title: Connexin 32 and 43 promoter methylation in Helicobacter pylori-associated gastric tumorigenesis.
abstract: OBJECTIVE
  To explore the mechanism of abnormal Connexin (Cx) 32 and Cx43 expression in the gastric mucosa after Helicobacter pylori (H. pylori) infection.
  
  
  METHODS
  Biopsy specimens of gastric mucosa in different gastric carcinogenesis stages with H. pylori infection, that is, non-atrophic gastritis (NAG; n = 24), chronic atrophic gastritis (CAG; n = 25), intestinal metaplasia (IM; n = 28), dysplasia (DYS; n = 24), and gastric cancer (GC; n = 30), as well as specimens of normal gastric mucosa without H. pylori infection (NGM; n = 25), were confirmed by endoscopy and pathological examination. Cx32 and Cx43 mRNA expression was detected by real-time polymerase chain reaction (PCR). Cx32 and Cx43 promoter CpG island methylation status was determined by methylation-specific PCR (MSP), bisulfite PCR sequencing (BSP) and MassArray methods.
  
  
  RESULTS
  The relative mRNA expression levels in the gastric mucosa of patients with NGM, NAG, CAG, IM, DYS and GC were 0.146 ± 0.011, 0.133 ± 0.026, 0.107 ± 0.035, 0.039 ± 0.032, 0.037 ± 0.01 and 0.03 ± 0.011 for Cx32; and 0.667 ± 0.057, 0.644 ± 0.051, 0.624 ± 0.049, 0.555 ± 0.067, 0.536 ± 0.058 and 0.245 ± 0.121 for Cx43, respectively, which were gradually decreasing and significantly different (GC vs NGM: P < 0.001 for Cx32, P < 0.001 for Cx43). The promoter methylation levels in the gastric mucosa from NGM to GC stages by MSP were 38.8% ± 9.0%, 43.1% ± 9.4%, 56.5% ± 3.1%, 64.4% ± 9.7%, 72.5% ± 4.2% and 79.6% ± 6.8% for Cx32; and 49.0% ± 3.9%, 58.1% ± 5.0%, 66.5% ± 7.9%, 74.0% ± 8.8%, 78.3% ± 3.6% and 88.7% ± 6.2% for Cx43, respectively, which were gradually increasing and significantly different (P = 0.039, P = 0.019). The promoter methylation levels by BSP and MassArray exhibited similar trends. Cx32 and Cx43 mRNA expression was negatively correlated with promoter methylation status and gastric carcinogenesis stages (P < 0.001, P = 0.016).
  
  
  CONCLUSIONS
  Cx32 and Cx43 mRNA expression decreased gradually during H. pylori infection-associated gastric carcinogenesis, and it is associated with hypermethylation of these genes' promoter.
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Li-Hua
authors.initials: LH
authors.lastname: Huang
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Can-Xia
authors.initials: CX
authors.lastname: Xu
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Xiao
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Li
authors.initials: L
authors.lastname: Zhou
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Dan
authors.initials: D
authors.lastname: Cao
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Xiao-Min
authors.initials: XM
authors.lastname: Liu
authors.affiliation: Yu Wang, Can-Xia Xu, Jing Xiao, Li Zhou, Dan Cao, Xiao-Min Liu, Department of Gastroenterology, Third Xiangya Hospital, Central South University, Changsha 410013, Hunan Province, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Qi
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics*; D001706:Biopsy; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000473:pathology; D018031:Connexin 43 / Q000235:genetics*; D017630:Connexins / Q000235:genetics*; D018899:CpG Islands; D019175:DNA Methylation; D018450:Disease Progression; D015536:Down-Regulation; D005753:Gastric Mucosa / Q000382:microbiology / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D016481:Helicobacter Infections / Q000150:complications / Q000382:microbiology*; D016480:Helicobacter pylori / Q000472:pathogenicity*; D006801:Humans; D009367:Neoplasm Staging; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000032:analysis; D013274:Stomach Neoplasms / Q000235:genetics* / Q000382:microbiology* / Q000473:pathology; D000097002:Gap Junction beta-1 Protein
chemical_list: D014408:Biomarkers, Tumor; D018031:Connexin 43; D017630:Connexins; D012333:RNA, Messenger; D000097002:Gap Junction beta-1 Protein; C500216:GJA1 protein, human
keywords: Cx32; Cx43; DNA methylation; Gastric cancer; Helicobacter pylori