OBJECTIVETo explore the potential reporter gene assay for the detection of sodium channel-specific toxins in shellfish as an alternative for screening harmful algal bloom (HAB) toxins, considering the fact that the existing methods including HPLC and bioassay are inappropriate for identifying HAB toxins which poses a serious problem on human health and shellfish industry.

METHODSA reporter plasmid pEGFP-c-fos containing c-fos promoter and EGFP was constructed and transfected into T24 cells using LipofectAMINE 2000. Positive transfectants were screened by G418 to produce a pEGFP-c-fos-T24 cell line. After addition of increasing neurotoxic shellfish poison (NSP) or GTX2,3, primary components of paralytic shellfish poison (PSP), changes in expression of EGFP in the cell line were observed under a laser scanning confocal microscope and quantified with Image-pro Plus software.

RESULTSDose-dependent changes in the intensity of green fluorescence were observed for NSP in a range from 0 to 10 ng/mL and for GTX2,3 from 0 to 16 ng/mL.

CONCLUSIONpEGFP-c-fos-T24 can be applied in detecting HAB toxins, and cell-based assay can be used as an alternative for screening sodium channel-specific HAB toxins.

Animals ; Biological Assay ; Cell Line, Tumor ; Genes, Reporter ; physiology ; Green Fluorescent Proteins ; Harmful Algal Bloom ; physiology ; Humans ; Plasmids ; Proto-Oncogene Proteins c-fos ; genetics ; metabolism ; Shellfish ; analysis ; Sodium Channels ; Toxins, Biological ; chemistry ; toxicity

OBJECTIVETo investigate the effects of transforming growth factor beta (TGF ) on c-fos gene expression in hepatic stellate cells.

METHODSHepatic stellate cells (HSC-T6) were cultured in the medium containing different concentrations of TGF (0.2, 1, and 5 ng/ml), and cells were collected at different time points of incubation (8, 24, 48, and 72 h). The total RNA of the HSCs was isolated and c-fos gene expression level were measured by reverse transcription polymerase chain reaction.

RESULTSc-fos gene expression levels of HSCs cultured in the presence of low (0.2 ng/ml), moderate (1 ng/ml) and high (5 ng/ml) concentrations of TGF for 8, 24, 48 and 72 h were significantly greater than those of control group. The c-fos gene expression levels of HSCs increased gradually with the increment of TGF concentration, and significant differences in c-fos gene expression were found between the 3TGF groups.

CONCLUSIONTGF strongly up-regulates c-fos gene expression in hepatic stellate cells.

Animals ; Cells, Cultured ; Gene Expression ; drug effects ; Genes, fos ; Hepatic Stellate Cells ; drug effects ; Proto-Oncogene Proteins c-fos ; genetics ; metabolism ; Rats ; Transforming Growth Factor beta ; pharmacology

OBJECTIVETo investigate the changes of proto-oncogene c-fos, c-jun mRNA expression in angiotensin II (Ang II)-induced hypertrophy and effects of tanshinone II A (Tan) in the primary culture of neonatal rat cardiomyocytes.

METHODTwelve neonatal Wistar rats aged one day old of clean grade and both sexes were selected to isolate and culture cardiomyocytes. The cardiomyocytes were divided into: normal control group, Ang II (10(-6) mol x L(-1)) group, Ang II (10(-6) mol x L(-1)) +Tan (10(-8) g x L(-1)) group, Ang II (10(-6) mol x L(-1)) + valsartan (10(-6) mol x L(-1)) group, Tan (10(-8) g x L(-1)) group, valsartan (10(-6) mol x L(-1)) group. The cardiomyocyte size was determined by phase contrast microscope, the rate of protein synthesis in cardiomyocytes was measured by 3H-leucine incorporation. The c-fos, c-jun mRNA expression of cardiomyocytes were assessed using reverse transcription polymerase chain reaction (RT-PCR).

RESULTAng II was added to the culture medium and 30 min later, the c-fos, c-jun mRNA expression of cardiomyocytes increased significantly (P < 0. 01). After Ang II took effect for 24 h, the rate of protein synthesis in Ang II group increased more prominently than that in normal control group (P < 0.01). After Ang II took effect for 7 days, the size of cardiomyocyte in Ang II group increased obviously (P < 0. 05). If tanshinone II or valsartan was added to the culture medium before Ang II, both of them could inhibit the increase of c-fos, c-jun mRNA expression (P < 0.01), cardiomyocyte protein synthesis rate (P < 0.01), and cardiomyocyte size (P < 0.05) induced by Ang II.

CONCLUSIONTanshinone II could ameliorate Ang II-induced cardiomyocytes hypertrophy by inhabiting c-fos, c-jun mRNA expression.

Angiotensin II ; biosynthesis ; pharmacology ; Animals ; Cardiomegaly ; chemically induced ; metabolism ; pathology ; Diterpenes, Abietane ; Gene Expression Regulation ; drug effects ; Genes, fos ; genetics ; Genes, jun ; genetics ; Myocytes, Cardiac ; drug effects ; metabolism ; pathology ; Phenanthrenes ; pharmacology ; Proto-Oncogene Proteins c-fos ; genetics ; Proto-Oncogene Proteins c-jun ; genetics ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar ; Tetrazoles ; pharmacology ; Valine ; analogs & derivatives ; pharmacology ; Valsartan

Exposure to light can induce photoreceptor cell death and exacerbate retinal degeneration. In this study, mice with genetic knockout of several genes, including rhodopsin kinase (Rhok-/-), arrestin (Sag-/-), transducin (Gnat1-/-), c-Fos (c-Fos-/-) and arrestin/transducin (Sag-/-/Gnat1-/-), were examined. We measured the expression levels of thousands of genes in order to investigate their roles in phototransduction signaling in light-induced retinal degeneration using DNA microarray technology and then further explored the gene network using pathway analysis tools. Several cascades of gene components were induced or inhibited as a result of corresponding gene knockout under specific light conditions. Transducin deletion blocked the apoptotic signaling induced by exposure to low light conditions, and it did not require c-Fos/AP-1. Deletion of c-Fos blocked the apoptotic signaling induced by exposure to high intensity light. In the present study, we identified many gene transcripts that are essential for the initiation of light-induced rod degeneration and proposed several important networks that are involved in pro- and anti-apoptotic signaling. We also demonstrated the different cascades of gene components that participate in apoptotic signaling under specific light conditions.
Animals ; Apoptosis/radiation effects ; G-Protein-Coupled Receptor Kinase 1/genetics ; GTP-Binding Protein alpha Subunits/genetics ; *Gene Expression Profiling ; Genes, fos/genetics ; Light/adverse effects ; Light Signal Transduction/*genetics/physiology/radiation effects ; Mice ; Mice, Knockout ; Oligonucleotide Array Sequence Analysis ; Retina/metabolism/pathology/radiation effects ; Retinal Degeneration/etiology/*genetics/physiopathology ; Transducin/genetics

OBJECTIVE: To characterize the antisense c-fos oligonucleotides that control the expression of immediate-early gene c-fos in retina in order to better understand the mechanism by which antisense c-fos oligonucleotides induced myopia. In this study the signal transduction in the pathway linking visual experience and the regulation of the eye's growth was investigated. METHODS: Thirty-one 3-week guinea pigs were assigned into 3 groups: antisense and sense c-fos oligonucleotides were intravitreally injected every 3 days to the eyes of the experimental guinea pigs at different concentrations; and saline vehicle to control guinea pigs in the same way. The refraction and axial length of the eyes were measured before and after the treatment, and the immediate-early gene c-fos expression in the retina was quantified by immunohistochemistry and RT-PCR. RESULTS: The moderate myopia was induced in high (1 nmol) and low (0.1 nmol) level of antisense c-fos oligonucleotide intravitreous injection (-5.425 D and -5.575 D, respectively) compared with the control ateral eyes. The refraction and axial length of the treated eyes increased, and the expression of immediate-early gene c-fos decreased significantly in the antisense c-fos oligonucleotides intravitreously injected eyes compared with the sense c-fos oligonucleotide intravitreously and saline vehicle injected eyes (P<0.01). The refraction and axial length were of no statistically significant differences among the sense c-fos oligonucleotides-treated eyes and saline-treated eyes and non-treated eyes (P>0.05). CONCLUSION The obvious myopia can be induced by antisense c-fos oligonucleotides in guinea pigs; antisense c-fos oligonucleotides inhibit c-fos expression in the retina. Immediate-early gene c-fos may be a potential factor in the prevention of myopia and plays an important role in the signal transduction of the retina.
Animals ; Genes, Immediate-Early ; genetics ; Guinea Pigs ; Immunohistochemistry ; Microinjections ; Myopia ; chemically induced ; genetics ; physiopathology ; Oligonucleotides, Antisense ; administration & dosage ; genetics ; toxicity ; Proto-Oncogene Proteins c-fos ; biosynthesis ; genetics ; RNA, Messenger ; genetics ; metabolism ; Random Allocation ; Retina ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction ; physiology

OBJECTIVETo study the effects of selenium on DNA damage, apoptosis and c-myc, c-fos, and c-jun expression in rat hepatocytes.

METHODSSodium selenite at the doses of 5, 10, and 20 micromol/kg was given to rats by i.p. and there were 5 male SD rats in each group. Hepatocellular DNA damage was detected by single cell gel electrophoresis (or comet assay). Hepatocellular apoptosis was determined by TUNEL (TdT-mediated dUTP nick end labelling) and flow cytometry. C-myc, c-fos, and c-jun expression in rat hepatocytes were assayed by Northern dot hybridization. C-myc, c-fos, and c-jun protein were detected by immunohistochemical method.

RESULTSAt the doses of 5, 10, and 20 micromol/kg, DNA damage was induced by sodium selenite in rat hepatocytes and the rates of comet cells were 34.40%, 74.80%, and 91.40% respectively. Results also showed an obvious dose-response relationship between the rates of comet cells and the doses of sodium selenite (r=0.9501, P<0.01). Sodium selenite at the doses of 5, 10, and 20 micromol/kg caused c-myc, c-fos, and c-jun overexpression obviously. The positive brown-yellow signal for proteins of c-myc, c-fos, and c-jun was mainly located in the cytoplasm of hepatocytes with immunohistochemical method. TUNEL-positive cells were detected in selenium-treated rat livers. Apoptotic rates (%) of selenium-treated liver cells at the doses of 5, 10, and 20 micromol/kg were (3.72 +/- 1.76), (5.82 +/- 1.42), and (11.76 +/- 1.87) respectively, being much higher than those in the control. Besides an obvious dose-response relationship between apoptotic rates and the doses of sodium selenite (r=0.9897, P<0.01), these results displayed a close relationship between DNA damage rates and apoptotic rates, and the relative coefficient was 0.9021, P<0.01.

CONCLUSIONSelenium at 5-20 micromol/kg can induce DNA damage, apoptosis, and overexpression of c-myc, c-fos, and c-jun in rat hepatocytes.

Animals ; Apoptosis ; drug effects ; Blotting, Northern ; Comet Assay ; DNA Damage ; Dose-Response Relationship, Drug ; Genes, fos ; drug effects ; genetics ; Genes, jun ; drug effects ; genetics ; Genes, myc ; drug effects ; genetics ; Hepatocytes ; drug effects ; pathology ; Male ; Nucleic Acid Hybridization ; Rats ; Rats, Sprague-Dawley ; Selenium ; pharmacology ; Sodium Selenite ; pharmacology

OBJECTIVETo explore the mechanism of electroacupuncture at "Neiguan" (PC 6) improving acute myocardial ischemia.

METHODSThe rats were randomly divided into a sham operation group, a myocardial ischemia model group and a myocardial ischemia model plus electroacupuncture group. The acute myocardial ischemia model was developed byligation of the descending anterior branch of the coronary artery, and electroacupuncture was given at bilateral "Neiguan" (PC 6). Serum myocardial enzymes was determined by biochemical method and the expression of c-fos mRNA in myocardium was detected by using reverse transcription polymerase chain reaction (RT-PCR).

RESULTSThe activities of serum myocardial enzymes and the expression of c-fos mRNA in ischemic myocardium were significantly increased as compared with those in the sham operation group (P < 0.05), and after electroacupuncture they were significantly decreased (P < 0.05).

CONCLUSIONThe mechanism of electroacupuncture at "Neiguan" (PC 6) improving acute myochadial ischemia is possibly related with down-regulation of expression of c-fos mRNA in myocardium.

Acupuncture Points ; Animals ; Electroacupuncture ; Genes, fos ; Humans ; Myocardial Ischemia ; genetics ; Myocardium ; metabolism ; Rats

OBJECTIVEIt was reported previously that genistein could inhibit proliferation of human ovarian carcinoma cell line SKOV(3), but mechanism was not clear. There is a close relationship between EGFR mediated signal transduction pathway and the development of ovarian tumor. This study aimed to investigate the effects of genistein on the EGFR mediated signal transduction pathway and to clarify its mechanisms of proliferation inhibition on human ovarian carcinoma cell line SKOV(3) and its xenograft in nude mice.

METHODSThe expression of c-erbB-2 protein was determined using immunocytochemistry. The expressions of c-jun and c-fos protein were determined using Western blotting. The expression of c-erbB-2, c-raf-1, c-jun and c-fos mRNA were tested by reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSThe expression of c-erbB-2, c-raf-1 and its downstream gene c-jun and c-fos were decreased at mRNA level in the 20 micromol/L genistein group. The expression of c-erbB-2 protein were decreased, its average absorbency (A) were decreased after treatment of SKOV(3) with 20 micromol/L genistein for 48 h, reached at 0.42 +/- 0.02 (P < 0.05). Western blotting demonstrated that the expressions of c-jun and c-fos protein were decreased gradually after being treated with 20 micromol/L genistein for 12 - 72 h.

CONCLUSIONSGenistein could down-regulate the expression of two key genes, c-erbB-2 and c-raf-1 at mRNA and protein level in the EGFR mediated signal transduction pathway, and down-regulate the expression of its downstream nuclear transcription factors c-jun and c-fos at mRNA and protein level. It is suggested that interfering the expressions of some key signal molecules in EGFR mediated signal transduction system by genistein may account for its molecular foundation of proliferation inhibition in ovarian carcinoma.

Animals ; Antineoplastic Agents ; pharmacology ; Cell Line, Tumor ; Female ; Gene Expression Regulation, Neoplastic ; Genes, fos ; Genes, jun ; Genistein ; pharmacology ; Humans ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; Ovarian Neoplasms ; metabolism ; pathology ; Proto-Oncogene Proteins c-fos ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-jun ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-raf ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Receptor, Epidermal Growth Factor ; metabolism ; Receptor, ErbB-2 ; biosynthesis ; genetics ; Signal Transduction ; drug effects

To observe the effects of Panax Notoginosides (PNS) on up-regulation of AP-1 family transcription factors NF-E2, c-jun and c-fos for exploring intracellular signal pathway of PNS in hematopoietic cells, four human hematopoietic cells lines including myeloid HL-60, erythroid K562, megakaryoid CHRF-288 and Meg-01 were incubated in the presence of PNS for 14 days. The nuclear protein of cells were extracted and analyzed by Western blot with antibodies against NF-E2, c-fos and c-jun. Electrophoretic mobility shift assay (EMSA) was performed by using (32)P labeled AP-1 consensus oligonucleotide which contains binding site for NF-E2, c-jun and c-fos. The results showed that the transcription factors NF-E2, c-jun and c-fos of AP-1 family could be induced by PNS. Western blot demonstrated that the nuclear protein of both NF-E2 and c-jun in four cell lines treated by PNS were increased by 1.5-2.5- and 2.0-3.0-fold over untreated cells respectively. The c-fos protein in three cell lines of K562, CHRF-288 and Meg-01 was also elevated by 2.0-3.0-fold respectively, while c-fos protein in HL-60 cells was no detectable difference after PNS treatment. EMSA results in four cell lines indicated that AP-1 binding activity initiated by PNS was apparently elevated to form higher density band of AP-1-DNA complex. In conclusion, the intracellular transcription regulation initiated by PNS was involved in transcription factors NF-E2, c-jun and c-fos of AP-1 family members, which could play an important role in the up-regulation of genes expression related to proliferation and differentiation of hematopoietic cells.
DNA ; metabolism ; DNA-Binding Proteins ; genetics ; Erythroid-Specific DNA-Binding Factors ; Gene Expression Regulation ; drug effects ; Genes, fos ; Genes, jun ; Ginsenosides ; pharmacology ; HL-60 Cells ; Humans ; K562 Cells ; NF-E2 Transcription Factor ; NF-E2 Transcription Factor, p45 Subunit ; Panax ; Transcription Factor AP-1 ; metabolism ; Transcription Factors ; genetics ; Up-Regulation

OBJECTIVETo study the mechanism of Shu Di-huang in improving the function of learning and memory.

METHODOn the rats model with thalamic arcuate nucleus dameged by MSG, the improving function of Shu Di-huang on learning and memory was observed by step down task and Morris water maze task, and the expression of c-fos and NGF in hippocampi was observed by immunohistochemical means.

RESULTShu Di-huang could decrease the times of mistakes and prolong the incubation period in step down task, and shorten the incubation period of seeking the platform, and improve the percentage rate through the platform position in Morris water maze task. Shu Di-huang also increase the expression of hippocampal NGF, c-fos.

CONCLUSIONShu Di-huang can improve the function of learning and memory of MSG rats, and its mechanism may be related with the increase of the expression of hippocampal c-fos and NGF.

Animals ; Arcuate Nucleus of Hypothalamus ; drug effects ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Genes, fos ; Hippocampus ; metabolism ; Male ; Maze Learning ; drug effects ; Memory ; drug effects ; Nerve Growth Factor ; biosynthesis ; genetics ; Plants, Medicinal ; chemistry ; Proto-Oncogene Proteins c-fos ; biosynthesis ; genetics ; Rats ; Rats, Sprague-Dawley ; Rehmannia ; chemistry ; Sodium Glutamate