The androgen receptor (AR) regulates ligand-dependent gene transcription upon binding specific DNA sequences. The AR conveys both trans-activation and trans-repression functions, which together contribute to prostate cellular growth, differentiation, and apoptosis. Like histone H3, the AR is post-translationally modified by both acetylation and phosphorylation. The histone acetyltransferase p300 transactivates the AR and directly acetylates the AR in vitro at a conserved motif. Point mutations of the AR acetylation motif that abrogate acetylation reduce trans-activation by p300 without affecting the trans-repression function of the AR. The current studies assessed the functional relationship between acetylation and phosphorylation of the AR. Herein trans-activation of the AR acetylation site mutants were enhanced by the p42/ p44 MAPK pathway but were defective in regulation by protein kinase A (PKA) signaling. PKA inhibition augmented ARwt activity but not AR acetylation mutant gene reporter activity and association at an androgen response element in chromatin immunoprecipitation assays. Mutations of the lysine residues at the AR acetylation site reduced trichostatin A (TSA) responsiveness and ligand-induced phosphorylation of the AR. The AR acetylation site mutant formed ligandinduced phosphorylation-dependent isoforms with distinguishable characteristics from wild type AR as determined with two-dimensional electrophoresis. Conversely, point mutation of a subset of AR phosphorylation sites reduced trichostatin A responsiveness and trans-activation by histone acetyltransferases. Together these studies suggest that acetylation and phosphorylation of the AR are linked events and that the conserved AR lysine motif contributes to a select subset of pathways governing AR activity.Steroid receptors, including the androgen receptor (AR), 1 are members of the nuclear receptor superfamily that generally function as ligand-dependent transcriptional regulators (1). In the absence of ligand, interactions with corepressors maintain the receptor in an inactive state. The AR is expressed in a variety of cell types and plays an important role in development, male sexual differentiation, and prostate cellular proliferation. The functional domains of the AR (termed A-F) are conserved with other members of the "classic" receptor subclass. The C-terminal region of the AR, including the hinge region and ligand-binding domain (LBD) is responsible for ligand binding and dimerization. The well conserved DNA binding domain consists of 68 amino acids with two zinc finger structures. The N-terminal region contributes to transcriptional activation through its activation function 1 (2). In contrast to several other hormone-regulated nuclear receptors, the AR lacks an intrinsic activation function 2 function in the LBD. The LBD, which consists of twelve ␣-helices, projects away from the hormone-binding pocket in the absence of ligand and undergoes substantial conformational changes in the presence of ligand (3). The folding of the most C-ter...
Lead (Pb) toxicity is one of the commonest environmental problems in our life; it causes many reversible and irreversible changes in our tissues. This study was carried out to investigate the effect of meso-2,3-dimercaptosuccinic acid (DMSA) on treatment of oxidative stress caused by lead poisoning in rabbits. Lead acetate (Pb(Ac)(2)) was orally administrated to rabbits for 21 days and then treated by DMSA for another 21 days. The effect of this treatment was investigated by measuring 2 of the apoptosis proteins p53 and Bcl2. Also, the auto-oxidation rate and their histopathological changes in brain, bone and liver were investigated. Hemoglobin auto-oxidation rate is measured as well as histopathological study of liver. Our data indicate that exposure to rabbits to Pb(Ac)(2) caused a significant increase of apoptosis protein p53 and decrease in the antiapoptotic BCl2 proteins.
BackgroundNon invasive approaches will likely be increasing utilized to assess liver fibrosis. This work provides a new non invasive index to predict liver fibrosis induced in mice.MethodsFibrosis was generated by thioacetamide (TAA), chronic intake of ethanol, or infection with S. mansoni in 240 mice. Both progression and regression of fibrosis (after treatment with silymarin and/or praziquantel) were monitored. The following methods were employed: (i) The METAVIR system was utilized to grade and stage liver inflammation and fibosis; (ii) Determination of hepatic hydroxyproline and collagen; and (iii) Derivation of a new hepatic fibrosis index from the induced changes, and its prospective validation in a group of 70 mice.ResultsThe index is composed of 4 serum variable including total proteins, γ-GT, bilirubin and reduced glutathione (GSH), measured in diseased, treated and normal mice. These parameters were highly correlated with both the histological stage and the grade. They were combined in a logarithmic formula, which non-invasively scores the severity of liver fibrosis through a range (0 to 2), starting with healthy liver (corresponding to stage 0) to advanced fibrosis (corresponding stage 3).Receiver operating characteristic curves (ROC) for the accuracy of the index to predict the histological stages demonstrated that the areas under the curve (AUC) were 0.954, 0.979 and 0.99 for index values corresponding to histological stages 1, 2 and 3, respectively. Also, the index was correlated with stage and grade, (0.947 and 0.859, respectively). The cut off values that cover the range between stages 0-1, 1-2 and 2-3 are 0.4, 1.12 and 1.79, respectively. The results in the validation group confirmed the accuracy of the test. The AUROC was 0.869 and there was good correlation with the stage of fibrosis and grade of inflammation.ConclusionThe index fulfils the basic criteria of non-invasive marker of liver fibrosis since it is liver-specific, easy to implement, reliable, and inexpensive. It proved to be accurate in discriminating precirrhotic stages.
Non-visual arrestins (β-arrestins) are endocytic proteins that mediate agonist activated GPCRs internalization and signaling pathways in an independent manner. The involvement of β-arrestins in cancer invasion and metastasis is increasingly reported. So, it is hypothesized that inhibition of β-arrstins may diminish the survival chances of cancer cells. This study aimed to evaluate the in vitro impact of inhibiting β-arrestins on the autophagic and/or apoptotic responsiveness of breast cancer cells.We used Barbadin to selectively inhibit β-Arr/AP2 interaction in AVP stimulated V2R receptor of triple negative breast cancer cells (MDA MB-231). Autophagy was assessed by the microtubule-associated protein 1 light chain 3-II (LC3II), apoptosis was measured by Annexin-V/PI staining and cell cycle distribution was investigated based upon the DNA content using ow cytometry. Barbadin reduced cell viability to 69.1% and increased the autophagy marker LC3 II and its autophagic effect disappeared in cells transiently starved in Earle's balanced salt solution (EBSS). Also, Barbadin mildly enhanced the expression of P62 mRNA and arrested 63.7% of cells in G0/G1 phase. In parallel, the drug induced apoptosis in 29.9% of cells (by AV/PI) and 27.8% of cells were trapped in sub-G1 phase. The apoptotic effect of Barbadin was enhanced when autophagy was inhibited by the PI3K inhibitor (Wortmannin).Conclusively, the data demonstrate the dual autophagic and apoptotic effects of β-βArr/AP2 inhibition in triple negative breast cancer cells. These observations nominate β-Arrs as selective targets in breast cancer treatment.Thus, this work was designed to explore the autophagic and apoptosis effects of the inhibition of β-Arr/AP2 interaction in hormonally stimulated breast cancer cells. Materials And MethodsKey Reagents: Barbadin (3-amino-5-(4-benzylphenyl)-3H,4H-thieno[2,3-d]pyrimidin-4-one) (Cat No. B118250), Arginine Vasopressin acetic acid salt (AVP) (Cat No. V991535) and Wortmannin (Cat No. W499400), were purchased from Toronto Research Chemicals, Toronto, Ontario, Canada). Trichostatin A (TSA) and dimethyl sulfoxide (DMSO) were from Sigma Chemicals, USA. Earle's balanced salt solution (EBSS) and other cell culture reagents (DMEM 4.5 g/L glucose with L glutamine, penicillin/streptomycin, fetal bovine serum (FBS) and Trypsin/EDTA) were from Lonza Pharma & Biotech. Cell culture and treatment MDA MB-231 cells [16] was generously provided by Department of Cancer Biology, NCI, Cairo University.Cells were cultured in Dulbecco's modi ed Eagle's Minimal Essential Medium (DMEM) supplemented with 10% heat inactivated FBS, 1% Penicillin/Streptomycin in a humidi ed atmosphere of 95% air and 5% CO 2 at 37 °C. Initially, cells were seeded with a low cell density then subcultured with particular densities in 100 mm, 6 wells, or 96 wells plates according to the experimental settings. Cell treatmentsAutophagy was induced by glucose oxygen deprivation, where cells were starved for 4 hours in EBSS, which does not contain nutrients nor growth factors, at...
The study reports the antiproliferative and apoptosis-mediated cytotoxic effects of green tea and ginger polyphenolic extracts on human H460 cell line, indicating their promising chemopreventive effect against lung cancer.
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