Search PubMed⌕ Search

Biomedical subjects

H Stein

Publications and source records attributed to H Stein.

At least 55 records · Page 3Linked to original sources
PubMed2002

Hodgkin's lymphoma: the pathologist's viewpoint.

Despite its well known histological and clinical features, Hodgkin's lymphoma (HL) has recently been the object of intense research activity, leading to a better understanding of its phenotype, molecular characteristics, histogenesis, and possible mechanisms of lymphomagenesis. There is complete consensus on the B cell derivation of the tumour in most cases, and on the relevance of Epstein-Barr virus infection and defective cytokinesis in at least a proportion of patients. The REAL/WHO classification recognises a basic distinction between lymphocyte predominance HL (LP-HL) and classic HL (CHL), reflecting the differences in clinical presentation and behaviour, morphology, phenotype, and molecular features. CHL has been classified into four subtypes: lymphocyte rich, nodular sclerosing, with mixed cellularity, and lymphocyte depleted. The borders between CHL and anaplastic large cell lymphoma have become sharper, whereas those between LP-HL and T cell rich B cell lymphoma remain ill defined. Treatments adjusted to the pathobiological characteristics of the tumour in at risk patients have been proposed and are on the way to being applied.

Diagnosis, Differential↗
PubMed2001

Regulation of the intestinal mucin MUC2 gene expression in vivo: evidence for the role of promoter methylation.

In the present work we investigated the in vivo regulation of the mucin gene MUC2, which is overexpressed in all mucinous colorectal carcinomas. The inhibition of methylation by 5-azadeoxycytidine induces de novo expression of MUC2 in the colon carcinoma cell line COLO 205. The expression is retained in xenograft tissue and the cells give rise to MUC2-expressing tumours in nude mice. The strong expression of MUC2 in the normal human goblet cells and in the tissue of human mucinous colorectal carcinomas is associated with the average methylation of about 50% at every investigated CpG site of the MUC2 promoter. In contrast, MUC2 promoter in the non-expressing normal columnar cells and in the non-mucinous carcinoma tissue is methylated to nearly 100%. These data show that (i) low methylation of MUC2 promoter is associated with MUC2 expression in vivo and (ii) the pattern of MUC2 promoter methylation in the normal goblet or columnar cells most closely resembles that in mucinous or non-mucinous colorectal carcinomas, respectively. They indicate that MUC2 expression in vivo is regulated by promoter methylation and support the hypothesis that cells with goblet-like differentiation give rise to mucinous colonic carcinomas.

Animals↗
PubMed2001

Structure of the Hodgkin's lymphoma-associated human CD30 gene and the influence of a microsatellite region on its expression in CD30(+) cell lines.

The CD30 antigen is a member of the tumor necrosis factor receptor (TNFR) family which is overexpressed on the surface of the tumor cells of Hodgkin's lymphoma, anaplastic large cell lymphoma (ALCL), and embryonal carcinoma of the testis. In this study the entire cd30 gene which is more than 24000 bp long and organized in eight exons was characterized by analyzing cosmid and phage lambda clones from human placental libraries with long-range polymerase chain reaction (PCR) and sequencing. Differences to other genes of the TNFR family were detected in the region encoding the extracellular domain of the cd30 gene. In nearly all other TNFR genes, the coding region of each cysteine-rich repeat is interrupted by one intron, i.e., the 3-4 cysteine-rich repeats of these receptors are encoded by at least 4-5 exons, whereas the six cysteine-rich repeats of the cd30 gene are encoded by two exons, i.e., each of these exons encode three cysteine-rich repeats. In addition, we also found a genetic polymorphism of tetranucleotide ATCC-repeats in the 5' part of the CD30 promoter. This region was amplified by PCR from seven CD30 overexpressing human lymphoid cell lines and five human tissues with an absent or very low CD30 expression. The amplification products showed length differences of more than 550 bp. The number of the ATCC-repeats was higher in CD30(+) cell lines than in normal tissues. Comparison of the individual PCR products in reporter gene assays revealed that the CD30 promoter activity increased with the length of this polymorphic region up to eightfold. The data suggest that the number of ATCC-repeats in the 5' region of the CD30 promoter modulates the regulation of CD30 expression.

Base Sequence↗
PubMed2001

Defective octamer-dependent transcription is responsible for silenced immunoglobulin transcription in Reed-Sternberg cells.

The absence of immunoglobulin (Ig) expression in B-cell-derived Hodgkin and Reed-Sternberg (HRS) cells of classical Hodgkin disease (cHD) was initially suggested to be caused by crippling mutations in the Ig promoter or coding region. More recent investigations have, however, challenged this concept. This study addressed the role of mutations in the Ig promoter region in HRS cells. Nine cases of cHD and 3 B-cell-derived HD lines were analyzed for mutations in the TATA box and octamer motif of the Ig promoter. Mutations in the octamer motif were found in only 1 of the 9 cases and in 1 of the 3 HD cell lines (L1236). Furthermore, in all cases either a complete lack or strong reduction in the expression of the Oct2 transcription factor and the BOB.1/OBF.1 coactivator were found. The relevance of the rare promoter mutations was investigated by assaying the activity of Ig promoter reporter constructs transfected into the HD cell line L1236, which harbors a mutated octamer motif. These Ig reporter constructs were completely inactive in L1236 cells; however, their activity could be reconstituted by the cotransfection of a BOB.1/OBF.1 expression vector. The additional transfection with an Oct2 expression vector did not further enhance the Ig promoter activity. The conclusions drawn from these results are that crippling mutations in the Ig promoter and coding region are not the sole cause for the lack of Ig expression in HRS cells and that defects in the transcription machinery such as absence of BOB.1/OBF.1 are more important for this phenomenon.

Carrier Proteins↗
PubMed2001

Analysis of BCL-6 mutations in classic Hodgkin disease of the B- and T-cell type.

BCL-6 is essential for germinal center formation and thus for affinity maturation of immunoglobulin (Ig) genes by somatic mutations. The 5'-noncoding region of the BCL-6 gene is even a target for the mutation machinery. Translocations of the BCL-6 gene to heterologous promoters and mutations of its 5'-noncoding regulatory region were reported to be potential mechanisms for deregulating BCL-6 expression and for playing a role in the genesis of non-Hodgkin lymphoma. In line with this hypothesis is the observation that B-cell lymphoma with somatic mutations, such as diffuse large B-cell lymphoma and follicular lymphoma, also carry BCL-6 mutations, some of which are recurrently detectable. Classic Hodgkin disease (cHD) is also derived from B cells with high loads of somatic mutations and thus a further candidate for BCL-6 mutations. To determine the presence and potential role of BCL-6 mutations in cHD, the 5'-noncoding BCL-6 proportion of single Hodgkin and Reed-Sternberg (HRS) cells from 6 cases of cHD and 6 cases of HD-derived cell lines was analyzed. All B-cell-derived HD cases and cell lines harbored BCL-6 mutations. In contrast, both T-cell-derived HD cases and cell lines were devoid of BCL-6 mutations. With only one exception, there were no lymphoma-specific recurrent BCL-6 mutations detected, and BCL-6 protein was absent from the HRS cells of most cases. In conclusion, (1) somatic BCL-6 mutations are restricted to cHD cases of B-cell origin, and (2) the BCL-6 mutations represent mostly irrelevant somatic base substitutions without consequences for BCL-6 protein expression and the pathogenesis of cHD.

5' Untranslated Regions↗
PubMed2001

The diversity of rearranged immunoglobulin heavy chain variable region genes in peripheral blood B cells of preterm infants is restricted by short third complementarity-determining regions but not by limited gene segment usage.

The immunoglobulin diversity is restricted in fetal liver B cells. This study examined whether peripheral blood B cells of extremely preterm infants show similar restrictions (overrepresentation of some gene segments, short third complementarity-determining regions [CDR3]). DNA of rearranged immunoglobulin heavy chain genes was amplified by polymerase chain reaction, cloned, and sequenced. A total of 417 sequences were analyzed from 6 preterm infants (25-28 weeks of gestation), 6 term infants, and 6 adults. Gene segments from the entire V(H) and D(H) gene locus were rearranged in preterm infants, even though the D(H)7-27 segment was overrepresented (17% of rearrangements) compared to term infants (7%) and adults (2%). CDR3 was shorter in preterm infants (40 +/- 10 nucleotides) than in term infants (44 +/- 12) and adults (48 +/- 14) (P <.001) due to shorter N regions. Somatic mutations were exclusively found in term neonates and adults (mutational frequency 0.8% and 1.8%). We conclude that preterm infants have no limitations in gene segment usage, whereas the diversity of CDR3 is restricted throughout gestation.

Adult↗
PubMed2001

Down-regulation of BOB.1/OBF.1 and Oct2 in classical Hodgkin disease but not in lymphocyte predominant Hodgkin disease correlates with immunoglobulin transcription.

In contrast to the tumor cells (L&H cells) of lymphocyte predominant Hodgkin disease (LPHD), Hodgkin and Reed-Sternberg (HRS) cells of classical Hodgkin disease (cHD) are unable to transcribe immunoglobulin, despite the presence of rearranged immunoglobulin genes. Although initial studies have suggested crippling immunoglobulin gene mutations to be the cause of absent immunoglobulin expression in cHD, recent work of our group has demonstrated an impaired activation of the immunoglobulin promoter as a superior mechanism. As immunoglobulin transcription is mainly regulated by the B-cell transcription factors Oct2 and BOB.1/OBF.1, we analyzed 35 cases of LPHD, 32 cases of cHD, and 2 Hodgkin disease cell lines for the expression of these transcription factors and also in parallel for immunoglobulin expression. Our results demonstrate an absence of Oct2 and/or BOB.1/OBF.1 in cHD and a striking overexpression of Oct2 in LPHD. Immunoglobulin expression was lacking in cHD but present in LPHD. Furthermore, the reintroduction of BOB.1/OBF.1 and Oct2 into cultured HRS cells restored the activity of cotransduced immunoglobulin promoter constructs. Our findings dismiss the concept that the different immunoglobulin expression in cHD and LPHD is due to disrupting mutations of immunoglobulin V genes in cHD but is most likely due to a down-regulation of Oct2 and/or BOB.1/OBF.1. This study further revealed Oct2 as a new and valuable marker for the identification of L&H cells and their distinction from HRS cells. The impairment of immunoglobulin transcription with a down-regulated synthesis of Oct2 and BOB.1/OBF.1 is the first established general recurrent defect found in HRS cells.

Cell Line↗
PubMed2001

Analysis of single EBER-positive and negative tumour cells in EBV-harbouring B-cell non-Hodgkin lymphomas.

In situ hybridization for Epstein-Barr virus (EBV) encoded small nuclear RNAs (EBERs) is the method of choice for the detection of EBV infection at the single cell level. With the application of this technique it was shown that non-Hodgkin's lymphomas of B-cell type may be associated with an EBV infection of tumour cells. Interestingly, in many EBV-positive cases, only a proportion of the tumour cell population has been found to be EBER-positive. To clarify whether EBV is absent or whether EBER gene expression is downregulated in EBER-negative tumour cells, single EBER-positive and negative tumour cells were isolated from paraffin sections from four B-cell non-Hodgkin's lymphoma cases with partial EBER expression in the neoplastic cells. These single cells were then screened for the presence of EBV DNA by a nested single cell PCR. EBV DNA was undetectable in all but one of the 86 EBER-negative cells, whereas in the EBER-positive cells EBV-specific DNA amplificates could be generated following single cell PCR. This finding prompts the conclusion that the inability to detect EBERs in a proportion of tumour cells is not due to a down-regulation of gene expression but to a real absence of EBV from these cells. This partial absence of EBV is thought to be caused by a loss of the EBV episomes during cell division, rather than by infection of only a proportion of the tumour cells.

Cell Separation↗
PubMed2001

Secondary ocular involvement in systemic "memory" B-cell lymphocytic leukemia.

OBJECTIVE: Recently, B-cell chronic lymphocytic leukemia (B-CLL) has been subdivided into "naive" B-CLL and "memory" B-CLL on the basis of the presence of somatic mutations in the variable region of the immunoglobulin heavy chain gene (IgH). The aim of the current paper was to report the clinical, histopathologic, and molecular biologic findings of intraocular and ocular adnexal involvement in a patient with systemic B-CLL. DESIGN: Case report. INTERVENTION: Treatment consisted of systemic chemotherapy, conjunctival biopsies, and, ultimately, enucleation of the left eye. METHODS: Histopathologic findings of a bone marrow biopsy, the conjunctival biopsies, and the enucleated eye were compared. Further, extensive immunohistochemistry, polymerase chain reaction (PCR) for the detection of heavy chain (IgH) gene rearrangement, gene scan analysis, and DNA sequencing were performed on all tissues. RESULTS: The tumor manifestations in all specimens demonstrated similar morphologic and immunophenotypic characteristics, consistent with the diagnosis of B-CLL. Immunoglobulin-H PCR and gene scan analysis showed that the B-CLL infiltrates consisted of B cells derived from the same clone. DNA sequencing demonstrated the presence of eight somatic mutations in the variable region of IgH, consistent with "memory" B-CLL. CONCLUSIONS: Secondary ocular manifestations of B-CLL occur relatively commonly during disease progression. In the current case of memory B-CLL, ocular manifestation of the disease occurred 16 years after initial diagnosis, agreeing with clinical studies suggesting that a less aggressive course is seen in "memory" B-CLL than its counterpart, "naive" B-CLL. Somatic mutation analysis in the variable region of IgH in B-CLL should be part of routine staging investigations to aid the prediction of the individual clinical course in B-CLL patients and to determine new therapeutic strategies.

Antibodies, Monoclonal↗
PubMed2001

Recovery of muscles of old rats after hindlimb immobilisation by external fixation is impaired compared with those of young rats.

The right hindlimbs of 24-month-old female Wistar rats were immobilised for 4weeks using external fixation of the knee joint. In a further group, after the external fixation was removed, the rats were allowed to remobilise for an additional 4weeks. Hindlimb immobilisation for 4weeks caused a 32-42% reduction in wet weights of the hindlimb muscles of the rats as compared to those of the contralateral non-immobilised legs. After 4weeks of remobilisation the hindlimb muscles had not returned to the "control" weights. Biochemical changes in the gastrocnemius muscle resulting from the external fixation showed greatly elevated acid phosphatase activities (33.2%) and markedly reduced creatine phosphokinase activities (17.2%), which did not recover to preimmobilisation values after 4weeks of remobilisation. Light and transmission electron microscopy showed that remobilisation for 4weeks (after external fixation) resulted in only partial morphological restoration of the damage to the muscles in these aged rats. A comparison of similar hindlimb external fixation and remobilisation in young (6months old) rats showed that remobilisation caused a substantial recovery in biochemical parameters in both age groups, with the muscles of the young group (but not the old group) often reaching almost complete recovery accompanied by morphological restoration. We conclude that the net gain in the recovery period of biochemical and morphological parameters is significantly greater in the young rats compared to the old rats indicating that muscle metabolism and capacity for recovery from disuse atrophy is impaired with ageing.

Aging↗
PubMed2001

Osteoprogenitor cells of mature human skeletal muscle tissue: an in vitro study.

The presence of osteogenic progenitors in human skeletal muscle is suggested by the formation of ectopic bone in clinical and experimental conditions, but their direct identification has not yet been demonstrated. The aims of this study were to identify osteogenic progenitor cells in human skeletal muscle tissue and to expand and characterize them in culture. Specimens of gracilis and semitendinosus muscle were obtained from young adults and digested to separate the connective tissue and satellite cell fractions. The cells were cultured and characterized morphologically and immunohistochemically using antibodies known to be reactive with primitive osteoprogenitor cells, pericytes, intermediate filaments, and endothelial cells. Alkaline phosphatase activity and osteocalcin gene expression were also determined. In the early stages of culture, the connective tissue cells obtained were highly positive for primitive osteoprogenitor cell and for pericyte markers. Alkaline phosphatase activity was detectable at early stages of culture and rose as a function of time, whereas primitive osteoprogenitor cell markers declined and osteocalcin mRNA expression became detectable by reverse transcriptase-polymerase chain reaction (RT-PCR). It is shown that human skeletal muscle connective tissue contains osteogenic progenitor cells. Their identification as pericytes, perivascular cells with established osteogenic potential, suggests a cellular link between angiogenesis and bone formation in muscle tissue. These cells are easily cultured and expanded in vitro by standard techniques, providing an alternative source of osteogenic progenitor cells for possible cell-based therapeutic use in certain conditions.

Actins↗
PubMed2001

Her-2/neu gene amplification, elevated mRNA expression, and protein overexpression in the metaplasia-dysplasia-adenocarcinoma sequence of Barrett's esophagus.

SUMMARY: The importance of alterations of the Her-2/neu oncogene in the tumorigenesis of Barrett's adenocarcinoma (BCA) is discussed controversially. In the present study, we evaluated for the first time the Her-2/neu status in the metaplasia-dysplasia-adenocarcinoma sequence of BCA simultaneously at the DNA, mRNA, and protein level using resection specimens of 25 patients. The locus-specific Her-2/neu gene status was quantified by performing fluorescence in situ hybridization, and information about the ploidy status of chromosome 17 was obtained. Tissue sections from the same areas were used for quantitative RT-PCR (TaqMan RT-PCR) of laser-microdissected tumor cells and for immunohistochemistry to quantify Her-2/neu mRNA and oncoprotein expression. Her-2/neu gene amplification was observed in 35% of BCA, and all of these samples showed strong overexpression of both mRNA and oncoprotein. A polysomy 17 without Her-2/neu gene amplification was observed in 52% of BCA, showing a normal or moderately elevated mRNA expression and no or weak immunopositivity. From 13 areas of high-grade dysplasia (HGD) we found four to be amplified for the Her-2/neu locus, whereas five showed a polysomy 17. All four samples of HGD areas with Her-2/neu gene amplification displayed mRNA and strong oncoprotein overexpression; however, lower mRNA levels were seen than in the amplified BCA areas. None of the samples with low-grade dysplasia (LGD) showed a locus-specific Her-2/neu amplification, but polysomy 17 was present in four of eight cases. No changes were detected in BCA-associated intestinal metaplasia and squamous epithelium. In summary, only a locus-specific Her-2/neu gene amplification was associated with strong mRNA overexpression and strong membranous Her-2/neu immunostaining in BCA and HGD. A chromosome 17 polysomy, as found in the majority of BCA, led to no or weak mRNA overexpression and no or weak immunopositivity. In the metaplasia-dysplasia-adenocarcinoma sequence, a chromosome 17 polysomy without Her-2/neu gene amplification was already present in LGD. This may be a result of an early polyploidization, preceding the later genetic events, such as Her-2/neu gene amplification in HGD and BCA.

Adenocarcinoma↗
PubMed2001

Extranodal mantle cell lymphoma mimicking marginal zone cell lymphoma.

AIM: We report a case of mantle cell lymphoma masquerading as a marginal zone cell lymphoma. METHODS AND RESULTS: In the initial manifestation in the palatine tonsils, the neoplastic cells were found to grow exclusively within the marginal zones of secondary follicles which showed a preserved mantle zone. The few immunostains performed showed a B-cell phenotype including an immunoglobulin light chain restriction. The extranodal manifestation, the growth pattern, and the immunophenotype led to the diagnosis of an extranodal marginal zone B-cell non-Hodgkin's lymphoma (NHL). The specimen from the relapse occurring 8 months later exhibited diffuse monomorphous cells co-expressing B-cell antigens and CD5, CD43 and cyclin D1, leading to the diagnosis of mantle cell lymphoma. Re-investigation of the initial biopsy revealed that the neoplastic cells within the marginal zones had a mantle cell lymphoma immunophenotype expressing cyclin D1, the immunoglobulin heavy chains IgD and IgM and partly CD5. Both lesions harboured identical clonal immunoglobulin gene rearrangements proving that they represented different manifestations of the same lymphoma. CONCLUSION: This case emphasizes the importance of broad immunohistological investigation of B-cell NHLs involving the marginal zone.

Antigens, CD↗