Intracellular mechanisms responsible for exercise-induced suppression of macrophage antigen presentation.

In a previous study, we demonstrated that exhaustive exercise suppressed peritoneal macrophage antigen presentation (AP). In this study, we explored the intracellular mechanism(s) responsible for this suppression. Pathogen-free male BALB/c mice (8 +/- 2 wk) were randomly assigned to either home cage control (HCC) or exhaustive exercise stress (Exh, 18-30 m/min for 3 h/day) treatment groups. The mice underwent treatments for a period of 4 days during induced peritoneal thioglycollate inflammation. Elicited macrophages were harvested, purified, and incubated with chicken ovalbumin (C-Ova, 2. 5 and 10 mg/ml) for 18 h. After macrophages were washed, they were cocultured with C-Ova-specific T cells for 48 h at which time the supernates were harvested and analyzed via ELISA for interleukin (IL)-2 as an indication of macrophage AP. There was no significant (P > 0.05) difference in macrophage AP between cells fixed with paraformaldehyde vs. those that remained unfixed, suggesting that Exh did not affect production of soluble factors influencing macrophage AP (i.e., IL-1, IL-4, PGE(2)). The ability of macrophages to generate C-Ova immunogenic peptides was analyzed using FITC-labeled C-Ova, which shows fluorescence only when degraded intracellularly. There was a significant ( approximately 20%, P < 0. 05) suppression in fluorescence in the Exh compared with HCC, indicating a possible defect in the ability of macrophages from Exh to degrade C-Ova into immunogenic peptides. Macrophages were also incubated with C-Ova immunogenic peptide in a manner identical to that for native C-Ova. We found a similar suppression ( approximately 22-38%, P < 0.05) in macrophage AP using a C-Ova peptide when compared with native C-Ova in the Exh group, indicating reduced major histocompatibility complex (MHC) II loading and/or C-Ova-MHC II complex cell surface expression. In conclusion, these data indicate an intracellular defect in the macrophage antigen processing pathway induced by Exh.     

Leishmania-induced inhibition of macrophage antigen presentation analyzed at the single-cell level

A number of studies have previously examined the capacity of intracellular Leishmania parasites to modulate the capacity of macrophages to process and present Ags to MHC class II-restricted CD4(+) T cells. However, the bulk culture approaches used for assessing T cell activation make interpretation of some of these studies difficult. To gain a more precise understanding of the interaction between Leishmania-infected macrophages and effector T cells, we have analyzed various parameters of T cell activation in individual macrophage-T cell conjugates. Leishmania-infected macrophages efficiently stimulate Ag-independent as well as Ag-dependent, TCR-mediated capping of cortical F-actin in DO.11 T cells. However, infected macrophages are less efficient at promoting the sustained TCR signaling necessary for reorientation of the T cell microtubule organizing center and for IFN-gamma production. A reduced ability to activate these T cell responses was not due to altered levels of surface-expressed MHC class II-peptide complexes. This study represents the first direct single-cell analysis of the impact of intracellular infection on the interaction of macrophages with T cells and serves to emphasize the subtle influence Leishmania has on APC function.     

Leishmania donovani affects antigen presentation of macrophage by disrupting lipid rafts

Leishmania donovani-infected splenic macrophages and P388D1 (P388D1(I)) failed to activate T cells in response to low dose of exogenous peptide. The membrane fluidity of P388D1(I) was greater than that of the normal counterpart P388D1(N), but could be reduced either by exposing the cell below phase transition point or by loading cholesterol into membrane (L-P388D1(I)), and this was associated with enhanced Ag-presenting ability of P388D1(I). Presentation of endogenous leishmanial Ag, kinetoplastid membrane protein-11, was also defective, but could be corrected by loading cholesterol into membrane. Because membrane rafts are important for Ag presentation at a low peptide dose, raft architecture of P388D1(I) was studied using raft (CD48 and cholera toxin-B) and non-raft (CD71) markers in terms of their colocalization with I-A(d). Binding of anti-CD48 mAb and cholera toxin B subunit decreased significantly in P388D1(I), and consequently, colocalization with I-A(d) was not seen, but this could be restored in L-P388D1(I). Conversely, colocalization between I-A(d) and CD71 remained unaffected regardless of the presence or the absence of intracellular parasites. P388D1(N) and L-P388D1(I), but not P388D1(I), formed peptide-dependent synapse with T cells quite efficiently and this was found to be corroborated with both intracellular Ca2+ mobilization in T cells and IL-2 production. This indicated that intracellular parasites disrupt the membrane rafts, possibly by increasing the membrane fluidity, which could be corrected by making the membrane rigid. This may be a strategy that intracellular L. donovani adopts to evade host immune system.     

Species-dependent regulation of monocyte/macrophage Ia antigen expression and antigen presentation by prostaglandin E

The expression of Ia antigen by various murine and human macrophage populations and the ability of prostaglandins of the E series to regulate Ia antigen expression were explored. Monocytes and macrophages from human and murine populations demonstrated a dichotomy in the expression of Ia antigen. Both human monocytes and macrophages expressed elevated levels of Ia antigen compared to their murine counterpart. Murine macrophages appear to express elevated levels of Ia antigen only when actively interacting with T lymphocytes in vivo or with lymphokines in vitro. Prostaglandins of the E series can suppress murine macrophage Ia antigen expression, but have little effect on the expression of Ia antigen by human monocytes and macrophages. Also, prostaglandins of the E series do not modulate the ability of human monocytes to present antigen to autologous lymphocytes when studied over a broad concentration range. These data suggest that prostaglandin E compounds do not profoundly affect human monocyte/macrophage Ia antigen expression or human monocyte antigen presenting activity.     

Enhanced antigen presentation using human Fc gamma receptor (monocyte/macrophage)-specific immunogens.

A major new challenge for vaccine development is to target APC such as monocytes and macrophages for efficient Ag processing and presentation. It has been shown that Fc gamma R-mediated uptake of Ag-antibody complexes can enhance Ag presentation by myeloid cells at least 100-fold, and directing Ag to Fc gamma R in mice brings about a substantial increase in the effectiveness of immunization while eliminating the requirement for adjuvant. It has not been determined which of the three subclasses of human Fc gamma R on myeloid cells (Fc gamma RI, Fc gamma RII, or Fc gamma RIII) function to enhance Ag presentation. We have targeted our Ag (TT) to each of the three subclasses of human Fc gamma R on monocytes using Fc gamma R subclass-specific mAb-TT conjugates, and have measured TT presentation by monitoring T cell proliferation in response to TT. In addition, we have examined enhanced Ag presentation mediated by a human IgG1 (HIgG1) anti-TT mAb. All anti-Fc gamma R-TT conjugates enhanced Ag presentation. HIgG1 anti-TT, in monomeric form, enhanced Ag presentation through Fc gamma RI only. Anti-Fc gamma RI-Ag conjugates appear to be optimal for application as vaccines. They are monocyte/macrophage-specific, are very efficiently processed and presented, and enhance Ag presentation despite occupation of Fc gamma RI with HIgG.     

delta 9-Tetrahydrocannabinol suppresses macrophage extrinsic antiherpesvirus activity.

The effect of delta 9-tetrahydrocannabinol (THC), the major psychoactive component of marijuana, on macrophage intrinsic and extrinsic antiherpesvirus activities was examined. THC had no effect on the capacity of the macrophage-like cells RAW264.7, J774A.1, and P388D1 to take up virus. In addition, replication of virus within macrophages did not occur regardless of drug treatment, indicating that THC had no effect on macrophage intrinsic antiviral activity. In contrast, the cannabinoid exerted a dose-dependent inhibition of macrophage extrinsic antiviral activity. This activity describes that macrophage function by which these cells suppress virus replication within xenogeneic cells in an interferon-independent manner. The inhibitory effect of THC on extrinsic antiviral activity was greatest on RAW264.7 and J774A.1 cells, followed by P388D1 cells. These macrophage-like cells regained their extrinsic antiviral activity in a time-related fashion following removal of the drug. These results indicate that THC inhibits macrophage extrinsic antiherpesvirus activity, but has no effect on intrinsic antiviral activity. However, the suppressive effect of THC on extrinsic antiviral activity is reversible upon removal of the drug.     

Antigen presentation the macrophage way

Multiphoton intravital microscopy of lymph nodes has revolutionized the study of the early events leading to induction of acquired immunity. Using this technique, Junt et al. (2007) report in a recent issue of Nature that macrophages of the mouse lymph node subcapsular sinus facilitate B cell activation in vivo by collecting and displaying native antigen.     

MHC class II-restricted presentation of intracellular antigen.

An endogenously produced immunoglobulin light chain (lambda 2(315] is processed and presented to T cells in association with major histocompatibility complex (MHC) class II molecules. Using transfectants producing variant forms of lambda 2(315) that are neither expressed on the cell surface nor secreted, we demonstrate that intracellular lambda 2(315), which has never been exported outside of the cell, is the source of processed lambda 2(315) idiotype. This challenges the currently accepted paradigm that endogenous antigens are only presented by MHC class I molecules. Variants of lambda 2(315) protein that are retained in the endoplasmic recticulum (ER) are also presented. Variants that are expressed in the cytosol as well as those that are transported into the nucleus rather than the ER are not presented. Thus, the ER is likely to be the processing compartment.     

Selective macrophage ascorbate deficiency suppresses early atherosclerosis

To test whether severe ascorbic acid deficiency in macrophages affects progression of early atherosclerosis, we used fetal liver cell transplantation to generate atherosclerosis-prone apolipoprotein E-deficient (apoE(-/-)) mice that selectively lacked the ascorbate transporter (SVCT2) in hematopoietic cells, including macrophages. After 13 weeks of chow diet, apoE(-/-) mice lacking the SVCT2 in macrophages had surprisingly less aortic atherosclerosis, decreased lesion macrophage numbers, and increased macrophage apoptosis compared to control-transplanted mice. Serum lipid levels were similar in both groups. Peritoneal macrophages lacking the SVCT2 had undetectable ascorbate; increased susceptibility to H(2)O(2)-induced mitochondrial dysfunction and apoptosis; decreased expression of genes for COX-2, IL1β, and IL6; and decreased lipopolysaccharide-stimulated NF-κB and antiapoptotic gene expression. These changes were associated with decreased expression of both the receptor for advanced glycation end products and HIF-1α, either or both of which could have been the proximal cause of decreased macrophage activation and apoptosis in ascorbate-deficient macrophages.     

Autophagy and antigen presentation

CD4(+) T cells co-ordinate adaptive immunity and are required for immunological memory establishment and maintenance. They are thought to primarily recognize extracellular antigens, which are endocytosed, processed by lysosomal proteases and then presented on major histocompatibility complex (MHC) class II. However, recent studies have demonstrated that viral, tumour and autoantigens can gain access to this antigen presentation pathway from within cells by autophagy. This review will discuss the autophagic pathways that contribute to endogenous MHC class II antigen processing. Furthermore, potential characteristics of autophagy substrates, qualifying them to access these pathways, and regulation of autophagy will be considered. Finally, I will suggest how antigen presentation after autophagy might contribute to immune surveillance of infected and transformed cells.     

Common principles in protein folding and antigen presentation.

The regular recurrence of hydrophobic amino acid residues along a peptide sequence determines the formation of a longitudinal hydrophobic strip when the peptide forms an alpha-helix. An understanding of the ways this may affect both folding of nascent proteins and antigen presentation should facilitate vaccine and therapeutics design.     

Host glycans and antigen presentation

The cell-mediated adaptive immune response depends upon the activation of T cells via recognition of antigen in the context of a major histocompatibility complex (MHC) molecule. Although studies have shown that alterations in T cell receptor glycosylation reduces the activation threshold, the data on MHC is far less definitive. Here, we discuss the data on MHC glycosylation and the role the glycans might play during the adaptive host response.     

Intracellular interference with antigen presentation

Murine peritoneal macrophages were cultured with heat-killed Listeria monocytogenes organisms and then with the protein hen egg white lysozyme. Hen egg lysozyme is well known to need intracellular processing for presentation to T cells. The presentation to T cells of lysozyme was affected despite no reduction in the amount taken up or catabolized by the macrophage. This interference with Ag presentation was not found if the macrophages were cultured with lysozyme before the Listeria pulse. The interference with Ag presentation induced by Listeria was found for a second Ag (conalbumin). Uptake of Listeria did not affect the presentation of the lysozyme peptide 46-61, indicating that MHC class II molecules were available at the macrophage surface. Other materials that are retained in the macrophages affected presentation of lysozyme but not of the processed peptide. These included SRBC, dextran, sucrose, cellobiose, polyvinyl pyrrolidone, sodium dextran sulfate, Ficoll, and polyethylene glycol. Except for SRBC, which were not tested, the remaining molecules did not interfere with presentation of 46-61 by formaldehyde-fixed macrophages, an indication that they did not affect the peptide interaction with class II molecules. Finally, uptake of latex beads did not affect presentation of lysozyme. We conclude that retention in the macrophage of a variety of soluble or particulate molecules can interfere with the intracellular events that result in the creation of an immunogenic determinant. This interference is independent of the catabolism of the Ag or of the availability of class II molecules to bind peptides.