| Objective: As one of the top ten cancers, lymphoma accounts for about a half of the blood system tumors diagnosed each year, and its incidence increases year by year. Lymphoma are divided into B-cell lymphoma and T-cell lymphoma according to the origin of cell and diffuse large B-cell lymphoma(DLBCL) is the most common type of B-cell lymphoma. Although the initial symptoms of lymphoma are various, the most common ones are painless progressive lymphadenectasis, local mass and the compression symptoms of the relevant organs. The highly invasive types of lymphoma progress rapidly, while combined chemotherapy helps only part of these patients to get sustained remission and has poor effect on most of them. Therefore,it’s quite necessary to pay attention to the etiology research, early diagnosis and targeted therapy of lymphoma. The further developments of tumour research have found that gene mutation as well as epigenetics mechanism might play important roles in the occurrence and development of tumour. As an important part of epigenetics, DNA methylation is becoming a hotspot of tumour research because of its characteristic of reversible. RASSF5( also called NORE1 or RAPL) which located at chromosome 1q32.1 is a member of Ras relevant family( RASSF1-RASSF10). RASSF5 can be divided into three different transcripts as RASSF5 A, RASSF5 B and RASSF5 C according to the difference of alternative splicing and promoter. In this research, the peripheral blood RASSF5 A gene promoter methylation status and m RNA expression levels in patients and normal people as well as the relationship between the test results and clinical data were discussed by the application of methylation-specific polymerase chain reaction(Methylation-Specific PCR, MSP) and reverse transcription- polymerase chain reaction(Reverse Transcriptase-PCR, RT-PCR), and which provided a theoretical basis for lymphoma pathogenesis, diagnosis and treatment.Methods: 1 Methylation-specific PCR(MSP) method was used to examine the methylation status of RASSF5 A gene in diffuse large B cell lymphoma, T cell lymphoma patients and healthy human periphral blood. The relationship between RASSF5 A gene methylation and clincal data was further analyzed. 2 Reverse Transcriptase-PCR(RT-PCR) method was used to detect the m RNA expression of RASSF5 A gene in diffuse large B cell lymphoma, T cell lymphoma patients and healthy human periphral blood. The relationship between RASSF5 A gene expression and clincal data was further analyzed. 3 SPSS19.0 was used to analyze the results of experiments.Results: 1 The expression and methylation status of RASSF5 A gene in peripheral blood of diffuse large B cell lymphoma patients 1.1 The methylation frequency of RASSF5 A in diffuse large B cell lymphoma patients peripheral blood(48/74, 64.9%) was significantly higher than that in healthy human periphral blood(3/42, 7.1%)(P<0.05). Methylation frequency of RASSF5 A gene was associated with LDH, IPI, and Ki-67(P<0.05). Methylation status of RASSF5 A gene was not associated with age, gender, bone marrow and extranodal involvment or not, A/B symptom, GCB/Non-GCB, clinical stage and β2 microglobulin(P>0.05). 1.2 The m RNA expression of RASSF5 A gene in diffuse large B cell lymphoma patients peripheral blood(0.54±0.17) was significantly lower than that in healthy human periphral blood(0.86±0.10)(P<0.05). The expression of RASSF5 A gene was correlated with IPI, extranodal involvment, however, it was not associated with age, gender, bone marrow or not, A/B symptom, GCB/Non-GCB, clinical stage, LDH, Ki-67, β2 microglobulin(P>0.05). 1.3 The m RNA expression of RASSF5 A gene in diffuse large B cell lymphoma with promoter methylation of the gene(0.51±0.18) was significantly lower than that in diffuse large B cell lymphoma with unmethylation of the gene(0.60±0.17)(P<0.05). 2 The expression and methylation status of RASSF5 A gene in peripheral blood of T cell lymphoma patients 2.1 The methylation frequency of RASSF5 A in T cell lymphoma patients peripheral blood(31/42, 73.8%) was significantly higher than that in healthy human periphral blood(3/42, 7.1%)(P<0.05). Methylation frequency of RASSF5 A gene was associated with clinical stage, extranodal involvment, Ki-67(P<0.05). Methylation status of RASSF5 A gene was not associated with age, gender, bone marrow and or not, A/B symptom, LDH, β2 microglobulin(P>0.05). 2.2 The m RNA expression of RASSF5 A gene in T cell lymphoma patients peripheral blood(0.52±0.18) was significantly lower than that in healthy human periphral blood(0.86±0.10)(P<0.05). The expression of RASSF5 A gene was correlated with clinical stage, extranodal involvment(P<0.05). however, it was not associated with age, gender, bone marrow or not, A/B symptom, LDH, Ki-67, β2 microglobulin(P>0.05). 2.3 The m RNA expression of RASSF5 A gene in T cell lymphoma with promoter methylation of the gene(0.50±0.15) was significantly lower than that in T cell lymphoma with unmethylation of the gene(0.63±0.12)(P<0.05).Conclusions: 1 The promoter methylation frequency of RASSF5 A in diffuse large B cell lymphoma and T cell lymphoma peripheral blood was significantly higher than that in healthy human, which suggests that promoter methylation of RASSF5 A gene may be associated with the occurrence of diffuse large B cell lymphoma and T cell lymphoma. 2 The expression of RASSF5 A in diffuse large B cell lymphoma and T cell lymphoma peripheral blood was lower than that in healthy human, which suggests that RASSF5 A gene may act as a tumor suppressor gene in diffuse large B cell lymphoma and T cell lymphoma. 3 The relatively low expression and the hypermethylation of RASSF5 A gene were detected in diffuse large B cell lymphoma and T cell lymphoma peripheral blood, which suggests that aberrant promoter hypermethylation of RASSF5 A gene may be one of the important mechanisms for its inactivation in diffuse large B cell lymphoma and T cell lymphoma. 4 The methylation status of RASSF5 A gene in diffuse large B cell lymphoma was correlated with IPI,LDH and Ki-67, the T cell lymphoma promoter methylation was correlated with clinical stage, extranodal involvment and Ki-67, which suggests that promoter methylation of RASSF5 A may be associated with tumor invasiveness, tumor malignant process and tumor prognosis. |
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