Role Of P38 MAP Kinase Research Articles

Daily ketoprofen restores AIH‐induced phrenic long‐term facilitation after prolonged chronic intermittent hypoxia in spinal intact and chronically injured rats

Acute intermittent hypoxia (AIH) elicits a form of spinal, respiratory motor plasticity known as phrenic long‐term facilitation (pLTF). In recent years, this form of plasticity has been leveraged for therapeutic benefit to restore respiratory (and non‐respiratory) motor function after spinal cord injury (SCI). While therapeutic AIH shows tremendous promise as a non‐invasive and easy‐to administer rehabilitation strategy, its efficacy may be undermined by factors such as inflammation (Huxtable et al., 2011). For example, a single night of chronic intermittent hypoxia (CIH) simulating that experienced by an individual with moderate sleep apnea (2 min hypoxic episodes; 2 min intervals; 8 hrs) abolishes AIH‐induced pLTF through a mechanism that requires systemic inflammation and spinal p38 MAP kinase activity (Huxtable et al., 2015). Similarly, prolonged CIH (7‐28 days) also abolishes pLTF (Gonzalez‐Rothi et al., 2021; El Chami and colleagues, unpublished), though the underlying mechanisms have not yet been defined. Since CIH induces time‐dependent cascades of distinct spinal inflammatory gene expression (Smith et al., 2013), we hypothesize that prolonged CIH (versus a single night) blunts pLTF through an inflammation‐dependent mechanism, although the specific pathways may differ with cumulative/repeated exposures. Further, since respiratory insufficiency is a leading cause of morbidity and mortality following cervical SCI and sleep disordered breathing/sleep apnea are highly prevalent (nearly 80% in cervical SCI), it is important to understand how untreated sleep apnea may interact with and/or undermine the efficacy of therapeutic interventions like AIH in chronic SCI. Thus, we are exploring the impact of anti‐inflammatory administration on AIH‐induced pLTF in both intact rats and rats with chronic (12 wks) C3/C4 mid‐cervical spinal cord contusion, preconditioned with 7 days of moderate CIH. In intact rats, daily administration of the non‐steroidal anti‐inflammatory drug, ketoprofen rescued expression of pLTF following prolonged CIH (70.4% increase from baseline, p=0.0039 vs. vehicle). Interestingly, a single dose of ketoprofen delivered just prior to pLTF assessments did not restore expression of plasticity (19.5% increase, p=0.8842 vs. vehicle), suggesting a shift in the underlying mechanisms by which CIH impacts phrenic motor plasticity that occurs with cumulative exposure over time. Similarly, in rats with chronic cSCI, daily ketoprofen rescued expression of pLTF following prolonged CIH (62.9% increase from baseline, p=0.0348 vs. vehicle). Upcoming experiments will explore the role of p38 MAP kinase specifically after prolonged CIH. Improved understanding of factors that undermine plasticity, (e.g., CIH, inflammation), will enable us to identify strategies to mitigate deleterious effects and optimize functional benefits, which is essential for effectively translating promising interventions like therapeutic AIH to clinical practice.

Daily administration of ketoprofen restores AIH‐induced phrenic long term facilitation with prolonged chronic intermittent hypoxia

Acute intermittent hypoxia (AIH) elicits a form of spinal, respiratory motor plasticity known as phrenic long-term facilitation (pLTF). Preconditioning with a single day of chronic intermittent hypoxia (8 hours, 2 min hypoxic episodes with 2 min intervals) abolishes AIH-induced pLTF through a mechanism that requires systemic inflammation and spinal p38 MAP kinase activation (Huxtable et al., 2015). pLTF is restored by high-dose, systemic administration of the non-steroidal anti-inflammatory drug, ketoprofen, or spinal p38 MAP kinase inhibition. Similarly, preconditioning with 7 days of CIH (8 hours per day, 2 min episodes, 2 min intervals) abolishes pLTF, but we observed that phospho-p38 within phrenic motor neurons is unchanged from control rats (Gonzalez-Rothi and colleagues, unpublished). Thus, mechanisms whereby 7 days of CIH undermines AIH-induced pLTF are unclear. Since CIH induces time-dependent spinal inflammatory gene expression profiles over time (Smith et al., 2013), we hypothesize that prolonged CIH (versus a single day) blunts LTF through an inflammation-dependent mechanism that is independent of p-38MAP kinase. Thus, in ongoing experiments, the impact of anti-inflammatory drugs on AIH-induced pLTF in rats preconditioned with prolonged CIH (7 days). pLTF was restored when ketoprofen was administered each day just prior to the IH7 exposure (70% increase, p=.0013 vs. vehicle), but not when a single dose of ketoprofen was delivered just prior to assessment of pLTF (19% increase, p=0.9975 vs. vehicle). Upcoming experiments will explore the role of p38 MAP kinase with prolonged CIH exposure. These experiments are the first to explore shifting mechanisms whereby prolonged CIH and associated inflammation undermine phrenic motor plasticity. Such experiments are essential since chronic intermittent hypoxia is a hallmark of sleep apnea, a condition highly prevalent in individuals living with neurological disorders (spinal cord injury, ALS, etc.). It is therefore important to identify and mitigate factors which may undermine the potential for therapeutic interventions (i.e., repetitive AIH) to harness endogenous plasticity and ultimately improve breathing.

The sex-specific role of p38 MAP kinase in CNS autoimmunity is regulated by estrogen receptor alpha

The sex-specific role of p38 MAP kinase in CNS autoimmunity is regulated by estrogen receptor alpha

Role of p38 MAP kinase signaling pathways in storage and voiding dysfunction in mice with spinal cord injury.

To investigate the role of p38 MAP kinase in lower urinary tract dysfunction in mice with spinal cord injury (SCI). Cystometry and external urethral sphincter-electromyography were performed under an awake condition in 4-week SCI female mice. Two weeks after SCI, a catheter connected to an osmotic pump filled with a p38 mitogen-activated protein kinase (MAPK) inhibitor or artificial cerebrospinal fluid (CSF) was implanted into the intrathecal space of L6-S1 spinal cord for continuous intrathecal instillation at infusion rate of 0.51 μL/h for 2 weeks before the urodynamic study. L6 dorsal root ganglia were then removed from CSF and p38 MAPK inhibitor-treated SCI mice as well as from CSF-treated normal (spinal intact) mice to evaluate the levels of transient receptor potential cation channel subfamily V member 1 (TRPV1), tumor necrosis factor-α (TNF-α), and inducible nitric oxide synthase (iNOS) transcripts by real-time polymerase chain reaction. In p38 MAPK inhibitor-treated SCI mice, nonvoiding contractions during bladder filling, bladder capacity, and post-void residual volume were significantly reduced while micturition pressure and voiding efficiency were significantly increased in comparison to these measurements in CSF-treated SCI mice. The expression of TRPV1, TNF-α, and iNOS messenger RNA was increased in SCI mice compared with expression in spinal intact mice and significantly decreased after p38 MAPK inhibitor treatment. The p38 MAPK signaling pathway in bladder sensory neurons or in the spinal cord plays an important role in storage and voiding problems such as detrusor overactivity and inefficient voiding after SCI.

Role of p38alpha/beta MAP Kinase in Cell Susceptibility to Clostridium sordellii Lethal Toxin and Clostridium difficile Toxin B.

Lethal Toxin from Clostridium sordellii (TcsL), which is casually involved in the toxic shock syndrome and in gas gangrene, enters its target cells by receptor-mediated endocytosis. Inside the cell, TcsL mono-O-glucosylates and thereby inactivates Rac/Cdc42 and Ras subtype GTPases, resulting in actin reorganization and an activation of p38 MAP kinase. While a role of p38 MAP kinase in TcsL-induced cell death is well established, data on a role of p38 MAP kinase in TcsL-induced actin reorganization are not available. In this study, TcsL-induced Rac/Cdc42 glucosylation and actin reorganization are differentially analyzed in p38alpha−/− MSCV empty vector MEFs and the corresponding cell line with reconstituted p38alpha expression (p38alpha−/− MSCV p38alpha MEFs). Genetic deletion of p38alpha results in reduced susceptibility of cells to TcsL-induced Rac/Cdc42 glucosylation and actin reorganization. Furthermore, SB203580, a pyridinyl imidazole inhibitor of p38alpha/beta MAP kinase, also protects cells from TcsL-induced effects in both p38−/− MSCV empty vector MEFs and in p38alpha−/− MSCV p38alpha MEFs, suggesting that inhibition of p38beta contributes to the protective effect of SB203580. In contrast, the effects of the related C. difficile Toxin B are responsive neither to SB203580 treatment nor to p38alpha deletion. In conclusion, the protective effects of SB203580 and of p38alpha deletion are likely not based on inhibition of the toxins’ glucosyltransferase activity rather than on inhibited endocytic uptake of specifically TcsL into target cells.

The role of p38 MAP-kinase in stress-induced senescence of human endometrium-derived mesenchymal stem cells

Our recent findings demonstrate that human endometrium-derived mesenchymal stem cells (hMESCs) respond to sublethal oxidative stress by stress-induced premature senescence via the АТМ/Chk2/p53/p21/Rb pathway. Application of SB203580 (SB) inhibitor suggested p38 MAP-kinase involvement in the senescence progression. However, there are several disadvantages concerning this inhibitor: (1) SB is toxic and hardly suitable for in vivo experiments and (2) poor kinase selectivity profile of SB complicates interpretation of the obtained data. Here, to confirm the involvement of p38 in H2O2-induced hMESCs senescence, we applied another highly specific p38 inhibitor, BIRB796 (BIRB). In the presence of BIRB, the cell size decreased, the level of reactive oxygen species reduced, proliferation partially resumed, and Rb phosphorylation level increased in comparison to H2O2-treated hMESCs. Summarizing these results, we can postulate p38 involvement in H2O2-induced senescence of hMESCs and suggest p38 inhibition as a promising approach in prevention of premature senescence.

Budesonide Inhibits Intracellular Infection with Non-Typeable Haemophilus influenzae despite Its Anti-Inflammatory Effects in Respiratory Cells and Human Lung Tissue: A Role for p38 MAP Kinase

Background: Inhaled corticosteroids (ICS) are widely used in the treatment of obstructive lung diseases. Recent data suggest a higher pneumonia risk in chronic obstructive pulmonary disease (COPD) patients treated with ICS. Objective: Since non-typeable Haemophilus influenzae (NTHi) is the most common pathogen associated with acute exacerbations of COPD, we investigated the effects of budesonide (BUD) on NTHi-induced inflammation and invasive infection. Methods: The alveolar epithelial cell line A549 and specimens of human lung tissue (HLT) were used in our experiments. Intracellular infection was determined by a lysis/culture assay of infected cells. Activated p38 mitogen-associated protein kinase (MAPK) was assessed using Western blotting and immunohistochemistry, expression of toll-like receptor 2 (TLR2) was determined by PCR, and CXCL-8 levels were measured using ELISA. Immunohistochemistry was used for detection of CXCL-8, platelet-activating factor receptor (PAF-R) and NTHi. Results: BUD significantly reduced CXCL-8 secretion in A549 cells and lung tissue infected with NTHi. Furthermore, BUD decreased the expression of PAF-R in HLT and A549 cells. In A549 cells and HLT, BUD inhibited intracellular infection and - synergistically with NTHi - increased the expression of TLR2 (in A549 cells). TLR2 stimulation did not influence the intracellular infection of A549 cells, but p38 MAPK inhibition resulted in a significant reduction of infection. Conclusion: The present study adds new insights into the effects of glucocorticoids on pulmonary host defence after NTHi infection. Although the inflammatory response to infection is suppressed by BUD, interestingly, the intracellular infection is also inhibited. This effect seems to depend on the inhibition of p38 MAPK - a key enzyme in many pro-inflammatory pathways - as well as of PAF-R expression.

The Small GTPase RhoB Regulates TNFα Signaling in Endothelial Cells

The inflammatory response of endothelial cells triggered by cytokines such as TNFα and IL1β plays a pivotal role in innate immunity. Upon pro-inflammatory cytokine stimulation, endothelial cells produce chemokines and cytokines that attract and activate leukocytes, and express high levels of leukocyte adhesion molecules. This process is mediated by intracellular signaling cascades triggered by activation of e.g. the TNFα receptor (TNFR) that lead to the activation of the NFκB transcription factor and of MAP kinases, which in turn activate inflammatory gene transcription. We found that the small GTPase RhoB was strongly and rapidly upregulated in primary human endothelial cells by TNFα, IL1β and LPS. We subsequently investigated the role of RhoB in the regulation of TNFR signaling in endothelial cells by silencing RhoB expression with siRNA. We provide evidence that the TNFα-induced activation of p38 MAP kinase is strongly dependent on RhoB, but not on RhoA, while JNK activation is regulated by both RhoB and RhoA. Consistent with the important role of p38 MAP kinase in inflammation, we demonstrate that loss of RhoB impairs TNFα-induced ICAM-1 expression and reduces cell production of IL6 and IL8. In addition, we show that RhoB silencing alters the intracellular traffic of TNFα after endocytosis. Since RhoB is a known regulator of the intracellular traffic of membrane receptors, our data suggest that RhoB controls TNFα signaling through the regulation of the TNFR traffic.

Abstract LB-352: DUSP26 inhibition: a new therapeutic pathway in neuroblastoma.

Abstract Introduction: Dual specificity phosphatase 26 (DUSP26) is overexpressed in high risk neuroblastoma (NB) and contributes to chemoresistance by inhibiting p53 function. In vitro, DUSP26 has also been shown to effectively inhibit p38 MAP kinase. We hypothesize that inhibiting DUSP26 will result in decreased NB cell growth in a p53 and/or p38 mediated manner. Methods: We developed 2 small hairpin RNA's (shRNA) against DUSP26 and stably transduced them into NB cell lines. With these cell lines, we studied the effects of DUSP26 knockdown on anchorage independent growth, cell cycle, and in vivo tumor growth using an orthotopic NB mouse model. We then tested the effects of a known DUSP26 inhibitor, NSC-87877 (NSC), on NB cytotoxicity, in combination with doxorubicin and SB203580 (p38 inhibitor), and in vivo as a single agent. Results: Both sh-RNA sequences achieved >80% knockdown of DUSP26 expression. Colony formation in soft agar was significantly reduced (p<0.05) in the sh-DUSP26 cell lines compared to control cell lines. Cell cycle analysis was also performed comparing the sh-Control and sh-DUSP26 cell lines that showed a G1 cell cycle arrest (46.8% vs. 80.6%, p<0.03). In vivo, SH-SY5Y cells transduced with sh-DUSP26 had significantly lower tumor weights after 4 weeks of growth compared to the sh-Control transduced cells (1.8gm vs. 4.2gm, p<0.01). Cytotoxicity assays testing NSC were performed on multiple NB cell lines. After 24 to 48 hours of exposure, NSC caused significantly decreased (p<0.005) cell viability at 10µM in IMR32 (18% viable), NB-19 (40%), LAN-1 (41%), SH-SY5Y (53%), and SK-N-SH (38 %). In contrast, the SHEP cell line, known to have very little DUSP26 expression, actually grew in the setting of NSC treatment (200%, p<0.005). In combination with doxorubicin (0.025 µM), NSC resulted in a significant additive decrease (p<0.05) in cell viability for the MYCN amplified cell lines (IMR-32, 26% less; LAN-1, 24%; NB-19, 20%). The p53 pathway was evaluated by Western blot using both DUSP26 inhibition techniques, shRNA and NSC. Increased p53 stability and phosphorylation at Ser46 demonstrated a possible role for p38 MAP kinase. With NSC treatment, cells displayed increased p38 phosphorylation and PARP cleavage. This was further supported with the reversal of NSC-mediated cytotoxity (p<0.01) by the p38 inhibitor SB203580 on IMR32 (54% vs. 124% viability) and NB-19 (69% vs. 102% viability). After 2 weeks of tumor growth in vivo, NSC was administered to mice once daily at 15mg/kg for a total of 21 days. Tumor weights were significantly decreased (p<0.03) in the treated cohort of mice (0.1 gm, n=3) compared to placebo control (0.8 gm, n=2). Conclusion: DUSP26 is an essential protein for NB cell growth in vitro and in vivo. By inhibiting DUSP26, pro-apoptotic pathways, such as p53 and/or p38, potentially become disinhibited. DUSP26 targeted therapy may prove to be a useful single agent or adjuvant to current chemotherapy for NB. Citation Format: Irene T. Ma, Yihui Fan, Roma H. Patel, Jin Cheng, Eugene S. Kim, Jason M. Shohet, Jianhua Yang, Sanjeev A. Vasudevan. DUSP26 inhibition: a new therapeutic pathway in neuroblastoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr LB-352. doi:10.1158/1538-7445.AM2013-LB-352

Involvement of p38 MAP kinase, but not ERK‐1/2, in histamine‐induced endothelial actin reorganization and barrier disruption

We tested the hypothesis that histamine disrupts the endothelial barrier by activation of the p38 MAP kinase and ERK‐1/2. Transendothelial electrical resistance (TER) of human umbilical vein endothelial cells (HUVEC) grown on small gold electrodes served as an index of barrier function, before and after addition of 10 μM histamine. The role of p38 MAP kinase was tested with 6 μM SB203580 and ERK‐1/2 with 10 μM PD98059 or 1 μM U0126. Specific, dual phosphorylation of p38 MAP kinase or ERK‐1/2 on their activation sites was detected by Western blotting. Dynamics of GFP‐actin and VE‐cadherin‐GFP expressed in HUVEC were also evaluated. The results show that histamine increased phosphorylation of both p38 MAP kinase and ERK‐1/2, which was blocked by pretreatment with SB203580, or PD98059/U0126, respectively. Histamine‐induced decreases in TER were inhibited by SB203580, but not affected by PD98059 or U0126 pretreatment. Histamine did not change VE‐cadherin‐GFP organization, but did briefly stop GFP‐actin‐rich edge protrusions. However, these protrusions were not decreased by histamine when the cells were pretreated with SB203580. The data suggest that p38 MAP kinase, but not ERK‐1/2, mediates histamine‐induced endothelial barrier disruption by reducing endothelial cell spreading motions. Supported by NIH grants R01HL098215 and P20GM103424, NSF grant HRD0928797, and the Louisiana Board of Regents Support Fund.

Geraniol attenuates 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced oxidative stress and inflammation in mouse skin: Possible role of p38 MAP Kinase and NF-κB

Abnormal production of reactive oxygen species (ROS) and proinflammatory cytokines often act as trigger for development of most of the chronic human diseases including cancer via up-regulation of transcription factors and activation of MAP kinases. We investigated the protective effects of geraniol (GOH) against 12-O-tetradecanoyl phorbol-13-acetate (TPA) induced oxidative and inflammatory responses, expression of p38MAPK, NF-κB and COX-2 in mouse skin. Animals were divided into four groups I-IV (n=6). Group II and III received topical application of TPA at the dose of 10nmol/0.2ml of acetone/animal/day, for two days. Group III was pre-treated with GOH (250μg) topically 30min prior to each TPA administration. While group I and IV were given acetone (0.2ml) and GOH respectively. Our results show that GOH significantly inhibited TPA induced lipid peroxidation (LPO), inflammatory responses, proinflammatory cytokine release, up regulates reduced glutathione (GSH) content and the activity of different antioxidant enzymes. Interestingly, GOH also inhibited TPA induced altered activity of p38MAPK. Further, TPA induced altered expression of NF-κB (p65) and COX-2 was also attenuated by GOH. Thus, our results suggest that GOH attenuates early tumor promotional changes, and it may serve as one of the various ways to prevent carcinogenesis.

Analysis of the role of p38 MAP kinase in epidermal growth factor-induced JB6 Cl41 cell transformation by cDNA array

To further explore the mechanism of p38 MAP kinase in regulation of JB6 Cl41 cell transformation. cDNA array was employed to scan the differential expression genes between DN-p38 cells and CMV-neo JB6 Cl41 cells after EGF stimuli. We found that up-expression genes including oncogenes and tumor suppressor genes, p53-associated protein, transcription repressors, apoptosis-associated genes, and growth arrest and DNA damage-inducible protein 153 were detected in DN-p38 cells, but low expression in CMV-neo JB6 Cl41 cells after EGF treatment. Meanwhile, some proto-oncogenes, such as c-Myc, and signal transducer and activator of transcription 1 (STAT1) were lowly expressed in EGF-stimulated DN-p38 cells, but had relatively high expression level in CMV-neo JB6 Cl41 cells under the same stimuli. Four of the differential expression genes were further confirmed by quantitative RT-PCR analysis. Our results indicate that p38 MAP kinase is involved in EGF-induced JB6 Cl41 cell transformation through effecting on more genes expression levels including transcription factors, proto-oncogene, apoptosis-related genes and growth arrest genes.

1449 P38 MAP KINASE ASSOCIATES WITH ANDROGEN RECEPTOR (AR) AND MODULATES AR SIGNALING IN PROSTATE CANCER CELLS

You have accessJournal of UrologyProstate Cancer: Basic Research1 Apr 20111449 P38 MAP KINASE ASSOCIATES WITH ANDROGEN RECEPTOR (AR) AND MODULATES AR SIGNALING IN PROSTATE CANCER CELLS Sweaty Koul, Douglas Lim, Fernando J. Kim, Randall B. Meacham, and Hari Koul Sweaty KoulSweaty Koul Aurora, CO More articles by this author , Douglas LimDouglas Lim Aurora, CO More articles by this author , Fernando J. KimFernando J. Kim Aurora, CO More articles by this author , Randall B. MeachamRandall B. Meacham Aurora, CO More articles by this author , and Hari KoulHari Koul Aurora, CO More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.1362AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES p38 MAP kinase signal transduction pathway is activated by various forms of cellular stress. While, in many tissues activation of p38 MAP kinase is associated with apoptosis, in some tissues p38 activation is critical for survival. Indeed activation of p38 MAP kinase pathway has been shown in several tumor types, but the role of p38 MAP kinase to prostate cancer biology and castrate resistant PCa has not been clearly delineated. Present studies aimed at understanding the role of p38 MAP kinase pathway in prostate cancer with special emphasis on Androgen Receptor signaling. METHODS LNCaP cells, were grown at high densities in RPMI medium supplemented with 10% Fetal Calf Serum, 1mM Sodium Pyruvate and antibiotics and maintained at 37 °C in 5% CO2 incubator. Western blot analysis was used to measure amount and activity of p38 MAP kinase, and Luciferase reporter assays were used to determine the activity of Androgen Receptor proteins. Immuno precipitation and Western blot combination was used to probe interaction of AR with p38 MAP kinase. Chemical inhibitor, dominant negative expression and shRNA to p38 MAP kinase was used to regulate p38 MAP kinase activity and levels. Well established protocols in the lab probed the effects of these treatments on cell viability, invasion, clonogenic activity and invasive behavior of the cells. RESULTS We observed that suppression of p38 MAP kinase activity by a variety of approaches decreased viability and growth of LNCaP cells in culture. We also observed that androgen depletion was associated with increased activation of p38 MAP kinase activity and inhibition of p38 MAP kinase sensitized these cells to androgen depletion. Our results show that activation of p38 MAP kinase during androgen depletion was associated with androgen independent AR activation. Results of IP experiments show for the first time that AR co-localized withp38 MAP kinase and finally and most surprisingly, stable suppression of p38 MAP kinase using Lentiviral shRNA approach resulted in complete loss of AR activity and AR protein levels. CONCLUSIONS These data show, for the first time, critical requirement for p38 MAP kinase in androgen receptor signaling in prostate cancer cells; and show for the first time, in any cell type, the regulation of AR expression by p38 MAP kinase signaling pathway. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e581 Peer Review Report Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Sweaty Koul Aurora, CO More articles by this author Douglas Lim Aurora, CO More articles by this author Fernando J. Kim Aurora, CO More articles by this author Randall B. Meacham Aurora, CO More articles by this author Hari Koul Aurora, CO More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

The Involvement of Smac/DIABLO, p53, NF-kB, and MAPK Pathways in Apoptosis of Keratinocytes from Perilesional Vitiligo Skin: Protective Effects of Curcumin and Capsaicin

Oxidative stress has been suggested as the initial pathogenetic event in melanocyte degeneration in vitiligo. Our previous results indicate that keratinocytes from perilesional skin show the features of damaged cells. In the present study, biopsies were taken from the perilesional skin of 12 patients suffering from nonsegmental vitiligo. The intracellular pathways involved in keratinocyte damage and apoptosis and the antioxidant protection of curcumin and capsaicin in these cells were investigated. In keratinocytes from perilesional vitiligo skin, we observed high levels of activated p38, NF-kB p65 subunit, p53, and Smac/DIABLO proteins. In contrast, low levels of ERK phosphorylation were present. To investigate the relationship between these pathways, we used specific inhibitors and evaluated the alteration of each pathway. For the first time, our study demonstrates the pivotal role of p38 MAP kinase as an upstream signal of perilesional keratinocyte damage, and the important contribution of p38 and NF-kB on p53 accumulation. Curcumin and capsaicin also increase ERK phosphorylation, thus inhibiting apoptosis. Moreover, pretreatment with such natural antioxidants inhibited caspase activation, increased total antioxidant capacity, repressed intracellular ROS generation and lipid peroxidation, and improved mitochondrial activity. These results suggest that antioxidants might represent an alternative approach to protect against vitiligo progression.

A Theoretical Investigation on the Pleiotropic Effects of Statins as p38 MAP Kinase Ligands

Pleiotropic effects of therapeutic statins have been frequently reported. The role of p38 MAP kinase has been advanced. In this paper, atorvastatin (containing a terphenylic system), rosuvastatin and fluvastatin, which are important drugs of the statin class, were docked into the active site of p38 MAP kinase where the selective and highly potent inhibitor SB 203580, a fluoro-phenyl pyridinylimidazole derivative, possessing a terphenylic system, binds. The protein-statin complexes near the ATP binding pocket reinforce the hypothesis of p38 MAP kinase as one possible molecular target for statin drugs. Keywords: Molecular docking, p38 MAP kinase, Pleiotropic effects, Statins

Modulatory effect of interleukin-1α on expression of structural matrix proteins, MMPs and TIMPs in human cardiac myofibroblasts: Role of p38 MAP kinase

The proinflammatory cytokine interleukin-1 (IL-1) elicits catabolic effects on the myocardial extracellular matrix (ECM) early after myocardial infarction but there is little understanding of its direct effects on cardiac myofibroblasts (CMF), or the role of p38 mitogen-activated protein kinase (MAPK). We used a focused RT-PCR microarray to investigate the effects of IL-1α on expression of 41 ECM genes in CMF cultured from different patients, and explored regulation by p38 MAPK.IL-1α (10ng/ml, 6h) had minimal effect on mRNA expression of structural ECM proteins, including collagens, laminins, fibronectin and vitronectin. However, it induced marked increases in expression of specific ECM proteases, including matrix metalloproteinases MMP-1 (collagenase-1), MMP-3 (stromelysin-1), MMP-9 (gelatinase-B) and MMP-10 (stromelysin-2). Conversely, IL-1α reduced mRNA and protein expression of ADAMTS1, a metalloproteinase that suppresses neovascularization. IL-1α increased expression of TIMP-1 slightly, but not TIMP-2. Data for MMP-1, MMP-2, MMP-3, MMP-9, MMP-10 and ADAMTS1 were confirmed by quantitative real-time RT-PCR. Tumor necrosis factor-alpha (TNFα), another important myocardial proinflammatory cytokine, did not alter expression of these metalloproteinases. IL-1α strongly activated the p38 MAPK pathway in human CMF. Pharmacological inhibitors of p38-α/β (SB203580) or p38-α/β/γ/δ (BIRB-0796) reduced MMP-3 and ADAMTS1 mRNA expression, but neither inhibitor affected MMP-9 levels. MMP-1 and MMP-10 expression were inhibited by BIRB-0796 but not SB203580, suggesting roles for p38-γ/δ.In summary, IL-1α induces a distinct pattern of ECM protein and protease expression in human CMF, in part regulated by distinct p38 MAPK subtypes, affirming the key role of IL-1α and CMF in post-infarction cardiac remodeling.

Evaluating the role of p38 MAP kinase in growth of Werner syndrome fibroblasts

The accelerated aging of Werner syndrome (WS) fibroblasts can be prevented by treatment with the p38 kinase inhibitor SB203580. If accelerated cellular senescence underlies the premature ageing features seen in this human aging model, then p38 inhibitors may have therapeutic potential in WS. However, SB203580 can inhibit in vitro several kinases that are involved in control of cellular growth, in particular, c-Raf1, CK1, and RIPK2. Thus, a better understanding of the role of this inhibitor in WS cells is required. Here we use a combination of kinase inhibitors and small intefering RNA-induced gene knockdown to show that it is inhibition of the stress-induced p38 MAP kinase that is the most plausible explanation for the effects of SB203580 on the growth of WS cells. As the development of highly selective p38 inhibitors with low toxicity is a major effort of the pharmaceuticals sector, these studies help pave the way for possible therapeutics for WS.

Induction of interleukin‐10 expression through Fcα receptor in human monocytes and monocyte‐derived dendritic cells: role of p38 MAPKinase

As previously reported by others for immunoglobulin (Ig)G, we observed that IgA can induce interleukin (IL)-10 expression in human monocytes. In this study, we explored the molecular mechanisms of IL-10 induction by IgA in monocytes and monocyte-derived dendritic cells (MD-DCs). Monomeric IgA induced IL-10 production in monocytes and this production was further increased upon IgA cross-linking. Similar IL-10 responses were observed in monocytes and autologous MD-DCs, and were inhibited (by approximately 77%) by preincubation with a blocking mAb to FcalphaRI. IL-10 induction by IgA correlated with activation of MAPKinases ERK1/2, p38 and JNK, whereas only p38-inhibitor SB-203580 inhibited IL-10 induction. Upon IgA stimulation, AP-1, NFkappaB and Sp1 transcription factors were activated and inhibitors of NFkappaB and of Sp1 suppressed IgA-driven transcriptional activation of IL-10. In addition, p38 MAPK activation appeared that it was required to control nuclear translocation of NFkappaB and Sp1 upon IgA stimulation. Therefore, in human monocytes and MD-DCs the mechanisms of IL-10 induction by IgA involve p38 MAPK-dependent recruitment of both NFkappaB and Sp1.

Molecular determinants of the antitumor effects of trichostatin A in pancreatic cancer cells

To gain molecular insights into the action of the histone deacetylase inhibitor (HDACI) trichostatin-A (TSA) in pancreatic cancer (PC) cells. Three PC cell lines, BxPC-3, AsPC-1 and CAPAN-1, were treated with various concentrations of TSA for defined periods of time. DNA synthesis was assessed by measuring the incorporation of 5-bromo-2'-deoxyuridine. Gene expression at the level of mRNA was quantified by real-time polymerase chain reaction. Expression and phosphorylation of proteins was monitored by immunoblotting, applying an infrared imaging technology. To study the role of p38 MAP kinase, the specific enzyme inhibitor SB202190 and an inactive control substance, SB202474, were employed. TSA most efficiently inhibited BrdU incorporation in BxPC-3 cells, while CAPAN-1 cells displayed the lowest and AsPC-1 cells an intermediate sensitivity. The biological response of the cell lines correlated with the increase of histone H3 acetylation after TSA application. In BxPC-3 cells (which are wild-type for KRAS), TSA strongly inhibited phosphorylation of ERK 1/2 and AKT. In contrast, activities of ERK and AKT in AsPC-1 and CAPAN-1 cells (both expressing oncogenic KRAS) were not or were only modestly affected by TSA treatment. In all three cell lines, but most pronounced in BxPC-3 cells, TSA exposure induced an activation of the MAP kinase p38. Inhibition of p38 by SB202190 slightly but significantly diminished the antiproliferative effect of TSA in BxPC-3 cells. Interestingly, only BxPC-3 cells responded to TSA treatment by a significant increase of the mRNA levels of bax, a pro-apoptotic member of the BCL gene family. Finally, in BxPC-3 and AsPC-1 cells, but not in the cell line CAPAN-1, significantly higher levels of the cell cycle inhibitor protein p21(Waf1) were observed after TSA application. The biological effect of TSA in PC cells correlates with the increase of acetyl-H3, p21(Waf1), phospho-p38 and bax levels, and the decrease of phospho-ERK 1/2 and phospho-AKT.

Hemodynamic Responses to Prenatal Stress and/or Restraint Stress in Rats

We reported that a combination of prenatal stress (PS) and restraint stress (PS+R) resulted in echocardiographic evidence of myocardial dysfunction in rats. We now report hemodynamic measurements in PS, R, or PS+R. PS, R and PS+R blunted baseline systolic and diastolic function under anesthesia. However, no differences were observed in +dp/dt and ‐dp/dt in PS or R in awake state. In contrast, the differences in +dp/dt and ‐dp/dt persisted with the PS+R in the awake state. Attenuated responses to isoproterenol were still apparent in PS, R and PS+R. The p38 MAPK inhibitor, SB 203580, reversed the blunted +dp/dt and ‐dp/dt. These findings support a central role for p38 MAP kinase in stress‐induced reversible myocardial dysfunction.Mean±SE; * p<0.05 ,** p<0.01, control+R vs PS+R; # p<0.01, control vs PS+R