Toshimi Otsuka

Influence of HRH2 promoter polymorphism on aberrant DNA methylation of DAPK and CDH1 in the gastric epithelium

BackgroundAberrant methylation patterns in CpG island are known to be influential in gene silencing. Histamine plays important physiological roles in the upper gastrointestinal tract and acts via the H2 receptor. We report an investigation into the effect of HRH2 promoter polymorphism (rs2607474 G > A) on the methylation of DAPK and CDH1.MethodsNon cancerous gastric mucosa samples were obtained from 115 subjects with gastric cancer (GC) and 412 non-cancer subjects (non-GC). Methylation status of genes was determined by MSP. The genotyping of rs2607474 was performed by PCR-SSCP.ResultsMethylation of DAPK and CDH1 was observed in 296 and 246 subjects, respectively. The frequency of CDH1 methylation in the subjects with GC was significantly lower in cancer lesion than in non cancerous mucosa, whereas that of DAPK methylation was not different. The allelic distribution of rs2607474 was 401GG, 119GA and 7AA. The GG homozygote was associated with a significantly increased risk for methylation of both DAPK and CDH1 (p < 0.0001 and p = 0.0009, respectively). In the non-GC subjects or more than 60 years of age, GG homozygote was more closely associated with both DAPK and CDH1 methylation. However, this genotype did not show an increased risk for the development of methylation of both genes in patients with GC. In H. pylori negative subjects, GG homozygote showed an increased risk for the methylation of both DAPK and CDH1 (p = 0.0074 and p = 0.0016, respectively), whereas this genotype was associated with an increased risk for the development of DAPK methylation in H. pylori positive subjects (p = 0.0018). In addition, in subjects older than 60 years of age, atrophy and metaplasia scores were significantly higher in the GG homozygote (p = 0.011 and p = 0.039, respectively) and a significant correlation was observed between age and atrophy or metaplasia.ConclusionsOur results suggest that rs2607474 GG homozygote confers a significantly increased risk for age- and inflammation-related DAPK and CDH1 methylation.

Genetic polymorphisms of IL17A and pri-microRNA-938, targeting IL17A 3'-UTR, influence susceptibility to gastric cancer.

We report an association between gastric cancer (GC) and polymorphisms in IL17A, rs2275913 (-197 G > A), rs3748067 (*1249 C > T), and pri-miR-938, rs2505901 (T > C). We employed the multiplex PCR-SSCP method to detect gene polymorphisms in 337 GC cases and 587 controls. The minor allele frequency of rs2275913 was significantly higher, and those of rs3748067 and rs2505901 significantly lower, in GC cases than controls. The rs2275913 AA homozygote was associated with an increased risk (OR, 2.38; 95%CI, 1.63-3.46; p < 0.0001) for the development of both intestinal and diffuse types of GC. The rs3748067 T polymorphism was associated with a decreased risk for intestinal GC (OR, 0.511; 95%CI, 0.272-0.962; p = 0.037), whereas rs2505901 C locus carried a decreased risk overall for GC (OR, 0.733; 95%CI, 0.545-0.985; p = 0.039). In addition, rs3748067 T allele was inversely correlated with lymph node metastasis. Our results suggest that polymorphisms in both IL17A and pri-miR-938 contribute to cancer risk susceptibility and therefore can affect the development of gastric cancer.

Association between functional promoter polymorphisms of macrophage migration inhibitory factor (MIF) gene and ulcerative colitis in Japan.

Macrophage migration inhibitory factor (MIF) is a key proinflammatory mediator. Two functional polymorphisms were identified in the promoter region of MIF gene. We attempted to clarify the associations between these polymorphisms and ulcerative colitis (UC). The study was performed in 111 patients with UC and 209 subjects without UC. We employed the PCR-SSCP method to detect gene polymorphisms. Overall, 5/5-CATT genotype was a decreased risk for the development of UC (OR, 0.51; 95% CI, 0.26-0.99). In addition, 7/7-CATT genotype was significantly associated with chronic continuous phenotype and distal colitis phenotype (OR, 5.49; 95% CI, 1.19-25.3, and OR, 6.10; 95% CI, 1.32-28.2, respectively), whereas 5/5-CATT genotype had an inhibitory effect on the development of UC after 20years of age (OR, 0.33; 95% CI, 0.14-0.82). On the other hand, G-173C polymorphism did not affect the susceptibility to and the phenotypes of UC. Our results suggested that tetranucleotide CATT repeat of MIF gene promoter may be associated with the development of UC and the severity of inflammation in patients with UC.

Functional promoter polymorphisms of NFKB1 influence susceptibility to the diffuse type of gastric cancer

In the present study, we report an association between gastric cancer and polymorphisms in NFKB1 (rs28362941 and rs78696119). We employed the PCR-SSCP method to detect gene polymorphisms in 479 gastric cancer cases and 880 controls. The rs28362941 del/del homozygote was significantly associated with gastric cancer development; in particular it was closely associated with diffuse type gastric cancer. The rs78696119 GG homozygote was also associated with the diffuse type of gastric cancer. In young subjects, both polymorphisms were significantly associated with the development of gastric cancer. In addition, both polymorphisms were related to tumor progression such as tumor invasion and lymph node metastasis. The inflammatory cell infiltration into non-cancerous gastric mucosa was greater in the subjects with the rs28362491 del/del or rs78696119 GG genotype when compared to those with the other genotypes. In conclusion, functional polymorphisms of NFKB1 are associated with an increased risk of gastric cancer; in particular they are closely associated with the development of diffuse type of gastric cancer via severe gastric inflammation. These polymorphisms also appear to be associated with gastric cancer progression.

NFKB1 polymorphism is associated with age-related gene methylation in Helicobacter pylori-infected subjects

CpG island aberrant methylation is shown to be an important mechanism in gene silencing. The important role of NF-κB in the inflammatory response to H. pylori colonization has been indicated. We investigated the influence of NFKB1 polymorphisms, -94 ins/del (rs28362491) and -449 C>G (rs72696119), on the aberrant gene methylation under H. pylori infection. Gastric mucosal samples were obtained from sub-subjects without malignancies. Methylation status of genes (p14ARF, p16INK4a, DAPK and CDH1) was determined by methylation-specific PCR (MSP). The genotyping of NFKB1 was performed by PCR-SSCP. There was a strong allelic association between rs28362491 and rs72696119, and all H. pylori-infected -94 del/del homozygotes had a -449 GG genotype. The -94 del/del homozygosity was significantly associated with risk for development of CpG island high methylation (CIHM) (two or more gene methylations), especially DAPK and CDH1 methylations, and the number of methylated genes was significantly higher in -94 del/del homozygotes than in ins/del and ins/ins (ins carrier) H. pylori-infected elder subjects. In addition, this methylated gene number was significantly increased with age in H. pylori-infected del/del homozygotes, but not in infected ins carriers. Furthermore, the inflammation score was significantly higher in H. pylori-infected del/del homozygotes compared to ins carriers. NFKB1 -94 ins/del ATTG polymorphism (rs28362491) was significantly associated with the increased risk for the development of age-related gene methylation in non-cancerous gastric mucosa under H. pylori-induced inflammation.

Genetic polymorphisms of MAFK, encoding a small Maf protein, are associated with susceptibility to ulcerative colitis in Japan

AIM To investigate whether single nucleotide polymorphisms in maf protein K (MAFK), which encodes the MAFK, lead to increased susceptibility to ulcerative colitis in the Japanese population. METHODS This case control study examined the associations between MAFK single nucleotide polymorphisms (rs4268033 G>A, rs3735656 T>C and rs10226620 C>T) and ulcerative colitis susceptibility in 174 patients with ulcerative colitis (UC) cases, and 748 subjects without no lower abdominal symptoms, diarrhea or hematochezia (controls). In addition, as the second controls, we set 360 subjects, who have an irregular bowel movement without abnormal lower endoscopic findings (IBM controls). RESULTS The genotype frequency of rs4268033 AA and allelic frequency of the rs4268033A allele were significantly higher in the UC cases than in both controls (P = 0.0005 and < 0.0001, P = 0.015 and 0.0027 vs controls and IBM controls, respectively). Logistic regression analysis after adjustment for age and gender showed that the rs4268033 AA and rs3735656 CC genotypes were significantly associated with susceptibility to UC development (OR = 2.63, 95%CI: 1.61-4.30, P = 0.0001 and OR = 1.81; 95%CI: 1.12-2.94, P = 0.015, respectively). Similar findings were observed by the comparison with IBM controls. In addition, the rs4268033 AA genotype was significantly associated with all phenotypes of UC except early onset. There was no significant association between rs10226620 and ulcerative colitis. CONCLUSION Our results provide the first evidence that MAFK genetic polymorphisms are significantly associated with susceptibility to UC development. In particular, rs4268033 is closely associated with an increased risk for the development of UC.

Association between receptor interacting serine/threonine kinase 2 polymorphisms and gastric cancer susceptibility

The present study aimed to investigate whether single nucleotide polymorphisms in receptor interacting serine/threonine kinase 2 (RIPK2), which encodes a component of the nucleotide binding oligomerization domain containing 2-RIP2 pathway, may compromise the innate immune response to Helicobacter pylori infection, leading to increased susceptibility to gastric cancer in the Japanese population. The present case control study investigated the associations between RIPK2 single nucleotide polymorphisms and gastric mucosal inflammation, atrophy and cancer susceptibility in 528 patients with gastric cancer and 697 patients without gastric malignancies on upper gastro-duodenal endoscopy. Overall, the RIPK2 rs16900627 minor allele was significantly associated with the susceptibility to gastric cancer [OR, 1.37; 95% confidence interval (CI), 1.06-1.77; P=0.016], particularly of the intestinal type (OR, 1.53; 95% CI, 1.13-2.07; P=0.0062). It was also significantly associated with gastric mucosal atrophy (OR, 1.83; 95% CI, 1.14-2.93; P=0.011). When assessing the severity of chronic gastritis using the updated Sydney system, the activity and inflammation scores, as well as atrophy and metaplasia scores, were significantly higher in rs16900627 minor allele carriers compared with wild-type homozygotes. In patients younger than 60 years old, the pepsinogen I/II ratio was significantly lower in rs16900627 minor allele carriers compared with wild-type homozygotes (P=0.037). The rs16900627 minor allele is associated with the severity of gastric mucosal inflammation and the development of gastric mucosal atrophy. Carriers of this allele may have an increased risk for the development of gastric cancer, particularly of the intestinal type.

Su1501 Morphological Changes of Intraductal Papillary Mucinous Neoplasm of the Pancreas Are Not Enough for Indicators of the Development of Pancreatic Carcinoma During Follow Up

G A A b st ra ct s receiver operator characteristics curve. The factors associated with malignant IPMNs were determined by logistic regression analysis for BD-IPMN and MD-IPMN separately. Diagnostic accuracy of the determined predictors of malignancy was calculated. Results: Total 241 patients consisted of 202 BD-IPMN and 39 MD-IPMN were included. Cutoff values of cyst size, MPD diameter, and nodule size were determined for BD-IPMN as 30 mm, 6 mm, and 10 mm, respectively, whereas for MD-IPMN as 24 mm, 12 mm and 10 mm, respectively. For BD-IPMN, nodule size ≥10 mm (OR=176.97, p<0.0001) and positive cytology (OR127.14, p<0.0001) were significantly associated with malignant IPMN (Table 1). For MD-IPMN, nodule size ≥10 mm (OR=16.07, p=0.002) and positive cytology (OR=160.00, p<0.0001) were significantly associated with malignant IPMN. Sensitivity, specificity, and overall diagnostic accuracy of nodule size ≥10 mm alone were 59%, 98%, 90%, respectively, and positive cytology alone were 83%, 96%, 94%, respectively (Table 2). In combination of nodule size ≥10 mm and/or positive cytology, sensitivity, specificity, and overall diagnostic accuracy were 96%, 95%, and 95%, respectively. Conclusions: The common predictors, nodule size ≥10 mm and positive cytology, were demonstrated to be appropriate for both BD-IPMN and MD-IPMN. In combination of nodule size ≥10 mm and/or positive cytology as predictors of malignancy, diagnostic accuracy was shown to be excellent. Table 1. Factors associated with malignant IPMN determined by logistic regression analysis.

Mo1366 Genetic Polymorphisms of MAFK, Encoding Small Maf Protein MAFK, Are Associated With the Susceptibility to Ulcerative Colitis

[Background] The small Maf proteins have emerged as crucial regulators of mammalian gene expression. Small Mafs do not contain an obvious transcriptional activation domain. However, it has been clear that small Maf proteins are regulated transcription factors, such as Nrf2 or Bach1. MafK, one of the small Maf proteins, is encoded by MAFK gene. Within 20kbp around MAFK gene, there are two linkage disequilibrium blocks, with above 0.05 of HWE p value and above 0.05 of minor allele frequency. In this study, we investigated an association between ulcerative colitis (UC) and each tag polymorphism, rs4268033G.A and rs3735656 (*910T.C), and the other one rs10226620 (*1506T.C). [Material and Methods] The studied population comprised 922 subjects, including patients with UC (UC cases, n=174), who were enrolled Fujita Health University Hospital, and the subjects without UC (controls, n=748). We employed the PCR-SSCP method to detect a gene polymorphism. [Results] In controls, mean age was 57.1 years old and male/female ratio was 438/310. In UC cases, mean age was 40.3 years old, mean age of onset was 33.0 years old and male/ female ratio was 98/76. The distributions of rs4268033, rs3735656 and rs10226620 in controls were 366GG, 324GA and 58AA (HWE p=0.25), 329TT, 346TC and 73CC (HWE p=0.21) and 345TT, 328TC and 75CC (HWE p=0.87) respectively, whereas the distributions in UC cases was 80GG, 66GA and 28AA, 80TT, 69TC and 25CC, 80TT, 72TC and 22CC, respectively. The genotype frequency of rs4268033 AA and allelic frequency of rs4268033A allele were significantly higher in UC cases than in controls (p=0.0013 and 0.045 by Fisher). By logistic regression analysis after adjustment for age and gender, rs4268033 AA and rs3735656 CC genotypes were significantly associated with the susceptibility to UC development (OR, 2.62; 95%CI, 1.54-4.48; p=0.0004 and OR, 1.79; 95%CI, 1.05-3.04; p=0.031, respectively). In addition, rs4268033 AA genotype was significantly associated with all phenotypes of UC (i.e. total colitis phenotype: OR, 2.58; 95%CI, 1.29-5.17; p=0.0073, not total colitis: OR, 2.39; 95%CI, 1.22-4.72; p=0.012). The rs3735656CC genotype was significantly associated with steroid-independent or not required phenotype of UC (OR, 1.87; 95%CI, 1.08-3.26; p=0.026). There was no significant association between rs10226620 and UC. [Conclusion] Our results provided the first evidence that MAFK gene polymorphisms were significantly associated with the susceptibility to UC development. In particular, rs4268033 was closely associated with an increased risk for the development of UC.

Mo1131 Genetic Polymorphism of MAFK (Rs4268033) Is Associated With the Susceptibility to Gastric Cancer in Japan

[Background] The small Maf proteins have emerged as crucial regulators of mammalian gene expression. Small Mafs do not contain an obvious transcriptional activation domain. However, it has been clear that small Maf proteins are regulated transcription factors, such as Nrf2 or Bach1. MafK, one of the small Maf proteins, is encoded by MAFK gene. Within 20kbp around MAFK gene, there are two linkage disequilibrium blocks, with above 0.05 of HWE p value and above 0.05 of minor allele frequency. In this study, we investigated an association between gastric cancer (UC) and each tag polymorphism, rs4268033G .A and rs3735656 (*910T.C), and the other one rs10226620 (*1506T.C). [Material and Methods] The study was performed in 334 patients with gastric cancer (GC cases) and 661 subjects with no evidence of gastric malignancies (controls) on upper gastro-duodenal endoscopy. We employed the PCR-SSCP method to detect the gene polymorphism. [Results] The mean age, male/female ratio and Helicobacter pylori (HP) positive ratio in GC cases were significantly higher than those in controls. The distributions of rs4268033, rs3735656 and rs10226620 in controls were 326GG, 284GA and 51AA (HWE p=0.35), 291TT, 304TC and 66CC (HWE p=0.33) and 304TT, 288TC and 69CC (HWE p=0.93) respectively, whereas the distributions in GC cases was 127GG, 163GA and 44AA, 142TT, 153TC and 39CC, 144TT, 154TC and 36CC, respectively. The allelic frequency of rs4268033 Awas significantly higher in GC cases than in controls ‘(37.6% vs. 29.2%, p=0.0002). Overall, rs4268033 minor variant had a significantly increased risk for the development of gastric cancer by logistic regression analysis after adjustment for gender, age and HP infection status (recessive genetic model: OR, 1.72 95%CI, 1.10-2.69; p=0.016 and dominant genetic model: OR, 1.61; 95%CI, 1.21-2.14; p=0.0009), whereas no significant association was found between gastric cancer susceptibility and rs3735656 or rs10226620. The rs4268033 minor allele was more strongly associated with intestinal type of gastric cancer (recessive genetic model: OR, 2.12; 95%CI, 1.27-3.54; p=0.0040 and dominant genetic model: OR, 1.66; 95%CI, 1.17-2.35; p=0.0042), and slightly associated with diffuse type of gastric cancer (dominant genetic model: OR, 1.50; 95%CI, 1.02-2.21; p=0.040). In addition, rs4268033 minor allele was significantly associated with non-cardiac gastric cancer (recessive genetic model: OR, 1.76; 95%CI, 1.11-2.77; p=0.015 and dominant genetic model: OR, 1.68; 95%CI, 1.252.25; p=0.0005). [Conclusion] Our results provided the first evidence that MAFK gene polymorphism is significantly associated with the susceptibility to gastric cancer development. The rs4268033 minor allele is closely associated with an increased risk for the development of gastric cancer.