Study On Role Of MicroRNAs In Hypoxic Microenvironment And Solid Tumor

Hypoxia remains a major characteristic of solid tumors and defined but usually referring to tissues at oxygen levels below 5 to 10 mmHg. Malignant cell growth requires the presence of a local vascular network that supplies oxygen and nutrients. However, a highly proliferating mass of tumor cells develops faster than the vasculature, and tumor cells rapidly meet up with an avascular environment deficient in oxygen. Hypoxia affects a variety of tumor cell properties such as cell growth, apoptosis, neovascularization, metastasis and sensitivity to treatment. Therefore, how hypoxic environments are generated in tumor tissues and how cells respond to hypoxia are essential questions in understanding tumor progression and a better understanding of these processes may lead to novel strategies for diagnosis and treatment. microRNAs (miRNAs) are about 22 nucleotide RNA molecules that participate in a wide variety of physiological and pathological cellular processes such as cell differentiation, proliferation, death, metabolism and more recently tumorigenesis. Recent studies have established a link between a distinct miRNAs expression profile with hypoxia, and global miRNAs expression changes have been described to occur in human cancers and in some cases shown to correlate with the clinicopathological features of the tumor. However, there is still few mechanism has been proposed to date for these profile alterations and relatively little is known about miRNAs regulation and response to hypoxia.In order to propose the mechanism of miRNAs response to hypoxia and involve in tumorigenesis or tumor progression, we screened the miRNA expression patterns in colorectal cancer (CRC) cell line cultured in diverse oxygen pressure using a high-resolution mapping assay, miRNA microarray and identified a specific group of hypoxia-related miRNAs with aberrant expression level. And then, the regulatory and acting mechanism of some important hypoxia-related miRNAs was investigated in vitro and in vivo. We found the expression levels of miR-17-92 cluster were reduced in hypoxia-treated cells containing wild-type p53, but were unchanged in hypoxia-treated p53-deficient cells. The repression of miR-17-92 cluster under hypoxia is independent of c-Myc, a promised transcription regulator of miR-17-92 cluster. Luciferase reporter assays mapped the region responding to p53-mediated repression to a p53-binding site in the proximal region of the miR-17-92 promoter. Chromatin immunoprecipitation (ChIP), Re-ChIP and gel retardation assays revealed that the binding sites for p53-and the TATA-binding protein (TBP)overlap within the promoter; these proteins were found to compete for binding. Over-express miR-17-92 cluster markedly inhibits hypoxia-induced apoptosis, whereas blocked miR-17-5p and miR-20a sensitize the cells to hypoxia-induced apoptosis. These data indicated that p53-mediated repression of miR-17-92 expression likely has an important function in hypoxia-induced apoptosis, and thus further our understanding of the tumor suppressive function of p53. Furthermore, we also demonstrate that the levels of miR-15-16 cluster are remarkably decreased under hypoxia and the low-expression favors colorectal cancer cell apoptosis, proliferative and especially invasive/metastatic behavior. Intratumoral administration of miRNAs shows that high levels of miR-15-16 cluster can significantly present angiogenesis and metastasis in a CRC nude mouse model, fibroblast growth factor-2 (FGF2), which regulates tissue angiogenesis, was identified as the direct and functional target of miR-15-16 both in vitro and in vivo. We also correlated the expression of miR-15-16 cluster with distant metastasis and clinical outcomes in malignant tissues. These data indicated that down-regulation of miR-15-16 cluster expression induced by hypoxia in CRC promotes tumor angiogenesis and distant metastasis by enhancement of FGF2 expression. In a word, our study provides a better understanding on the specificity of hypoxia and thus may be helpful in early diagnosis and targeting therapy of solid tumors...