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.

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.

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.

CD14-159C/T polymorphism in the development of delayed skin hypersensitivity to tuberculin.

The skin tuberculin test (TST), an example of a delayed-type hypersensitivity (DTH) reaction, is based on measuring the extent of skin induration to mycobacterial tuberculin (PPD). Little is known about the genetic basis of TST reactivity, widely used for diagnosing TB infection. The study investigated the relationship of the single base change polymorphic variants in CD14 gene (CD14(-159C/T)) with the development of DTH to PPD in BCG-vaccinated Polish Caucasian individuals. We found persistent lack of TST reactivity in about 40% of healthy subjects despite receiving more than one dose of BCG. The TST size was negatively correlated with the number of BCG inoculations. The distribution of C/T genotype was significantly more frequent among TST-negative compared with TST-positive individuals. The concentration of serum sCD14 was positively associated with mCD14 expression, but not with the TST status or CD14(-159C/T) polymorphism. A significant increase in mCD14 expression and serum sCD14 levels was found in TB group. We hypothesize that CD14(-159C/T) polymorphic variants might be one of genetic components in the response to attenuated M. bovis BCG bacilli.

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.

Interferon-gamma assay in combination with tuberculin skin test are insufficient for the diagnosis of culture-negative pulmonary tuberculosis.

OBJECTIVE: Early diagnosis of infectious cases and treatment of tuberculosis (TB) are important strategies for reducing the incidence of this disease. Unfortunately, traditional TB diagnostic methods are time-consuming and often unreliable. This study compared the accuracy and reliability of the tuberculin skin test (TST) and interferon (IFN)-γ-based assay (IGRA) for the diagnosis of active pulmonary TB Polish cases that could or could not be confirmed by M. tuberculosis (M.tb) culture. METHODS: In total, 126 adult patients with clinically active TB or non-mycobacterial, community-acquired lung diseases (NMLD) hospitalised at the Regional Specialised Hospital of Tuberculosis, Lung Diseases and Rehabilitation in Tuszyn, Poland were enrolled in the present study. Sensitivity, specificity, positive predicted value (PPV), negative predicted value (NPV), and analytic accuracy (Acc) of TST and IGRA testing for the diagnosis of culture-positive and culture-negative TB patients were calculated. The quantities of IFN-γ produced in the response to M.tb specific antigens (TB Ag - Nil) in the cultures of blood from patients with active TB and NMLD patients were also analysed. RESULTS: The IGRA sensitivity in culture-positive and culture-negative TB patients was similar, measuring 65.1% and 55.6%, respectively. The sensitivity of TST did not differ from the parameters designated for IGRA, measuring 55.8% in culture-positive and 64.9% in culture-negative TB. The sensitivity of TST and IGRA was age-dependent and decreased significantly with the age of the patients. No differences in the frequency or intensity of M.tb-stimulated IFN-γ production, as assessed by IGRA testing between culture-positive and culture-negative TB were noticed. Significantly lower concentrations of IFN-γ were observed in patients with advanced TB forms compared with those with mild or moderate TB pathologies. CONCLUSIONS: Our results do not show that a combination of IGRA and TST might be a step forward in the diagnosis of culture-negative TB cases. However, M. tuberculosis-stimulated IFN-γ levels might help to assess the extent of pulmonary TB lesions.

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.