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Biomedical subjects

R W Dutton

Publications and source records attributed to R W Dutton.

At least 19 recordsLinked to original sources
PubMed2001

Activated antigen-specific CD8+ T cells persist in the lungs following recovery from respiratory virus infections.

The poor correlation between cellular immunity to respiratory virus infections and the numbers of memory CD8(+) T cells in the secondary lymphoid organs suggests that there may be additional reservoirs of T cell memory to this class of infection. Here we identify a substantial population of Ag-specific T cells in the lung that persist for several months after recovery from an influenza or Sendai virus infection. These cells are present in high numbers in both the airways and lung parenchyma and can be distinguished from memory cell populations in the spleen and peripheral lymph nodes in terms of the relative frequencies among CD8(+) T cells, activation status, and kinetics of persistence. In addition, these cells are functional in terms of their ability to proliferate, express cytolytic activity, and secrete cytokines, although they do not express constitutive cytolytic activity. Adoptive transfer experiments demonstrated that the long-term establishment of activated T cells in the lung did not require infection in the lung by a pathogen carrying the inducing Ag. The kinetics of persistence of Ag-specific CD8(+) T cells in the lung suggests that they play a key role in protective cellular immunity to respiratory virus infections.

Animals↗
PubMed2001

Immunopotentiating role of IFN-gamma in early and late stages of type 1 CD8 effector cell-mediated tumor rejection.

Type 1 cytolytic CD8 effector T cells (Tc1) characteristically secrete IFN-gamma. Using an OVA-transfected B16 melanoma lung tumor model, we show that OVA Ag-specific Tc1 cells mediate a reduction in tumor growth that significantly prolongs survival in tumor-bearing mice. Transfer of Tc1 cells from OT-I mice crossed to IFN-gamma-KO mice showed that IFN-gamma-deficient Tc1 effector cells were less therapeutically effective than corresponding cells from wildtype mice. Therapeutic effects were dependent, in part, on effector cell-derived IFN-gamma, which not only induced elevated levels of lung-derived IP-10 and RANTES chemokine message in vivo, but also increased the local accumulation of activated host-derived CD4(+)/CD44(High), CD8(+)/CD44(High), and non-T-immune cell populations at the tumor site. Over time, the numbers of host-derived immune cells increased in the lung, which correlated with an elevated production of IP-10 and RANTES and a continued reduction in tumor burden. Conversely, donor Tc1 cell numbers markedly diminished at corresponding times, suggesting that prolonged therapeutic responses were due to the presence of host-derived antitumor mechanisms. Moreover, adoptive transfer of IFN-gamma-deficient Tc1 cells into tumor-bearing IFN-gamma-KO recipients showed that both recipient and donor-derived IFN-gamma play a significant role in Tc1-mediated responses and that Tc1 effector cell immunotherapy is predominantly mediated by IFN-gamma-dependent mechanisms.

Adjuvants, Immunologic↗
PubMed2001

Production of interferon-gamma by influenza hemagglutinin-specific CD8 effector T cells influences the development of pulmonary immunopathology.

This study examined the inflammation, lung function impairment, and immune protection associated with either wild-type or interferon (IFN)-gamma-deficient Tc1- or Tc2-CD8 effector cells responding to influenza pneumonia. The adoptive transfer of influenza hemagglutinin-specific Tc1 effectors afforded protection and elicited only minimal impairment of lung function. IFN-gamma-deficient Tc1 effector cells were equally protective, but were associated with an eosinophil influx and slightly more lung function impairment early in the response. Relative to Tc1, Tc2 effector cells were less protective, elicited an eosinophil influx and a greater impairment of lung functions. IFN-gamma-deficient Tc2 effector cells were not protective and were associated with the severest impairment of lung function throughout the response, an accumulation of neutrophils, and extensive pulmonary vasculitis and alveolar hemorrhaging. Deletion of IFN-gamma was associated with a delay in effector cell recruitment and the elicitation of a more intense inflammatory response that resulted in more severe lung function impairment in the recipients of either Tc1 or Tc2 IFN-gamma-deficient effector cells. Thus, during influenza infections, IFN-gamma production by the responding CD8 T cells is associated with effector cell recruitment and mitigation of the associated inflammation and of the resulting impairment in lung functions but is not necessary for optimal protection.

Animals↗
PubMed2000

The effects of prolonged administration of 5-bromodeoxyuridine on cells of the immune system.

We have determined the in vivo effect of 5-bromodeoxyuridine (BrdU) administered to mice in the drinking water for various lengths of time on the performance of T and B lymphocytes in a number of experimental protocols. Young mice continuously exposed to BrdU fail to gain weight, and the lymphocytes recovered after a prolonged period of exposure are fewer in number than in control mice. The recovery of normal levels of T and B lymphocytes after irradiation is severely impaired. Ag-specific cells responding to Ag in an adoptive transfer model fail to expand as much in the presence of BrdU as in the absence, and the Ag-specific effectors produced in the presence of BrdU are less able to secrete cytokines upon restimulation in vitro. Polarized populations of Tc1 and Tc2 effectors generated in vitro proliferate less in the presence of BrdU, and the resulting effectors make less cytokines per cell upon restimulation. Thus, the incorporation of BrdU into T or B lymphocytes can, under some circumstances, seriously impair the performance of the labeled cells, and these findings raise a note of caution in the interpretation of studies that make use of long-term exposure to BrdU.

Administration, Oral↗
PubMed2000

Type 1 and type 2 CD8+ effector T cell subpopulations promote long-term tumor immunity and protection to progressively growing tumor.

Cytolytic CD8+ effector cells fall into two subpopulations based on cytokine secretion. Type 1 CD8+ T cells (Tc1) secrete IFN-gamma, whereas type 2 CD8+ T cells (Tc2) secrete IL-4, IL-5, and IL-10. Using an OVA-transfected B16 lung metastases model, we assessed the therapeutic effects of adoptively transferred OVA-specific Tc1 and Tc2 subpopulations in mice bearing established pulmonary malignancy. Effector cell-treated mice exhibiting high (5 x 105) tumor burdens experienced significant (p < 0.05) delays in mortality compared with those of untreated control mice, whereas high proportions (70-90%) of mice receiving therapy with low (1 x 105) tumor burdens survived indefinitely. Long-term tumor immunity was evident by resistance to lethal tumor rechallenge, heightened levels of systemic OVA Ag-specific CTL responses ex vivo, and detection of long-lived TCR transgene-positive donor cells accompanied by an elevation in the total numbers of CD8+ CD44high activated and/or memory T cells at sites of tumor growth. Long-lasting protection by Tc2 and Tc1 effector cells were dependent, in part, on both the level of tumor burden and effector cell-derived IL-4, IL-5, and IFN-gamma, respectively. We conclude that Tc1 and Tc2 effector cells provide immunity by different mechanisms that subsequently potentiate host-derived antitumor responses.

Adoptive Transfer↗
PubMed1999

Naive, effector, and memory CD8 T cells in protection against pulmonary influenza virus infection: homing properties rather than initial frequencies are crucial.

The goal of adoptive immunotherapy is to target a high number of persisting effector cells to the site of a virus infection or tumor. In this study, we compared the protective value of hemagglutinin peptide-specific CD8 T cells generated from the clone-4 TCR-transgenic mice, defined by different stages of their differentiation, against lethal pulmonary influenza infection. We show that the adoptive transfer of high numbers of Ag-specific unprimed, naive CD8 T cells failed to clear the pulmonary virus titer and to promote host survival. The same numbers of in vitro generated primary Ag-specific Tc1 effector cells, producing high amounts of IFN-gamma, or resting Tc1 memory cells, generated from these effectors, were protective. Highly activated CD62Llow Tc1 effectors accumulated in the lung with rapid kinetics and most efficiently reduced the pulmonary viral titer early during infection. The resting CD62Lhigh naive and memory populations first increased in cell numbers in the draining lymph nodes. Subsequently, memory cells accumulated more rapidly and to a greater extent in the lung lavage as compared with naive cells. Thus, effector cells are most effective against a localized virus infection, which correlates with their ability to rapidly distribute at the infected tissue site. The finding that similar numbers of naive Ag-specific CD8 T cells are not protective supports the view that qualitative differences between the two resting populations, the naive and the memory population, may play a major role in their protective value against disease.

Adoptive Transfer↗
PubMed1999

CD28, IL-2-independent costimulatory pathways for CD8 T lymphocyte activation.

We investigate, here, the mechanism of the costimulatory signals for CD8 T cell activation and confirm that costimulation signals via CD28 do not appear to be required to initiate proliferation, but provide survival signals for CD8 T cells activated by TCR ligation. We show also that IL-6 and TNF-alpha can provide alternative costimulatory survival signals. IL-6 and TNF-alpha costimulate naive CD8 T cells cultured on plate-bound anti-CD3 in the absence of CD28 ligation. They act directly on sorted CD8-positive T cells. They also costimulate naive CD8 T cells from Rag-2-deficient mice, bearing transgenic TCRs for HY, which lack memory cells, a potential source of IL-2 secretion upon activation. IL-6 and TNF-alpha provide costimulation to naive CD8 T cells from CD28, IL-2, or IL-2Ralpha-deficient mice, and thus function in the absence of the B7-CD28 and IL-2 costimulatory pathways. The CD8 T cell generated via the anti-CD3 plus IL-6 and TNF-alpha pathway have effector function in that they express strong cytolytic activity on Ag-specific targets. They secrete only very small amounts of any of the cytokines tested upon restimulation with peptide-loaded APC. The ability of the naive CD8 T cells to respond to TCR ligation and costimulatory signals from IL-6 and TNF-alpha provides a novel pathway that can substitute for signals from CD4 helper cells or professional APC. This may be significant in the response to viral Ags, which can be potentially expressed on the surface of any class I MHC-expressing cell.

Adjuvants, Immunologic↗
PubMed1999

Therapeutic effects of tumor-reactive type 1 and type 2 CD8+ T cell subpopulations in established pulmonary metastases.

Cytolytic CD8+ T cells fall into two subpopulations based on cytokine-secretion. Type 1 CD8+ cells (Tc1) characteristically secrete IFN-gamma, whereas type 2 CD8+ cells (Tc2) secrete IL-4 and IL-5. We assessed the relative therapeutic effects of adoptively transferred OVA-specific Tc1 and Tc2 CD8+ cells in mice bearing established OVA-transfected B16 melanoma lung metastases. Both Tc1 and Tc2 subpopulations mediated a reduction in lung tumor growth that subsequently prolonged survival times in mice with both early (day 7) and more advanced (day 14) levels of tumor development. CD8+ T cell populations recovered from spleens of tumor-bearing mice receiving Tc1 or Tc2 cells showed markedly enhanced tumor Ag-specific cytolytic and cytokine-releasing activities that correlated with delays in tumor cell growth and progression. Initially, both tumor-reactive Tc1 and Tc2 effector cells accumulated at the tumor site with nearly equal frequency. Tc1 cells persisted, whereas Tc2 cell numbers progressively diminished over time. Titration of Tc1 and Tc2 effector cells showed that protection was dose dependent with the former being 5-fold more effective. Tc2 cells achieved a comparable reduction in lung tumor cell growth at higher concentrations of cell transfer. Tc1 effectors from IFN-gamma-deficient mice were less therapeutically effective than wild-type mice, but there was no significant reduction in activity between corresponding Tc2 populations. We speculate that the effectiveness of Tc1 and Tc2 cells may depend on different mechanisms. These studies suggest a potential role for Tc1 and Tc2 CD8+ subpopulations in tumor regression and immunotherapy.

Adoptive Transfer↗
PubMed1999

Migration kinetics and final destination of type 1 and type 2 CD8 effector cells predict protection against pulmonary virus infection.

The requirements for CD8 T cells to provide protection against a localized virus infection in models of adoptive immunotherapy are not well defined. Here we investigated the protective value of defined in vitro-generated hemagglutinin (HA) peptide-specific primary CD8 T cell effectors from the clone 4 T cell receptor transgenic mice, secreting type 1 or type 2 cytokines, against pulmonary infection with whole influenza virus. Cytotoxic T lymphocytes producing type 1 and type 2 cytokine (Tc1 and Tc2) populations were equally cytolytic, but Tc1 effectors and not Tc2 effectors reduced the pulmonary virus titer early during infection. Host recovery mediated by Tc1 effectors was found to be independent of interferon gamma production. Tc2 effectors entered the lung with delayed kinetics as compared with Tc1 effectors, and after lung entry Tc2 effector cells did not localize near the infected airway epithelium as did Tc1 effectors but were found within clusters of inflammatory cells distant from the epithelium. We also show that the expression of several chemokine receptors was selectively regulated in the Tc1 and Tc2 subsets. Thus, the protective value of a CD8 cell population against pulmonary influenza virus infection is strongly correlated with its ability to exert its effector potential at the site of virus infection.

Animals↗
PubMed1998

In vivo persistence of CD8 polarized T cell subsets producing type 1 or type 2 cytokines.

Naive CD8 T cells can be polarized into effectors producing the type 1 cytokines IFN-gamma and IL-2 or the type 2 cytokines IL-4, IL-5, and IL-10, respectively. To study whether the polarized cytokine phenotype of the effectors is stable, we generated highly cytotoxic hemagglutinin (HA) peptide-specific CD8 Tc1 and Tc2 (cytotoxic CD8 T cells producing type 1 or type 2 cytokines) effectors from Clone-4 TCR-transgenic mice, which were adoptively transferred into syngeneic adult thymectomized irradiated and bone marrow-reconstituted recipients. The highly activated blast-size, CD25+ Tc1 and Tc2 effectors gave rise to homogeneous resting CD25- CD44(high) Ly6C(high) Ag-specific populations, which persisted for at least 13 wk after adoptive transfer. These memory CD8 T cells, recovered 13 wk after transfer of Tc1 or Tc2 effectors, still produced either the type 1 or type 2 cytokines, i.e., IFN-gamma, or IL-4 and IL-5, respectively, upon restimulation with APCs loaded with the HA peptide, but not in the absence of Ag. The amounts of IL-2 detected in the supernatants of Tc1 and Tc2 memory populations were comparable to that in memory CD4 cells, and both Tc1 and Tc2 memory cells became cytotoxic upon restimulation. Thus, cytokine-polarized CD8 memory T cells are a source of a variety of cytokines, which were classically considered helper cytokines, opening new perspectives on their function as regulatory cells in an immune response.

Adoptive Transfer↗
PubMed1998

T cell memory.

Immunological memory can be defined as the faster and stronger response of an animal that follows reexposure to the same antigen. By this definition, it is an operational property of the whole animal or the immune system. Memory cells express a different pattern of cell surface markers, and they respond in several ways that are functionally different from those of naive cells. Murine memory cells are CD44 high and low in the expression of activation markers such as CD25 (IL-2R), whereas human memory cells are CD45RA-, CD45RO+. In contrast to naive cells, memory cells secrete a full range of T cell cytokines and can be polarized to secrete particular restricted patterns of secretion for both CD4 and CD8 T cells. The requirements for the activation of memory cells for proliferation and cytokine production are not quite as strict as those of naive cells, but costimulation in the broad sense is required for optimum responses and for responses to suboptimum antigen concentrations. It would appear that memory cells can persist in the absence of antigenic stimulation and persist as nondividing cells. Reencounter with the same antigen can expand the population to a new, stable, higher level and generate a separate population of CD44 high effectors that may be required for protection, while competition from other antigens can drive it down to a lower stable level. It is unclear how or where memory cells arise, but once generated they have different pathways of recirculation and homing.

Animals↗
PubMed1997

Unequal death in T helper cell (Th)1 and Th2 effectors: Th1, but not Th2, effectors undergo rapid Fas/FasL-mediated apoptosis.

T helper cell (Th) 1, but not Th2, effectors undergo rapid Fas/Fas ligand (FasL)-mediated, activation-induced cell death upon restimulation with antigen. Unequal apoptosis is also observed without restimulation, after a longer lag period. Both effectors undergo delayed apoptosis induced by a non-Fas-mediated pathway. When Th1 and Th2 effectors are co-cultured, Th2 effectors survive preferentially, suggesting the responsible factor(s) is intrinsic to each population. Both Th1 and Th2 effectors express Fas and FasL, but only Th2 effectors express high levels of FAP-1, a Fas-associated phosphatase that may act to inhibit Fas signaling. The rapid death of Th1 effectors leading to selective Th2 survival provides a novel mechanism for differential regulation of the two subsets.

Animals↗
PubMed1995

Relative perforin- and Fas-mediated lysis in T1 and T2 CD8 effector populations.

IFN-gamma-secreting T1 and IL-5-secreting T2 subsets of CD8 effectors were generated in vitro using freshly isolated cells from wild-type and perforin knockout mice stimulated with allogeneic Ag-presenting cells. Both T1 and T2 effectors from wild-type mice exhibited perforin-mediated cytolysis. T1, but not T2, populations from perforin knockout mice displayed significant lysis by the Fas-mediated pathway. Th1 cells have recently been shown to be regulated by Fas and we speculate that Fas-mediated mechanisms are involved in the regulation of both Th1 and T1 populations of T cells.

Animals↗
PubMed1994

Generation of polarized antigen-specific CD8 effector populations: reciprocal action of interleukin (IL)-4 and IL-12 in promoting type 2 versus type 1 cytokine profiles.

We have generated primary effector populations from naive CD8 T cells in response to antigen and determined their patterns of cytokine secretion upon restimulation. The effect of exogenous factors on the effector generation was examined and compared with responses of antigen-specific CD4 effectors generated under comparable conditions. CD8 cells from bm1 mice were stimulated with C57BL/6 (B6) antigen presenting cells (APCs) bearing allogeneic class I and CD8 cells from female severe combined immunodeficiency (SCID) B6 mice, transgenic for a T cell receptor alpha/beta (TCR-alpha/beta) that recognizes H-Y on Db, were stimulated with APCs from male mice. In parallel, CD4 cells from bm12 mice were stimulated with alloantigen and CD4 cells from V beta 3/V alpha 11 TCR transgenics were stimulated with a peptide of pigeon cytochrome c on IEk. T cells from both transgenic mice were of naive phenotype whereas normal mice contained 10-20% memory cells. Effector CD8 populations generated were L-selectin low, CD45RB high, and CD44 high. Naive CD8 cells from SCID anti-H-Y mice made little or no cytokine immediately upon stimulation in contrast to naive CD4 which produced large amounts of interleukin 2 (IL-2). Both populations, however, generated primary effectors over 4-5 d that made substantial quantities of many cytokines upon restimulation. Both CD8 and CD4 effectors produced similar patterns of cytokines with alloantigen or specific antigen. Cytokines present during naive CD8 stimulation influenced the cytokine secretion profile of the effectors, as previously shown for CD4 cells, although secretion by CD8 effectors was generally lower than that of CD4 effectors. CD8 cells cultured with IL-2 alone made predominantly interferon gamma (IFN-gamma) and no IL-4 or IL-5, similar to CD4 cells. Priming with IFN-gamma increased IFN-gamma secretion from CD4 effectors, but had little if any effect on CD8 cells. In contrast, priming with IL-12 generated CD8 effectors, as well as CD4 effectors, producing elevated quantities of IFN-gamma, with similar levels from both the CD4 and CD8 populations. The presence of IL-4 during effector cell generation promoted synthesis of IL-4 and IL-5 from both CD8 and CD4 cells while downregulating IFN-gamma secretion. CD8 cells made only small amounts of IL-4, more than 100-fold less than CD4 cells, whereas significant levels of IL-5 were induced, only 3-10-fold lower than from CD4.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Monoclonal antibody that distinguishes between a phosphorylated, beta 2-microglobulin-associated, and a free, nonphosphorylated, chain of MHC class I.

We have shown that a mAb, 7.2.14, recognizes a conserved sequence in exon 7 of a number of murine MHC class I molecules. 7.2.14 binding is abolished when the molecule is phosphorylated, presumably at a serine residue in exon 7, whereas treatment of material in cell lysates with alkaline phosphatase increases the intensity of the binding. A genomic construct containing Dd was transfected into human fibroblasts and a clonal cell line expressing high levels of surface MHC was selected. Cell lysates were prepared from surface-iodinated cells and analyzed by using a panel of antibodies. An apparent size heterogeneity was detected in the MHC class I gene product precipitated by different anti-class I MHC antibodies, suggesting that more than one conformational species of Dd was present. This was further investigated regarding the molecules precipitated by antibodies 34-2-12, M1/42, and 7.2.14. After preclearing of surface-iodinated cell lysates by using one antibody, challenge with the others still precipitated a Dd molecule, confirming that there were three independent conformations of the Dd gene product. A similar complexity could be observed in the lysates of surface-labeled spleen cells from C57BL/6 mice. A major polypeptide at approximately 48 to 50 kDa, representing the MHC H chain, was seen, and one or two as yet unidentified but strongly associated polypeptides at 41 kDa and 56 kDa were also visible. Sequential clearing of surface-iodinated material with one antibody followed by precipitation with the other confirmed that the 7.2.14-reactive material was distinct from that which reacted with M1/42. We propose that the 7.2.14-reactive 50-kDa band is the nonphosphorylated form of class I MHC, which exists in a conformation different from that of the conventional 48-kDa, phosphorylated, beta 2-microglobulin-associated entity.

Amino Acid Sequence↗