Anti-p70 S6K (R365) Antibody (A25593)

Hepatic stellate cells (HSCs) are liver-specific pericytes that are recruited to vessels and secret pro-angiogenic cytokines, and thus actively involved in pathological vascularization during liver fibrosis. Peroxisome proliferator-activated receptor-γ (PPARγ) is a switch molecule controlling HSC activation. We investigated PPARγ regulation of angiogenic signal transduction and the molecular mechanisms involved in HSCs. Primary rat HSCs and liver sinusoidal endothelial cells (LSECs) were isolated and used in this study. Boyden chamber and tubulogenesis assays, identified that focal adhesion kinase (FAK)-RhoA signaling activated by platelet-derived growth factor (PDGF) was required for HSC motility and the associated vascularization. PDGF also stimulated vascular endothelial growth factor (VEGF) expression and HSC-driven vascularization through signals mediated by extracellular signal-regulated kinase (ERK) and mammalian target of rapamycin (mTOR). Gain- and loss-of-function analyses demonstrated that activation of PPARγ interrupted FAK-RhoA, ERK and mTOR cascades and inhibited HSC-based vascularization. Molecular evidence further revealed that PPARγ attenuation of HSC angiogenic properties was dependent on inhibition of PDGF-β receptor expression. We concluded that PPARγ inhibited angiogenic signal transduction through transrepression of PDGF-β receptor leading to reduced HSC motility, reduced VEGF expression, and thereby attenuated HSC-driven angiogenesis. PPARγ could be a molecular target for preventing vascular remolding in hepatic fibrosis.

Excessive βAR stimulation is an independent factor in inducing pathological cardiac hypertrophy. Here, we report miR-145 regulates both expression and localization of GATA6, thereby protecting the heart against cardiomyocyte hypertrophy induced by isoproterenol (ISO). The protective activity of miR-145 was associated with down-regulation of ANF, BNP and β-MHC expression, a decreased rate of protein synthesis, inhibited cardiomyocyte growth and the modulation of several signaling pathways including ERK1/2, JNK and Akt-GSK3β. The anti-hypertrophic effect was abrogated by exogenous over-expression of transcription factor GATA6 which was further identified as a direct target of miR-145. In addition, GSK3β antagonists, LiCl and TDZD8, restored the nuclear accumulation of GATA6, which was attenuated by miR-145 Finally, we observed a dynamic pattern of miR-145 expression in ISO-treated NRCMs and in the hearts of TAC mice. Together, our results identify miR-145 as an important regulator in cardiac hypertrophy.

Pancreatic cancer remains the fourth most common cause of cancer-related death in the United States. Potent therapeutic strategies are urgently needed for pancreatic cancer. Cucurmosin is a novel type 1 ribosome-inactivating protein (RIP) isolated from the sarcocarp of Cucurbita moschata (pumpkin). Due to its cytotoxicity, cucurmosin can inhibit tumor cell proliferation through induction of apoptosis on tumor cells, but the specific mechanism is still unclear. We explored the function of cucurmosin in BxPC-3 pancreatic cancer cells using multiple cellular and molecular approaches such as 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry, reverse transcription polymerase chain reaction (RT-PCR), Western blotting and transmission electron microscopy for observing typical changes and formation of apoptotic bodies. We found that cucurmosin inhibited the proliferation of BxPC-3 cells in a time- and dose-dependent manner, and increased the cell population in the G0-G1 phase. With increasing concentration of cucurmosin, the expression of EGFR, p-PI3K, Akt, p-Akt, mTOR, p-mTOR, P70S6K-α, p-P70S6K-α, 4E-BP1 and p-4E-BP1 at the protein level was decreased, whereas the expression of p-Bad and caspase-9 was elevated. However, the mRNA expression of EGFR did not change. These findings suggest that cucurmosin can down-regulate the expression of EGFR by targeting. Cucurmosin induces the apoptosis of BxPC-3 pancreatic cancer cells via the PI3K/Akt/mTOR signaling pathway.

To study the roles of microRNA-223 (miR-223) in regulation of cell growth, we established a miR-223 over-expression model in HeLa cells infected with miR-223 by Lentivirus pLL3.7 system. We observed in this model that miR-223 significantly suppressed the proliferation, growth rate, colony formation of HeLa cells in vitro, and in vivo tumorigenicity or tumor formation in nude mice. To investigate the mechanisms involved, we scanned and examined the potential and putative target molecules of miR-223 by informatics, quantitative PCR and Western blot, and found that insulin-like growth factor-1 receptor (IGF-1R) was the functional target of miR-223 inhibition of cell proliferation. Targeting IGF-1R by miR-223 was not only seen in HeLa cells, but also in leukemia and hepatoma cells. The downstream pathway, Akt/mTOR/p70S6K, to which the signal was mediated by IGF-1R, was inhibited as well. The relative luciferase activity of the reporter containing wild-type 3'UTR(3'untranslated region) of IGF-1R was significantly suppressed, but the mutant not. Silence of IGF-1R expression by vector-based short hairpin RNA resulted in the similar inhibition with miR-223. Contrarily, rescued IGF-1R expression in the cells that over-expressed miR-223, reversed the inhibition caused by miR-223 via introducing IGF-1R cDNA that didn't contain the 3'UTR. Meanwhile, we also noted that miR-223 targeted Rasa1, but the downstream molecules mediated by Rasa1 was neither targeted nor regulated. Therefore we believed that IGF-1R was the functional target for miR-223 suppression of cell proliferation and its downstream PI3K/Akt/mTOR/p70S6K pathway suppressed by miR-223 was by targeting IGF-1R.

Infusion (i.c.v.) of beta-amyloid 25-35 (Abeta(25-35)) stimulates proliferation of progenitor cells in the hippocampal dentate gyrus (DG) of adult male mice, but a large population of the newborn cells will die in the 2nd week after birth, a critical period for neurite growth. Neurosteroid dehydroepiandrosterone (DHEA) has been demonstrated to promote neurite growth. Herein, we report that the DHEA-treatment on 6-12 days after BrdU-injection (BrdU-D(6-12)) dose-dependently attenuates the loss of newborn neurons induced by Abeta(25-35)-infusion. The DHEA-neuroprotection was blocked by the sigma(1) receptor antagonist NE100 and mimicked by the sigma(1) receptor agonist PRE084 when administered on BrdU-D(6-12). The DHEA-action was sensitive to the PI3K inhibitor LY294002 and the mammalian target of rapamycin (mTOR) inhibitor rapamycin. The Abeta(25-35)-infusion decreased the levels of Akt, mTOR and p70S6k phosphorylation, which could be rescued by DHEA-treatment in a sigma(1) receptor-dependent manner. Furthermore, the Abeta(25-35)-infusion led to a decrease in the dendritic density and length of doublecortin positive cells in the DG, which also was improved by the DHEA-treatment on BrdU-D(6-12). These findings suggest that DHEA prevents the Abeta(25-35)-impaired survival and dendritic growth of newborn neurons through a sigma(1) receptor-mediated modulation of PI3K-Akt-mTOR-p70S6k signaling.