The Effect Of LncRNA UCA1 Cell Autophagy And Colon Cancer Cell Proliferation Apoptosis

Cell autophagy is a degrading way of cytoplasmic vacuolated lysosomess.It plays an important role in maintaining cell structure and functional stability,as well as tumor,cardiovascular disease,neurodegenerative diseases and so on,which by removing damaged organelles and misfolded proteins.Long non-coding RNA(lnc RNA)is a class of RNA between 200 bp to 100 kb in length.Once it recognised as “Transcriptional Noise” or “Dark Matter”.Recent studies have shown that lnc RNA has an effect on tumor development,cell proliferation and autophagy regulation.Urothelial carcinoma associated 1(UCA1)was originally found in human bladder cancer cell and involved in the growth and metastasis of a variety of tumors.However its research is less in colon cancer.The relationship between UCA1 and autophagy,and whether the regulation of cell autophagy affects the proliferation and apoptosis of colon cancer cells need to be explored.So this study analyzes the relationship between UCA1 and autophagy,discusses the effect of UCA1 on the proliferation and apoptosis of colon cancer cells.Thus,these provide the foundation for elucidating the role of UCA1 in autophagy with cells and in colon cancer.The main projects are as follows:1.The effects of UCA1 on autophagy and AKT/m TOR signalingThe 293 T cells were stimulated with LPS(1 ?g/m L)for 0 h,2 h,4 h,8 h,12 h and 24 h.At the same time,the expression of UCA1 and autophagy-related genes LC3,AKT and m TOR were detected by q PCR and the correlation was analyzed by SPSS software.Then 293 T cells were transfected with si-UCA1 and si-NC at a concentration of 50 n M for 24 h.q PCR and the Western Blot were used to detect the expression levels of LC3,ATG3,p62,AKT and m TOR.It was found that the expression levels of UCA1 and LC3 were significantly up-regulated,AKT and m TOR were significantly down-regulated at 4 hour and 6 hour.In addition,UCA1 was negatively correlated with AKT and m TOR,and was positively correlated with LC3.Down-regulated of UCA1,p62,AKT,m TOR was significantly up-regulated,while LC3 and ATG3 were significantly down-regulated.It shows that knockdown UCA1 can inhibit cell autophagy.2.The effects of UCA1 on miR-23 a expressionThe binding sites of UCA1 and miR-23 a were analyzed by bioinformatics,and miR-23 a mimics / mimics NC,miR-23 a inhibitor / inhibitor NC were transfected into 293 T cells.Then,q PCR was used to detect the expression of UCA1.At the same time,si-UCA1 and si-NC were transfected into 293 T cells,and the relative expression of Pri-miR-23 a,Pre-miR-23 a and Mature miR-23 a was detected by q PCR.The results showed that UCA1 and miR-23 a have a binding site and negatively correlated.And we found UCA1 down,and only the relative expression of Mature miR-23 a was significantly increased.That indicates that UCA1 is negatively correlated with miR-23's expression and is involved in its post-transcriptional regulation.3.The effects of UCA1 on proliferation and apoptosis of 293 T cellsThe si-UCA1 and si-NC were infected into 293 T cells and treated with LPS.Then,cell proliferation,apoptosis and cycling were measured by CCK-8 experiment and flow cytometry.Compared with the control group,we found that down-regulated of UCA1 resulted in obvious decreased of the cell proliferation level at 4 d-5 d,significantly increased of apoptosis levels,and the cell cycle block in G2 phase.The result indicate that down-regulated of UCA1 can inhibit cell proliferation,promote the cell cycle block in G2 phase and cell apoptosis.4.The expression of UCA1 in colon cancer cells and its effect on autophagyThe q PCR was used to detect the expression of UCA1 in normal colon epithelial cells NCM460 and colon cancer cell lines,such as Lo Vo,HT-29,HCT-116,Caco-2.Then,we transfected in Caco-2 with the same concentration of si-NC,si-UCA1 and si-UCA1 with RAPA(autophagy activator,100 nmol/L).Then we used Western Blot to detect LC3 protein expression after 24 h.The results showed that the expression level of UCA1 in colon cancer cells was significantly higher than that in the control group,especially Caco-2 cells.And after the interference of UCA1,LC3 II protein significantly down,RAPA is significantly reversed.Which suggests that down-regulated of UCA1 can inhibit the occurrence of autophagy in colon cancer cells.5.The effects of UCA1 on Caco-2 proliferation and apoptosis by regulating cell autophagyThe si-UCA1,si-UCA1 with RAPA and si-UCA1 with 3-MA(autophagy inhibitor,5 mmol/L)were transfected into Caco-2.Then,the cell proliferation,apoptosis and cycling were measured by CCK-8 experiment and flow cytometry.The results showed that the cell proliferation level decreased significantly,the apoptosis level was significantly increased and the cell cycle occurred G2 phase arrest after UCA1 interfered.But 3-MA can promote this effect,and RAPA plays a role in inhibiting.This indicated that down-regulated of UCA1 in Caco-2 cells can inhibit cell autophagy.Thereby it can inhibit cell proliferation,promote cell apoptosis and block cell cycle G2 phase.In summary: UCA1 is closely related to cell autophagy and may affect cell autophagy by AKT / m TOR signaling pathway;UCA1 can also regulate the expression of miR-23 a at post-transcriptional level;Knockdown of UCA1 can change cell activity and it can inhibit cell proliferation promoting cell apoptosis and block cell cycle G2 phase by inhibiting cell autophagy.