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Biomedical subjects

S C Knight

Publications and source records attributed to S C Knight.

At least 109 records · Page 6Linked to original sources
PubMed1990

HIV I infection of dendritic cells.

Dendritic cells (DC) from human peripheral blood are susceptible to productive and probably to latent infection with HIV-I. Infection of DC also occurs in vivo since in HIV-seropositive individuals Langerhans' cells of the skin and DC from peripheral blood, (in preparation) are infected. In peripheral blood 3-25% of DC, identified as large, low-density cells lacking monocyte markers, are infected as judged by in situ hybridization with an HIV probe. This contrasts with the lower proportion (< 0.2%) of other cells infected. DC exposed to HIV in vitro or in vivo fail to present other antigens or mitogens to stimulate T cells. This functional defect in infected DC is not blocked by the presence of soluble CD4 antigen and occurs in the absence of T cell infection suggesting a block at the level of the antigen-presenting cell itself. Infection, depletion and dysfunction of DC in HIV seropositive patients is already present in asymptomatic individuals and this precedes the appearance of T cell defects. We speculate that loss of functional DC may be a fundamental defect leading to a block in recruitment of resting T cells into immune responses. In contrast to the HIV-induced impairment of antigen presentation by DC, these cells were potent stimulators of responses to the HIV antigens themselves. Normal DC infected with HIV in vitro stimulated primary proliferative and cytotoxic T cell responses (in preparation). These were produced in cells from individuals expressing a range of different MHC types but the cytotoxic cells, once produced, killed autologous but not allogeneic, infected T cell blasts. Primary response to viral peptides can also be produced suggesting that this system may be useful for identifying immunogenic epitopes of HIV using cells from sero-negative, non-immunocompromised individuals.

CD4 Antigens↗
PubMed1990

Antigenic competition in contact sensitivity. Evidence for changes in dendritic cell migration and antigen handling.

The frequency and antigen-bearing characteristics of dendritic cells (DC) within draining lymph nodes have been examined during antigenic competition in contact sensitivity. Pre-exposure of mice to oxazolone on the flank resulted in a marked depression of subsequent fluorescein isothiocyanate (FITC)-induced lymph node cell (LNC) proliferation. Antigenic competition was associated with an increased frequency of DC in the draining lymph nodes, but also with a reduced amount of antigen per cell. Thus, at 24 hr the draining nodes of FITC-challenged mice previously exposed to oxazolone exhibited an increased number of DC compared with control animals. Flow cytometric analysis revealed, however, that the percentage of antigen-bearing DC was reduced and that the median amount of antigen borne by DC was lower. Since exposure to oxazolone caused a significant increase in the frequency of DC in distant nodes, the changes observed in antigenic competition may, at least in part, be attributable to systemic effects on DC migration following application of the first antigen. These data indicate that the reduced primary proliferative response which characterizes antigenic competition in contact sensitivity is associated with, and may result from, induced changes in dendritic cell behaviour.

Animals↗
PubMed1990

Systemic migration of dendritic cells during contact sensitization.

Twenty-four hours after skin painting mice on the flank with the contact sensitizer fluorescein isothiocyanate (FITC), the number of dendritic cells (DC) increased sharply, not only in draining but also in contralateral (CLN) and distant lymph nodes. High levels of antigen were detected on up to 50% of DC isolated from draining lymph nodes (DLN), and these cells were potent stimulators of naive T cells in vitro. Less than 3% of DC from contralateral and distant lymph nodes carried detectable antigen and did not induce significant T-cell proliferation. A significant number of DC had migrated to draining, contralateral and distant lymph nodes without acquiring detectable antigen. This indicates that there is a systemic signal causing the movement of DC to lymph nodes. This appears to be independent of mature T cells, as the systemic migration of DC also occurred in nude mice.

Animals↗
PubMed1990

Dendritic cell infection, depletion and dysfunction in HIV-infected individuals.

Immune responses in resting T cells are initiated by the presentation of antigen by bone marrow-derived dendritic cells (DC). Normal DC are susceptible to infection with human immunodeficiency virus (HIV) in vitro (Patterson & Knight, 1987) and this blocks their capacity to stimulate T-cell responses to other antigens (Macatonia, Patterson & Knight, 1989a). To study the relationship between HIV and DC in patients and its relevance to the pathogenesis of disease, DC have been isolated from the blood of individuals in the different clinical categories, counted, examined for the presence of virus genome and their antigen-presenting capacity measured. Infection, depletion and impaired function of DC occur in early HIV infection. HIV seropositive patients who were asymptomatic and those with symptoms of disease had significantly reduced numbers of DC, but patients with persistent generalized lymphadenopathy had normal numbers. Between 3% and 21% of DC, identified as large low-density cells not bearing monocyte, lymphocyte or natural killer cell markers, were infected with HIV, as indicated by in situ hybridization. Less than 0.12% of the lymphocytes or monocytes were infected. The DC from infected individuals were poor at enhancing responses to the mitogen concanavalin A (Con A). They also caused low levels of stimulation in allogeneic lymphocytes in mixed leucocyte cultures. By contrast, T cells from asymptomatic patients gave normal T-cell responses to uninfected allogeneic DC, although those from acquired immunodeficiency syndrome (AIDS) patients did show reduced responsiveness. Defects in DC thus precede both the appearance of symptoms and changes in T cells and may be instrumental in the development of AIDS. Furthermore, since DC numbers and function differ at different stages of disease, monitoring these may contribute to clinical assessment and lead to new therapeutic approaches.

Cell Count↗
PubMed1989

Primary stimulation by dendritic cells induces antiviral proliferative and cytotoxic T cell responses in vitro.

We used well-gassed hanging drop (20 microliters) cultures with high concentrations of purified T cells from normal BALB/c mice to examine whether dendritic cells (DC) can induce primary antiviral proliferative T cell responses and generate virus-specific CTL. We found that DC exposed to infectious influenza virus in vitro or in vivo in small numbers (0.1-1%) resulted in strong proliferation of responder T cells within 3 d, and this was strongly inhibited by antibodies to class II MHC molecules. In addition, in 5-d cultures, the influenza-treated DC generated CTL specifically able to lyse influenza-infected syngeneic target cells bearing MHC class I antigens. The most potent nucleoprotein (NP) epitope recognized by BALB/c CTL is peptide 147-158 (Arg156-) and influenza-infected DC in vitro stimulated CTL recognizing this peptide, thus mimicking the response in mice primed by intranasal influenza infection. We also induced T cell proliferation and virus-specific CTL in cultures of normal T cells by stimulating with DC pulsed with the natural NP sequence 147-158 or the potent peptide 147-158 (Arg156-). Small numbers of peritoneal exudate cells, after activation with Con A to produce class II MHC expression and after removal of DC with a specific mAb (33DI), did not lead to primary CTL generation but initiated secondary stimulation in vitro. Our results using the hanging drop culture method and DC as APC have implications for studying the T cell repertoire for viral components in humans without the necessity of previous immunization.

Animals↗
PubMed1989

Lithium and hydrocortisone interactions on cell growth and gene expression in human promyelocytic leukemia (HL60).

Lithium (Li) and hydrocortisone (HC) modify gene expression as well as stimulate human hematopoiesis in vivo and in vitro. The responses of blood-forming tissues, to these natural substances individually, are closely similar. However, clonogenic co-culture experiments, using normal human bone marrow cells, have revealed that the effects of Li and HC are not additive but rather mutually inhibitory. Studies of HL-60 cells in suspension indicate that growth of this human promyelocytic leukemic line is enhanced by HC at 10(-6) M, a "therapeutic" plasma concentration. This phenomenon is abrogated by the co-addition of Li at 3 x 10(-4) M, likewise a "therapeutic" plasma concentration. Neither substrate exerts any influence on the morphological or cytochemical features of differentiation which accompany exposure of HL-60 cells to dimethyl sulphoxide (DMSO), retinoic acid (RA), tetradecanoyl phorbol acetate (TPA) or sodium butyrate. In contrast, the expression of c-fms, a cellular proto-oncogene which codes for the CSF-1 (monocyte-macrophage colony-stimulating factor) surface membrane receptor, is modified by Li in the presence of the differentiation induction agents DMSO and RA which promote the development of mature granulocytes from HL-60 cells. These observations afford further insights into the humoral mechanisms which modulate the expression of genes involved in the regulation of hematopoiesis.

Cell Differentiation↗
PubMed1989

Dendritic cells and T cells transfer sensitization for delayed-type hypersensitivity after skin painting with contact sensitizer.

The cells involved in the development of delayed-type hypersensitivity (DTH) to the contact sensitizer fluorescein isothiocyanate (FITC) were examined. Cells used for transferring sensitization were obtained from donor mice up to 5 days following skin painting with FITC. Recipient mice were sensitized by footpad injections of dendritic cells (DC) obtained from donor lymph nodes up to 3 days following skin painting, when DC expressed high levels of antigen. DTH, assessed by ear swelling 24 hr following ear challenge with FITC, was detected when recipient mice were challenged 5 days after transfer of DC, but not when ear challenged immediately after transfer. Removal of donor DC using the cytotoxic antibody 33D1, plus complement, either from the DC-enriched population or from whole lymph node cells 24 hr after skin painting, abolished the capacity to transfer DTH. Purified T lymphocytes obtained from donor mice between Days 3 and 5 after skin painting, transferred DTH when recipients were ear challenged with FITC 5 days after footpad injections. DTH also occurred in mice ear challenged with antigen immediately after receiving footpad injections of either normal or irradiated T cells obtained from donors 4 days after skin painting. B cells and macrophages did not transfer sensitization for DTH throughout the time-course. Therefore an early stage in the immune response, where antigen-bearing DC initiated DTH, was distinguished from a later stage, where T cells transferred sensitization.

Adjuvants, Immunologic↗
PubMed1989

The effect of retinoids on dendritic cell function.

The effect of retinoid administration on the antigen presenting function of mouse dendritic cells (DC) was assessed. Culturing spleen cells with retinoic acid (10(-10)-10(-4) M) had no effect on the numbers of DC separated from these cultures. However, DC isolated from retinoic-acid-treated cultures were less stimulatory than DC from untreated cultures when added to allogeneic lymphocytes in a mixed leucocyte culture. Allogeneic stimulation by DC was also inhibited by pulsing separated DC with retinoic acid (10(-6)-10(-4) M) for 2 h. Pulsing DC with lower doses of retinoic acid (10(-14)-10(-20) M) enhanced this response. DC isolated from animals maintained on VAA-enriched diets had a reduced capacity to stimulate allogeneic lymphocytes. The response of unseparated lymph node cells pulsed with retinoic acid to untreated allogeneic DC was inhibited by 10(-10)-10(-4) M retinoic acid but enhanced by lower doses (10(-14) M). However, the inhibitory effect of retinoids on the function of responding lymphoid populations was abolished on removal of DC from responding cells. The results indicate that immunomodulation by retinoids could occur via an effect on the efficiency of antigen presentation by DC.

Animals↗
PubMed1989

Suppression of immune responses by dendritic cells infected with HIV.

Evidence of human immunodeficiency virus (HIV) replication both in the skin Langerhans' cells of AIDS patients (Tschachler et al., 1987) and in normal, peripheral blood dendritic cells (DC) (Patterson & Knight, 1987; Knight & Patterson, 1989) suggests that infection of these antigen-presenting cells may contribute to the immunosuppression seen in AIDS. Support for this hypothesis is now provided by experiments in which the capacity of DC infected in vitro to present mitogen to normal syngeneic lymphocytes was measured. Infecting DC with HIV before culturing with lymphocytes inhibited mitogen-stimulated cell proliferation. Viral DNA was detected in DC in these cultures by in situ hybridization but, in addition, HIV was also present in a small proportion of lymphocytes. However, introducing an inhibitor of virus replication, 2',3' dideoxyadenosine, after infection of the DC but before culturing with lymphocytes, blocked growth of HIV in lymphocytes. In these latter experiments mitogen proliferation responses were still suppressed. Infection of DC could, therefore, cause immunosuppression in AIDS, both by direct effect on antigen-presentation and by the transfer of HIV to T cells.

Acquired Immunodeficiency Syndrome↗
PubMed1989

Class II antigens on dendritic cells from the synovial fluids of patients with inflammatory arthritis.

Dendritic cells were enriched from synovial fluids (SF) of patients with inflammatory arthritis and studied by immunogold labelling and electron microscopy for expression of histocompatability antigens of the HLA-D locus. Dendritic cells from SF were larger than most of these from peripheral blood with a more extensive Golgi region and more lysosomes and microfilaments. Class II histocompatability antigens HLA-DR, -DP, -DQ and that labelled by the antibody RFDI were abundant on the dendritic cells. The macrophages in the enriched cells showed labelling for DR but little labelling with the other antibodies. DR, DP and RFDI were often concentrated at areas of contact between dendritic and other cells (other dendritic cells, macrophages or lymphocytes). On incubating labelled cells at 37 degrees C for 30 min many macrophages lost their DR label but dendritic cells always retained some surface label. Some gold labelling DR and DP was found in characteristic channels between the veils and became internalized in membrane-bound structures. A small proportion of the RFDI label internalized in areas resembling coated pits. Less DQ label internalized and appeared on vesicles inside vacuoles. Material bound to different class II molecules may thus be internalized or processed differently by dendritic cells. The presence in inflammatory lesions of large activated dendritic cells with high expression of class II antigens suggests that these cells could be presenting antigen to lymphocytes within the joints.

Arthritis↗
PubMed1988

Induction of autoimmunity with dendritic cells: studies on thyroiditis in mice.

The initiation and maintenance of thyroid autoimmunity by professional antigen-presenting cells were assessed by observing thyroiditis and induction of IgG antibodies to thyroglobulin (Tg). Dendritic cells (DC) were purified from spleens of CBA mice and T cells removed with anti-Thy 1 and complement. Some DC were pulsed with 25-500 micrograms/ml of mouse Tg in vitro and normal syngeneic mice received injections of 10(5) cells intravenously. In untreated animals only 1 thyroid out of 40 showed a lymphocyte infiltrate and antibody to Tg was rarely seen. In animals receiving normal DC without Tg, lymphocyte infiltration was seen 2-6 weeks later in 5 out of 33 thyroids and some animals produced low levels of antibody to thyroglobulin (8 of 33 animals). DC pulsed with 500 micrograms Tg/ml in vitro caused thyroid infiltration in 6 out of 15 animals but did not increase the incidence of anti-Tg antibodies. Lower doses had no effect. When 10(5) DC were given from animals with experimental allergic thyroiditis (EAT, induced with Tg in complete Freund's adjuvant, CFA) more than half of the recipient animals showed thyroiditis (8 out of 15) and autoantibody production (12 of 15 animals). DC may therefore play a role in the initiation and maintenance of autoimmunity by providing a stimulus for antigen-specific T cells.

Animals↗
PubMed1988

Dendritic cells and viruses.

Dendritic cells (DC) are potent at presenting viral antigens in the initiation of both primary and secondary responses. DC in the lymph nodes draining the site of infection with HSV express surface antigen and can stimulate proliferation of sensitised lymphocytes. In secondary stimulation, nonresponsiveness in cytotoxic T cell assays of cells from mice primed to Moloney virus was also overcome by stimulation in vitro with virus-pulsed DC. Marked primary proliferative and cytotoxic T cell responses were previously found only to alloantigens and to haptens, both presented on the surface of DC However, DC exposed in vitro or in vivo to influenza virus stimulated primary proliferative and cytotoxic T cell responses in normal syngeneic lymphocytes in 20-microliters hanging drop cultures (Macatonia, Taylor, Knight and Askonas, in press). This provides a method for analysing primary responses to viruses in vitro without the necessity of using pre-sensitised donors. Although DC may present HIV antigens to lymphocytes the DC are also susceptible to infection with HIV. This occurs in vivo as evidenced by the infection of Langerhans cells in AIDS patients. This infection of DC may not only compromise their function in antigen presentation but also act as a reservoir of virus which is handed on to T cells during the close clustering of the presenting cells with the T cells during the initiation of the immune response.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Blocking of acquisition and presentation of antigen by dendritic cells with cyclosporine. Studies with fluorescein isothiocyanate.

The effect of cyclosporine on the acquisition and presentation of antigen by dendritic cells (DC) was examined. Mice skin-painted with the contact sensitizer FITC received 100 mg/kg of CsA orally on the day before and the day of sensitization. This blocked the development of delayed hypersensitivity as measured by ear-swelling on challenge with antigen on day 6. The antigen-presenting DC in the draining lymph nodes 24 hr after skin painting were studied. The numbers of DC within the lymph node more than doubled after skin painting and this was not altered by the treatment with CsA. The DC from normal skin-painted animals showed a biphasic distribution of antigen, with more than half the cells acquiring high levels of antigen and the remainder having low amounts. In the animals treated with CsA most cells had low levels of antigen. The DC from untreated animals stimulated primary proliferative responses of syngeneic lymphocytes in vitro and initiated delayed hypersensitivity in recipient animals, but the DC from CsA-treated animals did not stimulate immune responses. Cyclosporine may, therefore, prevent acquisition and presentation of antigen by DC in addition to any direct effects on T and B cells.

Animals↗
PubMed1987

Localization of antigen on lymph node dendritic cells after exposure to the contact sensitizer fluorescein isothiocyanate. Functional and morphological studies.

We have examined the cells involved in the development of contact sensitivity to FITC in CBA mice. After skin painting with antigen, the number of dendritic cells (DC) in the draining lymph nodes increased by 30 min, was maximal at 48 h, and returned to normal by 6 d. Derivation of some DC from Langerhans' cells of the skin was indicated from the presence of Birbeck granules observed in some DC isolated 24 h after skin painting. The DC acquired FITC and by 8 h there were two populations, one highly fluorescent and the other less fluorescent. The highly fluorescent cells were present between 8 h and 3 d after sensitization, and during this period the DC were potent at initiating primary proliferative responses of normal syngeneic T lymphocytes in vitro. Between days 3 and 5 the numbers of lymphocytes in the draining lymph node increased. During this period purified T lymphocytes did not express detectable levels of antigen, but enriched B cell populations expressed antigen transiently on day 1, 2, or 3 after exposure to antigen. The results showed that, during a 3-d period after exposure to antigen, DC expressed antigen and stimulated T cell proliferation. We speculate that low amounts of FITC binding selectively to veiled cells or lymph node DC in the first hours after exposure to antigen are not immunogenic but that Langerhans' cells acquire high levels of antigen, enter the nodes, and initiate immune responses.

Actin Cytoskeleton↗
PubMed1987

Susceptibility of human peripheral blood dendritic cells to infection by human immunodeficiency virus.

Preparations of human peripheral blood dendritic cells have been infected with human immunodeficiency virus (HIV). After 5 days in culture they were examined by electron microscopy. Virus was observed budding from the plasma membrane of dendritic cells and mature virions were observed on the cell surface. In addition, a second cell type, similar in morphology to 'classical' dendritic cells but containing numerous cytoplasmic granules, was also found to support replication of the virus. We speculate that the growth of HIV in dendritic cells could cause immunosuppression by impairing antigen presentation.

Cytoplasmic Granules↗