Cryptococcus neoformans and Candida albicans regulate CD4 expression on human monocytes

This study examined the capability of Candida albicans and Cryptococcus neoformans to modulate CD4 expression on human monocytes. C. albicans and an acapsular strain of C. neoformans induced higher levels of CD4 expression than encapsulated strains. Purified glucuronoxylomannan did not regulate CD4 expression on monocytes, but down-regulation of CD4 expression compared with stimulation by acapsular C. neoformans alone was observed when glucuronoxylomannan was used in combination with acapsular C. neoformans. The ability of opsonic factors to facilitate fungal-mediated CD4 overexpression suggests that binding or internalization (or both) of the yeast cells is a critical event. Protein synthesis was required, excluding redistribution of the intracellular pool of CD4 receptors to the cellular surface as the sole possible mechanism. Results demonstrate a new effect of fungi on professional phagocytic cells and raise the possibility that modulation of CD4 could influence gp120-mediated human immunodeficiency virus entry.     

Reduced IL-12 production by monocytes upon ultraviolet-B irradiation selectively limits activation of T helper-1 cells

The capacity of APC to stimulate the proliferation of human peripheral blood T cells decreases upon ultraviolet-B (UVB) irradiation. The aim of this study was to investigate whether all T cell subsets are equally sensitive to this reduced APC function. Established human Th1, Th2, and Th0 clones were stimulated with monocytes in a soluble CD3 mAb-mediated assay that is dependent on the presence of APC. Monocytes were exposed to low nonlethal doses of UVB radiation before coculture with T cells. UVB irradiation inhibited the capacity of monocytes to stimulate the proliferation and IFN-gamma production of Th1 cells in a dose-related fashion. In contrast, UVB-treated monocytes induced normal proliferation and IL-4 production in Th2 cells. Stimulation of Th0 cell proliferation by UVB-irradiated monocytes was normal, but a preferential suppression of IFN-gamma production was observed, thus leading to a more Th2-like cytokine response. The loss of Th1 proliferation upon stimulation with UVB-irradiated monocytes could be overcome by rIL-2; however, IFN-gamma production remained suppressed. IFN-gamma production could be completely restored by rIL-12, whereas the addition of IL-1 beta, TNF-alpha, or indomethacin had no such effect, nor did the addition of mAb to CD28, added to compensate for the reduced B7 expression of UVB-irradiated monocytes. Monocytes exposed to UVB radiation exhibited reduced expression of mRNA for the IL-1 2 subunits p35 and p40 and suppressed production of the IL-12 p70 protein. Our results thus indicate that UVB irradiation of APC selectively impairs Th1-like responses, a phenomenon caused by the UVB-induced suppression of monocyte IL-12 production.     

IL-15 promotes IL-12 production by human monocytes via T cell-dependent contact and may contribute to IL-12-mediated IFN-gamma secretion by CD4+ T cells in the absence of TCR ligation

At inflammatory sites, the number of activated bystander T cells exceeds that of Ag-activated T cells. We investigated whether IL-15, a monocyte-derived cytokine that shares several biologic activities with IL-2, may contribute to bystander T cell activation in the absence of IL-2 and triggering Ag. The addition of IL-15 to cocultures of monocytes and T cells stimulates CD4+ but not CD8+ T cells to produce IFN-gamma. IFN-gamma production requires endogenous IL-12, the production of which in turn is dependent upon CD40/CD154 interactions between CD4+ T cells and monocytes. Indeed, non-TCR-activated CD4+ but not CD8+ T cells express significant levels of CD154. IL-15 may enhance IFN-gamma in this system by up-regulating CD40 expression on monocytes and IL-12Rbeta1 expression on CD4+ T cells. Conversely, using neutralizing anti-IL-15 mAb, we show that the ability of IL-12 to augment IFN-gamma secretion is partly mediated by endogenous IL-15. Finally, in the absence of monocytes, a synergistic effect between exogenous IL-12 and IL-15 is necessary to induce IFN-gamma production by purified CD4+ T cells, while IL-15 alone induces T cell proliferation. It is proposed that this codependence between IL-12 and IL-15 for the activation of inflammatory T cells may be involved in chronic inflammatory disorders that are dominated by a Th1 response. In such a response, a self-perpetuating cycle of inflammation is set forth, because IL-15-stimulated CD4+ T cells may activate monocytes to release IL-12 that synergizes with IL-15 to induce IL-12 response and IFN-gamma production.     

Direct activity of human T lymphocytes and natural killer cells against Cryptococcus neoformans

Lymphocytes constitute a critical component of host defenses against cryptococcosis. Previously, we demonstrated that human lymphocytes cultured with interleukin-2 formed conjugates with, and directly inhibited the growth of, Cryptococcus neoformans. Here, we explore the anticryptococcal activity of freshly isolated, highly purified populations of human peripheral blood lymphocytes. Lymphocytes were incubated with encapsulated C. neoformans for 24 h, after which the lymphocytes were lysed, dilutions and spread plates were made, and CFU were counted. Fungistasis was determined by comparing growth in wells with and without lymphocytes. Nylon wool-nonadherent peripheral blood mononuclear cells (NWNA PBMC) were highly fungistatic, even if either T cells or natural killer (NK) cells were depleted by panning. A mixed population of T cells and NK cells, obtained by rosetting NWNA PBMC with sheep erythrocytes, completely inhibited cryptococcal growth, whereas the nonrosetting cells had little fungistatic activity. CD4+, CD8+, and CD16/56+ lymphocytes, isolated by positive immunoselection, had potent growth-inhibitory activity. In contrast, purified B cells had no activity. Fungistasis was seen even in the absence of opsonins. Antifungal activity was markedly diminished when surface receptors on NWNA PBMC were cleaved by treatment with trypsin or bromelain. Supernatants from stimulated lymphocytes or concentrated lymphocyte sonicates were not active. Lymphocyte-mediated fungistasis was seen with two different strains of C. neoformans. CD4+, CD8+, and CD16/56+ lymphocytes formed conjugates with C. neoformans, as observed under Nomarski differential interference contrast microscopy and videomicroscopy. These data demonstrate that freshly isolated peripheral blood T cells and NK cells have the capacity to bind and directly inhibit the growth of C. neoformans.     

T cell-derived IL-4 and dendritic cell-derived IL-12 regulate the lymphokine-producing phenotype of alloantigen-primed naive human CD4 T cells

Previous studies on human Th subset development were restricted to the analysis of naive T cells activated with anti-CD3 mAb in the absence of physiologic APC. In this study, we have analyzed the role of cytokines and physiologic APC on T cell maturation in an Ag-specific system, in which naive neonatal CD4 T cells were primed with allogeneic dendritic cells (DC). We found that the cytokine profile of primed cells was dependent upon 1) the ratio between T cells and allogeneic DC and 2) the endogenous production of IL-4 and IL-12. Neutralization of IL-4 during primary MLR increased IFN-gamma production at priming and shifted the phenotype of primed cells from Th0 to Th1. These effects were IL-12 dependent, in that they were suppressed by anti-IL-12 Abs. The production of IL-12 in primary MLR was further evidenced by the presence of IL-12 p40 in the culture supernatant fluids. IL-12 production was suppressed by exogenous IL-4 and increased by anti-IL-4 blocking mAbs, indicating that endogenous IL-4 down-regulated IL-12 production by DC. Finally, IL-12 was produced as a result of T cell/DC interaction involving the CD40/CD40 ligand and CD28/B7 costimulation pathways, as revealed by the inhibitory effect of anti-CD40 ligand mAb and CTLA-4Ig. These observations suggest that in neutral conditions, Ag presentation by DC results in the coordinate production of naive T cell-derived IL-4 and DC-derived IL-12 that in concert shape the cytokine profile of Th cells.     

Impaired production of IL-12 in systemic lupus erythematosus. I. Excessive production of IL-10 suppresses production of IL-12 by monocytes

Currently, primary osteoporosis is the most frequent metabolic disease in women after menopause [1]. The resulting loss of bone mass is accompanied by an increased risk of skeletal fragility. One reason for the development of osteoporosis might be an impaired function of mature osteoblasts. To evaluate the involvement of specific growth factors in bone remodeling, cell cultures of osteoblastic cells derived from nonosteoporotic and osteoporotic postmenopausal women were established. The influences of TGF beta-1 and IGF-I on proliferation and mRNA expression of TGF beta-1 were investigated by [3H]-thymidine incorporation and competitive RT-PCR. We found IGF-I to have no significant effect on proliferation in cells of osteoporotic and nonosteoporotic patients. In contrast, differences were found in TGF beta-1 mRNA expression after application of IGF-I. Application of TGF beta-1 enhanced its own mRNA expression in both groups in a similar manner. Whereas the proliferation of cells of nonosteoporotic patients was inhibited by (10(-10) M) TGF beta-1, this treatment led to an increased proliferation of cells of osteoporotic patients.     

Encapsulation of Cryptococcus neoformans with glucuronoxylomannan inhibits the antigen-presenting capacity of monocytes

PURPOSE: The combination of carmustine (BCNU), dacarbazine (DTIC), cisplatin (DDP), and tamoxifen (Tam) has been reported in small series to provide a response rate of 50%, but with significant myelosuppression and risk of thromboembolic complications. We performed this phase II study to assess the antitumor activity and important toxicities of this combination in the cooperative group setting. PATIENTS AND METHODS: Seventy-nine eligible patients were treated with BCNU 150 mg/m2/d, every 6 weeks, DTIC 220 mg/m2/d on days 1 to 3 every 3 weeks, DDP 25 mg/m2/d on days 1 to 3 every 3 weeks, and Tam 20 mg orally daily throughout treatment. Treatment cycles were repeated every 6 weeks in responding or stable patients for a maximum duration of 1 year. RESULTS: Twelve objective responses were achieved (response rate 15%, 95% confidence interval 8%-25%). Five responses were complete (CR) and seven were partial (PR). The median response duration was 8+ (range, 4-19+) months, (16+ [4-19+] for CR and 8+ [4-11] for PR), and the median survival of the entire group was 9 months. The toxicities were predominantly neutropenia and thrombocytopenia. Four patients developed thromboembolic events. Two patients died while on protocol therapy, one with complications of neutropenia, and the other with disease progression. CONCLUSION: The activity of this regimen is in the range reported for single agents or DTIC plus DDP, and the addition of BCNU and Tam appears to increase toxicity. We do not recommend this combination for routine treatment of advanced melanoma or as the control arm in randomized studies of combination therapy.     

Effects of a corticosteroid, budesonide, on production of bioactive IL-12 by human monocytes

Although Corynebacterium minutissimum is well-known as the cause of erythrasma, it is noted as the etiologic agent of nondermatologic disease only rarely. We document this organism as a cause of central venous catheter-associated bacteremia and report the use of pulsed-field gel electrophoresis to characterize its molecular epidemiology.     

Increased production of interleukin-10 by human blood monocytes stimulated with Mycobacterium avium-intracellulare complex

BACKGROUND: Overseas screening of immigrants and refugees applying for a visa to the United States identifies foreign-born individuals who are at high risk for tuberculosis (TB) or who have active TB. The system's effectiveness relies on further medical evaluation and follow-up of foreign-born individuals after their arrival in the United States. METHODS: Retrospective cohort study of 893 immigrants and refugees who arrived in the United States from July 1, 1992, through December 31, 1993, with a destination of San Francisco, Calif, and a referral for further medical evaluation. MAIN OUTCOME MEASURES: Time to report to the local health department after arrival and the yield of active and preventable cases of TB from follow-up medical evaluations. RESULTS: Median time from arrival in the United States to seeking care in San Francisco was 9 days (range, 1-920 days). Of 745 immigrants and refugees (83.4%) who sought further medical evaluation, 51 (6.9%) had active TB and 296 (39.7%) were candidates for preventive therapy. Being a refugee was an independent predictor of failure to seek further medical evaluation in the United States. Class B-1 disease status based on overseas TB screening (odds ratio, 3.5; 95% confidence interval, 2.0-6.2) and being from mainland China (odds ratio, 4.4; 95% confidence interval, 1.9-9.9) were independent predictors of TB diagnosed in San Francisco. CONCLUSIONS: Timely, adequate medical evaluation and follow-up care of immigrants and refugees has a relatively high yield and should be a high priority for TB prevention and control programs.     

Differential production of IL-10 by T cells and monocytes of HIV-infected individuals: association of IL-10 production with CD28-mediated immune responsiveness

The present study determines the proportions of unmyelinated cutaneous axons at the dermal-epidermal junction in glabrous skin and of myelinated and unmyelinated axons in the sural and medial plantar nerves that immunostain for subunits of the ionotropic glutamate receptors. Approximately 20% of the unmyelinated cutaneous axon profiles at the dermal-epidermal junction immunostain for either N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), or kainate receptor subunits. These findings are consistent with previous observations that NMDA and non-NMDA antagonists ameliorate nociceptive behaviors that result from noxious peripheral stimulation. In the sural nerve, where the large majority of myelinated fibers are sensory, approximately half of the myelinated axon profiles immunostain for the NMDA receptor 1 (R1) subunit, 28% immunostain for the glutamate receptor 1 (GluR1) AMPA subunit, and 11% for the GluR5,6,7 kainate subunits. Even higher proportions immunostain for these receptors in the medial plantar nerve, a mixed sensory and motor nerve. In the sural nerve, 20% of the unmyelinated axon profiles immunostain for NMDAR1 and only 7% label for GluR1 or GluR5,6,7. Because the sural nerve innervates hairy skin, these data suggest that glutamate will activate a higher proportion of unmyelinated axons in glabrous skin than in hairy skin. Measurements of fiber diameters indicate that all sizes of myelinated axon profiles, including Adelta and Abeta, are positively labeled for the ionotropic receptors. The presence of glutamate receptors on large-diameter myelinated axons suggests that these mechanosensitive receptors, presumably transducing touch and pressure, may also respond to local glutamate and thus be chemosensitive.     

CD4-mediated inhibiton of IL-2 production in activated T cells

The role of CD4 in T cell activation has been attributed to its capacity to increase the avidity of interaction with APC and to shuttle associated Lck to the TCR/CD3 activation complex. The results presented in this study demonstrate that ligation of CD4 inhibits ongoing responses of preactivated T cells. Specifically, delayed addition of CD4-specific mAb is shown to inhibit Ag- or mAb-induced responses of both primary T cells and T cell clonal variants. The Ag responses of the latter are independent of the adhesion provided by CD4; thus the observed inhibition is not due to blocking CD4-MHC interactions. Further, analysis of the clonal variants demonstrates that CD4-associated Lck is not essential for the inhibition observed, as anti-CD4 inhibits responses of clonal variants, expressing a form of CD4 unable to associate with Lck (double cysteine-mutated CD4). The inhibition is counteracted by the addition of exogenous IL-2, demonstrating that the block is not due to a lesion in IL-2 utilization, rather its production. It is demonstrated that the delayed addition of anti-CD4 results in a rapid reduction in steady-state levels of IL-2 mRNA in both primary T cells and clonal variants.     

Urocanic acid enhances IL-10 production in activated CD4+ T cells

The immunosuppressive effects of UV radiation have been well documented. This suppression has been attributed to the action of the cis form of urocanic acid (UCA), a photoproduct of trans-UCA, a natural constituent of the skin. Here, we show that mouse spleen cells preincubated with cis-UCA have a diminished proliferative response to allogeneic cells in MLC and to stimulation with anti-CD3 mAb. Cells preincubated with cis-UCA also had a decreased ability to serve as APC and to stimulate the proliferation of allogeneic lymphocytes in MLC. Simultaneously, the production of IL-2 and IFN-gamma by cells preincubated with cis-UCA was decreased. However, IL-10 gene expression and IL-10 protein secretion by spleen cells stimulated in the presence of cis-UCA were significantly enhanced. The principal cell population displaying the cis-UCA-induced elevated production of IL-10 was CD4+ T cells, which were shown to be a direct target of cis-UCA action. This was also supported by the observation that production of IL-10 by stimulated splenic non-T cells or by macrophages was not altered by cis-UCA. The enhanced production of IL-10 by activated CD4+ T cells may represent a novel pathway of UVB radiation-induced, cis-UCA-mediated immunosuppression. We suggest that the elevated production of IL-10 by activated CD4+ T cells may account for the suppressor T cell phenomena described in UV-irradiated recipients.     

Differential stimulation of interleukin-12 (IL-12) and IL-10 by live and killed Helicobacter pylori in vitro and association of IL-12 production with gamma interferon-producing T cells in the human gastric mucosa

The objective of these experiments was to examine the ability of Helicobacter pylori to stimulate interleukin-10 (IL-10) or IL-12 and select for either Th1 or Th2 cells. Gastric biopsy specimens were collected from patients who were categorized with respect to the presence of H. pylori and gastric disease as well as their age, gender, medications, and other factors. As Th1 and Th2 cells are selected by IL-12 and IL-10, respectively, biopsy specimens were screened for mRNA and protein for these cytokines. Although mRNA for IL-12 and IL-10 was detected in biopsy specimens obtained from both infected and uninfected patients, IL-12 protein predominated. Levels of IL-10 and IL-12 in gastric tissue did not change in response to infection. Moreover, gamma interferon (IFN-gamma)-producing T cells were found in both the infected and the uninfected gastric mucosa. Stimulation of peripheral blood leukocytes from either infected or uninfected donors with various concentrations of live or killed H. pylori induced immunoreactive IL-12 and IL-10. After stimulation with live H. pylori, IL-12 levels increased more than 30-fold, whereas IL-10 levels increased only 2- to 5-fold, compared to cells stimulated with medium alone. Interestingly, killed H. pylori induced significantly more IL-10 (P < 0.05) than live H. pylori, while recombinant urease only induced IL-10. These results demonstrate that live H. pylori selectively stimulates the induction of IL-12 and Th1 cells that produce IFN-gamma, whereas preparations used in oral vaccines induce more IL-10 and may favor Th2 cell responses.     

Antigen-stimulated IL-4, IL-13 and IFN-gamma production by human T cells at a single-cell level

To understand the intricate balance and the coordinate expression of the Th1 and Th2 cytokines following a natural mode of T cell triggering, antigen-stimulated IL-4, IL-13 and IFN-gamma production was studied in primary peripheral blood mononuclear cell cultures at a single-cell level. Cells from filariasis patients who respond to parasite antigen by producing not only IFN-gamma but also IL-4 and IL-13 were stimulated with Brugia malayi adult worm antigen and analyzed for co-expression of cytokines by intracellular staining. IL-4 and IL-13 were frequently co-expressed (54% of IL-4+ cells stained for IL-13 and 29% of IL-13+ cells expressed IL-4 at all time points), whereas IFN-gamma expression was totally segregated from both IL-4 and IL-13. These data indicate that in human peripheral T cells the co-expression of the dominant Th1 and Th2 cytokines within a single cell is a rare event and that IL-13 is clearly more frequently associated with a Th2 than a Th1 type response in primary T cell cultures.     

IL-12 up-regulates IL-18 receptor expression on T cells, Th1 cells, and B cells: synergism with IL-18 for IFN-gamma production

IL-18 is a product of macrophages and with IL-12 strikingly induces IFN-gamma production from T, B, and NK cells. Furthermore, IL-18 and 1L-12 synergize for IFN-gamma production from Th1 cells, although this combination fails to affect Th2 cells. In this study, we show that IL-12 and IL-18 promptly and synergistically induce T and B cells to develop into IFN-gamma-producing cells without engaging their Ag receptors. We also studied the mechanism underlying differences in IL-18 responsiveness between Th1 and Th2 cells. Pretreatment of T or B cells with IL-12 rendered them responsive to IL-18, which induces cell proliferation and IFN-gamma production. These IL-12-stimulated cells had both high and low affinity IL-18R and an increased IL-18R mRNA expression. In particular, IL-12-stimulated T cells strongly and continuously expressed IL-18R mRNA. However, when T cells developed into Th1 cells after stimulation with anti-CD3 and IL-12, they lowered this IL-12-induced-IL-18R mRNA expression. Then, such T cells showed a dominant response to anti-CD3 by IFN-gamma production when they were subsequently stimulated with anti-CD3 and IL-18. In contrast, Th2 cells did not express IL-18R mRNA and failed to produce IFN-gamma in response to anti-CD3 and IL-18, although they produced a substantial amount of IFN-gamma in response to anti-CD3 and IL-12. However, when Th1 and Th2 cells were stimulated with anti-CD3, IL-12, and IL-18, only the Th1 cells markedly augmented IFN-gamma production in response to IL-18, suggesting that IL-18 responsiveness between Th1 and Th2 cells resulted from their differential expression of IL-18R.     

The cell wall and membrane of Cryptococcus neoformans possess a mitogen for human T lymphocytes

The mechanism of human T-lymphocyte activation by the pathogenic yeast Cryptococcus neoformans has not been established. Previous investigations have suggested that C. neoformans contains a mitogen for T lymphocytes, while other investigators have attributed lymphocyte proliferation in vitro to a recall antigen. Because of the potential importance of the mechanism of T-cell activation for our understanding of the immune response to C. neoformans, the present studies were performed to determine whether C. neoformans contains a mitogen for T lymphocytes. C. neoformans stimulates fetal blood lymphocytes to proliferate and stimulates proliferation of CD45RA+ cells from adults, indicating that it stimulates naive T cells. The T-cell response to C. neoformans was dependent upon the presence of accessory cells. However, allogeneic cells were sufficient for accessory cell function, indicating that the response was not major histocompatibility complex restricted. The percentage of T cells in the cell cycle was higher than that with the recall antigen tetanus toxoid but lower than that with the mitogenic lectin phytohemagglutinin A or the superantigen Staphylococcus enterotoxin B. Precursor frequency analysis established that 1 in 7,750 +/- 2, 270 T cells proliferated in response to the cryptococcal cell wall and membrane. Compared to the case for most mitogens or superantigens, the proliferative response is late and the number of T cells that enter the cell cycle and the precursor frequency are low, indicating that the mitogenic effect is modest. However, the mitogenic effect of C. neoformans should be considered when interpreting the immune response to C. neoformans, since even weak mitogens can have profound effects on host defense.     

Characterization of anticapsular monoclonal antibodies that regulate activation of the complement system by the Cryptococcus neoformans capsule

PURPOSE: To evaluate the effectiveness of adding interferon (IFN) alfa-2b to chemotherapy in the induction treatment of low-grade non-Hodgkin's lymphoma (NHL), and to assess the role of maintenance IFN. PATIENTS AND METHODS: A multicenter, two-phase controlled trial with double randomization was conducted in 155 patients with low-grade NHL. In the first randomization, 78 patients received cyclophosphamide, vincristine, and prednisone (CVP) and IFN, 3 MU/m2 three times a week for 3 months, and 77 patients received CVP alone. Responding patients were randomized to receive IFN for 1 year versus observation. RESULTS: Of 144 assessable patients, 73 received CVP + IFN and 71 received CVP. Responses were similar: CVP + IFN 79% versus CVP 76% (P = .62). The number of patients who did not complete the treatment was higher in the CVP + IFN group than in the CVP group (18% v 4%; P = .009), although the received dose-intensity of chemotherapy was comparable. Duration of response and progression-free survival (PFS) were significantly higher in the CVP + IFN group than in the CVP group (P = .0004). However, we observed no differences in overall survival (OS) (P = .30), with a median follow-up for the surviving patients of 3 years. Grade 3/4 granulocytopenia was the most frequent toxicity and was similar in both groups (33% v32%). Eighty-three (74%) of the 112 responding patients were randomized to maintenance IFN or observation. The duration of response was similar between 42 patients that received IFN compared with 41 control patients (P = .83), independently of treatment previously administered. CONCLUSION: Adding IFN alfa-2b to induction CVP in low-grade NHL did not induce a higher response rate, but it significantly increased the duration of the responses. We found significant differences in PFS that favored the patients who received CVP + IFN, but not in OS. To date, no additional benefit has been seen from the administration of IFN for maintenance.