The Roles Of MiR-337 And Autophagy Related Gene LC3B,p62,p53 In Human Retinoblastoma

Dysfunction of autophagy has been implicated in development and progression of diverse human cancers.However,the exact role and mechanism of autopahgy has not been fully understood in human cancers,especially in retinoblastoma(Rb).Recent studies suggest that dysfunction of cell autophagic acitivity plays an important role in various human diseases,including neurodegenerative diseases,ageing,and cancer.However,the exact role of autophagy in tumor development and progression is not fully understood,even be controversial.Some studies demonstrated that autophagy plays an oncogene role in diverse cancers for its pro-survival capacity for cancer cells.This hypothesis is that autophagy can help cancer cells survive and proliferate under stressful conditions such as insufficient energy and resources.On the contrary,other studies revealed that autophagy plays a tumor suppressor role in cancer initiation.For example,allelic loss of the essential autophagy gene,Beclin-1,was significantly associated with high frequency of diverse human malignant tumors including ovarian,breast,and prostate cancer.Whereas LC3 and p62 promoted tumor progression.We determined the autophagy activity in human Rb tissues by assessing the autophagy markers microtubule-associated protein light chain 3B(LC3B)and SQSTM1(p62)in formalin fixed and paraffin embedded human tissue by immunohistochemistry and then associated their expression with patient clinicopathological features.We further explored the correlation between the expression of LC3 B and p62 and the expression of cytoplasmic p53,a newly identified autophagy suppressor,in Rb tissues.LC3 B is one of the most widely used markers to monitor autophagy.The other one of the best-studied autophagy regulator is p62,also named sequestosome1.P62 participates in autophagy-dependent elimination of many different cargos,such as ubiqutinated protein aggregates and bacteria.Due to interact with LC3,p62 is constantly degraded via autophagy and autophagy inhibition results in the accumulation of p62 positive aggregates.Therefore,monitoring of p62 degradation is used to measure autophagic flux under different conditions.Detection of LC3 B and p62 to monitor the level of autophagy can be performed by many techniques,such as western blotting and GFP-LC3 fluorescence microscopy,and so on.Interestingly,a recent study by Schl?fli et al demonstrated that immunohistochemistry staining of LC3 B and p62 on formalin fixed and paraffin embedded human tissues is specific and reliable to measure the level of cell autophagy.The principal tumor suppressor p53,a nuclear transcriptional regulatory protein,is involved in DNA damage,cell cycle,apoptosis and oncogene activation.Recently,an emerging area of research unravels the additional activities and functions of p53 in the cytoplasm,where p53 can induces cell apoptosis and inhibits autophagy.Furthermore,to investigate the regulatory machnism of p53 on cell autophagy,we used Targetsan prediction and experimental verification to determine the targeting miRNA of p53.Our data revealed that the expression of LC3 B and p62,was significantly associated with disease progression and tumor invasion of Rb.Furthermore,we also revealed that cytoplasmic expression of p53 was inversely associated with the behavior of tumor invasion.Finally,Spearman correlation analysis demonstrated that cytoplasmic expression of p53 was significantly and inversely correlated to expression of both LC3 B and p62.Autophagy might play an important role in human Rb progression and LC3 B and p62 may be useful predictors of disease progression in patients with Rb.Futhermore,we determined that in human Rb,p53 suppressed the expression of ATG5,ATG7,ATG12,Beclin1,LC3 and p62.P53 suppressed cell autophagic acitivity in Rb.Moreover,we documented that p53 was a direct target of miR-337 and miR-337 suppressed the expression of p53.In human Rb,miR-337 promoted cell autophagic acitivity.In conclusion,our data demonstrated that cell autophagy play an important role in human Rb development and progression.And expression of LC3 B and p62 was effect biomarker for disease progression and diagnosis of Rb.P53 could inhibit the autophgay by inhibit the expression of ATG5,ATG7,ATG12,Beclin1,LC3 and p62 in Rb cells.Finally,miR-337 could increase cell autophagy by targeting p53 in Rb cells.