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J W Murphy

Publications and source records attributed to J W Murphy.

At least 37 records · Page 2Linked to original sources
PubMed1998

Type 1 and type 2 cytokines: from basic science to fungal infections.

At the present time, it is clear that Th1 responses afford protection against the fungi; however, the development, maintenance and function of the protective immune responses are complex mechanisms and are influenced by multiple factors. The route of infection has been shown to affect initial cytokine production and, consequently, the induction of protective Th1 responses. The ability of different isolates of the same fungal agent to induce and sustain a protective response has also been emphasized. Protective immune responses have been shown to vary in genetically different mouse strains after infection. In addition, these protective responses, such as cellular influx and cytokine production, also vary within the same animal depending on the tissue infected. The functional dominance of certain cytokines over others in influencing development and maintenance of protective responses has been discussed. Certain cytokines may act differently in hosts lacking important components of their innate or immune repertoire. It is evident from these presentations that a more comprehensive understanding of the protective mechanisms against different fungal agents is emerging. However, there is still much to learn before cytokine modulatory therapy can be used effectively without risk in the human host.

Animals↗
PubMed1997

MIP-1 alpha contributes to the anticryptococcal delayed-type hypersensitivity reaction and protection against Cryptococcus neoformans.

Leukocyte infiltration into infected tissues is essential for the clearance of microorganisms. In animals with a cell-mediated immune (CMI) response to the infectious agent, as opposed to naive animals, leukocyte migration is greatly enhanced into sites of the organism or antigen. The role of the,chemotactic cytokine or chemokine, macrophage inflammatory protein-1 alpha (MIP-1 alpha), in the expression phase of the CMI response and in protection against Cryptococcus neoformans was assessed. With the use of a gelatin sponge model in mice as a means of detecting an anti-cryptococcal delayed-type hypersensitivity (DTH) reaction, we found that MIP-1 alpha levels in fluids from cryptococcal antigen (CneF)-injected sponges in immunized mice (DTH-reactive sponges) were significantly increased over levels of MIP-1 alpha in fluids from saline-injected control sponges at 12 and 24-30 h after injection. MIP-1 alpha levels peaked before increases in neutrophils and lymphocytes in the DTH-reactive sponges, suggesting that MIP-1 alpha was responsible, at least in part, for attracting these leukocyte types. Immunized mice treated with neutralizing antibody to MIP-1 alpha before sponge injection with CneF had reduced numbers of neutrophils and lymphocytes in the DTH-reactive sponges and showed reduced clearance of C. neoformans from the lungs, spleens, livers, and brains when compared with controls. Furthermore, injection of rmMIP-1 alpha into sponges in naive mice resulted in an increase in the influx of neutrophils and lymphocytes into the sponges compared with saline-injected sponges. Together our findings provide solid evidence that MIP-1 alpha is a component of the anticryptococcal DTH reaction. In addition, MIP-1 alpha influences neutrophil influx and attracts lymphocytes into the DTH reaction site. Finally, we showed that MIP-1 alpha plays a role in protection against C. neoformans.

Animals↗
PubMed1997

Kinetics of cellular infiltration and cytokine production during the efferent phase of a delayed-type hypersensitivity reaction.

Cell-mediated immunity is a primary host resistance mechanism against many infectious organisms and is responsible for leucocyte recruitment to the infection site. Delayed-type hypersensitivity (DTH) reactions are in vivo correlates of cell-mediated immunity and have long been used to assess the level of cell-mediated immune (CMI) responsiveness to specific antigens. It has been difficult to study the kinetics of cellular influx and cytokine composition at the site of an on-going CMI reaction. Consequently, knowledge of the sequential events occurring during the efferent phase of a CMI response is incomplete. Here we report on the use of a gelatin sponge model for evaluating the progression of events during the effector phase of a DTH reaction to antigens of the mycotic organism Cryptococcus neoformans. Previously, we have shown that 24 hr after antigen injection into sponges in infected or immune mice, the leucocyte types infiltrating the sponges are consistent with a classical murine DTH reaction. Through kinetic studies, we show here that neutrophils are the first leucocytes to appear in DTH-reactive sponges, followed by increases in lymphocyte and then monocyte numbers. Tumour necrosis factor (TNF), interleukin-2 (IL-2), interferon-gamma (IFN-gamma) and IL-5 were elevated in DTH-reactive sponges compared with control sponges, and each cytokine had a relatively unique temporal profile. IL-4 was not detectable in the sponges. Together our data indicate that the expression of a CMI response comprises a well-regulated sequential influx of leucocytes that contribute to the lymphokine composition of the reaction.

Animals↗
PubMed1997

Secretion of the C3 component of complement by peritoneal cells cultured with encapsulated Cryptococcus neoformans.

Two isolates of Cryptococcus neoformans were identified as being widely divergent in pathogenic potential after intratracheal infection of mice. These isolates differed in their ability to upregulate capsule synthesis when grown under tissue culture conditions, and this property correlated with virulence. We postulated that differential capsule synthesis may cause differential stimulation of macrophages to produce products such as complement components. To test this hypothesis, heat-killed yeast cells were incubated with normal mouse peritoneal cells (PC) before the level of C3 secreted was determined. Cryptococcal stimulants were grown on mycological agar, which does not promote capsule synthesis, or in RPMI 1640 at 37 degrees C in an atmosphere of 5% CO2, which stimulates capsule synthesis, to determine the role that the capsule plays in the induction of C3 secretion. C3 levels were elevated in cultures containing cryptococci grown in RPMI 1640 at 37 degrees C in an atmosphere of 5% CO2, and the level of C3 detected was correlated with the amount of capsule expressed by the yeast cell stimulant. Nonencapsulated mutants of C. neoformans did not stimulate C3 secretion. Purified capsular polysaccharide (glucuronoxylomannan [GXM]) also stimulated the PC to secrete C3. Two signals were required before GXM stimulated C3 secretion. The second signal was identified as endotoxin present in small amounts (0.06 ng per ml) in tissue medium. Endotoxin may provide a priming stimulus for PC to express receptors or other cytokines needed for effective stimulation of C3. These experiments show that enhancement of C3 secretion by C. neoformans is due to GXM and is correlated with the virulence of the cryptococcal isolate.

Animals↗
PubMed1997

Direct interactions of human natural killer cells with Cryptococcus neoformans inhibit granulocyte-macrophage colony-stimulating factor and tumor necrosis factor alpha production.

Human natural killer (NK) cells and T lymphocytes can bind to and inhibit the growth of the yeast-like organism Cryptococcus neoformans. Binding of target cells to NK or T cells also has the potential to modulate cytokine production by the effector cells. In this study, we assessed the ability of C. neoformans to modulate NK cell production, or in some cases T-cell production, of granulocyte-macrophage colony-stimulating factor (GM-CSF) or tumor necrosis factor alpha (TNF-alpha). We found that freshly isolated human NK cells from most individuals make GM-CSF and TNF-alpha constitutively when cultured in vitro. The addition of C. neoformans to T-cell fractions which do not make GM-CSF constitutively did not affect GM-CSF production, but the addition of C. neoformans to NK cell fractions significantly reduced the amounts of GM-CSF produced in most NK cell samples. The reduction in the amount of GM-CSF in C. neoformans-NK cell cocultures could not be attributed to loss of lymphocyte viability or to C. neoformans adsorbing or degrading the cytokine and was dependent on direct contact between the NK cells and cryptococcal cells. GM-CSF was not the only cytokine to be down-regulated. TNF-alpha production was also diminished when NK cells were incubated with C. neoformans. The regulation of both cytokines was at the transcriptional level because GM-CSF and TNF-alpha mRNA levels were lower in NK cell samples incubated with C. neoformans than in NK cell samples incubated without C. neoformans. Diminished production of constitutively produced cytokines resulting from the interaction of NK cells with cryptococcal cells has the potential to affect phagocytic cells in the immediate regional environment and to damp the immune response.

Cryptococcus neoformans↗
PubMed1997

Cryptococcal polysaccharides bind to CD18 on human neutrophils.

CD18, the beta chain of the beta 2 integrin family of adhesion molecules, is associated with three different alpha chains (CD11a, -b, and -c) and is expressed on the surface of all types of leukocytes. CD18-containing molecules are up-regulated on the surface of neutrophils (polymorphonuclear cells [PMN]) in response to chemotactic agents and are implicated in mediating adhesion to an inflamed endothelium, which is a prerequisite to migration of PMN into infected tissues. In a previous study, we found that a cryptococcal culture filtrate (CneF), when injected into the bloodstream of mice to simulate the antigenemia in cryptococcosis, inhibits PMN accumulation at the site of an inflammatory stimulus. In the present study, we assessed the ability of CneF and its individual components, i.e., glucuronoxylomannan (GXM), galactoxylomannan (GalXM), and mannoprotein (MP), to interact with CD18 on human PMN. CneF labeled with 14C was shown to bind to human PMN in a dose-dependent manner. Pretreatment of PMN with anti-CD18, but not an isotype-matched control monoclonal antibody (MAb) or anti-CD11a MAb, blocked the binding of 14C-labeled CneF to PMN. In addition, CneF, GXM, and GalXM but not MP significantly blocked the binding of the anti-CD18 MAb to CD18 on the surface of unactivated and formyl methionyl leucyl phenylalanine-activated PMN as determined by indirect immunofluorescence staining and flow cytometric analysis. In the same experiments, the cryptococcal polysaccharides did not affect the binding of an anti-CD11a or anti-L-selectin MAb to the surface of PMN at 4 degrees C. The results suggest that CneF and its components GXM and GalXM bind to CD18 on human PMN. Based on our findings, we propose that CD18 is a possible molecular target of cryptococcal polysaccharides and that binding of the polysaccharides to CD18 has the potential to inhibit leukocyte infiltration into inflammatory sites.

Antibodies, Monoclonal↗
PubMed1996

Cryptococcal polysaccharides induce L-selectin shedding and tumor necrosis factor receptor loss from the surface of human neutrophils.

High titers of cryptococcal polysaccharides in the serum and spinal fluid and the lack of cellular infiltrates in the infected tissues are hallmarks of disseminated cryptococcosis. Cryptococcal polysaccharides given intravenously to mice inhibit the influx of leukocytes into sites injected with inflammatory mediators. The purpose of this investigation was to determine if cryptococcal polysaccharides, i.e., glucuronoxylomannan (GXM), galactoxylomannan, and mannoprotein, affect expression of molecules on the surface of neutrophils that are important in extravasation. GXM in the absence of serum was shown to induce human neurophils to shed L-selectin, a molecule needed in the first step of neutrophil movement into tissues. In the presence of serum, GXM caused a further shedding of L-selectin. Shedding of L-selectin was evident by reduced amounts of L-selectin on the neutrophils treated with GXM and by increased levels of soluble L-selectin in the GXM-treated neutrophil supernatants. GXM also stimulated neutrophils to have reduced expression of TNF receptor. In contrast, GXM-treated neutrophils showed increased levels of CD15 and CD11b, and unchanged CD16 expression. In the absence of serum, galactoxylomannan and mannoprotein did not affect L-selectin, TNF receptor, CD15, CD11b, or CD16 on neutrophils but did induce loss of L-selectin in the presence of serum. Our results indicate that cryptococcal polysaccharides, especially GXM, can cause shedding of L-selectin from the surface of neutrophils, and this may prevent neutrophils from attaching to the endothelial cell surfaces. Blockage of this early step in cell migration from the vessels into tissues may be responsible in part for reduced cellular infiltration into infected tissues of individuals with disseminated cryptococcosis.

Antigens, CD↗
PubMed1995

K562 cell strains differ in their response to poliovirus infection.

Poliovirus readily establishes a persistent infection in the K562-Mu erythroleukemia cell strain. In this study, three additional K562 cell strains were analyzed for their responses to poliovirus infection and found to be quite variable. K562 cells obtained from the ATCC established a persistent infection, similar to the K562-Mu cell strain, while the majority of cells from two other strains, K562-KI and K562-We, were killed by 4 or 11 days postinfection (p.i.), respectively. Several characteristics of the uninfected and infected cell strains were examined to determine if differences existed which could explain the dramatically different responses to infection. Since K562 cell strains can differentiate toward several cell lineages, the four strains were analyzed for physical and functional likeness to the original K562 cell line using well-established functional criteria to determine whether gross changes in differentiation state had occurred. Based on the lack of MHC class I antigen expression and a dose-dependent increase in globin synthesis in response to hemin, all three laboratory K562 cell strains were indistinguishable from the ATCC reference strain. Surface poliovirus receptor levels were also similar in all K562 cell strains, although four- to fivefold lower than those in HeLa cells. Most biochemical events in virus replication either were very similar among K562 cells or were slightly variable and did not correlate with the degree of cell killing. These included levels of virus production, levels of viral protein produced, and processing and turnover of viral polypeptides. The key difference between the cell strains which consistently correlated with cell killing was the degree of virus-induced host translation shutoff, which was always greatest in the most virus-sensitive K562-KI cells. In addition, levels of 2Apro produced in K562 cell strains did not appear to correlate with the levels of host protein shutoff. A related and novel finding in these studies which also strongly correlated with the outcome of infection was the ability of levels of intact p220 to recover by 24 hr p.i. in virus-resistant K562-Mu and -ATCC cells. These data suggest that the key determinants of outcome of infection in this cell model are cytoplasmic host factors related to cytopathology and not factors which may modulate levels of viral protein synthesis or RNA synthesis.

Blotting, Northern↗
PubMed1995

Direct antimicrobial activity of T cells.

T cells are generally thought to contribute to antimicrobial activity either by releasing lymphokines, which recruit and activate other cell types, or by major histocompatibility complex (MHC)-restricted lysis of infected host cells. Recently, it has become apparent that T cells can also mediate antimicrobial activity by direct interaction with microbial targets. Such interactions, which can be either antigen specific or nonspecific, occur in the apparent absence of MHC restriction and do not require the presence of other host cells. Microbial targets recognized by T cells include fungi, parasites and bacteria. Here, Stuart Levitz, Herbert Mathews and Juneann Murphy discuss the direct antimicrobial activity of T cells and speculate on its in vivo relevance.

Animals↗
PubMed1995

Intravascular cryptococcal culture filtrate (CneF) and its major component, glucuronoxylomannan, are potent inhibitors of leukocyte accumulation.

Disseminated cryptococcosis is characterized by high titers of cryptococcal polysaccharides in serum and minimal cellular infiltrates in infected tissues of patients. The main objective of this study was to determine whether the circulating cryptococcal polysaccharides could contribute to the lack of cellular infiltration into infected tissues. To assess this possibility, a gelatin sponge implantation model was used. We found that intravenous (i.v.) injection of mice with cryptococcal culture filtrate antigen (CneF) inhibited migration of leukocytes (neutrophils, lymphocytes, and monocytes) into the intrasponge sites of acute inflammation induced by CneF, tumor necrosis factor alpha, or formylmethionyl leucyl phenylalanine. In addition, i.v. administration of CneF inhibited leukocyte migration into the intrasponge sites of a cell-mediated immune reaction irrespective of whether the delayed-type hypersensitivity response was to CneF or the mycobacterial antigen purified protein derivative. Glucuronoxylomannan, a major constituent of CneF and a major cryptococcal antigen detected in the sera of patients with disseminated cryptococcosis, when given i.v. to mice, inhibited leukocyte migration into the sponges. Our results suggest that the minimal cellular infiltrates observed in infected tissues of cryptococcosis patients may be due, in part, to the circulating cryptococcal polysaccharide functioning as we have demonstrated in the mouse model. Furthermore, the high titers of cryptococcal antigen in the sera of patients may diminish leukocyte migration in response to stimuli other than Cryptococcus neoformans, a point that may be relevant in AIDS patients with cryptococcosis.

Animals↗
PubMed1995

Direct anticryptococcal activity of lymphocytes from Cryptococcus neoformans-immunized mice.

Assessment of the direct anticryptococcal activity of murine lymphocytes from both Cryptococcus neoformans-immunized and control mice was the focus of this investigation. We demonstrate that at a 2:1 effector cell-to-cryptococcal target cell ratio, effector cell populations comprised of alpha beta T-cell receptor-positive T lymphocytes (98 to 99% CD3+) from C. neoformans-immunized mice inhibited the growth of cryptococcal cells better than similar populations of lymphocytes from nonimmunized control mice. Almost immediately after mixing of cryptococci with the effector cells, C. neoformans-lymphocyte conjugates were observed. The percentage of conjugates increased over the first 30 min of incubation and then remained constant over the next 1.5 h. T-lymphocyte-enriched populations from C. neoformans-immunized mice formed significantly greater percentages of conjugates with cryptococci than control T lymphocytes at each time period that assessment was made. For growth inhibition to occur, direct contact between the effector and target cells was necessary, as evidenced by abrogation of cryptococcal growth inhibition when lymphocyte and cryptococcal cell populations were separated by a porous membrane during the growth inhibition assay. Vital staining of cryptococci after incubation with the T-cell-enriched populations showed that the T lymphocytes killed the cryptococcal cells.

Animals↗
PubMed1995

Effects of immunization with Cryptococcus neoformans cells or cryptococcal culture filtrate antigen on direct anticryptococcal activities of murine T lymphocytes.

Immunizing CBA/J mice with intact Cryptococcus neoformans cells or with a cryptococcal culture filtrate antigen (CneF) induces an anticryptococcal delayed-type hypersensitivity response. Recently, it has been shown that two phenotypically different T-cell populations are responsible for delayed-type hypersensitivity reactivity in mice immunized with intact cryptococcal cells, whereas only one of those populations is present in mice immunized with soluble cryptococcal antigens in complete Freund's adjuvant (CFA). The purpose of this study was to determine if differences occur with regard to direct anticryptococcal activity between T-lymphocyte-enriched populations from mice immunized with intact viable or dead cryptococcal cells and similar cell populations from mice immunized with the soluble cryptococcal culture filtrate antigen, CneF, emulsified in CFA. The percentage of lymphocytes which form conjugates with C. neoformans and the percentage of cryptococcal growth inhibition in vitro are greater with T-lymphocyte-enriched populations from mice sublethally infected with C. neoformans or from mice immunized with intact heat-killed cryptococcal cells in the presence or absence of CFA than with lymphocyte populations from mice immunized with CneF-CFA. Enhanced anticryptococcal activity of T lymphocytes could be induced by immunizing mice with heat-killed C. neoformans cells of serotype A, B, C, or D as well as by immunizing with a similar preparation of an acapsular C. neoformans mutant but not by immunizing with CFA emulsified with CneF prepared from any one of the C. neoformans isolates. These data indicate that the soluble cryptococcal culture filtrate antigens do not induce the same array of functional T lymphocytes as whole cryptococcal cells.

Animals↗
PubMed1995

Effects of the two varieties of Cryptococcus neoformans cells and culture filtrate antigens on neutrophil locomotion.

Cryptococcus neoformans var. gattii (serotype B and C) isolates have a relative predilection for immunocompetent hosts, and C. neoformans var. neoformans (serotype A and D) isolates have a relative predilection for immunocompromised hosts, suggesting that normal host resistance to the former may be relatively inefficient compared with that to the latter variety. In order to assess the possibility that normal cellular host defense is inadequate in protecting against C. neoformans var. gattii, we compared the two varieties of C. neoformans cells and their culture filtrate antigens (CneF) with respect to effects on neutrophil (polymorphonuclear leukocyte [PMN]) locomotion. In a 48-well modified Boyden chamber, the cells and CneF of C. neoformans var. neoformans (serotype A and D) isolates stimulated chemotaxis and chemokinesis of human PMN and activated a complement component(s) in pooled human serum to become a chemoattractant(s) for human PMN. In contrast, the cells and CneF of C. neoformans var. gattii (serotype B and C) isolates did not stimulate chemotaxis or chemokinesis in human PMN but rather inhibited chemokinesis and chemotactic responses of PMN to pooled human serum and formylmethionyl leucyl phenylalanine. Neither of the CneF from the C. neoformans var. gattii isolates was cytotoxic to PMN. Furthermore, with the mouse model, we found that CneF from C. neoformans var. neoformans caused migration of PMN into gelatin sponges implanted in naive and immunized mice, whereas CneF from C. neoformans var. gattii inhibited PMN migration into sponges. Our results, combined with findings of others showing reduced PMN infiltration in lungs of mice infected with C. neoformans var. gattii compared with PMN infiltration in lungs of mice infected with C. neoformans var. neoformans, indicate that the relative inadequacy of normal host resistance mechanisms to prevent infection with C. neoformans var. gattii results, in part, from inhibition of PMN migration to the site of the organism.

Animals↗
PubMed1995

Factors associated with premature termination of psychotherapy by children.

This study reports findings on factors associated with children who terminate psychotherapy prematurely. The sample was composed of 45 children, with an average age of 13.5 years. It was found that children are more likely to remain in treatment if the therapist is perceived as professional and caring, and if the treatment plan is adequately explained.

Adolescent↗
PubMed1994

Cytokines in the host response to mycotic agents.

In summary, different approaches have been taken to understand cytokine responses to different fungal infections. Singer-Vermes and co-investigators indirectly examined cytokine responses to paracoccidioidomycosis by studying the types of cellular and humoral immune responses that were induced in resistant and susceptible mouse strains. Their results implicated Th1 cell responses in the resistant mouse strain and Th2 cell responses in the mouse strain susceptible to paracoccidioidomycosis. By measuring cytokine production and through cytokine depletion experiments, Wu-Hsieh showed that besides IFN gamma, TNF alpha was important in host defences against the intracellular pathogen, H. capsulatum. Both cytokines play important roles in the regulation of other cytokines. In histoplasmosis, the dynamics of the complex interactions amongst cytokines govern the efficiency of host clearance of the fungus from tissues. Ferrante and collaborators, examining TNF alpha and TNF alpha receptors on neutrophils presented data showing that TNF alpha plays an important role in the activation of neutrophils for anti-Candida activity. Through the detection of cytokine mRNAs with RT-PCR, Moser and co-workers found that cytokine mRNAs of macrophage origin were produced preferentially in the lungs of mice infected with Histoplasma or Blastomyces. A great challenge still lies ahead of us. It is well understood that the interactions of cytokines are extremely complex at the levels of the induction and expression of the immune responses as well as on effects on natural cellular defences. Work accomplished thus far has laid the ground work for future studies in the effort to dissect host cytokine responses and to understand the roles of cytokines in protection against fungal infections.

Animals↗
PubMed1994

Regulation of cytokine production during the expression phase of the anticryptococcal delayed-type hypersensitivity response.

Effects of both positive and negative regulatory T cells on cellular infiltration and cytokine production during the expression phase of the anticryptococcal immune response were examined. Tamp cells, which are induced by cryptococcal antigen, significantly amplify the anticryptococcal delayed-type hypersensitivity response, whereas a cascade of T suppressor (Ts) cells inhibits the response and decreases the clearance of Cryptococcus neoformans during an infection. By using the gelatin sponge implantation model, we found that Tamp cells do not stimulate a significant increase in cellular infiltration into the sponges in response to cryptococcal antigen compared with that into delayed-type hypersensitivity-reactive sponges in immune control mice. However, Tamp cells do stimulate significant increases in the production of gamma interferon and interleukin-2 (IL-2) in the antigen-injected sponges over the level of the representative cytokine in antigen-injected sponges from the immune control mice. Likewise, Ts1 cells, induced with cryptococcal antigen, do not significantly affect antigen-induced cellular infiltration into sponges in immune mice. In contrast, decreased levels of gamma interferon and IL-2 are observed in antigen-injected sponges from Ts1-cell-recipient, immunized mice compared with those of the positive immune controls. The presence of either Tamp or Ts1 cells in immunized mice stimulates increased production of IL-5 but not IL-4 over that of the positive immune controls.

Animals↗