The molecular mechanism of the cell-cycle machinery in uterine leiomyoma has not yet been fully elucidated. Among the various types of cell-cycle regulators, p27(Kip1)(p27) is considered to be a potent tumor suppressor. To provide further molecular basis for understanding the progression of uterine leiomyoma, our objective was to evaluate the expression level of p27 in normal myometrium and uterine leiomyoma tissue and its effect on cytogenic growth. Western blot analysis, real-time polymerase chain reaction (PCR) and immunohistochemical staining revealed that p27 protein and messenger RNA were down-regulated in uterine leiomyoma tissue and cultured cells compared to normal myometerium. Full-length human p27 cDNA was transferred using a replication-deficient recombinant adenoviral vector (Ad.p27) into uterine leiomyoma cells and evaluated the effect on cell proliferation. Transfection of Ad.p27 into uterine leiomyoma cells resulted in the induction of apoptosis, reduction in viability and proliferation of uterine leiomyoma cells. Our results suggest a new paradigm that down-regulated p27 protein expression is the possible underlying mechanism for the growth of uterine leiomyoma and over-expression of p27 induces cell death. This study provides better understanding of the control exerted by p27 in regulating growth and disease progression of uterine leiomyoma.
Adult ; Cell Cycle ; Cell Proliferation ; Female ; Humans ; Intracellular Signaling Peptides and Proteins/antagonists & inhibitors/genetics/*physiology ; Leiomyoma/*pathology ; Middle Aged ; RNA, Messenger/analysis ; Uterine Neoplasms/*pathology

OBJECTIVE: To observe whether HSP70 could protect against H2O2-induced apoptosis in C2C12 myogenic cells by inhibiting Smac release from the mitochondria. METHODS: HSP70 gene and full length Smac gene was transiently transfected in C2C12 myogenic cells by lipofectamine and the protein levels of HSP70 and Smac were analysed by Western blotting. Hoechst 33 258 staining was used to examine cell morphological changes and to calculate percentage of apoptotic nuclei. DNA ladder pattern on agarose gel electrophoresis was used to observe the DNA fragmentation. Activities of caspase-3 and caspase-9 were assayed with Western blotting. The release of Smac from the mitochondria to the cytoplasm was observed by immunofluorescence. RESULTS: H2O2 ( 0.5 mmol/L ) activated caspase-3, caspase-9 8 h after the treatment and specific morphological changes of apoptosis 12 h after the treatment, and overexpression of Smac significantly promoted H2O2-induced activation of caspase-3, caspase-9 and apoptosis in C2C12 myogenic cells. HSP70 overexpression significantly inhibited H2O2-induced and Smac-promoted apoptosis, as shown by no specific DNA ladder pattern in agarose gel electrophoresis, decrease of percentage of apoptotic nuclei, and marked inactivation of caspase-3 and caspase-9. HSP70 could inhibit the release of Smac from the mitochondria to the cytoplasm 2 h after the treatment by H2O2. CONCLUSION HSP70 inhibits Smac release from the mitochondria and protects against H2O2-induced apoptosis in C2C12 myogenic cells.
Apoptosis ; drug effects ; Apoptosis Regulatory Proteins ; Cells, Cultured ; HSP70 Heat-Shock Proteins ; pharmacology ; Humans ; Hydrogen Peroxide ; Intracellular Signaling Peptides and Proteins ; metabolism ; Mitochondria, Heart ; drug effects ; metabolism ; Mitochondrial Proteins ; antagonists & inhibitors ; metabolism ; Myoblasts ; metabolism ; Myocytes, Cardiac ; drug effects ; metabolism

Animals ; Carcinoma, Non-Small-Cell Lung ; drug therapy ; metabolism ; Humans ; Intracellular Signaling Peptides and Proteins ; antagonists & inhibitors ; metabolism ; Lung Neoplasms ; drug therapy ; metabolism ; Protein-Serine-Threonine Kinases ; antagonists & inhibitors ; metabolism ; Signal Transduction ; drug effects ; Sirolimus ; therapeutic use ; TOR Serine-Threonine Kinases

CSN1 is a component of the COP9 signalosome (CSN), a conserved protein complex with pleiotropic functions in many organs and cell types. CSN regulates ubiquitinproteasome dependent protein degradation via the deneddylation and the associated deubiquitination activities. In addition, CSN associates with protein kinases and modulates cell signaling, particularly the activator protein 1 (AP-1) pathway. We have shown previously that CSN1 suppresses AP-1 transcription activity and inhibits ultraviolet (UV) and serum activation of c-fos expression. Here we show that CSN1 can inhibit phosphorylation of proto-oncogene c-Jun product and repress c-Jun dependent transcription. Further, CSN1 dramatically downregulates ectopic expression of c-Jun N-terminal kinase 1 (JNK1) in cultured cells. The decline in JNK1 is not caused by excessive proteolysis or by 3' UTR-dependent mRNA instability, but by CSN1-dependent repression of one or multiple steps in transcriptional and posttranscriptional mechanisms. Thus, in contrast to CSN5/Jab1, which promotes AP-1 activity, CSN1 displays a negative effect on the AP-1 pathway. Finally, we discuss about the dynamic equilibrium of the CSN complexes in regulation of the AP-1 pathway.
3' Untranslated Regions ; Animals ; COP9 Signalosome Complex ; Cell Line ; Down-Regulation ; Humans ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Mice ; Mitogen-Activated Protein Kinase 8 ; metabolism ; Phosphorylation ; Proto-Oncogene Proteins c-jun ; antagonists & inhibitors ; metabolism ; Transcription Factor AP-1 ; metabolism

Oxidative stress and apoptosis are the key factors that limit the hypothermic preservation time of donor hearts to within 4-6 h. The aim of this study was to investigate whether the histone deacetylase 3 (HDAC3) inhibitor RGFP966 could protect against cardiac injury induced by prolonged hypothermic preservation. Rat hearts were hypothermically preserved in Celsior solution with or without RGFP966 for 12 h followed by 60 min of reperfusion. Hemodynamic parameters during reperfusion were evaluated. The expression and phosphorylation levels of mammalian STE20-like kinase-1 (Mst1) and Yes-associated protein (YAP) were determined by western blotting. Cell apoptosis was measured by the terminal deoxynucleotidyl-transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) method. Addition of RGFP966 in Celsior solution significantly inhibited cardiac dysfunction induced by hypothermic preservation. RGFP966 inhibited the hypothermic preservation-induced increase of the phosphorylated (p)-Mst1/Mst1 and p-YAP/YAP ratios, prevented a reduction in total YAP protein expression, and increased the nuclear YAP protein level. Verteporfin (VP), a small molecular inhibitor of YAP-transcriptional enhanced associate domain (TEAD) interaction, partially abolished the protective effect of RGFP966 on cardiac function, and reduced lactate dehydrogenase activity and malondialdehyde content. RGFP966 increased superoxide dismutase, catalase, and glutathione peroxidase gene and protein expression, which was abolished by VP. RGFP966 inhibited hypothermic preservation-induced overexpression of B-cell lymphoma protein 2 (Bcl-2)-associated X (Bax) and cleaved caspase-3, increased Bcl-2 mRNA and protein expression, and reduced cardiomyocyte apoptosis. The antioxidant and anti-apoptotic effects of RGFP966 were cancelled by VP. The results suggest that supplementation of Celsior solution with RGFP966 attenuated prolonged hypothermic preservation-induced cardiac dysfunction. The mechanism may involve inhibition of oxidative stress and apoptosis via inactivation of the YAP pathway.
Acrylamides/pharmacology* ; Animals ; Apoptosis/drug effects* ; Cryopreservation ; Disaccharides/pharmacology* ; Electrolytes/pharmacology* ; Glutamates/pharmacology* ; Glutathione/pharmacology* ; Heart/physiology* ; Heart Transplantation/methods* ; Hepatocyte Growth Factor/antagonists & inhibitors* ; Histidine/pharmacology* ; Histone Deacetylase Inhibitors/pharmacology* ; Intracellular Signaling Peptides and Proteins/antagonists & inhibitors* ; Male ; Mannitol/pharmacology* ; Oxidative Stress/drug effects* ; Phenylenediamines/pharmacology* ; Proto-Oncogene Proteins/antagonists & inhibitors* ; Rats ; Rats, Sprague-Dawley ; Signal Transduction/drug effects* ; YAP-Signaling Proteins

Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Chromones ; pharmacology ; Cyclin-Dependent Kinase Inhibitor p27 ; antagonists & inhibitors ; physiology ; Endometrial Neoplasms ; drug therapy ; pathology ; Estradiol ; pharmacology ; Female ; G1 Phase ; drug effects ; Humans ; Intracellular Signaling Peptides and Proteins ; antagonists & inhibitors ; physiology ; Morpholines ; pharmacology ; Phosphatidylinositol 3-Kinases ; antagonists & inhibitors ; physiology ; Proto-Oncogene Proteins c-akt ; antagonists & inhibitors ; physiology ; Signal Transduction ; drug effects ; physiology

The present study aimed to investigate the effects of humanin (HN) on primary cortical neuronal apoptosis induced by Abeta31-35, and explore the potential mechanisms. Cultured cortical neurons were pretreated with different concentrations of HN (5, 10, 20 micromol/L) for different time period (0, 8 and 16 h) respectively, and then exposed to Abeta31-35 (25 micromol/L) for additional 24 h and the neuronal apoptosis was examined by morphological analysis, flow cytometric assays and TUNEL staining. Caspase activities were measured using a spectrophotometer. Bax expression was measured by Western blot. The results were as follows. (1) Pretreatment with HN (20 micromol/L) for 16 h significantly prevented Abeta31-35-induced apoptosis in cortical neurons; (2) HN significantly decreased Abeta31-35-induced elevation of caspase-3 and -9 activities; (3) HN suppressed Abeta31-35-induced translocation of Bax from the cytosol to mitochondria, but had no effect on overall Bax expression. In conclusions, HN attenuated Abeta31-35-induced cortical neuronal apoptosis by blocking intrinsic caspase-dependent apoptotic pathways.
Amyloid beta-Peptides ; antagonists & inhibitors ; toxicity ; Animals ; Animals, Newborn ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Caspase 9 ; metabolism ; Cells, Cultured ; Cerebral Cortex ; cytology ; pathology ; Intracellular Signaling Peptides and Proteins ; pharmacology ; Neurons ; cytology ; pathology ; Neuroprotective Agents ; pharmacology ; Peptide Fragments ; antagonists & inhibitors ; toxicity ; Rats ; Rats, Sprague-Dawley

BACKGROUNDIt has been reported that there is a significant difference in the local tissue concentration of transforming growth factor (TGF)-β1 between chronic rhinosinusitis without nasal polyps (CRSsNP) and chronic rhinosinusitis without nasal polyps (CRSwNP) patients. TGF-β has been reported to play an important role in regulating epithelial cell repair in lower airway remodeling and may be a critical factor involved in the remodeling process of chronic rhinosinusitis (CRS).

METHODSEthmoidal mucosal samples collected from CRS and healthy control patients were analyzed for TGF-β1, TGF-β receptor I, TGF-β receptor II, Smad3, phospho-Smad3, Smad7, and Smad anchor for receptor activation by Western blotting analysis. The proliferation of sinonasal epithelial cells at baseline and after TGF-β1 and/or EGF stimulation was evaluated by the MTT assay.

RESULTSIn CRSsNP, TGF-β1, TGF-β receptor I, TGF-β receptor II, and Smad3 protein levels were significantly higher than controls. In CRSwNP, TGF-β1, Smad3, and pSmad3 protein levels were significantly lower than controls. Smad7 protein was significantly higher in CRS than controls. In vitro experiments demonstrated that the baseline proliferation levels of sinonasal epithelial cells were lower in CRS than controls.

CONCLUSIONSCRSwNP is characterized by a lower level of TGF-signaling compared with the control. In CRSsNP, although the upstream signaling of TGF-β was enhanced, the high Smad7 protein expression may restrain the downstream signaling components (e.g., pSmad3) and the TGF-β antiproliferative effect on sinonasal epithelium. The difference in the local tissue concentration of TGF-β1 between CRSsNP and CRSwNP patients did not result in significant differences in epithelial proliferation.

Adult ; Aged ; Benzamides ; pharmacology ; Cells, Cultured ; Dioxoles ; pharmacology ; Female ; Humans ; Intracellular Signaling Peptides and Proteins ; metabolism ; Male ; Middle Aged ; Protein-Serine-Threonine Kinases ; antagonists & inhibitors ; metabolism ; Receptors, Transforming Growth Factor beta ; antagonists & inhibitors ; metabolism ; Serine Endopeptidases ; metabolism ; Signal Transduction ; drug effects ; Sinusitis ; metabolism ; Smad3 Protein ; metabolism ; Smad7 Protein ; metabolism ; Transforming Growth Factor beta ; metabolism ; Transforming Growth Factor beta1 ; antagonists & inhibitors ; metabolism ; Young Adult

AIMTo identify flutamide regulated genes in the rat ventral prostate.

METHODSTotal RNA from ventral prostates of control and flutamide treated rats were isolated. Differentially expressed transcripts were identified using differential display reverse transcriptase polymerase chain reaction. The effect of castration on the expression of flutamide-regulated transcripts was studied.

RESULTSWe have identified beta2-microglobulin, cytoplasmic FMR1 interacting protein 2 and pumilio 1 as flutamide induced and spermine binding protein and ribophorin II as flutamide repressed targets in the rat ventral prostate. Although flutamide treatment caused an induction of pumilio 1 mRNA, castration had no effect.

CONCLUSIONCastration and flutamide treatments exert differential effects on gene expression. Flutamide might also have direct AR independent effects, which might have implications in the emergence of androgen independent prostate cancer and the failure of flutamide therapy.

Adaptor Proteins, Signal Transducing ; drug effects ; metabolism ; Androgen Antagonists ; pharmacology ; Animals ; Flutamide ; pharmacology ; Gene Expression Regulation ; drug effects ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Male ; Membrane Proteins ; genetics ; metabolism ; Orchiectomy ; Prostate ; drug effects ; metabolism ; RNA-Binding Proteins ; metabolism ; Rats ; Rats, Wistar ; beta 2-Microglobulin ; drug effects ; metabolism

Allergic diseases have become global social health problems. The binding of IgE with its high affinity receptor FcepsilonRI plays a key step in I-type allergy. Recently, more and more key molecules on the IgE/FcepsilonRI signaling transduction pathway were to be the drug candidates against allergic diseases, with in-depth study of FcepsilonRI signal pathway gradually. The main drugs include molecule antibodies, peptides, vaccines, fusion proteins, small molecules, and other drugs related to IgE/FcepsilonRI. The recent progress in the study of mechanisms of representative drugs targeting on IgE/FcepsilonRI signaling pathway was reviewed in this article.
Aminophenols ; pharmacology ; therapeutic use ; Animals ; Anti-Allergic Agents ; pharmacology ; therapeutic use ; Antibodies, Anti-Idiotypic ; pharmacology ; therapeutic use ; Antibodies, Monoclonal, Humanized ; pharmacology ; therapeutic use ; Humans ; Hypersensitivity ; drug therapy ; immunology ; Immunoglobulin E ; metabolism ; Intracellular Signaling Peptides and Proteins ; antagonists & inhibitors ; Molecular Targeted Therapy ; Omalizumab ; Protein-Tyrosine Kinases ; antagonists & inhibitors ; Pyrimidines ; pharmacology ; therapeutic use ; Receptors, IgE ; metabolism ; Signal Transduction ; Syk Kinase