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G Dennert

Publications and source records attributed to G Dennert.

At least 73 records · Page 4Linked to original sources
PubMed1984

Reorientation of the microtubule-organizing center and the Golgi apparatus in cloned cytotoxic lymphocytes triggered by binding to lysable target cells.

By immunofluorescence observations with cell couples of cloned murine cytotoxic T lymphocytes (CTL) and target cells, evidence is presented for a rapid reorientation of the microtubule-organizing center (MTOC) and the Golgi apparatus (GA) in the effector cell (but not in the target cell) toward the contact area with the target. The reorientation of the MTOC/GA and the cytotoxic activity of the CTL were inhibited reversibly by nocodazole, a microtubule-disrupting agent. In lectin-formed cell couples of CTL and neuraminidase-treated target cells, the MTOC in essentially all of the CTL was oriented toward the effector-target contact area of a lysable target cell, but was left randomly oriented with a nonlysable target cell. A similar random orientation of the effector-MTOC was also observed in cell couples of cloned natural killer cells and nonlysable targets. These findings indicate that the repositioning of the MTOC and the GA, which is shared by CTL and natural killer cells, is an essential and early event in the onset of the cytolytic mechanism. It is suggested that this reorientation serves the purpose of directing to the bound target cell secretory vesicles derived from the GA that contain cytotoxic substances.

Animals↗
PubMed1984

Glycosaminoglycan profiles in cloned granulated lymphocytes with natural killer function and in cultured mast cells: their potential use as biochemical markers.

Proteoglycans from three cloned, granulated lymphocyte cell lines with natural killer (NK) function (NKB61A2, HY-3, H-1) and one mast cell line (PT-18) were labeled with [35S]sulfate. [35S]proteoglycans were extracted in 1 M NaCl with protease inhibitors to preserve their native structure and were separated from unincorporated [35S]sulfate by Sephadex G-25 chromatography. [35S]proteoglycans from all four cell lines were chromatographed over Sepharose 4B and were found to have a similar range of m.w. The [35S]glycosaminoglycans from each cell line were then separated from parent proteoglycans by treatment with 0.5 M NaOH. The [35S]glycosaminoglycans from the three lymphocyte cell lines exhibited a similar m.w. as assessed by Sepharose 4B gel filtration, whereas the [35S]glycosaminoglycans from the mast cell line chromatographed as a smaller m.w. molecule. [35S )glycosaminoglycan charge characteristics were evaluated with DEAE C1-6B ion exchange chromatography. The consistency of the elution patterns was determined by using [35S]glycosaminoglycans obtained from radiolabelings of each cell line separated by 6 mo in culture. Each NK lymphocyte cell line reproducibly produced two distinct [35S]glycosaminoglycan chains that eluted in two regions well before the commercial heparin marker. The proportions of each chain were dependent upon the specific cell line. The mast cell line produced a single [35S]glycosaminoglycan chain, which eluted overlapping the internal commercial heparin marker, consistent with its higher charge characteristics. [35S]glycosaminoglycans from all cell lines were identified as chondroitin sulfates with the use of specific polysaccharidases. The NK lymphocyte glycosaminoglycans contained chondroitin 4-sulfate disaccharides. The mast cell glycosaminoglycans contained oversulfated disaccharides and chondroitin 4-sulfate disaccharides. Thus, each granulated NK lymphocyte cell line produced chondroitin sulfate glycosaminoglycans that were characteristic of that cell line and of different composition and less charge than those produced by cultured mast cells. These findings demonstrate that glycosaminoglycan profiles are useful biochemical markers in the characterization of diverse granulated cell lines including NK lymphocytes and mast cells.

Animals↗
PubMed1983

Isolation and partial characterization of human T-cell growth factor.

Human T-cell growth factor (TCGF) has been isolated from conditioned media of the Jurkat T-leukemia cell line. Using a high-efficiency isolation procedure involving hollow fiber concentration, gel filtration and 3 steps of reverse-phase HPLC we obtained 100 to 600 pmol TCGF per liter of conditioned medium. Jurkat cell-derived TCGF (jTCGF) has a molecular weight of 15,750. The amino acid composition of jTCGF agrees well with that derived from the cDNA sequence coding for this protein (Taniguchi et al, Nature 302, 305, 1983). jTCGF is highly active in vitro in stimulating the proliferation of T-cells as measured by 3H-thymidine incorporation into DNA (half-maximal stimulation with 3 fmol/100 microliters well).

Amino Acids↗
PubMed1983

Cytolysis by H-2-specific T killer cells. Assembly of tubular complexes on target membranes.

Cloned T killer cells derived from one-way mixed lymphocyte reactions were characterized with regard to their Lyt phenotype and specificity. Two clones of Lyt-1-2+ phenotype that recognized H-2Dd were selected and examined for their cytolytic function by negative staining and thin section electron microscopy. When incubated with the H-2d target S194, they assemble two types of tubular complexes, polyperforin 1 and 2. Both structures appear to arise by polymerization of precursors that may originate in dense granules and/or Golgi of T killer cells. Polyperforins appear to be associated with vesicles that are released during the lytic reaction and transferred to target membranes as shown by immunoelectron microscopy. Since there is a close correlation between target lysis and the appearance of polyperforins on target membranes, it is suggested that polyperforins take part in the cell-mediated killing reaction. Although polyperforins are different in size and several molecular properties from complement, there are striking similarities between these circular complexes and polyperforin (C9). It is therefore possible that they belong to a closely related family of cytolytic effector molecules.

Animals↗
PubMed1983

Polarization of the Golgi apparatus and the microtubule-organizing center within cloned natural killer cells bound to their targets.

In cell couples formed between a cloned murine natural killer (NK) cell and either of two target cells, we have obtained evidence by immunofluorescence observations for a rapid coordinate repositioning of two organelles, the microtubule organizing center and the perinuclear Golgi apparatus, inside the NK cell to face the region of contact with the target cell. With microtubule-disrupting drugs, which also cause a dispersion of the Golgi apparatus throughout the periphery of the NK cell, it was found that target cell lysis is reversibly inhibited. It is proposed that the coordinate repositioning of the two organelles serves the function of directing secretory vesicles derived from the Golgi apparatus to the bound target cells, the secretory vesicles containing components that induce target cell lysis.

Animals↗
PubMed1981

Cloned cell lines with natural killer activity. Specificity, function, and cell surface markers.

Cell lines with natural killer (NK) activity grown from native spleen cells cultured in medium conditioned by spleen cells proliferating in the presence of concanavalin A (Con A) were characterized. One NK cell line was cloned and assayed on several human and mouse NK-sensitive targets to analyze whether target specificities segregate upon cloning. Results showed that NK clones display target specificities identical to NK cells in normal spleen. This suggests that NK cells have no clonally distributed specific receptors to a given target. They may, however, have receptors which recognize identical antigens on all NK-sensitive targets or may possess multiple receptors for different target specificities. NK lines could not be demonstrated to possess activity in antibody-dependent cell-mediated cytotoxicity, nor did they effect mutual lysis. In the presence of Con A, NK cells exhibited dramatically enhanced lysis of NK-sensitive targets but only a slight increase in lysis of NK-insensitive targets. This indicates that the degree of lysis of an NK target is a function of two variables: effector binding to the target and target sensitivity to lysis. Furthermore, it suggests that the affinity of the putative antigen receptors on NK effectors must be rather weak. Cell surface marker analysis reveals that NK cell lines are Thy 1.2+, Lt-1-2-, T200+, asialo GM1+, and asialo GM2+. These markers distinguish NK cells from cytolytic thymus-derived lymphocytes, without resolving the question of classification within a give hematopoietic cell lineage.

Animals↗
PubMed1981

Fine specificity mapping of two allospecific T cell lines: recognition of private specificities in the H-2 IA subregion.

Two allospecific T-cell lines, C.C3.11.75 (H-2d anti-H-2k) and B6.C.7.76 (H-2b anti-H-2d) established from mixed lymphocyte cultures, were selected by continuous antigenic stimulation over several years. Both cell lines proliferated to alloantigen, provided allohelp to B cells in the humoral antibody response, and one of them showed cytolytic activity. Using spleen cells from various congenic mouse strains, the specificity of antigen recognition was examined in order to examine whether there is a correlation between T cell function and antigen recognition. Results showed that in all functional assays. This shows that there is no obligatory correlation between T cell function and antigen recognition and may suggest that the T cell receptor repertoire is biased towards recognizing private specificities.

Animals↗
PubMed1981

The Lyt phenotype of a long-term allospecific T cell line. Both helper and killer activities to IA are mediated by Ly-1 cells.

The relationship between surface antigen expression and function in the long-term allospecific T cell line C.C3.11.75 was examined by flow microfluorometry, antiserum plus complement depletion and cell sorting. T cells of the line expressed Lyt-1, but little or no Lyt-2 antigens. Proliferation to cells bearing Iak determinants, generation of T cell-replacing nonspecific helper factor in response to Iak-positive cells and the killing of Iak-positive targets were dependent only on Ly-1 cells. No obvious heterogeneity was found in this cell line despite its disparate functional activities. The fact that Lyt-2 molecules need not be present for killing directed against Ia determinants indicates that such molecules are not obligatory for the induction or delivery of killing and raises the question of what the role of Lyt molecules in T cell recognition or function might be.

Animals↗
PubMed1981

Culture supernatants of a stimulated T-cell line have helper activity that acts synergistically with interleukin 2 in the response of B cells to antigen.

Culture supernatants of an antigen-stimulated long-term alloreactive T cell line, C.C3.11.75, contain a T-cell-replacing factor (TRF) activity for the B-cell response to antigen. These same supernatants show little activity in the T-cell growth assay or the costimulator assay. TRF activity was assayed by using spleen cells that were rigorously depleted of both T cells and macrophages. In this assay preparations containing interleukin 2 and supernatants from stimulated C.C3.11.75 cells are relatively inactive if added alone but show marked synergy when added together. We conclude that the C.C3.11.75 TRF activity is not due to interleukin 1 or to interleukin 2 but to a third factor provisionally designated as (DL)TRF. This activity may be equivalent to the (late-acting) TRF described by Schimpl and Wecker. Evidence is presented suggesting that the helper activity (DL)TRF is a product of the T-cell line.

Animals↗
PubMed1981

T cells may express multiple activities: specific allohelp, cytolysis, and delayed-type hypersensitivity are expressed by a cloned T-cell line.

A T-cell line that recognizes private specificities encoded in the H-2 IA subregion in the proliferative response, cytotoxic activity, delayed-type hypersensitivity, and allohelp reaction with B cells has been cloned. The individual clones were assayed for ability to express the activities of the parent line. Results show that the cloned sublines tested are able to perform all the activities of the parent line. It is suggested that T cells may express multiple activities and that the apparent monofunctionality of T cells is due to regulation of these activities.

Animals↗
PubMed1981

T cell lines active in the delayed-type hypersensitivity reaction (DTH).

Two allospecific uncloned T cell lines, C.C3.11.75 (H-2d anti-H-2k) and B6-C.7.76 (H-2b anti-H-2d), were assayed for their ability to elicit a DTH reaction in vivo. It was shown that both cell lines, C.C3.11.75 being cytolytic and B6-C.7.76 being noncytolytic, evoke a strong DTH reaction. The reaction was in tempo and histology typical for a DTH reaction and could not be distinguished from a DTH reaction elicited by in vivo generated T cells active in a GVH reaction. Both cell lines recognize a private specificity encoded in the H-2 IA subregion. Cell proliferation of the responder cells did not appear to be critical in the reaction nor are T cells from the host recruited during the reaction. The DTH reaction can be assayed in allogeneic hosts demonstrating a complete lack of H-2 restriction in the reaction even when presumably antigen presenting macrophages are removed from the stimulator cell population. Furthermore, tumor cells expressing IA antigens could be employed to cause the reaction in both syngeneic and allogeneic recipients. Cloned sublines of one of the lines are also active in the reaction.

Animals↗
PubMed1980

Functional Thy-1+ cells in cultures of spleen cells from nu/nu mice.

Thy-1+ lymphocytes, detectable by quantitative serum absorption, arise in cultures of spleen cells from congenitally athymic (nu/nu) mice supplemented with supernatants from cultures of normal spleen cells stimulated with the T cell mitogen concanavalin A. Pretreatment of nu/nu spleen cells with appropriate anti-Thy-1 alloantibody and complement prior to culture reduces their capacity to generate Thy-1+ cells by about 90%. This shows that the majority of cells proliferating in these cultures are descendants of Thy-1+ cells which can be detected in the original nu/nu spleens. Experiments aimed at exploring the function of these Thy-1+ cells after culture in conditioned medium revealed that within one or two days after culture initiation, strong cooperative activity for a humoral response to xenogeneic erythrocytes can be detected. In mixtures of bone marrow-derived lymphocytes from various mouse strains with cultured nu/nu spleen cells, it was observed that T-B cooperation is not H-2-restricted. Attempts at inducing T cell-mediated cytotoxicity to alloantigens in such cultures of nu/nu spleen cells were unsuccessful. In contrast, nonspecific cytotoxicity which was attributed to natural killer cells was regularly observed and could be maintained in these cultures over extended periods.

Animals↗