Micro Rnas Regulate The Mechanism Of C-phycocyanin-mediated Inhibition Of Non-small Cell Lung Cancer

C-phycocyanin(C-PC)is a blue,non-toxic,naturally active protein that is easily soluble in water and exists in algae such as cyanobacteria and spirulina.As a natural food colorant,C-PC also has the physiological function of antitumor activity.In China,the incidence and mortality of lung cancer ranks first among malignant tumors for a long time.Among them,non-small cell lung cancer(NSCLC)is the most common.mi RNAs(micro RNAs)are a class of endogenous,small,single-stranded non-coding RNAs consisting of about 19-23 nucleotides.They can pair with the 3 'UTR region of the target gene,So that expression levels are suppressed after transcription.The expression of mi RNAs in tumors is highly specific.Many studies have found that mi RNAs are related to the development of many cancers,including NSCLC.Therefore,this paper actively explores mi RNAs involved in the regulation of cellular physiological processes during the inhibition of NSCLC by PC.It is necessary to find the regulation mode of mi RNAs-target genes.This research can provide a new perspective for the study of the mechanism of PC against NSCLC.In this research,we used four typical non-small cell lung cancer cells A549,H1299,H460,and LTEP-a2 as the cell model.Previous studies have found that PC can inhibit the physiological activities of NSCLC,including the proliferation and migration of cells in vitro,and also promote apoptosis.In order to explore its inhibitory pathway,differential mi RNAstarget genes were screened by m RNA-mi RNA transcriptomics sequencing technology.First,two key differential proteins,RIPK1 and TIRAP,were screened upstream of the significantly different NF-?B signaling pathway,and the protein expression was specifically silenced by small interfering RNA technology.Cells were measured by MTT method,clone formation experiment,and flow cytometry and other methods were used to detect the in vitro proliferation ability of cells after silencing RIPK1 and TIRAP protein expression,respectively.The results show that silencing RIPK1 and silencing TIRAP can significantly inhibit the proliferation of non-small cell lung cancer in vitro.The results of flow cytometry double staining experiments also demonstrated that silencing RIPK1 and TIRAP can promote apoptosis.At the same time,Western Blot detected the expression of key proteins in the NF-?B signaling pathway and found that it can reduce the phosphorylation level of some proteins.Immediately afterwards,an overexpression recombinant plasmid was constructed to verify the biological functions of RIPK1 and TIRAP in both directions.Recombinant plasmids were transfected into cells,which significantly enhanced the cell's ability to proliferate and migrate in vitro.mi RNA-Seq searches for mi RNAs that may be directly targeted.Through bioinformatics prediction and double luciferase experiments,we found for the first time that RIPK1 is a direct target gene of mi R-642a-5p,and TIRAP is a target gene of mi R-3150a-3p,mi R-6883-3p,mi R-627-5p.The high expression of mi RNA in cells inhibits the protein expression of target genes.Further research found that the overexpression of mi R-642a-5p,mi R-3150a-3p,mi R-6883-3p,mi R-627-5p,respectively,resulted in the same results as the silencing-related target proteins RIPK1 and TIRAP.In addition,in vivo experiments were performed using a nude mouse subcutaneous tumor formation model.Nude mice administered with phycocyanin had a significant decrease in tumor formation ability.The expression of RIPK1,TIRAP protein,and mi RNA in the tumor body also basically matched the results of cell experiments.In summary,this study can provide a certain theoretical basis for the application of phycocyanin functional foods and the treatment of non-small cell lung cancer.