| Object: to assess the contents of interferon-gamma(IFN-γ), Tumor necrosis factor-alpha (TNF-α), interleukin-8 (IL-8), interleukin-17 (IL-17) in the plasma and sputum of pulmonary tuberculosis patients with initial treatment, and the messenger RNA (mRNA) expression of these cytokines in patients with pulmonary tuberculosis(PTB); to observe the variations and features of these cytokines; to analyze the relevant influencing factors; to understand the immunologic mechanism of the anti-tuberculosis process; to estimate the value of the cytokines in clinical application; finally to provide the basis of the prehension of the immunological mechanisms of PTB, the auxiliary tuberculosis diagnosis, and the evaluation for the efficacy of anti-TB treatment.Method: The subjects were 44 patients with secondary pulmonary tuberculosis initially treated, and there were 43 normal controls and 18 lung cancer patients as control groups. To detect the levels of IFN-γ, IL-17, TNF-αand IL-8 in plasma and sputum of the three target groups with ELISA; to check out the expression of the mRNA in PBMC and cells in the sputum of the three groups by RT-PCR; in the normal control group, to distinguish the latent tuberculosis infection(LTBI) by Enzyme-Linked Immune spot (ELISPOT)assay; to check out the mycobacterial load in sputum of the patients with Fluorescent Quantitation Polymerase Chain Reaction(FQ-PCR); Statistical analysis was performed to analyze the results and the relevant factors, and the diagnostic value of the four cytokines were also analyzed.Result:1. In the normal control group, IFN-γlevels in Plasma and sputum was similar (P>0.05), while plasma IL-17, TNF-α, IL-8 levels were significantly higher than those in sputum (P<0.01). In addition, the expression of the mRNA in PBMC and in sputum cells of IFN-γ, IL-17, IL-8 were close (P>0.05), the mRNA expression of TNF-αin PBMC was significantly higher than that of sputum cells (P<0.05).2. Comparing with the normal control group, levels of IFN-γ,IL-8 in plasma of pulmonary tuberculosis patients were higher and levels of IL-17, TNF-αin plasma were lower; levels of IFN-γin sputum were lower and levels of IL-17,TNF-α,IL-8 in sputum of pulmonary tuberculosis patients were higher. In the pulmonary tuberculosis group, comparing with the levels in plasma, levels of IL-17 and TNF-αin sputum were higher and levels of IFN-γand IL-8 in sputum were lower(P<0.01 or P<0.05).3. Influencing factors: (1) In the normal control group, The contents of the four cytokines IFN-γ, IL-17, TNF-α, IL-8 in plasma and sputum from the LTBI group were closed to that of the healthy groups(P>0.05). (2)only the expression of the mRNA of IFN-γ, IL-17in PBMC from PTB patients were higher than those in the normal control group (P <0.05), the expression of the mRNA of TNF-α, IL-8 in PTB patients were similar to those in the normal control group (P> 0.05).(3) The contents of IFN-γ(r=0.843)and IL-8(r=0.861) in plasma had a positive correlation with the mycobacteria level, the plasma contents of TNF-α(r=-0.788) and IL-17(r=-0.816) had a negative correlation with the mycobacteria level (P<0.01); The contents of IFN-γ(r=-0.893) in sputum had a negative correlation with the mycobacteria level, the sputum TNF-α,IL-8,IL-17 contents had a positive correlation with the mycobacterial level, (r values was 0.631,0.889,0.756, respectively)(P<0.01).(4) The levels of the four cytokines in plasma and sputum from the PTB group with cavitation were close to those without it(P>0.05).(5) The contents of IFN-γand IL-8 in the sputum of those patients whose peripheral blood lymphocyte counts were lower, were lower than the group whose peripheral blood lymphocyte counts was normal(P<0.01 and P<0.05 respectively), while the contents of TNF-α,IL-17 in sputum carried no difference between the two groups(P>0.05).(6) For the PTB patients, two weeks after the treatment, the contents of the TNF-αand IL-17 in plasma were higher and the contents of the IL-8 in plasma were lower than before, which carried a statistical difference(P<0.05). The contents of the IFN-γin the plasma and the four cytokines in sputum were similar before and after the treatment(P>0.05).4. Comparing with the lung cancer group, the levels of IFN-γ, IL-8 in the plasma of PTB patients were higher, and those of IL-17 and TNF-αwere lower(P<0.01 or P<0.05); the level of IFN-αin their sputum was lower, while levels of IL-17 and TNF-αin their sputum were higher(P<0.01 or P<0.05); the contents of IL-8 in sputum carried no statistical difference between these two groups(P>0.05). Receiver operating characteristic curve (ROC curve) analysis suggested that when the contents of IFN-γ, TNF-α, IL-8 and IL-17 in plasma and sputum were transformed into the area under the curve (AUC) which were 0.778～0.958, it could be an operative diagnosis index(P<0.01). The sensitivities were all above 68%, specificities were all above 66% and accuracies were all above 76%. Among them, the sensitivity of sputum TNF-αwas the highest, 90%; IL-17 in sputum carried the highest specificity, 97%; IL-17 in sputum also carried the highest diagnostic accuracy, which was 93%. The combination indexes of the cytokines in plasma and sputum could improve the sensitivity and specificity of a diagnosis.Conclusion:1. In the normal control group, The level of IFN-γin plasma which represents the general immunity status is consistent with that in sputum that represents the local immunity status. However, differences between the levels of IL-17, TNF-α, IL-8 in plasma and in sputum suggested that their participation in the local immunity is lower than that in the systemic immunity.2. IFN-γ, IL-17,TNF-αand IL-8 were closely related to the incidence and the development of tuberculosis. Among them, the increase of specific immunity which resulted from the increasing level of IFN-γin the plasma of PTB patients can hardly compensate for the reducing level of some specific immunity resulting from the lessening level of TNF-αand IL-17. And it is also difficult to antagonize the immune damage caused by the increasing level of IL-8.The increasing levels of IL-17,TNF-αand IL-8 in the sputum of PTB patients, the reducing level of IFN-γare closely related to the local lesion formation in an infection site caused by Both inhibition of the activation of macrophages and mediation of the immune injury.3. The levels of IFN-γ,IL-17,TNF-αand IL-8 in plasma and in sputum of the patients with PTB were affected by many factors. Among them, IFN-γand IL-8's high expressions of the mRNA in plasma was in correlation with the increasing level of plasma. Bacterial counts carried an influence on the increasing levels of IFN-γ,IL-8 in plasma and the reducing levels of IL-17,TNF-αin plasma, bacterial counts affected the reduction of IFN-γand the increase of IL-17,IL-8, TNF-αin sputum. Low levels of lymphocytes had a certain extent to the reducing level of IL-8 and the increase of IFN-γin sputum. Anti-tuberculosis treatment in patients can make the plasma cytokine levels to be nearly normal. The latent Mycobacterium tuberculosis infection had no significant effect on the four cytokines in plasma and sputum, and the prominent role of cytokines had not been seen on cavity formation.4. Detection of the levels of IFN-γ, IL-17, TNF-αand IL-8 in plasma and sputum in Patients with pulmonary tuberculosis carries some certain clinical significance. The changes of the cytokines levels in plasma before and after treatment indicate that dynamic monitoring or observation the four cytokines levels in plasma may serve as early assessment markers for the efficacy of anti-TB treatment. The cutoff values which we can get from the application of ROC curve can be used as diagnosis indicators for tuberculosis, especially for culture negative pulmonary tuberculosis.
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