Study On Derivation Of H/RS Cell And Its Clonal Relation With The Backgroun Lymphocytes In Classical Hodgkin's Lymphoma

Objective Hodgkin's lymphoma is characterized by its unique histomorphology: sporadic and isolated tumor cells are surrounded by reactive background cells hundreds of times more than them, which is different very much from the non Hodgkin's lymphoma. On this, it has been attracting a lot of medicine workers being engaged in studying it from almost all aspects of it for a hundred and sixty years. There were two focuses recently: (1) which cell lineage does the I-l/RS cell derive from after all? (2) What relation is there between the H/RS cell and its background cells? Being based on them, we creatively take usage of single-cell study on paraffin section, combined with lgH and TCR 13 gene arrangements detection and in situ detection of DNA after PCR, to spread out a study on clonality of background lymphocytes in cHL from a new direction, starting with detection of gene clonities of H/RS and background lymphocytes. That is which derivation does [1/RS cell come from and whether there is a same clone cell as the J-l/RS cell in the background? Design Every mature lymphocyte has its special variable zone after IgH gene or TCR f3 gene rearrangement, which means diversity, so as to a lymphocyte tumor should have a monoclonal Igll gene or TCR 13 gene rearrangement. According to this, we can check its rearrangement of IgIl or TCR gene to explore this suspicion. (1) Firstly, protein expressions of CD 15, CD3O, CD2O, CD45RO and EBV were detected in 32 cases of cHL to verify the diagnosis. (2) Following this, we detected the rearrangements of IgI-I gene and TCR 13 gene by PCR. (3) Then we synthesized Dig marked probes taking usage of PCR on the purified JgH PCR products, with which we proceeded in situ detectioii of DNA after PCR in two cases. (4) But it couldn抰 be trusted thoroughly except the disturbances were excluded. At last we selected a case for gene analysis of single-cell harvested by micromanipulation which had been stained by CD3O for position. Results (1) Of them, 11 cases were LrHL, 2 cases NSHL, 14 cases MCHL, 5 eases LDHL. (2) 25 /32(78.1%) were CDI5 positive, 30/32(93.8%) were CD3O positive, 6/32(18.8%) were CD2O positive and 7/32(21.9%) were CD45RO positive, 20/32(62.5%) were EBV positive. (3) Subsequently 10/32(31.3%) revealed single clonal bands of JgH gene and 3/32(9.1%)ofTCR 13 gene. (4) We established the method of in situ detection of DNA after PCR in paraffin section and found brown staining particles not only in H/RS cells? nucleuses but also in the surrounding background lymphocytes? (5) We built up the method of single-cell study in paraffin section, with which we acquainted monoclonal band and sequenced them successfully. Conclusion From above study, it can show initially the following conclusions: (1) The diagnosis and classification can be basically verified by all these expressed markers of Hodgkin抯 lymphoma. (2) Through the detection of e]onal rearrangement of JgH gene by PCR, we can draw a conclusion that partial cHL are derived from B cell, but can抰 exclude the possibility which is from the background lymphocytes. The detection of 3 cases clonal rearrangements of TCR 13 give a certain support to the T cell derivation. (3). According to in situ detection of DNA after PCR by using Dig marked lgH PCR products probes, it indicates that there is a homogeny between the partial background lymphocytes and H/RS cells. But it will be furtherly verified by designing the specific oligonucleotide probe to...