Prophylaxis of implant-related infections by local release of vancomycin from a hydrogel in rabbits.

Local prophylaxis with antibiotic-loaded bone cement is a successful method to prevent post-operative infections in patients receiving orthopaedic implants. No comparable method is available for uncemented implants. Therefore, a hydrogel consisting of hyaluronic and polylactic acids was evaluated in a rabbit model for delivery of antimicrobial agents to prevent post-operative infections. In a pilot study, the suitability of the in vivo model was assessed by testing the hydrogel as carrier material for antimicrobial agents.In the main study, the antimicrobial-agent-loaded hydrogel was evaluated for infection prophylaxis. Rabbits received a titanium rod intramedullary in the tibia after contamination with Staphylococcus aureus. The rods were coated with unloaded hydrogel (Gel), hydrogel loaded with 2 % (Van2) or 5 % vancomycin (Van5), bioactive glass (BAG) or N-acetyl-L-cysteine (NAC). To analyse the infection severity after 28 d, histopathological, bacteriological, micro-computed tomographic and haematological analyses were performed. In the pilot study, the Van5 group had less infection (0/6 infected) as compared to the Gel group (5/5, p = 0.000) and the in vivo model was deemed suitable. In the main study, in the Van2 and Van5 groups, the number of infected animals was lower [1/6 (p = 0.006) and 2/6 (p = 0.044) infected, respectively]. In contrast, BAG and NAC groups showed no infection reduction (5/6 both groups, p = 0.997). The hydrogel can be used as a local carrier of vancomycin for prophylaxis of implant-related infections.The present study showed promising results for local delivery of antibacterial agents by hydrogel to prevent implant-related infections.

[Pathophysiology of implant-associated infections: From biofilm to osteolysis and septic loosening]. / Pathophysiologie der implantatassoziierten Infektion : Vom Biofilm zur Osteolyse und septischen Lockerung.

Biofilm formation is the key factor in the pathogenesis of implant-associated infections. The most common pathogens isolated are Staphylococcus species, opportunists belonging to the physiological flora of the skin. Biofilm formation starts with the adhesion of bacteria and colonisation preferentially occurs on the surfaces of the foreign body material. As an interactive symbiotic "city of microbes," biofilm formation represents an efficient survival strategy for bacteria. In clinically apparent infections the biofilm induces a local host response with infiltration of phagocytic immune cells. The proinflammatory microenvironment results in a stimulation of osteoclastogenesis, with local osteolysis, and finally septic loosening of the implant. According to the biofilm theory, retaining the implant in primary revision surgery is only recommended in early-stage infections with a stable implant; in late-stage infections, or when loosening occurs, the implant should be removed. Results of previous anti-biofilm therapies have not been satisfactory; therefore, current research is focused on prevention strategies, especially the modification of implant surfaces. Basic knowledge of the underlying pathophysiology is a prerequisite for the development of innovative interdisciplinary therapy and prevention strategies; in this context, essential aspects of biofilm formation, its consequences, and its relevance to diagnosis and therapy are described and discussed.

Bacterial quorum sensing molecule induces chemotaxis of human neutrophils via induction of p38 and leukocyte specific protein 1 (LSP1).

When bacteria colonize surfaces, they socialize and form biofilms. This process is well regulated and relies on the communication among the bacteria via so-called "quorum sensing molecules". Among those, N-(3-oxododecanoyl)-L-homoserine lactone (AHL-12), generated by Pseudomonas aeruginosa and other Gram-negative bacteria, activates not only bacteria but also interacts with mammalian cells. Among others, it activates phagocytic cells and - as we had shown previously - it is chemotactic for human polymorphonuclear neutrophils (PMN) in vitro. In the present study, we analyzed the signalling pathway of AHL-12 in PMN. We focused on the mitogen activated protein (MAP) kinase p38, because SB203580, an inhibitor of p38, prevented the AHL-12 induced chemotaxis. We found that in response to AHL-12, p38 was phosphorylated within minutes, as was its downstream target, the MAPKAP-Kinase-2 (MK2). In PMN, the major substrate of MK2 is the leukocyte specific protein 1 (LSP1), which binds to F-actin and participates directly in actin polymerization and cell migration. In response to AHL-12, LSP1 was phosphorylated and co-localized with F-actin in polarized PMN, suggesting that AHL-12-induced migration depended on p38 and LSP1 activation.

Expression of CD57 on CD8+ T lymphocytes of patients with Wegener's granulomatosis and microscopic polyangiitis: evidence for continuous activation of CD8+ cells.

OBJECTIVES: To gain insight into the immune pathogenesis of Wegener's granulomatosis (WG) and microscopic polyangiitis (MPA), the prevalence of circulating CD8+ T lymphocytes expressing CD57 as a marker for previous activation was analyzed. METHODS: Receptor expression of CD57 was measured in CD8+ T cells of patients with active disease (n=5) by cytofluorometry and compared with expression in patients in remission (n=80) and in age-matched healthy donors (n=34). The results were compared to clinical parameters including severity and duration of the disease. RESULTS: CD8+CD57+ were detected in patients with WG and MPA and in healthy donors as well and increased considerably with age. Compared to age-matched healthy donors, the prevalence of CD8+CD57+ was increased in the younger patients (up to 40 y). In most patients a high percentage of CD8+CD57+ coincided with severe disease and multiple organ involvement, while low CD8+CD57+ percentage was seen in patients with limited disease or in patients in complete remission. In patients with smoldering disease, the percentage of CD8+CD57+ increased with time. High numbers of CD8+CD57+ correlated with low CD4:CD8 ratio. CONCLUSIONS: In patients with WG and MPA a population of CD8+CD57+ expand, identifying terminally differentiated CD8+ cells. The prevalence of CD57+ cells was related to the course of disease. So far, the function of CD57 on CD8+ cells is not understood. However, these cells might produce certain cytokines, which play a role in the pathogenesis of AAV. The data support the hypothesis that CD8+ T cells are activated in the context of primary vasculitides.

The extracellular polymer substance of Pseudomonas aeruginosa: too slippery for neutrophils to migrate on?

PURPOSE: Biofilm formation is increasingly recognized as the cause of persistent infections and there is evidence that P. aeruginosa organized into biofilms are quite resistant toward host defence mechanisms, particularly against an attack by polymorphonuclear neutrophils (PMN). Apparently, the migration of PMN through the biofilms is impaired, and thus the bactericidal activity remains highly localized. The aim of this study was to directly investigate the interaction of PMN with the biofilm and the extracted extracellular polymeric substance (EPS) of P. aeruginosa. MATERIAL AND METHODS: Chemotaxis and random migration of PMN through P. aeruginosa biofilms was tested, as was their migration through and along the EPS. RESULTS: We found that the EPS and mature biofilms, but not immature or developing ones, reduced the chemotactic migration of PMN. On EPS, rather than immobilize the cells, their random, spontaneous migration was enhanced. CONCLUSION: We propose that on EPS, the PMN lose their capacity to sense the direction and just slide over the EPS in a disoriented manner.

Combating implant infections. Remarks by a women's team.

Research on implant infections requires cooperative efforts and integration between basic and clinical expertises. An international group of women scientists is acting together in this field. The main research topics of the participants of this group are described. Formation of bacterial biofilms, antibiotic resistance and production of virulence factors like adhesins and toxins are investigated. New biomaterials, coatings and drugs designed to inhibit microbial adhesion are evaluated, and infection-resistant biomaterials are under study, such as a novel heparinizable polycarbonate-urethane (Bionate) or incorporation of diamino-diamide-diol (PIME) to reduce bacterial attachment. The correlation between biofilm production and the accessory-gene-regulator (agr) is investigated in Staphylococcus aureus. The ability to form biofilm has also been shown to be one of the important virulence factors of Enterococcus faecalis, favouring colonization of inert and biological surfaces. The study of quorum sensing has led to the discovery of a quorum sensing inhibitor termed RIP that suppresses staphylococcal biofilm and infections. The immune response and the local defence mechanisms of the host against implant-associated infections, activation and infiltration of immunocompetent cells into the sites of infection have been studied in patients with implant-associated osteomyelitis. Production of monoclonal antibodies (mAbs) as possible vaccines against the staphylococcal collagen-binding MSCRAMMs is in progress.

Expression of the CXCR6 on polymorphonuclear neutrophils in pancreatic carcinoma and in acute, localized bacterial infections.

The chemokine receptor CXCR6 has been described on lymphoid cells and is thought to participate in the homing of activated T-cells to non-lymphoid tissue. We now provide evidence that the chemokine receptor CXCR6 is also expressed by activated polymorphonuclear neutrophils (PMN) in vivo: Examination of biopsies derived from patients with pancreatic carcinoma by confocal laser scan microscopy revealed a massive infiltration of PMN that expressed CXCR6, while PMN of the peripheral blood of these patients did not. To answer the question whether CXCR6 expression is a property of infiltrated and activated PMN, leucocytes were collected from patients with localized soft tissue infections in the course of the wound debridement. By cytofluorometry, the majority of these cells were identified as PMN. Up to 50% of these PMN were also positive for CXCR6. Again, PMN from the peripheral blood of these patients were nearly negative for CXCR6, as were PMN of healthy donors. In a series of in vitro experiments, up-regulation of CXCR6 on PMN of healthy donors by a variety of cytokines was tested. So far, a minor, although reproducible, effect of tumour necrosis factor (TNFalpha) was seen: brief exposure with low-dose TNFalpha induced expression of CXCR6 on the surface of PMN. Furthermore, we could show an increased migration of PMN induced by the axis CXCL16 and CXCR6. In summary, our data provide evidence that CXCR6 is not constitutively expressed on PMN, but is up-regulated under inflammatory conditions and mediates migration of CXCR6-positive PMN.

Abnormal crosstalk between pancreatic acini and macrophages during the clearance of apoptotic cells in chronic pancreatitis.

In chronic pancreatitis (CP), both the progressive loss of acinar parenchyma and aggressive fibro-inflammatory reactions ultimately lead to irreversible organ destruction. Dying cells are normally removed by macrophages and elimination is associated with anti-inflammatory cytokine switch. We investigated whether defective clearance of damaged acini by macrophages such as compromised phagocytosis or altered cytokine reaction occurs in CP and thus represents a causative link between acinar loss and fibro-inflammation. In a checkerboard-like co-culture system, we assessed normal and CP macrophages for their phagocytic and cytokine responses to dying pancreatic acinar cells of normal or CP origin by FACS, confocal microscopy, QRT-PCR, and ELISA. In CP, phagocytosis of apoptotic acini by macrophages was not impaired; however, the associated cytokine responses were gradually perturbed. Most interestingly, only normal acini suppressed TGFbeta1 expression and accumulation specifically in normal macrophage cultures, while CP acini lost this ability. Both types of apoptotic acini induced pro-inflammatory cytokine bursts of varying strength in both types of macrophages; however, the most significant difference (more than 50-fold higher expression of IL-1beta, IL-6, and IL-8) was evident between CP/CP and normal/normal combinations, indicating that acinar and macrophage alterations synergistically lead to the ultimate CP-specific bias. In combination with in situ data comparing circulating inflammatory cells to pancreatic resident ones, our results indicate that cytokine expression in inflammatory cells undergoes spatiotemporal modulation, most likely through a successive interplay of acinar, stromal, and circulating factors. Thus, clearance of injured pancreatic acini by macrophages is associated with a unique cytokine reaction which may constitute a basis for progression of SAPE (sentinel acute pancreatitis event) to the irreversible fibro-inflammation in CP.

T lymphocytes in patients with primary vasculitis: expansion of CD8+ T cells with the propensity to activate polymorphonuclear neutrophils.

OBJECTIVES: To gain insight into the immune pathogenesis of primary ANCA-associated vasculitides, the prevalence of circulating T lymphocytes expressing CD11b as a marker for activation was analysed in patients with WG or microscopic polyangiitis. METHODS; Receptor expression and IFNgamma synthesis were measured in T cells of patients with active disease by cytofluorometry and compared with expression in patients in remission and in healthy donors. RESULTS: During active disease, a small but conspicuous population of CD8+CD28+CD11b+ was found which produced IFNgamma. In healthy donors and in patients in remission or undergoing immunosuppressive therapy, CD11b was exclusively associated with CD8+CD28- cells, the latter being more frequent in patients with long-lasting or severe disease. In vitro experiments confirmed that CD11b is up-regulated when T cells are activated. After multiple rounds of restimulation, the CD11b expression persists whereas CD28 expression is lost, compatible with the notion that CD8+CD28+CD11b+ represents a transient phenotype in the course of T-cell activation. The IFNgamma-producing T cells activated polymorphonuclear neutrophils (PMN) to express MHC class II, thus generating the same PMN phenotype as in patients with active ANCA-associated vasculitis. A similar PMN phenotype could be generated by cultivation with supernatants of activated T cells or by IFNgamma alone, but not by antibodies to proteinase 3. CONCLUSIONS: In active primary vasculitis, a small population of CD8+ T cells, identified by the expression of CD11b, expands, producing IFNgamma. These T cells could activate PMN, thus generating a long-living and potentially destructive PMN phenotype.

[Implant-associated post-traumatic osteomyelitis. Bacterial biofilms and the immune defence as protagonists of the local inflammatory process]. / Die implantatassoziierte posttraumatische Osteitis. Bakterielle Biofilme und Infektabwehr als Protagonisten der lokalen Entzündungsreaktion.

BACKGROUND: Formation of bacterial biofilms on implants is a severe complication following orthopaedic surgery. In the present study we addressed the role of the immune response, particularly with regard to the pathogenesis of the disease. METHODS: In a prospective study comprising 74 patients with implant-associated post-traumatic osteomyelitis, peripheral blood cells as well as cells recovered from the infected site during surgery were characterised phenotypically and functionally. RESULTS: We found massive infiltration of polymorphonuclear neutrophils (PMN), which were highly activated, particularly regarding their bactericidal potential, such as increased production of superoxides and upregulation of activation-associated surface receptors. CONCLUSION: PMN are activated in response to the implant-associated osteomyelitis; they also infiltrate the infected tissue, but cannot control the infection. By release of their cytotoxic entities they could contribute to tissue destruction and eventually to osteolysis.

Cellular inflammatory response to persistent localized Staphylococcus aureus infection: phenotypical and functional characterization of polymorphonuclear neutrophils (PMN).

Persistent, localized Staphylococcus aureus infections, refractory to antibiotic treatment, can result in massive tissue destruction and surgical intervention is often the only therapeutic option. In that context, we investigated patients with S. aureus-induced infection at various sites, apparent as either olecranon bursitis, empyema of the knee joint or soft tissue abscess formation. As expected, a prominent leucocyte infiltrate was found, consisting predominantly of polymorphonuclear neutrophils (PMN) (up to 75%) and to a lesser extent of T lymphocytes and natural killer (NK) cells. In line with their bactericidal capacity, PMN expressed the high-affinity receptor for IgG, CD64 and the lipopolysaccharide (LPS) receptor CD14; moreover, the oxygen radical production in response to the bacterial peptide f-MLP was enhanced, while chemotactic activity was greatly reduced. The more intriguing finding, however, was that a portion of PMN had acquired major histocompatibility complex (MHC) class II antigens and CD83, indicative of a transdifferentiation of PMN to cells with dendritic-like characteristics. Of note is that a similar transdifferentiation can be induced in PMN in vitro, e.g. by gamma interferon or by tumour necrosis factor alpha. Co-cultivation of transdifferentiated PMN with autologous T lymphocytes resulted in prominent T cell proliferation, provided that S. aureus enterotoxin A was added. Taken together, persistent S. aureus infection induces PMN to acquire characteristics of dendritic cells, which in turn might promote the local immune response.

Implant-associated posttraumatic osteomyelitis: collateral damage by local host defense?

Infections following osteosynthesis or total joint replacement, also known as ''implant-associated posttraumatic osteomyelitis'', represent a major complication in orthopedic and trauma surgery. While the formation of bacterial biofilms on the implanted osteosynthesis materials is generally accepted as cause of the persistent infection, the molecular mechanisms leading to the progressive and destructive local inflammatory process and eventually to bone degradation, the osteolysis, have not been delineated. Here we provide evidence supporting the hypothesis that it is not the infection per se that causes tissue degradation and osteolysis, but rather the cytotoxic, proteolytic, and proinflammatory effector functions of cells of the host defense, particularly of the infiltrating polymorphonuclear neutrophils.

The pathogenesis of ANCA-associated vasculitides: old hypotheses and new insights.

The pathomechanism of the ANCA-associated vasculitides is discussed in light of the abstracts presented at the ANCA- and Vasculitis Workshop 2005 in Heidelberg!

Transdifferentiation of polymorphonuclear neutrophils to dendritic-like cells at the site of inflammation in rheumatoid arthritis: evidence for activation by T cells.

OBJECTIVES: To investigate infiltrated cells in the synovial fluid (SF) of inflamed joints of patients with rheumatoid arthritis (RA), with special reference to polymorphonuclear neutrophils (PMN) and their interaction with T cells. METHODS: Expression on PMN of activation associated receptors CD14, CD64, CD83, and major histocompatibility complex (MHC) class II was examined in the SF of 15 patients with RA, as were the infiltrated T cells. SF cytokines were determined by enzyme linked immunosorbent assay (ELISA). To mimic the in vivo situation, co-culture experiments were carried out using PMN and T cells of healthy donors. RESULTS: The SF contained activated T lymphocytes and abundant PMN. SF PMN expression of CD14 and CD64 was enhanced compared with peripheral blood. Of special interest was the observation that only the SF PMN expressed MHC class II antigens and CD83. Exposure to SF, which contained considerable amounts of cytokines (for example, interferon gamma (IFNgamma), tumour necrosis factor alpha, and interleukin 2), induced a similar receptor pattern on blood derived PMN of healthy donors. Furthermore, PMN acquired MHC class II and CD83 within 24 to 48 hours, when co-cultured with autologous T cells or T cell lines. This effect was also achieved by T cell supernatants, was dependent on protein synthesis, and could be inhibited by antibodies against IFNgamma. CONCLUSIONS: SF PMN from patients with RA undergo major alterations, including transdifferentiation to cells with dendritic-like characteristics, probably induced by T cell derived cytokines. Because MHC class II positive PMN are known to activate T cells, the mutual activation of PMN and T cells might contribute to the perpetuation of the local inflammatory process, and eventually to the destructive process in RA.

Genetic deficiency of CD16, the low-affinity receptor for immunoglobulin G, has no impact on the functional capacity of polymorphonuclear neutrophils.

BACKGROUND: Of the three receptors for immunoglobulin G (IgG), the low-affinity receptor CD16 is constitutively expressed on polymorphonuclear neutrophils (PMNs), monocytes and NK-cells. CD16 participates in various effector functions, notably phagocytosis of opsonized particles or of immune complexes, and in antibody-dependent cellular cytotoxicity (ADCC). In the present study we report a case of total CD16 deficiency on PMNs and monocytes. DESIGN: Polymorphonuclear neutrophils, monocytes and NK-cells were analyzed for surface-receptor expression by cytofluorometry and laser scan microscopy. Moreover, CD16-specific mRNA was assessed by RT-PCR. As functional parameters, phagocytosis of opsonized bacteria was tested, as was superoxide production. RESULTS: Polymorphonuclear neutrophils and monocytes totally deficient in CD16 were detected by chance in an apparently healthy individual. Further analysis revealed that two more members of his family, his father and sister, were also deficient in CD16. All were healthy and there was no evidence of an increased frequency, or of exceptionally severe or persistent infections. Despite the lack of CD16, phagocytosis of antibody-coated bacteria was within the normal range, as was the superoxide production. CONCLUSION: Deficiency of CD16 does not compromise the host defence. Apparently, the other receptors for IgG, CD32 and CD64, can compensate for the lack of CD16.

Up-regulation of the dendritic cell marker CD83 on polymorphonuclear neutrophils (PMN): divergent expression in acute bacterial infections and chronic inflammatory disease.

Upon cultivation with interferon-gamma (IFN-gamma ) and granulocyte/macrophage-colony stimulating factor (GM-CSF) polymorphonuclear neutrophils (PMN) acquire characteristics of dendritic cells, including expression of major histocompatibility complex (MHC) class II antigens, of the co-stimulatory antigens CD80, CD86 and of CD83, the latter considered to be specific for dendritic cells. Dendritic-like PMN were also able to present to T cells antigens in a MHC class II-restricted manner. To assess whether dendritic-like PMN are also generated in vivo, cells of patients with acute bacterial infections and of patients with chronic inflammatory diseases (primary vasculitis) were tested. During acute infection up to 80% of PMN acquired CD83, but remained negative for MHC class II, CD80 or CD86. PMN of patients with primary vasculitis expressed MHC class II antigens, CD80 and CD86, but not CD83, indicating that up-regulation of MHC class II and of CD83 are not necessarily linked to each other. Indeed, parallel studies with PMN of healthy donors showed that while IFN-gamma and granulocyte/macrophage colony stimulating factor (GM-CSF) induced both, MHC class II and CD83, tumour necrosis factor (TNF)-alpha selectively induced de novo synthesis of CD83. The function of CD83 on PMN is still elusive. A participation in the MHC class II-restricted antigen presentation could be ruled out, consistent with the segregation of MHC class II and CD83 expression. Regardless, however, of its function, CD83 expression could serve as a marker to differentiate between acute and chronic inflammation.

Polymorphonuclear neutrophils in Wegener's granulomatosis acquire characteristics of antigen presenting cells.

BACKGROUND: Constitutive expression of major histocompatibility complex (MHC) class II antigens and of the co-stimulatory receptors CD80 and CD86 is restricted to professional antigen presenting cells. Polymorphonuclear neutrophils (PMN) of healthy donors are negative for those antigens. Our recent study, however, found that PMN of patients with active Wegener's granulomatosis acquired MHC class II antigens. METHODS: To continue and extend the previous study results, PMN and monocytes of 60 patients with Wegener's granulomatosis, 24 patients with microscopic polyangiitis (MPA), 20 patients with acute bacterial infection, and 53 healthy donors were analyzed for the expression of MHC class II antigens as well as of CD80 and CD86. Moreover, induction on PMN of MHC class II expression was studied, as was antigen presentation as a possible functional consequence. RESULTS: PMN of patients with acute, active Wegener's granulomatosis expressed MHC class II antigens, CD80 and CD86; on monocytes up-regulation of MHC class II was seen. In contrast, PMN of patients with inactive disease, or with relapse, patients with microscopic polyangiitis or with bacterial infections expressed neither MHC class II, nor CD80 or CD86. PMN of healthy donors acquired these antigens when cultured in the presence of T cells or T cell-derived cytokines. The PMN were then able to present to T cell antigens in a MHC-class II restricted manner. CONCLUSION: During active disease, the PMN of patients with Wegener's granulomatosis acquire characteristics of antigen presenting cells, whereas the PMN of patients with MPA or bacterial infection do not. The finding reflects differences in the pattern of the respective inflammatory response and suggests new effector functions of PMN. Moreover, MHC class II expression on PMN could serve as a novel marker for active Wegener's granulomatosis.

Transdifferentiation of polymorphonuclear neutrophils: acquisition of CD83 and other functional characteristics of dendritic cells.

Polymorphonuclear neutrophils (PMN) are in the first line of defense against bacterial infections. They are considered to be end-differentiated cells undergoing constitutive apoptosis within hours after release from the bone marrow. During pathological events, however, their life span is extended in conjunction with morphological and functional alterations indicative of a transdifferentiation of mature PMN. To further characterize differentiated PMN, the alterations seen in vivo were reproduced by cultivating PMN of healthy donors with either gamma-interferon, granulocyte/macrophage colony stimulating factor, or a combination thereof. Thus cultivated cells escaped from apoptosis, and protein synthesis was induced, notably of the major histocompatibility complex (MHC) class II antigens, CD80 and CD86. Moreover, CD83, thought to be specific for dendritic cells was synthesized, while typical markers of PMN, including CD66b, CD11a/CD11b/CD11c, CD15, CD18 were preserved. A profound alteration of both cellular morphology and of function was seen: the cultivated PMN lost their chemotactic activity but had acquired the ability to present to T-cells a peptide antigen in a MHC class II restricted manner. The data lead to the conclusion that mature PMN can differentiate further to cells with characteristics of DCs, thereby connecting PMN to the specific T-cell response.

Dietary wheat germ agglutinin modulates ovalbumin-induced immune responses in Brown Norway rats.

The trend towards an increased consumption of minimally processed plant food results in a higher intake of non-nutritive compounds such as lectins. Lectins are typically globular proteins that are resistant to digestion in the gastrointestinal tract. They affect the integrity of the intestinal epithelium and the absorption of dietary antigens, and induce the release of allergic mediators from mast cells in vitro. Based on this information we have studied whether dietary wheat germ agglutinin (WGA) could be involved in triggering food allergies. Brown Norway rats were immunized intraperitoneally using ovalbumin (OVA; 10 microg/rat) and 10 d later treated for five consecutive days with WGA (10 mg/rat per d) administered intragastrically. Rats were then orally challenged with OVA (100 microg/rat) 1 h after the last WGA application, and blood was collected 4 h later. Immunological responses (anti-OVA immunoglobulins E and G, rat mast cell protease II, interferon-gamma and lymphocyte proliferation) were measured and lymphocyte subpopulations were determined. In immunized rats WGA treatment resulted in increased serum rat mast cell protease II concentrations (pre-challenge 0.26 (SE 0.08) microg/ml, post-challenge 0.49 (SE 0.09) microg/ml; P < 0.01) 4 h after the OVA challenge. After 5 d serum concentrations of anti-OVA immunoglobulin E were significantly increased only in the immunized controls (absorbance at 405 nm on days 14 and 19 was 0.09 (SE 0.008) and 0.24 (SE 0.046) respectively; P = 0.02), while in WGA-treated rats no significant increase was seen (0.08 (SE 0.004) and 0.15 (SE 0.037 respectively; P = 0.14). CD4+ : CD8+ T lymphocytes in the spleen was significantly increased at this time (OVA 1.1 (SD 0.2), 1.4 (sd 0.1), P < 0.05). The treatment did not impair the proliferation and interferon-gamma production of mesenteric lymphocytes. In conclusion, these data suggest that high dietary intake of lectins such as WGA may affect the allergic response towards oral antigens in the gut-associated lymphoid tissue.

The complement receptor 3, CR3 (CD11b/CD18), on T lymphocytes: activation-dependent up-regulation and regulatory function.

The complement receptor 3 (CR3; CD11b/CD18) is present exclusively on leukocytes, particularly on NK cells, monocytes and polymorphonuclear neutrophils. Approximately 10% of peripheral T lymphocytes and, as we found now mainly CD8(+) cells, expressed CD11b. Upon stimulation, however, expression of CD11b was up-regulated also on CD4(+) cells. Stimulation of T cells either by cross-linked anti-CD3 and IL-2 or by mononuclear cells and mitogen yielded up to 28% CD11b(+) T cells. The majority of CD11b(+) T cells also expressed CD56. T cell lines established from healthy donors were also found to express CR3. When restimulated up to 90% of cells became positive for CD11b making those cells an ideal tool for studying the functional role of CD11b. Antibodies to CD11b and bona fide ligands for the complement receptor inhibited the anti-CD3-induced T cell proliferation and as well as IL-2 release. In contrast, proliferation of a CD11b(-) T cell line was not inhibited. Taken together, our data indicate an activation-dependent expression of the complement receptor on T cells and suggest a regulatory function.