Activation Of Extracellular Signal-regulated Kinase Signaling Pathway Research Articles

Anserine/Carnosine-Rich Extract from Thai Native Chicken Suppresses Melanogenesis via Activation of ERK Signaling Pathway.

Skin hyperpigmentation is an aesthetic problem that leads to psychosocial issues. Thus, skin whitening agents from agro- and poultry-industrial co-products are considered high economic value ingredients of interest for sustainable application. Therefore, this study aimed to determine the cosmeceutical potential of anserine/carnosine-rich chicken extract (ACCE) from the Thai native chicken Pradu Hang Dam Mor Kor 55 (PD) meat. The chemical composition was identified and quantified using the HPLC-UV method. Then, the antioxidation potential of the extract was compared to that of L-anserine and L-carnosine, using 1,1-diphenyl-2-picrylhydrazyl assay and shikonin-induced production of reactive oxygen species in CCD-986Sk cell models, and the anti-melanogenesis effect in the MNT-1 melanoma cell line model was investigated. Furthermore, related mechanisms were identified using colorimetric tyrosinase assay and the Western blot technique. The ACCE was composed of L-anserine and L-carnosine as two major constituents. In a dose-dependent manner, ACCE, L-anserine, and L-carnosine manifested significant antioxidation potential and significant reduction of melanin production. Activation of the extracellular signal-regulated kinase (ERK) signaling pathway and inhibition of tyrosinase activity of ACCE were demonstrated as the mechanisms of the anti-melanogenesis effect. In conclusion, ACCE has been revealed as a potential cosmeceutical agent due to its antioxidation and anti-melanogenic activity in association with L-anserine and L-carnosine composition and biomolecular regulating ability. Therefore, further studies and development should be considered to support the utilization of anserine/carnosine-rich chicken extract in the cosmetic industry for economic value creation and sustainability.

Effects of miR-490-5p targeting CDK1 on proliferation and apoptosis of colon cancer cells via ERK signaling pathway.

The aim of this study was to investigate the effects of micro ribonucleic acid (miR)-490-5p on the proliferation and apoptosis of colon cancer cells, and to explore its potential mechanism. The mRNA expression of miR-490-5p in 30 pairs of colon cancer tissues and adjacent normal tissues was detected via reverse transcription-polymerase chain reaction (RT-PCR). Human colon cancer SW480 cell lines were cultured in vitro and divided into Control group and miR-490-5p overexpression group (miR-490-5p mimic group). The nonsense sequence and miR-490-5p mimic were transfected using liposome transfection technique into colon cancer cells in control group and miR-490-5p mimic group, respectively. Cell proliferation and apoptosis in each group were then observed. At the same time, the effect of miR-490-5p on the growth of colon cancer in vivo was explored using subcutaneous tumorigenesis assay. The protein expressions of extracellular signal-regulated kinase (ERK)1/2 signaling pathway and cyclin-dependent kinase 1 (CDK1) were determined via Western blotting. Furthermore, immunohistochemical staining was performed to verify the protein expression of CDK1 in vivo. The expression of miR-490-5p in colon cancer tissues was significantly lower than that in adjacent normal tissues (p<0.05). After transfection with miR-490-5p mimic in vitro, EdU staining and colony formation assay showed that the proliferation ability of SW480 cells was significantly weakened (p<0.05). Meanwhile, the number of colonies in miR-490-5p mimic group was markedly less than that in Control group (p<0.05). The results of Western blotting revealed that overexpression of miR-490-5p remarkably up-regulated the Bax/Bcl-2 and C-Caspase3/T-Caspase3 ratios in cancer cells (p<0.05). Subsequent results indicated that the subcutaneous tumorigenesis of colon cancer cells was markedly inhibited by overexpression of miR-490-5p (p<0.05). According to the results of Western blotting, the activation of ERK signaling pathway and the protein expression of CDK1 were significantly suppressed by overexpression of miR-490-5p (p<0.05). In vivo experiments further revealed that the protein expression of CDK1 in colon cancer tissues increased significantly (p<0.05). MiR-490-5p was found lowly expressed in colon cancer patients. In addition, overexpression of miR-490-5p inhibited the proliferation and promoted the apoptosis of colon cancer cells via down-regulating CDK1 both in vitro and in vivo.

Anisodamine ameliorates ischemia/reperfusion-induced renal injury in rats through activation of the extracellular signal-regulated kinase (ERK) pathway and anti-apoptotic effect.

Anisodamine exerts significant protective effect on ischemia/reperfusion (I/R) injury in various organs. However, little is known about the mechanisms of anisodamine in renal I/R injury. Activation of extracellular regulated protein kinases (ERK) pathway promotes the repair of renal epithelial cells following oxidant injury. The present study investigated whether the renoprotective role of anisodamine against renal I/R injury in rats was associated with the activation of ERK signaling pathway. Male Sprague-Dawley (SD) rats were separated into the following groups: Sham-operated group, I/R group, anisodamine-treated group, PD98059 (MEK-1/ERK inhibitor)-treated group and anisodamine plus PD98059-treated group. A rat model of renal I/R was established by excising the right kidney and then clamping the left renal pedicle for 45 min followed by reperfusion for 24 h. Serum and renal tissue samples were obtained for assays of the associated morphological, molecular and biochemical parameters. Treatment with anisodamine ameliorated renal I/R injury, as evidenced by improvements of renal histology and kidney function, a decrease in paller's score and apoptosis index. Anisodamine also upregulated the phosphorylation levels of ERK1/2 and its downstream targets, including 90 ribosomal S6 kinase (p90rsk) and Bad, as well as the expression of antiapoptotic Bcl-2 protein, downregulated the expression levels of proapoptotic proteins Bax and cleaved-caspase-3, whereas these effects were greatly abolished by administration of PD98059. In conclusion, the results suggest that anisodamine prevents renal I/R injury in rats as a result of an activation of the ERK signaling pathway and anti-apoptotic properties.

Annexin A3 depletion overcomes resistance to oxaliplatin in colorectal cancer via the MAPK signaling pathway

Colorectal cancer (CRC) is a common disease with high mortality and morbidity. Annexin A3 (ANXA3) belongs to the structurally homologous family of Ca2+ and phospholipid-binding proteins. This study aimed to investigate the effects and potential mechanisms of ANXA3 on oxaliplatin (Ox) resistance in CRC. We generated two human CRC cell lines (HCT116/Ox and SW480/Ox) with acquired Ox resistance and determined their resistance properties. ANXA3 expression and cell apoptosis, migration and invasion also were evaluated. We found that cell viability of HCT116/Ox and SW480/Ox was higher than that in parental cells in the presence of Ox. ANXA3 was highly expressed in HCT116/Ox and SW480/Ox cells. ANXA3 downregulation diminished cell survival, migration and invasion, while increased the apoptosis of HCT116 and SW480 with or without Ox. Moreover, depletion of ANXA3 reduced cell viability and BrdU incorporation, increased cell apoptosis and c-caspase 3 expression in HCT116/Ox with or without Ox. A transwell assay determined that knockdown of ANXA3 impeded the migration and invasion of HCT116/Ox and SW480/Ox cells. Additionally, phosphorylation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) decreased upon ANXA3 depletion in HCT116/Ox cells, and ANXA3 silencing suppressed Ox-induced activation of ERK and JNK signaling pathway. ANXA3 downregulation reduced Ox resistance in CRC, and treatment with the ERK inhibitor PD098059 or JNK inhibitor SP600125 contributed to this process. These results indicate that silencing ANXA3 could overcome Ox resistance in CRC via the mitogen-activated protein kinase signaling pathway.

Activation of NPFF2 receptor stimulates neurite outgrowth in Neuro 2A cells through activation of ERK signaling pathway

Neurite outgrowth is an important process in neural regeneration and plasticity, especially after neural injury, and recent evidence indicates that several Gαi/o protein-coupled receptors play an important role in neurite outgrowth. The neuropeptide (NP)FF system contains two Gαi/o protein-coupled receptors, NPFF1 and NPFF2 receptors, which are mainly distributed in the central nervous system. The aim of the present study was to determine whether the NPFF system is involved in neurite outgrowth in Neuro 2A cells. We showed that Neuro 2A cells endogenously expressed NPFF2 receptor, and the NPFF2 receptor agonist dNPA inhibited cyclic adenosine monophosphate (cAMP) production stimulated by forskolin in Neuro 2A cells. We also demonstrated that NPFF and dNPA dose-dependently induced neurite outgrowth in Neuro 2A cells, which was completely abolished by the NPFF receptor antagonist RF9. Pretreatment with mitogen-activated protein kinase inhibitors PD98059 and U0126 decreased dNPA-induced neurite outgrowth. In addition, dNPA increased phosphorylation of extracellular signal-regulated kinase (ERK) in Neuro 2A cells, which was completely antagonized by pretreatment with U0126. Our results suggest that activation of NPFF2 receptor stimulates neurite outgrowth in Neuro 2A cells through activation of the ERK signaling pathway. Moreover, NPFF2 receptor may be a potential therapeutic target for neural injury and degeneration in the future.

Xenin-induced feeding suppression is not mediated through the activation of central extracellular signal-regulated kinase signaling in mice

Xenin is a gut hormone that reduces food intake by partly acting through the hypothalamus via neurotensin receptor 1 (Ntsr1). However, specific signaling pathways that mediate xenin-induced feeding suppression are not fully understood. Activation of Ntsr1 leads to the activation of the extracellular signal-regulated kinase (ERK). Hypothalamic ERK participates in the regulation of food intake by mediating the effect of hormonal signals. Therefore, we hypothesized that the anorectic effect of xenin is mediated by hypothalamic ERK signaling. To address this hypothesis, we compared levels of phosphorylation of ERK1/2 (pERK1/2) in the hypothalamus of both control and xenin-treated mice. The effect of xenin on ERK1/2 phosphorylation was also examined in mouse hypothalamic neuronal cell lines with or without Ntsr1. We also examined the effect of the blockade of central ERK signaling on xenin-induced feeding suppression in mice. The intraperitoneal (i.p.) injection of xenin caused a significant increase in the number of pERK1/2-immunoreactive cells in the hypothalamic arcuate nucleus. The majority of pERK1/2-positive cells expressed neuronal nuclei (NeuN), a marker for neurons. Xenin treatment increased pERK1/2 levels in one cell line expressing Ntsr1 but not another line without Ntsr1 expression. Both i.p. and intracerebroventricular (i.c.v.) injections of xenin reduced food intake in mice. The i.c.v. pre-treatment with U0126, a selective inhibitor of ERK1/2 upstream kinases, did not affect xenin-induced reduction in food intake. These findings suggest that although xenin activates ERK signaling in subpopulations of hypothalamic neurons, xenin does not require the activation of hypothalamic ERK signaling pathway to elicit feeding suppression.

Role of ERK signaling pathway in inhibition of OGD/R-induced apoptosis in rat cortical neurons by sevoflurane: the relationship with mPTP

Objective To evaluate the role of extracellular signal-regulated kinase(ERK)signaling pathway in inhibition of oxygen-glucose deprivation and restoration(OGD/R)-induced apoptosis in rat cortical neurons by sevoflurane and the relationship with mitochondrial permeability transition pore(mPTP). Methods The rat cortical neurons were cultured in vitro and seeded in 6-well or 12-well culture plates.The neurons were randomly divided into 4 groups(n=18 each)using a random number table: control group(group C); OGD/R group(group O); sevoflurane group(group OS); sevoflurane+ ERK1/2 inhibitor PD98059 group(group OSP). The neurons were subjected to O2-glucose deprivation for 90 min followed by restoration of O2-glucose supply for 24 h in group O. The neurons were incubated with 2% sevoflurane for 2 h after OGD/R in group OS.OGD/R was performed at 1 h after ERK1/2 inhibitor PD98059 30 μmol/L was added, and the neurons were incubated with 2% sevoflurane for 2 h after OGD/R in group OSP.At 24 h of restoration of O2-glucose supply, the expression of phosphorylated ERK1/2(p-ERK1/2)in neurons was measured by Western blot, the neuronal apoptosis was detected using Annexin V-FITC/PI double staining combined with flow cytometry, and the opening of mPTP was determined through measuring the optical density at 540 nm.The apoptosis rate was calculated. Results Compared with group C, the expression of p-ERK1/2 in neurons was significantly up-regulated, and the apoptosis rate and mPTP opening were significantly increased in group O(P<0.05). Compared with group O, the expression of p-ERK1/2 in neurons was significantly up-regulated, and the apoptosis rate and mPTP opening were significantly decreased in group OS(P<0.05). Compared with group OS, the expression of p-ERK1/2 in neurons was significantly down-regulated, and the apoptosis rate and mPTP opening were significantly increased in group OSP(P<0.05). Conclusion The mechanism by which sevoflurane inhibits OGD/R-induced apoptosis in rat cortical neurons is related to inhibition of mPTP opening after activation of ERK signaling pathway. Key words: Extracellular signal-regulated MAP kinases; Mitochondrial membrane transport proteins; Anesthetics, inhalation; Neurons; Apoptosis

Role of spinal ERK signaling pathway in reduction of remifentanil-induced hyperalgesia by electro-acupuncture at Zusanli in rats with incisional pain

Objective To evaluate the role of spinal extracellular signal-regulated kinase (ERK) signaling pathway in reduction of remifentanil-induced hyperalgesia by electro-acupuncture (EA) at Zusanli in rats with incisional pain. Methods Fifty male adult Sprague-Dawley rats, weighing 250–280 g, in which the intrathecal catheter was successfully placed without complications, were randomly divided into 5 groups (n= 10 each) using a random number table: control group (group C), remifentanil + incisional pain group (group RI), EA at acupoint group (group E), EA at non-acupoint group (group NE), and 1/2 ERK inhibitor U0126 + EA at acupoint group (group UE). Normal saline 0.1 ml· kg–1· min–l was infused intravenously for 60 min in group C. In RI, E, NE and UE groups, after the model of incisional pain was established, remifentanil 1.0 μg · kg–1· min–l was infused for 60 min, and in addition, EA (intensity 10 mA, frequency 4 Hz) of Zusanli lasting for 60 min was performed at the same time in E and UE groups, and EA was performed at the points 5 mm lateral to the acupoints of Zusanli on the operated side simultaneously in group NE.ERK1/2 inhibitor U0126 5 μg (in 5% dimethyl sulfoxide 10 μl) was injected intrathecally in group UE, and 5% dimethyl sulfoxide 10 μl was injected intrathecally in the other groups.The mechanical paw withdrawal threshold (MWT) was measured before remifentanil or normal saline infusion (T1), and at 2 h, 1, and 2 days after the end of infusion (T2-4). After MWT was measured at T4, the expression of ERK1/2 and phosphorylated ERK1/2 (p-ERK1/2) in spinal cord dorsal horns was measured by Western blot. Results Compared with group C, the MWT was significantly decreased at T2-4 in RI, E, NE and UE groups, and the expression of p-ERK1/2 was up-regulated in RI, E and NE groups (P 0.05). Compared with group E, the MWT was significantly increased at T2-4, and the expression of p-ERK1/2 was down-regulated in group UE (P<0.05). Conclusion The mechaism by which EA at Zusanli reduces hyperalgesia induced by remifentanil in rats with incisional pain is related to inhibited activation of ERK signaling pathway in the spinal cord . Key words: Extracellular signal-regulated MAP kinases; Spinal cord; Electric stimulation therapy; POINT ST36 (ZUSANLI); Piperidines; Pain, postoperative; Hyperalgesia

Inhibitory effect of apigenin on proliferation and invasion of human malignant melanoma cells

Objective To evaluate the inhibitory effect of apigenin on malignant melanoma in vitro, and to investigate its mechanisms. Methods Human malignant melanoma cell lines A375 and C8161 were divided into test groups and control group separately. Cells in the test groups were treated with apigenin at different concentrations, while cells in the control group were treated with dimethyl sulfoxide, for different durations. Subsequently, methyl thiazolyl tetrazolium（MTT）assay, wound healing assay and Matrigel invasion assay were carried out to estimate cellular proliferative activity, migratory activity and invasive activity, respectively, and scanning electron microscopy（SEM）was used to observe morphology of melanocyte dendrites. Flow cytometry using annexin-V/propidium iodide（PI）staining was performed to detect cell apoptosis, propidium iodide（PI）staining to analyze cell cycle, and Western blot to measure the expressions of proteins related to apoptosis and involved in the extracellular signal-regulated kinase（ERK）signaling pathway. Results MTT assay showed significant differences in cellular proliferative activity between the test groups and control group（all P < 0.05）. The proliferation of A375 and C8161 cells was inhibited by apigenin in a dose-dependent manner when the concentration of apigenin was 10-40 mg/L, and in a time-dependent manner when the treatment duration varied from 0 to 48 hours. The half-maximal inhibitory concentration（IC50）of apigenin at 24 hours was 25 mg/L for both A375 and C8161 cells. Wound healing assay showed that the migration of A375 and C8161 cells was significantly decelerated after 24-hour treatment with apigenin of 10, 20 and 25 mg/L compared with the control cells（all P < 0.01）. Matrigel invasion assay demonstrated that the number of invasive cells was significantly smaller in A375 and C8161 cells treated with apigenin of 10, 20 and 25 mg/L for 72 hours than in the control cells（all P< 0.01）. SEM showed that the dendrits of both A375 and C8161 cells became thinner and longer after treatment with 25 mg/L apigenin for 24 hours, with the length of dendrits being（23.30 ± 2.62）μm and（16.50 ± 1.62）μm respectively, compared to（12.38 ± 2.27）μm and（9.36 ± 2.51）μm respectively in the control groups（both P < 0.01）. After treatment with apigenin of 10 and 25 mg/L for 24 hours, a significant increase was observed in apoptosis rate in both A375 cells（3.30% ± 0.82% vs. 0.40% ± 0.07%, P< 0.01; 10.00% ± 0.60% vs. 4.00% ± 0.70%, P< 0.01）and C8161 cells（13.10% ± 1.45% vs. 7.27% ± 1.31%; 25.77% ± 2.40% vs. 7.27% ± 1.31%; both P < 0.01）compared with the control cells. Both A375 and C8161 cells were arrested in G2/M phase after treatment with 25 mg/L apigenin for 24 hours, with the percentage of cells in G2/M phase being 48.70% ± 3.04% and 31.10% ± 1.90% respectively, compared to 21.30% ± 0.75% and 25.06% ± 2.12% respectively in the control groups（both P< 0.01）. Western blot showed an increase in the expressions of apoptosis-related proteins including cleaved caspase-3 and cleaved poly ADP-ribose polymerase（PARP）with the activation of ERK signaling pathway in both A375 and C8161 cells after 24-hour treatment with 25 mg/L apigenin compared with the control groups. Conclusions Apigenin can inhibit the proliferation, migration and invasion of, but induce apoptosis and cell cycle arrest in human malignant melanoma cells, likely by regulating the expression of ERK signaling pathway-related proteins. Key words: Apigenin; Melanoma, experimental; Cell proliferation; Cell movement; Extracellular signal-regulated MAP kinases

Induction of neurite outgrowth in PC12 cells by artemisinin through activation of ERK and p38 MAPK signaling pathways

Growth of neurite processes is a critical step in neuronal development, regeneration, differentiation, and response to injury. The discovery of compounds that can stimulate neurite formation would be important for developing new therapeutics against both neurodegenerative disorders and trauma-induced neuronal injuries. Semisynthetic derivatives of artemisinin, an active compound in Artemisia annua, have been effectively used in malaria treatment, but they have been shown to possess neurotoxic potential. In this study, we found unexpectedly that artemisinin and its derivatives induced neurite outgrowth of PC12 cells. Artemisinins containing an endoperoxide bridge such as artemisinin and dihydroartemisinin induced growth of neurite processes at concentrations that were slightly cytotoxic, artemisinin having the most potent maximal effect among them. Deoxyartemisinin, which lacks the endoperoxide bridge, was ineffective. Artemisinin-treated cells expressed increased levels of the neuronal marker βIII-tubulin. Artemisinin upregulated phosphorylation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK), critical signaling molecules in neuronal differentiation. Consistent with activation of the two MAPKs, neurite outgrowth induced by artemisinin was inhibited by the MAPK/ERK kinase inhibitor PD98059 and the p38 MAPK inhibitor SB203580. Artemisinin also induced phosphorylation of cyclic AMP response element-binding protein (CREB) that was almost completely attenuated by PD98059 but not by SB203580. Taken together, our results indicate that artemisinin and its derivatives containing the endoperoxide bridge induced differentiation of PC12 cells toward a neuronal phenotype and suggest that both activation of ERK signaling pathway, which leads to CREB phosphorylation, and activation of p38 MAPK signaling pathway are involved in this process.

Abstract 1436: Interleukin-17A stimulates expression of chemokines and cytokines in prostatic epithelial cells via nuclear factor-kappaB and extracellular signal-regulated kinase pathways

Abstract T helper 17 (Th17) cells secret interleukin-17A (IL-17A) and interleukin-17F. Th17 cells and IL-17A are increased in prostate tumors. The role of IL-17A in prostate cancer is not clear. The objective of this in-vitro study was to investigate the effects of IL-17A on prostatic epithelial cells. Cell growth assay, Western blot, and real-time quantitative reverse transcription- polymerase chain reaction (qRT-PCR) analysis were used to measure the effects of IL-17A on the immortalized normal human prostatic epithelial cell lines (RWPE-1 and pRNS-1-1), human high-grade prostatic intraepithelial neoplasia (PIN) cell line, human prostate cancer LNCaP cell line, and mouse prostate cancer TRAMP-C1 cell line. We found that recombinant human IL-17A (20 ng/ml) did not affect cell growth rate in any of the prostatic cell lines studied. IL-17A did not significantly activate Nuclear Factor-kappaB (NF-kappaB) or Extracellular signal-Regulated Kinase (ERK) signaling pathway in RWPE-1, pRNS-1-1, or PIN cells. IL-17A activated NF-kappaB and/or ERK signaling pathways in LNCaP and TRAMP-C1 cells. When IL-17RC (receptor of IL-17A) was overexpressed in PIN and LNCaP cells, activation of NF-kappaB or ERK signaling pathway by IL-17A was significantly enhanced. IL-17A modestly induced mRNA expression of chemokines (CXCL1 and CXCL2) in RWPE-1, pRNS-1-1, and PIN cells. IL-17A induced mRNA expression of chemokines (CXCL1, CXCL2, CCL2, CCL5, CCL7, and CCL20) and cytokine IL-6 in LNCaP cells. In mouse TRAMP-C1 cells, IL-17A induced mRNA expression of chemokines (CXCL1, CXCL2, CXCL5, CCL2, and CCL5) and cytokine IL-6. When IL-17RC was overexpressed in PIN and LNCaP cells, chemokine expression was significantly enhanced. Our findings suggest that although IL-17A has no effects on cell growth in-vitro, it may modulate the stroma-epithelial interaction in tumor microenvironment in-vivo via stimulation of expression of chemokines and cytokines. Acknowledgement: DoD W81XWH-05-1-0567, NIH/NCRR 2P20 RR020152-06, and LCRC Fund. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1436.

Activation of spinal ERK signaling pathway contributes to pain-related responses induced by scorpion Buthus martensi Karch venom

It has been demonstrated that spontaneous nociceptive behaviors, cutaneous hyperalgesia and paw edema can be induced by intraplantar injection of scorpion Buthus martensi Karch (BmK) venom in rats. In the present study, activation of spinal extracellular signal-regulated kinase (ERK) signaling pathway and its contribution to pain-related responses induced by scorpion BmK venom were investigated. It was found that ERK was activated not only in the superficial layers but also in deep layers of L4–L5 spinal cord dorsal horn, which started at 2 min, peaked at 30–60 min and almost disappeared at 4 h following intraplantar injection of BmK venom. Intrathecal injection of U0126 (0.1, 1.0 and 10 μg), a widely used specific MAP kinase kinase (MEK) inhibitor, suppressed spontaneous nociceptive responses and reduced primary heat hyperalgesia and bilateral mechanical hyperalgesia induced by BmK venom. In addition, BmK venom-induced spinal c-Fos expression could be inhibited by U0126 dose-dependently. Intrathecal delivery of NMDA receptor antagonist (5R, 10S)-(+)-5-methyl-10, 11-dihydro-5H-dibenzo [a,d]-cyclohepten-5-10-imine hydrogen maleate (MK-801) and the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) could partially inhibit activation of spinal ERK induced by BmK venom at 30 min. Thus, activation of ERK in spinal cord dorsal horn, partially mediated by NMDA and non-NMDA receptor, potentially contributes to BmK venom-induced pain-related behaviors.

Epidermal growth factor and transforming growth factor-β1 enhance HK-2 cell migration through a synergistic increase of matrix metalloproteinase and sustained activation of ERK signaling pathway

Epidermal growth factor (EGF) and transforming growth factor-β1 (TGF-β1), upregulated in renal diseases, have a combinational effect on epithelial–mesenchymal transformation (EMT) of renal proximal tubular cells. The aim of this study was to examine the mechanism regarding the combinational effect of EGF and TGF-β1 on cell migration following EMT. The results demonstrated that EGF (10 ng/ml) and TGF-β1 (3 ng/ml) synergistically increased cell migration, accompanied by an increase in matrix metalloproteinase-9 ( MMP-9) gene expression, production and activity. Inhibition of MMP-9 production and activity by an MMP-2/MMP-9-specific inhibitor blocked the synergistic effect of EGF and TGF-β1 on cell migration. The kinetic profile of extracellular signal-regulated kinase (ERK) signals demonstrated that ERK1/2 activation was rapidly and strongly induced by EGF but delayed and less marked by TGF-β1 stimulation. In contrast, co-administration of EGF and TGF-β1 caused an early pronounced and persistent ERK1/2 activation. Inhibition of the ERK1/2 activity by PD98059 abrogated the synergistic effect of EGF and TGF-β1 on cell migration, MMP-9 production and activity, indicating that EGF and TGF-β1 converged at the ERK signaling pathway to mediate cell migration. This study demonstrates that EGF and TGF-β1 synergistically stimulate proximal tubular cell migration through the increased MMP-9 function and enhanced ERK1/2 activation.

Activation of the spinal ERK signaling pathway contributes naloxone-precipitated withdrawal in morphine-dependent rats

Extracellular signal-regulated kinase (ERK), a mitogen-activated protein kinases (MAPK), transduces a broad range of extracellular stimuli into diverse intracellular responses. Recent studies have showed that ERK activation in the supraspinal level involved in the development of drug dependence, especially in psychological dependence. In this study, we reported that the spinal ERK signaling pathway was activated by chronic morphine injection. There was a further increase in ERK activation after naloxone-precipitated withdrawal. Furthermore, attenuation of the spinal ERK phosphorylation by intrathecal a MAPK kinase (MEK) inhibitor U0126 or knockdown of the spinal ERK by antisense oligonucleotides not only decreased the scores of morphine withdrawal, but also attenuated withdrawal-induced allodynia, which were accompanied by decreased ERK phosphorylation in the spinal cord. The spinal ERK inhibition or knockdown also reduced morphine withdrawal-induced phosphorylation of cAMP response element binding protein (CREB), which is one of the important downstream substrates of ERK pathway, and Fos expression. The involvement of the spinal ERK in morphine withdrawal was supported by our finding that intrathecal N-methyl- d-aspartate receptor antagonist MK-801 or protein kinase C inhibitor chelerythrine chloride suppressed withdrawal-induced ERK activation in the spinal cord and attenuated morphine withdrawal symptoms. These findings suggest activation of the spinal ERK signaling pathway contributes naloxone-precipitated withdrawal in morphine-dependent rats.

Potential use of nerve growth factor to treat Alzheimer's disease

Growth of neurite processes is a critical step in neuronal development, regeneration, differentiation, and response to injury. The discovery of compounds that can stimulate neurite formation would be important for developing new therapeutics against both neurodegenerative disorders and trauma-induced neuronal injuries. Semisynthetic derivatives of artemisinin, an active compound in Artemisia annua, have been effectively used in malaria treatment, but they have been shown to possess neurotoxic potential. In this study, we found unexpectedly that artemisinin and its derivatives induced neurite outgrowth of PC12 cells. Artemisinins containing an endoperoxide bridge such as artemisinin and dihydroartemisinin induced growth of neurite processes at concentrations that were slightly cytotoxic, artemisinin having the most potent maximal effect among them. Deoxyartemisinin, which lacks the endoperoxide bridge, was ineffective. Artemisinin-treated cells expressed increased levels of the neuronal marker βIII-tubulin. Artemisinin upregulated phosphorylation of extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK), critical signaling molecules in neuronal differentiation. Consistent with activation of the two MAPKs, neurite outgrowth induced by artemisinin was inhibited by the MAPK/ERK kinase inhibitor PD98059 and the p38 MAPK inhibitor SB203580. Artemisinin also induced phosphorylation of cyclic AMP response element-binding protein (CREB) that was almost completely attenuated by PD98059 but not by SB203580. Taken together, our results indicate that artemisinin and its derivatives containing the endoperoxide bridge induced differentiation of PC12 cells toward a neuronal phenotype and suggest that both activation of ERK signaling pathway, which leads to CREB phosphorylation, and activation of p38 MAPK signaling pathway are involved in this process.