Role of Shikonin modulation of TLRs signalling pathway on lung inflammation,immune function and inflammatory factors in Mycobacterium tuberculosis-infected mice

LI Hongmei; LAI Min; LIU Hua; TAN Hui

doi:10.3969/j.issn.1007-8134.2024.02.009

Abstract：Objective To analyse the effects and mechanism of action of Shikonin on pulmonary inflammation,immune function and inflammatory factors in Mycobacterium tuberculosis-infected mice. Methods Eighty-seven SPF C57BL/6 mice were randomly selected,and 15 mice were randomly selected as the control group. The remaining 72 mice were established with Mycobacterium tuberculosis infection,and 45 mice were successfully established,with a success rate of 62.50％. They were randomly divided into model group,shikonin group,shikonin+Toll-like receptor 4 (Toll-like receptor 4,TLR4),Myeloid differentiation factor 88 (MyD 88),nuclear transcription factor κB (nuclear transcription factor κB,NF-κB,NF-κB) P65 signal pathway agonist group (shikonin+LPS group) with 15 rats in each group,and shikonin group was intraperitoneally injected with 40 mg/kg shikonin. In shikonin+LPS group,40 mg/kg shikonin and 0.25 mg/kg LPS;were injected intraperitoneally;The control group and model group were injected with the same amount of normal saline intraperitoneally. After the last administration,the lung weight index,the amount of tuberculosis bacteria in lung tissue,the pathological changes of lung tissue,the indexes of immune function,the expression levels of inflammatory factors and pathway-related factors were observed. Results Compared with the control group,the lung weight index,lung lesion index and the amount of tuberculosis bacteria in lung tissue in the model group were significantly increased (P<0.05). Compared with the model group,the lung weight index,lung lesion index and pulmonary tuberculosis load in shikonin group were significantly reduced (P<0.05). Compared with the shikonin group,the lung weight index,lung lesion index and pulmonary tuberculosis load in the shikonin+LPS group were significantly increased (P<0.05). In the control group,the lung tissue structure is normal,the cells are closely arranged,the alveolar structure is complete,and no inflammatory cell infiltration is found. In the model group,the lung tissue structure was seriously damaged,the number of alveoli decreased,a large number of inflammatory cells infiltrated,and tuberculosis nodules were formed. In shikonin group,the lung tissue structure was basically complete,and inflammatory cell infiltration was occasionally seen. The lung tissue structure,cell arrangement,inflammatory cell infiltration and tuberculosis nodules in shikonin+LPS group are similar to those in model group,but better than those in model group. Compared with the control group,the levels of CD3+T lymphocytes,CD4+T lymphocytes,CD4+/CD8+and interleukin-10 (IL-10) in the model group were significantly decreased,while CD8+T lymphocytes and interleukin-6 (IL-6),Tumor necrosis factor-α (TNF-α),TLR4,MyD88 and NF-κB p65 all increased significantly (P<0.05). Compared with the model group,the expression levels of CD3+T lymphocytes,CD4+T lymphocytes,CD4+/CD8+and IL-10 in shikonin group were significantly increased,while the expression levels of CD8+T lymphocytes,IL-6,TNF-α,TLR4,MyD88 and NF-κB p65 were significantly decreased (P<0.05). Compared with shikonin group,the expression levels of CD3+T lymphocytes,CD4+T lymphocytes,CD4+/CD8+and IL-10 in shikonin+LPS group were significantly decreased,while the expression levels of CD8+T lymphocytes,IL-6,TNF-α,TLR4,MyD88 and NF-κB p65 were significantly increased (P<0.05). Conclusion Comfreyin may play a certain lung-protective role by inhibiting the TLR4/MyD88/NF-κB p65 signalling pathway to improve Mycobacterium tuberculosis charge in the lung tissues of Mycobacterium tuberculosis-infected mice,suppressing inflammatory responses in the lungs,and improving immune function.