OBJECTIVETo detect protein expression of ERK(1), ERK(2), JNK(1), p38 and MEK(1), MEK(2) in human hepatocellular carcinoma and adjacent non-neoplastic liver.

METHODSIn 16 surgically resected hepatocellular carcinoma and para-carcinoma tissues, Western blotting was used to detect expression of ERK(1), ERK(2), JNK(1), p38 and MEK(1), MEK(2).

RESULTSIn all cases, ERK(1), ERK(2), p38 expression in hepatocellular carcinoma was significantly higher than that in para-carcinoma: integral optic density (IOD) of ERK(1) was 300 +/- 98 in carcinoma and 98 +/- 48 in para-carcinoma tissues (t = 2.519, P < 0.01); IOD of ERK(2) was 587 +/- 83 in carcinoma and 232 +/- 96 in para-carcinoma tissues (t = 2.745, P < 0.01); IOD of p38 was 270 +/- 85 in carcinoma and 107 +/- 88 in para-carcinoma tissues (t = 2.491, P < 0.01). JNK(1) expression in hepatocellular carcinoma was significantly lower than that in para-carcinoma; IOD of JNK(1) was 111 +/- 93 in carcinoma and 292 +/- 109 in para-carcinoma tissues (t = 2.473, P < 0.01). Protein levels of MEK(1) and MEK(2) in carcinoma were significantly higher than in para-carcinoma. IOD of MEK(1) was 1 418 +/- 244 in carcinoma and 806 +/- 90 in para-carcinoma tissues (t = 2.546, P < 0.01). IOD of MEK(2) was 1 041 +/- 122 in carcinoma and 468 +/- 40 in para-carcinoma tissues (t = 2.861, P < 0.01).

CONCLUSIONSERK(1), ERK(2), MEK(1) and MEK(2) in the signal transduction pathway for cell proliferation are significantly overexpressed and the expression of JNK(1) is lower in hepatocellular carcinoma. Their unbalance is one of the important reasons for the over growth and infinite proliferation of the hepatocellular carcinoma cell. The p38 and JNK(1) may be activated by different pathway.

Adult ; Aged ; Carcinoma, Hepatocellular ; enzymology ; Enzyme Activation ; Female ; Humans ; JNK Mitogen-Activated Protein Kinases ; Liver Neoplasms ; enzymology ; MAP Kinase Kinase 1 ; Male ; Middle Aged ; Mitogen-Activated Protein Kinase Kinases ; analysis ; Mitogen-Activated Protein Kinases ; metabolism ; Protein-Serine-Threonine Kinases ; analysis

Aluminum nanoparticles (Al-NPs) are one of the most widely used nanomaterial in cosmetics and medical materials. For this reason, Al-NP exposure is very likely to occur via inhalation in the environment and the workplace. Nevertheless, little is known about the mechanism of Al-NP neurotoxicity via inhalation exposure. In this study, we investigated the effect AL-NPs on the brain. Rats were exposed to Al-NPs by nasal instillation at 1 mg/kg body weight (low exposure group), 20 mg/kg body weight (moderate exposure group), and 40 mg/kg body weight (high exposure group), for a total of 3 times, with a 24-hr interval after each exposure. Inductively coupled plasma mass spectrometry (ICP-MS) analysis indicated that the presence of aluminum was increased in a dose-dependent manner in the olfactory bulb (OFB) and the brain. In microarray analysis, the regulation of mitogen-activated protein kinases (MAPK) activity (GO: 0043405), including Ptprc, P2rx7, Map2k4, Trib3, Trib1, and Fgd4 was significantly over-expressed in the treated mice than in the controls (p = 0.0027). Moreover, Al-NPs induced the activation of ERK1 and p38 MAPK protein expression in the brain, but did not alter the protein expression of JNK, when compared to the control. These data demonstrate that the nasal exposure of Al-NPs can permeate the brain via the olfactory bulb and modulate the gene and protein expression of MAPK and its activity.
Aluminum* ; Animals ; Body Weight ; Brain* ; Inhalation ; Inhalation Exposure ; Mass Spectrometry ; Mice ; Microarray Analysis ; Mitogen-Activated Protein Kinases ; Nanoparticles* ; Nanostructures ; Olfactory Bulb ; p38 Mitogen-Activated Protein Kinases ; Plasma ; Rats*

The ras, is a G-like protein that controls the mitogen-activated protein kinase (MAPK) pathway involved in control and differentiation of cell growth. MAPK is a key component of its signaling pathway and the aberrant activation may play an important role in the transformation process. To better understand roles of ras in the activation of MAPKs, we have established ras transformed NIH3T3 fibroblast cell line, and analyzed the MAPK module. The ras transformed cells formed numerous spikes at the edges of cells and showed loss of contact inhibition. The levels of ERK1/2 MAPKs as revealed by Western blot analysis were not significantly different between ras transformed and non-transformed cells. However, phosphorylation of ERK MAPKs and the level of MEK were significantly increased although the heavily expressed level of Raf-1, an upstream component of MAPK pathway was unchanged in ras transformed NIH3T3 cells. The sedimentation profile of the MAPK module kinases in a glycerol gradient showed the presence of a rather homogeneous species of multimeric forms of ERK1/2 and MEK as indicated by the narrow distribution peak areas. The broad sedimentation profile of the Raf-1 in a glycerol gradient may suggest possible heterologous protein complexes but the identification of interacting molecules still remains to be identified in order to understand the organization of the MAPK signal transduction pathway.
3T3 Cells ; Animal ; Cell Transformation, Neoplastic* ; Genes, ras* ; Mice ; Mitogen-Activated Protein Kinase Kinases/analysis ; Mitogen-Activated Protein Kinases/analysis ; Phosphorylation ; Protein-Serine-Threonine Kinases/analysis* ; Proto-Oncogene Proteins c-raf/analysis ; p42 MAP Kinase/analysis

OBJECTIVETo investigate the effects of mitogen activated protein kinase (MAPK) signal transduction pathways on heat shock protein 70 (HSP70) gene expression in endothelial cells exposed to benzo(a)pryene (BaP).

METHODSPorcine aortic endothelial cells were pre-treated or by PD98059 (10 micro mol/L) or SB203580 (20 micro mol/L) for 1 hour, then treated with different concentrations of BaP (0, 0.1, 0.5, 1.0, 5.0 and 10.0 micro mol/L) for 24 hours respectively;Expression levels of three phosphorylated MAPKs [extracellular signal regulated protein kinase (ERK), c-Jun amino-terminal kinase (JNK), and p38] and HSP70 were determined by Western-blot.

RESULTSThe three phosphorylated MAPKs expressional levels especially p-ERK1 had different extents of changes with dose-response relationship under BaP exposure. BaP inhibited the expression of HSP70, which significantly decreased in medium and high dose group (>or= 1.0 micro mol/L) but did not decrease in control group (P < 0.05). Although the inhibitor of ERK (PD98059) could partly weaken the inhibited effects of BaP on HSP70 expression, HSP70 expression levels of endothelial cells pre-treated with PD98059 were still significantly lower than that of control cells (P < 0.05).

CONCLUSIONERK1 pathway might play some roles in HSP70 gene expression in endothelial cells exposed to BaP, and other unknown signal pathways might also have some effects on this process.

Animals ; Benzo(a)pyrene ; toxicity ; Blotting, Western ; Dose-Response Relationship, Drug ; Endothelial Cells ; drug effects ; metabolism ; Enzyme Inhibitors ; pharmacology ; Flavonoids ; pharmacology ; HSP70 Heat-Shock Proteins ; analysis ; Imidazoles ; pharmacology ; JNK Mitogen-Activated Protein Kinases ; MAP Kinase Kinase 4 ; Mitogen-Activated Protein Kinase Kinases ; analysis ; Mitogen-Activated Protein Kinases ; analysis ; antagonists & inhibitors ; Pyridines ; pharmacology ; Signal Transduction ; physiology ; Swine ; p38 Mitogen-Activated Protein Kinases

Animals ; DNA-Binding Proteins ; analysis ; Liver Cirrhosis, Experimental ; etiology ; metabolism ; Male ; Mitogen-Activated Protein Kinase 3 ; Mitogen-Activated Protein Kinases ; analysis ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; Smad3 Protein ; Trans-Activators ; analysis

OBJECTIVETo explore the functional effects of MAPK pathway in the pathogenesis of human osteosarcoma.

METHODSGene microarray (Human Genome U133A, Affymetrix) was used to screen the differential expression of genes involved in MAPK pathway between osteosarcoma cell lines and 3 osteoblastic cell lines. KEGG metabolic pathway analysis was performed among significantly increased or decreased genes using the MATLAB software. Immunohistochemical technique was used to detect the expressions of ERK1/2, JNK and p38 proteins among 48 osteosarcoma and benign 24 osteoblastic tumor samples.

RESULTSUsing an entrance limit of > or = 2.0, 18 differentially expressed MAPK pathway-related genes were selected (10 up-regulated, 8 down-regulated) to mapped to the MAPK pathway of KEGG which are all important node genes. The positive rates of ERK1/2, JNK and p38 proteins were 83.3% (40/48), 72.9% (35/48) and 85.4% (41/48) in osteosarcomas,and 12.5% (3/24), 8.3% (2/24) and 16.7% (4/24) in the control group, respectively. The positive rates and expression intensities were statistically different between the 2 groups (P<0.01).

CONCLUSIONMAPK pathway plays an important role in the pathogenesis of osteosarcoma. ERK, JNK and p38 form an intercoordinating network and regulate the cell proliferation, differentiation, apoptosis, invasion and migration in osteosarcoma.

Adolescent ; Adult ; Aged ; Bone Neoplasms ; genetics ; metabolism ; pathology ; Cell Line, Tumor ; Child ; Female ; Gene Expression Profiling ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Male ; Middle Aged ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Mitogen-Activated Protein Kinases ; metabolism ; Oligonucleotide Array Sequence Analysis ; Osteoblastoma ; genetics ; metabolism ; pathology ; Osteosarcoma ; genetics ; metabolism ; pathology ; Signal Transduction ; Young Adult ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVESTo explorer the change of several signal pathway and their signal after liver transplantation.

METHODSClassified 34 punctured donor liver samples and 10 normal liver samples as A (no rejection) groups, B (mild/moderate acute rejection) groups, C (serious acute rejection) groups, D (chronic rejection/fibrosis) groups and E (control) groups, MAPK, Ras and p53 were performed immunohistochemistry analysis and image analysis. MAPK and Ras were performed in situ hybridizition. Then image analysis was performed.

RESULTSThe protein expression of MAPK, Ras, increase by turns of A, B and C groups (1.42+/-0.28, 3.88+/-0.87, 6.68+/-0.57 in MAPK; 1.27+/-0.12, 2.80+/-0.30, 3.93+/-0.20 in Ras; corresponding), and decrease by turns of D and E groups (1.49+/-0.37, 0.88+/-0.20 in MAPK; 1.47+/-0.21, 1.01+/-0.12 in Ras; corresponding, F=178.39 in MAPK and 320.59 in Ras, groups B, C vs groups A, D, E, P<0.001 in MAPK and Ras), The protein expression of p53 is higher in treated groups (The results of groups A to E are 2.09+/-0.13, 2.39+/-0.11, 2.03+/-0.19, 2.26+/-0.18 and 0.35+/-0.08, corresponding, F=360.08, groups E vs groups A, B, C, D, P<0.001). Expression of MAPK, Ras mRNA is as same as that of protein.

CONCLUSIONThe MAPKs pathway has role in rejection response after liver transplantation. And it seemed that the MAPKs and p53 are one regulation mechanism for protecting the hepatocyte from damage after liver transplantation.

Humans ; Immunohistochemistry ; In Situ Hybridization ; Liver Transplantation ; MAP Kinase Signaling System ; Mitogen-Activated Protein Kinases ; analysis ; Signal Transduction ; physiology ; Tumor Suppressor Protein p53 ; analysis ; ras Proteins ; analysis

OBJECTIVETo observe the effects of hippocampal Abeta42 deposition on the expression of inflammatory cytokines and phosphorylated MAPK signal molecules as well as the intervention of AD by total glucosides of paeony (TGP).

METHOD12 week-old female SD rats were stereotactic injected one-time with a fibrillar Abeta42 positioning hippocampus to replicate AD pathology model and interfered with TGP. The expression of inflammatory cytokines and phosphorylated MAPK pathway signaling molecules were observed by immunohistochemistry (SABC), and SABC images were analyzed by image analysis software.

RESULTCompared with the control group, the IL-1beta, IL-6 and p-p38, p-JNK, p-MEK3/6 positive stained areas of AD pathology model group increased and their staining intensity decreased (the protein expression quantity inversely proportional to the staining intensity), while the IL-1beta, IL-6 and p-p38, p-JNK, p-MEK3/6 positive stained areas of the treatment groups decreased and their staining intensity increased compared with AD pathology model group.

CONCLUSIONAbeta42 deposition in hippocampus can induce the brain inflammation and the over-expression of IL-1beta, IL-6 and p-p38, p-JNK, p-MEK3/6. Inhibiting the over-expression of inflammatory cytokines and phosphorylated MAPK signaling molecules may be a major antagonistic mechanism of TGP against AD.

Alzheimer Disease ; drug therapy ; Amyloid beta-Peptides ; metabolism ; toxicity ; Animals ; Cytokines ; analysis ; Female ; Glucosides ; pharmacology ; therapeutic use ; Hippocampus ; metabolism ; JNK Mitogen-Activated Protein Kinases ; metabolism ; MAP Kinase Signaling System ; drug effects ; Mitogen-Activated Protein Kinases ; metabolism ; Paeonia ; chemistry ; Peptide Fragments ; metabolism ; toxicity ; Phosphorylation ; Rats ; Rats, Sprague-Dawley ; p38 Mitogen-Activated Protein Kinases ; metabolism

Epidermal keratinocyte differentiation is a tightly regulated stepwise process that requires protein kinase C (PKC) activation. Studies on cultured mouse keraitnocytes induced to differentiate with Ca2+ have indirectly implicated the involvement of PKC alpha isoform. When PKC alpha was overexpressed in undifferentiated keratinocytes using adenoviral system, expressions of differentiation markers such as loricrin, filaggrin, keratin 1 (MK1) and keratin 10 (MK10) were increased, and ERK1/2 phosphorylation was concurrently induced without change of other MAPK such as p38 MAPK and JNK1/2. Similarly, transfection of PKC alphakinase active mutant (PKC alpha- CAT) in the undifferentiated keratinocyte, but not PKC beta-CAT, also increased differentiation marker expressions. On the other hand, PKC alphadominant negative mutant (PKC beta-KR) reduced Ca2+ -mediated differentiation marker expressions, while PKC beta-KR did not, suggesting that PKC alphais responsible for keratinocyte differentiation. When downstream pathway of PKC alphain Ca2+ - mediated differentiation was examined, ERK1/2, p38 MAPK and JNK1/2 phosphorylations were increased by Ca2+ shift. Treatment of keratinocytes with PD98059, MEK inhibitor, and SB20358, p38 MAPK inhibitor, before Ca2+ shift induced morphological changes and reduced expressions of differentiation markers, but treatment with SP60012, JNK1/2 inhibitor, did not change at all. Dominant negative mutants of ERK1/2 and p38 MAPK also inhibited the expressions of differentiation marker expressions in Ca2+ shifted cells. The above results indicate that both ERK1/2 and p38 MAPK may be involved in Ca2+- mediated differentiation, and that only ERK1/2 pathway is specific for PKCa-mediated differentiation in mouse keratinocytes.
Animals ; Calcium/pharmacology/physiology ; Cell Differentiation/physiology ; Intermediate Filament Proteins/analysis/metabolism ; Keratinocytes/cytology/*enzymology ; Membrane Proteins/analysis/metabolism ; Mice ; Mitogen-Activated Protein Kinase 1/*metabolism ; Mitogen-Activated Protein Kinase 3/*metabolism ; Phosphorylation ; Protein Kinase C/genetics/*physiology ; Research Support, Non-U.S. Gov't ; p38 Mitogen-Activated Protein Kinases/metabolism

OBJECTIVETo examine the effect of SH2A gene in cell signal transduction and its subcellular localization.

METHODSRT-PCR method was used to amplify the coding sequence of SH2A gene. Eukaryotic recombined expression vector pcDNA 3.1-SH2A was constructed, and then Bel7402 cell and COS7 cell transfected by liposome. Multiple kinase assay was performed to examine the activity of protein kinase (PKC), mitogen activated protein kinase (MAPK), tyrosine protein kinase (TPK) in the transfected cells. Meantime, pEGFP-SH2A vector was also constructed and the cells transfected with it were examined by fluorescent microscopy.

RESULTSRecombined expression vector pcDNA3.2-SH2A and pEGFP-SH2A contained the coding sequence of SH2A cDNA. In both cell lines expressing SH2A gene, the cytoplasm PKC activity decreased by 40% or so, but no apparent alteration was found in MAPK and TPK activity. SH2A gene was found localized in the cytoplasm of transfected cells under fluorescent microscope.

CONCLUSIONSH2A gene may act as an inhibiting factor in PKC signal transduction, and it is localized in cytoplasm.

Animals ; COS Cells ; Cytoplasm ; chemistry ; Humans ; Membrane Proteins ; analysis ; genetics ; physiology ; Mitogen-Activated Protein Kinases ; metabolism ; Protein Kinase C ; physiology ; Signal Transduction ; physiology ; Transfection