Glioma is a highly vascular, very aggressive and extremely invasive primary brain tumor. Hypoxia induces changes in glioma and its microenvironment, which leads to increased aggressiveness and resistance to chemotherapy and radiation . Studies have shown that large areas of hypoxia within glioma correlates inversely
with the patient’s outcome and survival [1–4]. ADAM17 (A Disintegrin and Metalloproteinase-17) DMXAA concentration also called TACE (TNF-alpha converting enzyme) plays a pivotal role in the processing of numerous growth factor proteins, and has emerged as a new therapeutic target in several tumor types [5–8]. Recent studies showed that when ADAM17 is either inhibited or suppressed there is attenuation in tumor invasiveness and malignancy, resulting in a better outcome for breast cancer patients [9, 10]. Low levels of oxygen (hypoxia) initiates cellular invasive processes that occur under physiological and pathological conditions such as tumor invasiveness and metastasis . Specificity transcription protein-1 (Sp1) is believed to play an important role in the transcription of many genes involved in cancer that have an abundance of GC boxes in their promoter region [12–15]. Currently, the role of Sp1 in ADAM17 expression and activity is unknown, but it is known the ADAM17 promoter region
contains GC-rich sequences highly complementary to the Sp1 DNA-binding site . Hypoxia induces expression Trichostatin A price of ADAM17 and increases invasiveness of glioma in vitro . In this study, we investigated if Sp1 protein plays a role in ADAM17 transcription, GABA Receptor and if Sp1 regulates hypoxic-induced ADAM17 expression in U87 human glioma cells. In addition, we examined the function of Sp1 in tumor invasiveness under normoxic and GSK1838705A order hypoxic conditions. Methods Cell culture The U87 tumor cell line was obtain from American Type Culture Collection (ATCC) The cells were grown in DMEM (Dulbeco Modified Essential Medium) which contained 10%
FBS (Fetal Bovine Serum),100 IU/mL penicillin, 100 μg/mL streptomycin (Life Technologies). The cells were passed once a week after trypsinization (0.05% trypsin-ethylenediaminetetraacetic acid; Life Technology). Hypoxic culture conditions The hypoxia experiments were performed in an anaerobic chamber (model 1025; Forma Scientific) which was saturated with 85%N2/10%H2/5%CO2. The temperature in the anaerobic chamber was set at 37°C and the oxygen level was below 1%. The media was changed before the experiment with DMEM low glucose and 10% FBS. The cells were harvested at 8, 12, 16 and 20 hours. In parallel with the hypoxic culture, normoxic culture was harvested as well to serve as a control for all assays.