| Objective: DNA methylation is an important mechanism for regulating gene expression,and it plays an important role in tumor occurrence and development. Studies have shown that, in aspect of tumor diagnosis, prognosis and therapeutic evaluation,DNA methylation is a potential value of tumor markers. In this study, we create a real-time quantitative PCR methylation detection method to detect the expression of GSTP1 gene methylation in free DNA of peripheral blood in patients with prostate cancer, and to investigate its value in early diagnosis of prostate cancer.Methods: Collected peripheral blood samples of patients hospitalized, each patient two tubes, about 2ml / tube,then cryopreserved them in-80 ? ultra-low temperature freezer.Wherein, the samples met the inclusion criteria were: 58 endocrine therapy patients with prostate cancer as group A whose age fluctuated from 61 to 82 years, the average age was 72 years; 42 cases visited the hospital because of "difficulty urinating, found elevated PSA" as group B, and 23 patients were diagnosed with prostate cancer(B1), 19 patients were diagnosed with non prostate cancer(B2), the age of group B fluctuated from 55 to 78 years, the average age was 70 years old; 34 cases male patients of non-prostate disease as group C, whose PSA values within the normal reference range,for the same period due to urethral stricture, kidney stones, renal cysts and other benign diseases of treatment, the age fluctuated from 58 to 76 years, the average age was 68 years old. For the three groups, we detected GSTP1 gene qualitative expression with methylation-specific PCR method and GSTP1 methylation relative expression with quantitative PCR, then analyzed the relationship with the main statistical of clinical and pathological features of prostate patients.Results: 1. The relative expression(M / U) of GSTP1 methylation of free DNA in peripheral blood of patients in group A was 0.330973310 ± 0.1848315242, the relative expression of B2 was 1.585767684 ± 0.8382501872, the relative expression of group C was 2.354086941 ± 2.1664719163, Welch test P = 0.000 <0.05, so A, B2, C three groups were not all equal at overall mean. Pairwise comparisons, A, B2 overall mean were significant difference(P = 0.000); A, C groups overall mean were significantly different (P = 0.000); B2, C overall mean were not significant difference(P = 0.230). We considered methylation ratio of GSTP1 gene of group A was less than B2, C two groups.2. The relative expression(M / U) of GSTP1 methylation of group B1 was 0.304547000± 0.1905957866, comparative analysis with group B2, Welch test P = 0.000 <0.05, the overall means were significant difference between the two groups. 3. Analyszed group A and B1 using independent sample t test, P = 0.806 > 0.05 was, the overall means were no significant difference between the two groups. 4. Correlation analyszed the degree of GSTP1 gene methylation of prostate cancer patients with corresponding PSA,Kendall'tau_b, Spearman'rho two non-parametric methods test, Kendall correlation coefficient was 0.198, Spearman correlation coefficient was 0.263, both P <0.05,indicated the correlation was not significant(correlation coefficient is less than 0.5)between the relative expression of GSTP1 gene methylation of free DNA in peripheral blood of patients with prostate cancer and PSA. 5. The overall means of GSTP1 methylation were not significant difference(P = 0.930) between Gleason score of levels5 to 7 points and 8 to 10, the overall means were not significant difference(P = 0.241)between risk classification of low-risk and high-risk.Conclusion: 1. Methylated and unmethylated expression of GSTP1 gene in free DNA are present in prostate cancer patients and benign prostatic patients; 2 The ratio values(M / U) of prostate cancer patients is lower than patients with benign lesions(BPH etc.),presumably feedback protective effect against tumors of the body; 3. endocrine therapy for prostate cancer patients has no effect to relative expression of GSTP1 gene methylation in free DNA; 4. the correlation was not significant(r <0.5) between the relative expression of GSTP1 gene methylation in free DNA of peripheral blood of prostate cancer patients and PSA, which can be used as an independent biomarker for screening prostate cancer; 5. Though detecting the relative expression of GSTP1 gene methylation in patients' s peripheral blood can not predict Gleason score and risk grade of prostate cancer, but it has a certain value to improve the accuracy of early diagnosis of prostate cancer. |
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