Involvement Of P38 Mitogen-activated Protein Kinase Research Articles

Interleukin‐17A Stimulates Migration of Periodontal Ligament Fibroblasts Via p38 MAPK/NF‐κB ‐Dependent MMP‐1 Expression

Interleukin-17 (IL-17) is a cytokine secreted predominantly by Th17 cells. Although IL-17 is primarily associated with the induction of tissue inflammation, the other biological functions of IL-17, including its wound-healing functions, have yet to be thoroughly explored. Fibroblast proliferation and migration play essential roles in periodontal wound-healing responses. In this study, we report that IL-17A can increase the migration and expression of matrix metalloproteinase (MMP)-1 in human periodontal ligament (PDL) fibroblasts but has no effect on PDL fibroblast proliferation. IL-17A-induced MMP-1 expression led to cell migration, which was attenuated by pre-treatment with IL-17 receptor neutralizing antibody and small interfering RNA (siRNA) for MMP-1. The IL-17A-induced cell migration was also attenuated by its tissue inhibitor of matrix metalloproteinase (TIMP)-1. In addition, a p38 mitogen-activated protein kinase (MAPK) inhibitor (SB203580) inhibited IL-17A-induced increase of the migration and MMP-1 upregulation of PDL fibroblasts. The involvement of p38 MAPK in IL-17A-induced MMP-1 expression and cell migration was further confirmed by transfection of p38α siRNA. A nuclear factor kappaB (NF-κB) inhibitor (pyrrolidine dithiocarbamate) also suppressed the cell migration and MMP-1 expression enhanced by IL-17A. Moreover, transfection with p38α siRNA inhibited IL-17A-induced NF-κB nuclear translocation as well as NF-κB binding activity. Our results suggest that IL-17A enhances the migration of PDL fibroblasts by increasing MMP-1 expression through the IL-17 receptor, p38 MAPK, and NF-κB signal transduction pathways.

Dual role of the p38 MAPK/cPLA 2 pathway in the regulation of platelet apoptosis induced by ABT-737 and strong platelet agonists

p38 Mitogen-activated protein (MAP) kinase is involved in the apoptosis of nucleated cells. Although platelets are anucleated cells, apoptotic proteins have been shown to regulate platelet lifespan. However, the involvement of p38 MAP kinase in platelet apoptosis is not yet clearly defined. Therefore, we investigated the role of p38 MAP kinase in apoptosis induced by a mimetic of BH3-only proteins, ABT-737, and in apoptosis-like events induced by such strong platelet agonists as thrombin in combination with convulxin (Thr/Cvx), both of which result in p38 MAP kinase phosphorylation and activation. A p38 inhibitor (SB202190) inhibited the apoptotic events induced by ABT-737 but did not influence those induced by Thr/Cvx. The inhibitor also reduced the phosphorylation of cytosolic phospholipase A2 (cPLA2), an established p38 substrate, induced by ABT-737 or Thr/Cvx. ABT-737, but not Thr/Cvx, induced the caspase 3-dependent cleavage and inactivation of cPLA2. Thus, p38 MAPK promotes ABT-737-induced apoptosis by inhibiting the cPLA2/arachidonate pathway. We also show that arachidonic acid (AA) itself and in combination with Thr/Cvx or ABT-737 at low concentrations prevented apoptotic events, whereas at high concentrations it enhanced such events. Our data support the hypothesis that the p38 MAPK-triggered arachidonate pathway serves as a defense mechanism against apoptosis under physiological conditions.

Role of Neuronal Nitric Oxide Synthase in the Cardiac Ischemia Reperfusion in Mice

Several studies have demonstrated the role of endothelial and inducible nitric oxide synthase in cardiac ischemia reperfusion(IR). However, the role of neuronal nitric oxide synthase (nNOS) in IR is still controversial. The present study was designed to explore the possible involvement of nNOS in cardiac IR. nNOS-/- knockout (KO) and wild type C57 (WT) mice were subjected to 45 min of ischemia by left descending branch of coronary artery ligation followed by 3 h reperfusion, which plasma was collected for creatine kinase (CK) and lactate dehydrogenase (LDH) measurements, terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining and measurements of activities of caspase-3, -8, -9, phospho-p38, -ERK, -JNK mitogen-activated protein kinase (MAPK) and phospho-nNOS, phospho-eNOS and iNOS. IR induced cardiac tissue apoptosis by increases of TUNEL staining and activities of caspase-3, -8, and -9, accompanied with increase of CK and LDH concentration and phosphorylation of p38, ERK and JNK MAPK and phospho-nNOS, phospho-eNOS and iNOS in both mouse strains. However, IR induced increases of TUNEL staining and activities of caspase-3, -8 and -9, and CK and LDH concentrations and activation of p38 MAPK were markedly lower in KO mice compared with WT mice. But the phosphorylation of eNOS was significantly higher compared with WT IR group (P < 0.05). The data obtained suggest that nNOS exacerbates IR-induced injury maybe involving p38 MAPK activation.

4-Hydroxy-2-Nonenal Enhances Tissue Factor Activity in Human Monocytic Cells via p38 Mitogen-Activated Protein Kinase Activation-Dependent Phosphatidylserine Exposure

4-hydroxy-2-nonenal (HNE) is one of the major aldehydes formed during lipid peroxidation and is believed to play a role in the pathogenesis of atherosclerosis. The objective of the present study is to investigate the effect of HNE on tissue factor (TF) procoagulant activity expressed on cell surfaces. TF activity and antigen levels on intact cells were measured using factor Xa generation and TF monoclonal antibody binding assays, respectively. Exposure of phosphatidylserine on the cell surface was analyzed using thrombin generation assay or by binding of a fluorescent dye-conjugated annexin V. 2',7'-dichlorodihydrofluorescein diacetate was used to detect the generation of reactive oxygen species. Our data showed that HNE increased the procoagulant activity of unperturbed THP-1 cells that express traces of TF antigen, but had no effect on unperturbed endothelial cells that express no measurable TF antigen. HNE increased TF procoagulant activity but not TF antigen of both activated monocytic and endothelial cells. HNE treatment generated reactive oxygen species, activated p38 mitogen-activated protein kinase, and increased the exposure of phosphatidylserine at the outer leaflet in THP-1 cells. Treatment of THP-1 cells with an antioxidant, N-acetyl cysteine, suppressed the above HNE-induced responses and negated the HNE-mediated increase in TF activity. Blockade of p38 mitogen-activated protein kinase activation inhibited HNE-induced phosphatidylserine exposure and increased TF activity. HNE increases TF coagulant activity in monocytic cells through a novel mechanism involving p38 mitogen-activated protein kinase activation that leads to enhanced phosphatidylserine exposure at the cell surface.

Immunoexpression of p38 Mitogen-Activated Proteinkinase in Patients with Aggressive and Chronic Periodontitis

The influence of p38 mitogen-activated proteinkinase (MAPK) expression in the development and progression of periodontal disease is currently under investigation. The aim of the present study is to investigate whether the p38 MAPK expression in gingival tissues correlates with IL-1β levels in gingival crevicular fluid (GCF). Twenty patients with generalized aggressive periodontitis (GAgP), 15 patients with generalized chronic periodontitis (GCP) and 10 healthy subjects (H) were enrolled in the study. Clinical data, gingival tissue biopsies and GCF samples were collected. The expression of p38 was investigated by immunohistochemistry. The levels of IL-1β in GCF were measured using ELISA. Mean clinical parameters and GCF volumes were statistically higher in patients with GAgP and GCP compared to H subjects. Higher levels of IL-1β were found in both periodontitis groups. The p38 expression was significantly increased in inflamed gingival tissues. There were no statistically significant differences in levels of IL-1β and p38 expression between subjects with GAgP and GCP. Our data support the hypothesis that MAPK signaling pathway is an additional player in the pathogenesis of periodontitis. This is the first report to evaluate the involvement of p38 MAPK in patients with GAgP and GCP which might be, in part, considered of value in understanding disease mechanisms.

Modulation of the number and functions of endothelial progenitor cells by interleukin 1β in the peripheral blood of pigs

Endothelial progenitor cells (EPCs) have therapeutic potential for the treatment of organ ischemia following trauma or sepsis, frequently associated with inflammatory conditions. We aimed to investigate the effects of interleukin 1β (IL-1β) on the properties of EPCs and explore its possible relationship with p38 mitogen-activated protein kinase (MAPK). EPCs were isolated from peripheral blood of a porcine model and were characterized. Effects of IL-1β on cell number, proliferation, migration, adhesion, and angiogenic function of EPCs were evaluated in a time- and dose-dependent manner. The activity of p38 MAPK in EPCs was measured by Western blot. Moreover, the effects of SB203580, a specific p38 MAPK inhibitor, on levels of p38 MAPK phosphorylation and the number and functions of EPCs under IL-1β conditions were examined. Incubation of EPCs with IL-1β (5 ng/mL) for 5 days and with IL-1β (0.05-50 ng/mL) for 48 hours induced a significant reduction in EPC numbers and proliferation, respectively (p < 0.01 vs. control cells). The capacities for migration, adhesion, and angiogenic function of EPCs were also reduced in a time- and dose-dependent manner. IL-1β induced dose- and time-dependent activation of p38 MAPK in EPCs. Moreover, inhibition of p38 MAPK by SB203580 significantly increased the total number of EPCs by twofold as compared with the IL-1β-alone group (p < 0.01) and blocked the ability of IL-1β to impair the functional response of EPCs. These results demonstrate that there is a negative cause-effect relationship between IL-1β and EPCs. Thus, IL-1β inhibits EPC proliferation, migration, adhesion, and tube formation by a mechanism, which involves p38 MAPK signaling in regulating the number and functions of EPCs in vitro.

Blueberry protects LPS-stimulated BV-2 microglia through inhibiting activities of p38 MAPK and ERK1/2

The effects of blueberry extract (BE) were investigated on activities of p38 mitogen-activated protein kinase (MAPK) and extracellular extracellular signal regulated kinase 1/2 (ERK1/2) in mouse BV-2 microglia stimulated lipopolysaccharide (LPS). BV-2 microglia were cultured for 24 h in the presence of 1 μg/mL LPS either with or without BE preincubation for 0, 1, and 12 h. BE relieved repression of cell proliferation, and reduced cell death. These alterations caused by BE addition accompanied reduction of radical oxygen species (ROS). There were also significant decreases in the levels of both transcripts, and proteins of inducible nitric oxide synthase (iNOS) and peroxiredoxin 1 (Prx1) genes in the BE-treated cells, suggesting that BE inhibit the ROS-induced gene expression. The antioxidant effects of BE appeared to involve p38 MAPK and ERK1/2 signaling pathways since BE reduced both kinase activities by inhibiting phosphorylation. Taken these results together, BE protects LPS-stimulated BV-2 microglia by reducing cell death, ROS accumulation, and activities of p38 MAPK and ERK1/2.

Accumulating microglia phagocytose injured neurons in hippocampal slice cultures: involvement of p38 MAP kinase.

In this study, microglial migration and phagocytosis were examined in mouse organotypic hippocampal slice cultures, which were treated with N-methyl-D-aspartate (NMDA) to selectively injure neuronal cells. Microglial cells were visualized by the expression of enhanced green fluorescent protein. Daily observation revealed microglial accumulation in the pyramidal cell layer, which peaked 5 to 6 days after NMDA treatment. Time-lapse imaging showed that microglia migrated to the pyramidal cell layer from adjacent and/or remote areas. There was no difference in the number of proliferating microglia between control and NMDA-treated slices in both the pyramidal cell layer and stratum radiatum, suggesting that microglial accumulation in the injured areas is mainly due to microglial migration, not to proliferation. Time-lapse imaging also showed that the injured neurons, which were visualized by propidium iodide (PI), disappeared just after being surrounded by microglia. Daily observation revealed that the intensity of PI fluorescence gradually attenuated, and this attenuation was suppressed by pretreatment with clodronate, a microglia toxin. These findings suggest that accumulating microglia phagocytosed injured neurons, and that PI fluorescence could be a useful indicator for microglial phagocytosis. Using this advantage to examine microglial phagocytosis in living slice cultures, we investigated the involvements of mitogen-activated protein (MAP) kinases in microglial accumulation and phagocytosis. p38 MAP kinase inhibitor SB203580, but not MAP kinase/extracellular signal-regulated kinase inhibitor PD98059 or c-Jun N-terminal kinase inhibitor SP600125, suppressed the attenuation of PI fluorescence. On the other hand, microglial accumulation in the injured areas was not inhibited by any of these inhibitors. These data suggest that p38 MAP kinase plays an important role in microglial phagocytosis of injured neurons.

Overexpression of Aurora-A Enhances Invasion and Matrix Metalloproteinase-2 Expression in Esophageal Squamous Cell Carcinoma Cells

Esophageal squamous cell carcinoma (ESCC) is one of the most aggressive cancers, and metastasis is the principal cause of death in ESCC patients. It has been shown that amplification and overexpression of mitotic serine/threonine kinase Aurora-A occur in several types of human tumors, including ESCC. Moreover, increase in expression levels of Aurora-A has been predicted to correlate with the grades of tumor differentiation and invasive capability. However, the mechanisms by which Aurora-A mediates its invasive effects still remain elusive. In this article, we showed that Aurora-A overexpression significantly increased cell migration and invasion as well as secretion and expression of matrix metalloproteinase-2 (MMP-2). Conversely, siRNA-mediated knockdown of Aurora-A expression in human ESCC cells led to inhibition of cell invasiveness as well as secretion and expression of MMP-2. In addition, Aurora-A overexpression increased phosphorylation levels of p38 mitogen-activated protein kinase (MAPK) and Akt, and the knockdown of Aurora-A by siRNA decreased the activity of p38 MAPK and Akt. Moreover, the blocking of the activity of above kinases using chemical inhibitors suppressed the ability of Aurora-A to induce MMP-2 secretion and expression as well as cell invasion. These data show that overexpression of Aurora-A contributes to the malignancy development of ESCC by enhancing tumor cell invasion as well as MMP-2 activity and expression, which can occur through signaling pathways involving p38 MAPK and Akt protein kinases. Taken together, these studies provide a molecular basis for promoting the role of Aurora-A in malignancy development of ESCC.

Inhibition of p38 MAPK Reduces Expression of Vascular Endothelial Growth Factor in Allergic Airway Disease

The p38 mitogen-activated protein kinase (MAPK) appears to play an important role in various pathophysiological responses and has been suggested to be involved in many processes considered critical to the inflammatory response and tissue remodeling. Bronchial asthma is a chronic inflammatory disorder of the airway accompanied by increased vascular permeability. Vascular endothelial growth factor (VEGF) is a potent stimulator of bronchial inflammation, airway remodeling, and physiologic dysregulation that augments antigen sensitization and T-helper type 2 cell (Th2)-mediated inflammation in allergic airway diseases. However, there are little data on the relationship between p38 MAPK signaling and VEGF expression in allergic airway disease. This study aimed to investigate the role of p38 MAPK on the pathogenesis of allergic airway disease, more specifically in VEGF expression. Using ovalbumin (OVA)-inhaled mice and a selective p38 MAPK inhibitor, SB 239063, the involvement of p38 MAPK in allergen-induced VEGF expression in the airway was evaluated. The increases of phosphorylation of p38 MAPK, VEGF protein expression, and vascular permeability in the lung after OVA inhalation were decreased substantially by the administration of SB 239063. In addition, SB 239063 significantly reduced the increase of Th2 cytokines and OVA-specific IgE. The inhibition of p38 MAPK or VEGF signaling prevented and also decreased the increases in the number of inflammatory cells and airway hyperresponsiveness in OVA-induced allergic airway disease. These results indicate that inhibition of p38 MAPK may attenuate allergen-induced airway inflammation and vascular leakage through modulation of VEGF expression in mice.

Abstract 16962: Angiotensin II Limits NO Production by Upregulating Arginase through a Mitogen Activated Protein Kinase-Activating Transcription Factor-2 pathway

Nitric oxide (NO) produced by endothelial nitric oxide synthase (NOS) is needed for normal vascular function. During hypertension, diabetes or aging, elevated levels of arginase (ARG) can compete with NOS for available L-arginine thus reducing vascular NO production. Elevated angiotensin II (Ang II) is a key participant of endothelial dysfunction in many cardiovascular diseases and has been linked to elevated ARG activity. In this study we explored the signaling pathway leading to increased ARG expression/activity in responses to Ang II in bovine aortic endothelial cells (BAEC). Treatment of BAEC with Ang II (10 -7 M, 24 hrs) caused a 40±6% increase (p&lt;0.05) in ARG activity. This was accompanied by 30±8% decrease (p&lt;0.05) in NO production. Our studies indicate involvement of p38 mitogen activated protein kinase (MAPK) in Ang II-induced ARG upregulation and reduced NO production as p38 MAPK inhibitor SB202190 (2 μM) prevented the Ang II-induced increase in ARG activity. Our mouse studies also show involvement of p38 MAPK in Ang II-induced vascular dysfunction associated with elevated ARG activity and ARG expression. Ang II (42 µg/kg/h, sc, 2 wks) caused impaired EC-dependent vasorelaxation in mouse aorta (55±7% vs. 75±8% for control). This impairment was prevented by treatment with p38 inhibitor SB203580 (5µg/kg/day). Ang II also caused a 6.2 fold increase in vascular ARG activity/expression that was completely prevented by p38 MAPK inhibition. Additionally, treatment of BAEC with Ang II causes phosphorylation of activating transcription factor-2 (ATF-2) by 10 minutes with a peak at 30 minutes (3.9 fold). Transfection of BAEC with ATF-2 siRNA prevents Ang II-induced increases in ARG activity/expression and maintains NO production. These results indicate that ATF-2 is necessary for enhanced expression of ARG by Ang II. Collectively our results indicate that Ang II increases endothelial ARG activity/expression through a p38 MAPK-ATF-2 pathway leading to reduced NO production and endothelial dysfunction. Targeting these signaling steps might be therapeutic points for preventing vascular endothelial dysfunction associated with elevated ARG activity/expression.

Translocation and oligomerization of Bax is regulated independently by activation of p38 MAPK and caspase-2 during MN9D dopaminergic neurodegeneration

Bax is translocated into the mitochondrial membrane and oligomerized therein to initiate mitochondrial apoptotic signaling. Our previous study indicated that reactive oxygen species (ROS)-mediated activation of mitogen-activated protein kinase (MAPK) and caspase is critically involved in 6-hydroxydopamine (6-OHDA)-mediated neurodegeneration. Here, we specifically attempted to examine whether and how these death signaling pathways may be linked to Bax translocation and oligomerization. We found that 6-OHDA treatment triggered translocation and oligomerization of Bax onto the mitochondria in MN9D dopaminergic neuronal cells. These events preceded cytochrome c release into the cytosol. Cross-linking assay revealed that co-treatment with a ROS scavenger or a pan-caspase inhibitor inhibited 6-OHDA-induced Bax oligomerization. Among several candidates of ROS-activated MAPKs and caspases, we found that co-treatment with PD169316 or VDVAD specifically inhibited 6-OHDA-induced Bax oligomerization, suggesting critical involvement of p38 MAPK and caspase-2. Consequently, overexpression of a dominant negative form of p38 MAPK or a shRNA-mediated knockdown of caspase-2 indeed inhibited 6-OHDA-induced Bax oligomerization. However, activation of p38 MAPK and caspase-2 was independently linked to oligomerization of Bax. This specificity was largely confirmed with a Bax 6A7 antibody known to detect activated forms of Bax on the mitochondria. Taken together, our data suggest that there is an independent amplification loop of Bax translocation and oligomerization via caspase-2 and p38 MAPK during ROS-mediated dopaminergic neurodegeneration.

Adenosine A1 and A3 receptor agonists reduce hypoxic injury through the involvement of P38 MAPK

Activation of either the A(1) adenosine receptor (A(1)R) or the A(3) adenosine receptor (A(3)R), by their specific agonists CCPA and Cl-IB-MECA, respectively, protects cardiac cells in culture against ischemic injury. Yet the full protective mechanism remains unclear. In this study, we therefore examined the involvement of p38 mitogen-activated protein kinase (MAPK) and extracellular signal-regulated kinases (ERK) phosphorylation in this protective intracellular signaling mechanism. Furthermore, we investigated whether p38 MAPK phosphorylation occurs upstream or downstream from the opening of mitochondrial ATP-sensitive potassium (K(ATP)) channels. The role of p38 MAPK activation in the intracellular signaling process was studied in cultured cardiomyocytes subjected to hypoxia, that were pretreated with CCPA or Cl-IB-MECA or diazoxide (a mitochondrial K(ATP) channel opener) with and without SB203580 (a specific inhibitor of phosphorylated p38 MAPK). Cardiomyocytes were also pretreated with anisomycin (p38 MAPK activator) with and without 5-hydroxy decanoic acid (5HD) (a mitochondrial K(ATP) channel blocker). SB203580 together with the CCPA, Cl-IB-MECA or diazoxide abrogated the protection against hypoxia as shown by the level of ATP, lactate dehydrogenase (LDH) release, and propidium iodide (PI) staining. Anisomycin protected the cardiomyocytes against ischemic injury and this protection was abrogated by SB203580 but not by 5HD. Conclusions Activation of A(1)R or A(3)R by CCPA or Cl-IB-MECA, respectively, protects cardiomyocytes from hypoxia via phosphorylation of p38 MAPK, which is located downstream from the mitochondrial K(ATP) channel opening. Elucidating the signaling pathway by which adenosine receptor agonists protect cardiomyocytes from hypoxic damage, will facilitate the development of anti ischemic drugs.

Evidence that adiponectin receptor 1 activation exacerbates ischemic neuronal death

Background-Adiponectin is a hormone produced in and released from adipose cells, which has been shown to have anti-diabetic and anti-inflammatory actions in peripheral cells. Two cell surface adiponectin receptors (ADRs) mediate the majority of the known biological actions of adiponectin. Thus far, ADR expression in the brain has been demonstrated in the arcuate and the paraventricular nucleus of hypothalamus, where its activation affects food intake. Recent findings suggest that levels of circulating adiponectin increase after an ischemic stroke, but the role of adiponectin receptor activation in stroke pathogenesis and its functional outcome is unclear.Methods-Ischemic stroke was induced in C57BL/6 mice by middle cerebral artery occlusion (MCAO) for 1 h, followed by reperfusion. Primary cortical neuronal cultures were established from individual embryonic neocortex. For glucose deprivation (GD), cultured neurons were incubated in glucose-free Locke's medium for 6, 12 or 24 h. For combined oxygen and glucose deprivation (OGD), neurons were incubated in glucose-free Locke's medium in an oxygen-free chamber with 95% N2/5% CO2 atmosphere for either 3, 6, 9, 12 or 24 h. Primary neurons and brain tissues were analysed for Adiponectin and ADRs using reverse transcriptase polymerase chain reaction (RT-PCR), immunoblot and immunochemistry methods.Results-Cortical neurons express ADR1 and ADR2, and that the levels of ADR1 are increased in neurons in response to in vitro or in vivo ischemic conditions. Neurons treated with either globular or trimeric adiponectin exhibited increased vulnerability to oxygen and glucose deprivation which was associated with increased activation of a pro-apoptotic signaling cascade involving p38 mitogen-activated protein kinase (p38MAPK) and AMP-activated protein kinase (AMPK).Conclusions-This study reveals a novel pathogenic role for adiponectin and adiponectin receptor activation in ischemic stroke. We show that cortical neurons express ADRs and reveal a pro-apoptotic role for ADR1 activation in neurons, which may render them vulnerable to ischemic death.

Chitosan oligosaccharides protect human umbilical vein endothelial cells from hydrogen peroxide-induced apoptosis

This study aimed to investigate the protective effect of chitosan oligosaccharides (COS) on human umbilical vein endothelial cell (HUVEC) apoptosis induced by hydrogen peroxide (H(2)O(2)). We found that COS not only reversed the decrease of cell viability and proliferation activity, but ameliorated nuclear chromatin damage in H(2)O(2)-induced HUVECs. Additionally, COS contributed to the decrease of cytosolic Ca(2+) level, increase of mitochondrial membrane potential, up-regulation of Bcl-2 mRNA, down-regulation of Bax mRNA, reduction of cleaved caspase-3 protein, inhibition of phosphorylated p38 mitogen-activated protein kinase (MAPK) and induction of phosphorylated Akt in HUVECs. Further, H(2)O(2)-induced caspase-3 activation could be suppressed by p38 MAPK inhibitor (SB203580) and up-regulated by phosphatidylinositol-3-kinase (PI3K) inhibitor (LY294002), indicating the involvement of p38 MAPK and PI3K/Akt signaling pathways. In conclusion, the results show that COS may effectively inhibit the H(2)O(2)-induced HUVEC apoptosis. (C) 2010 Elsevier Ltd. All rights reserved.

ACh receptors link two signaling pathways to neuroprotection against glutamate‐induced excitotoxicity in isolated RGCs

Previous studies have reported that activation of nicotinic acetylcholine (ACh) receptors (nAChRs) on cultured pig retinal ganglion cells (RGCs) has a neuroprotective effect against glutamate-induced excitotoxicity. However, the mechanism linking nAChRs to neuroprotection is unknown. Here, we tested the hypothesis that signaling cascades involving p38 mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) --> Akt are involved in linking activation of nAChRs to neuroprotection in isolated pig RGCs. In ELISA studies, regulation of phosphorylated p38 MAPK and Akt were analyzed after inducing excitotoxicity or neuroprotection in the presence and absence of specific inhibitors for p38 MAPK and PI3K. ELISA results demonstrated that ACh significantly increased phosphorylated Akt and decreased p38 MAPK. Glutamate increased phosphorylated p38 MAPK but had no significant effect on phosphorylated Akt. Other ELISA studies using p38 MAPK and PI3K inhibitors also supported the hypothesis that ACh up-regulated Bcl-2 levels downstream from PI3K and Akt, whereas glutamate down-regulated Bcl-2 levels downstream from p38 MAPK. RGC survival was subsequently assessed by culturing RGCs in conditions to induce excitotoxicity or neuroprotection in the presence or absence of specific inhibitors of p38 MAPK or PI3K. The p38 MAPK inhibitor significantly decreased the number of RGCs that died by glutamate-induced excitotoxicity but had no effect on the number of cells that survived because of ACh-induced neuroprotection. PI3K inhibitors significantly decreased cell survival caused by ACh-induced neuroprotection but had no effect on cell death caused by glutamate-induced excitotoxicity. These results demonstrate that glutamate mediates excitotoxicity through the p38 MAPK signaling pathway and that ACh provides neuroprotection by stimulating the PI3K --> Akt --> Bcl-2 signaling pathway and inhibiting the p38 MAPK --> Bcl-2 pathway.

Correction: Zhou, Z.-B. et al. Receptor-Mediated Vascular Smooth Muscle Migration Induced by LPA Involves p38 Mitogen-Activated Protein Kinase Pathway Activation. Int. J. Mol. Sci. 2009, 10, 3194-3208

Due to personal reasons, I left the research group. In accordance with the regulations of the funding institution, Xiamen Health Administration, China, I hereby declare a withdrawal of my signature, Zhi-Jun Zhang, from this paper. [...]

The p38 MAPK inhibitors for the treatment of inflammatory diseases and cancer

Background: The p38 mitogen-activated protein kinase (MAPK) is activated by various pro-inflammatory and stressful stimuli. Mounting evidence suggests that the p38 MAPK signaling cascade is involved in various biological responses other than inflammation such as cell proliferation, differentiation, apoptosis and invasion, suggesting that the p38 MAPK can serve as a potential therapeutic target for the treatment of not only inflammatory diseases but also cancer. Methods: The unique characteristics of p38 MAPK are summarized with regard to activation and function of p38 MAPK signaling cascades. We then discuss the involvement of p38 MAPK in diseases and the implications of the possible therapeutic use of p38 MAPK inhibitors. The p38 MAPK inhibitors that have been used in the in vitro/in vivo systems as well as in the clinical trials are summarized. Results/conclusion: The p38 MAPK plays an important role in key cellular processes related to inflammation and cancer. Understanding the signal transduction mechanisms and gene regulation by p38 MAPK provides useful information in the development of p38 MAPK inhibitors with therapeutic benefits with reduced side effects. In this review, we summarize and present the list of p38 MAPK inhibitors in in vitro/in vivo studies as well as in clinical trials.

Curcumin-induced apoptosis in ovarian carcinoma cells is p53-independent and involves p38 mitogen-activated protein kinase activation and downregulation of Bcl-2 and survivin expression and Akt signaling

New cytotoxic agents are urgently needed for the treatment of advanced ovarian cancer because of the poor long-term response of this disease to conventional chemotherapy. Curcumin, obtained from the rhizome of Curcuma longa, has potent anticancer activity; however, the mechanism of curcumin-induced cytotoxicity in ovarian cancer cells remains a mystery. In this study we show that curcumin exhibited time- and dose-dependent cytotoxicity against monolayer cultures of ovarian carcinoma cell lines with differing p53 status (wild-type p53: HEY, OVCA429; mutant p53: OCC1; null p53: SKOV3). In addition, p53 knockdown or p53 inhibition did not diminish curcumin killing of HEY cells, confirming p53-independent cytotoxicity. Curcumin also killed OVCA429, and SKOV3 cells grown as multicellular spheroids. Nuclear condensation and fragmentation, as well as DNA fragmentation and poly (ADP-ribose) polymerase-1 cleavage in curcumin-treated HEY cells, indicated cell death by apoptosis. Procaspase-3, procaspase-8, and procaspase-9 cleavage, in addition to cytochrome c release and Bid cleavage into truncated Bid, revealed that curcumin activated both the extrinsic and intrinsic pathways of apoptosis. Bax expression was unchanged but Bcl-2, survivin, phosphorylated Akt (on serine 473), and total Akt were downregulated in curcumin-treated HEY cells. Curcumin also activated p38 mitogen-activated protein kinase (MAPK) without altering extracellular signal-regulated kinase 1/2 activity. We conclude that p53-independent curcumin-induced apoptosis in ovarian carcinoma cells involves p38 MAPK activation, ablation of prosurvival Akt signaling, and reduced expression of the antiapoptotic proteins Bcl-2 and survivin. These data provide a mechanistic rationale for the potential use of curcumin in the treatment of ovarian cancer.

Receptor-Mediated Vascular Smooth Muscle Migration Induced by LPA Involves p38 Mitogen-Activated Protein Kinase Pathway Activation

Lysophosphatidic acid (LPA), a naturally occurring glycerophospholipid, can evoke various biological responses, including cell migration, proliferation and survival, via activation of G protein-coupled receptors (GPCRs). However, the role of LPA receptors and details of LPA signaling in migration are largely unexplored. In this study we detect the expression of LPA1 and LPA3 receptors in rat aortic smooth muscle cells (RASMCs). LPA stimulated RASMCs migration in a dose-dependent manner and induced the phosphorylation of p38 mitogen-activated protein kinase (p38MAPK) and extracellular signal-regulated kinase (ERK). LPA-induced cell migration was significantly inhibited by specific LPA1/LPA3-receptor antagonist Dioctylglycerol pyrophosphate (8:0) (DGPP8.0) at higher concentration. Migration of cells toward LPA was partially, but significantly, reduced in the presence of SB-203580, a p38 MAPK inhibitor, but not PD98059, an ERK inhibitor. In addition, pertussis toxin (PTX), a Gi protein inhibitor, induced an inhibitory effect on p38 MAPK, ERK phosphorylation and RASMCs migration. These data suggest that LPA-induced migration is mediated through the Gi-protein-coupled LPA1 receptor involving activation of a PTX-sensitive Gi / p38MAPK pathway.