| Part 1: Detection of m6 A levels in the whole RNA of peripheral blood in patients with acute myeloid leukemia Objective: To explore the changes of m6A(N6-methyladenosine)levels in the whole RNA of peripheral blood and the change methylation levels after treatment with decitabine in patients with acute myeloid leukemia(AML),and analyse the relationship betwewn the clinical features and the prognosis.Methods: Total of 40 cases of AML patients(22 males,18 females,median age 60 years)were enrolled in this study from Jan,2015 to Mar,2017.The patients were treated with chemotherapy based with decitabine(15-20mg/(m2·d)×3~5d).All the AML patients were divided into three groups according to their clinical status which were de novo group(15 cases),relapsed/ refractory group(8 cases),remission group(17 cases)which was also used as a positive control.The peripheral blood were collected at-1,7 and 14 day of combination chemotherapy from all the patients.The m6 A levels was detected by using the of colorimetric method.The other 18 patients with iron deficiency anemia were as the normal control.Result: 1)The m6 A levels in de novo AML,relapsed/refrctory AML,postivie and normal controls were 0.541±0.099 ng,0.320±0.048 ng,0.245±0.036 ng and 0.263±0.040 ng respectively.The m6 A level of newly diagnosed and relapsed patients was higher than those in normal and positive controls(p<0.05),the newly diagnosed group is the highest in all groups(p <0.05).2)After teratment with decitabine,In de novo patients post-treatment 7 day and 14 days,m6 A content was 0.252±0.030 ng,and 0.241±0.057 ng,significant differences(p <0.05);relapsed/ refractory patients after treatment(7 day and 14 days)m6A content was 0.251±0.033 ng and 0.227±0.030 ng,of which post-treatment 7day and 14 days than per-treatment,but insignificantly decreased(p >0.05).positive control group post-treatment 7 day and 14 days m6 A content with per-treatment significantly decreased(p<0.05);3)After treatment,patients who were CR and NR,the m6 A content was 0.545±0.168 and 0.386±0.090,respectively.Compared with the control group,CR group close to NR,which significantly higher than normal(p< 0.05).4)Per-treatment m6 A levels no significant correlation with gender,age,FAB type,original cell,white blood cell count,hemoglobin,and platelet.5)Per-treatment m6 A levels has an effect on OS and PFS of AML,whose m6 A levles below 0.359 have a longer time of OS and DFS.Conclusion: The genome methylation in the AML patients of de nove and relapsed/refractory was increase,post-treatment with decitabin the methylation of genome was reduced and close to normal after remission;the OS and DFS is shorter in the patients who has a highter level of global methylation.Part 2: Detection the expression of UCK1、UCK2 and DCK genes of peripheral blood in patients with acute myeloid leukemiaObjective: To explore the changes of the expression level of UCK1、UCK2 and DCK genes in the peripheral blood and the change methylation levels after treatment with decitabine in patients with acute myeloid leukemia(AML),and analyse the relationship betwewn the clinical features and the prognosis.Methods: Total of 40 cases of AML patients(22 males,18 females,median age 60 years)were enrolled in this study from Jan,2015 to Mar,2017.The patients were treated with chemotherapy based with decitabine(15-20mg/(m2·d)×3~5d).All the AML patients were divided into three groups according to their clinical status which were de novo group(15 cases),relapsed/ refractory group(8 cases),remission group(17 cases)which was also used as a positive control.The peripheral blood were collected at-1,7 and 14 day of combination chemotherapy from all the patients.The gene expression levels was detected by using the of q RT-PCR method.The other 18 patients with iron deficiency anemia as the normal control.Result: 1)the ΔCt value of DCK gene in AML patients before treated shows: postivie and normal controls were higher than the de novo AML and relapsed/refrctory AML respectively.The relapsed patients get the lowest expression of DCK gene(p <0.05),in the contrary,the ΔCt value of UCK2 gene in AML patients in de novo AML and relapsed/refrctory AML higher than the postivie and normal controls respectively.The relapsed patients get the highest expression of UCK2 gene(p <0.05);while the expression of UCK1 in four groups are similar(p >0.05).2)After teratment with decitabine,In de novo patients post-treatment 7 day and 14 days,the ΔCt value of DCK gene was 7.984±0.836,and 12.842±1.58,significant differences(p<0.05),while the ΔCt value of UCK1 and UCK2 genes were similar(p>0.05);in relapsed/ refractory groups after treatment(7 day and 14 days),the ΔCt value of DCK gene was 5.370±0.707 and 6.097±0.513,of which post-treatment 7day and 14 days than per-treatment,but insignificantly decreased(p >0.05),the ΔCt value of UCK1 and UCK2 genes were similar(p>0.05);in positive control group post-treatment 7 day and 14 days,the ΔCt values of UCK1、 UCK2 and DCK genes were similar(p>0.05);3)Per-treatment expression of UCK1、UCK2 and DCK have no significant correlation with gender,age,FAB type,original cell,white blood cell count,hemoglobin,and platelet.5)Per-treatment expression levels of DCK gene has an effect on OS and PFS of AML,whose m6 A levlesup 4.71 have a longer time of OS and DFS.Conclusion: The expression of DCK in the AML patients of de nove and relapsed/refractory was increased;while the expression of UCK2 in both was decreased;post-treatment with decitabin the expression of DCK was reduced and close to normal after remission;the OS and DFS is shorter in the patients who has a highter expression level of DCK gene. |
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