RPMI-1640medium without acitretin.After treatment,cells were submitted to the following experiments:（1）Morphological study with a inverted microscope.（2）Apoptosis detection with a Annexin V-FITC kit under the confocal microscope.（3）Transcriptomic analysis including differentially expressed gene analysis,GO and KEGG enrichment analysis,after transcriptomic sequencing with RNA extraction from SCL-1 cells with a Trizol kit.（4）Differentially expressed gene partly verification with RT-q PCR.Results:（1）Compared with the control group,the SCL-1 cells in the acitretin treatment group had obvious changes in cell morphology,characterized by a significant decrease in cell density,an increase in cell volume,rough cytoplasm and increased accumulation of intracellular granules.（2）Acitretin treatment induced obvious early apoptosis and late apoptosis in the treatment group compared with the control group.（3）Transcriptomic data analysis revealed a total of 503 differentially expressed genes enriched in SCL-1 cells,including 205 up-regulated differential genes and 298down-regulated genes（|log2FC|≥1 and FDR<0.05）.The results of GO annotation enrichment analysis showed that differential genes were enriched in a variety of cell components and functions such as metabolism,signal,proliferation,differentiation,immunity,migration,etc.The KEGG pathway enrichment analysis indicated that the differential genes were mainly enriched in IL-17 signaling pathway,TNF signaling pathway,fluid shear stress and atherosclerosis,cytokine-cytokine receptor interaction,MAPK signaling pathway and so on.The KEGG enrichment analysis network map also suggested that MAPK signaling pathway and IL-17 signaling pathway were the core node pathways in the enriched pathways.In addition,compared with the control group,HBEGF,AKAP12,CYP26B1,DHRS9 and DHRS3 gene expressions were strongly up-regulated,and SOX2 gene expression was strongly down-regulated in the acitretin treatment group.（4）The expression trends of some genes,such as IL-17RE,DUPS1 and DUPS5 genes detected by RT-q PCR validation was completely consistent with the gene expression trends in RNA-Seq.Conclusion:（1）Acitretin can cause morphological changes of SCL-1 cells in vitro.（2）Acitretin can promote early and late apoptosis of SCL-1 cells in vitro.（3）Cell transcriptomic analysis suggests that acitretin may inhibit the growth of SCL-1cells in vitro by affecting various cellular components and functions,and regulating the genes expression and the transduction function of signaling pathways related to apoptosis and proliferation. |
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