Abstract
PurposeThe prognosis of pancreatic cancer ranks among the worst of all cancer types, which is primarily due to the fact that during the past decades little progress has been made in its diagnosis and treatment. Here, we set out to investigate the role of microRNA 138 (miR-138-5p) in the regulation of pancreatic cancer cell growth and to assess its role as putative therapeutic target.MethodsqRT-PCR was used to examine the expression of miR-138-5p in 8 pancreatic cancer cell lines and 18 primary human pancreatic cancer samples. A lentivirual vector containing miR-138-5p mimics (lv-miR-138-5p) was used to exogenously over-express miR-138-5p in the pancreatic cancer cells lines Capan-2 and PANC-1. The effect of this over-expression on cell proliferation was examined using an in vitro propidium iodide fluorescence assay. Capan-2 cells exogenously over-expressing miR-138-5p were transplanted into nude mice to examine its in vivo effect on tumor growth. A predicted target of miR-138-5p (FOXC1) was first validated using a luciferase assay and, subsequently, down-regulated by siRNA to assess its effect on pancreatic cancer cell growth.ResultsWe found that miR-138-5p was markedly down-regulated in both pancreatic cancer cell lines and primary human pancreatic cancer samples, compared to a human pancreas ductal epithelial (HPDE) cell line and normal pancreatic tissues, respectively (P < 0.05). In addition, we found that in the pancreatic cancer cells lines Capan-2 and PANC-1 lentiviral transfection of miR-138-5p mimicked up-regulation of the endogenous expression of miR-138-5p and, concomitantly, inhibited cancer cell proliferation (P < 0.05). The exogenous over-expression of miR-138-5p also led to a significant inhibition of tumor formation in vivo. Using a luciferase assay, we found that miR-138-5p directly targets FOXC1. In conformity with this notion, we found that FOXC1 was down-regulated upon miR-138-5p over-expression in pancreatic cancer cells. Finally, we found that silencing of FOXC1 by siRNA had an inhibitory effect on pancreatic cancer cell growth.ConclusionsOur data indicate that miR-138-5p may play an important role in regulating pancreatic cancer cell growth, possibly through targeting FOXC1. Over-expression of miR-138-5p may serve as a novel approach for the treatment of patients with pancreatic cancer.
Highlights
Chao Yu and Min Wang contributed .C
Through bioinformatic analyses, including TargetScan, Pictar and miRANDA, we found that Forkhead box C1 (FOXC1) may serve as a direct target of miR-138-5p (Fig. 4a)
We found by Quantitative real-time reverse transcription-PCR (qRT-PCR) that FOXC1 is up-regulated in Capan-2 and PANC-1 cells (Fig. 4c), as well as in primary pancreatic cancer tissues (Fig. 4d), compared to normal pancreatic cells (HPDE) and nonmalignant clinical tissues, respectively
Summary
MiR-138-5p, one of the mature isoforms of the miR-138 family, has previously been shown to play a critical role in many types of cancer [33,34,35]. Unlike miR-138-5p, the other known mature isoform of the miR-138 family, miR-138-1-3p, does not appear to exert any relevant biological functions in humans (based on search results of Google Scholar and PubMed). We used qRT-PCR to assess the expression level of miR-138-5p in 9 pancreatic cancer cell lines, a human pancreas ductal epithelial cell line (HPDE) and a primary culture of normal pancreatic epithelial cells (Fig. 1a). We found that in all the pancreatic cancer cell lines tested the expression level of miR-138-5p was significantly lower than in the HPDE or primary normal pancreatic epithelial cells. The results obtained showed that miR-138-5p was significantly up-regulated in both transfected cell lines (Fig. 2a, P
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