The Expressions And Relationship Of Cytoplasmic Rap1 And Nuclear NF-κBp65 Expression In Gastric Carcinoma

AimGastric cancer is a common malignant tumor of the human digestive tract,which seriously threatens to human health A retrospective sampling survey on malignant tumors in China found that the incidence rate from gastric cancer ranked second and the mortality rate from gastric cancer ranked third.The vast majority of patients is in the advanced gastric cancer.The diagnosis、treatment and prognosis of gastric cancer is still not optimistic.It is important to do basic and clinical research of gastric cancer. Mammalian telomeric chromatin localized at the ends of Chromosomes,is composed of tandem repeats of the TTAGGG sequence bound by a specialized multiprotein complex known as shelterin.Shelterin complex, consisting of TRF1, TRF2, TPP1, RAP1, TIN2, and POT1.Telomeres protect the ends from fusion、degradation and recombination.As cells divide, telomeres gradually shorten to a certain extent and DNA semiconservative replication cannot continue,which leads the cells to senescence and death.Shelterin complex has been shown to regulate telomere function and be involved in cancer and aging.Recent studies suggest shelterin complex has the telomeric and extra-telomeric functions.Rap1 is a TRF2-interacting protein in the shelterin.Human cells contain a considerable fraction of Rap1 in the cytoplasm. The cytoplasmic fraction of Rap1 was not affected by TRF2 levels and may not be associated with TRF2.Nucleus Rap1 expression was significantly suppressed after TRF2 knockout.Rap1 in the cytoplasm controls gene expression through binding to extratelomeric sites of the(TTAGGG)2 consensus motif.Teo et al. reported an unanticipated role for cytoplasmic Rap1 in the modulation of NF-κB signaling.Rap1 associates with IκB kinases(IKK) to phosphorylate the p65 subunit of NF-κB.Human breast cancers with NF-κB hyperactivity show elevated levels of cytoplasmic Rap1.Repression of Rap1 sensitizes breast cancer cells to apoptosis.However, there is still little information available on the role and function of Rap1 in gastric carcinoma.This study investigates the expression of cytoplasmic Rap1 and nuclear NF-κB p65 in gastric cancer、paracancerous tissues and normal gastric mucosa tissues and analyze the relationship with clinicopathological features.This study discusses the possible role and clinical significance of the two proterin in gastric carcinoma. Materisls and methodsThe gastric cancer tissues、tumor-adjacent tissues( less than 3 cm from the edge of gastric carcinoma tissues) and normal gastric tissues( more than 3 cm away from the edge of gastric carcinoma tissues) were selected from 90 patients,undergoing surgical resection from April 2013 to July 2015 at the First Affiliated Hospital of Zhengzhou University.All the patient did not receive thermotherapy、radiotherapy and biotherapy prior to surgery.There were 47 males and 43 females, aged between 32 and 80 years(medianage 54 years).Thirty-five cases were at stage TNMⅠ~Ⅱ,55 cases were at stage TNM Ⅲ ~ Ⅳ.30 cases were classified as well-differentiated adenocarcinomas,28 were moderately differentiated adenocarcinomas, and 32 were poorly differentiated adenocarcinomas.Lymphadenopathy was found in 52 cases. In the depth of invasion,62 cases were of serosal invasion, 28 cases were without serosal infiltration.All studies were approved by the First Affiliated Hospital of Zhengzhou University.Written consents were obtained from each patient.All specimens were fixed in 40 g/L formaldehyde.After embedded in paraffin,four-micrometer-thick sections were cut of the tissue.The sections were deparaffinized, rehydrated and processed using the mmunohistochemical staining method,in accordance with the instruction of test kits.The known positive gastric cancer tissue samples were as the positive control, with PBS buffer as a negative control.SPSS17.0 software was used for statistical analysis.The differences between groups were compared using Chi-squared test and their correlations were annlyzed using Spearman rank correlation analysis.A P value of <0.05 is considered statistically significant. Results(1) Rap1 was expressed in the cytoplasm and nucleus.The positive expression of Rap1 protein was located in the cytoplasm as brown or dark brown granules.More Cytoplasm Rap1 expression was observed in gastric cancer tissues,compared to normal gastric tissue and tumor-adjacent tissue.The positive rate of Rap1 expression in normal gastric tissue,tumor-adjacent tissue and the gastric cancer tissues was 22.2%,37.8% and 75.6%. The differences between the groups had statistical significance( P < 0.05).(2)NF-κBp65 was expressed in the cytoplasm and nucleus.The positive expression of Rap1 protein was located in the nucleus as brown or dark brown granules.The positive rate of NF-κBp65 expression in normal gastric tissue,tumor-adjacent tissue and the gastric cancer tissues was 24.5%,46.7% and 70.0%. The differences between the groups had statistical significance( P < 0.05).(3)Cytoplasmic Rap1 levels correlated positively with differentiation grade, lymph node metastasis,depth of invasion and Tumor-Node-Metastasis stage with a statistic P < 0.05 in each parameter. No correlation was observed between cytoplasmic Rap1 level and age or gender. Nuclear NF-κBp65 levels correlated positively with differentiation grade, lymph node metastasis,depth of invasion and Tumor-Node-Metastasis stage with a statistic P < 0.05 in each parameter. No correlation was observed between Nuclear NF-κBp65 level and age or gender.(4)High cytoplasmic Rap1 expression levels was significantly correlated with high nuclear NF-κB levels(r=0.241,P<0.05). Conclusion(1) Significantly higher expression of cytoplasmic Rap1 was observed in gastric cancer tissues,compared to normal gastric tissue and tumor-adjacent tissue.The expression of cytoplasmic Rap1 was associated with differentiation grade,lymph node metastasis,depth of invasion and Tumor-Node-Metastasis stage,but not with age and gender.Cytoplasmic Rap1 may be involved in the carcinogenesis and progression of gastric cancer.(2) Significantly higher expression of Nuclear NF-κBp65 was observed in gastric cancer tissues,compared to normal gastric tissue and tumor-adjacent tissue.The expression of Nuclear NF-κBp65 was associated with differentiation grade,lymph node metastasis,depth of invasion and Tumor-Node-Metastasis stage,but not with age and gender.Nuclear NF-κBp65 may be involved in the carcinogenesis and progression of gastric cancer.(3) There was a positive correlation between the expressions of cytoplasmic Rap1 and that of Nuclear NF-κBp65 in gastric adenocarcinoma.Cytoplasmic Rap1 may correlates with the NF-κB signaling pathway in the gastric cancer.They may be synergistically involved in the carcinogenesis of gastric cancer.