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Extracellular signal-regulated kinase 1/2 (ERK1/2) is a serine/threoninekinase involved in the signal transduction cascade of Ras-Raf-mitogen-activated protein kinase (MEK)-ERK.It participates in the cell growth,proliferation and even invasion by regulating gene transcription and expression.The occurrence of a variety of diseases such as lung cancer,liver cancer,ovarian cancer,cervical cancer,endometriosis,and preeclampsia,as well the metastasis and disease progression,is closely associated with the regulation of cell invasion by ERK1/2 signaling pathway.Therefore,exploring the regulation of ERK1/2 signaling on cell invasion and its role in pathogenesis of diseases may help to develop more effective treatment schemes.This article introduces recent progress in the regulation of ERK1/2 signaling on cell invasion and the role of such regulation in diseases,with a view to give new insights into the clinical treatment of ERK 1/2-related diseases.
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Female , Pregnancy , Humans , Mitogen-Activated Protein Kinase 3 , Signal Transduction , Mitogen-Activated Protein Kinases , Cell Cycle , Cell ProliferationABSTRACT
Objective:To clarify the neuroprotective effects of neural cell adhesion molecule (NCAM) derived peptide P2 on in vitro cultured neuron and ischemic stroke rat. Methods:Primary cortical neurons were extracted and cultured, and CCK-8 method was used to observe the protective effect of different concentrations of P2 on cortical neurons under oxygen-glucose deprivation (OGD) conditions.The levels of apoptosis-related proteins and extracellular signal regulated kinase 1/2 (Erk1/2) were observed by Western blot. Clean grade male SD rats were selected for animal experiments. The middle cerebral artery occlusion (MCAO) method was used to establish the rat model of cerebral ischemia/reperfusion injury. The rats with successful model were divided into sham operation group, MCAO group and MCAO+ P2 group according to the random number table, with 12 rats in each group. After operation, rats in MCAO+ P2 group were subcutaneously injected with 1 mg/kg P2 once a day until 14 days after operation, and rats in the other two groups were subcutaneously injected with 0.9% sodium chloride solution of the same volume.Beam-walking test was used to evaluate the motor function of rats.Immunofluorescence staining and Western blot were used to detect the in-situ apoptosis of neuronal cells and the expression of Erk1/2 in ischemic penumbra of rat brains, respectively. All statistical analyses were performed using SPSS 22.0.Repeated measurement ANOVA was used to evaluate the beam-walking experimental data, and one-way ANOVA were used to analyze other experimental data among multiple groups.Results:Compared with OGD group, 0.5, 1.0 and 2.0 μmol/L P2 improved the activity of neurons under OGD conditions, of which 1 μmol/L P2 had the best effect ((2.436±0.284), (1.551±0.410), P<0.05). Western blot showed that the protein levels of bax ((76.120±3.232)%, (88.965±5.208)%, P<0.05), cleaved caspase-3 ((76.736±4.306)%, (97.781±8.111)%, P<0.05) and cleaved caspase-9 ((88.833±6.581)%, (104.962±4.788)%, P<0.05) in 1 μmol/L P2 treated group were all lower than those in OGD group, while the protein levels of bcl-2 ((56.146±3.882)%, (43.170±6.945)%, P<0.05) and phosphorylated Erk1/2 ((73.583±8.557)%, (55. 219±4.615)%, P<0.05) in 1 μmol/L P2 treated group were both higher than those in OGD group. Compared with MCAO group, on the 14th day after P2 intervention, the slip ratio of hindlimb of the paralyzed hind limbs of rats was lower ((23.438±11.540)%, (41.733±13.631)%, P<0.05), the apoptosis rate of neurons around the focus was lower ((13.144±6.485)%, (26. 699±6. 402)%, P<0.05), and the level of phosphorylated Erk1/2 protein in the brain tissues around the infarct focus was higher ((74.062±7.458)%, (53.327±7.093)%, P<0.05). Conclusion:Low doses of neural cell adhesion molecule derived peptide P2 exert neuroprotective effects on OGD neurons and ischemic stroke rats. The underlying mechanism may be related to the activation of Erk.
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Objective:To investigate changes in circadian gene cryptochrome 2 (CRY2) expression in mouse models of psoriasis and HaCaT cells, and to explore underlying mechanisms.Methods:Imiquimod-induced mouse model experiment: 12 C57BL/6 female mice were randomly and equally divided into imiquimod group receiving topical imiquimod treatment for 5 consecutive days and control group receiving no treatment; these mice were sacrificed on day 6, skin tissues were resected from the back of mice, and immunofluorescence staining was performed to determine the CRY2 expression in the epidermis. HaCaT cell transfection experiment: HaCaT cells with small interfering RNA (siRNA) -mediated knockdown of CRY2 served as siRNA-CRY2 group, and siRNA-NC group as control group; 5-ethynyl-2′-deoxyuridine (EdU) staining was performed to evaluate the proliferative activity of the HaCaT cells, real-time fluorescence-based quantitative PCR (qPCR) to determine the mRNA expression of chemokines in the HaCaT cells, and Western blot analysis to determine phosphorylation levels of extracellular signal-regulated kinase 1/2 (ERK1/2) . Tumor necrosis factor-α (TNF-α) -stimulated animal and cell experiments: 12 C57BL/6 female mice were randomly and equally divided into TNF-α group subcutaneously injected with TNF-α solution in the ear for 6 days, and phosphate buffered saline (PBS) group subcutaneously injected with the same amount of PBS; the mice were sacrificed on day 7, skin tissues were resected from the ear of mice, and immunofluorescence staining was conducted to determine the CRY2 expression in the epidermis; CRY2-knockdown HaCaT cells stimulated with 50 ng/ml TNF-α for 12 hours served as siRNA-CRY2 + TNF-α group, and siRNA-NC + TNF-α group as control group; qPCR was performed to determine the mRNA expression of chemokines in HaCaT cells in the above groups. Statistical analysis was carried out by using two-independent-sample t test. Results:Immunofluorescence staining showed that the CRY2 protein expression was significantly lower in the mouse dorsal epidermis in the imiquimod group (0.94 ± 0.23) than in the control group (2.30 ± 0.25, t = 3.99, P = 0.016) . Compared with the siRNA-NC group, the siRNA-CRY2 group showed significantly increased proportions of EdU-positive cells (48.13% ± 10.97% vs. 38.23% ± 0.81%, t = 5.00, P = 0.007) , mRNA expression levels of chemokines CXCL1 and CXCL8, as well as significantly increased phosphorylated (p) -ERK1/2 protein expression levels (all P < 0.05) , while there were no significant differences in the CCL20 mRNA expression or ERK1/2 protein expression between the two groups (both P > 0.05) . Immunofluorescence staining showed significantly decreased CRY2 protein expression level in the mouse ear epidermis in the TNF-α group (0.37 ± 0.34) compared with the PBS group (2.04 ± 0.17, t = 4.38, P = 0.012) ; the relative mRNA expression levels of chemokines CXCL1, CXCL8, and CCL20 in HaCaT cells were significantly higher in the siRNA-CRY2 + TNF-α group than in the siRNA-NC + TNF-α group (all P < 0.05) . Conclusion:CRY2 was markedly underexpressed in psoriasis, which might promote the proliferation of keratinocytes and expression of chemokines CXCL1, CXCL8 and CCL20, and TNF-α might be an upstream cytokine that could downregulate CRY2 expression.
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The mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway is widely activated by a variety of extracellular stimuli, and its dysregulation is associated with the proliferation, invasion, and migration of cancer cells. ERK1/2 is located at the distal end of this pathway and rarely undergoes mutations, making it an attractive target for anticancer drug development. Currently, an increasing number of ERK1/2 inhibitors have been designed and synthesized for antitumor therapy, among which representative compounds have entered clinical trials. When ERK1/2 signal transduction is eliminated, ERK5 may provide a bypass route to rescue proliferation, and weaken the potency of ERK1/2 inhibitors. Therefore, drug research targeting ERK5 or based on the compensatory mechanism of ERK5 for ERK1/2 opens up a new way for oncotherapy. This review provides an overview of the physiological and biological functions of ERKs, focuses on the structure-activity relationships of small molecule inhibitors targeting ERKs, with a view to providing guidance for future drug design and optimization, and discusses the potential therapeutic strategies to overcome drug resistance.
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OBJECTIVE@#To investigate the influence of chronic masseter hyperalgesia induced by 17β-estradiol (E2) and experimental occlusal interference (EOI) on underlying mechanism in hippocampus of ovariectomized (OVX) rats.@*METHODS@#In the study, 32 OVX rats were randomly divided into 4 groups (8 rats/group): The control group was OVX group, and 0 μg/d E2 (vehicle) injection was started 7 d after OVX without EOI; in the experimental group (1) OVX + E2 group, 80 μg/d E2 injection was started 7 d after OVX without EOI; in the experimental group (2) OVX + EOI group, vehicle injection was started 7 d after OVX and EOI was applied 17 d after OVX; in the experimental group (3) OVX + E2 + EOI group, 80 μg/d E2 injection was started 7 d after OVX and EOI was applied 17 d after OVX. Bilateral masseter muscle mechanical withdrawal thresholds were measured before OVX, 7 days after OVX (before E2 injection), 17 days after OVX (10 days after E2 injection and before EOI) and 24 days after OVX (7 days after EOI). Immunofluorescence staining was used to reveal phospho-extracellular signal regulated kinase 1/2 (p-ERK1/2)-positive neurons in CA3 of hippocampus. The protein expression of p-ERK1/2 in hippocampus was detected using Western Blot.@*RESULTS@#Compared with the control group [left side: (135.3±8.5) g, right side: (135.4±10.8) g], bilateral masseter muscle mechanical withdrawal thresholds of OVX+E2 group [left side: (113.3±5.6) g, right side: (112.5 ± 5.6) g] and OVX+EOI group [left side: (93.3±5.4) g, right side: 90.8±5.5) g] were decreased (P < 0.01). Bilateral masseter muscle mechanical withdrawal thresholds were significantly lower in OVX+E2+EOI group [left side: (81.2±6.2) g, right side: 79.8±7.7) g] than in the control, OVX+E2 and OVX+EOI groups (P < 0.05). The proportion of p-ERK1/2 positive neurons in the CA3 region of the hippocampus was increased in the control, OVX+E2, OVX+EOI and OVX+E2+EOI groups in turn, and the difference between the groups was statistically significant (P < 0.05). p-ERK1/2 protein expression was increased in the control, OVX+E2 and OVX+EOI groups in turn, but the difference was not statistically significant (P>0.05). p-ERK1/2 expression was significantly higher in OVX+E2+EOI group than in the other three groups (P < 0.05).@*CONCLUSION@#High concentration of E2 could exacerbated EOI-induced chronic masseter hyperalgesia in ovariectomized rats, and its central mechanism may be related to the upregulation of the phosphorylation of ERK1/2 in hippocampus.
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Animals , Female , Humans , Rats , Estradiol , Hippocampus , Hyperalgesia/chemically induced , Masseter Muscle , Ovariectomy , Rats, Sprague-DawleyABSTRACT
Objective: To research the effect of matrine on the proliferation and migration of HepG2 cells through extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway. Methods: HepG2 cell was selected and divided into blank control group, experimental group (matrine 1, 2, and 4 mg/mL), and positive control group (PD98059, ERK1/2 inhibitor). MTT measure was used to detect the effective time and concentration which matrine inhibits HepG2 cells. After 24 h, the effect of effective concentration of matrine on the of morphological changing HepG2 cells was observed. The invasion ability was assayed by transwell method, the expression of ERK1/2 and pERK1/2 were detected through Western blot, and reverse transcription polymerase chain reaction was used to test the expression level of ERK1/2 mRNA. Results: With the increase of matrine concentration, the number of adherent HepG2 cells gradually decreased, the morphologic changes gradually became spherical, some cell morphology was incomplete, and even cell fragments appeared. The proliferation and invasion ability of HepG2 cells decreased. The expression of ERK1/2, pERK1/2, and ERK1/2 mRNA downregulated with the increase of matrine concentration (P < 0.05). Conclusion: Matrine inhibits the proliferation and migration of HepG2 cells by downregulating the ERK1/2 signaling pathway.
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Total Panax notoginseng saponin (TPNS) is the main bioactivity compound derived from the roots and rhizomes of Panax notoginseng (Burk.) F.H. Chen. The aim of this study was to investigate the effectiveness of TPNS in treating vascular neointimal hyperplasia in rats and its mechanisms. Male Sprague-Dawley rats were randomly divided into five groups, sham (control), injury, and low, medium, and high dose TPNS (5, 10, and 20 mg/kg). An in vivo 2F Fogarty balloon-induced carotid artery injury model was established in rats. TPNS significantly and dose-dependently reduced balloon injury-induced neointimal area (NIA) (P<0.001, for all doses) and NIA/media area (MA) (P<0.030, for all doses) in the carotid artery of rats, and PCNA expression (P<0.001, all). The mRNA expression of smooth muscle (SM) α-actin was significantly increased in all TPNS groups (P<0.005, for all doses) and the protein expression was significantly increased in the medium (P=0.006) and high dose TPNS (P=0.002) groups compared to the injury group. All the TPNS doses significantly decreased the mRNA expression of c-fos (P<0.001). The medium and high dose TPNS groups significantly suppressed the upregulation of pERK1/2 protein in the NIA (P<0.025) and MA (P<0.004). TPNS dose-dependently inhibited balloon injury-induced activation of pERK/p38MAPK signaling in the carotid artery. TPNS could be a promising agent in inhibiting cell proliferation following vascular injuries.
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Animals , Male , Rats , Saponins/pharmacology , Carotid Artery Injuries/prevention & control , p38 Mitogen-Activated Protein Kinases/metabolism , Panax notoginseng/drug effects , Neointima/pathology , Immunohistochemistry , Signal Transduction , Up-Regulation , Rats, Sprague-Dawley , Carotid Artery Injuries/etiology , Real-Time Polymerase Chain Reaction , HyperplasiaABSTRACT
Objective To explore the value of extracellular signal-regulated kinase (ERK)-1/2 and p-ERK-1/2 expression in predicting the efficacy of neoadjuvant chemotherapy in cervical squamous cell carcinoma.Methods Fifty-seven patients with cervical squamous cell carcinoma who received neoadjuvant chemotherapy from May 2016 to May 2018 in Bazhong Central Hospital of Sichuan Province were selected.The incidence of adverse reactions during treatment was analyzed.The expression of ERK 1/2 and p-ERK 1/2 in cervical squamous cell carcinoma tissues before and after treatment was compared,and the expression levels of ERK 1/2 and p-ERK 1/2 and the effectiveness of neoadjuvant chemotherapy were analyzed.Results After treatment,the effective rate was 68.42％ (39/57),and the incidence of adverse reactions was 24.56％ (14/57).Before treatment,the positive rate of ERK1/2 was 87.72％ (50/57),after treatment,the positive rate of ERK 1/2 was 31.58％ (18/57),and there were statistical differences (P＜ 0.01).Before treatment,the positive rate of p-ERK 1/2 was 75.44％ (43/57),after treatment,the positive rate of p-ERK 1/2 was 19.30％ (11/57),and there was statistical difference (P ＜ 0.01).The effective rate after treatment in pre-treatment ERK 1/2 positive group was significantly lower than that in pre-treatment ERK 1/2 negative group:53.12％ (17/32) vs.88.00％ (22/25),and there was statistical difference (P ＜ 0.05).The effective rate after treatment in pre-treatment p-ERK 1/2 positive group was significantly lower than that in pre-treatment p-ERK 1/2 negative group:60.47％ (26/43) vs.13/14,and there was statistical difference (P ＜ 0.05).Conclusions ERK 1/2 and p-ERK 1/2 in cervical squamous cell carcinoma can predict the efficacy of neoadjuvant chemotherapy.
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Objective@#To explore the value of extracellular signal-regulated kinase (ERK)-1/2 and p-ERK-1/2 expression in predicting the efficacy of neoadjuvant chemotherapy in cervical squamous cell carcinoma.@*Methods@#Fifty-seven patients with cervical squamous cell carcinoma who received neoadjuvant chemotherapy from May 2016 to May 2018 in Bazhong Central Hospital of Sichuan Province were selected. The incidence of adverse reactions during treatment was analyzed. The expression of ERK 1/2 and p-ERK 1/2 in cervical squamous cell carcinoma tissues before and after treatment was compared, and the expression levels of ERK 1/2 and p-ERK 1/2 and the effectiveness of neoadjuvant chemotherapy were analyzed.@*Results@#After treatment, the effective rate was 68.42% (39/57), and the incidence of adverse reactions was 24.56% (14/57). Before treatment, the positive rate of ERK1/2 was 87.72% (50/57), after treatment, the positive rate of ERK 1/2 was 31.58% (18/57), and there were statistical differences (P<0.01). Before treatment, the positive rate of p-ERK 1/2 was 75.44% (43/57), after treatment, the positive rate of p-ERK 1/2 was 19.30% (11/57), and there was statistical difference (P<0.01). The effective rate after treatment in pre-treatment ERK 1/2 positive group was significantly lower than that in pre-treatment ERK 1/2 negative group: 53.12% (17/32) vs. 88.00% (22/25), and there was statistical difference (P<0.05). The effective rate after treatment in pre-treatment p-ERK 1/2 positive group was significantly lower than that in pre-treatment p-ERK 1/2 negative group: 60.47% (26/43) vs. 13/14, and there was statistical difference (P<0.05).@*Conclusions@#ERK 1/2 and p-ERK 1/2 in cervical squamous cell carcinoma can predict the efficacy of neoadjuvant chemotherapy.
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Objective To observe the effect of edaravone (EDA) on the expressions of malondialdehyde (MDA), phosphorylated extracellular signal-regulated kinase (p-ERK), nuclear factor kappa-B cells (NF-κB) and Collagen-III in rat model of unilateral ureteral obstruction (UUO), and discuss the protective effect of EDA on renal-interstitial fibrosis. Methods Fifty-four male SD rats were randomly divided into three groups (18 each): sham group (normal control), UUO group and EDA group. Rats in UUO group and EDA group received UUO operation. One day before UUO operation, rats in EDA group received intraperitoneal injection of 3.75mg/(kg·d) EDA, and those in sham group and UUO group were injected with equivalent normal saline. Rats were sacrificed in three batches at the 3rd, 7th and 14th day after operation, six from each group in each batch. The renal pathological changes were observed via HE staining and Masson staining. The contents of MDA in renal tissues were determined by spectrophotometry, and the expression levels of p-ERK1/2, NF-κB and collagen-III were detected by immunohistochemistry. Results HE and Masson staining showed no significant changes of MDA content, renal tubules and interstitium in sham group at 3rd, 7th and 14th day, but obvious changes of interstitium and increased fibre deposition in UUO group and EDA group compared with sham group. Compared with UUO group, the lesions of renal tubular and interstitium significantly reduced and the interstitial fibrosis deposition decreased in EDA group. Spectrophotometry indicated that the MDA contents showed no significant difference between EDA group and UUO group [(9.261±0.496)nmol/mg prot and (10.143±0.301)nmol/mg prot] on the 3rd day after operation, but was higher than in sham group [(6.918±1.120)nmol/mg prot]. The MDA contents at the 7th and 14th day after surgery in EDA group [(11.545±0.620)nmol/mg prot and (15.203±0.512)nmol/mg prot] were significantly lower than in UUO group [(13.405±0.612)nmol/mg prot and (18.133±1.684)nmol/mg prot]. Immunohistochemistry analysis showed that no significant difference existed in the expressions of p-ERK1/2, NF-κB and Collagen III at the 3 time points in sham group; At the 3rd day after surgery, the expressions of p-ERK1/2, NF-κB and Collagen III were higher in EDA group (19 055.7±1866.1, 11 217.0±989.9 and 9724.1±341.9) than in sham group (14 100.3±1822.7, 7975.1±2058.5 and 6890.0±2389.9, P0.05). At the 7th and 14th day after surgery, the expression level of p-ERK1/2 was significantly lower in EDA group (26 228.0±523.0 and 30 647.1±583.8) than in UUO group (28 254.9±886.6 and 33 240.3±1330.4); the expression level of NF-κB was obviously lower in EDA group (16 374.4±1045.3 and 21 111.2±1022.5) than in UUO group (18 799.4±357.0 and 27 125.2±2873.3); the expression level of Collagen-III was markedly lower in EDA group (12 470.9±506.1 and 19 615.8±1120.1) than in UUO group (15 049.9±1372.3 and 22 868.9±1889.2), all the differences were of statistical significance (P<0.05). Conclusion Oxidative stress and the signaling pathway of p-ERK1/2 and NF-κB may play a role in the progress of renal interstitial fibrosis (RIF) in UUO rat model, and EDA may have an antifibrosis effect by down-regulating the levels of oxidative stress in kidney tissue and inhibiting the signaling pathway of p-ERK1/2 and NF-κB.
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Objective To investigate the function of cAMP response element binding protein (CREB) and extracellular signal-regulated kinase (ERK1/2) in osteoclast differentiation mediated by Ca2+/calmodulin-dependent kinase Ⅱ (CaMK Ⅱ)δ,and elucidate the molecular mechanism thereof.Methods CaMK Ⅱδ RNA interference lentivirus vector was constructed and mouse RAW264.7 cells were transfected with the virus to determine the interference efficiency.After virus transfection,RAW264.7 cells were treated with 50ng/ml receptor activator of nuclear factor κB ligand (RANKL) and the phosphorylation levels of CREB and ERK1/2 were detected at different time points.The cells were also treated with PD98059,an ERK1/2 inhibitor,to determine the effect of ERK1/2 signal blocking on the expression of nuclear factor activated T-cells cytoplasmic 1 (NFATc1) and osteoclast differentiation.Results Interference efficiency of recombinant CaMK Ⅱδ virus vector was 77.2％ at mRNA level and 70.2％ at protein level.CaMK Ⅱδ RNA interference significantly suppressed phosphorylation of CREB and ERK1/2,and the levels ofp-CREB and p-ERK1/2 were down-regulated by 21％-55％ and 55％-64％,respectively.ERK1/2 inhibitor significantly down-regulated the protein expression of NFATc1,and the number of osteoclast,the number and size of bone resorption lacunae decreased by 39.3％,50.0％ and 52.3％,respectively.Conclusion CaMK Ⅱδ RNA interference may significantly suppress the phosphorylation of CREB and ERK1/2,and CREB and ERK1/2 have mediated the CaMK Ⅱδ-induced osteoclast differentiation.
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Objective To investigate the effects of intrathecal injection of Roscovitine on the pain behavior and the expression of pERK1/2 and pPPARγ receptor in spinal cord in rats with neuropathic pain.Methods Fifty-two male adult Sprague-Dawley rats of 220～ 250 g were randomly divided into 4 groups:Sham+DMSO group (dimethyl sulfoxide,S+D group),Sham+Roscovitine (S+R) group,CCI+ DMSO group (I+D),CCI+ Roscovitine (I+R) group.The corresponding drugs were administered by intrathecal injection from the seventh day after CCI once a day for 14 days.The right hind paw mechanical threshold value (PMWT) was measured by Von Frey filament and paw thermal withdrawal latency(PWTL) was measured by Hargreaves methods before 1 day and 3 d,7 d,10 d,14 d after surgery.The spinal cord lumbar enlargement was taken at 14 d after surgery,and Cdk5,p35,phosphorylated ERK protein (pERK),total ERK protein (ERK),phosphorylated PPARγprotein (pPPARγ) and total PPARγ protein (PPARγ) were detecteded by Western blot.Results Roscovitine was administered by intrathecal injection alleviated mechanical allodynia and thermal hyperalgesia of CCI rats.Compared with I+D group,PWMT in I+R group at 7 d,10 d,14 d after administration((4.65±0.08)g,(6.47±0.12)g,(7.90±0.19)g,) vs ((3.71 ±0.06)g,(2.45±0.17)g,(2.31±0.15)g) (Fgroup =505.71,P＜0.05,Ftime×group =15.33,P＜0.05),PWTL((9.22±0.33) s,(13.52±0.43) s,(12.63±0.88) s) vs ((8.02±0.20) s,(5.90±0.28) s,(5.40±0.38) s) (Fgroup =355,P＜0.05,F time×group =8.589,P＜0.05) were significantly increased.Compared with I+D group(p35 (0.58±0.02),pERK (1.12±0.13),pPPARγ (0.77±0.16)),p35 (0.46±0.04,F=11.06,P＜0.05) and pERK(0.79±0.11,F =22.91,P＜ 0.05) in I+ R group,were significantly dereased,and the expression of pPPARγ(0.99±0.13,F =17.62,P＜0.05))was significantly increased.Conclusion Intrathecal injection Roscovitine can alleviate both mechanical allodynia and thermal hyperalgesia in rats with neuropathic pain,and its mechanisms may be related to the inhibition of Cdk5/p35 and ERK activity,enhancement of PPARγ activity in the spinal dorsal horn.
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AIM: To investigate whether inactivation of extracellular signal-regulated kinase 1/2 ( Erk1/2 ) will affect the function of fibroblast growth factor 21 (FGF21) to regulate glucose and lipid metabolism .METHODS:Male db/db mice (8 weeks old) were treated with U0126 (an inhibitor of Erk1/2 kinase) for 1 week, and then treated with re-combinant human FGF21 protein and adenovirus-mediated FGF21 (Ad-FGF21).The profile changes of blood glucose and blood lipid were evaluated at 120 min or 4 weeks after FGF21 administration.Meanwhile, the molecular mechanism was ex-plored by in vitro study.RESULTS: Treatment of db/db mice with recombinant human FGF21 protein significantly re-duced blood glucose and triglyceride levels at 120 min after FGF21 administration , but these changes were comparable in U0126-treated mice .Furthermore , abnormal glucose and triglyceride levels , and glucose and insulin tolerance were strong-ly improved in db/db mice as accompanied with decreasing body fat content after 4 weeks of ad-FGF21 administration .In-terestingly, treatment with or without U0126 did not influence these effects of FGF21.Mechanically, treatment with Ad-FGF21 significantly upregulated the protein levels of p-Erk1/2 and peroxisome proliferator-activated receptor γ( PPARγ) as well as the expression of adiponectin at mRNA and protein levels in adipose tissues .However , treatment with or without U0126 did not change the profiles .On the other hand , in vitro experiments also indicated that treatment of adipocytes with recombinant human FGF 21 protein significantly activated Erk 1/2 phosphorylation , and upregulated the expression levels of PPARγand adiponectin (P<0.05).However, pre-administration of U0126 did not affect the profiles.CONCLUSION:Pharmaceutical inactivation of Erk 1/2 by U0216 does not affect the biological function of FGF 21 to regulate blood glucose balance and improve abnormal blood lipids in vivo.
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Objective To study the influence of glycyrrhetinic acid (GA) on bronchial asthma (BA) smooth muscle proliferation and apoptosis as well as inflammatory factor expression and its molecular mechanism. Methods Male SD guinea pigs were selected and made into asthma models, bronchial asthma smooth muscle cells were cultured and divided into BA group, GA group and GA + LM group that were treated with serum-free RPMI1640 culture medium, serum-free RPMI1640 culture medium containing 50 ng/mL glycyrrhetinic acid, serum-free RPMI1640 culture medium containing 50 ng/mL glycyrrhetinic acid and 100 ng/mL LM22B-10 respectively; normal guinea pigs were collected and bronchial smooth muscle cells were cultured as control group. The cell proliferation activity as well as the expression of proliferation and apoptosis genes, inflammatory factors and p-ERK1/2 was determined. Results Proliferation activity value and mRNA expression of Bcl-2, TNF-α IL-4, IL-6, YKL-40, protein expression of p-ERK1/2 of airway smooth muscle cell in BA group were significantly higher than those of control group while mRNA expression levels of Bax, caspase-9 as well as caspase-3 were significantly lower than that of control group (P < 0.05); proliferation activity value and mRNA expression of Bcl-2, TNF-α IL-4, IL-6, YKL-40, protein expression of p-ERK1/2 of airway smooth muscle cell in GA group were significantly lower than those of BA group (P < 0.05) while the mRNA expression levels of Bax, caspase-9 as well as caspase-3 were significantly higher than those of BA group (P < 0.05); proliferation activity value and mRNA expression of Bcl-2, TNF-α IL-4, IL-6, YKL-40 of airway smooth muscle cell in GA + LM group were significantly higher than those of GA group (P < 0.05) while mRNA expression levels of Bax, caspase-9 as well as caspase-3 were significantly lower that of GA group (P < 0.05). Conclusion GA can inhibit the proliferation of bronchial smooth muscle cells and reduce the expression of inflammatory factors by inhibiting the phosphorylation of ERK1/2.
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Objective To explore the effects of acupuncture-rehabilitation therapy on neurological function recovery and extracellular signal-regulated kinase 1/2 (ERK1/2) signaling pathway activation after permanent focal cerebral ischemia in rats. Methods Ninty male Sprague-Dawley rats were divided into five groups, namely sham group, model group, acupuncture group, rehabilitation group and acupunc-ture-rehabilitation group, and 18 in each group. Each group was further divided into 3 days, 7 days and 14 days subgroups (n=6). Their mid-dle cerebral arteries were occluded except those of sham group. The sham and model groups accepted no treatment, while the acupuncture group accepted cluster needling of scalp acupuncture, rehabilitation group accepted treadmill training, and the acupuncture-rehabilitation group accepted both acupuncture and treadmill training. They were assessed with modified Neurologic Severity Score (mNSS) 3, 7 and 14 days after modeling, while the expression of ERK1/2 and p-ERK1/2 were determined with Western blotting. Results The neurologic severi-ty score reduced in all three treatment groups (P<0.05) compared with that of the model group at every time point, and was the least in the acupuncture-rehabilitation group (P<0.05) 7 and 14 days after modeling among the treatment groups. Meanwhile, the expression of p-ERK1/2 increased in all three treatment groups (P<0.05), and was the most in the acupuncture-rehabilitation group (P<0.05), as well as the rate of (p- ERK1/2)/(ERK1/2) (P<0.05). Conclusion Acupuncture- rehabilitation therapy can promote the neurological function recovery, which may be associated with activation of ERK1/2 signaling pathway.
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Objective:To observe effect of acupuncture combined with hypothermia therapy on MAPK/ERK pathway and apoptosis related factorsin rats suffered cerebral ischemia reperfusion and to explore underlying mechanisms.Methods:Middle cerebral artery ischemia model were established.Ninety SD rats were randomly assigned into a blank group,a control group,a model group,an acupuncture group,a mild hypothermia group,and an acupuncture with hypothermia group.After 72 h treatment,nervefunction defect scores were observed,and infarction area percent was detected by 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) staining;expressions of Bcl-2 and Bax were examined by immunohistochemistry;apoptotic cells were detected by TUNEL assay;and expression levels of phospho-mitogen-activated protein kinase(p-MEK2) and phospho-extracellular signal regulated kinase 1/2 (p-ERK1/2) in the rats' hippocampus ischemic side were determined by Western blot.Results:In the rats of the model group,the neural function defect scores,the infarction area percent,the expression level of Bax,and apoptotic cells increased,while the level of Bcl-2 decreased significantly.The level of p-MEK2 and p-ERK1/2 increased obviously compared with the blank and control groups (P＜0.05 or P＜0.01).After treatment with acupuncture and hypothermia,the neural function defect scores,infarction area percent,and the level ofBax,apoptotic cells and the levels of p-MEK2 and p-ERK1/2 were significantly decreased,while the level of Bcl-2 in the treatment group was significantly elevated (P＜0.05 or P＜0.01) compared with the model group.Compared with the acupuncture group or the hypothermia group,the neural function defect scores and the levels of p-MEK2 and p-ERK1/2 in the acupuncture combined with hypothermia group were significantly reduced (P＜0.05 or P＜0.01).Conclusion:Acupuncture and hypothermia therapy can improve cerebral function,and reduce the cerebral injury through down-regulation of Bax level,and up-regulation of Bcl-2 level,which is related to reducing the levels of p-MEK2 and p-ERK1/2.The therapeutic effects on cerebral ischemia reperfusion injury for combination of acupuncture with hypothermia are better than those with single application of acupuncture or hypothermia.
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<p><b>OBJECTIVE</b>To analyze the effects of salvianolate on myocardial infarction in a murine in vivo model of ischemia and reperfusion (I/R) injury.</p><p><b>METHODS</b>Myocardial I/R injury model was constructed in mice by 30 min of coronary occlusion followed by 24 h of reperfusion and pretreated with salvianolate 30 min before I/R (SAL group). The SAL group was compared with SHAM (no I/R and no salvianolate), I/R (no salvianolate), and ischemia preconditioning (IPC) groups. Furthermore, an ERK1/2 inhibitor PD98059 (1 mg/kg), and a phosphatidylinositol-3-kinase (PI3-K) inhibitor, LY294002 (7.5 mg/kg), were administered intraperitoneal injection (i.p) for 30 min prior to salvianolate, followed by I/R surgery in LY and PD groups. By using a double staining method, the ratio of the infarct size (IS) to left ventricle (LV) and of risk region (RR) to LV were compared among the groups. Correlations between IS and RR were analyzed. Western-blot was used to detect the extracellular signal-regulated kinase 1/2 (ERK1/2) and protein kinase B (AKT) phosphorylation changes.</p><p><b>RESULTS</b>There were no significant differences between RR to LV ratio among the SHAM, I/R, IPC and SAL groups (P>0.05). The SAL and IPC groups had IS of 26.1%±1.4% and 22.3%±2.9% of RR, respectively, both of which were significantly smaller than the I/R group (38.5%±2.9% of RR, P<0.05, P<0.01, respectively). Moreover, the phosphorylation of ERK1/2 was increased in SAL group (P<0.05), while AKT had no significant change. LY294002 further reduced IS, whereas the protective role of salvianolate could be attenuated by PD98059, which increased the IS. Additionally, the IS was not linearly related to the RR (r=0.23, 0.45, 0.62, 0.17, and 0.52 in the SHAM, I/R, SAL, LY and PD groups, respectively).</p><p><b>CONCLUSION</b>Salvianolate could reduce myocardial I/R injury in mice in vivo, which involves an ERK1/2 pathway, but not a PI3-K signaling pathway.</p>
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Animals , Male , Blotting, Western , Cardiotonic Agents , Pharmacology , Therapeutic Uses , Flavonoids , Pharmacology , Heart Ventricles , Pathology , MAP Kinase Signaling System , Mice, Inbred C57BL , Mitogen-Activated Protein Kinase 1 , Metabolism , Mitogen-Activated Protein Kinase 3 , Metabolism , Myocardial Reperfusion Injury , Drug Therapy , Pathology , Organ Size , Phosphorylation , Plant Extracts , Chemistry , Pharmacology , Therapeutic Uses , Protein Kinase Inhibitors , Pharmacology , Staining and LabelingABSTRACT
OBJECTIVE@#To study the influence of glycyrrhetinic acid (GA) on bronchial asthma (BA) smooth muscle proliferation and apoptosis as well as inflammatory factor expression and its molecular mechanism.@*METHODS@#Male SD guinea pigs were selected and made into asthma models, bronchial asthma smooth muscle cells were cultured and divided into BA group, GA group and GA + LM group that were treated with serum-free RPMI1640 culture medium, serum-free RPMI1640 culture medium containing 50 ng/mL glycyrrhetinic acid, serum-free RPMI1640 culture medium containing 50 ng/mL glycyrrhetinic acid and 100 ng/mL LM22B-10 respectively; normal guinea pigs were collected and bronchial smooth muscle cells were cultured as control group. The cell proliferation activity as well as the expression of proliferation and apoptosis genes, inflammatory factors and p-ERK1/2 was determined.@*RESULTS@#Proliferation activity value and mRNA expression of Bcl-2, TNF-α, IL-4, IL-6, YKL-40, protein expression of p-ERK1/2 of airway smooth muscle cell in BA group were significantly higher than those of control group while mRNA expression levels of Bax, caspase-9 as well as caspase-3 were significantly lower than that of control group (P < 0.05); proliferation activity value and mRNA expression of Bcl-2, TNF-α, IL-4, IL-6, YKL-40, protein expression of p-ERK1/2 of airway smooth muscle cell in GA group were significantly lower than those of BA group (P < 0.05) while the mRNA expression levels of Bax, caspase-9 as well as caspase-3 were significantly higher than those of BA group (P < 0.05); proliferation activity value and mRNA expression of Bcl-2, TNF-α, IL-4, IL-6, YKL-40 of airway smooth muscle cell in GA + LM group were significantly higher than those of GA group (P < 0.05) while mRNA expression levels of Bax, caspase-9 as well as caspase-3 were significantly lower that of GA group (P < 0.05).@*CONCLUSION@#GA can inhibit the proliferation of bronchial smooth muscle cells and reduce the expression of inflammatory factors by inhibiting the phosphorylation of ERK1/2.
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<p><b>OBJECTIVE</b>This study aimed to investigate the mechanism of cyclic stretch that promotesthe osteogenic differentiation of human periodontal ligament cells (hPDLCs) through the mediation of extracellular-signal-regulated kinase 1/2 (ERK1/2).</p><p><b>METHODS</b>hPDLCs were isolated through the explant method and cultured in vitro. hPDLCs were mechanically stimulated by a multi-channel cell-stress-loading system for 1, 3, 6, 12, and 24 h. The magnitude of stretch was 10% deformation, and the frequency was 0.5 Hz. Nonloaded cells were used as control group. ERK1/2 activation was blocked by U0126, a specific ERK1/2 pathway inhibitor. Additionally, hPDLCs were transfected with adenoviral vector encoding dominant negative ERK1/2 (DN-ERK1/2) to continuouslyinhibit ERK1/2 activation. The mRNA and protein levels of target geneswere detected through real-time polymerase chain reaction and Western blot.</p><p><b>RESULTS</b>Cyclic stretching promoted the expression of ERK1/2, osteocalcin (OCN) mRNA, and bone sialoprotein (BSP) mRNA. The expression of runt-related transcription factor (Runx) 2 protein and mRNA also increased at 3 and 6 h of cyclic stretching. The inhibition of ERK1/2 by U0126 and DN-ERK1/2 suppressed the expressionof Runx2 mRNA, OCN mRNA, BSP mRNA, Runx 2 protein, and p-ERK1/2 protein relative to that in stretched cells without the ERK1/2 inhibitor.</p><p><b>CONCLUSIONS</b>ERK1/2 is a critical molecule in the mediation ofthe osteogenic differentiation of hPDLCs under mechanical stimulation. ERK1/2 activation induced the elevation of Runx2 protein levels, which may be involved in the stretch-induced expressions of OCN and BSP.</p>
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Objective To investigate the regulation on extracellular signal regulated kinase (ERK )1/2 and p38 signaling pathways and immune expression by moist exposed burn therapy/moist exposed burn ointment(MEBT/MEBO)in diabetic foot and their relationship ,to explore the repair mechanism of MEBT/MEBO on diabetic foot ulcers .Methods Totally 40 diabetic foot pa‐tients were treated by MEBT/MEBO ,to take wound tissue before and after treatment and detect the expression of ERK 1/2 ,p38 , MAPKK6 ,c‐myc ,Akt ,ATF2 ,IgA ,IgM ,IgG ,C3c and C4c by immunohistochemistry ,to investigate their relationship .Results Af‐ter treatment with MEBT/MEBO ,the area of foot wounds in 39 patients was reduced in different degree .Only one patient had no obvious change .14 patients(35 .00% ) were markedly effective ,25 patients(62 .50% ) and 1 patient(2 .50% ) were ineffective .Before andaftertreatment,allpositiveexpression,positiveimmunoreactivity(anyindex)andpositiveexpression(specificindex)ofsignal pathway molecules were statistically significant (P<0 .05) .While the positive rate of molecular expression in wound pathway in‐creased ,the positive expression rate of immune factor increased .Before treatment ,a small amount of immune factors were found in the wound tissue .After treatment ,the immune factors IgA ,IgM ,IgG ,C3c ,C4c were distributed widely and diffusely .Before treat‐ment ,the wound tissue showed a very small number of signal molecules .After treatment ,the signal pathway molecules MEBT/MEBO and p38 ,MAPKK6 ,c‐myc ,Akt ,ATF2 showed broad and diffuse distribution .Conclusion MEBT/MEBO may promote the expression of ERK1/2 and p38 signaling molecules and immune in diabetic foot ,p38 and ERK1/2 signaling pathway may promote the healing of diabetic foot wound by increasing the expression of immune .