The Role Of Mir-148a In Pancreatic Cancer Cells

BackgroundPancreatic cancer is one of leading causes of cancer-related deaths owing to latediagnosis,fast tumor progression and low response rate to recent chemotherapeuticstrategies. Hence, further investigation of pancreatic cancer biology is warranted to gainplausible insights into the pathophysiologic mechanisms underlying Pancreaticpancreatic cancer development. Recently, a new class of RNA regulatory genes knownas microRNAs (microRNAs) has been found to introduce a whole new layer of generegulation in eukaryotes. MicroRNAs are endogenous non-coding RNAs, consisting of19to24nucleotides in length. They play an important role in regulating geneexpression by base-pairing to the complementary sites on the target mRNAs, thusblocking the translation or triggering the degradation of the target mRNAs.Mir-148a hasbeen earlier described to be down-regulated in several types of solid cancers.Thegenerally observed reduction in miR-148a abundance in several cancers lead to theassumption that miR-148a confers a tumor suppressive role and is important fortumorigenesis.Objective1. To investigate the functional role of deregulated miR-148a in pancreatic cancer.2.To find the downstream target gene of miR148a in pancreatic cancer cell linesPANC-1、BXPC-3.3. To Observed observe the effect between of the siRNA targeting ErbB3in the pancreatic cancer cell lines genes inhibition and to compare with that ofoverexpression and overexpression of miR-148a in the pancreatic cancer celllines.Methods1. MTT assaies were used to determine cellular PANC-1、BXPC-3carcinoma cellsproliferation.2. Migration assays were done using a modified transwell chamber system ofPANC-1、BXPC-3carcinoma cells3. Construction of luciferase reporter gene.The putative downstream target gene ofmiR148a was found through bio-informatics analysis. PANC-1、 BXPC-3panceratic cancer cells were transfected with the ErbB33’-UTR reporter plasmid.4. Then the activity of renilla and firefly luciferase was assessed using thedual-luciferaseH reporter assay system,Taqman PCR assay was used to assessmiR-148a expression.5. The expression of ErbB3was detected using western blot and RT-qpcr analysis.6. Cells were transfected with ErbB3RNAi by using Lipofectamine2000.Results1. MiR-148a inhibits proliferation of PANC-1、BXPC-3cells2. MiR-148a inhibits migration of PANC-1、BXPC-3cells.3. Three internet-based algorithms identified miR-148a as potentially binding with theErbB33‘-untranslated region.The result of luciferase activity assay revealed that ErbB3might be the direct target gene of miR-148a.4. Overexpressed ErbB3inversely correlated with miR-148a in pancreatic cancerPANC-1、BXPC-3cells.5. Transfection of the cell lines with miR-148a decreased the level of ErbB3protein andmRNA.6. Silencing of ErbB3with RNAi inhibited the growth of pancreatic cancer cells. 7. The inhibition of the growth of pancreatic cancer cells lines was miR-148a slightlybetter than direct interference with siRNA of ErbB3. in pancreatic cancer cell lines.Conclusions1. MiR-148a inhibits the PANC-1、BXPC-3cells proliferation and migration ofPANC-1、BXPC-3cells.2. ErbB3might be the direct target gene of miR-148a.3. Silencing of ErbB3with RNAi inhibited the growth of pancreatic cancer cells. Theinhibition of the growth of pancreatic cancer cells lines was slightly better thandirect interference with siRNA of ErbB3.But,the inhibition of miR-148a slightly better than direct interference with siRNA inpancreatic cancer cell lines.