Control of IgE responses. V. Oral administration of a synthetic derivative of the inner bacterial cell wall (SDZ 280.636) to sensitized mice induces isotype specific suppression of peak hapten specific IgE antibody forming cell responses, serum IgE levels and immediate hypersensitivity responses.

SDZ 280.636, a nontoxic diacyl glycerol derivative of muramyl dipeptide (MDP), a component of the inner bacterial cell wall, which is suitable for use in man, suppressed hapten specific IgE antibody forming cell (AFC) responses in spleen, serum levels of hapten specific IgE and hapten specific immediate hypersensitivity (i.h.) responses in skin, when fed to mice at the peak of a hapten specific IgE AFC response. In addition, serum levels of IL-6 appeared increased while IFN gamma was decreased. To induce these IgE responses, BALB/c mice were injected i.p. with BPO-KLH (benzylpenicilloyl-keyhole limpet hemocyanin) (10 micrograms) in aluminum hydroxide gel (alum) on days 0, 21 and 42. Mice were fed (gavage) with either MDP or SDZ 280.636 (1.0 or 10 mg/kg) on day 44, or on days 44, 46 and 48, and killed on days 46 or 50. Numbers of BPO specific AFC in spleen, and serum levels of BPO specific immunoglobulins (IgG1, IgE and IgA) were determined (ELISPOT assay, ELISA). In addition, BPO specific IH responses were measured in these animals. Mice were injected in the right pinna with BPO-BSA (0.1 microgram) and in the left pinna with an equal volume of saline (0.05 ml). At 2 hr, pinnae were measured using a micrometer caliper. We found that 1 feeding with either MDP or SDZ 280.636 abrogated IgE AFC responses and dramatically suppressed serum levels of IgE, both in isotype specific fashion, and suppressed IH responses (> 50%). 3 feedings with SDZ 280.636 also abrogated IgE AFC responses and further decreased serum levels of IgE. In contrast to SDZ 280.636, 3 treatments with MDP had opposite effects in that IgE AFC responses and serum levels of IgE dramatically increased. A single treatment with SDZ 280.636 appeared to increase serum levels of IL-6 up to three fold, while IFN gamma levels decreased. Our data suggest that SDZ 280.636 may be useful in the therapeutic and prophylactic management of human atopic disease such as allergic rhinitis, asthma, and other atopic diseases.

Suppression of in vivo IgE and tissue IL-4 mRNA induction by SDZ 280.636, a synthetic muramyl dipeptide derivative.

Modulation of IgE isotype expression on B cells is one of the numerous effects of muramyl peptides on the regulation of the immune system. A non toxic diacyl glycerol derivative of muramyl dipeptide (MDP), in which the L-alanine is replaced by L-threonine (MDP-Threo-GDP; SDZ 280.636), is currently under investigation as lead compound for the development of an anti-allergic drug. In this report, the modulatory effect of orally administered SDZ 280.636 in a murine model on polyclonally induced IgE levels is described. In this model, mice are injected i.v. with goal anti mouse IgD (GAMD) and challenged three to four weeks later with goal IgG (GIG). Both the primary and secondary immune responses lead to an increase of serum IgE levels. We demonstrate the efficacy of this muramyl dipeptide derivative in selectively inhibiting a polyclonal IgE response in GAMD-primed, GIG challenged mice without affecting the levels of other immunoglobulin classes. It is further shown that the induction of interleukin 4 (IL-4) gene transcript levels in lymphoid organs, which is observed as a consequence of boosting GAMD pretreated mice with GIG, is selectively suppressed in gut associated lymphoid tissues (GALT) and mesenteric lymph nodes but not in spleen. In contrast, interleukin 13 (IL-13) mRNA levels are not affected by SDZ 280.636. The findings that SDZ 280.636 inhibits polyclonal IgE responses and suppresses IL-4, but not IL-13 mRNA expression point towards differences in the regulatory pathways of IL-4 and IL-13 gene transcription in lymphoid organs. Thus the mechanism of action appears to involve a specific suppression of IL-4 gene transcription in cells occurring in Peyer's patches and mesenteric lymph nodes which are among the first constituents of the immune system encountered by an orally administered drug.

Kinetics and isotype profile of rheumatoid factor production during viral infection: organ distribution of antibody secreting cells.

The kinetics and isotype profile of influenza virus-specific IgG antibodies were studied in correlation with the serum titre of IgG-reactive autoantibodies. An increased level of IgG isotype-specific, rheumatoid factor-type autoantibody secretion was observed in the late phase of the virus-specific memory response. These rheumatoid factors were specific for the IgG2a and IgG1 subclasses which dominated the anti-viral antibody response. As revealed by a preparative immunosorbent technique combined with isotype quantitation the majority of IgG2a- or IgG1-bound immunoglobulins isolated from the serum of virus-infected mice belonged to the same subclass as the target antibody. Comparison of the kinetics of appearance and the number of IgM-, IgG- and IgA-type IgG2a-reactive autoantibody secreting cells during the primary and memory anti-viral antibody responses showed isotype switch of IgM rheumatoid factor secreting cells predominantly to IgA. Localization of IgM and IgA antibody secreting cells demonstrated the wide organ distribution of IgM-type rheumatoid factor secreting cells. On the contrary, IgA rheumatoid factor production was observed only in Peyer's patches and at the site of the local virus-specific immune response, i.e. in mediastinal lymph nodes and in the lung. These results demonstrate that B cells specific for self IgG are activated and differentiated in concert with the viruspecific antibody response in similar microenvironments. The predominant involvement of the mediastinal lymph nodes and the spleen in the production of IgG2a-specific IgM-type autoantibodies suggest a regulatory function of this type of autoantibodies in modulating IgG2a production in both systemic and local anti-viral immune responses. The results also suggest a strictly regulated rheumatoid factor production which, however, can be unbalanced by repeated viral infections resulting in the escape of high affinity, isotype-switched autoantibodies.

Oral administration of muramyl dipeptide into mice modulates cell proliferation, immunoglobulin synthesis and cytokine mRNA levels in gut associated lymphoid tissues.

Muramyl peptides (MDPs) are synthetic immunostimulants capable of potentiating a multitude of immune functions following parenteral administration into a host. The parent molecule MDP was also found to exert certain activities when applied via the oral route. Thus, we have studied the effect of oral treatment of mice with MDP on the lymphoproliferative responses, immunoglobulin secretion and cytokine induction in gut-associated lymphoid tissues (GALT) employing lymphocyte transformation test, ELISA and RT-PCR, respectively. Cells derived from Peyer's patches (PP) of mice orally primed with MDP were found to have enhanced proliferative responses to different mitogens and to secrete significantly more IgG, IgM and IgA immunoglobulins than cells from unprimed mice. These effects were not observed with cells derived from mesenteric lymph nodes (MLN) or spleens of MDP-primed mice. However, oral administration of MDP resulted in the up-regulation of interleukin-6 (IL-6) mRNA in MLN and down-regulation of IL-4 and tumour necrosis factor-alpha (TNF-alpha) mRNAs in MLN, spleens and PP. These studies suggest that selective modulations of GALT responses by orally administered MDP are achievable and imply that these agents may be useful in the therapy and prophylaxis of allergic diseases.

Some aspects of IL-8 pathophysiology. III: Chemokine interaction with endothelial cells.

Chemokines have been convincingly implicated in driving leukocyte emigration in different inflammatory reactions. However, the cellular and molecular mechanisms of chemokine involvement in leukocyte emigration are not clear. We and others suggested that leukocyte adhesion to the endothelium and transmigration are induced by chemokines immobilized on the endothelial cell surface. This would require the presence of specific chemokine binding sites in this microanatomical location. Using an in situ binding assay we demonstrated the presence of binding sites for interleukin-8 (IL-8) and RANTES, but not monocyte inflammatory protein-1 alpha on the endothelium of postcapillary venules and small veins in human skin. In contrast, venules and veins in various anatomical locations showed dramatically differing IL-8 binding patterns. The subcellular distribution of IL-8 in the venular endothelial cells following its in vivo and ex vivo injections was studied by use of electron microscopy. Our results suggest that IL-8 was internalized by the endothelial cells, transported transcellularly via plasmalemmal vesicles, and released onto the luminal surface where it appeared located preferentially on tips of membrane protrusions. We were unable to study the endothelial IL-8 binding or transport in vitro because all the in vitro propagated endothelial cell lines and primary endothelial cells tested lacked IL-8 binding sites. This includes human umbilical vein endothelial cells (HUVECs), which also did not bind IL-8 in situ. However, HUVECs provided a satisfactory in vitro system to study the secretion of IL-8 by the endothelial cells. Two possible alternative pathways were described: secretion directly from the Golgi apparatus or following storage in Weibel-Palade bodies.

IgG isotype-specific auto-antibodies bind preferentially to cross-linked membrane Ig.

Under equilibrium conditions, the affinities of five anti-IgG2a mAb isolated from virus-infected mice were comparable to other high-affinity auto-antibodies. Similar to rheumatoid factors, these anti-IgG2a auto-antibodies bound to aggregated or complexed IgG2a with 50 to 1500-fold higher avidity than their monomeric counterparts. Despite their high functional affinity to IgG2a, flow cytometric analysis revealed no binding or marginal mAb binding to four distinct lines of B cells expressing different densities of membrane-anchored IgG2a. If, however, surface IgG2a was cross-linked by polyclonal light chain-specific antibodies, IgM and IgA mAb binding resulted, and was detected as an increase in mean fluorescence intensity compared with isotype-matched control antibodies. The binding of one IgM mAb to cross-linked IgG2a patches of the cell surface was also visualized by confocal microscopy. Pretreatment of cells with aggregated IgG2a caused increased fluorescence intensity, demonstrating that the IgM and IgA mAb were also able to interact with IgG2a aggregates bound on the B cell surface via Fc gamma RIIB. It also permitted efficient co-ligation of the aggregated B cell receptors (BCR) with Fc gamma RIIB-fixed immune complexes known to deliver a negative signal in B cell activation. Cross-linking of IgG2a complexes bound to Fc gamma RI on macrophages or dendritic cells with antigen-specific BCR and/or T cells via their Fc gamma RIIB may accelerate the physical contact of cells involved in the antigen-specific response.(ABSTRACT TRUNCATED AT 250 WORDS)

Induction in mice of serum IgE levels after treatment with anti-mouse IgD antibodies is preceded by differential modulation of tissue cytokine gene transcription.

Injection of mice with purified goat anti-mouse IgD (GAMD) leads to an interleukin (IL)-4-dependent increase of serum IgE levels. Challenge of GAMD-primed mice with goat IgG (GIG) initiates a secondary immune response with elevated serum IgE. In this report, kinetic cytokine transcript profiles of murine lymphoid tissues in response to primary i.v. GAMD treatment, as well as GIG challenge are presented. For the first time, gene transcription patterns of the recently described cytokines IL-12 and IL-13 are shown and compared with the corresponding patterns for other cytokine genes involved in IgE regulation, i.e. IL-4, and interferon (IFN)-gamma. After GAMD injection, two groups of induction profiles were observed in spleen, mesenteric lymph nodes and Peyer's patches: while IL-4 and IL-12p35 gene transcription was strongly enhanced, IFN-gamma, IL-12p40 and IL-13 mRNA were only moderately induced. Generally, maximal mRNA induction was measured on days 3 to 4 after GAMD treatment. The data demonstrate a clear-cut difference between the IL-4 and IL-13 response on the transcriptional level although both gene products show similar biological activities. The cytokine mRNA profiles support the assumption of IL-4 playing the central role in generating an IgE response. However, they do not reflect a strict Th1 versus Th2 cytokine gene transcription pattern but rather point towards a concerted action of various, partially antagonizing cytokines with respect to the regulation of IgE synthesis. IL-12 may, possibly via stimulation of IFN-gamma synthesis, represent a counterbalancing factor in the IL-4-mediated IgE response.

Immunopharmacological activities and clinical development of muramyl peptides with particular emphasis on murabutide.

Certain immunopharmacological activities of muramyl peptides have been associated with inflammatory and undesirable side-effects typically observed following the administration of the prototype molecule muramyl dipeptide. This activity is now demonstrated not to be linked to a direct activation of inflammatory processes in endothelial cells. Neither MDP nor other structural derivatives were able to induce inflammatory cytokines release or E-selectin gene expression in cultured human umbilical vein endothelial cells. However, oral administration of muramyl peptides has been reported to induce certain biological effects, including the downregulation of anamnestic, antigen-specific IgE responses, which are not observed following parenteral administration. We elaborate on these findings and extend them to show the efficacy of a new muramyl peptide in suppressing polyclonally induced serum IgE levels in anti-IgD-treated mice. The comparative effects of muramyl peptides, selected for clinical development, on the induction of cytokines in human whole blood are then presented at the level of mRNA accumulation and protein secretion. Moreover, the cytokine profile induced in vitro and in vivo by the combination of the safe immunostimulant, Murabutide, with interferon-alpha is examined. This combination reveals a selective and beneficial synergistic activity and induces anti-inflammatory cytokines in the absence of synergistic toxicity. The potential and the implications for the use of a therapeutic combination of an immunostimulant with a cytokine are discussed.

IgG isotype distribution of local and systemic immune responses induced by influenza virus infection.

The IgG isotype profile of the influenza virus-specific immune response was studied by quantitation of serum antibody (Ab) levels in correlation with the enumeration of antibody-secreting cells (ASC) detected in the lung, spleen, mediastinal lymph nodes (MLN), Peyer's patches and bone marrow (BM). Distinct isotypic patterns for serum Ab and Ab produced by cells present at or close to the site of infection were found after primary or repeated infections. An elevated number of IgM ASC was found after primary challenge in the spleen, lung and MLN. In contrast, the site of IgA and IgG production is restricted to the lung and lymph nodes draining the site of infection. In these organs IgA, IgG2a and IgG1 ASC are found as a result of primary virus infection while viral challenge induces mostly activation of IgA-producing cells and secretion of IgA to the lung lavage. In contrast, the majority (80-90%) of Ab detected in the serum belong to the IgG2a subclass and their serum level is maintained at a high level during the whole period of the response. The relative level of virus-specific serum IgG2a in correlation with the production of IgG2a Ab found predominantly in MLN and lung is highly dependent on the viral dose used for priming or challenge. As IgG2a ASC can be detected at relatively low numbers in the spleen and BM these results suggest that the production of the dominant IgG2a isotype of serum Ab occurs close to the viral challenge site. These data, however, point to distinct isotypic regulation in systemic versus local virus-specific Ab responses.

Control of IgE responses. II. Isotype specific suppression of peak hapten specific IgE antibody forming cell responses in BPO-KLH sensitized mice after oral administration of muramyldipeptide or murabutide.

Muramyldipeptide (MDP) and murabutide (MB), a pyrogen free derivative of MDP, suppressed BPO specific IgE antibody forming cell (AFC) responses in vivo. To induce IgE responses, BALB/c mice were injected intraperitoneally (i.p.) with BPO-KLH (10 micrograms) in alum on days 0 and 21, or on days 0, 21 and 42. On day 44, mice were fed (gavage) or injected subcutaneously (s.c.) with MDP or MB (0.1-500 mg/kg). Mice were killed on days 45-70, and the numbers of BPO specific IgM, IgG1, IgE, and IgA antibody forming cells (AFC) in lymphoid organs determined in ELISPOT assay. With either immunization schedule, oral treatment with MDP or MB on day 44 suppressed BPO specific IgE AFC responses within 48 h (65-100%). With both molecules, the suppression was IgE isotype specific, dose dependent and transient. The suppression was also route specific since it was obtained only when MDP or MB was given by gavage, and not when injected s.c. These results show that peak antigen specific IgE responses can be suppressed in vivo, in isotype specific fashion, by a clearly defined class of molecules, one of which, MB, is a candidate for clinical studies in man. Pharmacologic agents of this type may be suitable for use in the therapeutic or prophylactic suppression of IgE and, hence, in the therapy of IgE mediated diseases such as allergic rhinitis, asthma, and other atopic diseases.

Control of IgE responses. III. IL-6 and IFN-alpha are isotype-specific regulators of peak BPO-specific IgE antibody-forming cell responses in mice.

The ability of cytokines (IL-4, IL-5, IL-6, IFN-alpha, IFN-gamma, TNF-alpha, GmCSF) to regulate peak benzylpenicilloyl (BPO)-specific IgE antibody-forming cell (AFC) responses was investigated. These responses were induced in BALB/c mice by ip injection of BPO-keyhole limpet hemocyanin (BPO-KLH; 10 micrograms) in aluminum hydroxide gel on Days 0, 21, and 42. On Day 44, or on Days 43, 44, and 45, mice were injected sc with varying doses of cytokine or anti-cytokine antibody. On Day 46, the numbers of BPO-specific AFC (IgM, IgG1, IgE and IgA) in spleen were determined ex vivo in enzyme-linked immunosorbent spot assay. Among the cytokines tested, only IL-6 suppressed BPO-specific IgE AFC responses in an isotype-specific fashion (60-90%). However, treatment of mice with anti-IL-6 also suppressed these responses, suggesting that IL-6 can either suppress or increase peak antigen specific IgE responses, depending upon its concentration. Among the cytokines tested, only IFN-alpha increased BPO-specific IgE AFC responses in an isotype-specific fashion. Since treatment with anti-IFN-alpha suppressed these responses, it appears that IFN-alpha is required to maintain peak antigen-specific IgE AFC responses. IL-4 or IFN-gamma nonspecifically suppressed responses of all isotypes. Treatment with anti-IL-4 also suppressed IgE responses, suggesting that this cytokine is required to maintain peak antigen specific IgE responses. Treatment with anti-IFN-gamma increased IgE responses, indicating that IFN-gamma suppresses peak antigen-specific IgE responses.

Control of IgE responses. 4. Isotype-specific suppression of peak BPO-specific IgE antibody-forming cell responses and of BPO-specific IgE in serum by muramyldipeptide or murabutide after administration to mice by gavage.

Muramyldipeptide (MDP) and murabutide (MB) suppressed hapten-specific IgE antibody-forming cell (AFC) responses in vivo. IgE responses were induced in BALB/c mice by intraperitoneal injection with benzylpenicilloyl-keyhole limpet hemocyanin (BPO-KLH) (10 micrograms) in aluminum hydroxide gel (Alum) on days 0, 21 and 42. On day 44, mice were fed (gavage) or injected subcutaneously with varying concentrations of MDP or MB (0.1-500 mg/kg). The mice were killed on days 45-70, and the numbers of BPO-specific IgM, IgG1, IgE, and IgA AFC in various lymphoid organs were determined in an enzyme-linked immunosorbent spot (ELISPOT) assay. In addition, levels of BPO-specific IgE in serum were determined by ELISA. Data are expressed as AFC/10(7) cells or as micrograms/ml. Feeding with MDP or MB on day 44 suppressed BPO-specific IgE AFC responses and serum levels of BPO-specific IgE within 48 h (day 46) (65-100% and approximately 50% decrease, respectively). With both molecules, the suppression was IgE isotype-specific, dose-dependent and transient. The suppression was also route-specific since it was obtained only when MDP or MB were given by gavage, and not when injected subcutaneously. These results show that peak antigen-specific IgE responses can be downregulated in vivo, in isotype-specific fashion, by a clearly defined class of molecules, MDP and MB, one of which, MB, is a candidate for clinical studies in man. The mechanism of suppression probably involves the modulation of gut-associated lymphoid tissue and mucosal immunity. The clinical implications are that pharmacologic agents of this type may be suitable for use in the therapeutic or prophylactic downregulation of IgE and, hence, in the therapy of IgE-mediated diseases in man such as allergic rhinitis, asthma, and other atopic diseases.

Selection of a muramyl peptide based on its lack of activation of nuclear factor-kappa B as a potential adjuvant for AIDS vaccines.

Activation of the cellular transcription factor nuclear factor-kappa B (NF-kappa B) by cytokines and other immunostimulants has been tightly linked with enhanced replication of human immunodeficiency virus-type 1 (HIV-1) in infected cells. Various immunomodulators are currently being examined in animal and human trials for their suitability as adjuvants in potential vaccines against acquired immunodeficiency syndrome (AIDS). It may prove to be beneficial to select adjuvants that do not induce NF-kappa B activation and particularly if the vaccines are to be aimed at seropositive individuals. We have examined a battery of synthetic immunostimulants of the muramyl peptide family for their ability to activate NF-kappa B in human and mouse cell lines. In this report, we demonstrate selective activation of NF-kappa B in different cell lines and by different muramyl peptides possessing immunostimulatory activities. The mechanism of such activation is apparently via production of reactive oxygen intermediates (ROI) since pretreatment of cells with antioxidants blocked subsequent activation of NF-kappa B. However, among all the molecules tested only one lipophilic, non-pyrogenic adjuvant active muramyl peptide showed a complete lack of NF-kappa B activation in all cell lines tested. This molecule could well become the adjuvant of choice in future AIDS vaccines.

Origin and fate of IgE-bearing lymphocytes. II. Modulation of IgE isotype expression on Peyer's patch cells by feeding with certain bacteria and bacterial cell wall components or by thymectomy.

Mechanisms regulating the appearance of sIgE+ B lymphocytes appear to be lacking in adult germfree (GF) rats in that their Peyer's patches (PP) contain high numbers of cells with sIgE (approximately 15% of total cells), one-half of which simultaneously express sIgA, whereas sIgE+ cells are absent from PP of conventional rats (less than 1%). GF rat PP also contain elevated numbers of sIgA+ cells and decreased numbers of sIgM+ cells, with elevated numbers of sThy-1+ RT 7.1+ Ig- T cells, and reduced numbers of sThy-1- RT 7.1+ Ig- T cells. The cellular composition of PP of GF rats was converted to that resembling a conventional rat within 18 h after either 1) use of standard (unautoclaved) food; 2) feeding with certain bacteria (Clostridium difficile, Corynebacterium pseudodiphtheriticum, Mycobacterium tuberculosis, and Klebsiella pneumoniae), in either live or heat-killed, but not autoclaved form; or with certain bacterial cell wall components: murein (peptidoglycan), and its synthetic derivatives, muramyltripeptide phosphatidylethanolamine and desmethyl-muramyldipeptide, but not with LPS, core lipid A or lipoprotein; there was no effect if any bacterial cell wall component was injected i.v.; or 3) thymectomy. Each procedure resulted in elimination of sIgE+ B cells and normalization of the other surface isotypes, and loss of sThy-1+ RT 7.1+ Ig- T cells and normalization of sThy-1- RT 7.1+ Ig- T cells. Irrespective of treatment, no sIgE+ cells were detected in bone marrow, thymus, other lymphoid organs or blood, excluding the possibility that the elimination of these cells from PP was associated with their redistribution to other sites. Thus, exposure to gut flora and bacterial peptidoglycan components may have resulted in IgE isotype switching, either directly or through the mediation of accessory and/or sThy-1+ RT 7.1+ regulatory T cells. The sites in which sIgE+ B cells are down-regulated appear to be PP.

Control of IgE responses.

Peyer's patches (PP) in germ-free rats (GF) and in the hyper-IgE syndrome patient (HIES) differ from their conventional rat (C) and healthy human (HH) counterparts in that GF rats contained fewer (two-fold) PP and none was detected in HIES. Existing PP in GF rats had reduced cellularity (three-fold) and different B and T cell subsets: high numbers of IgE-bearing (sIgE+) B cells (approximately 15% of total cells), one-half of which also expressed sIgA, were present in GF rat PP while none was detected in C rat PP (less than 1%). GF rat PP also contained elevated numbers of sIgA+ cells and decreased sIgM+ cells, with elevated numbers of sThy 1+ RT 7.1+ Ig- T cells (suppressor phenotype) and reduced sThy 1- RT 7.1+ Ig- T cells (helper phenotype). The cellular composition of GF rat PP was converted to that resembling a C rat within 18 hr after (a) use of standard (unautoclaved) chow; (b) feeding with certain bacteria or "working" bacterial cell wall components (BCWC) and synthetic derivatives, murein, MTP-PE, and norMDP, but not with LPS, core lipid A, or lipoprotein; BCWC had no effect if injected intravenously; or (c) thymectomy. Each procedure resulted in (i) elimination of sIgE+ B cells and normalization of the other isotypes, and (ii) loss of T suppressor cells and normalization of T helper cells. After treatments, no sIgE+ cells were detected in bone marrow (BM), thymus, other lymphoid organs, or blood. PP were not detected in HIES, although they were present in HH (approximately 10/individual). P blood contained two distinct sIgE+ B cell subpopulations, the apparent source of which was mesenteric lymph node (MLN), the only organ in which high numbers of these cells (35%) (five nodes examined) were detected; far fewer IgE+ cells were found in spleen (less than 5%), and none was detected in BM, thymus, other LN, or appendix, which was virtually acellular. Virtually no IgE secreting plasma cells were detected in MLN, spleen, appendix, other lymphoid organs, or in gut lamina propria. IgE+ B cells in MLN were not detected in follicles (classical B cell areas); instead, they were found in high numbers in the thymus-dependent area and in medulla. Most follicles (greater than 98%) in MLN and spleen contained intercellular IgE complexed to bacterial antigen and/or CD23 (IgE-binding factor? antigen?), but contained no germinal centers.(ABSTRACT TRUNCATED AT 400 WORDS)

Modulation of interferon-gamma-induced major histocompatibility (MHC) and CD14 antigen changes by lipophilic muramyltripeptide MTP-PE in human monocytes.

The lipophilic muramylpeptide derivative muramyltripeptide-phosphatidylethanolamine (MTP-PE, 0.05 to 5 micrograms/ml) and human recombinant interferon-gamma (rIFN-gamma, 1 to 100 U/ml) were applied singly or in combination to fresh human mononuclear blood leucocytes in vitro. After 15 to 72 hr incubation, culture- and drug-induced changes in beta 2-microglobulin (MHC class I associated), HLA-DR (MHC class II), and Leu-M3 (CD14) antigen expression were investigated by flow cytometry; changes in monocyte morphology (forward light scatter and side scatter) were assessed by scatter analysis. It was found that (1) rIFN-gamma caused a simultaneous down-regulation of the CD14 antigen and an up-regulation of MHC class I and class II molecules on the surface of cultured monocytes; (2) MTP-PE, which by itself failed to influence the expression of these antigens, synergized with rIFN-gamma in increasing MHC antigens and reducing CD14; (3) at high concentrations rIFN-gamma reduced monocyte viability to a small but significant extent and this effect was further potentiated by MTP-PE; and (4) untreated monocytes in culture showed an apparently MTP-PE-insensitive increase in size, density, and beta 2-microglobulin, HLA-DR, and CD14 antigen expression. The influence of MTP-PE on rIFN-gamma-induced surface marker changes may contribute to its immunoadjuvant activity in vivo.

Induction of tumouricidal leucocytes by the intranasal application of MTP-PE, a lipophilic muramyl peptide.

Single intranasal applications of MTP-PE, a lipophilic muramyl peptide, induce tumouricidal and tumouristatic leucocytes in the lungs of rats. In ex vivo assays the tumouristatic activity was detectable for 8 days after drug administration. By separation of the effector cells on Ficoll-Hypaque gradients, it was shown that both neutrophils and macrophages are responsible for this activity. Using the B16/BL6 melanoma system in mice, there was a high survival rate after repeated intranasal applications of MTP-PE.

Nor-MDP, saponin, corynebacteria, and pertussis organisms as immunological adjuvants in experimental malaria vaccination of macaques.

Vaccination of primates against malaria using antigen derived from erythrocytic parasite stages has been most successful where Freund's complete adjuvant has been employed. Since this adjuvant is clinically unacceptable its replacement is a matter of urgency.In the present work a muramyldipeptide derivative (nor-MDP) given in mineral oil has proved to be partially effective as an adjuvant for merozoite vaccination of Macaca mulatta against Plasmodium knowlesi, and saponin has proved to be effective in similar vaccination of M. fascicularis.