Expression Of Complement C1s In Esophageal Squamous Cell Carcinoma And Its Biological Role

Objective:To analyze the expression of complement C1 s in esophageal squamous carcinoma(ESCC)and its clinicopathological characteristics in patients with ESCC by means of bioinformatics.The influence of the change of complement C1 s expression on the proliferation and apoptosis of esophageal squamous cell cells was explored in vitro.The biological function of complement C1 s in ESCC was studied to lay theoretical and clinical foundation for the diagnosis and treatment of ESCC.Methods:1)The expression of complement C1 s in ESCC cancerous tissues,normal tissues or paracancerous tissues from GEO database was statistically analyzed.The expression of C1 s in cancerous and paracancerous tissues from Taizhou People’s Hospital was detected by immunohistochemical method,and the expression of C1 s in ESCC patients was verified by high throughput sequencing.Clinicopathologic data were collected from the TCGA database,and the correlation between clinical features and C1 s expression in patients with ESCC was statistically analyzed.2)C1s si RNA interference plasmid was screened.Human normal esophageal epithelial cell line ET-1A and esophageal squamous cell line TE-2 with low expression of complement C1 s were constructed.Three constructed C1 s si RNA interference sequences were transfected into HT-1A and TE-2 respectively.The cell transfection rate was further observed by fluorescence microscopy,and the interference efficiency of C1 s si RNA interference plasmid was detected by RT-q PCR.3)The effect of knocking down C1 s on the proliferation ability of ET-1A and TE-2 cells was detected by CCK-8 technique at the cellular level.And the effects of C1 s on Wnt1/ β-catenin pathway and apoptosis-related signal pathways were verified by Western-blot technique.Results:1)The expression of complement C1 s in ESCC was higher than that in normal or paracancerous tissues(P < 0.001)by the analysis of GSE77861 and GSE161533 data.The complement C1 s was highly expressed in tumor cells of patients with ESCC by immunohistochemistry,and its high expression in tumor cells of patients with ESCC was confirmed by high throughput sequencing.The clinical data of 82 patients with ESCC were analyzed by TCGA database.The high expression of complement C1 s was positively correlated with the race,residual focus and tumor location of ESCC patients,but not with other clinicopathological features.2)The fluorescence microscope observation showed that the positive rate of transfection was higher than 90% after si RNA transfection.The knockout si RNA sequence was selected as the condition for subsequent cell treatment combining with RT-q PCR results.3)CCK-8 experiment showed that the proliferation ability of HET-1A and TE-2 cells was down-regulated after knocking down C1 s,which was significantly weaker than that of normal esophageal cells HET-1A and ESCC TE-2.Flow cytometry showed that the apoptosis levels of HET-1A and TE-2 cells were up-regulated after knocking down C1 s,which was significantly higher than that of normal esophageal cells HET-1A and ESCC TE-2.After C1 s knockout,HET-1A cells inhibited cell proliferation by down-regulating the expression of Wnt1/ β-catenin pathway,and promoted cell apoptosis by down-regulating the expression of Bcl2 and up-regulating the expression of Cleaved-caspase3.TE-2 cells inhibited cell proliferation by down-regulating the expression of Wnt1/ β-catenin pathway,and promoted apoptosis by down-regulating the expression of Bcl2 and upregulating the expression of Bax and Cleaved-caspase3.Conclusion:The expression of complement C1 s in ESCC is significantly higher than that in paracancerous tissues,and the high expression of complement C1 s is positively correlated with the race,residual focus and tumor location of ESCC.Complement C1 s is involved in the process of proliferation and apoptosis of ESCC,and knocking down C1 s can significantly inhibit the proliferation of ESCC and promote the apoptosis of ESCC.C1 s can inhibit cell proliferation by regulating Wnt1/ β-catenin pathway,and promote cell apoptosis by regulating the expression of Bcl2,Bax and Cleaved-caspase3.