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G Klein

Publications and source records attributed to G Klein.

At least 271 records · Page 15Linked to original sources
PubMed1995

Downregulation of c-myc expression after heat shock in human B-cell lines is independent of 5' mRNA sequences.

The effect of heat-shock on the expression of c-myc genes in different chromosomal contexts was investigated in a panel of human B-lymphoid cell lines. Burkitt's lymphoma cell lines with c-myc translocation breakpoints upstream of the first exon, within the exon itself, or in the first intron showed downregulation of c-myc levels as did a cell line without any translocation. The c-myc mRNA of cell lines with translocation breakpoints within the c-myc gene have previously been reported to have prolonged half-lives. After heat shock, the levels of these mRNA species were reduced with similar kinetics as the normal c-myc mRNA. An exception was an RNA species where the only c-myc sequences are derived from exon 1, showing that sequences from this part of the c-myc gene are not sufficient to mediate the rapid downregulation. Nuclear run-on analysis did not show reduced transcription of c-myc after heat shock and a comparison of cytoplasmic and total RNA did not indicate accumulation of longer, unspliced c-myc mRNA species. These observations suggest a posttranscriptional, cytoplasmic downregulation targeting exons 2 and/or 3. B-lymphoma lines transfected with a hsp70 promoter-linked c-myc gene were deficient in their ability to reinitiate proliferation after heat shock, providing a physiological rationale for the normal downregulation of c-myc after this type of physical stress.

B-Lymphocytes↗
PubMed1995

[Noninvasive monitoring of regional cerebral oxygen saturation--experiences in critical care medicine].

Regional cerebral oxygen saturation (rSO2) was studied in 15 healthy volunteers aged between 23 and 53 years (mean age: 34.2 +/- 8.7 years) and in 17 intensive care patients (mean age: 37.2 +/- 18.6 years; range 17-70). An infrared spectroscopy technique (INVOS 3100, Somanetics Co.) was used. The results were analysed in terms of age, heart rate, temperature at the recording site, blood pressure and peripheral oxygen saturation. The mean +/- SD of rSO2 was 69.4% +/- 7.0% in the healthy volunteers and 66.4% +/- 7.4% in intensive care patients (65.3% +/- 6.2% in 7 patients with a cerebral lesion, 67.1% +/- 8.4% in 10 patients with no cerebral lesion). Individual examples are presented to show that infrared spectroscopy is a useful non-invasive method of investigating cerebral oxygenation. However, regional pathological-anatomical limitations of the technique, and possible extracerebral artefacts make necessary a critical interpretation of the rSO2 values.

Adolescent↗
PubMed1995

Laparoscopic pelvic lymph node dissection combined with real-time interactive transrectal ultrasound guided transperineal radioactive seed implantation of the prostate.

Laparoscopic pelvic lymph node dissection with real-time interactive transrectal ultrasound guided transperineal radioactive seed implantation is a new method of treatment for localized carcinoma of the prostate. A total of 58 patients with clinically confined prostate cancer and negative seminal vesicle biopsies underwent staging laparoscopic pelvic lymph node dissection immediately followed by prostate implantation: 50 had 125iodine and 8 had 103palladium implants. Mean operating time was 226 minutes (range 120 to 475), mean blood loss was 57 cc (range 5 to 400) and average hospital stay was 2.2 days (range 0.5 to 8). At a mean followup of 12 months (range 6 to 24), complications included proctitis in 1.7% of the cases, dysuria in 24%, nocturia in 21% and outlet obstruction in 17%. Erectile function remained unchanged. Prostate volume decreased to 58.9% of the pretreatment value by 12 months and to 44.3% by 24 months. Mean prostate specific antigen level was 18.4 +/- 26.3 ng./ml. before treatment, 3.4 +/- 3.9 ng./ml. at 6 months, 2.3 +/- 2.3 ng./ml. at 12 months and 4.9 +/- 6.0 ng./ml. at 24 months (1.2 +/- 1.0 ng./ml. for patients with no evidence of disease). Of the patients 15.8% had local failure at 18 to 24 months as determined by positive transrectal ultrasound guided biopsy. Five of 58 patients (8.6%) had persistently elevated prostate specific antigen levels, only 1 of whom had a positive biopsy. Laparoscopic pelvic lymph node dissection with transrectal ultrasound guided implantation is a safe and promising mode of therapy for patients with localized prostate cancer.

Aged↗
PubMed1995

Differential expression of adhesion molecules in acute leukemia.

Interactions between hematopoietic cells and the stromal microenvironment are mediated by membrane-bound adhesion molecules. As the expression patterns of these molecules may alter the adhesive qualities of leukemic blasts, leukemic samples were investigated for the expression of beta 1-, beta 2-, beta 3-integrins, CD44, the three selectins and several members of the immunoglobulin family. CD44 (167/169), LFA-3 (158/169), the beta 1-integrins VLA-4 (120/123) and VLA-5 (45/51) and the beta 2-integrin LFA-1 (149/157) were found on > 70% of blasts in most cases of leukemias. Other molecules were restricted to specific differentiation stages and lineage. The beta 2-integrins Mac-1 (CD11b/CD18) and gp 150,95 (CD11c/CD18) were preferentially expressed on M4 and M5 subtypes, and NCAM (CD56) was only found on a subset of acute myeloid leukemias (17/113). Unexpectedly, the beta 1-integrins VLA-1 (1/51), VLA-2 (18/123), VLA-3 (5/43), VLA-6 (15/29) and the E-selectin (2/47) were expressed on > 70% blasts on a subset of leukemias of varied phenotype. These molecules were absent on normal CD34+ bone marrow precursors. The simultaneous analysis generally revealed a higher percentage of positive blasts in the blood than in bone marrow. Our observations therefore suggest that in leukemia these antigens are displayed on a non-adherent population that is defective and is unable to convert to an adherent, functionally active conformational state.

Acute Disease↗
PubMed1995

Xenogenization.

Explore the source record for details and available documents.

Animals↗
PubMed1995

bcl-2 inhibits wild-type p53-triggered apoptosis but not G1 cell cycle arrest and transactivation of WAF1 and bax.

We have earlier shown that wild-type (wt) p53 expressed from a temperature-sensitive construct (ts p53) triggers apoptosis in the v-myc retrovirus-induced, p53-negative T-cell lymphoma line J3D (Y. Wang et al., Cell Growth & Differ., 4: 467-473, 1993). We also found that constitutive bcl-2 expression inhibits wt p53-triggered apoptosis in these cells (Y. Wang et al., Oncogene, 8: 3427-3431, 1993). Here we demonstrate that more than 90% of the ts p53-transfected J3D cells were arrested in G1 at 18 h after induction of wt p53 expression by temperature shift to 32 degrees C. At this time, at least 80% of the cells remained viable. After 30 h at 32 degrees C, around 50% of the cells had died by apoptosis, while most of the remaining cells were still alive in G1, indicating that p53-induced apoptosis occurred following G1 arrest. The G1 cell cycle arrest at 18 h after temperature shift to 32 degrees C was reversible, as shown by the fact that the cells readily resumed exponential growth following temperature shift back to 37 degrees C, although viability dropped from around 80 to 65%. Expression of both WAF1 and bax mRNA was induced by wt p53 in both the ts p53 and ts p53/bcl-2 transfected cells. The kinetics of G1 cell cycle arrest at 32 degrees C was similar in both the ts p53 and the ts p53/bcl-2 double transfectants.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1995

Burkitt's lymphoma is a malignancy of mature B cells expressing somatically mutated V region genes.

BACKGROUND: The developmental stage from which stems the malignant B cell population in Burkitt's lymphoma (BL) is unclear. An approach to answering this question is provided by the sequence analysis of rear-ranged immunoglobulin (Ig) variable region (V) genes from BL for evidence of somatic mutations, together with a phenotypic characterization. As somatic hypermutation of Ig V region genes occurs in germinal center B cells, somatically mutated Ig genes are found in germinal center B cells and their descendents. MATERIALS AND METHODS: Rearranged V kappa region genes from 10 kappa-expressing sporadic and endemic BL-derived cell lines (9 IgM and 1 IgG positive) and three kappa-expressing endemic BL biopsy specimens were amplified by polymerase chain reaction and sequenced. In addition, VH region gene sequences from these cell lines were determined. RESULTS: All BL cell lines and the three biopsy specimens carried somatically mutated V region genes. The average mutation frequency of rearranged V kappa genes from eight BL cell lines established from sporadic BL was 1.8%. A higher frequency (6%) was found in five endemic cases (three biopsy specimens and two BL cell lines). CONCLUSIONS: The detection of somatic mutations in the rearranged V region genes suggests that both sporadic and endemic BL represent a B-cell malignancy originating from germinal center B cells or their descendants. Interestingly, the mutation frequency detected in sporadic BL is in a range similar to that characteristic for IgM-expressing B cells in the human peripheral blood and for mu chain-expressing germinal center B cells, whereas the mutation frequency found in endemic BL is significantly higher.

B-Lymphocytes↗
PubMed1994

Molecular requirements for rapid plasmacytoma and pre-B lymphoma induction by Abelson murine leukemia virus in myc-transgenic mice.

Mutants and fusion products of the c-abl gene were used to define some of the molecular requirements for rapid plasmacytoma (PC) and pre-B-lymphoma induction in pristane-treated N-myc transgenic BALB/c mice. A-MuLV induced PCs in 21 of 25 mice with a mean post-pristane latency period of 46 +/- 9 days, compared to 134 +/- 25 days in controls exposed to pristane alone. delta XB, a mutant of type IV c-abl with a deletion of the SH3 domain, was equally effective in inducing PCs in 7 of 7 mice with a latency period of 49 +/- 7 days, indicating that gag sequences are not required for rapid PC induction. The delta XB delta Nar mutant that carried a large C-terminal deletion in addition showed only a negligible activity, if any, suggesting that PC acceleration requires the C-terminal domain in the same way as lymphoid transformation and in contrast to fibroblast transformation. BCR-ABL fusion constructs encoding an 185-kDa protein as in acute leukemia, or a 210-kDa protein as in chronic myelocytic leukemia (CML), did not accelerate pristane-induced PC development in the N-myc transgenic mice, in contrast to their known ability to immortalize lymphoid cells in vitro. Only one of 14 non-transgenic littermates developed a pre-B lymphoma after A-MuLV infection, and none of 10 normal littermates infected with delta XB virus developed a construct-carrying tumor. This result suggests that PC acceleration is due to co-operative interaction of the N-myc transgene and activated abl. Infection of N-myc transgenic bone marrow or spleen cells with A-MuLV in vitro led to the outgrowth of pre-B lymphomas after transplantation to pristane-treated BALB/c recipients. The lymphoma-inducing activity of A-MuLV depends on its high titer, since diluted A-MuLV or the lower-titered delta XB induced only PCs under the same conditions. The v-abl, delta XB and BCR-ABL-carrying viruses generated immortalized lymphoblastoid lines in vitro, regardless of the presence of the N-myc transgene, suggesting that lymphoid transformation is a direct function of appropriate abl sequences in contrast to PC acceleration.

Animals↗
PubMed1994

Endogenous TGF-beta contributes to the induction of the EBV lytic cycle in two Burkitt lymphoma cell lines.

A low proportion of cells in the BL lines P3HR-I and Akata enter spontaneously into the EBV lytic cycle, detectable by the expression of early antigens (EA). We found that both lines produce the active and inactive forms of TGF beta. It was shown earlier that a larger number of cells can be induced to enter the lytic cycle by exposing P3HR-I to phorbol esters and n-butyrate and the surface IgG-positive Akata cells to anti-IgG. We now show that the same treatments raise the level of active TGF beta release. Exposure to anti-TGF beta antibodies reduced EA induction by 75-85%. Our results indicate that induction of the viral productive cycle by the above-mentioned reagents is at least partly dependent on the activation of endogenous TGF beta production.

Burkitt Lymphoma↗
PubMed1994

Effect of beta-lymphocyte- and NPC-derived EBV-LMP1 gene expression on in vitro growth and differentiation of human epithelial cells.

The effect of expression of the Epstein-Barr-virus (EBV) latent membrane protein (LMP1) derived from B-lymphocytes (B) and nasopharyngeal carcinoma (NPC) (C) on the in vitro growth and differentiation of a human keratinocyte line, Rhek-1, was analyzed in clonal growth and in in vitro differentiation assays. In contrast to the polygonal parental cells, the B-LMP1-expressing sublines were spindle-shaped while the C-LMP1-expressing cells were pleomorphic. Both B- and C-LMP1-expressing sublines showed increased proliferation as evidenced by: (1) higher colony-forming efficiency (CFE) and larger colony size at reduced serum levels; (2) an increased number of epithelial cell layers formed in the air-liquid-interface culture system and (3) increased expression of proliferative cell nuclear antigen (PCNA). At low serum concentration, the C-LMP1-expressing sublines formed larger colonies than those expressing B-LMP1. In the air-liquid-interface culture system, both B- and C-LMP1-expressing lines showed reduced epithelial differentiation resulting in reduced stratification and reduced involucrin expression similar to those of the cancer cell line, Siha. The results of the present study indicate that the expression of LMP1 in human keratinocytes is associated with morphological transformation and predisposes these cells to a more neoplastic phenotype. The structural difference between the 2 genes responsible for the functional differences and transforming ability will be pinpointed in further experiments.

Antigens, Viral↗
PubMed1994

Sequence-specific methylation inhibits the activity of the Epstein-Barr virus LMP 1 and BCR2 enhancer-promoter regions.

We reported earlier that variable expression of the Epstein-Barr virus (EBV) encoded membrane protein LMP 1 in nasopharyngeal carcinoma and the host-cell-phenotype-dependent activity of the BCR2 promoter (one of the possible initiator sites for transcripts of Epstein-Barr nuclear antigens) in Burkitt's lymphoma (BL) lines can be related to the methylation status of the 5'-flanking regulatory regions of the BNLF 1 and BCR2 promoter, respectively. Here we report that clones of the BL line Mutu that differ in expression of LMP 1 also show a differential methylation pattern of the LMP 1 regulatory sequences: this region is hypomethylated in an LMP 1 expressing (group III) clone but methylated in a group I clone that does not express LMP 1. We introduced in vitro methylated reporter plasmids carrying BNLF 1 and BCR2 enhancer-promoter sequences into the BL line Raji and found that overall methylation of 5'-CG-3' sequences by the Spiroplasma methylase Sssl significantly reduced their activity compared to unmethylated or mock-methylated controls. Methylation of 5-CCGG-3' sequences by Hpall methyltransferase gave similar results. On the contrary, methylation of 5'GCGC-3' sequences by Hhall methyltransferase gave similar results. On the contrary, methylation of 5'-GCGC-3' sequences by Hpal methyltransferase resulted only in a moderate reduction of BNLF 1 enhancer-promoter activity. These data support the notion that methylation at discrete sites within control regions of latent, growth-transformation associated EBV genes may contribute to silencing their expression.

Burkitt Lymphoma↗
PubMed1994

Construction of representative NotI linking libraries specific for the total human genome and for human chromosome 3.

NotI linking clones represent valuable tools for both physical and genetic mapping. Using procedures that we have previously described, several chromosome 3-specific NotI linking libraries have been constructed. Here, we describe the construction of six independent NotI linking libraries specific for the total human genome. These libraries were made using three different vectors and two combinations of restriction enzymes. Altogether, these six libraries contain more than 1 million recombinant phages. Considering that the human genome contains about 3000-5000 NotI sites, it is likely that all clonable NotI sites are present in these libraries. Two of the six libraries were transferred into plasmid form. At the same time, a chromosome 3-specific EcoRI-NotI library (NRL1) was constructed. This library considerably increases the representation of cloned NotI sites in combination with previously constructed libraries that were made using BamHI-NotI digestion. All libraries are available on request.

Blotting, Southern↗
PubMed1994

Three exceptional IgH/myc-translocation-carrying rat immunocytomas have breakpoints 50 to 80 kb 5' of c-myc.

The spontaneously arising immunocytoma of the Louvain rat (RIC) carries a consistent chromosomal translocation between chromosomes 6 and 7. This translocation juxtaposes immunoglobulin heavy chain and c-myc sequences. In an earlier study on 14 RIC tumors, we found that the translocation breakpoint is located within 1.5 kb immediately upstream of c-myc in 10 of the tumors. Here we describe 3 exceptional tumors that had no rearrangement within 20 kb 5' of c-myc. Using pulsed-field gel electrophoresis we show that the translocation breakpoints in these tumors are located 50-80 kb 5' of c-myc and that c-myc rearranges to the 3' end of the IgH cluster.

Animals↗
PubMed1994

p53 binds single-stranded DNA ends and catalyzes DNA renaturation and strand transfer.

The p53 tumor-suppressor protein has previously been shown to bind double-stranded and single-stranded DNA. We report that the p53 protein can bind single-stranded DNA ends and catalyze DNA renaturation and DNA strand transfer. Both a bacterially expressed wild-type p53 protein and a glutathione S-transferase-wild-type p53 fusion protein catalyzed renaturation of different short (25- to 76-nt) complementary single-stranded DNA fragments and promoted strand transfer between short (36-bp) duplex DNA and complementary single-stranded DNA. Mutant p53 fusion proteins carrying amino acid substitutions Glu-213, Ile-237, or Tyr-238, derived from mutant p53 genes of Burkitt lymphomas, failed to catalyze these reactions. Wild-type p53 had significantly higher binding affinity for short (36- to 76-nt) than for longer (> or = 462-nt) single-stranded DNA fragments in an electrophoretic mobility-shift assay. Moreover, electron microscopy showed that p53 preferentially binds single-stranded DNA ends. Binding of DNA ends to p53 oligomers may allow alignment of complementary strands. These findings suggest that p53 may play a direct role in the repair of DNA breaks, including the joining of complementary single-stranded DNA ends.

Base Sequence↗
PubMed1994

Identification of wild-type and mutant p53 peptides binding to HLA-A2 assessed by a peptide loading-deficient cell line assay and a novel major histocompatibility complex class I peptide binding assay.

Mutations of the p53 gene are the most frequently observed genetic changes in human cancers; often leading to an overexpression of the wild-type (wt) p53 protein. Demonstrable T cell reactivity against tumor cells overexpressing wt or mutant p53-derived peptides could support the application of such epitopes in cancer immunotherapies. As the binding of peptide to MHC class I molecules is a prerequisite for antigen-specific T cell recognition, we evaluated the ability of wt and mutant p53 peptides to bind to HLA-A2.1 using two independent flow cytometry-based assay systems, the T2 major histocompatibility complex (MHC) class I peptide stabilization assay (stabilization assay) and the peptide-induced MHC class I reconstitution assay (reconstitution assay). The twenty selected wt sequences each conformed to the previously reported HLA-A2.1 peptide binding motif. Seven of the wt p53 and 2/13 mutant p53 peptides derived from the previously chosen wt peptides bound to HLA-A2.1 in both the stabilization and the reconstitution assays. An additional six wt and six mutant p53 peptides, presumably exhibiting lower affinity for HLA-A2.1, were identified only in the reconstitution assay. Those p53 peptides binding HLA-A2.1 may provide useful immunogens for the generation of HLA-A2.1-restricted cytolytic T lymphocytes in vitro and in vivo.

Amino Acid Sequence↗
PubMed1994

Functional and molecular characterization of B cell-responsive V delta 1+ gamma delta T cells.

Cells expressing the V delta 1+ gene segment are a minor gamma delta T cell population in human peripheral blood but predominate in epithelial and (inflamed) tissues. The characteristic dendritic-like morphology of these gamma delta T cells is consistent with their putative immune surveillance role in epithelia. Their function, however, remains unknown. We and others previously reported that a subset of V delta 1+ gamma delta T cells proliferates after stimulation with Epstein-Barr virus (EBV)-transformed B lymphoblastoid cell lines (LCL), but not with fresh peripheral blood-derived B cells. These responses were independent of the type of T cell receptor (TcR) gamma chain co-expressed with the V delta 1 chain. The in vivo relevance of this LCL-mediated activation as well as the nature of the stimulatory ligand on the LCL is not well established. In this study, we tested the proliferative response of V delta 1+ LCL-responsive T cells against non-EBV-transformed B cells, activated through CD40 by murine EL4 B5 cells, and to a panel of B cell lines differing in the expression of EBV nuclear antigen proteins and adhesion/co-stimulatory molecules. The role of the Epstein-Barr virus-derived antigen in the induction of this response could be excluded as the activated (non-EBV-transformed) peripheral blood B cells were also able to induce a proliferative response in the LCL-responsive V delta 1+ T cells. Therefore, the stimulatory ligand on B cells is of cellular rather than of viral origin, and its expression is up-regulated upon activation of B cells. The expression of B7 and CD39 molecules on the surface of activated B cells appeared to be crucial since antibodies to these structures could block the induction of proliferation of the V delta 1+ T cells. Finally, we investigated the diversity of the responding V delta 1+ gamma delta T cell clones by sequence analysis of the TcR delta junctional regions. No restricted V-D-J sequences were found among the LCL-responsive V delta 1+ T cell clones, arguing strongly against a mono- or oligoclonal V delta 1+ gamma delta T cell response to LCL. These findings may explain the presence of polyclonally activated V delta 1+ T cells in inflamed tissues where activated B cells are often present.

Adenosine Triphosphatases↗