Analysis of antigen receptor genes in Hodgkin's disease.

AIM--To analyse the configuration of the antigen receptor genes in Hodgkin's disease. METHODS--DNA extracted from 45 samples of Hodgkin's disease was analysed using Southern blotting and DNA hybridisation, using probes to the joining region of the immunoglobulin heavy chain gene, the constant region of kappa immunoglobulin light chain gene, and the constant region of the beta chain of the T cell receptor gene. RESULTS--A single case of nodular sclerosing disease showed clonal rearrangement of the immunoglobulin heavy and light chain genes, all other samples having germline immunoglobulin genes. The nature of the clonal population in the diseased tissue is uncertain, because the intensity of the rearranged bands did not correlate with the percentage of Reed-Sternberg cells present. The T cell receptor genes were in germline configuration in all the samples. CONCLUSIONS--Antigen receptor gene rearrangement is a rare finding in unselected cases of Hodgkin's disease.     

Clonal gene rearrangement patterns correlate with immunophenotype and clinical parameters in patients with angioimmunoblastic lymphadenopathy.

T cell receptor beta (TcR beta) chain gene rearrangements have been reported in cases of angioimmunoblastic lymphadenopathy (AILD) and provided evidence for the presence of clonal T cell proliferations in this disorder. Twenty-three cases of AILD and two cases of hyperimmune reaction (HR) were investigated. In the two HR cases, essentially the same histologic pattern was present as in AILD but lymph node follicles were hyperplastic. Both HR cases showed germline configuration for the TcR and immunoglobulin heavy chain (IgH) genes. All other patients diagnosed with AILD had clonal rearrangements for TcR gamma and beta chain genes. In addition, seven out of these cases had clonally rearranged their IgH genes. These two different rearrangement patterns (TcR with or without Ig gene rearrangement) correlated to immunohistochemical and clinical data. Cases with TcR but without Ig gene rearrangements (group I) exclusively showed CD4+ proliferating T cells, whereas those cases with TcR and Ig gene rearrangements had significantly elevated numbers of CD8+ proliferating cells (group II). Group II patients significantly more often presented with hemolytic anemia and went into transient remission spontaneously or under steroid treatment. Group I patients, however, had a higher response to chemotherapy and a longer survival time. These data show that, based on different rearrangement patterns, it is possible to divide AILD into two different groups with distinct immunophenotypic properties and differences in clinical parameters. Immunogenotyping in AILD thus will have prognostic and therapeutic implications.     

Evaluation of biotinylated DNA probes for the detection of gene rearrangements in clinical specimens

To determine whether a nonisotopic procedure is suitable for analyzing clinical specimens for gene rearrangements, the authors hybridized DNA from 15 specimens of lymphoid tissue with biotinylated DNA probes directed to J beta I + J beta II (T-cell receptor beta chain gene), JH (immunoglobulin gene heavy chain J region), and J kappa (immunoglobulin gene kappa light chain J region). Five cases of benign lymphoid hyperplasia, one case of dermatopathic lymphadenopathy, and one case of small noncleaved follicular center cell lymphoma had germline hybridization patterns when digested with Bam HI, Eco RI, and Hind III restriction endonucleases. Four cases of B-cell lymphoma and three cases of T-cell lymphoma had clearly detectable rearrangements of the genes for immunoglobulin or the T-cell receptor or both. One case of dermatopathic lymphadenopathy had a faint, clonal rearrangement of the T-cell receptor after digestion with Eco RI and Bam III. The authors conclude that biotinylated DNA probes can be useful for analyzing gene rearrangements in clinical specimens.     

Clonality of T cell and phenotypically undefined lymphoid neoplasms: the value of genotypic analyses.

The value of genotypic analysis for routine assessment of leukaemia and lymphoma was shown by the findings in a selected series of 30 cases. T cell receptor (TcR) gene rearrangements were observed in six out of nine cases of CD3+ CD8+ lymphocytoses and provided clear evidence for clonality in this group. The T cell proliferations in two of the remaining cases masked B cell lymphocytic leukaemia and hairy cell leukaemia, while in the third case no cause was found for the polyclonal proliferation. Heterogeneity of phenotype and genotype were observed in peripheral T cell lymphomas: one out of six cases showed TcR gene rearrangement, one case retained its germline configuration, a further case masked B cell lymphoma and the remainder were polyclonal. Genotypic analysis was helpful in the analysis of a tumour of mixed T cell and myeloid phenotype which was shown to be germline for TcR and immunoglobulin genes, consistent with a myeloid origin. Two histiocytic tumours were found to have clonal rearrangement of TcR genes. Nine out of 11 B cell tumours showed immunoglobulin gene rearrangement. It is concluded that genetic analyses are useful in the analysis of T cell, histiocytic, and B cell tumours in which an immunoglobulin phenotype cannot be defined.     

Immunoglobulin gene rearrangements in Hodgkin's disease

An initial survey of biopsy specimens from 16 cases of Hodgkin's disease revealed clonal immunoglobulin gene rearrangements in one specimen, which contained large numbers of Reed-Sternberg (R-S) cells. As a result of this finding, the configuration of immunoglobulin and T-cell receptor gene DNA was investigated in biopsy tissues from other cases that were histologically and immunophenotypically consistent with Hodgkin's disease and contained numerous R-S cells. In six of seven such specimens (all of the nodular sclerosing subtype), selected solely on the basis of high R-S cell content and sufficient frozen tissue for study, at least one immunoglobulin gene was found to be rearranged in a clonal manner. Additionally, tissue samples obtained at two different time points from the original patient who showed immunoglobulin gene rearrangements revealed identical patterns of rearrangement. In the majority of cases, only a single gene showed rearrangement, and the rearranged bands in Southern blot autoradiograms were usually considerably less intense than the germline bands. No rearrangements of T-cell receptor DNA were detected in any case with a probe for the beta T-cell receptor gene. The results suggest that clonal cell populations possessing uniform immunoglobulin gene rearrangements are present in tissue in some cases of Hodgkin's disease. It is not possible to determine which cells contain these rearranged genes, but the increased incidence of detectable rearrangements in cases with high numbers of R-S cell raises the possibility that immunoglobulin gene rearrangement occurs in these cells.     

Regressing atypical histiocytosis: a review and critical appraisal

Regressing atypical histiocytosis (RAH) has been defined as a primary cutaneous neoplasm composed of atypical histiocytes. In this study, ten cases of RAH were available for review including the two first reported cases. In addition, one new case was studied immunocytologically, for immunoglobulin and T cell receptor gene rearrangement, and for DNA ploidy analysis. Histologic study of ten cases permitted recognition of microscopic features both common and uncommon to RAH. Clinical follow-up of eight cases suggests an indolent course but with probable substantial long-term risk for development of systemic lymphoma. The histiocytic origin of RAH must now be considered questionable because the results of immunologic phenotyping and the discovery of rearrangement of T cell receptor beta- and gamma-chain genes found in the newly studied case indicate that this primary cutaneous neoplasm, previously considered histiocytic, is most probably of T cell lineage.     

Study of the immunohistochemistry and T cell clonality of enteropathy-associated T cell lymphoma.

Specimens from 23 patients with enteropathy-associated T cell lymphoma were studied by immunohistochemistry after antigen retrieval. Specimens from 14 of these patients were investigated for the presence of clonal T cell gene rearrangements in both the tumor and the adjacent enteropathic intestine by the polymerase chain reaction. Primers for T cell receptor beta and gamma genes were used in a combination that permits the identification of approximately 90% of T cell receptor rearrangements. Clonal rearrangements of the T cell receptor were found in 13 of the 14 tumors studied. Specimens of enteropathic bowel resected with the tumor, but showing no morphological or immunohistochemical evidence of tumor involvement, showed clonal T cell receptor gene rearrangements in 11 cases. In 10 of these, the amplified DNA was of the same molecular weight in the enteropathic bowel as in the corresponding tumor. In 2 cases, sequencing the polymerase chain reaction product showed identical T cell receptor gene rearrangements in the tumor and in the adjacent intestine. Uniform staining for p53 was seen in 22 of the 23 tumors. In 9 of 19 cases studied, collections of small lymphocytes in the enteropathic bowel expressed p53. In all but one of these specimens, a clonal rearrangement of the T cell receptor genes was identified. We interpret these findings as support for the concept that enteropathy-associated T cell lymphoma arises on a background of gluten-sensitive enteropathy with evolution of neoplastic T cell clones from the reactive T cell population present in the enteropathic bowel.     

Most null large cell lymphomas are B lineage neoplasms

DNA of immunoglobulin and the beta T cell receptor genes was analyzed for rearrangements in 34 diffuse large cell lymphomas that failed to express immunoglobulins or T cell antigens. Twenty-eight cases had both heavy and light chain immunoglobulin rearrangements, two cases had only heavy chain gene rearrangements, three cases had only light chain gene rearrangements, and one case failed to show rearrangements for any of the immunoglobulin genes. None of the cases showed rearrangements for the beta T cell receptor gene. These results indicate that the vast majority of diffuse large cell lymphomas that lack definitive B or T cell phenotypic markers are actually B cell in origin.     

急性T淋巴细胞白血病中Tal~d重排的结合部顺序研究

应用多聚酶链反应和ＤＮＡ测序技术，对２８例急性Ｔ急性淋巴细胞白血病（Ｔ－ＡＬＬ）患者进行ＳＩＬ－ＴＡＬ－１重排的检测和重排结合部的顺序分析。结果发现，５例有ＳＩＬ－ＴＡＬ－１融合基因，对其中４例的ＤＮＡ重排结合部分析表明均为Ｔａｌｄ１信号所介导、且存在着Ｎ顺序，证实Ｔａｌｄ１的重排是由免疫球蛋白／Ｔ细胞受体基因Ｖ－（Ｄ）－Ｊ重组酶系的错误所致，在Ｔ－ＡＬＬ的发病中具有重要的意义。     

Nasal T-cell lymphoma associated with hemophagocytic syndrome. Immunohistochemical and genotypic studies.

A 53-year-old man with nasal T-cell lymphoma exhibited hemophagocytic syndrome as a terminal event. Immunohistochemical studies revealed that neoplastic cells were derived from T cells. Genotypic analysis of DNA samples that were obtained from the frozen tissue specimens demonstrated clonal rearrangements of the T-cell receptor beta-chain genes. No rearrangement was observed in the immunoglobulin heavy-chain gene. To our knowledge, only three cases of nasal T-cell lymphoma with hemophagocytic syndrome have been reported before the present case. All of these cases occurred in Oriental patients. The present report suggests the beneficial effect of high-dose glucocorticoid therapy on the prolongation of survival compared with that of the other three cases.     

T-cell lymphomas in adults: a clinicopathological study of eighteen cases

Eighteen cases of adult T-cell lymphoma have been studied with respect to clinical presentation, response to treatment, histology, enzyme histochemistry, immunocytochemistry, and gene rearrangement. Seven cases (39 per cent) presented at extra-nodal sites, and the age range was from 18 to 79. Treatment was with combination chemotherapy in most cases, and 11 of the 18 patients died within the three year follow-up period. The lymphomas were classified morphologically into six types; T-lymphoblastic lymphoma (TLL), angioimmunoblastic lymphadenopathy (AIL)-like T-cell lymphoma, T-zone lymphoma, pleomorphic mixed medium and large cell T-cell lymphoma (PMMLC), pleomorphic large cell T-cell lymphoma (PLC), and monomorphic large cell T-cell lymphoma (MLC). Enzyme histochemistry was found to be of limited value in the identification of T-cell lymphomas. Immunocytochemistry showed a degree of correlation between the immunological profile and morphology, with cases in the PLC and MLC groups showing limited expression of T-cell antigens. Re-arrangement of the beta chain of the T-cell receptor gene was detected in 12 of the 14 cases studied, and all showed germ-line immunoglobulin genes. The study emphasizes the varied morphological and clinical appearances of adult T-cell lymphoma.     

Rearrangement of immunoglobulin and T-cell receptor genes in Hodgkin''s disease.

The precise cellular origin of the malignant cell population in Hodgkin's disease (HD) is unknown. Recent application of Southern blotting techniques to detect clonal rearrangements of immunoglobulin (Ig) and T-cell receptor (TCR) genes has yielded conflicting results. The authors report the detailed analysis of tumor tissue DNA obtained from 18 cases of HD using Ig and TCR gene probes. The distribution of HD subtypes was similar to that in other series. Samples were examined for rearrangement by means of multiple restriction enzymes with specific probes for the Ig heavy chain, Ig kappa, Ig lambda, TCR beta, and TCR gamma loci. Only germline bands were detected in all 18 cases with the Ig gene probes and in 15 of 18 cases with the TCR probes. In 2 cases blot analysis suggested a predominance of polyclonal (or oligoclonal) T cells. In 1 case monoclonal rearrangement of the TCR beta gene was detected. Based on the intensity of the rearrangement and the small percentage of Reed-Sternberg (R-S) cells in this case, the clonal population detected was most likely not the R-S cell itself. The data do not support the frequent occurrence of Ig or TCR monoclonal gene rearrangement in HD.     

Polyclonal rearrangement of the T-cell antigen receptor genes in Hodgkin's disease: implications for diagnosis.

Distinction between Hodgkin's disease and peripheral T-cell lymphoma can be difficult, based on routine morphologic and immunohistologic stains. To assess the utility of Southern blot analysis, we investigated the genes for the beta (T beta) and gamma (T gamma) chains of the T-cell antigen receptor and immunoglobin heavy and light chains in 15 unselected cases of Hodgkin's disease. Following digestion with the restriction enzyme EcoRI, the intensity of the germline band corresponding to the first constant region of T beta was reduced when compared with the intensity of the second constant region germline band, a pattern consistent with polyclonal rearrangement of this gene. Hybridization of EcoRI-digested DNA with a probe recognizing the joining regions of T gamma revealed multiple rearranged bands corresponding to the known limited recombinatorial possibilities of this gene. Clonal rearrangement of T beta and T gamma was never demonstrated, although the banding pattern seen with T gamma could easily be misinterpreted as such. The findings in Hodgkin's disease were identical to those obtained in hyperplastic lymph nodes, normal thymuses, and normal peripheral blood enriched for T-cells. We then examined the status of T-cell receptor and immunoglobulin genes in six cases that could not be definitely classified as either Hodgkin's disease or T-cell lymphoma by either morphology or immunohistology. Clonal T-cell receptor gene rearrangement was found in three cases, supporting the diagnosis of T-cell lymphoma. Our study confirms the polyclonal lineage of the major fraction of T-cells and B-cells in Hodgkin's disease, a finding that may have diagnostic importance.     

Frequent immunoglobulin and T-cell receptor gene rearrangements in "histiocytic" neoplasms

The authors have analyzed the DNA of immunoglobulin and T-cell receptor genes in a series of 6 malignancies which were judged to be of histiocytic derivation on the basis of morphologic criteria. They found that 4 of these cases showed rearrangements of the beta T-cell receptor genes in spite of the lack of any specific immunohistochemical markers for B or T cells. One case showed rearrangements of both heavy and light chain immunoglobulin genes and probably represents either a sinusoidal large cell lymphoma or a B-cell lymphoma with activation of histiocytes simulating malignant histiocytosis. A single case lacked both immunoglobulin and T-cell receptor rearrangements consistent with immunologic analyses that suggested its origin from an interdigitating reticulum cell. The result of this study in conjunction with the authors' previous immunologic observations suggests that many presumed histiocytic malignancies actually represent T-cell lymphomas. Alternatively, beta T-cell receptor rearrangement may be a common feature of tumors that show monocyte/histiocytic differentiation.     

Rearrangements of T cell receptor loci can be found only rarely in B lymphoid cells

We have studied the rearrangement status of the T cell receptor genes in 64 B lymphoid cell lines, and we found that, unlike the immunoglobulin heavy chain genes in T lymphocytes, T cell receptor beta and related gamma chain genes are almost always in germ-line configuration in B lymphoid cells. The only exception was a myeloma MOPC511 (IgA, chi) which contained all T cell receptor genes, beta 1, beta 2, gamma 1, gamma 2, gamma 3 and alpha, in rearranged configuration in both homologous chromosomes. This exception supports the concept that all immunoglobulin and T cell receptor genes exploit the same recombinase to build their complete variable regions. Obviously, in MOPC511 cells the regulation, which confers the tissue specificity i.e. T vs. B lymphocytes, has failed.     

Sensitivity of PCR in detecting monoclonal B cell proliferations.

AIMS--To evaluate the rapid detection of various forms of monoclonal B cell proliferations by using the polymerase chain reaction (PCR) to identify clonal immunoglobulin heavy chain genomic rearrangements. METHODS--Thirty four B cell lymphomas defined by morphology, immunophenotyping, and positive immunoglobulin heavy chain gene rearrangements detected by Southern blot analysis were examined. An additional 22 cases representing miscellaneous lymphoproliferative and non-lymphoproliferative disorders were also studied. RESULTS--Monoclonal rearrangements were identified in 19 (56%) cases of B cell lymphoma. The method was less sensitive in the detection of follicular centre cell lymphomas (15 of 28, or 54%) than non-follicular centre cell lesions (four of six, or 67%). Monoclonal rearrangement was not identified in 19 control cases, including T cell lymphomas, Hodgkin's disease, reactive lymphadenopathy and metastatic carcinoma. Three cases showed positive immunoglobulin gene rearrangement by PCR but were negative on Southern blotting. Two of these cases had definite clinical, morphological, and immunophenotypic evidence of monoclonal B cell proliferation suggesting that PCR could, on occasion, pick up cases missed by Southern blotting and that the two methods are complementary in clonal lymphoproliferative disease diagnosis. The third case represented a "false positive" PCR reaction involving a colonic adenocarcinoma. CONCLUSIONS--PCR analysis, using the primer sequences outlined in this study, will detect about 55% of clonal lymphoproliferative proliferations with increased sensitivity for non-follicular centre cell lesions. With these levels of detection in mind, this testing strategy can still be especially useful in cases which prove diagnostically problematic with standard morphological and immunophenotypic analysis, and in instances where the quantity and type of diagnostic material is limiting (needle aspirates and cellular fluids).     

Acute lymphoblastic leukaemia: correlation between morphological/immunohistochemical and molecular biological findings in bone marrow biopsy specimens.

BACKGROUND: Although numerous antibodies suitable for use on paraffin wax embedded sections are available for the subtyping of acute leukaemia (acute myelogenous leukaemia (AML) and acute lymphoblastic leukaemia (ALL)) in bone marrow biopsy sections, unequivocal identification of the cell line involved is sometimes impossible. METHODS: Forty eight formalin fixed, paraffin wax embedded bone marrow biopsy specimens that had been decalcified in EDTA were investigated, including 42 thought to exhibit ALL on the basis of bone marrow smears. Five specimens exhibited AML and one biphenotypic leukaemia, as diagnosed immunohistochemically in bone marrow biopsies. Immunostaining was performed with antibodies against relatively specific B and T cell antigens. The blasts were investigated for rearrangements of the immunoglobulin heavy chain (IgH) and the T cell antigen receptor (TCR) genes. RESULTS: Amplifiable DNA was obtained from all 48 specimens. An IgH gene rearrangement was detected in 20 of 23 c-ALL specimens. Four of seven T cell ALL (T-ALL) specimens had a TCR-gamma gene rearrangement, and the one B cell ALL (B-ALL) specimen exhibited a clonal IgH gene. Three of four cases of unclassifiable ALL could be assigned to the B cell lineage on the basis of gene rearrangement analysis. Seven cases originally diagnosed in smears as ALL were rediagnosed as AML (n = 5) or biphenotypic leukaemia (n = 2) because of immunohistochemical reactivity for myeloperoxidase or lysozyme. Two of these AML cases and two of three cases of biphenotypic leukaemia exhibited a monoclonal IgH gene rearrangement. CONCLUSIONS: Acute leukaemia can be subtyped in bone marrow sections with a limited panel of antibodies suitable for use on paraffin wax embedded sections (against CD3, CD10, CD20, CD79a, myeloperoxidase, and lysozyme). In patients with ALL and a diagnostically equivocal immunophenotype, gene rearrangement analysis might indicate whether the B or T cell lineage is involved.     

Genotypic analysis in large cell lymphomas expressing a restricted set of differentiation antigens

Immunophenotyping and immunogenotyping were performed in a series of 8 large cell lymphomas exhibiting anaplastic or "histiocytic" morphology and displaying an uncertain phenotype due to a restricted number of differentiation antigens. 6 cases expressed the Ki-1 antigen. 4 cases expressed one or two B-cell markers and contained rearrangements of the immunoglobulin genes. One of them also exhibited a T-cell receptor (TCR) beta gene rearrangement. 3 cases expressed a single T-cell differentiation antigen. Among them, only 1 displayed both gamma and beta TCR gene rearrangement; 1 only contained a gamma TCR gene rearrangement and 1 completely lacked clonal rearrangements. The eight cases expressed an inconclusive immunophenotype due to an abundant population of reactive cells but showed an immunoglobulin gene rearrangement. In conclusion, 5 out of the 8 unusual lymphomas studied here could be characterized by immunogenotyping. This approach was, however, inconclusive in the 3 remaining cases, whose lineage and differentiation stage remain poorly defined.     

血液系统肿瘤患者外周血细胞IgH基因和TCRγ基因的检测及意义

检测各种血液系统肿瘤患者外周血细胞免疫球蛋白重链基因 (IgH )和T细胞受体γ基因 (TCRγ )克隆性重排并探讨其意义。通过多聚酶链式反应 (PCR )方法检测 32例非霍奇金淋巴瘤 (NHL )、 18例急性髓性白血病 (AML )、 2 4例多发性骨髓瘤 (MM )、 8例急性淋巴细胞白血病 (ALL )及 6例慢性淋巴细胞白血病 (CLL )患者外周血细胞IgH及TCRγ克隆性基因重排。结果表明 ,NHL、AML、MM、ALL及CLL患者中IgH克隆性重排率分别为 37 5 0 %、 2 2 2 2 %、 83 33%、 12 5 0 %和 16 6 7% ;TCRγ基因克隆性重排率分别为 6 2 5 0 %、 5 0 0 0 %、 5 4 17%、 5 0 0 0 %及 5 0 0 0 %。在B型、T型NHL中 ,IgH克隆性重排率分别为 31 5 8%及 6 6 6 7% ;TCRγ克隆性重排率分别为 47 37%及 6 6 6 7%。AML中IgH克隆性重排阳性者的初治完全缓解率(CR ) (5 0 0 0 % )与IgH重排阴性的初治CR率 (5 0 0 0 % )无显著差异 (P >0 0 5 )。TCRγ克隆性重排阳性者与阴性者的初治CR率 (均为 44 44 % )亦无显著差异 (P >0 0 5 )。IgH及TCRγ基因克隆性重排不具有细胞谱系的特异性 ,但通过检测外周血IgH、TCRγ克隆性基因重排对NHL有辅助诊断意义 ,并且可作为监测微小残留病壮 (MRD )的手段。