Vascular endothelial growth factor (VEGF) promotes angiogenesis. VEGF has different isoforms with different functions that are generated by alternative splicing (AS).There are several proteins that participate in splicing, including serine-arginine-rich splicing factors (SRSF).SRSF are activated by kinases such as serine arginine rich protein kinases (SRPKs). Hypoxia has been shown to upregulate VEGF in several cancers. Others have demonstrated that hypoxia up-regulates SRSF proteins, which subsequently increases just the VEGF 121 isoforms in endometrial carcinoma cells. We previously showed that in hypoxic leukemia cells VEGF isoforms were spliced differently and VEGF 121 levels were inversely correlated with levels of WT1, a transcription factor. In acute lymphoblastic leukemia cells, VEGF165 and WT1 were significantly increased by hypoxia. In chronic myelogenous leukemia cells, hypoxia increased both VEGF165 and VEGF121 levels with little change in WT1 levels. Since others have shown that WT1 suppresses SRPK1 levels in sarcoma cells, our data in leukemia cells were consistent with an inverse relation between WT1 and VEGF 121 levels. The aim of this study was to determine if hypoxic conditions activated SRPK1/SRPK2 in leukemic cells to alter splicing of VEGF isoforms. The results indicated that hypoxia does not change mRNA levels of SRPK1/SRPK2, although protein levels need to be examined. Another possible explanation could be that other splicing kinases may be responsible for altering the VEGF 121 levels in leukemia cells. A better understanding of the mechanism of AS of VEGF could lead to new therapy targeting specific isoforms.
Hypoxic tumor microenvironment alters Vascular Endothelial Growth Factor isoform expression in cancer cells03/11/2015
Vascular endothelial growth factor (VEGF) is necessary for angiogenesis and tumor growth. VEGF splice isoforms have distinct characteristics, with VEGF121 being more diffusible, thus allowing for more distant angiogenic signaling, and VEGF165 and VEGF189 being found within the cell and the extracellular matrix. We hypothesized that VEGF expression in cancer cells would increase under hypoxia and that the VEGF 121 isoform would be enhanced. We examined VEGF mRNA levels and relative isoform ratios in cells grown in hypoxia or treated with CoCl2 to mimic hypoxia. Prostate cancer (PC3, LNCaP) and leukemia (MOLT-4, K562) cells were initially cultured under normoxic conditions with 20% O2 and then placed in a hypoxia chamber with 1% O2 or treated with CoCl2. RNA was extracted from treated and control cells, and quantitative real-time PCR analysis was performed to analyze VEGF expression under the different conditions. Our findings suggest that hypoxia increased both VEGF121 and VEGF165 expression in prostate cancer and leukemia cells. These results of elevated VEGF121 indicate the potential for lethal metastatic tumor growth distant from the primary tumor site. Overall, this work highlights the role of the hypoxic tumor microenvironment in regulating the functionally distinct VEGF isoforms in cancer cells.