Immunosuppressive Molecule Regulate The Pathogenesis Of Active Tuberculosis And Its Application In Clinical Diagnosis

[Objective] Assess the immune function of tuberculosis patients and healthy people.[Methods] Fresh peripheral blood was collected from 23 healthy volunteers and 38 TB patients.Peripheral blood mononuclear cells(PBMCs)were isolated by Ficoll-Hypaque density gradient centrifugation.Flow cytometry was used to assess cellular activating molecules,inhibitory molecules,and cellular functions.The concentrations of IFN-?,IL-10,IL-12p70,IL-13,IL-1?,IL-2,IL-4,IL-6,IL-8 and TNF-? in serum were detected by electrochemiluminescence method.[Results] The secretion of IFN-?,IL-10,IL-12p70,IL-13,IL-1?,IL-2,IL-4,IL-6,IL-8 and TNF-? in tuberculosis patients was significantly higher than that in healthy controls.The effector cell function of tuberculosis patients showed a general weakening trend,including CD4+,CD8+,NK,DC,and Mon.However,regulatory T cells(Tregs)and tuberculosis-specific CD4+,CD8+ and NK cell functions are significantly enhanced.CTLA-4,TIM-3,TIGIT,and PD-1 were significantly higher in CD4+,CD8+,NK,and Tregs than in healthy volunteers.[Conclusion] The immune function of tuberculosis patients involves a variety of inflammatory factors,cells and molecules,which are heterogeneous.The imbalance of cell function in tuberculosis patients,that is,the effector cell function is significantly weakened,and the inhibitory cell function is enhanced.[Objective] Secondly,we study the effect of TIGIT in the role and mechanisms of regulatory T and T cells in active tuberculosis.[Methods] 85 healthy volunteers and 52 TB patients were collected.The relationship between TIGIT expression and activation and proliferation of Tregs and CD4+ effector cells(Teffs)was detected by flow cytometry.The TIGIT Fc fusion protein or CD155 fusion protein or control Ig G was incubated with PBMCs of tuberculosis patients to observe the effects of blocking or activating TIGIT signaling pathway on Tregs and Teffs cell activation,cytokine secretion and proliferation function.Finally,blocking the TIGIT signaling pathway of CD4+ T cells and observing the ability of macrophages to control the growth of Mtb[Results] TIGIT in tuberculosis patients is highly expressed on Tregs and is positively correlated with the activation and function of Tregs.Blocking the TIGIT signaling pathway alone can reduce the function of Tregs,including IL-10 secretion and proliferation,but has no effect on Teffs function.Although blocking TIGIT has no effect on Teffs,blocking the TIM-3 signaling pathway restores Teffs function.In addition,the TIGIT pathway that blocks CD4+ T cells enhances the ability of macrophages to control Mtb growth.[Conclusion] TIGIT expressed on CD4+ T cells is likely to play a role in tuberculosis mainly through Tregs rather than Teffs.[Objective] The effect of inhibitory molecules on T-SPOT results was investigated.[Methods] Incorporate 32 healthy volunteers,24 patients with tuberculosis T-SPOT high response(TBAg/PHA?0.3)and 20 patients with tuberculosis T-SPOT false negative/low value(TBAg/PHA<0.3).Fresh peripheral blood was collected and cell functions and expression of TIM-3 and PD-1of CD4+,CD8+ and NK were detected by flow cytometry.TIM-3 and PD-1 blocking antibodies were added to tuberculosis T-SPOT false negative/low value PBMCs to observe the effect of blocking signal pathway on T-SPOT results.[Results] Non-specific CD4,CD8,and NK cell functions were significantly lower in tuberculosis patients than in healthy controls,and T-SPOT high-response patients did not differ from T-SPOT low-response patients.However,tuberculosis-specific CD4+,CD8+,and NK cells in T-SPOT hyperresponsive patients are more potent than T-SPOT false-negative/low-value patients.The expression of CD8+PD-1+,NK PD-1+ and NK TIM-3+ in the T-SPOT false negative/low value group was significantly higher than that in the T-SPOT hyperresponsive patient.Combination blocking of TIM-3 and PD-1 signaling pathways partially can increase IFN-? secretion levels and T-SPOT spot counts.[Conclusion] Overexpression of TIM-3 and PD-1 can lead to T-SPOT false negative/low value,blocking TIM-3 and PD-1 signaling pathways can partially correct T-SPOT false negatives and low ratios to improve T-SPOT accuracy.