p~(38)MAPK在IL-18诱导肾小管上皮细胞转分化中的作用

目的:白细胞介素18(IL-18)可诱导肾小管上皮细胞转分化,本研究探讨其是否是通过p38MAPK途径而起作用。方法:应用不同浓度的p38MAPK通路特异性阻断剂SB203580(0、5、10、20μmol/L)预孵育人近端肾小管上皮细胞(HK-2细胞)30min后,加入IL-18(100ng/ml)共培养24、48、72h。应用RT-PCR法检测α-平滑肌肌动蛋白(α-SMA)mRNA的表达水平;应用ELISA法测定细胞浆中α-SMA蛋白质含量。结果:SB203580呈剂量依赖性地抑制IL-18诱导的HK-2细胞α-SMA基因表达(P0.05)。结论:p38MAPK通路是调控IL-18诱导肾小管上皮细胞转分化的主要信号通路之一。     

Prostaglandin E2 induces MUC8 gene expression via a mechanism involving ERK MAPK/RSK1/cAMP response element binding protein activation in human airway epithelial cells

MUC8 gene expression is overexpressed in nasal polyp epithelium and is also increased by treatment with inflammatory mediators in nasal epithelial cells. These data suggest that MUC8 may be one of important mucin genes expressed in human airway. However, the mechanisms of various inflammatory mediator-induced MUC8 gene expression in normal nasal epithelial cells remain unclear. We examined the mechanism by which prostaglandin E(2) (PGE2), an arachidonic acid metabolite, increases MUC8 gene expression levels. Here, we show that ERK mitogen-activated protein kinase is essential for PGE2-induced MUC8 gene expression in normal human nasal epithelial cells and that p90 ribosomal S 6 protein kinase 1 (RSK1) mediates the PGE2-induced phosphorylation of cAMP-response element binding protein. Our results also indicate that cAMP-response element at the -803 region of the MUC8 promoter is an important site of PGE2-induced MUC8 gene expression. In conclusion, this study gives insights into the molecular mechanism of PGE2-induced MUC8 gene expression in human airway epithelial cells.     

Japanese encephalitis virus down-regulates thioredoxin and induces ROS-mediated ASK1-ERK/p38 MAPK activation in human promonocyte cells

Japanese encephalitis virus (JEV), a mosquito-borne flavivirus, causes severe neurological disease with high mortality. Molecular mechanisms of JEV pathogenesis such as upstream apoptotic processes and pathways are not yet completely resolved or understood. In this study, JEV replication in human promonocyte cells induced time-dependent apoptosis and activated virus dose-dependent caspases 3, 8 and 9. Proteomic analysis demonstrated up- and down-regulated (more or less than 1.5-fold) proteins in JEV-infected promonocyte cells. Biological process categorization showed processes of antioxidation, free radical removal, and sulfur redox metabolism entailed many identified up- and down-regulated proteins. Down-regulation of thioredoxin, confirmed by using Western blotting, was involved in the apoptosis process of the oxidative stress response pathway. JEV infection caused increased intracellular ROS production and activation of ASK1-ERK/p38 MAPK signaling. ERK/p38 MAPK inhibitor PD98059 treatment definitely suppressed this apoptosis. Down-regulation of thioredoxin, increased intracellular ROS, and activation of ASK1-ERK/p38 MAPK signaling all were associated with JEV-induced apoptosis. These results are suggestive of an oxidative stress-pathway as a key element of JE pathogenesis.     

Interleukin-13 induces thymus and activation-regulated chemokine (CCL17) in human peripheral blood mononuclear cells

BACKGROUND: In allergic inflammation involving allergic rhinitis, the predominance of Th(2) lymphocytes is one of the primary causal agents in promotion of the allergic condition. Thymus and activation-regulated chemokine (TARC/CCL17) is a recently identified chemokine that induces the development of Th(2) lymphocytes. One of the sources of TARC has been reported to be peripheral blood mononuclear cells (PBMCs). OBJECTIVE: We investigated TARC production from PBMCs by the stimulation of specific antigens and Th(2) type cytokines. METHOD: PBMCs were isolated from both allergic rhinitis patients and healthy volunteers. PBMCs were incubated with cytokine. TARC mRNA expression was examined by real time PCR methods and the amount of TARC production was examined by ELISA. RESULTS: IL-13 was found to be the most potent inducer for TARC mRNA expression and protein production in PBMCs. Furthermore, tumour necrosis factor alpha and IL-13 synergistically induce TARC. The amount of TARC from allergic rhinitis patients was significantly larger than that from healthy volunteers. Moreover, TARC was induced by a specific antigen, and was 35% inhibited by an anti-IL-13 neutralizing antibody. CONCLUSION: These results indicate that IL-13 is important in TARC mediated Th(2) lymphocytes infiltration in the nasal mucosa.     

Interleukin-1beta induces chronic activation and de novo synthesis of neutral ceramidase in renal mesangial cells.

The lipid signaling molecule ceramide is formed by the action of acid and neutral sphingomyelinases and degraded by acid and neutral ceramidases. Short-term stimulation of mesangial cells with the pro-inflammatory cytokine interleukin-1beta (IL-1beta) leads to a rapid and transient increase in neutral sphingomyelinase activity (Kaszkin, M., Huwiler, A., Scholz, K., van den Bosch, H., and Pfeilschifter, J. (1998) FEBS Lett. 440, 163-166). In this study, we report on a second delayed peak of activation occurring after hours of IL-1beta treatment. This second phase of activation was first detectable after 2 h of treatment and steadily increased over the next 2 h, reaching maximal values after 4 h. In parallel, a pronounced increase in neutral ceramidase activity was observed, accounting for a constant or even decreased level of ceramide after long-term IL-1beta treatment, despite continuous sphingomyelinase activation. The increase in neutral ceramidase activity was due to expressional up-regulation, as detected by an increase in mRNA levels and enhanced de novo protein synthesis. The increase in neutral ceramidase protein levels and activity could be blocked dose- dependently by the p38 MAPK inhibitor SB 202190, whereas the classical MAPK pathway inhibitor U0126 and the protein kinase C inhibitor Ro 318220 were ineffective. Moreover, cotreatment of cells for 24 h with IL-1beta and SB 202190 led to an increase in ceramide formation. Interestingly, IL-1beta-stimulated neutral ceramidase activation was not reduced in mesangial cells isolated from mice deficient in MAPK-activated protein kinase-2, which is a downstream substrate of p38 MAPK, thus suggesting that the p38 MAPK-mediated induction of neutral ceramidase occurs independently of the MAPK-activated protein kinase-2 pathway. In summary, our results suggest a biphasic regulation of sphingomyelin hydrolysis in cytokine-treated mesangial cells with delayed de novo synthesis of neutral ceramidase counteracting sphingomyelinase activity and apoptosis. Neutral ceramidase may thus represent a novel cytoprotective enzyme for mesangial cells exposed to inflammatory stress conditions.     

Interleukin-4 induces specific pp-GalNAc-T expression and alterations in mucin O-glycosylation in colonic epithelial cells

Mucus hypersecretion occurs as a consequence of the Th2 immune response in epithelia, yet it was not previously known whether the degree of O-glycosylation was modulated under such conditions. A colonic carcinoma cell line LS174T was used to assess the effect of interleukin (IL)-4 on the mRNA levels of eight pp-GalNAc-Ts. A three- to four-fold increase in pp-GalNAc-T1, T4, and T7 levels was observed. Lysates of untreated or IL-4-treated cells were examined for their ability to transfer GalNAc residues onto a peptide corresponding to the tandem repeat portion of human MUC2. The number of incorporated GalNAc residues was greater after incubation with lysates of IL-4-treated cells than with lysates of untreated cells. Mucin-like large glycoproteins secreted by IL-4-treated cells had higher binding capacity to PNA and VVA-B₄ than those secreted by untreated cells. The results indicated that IL-4-treated LS174T cells are able to produce mucins with a higher degree of O-glycosylation than untreated counterparts.     

17beta -Estradiol modulates mechanical strain-induced MAPK activation in mesangial cells.

Gender is an important determinant of clinical outcome across a broad spectrum of kidney diseases, but the mechanism(s) responsible for the protective effect of female gender have not been fully elucidated. Remnant kidney glomerular injury is limited in female rats compared with male rats despite similar elevations in glomerular capillary pressure. In vitro, mechanical strain leads to the activation of p44/42 mitogen-activated kinase (p44/42 MAPK) and Jun N-terminal kinase/stress-activated protein kinase (SAPK) in glomerular mesangial cells (MC). Accordingly, we studied the effect of 17beta-estradiol on mechanical strain-induced signal transduction in MC. Exposure of MC to mechanical strain increased p44/42 MAPK activation (3-fold) and SAPK activation (2.5-fold), and kinase activation was inhibited by pretreatment with 17beta-estradiol (10(minus sign8) to 10(minus sign11) m) for 24 h in a dose-dependent manner. Mechanical strain-induced nuclear translocation of p44/42 MAPK and SAPK and nuclear protein binding to AP-1 were also attenuated by 17beta-estradiol. The inhibitory effects of 17beta-estradiol were not reproduced by the cell-impermeable estrogen, BSA/17beta-estradiol, nor did preincubation with 17beta-estradiol lead to actin cytoskeleton disassembly or impaired stress fiber formation. However, 17beta-estradiol did increase base-line levels of the dual specificity phosphatase MKP-1. The inhibitory effects of 17beta-estradiol on p44/42 MAPK activation and SAPK activation, translocation, and AP-1 binding were all abrogated by the estrogen receptor antagonist, ICI-182,780. We conclude that attenuation of mechanical strain-induced MAPK activation by 17beta-estradiol is dependent on intracellular estrogen receptor. The attenuation of stretch-induced kinase activation may be due, at least in part, to an effect of 17beta-estradiol on MKP-1 expression. Together, these findings add insight into the protective effect of gender on renal disease progression.     

Carbon monoxide inhibits IL-17-induced IL-6 production through the MAPK pathway in human pulmonary epithelial cells

Interleukin (IL)-17 is a proinflammatory cytokine that is produced by activated memory CD4 T cells, which regulates pulmonary neutrophil emigration by the induction of CXC chemokines and cytokines. IL-17 constitutes a potential target for pharmacotherapy against exaggerated neutrophil recruitment in airway diseases. As a cytoprotective and anti-inflammatory gaseous molecule, carbon monoxide (CO) may also regulate IL-17-induced inflammatory responses in pulmonary cells. Herein, we examine the production of cytokine IL-6 induced by IL-17 and the effect of CO on IL-17-induced IL-6 production in human pulmonary epithelial cell A549. We first show that IL-17 can induce A549 cells to release IL-6 and that CO can markedly inhibit IL-17-induced IL-6 production. IL-17 activated the ERK1/2 MAPK pathway but did not affect p38 and JNK MAPK pathways. CO exposure selectively attenuated IL-17-induced ERK1/ERK2 MAPK activation without significantly affecting either JNK or p38 MAPK activation. Furthermore, in the presence of U0126 and PD-98059, selective inhibitors of MEK1/2, IL-17-induced IL-6 production was significantly attenuated. We conclude that CO inhibits IL-17-stimulated inflammatory response via the ERK1/2-dependent pathway.     

Methylglyoxal induces apoptosis through activation of p38 MAPK in rat Schwann cells

The formation of glucose-derived methylglyoxal (MG), a highly reactive dicarbonyl compound, is accelerated under diabetic conditions. We examined whether MG was capable of inducing apoptosis in Schwann cells (SCs), since recent studies have suggested a potential involvement of apoptotic cell death in the development of diabetic neuropathy. MG induced apoptosis in SCs in a dose-dependent manner, accompanied by a reduction of intracellular glutathione content and activation of the p38 MAPK. Inhibiting the p38 MAPK activation by SB203580 successfully suppressed the MG-induced apoptosis in SCs. Aminoguanidine and N-acetyl-l-cysteine also inhibited the MG-induced p38 MAPK activation and apoptosis along with restoration of the intracellular glutathione content. These results suggest a potential role for MG in SC injury through oxidative stress-mediated p38 MAPK activation under diabetic conditions, and it may serve as a novel insight into therapeutic strategies for diabetic neuropathy.     

Fluvastatin inhibits angiotensin II-induced nuclear factor kappa B activation in renal tubular epithelial cells through the p38 MAPK pathway

3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors has been shown to reduce the progression of renal disease independent of cholesterol-lowering effect, but the mechanism of potential protective effect remains unclear. Here, we investigate the effect of fluvastatin on activation of nuclear factor-κB (NF-κB) induced by angiotensin II (AngII) in rat kidney tubule epithelial cells (NRK-52E). Electrophoretic mobility shift assays (EMSA) was used to detect NF-κB activation. Phosphorylation of cellular p38 mitogen-activated protein kinase (p38MAPK) was determined by western blot analysis. AngII stimulated the DNA-binding activity of NF-κB and phosphorylation of p38MAPK in cultured NRK-52E cells in a dose-dependent (10⁻⁹–10⁻⁶ mol/l) manner (P < 0.01). AngII (10⁻⁶ mol/l) induced a rapid (5 min) increase of the p38MAPK phosphorylation. NF-κB DNA-binding activity was increased at as early as 30 min, peaked at 2 h after AngII treatment. This stimulatory effect of AngII on NF-κB was blocked by SB203580 (a specific inhibitor of p38MAPK). Incubation of cells with fluvastatin significantly inhibited the AngII-induced NF-κB activation in a dose-dependent (10⁻⁷–10⁻⁵ mol/l) manner (P < 0.05). Exogenous mevalonate (10⁻⁴mol/l) prevented the effect of fluvastatin on NF-κB activation. These results suggest the fluvastatin reduced AngII-induced NF-κB activation via the p38MAPK pathway in NRK-52E cells. The effect is at least partly due to blocking the biosynthesis of mevalonate.     

Asbestos exposure induces alveolar epithelial cell plasticity through MAPK/Erk signaling

The inhalation of asbestos fibers is considered to be highly harmful, and lead to fibrotic and/or malignant disease. Epithelial-to-mesenchymal transition (EMT) is a common pathogenic mechanism in asbestos associated fibrotic (asbestosis) and malignant lung diseases. The characterization of molecular pathways contributing to EMT may provide new possibilities for prognostic and therapeutic applications. The role of asbestos as an inducer of EMT has not been previously characterized. We exposed cultured human lung epithelial cells to crocidolite asbestos and analyzed alterations in the expression of epithelial and mesenchymal marker proteins and cell morphology. Asbestos was found to induce downregulation of E-cadherin protein levels in A549 lung carcinoma cells in 2-dimensional (2D) and 3D cultures. Similar findings were made in primary small airway epithelial cells cultured in 3D conditions where the cells retained alveolar type II cell phenotype. A549 cells also exhibited loss of cell-cell contacts, actin reorganization and expression of α-smooth muscle actin (α-SMA) in 2D cultures. These phenotypic changes were not associated with increased transforming growth factor (TGF)-β signaling activity. MAPK/Erk signaling pathway was found to mediate asbestos-induced downregulation of E-cadherin and alterations in cell morphology. Our results suggest that asbestos can induce epithelial plasticity, which can be interfered by blocking the MAPK/Erk kinase activity.