| Objective: Esophageal cancer(EC)is one of the most common malignant tumors of digestive system and it has a higher mortality rate in the world.There are obvious regional differences in EC and the morbidity of EC in China is significantly higher than that in other countries.Esophageal squamous cell carcinoma(ESCC)is the major pathological type of EC.Although in recent years,the effectiveness of ESCC in multidisciplinary treatment has improved,however,the 5 year survival rate is still low.Gastric cardia adenocarcinoma(GCA)is one of the frequent kinds of malignant tumors in the northern China.The mortality of GCA is still increasing in our country.It is hard to achieve in early diagnosis of the tumor so that majority of patients treated in hospital has been in the late stage.Yet so far,the pathogenesis of ESCC and GCA has not been completely clarified.Therefore,it is very important to find the early biomarkers of ESCC and GCA for prevention and treatment.Long non-coding RNA(lnc RNA)is a kind of non coding RNA with a length of 200 ~ 100000 nt,which can regulate gene expression.The roles of lnc RNAs in ESCC and GCA screened by the gene chip are still little known.There are few reports on the correlation of lnc RNAs with ESCC and GCA.Therefore,in this study,we aimed to detect the expression and methylation status of long non-coding RNA LOC100130476 in esophageal cancer cell lines,ESCC and GCA tissues,and to analyze the correlation between them.We further explored the relationship between clinical data including age,gender,lymph node metastasis,pathological differentiation,TNM stage and lnc RNA LOC100130476.Methods:1 Reverse transcription polymerase chain reaction(RT-PCR)method was used to detect the expression of lnc RNA LOC100130476 in four esophageal cancer cell lines(TE1,TE13,T.Tn,Eca109)treated and untreated with DNA methyltransferase inhibitor 5-Aza-2’-detoxycytidine(5-Aza-d C),72 ESCC tissue specimens and the corresponding normal tissue specimens and76 GCA tissue specimens and the corresponding normal tissue specimens.2 Methylation specific PCR(MSP)method was used to examine the methylation status of lnc RNA LOC100130476 in four esophageal cancer cell lines(TE1,TE13,T.Tn,Eca109)treated and untreated with DNA methyltransferase inhibitor 5-Aza-2’-detoxycytidine(5-Aza-d C),72 ESCC tissue specimens together with the corresponding normal tissue specimens and76 GCA tissue specimens together with the corresponding normal tissue specimens.3 SPSS13.0 was applied to analyze the results of experiments.Results:1 The expression and methylation status of lnc RNA LOC100130476 in esophageal cancer cell lines1.1 The expression of lnc RNA LOC100130476 in esophageal cancer cell lines treated and untreated with 5-Aza-d CThe expression of LOC100130476 was not detected in TE1,TE13,T.Tn cell lines.Weak positive expression of LOC100130476 was detected in Eca109 cell line.After 5-Aza-d C treatment,positive expression of LOC100130476 was detected in TE1,TE13,T.Tn cell lines and the expression level of LOC100130476 was raised in Eca109 cell line.1.2 The methylation status of lnc RNA LOC100130476 in esophageal cancer cell lines treated and untreated with 5-Aza-d CHypermethylation of LOC100130476 was detected in the four cell lines before 5-Aza-d C treatment.However,after the application of 5-Aza-d C,the methylation level of LOC100130476 was reduced in Ec109 cell line,and the other three cell lines demonstrated unmethylation status of the gene.2 The relationship between expression and methylation status of lnc RNA LOC100130476 together with ESCC clinical pathological characteristics2.1 The expression of lnc RNA LOC100130476 in ESCCLOC100130476 expression in ESCC tissues was significantly lower than that in corresponding normal tissues(0.49±0.09 vs 0.56±0.08),and the difference was statistically significant(t=-5.893,P<0.05).The expression of LOC100130476 was closely correlated with pathological differentiations and TNM stages(P<0.05),however,it was not associated with age,gender and lymph node metastasis(P>0.05).2.2 The methylation status of lnc RNA LOC100130476 in ESCCThe first exon region methylation frequency of LOC100130476 in ESCC tissues was significantly higher than that in corresponding normal tissues[69.4%(50/72)vs 26.4%(19/72)],and the difference was statistically significant(χ2=26.741,P < 0.05).It was closely related with pathological differentiations and TNM stages(P<0.05)and unrelated with age,gender and lymph node metastasis(P>0.05).The expression of LOC100130476 in ESCC tissues with LOC10013047 methylation was significantly lower than that in ESCC tissues without methylation of the gene(0.47±0.08 vs 0.55±0.08),and the difference was statistically significant(t=-4.099,P<0.05).3 The relationship between expression and methylation status of lnc RNA LOC100130476 together with GCA clinical pathological characteristics3.1 The expression of lnc RNA LOC100130476 in GCALOC100130476 expression in GCA tissues was significantly lower than that in corresponding normal tissues(0.61±0.22 vs 0.79±0.23),and the difference was statistically significant(t=-5.387,P<0.05).The expression of LOC100130476 was closely correlated with pathological differentiations and lymph node metastasis(P<0.05),however,it was not associated with age,gender and TNM stages(P>0.05).3.2 The methylation status of lnc RNA LOC100130476 in GCAThe first exon region methylation frequency of LOC100130476 in GCA tissues was significantly higher than that in corresponding normal tissues[64.5%(49/76)vs 23.7%(18/76)],and the difference was statisticallysignificant(χ2=25.649,P < 0.05).It was closely related with pathological differentiations and lymph node metastasis(P<0.05)and unrelated with age,gender and TNM stages(P>0.05).The expression of LOC100130476 in GCA tissues with LOC10013047 methylation was significantly lower than that in GCA tissues without methylation of the gene(0.56±0.21 vs 0.71±0.21),and the difference was statistically significant(t=-3.055,P<0.05).Conclusions:1 LOC100130476 may be associated with the occurrence and development of ESCC and GCA,and it may act as a tumor suppressor gene.2 The aberrant methylation status in LOC100130476 first exon region may be one of the main mechanisms in inactivating the gene in ESCC and GCA.3 The first exon region methylation frequency of LOC100130476 may be related to the malignant degree and prognosis of ESCC and GCA. |
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