The second step of immunoglobulin gene alteration consists of somatic hypermutation and class switch recombination. 80th are regulated by activation-induced cytidine deaminase (AID). Methods: Study on possible application of class switch recombination assays for immunoglobulin gene alteration via AID. Cell based assays using AID B lymphocyte and NIH3T3 cell carrying switch substrate; gene transfer using retrovirus system; FACS analysis; PCR and ELISA. Results: DNA sequencing for S region and gamma1CT are the most sensitive and accurate assays. However, gamma1CT assay seemed to be more reliable and applicable. Others are accurate assays but less applicable. Conclusion: gamma1CT determination is the best class switch recombination assay for immunoglobulin gene alteration via AID.
Immunoglobulins, Genes

No abstract available.
Genes, Immunoglobulin Heavy Chain* ; Immunoglobulin Heavy Chains* ; Immunoglobulins*

Anti-idiotype antibody (anti-id Ab) which recognizes idiotope in the variable region of immunoglobulin (Ig) can regulate Ab production by B cells in vivo and in vitro. Although it has been reported that anti-id Ab can suppress IgM production by lymphocytes or hybridoma cells without suppression of cell proliferation, the regulatory mechanism of anti-id Ab is not completely understood. We studied the effects of anti-id Ab on the production of IgG class anti-DNA Ab by hybridoma cells, on the proliferation of cells, and on the transcription levels of Ig genes. In contrast to suppressive effect of anti-id Ab on the production of IgM previously reported by others, stimulatory effects of anti-id Ab on the production of IgG by hybridoma cells as well as the proliferation of these .cells were observed. However, little effect of anti-id Ab on the transcription levels of Ig genes was observed. These results suggest that anti-id Ab can increase Ab production by stimulation of cell proliferation. Furthermore, these results suggest that the effect of anti-id Ab on the production of Ab may be determined by the difference in class of Ab produced by hybridoma cells following the treatment with anti-id Ab.
Antibody Formation* ; B-Lymphocytes ; Cell Proliferation ; DNA* ; Genes, Immunoglobulin ; Hybridomas* ; Immunoglobulin G ; Immunoglobulin M ; Immunoglobulins ; Lymphocytes

No abstract available.
B-Lymphocytes* ; Bone Marrow* ; Genes, Immunoglobulin* ; Immunoglobulins* ; Lymphoma, Non-Hodgkin* ; Multiplex Polymerase Chain Reaction*

BACKGROUND: In the early stages of non-Hodgkin lymphoma (NHL), it can be difficult to recognize minimal morphological changes in the bone marrow (BM). In particular, when the quality of the BM biopsy is poor, determining BM involvement is limited to microscopic findings on BM aspiration. In this study, we compared the results of clonal immunoglobulin (IG) gene rearrangements with BM morphology results in B-cell NHL patients who underwent BM analysis as a staging workup and evaluated the usefulness of the clonal IG gene rearrangements for staging. METHODS: Forty two B-cell NHL patients were analyzed. Clonal rearrangements of the IG heavy chain (IGH), kappa light chain (IGK) and lambda light chain (IGL) genes were detected using the IdentiClone(TM) Clonality assay (InVivoScribe Technologies, USA). Clinical characteristics and outcomes were evaluated based on the detection of monoclonal IG gene rearrangements. RESULTS: Monoclonal IG gene rearrangements were found in 9 of 42 patients (21.4%). Microscopic BM involvement was found in only 2 of 42 patients (4.8%). The monoclonality rate of IG genes in BM was correlated with clinical stage and the international prognostic index (P<0.01). Patients with monoclonal IG gene rearrangements in BM had a significantly higher relapse rate (P=0.014) and poorer overall survival at 2 yr (P<0.01). CONCLUSIONS: Clonality analysis of BM in B-cell NHL can contribute to identification of patients with occult BM involvement with a significantly poorer overall survival despite normal BM histology.
B-Lymphocytes* ; Biopsy ; Bone Marrow* ; Gene Rearrangement ; Genes, Immunoglobulin* ; Humans ; Immunoglobulins ; Lymphoma, Non-Hodgkin* ; Recurrence

BACKGROUND: T cell immunoglobulin mucin containing molecule (TIM)-3 is expressed in differentiated Th1 cells and is involved in the suppression of the cytokine production by these cells. However, the regulation of the expression of other T cell genes by TIM-3 is unclear. Herein, we attempted to identify differentially expressed genes in cells abundantly expressing TIM-3 compared to cells with low expression of TIM-3. METHODS: TIM-3 overexpressing cell clones were established by transfection of Jurkat T cells with TIM-3 expression vector. For screening of differentially expressed genes, gene fishing technology based on reverse transcription-polymerase chain reaction (RT-PCR) using an annealing control primer system was used. The selected candidate genes were validated by semi quantitative and real-time RT-PCR. RESULTS: The transcription of TIMP-1, IFITM1, PAR3 and CCL1 was different between TIM-3 overexpressing cells and control cells. However, only CCL1 transcription was significantly different in cells transiently transfected with TIM3 expression vector compared with control cells. CCL1 transcription was increased in primary human CD4+ T cells abundantly expressing TIM-3 but not in cells with low expression of TIM-3. CONCLUSION: CCL1 was identified as a differentially transcribed gene in TIM-3-expressing CD4+ T cells.
Clone Cells ; Gene Expression ; Genes, vif ; Humans ; Immunoglobulins ; Mass Screening ; Mucins ; T-Lymphocytes ; Th1 Cells ; Tissue Inhibitor of Metalloproteinase-1 ; Transfection

OBJECTIVE: To gain insights into structural characteristics of immunoglobulin kappa chain repertoire expressed in the inflammed synovium of rheumatoid arthritis (RA), we analyzed V kappa transcirpts expressed in the synovium of a patient with longstanding RA and compared to those expressed in the PBLs of RA and normal controls. METHODS: RT-PCR was done to amplify kappa chain transcripts from RA synovial lymphocytes and the cDNA sequences were compared to those from PBL of RA patient or normal control. RESULTS: Kappa chain repertoire from RA patient's synovial lymphocytes or PBL revealed increased somatic mutation and unusually long complementarity determining region (CDR) 3 compared to normal control. CONCLUSIONS: These changes in kappa chain repertoire in RA patient are suggesting that the antibody repertoire expressed in the synovium or PBL is unique and may be related with systemic dysregulation of B cell development
Arthritis, Rheumatoid ; Autoimmune Diseases* ; Complementarity Determining Regions ; DNA, Complementary ; Genes, Immunoglobulin* ; Humans ; Immunoglobulin kappa-Chains ; Immunoglobulins* ; Lymphocytes ; Synovial Membrane*

PURPOSE: To evaluate whether the levels of immunoglobulin G (IgG) antibody to Tanerella forsythia are associated with periodontal status. METHODS: Patients with a diagnosis of chronic periodontitis were considered candidates for the study; thus 80 chronic periodontitis patients and 28 healthy persons (control group) were invited to participate in this investigation. The presence of T. forsythia was detected by polymerase chain reaction (PCR) analysis using primers designed to target the respective 16S rRNA gene sequences. Peripheral blood was collected from each subject to identify the IgG1 and IgG2 serum antibodies against T. forsythia. All microbiological and immunological laboratory processes were completed blindly, without awareness of the clinical status of the study patients or of the periodontal sites tested. RESULTS: The bivariate analysis showed that lower mean levels of clinical attachment level (CAL) and probing depth were found in the presence of the IgG1 antibody titers against whole-cell T. forsythia; however, only the difference in CAL was statistically significant. In the presence of the IgG2 antibody titers against whole-cell T. forsythia, the periodontal parameters evaluated were higher but they did not show statistical differences, except for plaque. The unadjusted linear regression model showed that the IgG1 antibody against whole-cell T. forsythia in periodontitis patients was associated with a lower mean CAL (beta=-0.654; 95% confidence interval [CI], -1.27 to -0.28; P<0.05). This statistically significant association remained after adjusting for possible confounders (beta=-0.655; 95% CI, -1.28 to -0.29; P<0.05). On the other hand, smoking was a statistically significant risk factor in the model (beta=0.704; 95% CI, 0.24 to 1.38; P<0.05). CONCLUSIONS: Significantly lower mean levels of CAL were shown in the presence of the IgG1 antibody titers against whole-cell T. forsythia in periodontitis patients. Thus, the results of this study suggest that IgG1 antibody to T. forsythia may have been a protective factor from periodontitis in this sample.
Antibodies ; Chronic Periodontitis ; Diagnosis ; Forsythia* ; Genes, rRNA ; Hand ; Humans ; Immunoglobulin G ; Immunoglobulins* ; Linear Models ; Periodontal Diseases ; Periodontitis ; Polymerase Chain Reaction ; Risk Factors ; Smoke ; Smoking

BACKGROUND: The clonality of lymphoid infiltrates determined by polymerase chain reaction (PCR) for immunoglobulin heavy chain (IgH) or T cell receptor (TCR) genes is not only useful in confirming the diagnosis of malignant lymphoma but also in establishing the lineage of a clonal lymphoid proliferation. We analyzed the efficiency of PCR analyses for IgH and TCRgenes that have been routinely applied for the diagnosis of malignant lymphoma in our laboratory. METHODS: Paraffin sections of 200 cases were analyzed by seminested PCR. Primers were FRIIIA-LJH/VLJH consensus primer for IgH gene and V-J consensus primer for TCR gene. The cases showing negative results by PCR for TCR gene were further analyzed by multiplex V family primers with heteroduplex analysis. RESULTS: PCR approach for IgH gene allowed detection of clonality in 100% of cases with false positive rate of 0.3% and false negative rate of 0%. The combination of PCR for TCR consensus primers with multiplex V family primers allowed detection of clonality in 91% of cases with false positive rate of 0.6% and false negative rate of 10.3%. CONCLUSIONS:Combined analysis of IgH and TCR gene rearragnements by the PCR technique followed by heteroduplex analysis can be a useful diagnostic adjunct to determine the clonality of various lymphoproliferative diseases with high sensitivity. But clinical, morphological and immunophenotypical correlation should be considered to reach the final diagnosis due to a few false positive cases.
Consensus ; Diagnosis ; Gene Rearrangement* ; Genes, T-Cell Receptor ; Heteroduplex Analysis ; Humans ; Immunoglobulin Heavy Chains* ; Immunoglobulins* ; Lymphoma ; Paraffin ; Polymerase Chain Reaction ; Receptors, Antigen, T-Cell* ; T-Lymphocytes*

BACKGROUND: Immunoglobulin (Ig) gene rearrangement analysis is a useful additional tool to detect clonality of B-lymphoproliferative disease and the method to detect immunoglobulin gene rearrangement is required the high sensitivity and specificity. BIOMED-2 multiplex PCR was designed for the evaluation of molecular clonality of lymphoid lesions. We evaluated the usefulness of the BIOMED-2 multiplex PCR by comparing it with conventional nested PCR. METHODS: Sixteen patients with malignant lymphoma and 5 with reactive lymph nodes were examined to assess the sensitivity, specificity, and accuracy between conventional nested PCR and BIOMED-2. All 3 tests performed using the BIOMED-2 kit for immunoglobulin (Ig) heavy chain (IGH) gene, Igkappa light chain (IGK) gene, and Iglambda light chain (IGL) gene, were used to analyze clonality. RESULTS: Both the methods showed 100% specificity (95% confidence interval, 56.6-100.0). The combination of IGH and IGK BIOMED-2 tests with or without IGL revealed the highest sensitivity (87.5%; range, 64.0-96.5%) and accuracy (90%; range, 0.70-0.97). Compared to the conventional method, the BIOMED-2 test for IGH showed a higher sensitivity (62.5%; range, 38.6-81.5%) and accuracy (71%, 0.50-0.86). CONCLUSIONS: These results suggest that, compared to the conventional method, BIOMED-2 has higher sensitivity and allows for easier interpretation while evaluating the clonality of B-lymphoproliferative disease.
Gene Rearrangement ; Genes, Immunoglobulin ; Hematologic Neoplasms ; Humans ; Immunoglobulins ; Light ; Lymph Nodes ; Lymphoma ; Multiplex Polymerase Chain Reaction ; Polymerase Chain Reaction ; Sensitivity and Specificity