Protein P38 Mitogen-activated Protein Kinase Research Articles

Dietary soybean β‐conglycinin suppresses growth performance and inconsistently triggers apoptosis in the intestine of juvenile grass carp ( Ctenopharyngodon idella ) in association with ROS‐mediated MAPK signalling

The current study aimed to investigate the effects of dietary soybean beta-conglycinin on growth performance and intestine apoptosis in juvenile grass carp (Ctenopharyngodon idella). For fish fed with the 80 g beta-conglycinin/kg diet for 7 weeks, the specific growth rate and feed intake were decreased. In the proximal intestine, dietary beta-conglycinin did not induce DNA fragmentation, tended to decrease the reactive oxygen species (ROS) content, and decreased ROS-generating enzyme (NADPH oxidase [NOX]) activity. Subsequently, in the mid-intestine, dietary beta-conglycinin caused DNA fragmentation, tended to increase the ROS content, increased caspase-3, caspase-8 and caspase-9 activities, upregulated the mRNA levels of proapoptotic molecules (apoptotic protease-activating factor-1 [Apaf1] and Bcl-2-associated X protein [BAX]) and mitogen-activated protein kinase (MAPK)-related signal molecules (Jun N-terminal kinase (JNK) and p38 MAPK) and increased the protein levels of p38 MAPK and phospho-p38 MAPK. Moreover, in the distal intestine, dietary beta-conglycinin induced DNA fragmentation, elevated NOX activity and the ROS content and increased caspase-3, caspase-8 and caspase-9 activities, death ligand (TNF-alpha) mRNA expression level, and p38 MAPK and phospho-p38 MAPK protein levels. In summary, dietary soybean beta-conglycinin suppressed fish growth and inconsistently caused apoptosis among the different intestinal segments which was partially associated with ROS-mediated MAPK signalling.

The effect of a high fat diet in the regulation of MAP kinase in left ventricle fibrosis

IntroductionIt is well known that obesity contributes to the development of systemic inflammatory responses, which in turn may be involved in the process of interstitial fibrosis and left ventricular (LV) remodeling. Activation of proinflammatory factors such as transforming growth factor β (TGF‐β) can directly stimulate mitogen‐activated protein kinase (MAPK) p38 and JNK.PurposeThe aim of this study was to evaluate the level of TGF‐β, MAPK p38 and JNK in the LV in Sprague Dawley (SPRD) rats maintained on a high fat diet (HFD).MethodsThe SPRD rats were maintained on a normal fat diet (NFD) or on a HFD from 4 weeks of age for 12 weeks (NFD 16‐week‐old rats, NFD 16‐wk; or HFD 16‐week‐old rats, HFD 16‐wk) or for 16 weeks (NFD 20‐week‐old rats, NFD 20‐wk; or HFD 20‐week‐old rats, HFD 20‐wk). At the end of the experiment, blood and LV were collected from all rats for further analysis (Real‐Time PCR, histopathological and immunohistochemical analysis). Consent was obtained from The Local Animal Research Ethics Committee (33/2011).ResultsThe LV fibrosis area in relation to the entire LV area was significantly higher in the HFD 16‐wk rats and the HFD 20‐wk rats compared with the NFD 16‐wk rats and the NFD 20‐wk rats, respectively. TGF‐β mRNA expression did not differ between the study groups of rats. However, p38 MAPK mRNA expression and c‐jun mRNA expression were significantly lower in the HFD 20‐wk rats than in the HFD 16‐wk rats. TGF‐β type II receptor (TβRII) protein demonstrated only cytoplasmic reactivity, while p38 MAPK protein and c‐jun protein showed both nuclear and cytoplasmic reactivity. The p38 MAPK nuclear reaction did not differ significantly between the study groups of rats. In contrast, the cytoplasmic response of the p38 MAPK was dependent on the administration of both HFD in two time intervals to the groups of rats. Significant differences in the nuclear reaction for c‐jun were observed between the exanimated groups of rats. The c‐jun nuclear expression was significantly lower in the HFD 20‐wk rats than in the HFD 16‐wk rats.ConclusionA high fat diet caused an increase in LV fibrosis in SPRD rats. Additionally, extension by the next 4 weeks the duration of HFD intake did not increase the LV fibrosis area. Moreover, high fat diet in both intervals significantly influences the expression of p38 MAPK and JNK in the LV. However, demonstrating their potential involvement in the processes of interstitial myocardial fibrosis and left ventricular remodeling requires further research.Support or Funding InformationThis study was financially supported by statutory funds from the Medical University of Warsaw (1MA/N/2018).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

The role of high fat diet in the regulation of MAP kinases activity in left ventricular fibrosis

It is well known that obesity contributes to the development of systemic inflammatory responses, which in turn may be involved in the process of interstitial fibrosis and left ventricular (LV) remodelling. Activation of pro-inflammatory factors such as transforming growth factor β (TGF-β) can directly stimulate mitogen-activated protein kinase (MAPK) p38 and JNK. The aim of the study was to evaluate the level of TGF-β and MAPK p38 and JNK in the LV in Sprague Dawley (SPRD) rats maintained on a high fat diet (HFD).The SPRD rats from 4 weeks of age were on a normal fat diet (NFD) or a HFD for 12 weeks (NFD-16-week-old rats, NFD 16-wk; or HFD-16-week-old rats, HFD 16-wk) or 16 weeks (NFD-20-week-old rats, NFD 20-wk; or HFD-20-week-old rats, HFD 20-wk). At the end of the experiment, blood and LV were collected from all rats for further analysis (biochemical, Real Time PCR and immunohistochemical analysis). TGF-β mRNA expression did not differ between the study groups of rats. However, p38 MAPK mRNA expression was significantly lower in the HFD 20-wk rats than in both the HFD 16-wk rats and the NFD 20-wk rats. c-jun mRNA expression was significantly higher in the HFD 16-wk rats than in the NFD 16-wk rats. There was significantly lower expression of c-jun mRNA in the HFD 20-wk rats and in the NFD 20-wk rats than in the HFD 16-wk rats and in the NFD 16-wk rats, respectively. TGF-β type II receptor (TβRII) protein demonstrated only cytoplasmic reactivity, while p38 MAPK protein and c-jun protein showed both nuclear and cytoplasmic reactivity. The results suggest that a high fat diet and in two time intervals significantly influence the expression of p38 MAPK and JNK in the LV. However, demonstrating their potential involvement in the processes of interstitial myocardial fibrosis and left ventricular remodeling requires further research.

Transcriptomic analysis reveals that enterovirus F strain SWUN-AB001 infection activates JNK/SAPK and p38 MAPK signaling pathways in MDBK cells

BackgroundEnteroviruses (Picornaviridae family) have been widely detected in the feces from cattle with diarrhea. However, the mechanisms responsible for the pathogenicity of enteroviruses in cattle remain unclear. Recently, we isolated a novel EV-F7 strain called SWUN-AB001 from diarrheal yak (Bos grunniens) feces. To explore the pathogenic mechanisms of this novel virus, we used a transcriptomics approach to find genes with differential expression patterns in Madin-Darby bovine kidney (MDBK) cells during infection with SWUN-AB001 over time.ResultsMDBK cells were sampled at 12 and 24 h post-infection (hpi) to represent the early and late stages of a SWUN-AB001 infection. Compared with the non-infected cells, 19 and 1050 differentially expressed genes (DEGs) were identified at 12 and 24 hpi, respectively. These DEGs were associated with disease, signal transduction, cellular process and cytokine signaling categories. At 24 hpi, the pathway enrichment analysis revealed that signal pathways such as c-Jun NH2-terminal kinase/ stress-activated protein kinase (JNK/SAPK) and mitogen-activated protein kinase (MAPK) pathways and cytokine-cytokine receptor interactions were associated with the interactions occurring between EV-F7 and MDBK cells. Our additional western blot analysis showed that the phosphorylation levels of JNK/SAPK and p38 MAPK proteins increased significantly in the MDBK cells at 24 hpi. The result indicated that infection with EV-F7 could activate JNK/SAPK and p38 MAPK pathways in MDBK cells, and possibly trigger large-scale cytokine production.ConclusionOur transcriptome analysis provides useful initial data towards better understanding of the infection mechanisms used by EV-F7, while highlighting the potential molecular relationships occurring between the virus and the host’s cellular components.

Activation of c-Jun N-terminal kinase and p38 after cerebral ischemia upregulates cerebral sodium-glucose transporter type 1

Cerebral ischemic stress increases cerebral sodium-glucose transporter type 1 (SGLT-1). However, the mechanism by which cerebral ischemia leads to the up-regulation of SGLT-1 remains unclear. In peripheral tissue, the activation of mitogen-activated protein kinases (MAPKs) increases SGLT-1. MAPK pathways [c-Jun N-terminal kinase (JNK), p38 MAPK, and extracellular signal-regulated protein kinase (ERK)] are activated by cerebral ischemic stress. Therefore, we confirmed the involvement of MAPKs in the up-regulation of cerebral SGLT-1 after cerebral ischemia. Male ddY mice were subjected to middle cerebral artery occlusion (MCAO). Protein expression was assessed by western blotting. Mice received an intracerebroventricular (i.c.v.) injection of SP600125 (JNK inhibitor), SB203580 (p38 inhibitor), and PD98059 (MEK inhibitor) immediately after reperfusion. The infarction and behavioral abnormalities were assessed on days 1 and 3 after MCAO. The MAPK inhibitors suppressed the activation of JNK, p38, and ERK 3 h after MCAO. SP600125 and SB203580 administration ameliorated cerebral ischemic neuronal damage, whereas PD98059 administration exacerbated cerebral ischemic neuronal damage. SP600125 and SB203580 significantly suppressed the increase in SGLT-1 12 h after MCAO. PD98059 had no effect on SGLT-1 expression after MCAO. Our results indicate that the activation of JNK and p38 participate in the up-regulation of cerebral SGLT-1 after MCAO.

Continuous Blood Purification Ameliorates Multiple Organ Failure Through Inhibiting the Activation of the P38 MAPK Signaling Pathway in a Rat Model

Background/Aims: Multiple organ failure (MOF) is a primary threat to the survival of patients with systemic inflammation. Blood purification is employed in the treatment of MOF, as an artificial kidney or artificial liver. This study focuses on the effects of continuous blood purification (CBP) on ameliorating MOF through regulating the p38 mitogen-activated protein kinase (MAPK) signaling pathway in a rat model. Methods: A rat model of MOF was successfully established by endotoxin injection after hemorrhagic shock resuscitation. The mRNA expressions of inducible nitric oxide synthase (iNOS) and p38 MAPK of liver, kidney, and lung tissues in each group were measured by RT-qPCR at each measuring time point. To evaluate the activation of p38 MAPK signaling pathway, protein levels of phosphorylated p38 (p-p38) MAPK and p38 MAPK was measured by western blot analysis. The serum levels of nitric oxide and TNF-α were determined. Results: After CBP treatment, the levels of SGPT, SGOT, Cr, and BUN were significantly declined, while the PaO₂ value was increased. Expressions of p38 MAPK mRNA, iNOS mRNA, p-p38 MAPK protein and p38 MAPK protein, and nitric oxide and TNF-α levels were markedly elevated in MOF, an effect blunted by CPB. Meanwhile, pathological sections of liver, kidney, and lung tissues after CPB treatment ameliorated swelling and inflammation. Conclusion: Our study proved that CBP could downregulate the p38 MAPK signaling pathway, suppress iNOS expression, reduced the serum levels of nitric oxide and TNF-α, thus ameliorate symptom of MOF.

Resveratrol protects neuronal cells from isoflurane-induced inflammation and oxidative stress-associated death by attenuating apoptosis via Akt/p38 MAPK signaling.

The aim of the present study was to determine whether resveratrol protects neuronal cells from inflammation and isoflurane-induced oxidative stress-associated death via attenuating apoptosis via Akt/p38 mitogen-activated protein kinase (MAPK) signaling. The PC12 rat pheochromocytoma cell line was treated with 2% isoflurane + 21% O2 + 5% CO2 for 6 h and pre-treated with resveratrol (0-1,000 µM) for 0, 24 or 48 h prior to isoflurane treatment. An MTT assay, flow cytometry and ELISA of tumor necrosis factor-α, interleukin-6, malondialdehyde and superoxide dismutase revealed that resveratrol reduced growth inhibition, restrained apoptosis and suppressed inflammation and oxidative stress induced by isoflurane in PC12 cells. Pretreatment with resveratrol effectively reduced caspase-3 activity and inducible nitric oxide synthase protein expression in isoflurane-induced PC12 cells. In addition, western blot analysis demonstrated that resveratrol treatment significantly attenuated isoflurane-induced decreases in the activated phosphorylated (p)-Akt/Akt ratio and increases in the p-p38/p38 MAPK protein ratio in PC12 cells. These findings indicated that resveratrol was able to protect neuronal cells from isoflurane-induced inflammation and oxidative stress-associated death by attenuating apoptosis via Akt/p38 MAPK signaling.

Protective effects of probucol on Ox-LDL-induced epithelial-mesenchymal transition in human renal proximal tubular epithelial cells via LOX‑1/ROS/MAPK signaling.

Oxidized low-density lipoprotein (Ox-LDL), as a strong oxidant, results in renal injury through multiple mechanisms. The aim of the present study was to determine the injury effects of Ox‑LDL and the potential protective effects of the antioxidant reagent probucol on epithelial‑mesenchymal transition (EMT) in human renal proximal tubular epithelial cells (HK‑2) and to further explore the role and interrelation of lectin‑like oxidized low‑density lipoprotein receptor‑1 (LOX‑1), reactive oxygen species (ROS) and mitogen‑activated protein kinase (MAPK) pathway. In the present study, concentrations of 0‑100µg/ml Ox‑LDL were used to induce HK‑2 cell EMT. Then, probucol (20µmol/l) and the LOX‑1 inhibitor, polyinosinic acid (250µg/ml), were also used to pretreat HK‑2 cells. Intracellular ROS activity was evaluated using the specific probe 2',7'‑dichlorodihydrofluorescein diacetate (DCFH‑DA). Concentration of nitric oxide (NO) was determined using a biochemical colorimetric method. Expression of E‑cadherin, α‑smooth muscle actin (SMA), LOX‑1, NADPH oxidase 4 (NOX4), cytochrome b‑245 α chain (p22phox), extracellular signal‑regulated kinase (ERK), and p38 MAPK protein levels were examined by western blotting. The results revealed that Ox‑LDL induced the expression of LOX‑1 and α‑SMA and reduced the expression of E‑cadherin in a dose‑dependent manner, and these effects were inhibited by polyinosinic acid or probucol pretreatment. Stimulation with 50µg/ml Ox‑LDL induced the expression of NOX4 and p22phox and increased intracellular ROS activity, but NO production in the cell supernatants was not affected. The Ox‑LDL‑mediated increases in Nox4 and p22phox expression and in ROS activity were inhibited by probucol pretreatment. Further investigations into the underlying molecular pathways demonstrated that ERK and p38 MAPK were activated by Ox‑LDL stimulation and then inhibited by probucol pretreatment. The findings of the present study therefore suggest that Ox‑LDL induced EMT in HK‑2 cells, the mechanism of which may be associated with LOX‑1‑related oxidative stress via the ERK and p38 MAPK pathways. Notably, pretreatment with probucol inhibited the Ox‑LDL‑induced oxidative stress by reducing the expression of LOX‑1, and blocked the progression of EMT.

Atorvastatin inhibits cholesterol-induced caspase-3 cleavage through down-regulation of p38 and up-regulation of Bcl-2 in the rat carotid artery.

SummaryAim:Atherosclerotic lesions in the carotid arteries lead to a broad range of cerebrovascular disorders such as vascular dementia and ischaemic stroke. Recent studies have verified the beneficial role of atorvastatin (AV) in atherosclerosis. Despite a large body of studies, the mechanisms underlying this effect have not been completely explained. In this study, several experiments were performed on atherosclerotic rat models to investigate the anti-inflammatory and anti-apoptotic effect of AV in the carotid artery.Methods:In this experimental study, 40 male Wistar rats (250 ± 25 g) were randomly divided into four groups: rats on a normal diet (ND; n = 10); a high-cholesterol diet (HD; n = 10); a high-cholesterol diet plus AV (HD + AV; n = 10); and the AV control group (AV; n = 10). Cleavage of caspase-3 protein, expression of B-cell lymphoma 2 (Bcl-2) as well as phosphorylation of p38 mitogen-activated protein kinase (MAPK) were determined by immunoblotting assay in the carotid artery homogenate. Plasma atherogenic indices, including total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) were measured by colorimetric assay at the end of the experiment. Plasma levels of oxidised LDL (oxLDL) were measured by sandwich enzyme-linked immunosorbent assay (ELISA).Results:After eight weeks of feeding with a high-cholesterol diet, an elevated level of oxLDL was observed in the plasma in the HD group compared with the ND group [214.42 ± 17.46 vs 69.13 ± 9.92 mg/dl (5.55 ± 0.45 vs 1.78 ± 0.26 mmol/l); p < 0.01]. AV administration significantly reduced oxLDL levels in the HD + AV compared to the HD group [126.52 ± 9.46 vs 214.42 ± 17.46 mg/dl (3.28 ± 0.25 vs 5.55 ± 0.45 mmol/l); p < 0.01]. Results also showed that compared with the HC group, the HC + AV group had lower levels of p38 phosphorylation (p < 0.05) and higher levels of Bcl-2 expression (p < 0.05). Lower levels of cleaved caspase-3 were observed in the HC + AV group in comparison with the HC group (p < 0.05).Conclusions:The resultant data suggest that the anti-apoptotic effect of AV could be partially mediated by the pro-inflammatory protein p38 MAPK and the anti-apoptotic protein Bcl-2 in the rat carotid artery. Atorvastatin can therefore be considered a target drug in the prevention or development of atherosclerotic events.

Protective effects of taurine against renal ischemia/reperfusion injury in rats by inhibition of gelatinases, MMP-2 and MMP-9, and p38 mitogen-activated protein kinase signaling

Dysregulated expression of matrix metalloproteinases (MMPs) is closely associated with the pathogenesis of renal ischemia/reperfusion injury (I/R). The production of excessive reactive oxygen species (ROS) causes tissue damage. Increased ROS production causes activation of p38 mitogen-activated protein kinase (MAPK) signaling, which participates in gene regulation of MMPs, especially MMP-2 and MMP-9 (gelatinases). Taurine (2-aminoethanesulfonic acid) in mammalian cells functions in bile acid conjugation, maintenance of calcium homeostasis, osmoregulation, membrane stabilization, and antioxidation, antiinflammatory, and antiapoptotic action. We investigated the effects of taurine and the possible role of p38 MAPK signaling on regulation of MMP-2 and MMP-9 in a renal I/R injury model in rats. Rats were divided into three groups: sham, I/R, and I/R + taurine treated. After a right nephrectomy, I/R was induced by clamping the left renal pedicle for 1 h followed by 6 h reperfusion. Taurine was administered 45 min prior to induction of ischemia. Renal function was assessed by serum creatinine and blood urea nitrogen (BUN) levels. Tubule injury and structural changes were evaluated by light microscopy. Malondialdehyde (MDA) levels were analyzed by high performance liquid chromatography (HPLC). Superoxide dismutase (SOD) activity levels were measured using a colorimetric kit. mRNA expression of MMP-2 and MMP-9 was determined by real-time polymerase chain reaction. MMP-2 and MMP-9 activities were measured using a fluorimetric kit. Phosphorylated p38 (p-p38) and total p38 MAPK protein expressions were evaluated by western blot. Taurine pretreatment significantly attenuated renal dysfunction and histologic damage, such as renal tubule dilation and loss of brush borders. The pretreatment also decreased the MDA level and attenuated the reduction of SOD activity in the kidney during I/R. Taurine pretreatment also decreased significantly both MMP-2 and MMP-9 mRNA expression and MMP-9 activity induced by I/R. In addition, the activity of p38 MAPK signaling was down-regulated significantly by taurine administration. Inhibition of MMP-2 and MMP-9 expression and MMP-9 activity caused by taurine may be associated with suppression of p38 MAPK activation during I/R induced renal injury in rats. Therefore, taurine administration may prove to be a strategy for attenuating renal I/R injury.

Rumen-derived lipopolysaccharide provoked inflammatory injury in the liver of dairy cows fed a high-concentrate diet.

Rumen-derived lipopolysaccharide (LPS) is translocated from the rumen into the bloodstream when subacute ruminal acidosis (SARA) occurs following long-term feeding with a high-concentrate (HC) diet in dairy cows. The objective of this study was to investigate the mechanism of inflammatory responses in the liver caused by HC diet feeding. We found that SARA was induced in dairy cows when rumen pH below 5.6 lasted for at least 3 h/d with HC diet feeding. Also, the LPS levels in the portal and hepatic veins were increased significantly and hepatocytes were impaired as well as the liver function was inhibited during SARA condition. Meanwhile, the mRNA expression of immune genes including TNF receptor associated factor 6 (TRAF6), nuclear factor-kappa B (NF-κB), p38 mitogen-activated protein kinase (MAPK), extracellular regulated protein kinases (ERK) MAPK, Interleukin-1 (IL-1) and serum amyloid A (SAA) in the liver were significantly increased in SARA cows. Moreover, the phosphorylation level of NF-κB p65 and p38 MAPK proteins in the liver and the concentration of Tumor Necrosis Factor (TNF-α), Interleukin-1β (IL-1β) and Interleukin-6 (IL-6) in peripheral blood were obviously increased under SARA condition. In conclusion, the inflammatory injury in the liver caused by LPS that traveled from the digestive tract to the liver through the portal vein after feeding with a HC diet.

Effects of p38 mitogen-activated protein kinase on lung ischemia-reperfusion injury in diabetic rats

BackgroundLung ischemia-reperfusion injury (LIRI) is a pathologic process that is observed in several clinical conditions, and p38 mitogen-activated protein kinase (MAPK) is involved. Diabetes mellitus (DM) results in an increased incidence of ischemia-induced organ damage. The aims of this study were to examine the effects of DM on LIRI in a rat model of DM and to explore the possible mechanisms in relation to the p38 MAPK pathway. MethodsForty rats were randomly divided into the following five groups (n = 8 each): a control + sham group, a control + IR group (CIR), a DM + sham group, a DM + IR group (DIR), and a DM + IR + SB203580 group. The control and streptozotocin-induced diabetic rats underwent a sham operation or left hilum occlusion for 90 min followed by reperfusion for 4 h. SB203580 was used to inhibit the p38 MAPK pathway. The pulmonary oxygenation index, inflammatory cytokines in the serum, lung edema, histopathology, oxidant stress, apoptosis, and phosphorylated/total-p38 MAPK protein levels were measured. ResultsThe DIR group displayed greater concentrations of tumor necrosis factor-α, interleukin-6, and intercellular adhesion molecule-1 and increases in the wet weight-to-dry weight ratio, lung injury scores, malondialdehyde levels, and cellular apoptosis, and these effects were accompanied by lower pulmonary oxygenation compared with the CIR group (P < 0.05). In the DIR group, the expression levels of p38 MAPK protein were significantly upregulated compared with those of the CIR group. Additionally, all of these alterations were attenuated in the DM + IR + SB203580 group compared with the DIR group. ConclusionsDiabetes exacerbates LIRI by activating the p38 MAPK pathway.

Molecular Characterization and Expression Analysis of P38 MAPK Gene and Protein in Aquatic Midge, Chironomus riparius (Diptera: Chironomidae), Exposed to Environmental Contaminants.

P38 Mitogen-activated protein kinase (MAPK), an important signaling protein involved in various cellular processes, including stress responses, has been well characterized in model organisms. P38 has been identified in a number of insects, including the genus Drosophila; however, its homologue in Chironomus riparius has not yet been identified. In this study, we identified and characterized p38 MAPK (Crp38) gene in C. riparius using a transcriptome database that was previously generated 454 GS-FLX pyrosequencing. Comparative and phylogenetic analyses were performed using the p38 homologue of other species, such as Drosophila melanogaster, Aedes aegypti, Bombyx mori, Caenorhabditis elegans, Homo sapiens, etc. Furthermore, to test its potential as a biomarker of environmental contamination, Crp38 gene expression was analyzed upon exposure to nonylphenol (NP), silver nanoparticles (AgNPs), and cadmium (Cd). Crp38 gene expression was up- or down-regulated depending on the concentration and exposure duration of chemicals. These results show the role of Crp38 gene in defense against environmental stresses, as well as its potential use as a biomarker for various environmental pollutants. We further synthesized p38 antibody based on the predicted amino acid sequence deduced from Crp38 cDNA and, using this customized antibody, examined p38 protein expression in Cd exposed C. riparius. Although transcriptional alteration was not translated to the protein level, this result showed the possible application of a protein level functional study using cDNA sequence information from next-generation sequencing database in nonmodel organisms.

Effects of the MAPK pathway and the expression of CAR in a murine model of viral myocarditis.

The pathogenesis of viral myocarditis (VMC) is not fully understood. This study aimed to examine the relationship between coxsackie-adenovirus receptor (CAR) and the p38 mitogen activated protein kinase (MAPK) pathway mechanisms in a mouse model. Three groups of mice were established: 5 mice in a control group injected with saline, 15 in the model group injected with coxsackie virus B3 (CVB) and 15 in the intervention group injected with CVB3 but treated with the p38 MAPK inhibitor SB203580. Mice were sacrificed at days 1, 5, 10, 15 and 30 and cardiac tissues were isolated to perform the tests. Quantitative PCR and western blot analysis showed CAR mRNA and protein expression levels were highest in the model group at all time-points (P<0.05). The expression levels of p38 MAPK protein by western blot analysis at days 1, 5 and 10 were obviously higher in the model group (P>0.05). H&E staining used to observe myocardial pathological changes showed the inflammatory infiltration was also higher in the model group at all the time-points (P<0.05). Our results show a direct relationship between CAR and p38 MAPK levels, and since the p38 MAPK inhibitor treatment resulted in reduced levels of CAR as well as lower inflammatory infiltration, it is possible that the signaling pathway may mediate CAR expression during the pathogenesis of VMC.

Tanshinone IIA pretreatment attenuates ischemia/reperfusion-induced renal injury

Tanshinone IIA is a chemical compound extracted from the root of traditional Chinese herb Salvia miltiorrhiza Bunge. Tanshinone IIA has been suggested to possess anti-inflammatory activity and antioxidizing capability. Recently, accumulating results have indicated the antitumor activity of tanshinone IIA; thus, it has attracted increasing attention. In addition, tanshinone IIA has been indicated to attenuate ischemia/reperfusion induced renal injury (I/RIRI); however, little is known regarding the underlying mechanisms involved in this process. In the present study an I/RIRI rat model was used to analyze the effects of tanshinone IIA on myeloperoxidase (MPO), TNF-α and IL-6 activities using ELISA kits. Furthermore, macrophage migration inhibitory factor (MIF), cleaved caspase-3, B-cell lymphoma 2 (Bcl-2) and p38 mitogen-activated protein kinase (MAPK) protein expression levels were evaluated using western blot analysis. The results indicated that tanshinone IIA protected renal function in I/RIRI rats. ELISA demonstrated that tanshinone IIA significantly reduced MIF, TNF-α and IL-6 activities in I/RIRI rats. Western blot analysis showed that tanshinone IIA significantly suppressed MIF, cleaved caspase-3 and p38 MAPK protein expression levels in I/RIRI rats. The present results suggest that tanshinone IIA pretreatment attenuates I/RIRI via the downregulation of MPO expression, inflammation, MIF, cleaved caspase-3 and p38 MAPK.

Molecular Changes Involving MEK3–p38 MAPK Activation in Chronic Masticatory Myalgia

The exact mechanism underlying chronic masticatory myalgia (CMM), a conspicuous symptom in temporomandibular disorders, remains unclear. This investigation compared gene expression profiles between CMM patients and healthy subjects. Peripheral blood leukocytes were collected in 8 cases and 8 controls and subjected to whole genome microarray analyses. Data were analyzed with Gene Ontology and interactive pathways analyses. According to Gene Ontology analysis, categories such as ion transport, response to stimuli, and metabolic process were upregulated. The pathway analysis suggested overexpression of the mitogen-activated protein kinase (MAPK) pathway in CMM patients and to a higher degree in a pathway network. Overexpression of representative members of the MAPK pathway—including MAPK kinase 3 (MEK3), calcium voltage-gated channel auxiliary subunit gamma 2 (CACNG2), and growth arrest and DNA damage–inducible gamma (GADD45G)—was validated with real-time polymerase chain reaction. The upregulation of MEK3 was negatively correlated with the age of the CMM group. In the next step, the authors focused on MEK3, the gene that exhibited the greatest degree of differential expression, and its downstream target protein p38 MAPK. The results revealed upregulation of MEK3, as well as phosphorylated MEK3 and phosphorylated p38 MAPK, in CMM patients. These results provide a “fingerprint” for mechanistic studies of CMM in the future and highlight the importance of MEK3–p38 MAPK activation in CMM.

P38 MAPK Inhibitor Administration Provides Protection against Acute Lung Injury in Rats with Intestinal Ischemia Reperfusion Via Downregulating Interlukin-6 Expression

Background: Intestinal ischemia-reperfusion (II/R) can lead to indirect lung injury. Previous studies have shown that p38 kinase may be involved in acute lung injury (ALI). However, the exact role of p38 mitogen-activated protein kinase (MAPK) and possible mechanisms in lungs are unknown and no specific treatment exists for II/R-induced ALI. The purpose of this study was to investigate the protective role of p38 MAPK inhibition in the regulation of the inflammatory response to ALI induced by II/R in rats.Methods: II/R was induced by occlusion of the superior mesenteric artery (SMA) and coeliac artery (CA) for 40 minutes and subsequent reperfusion for 0, 8, 16, 24 hours. SB239063, a specific inhibitor of p38 MAPK, was injected (10 mg/kg) intraperitoneally 60 minutes before the operation. The severity of ALI was measured by histology analysis (hematoxylin-eosin [HE] staining and ALI scoring) and lung edema (lung wet/dry weight ratio). Quantitative polymerase chain reaction (qPCR), western blot (WB), and immunofluorescent staining were employed to assess expression and location of interleukin-6 (IL-6) and p38 MAPK, respectively.Results: ALI including lung edema, alveolar collapse, hemorrhage, exudation and infiltration of inflammatory cells in the lungs was induced by II/R and expression levels of IL-6 and p38 MAPK protein in the lung tissues were significantly increased at 8 and 16 hours after reperfusion, respectively compared with that in the sham group. SB239063 administration significantly down-regulated p38 MAPK and IL-6 concentration in the lung specimens, therefore, protected effectively lung tissues from injury after II/R.Conclusions: These findings indicated that p38 MAPK inhibition may downregulate the expression of IL-6 to protect lung from ALI induced by II/R, which could be used as a potential target for the prevention or treatment of ALI caused by II/R. Citation: Liu-Lin Xiong, Yun-Xia Zuo, Chao-Zhi Luo, Jie Yu, Qing-Jie Xia, Fei-Fei Xu, et al. p38 MAPK inhibitor administration provides protection against acute lung injury in rats with intestinal ischemia reperfusion via downregulating interlukin-6 expression. J Anesth Perioper Med 2016; 3: 27-34. doi: 10.24015/JAPM.2016.0004This is an open-access article, published by Evidence Based Communications (EBC). This work is licensed under the Creative Commons Attribution 4.0 International License, which permits unrestricted use, distribution, and reproduction in any medium or format for any lawful purpose. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Possible Mechanisms of Di(2-ethylhexyl) Phthalate-Induced MMP-2 and MMP-9 Expression in A7r5 Rat Vascular Smooth Muscle Cells.

Proliferation and migration of vascular smooth muscle cells (VSMC) are important in the development and/or progression of many cardiovascular diseases, including atherosclerosis. Evidence shows that matrix metalloproteinase (MMP)-2 and MMP-9 are related to the pathogenesis of atherosclerosis. The expressions of MMP-2 and MMP-9 in atherosclerosis are regulated via various pathways, such as p38 mitogen activated protein kinase (MAPK), extracellular signal regulated kinase 1 and 2 (ERK1/2), Akt, and nuclear factor kappa (NF-κB). Di(2-ethylhexyl) phthalate (DEHP) has been shown to induce atherosclerosis by increasing tumor necrosis factor (TNF)-α, interleukin (IL)-6, and intercellular adhesion molecule (ICAM) productions. However, whether DEHP poses any effects on MMP-2 or MMP-9 expression in VSMC has not yet been answered. In our studies, rat aorta VSMC was treated with DEHP (between 2 and 17.5 ppm) and p38 MAPK, ERK1/2, Akt, NF-κB, and MMP-2 and MMP-9 proteins and activities were measured. Results showed that the presence of DEHP can induce higher MMP-2 and MMP-9 expression than the controls. Similar results on MMP-regulating proteins, i.e., p38 MAPK, ERK1/2, Akt, and NF-κB, were also observed. In summary, our current results have showed that DEHP can be a potent inducer of atherosclerosis by increasing MMP-2 and MMP-9 expression at least through the regulations of p38 MAPK, ERK1/2, Akt, and NF-κB.

Lapatinib increases motility of triple-negative breast cancer cells by decreasing miRNA-7 and inducing Raf-1/MAPK-dependent interleukin-6.

Lapatinib, a dual epidermal growth factor receptor (EGFR) and HER2 tyrosine kinase inhibitor (TKI), has been approved for HER2-positive breast cancer patients. Nevertheless, its inhibitory effect on EGFR did not deliver clinical benefits for triple-negative breast cancer (TNBC) patients even EGFR overexpression was frequently found in this disease. Moreover, lapatinib was unexpectedly found to enhance metastasis of TNBC cells, but the underlying mechanisms are not fully understood. In this study, we explored that the level of interleukin-6 (IL-6) was elevated in lapatinib-treated TNBC cells. Treatment with IL-6 antibody abolished the lapatinib-induced migration. Mechanistically, the signaling axis of Raf-1/mitogen-activated protein kinases (MAPKs), c-Jun N-terminal kinases (JNKs), p38 MAPK, and activator protein 1 (AP-1) was activated in response to lapatinib treatment to induce IL-6 expression. Furthermore, our data showed that microRNA-7 directly binds and inhibits Raf-1 3'UTR activity, and that down-regulation of miR-7 by lapatinib contributes to the activation of Raf-1 signaling pathway and the induction of IL-6 expression. Our results not only revealed IL-6 as a key regulator of lapatinib-induced metastasis, but also explored the requirement of miR7/Raf-1/MAPK/AP-1 axis in lapatinib-induced IL-6 expression.

Excessive L-cysteine induces vacuole-like cell death by activating endoplasmic reticulum stress and mitogen-activated protein kinase signaling in intestinal porcine epithelial cells.

High intake of dietary cysteine is extremely toxic to animals and the underlying mechanism remains largely unknown. This study was conducted to test the hypothesis that excessive L-cysteine induces cell death by activating endoplasmic reticulum (ER) stress and mitogen-activated protein kinase (MAPK) signaling in intestinal porcine epithelial cells. Jejunal enterocytes were cultured in the presence of 0-10 mmol/L L-cysteine. Cell viability, morphologic alterations, mRNA levels for genes involved in ER stress, protein abundances for glucose-regulated protein 78, C/EBP homologous protein (CHOP), alpha subunit of eukaryotic initiation factor-2 (eIF2α), extracellular signal-regulated kinase (ERK1/2), p38 MAPK, and c-Jun N-terminal protein kinase (JNK1/2) were determined. The results showed that L-cysteine (5-10 mmol/L) reduced cell viability (P < 0.05) and led to vacuole-like cell death in intestinal porcine epithelial cells. These adverse effects of L-cysteine were not affected by the autophagy inhibitor 3-methyladenine. The protein abundances for CHOP, phosphorylated (p)-eIF2α, p-JNK1/2, p-p38 MAPK, and the spliced form of XBP-1 mRNA were enhanced (P < 0.05), whereas those for p-ERK1/2 were reduced (P < 0.05). Collectively, excessive L-cysteine induces vacuole-like cell death via the activation of ER stress and MAPK signaling in small intestinal epithelial cells. These signaling pathways may be potential targets for developing effective strategies to prevent the toxicity of dietary cysteine.