engleski

Molecular insight into the diagnosis of lymphoma

Some malignant lymphoma may recapitulate benign architecture, on the other hand, some benign infiltrates appear malignant. This makes them very difficult to distinguish and diagnose with routine histopathological methods. Since a monoclonal proliferation is highly suggestive of neoplasia, the best method of diagnosis is to establish the clonal profile of the lymphocyte population. Immunostaining can be of great help, but can be very difficult to interpret, dependent on immunoglobulin production by lymphoma cells, as well as the techniques (antibodies) used. Demonstration of monoclonality in lymphocyte population can be achieved by DNA analysis using Southern blotting, which is a rather time consuming technique requiring high quality DNA and radioisotopes. PCR (polymerase chain reaction) is, on the other hand, extremely sensitive, highly specific method of detecting a target DNA sequence that can be present in a low copy number. During differentiation of B lymphocytes V, D, J and C segments of immunoglobulin heavy chain gene (IgH) are brought together in the specific and authentic mode in each B-cell allowing production of antibodies of extensive variability. Tumours, however, initially arise as single transformed cells; therefore the IgH gene in such cells are rearranged in the unique manner, and antibody made by such B-cell tumours are homogeneous. Using PCR primers complementary to relatively conserved regions called frameworks (FR1-FR3) laying among hyper variable regions (CDR1-CDR3) unable us to detect monoclonal versus polyclonal B-cell population. The length of the PCR product with these primers is unique if we deal with a monoclonal population. On the contrary, a polyclonal population gives PCR products of a different size. In this retrospective study we used semi-nested PCR to analyse 38 paraffin embedded specimens. All of them had been evaluated previously by pathohistological and immunophenotypic criteria at the Department of Pathology, Merkur Hospital, Zagreb. A number of polyclonal (PBL and tonsils from healthy donors) and monoclonal cells (PBL from CLL patients, Raji cell line) were analysed as controls. Clonality was successfully determined in all specimens. Our results support the concept that molecular techniques such as PCR provide a helpful approach for detection of monoclonal immunoglobulin rearrangements in malignant lymphoma. This is especially true for border cases, but always in the combination with other pathohistological methods.

malignant lymphoma; monoclonalitiy; PCR

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