Biomedical subjects

A Bräuninger

Publications and source records attributed to A Bräuninger.

At least 19 recordsLinked to original sources

PubMed2005

Insights into the multistep transformation process of lymphomas: IgH-associated translocations and tumor suppressor gene mutations in clonally related composite Hodgkin's and non-Hodgkin's lymphomas.

Clonally related composite lymphomas of Hodgkin's lymphoma (HL) and Non-Hodgkin's lymphoma (NHL) represent models to study the multistep transformation process in tumorigenesis and the development of two distinct tumors from a shared precursor. We analyzed six such lymphomas for transforming events. The HLs were combined in two cases with follicular lymphoma (FL), and in one case each with B-cell chronic lymphocytic leukemia, splenic marginal zone lymphoma, mantle cell lymphoma (MCL) and diffuse large B-cell lymphoma (DLBCL). In the HL/FL and HL/MCL combinations, BCL2/IGH and CCND1/IGH translocations, respectively, were detected in both the HL and NHL. No mutations were found in the tumor suppressor genes FAS, NFKBIA and ATM. The HL/DLBCL case harbored clonal replacement mutations of the TP53 gene on both alleles exclusively in the DLBCL. In conclusion, we present the first examples of molecularly verified IgH-associated translocations in HL, which also show that BCL2/IGH or CCND1/IGH translocations can represent early steps in the pathogenesis of composite HL/FL or HL/MCL. The restriction of the TP53 mutations to the DLBCL in the HL/DLBCL case exemplifies a late transforming event that presumably happened in the germinal center and affected the fate of a common lymphoma precursor cell towards development of a DLBCL.

Cell Transformation, Neoplastic↗

PubMed2005

Activation induced cytidine deaminase expression in lymphocyte predominant Hodgkin lymphoma.

BACKGROUND: The lymphocytic and histiocytic (L&H) cells of lymphocyte predominant Hodgkin lymphoma (HL) originate from germinal centre B cells and carry mutated V gene rearrangements, usually with intraclonal diversity. It is unclear whether intraclonal V gene diversification by somatic hypermutation, which is strictly dependent on the enzyme activation induced cytidine deaminase (AID), is restricted to the early phase of lymphoma clone expansion and later silenced, or whether it remains active throughout malignant proliferation. AIMS: To analyse whether AID is expressed in L&H cells as an indicator of active somatic hypermutation in the tumour cells. METHODS: L&H cells from lymphocyte predominant HL cases and centroblasts from lymphadenites were micromanipulated and analysed for AID expression by quantitative real time polymerase chain reaction. RESULTS: The AID transcription level was higher than background in three of the six lymphocyte predominant HL cases, although it was lower than that seen in centroblasts. CONCLUSIONS: Somatic hypermutation may remain active in L&H cells in a considerable proportion of cases, increasing the risk of acquiring further transforming mutations.

Cytidine Deaminase↗

PubMed2002

Biology of Hodgkin's lymphoma.

Significant progress has been made in recent years in our understanding of the cellular origin of Hodgkin and Reed-Sternberg (HRS) cells in Hodgkin's lymphoma (HL). It is now clear that in most instances HRS cells represent clonal populations of transformed germinal centre (GC) B cells. While the tumour cells in the lymphocyte predominant type of the disease resemble mutating and antigen-selected GC B cells, there is evidence that HRS cells in classical HL originate from pre-apoptotic GC B cells. HRS cells of the recently defined novel subtype lymphocyte-rich classical HL moleculary resemble HRS cells of the other types of classical HL, but there appear to be phenotypic differences. In rare cases, HRS cells derive from T cells. In contrast to previous speculations, cell fusion apparently does not play a role in the generation of the tumour clone. By gene expression profiling of HL cell lines, it became evident that HRS cells have lost most of the B cell-typical gene expression program, which may explain why these cells can persist without B cell receptor expression and which suggests that at least one of the transforming events involved in HL pathogenesis affects a master regulator of cell lineage identity.

Animals↗

PubMed2001

Survival and clonal expansion of mutating "forbidden" (immunoglobulin receptor-deficient) epstein-barr virus-infected b cells in angioimmunoblastic t cell lymphoma.

Angioimmunoblastic lymphadenopathy with dysproteinemia (AILD) is a peculiar T cell lymphoma, as expanding B cell clones are often present besides the malignant T cell clones. In addition, large numbers of Epstein-Barr virus (EBV)-infected B cells are frequently observed. To analyze the differentiation status and clonal composition of EBV-harboring B cells in AILD, single EBV-infected cells were micromanipulated from lymph nodes of six patients with frequent EBV(+) cells and their rearranged immunoglobulin (Ig) genes analyzed. Most EBV-infected B cells carried mutated Ig genes, indicating that in AILD, EBV preferentially resides in memory and/or germinal center B cells. EBV(+) B cell clones observed in all six cases ranged from small polyclonal to large monoclonal expansions and often showed ongoing somatic hypermutation while EBV(-) B cells showed little tendency for clonal expansion. Surprisingly, many members of expanding B cell clones had acquired destructive mutations in originally functional V gene rearrangements and showed an unfavorable high load of replacement mutations in the framework regions, indicating that they accumulated mutations over repeated rounds of mutation and division while not being selected through their antigen receptor. This sustained selection-free accumulation of somatic mutations is unique to AILD. Moreover, the survival and clonal expansion of "forbidden" (i.e., Ig-deficient) B cells has not been observed before in vivo and thus represents a novel type of viral latency in the B cell compartment. It is likely the interplay between the microenvironment in AILD lymph nodes and the viral transformation that leads to the survival and clonal expansion of Ig-less B cells.

Adult↗

PubMed2001

B-cell development in progressively transformed germinal centers: similarities and differences compared with classical germinal centers and lymphocyte-predominant Hodgkin disease.

Progressively transformed germinal centers (PTGCs) are histologic structures mainly composed of small resting B cells and intermingled proliferating centroblast-like cells. The B-cell differentiation processes within PTGCs and their relation to classical germinal centers (GC) and to lymphocyte-predominant Hodgkin disease (LPHD), with which PTGCs are often associated, are largely unknown. To address these issues, single small resting (Ki67-) and proliferating (Ki67+) centroblast-like cells were isolated from 7 PTGCs of 5 lymph nodes, and rearranged immunoglobulin genes were amplified and sequenced. Most small resting B cells were clonally unrelated, and most carried unmutated immunoglobulin gene rearrangements resembling mantle zone B cells. Small resting B cells with mutated immunoglobulin gene rearrangements may represent centrocytes, memory B cells, or both. Among the centroblast-like Ki67+ cells, expanded B-cell clones were observed in 6 of 7 PTGCs analyzed. Clonally related V region genes showed extensive intraclonal diversity, and the mutation pattern indicated stringent selection of the cells for the expression of functional antigen receptors. Thus, somatic hypermutation, clonal expansion, and selection occur also in the disorganized PTGC microenvironment, as in classical GCs. In lymph nodes affected by PTGCs, no clonal expansion across the borders of individual PTGCs was observed, distinguishing PTGCs from LPHD.

Adolescent↗

PubMed2001

Evidence that Hodgkin and Reed-Sternberg cells in Hodgkin disease do not represent cell fusions.

In most cases, Hodgkin and Reed-Sternberg (HRS) cells of classical Hodgkin disease (HD) carry rearranged immunoglobulin (Ig) genes and thus derive from B cells. In rare cases, HRS cells originate from T cells. However, based on the unusual immunophenotype of HRS cells, often showing coexpression of markers typical for different hematopoietic lineages, and the regular detection of numerical chromosomal abnormalities, it has been speculated that HRS cells might represent cell fusions. Five cases of HD with 2 rearranged IgH alleles were analyzed for the presence of additional IgH alleles in germline configuration as a potential footprint of a cell fusion between a B and a non-B cell. Similarly, one case of T-cell-derived HD with biallelic T-cell receptor beta (TCRbeta) rearrangements was studied for the presence of unrearranged TCRbeta alleles. In none of the 6 cases was evidence for additional IgH (or TCRbeta) alleles obtained, strongly arguing against a role of cell fusion in HRS cell generation.

Cell Fusion↗

PubMed2001

Regulation of immunoglobulin light chain gene rearrangements during early B cell development in the human.

Southern blot analyses of immunoglobulin light chain gene rearrangements in human leukemias and myelomas indicated that lambda loci in kappa-producing cells are largely unrearranged while kappa loci in lambda producers are often rearranged and inactivated by rearrangements of the kappa-deleting element (KDE). For a systematic analysis of the regulation of light chain rearrangements during early B cell development in normal human B cells also considering functionality of the rearrangements, we used FACS-sorted single naive kappa- and lambda-expressing B cells from peripheral blood of healthy humans. V(kappa)J(kappa) and V(lambda)J(lambda) joints and rearrangements involving the KDE were amplified simultaneously from single cells and sequenced. Whereas only 2 - 3 % of kappa-expressing cells carry V(lambda)J(lambda) joints, nearly all lambda-expressing cells have rearranged kappa loci and indeed carry V(kappa)J(kappa) joints. The V(kappa)J(kappa) joints in lambda-expressing cells exhibit preferential J(kappa)4 and J(kappa)5 over J(kappa)1 and J(kappa)2 usage compared to kappa-expressing cells. Thirty percent of the V(kappa)J(kappa) joints in lambda producers are rearranged in-frame. These data indicate extensive sequential V(kappa)-J(kappa) rearrangements and inactivation of functional V(kappa)J(kappa) joints in lambda-expressing cells, presumably before V(lambda)J(lambda) joining.

B-Lymphocytes↗

PubMed2001

Receptor revision plays no major role in shaping the receptor repertoire of human memory B cells after the onset of somatic hypermutation.

In order to determine whether V gene replacement accompanies somatic hypermutation in the germinal center (GC) reaction in the human, we analyzed V(kappa)J(kappa) and V(lambda)J(lambda) joints and the kappa-deleting element in single lambda(+) naive and post GC B cells for rearrangements at the kappa and lambda loci. Among 265 lambda(+) post GC B cells, not a single unequivocal and only two potential examples of a cell that switched to lambda light chain expression after accumulation of (unfavorable) mutations in its productive V(kappa) rearrangement were observed. Taking the PCR efficiency into account, the frequency of such cells is likely below 3 %. In addition, heavy and light chain gene rearrangements were amplified and sequenced from the oligoclonal population of IgD-only peripheral blood post GC B cells which display extensive intraclonal sequence diversity. Among 61 IgD-only B cells belonging to 15 clones with intraclonal diversity, no combination of V gene rearrangements indicating receptor revision during clonal expansion was observed. Moreover, among 124 and 49 V(H) genes amplified from IgD-only and class-switched B cells, respectively, not a single example of V(H) revision through V(H) hybrid generation was detected. These results suggest that in the human GC reaction V gene replacement either does not usually accompany somatic hypermutation or is mostly counterselected.

B-Lymphocytes↗

PubMed2001

Cytogenetic and molecular characterization of a patient with simultaneous B-cell chronic lymphocytic leukemia and peripheral T-cell lymphoma.

A patient is described who developed a peripheral T-cell lymphoma (PTCL) after a 6-year history of B-cell chronic lymphocytic leukemia (B-CLL). The progression of the T-cell disease spreading to pleura and skin terminated the course of the disease. A cytogenetic analysis performed six years after the first onset of the B-CLL showed the presence of two clones, one with trisomy 12 and another with inv(14)(q11q32.1) and trisomy 8. Combined immunophenotyping and fluorescence in situ hybridization demonstrated that only CD19+ cells contained a trisomy 12, whereas CD3+ cells contained a trisomy 8. Analyses of IgH and TCR rearrangements in single micromanipulated B- and T-cells lacked evidence for a clonal relation between B-CLL and PTCL cells. Based on our findings, we discuss the different hypotheses which might explain the development of simultaneous PTCL and B-CLL.

Aged↗

PubMed2001

Molecular single-cell analysis of Hodgkin- and Reed-Sternberg cells harboring unmutated immunoglobulin variable region genes.

Hodgkin- and Reed-Sternberg (H/RS) cells in classical Hodgkin's disease of the B lineage are the clonal progeny of antigen-experienced B cells harboring highly mutated immunoglobulin variable (V) region genes. Based on the detection of obviously destructive somatic mutations in a fraction of cases, we speculated that H/RS cells may be derived from a pre-apoptotic germinal center B cell. Seemingly contradicting this speculation, we present here the first case of classical Hodgkin's disease with H/RS cells harboring unmutated, potentially functional V region genes, which may indicate the derivation of the H/RS clone from a naive B cell. However, germinal center founder cells, which have not yet acquired somatic mutations, already have the intrinsic propensity to die by apoptosis. Thus, the rare occurrence of H/RS cells with unmutated V genes is expected if the H/RS cells are derived from the pool of pre-apoptotic germinal center B cells.

Aged↗

PubMed2000

Molecular single-cell analysis of the clonal relationship of small Epstein-Barr virus-infected cells and Epstein-Barr virus-harboring Hodgkin and Reed/Sternberg cells in Hodgkin disease.

Epstein-Barr virus (EBV) can be detected in the tumor cells of approximately 40% of cases of classical Hodgkin disease (cHD). Clonality studies suggest that infection of the neoplastic Hodgkin and Reed/Sternberg (HRS) cells occurs before tumor clone expansion. In EBV-positive cases, variable numbers of EBER-positive small B cells are sometimes also observed that immunohistologically differ from the neoplastic cells by lack of CD30 and latent membrane protein 1 expression. To analyze the clonal relationship between these EBV(+) cells and the HRS cells, single EBV-infected CD30(-) B cells, as well as HRS cells from 3 cases of EBV-positive cHD were micromanipulated, their immunoglobulin gene rearrangements amplified and then compared with each other. In 2 cases, all small EBV-infected cells were clonally unrelated to the HRS cells. In a third case, 2 of 29 small CD30(-) cells were found to carry HRS cell-specific rearrangements. Thus, small CD30(-) EBV-infected B cells in cHD belong to the HRS tumor clone rarely, if at all. In all cases, small clones unrelated to the HRS cell clones were identified among the small EBV(+) CD30(-) cells. The vast majority of small EBV(+) CD30(-) B cells was found to carry somatically mutated V region genes, indicating that in lymph nodes of patients with HD, like in the peripheral blood of healthy individuals, EBV persists in memory B cells.

Adult↗

PubMed2000

Somatic mutations of the CD95 gene in Hodgkin and Reed-Sternberg cells.

Hodgkin and Reed-Sternberg (H/RS) cells in classical Hodgkin's disease (cHD) are thought to be derived from preapoptotic germinal center B cells. However, little is known about the transforming events rescuing the precursor of the H/RS cells from apoptosis. Given the importance of CD95 (Apo-1/Fas)-mediated apoptosis for negative selection within the germinal center, single micromanipulated H/RS cells from 10 cases of cHD were analyzed for somatic mutations within the CD95 gene. Three clonal mutations within the 5' regions were amplified from single H/RS cells in one case. From H/RS cells of another case, two mutations within the last exon coding for the death domain were detected. About half of these H/RS cells carried a monoallelic stop-codon; the remaining tumor cells harbored a monoallelic replacement mutation. Both mutations likely impair CD95 function. Because all these H/RS cells also bear clonal mutations inactivating the IkappaB alpha gene, the IkappaB alpha mutations occurred earlier than those of the CD95 gene in the sequence of transforming events leading to cHD. In conclusion, somatic mutations of the CD95 gene occur in a fraction of cHD cases and may favor the escape of the precursor of the H/RS clone from apoptosis.

Adult↗

PubMed2000

Identification and functional characterization of the human and murine fibroblast growth factor receptor 4 promoters.

Fibroblast growth factor receptors (FGFRs) play crucial roles in signal transduction of adult tissues and during embryonic development. To study the transcriptional control, we isolated and characterized the promoter of human FGFR4. Two transcription initiation sites were identified. The deletion analysis in different cell types defined a core promoter reaching from -9 to -198, lacking TATA and CCAAT boxes but displaying high GC content (77%) in a stretch of 300 bp upstream of the major mRNA start. This region harbors multiple binding motifs for transcription factors. Moreover, the region between -1085 and -1140 contains a potential repressor element, which downregulates transcriptional activity. To identify conserved regulatory elements, we isolated and analyzed also the murine FGFR4 promoter. Only one transcription start was identified using RNase protection assays. Sequence alignment of human and mouse shows a striking similarity in the core promoter region of both genes, encompassing conserved transcription factor binding sites and a splice acceptor site. Furthermore, the region containing the putative repressor element is also conserved suggesting a functional role for gene expression.

Animals↗

PubMed2000

Rare occurrence of classical Hodgkin's disease as a T cell lymphoma.

Recent work identified Hodgkin and Reed-Sternberg (H/RS) cells in classical Hodgkin's disease (cHD) as clonal progeny of mature B cells. Therefore, it is generally assumed that cHD homogenously represents a B cell lymphoma. In a subset of cHD, however, H/RS cells expressing T cell-associated proteins may be candidates for alternative lineage derivation. Single H/RS cells with cytotoxic T cell phenotype were micromanipulated from three cases of cHD and analyzed by single cell polymerase chain reaction for immunoglobulin heavy (IgH) and light chain (IgL) gene rearrangements, T cell receptor (TCR)-beta gene rearrangements, and germline configuration of the IgH and TCR-beta loci. H/RS cells from two cases of cHD harbored clonal, somatically mutated Ig gene rearrangements, whereas TCR-beta loci were in germline configuration. In contrast, H/RS cells from an additional case harbored clonal TCR-beta variable/diversity/joining (VDJ) and DJ gene rearrangements, whereas the IgH locus was in germline configuration on both alleles. Thus, in two cases of cHD with H/RS cells expressing cytotoxic T cell molecules, the tumor cells are derived from mature B cells that aberrantly express T cell markers. In a third case, however, H/RS cells were derived from a T cell, demonstrating that cHD can also occur as a T cell lymphoma.

Adult↗

PubMed2000

Clonal deleterious mutations in the IkappaBalpha gene in the malignant cells in Hodgkin's lymphoma.

Members of the nuclear factor (NF)-kappaB family of transcription factors play a crucial role in cellular activation, immune responses, and oncogenesis. In most cells, they are kept inactive in the cytosol by complex formation with members of the inhibitor of NF-kappaB (IkappaB) family, whose degradation activates NF-kappaB in response to diverse stimuli. In Hodgkin's lymphoma (HL), high constitutive nuclear activity of NF-kappaB is characteristic of the malignant Hodgkin and Reed-Sternberg (H/RS) cells, which occur at low number in a background of nonneoplastic inflammatory cells. In single H/RS cells micromanipulated from histological sections of HL, we detect clonal deleterious somatic mutations in the IkappaBalpha gene in two of three Epstein-Barr virus (EBV)-negative cases but not in two EBV-positive cases (in which a viral oncogene may account for NF-kappaB activation). There was no evidence for IkappaBalpha mutations in two non-HL entities or in normal germinal center B cells. This study establishes deleterious IkappaBalpha mutations as the first recurrent genetic defect found in H/RS cells, indicating a role of IkappaBalpha defects in the pathogenesis of HL and implying that IkappaBalpha is a tumor suppressor gene.

Adult↗

PubMed2000

CD5-positive B cells in healthy elderly humans are a polyclonal B cell population.

B cell chronic lymphocytic leukemia (B-CLL) is a disease of the elderly and is characterized by a malignant clone of CD5+ B cells. In old mice, clonal expansions of CD5+ B cells are a common feature, and these animals frequently develop B-CLL. To investigate whether clonal expansion of CD5+ B cells also occurs in elderly humans, predisposing for the development of B-CLL, we analyzed VH gene rearrangements of CD5+ B cells from blood samples of four healthy, 65-82-years-old volunteers as markers of clonality. CD5+ and CD5-B cells were obtained by cell sorting, CDRIII of rearranged VH genes were amplified by polymerase chain reaction, and fragment length analysis was performed. All samples demonstrated a polyclonal pattern of VH gene length distribution. In addition, VH gene rearrangements were amplified and sequenced from sorted single cells of two of the donors. No clonally related CD5+ or CD5- B cells were observed. Thus, development of dominant clones of CD5+ peripheral blood B cells is unlikely to be a common trait of elderly individuals.

Aging↗

PubMed1999

Identification of common germinal-center B-cell precursors in two patients with both Hodgkin's disease and non-Hodgkin's lymphoma.

BACKGROUND: Hodgkin's disease and non-Hodgkin's B-cell lymphoma occasionally occur in the same patient. The identification of a common precursor of the two types of lymphoma would show definitively that Reed-Sternberg cells originate from B cells. METHODS: We studied lymphomas from two patients, one with a composite lymphoma (classic Hodgkin's disease and a follicular lymphoma in the same lymph node) and the other with a T-cell-rich B-cell lymphoma that was followed by classic Hodgkin's disease. Single Reed-Sternberg cells and non-Hodgkin's lymphoma cells from frozen sections were micromanipulated. The rearranged immunoglobulin variable-region genes (V genes) of the heavy and light chains were amplified by the polymerase chain reaction from genomic DNA and sequenced. RESULTS: In both patients, the Reed-Sternberg cells were related clonally to the non-Hodgkin's lymphoma B cells. The V genes carried somatic mutations (a hallmark of germinal-center B cells and their descendants). In both patients, some somatic mutations were shared by the Reed-Sternberg and non-Hodgkin's lymphoma cells, whereas other somatic mutations were found exclusively in one or the other cell type. CONCLUSIONS: In two patients with classic Hodgkin's disease and non-Hodgkin's B-cell lymphoma, we identified a common B-cell precursor, probably a germinal-center B-cell, for both lymphomas. This finding suggests that the two types of lymphoma underwent both shared and distinct transforming events and provides proof of the B-cell derivation of Reed-Sternberg cells in classic Hodgkin's disease.

Aged↗

PubMed1999

Molecular analysis of single B cells from T-cell-rich B-cell lymphoma shows the derivation of the tumor cells from mutating germinal center B cells and exemplifies means by which immunoglobulin genes are modified in germinal center B cells.

T-cell-rich B-cell lymphoma (TCRBCL) belongs to the group of diffuse large cell lymphomas (DLL). It is characterized by a small number of tumor B cells among a major population of nonmalignant polyclonal T cells. To identify the developmental stage of the tumor progenitor cells, we micromanipulated the putative neoplastic large CD20(+) cells from TCRBCLs and amplified and sequenced immunoglobulin (Ig) V gene rearrangements from individual cells. In six cases, clonal Ig heavy, as well as light chain, gene rearrangements were amplified from the isolated B cells. All six cases harbored somatically mutated V gene rearrangements with an average mutation frequency of 15.5% for heavy (VH) and 5.9% for light (VL) chains and intraclonal diversity based on somatic mutation. These findings identify germinal center (GC) B cells as the precursors of the transformed B cells in TCRBCL. The study also exemplifies various means how Ig gene rearrangements can be modified by GC B cells or their malignant counterparts in TCRBCL: In one case, the tumor precursor may have switched from kappa to lambda light chain expression after acquiring a crippling mutation within the initially functional kappa light chain gene. In another case, the tumor cells harbor two in-frame VH gene rearrangements, one of which was rendered nonfunctional by somatic mutation. Either the tumor cell precursor entered the GC with two potentially functional in-frame rearrangements or the second VHDHJH rearrangement occurred in the GC after the initial in-frame rearrangement was inactivated by somatic mutation. Finally, in each of the six cases, at least one cell contained two (or more) copies of a clonal Ig gene rearrangement with sequence variations between these copies. The presence of sequence variants for V region genes within single B cells has so far not been observed in any other normal or transformed B lymphocyte. Fluorescence in situ hybridization (FISH) points to a generalized polyploidy of the tumor cells.

Adult↗