Epstein-Barr Virus Infection And Clonal Rearrangement And Molecular Characteristics Of Ig/TCR In NHL

Epstein-Barr virus is a y-1herpes virus, which related to a variety of tumors and other diseases, such as Hodgkin’s lymphomas (HL), Burkitt lymphoma (BL), immunodeficiency associated lymphoma, senile DLBCL and nasal type N/KT cell lymphoma, infectious mononucleosis, chronic active EBV infection and so on. According to the classification criteria of carcinogenic factor from National Cancer Institute, EBV was listed in the first group of carcinogenic factor. However, the potential pathogenic mechanism is extremely complicated; EBV may cause abnormalities of the host cell directly and indirectly through integrating its own genome into the host cell genome or expressing various gene products.Ig/TCR gene rearrangement is a normal physiological process of mature lymphocytes. Ig/TCR gene is composed of4parts of genes, including variable area (V), high change area (D), combining area (J) and constant area (C). V, D and J gene conducted a completely rearrangement during the development of B lymphocytes and thymus T cells and formatted a functional Ig/TCR gene, known as the gene rearrangement. Somatic hypermutant (SHM) was happened in germinal center (GC) so mutational analysis of Ig gene may help to determine the source of the B cell lymphoma. Polymorphisms of CDR3determined the specificity of TCR gene. Biased VH gene usage and selective expression of TCR gene families suggested that some tumor-associated specific antigens may play a role in the pathgenesis and development of lymphomas. As a lymphoma-associated virus, it is significant to find the relationship between EBV infection and molecular characteristics of Ig/TCR gene in NHL.Using nested PCR technology and electrophoretic grosal gel, Ig gene rearrangement of50NHL cases (mainly including DLBCL, FL, MALT and CLL) were performed. Ig gene of49cases (98.0%) were clonal rearrangement with FR3A primers, and with FR2A primers, Ig gene of39cases (78.0%) were clonal rearrangement, and combining with FR3A and FR2A primers, Ig gene of49cases were clonal rearrangement. In9cases of T-NHL, tested with the TCRyl and TCRy2primers, both positive percent was100%. Meanwhile, we used EBER-ISH technique to detect EBV infection in39cases of B-NHL,9cases were EBV infected, all of9cases were DLBCL. In addition,7cases in total9cases of NK/T-NHL were EBV infected with77.8%positive rate. We used fluorescent to mark primer for PCR based Genescans. With FR3A/LJH/VLJH as primers,15cases (43.8%) in31B-NHL were monoclonal rearrangements,6cases (19.4%) were biclonal rearrangements,5cases (16.1%) were oligoclonal rearrangements and5cases (16.1%) were multiple clonal rearrangements. With FR2A/LJH/VLJH as primers,21cases (70.0%) in30B-NHL were monoclonal rearrangements,2cases (6.6%) were biclonal rearrangements,3cases (10.0%) were oligoclonal rearrangements, and4cases (13.3%) were multiple clone rearrangements. With TCRyl primer,2cases in6T-NHL was monoclonal,1case was biclonal rearrangement and3cases were polyclonal. With TCRy2primer,1case in6T-NHL was monoclonal,1case was oligoclonal, and4cases were polyclonal.17samples of NHL were sequenced and Ig/TCR BLAST analysised,15cases carried mutated Ig gene with point mutations and nucleotide substitutions. Biased VH family usages in NHL were found,8of17VH usages belonged to VH4family and8of 17VH usages belonged to VH3family. Among VH4family, VH4-34gene segment were used4times (25.0%) in16samples, and VH4-61gene were used3times (18.8%), which was comfirmed the results of our previous study that biased VH family usages in DLBCL with VH4-34(4/17) and VH4-61segements (4/17)In conclusion, EBV infection rate in NHL lymphoma samples of our experiment group was42.0%. Positive infection rate was27.3%among33DLBCL samples, and8of9EBV infected DLBCL patients were older than50years old. The size and quantity of the DNA fragments could be accurately detected by PCR-GeneScan, but the diagnosis of lymphoma still need combining with morphological, immunohistochemical and clinical manifestations. The mutation status of IgVH gene is helpful to determine the source of B cell lymphoma, only2patients with DLBCL in this group carried unmutated IgVH genes, and the rest patients carried mutated IgVH genes, meaning germinal center (GC) or post GC sources. Biased gene family usage was found:VH4-34gene segment were used4times (25.0%) in16samples, and VH4-61gene were used3times (18.8%), which suggests that the pathogenesis of MALT lymphoma and DLBCL may be associated with certain antigen, but the correlation of EB virus still needs more samples.