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Bmi-1 silencing on neural stem cell proliferation and its mechanism of action

Author: JiHuiJun
Tutor: SunPeng
School: Qingdao University
Course: Neurosurgery
Keywords: Bmi-1 RNAi P16INK4a Neural Stem Cell Striatum
CLC: R741
Type: Master's thesis
Year: 2009
Downloads: 85
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Abstract


Objective Bmi-1 is a proto-oncogene belonging to the Polycomb group（PcG）.It is directly involved in the regulation of growth and proliferation of cells,and necessary for the self-renewal of stem cells.Transfect the neural stem cells with Bmi-1 and green fluorescent protein（GFP）co—expressing lentiviral vector.Observe the knock-down Bmi-1 expression and detect the effect on its possible downstream target gene expression of Bmi-1 and explore the relation of Bmi-1 and proliferation of cells.Methods Bmi-1 and green fluorescent protein（GFP） co—expressing lentiviral vector were transfected into neural stem cells which had been divided into the control group、non-carrier group and groups at 7D after transfection.Observe the changes of proliferation ability and cell sensence of neural stem cells before and after transfection。At same time,SYBR GreenⅠreal-time RT-PCR was used to quantitate Bmi-1,P16INK4a et al mRNA before and after transfection.Results①Cloning efficiency of the control group was 7.6%±0.2%,cloning efficiency of the bmi-1-shRNA group was 1.4%±0.3%（p＜0.05）;Brdu incorporation of the control group was 44.4%±9.3%,Brdu incorporation of bmi-1-shRNA group was 14.1%±6.7% （p＜0.05）.②Cell sensence of the neural stem cells after transfection was significantly promoted;③Compared with that of the control group,the inhibition rates of Bmi-1 mRNA at 7D detected by real-time PCR was 45%,and PI6INK4a mRNA expression was increased by 10 or 12 folds（p＜0.05）.Conclusions Bmi-1 shRNA can efficiently knock-down Bmi-1 mRNA expression in neural stem cells.Bmi-1 under-expression in neural stem cells has increased P16INK4a mRNA expression.Bmi-1 is the key factor of keeping the self-renewal of the neural stem cells.

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