| Rearrangement of the immunoglobulin heavy chain (IgH) gene and T cell receptor (TCR) gene occurs separately in all normal developing B and T lymphocytes. Each normal lymphocyte undergoes a unique rearrangement, the differences among each cell resulting in a polyclonal in benign lymphoproliferative disease. As we know, in malignant tumor, all tumor cells originate from the monoclone .meanwhile , IgH gene and/or TCR gene monoclonal rearrangement can be shown in non-Hodgkin's lymphoma(NHL). This is the molecular basis to differentiate malignant lymphoma from benign lymphoproliferative disease. The application of polymerase chain reaction (PCR) for detection of gene rearrangement has been carried out widely in recent years. The consensus V and J primers PCR used to detect IgH are always designed directed to the relatively conservative amino acid sequences of the FR1, FR2, FR3, and J region. The primers with the highest detection rate and the most popular choice are directed against a region termed FR3 region of the various VH genes. FR3-directed primers FR3A detect approximately 60% of clonal B cell malignancies. FR3A PCR sometimes cannot amplify because of CDR region displaying somatic mutation. The addition of other framework regions will improve the detection rate of this test. The locus of the TCRy region is relatively simple, the rearrangement of TCRγ occurs at early stage of T cell maturation, and it can be detected in almost all the T cell lymphoma, so TCRγ become the favorite targets for detection of monoclonal T cells. In T-cell lymphoma, the detection rate of PCR is approximately 60% using primer directed TCRy, and the detection rate is approximately 40% with primer directed TCR#. Using combination of TCRγ and TCRβ primers will elevate detection rate to 70%-80%. Gene monoclonalrearrangement in malignant lymphoma sometimes may not be lineage specific. Approximately 10% mature B or T-cell lymphomas show dual rearrangements. Dual rearrangements and its relationship to malignant transformation remain to be elucidated. Both at home and abroad there are little papers reporting whether the biologic behaviors, such as proliferation, are different between dual rearrangements and correct rearrangement. The common methods of detection PCR products are agarose and polyacrylamide gel electrophoresis, which may somewhat induce to subjective interpretation of the results. Therefore, we try to use gel scan method so that we can make our interpretation based on a relative objective standard.PCR analysis of IgH and TCRγ, TCRβ gene rearrangement had been carried out in 125 cases of NHL to evaluate detection rate of clonality gene rearrangement and dual rearrangements with different primers in various types of NHL. At the same time we tried to find the most effective clonality rearrangement detection method, and to elucidate the significance of detection of clonality rearrangement of IgH gene and TCR gene in the NHL diagnosis. Immunohistochemistry was performed with ki-67 antibody in 117 case of NHL. We tried to find out whether or not the proliferation in dual rearrangements is different from that of correct rearrangement. 13 cases of benign lymphoid tissues (5 cases of reactive proliferative lymph node, 3 cases of tonsillitis, 5 cases of normal peripheral blood mononuclear cells) and 65 cases of IgH gene monoclonal rearrangement proved by FR3APCR and PAGE, were all analyzed by gel scan method. To find and prove the relatively objective standard of monoclonal rearrangement, the sensitivity of PCR detection of IgH FR3A and TCRγ was also analyzed.Materials and Methods:1. Patients: 125 cases of NHL were obtained from our affiliated hospitals and other local hospitals from the year of 1997 to 2002. All specimens were fixed in 10% formalin and then embedded in paraffin. Sections were stained by hemotoxylin and eosin and for immunohistochemistrical detection. All cases were divided into T- and B- NHL on the basis of their immunophenotypes and morphologies according to the WHO classification.2. |
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