Effect of Cyclophosphamide Treatment on Central and Effector Memory T Cells in Mice.

Immunological memory is a key feature of adaptive immunity. It provides the organism with long-lived and robust protection against infection. The important question is whether cyclophosphamide (CP), as immunosuppressive agent used in cancer therapy and in some autoimmune diseases, may act on the memory T-cell population. We investigated the effect of CP on the percentage of central memory T cells (TCM) and effector memory T cells (TEM) in the mouse model of CP-induced immunosuppression (8-10-week-old male C57BL/6 mice CP treated for 7 days at the daily dose of 50 µg/g body weight [bw], manifested the best immunosuppression status, as compared to lower doses of CP: 10 or 20 µg/g bw). The CP induced a significant decrease in the percentage of CD8+ (TCM), compared to nonimmunosuppressed mice. This effect was not observed in the case of CD4+ TCM population. The percentage of gated TEM with CD4 and CD8 phenotype was significantly decreased in CP-treated mice, as compared to the control ones. Taken together, the above data indicate that CP-induced immunosuppression in mice leads to a reduction in the abundance of central memory cells possessing preferentially CD8+ phenotype as well as to a reduction in the percentage of effector memory cells (splenocytes both CD4+ and CD8+), compared to the cells from nonimmunosuppressed mice. These findings in mice described in this article may contribute to the understanding of the complexity of the immunological responses in humans and extend research on the impact of the CP model of immunosuppression in mice and memory T-cell populations.

Mycobacteria-derived biomarkers for tuberculosis diagnosis.

Tuberculosis (TB) remains an escalating problem worldwide. The current diagnostic methods do not always guarantee reliable diagnosis. TB treatment is a time-consuming process that requires the use of several chemotherapeutics, to which mycobacteria are becoming increasingly resistant. This article focuses on the potential utility of biomarkers of mycobacterial origin with potential implications for TB diagnosis. Properly standardized indicators could become new diagnostic tools, improving and streamlining the identification of Mycobacterium tuberculosis infection and the implementation of appropriate therapy. These markers can also potentially provide a quick confirmation of effectiveness of new anti-mycobacterial drugs and TB vaccines, leading to a possible application in practice.

Two-Year Follow-up Study of Mycobacterium tuberculosis Antigen-Driven IFN-γ Responses and Macrophage sCD14 Levels After Tuberculosis Contact.

Clinical data regarding the prediction of active tuberculosis (TB) development in close TB contacts are scarce. To address this problem, we performed a 2-year follow-up study of Mycobacterium tuberculosis (M.tb) antigen-driven IFN-gamma responses and serum levels of soluble macrophage CD14 receptor in individuals with recent or prolonged M.tb exposure. Between June 2011 and June 2013, we studied 60 healthy Polish adults with recent household or long-term work TB contact and individuals without known M.tb exposure. All of them underwent baseline and repeated testing with IGRA (IFN-gamma release assay) and serum sCD14 ELISA quantification. Frequencies of IGRA results differed at the baseline and follow-up testing. IGRA reversions were noticed in almost one-third of Work TB Contacts and no participants from the Household TB Contact group. IGRA conversions were found in 40 % of Household TB Contacts. No correlation between the IGRA result and the sCD14 level was observed. IFN-γ variability has important implications for clinical practice and requires caution in interpreting the results to distinguish new infections from nonspecific inter-individual variations in cytokine responses. The impairment of IFN-γ response in some individuals with prolonged M.tb exposure representing a resistant immune status does not allow considering IGRA results as reliable and credible. Monitoring the serum sCD14 level can reduce the likelihood of a false prediction of active TB development in close TB contacts showing an M.tb-specific increase in the IFN-gamma production in repeated IGRA testing.

Monocyte signal transduction receptors in active and latent tuberculosis.

The mechanisms that promote either resistance or susceptibility to TB disease remain insufficiently understood. Our aim was to compare the expression of cell signaling transduction receptors, CD14, TLR2, CD206, and ß2 integrin LFA-1 on monocytes from patients with active TB or nonmycobacterial lung disease and healthy individuals with M.tb latency and uninfected controls to explain the background of the differences between clinical and subclinical forms of M.tb infection. A simultaneous increase in the expression of the membrane bound mCD14 receptor and LFA-1 integrin in patients with active TB may be considered a prodrome of breaking immune control by M.tb bacilli in subjects with the latent TB and absence of clinical symptoms.

Interaction of Helicobacter pylori with C-type lectin dendritic cell-specific ICAM grabbing nonintegrin.

In this study we asked whether Helicobacter pylori whole cells and lipopolysaccharide (LPS) utilize sugar moieties of Lewis (Le) antigenic determinants to interact with DC-SIGN (dendritic cell specific ICAM grabbing nonintegrin) receptor on dendritic cells (DCs). For this purpose the soluble DC-SIGN/Fc adhesion assay and the THP-1 leukemia cells with induced expression of DC-SIGN were used. We showed that the binding specificity of DC-SIGN with H. pylori Le(X/Y) positive whole cells and H. pylori LPS of Le(X/Y) type was fucose dependent, whereas in Le(XY) negative H. pylori strains and LPS preparations without Lewis determinants, this binding was galactose dependent. The binding of soluble synthetic Le(X) and Le(Y) to the DC-SIGN-like receptor on THP-1 cells was also observed. In conclusion, the Le(XY) dependent as well as independent binding of H. pylori whole cells and H. pylori LPS to DC-SIGN was described. Moreover, we demonstrated that THP-1 cells may serve as an in vitro model for the assessment of H. pylori-DC-SIGN interactions mediated by Le(X) and Le(Y) determinants.

Helicobacter pylori antigens as potential modulators of lymphocytes' cytotoxic activity.

Helicobacter pylori (H.p) colonizes human gastric mucosa and causes gastric and duodenal ulcer disease or gastric cancer. Various H.p compounds may modulate the host immune response in regards to tolerance of the infection or disease development. The aim of this study was to determine whether H.p lipopolysaccharide (LPS) and glycine acid extract antigens (GE) or E. coli LPS influence the cytotoxic activity of peripheral blood lymphocytes from H.p infected - H.p (+) or uninfected - H.p (-) individuals, in the presence or absence of exogenous interleukin (IL)12. Individual H.p status was defined by the urea breath test. Lymphocytes, stimulated or not with H.p, and control antigens, with or without IL-12, were used as effector cells and epithelial HeLa cells as targets. The cytotoxicity of lymphocytes was expressed as the percentage of dead target cells unable to reduce tetrazolium salt. The supernatants from HeLa/lymphocyte cultures were used for detection of the cellular cytotoxicity markers granzyme B and caspase 8. The natural cytotoxic activity of lymphocytes from H.p (+) was less than that of H.p (-) donors. This may have been due to fewer natural killer cells of CD3(-) CD56(+) Nkp46(+) phenotype in H.p (+) in comparison to H.p (-) subjects. H.p GE and standard E. coli LPS enhanced the cytotoxicity of lymphocytes towards target cells whereas H.p LPS downregulated this activity. The decrease in lymphocyte cytotoxicity in response to H.p LPS correlated with a lack of IL-2 and IL-12 production, inhibition of interferon-γ production, and low IL-10 secretion by mononuclear leukocytes. IL-12 significantly enhanced the natural as well as H.p LPS and H.p GE driven cytotoxic capacity of lymphocytes. In conclusion, H.p LPS may negatively modulate natural cytotoxic activity and cytokine secretion by immunocompetent cells and thus be involved in the maintenance of infection and development of gastric pathologies.

Latent M. tuberculosis infection--pathogenesis, diagnosis, treatment and prevention strategies.

One third of the earths population is infected with Mycobacterium tuberculosis (Mtb), but only 5-10% of the infected individuals develop active tuberculosis (TB) over their lifetime. The remaining 90-95% stay healthy and are called latently infected individuals. They are the biggest reservoir of the tubercle bacilli and identifying the cases of latent TB is a part of the global plan of TB control. From the clinical point of view detection of latent TB infections (LTBI) in individuals with the highest active TB risk including cases of HIV infection, autoimmune inflammatory diseases or cancer, is a priority. This review summarizes the recent findings in the pathogenesis of latent TB, its diagnosis, treatment and prevention.

Mycobacterium bovis Wild-Type BCG or Recombinant BCG Secreting Murine IL-18 (rBCG/IL-18) Strains in Driving Immune Responses in Immunocompetent or Immunosuppressed Mice.

Mycobacterium tuberculosis infections remain a global health problem in immunosuppressed patients. The effectiveness of BCG (Bacillus Calmette−Guérin), an anti-tuberculosis vaccine, is unsatisfactory. Finding a new vaccine candidate is a priority. We compared numerous immune markers in BCG-susceptible C57BL/6 and BCG-resistant C3H mice who had been injected with 0.9% NaCl (control) or with wild-type BCG or recombinant BCG secreting interleukin (IL)-18 (rBCG/IL-18) and in immunized mice who were immunocompromised with cyclophosphamide (CTX). The inoculation of rBCG/IL-18 in immunocompetent mice increased the percentage of bone marrow myeloblasts and promyelocytes, which were further elevated in the rBCG/IL-18/CTX-treated mice: C57BL/6 mice­3.0% and 11.4% (control) vs. 18.6% and 42.4%, respectively; C3H mice­1.1% and 7.7% (control) vs. 18.4% and 44.9%, respectively, p < 0.05. The bone marrow cells showed an increased mean fluorescence index (MFI) in the CD34 adhesion molecules: C57BL/6 mice­4.0 × 103 (control) vs. 6.2 × 103; C3H mice­4.0 × 103 (control) vs. 8.0 × 103, p < 0.05. Even in the CTX-treated mice, the rBCG/IL-18 mobilized macrophages for phagocytosis, C57BL/6 mice­4% (control) vs. 8%; C3H mice­2% (control) vs. 6%, and in immunocompetent mice, C57BL/6 induced the spleen homing of effector memory CD4+ and CD8+ T cells (TEM), 15% (control) vs. 28% and 8% (control) vs. 22%, respectively, p < 0.05. In conclusion, rBCG/IL-18 effectively induced selected immune determinants that were maintained even in immunocompromised mice.

[Recognition of mycobacterial antigens by phagocytes]. / Rozpoznawanie antygenów pratków przez fagocyty.

Recognition of mycobacterial antigens by receptors of phagocytes is not only a key element of the first line of defense, but also an important link to the specific phase of the immune response. The immune response is based on the existence of a number of pattern recognition receptors (PRR) that recognize conservative microbial structures called pathogen-associated molecular patterns (PAMP). These receptors are involved in the processes of opsonization and phagocytosis of pathogens, activation of the complement system, induction of apoptosis and signal transduction cell systems. The initiated signal cascade is supposed to lead to the mobilization of immune forces against the penetrating pathogen and is aimed at its fast elimination from the body. Understanding the role of these receptors in the antimycobacterial immune response appears to be fully justified in view of their potential application in distinguishing persons particularly sensitive to tuberculosis as well as in the development of new generation vaccines for prophylaxis and therapy and new biomarkers for improvement of the difficult and time-consuming diagnosis of mycobacterial infections.

[Nontuberculous mycobacteria: M. marinum, M. ulcerans, M. xenopi - brief characteristics of the bacteria and diseases caused by them]. / Pratki niegruzlicze: M. marinum, M. ulcerans, M. xenopi ­ krótka charakterystyka drobnoustrojów i zmian klinicznych przez nie wywolywanych.

Mycobacterium is a variable group of acid-fast bacillus which contains pathogenic bacteria causing tuberculosis (MTC - Mycobacterium tuberculosis complex) and leprosy (M. leprae) as well as numerous nontuberculous mycobacteria (NTM) causing diseases mostly in people with immunodeficiency, although some NTM strains are capable of causing illnesses in non-immunocompromised patients. This group includes for example Mycobacterium marinum, Mycobacterium ulcerans and Mycobacterium xenopi. These microorganisms are environmental mycobacteria, present in developing countries of Africa, but they may also be transferred to other continents. The most common symptoms of diseases caused by these species are skin lesions (hyperpigmentation, tumors, ulcers) and arthritis. Because of the rarity of their occurrence, these mycobacteria are relatively poorly known. Effective ways of treating the diseases caused by these bacilli are still under study.

[Vaccination against M. tuberculosis - what next after BCG?]. / Szczepienia przeciwpratkowe ­ BCG i co dalej?

Tuberculosis (TB) still remains a huge global health problem. An increase in TB has been observed in many parts of the world, especially in poor and densely populated sub-Saharan Africa and Asia. Tuberculosis affects not only the developing countries but also the relatively wealthy regions of Europe, particularly Eastern Europe, where drug-resistant mycobacterial strains are increasingly reported. Control of tuberculosis expansion is very difficult. It requires the long-term use of anti-mycobacterial drugs. Additionally, the HIV epidemic and the phenomenon of multi-drug resistance are assumed to be responsible for the increase in TB cases. Therefore the most reasonable form of anti-TB protection seems to be effective vaccination. At the beginning of the twentieth century the BCG vaccine was introduced into general use as the first and so far the only immune protector against tuberculosis. Now it is known that this vaccine is not powerful enough and induces protection at a relatively low level. Hence ongoing research on the development of a more powerful anti-mycobacterial vaccine is still needed. Many of the new formulations are in phase II or III of clinical trials and the results are promising. The search for new vaccines involves several strategies: modified virulence-attenuated Mycobacterium tuberculosis strains, recombination of attenuated M. bovis BCG bacilli, immunogenic mycobacterial proteins and DNA encoding selected proteins as well as unrelated microorganisms used as carriers of mycobacterial antigens. The wide range of concepts is extremely important because new vaccines should serve for immunization of the broadest possible population, not only healthy individuals but also those who are immunocompromised.

Cytokine Biosignature of Active and Latent Mycobacterium Tuberculosis Infection in Children.

None of the currently used diagnostic tools are efficient enough in diagnosing Mycobacterium tuberculosis (M.tb) infection in children. The study was aimed to identify cytokine biosignatures characterizing active and latent tuberculosis (TB) in children. Using a multiplex bead-based technology, we analyzed the levels of 53 Th17-related cytokines and inflammatory mediators in sera from 216 BCG-vaccinated children diagnosed with active TB (TB) or latent TB (LTBI) as well as uninfected controls (HC). Children with active TB, compared to HC children, showed reduced serum levels of IL-17A, MMP-2, OPN, PTX-3, and markedly elevated concentrations of APRIL/TNFSF13. IL-21, sCD40L, MMP-2, and IL-8 were significantly differentially expressed in the comparisons between groups: (1) HC versus TB and LTBI (jointly), and (2) TB versus LTBI. The panel consisting of APRIL/TNFSF13, sCD30/TNFRSF8, IFN-α2, IFN-γ, IL-2, sIL-6Rα, IL-8, IL-11, IL-29/IFN-λ1, LIGHT/TNFSF14, MMP-1, MMP-2, MMP-3, osteocalcin, osteopontin, TSLP, and TWEAK/TNFSF12 possessed a discriminatory potential for the differentiation between TB and LTBI children. Serum-based host biosignatures carry the potential to aid the diagnosis of childhood M.tb infections. The proposed panels of markers allow distinguishing not only children infected with M.tb from uninfected individuals but also children with active TB from those with latent TB.

Decrease of IL-5 Production by Naive T Cells Cocultured with IL-18-Producing BCG-Pulsed Dendritic Cells from Patients Allergic to House Dust Mite.

The only currently available anti-tuberculosis vaccine, Bacillus Calmette-Guérin (BCG), has been reported to also protect against unrelated diseases, including inflammatory diseases such as allergic asthma. Recombinant BCG strains that produce IL-18 have been shown to enhance Th1 responses over non-recombinant BCG and to reduce IL-5 production and bronchoalveolar eosinophilia in mice. However, their ability to decrease the immune polarization of human Th2 cells is not known. Here, we show that BCG and recombinant BCG producing human IL-18 (rBCG-hIL-18) induced the maturation of Der p 1-stimulated monocyte-derived dendritic cells (MD-DCs) from healthy controls and from patients allergic to house dust mites. After incubation with mycobacteria and Der p 1, MD-DCs produced significantly more IL-23 and IP-10 but had no effect on IL-12p70 or IL-10 production compared to Der p 1-pulsed MD-DCs in the absence of mycobacteria. In the presence of Der p 1, BCG- and rBCG-hIL-18-pulsed MD-DCs cocultured with naive, but not with memory T cells from allergic patients, resulted in a decrease in IL-5 production compared to non-pulsed MD-DCs cultured in the presence of Der p 1. BCG, and especially rBCG-hIL-18, may thus be potential therapeutic tools to reduce exacerbated Th2 responses in patients with allergic asthma.

IL-18 and related function proteins associated with tuberculosis severity and screening for active TB among patients with non-mycobacterial community-acquired pneumonia (CAP).

BACKGROUND: Differentiation of active pulmonary tuberculosis (TB) from non-mycobacterial community-acquired pneumonia (CAP) still remains a diagnostic challenge. OBJECTIVE: The study aimed to quantify the IL-18, IFN-γ, IL-18BP, IL-37, and IP-10 levels in serum and Mycobacterium tuberculosis (M.tb) antigens-stimulated blood cultures from TB or CAP patients and explore if the proteins can be a useful basis for discriminating these diseases. METHODS: In total, 124 Polish adults, including mild/moderate (M/MTB) or advanced (ATB) TB patients, and CAP patients, were enrolled in the study. The concentrations of IL-18, IL-18BP, IFN-γ, IL-37, and IP-10 in sera and M.tb-stimulated cultures were measured by ELISA. RESULTS: The most specific and sensitive serum proteins discriminating TB from CAP were IP-10 and IL-18BP; however, IP-10 had the highest AUC in the ROC curve for the diagnosis. Serum IP-10 and IL-18BP levels increased significantly in M/MTB or ATB groups. The IL-18BP elevation in ATB group was accompanied by an increase in IL-18. No single protein measured in M.tb-stimulated cultures differed TB from CAP patients. CONCLUSIONS: The combined analysis of serum IL-18BP and IP-10 might be considered as an auxiliary tool in the differentiation of TB from CAP.

Interleukin-18, Functional IL-18 Receptor and IL-18 Binding Protein Expression in Active and Latent Tuberculosis.

A thorough understanding of the processes modulating the innate and acquired immune response to Mycobacterium tuberculosis (M.tb) infection in the context of gene expression is still a scientific and diagnostic problem. The study was aimed to assess IL-18, IL-18 binding protein (IL-18BP), IL-18R, IFN-γ, and IL-37 mRNA expression in patients with active tuberculosis (ATB) and healthy volunteers with latent M.tb-infection (LTB) or M.tb-uninfected healthy controls (Control). The relative mRNA expression was assessed in the buffy coat blood fraction using the qPCR method. In total, 97 BCG-vaccinated Polish adults were enrolled in the study. The relative expression of IL-18 and IL-18BP mRNA was significantly elevated in the ATB and LTB groups. In ATB, but not LTB individuals, the overexpression of IL-18 and IL-18BP, as well as a significant increase in IFN-γ mRNA expression, might be considered as a manifestation of active tuberculosis disease. No statistically significant differences were observed in the IL-37 mRNA expression among the studied groups. Particularly noteworthy is the outstanding reduction in the relative expression of IL-18R mRNA in the LTB group as compared to the ATB and Control group. Reduced expression of IL-18R in LTB group may, at least partially, prevent the development of a pathological inflammatory reaction and promote the maintenance of homeostatic conditions between host immunity and M.tb.

Interleukin-1b and interleukin-1 receptor inhibitor gene cluster polymorphisms in patients with coronary artery disease after percutaneous angioplasty or coronary artery bypass grafting.

BACKGROUND: Pro-inflammatory cytokine interleukin-1b (IL-1b) plays a role in atherosclerosis. The results of several studies on the association between polymorphism of the IL-1b gene cluster and the course of coronary atherosclerosis have been inconclusive. AIM: To investigate retrospectively whether the patients with the most common variants of polymorphism of the IL-1b gene cluster differ with respect to localisation and extent of coronary atherosclerosis to a degree which may influence the treatment strategy. METHODS: Ninety-two consecutive out-patients (age 39-83, male sex 74%) with coronary artery disease confirmed by angiography were included. In this group, 23 patients underwent coronary artery bypass grafting (CABG) and 69 percutaneous coronary interventions (PCI) of whom in 16 repeated treatment was performed. The polymorphisms of the IL-1b gene - transition C/T at -511 and -31 position - as well as of the IL-1 receptor antagonist gene (IL-1RN) - an 86-base pair variable-number tandem repeat in intron 2 - were determined by PCR. Out of the 54 theoretically possible combinations of polymorphisms, 17 were found in the studied group. The three most common combinations of polymorphisms were selected. The fraction of patients treated by means of primary or elective percutaneous coronary intervention (pPCI, ePCI) and by means of CABG were compared between the subgroups with one of the 3 most common combinations of polymorphisms. RESULTS: The most frequent combinations of polymorphisms were - Variant A: -31 C/T, -511C/T, RN 1/1 - 32.6%; Variant B: -31T/T, -511C/C, RN 1/1 - 27.1%; Variant C: -31C/T, -11C/T, RN 1/2 - 10.8%. The remaining patients (29.5%) represented 14 variants present in very small subgroups consisting only of 1, 2 or 3 persons. Statistical analysis showed that patients with the second most common variant of studied polymorphisms (variant B) were significantly more frequently treated with CABG in comparison to the two other variants. Also, repeated PCI was most frequent in this subgroup. CONCLUSION: The data presented here suggest that carriers of the two relatively frequent variants of the IL-1b gene at -31 and -511 position, i.e. -31TT and -511CC, are at a higher risk of developing coronary artery disease requiring surgical treatment or two-stage percutaneous angioplasty.

Lack of significant effect of interleukin-18 gene variants on tuberculosis susceptibility in the Polish population.

BACKGROUND: Polymorphisms in genes encoding cytokines are known to determine susceptibility to Mycobacterium tuberculosis (M.tb) infection. In particular, interleukin-18 (IL-18), an inducer of interferon-gamma (IFN-γ), playing an important role in anti-mycobacterial immune responses, may influence the risk of developing active tuberculosis (TB). AIM: A case-control study was performed to investigate whether two promoter polymorphisms of the IL-18 gene at positions -137G/A (rs187238) and -607A/C (rs1946518) affect the serum level of IL-18 and might be associated with genetic susceptibility to tuberculosis (TB) in the Polish population. METHODS: Two IL-18 gene promoter SNPs were detected by an allele-specific polymerase chain reaction. Serum IL-18 levels were measured immunoenzymatically using Human Total IL-18 ELISA DuoSet. RESULTS: A single-gene analysis showed no differences either in allele or genotype frequencies of the studied SNPs between TB patients and healthy controls. No significant differences in the frequencies of any of the haplotypes between TB patients and healthy controls were found. None of the polymorphic variants of IL-18(-137G/A) or IL-18(-607A/C) SNP was associated with IL-18 producing capability. CONCLUSION: Our results suggest that IL-18(-137G/A) and IL-18(-607A/C) polymorphisms may not be risk factors for susceptibility to TB in the Polish population. Increased serum IL-18 level observed in TB patients has no genetic background, but is a consequence of M.tb infection. Further studies with a higher sample size are required to confirm these findings.

IL-18/IL-37/IP-10 signalling complex as a potential biomarker for discriminating active and latent TB.

BACKGROUND: Currently, there are serious limitations in the direct diagnosis of active tuberculosis (ATB). We evaluated the levels of the IL-18/IL-37/IP-10 signalling complex proteins in Mycobacterium tuberculosis (M.tb)-specific antigen-stimulated QuantiFERON® Gold In-Tube (QFT) cultures and in serum samples from ATB patients, healthy individuals with latent M.tb infection (LTBI) and healthy controls (HC) to examine whether combined analyses of these proteins were useful in the differentiation of M.tb states. METHODS: The concentrations of IL-18, IL-18BP, IFN-γ, IL-37 and IP-10 in the serum and QFT supernatants were measured using specific enzyme-linked immunosorbent assay (ELISA) kits. Free IL-18 levels were calculated using the law of mass action. RESULTS: Increased concentrations of total and free IL-18, IL-18BP, IFN-γ and IP-10 in the sera of ATB patients were detected. These increases were not counterbalanced by the overproduction of IL-37. Complex co-expression of serum IL-18BP and IL-37, IP-10 and IFN-γ was identified as the highest discriminative biomarker set for the diagnosis of ATB. CONCLUSIONS: Our results suggest that the IL-18 signalling complex might be exploited by M. tuberculosis to expand the clinical manifestations of pulmonary TB. Therefore, direct analysis of the serum components of the IL-18/IL-37 signalling complex and IP-10 may be applicable in designing novel diagnostic tests for ATB.

Anti-phagocytic activity of Helicobacter pylori lipopolysaccharide (LPS)--possible modulation of the innate immune response to these bacteria.

The Helicobacter pylori infections are followed by an infiltration of the gastric mucosa by neutrophils and macrophages. Accumulation of phagocytes enables them to interact with H. pylori, but a great number of infected subjects cannot eradicate these bacteria. The H. pylori inhibits its own uptake by blocking the function of phagocytes. The anti-phagocytic mechanism depends on bacterial surface structures and the presence of the cag pathogenicity island (PAI). The role of H. pylori lipopolysaccharide (LPS), during phagocytosis of these bacteria is not clear. LPS may mediate direct bacteria/phagocyte interactions and it may also regulate antibacterial activity of the phagocytes. In this study we investigated the influence of H. pylori LPS on phagocytosis of these bacteria. The H. pylori LPS inhibited an ingestion of these microbes by human peripheral blood granulocytes. This was correlated with a diminished ability of phagocytes to reduce MTT-tetrazolium salt. The anti-phagocytic effect of H. pylori LPS was reduced by recombinant lipopolysaccharide binding protein (rLBP). It is possible that in vivo H. pylori LPS may diminish elimination of these bacteria from the gastric mucosa promoting an infection persistence. However, LBP may modulate the uptake of H. pylori due to neutralization of anti-phagocytic effect of its LPS.

Tuberculin skin test reaction is related to memory, but not naive CD4+ T cell responses to mycobacterial stimuli in BCG-vaccinated young adults.

Bacillus Calmette-Guérin (BCG) is the only vaccine available against tuberculosis and the tuberculin skin test (TST) is the most widely used method to detect BCG take. However, subjects may remain TST-negative, even after several BCG administrations. To investigate some of the potential reasons underlying this inability of developing tuberculin sensitivity in response to BCG we compared the effect of different mycobacterial stimuli in the groups differently responding to tuberculin. TST was performed on 71 healthy adults aged 25-30 years, who had received BCG in their childhood, and considered TST-positive at ≥10 mm. Dendritic cells (DCs) were incubated with PPD, live BCG or rBCGhIL-18, producing human IL-18. The latter strain was used to investigate whether the production of IL-18 could overcome some of the immune read-out limitations in the TST-negative subjects. CD86, CD80, CD40, and DC-specific intracellular adhesion molecule-3 grabbing nonintegrin (DC-SIGN) expression was analysed by flow cytometry and IL-10, IL-23 and IP-10 secretion in culture supernatants by ELISA. In DCs-T cell co-cultures with naive and memory CD4+ T cells, the IFN-γ and IL-10 levels were determined by ELISA. We found no difference in IL-10 and IFN-γ production by naive T cells between the TST-negative and TST-positive subjects. However, IFN-γ was produced in significantly higher amounts by memory T cells incubated with PPD, BCG or rBCGhIL-18-pulsed DCs in TST-positive than in TST-negative subjects, whereas the numbers of the IFN-γ-producing T cells were similar in both groups. This difference may be partially due to a decreased CD40 and enhanced reduction in DC-SIGN expression by DCs of TST-negative versus TST-positive subjects. A strong effect of IL-18 expression by rBCGhIL-18 on IL-23 production by the DC was seen in both groups, which likely was the reason for the increased IFN-γ production by naïve T cells upon incubation with mycobacteria-pulsed DC, regardless of the TST status.