| Objective: Oral squamous cell cancer(OSCC)is one of the most common type of oral and maxillofacial tumor,accounting for 3% of human malignancies.According to statistics,every year about 46500 new cases of oral cancer in our country,the onset age wasbetween 40 to 60 years old,more men than women.As the medical level gradually improve,although multidisciplinary sequence treatment has greatly promoted the oral cancer treatment in a certain extent,the five-year survival rate is still at 55%-60%.As is known to all,the prognosis of OSCC depends on clinical stage,histological grade,lymph node metastasis and other factors,but most of the patients at the already is late when they were see a doctor.So the early diagnosis and early treatment of oral canceris very important.The study found that in addition to Genetic factors of the gene mutation and gene deletions,the change of DNA methylation and other epigenetic is closely related with the occurrence and development of tumors.Clinical application of epigenetic markers can do early diagnosis of oral squamous cell carcinomas,to predict the prognosis,and thus improve the patient's quality of life,improve the survival rate of patients.The naked cuticle NKD family includes naked cuticle homolog 1(NKD1)and 2(NKD2).Loss of heterozygosity has been frequently found in the regions where NKD1,NKD2 related in different types of tumors,including liver cancer,gastric cancer and breast cancer.The promoter region of NKD2 is hypermethylated in glioblastoma cells.A research have been reported in the magzaine that mRNA expression and the hypermethylation of NKD1,NKD2 gene in different types of tumors,such as osteosarcoma cancer,breast cancer.And we don't know its expression level and methylation status in OSCC.This article aims to analyze the expression and methylation of NKD1,NKD2 gene and know their differences in human OSCC tissues andcorresponding normaltissues.According to statistical analysis on the data of methylation results,clinical data,expression level,we can explore the function of NKD1,NKD2 in the process of OSCC occurrence and development.The purpose of this article is to provide new ideas,treatment and diagnosis of OSCC.Methods:1 The 52 patients with primary oral squamous cell carcinoma who were treated with excision surgery during Sep.2015 to Dem.2016 in the Fourth Hospital of Hebei Medical University were elected,and were compared with the corresponding adjacent normal oral mucosa tissues.The cancer tissues were proved to be squamous cell carcinoma and adjacent tissues were proved to be non-tumor tissue by pathological detection.All the tissues were used immediately after surgery and cryopreserved in-80? refrigerator.2 Reverse Transcrpition Polymerase Chain Reaction(RT-PCR)was used to detect the relative mRNA expression of NKD1,NKD2 gene in 52 OSCC and Normal tissue samples.3 The methylation status of NKD1,NKD2 gene promoter regions CpG island in 52 OSCC and normal tissue specimens was identified by methylation-specific Polymerase Chain Reaction(MSP).4 SPSS 21.0 software was employed to analyze all data.Paired samples t test,two-sample t test or separate variance estimation t test was used for detection of gene mRNA expression.Linear correlation was used for correlations of two genes mRNA expression.?2 test or Fisher's exact test was used for detection of gene methylation.Spearman rand correlation test was used for correlations of gene methylation,within which P<0.05 set for statistically differences.Results:1 The relative expression quantity of NKD1 gene mRNA in oral squamous cell carcinoma tissues was 0.50±0.12,significantly lower than that in corresponding oral mucosa tissues 0.66±0.18,the difference had statistical significance(t=19.934,P <0.001).In the meanwhile,the mRNA expression ofNKD1 gene in?+?group was 0.53±0.12,which was significantly higher than?+?group 0.44±0.10(t=-3.041,P=0.004).The mRNA expression of NKD1 gene in lymph node metastasis group was 0.44±0.10,which was lower than that in none of lymph node metastasis group 0.52±0.12(t=2.469,P=0.017).However,there was no relationship between the relative expression quantity of NKD1 mRNA and clinical data,such as age,gender and smoking(P>0.05).2 The relative expression quantity of NKD2 gene mRNA in oral squamous cell carcinoma tissues was 0.42±0.10,significantly lower than that in corresponding oral mucosa tissues0.62±0.12.The difference had statistical significance(t=10.915,P < 0.001).The mRNA expression of NKD2 gene in?+?group was 0.44±0.10,which was significantly higher than ?+?group0.37±0.05(t=-3.739,P < 0.001).The mRNA expression of NKD2 gene in lymphnode metastasis group was 0.37±0.07,which was lower than that in none of lymph node metastasis group 0.45±0.10(t=2.873,P=0.006).And the relative expression quantity of NKD2 mRNA in smoking group was0.38±0.09,significantly lower than that in no smoking group 0.45±0.08(t=-3.395,P=0.001).However,there was no relationship between the relative expression quantity of NKD2 mRNA and clinical data,such as age and gender(P>0.05).3 In the oral squamous cell carcinoma tissues,the frequency of NKD1 gene promoter region methylation was 48.08%(25/52),significantly higher than that in corresponding normal oral mucosa tissues26.92%(14/52).The difference had statistical significance(?2=4.964,P=0.026).In the meanwhile,the frequency of NKD1 gene promoter region methylation in clinical stage?+?was35.48%(11/31),significantly lower than that in clinical stage?+?66.67(14/21)(?2=4.877,P=0.027).The frequency of NKD1 gene promoter region methylation in lymph node metastasis group was70.59%(12/17),significantly higher than that none of lymph node metastasis group37.14%(13/35)(?2=5.127,P=0.024).However,there was no relationship between the frequency of NKD1 gene promoter region methylation and clinical data,such as age,gender and smoking(P>0.05).4 In the oral squamous cell carcinoma tissues,the frequency of NKD2 gene promoter region methylation was 44.23%(23/52),significantly higher than that in corresponding normal oral mucosa tissues21.15%(11/52).The difference had statistical significance(?2=6.292,P=0.012).In the meanwhile,the frequency of NKD2 gene promoter region methylation in clinical stage?+?was 29.03%(9/31),significantly lower than that in clinical stage?+?66.67%(14/21)(?2=7.188,P=0.007).The frequency of NKD2 gene promoter region methylation in lymph node metastasis group was 64.71%(11/17),significantly higher than that none of lymph node metastasis group34.29%(12/35)(?2=4.293,P=0.038).The frequency of NKD2 gene promoter region methylation in smoking group was 57.14%(16/28),significantly higher than that no smoking group 29.17%(7/24)(?2=4.1,P=0.043).However,there was no relationship between the frequency of NKD2 gene promoter region methylation and clinical data,such as age and gender(P>0.05).5 In the oral squamous cell carcinoma tissues,the relative expression quantity of NKD1 mRNA in methylated tissues was 0.44±0.11,which was lower than that in unmethylated tissues 0.54±0.11.There was significancein statistics(t=-3.344,P=0.002).And the relative expression quantity of NKD2 mRNA in methylated tissues was 0.36±0.06,which was lower than that in unmethylated tissues 0.47±0.09.There was significance in statistics(t=-4.934,P<0.001).6 In the oral squamous cell carcinoma tissues,the cases of both NKD1,NKD2 gene promoter region methylation was 16,and the cases of both NKD1,NKD2 gene promoter region ummethylation was 20.The expressions of NKD1 and NKD2 in the oral squamous cell carcinoma was positively correlated(r=0.038,P=0.005).The expressions of NKD1 and NKD2 mRNA in the oral squamous cell carcinoma was positively correlated(r=0.312,P <0.024).Conclusions:1 The result suggests that NKD1,NKD2 gene low expression or silence may leads to the occurrence and progression of the oral cancer occurrence andprogression of the oral cancer.The relative expression quantity of NKD1 gene mRNA is associated with clinical stage and lymph node metastasis,and the NKD2 gene mRNA expression quantity is associated with clinical stage,lymph node metastasis and smoking.2 The result indicates that the hypermethylation of the NKD1,NKD2 gene occuring of oral promoter may be one of the important mechanism in the cancer.At the same time,NKD1 gene promoter region methylation status is associated with clinical stage and lymphnode metastasis.And NKD2 gene promoter region methylation status is associated with clinical stage,ymphnode metastasis and smoking.3 The result suggests that NKD1,NKD2 gene promoter regions CpG island hypermethylationcould be one of the important reason for its transcriptional silencing.4 In oral squamous cell carcinomas NKD1 and NKD2 gene may exist in the process of occurrence and development of synergy. |
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