| Background:Multiple myeloma (MM) is a neoplasm of a terminally differentiated B-cell,characterized by the slow proliferation of malignant plasma cells in the bone marrow. Much of our current understanding of the biology of MM has been obtained by studying MM-derived cell lines. Human myeloma cell lines were shown to be suitable model systems for use in various fields of the biological sciences. However,it is has proved very difficult to establish cell lines from plasma cell dyscrasias. There were only three MM cell lines have been established in our country,furthermore,these cell lines can not secret M proteins and have not been characted systematically;The origin of the MM progenitor cell has been extensively debated over the last decade .If the cell of origin has experienced antigen selection and somatic hypermutation,before neoplastic transformation,the ratio of amino acid replacement mutation (R)/amino acid silence mutation (S) in complementerity-determinng regions (CDRs) should be significantly greater than 2.9,the ratio of R:S in framework regions (FRS) should be significantly lower than 2.9. Each B cell clone has its unique variable region of immunoglobulin heavy chain gene,the origin of MM cell can be elucidated by studying the Vugene. Basing upon what mentioned above,we attemp to establish a MM cell line CZ-land analyse its biological characteristics,then analyse the VH genes of CZ-1 cells and peripheral b100d mononuclear cells of 98 MM patients in order to analyse the usage of VH gene famlies from the available repertoires of the MM cells,then elucidate the origin of the MM progenitor cell by analysing the charactertics of the mutation in VH..Objective:To establish a novel MM cell line CZ-1 and analyse its biological characteristics;to analyse the usage of VH gene families from the available repertoire of the MM cells and elucidate the origin of the MM progenitor cell byanalysis the charactertics of the gene mutation in VH.Methods:Mononuclear cells isolated from the peripheral b100d and bone marrow (BM) of a advanced MM patient (A light chain type) were incubated by liquid cell culture to establish CZ-1 cell line;the morphologies of the cells were determined by Wright-Giemsa-stainings and cytochemical stainings;the immunophenotypes of the cells were analyzed by flow cytometry;the cytogenetic analysis was performed by chromosome RHG-banding method;Quantitative fluorescent polymerase chain reaction was used to detected EBV DNA;The DNA of the CZ-1 cells and the peripheral b100d mononuclear cells of 98 MM patients was amplified by PCR technology using unique primers to variable region 1 to variable region 6 (Vnl-VH6),the purified PCR products were inserted into pMDIS-T vector,then transfected in JM109 bacteria,the expected DNA fragments were sequenced by dideoxy chain termination method,then compared with the correspondng germ line gene sequences. Results:CZ-1 cells can survive and proliferate not only when the bone marrow strumal cells or the human skin fibroblast cells were used as the feeder cells but also when the culture supernatants of the feeder cells were used as conditioned medium;CZ-l cells secret A light chain;CZ-1 cell is negative for EBV,Wright-Giemsa-stained CZ-1 cells showed typical plasma cell morphology with abundant cytoplasm and one to three nuclei;The cytochemical staining of CZ-1 cells showed the following reactivity pattern:positive for acid phosphatase and periodic acid Schiff-,negative for myeloperoxidase-,The immunoprofile of CZ-lcells corresponds to that typically in MM cells:positive for CD28x CD56 CD3S,CD138 CD49d CD 10,CD4 CD53 CD54,negative for CD5 CD19 CD34 CD40 CD40L CD95 CD95L;The sequences of the rearranged VH genes of the CZ-1 cells from both sources are identical;The cytogenetic analysis showed i (lq+),8q,13q,i (17q),i (18q),+M chromosome abnormality;The Vngene sequences of the CZ-1 cells from both sources are identical. The turn of the usage of VH genes from the available repertoirein multiple myeloma is VH3>VH1>VH4>VH2>VH5>VH6;The R/ S ratio in CDRs i...
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