Hypermethylation of the p16 gene promoter in pterygia and its association with the expression of DNA methyltransferase 3b.
OBJECTIVE
A pterygium has long been considered as a degenerative condition. After p53 protein was found to be abnormally expressed in the epithelium, researchers suggested that a pterygium may be a tumor, but additional evidence is required to support this hypothesis. Aberrant methylation of the p16 gene (CDKN2A) promoter and resultant gene silencing play important roles in the pathogenesis of many types of human cancers. The purpose of this study was to investigate hypermethylation of the p16 promoter in pterygia and the relationship between this hypermethylation and the expression of p16 and DNA methyltransferase 3b (DNMT3b) proteins.


METHODS
We studied the methylation status of p16 and the expression of p16 and DNMT3b proteins by performing methylation-specific polymerase chain reaction and immunohistochemistry, respectively, in specimens of 129 pterygia and 16 normal conjunctiva. The results were statistically analyzed.


RESULTS
Hypermethylation of the p16 gene promoter was detected in 21 (16.3%) of 129 pterygial specimens. Among them, 46 (35.7%) were positive for p16 protein expression, and 83 (64.3%) were negative. Staining for p16 was limited to the nuclei of the epithelial layer. We observed a significant reverse correlation between hypermethylation of the p16 promoter and the expression of p16 protein (p=0.006). Thirty-eight (29.5%) pterygial specimens were positive for DNMT3b protein expression, and 91 (70.5%) were negative. DNMT3b staining was limited to the nuclei of the epithelial layer. A significant correlation was found between hypermethylation of the p16 promoter and the expression of DNMT3b protein (p<0.001).


CONCLUSIONS
The p16 gene promoter was hypermethylated in pterygia, and this hypermethylation was strongly linked to expression of the positive expression of DNMT3b protein and to the suppression of p16 protein. These data provided molecular evidence that methylation occurs in pterygia and that it may play a role in the their development.
Chen
Pei-Liang
PL
Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan. t3273@so-net.net.tw <t3273@so-net.net.tw>
Cheng
Ya-Wen
YW
Chiang
Chun-Chi
CC
Tseng
Sung Huei
SH
Chau
Pak Sam
PS
Tsai
Yi-Yu
YY
D000368:Aged; D002467:Cell Nucleus / Q000378:metabolism; D003228:Conjunctiva / Q000378:metabolism; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000378:metabolism; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000378:metabolism*; D019175:DNA Methylation; D004848:Epithelium / Q000378:metabolism; D005260:Female; D019942:Genes, p16; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D007231:Infant, Newborn; D008297:Male; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011625:Pterygium / Q000201:enzymology* / Q000235:genetics* / Q000378:metabolism; D013194:Staining and Labeling; D014018:Tissue Distribution; D000097584:DNA Methyltransferase 3B
D019941:Cyclin-Dependent Kinase Inhibitor p16; D004248:DNA (Cytosine-5-)-Methyltransferases; D000097584:DNA Methyltransferase 3B