The role of p38 protein kinase in mouse responses to low-intensity electromagnetic radiation of the centimeter range

The role of p38 mitogen-activated protein kinase in regulating the cell responses to ultralow-intensity centimeter microwaves (8.15–18.0 GHz, 1 µW/cm², 1 h) was studied in male BalbC mice. Mice were tested for the level of protein phosphorylation in the NF-κB (p65 and IKK), JNK, and IRF3 signaling cascades expression of TLR4 and stress-inducible heat shock proteins Hsp72 and Hsp90 in splenic lymphocytes and pro- and anti-inflammatory cytokines and IL-10 in the blood serum. An inhibitor of the p38 signaling pathway (p38 inhibitor XI) was shown to reduce the sensitivity to ultralow-intensity ultrahigh-frequency electromagnetic radiation. This was evident from a decrease in radiation-induced activation of the NF-κB signaling pathway, expression of Hsp72 and Hsp90 in splenic cells, and an accumulation of proinflammatory cytokines (IL-6, IL-17, TNF-α, and IFN-γ) in the blood serum of irradiated mice pretreated with p38 inhibitor XI. However, p38 inhibitor XI did not attenuate the activation of the p38 and IRF3 signaling pathways and overexpression of TLR4 in splenic cells of irradiated mice. It was assumed that p38 mitogenactivated protein kinase is involved in regulating the nonspecific defense responses does not determine the murine sensitivity to ultralow-intensity electromagnetic waves of the centimeter range.     

The Role of p38 and CK2 Protein Kinases in the Response of RAW 264.7 Macrophages to Lipopolysaccharide

The role of protein kinases p38 and CK2 (casein kinase II) in the response of RAW 264.7 macrophages to the lipopolysaccharide (LPS) from gram-negative bacteria was studied. Using specific p38 and CK2 inhibitors (p38 MAP kinase Inhibitor XI and casein kinase II Inhibitor III, respectively), we investigated the effects of these protein kinases on (i) LPS-induced activation of signaling pathways involving nuclear factor κB (NF-κB), stress-activated protein kinase/c-Jun N-terminal kinase (SAPK/JNK), p38, and interferon regulatory factor 3 (IRF3); (ii) expression of Toll-like receptor 4 (TLR4) and inducible heat-shock proteins HSP72 and HSP90; and (iii) production of interleukins IL-1α, IL-1β, IL-6, tumor necrosis factor α, and IL-10. Activation of the proapoptotic signaling in the macrophages was evaluated from the ratio between the active and inactive caspase-3 forms and p53 phosphorylation. Six hours after LPS addition (2.5 μg/ml) to RAW 264.7 cells, activation of the TLR4 signaling pathways was observed that was accompanied by a significant increase in phosphorylation of IκB kinase α/β, NF-κB (at both Ser536 and Ser276), p38, JNK, and IRF3. Other effects of macrophage incubation with LPS were an increase in the contents of TLR4, inducible heat-shock proteins (HSPs), and pro- and anti-inflammatory cytokines, as well as slight activation of the pro-apoptotic signaling in the cells. Using inhibitor analysis, we found that during the early response of macrophages to the LPS, both CK2 and p38 modulate activation of MAP kinase and NF-κB signaling pathways and p65 phosphorylation at Ser276/Ser536 and cause accumulation of HSP72, HSP90 and the LPS-recognizing receptor TLR4. Suppression of the p38 MAP kinase and CK2 activities by specific inhibitors (Inhibitor XI and Inhibitor III, respectively) resulted in the impairment of the macrophage effector function manifested as a decrease in the production of the early-response proinflammatory cytokines and disbalance between the pro- and anti-apoptotic signaling pathways leading presumably to apoptosis development. Taken together, our data indicate the inefficiency of therapeutic application of p38 and CK2 inhibitors during the early stages of inflammatory response.     

Role of mitogen-activated protein kinases and tyrosine kinases on IL-1-Induced NF-kappaB activation and iNOS expression in bovine articular chondrocytes.

Nitric oxide (NO), produced by the inducible isoform of the NO synthase (iNOS), plays an important role in the pathophysiology of arthritic diseases. This work aimed at elucidating the role of the mitogen-activated protein kinases (MAPK), p38MAPK and p42/44MAPK, and of protein tyrosine kinases (PTK) on interleukin-1beta (IL-1)-induced iNOS expression in bovine articular chondrocytes. The specific inhibitor of the p38MAPK, SB 203580, effectively inhibited IL-1-induced iNOS mRNA and protein synthesis, as well as NO production, while the specific inhibitor of the p42/44MAPK, PD 98059, had no effect. These responses to IL-1 were also inhibited by treatment of the cells with the tyrosine kinase inhibitors, genistein and tyrphostin B42, which also prevented IL-1-induced NF-kappaB activation. The p38MAPK inhibitor, SB 203580, had no effect on IL-1-induced NF-kappaB activation. Finally, the p42/44MAPK inhibitor, PD 98059, prevented IL-1-induced AP-1 activation in a concentration that did not inhibit iNOS expression. In conclusion, this study shows that (1) PTK are part of the signaling pathway that leads to IL-1-induced NF-kappaB activation and iNOS expression; (2) the p38MAPK cascade is required for IL-1-induced iNOS expression; (3) the p42/44MAPK and AP-1 are not involved in IL-1-induced iNOS expression; and (4) NF-kappaB and the p38MAPK lie on two distinct pathways that seem to be independently required for IL-1-induced iNOS expression. Hence, inhibition of any of these two signaling cascades is sufficient to prevent iNOS expression and the subsequent production of NO in articular chondrocytes.     

Heat shock protein hsp72 is a negative regulator of apoptosis signal-regulating kinase 1

Heat shock protein 72 (Hsp72) is thought to protect cells against cellular stress. The protective role of Hsp72 was investigated by determining the effect of this protein on the stress-activated protein kinase signaling pathways. Prior exposure of NIH 3T3 cells to mild heat shock (43 degrees C for 20 min) resulted in inhibition of H(2)O(2)-induced activation of apoptosis signal-regulating kinase 1 (ASK1). Overexpression of Hsp72 also inhibited H(2)O(2)-induced activation of ASK1 as well as that of downstream kinases in the p38 mitogen-activated protein kinase (MAPK) signaling cascade. Recombinant Hsp72 bound directly to ASK1 and inhibited ASK1 activity in vitro. Furthermore, coimmunoprecipitation analysis revealed a physical interaction between endogenous Hsp72 and ASK1 in NIH 3T3 cells exposed to mild heat shock. Hsp72 blocked both the homo-oligomerization of ASK1 and ASK1-dependent apoptosis. Hsp72 antisense oligonucleotides prevented the inhibitory effects of mild heat shock on H(2)O(2)-induced ASK1 activation and apoptosis. These observations suggest that Hsp72 functions as an endogenous inhibitor of ASK1.     

Down-regulation of cytokine-induced interleukin-8 requires inhibition of p38 mitogen-activated protein kinase (MAPK) via MAPK phosphatase 1-dependent and -independent mechanisms

Down-regulation of overabundant interleukin (IL)-8 present in cystic fibrosis (CF) airways could ease excessive neutrophil burden and its deleterious consequences for the lung. IL-8 production in airway epithelial cells, stimulated with e.g. inflammatory cytokines IL-1β and tumor necrosis factor (TNF)-α, is regulated by several signaling pathways including nuclear factor (NF)-κB and p38 mitogen-activated protein kinase (MAPK). We previously demonstrated that the anti-inflammatory drugs dexamethasone and ibuprofen suppress NF-κB; however, only dexamethasone down-regulates cytokine-induced IL-8, highlighting the importance of non-NF-κB mechanisms. Here, we tested the hypothesis that down-regulation of cytokine-induced IL-8 requires modulation of the MAPK phosphatase (MKP)-1/p38 MAPK/mRNA stability pathway. The effects of dexamethasone (5 nm) and ibuprofen (480 μm) on this pathway and IL-8 were studied in CF (CFTE29o-, CFBE41o-) and non-CF (1HAEo-) airway epithelial cells. We observed that dexamethasone, but not ibuprofen, destabilizes IL-8 mRNA and up-regulates MKP-1 mRNA. Further, siRNA silencing of MKP-1, via p38 MAPK, leads to IL-8 overproduction and diminishes the anti-IL-8 potential of dexamethasone. However, MKP-1 overexpression does not significantly alter IL-8 production. By contrast, direct inhibition of p38 MAPK (inhibitor SB203580) efficiently suppresses IL-8 with potency comparable with dexamethasone. Similar to dexamethasone, SB203580 decreases IL-8 mRNA stability. Dexamethasone does not affect p38 MAPK activation, which excludes its effects upstream of p38 MAPK. In conclusion, normal levels of MKP-1 are necessary for a full anti-IL-8 potential of pharmacological agents; however, efficient pharmacological down-regulation of cytokine-induced IL-8 also requires direct effects on p38 MAPK and mRNA stability independently of MKP-1.     

姜黄素对过度训练大鼠脾脏炎症反应的调控作用及其机制

目的:研究姜黄素调控Toll-样受体4(TLR4)-p38丝裂原活化蛋白激酶(p38 MAPK)/核因子κB(NF-κB)信号通路缓解过度训练大鼠脾脏炎症反应的作用及其机制.方法:7周龄SPF级雄性Wistar大鼠分为安静对照组(C组,n=12)、过度训练模型组(OM组,n=11)、姜黄素+模型组(COM组,n=14)...     

Proteomic identification of Bcl2-associated athanogene 2 as a novel MAPK-activated protein kinase 2 substrate

The p38 MAPK cascade is activated by various stresses or cytokines. Downstream of p38 MAPKs, there are diversification and extensive branching of signaling pathways. Fluorescent two-dimensional difference gel electrophoresis of phosphoprotein-enriched samples from HeLa cells in which p38 MAPK activity was either suppressed or activated enabled us to detect approximately 90 candidate spots for factors involved in p38-dependent pathways. Among these candidates, here we identified four proteins including Bcl-2-associated athanogene 2 (BAG2) by peptide mass fingerprintings. BAG family proteins are highly conserved throughout eukaryotes and regulate Hsc/Hsp70-mediated molecular chaperone activities and apoptosis. The results of two-dimensional immunoblots suggested that the phosphorylation of BAG2 was specifically controlled in a p38 MAPK-dependent manner. Furthermore, BAG2 was directly phosphorylated at serine 20 in vitro by MAPK-activated protein kinase 2 (MAPKAP kinase 2), which is known as a primary substrate of p38 MAPK and mediates several p38 MAPK-dependent processes. We confirmed that MAPKAP kinase 2 is also required for phosphorylation of BAG2 in vivo. Thus, p38 MAPK-MAPKAP kinase 2-BAG2 phosphorylation cascade may be a novel signaling pathway for response to extracellular stresses.     

Enterovirus 71 induces COX-2 expression via MAPKs,NF-κB,and AP-1 in SK–N–SH cells: Role of PGE2 in viral replication

The enterovirus 71 (EV71) causes severe neurological diseases that were mediated through cyclooxygenase-2 (COX-2) expression in brain. However, the mechanisms underlying EV71-initiated intracellular signaling pathways leading to COX-2 expression remain unknown in neurons. Here we report that exposure of SK–N–SH cells to EV71 increased COX-2 expression and PGE₂ generation in a time- and virus titer-dependent manner, revealed by Western blot, real-time PCR, and PGE₂ analyses. These EV71-induced responses were mediated through activation of p42/p44 MAPK, p38 MAPK, JNK, NF-κB, and AP-1, revealed by using selective pharmacological inhibitors or transfection with respective siRNAs. Consistently, EV71-stimulated translocation of NF-κB into the nucleus and degradation of IκBα in the cytosol was blocked by pretreatment with the selective inhibitors of MEK1/2 (U0126) and NF-κB (Bay11-7085), respectively, suggesting that MEK1/2-p42/p44 MAPK cascade linking to NF-κB was involved in COX-2 expression. In addition, EV71-induced AP-1 subunits (c-jun and c-fos mRNA) expression was also attenuated by pretreatment with a selective JNK inhibitor SP600125, suggesting that JNK cascade linking to AP-1 was involved in COX-2 expression induced by EV71. These findings suggested that up-regulation of COX-2 associated with the release of PGE₂ from EV71-infected SK–N–SH cells which was mediated through activation of p38 MAPK, JNK, p42/p44 MAPK, NF-κB, and AP-1 pathways.     

Critical role of TRIF and MyD88 in Mycobacterium tuberculosis Hsp70-mediated activation of dendritic cells

As a potent immune regulator, heat shock protein 70 derived from Mycobacterium tuberculosis (Mtb Hsp70) has adjuvant effect and activates immune cells such as macrophages and dendritic cells (DCs). Although Toll-like receptors (TLRs) are known to involve in DCs activation by Mtb Hsp70, there is still a controversy and the underlying mechanism is not well understood. In this study, we examined whether TRIF and MyD88, the core adaptor molecules for TLRs signaling, regulate Mtb Hsp70-induced DCs activation. Although Mtb Hsp70 produced substantial level of cytokines (IL-6, IL-12p40, and TNF-α) in TRIF-deficient DCs in a dose-dependent manner, each level was significantly lower than that in WT cells. The cytokines production was almost abolished in MyD88-deficient DCs. Consistent with cytokine results, Mtb Hsp70-induced activation of NF-κB and MAPKs was also impaired in both TRIF- and MyD88-deficient DCs, as compared with WT cells. Inhibitor assay revealed that NF-κB, ERK, and JNK, but not p38, regulate Mtb Hsp70-induced production of cytokines. In addition, the up-regulation of co-stimulatory molecules and MHC class II was mostly TRIF-dependent in DCs in response Mtb Hsp70, whereas MyD88 was only partially involved. Finally, mixed leukocytes reaction (MLR) assay revealed that both TRIF and MyD88 are critical for DCs ability promoted by Mtb Hsp70 to differentiate naïve T cells into effector T cells of producing IFN-γ. Our findings suggest that both TRIF and MyD88 are essential for the activation and maturation of DCs in response to Mtb Hsp70.     

Up-regulation of IL-23 p19 expression in human periodontal ligament fibroblasts by IL-1β via concurrent activation of the NF-κB and MAPKs/AP-1 pathways

Interleukin (IL)-23 is an essential cytokine involved in the expansion of a novel CD4(+) T helper subset known as Th17, which has been implicated in the pathogenesis of periodontitis recently. Our previous study first identified specialized human periodontal ligament fibroblasts (hPDLFs) as an important production source of IL-23. The present study was undertaken to investigate the effects of the pro-inflammatory and Th17-polarizing mediator IL-1β on hPDLFs-mediated IL-23 p19 production, and the molecular mechanism involved. IL-23 p19 expression was in situ detected in IL-1β-stimulated hPDLFs. IL-1β was capable of stimulating the expression of IL-23 p19 mRNA and protein in cultured hPDLFs, which was attenuated by IL-1 receptor antagonist (IL-1Ra) or myeloid differentiation primary response gene 88 (MyD88) inhibitor. Meanwhile, inhibitors of p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase (ERK) 1/2, c-Jun-N-terminal kinase (JNK), activator protein-1 (AP-1), or nuclear factor-kappaB (NF-κB) significantly suppressed IL-23 p19 production from IL-1β-stimulated hPDLFs. Moreover, IL-1β-initiated AP-1 activation was blocked by p38 MAPK, ERK 1/2, or JNK inhibition, whereas NF-κB activity remained unaltered by all the above pathway specific inhibitors. Thus, these results provide evidence that Th17-polarizing mediator IL-1β up-regulated the expression of IL-23 p19 in hPDLFs via NF-κB signaling and MAPKs-dependent AP-1 pathways. Taken together, our findings indicate that IL-1Ra may be used therapeutically to inhibit Th17-driven inflammatory diseases including periodontitis.     

MAPKs and NF-κB differentially regulate cytokine expression in the diaphragm in response to resistive breathing: the role of oxidative stress

Inspiratory resistive breathing (IRB) induces cytokine expression in the diaphragm. The mechanism of this cytokine induction remains elusive. The roles of MAPKs and NF-κB and the impact of oxidative stress in IRB-induced cytokine upregulation in the diaphragm were studied. Wistar rats were subjected to IRB (50% of maximal inspiratory pressure) via a two-way nonrebreathing valve for 1, 3, or 6 h. Additional groups of rats subjected to IRB for 6 h were randomly assigned to receive either solvent or N-acetyl-cysteine (NAC) or inhibitors of NF-κB (BAY-11-7082), ERK1/2 (PD98059), and P38 MAPK (SB203580) to study the effect of oxidative stress, NF-κB, and MAPKs in IRB-induced cytokine upregulation in the diaphragm. Quietly breathing animals served as controls. IRB upregulated cytokine (IL-6, TNF-α, IL-10, IL-2, IL-1β) protein levels in the diaphragm and resulted in increased activation of MAPKs (P38, ERK1/2) and NF-κB. Inhibition of NF-κB and ERK1/2 blunted the upregulation of all cytokines except that of IL-6, which was further increased. P38 inhibition attenuated all cytokine (including IL-6) upregulation. Both P38 and ERK1/2 inhibition decreased NF-κB/p65 subunit phosphorylation. NAC pretreatment blunted IRB-induced cytokine upregulation in the diaphragm and resulted in decreased ERK1/2, P38, and NF-κB/p65 phosphorylation. In conclusion, IRB-induced cytokine upregulation in the diaphragm is under the regulatory control of MAPKs and NF-κB. IL-6 is regulated differently from all other cytokines through a P38-dependent and NF-κB independent pathway. Oxidative stress is a stimulus for IRB-induced cytokine upregulation in the diaphragm.     

CCL27 expression is regulated by both p38 MAPK and IKKβ signalling pathways

The skin-specific chemokine CCL27 is believed to play a pivotal role in establishing the inflammatory infiltrate characteristic for common inflammatory skin diseases. Through binding to the chemokine receptor 10 (CCR10), CCL27 mediates inflammation by promoting lymphocyte migration into the skin. Little is known about the regulation of CCL27 gene expression. The purpose of our study was to investigate the regulation of the IL-1β-induced CCL27 gene expression in normal human keratinocytes (NHEK). Preincubation of NHEK with the inhibitory κB (IκB) kinase (IKK) inhibitor, SC-514, or the p38 mitogen-activated protein kinase (MAPK) inhibitor, SB202190, revealed a profound reduction in both CCL27 mRNA and CCL27 protein expression indicating the significance of these pathways in the regulation of CCL27 expression. Furthermore, the impact of inhibitors of mitogen- and stress-activated kinase 1 (MSK1) or the mitogen-activated protein kinase-interacting kinases (Mnk1+2), downstream kinases of p38 MAPK, on IL-1β-induced CCL27 expression in NHEK were investigated. We identified seven NF-κB binding elements upstream from the CCL27 gene start codon using electrophoretic mobility shift assay (EMSA). Supershift analyses demonstrated the involvement of the p50/p65 NF-κB heterodimer. We conclude that IL-1β-induced CCL27 gene expression in NHEK is regulated through the p38 MAPK/MSK1/Mnk1+2 as well as the IKKβ/NF-κB signalling pathways.     

Salidroside promotes rat spinal cord injury recovery by inhibiting inflammatory cytokine expression and NF-κB and MAPK signaling pathways

Spinal cord injury (SCI) is a public health problem in the world. The SCI usually triggers an excessive inflammatory response that brings about a secondary tissue wreck leading to further cellular and organ dysfunction. Hence, there is great potential of reducing inflammation for therapeutic strategies of SCI. In this study, we aim to investigate if Salidroside (SAD) exerts an anti-inflammatory effect and promotes recovery of motor function on SCI through suppressing nuclear factor-κB (NF-κB) and the mitogen-activated protein kinase (MAPK) pathways. In vitro, real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) were used to examine the inhibitory effect of SAD on the expression and release of interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) activated by lipopolysaccharide (LPS) in astrocytes. In addition, SAD was found to inhibit NF-κB, p38 and extracellular-regulated protein kinases (ERK) signaling pathways by western blot analysis. Further, in vivo study showed that SAD was able to improve hind limb motor function and reduce tissue damage accompanied by the suppressed expression of inflammatory cytokines IL-1β, IL-6, and TNF-α. Overall, SAD could reduce the inflammatory response and promote motor function recovery in rats after SCI by inhibiting NF-κB, p38, and ERK signaling pathways.     

Inflammatory gene expression by human colonic smooth muscle cells

Intestinal mucosal cells and invading leukocytes produce inappropriate levels of cytokines and chemokines in human colitis. However, smooth muscle cells of the airway and vasculature also synthesize cytokines and chemokines. To determine whether human colonic myocytes can synthesize proinflammatory mediators, strips of circular smooth muscle and smooth muscle cells were isolated from human colon. Myocytes and muscle strips were stimulated with 10 ng/ml of IL-1beta, TNF-alpha, and IFN-gamma, respectively. Expression of mRNA for IL-1beta, IL-6, IL-8, and cyclooxygenase-2 (COX-2) was induced within 2 h and continued to increase for 8-12 h. Regulated on activation, normal T cell-expressed and -secreted (RANTES) mRNA expression was slower, appearing at 8 h and increasing linearly through 20 h. Expression of all five mRNAs was inhibited by 0.1 microM MG-132, a proteosome inhibitor that blocks NF-kappaB activation. Expression of IL-1beta, IL-6, IL-8, and COX-2 mRNA was reduced by 30 microM PP1, an Src family tyrosine kinase inhibitor, and by 25 microM SB-203580, a p38 MAPK inhibitor. MAPK/extracellular regulated kinase-1 inhibitor PD-98059 (25 microM) was much less effective. In conclusion, human colonic smooth muscle cells can synthesize and secrete interleukins (IL-1beta and IL-6) and chemokines (IL-8 and RANTES) and upregulate expression of COX-2. Regulation of cytokine, chemokine, and COX-2 mRNA depends on multiple signaling pathways, including Src-family kinases, extracellular regulated kinase, p38 MAPKs, and NF-kappaB. SB-203580 was a consistent, efficacious inhibitor of inflammatory gene expression, suggesting an important role of p38 MAPK in synthetic functions of human colonic smooth muscle.     

Cutting edge: IL-4 induces suppressor of cytokine signaling-3 expression in B cells by a mechanism dependent on activation of p38 MAPK

The signaling cascade initiated by IL-4 is classically divisible into two major pathways: one mediated by STAT6, and the other by insulin receptor substrates-1 and -2 via activation of PI3K. In murine splenic B cells, the suppressor of cytokine signaling (SOCS)3 is inducible by IL-4 via a mechanism independent of STAT6 and PI3K. SOCS3 expression increases 9-fold within 5 h of IL-4 treatment. This induction occurs normally in B cells deficient in STAT6 and is unaffected by pretreatment with the PI3K inhibitor wortmannin, or with the ERK pathway inhibitor, PD98059. However, the IL-4 induction of SOCS3 is blocked by inhibitors of either the JNK or p38 MAPK pathways (SP600125 and SB203580, respectively). Direct examination of these pathways reveals rapid, IL-4-directed activation of p38 MAPK, uncovering a previously unappreciated pathway mediating IL-4 signal transduction.     

Cry1Ac toxin induces macrophage activation via ERK1/2, JNK and p38 mitogen-activated protein kinases

The Cry1Ac toxin from Bacillus thuringiensis is used commercially as a bio-insecticide and is expressed in transgenic plants that are used for human and animal consumption. Although it was originally considered innocuous for mammals, the Cry1Ac toxin is not inert and has the ability to induce mucosal and systemic immunogenicity. Herein, we examined whether the Cry1Ac toxin promotes macrophage activation and explored the signalling pathways that may mediate this effect. Treatment of primary and RAW264.7 macrophages with the Cry1Ac toxin resulted in upregulation of the costimulatory molecules CD80, CD86 and ICOS-L and enhanced production of nitric oxide, the chemokine MCP-1 and the proinflammatory cytokines TNF-α and IL-6. Remarkably, the Cry1Ac toxin induced phosphorylation of the mitogen-activated protein kinases (MAPKs) ERK1/2, JNK and p38 and promoted nuclear translocation of nuclear factor-kappa B (NF-κB) p50 and p65. p38 and ERK1/2 MAPKs were involved in this effect, as indicated by the Cry1Ac-induced upregulation of CD80 and IL-6 and TNF-α abrogation by the p38 MAPK inhibitor SB203580. Furthermore, treatment the MEK1/2 kinase inhibitor PD98059 blocked increases in MCP-1 secretion and augmented Cry1Ac-induced ICOS-L upregulation. These data demonstrate the capacity of the Cry1Ac toxin to induce macrophage activation via the MAPK and NF-κB pathways.     

Impairment of cellular immunity is associated with overexpression of heat shock protein 70 in neonatal pigs with intrauterine growth retardation

Neonates with intrauterine growth retardation (IUGR) are susceptible to decreases in cellular immunity. In recent years, a growing body of evidence indicates that Hsp70 may serve as a danger signal to the innate immune system and promote receptor-mediated apoptosis. Using neonatal pigs with IUGR, we investigated immune function of pigs and expression of heat shock protein 70 (Hsp70), nuclear factor-kappa B (NF-κB), and forkhead box O 3a (FoxO3a) in the intestinal tract. Samples from the blood, duodenum, jejunum, and ileum of normal body weight (NBW) piglets and IUGR piglets were collected at day 7 after birth. Furthermore, to test whether Hsp70 is associated with regulation of NF-κB and FoxO3a, Hsp70 was silenced using small RNA interference (siRNA) in IEC-6 cells. Body and intestinal weights were lower in IUGR piglets than in NBW piglets (p?相似文献   

PKD, PKD2, and p38 MAPK mediate Hsp27 serine-82 phosphorylation induced by neurotensin in pancreatic cancer PANC-1 cells

It is widely recognized that Hsp27 is a downstream substrate of the p38 MAPK cascade whereas the role of PKD family members in mediating receptor-stimulated Hsp27 Ser-82 phosphorylation has not been evaluated. Here, we show that neurotensin induced a rapid and striking increase in Hsp27 Ser-82 phosphorylation in PANC-1 cells, which was closely correlated with stimulation of activation loop phosphorylation of PKDs and p38 MAPK Thr180/Tyr182 phosphorylation. Treatment of PANC-1 cells with either the selective PKC inhibitor GF-I or the p38 MAPK inhibitor SB202190 partially reduced neurotensin-induced Hsp27 Ser-82 phosphorylation. However, treatment of the cells with a combination of GF-I and SB202190 virtually abolished neurotensin-induced Hsp27 Ser-82 phosphorylation. Overexpression of PKD in stably transfected PANC-1 cells increased the magnitude and prolonged the duration of Hsp27 Ser-82 phosphorylation in response to neurotensin. Either PKD or PKD2 gene silencing utilizing siRNAs targeting distinct PKD or PKD2 sequences reduced neurotensin-stimulated Hsp27 Ser-82 phosphorylation, but cotransfection of siRNAs targeting both, PKD and PKD2, markedly decreased neurotensin-induced Hsp27 Ser-82 phosphorylation. Knockdown of PKD and PKD2 abolished Hsp27 phosphorylation in cells treated with SB202190. Thus, neurotensin induces Hsp27 Ser-82 phosphorylation through p38 MAPK- and PKC/PKD-dependent pathways in PANC-1 cells. Our results demonstrate, for the first time, that neurotensin induces a striking increase in Hsp27 phosphorylation on Ser-82 in PANC-1 cells through convergent p38 MAPK, PKD, and PKD2 signaling.     

Porphyromonas gingivalis lipopolysaccharide regulates interleukin (IL)-17 and IL-23 expression via SIRT1 modulation in human periodontal ligament cells

Increased interleukin (IL)-17 and IL-23 levels exist in the gingival tissue of periodontitis patients, but the precise molecular mechanisms that regulate IL-17 and IL-23 production remain unknown. The aim of this study was to explore the role of SIRT1 signaling on Porphyromonas gingivalis lipopolysaccharide (LPS)-induced IL-17 and IL-23 production in human periodontal ligament cells (hPDLCs). IL-17 and IL-23 production was significantly increased in LPS-treated cells. LPS treatment also led to the upregulation of SIRT1 mRNA and protein expression. LPS-induced IL-17 and IL-23 upregulation was attenuated by pretreatment with inhibitors of phosphoinositide 3-kinase (PI3K), p38, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), mitogen-activated protein kinase (MAPK), and NF-κB, as well as neutralizing antibodies against Toll-like receptors (TLRs) 2 and 4. Sirtinol treatment (a known SIRT1 inhibitor) or SIRT1 knockdown by small interfering RNA blocked LPS-stimulated IL-17 and IL-23 expression. Further investigation showed that LPS decreased osteoblast markers (i.e., ALP, OPN, and BSP) and concomitantly increased osteoclast markers (i.e., RANKL and M-CSF). This response was attenuated by inhibitors of the PI3K, p38, ERK, JNK, NF-κB, and SIRT1 pathways. These findings, for the first time, suggest that human periodontopathogen P. gingivalis LPS is implicated in periodontal disease bone destruction and may mediate IL-17 and IL-23 release from hPDLCs. This process is dependent, at least in part, on SIRT1-Akt/PI3K-MAPK-NF-κB signaling.