Objective To investigate whether p38 mitogen activated protein kinase (p38MAPK) inhibitor can reduce acute lung injury (ALI) caused by lipopolysaccharide (LPS) by regulating Th17/Treg balance. Methods Balb/c mice were randomly divided into a control group, an ALI group and an intervention group. The mice in the control group were injected with phosphate-buffered saline, the mice in the ALI group were intraperitoneally injected with 40 mg/kg LPS, and the mice in the intervention group were injected with SB203580 (0.5 mg/kg, 1 mg/kg, 2 mg/kg, 5 mg/kg) intraperitoneally 1 h prior to the intraperitoneal injection of LPS. All mice were killed on 12 h later respectively. Hematoxylin-eosinstin staining was used to observe the pathological changes of lung tissue, and cell classification, counting, and total protein levels in bronchoalveolar lavage fluid (BALF) were detected. Transcript expression of forkhead box p3 (Foxp3) and retinoic acid receptor-related orphan receptor-γt (RORγt) was detected by real-time polymerase chain reaction. Interleukin (IL)-6, IL-10, IL-17, IL-23 and transforming growth factor-β (TGF-β) in lung tissue and IL-6, tumor necrosis factor-α (TNF-α) in serum were measured by enzyme-linked immunosorbent assay. The Th17 and Treg subset distribution in spleen was determined by flow cytometry. Results Histopathological examination showed that LPS induced inflammatory cell infiltration in lung tissue, increased cell count and protein levels in BALF (P<0.05), and increased proportion of neutrophils and monocytes in the ALI mice. SB203580 significantly attenuated tissue injury of the lungs in LPS-induced ALI mice. Serum levels of IL-6 and TNF-α in the ALI group were significantly higher than those in the control group, and inflammatory cytokines were decreased after SB203580 intervention. Compared with the ALI group, the production of inflammatory cytokines associate with Th17, including IL-17, IL-23, RORγt was inhibited, and the production of cytokines associate with Treg, such as IL-10 and Foxp3 in lung tissue was increased in the intervention group in a concentration-dependent manner with SB203580. After SB203580 intervention, Th17/Treg ratio was significantly decreased compared with the LPS group (P<0.05). Conclusion p38MAPK inhibitor can reduce LPS-induced ALI by regulating the imbalance of Treg cells and Th17 cells.
Objective To investigate changes of TLR2 mRNA expression in lung of a mouse model of Chlamydia Pneumoniae pneumonitis, and to explore the possible mechanism of signal transduction. Methods Ninety-six male C3H/HeJ mice were randomly divided into four groups as follows: a control group, a model group, a SB203580 intervened group, and a pyrrolidine dithiocarbamate( PDTC) intervened group. Chlamydia Pneumoniae pneumonitis was induced by intranasally inoculated with 4. 0 ×106 IFU/mL of C. Pneumoniae per mouse in the model group and two intervened groups. Then the intervened groups were intraperitoneally injected with the p38MAPK inhibitor SB203580 and nuclear factor kappa B ( NF-κB)inhibitor PDTC, respectively. Six mice in each group were randomly killed in 1st, 4th, 7th and 14th day. The expressional changes of TLR2 mRNA in the mice lung tissue were measured by semi-quantitative RT-PCR. The concentrations of TNF-α in the lung homogenate were measured by ELISA. Results TLR2 mRNA expression in the lung tissue significantly increased after C. Pneumoniae infection, peaking at 4th and 7th days, then decreased after 14th day. Tumor necrosis factor-α( TNF-α) was also elevated in the lung tissue after C. Pneumoniae challenging. Both SB203580 and PDTC treatment effectively inhibited TNF-αand TLR2 mRNA expressions in lung. The inhibitory effect was more obvious by SB203580 treatment. Conclusion C. Pneumoniae can upregulate the expressions of TLR2 and TNF-α in lung, and TLR2/MAPK and TLR2 /NF-κB signal pathways may be involved in Chlamydia Pneumoniae pneumonitis.
Objective To investigate whether Chlamydia pneumoniae alters the expression of TLR2 mRNA and p38 MAPK mRNA in mice with Chlamydia pneumoniae infection in TLR2-p38 MAPK-dependent pathway, subsequently leading to the release of cytokines. Methods Seventy-two male C3H/HeJmice were randomly divided into three groups as follow: a normal control group, a C. pneumoniae-inoculated group, and a C. pneumoniae-inoculated with SB203580 treatment group. The mice in the three groups were sacrificed on 1st, 4th, 7th, 14th day separately, and lung tissues were sampled for measurement. The expression changes of TLR2 mRNA and p38 MAPK mRNA in the mice lung tissue were measured by semi-quantitative RT-PCR. The concentrations of TNF-αin the lung tissue were measured by ELISA.Results Compared with those in the normal group, the expressions of TLR2 mRNA and p38MAPK mRNA in the lung tissue increased quickly after C. pneumoniae infection, which was especially obvious on day 4 and on day 7, the expression level of TLR2 mRNA on day 7 was markedly higher than that of the normal group [ ( 7. 24 ±1. 78) mg/L vs.( 0. 64 ±0. 14) mg/L, P lt;0. 05] ; The expression level of p38 MAPK mRNA on day 4 was markedly higher than that of the normal group [ ( 9. 267 ±1. 813) mg/L vs. ( 3. 734 ±0. 946) mg/L, P lt;0. 05] . After 14 days, C. pneumoniae infection of mice was attenuated, the concentration of TNF-α in the lung tissue increased, and was clearly higher than that of the normal control group, peaking on day 4 [ ( 77. 29 ±9. 66) pg/mg] . Treatment with SB203580 could effectively inhibit TLR2 mRNA and p38 MAPK mRNA expression in lung, which was especially obvious on day 4 and on day 7. The expression level of TLR2 mRNA on day 7 was ( 0. 269 ±0. 09) mg/L, and the expression level of p38 MAPK mRNA on day 7 [ ( 0. 002 ±0. 001) mg/L] was even more obviously attenuated, the concentration of TNF-α in the lung tissue markedly decreased when compared with that in the infected group, and its concentration on day 4 [ ( 25. 76 ±3. 49) pg/mg] lowered more clearly. Conclusions The alteration of TLR2-p38 MAPKdependent signal pathway in lungs is closely connected with Chlamydia pneumoniae infection. SB203580 treatment can effectively controll the elevation of TLR2 mRNA and p38 MAPK mRNA expressions in lung. It can effectively control the TLR2-MAPK signal transduction pathway.
Abstract： Objective To investigate the acute cardioprotective effect of 17b-estradiol (17b-E2) against severe myocardial ischemia/reperfusion (I/R) injury in rabbits and the mechanism of the effect. Methods We established the model of myocardial I/R in vivo by occluding the left anterior descending coronary artery of the rabbits (who underwent coronary occlusion for 40 minutes followed by 3 hours of reperfusion). Twentyfour New Zealand white male rabbits were randomly divided into two groups with 12 in each group. Before coronary occlusion, 1 ml of ethanol or 17b-E2 at 10 μg/kg was administered intravenously to the rabbits in the control group and the experimental group respectively. The serum levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were measured by enzymelinked immunosorbent assay (ELISA) at the following time points: before occlusion, 40 minutes after occlusion, 1 hour, 2 hours and 3 hours after reperfusion. Activation of p38 mitogen activated protein kinase(MAPK) was determined by Western blotting analysis, and apoptosis of cardiocytes was identified by terminal deoxynucleotidlyl transferase mediated deoxyuridinebiotin dUTP Nick End Labeline (TdT)mediated dNTP nick end labeling (TUNEL) staining. Results During myocardial ischemia, TNF-α decreased significantly in the experimental group compared with the control group (F=0.007,P=0.001), while there was no difference in IL-6 between the two groups (F=0.616,P=0.095). During the process of reperfusion, the levels of TNF-α and IL-6 in the experimental group were significantly lower than those in the control group (Plt;0.01). Besides, the activation of p38 MAPK and apoptotic index for the experimental group were also lower (45.07%±2.73% vs. 61.25%±2.41%, t＝－15.398, P=0.000; 11.21%±3.85% vs. 22.02%±4.49%, t＝－6.332, P=0.000). Conclusion The cardioprotective effect of 17b-E2 against myocardial I/R may be attributed to its antiinflammatory and antiapoptotic properties, which is probably associated with the inhibition of 17bE2 on p38MAPK activity.
目的:探讨表没食子儿茶素没食子酸酯(EGCG)对乳腺癌细胞MCF7生长的影响及对乳腺癌细胞MDAMB231迁移的影响。方法:MCF7细胞培养贴壁之后,加入EGCG处理,2d后收集蛋白,采用Western Blot检测磷酸化p38丝裂原活化蛋白激酶(phosphop38MAPK)的表达;同样处理后收集活细胞,用细胞计数法检测细胞的存活;取对数生长期的MDAMB231细胞,分至6孔板培养,使用EGCG处理后,采用细胞划线法探测乳腺癌细胞的迁移。结果:使用EGCG处理乳腺癌细胞后,phosphop38MAPK的表达降低,EGCG处理乳腺癌细胞4d后其增殖率降低50%,迁移活性降低。结论:EGCG处理乳腺癌细胞能抑制肿瘤细胞的生长以及迁移,这与p38MAPK信号通路相关。