The WNK2 Gene Methylation State And MRNA Expression In Renal Cell Carcinoma

Objective: Renal Cell Carcinoma(Renal Cell Carcinoma, RCC) is one of the most common tumor in the urinary system and has the high malignant degree, moreover this tumor has a tendency of distant metastases, and in recent years, the incidence of RCC is on the increase. So far, the mechanism of RCC occurrence remains obscure. Epidemiological investigation showed that some factors are perceived as contributing to RCC, such as smoking, obesity, hypertension, diabetes, heredity and so on. One of the most important factors of tumor occurrence is anti-oncogene inactivation. With the constantly development of Epigenetic, several epigenetic modifications have been found, such as DNA methylation, histone modification, chromatin remodeling, RNA interference and so on. The DNA methylation is one of the most important epigenetic modifications in tumor. Most views believe that DNA promoter methylation can result in the anti-oncogene inactivation. The study found that almost all protein kinases have a highly conserved lysine in subdomain II, but the WNK kinases lack this highly conserved lysine, hence the name “ With no K ” kinases( K stands for lysine), WNK is the abbreviation for “ With no K ”. WNK2 is a member of the WNK family, WNK2 gene resides on the chromosome 9q22.31, which is a newly discovered anti-oncogene. WNK2 gene regulates the cell cycle mainly through a signalling cross-talk between the EGF induced Ras/Raf/MEK/ERK signaling pathway and Rac/PAK signaling pathway. Initially, silencing of WNK2 expression via promoter hypermethylation was first reported in gliomas, and the WNK2 promoter hypermethylation was also found in meningioma and pancreatic ductal adenocarcinoma. But the research between WNK2 gene and RCC was not been reported. In this research, RCC tissue, tumor-adjacent tissue and normal kidney tissue specimens were used to study WNK2 gene methylation state andm RNA expression level. In this research, the relationship between the WNK2 gene and the clinical date was analysed, and the function of WNK2 gene in the process of RCC occurrence was investigated. The aim of this research is to provide novel idea for the pathogeny, diagnosis and therapy.Methods:1 WNK2 promoter methylation state testing: The WNK2 gene methylation state of 58 cases of RCC tissue and corresponding tumor-adjacent tissue and normal kidney tissue were tested by Methylation-specific PCR(Methylation Specific PCR, MSP). Meanwhile, the study aims to investigate the function of the WNK2 gene methylation in development of RCC, and analyse the relationship between WNK2 promoter methylation and clinical date.2 WNK2 gene mRNA relative expression quantity testing: Relative expression quantity of WNK2 gene in 58 cases of RCC tissue and corresponding tumor-adjacent tissue and normal kidney tissue was analysed by reverse transcription-PCR(RT-PCR). And the relationship between WNK2 m RNA relative expression quantity in RCC tissue and clinical date was detected in the research.3 The SPSS 20.0 was used for statistical analysis.Results:1 The WNK2 gene methylation state in RCC tissuesThere are three different methylation state of WNK2 gene in RCC tissues: methylated state, hemimethylated state and unmethylated state.2 The relationship between WNK2 methylation state in RCC tissues and clinical date.2.1 Comparing the methylation states in RCC tissues, tumor-adjacent tissues and normal kidney tissues.Positive ratio of WNK2 gene DNA promoter methylation in RCC tissues and tumor-adjacent tissues and normal kidney tissues are 65.5%(38/58), 34.4%(20/58) and 10.3%(6/58), respectively. There is significant difference among the three groups(X2=38.161,P=0.000). RCC tissues methylationpositive ratio was significant higher than the tumor-adjacent tissues(X2=11.172,P=0.001) and the normal kidney tissues(X2=37.495,P=0.000). 2.2 The relationship between the WNK2 promoter methylation state in RCC tissues and clinical date.In the 58 RCC tissues, there is no obviously relation between WNK2 promoter methylation state and gender(P=0.820), age(P=0.134), various clinical stages(P=1.000), tumor size(P=0.718).3 The relative expression quantity of WNK2 m RNA in 58 RCC tissues and corresponding tumor-adjacent tissues and normal kidney tissues, and the relationship between the relative expression quantity of WNK2 gene in RCC tissues and clinical date.3.1 The relative expression quantity of WNK2 m RNA in 58 RCC tissues, 58 tumor-adjacent tissues and 58 normal kidney tissues are 0.54±0.03 、0.66±0.05、0.68±0.03, respectively. There is obviously difference among the three groups(P<0.05). It is obviously that WNK2 m RNA relative expression quantity in RCC tissues lower than in tumor-adjacent tissues(P<0.05) and normal kidney tissues(P<0.05).3.2 There is no obviously correlation between WNK2 m RNA relative expression quantity and gender(P=0.142), age(P=0.217), tumor size(P=0.145).4 The relative expression quantity of WNK2 in methylation group in RCC tissues is 0.52±0.04, which is lower than in unmethylation group(0.57±0.02), there is significantly difference between methylation group and unmethylation group.Conclusions:1 The WNK2 promoter methylation positive ratio in normal kidney tissues, tumor-adjacent tissues and RCC tissues in turn increase, suggesting that WNK2 promoter methylation may participate in the tumorigenesis of RCC.2 The relative expression quantity of WNK2 gene in normal kidney tissues, tumor-adjacent tissues and RCC tissues in turn decrease, indicatingthat downregulation of the WNK2 gene expression may cause the tumorigenesis of RCC.3 In RCC tissues, the relative expression quantity of WNK2 in methylation group is lower than in unmethylation group, there is significantly difference between methylation group and unmethylation group, suggesting that WNK2 promoter hypermethylation may be a factor which can cause the downregulation of WNK2 gene m RNA expression.