| Objective: Lymphoma is malignancies of hematological. Lymphoma is divided into two categories including Hodgkin lymphoma and Non-Hodgkin lymphoma(NHL) according to the organization and pathology. NHL includes numerous subtypes. The prognosis of each subtype is different and there are several criteria for the evaluation of the prognosis of each subtype. At present, International prognostic index(IPI score) is commonly used to evaluate the prognosis of the non Hodgkin lymphoma, But has a limitation and still can not meet the clinical needs. New prognostic indicators need to be explored to guide the individualized treatment of NHL. Microsatellite(MS) is a highly polymorphic ? short and tandem repeat nucleotide sequence exists in prokaryotic and eukaryotic genomes. Microsatellite instability(MSI)refers to repetitive unit insertions or deletions caused by Changes in the length of the MSI and repeat sequences most in 1-6bp. MSI is a form of genetic instability th at includes n MSI(nucleus MSI)and mt MSI(mitochondria MSI). Recent investigations have showed that mt MSl also plays an important role in the occurrence and development. Our studies have found that 5892?8272?8280?8281?8289?9777 site of mitochondrial DNA non-D-loop have no significant correlation with the occurrence and prognosis of renal tumors except for the 8272 locus in our previous studies. There are significant correlation between the occurrence and prognosis of renal tumors. The Studies of the association of mt MSI with lymphoma have not been reported at home and abroad. This study aim to study the relationship between MSI which was caused by GGGGGTAGA9-bp insertions/deletions of mtDNA 8272 loci and clinical features and prognosis of NHL. We want to expore a possible prognostic indicator of NHL.Methods: 1 The sample: The cases were collected from the 139 cases of NHL patients who were diagnosed with immunohistochemical at Fourth Hospital of Hebei Medical University hematology department between January 2005 and January 2010, The patients' age were between 14-81 years old, median age was 49 years old,in which T cell lymphoma patients accounted for 69 cases(69/139 49.6%), mainly include ALK T-cells lymphoma, peripheral T-cells lymphoma(non specified type), T-lymphoblastoid cell lymphoma; and B-cell lymphoma(70 patients) accounted for(70/139 50.4%), the inert lymphoma of a total of 28 cases, including small lymphocytic lymphoma, follicular lymphoma and marginal lymphoma, invasion(diffuse large B-cell lymphoma and mantle cell lymphoma, B-lymphoblastoid cell lymphoma) were 42 cases. Peripheral venous blood samples were collected from the patients. Normal control subjects are peripheral venous blood samples of 98 healthy adult who were visited Fourth Hospital medical examination center of Hebei Medical University. 2 Study methods: 2.1 DNA extraction: The mtDNA of experimental group and normal control group were extracted by using the GENMED method and it will be placed in the refrigerator of- 80? and used as backup. 2.2 The polymerase chain reaction(PCR), denaturing polyacrylamide gel and sequenced:The target gene fragment was amplified by PCR and it was confirmed by gel electrophoresis. The PCR amplification product(amplified fragments are 200bp) was sent to the Shanghai Generay Biotech CO, Ltd. To obtain gene sequence of 6 sites(5892, 8272, 8280, 8281, 8289, 9777)in the mtDNA non-D-loop, and then compare with Normal human mitochondrial gene sequences, the purpose is to obtain ins/del cases and used as a next step analysis. we found that the association between mtDNA non-D-loop(5892, 8280, 8281, 8289, 9777)five loci insertion/deletion and prognosis of NHL had no correlation through Pre experiment. Therefore we study the association between mtDNA non-D-loop 8272 sites insertion/deletion and lymphoma prognostic. 2.3 Statistical analysis: Spss 19.0 software was used for statistics calculation. Probability of P < 0.05 was considered statistically signify. The survival curve was calculated using the Kaplan–Meier method. Multivariate survival analysis was performed using the Cox proportional hazards model. We analysis the rate different of the mtDNA8272 9-bp insertion / deletion in the different groups by using chi square test and Fisher's exact probability method.Results: 1 Comparison of the mtDNA8272 insertion/deletion rate between the experimental group and the control group there are 139 patients of NHL in the experimental group, among which 16 cases occurred mtDNA8272 GGGGGTAGA insertion/deletion. there are 98 cases patients in the control group, among which 5 cases occurred GGGGGTAGA deletion. P = 0.087, the difference was not statistically significant. 2 there are 9 cases(13.0%) occurred mtDNA 8272 GGGGGTAGA ins/del in T cell lymphoma and B cell lymphoma have 7 cases(10%). The occur rate was not statistically significant. 3 Mt DNA8272 GGGGGTAGA insertion / deletion was found 3 cases(7.9%) in aggressive B-cell lymphoma, and we found 4 cases(12.5%) in indolent B-cell lymphoma, P=0.403, the difference was not statistically significant(X2 test). 4 The relationship between whether occur mtDNA8272GGGGGTAGA9-bp insertion/deletion and the prognosis of NHL, the 5 year survival rate was 33.3%, 26.7%, P > 0.05, the difference was not statistically significant. 5 The relationship between the clinical features of T-cell lymphoma and B-cell lymphoma and mtDNA8272 GGGGGTAGA insertion/deletion phenomenon occurred rate(Fisher 's exact probability method). Patients were categorized according to the clinical characteristics(age, gender, IPI score, LDH, HBDH, stage, pathological type, ?2, whether bone marrow infiltration), compared whether occur mtDNA8272GGGGGTAGA insertion/deletion phenomenon and T-cell lymphoma and B-cell lymphoma, the difference was not statistically significant, P > 0.05. 6 The 5 year survival rate of T-cell lymphoma was 55.6% in mtDNA8272 GGGGGTAGA insertion / deletion and the 5 year survival rate was 16.4% in mtDNA8272 GGGGGTAGA sequence normal, P = 0.046, the difference is statistically significant. The 5 year survival rate of B-cell lymphoma was 42.9% in mtDNA8272 GGGGGTAGA insertion / deletion and the 5 year survival rate was 36.8%, P = 0.672, the difference was not statistically significant. 7 Multivariate analysis in the outcome of T-cell lymphoma mtDNA8272GGGGGTAGA9-bp insertion/deletions, IPI score, LDH, HBDH, staging, typing, ?2 microglobulin and whether bone marrow infiltration of the T-cell lymphoma patients be included in the Cox regression analysis, the aim is to study the influence of multiple factors on the survival time(5 years survival rate), we found that mtDNA 8272 GGGGGTAGA ins/del and LDH were statistically significant..Conclusions: 1 There was no correlation between the whether occurrence of mtDNA8272 GGGGGTAGA insertion / deletion and the risk of non-Hodgkin's lymphoma. 2 There was non correlation between mtDNA8272 GGGGGTAGA insertion / deletion and the subtypes of NHL and the clinical features of patients with different subtypes of lymphoma. 3 There was non correlation between 8272 GGGGGTAGA ins/del and the outcome of total non Hodgkin lymphoma lymphoma and B-cell lymphoma; It was considered correlation between 8272GGGGGTAGAins/del and T-cell lymphoma. 4 8272GGGGGTAGAins/del and LDH can be used as an independent prognostic marker for T-cell lymphoma. The cases of 8272 GGGGGTAGA ins/del and LDH is normal have a better prognosis. |
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