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J Lorenzen

Publications and source records attributed to J Lorenzen.

At least 91 records · Page 5Linked to original sources
PubMed1994

Detection of T-cell clonality in paraffin-embedded tissues.

In lymph node diagnosis, difficulties are frequently encountered with the differential diagnosis of reactive and neoplastic T-cell proliferations. Immunohistochemistry is of limited use, and fresh frozen material for DNA studies is not available in many cases. We have established a polymerase chain reaction (PCR) technique to amplify rearranged T-cell receptor (TCR)-gamma sequences from paraffin-embedded material. Our method differs from other techniques previously described in that it uses four sets of family-specific variable (V)gamma primers. Clonality of the investigated T cells is reflected not only by the varying lengths of amplified products but also by differences in the relative amount of rearranged V gamma families. Preliminary studies indicate that this approach can provide information about the presence of predominant T-cell clones within the sample and thus help to classify the lymph node lesion. With this technique we were able to confirm clonality in seven of 12 T-cell lymphomas in paraffin-embedded tissues.

Adult↗
PubMed1994

[Detection of T-cell receptor gamma chain V gene rearrangements in angioimmunoblastic lymphadenopathy type T-cell lymphoma by PCR and family specific primers].

Using PCR method and family specific primers for T-cell receptor gamma chain V gene, the T-cell colonalities of 12 cases of AILD-TCL were analyzed. The results showed monoclonal DNA amplification patterns of TCR gamma in all 12 cases (13 specimens), whereas the DNA from normal peripheral lymphocytes and 2 samples of atypical lymphoid hyperplasia showed polyclonal rearrangement patterns. Our results suggest that AILD-TCL is a monoclonal proliferative disorder originating from T-cell. The PCR method and family specific primers could successfully amplicate the template DNA from paraffin sections, so it is a sensitive, simple and rapid method for routine hematopathologic studies.

Adult↗
PubMed1994

An unusual pattern of lymph node involvement in mycosis fungoides simulating neoplastic follicles.

We describe an unusual pattern of lymph node involvement in a case of mycosis fungoides. The neoplastic proliferation formed pale nodules composed of small, irregular lymphoid cells, features that, on routine paraffin sections, suggested neoplastic follicles and a B-cell phenotype. The correct diagnosis was established by immunohistochemical studies that demonstrated a T-cell phenotype. Polymerase chain reaction for rearrangement of the T-cell receptor gamma gene confirmed that the proliferation was monoclonal. This case demonstrates that both architectural pattern and cytologic features may be unreliable in predicting immunologic phenotype. Immunologic marker studies and careful attention to the clinical history are vital to classify lymphoproliferative disorders correctly.

Aged↗
PubMed1993

Growth factors: regulation of normal and neoplastic growth.

This paper presents an overview of current trends in growth factor research. The first part of the review considers the current classification of growth factors and their receptors. A model of cell proliferation regulation by growth factors is then presented. The final section reviews the latest concepts of the involvement of growth factors in the development of neoplasia.

Cell Division↗
PubMed1993

Demonstration of light chain mRNA in Hodgkin's disease.

The lineage of Hodgkin and Reed-Sternberg cells is still unclear. Detection of both immunoglobulin light chains in Hodgkin and Reed-Sternberg cells by immunohistochemistry is a well-known phenomenon. However, up to now, in situ hybridization techniques have failed to demonstrate light chain messenger(m) RNA in Hodgkin and Reed-Sternberg cells. In this investigation, we have analysed 26 cases of Hodgkin's disease (nodular lymphocyte predominant Hodgkin's disease, mixed cellularity, and nodular sclerosis type) using digoxigenin-labelled oligonucleotide probes for kappa and lambda light chains by in situ hybridization. In nearly half of the cases of nodular lymphocyte predominant Hodgkin's disease and in one case of mixed cellularity type, mRNA for only one light chain could be clearly demonstrated in the lymphocytic and histiocytic cells, Hodgkin, and Reed-Sternberg cells. These results support the idea that at least some cases of Hodgkin's disease are B-cell neoplasms.

Hodgkin Disease↗
PubMed1993

Saturated and unsaturated repair of DNA strand breaks in CHO cells after X-irradiation with doses ranging from 3 to 90 Gy.

The kinetics of DNA strand break repair was studied in exponentially-growing CHO cells after X-irradiation with doses of 3, 9, 30, 60 and 90 Gy. DNA strand breaks were measured using the alkaline unwinding technique. For all X-ray doses applied the kinetics of DNA strand break repair consisted of fast, intermediate and slow phases. The latter, which was interpreted as the repair kinetic of DNA double-strand breaks, was best described by an exponential decline. The actual repair half-time of double-strand break repair, tau dsb, was obtained from the slope of the slow component after subtracting the number of non-reparable breaks measured 24 h after irradiation. This half-time was found to be independent of the dose applied with a mean value of tau dsb = 168 +/- 10 min. This result indicated that the repair of double-strand breaks was unsaturated for doses up to 90 Gy. The repair kinetics of the breaks of the fast and intermediate phases were found to be dependent on the dose applied. These kinetics were associated with the induction and repair of primary and secondary single-strand breaks, the latter possibly generated by enzymatic incision at damaged bases. Analysis of these curves using the Michaelis-Menten equation showed that the half-time of enzymatic incision, tau in, and the half-time at which both primary and secondary single-strand breaks were rejoined, tau rep, varied with the amount of damage present in the cell. tau in Increased from a minimum value tau in,min = 13 +/- 2 min proportionally to the number of base damage with a rate of alpha in = 0.138 +/- 0.015 min Gy-1, and tau rep from a minimum value tau rep,min = 1.4 +/- 0.2 proportionally to the number of single-strand breaks with a rate of alpha rep = 0.038 +/- 0.001 min Gy-1. The enzymatic incision was unsaturated for doses up to about 30 Gy, whereas the repair of single-strand breaks was unsaturated only for doses up to about 10 Gy. Up to these doses the increase in the half-time tau in and tau rep was so small that, within the range of experimental errors, the parameters may be approximated by constant values. From the results it is concluded that for CHO cells the continuously-bending dose-response curve obtained for radiation-induced killing cannot be attributed to a saturated repair.

Animals↗
PubMed1993

Expression of the proliferating cell nuclear antigen in the different types of Hodgkin's disease.

Thirty-eight cases of Hodgkin's disease (HD, lymphocyte-predominant, n = 10; nodular sclerosis, n = 10; mixed cellularity, n = 10; lymphocyte depletion, n = 8) were investigated with the antibody PC10 directed against the proliferating cell nuclear antigen (PCNA) with B- and T-cell markers using a double-staining technique in paraffin-embedded material. It could be shown that nearly all (95-97%) Hodgkin's and Reed-Sternberg (HRS) cells and their variants were PCNA-positive regardless of the type of HD. There was only a low number of PCNA-positive lymphocytes (2.8-3.4%) in all types mostly consisting of MT1-positive T lymphocytes. In contrast to the other types, lymphocyte-predominant type showed a relatively high percentage (5%) of Leu-7-positive lymphocytes. The high percentage of PCNA-positive HRS cells correlates with their malignant nature, and might be another example of dysregulated expression of PCNA.

Antigens, Differentiation↗
PubMed1992

Expression of the bcl-2 oncogene product and chromosomal translocation t(14;18) in Hodgkin's disease.

The B-cell lymphoma/leukemia oncogene bcl-2 takes part in crucial regulatory events in B-cell maturation and differentiation. The reciprocal chromosomal translocation t(14;18), leading to overexpression of this oncogene, can be found in the majority of follicular lymphomas and much less frequently in B-cell leukemias and diffuse lymphomas. We have studied the expression of this protein in different subtypes of Hodgkin's disease using monoclonal antibodies directed against a formalin-resistant epitope of the bcl-2 protein and also have investigated these cases by polymerase chain reaction for evidence of the t(14;18) translocation. We were particularly interested to determine whether nodular paragranuloma (lymphocyte-predominant, nodular), which differs from other subtypes of Hodgkin's disease by virtue of the B-cell nature of its malignant cell population, is characterized by expression of the bcl-2 protein. Our data indicate that only a small number of nodular paragranulomas express the bcl-2 protein and that the expression is not specific for this type of Hodgkin's disease. In a smaller number of cases this expression of bcl-2 could be explained by the presence of the translocation t(14;18).

Antibodies, Monoclonal↗
PubMed1992

A Rhizobium meliloti lipopolysaccharide mutant altered in competitiveness for nodulation of alfalfa.

A transposon Tn5-induced mutant of Rhizobium meliloti Rm2011, designated Rm6963, showed a rough colony morphology on rich and minimal media and an altered lipopolysaccharide (LPS). Major differences from the wild-type LPS were observed in (i) hexose and 2-keto-3-deoxyoctonate elution profiles of crude phenol extracts chromatographed in Sepharose CL-4B, (ii) silver-stained sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis patterns of crude and purified LPS fractions, and (iii) immunoreactivities otherwise present in purified LPS of the parental strain Rm2011. In addition, Rm6963 lost the ability to grow in Luria-Bertani medium containing the hydrophobic compounds sodium deoxycholate or SDS and showed a decrease in survival in TY medium supplemented with high calcium concentrations. The mutant also had altered symbiotic properties. Rm6963 formed nodules that fixed nitrogen but showed a delayed or even reduced ability to nodulate the primary root of alfalfa without showing changes in the position of nodule distribution profiles along the roots. Furthermore, 2 to 3 weeks after inoculation, plants nodulated by Rm6963 were smaller than control plants inoculated with wild-type bacteria in correlation with a transient decrease in nitrogen fixation. In most experiments, the plants recovered later by expressing a full nitrogen-fixing phenotype and developing an abnormally high number of small nodules in lateral roots after 1 month. Rm6963 was also deficient in the ability to compete for nodulation. In coinoculation experiments with equal bacterial numbers of both mutant and wild-type rhizobia, only the parent was recovered from the uppermost root nodules. A strain ratio of approximately 100 to 1 favoring the mutant was necessary to obtain an equal ratio (1:1) of nodule occupancy. These results show that alterations in Rm6963 which include LPS changes lead to an altered symbiotic phenotype during the association with alfalfa that affects the timing of nodule emergence, the progress of nitrogen fixation, and the strain competitiveness for nodulation.

Calcium↗
PubMed1992

[Which are the proliferating cells in Hodgkin's disease?].

This investigation characterizes the proliferating cells in Hodgkin's disease. We used the antibody PC 10 which reacts with the proliferating cell nuclear antigen (PCNA) and works on paraffin sections in combination with B- and T-cell markers in a double-staining technique. In all 38 cases of Hodgkin's disease (lymphocyte predominant type n = 10, nodular sclerosis type n = 10, mixed cellularity type n = 10, lymphocyte depletion type n = 8) a high percentage of the Hodgkin and Sternberg-Reed cells (95%-97%) expressed PCNA. There was no statistical significance between the different types. In contrast, only a small amount of the reactive lymphocytes (2.8%-3.4%) demonstrated positivity for PCNA in the four types of Hodgkin's disease. Nearly all of these lymphocytes were T-lymphocytes.

Antigens, CD↗
PubMed1992

[Bcl-2 in potentials precursors of nodular paragranuloma and its dedifferentiated variant (large cell B-lymphoma)].

We have investigated seven cases of large cell lymphomas (LCL) developing simultaneously or metachronously to nodular lymphocyte-predominant Hodgkin's disease (nodular paragranuloma, NP) for the presence of EBV and the chromosomal translocation t(14;18) by use of the polymerase chain reaction (PCR). The expression of the bcl-2 oncogene product in these cases and in five cases of progressive transformation of germinal centres as a potential precursor of NP was detected immunohistochemically with the monoclonal antibodies bcl-2-100 and bcl-2-124. All cases investigated were negative for EBV genomic material. The chromosomal translocation t(14;18) was also absent. Expression of the bcl-2 oncogene could be detected only in one case of nodular paragranuloma and in an unrelated case of LCL. Hence, LCL developing out of NP differ from other germinal center derived high-grade lymphomas.

Adult↗
PubMed1992

[Detection of clonal immunoglobulin rearrangements in paraffin embedded tissues by PCR].

A polymerase chain reaction based method was established using immunoglobulin VH and VK gene rearrangements as markers to detect clonal B cell populations in paraffin embedded sections. Family specific VH and VK primers are used in separate reactions together with the corresponding J primers to amplify rearranged VH and VK genes from genomic DNA. This allows to distinguish clonal from polyclonal B cell populations in most of the cases. The method may be helpful in routine diagnosis of B cell NHL and some morphological and immunohistochemical difficult cases.

Base Sequence↗
PubMed1991

Activation mechanism of rabbit skeletal muscle myosin light chain kinase. 5'-p-fluorosulfonylbenzoyl adenosine as a probe of the MgATP-binding site of the calmodulin-bound and calmodulin-free enzyme.

5'-p-fluorosulfonylbenzoyl adenosine (FSBA), an ATP-like affinity labelling reagent, reacted with rabbit skeletal muscle myosin light chain kinase (skMLCK) and its calmodulin complex in a site-specific manner. Reaction was dependent upon the presence of the adenosine moiety of FSBA, saturated with increasing FSBA, was inhibited by MgATP, and was accompanied by stoichiometric incorporation of [14C]FSBA. The kinetic constants describing the reaction were similar for skMLCK and its calmodulin complex: k3 = -0.040 min-1 and -0.038 min-1, and Ki = 0.18 mM and 0.40 mM, respectively. It is concluded that the MgATP-binding site on skMLCK remains accessible at all times and maintains a near constant conformation.

Adenosine↗
PubMed1991

Basic fibroblast growth factor and interleukins 4 and 6 stimulate the release of IFN-gamma by individual NK cells.

Both the secretory and cytotoxic activity of natural killer (NK) cells are known to be regulated by such cytokines as interleukin-2 (IL-2) and interferon-gamma (IFN-gamma). In the present study we have used the reverse hemolytic plaque assay to investigate either the direct effects of the protein kinase activator, phorbol myristate acetate (PMA), or exposure to recombinant human interleukins 2, 4, and 6 (IL-2, IL-4, and IL-6) tumour necrosis factor alpha (TNF-alpha) and basic fibroblast growth factor (bFGF) on the release of IFN-gamma by individual, immunoidentified NK cells isolated from peripheral blood. This sensitive immunoassay was adapted and coupled with immunocytochemistry not only to immunophenotype and enumerate cells secreting IFN-gamma in a given cell population, but also to quantify the amount of this cytokine released per individual cell. These studies have confirmed mononuclear cells with the morphology of large granular lymphocytes and the immunophenotype of CD3-/CD16+ NK cells to be the predominant source of spontaneously released IFN-gamma in vitro. In contrast to this, fewer than 2% of the CD3+ T cells secreted detectable levels of this cytokine during the assay, irrespective of the stimulus applied. Whilst TNF-alpha had no significant effect on IFN-gamma release by NK cells, a 6-hr exposure to IL-2 or PMA stimulated an increase in the amount secreted per single cell. Furthermore, bFGF and interleukins 4 and 6 elicited a marked, dose-dependent stimulation of IFN-gamma secretion by this cell type. However, exposure to these cytokines did not alter the number of cells capable of releasing detectable levels of IFN-gamma during the assay. These studies demonstrate that (i) both the spontaneous and stimulated release of IFN-gamma by NK cells can be visualized and quantified at the single-cell level using this sensitive immunoassay, and (ii) bFGF and interleukins 2, 4, and 6, but not TNF-alpha, are potent stimulants of IFN-gamma secretion by CD3-/CD16+ NK cells.

Antigens, CD↗
PubMed1991

Human tumour-associated NK cells secrete increased amounts of interferon-gamma and interleukin-4.

Numerous interactions between malignant and stromal/inflammatory cells take place within solid human tumours, which are mediated, in part, by the release of signalling proteins called cytokines. In the present study, we have compared the secretion of two important immunomodulatory cytokines, IFN-gamma and IL-4 by individual, immunophenotyped NK cells freshly isolated from either malignant tumour biopsies, or peripheral blood samples from patients with ductal invasive breast cancer. Due to the marked heterogeneity amongst cells isolated from these clinical samples, we have employed a technique called the reverse haemolytic plaque assay to identify and enumerate cytokine-secreting cells at the single cell level. Our data indicate that NK cells isolated directly from the tumour site secrete more IFN-gamma and IL-4 than NK cells from the blood of the same patients. However, a greater proportion of CD16+ cells from both sources in breast cancer patients secreted IFN-gamma than of those from the blood of healthy donors. We also show that factors secreted by the human breast cell lines, MCF-7 and MDA-231 PN9, were able to mimic the stimulatory influence of the tumour microenvironment on secretory activity of NK cells.

Antigens, CD↗
PubMed1990

Measurement of cytokine release by human cells. A quantitative analysis at the single cell level using the reverse haemolytic plaque assay.

The reverse haemolytic plaque assay has been adapted to detect and measure the release of such cytokines as interleukin-1, -2 and -6, GM colony-stimulating factor or interferon-gamma by individual human cells derived from either peripheral blood or enzymatically dispersed breast carcinomas. Since each of these peptides is released by more than one cell type, this in vitro assay has been coupled with immunocytochemistry to identify the particular cell type(s) contributing to the release of each cytokine. This technique is useful in (i) obviating the need for purification of a given cell type prior to estimating cytokine release, and (ii) evaluating quantitative differences in secretion amongst cells of a particular type. Such a method has the additional advantage over most alternative methods applied at the single cell level in that the cells remain viable at the end of the assay and can be used in further studies. This assay thus provides a powerful new tool in the investigation of the role of cytokines in both the normal modulation of the immune system and the development of such diseases as neoplasia.

Biological Factors↗