Effects Of LRP16 On Proliferation And Apoptosis Of Esophageal Squamous Cell Carcinoma And Its Relationship With Hippo Pathway

Background and purpose:Esophageal cancer(EC)is the seventh most common malignant tumor in the world,of which about 50%of esophageal cancer occurs in China,especially in parts of Henan.The most common EC is esophageal squamous cell carcinoma(ESCC),and the cure rate of ESCC is very low.So far,surgery and resection are the main treatment methods of ESCC,but the five-year survival rate after surgery is only 15-25%.What is more serious is that with the improvement of living standards and the extension of human life expectancy,the number of elderly patients with esophageal cancer has increased year by year,and the incidence of ESCC generally has an upward trend.Therefore,there is an urgent need to explore and discover new targeting molecules and molecular mechanisms related to the development and progress of ESCC,in order to improve the diagnosis and treatment of ESCC.The latest research results show that Leukemia-associated protein 16(LRP16)is a nuclear protein factor associated with tumorigenesis,and also confirms that LRP16 is a co-activator of ERα.At the same time,studies have also shown that ERα can activate the Hippo pathway in ESCC,causing the cells to become disordered,the cells will overproliferate,the apoptosis will be weakened,and the proliferation of tissues and organs will increase,leading to tumors.However,the expression of LRP16 in ESCC has not been reported,and its functional role in ESCC is temporarily unclear.Based on the above theoretical basis,this experiment proposed that LRP16 protein is involved in the occurrence and development of esophageal cancer and is related to the Hippo pathway.Methods:1.The immunohistochemical sections of the Department of Pathology of the first affiliated Hospital of Zhengzhou University were collected,and the expression of LRP16 gene in ESCC,atypical hyperplasia and normal tissues was detected by immunohistochemistry.The relationship between LRP16 gene and TNM stage,distant metastasis and depth of invasion was analyzed by patient information.2.A cell line with the highest relative expression of LRP16 among ESCC cell lines was selected for LRP16 gene silencing experiment.The plasmid was transfected into esophageal cancer cell line,and the stable transgenic strain was screened by genetic mycin(G418).Verified by qRT-PCR and Western blot techniques,the LRP16 knock-down stable transgenic strain was obtained.3.The effect of knocking down LRP16 on apoptosis was detected by flow cytometry,the proliferation of ECSS cells was detected by CCK-8 value-added technique,and the repair ability of ESCC cells was detected by scratch test.4.qRT-PCR and Western blot experiments were used to analyze the changes of the expression of factors related to Hippo signal pathway before and after knocking down LRP16.Results:1.This test proves that the expression of LRP16 in ESCC tissues is much higher than that in normal tissues,and it has obvious differential expression.The expression of atypical hyperplasia tissues is higher than that of normal tissues.There is a significant difference in expression between the three.The expression level of LRP16 has nothing to do with gender and age,and is related to the depth of invasion,TNM stage and lymph node metastasis,which means that LRP16 is involved in the metastasis and spread of esophageal squamous cell carcinoma.2.Among the 5 ESCC cell lines,TE-1 cells have the highest relative expression of LRP16 protein,which can be used as an experimental object.and the results of fluorescence expression showed that the sh-LRP16 plasmid was successfully transferred into TE-1ESCC cell line.The results of qRT-PCR and Western blot experiments showed that the TE-1 ESCC line with low LRP16 was successfully constructed.3.In the ESCC with low LRP16 expression,the apoptosis and proliferation decreased,the cell cycle was blocked in G2/M phase,and the mobility and invasion ability decreased at the same time.4.After knocking down LRP16,the expression of ERα and ERβ decreased,the expression of YAP1 decreased,and the expression of MST increased,confirming that LRP16 promotes the development of ESCC through Hippo signaling pathway.Conclusion:Our experiments confirmed that LRP16 is expressed in ESCC and is related to the depth of tumor invasion,TNM stage,and lymph node metastasis.This study initially explained that silencing LRP16 can inhibit the proliferation and apoptosis of ESCC cells,block the cell cycle at G2/M phase,reduce the ability of cell migration,and LRP16 participates in the regulation of Hippo signaling pathway.It can regulate and inhibit the progression of ESCC by inhibiting YAP1 and up-regulating MST1/2.This provides a new idea for the diagnosis,drug treatment and early prevention of ESCC in the future.