Analysis of p16 Gene Mutation, Deletion and Methylation in Patients with Arseniasis Produced by Indoor Unventilated-Stove Coal Usage in Guizhou, China

Abstract

The aim of this study was to determine p16 gene mutation, deletion, and promoter 5′ CpG island hypermethylation in peripheral blood mononuclear leukocyte of patients with arseniasis as attributed to exposure to indoor unventilated coal stove. The role of the aberrant change of p16 gene in the induction and development of carcinogenesis in endemic arsenisiasis region in China was also examined. Polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP), multiplex PCR (mPCR), methylation-specific PCR (MSP), and sequencing techniques were performed to detect (1) mutation of the p16 gene exon 2, (2) homozygous deletion of the p16 gene exon 1 and exon 2, and (3) hypermethylation of the promoter CpG island in peripheral blood mononuclear leukocyte of patients with arseniasis. Results showed no mutation was found in exon 2 of p16 gene. The homozygous deletion frequency of p16 gene was 5 and 15% in control and arseniasis patients, respectively. The homozygous deletion occurred mainly in exon 2, with significant deletion frequencies of 9, 13, and 20% in mild, intermediate, and severe arseniasis groups. The significant homozygous deletion frequency was 9 and 39% in noncarcinoma and carcinoma individuals. The positive rate of p16 gene promoter CpG island hyermethylation was 42 and 2% in the exposed group and the control group, respectively. The positive rate was 26, 42, and 50% in mild, intermediate, and severe arseniasis. The marked different positive rate was 22 and 56% in noncarcinoma and carcinoma individuals, respectively. In conclusion, homozygous deletion and hypermethylation of p16 gene may play an important role in the initiation and development of manifestations seen in endemic arseniasis including carcinogenesis.

This study was supported by the National Natural Science Foundation of China(30460123); by the Scientific and Technique Talented Persons & Nomarch Special Foundation of Guizhou, China ([2004]07); and by the Natural Science Foundation of Guizhou, China ([2005] 400115). We acknowledge the 44th hospital of PLA, China, Xinren Hygienic and Anti-Epidemic Station for assistance in the investigation. In particular, we thank the villagers from Xinren county for their participation and corporation in this investigation.