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R Kiessling

Publications and source records attributed to R Kiessling.

At least 73 records · Page 4Linked to original sources
PubMed1995

Analysis of gamma delta V region usage in normal and diseased human intestinal biopsies and peripheral blood by polymerase chain reaction (PCR) and flow cytometry.

The intestinal population of gamma delta T cell receptor (TCR)-bearing cells was characterized with regard to V delta and V gamma subtype expression. For this purpose, we utilized V gene-specific PCR of mRNA prepared from intestinal biopsies. Predominant expression of the V delta 1 subtype was demonstrated in the small intestine of patients with coeliac disease and in the inflamed colon of patients with inflammatory bowel diseases (IBD: ulcerative colitis and Crohn's disease) as well as in colon biopsies taken from macroscopically normal areas of colon. Although intestinal gamma delta T cells preferentially expressed V delta 1, other V delta transcripts could be detected, of which V delta 2 and V delta 5 were commonly expressed. Analysis of biopsies from mesenteric lymph nodes demonstrated a V delta repertoire similar to the mucosa. In peripheral blood on the other hand, high expression of both V delta 2 and V delta 1 was found. The predominant expression of V delta 1 transcripts in the intestinal mucosa of IBD patients correlated well with protein cell surface expression as analysed by flow cytometry using V delta 1- and V delta 2-specific antibodies. Selective expansion of gamma delta T cells could not be demonstrated within the inflamed mucosa as shown by mRNA analysis and flow cytometry. Instead, IBD patients demonstrated a decreased proportion of TCR gamma delta-carrying T cells in the inflamed mucosa compared with macroscopically normal area of colon. On the other hand, a significantly increased percentage of T cells bearing the gamma delta TCR was found in peripheral blood of patients with Crohn's disease compared with healthy individuals, indicating that local mucosal inflammation may influence the circulating gamma delta T cell population.

Base Sequence↗
PubMed1995

Engagement of MHC class I proteins on natural killer cells inhibits their killing capacity.

We have studied whether engagement of MHC class I (MHC-I) molecules on natural killer (NK) cells can influence the NK killing activity. Human NK effector cells, enriched by nylon wool passage, were incubated with monoclonal antibodies (MoAb) to MHC-I followed by cross-linking with secondary rabbit anti mouse Ig or streptavidin. Cross linking of MHC-I molecules on NK cells resulted in a clear inhibition of the NK activity against the target cells K562, Molt-4 and U937. The inhibitory effect was selective for MHC-I and was not seen with MoAb to MHC-II or CD56 molecules. The inhibition was not mediated via Fc receptors since F(ab)2 fragments of the MHC-I MoAb W6/32 were as effective as the intact antibody. The best inhibition of NK activity was obtained using biotin-labelled F(ab)2 fragments of W6/32 and streptavidin as a cross-linker, where up to 70% reduction in NK cell activity was observed. Antibody dependent cellular cytotoxicity (ADCC) was also inhibited by cross-linking MHC-I molecules on the effector cells. The results show that antibody mediated cross-linking of MHC-I proteins on NK cells can inhibit their killing capacity. This indicates that MHC-I molecules on NK cells can be involved in the regulation of NK cytotoxicity, perhaps by transmitting inhibitory signals into the NK cell.

Antibodies, Monoclonal↗
PubMed1994

Interleukin 10 pretreatment protects target cells from tumor- and allo-specific cytotoxic T cells and downregulates HLA class I expression.

Interleukin 10 (IL-10) is a cytokine with a variety of reported effects including inhibition of monocyte major histocompatibility complex (MHC) class II-dependent antigen presentation, type 1 helper T cell cytokine production, and inhibition of T cell proliferation. Herein we report the effect of IL-10 pretreatment on antigen presentation to tumor- and allo-specific CD8+ cytotoxic T lymphocytes (CTL). Prior incubation of human melanoma cells with recombinant IL-10 (rIL-10) for 48-72 h resulted in a dose-dependent, up to 100% inhibition, of autologous CTL-mediated, HLA-A2.1-restricted, tumor-specific lysis. Allo-specific CTL cytotoxicity against Epstein-Barr virus-transformed lymphoblastoid cell lines (LCL) was also inhibited, demonstrating a protective effect also on lymphoid cells. In contrast, IL-10 pretreatment of allogeneic LCL or K562 targets had either no effect or slightly enhanced cytotoxic activity mediated by freshly isolated or IL-2-activated natural killer cells. Flow cytometric analysis with monoclonal antibodies against HLA-A2, or nonpolymorphic determinants of MHC class I proteins, revealed a 20-50% reduction in cell-surface expression, whereas intercellular adhesion molecules 1, and 2, and lymphocyte function-associated antigen 3 levels were not affected. In addition, relative to untreated target cells, IL-10 pretreated tumor cells were unaltered in their capacity to affect CTL-mediated lysis by cold target inhibition, demonstrating that the effect of IL-10 is unrelated to the initial binding of CTL to their targets. These results are compatible with an effect of IL-10 on the MHC class I antigen presentation pathway, and suggest a novel mechanism of immune tolerance, based on escape from CTL-mediated tumor and allo-transplant rejection.

Antibodies, Monoclonal↗
PubMed1994

The Epstein-Barr virus latent membrane protein-1 (LMP1) induces interleukin-10 production in Burkitt lymphoma lines.

Human interleukin-10 (h-IL-10) is a pleiotropic cytokine with stimulatory activity on B-lymphocytes. Recent evidence indicates that infection with Epstein-Barr virus (EBV) induces h-IL-10 production in B-cells and that this cytokine may contribute to EBV-induced B-cell transformation. It is not known whether h-IL-10 induction by EBV correlates with distinct phenotypic features of the infected cells or with the expression of particular viral genes. We have approached these questions by investigating the expression of h-IL-10 mRNA in a panel of B-cell lines including: in vitro EBV-transformed lymphoblastoid cell lines (LCLs), EBV-carrying Burkitt lymphoma (BL) lines, EBV-negative BL lines and their sublines infected with different EBV strains, or transfected with the transformation-associated viral gene. h-IL-10 mRNA was detected by reverse-transcriptase-assisted (RT)-PCR in a subset of EBV-negative BLs and in all EBV-positive BL lines and LCLs investigated except Daudi. This cell line carries an EBV nuclear antigen (EBNA)-2 gene-defective virus strain. h-IL-10 mRNA was induced by conversion of 3 EBV-negative and h-IL-10-negative BL lines (BL41, BL47 and BL49) with the transforming, B95.8-derived EBV strain. P3HR-I virus convertants that do not express the viral EBNA-2 and the EBV latent membrane protein (LMP)-1, and fail to progress towards a LCL-like cell phenotype, showed no evidence of h-IL-10 up-regulation. Expression of LMP1 was sufficient to induce h-IL-10 mRNA in transfected sublines of the EBV-negative DG75 and BL41 cell lines, whereas expression of EBNA1, 2, 5, or 6 had no effect. h-IL-10 was detected in the culture supernatants of the LMP1 transfectants by specific ELISA assays. The present findings confirm the role of LMP1 in the transactivation of a wide variety of cellular genes which may be involved in EBV-induced B-cell transformation.

Antigens, Viral↗
PubMed1993

Decreased expression of the signal-transducing zeta chains in tumor-infiltrating T-cells and NK cells of patients with colorectal carcinoma.

An impaired immune response is frequently observed in cancer patients and tumor-bearing mice. T-cells from mice with an experimental colon carcinoma were recently shown to express T-cell receptors that completely lacked the signal-transducing molecule CD3 zeta. Here, we have investigated the expression of the signal-transducing molecule zeta on lymphocytes from 14 patients with colorectal carcinomas using flow cytometric analysis of permeabilized cells with a monoclonal antibody (TIA-2; IgG1) specific for the cytoplasmic domain of the zeta chain as well as with immunoprecipitation and analysis on diagonal gel electrophoresis. We demonstrate that T-cells isolated from the tumors of the patients express significantly less CD3 zeta than T-cells in the peripheral blood of the same patients and that the peripheral blood of the patients express decreased levels of zeta chains, as compared to the levels found in lymphocytes from healthy controls. This decreased expression was also observed on zeta chains associated with the low affinity Fc receptor for IgG found in tumor-infiltrating NK cells (Fc gamma RIIIA alpha; CD16).

Colorectal Neoplasms↗
PubMed1993

Immunization against tumor and minor histocompatibility antigens by eluted cellular peptides loaded on antigen processing defective cells.

Material eluted from RMA lymphoma or B6 spleen cells under acid conditions was fractionated by reverse phase high-performance liquid chromatography, and tested for ability to restore the sensitivity to cytotoxic T lymphocytes of the processing/presentation defective mutant line RMA-S. This allowed identification of three fractions (termed M1, M2 and M3) carrying B6 antigens recognized by cytotoxic T lymphocytes (CTL) elicited across the minor histocompatibility barrier A.BY anti-B6 (both H-2b) and one fraction (termed T1) carrying a tumor antigen recognized by B6 anti-RMA CTL. By parallel runs of material from cell lysates over major histocompatibility complex class I affinity columns, the M2 and M3 antigens were defined as Kb restricted, and M1 and T1 as Db restricted. Isolated fractions loaded onto RMA-S cells could be used to prime anti-minor histocompatibility antigen and tumor CTL in vivo. They could also be used for in vitro restimulation of spleen cells from mice that had been primed either by antigen-loaded RMA-S, or by wild-type RMA tumor cells and B6 splenocytes. The CTL generated by these methods were specific for the loading antigen, and they also recognized the antigen on the "physiological" target, i.e. RMA or B6 lymphoblasts. This system based on RMA-S as an immunization and target antigen reporter cell may be used for dissection of complex CTL responses, e.g. in studies of clonal composition and epitope dominance, or for studies of tumors that are poor stimulators of immunity.

Animals↗
PubMed1993

Cytotoxic activity and T cell receptor repertoire in tumor-infiltrating lymphocytes of adrenal cell carcinomas.

The cytotoxic activity and T cell receptor (TCR) V beta repertoire in tumor-infiltrating lymphocytes (TIL) of three primary adrenal cell carcinomas were analyzed. Fresh, non-cultured TIL from two of the three tumors showed low but significant lysis of the autologous tumor, and for one of the patients this activity was strongly enhanced upon culture in interleukin-2. An allogeneic adrenal cell carcinoma line and the K562 or Daudi targets included as controls were not killed. Phenotypic analysis of freshly isolated TIL demonstrated that the cells from the two patients that demonstrated cytolytic capacity mainly consisted of CD45RO+ T cells. In vitro cultured TIL lines from these patients demonstrated a high percentage of CD8+ cells expressing either the V beta 6 gene or the V beta 8 gene product, as measured with a panel of mAb specific for TCR V alpha and V beta gene products. Analysis of the TCR V beta gene mRNA expression in freshly isolated non-cultured TIL, using a polymerase-chain-reaction-assisted cDNA-amplification assay, confirmed the strong expression of the genes coding for the TCR V beta 6 or the V beta 8. This assay also demonstrated a more restricted TCR V beta gene usage in the TIL as compared to peripheral blood lymphocytes from the same patient.

Adrenal Gland Neoplasms↗
PubMed1993

Restricted T cell receptor V-beta and J-beta usage in T cells from interleukin-2-cultured lymphocytes of ovarian and renal carcinomas.

Tumour-infiltrating lymphocytes (TIL) are often observed in human tumours and their presence has been correlated with a better prognosis. It has been suggested that TIL are enriched for tumour-specific cytotoxic cells, and TIL activated and expanded in vitro by interleukin-2 (IL-2) are currently used in the therapy of human cancer. We have studied the T cell repertoire in IL-2-expanded TIL cells from patients with ovarian and renal carcinoma using T-cell-receptor-V-beta-specific monoclonal antibodies and a polymerase-chain-reaction-based Southern blot technique for analysis of J-beta usage. In TIL lines derived from three of nine patients with ovarian carcinomas and from two of eight patients with renal carcinomas, selective usage of the V-beta 6 or V-beta 5 T-cell receptor gene products was found. The majority of the cells were CD4+, with up to 40% of the T cells utilizing the same V-beta gene. T-cell lines derived from peripheral blood lymphocytes from patients or healthy donors contained normal levels of V-beta subsets. Only moderate levels of V-beta 6+ T cells were detected from freshly isolated TIL and the increase of this subpopulation appeared as a result of in vitro culture. The level of clonal restriction, as measured by the usage of J-beta gene segments within the V-beta 5 or V-beta 6 families, was analysed using a recently developed technique based on the polymerase chain reaction. Evidence for restricted J-beta usage was detected only in TIL expanded in vitro, while this was not the case in freshly isolated tumour-derived lymphocytes or T cell lines obtained from peripheral blood lymphocytes. The presence of a population with biased T cell receptor expression in cells derived from tumour tissue could be explained by their activation in vivo as a result of contact with tumour antigens and should be taken into consideration when discussing the therapeutic efficiency of IL-2-expanded TIL.

Antibodies, Monoclonal↗
PubMed1992

Selective expression of interleukin 10, interferon gamma, and granulocyte-macrophage colony-stimulating factor in ovarian cancer biopsies.

The variable clinical response seen with most cancer immunotherapy suggests that there is a large interindividual variation in immunologic response to tumors. One of the key functional parameters of an immune response is the local production of cytokines. As a method to survey the immune status of tumor-infiltrating cells, we have investigated the constitutive expression of cytokine mRNA in biopsies from epithelial ovarian carcinomas by using a PCR-assisted mRNA amplification assay. Using a set of cytokine-specific primers for 10 different cytokines, we have found selective expression of interleukin 10 (IL-10), granulocyte-macrophage colony-stimulating factor, and interferon gamma mRNA in ovarian tumor tissue as compared to normal ovaries and ovarian tumor cell lines. Such differences could not be explained by the extent of T-cell infiltration, since comparing samples with the same intensity of T-cell receptor (TCR) constant region alpha-chain product from the tumor and normal biopsies demonstrated different cytokine patterns. No IL-2 gene expression was detected in the tumor biopsies. IL-2 mRNA, however, became expressed after stimulation of the tumor-derived cells via the CD3 molecule but not after growth in recombinant IL-2 alone. Using the same methodology, we also analyzed the TCR variable region beta-chain gene repertoire. No restriction or biased expression of these genes was observed.

Base Sequence↗
PubMed1992

T cell receptor diversity and activation markers in the V delta 1 subset of rheumatoid synovial fluid and peripheral blood T lymphocytes.

In the present study we have characterized the gamma/delta T cell receptor (TcR) population in synovial fluid (SF) and peripheral blood (PB) of patients with chronic inflammatory arthritis. By double staining we have shown that (a) synovial V delta 1+ cells have a high expression of activation markers CD45R0 ("memory cells") and HLA-DR as compared to PB, indicating a preactivated population of V delta 1-carrying T cells in vivo and (b) interleukin 2-induced expansion of synovial cells yields a high proportion of gamma/delta in most samples expressing predominantly the V delta 1 TcR. Junctional sequence analysis of the TcR delta chain from interleukin 2-expanded PB cell lines demonstrated a polyclonal V delta 1 population in three out of three samples. In SF cell lines three out of four samples were polyclonally expanded. In SF from one patient, however, a limited repertoire of expressed V delta 1 genes was found. Altogether, our data demonstrate the presence of preactivated V delta 1-expressing cells in the synovial compartment. This V delta 1 population is predominantly polyclonal, except in one patient where oligoclonally expanded V delta 1 cells were detected.

Antigens, CD↗
PubMed1992

Integrins and other adhesion molecules on lymphocytes from synovial fluid and peripheral blood of rheumatoid arthritis patients.

Cell and matrix adhesion of lymphocytes participates in homing, migration and accumulation of these cells in inflamed tissues as well as in the generation of immune and inflammatory responses. In inflamed joints of rheumatoid arthritis (RA) patients, lymphocytes accumulate in the synovial membrane and the synovial fluid. In the present study we have analyzed the expression of integrins and other adhesion molecules in synovial fluid lymphocytes (RA-SFL) and paired peripheral blood lymphocytes (RA-PBL) from 21 RA patients by immunofluorescence flow cytometry. We have also investigated the expression of these adhesion molecules on peripheral blood lymphocytes obtained from 13 sex- and age-matched healthy controls (CO-PBL). RA-SFL, which consisted mostly of T cells, showed higher expression of the integrin subunits beta 1 (CD29), VLA-1 alpha, -3 alpha, -4 alpha, -5 alpha and -6 alpha when compared to RA-PBL. In turn, RA-PBL showed lower expression of these molecules than CO-PBL. The expression of the immunoglobulin-related molecules CD2, CD54 (ICAM-1) and CD58 (LFA-3) was higher on RA-SFL when compared to RA-PBL or CO-PBL, and similar results were obtained with the beta 2 integrin subunits CD11a and CD18. In contrast, L-selectin (LECAM-1) and ICAM-2 were expressed at much lower levels on RA-SFL than on RA-PBL or CO-PBL. CD44, a receptor for hyaluronic acid and collagen, was expressed by most RA-SFL, RA-PBL and CO-PBL cells but at higher density on RA-SFL. The results indicate that RA-SFL express a distinct array of adhesion molecules, similar to the one of memory T lymphocytes. This characteristic phenotype may contribute to the lymphocytic infiltration of the synovium and to the pathogenesis of RA.

Adult↗
PubMed1992

Diverse effect of cytokine treatment of tumor cells on specific versus non-specific cytotoxicity.

The effect of IFN-gamma and TNF-alpha treatment of an ovarian carcinoma line on the sensitivity to lysis by specific CTL clones and non-specific Tumor Associated Lymphocytes (TAL), isolated from the ascites fluid, was analyzed. The in vitro established TAL line displayed a non-specific lytic activity against the autologous tumor as well as against several allogeneic tumor lines. Pretreatment with IFN-gamma alone, or in combination with TNF-alpha, rendered the carcinoma line less susceptible to lysis by the autologous TAL line. Conversely, susceptibility to lysis by tumor specific T cell clones, isolated from the TAL line, was increased as a result of cytokine pretreatment. Several TCR-alpha/beta+, CD8+ T-cell clones showing a more specific pattern of lysis against the autologous tumor were isolated. Lysis of the autologous tumor by these clones involved the TCR-alpha/beta via a MHC-class I restricted mechanism dependent on the adhesion molecules ICAM-1 and LFA-3, as inferred from antibody blocking studies. The enhanced sensitivity to specific CTL clones seen after cytokine treatment may be related to the enhanced expression of ICAM-1 molecules on the ovarian carcinoma. These results have implications for cytokine based immunotherapy, where IFN-gamma may enhance the effects of tumor associated specific CTL while decreasing that of non-specific effector cells.

Ascitic Fluid↗
PubMed1992

Induction of human hsp60 expression in monocytic cell lines.

Both bacterial and mammalian heat shock proteins (HSP) are recognized by some T cells, and hsp60 recognition has been implicated in rheumatoid arthritis. We have developed a model to study the induction of hsp60 in human monocytic cell lines. An anti-mycobacterial hsp65 mAb (ML30), cross-reacting with human hsp60 was used to screen 21 human tumor cell lines in Western blot analysis. All T cell and B cell lymphomas constitutively expressed hsp60 protein at moderate to high levels, while little or no hsp60 protein was detected in two monocytic leukemia lines. Moderate to high levels of hsp60 mRNA and protein could be induced in the THP-I monocytic leukemia cell line by heat shock, retinoic acid, interferon (IFN)-gamma or tumor necrosis factor (TNF)-alpha treatment, the highest levels obtained with a combination of IFN-gamma/TNF-alpha. This was also seen using two rabbit anti-hsp60 antisera directed against the N-terminal or C-terminal part of the human hsp60 protein. The determinants detected by the ML30 mAb or the two rabbit anti-hsp60 antisera were not cell surface expressed, as measured with immunofluorescence (FACS) analysis on control cultured or cytokine treated cell lines. This could be a useful model for studies related to the induction of hsp60 in human cells.

Cytokines↗
PubMed1992

Soluble CD4 suppress the antigen driven proliferative response of certain T cell clones specific for mycobacteria and for peptides by mycobacterial heat shock proteins.

We have investigated the effect of soluble recombinant CD4 (sCD4) on the antigen specific (BCG, peptides of mycobacterial 65 kDa hsp) responses of T cell lines of T cell clones. The majority of the antigen specific clones could be suppressed in their antigen driven response by the addition of sCD4, while others, including the parental polyclonal T cell line, were not. The suppression of the specific T cell response was reversed by the addition of anti-CD3, did not affect the proliferative response to IL-2, and was independent of the amount of antigen. A decreased capacity to produce IFN-gamma in response to the antigen by the addition of sCD4 was seen only with those clones that were also inhibited in their specific proliferative response. This model may be used to delineate further the interaction between T cells and the antigen presenting cell, and the finding may limit the possible in vivo use of sCD4 in the therapy of human immunodeficiency virus (HIV) infections.

Bacterial Proteins↗
PubMed1991

Increased susceptibility of IFN-gamma-treated neuroblastoma cells to lysis by lymphokine-activated killer cells: participation of ICAM-1 induction on target cells.

We have investigated the effect of interferon-gamma (IFN-gamma) treatment of neuroblastoma cells on the susceptibility to lysis by lymphokine-activated killer (LAK) cells and examined the participation of cell-adhesion molecules on the target cells in LAK cell lysis. Untreated neuroblastoma cells expressed lymphocyte-function-associated antigen 3 (LFA-3) and neural-cell-adhesion molecule (NCAM), but did not express MHC-class-I, MHC-class-II, or intercellular-adhesion molecule I (ICAM-I). IFN-gamma treatment of neuroblastoma cells induced the expression of MHC-class-I and ICAM-I antigens, but did not affect the expression of MHC-class-II, LFA-3, and NCAM. This was accompanied by an increased susceptibility to lysis by LAK cells. Anti-ICAM-I antibody inhibited partially the increased sensitivity of IFN-gamma-treated neuroblastoma cells to LAK cell lysis, and blocked completely the increase in binding of LAK cells observed after IFN-gamma treatment of the target cells. These results suggest that the increased LAK sensitivity of IFN-gamma-treated neuroblastoma cells is partially attributable to the induction of ICAM-I on neuroblastoma cells and indicate that post-binding events also play a role in the increased sensitivity to LAK cell lysis observed after IFN-gamma treatment.

Cell Adhesion Molecules↗
PubMed1991

In vivo administration of low-dose human interleukin-2 induces lymphokine-activated killer cells for enhanced cytolysis in vitro.

We have examined the effect of the intradermal administration of IL-2 on the generation of natural killer (NK) cell and lymphokine-activated killer (LAK) cell activity. Peripheral blood mononuclear cells (PBMC) obtained from borderline lepromatous (BL) and lepromatous leprosy (LL) patients and normal volunteers prior to and after IL-2 injection were stimulated in vitro with IL-2 and their cytolytic activities compared against 51Cr labeled target K562 cells, Daudi cells, and monocytes. Before IL-2 administration, PBMC obtained from BL/LL patients and normal volunteers possessed similar levels of NK cell activity indicating that the NK cell activity of the BL/LL patients was intact. LAK cell activity was induced with IL-2 in vitro in both BL/LL patients and in normal volunteers. The level of LAK cell activity in BL/LL patients was, however, suboptimal. A single intradermal dose of 25 micrograms IL-2 had no effect on the phenotype of circulating mononuclear cells in either patients or normal volunteers. However, 6-12 days after IL-2 injection and subsequent restimulation of the PBMC with IL-2 in vitro, cytolytic activity of LAK cells obtained from the BL/LL patients was enhanced while cells from normal volunteers expressed the same high levels of activity as observed before IL-2 injection.

Cells, Cultured↗
PubMed1991

Generation of species-specific DNA probes for Leishmania aethiopica.

We report here the cloning of kinetoplast DNA (kDNA) sequences from Leishmania aethiopica in order to develop a specific and sensitive method for the identification of the parasite. Analysis of the cloned kDNA sequences showed different taxonomic specificities demonstrating sequence diversity within the kinetoplast DNA. Cloned whole minicircle hybridized with all Old World Leishmania species tested. Some cloned fragments of minicircle kDNA hybridized with Leishmania species causing cutaneous leishmaniasis in the Old World, but not with the viscerotropic species. Two L. aethiopica-specific clones were found. These clones hybridized with all L. aethiopica isolates tested, but did not react with other Leishmania species. The nucleotide sequence of the L. aethiopica-specific R3 clone is presented. Clones hybridizing with only some of the L. aethiopica isolates were also identified, although none of them showed specificity only for isolates causing localized (LCL) or diffuse (DCL) form of cutaneous leishmaniasis in Ethiopia.

Animals↗