Abstract
Objective To study the effect of increasing apoptosis by antisence miR - 221/222upregulating PUMA expression on U251 human glioma cell and its mechanism. Methods Oligofectamine was used to transfect miRNA- 221/222 antisense oligonucleotides to knock down the miR- 221/221expression level of U251 human glioma cell line in vitro. Flow cytometry was used to measure early apoptosis by Annexin V staining. Caspase 3/7 activity assay accessed Caspase 3 activity. Changes in mitochondrial membrane potential were determinated by JC - 1 Staining. Translocation of Bax was detected by immunofluorescence. Western blot assay validated expression of apoptotic protein. Results After miR -221/222 was knocked - down, the apoptosis was increased, Caspase 3 activity was enhanced, mitochondrial membrane potential was significantly decreased and Bax translocated from the cytosol to the mitochondria.Western blot assay showed that the expression of PUMA, Caspase 3 and Bax were up - regulated and the expression of bcl - 2 was down - regulated in the anti - miR - 221/222 group. Conclusions By upregulating PUMA, anti- miR-221/222 chould increase the apoptosis of U251 human glioma cell line. Key words: Glioma; miR - 221; miR - 222; PUMA; Apoptosis
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