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Biomedical subjects

R Kiessling

Publications and source records attributed to R Kiessling.

At least 109 records · Page 6Linked to original sources
PubMed1988

In vivo function of natural killer cells as regulators of myeloid precursor cells in the spleen.

In order to study the in vivo effect that natural killer (NK) cells may have on hematopoietic precursor cells in the autologous host, NK-depleted mice were constructed by injection of an anti-NK-1.1 monoclonal antibody. Only the spleen, and not the bone marrow, of such NK-depleted mice shows an increase in the number of myeloid precursur cells able to form colonies in soft agar as compared to control mice with normal levels of NK activity. The increase in number of hematopoietic immature cells due to lack of a regulatory cell (i.e. NK cell) is selective for the committed myeloid precursur cells, and not evident for erythroid or pluripotent progenitor cells. This is direct evidence in support of the hypothesis that NK cells in the autologous host serve as a regulator of extramedullary myelopoiesis.

Animals↗
PubMed1988

Cellular, humoral, and gamma interferon responses to Mycobacterium leprae and BCG antigens in healthy individuals exposed to leprosy.

Protective immunity against mycobacteria is dependent on antigen-specific T cells. The antibodies induced upon immunization with mycobacteria have no apparent role in host protection. Serological techniques have detected some antigens that are also recognized by human T cells but may fail to recognize others. Potentially, there may be differences in the epitopes seen by the T and B cell anti-mycobacterial antigen repertoires. We have screened the different components of sonicated BCG or Mycobacterium leprae that were separated according to their molecular weight (MW) by SDS-PAGE and then electroblotted on nitrocellulose paper. The blots were cut into squares and tested directly in a T cell proliferation assay. Our results indicate that peripheral T cells of healthy leprosy patient contacts respond preferentially to the lower MW (less than 70,000) and not the higher MW fractions of M. leprae and BCG, in contrast to the humoral response of these same individuals. The most important fractions in inducing a lymphoproliferative response were in the regions of 11-16 kDa of BCG and M. leprae and to the 22-26 kDa region of M. leprae. These fractions appeared to represent molecular weight regions that were in some instances clearly distinct from previously defined antigens. It was further shown that lymphoproliferation in response to mycobacterial fractions correlated with the production of gamma interferon, a lymphokine required for macrophage activation and elimination of mycobacteria. These studies allow the direct assessment of antigens involved in protective T cell-mediated immunity, and should be helpful in selecting relevant antigens for skin testing and immunization.

Antibody Formation↗
PubMed1987

An active T-cell-independent mechanism enhances MHC class I transcription and expression on a mouse T-cell lymphoma in vivo.

The present study focuses on the mechanism underlying changes in H-2 cell surface antigen expression after passage of the in vitro grown YAC-1 lymphoma as an ascites tumor. The increase in cell-surface expression correlated with elevated levels of class I transcripts as revealed by Northern blots. The enhanced H-2 expression was also seen with a cloned YAC-1 line, and not until 2 weeks after in vitro explantation had levels of H-2 decreased to those on the in vitro established YAC-1. Arguing for the necessity of a mature functioning immune system, suckling mice were unable to increase H-2 expression on inoculated lymphoma cells. Also pretreatment with cyclophosphamide or irradiation abolished the capacity of adult mice to increase cell surface H-2 on YAC-1 cells. A functioning T-cell system was not required for H-2 enhancement to occur since athymic nude mice were fully competent. The possible significance of an active T-cell-independent host mechanism which enhances tumor H-2 expression at the transcriptional level is discussed.

Age Factors↗
PubMed1987

Interferon gamma induces lung colonization by intravenously inoculated B16 melanoma cells in parallel with enhanced expression of class I major histocompatibility complex antigens.

Treatment of H-2-deficient nonmetastatic B16 melanoma cells with physiological doses of interferon gamma (IFN-gamma) reduced cellular growth in vitro but induced a shift to the lung-colonizing phenotype as assessed after intravenous injection of the treated cells. As little as 1 antiviral unit of recombinant IFN-gamma per ml induced B16 cells to form 3-40 pulmonary metastases in each injected mouse, whereas a 1000-fold higher concentration of IFN-beta was required to see similar effects. IFN-gamma may induce cell-surface molecules that contribute to the metastatic ability of the tumor cells. The efficient enhancement of metastatic ability after IFN-gamma treatment of the B16 cells was paralleled by an increased H-2 antigen expression and decreased sensitivity to natural killer cells. The experiments support the idea that metastasis may not depend exclusively on stable genetic changes or heterogeneity within a tumor population but may be also influenced through the modulation of the phenotype by physiological or pharmacological agents. The results are also discussed with regard to the role of different effector cells in tumor cell clearance and in relation to lymphokine-based strategies for therapy.

Animals↗
PubMed1986

An immunofluorescent method for identifying individual IFN-gamma-producing lymphocytes.

An immunofluorescent staining method was developed for detecting human IFN-gamma-producing cells in single cell suspensions. Mononuclear leukocytes, stimulated in vitro to produce IFN-gamma, were fixed and made permeable. The cytoplasmic presence of IFN-gamma was visualized by indirect immunofluorescence using IFN-gamma-specific mouse monoclonal antibodies. The staining was found to be specific for IFN-gamma and allowed the detection of newly synthesized rather than internalized IFN-gamma molecules. The cytoplasmic fluorescence appeared locally in a polar, juxtanuclear position, which overlapped the Golgi apparatus, probably reflecting the glycosylation site of the newly formed IFN-gamma molecules. Two-colour staining experiments showed that the method is useful not only for the detection and enumeration but also for the phenotypic characterization of IFN-gamma-producing cells.

Antibodies, Monoclonal↗
PubMed1986

Phenotypic characterization of individual interferon-gamma-producing cells after OKT3 antibody activation.

This is the first study using a direct approach to determine the phenotype of interferon gamma (IFN-gamma)-producing cells after polyclonal T cell activation. Blood mononuclear cells (MNC) were cultured and stimulated by OKT3 antibody to produce IFN-gamma. They were then stained with a panel of monoclonal antibodies with specificity for cell surface antigens, fixed and permeabilized in suspension and stained for cytoplasmic IFN-gamma using monoclonal antibodies and indirect immunofluorescence technique. IFN-gamma appeared mainly as a local, polar accumulation in the cytoplasm in a juxtanuclear position, probably identical to the Golgi apparatus. After a culture period of 20 h, 6-11% and 0.2-0.6% of the OKT3-activated MNC from adults or newborns, respectively, produced IFN-gamma in cultures. Unstimulated cultures contained maximally 0.1% IFN-gamma-positive MNC. The majority of the IFN-gamma-producing MNC were T cells and expressed the T11 antigen and receptors for interleukin 2. To our surprise most of these T cells also carried receptors for complement 3bi (OKM1) but did not express class II molecules. Less than 50% displayed T4 or T8 antigens in three out of the four experiments performed. Natural killer cells, as recognized by Leu 7 or B73.1 antibodies, contributed only marginally to the IFN-gamma synthesis whereas B cells (OKB7) and monocytes (Leu M3) showed no production of IFN-gamma.

Antibodies, Monoclonal↗
PubMed1986

Mechanisms of lymphocytic choriomeningitis virus-induced hemopoietic dysfunction.

Results of this study showed that lymphocytic choriomeningitis virus infection causes a marked activation of natural killer (NK) cells not only in the spleen but also in the bone marrow. This activity reached its peak at about day 3 of infection and declined after days 6 to 7. Enhanced NK cell activity was found to correlate with decreased receptivity for syngeneic stem cells in bone marrow and spleen, with the notable exception that decreased receptivity persisted longer in bone marrow. Treatment of infected recipients with anti-asialo GM1 (ganglio-N-tetraosylceramide) significantly increased the receptivity for syngeneic hemopoietic cells. These findings are consistent with the hypothesis that NK cell activation causes rejection of syngeneic stem cells, thus resulting in hemopoietic depression. To understand the mechanisms behind the prolonged decrease in bone marrow receptivity (and bone marrow function in the intact mouse) mentioned above, we followed the changes in the number of pluripotential stem cells (CFU-S) circulating in the peripheral blood and in endogenous spleen colonies in irradiated mice, the limbs of which were partially shielded. It was found that following a marked early decline, both parameters increased to normal or supranormal levels at about day 9 after infection. Because the bone marrow pool of CFU-S is only about 20% of normal at this time after infection, a marked tendency for CFU-S at this stage in the infection to migrate from the bone marrow to the spleen is suggested. It seems, therefore, that as NK cell activity declines, the spleen regains the ability to support growth of hemopoietic cells and the bone marrow resumes an elevated export of stem cells to the spleen. This diversion of hemopoiesis could explain both the long-standing deficiencies of the bone marrow compartment and the prolonged decrease in the receptivity of this organ.

Animals↗
PubMed1986

Interferon and butyrate treatment leads to a decreased sensitivity of NK target cells to lysis by homologous but not by heterologous effector cells.

Human K-562 and HHMS cells were pretreated with human recombinant interferon (IFN)-gamma and used as targets in NK assays against human and murine effector cells. A protective effect against NK lysis was observed only in the homologous assay, whereas no change or even a slight increase in NK sensitivity against heterologous effector cells was found. In cold target inhibition experiments IFN-treatment of K-562 cells led to a decrease in their capacity to act as competitors in the homologous NK assay, leaving their inhibitory capacity unaltered in the heterologous assay. In accordance with results observed using human NK targets, murine YAC-1 cells treated with mouse recombinant IFN-gamma did not lose their susceptibility to human NK cells. However, they were markedly less susceptible to lysis mediated by murine effectors. Butyrate, another compound causing decreased sensitivity of K-562 cells for human natural killing, also failed to reduce the susceptibility against murine NK cells. The results indicate that the NK-resistant tumor target phenotype caused by IFN or differentiation-inducing agents can only be detected by homologous but not by heterologous effector cells. This suggests that major differences exist between the inter- and intraspecies NK killing mechanisms.

Animals↗
PubMed1986

Genetic studies on a tumor growth-inhibitory factor in serum of normal mice and its relationship to NK activity.

It has been shown that normal mouse serum contains a tumor growth-inhibitory factor (GIF). and that strain-dependent levels of GIF correlate with mouse NK activity. To further analyze the genetic control of GIF we have studied the growth-inhibitory activity of normal mouse serum from 8 different mouse strains and their F1 hybrids. A sensitive method using a chromogenic substrate for an endogenous lysosomal enzyme was used to measure the inhibitory activity of normal mouse serum on the mouse B16 melanoma. The highest level of GIF was found in old mice, lower activity in serum of young animals and no activity in suckling mice. To compare the genetic control of GIF and NK, spleen NK activity against B16 as well as YAC-1 targets was measured in parallel in the same animals. Confirming previous results we found the H-2k strains CBA and C3H to have high levels of GIF as well as NK activity, while the strain A/Sn and the A congenic strain A.SW had low levels of both activities. Experiments with H-2d and H-2b strains, however, showed that GIF and NK had a different genetic control; thus the DBA/2 and Balb/c strains had considerably higher GIF activity than the C57B1 and Leaden strains, while the reverse was true for NK activity. In F1 hybrid crosses between strains with high and low activity, high activity was inherited as a dominant trait for both GIF and NK. A backcross analysis in (A X CBA) X A backcross mice, segregating for NK and GIF showed that the two activities did not cosegregate. These studies therefore demonstrate that GIF and NK activity are under different genetic control, and do not support any direct or simple relationship between GIF and NK cells.

Animals↗
PubMed1985

Interferon-gamma is a strong modulator of NK susceptibility and expression of beta 2-microglobulin but not of transferrin receptors of K562 cells.

The human cell line K562 was treated with human natural leukocyte interferon (IFN-alpha) and recombinant immune interferon (IFN-gamma). Cell cultures exposed to both types of IFNs displayed a reduced susceptibility to the cytotoxic activity of human PBL (NK activity). While this effect occurred preferentially at high doses of IFN-alpha, as little as 10 U/ml of IFN-gamma caused a marked decrease in susceptibility to NK-cell-mediated lysis. Using a monoclonal antibody against human beta2-microglobulin (beta2M) a low level of specific binding to K562 cells was detected. The binding increased after treatment with IFN-alpha (1.4-fold) and IFN-gamma (1.7-fold). The expression of transferrin receptors (TR) was not changed significantly. A hybrid cell line between K562 and a Burkitt's lymphoma-derived cell line displayed a similar pattern of response to IFN-alpha and IFN-gamma as did K562, when effects on NK susceptibility, beta2M expression, and TR expression were studied. The Burkitt's lymphoma line PUT showed no consistent changes in expression of beta2M and TR. These results demonstrate that IFN-gamma is highly efficient in modulating the NK susceptibility, and the expression of beta2M on K562. The presented data do not support a role for expression of TR as the only property that determines the degree of NK susceptibility, since there was no correlation between NK susceptibility and TR expression among the cell lines tested or when IFN-treated and untreated cells were compared.

Cell Line↗
PubMed1985

Comparison between murine natural antibodies and natural killer cells: recognition of separate target structures as revealed by differential in vitro expression and dependence on glycosylation.

The specificity of complement-fixing, cytotoxic antibodies against the YAC lymphoma in sera of normal young adult (A X C57BL)F1 mice was studied. In vivo-maintained, immunoselected sublines of the YAC lymphoma expressed low amounts of the natural antibody (NAb) target structure. These cell lines were also resistant to natural killer (NK) cell-mediated lysis. After 2-3 weeks of in vitro culture the immunoselected cell lines became NK sensitive, but they remained resistant to NAb. When several independently derived variants selected for low NK sensitivity were tested for their ability to absorb NAb, the degree of absorption varied considerably among the variants. NAb could be inhibited by purified C-type virus particles and also by bacterial sonicates and various glycoprotein preparations. Treatment of target cells with tunicamycin, an inhibitor of asparagine-linked glycosylation, decreased the sensitivity to NAb lysis but had no impact on NK sensitivity. Thus the results indicated that a) NAb and NK cells recognized separate target structures and b) the target structure(s) for NAb but not for NK cells were saccharides of the N-glycosidic type.

Animals↗
PubMed1985

YAC-1 MHC class I variants reveal an association between decreased NK sensitivity and increased H-2 expression after interferon treatment or in vivo passage.

Two H-2 negative variants of the YAC-1 lymphoma were selected by mutagenization and sequential in vitro selections and compared with wild-type cells for changes in NK sensitivity and H-2 expression after interferon treatment or in vivo passage. The H-2 negative variants and the low H-2 expressor YAC-1 wild-type cells had similar NK sensitivity. However, IFN-beta or recombinant IFN-gamma pretreatments increased the H-2 expression of YAC-1 and protected them from NK lysis, whereas the H-2 variants, which remained H-2 negative, were not protected and often more sensitive to NK lysis. The H-2 variants were similarly susceptible as wild-type cells to three other cellular effects of interferon: protection from virus infection, modulation of Con A capping, and inhibition of cell proliferation. Thus, the only interferon-mediated effect that distinguished the H-2 negative variants from wild-type cells was the inability of the former to increase their H-2 expression and decrease their NK sensitivity. The wild-type YAC-1 line showed increased H-2 expression and decreased NK sensitivity after in vivo passage. In contrast, in vivo passaged H-2 variants showed no reexpression of H-2, and remained NK sensitive. The altered responses to interferon and in vivo passage were specific for loss or down-regulation of H-2, because Thy-1 loss (H-2 positive) YAC-1 variants behaved as the wild-type cells in all respects. This study supports the hypothesis that NK cells may function in vivo to eliminate host cells that fail to express H-2 after interferon stimulation during an immune response; such cells are a potential threat because they may escape recognition by T lymphocytes despite the expression of viral or tumor-associated antigens.

Animals↗
PubMed1983

Gamma-interferon (IFN-gamma) produced during effector and target interactions renders target cells less susceptible to NK-cell-mediated lysis.

Human mitomycin C-treated PBL were mixed with cells of an NK sensitive hybrid cell line (PUTKO-I). A fraction of tumor cells survived this treatment and could be recovered from the cultures. These surviving cells were completely NK-resistant and this property persisted for 2-3 weeks after cultivation in fresh medium. Treatment of a clone (C13) of PUTKO-I with PBL-PUTKO mixed lymphocyte-tumor-cell culture (MLTC) supernatants resulted in a marked reduction in NK sensitivity after 8-12 h of treatment. The kinetics of induction of NK resistance by MLTC supernatants was similar to that of purified IFN-gamma and was faster than for IFN-alpha. The active component in the supernatants was characterized as a mixture of IFN-gamma and IFN-alpha based on neutralization of activity with specific antisera. The role of mycoplasma contamination was investigated and it was found that cell lines free of detectable mycoplasma stimulated production of NK-protective activity by PBL and this activity was neutralized by anti-IFN-gamma serum. Separation of PBL on discontinuous Percoll gradients demonstrated a correlation between the NK activity of cell fractions and their ability to produce IFN in response to tumor cells. Taken together, the selection-dependent variations in NK sensitivity, the kinetics of IFN production and induction of resistance suggest that tumor cells may be able to escape elimination by NK cells due to protection by IFN produced by the effector-cell-containing population.

Cell Line↗
PubMed1983

Incidence and type of tumors induced by DMBA carcinogenesis in H-2 congenic strains on a B10 background.

Five congenic strains on B10 background (B10, B10.S, B10.G, B10.BR and B10.A) were investigated for their susceptibility to tumor induction by 9,10 dimethyl-1,2 benzanthracene (DMBA). The DMBA was administered via a stomach tube five times at weekly intervals (total 5 mg). Two types of malignancies predominated: diffuse lymphocytic lymphomas with or without thymic involvement, and epidermal tumors. B10 (H-2b) was the most resistant strain when total tumor incidences were compared. B10.S (H-2s) had the highest incidence of both lymphomas and epithelial tumors and B10.G was significantly more susceptible to thymic lymphomas than B10. In B10.S mice thymic lymphomas were more frequent in males than in females, the reverse being true for tumors of epithelial origin. No correlation between tumor incidence and in vitro NK activity was demonstrated.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1983

Interferon-induced increase in neuraminidase-releasable sialic acid and glycosphingolipid metabolism in mouse lymphoma and L1210 leukemic cell lines: correlation with susceptibility to natural killer cell-mediated lysis.

Changes in sialoglycoconjugates and glycosphingolipid (GSL)5 metabolism were demonstrated in mouse EL4, P52 and YAC-1 lymphoma and L1210 leukemia cell lines treated with beta-interferon (IFN). Expression of cell surface (neuraminidase-releasable) sialic acid on IFN-treated cells was markedly elevated (three- to six-fold). The increase in neuraminidase-releasable sialic acid is contributed by sialoglycoproteins and particularly by cell-surface gangliosides in IFN-treated cells. Incorporation of [3H]-galactose into all GSL was elevated in IFN-treated cells. Thin-layer chromatographic analysis of GSL of IFN-treated cells showed an increase in several GSL homologues with striking changes in ganglioside with chromatographic migration of GM2, GM1, and GD1a relative to control cells. IFN-treated tumor-cell lines became resistant to lysis by virus-induced IFN-activated natural killer (NK) cells, as shown previously, but addition of neuraminidase to IFN-treated and untreated cells caused only a moderate increase in NK-sensitivity. This suggests that IFN-mediated protection of target cells from NK lysis was not due to a preferential masking of target structure by elevated levels of sialic acid. These membrane-associated changes in GSL and sialic acid in IFN-treated cells may be potentially significant, because a correlation between certain GSL expression, sialic acid phenotype and susceptibility of target cells to NK-cell-mediated lysis have been found in several other systems.

Animals↗