IN VITRO INVESTIGATION OF THE EFFECTIVENESS OF NATURAL KILLER CELLS ON GLIOBLASTOMA TUMOR CELLS DIFFERENTIATED FROM CORD BLOOD HEMATOPOIETIC STEM CELLS
Project no. 91 · Status: Kapanmış · 2017
Summary
Glioblastoma is the most common and the most malignant primary adult brain tumor. Median survival is 15 months and the 5-year survival rate is 4–5% despite the current treatment options such as surgery, radiotherapy, and chemotherapy. Cord blood is rich in source of CD34+ hematopoietic stem cell (HSC). Cord blood derived HSCs (CB-HSC) are superior to bone marrow derived HSCs in aspect of quantity and proliferation capacity. CB-HSC and umbilical cord mesenchymal stem cell (UC-MSC) due to capability of multipotency, they have immense potential to be used in different fields of medicine. Natural killer cells (NK) originate from bone marrow and known for its antitumoral and antiviral properties. NK cells use cytotoxic proteins, such as Perforin and Granzyme B, and apoptosis-inducing receptors as well as antibody dependent cytotoxicity to eliminate tumor cells. The aim of this study is to differentiate NK cells from CD34+ HSCs on UC-MSC feeder layer and co-culture these NK cells with T98, an immortalized glioblastoma cell-line, to test their antitumoral effects. UC-MSC are treated with mitomycin-c to inhibit its proliferation prior to use it as feeder-layer. CD34+ CB-HSCs are differentiated to NK cells on this feeder-layer derived from UC-MSC. Later NK cell characterization is performed by immunohistochemistry using indirect immunoperoxidase method. Differentiated NK cells from CD34+ HSCs on UC-MSC feeder layer are co-coltured with T98 glioblastoma cells to determine its antitumoral effect. Annexin V method is performed to determine induction of apoptosis. RT-PCR is performed to analyze expression of K-ras, P53, TGFBR2 cancer related genes as well as Oct 3/4 and Nanog genes related to cancer stem cells which are responsible for tumor development, invasion and metastasis. Based on the results obtained in this in vitro study, NK cells represent an efficient and effective cellular immunotherapeutic approach against glioblastoma