Background: Esophageal squamous cell carcinoma (ESCC) is the fourth most common cause of cancer death in China. Long noncoding RNAs have emerged as critical regulators in cancer. Long intergenic noncoding RNA-p21, a kind of Long noncoding RNAs, LincRNA-p21  have been discussed dysregulated in several cancers, but its role in ESCC remains unknown. This study investigated the role of LincRNA-p21  in ESCC.
Materials and methods: The LincRNA-p21  expression level and its association with esophageal cancer was determined in 64 tumor tissues of esophageal squamous cell carcinoma patients and cells using quantitative real-time reverse transcription PCR. Fluorescence in situ hybridization of single-RNA molecular probes was used to determine subcellular localization of LincRNA-p21 . CCK8 and EdU assays were used for proliferation assay, flow cytometry was performed for apoptosis and cell-cycle distribution, and 24-well Mill cell chamber was made for measuring the abilities of migration and invasion after transfected with lentivirus-expressing LincRNA-p21  in EC109 cells. Then, quantitative real-time reverse transcription PCR and Western blot detected the expression of p21. Further, UC2288, an inhibitor of p21, was used to decrease the level of p21, and flow cytometry was used to detect cell cycle. Finally, screening for differential pathways from microarray analysis and expression of p53 and cyclin D were detected by Western blot.
Results: LincRNA-p21  expression level was remarkably lower in tumor tissues versus nontumor tissues and lower in EC109 cells versus Het-1A cells. Statistical analysis found that LincRNA-p21  might enhance the risk of ESCC. We observed that LincRNA-p21  was expressed both in the nucleus and cytoplasm, and a larger proportion of LincRNA-p21  was observed in the cytoplasm. The results demonstrated that upregulating the expression of LincRNA-p21  could inhibit cell proliferation, migration, invasion, and the transition of cell cycle from G1 and promoted apoptosis of EC109. Then, we found that LincRNA-p21  promotes the expression of p21. Decreasing the level of p21 revealed that cell-cycle arrest was restored. Pathway analysis found p53 pathway was downregulated, and upregulation of LincRNA-p21  inhibited the expression of cyclin D.
Conclusion: Our study suggests that LincRNA-p21  plays as a tumor inhibitor in ESCC development and LincRNA-p21  might induce G1 arrest through p53 signal pathway.
Keywords: esophageal squamous-cell carcinoma, LincRNA-p21 , G1 arrest, p53 pathway, Fluorescence in Situ Hybridization