OBJECTIVETo study the anti-cancer effect of matrine (Mat) on U937 cell line and its possible molecular mechanism.

METHODThe cells were cultured in medium containing either 0.1, 0.2, 0.3, 0.4, or 0.5 g x L(-1) of Mat. The morphological alteration was observed by inverted microscopy and electron microscopy. Cell proliferation was analyzed by Try pan blue staining and MTT. The method of Western Blot was used to detect phosphorylation activity of MAPK.

RESULTMatrine had a significant inhibitory effect on proliferation of U937 cell line at the concentration of 0.2 g x L(-1). Treated with matrine of 0.2 g x L(-1) for 48 h, U937 cells became smaller and appeared more round than previously. The number of U937 cells showing apoptosis increased with elevation of the concentration of the matrine. Matrine had an ability of inhibiting the activity of ERK and increasing the activities of p38 and JNK to some degree in U937 cells.

CONCLUSIONMatrine can inhibit the proliferation of U937 cell line in vitro and induce its apoptosis possibly through inhibiting the activity of ERK and increasing the activities of p38 and JNK in U937 cells.

Alkaloids ; pharmacology ; Apoptosis ; drug effects ; Cell Proliferation ; drug effects ; Humans ; MAP Kinase Signaling System ; drug effects ; Quinolizines ; pharmacology ; U937 Cells

OBJECTIVETo explore the effects of MAPK antagonist on TPO stimulated UT7 cell proliferation and differentiation, and to elucidate the mechanism of TPO functioning on UT7 cells.

METHODSEGFP pMSCV and MEK 1 pMSCV MEK 1 plasmids were transferred into UT7 cells. Phosphorylated MEK1 of UT7 cells was examined by Western blot. The proliferation and CD41 expression of UT7 cells transfected with mutant (ser222A) MEK1 or exposed to PD98059 were examined.

RESULTS(1) 60.73% EGFP positive cells were obtained in retroviral vector MEK1 pMSCV transfected UT7cells. (2) In different time of TPO stimulating UT7 cells, the level of phosphorylated MEK1 was lower in experiment group than in control group. In experiment group, the level of phosphorylated MEK1 was decreased after stimulated by TPO for 1 hour, and almost disappeared after stimulated for 3 hours. (3) The effect of TPO on UT7 cell proliferation was inhibited by PD98059 and the transfected mutation MEK1 gene. The proliferation rate was 98.58% in DMSO control group, 39.00% in PD98059 group (P < 0.05), 102.13% in EGFP pMSCV group, and 48.94% in MEK1pMSCV (P < 0.05). (4) The CD41 expression on UT7 was inhibited by PD98059 and the transfected mutation MEK1 gene.

CONCLUSIONPhosphorylation of MEK1 in UT7 cells can be induced by TPO. There was a relationship between the TPO stimulating time and phosphorylation of MEK1. The effects of TPO on UT7 cell proliferation and CD41 expression is mediated by MAPK signal transduction pathway.

Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Flavonoids ; pharmacology ; Humans ; MAP Kinase Kinase 1 ; drug effects ; metabolism ; MAP Kinase Signaling System ; drug effects ; Megakaryocytes ; cytology ; drug effects ; metabolism

Animals ; Apoptosis ; Endotoxins/toxicity* ; Heart/drug effects* ; MAP Kinase Signaling System ; Myocardium/pathology* ; Rats ; Rats, Wistar ; Receptors, Calcium-Sensing/metabolism*

BACKGROUNDObstructive sleep apnea (OSA) has been recognized as an independent risk factor for systemic hypertension. The study investigated the functional consequences of chronic intermittent hypoxia (CIH) on aortic constriction induced by angiotensin II (Ang II) and the possible signaling involving ERK1/2 and contractile proteins such as myosin light chain kinase (MLCK), myosin phosphatase targeting subunit (MYPT1) and myosin light chain (MLC).

METHODSMale Wistar rats were randomly divided into CIH group and normoxia group and exposed to either CIH procedure or air-air cycles. Phosphorylation of ERK1/2, MYPT1 and MLC was assessed by Western blotting following constrictor studies in the presence or absence of PD98059 (10 µmol/L).

RESULTSCIH-exposure resulted in more body weight gain and elevated blood pressure, which could be attenuated by pretreatment with PD98059. Endothelium-removed aortic rings from CIH rats exhibited higher constrictor sensitivity to Ang II (Emax: (138.56 ± 5.78)% versus (98.45±5.31)% of KCl; pD2: 7.98 ± 0.14 versus 8.14 ± 0.05, respectively). CIH procedure exerted complex effects on ERK expressions (total ERK1/2 decreased whereas the ratio of phosphorylated to total ERK1/2 increased). CIH aortas had higher MLCK mRNA and basal phosphorylation of MYPT1 and MLC. In parallel to greater increases in phosphorylation of ERK1/2, MYPT1 and MLC, Ang II-induced aortic constriction was significantly enhanced in CIH rats, which was largely reversed by PD98059. However vascular constriction of normoxia rats remained unchanged despite similar but smaller changing tendency of proteins phosphorylation.

CONCLUSIONThese data suggest that CIH exposure results in aortic hyperresponsiveness to Ang II, presumably owing to more activated ERK1/2 signaling pathway.

Angiotensin II ; pharmacology ; Animals ; Aorta ; drug effects ; Flavonoids ; pharmacology ; Hypoxia ; physiopathology ; MAP Kinase Signaling System ; drug effects ; Male ; Phosphorylation ; drug effects ; Rats ; Rats, Wistar ; Vasoconstriction ; drug effects

The study was aimed to investigate the effects of bortezomib (BTZ) on the expression of ERK, JNK and P38 in daunorubicin (DNR)-resistant K562 cells (K562/DNR) and to clarify the molecular mechanism of BTZ in reversing the drug-resistance in leukemic cells. The K562/DNR cells and the cellular toxicity of BTZ was determined by MTT, then 4 µg/L of BTZ was chosen to do the experiment. The expression of ERK, JNK, p38 and P-gp of K562/DNR cells treated with DNR only or DNR combined with BTZ for 12, 24 and 36 hours was detected by Western blot. The apoptosis rate in each group was assayed by flow cytometry. The results showed that as compared with DNR group, the expression of P-ERK, P-P38 and P-gp was significantly suppressed (p < 0.05) and the expression of P-JNK was significantly enhanced (p < 0.05) in the cells treated with DNR combined with BTZ. There was no change in the expression of total ERK, P38 and JNK. The effect increased with the prolonging of time. Meanwhile, the apoptosis rate in cells treated with DNR combined with BTZ increased compared with DNR only. It is concluded that the BTZ can reverse the drug resistance in K562/DNR cells by MAPK signaling pathway and increase the apoptosis of leukemic cells. The effect shows the characteristics of time-dependent manner.
Apoptosis ; drug effects ; Boronic Acids ; pharmacology ; Bortezomib ; Drug Resistance, Multiple ; drug effects ; Drug Resistance, Neoplasm ; drug effects ; Humans ; K562 Cells ; MAP Kinase Signaling System ; Pyrazines ; pharmacology

OBJECTIVETo explore the effect of reinforcing qi for resolving masses method (RQRMM) on expressions of extracellular signal regulated kinase 2 (MEK2) and phosphorylation extracellular signal regulated kinase (p-ERK) protein in estrogen induced uterine leiomyoma model Guinea pigs' uterine tissue.

METHODSGuinea pigs were randomly divided into five groups, i.e., the model group, the high dose group, the middle dose group, the low dose group, and the Western medicine group (mifepristone). The normal control group was set up. The uterine leiomyoma model in guinea pigs was established by castrating and subcutaneous injecting estradiol (E2). The protein expression levels of MEK2 and p-ERK of guinea pigs' uterine tissues were detected by immunohistochemical assay.

RESULTSThe protein expressions of MEK2 and p-ERK in the uterine muscular tissue of Guinea pigs' uterine tissue were higher in the model group than in the normal group (P < 0.01). They decreased to some degree in the high dose group, the middle dose group, and the low dose group. Of them, the protein expressions of MEK2 and p-ERK were significantly lower in the high dose group than in the model group and the Western medicine group (P < 0.01).

CONCLUSIONRQRMM could treat uterine leiomyoma possibly through intervening the MAPK/ERK cell signal pathway to inhibit the proliferation of myoma cells.

Animals ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Estrogens ; adverse effects ; Female ; Guinea Pigs ; MAP Kinase Kinase 2 ; metabolism ; MAP Kinase Signaling System ; drug effects ; Phytotherapy ; methods ; Signal Transduction ; Uterine Neoplasms ; chemically induced ; drug therapy ; metabolism ; Uterus ; metabolism

The purpose of this study was to investigate the effect and molecular mechanism of metformin (Met) on biological characteristics of acute promyelocytic leukemia (APL) cell line NB4. NB4 cells were treated with various concentrations of Met for different time, MTT method was used to detect cell proliferation, the alteration of cell apoptosis was analyzed by flow cytometry, and the change of cell adhesion ability was examined by cell adhesion assay. NB4 cells were pretreated with U0126, a specific inhibitor for extracellular signal-regulated kinase (ERK) phosphorylation, ERK phosphorylation was assessed by Western blot analysis, apoptosis and cell adhesion ability were evaluated by flow cytometry and cell adhesion test respectively. The results showed that Met could inhibit the cell proliferation, induce the cell apoptosis and increase the ability of cell adhesion. The pretreatment of NB4 cells with 5 µmol/L U0126 could effectively inhibit the phosphorylation of ERK, and reduce cell apoptosis and adhesion induced by 5 mmol/L Met. It is concluded that Met can inhibit the proliferation and promote the apoptosis and adhesion of NB4 cells. MEK/ERK signaling pathway may be one of the molecular mechanisms of metformin on NB4 cells.
Apoptosis ; drug effects ; Cell Adhesion ; drug effects ; Cell Line, Tumor ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Humans ; Leukemia, Promyelocytic, Acute ; metabolism ; pathology ; MAP Kinase Signaling System ; drug effects ; Metformin ; pharmacology ; Mitogen-Activated Protein Kinase Kinases ; metabolism ; Phosphorylation

OBJECTIVETo explore the effect of extracellular signal regulated kinase 1/2 (ERK1/2) on edaravone (EDA)-triggered protection against myocardial toxicity induced by isoprenaline (ISO) in H9c2 myocardial cells (H9c2 cells).

METHODSH9c2 cells were exposed to ISO at different concentrations to establish a cardiac toxicity model induced by persistent excitation of β1 receptor. EDA was added before ISO as a pretreatment. PD-98059, an ERK1/2 inhibitor, was administered 1 h prior to EDA to inhibit the phosphorylation of ERK1/2. Cell viability was measured using cell counter kit (CCK-8). The expressions of p-ERK1/2 and t-ERK1/2 were tested by Western blotting. Mitochondrial membrane potential (MMP) was detected by Rhodamine123 (Rh123) staining and photofluorography.

RESULTSExposure of H9c2 cells to 80 µmol/L ISO for 24 h down-regulated ERK1/2 phosphorylation and repressed MMP. Pretreatment with 10-40 µmol/L EDA for 1 h inhibited ISO-induced myocardial toxicity and pretreatment of 40 µmol/L EDA partially rescued ERK1/2 phosphorylation and MMP level. PD-98059 abolished cardiac protection of EDA, leading to myocardial toxicity and MMP loss.

CONCLUSIONEDA can protect H9c2 cells against myocardial injury induced by ISO by suppressing ISO-triggered inhibition of ERK1/2 activation.

Animals ; Antipyrine ; analogs & derivatives ; pharmacology ; Cell Line ; Flavonoids ; pharmacology ; Isoproterenol ; toxicity ; MAP Kinase Signaling System ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Myocytes, Cardiac ; drug effects ; metabolism ; Phosphorylation ; Rats

OBJECTIVETo explore the role of the P38 signaling pathway in the apoptosis of arsenic trioxide (As2 O3)-induced androgen-independent prostate cancer PC-3 cells.

METHODSAndrogen-independent prostate cancer PC-3 cells were treated with different concentrations of As2 O3 for 24, 48 and 72 hours. The inhibitory effect of As2 O3 on the cell growth was measured by MTT, the expression of p- P38 detected by Western blot, and the rate of cell apoptosis determined by Annexin V and PI double staining before and after interfering the P38 signaling pathway by SB203580, a highly selective P38 inhibitor.

RESULTSAs2 O3 inhibited the proliferation of PC-3 cells in a concentration- and time-dependent manner, and quickly activated P38 phosphorylation, thus giving full play to its biological activities. After 24 hours of treatment with As2 O3 at the concentrations of 2, 10 and 20 micromol/L, the apoptosis rates of the PC-3 cells were (18.9 +/- 0.43), (24.7 +/- 0.29) and (49.7 +/- 1.79)%, respectively, which were reduced to (14.8 +/- 0.81), (22.1 +/- 0.51) and (39.6 +/- 1.74)% after interfering the P38 pathway with SB203580. Inhibition of the P38 pathway significantly reduced the apoptosis of the PC-3 cells induced by As2 O3 (P < 0.05).

CONCLUSIONAs2 O3 can induce the apoptosis of prostate cancer PC-3 cells by activating the P38 signaling pathway, and interfering the P38 signaling pathway can reduce their apoptosis, which suggests that the P38 signaling pathway is involved in the apoptosis of As2 O3-induced androgen-independent prostate cancer PC-3 cells.

Arsenicals ; pharmacology ; Cell Line, Tumor ; Humans ; MAP Kinase Signaling System ; drug effects ; Male ; Oxides ; pharmacology ; Prostatic Neoplasms ; pathology ; Signal Transduction ; drug effects ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVETo investigate the effects of GnRH analogues GnRHa and GnRHant on the MAPK pathway in rat Leydig cells.

METHODSRat Leydig cells were primarily cultured for 24 hours in vitro and serum-starved for 2 hours, followed by treatment with GnRHa (10(-7) mol/L) or GnRHant (10(-6) mol/L) for 0, 5, 15, 30, 60 and 90 minutes, with the 0 min group as the control. Then the protein levels of phosphorylated ERK (p-ERK) and phosphorylated p38 (p-p38) were detected by Western blot, and that of p-ERK determined by the same means after co-incubation of GnRHa or GnRHant with the PKC inhibitor GF109203X at 1, 5, 10 and 20 micromol/L.

RESULTSAfter stimulation of the Leydig cells with GnRHa or GnRHant for different times, the protein level of p-p38 showed no significant difference from that of the control group (P > 0.05). Then the Leydig cells were treated with GF109203X at different concentrations for 20 minutes and with addition of GnRHa for another 10 minutes. The level of p-ERK was significantly decreased (P < 0.05) by GF109203X at 10 and 20 micromol/L. Compared with the control, the p-ERK expression was increased by 65% at 15 minutes (P < 0.05) in the GnRHant stimulation group, by 81% (to the peak) at 30 minutes (P < 0.05), began to fall at 60 minutes, and returned to the base level at 90 minutes. The p-ERK level exhibited no significant difference from that of the control (P > 0.05) after treatment of the Leydig cells with different concentrations of GF109203X for 20 minutes and then with GnRHant for 30 minutes.

CONCLUSIONThe ERK MAPK activation induced by GnRHa depends on the PKC pathway, but not that induced by GnRHant. The p-38 MAPK pathway may not be involved in the effect of GnRH analogues on rat Leydig cells.

Animals ; Cells, Cultured ; Gonadotropin-Releasing Hormone ; analogs & derivatives ; pharmacology ; Leydig Cells ; drug effects ; metabolism ; MAP Kinase Signaling System ; drug effects ; Male ; Rats ; Rats, Sprague-Dawley