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 ligand-induced 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.

Cell-therapy modalities using mesenchymal stem (MSCs) in experimental strokes are being investigated due to the role of MSCs in neuroprotection and regeneration. It is necessary to know the sequence of events that occur during stress and how MSCs complement the rescue of neuronal cell death mediated by [Ca²⁺]_i and reactive oxygen species (ROS). In the current study, SH-SY5Y-differentiated neuronal cells were subjected to in vitro cerebral ischemia-like stress and were experimentally rescued from cell death using an MSCs/neuronal cell coculture model. Neuronal cell death was characterized by the induction of proinflammatory tumor necrosis factor (TNF)-α, interleukin (IL)-1β and -12, up to 35-fold with corresponding downregulation of anti-inflammatory cytokine transforming growth factor (TGF)-β, IL-6 and -10 by approximately 1 to 7 fold. Increased intracellular calcium [Ca²⁺]_i and ROS clearly reaffirmed oxidative stress-mediated apoptosis, while upregulation of nuclear factor NF-κB and cyclo-oxygenase (COX)-2 expressions, along with ~41% accumulation of early and late phase apoptotic cells, confirmed ischemic stress-mediated cell death. Stressed neuronal cells were rescued from death when cocultured with MSCs via increased expression of anti-inflammatory cytokines (TGF-β, 17%; IL-6, 4%; and IL-10, 13%), significantly downregulated NF-κB and proinflammatory COX-2 expression. Further accumulation of early and late apoptotic cells was diminished to 23%, while corresponding cell death decreased from 40% to 17%. Low superoxide dismutase 1 (SOD1) expression at the mRNA level was rescued by MSCs coculture, while no significant changes were observed with catalase (CAT) and glutathione peroxidase (GPx). Interestingly, increased serotonin release into the culture supernatant was proportionate to the elevated [Ca²⁺]_i and corresponding ROS, which were later rescued by the MSCs coculture to near normalcy. Taken together, all of these results primarily support MSCs-mediated modulation of stressed neuronal cell survival in vitro.

Background and Objective: Although the anticancer potentials of water-insoluble drugs are improved by nanoformulation, other intervening factors may contribute in the drug efficacy. This work was designated to explore the effect of paclitaxel-loaded Poly(Lactic-co-Glycolic Acid) (PLGA) nanoparticles on the viability of cancer cells, the expression of Taxol Resistance gene I (TXR1) and paclitaxel metabolizing genes. Methods: Paclitaxel loaded PLGA Nanoparticles (PTX-NPs) were prepared, physically characterized and used in the treatment of breast adenocarcinoma cells (MCF-7) and hepatoma cells (HepG2). Cells viability and apoptosis were investigated. In parallel, RNA was isolated, reverse transcribed and used to monitor the expression levels of TXR1, CYP 3A4 and CYP2C8 genes. Results: PTX-NPs were characterized by transmission electron microscopy to be of a nano-size sphere-like shape. FTIR analysis revealed good coupling between PTX and PLGA. The encapsulation efficiency was 99% and the drug release demonstrated a progressive releasing phase followed by slower and sustained releasing phases. Although HepG2 cells demonstrated more resistance to PTX than MCF-7 cells, both cell types were more responsive to PTX-NPS compared to PTX. The IC50 values decreased from 19.3 to 6.7 in breast cancer cells and from 42.5 to 13.1μg/ml in hepatoma cells. The apoptosis was the key mechanism in both cells, where at least 44% of cells underwent apoptosis. The expression of TXR1 decreased when either cells were treated with PTX-NPs, respectively, meanwhile the expressions of CYP3A4 and CYP2C8 were increased. Conclusion: Taken together, this in vitro study reports the associations between the enhanced responsiveness of MCF-7 and HepG2 cells to PLGA-loaded paclitaxel nanoparticles and the accompanying decrease in the cells resistance to the PTX and its enhanced metabolism.

Breast cancer cells abnormally express vasopressin (AVP) and its receptors. The effect of AVP is largely orchestrated through its downstream signaling and by receptor-mediated endocytosis (RME), in which Dynamin 2 (Dyn2) plays an integral role in vesicle closure. In this work, luminal A breast cancer cells were treated with AVP, and then Dynasore (DYN) was employed to inhibit Dyn2 to explore the combined effect of AVP and Dyn2 inhibition on the survival of breast cancer cells. The results revealed that DYN alone demonstrated a concentration-dependent cytotoxic effect in AVP untreated cells. Apoptosis developed in 29.7 and 30.3% of cells treated with AVP or AVP+DYN, respectively, compared to 32.5% in cells treated with Wortmannin (Wort, a selective PI3K pathway inhibitor). More apoptosis was observed when cells were treated with DYN+Wort in presence or absence of exogenous AVP. Besides, 2 or 4- fold increases in the expression of Bax and Caspase-3, were observed in cells exposed to AVP in absence or presence of DYN, respectively. This was associated with higher levels of the autophagy marker (LC3II protein). Meanwhile, the activation of Akt protein, sequentially decreased in the same pattern. Cell’s invasion decreased when they were exposed to AVP alone or combined with DYN or/and Wort. Conclusively, although many reports suggested the proliferative effect of AVP, the results predict the antiproliferative and antimetastatic effects of 100 nM AVP in luminal A breast cancer cells. However, the hormone did not enhance the cytotoxic effect of Dyn 2 or PI3K pathway inhibition. Graphical Abstract Summary of the Dynamin 2 independent AVP antiproliferative effects. Breast cancer cells expresses AVP as a Prohormone (A). At high dose of AVP, the hormone is liganded with AVP receptor (B) to initiate RME, where the endosomed complex (C) is degraded through the endosome-lysosome system, as a part of signal management. These events consume soluble Dyn2 in neck closure and vesicle fission (D). This makes the cells more substitutable to the direct apoptotic effect of DYN (E). Alternatively, at lower AVP doses the liganded AVP may initiate cAMP-mediated downstream signaling (F) and cellular proliferation. In parallel, Wort inhibits PIP2-PIP3 conversion (G) and the subsequent inhibition of PI3K/Akt/mTOR pathway leading to cell death.