| Objective: The aim of present study is (1) to examin thepossible role of Wnt signaling pathway in the pathogenesis of the rightcolon serrated polyps.(2) to explore the mechanism of activation andregulation of Wnt signaling pathway in the right colon serrated polyps.Methods:216cases of fresh polyps were collected during polypectomyor endoscopic biopsy from2011January to2012November in theendoscopy center of the Affiliated Hospital of Luzhou Medical College,10cases of normal colon mucosa collected as control. These tissues weredivided into7groups,including right-sided hyperplastic polyp(RHP),sessile serrated adenoma (SSA), right-sided traditional adenoma (RTA),right-sided colorectal cancer (RCRC), left-sided hyperplastic polyp (LHP),left-sided traditional adenoma (LTA) and left-sided colorectal cancer(LCRC). The right hemicolon includes the ascending colon, cecum andtransvers colon,and the left half colon includs the rectum, sigmoid colonand descending colon.(1) The expression of β-catenin, APC and c-myc invarious types of polyps were examined by immunohistochemistry (SPmethod) and immunofluorescence (indirect method).(2)After extractionof DNA from fresh tissues, the exon3of β-catenin and the exon15ofAPC were amplified using PCR method, followed by sequencing todetect the gene mutation.(3)After bisulfite modification of DNA in samples, methylated specific PCR using methylated and non methylatedprimers were performed. The methylation status of each specimen weredetermined by gel electrophoresis of PCR products.5HPs and5SSAsfrom the right half colon and right were selected to analysis by bisulfite-sequencing. After amplification of APC promoter, products were purifiedand connected to the T vector, which was then transformed into competentcells of escherichia TOP10coli. Plasmids were extracted for sequencing.The methylation status of each CpG site in APC promoter was analyzed.Results:(1) Expressions of APC, β-catenin and c-myc: The nuclearstaining rate of β-catenin was7.9%in the group of RHP,28.6%in SSA,significantly higher than that in LHP (3.3%, P<0.05). The cytoplasmicstaining rate of β-catenin was21.4%in RHPs,33.3%in SSAs,significantly higher than that in LHP (13.3%, P<0.05). The expression ofAPC in the RHPs (72.2±17.0), SSAs (68.4±13.1) and LHPs (53.7±26.5)were higher than that in TA (left,18.7±23.8; right,21.8±22.5) and CRC(left,31.4±21.5; right,34.4±19.7)(P<0.05). There was no significantdifferences of the expression of c-myc among the three groups of RHP(28.9±21.1),SSA (35.7±17.5) and LHP(39.0±16.3). The expression of c-myc in TA s(left,76.9±16.0; right,76.9±17.5) and CRCs (left,78.0±16.9;right,80.3±16.4) were significantly higher than that in RHPs(28.9±21.1),SSAs(35.7±17.5) and LHPs (39.0±16.3)（P<0.05).(2) Gene mutationsin β-catenin exon3and APC exon15: We found no gene mutation of β- catenin in HPs and SSAs(0/58and0/21,respectively). The mutation rateof β-catenin in TAs was2.9%（left）and6.3%（right）,in CRCs5.7%（left）and5.7%（right）.The mutation rates of APC in LCRCs, LTAs,RCRCs and RTAs were65.7%,57.1%,60.0%and78.1%,respectively. Inaddition, the mutation rates of APC in RHPs (67.9%) and SSAs (57.1%)were significantly higher than that in LHPs (36.7%, P<0.05).(3) APCpromoter methylation status: By the MSP method, we found thatAPC methylation in RHPs (14.3%) and SSAs(23.8%) were significantlylower than that in LHPs (36.7%) and TAs (left,37.1%; right,34.3%),(P<0.05). By BSP method, we confirmed the hypo-methylation statusof APC promoter in RHPs and SSAs. In addition, we found that APCmutations and APC methylation were negatively correlated (Spearman’sR=0.318, P<0.01). Moreover, APC expressions were negativelycorrelated with APC methylation (Spearman’s R=0.283, P<0.01).Conclusions:(1) Wnt signaling pathway in the right colon serrated lesionsis activated. Wnt signaling pathway may play an important role in the rightserrated pathway.(2).The activation of Wnt signaling pathway in the rightcolon serrated lesions may be related to the mutations of APC gene, insteadof APC promoter methylation. |
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