Establishment of an N-2a cell line stably expressing mouse galanin and the effect of over-expressed galanin on the proliferation and apoptosis of N-2a cell.
- Author:
Man RONG
1
;
Rui-hu ZHANG
;
Tian-fu LIU
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Apoptosis; Cell Line, Tumor; Cell Proliferation; Galanin; biosynthesis; Mice; RNA, Messenger; biosynthesis
- From: Acta Academiae Medicinae Sinicae 2013;35(5):524-529
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo construct an N-2a cell line stably expressing PcDNA 3.1-platelet derived growth factor-galanin (GAL) and explore the effect of over-expressed GAL on proliferation and apoptosis of N-2a cell in vitro.
METHODSThe vector containing the target gene was transfected into N-2a cells by liposome, and cell clones stably over-expressing GAL was obtained via G418 screening. GAL mRNA and protein levels were determined by reverse transcriotion-polymerase chain reaction (RT-PCR) and Western blot. The proliferation of N-2a cells was detected by MTT.The cell cycle and apoptosis were detected by flow cytometry.
RESULTSRT-PCT and Western blot indicated that GAL genes were highly expressed in the transfected N-2a cells (i.e.GAL-N-2a). As shown by MTT, the proliferation of the N-2a cells transfected with PcDNA 3.1-PDGF-GAL was significantly slower than the control group(P<0.05). Compared with the non-transfected cells in the control group, the N-2a cells with endogenously overexpressed GAL were arrested at the G0/G1 phases, and the over-expressed GAL protein significantly induced the N-2a cell apoptosis in a concentration-dependent fashion.
CONCLUSIONEukaryotic expression vector PcDNA 3.1-PDGF-GAL can encode the expression of GAL in N-2a cells. Aslo, it can inhibit cell proliferation and promote the cell apoptosis.