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. 2007 Jul 3:7:120.
doi: 10.1186/1471-2407-7-120.

Association of diminished expression of RASSF1A with promoter methylation in primary gastric cancer from patients of central China

Mei Ye  1 , Bing Xia, Qiusha Guo, Feng Zhou, Xiaolian Zhang
Affiliations

Association of diminished expression of RASSF1A with promoter methylation in primary gastric cancer from patients of central China

Mei Ye et al. BMC Cancer. .

Abstract

Background: Although methylation-mediated inactivation of expression of RASSF1A, a candidate tumor suppressor gene, has been observed in several human cancers, the data concerning alteration of RASSF1A expression and methylation in Chinese primary gastric cancer are scarce. Moreover, direct evidence showing the association between protein expression of RASSF1A and primary human cancers is lacking. The aim of this study was to investigate RASSF1A expression in tissue of primary gastric cancer (GC) at mRNA and protein levels, and to establish the possible relationship between DNA methylation status and protein expression of RASSF1A in Chinese.

Methods: Fifty-four patients with primary gastric cancers were included in the study of RASSF1A mRNA expression and methylation status between the cancer tissue and the corresponding adjacent normal tissue. 20 out of 54 patients were included for study of RASSF1A protein expression. The expression of RASSF1A at mRNA and protein levels was determined by RT-PCR and Western-blotting, respectively. The RASSF1A promoter methylation was detected by methylation-specific PCR.

Results: RASSF1A mRNA and protein expressions in GC were reduced significantly with comparison to the corresponding normal tissues (OD value: 0.2589 +/- 0.2407 vs 0.5448 +/- 0.2971, P < 0.0001; 0.1874 +/- 0.0737 vs 0.6654 +/- 0.2201, P < 0.0001, respectively). Methylation frequency of RASSF1A in primary GC is higher than that in the corresponding normal tissues (66.7% vs. 14.8%, P < 0.0001). Furthermore, RASSF1A mRNA expression in methylation group of GC was further reduced when compared to the unmethylation group of GC (0.1384 +/- 0.1142 vs. 0.5018 +/- 0.2463, P < 0.0001).

Conclusion: Expression of RASSF1A was reduced in tissue of GC at mRNA and protein levels. Diminished expression of RASSF1A was associated with the promoter methylation.

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Figures

Figure 1
Figure 1
Expression of RAASF1A mRNA in primary gastric cancer (T) and corresponding normal tissue (N). RT-PCR products for RASSF1A were quantified by densitometric scanning of in ethidium bromide-stained gels compared with GAPDH.
Figure 2
Figure 2
Protein expression of RASSF1A in primary gastric cancer (T) and corresponding normal tissue (N). Protein level of RASSF1A was quantified by Western-blotting compared with GAPDH.
Figure 3
Figure 3
Methylation of RASSFIA in primary gastric cancer and corresponding normal tissue by MSP. Lane 3, 4 and lane 6, 7 show a representative result from a tumor and normal sample, respectively. U: amplified product with primer recognizing unmethylated sequences; M: amplified product with primer recognizing methylated sequences. Genomic DNA, methylated in vitro by CpG methylase (Sss I) was used as a positive control. Water blank was used as a negative control. The PCR products were resolved on a 8% polyacrylamide gels
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