鼠尾草酚通过调控TAK1活性影响慢性阻塞性肺疾病小鼠的炎症反应OA

Objective:To investigate the effects of carnosol on pulmonary inflammation and fibrosis in a murine model of chronic obstructive pulmonary disease(COPD)and to elucidate whether these effects are mediated through the regulation of TAK1 activity.Methods:30 male C57BL/6 mice were randomly allocated into five groups:normal control,COPD model,low-dose carnosol(2.5 mg/kg),high-dose carnosol(5 mg/kg),and dexamethasone positive control(1 mg/kg),with 6 mice in each group.The COPD model was established through intratracheal lipopolysaccharide(LPS)instillation combined with cigarette smoke exposure and intraperitoneal injection of cigarette smoke extract(CSE).Modeling was accompanied by drug intervention,which was administered over a consecutive 7-day period.24 hours following the final administration,serum,bronchoalveolar lavage fluid(BALF),and lung tissues were collected.Hematoxylin-eosin(HE)staining was performed to evaluate pulmonary histopathological changes,while Masson's trichrome staining was utilized to assess collagen deposition.Serum and BALF levels of tumor necrosis factor-α(TNF-α)and interleukin-1β(IL-1β)were quantified by enzyme-linked immunosorbent assay(ELISA).Western blot analysis was conducted to determine the protein expression levels of phosphorylated TAK1(p-TAK1),phosphorylated P38(p-P38),phosphorylated P65(p-P65),phosphorylated c-Jun N-terminal kinase(p-JNK),and inhibitor of NF-κB α(IκBα)in lung tissues.Real-time quantitative polymerase chain reaction(RT-qPCR)was employed to measure TAK1 mRNA expression.Results:Compared with the normal control group,the COPD model group exhibited severe pulmonary histopathological damage characterized by massive inflammatory cell infiltration and substantial collagen fiber deposition,and an increase in mean linear intercept(MLI)(P<0.001).Serum and BALF levels of TNF-α and IL-1β were elevated(P<0.001).Furthermore,the protein expression levels of p-TAK1,p-P38,p-P65,and p-JNK,along with the TAK1 mRNA expression in lung tissues,were upregulated(P<0.001).Meanwhile,the IκBα protein expression was down-regulated(P<0.001).Compared with the model group,low-dose carnosol attenuated histopathological injury in lung tissue,suppressed inflammatory cell infiltration,and mitigated collagen deposition(P<0.05),the MLI was reduced(P<0.05).No significant differences were observed in the concentrations of TNF-α and IL-1β in serum or BALF(P>0.05).At the protein level,p-TAK1 and p-P38 was downregulated(P<0.01),whereas IκBα expression was upregulated(P<0.01).In contrast,no statistically significant changes were detected in the p-JNK or p-P65(P>0.05),the mRNA expression level of TAK1 was down-regulated(P<0.01).In the high-dose carnosol group,histopathological lung injury,inflammatory cell infiltration,and collagen deposition were further attenuated(P<0.001),and MLI was further decreased(P<0.001).TNF-α and IL-1β levels in both serum and BALF were lowered(P<0.05).The levels of p-TAK1,p-JNK,p-P38,and p-P65 were all downregulated(P<0.01),IκBα protein expression was further enhanced(P<0.001),and TAK1 mRNA expression was markedly suppressed(P<0.001).Relative to the low-dose carnosol group,the high-dose group exhibited significantly milder histopathological alterations,less pronounced inflammatory infiltration,and reduced collagen accumulation(P<0.01),along with a shorter MLI(P<0.05).No intergroup differences were observed in serum or BALF TNF-α and IL-1β levels(P>0.05).The levels of p-TAK1,p-P38 and p-P65 were further diminished(P<0.05),there was no difference in the expression levels of p-JNK and total IκBα protein(P>0.05).TAK1 mRNA expression was also lower in the high-dose group(P<0.001).The dexamethasone group demonstrated therapeutic efficacy comparable to that of the high-dose carnosol group across all major endpoints—including attenuation of lung histopathology,suppression of inflammation and fibrosis,reduction of systemic and local pro-inflammatory cytokines,and inhibition of TAK1 activation and downstream NF-κB/MAPK signaling.There was no statistically significant difference in the levels of MLI and TAK1 mRNA between the two groups(P>0.05).Conclusion:Carnosol dose-dependently attenuates pulmonary inflammation and fibrosis in COPD mice by inhibiting TAK1 transcription and phosphorylation,consequently blocking the downstream NF-κB/MAPK signaling cascade and ameliorating pulmonary histopathological damage.