[Effect of siRNA targeting centromere protein-A gene on biological behavior of HepG2 cells].

Abstract

OBJECTIVE:

To study the influence of siRNA inhibition of CENP-A expression on the biological behavior of HepG2 cells.

METHODS:

Three pairs of 21 bp reverse repeated motifs of CENP-A target sequence with 9 spacer were synthesized and inserted into vector pSilencer 2.1-U6 neo to generate siRNA eukaryotic expression plasmids. After stable transfection into HepG2 cells, cell growth, apoptosis, cell cycles and plate clone forming efficiency were investigated. Expressions of CENP-A mRNA was monitored by the reverse transcriptase polymerase chain reaction (RT-PCR). The protein expression of CENP-A, bcl-2, Bax, p53, p21waf1 and mdm2 were detected by Western-blotting.

RESULTS:

Two eukaryotic expression plasmids with significant siRNA specific inhibition to the CENP-A gene were created. Compared with control cells, HepG2 cells transfected with the constructs showed G1 phase delay (P < 0.01) and cell number decrease in the S phase (P < 0.001), along with an increased apoptotic rate (P = 0.003), significant increase of Bax expression and decreased bcl-2 expression (P< or =0.001). The protein expressions of p21waf1 was higher and mdm2 was lower than those of the control groups. However, the wild type p53 protein expression was not effected by CENP-A siRNA.

CONCLUSIONS:

An altered expression of CENP-A may be related to the proliferation of hepatocellular carcinoma through cell cycle regulation involving an altered bcl-2/Bax expression, that may be p53 independent.

• Authors: Yong-mei Li, Zhi Zhu, Ying Chen, Zhi-gang Luo, Min Shi, Ming-hua Zhu
• Journal: Zhonghua bing li xue za zhi = Chinese journal of pathology (Zhonghua Bing Li Xue Za Zhi) Vol. 37 Issue 2 Pg. 124-8 (Feb 2008) ISSN: 0529-5807 [Print] China
• PMID: 18681325 (Publication Type: Journal Article)

Chemical References

• Autoantigens
• CENPA protein, human
• Centromere Protein A
• Chromosomal Proteins, Non-Histone
• RNA, Small Interfering

Topics

• Autoantigens (drug effects, genetics)
• Carcinoma, Hepatocellular (pathology)
• Cell Line, Tumor
• Centromere Protein A
• Chromosomal Proteins, Non-Histone (drug effects, genetics)
• Gene Expression Regulation, Neoplastic (drug effects)
• Hep G2 Cells
• Humans
• RNA Interference (drug effects)
• RNA, Small Interfering (drug effects, pharmacology)
• Reverse Transcriptase Polymerase Chain Reaction
• Tumor Cells, Cultured