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Sample records for akt signaling activates

  1. Akt1 signaling coordinates BMP signaling and β-catenin activity to regulate second heart field progenitor development.

    PubMed

    Luo, Wen; Zhao, Xia; Jin, Hengwei; Tao, Lichan; Zhu, Jingai; Wang, Huijuan; Hemmings, Brian A; Yang, Zhongzhou

    2015-02-15

    Second heart field (SHF) progenitors exhibit continued proliferation and delayed differentiation, which are modulated by FGF4/8/10, BMP and canonical Wnt/β-catenin signaling. PTEN-Akt signaling regulates the stem cell/progenitor cell homeostasis in several systems, such as hematopoietic stem cells, intestinal stem cells and neural progenitor cells. To address whether PTEN-Akt signaling is involved in regulating cardiac progenitors, we deleted Pten in SHF progenitors. Deletion of Pten caused SHF expansion and increased the size of the SHF derivatives, the right ventricle and the outflow tract. Cell proliferation of cardiac progenitors was enhanced, whereas cardiac differentiation was unaffected by Pten deletion. Removal of Akt1 rescued the phenotype and early lethality of Pten deletion mice, suggesting that Akt1 was the key downstream target that was negatively regulated by PTEN in cardiac progenitors. Furthermore, we found that inhibition of FOXO by Akt1 suppressed the expression of the gene encoding the BMP ligand (BMP7), leading to dampened BMP signaling in the hearts of Pten deletion mice. Cardiac activation of Akt also increased the Ser552 phosphorylation of β-catenin, thus enhancing its activity. Reducing β-catenin levels could partially rescue heart defects of Pten deletion mice. We conclude that Akt signaling regulates the cell proliferation of SHF progenitors through coordination of BMP signaling and β-catenin activity.

  2. Constitutively expressed COX-2 in osteoblasts positively regulates Akt signal transduction via suppression of PTEN activity.

    PubMed

    Li, Ching-Ju; Chang, Je-Ken; Wang, Gwo-Jaw; Ho, Mei-Ling

    2011-02-01

    Cyclooxygenase-2 (COX-2) is thought to be an inducible enzyme, but increasing reports indicate that COX-2 is constitutively expressed in several organs. The status of COX-2 expression in bone and its physiological role remains undefined. Non-selective non-steroidal anti-inflammatory drugs (NSAIDs) and selective COX-2 inhibitors, which commonly suppress COX-2 activity, were reported to suppress osteoblast proliferation via Akt/FOXO3a/p27(Kip1) signaling, suggesting that COX-2 may be the key factor of the suppressive effects of NSAIDs on proliferation. Although Akt activation correlates with PTEN deficiency and cell viability, the role of COX-2 on PTEN/Akt regulation remains unclear. In this study, we hypothesized that COX-2 may be constitutively expressed in osteoblasts and regulate PTEN/Akt-related proliferation. We examined the localization and co-expression of COX-2 and p-Akt in normal mouse femurs and in cultured mouse (mOBs) and human osteoblasts (hOBs). Our results showed that osteoblasts adjacent to the trabeculae, periosteum and endosteum in mouse femurs constitutively expressed COX-2, while COX-2 co-expressed with p-Akt in osteoblasts sitting adjacent to trabeculae in vivo, and in mOBs and hOBs in vitro. We further used COX-2 siRNA to test the role of COX-2 in Akt signaling in hOBs; COX-2 silencing significantly inhibited PTEN phosphorylation, enhanced PTEN activity, and suppressed p-Akt level and proliferation. However, replenishment of the COX-2 enzymatic product, PGE2, failed to reverse COX-2-dependent Akt phosphorylation. Furthermore, transfection with recombinant human COX-2 (rhCOX-2) significantly reversed COX-2 siRNA-suppressed PTEN phosphorylation, but this effect was reduced when the enzymatic activity of rhCOX-2 was blocked. This finding indicated that the effect of COX-2 on PTEN/Akt signaling is not related to PGE2 but still dependent on COX-2 enzymatic activity. Conversely, COX-1 silencing did not affect PTEN/Akt signaling. Our findings provide

  3. Antitumor Effects of Fucoidan on Human Colon Cancer Cells via Activation of Akt Signaling.

    PubMed

    Han, Yong-Seok; Lee, Jun Hee; Lee, Sang Hun

    2015-05-01

    We identified a novel Akt signaling mechanism that mediates fucoidan-induced suppression of human colon cancer cell (HT29) proliferation and anticancer effects. Fucoidan treatment significantly inhibited growth, induced G1-phase-associated upregulation of p21WAF1 expression, and suppressed cyclin and cyclin-dependent kinase expression in HT29 colon cancer cells. Additionally, fucoidan treatment activated the Akt signaling pathway, which was inhibited by treatment with an Akt inhibitor. The inhibition of Akt activation reversed the fucoidan-induced decrease in cell proliferation, the induction of G1-phase-associated p21WAF1 expression, and the reduction in cell cycle regulatory protein expression. Intraperitoneal injection of fucoidan reduced tumor volume; this enhanced antitumor efficacy was associated with induction of apoptosis and decreased angiogenesis. These data suggest that the activation of Akt signaling is involved in the growth inhibition of colon cancer cells treated with fucoidan. Thus, fucoidan may serve as a potential therapeutic agent for colon cancer.

  4. Phosphatidylinositol 3-kinase/Akt signaling enhances nuclear localization and transcriptional activity of BRCA1

    SciTech Connect

    Hinton, Cimona V.; Fitzgerald, Latricia D.; Thompson, Marilyn E. . E-mail: methompson@mmc.edu

    2007-05-15

    Signaling pathways involved in regulating nuclear-cytoplasmic distribution of BRCA1 have not been previously reported. Here, we provide evidence that heregulin {beta}1-induced activation of the Akt pathway increases the nuclear content of BRCA1. First, treatment of T47D breast cancer cells with heregulin {beta}1 results in a two-fold increase in nuclear BRCA1 as assessed by FACS analysis, immunoblotting and immunofluorescence. This heregulin-induced increase in nuclear BRCA1 is blocked by siRNA-mediated down-regulation of Akt. Second, mutation of threonine 509 in BRCA1, the site of Akt phosphorylation, to an alanine, attenuates the ability of heregulin to induce BRCA1 nuclear accumulation. These data suggest that Akt-catalyzed phosphorylation of BRCA1 is required for the heregulin-regulated nuclear concentration of BRCA1. Because most functions ascribed to BRCA1 occur within the nucleus, we postulated that phosphorylation-dependent nuclear accumulation of BRCA1 would result in enhanced nuclear activity, specifically transcriptional activity, of BRCA1. This postulate is affirmed by our observation that the ability of BRCA1 to transactivate GADD45 promoter constructs was enhanced in T47D cells treated with heregulin {beta}1. Furthermore, the heterologous expression of BRCA1 in HCC1937 human breast cancer cells, which have constitutively active Akt, also induces GADD45 promoter activity, whereas the expression of BRCA1 in which threonine 509 has been mutated to an alanine is able to only minimally induce promoter activity. These findings implicate Akt in upstream events leading to BRCA1 nuclear localization and function.

  5. Subcutaneous adipocytes promote melanoma cell growth by activating the Akt signaling pathway: role of palmitic acid.

    PubMed

    Kwan, Hiu Yee; Fu, Xiuqiong; Liu, Bin; Chao, Xiaojuan; Chan, Chi Leung; Cao, Huihui; Su, Tao; Tse, Anfernee Kai Wing; Fong, Wang Fun; Yu, Zhi-Ling

    2014-10-31

    Tumorigenesis involves constant communication between tumor cells and neighboring normal cells such as adipocytes. The canonical function of adipocytes is to store triglyceride and release fatty acids for other tissues. This study was aimed to find out if adipocytes promoted melanoma cell growth and to investigate the underlying mechanism. Here we isolated adipocytes from inguinal adipose tissue in mice and co-cultured with melanoma cells. We found that the co-cultured melanoma had higher lipid accumulation compared with mono-cultured melanoma. In addition, fluorescently labeled fatty acid BODIPY® FLC16 signal was detected in melanoma co-cultured with the adipocytes that had been loaded with the fluorescent dye, suggesting that the adipocytes provide fatty acids to melanoma cells. Compared with mono-cultured melanoma, co-cultured melanoma cells had a higher proliferation and phospho-Akt (Ser-473 and Thr-450) expression. Overexpression of Akt mutants in melanoma cells reduced the co-culture-enhanced proliferation. A lipidomic study showed that the co-cultured melanoma had an elevated palmitic acid level. Interestingly, we found that palmitic acid stimulated melanoma cell proliferation, changed the cell cycle distribution, and increased phospho-Akt (Ser-473 and Thr-450) and PI3K but not phospho-PTEN (phosphophosphatase and tensin homolog) expressions. More importantly, the palmitic acid-stimulated proliferation was further enhanced in the Akt-overexpressed melanoma cells and was reduced by LY294002 or knockdown of endogenous Akt or overexpression of Akt mutants. We also found that palmitic acid-pretreated B16F10 cells were grown to a significantly larger tumor in mice compared with control cells. Taken together, we suggest that adipocytes may serve as an exogenous source of palmitic acid that promotes melanoma cell growth by activating Akt.

  6. Subcutaneous Adipocytes Promote Melanoma Cell Growth by Activating the Akt Signaling Pathway

    PubMed Central

    Kwan, Hiu Yee; Fu, Xiuqiong; Liu, Bin; Chao, Xiaojuan; Chan, Chi Leung; Cao, Huihui; Su, Tao; Tse, Anfernee Kai Wing; Fong, Wang Fun; Yu, Zhi-Ling

    2014-01-01

    Tumorigenesis involves constant communication between tumor cells and neighboring normal cells such as adipocytes. The canonical function of adipocytes is to store triglyceride and release fatty acids for other tissues. This study was aimed to find out if adipocytes promoted melanoma cell growth and to investigate the underlying mechanism. Here we isolated adipocytes from inguinal adipose tissue in mice and co-cultured with melanoma cells. We found that the co-cultured melanoma had higher lipid accumulation compared with mono-cultured melanoma. In addition, fluorescently labeled fatty acid BODIPY® FLC16 signal was detected in melanoma co-cultured with the adipocytes that had been loaded with the fluorescent dye, suggesting that the adipocytes provide fatty acids to melanoma cells. Compared with mono-cultured melanoma, co-cultured melanoma cells had a higher proliferation and phospho-Akt (Ser-473 and Thr-450) expression. Overexpression of Akt mutants in melanoma cells reduced the co-culture-enhanced proliferation. A lipidomic study showed that the co-cultured melanoma had an elevated palmitic acid level. Interestingly, we found that palmitic acid stimulated melanoma cell proliferation, changed the cell cycle distribution, and increased phospho-Akt (Ser-473 and Thr-450) and PI3K but not phospho-PTEN (phosphophosphatase and tensin homolog) expressions. More importantly, the palmitic acid-stimulated proliferation was further enhanced in the Akt-overexpressed melanoma cells and was reduced by LY294002 or knockdown of endogenous Akt or overexpression of Akt mutants. We also found that palmitic acid-pretreated B16F10 cells were grown to a significantly larger tumor in mice compared with control cells. Taken together, we suggest that adipocytes may serve as an exogenous source of palmitic acid that promotes melanoma cell growth by activating Akt. PMID:25228694

  7. Myostatin signaling regulates Akt activity via the regulation of miR-486 expression.

    PubMed

    Hitachi, Keisuke; Nakatani, Masashi; Tsuchida, Kunihiro

    2014-02-01

    Myostatin, also known as growth and differentiation factor-8, is a pivotal negative regulator of skeletal muscle mass and reduces muscle protein synthesis by inhibiting the insulin-like growth factor-1 (IGF-1)/Akt/mammalian target of rapamycin (mTOR) pathway. However, the precise mechanism by which myostatin inhibits the IGF-1/Akt/mTOR pathway remains unclear. In this study, we investigated the global microRNA expression profile in myostatin knockout mice and identified miR-486, a positive regulator of the IGF-1/Akt pathway, as a novel target of myostatin signaling. In myostatin knockout mice, the expression level of miR-486 in skeletal muscle was significantly increased. In addition, we observed increased expression of the primary transcript of miR-486 (pri-miR-486) and Ankyrin 1.5 (Ank1.5), the host gene of miR-486, in myostatin knockout mice. In C2C12 cells, myostatin negatively regulated the expression of Ank1.5. Moreover, canonical myostatin signaling repressed the skeletal muscle-specific promoter activity of miR-486/Ank1.5. This repression was partially mediated by the E-box elements in the proximal region of the promoter. We also show that overexpression of miR-486 induced myotube hypertrophy in vitro and that miR-486 was essential to maintain skeletal muscle size both in vitro and in vivo. In addition, inhibition of miR-486 led to a decrease in Akt activity in C2C12 myotubes. Our findings indicate that miR-486 is one of the intermediary molecules connecting myostatin signaling and the IGF-1/Akt/mTOR pathway in the regulation of skeletal muscle size.

  8. AKT/PKB Signaling: Navigating Downstream

    PubMed Central

    Manning, Brendan D.; Cantley, Lewis C.

    2009-01-01

    The serine/threonine kinase Akt, also known as protein kinase B (PKB), is a central node in cell signaling downstream of growth factors, cytokines, and other cellular stimuli. Aberrant loss or gain of Akt activation underlies the pathophysiological properties of a variety of complex diseases, including type-2 diabetes and cancer. Here, we review the molecular properties of Akt and the approaches used to characterize its true cellular targets. In addition, we discuss those Akt substrates that are most likely to contribute to the diverse cellular roles of Akt, which include cell survival, growth, proliferation, angiogenesis, metabolism, and migration. PMID:17604717

  9. Deoxycholyltaurine Rescues Human Colon Cancer Cells From Apoptosis by Activating EGFR-Dependent PI3K/Akt Signaling

    PubMed Central

    Raufman, Jean-Pierre; Shant, Jasleen; Guo, Chang Yue; Roy, Sanjit; Cheng, Kunrong

    2010-01-01

    Recent studies indicate that secondary bile acids promote colon cancer cell proliferation but their role in maintaining cell survival has not been explored. We found that deoxycholyltaurine (DCT) markedly attenuated both unstimulated and TNF-α-stimulated programmed cell death in colon cancer cells by a phosphatidylinositol 3-kinase (PI3K)-dependent mechanism. To examine the role of bile acids and PI3K signaling in maintaining colon cancer cell survival, we explored the role of signaling downstream of bile acid-induced activation of the epidermal growth factor receptor (EGFR) in regulating both apoptosis and proliferation of HT-29 and H508 human colon cancer cells. DCT caused dose- and time-dependent Akt (Ser473) phosphorylation, a commonly used marker of activated PI3K/Akt signaling. Both EGFR kinase and PI3K inhibitors attenuated DCT-induced Akt phosphorylation and Akt activation, as demonstrated by reduced phosphorylation of a GSK-3-paramyosin substrate. Transfection of HT-29 cells with kinase-dead EGFR (K721M) reduced DCT-induced Akt phosphorylation. In HT-29 cells, EGFR and PI3K inhibitors as well as transfection with dominant negative AKT attenuated DCT-induced cell proliferation. DCT-induced PI3K/Akt activation resulted in downstream phosphorylation of GSK-3 (Ser21/9) and BAD (Ser136), and nuclear translocation (activation) of NF-κB, thereby confirming that DCT-induced activation of PI3K/Akt signaling regulates both proproliferative and prosurvival signals. Collectively, these results indicate that DCT-induced activation of post-EGFR PI3K/Akt signaling stimulates both colon cancer cell survival and proliferation. PMID:18064605

  10. Zinc induces cell death in immortalized embryonic hippocampal cells via activation of Akt-GSK-3beta signaling.

    PubMed

    Min, Young Kyu; Lee, Jong Eun; Chung, Kwang Chul

    2007-01-15

    Zinc is an essential catalytic and structural element of many proteins and a signaling messenger that is released by neuronal activity at many central excitatory synapses. Excessive synaptic release of zinc followed by entry into vulnerable neurons contributes severe neuronal cell death. We have previously observed that zinc-induced neuronal cell death is accompanied by Akt activation in embryonic hippocampal progenitor (H19-7) cells. In the present study, we examined the role of Akt activation and its downstream signaling events during extracellular zinc-induced neuronal cell death. Treatment of H19-7 cells with 10 microM of zinc plus zinc ionophore, pyrithione, led to increased phosphorylation of Akt at Ser-473/Thr-308 and increased Akt kinase activity. Zinc-induced Akt activation was accompanied by increased Tyr-phosphorylated GSK-3beta as well as increased GSK-3beta kinase activity. Transient overexpression of a kinase-deficient Akt mutant remarkably suppressed GSK-3beta activation and cell death. Furthermore, tau phosphorylation, but not the degradation of beta-catenin, was dependent upon zinc-induced GSK-3beta activation and contributed to cell death. The current data suggest that, following exposure to zinc, the sequential activation of Akt and GSK-3beta plays an important role directing hippocampal neural precursor cell death.

  11. SC79 protects retinal pigment epithelium cells from UV radiation via activating Akt-Nrf2 signaling

    PubMed Central

    Cao, Guo-fan; Cao, Cong; Jiang, Qin

    2016-01-01

    Excessive Ultra-violet (UV) radiation causes oxidative damages and apoptosis in retinal pigment epithelium (RPE) cells. Here we tested the potential activity of SC79, a novel small molecule activator of Akt, against the process. We showed that SC79 activated Akt in primary and established (ARPE-19 line) RPE cells. It protected RPE cells from UV damages possibly via inhibiting cell apoptosis. Akt inhibition, via an Akt specific inhibitor (MK-2206) or Akt1 shRNA silence, almost abolished SC79-induced RPE cytoprotection. Further studies showed that SC79 activated Akt-dependent NF-E2-related factor 2 (Nrf2) signaling and inhibited UV-induced oxidative stress in RPE cells. Reversely, Nrf2 shRNA knockdown or S40T mutation attenuated SC79-induced anti-UV activity. For the in vivo studies, we showed that intravitreal injection of SC79 significantly protected mouse retina from light damages. Based on these results, we suggest that SC79 protects RPE cells from UV damages possibly via activating Akt-Nrf2 signaling axis. PMID:27517753

  12. SC79 protects retinal pigment epithelium cells from UV radiation via activating Akt-Nrf2 signaling.

    PubMed

    Gong, Yi-Qing; Huang, Wei; Li, Ke-Ran; Liu, Yuan-Yuan; Cao, Guo-Fan; Cao, Cong; Jiang, Qin

    2016-09-13

    Excessive Ultra-violet (UV) radiation causes oxidative damages and apoptosis in retinal pigment epithelium (RPE) cells. Here we tested the potential activity of SC79, a novel small molecule activator of Akt, against the process. We showed that SC79 activated Akt in primary and established (ARPE-19 line) RPE cells. It protected RPE cells from UV damages possibly via inhibiting cell apoptosis. Akt inhibition, via an Akt specific inhibitor (MK-2206) or Akt1 shRNA silence, almost abolished SC79-induced RPE cytoprotection. Further studies showed that SC79 activated Akt-dependent NF-E2-related factor 2 (Nrf2) signaling and inhibited UV-induced oxidative stress in RPE cells. Reversely, Nrf2 shRNA knockdown or S40T mutation attenuated SC79-induced anti-UV activity. For the in vivo studies, we showed that intravitreal injection of SC79 significantly protected mouse retina from light damages. Based on these results, we suggest that SC79 protects RPE cells from UV damages possibly via activating Akt-Nrf2 signaling axis.

  13. Akt-mTORC1 signaling regulates Acly to integrate metabolic input to control of macrophage activation

    PubMed Central

    Covarrubias, Anthony J; Aksoylar, Halil Ibrahim; Yu, Jiujiu; Snyder, Nathaniel W; Worth, Andrew J; Iyer, Shankar S; Wang, Jiawei; Ben-Sahra, Issam; Byles, Vanessa; Polynne-Stapornkul, Tiffany; Espinosa, Erika C; Lamming, Dudley; Manning, Brendan D; Zhang, Yijing; Blair, Ian A; Horng, Tiffany

    2016-01-01

    Macrophage activation/polarization to distinct functional states is critically supported by metabolic shifts. How polarizing signals coordinate metabolic and functional reprogramming, and the potential implications for control of macrophage activation, remains poorly understood. Here we show that IL-4 signaling co-opts the Akt-mTORC1 pathway to regulate Acly, a key enzyme in Ac-CoA synthesis, leading to increased histone acetylation and M2 gene induction. Only a subset of M2 genes is controlled in this way, including those regulating cellular proliferation and chemokine production. Moreover, metabolic signals impinge on the Akt-mTORC1 axis for such control of M2 activation. We propose that Akt-mTORC1 signaling calibrates metabolic state to energetically demanding aspects of M2 activation, which may define a new role for metabolism in supporting macrophage activation. DOI: http://dx.doi.org/10.7554/eLife.11612.001 PMID:26894960

  14. Akt-mTORC1 signaling regulates Acly to integrate metabolic input to control of macrophage activation.

    PubMed

    Covarrubias, Anthony J; Aksoylar, Halil Ibrahim; Yu, Jiujiu; Snyder, Nathaniel W; Worth, Andrew J; Iyer, Shankar S; Wang, Jiawei; Ben-Sahra, Issam; Byles, Vanessa; Polynne-Stapornkul, Tiffany; Espinosa, Erika C; Lamming, Dudley; Manning, Brendan D; Zhang, Yijing; Blair, Ian A; Horng, Tiffany

    2016-02-19

    Macrophage activation/polarization to distinct functional states is critically supported by metabolic shifts. How polarizing signals coordinate metabolic and functional reprogramming, and the potential implications for control of macrophage activation, remains poorly understood. Here we show that IL-4 signaling co-opts the Akt-mTORC1 pathway to regulate Acly, a key enzyme in Ac-CoA synthesis, leading to increased histone acetylation and M2 gene induction. Only a subset of M2 genes is controlled in this way, including those regulating cellular proliferation and chemokine production. Moreover, metabolic signals impinge on the Akt-mTORC1 axis for such control of M2 activation. We propose that Akt-mTORC1 signaling calibrates metabolic state to energetically demanding aspects of M2 activation, which may define a new role for metabolism in supporting macrophage activation.

  15. Activation of the EGFR/Akt/NF-κB/cyclinD1 survival signaling pathway in human cholesteatoma epithelium.

    PubMed

    Liu, Wei; Yin, Tuanfang; Ren, Jihao; Li, Lihua; Xiao, Zian; Chen, Xing; Xie, Dinghua

    2014-02-01

    Cholesteatoma is a benign keratinizing squamous epithelial lesion characterized by the hyper-proliferation of keratinocytes with abundant production of keratin debris in the middle ear. The epidermal growth factor receptor (EGFR)/Akt/nuclear factor-kappa B (NF-κB)/cyclinD1 signaling pathway is one of the most important pathways in regulating cell survival and proliferation. We hypothesized that the EGFR/Akt/NF-κB/cyclinD1 signaling pathway may be activated and involved in the cellular hyperplasia mechanism in acquired cholesteatoma epithelium. Immunohistochemical staining of phosphorylated EGFR (p-EGFR), phosphorylated Akt (p-Akt), activated NF-κB and cyclinD1 protein was performed in 40 cholesteatoma samples and 20 samples of normal external auditory canal (EAC) epithelium. Protein expression of p-EGFR, p-Akt, activated NF-κB and cyclinD1 in cholesteatoma epithelium was significantly increased when compared with normal EAC epithelium (p < 0.01). In cholesteatoma epithelium, a significant positive association was observed between p-EGFR and p-Akt expression and between the expressions of p-Akt and NF-κB, NF-κB and cyclinD1, respectively (p < 0.01). No significant relationships were observed between the levels of investigated proteins and the degree of bone destruction (p > 0.05). The increased protein expression of p-EGFR, p-Akt, NF-κB and cyclinD1 and their associations in cholesteatoma epithelium suggest that the EGFR/Akt/NF-κB/cyclinD1 survival signaling pathway is active and may be involved in the regulatory mechanisms of cellular hyperplasia in cholesteatoma epithelium.

  16. Effects of AFP-activated PI3K/Akt signaling pathway on cell proliferation of liver cancer.

    PubMed

    Zheng, Lu; Gong, Wei; Liang, Ping; Huang, XiaoBing; You, Nan; Han, Ke Qiang; Li, Yu Ming; Li, Jing

    2014-05-01

    This study aims to investigate effects of alpha-fetoprotein (AFP)-activated phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signaling pathway on hepatocellular carcinoma cell proliferation. Active cirrhosis patients after hepatitis B infection (n = 20) and viral hepatitis patients with hepatocellular carcinoma (HCC) (n = 20) were selected as the subjects of the present study. Another 20 healthy subjects were selected as the control group. The serum AFP expression and liver tissue PI3K and Akt gene mRNA expression were detected. The hepatoma cell model HepG2 which had a stable expression of AFP gene was used. Real-time quantitative PCR and Western blot and other methods were used to analyze the intracellular PI3K and Akt protein levels. Compared with control group and cirrhosis group, the serum AFP levels in HCC group significantly increased, and the tissue PI3K and Akt mRNA expression also significantly increased. HepG2 cells were intervened using AFP, in which the PIK and Akt protein expression significantly increased. After intervention by use of AFP monoclonal antibodies or LY294002 inhibitor, the PIK and Akt protein expression in HepG2 cell was significantly decreased (P < 0.05). AFP can promote the proliferation of hepatoma cells via activation of PI3K/Akt signaling pathway.

  17. Epigenetic silencing mediated through activated PI3K/AKT signaling in breast cancer.

    PubMed

    Zuo, Tao; Liu, Ta-Ming; Lan, Xun; Weng, Yu-I; Shen, Rulong; Gu, Fei; Huang, Yi-Wen; Liyanarachchi, Sandya; Deatherage, Daniel E; Hsu, Pei-Yin; Taslim, Cenny; Ramaswamy, Bhuvaneswari; Shapiro, Charles L; Lin, Huey-Jen L; Cheng, Alfred S L; Jin, Victor X; Huang, Tim H-M

    2011-03-01

    Trimethylation of histone 3 lysine 27 (H3K27me3) is a critical epigenetic mark for the maintenance of gene silencing. Additional accumulation of DNA methylation in target loci is thought to cooperatively support this epigenetic silencing during tumorigenesis. However, molecular mechanisms underlying the complex interplay between the two marks remain to be explored. Here we show that activation of PI3K/AKT signaling can be a trigger of this epigenetic processing at many downstream target genes. We also find that DNA methylation can be acquired at the same loci in cancer cells, thereby reinforcing permanent repression in those losing the H3K27me3 mark. Because of a link between PI3K/AKT signaling and epigenetic alterations, we conducted epigenetic therapies in conjunction with the signaling-targeted treatment. These combined treatments synergistically relieve gene silencing and suppress cancer cell growth in vitro and in xenografts. The new finding has important implications for improving targeted cancer therapies in the future.

  18. Epigenetic Silencing Mediated Through Activated PI3K/AKT Signaling in Breast Cancer

    PubMed Central

    Zuo, Tao; Liu, Ta-Ming; Lan, Xun; Weng, Yu-I; Shen, Rulong; Gu, Fei; Huang, Yi-Wen; Liyanarachchi, Sandya; Deatherage, Daniel E.; Hsu, Pei-Yin; Taslim, Cenny; Ramaswamy, Bhuvaneswari; Shapiro, Charles L.; Lin, Huey-Jen L.; Cheng, Alfred S.L.; Jin, Victor X.; Huang, Tim H.-M.

    2011-01-01

    Trimethylation of histone 3 lysine 27 (H3K27me3) is a critical epigenetic mark for the maintenance of gene silencing. Additional accumulation of DNA methylation in target loci is thought to cooperatively support this epigenetic silencing during tumorigenesis. However, molecular mechanisms underlying the complex interplay between the two marks remain to be explored. Here we demonstrate that activation of PI3K/AKT signaling can be a trigger of this epigenetic processing at many downstream target genes. We also find that DNA methylation can be acquired at the same loci in cancer cells, thereby reinforcing permanent repression in those losing the H3K27me3 mark. Because of a link between PI3K/AKT signaling and epigenetic alterations, we conducted epigenetic therapies in conjunction with the signaling-targeted treatment. These combined treatments synergistically relieve gene silencing and suppress cancer cell growth in vitro and in xenografts. The new finding has important implications for improving targeted cancer therapies in the future. PMID:21216892

  19. Berberine ameliorates hyperglycemia in alloxan-induced diabetic C57BL/6 mice through activation of Akt signaling pathway.

    PubMed

    Xie, Xi; Li, Wenyuan; Lan, Tian; Liu, Weihua; Peng, Jing; Huang, Kaipeng; Huang, Juan; Shen, Xiaoyan; Liu, Peiqing; Huang, Heqing

    2011-01-01

    Recently, it is implicated that the abnormality of Akt signaling pathway is involved in the diabetic pathology. Previous studies have demonstrated that berberine could decrease blood glucose by elevating liver glycogen synthesis. However, the underlying mechanism is still unclear. In the present study, we investigated the effects of berberine on fasting blood glucose, liver glycogen, Akt, Glycogen synthase kinase-3, glucokinase and insulin receptor substrate (IRS) in alloxan-induced diabetic mice, exploring its possible hypoglycemic mechanism. We found that in alloxan-induced diabetic mice, the high blood glucose was significantly lowered by berberine treatment. Liver glycogen content, the expression and activity of glucokinase and the phosphorylated Akt and IRS were all significantly reduced in diabetic mice whereas berberine blocked these changes. Berberine also depressed the increasing of phosphorylated GSK-3β in diabetic mice. Collectively, Berberine upregulates the activity of Akt possibly via insulin signaling pathway, eventually lowering high blood glucose in alloxan-induced diabetic mice.

  20. Control of macrophage metabolism and activation by mTOR and Akt signaling

    PubMed Central

    Covarrubias, Anthony J.; Aksoylar, H. Ibrahim; Horng, Tiffany

    2015-01-01

    Macrophages are pleiotropic cells that assume a variety of functions depending on their tissue of residence and tissue state. They maintain homeostasis as well as coordinate responses to stresses such as infection and metabolic challenge. The ability of macrophages to acquire diverse, context-dependent activities requires their activation (or polarization) to distinct functional states. While macrophage activation is well understood at the level of signal transduction and transcriptional regulation, the metabolic underpinnings are poorly understood. Importantly, emerging studies indicate that metabolic shifts play a pivotal role in control of macrophage activation and acquisition of context-dependent effector activities. The signals that drive macrophage activation impinge on metabolic pathways, allowing for coordinate control of macrophage activation and metabolism. Here we discuss how mTOR and Akt, major metabolic regulators and targets of such activation signals, control macrophage metabolism and activation. Dysregulated macrophage activities contribute to many diseases, including infectious, inflammatory, and metabolic diseases and cancer, thus a better understanding of metabolic control of macrophage activation could pave the way to the development of new therapeutic strategies. PMID:26360589

  1. Control of macrophage metabolism and activation by mTOR and Akt signaling.

    PubMed

    Covarrubias, Anthony J; Aksoylar, H Ibrahim; Horng, Tiffany

    2015-08-01

    Macrophages are pleiotropic cells that assume a variety of functions depending on their tissue of residence and tissue state. They maintain homeostasis as well as coordinate responses to stresses such as infection and metabolic challenge. The ability of macrophages to acquire diverse, context-dependent activities requires their activation (or polarization) to distinct functional states. While macrophage activation is well understood at the level of signal transduction and transcriptional regulation, the metabolic underpinnings are poorly understood. Importantly, emerging studies indicate that metabolic shifts play a pivotal role in control of macrophage activation and acquisition of context-dependent effector activities. The signals that drive macrophage activation impinge on metabolic pathways, allowing for coordinate control of macrophage activation and metabolism. Here we discuss how mTOR and Akt, major metabolic regulators and targets of such activation signals, control macrophage metabolism and activation. Dysregulated macrophage activities contribute to many diseases, including infectious, inflammatory, and metabolic diseases and cancer, thus a better understanding of metabolic control of macrophage activation could pave the way to the development of new therapeutic strategies.

  2. Withaferin A inhibits matrix metalloproteinase-9 activity by suppressing the Akt signaling pathway.

    PubMed

    Lee, Dae Hyung; Lim, In-Hye; Sung, Eon-Gi; Kim, Joo-Young; Song, In-Hwan; Park, Yoon Ki; Lee, Tae-Jin

    2013-08-01

    Withaferin A (Wit A), a steroidal lactone isolated from Withania somnifera, exhibits anti-inflammatory, immuno-modulatory and anti-angiogenic properties and antitumor activities. In the present study, we investigated the effects of Wit A on protease-mediated invasiveness of the human metastatic cancer cell lines Caski and SK-Hep1. We found that treatment with Wit A resulted in marked inhibition of the TGF‑β‑induced increase in expression and activity of matrix metalloproteinase (MMP)‑9 in Caski cell line. These effects of Wit A were dose-dependent and showed a correlation with suppression of MMP‑9 mRNA expression levels. Treatment with Wit A resulted in an ~1.6-fold induction of MMP-9 promoter activity, which was also suppressed by treatment with Wit A in Caski cells. We found that treatment with Wit A resulted in inhibition of TGF‑β‑induced phosphorylation of Akt, which was involved in the downregulation of expression of MMP-9 at the protein level. Introduction with constitutively active (CA)‑Akt resulted in a partial increase in the secretion of TGF-β-induced MMP-9 blocked by treatment with Wit A in Caski cells. According to these results, Wit A may inhibit the invasive and migratory abilities of Caski cells through a reduction in MMP-9 expression through suppression of the pAkt signaling pathway. These findings indicate that use of Wit A may be an effective strategy for control of metastasis and invasiveness of tumors.

  3. Akt signaling leads to stem cell activation and promotes tumor development in epidermis.

    PubMed

    Segrelles, Carmen; García-Escudero, Ramón; Garín, Maria I; Aranda, Juan F; Hernández, Pilar; Ariza, José M; Santos, Mirentxu; Paramio, Jesús M; Lorz, Corina

    2014-07-01

    Hair follicle stem cells (HF-SCs) alternate between periods of quiescence and proliferation, to finally differentiate into all the cell types that constitute the hair follicle. Also, they have been recently identified as cells of origin in skin cancer. HF-SCs localize in a precise region of the hair follicle, the bulge, and molecular markers for this population have been established. Thus, HF-SCs are good model to study the potential role of oncogenic activations on SC physiology. Expression of a permanently active form of Akt (myrAkt) in basal cells leads to Akt hyperactivation specifically in the CD34(+)Itga6(H) population. This activation causes bulge stem cells to exit from quiescence increasing their response to proliferative stimuli and affecting some functions such as cell migration. HF-SC identity upon Akt activation is preserved; in this sense, increased proliferation does not result in stem cell exhaustion with age suggesting that Akt activation does not affect self-renewal an important aspect for normal tissue maintenance and cancer development. Genome-wide transcriptome analysis of HF-SC isolated from myrAkt and wild-type epidermis underscores changes in metabolic pathways characteristic of cancer cells. These differences manifest during a two-step carcinogenesis protocol in which Akt activation in HF-SCs results in increased tumor development and malignant transformation.

  4. Bamboo leaf extract ameliorates diabetic nephropathy through activating the AKT signaling pathway in rats.

    PubMed

    Ying, Changjiang; Mao, Yizhen; Chen, Lei; Wang, Shanshan; Ling, Hongwei; Li, Wei; Zhou, Xiaoyan

    2017-03-27

    Cleaved Caspase-3 levels in STZ-diabetic rats. In conclusion, our study suggested that bamboo leaf extract ameliorated DN in diabetic rats, and this protective effect is possibly related to suppressing oxidative stress through activating AKT signaling pathway. Bamboo leaf extract treatment may be a potential promising therapy for DN.

  5. Oxidative stress induces proliferation of colorectal cancer cells by inhibiting RUNX3 and activating the Akt signaling pathway.

    PubMed

    Kang, Kyoung Ah; Kim, Ki Cheon; Bae, Suk Chul; Hyun, Jin Won

    2013-11-01

    We recently reported that the tumor suppressor Runt-related transcription factor 3 (RUNX3) is silenced in colorectal cancer cells via oxidative stress-induced hypermethylation of its promoter. The resulting downregulation of RUNX3 expression influences cell proliferation. Activation of the Akt signaling pathway is also associated with cell survival and proliferation; however, the effects of oxidative stress on the relationship between RUNX3 and Akt signaling are largely unknown. Therefore, this study investigated the mechanisms involved in cell proliferation caused by oxidative stress-induced silencing of RUNX3. The levels of RUNX3 mRNA and protein were downregulated in response to treatment of the human colorectal cancer cell line SNU-407 with H2O2. Treatment of the cells with H2O2 also upregulated Akt mRNA and protein expression, and inhibited the binding of RUNX3 to the Akt promoter. The inverse correlation between the expression levels of RUNX3 and Akt in H2O2-treated cells was also associated with nuclear translocation of β-catenin and upregulation of cyclin D1 expression, which induced cell proliferation. H2O2 treatment also increased the binding of β-catenin to the cyclin D1 promoter. The results presented here demonstrate that reactive oxygen species silence the tumor suppressor RUNX3, enhance the Akt-mediated signaling pathway, and promote the proliferation of colorectal cancer cells.

  6. Intracellular calcium promotes radioresistance of non-small cell lung cancer A549 cells through activating Akt signaling.

    PubMed

    Wang, Yiling; He, Jiantao; Zhang, Shenghui; Yang, Qingbo

    2017-03-01

    Radiotherapy is a major therapeutic approach in non-small cell lung cancer but is restricted by radioresistance. Although Akt signaling promotes radioresistance in non-small cell lung cancer, it is not well understood how Akt signaling is activated. Since intracellular calcium (Ca(2+)) could activate Akt in A549 cells, we investigated the relationship between intracellular calcium (Ca(2+)) and Akt signaling in radioresistant A549 cells by establishing radioresistant non-small cell lung cancer A549 cells. The radioresistant cell line A549 was generated by dose-gradient irradiation of the parental A549 cells. The cell viability, proliferation, and apoptosis were, respectively, assessed using the cell counting kit-8, EdU labeling, and flow cytometry analysis. The phosphorylation of Akt was evaluated by Western blotting, and the intracellular Ca(2+) concentration was assessed by Fluo 4-AM. The radioresistant A549 cells displayed mesenchymal morphology. After additional irradiation, the radioresistant A549 cells showed decreased cell viability and proliferation but increased apoptosis. Moreover, the intracellular Ca(2+) concentration and the phosphorylation level on the Akt473 site in radioresistant A549 cells were higher than those in original cells, whereas the percentage of apoptosis in radioresistant A549 cells was less. All these results could be reversed by verapamil. In conclusion, our study found that intracellular Ca(2+) could promote radioresistance of non-small cell lung cancer cells through phosphorylating of Akt on the 473 site, which contributes to a better understanding on the non-small cell lung cancer radioresistance, and may provide a new target for radioresistance management.

  7. Modulation in Activation and Expression of PTEN, Akt1, and PDK1: Further Evidence Demonstrating Altered Phosphoinositide 3-kinase Signaling in Postmortem Brain of Suicide Subjects

    PubMed Central

    Dwivedi, Yogesh; Rizavi, Hooriyah S.; Zhang, Hui; Roberts, Rosalinda C.; Conley, Robert R.; Pandey, Ghanshyam N.

    2010-01-01

    Background Phosphoinositide 3-kinase (PI 3-K) signaling plays a crucial role in neuronal growth and plasticity. Recently, we demonstrated that suicide brain is associated with decreased activation and expression of selective catalytic and regulatory subunits of PI 3-K. The present investigation examined the regulation and functional significance of compromised PI 3-K in suicide brain at the level of upstream phosphatase and tensin homolog on chromosome ten (PTEN) and downstream substrates 3-phosphoinositide-dependent kinase 1 (PDK1) and Akt. Method mRNA expression of Akt1, Akt3, PTEN, and PDK1 by competitive RT-PCR; protein expression of Akt1, Akt3, PTEN, PDK1, phosphorylated-Akt1 (Ser473), phosphorylated-Akt1(Thr308), phosphorylated-PDK1, and phosphorylated-PTEN by Western blot; and catalytic activities of Akt1, Akt3, and PDK1 by enzymatic assays were determined in prefrontal cortex (PFC) and hippocampus obtained from suicide subjects and nonpsychiatric controls. Results No significant changes in the expression of Akt1 or Akt3 were observed; however, catalytic activity of Akt1, but not of Akt3, was decreased in PFC and hippocampus of suicide subjects, which was associated with decreased phosphorylation of Akt1 at Ser473 and Thr308. The catalytic activity of PDK1 and the level of phosphorylated-PDK1 were also decreased in both brain areas without any change in expression levels of PDK1. On the other hand, mRNA and protein expression of PTEN was increased, whereas the level of phosphorylated-PTEN was decreased. Conclusion Our study demonstrates abnormalities in PI 3-K signaling at several levels in brain of suicide subjects and suggests the possible involvement of aberrant PI 3-K/Akt signaling in the pathogenic mechanisms of suicide. PMID:20163786

  8. Dysfunctional MnSOD leads to redox dysregulation and activation of prosurvival AKT signaling in uterine leiomyomas

    PubMed Central

    Vidimar, Vania; Gius, David; Chakravarti, Debabrata; Bulun, Serdar E.; Wei, Jian-Jun; Kim, J. Julie

    2016-01-01

    AKT signaling promotes cell growth and survival and is often dysregulated via multiple mechanisms in different types of cancer, including uterine leiomyomas (ULMs). ULMs are highly prevalent fibrotic tumors that arise from the smooth muscular layer of the uterus, the myometrium (MM). ULMs pose a major public health issue because they can cause severe morbidity and poor pregnancy outcomes. ‬We investigate the mechanisms driving ULM growth and survival via aberrant activation of AKT. We demonstrate that an acetylation-mediated impairment of the manganese superoxide dismutase (MnSOD) activity is prevalent in ULM cells compared to the normal-matched MM from the same patients. This impairment increases the levels of superoxide and oxidative stress, which activate AKT via oxidative inactivation of the phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Redox activation of AKT promotes ULM cell survival under conditions of moderate but persistent oxidative stress that are compatible with ULM’s prooxidative microenvironment. Moreover, because of impaired MnSOD activity, ULM cells are sensitive to high levels of reactive oxygen species (ROS) and superoxide-generating compounds, resulting in decreased ULM cell viability. On the contrary, MM cells with functional MnSOD are more resistant to high levels of oxidants. This study demonstrates a causative role of acetylation-mediated MnSOD dysfunction in activating prosurvival AKT signaling in ULMs. The specific AKT and redox states of ULM cells provide a potential novel therapeutic rationale to selectively target ULM cells because of their defective ROS-scavenging system.‬‬‬‬‬‬‬‬ PMID:27847869

  9. Erbin loss promotes cancer cell proliferation through feedback activation of Akt-Skp2-p27 signaling

    SciTech Connect

    Huang, Hao; Song, Yuhua; Wu, Yan; Guo, Ning; Ma, Yuanfang; Qian, Lu

    2015-07-31

    Erbin localizes at the basolateral membrane to regulate cell junctions and polarity in epithelial cells. Dysregulation of Erbin has been implicated in tumorigenesis, and yet it is still unclear if and how disrupted Erbin regulates the biological behavior of cancer cells. We report here that depletion of Erbin leads to cancer cell excessive proliferation in vitro and in vivo. Erbin deficiency accelerates S-phase entry by down-regulating CDK inhibitors p21 and p27 via two independent mechanisms. Mechanistically, Erbin loss promotes p27 degradation by enhancing E3 ligase Skp2 activity though augmenting Akt signaling. Interestingly, we also show that Erbin is an unstable protein when the Akt-Skp2 signaling is aberrantly activated, which can be specifically destructed by SCF-Skp2 ligase. Erbin loss facilitates cell proliferation and migration in Skp2-dependent manner. Thus, our finding illustrates a novel negative feedback loop between Erbin and Akt-Skp2 signaling. It suggests disrupted Erbin links polarity loss, hyperproliferation and tumorigenesis. - Highlights: • Erbin loss leads to cancer cell excessive proliferation in vitro and in vivo. • Erbin loss accelerates cell cycle though down-regulating p21 and p27 expression. • Erbin is a novel negative modulator of Akt1-Skp2-p27 signaling pathway. • Our study suggests that Erbin loss contributes to Skp2 oncogenic function.

  10. Juglanthraquinone C Induces Intracellular ROS Increase and Apoptosis by Activating the Akt/Foxo Signal Pathway in HCC Cells.

    PubMed

    Hou, Ya-Qin; Yao, Yao; Bao, Yong-Li; Song, Zhen-Bo; Yang, Cheng; Gao, Xiu-Li; Zhang, Wen-Jing; Sun, Lu-Guo; Yu, Chun-Lei; Huang, Yan-Xin; Wang, Guan-Nan; Li, Yu-Xin

    2016-01-01

    Juglanthraquinone C (JC), a naturally occurring anthraquinone extracted from Juglans mandshurica, could induce apoptosis of cancer cells. This study aims to investigate the detailed cytotoxicity mechanism of JC in HepG2 and BEL-7402 cells. The Affymetrix HG-U133 Plus 2.0 arrays were first used to analyze the mRNA expression exposed to JC or DMSO in HepG2 cells. Consistent with the previous results, the data indicated that JC could induce apoptosis and hyperactivated Akt. The Western blot analysis further revealed that Akt, a well-known survival protein, was strongly activated in HepG2 and BEL-7402 cells. Furthermore, an obvious inhibitory effect on JC-induced apoptosis was observed when the Akt levels were decreased, while the overexpression of constitutively active mutant Akt greatly accelerated JC-induced apoptosis. The subsequent results suggested that JC treatment suppressed nuclear localization and increased phosphorylated levels of Foxo3a, and the overexpression of Foxo3a abrogated JC-induced apoptosis. Most importantly, the inactivation of Foxo3a induced by JC further led to an increase of intracellular ROS levels by suppressing ROS scavenging enzymes, and the antioxidant N-acetyl-L-cysteine and catalase successfully decreased JC-induced apoptosis. Collectively, this study demonstrated that JC induced the apoptosis of hepatocellular carcinoma (HCC) cells by activating Akt/Foxo signaling pathway and increasing intracellular ROS levels.

  11. Stem cell factor (SCF) protects osteoblasts from oxidative stress through activating c-Kit-Akt signaling

    SciTech Connect

    Yang, Lei; Wu, Zhong; Yin, Gang; Liu, Haifeng; Guan, Xiaojun; Zhao, Xiaoqiang; Wang, Jianguang; Zhu, Jianguo

    2014-12-12

    Highlights: • SCF receptor c-Kit is functionally expressed in primary and transformed osteoblasts. • SCF protects primary and transformed osteoblasts from H{sub 2}O{sub 2}. • SCF activation of c-Kit in osteoblasts, required for its cyto-protective effects. • c-Kit mediates SCF-induced Akt activation in cultured osteoblasts. • Akt activation is required for SCF-regulated cyto-protective effects in osteoblasts. - Abstract: Osteoblasts regulate bone formation and remodeling, and are main target cells of oxidative stress in the progression of osteonecrosis. The stem cell factor (SCF)-c-Kit pathway plays important roles in the proliferation, differentiation and survival in a range of cell types, but little is known about its functions in osteoblasts. In this study, we found that c-Kit is functionally expressed in both osteoblastic-like MC3T3-E1 cells and primary murine osteoblasts. Its ligand SCF exerted significant cyto-protective effects against hydrogen peroxide (H{sub 2}O{sub 2}). SCF activated its receptor c-Kit in osteoblasts, which was required for its cyto-protective effects against H{sub 2}O{sub 2}. Pharmacological inhibition (by Imatinib and Dasatinib) or shRNA-mediated knockdown of c-Kit thus inhibited SCF-mediated osteoblast protection. Further investigations showed that protection by SCF against H{sub 2}O{sub 2} was mediated via activation of c-Kit-dependent Akt pathway. Inhibition of Akt activation, through pharmacological or genetic means, suppressed SCF-mediated anti-H{sub 2}O{sub 2} activity in osteoblasts. In summary, we have identified a new SCF-c-Kit-Akt physiologic pathway that protects osteoblasts from H{sub 2}O{sub 2}-induced damages, and might minimize the risk of osteonecrosis caused by oxidative stress.

  12. Metastasis and AKT activation.

    PubMed

    Qiao, Meng; Sheng, Shijie; Pardee, Arthur B

    2008-10-01

    Metastasis is responsible for 90% of cancer patient deaths. More information is needed about the molecular basis for its potential detection and treatment. The activated AKT kinase is necessary for many events of the metastatic pathway including escape of cells from the tumor's environment, into and then out of the circulation, activation of proliferation, blockage of apoptosis, and activation of angiogenesis. A series of steps leading to metastatic properties can be initiated upon activation of AKT by phosphorylation on Ser-473. These findings lead to the question of how this activation is connected to metastasis. Activated AKT phosphorylates GSK-3beta causing its proteolytic removal. This increases stability of the negative transcription factor SNAIL, thereby decreasing transcription of the transmembrane protein E-cadherin that forms adhesions between adjacent cells, thereby permitting their detachment. How is AKT hyperactivated in metastatic cells? Increased PI3K or TORC2 kinase activity- or decreased PHLPP phosphatase could be responsible. Furthermore, a positive feedback mechanism is that the decrease of E-cadherin lowers PTEN and thereby increases PIP3, further activating AKT and metastasis.

  13. FAM83B-mediated activation of PI3K/AKT and MAPK signaling cooperates to promote epithelial cell transformation and resistance to targeted therapies

    PubMed Central

    Cipriano, Rocky; Miskimen, Kristy L.S.; Bryson, Benjamin L.; Foy, Chase R.; Bartel, Courtney A.; Jackson, Mark W.

    2013-01-01

    Therapies targeting MAPK and AKT/mTOR signaling are currently being evaluated in clinical trials for several tumor types. However, recent studies suggest that these therapies may be limited due to acquired cancer cell resistance and a small therapeutic index between normal and cancer cells. The identification of novel proteins that are involved in MAPK or AKT/mTOR signaling and differentially expressed between normal and cancer cells will provide mechanistically distinct therapeutic targets with the potential to inhibit these key cancer-associated pathways. We recently identified FAM83B as a novel, previously uncharacterized oncogene capable of hyperactivating MAPK and mTOR signaling and driving the tumorigenicity of immortalized human mammary epithelial cells (HMEC). We show here that elevated FAM83B expression also activates the PI3K/AKT signaling pathway and confers a decreased sensitivity to PI3K, AKT, and mTOR inhibitors. FAM83B co-precipitated with the p85α and p110α subunits of PI3K, as well as AKT, and increased p110α and AKT membrane localization, consistent with elevated PI3K/AKT signaling. In tumor-derived cells harboring elevated FAM83B expression, ablation of FAM83B decreased p110α and AKT membrane localization, suppressed AKT phosphorylation, and diminished proliferation, AIG, and tumorigenicity in vivo. We propose that the level of FAM83B expression may be an important factor to consider when combined therapies targeting MAPK and AKT/mTOR signaling are used. Moreover, the identification of FAM83B as a novel oncogene and its integral involvement in activating PI3K/AKT and MAPK provides a foundation for future therapies aimed at targeting FAM83B in order to suppress the growth of PI3K/AKT- and MAPK-driven cancers. PMID:23676467

  14. Fucoidan/FGF-2 induces angiogenesis through JNK- and p38-mediated activation of AKT/MMP-2 signalling

    SciTech Connect

    Kim, Beom Su; Park, Ji-Yun; Kang, Hyo-Jin; Kim, Hyung-Jin; Lee, Jun

    2014-08-08

    Graphical abstract: Schematic diagram of the angiogenic activity mechanism by FGF-2/fucoidan treatment in HUVECs. Fucoidan enhances the FGF-2-induced phosphorylation of p38, JNK, and ERK MAPKs. However, p38 and JNK were involved in AKT phosphorylation and MMP-2 activation and resulted in enhanced angiogenic activity, such as tube formation and migration, in HUVECs. - Highlights: • The angiogenic activity of fucoidan in HUVECs was explored. • Fucoidan enhanced HUVEC proliferation, migration, and tube formation. • Fucoidan enhanced angiogenesis through p38 and JNK but not ERK in HUVECs. • Fucoidan targeted angiogenesis-mediated AKT/MMP-2 signalling in HUVECs. - Abstract: Angiogenesis is an important biological process in tissue development and repair. Fucoidan has previously been shown to potentiate in vitro tube formation in the presence of basic fibroblast growth factor (FGF-2). However, the underlying molecular mechanism remains largely unknown. This study was designed to investigate the action of fucoidan in angiogenesis in human umbilical vein endothelial cells (HUVECs) and to explore fucoidan-signalling pathways. First, we evaluated the effect of fucoidan on cell proliferation. Matrigel-based tube formation and wound healing assays were performed to investigate angiogenesis. Matrix metalloproteinase-2 (MMP-2) mRNA expression and activity levels were analysed by reverse transcription polymerase chain reaction (RT-PCR) and zymography, respectively. Additionally, phosphorylation of mitogen-activated protein kinases (MAPKs) and protein kinase B (AKT) was detected by Western blot. The results indicate that fucoidan treatment significantly increased cell proliferation in the presence of FGF-2. Moreover, compared to the effect of FGF-2 alone, fucoidan and FGF-2 had a greater effect on tube formation and cell migration, and this effect was found to be synergistic. Furthermore, fucoidan enhanced the phosphorylation of extracellular signal-regulated kinase (ERK

  15. Zinc promotes proliferation and activation of myogenic cells via the PI3K/Akt and ERK signaling cascade

    SciTech Connect

    Ohashi, Kazuya; Nagata, Yosuke; Wada, Eiji; Zammit, Peter S.; Shiozuka, Masataka; Matsuda, Ryoichi

    2015-05-01

    Skeletal muscle stem cells named muscle satellite cells are normally quiescent but are activated in response to various stimuli, such as injury and overload. Activated satellite cells enter the cell cycle and proliferate to produce a large number of myogenic progenitor cells, and these cells then differentiate and fuse to form myofibers. Zinc is one of the essential elements in the human body, and has multiple roles, including cell growth and DNA synthesis. However, the role of zinc in myogenic cells is not well understood, and is the focus of this study. We first examined the effects of zinc on differentiation of murine C2C12 myoblasts and found that zinc promoted proliferation, with an increased number of cells incorporating EdU, but inhibited differentiation with reduced myogenin expression and myotube formation. Furthermore, we used the C2C12 reserve cell model of myogenic quiescence to investigate the role of zinc on activation of myogenic cells. The number of reserve cells incorporating BrdU was increased by zinc in a dose dependent manner, with the number dramatically further increased using a combination of zinc and insulin. Akt and extracellular signal-regulated kinase (ERK) are downstream of insulin signaling, and both were phosphorylated after zinc treatment. The zinc/insulin combination-induced activation involved the phosphoinositide 3-kinase (PI3K)/Akt and ERK cascade. We conclude that zinc promotes activation and proliferation of myogenic cells, and this activation requires phosphorylation of PI3K/Akt and ERK as part of the signaling cascade. - Highlights: • Zinc has roles for promoting proliferation and inhibition differentiation of C2C12. • Zinc promotes activation of reserve cells. • Insulin and zinc synergize activation of reserve cells. • PI3K/Akt and ERK cascade affect zinc/insulin-mediated activation of reserve cells.

  16. Increase in reactive oxygen species and activation of Akt signaling pathway in neuropathic pain.

    PubMed

    Guedes, Renata P; Araújo, Alex S R; Janner, Daiane; Belló-Klein, Adriane; Ribeiro, Maria Flávia M; Partata, Wania A

    2008-12-01

    Neuropathic pain occurs as a result of peripheral or central nervous system injury. Its pathophysiology involves mainly a central sensitization mechanism that may be correlated to many molecules acting in regions involved in pain processing, such as the spinal cord. It has been demonstrated that reactive oxygen species (ROS) and signaling molecules, such as the serine/threonine protein kinase Akt, are involved in neuropathic pain mechanisms. Thus, the aim of this study was to provide evidence of this relationship. Sciatic nerve transection (SNT) was used to induce neuropathic pain in rats. Western blot analysis of Akt and 4-hydroxy-2-nonenal (HNE)-Michael adducts, and measurement of hydrogen peroxide (H(2)O(2)) in the lumbosacral spinal cord were performed. The main findings were found seven days after SNT, when there was an increase in HNE-Michael adducts formation, total and p-Akt expression, and H(2)O(2) concentration. However, one and 15 days after SNT, H(2)O(2) concentration was raised in both sham (animals that were submitted to surgery without nerve injury) and SNT groups, showing the high sensibility of this ROS to nociceptive afferent stimuli, not only to neuropathic pain. p-Akt also increased in sham and SNT groups one day post injury, but at 3 and 7 days the increase occurred exclusively in SNT animals. Thus, there is crosstalk between intracellular signaling pathways and ROS, and these molecules can act as protective agents in acute pain situations or play a role in the development of chronic pain states.

  17. PPAR-γ agonist stabilizes KLF4 protein via activating Akt signaling and reducing KLF4 ubiquitination

    SciTech Connect

    Sun, Yan; Zheng, Bin; Zhang, Xin-hua; He, Ming; Guo, Zong-wei; Wen, Jin-kun

    2014-01-10

    Highlights: •PPAR-γ increases KLF4 protein level but does not influence KLF4 gene transcription. •The increase of KLF4 protein levels induced by pioglitazone is PPAR-γ-dependent. •Pioglitazone stabilizes KLF4 protein via activating Akt signaling and reducing KLF4 ubiquitination. -- Abstract: Peroxisome proliferator activated receptor γ (PPAR-γ) plays important roles in cell cycle regulation, differentiation and apoptosis. Krüppel-like factor 4 (KLF4) modulates vascular smooth muscle cell (VSMC) phenotype. Both KLF4 and PPAR-γ are involved in VSMC proliferation and differentiation. However, the actual relationship between KLF4 and PPAR-γ in VSMCs is not clear. In this study, we found that PPAR-γ agonist pioglitazone increases KLF4 protein levels but does not influence KLF4 gene transcription. PPAR-γ overexpression increases, while PPAR-γ knockdown reduces KLF4 expression, suggesting that the increase in KLF4 protein levels induced by pioglitazone is PPAR-γ-dependent. Further study showed that pioglitazone enhances KLF4 protein stability through reducing KLF4 ubiquitination. Furthermore, we demonstrated that stabilization of KLF4 by pioglitazone was related to the activation of Akt signaling pathway. Taken together, we revealed that PPAR-γ agonist pioglitazone stabilizes KLF4 protein via activating Akt signaling and reducing KLF4 ubiquitination, providing further insights into PPAR-γ and KLF4 in regulating each other’s expression in VSMCs.

  18. Dystrophin glycoprotein complex-associated Gbetagamma subunits activate phosphatidylinositol-3-kinase/Akt signaling in skeletal muscle in a laminin-dependent manner.

    PubMed

    Xiong, Yongmin; Zhou, Yanwen; Jarrett, Harry W

    2009-05-01

    Previously, we showed that laminin-binding to the dystrophin glycoprotein complex (DGC) of skeletal muscle causes a heterotrimeric G-protein (Galphabetagamma) to bind, changing the activation state of the Gsalpha subunit. Others have shown that laminin-binding to the DGC also leads to Akt activation. Gbetagamma, released when Gsalpha is activated, is known to bind phosphatidylinositol-3-kinase (PI3K), which activates Akt in other cells. Here, we investigate whether muscle Akt activation results from Gbetagamma, using immunoprecipitation and immunoblotting, and purified Gbetagamma. In the presence of laminin, PI3K-binding to the DGC increases and Akt becomes phosphorylated and activated (pAkt), and glycogen synthase kinase is phosphorylated. Antibodies, which specifically block laminin-binding to alpha-dystroglycan, prevent PI3K-binding to the DGC. Purified bovine brain Gbetagamma also caused PI3K and Akt activation. These results show that DGC-Gbetagamma is binding PI3K and activating pAkt in a laminin-dependent manner. Mdx mice, which have greatly diminished amounts of DGC proteins, display elevated pAkt signaling and increased expression of integrin beta1 compared to normal muscle. This integrin binds laminin, Gbetagamma, and PI3K. Collectively, these suggest that PI3K is an important target for the Gbetagamma, which normally binds to DGC syntrophin, and activates PI3K/Akt signaling. Disruption of the DGC in mdx mouse is causing dis-regulation of the laminin-DGC-Gbetagamma-PI3K-Akt signaling and is likely to be important to the pathogenesis of muscular dystrophy. Upregulating integrin beta1 expression and activating the PI3K/Akt pathway in muscular dystrophy may partially compensate for the loss of the DGC. The results suggest new therapeutic approaches to muscle disease.

  19. Stem cell factor (SCF) protects osteoblasts from oxidative stress through activating c-Kit-Akt signaling.

    PubMed

    Yang, Lei; Wu, Zhong; Yin, Gang; Liu, Haifeng; Guan, Xiaojun; Zhao, Xiaoqiang; Wang, Jianguang; Zhu, Jianguo

    2014-12-12

    Osteoblasts regulate bone formation and remodeling, and are main target cells of oxidative stress in the progression of osteonecrosis. The stem cell factor (SCF)-c-Kit pathway plays important roles in the proliferation, differentiation and survival in a range of cell types, but little is known about its functions in osteoblasts. In this study, we found that c-Kit is functionally expressed in both osteoblastic-like MC3T3-E1 cells and primary murine osteoblasts. Its ligand SCF exerted significant cyto-protective effects against hydrogen peroxide (H₂O₂). SCF activated its receptor c-Kit in osteoblasts, which was required for its cyto-protective effects against H₂O₂. Pharmacological inhibition (by Imatinib and Dasatinib) or shRNA-mediated knockdown of c-Kit thus inhibited SCF-mediated osteoblast protection. Further investigations showed that protection by SCF against H₂O₂ was mediated via activation of c-Kit-dependent Akt pathway. Inhibition of Akt activation, through pharmacological or genetic means, suppressed SCF-mediated anti-H₂O₂ activity in osteoblasts. In summary, we have identified a new SCF-c-Kit-Akt physiologic pathway that protects osteoblasts from H₂O₂-induced damages, and might minimize the risk of osteonecrosis caused by oxidative stress.

  20. Fucoidan/FGF-2 induces angiogenesis through JNK- and p38-mediated activation of AKT/MMP-2 signalling.

    PubMed

    Kim, Beom Su; Park, Ji-Yun; Kang, Hyo-Jin; Kim, Hyung-Jin; Lee, Jun

    2014-08-08

    Angiogenesis is an important biological process in tissue development and repair. Fucoidan has previously been shown to potentiate in vitro tube formation in the presence of basic fibroblast growth factor (FGF-2). However, the underlying molecular mechanism remains largely unknown. This study was designed to investigate the action of fucoidan in angiogenesis in human umbilical vein endothelial cells (HUVECs) and to explore fucoidan-signalling pathways. First, we evaluated the effect of fucoidan on cell proliferation. Matrigel-based tube formation and wound healing assays were performed to investigate angiogenesis. Matrix metalloproteinase-2 (MMP-2) mRNA expression and activity levels were analysed by reverse transcription polymerase chain reaction (RT-PCR) and zymography, respectively. Additionally, phosphorylation of mitogen-activated protein kinases (MAPKs) and protein kinase B (AKT) was detected by Western blot. The results indicate that fucoidan treatment significantly increased cell proliferation in the presence of FGF-2. Moreover, compared to the effect of FGF-2 alone, fucoidan and FGF-2 had a greater effect on tube formation and cell migration, and this effect was found to be synergistic. Furthermore, fucoidan enhanced the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38, and AKT. MMP-2 activation was also significantly increased. Specific inhibitors of p38 (SB203580) and JNK (SP600125) inhibited tube formation and wound healing, while an ERK inhibitor (PD98059) did not. MMP-2 activation and AKT phosphorylation were also attenuated and associated with the suppression of p38 and JNK phosphorylation, but not with that of ERK. These results indicate that fucoidan, in the presence of FGF-2, induces angiogenesis through AKT/MMP-2 signalling by activating p38 and JNK. These findings provide basic molecular information on the effect of fucoidan on angiogenesis in the presence of FGF-2.

  1. Alpha-fetoprotein activates AKT/mTOR signaling to promote CXCR4 expression and migration of hepatoma cells.

    PubMed

    Zhu, Mingyue; Guo, Junli; Xia, Hua; Li, Wei; Lu, Yan; Dong, Xu; Chen, Yi; Xie, Xieju; Fu, Shigan; Li, Mengsen

    2015-01-01

    CXCR4, stromal cell-derived factor-1α(SDF 1α) receptor, stimulates growth and metastasis of hepatocellular carcinoma (HCC). Alpha-fetoprotein(AFP) governs the expression of some metastasis-related genes. Here we report that AFP and CXCR4 levels correlated in HCC tissues. AFP-expressing vectors induced CXCR4. In agreement, AFP depletion by siRNA decreased CXCR4. AFP co-localized and interacted with PTEN, thus inducing CXCR4 by activating AKT(Ser473) phosphorylation. In turn, phospho-mTOR(Ser2448) entered the nucleus and bound the CXCR4 gene promoter. Thus, AFP promoted migration of HCC cells. In concusion, AFP induced CXCR4 by activating the AKT/mTOR signal pathway.

  2. Alpha-fetoprotein activates AKT/mTOR signaling to promote CXCR4 expression and migration of hepatoma cells

    PubMed Central

    Li, Wei; Lu, Yan; Dong, Xu; Chen, Yi; Xie, Xieju; Fu, Shigan; Li, Mengsen

    2015-01-01

    CXCR4, stromal cell-derived factor-1α(SDF 1α) receptor, stimulates growth and metastasis of hepatocellular carcinoma (HCC). Alpha-fetoprotein(AFP) governs the expression of some metastasis-related genes. Here we report that AFP and CXCR4 levels correlated in HCC tissues. AFP-expressing vectors induced CXCR4. In agreement, AFP depletion by siRNA decreased CXCR4. AFP co-localized and interacted with PTEN, thus inducing CXCR4 by activating AKT(Ser473) phosphorylation. In turn, phospho-mTOR(Ser2448) entered the nucleus and bound the CXCR4 gene promoter. Thus, AFP promoted migration of HCC cells. In concusion, AFP induced CXCR4 by activating the AKT/mTOR signal pathway. PMID:25815363

  3. BCL-3 expression promotes colorectal tumorigenesis through activation of AKT signalling

    PubMed Central

    Urban, Bettina C; Collard, Tracey J; Eagle, Catherine J; Southern, Samantha L; Greenhough, Alexander; Hamdollah-Zadeh, Maryam; Ghosh, Anil; Paraskeva, Christos; Silver, Andrew; Williams, Ann C

    2016-01-01

    Objective Colorectal cancer remains the fourth most common cause of cancer-related mortality worldwide. Here we investigate the role of nuclear factor-κB (NF-κB) co-factor B-cell CLL/lymphoma 3 (BCL-3) in promoting colorectal tumour cell survival. Design Immunohistochemistry was carried out on 47 tumour samples and normal tissue from resection margins. The role of BCL-3/NF-κB complexes on cell growth was studied in vivo and in vitro using an siRNA approach and exogenous BCL-3 expression in colorectal adenoma and carcinoma cells. The question whether BCL-3 activated the AKT/protein kinase B (PKB) pathway in colorectal tumour cells was addressed by western blotting and confocal microscopy, and the ability of 5-aminosalicylic acid (5-ASA) to suppress BCL-3 expression was also investigated. Results We report increased BCL-3 expression in human colorectal cancers and demonstrate that BCL-3 expression promotes tumour cell survival in vitro and tumour growth in mouse xenografts in vivo, dependent on interaction with NF-κB p50 or p52 homodimers. We show that BCL-3 promotes cell survival under conditions relevant to the tumour microenvironment, protecting both colorectal adenoma and carcinoma cells from apoptosis via activation of the AKT survival pathway: AKT activation is mediated via both PI3K and mammalian target of rapamycin (mTOR) pathways, leading to phosphorylation of downstream targets GSK-3β and FoxO1/3a. Treatment with 5-ASA suppressed BCL-3 expression in colorectal cancer cells. Conclusions Our study helps to unravel the mechanism by which BCL-3 is linked to poor prognosis in colorectal cancer; we suggest that targeting BCL-3 activity represents an exciting therapeutic opportunity potentially increasing the sensitivity of tumour cells to conventional therapy. PMID:26033966

  4. DYRK1B blocks canonical and promotes non-canonical Hedgehog signaling through activation of the mTOR/AKT pathway

    PubMed Central

    Singh, Rajeev; Dhanyamraju, Pavan Kumar; Lauth, Matthias

    2017-01-01

    Hedgehog (Hh) signaling plays important roles in embryonic development and in tumor formation. Apart from the well-established stimulation of the GLI family of transcription factors, Hh ligands promote the phosphorylation and activation of mTOR and AKT kinases, yet the molecular mechanism underlying these processes are unknown. Here, we identify the DYRK1B kinase as a mediator between Hh signaling and mTOR/AKT activation. In fibroblasts, Hh signaling induces DYRK1B protein expression, resulting in activation of the mTOR/AKT kinase signaling arm. Furthermore, DYRK1B exerts positive and negative feedback regulation on the Hh pathway itself: It negatively interferes with SMO-elicited canonical Hh signaling, while at the same time it provides positive feed-forward functions by promoting AKT-mediated GLI stability. Due to the fact that the mTOR/AKT pathway is itself subject to strong negative feedback regulation, pharmacological inhibition of DYRK1B results in initial upregulation followed by downregulation of AKT phosphorylation and GLI stabilization. Addressing this issue therapeutically, we show that a pharmacological approach combining a DYRK1B antagonist with an mTOR/AKT inhibitor results in strong GLI1 targeting and in pronounced cytotoxicity in human pancreatic and ovarian cancer cells. PMID:27903983

  5. FoxM1 promotes breast tumorigenesis by activating PDGF-A and forming a positive feedback loop with the PDGF/AKT signaling pathway.

    PubMed

    Yu, Guanzhen; Zhou, Aidong; Xue, Jianfei; Huang, Chen; Zhang, Xia; Kang, Shin-Hyuk; Chiu, Wen-Tai; Tan, Christina; Xie, Keping; Wang, Jiejun; Huang, Suyun

    2015-05-10

    The autocrine platelet-derived growth factor (PDGF)/PDGF receptor (PDGFR) signaling pathway promotes breast cancer tumorigenesis, but the mechanisms for its dysregulation in breast cancer are largely unknown. In the study, we identified PDGF-A as a novel transcriptional target of FoxM1. FoxM1 directly binds to two sites in the promoter of PDGF-A and activates its transcription. Mutation of these FoxM1-binding sites diminished PDGF-A promoter activity. Increased FoxM1 resulted in the upregulation of PDGF-A, which led to activation of the AKT pathway and increased breast cancer cell proliferation and tumorigenesis, whereas knockdown of FoxM1 does the opposite. Blocking AKT activation with a phosphoinositide 3-kinase/AKT inhibitor decreased FoxM1-induced cell proliferation. Moreover, PDGF/AKT pathway upregulates the expression of FoxM1 in breast cancer cells. Knockdown of PDGF-A or blockade of AKT activation inhibited the expression of FoxM1 in breast cancer cells. Furthermore, expression of FoxM1 significantly correlated with the expression of PDGF-A and the activated AKT signaling pathway in human breast cancer specimens. Our study demonstrates a novel positive regulatory feedback loop between FoxM1 and the PDGF/AKT signaling pathway; this loop contributes to breast cancer cell growth and tumorigenesis.

  6. Methylmercury, an environmental electrophile capable of activation and disruption of the Akt/CREB/Bcl-2 signal transduction pathway in SH-SY5Y cells

    PubMed Central

    Unoki, Takamitsu; Abiko, Yumi; Toyama, Takashi; Uehara, Takashi; Tsuboi, Koji; Nishida, Motohiro; Kaji, Toshiyuki; Kumagai, Yoshito

    2016-01-01

    Methylmercury (MeHg) modifies cellular proteins via their thiol groups in a process referred to as “S-mercuration”, potentially resulting in modulation of the cellular signal transduction pathway. We examined whether low-dose MeHg could affect Akt signaling involved in cell survival. Exposure of human neuroblastoma SH-SY5Y cells of up to 2 μM MeHg phosphorylated Akt and its downstream signal molecule CREB, presumably due to inactivation of PTEN through S-mercuration. As a result, the anti-apoptotic protein Bcl-2 was up-regulated by MeHg. The activation of Akt/CREB/Bcl-2 signaling mediated by MeHg was, at least in part, linked to cellular defence because either pretreatment with wortmannin to block PI3K/Akt signaling or knockdown of Bcl-2 enhanced MeHg-mediated cytotoxicity. In contrast, increasing concentrations of MeHg disrupted Akt/CREB/Bcl-2 signaling. This phenomenon was attributed to S-mercuration of CREB through Cys286 rather than Akt. These results suggest that although MeHg is an apoptosis-inducing toxicant, this environmental electrophile is able to activate the cell survival signal transduction pathway at lower concentrations prior to apoptotic cell death. PMID:27357941

  7. β-Caryophyllene Pretreatment Alleviates Focal Cerebral Ischemia-Reperfusion Injury by Activating PI3K/Akt Signaling Pathway.

    PubMed

    Zhang, Qian; An, Ruidi; Tian, Xiaocui; Yang, Mei; Li, Minghang; Lou, Jie; Xu, Lu; Dong, Zhi

    2017-02-24

    β-Caryophyllene (BCP) has been reported to be protective against focal cerebral ischemia-reperfusion (I/R) injury by its anti-oxidative and anti-inflammatory features. Recent study demonstrates that the BCP exhibits potential neuroprotection against I/R injury induced apoptosis, however, the mechanism remains unknown. Therefore, we investigate the underlying anti-apoptotic mechanism of BCP pretreatment in I/R injury. Sprague-Dawley rats (pretreated with BCP suspensions or solvent orally for 7 days) were subjected to transient Middle Cerebral Artery Occlusion (MCAO) for 90 min, followed by 24 h reperfusion. Results showed that BCP pretreatment improved the neurologic deficit score, lowered the infarct volume and decreased number of apoptotic cells in the hippocampus. Moreover, in western blot and RT-qPCR detections, BCP pretreatment down-regulated the expressions of Bax and p53, up-regulated the expression of Bcl-2, and enhanced the phosphorylation of Akt on Ser473. Blockage of PI3K activity by wortmannin not only abolished the BCP-induced decreases in infarct volume and neurologic deficit score, but also dramatically abrogated the enhancement of AKt phosphorylation. Our results suggested that BCP pre-treatment protects against I/R injury partly by suppressing apoptosis via PI3K/AKt signaling pathway activation.

  8. Acadesine Inhibits Tissue Factor Induction and Thrombus Formation by Activating the Phosphoinositide 3-Kinase/Akt Signaling Pathway

    PubMed Central

    Zhang, Weiyu; Wang, Jianguo; Wang, Huan; Tang, Rong; Belcher, John D.; Viollet, Benoit; Geng, Jian-Guo; Zhang, Chunxiang; Wu, Chaodong; Slungaard, Arne; Zhu, Chuhong; Huo, Yuqing

    2013-01-01

    Objective Acadesine, an adenosine-regulating agent and activator of AMP-activated protein kinase, has been shown to possess antiinflammatory activity. This study investigated whether and how acadesine inhibits tissue factor (TF) expression and thrombus formation. Methods and Results Human umbilical vein endothelial cells and human peripheral blood monocytes were stimulated with lipopolysaccharide to induce TF expression. Pretreatment with acadesine dramatically suppressed the clotting activity and expression of TF (protein and mRNA). These inhibitory effects of acadesine were unchanged for endothelial cells treated with ZM241385 (a specific adenosine A2A receptor antagonist) or AMP-activated protein kinase inhibitor compound C, and in macrophages lacking adenosine A2A receptor or α1–AMP-activated protein kinase. In endothelial cells and macrophages, acadesine activated the phosphoinositide 3-kinase/Akt signaling pathway, reduced the activity of mitogen-activated protein kinases, and consequently suppressed TF expression by inhibiting the activator protein-1 and NF-κB pathways. In mice, acadesine suppressed lipopolysaccharide-mediated increases in blood coagulation, decreased TF expression in atherosclerotic lesions, and reduced deep vein thrombus formation. Conclusion Acadesine inhibits TF expression and thrombus formation by activating the phosphoinositide 3-kinase/Akt pathway. This novel finding implicates acadesine as a potentially useful treatment for many disorders associated with thrombotic pathology, such as angina pain, deep vein thrombosis, and sepsis. PMID:20185792

  9. Erythropoietin pretreatment suppresses inflammation by activating the PI3K/Akt signaling pathway in myocardial ischemia-reperfusion injury

    PubMed Central

    RONG, REN; XIJUN, XIAO

    2015-01-01

    Erythropoietin (EPO), a glycoprotein originally known for its important role in the stimulation of erythropoiesis, has recently been shown to have significant protective effects in animal models of kidney and intestinal ischemia-reperfusion injury (IRI). However, the mechanism underlying these protective effects remains unclear. The aim of the current study was to evaluate the effects of EPO on myocardial IRI and to investigate the mechanism underlying these effects. A total of 18 male Sprague Dawley rats were randomly divided into three groups, namely the sham, IRI-saline and IRI-EPO groups. Rats in the IRI-EPO group were administered 5,000 U/kg EPO intraperitoneally 24 h prior to the induction of IRI. IRI was induced by ligating the left descending coronary artery for 30 min, followed by reperfusion for 3 h. Pathological changes in the myocardial tissue were observed and scored. The levels of the proinflammatory cytokines, interleukin (IL)-6, IL-1β and tumor necrosis factor (TNF)-α, were evaluated in the serum and myocardial tissue. Furthermore, the effects of EPO on phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) signaling and EPO receptor (EPOR) phosphorylation were measured. Pathological changes in the myocardial tissue, increased expression levels of TNF-α, IL-6 and IL-1β in the myocardium, and increased serum levels of these mediators, as a result of IRI, were significantly decreased by EPO pretreatment. The effects of EPO were found to be associated with the activation of PI3K/Akt signaling, which suppressed the inflammatory responses, following the initiation of EPOR activation by EPO. Therefore, EPO pretreatment was demonstrated to decrease myocardial IRI, which was associated with activation of EPOR, subsequently increasing PI3K/Akt signaling to inhibit the production and release of inflammatory mediators. Thus, the results of the present study indicated that EPO may be useful for preventing myocardial IRI. PMID:26622330

  10. Cruciferous vegetable phytochemical sulforaphane affects phase II enzyme expression and activity in rat cardiomyocytes through modulation of Akt signaling pathway.

    PubMed

    Leoncini, Emanuela; Malaguti, Marco; Angeloni, Cristina; Motori, Elisa; Fabbri, Daniele; Hrelia, Silvana

    2011-09-01

    The isothiocyanate sulforaphane (SF), abundant in Cruciferous vegetables, is known to induce antioxidant/detoxification enzymes in many cancer cell lines, but studies focused on its cytoprotective action in nontransformed cells are just at the beginning. Since we previously demonstrated that SF elicits cardioprotection through an indirect antioxidative mechanism, the aim of this study was to analyze the signaling pathways through which SF exerts its protective effects. Using cultured rat cardiomyocytes, we investigated the ability of SF to activate Akt/protein kinase B (PKB) and extracellular signal-regulated kinase 1 and 2 (ERK1/2) signaling pathways, which are implicated in cardiac cell survival, and to increase the phosphorylation of Nuclear factor E2-related factor 2 (Nrf2) and its binding to the antioxidant response element. By means of specific inhibitors, we demonstrated that the Phosphatidylinositol 3-kinase (PI3K)/Akt pathway represents a mechanism through which SF influences both expression and activity of glutathione reductase, glutathione-S-transferase, thioredoxin reductase, and NAD(P)H:quinone oxidoreductase-1, analyzed by western immunoblotting and spectrophotometric assay, respectively, and modulates Nrf2 binding and phosphorylation resulting in a cytoprotective action against oxidative damage. Results of this study confirm the importance of phase II enzymes modulation as cytoprotective mechanism and support the nutritional assumption of Cruciferous vegetables as source of nutraceutical cardioprotective agents.

  11. The mitochondrial-derived peptide humanin activates the ERK1/2, AKT, and STAT3 signaling pathways and has age-dependent signaling differences in the hippocampus.

    PubMed

    Kim, Su-Jeong; Guerrero, Noel; Wassef, Gabriella; Xiao, Jialin; Mehta, Hemal H; Cohen, Pinchas; Yen, Kelvin

    2016-07-26

    Humanin is a small secreted peptide that is encoded in the mitochondrial genome. Humanin and its analogues have a protective role in multiple age-related diseases including type 2 diabetes and Alzheimer's disease, through cytoprotective and neuroprotective effects both in vitro and in vivo. However, the humanin-mediated signaling pathways are not well understood. In this paper, we demonstrate that humanin acts through the GP130/IL6ST receptor complex to activate AKT, ERK1/2, and STAT3 signaling pathways. Humanin treatment increases phosphorylation in AKT, ERK 1/2, and STAT3 where PI3K, MEK, and JAK are involved in the activation of those three signaling pathways, respectively. Furthermore, old mice, but not young mice, injected with humanin showed an increase in phosphorylation in AKT and ERK1/2 in the hippocampus. These findings uncover a key signaling pathway of humanin that is important for humanin's function and also demonstrates an age-specific in vivo effect in a region of the brain that is critical for memory formation in an age-dependent manner.

  12. The mitochondrial-derived peptide humanin activates the ERK1/2, AKT, and STAT3 signaling pathways and has age-dependent signaling differences in the hippocampus

    PubMed Central

    Kim, Su-Jeong; Guerrero, Noel; Wassef, Gabriella; Xiao, Jialin; Mehta, Hemal H.; Cohen, Pinchas; Yen, Kelvin

    2016-01-01

    Humanin is a small secreted peptide that is encoded in the mitochondrial genome. Humanin and its analogues have a protective role in multiple age-related diseases including type 2 diabetes and Alzheimer's disease, through cytoprotective and neuroprotective effects both in vitro and in vivo. However, the humanin-mediated signaling pathways are not well understood. In this paper, we demonstrate that humanin acts through the GP130/IL6ST receptor complex to activate AKT, ERK1/2, and STAT3 signaling pathways. Humanin treatment increases phosphorylation in AKT, ERK 1/2, and STAT3 where PI3K, MEK, and JAK are involved in the activation of those three signaling pathways, respectively. Furthermore, old mice, but not young mice, injected with humanin showed an increase in phosphorylation in AKT and ERK1/2 in the hippocampus. These findings uncover a key signaling pathway of humanin that is important for humanin's function and also demonstrates an age-specific in vivo effect in a region of the brain that is critical for memory formation in an age-dependent manner. PMID:27384491

  13. {delta}-Opioid receptor-stimulated Akt signaling in neuroblastoma x glioma (NG108-15) hybrid cells involves receptor tyrosine kinase-mediated PI3K activation

    SciTech Connect

    Heiss, Anika; Ammer, Hermann; Eisinger, Daniela A.

    2009-07-15

    {delta}-Opioid receptor (DOR) agonists possess cytoprotective properties, an effect associated with activation of the 'pro-survival' kinase Akt. Here we delineate the signal transduction pathway by which opioids induce Akt activation in neuroblastoma x glioma (NG108-15) hybrid cells. Exposure of the cells to both [D-Pen{sup 2,5}]enkephalin and etorphine resulted in a time- and dose-dependent increase in Akt activity, as measured by means of an activation-specific antibody recognizing phosphoserine-473. DOR-mediated Akt signaling is blocked by the opioid antagonist naloxone and involves inhibitory G{sub i/o} proteins, because pre-treatment with pertussis toxin, but not over-expression of the G{sub q/11} scavengers EBP50 and GRK2-K220R, prevented this effect. Further studies with Wortmannin and LY294002 revealed that phophoinositol-3-kinase (PI3K) plays a central role in opioid-induced Akt activation. Opioids stimulate Akt activity through transactivation of receptor tyrosine kinases (RTK), because pre-treatment of the cells with inhibitors for neurotrophin receptor tyrosine kinases (AG879) and the insulin-like growth factor receptor IGF-1 (AG1024), but not over-expression of the G{beta}{gamma} scavenger phosducin, abolished this effect. Activated Akt translocates to the nuclear membrane, where it promotes GSK3 phosphorylation and prevents caspase-3 cleavage, two key events mediating inhibition of cell apoptosis and enhancement of cell survival. Taken together, these results demonstrate that in NG108-15 hybrid cells DOR agonists possess cytoprotective properties mediated by activation of the RTK/PI3K/Akt signaling pathway.

  14. Expression of constitutively active Akt/protein kinase B signals GLUT4 translocation in the absence of an intact actin cytoskeleton.

    PubMed

    Eyster, Craig A; Duggins, Quwanza S; Olson, Ann Louise

    2005-05-06

    The actin cytoskeleton has been shown to be required for insulin-dependent GLUT4 translocation; however, the role that the actin network plays is unknown. Actin may play a role in formation of an active signaling complex, or actin may be required for movement of vesicles to the plasma membrane surface. To distinguish between these possibilities, we examined the ability of myr-Akt, a constitutively active form of Akt that signals GLUT4 translocation to the plasma membrane in the absence of insulin, to signal translocation of an HA-GLUT4-GFP reporter protein in the presence or absence of an intact cytoskeleton in 3T3-L1 adipocytes. Expression of myr-Akt signaled the redistribution of the GLUT4 reporter protein to the cell surface in the absence or presence of 10 microm latrunculin B, a concentration sufficient to completely inhibit insulin-dependent redistribution of the GLUT4 reporter to the cell surface. These data suggest that the actin network plays a primary role in organization of the insulin-signaling complex. To further support this conclusion, we measured the activation of known signaling proteins using a saturating concentration of insulin in cells pretreated without or with 10 microm latrunculin B. We found that latrunculin treatment did not affect insulin-dependent tyrosine phosphorylation of the insulin receptor beta-subunit and IRS-1 but completely inhibited activation of Akt/PKB enzymatic activity. Phosphorylation of Akt/PKB at Ser-473 and Thr-308 was inhibited by latrunculin B treatment, indicating that the defect in signaling lies prior to Akt/PKB activation. In summary, our data support the hypothesis that the actin network plays a role in organization of the insulin-signaling complex but is not required for vesicle trafficking and/or fusion.

  15. Regulator of G protein signaling 1 suppresses CXCL12-mediated migration and AKT activation in RPMI 8226 human plasmacytoma cells and plasmablasts.

    PubMed

    Pak, Hyo-Kyung; Gil, Minchan; Lee, Yoonkyung; Lee, Hyunji; Lee, A-Neum; Roh, Jin; Park, Chan-Sik

    2015-01-01

    Migration of plasma cells to the bone marrow is critical factor to humoral immunity and controlled by chemokines. Regulator of G protein signaling 1 (RGS1) is a GTPase-activating protein that controls various crucial functions such as migration. Here, we show that RGS1 controls the chemotactic migration of RPMI 8226 human plasmacytoma cells and human plasmablasts. LPS strongly increased RGS1 expression and retarded the migration of RPMI 8226 cells by suppressing CXCL12-mediated AKT activation. RGS1 knockdown by siRNA abolished the retardation of migration and AKT suppression by LPS. RGS1-dependent regulation of migration via AKT is also observed in cultured plasmablasts. We propose novel functions of RGS1 that suppress AKT activation and the migration of RPMI 8226 cells and plasmablasts in CXCL12-mediated chemotaxis.

  16. Isorhamnetin inhibits cell proliferation and induces apoptosis in breast cancer via Akt and mitogen-activated protein kinase kinase signaling pathways

    PubMed Central

    HU, SHAN; HUANG, LIMING; MENG, LIWEI; SUN, HE; ZHANG, WEI; XU, YINGCHUN

    2015-01-01

    Breast cancer is the most common cause of female cancer-associated mortality. Although treatment options, including chemotherapy, radiotherapy and surgery have led to a decline in the mortality rates associated with breast cancer, drug resistance remains one of the predominant causes for poor prognosis and high recurrence rates. The present study investigated the potential effects of the natural product, isorhamnetin on breast cancer, and examined the effects of isorhamnetin on the Akt/mammalian target of rapamycin (mTOR) and the mitogen-activated protein kinase (MAPK)/MAPK kinase (MEK) signaling cascades, which are two important signaling pathways for endocrine therapy resistance in breast cancer. The results of the present study indicate that isorhamnetin inhibits cell proliferation and induces cell apoptosis. In addition, isorhamnetin was observed to inhibit the Akt/mTOR and the MEK/extracellular signal-regulated kinase phosphorylation cascades. The inhibition of these two signaling pathways was attenuated by the two Akt and MEK1 inhibitors, but not by the nuclear factor-κB inhibitor. Furthermore, epidermal growth factor inhibited the effects of isorhamnetin via activation of the Akt and MEK signaling pathways. These results indicate that isorhamnetin exhibits antitumor effects in breast cancer, which are mediated by the Akt and MEK signaling pathways. PMID:26502751

  17. Isorhamnetin inhibits cell proliferation and induces apoptosis in breast cancer via Akt and mitogen‑activated protein kinase kinase signaling pathways.

    PubMed

    Hu, Shan; Huang, Liming; Meng, Liwei; Sun, He; Zhang, Wei; Xu, Yingchun

    2015-11-01

    Breast cancer is the most common cause of female cancer-associated mortality. Although treatment options, including chemotherapy, radiotherapy and surgery have led to a decline in the mortality rates associated with breast cancer, drug resistance remains one of the predominant causes for poor prognosis and high recurrence rates. The present study investigated the potential effects of the natural product, isorhamnetin on breast cancer, and examined the effects of isorhamnetin on the Akt/mammalian target of rapamycin (mTOR) and the mitogen-activated protein kinase (MAPK)/MAPK kinase (MEK) signaling cascades, which are two important signaling pathways for endocrine therapy resistance in breast cancer. The results of the present study indicate that isorhamnetin inhibits cell proliferation and induces cell apoptosis. In addition, isorhamnetin was observed to inhibit the Akt/mTOR and the MEK/extracellular signal-regulated kinase phosphorylation cascades. The inhibition of these two signaling pathways was attenuated by the two Akt and MEK1 inhibitors, but not by the nuclear factor-κB inhibitor. Furthermore, epidermal growth factor inhibited the effects of isorhamnetin via activation of the Akt and MEK signaling pathways. These results indicate that isorhamnetin exhibits antitumor effects in breast cancer, which are mediated by the Akt and MEK signaling pathways.

  18. AKT/GSK3 signaling pathway and schizophrenia

    PubMed Central

    Emamian, Effat S.

    2012-01-01

    transcription factors related to schizophrenia will be reviewed to see how hypo-activity of AKT signaling pathway may impact such transcriptional mechanisms. PMID:22435049

  19. Betulinic acid protects against cerebral ischemia/reperfusion injury by activating the PI3K/Akt signaling pathway.

    PubMed

    Jiao, Shujie; Zhu, Hongcan; He, Ping; Teng, Junfang

    2016-12-01

    Betulinic acid (BA), a naturally occurring pentacyclic lupane group triterpenoid, has been demonstrated to protect against ischemia/reperfusion-induced renal damage. However, the effects of BA on cerebral ischemia/reperfusion (I/R) injury remain unclear. Hence, this study was to investigate the effects of BA on oxygen and glucose deprivation/reperfusion (OGD/R) induced neuronal injury in rat hippocampal neurons. Our results showed that BA pretreatment greatly attenuated OGD/R-induced neuronal injury. BA also inhibited OGD/R-induced intracellular ROS production and MDA level in rat hippocampal neurons. Furthermore, the down-regulation of Bcl-2, up-regulation of Bax and the consequent activation of caspase-3 induced by OGD/R were reversed by BA pretreatment. Mechanistic studies demonstrated that BA pretreatment up-regulated the expression levels of p-PI3K and p-Akt in hippocampal neurons induced by OGD/R. Taken together, these data suggested that BA inhibits OGD/R-induced neuronal injury in rat hippocampal neurons through the activation of PI3K/Akt signaling pathway.

  20. A novel 7-bromoindirubin with potent anticancer activity suppresses survival of human melanoma cells associated with inhibition of STAT3 and Akt signaling.

    PubMed

    Liu, Lucy; Kritsanida, Marina; Magiatis, Prokopios; Gaboriaud, Nicolas; Wang, Yan; Wu, Jun; Buettner, Ralf; Yang, Fan; Nam, Sangkil; Skaltsounis, Leandros; Jove, Richard

    2012-11-01

    STAT3 and Akt signaling have been validated as potential molecular targets for treatment of cancers including melanoma. These small molecule inhibitors of STAT3 or Akt signaling are promising for developing anti-melanoma therapeutic agents. MLS-2438, a novel 7-bromoindirubin, a derivative of the natural product indirubin, was synthesized with a bromo-group at the 7-position on one indole ring and a hydrophilic group at the 3'-position on the other indole ring. We tested the anticancer activity of MLS-2438 and investigated its mechanism of action in human melanoma cell lines. Here, we show that MLS-2438 inhibits viability and induces apoptosis of human melanoma cells associated with inhibition of STAT3 and Akt signaling. Several pro-apoptotic Bcl-2 family proteins are involved in the MLS-2438 mediated apoptosis. MLS-2438 inhibits Src kinase activity in vitro and phosphorylation of JAK2, Src, STAT3 and Akt in cultured cancer cells. In contrast to the decreased phosphorylation levels of JAK2, Src, STAT3 and Akt, phosphorylation levels of the MAPK (Erk1/2) signaling protein were not reduced in cells treated with MLS-2438. These results demonstrate that MLS-2438, a novel natural product derivative, is a Src inhibitor and potentially regulates kinase activity of JAK2 and Akt in cancer cells. Importantly, MLS-2438 suppressed tumor growth with low toxicity in a mouse xenograft model of human melanoma. Our findings support further development of MLS-2438 as a potential small-molecule therapeutic agent that targets both STAT3 and Akt signaling in human melanoma cells.

  1. Dodeca-2(E),4(E)-dienoic acid isobutylamide enhances glucose uptake in 3T3-L1 cells via activation of Akt signaling.

    PubMed

    Choi, Kyeong-Mi; Kim, Wonkyun; Hong, Jin Tae; Yoo, Hwan-Soo

    2017-02-01

    Dodeca-2(E),4(E)-dienoic acid isobutylamide (DDI), an alkamide derived from the plant Echinacea purpurea, promotes adipocyte differentiation and activates peroxisome proliferator-activated receptor γ, which is associated with enhanced insulin sensitivity. In the present study, we investigated whether DDI may increase glucose uptake through activation of the insulin signaling pathway in 3T3-L1 adipocytes. DDI increased insulin-stimulated glucose uptake, and expression and translocation of glucose transporter 4 in adipocytes treated with sub-optimal levels of insulin. Additionally, DDI enhanced Akt phosphorylation, whereas phosphoinositide 3-kinase/Akt inhibitors suppressed DDI-induced glucose uptake. These results suggest that DDI may improve insulin sensitivity through the activation of Akt signaling, which leads to enhanced glucose uptake.

  2. Metastasis and AKT activation.

    PubMed

    Sheng, Shijie; Qiao, Meng; Pardee, Arthur B

    2009-03-01

    Metastasis, responsible for 90% of cancer patient deaths, is an inefficient process because many tumor cells die. The survival of metastatic tumor cells should be considered as a critical therapeutic target. This review provides a new perspective regarding the role of AKT in tumor survival, and the rationale to target AKT in anti-metastasis therapies.

  3. Liraglutide reduces lipogenetic signals in visceral adipose of db/db mice with AMPK activation and Akt suppression.

    PubMed

    Shao, Yimin; Yuan, Geheng; Zhang, Junqing; Guo, Xiaohui

    2015-01-01

    Liraglutide, a glucagon-like peptide-1 analog, has been proved to reduce body weight and visceral adipose tissue (VAT) in human studies. In this study, we aimed at examining lipogenetic signal changes in VAT after weight-loss with liraglutide in db/db mice. The mice were divided into two groups: liraglutide-treated group (n=14, 8-week-old, fasting glucose. >10 mmol/L, liraglutide 300 μg/kg twice a day for 4 weeks) and control group (n=14, saline). We found body weight gain and food intake were reduced after liraglutide treatment (P<0.05). Compared to the control group, the VAT weights were significantly lower in the treated group (2.32±0.37 g versus 3.20±0.30 g, P<0.01) than that in control group. In VAT, compared with control group, the lipogenetic transcription factors PPARγ and C/EBPα expressions were both reduced with pAMPK and pACC increased 3.5-fold and 2.31-fold respectively, while pAkt and pP38MAPK were reduced 0.38-fold and 0.62-fold respectively (P<0.01). In conclusion, VAT was reduced after weight loss with AMPK activation and Akt suppression with liraglutide treatment, which was associated with reduction of lipogenetic process in VAT.

  4. IL-8 induces the epithelial-mesenchymal transition of renal cell carcinoma cells through the activation of AKT signaling

    PubMed Central

    Zhou, Nan; Lu, Fuding; Liu, Cheng; Xu, Kewei; Huang, Jian; Yu, Dexin; Bi, Liangkuan

    2016-01-01

    The epithelial-mesenchymal transition (EMT) process has increasingly been examined due to its role in the progression of human tumors. Renal cell carcinoma (RCC) is one of the most common urological tumors that results in patient mortality. Previous studies have demonstrated that the EMT process is closely associated with the metastasis of RCC; however, the underlying molecular mechanism has not been determined yet. The present study revealed that interleukin (IL)-8 was highly expressed in metastatic RCC. IL-8 could induce the EMT of an RCC cell line by enhancing N-cadherin expression and decreasing E-cadherin expression. Furthermore, IL-8 could induce AKT phosphorylation, and the phosphatidylinositol-4,5-bisphosphate 3-kinase inhibitor LY294002 could inhibit the EMT of RCC cells that was induced by IL-8. Therefore, these results suggest that IL-8 is able to promote the EMT of RCC through the activation of the AKT signal transduction pathway, and this may provide a possible molecular mechanism for RCC metastasis. PMID:27588140

  5. Bis(hinokitiolato)zinc complex ([Zn(hkt)2]) activates Akt/protein kinase B independent of insulin signal transduction.

    PubMed

    Naito, Yuki; Yoshikawa, Yutaka; Masuda, Kazufumi; Yasui, Hiroyuki

    2016-07-01

    Since many Zn complexes have been developed to enhance the insulin-like activity and increase the exposure and residence of Zn in the animal body, these complexes are recognized as one of the new candidates with action mechanism different from existing anti-diabetic drugs. However, the molecular mechanism by which Zn complexes exert an anti-DM effect is unknown. Therefore, we evaluated the activity of Zn complexes, especially related to the phosphorylation of insulin signaling pathway components. We focused on the insulin-like effects of the bis(hinokitiolato)zinc complex, [Zn(hkt)2], using 3T3-L1 adipocytes. [Zn(hkt)2] was taken up by cells and induced Akt phosphorylation in a time-dependent manner. Additionally, it showed inhibitory activity against PTP1B and PTEN, which are major negative regulators of insulin signaling. It did not promote the phosphorylation of IR (insulin receptor)-β or IRS (insulin receptor substrate)-1 by itself, but in combination with insulin, it enhanced the phosphorylation of IRβ. We conclude that [Zn(hkt)2] has effects on the proteins of insulin signaling pathway without insulin receptor mediation, and [Zn(hkt)2] promotes insulin function and shows the anti-DM effects. Thus, [Zn(hkt)2] may be the basis for improved DM treatments.

  6. Akt phosphorylates and regulates the osteogenic activity of Osterix.

    PubMed

    Choi, You Hee; Jeong, Hyung Min; Jin, Yun-Hye; Li, Hongyan; Yeo, Chang-Yeol; Lee, Kwang-Youl

    2011-08-05

    Osterix (Osx), a zinc-finger transcription factor is required for osteoblast differentiation and new bone formation during embryonic development. Akt is a member of the serine/threonine-specific protein kinase and plays important roles in osteoblast differentiation. The function of Osterix can be also modulated by post-translational modification. But, the precise molecular signaling mechanisms between Osterix and Akt are not known. In this study, we investigated the potential regulation of Osterix function by Akt in osteoblast differentiation. We found that Akt phosphorylates Osterix and that Akt activation increases protein stability, osteogenic activity and transcriptional activity of Osterix. We also found that BMP-2 increases the protein level of Osterix in an Akt activity-dependent manner. These results suggest that Akt activity enhances the osteogenic function of Osterix, at least in part, through protein stabilization and that BMP-2 regulates the osteogenic function of Osterix, at least in part, through Akt.

  7. Statins upregulate cystathionine γ-lyase transcription and H2S generation via activating Akt signaling in macrophage.

    PubMed

    Xu, Yuan; Du, Hua-Ping; Li, Jiaojiao; Xu, Ran; Wang, Ya-Li; You, Shou-Jiang; Liu, Huihui; Wang, Fen; Cao, Yong-Jun; Liu, Chun-Feng; Hu, Li-Fang

    2014-09-01

    Hydrogen sulfide (H2S), the third gaseous transmitter, is implicated in various pathophysiologic processes. In the cardiovascular system, H2S exerts effects of cardioprotection, vascular tone regulation, and atherogenesis inhibition. Recent studies demonstrated that atorvastatin, the inhibitor of 3-hydroxyl-3-methyl coenzyme A reductase, affected H2S formation in kidney and other organs. However, the underlying mechanisms are not fully understood. In this study, we examined the effects of three different statins (fluvastatin, atorvastatin and pravastatin) on H2S formation in raw264.7 macrophages. There was a remarkable rise in H2S level in fluvastatin- and atorvastatin-stimulated macrophages, while pravastatin failed to show any significant effect on it. Moreover, fluvastatin and atorvastatin enhanced the mRNA and protein expression of cystathionine γ-lyase (CSE) in dose- and time-dependent manners. Fluvastatin also markedly enhanced the CSE activity. However, fluvastatin did not alter the mRNA or protein expression of another H2S-producing enzyme 3-mercaptopyruvate sulfurtransferase. Blockade of CSE with its inhibitor dl-propargylglycine (PAG) or siRNA markedly reduced the H2S level in fluvastatin-stimulated macrophages. In addition, fluvastatin elevated Akt phosphorylation, which occurred as early as 15 min after treatment, peaked at 1h, and lasted at least 3h. Both PI3K inhibitor LY294002 (10 μM) and Akt inhibitor perifosine (10μM) were able to reverse the increases of CSE mRNA and H2S production in fluvastatin-stimulated macrophages. Last, we showed that fluvastatin reduced the mRNA levels of pro-inflammatory molecules such as IL-1β and MCP-1 in LPS-treated macrophages, which were completely reversed by CSE inhibitor PAG. Taken together, the findings demonstrate that statins may up-regulate CSE expression/activity and subsequently elevate H2S generation by activating Akt signaling pathway and also imply that CSE-H2S pathway plays a critical role in the anti

  8. Sericin can reduce hippocampal neuronal apoptosis by activating the Akt signal transduction pathway in a rat model of diabetes mellitus☆

    PubMed Central

    Chen, Zhihong; He, Yaqiang; Song, Chengjun; Dong, Zhijun; Su, Zhejun; Xue, Jingfeng

    2012-01-01

    In the present study, a rat model of type 2 diabetes mellitus was established by continuous peritoneal injection of streptozotocin. Following intragastric perfusion of sericin for 35 days, blood glucose levels significantly reduced, neuronal apoptosis in the hippocampal CA1 region decreased, hippocampal phosphorylated Akt and nuclear factor kappa B expression were enhanced, but Bcl-xL/Bcl-2 associated death promoter expression decreased. Results demonstrated that sericin can reduce hippocampal neuronal apoptosis in a rat model of diabetes mellitus by regulating abnormal changes in the Akt signal transduction pathway. PMID:25767499

  9. Icariin stimulates angiogenesis by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways in human endothelial cells

    SciTech Connect

    Chung, Byung-Hee; Kim, Jong-Dai; Kim, Chun-Ki; Kim, Jung Huan; Won, Moo-Ho; Lee, Han-Soo; Dong, Mi-Sook; Ha, Kwon-Soo; Kwon, Young-Geun; Kim, Young-Myeong

    2008-11-14

    We investigated the molecular effect and signal pathway of icariin, a major flavonoid of Epimedium koreanum Nakai, on angiogenesis. Icariin stimulated in vitro endothelial cell proliferation, migration, and tubulogenesis, which are typical phenomena of angiogenesis, as well as increased in vivo angiogenesis. Icariin activated the angiogenic signal modulators, ERK, phosphatidylinositol 3-kinase (PI3K), Akt, and endothelial nitric oxide synthase (eNOS), and increased NO production, without affecting VEGF expression, indicating that icariin may directly stimulate angiogenesis. Icariin-induced ERK activation and angiogenic events were significantly inhibited by the MEK inhibitor PD98059, without affecting Akt and eNOS phosphorylation. The PI3K inhibitor Wortmannin suppressed icariin-mediated angiogenesis and Akt and eNOS activation without affecting ERK phosphorylation. Moreover, the NOS inhibitor NMA partially reduced the angiogenic activity of icariin. These results suggest that icariin stimulated angiogenesis by activating the MEK/ERK- and PI3K/Akt/eNOS-dependent signal pathways and may be a useful drug for angiogenic therapy.

  10. Mefloquine exerts anticancer activity in prostate cancer cells via ROS-mediated modulation of Akt, ERK, JNK and AMPK signaling

    PubMed Central

    YAN, KUN-HUANG; YAO, CHIH-JUNG; HSIAO, CHI-HAO; LIN, KE-HSUN; LIN, YUNG-WEI; WEN, YU-CHING; LIU, CHUNG-CHI; YAN, MING-DE; CHUANG, SHUANG-EN; LAI, GI-MING; LEE, LIANG-MING

    2013-01-01

    Mefloquine (MQ) is a prophylactic anti-malarial drug. Previous studies have shown that MQ induces oxidative stress in vitro. Evidence indicates that reactive oxygen species (ROS) may be used as a therapeutic modality to kill cancer cells. This study investigated whether MQ also inhibits prostate cancer (PCa) cell growth. We used sulforhodamine B (SRB) staining to determine cell viability. MQ has a highly selective cytotoxicity that inhibits PCa cell growth. The antitumor effect was most significant when examined using a colony formation assay. MQ also induces hyperpolarization of the mitochondrial membrane potential (MMP), as well as ROS generation. The blockade of MQ-induced anticancer effects by N-acetyl cysteine (NAC) pre-treatment confirmed the role of ROS. This indicates that the MQ-induced anticancer effects are caused primarily by increased ROS generation. Moreover, we observed that MQ-mediated ROS simultaneously downregulated Akt phosphorylation and activated extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and adenosine monophosphate-activated protein kinase (AMPK) signaling in PC3 cells. These findings provide insights for further anticancer therapeutic options. PMID:23760395

  11. Calcium oxalate crystals induces tight junction disruption in distal renal tubular epithelial cells by activating ROS/Akt/p38 MAPK signaling pathway.

    PubMed

    Yu, Lei; Gan, Xiuguo; Liu, Xukun; An, Ruihua

    2017-11-01

    Tight junction plays important roles in regulating paracellular transports and maintaining cell polarity. Calcium oxalate monohydrate (COM) crystals, the major crystalline composition of kidney stones, have been demonstrated to be able to cause tight junction disruption to accelerate renal cell injury. However, the cellular signaling involved in COM crystal-induced tight junction disruption remains largely to be investigated. In the present study, we proved that COM crystals induced tight junction disruption by activating ROS/Akt/p38 MAPK pathway. Treating Madin-Darby canine kidney (MDCK) cells with COM crystals induced a substantial increasing of ROS generation and activation of Akt that triggered subsequential activation of ASK1 and p38 mitogen-activated protein kinase (MAPK). Western blot revealed a significantly decreased expression of ZO-1 and occludin, two important structural proteins of tight junction. Besides, redistribution and dissociation of ZO-1 were observed by COM crystals treatment. Inhibition of ROS by N-acetyl-l-cysteine (NAC) attenuated the activation of Akt, ASK1, p38 MAPK, and down-regulation of ZO-1 and occludin. The redistribution and dissociation of ZO-1 were also alleviated by NAC treatment. These results indicated that ROS were involved in the regulation of tight junction disruption induced by COM crystals. In addition, the down-regulation of ZO-1 and occludin, the phosphorylation of ASK1 and p38 MAPK were also attenuated by MK-2206, an inhibitor of Akt kinase, implying Akt was involved in the disruption of tight junction upstream of p38 MAPK. Thus, these results suggested that ROS-Akt-p38 MAPK signaling pathway was activated in COM crystal-induced disruption of tight junction in MDCK cells.

  12. Angiotensin-converting enzyme 2/angiotensin-(1–7)/Mas axis activates Akt signaling to ameliorate hepatic steatosis

    PubMed Central

    Cao, Xi; Yang, Fangyuan; Shi, Tingting; Yuan, Mingxia; Xin, Zhong; Xie, Rongrong; Li, Sen; Li, Hongbing; Yang, Jin-Kui

    2016-01-01

    The classical axis of renin-angiotensin system (RAS), angiotensin (Ang)-converting enzyme (ACE)/Ang II/AT1, contributes to the development of non-alcoholic fatty liver disease (NAFLD). However, the role of bypass axis of RAS (Angiotensin-converting enzyme 2 (ACE2)/Ang-(1–7)/Mas) in hepatic steatosis is still unclear. Here we showed that deletion of ACE2 aggravates liver steatosis, which is correlated with the increased expression of hepatic lipogenic genes and the decreased expression of fatty acid oxidation-related genes in the liver of ACE2 knockout (ACE2−/y) mice. Meanwhile, oxidative stress and inflammation were also aggravated in ACE2−/y mice. On the contrary, overexpression of ACE2 improved fatty liver in db/db mice, and the mRNA levels of fatty acid oxidation-related genes were up-regulated. In vitro, Ang-(1–7)/ACE2 ameliorated hepatic steatosis, oxidative stress and inflammation in free fatty acid (FFA)-induced HepG2 cells, and what’s more, Akt inhibitors reduced ACE2-mediated lipid metabolism. Furthermore, ACE2-mediated Akt activation could be attenuated by blockade of ATP/P2 receptor/Calmodulin (CaM) pathway. These results indicated that Ang-(1–7)/ACE2/Mas axis may reduce liver lipid accumulation partly by regulating lipid-metabolizing genes through ATP/P2 receptor/CaM signaling pathway. Our findings support the potential role of ACE2/Ang-(1–7)/Mas axis in prevention and treatment of hepatic lipid metabolism. PMID:26883384

  13. Phosphatidylserine is a critical modulator for Akt activation

    PubMed Central

    Huang, Bill X.; Akbar, Mohammed; Kevala, Karl

    2011-01-01

    Akt activation relies on the binding of Akt to phosphatidylinositol-3,4,5-trisphosphate (PIP3) in the membrane. Here, we demonstrate that Akt activation requires not only PIP3 but also membrane phosphatidylserine (PS). The extent of insulin-like growth factor–induced Akt activation and downstream signaling as well as cell survival under serum starvation conditions positively correlates with plasma membrane PS levels in living cells. PS promotes Akt-PIP3 binding, participates in PIP3-induced Akt interdomain conformational changes for T308 phosphorylation, and causes an open conformation that allows for S473 phosphorylation by mTORC2. PS interacts with specific residues in the pleckstrin homology (PH) and regulatory (RD) domains of Akt. Disruption of PS–Akt interaction by mutation impairs Akt signaling and increases susceptibility to cell death. These data identify a critical function of PS for Akt activation and cell survival, particularly in conditions with limited PIP3 availability. The novel molecular interaction mechanism for Akt activation suggests potential new targets for controlling Akt-dependent cell survival and proliferation. PMID:21402788

  14. Isoflurane Promotes Non-Small Cell Lung Cancer Malignancy by Activating the Akt-Mammalian Target of Rapamycin (mTOR) Signaling Pathway

    PubMed Central

    Zhang, Wenhua; Shao, Xueqian

    2016-01-01

    Background Lung cancer is one of the leading causes of cancer mortalities worldwide, and non-small cell lung cancer (NSCLC) accounts for the majority of all lung cancer cases. Surgery remains one of the front-line treatment options for NSCLC, but events within the perioperative period were found to affect cancer prognosis, such as anesthesia procedures. Isoflurane, a commonly used volatile anesthetic, enhances the malignant potential of renal, prostate, and ovarian cancer cells, but its effects on NSCLC development have not been previously reported. Material/Methods CCK-8 and MTT cell proliferation assays were used to analyze NSCLC cell proliferation. Metastatic ability was examined by wound healing and transwell assays. We used Western blot analysis to study the mechanism of effect of Isoflurane in NSCLC development. Results We demonstrated that isoflurane promotes proliferation, migration and invasiveness of NSCLC cells, as well as upregulation of the Akt-mTOR signaling pathway in NSCLC cells. Pharmacological inhibition of Akt-mTOR signaling abolished the ability of isoflurane to promote proliferation, migration, and invasion of NSCLC cells, indicating that isoflurane promotes NSCLC cell malignancy by activating the Akt-mTOR signaling pathway. Conclusions Isoflurane promotes NSCLC proliferation, migration and invasion by activating the Akt-mTOR signaling pathway. PMID:27897153

  15. Antiapoptotic effects of erythropoietin in differentiated neuroblastoma SH-SY5Y cells require activation of both the STAT5 and AKT signaling pathways.

    PubMed

    Um, Moonkyoung; Lodish, Harvey F

    2006-03-03

    The hematopoietic cytokine erythropoietin (Epo) prevents neuronal death during ischemic events in the brain and in neurodegenerative diseases, presumably through its antiapoptotic effects. To explore the role of different signaling pathways in Epo-mediated antiapoptotic effects in differentiated human neuroblastoma SH-SY5Y cells, we employed a prolactin receptor (PrlR)/erythropoietin receptor (EpoR) chimera system, in which binding of prolactin (Prl) to the extracellular domain activates EpoR signaling in the cytosol. On induction of apoptosis by staurosporine, Prl supports survival of the SH-SY5Y cells expressing the wild-type PrlR/EpoR chimera. In these cells Prl treatment strongly activates the STAT5, AKT, and MAPK signaling pathways and induces weak activation of the p65 NF-kappaB factor. Selective mutation of the eight tyrosine residues of the EpoR cytoplasmic domain results in impaired or absent activation of either STAT5 (mutation of Tyr(343)) or AKT (mutation of Tyr(479)) or both (mutation of all eight tyrosine residues). Most interestingly, Prl treatment does not prevent apoptosis in cells expressing mutant PrlR/EpoR chimeras in which either the STAT5 or the AKT signaling pathways are not activated. In contrast, ERK 1/2 is fully activated by all mutant PrlR/EpoR chimeras, comparable with the level seen with the wild-type PrlR/EpoR chimera, implying that activation of the MAPK signaling pathway per se is not sufficient for antiapoptotic activity. Therefore, the antiapoptotic effects of Epo in neuronal cells require the combinatorial activation of multiple signaling pathways, including STAT5, AKT, and potentially MAPK as well, in a manner similar to that observed in hematopoietic cells.

  16. TC21 mediates transformation and cell survival via activation of phosphatidylinositol 3-kinase/Akt and NF-kappaB signaling pathway.

    PubMed

    Rong, Rong; He, Qin; Liu, Yusen; Sheikh, M Saeed; Huang, Ying

    2002-02-07

    The signaling pathways of TC21-mediated transformation and cell survival are not well-established. In this study, we have investigated the role of PI3-K/Akt signaling pathway in oncogenic-TC21-mediated transformation and cell survival. We found that oncogenic-TC21 stimulated the PI3-K activity. This was associated with the activation of Akt, a key component of PI3-K signaling pathway. We also found that TC21 interacted and formed complex with PI3-K. Mutations in the GTP-binding region of TC21, which enhanced GTP-binding potential of this protein, also stimulated its association with PI3-K, suggesting that PI3-K may preferentially interact with the GTP-bound form. Suppression of PI3-K and Akt by specific inhibitors LY294002 and Wortmannin reversed TC21-induced transformation. Likewise, inhibition of PI3-K activity by the PI3-K phosphotase PTEN reduced TC21-mediated focus formation in NIH3T3 cells. Investigation of TC21's effect on cell survival revealed that mutant-TC21 expressing cells were more resistant to etoposide- and cisplatin-induced cell death, and this was associated with the activation of anti-apoptotic protein NF-kappaB, a downstream target of Akt. Treatment of PI3-K inhibitor LY294002 significantly suppressed TC21-mediated NF-kappaB activation. In conclusion, we have identified PI3-K as an effector of TC21 and demonstrated that the PI3-K/Akt signaling pathway plays important roles in TC21-mediated transformation and cell survival.

  17. Galangin Activates the ERK/AKT-Driven Nrf2 Signaling Pathway to Increase the Level of Reduced Glutathione in Human Keratinocytes.

    PubMed

    Madduma Hewage, Susara Ruwan Kumara; Piao, Mei Jing; Kang, Kyoung Ah; Ryu, Yea Seong; Fernando, Pattage Madushan Dilhara Jayatissa; Oh, Min Chang; Park, Jeong Eon; Shilnikova, Kristina; Moon, Yu Jin; Shin, Dae O; Hyun, Jin Won

    2016-11-08

    Previously, we demonstrated that galangin (3,5,7-trihydroxyflavone) protects human keratinocytes against ultraviolet B (UVB)-induced oxidative damage. In this study, we investigated the effect of galangin on induction of antioxidant enzymes involved in synthesis of reduced glutathione (GSH), and investigated the associated upstream signaling cascades. By activating nuclear factor-erythroid 2-related factor (Nrf2), galangin treatment significantly increased expression of glutamate-cysteine ligase catalytic subunit (GCLC) and glutathione synthetase (GSS). This activation of Nrf2 depended on extracellular signal-regulated kinases (ERKs) and protein kinase B (AKT) signaling. Inhibition of GSH in galangin-treated cells attenuated the protective effect of galangin against the deleterious effects of UVB. Our results reveal that galangin protects human keratinocytes by activating ERK/AKT-Nrf2, leading to elevated expression of GSH-synthesizing enzymes.

  18. SMND-309 promotes neuron survival through the activation of the PI3K/Akt/CREB-signalling pathway.

    PubMed

    Wang, Youlei; Zhang, Jinjin; Han, Meng; Liu, Bo; Gao, Yulin; Ma, Peng; Zhang, Songzi; Zheng, Qingyin; Song, Xiaodong

    2016-10-01

    Context In clinical practice, the promotion of neuron survival is necessary to recover neurological functions after the onset of stroke. Objective This study aimed to investigate the post-ischaemic neuroprotective effect of SMND-309, a novel metabolite of salvianolic acid, on differentiated SH-SY5Y cells. Materials and methods SH-SY5Y cells were differentiated by pre-treating with 5 μM all-trans-retinoic acid for 6 d. The differentiated SH-SY5Y cells were exposed to oxygen-glucose deprivation (OGD) for 2 h and reperfusion (R) for 24 h to induce OGD/R injury. After OGD injury, differentiated SH-SY5Y cells were treated with or without SMND-309 (5, 10, 20 μM) for another 24 h. Cell viability was detected through Cell counting kit-8 assay and lactate dehydrogenase leakage assay. Apoptosis was evaluated through flow cytometry, caspase-3 activity assay. Changes in protein levels were assessed through Western blot. Results SMND-309 ameliorated the degree of injury in the differentiated SH-SY5Y cells by increasing cell viabilities (5 μM, 65.4% ± 4.1%; 10 μM, 69.8% ± 3.7%; 20 μM, 75.3% ± 5.1%) and by reducing LDH activity (20 μM, 2.5 fold) upon OGD/R stimulation. Annexin V-fluorescein isothiocyanate/propidium iodide staining results suggested that apoptotic rate of differentiated SH-SY5Y cells decreased from 43.8% induced by OGD/R injury to 19.2% when the cells were treated with 20 μM SMND-309. SMND-309 significantly increased the Bcl-2 level of the injured differentiated SH-SY5Y cells but decreased the caspase-3 activity of these cells by 1.6-fold. In contrast, SMND-309 did not affect the Bax level of these cells. SMND-309 evidently increased the protein expression of BDNF when Akt and CREB were activated. This function was antagonized by the addition of LY294002. Conclusion SMND-309 can prevent neuronal cell death in vitro. This process may be related to the activation of the PI3K/Akt/CREB-signalling pathway.

  19. Asiatic acid enhances Nrf2 signaling to protect HepG2 cells from oxidative damage through Akt and ERK activation.

    PubMed

    Qi, Zhimin; Ci, Xinxin; Huang, Jingbo; Liu, Qinmei; Yu, Qinlei; Zhou, Junfeng; Deng, Xuming

    2017-04-01

    Asiatic acid (AA), a natural triterpene isolated from the plant Centella asiatica, have antioxidative potential, but the molecular mechanism of AA against oxidative stress remains unclear. Our study was performed to investigate the antioxidative effect of AA against oxidative stress and the antioxidative mechanism in tert-butyl hydroperoxide (t-BHP) -stimulated the HepG2 cells. The results showed that AA suppressed t-BHP-induced cytotoxicity, apoptosis, and reactive oxygen species (ROS) generation. Additionally, AA activated the nuclear factor erythroid 2-related factor 2 (Nrf2) signal, which was closely related to induction Nrf2 nuclear translocation, reduction the expression of Keap1 and up-regulation the activity of the antioxidant response element (ARE). Meanwhile, activation of Nrf2 signal upregulated the protein expressions of antioxidant genes, including heme oxygenase-1 (HO-1), NAD(P)H: quinone oxidase (NQO-1), and glutamyl cysteine ligase catalytic subunit (GCLC). Excitingly, Knockout of Nrf2 almost abolished AA-mediated antioxidant activity and cytoprotection against t-BHP. Further studies showed the mechanism underlying that AA induced Nrf2 activation in HepG2 cells via Akt and ERK signal activation. We found Akt and ERK inhibitors treatment attenuated AA-mediated Nrf2 nuclear translocation. Furthermore, treatment with either Akt or ERK inhibitor also decreased AA-mediated cytoprotection against t-BHP-induced cellular damage. Collectively, these results presented in this study indicate that AA has the protective effect against t-BHP-induced cellular damage and oxidative stress by modulating Nrf2 signaling through activating the signals of Akt and ERK.

  20. TNF-{alpha} promotes human retinal pigment epithelial (RPE) cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression through activation of Akt/mTORC1 signaling

    SciTech Connect

    Wang, Cheng-hu; Cao, Guo-Fan; Jiang, Qin; Yao, Jin

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer TNF-{alpha} induces MMP-9 expression and secretion to promote RPE cell migration. Black-Right-Pointing-Pointer MAPK activation is not critical for TNF-{alpha}-induced MMP-9 expression. Black-Right-Pointing-Pointer Akt and mTORC1 signaling mediate TNF-{alpha}-induced MMP-9 expression. Black-Right-Pointing-Pointer SIN1 knockdown showed no significant effect on MMP-9 expression by TNF-{alpha}. -- Abstract: Tumor necrosis factor-alpha (TNF-{alpha}) promotes in vitro retinal pigment epithelial (RPE) cell migration to initiate proliferative vitreoretinopathy (PVR). Here we report that TNF-{alpha} promotes human RPE cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression. Inhibition of MMP-9 by its inhibitor or its neutralizing antibody inhibited TNF-{alpha}-induced in vitro RPE cell migration. Reversely, exogenously-added active MMP-9 promoted RPE cell migration. Suppression Akt/mTOR complex 1(mTORC1) activation by LY 294002 and rapamycin inhibited TNF-{alpha}-mediated MMP-9 expression. To introduce a constitutively active Akt (CA-Akt) in cultured RPE cells increased MMP-9 expression, and to block mTORC1 activation by rapamycin inhibited its effect. RNA interference (RNAi)-mediated silencing of SIN1, a key component of mTOR complex 2 (mTORC2), had no effect on MMP-9 expression or secretion. In conclusion, this study suggest that TNF-{alpha} promotes RPE cell migration by inducing MMP-9 expression through activation of Akt/ mTORC1, but not mTORC2 signaling.

  1. Breast Cancer Chemoresistance Mechanisms Through PI 3-Kinase and Akt Signaling

    DTIC Science & Technology

    2014-05-01

    discovered that the Akt pathway modulates breast cancer cell survival in response to genotoxic agents, and discovered a new substrate of Akt, MERIT40, that...12 3         INTRODUCTION Genotoxic chemotherapy agents are used to...resistance to genotoxic chemotherapy agents is activation of the PI3K/Akt signaling cascade. We proposed that genotoxic drugs induce the activation of

  2. Galectin-1 induces hepatocellular carcinoma EMT and sorafenib resistance by activating FAK/PI3K/AKT signaling

    PubMed Central

    Zhang, P-F; Li, K-S; Shen, Y-h; Gao, P-T; Dong, Z-R; Cai, J-B; Zhang, C; Huang, X-Y; Tian, M-X; Hu, Z-Q; Gao, D-M; Fan, J; Ke, A-W; Shi, G-M

    2016-01-01

    Galectin-1 (Gal-1) is involved in several pathological activities associated with tumor progression and chemoresistance, however, the role and molecular mechanism of Gal-1 activity in hepatocellular carcinoma (HCC) epithelial–mesenchymal transition (EMT) and sorafenib resistance remain enigmatic. In the present study, forced Gal-1 expression promoted HCC progression and sorafenib resistance. Gal-1 elevated αvβ3-integrin expression, leading to AKT activation. Moreover, Gal-1 overexpression induced HCC cell EMT via PI3K/AKT cascade activation. Clinically, our data revealed that Gal-1 overexpression is correlated with poor HCC survival outcomes and sorafenib response. These data suggest that Gal-1 may be a potential therapeutic target for HCC and a biomarker for predicting response to sorafenib treatment. PMID:27100895

  3. AKT Regulates BRCA1 Stability in Response to Hormone Signaling

    PubMed Central

    Nelson, Andrew C.; Lyons, Traci R.; Young, Christian D.; Hansen, Kirk C.; Anderson, Steven M.; Holt, Jeffrey T.

    2015-01-01

    BRCA1, with its binding partner BARD1, regulates the cellular response to DNA damage in multiple tissues, yet inherited mutations within BRCA1 result specifically in breast and ovarian cancers. This observation, along with several other lines of evidence, suggests a functional relationship may exist between hormone signaling and BRCA1 function. Our data demonstrates that AKT activation promotes the expression of BRCA1 in response to estrogen and IGF-1 receptor signaling. Further, we have identified a novel AKT phosphorylation site in BRCA1 at S694 which is responsive to activation of these signaling pathways. This rapid increase in BRCA1 protein levels appears to occur independently of new protein synthesis and treatment with the clinically utilized proteasome inhibitor bortezomib similarly leads to a rapid increase in BRCA1 protein levels. Together, these data suggest that AKT phosphorylation of BRCA1 increases total protein expression by preventing proteasomal degradation. AKT activation also appears to support nuclear localization of BRCA1, and co-expression of activated AKT with BRCA1 decreases radiation sensitivity, suggesting this interaction has functional consequences for BRCA1's role in DNA repair. We conclude that AKT regulates BRCA1 protein stability and function through direct phosphorylation of BRCA1. Further, the responsiveness of the AKT-BRCA1 regulatory pathway to hormone signaling may, in part, underlie the tissue specificity of BRCA1 mutant cancers. Pharmacological targets within this pathway could provide strategies for modulation of BRCA1 protein, which may prove therapeutically beneficial for the treatment of breast and ovarian cancers. PMID:20085797

  4. Activation of PI3K/Akt signaling in rostral ventrolateral medulla impairs brain stem cardiovascular regulation that underpins circulatory depression during mevinphos intoxication.

    PubMed

    Tsai, Ching-Yi; Chang, Alice Y W; Chan, Julie Y H; Chan, Samuel H H

    2014-03-01

    As the most widely used pesticides in the globe, the organophosphate compounds are understandably linked with the highest incidence of suicidal poisoning. Whereas the elicited toxicity is often associated with circulatory depression, the underlying mechanisms require further delineation. Employing the pesticide mevinphos as our experimental tool, we evaluated the hypothesis that transcriptional upregulation of nitric oxide synthase II (NOS II) by NF-κB on activation of the PI3K/Akt cascade in the rostral ventrolateral medulla (RVLM), the brain stem site that maintains blood pressure and sympathetic vasomotor tone, underpins the circulatory depressive effects of organophosphate poisons. Microinjection of mevinphos (10 nmol) bilaterally into the RVLM of anesthetized Sprague-Dawley rats induced a progressive hypotension that was accompanied sequentially by an increase (Phase I) and a decrease (Phase II) of an experimental index for the baroreflex-mediated sympathetic vasomotor tone. There were also progressive augmentations in PI3K or Akt enzyme activity and phosphorylation of p85 or Akt(Thr308) subunit in the RVLM that were causally related to an increase in NF-κB transcription activity and elevation in NOS II or peroxynitrite expression. Loss-of-function manipulations of PI3K or Akt in the RVLM significantly antagonized the reduced baroreflex-mediated sympathetic vasomotor tone and hypotension during Phase II mevinphos intoxication, and blunted the increase in NF-κB/NOS II/peroxynitrite signaling. We conclude that activation of the PI3K/Akt cascade, leading to upregulation of NF-κB/NOS II/peroxynitrite signaling in the RVLM, elicits impairment of brain stem cardiovascular regulation that underpins circulatory depression during mevinphos intoxication.

  5. Reactive oxygen species mediate Cr(VI)-induced carcinogenesis through PI3K/AKT-dependent activation of GSK-3β/β-catenin signaling

    SciTech Connect

    Son, Young-Ok; Pratheeshkumar, Poyil; Wang, Lei; Wang, Xin; Fan, Jia; Kim, Dong-Hern; Lee, Ju-Yeon; Zhang, Zhuo; Lee, Jeong-Chae; Shi, Xianglin

    2013-09-01

    Cr(VI) compounds are known human carcinogens that primarily target the lungs. Cr(VI) produces reactive oxygen species (ROS), but the exact effects of ROS on the signaling molecules involved in Cr(VI)-induced carcinogenesis have not been extensively studied. Chronic exposure of human bronchial epithelial cells to Cr(VI) at nanomolar concentrations (10–100 nM) for 3 months not only induced cell transformation, but also increased the potential of these cells to invade and migrate. Injection of Cr(VI)-stimulated cells into nude mice resulted in the formation of tumors. Chronic exposure to Cr(VI) increased levels of intracellular ROS and antiapoptotic proteins. Transfection with catalase or superoxide dismutase (SOD) prevented Cr(VI)-mediated increases in colony formation, cell invasion, migration, and xenograft tumors. While chronic Cr(VI) exposure led to activation of signaling cascades involving PI3K/AKT/GSK-3β/β-catenin and PI3K/AKT/mTOR, transfection with catalase or SOD markedly inhibited Cr(VI)-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the Cr(VI)-mediated increase in total and active β-catenin proteins and colony formation. In particular, Cr(VI) suppressed autophagy of epithelial cells under nutrition deprivation. Furthermore, there was a marked induction of AKT, GSK-3β, β-catenin, mTOR, and carcinogenic markers in tumor tissues formed in mice after injection with Cr(VI)-stimulated cells. Collectively, our findings suggest that ROS is a key mediator of Cr(VI)-induced carcinogenesis through the activation of PI3K/AKT-dependent GSK-3β/β-catenin signaling and the promotion of cell survival mechanisms via the inhibition of apoptosis and autophagy. - Highlights: • Chronic exposure to Cr(VI) induces carcinogenic properties in BEAS-2B cells. • ROS play an important role in Cr(VI)-induced tumorigenicity of BEAS-2B cells. • PI3K/AKT/GSK-3β/β-catenin signaling involved in Cr

  6. Compensation of the AKT signaling by ERK signaling in transgenic mice hearts overexpressing TRIM72

    SciTech Connect

    Ham, Young-Mi; Mahoney, Sarah Jane

    2013-06-10

    The AKT and ERK signaling pathways are known to be involved in cell hypertrophy, proliferation, survival and differentiation. Although there is evidence for crosstalk between these two signaling pathways in cellulo, there is less evidence for cross talk in vivo. Here, we show that crosstalk between AKT and ERK signaling in the hearts of TRIM72-overexpressing transgenic mice (TRIM72-Tg) with alpha-MHC promoter regulates and maintains their heart size. TRIM72, a heart- and skeletal muscle-specific protein, downregulates AKT-mTOR signaling via IRS-1 degradation and reduces the size of rat cardiomyocytes and the size of postnatal TRIM72-Tg hearts. TRIM72 expression was upregulated by hypertrophic inducers in cardiomyocytes, while IRS-1 was downregulated by IGF-1. TRIM72 specifically regulated IGF-1-dependent AKT-mTOR signaling, resulting in a reduction of the size of cardiomyocytes. Postnatal TRIM72-Tg hearts were smaller than control-treated hearts with inhibition of AKT-mTOR signaling. However, adult TRIM72-Tg hearts were larger than of control despite the suppression of AKT-mTOR signaling. Activation of ERK, PKC-α, and JNK were observed to be elevated in adult TRIM72-Tg, and these signals were mediated by ET-1 via the ET receptors A and B. Altogether, these results suggest that AKT signaling regulates cardiac hypertrophy in physiological conditions, and ERK signaling compensates for the absence of AKT signaling during TRIM72 overexpression, leading to pathological hypertrophy. -- Highlights: • TRIM72 inhibits AKT signaling through ubiquitination of IRS-1 in cardiac cells. • TRIM72 regulates the size of cardiac cells. • TRIM72 regulates size of postnatal TRIM72-overexpressing transgenic mice hearts. • Adult TRIM72-overexpressing transgenic mice hearts showed cardiac dysfunction. • Adult TRIM72 transgenic mice hearts showed higher expression of endothelin receptors.

  7. Simvastatin inhibits glucose uptake activity and GLUT4 translocation through suppression of the IR/IRS-1/Akt signaling in C2C12 myotubes.

    PubMed

    Li, Weihua; Liang, Xiaojing; Zeng, Zhipeng; Yu, Kaizhen; Zhan, Shaopeng; Su, Qiang; Yan, Yinzhi; Mansai, Huseen; Qiao, Weitong; Yang, Qi; Qi, Zhongquan; Huang, Zhengrong

    2016-10-01

    Simvastatin,a 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase inhibitor, is clinically used in the prevention and treatment of cardiovascular diseases. Numerous studies demonstrate that statins increase the risk of new-onset diabetes in long-term therapy, but mechanisms underpinning this effect are still unclear. Here, we investigated whether simvastatin inhibited the glucose uptake activity and the underlying mechanisms in C2C12 myotubes. Our studies showed that simvastatin significantly inhibited glucose uptake activity and GLUT4 translocation, whereas the effect was reversible with mevalonolactone (ML), which acts as an intermediate of cholesterol synthesis pathway. Mechanistically, the inhibition of glucose uptake and GLUT4 translocation elicited by simvastatin were associated with the suppression of the insulin receptor (IR)/IR substrate (IRS)/Akt signaling cascade. Simvastatin suppressed the phosphorylation of IR, IRS-1 and Akt, and total expression of IR or IRS-1, but did not affect Akt. Furthermore, simvastatin decreased Rac1 GTP binding. In conclusion, our findings indicate that simvastatin suppresses glucose uptake activity and GLUT4 translocation via IR-dependent IRS-1/PI3K/Akt pathway. These results provide an important new insight into the mechanism of statins on insulin sensitivity which may be associated with new-onset diabetes.

  8. Protection afforded by quercetin against H2O2-induced apoptosis on PC12 cells via activating PI3K/Akt signal pathway.

    PubMed

    Chen, Liang; Sun, Lejin; Liu, Zhene; Wang, Hongxia; Xu, Cunli

    2016-01-01

    Cell damage and apoptosis induced by oxidative stress have been involved in various neurodegenerative diseases. This study aims to explore the neuro-protective effects of quercetin on PC12 cells apoptosis induced by hydrogen peroxide (H(2)O(2)) and the underlying mechanisms. The cell viability was detected, as well as the production of reactive oxygen species (ROS), lactate dehydrogenase (LDH) leakage, and the activities of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and malondialdehyde (MDA) of the cells in control, H(2)O(2) and quercetin groups. It finally turned out that quercetin might protect PC12 cells against the negative effect of H(2)O(2) by decreasing of LDH release, ROS concentration and MDA level and regaining the GSH-Px and SOD activities. To investigate the mechanism, LY294002 was introduced, the phosphatidylinositol-3-kinase (PI3K) inhibitor. Bax/Bcl-2 ratio and Akt phosphorylation (p-Akt) were examined by Western blot analysis. The data showed that LY294002 almost had the same effects with H(2)O(2), which was also significantly reversed by quercetin could enhance Bax/Bcl-2 ratio and adjust the p-Akt expression, which indicated quercetin might protect PC12 cells against the negative effect of H(2)O(2) via activating the PI3K/Akt signal pathway.

  9. PI3K/AKT Signaling Regulates Bioenergetics in Immortalized Hepatocytes

    PubMed Central

    Li, Chen; Li, Yang; He, Lina; Agarwal, Amit R.; Zeng, Ni; Cadenas, Enrique; Stiles, Bangyan L.

    2013-01-01

    Regulation of cellular bioenergetics by PI3K/AKT signaling was examined in isogenic hepatocyte cell lines lacking the major inhibitor of PI3K/AKT signaling, PTEN (phosphatase and tensin homolog deleted on Chromosome 10). PI3K/AKT signaling was manipulated using the activator (IGF-1) and the inhibitor (LY 294002) of the PI3K/AKT pathway. Activation of PI3K/AKT signaling resulted in an enhanced anaerobic glycolysis and mitochondrial respiration. AKT, when phosphorylated and activated, translocated to mitochondria and localized within the membrane structure of mitochondria, where it phosphorylated a number of mitochondrial residence proteins including the subunits α and β of ATP synthase. Inhibition of GSK3β by either phosphorylation by AKT or lithium chloride resulted in activation of pyruvate dehydrogenase, i.e., decrease of its phosphorylated form. AKT-dependent phosphorylation of ATP synthase subunits α and β resulted in an increased complex activity. AKT translocation to mitochondria was associated with an increased expression and activity of complex I. These data suggest that the mitochondrial signaling pathway AKT-GSK3β-PDH, AKT-dependent phosphorylation of ATP synthase, and upregulation of mitochondrial complex I expression and activity are involved in the control of mitochondrial bioenergetics by increasing substrate availability and regulating the mitochondrial catalytic/energy-transducing capacity. PMID:23376468

  10. Fucoidan Induces ROS-Dependent Apoptosis in 5637 Human Bladder Cancer Cells by Downregulating Telomerase Activity via Inactivation of the PI3K/Akt Signaling Pathway.

    PubMed

    Han, Min Ho; Lee, Dae-Sung; Jeong, Jin-Woo; Hong, Su-Hyun; Choi, Il-Whan; Cha, Hee-Jae; Kim, Suhkmann; Kim, Heui-Soo; Park, Cheol; Kim, Gi-Young; Moon, Sung-Kwon; Kim, Wun-Jae; Hyun Choi, Yung

    2017-02-01

    Preclinical Research Fucoidan, a sulfated polysaccharide, is a compound found in various species of seaweed that has anti-viral, anti-bacterial, anti-oxidant, anti-inflammatory, and immunomodulatory activities; however, the underlying relationship between apoptosis and anti-telomerase activity has not been investigated. Here, we report that fucoidan-induced apoptosis in 5637 human bladder cancer cells was associated with an increase in the Bax/Bcl-2 ratio, the dissipation of the mitochondrial membrane potential (MMP, Δψm), and cytosolic release of cytochrome c from the mitochondria. Under the same experimental conditions, fucoidan-treatment decreased hTERT (human telomerase reverse transcriptase) expression and the transcription factors, c-myc and Sp1. This was accompanied by decreased telomerase activity. Fucoidan-treatment also suppressed activation of the PI3K/Akt signaling pathway. Inhibition of PI3K/Akt signaling enhanced fucoidan-induced apoptosis and anti-telomerase activity. Meanwhile, fucoidan treatment increased the generation of intracellular ROS, whereas the over-elimination of ROS by N-acetylcysteine, an anti-oxidant, attenuated fucoidan-induced apoptosis, inhibition of hTERT, c-myc, and Sp1 expression, and reversed fucoidan-induced inactivation of the PI3K/Akt signaling pathway. Collectively, these data indicate that the induction of apoptosis and the inhibition of telomerase activity by fucoidan are mediated via ROS-dependent inactivation of the PI3K/Akt pathway. Drug Dev Res 78 : 37-48, 2017.   © 2016 Wiley Periodicals, Inc.

  11. Carbohydrate-binding motif in Chitinase 3-like 1 (CHI3L1/YKL-40) specifically activates Akt signaling pathway in colonic epithelial cells

    PubMed Central

    Chen, Chun-Chuan; Llado, Victoria; Eurich, Katrin; Tran, Hoa T.; Mizoguchi, Emiko

    2011-01-01

    Host-microbial interactions play a key role during the development of colitis. We have previously shown that chinase 3-like 1 (CHI3L1) is an inducible molecule overexpressed in colonic epithelial cells (CECs) under inflammatory conditions. In this study, we found that chitin-binding motif (CBM) of CHI3L1 is specifically associated with the CHI3L1-mediated activation of the Akt-signaling in CEC by transfecting the CBM-mutant CHI3L1 vectors in SW480 CECs. Downstream, CHI3L1 enhanced the secretion of IL-8 and TNFα in a dose-dependent manner. We previously show that 325 through 339 amino-acids in CBM are crucial for the biological function of CHI3L1. Here we demonstrated that 325th–339th residues of CBM in CHI3L1 is a critical region for the activation of Akt, IL-8 production, and for a specific cellular localization of CHI3L1. In conclusion, CBM region of CHI3L1 is critical in activating Akt signaling in CECs, and the activation may be associated with the development of chronic colitis. PMID:21546314

  12. Squamosamide derivative FLZ protected dopaminergic neuron by activating Akt signaling pathway in 6-OHDA-induced in vivo and in vitro Parkinson's disease models.

    PubMed

    Bao, Xiu-Qi; Kong, Xiang-Chen; Kong, Li-Bing; Wu, Liang-Yu; Sun, Hua; Zhang, Dan

    2014-02-14

    Parkinson's disease (PD) is a neurodegenerative disease affecting up to 80% of dopaminergic neurons in the nigrostriatal pathway. FLZ, a novel synthetic squamosamide derivative from a Chinese herb, has been shown to have neuroprotective effects in experimental PD models. In this study, we carried out a set of in vitro and in vivo experiments to address the neuroprotective effect of FLZ and related mechanism. The results showed that FLZ significantly improved motor dysfunction and dopaminergic neuronal loss of rats injured by 6-hydroxydopamine (6-OHDA). The beneficial effects of FLZ attributed to the elevation of dopaminergic neuron number, dopamine level and tyrosine hydroxylase (TH) activity. Mechanistic study showed that FLZ protected TH activity and dopaminergic neurons through decreasing α-synuclein (α-Syn) expression and the interaction between α-Syn and TH. Further studies indicated the involvement of phosphoinositide 3-kinases (PI3K)/Akt signaling pathway in the protective effect of FLZ since it showed that blocking PI3K/Akt signaling pathway prevented the expression of α-Syn and attenuated the neuroprotection of FLZ. In addition, FLZ treatment reduced the expression of RTP801, an important protein involved in the pathogenesis of PD. Taken together, these results revealed that FLZ suppressed α-Syn expression and elevated TH activity in dopaminergic neuron through activating Akt survival pathway in 6-OHDA-induced PD models. The data also provided evidence that FLZ had potent neuroprotecive effects and might become a new promising agent for PD treatment.

  13. Cadmium induces carcinogenesis in BEAS-2B cells through ROS-dependent activation of PI3K/AKT/GSK-3β/β-catenin signaling

    SciTech Connect

    Son, Young-Ok; Wang, Lei; Poyil, Pratheeshkumar; Budhraja, Amit; Hitron, J. Andrew; Zhang, Zhuo; Lee, Jeong-Chae; Shi, Xianglin

    2012-10-15

    Cadmium has been widely used in industry and is known to be carcinogenic to humans. Although it is widely accepted that chronic exposure to cadmium increases the incidence of cancer, the mechanisms underlying cadmium-induced carcinogenesis are unclear. The main aim of this study was to investigate the role of reactive oxygen species (ROS) in cadmium-induced carcinogenesis and the signal transduction pathways involved. Chronic exposure of human bronchial epithelial BEAS-2B cells to cadmium induced cell transformation, as evidenced by anchorage-independent growth in soft agar and clonogenic assays. Chronic cadmium treatment also increased the potential of these cells to invade and migrate. Injection of cadmium-stimulated cells into nude mice resulted in the formation of tumors. In contrast, the cadmium-mediated increases in colony formation, cell invasion and migration were prevented by transfection with catalase, superoxide dismutase-1 (SOD1), or SOD2. In particular, chronic cadmium exposure led to activation of signaling cascades involving PI3K, AKT, GSK-3β, and β-catenin and transfection with each of the above antioxidant enzymes markedly inhibited cadmium-mediated activation of these signaling proteins. Inhibitors specific for AKT or β-catenin almost completely suppressed the cadmium-mediated increase in total and active β-catenin proteins and colony formation. Moreover, there was a marked induction of AKT, GSK-3β, β-catenin, and carcinogenic markers in tumor tissues formed in mice after injection with cadmium-stimulated cells. Collectively, our findings suggest a direct involvement of ROS in cadmium-induced carcinogenesis and implicate a role of AKT/GSK-3β/β-catenin signaling in this process. -- Highlights: ► Chronic exposure to cadmium induces carcinogenic properties in BEAS-2B cells. ► ROS involved in cadmium-induced tumorigenicity of BEAS-2B cells. ► Cadmium activates ROS-dependent AKT/GSK-3β/β-catenin-mediated signaling. ► ROS

  14. Novel pathway in Bcr-Abl signal transduction involves Akt-independent, PLC-gamma1-driven activation of mTOR/p70S6-kinase pathway.

    PubMed

    Markova, B; Albers, C; Breitenbuecher, F; Melo, J V; Brümmendorf, T H; Heidel, F; Lipka, D; Duyster, J; Huber, C; Fischer, T

    2010-02-04

    In chronic myeloid leukemia, activation of the phosphoinositide 3-kinase (PI3K)/Akt pathway is crucial for survival and proliferation of leukemic cells. Essential downstream molecules involve mammalian target of rapamycin (mTOR) and S6-kinase. Here, we present a comprehensive analysis of the molecular events involved in activation of these key signaling pathways. We provide evidence for a previously unrecognized phospholipase C-gamma1 (PLC-gamma1)-controlled mechanism of mTOR/p70S6-kinase activation, which operates in parallel to the classical Akt-dependent machinery. Short-term imatinib treatment of Bcr-Abl-positive cells caused dephosphorylation of p70S6-K and S6-protein without inactivation of Akt. Suppression of Akt activity alone did not affect phosphorylation of p70-S6K and S6. These results suggested the existence of an alternative mechanism for mTOR/p70S6-K activation. In Bcr-Abl-expressing cells, we detected strong PLC-gamma1 activation, which was suppressed by imatinib. Pharmacological inhibition and siRNA knockdown of PLC-gamma1 blocked p70S6-K and S6 phosphorylation. By inhibiting the Ca-signaling, CaMK and PKCs we demonstrated participation of these molecules in the pathway. Suppression of PLC-gamma1 led to inhibition of cell proliferation and enhanced apoptosis. The novel pathway proved to be essential for survival and proliferation of leukemic cells and almost complete cell death was observed upon combined PLC-gamma1 and Bcr-Abl inhibition. The pivotal role of PLC-gamma1 was further confirmed in a mouse leukemogenesis model.

  15. Quercetin attenuates cell apoptosis in focal cerebral ischemia rat brain via activation of BDNF-TrkB-PI3K/Akt signaling pathway.

    PubMed

    Yao, Rui-Qin; Qi, Da-Shi; Yu, Hong-Li; Liu, Jing; Yang, Li-Hua; Wu, Xiu-Xiang

    2012-12-01

    Many studies have demonstrated that apoptosis play an important role in cerebral ischemic pathogenesis and may represent a target for treatment. Neuroprotective effect of quercetin has been shown in a variety of brain injury models including ischemia/reperfusion. It is not clear whether BDNF-TrkB-PI3K/Akt signaling pathway mediates the neuroprotection of quercetin, though there has been some reports on the quercetin increased brain-derived neurotrophic factor (BDNF) level in brain injury models. We therefore first examined the neurological function, infarct volume and cell apoptosis in quercetin treated middle cerebral artery occlusion (MCAO) rats. Then the protein expression of BDNF, cleaved caspase-3 and p-Akt were evaluated in either the absence or presence of PI3K inhibitor (LY294002) or tropomyosin receptor kinase B (TrkB) receptor antagonist (K252a) by immunohistochemistry staining and western blotting. Quercetin significantly improved neurological function, while it decreased the infarct volume and the number of TdT mediated dUTP nick end labeling positive cells in MCAO rats. The protein expression of BDNF, TrkB and p-Akt also increased in the quercetin treated rats. However, treatment with LY294002 or K252a reversed the quercetin-induced increase of BDNF and p-Akt proteins and decrease of cleaved caspase-3 protein in focal cerebral ischemia rats. These results demonstrate that quercetin can decrease cell apoptosis in the focal cerebral ischemia rat brain and the mechanism may be related to the activation of BDNF-TrkB-PI3K/Akt signaling pathway.

  16. l-carnitine protects human hepatocytes from oxidative stress-induced toxicity through Akt-mediated activation of Nrf2 signaling pathway.

    PubMed

    Li, Jinlian; Zhang, Yanli; Luan, Haiyun; Chen, Xuehong; Han, Yantao; Wang, Chunbo

    2016-05-01

    In our previous study, l-carnitine was shown to have cytoprotective effect against hydrogen peroxide (H2O2)-induced injury in human normal HL7702 hepatocytes. The aim of this study was to investigate whether the protective effect of l-carnitine was associated with the nuclear factor erythroid 2 (NFE2)-related factor 2 (Nrf2) pathway. Our results showed that pretreatment with l-carnitine augmented Nrf2 nuclear translocation, DNA binding activity and heme oxygenase-1 (HO-1) expression in H2O2-treated HL7702 cells, although l-carnitine treatment alone had no effect on them. Analysis using Nrf2 siRNA demonstrated that Nrf2 activation was involved in l-carnitine-induced HO-1 expression. In addition, l-carnitine-mediated protection against H2O2 toxicity was abrogated by Nrf2 siRNA, indicating the important role of Nrf2 in l-carnitine-induced cytoprotection. Further experiments revealed that l-carnitine pretreatment enhanced the phosphorylation of Akt in H2O2-treated cells. Blocking Akt pathway with inhibitor partly abrogated the protective effect of l-carnitine. Moreover, our finding demonstrated that the induction of Nrf2 translocation and HO-1 expression by l-carnitine directly correlated with the Akt pathway because Akt inhibitor showed inhibitory effects on the Nrf2 translocation and HO-1 expression. Altogether, these results demonstrate that l-carnitine protects HL7702 cells against H2O2-induced cell damage through Akt-mediated activation of Nrf2 signaling pathway.

  17. MiR-20a Induces Cell Radioresistance by Activating the PTEN/PI3K/Akt Signaling Pathway in Hepatocellular Carcinoma

    SciTech Connect

    Zhang, Yuqin; Zheng, Lin; Ding, Yi; Li, Qi; Wang, Rong; Liu, Tongxin; Sun, Quanquan; Yang, Hua; Peng, Shunli; Wang, Wei; Chen, Longhua

    2015-08-01

    Purpose: To investigate the role of miR-20a in hepatocellular carcinoma (HCC) cell radioresistance, which may reveal potential strategies to improve treatment. Methods and Materials: The expression of miR-20a and PTEN were detected in HCC cell lines and paired primary tissues by quantitative real-time polymerase chain reaction. Cell radiation combined with colony formation assays was administrated to discover the effect of miR-20a on radiosensitivity. Bioinformatics prediction and luciferase assay were used to identify the target of miR-20a. The phosphatidylinositol 3-kinase inhibitor LY294002 was used to inhibit phosphorylation of Akt, to verify whether miR-20a affects HCC cell radioresistance through activating the PTEN/PI3K/Akt pathway. Results: MiR-20a levels were increased in HCC cell lines and tissues, whereas PTEN was inversely correlated with it. Overexpression of miR-20a in Bel-7402 and SMMC-7721 cells enhances their resistance to the effect of ionizing radiation, and the inhibition of miR-20a in HCCLM3 and QGY-7701 cells sensitizes them to it. PTEN was identified as a direct functional target of miR-20a for the induction of radioresistance. Overexpression of miR-20a activated the PTEN/PI3K/Akt signaling pathway. Additionally, the kinase inhibitor LY294002 could reverse the effect of miR-20a–induced radioresistance. Conclusion: MiR-20a induces HCC cell radioresistance by activating the PTEN/PI3K/Akt pathway, which suggests that miR-20a/PTEN/PI3K/Akt might represent a target of investigation for developing effective therapeutic strategies against HCC.

  18. Relaxin attenuates aristolochic acid induced human tubular epithelial cell apoptosis in vitro by activation of the PI3K/Akt signaling pathway.

    PubMed

    Xie, Xiang-Cheng; Zhao, Ning; Xu, Qun-Hong; Yang, Xiu; Xia, Wen-Kai; Chen, Qi; Wang, Ming; Fei, Xiao

    2017-04-06

    Aristolochic acid nephropathy remains a leading cause of chronic kidney disease (CKD), however few treatment strategies exist. Emerging evidence has shown that H2 relaxin (RLX) possesses powerful antifibrosis and anti-apoptotic properties, therefore we aimed to investigate whether H2 relaxin can be employed to reduce AA-induced cell apoptosis. Human proximal tubular epithelial (HK-2) cells exposed to AA-I were treated with or without administration of H2 RLX. Cell viability was examined using the WST-8 assay. Apoptotic morphologic alterations were observed using the Hoechst 33342 staining method. Apoptosis was detected using flow cytometry. The expression of caspase 3, caspase 8, caspase 9, ERK1/2, Bax, Bcl-2, and Akt proteins was determined by Western blot. Co-treatment with RLX reversed the increased apoptosis observed in the AA-I only treated group. RLX restored expression of phosphorylated Akt which found to be decreased in the AA-I only treated cells. RLX co-treatment led to a decrease in the Bax/Bcl-2 ratio as well as the cleaved form of caspase-3 compared to the AA-I only treated cells. This anti-apoptotic effect of RLX was attenuated by co-administration of the Akt inhibitor LY294002. The present study demonstrated H2 RLX can decrease AA-I induced apoptosis through activation of the PI3K/Akt signaling pathway.

  19. Redox-Sensitive Oxidation and Phosphorylation of PTEN Contribute to Enhanced Activation of PI3K/Akt Signaling in Rostral Ventrolateral Medulla and Neurogenic Hypertension in Spontaneously Hypertensive Rats

    PubMed Central

    Wu, Kay L.H.; Wu, Chiung-Ai; Wu, Chih-Wei; Chan, Samuel H.H.; Chang, Alice Y.W.

    2013-01-01

    Abstract Aims: The activity of phosphoinositide 3-kinase (PI3K)/serine/threonine protein kinase (Akt) is enhanced under hypertension. The phosphatase and tensin homolog deleted on chromosome 10 (PTEN) is a negative regulator of PI3K signaling, and its activity is redox-sensitive. In the rostral ventrolateral medulla (RVLM), which is responsible for the maintenance of blood pressure, oxidative stress plays a pivotal role in neurogenic hypertension. The present study evaluated the hypothesis that redox-sensitive inactivation of PTEN results in enhanced PI3K/Akt signaling in RVLM, leading to neurogenic hypertension. Results: Compared to age-matched normotensive Wistar-Kyoto (WKY) rats, PTEN inactivation in the form of oxidation and phosphorylation were greater in RVLM of spontaneously hypertensive rats (SHR). PTEN inactivation was accompanied by augmented PI3K activity and PI3K/Akt signaling, as reflected by the increase in phosphorylation of Akt and mammalian target of rapamycin. Intracisternal infusion of tempol or microinjection into the bilateral RVLM of adenovirus encoding superoxide dismutase significantly antagonized the PTEN inactivation and blunted the enhanced PI3K/Akt signaling in SHR. Gene transfer of PTEN to RVLM in SHR also abrogated the enhanced Akt activation and promoted antihypertension. Silencing PTEN expression in RVLM with small-interfering RNA, on the other hand, augmented PI3K/Akt signaling and promoted long-term pressor response in normotensive WKY rats. Innovation: The present study demonstrated for the first time that the redox-sensitive check-and-balance process between PTEN and PI3K/Akt signaling is engaged in the pathogenesis of hypertension. Conclusion: We conclude that an aberrant interplay between the redox-sensitive PTEN and PI3k/Akt signaling in RVLM underpins neural mechanism of hypertension. Antioxid. Redox Signal. 18, 36–50. PMID:22746319

  20. Oncogenic S1P signalling in EBV-associated nasopharyngeal carcinoma activates AKT and promotes cell migration through S1P receptor 3.

    PubMed

    Lee, Hui Min; Lo, Kwok-Wai; Wei, Wenbin; Tsao, Sai Wah; Chung, Grace Tin Yun; Ibrahim, Maha Hafez; Dawson, Christopher W; Murray, Paul G; Paterson, Ian C; Yap, Lee Fah

    2017-02-27

    Undifferentiated nasopharyngeal carcinoma (NPC) is a cancer with high metastatic potential that is consistently associated with Epstein-Barr virus (EBV) infection. In this study, we have investigated the functional contribution of sphingosine-1-phosphate (S1P) signalling to the pathogenesis of NPC. We show that EBV infection or ectopic expression of the EBV-encoded latent genes (EBNA1, LMP1 and LMP2A) can up-regulate sphingosine kinase 1 (SPHK1), the key enzyme that produces S1P, in NPC cell lines. Exogenous addition of S1P promotes the migration of NPC cells through the activation of AKT; shRNA knockdown of SPHK1 resulted in a reduction in the levels of activated AKT and inhibition of cell migration. We also show that S1P receptor 3 (S1PR3) mRNA is over-expressed in EBV-positive NPC patient-derived xenografts and a subset of primary NPC tissues, and that knockdown of S1PR3 suppressed the activation of AKT and the S1P-induced migration of NPC cells. Taken together, our data point to a central role for EBV in mediating the oncogenic effects of S1P in NPC and identify S1P signalling as a potential therapeutic target in this disease.

  1. Salidroside inhibits oxygen glucose deprivation (OGD)/re-oxygenation-induced H9c2 cell necrosis through activating of Akt-Nrf2 signaling.

    PubMed

    Zheng, Koulong; Sheng, Zhenqiang; Li, Yefei; Lu, Huihe

    2014-08-15

    Oxygen glucose deprivation (OGD)/re-oxygenation has been applied to cultured cardiomyocytes to create a cellular model of ischemic heart damage. In the current study, we explored the potential role of salidroside against OGD/re-oxygenation-induced damage in H9c2 cardiomyocytes, and studied the underlying mechanisms. We found that OGD/re-oxygenation primarily induced necrosis in H9c2 cells, which was inhibited by salidroside. Salidroside suppressed OGD/re-oxygenation-induced reactive oxygen species (ROS) production, p53 mitochondrial translocation and cyclophilin D (Cyp-D) association as well as mitochondrial membrane potential (MMP) decrease in H9c2 cells. Meanwhile, salidroside activated Akt and promoted transcription of NF-E2-related factor 2 (Nrf2)-regulated genes (heme oxygenase-1 (HO-1) and quinone oxidoreductase 1 (NQO-1)). Significantly, Nrf2 shRNA knockdown or Akt inhibitors (LY 294002 and wortmannin) not only prevented salidroside-induced HO-1/NQO-1 transcription, but also alleviated salidroside-mediated cytoprotective effect against OGD/re-oxygenation in H9c2 cells. These observations suggest that salidroside activates Nrf2-regulated anti-oxidant signaling, and protects against OGD/re-oxygenation-induced H9c2 cell necrosis via activation of Akt signaling.

  2. Gremlin promotes retinal pigmentation epithelial (RPE) cell proliferation, migration and VEGF production via activating VEGFR2-Akt-mTORC2 signaling

    PubMed Central

    Liu, Yuan; Chen, Zhijun; Cheng, Haixia; Chen, Juan; Qian, Jing

    2017-01-01

    Retinopathy of prematurity (ROP) is characterized by late-phase pathologic retinal vasoproliferation. Gremlin is a novel vascular endothelial growth factors (VEGF) receptor 2 (VEGFR2) agonist and promotes angiogenic response. We demonstrated that gremlin expression was significantly increased in retinas of ROP model mice, which was correlated with VEGF upregulation. In retinal pigmentation epithelial (RPE) cells, gremlin activated VEGFR2-Akt-mTORC2 (mammalian target of rapamycin complex 2) signaling, and promoted cell proliferation, migration and VEGF production. VEGFR inhibition (by SU5416) or shRNA knockdown almost abolished gremlin-mediated pleiotropic functions in RPE cells. Further, pharmacological inhibition of Akt-mTOR, or shRNA knockdown of key mTORC2 component (Rictor or Sin1) also attenuated gremlin-exerted activities in RPE cells. We conclude that gremlin promotes RPE cell proliferation, migration and VEGF production possibly via activating VEGFR2-Akt-mTORC2 signaling. Gremlin could be a novel therapeutic target of ROP or other retinal vasoproliferation diseases. PMID:27894090

  3. PDGF-D/PDGFRβ promotes tongue squamous carcinoma cell (TSCC) progression via activating p38/AKT/ERK/EMT signal pathway.

    PubMed

    Zhang, Hong; Sun, Jia-Dong; Yan, Ling-Jian; Zhao, Xiao-Peng

    2016-09-16

    Platelet-derived growth factor D (PDGF-D) signaling plays significant roles during the development and progression of human malignancies via interacting with the receptor of PDGF-D (PDGFR). Meanwhile, the majority of human tumor metastasis is closely associated with epithelial-mesenchymal transition (EMT). However, the underlying mechanism between PDGF-D/PDGFR signaling and EMT which involved in tumor metastasis remain dismal. This study aimed to investigate the role of PDGF-D signaling during EMT process of tongue squamous cell carcinoma (TSCC). In our study, the expression of PDGF-D and PDGFR were examined in primary TSCC samples and the expression of PDGF-D was also determined in TSCC cell lines. In addition, the correlation between PDGF-D expression and TSCC aggressive histopathological features was analyzed. Our results implied that upregulation of PDGFRβ in UM1 cells induced with exogenous PDGF-D can remarkably promote tumor cells invasiveness; conversely, when using small interfering RNA (siRNA), the invasiveness can be severely prohibited. Furthermore, PDGF-D downstream signal molecules p38, AKT, ERK and EMT biomarkers (E-cadherin, N-cadherin, Vimentin and snail) were measured using Western blot. Our results showed that PDGF-D can induce p38, AKT and ERK phosphorylation; downregulate epithelial markers and upregulate mesenchymal markers. On the contrary, PDGFRβ siRNA significantly prohibited p38, AKT and ERK phosphorylation; inhibited EMT process. Function analysis revealed that PDGFRβ siRNA obviously interfered with UM1 cell migration and invasion, according to transwell and wound healing assay. In conclusion, this study suggested that EMT process can be triggered by the PDGF-D/PDGFRβ axis in TSCC, and then involved in the tumor cell invasion via activation of p38/AKT/ERK/EMT pathway.

  4. Crop milk protein is synthesised following activation of the IRS1/Akt/TOR signalling pathway in the domestic pigeon (Columba livia).

    PubMed

    Hu, X-C; Gao, C-Q; Wang, X-H; Yan, H-C; Chen, Z-S; Wang, X-Q

    2016-12-01

    The experiment was conducted to study whether insulin receptor substance 1 (IRS1) / Protein kinase B (Akt)/target of the rapamycin (TOR) signalling pathway activation stimulates crop milk protein synthesis in the domestic pigeon (Columba livia). Crop milk was collected from ten 1-d-old squabs and analysed for nutrient content. During the non-breeding period and the first day of lactation, blood samples were collected from 5 pairs of breeding pigeons and the levels of prolactin and insulin were determined. Crop samples were collected from 5 pairs of breeders at d 14 and 16 of the incubation period and d 1, 3 and 7 of the lactation period. Crop samples were evaluated for changes in crop weight and thickness and changes in the expression patterns of IRS1/Akt/TOR signalling pathway-related proteins. The results demonstrated that prolactin induces a gradual increase in the relative weight and thickness of the crop, with crops reaching a maximum size at the third day of lactation. Pigeon crop milk contains 64.1% crude protein and 29.7% crude fat based on dry weight. Serum prolactin and insulin levels in the lactation period were significantly higher than those in the non-breeding period. Compared with non-breeding pigeons, the expression of the phosphorylated IRS1 phosphorylated Akt, phosphorylated TOR, phosphorylated ribosomal protein S6 kinase, phosphorylated S6, phosphorylated eukaryotic initiation factor 4E binding protein 1 and eukaryotic initiation factor 4E were significantly up-regulated in the crop of pigeons in the lactation period. In conclusion, prolactin might induce changes in crop tissue and form the physiological structure for crop milk synthesis. Furthermore, the synthesis of crop milk protein is regulated by activation of the IRS1/Akt/TOR signalling pathway.

  5. Paeonia lactiflora Pall. protects against ANIT-induced cholestasis by activating Nrf2 via PI3K/Akt signaling pathway

    PubMed Central

    Ma, Xiao; Zhao, Yan-ling; Zhu, Yun; Chen, Zhe; Wang, Jia-bo; Li, Rui-yu; Chen, Chang; Wei, Shi-zhang; Li, Jian-yu; Liu, Bing; Wang, Rui-lin; Li, Yong-gang; Wang, Li-fu; Xiao, Xiao-he

    2015-01-01

    Background Paeonia lactiflora Pall. (PLP), a traditional Chinese herbal medicine, has been used for hepatic disease treatment over thousands of years. In our previous study, PLP was shown to demonstrate therapeutic effect on hepatitis with severe cholestasis. The aim of this study was to evaluate the antioxidative effect of PLP on alpha-naphthylisothiocyanate (ANIT)-induced cholestasis by activating NF-E2-related factor 2 (Nrf2) via phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. Materials and methods Liquid chromatography-mass spectrometry (LC-MS) was performed to identify the main compounds present in PLP. The mechanism of action of PLP and its therapeutic effect on cholestasis, induced by ANIT, were further investigated. Serum indices such as total bilirubin (TBIL), direct bilirubin (DBIL), aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), γ-glutamyl transpeptidase (γ-GT), and total bile acid (TBA) were measured, and histopathology of liver was also performed to determine the efficacy of treatment with PLP. Moreover, in order to illustrate the underlying signaling pathway, liver glutathione (GSH) content and mRNA or protein levels of glutamate-cysteine ligase catalytic subunit (GCLc), glutamate-cysteine ligase modulatory subunit (GCLm), Akt, heme oxygenase-1 (HO-1), NAD(P)H/quinone oxidoreductase 1 (Nqo1), and Nrf2 were further analyzed. In addition, validation of PLP putative target network was also performed in silico. Results Four major compounds including paeoniflorin, albiflorin, oxypaeoniflorin, and benzoylpaeoniflorin were identified by LC-MS analysis in water extract of PLP. Moreover, PLP could remarkably downregulate serum levels of TBIL, DBIL, AST, ALT, ALP, γ-GT, and TBA, and alleviate the histological damage of liver tissue caused by ANIT. It enhanced antioxidative system by activating PI3K/Akt/Nrf2 pathway through increasing Akt, Nrf2, HO-1, Nqo1, GCLc, and GCLm expression. The putative

  6. CB2 cannabinoid receptor activation promotes colon cancer progression via AKT/GSK3β signaling pathway

    PubMed Central

    Martínez-Martínez, Esther; Martín-Ruiz, Asunción; Martín, Paloma; Calvo, Virginia; Provencio, Mariano; García, José M.

    2016-01-01

    The pharmacological activation of the cannabinoid receptor type 2, CB2, has been shown to elicit anti-tumoral mechanisms in different cancer types. However, little is known about its endogenous role in tumor pathophysiology, and different studies have attributed pro-tumorigenic properties to this receptor. In a previous work, we showed that CB2 expression is a poor prognostic factor in colon cancer patients. Here we report that activation of CB2 with low doses of specific agonists induce cell proliferation and favor the acquisition of aggressive molecular features in colon cancer cells. We show that sub-micromolar concentrations of CB2-specific agonists, JWH-133 and HU-308, promote an increase in cell proliferation rate through the activation of AKT/PKB pathway in colon cancer in vitro and in vivo. AKT activation promotes GSK3β inhibition and thus, a more aggressive cell phenotype with the subsequent elevation of SNAIL levels, E-cadherin degradation and β-catenin delocalization from cell membrane. Taken together, our data show that CB2 activation with sub-micromolar doses of agonists, which could be more similar to endogenous levels of cannabinoids, promote colon cancer progression, implicating that CB2 could have a pro-tumorigenic endogenous role in colon cancer. PMID:27634891

  7. Membrane Heterogeneity in Akt Activation in Prostate Cancer

    DTIC Science & Technology

    2009-11-01

    construct from the top, Fig. 1), proved to be incompatible with Akt1 activity. The prenylation signal derived from K- Ras rendered Akt1 catalytically...with either cyclo- dextrin (CD) or water-soluble cholesterol ( Chol ) or with CD followed by cholesterol treatment (CD Chol ). Cells incubated in serum

  8. Membrane Heterogeneity in Akt Activation in Prostate Cancer

    DTIC Science & Technology

    2009-07-01

    variant (third construct from the top, Fig. 1), proved to be incompatible with Akt1 activity. The prenylation signal derived from K- Ras rendered Akt1...water-soluble cholesterol ( Chol ) or with CD followed by cholesterol treatment (CD Chol ). Cells incubated in serum-free medium served as controls

  9. Andrographolide Inhibits Nuclear Factor-κB Activation through JNK-Akt-p65 Signaling Cascade in Tumor Necrosis Factor-α-Stimulated Vascular Smooth Muscle Cells

    PubMed Central

    Chen, Yu-Ying; Hsieh, Cheng-Ying; Lee, Lin-Wen; Sheu, Joen-Rong

    2014-01-01

    Critical vascular inflammation leads to vascular dysfunction and cardiovascular diseases, including abdominal aortic aneurysms, hypertension, and atherosclerosis. Andrographolide is the most active and critical constituent isolated from the leaves of Andrographis paniculata, a herbal medicine widely used for treating anti-inflammation in Asia. In this study, we investigated the mechanisms of the inhibitory effects of andrographolide in vascular smooth muscle cells (VSMCs) exposed to a proinflammatory stimulus, tumor necrosis factor-α (TNF-α). Treating TNF-α-stimulated VSMCs with andrographolide suppressed the expression of inducible nitric oxide synthase in a concentration-dependent manner. A reduction in TNF-α-induced c-Jun N-terminal kinase (JNK), Akt, and p65 phosphorylation was observed in andrographolide-treated VSMCs. However, andrographolide affected neither IκBα degradation nor p38 mitogen-activated protein kinase or extracellular signal-regulated kinase 1/2 phosphorylation under these conditions. Both treatment with LY294002, a phosphatidylinositol 3-kinase/Akt inhibitor, and treatment with SP600125, a JNK inhibitor, markedly reversed the andrographolide-mediated inhibition of p65 phosphorylation. In addition, LY294002 and SP600125 both diminished Akt phosphorylation, whereas LY294002 had no effects on JNK phosphorylation. These results collectively suggest that therapeutic interventions using andrographolide can benefit the treatment of vascular inflammatory diseases, and andrographolide-mediated inhibition of NF-κB activity in TNF-α-stimulated VSMCs occurs through the JNK-Akt-p65 signaling cascade, an IκBα-independent mechanism. PMID:25114952

  10. Desmocollin 3 mediates follicle stimulating hormone-induced ovarian epithelial cancer cell proliferation by activating the EGFR/Akt signaling pathway.

    PubMed

    Yang, Xiao; Wang, Jing; Li, Wen-Ping; Jin, Zhi-Jun; Liu, Xiao-Jun

    2015-01-01

    Follicle-stimulating hormone (FSH) is associated with the pathogenesis of ovarian cancer. We sought to explore whether desmocollin 3 (Dsc3) mediates FSH-induced ovarian epithelial cancer cell proliferation and whether the EGFR/Akt signaling pathway may be involved in this process. Dsc3 positivity in ovarian tissue specimens from 72 patients was assessed by immunohistochemistry. The positive expression rates of Dsc3 were similar in ovarian cancer tissues (24/31:77.4%) and borderline ovarian tumor tissues (18/22:81.8%) (P>0.05), but were significantly higher in these cancerous tissues than in benign ovarian cyst tissues (3/19:15.8%) (P<0.05). Consistently, the expression of Dsc3 in four out of five ovarian cancer cells (HO8910, Skov3ip, Skov and Hey cells, but not ES-2 and in borderline ovarian MCV152 tumor cells was higher than in the immortalized ovarian epithelial cell line, Moody. FSH up-regulated the expression of Dsc3 and EGFR in a dose- and time-dependent manner. Furthermore, a converse relationship between the expression of Dsc3, EFGR and PI3K/Akt signaling was elucidated using RNA interference and PI3K/Akt inhibitor in the absence and presence of FSH. A role for these proteins in FSH-induced cell proliferation was verified, highlighting their interdependence in mediating ovarian cancer cell function. These results suggest that Dsc3 can mediate FSH-induced ovarian cancer cell proliferation by activating the EGFR/Akt signaling pathway.

  11. Jaceosidin, a natural flavone, promotes angiogenesis via activation of VEGFR2/FAK/PI3K/AKT/NF-κB signaling pathways in endothelial cells.

    PubMed

    Lee, Tae Hoon; Jung, Hana; Park, Keun Hyung; Bang, Myun Ho; Baek, Nam-In; Kim, Jiyoung

    2014-10-01

    Angiogenesis, the growth of new blood vessels from pre-existing vasculature, plays an important role in physiological and pathological processes such as embryonic development wound healing and revascularization of tissues after exposure to ischemia. We investigated the effects of jaceosidin, a main constituent of medicinal herbs of the genus Artemisia, on angiogenesis and signaling pathways in endothelial cells. Jaceosidin stimulated proliferation, migration and tubulogenesis of ECs as well as ex vivo sprouting from aorta rings, which are phenomena typical of angiogenesis. Jaceosidin activated vascular endothelial growth factor receptor 2 (VEGFR2, FLk-1/KDR) and angiogenic signaling molecules such as focal adhesion kinase, phosphatidylinositol 3-kinase, and its downstream target, the serine-threonine kinase AKTWe also demonstrated that jaceosidin activated the NF-κB-driven expression of a luciferase reporter gene and NF-κB binding to DNA. Jaceosidin-induced proliferation and migration of human umbilical vascular endothelial cells were strongly inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002 and NF-κB inhibitor BAY11-7082, indicating that the PI3K/AKT/NF-κB signaling pathway is involved in jaceosidin-induced angiogenesis. Our results suggest that jaceosidin stimulates angiogenesis by activating the VEGFR2/FAK/PI3K/AKT/NF-κB signaling pathway and that it may be useful in developing angiogenic agents to promote the growth of collateral blood vessels in ischemic tissues.

  12. Morphine mediates a proinflammatory phenotype via μ-opioid receptor-PKCɛ-Akt-ERK1/2 signaling pathway in activated microglial cells.

    PubMed

    Merighi, Stefania; Gessi, Stefania; Varani, Katia; Fazzi, Debora; Stefanelli, Angela; Borea, Pier Andrea

    2013-08-15

    Anti-nociceptive tolerance to opioids severely limits their clinical efficacy for the treatment of chronic pain syndromes. Glia has a central role in the development of morphine tolerance. Here, we characterized the receptor-proximal signaling events that link μ-opioid receptors to activation of Akt and ERKs in lipopolysaccharide (LPS)-stimulated murine microglial cells with the aim to define the molecular mechanism contributing to the ability of morphine to increase inflammatory mediators such as nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-1β and IL-6 in activated microglial cells. In particular, the role of PKCɛ isoform in μ-opioid-induced inflammatory response in microglia was investigated. The results indicate that morphine increases the LPS-induced expression and activation of PKCɛ and stimulates Akt pathway upstream of ERK1/2 and iNOS. Furthermore, we found that morphine enhanced the release of IL-1β, TNF-α, IL-6, and of NO via μ-opioid receptor-PKCɛ signaling pathway in activated microglial cells, mediating a proinflammatory phenotype in mouse microglial cells. Together, these data suggest that the modulation of μ-opioid receptor signaling on microglia through PKCɛ selective inhibition may provide a means to attenuate glial activation and, as a consequence, to treat opioid development of tolerance and dependence.

  13. Negative Immune Regulator TIPE2 Promotes M2 Macrophage Differentiation through the Activation of PI3K-AKT Signaling Pathway

    PubMed Central

    Geng, Wenwen; Chen, Youhai H.; Zhang, Cui

    2017-01-01

    Macrophages play important roles in the regulation of the innate and adaptive immune responses. Classically activated macrophages and alternatively activated macrophages are the two major forms of macrophages and have opposing functionalities. Tumor necrosis factor-α-induced protein 8–2 is expressed primarily by immune cells and negatively regulates type 1 innate and adaptive immune responses to maintain immune tolerance. While previous studies indicate that TIPE2 promotes M2 but inhibits M1 macrophage differentiation, the underlying molecular mechanism by which TIPE2 promotes M2 macrophage differentiation remains unclear. Our current study shows that TIPE2-deficient bone-marrow cells are defective in IL-4 induced M2 macrophage differentiation in vitro. Mechanistic studies revealed that TIPE2 promotes phosphoinositide metabolism and the activation of the down-stream AKT signaling pathway, which in turn leads to the expression of markers specific for M2 macrophages. In addition, our results showed that Tipe2-deficiency does not affect the activation of the JAK-STAT6 signaling pathway that also plays an important role during M2 macrophage differentiation. Taken together, these results indicate that TIPE2 promotes M2 macrophage differentiation through the activation of PI3K-AKT signaling pathway, and may play an important role during the resolution of inflammation, parasite control, as well as tissue repair. PMID:28122045

  14. Pseudolaric acid B exerts antitumor activity via suppression of the Akt signaling pathway in HeLa cervical cancer cells.

    PubMed

    Li, Mingqun; Hong, Li

    2015-08-01

    Pseudolaric acid B (PAB) is a diterpene acid isolated from the bark of the root and trunk of Pseudolarix kaempferi Gordon (Pinaceae), which has demonstrated cytotoxic effects against various types of cancer. However, the mechanisms underlying the anticancer effects of PAB have remained to be elucidated. In the present study, the effects of PAB on the viability and apoptosis of HeLa cells were investigated by MTT assay, flow cytometric analysis of Annexin V-fluorescein isothiocyanate/propidium iodide staining, Rhodamine 123 staining and western blot analysis. The results demonstrated that PAB had antiproliferative and apoptosis-inducing effects on HeLa cells. PAB markedly inhibited HeLa cell viability in a time- and concentration-dependent manner. Flow cytometric analysis indicated that PAB induced apoptosis in HeLa cells in a dose-dependent manner. Treatment with PAB suppressed the expression of anti-apoptotic factor B cell lymphoma-2, and promoted the expression of pro-apoptotic factor Bcl-2-associated X protein. In addition, PAB induced an increase in Caspase-3 activity and loss of mitochondrial membrane potential, suggesting that this apoptosis may be mediated by mitochondrial pathways. Furthermore, the results of western blot analysis indicated that PAB was able to reduce Akt phosphorylation, thereby inhibiting the Akt pathway. These results suggested that PAB inhibited cell proliferation and induced apoptosis in HeLa cells, and that the anti-tumor effects of PAB were associated with inhibition of the Akt pathway. In conclusion, the results of the present study suggested that PAB may represent a novel therapeutic strategy for the treatment of human cervical cancer. However, additional studies are required to investigate the underlying apoptotic mechanisms.

  15. Rumex acetosa L. induces vasorelaxation in rat aorta via activation of PI3-kinase/Akt- AND Ca(2+)-eNOS-NO signaling in endothelial cells.

    PubMed

    Sun, Y Y; Su, X H; Jin, J Y; Zhou, Z Q; Sun, S S; Wen, J F; Kang, D G; Lee, H S; Cho, K W; Jin, S N

    2015-12-01

    Rumex acetosa L. (RA) (Polygonaceae) is an important traditional Chinese medicine (TCM) commonly used in clinic for a long history in China and the aerial parts of RA has a wide variety of pharmacological actions such as diuretic, anti-hypertensive, anti-oxidative, and anti-cancer effects. However, the mechanisms involved are to be defined. The purpose of the present study was to evaluate the vasorelaxant effect and define the mechanism of action of the ethanol extract of Rumex acetosa L. (ERA) in rat aorta. ERA was examined for its vascular relaxant effect in isolated phenylephrine-precontracted rat thoracic aorta and its acute effects on arterial blood pressure. In addition, the roles of the nitric oxide synthase (NOS)-nitric oxide (NO) signaling in the ERA-induced effects were tested in human umbilical vein endothelial cells (HUVECs). The phosphorylation levels of Akt and eNOS were assessed by Western blot analysis in the cultured HUVECs. ERA induced endothelium-dependent vasorelaxation. The ERA-induced vasorelaxation was abolished by L-NAME (an NOS inhibitor) or ODQ (a sGC inhibitor), but not by indomethacin. Inhibition of PI3-kinase/Akt signaling pathway markedly reduced the ERA-induced vasorelaxation. In HUVECs, ERA increased NO formation in a dose-dependent manner, which was inhibited by L-NAME and by removing extracellular Ca(2+). In addition, ERA promoted phosphorylation of Akt and eNOS, which was prevented by wortmannin and LY294002, indicating that ERA induces eNOS phosphorylation through the PI3-kinase/Akt pathway. Further, in anesthetized rats, intravenously administered ERA decreased arterial blood pressure in a dose-dependent manner through an activation of the NOS-NO system. In summary, the ERA- induced vasorelaxation was dependent on endothelial integrity and NO production, and was mediated by activation of both the endothelial PI3-kinase/Akt- and Ca(2+)-eNOS-NO signaling and muscular NO-sGC-cGMP signaling.

  16. The sonic hedgehog signaling pathway stimulates anaplastic thyroid cancer cell motility and invasiveness by activating Akt and c-Met.

    PubMed

    Williamson, Ashley J; Doscas, Michelle E; Ye, Jin; Heiden, Katherine B; Xing, Mingzhao; Li, Yi; Prinz, Richard A; Xu, Xiulong

    2016-03-01

    The sonic hedgehog (Shh) pathway is highly activated in thyroid neoplasms and promotes thyroid cancer stem-like cell phenotype, but whether the Shh pathway regulates thyroid tumor cell motility and invasiveness remains unknown. Here, we report that the motility and invasiveness of two anaplastic thyroid tumor cell lines, KAT-18 and SW1736, were inhibited by two inhibitors of the Shh pathway (cyclopamine and GANT61). Consistently, the cell motility and invasiveness was decreased by Shh and Gli1 knockdown, and was increased by Gli1 overexpression in KAT-18 cells. Mechanistic studies revealed that Akt and c-Met phosphorylation was decreased by a Gli1 inhibitor and by Shh and Gli1 knockdown, but was increased by Gli1 overexpression. LY294002, a PI-3 kinase inhibitor, and a c-Met inhibitor inhibited the motility and invasiveness of Gli1-transfected KAT-18 cells more effectively than the vector-transfected cells. Knockdown of Snail, a transcription factor regulated by the Shh pathway, led to decreased cell motility and invasiveness in KAT-18 and SW1736 cells. However, key epithelial-to-mesenchymal transition (EMT) markers including E-cadherin and vimentin as well as Slug were not affected by cyclopamine and GANT61 in either SW1736 or WRO82, a well differentiated follicular thyroid carcinoma cell line. Our data suggest that the Shh pathway-stimulated thyroid tumor cell motility and invasiveness is largely mediated by AKT and c-Met activation with little involvement of EMT.

  17. Notch-1 signaling activates NF-κB in human breast carcinoma MDA-MB-231 cells via PP2A-dependent AKT pathway.

    PubMed

    Li, Li; Zhang, Jing; Xiong, Niya; Li, Shun; Chen, Yu; Yang, Hong; Wu, Chunhui; Zeng, Hongjuan; Liu, Yiyao

    2016-04-01

    Breast cancer has a high incidence in the world and is becoming a leading cause of death in female patients due to its high metastatic ability. High expression of Notch-1 and its ligand Jagged-1 correlates with poor prognosis in breast cancer. Our previous work has shown that Notch-1 signaling pathway upregulates NF-κB transcriptional activity and induces the adhesion, migration and invasion of human breast cancer cell line MDA-MB-231. However, the role of Notch-1 in NF-κB activation is still poorly understood. Here, we aim to understand the exact mechanism that Notch-1 regulates NF-κB activity. In MDA-MB-231 cells where Notch-1 is constitutively activated, the phosphorylation of p85 and AKT (Tyr308/Ser473) is upregulated, indicating PI3K/AKT pathway is activated. Notch-1 activation caused the increase of PP2A phosphorylation at Tyr307, indicating Notch-1 inhibits PP2A activity. NF-κB transcriptional activity was evaluated by dual-luciferase reporter assay, and the results showed that, while silencing of Notch-1, PP2A activity was upregulated and NF-κB activity was downregulated, whereas PP2A inhibitor okadaic acid (OA) restored NF-κB activity. Immunofluorescence and Western blots showed that OA treatment antagonized the decrease of p65 nuclear translocation caused by Notch-1 silencing. Moreover, OA treatment also upregulated MMP-2, MMP-9 and VEGF mRNA expression levels, indicating OA rescues Notch-1 silencing that caused low cell invasion. Taken together, our results suggest that Notch-1-activating PI3K/AKT/NF-κB pathway is PP2A dependent; PP2A may be a potential therapeutic target in breast cancer.

  18. Protein O-fucosyltransferase 1 promotes trophoblast cell proliferation through activation of MAPK and PI3K/Akt signaling pathways.

    PubMed

    Liu, Chang; Liang, Xiaohua; Wang, Jiao; Zheng, Qin; Zhao, Yue; Khan, Muhammad Noman; Liu, Shuai; Yan, Qiu

    2017-04-01

    Protein O-fucosylation is an important glycosylation modification and plays an important role in embryonic development. Protein O-fucosyltransferase 1 (poFUT1) is an essential enzyme that catalyzes the synthesis of protein O-fucosylation. Our previous studies showed that poFUT1 promoted trophoblast cell migration and invasion at the fetal-maternal interface, but the role of poFUT1 in trophoblast cells proliferation remains unclear. Here, immunohistochemistry data showed that poFUT1 and PCNA levels were decreased in abortion patient's trophoblasts compared with women with normal pregnancies. Our results also showed that poFUT1 promoted trophoblast cell proliferation by CCK-8 assay and cell cycle analysis. PoFUT1 increased the phosphorylation of ERK1/2, p38 MAPK, and PI3K/Akt, while inhibitors of ERK1/2(PD98059), p38 MAPK(SB203580), and PI3K (LY294002) prevented ERK1/2, p38 MAPK, and Akt phosphorylation. Moreover, poFUT1 stimulation of trophoblast cells proliferation correlated with increased cell cycle progression by promoting cells into S-phase. The underlying mechanism involved increased cyclin D1, cyclin E, CDK 2, CDK 4, and pRb expression and decreased levels of the cyclin-dependent kinase inhibitors p21 and p27, which were blocked by inhibitors of the upstream signaling molecules MAPK and PI3K/Akt. In conclusion, poFUT1 promotes trophoblast cell proliferation by activating MAPK and PI3K/Akt signaling pathways.

  19. HO-1 attenuates hippocampal neurons injury via the activation of BDNF-TrkB-PI3K/Akt signaling pathway in stroke.

    PubMed

    Qi, Dashi; Ouyang, Changjie; Wang, Yulan; Zhang, Shichun; Ma, Xijuan; Song, YuanJian; Yu, HongLi; Tang, Jiali; Fu, Wei; Sheng, Lei; Yang, Lihua; Wang, Mei; Zhang, Weihao; Miao, Lei; Li, Tengteng; Huang, Xiaojing; Dong, Hongyan

    2014-08-19

    Although recent studies have found that HO-1 plays an important role in neuronal survival, little is known about the precise mechanisms occurring during cerebral ischemia/reperfusion (I/R). Therefore, the aim of this study was to investigate the neuroprotective mechanisms of HO-1 against ischemic brain injury induced by cerebral I/R and to explore whether the BDNF-TrkB-PI3K/Akt signaling pathway contributed to the protection provided by HO-1. Over-expressed HO-1 plasmids were employed to induce the overexpression of HO-1 through hippocampi CA1 injection 5 days before the cerebral I/R animal model was induced by four-vessel occlusion for 15 min transient ischemia and followed by reperfusion in Sprague-Dawley rats. Immunoblotting was carried out to examine the expression of the related proteins, and HE-staining was used to detect the percentage of living neurons in the hippocampal CA1 region. The results showed that over-expressed HO-1 could significantly protect neurons against cerebral I/R. Furthermore, the protein expression of BDNF, TrkB and p-Akt also increased in the rats treated with over-expressed HO-1 plasmids. However, treatment with tropomyosin receptor kinase B (TrkB) receptor antagonist (K252a) reversed the HO-1-induced increase in BDNF and p-Akt protein levels and decreased the level of cleaved caspase-3 protein in I/R rats. In summary, our results imply that HO-1 can decrease cell apoptosis in the I/R rat brain and that the mechanism may be related to the activation of the BDNF-TrkB-PI3K/Akt signaling pathway.

  20. Id-1 promotes tumorigenicity and metastasis of human esophageal cancer cells through activation of PI3K/AKT signaling pathway.

    PubMed

    Li, Bin; Tsao, Sai Wah; Li, Yuk Yin; Wang, Xianghong; Ling, Ming Tat; Wong, Yong Chuan; He, Qing Yu; Cheung, Annie L M

    2009-12-01

    Id-1 (inhibitor of differentiation or DNA binding) is a helix-loop-helix protein that is overexpressed in many types of cancer including esophageal squamous cell carcinoma (ESCC). We previously reported that ectopic Id-1 expression activates the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) signaling pathway in human esophageal cancer cells. In this study, we confirmed a positive correlation between Id-1 and phospho-AKT (Ser473) expressions in ESCC cell lines, as well as in ESCC on a tissue microarray. To investigate the significance of Id-1 in esophageal cancer progression, ESCC cells with stable ectopic Id-1 expression were inoculated subcutaneously into the flank of nude mice and were found to form larger tumors that showed elevated Ki-67 proliferation index and increased angiogenesis, as well as reduced apoptosis, compared with control cells expressing the empty vector.The Id-1-overexpressing cells also exhibited enhanced metastatic potential in the experimental metastasis assay. Treatment with the PI3K inhibitor LY294002 attenuated the tumor promotion effects of Id-1, indicating that the effects were mediated by the PI3K/AKT signaling pathway. In addition, our in vitro experiments showed that ectopic Id-1 expression altered the expression levels of markers associated with epithelial-mesenchymal transition and enhanced the migration ability of esophageal cancer cells. The Id-1-overexpressing ESCC cells also exhibited increased invasive potential, which was in part due to PI3K/AKT-dependent modulation of matrix metalloproteinase-9 expression. In conclusion, our results provide the first evidence that Id-1 promotes tumorigenicity and metastasis of human esophageal cancer in vivo and that the PI3K inhibitor LY294002 can attenuate these effects.

  1. Holo-APP and G-protein-mediated signaling are required for sAPPα-induced activation of the Akt survival pathway

    PubMed Central

    Milosch, N; Tanriöver, G; Kundu, A; Rami, A; François, J-C; Baumkötter, F; Weyer, S W; Samanta, A; Jäschke, A; Brod, F; Buchholz, C J; Kins, S; Behl, C; Müller, U C; Kögel, D

    2014-01-01

    Accumulating evidence indicates that loss of physiologic amyloid precursor protein (APP) function leads to reduced neuronal plasticity, diminished synaptic signaling and enhanced susceptibility of neurons to cellular stress during brain aging. Here we investigated the neuroprotective function of the soluble APP ectodomain sAPPα (soluble APPα), which is generated by cleavage of APP by α-secretase along the non-amyloidogenic pathway. Recombinant sAPPα protected primary hippocampal neurons and SH-SY5Y neuroblastoma cells from cell death induced by trophic factor deprivation. We show that this protective effect is abrogated in neurons from APP-knockout animals and APP-depleted SH-SY5Y cells, but not in APP-like protein 1- and 2- (APLP1 and APLP2) depleted cells, indicating that expression of membrane-bound holo-APP is required for sAPPα-dependent neuroprotection. Trophic factor deprivation diminished the activity of the Akt survival pathway. Strikingly, both recombinant sAPPα and the APP-E1 domain were able to stimulate Akt activity in wild-type (wt) fibroblasts, SH-SY5Y cells and neurons, but failed to rescue in APP-deficient neurons or fibroblasts. The ADAM10 (a disintegrin and metalloproteinase domain-containing protein 10) inhibitor GI254023X exacerbated neuron death in organotypic (hippocampal) slice cultures of wt mice subjected to trophic factor and glucose deprivation. This cell death-enhancing effect of GI254023X could be completely rescued by applying exogenous sAPPα. Interestingly, sAPPα-dependent Akt induction was unaffected in neurons of APP-ΔCT15 mice that lack the C-terminal YENPTY motif of the APP intracellular region. In contrast, sAPPα-dependent rescue of Akt activation was completely abolished in APP mutant cells lacking the G-protein interaction motif located in the APP C-terminus and by blocking G-protein-dependent signaling with pertussis toxin. Collectively, our data provide new mechanistic insights into the physiologic role of APP in

  2. Holo-APP and G-protein-mediated signaling are required for sAPPα-induced activation of the Akt survival pathway.

    PubMed

    Milosch, N; Tanriöver, G; Kundu, A; Rami, A; François, J-C; Baumkötter, F; Weyer, S W; Samanta, A; Jäschke, A; Brod, F; Buchholz, C J; Kins, S; Behl, C; Müller, U C; Kögel, D

    2014-08-28

    Accumulating evidence indicates that loss of physiologic amyloid precursor protein (APP) function leads to reduced neuronal plasticity, diminished synaptic signaling and enhanced susceptibility of neurons to cellular stress during brain aging. Here we investigated the neuroprotective function of the soluble APP ectodomain sAPPα (soluble APPα), which is generated by cleavage of APP by α-secretase along the non-amyloidogenic pathway. Recombinant sAPPα protected primary hippocampal neurons and SH-SY5Y neuroblastoma cells from cell death induced by trophic factor deprivation. We show that this protective effect is abrogated in neurons from APP-knockout animals and APP-depleted SH-SY5Y cells, but not in APP-like protein 1- and 2- (APLP1 and APLP2) depleted cells, indicating that expression of membrane-bound holo-APP is required for sAPPα-dependent neuroprotection. Trophic factor deprivation diminished the activity of the Akt survival pathway. Strikingly, both recombinant sAPPα and the APP-E1 domain were able to stimulate Akt activity in wild-type (wt) fibroblasts, SH-SY5Y cells and neurons, but failed to rescue in APP-deficient neurons or fibroblasts. The ADAM10 (a disintegrin and metalloproteinase domain-containing protein 10) inhibitor GI254023X exacerbated neuron death in organotypic (hippocampal) slice cultures of wt mice subjected to trophic factor and glucose deprivation. This cell death-enhancing effect of GI254023X could be completely rescued by applying exogenous sAPPα. Interestingly, sAPPα-dependent Akt induction was unaffected in neurons of APP-ΔCT15 mice that lack the C-terminal YENPTY motif of the APP intracellular region. In contrast, sAPPα-dependent rescue of Akt activation was completely abolished in APP mutant cells lacking the G-protein interaction motif located in the APP C-terminus and by blocking G-protein-dependent signaling with pertussis toxin. Collectively, our data provide new mechanistic insights into the physiologic role of APP in

  3. G protein-coupled receptors (GPCRs) That Signal via Protein Kinase A (PKA) Cross-talk at Insulin Receptor Substrate 1 (IRS1) to Activate the phosphatidylinositol 3-kinase (PI3K)/AKT Pathway.

    PubMed

    Law, Nathan C; White, Morris F; Hunzicker-Dunn, Mary E

    2016-12-30

    G protein-coupled receptors (GPCRs) activate PI3K/v-AKT thymoma viral oncoprotein (AKT) to regulate many cellular functions that promote cell survival, proliferation, and growth. However, the mechanism by which GPCRs activate PI3K/AKT remains poorly understood. We used ovarian preantral granulosa cells (GCs) to elucidate the mechanism by which the GPCR agonist FSH via PKA activates the PI3K/AKT cascade. Insulin-like growth factor 1 (IGF1) is secreted in an autocrine/paracrine manner by GCs and activates the IGF1 receptor (IGF1R) but, in the absence of FSH, fails to stimulate YXXM phosphorylation of IRS1 (insulin receptor substrate 1) required for PI3K/AKT activation. We show that PKA directly phosphorylates the protein phosphatase 1 (PP1) regulatory subunit myosin phosphatase targeting subunit 1 (MYPT1) to activate PP1 associated with the IGF1R-IRS1 complex. Activated PP1 is sufficient to dephosphorylate at least four IRS1 Ser residues, Ser(318), Ser(346), Ser(612), and Ser(789), and promotes IRS1 YXXM phosphorylation by the IGF1R to activate the PI3K/AKT cascade. Additional experiments indicate that this mechanism also occurs in breast cancer, thyroid, and preovulatory granulosa cells, suggesting that the PKA-dependent dephosphorylation of IRS1 Ser/Thr residues is a conserved mechanism by which GPCRs signal to activate the PI3K/AKT pathway downstream of the IGF1R.

  4. Jujuboside A Protects H9C2 Cells from Isoproterenol-Induced Injury via Activating PI3K/Akt/mTOR Signaling Pathway

    PubMed Central

    Han, Dandan; Wan, Changrong; Liu, Fenghua; Xu, Xiaolong; Jiang, Linshu; Xu, Jianqin

    2016-01-01

    Jujuboside A is a kind of the saponins isolated from the seeds of Ziziphus jujuba, which possesses multiple biological effects, such as antianxiety, antioxidant, and anti-inflammatory effects; however, its mediatory effect on isoproterenol-stimulated cardiomyocytes has not been investigated yet. In this study, we tried to detect the protective effect and potential mechanism of JUA on ISO-induced cardiomyocytes injury. H9C2 cells were treated with ISO to induce cell damage. Cells were pretreated with JUA to investigate the effects on the cell viability, morphological changes, light chain 3 conversion, and the activation of PI3K/Akt/mTOR signaling pathway. Results showed that ISO significantly inhibited the cell viability in a time- and dose-dependent manner. JUA pretreatment could reverse the reduction of cell viability and better the injury of H9C2 cells induced by ISO. Western blot analysis showed that JUA could accelerate the phosphorylation of PI3K, Akt, and mTOR. Results also indicated that JUA could significantly decrease the ratio of microtubule-associated protein LC3-II/I in H9C2 cells. Taken together, our research showed that JUA could notably reduce the damage cause by ISO via promoting the phosphorylation of PI3K, Akt, and mTOR and inhibiting LC3 conversion, which may be a potential choice for the treatment of heart diseases. PMID:27293469

  5. PTEN mutations and activation of the PI3K/Akt/mTOR signaling pathway in papillary tumors of the pineal region.

    PubMed

    Goschzik, Tobias; Gessi, Marco; Denkhaus, Dorota; Pietsch, Torsten

    2014-08-01

    Papillary tumors of the pineal region (PTPR) are recognized as a distinct entity in the World Health Organization classification of CNS tumors. Papillary tumors of the pineal region frequently show loss of chromosome 10, but no studies have investigated possible target genes on this chromosome. Chromosome 10 harbors the PTEN (phosphatase and tensin homolog) gene, the inactivation of which, by mutation or epigenetic silencing, has been observed in different brain tumors, including high-grade gliomas. In this study, we investigated copy number changes by molecular inversion probe (MIP) analysis and the mutational status of PTEN in 13 PTPR by direct sequencing. MIP analysis of 5 PTPR showed chromosome 10 loss in all cases. In addition, there were losses of chromosomes 3, 14, 22, and X, and gains of whole chromosomes 8, 9, and 12 in more than 1 case. One case had a homozygous PTEN deletion; and 2 point mutations in exon 7 of PTEN (G251D and Q261stop) were found. Immunohistochemistry revealed decrease or loss of the PTEN protein and increased expression of p-Akt and p-S6. These results indicated that PTEN mutations and activation of the PI3K/Akt/mTOR signaling pathway may play a role in the biology of PTPR. This evidence may lead to the possible use of PI3K/Akt/mTOR inhibitors in therapy for patients with PTPR.

  6. Fisetin inhibits UVB-induced cutaneous inflammation and activation of PI3K/AKT/NFκB signaling pathways in SKH-1 hairless mice.

    PubMed

    Pal, Harish Chandra; Athar, Mohammad; Elmets, Craig A; Afaq, Farrukh

    2015-01-01

    Solar ultraviolet B (UVB) radiation has been shown to induce inflammation, DNA damage, p53 mutations and alterations in signaling pathways eventually leading to skin cancer. In this study, we investigated whether fisetin reduces inflammatory responses and modulates PI3K/AKT/NFκB cell survival signaling pathways in UVB-exposed SKH-1 hairless mouse skin. Mice were exposed to 180 mJ cm(-2) of UVB radiation on alternate days for a total of seven exposures, and fisetin (250 and 500 nmol) was applied topically after 15 min of each UVB exposure. Fisetin treatment to UVB-exposed mice resulted in decreased hyperplasia and reduced infiltration of inflammatory cells. Fisetin treatment also reduced inflammatory mediators such as COX-2, PGE2 as well as its receptors (EP1-EP4) and MPO activity. Furthermore, fisetin reduced the level of inflammatory cytokines TNFα, IL-1β and IL-6 in UVB-exposed skin. Fisetin treatment also reduced cell proliferation markers as well as DNA damage as evidenced by increased expression of p53 and p21 proteins. Further studies revealed that fisetin inhibited UVB-induced expression of PI3K, phosphorylation of AKT and activation of the NFκB signaling pathway in mouse skin. Overall, these data suggest that fisetin may be useful against UVB-induced cutaneous inflammation and DNA damage.

  7. Activation of focal adhesion kinase by Salmonella suppresses autophagy via an Akt/mTOR signaling pathway and promotes bacterial survival in macrophages.

    PubMed

    Owen, Katherine A; Meyer, Corey B; Bouton, Amy H; Casanova, James E

    2014-06-01

    Autophagy has emerged as an important antimicrobial host defense mechanism that not only orchestrates the systemic immune response, but also functions in a cell autonomous manner to directly eliminate invading pathogens. Pathogenic bacteria such as Salmonella have evolved adaptations to protect themselves from autophagic elimination. Here we show that signaling through the non-receptor tyrosine kinase focal adhesion kinase (FAK) is actively manipulated by the Salmonella SPI-2 system in macrophages to promote intracellular survival. In wild-type macrophages, FAK is recruited to the surface of the Salmonella-containing vacuole (SCV), leading to amplified signaling through the Akt-mTOR axis and inhibition of the autophagic response. In FAK-deficient macrophages, Akt/mTOR signaling is attenuated and autophagic capture of intracellular bacteria is enhanced, resulting in reduced bacterial survival. We further demonstrate that enhanced autophagy in FAK(-/-) macrophages requires the activity of Atg5 and ULK1 in a process that is distinct from LC3-assisted phagocytosis (LAP). In vivo, selective knockout of FAK in macrophages resulted in more rapid clearance of bacteria from tissues after oral infection with S. typhimurium. Clearance was correlated with reduced infiltration of inflammatory cell types into infected tissues and reduced tissue damage. Together, these data demonstrate that FAK is specifically targeted by S. typhimurium as a novel means of suppressing autophagy in macrophages, thereby enhancing their intracellular survival.

  8. Cellular context–mediated Akt dynamics regulates MAP kinase signaling thresholds during angiogenesis

    PubMed Central

    Hellesøy, Monica; Lorens, James B.

    2015-01-01

    The formation of new blood vessels by sprouting angiogenesis is tightly regulated by contextual cues that affect angiogeneic growth factor signaling. Both constitutive activation and loss of Akt kinase activity in endothelial cells impair angiogenesis, suggesting that Akt dynamics mediates contextual microenvironmental regulation. We explored the temporal regulation of Akt in endothelial cells during formation of capillary-like networks induced by cell–cell contact with vascular smooth muscle cells (vSMCs) and vSMC-associated VEGF. Expression of constitutively active Akt1 strongly inhibited network formation, whereas hemiphosphorylated Akt1 epi-alleles with reduced kinase activity had an intermediate inhibitory effect. Conversely, inhibition of Akt signaling did not affect endothelial cell migration or morphogenesis in vSMC cocultures that generate capillary-like structures. We found that endothelial Akt activity is transiently blocked by proteasomal degradation in the presence of SMCs during the initial phase of capillary-like structure formation. Suppressed Akt activity corresponded to the increased endothelial MAP kinase signaling that was required for angiogenic endothelial morphogenesis. These results reveal a regulatory principle by which cellular context regulates Akt protein dynamics, which determines MAP kinase signaling thresholds necessary drive a morphogenetic program during angiogenesis. PMID:26023089

  9. Akt phosphorylates Tal1 oncoprotein and inhibits its repressor activity.

    PubMed

    Palamarchuk, Alexey; Efanov, Alexey; Maximov, Vadim; Aqeilan, Rami I; Croce, Carlo M; Pekarsky, Yuri

    2005-06-01

    The helix-loop-helix transcription factor Tal1 is required for blood cell development and its activation is a frequent event in T-cell acute lymphoblastic leukemia. The Akt (protein kinase B) kinase is a key player in transduction of antiapoptotic and proliferative signals in T cells. Because Tal1 has a putative Akt phosphorylation site at Thr90, we investigated whether Akt regulates Tal1. Our results show that Akt specifically phosphorylates Thr90 of the Tal1 protein within its transactivation domain in vitro and in vivo. Coimmunoprecipitation experiments showed the presence of Tal1 in Akt immune complexes, suggesting that Tal1 and Akt physically interact. We further showed that phosphorylation of Tal1 by Akt causes redistribution of Tal1 within the nucleus. Using luciferase assay, we showed that phosphorylation of Tal1 by Akt decreased repressor activity of Tal1 on EpB42 (P4.2) promoter. Thus, these data indicate that Akt interacts with Tal1 and regulates Tal1 by phosphorylation at Thr90 in a phosphatidylinositol 3-kinase-dependent manner.

  10. Anti-Osteoclastogenic Activity of Praeruptorin A via Inhibition of p38/Akt-c-Fos-NFATc1 Signaling and PLCγ-Independent Ca2+ Oscillation

    PubMed Central

    Choi, Sik-Won; Moon, Seong-Hee; Park, Young Sik; Ryu, Byung Jun; Oh, Jaemin; Kim, Min Seuk; Erkhembaatar, Munkhsoyol; Son, Young-Jin; Kim, Seong Hwan

    2014-01-01

    Background A decrease of bone mass is a major risk factor for fracture. Several natural products have traditionally been used as herbal medicines to prevent and/or treat bone disorders including osteoporosis. Praeruptorin A is isolated from the dry root extract of Peucedanum praeruptorum Dunn and has several biological activities, but its anti-osteoporotic activity has not been studied yet. Materials and Methods The effect of praeruptorin A on the differentiation of bone marrow–derived macrophages into osteoclasts was examined by phenotype assay and confirmed by real-time PCR and immunoblotting. The involvement of NFATc1 in the anti-osteoclastogenic action of praeruptorin A was evaluated by its lentiviral ectopic expression. Intracellular Ca2+ levels were also measured. Results Praeruptorin A inhibited the RANKL-stimulated osteoclast differentiation accompanied by inhibition of p38 and Akt signaling, which could be the reason for praeruptorin A-downregulated expression levels of c-Fos and NFATc1, transcription factors that regulate osteoclast-specific genes, as well as osteoclast fusion-related molecules. The anti-osteoclastogenic effect of praeruptorin A was rescued by overexpression of NFATc1. Praeruptorin A strongly prevented the RANKL-induced Ca2+ oscillation without any changes in the phosphorylation of PLCγ. Conclusion Praeruptorin A could exhibit its anti-osteoclastogenic activity by inhibiting p38/Akt-c-Fos-NFATc1 signaling and PLCγ-independent Ca2+ oscillation. PMID:24586466

  11. Inhibition of c-Met activation sensitizes osteosarcoma cells to cisplatin via suppression of the PI3K-Akt signaling.

    PubMed

    Wang, Kelai; Zhuang, Yan; Liu, Chunlan; Li, Yang

    2012-10-01

    Osteosarcoma is a common malignant bone tumor. Cisplatin (CDDP) achieves a high response rate in osteosarcoma. However, osteosarcoma usually exhibits cisplatin resistance. Many members of receptor tyrosine kinases (RTKs)(1) have been demonstrated to be overexpressed and constitutively activated in various tumors including osteosarcoma, resulting in malignant progression and insensitivity to chemotherapy. Hepatocyte growth factor receptor (HGFR/c-Met) also appears overexpressed and activated in osteosarcoma cells. Nevertheless, which role of c-Met activation in cisplatin efficacy against osteosarcoma cells remains still elusive. This study found that inhibition of c-Met activity by PHA-665752 or blockade of the interaction of autocrined HGF with c-Met with neutralizing anti-HGF antibody promoted cisplatin efficacy in osteosarcoma cells, while addition of recombinant human HGF (rh-HGF) counteracts cisplatin cytotoxicity. Specifically, we demonstrated that inhibition of c-Met activity led to suppression of the PI3K-Akt pathway, thus enhancing cisplatin chemosensitivity. Our study clearly suggests that inhibition of c-Met activity can effectively sensitize osteosarcoma cells to cisplatin via suppression of the PI3K-Akt signaling.

  12. Impaired translocation and activation of mitochondrial Akt1 mitigated mitochondrial oxidative phosphorylation Complex V activity in diabetic myocardium.

    PubMed

    Yang, Jia-Ying; Deng, Wu; Chen, Yumay; Fan, Weiwei; Baldwin, Kenneth M; Jope, Richard S; Wallace, Douglas C; Wang, Ping H

    2013-06-01

    Insulin can translocate Akt to mitochondria in cardiac muscle. The goals of this study were to define sub-mitochondrial localization of the translocated Akt, to dissect the effects of insulin on Akt isoform translocation, and to determine the direct effect of mitochondrial Akt activation on Complex V activity in normal and diabetic myocardium. The translocated Akt sequentially localized to the mitochondrial intermembrane space, inner membrane, and matrix. To confirm Akt translocation, in vitro import assay showed rapid entry of Akt into mitochondria. Akt isoforms were differentially regulated by insulin stimulation, only Akt1 translocated into mitochondria. In the insulin-resistant Type 2 diabetes model, Akt1 translocation was blunted. Mitochondrial activation of Akt1 increased Complex V activity by 24% in normal myocardium in vivo and restored Complex V activity in diabetic myocardium. Basal mitochondrial Complex V activity was lower by 22% in the Akt1(-/-) myocardium. Insulin-stimulated Complex V activity was not impaired in the Akt1(-/-) myocardium, due to compensatory translocation of Akt2 to mitochondria. Akt1 is the primary isoform that relayed insulin signaling to mitochondria and modulated mitochondrial Complex V activity. Activation of mitochondrial Akt1 enhanced ATP production and increased phosphocreatine in cardiac muscle cells. Dysregulation of this signal pathway might impair mitochondrial bioenergetics in diabetic myocardium.

  13. Flotillin-2 promotes metastasis of nasopharyngeal carcinoma by activating NF-κB and PI3K/Akt3 signaling pathways

    PubMed Central

    Liu, Jie; Huang, Wei; Ren, Caiping; Wen, Qiuyuan; Liu, Weidong; Yang, Xuyu; Wang, Lei; Zhu, Bin; Zeng, Liang; Feng, Xiangling; Zhang, Chang; Chen, Huan; Jia, Wei; Zhang, Lihua; Xia, Xiaomeng; Chen, Yuxiang

    2015-01-01

    Lipid raft proteins have been confirmed to be important in cell signal transduction. Some reports have shown that the aberrant expression of lipid raft proteins is associated with malignant phenotypes in some cancers. However, the role of the lipid raft protein flotillin-2 (Flot-2) in nasopharyngeal carcinoma (NPC) remains to be comprehensively characterized. Here, overexpression of Flot-2 in NPC tissues and cell lines was detected by immunostaining, and Flot-2 expression was found to be positively associated with NPC metastasis. Furthermore, inhibiting Flot-2 expression impaired the malignancy of the highly metastatic NPC cell line 5-8F by constraining its growth and proliferation, mobility and migration, and decreasing the capacity of 5-8F cells to metastasize in nude mice. In contrast, forced overexpression of Flot-2 increased the malignancy of 6-10B, a non-metastatic NPC cell line that weakly expresses Flot-2. Moreover, in 5-8F-shFlot-2 cells, which have inhibited Flot-2 expression, the NF-κB and PI3K/Akt3 pathways were inactivated. Subsequently, MMPs expression were decreased, and Foxo1 activity was increased. In addition, enhanced NF-κB and PI3K/Akt3 activities were observed in Flot-2 overexpressing 6-10B cells. Thus, Flot-2 exerts a pro-neoplastic role in NPC and is involved in tumor progression and metastasis. Moreover, Flot-2 exerts its role through NF-κB and PI3K/Akt3 signaling. PMID:26206082

  14. Ankyrin repeat and suppressor of cytokine signaling (SOCS) box-containing protein (ASB) 15 alters differentiation of mouse C2C12 myoblasts and phosphorylation of mitogen-activated protein kinase and Akt.

    PubMed

    McDaneld, T G; Spurlock, D M

    2008-11-01

    Ankyrin repeat and suppressor of cytokine signaling box-containing protein (ASB) 15 is a novel ASB gene family member predominantly expressed in skeletal muscle. We have previously reported that overexpression of ASB15 delays differentiation and alters protein turnover in mouse C(2)C(12) myoblasts. However, the extent of ASB15 regulation of differentiation and molecular pathways underlying this activity are unknown. The extracellular signal-regulated kinase (Erk) 1/2 and phosphatidylinositol-3 kinase-Akt (PI3K/Akt; Akt is also known as protein kinase B) signaling pathways have a role in skeletal muscle growth. Activation (phosphorylation) of the Erk1/2 signaling pathway promotes proliferation, whereas activation of the PI3K/Akt signaling pathway promotes myoblast differentiation. Accordingly, we tested the hypothesis that ASB15 controls myoblast differentiation through its regulation of these kinases. Stably transfected myoblasts overexpressing ASB15 (ASB15+) demonstrated decreased differentiation, whereas attenuation of ASB15 expression (ASB15-) increased differentiation. However, ASB15+ cells had less abundance of the phosphorylated mitogen-activated protein kinase (active) form, despite decreased differentiation relative to control myoblasts (ASB15Con). The mitogen-activated protein kinase kinase inhibitor, U0126, effectively decreased mitogen-activated protein kinase phosphorylation and stimulated differentiation in ASB15- and ASB15Con cells. However, inhibition of the Erk1/2 pathway was unable to overcome the inhibitory effect of overexpressing ASB15 on differentiation (ASB15+), suggesting that the Erk1/2 pathway is likely not the predominant mediator of ASB15 activity on differentiation. Expression of ASB15 also altered phosphorylation of the PI3K/Akt pathway, as ASB15+ and ASB15- cells had decreased and increased Akt phosphorylation, respectively. These data were consistent with observed differences in differentiation. Administration of IGF-I, a PI3K/Akt

  15. Raloxifene activates G protein-coupled estrogen receptor 1/Akt signaling to protect dopamine neurons in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine mice.

    PubMed

    Bourque, Mélanie; Morissette, Marc; Di Paolo, Thérèse

    2014-10-01

    Raloxifene, used in the clinic, is reported to protect brain dopaminergic neurons in mice. Raloxifene was shown to mediate an effect through the G protein-coupled estrogen receptor 1 (GPER1). We investigated if raloxifene neuroprotective effect in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated male mice is mediated through GPER1 by using its antagonist G15. Striatal concentrations of dopamine, 3,4-dihydroxyphenylacetic acid, homovanillic acid to dopamine ratio as well as dopamine transporter and vesicular monoamine transporter 2 showed that raloxifene neuroprotection of dopaminergic neurons was blocked by G15. Protection by raloxifene was accompanied by activation of striatal Akt signaling (but not ERK1/2 signaling) and increased Bcl-2 and brain-derived neurotrophic factor levels; these effects were abolished by coadministration with G15. The effect of raloxifene was not mediated through increased levels of 17β-estradiol. MPTP mice had decreased plasma testosterone, dihydrotestosterone, and 3β-diol levels; this was prevented in raloxifene-treated MPTP mice. Our results suggest that raloxifene acted through GPER1 to mediate Akt activation, increase Bcl-2 and brain-derived neurotrophic factor levels, and protection of dopaminergic neurons and plasma androgens.

  16. Berberine reduces ischemia/reperfusion-induced myocardial apoptosis via activating AMPK and PI3K-Akt signaling in diabetic rats.

    PubMed

    Chen, Keke; Li, Guohua; Geng, Fenghao; Zhang, Zhao; Li, Jiani; Yang, Min; Dong, Ling; Gao, Feng

    2014-06-01

    Diabetes increases the risk of cardiovascular diseases. Berberine (BBR), an isoquinoline alkaloid used in Chinese medicine, exerts anti-diabetic effect by lowering blood glucose and regulating lipid metabolism. It has been reported that BBR decreases mortality in patients with chronic congestive heart failure. However, the molecular mechanisms of these beneficial effects are incompletely understood. In the present study, we sought to determine whether BBR exerts cardioprotective effect against ischemia/reperfusion (I/R) injury in diabetic rats and the underlying mechanisms. Male Sprague-Dawley rats were injected with low dose streptozotocin and fed with a high-fat diet for 12 weeks to induce diabetes. The diabetic rats were intragastrically administered with saline or BBR (100, 200 and 400 mg/kg/d) starting from week 9 to 12. At the end of week 12, all rats were subjected to 30 min of myocardial ischemia and 3 h of reperfusion. BBR significantly improved the recovery of cardiac systolic/diastolic function and reduced myocardial apoptosis in diabetic rats subjected to myocardial I/R. Furthermore, in cultured neonatal rat cardiomyocytes, BBR (50 μmol/L) reduced hypoxia/reoxygenation-induced myocardial apoptosis, increased Bcl-2/Bax ratio and decreased caspase-3 expression, together with enhanced activation of PI3K-Akt and increased adenosine monophosphate-activated protein kinase (AMPK) and eNOS phosphorylation. Pretreatment with either PI3K/Akt inhibitor wortmannin or AMPK inhibitor Compound C blunted the anti-apoptotic effect of BBR. Our findings demonstrate that BBR exerts anti-apoptotic effect and improves cardiac functional recovery following myocardial I/R via activating AMPK and PI3K-Akt-eNOS signaling in diabetic rats.

  17. Notoginsenoside R1 ameliorates podocyte injury in rats with diabetic nephropathy by activating the PI3K/Akt signaling pathway

    PubMed Central

    Huang, Guodong; Lv, Jianzhen; Li, Tongyu; Huai, Guoli; Li, Xiang; Xiang, Shaowei; Wang, Longlong; Qin, Zhenlin; Pang, Jianli; Zou, Bingyu; Wang, Yi

    2016-01-01

    The present study was designed to examine the protective effect of notoginsenoside R1 (NR1) on podocytes in a rat model of streptozotocin (STZ)-induced diabetic nephropathy (DN), and to explore the mechanism responsible for NR1-induced renal protection. Diabetes was induced by a single injection of STZ, and NR1 was administered daily at a dose of 5 mg/kg (low dose), 10 mg/kg (medium) and 20 mg/kg (high) for 16 weeks in Sprague-Dawley rats. Blood glucose levels, body weight and proteinuria were measured every 4 weeks, starting on the day that the rats received NR1. Furthermore, on the day of sacrifice, blood, urine and kidneys were collected in order to assess renal function according to general parameters. Pathological staining was performed to evaluate the renal protective effect of NR1, and the expression of the key slit diaphragm proteins, namely neprhin, podocin and desmin, were evaluated. In addition, the serum levels of inflammatory cytokines [tumor necrosis factor-α (TNF-α), tumor growth factor-β1 (TGF-β1), interleukin (IL)-1 and IL-6] as well as an anti-inflammatory cytokine (IL-10) were assessed, and the apoptosis of podocytes was quantified. Finally, the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway and the involvement of nuclear factor-κB (NF-κB) inactivation was further analyzed. In this study, NR1 improved renal function by ameliorating histological alterations, increasing the expression of nephrin and podocin, decreasing the expression of desmin, and inhibiting both the inflammatory response as well as the apoptosis of podocytes. Furthermore, NR1 treatment increased the phosphorylation of both PI3K (p85) and Akt, indicating that activation of the PI3K/Akt signaling pathway was involved. Moreover, NR1 treatment decreased the phosphorylation of NF-κB (p65), suggesting the downregulation of NF-κB. This is the first study to the best of our knowledge, to clearly demonstrate that NR1 treatment ameliorates podocyte injury by inhibiting both

  18. EPO-dependent activation of PI3K/Akt/FoxO3a signalling mediates neuroprotection in in vitro and in vivo models of Parkinson's disease.

    PubMed

    Jia, Yu; Mo, Shi-Jing; Feng, Qi-Qi; Zhan, Ma-Li; OuYang, Li-Si; Chen, Jia-Chang; Ma, Yu-Xin; Wu, Jia-Jia; Lei, Wan-Long

    2014-05-01

    Erythropoietin (EPO) may become a potential therapeutic candidate for the treatment of the neurodegenerative disorder -- Parkinson's disease (PD), since EPO has been found to prevent neuron apoptosis through the activation of cell survival signalling. However, the underlying mechanisms of how EPO exerts its neuroprotective effect are not fully elucidated. Here we investigated the mechanism by which EPO suppressed 6-hydroxydopamine (6-OHDA)-induced neuron death in in vitro and in vivo models of PD. EPO knockdown conferred 6-OHDA-induced cytotoxicity. This effect was reversed by EPO administration. Treatment of PC12 cells with EPO greatly diminished the toxicity induced by 6-OHDA in a dose- and time-dependent manner. EPO effectively reduced apoptosis of striatal neurons and induced a significant improvement on the neurological function score in the rat models of PD. Furthermore, EPO increased the expression of phosphorylated Akt and phosphorylated FoxO3a, and abrogated the 6-OHDA-induced dysregulation of Bcl-2, Bax and Caspase-3 in PC12 cells and in striatal neurons. Meanwhile, the EPO-dependent neuroprotection was notably reversed by pretreatment with LY294002, a specific inhibitor of phosphatidylinositol 3-kinase (PI3K). Our data suggest that PI3K/Akt/FoxO3a signalling pathway may be a possible mechanism involved in the neuroprotective effect of EPO in PD.

  19. Ergosterol peroxide activates Foxo3-mediated cell death signaling by inhibiting AKT and c-Myc in human hepatocellular carcinoma cells

    PubMed Central

    Hu, Liming; Du, William W.; Jiao, Chunwei; Pan, Honghui; Sdiri, Mouna; Wu, Nan; Xie, Yizhen; Yang, Burton B.

    2016-01-01

    Sterols are the important active ingredients of fungal secondary metabolites to induce death of tumor cells. In our previous study, we found that ergosterol peroxide (5α, 8α-epidioxiergosta-6, 22-dien-3β-ol), purified from Ganoderma lucidum, induced human cancer cell death. Since the amount of purified ergosterol peroxide is not sufficient to perform in vivo experiments or apply clinically, we developed an approach to synthesize ergosterol peroxide chemically. After confirming the production of ergosterol peroxide, we examined the biological functions of the synthetic ergosterol peroxide. The results showed that ergosterol peroxide induced cell death and inhibited cell migration, cell cycle progression, and colony growth of human hepatocellular carcinoma cells. We further examined the mechanism associated with this effect and found that treatment with ergosterol peroxide increased the expression of Foxo3 mRNA and protein in HepG2 cells. The upstream signal proteins pAKT and c-Myc, which can inhibit Foxo3 functions, were clearly decreased in HepG2 cells treated with ergosterol peroxide. The levels of Puma and Bax, pro-apoptotic proteins, were effectively enhanced. Our results suggest that ergosterol peroxide stimulated Foxo3 activity by inhibiting pAKT and c-Myc and activating pro-apoptotic protein Puma and Bax to induce cancer cell death. PMID:27058618

  20. PDGFRα signaling in the primary cilium regulates NHE1-dependent fibroblast migration via coordinated differential activity of MEK1/2–ERK1/2–p90RSK and AKT signaling pathways

    PubMed Central

    Clement, Ditte L.; Mally, Sabine; Stock, Christian; Lethan, Mette; Satir, Peter; Schwab, Albrecht; Pedersen, Stine F.; Christensen, Søren T.

    2013-01-01

    Summary In fibroblasts, platelet-derived growth factor receptor alpha (PDGFRα) is upregulated during growth arrest and compartmentalized to the primary cilium. PDGF-AA mediated activation of the dimerized ciliary receptor produces a phosphorylation cascade through the PI3K–AKT and MEK1/2–ERK1/2 pathways leading to the activation of the Na+/H+ exchanger, NHE1, cytoplasmic alkalinization and actin nucleation at the lamellipodium that supports directional cell migration. We here show that AKT and MEK1/2–ERK1/2–p90RSK inhibition reduced PDGF-AA-induced cell migration by distinct mechanisms: AKT inhibition reduced NHE1 activity by blocking the translocation of NHE1 to the cell membrane. MEK1/2 inhibition did not affect NHE1 activity but influenced NHE1 localization, causing NHE1 to localize discontinuously in patches along the plasma membrane, rather than preferentially at the lamellipodium. We also provide direct evidence of NHE1 translocation through the cytoplasm to the leading edge. In conclusion, signals initiated at the primary cilium through the PDGFRαα cascade reorganize the cytoskeleton to regulate cell migration differentially through the AKT and the MEK1/2–ERK1/2–p90RSK pathways. The AKT pathway is necessary for initiation of NHE1 translocation, presumably in vesicles, to the leading edge and for its activation. In contrast, the MEK1/2–ERK1/2–p90RSK pathway controls the spatial organization of NHE1 translocation and incorporation, and therefore specifies the direction of the leading edge formation. PMID:23264740

  1. PDGFRα signaling in the primary cilium regulates NHE1-dependent fibroblast migration via coordinated differential activity of MEK1/2-ERK1/2-p90RSK and AKT signaling pathways.

    PubMed

    Clement, Ditte L; Mally, Sabine; Stock, Christian; Lethan, Mette; Satir, Peter; Schwab, Albrecht; Pedersen, Stine F; Christensen, Søren T

    2013-02-15

    In fibroblasts, platelet-derived growth factor receptor alpha (PDGFRα) is upregulated during growth arrest and compartmentalized to the primary cilium. PDGF-AA mediated activation of the dimerized ciliary receptor produces a phosphorylation cascade through the PI3K-AKT and MEK1/2-ERK1/2 pathways leading to the activation of the Na(+)/H(+) exchanger, NHE1, cytoplasmic alkalinization and actin nucleation at the lamellipodium that supports directional cell migration. We here show that AKT and MEK1/2-ERK1/2-p90(RSK) inhibition reduced PDGF-AA-induced cell migration by distinct mechanisms: AKT inhibition reduced NHE1 activity by blocking the translocation of NHE1 to the cell membrane. MEK1/2 inhibition did not affect NHE1 activity but influenced NHE1 localization, causing NHE1 to localize discontinuously in patches along the plasma membrane, rather than preferentially at the lamellipodium. We also provide direct evidence of NHE1 translocation through the cytoplasm to the leading edge. In conclusion, signals initiated at the primary cilium through the PDGFRαα cascade reorganize the cytoskeleton to regulate cell migration differentially through the AKT and the MEK1/2-ERK1/2-p90(RSK) pathways. The AKT pathway is necessary for initiation of NHE1 translocation, presumably in vesicles, to the leading edge and for its activation. In contrast, the MEK1/2-ERK1/2-p90(RSK) pathway controls the spatial organization of NHE1 translocation and incorporation, and therefore specifies the direction of the leading edge formation.

  2. The PI3K/Akt signal hyperactivates Eya1 via the SUMOylation pathway

    PubMed Central

    Sun, Ye; Kaneko, Satoshi; Li, Xiaokun; Li, Xue

    2014-01-01

    Eya1 is a conserved critical regulator of organ-specific stem cells. Ectopic Eya1 activities, however, promote transformation of mammary epithelial cells. Signals that instigate Eya1 oncogenic activities remain to be determined. Here, we show that Akt1 kinase physically interacts with Eya1 and phosphorylates a conserved consensus site of the Akt kinase. PI3K/Akt signaling enhances Eya1 transcription activity, which largely attributes to the phosphorylation-induced reduction of Eya1 SUMOylation. Indeed, SUMOylation inhibits Eya1 transcription activity; and pharmacologic and genetic activation of PI3K/Akt robustly reduces Eya1 SUMOylation. Wild type but not Akt phosphorylation site mutant Eya1 variant rescues the cell migratory phenotype of EYA1-silenced breast cancer cells, highlighting the importance of Eya1 phosphorylation. Furthermore, knockdown EYA1 sensitizes breast cancer cells to the PI3K/Akt1 inhibitor and irradiation treatments. Thus, the PI3K/Akt signal pathway activates Eya1. These findings further suggest that regulation of SUMOylation by PI3K/Akt signaling is likely an important aspect of tumorigenesis. PMID:24954506

  3. K6PC-5, a novel sphingosine kinase 1 (SphK1) activator, alleviates dexamethasone-induced damages to osteoblasts through activating SphK1-Akt signaling.

    PubMed

    Ji, Feng; Mao, Li; Liu, Yuanyuan; Cao, Xiaojian; Xie, Yue; Wang, Shouguo; Fei, Haodong

    2015-03-13

    Long-term glucocorticoid usage is a common cause of non-traumatic femoral head osteonecrosis. Glucocorticoids (i.e. dexamethasone (Dex)) could directly induce damages to osteoblasts. In the current study, we investigated the potential activity of K6PC-5 [N-(1,3-dihydroxyisopropyl)-2-hexyl-3-oxo-decanamide], a novel sphingosine kinase 1 (SphK1) activator, against this process. Our data revealed that both osteoblastic-like MC3T3-E1 cells and primary murine osteoblasts were responsible to K6PC-5. K6PC-5 activated SphK1, increased sphingosine-1-phosphate (S1P) production and induced Akt phosphorylation in cultured osteoblasts. Functionally, K6PC-5 protected osteoblasts from Dex-induced apoptosis and necrosis. Such signaling and functional effects by K6PC-5 were prevented by the SphK1 inhibitor N,N-dimethylsphingosine (DMS), and by SphK1-siRNAs. On the other hand, exogenously-added S1P activated Akt and reduced Dex-induced osteoblast damages. LY294002 and MK-2206, two established Akt inhibitors, alleviated K6PC-5- or S1P-mediated osteoblast protection against Dex. Together, our results suggest that K6PC-5 alleviates Dex-induced osteoblast injuries through activating SphK1-Akt signaling. K6PC-5 might be further investigated in animal or clinical studies for its anti-glucocorticoids-associated osteonecrosis potential.

  4. Roles of oxidative stress and Akt signaling in doxorubicin cardiotoxicity

    SciTech Connect

    Ichihara, Sahoko . E-mail: saho@gene.mie-u.ac.jp; Yamada, Yoshiji; Kawai, Yoshichika; Osawa, Toshihiko; Furuhashi, Koichi; Duan Zhiwen; Ichihara, Gaku

    2007-07-20

    Cardiotoxicity is a treatment-limiting side effect of the anticancer drug doxorubicin (DOX). We have now investigated the roles of oxidative stress and signaling by the protein kinase Akt in DOX-induced cardiotoxicity as well as the effects on such toxicity both of fenofibrate, an agonist of peroxisome proliferator-activated receptor-{alpha}, and of polyethylene glycol-conjugated superoxide dismutase (PEG-SOD), an antioxidant. Mice injected intraperitoneally with DOX were treated for 4 days with fenofibrate or PEG-SOD. Fenofibrate and PEG-SOD each prevented the induction of cardiac dysfunction by DOX. Both drugs also inhibited the activation of the transcription factor NF-{kappa}B and increase in lipid peroxidation in the left ventricle induced by DOX, whereas only PEG-SOD inhibited the DOX-induced activation of Akt and Akt-regulated gene expression. These results suggest that fenofibrate and PEG-SOD prevented cardiac dysfunction induced by DOX through normalization of oxidative stress and redox-regulated NF-{kappa}B signaling.

  5. Homocysteine enhances MMP-9 production in murine macrophages via ERK and Akt signaling pathways

    SciTech Connect

    Lee, Seung Jin; Lee, Yi Sle; Seo, Kyo Won; Bae, Jin Ung; Kim, Gyu Hee; Park, So Youn; Kim, Chi Dae

    2012-04-01

    Homocysteine (Hcy) at elevated levels is an independent risk factor of cardiovascular diseases, including atherosclerosis. In the present study, we investigated the effect of Hcy on the production of matrix metalloproteinases (MMP) in murine macrophages. Among the MMP known to regulate the activities of collagenase and gelatinase, Hcy exclusively increased the gelatinolytic activity of MMP-9 in J774A.1 cells as well as in mouse peritoneal macrophages. Furthermore, this activity was found to be correlated with Western blot findings in J774A.1 cells, which showed that MMP-9 expression was concentration- and time-dependently increased by Hcy. Inhibition of the ERK and Akt pathways led to a significant decrease in Hcy-induced MMP-9 expression, and combined treatment with inhibitors of the ERK and Akt pathways showed an additive effects. Activity assays for ERK and Akt showed that Hcy increased the phosphorylation of both, but these phosphorylation were not affected by inhibitors of the Akt and ERK pathways. In line with these findings, the molecular inhibition of ERK and Akt using siRNA did not affect the Hcy-induced phosphorylation of Akt and ERK, respectively. Taken together, these findings suggest that Hcy enhances MMP-9 production in murine macrophages by separately activating the ERK and Akt signaling pathways. -- Highlights: ► Homocysteine (Hcy) induced MMP-9 production in murine macrophages. ► Hcy induced MMP-9 production through ERK and Akt signaling pathways. ► ERK and Akt signaling pathways were activated by Hcy in murine macrophages. ► ERK and Akt pathways were additively act on Hcy-induced MMP-9 production. ► Hcy enhances MMP-9 production in macrophages via activation of ERK and Akt signaling pathways in an independent manner.

  6. mTOR Activation by PI3K/Akt and ERK Signaling in Short ELF-EMF Exposed Human Keratinocytes

    PubMed Central

    Patruno, Antonia; Pesce, Mirko; Grilli, Alfredo; Speranza, Lorenza; Franceschelli, Sara; De Lutiis, Maria Anna; Vianale, Giovina; Costantini, Erica; Amerio, Paolo; Muraro, Raffaella; Felaco, Mario; Reale, Marcella

    2015-01-01

    Several reports suggest that ELF-EMF exposures interact with biological processes including promotion of cell proliferation. However, the molecular mechanisms by which ELF-EMF controls cell growth are not completely understood. The present study aimed to investigate the effect of ELF-EMF on keratinocytes proliferation and molecular mechanisms involved. Effect of ELF-EMF (50 Hz, 1 mT) on HaCaT cell cycle and cells growth and viability was monitored by FACS analysis and BrdU assay. Gene expression profile by microarray and qRT-PCR validation was performed in HaCaT cells exposed or not to ELF-EMF. mTOR, Akt and MAPKs expressions were evaluated by Western blot analysis. In HaCaT cells, short ELF-EMF exposure modulates distinct patterns of gene expression involved in cell proliferation and in the cell cycle. mTOR activation resulted the main molecular target of ELF-EMF on HaCaT cells. Our data showed the increase of the canonical pathway of mTOR regulation (PI3K/Akt) and activation of ERK signaling pathways. Our results indicate that ELF-EMF selectively modulated the expression of multiple genes related to pivotal biological processes and functions that play a key role in physio-pathological mechanisms such as wound healing. PMID:26431550

  7. Effects of SIRT1 gene knock-out via activation of SREBP2 protein-mediated PI3K/AKT signaling on osteoarthritis in mice.

    PubMed

    Yu, Fei; Zeng, Hui; Lei, Ming; Xiao, De-Ming; Li, Wei; Yuan, Hao; Lin, Jian-Jing

    2016-10-01

    This study investigated the effects of SIRT1 gene knock-out on osteoarthritis in mice, and the possible roles of SREBP2 protein and the PI3K/AKT signaling pathway in the effects. Mice were randomly divided into a normal group and a SIRT1 gene knock-out group (6 mice in each group). In these groups, one side of the knee anterior cruciate ligament was traversed, and the ipsilateral medial meniscus was cut to establish an osteoarthritis model of knee joint. The countralateral synovial bursa was cut out, serving as controls. The knee joint specimens were then divided into four groups: SIRT1(+/+) control group (group A, n=6); SIRT1(+/+) osteoarthritis group (group B, n=6); SIRT1(-/-) control group (group C, n=6); SIRT1(-/-) osteoarthritis group (group D, n=6). HE staining, Masson staining, Safranin O-Fast Green staining and Van Gieson staining were used to observe the morphological changes in the articular cartilage of the knee. Immunohistochemical staining was employed to detect the expression of SIRT1, SREBP2, VEGF, AKT, HMGCR and type II collagen proteins. SA-β-gal staining was utilized to evaluate chondrocyte aging. The results showed clear knee joint cartilage destruction and degeneration in the SIRT1(-/-) osteoarthritis group. The tidal line was twisted and displaced anteriorly. Type II collagen was destroyed and distributed unevenly. Compared with the SIRT1(+/+) osteoarthritis group and SIRT1(-/-) control group, SIRT1 protein expression was not obviously changed in the SIRT1(-/-) osteoarthritis group (P>0.05), while the expression levels of the SREBP2, VEGF and HMGCR proteins were significantly increased (P<0.05) and the levels of AKT and type II collagen proteins were significantly decreased (P<0.05). SIRT1 gene knock-out may aggravate cartilage degeneration in osteoarthritis by activating the SREBP2 protein-mediated PI3K/AKT signalling pathway, suggesting that SIRT1 gene may play a protective role against osteoarthritis.

  8. Immunohistochemical Analysis of the Activation Status of the Akt/mTOR/pS6 Signaling Pathway in Oral Lichen Planus

    PubMed Central

    Prodromidis, Georgios; Nikitakis, Nikolaos G.; Sklavounou, Alexandra

    2013-01-01

    Introduction. Aberrations of the Akt/mTOR/pS6 pathway have been linked to various types of human cancer, including oral squamous cell carcinoma (OSCC). The purpose of this study was to evaluate the activation status of Akt, mTOR, and pS6 in oral lichen planus (OLP) in comparison with oral premalignant and malignant lesions and normal oral mucosa (NM). Materials and Methods. Immunohistochemistry for p-Akt, p-mTOR, and phospho-pS6 was performed in 40 OLP, 20 oral leukoplakias (OL), 10 OSCC, and 10 control samples of NM. Results. Nuclear p-Akt expression was detected in the vast majority of cases in all categories, being significantly higher in OL. Cytoplasmic p-Akt and p-mTOR staining was present only in a minority of OLP cases, being significantly lower compared to OL and OSCC. Phospho-pS6 showed cytoplasmic positivity in most OLP cases, which however was significantly lower compared to OL and OSCC. Conclusions. Overall, cytoplasmic p-Akt, p-mTOR, and phospho-pS6 levels appear to be significantly lower in OLP compared to OL and OSCC. However, the expression of these molecules in a subset of OLP cases suggests that activation of Akt/mTOR/pS6 may occur in the context of OLP, possibly contributing to the premalignant potential of individual cases. PMID:24228033

  9. HPV16 E6-E7 induces cancer stem-like cells phenotypes in esophageal squamous cell carcinoma through the activation of PI3K/Akt signaling pathway in vitro and in vivo

    PubMed Central

    Xi, Ruxing; Pan, Shupei; Chen, Xin; Hui, Beina; Zhang, Li; Fu, Shenbo; Li, Xiaolong; Zhang, Xuanwei; Gong, Tuotuo; Guo, Jia; Zhang, Xiaozhi; Che, Shaomin

    2016-01-01

    High-risk human papillomavirus (HPV), especially HPV16, correlates with cancerogenesis of human esophageal squamous cell carcinoma (ESCC) and we have reported that HPV16 related with a poor prognosis of ESCC patients in China. We aim to investigate the potential role and mechanism of HPV16 in ESCC development and progress. Our following researches demonstrated that ESCC cells which were stably transfected by HPV16 E6-E7 lentiviral vector showed a remarkable cancer stem-like cells (CSCs) phenotype, such as: migration, invasion, spherogenesis, high expression of CSCs marker in ESCC---p75NTR, chemoresistance, radioresistance, anti-apoptosis ability in vitro and cancerogenesis in vivo. HPV16 E6-E7 induced PI3K/Akt signaling pathway activation and this affect could be effectively inhibited by LY294002, a specific PI3K inhibitor. It was also indicated that the inhibition of PI3K/Akt signaling pathway by PI3K and Akt siRNA reverse the effect which induced by HPV16 E6-E7 in ESCC cells. Taken together, the present study demonstrates that HPV16 E6-E7 promotes CSCs phenotype in ESCC cells through the activation of PI3K/Akt signaling pathway. Targeting the PI3K/Akt signaling pathway in HPV16 positive tissues is an available therapeutic for ESCC patients. PMID:27489353

  10. HPV16 E6-E7 induces cancer stem-like cells phenotypes in esophageal squamous cell carcinoma through the activation of PI3K/Akt signaling pathway in vitro and in vivo.

    PubMed

    Xi, Ruxing; Pan, Shupei; Chen, Xin; Hui, Beina; Zhang, Li; Fu, Shenbo; Li, Xiaolong; Zhang, Xuanwei; Gong, Tuotuo; Guo, Jia; Zhang, Xiaozhi; Che, Shaomin

    2016-08-30

    High-risk human papillomavirus (HPV), especially HPV16, correlates with cancerogenesis of human esophageal squamous cell carcinoma (ESCC) and we have reported that HPV16 related with a poor prognosis of ESCC patients in China. We aim to investigate the potential role and mechanism of HPV16 in ESCC development and progress. Our following researches demonstrated that ESCC cells which were stably transfected by HPV16 E6-E7 lentiviral vector showed a remarkable cancer stem-like cells (CSCs) phenotype, such as: migration, invasion, spherogenesis, high expression of CSCs marker in ESCC---p75NTR, chemoresistance, radioresistance, anti-apoptosis ability in vitro and cancerogenesis in vivo. HPV16 E6-E7 induced PI3K/Akt signaling pathway activation and this affect could be effectively inhibited by LY294002, a specific PI3K inhibitor. It was also indicated that the inhibition of PI3K/Akt signaling pathway by PI3K and Akt siRNA reverse the effect which induced by HPV16 E6-E7 in ESCC cells. Taken together, the present study demonstrates that HPV16 E6-E7 promotes CSCs phenotype in ESCC cells through the activation of PI3K/Akt signaling pathway. Targeting the PI3K/Akt signaling pathway in HPV16 positive tissues is an available therapeutic for ESCC patients.

  11. Rutin attenuates H2O2-induced oxidation damage and apoptosis in Leydig cells by activating PI3K/Akt signal pathways.

    PubMed

    Sun, Jianhua; Wang, Heng; Liu, Bei; Shi, Wenhao; Shi, Juanzi; Zhang, Zhou; Xing, Junping

    2017-04-01

    Oxidative stress is a primary factor in the pathology of male infertility. The strong antioxidative capacity of rutin has been proven by numerous studies, but a protective role in the context of male reproduction remains to be elucidated. To explore the biological role of rutin in protecting male reproductive function and the potential underlying mechanism, H2O2-induced Leydig cells were used as a cell model of oxidation damage. Our findings showed that rutin at concentrations of 10, 20, and 40μmol/L remarkably increased cell survival rate of H2O2-induced Leydig cells to 70.1%, 86.8%, and 80.3% respectively. Next, rutin with concentrations of 10, 20, and 40μmol/L decreased reactive oxygen species (ROS) and malondialdehyde (MDA) levels but increased the levels of glutathione (GSH) and testosterone in H2O2-induced Leydig cells. The activities of superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) were remarkably increased by rutin treatment with concentrations of 20 and 40μmol/L, but glutathione peroxidase (GSH-Px) activity was notably decreased. Moreover, rutin with concentrations of 10, 20, and 40μmol/L increased Bcl-2 protein levels but decreased protein levels of Bax and caspase-3. Furthermore, 20μmol/L rutin significantly abrogated the decrease in levels of phosphoinositide 3-kinase (PI3K) and phosphorylated serine/threonine kinase (p-AKT) induced by H2O2. Pretreatment with LY294002, a PI3K inhibitor, antagonized protective action of 20μmol/L rutin against H2O2-induced cell activities, intracellular oxidant, testosterone, antioxidant enzyme activities, and the apoptosis related protein expression. Taken together, these results suggest that rutin attenuates H2O2-induced oxidation damage and apoptosis in Leydig cells by activating PI3K/Akt signal pathways, providing a promising strategy to decrease oxidative stress associated with male infertility.

  12. (+)-Catechin Attenuates NF-κB Activation Through Regulation of Akt, MAPK, and AMPK Signaling Pathways in LPS-Induced BV-2 Microglial Cells.

    PubMed

    Syed Hussein, Sharifah Salwa; Kamarudin, Muhamad Noor Alfarizal; Kadir, Habsah Abdul

    2015-01-01

    (+)-Catechin is a flavanol that possesses various health and medicinal values, which include neuroprotection, anti-oxidation, antitumor and antihepatitis activities. This study investigated the modulatory effects of (+)-catechin on the lipopolysaccharides (LPS)-stimulated BV-2 cells. (+)-catechin attenuated LPS-induced inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and inhibited microglial NO and ROS production. Additionally, (+)-catechin suppressed the production of tumor necrosis factor-α (TNF-α) and interleukin (IL)-6, while augmenting IL-4. (+)-catechin attenuated LPS-induced nuclear factor-κB (NF-κB) p65 nuclear translocation via the inhibition of IκB-α phosphorylation. Moreover, (+)-catechin blocked the activation of Akt and its inhibition was shown to play a crucial role in LPS-induced inflammation in BV-2 microglial cells. (+)-catechin also attenuated the LPS-induced phosphorylation of extracellular signal-regulated kinase (ERK1/2), and p-38 mitogen activated protein kinases (p38 MAPK) and specific inhibitors of ERK1/2 (UO126) and p38 MAPK (SB202190) subsequently down-regulated the expression of the proinflammatory mediators iNOS and COX-2. Further mechanistic study revealed that (+)-catechin acted through the amelioration of the LPS-induced suppression of adenosine monophosphate-activated protein kinase (AMPK) activity. Taken together, our data indicate that (+)-catechin exhibits anti-inflammatory effects in BV-2 cells by suppressing the production of proinflammatory mediators and mitigation of NF-κB through Akt, ERK, p38 MAPK, and AMPK pathways.

  13. Dissecting signalling by individual Akt/PKB isoforms, three steps at once.

    PubMed

    Osorio-Fuentealba, Cesar; Klip, Amira

    2015-09-01

    The serine/threonine kinase Akt/PKB (protein kinase B) is key for mammalian cell growth, survival, metabolism and oncogenic transformation. The diverse level and tissue expression of its three isoforms, Akt1/PKBα, Akt2/PKBβ and Akt3/PKBγ, make it daunting to identify isoform-specific actions in vivo and even in isolated tissues/cells. To date, isoform-specific knockout and knockdown have been the best strategies to dissect their individual overall functions. In a recent article in the Biochemical Journal, Kajno et al. reported a new strategy to study isoform selectivity in cell lines. Individual Akt/PKB isoforms in 3T3-L1 pre-adipocytes are first silenced via shRNA and stable cellular clones lacking one or the other isoform are selected. The stably silenced isoform is then replaced by a mutant engineered to be refractory to inhibition by MK-2206 (Akt1(W80A) or Akt2(W80A)). Akt1(W80A) or Akt2(W80A) are functional and effectively recruited to the plasma membrane in response to insulin. The system affords the opportunity to acutely control the activity of the endogenous non-silenced isoform through timely addition of MK-2206. Using this approach, it is confirmed that Akt1/PKBα is the preferred isoform sustaining adipocyte differentiation, but both Akt1/PKBα and Akt2/PKBβ can indistinctly support insulin-dependent FoxO1 (forkhead box O1) nuclear exclusion. Surprisingly, either isoform can also support insulin-dependent glucose transporter (GLUT) 4 translocation to the membrane, in contrast with the preferential role of Akt2/PKBβ assessed by knockdown studies. The new strategy should allow analysis of the plurality of Akt/PKB functions in other cells and in response to other stimuli. It should also be amenable to high-throughput studies to speed up advances in signal transmission by this pivotal kinase.

  14. Activation of PI3K/Akt/mTOR signaling in the tumor stroma drives endocrine therapy-dependent breast tumor regression

    PubMed Central

    Polo, María Laura; Riggio, Marina; May, María; Rodríguez, María Jimena; Perrone, María Cecilia; Stallings-Mann, Melody; Kaen, Diego; Frost, Marlene; Goetz, Matthew; Boughey, Judy; Lanari, Claudia; Radisky, Derek; Novaro, Virginia

    2015-01-01

    Improved efficacy of neoadjuvant endocrine-targeting therapies in luminal breast carcinomas could be achieved with optimal use of pathway targeting agents. In a mouse model of ductal breast carcinoma we identify a tumor regressive stromal reaction that is induced by neoadjuvant endocrine therapy. This reparative reaction is characterized by tumor neovascularization accompanied by infiltration of immune cells and carcinoma-associated fibroblasts that stain for phosphorylated ribosomal protein S6 (pS6), downstream the PI3K/Akt/mTOR pathway. While tumor variants with higher PI3K/Akt/mTOR activity respond well to a combination of endocrine and PI3K/Akt/mTOR inhibitors, tumor variants with lower PI3K/Akt/mTOR activity respond more poorly to the combination therapy than to the endocrine therapy alone, associated with inhibition of stromal pS6 and the reparative reaction. In human breast cancer xenografts we confirm that such differential sensitivity to therapy is primarily determined by the level of PI3K/Akt/mTOR in tumor cells. We further show that the clinical response of breast cancer patients undergoing neoadjuvant endocrine therapy is associated with the reparative stromal reaction. We conclude that tumor level and localization of pS6 are associated with therapeutic response in breast cancer and represent biomarkers to distinguish which tumors will benefit from the incorporation of PI3K/Akt/mTOR inhibitors with neoadjuvant endocrine therapy. PMID:26098779

  15. Activation of GRs-Akt-nNOs-NR2B signaling pathway by second dose GR agonist contributes to exacerbated hyperalgesia in a rat model of radicular pain.

    PubMed

    Zhang, Jing; Zhang, Wei; Sun, Yu'e; Liu, Yue; Song, Lihua; Ma, Zhengliang; Gu, Xiaoping

    2014-06-01

    Central Akt, neuronal nitric oxide synthase (nNOS) and N-methyl-D-aspartate receptor subunit 2B (NR2B) play key roles in the development of neuropathic pain. Here we investigate the effects of glucocorticoid receptors (GRs) on the expression and activation of spinal Akt, nNOS and NR2B after chronic compression of dorsal root ganglia (CCD). Thermal hyperalgesia test and mechanical allodynia test were used to measure rats after intrathecal injection of GR antagonist mifepristone or GR agonist dexamethasone for 21 days postoperatively. Expression of spinal Akt, nNOS, NR2B and their phosphorylation state after CCD was examined by western blot. The effects of intrathecal treatment with dexamethasone or mifepristone on nociceptive behaviors and the corresponding expression of Akt, nNOS and NR2B in spinal cord were also investigated. Intrathecal injection of mifepristone or dexamethasone inhibited PWMT and PWTL in CCD rats. However, hyperalgesia was induced by intrathecal injection of dexamethasone on days 12 to 14 after surgery. Treatment of dexamethasone increased the expression and phosphorylation levels of spinal Akt, nNOS, GR and NR2B time dependently, whereas administration of mifepristone downregulated the expression of these proteins significantly. GRs activated spinal Akt-nNOS/NR2B pathway play important roles in the development of neuropathic pain in a time-dependent manner.

  16. ON 01910.Na (rigosertib) inhibits PI3K/Akt pathway and activates oxidative stress signals in head and neck cancer cell lines

    PubMed Central

    Prasad, Anil; Khudaynazar, Nagina; Tantravahi, Ramana V.; Gillum, Amanda M.; Hoffman, Benjamin S.

    2016-01-01

    Squamous cell carcinoma of the head and neck (HNSCC) is characterized by high morbidity and mortality. Treatment failure, drug resistance and chemoradiation toxicity have necessitated the development of alternative treatment strategies. Styryl benzyl sulfones, a family of novel small molecule inhibitors, are being evaluated as anti-neoplastic agents in multiple clinical trials. The activity of these compounds has been well characterized in several preclinical tumor studies, but their activity has yet to be fully examined in HNSCC. We tested ON 01910.Na (rigosertib), a styryl benzyl sulfone in late-stage development, in HNSCC preclinical models. Rigosertib induced cytotoxicity in both HPV(+) and HPV(−) HNSCC cells in a dose-dependent manner. Characterization of the underlying molecular mechanism indicated that rigosertib induced inhibition of the PI3K/Akt/mTOR pathway, induced oxidative stress resulting in increased generation of reactive oxygen species (ROS), and activated extracellular signal-regulated kinases (ERK1/2) and c-Jun NH2-terminal kinase (JNK). Increased phosphorylation and cytoplasmic translocation of ATF-2 were also observed following rigosertib treatment. These changes in cell signaling led us to consider combining rigosertib with HNSCC standard-of-care therapies, such as cisplatin and radiation. Our study highlights the promising preclinical activity of rigosertib in HNSCC irrespective of HPV status and provides a molecular basis for rigosertib in combination with standard of care agents for HNSCC. PMID:27764820

  17. Phosphodiesterase 5a Inhibition with Adenoviral Short Hairpin RNA Benefits Infarcted Heart Partially through Activation of Akt Signaling Pathway and Reduction of Inflammatory Cytokines

    PubMed Central

    Jin, Zhe; Zhang, Jian; Paul, Christian; Wang, Yigang

    2015-01-01

    Introduction Treatment with short hairpin RNA (shRNA) interference therapy targeting phosphodiesterase 5a after myocardial infarction (MI) has been shown to mitigate post-MI heart failure. We investigated the mechanisms that underpin the beneficial effects of PDE5a inhibition through shRNA on post-MI heart failure. Methods An adenoviral vector with an shRNA sequence inserted was adopted for the inhibition of phosphodiesterase 5a (Ad-shPDE5a) in vivo and in vitro. Myocardial infarction (MI) was induced in male C57BL/6J mice by left coronary artery ligation, and immediately after that, the Ad-shPDE5a was injected intramyocardially around the MI region and border areas. Results Four weeks post-MI, the Ad-shPDE5a-treated mice showed significant mitigation of the left ventricular (LV) dilatation and dysfunction compared to control mice. Infarction size and fibrosis were also significantly reduced in Ad-shPDE5a-treated mice. Additionally, Ad-shPDE5a treatment decreased the MI-induced inflammatory cytokines interleukin (IL)-1β, IL-6, tumor necrosis factor-α, and transforming growth factor-β1, which was confirmed in vitro in Ad-shPDE5a transfected myofibroblasts cultured under oxygen glucose deprivation. Finally, Ad-shPDE5a treatment was found to activate the myocardial Akt signaling pathway in both in vivo and in vitro experiments. Conclusion These findings indicate that PDE5a inhibition by Ad-shPDE5a via the Akt signal pathway could be of significant value in the design of future therapeutics for post-MI heart failure. PMID:26709517

  18. FTY720 prevents ischemia/reperfusion injury-associated arrhythmias in an ex vivo rat heart model via activation of Pak1/Akt signaling.

    PubMed

    Egom, E Eroume A; Ke, Yunbo; Musa, Hanny; Mohamed, Tamer M A; Wang, Tao; Cartwright, Elizabeth; Solaro, R John; Lei, Ming

    2010-02-01

    Recent studies demonstrated a role of sphingosine-1-phosphate (S1P) in the protection against the stress of ischemia/reperfusion (I/R) injury. In experiments reported here, we have investigated the signaling through the S1P cascade by FTY720, a sphingolipid drug candidate displaying structural similarity to S1P, underlying the S1P cardioprotective effect. In ex vivo rat heart and isolated sinoatrial node models, FTY720 significantly prevented arrhythmic events associated with I/R injury including premature ventricular beats, VT, and sinus bradycardia as well as A-V conduction block. Real-time PCR and Western blot analysis demonstrated the expression of the S1P receptor transcript pools and corresponding proteins including S1P1, S1P2, and S1P3 in tissues dissected from sinoatrial node, atrium and ventricle. FTY720 (25 nM) significantly blunted the depression of the levels of phospho-Pak1 and phospho-Akt with ischemia and with reperfusion. There was a significant increase in phospho-Pak1 levels by 35%, 199%, and 205% after 5, 10, and 15 min of treatment with 25 nM FTY720 compared with control nontreated myocytes. However, there was no significant difference in the levels of total Pak1 expression between nontreated and FTY720 treated. Phospho-Akt levels were increased by 44%, 63%, and 61% after 5, 10, and 15 min of treatment with 25 nM FTY720, respectively. Our data provide the first evidence that FTY720 prevents I/R injury-associated arrhythmias and indicate its potential significance as an important and new agent protecting against I/R injury. Our data also indicate, for the first time, that the cardioprotective effect of FTY720 is likely to involve activation of signaling through the Pak1.

  19. Activation of phosphatidylinositol 3-kinase/Akt-mammalian target of Rapamycin signaling pathway in the hippocampus is essential for the acquisition of morphine-induced place preference in rats.

    PubMed

    Cui, Yue; Zhang, X Q; Cui, Y; Xin, W J; Jing, J; Liu, X G

    2010-11-24

    Hippocampus is a critical structure for the acquisition of morphine-induced conditioned place preference (CPP), which is a usual learning paradigm for assessing drug reward. However, the precise mechanisms remain largely unknown. Phosphatidylinositol 3-kinase (PI3K) and its downstream targets, including Akt, mammalian target of Rapamycin (mTOR) and 70-kDa ribosomal S6 kinase (p70S6K), are critical molecules implicated in learning and memory. Here, we tested the role of PI3K/Akt-mTOR-p70S6K signaling pathway in morphine-induced CPP in the hippocampus. Our results showed that the acquisition of morphine CPP increased phosphorylation of Akt in the hippocampal CA3, but not in the nucleus accumbens (NAc), the ventral tegmental area (VTA) or the CA1. Moreover, the phosphorylated Akt exclusively expressed in the CA3 neurons. Likewise, levels of phosphorylated mTOR and p70S6K were significantly enhanced in the CA3 following morphine CPP. The alterations of these phosphorylated proteins are positively correlated with the acquisition of morphine CPP. More importantly, microinjection of PI3K inhibitor (LY294002) or mTOR inhibitor (Rapamycin) into the CA3 prevented the acquisition of CPP and inhibited the activation of PI3K-Akt signaling pathway. In addition, pre-infusion of β-FNA (β-funaltrexamine hydrochloride), a selective irreversible μ opioid receptor antagonist, into CA3 significantly prevented the acquisition of CPP and impaired Akt phosphorylation. All these results strongly implied that the PI3K-Akt signaling pathway activated by μ opioid receptor in hippocampal CA3 plays an important role in acquisition of morphine-induced CPP.

  20. Crocin Inhibits Oxidative Stress and Pro-inflammatory Response of Microglial Cells Associated with Diabetic Retinopathy Through the Activation of PI3K/Akt Signaling Pathway.

    PubMed

    Yang, Xinguang; Huo, Fuquan; Liu, Bei; Liu, Jing; Chen, Tao; Li, Junping; Zhu, Zhongqiao; Lv, Bochang

    2017-02-25

    Diabetic retinopathy (DR) is a serious microvascular complication of diabetes mellitus that is closely associated with the degeneration and loss of retinal ganglion cells (RGCs) caused by diabetic microangiopathy and subsequent oxidative stress and an inflammatory response. Microglial cells are classed as neurogliocytes and play a significant role in neurodegenerative diseases. Over-activated microglial cells may cause neurotoxicity and induce the death and apoptosis of RGCs. Crocin is one of the two most pharmacologically bioactive constituents in saffron. In the present study, we focused on the role of microglial cells in DR, suggesting that DR may cause the over-activation of microglial cells and induce oxidative stress and the release of pro-inflammatory factors. Microglial cells BV-2 and N9 were cultured, and high-glucose (HG) and free fatty acid (FFA) were used to simulate diabetes. The results showed that HG-FFA co-treatment caused the up-regulated expression of CD11b and Iba-1, indicating that BV-2 and N9 cells were over-activated. Moreover, oxidative stress markers and pro-inflammatory factors were significantly enhanced by HG-FFA treatment. We found that crocin prevented the oxidative stress and pro-inflammatory response induced by HG-FFA co-treatment. Moreover, using the PI3K/Akt inhibitor LY294002, we revealed that PI3K/Akt signaling plays a significant role in blocking oxidative stress, suppressing the pro-inflammatory response, and maintaining the neuroprotective effects of crocin. In total, these results provide a new insight into DR and DR-induced oxidative stress and the inflammatory response, which provide a potential therapeutic target for neuronal damage, vision loss, and other DR-induced complications.

  1. Multiple roles and therapeutic implications of Akt signaling in cancer

    PubMed Central

    Calvo, Emiliano; Bolós, Victoria; Grande, Enrique

    2009-01-01

    The prominence of the PI3K-Akt signaling pathway in several tumors indicates a relationship with tumor grade and proliferation. Critical cellular processes are driven through this pathway. More detailed knowledge of the pathogenesis of tumors would enable us to design targeted drugs to block both membrane tyrosine kinase receptors and the intracellular kinases involved in the transmission of the signal. The newly approved molecular inhibitors sunitinib (an inhibitor of vascular endothelial growth factor receptor, platelet-derived growth factor receptor, and other tyrosine kinase receptors), sorafenib (a serine–threonine kinase inhibitor that acts against B-Raf) and temsirolimus (an mTOR inhibitor) shown clinical activity in advanced kidney cancer. Chronic myeloid leukemia has changed its natural history thanks to imatinib and dasatinib, both of which inhibit the intracellular bcr/abl protein derived from the alteration in the Philadelphia chromosome. Intracellular pathways are still important in cancer development and their blockade directly affects outcome. Cross-talk has been observed but is not well understood. Vertical and horizontal pathway blockade are promising anticancer strategies. Indeed, preclinical and early clinical data suggest that combining superficial and intracellular blocking agents can synergize and leverage single-agent activity. The implication of the Akt signaling pathway in cancer is well established and has led to the development of new anticancer agents that block its activation. PMID:20616901

  2. Integrin β1, myosin light chain kinase and myosin IIA are required for activation of PI3K-AKT signaling following MEK inhibition in metastatic triple negative breast cancer

    PubMed Central

    Choi, Cheolwon; Kwon, Junyeob; Lim, Sunyoung; Helfman, David M.

    2016-01-01

    The effectiveness of targeted therapies against the Ras-ERK signaling pathway are limited due to adaptive resistance of tumor cells. Inhibition of the Ras-ERK pathway can result in activation of the PI3K-AKT pathway, thereby diminishing the therapeutic effects of targeting ERK signaling. Here we investigated the crosstalk between the Ras-ERK and PI3K-AKT pathways in MDA-MB-231 breast cancer cell lines that have a preference to metastasize to lung (LM2), brain (BrM2) or bone (BoM2). Inhibition of the Ras-ERK pathway reduced motility in both parental and BoM2 cells. In contrast, inhibition of the Ras-ERK pathway in BrM2 and LM2 cells resulted in activation of PI3K-AKT signaling that was responsible for continued cell motility. Analysis of the cross talk between Ras-ERK and PI3K-AKT signaling pathways revealed integrin β1, myosin light chain kinase (MLCK) and myosin IIA are required for the activation of PI3K-AKT following inhibition of the Ras-ERK pathway. Furthermore, feedback activation of the PI3K-AKT pathway following MEK suppression was independent of the epidermal growth factor receptor. Thus, integrin β1, MLCK, and myosin IIA are factors in the development of resistance to MEK inhibitors. These proteins could provide an opportunity to develop markers and therapeutic targets in a subgroup of triple negative breast cancer (TNBC) that exhibit resistance against MEK inhibition. PMID:27563827

  3. AKT/GSK3β signaling pathway is critically involved in human pluripotent stem cell survival

    PubMed Central

    Romorini, Leonardo; Garate, Ximena; Neiman, Gabriel; Luzzani, Carlos; Furmento, Verónica Alejandra; Guberman, Alejandra Sonia; Sevlever, Gustavo Emilio; Scassa, María Elida; Miriuka, Santiago Gabriel

    2016-01-01

    Human embryonic and induced pluripotent stem cells are self-renewing pluripotent stem cells (PSC) that can differentiate into a wide range of specialized cells. Basic fibroblast growth factor is essential for PSC survival, stemness and self-renewal. PI3K/AKT pathway regulates cell viability and apoptosis in many cell types. Although it has been demonstrated that PI3K/AKT activation by bFGF is relevant for PSC stemness maintenance its role on PSC survival remains elusive. In this study we explored the molecular mechanisms involved in the regulation of PSC survival by AKT. We found that inhibition of AKT with three non-structurally related inhibitors (GSK690693, AKT inhibitor VIII and AKT inhibitor IV) decreased cell viability and induced apoptosis. We observed a rapid increase in phosphatidylserine translocation and in the extent of DNA fragmentation after inhibitors addition. Moreover, abrogation of AKT activity led to Caspase-9, Caspase-3, and PARP cleavage. Importantly, we demonstrated by pharmacological inhibition and siRNA knockdown that GSK3β signaling is responsible, at least in part, of the apoptosis triggered by AKT inhibition. Moreover, GSK3β inhibition decreases basal apoptosis rate and promotes PSC proliferation. In conclusion, we demonstrated that AKT activation prevents apoptosis, partly through inhibition of GSK3β, and thus results relevant for PSC survival. PMID:27762303

  4. Lentiviral shRNA against KCa3.1 inhibits allergic response in allergic rhinitis and suppresses mast cell activity via PI3K/AKT signaling pathway

    PubMed Central

    Lin, Hai; Zheng, Chunquan; Li, Jing; Yang, Chen; Hu, Li

    2015-01-01

    Calcium-activated potassium ion channel-3.1 (KCa3.1) plays a pivotal role in the potassium-calcium exchange involved in atopy. This study aimed to explore the impact of lentiviral-mediated shRNA silencing KCa3.1 on allergic response in a murine allergic rhinitis (AR) model. The BALB/c mice were divided into four groups: untreated AR group, negative control AR group, lentiviral KCa3.1-shRNA treated AR group and normal control group. Concentrations of ovalbumin (OVA)-specific IgE, histamine and leukotrienes C4 (LTC4) in serum, and IL-4, IL-9 and IL-17 in nasal lavage fluid (NLF) were analyzed. Goblet cells and mast cells were counted. KCa3.1 positive cells were counted after immunolabelling by immunofluorescence method. KCa3.1, Mucin 5AC (MUC5AC), and tryptase mRNA levels were determined using real-time polymerase chain reaction. Furthermore, P815 cell line was used to explore the role and mechanism of lentiviral KCa3.1-shRNA on mast cells. The results showed that LV-KCa3.1-shRNA intervention effectively attenuated allergic responses in LV-KCa3.1-shRNA treated mice. LV-KCa3.1-shRNA intervention effectively suppressed KCa3.1 levels and phosphorylation of AKT in P815 cells, leading to the downregulation of tryptase, IL-6 and IL-8 levels. LV-KCa3.1-shRNA intervention effectively attenuated the allergic responses in AR and suppressed mast cell activity by inhibiting PI3K/AKT signaling pathway. PMID:26272420

  5. Blockade of the phosphatidylinositol-3-kinase-Akt signaling pathway enhances the induction of apoptosis by microtubule-destabilizing agents in tumor cells in which the pathway is constitutively activated.

    PubMed

    Fujiwara, Yusuke; Hosokawa, Yoshihisa; Watanabe, Kazushi; Tanimura, Susumu; Ozaki, Kei-ichi; Kohno, Michiaki

    2007-03-01

    Constitutive activation of the phosphatidylinositol-3-kinase (PI3K)-Akt signaling pathway is associated with the neoplastic phenotype in many human tumor cell types. Given the antiapoptotic role of this pathway, we examined whether its specific blockade might sensitize human tumor cells to the induction of apoptosis by various anticancer drugs. Although specific blockade of the PI3K-Akt pathway alone with inhibitors such as LY294002 did not induce cell death, it resulted in marked and selective enhancement of the induction of apoptosis by microtubule-destabilizing agents such as vincristine. This effect was apparent only in tumor cells in which the PI3K-Akt pathway is constitutively activated. Blockade of the PI3K-Akt pathway induced the activation of glycogen synthase kinase-3beta, which phosphorylates microtubule-associated proteins such as tau and thereby reduces their ability to bind and stabilize microtubules. The consequent destabilization of microtubules induced by the inhibition of PI3K-Akt signaling appeared to increase their sensitivity to low concentrations of microtubule-destabilizing agents that alone do not lead to the disruption of cytoplasmic microtubules in tumor cells. Such a synergistic effect on microtubule integrity was not apparent for stable microtubules in the neurites of neuronal cells. These results suggest that the administration of a combination of a PI3K-Akt pathway inhibitor and a microtubule-destabilizing agent is a potential chemotherapeutic strategy for the treatment of tumor cells in which this signaling pathway is constitutively activated.

  6. 4-Ketopinoresinol, a novel naturally occurring ARE activator, induces the Nrf2/HO-1 axis and protects against oxidative stress-induced cell injury via activation of PI3K/AKT signaling.

    PubMed

    Chen, Huang-Hui; Chen, Yu-Tsen; Huang, Yen-Wen; Tsai, Hui-Ju; Kuo, Ching-Chuan

    2012-03-15

    The Nrf2/ARE pathway plays an important role in inducing phase II detoxifying enzymes and antioxidant proteins and has been considered a potential target for cancer chemoprevention because it eliminates harmful reactive oxygen species or reactive intermediates generated from carcinogens. The objectives of this study were to identify novel Nrf2/ARE activators and to investigate the mechanistic signaling pathway involved in the activation of Nrf2-mediated cytoprotective effects against oxidative-induced cell injury. A stable ARE-driven luciferase reporter cell line was established to screen a potentially cytoprotective compound. 4-Ketopinoresinol (4-KPR), the (α-γ) double-cyclized type of lignan obtained from adlay (Coix lachryma-jobi L. var. ma-yuen Stapf), activates ARE-driven luciferase activity more effectively than the classical ARE activator tert-butylhydroquinone. 4-KPR treatment resulted in a transient increase in AKT phosphorylation and subsequent phosphorylation and nuclear translocation of Nrf2, along with increased expression of ARE-dependent cytoprotective genes, such as heme oxygenase-1 (HO-1), aldo-keto reductases, and glutathione synthetic enzyme. 4-KPR suppresses oxidative stress-induced DNA damage and cell death via upregulation of HO-1. Inhibition of PI3K/AKT signaling by chemical inhibitors or RNA interference not only suppressed 4-KPR-induced Nrf2/HO-1 activation, but also eliminated the cytoprotective effect against oxidative damage. These observations in an ARE-regulated gene system suggest that 4-KPR is a novel Nrf2/ARE-mediated transcription activator, activates the Nrf2/HO-1 axis, and protects against oxidative stress-induced cell injury via activation of PI3K/AKT signaling.

  7. Coactivation of the PI3K/Akt and ERK signaling pathways in PCB153-induced NF-κB activation and caspase inhibition

    SciTech Connect

    Liu, Changjiang; Yang, Jixin; Fu, Wenjuan; Qi, Suqin; Wang, Chenmin; Quan, Chao; Yang, Kedi

    2014-06-15

    Polychlorinated biphenyls (PCBs) are a group of persistent and widely distributed environmental pollutants that have various deleterious effects, e.g., neurotoxicity, endocrine disruption and reproductive abnormalities. In order to verify the hypothesis that the PI3K/Akt and MAPK pathways play important roles in hepatotoxicity induced by PCBs, Sprague–Dawley (SD) rats were dosed with PCB153 intraperitoneally at 0, 4, 16 and 32 mg/kg for five consecutive days; BRL cells (rat liver cell line) were treated with PCB153 (0, 1, 5, and 10 μM) for 24 h. Results indicated that the PI3K/Akt and ERK pathways were activated in vivo and in vitro after exposure to PCB153, and protein levels of phospho-Akt and phospho-ERK were significantly increased. Nuclear factor-κB (NF-κB) activation and caspase-3, -8 and -9 inhibition caused by PCB153 were also observed. Inhibiting the ERK pathway significantly attenuated PCB153-induced NF-κB activation, whereas inhibiting the PI3K/Akt pathway hardly influenced phospho-NF-κB level. However, inhibiting the PI3K/Akt pathway significantly elevated caspase-3, -8 and -9 activities, while the ERK pathway only synergistically regulated caspase-9. Proliferating cell nuclear antigen (PCNA), a reliable indicator of cell proliferation, was also induced. Moreover, PCB153 led to hepatocellular hypertrophy and elevated liver weight. Taken together, PCB153 leads to aberrant proliferation and apoptosis of hepatocytes through NF-κB activation and caspase inhibition, and coactivated PI3K/Akt and ERK pathways play critical roles in PCB153-induced hepatotoxicity. - Highlights: • PCB153 led to hepatotoxicity through NF-κB activation and caspase inhibition. • The PI3K/Akt and ERK pathways were coactivated in vivo and in vitro by PCB153. • The ERK pathway regulated levels of phospho-NF-κB and caspase-9. • The PI3K/Akt pathway regulated levels of caspase-3, -8 and -9.

  8. Hesperidin attenuates learning and memory deficits in APP/PS1 mice through activation of Akt/Nrf2 signaling and inhibition of RAGE/NF-κB signaling.

    PubMed

    Hong, Yan; An, Zhongping

    2015-09-21

    Numerous studies have demonstrated that oxidative stress and inflammation play a pivotal role in the pathophysiology of Alzheimer disease (AD). Hesperidin (HP) has various pharmacological effects including anti-oxidative, anti-inflammatory and neuroprotective properties. In this study, APP/PS1 mice were used to evaluate the neuroprotective effects of HP. We reported that intragastric administration of HP (40 mg/kg) for 90 days significantly attenuated cognitive impairment in APP/PS1 mice. HP treatment suppressed oxidative stress by reducing the levels of ROS, LPO, protein carbonyl and 8-OHdG and increasing the activity of HO-1, SOD, catalase, and GSH-Px. HP treatment also inhibited inflammation by decreasing the levels of TNF-α, C-reactive protein and MCP-1 and reducing the activity of NF-κB. Moreover, HP could reverse the decreased phosphorylation of Akt, the decreased phosphorylation of GSK-3β, the lessened Nrf2 and the reduced expression of HO-1. HP could also inhibit the increased the RAGE expression, the enhanced phosphorylation of IκBα, and the augmented nuclear translocation of NF-κB/p65 in cortex of APP/PS1 mice. Taken together, HP suppresses oxidative stress and inflammation via activation of Akt/Nrf2 signaling and inhibition of RAGE/NF-κB signaling and further confers neuroprotection.

  9. Phospholipase D2 mediates survival signaling through direct regulation of Akt in glioblastoma cells.

    PubMed

    Bruntz, Ronald C; Taylor, Harry E; Lindsley, Craig W; Brown, H Alex

    2014-01-10

    The lack of innovative drug targets for glioblastoma multiforme (GBM) limits patient survival to approximately 1 year following diagnosis. The pro-survival kinase Akt provides an ideal target for the treatment of GBM as Akt signaling is frequently activated in this cancer type. However, the central role of Akt in physiological processes limits its potential as a therapeutic target. In this report, we show that the lipid-metabolizing enzyme phospholipaseD(PLD) is a novel regulator of Akt inGBM.Studies using a combination of small molecule PLD inhibitors and siRNA knockdowns establish phosphatidic acid, the product of the PLD reaction, as an essential component for the membrane recruitment and activation of Akt. Inhibition of PLD enzymatic activity and subsequent Akt activation decreases GBM cell viability by specifically inhibiting autophagic flux. We propose a mechanism whereby phosphorylation of beclin1 by Akt prevents binding of Rubicon (RUN domain cysteine-rich domain containing beclin1-interacting protein), an interaction known to inhibit autophagic flux. These findings provide a novel framework through which Akt inhibition can be achieved without directly targeting the kinase.

  10. Phospholipase D2 Mediates Survival Signaling through Direct Regulation of Akt in Glioblastoma Cells*♦

    PubMed Central

    Bruntz, Ronald C.; Taylor, Harry E.; Lindsley, Craig W.; Brown, H. Alex

    2014-01-01

    The lack of innovative drug targets for glioblastoma multiforme (GBM) limits patient survival to approximately 1 year following diagnosis. The pro-survival kinase Akt provides an ideal target for the treatment of GBM as Akt signaling is frequently activated in this cancer type. However, the central role of Akt in physiological processes limits its potential as a therapeutic target. In this report, we show that the lipid-metabolizing enzyme phospholipase D (PLD) is a novel regulator of Akt in GBM. Studies using a combination of small molecule PLD inhibitors and siRNA knockdowns establish phosphatidic acid, the product of the PLD reaction, as an essential component for the membrane recruitment and activation of Akt. Inhibition of PLD enzymatic activity and subsequent Akt activation decreases GBM cell viability by specifically inhibiting autophagic flux. We propose a mechanism whereby phosphorylation of beclin1 by Akt prevents binding of Rubicon (RUN domain cysteine-rich domain containing beclin1-interacting protein), an interaction known to inhibit autophagic flux. These findings provide a novel framework through which Akt inhibition can be achieved without directly targeting the kinase. PMID:24257753

  11. Angiotensin II Signaling in Human Preadipose Cells: Participation of ERK1,2-Dependent Modulation of Akt

    PubMed Central

    Dünner, Natalia; Quezada, Carolina; Berndt, F. Andrés; Cánovas, José; Rojas, Cecilia V.

    2013-01-01

    The renin-angiotensin system expressed in adipose tissue has been implicated in the modulation of adipocyte formation, glucose metabolism, triglyceride accumulation, lipolysis, and the onset of the adverse metabolic consequences of obesity. As we investigated angiotensin II signal transduction mechanisms in human preadipose cells, an interplay of extracellular-signal-regulated kinases 1 and 2 (ERK1,2) and Akt/PKB became evident. Angiotensin II caused attenuation of phosphorylated Akt (p-Akt), at serine 473; the p-Akt/Akt ratio decreased to 0.5±0.2-fold the control value without angiotensin II (p<0.001). Here we report that the reduction of phosphorylated Akt associates with ERK1,2 activities. In the absence of angiotensin II, inhibition of ERK1,2 activation with U0126 or PD98059 resulted in a 2.1±0.5 (p<0.001) and 1.4±0.2-fold (p<0.05) increase in the p-Akt/Akt ratio, respectively. In addition, partial knockdown of ERK1 protein expression by the short hairpin RNA technique also raised phosphorylated Akt in these cells (the p-Akt/Akt ratio was 1.5±0.1-fold the corresponding control; p<0.05). Furthermore, inhibition of ERK1,2 activation with U0126 prevented the reduction of p-Akt/Akt by angiotensin II. An analogous effect was found on the phosphorylation status of Akt downstream effectors, the forkhead box (Fox) proteins O1 and O4. Altogether, these results indicate that angiotensin II signaling in human preadipose cells involves an ERK1,2-dependent attenuation of Akt activity, whose impact on the biological functions under its regulation is not fully understood. PMID:24098385

  12. Infectious bursal disease virus activates the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway by interaction of VP5 protein with the p85{alpha} subunit of PI3K

    SciTech Connect

    Wei Li; Hou Lei; Zhu Shanshan; Wang Jing; Zhou Jiao; Liu Jue

    2011-08-15

    Phosphatidylinositol 3-kinase (PI3K)/Akt signaling is commonly activated upon virus infection and has been implicated in the regulation of diverse cellular functions such as proliferation and apoptosis. The present study demonstrated for the first time that infectious bursal disease virus (IBDV), the causative agent of a highly contagious disease in chickens, can induce Akt phosphorylation in cultured cells, by a mechanism that is dependent on PI3K. Inhibition of PI3K activation greatly enhanced virus-induced cytopathic effect and apoptotic cell death as evidenced by cleavage of poly-ADP ribose polymerase and activation of caspase-3. Investigations into the mechanism of PI3K/Akt activation revealed that IBDV activates PI3K/Akt signaling through binding of the non-structural protein VP5 to regulatory subunit p85{alpha} of PI3K resulting in the suppression of premature apoptosis and improved virus growth after infection. The results presented here provide a basis for understanding molecular mechanism of IBDV infection.

  13. Positive feedback loop of IL-1β/Akt/RARα/Akt signaling mediates oncogenic property of RARα in gastric carcinoma

    PubMed Central

    Huang, Gui-Li; Liu, Yu; Shen, Jin-Xing; Zhou, Pan; Liu, Wen-Ming; Shen, Dong-Yan

    2017-01-01

    Abnormal expression and function of retinoic acid receptor α (RARα) have been reported to be associated with various cancers including acute promyelocytic leukemia and hepatocellular carcinoma. However, the role and the mechanism of RARα in gastric carcinoma (GC) were unknown. Here, the expression of RARα was frequently elevated in human GC tissues and cell lines, and its overexpression was closely correlated with tumor size, lymph node metastasis and clinical stages in GC patients. Moreover, RARα overexpression was related with pathological differentiation. Functionally, RARα knockdown inhibited the proliferation and metastasis of GC cells, as well as enhanced drug susceptibility both in vitro and in vivo. Additionally, RARα knockdown suppressed GC progression through regulating the expression of cell proliferation, cell cycle, invasion and drug resistance associated proteins, such as PCNA, CyclinB1, CyclinD2, CyclinE, p21, MMP9 and MDR1. Mechanistically, the above oncogenic properties of RARα in GC were closely associated with Akt signaling activation. Moreover, overexpression of RARα was induced by IL-1β/Akt signaling activation, which suggested a positive feedback loop of IL-1β/Akt/RARα/Akt signaling in GC. Taken together, we demonstrated that RARα was frequently elevated in GC and exerted oncogenic properties. It might be a potential molecular target for GC treatment. PMID:28035062

  14. PTEN differentially regulates expressions of ICAM-1 and VCAM-1 through PI3K/Akt/GSK-3β/GATA-6 signaling pathways in TNF-α-activated human endothelial cells.

    PubMed

    Tsoyi, Konstantin; Jang, Hwa Jin; Nizamutdinova, Irina Tsoy; Park, Kyungok; Kim, Young Min; Kim, Hye Jung; Seo, Han Geuk; Lee, Jae Heun; Chang, Ki Churl

    2010-11-01

    Phosphotase and tensin homolog deleted on chromosome 10 (PTEN) is a potent negative regulator of PI3K/Akt pathway. Here, we tried to elucidate the role of PTEN in the regulation of endothelial adhesion molecules, vascular cell adhesion molecule (VCAM)-1 and intracellular adhesion molecule (ICAM)-1, induced by TNF-α in human endothelial cells (ECs). Transfection with PTEN overexpressing vector resulted in the significant decrease in phosphorylation of Akt in TNF-α-treated ECs. PTEN strongly inhibited VCAM-1 but not ICAM-1, however this inhibitory effect was reversed by co-transfection with constitutively active-Akt (CA-Akt-HA) in TNF-α-stimulated ECs. Additionally, silencing of PTEN with specific siRNA showed significant increase of phosphor-Akt compared with TNF-α alone treated ECs. siPTEN significantly upregulated VCAM-1 but was indifferent to ICAM-1 in TNF-α-treated cells. Further, chromatin immunoprecipitation (ChIP) assay showed that PTEN targets GATA-6 but not IRF-1 binding to VCAM-1 promoter. In addition, GATA-6 is associated with glycogen synthesis kinase-3beta (GSK-3β) which is in turn regulated by PTEN-dependent Akt activity. Finally, PTEN significantly prevented monocyte adhesion to TNF-α-induced ECs probably through VCAM-1 regulation. It is concluded that PTEN selectively inhibits expression of VCAM-1 but not ICAM-1 through modulation of PI3K/Akt/GSK-3β/GATA-6 signaling cascade in TNF-α-treated ECs.

  15. Cisplatin triggers atrophy of skeletal C2C12 myotubes via impairment of Akt signalling pathway and subsequent increment activity of proteasome and autophagy systems

    SciTech Connect

    Fanzani, Alessandro Zanola, Alessandra; Rovetta, Francesca; Rossi, Stefania; Aleo, Maria Francesca

    2011-02-01

    Cisplatin (cisPt) is an antineoplastic drug which causes an array of adverse effects on different organs and tissues, including skeletal muscle. In this work we show that cisPt behaves as a potent trigger to activate protein hypercatabolism in skeletal C2C12 myotubes. Within 24 h of 50 {mu}M cisPt administration, C2C12 myotubes displayed unchanged cell viability but showed a subset of hallmark signs typically recognized during atrophy, including severe reduction in body size, repression of Akt phosphorylation, transcriptional up-regulation of atrophy-related genes, such as atrogin-1, gabarap, beclin-1 and bnip-3, and loss of myogenic markers. As a consequence, proteasomal activity and formation of autophagosomes were remarkably increased in cisPt-treated myotubes, but forced stimulation of Akt pathway, as obtained through insulin administration or delivery of a constitutively activated Akt form, was sufficient to counter the cisPt-induced protein breakdown, leading to rescue of atrophic size. Overall, these results indicate that cisPt induces atrophy of C2C12 myotubes via activation of proteasome and autophagy systems, suggesting that the Akt pathway represents one sensitive target of cisPt molecular action in skeletal muscle.

  16. Activated AKT regulates NF-kappaB activation, p53 inhibition and cell survival in HTLV-1-transformed cells.

    PubMed

    Jeong, Soo-Jin; Pise-Masison, Cynthia A; Radonovich, Michael F; Park, Hyeon Ung; Brady, John N

    2005-10-06

    AKT activation enhances resistance to apoptosis and induces cell survival signaling through multiple downstream pathways. We now present evidence that AKT is activated in HTLV-1-transformed cells and that Tax activation of AKT is linked to NF-kappaB activation, p53 inhibition and cell survival. Overexpression of AKT wild type (WT), but not a kinase dead (KD) mutant, resulted in increased Tax-mediated NF-kappaB activation. Blocking AKT with the PI3K/AKT inhibitor LY294002 or AKT SiRNA prevented NF-kappaB activation and inhibition of p53. Treatment of C81 cells with LY294002 resulted in an increase in the p53-responsive gene MDM2, suggesting a role for AKT in the Tax-mediated regulation of p53 transcriptional activity. Further, we show that LY294002 treatment of C81 cells abrogates in vitro IKKbeta phosphorylation of p65 and causes a reduction of p65 Ser-536 phosphorylation in vivo, steps critical to p53 inhibition. Interestingly, blockage of AKT function did not affect IKKbeta phosphorylation of IkappaBalpha in vitro suggesting selective activity of AKT on the IKKbeta complex. Finally, AKT prosurvival function in HTLV-1-transformed cells is linked to expression of Bcl-xL. We suggest that AKT plays a role in the activation of prosurvival pathways in HTLV-1-transformed cells, possibly through NF-kappaB activation and inhibition of p53 transcription activity.

  17. HER2-induced metastasis is mediated by AKT/JNK/EMT signaling pathway in gastric cancer

    PubMed Central

    Choi, Yiseul; Ko, Young San; Park, Jinju; Choi, Youngsun; Kim, Younghoon; Pyo, Jung-Soo; Jang, Bo Gun; Hwang, Douk Ho; Kim, Woo Ho; Lee, Byung Lan

    2016-01-01

    AIM To investigated the relationships between HER2, c-Jun N-terminal kinase (JNK) and protein kinase B (AKT) with respect to metastatic potential of HER2-positive gastric cancer (GC) cells. METHODS Immunohistochemistry was performed on tissue array slides containing 423 human GC specimens. Using HER2-positve GC cell lines SNU-216 and NCI-N87, HER2 expression was silenced by RNA interference, and the activations of JNK and AKT were suppressed by SP600125 and LY294002, respectively. Transwell assay, Western blot, semi-quantitative reverse transcription-polymerase chain reaction and immunofluorescence staining were used in cell culture experiments. RESULTS In GC specimens, HER2, JNK, and AKT activations were positively correlated with each other. In vitro analysis revealed a positive regulatory feedback loop between HER2 and JNK in GC cell lines and the role of JNK as a downstream effector of AKT in the HER2/AKT signaling pathway. JNK inhibition suppressed migratory capacity through reversing EMT and dual inhibition of JNK and AKT induced a more profound effect on cancer cell motility. CONCLUSION HER2, JNK and AKT in human GC specimens are positively associated with each other. JNK and AKT, downstream effectors of HER2, co-operatively contribute to the metastatic potential of HER2-positive GC cells. Thus, targeting of these two molecules in combination with HER2 downregulation may be a good approach to combat HER2-positive GC. PMID:27895401

  18. Apamin inhibits PDGF-BB-induced vascular smooth muscle cell proliferation and migration through suppressions of activated Akt and Erk signaling pathway.

    PubMed

    Kim, Jung-Yeon; Kim, Kyung-Hyun; Lee, Woo-Ram; An, Hyun-Jin; Lee, Sun-Jae; Han, Sang-Mi; Lee, Kwang-Gill; Park, Yoon-Yub; Kim, Kee-Sik; Lee, Young-Soo; Park, Kwan-Kyu

    2015-07-01

    The increased proliferation and migration of vascular smooth muscle cells (VSMC) are key process in the development of atherosclerosis lesions. Platelet-derived growth factor (PDGF) initiates a multitude of biological effects that contribute to VSMC proliferation and migration. Apamin, a component of bee venom, has been known to block the Ca(2+)-activated K(+) channels. However, the effects of apamin in the regulation PDGF-BB-induced VSMC proliferation and migration has not been identified. In this study, we investigate the inhibitory effect of apamin on PDGF-BB-induced VSMC proliferation and migration. Apamin suppressed the PDGF-BB-induced VSMC proliferation and migration with no apparent cytotoxic effect. In accordance with these findings, apamin induced the arrest of cell cycle progression at G0/G1 phase. Apamin also decreased the expressions of G0/G1 specific regulatory proteins including proliferating cell nuclear antigen (PCNA), cyclin D1, cyclin-dependent kinases (CDK) 4, cyclin E and CDK2, as well as increased the expression of p21(Cip1) in PDGF-BB-induced VSMC. Moreover, apamin inhibited PDGF-BB-induced phosphorylation of Akt and Erk1/2. These results suggest that apamin plays an important role in prevention of vascular proliferation and migration through the G0/G1 cell cycle arrest by PDGF signaling pathway. Thus, apamin may be a promising candidate for the therapy of atherosclerosis.

  19. Gli1 promotes colorectal cancer metastasis in a Foxm1-dependent manner by activating EMT and PI3K-AKT signaling

    PubMed Central

    Zhang, Yue; Ji, Bing; Wang, Sen; Sun, Ye; Zhu, Chunyan; Zhang, Dongsheng; Sun, Yueming

    2016-01-01

    Colorectal cancer(CRC) is one of the most commonly diagnosed cancers in human beings and metastasis is the main death reason. Recently, Gli1 has been reported to be a key regulator of various cancer biologies and genes expressions. However, the detailed molecular mechanism of Gli1 in CRC metastasis remains largely unknown. In this study, we aimed to investigate the role of Gli1 in CRC metastasis. We used qRT-PCR, Immunohistochemistry and Western blot to test the expression levels of Gli1, Foxm1 and other target genes in the tissues and cells; Lentivirus stable transfection to change the expression levels of Gli1 and Foxm1; Wound-healing, cell invasion, migration assays and tail vein metastatic assay to test the role of Gli1 in CRC metastasis in vitro and vivo. We demonstrated that Gli1 was significantly overexpressed in colorectal cancer tissues and cells. Foxm1 level had a positive correlation with Gli1. Furthermore, we found that Gli1 promotes colorectal cancer cells metastasis in a Foxm1-dependent manner by activating EMT and PI3K-AKT signaling. Thus, we proved that Gli1 plays important role in CRC metastasis and provided a new visual field on the therapy of CRC metastasis. PMID:27863385

  20. TMEFF2 modulates the AKT and ERK signaling pathways

    PubMed Central

    Chen, Xiaofei; Ruiz-Echevarría, Maria J

    2013-01-01

    The transmembrane protein with epidermal growth factor (EGF) and two follistatin (FS) motifs 2 (TMEFF2) has a limited tissue distribution with strong expression only in brain and prostate. While TMEFF2 is overexpressed in prostate cancer indicating an oncogenic role, several studies indicate a tumor suppressor role for this protein. This dual mode of action is, at least in part, the result of metalloproteinase-dependent shedding that generates a soluble TMEFF2 ectodomain with a growth promoting function. While recent studies have shed some light on the biology of different forms of TMEFF2, little is known about the molecular mechanisms that influence its oncogenic/tumor suppressive function. In several non-prostate cell lines, it has been shown that a recombinant form of the TMEFF2 ectodomain can interact with platelet derived growth factor (PDGF)-AA to suppress PDGF receptor signaling and can promote ErbB4 and ERK1/2 phosphorylation. However, the role of the full length TMEFF2 in these pathways has not been examined. Using prostate cell lines, here we examine the role of TMEFF2 in ERK and Akt activation, two pathways implicated in prostate cancer progression and that have been shown to cross talk in several cancers. Our results show that different forms of TMEFF2 distinctly affect Akt and ERK activation and this may contribute to a different cellular response of either proliferation or tumor suppression. PMID:23936739

  1. Notoginsenoside R1 attenuates glucose-induced podocyte injury via the inhibition of apoptosis and the activation of autophagy through the PI3K/Akt/mTOR signaling pathway

    PubMed Central

    Huang, Guodong; Zou, Bingyu; Lv, Jianzhen; Li, Tongyu; Huai, Guoli; Xiang, Shaowei; Lu, Shilong; Luo, Huan; Zhang, Yaping; Jin, Yi; Wang, Yi

    2017-01-01

    Injury to terminally differentiated podocytes contributes ignificantly to proteinuria and glomerulosclerosis. The aim of this study was to examine the protective effects of notoginsenoside R1 (NR1) on the maintenance of podocyte number and foot process architecture via the inhibition of apoptosis, the induction of autophagy and the maintenance pf podocyte biology in target cells. The effects of NR1 on conditionally immortalized human podocytes under high glucose conditions were evaluated by determining the percentage apoptosis, the percentage autophagy and the expression levels of slit diaphragm proteins. Our results revealed that NR1 protected the podocytes against high glucose-induced injury by decreasing apoptosis, increasing autophagy and by promoting cytoskeletal recovery. The phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling pathway was further investigated in order to elucidate the mechanisms responsible for the protective effects of NR1 on podocytes. Our data indicated that treatment with NR increased the phosphorylation levels of PI3K, Akt and mTOR, leading to the activation of the PI3K/Akt/mTOR signaling pathway in podocytes. To the best of our knowledge, this is the first in vitro study to demonstrate that NR1 protects podocytes by activating the PI3K/Akt/mTOR pathway. PMID:28112381

  2. PI3K/Akt/mTOR signaling pathway in cancer stem cells: from basic research to clinical application

    PubMed Central

    Xia, Pu; Xu, Xiao-Yan

    2015-01-01

    Cancer stem cells (CSCs) are a subpopulation of tumor cells that possess unique self-renewal activity and mediate tumor initiation and propagation. The PI3K/Akt/mTOR signaling pathway can be considered as a master regulator for cancer. More and more recent studies have shown the links between PI3K/Akt/mTOR signaling pathway and CSC biology. Herein, we provide a comprehensive review on the role of signaling components upstream and downstream of PI3K/Akt/mTOR signaling in CSC. In addition, we also summarize various classes of small molecule inhibitors of PI3K/Akt/mTOR signaling pathway and their clinical potential in CSC. Overall, the current available data suggest that the PI3K/Akt/mTOR signaling pathway could be a promising target for development of CSC-target drugs. PMID:26175931

  3. Canstatin inhibits hypoxia-induced apoptosis through activation of integrin/focal adhesion kinase/Akt signaling pathway in H9c2 cardiomyoblasts

    PubMed Central

    Yamawaki, Hideyuki

    2017-01-01

    A hypoxic stress which causes apoptosis of cardiomyocytes is the main problem in the ischemic heart disease. Canstatin, a non-collagenous fragment of type IV collagen α2 chain, is an endogenous anti-angiogenic factor. We have previously reported that canstatin has a cytoprotective effect on cardiomyoblasts. In the present study, we examined the effects of canstatin on hypoxia-induced apoptosis in H9c2 cardiomyoblasts. Cell counting assay was performed to determine a cell viability. Western blotting was performed to detect expression of cleaved casepase-3 and phosphorylation of focal adhesion kinase (FAK) and Akt. Immunocytochemical staining was performed to observe a distribution of αv integrin. Hypoxia (1% O2, 48 h) significantly decreased cell viability and increased cleaved caspase-3 expression. Canstatin (10–250 ng/ml) significantly inhibited these changes in a concentration-dependent manner. Cilengitide (1 μM), an αvβ3 and αvβ5 integrin inhibitor, significantly prevented the protective effects of canstatin on cell viability. Canstatin significantly increased phosphorylation of FAK and Akt under hypoxic condition, which were inhibited by cilengitide. LY294002, an inhibitor of phosphatidylinositol-3 kinase/Akt pathway, suppressed the canstatin-induced Akt phosphorylation and reversed the protective effects of canstatin. It was observed that hypoxia caused a localization of αv integrin to focal adhesion. In summary, we for the first time clarified that canstatin inhibits hypoxia-induced apoptosis via FAK and Akt pathways through activating integrins in H9c2 cardiomyoblasts. PMID:28235037

  4. PDGF inactivates forkhead family transcription factor by activation of Akt in glomerular mesangial cells.

    PubMed

    Ghosh Choudhury, Goutam; Lenin, Mahimainathan; Calhaun, Cheresa; Zhang, Jian-Hua; Abboud, Hanna E

    2003-02-01

    Regulation of the forkhead domain transcription factors by PDGF has not been studied. In this report, we investigated the role of PDGF-induced Akt in regulating forkhead domain protein FKHRL1 in glomerular mesangial cells. PDGF increased phosphorylation of FKHRL1 in a time- and PI 3 kinase-dependent manner. Expression of dominant negative Akt by adenovirus-mediated gene transfer blocked PDGF-induced FKHRL1 phosphorylation. PDGF inhibited transcription of a forkhead DNA binding element-driven reporter gene. This inhibition was mimicked by constitutively active myristoylated Akt. Moreover, FKHR1-mediated transcription of the reporter gene was completely attenuated by both PDGF and Myr-Akt. One of the targets of forkhead transcription factors is the proapoptotic Fas ligand (FasL) gene. PDGF, as well as Myr-Akt, inhibited transcription of FasL. In contrast, inhibition of PI 3 kinase and dominant negative Akt increased FasL gene transcription, suggesting that suppression of PI 3 kinase/Akt signalling may induce apoptosis in mesangial cells via upregulation of FasL expression. However, expression of dominant negative Akt by adenovirus did not induce apoptosis in mesangial cells, suggesting that Akt-independent antiapoptotic mechanisms also exist. Together, our data demonstrate for the first time that PDGF inactivates forkhead domain transcription factor by Akt-dependent phosphorylation and that suppression of Akt signalling is not sufficient to induce apoptosis in mesangial cells.

  5. Restoration of SHIP activity in a human leukemia cell line downregulates constitutively activated phosphatidylinositol 3-kinase/Akt/GSK-3beta signaling and leads to an increased transit time through the G1 phase of the cell cycle.

    PubMed

    Horn, S; Endl, E; Fehse, B; Weck, M M; Mayr, G W; Jücker, M

    2004-11-01

    The inositol 5-phosphatase SHIP (SHIP-1) is a negative regulator of signal transduction in hematopoietic cells and targeted disruption of SHIP in mice leads to a myeloproliferative disorder. We analyzed the effects of SHIP on the human leukemia cell line Jurkat in which expression of endogenous SHIP protein is not detectable. Restoration of SHIP expression in Jurkat cells with an inducible expression system caused a 69% reduction of phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) and a 65% reduction of Akt kinase activity, which was associated with reduced phosphorylation of glycogen synthase kinase 3beta (GSK-3beta) (Ser-9) without changing the phosphorylation of Bad (Ser-136), FKHR (Ser-256) or MAPK (Thr-202/Tyr-204). SHIP-expressing Jurkat cells showed an increased transit time through the G1 phase of the cell cycle, but SHIP did not cause a complete cell cycle arrest or apoptosis. Extension of the G1 phase was associated with an increased stability of the cell cycle inhibitor p27(Kip1) and reduced phosphorylation of the retinoblastoma protein Rb at serine residue 780. Our data indicate that restoration of SHIP activity in a human leukemia cell line, which has lost expression of endogenous SHIP, downregulates constitutively activated phosphatidylinositol 3-kinase/Akt/GSK-3beta signaling and leads to an increased transit time through the G1 phase of the cell cycle.

  6. The protective effects of Chinese herb-Taikong Yangxin Prescription on the atrophic remodeling of cardiac muscle in rats induced by hindlimb unloading through activating Akt/GSK-3beta signaling pathway

    NASA Astrophysics Data System (ADS)

    Ming, Yuan; Min, Yuan; Jianfeng, Zhang; Zhili, Li; Huijuan, Wang; Desheng, Wang; Yinghui, Li; Yongzhi, Li; Shizhong, Jiang

    Objective To test the hypothesis that traditional Chinese herb-TaiKong Yangxin Prescrip-tion can activate the Akt/GSK-3β signaling pathway and alleviate the atrophic remodeling of cardiac muscle in rats induced by hindlimb unloading. Methods The physiological effects of simulated microgravity was induced by 7d hindlimb unloading in rats. TaiKong Yangxin Pre-scription was given daily by gastric irrigation as countermeasure against effects of simulated microgravity. The frozen sections of left ventricular cardiac muscles were stained by FITC la-beled lectin and visualized by laser scanning confocal microscopy, the cross section areas(CSA) of cardiomyocytes were calculated by IPP6.0 Image software. The protein expression of TnI, phosphorylation level of Akt and GSK-3β were measured by Western blot. Results Simulated microgravity decreased the CSA of cardiomyocytes and protein expression of TnI in left ven-tricular cardiac muscles, inhibited the phosphorylation level of Akt at serine 473 and GSK-3β at serine 9. The traditional Chinese herb-TaiKong Yangxin Prescription alleviated the atrophic remodeling of cardiac muscles, reversed the declined protein expression of TnI and phosphoryla-tion levels of Akt at serine 473 and GSK-3β at serine 9 in hindlimb-unloading rats. Conclusion The traditional Chinese herb-TaiKong Yangxin Prescription has significant countermeasure effects on the atrophic remodeling of cardiac muscle induced by hindlimb unloading in rats, in which activating Akt/GSK-3β signaling pathway plays an important role.(Funded by Advanced space medico-engineering research project of China, grant NO. 2005SY5206005 and SJ200801)

  7. Mutual inhibition of insulin signaling and PHLPP-1 determines cardioprotective efficiency of Akt in aged heart

    PubMed Central

    Xing, Yuan; Sun, Wanqing; Wang, Yishi; Gao, Feng; Ma, Heng

    2016-01-01

    Insulin protects cardiomyocytes from myocardial ischemia/reperfusion (MI/R) injury through activating Akt. However, phosphatase PHLPP-1 (PH domain leucine-rich repeat protein phosphatase-1) dephosphorylates and inactivates Akt. The balanced competitive interaction of insulin and PHLPP-1 has not been directly examined. In this study, we have identified the effect of mutual inhibition of insulin signaling and PHLPP-1 on the cardioprotective efficiency of Akt in aged heart. Young (3 mon) and aged (20 mon) Sprague Dawley (SD) rats were subjected to MI/R in vivo. The PHLPP-1 level was higher in aged vs. young hearts at base. But, insulin treatment failed to decrease PHLPP-1 level during reperfusion in the aged hearts. Consequently, the cardioprotection of insulin-induced Akt activation was impaired in aged hearts, resulting in more susceptible to MI/R injury. In cultured rat ventricular myocytes, PHLPP-1 knockdown significantly enhanced insulin-induced Akt phosphorylation and reduced simulated hypoxia/reoxygenation-induced apoptosis. Contrary, PHLPP-1 overexpression terminated Akt phosphorylation and deteriorated myocytes apoptosis. Using in vivo aged animal models, we confirmed that cardiac PHLPP-1 knockdown or enhanced insulin sensitivity by exercise training dramatically increased insulin-induced Akt phosphorylation. Specifically, MI/R-induced cardiomyocyte apoptosis and infarct size were decreased and cardiac function was increased. More importantly, we found that insulin regulated the degradation of PHLPP-1 and insulin treatment could enhance the binding between PHLPP-1 and β-transducin repeat-containing protein (β-TrCP) to target for ubiquitin-dependent degradation. Altogether, we have identified a new mechanism by which insulin suppresses PHLPP-1 to enhance Akt activation. But, aged heart possesses lower insulin effectiveness and fails to decrease PHLPP-1 during MI/R, which subsequently limited Akt activity and cardioprotection. PHLPP-1 could be a promising

  8. Vascular neuroprotection via TrkB- and Akt-dependent cell survival signaling.

    PubMed

    Guo, Shuzhen; Som, Angel T; Waeber, Christian; Lo, Eng H

    2012-11-01

    The cerebral endothelium can be a vital source of signaling factors such as brain-derived neurotrophic factor that defends the neuronal parenchyma against stress and injury. But the underlying mechanisms remain to be fully defined. Here, we use cell models to ask how vascular neuroprotection is sustained. Human brain endothelial cells were grown in culture, and conditioned media were transferred to primary rat cortical neurons. Brain endothelial cell-conditioned media activated neuronal Akt signaling and protected neurons against hypoxia and oxygen-glucose deprivation. Blockade of Akt phosphorylation with the PI3-kinase inhibitor LY294002 negated this vascular neuroprotective effect. Upstream of Akt signaling, the brain-derived neurotrophic factor receptor TrkB (neurotrophic tyrosine kinase receptor, type 2) was involved because depletion with TrkB/Fc eliminated the ability of endothelial-conditioned media to protect neurons against hypoxia. Downstream of Akt signaling, activation of GSK-3β (glycogen synthase kinase 3 beta), caspase 9, caspase 3 and Bad pathways were detected. Taken together, these findings suggest that the molecular basis for vascular neuroprotection involves TrkB-Akt signaling that ameliorates neuronal apoptosis. Further investigation of these mechanisms may reveal new approaches for augmenting endogenous vascular neuroprotection in stroke, brain injury, and neurodegeneration.

  9. Activation of sonic hedgehog signaling enhances cell migration and invasion by induction of matrix metalloproteinase-2 and -9 via the phosphoinositide-3 kinase/AKT signaling pathway in glioblastoma.

    PubMed

    Chang, Liang; Zhao, Dan; Liu, Hui-Bin; Wang, Qiu-Shi; Zhang, Ping; Li, Chen-Long; Du, Wen-Zhong; Wang, Hong-Jun; Liu, Xing; Zhang, Zhi-Ren; Jiang, Chuan-Lu

    2015-11-01

    Aberrant hedgehog signaling contributes to the development of various malignancies, including glioblastoma (GBM). However, the potential mechanism of hedgehog signaling in GBM migration and invasion has remained to be elucidated. The present study showed that enhanced hedgehog signaling by recombinant human sonic hedgehog N‑terminal peptide (rhSHH) promoted the adhesion, invasion and migration of GBM cells, accompanied by increases in mRNA and protein levels of matrix metalloproteinase‑2 (MMP‑2) and MMP‑9. However, inhibition of hedgehog signaling with cyclopamine suppressed the adhesion, invasion and migration of GBM cells, accompanied by decreases in mRNA and protein levels of MMP‑2 and ‑9. Furthermore, it was found that MMP‑2- and MMP‑9-neutralizing antibodies or GAM6001 reversed the inductive effects of rhSHH on cell migration and invasion. In addition, enhanced hedgehog signaling by rhSHH increased AKT phosphorylation, whereas blockade of hedgehog signaling decreased AKT phosphorylations. Further experiments showed that LY294002, an inhibitor of phosphoinositide-3 kinase (PI3K), decreased rhSHH‑induced upregulation of MMP‑2 and ‑9. Finally, the protein expression of glioblastoma-associated oncogene 1 was positively correlated with levels of phosphorylated AKT as well as protein expressions of MMP‑2 and ‑9 in GBM tissue samples. In conclusion, the present study indicated that the hedgehog pathway regulates GBM-cell migration and invasion by increasing MMP-2 and MMP-9 production via the PI3K/AKT pathway.

  10. Interleukin-11 promotes epithelial-mesenchymal transition in anaplastic thyroid carcinoma cells through PI3K/Akt/GSK3β signaling pathway activation

    PubMed Central

    Jiang, Yue; Sun, Ruimei; Chen, Xue; Chu, Hongying; Zeng, Musheng; Sun, Chuanzheng

    2016-01-01

    Metastasis is the major cause of treatment failure in anaplastic thyroid carcinoma (ATC) patients. In the preliminary study, we demonstrated that interleukin (IL)-11 expression is positively correlated with distant metastasis in ATC. However, the mechanisms underlying remain largely unknown. Here, we found that cobalt chloride (a hypoxia mimetic) promoted IL-11 expression via HIF-1α activation. Furthermore, the resultant increase in IL-11 expression significantly induced epithelial-mesenchymal transition (EMT) in ATC cells, accompanied by Akt/GSK3β pathway activation and increased invasive and migratory abilities. Conversely, HIF-1α or IL-11 knockdown, or treating cells with a neutralizing antibody against IL-11, a PI3K inhibitor, or Akt inhibitor V, significantly suppressed the induction of EMT and counteracted the enhancements in invasive and migratory abilities. These results indicate that hypoxia increases IL-11 secretion in ATC cells via HIF-1α induction and that IL-11 then induces EMT in these cells via the PI3K/Akt/GSK3β pathway, ultimately improving their invasive and migratory potential. This study elucidates the prometastatic role played by IL-11 in ATC metastasis and indicates it as a potential target for the treatment of cancer metastasis. However, many questions remain to be explored. PMID:27487122

  11. The neuroprotective effect of a novel agent N2 on rat cerebral ischemia associated with the activation of PI3K/Akt signaling pathway.

    PubMed

    Huang, Jinru; Kodithuwakku, Nandani Darshika; He, Wei; Zhou, Yi; Fan, Wenxiang; Fang, Weirong; He, Guangwei; Wu, Qiang; Chu, Shaoxing; Li, Yunman

    2015-08-01

    Ischemic stroke is the third leading cause of death and the main reason for severe disabilities in the world today. N2, 4 - (2 - (1H - imidazol - 1 - yl) ethoxy) - 3 - methoxybenzoic acid is considered as a novel potent agent for cerebral ischemia due to its effect in preventing neuronal cell death after ischemic stroke. In the present study, we investigated the post-ischemic neuroprotective effect of N2 and its underlying mechanisms. Using a MCAO rat model, we found that N2 reversed brain infarct size, reduced cerebral edema and decreased the neurological deficit score significantly. Moreover, N2 diminished TUNEL positive cells, down-regulated bax expression and up-regulated bcl-2 expression notably. In addition, we evaluated the oxygen glucose deprivation/reoxygenation (OGD/R) injury induced neuron cell death in rat primary cortical neuron and assessed the neuroprotective effect of our drug. N2 increased cell viability, ameliorated neuron cell injury by decreasing LDH activity, and inhibited cell apoptotic rate while suppressed apoptotic signaling via inhibiting the bax expression, and elevating the bcl-2 expression. Furthermore, the neuroprotective effect of N2 was associated with the PI3K/Akt pathway which was proved by the use of PI3K inhibitor LY294002. The combination of our findings disclosed that N2 can be used as an effective neuroprotective agent for ischemic stroke due to its significant effect on preventing neuronal cell death after cerebral ischemia both in vivo and in vitro and the effectiveness was dose dependent.

  12. Cobalt chloride stimulates phosphoinositide 3-kinase/Akt signaling through the epidermal growth factor receptor in oral squamous cell carcinoma.

    PubMed

    Ryu, Mi Heon; Park, Jeong Hee; Park, Ji Eun; Chung, Jin; Lee, Chang Hun; Park, Hae Ryoun

    2010-04-01

    Tumor cells are often found under hypoxic conditions due to the rapid outgrowth of their vascular supply, and, in order to survive hypoxia, these cells induce numerous signaling factors. Akt is an important kinase in cell survival, and its activity is regulated by the upstream phosphoinositide 3-kinase (PI3K) and receptor tyrosine kinases (RTKs). In this study, we examined Akt activation and RTKs/PI3K/Akt signaling using the hypoxia-mimetic cobalt chloride in oral squamous carcinoma cells. Cobalt chloride increases Akt phosphorylation in both a dose- and time-dependent manner. Blocking the activation of the PI3K/Akt pathway using LY294002 abolished Akt activation in response to cobalt chloride, suggesting that Akt phosphorylation by cobalt chloride is dependent on PI3K. In addition, activation of the PI3K/Akt pathway seems to rely on the epidermal growth factor receptor (EGFR), since the inhibition of EGFR attenuated cobalt chloride-induced Akt activation. The results in this study also demonstrate that cobalt chloride increases EGFR protein levels and induces oral squamous cell carcinoma cells to enter S phase.

  13. Testosterone regulation of Akt/mTORC1/FoxO3a Signaling in Skeletal Muscle

    PubMed Central

    White, James P.; Gao, Song; Puppa, Melissa J.; Sato, Shuichi; Welle, Stephen L.; Carson, James A.

    2012-01-01

    Low endogenous testosterone production, known as hypogonadism is commonly associated with conditions inducing muscle wasting. Akt signaling can control skeletal muscle mass through mTOR regulation of protein synthesis and FoxO regulation of protein degradation, and this pathway has been previously identified as a target of androgen signaling. However, the testosterone sensitivity of Akt/mTOR signaling requires further understanding in order to grasp the significance of varied testosterone levels seen with wasting disease on muscle protein turnover regulation. Therefore, the purpose of this study is to determine the effect of androgen availability on muscle Akt/mTORC1/FoxO3a regulation in skeletal muscle and cultured C2C12 myotubes. C57BL/6 mice were either castrated for 42 days or castrated and treated with the nandrolone decanoate (ND) (6 mg/kg bw/wk). Testosterone loss (TL) significantly decreased volitional grip strength, body weight, and gastrocnemius (GAS) muscle mass, and ND reversed these changes. Related to muscle mass regulation, TL decreased muscle IGF-1 mRNA, the rate of myofibrillar protein synthesis, Akt phosphorylation, and the phosphorylation of Akt targets, GSK3β, PRAS40 and FoxO3a. TL induced expression of FoxO transcriptional targets, MuRF1, atrogin1 and REDD1. Muscle AMPK and raptor phosphorylation, mTOR inhibitors, were not altered by low testosterone. ND restored IGF-1 expression and Akt/mTORC1 signaling while repressing expression of FoxO transcriptional targets. Testosterone (T) sensitivity of Akt/mTORC1 signaling was examined in C2C12 myotubes, and mTOR phosphorylation was induced independent of Akt activation at low T concentrations, while a higher T concentration was required to activate Akt signaling. Interestingly, low concentration T was sufficient to amplify myotube mTOR and Akt signaling after 24h of T withdrawal, demonstrating the potential in cultured myotubes for a T initiated positive feedback mechanism to amplify Akt

  14. Akt kinase C-terminal modifications control activation loop dephosphorylation and enhance insulin response

    PubMed Central

    Chan, Tung O.; Zhang, Jin; Tiegs, Brian C.; Blumhof, Brian; Yan, Linda; Keny, Nikhil; Penny, Morgan; Li, Xue; Pascal, John M.; Armen, Roger S.; Rodeck, Ulrich; Penn, Raymond B.

    2015-01-01

    The Akt protein kinase, also known as protein kinase B, plays key roles in insulin receptor signalling and regulates cell growth, survival and metabolism. Recently, we described a mechanism to enhance Akt phosphorylation that restricts access of cellular phosphatases to the Akt activation loop (Thr308 in Akt1 or protein kinase B isoform alpha) in an ATP-dependent manner. In the present paper, we describe a distinct mechanism to control Thr308 dephosphorylation and thus Akt deactivation that depends on intramolecular interactions of Akt C-terminal sequences with its kinase domain. Modifications of amino acids surrounding the Akt1 C-terminal mTORC2 (mammalian target of rapamycin complex 2) phosphorylation site (Ser473) increased phosphatase resistance of the phosphorylated activation loop (pThr308) and amplified Akt phosphorylation. Furthermore, the phosphatase-resistant Akt was refractory to ceramide-dependent dephosphorylation and amplified insulin-dependent Thr308 phosphorylation in a regulated fashion. Collectively, these results suggest that the Akt C-terminal hydrophobic groove is a target for the development of agents that enhance Akt phosphorylation by insulin. PMID:26201515

  15. Cytoplasmic localization of wild-type survivin is associated with constitutive activation of the PI3K/Akt signaling pathway and represents a favorable prognostic factor in patients with acute myeloid leukemia

    PubMed Central

    Serrano-López, Juana; Serrano, Josefina; Figueroa, Vianihuini; Torres-Gomez, Antonio; Tabares, Salvador; Casaño, Javier; Fernandez-Escalada, Noemi; Sánchez-Garcia, Joaquín

    2013-01-01

    Survivin is over-expressed in most hematologic malignancies but the prognostic significance of the subcompartmental distribution of wild-type or splicing variants in acute myeloid leukemia has not been addressed yet. Using western blotting, we assessed the expression of wild-type survivin and survivin splice variants 2B and Delta-Ex3 in nuclear and cytoplasmic protein extracts in samples taken from 105 patients at the time of their diagnosis of acute myeloid leukemia. Given that survivin is a downstream effector of the PI3K/Akt signaling pathway, survivin expression was also correlated with pSer473-Akt. Wild-type survivin and the 2B splice variant were positive in 76.3% and 78.0% of samples in the nucleus, cytoplasm or both, whereas the Delta-Ex3 isoform was only positive in the nucleus in 37.7% of samples. Cytoplasmic localization of wild-type survivin was significantly associated with the presence of high levels of pSer473-Akt (P<0.001). Inhibition of the PI3K/Akt pathway with wortmannin and Ly294002 caused a significant reduction in the expression of cytoplasmic wild-type survivin. The presence of cytoplasmic wild-type survivin and pSer473-Akt was associated with a lower fraction of quiescent leukemia stem cells (P=0.02). The presence of cytoplasmic wild-type survivin and pSer473-Akt were favorable independent prognostic factors. Moreover, the activation of the PI3K/Akt pathway with expression of cytoplasmic wild-type survivin identified a subgroup of acute myeloid leukemia patients with an excellent outcome (overall survival rate of 60.0±21.9% and relapse-free survival of 63.0±13.5%). Our findings suggest that cytoplasmic wild-type survivin is a critical downstream effector of the PI3K/Akt pathway leading to more chemosensitive cells and a more favorable outcome in acute myeloid leukemia. PMID:23812937

  16. Inhibitors beta-amyloid-induced toxicity by modulating the Akt signaling pathway.

    PubMed

    Nakagami, Yasuhiro

    2004-12-01

    The Akt signaling pathway plays a crucial role in neuronal survival, leading to inhibition of apoptosis. Many stimulants including neurotrophins are reported to activate this pathway in preclinical studies; however, there are no drugs for neurodegenerative diseases adopting such a concept on the market so far. Among neurodegenerative diseases, Alzheimer's disease is the most common and characterized by senile plaques and neurofibrillary tangles, which consist of beta-amyloid and hyperphosphorylated tau, respectively. Recent studies suggest that activation of Akt inhibits toxicity of beta-amyloid and formation of neurofibrillary tangles, leading to protection of neurons against apoptosis. This review discusses the possibility of treatment of Alzheimer's disease by activating the Akt signaling pathway.

  17. Exposure to Ionizing Radiation Causes Long-Term Increase in Serum Estradiol and Activation of PI3K-Akt Signaling Pathway in Mouse Mammary Gland

    SciTech Connect

    Suman, Shubhankar; Johnson, Michael D.; Fornace, Albert J.; Datta, Kamal

    2012-10-01

    Purpose: Exposure to ionizing radiation is an established risk factor for breast cancer. Radiation exposure during infancy, childhood, and adolescence confers the highest risk. Although radiation is a proven mammary carcinogen, it remains unclear where it acts in the complex multistage process of breast cancer development. In this study, we investigated the long-term pathophysiologic effects of ionizing radiation at a dose (2 Gy) relevant to fractionated radiotherapy. Methods and Materials: Adolescent (6-8 weeks old; n = 10) female C57BL/6J mice were exposed to 2 Gy total body {gamma}-radiation, the mammary glands were surgically removed, and serum and urine samples were collected 2 and 12 months after exposure. Molecular pathways involving estrogen receptor-{alpha} (ER{alpha}) and phosphatidylinositol-3-OH kinase (PI3K)-Akt signaling were investigated by immunohistochemistry and Western blot. Results: Serum estrogen and urinary levels of the oncogenic estrogen metabolite (16{alpha}OHE1) were significantly increased in irradiated animals. Immunostaining for the cellular proliferative marker Ki-67 and cyclin-D1 showed increased nuclear accumulation in sections of mammary glands from irradiated vs. control mice. Marked increase in p85{alpha}, a regulatory sub-unit of the PI3K was associated with increase in Akt, phospho-Akt, phospho-BAD, phospho-mTOR, and c-Myc in irradiated samples. Persistent increase in nuclear ER{alpha} in mammary tissues 2 and 12 months after radiation exposure was also observed. Conclusions: Taken together, our data not only support epidemiologic observations associating radiation and breast cancer but also, specify molecular events that could be involved in radiation-induced breast cancer.

  18. Functional proteomic analysis of a three-tier PKCepsilon-Akt-eNOS signaling module in cardiac protection.

    PubMed

    Zhang, Jun; Baines, Christopher P; Zong, Chenggong; Cardwell, Ernest M; Wang, Guangwu; Vondriska, Thomas M; Ping, Peipei

    2005-02-01

    Cardiac protective signaling networks have been shown to involve PKCepsilon. However, the molecular mechanisms by which PKCepsilon interacts with other members of these networks to form task-specific modules remain unknown. Among 93 different PKCepsilon-associated proteins that have been identified, Akt and endothelial nitric oxide (NO) synthase (eNOS) are of importance because of their independent abilities to promote cell survival and prevent cell death. The simultaneous association of PKCepsilon, Akt, and eNOS has not been examined, and, in particular, the formation of a module containing these three proteins and the role of such a module in the regulation of NO production and cardiac protection are unknown. The present study was undertaken to determine whether these molecules form a signaling module and, thereby, play a collective role in cardiac signaling. Using recombinant proteins in vitro and PKCepsilon transgenic mouse hearts, we demonstrate the following: 1) PKCepsilon, Akt, and eNOS interact and form signaling modules in vitro and in the mouse heart. Activation of either PKCepsilon or Akt enhances the formation of PKCepsilon-Akt-eNOS signaling modules. 2) PKCepsilon directly phosphorylates and enhances activation of Akt in vitro, and PKCepsilon activation increases phosphorylation and activation of Akt in PKCepsilon transgenic mouse hearts. 3) PKCepsilon directly phosphorylates eNOS in vitro, and this phosphorylation enhances eNOS activity. Activation of PKCepsilon in vivo increased phosphorylation of eNOS at Ser(1177), indicating eNOS activation. This study characterizes, for the first time, the physical, as well as functional, coupling of PKCepsilon, Akt, and eNOS in the heart and implicates these PKCepsilon-Akt-eNOS signaling modules as critical signaling elements during PKCepsilon-induced cardiac protection.

  19. Resveratrol Regulates Activated Hepatic Stellate Cells by Modulating NF-κB and the PI3K/Akt Signaling Pathway.

    PubMed

    Zhang, De-Quan; Sun, Peng; Jin, Quan; Li, Xia; Zhang, Yu; Zhang, Yu-Jing; Wu, Yan-Ling; Nan, Ji-Xing; Lian, Li-Hua

    2016-01-01

    In the present study, we investigated whether resveratrol could suppress the hepatic fibrogenesis in activated hepatic stellate cells. The immortalized rat hepatic stellate cells, t-HSC/Cl-6, were treated with resveratrol 1 h prior to lipopolysaccharide (LPS, 1 μg/mL). Resveratrol decreased t-HSC/Cl-6 cell viability at much lower concentrations within 24 h. Resveratrol pretreatment also decreased the LPS-induced protein expression of α-SMA and collagen I. In addition, resveratrol significantly reduced the protein expression of Toll-like receptor 4 (TLR4) and myeloid differentiation primary response gene 88 (MyD88), and the expression of phosphorylated phosphatidylinositol 3-kinase (PI3K) and phosphorylated serine/threonine kinase B (Akt). Moreover, resveratrol markedly blocked the translocation of nuclear factor (NF)-κB in LPS-activated HSCs. Furthermore, resveratrol inhibited HSCs activation through stimulating LXRβ, but did not influence LXRα. Overall, we conclude that the antifibrotic effect of resveratrol is the result of blocking NF-κB activation and PI3K/Akt phosphorylation, which inhibits HSC activation to obstruct liver fibrosis. Thus, resveratrol may be a natural agent for preventing hepatic fibrosis.

  20. O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling

    PubMed Central

    Shi, Jianhua; Gu, Jin-hua; Dai, Chun-ling; Gu, Jianlan; Jin, Xiaoxia; Sun, Jianming; Iqbal, Khalid; Liu, Fei; Gong, Cheng-Xin

    2015-01-01

    Apoptosis plays an important role in neural development and neurological disorders. In this study, we found that O-GlcNAcylation, a unique protein posttranslational modification with O-linked β-N-acetylglucosamine (GlcNAc), promoted apoptosis through attenuating phosphorylation/activation of AKT and Bad. By using co-immunoprecipitation and mutagenesis techniques, we identified O-GlcNAc modification at both Thr308 and Ser473 of AKT. O-GlcNAcylation-induced apoptosis was attenuated by over-expression of AKT. We also found a dynamic elevation of protein O-GlcNAcylation during the first four hours of cerebral ischemia, followed by continuous decline after middle cerebral artery occlusion (MCAO) in the mouse brain. The elevation of O-GlcNAcylation coincided with activation of cell apoptosis. Finally, we found a negative correlation between AKT phosphorylation and O-GlcNAcylation in ischemic brain tissue. These results indicate that cerebral ischemia induces a rapid increase of O-GlcNAcylation that promotes apoptosis through down-regulation of AKT activity. These findings provide a novel mechanism through which O-GlcNAcylation regulates ischemia-induced neuronal apoptosis through AKT signaling. PMID:26412745

  1. Inhibition of fatty acid synthase suppresses U-2 OS cell invasion and migration via downregulating the activity of HER2/PI3K/AKT signaling pathway in vitro

    SciTech Connect

    Wang, Tao Fang; Wang, Heng; Peng, Ai Fen; Luo, Qing Feng; Liu, Zhi Li; Zhou, Rong Ping; Gao, Song; Zhou, Yang; Chen, Wen Zhao

    2013-10-18

    Highlights: •We investigate the relationship between FASN and HER2 or p-HER2 by IHC in OS tissues. •We construct FASN-specific RNAi plasmid. •Inhibiting FASN down-regulates HER2/PI3K/AKT cell signaling in U-2 OS. •Inhibiting FASN blocks U-2 OS cell invasion and migration. -- Abstract: FASN plays an important role in the malignant phenotype of various tumors. Our previous studies show that inhibition FASN could induce apoptosis and inhibit proliferation in human osteosarcoma (OS) cell in vivo and vitro. The aim in this study was to investigate the effect of inhibition FASN on the activity of HER2/PI3K/AKT axis and invasion and migration of OS cell. The expression of FASN, HER2 and p-HER2(Y1248) proteins was detected by immunohistochemistry in OS tissues from 24 patients with pulmonary metastatic disease, and the relationship between FASN and p-HER2 as well as HER2 was investigated. The results showed that there was a positive correlation between FASN and HER2 as well as p-HER2 protein expression. The U-2 OS cells were transfected with either the FASN specific RNAi plasmid or the negative control RNAi plasmid. FASN mRNA was measured by RT-PCR. Western blot assays was performed to examine the protein expression of FASN, HER2, p-HER2(Y1248), PI3K, Akt and p-Akt (Ser473). Migration and invasion of cells were investigated by wound healing and transwell invasion assays. The results showed that the activity of HER2/PI3K/AKT signaling pathway was suppressed by inhibiting FASN. Meanwhile, the U-2OS cells migration and invasion were also impaired by inhibiting the activity of FASN/HER2/PI3K/AKT. Our results indicated that inhibition of FASN suppresses OS cell invasion and migration via down-regulation of the “HER2/PI3K/AKT” axis in vitro. FASN blocker may be a new therapeutic strategy in OS management.

  2. Non-immunosuppressive triazole-based small molecule induces anticancer activity against human hormone-refractory prostate cancers: the role in inhibition of PI3K/AKT/mTOR and c-Myc signaling pathways

    PubMed Central

    Chan, She-Hung; Hsu, Jui-Ling; Liu, Shih-Ping; Chan, Mei-Ling; Yu, Chia-Chun; Hsu, Lih-Ching; Chou, Yen-Lin; Chang, Wei-Ling; Hou, Duen-Ren; Guh, Jih-Hwa

    2016-01-01

    A series of triazole-based small molecules that mimic FTY720-mediated anticancer activity but minimize its immunosuppressive effect have been produced. SPS-7 is the most effective derivative displaying higher activity than FTY720 in anti-proliferation against human hormone-refractory prostate cancer (HRPC). It induced G1 arrest of cell cycle and subsequent apoptosis in thymidine block-mediated synchronization model. The data were supported by a decrease of cyclin D1 expression, a dramatic increase of p21 expression and an associated decrease in RB phosphorylation. c-Myc overexpression replenished protein levels of cyclin D1 indicating that c-Myc was responsible for cell cycle regulation. PI3K/Akt/mTOR signaling pathways through p70S6K- and 4EBP1-mediated translational regulation are critical to cell proliferation and survival. SPS-7 significantly inhibited this translational pathway. Overexpression of Myr-Akt (constitutively active Akt) completely abolished SPS-7-induced inhibitory effect on mTOR/p70S6K/4EBP1 signaling and c-Myc protein expression, suggesting that PI3K/Akt serves as a key upstream regulator. SPS-7 also demonstrated substantial anti-tumor efficacy in an in vivo xenograft study using PC-3 mouse model. Notably, FTY720 but not SPS-7 induced a significant immunosuppressive effect as evidenced by depletion of marginal zone B cells, down-regulation of sphingosine-1-phosphate receptors and a decrease in peripheral blood lymphocytes. In conclusion, the data suggest that SPS-7 is not an immunosuppressant while induces anticancer effect against HRPC through inhibition of Akt/mTOR/p70S6K pathwaysthat down-regulate protein levels of both c-Myc and cyclin D1, leading to G1 arrest of cell cycle and subsequent apoptosis. The data also indicate the potential of SPS-7 since PI3K/Akt signalingis responsive for the genomic alterations in prostate cancer. PMID:27769069

  3. Activation of endoplasmic reticulum stress promotes autophagy and apoptosis and reverses chemoresistance of human small cell lung cancer cells by inhibiting the PI3K/AKT/mTOR signaling pathway

    PubMed Central

    Yu, Xin-Shuang; Du, Juan; Fan, Yu-Jun; Liu, Feng-Jun; Cao, Li-Li; Liang, Ning; Xu, De-Guo; Zhang, Jian-Dong

    2016-01-01

    Objective This study aims to investigate the effects of endoplasmic reticulum stress (ERS) on autophagy, apoptosis and chemoresistance of human small cell lung cancer (SCLC) cells via the PI3K/AKT/mTOR signaling pathway. Results The expressions of ERS-related proteins (PEAK, eIF2α and CHOP) up-regulated, autophagy-related proteins (LC3, LC3-II and Beclin1) and apoptosis-related proteins (Bax and procaspase-3) down-regulated in NCI-H446 and H69 cells after tunicamycin treatment for 24 h. Compared with the blank group, the tunicamycin, BEZ235 and tunicamycin + BEZ235 groups exhibited decreased expressions of p-PI3K, p-AKT and p-mTOR, and increased expressions of autophagy-related proteins (LC3, LC3-II and Beclin1) and apoptosis proteins (Bax and procaspase-3), and the most obvious changes were observed in the tunicamycin + BEZ235 group. Materials and Methods CCK-8 assay was applied to select the best cell line from five SCLC cell lines (NCI-H446, H69, H526, H146 and H209). Finally, NCI-H446 and H69 cells were selected for further experiments. NCI-H446/CDDP and H69/CDDP were selected and divided into the blank group, tunicamycin (an ESR inducer) group, BEZ235 (inhibitors of PI3K/AKT/mTOR pathway) group and tunicamycin + BEZ235 group. Cell apoptosis was detected by flow cytometry. Autophagy was observed by fluorescence microscopy and flow cytometry. Western blotting was used to detect the expressions of ERS-related proteins, autophagy-related proteins, apoptosis-related proteins and PI3K/AKT/mTOR pathway-related proteins. Conclusions Our findings provide evidence that the activation of ERS could promote autophagy and apoptosis and reverse chemoresistance of human SCLC cells by inhibiting the PI3K/AKT/mTOR pathway. PMID:27765907

  4. The Asian-American variant of human papillomavirus type 16 exhibits higher activation of MAPK and PI3K/AKT signaling pathways, transformation, migration and invasion of primary human keratinocytes.

    PubMed

    Hochmann, Jimena; Sobrinho, João S; Villa, Luisa L; Sichero, Laura

    2016-05-01

    Asian-American (AA) HPV-16 variants are associated with higher risk of cancer. Abnormal activation of intracellular signaling play a critical role in cancer development and progression. Our aim was to elucidate mechanisms underlying the higher oncogenic potential attributed to AA variant. We evaluated activation of MAPK and PI3K/AKT pathways in primary human keratinocytes (PHKs) transduced with E6/E7 of three HPV-16 variants: E-P, AA, E-350G. Phenotypes examined included migration, anchorage independent growth and invasion. AA PHKs presented the highest levels of active proteins involved in all cascades analyzed: MAPK-ERK, MAPK-p38 and PI3K-AKT. AA PHKs were more efficient in promoting anchorage independent growth, and in stimulating cell migration and invasion. MEK1 inhibition decreased migration. The mesenchymal phenotype marker vimentin was increased in AA PHKs. Our results suggest that MEK1, ERK2, AKT2 hyperactivation influence cellular behavior by means of GSK-3b inactivation and EMT induction prompting AA immortalized PHKs to more efficiently surpass carcinogenesis steps.

  5. Nitric Oxide Synthase and Breast Cancer: Role of TIMP-1 in NO-mediated Akt Activation

    PubMed Central

    Ridnour, Lisa A.; Barasch, Kimberly M.; Windhausen, Alisha N.; Dorsey, Tiffany H.; Lizardo, Michael M.; Yfantis, Harris G.; Lee, Dong H.; Switzer, Christopher H.; Cheng, Robert Y. S.; Heinecke, Julie L.; Brueggemann, Ernst; Hines, Harry B.; Khanna, Chand; Glynn, Sharon A.; Ambs, Stefan; Wink, David A.

    2012-01-01

    Prediction of therapeutic response and cancer patient survival can be improved by the identification of molecular markers including tumor Akt status. A direct correlation between NOS2 expression and elevated Akt phosphorylation status has been observed in breast tumors. Tissue inhibitor matrix metalloproteinase-1 (TIMP-1) has been proposed to exert oncogenic properties through CD63 cell surface receptor pathway initiation of pro-survival PI3k/Akt signaling. We employed immunohistochemistry to examine the influence of TIMP-1 on the functional relationship between NOS2 and phosphorylated Akt in breast tumors and found that NOS2-associated Akt phosphorylation was significantly increased in tumors expressing high TIMP-1, indicating that TIMP-1 may further enhance NO-induced Akt pathway activation. Moreover, TIMP-1 silencing by antisense technology blocked NO-induced PI3k/Akt/BAD phosphorylation in cultured MDA-MB-231 human breast cancer cells. TIMP-1 protein nitration and TIMP-1/CD63 co-immunoprecipitation was observed at NO concentrations that induced PI3k/Akt/BAD pro-survival signaling. In the survival analysis, elevated tumor TIMP-1 predicted poor patient survival. This association appears to be mainly restricted to tumors with high NOS2 protein. In contrast, TIMP-1 did not predict poor survival in patient tumors with low NOS2 expression. In summary, our findings suggest that tumors with high TIMP-1 and NOS2 behave more aggressively by mechanisms that favor Akt pathway activation. PMID:22957045

  6. Pseudoginsenoside-F11 (PF11) exerts anti-neuroinflammatory effects on LPS-activated microglial cells by inhibiting TLR4-mediated TAK1/IKK/NF-κB, MAPKs and Akt signaling pathways.

    PubMed

    Wang, Xiaoxiao; Wang, Chunming; Wang, Jiming; Zhao, Siqi; Zhang, Kuo; Wang, Jingmin; Zhang, Wei; Wu, Chunfu; Yang, Jingyu

    2014-04-01

    Pseudoginsenoside-F11 (PF11), an ocotillol-type ginsenoside, has been shown to possess significant neuroprotective activity. Since microglia-mediated inflammation is critical for induction of neurodegeneration, this study was designed to investigate the effect of PF11 on activated microglia. PF11 significantly suppressed the release of ROS and proinflammatory mediators induced by LPS in a microglial cell line N9 including NO, PGE2, IL-1β, IL-6 and TNF-α. Moreover, PF11 inhibited interaction and expression of TLR4 and MyD88 in LPS-activated N9 cells, resulting in an inhibition of the TAK1/IKK/NF-κB signaling pathway. PF11 also inhibited the phosphorylation of Akt and MAPKs induced by LPS in N9 cells. Importantly, PF11 significantly alleviated the death of SH-SY5Y neuroblastoma cells and primary cortical neurons induced by the conditioned-medium from activated microglia. At last, the effect of PF11 on neuroinflammation was confirmed in vivo: PF11 mitigated the microglial activation and proinflammatory factors expression obviously in both cortex and hippocampus in mice injected intrahippocampally with LPS. These findings indicate that PF11 exerts anti-neuroinflammatory effects on LPS-activated microglial cells by inhibiting TLR4-mediated TAK1/IKK/NF-κB, MAPKs and Akt signaling pathways, suggesting its therapeutic implication for neurodegenerative disease associated with neuroinflammation.

  7. Cross Talk between the Akt and p38α Pathways in Macrophages Downstream of Toll-Like Receptor Signaling

    PubMed Central

    McGuire, Victoria A.; Gray, Alexander; Monk, Claire E.; Santos, Susana G.; Lee, Keunwook; Aubareda, Anna; Crowe, Jonathan; Ronkina, Natalia; Schwermann, Jessica; Batty, Ian H.; Leslie, Nick R.; Dean, Jonathan L. E.; O'Keefe, Stephen J.; Boothby, Mark; Gaestel, Matthias

    2013-01-01

    The stimulation of Toll-like receptors (TLRs) on macrophages by pathogen-associated molecular patterns (PAMPs) results in the activation of intracellular signaling pathways that are required for initiating a host immune response. Both phosphatidylinositol 3-kinase (PI3K)–Akt and p38 mitogen-activated protein kinase (MAPK) signaling pathways are activated rapidly in response to TLR activation and are required to coordinate effective host responses to pathogen invasion. In this study, we analyzed the role of the p38-dependent kinases MK2/3 in the activation of Akt and show that lipopolysaccharide (LPS)-induced phosphorylation of Akt on Thr308 and Ser473 requires p38α and MK2/3. In cells treated with p38 inhibitors or an MK2/3 inhibitor, phosphorylation of Akt on Ser473 and Thr308 is reduced and Akt activity is inhibited. Furthermore, BMDMs deficient in MK2/3 display greatly reduced phosphorylation of Ser473 and Thr308 following TLR stimulation. However, MK2/3 do not directly phosphorylate Akt in macrophages but act upstream of PDK1 and mTORC2 to regulate Akt phosphorylation. Akt is recruited to phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the membrane, where it is activated by PDK1 and mTORC2. Analysis of lipid levels in MK2/3-deficient bone marrow-derived macrophages (BMDMs) revealed a role for MK2/3 in regulating Akt activity by affecting availability of PIP3 at the membrane. These data describe a novel role for p38α-MK2/3 in regulating TLR-induced Akt activation in macrophages. PMID:23979601

  8. K-RAS(V12) Induces Autocrine Production of EGFR Ligands and Mediates Radioresistance Through EGFR-Dependent Akt Signaling and Activation of DNA-PKcs

    SciTech Connect

    Minjgee, Minjmaa; Toulany, Mahmoud; Kehlbach, Rainer; Giehl, Klaudia; Rodemann, H. Peter

    2011-12-01

    Purpose: It is known that postirradiation survival of tumor cells presenting mutated K-RAS is mediated through autocrine activation of epidermal growth factor receptor (EGFR). In this study the molecular mechanism of radioresistance of cells overexpressing mutated K-RAS(V12) was investigated. Methods and Materials: Head-and-neck cancer cells (FaDu) presenting wild-type K-RAS were transfected with empty vector or vector expressing mutated K-RAS(V12). The effect of K-RAS(V12) on autocrine production of EGFR ligands, activation of EGFR downstream pathways, DNA damage repair, and postirradiation survival was analyzed. Results: Conditioned medium collected from K-RAS(V12)-transfected cells enhanced activation of the phosphatidylinositol-3-kinase-Akt pathway and increased postirradiation survival of wild-type K-RAS parental cells when compared with controls. These effects were reversed by amphiregulin (AREG)-neutralizing antibody. In addition, secretion of the EGFR ligands AREG and transforming growth factor {alpha} was significantly increased upon overexpression of K-RAS(V12). Expression of mutated K-RAS(V12) resulted in an increase in radiation-induced DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylation at S2056. This increase was accompanied by increased repair of DNA double-strand breaks. Abrogation of DNA-PKcs phosphorylation by serum depletion or AREG-neutralizing antibody underscored the role of autocrine production of EGFR ligands, namely, AREG, in regulating DNA-PKcs activation in K-RAS mutated cells. Conclusions: These data indicate that radioresistance of K-RAS mutated tumor cells is at least in part due to constitutive production of EGFR ligands, which mediate enhanced repair of DNA double-strand breaks through the EGFR-phosphatidylinositol-3-kinase-Akt cascade.

  9. Cyclophilin A as a downstream effector of PI3K/Akt signalling pathway in multiple myeloma cells.

    PubMed

    Lin, Zuo-Lin; Wu, Hsin-Jou; Chen, Jin-An; Lin, Kuo-Chih; Hsu, Jung-Hsin

    2015-12-01

    Cyclophilin A (Cyp A), a member of the peptidyl-prolyl isomerase (PPI) family, may function as a molecular signalling switch. Comparative proteomic studies have identified Cyp A as a potential downstream target of protein kinase B (Akt). This study confirmed that Cyp A is a downstream effector of the phosphatidylinositide 3-kinase (PI3K)/Akt signalling pathway. Cyp A was highly phosphorylated in response to interleukin-6 treatment, which was consistent with the accumulation of phosphorylated Akt, suggesting that Cyp A is a phosphorylation target of Akt and downstream effector of the PI3K/Akt pathway. Cyclosporine A (CsA), a PPI inhibitor, inhibited the growth of multiple myeloma (MM) U266 cells. Moreover, CsA treatment inhibited the activation of the signal transducer and activator of transcription 3 (STAT3) in MM U266 cells. Several Cyp A mutants were generated. Mutants with mutated AKT phosphorylation sites increased the G1 phase arrest in MM U266 cells. The other mutants that mimicked the phosphorylated state of Cyp A decreased the percentage of G1 phase. These results demonstrated that the states of phosphorylation of Cyp A by Akt can influence the progress of the cell cycle in MM U266 cells and that this effect is probably mediated through the Janus-activated kinase 2/STAT3 signalling pathway.

  10. Gallic acid inhibits gastric cancer cells metastasis and invasive growth via increased expression of RhoB, downregulation of AKT/small GTPase signals and inhibition of NF-κB activity

    SciTech Connect

    Ho, Hsieh-Hsun; Chang, Chi-Sen; Ho, Wei-Chi; Liao, Sheng-You; Lin, Wea-Lung; Wang, Chau-Jong

    2013-01-01

    Our previous study demonstrated the therapeutic potential of gallic acid (GA) for controlling tumor metastasis through its inhibitory effect on the motility of AGS cells. A noteworthy finding in our previous experiment was increased RhoB expression in GA-treated cells. The aim of this study was to evaluate the role of RhoB expression on the inhibitory effects of GA on AGS cells. By applying the transfection of RhoB siRNA into AGS cells and an animal model, we tested the effect of GA on inhibition of tumor growth and RhoB expression. The results confirmed that RhoB-siRNA transfection induced GA to inhibit AGS cells’ invasive growth involving blocking the AKT/small GTPase signals pathway and inhibition of NF-κB activity. Finally, we evaluated the effect of GA on AGS cell metastasis by colonization of tumor cells in nude mice. It showed GA inhibited tumor cells growth via the expression of RhoB. These data support the inhibitory effect of GA which was shown to inhibit gastric cancer cell metastasis and invasive growth via increased expression of RhoB, downregulation of AKT/small GTPase signals and inhibition of NF-κB activity. Thus, GA might be a potential agent in treating gastric cancer. Highlights: ► GA could downregulate AKT signal via increased expression of RhoB. ► GA inhibits metastasis in vitro in gastric carcinoma. ► GA inhibits tumor growth in nude mice model.

  11. ROS-mediated Activation of AKT Induces Apoptosis Via pVHL in Prostate Cancer Cells

    PubMed Central

    Chetram, Mahandranauth A.; Bethea, Danaya A.; Jones, Kia J.; Don-Salu-Hewage, Ayesha S.; Odero-Marah, Valerie A.; Hinton, Cimona V.

    2013-01-01

    Reactive oxygen species (ROS) play a central role in oxidative stress, which leads to the onset of diseases, such as cancer. Furthermore, ROS contributes to the delicate balance between tumor cell survival and death. However, the mechanisms by which tumor cells decide to elicit survival or death signals during oxidative stress are not completely understood. We have previously reported that ROS enhanced tumorigenic functions in prostate cancer cells, such as transendothelial migration and invasion, which depended on CXCR4 and AKT signaling. Here, we report a novel mechanism by which ROS facilitated cell death through activation of AKT. We initially observed that ROS increased expression of phosphorylated AKT (p-AKT) in 22Rv1 human prostate cancer cells. The tumor suppressor PTEN, a negative regulator of AKT signaling, was rendered catalytically inactive through oxidation by ROS, although the expression levels remained consistent. Despite these events, cells still underwent apoptosis. Further investigation into apoptosis revealed that expression of the tumor suppressor pVHL increased, and contains a target site for p-AKT phosphorylation. pVHL and p-AKT associated in vitro, and knockdown of pVHL rescued HIF1α expression and the cells from apoptosis. Collectively, our study suggests that in the context of oxidative stress, p-AKT facilitated apoptosis by inducing pVHL function. PMID:23315288

  12. Melatonin inhibits AP-2β/hTERT, NF-κB/COX-2 and Akt/ERK and activates caspase/Cyto C signaling to enhance the antitumor activity of berberine in lung cancer cells.

    PubMed

    Lu, Jian-Jun; Fu, Lingyi; Tang, Zhipeng; Zhang, Changlin; Qin, Lijun; Wang, Jingshu; Yu, Zhenlong; Shi, Dingbo; Xiao, Xiangsheng; Xie, Fangyun; Huang, Wenlin; Deng, Wuguo

    2016-01-19

    Melatonin, a molecule produced throughout the animal and plant kingdoms, and berberine, a plant derived agent, both exhibit antitumor and multiple biological and pharmacological effects, but they have never been combined altogether for the inhibition of human lung cancers. In this study, we investigated the role and underlying mechanisms of melatonin in the regulation of antitumor activity of berberine in lung cancer cells. Treatment with melatonin effectively increased the berberine-mediated inhibitions of cell proliferation, colony formation and cell migration, thereby enhancing the sensitivities of lung cancer cells to berberine. Melatonin also markedly increased apoptosis induced by berberine. Further mechanism study showed that melatonin promoted the cleavage of caspse-9 and PARP, enhanced the inhibition of Bcl2, and triggered the releasing of cytochrome C (Cyto C), thereby increasing the berberine-induced apoptosis. Melatonin also enhanced the berberine-mediated inhibition of telomerase reverses transcriptase (hTERT) by down-regulating the expression of AP-2β and its binding on hTERT promoter. Moreover, melatonin enhanced the berberine-mediated inhibition of cyclooxygenase 2 (COX-2) by inhibiting the nuclear translocation of NF-κB and its binding on COX-2 promoter. Melatonin also increased the berberine-mediated inhibition of the phosphorylated Akt and ERK. Collectively, our results demonstrated that melatonin enhanced the antitumor activity of berberine by activating caspase/Cyto C and inhibiting AP-2β/hTERT, NF-κB/COX-2 and Akt/ERK signaling pathways. Our findings provide new insights in exploring the potential therapeutic strategies and novel targets for lung cancer treatment.

  13. Erythropoietin alleviates hepatic insulin resistance via PPARγ-dependent AKT activation

    PubMed Central

    Ge, Zhijuan; Zhang, Pengzi; Hong, Ting; Tang, Sunyinyan; Meng, Ran; Bi, Yan; Zhu, Dalong

    2015-01-01

    Erythropoietin (EPO) has beneficial effects on glucose metabolism and insulin resistance. However, the mechanism underlying these effects has not yet been elucidated. This study aimed to investigate how EPO affects hepatic glucose metabolism. Here, we report that EPO administration promoted phosphatidylinositol 3-kinase (PI3K)/AKT pathway activation in palmitic acid (PA)-treated HepG2 cells and in the liver of high-fat diet (HFD)-fed mice, whereas adenovirus-mediated silencing of the erythropoietin receptor (EPOR) blocked EPO-induced AKT signalling in HepG2 cells. Importantly, a peroxisome proliferator-activated receptor γ (PPARγ) antagonist and PPARγ small interfering RNA (siRNA) abrogated the EPO-induced increase in p-AKT in HepG2 cells. Lentiviral vector-mediated hepatic PPARγ silencing in HFD-fed C57BL/6 mice impaired EPO-mediated increases in glucose tolerance, insulin sensitivity and hepatic AKT activation. Furthermore, EPO activated the AMP-activated protein kinase (AMPK)/sirtuin 1 (SIRT1) signalling pathway, and AMPKα and SIRT1 knockdown each attenuated the EPO-induced PPARγ expression and deacetylation and PPARγ-dependent AKT activation in HepG2 cells. In summary, these findings suggest that PPARγ is involved in EPO/EPOR-induced AKT activation, and targeting the PPARγ/AKT pathway via EPO may have therapeutic implications for hepatic insulin resistance and type 2 diabetes. PMID:26643367

  14. Leukemia-associated mutations in SHIP1 inhibit its enzymatic activity, interaction with the GM-CSF receptor and Grb2, and its ability to inactivate PI3K/AKT signaling.

    PubMed

    Brauer, Helena; Strauss, Julia; Wegner, Wiebke; Müller-Tidow, Carsten; Horstmann, Martin; Jücker, Manfred

    2012-11-01

    The inositol 5-phosphatase SHIP1 is a negative regulator of the PI3K/AKT pathway, which is constitutively activated in 50-70% of acute myeloid leukemias (AML). Ten different missense mutations in SHIP1 have been described in 3% of AML patients suggesting a functional role of SHIP1 in AML. Here, we report the identification of two new SHIP1 mutations T162P and R225W that were detected in 2 and 1 out of 96 AML patients, respectively. The functional analysis of all 12 AML-associated SHIP1 mutations, one ALL-associated SHIP1 mutation (Q1076X) and a missense SNP (H1168Y) revealed that two mutations i.e. Y643H and P1039S abrogated the ability of SHIP1 to reduce constitutive PI3K/AKT signaling in Jurkat cells. The loss of function of SHIP1 mutant Y643H which is localized in the inositol phosphatase domain was due to a reduction of the specific activity by 84%. Because all other SHIP1 mutants had a normal enzymatic activity, we assumed that these SHIP1 mutants may be functionally impaired due to a loss of interaction with plasma membrane receptors or adapter proteins. In agreement with this model, we found that the SHIP1 mutant F28L located in the FLVR motif of the SH2 domain was incapable of binding tyrosine-phosphorylated proteins including the GM-CSF receptor and that the SHIP1 mutant Q1076X lost its ability to bind to the C-terminal SH3 domain of the adapter protein Grb2. In addition, SHIP1 mutant P1039S which does not reduce PI3K/AKT signaling anymore is located in a PXXP SH3 domain consensus binding motif suggesting that mutation of the conserved proline residue interferes with binding of SHIP1 to a so far unidentified SH3 domain containing protein. In summary, our data indicate that SHIP1 mutations detected in human leukemia patients impair the negative regulatory function of SHIP1 on PI3K/AKT signaling in leukemia cells either directly by reduced enzymatic activity or indirectly by disturbed protein interaction with tyrosine-phosphorylated membrane receptors or

  15. Activation of p53/p21/PUMA alliance and disruption of PI-3/Akt in multimodal targeting of apoptotic signaling cascades in cervical cancer cells by a pentacyclic triterpenediol from Boswellia serrata.

    PubMed

    Bhushan, Shashi; Malik, Fayaz; Kumar, Ajay; Isher, Harpreet Kaur; Kaur, Indu Pal; Taneja, Subhash Chandra; Singh, Jaswant

    2009-12-01

    Cervical carcinoma is a growing menace to women health worldwide. This study reports the apoptotic cell death in human cervical cancer HeLa and SiHa cells by a pentacyclic triterpenediol (TPD) from Boswellia serrata by a mechanism different from reported in HL-60 cells. It caused oxidative stress by early generation of nitric oxide and reactive oxygen species that robustly up regulated time-dependent expression of p53/p21/PUMA while conversely abrogating phosphatidylinositol-3-kinase (PI3K)/Akt pathways in parallel. TPD also decreased the expression of PI3K/pAkt, ERK1/2, NF-kappaB/Akt signaling cascades which coordinately contribute to cancer cell survival through these distinct pathways. The tumor suppressor p53 pathway predominantly activated by TPD further up-regulated PUMA, which concomitantly decreased the Bcl-2 level, caused mitochondrial membrane potential loss with attendant translocation of Bax and drp1 to mitochondria and release of pro-apoptotic factors such as cytochrome c and Smac/Diablo to cytosol leading to caspases-3 and -9 activation. In addition both the phospho-p53 and p21 were found to accumulate heavily in the nuclear fraction with attendant decrease in topoisomarase II and survivin levels. On the contrary, TPD did not affect the extrinsic signaling transduction pathway effectively through apical death receptors. Interestingly, N-acetyl cysteine, ascorbate and s-methylisothiourea (sMIT) rescued cells significantly from TPD induced DNA damage and caspases activation. TPD may thus find usefulness in managing and treating cervical cancer.

  16. Deuterohemin-AlaHisLys mitigates the symptoms of rats with non-insulin dependent diabetes mellitus by scavenging reactive oxygen species and activating the PI3-K/AKT signal transduction pathway.

    PubMed

    Lei, Liyan; Zhang, Guangji; Li, Pengfei; Zhang, Yuan; Guo, Youming; Zhang, Wenqi; Zhang, Wenbo; Hu, Bing; Wang, Liping

    2014-09-05

    Damage to pancreatic β-cells plays an important role in the development of type 2 diabetes, and oxidative stress is a likely contributor. In the present study, we investigated the effect of deuterohemin-AlaHisLys (DhHP-3), a microperoxidase-11 mimic, on rats with non-insulin dependent diabetes mellitus and examined the action mechanisms of DhHP-3. The induced hyperglycemia, glucose intolerance, and insulin resistance in diabetic rats were associated with increased oxidative stress and damage to pancreatic islets. DhHP-3 (3 mg/kg) ameliorated hyperglycemia and insulin resistance, protected pancreas islet, decreased the content of malondialdehyde, and increased the activity of superoxide dismutase in plasma and pancreatic tissue by reducing ROS levels. Furthermore, DhHP-3 stimulated the proliferation of INS-1 cells and inhibited apoptosis by activating the phosphatidylinositol 3-kinase/protein kinase B (PI3-K/AKT) signaling pathway. Our results demonstrated for the first time that DhHP-3 decreased blood glucose level in rats with non-insulin dependent diabetes mellitus, scavenged reactive oxygen species, activated the PI3-K/AKT signaling pathway, and protected pancreatic β-cells against apoptosis.

  17. Cervical spinal erythropoietin induces phrenic motor facilitation via ERK and Akt signaling

    PubMed Central

    Dale, Erica A.; Satriotomo, Irawan; Mitchell, Gordon S.

    2012-01-01

    Erythropoietin (EPO) is typically known for its role in erythropoiesis, but is also a potent neurotrophic/neuroprotective factor for spinal motor neurons. Another trophic factor regulated by Hypoxia-Inducible Factor-1, vascular endothelial growth factor (VEGF), signals via ERK and Akt activation to elicit long-lasting phrenic motor facilitation (pMF). Since EPO also signals via ERK and Akt activation, we tested the hypothesis that EPO elicits similar pMF. Using retrograde labeling and immunohistochemical techniques, we demonstrate in adult, male, Sprague-Dawley rats that EPO and its receptor, EPO-R, are expressed in identified phrenic motor neurons. Intrathecal EPO at C4 elicits long-lasting pMF; integrated phrenic nerve burst amplitude increased >90 min post-injection (63±12% baseline 90 min post-injection; p<0.001). EPO increased phosphorylation (and presumed activation) of ERK (1.6 fold vs controls; p<0.05) in phrenic motor neurons; EPO also increased pAkt (1.6 fold vs controls; p<0.05). EPO-induced pMF was abolished by the MEK/ERK inhibitor U0126 and the PI3 kinase/Akt inhibitor LY294002, demonstrating that ERK MAP kinases and Akt are both required for EPO-induced pMF. Pre-treatment with U0126 and LY294002 decreased both pERK and pAkt in phrenic motor neurons (p<0.05), indicating a complex interaction between these kinases. We conclude that EPO elicits spinal plasticity in respiratory motor control. Since EPO expression is hypoxia-sensitive, it may play a role in respiratory plasticity in conditions of prolonged or recurrent low oxygen. PMID:22539857

  18. IDH1 R132H Mutation Enhances Cell Migration by Activating AKT-mTOR Signaling Pathway, but Sensitizes Cells to 5-FU Treatment as NADPH and GSH Are Reduced

    PubMed Central

    Qiu, Jiangdong; Huang, Keting; Wu, Mindan; Xia, Chunlin

    2017-01-01

    Aim of study Mutations of isocitrate dehydrogenase 1 and 2 (IDH1 and IDH2) gene were recently discovered in vast majority of World Health Organization (WHO) grade II/III gliomas. This study is to understand the effects of IDH1 R132H mutation in gliomagenesis and to develop new strategies to treat glioma with IDH1 R132H mutation. Materials and methods Over expression of IDH1 R132H in U87MG cells was done by transfecting cells with IDH1 R132H plasmid. MTT assay, scratch repair assay and western blot were performed to study effects of IDH1 R132H mutation on cell proliferation, migration, regulating AKT-mTOR signaling pathway and cell death respectively. NADP+/NADPH and GSH quantification assays were performed to evaluate effects of IDH1 R132H mutation on the production of antioxidant NADPH and GSH. Results We found that over expression of IDH1 R132H mutation decreased cell proliferation consistent with previous reports; however, it increased cell migration and enhanced AKT-mTOR signaling pathway activation. Mutations in isocitrate dehydrogenase (IDH) 1 also change the function of the enzymes and cause them to produce 2-hydroxyglutarate and not produce NADPH. We tested the level of NADPH and GSH and demonstrated that IDH1 R132H mutant stable cells had significantly low NADPH and GSH level compared to control or IDH1 wild type stable cells. The reduced antioxidants (NADPH and GSH) sensitized U87MG cells with IDH R132H mutant to 5-FU treatment. Conclusion Our study highlights the important role of IHD1 R132H mutant in up- regulating AKT-mTOR signaling pathway and enhancing cell migration. Furthermore, we demonstrate that IDH1 R132H mutation affects cellular redox status and sensitizes gliomas cells with IDH1 R132H mutation to 5FU treatment. PMID:28052098

  19. Crosstalk between Akt/GSK3β signaling and dynamin-1 regulates clathrin-mediated endocytosis

    PubMed Central

    Reis, Carlos R; Chen, Ping-Hung; Srinivasan, Saipraveen; Aguet, François; Mettlen, Marcel; Schmid, Sandra L

    2015-01-01

    Clathrin-mediated endocytosis (CME) regulates signaling from the plasma membrane. Analysis of clathrin-coated pit (CCP) dynamics led us to propose the existence of a rate-limiting, regulatory step(s) that monitor the fidelity of early stages in CCP maturation. Here we show that nascent endocytic vesicles formed in mutant cells displaying rapid, dysregulated CME are defective in early endosomal trafficking, maturation and acidification, confirming the importance of this “checkpoint.” Dysregulated CME also alters EGF receptor signaling and leads to constitutive activation of the protein kinase Akt. Dynamin-1, which was thought to be neuron specific, is activated by the Akt/GSK3β signaling cascade in non-neuronal cells to trigger rapid, dysregulated CME. Acute activation of dynamin-1 in RPE cells by inhibition of GSK3β accelerates CME, alters CCP dynamics and, unexpectedly, increases the rate of CCP initiation. CRISPR-Cas9n-mediated knockout and reconstitution studies establish that dynamin-1 is activated by Akt/GSK3β signaling in H1299 non-small lung cancer cells. These findings provide direct evidence for an isoform-specific role for dynamin in regulating CME and reveal a feed-forward pathway that could link signaling from cell surface receptors to the regulation of CME. PMID:26139537

  20. Pharmacological manipulation of the akt signaling pathway regulates myxoma virus replication and tropism in human cancer cells.

    PubMed

    Werden, Steven J; McFadden, Grant

    2010-04-01

    Viruses have evolved an assortment of mechanisms for regulating the Akt signaling pathway to establish a cellular environment more favorable for viral replication. Myxoma virus (MYXV) is a rabbit-specific poxvirus that encodes many immunomodulatory factors, including an ankyrin repeat-containing host range protein termed M-T5 that functions to regulate tropism of MYXV for rabbit lymphocytes and certain human cancer cells. MYXV permissiveness in these human cancer cells is dependent upon the direct interaction between M-T5 and Akt, which has been shown to induce the kinase activity of Akt. In this study, an array of compounds that selectively manipulate Akt signaling was screened and we show that only a subset of Akt inhibitors significantly decreased the ability of MYXV to replicate in previously permissive human cancer cells. Furthermore, reduced viral replication efficiency was correlated with lower levels of phosphorylated Akt. In contrast, the PP2A-specific phosphatase inhibitor okadaic acid promoted increased Akt kinase activation and rescued MYXV replication in human cancer cells that did not previously support viral replication. Finally, phosphorylation of Akt at residue Thr308 was shown to dictate the physical interaction between Akt and M-T5, which then leads to phosphorylation of Ser473 and permits productive MYXV replication in these human cancer cells. The results of this study further characterize the mechanism by which M-T5 exploits the Akt signaling cascade and affirms this interaction as a major tropism determinant that regulates the replication efficiency of MYXV in human cancer cells.

  1. Optogenetic clustering of CNK1 reveals mechanistic insights in RAF and AKT signalling controlling cell fate decisions

    PubMed Central

    Fischer, Adrian; Warscheid, Bettina; Weber, Wilfried; Radziwill, Gerald

    2016-01-01

    Scaffold proteins such as the multidomain protein CNK1 orchestrate the signalling network by integrating and controlling the underlying pathways. Using an optogenetic approach to stimulate CNK1 uncoupled from upstream effectors, we identified selective clusters of CNK1 that either stimulate RAF-MEK-ERK or AKT signalling depending on the light intensity applied. OptoCNK1 implemented in MCF7 cells induces differentiation at low light intensity stimulating ERK activity whereas stimulation of AKT signalling by higher light intensity promotes cell proliferation. CNK1 clustering in response to increasing EGF concentrations revealed that CNK1 binds to RAF correlating with ERK activation at low EGF dose. At higher EGF dose active AKT binds to CNK1 and phosphorylates and inhibits RAF. Knockdown of CNK1 protects CNK1 from this AKT/RAF crosstalk. In C2 skeletal muscle cells CNK1 expression is induced with the onset of differentiation. Hence, AKT-bound CNK1 counteracts ERK stimulation in differentiated but not in proliferating cells. Ectopically expressed CNK1 facilitates C2 cell differentiation and knockdown of CNK1 impaired the transcriptional network underlying C2 cell differentiation. Thus, CNK1 expression, CNK1 clustering and the thereto related differential signalling processes decide on proliferation and differentiation in a cell type- and cell stage-dependent manner by orchestrating AKT and RAF signalling. PMID:27901111

  2. c-Src activation promotes nasopharyngeal carcinoma metastasis by inducing the epithelial-mesenchymal transition via PI3K/Akt signaling pathway: a new and promising target for NPC

    PubMed Central

    Lu, Jinping; Xia, Weixiong; Yu, Yahui; Peng, Yongjian; Wang, Li; Wang, Gang; Ye, Yanfang; Yang, Jing; Liang, Hu; Kang, Tiebang; Lv, Xing

    2016-01-01

    Aberrant activation of cellular Src (c-Src), a non-receptor tyrosine kinase, could promote cancer progression through activating its downstream signaling pathways. However, the roles of c-Src and phosphorylated-Src (p-Src) in nasopharyngeal carcinoma (NPC) progression are rarely investigated. Herein, we have identified high c-Src concentrations in the serum of NPC patients with distant metastasis using high-throughput protein microarrays. Levels of c-Src in serum and p-Src in human primary NPC samples were unfavorable independent prognostic factors for cancer-specific survival, disease-free survival, and distant metastasis-free survival. Depletion or inactivation of c-Src in NPC cells using sgRNA with CRISPR/Cas9 system or PP2 decreased cell viability, colony formation, migration and invasion in vitro and metastasis in vivo. In contrast, these malignancies could be up-regulated by overexpressed c-Src in a NPC cell line with low-metastasis potential. Furthermore, p-Src was involved in promoting NPC cell metastasis by inducing the epithelial-mesenchymal transition (EMT) process via activating the PI3K/Akt pathway and cytoskeleton remodeling. The p-Src-induced EMT process could be retarded by PP2, which mediated by down-regulating the PI3K/Akt pathway. In conclusion, elevated levels of c-Src in serum and p-Src in primary NPC tissue correlated with poor outcomes of NPC patients. And aberrant activation of c-Src facilitated NPC cells with malignant potential, especially metastasis ability, which mediated by the PI3K/Akt pathway activation and sequentially induced the EMT process. These findings unveiled a promising approach for targeted therapy of advanced NPC. PMID:27078847

  3. Akt and mTOR in B Cell Activation and Differentiation.

    PubMed

    Limon, Jose J; Fruman, David A

    2012-01-01

    Activation of phosphoinositide 3-kinase (PI3K) is required for B cell proliferation and survival. PI3K signaling also controls key aspects of B cell differentiation. Upon engagement of the B cell receptor (BCR), PI3K activation promotes Ca(2+) mobilization and activation of NFκB-dependent transcription, events which are essential for B cell proliferation. PI3K also initiates a distinct signaling pathway involving the Akt and mTOR serine/threonine kinases. It has been generally assumed that activation of Akt and mTOR downstream of PI3K is essential for B cell function. However, Akt and mTOR have complex roles in B cell fate decisions and suppression of this pathway can enhance certain B cell responses while repressing others. In this review we will discuss genetic and pharmacological studies of Akt and mTOR function in normal B cells, and in malignancies of B cell origin.

  4. MicroRNA-130a alleviates human coronary artery endothelial cell injury and inflammatory responses by targeting PTEN via activating PI3K/Akt/eNOS signaling pathway

    PubMed Central

    Song, Chun-Li; Liu, Bin; Shi, Yong-Feng; Liu, Ning; Yan, You-You; Zhang, Ji-Chang; Xue, Xin; Wang, Jin-Peng; Zhao, Zhuo; Liu, Jian-Gen; Li, Yang-Xue; Zhang, Xiao-Hao; Wu, Jun-Duo

    2016-01-01

    Our study aims to investigate the roles of microRNA-130a (miR-130a) in human coronary artery endothelial cells (HCAECs) injury and inflammatory responses by targeting PTEN through the PI3K/Akt/eNOS signaling pathway. HCAECs were treated with 1.0 mmol/L homocysteine (HCY) and assigned into eight groups: the blank group, the negative control (NC) group, the miR-130a mimics group, the miR-130a inhibitors group, the si-PTEN group, the Wortmannin group, the miR-130a inhibitors + si-PTEN group and the miR-130a mimics + Wortmannin group. Luciferase reporter gene assay was used to validate the relationship between miR-130a and PTEN. The expressions of miR-130a, PTEN and PI3K/Akt/eNOS signaling pathway-related proteins were detected by qRT-PCR assay and Western blotting. MTT assay and Hoechst 33258 staining were adopted to testify cell growth and apoptosis. The NO kit assay was used to detect the NO release. ELISA was conducted to measure serum cytokine levels. Luciferase reporter gene assay confirmed the target relationship between miR-130a and PTEN. Compared with the blank and NC groups, the miR-130a mimics and si-PTEN groups showed significant increases in the expressions of PI3K/Akt/eNOS signaling pathway-related proteins, cell viability and the NO release, while serum cytokine levels and cell apoptosis were decreased; by contrast, an opposite trend was observed in miR-130a inhibitors and Wortmannin groups. However, no significant difference was found in the miR-130a inhibitors + si-PTEN and miR-130a mimics + Wortmannin groups when compared with the blank group. These results indicate that miR-130a could alleviate HCAECs injury and inflammatory responses by down-regulating PTEN and activating PI3K/Akt/eNOS signaling pathway. PMID:27713121

  5. Orexins protect neuronal cell cultures against hypoxic stress: an involvement of Akt signaling.

    PubMed

    Sokołowska, Paulina; Urbańska, Anna; Biegańska, Kaja; Wagner, Waldemar; Ciszewski, Wojciech; Namiecińska, Magdalena; Zawilska, Jolanta B

    2014-01-01

    Orexins A and B are peptides produced mainly by hypothalamic neurons that project to numerous brain structures. We have previously demonstrated that rat cortical neurons express both types of orexin receptors, and their activation by orexins initiates different intracellular signals. The present study aimed to determine the effect of orexins on the Akt kinase activation in the rat neuronal cultures and the significance of that response in neurons subjected to hypoxic stress. We report the first evidence that orexins A and B stimulated Akt in cortical neurons in a concentration- and time-dependent manner. Orexin B more potently than orexin A increased Akt phosphorylation, but the maximal effect of both peptides on the kinase activation was very similar. Next, cultured cortical neurons were challenged with cobalt chloride, an inducer of reactive oxygen species and hypoxia-mediated signaling pathways. Under conditions of chemical hypoxia, orexins potently increased neuronal viability and protected cortical neurons against oxidative stress. Our results also indicate that Akt kinase plays an important role in the pro-survival effects of orexins in neurons, which implies a possible mechanism of the orexin-induced neuroprotection.

  6. GIV/girdin links vascular endothelial growth factor signaling to Akt survival signaling in podocytes independent of nephrin.

    PubMed

    Wang, Honghui; Misaki, Taro; Taupin, Vanessa; Eguchi, Akiko; Ghosh, Pradipta; Farquhar, Marilyn G

    2015-02-01

    Podocytes are critically involved in the maintenance of the glomerular filtration barrier and are key targets of injury in many glomerular diseases. Chronic injury leads to progressive loss of podocytes, glomerulosclerosis, and renal failure. Thus, it is essential to maintain podocyte survival and avoid apoptosis after acute glomerular injury. In normal glomeruli, podocyte survival is mediated via nephrin-dependent Akt signaling. In several glomerular diseases, nephrin expression decreases and podocyte survival correlates with increased vascular endothelial growth factor (VEGF) signaling. How VEGF signaling contributes to podocyte survival and prevents apoptosis remains unknown. We show here that Gα-interacting, vesicle-associated protein (GIV)/girdin mediates VEGF receptor 2 (VEGFR2) signaling and compensates for nephrin loss. In puromycin aminonucleoside nephrosis (PAN), GIV expression increased, GIV was phosphorylated by VEGFR2, and p-GIV bound and activated Gαi3 and enhanced downstream Akt2, mammalian target of rapamycin complex 1 (mTORC1), and mammalian target of rapamycin complex-2 (mTORC2) signaling. In GIV-depleted podocytes, VEGF-induced Akt activation was abolished, apoptosis was triggered, and cell migration was impaired. These effects were reversed by introducing GIV but not a GIV mutant that cannot activate Gαi3. Our data indicate that after PAN injury, VEGF promotes podocyte survival by triggering assembly of an activated VEGFR2/GIV/Gαi3 signaling complex and enhancing downstream PI3K/Akt survival signaling. Because of its important role in promoting podocyte survival, GIV may represent a novel target for therapeutic intervention in the nephrotic syndrome and other proteinuric diseases.

  7. Cyproheptadine-induced myeloma cell apoptosis is associated with inhibition of the PI3K/AKT signaling.

    PubMed

    Li, Jie; Cao, Biyin; Zhou, Shunye; Zhu, Jingyu; Zhang, Zubin; Hou, Tingjun; Mao, Xinliang

    2013-12-01

    Recent studies revealed that the anti-allergic cyproheptadine displays anti-blood cancer activity. However, its mechanism is still elusive. In this study, cyproheptadine was found to decrease the expression of anti-apoptotic proteins, including Bcl-2, Mcl-1, and XIAP. More importantly, cyproheptadine-induced apoptosis was accompanied by suppressing AKT activation in myeloma cells. In the subsequent study, cyproheptadine was found to inhibit insulin-like growth factor 1-triggered AKT activation in a time- and concentration-dependent manner. Specifically, cyproheptadine blocked AKT translocation from nuclei for phosphorylation. This inhibition led to suppressed activation of p70S6K and 4EBP1, two key downstream signaling proteins in the PI3K/AKT pathway. However, cyproheptadine did not display inhibition on activation of IGF-1R or STAT3, possible upstream signals of AKT activation. These results further demonstrated that cyproheptadine suppresses the PI3K/AKT signaling pathway, which is probably critical for cyproheptadine-induced MM cell apoptosis.

  8. Clusterin facilitates metastasis by EIF3I/Akt/MMP13 signaling in hepatocellular carcinoma

    PubMed Central

    Wang, Ning; Luo, Qin; Zhang, Yurong; Gao, Dongmei; Jiang, Kai; Gu, Dishui; Shen, Qiujing; Huo, Xisong; Hu, Fangyuan; Ge, Tianxiang; Zhao, Fangyu; Chu, Wei; Shu, Huiqun; Yao, Ming; Cong, Wenming; Qin, Wenxin

    2015-01-01

    Clusterin (CLU) is a stress-induced chaperone that confers proliferative and survival advantages to cancer cells. However, effects and molecular mechanisms of CLU in hepatocellular carcinoma (HCC) metastasis are still unknown. In this study, HCC tissue array (n = 198) was utilized to investigate correlation between CLU expression and clinicopathological features. Overexpression of CLU in HCC tissues was correlated with shorter overall survival and higher tumor recurrence. In vitro and in vivo assays demonstrated that silencing CLU attenuated the invasion and metastasis of HCC cells, whereas ectopic overexpression of CLU resulted in the forced metastasis of HCC cells. We also revealed that CLU activated Akt signaling through complexing with eukaryotic translation initiation factor 3 subunit I (EIF3I), which in turn promoted matrix metalloproteinase 13 (MMP13) expression and HCC metastasis. Positive correlations between CLU and MMP13, p-Akt, or EIF3I were found in HCC tissues. We further observed that CLU knockdown using the CLU inhibitor OGX-011 significantly suppressed HCC metastasis in two metastatic models through inhibiting EIF3I/Akt/MMP13 signaling. These findings indicate that CLU is an independent predictive factor for prognosis of HCC and it facilitates metastasis through EIF3I/Akt/MMP13 signaling. CLU suppression using OGX-011 may represent a promising therapeutic option for suppressing HCC metastasis. PMID:25609201

  9. Optogenetic activation reveals distinct roles of PIP3 and Akt in adipocyte insulin action.

    PubMed

    Xu, Yingke; Nan, Di; Fan, Jiannan; Bogan, Jonathan S; Toomre, Derek

    2016-05-15

    Glucose transporter 4 (GLUT4; also known as SLC2A4) resides on intracellular vesicles in muscle and adipose cells, and translocates to the plasma membrane in response to insulin. The phosphoinositide 3-kinase (PI3K)-Akt signaling pathway plays a major role in GLUT4 translocation; however, a challenge has been to unravel the potentially distinct contributions of PI3K and Akt (of which there are three isoforms, Akt1-Akt3) to overall insulin action. Here, we describe new optogenetic tools based on CRY2 and the N-terminus of CIB1 (CIBN). We used these 'Opto' modules to activate PI3K and Akt selectively in time and space in 3T3-L1 adipocytes. We validated these tools using biochemical assays and performed live-cell kinetic analyses of IRAP-pHluorin translocation (IRAP is also known as LNPEP and acts as a surrogate marker for GLUT4 here). Strikingly, Opto-PIP3 largely mimicked the maximal effects of insulin stimulation, whereas Opto-Akt only partially triggered translocation. Conversely, drug-mediated inhibition of Akt only partially dampened the translocation response of Opto-PIP3 In spatial optogenetic studies, focal targeting of Akt to a region of the cell marked the sites where IRAP-pHluorin vesicles fused, supporting the idea that local Akt-mediated signaling regulates exocytosis. Taken together, these results indicate that PI3K and Akt play distinct roles, and that PI3K stimulates Akt-independent pathways that are important for GLUT4 translocation.

  10. Role of Akt signaling in resistance to DNA-targeted therapy

    PubMed Central

    Avan, Abolfazl; Narayan, Ravi; Giovannetti, Elisa; Peters, Godefridus J

    2016-01-01

    The Akt signal transduction pathway controls most hallmarks of cancer. Activation of the Akt cascade promotes a malignant phenotype and is also widely implicated in drug resistance. Therefore, the modulation of Akt activity is regarded as an attractive strategy to enhance the efficacy of cancer therapy and irradiation. This pathway consists of phosphatidylinositol 3 kinase (PI3K), mammalian target of rapamycin, and the transforming serine-threonine kinase Akt protein isoforms, also known as protein kinase B. DNA-targeted agents, such as platinum agents, taxanes, and antimetabolites, as well as radiation have had a significant impact on cancer treatment by affecting DNA replication, which is aberrantly activated in malignancies. However, the caveat is that they may also trigger the activation of repairing mechanisms, such as upstream and downstream cascade of Akt survival pathway. Thus, each target can theoretically be inhibited in view of improving the potency of conventional treatment. Akt inhibitors, e.g., MK-2206 and perifosine, or PI3K modulators, e.g., LY294002 and Wortmannin, have shown some promising results in favor of sensitizing the cancer cells to the therapy in vitro and in vivo, which have provided the rationale for incorporation of these novel agents into multimodality treatment of different malignancies. Nevertheless, despite the acceptable safety profile of some of these agents in the clinical studies, with regard to the efficacy, the results are still too preliminary. Hence, we need to wait for the upcoming data from the ongoing trials before utilizing them into the standard care of cancer patients. PMID:27777878

  11. Connective tissue growth factor induces cardiac hypertrophy through Akt signaling

    SciTech Connect

    Hayata, Nozomi; Fujio, Yasushi; Yamamoto, Yasuhiro; Iwakura, Tomohiko; Obana, Masanori; Takai, Mika; Mohri, Tomomi; Nonen, Shinpei; Maeda, Makiko; Azuma, Junichi

    2008-05-30

    In the process of cardiac remodeling, connective tissue growth factor (CTGF/CCN2) is secreted from cardiac myocytes. Though CTGF is well known to promote fibroblast proliferation, its pathophysiological effects in cardiac myocytes remain to be elucidated. In this study, we examined the biological effects of CTGF in rat neonatal cardiomyocytes. Cardiac myocytes stimulated with full length CTGF and its C-terminal region peptide showed the increase in cell surface area. Similar to hypertrophic ligands for G-protein coupled receptors, such as endothelin-1, CTGF activated amino acid uptake; however, CTGF-induced hypertrophy is not associated with the increased expression of skeletal actin or BNP, analyzed by Northern-blotting. CTGF treatment activated ERK1/2, p38 MAPK, JNK and Akt. The inhibition of Akt by transducing dominant-negative Akt abrogated CTGF-mediated increase in cell size, while the inhibition of MAP kinases did not affect the cardiac hypertrophy. These findings indicate that CTGF is a novel hypertrophic factor in cardiac myocytes.

  12. Lithium potentiates GSK-3β activity by inhibiting phosphoinositide 3-kinase-mediated Akt phosphorylation

    SciTech Connect

    Tian, Nie; Kanno, Takeshi; Jin, Yu; Nishizaki, Tomoyuki

    2014-07-18

    Highlights: • Lithium suppresses Akt activity by reducing PI3K-mediated Akt phosphorylation. • Lithium enhances GSK-3β activity by reducing Akt-mediated GSK-3β phosphorylation. • Lithium suppresses GSK-3β activity through its direct inhibition. - Abstract: Accumulating evidence has pointed to the direct inhibitory action of lithium, an anti-depressant, on GSK-3β. The present study investigated further insight into lithium signaling pathways. In the cell-free assay Li{sub 2}CO{sub 3} significantly inhibited phosphoinositide 3-kinase (PI3K)-mediated phosphorylation of Akt1 at Ser473, but Li{sub 2}CO{sub 3} did not affect PI3K-mediated PI(3,4,5)P{sub 3} production and 3-phosphoinositide-dependent protein kinase 1 (PDK1)-mediated phosphorylation of Akt1 at Thr308. This indicates that lithium could enhance GSK-3β activity by suppressing Akt-mediated Ser9 phosphorylation of GSK-3β in association with inhibition of PI3K-mediated Akt activation. There was no direct effect of Li{sub 2}CO{sub 3} on Akt1-induced phosphorylation of GSK-3β at Ser9, but otherwise Li{sub 2}CO{sub 3} significantly reduced GSK-3β-mediated phosphorylation of β-catenin at Ser33/37 and Thr41. This indicates that lithium directly inhibits GSK-3β in an Akt-independent manner. In rat hippocampal slices Li{sub 2}CO{sub 3} significantly inhibited phosphorylation of Akt1/2 at Ser473/474, GSK-3β at Ser9, and β-catenin at Ser33/37 and Thr41. Taken together, these results indicate that lithium exerts its potentiating and inhibiting bidirectional actions on GSK-3β activity.

  13. Activation of PI3Kγ/Akt pathway increases cardiomyocyte HMGB1 expression in diabetic environment

    PubMed Central

    Song, Jia; Liu, Qian; Tang, Han; Tao, Aibin; Wang, Hao; Kao, Raymond; Rui, Tao

    2016-01-01

    Background The high mobility group box 1 (HMGB1) protein mediates the cardiomyocyte–cardiac fibroblast interaction that contributes to induction of myocardial fibrosis in diabetes mellitus (DM). In the present study, we aim to investigate the intracellular signaling pathway that leads to cardiomyocyte HMGB1 expression under a diabetic environment. Results HMGB1 expression is increased in high concentration of glucose (HG)-conditioned cardiomyocytes. Challenging cardiomyocytes with HG also increased PI3Kγ and Akt phosphorylation. Inhibition of PI3Kγ (CRISPR/Cas9 knockout plasmid or AS605240) prevented HG-induced Akt phosphorylation and HMGB1 expression by the cardiomyocytes. In addition, inhibition of Akt (Akt1/2/3 siRNA or A6730) attenuated HG-induced HMGB1 production. Finally, challenging cardiomyocytes with HG resulted in increased reactive oxygen species (ROS) production. Treatment of cardiomyocytes with an antioxidant (Mitotempo) abolished HG-induced PI3Kγ and Akt activation, as well as HMGB1 production. Materials and Methods Isolated rat cardiomyocytes were cultured with a high concentration of glucose. Cardiomyocyte phosphatidylinositol 3-kinase gamma (PI3Kγ) and Akt activation were determined by Western blot. Cardiomyocyte HMGB1 production was evaluated with Western blot and enzyme-linked immunosorbent assay (ELISA), while cardiomyocyte oxidative stress was determined with a DCFDA fluorescence probe. Conclusions Our results suggest that the cardiomyocytes incur an oxidative stress under diabetic condition, which subsequently activates the PI3Kγ/Akt cell-signaling pathway and further increases HMGB1 expression. PMID:27821807

  14. FANCI is a negative regulator of Akt activation.

    PubMed

    Zhang, Xiaoshan; Lu, Xiaoyan; Akhter, Shamima; Georgescu, Maria-Magdalena; Legerski, Randy J

    2016-01-01

    Akt is a critical mediator of the oncogenic PI3K pathway, and its activation is regulated by kinases and phosphatases acting in opposition. We report here the existence of a novel protein complex that is composed minimally of Akt, PHLPP1, PHLPP2, FANCI, FANCD2, USP1 and UAF1. Our studies show that depletion of FANCI, but not FANCD2 or USP1, results in increased phosphorylation and activation of Akt. This activation is due to a reduction in the interaction between PHLPP1 and Akt in the absence of FANCI. In response to DNA damage or growth factor treatment, the interactions between Akt, PHLPP1 and FANCI are reduced consistent with the known phosphorylation of Akt in response to these stimuli. Furthermore, depletion of FANCI results in reduced apoptosis after DNA damage in accord with its role as a negative regular of Akt. Our findings describe an unexpected function for FANCI in the regulation of Akt and define a previously unrecognized intersection between the PI3K-Akt and FA pathways.

  15. Exercise regulates Akt and glycogen synthase kinase-3 activities in human skeletal muscle.

    PubMed

    Sakamoto, Kei; Arnolds, David E W; Ekberg, Ingvar; Thorell, Anders; Goodyear, Laurie J

    2004-06-25

    Activation of Akt and deactivation of GSK3 are critical signals regulating a number of cellular processes in multiple systems. Whether physical exercise alters Akt and GSK3 activity in human skeletal muscle is controversial. beta-Catenin, a GSK3 substrate and important Wnt signaling protein that alters gene transcription, has not been investigated in human skeletal muscle. In the present study, eight healthy human subjects performed 30min of cycling exercise at 75% of maximum workload (submaximal) followed by 6 bouts of 60s at 125% maximum workload (maximal). Biopsies of vastus lateralis muscle were taken at rest (basal), and within 15s following cessation of the submaximal and maximal exercise bouts. Exercise at both submaximal and maximal intensities significantly increased Akt activity (40% and 110%, respectively). Increases in Akt activity were accompanied by increases in Akt Thr(308) and Ser(473) phosphorylation, decreased GSK3alpha activity ( approximately 30% at both intensities), and increased phosphorylation of GSK3alpha Ser(21). Exercise at both intensities also decreased beta-catenin Ser(33/37)Thr(41) phosphorylation (50-60% at both intensities). These results demonstrate that Akt, GSK3, and beta-catenin signaling are regulated by exercise in human skeletal muscle, and as such identify them as possible molecular mediators of exercise's effect on metabolic and transcriptional processes in skeletal muscle.

  16. Active form of AKT controls cell proliferation and response to apoptosis in hepatocellular carcinoma

    PubMed Central

    KUNTER, IMGE; ERDAL, ESRA; NART, DENIZ; YILMAZ, FUNDA; KARADEMIR, SEDAT; SAGOL, OZGUL; ATABEY, NESE

    2014-01-01

    Hepatocellular carcinoma (HCC) is the third most common cause of cancer-related mortality worldwide. Deregulation of the AKT signaling pathway has been found in HCC. However, the effect of AKT activation on the proliferation and apoptosis in HCC is not clear. Herein, expression of phosphorylated form of AKT (Ser 473) was investigated in HCC tumor (n=73), cirrhosis (n=17), normal liver (n=22) samples and in HCC cell lines (n=8). The results showed that expression of p-AKT was higher in tumor (53%) than in cirrhotic tissues (12%) while it was absent in normal liver (p<0.0001). p-AKT expression was also associated with number of tumor nodules and differentiation status (p<0.05). LY294002 induced cell cycle arrest at G0/G1 in SNU-449 and Mahlavu cells by decreasing expression of CDK2, CDK4, CycD1, CycD3, CycE, CycA and increasing expression of p21 and p27 as well; it also caused a decrease in the E2F1 transcriptional activity through declining phosphorylated Rb. LY294002 did not affect the basal level of apoptosis; however, it amplified cisplatin-induced apoptosis in SNU-449 cells. When the p-AKT level was decreased specifically after transfection with the DN-AKT plasmid, SNU-449 cells became more sensitive to cisplatin-induced apoptosis. HuH-7 cells with no basal p-AKT, were markedly affected by the treatment of doxorubicin. Thus, Akt signaling controls growth and chemical-induced apoptosis in HCC and p-AKT may be a potential target for therapeutic interventions in HCC patients. PMID:24337632

  17. Myogenic Akt signaling attenuates muscular degeneration, promotes myofiber regeneration and improves muscle function in dystrophin-deficient mdx mice

    PubMed Central

    Kim, Michelle H.; Kay, Danielle I.; Rudra, Renuka T.; Chen, Bo Ming; Hsu, Nigel; Izumiya, Yasuhiro; Martinez, Leonel; Spencer, Melissa J.; Walsh, Kenneth; Grinnell, Alan D.; Crosbie, Rachelle H.

    2011-01-01

    Duchenne muscular dystrophy, the most common form of childhood muscular dystrophy, is caused by X-linked inherited mutations in the dystrophin gene. Dystrophin deficiencies result in the loss of the dystrophin–glycoprotein complex at the plasma membrane, which leads to structural instability and muscle degeneration. Previously, we induced muscle-specific overexpression of Akt, a regulator of cellular metabolism and survival, in mdx mice at pre-necrotic (<3.5 weeks) ages and demonstrated upregulation of the utrophin–glycoprotein complex and protection against contractile-induced stress. Here, we found that delaying exogenous Akt treatment of mdx mice after the onset of peak pathology (>6 weeks) similarly increased the abundance of compensatory adhesion complexes at the extrasynaptic sarcolemma. Akt introduction after onset of pathology reverses the mdx histopathological measures, including decreases in blood serum albumin infiltration. Akt also improves muscle function in mdx mice as demonstrated through in vivo grip strength tests and in vitro contraction measurements of the extensor digitorum longus muscle. To further explore the significance of Akt in myofiber regeneration, we injured wild-type muscle with cardiotoxin and found that Akt induced a faster regenerative response relative to controls at equivalent time points. We demonstrate that Akt signaling pathways counteract mdx pathogenesis by enhancing endogenous compensatory mechanisms. These findings provide a rationale for investigating the therapeutic activation of the Akt pathway to counteract muscle wasting. PMID:21245083

  18. Glycitin regulates osteoblasts through TGF-β or AKT signaling pathways in bone marrow stem cells

    PubMed Central

    Zhang, Liyan; Chen, Jiying; Chai, Wei; Ni, Min; Sun, Xin; Tian, Dan

    2016-01-01

    The aim of the present study was to examine the effect of glycitin on the regulation of osteoblasts from bone marrow stem cells (BMSCs) through transforming growth factor (TGF)-β or protein kinase B (AKT) signaling pathways. BMSCs were extracted from New Zealand white rabbits and used to analyze the effect of glycitin on BMSCs. BMSCs were cleared using xylene and observed via light microscopy. BMSCs were subsequently induced with glycitin (0.01, 0.5, 1, 5 and 10 µM) for 7 days, and stained with Oil Red O. The mechanism of action of glycitin on BMSCs was investigated, in which contact with collagen type I (Col I), alkaline phosphatase (ALP), TGF-β and AKT was studied. Firstly, BMSCs appeared homogeneously mazarine blue, and which showed that BMSCs were successful extracted. Administration of glycitin increased cell proliferation and promoted osteoblast formation from BMSCs. Furthermore, glycitin activated the gene expression of Col I and ALP in BMSCs. Notably, glycitin suppressed protein expression of TGF-β and AKT in BMSCs. These results indicated that glycitin may regulate osteoblasts through TGF-β or AKT signaling pathways in BMSCs. PMID:27882117

  19. Tetramethylpyrazine Analogue CXC195 Protects Against Dopaminergic Neuronal Apoptosis via Activation of PI3K/Akt/GSK3β Signaling Pathway in 6-OHDA-Induced Parkinson's Disease Mice.

    PubMed

    Chen, Lin; Cheng, Li; Wei, Xinbing; Yuan, Zheng; Wu, Yanmei; Wang, Shuaishuai; Ren, Zhiping; Liu, Xinyong; Liu, Huiqing

    2016-12-22

    Parkinson's disease (PD) is a progressive neurodegenerative disorder and characterized by motor system disorders resulting in loss of dopaminergic (DA) neurons. CXC195, a novel tetramethylpyrazine derivative, has been shown strongest neuroprotective effects due to its anti-apoptotic activity. However, whether CXC195 protects against DA neuronal damage in PD and the mechanisms underlying its beneficial effects are unknown. The purpose of our study was to investigate the potential neuroprotective role of CXC195 and to elucidate its mechanism of action against 6-hydroxydopamine (6-OHDA)-induced mouse model of PD. CXC195 administration improved DA neurodegeneration in PD mice induced by 6-OHDA. Our further findings confirmed treatment of CXC195 at the dose of 10 mg/kg significantly inhibited the apoptosis by decreasing the level of cleaved caspase-3 and Bax, and increasing the level of Bcl-2 in 6-OHDA-lesioned mice. Meanwhile, 6-OHDA also decreased the amount of phosphorylated Akt while increased GSK-3β activity (the amount of phosphorylated GSK-3β at Ser9 was decreased) which was prevented by CXC195. Wortmannin, a specific PI3K inhibitor, dramatically abolished the changes induced by CXC195. Our study firstly demonstrated that CXC195 protected against DA neurodegeneration in 6-OHDA-induced PD model by its anti-apoptotic properties and PI3K/Akt/GSK3β signaling pathway was involved in it.

  20. Computational Modeling of PI3K/AKT and MAPK Signaling Pathways in Melanoma Cancer

    PubMed Central

    Pappalardo, Francesco; Russo, Giulia; Candido, Saverio; Pennisi, Marzio; Cavalieri, Salvatore; Motta, Santo; McCubrey, James A.; Nicoletti, Ferdinando; Libra, Massimo

    2016-01-01

    Background Malignant melanoma is an aggressive tumor of the skin and seems to be resistant to current therapeutic approaches. Melanocytic transformation is thought to occur by sequential accumulation of genetic and molecular alterations able to activate the Ras/Raf/MEK/ERK (MAPK) and/or the PI3K/AKT (AKT) signalling pathways. Specifically, mutations of B-RAF activate MAPK pathway resulting in cell cycle progression and apoptosis prevention. According to these findings, MAPK and AKT pathways may represent promising therapeutic targets for an otherwise devastating disease. Result Here we show a computational model able to simulate the main biochemical and metabolic interactions in the PI3K/AKT and MAPK pathways potentially involved in melanoma development. Overall, this computational approach may accelerate the drug discovery process and encourages the identification of novel pathway activators with consequent development of novel antioncogenic compounds to overcome tumor cell resistance to conventional therapeutic agents. The source code of the various versions of the model are available as S1 Archive. PMID:27015094

  1. EGFR trans-activation mediates pleiotrophin-induced activation of Akt and Erk in cultured osteoblasts.

    PubMed

    Fan, Jian-Bo; Liu, Wei; Yuan, Kun; Zhu, Xin-Hui; Xu, Da-Wei; Chen, Jia-Jia; Cui, Zhi-Ming

    2014-05-09

    Pleiotrophin (Ptn) plays an important role in bone growth through regulating osteoblasts' functions. The underlying signaling mechanisms are not fully understood. In the current study, we found that Ptn induced heparin-binding epidermal growth factor (HB-EGF) release to trans-activate EGF-receptor (EGFR) in both primary osteoblasts and osteoblast-like MC3T3-E1 cells. Meanwhile, Ptn activated Akt and Erk signalings in cultured osteoblasts. The EGFR inhibitor AG1478 as well as the monoclonal antibody against HB-EGF (anti-HB-EGF) significantly inhibited Ptn-induced EGFR activation and Akt and Erk phosphorylations in MC3T3-E1 cells and primary osteoblasts. Further, EGFR siRNA depletion or dominant negative mutation suppressed also Akt and Erk activation in MC3T3-E1 cells. Finally, we observed that Ptn increased alkaline phosphatase (ALP) activity and inhibited dexamethasone (Dex)-induced cell death in both MC3T3-E1 cells and primary osteoblasts, such effects were alleviated by AG1478 or anti-HB-EGF. Together, these results suggest that Ptn-induced Akt/Erk activation and some of its pleiotropic functions are mediated by EGFR trans-activation in cultured osteoblasts.

  2. The Histamine H3 Receptor Differentially Modulates Mitogen-activated Protein Kinase (MAPK) and Akt Signaling in Striatonigral and Striatopallidal Neurons.

    PubMed

    Rapanelli, Maximiliano; Frick, Luciana R; Horn, Kyla D; Schwarcz, Rivka C; Pogorelov, Vladimir; Nairn, Angus C; Pittenger, Christopher

    2016-09-30

    The basal ganglia have a central role in motor patterning, habits, motivated behaviors, and cognition as well as in numerous neuropsychiatric disorders. Receptors for histamine, especially the H3 receptor (H3R), are highly expressed in the striatum, the primary input nucleus of the basal ganglia, but their effects on this circuitry have been little explored. H3R interacts with dopamine (DA) receptors ex vivo; the nature and functional importance of these interactions in vivo remain obscure. We found H3R activation with the agonist R-(-)-α-methylhistamine to produce a unique time- and cell type-dependent profile of molecular signaling events in the striatum. H3 agonist treatment did not detectably alter extracellular DA levels or signaling through the cAMP/DARPP-32 signaling pathway in either D1- or D2-expressing striatal medium spiny neurons (MSNs). In D1-MSNs, H3 agonist treatment transiently activated MAPK signaling and phosphorylation of rpS6 and led to phosphorylation of GSK3β-Ser(9), a novel effect. Consequences of H3 activation in D2-MSNs were completely different. MAPK signaling was unchanged, and GSK3β-Ser(9) phosphorylation was reduced. At the behavioral level, two H3 agonists had no significant effect on locomotion or stereotypy, but they dramatically attenuated the locomotor activation produced by the D1 agonist SKF82958. H3 agonist co-administration blocked the activation of MAPK signaling and the phosphorylation of rpS6 produced by D1 activation in D1-MSNs, paralleling behavioral effects. In contrast, GSK3β-Ser(9) phosphorylation was seen only after H3 agonist treatment, with no interactive effects. H3R signaling has been neglected in models of basal ganglia function and has implications for a range of pathophysiologies.

  3. PI3K/AKT Signaling Pathway Is Essential for Survival of Induced Pluripotent Stem Cells

    PubMed Central

    Hossini, Amir M.; Quast, Annika S.; Plötz, Michael; Grauel, Katharina; Exner, Tarik; Küchler, Judit; Stachelscheid, Harald; Eberle, Jürgen; Rabien, Anja

    2016-01-01

    Apoptosis is a highly conserved biochemical mechanism which is tightly controlled in cells. It contributes to maintenance of tissue homeostasis and normally eliminates highly proliferative cells with malignant properties. Induced pluripotent stem cells (iPSCs) have recently been described with significant functional and morphological similarities to embryonic stem cells. Human iPSCs are of great hope for regenerative medicine due to their broad potential to differentiate into specialized cell types in culture. They may be useful for exploring disease mechanisms and may provide the basis for future cell-based replacement therapies. However, there is only poor insight into iPSCs cell signaling as the regulation of apoptosis. In this study, we focused our attention on the apoptotic response of Alzheimer fibroblast-derived iPSCs and two other Alzheimer free iPSCs to five biologically relevant kinase inhibitors as well as to the death ligand TRAIL. To our knowledge, we are the first to report that the relatively high basal apoptotic rate of iPSCs is strongly suppressed by the pancaspase inhibitor QVD-Oph, thus underlining the dependency on proapoptotic caspase cascades. Furthermore, wortmannin, an inhibitor of phosphoinositid-3 kinase / Akt signaling (PI3K-AKT), dramatically and rapidly induced apoptosis in iPSCs. In contrast, parental fibroblasts as well as iPSC-derived neuronal cells were not responsive. The resulting condensation and fragmentation of DNA and decrease of the membrane potential are typical features of apoptosis. Comparable effects were observed with an AKT inhibitor (MK-2206). Wortmannin resulted in disappearance of phosphorylated AKT and activation of the main effector caspase-3 in iPSCs. These results clearly demonstrate for the first time that PI3K-AKT represents a highly essential survival signaling pathway in iPSCs. The findings provide improved understanding on the underlying mechanisms of apoptosis regulation in iPSCs. PMID:27138223

  4. Upregulation of Akt/NF-κB-regulated inflammation and Akt/Bad-related apoptosis signaling pathway involved in hepatic carcinoma process: suppression by carnosic acid nanoparticle.

    PubMed

    Tang, Bo; Tang, Fang; Wang, Zhenran; Qi, Guangying; Liang, Xingsi; Li, Bo; Yuan, Shengguang; Liu, Jie; Yu, Shuiping; He, Songqing

    Primary liver cancer is globally the sixth most frequent cancer, and the second leading cause of cancer death and its incidence is increasing in many countries, becoming a serious threat to human health. Many researches focused on the treatment and prevention of liver cancer. However, due to the underlying molecular mechanism of liver cancer still not fully understood, the studies and development of treatments were forced to be delayed. Akt has been suggested to play an essential role in the progression of inflammation response and apoptosis. Hence, in this study, Akt-knockout mice and cells of liver cancer were used as a model to investigate the molecular mechanism of Akt-associated inflammatory and apoptotic signaling pathway linked with NF-κB and Bcl-2-associated death promoter (Bad) for the progression of liver cancer. Carnosic acid (CA), as a phenolic diterpene with anticancer, antibacterial, antidiabetic, as well as neuroprotective properties, is produced by many species from Lamiaceae family. Administration of CA nanoparticles was sufficient to lead to considerable inhibition of liver cancer progression. The results indicated that, compared to the normal liver cells, the expression of Akt was significantly higher in liver cancer cell lines. Also, we found that Akt-knockout cancer cell lines modulated inflammation response and apoptosis via inhibiting NF-κB activation and inducing apoptotic reaction. Our results indicated that the downstream signals, including cytokines regulated by NF-κB and caspase-3-activated apoptosis affected by Bad, were re-modulated for knockout of Akt. And CA nanoparticles, acting as Akt-knockout, could inhibit inflammation and accelerate apoptosis in liver cancer by altering NF-κB activation and activating caspase-3 through Bad pathway. These findings demonstrated that the nanoparticulate drug CA performed its effective role owing to its ability to reduce inflammatory action and enhance apoptosis for the overexpression of NF

  5. Upregulation of Akt/NF-κB-regulated inflammation and Akt/Bad-related apoptosis signaling pathway involved in hepatic carcinoma process: suppression by carnosic acid nanoparticle

    PubMed Central

    Tang, Bo; Tang, Fang; Wang, Zhenran; Qi, Guangying; Liang, Xingsi; Li, Bo; Yuan, Shengguang; Liu, Jie; Yu, Shuiping; He, Songqing

    2016-01-01

    Primary liver cancer is globally the sixth most frequent cancer, and the second leading cause of cancer death and its incidence is increasing in many countries, becoming a serious threat to human health. Many researches focused on the treatment and prevention of liver cancer. However, due to the underlying molecular mechanism of liver cancer still not fully understood, the studies and development of treatments were forced to be delayed. Akt has been suggested to play an essential role in the progression of inflammation response and apoptosis. Hence, in this study, Akt-knockout mice and cells of liver cancer were used as a model to investigate the molecular mechanism of Akt-associated inflammatory and apoptotic signaling pathway linked with NF-κB and Bcl-2-associated death promoter (Bad) for the progression of liver cancer. Carnosic acid (CA), as a phenolic diterpene with anticancer, antibacterial, antidiabetic, as well as neuroprotective properties, is produced by many species from Lamiaceae family. Administration of CA nanoparticles was sufficient to lead to considerable inhibition of liver cancer progression. The results indicated that, compared to the normal liver cells, the expression of Akt was significantly higher in liver cancer cell lines. Also, we found that Akt-knockout cancer cell lines modulated inflammation response and apoptosis via inhibiting NF-κB activation and inducing apoptotic reaction. Our results indicated that the downstream signals, including cytokines regulated by NF-κB and caspase-3-activated apoptosis affected by Bad, were re-modulated for knockout of Akt. And CA nanoparticles, acting as Akt-knockout, could inhibit inflammation and accelerate apoptosis in liver cancer by altering NF-κB activation and activating caspase-3 through Bad pathway. These findings demonstrated that the nanoparticulate drug CA performed its effective role owing to its ability to reduce inflammatory action and enhance apoptosis for the overexpression of NF

  6. IFNγ-induced suppression of β-catenin signaling: evidence for roles of Akt and 14.3.3ζ

    PubMed Central

    Nava, Porfirio; Kamekura, Ryuta; Quirós, Miguel; Medina-Contreras, Oscar; Hamilton, Ross W.; Kolegraff, Keli N.; Koch, Stefan; Candelario, Aurora; Romo-Parra, Hector; Laur, Oskar; Hilgarth, Roland S.; Denning, Timothy L.; Parkos, Charles A.; Nusrat, Asma

    2014-01-01

    The proinflammatory cytokine interferon γ (IFNγ ) influences intestinal epithelial cell (IEC) homeostasis in a biphasic manner by acutely stimulating proliferation that is followed by sustained inhibition of proliferation despite continued mucosal injury. β-Catenin activation has been classically associated with increased IEC proliferation. However, we observed that IFNγ inhibits IEC proliferation despite sustained activation of Akt/β-catenin signaling. Here we show that inhibition of Akt/β-catenin–mediated cell proliferation by IFNγ is associated with the formation of a protein complex containing phosphorylated β-catenin 552 (pβ-cat552) and 14.3.3ζ. Akt1 served as a bimodal switch that promotes or inhibits β-catenin transactivation in response to IFNγ stimulation. IFNγ initially promotes β-catenin transactivation through Akt-dependent C-terminal phosphorylation of β-catenin to promote its association with 14.3.3ζ. Augmented β-catenin transactivation leads to increased Akt1 protein levels, and active Akt1 accumulates in the nucleus, where it phosphorylates 14.3.3ζ to translocate 14.3.3ζ/β-catenin from the nucleus, thereby inhibiting β-catenin transactivation and IEC proliferation. These results outline a dual function of Akt1 that suppresses IEC proliferation during intestinal inflammation. PMID:25079689

  7. Cyclic Compressive Stress Regulates Apoptosis in Rat Osteoblasts: Involvement of PI3K/Akt and JNK MAPK Signaling Pathways

    PubMed Central

    Jiang, Dawei; Wang, Tianchen; Zhang, Yinquan; Ma, Hui

    2016-01-01

    It is widely accepted that physiological mechanical stimulation suppresses apoptosis and induces synthesis of extracellular matrix by osteoblasts; however, the effect of stress overloading on osteoblasts has not been fully illustrated. In the present study, we investigated the effect of cyclic compressive stress on rat osteoblasts apoptosis, using a novel liquid drop method to generate mechanical stress on osteoblast monolayers. After treatment with different levels of mechanical stress, apoptosis of osteoblasts and activations of mitogen-activated protein kinases (MAPKs) and PI3-kinase (PI3K)/Akt signaling pathways were investigated. Osteoblasts apoptosis was observed after treated with specific inhibitors prior to mechanical stimulation. Protein levels of Bax/Bcl-2/caspase-3 signaling were determined using western blot with or without inhibitors of PI3K/Akt and phosphorylation of c-jun N-terminal kinase (JNK) MAPK. Results showed that mechanical stimulation led to osteoblasts apoptosis in a dose-dependent manner and a remarkable activation of MAPKs and PI3K/Akt signaling pathways. Activation of PI3K/Akt protected against apoptosis, whereas JNK MAPK increased apoptosis via regulation of Bax/Bcl-2/caspase-3 activation. In summary, the PI3K/Akt and JNK MAPK signaling pathways played opposing roles in osteoblasts apoptosis, resulting in inhibition of apoptosis upon small-magnitude stress and increased apoptosis upon large-magnitude stress. PMID:27806136

  8. Embryonic liver fordin is involved in glucose glycolysis of hepatic stellate cell by regulating PI3K/Akt signaling

    PubMed Central

    Tu, Wei; Ye, Jin; Wang, Zhi-Jun

    2016-01-01

    AIM To investigate the role of embryonic liver fordin (ELF) in liver fibrosis by regulating hepatic stellate cells (HSCs) glucose glycolysis. METHODS The expression of ELF and the glucose glycolysis-related proteins were evaluated in activated HSCs. siRNA was used to silence ELF expression in activated HSCs in vitro and the subsequent changes in PI3K/Akt signaling and glucose glycolysis-related proteins were observed. RESULTS The expression of ELF increased remarkably in HSCs of the fibrosis mouse model and HSCs that were cultured for 3 wk in vitro. Glucose glycolysis-related proteins showed an obvious increase in the activated HSCs, such as phosphofructokinase, platelet and glucose transporter 1. ELF-siRNA, which perfectly silenced the expression of ELF in activated HSCs, led to the induction of glucose glycolysis-related proteins and extracellular matrix (ECM) components. Moreover, pAkt, which is an important downstream factor in PI3K/Akt signaling, showed a significant change in response to the ELF silencing. The expression of glucose glycolysis-related proteins and ECM components decreased remarkably when the PI3K/Akt signaling was blocked by Ly294002 in the activated HSCs. CONCLUSION ELF is involved in HSC glucose glycolysis by regulating PI3K/Akt signaling. PMID:27784964

  9. Akt Regulates TNFα Synthesis Downstream of RIP1 Kinase Activation during Necroptosis

    PubMed Central

    McNamara, Colleen R.; Ahuja, Ruchita; Osafo-Addo, Awo D.; Barrows, Douglas; Kettenbach, Arminja; Skidan, Igor; Teng, Xin; Cuny, Gregory D.; Gerber, Scott; Degterev, Alexei

    2013-01-01

    Necroptosis is a regulated form of necrotic cell death that has been implicated in the pathogenesis of various diseases including intestinal inflammation and systemic inflammatory response syndrome (SIRS). In this work, we investigated the signaling mechanisms controlled by the necroptosis mediator receptor interacting protein-1 (RIP1) kinase. We show that Akt kinase activity is critical for necroptosis in L929 cells and plays a key role in TNFα production. During necroptosis, Akt is activated in a RIP1 dependent fashion through its phosphorylation on Thr308. In L929 cells, this activation requires independent signaling inputs from both growth factors and RIP1. Akt controls necroptosis through downstream targeting of mammalian Target of Rapamycin complex 1 (mTORC1). Akt activity, mediated in part through mTORC1, links RIP1 to JNK activation and autocrine production of TNFα. In other cell types, such as mouse lung fibroblasts and macrophages, Akt exhibited control over necroptosis-associated TNFα production without contributing to cell death. Overall, our results provide new insights into the mechanism of necroptosis and the role of Akt kinase in both cell death and inflammatory regulation. PMID:23469174

  10. Akt Regulates TNFα synthesis downstream of RIP1 kinase activation during necroptosis.

    PubMed

    McNamara, Colleen R; Ahuja, Ruchita; Osafo-Addo, Awo D; Barrows, Douglas; Kettenbach, Arminja; Skidan, Igor; Teng, Xin; Cuny, Gregory D; Gerber, Scott; Degterev, Alexei

    2013-01-01

    Necroptosis is a regulated form of necrotic cell death that has been implicated in the pathogenesis of various diseases including intestinal inflammation and systemic inflammatory response syndrome (SIRS). In this work, we investigated the signaling mechanisms controlled by the necroptosis mediator receptor interacting protein-1 (RIP1) kinase. We show that Akt kinase activity is critical for necroptosis in L929 cells and plays a key role in TNFα production. During necroptosis, Akt is activated in a RIP1 dependent fashion through its phosphorylation on Thr308. In L929 cells, this activation requires independent signaling inputs from both growth factors and RIP1. Akt controls necroptosis through downstream targeting of mammalian Target of Rapamycin complex 1 (mTORC1). Akt activity, mediated in part through mTORC1, links RIP1 to JNK activation and autocrine production of TNFα. In other cell types, such as mouse lung fibroblasts and macrophages, Akt exhibited control over necroptosis-associated TNFα production without contributing to cell death. Overall, our results provide new insights into the mechanism of necroptosis and the role of Akt kinase in both cell death and inflammatory regulation.

  11. Loss of Akt activity increases circulating soluble endoglin release in preeclampsia: identification of inter-dependency between Akt-1 and heme oxygenase-1

    PubMed Central

    Cudmore, Melissa J.; Ahmad, Shakil; Sissaoui, Samir; Ramma, Wenda; Ma, Bin; Fujisawa, Takeshi; Al-Ani, Bahjat; Wang, Keqing; Cai, Meng; Crispi, Fatima; Hewett, Peter W.; Gratacós, Eduard; Egginton, Stuart; Ahmed, Asif

    2012-01-01

    Aims Endothelial dysfunction is a hallmark of preeclampsia. Desensitization of the phosphoinositide 3-kinase (PI3K)/Akt pathway underlies endothelial dysfunction and haeme oxygenase-1 (HO-1) is decreased in preeclampsia. To identify therapeutic targets, we sought to assess whether these two regulators act to suppress soluble endoglin (sEng), an antagonist of transforming growth factor-β (TGF-β) signalling, which is known to be elevated in preeclampsia. Methods and results Vascular endothelial growth factor-A (VEGF-A), fibroblast growth factor (FGF-2), angiopoietin-1 (Ang-1), and insulin, which all activate the PI3K/Akt pathway, inhibited the release of sEng from endothelial cells. Inhibition of the PI3K/Akt pathway, by overexpression of phosphatase and tensin homolog (PTEN) or a dominant-negative isoform of Akt (Aktdn) induced sEng release from endothelial cells and prevented the inhibitory effect of VEGF-A. Conversely, overexpression of a constitutively active Akt (Aktmyr) inhibited PTEN and cytokine-induced sEng release. Systemic delivery of Aktmyr to mice significantly reduced circulating sEng, whereas Aktdn promoted sEng release. Phosphorylation of Akt was reduced in preeclamptic placenta and this correlated with the elevated level of circulating sEng. Knock-down of Akt using siRNA prevented HO-1-mediated inhibition of sEng release and reduced HO-1 expression. Furthermore, HO-1 null mice have reduced phosphorylated Akt in their organs and overexpression of Aktmyr failed to suppress the elevated levels of sEng detected in HO-1 null mice, indicating that HO-1 is required for the Akt-mediated inhibition of sEng. Conclusion The loss of PI3K/Akt and/or HO-1 activity promotes sEng release and positive manipulation of these pathways offers a strategy to circumvent endothelial dysfunction. PMID:21411816

  12. Protective Role of PI3-kinase/Akt/eNOS Signaling in Mechanical Stress Through Inhibition of p38 Mitogen-Activated Protein Kinase in Mouse Lung

    DTIC Science & Technology

    2010-01-01

    81. 6 Iwakiri Y. Tsai MH. McCabe TJ. Gratton JP. Fulton D. Groszmann RJ. et al. Phosphorylation of eNOS initiates excessive NO production in early...Crit care Med 2008; 177(Abstracts issue): A760. 21 Gratton JP. Morales-Ruiz M. Kureishi Y. Fulton D. Walsh K. Sessa WC. Akt down-regulation of p38

  13. Photoreceptor Neuroprotection: Regulation of Akt Activation Through Serine/Threonine Phosphatases, PHLPP and PHLPPL.

    PubMed

    Rajala, Raju V S; Kanan, Yogita; Anderson, Robert E

    2016-01-01

    Serine/threonine kinase Akt is a downstream effector of insulin receptor/PI3K pathway that is involved in many processes, including providing neuroprotection to stressed rod photoreceptor cells. Akt signaling is known to be regulated by the serine/threonine phosphatases, PHLPP (PH domain and leucine rich repeat protein phosphatase) and PHLPPL (PH domain and leucine rich repeat protein phosphatase-like). We previously reported that both phosphatases are expressed in the retina, as well as in photoreceptor cells. In this study, we examined the PHLPP and PHLPPL phosphatase activities towards non-physiological and physiological substrates. Our results suggest that PHLPP was more active than PHLPPL towards non-physiological substrates, whereas both PHLPP and PHLPP dephosphorylated the physiological substrates of Akt1 and Akt3 with similar efficiencies. Our results also suggest that knockdown of PHLPPL alone does not increase Akt phosphorylation, due to a compensatory increase of PHLPP, which results in the dephosphorylation of Akt. Therefore, PHLPP and PHLPPL regulate Akt activation together when both phosphatases are expressed.

  14. Snail promotes cell migration through PI3K/AKT-dependent Rac1 activation as well as PI3K/AKT-independent pathways during prostate cancer progression

    PubMed Central

    Henderson, Veronica; Smith, Basil; Burton, Liza J; Randle, Diandra; Morris, Marisha; Odero-Marah, Valerie A

    2015-01-01

    Snail, a zinc-finger transcription factor, induces epithelial-mesenchymal transition (EMT), which is associated with increased cell migration and metastasis in cancer cells. Rac1 is a small G-protein which upon activation results in formation of lamellipodia, the first protrusions formed by migrating cells. We have previously shown that Snail promotes cell migration through down-regulation of maspin tumor suppressor. We hypothesized that Snail's regulation of cell migration may also involve Rac1 signaling regulated by PI3K/AKT and/or MAPK pathways. We found that Snail overexpression in LNCaP and 22Rv1 prostate cancer cells increased Rac1 activity associated with increased cell migration, and the Rac1 inhibitor, NSC23766, could inhibit Snail-mediated cell migration. Conversely, Snail downregulation using shRNA in the aggressive C4–2 prostate cancer cells decreased Rac1 activity and cell migration. Moreover, Snail overexpression increased ERK and PI3K/AKT activity in 22Rv1 prostate cancer cells. Treatment of Snail-overexpressing 22Rv1 cells with LY294002, PI3K/AKT inhibitor or U0126, MEK inhibitor, decreased cell migration significantly, but only LY294002 significantly reduced Rac1 activity, suggesting that Snail promotes Rac1 activation via the PI3K/AKT pathway. Furthermore, 22Rv1 cells overexpressing Snail displayed decreased maspin levels, while inhibition of maspin expression in 22Rv1 cells with siRNA, led to increased PI3K/AKT, Rac1 activity and cell migration, without affecting ERK activity, suggesting that maspin is upstream of PI3K/AKT. Overall, we have dissected signaling pathways by which Snail may promote cell migration through MAPK signaling or alternatively through PI3K/AKT-Rac1 signaling that involves Snail inhibition of maspin tumor suppressor. This may contribute to prostate cancer progression. PMID:26207671

  15. Dioscin inhibits osteoclast differentiation and bone resorption though down-regulating the Akt signaling cascades

    SciTech Connect

    Qu, Xinhua; Zhai, Zanjing; Liu, Xuqiang; Li, Haowei; Ouyang, Zhengxiao; Wu, Chuanlong; Liu, Guangwang; Fan, Qiming; Tang, Tingting; Qin, An; Dai, Kerong

    2014-01-10

    Highlights: •A natural-derived compound, dioscin, suppresses osteoclast formation and bone resorption. •Dioscin inhibits osteolytic bone loss in vivo. •Dioscin impairs the Akt signaling cascades pathways during osteoclastogenesis. •Dioscin have therapeutic value in treating osteoclast-related diseases. -- Abstract: Bone resorption is the unique function of osteoclasts (OCs) and is critical for both bone homeostasis and pathologic bone diseases including osteoporosis, rheumatoid arthritis and tumor bone metastasis. Thus, searching for natural compounds that may suppress osteoclast formation and/or function is promising for the treatment of osteoclast-related diseases. In this study, we for the first time demonstrated that dioscin suppressed RANKL-mediated osteoclast differentiation and bone resorption in vitro in a dose-dependent manner. The suppressive effect of dioscin is supported by the reduced expression of osteoclast-specific markers. Further molecular analysis revealed that dioscin abrogated AKT phosphorylation, which subsequently impaired RANKL-induced nuclear factor-kappaB (NF-κB) signaling pathway and inhibited NFATc1 transcriptional activity. Moreover, in vivo studies further verified the bone protection activity of dioscin in osteolytic animal model. Together our data demonstrate that dioscin suppressed RANKL-induced osteoclast formation and function through Akt signaling cascades. Therefore, dioscin is a potential natural agent for the treatment of osteoclast-related diseases.

  16. Differential regulation of mTOR signaling determines sensitivity to AKT inhibition in diffuse large B cell lymphoma

    PubMed Central

    Ezell, Scott A.; Wang, Suping; Bihani, Teeru; Lai, Zhongwu; Grosskurth, Shaun E.; Tepsuporn, Suprawee; Davies, Barry R.; Huszar, Dennis; Byth, Kate F.

    2016-01-01

    Agents that target components of the PI3K/AKT/mTOR pathway are under investigation for the treatment of diffuse large B cell lymphoma (DLBCL). Given the highly heterogeneous nature of DLBCL, it is not clear whether all subtypes of DLBCL will be susceptible to PI3K pathway inhibition, or which kinase within this pathway is the most favorable target. Pharmacological profiling of a panel of DLBCL cell lines revealed a subset of DLBCL that was resistant to AKT inhibition. Strikingly, sensitivity to AKT inhibitors correlated with the ability of these inhibitors to block phosphorylation of S6K1 and ribosomal protein S6. Cell lines resistant to AKT inhibition activated S6K1 independent of AKT either through upregulation of PIM2 or through activation by B cell receptor (BCR) signaling components. Finally, combined inhibition of AKT and BTK, PIM2, or S6K1 proved to be an effective strategy to overcome resistance to AKT inhibition in DLBCL. PMID:26824321

  17. Effects of RAF inhibitors on PI3K/AKT signalling depend on mutational status of the RAS/RAF signalling axis

    PubMed Central

    Fritsche-Guenther, Raphaela; Witzel, Franziska; Kempa, Stefan; Brummer, Tilman; Sers, Christine; Blüthgen, Nils

    2016-01-01

    Targeted therapies within the RAS/RAF/MEK/ERK signalling axis become increasingly popular, yet cross-talk and feedbacks in the signalling network lead to unexpected effects. Here we look systematically into how inhibiting RAF and MEK with clinically relevant inhibitors result in changes in PI3K/AKT activation. We measure the signalling response using a bead-based ELISA, and use a panel of three cell lines, and isogenic cell lines that express mutant forms of the oncogenes KRAS and BRAF to interrogate the effects of the MEK and RAF inhibitors on signalling. We find that treatment with the RAF inhibitors have opposing effects on AKT phosphorylation depending on the mutational status of two important oncogenes, KRAS and BRAF. If these two genes are in wildtype configuration, RAF inhibitors reduce AKT phosphorylation. In contrast, if BRAF or KRAS are mutant, RAF inhibitors will leave AKT phosphorylation unaffected or lead to an increase of AKT phosphorylation. Down-regulation of phospho-AKT by RAF inhibitors also extends to downstream transcription factors, and correlates with apoptosis induction. Our results show that oncogenes rewire signalling such that targeted therapies can have opposing effects on parallel pathways, which depend on the mutational status of the cell. PMID:26799289

  18. IL-29 and IFN-α regulate the expression of MxA, 2',5'-OAS and PKR genes in association with the activation of Raf-MEK-ERK and PI3K-AKT signal pathways in HepG2.2.15 cells.

    PubMed

    Chai, Yu; Huang, Hai-Liang; Hu, Dao-Jun; Luo, Xin; Tao, Qian-Shan; Zhang, Xiao-Ling; Zhang, Sheng-Quan

    2011-01-01

    Interferons (IFNs) can activate the PI3K-AKT and Raf-MEK-ERK signal pathways and induce antiviral proteins (MxA, 2',5'-OAS and PKR) expression in specific cell lines. However, the relationship between those antiviral proteins expression and signal pathways remains unknown at present. Thus our experiments were designed to determine the exact relationship in HepG2.2.15 cell line. The results demonstrated that IFN-α and IL-29 were both able to activate PI3K-AKT and Raf-MEK-ERK signal pathways, and IFN-α up-regulated the expression of MxA, 2',5'-OAS and PKR whereas IL-29 increased mRNA expression of MxA and 2',5'-OAS and had no influence on PKR. Furthermore, MxA, 2',5'-OAS and PKR expression were down-regulated while PI3K-AKT signal pathway was blocked by LY294002. And MxA was up-regulated after Raf-MEK-ERK signal pathway being blocked by PD98059. These findings indicate that the expression of MxA, 2',5'-OAS and PKR are up-regulate by PI3K-AKT signal pathway, and Raf-MEK-ERK signal pathway has a negative regulatory effect on the expression of MxA and no significant effect on 2',5'-OAS and PKR.

  19. 14-3-3ζ up-regulates hypoxia-inducible factor-1α in hepatocellular carcinoma via activation of PI3K/Akt/NF-кB signal transduction pathway

    PubMed Central

    Tang, Yufu; Lv, Pengfei; Sun, Zhongyi; Han, Lei; Luo, Bichao; Zhou, Wenping

    2015-01-01

    14-3-3ζ protein, a member of 14-3-3 family, plays important roles in multiple cellular processes. Our previous study showed that 14-3-3ζ could bind to regulate the expression of hypoxia-inducible factor-1α (HIF-1α), which is induced by hypoxia and a crucial factor for induction of tumor metastasis. Moreover, we also have confirmed the response of 14-3-3ζ to hypoxia in our unpublished data as well. Thus, in the present study, we attempted to reveal that whether the regulation effect of 14-3-3ζ on HIF-1α functioned in a similar pattern as hypoxia. Stable regulation of 14-3-3ζ in human HCC cell line SMMC-772 and HCC-LM3 was achieved. The regulation of 14-3-3ζ on HIF-1α mRNA transcription was evaluated by luciferase activity assay and quantitative real-time PCR (qPCR). The effect of 14-3-3ζ on the production of HIF-1α and pathways determining HIF-1α’s response to hypoxia was assessed using western blotting assay. Our results showed that regulation of 14-3-3ζ expression influenced the activity of HIF-1α, phosphatidyl inositol 3-kinase (PI3K), Akt, extracellular signal-regulated kinase 1/2 (ERK1/2), and nuclear factor kappa B (NF-кB). Blocking of these pathways using indicated inhibitors revealed that 14-3-3ζ enhanced the production of HIF-1α via the activation of PI3K/Akt/NF-кB pathway, which was identical to hypoxia induced HIF-1α expression. For the first time, our study described the key role of 14-3-3ζ in the HIF-1α production in HCC cells. And the molecule exerted its function on HIF-1α both by directly binding to it and via PI3K/Akt/NF-кB signal transduction pathway. PMID:26884855

  20. Anti-proliferative effect of RCE-4 from Reineckia carnea on human cervical cancer HeLa cells by inhibiting the PI3K/Akt/mTOR signaling pathway and NF-κB activation.

    PubMed

    Bai, Caihong; Yang, Xiaojiao; Zou, Kun; He, Haibo; Wang, Junzhi; Qin, Huilin; Yu, Xiaoqin; Liu, Chengxiong; Zheng, Juyan; Cheng, Fan; Chen, Jianfeng

    2016-06-01

    Cervical cancer is the second leading cause of cancer deaths in women worldwide. In recent years, the studies find that inflammation is a critical component of tumor progression, and the ideal therapeutic methods should be aimed at the inflammation reaction triggers. (1β,3β,5β,25S)-spirostan-1,3-diol1-[α-L-rhamnopyranosyl-(1 → 2)-β-D-xylopyranoside] (RCE-4) was the main active composition of Reineckia carnea (Andr.) Kunth. It significantly induced apoptosis in cervical cancer Caski cells through the mitochondrial pathway in our previous studies; however, its underlying mechanism remains poorly understood. This study aimed to further evaluate the effect of RCE-4 on human cervical cancer HeLa cells. Based on this observation, we investigated the anti-cervical cancer effect of RCE-4 by modulating phosphatidylinositol 3-kinase/protein kinase-B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway, nuclear factor-kappa B (NF-κB) activation, and inflammation-related key factors in HeLa cells. The results indicated that the HeLa cell was the most sensitive with an IC50 of 7.01 μM; RCE-4 significantly promoted the release of cellular lactate dehydrogenase (LDH); increased DNA fragmentation and apoptosis; reduced PI3K, Akt, mTOR, and NF-κBp65 phosphorylation levels; increased the Bax and cleaved poly (ADP-ribose) polymerase (PARP) protein levels; suppressed Bcl-2 protein expression; elevated the Bax/Bcl-2 expression ratio; and decreased the interleukin-1 beta (IL-1β) and interleukin-6 (IL-6) mRNA expressions in HeLa cells in a concentration-dependent manner. These findings suggest that RCE-4 exerted beneficially anti-cervical cancer effect on HeLa cells, mainly inhibiting PI3K/Akt/mTOR signaling pathway phosphorylation and NF-κB activation, promoting HeLa cell apoptosis. Graphical abstract Anti-tumor effect of RCE-4 on HeLa cells.

  1. Hydrogen peroxide inhibits transforming growth factor-β1-induced cell cycle arrest by promoting Smad3 linker phosphorylation through activation of Akt-ERK1/2-linked signaling pathway

    SciTech Connect

    Choi, Jiyeon; Park, Seong Ji; Jo, Eun Ji; Lee, Hui-Young; Hong, Suntaek; Kim, Seong-Jin; Kim, Byung-Chul

    2013-06-14

    Highlights: •H{sub 2}O{sub 2} inhibits TGF-β1-induced cell cycle arrest. •H{sub 2}O{sub 2} induces Smad3 linker phosphorylation through Akt-ERK1/2 pathway. •H{sub 2}O{sub 2}-mediated suppression of TGF-β signal requires Smad3 linker phosphorylation. •This is a first report about interplay between H{sub 2}O{sub 2} and growth inhibition pathway. -- Abstract: Hydrogen peroxide (H{sub 2}O{sub 2}) functions as a second messenger in growth factor receptor-mediated intracellular signaling cascade and is tumorigenic by virtue of its ability to promote cell proliferation; however, the mechanisms underlying the growth stimulatory action of H{sub 2}O{sub 2} are less understood. Here we report an important mechanism for antagonistic effects of H{sub 2}O{sub 2} on growth inhibitory response to transforming growth factor-β1 (TGF-β1). In Mv1Lu and HepG2 cells, pretreatment of H{sub 2}O{sub 2} (0.05–0.2 mM) completely blocked TGF-β1-mediated induction of p15{sup INK4B} expression and increase of its promoter activity. Interestingly, H{sub 2}O{sub 2} selectively suppressed the transcriptional activation potential of Smad3, not Smad2, in the absence of effects on TGF-β1-induced phosphorylation of the COOH-tail SSXS motif of Smad3 and its nuclear translocation. Mechanism studies showed that H{sub 2}O{sub 2} increases the phosphorylation of Smad3 at the middle linker region in a concentration- and time-dependent manner and this effect is mediated by activation of extracellular signal-activated kinase 1/2 through Akt. Furthermore, expression of a mutant Smad3 in which linker phosphorylation sites were ablated significantly abrogated the inhibitory effects of H{sub 2}O{sub 2} on TGF-β1-induced increase of p15{sup INK4B}-Luc reporter activity and blockade of cell cycle progression from G1 to S phase. These findings for the first time define H{sub 2}O{sub 2} as a signaling molecule that modulate Smad3 linker phosphorylation and its transcriptional activity, thus providing

  2. Psoralidin, An Herbal Molecule Inhibits PI3K Mediated Akt Signaling In Androgen Independent Prostate Cancer (AIPC) Cells

    PubMed Central

    Kumar, Raj; Srinivasan, Sowmyalakshmi; Koduru, Srinivas; Pahari, Pallab; Rohr, Jürgen; Kyprianou, Natasha; Damodaran, Chendil

    2008-01-01

    The protein kinase Akt plays an important role in cell proliferation and survival in many cancers, including prostate cancer. Due to its kinase activity, it serves as a molecular conduit for inhibiting apoptosis and promoting angiogenesis in most cell types. In most of the prostate tumors, Akt signaling is constitutively activated due to the deletion or mutation of the tumor suppressor PTEN, which negatively regulates PI3K through lipid phosphatase activity. Recently, we identified a natural compound, psoralidin, which inhibits Akt phosphorylation and its consequent activation in androgen independent prostate cancer cells (AIPC). Furthermore, ectopic expression of Akt renders AIPC cells resistant chemotherapy; however, psoralidin overcomes Akt-mediated resistance and induces apoptosis in AIPC cells. While dissecting the molecular events, both upstream and downstream of Akt, we found that psoralidin inhibits PI3 kinase activation and transcriptionally represses the activation of NF-κB and its target genes (Bcl-2, Survivin, and Bcl-xL, etc.), which results in the inhibition of cell viability and induction of apoptosis in PC-3 and DU-145 cells. Interestingly, psoralidin selectively targets cancer cells, without causing any toxicity to normal prostate epithelial cells. In vivo xenograft assays substantiate these in vitro findings, and show psoralidin inhibits prostate tumor growth in nude mice. Our findings are of therapeutic significance in the management of prostate cancer patients with advanced or metastatic disease, as they provide new directions for the development of a phyotochemical-based platform for prevention and treatment strategies for AIPC. PMID:19223576

  3. δ-Tocopherol inhibits receptor tyrosine kinase-induced AKT activation in prostate cancer cells.

    PubMed

    Wang, Hong; Hong, Jungil; Yang, Chung S

    2016-11-01

    The cancer preventive activity of vitamin E is suggested by epidemiological studies and supported by animal studies with vitamin E forms, γ-tocopherol and δ-tocopherol (δ-T). Several recent large-scale cancer prevention trials with high dose of α-tocopherol, however, yielded disappointing results. Whether vitamin E prevents or promotes cancer is a serious concern. A better understanding of the molecular mechanisms of action of the different forms of tocopherols would enhance our understanding of this topic. In this study, we demonstrated that δ-T was the most effective tocopherol form in inhibiting prostate cancer cell growth, by inducing cell cycle arrest and apoptosis. By profiling the effects of δ-T on the cell signaling using the phospho-kinase array, we found that the most inhibited target was the phosphorylation of AKT on T308. Further study on the activation of AKT by EGFR and IGFR revealed that δ-T attenuated the EGF/IGF-induced activation of AKT (via the phosphorylation of AKT on T308 induced by the activation of PIK3). Expression of dominant active PIK3 and AKT in prostate cancer cell line DU145 in which PIK3, AKT, and PTEN are wild type caused the cells to be reflectory to the inhibition of δ-T, supporting that δ-T inhibits the PIK3-mediated activation of AKT. Our data also suggest that δ-T interferes with the EGF-induced EGFR internalization, which leads to the inhibition of the receptor tyrosine kinase-dependent activation of AKT. In summary, our results revealed a novel mechanism of δ-T in inhibiting prostate cancer cell growth, supporting the cancer preventive activity δ-T. © 2015 Wiley Periodicals, Inc.

  4. Myostatin regulates cardiomyocyte growth through modulation of Akt signaling.

    PubMed

    Morissette, Michael R; Cook, Stuart A; Foo, ShiYin; McKoy, Godfrina; Ashida, Noboru; Novikov, Mikhail; Scherrer-Crosbie, Marielle; Li, Ling; Matsui, Takashi; Brooks, Gavin; Rosenzweig, Anthony

    2006-07-07

    Myostatin is a highly conserved, potent negative regulator of skeletal muscle hypertrophy in many species, from rodents to humans, although its mechanisms of action are incompletely understood. Transcript profiling of hearts from a genetic model of cardiac hypertrophy revealed dramatic upregulation of myostatin, not previously recognized to play a role in the heart. Here we show that myostatin abrogates the cardiomyocyte growth response to phenylephrine in vitro through inhibition of p38 and the serine-threonine kinase Akt, a critical determinant of cell size in many species from drosophila to mammals. Evaluation of male myostatin-null mice revealed that their cardiomyocytes and hearts overall were slightly smaller at baseline than littermate controls but exhibited more exuberant growth in response to chronic phenylephrine infusion. The increased cardiac growth in myostatin-null mice corresponded with increased p38 phosphorylation and Akt activation in vivo after phenylephrine treatment. Together, these data demonstrate that myostatin is dynamically regulated in the heart and acts more broadly than previously appreciated to regulate growth of multiple types of striated muscle.

  5. Targeting the PI3K/AKT/mTOR Signaling Axis in Children with Hematologic Malignancies

    PubMed Central

    Barrett, David; Brown, Valerie I.; Grupp, Stephan A.; Teachey, David T.

    2014-01-01

    The phosphatidylinositiol 3-kinase (PI3K), AKT, mammalian target of rapamycin (mTOR) signaling pathway (PI3K/AKT/mTOR) is frequently dysregulated in disorders of cell growth and survival, including a number of pediatric hematologic malignancies. The pathway can be abnormally activated in childhood acute lymphoblastic leukemia (ALL), acute myelogenous leukemia (AML), and chronic myelogenous leukemia (CML), as well as in some pediatric lymphomas and lymphoproliferative disorders. Most commonly, this abnormal activation occurs as a consequence of constitutive activation of AKT, providing a compelling rationale to target this pathway in many of these conditions. A variety of agents, beginning with the rapamycin analogue (rapalog) sirolimus, have been used successfully to target this pathway in a number of pediatric hematologic malignancies. Rapalogs demonstrate significant preclinical activity against ALL, which has led to a number of clinical trials. Moreover, rapalogs can synergize with a number of conventional cytotoxic agents and overcome pathways of chemotherapeutic resistance for drugs commonly used in ALL treatment, including methotrexate and corticosteroids. Based on preclinical data, rapalogs are also being studied in AML, CML, and non-Hodgkin’s lymphoma. Recently, significant progress has been made using rapalogs to treat pre-malignant lymphoproliferative disorders, including the autoimmune lymphoproliferative syndrome (ALPS); complete remissions in children with otherwise therapy-resistant disease have been seen. Rapalogs only block one component of the pathway (mTORC1), and newer agents are under preclinical and clinical development that can target different and often multiple protein kinases in the PI3K/AKT/mTOR pathway. Most of these agents have been tolerated in early-phase clinical trials. A number of PI3K inhibitors are under investigation. Of note, most of these also target other protein kinases. Newer agents are under development that target both m

  6. Reduced signaling of PI3K-Akt and RAS-MAPK pathways is the key target for weight-loss-induced cancer prevention by dietary calorie restriction and/or physical activity.

    PubMed

    Standard, Joseph; Jiang, Yu; Yu, Miao; Su, Xiaoyu; Zhao, Zhihui; Xu, Jianteng; Chen, Jie; King, Brenee; Lu, Lizhi; Tomich, John; Baybutt, Richard; Wang, Weiqun

    2014-12-01

    Weight control through either dietary calorie restriction (DCR) or exercise has been associated with cancer prevention in animal models. However, the underlying mechanisms are not fully defined. Bioinformatics using genomics, proteomics and lipidomics was employed to elucidate the molecular targets of weight control in a mouse skin cancer model. SENCAR mice were randomly assigned into four groups for 10 weeks: ad-libitum-fed sedentary control, ad-libitum-fed exercise (AE), exercise but pair-fed isocaloric amount of control (PE) and 20% DCR. Two hours after topical TPA treatment, skin epidermis was analyzed by Affymetrix for gene expression, DIGE for proteomics and lipidomics for phospholipids. Body weights were significantly reduced in both DCR and PE but not AE mice versus the control. Among 39,000 transcripts, 411, 67 and 110 genes were significantly changed in DCR, PE and AE, respectively. The expression of genes relevant to PI3K-Akt and Ras-MAPK signaling was effectively reduced by DCR and PE but not AE as measured through GenMAPP software. Proteomics analysis identified ~120 proteins, with 27 proteins significantly changed by DCR, including up-regulated apolipoprotein A-1, a key antioxidant protein that decreases Ras-MAPK activity. Of the total 338 phospholipids analyzed by lipidomics, 57 decreased by PE including 5 phophatidylinositol species that serve as PI3K substrates. Although a full impact has not been determined yet, it appears that the reduction of both Ras-MAPK and PI3K-Akt signaling pathways is a cancer preventive target that has been consistently demonstrated by three bioinformatics approaches.

  7. Oxidized ATM promotes abnormal proliferation of breast CAFs through maintaining intracellular redox homeostasis and activating the PI3K-AKT, MEK-ERK, and Wnt-β-catenin signaling pathways.

    PubMed

    Tang, Shifu; Hou, Yixuan; Zhang, Hailong; Tu, Gang; Yang, Li; Sun, Yifan; Lang, Lei; Tang, Xi; Du, Yan-E; Zhou, Mingli; Yu, Tenghua; Xu, Liyun; Wen, Siyang; Liu, Chunming; Liu, Manran

    2015-01-01

    Abnormal proliferation is one characteristic of cancer-associated fibroblasts (CAFs), which play a key role in tumorigenesis and tumor progression. Oxidative stress (OS) is the root cause of CAFs abnormal proliferation. ATM (ataxia-telangiectasia mutated protein kinase), an important redox sensor, is involved in DNA damage response and cellular homeostasis. Whether and how oxidized ATM regulating CAFs proliferation remains unclear. In this study, we show that there is a high level of oxidized ATM in breast CAFs in the absence of double-strand breaks (DSBs) and that oxidized ATM plays a critical role in CAFs proliferation. The effect of oxidized ATM on CAFs proliferation is mediated by its regulation of cellular redox balance and the activity of the ERK, PI3K-AKT, and Wnt signaling pathways. Treating cells with antioxidant N-acetyl-cysteine (NAC) partially rescues the proliferation defect of the breast CAFs caused by ATM deficiency. Administrating cells with individual or a combination of specific inhibitors of the ERK, PI3K-AKT, and Wnt signaling pathways mimics the effect of ATM deficiency on breast CAF proliferation. This is mainly ascribed to the β-catenin suppression and down-regulation of c-Myc, thus further leading to the decreased cyclinD1, cyclinE, and E2F1 expression and the enhanced p21(Cip1) level. Our results reveal an important role of oxidized ATM in the regulation of the abnormal proliferation of breast CAFs. Oxidized ATM could serve as a potential target for treating breast cancer.

  8. Reduced signaling of PI3K-Akt and RAS-MAPK pathways are the key targets for weight loss-induced cancer prevention by dietary calorie restriction and/or physical activity

    PubMed Central

    Standard, Joseph; Jiang, Yu; Yu, Miao; Su, Xiaoyu; Zhao, Zhihui; Xu, Jianteng; Chen, Jie; King, Brenee; Lu, Lizhi; Tomich, John; Baybutt, Richard; Wang, Weiqun

    2014-01-01

    Weight control through either dietary calorie restriction (DCR) or exercise has been associated with cancer prevention in animal models. However, the underlying mechanisms are not fully defined. Bioinformatics using genomics, proteomics, and lipidomics were employed to elucidate the molecular targets of weight control in a mouse skin cancer model. SENCAR mice were randomly assigned into 4 groups for 10 weeks: ad lib-fed sedentary control, ad lib-fed exercise (AE), exercise but pair-fed isocaloric amount of control (PE), and 20% DCR. Two hours after topical TPA treatment, skin epidermis was analyzed by Affymetrix for gene expression, DIGE for proteomics, and lipidomics for phospholipids. Body weights were significantly reduced in both DCR and PE but not AE mice versus the control. Among 39,000 transcripts, 411, 67, and 110 genes were significantly changed in DCR, PE, and AE, respectively. The expression of genes relevant to PI3K-Akt and Ras-MAPK signaling was effectively reduced by DCR and PE but not AE as measured through GenMAPP software. Proteomics analysis identified ~120 proteins, with 27 proteins significantly changed by DCR, including upregulated apolipoprotein A-1, a key antioxidant protein that decreases Ras-MAPK activity. Of the total 338 phospholipids analyzed by lipidomics, 57 decreased by PE including 5 phophatidylinositol species that serve as PI3K substrates. Although a full impact has not been determined yet, it appears the reduction of both Ras-MAPK and PI3K-Akt signaling pathways are cancer preventive targets that have been consistently demonstrated by three bioinformatics approaches. PMID:25283328

  9. PI3K/Akt/mTOR signaling & its regulator tumour suppressor genes PTEN & LKB1 in human uterine leiomyomas

    PubMed Central

    Makker, Annu; Goel, Madhu Mati; Mahdi, Abbas Ali; Bhatia, Vikram; Das, Vinita; Agarwal, Anjoo; Pandey, Amita

    2016-01-01

    Background & objectives: Despite their high occurrence and associated significant level of morbidity manifesting as spectrum of clinical symptoms, the pathogenesis of uterine leiomyomas (ULs) remains unclear. We investigated expression profile of tumour suppressor genes PTEN (phosphatase and tensin homolog deleted on chromosome ten) and LKB1 (liver kinase B1), and key signaling components of P13K (phosphatidylinositol 3-kinase)/Akt (protein kinase B)/mTOR (mammalian target of rapamycin) pathway in leiomyomas and adjacent normal myometrium in women of reproductive age, to explore the possibility of targeting this pathway for future therapeutic implications. Methods: Real time PCR (qPCR) was used to quantify relative gene expression levels of PTEN, Akt1, Akt2, mTOR, LKB1 and VEGFA (vascular endothelial growth factor A) in leiomyoma as compared to adjacent normal myometrium. Immunohistochemistry was subsequently performed to analyze expression of PTEN, phospho-Akt, phospho-mTOR, phospho-S6, LKB1 and VEGFA in leiomyoma and adjacent normal myometrium. Results: Significant upregulation of PTEN (2.52 fold; P=0.03) and LKB1 (3.93 fold; P=0.01), and downregulation of VEGFA (2.95 fold; P=0.01) genes were observed in leiomyoma as compared to normal myometrium. Transcript levels of Akt1, Akt2 and mTOR did not vary significantly between leiomyoma and myometrium. An increased immunoexpression of PTEN (P=0.015) and LKB1 (P<0.001) and decreased expression of VEGFA (P=0.01) was observed in leiomyoma as compared to myometrium. Immunostaining for activated (phosphorylated) Akt, mTOR and S6 was absent or low in majority of leiomyoma and myometrium. Interpretation & conclusions: Upregulation of PTEN and LKB1 in concert with negative or low levels of activated Akt, mTOR and S6 indicates that PI3K/Akt/mTOR pathway may not play a significant role in pathogenesis of leiomyoma. PMID:27748285

  10. Highly Active Microbial Phosphoantigen Induces Rapid yet Sustained MEK/Erk- and PI-3K/Akt-Mediated Signal Transduction in Anti-Tumor Human γδ T-Cells

    PubMed Central

    Cardoso, Bruno A.; Lança, Telma; Grosso, Ana R.; deBarros, Ana; Martins, Leila R.; Barata, João T.; Silva-Santos, Bruno

    2009-01-01

    Background The unique responsiveness of Vγ9Vδ2 T-cells, the major γδ subset of human peripheral blood, to non-peptidic prenyl pyrophosphate antigens constitutes the basis of current γδ T-cell-based cancer immunotherapy strategies. However, the molecular mechanisms responsible for phosphoantigen-mediated activation of human γδ T-cells remain unclear. In particular, previous reports have described a very slow kinetics of activation of T-cell receptor (TCR)-associated signal transduction pathways by isopentenyl pyrophosphate and bromohydrin pyrophosphate, seemingly incompatible with direct binding of these antigens to the Vγ9Vδ2 TCR. Here we have studied the most potent natural phosphoantigen yet identified, (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP), produced by Eubacteria and Protozoa, and examined its γδ T-cell activation and anti-tumor properties. Methodology/Principal Findings We have performed a comparative study between HMB-PP and the anti-CD3ε monoclonal antibody OKT3, used as a reference inducer of bona fide TCR signaling, and followed multiple cellular and molecular γδ T-cell activation events. We show that HMB-PP activates MEK/Erk and PI-3K/Akt pathways as rapidly as OKT3, and induces an almost identical transcriptional profile in Vγ9+ T-cells. Moreover, MEK/Erk and PI-3K/Akt activities are indispensable for the cellular effects of HMB-PP, including γδ T-cell activation, proliferation and anti-tumor cytotoxicity, which are also abolished upon antibody blockade of the Vγ9+ TCR Surprisingly, HMB-PP treatment does not induce down-modulation of surface TCR levels, and thereby sustains γδ T-cell activation upon re-stimulation. This ultimately translates in potent human γδ T-cell anti-tumor function both in vitro and in vivo upon transplantation of human leukemia cells into lymphopenic mice, Conclusions/Significance The development of efficient cancer immunotherapy strategies critically depends on our capacity to maximize anti

  11. Multiple host kinases contribute to Akt activation during Salmonella infection.

    PubMed

    Roppenser, Bernhard; Kwon, Hyunwoo; Canadien, Veronica; Xu, Risheng; Devreotes, Peter N; Grinstein, Sergio; Brumell, John H

    2013-01-01

    SopB is a type 3 secreted effector with phosphatase activity that Salmonella employs to manipulate host cellular processes, allowing the bacteria to establish their intracellular niche. One important function of SopB is activation of the pro-survival kinase Akt/protein kinase B in the infected host cell. Here, we examine the mechanism of Akt activation by SopB during Salmonella infection. We show that SopB-mediated Akt activation is only partially sensitive to PI3-kinase inhibitors LY294002 and wortmannin in HeLa cells, suggesting that Class I PI3-kinases play only a minor role in this process. However, depletion of PI(3,4) P2/PI(3-5) P3 by expression of the phosphoinositide 3-phosphatase PTEN inhibits Akt activation during Salmonella invasion. Therefore, production of PI(3,4) P2/PI(3-5) P3 appears to be a necessary event for Akt activation by SopB and suggests that non-canonical kinases mediate production of these phosphoinositides during Salmonella infection. We report that Class II PI3-kinase beta isoform, IPMK and other kinases identified from a kinase screen all contribute to Akt activation during Salmonella infection. In addition, the kinases required for SopB-mediated activation of Akt vary depending on the type of infected host cell. Together, our data suggest that Salmonella has evolved to use a single effector, SopB, to manipulate a remarkably large repertoire of host kinases to activate Akt for the purpose of optimizing bacterial replication in its host.

  12. Kinetic analysis of platelet-derived growth factor receptor/phosphoinositide 3-kinase/Akt signaling in fibroblasts.

    PubMed

    Park, Chang Shin; Schneider, Ian C; Haugh, Jason M

    2003-09-26

    Isoforms of the serine-threonine kinase Akt coordinate multiple cell survival pathways in response to stimuli such as platelet-derived growth factor (PDGF). Activation of Akt is a multistep process, which relies on the production of 3'-phosphorylated phosphoinositide (PI) lipids by PI 3-kinases. To quantitatively assess the kinetics of PDGF receptor/PI 3-kinase/Akt signaling in fibroblasts, a systematic study of this pathway was performed, and a mechanistic mathematical model that describes its operation was formulated. We find that PDGF receptor phosphorylation exhibits positive cooperativity with respect to PDGF concentration, and its kinetics are quantitatively consistent with a mechanism in which receptor dimerization is initially mediated by the association of two 1:1 PDGF/PDGF receptor complexes. Receptor phosphorylation is transient at high concentrations of PDGF, consistent with the loss of activated receptors upon endocytosis. By comparison, Akt activation responds to lower PDGF concentrations and exhibits more sustained kinetics. Further analysis and modeling suggest that the pathway is saturated at the level of PI 3-kinase activation, and that the p110alpha catalytic subunit of PI 3-kinase contributes most to PDGF-stimulated 3'-PI production. Thus, at high concentrations of PDGF the kinetics of 3'-PI production are limited by the turnover rate of these lipids, while the Akt response is additionally influenced by the rate of Akt deactivation.

  13. CARMA1 is required for Akt-mediated NF-kappaB activation in T cells.

    PubMed

    Narayan, Preeti; Holt, Brittany; Tosti, Richard; Kane, Lawrence P

    2006-03-01

    Many details of the generic pathway for induction of NF-kappaB have been delineated, but it is still not clear how multiple, diverse receptor systems are able to converge on this evolutionarily conserved family of transcription factors. Recent studies have shown that the CARMA1, Bcl10, and MALT1 proteins are critical for coupling the common elements of the NF-kappaB pathway to the T-cell receptor (TCR) and CD28. We previously demonstrated a role for the serine/threonine kinase Akt in CD28-mediated NF-kappaB induction. Using a CARMA1-deficient T-cell line, we have now found that the CARMA complex is required for induction of NF-kappaB by Akt, in cooperation with protein kinase C activation. Furthermore, using a novel selective inhibitor of Akt, we confirm that Akt plays a modulatory role in NF-kappaB induction by the TCR and CD28. Finally, we provide evidence for a physical and functional interaction between Akt and CARMA and for Akt-dependent phosphorylation of Bcl10. Therefore, in T cells, Akt impinges upon NF-kappaB signaling through at least two separate mechanisms.

  14. Akt isoforms in vascular disease

    PubMed Central

    Yu, Haixiang; Littlewood, Trevor; Bennett, Martin

    2015-01-01

    The mammalian serine/threonine Akt kinases comprise three closely related isoforms: Akt1, Akt2 and Akt3. Akt activation has been implicated in both normal and disease processes, including in development and metabolism, as well as cancer and cardiovascular disease. Although Akt signalling has been identified as a promising therapeutic target in cancer, its role in cardiovascular disease is less clear. Importantly, accumulating evidence suggests that the three Akt isoforms exhibit distinct tissue expression profiles, localise to different subcellular compartments, and have unique modes of activation. Consistent with in vitro findings, genetic studies in mice show distinct effects of individual Akt isoforms on the pathophysiology of cardiovascular disease. This review summarises recent studies of individual Akt isoforms in atherosclerosis, vascular remodelling and aneurysm formation, to provide a comprehensive overview of Akt function in vascular disease. PMID:25929188

  15. Rapid activation of ERK1/2 and AKT in human breast cancer cells by cadmium

    SciTech Connect

    Liu Zhiwei; Yu Xinyuan; Shaikh, Zahir A.

    2008-05-01

    Cadmium (Cd), an endocrine disruptor, can induce a variety of signaling events including the activation of ERK1/2 and AKT. In this study, the involvement of estrogen receptors (ER) in these events was evaluated in three human breast caner cell lines, MCF-7, MDA-MB-231, and SK-BR-3. The Cd-induced signal activation patterns in the three cell lines mimicked those exhibited in response to 17{beta}-estradiol. Specifically, treatment of MCF-7 cells, that express ER{alpha}, ER{beta} and GPR30, to 0.5-10 {mu}M Cd for only 2.5 min resulted in transient phosphorylation of ERK1/2. Cd also triggered a gradual increase and sustained activation of AKT during the 60 min treatment period. In SK-BR-3 cells, that express only GPR30, Cd also caused a transient activation of ERK1/2, but not of AKT. In contrast, in MDA-MB-231 cells, that express only ER{beta}, Cd was unable to cause rapid activation of either ERK1/2 or AKT. A transient phosphorylation of ER{alpha} was also observed within 2.5 min of Cd exposure in the MCF-7 cells. While the estrogen receptor antagonist, ICI 182,780, did not prevent the effect of Cd on these signals, specific siRNA against hER{alpha} significantly reduced Cd-induced activation of ERK1/2 and completely blocked the activation of AKT. It is concluded that Cd, like estradiol, can cause rapid activation of ERK1/2 and AKT and that these signaling events are mediated by possible interaction with membrane ER{alpha} and GPR30, but not ER{beta}.

  16. PI3K/Akt signaling mediated Hexokinase-2 expression inhibits cell apoptosis and promotes tumor growth in pediatric osteosarcoma

    SciTech Connect

    Zhuo, Baobiao; Li, Yuan; Li, Zhengwei; Qin, Haihui; Sun, Qingzeng; Zhang, Fengfei; Shen, Yang; Shi, Yingchun; Wang, Rong

    2015-08-21

    Accumulating evidence has shown that PI3K/Akt pathway is frequently hyperactivated in osteosarcoma (OS) and contributes to tumor initiation and progression. Altered phenotype of glucose metabolism is a key hallmark of cancer cells including OS. However, the relationship between PI3K/Akt pathway and glucose metabolism in OS remains largely unexplored. In this study, we showed that elevated Hexokinase-2 (HK2) expression, which catalyzes the first essential step of glucose metabolism by conversion of glucose into glucose-6-phosphate, was induced by activated PI3K/Akt signaling. Immunohistochemical analysis showed that HK2 was overexpressed in 83.3% (25/30) specimens detected and was closely correlated with Ki67, a cell proliferation index. Silencing of endogenous HK2 resulted in decreased aerobic glycolysis as demonstrated by reduced glucose consumption and lactate production. Inhibition of PI3K/Akt signaling also suppressed aerobic glycolysis and this effect can be reversed by reintroduction of HK2. Furthermore, knockdown of HK2 led to increased cell apoptosis and reduced ability of colony formation; meanwhile, these effects were blocked by 2-Deoxy-D-glucose (2-DG), a glycolysis inhibitor through its actions on hexokinase, indicating that HK2 functions in cell apoptosis and growth were mediated by altered aerobic glycolysis. Taken together, our study reveals a novel relationship between PI3K/Akt signaling and aerobic glycolysis and indicates that PI3K/Akt/HK2 might be potential therapeutic approaches for OS. - Highlights: • PI3K/Akt signaling contributes to elevated expression of HK2 in osteosarcoma. • HK2 inhibits cell apoptosis and promotes tumor growth through enhanced Warburg effect. • Inhibition of glycolysis blocks the oncogenic activity of HK2.

  17. Gα13 negatively controls osteoclastogenesis through inhibition of the Akt-GSK3β-NFATc1 signalling pathway

    PubMed Central

    Wu, Mengrui; Chen, Wei; Lu, Yun; Zhu, Guochun; Hao, Liang; Li, Yi-Ping

    2017-01-01

    Many positive signalling pathways of osteoclastogenesis have been characterized, but negative signalling pathways are less well studied. Here we show by microarray and RNAi that guanine nucleotide-binding protein subunit α13 (Gα13) is a negative regulator of osteoclastogenesis. Osteoclast-lineage-specific Gna13 conditional knockout mice have a severe osteoporosis phenotype. Gna13-deficiency triggers a drastic increase in both osteoclast number and activity (hyper-activation), mechanistically through decreased RhoA activity and enhanced Akt/GSK3β/NFATc1 signalling. Consistently, Akt inhibition or RhoA activation rescues hyper-activation of Gna13-deficient osteoclasts, and RhoA inhibition mimics the osteoclast hyperactivation resulting from Gna13-deficiency. Notably, Gα13 gain-of-function inhibits Akt activation and osteoclastogenesis, and protects mice from pathological bone loss in disease models. Collectively, we reveal that Gα13 is a master endogenous negative switch for osteoclastogenesis through regulation of the RhoA/Akt/GSK3β/NFATc1 signalling pathway, and that manipulating Gα13 activity might be a therapeutic strategy for bone diseases. PMID:28102206

  18. Grape polyphenols inhibit Akt/mammalian target of rapamycin signaling and potentiate the effects of gefitinib in breast cancer.

    PubMed

    Castillo-Pichardo, Linette; Dharmawardhane, Suranganie F

    2012-01-01

    We recently reported that a combination of dietary grape polyphenols resveratrol, quercetin, and catechin (RQC), at low concentrations, was effective at inhibiting metastatic cancer progression. Herein, we investigate the molecular mechanisms of RQC in breast cancer and explore the potential of RQC as a potentiation agent for the epidermal growth factor receptor (EGFR) therapeutic gefitinib. Our in vitro experiments showed RQC induced apoptosis in gefitinib-resistant breast cancer cells via regulation of a myriad of proapoptotic proteins. Because the Akt/mammalian target of rapamycin (mTOR) signaling pathway is often elevated during development of anti-EGFR therapy resistance, the effect of RQC on the mTOR upstream effector Akt and the negative regulator AMP kinase (AMPK) was investigated. RQC was found to reduce Akt activity, induce the activation of AMPK, and inhibit mTOR signaling in breast cancer cells. Combined RQC and gefitinib decreased gefitinib resistant breast cancer cell viability to a greater extent than RQC or gefitinib alone. Moreover, RQC inhibited Akt and mTOR and activated AMPK even in the presence of gefitinib. Our in vivo experiments showed combined RQC and gefitinib was more effective than the individual treatments at inhibiting mammary tumor growth and metastasis in nude mice. Therefore, RQC treatment inhibits breast cancer progression and may potentiate anti-EGFR therapy by inhibition of Akt/mTOR signaling.

  19. AKT signaling is involved in fucoidan-induced inhibition of growth and migration of human bladder cancer cells.

    PubMed

    Cho, Tae-Min; Kim, Wun-Jae; Moon, Sung-Kwon

    2014-02-01

    We identified a novel mechanism of AKT signaling in the fucoidan-induced proliferation and migration of human urinary 5637 cancer cells. Fucoidan treatment showed a significant growth inhibition followed by G1-phase-associated up-regulation of p21WAF1 expression and suppression of cyclins and CDK expression in 5637 cells. Also, fucoidan treatment induced the activation of AKT signaling, which was inhibited by treatment with wortmannin, a PI3K-specific inhibitor. Blockade of the AKT function reversed the fucoidan-mediated inhibition of cell proliferation, the increased G1-phase-associated p21WAF1 expression, and the reduction of cell-cycle proteins. Moreover, treatment with fucoidan blocked migration and invasion of 5637 cells. This inhibition was attributed to decreased expression of MMP-9, which was mediated by down-regulation of AP-1 and NF-κB binding activity. Furthermore, wortmannin treatment abolished the decreased cell migration and invasion and the inhibition of MMP-9 expression via the suppression of NF-κB and AP-1 in fucoidan-treated cells. Similar results were observed in another bladder cancer T-24 cells treated with fucoidan. Finally, overexpression of the AKT gene inhibited the proliferation, migration and invasion of bladder cancer cells. These data suggest that the activation of AKT signaling is involved in growth inhibition and suppression of the migration and invasion of bladder cancer cells treated with fucoidan.

  20. CKIP-1 regulates macrophage proliferation by inhibiting TRAF6-mediated Akt activation

    PubMed Central

    Zhang, Luo; Wang, Yiwu; Xiao, Fengjun; Wang, Shaoxia; Xing, Guichun; Li, Yang; Yin, Xiushan; Lu, Kefeng; Wei, Rongfei; Fan, Jiao; Chen, Yuhan; Li, Tao; Xie, Ping; Yuan, Lin; Song, Lei; Ma, Lanzhi; Ding, Lujing; He, Fuchu; Zhang, Lingqiang

    2014-01-01

    Macrophages play pivotal roles in development, homeostasis, tissue repair and immunity. Macrophage proliferation is promoted by macrophage colony-stimulating factor (M-CSF)-induced Akt signaling; yet, how this process is terminated remains unclear. Here, we identify casein kinase 2-interacting protein-1 (CKIP-1) as a novel inhibitor of macrophage proliferation. In resting macrophages, CKIP-1 was phosphorylated at Serine 342 by constitutively active GSK3β, the downstream target of Akt. This phosphorylation triggers the polyubiquitination and proteasomal degradation of CKIP-1. Upon M-CSF stimulation, Akt is activated by CSF-1R-PI3K and then inactivates GSK3β, leading to the stabilization of CKIP-1 and β-catenin proteins. β-catenin promotes the expression of proliferation genes including cyclin D and c-Myc. CKIP-1 interacts with TRAF6, a ubiquitin ligase required for K63-linked ubiquitination and plasma membrane recruitment of Akt, and terminates TRAF6-mediated Akt activation. By this means, CKIP-1 inhibits macrophage proliferation specifically at the late stage after M-CSF stimulation. Furthermore, CKIP-1 deficiency results in increased proliferation and decreased apoptosis of macrophages in vitro and CKIP-1−/− mice spontaneously develop a macrophage-dominated splenomegaly and myeloproliferation. Together, these data demonstrate that CKIP-1 plays a critical role in the regulation of macrophage homeostasis by inhibiting TRAF6-mediated Akt activation. PMID:24777252

  1. Andrographolide suppresses endothelial cell apoptosis via activation of phosphatidyl inositol-3-kinase/Akt pathway.

    PubMed

    Chen, Jiun-Han; Hsiao, George; Lee, An-Rong; Wu, Chin-Chen; Yen, Mao-Hsiung

    2004-04-01

    Andrographolide (Andro), an active component isolated from the Chinese official herbal Andrographis paniculata, which has been reported to prevent oxygen radical production and thus prevent inflammatory diseases. In this study, we investigated the molecular mechanisms and signaling pathways by which Andro protects human umbilical vein endothelial cells (HUVECs) from growth factor (GF) deprivation-induced apoptosis. Results demonstrated that HUVECs undergo apoptosis after 18 hr of GF deprivation but that this cell death was suppressed by the addition of Andro in a concentration-dependent manner (1-100 microM). Andro suppresses the mitochondrial pathway of apoptosis by inhibiting release of cytochrome c into the cytoplasm and dissipation of mitochondrial potential (Deltapsi(m)), as a consequence, prevented caspase-3 and -9 activation. Treatment of endothelial cells with Andro-induced activation of the protein kinase Akt, an anti-apoptotic signal, and phosphorylation of BAD, a down-stream target of Akt. Suppression of Akt activity by wortmannin, by LY-294002 and by using a dominant negative Akt mutant abolished the anti-apoptotic effect of Andro. In contrast, the ERK1/2 activities were not affected by Andro. The ERK1/2 inhibitor, PD98059 failed to antagonize the protective effect of Andro. In conclusion, Andro exerts its anti-apoptotic potential via activation of the Akt-BAD pathway in HUVECs and thus may represent a candidate of therapeutic agent for atherosclerosis.

  2. Regulation of PI-3-Kinase and Akt Signaling in T Lymphocytes and Other Cells by TNFR Family Molecules

    PubMed Central

    So, Takanori; Croft, Michael

    2013-01-01

    Activation of phosphoinositide 3-kinase (PI3K) and Akt (protein kinase B) is a common response triggered by a range of membrane-bound receptors on many cell types. In T lymphocytes, the PI3K-Akt pathway promotes clonal expansion, differentiation, and survival of effector cells and suppresses the generation of regulatory T cells. PI3K activation is tightly controlled by signals through the T cell receptor (TCR) and the co-stimulatory receptor CD28, however sustained and periodic signals from additional co-receptors are now being recognized as critical contributors to the activation of this pathway. Accumulating evidence suggests that many members of the Tumor Necrosis Factor receptor (TNFR) superfamily, TNFR2 (TNFRSF1B), OX40 (TNFRSF4), 4-1BB (TNFRSF9), HVEM (TNFRSF14), and DR3 (TNFRSF25), that are constitutive or inducible on T cells, can directly or indirectly promote activity in the PI3K-Akt pathway. We discuss recent data which suggests that ligation of one TNFR family molecule organizes a signalosome, via TNFR-associated factor (TRAF) adapter proteins in T cell membrane lipid microdomains, that results in the subsequent accumulation of highly concentrated depots of PI3K and Akt in close proximity to TCR signaling units. We propose this may be a generalizable mechanism applicable to other TNFR family molecules that will result in a quantitative contribution of these signalosomes to enhancing and sustaining PI3K and Akt activation triggered by the TCR. We also review data that other TNFR molecules, such as CD40 (TNFRSF5), RANK (TNFRSF11A), FN14 (TNFRSF12A), TACI (TNFRSF13B), BAFFR (TNFRSF13C), and NGFR (TNFRSF16), contribute to the activation of this pathway in diverse cell types through a similar ability to recruit PI3K or Akt into their signaling complexes. PMID:23760533

  3. Palmitoylethanolamide Modulates Inflammation-Associated Vascular Endothelial Growth Factor (VEGF) Signaling via the Akt/mTOR Pathway in a Selective Peroxisome Proliferator-Activated Receptor Alpha (PPAR-α)-Dependent Manner

    PubMed Central

    Sarnelli, Giovanni; D’Alessandro, Alessandra; Iuvone, Teresa; Capoccia, Elena; Gigli, Stefano; Pesce, Marcella; Seguella, Luisa; Nobile, Nicola; Aprea, Giovanni; Maione, Francesco; de Palma, Giovanni Domenico; Cuomo, Rosario; Steardo, Luca; Esposito, Giuseppe

    2016-01-01

    Background and Aim Angiogenesis is emerging as a pivotal process in chronic inflammatory pathologies, promoting immune infiltration and prompting carcinogenesis. Ulcerative Colitis (UC) and Crohn’s Disease (CD) represent paradigmatic examples of intestinal chronic inflammatory conditions in which the process of neovascularization correlates with the severity and progression of the diseases. Molecules able to target the angiogenesis have thus the potential to synergistically affect the disease course. Beyond its anti-inflammatory effect, palmitoylethanolamide (PEA) is able to reduce angiogenesis in several chronic inflammatory conditions, but no data about its anti-angiogenic activity in colitis have been produced, yet. Methods The effects of PEA on inflammation-associated angiogenesis in mice with dextran sulphate sodium (DSS)-induced colitis and in patients with UC were assessed. The release of Vascular Endothelial Growth Factor (VEGF), the hemoglobin tissue content, the expression of CD31 and of phosphatidylinositol 3-kinase/Akt/mammalian-target-of-rapamycin (mTOR) signaling axis were all evaluated in the presence of different concentrations of PEA and concomitant administration of PPAR-α and -γ antagonists. Results Our results demonstrated that PEA, in a selective peroxisome proliferator activated receptor (PPAR)-α dependent mechanism, inhibits colitis-associated angiogenesis, decreasing VEGF release and new vessels formation. Furthermore, we demonstrated that the mTOR/Akt axis regulates, at least partly, the angiogenic process in IBD and that PEA directly affects this pathway. Conclusions Our results suggest that PEA may improve inflammation-driven angiogenesis in colonic mucosa, thus reducing the mucosal damage and potentially affecting disease progression and the shift towards the carcinogenesis. PMID:27219328

  4. Hydrogen peroxide inhibits transforming growth factor-β1-induced cell cycle arrest by promoting Smad3 linker phosphorylation through activation of Akt-ERK1/2-linked signaling pathway.

    PubMed

    Choi, Jiyeon; Park, Seong Ji; Jo, Eun Ji; Lee, Hui-Young; Hong, Suntaek; Kim, Seong-Jin; Kim, Byung-Chul

    2013-06-14

    Hydrogen peroxide (H2O2) functions as a second messenger in growth factor receptor-mediated intracellular signaling cascade and is tumorigenic by virtue of its ability to promote cell proliferation; however, the mechanisms underlying the growth stimulatory action of H2O2 are less understood. Here we report an important mechanism for antagonistic effects of H2O2 on growth inhibitory response to transforming growth factor-β1 (TGF-β1). In Mv1Lu and HepG2 cells, pretreatment of H2O2 (0.05-0.2 mM) completely blocked TGF-β1-mediated induction of p15(INK4B) expression and increase of its promoter activity. Interestingly, H2O2 selectively suppressed the transcriptional activation potential of Smad3, not Smad2, in the absence of effects on TGF-β1-induced phosphorylation of the COOH-tail SSXS motif of Smad3 and its nuclear translocation. Mechanism studies showed that H2O2 increases the phosphorylation of Smad3 at the middle linker region in a concentration- and time-dependent manner and this effect is mediated by activation of extracellular signal-activated kinase 1/2 through Akt. Furthermore, expression of a mutant Smad3 in which linker phosphorylation sites were ablated significantly abrogated the inhibitory effects of H2O2 on TGF-β1-induced increase of p15(INK4B)-Luc reporter activity and blockade of cell cycle progression from G1 to S phase. These findings for the first time define H2O2 as a signaling molecule that modulate Smad3 linker phosphorylation and its transcriptional activity, thus providing a potential mechanism whereby H2O2 antagonizes the cytostatic function of TGF-β1.

  5. Stichopus japonicus Polysaccharide, Fucoidan, or Heparin Enhanced the SDF-1α/CXCR4 Axis and Promoted NSC Migration via Activation of the PI3K/Akt/FOXO3a Signaling Pathway.

    PubMed

    Cui, Chao; Wang, Peng; Cui, Ningshan; Song, Shuliang; Liang, Hao; Ji, Aiguo

    2016-11-01

    Stichopus japonicus Polysaccharide (SJP) is a sulfated polysaccharide from the body wall of the sea cucumber, Stichopus japonicus. Fucoidan is a heparinoid compound that belongs to a family of sulfated polyfucose polysaccharides. Heparin is a glycosaminoglycan. SJP, fucoidan, and heparin profoundly promoted stromal cell-derived factor 1 alpha (SDF-1α)-induced neural stem cell (NSC) migration in a concentration-dependent manner. In addition, the basal migration capacity of cells was significantly promoted after incubation with SJP, fucoidan, or heparin. Interaction of SJP, fucoidan, or heparin with SDF-1α efficiently showed additive effects on the promotion of cell migration from the neurosphere. SJP, fucoidan, or heparin interaction with SDF-1α treatment could increase Nestin expression. SDF-1α modulated by SJP, fucoidan, or heparin activated the CXCR4 receptor and directed cellular migration via the activation of the PI3K/Akt/FOXO3a signaling pathway. Moreover, interaction of SJP, fucoidan, or heparin with SDF-1α effectively promoted NSC migration and induced SDF-1α and CXCR4 expressions. Results suggested that SJP, fucoidan, and heparin might be good candidates for alleviating injury-initiated signals to which NSCs respond.

  6. Phosphatidylinositol 3-kinase-AKT-mammalian target of rapamycin (PI3K-Akt-mTOR) signaling pathway in non-small cell lung cancer

    PubMed Central

    2015-01-01

    Non-small cell lung cancer (NSCLC) is a devastating disease with poor prognosis. Systemic chemotherapy has been the mainstay of treatment in advanced disease for many decades. Personalized targeted therapy such as epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) and crizotinib has significantly changed the treatment paradigm in NSCLC. The future success of development of molecular targeted therapy relies on the understanding of signal transduction pathways. The PI3K-Akt-mTOR pathway is commonly deregulated in human malignancy including NSCLC. Therefore, this pathway is a target for many therapeutic developments. This review will provide an overview of PI3K-Akt-mTOR signaling pathway, genetic alterations activating the pathway and clinical therapeutic development of pathway inhibitors. PMID:25870799

  7. Signaling Through the PI 3-K, Akt, and SGK Pathway in Breast Cancer Progression

    DTIC Science & Technology

    2011-10-01

    ANSI Std. Z39.18 The aggressive behavior of malignant breast cancer is determined by a complex array of signaling pathways that regulate cell...Akt signaling promotes cancer progression. Many of the enzymes that regulate PI 3-K signaling are frequently mutated in human breast cancer , thereby...K, PIK3CA, is the most frequently mutated oncogene in breast cancer . However, recent studies have demonstrated that distinct Akt isoforms can either

  8. Enhanced Cardiac Akt/Protein Kinase B Signaling Contributes to Pathological Cardiac Hypertrophy in Part by Impairing Mitochondrial Function via Transcriptional Repression of Mitochondrion-Targeted Nuclear Genes

    PubMed Central

    Wende, Adam R.; O'Neill, Brian T.; Bugger, Heiko; Riehle, Christian; Tuinei, Joseph; Buchanan, Jonathan; Tsushima, Kensuke; Wang, Li; Caro, Pilar; Guo, Aili; Sloan, Crystal; Kim, Bum Jun; Wang, Xiaohui; Pereira, Renata O.; McCrory, Mark A.; Nye, Brenna G.; Benavides, Gloria A.; Darley-Usmar, Victor M.; Shioi, Tetsuo; Weimer, Bart C.

    2014-01-01

    Sustained Akt activation induces cardiac hypertrophy (LVH), which may lead to heart failure. This study tested the hypothesis that Akt activation contributes to mitochondrial dysfunction in pathological LVH. Akt activation induced LVH and progressive repression of mitochondrial fatty acid oxidation (FAO) pathways. Preventing LVH by inhibiting mTOR failed to prevent the decline in mitochondrial function, but glucose utilization was maintained. Akt activation represses expression of mitochondrial regulatory, FAO, and oxidative phosphorylation genes in vivo that correlate with the duration of Akt activation in part by reducing FOXO-mediated transcriptional activation of mitochondrion-targeted nuclear genes in concert with reduced signaling via peroxisome proliferator-activated receptor α (PPARα)/PGC-1α and other transcriptional regulators. In cultured myocytes, Akt activation disrupted mitochondrial bioenergetics, which could be partially reversed by maintaining nuclear FOXO but not by increasing PGC-1α. Thus, although short-term Akt activation may be cardioprotective during ischemia by reducing mitochondrial metabolism and increasing glycolysis, long-term Akt activation in the adult heart contributes to pathological LVH in part by reducing mitochondrial oxidative capacity. PMID:25535334

  9. 8-Amino-adenosine induces loss of phosphorylation of p38 mitogen-activated protein kinase, extracellular signal-regulated kinase 1/2, and Akt kinase: role in induction of apoptosis in multiple myeloma.

    PubMed

    Ghias, Kulsoom; Ma, Chunguang; Gandhi, Varsha; Platanias, Leonidas C; Krett, Nancy L; Rosen, Steven T

    2005-04-01

    Multiple myeloma is a slowly proliferating B-cell malignancy that accumulates apoptosis-resistant and replication-quiescent cell populations, posing a challenge for current chemotherapeutics that target rapidly replicating cells. Multiple myeloma remains an incurable disease in need of new therapeutic approaches. The purine nucleoside analogue, 8-amino-adenosine (8-NH2-Ado), exhibits potent activity in preclinical studies, inducing apoptosis in several multiple myeloma cell lines. This cytotoxic effect requires phosphorylation of 8-NH2-Ado to its triphosphate form, 8-amino-ATP, and results in a concomitant loss of endogenous ATP levels. Here, we show the novel effect of 8-NH2-Ado on the phosphorylation status of key cellular signaling molecules. Multiple myeloma cells treated with 8-NH2-Ado exhibit a dramatic loss of phosphorylation of several important signaling proteins, including extracellular signal-regulated kinase 1/2, p38 mitogen-activated protein kinase, and Akt kinase. Cells depleted of ATP independent of 8-NH2-Ado do not exhibit the same decrease in phosphorylation of vital cellular proteins. Therefore, the significant shifts in endogenous ATP pools caused by 8-NH2-Ado treatment cannot account for the changes in phosphorylation levels. Instead, 8-NH2-Ado may influence the activity of select regulatory protein kinases and/or phosphatases, with preliminary data suggesting that protein phophatase 2A activity is affected by 8-NH2-Ado. The distinctive effect of 8-NH2-Ado on the phosphorylation status of cellular proteins is a novel phenomenon for a nucleoside analogue drug and is unique to 8-NH2-Ado among this class of drugs. The kinetics of 8-NH2-Ado-mediated changes in phosphorylation levels of critical prosurvival and apoptosis-regulating proteins suggests that the modulation of these proteins by dephosphorylation at early time points may be an important mechanistic step in 8-NH2-Ado-induced apoptosis.

  10. Targeting hyperactivated DNA-PKcs by KU0060648 inhibits glioma progression and enhances temozolomide therapy via suppression of AKT signaling

    PubMed Central

    Qu, Yanming; Zhang, Mingshan; Wang, Haoran; Zhang, Zhihua; Zhou, Wei; Fan, Xinyi; Yu, Chunjiang; Zhan, Qimin; Song, Yongmei

    2016-01-01

    The overall survival remains undesirable in clinical glioma treatment. Inhibition of DNA-PKcs activity by its inhibitors suppresses tumor growth and enhances chemosensitivity of several tumors to chemotherapy. However, whether DNA-PKcs could be a potential target in glioma therapy remains unknown. In this study, we reported that the hyperactivated DNA-PKcs was profoundly correlated with glioma malignancy and observe a significant association between DNA-PKcs activation and survival of the glioma patients. Our data also found that inhibition of DNA-PKcs by its inhibitor KU0060648 sensitized glioma cells to TMZ in vitro. Specifically, we demonstrated that KU0060648 interrupted the formation of DNA-PKcs/AKT complex, leading to suppression of AKT signaling and resultantly enhanced TMZ efficacy. Combination of KU0060648 and TMZ substantially inhibited downstream effectors of AKT. The in vivo results were similar to those obtained in vitro. In conclusion, this study indicated that inhibition of DNA-PKcs activity could suppress glioma malignancies and increase TMZ efficacy, which was mainly through regulation of the of AKT signaling. Therefore, DNA-PKcs/AKT axis may be a promising target for improving current glioma therapy. PMID:27487130

  11. Depression of p53-independent Akt survival signals after high-LET radiation in mutated p53 cells

    NASA Astrophysics Data System (ADS)

    Ohnishi, Takeo; Takahashi, Akihisa; Nakagawa, Yosuke

    Although mutations and deletions in the p53 tumor suppressor gene lead to resistance to low linear energy transfer (LET) radiation, high-LET radiation efficiently induces cell lethality and apoptosis regardless of the p53 gene status. Recently, it has been suggested that the induction of p53-independent apoptosis takes place through the activation of Caspase-9 which results in the cleavage of Caspase-3 and poly (ADP-ribose) polymerase (PARP). This study was designed to examine if high-LET radiation depresses the activities of serine/threonine protein kinase B (PKB, also known as Akt) and Akt-related proteins. Human gingival cancer cells (Ca9-22 cells) harboring a mutated p53 (mp53) gene were irradiated with 2 Gy of X-rays or Fe-ion beams. The cellular contents of Akt-related proteins participating in cell survival signals were analyzed with Western blotting analysis 1 h, 2 h, 3 h and 6 h after irradiation. Cell cycle distributions after irradiation were assayed with flow cytometric analysis.Akt-related protein levels were decreased when cells were irradiated with high-LET radiation. High-LET radiation increased G _{2}/M phase arrests and suppressed the progression of the cell cycle much more efficiently when compared to low-LET radiation. These results suggest that high-LET radiation enhances apoptosis through the activation of Caspase-3 and Caspase-9, and depresses cell growth by suppressing Akt-related signals, even in the mp53 cells.

  12. Adiponectin Induces Oncostatin M Expression in Osteoblasts through the PI3K/Akt Signaling Pathway

    PubMed Central

    Su, Chen-Ming; Lee, Wei-Lin; Hsu, Chin-Jung; Lu, Ting-Ting; Wang, Li-Hong; Xu, Guo-Hong; Tang, Chih-Hsin

    2015-01-01

    Rheumatoid arthritis (RA), a common autoimmune disorder, is associated with a chronic inflammatory response and unbalanced bone metabolism within the articular microenvironment. Adiponectin, an adipokine secreted by adipocytes, is involved in multiple functions, including lipid metabolism and pro-inflammatory activity. However, the mechanism of adiponectin performance within arthritic inflammation remains unclear. In this study, we observed the effect of adiponectin on the expression of oncostatin M (OSM), a pro-inflammatory cytokine, in human osteoblastic cells. Pretreatment of cells with inhibitors of phosphatidylinositol 3-kinase (PI3K), Akt, and nuclear factor (NF)-κB reduced the adiponectin-induced OSM expression in osteoblasts. Stimulation of the cells with adiponectin increased phosphorylation of PI3K, Akt, and p65. Adiponectin treatment of osteoblasts increased OSM-luciferase activity and p65 binding to NF-κB on the OSM promoter. Our results indicate that adiponectin increased OSM expression via the PI3K, Akt, and NF-κB signaling pathways in osteoblastic cells, suggesting that adiponectin is a novel target for arthritis treatment. PMID:26712749

  13. p38 MAPK and PI3K/AKT Signalling Cascades inParkinson’s Disease

    PubMed Central

    Jha, Saurabh Kumar; Jha, Niraj Kumar; Kar, Rohan; Ambasta, Rashmi K; Kumar, Pravir

    2015-01-01

    Parkinson's disease (PD) is a chronic neurodegenerative condition which has the second largest incidence rate among all other neurodegenerative disorders barring Alzheimer's disease (AD). Currently there is no cure and researchers continue to probe the therapeutic prospect in cell cultures and animal models of PD. Out of the several factors contributing to PD prognosis, the role of p38 MAPK (Mitogen activated protein-kinase) and PI3K/AKT signalling module in PD brains is crucial because the impaired balance between the pro- apoptotic and anti-apoptotic pathways trigger unwanted phenotypes such as microglia activation, neuroinflammation, oxidative stress and apoptosis. These factors continue challenging the brain homeostasis in initial stages thereby essentially assisting the dopaminergic (DA) neurons towards progressive degeneration in PD. Neurotherapeutics against PD shall then be targeted against the misregulated accomplices of the p38 and PI3K/AKT cascades. In this review, we have outlined many such established mechanisms involving the p38 MAPK and PI3K/AKT pathways which can offer therapeutic windows for the rectification of aberrant DA neuronal dynamics in PD brains. PMID:26261796

  14. Polycystin-1 Induces Resistance to Apoptosis through the Phosphatidylinositol 3-Kinase/Akt Signaling Pathway

    PubMed Central

    Boca, Manila; Distefano, Gianfranco; Boletta, Alessandra; Qian, Feng; Bhunia, Anil K.; Germino, Gregory G.

    2006-01-01

    Polycystin-1 (PC-1), the PKD1 gene product, is a large receptor whose expression in renal epithelial cells results in resistance to apoptosis and tubulogenesis, a model consistent with the phenotype observed in patients. This study links PC-1 expression to a signaling pathway that is known to be both antiapoptotic and important for normal tubulogenesis. This study found that PC-1 expression results in phosphorylation of Akt and downstream effectors and that phosphatidylinositol 3-kinase (PI3-K) inhibitors prevent this process. In addition, it is shown that dominant negative Akt can revert PC-1-induced protection from apoptosis. Furthermore, it was observed that increased PI3-K β activity in PC-1- expressing MDCK cells seems to be dependent on both tyrosine-kinase activity and heterotrimeric G proteins. It also was found that PC-1-induced tubulogenesis is inhibited by PI3-K inhibitors. Taken together, these data suggest that the PI3-K/Akt cascade may be a central modulator of PC-1 function and that its deregulation might be important in autosomal dominant polycystic kidney disease. PMID:16452497

  15. High fat diet reduces neuroprotection of isoflurane post-treatment: role of carboxyl-terminal modulator protein-Akt signaling

    PubMed Central

    Yu, Hai; Deng, Jiao; Zuo, Zhiyi

    2014-01-01

    Objective High fat diet (HFD) contributes to the increased prevalence of obesity and hyperlipidemia in young adults, a possible cause for their recent increase in stroke. Isoflurane post-treatment provides neuroprotection. We determined whether isoflurane post-treatment induced neuroprotection in HFD-fed mice. Design and Methods Six-week old CD-1 male mice were fed HFD or regular diet (RD) for 5 or 10 weeks. Their hippocampal slices (400 µm) were subjected to oxygen-glucose deprivation (OGD). Some slices were exposed to isoflurane for 30 min immediately after OGD. Some mice had a 90-min middle cerebral arterial occlusion and were post-treated with 2% isoflurane for 30 min. Results OGD time-dependently induced cell injury. This injury was dose-dependently reduced by isoflurane. The effect was apparent at 1% or 2% isoflurane in RD-fed mice but required 3% isoflurane in HFD-fed mice. HFD influenced the isoflurane effects in DG. OGD increased carboxyl-terminal modulator protein (CTMP), an Akt inhibitor, and decreased Akt signaling. Isoflurane reduced these effects. LY294002, an Akt activation inhibitor, attenuated the isoflurane effects. HFD increased CTMP and reduced Akt signaling. Isoflurane improved neurological outcome in the RD-fed mice but not in the HFD-fed mice. Conclusions HFD attenuated isoflurane post-treatment-induced neuroprotection possibly due to decreased prosurvival Akt signaling. PMID:25142024

  16. SDF-1α/CXCR4 Signaling in Lipid Rafts Induces Platelet Aggregation via PI3 Kinase-Dependent Akt Phosphorylation

    PubMed Central

    Hayashi, Moyuru; Kaneda, Mizuho; Iida, Kazuko; Shimonaka, Motoyuki; Hara, Takahiko; Arai, Morio; Koike, Yuichi; Yamamoto, Naomasa; Kasahara, Kohji

    2017-01-01

    Stromal cell-derived factor-1α (SDF-1α)-induced platelet aggregation is mediated through its G protein-coupled receptor CXCR4 and phosphatidylinositol 3 kinase (PI3K). Here, we demonstrate that SDF-1α induces phosphorylation of Akt at Thr308 and Ser473 in human platelets. SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the CXCR4 antagonist AMD3100 or the PI3K inhibitor LY294002. SDF-1α also induces the phosphorylation of PDK1 at Ser241 (an upstream activator of Akt), GSK3β at Ser9 (a downstream substrate of Akt), and myosin light chain at Ser19 (a downstream element of the Akt signaling pathway). SDF-1α-induced platelet aggregation is inhibited by pretreatment with the Akt inhibitor MK-2206 in a dose-dependent manner. Furthermore, SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the raft-disrupting agent methyl-β-cyclodextrin. Sucrose density gradient analysis shows that 35% of CXCR4, 93% of the heterotrimeric G proteins Gαi-1, 91% of Gαi-2, 50% of Gβ and 4.0% of PI3Kβ, and 4.5% of Akt2 are localized in the detergent-resistant membrane raft fraction. These findings suggest that SDF-1α/CXCR4 signaling in lipid rafts induces platelet aggregation via PI3K-dependent Akt phosphorylation. PMID:28072855

  17. SDF-1α/CXCR4 Signaling in Lipid Rafts Induces Platelet Aggregation via PI3 Kinase-Dependent Akt Phosphorylation.

    PubMed

    Ohtsuka, Hiroko; Iguchi, Tomohiro; Hayashi, Moyuru; Kaneda, Mizuho; Iida, Kazuko; Shimonaka, Motoyuki; Hara, Takahiko; Arai, Morio; Koike, Yuichi; Yamamoto, Naomasa; Kasahara, Kohji

    2017-01-01

    Stromal cell-derived factor-1α (SDF-1α)-induced platelet aggregation is mediated through its G protein-coupled receptor CXCR4 and phosphatidylinositol 3 kinase (PI3K). Here, we demonstrate that SDF-1α induces phosphorylation of Akt at Thr308 and Ser473 in human platelets. SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the CXCR4 antagonist AMD3100 or the PI3K inhibitor LY294002. SDF-1α also induces the phosphorylation of PDK1 at Ser241 (an upstream activator of Akt), GSK3β at Ser9 (a downstream substrate of Akt), and myosin light chain at Ser19 (a downstream element of the Akt signaling pathway). SDF-1α-induced platelet aggregation is inhibited by pretreatment with the Akt inhibitor MK-2206 in a dose-dependent manner. Furthermore, SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the raft-disrupting agent methyl-β-cyclodextrin. Sucrose density gradient analysis shows that 35% of CXCR4, 93% of the heterotrimeric G proteins Gαi-1, 91% of Gαi-2, 50% of Gβ and 4.0% of PI3Kβ, and 4.5% of Akt2 are localized in the detergent-resistant membrane raft fraction. These findings suggest that SDF-1α/CXCR4 signaling in lipid rafts induces platelet aggregation via PI3K-dependent Akt phosphorylation.

  18. Avian Reovirus Protein p17 Functions as a Nucleoporin Tpr Suppressor Leading to Activation of p53, p21 and PTEN and Inactivation of PI3K/AKT/mTOR and ERK Signaling Pathways

    PubMed Central

    Huang, Wei-Ru; Chiu, Hung-Chuan; Liao, Tsai-Ling; Chuang, Kuo-Pin; Shih, Wing-Ling; Liu, Hung-Jen

    2015-01-01

    Avian reovirus (ARV) protein p17 has been shown to regulate cell cycle and autophagy by activation of p53/PTEN pathway; nevertheless, it is still unclear how p53 and PTEN are activated by p17. Here, we report for the first time that p17 functions as a nucleoporin Tpr suppressor that leads to p53 nuclear accumulation and consequently activates p53, p21, and PTEN. The nuclear localization signal (119IAAKRGRQLD128) of p17 has been identified for Tpr binding. This study has shown that Tpr suppression occurs by p17 interacting with Tpr and by reducing the transcription level of Tpr, which together inhibit Tpr function. In addition to upregulation of PTEN by activation of p53 pathway, this study also suggests that ARV protein p17 acts as a positive regulator of PTEN. ARV p17 stabilizes PTEN by stimulating phosphorylation of cytoplasmic PTEN and by elevating Rak-PTEN association to prevent it from E3 ligase NEDD4-1 targeting. To activate PTEN, p17 is able to promote β-arrestin-mediated PTEN translocation from the cytoplasm to the plasma membrane via a Rock-1-dependent manner. The accumulation of p53 in the nucleus induces the PTEN- and p21-mediated downregulation of cyclin D1 and CDK4. Furthermore, Tpr and CDK4 knockdown increased virus production in contrast to depletion of p53, PTEN, and LC3 reducing virus yield. Taken together, our data suggest that p17-mediated Tpr suppression positively regulates p53, PTEN, and p21 and negatively regulates PI3K/AKT/mTOR and ERK signaling pathways, both of which are beneficial for virus replication. PMID:26244501

  19. Membrane Heterogeneity in Akt Activation in Prostate Cancer

    DTIC Science & Technology

    2009-03-01

    prenylation signal derived from K- Ras rendered Akt1 catalytically inactive when fused to the N-terminal PH-domain (Fig. 1). Fusion of this motif to...were treated with either cyclo- dextrin (CD) or water-soluble cholesterol ( Chol ) or with CD followed by cholesterol treatment (CD Chol ). Cells

  20. The HMG-CoA reductase inhibitor simvastatin activates the protein kinase Akt and promotes angiogenesis in normocholesterolemic animals

    PubMed Central

    Kureishi, Yasuko; Luo, Zhengyu; Shiojima, Ichiro; Bialik, Ann; Fulton, David; Lefer, David J.; Sessa, William C.; Walsh, Kenneth

    2010-01-01

    Recent studies suggest that statins can function to protect the vasculature in a manner that is independent of their lipid-lowering activity. We show here that statins rapidly activate the protein kinase Akt/PKB in endothelial cells. Accordingly, simvastatin enhanced phosphorylation of the endogenous Akt substrate endothelial nitric oxide synthase (eNOS), inhibited apoptosis and accelerated vascular structure formation in vitro in an Akt-dependent manner. Similar to vascular endothelial growth factor (VEGF) treatment, both simvastatin administration and enhanced Akt signaling in the endothelium promoted angiogenesis in ischemic limbs of normocholesterolemic rabbits. Therefore, activation of Akt represents a mechanism that can account for some of the beneficial side effects of statins, including the promotion of new blood vessel growth. PMID:10973320

  1. PI-103 and Quercetin Attenuate PI3K-AKT Signaling Pathway in T- Cell Lymphoma Exposed to Hydrogen Peroxide

    PubMed Central

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2016-01-01

    Phosphatidylinositol 3 kinase—protein kinase B (PI3K-AKT) pathway has been considered as major drug target site due to its frequent activation in cancer. AKT regulates the activity of various targets to promote tumorigenesis and metastasis. Accumulation of reactive oxygen species (ROS) has been linked to oxidative stress and regulation of signaling pathways for metabolic adaptation of tumor microenvironment. Hydrogen peroxide (H2O2) in this context is used as ROS source for oxidative stress preconditioning. Antioxidants are commonly considered to be beneficial to reduce detrimental effects of ROS and are recommended as dietary supplements. Quercetin, a ubiquitous bioactive flavonoid is a dietary component which has attracted much of interest due to its potential health-promoting effects. Present study is aimed to analyze PI3K-AKT signaling pathway in H2O2 exposed Dalton’s lymphoma ascite (DLA) cells. Further, regulation of PI3K-AKT pathway by quercetin as well as PI-103, an inhibitor of PI3K was analyzed. Exposure of H2O2 (1mM H2O2 for 30min) to DLA cells caused ROS accumulation and resulted in increased phosphorylation of PI3K and downstream proteins PDK1 and AKT (Ser-473 and Thr-308), cell survival factors BAD and ERK1/2, as well as TNFR1. However, level of tumor suppressor PTEN was declined. Both PI-103 & quercetin suppressed the enhanced level of ROS and significantly down-regulated phosphorylation of AKT, PDK1, BAD and level of TNFR1 as well as increased the level of PTEN in H2O2 induced lymphoma cells. The overall result suggests that quercetin and PI3K inhibitor PI-103 attenuate PI3K-AKT pathway in a similar mechanism. PMID:27494022

  2. PI-103 and Quercetin Attenuate PI3K-AKT Signaling Pathway in T- Cell Lymphoma Exposed to Hydrogen Peroxide.

    PubMed

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2016-01-01

    Phosphatidylinositol 3 kinase-protein kinase B (PI3K-AKT) pathway has been considered as major drug target site due to its frequent activation in cancer. AKT regulates the activity of various targets to promote tumorigenesis and metastasis. Accumulation of reactive oxygen species (ROS) has been linked to oxidative stress and regulation of signaling pathways for metabolic adaptation of tumor microenvironment. Hydrogen peroxide (H2O2) in this context is used as ROS source for oxidative stress preconditioning. Antioxidants are commonly considered to be beneficial to reduce detrimental effects of ROS and are recommended as dietary supplements. Quercetin, a ubiquitous bioactive flavonoid is a dietary component which has attracted much of interest due to its potential health-promoting effects. Present study is aimed to analyze PI3K-AKT signaling pathway in H2O2 exposed Dalton's lymphoma ascite (DLA) cells. Further, regulation of PI3K-AKT pathway by quercetin as well as PI-103, an inhibitor of PI3K was analyzed. Exposure of H2O2 (1mM H2O2 for 30min) to DLA cells caused ROS accumulation and resulted in increased phosphorylation of PI3K and downstream proteins PDK1 and AKT (Ser-473 and Thr-308), cell survival factors BAD and ERK1/2, as well as TNFR1. However, level of tumor suppressor PTEN was declined. Both PI-103 & quercetin suppressed the enhanced level of ROS and significantly down-regulated phosphorylation of AKT, PDK1, BAD and level of TNFR1 as well as increased the level of PTEN in H2O2 induced lymphoma cells. The overall result suggests that quercetin and PI3K inhibitor PI-103 attenuate PI3K-AKT pathway in a similar mechanism.

  3. TRIM14 regulates cell proliferation and invasion in osteosarcoma via promotion of the AKT signaling pathway

    PubMed Central

    Xu, Guoxing; Guo, Yongfei; Xu, Dabo; Wang, Yi; Shen, Yafeng; Wang, Feifei; Lv, Yuanyuan; Song, Fanglong; Jiang, Dawei; Zhang, Yinquan; Lou, Yi; Meng, Yake; Yang, Yongji; Kang, Yifan

    2017-01-01

    Recent studies have shown that some members of the tripartite motif-containing protein (TRIM) family serve as important regulators of tumorigenesis. However, the biological role of TRIM14 in osteosarcoma remains to be established. In this study, we showed that TRIM14 is upregulated in human osteosarcoma specimens and cell lines, and correlated with osteosarcoma progression and shorter patient survival times. Functional studies demonstrated that overexpression of TRIM14 enhances osteosarcoma cell proliferation, clone formation, cell cycle procession, migration and invasion in vitro and promotes tumor growth in vivo, and conversely, its silencing has the opposite effects. Furthermore, TRIM14 overexpression induced activation of the AKT pathway. Inhibition of AKT expression reversed the TRIM14-mediated promotory effects on cell growth and mobility, in addition to TRIM14-induced epithelial-to-mesenchymal transition (EMT) and cyclin D1 upregulation. Our findings collectively suggest that TRIM14 functions as an oncogene by upregulating the AKT signaling pathway in osteosarcoma cells, supporting its potential utility as a therapeutic target for this disease. PMID:28205534

  4. Inhibition of hydrogen peroxide signaling by 4-hydroxynonenal due to differential regulation of Akt1 and Akt2 contributes to decreases in cell survival and proliferation in hepatocellular carcinoma cells.

    PubMed

    Shearn, Colin T; Reigan, Philip; Petersen, Dennis R

    2012-07-01

    Dysregulation of cell signaling by electrophiles such as 4-hydroxynonenal (4-HNE) is a key component in the pathogenesis of chronic inflammatory liver disease. Another consequence of inflammation is the perpetuation of oxidative damage by the production of reactive oxidative species such as hydrogen peroxide. Previously, we have demonstrated Akt2 as a direct target of 4-HNE in hepatocellular carcinoma cells. In the present study, we used the hepatocellular carcinoma cell line HepG2 as model to understand the combinatorial effects of 4-HNE and hydrogen peroxide. We demonstrate that 4-HNE inhibits hydrogen peroxide-mediated phosphorylation of Akt1 but not Akt2. Pretreatment of HepG2 cells with 4-HNE prevented hydrogen peroxide stimulation of Akt-dependent phosphorylation of downstream targets and intracellular Akt activity compared with untreated control cells. Using biotin hydrazide capture, it was confirmed that 4-HNE treatment resulted in carbonylation of Akt1, which was not observed in untreated control cells. Using a synthetic GSK3α/β peptide as a substrate, treatment of recombinant human myristoylated Akt1 (rAkt1) with 20 or 40 μΜ 4-HNE inhibited rAkt1 activity by 29 and 60%, respectively. We further demonstrate that 4-HNE activates Erk via a PI3 kinase and PP2A-dependent mechanism leading to increased Jnk phosphorylation. At higher concentrations, 4-HNE decreased both cell survival and proliferation as evidenced by MTT assays and EdU incorporation as well as decreased expression of cyclin D1 and β-catenin, an effect only moderately increased by the addition of hydrogen peroxide. The ability of 4-HNE to exert combinatorial effects on Erk, Jnk, and Akt-dependent cell survival pathways provides additional insight into the mechanisms of cellular damage associated with chronic inflammation.

  5. The role of the PI3K-Akt signal transduction pathway in Autographa californica multiple nucleopolyhedrovirus infection of Spodoptera frugiperda cells

    SciTech Connect

    Xiao Wei; Yang Yi; Weng Qingbei; Lin Tiehao; Yuan Meijin; Yang Kai; Pang Yi

    2009-08-15

    Many viruses activate the phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway, thereby modulating diverse downstream signaling pathways associated with antiapoptosis, proliferation, cell cycling, protein synthesis and glucose metabolism, in order to augment their replication. To date, the role of the PI3K-Akt pathway in Baculovirus replication has not been defined. In the present study, we demonstrate that infection of Sf9 cells with Autographa californica multiple nucleopolyhedrovirus (AcMNPV) elevated cellular Akt phosphorylation at 1 h post-infection. The maximum Akt phosphorylation occurred at 6 h post-infection and remained unchanged until 18 h post-infection. The PI3K-specific inhibitor, LY294002, suppressed Akt phosphorylation in a dose-dependent manner, suggesting that AcMNPV-induced Akt phosphorylation is PI3K-dependent. The inhibition of PI3K-Akt activation by LY294002 significantly reduced the viral yield, including a reduction in budded viruses and occlusion bodies. The virus production was reduced only when the inhibitor was added within 24 h of infection, implying that activation of PI3K occurred early in infection. Correspondingly, both viral DNA replication and late (VP39) and very late (POLH) viral protein expression were impaired by LY294002 treatment; LY294002 had no effect on immediate-early (IE1) and early-late (GP64) protein expression. These results demonstrate that the PI3K-Akt pathway is required for efficient Baculovirus replication.

  6. Long-Term Administration of Dehydroepiandrosterone Accelerates Glucose Catabolism via Activation of PI3K/Akt-PFK-2 Signaling Pathway in Rats Fed a High-Fat Diet

    PubMed Central

    Kang, Jian; Ge, Chongyang; Yu, Lei; Li, Longlong; Ma, Haitian

    2016-01-01

    Dehydroepiandrosterone (DHEA) has a fat-reducing effect, while little information is available on whether DHEA regulates glucose metabolism, which would in turn affect fat deposition. To investigate the effects of DHEA on glucose metabolism, rats were administered a high-fat diet containing either 0 (HCG), 25 (HLG), 50 (HMG), or 100 (HHG) mg·kg-1 DHEA per day via gavage for 8 weeks. Results showed that long-term administration of DHEA inhibited body weight gain in rats on a high-fat diet. No statistical differences in serum glucose levels were observed, whereas hepatic glycogen content in HMG and HHG groups and muscle glycogen content in HLG and HMG groups were higher than those in HCG group. Glucokinase, malate dehydrogenase and phosphofructokinase-2 activities in HMG and HHG groups, pyruvate kinase and succinate dehydrogenase activities in HMG group, and pyruvate dehydrogenase activity in all DHEA treatment groups were increased compared with those in HCG group. Phosphoenolpyruvate carboxykinase and glycogen phosphorylase mRNA levels were decreased in HMG and HHG groups, whereas glycogen synthase-2 mRNA level was increased in HMG group compared with those in HCG. The abundance of Glut2 mRNA in HMG and HHG groups and Glut4 mRNA in HMG group was higher than that in HCG group. DHEA treatment increased serum leptin content in HMG and HHG groups compared with that in HCG group. Serum insulin content and insulin receptor mRNA level in HMG group and insulin receptor substrate-2 mRNA level in HMG and HHG group were increased compared with those in HCG group. Furthermore, Pi3k mRNA level in HMG and Akt mRNA level in HMG and HHG groups were significantly increased than those in HCG group. These data showed that DHEA treatment could enhance glycogen storage and accelerate glucose catabolism in rats fed a high-fat diet, and this effect may be associated with the activation of PI3K/Akt-PFK-2 signaling pathway. PMID:27410429

  7. Honokiol induces autophagy of neuroblastoma cells through activating the PI3K/Akt/mTOR and endoplasmic reticular stress/ERK1/2 signaling pathways and suppressing cell migration.

    PubMed

    Yeh, Poh-Shiow; Wang, Weu; Chang, Ya-An; Lin, Chien-Ju; Wang, Jhi-Joung; Chen, Ruei-Ming

    2016-01-01

    autophagy of neuroblastoma cells and consequent apoptosis through activating the PI3K/Akt/mTOR and ERS/ROS/ERK1/2 signaling pathways and suppressing cell migration. Thus, honokiol has potential for treating neuroblastomas.

  8. Non-tumorigenic epithelial cells secrete MCP-1 and other cytokines that promote cell division in breast cancer cells by activating ERα via PI3K/Akt/mTOR signaling.

    PubMed

    Riverso, Maria; Kortenkamp, Andreas; Silva, Elisabete

    2014-08-01

    Efforts in understanding the role of the microenvironment in the development of breast cancer have focused on tumor-stroma cross-talk, but the possibility that normal epithelial cells might also play a role in tumor progression has received little attention. Here, we show that non-tumorigenic human mammary epithelial cells (MCF10A and HMEC) secrete factors able to enhance the proliferation of estrogen receptor α (ERα) positive breast cancer cells (MCF7 and T47D) and suppress their ability to undergo apoptosis. Conditioned medium (CM) derived from MCF10A and HMEC cells was capable of activating ERα in a hormone-independent way, by phosphorylating ERα on Ser167. Co-exposure with PI3K and mTORC1 inhibitors significantly reduced the ERα Ser167 phosphorylation and suppressed the proliferation-enhancing effects of both 10A-CM and HMEC-CM on MCF7 cells. We show that MCF10A and HMEC secrete numerous cytokines, among them MCP-1, which was one of the most prevalent. MCP-1 was shown to have a role in the effects elicited by the 10A-CM. It activated the ERα by phosphorylating Ser167 via the PI3K/Akt/mTORC1 signaling pathway, an effect which was further confirmed by silencing the MCP-1 receptors, CCR2 and CCR4. To our knowledge, this is the first time MCP-1 has been shown to contribute to ERα signaling activation. These data suggest that normal mammary cells could have the capability of supporting the proliferation of breast cancer cells via paracrine interactions. A better understanding of the role of these cells may be useful for designing strategies for the prevention of tumor progression at early stages.

  9. PI-103 attenuates PI3K-AKT signaling and induces apoptosis in murineT-cell lymphoma.

    PubMed

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2017-05-01

    Aberrant activation of PI3K-AKT signaling in many pathological conditions including cancer has attracted much of interest for drug targeting. Various isoforms are known from three classes of PI3K. Targeting selective isoform is advantageous to overcome the global deleterious effects of drug. PI-103 is a specific inhibitor of p110α of class I PI3K. The present study is aimed to analyze anti-carcinogenic activity of PI-103 in Dalton's lymphoma ascite (DLA) cells. Result shows regression in cell proliferation and increased apoptosis in terms of increased Annexin V binding, nuclear fragmentation and active caspase 3 level. It is correlated with attenuation of PI3K-AKT signaling by PI-103 via downregulation of the level of p110α, phospho-p85α, phospho- AKT, and PKCα in DLA cells as well as in H2O2 induced DLA cells. Additionally, ROS accumulation is declined in H2O2 induced DLA cells. Overall result suggests that PI-103 attenuates PI3K-AKT signaling via induction of apoptosis in murine T-cell lymphoma.

  10. Activation of PI3K/Akt pathway limits JNK-mediated apoptosis during EV71 infection.

    PubMed

    Zhang, Hua; Li, Fengqi; Pan, Ziye; Wu, Zhijun; Wang, Yanhong; Cui, Yudong

    2014-11-04

    Apoptosis is frequently induced to inhibit virus replication during infection of Enterovirus 71 (EV71). On the contrary, anti-apoptotic pathway, such as PI3K/Akt pathway, is simultaneously exploited by EV71 to accomplish the viral life cycle. The relationship by which EV71-induced apoptosis and PI3K/Akt signaling pathway remains to be elucidated. In this study, we demonstrated that EV71 infection altered Bax conformation and triggered its redistribution from the cytosol to mitochondria in RD cells. Subsequently, cytochrome c was released from mitochondria to cytosol. We also found that c-Jun NH2-terminal kinase (JNK) was activated during EV71 infection. The JNK specific inhibitor significantly inhibited Bax activation and cytochrome c release, suggesting that EV71-induced apoptosis was involved into a JNK-dependent manner. Meanwhile, EV71-induced Akt phosphorylation involved a PI3K-dependent mechanism. Inhibition of the PI3K/Akt pathway enhanced JNK phosphorylation and the JNK-mediated apoptosis upon EV71 infection. Moreover, PI3K/Akt pathway phosphorylated apoptosis signal-regulating kinase 1 (ASK1) and negatively regulated the ASK1 activity. Knockdown of ASK1 significantly decreased JNK phosphorylation, which implied that ASK1 phosphorylation by Akt inhibited ASK1-mediated JNK activation. Collectively, these data reveal that activation of the PI3K/Akt pathway limits JNK-mediated apoptosis by phosphorylating and inactivating ASK1 during EV71 infection.

  11. Stat5 Promotes Survival of Mammary Epithelial Cells through Transcriptional Activation of a Distinct Promoter in Akt1▿

    PubMed Central

    Creamer, Bradley A.; Sakamoto, Kazuhito; Schmidt, Jeffrey W.; Triplett, Aleata A.; Moriggl, Richard; Wagner, Kay-Uwe

    2010-01-01

    The signal transducer and activator of transcription 5 (Stat5) plays a pivotal role in the proliferation, secretory differentiation, and survival of mammary epithelial cells. However, there is little information about Stat5 target genes that facilitate these biological processes. We provide here experimental evidence that the prolactin-mediated phosphorylation of Stat5 regulates the transcriptional activation of the Akt1 gene. Stat5 binds to consensus sequences within the Akt1 locus in a growth factor-dependent manner to initiate transcription of a unique Akt1 mRNA from a distinct promoter, which is only active in the mammary gland. Elevating the levels of active Akt1 restores the expression of cyclin D1 and proliferation of Jak2-deficient mammary epithelial cells, which provides evidence that Akt1 acts downstream of Jak/Stat signaling. The ligand-inducible expression of Stat5 in transgenic females mediates a sustained upregulation of Akt1 in mammary epithelial cells during the onset of postlactational involution. Stat5-expressing mammary glands exhibit a delay in involution despite induction of proapoptotic signaling events. Collectively, the results of the present study elucidate an underlying mechanism by which active Stat5 mediates evasion from apoptosis and self-sufficiency in growth signals. PMID:20385773

  12. Inhibition of Protein Kinase Akt1 by Apoptosis Signal-regulating Kinase-1 (ASK1) Is Involved in Apoptotic Inhibition of Regulatory Volume Increase*

    PubMed Central

    Subramanyam, Muthangi; Takahashi, Nobuyuki; Hasegawa, Yuichi; Mohri, Tatsuma; Okada, Yasunobu

    2010-01-01

    Most animal cell types regulate their cell volume after an osmotic volume change. The regulatory volume increase (RVI) occurs through uptake of NaCl and osmotically obliged water after osmotic shrinkage. However, apoptotic cells undergo persistent cell shrinkage without showing signs of RVI. Persistence of the apoptotic volume decrease is a prerequisite to apoptosis induction. We previously demonstrated that volume regulation is inhibited in human epithelial HeLa cells stimulated with the apoptosis inducer. Here, we studied signaling mechanisms underlying the apoptotic inhibition of RVI in HeLa cells. Hypertonic stimulation was found to induce phosphorylation of a Ser/Thr protein kinase Akt (protein kinase B). Shrinkage-induced Akt activation was essential for RVI induction because RVI was suppressed by an Akt inhibitor, expression of a dominant negative form of Akt, or small interfering RNA-mediated knockdown of Akt1 (but not Akt2). Staurosporine, tumor necrosis factor-α, or a Fas ligand inhibited both RVI and hypertonicity-induced Akt activation in a manner sensitive to a scavenger for reactive oxygen species (ROS). Any of apoptosis inducers also induced phosphorylation of apoptosis signal-regulating kinase 1 (ASK1) in a ROS-dependent manner. Suppression of (ASK1) expression blocked the effects of apoptosis, in hypertonic conditions, on both RVI induction and Akt activation. Thus, it is concluded that in human epithelial cells, shrinkage-induced activation of Akt1 is involved in the RVI process and that apoptotic inhibition of RVI is caused by inhibition of Akt activation, which results from ROS-mediated activation of ASK1. PMID:20048146

  13. Activation of the phosphatidylinositol 3-kinase/Akt pathway is involved in lipocalin-2-promoted human pulmonary artery smooth muscle cell proliferation.

    PubMed

    Wang, Guoliang; Ma, Ning; Meng, Liukun; Wei, Yingjie; Gui, Jingang

    2015-12-01

    Over-activated PI3K/Akt signaling, a pathway strongly related to cancer survival and proliferation, has been reported recently to be involved in pulmonary artery smooth muscle cell apoptosis and proliferation in pulmonary hypertension (PH). In this study, we observed greatly increased lipocalin-2 (Lcn2) expression accompanied with over-activated PI3K/Akt signaling in a standard rat model of PH induced by monocrotaline. In view of the close relationship between Lcn2 and PI3K/Akt pathway, we hypothesized that the up-regulated Lcn2 might be a trigger of over-activated PI3K/Akt signaling in PH. Our results showed that Lcn2 significantly activated the PI3K/Akt pathway (determined by augmented Akt phosphorylation and up-regulated Mdm2) and significantly promoted proliferation (assessed by Ki67 staining) in cultured human pulmonary artery smooth muscle cells. Furthermore, we demonstrated that inhibition of Akt phosphorylation (LY294002) abrogated the Lcn2-promoted proliferation in cultured human pulmonary artery smooth muscle cells. In conclusion, Lcn2 significantly promoted human pulmonary artery smooth muscle cell proliferation by activating PI3K/Akt pathway. Further study on the role and mechanism of Lcn2 will help explore novel therapeutic strategies based on attenuating over-activated PI3K/Akt signaling in PH.

  14. 5-Caffeoylquinic acid inhibits invasion of non-small cell lung cancer cells through the inactivation of p70S6K and Akt activity: Involvement of p53 in differential regulation of signaling pathways.

    PubMed

    In, Jae-Kyung; Kim, Jin-Kyu; Oh, Joa Sub; Seo, Dong-Wan

    2016-05-01

    In the present study, we investigated the effects and molecular mechanism of 5-caffeoylquinic acid (5-CQA), a natural phenolic compound isolated from Ligularia fischeri, on cell invasion, proliferation and adhesion in p53 wild-type A549 and p53-deficient H1299 non-small cell lung cancer (NSCLC) cells. 5-CQA abrogated mitogen-stimulated invasion, but not proliferation, in both A549 and H1299 cells. In addition, 5-CQA inhibited mitogen-stimulated adhesion in A549 cells only. Anti-invasive activity of 5-CQA in A549 cells was mediated by the inactivation of p70(S6K)-dependent signaling pathway. In contrast, in H1299 cells the inactivation of Akt was found to be involved in 5-CQA-mediated inhibition of cell invasion. Collectively, these findings demonstrate the pharmacological roles and molecular targets of 5-CQA in regulating NSCLC cell fate, and suggest further evaluation and development of 5-CQA as a potential therapeutic agent for the treatment and prevention of lung cancer.

  15. Germline activating AKT3 mutation associated with megalencephaly, polymicrogyria, epilepsy and hypoglycemia.

    PubMed

    Nellist, Mark; Schot, Rachel; Hoogeveen-Westerveld, Marianne; Neuteboom, Rinze F; van der Louw, Elles J T M; Lequin, Maarten H; Bindels-de Heus, Karen; Sibbles, Barbara J; de Coo, René; Brooks, Alice; Mancini, Grazia M S

    2015-03-01

    Activating germ-line and somatic mutations in AKT3 (OMIM 611223) are associated with megalencephaly-polymicrogyria-polydactyly-hydrocephalus syndrome (MPPH; OMIM # 615937) and megalencephaly-capillary malformation (MCAP; OMIM # 602501). Here we report an individual with megalencephaly, polymicrogyria, refractory epilepsy, hypoglycemia and a germline AKT3 mutation. At birth, head circumference was 43 cm (5 standard deviations above the mean). No organomegaly was present, but there was generalized hypotonia, joint and skin laxity, developmental delay and failure to thrive. At 6 months of age the patient developed infantile spasms that were resistant to antiepileptic polytherapy. Recurrent hypoglycemia was noted during treatment with adrenocorticotropic hormone but stabilized upon introduction of continuous, enriched feeding. The infantile spasms responded to the introduction of a ketogenic diet, but the hypoglycemia recurred until the diet was adjusted for increased resting energy expenditure. A novel, de novo AKT3 missense variant (exon 5; c.548T>A, p.(V183D)) was identified and shown to activate AKT3 by in vitro functional testing. We hypothesize that the sustained hypoglycemia in this patient is caused by increased glucose utilization due to activation of AKT3 signaling. This might explain the efficacy of the ketogenic diet in this individual.

  16. Activation of spinal chemokine receptor CXCR3 mediates bone cancer pain through an Akt-ERK crosstalk pathway in rats.

    PubMed

    Guan, Xue-Hai; Fu, Qiao-Chu; Shi, Dai; Bu, Hui-Lian; Song, Zhen-Peng; Xiong, Bing-Rui; Shu, Bin; Xiang, Hong-Bing; Xu, Bing; Manyande, Anne; Cao, Fei; Tian, Yu-Ke

    2015-01-01

    Previously, we showed that activation of the spinal CXCL9, 10/CXCR3 pathway mediated bone cancer pain (BCP) in rats. However, the cellular mechanism involved is poorly understood. Here, we found that the activated CXCR3 was co-localized with either neurons, microglia, and astrocytes in the spinal cord, or non-peptidergic-, peptidergic-, and A-type neurons in the dorsal root ganglion. The inoculation of Walker-256 mammary gland carcinoma cells into the rat's tibia induced a time-dependent phosphorylation of Akt and extracellular signal-regulated kinase (ERK1/2) in the spinal cord, and CXCR3 was necessary for the phosphorylation of Akt and ERK 1/2. Meanwhile, CXCR3 was co-localized with either pAkt or pERK1/2. Blockage of either Akt or ERK1/2 prevented or reversed the mechanical allodynia in BCP rats. Furthermore, there was cross-activation between PI3K/Akt and Raf/MEK/ERK pathway under the BCP condition. Our results demonstrated that the activation of spinal chemokine receptor CXCR3 mediated BCP through Akt and ERK 1/2 kinase, and also indicated a crosstalk between PI3K/Akt and Raf/MEK/ERK signaling pathways under the BCP condition.

  17. REDD1 enhances protein phosphatase 2A-mediated dephosphorylation of Akt to repress mTORC1 signaling

    PubMed Central

    Dennis, Michael D.; Coleman, Catherine S.; Berg, Arthur; Jefferson, Leonard S.; Kimball, Scot R.

    2014-01-01

    The protein kinase mTOR (mechanistic target of rapamycin) in complex 1 (mTORC1) promotes cell growth and proliferation in response to anabolic stimuli, including growth factors and nutrients. Growth factors activate mTORC1 by stimulating the kinase Akt, which phosphorylates and inhibits the tuberous sclerosis complex (TSC; which is comprised of TSC1, TSC2, and TBC1D7), thereby stimulating the mTORC1 activator Rheb. Here, we identified the mechanism through which REDD1 (regulated in DNA damage and development 1) represses the mTORC1 signaling pathway. We found that REDD1 promoted the protein phosphatase 2A (PP2A)-dependent dephosphorylation of Akt at Thr308 but not at Ser473. Consistent with previous studies showing that phosphorylation of Akt on Thr308, but not Ser473, is necessary for phosphorylation of TSC2, we observed a REDD1-dependent reduction in the phosphorylation of TSC2 and subsequently in the activity of Rheb. REDD1 and PP2A coimmunoprecipitated with Akt from wild-type but not REDD1-knockout mouse embryonic fibroblasts, suggesting that REDD1 may act as a targeting protein for the catalytic subunit of PP2A. Furthermore, binding to both Akt and PP2A was essential for REDD1 to repress signaling to mTORC1. Overall, the results demonstrate that REDD1 acts not just as a repressor of mTORC1, but also as a constant modulator of the phosphorylation of Akt in response to growth factors and nutrients. PMID:25056877

  18. MicroRNA-145 inhibits the activation of the mTOR signaling pathway to suppress the proliferation and invasion of invasive pituitary adenoma cells by targeting AKT3 in vivo and in vitro

    PubMed Central

    Zhou, Kai; Fan, Yan-Dong; Wu, Peng-Fei; Duysenbi, Serick; Feng, Zhao-Hai; Du, Guo-Jia; Zhang, Ting-Rong

    2017-01-01

    Purpose This study was designed to explore how miR-145 regulates the mTOR signaling pathway in invasive pituitary adenoma (IPA) by targeting AKT3. Methods A total of 71 cases of IPA tissues and 66 cases of non-IPA tissues were obtained in this study. In vitro, the IPA cells were assigned into blank control, empty plasmid, miR-145 mimic, miR-145 inhibitor, miR-145 mimic + rapamycin, miR-145 inhibitor + rapamycin and rapamycin groups. Quantitative real-time polymerase chain reaction (qRT-PCR) and Western blotting were performed to detect the protein expressions of PI3K, AKT3, mTOR mRNA and the mRNA expression of miR-145 both in vivo and in vitro. Additionally, the S6K and RPS6 mRNA and protein expressions as well as the relative phosphorylation levels were determined in vitro. MTT assay, flow cytometry and transwell assay were used to testify the cell proliferation, apoptosis and invasion ability, respectively. Results The IPA tissues exhibited significantly lower expression of miR-145 but higher PI3K, AKT3 and mTOR mRNA and protein expressions when compared with the non-IPA tissues. Compared with the blank control and empty plasmid groups, the miR-145 mimic group showed significantly decreased PI3K, AKT3, mTOR, S6K and RPS6 mRNA and protein expressions as well as phosphorylation levels; besides, the IPA cell proliferation, migration and invasion ability were strongly inhibited, accompanied with the increased number of apoptotic cells. In the miR-145 inhibitor group, the PI3K, AKT3, mTOR, S6K and RPS6 mRNA and protein expressions as well as the phosphorylation levels were significantly increased; cell proliferation, migration and invasion ability were remarkably elevated, accompanied with reduced apoptotic cell number. Conclusion The study demonstrates that miR-145 inhibits the mTOR signaling pathway to suppress the IPA cell proliferation and invasion and promotes its apoptosis by targeting AKT3. PMID:28352194

  19. Dissociation of ERK and Akt signaling in endothelial cell angiogenic responses to {beta}-amyloid

    SciTech Connect

    Magrane, Jordi; Christensen, Rial A.; Rosen, Kenneth M.; Veereshwarayya, Vimal; Querfurth, Henry W. . E-mail: hquerf01@granite.tufts.edu

    2006-04-15

    Cerebrovascular deposits of {beta}-amyloid (A{beta}) peptides are found in Alzheimer's disease and cerebral amyloid angiopathy with stroke or dementia. Dysregulations of angiogenesis, the blood-brain barrier and other critical endothelial cell (EC) functions have been implicated in aggravating chronic hypoperfusion in AD brain. We have used cultured ECs to model the effects of {beta}-amyloid on the activated phosphorylation states of multifunctional serine/threonine kinases since these are differentially involved in the survival, proliferation and migration aspects of angiogenesis. Serum-starved EC cultures containing amyloid-{beta} peptides underwent a 2- to 3-fold increase in nuclear pyknosis. Under growth conditions with sublethal doses of {beta}-amyloid, loss of cell membrane integrity and inhibition of cell proliferation were observed. By contrast, cell migration was the most sensitive to A{beta} since inhibition was significant already at 1 {mu}M (P = 0.01, migration vs. proliferation). In previous work, intracellular A{beta} accumulation was shown toxic to ECs and Akt function. Here, extracellular A{beta} peptides do not alter Akt activation, resulting instead in proportionate decreases in the phosphorylations of the MAPKs: ERK1/2 and p38 (starting at 1 {mu}M). This inhibitory action occurs proximal to MEK1/2 activation, possibly through interference with growth factor receptor coupling. Levels of phospho-JNK remained unchanged. Addition of PD98059, but not LY294002, resulted in a similar decrease in activated ERK1/2 levels and inhibition of EC migration. Transfection of ERK1/2 into A{beta}-poisoned ECs functionally rescued migration. The marked effect of extracellular A{beta} on the migration component of angiogenesis is associated with inhibition of MAPK signaling, while Akt-dependent cell survival appears more affected by cellular A{beta}.

  20. A novel signaling pathway associated with Lyn, PI 3-kinase and Akt supports the proliferation of myeloma cells

    SciTech Connect

    Iqbal, Mohd S.; Tsuyama, Naohiro; Obata, Masanori; Ishikawa, Hideaki

    2010-02-12

    Interleukin-6 (IL-6) is a growth factor for human myeloma cells. We have recently found that in myeloma cells the activation of both signal transducer and activator of transcription (STAT) 3 and extracellular signal-regulated kinase (ERK) 1/2 is not sufficient for the IL-6-induced proliferation, which further requires the activation of the src family kinases, such as Lyn. Here we showed that the Lyn-overexpressed myeloma cell lines had the higher proliferative rate with IL-6 and the enhanced activation of the phosphatidylinositol (PI) 3-kinase and Akt. The IL-6-induced phosphorylation of STAT3 and ERK1/2 was not up-regulated in the Lyn-overexpressed cells, indicating that the Lyn-PI 3-kinase-Akt pathway is independent of these pathways. The PI 3-kinase was co-precipitated with Lyn in the Lyn-overexpressed cells of which proliferation with IL-6 was abrogated by the specific inhibitors for PI 3-kinase or Akt, suggesting that the activation of the PI 3-kinase-Akt pathway associated with Lyn is indeed related to the concomitant augmentation of myeloma cell growth. Furthermore, the decreased expression of p53 and p21{sup Cip1} proteins was observed in the Lyn-overexpressed cells, implicating a possible downstream target of Akt. This study identifies a novel IL-6-mediated signaling pathway that certainly plays a role in the proliferation of myeloma cells and this novel mechanism of MM tumor cell growth associated with Lyn would eventually contribute to the development of MM treatment.

  1. High constitutive Akt2 activity in U937 promonocytes: effective reduction of Akt2 phosphorylation by the histamine H2-receptor and the β2-adrenergic receptor.

    PubMed

    Werner, Kristin; Neumann, Detlef; Seifert, Roland

    2016-01-01

    Histamine (HA) is approved for the treatment of acute myeloid leukemia (AML). Its antileukemic activity is related to histamine H2-receptor (H2R)-mediated inhibition of reactive oxygen species (ROS) production in myeloid cells facilitating survival of antineoplastic lymphocytes. The phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, which plays a crucial role in cell survival and proliferation, is constitutively activated in leukemic cells of most AML patients resulting in poor survival prognosis. In a proof-of-principle experiment using a human phosphorylated mitogen-activated protein kinase (MAPK) array, we found high phosphorylation levels of Akt2 in U937 promonocytes that was abrogated by HA or selective H2R agonists. The H2R and the β2-adrenergic receptor (β2AR) are Gs-protein-coupled receptors. Stimulation results in adenylyl cyclase activation followed by generation of the second messenger adenosine 3′,5′-cyclic monophosphate (cAMP). In our present study, we evaluated the pharmacological profile of the H2R and the β2AR regarding Akt2 phosphorylation at Ser474 via western blot analysis and ELISA and cAMP accumulation via HPLC-MS/MS in U937 promonocytes. H2R and β2AR agonists concentration-dependently decreased Akt2 phosphorylation at Ser474. Deviations of potencies and efficacies of agonists in Akt2 phosphorylation and cAMP accumulation assays indicated participation of cAMP-independent signaling in GPCR-induced reduction of Akt2 phosphorylation. Accordingly, our study supports the concept of functional selectivity of the H2R and the β2AR in U937 promonocytes. In summary, we extended the antileukemic mechanism of HA via H2R and revealed the potential of β2AR agonists, which are already approved in the treatment of bronchial asthma and chronic obstructive pulmonary disease, as antileukemic drugs.

  2. Physiological activation of Akt by PHLPP1 deletion protects against pathological hypertrophy

    PubMed Central

    Moc, Courtney; Taylor, Amy E.; Chesini, Gino P.; Zambrano, Cristina M.; Barlow, Melissa S.; Zhang, Xiaoxue; Gustafsson, Åsa B.; Purcell, Nicole H.

    2015-01-01

    Aims To examine the role of physiological Akt signalling in pathological hypertrophy through analysis of PHLPP1 (PH domain leucine-rich repeat protein phosphatase) knock-out (KO) mice. Methods and results To investigate the in vivo requirement for ‘physiological’ control of Akt activation in cardiac growth, we examined the effect of deleting the Akt phosphatase, PHLPP, on the induction of cardiac hypertrophy. Basal Akt phosphorylation increased nearly two-fold in the cardiomyocytes from PHLPP1 KO mice and physiological hypertrophy induced by swimming exercise was accentuated as assessed by increased heart size and myocyte cell area. In contrast, the development of pathophysiological hypertrophy induced by pressure overload and assessed by increases in heart size, myocyte cell area, and hypertrophic gene expression was attenuated. This attenuation coincided with decreased fibrosis and cell death in the KO mice. Cast moulding revealed increased capillary density basally in the KO hearts, which was further elevated relative to wild-type mouse hearts in response to pressure overload. In vitro studies with isolated myocytes in co-culture also demonstrated that PHLPP1 deletion in cardiomyocytes can enhance endothelial tube formation. Expression of the pro-angiogenic factor VEGF was also elevated basally and accentuated in response to transverse aortic constriction in hearts from KO mice. Conclusion Our data suggest that enhancing Akt activity by inhibiting its PHLPP1-mediated dephosphorylation promotes processes associated with physiological hypertrophy that may be beneficial in attenuating the development of pathological hypertrophy. PMID:25411382

  3. Coprinus comatus Cap Inhibits Adipocyte Differentiation via Regulation of PPARγ and Akt Signaling Pathway

    PubMed Central

    Jang, Sun-Hee; Kang, Suk Nam; Jeon, Beong-Sam; Ko, Yeoung-Gyu; Kim, Hong-Duck; Won, Chung-Kil; Kim, Gon-Sup; Cho, Jae-Hyeon

    2014-01-01

    This study assessed the effects of Coprinus comatus cap (CCC) on adipogenesis in 3T3-L1 adipocytes and the effects of CCC on the development of diet-induced obesity in rats. Here, we showed that the CCC has an inhibitory effect on the adipocyte differentiation of 3T3-L1 cells, resulting in a significant decrease in lipid accumulation through the downregulation of several adipocyte specific-transcription factors, including CCAAT/enhancer binding protein β, C/EBPδ, and peroxisome proliferator-activated receptor gamma (PPARγ). Moreover, treatment with CCC during adipocyte differentiation induced a significant down-regulation of PPARγ and adipogenic target genes, including adipocyte protein 2, lipoprotein lipase, and adiponectin. Interestingly, the CCC treatment of the 3T3-L1 adipocytes suppressed the insulin-stimulated Akt and GSK3β phosphorylation, and these effects were stronger in the presence of an inhibitor of Akt phosphorylation, LY294002, suggesting that CCC inhibited adipocyte differentiation through the down-regulation of Akt signaling. In the animal study, CCC administration significantly reduced the body weight and adipose tissue weight of rats fed a high fat diet (HFD) and attenuated lipid accumulation in the adipose tissues of the HFD-induced obese rats. The size of the adipocyte in the epididymal fat of the CCC fed rats was significantly smaller than in the HFD rats. CCC treatment significantly reduced the total cholesterol and triglyceride levels in the serum of HFD rats. These results strongly indicated that the CCC-mediated decrease in body weight was due to a reduction in adipose tissue mass. The expression level of PPARγ and phospho-Akt was significantly lower in the CCC-treated HFD rats than that in the HFD obesity rats. These results suggested that CCC inhibited adipocyte differentiation by the down-regulation of major transcription factor involved in the adipogenesis pathway including PPARγ through the regulation of the Akt pathway in 3T3

  4. Low Expression of CAPON in Glioma Contributes to Cell Proliferation via the Akt Signaling Pathway

    PubMed Central

    Gao, Shangfeng; Wang, Jie; Zhang, Tong; Liu, Guangping; Jin, Lei; Ji, Daofei; Wang, Peng; Meng, Qingming; Zhu, Yufu; Yu, Rutong

    2016-01-01

    CAPON is an adapter protein for nitric oxide synthase 1 (NOS1). CAPON has two isoforms in the human brain: CAPON-L (long form of CAPON) and CAPON-S (short form of CAPON). Recent studies have indicated the involvement of CAPON in tumorigenesis beyond its classical role in NOS1 activity regulation. In this study, we found that the protein levels of CAPON-S, but not than CAPON-L, were significantly decreased in glioma tissues. Therefore, we established lentivirus-mediated stable cell lines with CAPON-S overexpression or down-regulation, and investigated the role of CAPON-S in the proliferation of glioma cells by using CCK8, EdU, and flow cytometry assays. Overexpression of CAPON-S reduced the cell variability and the percentage of EdU-positive cells, and arrested the cells in the G1 phase in glioma cells. Silencing of CAPON by short-hairpin RNA showed the opposite effects. Furthermore, an intracellular signaling array revealed that overexpression of CAPON-S resulted in a remarkable reduction in the phosphorylation of Akt and S6 ribosomal protein in glioma cells, which was further confirmed by Western blot. These findings suggest that CAPON may function as a tumor suppressor in human brain glioma and that the inactivation of the Akt signaling pathway caused by CAPON-S overexpression may provide insight into the underlying mechanism of CAPON in glioma cell proliferation. PMID:27869735

  5. Akt signaling-associated metabolic effects of dietary gold nanoparticles in Drosophila

    NASA Astrophysics Data System (ADS)

    Wang, Bin; Chen, Nan; Wei, Yingliang; Li, Jiang; Sun, Li; Wu, Jiarui; Huang, Qing; Liu, Chang; Fan, Chunhai; Song, Haiyun

    2012-08-01

    Gold nanoparticles (AuNPs) are often used as vehicles to deliver drugs or biomolecules, due to their mild effect on cell survival and proliferation. However, little is known about their effect on cellular metabolism. Here we examine the in vivo effect of AuNPs on metabolism using Drosophila as a model. Drosophila and vertebrates possess similar basic metabolic functions, and a highly conserved PI3K/Akt/mTOR signaling pathway plays a central role in the regulation of energy metabolism in both organisms. We show that dietary AuNPs enter the fat body, a key metabolic tissue in Drosophila larvae. Significantly, larvae fed with AuNP show increased lipid levels without triggering stress responses. In addition, activities of the PI3K/Akt/mTOR signaling pathway and fatty acids synthesis are increased in these larvae. This study thus reveals a novel function of AuNPs in influencing animal metabolism and suggests its potential therapeutic applications for metabolic disorders.

  6. The Phosphatidylinositol 3-Kinase/Akt Pathway Regulates Transforming Growth Factor-β Signaling by Destabilizing Ski and Inducing Smad7*

    PubMed Central

    Band, Arja M.; Björklund, Mia; Laiho, Marikki

    2009-01-01

    Ski is an oncoprotein that negatively regulates transforming growth factor (TGF)-β signaling. It acts as a transcriptional co-repressor by binding to TGF-β signaling molecules, Smads. Efficient TGF-β signaling is facilitated by rapid proteasome-mediated degradation of Ski by TGF-β. Here we report that Ski is phosphorylated by Akt/PKB kinase. Akt phosphorylates Ski on a highly conserved Akt motif at threonine 458 both in vitro and in vivo. The phosphorylation of Ski at threonine 458 is induced by Akt pathway activators including insulin, insulin-like growth factor-1, and hepatocyte growth factor. The phosphorylation of Ski causes its destabilization and reduces Ski-mediated inhibition of expression of another negative regulator of TGF-β, Smad7. Induction of Smad7 levels leads to inactivation of TGF-β receptors and TGF-β signaling cascade, as indicated by reduced induction of TGF-β target p15. Therefore, Akt modulates TGF-β signaling by temporarily adjusting the levels of two TGF-β pathway negative regulators, Ski and Smad7. These novel findings demonstrate that Akt pathway activation directly impacts TGF-β pathway. PMID:19875456

  7. Novel pharmacodynamic biomarkers for MYCN protein and PI3K/AKT/mTOR pathway signaling in children with neuroblastoma.

    PubMed

    Smith, Jennifer R; Moreno, Lucas; Heaton, Simon P; Chesler, Louis; Pearson, Andrew D J; Garrett, Michelle D

    2016-04-01

    There is an urgent need for improved therapies for children with high-risk neuroblastoma where survival rates remain low. MYCN amplification is the most common genomic change associated with aggressive neuroblastoma and drugs targeting PI3K/AKT/mTOR, to activate MYCN oncoprotein degradation, are entering clinical evaluation. Our aim was to develop and validate pharmacodynamic (PD) biomarkers to evaluate both proof of mechanism and proof of concept for drugs that block PI3K/AKT/mTOR pathway activity in children with neuroblastoma. We have addressed the issue of limited access to tumor biopsies for quantitative detection of protein biomarkers by optimizing a three-color fluorescence activated cell sorting (FACS) method to purify CD45-/GD2+/CD56+ neuroblastoma cells from bone marrow. We then developed a novel quantitative measurement of MYCN protein in these isolated neuroblastoma cells, providing the potential to demonstrate proof of concept for drugs that inhibit PI3K/AKT/mTOR signaling in this disease. In addition we have established quantitative detection of three biomarkers for AKT pathway activity (phosphorylated and total AKT, GSK3β and P70S6K) in surrogate platelet-rich plasma (PRP) from pediatric patients. Together our new approach to neuroblastoma cell isolation for protein detection and suite of PD assays provides for the first time the opportunity for robust, quantitative measurement of protein-based PD biomarkers in this pediatric patient population. These will be ideal tools to support clinical evaluation of PI3K/AKT/mTOR pathway drugs and their ability to target MYCN oncoprotein in upcoming clinical trials in neuroblastoma.

  8. NF-κB and AKT signaling prevent DNA damage in transformed pre-B cells by suppressing RAG1/2 expression and activity.

    PubMed

    Ochodnicka-Mackovicova, Katarina; Bahjat, Mahnoush; Bloedjes, Timon A; Maas, Chiel; de Bruin, Alexander M; Bende, Richard J; van Noesel, Carel J M; Guikema, Jeroen E J

    2015-09-10

    In developing lymphocytes, expression and activity of the recombination activation gene protein 1 (RAG1) and RAG2 endonuclease complex is tightly regulated to ensure ordered recombination of the immunoglobulin genes and to avoid genomic instability. Aberrant RAG activity has been implicated in the generation of secondary genetic events in human B-cell acute lymphoblastic leukemias (B-ALLs), illustrating the oncogenic potential of the RAG complex. Several layers of regulation prevent collateral genomic DNA damage by restricting RAG activity to the G1 phase of the cell cycle. In this study, we show a novel pathway that suppresses RAG expression in cycling-transformed mouse pre-B cells and human pre-B B-ALL cells that involves the negative regulation of FOXO1 by nuclear factor κB (NF-κB). Inhibition of NF-κB in cycling pre-B cells resulted in upregulation of RAG expression and recombination activity, which provoked RAG-dependent DNA damage. In agreement, we observe a negative correlation between NF-κB activity and the expression of RAG1, RAG2, and TdT in B-ALL patients. Our data suggest that targeting NF-κB in B-ALL increases the risk of RAG-dependent genomic instability.

  9. Effect of eccentric exercise velocity on akt/mtor/p70(s6k) signaling in human skeletal muscle.

    PubMed

    Roschel, Hamilton; Ugrinowistch, Carlos; Barroso, Renato; Batista, Mauro A B; Souza, Eduardo O; Aoki, Marcelo S; Siqueira-Filho, Mario A; Zanuto, Ricardo; Carvalho, Carla R O; Neves, Manoel; Mello, Marco T; Tricoli, Valmor

    2011-04-01

    It has been suggested that muscle tension plays a major role in the activation of intracellular pathways for skeletal muscle hypertrophy via an increase in mechano growth factor (MGF) and other downstream targets. Eccentric exercise (EE) imposes a greater amount of tension on the active muscle. In particular, high-speed EE seems to exert an additional effect on muscle tension and, thus, on muscle hypertrophy. However, little is known about the effect of EE velocity on hypertrophy signaling. This study investigated the effect of acute EE-velocity manipulation on the Akt/mTORCI/p70(S6K) hypertrophy pathway. Twenty subjects were assigned to either a slow (20°·s(-1); ES) or fast EE (210°·s(-1); EF) group. Biopsies were taken from vastus lateralis at baseline (B), immediately after (T1), and 2 h after (T2) the completion of 5 sets of 8 repetitions of eccentric knee extensions. Akt, mTOR, and p70(S6K) total protein were similar between groups, and did not change postintervention. Further, Akt and p70(S6K) protein phosphorylation were higher at T2 than at B for ES and EF. MGF messenger RNA was similar between groups, and only significantly higher at T2 than at B in ES. The acute manipulation of EE velocity does not seem to differently influence intracellular hypertrophy signaling through the Akt/mTORCI/p70S6K pathway.

  10. Synthetic and endogenous cannabinoids protect retinal neurons from AMPA excitotoxicity in vivo, via activation of CB1 receptors: Involvement of PI3K/Akt and MEK/ERK signaling pathways.

    PubMed

    Kokona, Despina; Thermos, Kyriaki

    2015-07-01

    Cannabinoids have been suggested to protect retinal ganglion cells in different models of toxicity, but their effects on other retinal neurons are poorly known. We investigated the neuroprotective actions of the endocannabinoid N-arachidonoyl ethanolamine (Anandamide/AEA) and the synthetic cannabinoids R1-Methanandamide (MethAEA) and HU-210, in an in vivo retinal model of AMPA excitotoxicity, and the mechanisms involved in the neuroprotection. Sprague-Dawley rats were intravitreally injected with PBS or AMPA in the absence or presence of the cannabinoid agonists. Brain nitric oxide synthase (bNOS) and choline acetyltransferase (ChAT) immunoreactivity (IR), as well as TUNEL staining, assessed the AMPA-induced retinal amacrine cell loss and the dose-dependent neuroprotection afforded by cannabinoids. The CB1 receptor selective antagonist AM251 and the PI3K/Akt inhibitor wortmannin reversed the cannabinoid-induced neuroprotection, suggesting the involvement of CB1 receptors and the PI3K/Akt pathway in cannabinoids' actions. Experiments with the CB2 agonist JWH015 and [(3)H]CP55940 radioligand binding suggested that the CB2 receptor is not involved in the neuroprotection. AEA and HU-210 induced phosphorylation of Akt but only AEA induced phosphorylation of ERK1/2 kinases, as revealed by western blot analysis. To investigate the role of caspase-3 in the AMPA-induced cell death, the caspase-3 inhibitor Z-DEVD-FMK was co-injected with AMPA. Z-DEVD-FMK had no effect on AMPA excitotoxicity. Moreover, no difference was observed in the phosphorylation of SAPK/JNK kinases between PBS- and AMPA-treated retinas. These results suggest that endogenous and synthetic cannabinoids protect retinal amacrine neurons from AMPA excitotoxicity in vivo via a mechanism involving the CB1 receptors, and the PI3K/Akt and/or MEK/ERK1/2 signaling pathways.

  11. Dual inhibitory roles of geldanamycin on the c-Jun NH2-terminal kinase 3 signal pathway through suppressing the expression of mixed-lineage kinase 3 and attenuating the activation of apoptosis signal-regulating kinase 1 via facilitating the activation of Akt in ischemic brain injury.

    PubMed

    Wen, X-R; Li, C; Zong, Y-Y; Yu, C-Z; Xu, J; Han, D; Zhang, G-Y

    2008-10-15

    It is well documented that heat-shock protein (hsp90) plays an essential role in maintaining stability and activity of its clients. Recent studies have shown that geldanamycin (GA), an inhibitor of hsp90, could decrease the protein of mixed-lineage kinase (MLK) 3 and activate Akt; our previous research documented that MLK3 and Akt and subsequent c-Jun N-terminal kinase (JNK) were involved in neuronal cell death in ischemic brain injury. Here, we investigated whether GA could decrease the protein of MLK3 and activate Akt in rat four-vessel occlusion ischemic model. Our results showed that global cerebral ischemia followed by reperfusion could enhance the association of hsp90 with MLK3, the association of hsp90 with Src, and JNK3 activation. As a result, GA decreased the protein of MLK3 and down-regulated JNK activation. On the other hand, Src kinase was activated and phosphorylated Cbl, which then recruited the p85 subunit of phosphatidylinositol 3-kinase (PI-3K), resulting in PI-3K activation, and as a consequence increased Akt activation, which inhibited ASK1 activation and down-regulated JNK3 activation. In summary, our results indicated that GA showed a dual inhibitory role on JNK3 activation and exerted strong neuroprotection in vivo and in vitro, which provides a new possible approach for stroke therapy.

  12. Sea urchin akt activity is Runx-dependent and required for post-cleavage stage cell division.

    PubMed

    Robertson, Anthony J; Coluccio, Alison; Jensen, Sarah; Rydlizky, Katarina; Coffman, James A

    2013-05-15

    In animal development following the initial cleavage stage of embryogenesis, the cell cycle becomes dependent on intercellular signaling and controlled by the genomically encoded ontogenetic program. Runx transcription factors are critical regulators of metazoan developmental signaling, and we have shown that the sea urchin Runx gene runt-1, which is globally expressed during early embryogenesis, functions in support of blastula stage cell proliferation and expression of the mitogenic genes pkc1, cyclinD, and several wnts. To obtain a more comprehensive list of early runt-1 regulatory targets, we screened a Strongylocentrotus purpuratus microarray to identify genes mis-expressed in mid-blastula stage runt-1 morphants. This analysis showed that loss of Runx function perturbs the expression of multiple genes involved in cell division, including the pro-growth and survival kinase Akt (PKB), which is significantly underexpressed in runt-1 morphants. Further genomic analysis revealed that Akt is encoded by two genes in the S. purpuratus genome, akt-1 and akt-2, both of which contain numerous canonical Runx target sequences. The transcripts of both genes accumulate several fold during blastula stage, contingent on runt-1 expression. Inhibiting Akt expression or activity causes blastula stage cell cycle arrest, whereas overexpression of akt-1 mRNA rescues cell proliferation in runt-1 morphants. These results indicate that post-cleavage stage cell division requires Runx-dependent expression of akt.

  13. Targeted disruption of TC-PTP in the proliferative compartment augments STAT3 and AKT signaling and skin tumor development

    PubMed Central

    Lee, Hyunseung; Kim, Mihwa; Baek, Minwoo; Morales, Liza D.; Jang, Ik-Soon; Slaga, Thomas J.; DiGiovanni, John; Kim, Dae Joon

    2017-01-01

    Tyrosine phosphorylation is a vital mechanism that contributes to skin carcinogenesis. It is regulated by the counter-activities of protein tyrosine kinases (PTKs) and protein tyrosine phosphatases (PTPs). Here, we report the critical role of T-cell protein tyrosine phosphatase (TC-PTP), encoded by Ptpn2, in chemically-induced skin carcinogenesis via the negative regulation of STAT3 and AKT signaling. Using epidermal specific TC-PTP knockout (K14Cre.Ptpn2fl/fl) mice, we demonstrate loss of TC-PTP led to a desensitization to tumor initiator 7,12-dimethylbenz[a]anthracene (DMBA)-induced apoptosis both in vivo epidermis and in vitro keratinocytes. TC-PTP deficiency also resulted in a significant increase in epidermal thickness and hyperproliferation following exposure to the tumor promoter, 12-O-tetradecanoylphorbol-13-acetate (TPA). Western blot analysis showed that both phosphorylated STAT3 and phosphorylated AKT expressions were significantly increased in epidermis of TC-PTP-deficient mice compared to control mice following TPA treatment. Inhibition of STAT3 or AKT reversed the effects of TC-PTP deficiency on apoptosis and proliferation. Finally, TC-PTP knockout mice showed a shortened latency of tumorigenesis and significantly increased numbers of tumors during two-stage skin carcinogenesis. Our findings reveal that TC-PTP has potential as a novel target for the prevention of skin cancer through its role in the regulation of STAT3 and AKT signaling. PMID:28322331

  14. Liver Clock Protein BMAL1 Promotes de Novo Lipogenesis through Insulin-mTORC2-AKT Signaling*

    PubMed Central

    Zhang, Deqiang; Tong, Xin; Arthurs, Blake; Guha, Anirvan; Rui, Liangyou; Kamath, Avani; Inoki, Ken; Yin, Lei

    2014-01-01

    The clock protein BMAL1 (brain and muscle Arnt-like protein 1) participates in circadian regulation of lipid metabolism, but its contribution to insulin AKT-regulated hepatic lipid synthesis is unclear. Here we used both Bmal1−/− and acute liver-specific Bmal1-depleted mice to study the role of BMAL1 in refeeding-induced de novo lipogenesis in the liver. Both global deficiency and acute hepatic depletion of Bmal1 reduced lipogenic gene expression in the liver upon refeeding. Conversely, Bmal1 overexpression in mouse liver by adenovirus was sufficient to elevate the levels of mRNA of lipogenic enzymes. Bmal1−/− primary mouse hepatocytes displayed decreased levels of de novo lipogenesis and lipogenic enzymes, supporting the notion that BMAL1 regulates lipid synthesis in hepatocytes in a cell-autonomous manner. Both refed mouse liver and insulin-treated primary mouse hepatocytes showed impaired AKT activation in the case of either Bmal1 deficiency or Bmal1 depletion by adenoviral shRNA. Restoring AKT activity by a constitutively active mutant of AKT nearly normalized de novo lipogenesis in Bmal1−/− hepatocytes. Finally, Bmal1 deficiency or knockdown decreased the protein abundance of RICTOR, the key component of the mTORC2 complex, without affecting the gene expression of key factors of insulin signaling. Thus, our study uncovered a novel metabolic function of hepatic BMAL1 that promotes de novo lipogenesis via the insulin-mTORC2-AKT signaling during refeeding. PMID:25063808

  15. Repression of BIM mediates survival signaling by MYC and AKT in high-risk T-cell acute lymphoblastic leukemia.

    PubMed

    Reynolds, C; Roderick, J E; LaBelle, J L; Bird, G; Mathieu, R; Bodaar, K; Colon, D; Pyati, U; Stevenson, K E; Qi, J; Harris, M; Silverman, L B; Sallan, S E; Bradner, J E; Neuberg, D S; Look, A T; Walensky, L D; Kelliher, M A; Gutierrez, A

    2014-09-01

    Treatment resistance in T-cell acute lymphoblastic leukemia (T-ALL) is associated with phosphatase and tensin homolog (PTEN) deletions and resultant phosphatidylinositol 3'-kinase (PI3K)-AKT pathway activation, as well as MYC overexpression, and these pathways repress mitochondrial apoptosis in established T-lymphoblasts through poorly defined mechanisms. Normal T-cell progenitors are hypersensitive to mitochondrial apoptosis, a phenotype that is dependent on the expression of proapoptotic BIM. In a conditional zebrafish model, MYC downregulation induced BIM expression in T-lymphoblasts, an effect that was blunted by expression of constitutively active AKT. In human T-ALL cell lines and treatment-resistant patient samples, treatment with MYC or PI3K-AKT pathway inhibitors each induced BIM upregulation and apoptosis, indicating that BIM is repressed downstream of MYC and PI3K-AKT in high-risk T-ALL. Restoring BIM function in human T-ALL cells using a stapled peptide mimetic of the BIM BH3 domain had therapeutic activity, indicating that BIM repression is required for T-ALL viability. In the zebrafish model, where MYC downregulation induces T-ALL regression via mitochondrial apoptosis, T-ALL persisted despite MYC downregulation in 10% of bim wild-type zebrafish, 18% of bim heterozygotes and in 33% of bim homozygous mutants (P=0.017). We conclude that downregulation of BIM represents a key survival signal downstream of oncogenic MYC and PI3K-AKT signaling in treatment-resistant T-ALL.

  16. Sustained Activation of Akt Elicits Mitochondrial Dysfunction to Block Plasmodium falciparum Infection in the Mosquito Host

    PubMed Central

    Drexler, Anna L.; Antonova-Koch, Yevgeniya; Sakaguchi, Danielle; Napoli, Eleonora; Wong, Sarah; Price, Mark S.; Eigenheer, Richard; Phinney, Brett S.; Pakpour, Nazzy; Pietri, Jose E.; Cheung, Kong; Georgis, Martha; Riehle, Michael

    2013-01-01

    The overexpression of activated, myristoylated Akt in the midgut of female transgenic Anopheles stephensi results in resistance to infection with the human malaria parasite Plasmodium falciparum but also decreased lifespan. In the present study, the understanding of mitochondria-dependent midgut homeostasis has been expanded to explain this apparent paradox in an insect of major medical importance. Given that Akt signaling is essential for cell growth and survival, we hypothesized that sustained Akt activation in the mosquito midgut would alter the balance of critical pathways that control mitochondrial dynamics to enhance parasite killing at some cost to survivorship. Toxic reactive oxygen and nitrogen species (RNOS) rise to high levels in the midgut after blood feeding, due to a combination of high NO production and a decline in FOXO-dependent antioxidants. Despite an apparent increase in mitochondrial biogenesis in young females (3 d), energy deficiencies were apparent as decreased oxidative phosphorylation and increased [AMP]/[ATP] ratios. In addition, mitochondrial mass was lower and accompanied by the presence of stalled autophagosomes in the posterior midgut, a critical site for blood digestion and stem cell-mediated epithelial maintenance and repair, and by functional degradation of the epithelial barrier. By 18 d, the age at which An. stephensi would transmit P. falciparum to human hosts, mitochondrial dysfunction coupled to Akt-mediated repression of autophagy/mitophagy was more evident and midgut epithelial structure was markedly compromised. Inhibition of RNOS by co-feeding of the nitric-oxide synthase inhibitor L-NAME at infection abrogated Akt-dependent killing of P. falciparum that begins within 18 h of infection in 3–5 d old mosquitoes. Hence, Akt-induced changes in mitochondrial dynamics perturb midgut homeostasis to enhance parasite resistance and decrease mosquito infective lifespan. Further, quality control of mitochondrial function in the

  17. Resveratrol rescues hyperglycemia-induced endothelial dysfunction via activation of Akt

    PubMed Central

    Li, Jin-yi; Huang, Wei-qiang; Tu, Rong-hui; Zhong, Guo-qiang; Luo, Bei-bei; He, Yan

    2017-01-01

    Resveratrol (RSV), a phytoalexin, has shown to prevent endothelial dysfunction and reduce diabetic vascular complications and the risk of cardiovascular diseases. The aim of this study was to investigate the signaling mechanisms underlying the protecting effects of RSV against endothelial dysfunction during hyperglycemia in vitro and in vivo. Human umbilical vein endothelial cells (HUVECs) were treated with RSV, and then exposed to high glucose (HG, 30 mmol/L). Akt-Ser473 phosphorylation, eNOS-Ser1177 phosphorylation, and PTEN protein levels in the cells were detected using Western blot. For in vivo studies, WT and Akt−/− mice were fed a normal diet containing RSV (400 mg·kg−1·d−1) for 2 weeks, then followed by injection of STZ to induce hyperglycemia (300 mg/dL). Endothelial function was evaluated using aortic rings by assessing ACh-induced vasorelaxation. RSV (5–20 μmol/L) dose-dependently increased Akt-Ser473 phosphorylation, accompanied by increased eNOS-Ser1177 phosphorylation in HUVECs; these effects were more prominent under HG stimulation. Transfection with Akt siRNA abolished RSV-enhanced eNOS phosphorylation and NO release. Furthermore, RSV (5–20 μmol/L) dose-dependently decreased the levels of PTEN, which was significantly increased under HG stimulation, and PTEN overexpression abolished RSV-stimulated Akt phosphorylation in HG-treated HUVECs. Moreover, RSV dramatically increased 26S proteasome activity, which induced degradation of PTEN. In in vivo studies, pretreatment with RSV significantly increased Akt and eNOS phosphorylation in aortic tissues and ACh-induced vasorelaxation, and improved diabetes-induced endothelial dysfunction in wild-type mice but not in Akt−/− mice. RSV attenuates endothelial function during hyperglycemia via activating proteasome-dependent degradation of PTEN, which increases Akt phosphorylation, and consequentially upregulation of eNOS-derived NO production. PMID:27941804

  18. Acarbose Accelerates Wound Healing via Akt/eNOS Signaling in db/db Mice

    PubMed Central

    Yu, Jiawen; Sun, Yuannan; Ren, Guofei; Zhu, Jianjun

    2017-01-01

    Refractory wound is a dreaded complication of diabetes and is highly correlated with EPC dysfunction caused by hyperglycemia. Acarbose is a widely used oral glucose-lowering drug exclusively for T2DM. Previous studies have suggested the beneficial effect of acarbose on improving endothelial dysfunction in patients with T2DM. However, no data have been reported on the beneficial efficacy of acarbose in wound healing impairment caused by diabetes. We herein investigated whether acarbose could improve wound healing in T2DM db/db mice and the possible mechanisms involved. Acarbose hastened wound healing and enhanced angiogenesis, accompanied by increased circulating EPC number in db/db mice. In vitro, a reversed BM-EPC dysfunction was observed after the administration of acarbose in db/db mice, as reflected by tube formation assay. In addition, a significantly increased NO production was also witnessed in BM-EPCs from acarbose treated db/db mice, with decreased O2 levels. Akt inhibitor could abolish the beneficial effect of acarbose on high glucose induced EPC dysfunction in vitro, accompanied by reduced eNOS activation. Acarbose displayed potential effect in promoting wound healing and improving angiogenesis in T2DM mice, which was possibly related to the Akt/eNOS signaling pathway. PMID:28373902

  19. Combined inhibition of p38 and Akt signaling pathways abrogates cyclosporine A-mediated pathogenesis of aggressive skin SCCs

    SciTech Connect

    Arumugam, Aadithya; Walsh, Stephanie B.; Xu, Jianmin; Afaq, Farrukh; Elmets, Craig A.; Athar, Mohammad

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer p38 and Akt are the crucial molecular targets in the pathogenesis of SCCs in OTRs. Black-Right-Pointing-Pointer Combined inhibition of these targets diminished tumor growth by 90%. Black-Right-Pointing-Pointer Inhibition of these targets act through downregulating mTOR signaling pathway. -- Abstract: Non-melanoma skin cancers (NMSCs) are the most common neoplasm in organ transplant recipients (OTRs). These cancers are more invasive and metastatic as compared to those developed in normal cohorts. Previously, we have shown that immunosuppressive drug, cyclosporine A (CsA) directly alters tumor phenotype of cutaneous squamous cell carcinomas (SCCs) by activating TGF-{beta} and TAK1/TAB1 signaling pathways. Here, we identified novel molecular targets for the therapeutic intervention of these SCCs. We observed that combined blockade of Akt and p38 kinases-dependent signaling pathways in CsA-promoted human epidermoid carcinoma A431 xenograft tumors abrogated their growth by more than 90%. This diminution in tumor growth was accompanied by a significant decrease in proliferation and an increase in apoptosis. The residual tumors following the combined treatment with Akt inhibitor triciribine and p38 inhibitors SB-203580 showed significantly diminished expression of phosphorylated Akt and p38 and these tumors were less invasive and highly differentiated. Diminished tumor invasiveness was associated with the reduced epithelial-mesenchymal transition as ascertained by the enhanced E-cadherin and reduced vimentin and N-cadherin expression. Consistently, these tumors also manifested reduced MMP-2/9. The decreased p-Akt expression was accompanied by a significant reduction in p-mTOR. These data provide first important combinatorial pharmacological approach to block the pathogenesis of CsA-induced highly aggressive cutaneous neoplasm in OTRs.

  20. Effects of extracellular acid stimulation on rat vascular smooth muscle cell in Gas6/Axl or PI3K/Akt signaling pathway.

    PubMed

    Cui, Liwen; Bai, Yaling; Zhang, Junxia; Zhang, Shenglei; Xu, Jinsheng

    Recent studies have indicated that extracellular acid stimulation inhibited the calcification of vascular smooth muscle cells (VSMCs). Cell apoptosis played an important role in the occurrence and development of vascular calcification. We further explored the effects of Gas6/Axl or PI3K/Akt signaling pathway on the inhibition of rat VSMCs calcification in response to extracellular acid stimulation. Our study demonstrated that a high concentration of phosphorus induced apoptosis and calcification of VSMCs, decreased expression of Axl, and reduced phosphorylation of Akt. Stimulation of extracellular acid counteracted the effects as above by increasing the expression of Axl and Akt phosphorylation and decreasing the expression of activated Caspase3, which thereby decreased cell apoptosis and calcification. Moreover, the effects can be attenuated by PI3K inhibitor. Our study proved that extracellular acid stimulation played a vital role in the inhibition of rat VSMCs calcification and apoptosis in Gas6/Axl or PI3K/Akt signaling pathway.

  1. Fibroblast growth factor 4-induced migration of porcine trophectoderm cells is mediated via the AKT cell signaling pathway.

    PubMed

    Jeong, Wooyoung; Lee, Jieun; Bazer, Fuller W; Song, Gwonhwa; Kim, Jinyoung

    2016-01-05

    During early pregnancy, a well-coordinated communication network between the conceptus and maternal uterus is especially crucial in pigs in which there is a protracted pre-attachment phase prior to implantation. This network is regulated by an astonishing number of molecules such as growth factors. Fibroblast growth factor 4 (FGF4) is a multipotent growth factor that elicits diverse biological actions on various types of cells and tissues. In pigs, FGF4 and its receptors are expressed in the uterine endometrium and conceptus during early pregnancy, but less is known about the FGF4-mediated regulation of conceptus growth during peri-implantation period of pregnancy. Therefore, the aims of the present study were to investigate: 1) expression of endometrial FGF4 mRNA during early pregnancy; 2) up-regulation of FGF receptor expression in porcine trophectoderm (pTr) cells in response to FGF4; and 3) FGF-induced intracellular signaling and cellular activities in pTr cells. In vitro cultured pTr cells incubated with different concentrations of recombinant FGF4 (0-50 ng/ml) responded with a dose-dependent increase in AKT phosphorylation of 2.9-fold at 20 ng/ml FGF4. Within 30 min after treatment with 20 ng/ml FGF4, the abundances of p-AKT, p-P90RSK and p-RPS6 proteins increased 2.1-, 5.2- and 3.2-fold, respectively, and then returned to basal levels by 120 min. To ensure that the stimulatory effect of FGF4 on AKT signaling was p-AKT-dependent, pTr cells were pre-incubated with an AKT inhibitor (LY294002) for 1 h prior to FGF4 treatment. 20 μM of LY294002 decreased FGF4-induced p-AKT, p-P90RSK and p-RPS6 proteins. Immunofluorescence analyses revealed that p-RPS6 proteins were abundant within the cytoplasm of FGF4-treated cells, but present at basal levels in the presence of LY294002. Furthermore, FGF4 increased migration of pTr cells and LY294002 significantly reduced this effect. Results of the present study suggest that activation of the FGF receptor(s) on trophectoderm

  2. Effect of lithium on ventricular remodelling in infarcted rats via the Akt/mTOR signalling pathways.

    PubMed

    Lee, Tsung-Ming; Lin, Shinn-Zong; Chang, Nen-Chung

    2017-04-28

    Activation of phosphoinositide 3-kinase (PI3K)/Akt signalling is the molecular pathway driving physiological hypertrophy. As lithium, a PI3K agonist, is highly toxic at regular doses, we assessed the effect of lithium at a lower dose on ventricular hypertrophy after myocardial infarction (MI). Male Wistar rats after induction of MI were randomized to either vehicle or lithium (1 mmol/kg per day) for 4 weeks. The dose of lithium led to a mean serum level of 0.39 mM, substantially lower than the therapeutic concentrations (0.8-1.2 mM). Infarction in the vehicle was characterized by pathological hypertrophy in the remote zone; histologically, by increased cardiomyocyte sizes, interstitial fibrosis and left ventricular dilatation; functionally, by impaired cardiac contractility; and molecularly, by an increase of p-extracellular-signal-regulated kinase (ERK) levels, nuclear factor of activated T cells (NFAT) activity, GATA4 expression and foetal gene expressions. Lithium administration mitigated pathological remodelling. Furthermore, lithium caused increased phosphorylation of eukaryotic initiation factor 4E binding protein 1 (p-4E-BP1), the downstream target of mammalian target of rapamycin (mTOR). Blockade of the Akt and mTOR signalling pathway with deguelin and rapamycin resulted in markedly diminished levels of p-4E-BP1, but not ERK. The present study demonstrated that chronic lithium treatment at low doses mitigates pathological hypertrophy through an Akt/mTOR dependent pathway.

  3. [Signaling pathways mTOR and AKT in epilepsy].

    PubMed

    Romero-Leguizamon, C R; Ramirez-Latorre, J A; Mora-Munoz, L; Guerrero-Naranjo, A

    2016-07-01

    Introduccion. La via de señalizacion AKT/mTOR es un eje central en la regulacion celular, especialmente en las enfermedades neurologicas. En la epilepsia, se ha evidenciado su alteracion dentro de su proceso fisiopatologico. Sin embargo, aun no se han descrito todos los mecanismos de estas rutas de señalizacion, las cuales podrian abrir la puerta hacia nuevas investigaciones y estrategias terapeuticas, que finalmente permitan desarrollar tratamientos efectivos en enfermedades neurologicas como la epilepsia. Objetivo. Revisar las asociaciones existentes entre las rutas de señalizacion intracelular de mTOR y AKT en la fisiopatologia de la epilepsia. Desarrollo. La epilepsia es una enfermedad neurologica con un alto impacto epidemiologico en el mundo, por lo cual es de sumo interes la investigacion de los componentes fisiopatologicos que puedan generar nuevos tratamientos farmacologicos. En esta busqueda se han involucrado diferentes rutas de señalizacion intracelular en neuronas, como determinantes epileptogenos. Los avances en esta materia han permitido incluso la implementacion de nuevas estrategias terapeuticas exitosas y que abren el camino hacia nuevas investigaciones. Conclusiones. Mejorar los conocimientos respecto al papel fisiopatologico de la via de señalizacion mTOR/AKT en la epilepsia permite plantear nuevas investigaciones que ofrezcan nuevas alternativas terapeuticas para el tratamiento de la enfermedad. El uso de inhibidores de mTOR ha surgido en los ultimos años como una alternativa eficaz en el tratamiento de algunos tipos de epilepsias, pero es evidente la necesidad de seguir en la busqueda de nuevas terapias farmacologicas involucradas en estas vias de señalizacion.

  4. JNK/PI3K/Akt signaling pathway is involved in myocardial ischemia/reperfusion injury in diabetic rats: effects of salvianolic acid A intervention.

    PubMed

    Chen, Qiuping; Xu, Tongda; Li, Dongye; Pan, Defeng; Wu, Pei; Luo, Yuanyuan; Ma, Yanfeng; Liu, Yang

    2016-01-01

    Recent studies have demonstrated that diabetes impairs the phosphatidylinositol 3-kinase/Akt (PI3K/Akt) pathway, while insulin resistance syndrome has been associated with alterations of this pathway in diabetic rats after ischemia/reperfusion (I/R), and activation of C-jun N-terminal kinase (JNK) is involved. The present study was designed to investigate whether inhibiting JNK activity would partially restore the PI3K/Akt signaling pathway and protect against myocardial I/R injury in diabetic rats, and to explore the effect of intervention with salvianolic acid A (Sal A). The inhibitor of JNK (SP600125) and Sal A were used in type 2 diabetic (T2D) rats, outcome measures included heart hemodynamic data, myocardial infarct size, the release of lactate dehydrogenase (LDH), SERCA2a activity, cardiomyocyte apotosis, expression levels of Bcl-2, Bax and cleaved caspase-3, and the phosphorylation status of Akt and JNK. The p-Akt levels were increased after myocardial I/R in non-diabetic rats, while there was no change in diabetic rats. Pretreatment with the SP600125 and Sal A decreased the p-JNK levels and increased the p-Akt levels in diabetic rats with I/R, and heart hemodynamic data improved, infarct size and LDH release decreased, SERCA2a activity increased, Bax and cleaved caspase-3 expression levels decreased, and the expression of Bcl-2 and the Bcl-2/Bax ratio increased. Our results suggest that the JNK/PI3K/Akt signaling pathway is involved in myocardial I/R injury in diabetic rats and Sal A exerts an anti-apoptotic effect and improves cardiac function following I/R injury through the JNK/PI3K/Akt signaling pathway in this model.

  5. JNK/PI3K/Akt signaling pathway is involved in myocardial ischemia/reperfusion injury in diabetic rats: effects of salvianolic acid A intervention

    PubMed Central

    Chen, Qiuping; Xu, Tongda; Li, Dongye; Pan, Defeng; Wu, Pei; Luo, Yuanyuan; Ma, Yanfeng; Liu, Yang

    2016-01-01

    Recent studies have demonstrated that diabetes impairs the phosphatidylinositol 3-kinase/Akt (PI3K/Akt) pathway, while insulin resistance syndrome has been associated with alterations of this pathway in diabetic rats after ischemia/reperfusion (I/R), and activation of C-jun N-terminal kinase (JNK) is involved. The present study was designed to investigate whether inhibiting JNK activity would partially restore the PI3K/Akt signaling pathway and protect against myocardial I/R injury in diabetic rats, and to explore the effect of intervention with salvianolic acid A (Sal A). The inhibitor of JNK (SP600125) and Sal A were used in type 2 diabetic (T2D) rats, outcome measures included heart hemodynamic data, myocardial infarct size, the release of lactate dehydrogenase (LDH), SERCA2a activity, cardiomyocyte apotosis, expression levels of Bcl-2, Bax and cleaved caspase-3, and the phosphorylation status of Akt and JNK. The p-Akt levels were increased after myocardial I/R in non-diabetic rats, while there was no change in diabetic rats. Pretreatment with the SP600125 and Sal A decreased the p-JNK levels and increased the p-Akt levels in diabetic rats with I/R, and heart hemodynamic data improved, infarct size and LDH release decreased, SERCA2a activity increased, Bax and cleaved caspase-3 expression levels decreased, and the expression of Bcl-2 and the Bcl-2/Bax ratio increased. Our results suggest that the JNK/PI3K/Akt signaling pathway is involved in myocardial I/R injury in diabetic rats and Sal A exerts an anti-apoptotic effect and improves cardiac function following I/R injury through the JNK/PI3K/Akt signaling pathway in this model. PMID:27398138

  6. Contribution of insulin and Akt1 signaling to endothelial nitric oxide synthase in the regulation of endothelial function and blood pressure.

    PubMed

    Symons, J David; McMillin, Shawna L; Riehle, Christian; Tanner, Jason; Palionyte, Milda; Hillas, Elaine; Jones, Deborah; Cooksey, Robert C; Birnbaum, Morris J; McClain, Donald A; Zhang, Quan-Jiang; Gale, Derrick; Wilson, Lloyd J; Abel, E Dale

    2009-05-08

    Impaired insulin signaling via phosphatidylinositol 3-kinase/Akt to endothelial nitric oxide synthase (eNOS) in the vasculature has been postulated to lead to arterial dysfunction and hypertension in obesity and other insulin resistant states. To investigate this, we compared insulin signaling in the vasculature, endothelial function, and systemic blood pressure in mice fed a high-fat (HF) diet to mice with genetic ablation of insulin receptors in all vascular tissues (TTr-IR(-/-)) or mice with genetic ablation of Akt1 (Akt1-/-). HF mice developed obesity, impaired glucose tolerance, and elevated free fatty acids that was associated with endothelial dysfunction and hypertension. Basal and insulin-mediated phosphorylation of extracellular signal-regulated kinase 1/2 and Akt in the vasculature was preserved, but basal and insulin-stimulated eNOS phosphorylation was abolished in vessels from HF versus lean mice. In contrast, basal vascular eNOS phosphorylation, endothelial function, and blood pressure were normal despite absent insulin-mediated eNOS phosphorylation in TTr-IR(-/-) mice and absent insulin-mediated eNOS phosphorylation via Akt1 in Akt1-/- mice. In cultured endothelial cells, 6 hours of incubation with palmitate attenuated basal and insulin-stimulated eNOS phosphorylation and NO production despite normal activation of extracellular signal-regulated kinase 1/2 and Akt. Moreover, incubation of isolated arteries with palmitate impaired endothelium-dependent but not vascular smooth muscle function. Collectively, these results indicate that lower arterial eNOS phosphorylation, hypertension, and vascular dysfunction following HF feeding do not result from defective upstream signaling via Akt, but from free fatty acid-mediated impairment of eNOS phosphorylation.

  7. STC2 promotes head and neck squamous cell carcinoma metastasis through modulating the PI3K/AKT/Snail signaling

    PubMed Central

    Yang, Shuwen; Ji, Qinghai; Chang, Bin; Wang, Yan; Zhu, Yongxue; Li, Duanshu; Huang, Caiping; Wang, Yulong; Sun, Guohua; Zhang, Ling; Guan, Qing; Xiang, Jun; Wei, Wenjun; Lu, Zhongwu; Liao, Tian; Meng, Jiao; Wang, Ziliang; Ma, Ben; Zhou, Li; Wang, Yu; Yang, Gong

    2017-01-01

    The mammalian peptide hormone stanniocalcin 2 (STC2) plays an oncogenic role in many human cancers. However, the exact function of STC2 in human head and neck squamous cell carcinoma (HNSCC) is unclear. We aimed to examine the function and clinical significance of STC2 in HNSCC. Using in vitro and in vivo assays, we show that overexpression of STC2 suppressed cell apoptosis, promoted cell proliferation, migration, invasion, and cell cycle arrest at the G1/S transition. By contrast, silencing of STC2 inhibited these activities. We further show that STC2 upregulated the phosphorylation of AKT and enhanced HNSCC metastasis via Snail-mediated increase of vimentin and decrease of E-cadherin. These responses were blocked by silencing of STC2/Snail expression or inhibition of pAKT activity. Furthermore, clinical data indicate that high STC2 expression was associated with high levels of pAKT and Snail in tumor samples from HNSCC patients with regional lymph node metastasis (P < 0.01). Thus, we conclude that STC2 controls HNSCC metastasis via the PI3K/AKT/Snail signaling axis and that targeted therapy against STC2 may be a novel strategy to effectively treat patients with metastatic HNSCC. PMID:27863406

  8. 125I Seeds Radiation Induces Paraptosis-Like Cell Death via PI3K/AKT Signaling Pathway in HCT116 Cells

    PubMed Central

    Hu, Lelin; Wang, Hao; Zhao, Yong

    2016-01-01

    125I seeds brachytherapy implantation has been extensively performed in unresectable and rerecurrent rectal carcinoma. Many studies on the cancer-killing activity of 125I seeds radiation mainly focused on its ability to trigger apoptosis, which is the most well-known and dominant type of cell death induced by radiation. However our results showed some unique morphological features such as cell swelling, cytoplasmic vacuolation, and plasma membrane integrity, which is obviously different to apoptosis. In this study, clonogenic proliferation was carried out to assay survival fraction. Transmission electron microscopy was used to analyze ultrastructural and evaluate morphologic feature of HCT116 cells after exposure to 125I seeds radiation. Immunofluorescence analysis was used to detect the origin of cytoplasmic vacuoles. Flow cytometry analysis was employed to detect the size and granularity of HCT116 cells. Western blot was performed to measure the protein level of AIP1, caspase-3, AKT, p-Akt (Thr308), p-Akt (Ser473), and β-actin. We found that 125I seeds radiation activated PI3K/AKT signaling pathway and could trigger paraptosis-like cell death. Moreover, inhibitor of PI3K/AKT signaling pathway could inhibit paraptosis-like cell death induced by 125I seeds radiation. Our data suggest that 125I seeds radiation can induce paraptosis-like cell death via PI3K/AKT signaling pathway. PMID:28078301

  9. BRAIN OVERGROWTH IN DISORDERS OF RTK-PI3K-AKT SIGNALING: A MOSAIC OF MALFORMATIONS

    PubMed Central

    Hevner, Robert F.

    2014-01-01

    Disorders of brain overgrowth are significant causes of intractable epilepsy, intellectual disability, autism, and other complex neurological problems. The pathology of these disorders is sometimes striking and characteristic, as in hemimegalencephaly, but can also be subtle, as in autism. Recent genetic studies have shown that many diverse forms of brain overgrowth are caused by de novo mutations that increase activity in the receptor tyrosine kinase (RTK)–phosphatidylinositol-3-kinase (PI3K)–AKT signaling pathway, a key mediator of signaling by growth factors in the developing brain, such as fibroblast growth factors. In cases where mutations arise in postzygotic embryos, brain regions exhibit mosaic pathology that reflects the distribution of mutant cells, ranging from focal cortical dysplasia to lobar or hemispheric overgrowth. In turn, the histopathology of these disorders is also remarkably varied. The common underlying mechanisms of RTK-PI3K-AKT overactivation suggest new possibilities for drugs that inhibit this pathway. PMID:25432429

  10. PI3K/PTEN/Akt and TSC/mTOR signaling pathways, ovarian dysfunction, and infertility: an update.

    PubMed

    Makker, Annu; Goel, Madhu Mati; Mahdi, Abbas Ali

    2014-12-01

    Abnormalities in ovarian function, including defective oogenesis and folliculogenesis, represent a key female reproductive deficiency. Accumulating evidence in the literature has shown that the PI3K/PTEN/Akt and TSC/mTOR signaling pathways are critical regulators of ovarian function including quiescence, activation, and survival of primordial follicles, granulosa cell proliferation and differentiation, and meiotic maturation of oocytes. Dysregulation of these signaling pathways may contribute to infertility caused by impaired follicular development, intrafollicular oocyte development, and ovulation. This article reviews the current state of knowledge of the functional role of the PI3K/PTEN/Akt and TSC/mTOR pathways during mammalian oogenesis and folliculogenesis and their association with female infertility.

  11. Testosterone and Voluntary Exercise, Alone or Together Increase Cardiac Activation of AKT and ERK1/2 in Diabetic Rats

    PubMed Central

    Chodari, Leila; Mohammadi, Mustafa; Mohaddes, Gisou; Alipour, Mohammad Reza; Ghorbanzade, Vajiheh; Dariushnejad, Hassan; Mohammadi, Shima

    2016-01-01

    Background Impaired angiogenesis in cardiac tissue is a major complication of diabetes. Protein kinase B (AKT) and extracellular signal regulated kinase (ERK) signaling pathways play important role during capillary-like network formation in angiogenesis process. Objectives To determine the effects of testosterone and voluntary exercise on levels of vascularity, phosphorylated Akt (P- AKT) and phosphorylated ERK (P-ERK) in heart tissue of diabetic and castrated diabetic rats. Methods Type I diabetes was induced by i.p injection of 50 mg/kg of streptozotocin in animals. After 42 days of treatment with testosterone (2mg/kg/day) or voluntary exercise alone or in combination, heart tissue samples were collected and used for histological evaluation and determination of P-AKT and P-ERK levels by ELISA method. Results Our results showed that either testosterone or exercise increased capillarity, P-AKT, and P-ERK levels in the heart of diabetic rats. Treatment of diabetic rats with testosterone and exercise had a synergistic effect on capillarity, P-AKT, and P-ERK levels in heart. Furthermore, in the castrated diabetes group, capillarity, P-AKT, and P-ERK levels significantly decreased in the heart, whereas either testosterone treatment or exercise training reversed these effects. Also, simultaneous treatment of castrated diabetic rats with testosterone and exercise had an additive effect on P-AKT and P-ERK levels. Conclusion Our findings suggest that testosterone and exercise alone or together can increase angiogenesis in the heart of diabetic and castrated diabetic rats. The proangiogenesis effects of testosterone and exercise are associated with the enhanced activation of AKT and ERK1/2 in heart tissue.

  12. In vitro and in vivo activity of novel small-molecule inhibitors targeting the pleckstrin homology domain of protein kinase B/AKT.

    PubMed

    Moses, Sylvestor A; Ali, M Ahad; Zuohe, Song; Du-Cuny, Lei; Zhou, Li Li; Lemos, Robert; Ihle, Nathan; Skillman, A Geoffrey; Zhang, Shuxing; Mash, Eugene A; Powis, Garth; Meuillet, Emmanuelle J

    2009-06-15

    The phosphatidylinositol 3-kinase/AKT signaling pathway plays a critical role in activating survival and antiapoptotic pathways within cancer cells. Several studies have shown that this pathway is constitutively activated in many different cancer types. The goal of this study was to discover novel compounds that bind to the pleckstrin homology (PH) domain of AKT, thereby inhibiting AKT activation. Using proprietary docking software, 22 potential PH domain inhibitors were identified. Surface plasmon resonance spectroscopy was used to measure the binding of the compounds to the expressed PH domain of AKT followed by an in vitro activity screen in Panc-1 and MiaPaCa-2 pancreatic cancer cell lines. We identified a novel chemical scaffold in several of the compounds that binds selectively to the PH domain of AKT, inducing a decrease in AKT activation and causing apoptosis at low micromolar concentrations. Structural modifications of the scaffold led to compounds with enhanced inhibitory activity in cells. One compound, 4-dodecyl-N-(1,3,4-thiadiazol-2-yl)benzenesulfonamide, inhibited AKT and its downstream targets in cells as well as in pancreatic cancer cell xenografts in immunocompromised mice; it also exhibited good antitumor activity. In summary, a pharmacophore for PH domain inhibitors targeting AKT function was developed. Computer-aided modeling, synthesis, and testing produced novel AKT PH domain inhibitors that exhibit promising preclinical properties.

  13. Labdane diterpenes protect against anoxia/reperfusion injury in cardiomyocytes: involvement of AKT activation

    PubMed Central

    Cuadrado, I; Fernández-Velasco, M; Boscá, L; de las Heras, B

    2011-01-01

    Several labdane diterpenes exert anti-inflammatory and cytoprotective actions; therefore, we have investigated whether these molecules protect cardiomyocytes in an anoxia/reperfusion (A/R) model, establishing the molecular mechanisms involved in the process. The cardioprotective activity of three diterpenes (T1, T2 and T3) was studied in the H9c2 cell line and in isolated rat cardiomyocyte subjected to A/R injury. In both cases, treatment with diterpenes T1 and T2 protected from A/R-induced apoptosis, as deduced by a decrease in the percentage of apoptotic and caspase-3 active positive cells, a decrease in the Bcl-2/Bax ratio and an increase in the expression of antiapoptotic proteins. Analysis of cell survival signaling pathways showed that diterpenes T1 and T2 added after A/R increased phospho-AKT and phospho-ERK 1/2 levels. These cardioprotective effects were lost when AKT activity was pharmacologically inhibited. Moreover, the labdane-induced cardioprotection involves activation of AMPK, suggesting a role for energy homeostasis in their mechanism of action. Labdane diterpenes (T1 and T2) also exerted cardioprotective effects against A/R-induced injury in isolated cardiomyocytes and the mechanisms involved activation of specific survival signals (PI3K/AKT pathways, ERK1/2 and AMPK) and inhibition of apoptosis. PMID:22071634

  14. COMP-angiopoietin 1 increases proliferation, differentiation, and migration of stem-like cells through Tie-2-mediated activation of p38 MAPK and PI3K/Akt signal transduction pathways

    SciTech Connect

    Kook, Sung-Ho; Lim, Shin-Saeng; Cho, Eui-Sic; Lee, Young-Hoon; Han, Seong-Kyu; Lee, Kyung-Yeol; Kwon, Jungkee; Hwang, Jae-Won; Bae, Cheol-Hyeon; Seo, Young-Kwon; Lee, Jeong-Chae

    2014-12-12

    Highlights: • COMP-Ang1 induces Tie-2 activation in BMMSCs, but not in primary osteoblasts. • Tie-2 knockdown inhibits COMP-Ang1-stimulated proliferation and osteoblastogenesis. • Tie-2 knockdown prevents COMP-Ang1-induced activation of PI3K/Akt and p38 MAPK. • COMP-Ang1 induces migration of cells via activation of PI3K/Akt and CXCR4 pathways. • COMP-Ang1 stimulates in vivo migration of PDLSCs into a calvarial defect site of rats. - Abstract: Recombinant COMP-Ang1, a chimera of angiopoietin-1 (Ang1) and a short coiled-coil domain of cartilage oligomeric matrix protein (COMP), is under consideration as a therapeutic agent capable of inducing the homing of cells with increased angiogenesis. However, the potentials of COMP-Ang1 to stimulate migration of mesenchymal stem cells (MSCs) and the associated mechanisms are not completely understood. We examined the potential of COMP-Ang1 on bone marrow (BM)-MSCs, human periodontal ligament stem cells (PDLSCs), and calvarial osteoblasts. COMP-Ang1 augmented Tie-2 induction at protein and mRNA levels and increased proliferation and expression of runt-related transcription factor 2 (Runx2), osterix, and CXCR4 in BMMSCs, but not in osteoblasts. The COMP-Ang1-mediated increases were inhibited by Tie-2 knockdown and by treating inhibitors of phosphoinositide 3-kinase (PI3K), LY294002, or p38 mitogen-activated protein kinase (MAPK), SB203580. Phosphorylation of p38 MAPK and Akt was prevented by siRNA-mediated silencing of Tie-2. COMP-Ang1 also induced in vitro migration of BMMSCs and PDLSCs. The induced migration was suppressed by Tie-2 knockdown and by CXCR4-specific peptide antagonist or LY294002, but not by SB203580. Furthermore, COMP-Ang1 stimulated the migration of PDLSCs into calvarial defect site of rats. Collectively, our results demonstrate that COMP-Ang1-stimulated proliferation, differentiation, and migration of progenitor cells may involve the Tie-2-mediated activation of p38 MAPK and PI3K/Akt pathways.

  15. A Hot-spot of In-frame Duplications Activates the Oncoprotein AKT1 in Juvenile Granulosa Cell Tumors

    PubMed Central

    Bessière, Laurianne; Todeschini, Anne-Laure; Auguste, Aurélie; Sarnacki, Sabine; Flatters, Delphine; Legois, Bérangère; Sultan, Charles; Kalfa, Nicolas; Galmiche, Louise; Veitia, Reiner A.

    2015-01-01

    Background Ovarian granulosa cell tumors are the most common sex-cord stromal tumors and have juvenile (JGCTs) and adult forms. In a previous study we reported the occurrence of activating somatic mutations of Gαs, which transduces mitogenic signals, in 30% of the analyzed JGCTs. Methods We have searched for alterations in other proteins involved in ovarian mitogenic signaling. We focused on the PI3K–AKT axis. As we found mutations in AKT1, we analyzed the subcellular localization of the mutated proteins and performed functional explorations using Western-blot and luciferase assays. Findings We detected in-frame duplications affecting the pleckstrin-homology domain of AKT1 in more than 60% of the tumors occurring in girls under 15 years of age. The somatic status of the mutations was confirmed when peritumoral DNA was available. The JGCTs without duplications carried point mutations affecting highly conserved residues. Several of these substitutions were somatic lesions. The mutated proteins carrying the duplications had a non-wild-type subcellular distribution, with a marked enrichment at the plasma membrane. This led to a striking degree of AKT1 activation demonstrated by a strong phosphorylation level and by reporter assays. Interpretation Our study incriminates somatic mutations of AKT1 as a major event in the pathogenesis of JGCTs. The existence of AKT inhibitors currently tested in clinical trials opens new perspectives for targeted therapies for these tumors, which are currently treated with standard non-specific chemotherapy protocols. PMID:26137586

  16. Transcriptional activation of peroxisome proliferator-activated receptor-{gamma} requires activation of both protein kinase A and Akt during adipocyte differentiation

    SciTech Connect

    Kim, Sang-pil; Ha, Jung Min; Yun, Sung Ji; Kim, Eun Kyoung; Chung, Sung Woon; Hong, Ki Whan; Kim, Chi Dae; Bae, Sun Sik

    2010-08-13

    Research highlights: {yields} Elevated cAMP activates both PKA and Epac. {yields} PKA activates CREB transcriptional factor and Epac activates PI3K/Akt pathway via Rap1. {yields} Akt modulates PPAR-{gamma} transcriptional activity in concert with CREB. -- Abstract: Peroxisome proliferator-activated receptor-{gamma} (PPAR-{gamma}) is required for the conversion of pre-adipocytes. However, the mechanism underlying activation of PPAR-{gamma} is unclear. Here we showed that cAMP-induced activation of protein kinase A (PKA) and Akt is essential for the transcriptional activation of PPAR-{gamma}. Hormonal induction of adipogenesis was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor (LY294002), by a protein kinase A (PKA) inhibitor (H89), and by a Rap1 inhibitor (GGTI-298). Transcriptional activity of PPAR-{gamma} was markedly enhanced by 3-isobutyl-1-methylxanthine (IBMX), but not insulin and dexamethasone. In addition, IBMX-induced PPAR-{gamma} transcriptional activity was blocked by PI3K/Akt, PKA, or Rap1 inhibitors. 8-(4-Chlorophenylthio)-2'-O-methyl-cAMP (8-pCPT-2'-O-Me-cAMP) which is a specific agonist for exchanger protein directly activated by cAMP (Epac) significantly induced the activation of Akt. Furthermore, knock-down of Akt1 markedly attenuated PPAR-{gamma} transcriptional activity. These results indicate that both PKA and Akt signaling pathways are required for transcriptional activation of PPAR-{gamma}, suggesting post-translational activation of PPAR-{gamma} might be critical step for adipogenic gene expression.

  17. EGFR-AKT-mTOR activation mediates epiregulin-induced pleiotropic functions in cultured osteoblasts.

    PubMed

    Fan, Jian-Bo; Liu, Wei; Zhu, Xin-Hui; Yuan, Kun; Xu, Da-Wei; Chen, Jia-Jia; Cui, Zhi-Ming

    2015-01-01

    Epidermal growth factor (EGF) receptor (EGFR) emerges as an essential molecule for the regulating of osteoblast cellular functions. In the current study, we explored the effect of epiregulin, a new EGFR ligand, on osteoblast functions in vitro, and studied the underlying mechanisms. We found that epiregulin-induced EGFR activation in both primary osteoblasts and osteoblast-like MC3T3-E1 cells. Meanwhile, epiregulin activated AKT-mammalian target of rapamycin (mTOR) and Erk-mitogen-activated protein kinase (MAPK) signalings in cultured osteoblasts, which were blocked by EGFR inhibitor AG1478 or monoclonal antibody against EGFR (anti-EGFR). Further, in primary and MC3T3-E1 osteoblasts, epiregulin promoted cell proliferation and increased alkaline phosphatase activity, while inhibiting dexamethasone (Dex)-induced cell death. Such effects by epiregulin were largely inhibited by AG1478 or anti-EGFR. Notably, AKT-mTOR inhibitors, but not Erk inhibitors, alleviated epiregulin-induced above pleiotropic functions in osteoblasts. Meanwhile, siRNA depletion of Sin1, a key component of mTOR complex 2 (mTORC2), also suppressed epiregulin-exerted effects in MC3T3-E1 cells. Together, these results suggest that epiregulin-induced pleiotropic functions in cultured osteoblasts are mediated through EGFR-AKT-mTOR signalings.

  18. PI3K/Akt is involved in brown adipogenesis mediated by growth differentiation factor-5 in association with activation of the Smad pathway

    SciTech Connect

    Hinoi, Eiichi; Iezaki, Takashi; Fujita, Hiroyuki; Watanabe, Takumi; Odaka, Yoshiaki; Ozaki, Kakeru; Yoneda, Yukio

    2014-07-18

    Highlights: • Akt is preferentially phosphorylated in BAT and sWAT of aP2-GDF5 mice. • PI3K/Akt signaling is involved in GDF5-induced brown adipogenesis. • PI3K/Akt signaling regulates GDF5-induced Smad5 phosphorylation. - Abstract: We have previously demonstrated promotion by growth differentiation factor-5 (GDF5) of brown adipogenesis for systemic energy expenditure through a mechanism relevant to activating the bone morphological protein (BMP) receptor/mothers against decapentaplegic homolog (Smad)/peroxisome proliferator-activated receptor gamma co-activator 1α (PGC-1α) pathway. Here, we show the involvement of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in brown adipogenesis mediated by GDF5. Overexpression of GDF5 in cells expressing adipocyte protein-2 markedly accelerated the phosphorylation of Smad1/5/8 and Akt in white and brown adipose tissues. In brown adipose tissue from heterozygous GDF5{sup Rgsc451} mutant mice expressing a dominant-negative (DN) GDF5 under obesogenic conditions, the basal phosphorylation of Smad1/5/8 and Akt was significantly attenuated. Exposure to GDF5 not only promoted the phosphorylation of both Smad1/5/8 and Akt in cultured brown pre-adipocytes, but also up-regulated Pgc1a and uncoupling protein-1 expression in a manner sensitive to the PI3K/Akt inhibitor Ly294002 as well as retroviral infection with DN-Akt. GDF5 drastically promoted BMP-responsive luciferase reporter activity in a Ly294002-sensitive fashion. Both Ly294002 and DN-Akt markedly inhibited phosphorylation of Smad5 in the nuclei of brown pre-adipocytes. These results suggest that PI3K/Akt signals play a role in the GDF5-mediated brown adipogenesis through a mechanism related to activation of the Smad pathway.

  19. Early single Aspirin-triggered Lipoxin blocked morphine anti-nociception tolerance through inhibiting NALP1 inflammasome: Involvement of PI3k/Akt signaling pathway.

    PubMed

    Tian, Yu; Liu, Ming; Mao-Ying, Qi-Liang; Liu, Huan; Wang, Zhi-Fu; Zhang, Meng-Ting; Wang, Jun; Li, Qian; Liu, Shen-Bin; Mi, Wen-Li; Ma, Hong-Jian; Wu, Gen-Cheng; Wang, Yan-Qing

    2015-11-01

    Clinical usage of opioids in pain relief is dampened by analgesic tolerance after chronic exposure, which is related to opioid-associated neuroinflammation. In the current study, which is based on a chronic morphine tolerance rat model and sustained morphine treatment on primary neuron culture, it was observed that Akt phosphorylation, cleaved-Caspase-1-dependent NALP1 inflammasome activation and IL-1β maturation in spinal cord neurons were significantly enhanced by morphine. Moreover, treatment with LY294002, a specific inhibitor of PI3k/Akt signaling, significantly reduced Caspase-1 cleavage, NALP1 inflammasome activation and attenuated morphine tolerance. Tail-flick tests demonstrated that pharmacological inhibition on Caspase-1 activation or antagonizing IL-1β dramatically blocked the development of morphine tolerance. The administration of an exogenous analogue of lipoxin, Aspirin-triggered Lipoxin (ATL), caused a decline in Caspase-1 cleavage, inflammasome activation and mature IL-1β production and thus attenuated the development of morphine tolerance by inhibiting upstream Akt phosphorylation. Additionally, treatment with DAMGO, a selective μ-opioid receptor peptide, significantly induced Akt phosphorylation, Caspase-1 cleavage and anti-nociception tolerance, all of which were attenuated by ATL treatment. Taken together, the present study revealed the involvement of spinal NALP1 inflammasome activation in the development of morphine tolerance and the role of the μ-receptor/PI3k-Akt signaling/NALP1 inflammasome cascade in this process. By inhibiting this signaling cascade, ATL blocked the development of morphine tolerance.

  20. Down-Regulation of AKT Signalling by Ursolic Acid Induces Intrinsic Apoptosis and Sensitization to Doxorubicin in Soft Tissue Sarcoma

    PubMed Central

    Villar, Victor Hugo; Vögler, Oliver; Barceló, Francisca; Martín-Broto, Javier; Martínez-Serra, Jordi; Ruiz-Gutiérrez, Valentina; Alemany, Regina

    2016-01-01

    Several important biological activities have been attributed to the pentacyclic triterpene ursolic acid (UA), being its antitumoral effect extensively studied in human adenocarcinomas. In this work, we focused on the efficacy and molecular mechanisms involved in the antitumoral effects of UA, as single agent or combined with doxorubicin (DXR), in human soft tissue sarcoma cells. UA (5–50 μM) strongly inhibited (up to 80%) the viability of STS cells at 24 h and its proliferation in soft agar, with higher concentrations increasing apoptotic death up to 30%. UA treatment (6–9 h) strongly blocked the survival AKT/GSK3β/β-catenin signalling pathway, which led to a concomitant reduction of the anti-apoptotic proteins c-Myc and p21, altogether resulting in the activation of intrinsic apoptosis. Interestingly, UA at low concentrations (10–15 μM) enhanced the antitumoral effects of DXR by up to 2-fold, while in parallel inhibiting DXR-induced AKT activation and p21 expression, two proteins implicated in antitumoral drug resistance and cell survival. In conclusion, UA is able to induce intrinsic apoptosis in human STS cells and also to sensitize these cells to DXR by blocking the AKT signalling pathway. Therefore, UA may have beneficial effects, if used as nutraceutical adjuvant during standard chemotherapy treatment of STS. PMID:27219337

  1. AKT activation drives the nuclear localization of CSE1L and a pro-oncogenic transcriptional activation in ovarian cancer cells

    SciTech Connect

    Lorenzato, Annalisa; Biolatti, Marta; Delogu, Giuseppe; Capobianco, Giampiero; Farace, Cristiano; Dessole, Salvatore; Cossu, Antonio; Tanda, Francesco; Madeddu, Roberto; Olivero, Martina; Di Renzo, Maria Flavia

    2013-10-15

    The human homolog of the yeast cse1 gene (CSE1L) is over-expressed in ovarian cancer. CSE1L forms complex with Ran and importin-α and has roles in nucleocytoplasmic traffic and gene expression. CSE1L accumulated in the nucleus of ovarian cancer cell lines, while it was localized also in the cytoplasm of other cancer cell lines. Nuclear localization depended on AKT, which was constitutively active in ovarian cancer cells, as the CSE1L protein translocated to the cytoplasm when AKT was inactivated. Moreover, the expression of a constitutively active AKT forced the translocation of CSE1L from the cytoplasm to the nucleus in other cancer cells. Nuclear accrual of CSE1L was associated to the nuclear accumulation of the phosphorylated Ran Binding protein 3 (RanBP3), which depended on AKT as well. Also in samples of human ovarian cancer, AKT activation was associated to nuclear accumulation of CSE1L and phosphorylation of RanBP3. Expression profiling of ovarian cancer cells after CSE1L silencing showed that CSE1L was required for the expression of genes promoting invasion and metastasis. In agreement, CSE1L silencing impaired motility and invasiveness of ovarian cancer cells. Altogether these data show that in ovarian cancer cells activated AKT by affecting RanBP3 phosphorylation determines the nuclear accumulation of CSE1L and likely the nuclear concentration of transcription factors conveying pro-oncogenic signals. - highlights: • CSE1L is a key player in nucleocytoplasmic traffic by forming complex with Ran. • AKT phosphorylates RanBP3 that regulates the nucleocytoplasmic gradient of Ran. • The activated oncogenic AKT drives the nuclear accumulation of CSE1L. • CSE1L in the nucleus up-regulates genes conveying pro-oncogenic signals. • CSE1L might contribute to tumor progression driven by the activated oncogenic AKT.

  2. OSU-T315: a novel targeted therapeutic that antagonizes AKT membrane localization and activation of chronic lymphocytic leukemia cells

    PubMed Central

    Liu, Ta-Ming; Ling, Yonghua; Woyach, Jennifer A.; Beckwith, Kyle; Yeh, Yuh-Ying; Hertlein, Erin; Zhang, Xiaoli; Lehman, Amy; Awan, Farrukh; Jones, Jeffrey A.; Andritsos, Leslie A.; Maddocks, Kami; MacMurray, Jessica; Salunke, Santosh B.; Chen, Ching-Shih; Phelps, Mitch A.; Byrd, John C.

    2015-01-01

    Aberrant regulation of endogenous survival pathways plays a major role in progression of chronic lymphocytic leukemia (CLL). Signaling via conjugation of surface receptors within the tumor environmental niche activates survival and proliferation pathways in CLL. Of these, the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) pathway appears to be pivotal to support CLL pathogenesis, and pharmacologic inhibitors targeting this axis have shown clinical activity. Here we investigate OSU-T315, a compound that disrupts the PI3K/AKT pathway in a novel manner. Dose-dependent selective cytotoxicity by OSU-T315 is noted in both CLL-derived cell lines and primary CLL cells relative to normal lymphocytes. In contrast to the highly successful Bruton's tyrosine kinase and PI3K inhibitors that inhibit B-cell receptor (BCR) signaling pathway at proximal kinases, OSU-T315 directly abrogates AKT activation by preventing translocation of AKT into lipid rafts without altering the activation of receptor-associated kinases. Through this mechanism, the agent triggers caspase-dependent apoptosis in CLL by suppressing BCR, CD49d, CD40, and Toll-like receptor 9-mediated AKT activation in an integrin-linked kinase-independent manner. In vivo, OSU-T315 attains pharmacologically active drug levels and significantly prolongs survival in the TCL1 mouse model. Together, our findings indicate a novel mechanism of action of OSU-T315 with potential therapeutic application in CLL. PMID:25293770

  3. OSU-T315: a novel targeted therapeutic that antagonizes AKT membrane localization and activation of chronic lymphocytic leukemia cells.

    PubMed

    Liu, Ta-Ming; Ling, Yonghua; Woyach, Jennifer A; Beckwith, Kyle; Yeh, Yuh-Ying; Hertlein, Erin; Zhang, Xiaoli; Lehman, Amy; Awan, Farrukh; Jones, Jeffrey A; Andritsos, Leslie A; Maddocks, Kami; MacMurray, Jessica; Salunke, Santosh B; Chen, Ching-Shih; Phelps, Mitch A; Byrd, John C; Johnson, Amy J

    2015-01-08

    Aberrant regulation of endogenous survival pathways plays a major role in progression of chronic lymphocytic leukemia (CLL). Signaling via conjugation of surface receptors within the tumor environmental niche activates survival and proliferation pathways in CLL. Of these, the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) pathway appears to be pivotal to support CLL pathogenesis, and pharmacologic inhibitors targeting this axis have shown clinical activity. Here we investigate OSU-T315, a compound that disrupts the PI3K/AKT pathway in a novel manner. Dose-dependent selective cytotoxicity by OSU-T315 is noted in both CLL-derived cell lines and primary CLL cells relative to normal lymphocytes. In contrast to the highly successful Bruton's tyrosine kinase and PI3K inhibitors that inhibit B-cell receptor (BCR) signaling pathway at proximal kinases, OSU-T315 directly abrogates AKT activation by preventing translocation of AKT into lipid rafts without altering the activation of receptor-associated kinases. Through this mechanism, the agent triggers caspase-dependent apoptosis in CLL by suppressing BCR, CD49d, CD40, and Toll-like receptor 9-mediated AKT activation in an integrin-linked kinase-independent manner. In vivo, OSU-T315 attains pharmacologically active drug levels and significantly prolongs survival in the TCL1 mouse model. Together, our findings indicate a novel mechanism of action of OSU-T315 with potential therapeutic application in CLL.

  4. WISP1 overexpression promotes proliferation and migration of human vascular smooth muscle cells via AKT signaling pathway.

    PubMed

    Lu, Shun; Liu, Hao; Lu, Lihe; Wan, Heng; Lin, Zhiqi; Qian, Kai; Yao, Xingxing; Chen, Qing; Liu, Wenjun; Yan, Jianyun; Liu, Zhengjun

    2016-10-05

    Proliferation and migration of vascular smooth muscle cells (VSMCs) play crucial roles in the development of vascular restenosis. Our previous study showed that CCN4, namely Wnt1 inducible signaling pathway protein 1 (WISP1), significantly promotes proliferation and migration of rat VSMCs, but its mechanism remains unclear. This study aims to investigate whether and how WISP1 stimulates proliferation and migration of human VSMCs. Western blot analysis showed that FBS treatment increased WISP1 protein levels in human VSMCs in a dose-dependent manner. Overexpression of WISP1 using adenovirus encoding WISP1 (AD-WISP1) significantly increased proliferation rate of human VSMCs by 2.98-fold compared with empty virus (EV)-transfected cells, shown by EdU incorporation assay. Additionally, Scratch-induced wound healing assay revealed that adenovirus-mediated overexpression of WISP1 significantly increased cell migration compared with EV-transfected cells from 6h (4.56±1.14% vs. 11.23±2.25%, P<0.05) to 48h (25.25±5.51% vs. 97.54±13.12%, P<0.01) after injury. Transwell Migration Assay confirmed that WISP1 overexpression significantly promoted human VSMC migration by 2.25-fold compared with EV. Furthermore, WISP1 overexpression stimulated Akt signaling activation in human VSMCs. Blockage of Akt signaling by Akt inhibitor AZD5363 or PI3K inhibitor LY294002, led to an inhibitory effect of WISP1-induced proliferation and migration in human VSMCs. Moreover, we found that WISP1 overexpression stimulated GSK3α/β phosphorylation, and increased expression of cyclin D1 and MMP9 in human VSMCs, and this effect was abolished by AZD5363. Collectively, we demonstrated that Akt signaling pathway mediates WISP1-induced migration and proliferation of human VSMCs, suggesting that WISP1 may act as a novel potential therapeutic target for vascular restenosis.

  5. Regulation of PtdIns(3,4,5)P3/Akt signalling by inositol polyphosphate 5-phosphatases.

    PubMed

    Eramo, Matthew J; Mitchell, Christina A

    2016-02-01

    The phosphoinositide 3-kinase (PI3K) generated lipid signals, PtdIns(3,4,5)P3 and PtdIns(3,4)P2, are both required for the maximal activation of the serine/threonine kinase proto-oncogene Akt. The inositol polyphosphate 5-phosphatases (5-phosphatases) hydrolyse the 5-position phosphate from the inositol head group of PtdIns(3,4,5)P3 to yield PtdIns(3,4)P2. Extensive work has revealed several 5-phosphatases inhibit PI3K-driven Akt signalling, by decreasing PtdIns(3,4,5)P3 despite increasing cellular levels of PtdIns(3,4)P2. The roles that 5-phosphatases play in suppressing cell proliferation and transformation are slow to emerge; however, the 5-phosphatase PIPP [proline-rich inositol polyphosphate 5-phosphatase; inositol polyphosphate 5-phosphatase (INPP5J)] has recently been identified as a putative tumour suppressor in melanoma and breast cancer and SHIP1 [SH2 (Src homology 2)-containing inositol phosphatase 1] inhibits haematopoietic cell proliferation. INPP5E regulates cilia stability and INPP5E mutations have been implicated ciliopathy syndromes. This review will examine 5-phosphatase regulation of PI3K/Akt signalling, focussing on the role PtdIns(3,4,5)P3 5-phosphatases play in developmental diseases and cancer.

  6. MicroRNA-21 accelerates hepatocyte proliferation in vitro via PI3K/Akt signaling by targeting PTEN

    SciTech Connect

    Yan-nan, Bai; Zhao-yan, Yu; Li-xi, Luo; Jiang, Yi; Qing-jie, Xia

    2014-01-17

    Highlights: •miRNAs-expression patterns of primary hepatocytes under proliferative status. •miR-21 expression level peaked at 12 h after stimulated by EGF. •miR-21 drive rapid S phase entry of primary hepatocytes. •PI3K/Akt signaling was modulated via targeting PTEN by miR-21. -- Abstract: MicroRNAs (miRNAs) are involved in controlling hepatocyte proliferation during liver regeneration. In this study, we established the miRNAs-expression patterns of primary hepatocytes in vitro under stimulation of epidermal growth factor (EGF), and found that microRNA-21 (miR-21) was appreciably up-regulated and peaked at 12 h. In addition, we further presented evidences indicating that miR-21 promotes primary hepatocyte proliferation through in vitro transfecting with miR-21 mimics or inhibitor. We further demonstrated that phosphatidylinositol 3′-OH kinase (PI3K)/Akt signaling was altered accordingly, it is, by targeting phosphatase and tensin homologue deleted on chromosome 10, PI3K/Akt signaling is activated by miR-21 to accelerate hepatocyte rapid S-phase entry and proliferation in vitro.

  7. Restoration of Akt activity by the bisperoxovanadium compound bpV(pic) attenuates hippocampal apoptosis in experimental neonatal pneumococcal meningitis.

    PubMed

    Sury, Matthias D; Vorlet-Fawer, Lorianne; Agarinis, Claudia; Yousefi, Shida; Grandgirard, Denis; Leib, Stephen L; Christen, Stephan

    2011-01-01

    Pneumococcal meningitis causes apoptosis of developing neurons in the dentate gyrus of the hippocampus. The death of these cells is accompanied with long-term learning and memory deficits in meningitis survivors. Here, we studied the role of the PI3K/Akt (protein kinase B) survival pathway in hippocampal apoptosis in a well-characterized infant rat model of pneumococcal meningitis. Meningitis was accompanied by a significant decrease of the PI3K product phosphatidylinositol 3,4,5-trisphosphate (PIP(3)) and of phosphorylated (i.e., activated) Akt in the hippocampus. At the cellular level, phosphorylated Akt was decreased in both the granular layer and the subgranular zone of the dentate gyrus, the region where the developing neurons undergo apoptosis. Protein levels and activity of PTEN, the major antagonist of PI3K, were unaltered by infection, suggesting that the observed decrease in PIP(3) and Akt phosphorylation is a result of decreased PI3K signaling. Treatment with the PTEN inhibitor bpV(pic) restored Akt activity and significantly attenuated hippocampal apoptosis. Co-treatment with the specific PI3K inhibitor LY294002 reversed the restoration of Akt activity and attenuation of hippocampal apoptosis, while it had no significant effect on these parameters on its own. These results indicate that the inhibitory effect of bpV(pic) on apoptosis was mediated by PI3K-dependent activation of Akt, strongly suggesting that bpV(pic) acted on PTEN. Treatment with bpV(pic) also partially inhibited the concentration of bacteria and cytokines in the CSF, but this effect was not reversed by LY294002, indicating that the effect of bpV(pic) on apoptosis was independent of its effect on CSF bacterial burden and cytokine levels. These results indicate that the PI3K/Akt pathway plays an important role in the death and survival of developing hippocampal neurons during the acute phase of pneumococcal meningitis.

  8. Protective Effect of Tempol on Acute Kidney Injury Through PI3K/Akt/Nrf2 Signaling Pathway

    PubMed Central

    Zhang, Gensheng; Wang, Qiaoling; Zhou, Qin; Wang, Renjun; Xu, Minze; Wang, Huiping; Wang, Lei; Wilcox, Christopher S.; Liu, Ruisheng; Lai, En Yin

    2016-01-01

    Background/Aims Tempol is a protective antioxidant against ischemic injury in many animal models. The molecular mechanisms are not well understood. Nuclear factor erythroid 2-related factor (Nrf2) is a master transcription factor during oxidative stress, which is enhanced by activation of protein kinase C (PKC) pathway. Another factor, tubular epithelial apoptosis, is mediated by activation of phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB, Akt) signaling pathway during renal ischemic injury. We tested the hypothesis that tempol activates PKC or PI3K/Akt/Nrf2 pathways to transcribe many genes that coordinate endogenous antioxidant defense. Methods The right renal pedicle was clamped for 45 minutes and the left kidney was removed to study renal ischemia/reperfusion (I/R) injury in C57BL/6 mice. The response was assessed from serum parameters, renal morphology and renal expression of PKC, phosphorylated-PKC (p-PKC), Nrf2, heme oxygenase-1 (HO-1), Akt, phosphorylated-Akt (p-Akt), pro-caspase-3 and cleaved caspase-3 in groups of sham and I/R mice given vehicle, or tempol (50 or 100 mg/kg, intraperitoneal injection). Results The serum malondialdehyde (MDA, marker of reactive oxygen species) doubled and the BUN and creatinine increased 5- to 10-fold after I/R injury. Tempol (50 or 100 mg/kg) prevented the increases in MDA but only tempol (50 mg/kg) lessened the increases in BUN and creatinine and moderated the acute tubular necrosis. I/R did not change expression of PKC or p-PKC but reduced renal expression of Nrf2, p-Akt, HO-1 and pro-caspase-3 and increased cleaved caspase-3. Tempol (50 mg/kg) prevented these changes produced by I/R whereas tempol (100 mg/kg) had lesser or inconsistent effects. Conclusion Tempol (50 mg/kg) prevents lipid peroxidation and attenuates renal damage after I/R injury. The beneficial pathway apparently is not dependent on upregulation or phosphorylation of PKC, at lower tempol doses, does implicate upregulation of Akt with expression

  9. Arctigenin, a Natural Lignan Compound, Induces Apoptotic Death of Hepatocellular Carcinoma Cells via Suppression of PI3-K/Akt Signaling.

    PubMed

    Jiang, Xiaoxin; Zeng, Leping; Huang, Jufang; Zhou, Hui; Liu, Yubin

    2015-04-28

    In this study, we explored the cytotoxic effects of arctigenin, a natural lignan compound, on human hepatocellular carcinoma (HCC) cells and check the involvement of phosphatidylinositol 3-kinase (PI3-K)/Akt signaling. HCC cells were treated with different concentrations of arctigenin and cell viability and apoptosis were assessed. Manipulating Akt signaling was used to determine its role in the action of arctigenin. Arctigenin significantly inhibited the viability of HCC cells in a concentration-dependent manner. Arctigenin induced apoptosis and activation of caspase-9 and -3. Overexpression of a constitutively active Akt mutant blocked arctigenin-induced apoptosis. Combinational treatment with arctigenin and the PI3-K inhibitor LY294002 significantly enhanced apoptosis. Arctigenin reduced the expression of Bcl-xL, Mcl-1, and survivin and the phosphorylation of mTOR and S6K, which were significantly reversed by overexpression of constitutively active Akt. This is the first report about the anticancer activity of arctigenin in HCC cells, which is mediated by inactivation of PI3-K/Akt signaling.

  10. Dorsomorphin and LDN-193189 inhibit BMP-mediated Smad, p38 and Akt signalling in C2C12 cells.

    PubMed

    Boergermann, J H; Kopf, J; Yu, P B; Knaus, P

    2010-11-01

    Bone morphogenetic proteins (BMPs) are key regulators of cell fate decisions during embryogenesis and tissue homeostasis. BMPs signal through a coordinated assembly of two types of transmembrane serine/threonine kinase receptors to induce Smad1/5/8 plus non-Smad pathways, such as MAPK and Akt. The recent discovery of BMP receptor inhibitors opened new avenues to study specific BMP signalling and to delineate this effect from TGF-β and Activin signalling. Here we present comprehensive and quantitative analyses on both canonical and non-Smad mediated BMP signalling under Dorsomorphin (DM) and LDN-193189 (LDN) treatment conditions. We demonstrate for the first time, that both compounds affect not only the Smad but also the non-Smad signalling pathways induced by either BMP2, BMP6 or GDF5. The activation of p38, ERK1/2 and Akt in C2C12 cells was inhibited by DM and LDN. In addition "off-target" effects on all branches of BMP non-Smad signalling are presented. From this we conclude that the inhibition of BMP receptors by DM and more efficiently by LDN-193189 affects all known BMP induced signalling cascades.

  11. Stability and function of adult vasculature is sustained by Akt/Jagged1 signalling axis in endothelium

    PubMed Central

    Kerr, Bethany A.; West, Xiaoxia Z.; Kim, Young-Woong; Zhao, Yongzhong; Tischenko, Miroslava; Cull, Rebecca M.; Phares, Timothy W.; Peng, Xiao-Ding; Bernier-Latmani, Jeremiah; Petrova, Tatiana V.; Adams, Ralf H.; Hay, Nissim; Naga Prasad, Sathyamangla V.; Byzova, Tatiana V.

    2016-01-01

    The signalling pathways operational in quiescent, post-development vasculature remain enigmatic. Here we show that unlike neovascularization, endothelial Akt signalling in established vasculature is crucial not for endothelial cell (EC) survival, but for sustained interactions with pericytes and vascular smooth muscle cells (VSMCs) regulating vascular stability and function. Inducible endothelial-specific Akt1 deletion in adult global Akt2KO mice triggers progressive VSMC apoptosis. In hearts, this causes a loss of arteries and arterioles and, despite a high capillary density, diminished vascular patency and severe cardiac dysfunction. Similarly, endothelial Akt deletion induces retinal VSMC loss and basement membrane deterioration resulting in vascular regression and retinal atrophy. Mechanistically, the Akt/mTOR axis controls endothelial Jagged1 expression and, thereby, Notch signalling regulating VSMC maintenance. Jagged1 peptide treatment of Akt1ΔEC;Akt2KO mice and Jagged1 re-expression in Akt-deficient endothelium restores VSMC coverage. Thus, sustained endothelial Akt1/2 signalling is critical in maintaining vascular stability and homeostasis, thereby preserving tissue and organ function. PMID:26971877

  12. The inhibition of activated hepatic stellate cells proliferation by arctigenin through G0/G1 phase cell cycle arrest: persistent p27(Kip1) induction by interfering with PI3K/Akt/FOXO3a signaling pathway.

    PubMed

    Li, Ao; Wang, Jun; Wu, Mingjun; Zhang, Xiaoxun; Zhang, Hongzhi

    2015-01-15

    Proliferation of hepatic stellate cells (HSCs) is vital for the development of fibrosis during liver injury. In this study, we describe that arctigenin (ATG), a major bioactive component of Fructus Arctii, exhibited selective cytotoxic activity via inhibiting platelet-derived growth factor-BB (PDGF-BB)-activated HSCs proliferation and arrested cell cycle at G0/G1 phase, which could not be observed in normal human hepatocytes in vitro. The cyclin-dependent kinase (CDK) 4/6 activities could be strongly inhibited by ATG through down-regulation of cyclin D1 and CDK4/6 expression in early G1 phase arrest. In the ATG-treated HSCs, the expression level of p27(Kip1) and the formation of CDK2-p27(Kip1) complex were also increased. p27(Kip1) silencing significantly attenuated the effect of ATG, including cell cycle arrest and suppression of proliferation in activated HSCs. We also found that ATG suppressed PDGF-BB-induced phosphorylation of Akt and its downstream transcription factor Forkhead box O 3a (FOXO3a), decreased binding of FOXO3a to 14-3-3 protein, and stimulated nuclear translocation of FOXO3a in activated HSCs. Furthermore, knockdown of FOXO3a expression by FOXO3a siRNA attenuated ATG-induced up-regulation of p27(Kip1) in activated HSCs. All the above findings suggested that ATG could increase the levels of p27(Kip1) protein through inhibition of Akt and improvement of FOXO3a activity, in turn inhibited the CDK2 kinase activity, and eventually caused an overall inhibition of HSCs proliferation.

  13. Role of PI3K/Akt signaling in insulin-like growth factor-1 (IGF-1) skin tumor promotion.

    PubMed

    Wilker, Erik; Lu, Jerry; Rho, Okkyung; Carbajal, Steve; Beltrán, Linda; DiGiovanni, John

    2005-10-01

    Overexpression of human IGF-1 with the bovine keratin 5 (BK5) promoter (BK5.IGF-1 transgenic mice) induces persistent epidermal hyperplasia and leads to spontaneous skin tumor formation. In previous work, PI3K and Akt activities were found to be elevated in the epidermis of BK5.IGF-1 transgenic mice compared to nontransgenic littermates. In the present study, we examined the importance of the PI3K/Akt signaling pathway in mediating the skin phenotype and the skin tumor promoting action of IGF-1 in these mice. Western blot analyses with epidermal lysates showed that signaling components downstream of PI3K/Akt were altered in epidermis of BK5.IGF-1 mice. Increased phosphorylation of GSK-3 (Ser(9/21)), TSC2(Thr(1462)), and mTOR(Ser(2448)) was observed. In addition, hypophosphorylation and increased protein levels of beta-catenin were observed in the epidermis of BK5.IGF-1 mice. These data suggested that components downstream of Akt might be affected, including cell cycle machinery in the epidermis of BK5.IGF-1 mice. Protein levels of cyclins (D1, E, A), E2F1, and E2F4 were all elevated in the epidermis of BK5.IGF-1 mice. Also, immunoprecipitation experiments demonstrated an increase in cdk4/cyclin D1 and cdk2/cyclin E complex formation, suggesting increased cdk activity in the epidermis of transgenic mice. In further studies, the PI3K inhibitor, LY294002, significantly blocked IGF-1-mediated epidermal proliferation and skin tumor promotion in DMBA-initiated BK5.IGF-1 mice. In addition, inhibition of PI3K/Akt with LY294002 reversed many of the cell cycle related changes observed in untreated transgenic animals. Collectively, the current results supported the hypothesis that elevated PI3K/Akt activity and subsequent activation of one or more downstream effector pathways contributed significantly to the tumor promoting action of IGF-1 in the epidermis of BK5.IGF-1 mice.

  14. Analysis of miRNA profiles identified miR-196a as a crucial mediator of aberrant PI3K/AKT signaling in lung cancer cells.

    PubMed

    Guerriero, Ilaria; D'Angelo, Daniela; Pallante, Pierlorenzo; Santos, Mafalda; Scrima, Marianna; Malanga, Donatella; De Marco, Carmela; Ravo, Maria; Weisz, Alessandro; Laudanna, Carmelo; Ceccarelli, Michele; Falco, Geppino; Rizzuto, Antonia; Viglietto, Giuseppe

    2016-11-17

    Hyperactivation of the PI3K/AKT pathway is observed in most human cancer including lung carcinomas. Here we have investigated the role of miRNAs as downstream targets of activated PI3K/AKT signaling in Non Small Cell Lung Cancer (NSCLC). To this aim, miRNA profiling was performed in human lung epithelial cells (BEAS-2B) expressing active AKT1 (BEAS-AKT1-E17K), active PI3KCA (BEAS-PIK3CA-E545K) or with silenced PTEN (BEAS-shPTEN).Twenty-four differentially expressed miRNAs common to BEAS-AKT1-E17K, BEAS-PIK3CA-E545K and BEAS-shPTEN cells were identified through this analysis, with miR-196a being the most consistently up-regulated miRNA. Interestingly, miR-196a was significantly overexpressed also in human NSCLC-derived cell lines (n=11) and primary lung cancer samples (n=28).By manipulating the expression of miR-196a in BEAS-2B and NCI-H460 cells, we obtained compelling evidence that this miRNA acts downstream the PI3K/AKT pathway, mediating some of the proliferative, pro-migratory and tumorigenic activity that this pathway exerts in lung epithelial cells, possibly through the regulation of FoxO1, CDKN1B (hereafter p27) and HOXA9.

  15. Leptin reduces gentamicin-induced apoptosis in rat renal tubular cells via the PI3K-Akt signaling pathway.

    PubMed

    Chen, Yen-Cheng; Chen, Cheng-Hsien; Hsu, Yung-Ho; Chen, Tso-Hsiao; Sue, Yuh-Mou; Cheng, Chung-Yi; Chen, Tzen-Wen

    2011-05-11

    Leptin, a circulating hormone secreted mainly from adipose tissues, possesses protective effects on many cell types. Serum leptin concentration increases in patients with chronic renal failure and those undergoing maintenance dialysis. Gentamicin, a widely used antibiotic for the treatment of bacterial infection, can cause nephrotoxicity. In the present study, we intended to investigate the influence of leptin on apoptotic pathways and its mechanism in rat renal tubular cells treated with gentamicin. By using Annexin V-FITC/propidium iodide double staining, we found that leptin expressed a dose-dependent protective effect against gentamicin-induced apoptosis in rat renal tubular cells (NRK-52E) within 24h. Pretreatment of the cells with 50 or 100 ng/ml of leptin induced Bcl-2 and Bcl-x(L), increased the phosphorylation of Bad, and decreased the cleaved caspase-3 and caspase-9 in gentamicin-treated NRK-52E cells. Leptin also suppressed the activation of the transcription factor NF-κB and upregulated Akt activation in gentamicin-treated NRK-52E cells. We found that leptin activated the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway as demonstrated by the suppression of the anti-apoptotic effect of leptin by wortmannin. The treatment of wortmannin suppressed the leptin-induced phospho-Akt, Bcl-2, phospho-Bad as well as Bcl-x(L), and recovered the leptin-reduced cleaved caspase-3 and caspase-9. Based on our results, we suggested that leptin can attenuate gentamicin-induced apoptotic injury in rat renal tubular cells through PI3K/Akt signaling pathway.

  16. Integrin αIIb-Mediated PI3K/Akt Activation in Platelets

    PubMed Central

    Niu, Haixia; Chen, Xue; Gruppo, Ralph A.; Li, Ding; Wang, Yanhua; Zhang, Lin; Wang, Kemin; Chai, Weiran; Sun, Yueping; Ding, Zhongren; Gartner, T. Kent; Liu, Junling

    2012-01-01

    Integrin αIIbβ3 mediated bidirectional signaling plays a critical role in thrombosis and haemostasis. Signaling mediated by the β3 subunit has been extensively studied, but αIIb mediated signaling has not been characterized. Previously, we reported that platelet granule secretion and TxA2 production induced by αIIb mediated outside-in signaling is negatively regulated by the β3 cytoplasmic domain residues R724KEFAKFEEER734. In this study, we identified part of the signaling pathway utilized by αIIb mediated outside-in signaling. Platelets from humans and gene deficient mice, and genetically modified CHO cells as well as a variety of kinase inhibitors were used for this work. We found that aggregation of TxA2 production and granule secretion by β3Δ724 human platelets initiated by αIIb mediated outside-in signaling was inhibited by the Src family kinase inhibitor PP2 and the PI3K inhibitor wortmannin, respectively, but not by the MAPK inhibitor U0126. Also, PP2 and wortmannin, and the palmitoylated β3 peptide R724KEFAKFEEER734, each inhibited the phosphorylation of Akt residue Ser473 and prevented TxA2 production and storage granule secretion. Similarly, Akt phosphorylation in mouse platelets stimulated by the PAR4 agonist peptide AYPGKF was αIIbβ3-dependent, and blocked by PP2, wortmannin and the palmitoylated peptide p-RKEFAKFEEER. Akt was also phosphorylated in response to mAb D3 plus Fg treatment of CHO cells in suspension expressing αIIbβ3-Δ724 or αIIbβ3E724AERKFERKFE734, but not in cells expressing wild type αIIbβ3. In summary, SFK(s) and PI3K/Akt signaling is utilized by αIIb-mediated outside-in signaling to activate platelets even in the absence of all but 8 membrane proximal residues of the β3 cytoplasmic domain. Our results provide new insight into the signaling pathway used by αIIb-mediated outside-in signaling in platelets. PMID:23082158

  17. Astrocyte-conditioned medium protecting hippocampal neurons in primary cultures against corticosterone-induced damages via PI3-K/Akt signal pathway.

    PubMed

    Zhu, Ze-Hua; Yang, Ru; Fu, Xin; Wang, Yan-Qing; Wu, Gen-Cheng

    2006-10-09

    Prolonged or excessive exposure to corticosterone leads to neuronal damages in the brain regions, including hippocampus. We reported that astrocyte-conditioned medium (ACM) protected the neurons of the primary hippocampal cultures against the corticosterone-induced damages. Corticosterone added to the cultures resulted in a significant number of TUNEL-positive cells. However, corticosterone-induced TUNEL labeling was suppressed as for ACM-cultured neurons. To delineate the molecular basis underlying the neuroprotection of ACM, we assessed the activation of ERK1/2 and (PI3-K)/Akt signal pathways in response to corticosterone-induced neuronal damages. Western blot test revealed that corticosterone increased the phosphorylation of ERK1/2 and PI3-K/Akt in hippocampal neurons grown in Neurobasal medium supplemented with B27 and 500 microm L-glutamine (NBM+). Interestingly, the increase of phospho-ERK1/2 and Akt levels was much pronounced and the time course of phosphorylation was altered in ACM, suggesting that both signaling pathways might participate in ACM protection. Furthermore, the selective inhibitor of Akt, rather than ERK1/2, blocked the neuroprotective activity against corticosterone in ACM-cultured neurons. In summary, our data showed that ACM had a potent neuroprotective effect in cultured neurons. PI3-K/Akt signal pathway, but not ERK1/2, was involved in the protective activity against the corticosterone-induced damages.

  18. Quercetin regresses Dalton's lymphoma growth via suppression of PI3K/AKT signaling leading to upregulation of p53 and decrease in energy metabolism.

    PubMed

    Maurya, Akhilendra Kumar; Vinayak, Manjula

    2015-01-01

    Various oncogenes are associated with deregulation in cell proliferation, apoptosis, and cell survival, which ultimately cause cancerous growth. Phosphatidylinositol 3-kinase (PI3K) mediated signaling plays a key role in malignant transformation. Cell proliferation and cell survival of tumor cell are induced by hyper activation of PI3K, AKT1, glycolytic enzyme LDH-A, and inactivation of tumor suppressor gene p53. Dietary flavonoids such as quercetin are considered a powerful modulator of different cellular signaling pathways. The present study is focused on the role of quercetin on regulation of PI3K/AKT pathways in Dalton's lymphoma mice. Effect of quercetin was analyzed in ascite cells in terms of cell viability, glycolytic metabolism as well as expression, and level of PI3K (regulatory and catalytic subunit), AKT1, and p53 using standard methods. Results reflect hyperactivation of PI3K signaling in ascite cells of Dalton's lymphoma mice, leading to activation of AKT1 and inactivation of p53. Quercetin modulates the pathway toward suppression of lymphoma. Glycolytic metabolism was also downregulated by quercetin. Its tumor suppressor activity was confirmed by morphological parameters and longevity of mice. The findings suggest that quercetin may contribute to lymphoma prevention by downregulating PI3K-AKT1-p53 pathway as well as by glycolytic metabolism.

  19. Obesity Increases Vascular Senescence and Susceptibility to Ischemic Injury Through Chronic Activation of Akt and mTOR

    PubMed Central

    Wang, Chao-Yung; Kim, Hyung-Hwan; Hiroi, Yukio; Sawada, Naoki; Salomone, Salvatore; Benjamin, Laura E.; Walsh, Kenneth; Moskowitz, Michael A.; Liao, James K.

    2009-01-01

    Obesity and age are important risk factors for cardiovascular disease. However, the signaling mechanism linking obesity with age-related vascular senescence is unknown. Here we show that mice fed a high-fat diet show increased vascular senescence and vascular dysfunction compared to mice fed standard chow and are more prone to peripheral and cerebral ischemia. All of these changes involve long-term activation of the protein kinase Akt. In contrast, mice with diet-induced obesity that lack Akt1 are resistant to vascular senescence. Rapamycin treatment of diet-induced obese mice or of transgenic mice with long-term activation of endothelial Akt inhibits activation of mammalian target of rapamycin (mTOR)–rictor complex 2 and Akt, prevents vascular senescence without altering body weight, and reduces the severity of limb necrosis and ischemic stroke. These findings indicate that long-term activation of Akt-mTOR signaling links diet-induced obesity with vascular senescence and cardiovascular disease. PMID:19293429

  20. Endomembrane PtdIns(3,4,5)P3 activates the PI3K-Akt pathway.

    PubMed

    Jethwa, Nirmal; Chung, Gary H C; Lete, Marta G; Alonso, Alicia; Byrne, Richard D; Calleja, Véronique; Larijani, Banafshé

    2015-09-15

    PKB/Akt activation is a common step in tumour growth, proliferation and survival. Akt activation is understood to occur at the plasma membrane of cells in response to growth factor stimulation and local production of the phosphoinositide lipid phosphatidylinositol (3,4,5)-trisphosphate [PtdIns(3,4,5)P3] following phosphoinositide 3-kinase (PI3K) activation. The metabolism and turnover of phosphoinositides is complex--they act as signalling molecules as well as structural components of biological membranes. The localisation and significance of internal pools of PtdIns(3,4,5)P3 has long been speculated upon. By using transfected and recombinant protein probes for PtdIns(3,4,5)P3, we show that PtdIns(3,4,5)P3 is enriched in the nuclear envelope and early endosomes. By exploiting an inducible dimerisation device to recruit Akt to these compartments, we demonstrate that Akt can be locally activated in a PtdIns(3,4,5)P3-dependent manner and has the potential to phosphorylate compartmentally localised downstream substrates. This could be an important mechanism to regulate Akt isoform substrate specificity or influence the timing and duration of PI3K pathway signalling. Defects in phosphoinositide metabolism and localisation are known to contribute to cancer, suggesting that interactions at subcellular compartments might be worthwhile targets for therapeutic intervention.

  1. DMH1 increases glucose metabolism through activating Akt in L6 rat skeletal muscle cells.

    PubMed

    Xie, Xin; Xu, Xiao-Ming; Li, Na; Zhang, Yong-Hui; Zhao, Yu; Ma, Chun-Yan; Dong, De-Li

    2014-01-01

    DMH1(4-[6-(4-Isopropoxyphenyl)pyrazolo [1,5-a]pyrimidin-3-yl] quinoline) is a compound C analogue with the structural modifications at the 3- and 6-positions in pyrazolo[1,5-a]pyrimidine backbone. Compound C was reported to inhibit both AMPK and Akt. Our preliminary work found that DMH1 activated Akt. Since Akt was involved in glucose metabolism, we aimed to identify the effects of DMH1 on glucose metabolism in L6 rat muscle cells and the potential mechanism. Results showed that DMH1 increased lactic acid release and glucose consumption in L6 rat muscle cells in a dose-dependent manner. DMH1 activated Akt in L6 cells. Akt inhibitor inhibited DMH1-induced Akt activation and DMH1-induced increases of glucose uptake and consumption. DMH1 had no cytotoxicity in L6 cells, but inhibited mitochondrial function and reduced ATP production. DMH1 showed no effect on AMPK, but in the presence of Akt inhibitor, DMH1 significantly activated AMPK. Compound C inhibited DMH1-induced Akt activation in L6 cells. Compound C inhibited DMH1-induced increase of glucose uptake, consumption and lactic acid release in L6 cells. DMH1 inhibited PP2A activity, and PP2A activator forskolin reversed DMH1-induced Akt activation. We concluded that DMH1 increased glucose metabolism through activating Akt and DMH1 activated Akt through inhibiting PP2A activity in L6 rat muscle cells. In view of the analogue structure of DMH1 and compound C and the contrasting effects of DMH1 and compound C on Akt, the present study provides a novel leading chemical structure targeting Akt with potential use for regulating glucose metabolism.

  2. Distinct roles of AKT isoforms in regulating β1-integrin activity, migration, and invasion in prostate cancer

    PubMed Central

    Virtakoivu, Reetta; Pellinen, Teijo; Rantala, Juha K.; Perälä, Merja; Ivaska, Johanna

    2012-01-01

    AKT1 and AKT2 kinases have been shown to play opposite roles in breast cancer migration and invasion. In this study, an RNA interference screen for integrin activity inhibitors identified AKT1 as an inhibitor of β1-integrin activity in prostate cancer. Validation experiments investigating all three AKT isoforms demonstrated that, unlike in breast cancer, both AKT1 and AKT2 function as negative regulators of cell migration and invasion in PC3 prostate cancer cells. Down-regulation of AKT1 and AKT2, but not AKT3, induced activation of cell surface β1-integrins and enhanced adhesion, migration, and invasion. Silencing of AKT1 and AKT2 also resulted in increased focal adhesion size. Importantly, the mechanisms involved in integrin activity regulation were distinct for the two AKT isoforms. Silencing of AKT1 relieved feedback suppression of the expression and activity of several receptor tyrosine kinases, including EGFR and MET, with established cross-talk with β1-integrins. Silencing of AKT2, on the other hand, induced up-regulation of the microRNA-200 (miR-200) family, and overexpression of miR-200 was sufficient to induce integrin activity and cell migration in PC3 cells. Taken together, these data define an inhibitory role for both AKT1 and AKT2 in prostate cancer migration and invasion and highlight the cell type–specific actions of AKT kinases in the regulation of cell motility. PMID:22809628

  3. Histone Deacetylase Inhibitors Inhibit the Proliferation of Gallbladder Carcinoma Cells by Suppressing AKT/mTOR Signaling.

    PubMed

    Zhang, Peng; Guo, Zhiyong; Wu, Ying; Hu, Ronglin; Du, Jun; He, Xiaoshun; Jiao, Xingyuan; Zhu, Xiaofeng

    2015-01-01

    Gallbladder carcinoma is an aggressive malignancy with high mortality mainly due to the limited potential for curative resection and its resistance to chemotherapeutic agents. Here, we show that the histone deacetylase inhibitors (HDACIs) trichostatin-A (TSA) and suberoylanilide hydroxamic acid (SAHA) reduce the proliferation and induce apoptosis of gallbladder carcinoma cells by suppressing the AKT/mammalian target of rapamycin (mTOR) signaling. Gallbladder carcinoma SGC-996 cells were treated with different concentrations of TSA and SAHA for different lengths of time. Cell proliferation and morphology were assessed with MTT assay and microscopy, respectively. Cell cycle distribution and cell apoptosis were analyzed with flow cytometry. Western blotting was used to detect the proteins related to apoptosis, cell cycle, and the AKT/mTOR signaling pathway. Our data showed that TSA and SAHA reduced SGC-996 cell viability and arrested cell cycle at the G1 phase in a dose- and time-dependent manner. TSA and SAHA promoted apoptosis of SGC-996 cells, down-regulated the expression of cyclin D1, c-Myc and Bmi1, and decreased the phosphorylation of AKT, mTOR p70S6K1, S6 and 4E-BP1. Additionally, the mTOR inhibitor rapamycin further reduced the cell viability of TSA- and SAHA-treated SGC-996 cells and the phosphorylation of mTOR, whereas the mTOR activator 1,2-dioctanoyl-sn-glycero-3-phosphate (C8-PA) exerted the opposite influence. Our results demonstrate that histone deacetylase inhibitors (HDACIs) suppress the proliferation of gallbladder carcinoma cell via inhibition of AKT/mTOR signaling. These findings offer a mechanistic rationale for the application of HDACIs in gallbladder carcinoma treatment.

  4. Tetrandrine inhibits migration and invasion of rheumatoid arthritis fibroblast-like synoviocytes through down-regulating the expressions of Rac1, Cdc42, and RhoA GTPases and activation of the PI3K/Akt and JNK signaling pathways.

    PubMed

    Lv, Qi; Zhu, Xian-Yang; Xia, Yu-Feng; Dai, Yue; Wei, Zhi-Feng

    2015-11-01

    Tetrandrine (Tet), the main active constituent of Stephania tetrandra root, has been demonstrated to alleviate adjuvant-induced arthritis in rats. The present study was designed to investigate the effects of Tet on the migration and invasion of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS) and explore the underlying mechanisms. By using cultures of primary FLS isolated from synoviums of RA patients and cell line MH7A, Tet (0.3, 1 μmol·L(-1)) was proven to significantly impede migration and invasion of RA-FLS, but not cell proliferation. Tet also greatly reduced the activation and expressions of matrix degrading enzymes MMP-2/9, the expression of F-actin and the activation of FAK, which controlled the morphologic changes in migration process of FLS. To identify the key signaling pathways by which Tet exerts anti-migration effect, the specific inhibitors of multiple signaling pathways LY294002, Triciribine, SP600125, U0126, SB203580, and PDTC (against PI3K, Akt, JNK, ERK, p38 MAPK and NF-κB-p65, respectively) were used. Among them, LY294002, Triciribine, and SP600125 were shown to obviously inhibit the migration of MH7A cells. Consistently, Tet was able to down-regulate the activation of Akt and JNK as demonstrated by Western blotting assay. Moreover, Tet could reduce the expressions of migration-related proteins Rho GTPases Rac1, Cdc42, and RhoA in MH7A cells. In conclusion, Tet can impede the migration and invasion of RA-FLS, which provides a plausible explanation for its protective effect on RA. The underlying mechanisms involve the reduction of the expressions of Rac1, Cdc42, and RhoA, inhibition of the activation of Akt and JNK, and subsequent down-regulation of activation and/or expressions of MMP-2/9, F-actin, and FAK.

  5. Activating Akt1 mutations alter DNA double strand break repair and radiosensitivity.

    PubMed

    Oeck, S; Al-Refae, K; Riffkin, H; Wiel, G; Handrick, R; Klein, D; Iliakis, G; Jendrossek, V

    2017-02-17

    The survival kinase Akt has clinical relevance to radioresistance. However, its contributions to the DNA damage response, DNA double strand break (DSB) repair and apoptosis remain poorly defined and often contradictory. We used a genetic approach to explore the consequences of genetic alterations of Akt1 for the cellular radiation response. While two activation-associated mutants with prominent nuclear access, the phospho-mimicking Akt1-TDSD and the clinically relevant PH-domain mutation Akt1-E17K, accelerated DSB repair and improved survival of irradiated Tramp-C1 murine prostate cancer cells and Akt1-knockout murine embryonic fibroblasts in vitro, the classical constitutively active membrane-targeted myrAkt1 mutant had the opposite effects. Interestingly, DNA-PKcs directly phosphorylated Akt1 at S473 in an in vitro kinase assay but not vice-versa. Pharmacological inhibition of DNA-PKcs or Akt restored radiosensitivity in tumour cells expressing Akt1-E17K or Akt1-TDSD. In conclusion, Akt1-mediated radioresistance depends on its activation state and nuclear localization and is accessible to pharmacologic inhibition.

  6. Activating Akt1 mutations alter DNA double strand break repair and radiosensitivity

    PubMed Central

    Oeck, S.; Al-Refae, K.; Riffkin, H.; Wiel, G.; Handrick, R.; Klein, D.; Iliakis, G.; Jendrossek, V.

    2017-01-01

    The survival kinase Akt has clinical relevance to radioresistance. However, its contributions to the DNA damage response, DNA double strand break (DSB) repair and apoptosis remain poorly defined and often contradictory. We used a genetic approach to explore the consequences of genetic alterations of Akt1 for the cellular radiation response. While two activation-associated mutants with prominent nuclear access, the phospho-mimicking Akt1-TDSD and the clinically relevant PH-domain mutation Akt1-E17K, accelerated DSB repair and improved survival of irradiated Tramp-C1 murine prostate cancer cells and Akt1-knockout murine embryonic fibroblasts in vitro, the classical constitutively active membrane-targeted myrAkt1 mutant had the opposite effects. Interestingly, DNA-PKcs directly phosphorylated Akt1 at S473 in an in vitro kinase assay but not vice-versa. Pharmacological inhibition of DNA-PKcs or Akt restored radiosensitivity in tumour cells expressing Akt1-E17K or Akt1-TDSD. In conclusion, Akt1-mediated radioresistance depends on its activation state and nuclear localization and is accessible to pharmacologic inhibition. PMID:28209968

  7. Upregulation of AKT1 protein expression in forskolin-stimulated macrophage: evidence from ChIP analysis that CREB binds to and activates the AKT1 promoter.

    PubMed

    Misra, Uma Kant; Pizzo, Salvatore Vincent

    2007-03-01

    Recently, we reported that silencing CREB gene expression by RNAi significantly attenuates forskolin-induced activation of Akt1. We now provide evidence that forskolin-treatment causes transcriptional and translational upregulation of Akt1 in macrophages. Akt synthesis was demonstrated by [(14)C]leucine or [(35)S] incorporation into newly synthesized Akt1 protein. Akt protein levels increased by approximately 1.5-fold after only a 5 min exposure of macrophages to forskolin. Akt1 levels thereafter rapidly returned to basal values (t(1/2) approximately 15 min). Maximal upregulation of Akt1 occurred in cells treated with 10 microM forskolin. Forskolin-dependent Akt1 synthesis was abolished by pretreating the cells with CREB-directed dsRNA as demonstrated at both the message and protein level, as well as by determining the synthesis of [(35)S]-labeled Akt1 protein. The PKA inhibitor H-89, greatly attenuated forskolin-induced Akt1 synthesis. Transcriptional and translational inhibitors also greatly reduced Akt1 synthesis in forskolin-stimulated [(14)C]leucine-labeled macrophages. Using a chromatin immunoprecipitation assay, we demonstrate that CREB binds to a CRE binding domain of the Akt1 gene promoter. In conclusion, we show here for the first time transcriptional upregulation of Akt1 by CREB, based upon Akt1 protein synthesis and its modulation by transitional and translational inhibitors in forskolin-stimulated cells, Akt1 protein. and mRNA levels upon silencing CREB gene expression, and binding of CREB to the Akt1 gene promoter.

  8. Atorvastatin calcium inhibits phenotypic modulation of PDGF-BB-induced VSMCs via down-regulation the Akt signaling pathway.

    PubMed

    Chen, Shuang; Liu, Baoqin; Kong, Dehui; Li, Si; Li, Chao; Wang, Huaqin; Sun, Yingxian

    2015-01-01

    Plasticity of vascular smooth muscle cells (VSMCs) plays a central role in the onset and progression of proliferative vascular diseases. In adult tissue, VSMCs exist in a physiological contractile-quiescent phenotype, which is defined by lack of the ability of proliferation and migration, while high expression of contractile marker proteins. After injury to the vessel, VSMC shifts from a contractile phenotype to a pathological synthetic phenotype, associated with increased proliferation, migration and matrix secretion. It has been demonstrated that PDGF-BB is a critical mediator of VSMCs phenotypic switch. Atorvastatin calcium, a selective inhibitor of 3-hydroxy-3-methyl-glutaryl l coenzyme A (HMG-CoA) reductase, exhibits various protective effects against VSMCs. In this study, we investigated the effects of atorvastatin calcium on phenotype modulation of PDGF-BB-induced VSMCs and the related intracellular signal transduction pathways. Treatment of VSMCs with atorvastatin calcium showed dose-dependent inhibition of PDGF-BB-induced proliferation. Atorvastatin calcium co-treatment inhibited the phenotype modulation and cytoskeleton rearrangements and improved the expression of contractile phenotype marker proteins such as α-SM actin, SM22α and calponin in comparison with PDGF-BB alone stimulated VSMCs. Although Akt phosphorylation was strongly elicited by PDGF-BB, Akt activation was attenuated when PDGF-BB was co-administrated with atorvastatin calcium. In conclusion, atorvastatin calcium inhibits phenotype modulation of PDGF-BB-induced VSMCs and activation of the Akt signaling pathway, indicating that Akt might play a vital role in the modulation of phenotype.

  9. Activating PTEN by COX-2 inhibitors antagonizes radiation-induced AKT activation contributing to radiosensitization

    SciTech Connect

    Meng, Zhen; Gan, Ye-Hua

    2015-05-01

    Radiotherapy is still one of the most effective nonsurgical treatments for many tumors. However, radioresistance remains a major impediment to radiotherapy. Although COX-2 inhibitors can induce radiosensitization, the underlying mechanism is not fully understood. In this study, we showed that COX-2 selective inhibitor celecoxib enhanced the radiation-induced inhibition of cell proliferation and apoptosis in HeLa and SACC-83 cells. Treatment with celecoxib alone dephosphorylated phosphatase and tensin homolog deleted on chromosome ten (PTEN), promoted PTEN membrane translocation or activation, and correspondingly dephosphorylated or inactivated protein kinase B (AKT). By contrast, treatment with radiation alone increased PTEN phosphorylation, inhibited PTEN membrane translocation and correspondingly activated AKT in the two cell lines. However, treatment with celecoxib or another COX-2 selective inhibitor (valdecoxib) completely blocked radiation-induced increase of PTEN phosphorylation, rescued radiation-induced decrease in PTEN membrane translocation, and correspondingly inactivated AKT. Moreover, celecoxib could also upregulate PTEN protein expression by downregulating Sp1 expression, thereby leading to the activation of PTEN transcription. Our results suggested that COX-2 inhibitors could enhance radiosensitization at least partially by activating PTEN to antagonize radiation-induced AKT activation. - Highlights: • COX-2 inhibitor, celecoxib, could enhance radiosensitization. • Radiation induced PTEN inactivation (phosphorylation) and AKT activation. • COX-2 inhibitor induced PTEN expression and activation, and inactivated AKT. • COX-2 inhibitor enhanced radiosensitization through activating PTEN.

  10. Nuclear Akt2 opposes limbal keratinocyte stem cell self-renewal by repressing a FOXO-mTORC1 signaling pathway.

    PubMed

    Saoncella, Stefania; Tassone, Beatrice; Deklic, Erika; Avolio, Fabio; Jon, Cristina; Tornillo, Giusy; De Luca, Elisa; Di Iorio, Enzo; Piva, Roberto; Cabodi, Sara; Turco, Emilia; Pandolfi, Pier Paolo; Calautti, Enzo

    2014-03-01

    Signals downstream of Akt can either favor or oppose stem cell (SC) maintenance, but how this dual role can be achieved is still undefined. Using human limbal keratinocyte stem cells (LKSCs), a SC type used in transplantation therapies for corneal regeneration, we show that Akt signaling is prominent in SC populations both in vivo and in vitro, and that Akt1 promotes while Akt2 opposes SC self-renewal. Noteworthy, loss of Akt2 signaling enhances LKSC maintenance ex vivo, whereas Akt1 depletion anticipates SC exhaustion. Mechanistically, the antagonistic functions of Akt1 and Akt2 in SC control are mainly dictated by their differential subcellular distribution, being nuclear Akt2 selectively implicated in FOXO inhibition. Akt2 downregulation favors LKSC maintenance as a result of a gain of FOXO functions, which attenuates the mechanistic target of rapamycin complex one signaling via tuberous sclerosis one gene induction, and promotes growth factor signaling through Akt1. Consistently, Akt2 deficiency also enhances limbal SCs in vivo. Thus, our findings reveal distinct roles for nuclear versus cytosolic Akt signaling in normal epithelial SC control and suggest that the selective Akt2 inhibition may provide novel pharmacological strategies for human LKSC expansion in therapeutic settings and mechanistic research.

  11. Both mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 and phosphatidylinositide-3-OH kinase (PI3K)/Akt pathways regulate activation of E-twenty-six (ETS)-like transcription factor 1 (Elk-1) in U138 glioblastoma cells.

    PubMed

    Mut, Melike; Lule, Sevda; Demir, Ozlem; Kurnaz, Isil Aksan; Vural, Imran

    2012-02-01

    Epidermal growth factor (EGF) and its receptor (EGFR) have been shown to play a significant role in the pathogenesis of glioblastoma. In our study, the EGFR was stimulated with EGF in human U138 glioblastoma cells. We show that the activated mitogen-activated protein kinase (MAPK)/extracellular-signal-regulated kinases (ERK) 1/2 pathway phosphorylated the E twenty-six (ETS)-like transcription factor 1 (Elk-1) mainly at serine 383 residue. Mitogen-activated protein kinase kinase (MEK) 1/2 inhibitor, UO126 and ERK inhibitor II, FR180204 blocked the Elk-1 phosphorylation and activation. The phosphatidylinositide-3-OH kinase (PI3K)/Akt pathway was also involved in the Elk-1 activation. Activation of the Elk-1 led to an increased survival and a proliferative response with the EGF stimulation in the U138 glioblastoma cells. Knocking-down the Elk-1 using an RNA interference technique caused a decrease in survival of the unstimulated U138 glioblastoma cells and also decreased the proliferative response to the EGF stimulation. The Elk-1 transcription factor was important for the survival and proliferation of U138 glioblastoma cells upon the stimulation of EGFR with EGF. The MAPK/ERK1/2 and PI3K/Akt pathways regulated this response via activation of the Elk-1 transcription factor. The Elk-1 may be one of the convergence points for pathways located downstream of EGFR in glioblastoma cells. Utilization of the Elk-1 as a therapeutic target may lead to a novel strategy in treatment of glioblastoma.

  12. Berberine protects endothelial progenitor cell from damage of TNF-α via the PI3K/AKT/eNOS signaling pathway.

    PubMed

    Xiao, Min; Men, Li Na; Xu, Ming Guo; Wang, Guo Bing; Lv, Hai Tao; Liu, Cong

    2014-11-15

    Endothelial progenitor cells (EPCs) dysfunction is closely correlated with the coronary artery injury induced by Kawasaki disease (KD). The level of tumor necrosis factor-α (TNF-α) elevated significantly in acute phase of KD which can damage the functions of EPCs. The aim of this study was to investigate whether berberine (BBR) can protect EPCs from the inhibition caused by TNF-α via the PI3K (Phosphatidyl Inositol 3-kinase) /AKT (Serine/threonine protein kinase B) /eNOS (endothelial Nitric Oxide synthase) signaling pathway. The cell proliferative ability of EPCs was determined by MTT (methyl thiazolyl tetrazolium) assays. Nitric oxide (NO) level was determined in supernatants. The mRNA level of eNOS, PI3K and AKT were measured by Real Time-Polymerase Chain Reaction (RT-PCR), and the protein levels of eNOS, phospho-eNOS (p-eNOS), Akt, phospho-Akt (p-Akt) and PI3K were analyzed using Western-blot. The results demonstrated that TNF-α inhibits the proliferative ability of EPCs. However, BBR improves the proliferative activity of EPCs inhibited by TNF-α. Blockade of PI3K by 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (Ly294002) and blockade of eNOS by l-NAME (NG-Nitroarginine Methyl Ester) attenuates the effect of BBR. BBR can increase the level of PI3K/Akt/eNOS mRNA and the protein level of PI3K, p-Akt, eNOS and p-eNOS, which can be blocked by PI3K inhibitor (LY294002) and eNOS inhibitor (l-NAME). Therefore, we concluded that impaired EPCs proliferation could be reversed by BBR via the PI3K/AKT/eNOS signaling pathway.

  13. Pancreas-specific Pten deficiency causes partial resistance to diabetes and elevated hepatic AKT signaling.

    PubMed

    Tong, Zan; Fan, Yan; Zhang, Weiqi; Xu, Jun; Cheng, Jing; Ding, Mingxiao; Deng, Hongkui

    2009-06-01

    PTEN, a negative regulator of the phosphatidylinositol-3-kinase/AKT pathway, is an important modulator of insulin signaling. To determine the metabolic function of pancreatic Pten, we generated pancreas-specific Pten knockout (PPKO) mice. PPKO mice had enlarged pancreas and elevated proliferation of acinar cells. They also exhibited hypoglycemia, hypoinsulinemia, and altered amino metabolism. Notably, PPKO mice showed delayed onset of streptozotocin (STZ)-induced diabetes and sex-biased resistance to high-fat-diet (HFD)-induced diabetes. To investigate the mechanism for the resistance to HFD-induced hyperglycemia in PPKO mice, we evaluated AKT phosphorylation in major insulin-responsive tissues: the liver, muscle, and fat. We found that Pten loss in the pancreas causes the elevation of AKT signaling in the liver. The phosphorylation of AKT and its downstream substrate GSK3beta was increased in the liver of PPKO mice, while PTEN level was decreased without detectable excision of Pten allele in the liver of PPKO mice. Proteomics analysis revealed dramatically decreased level of 78-kDa glucose-regulated protein (GRP78) in the liver of PPKO mice, which may also contribute to the lower blood glucose level of PPKO mice fed with HFD. Together, our findings reveal a novel response in the liver to pancreatic defect in metabolic regulation, adding a new dimension to understanding diabetes resistance.

  14. Modification of Caffeic Acid with Pyrrolidine Enhances Antioxidant Ability by Activating AKT/HO-1 Pathway in Heart

    PubMed Central

    Ku, Hui-Chun; Lee, Shih-Yi; Yang, Kai-Chien; Kuo, Yueh-Hsiung; Su, Ming-Jai

    2016-01-01

    Overproduction of free radicals during ischemia/reperfusion (I/R) injury leads to an interest in using antioxidant therapy. Activating an endogenous antioxidant signaling pathway is more important due to the fact that the free radical scavenging behavior in vitro does not always correlate with a cytoprotection effect in vivo. Caffeic acid (CA), an antioxidant, is a major phenolic constituent in nature. Pyrrolidinyl caffeamide (PLCA), a derivative of CA, was compared with CA for their antioxidant and cytoprotective effects. Our results indicate that CA and PLCA exert the same ability to scavenge DPPH in vitro. In response to myocardial I/R stress, PLCA was shown to attenuate lipid peroxydation and troponin release more than CA. These responses were accompanied with a prominent elevation in AKT and HO-1 expression and a preservation of mnSOD expression and catalase activity. PLCA also improved cell viability and alleviated the intracellular ROS level more than CA in cardiomyocytes exposed to H2O2. When inhibiting the AKT or HO-1 pathways, PLCA lost its ability to recover mnSOD expression and catalase activity to counteract with oxidative stress, suggesting AKT/HO-1 pathway activation by PLCA plays an important role. In addition, inhibition of AKT signaling further abolished HO-1 activity, while inhibition of HO-1 signaling attenuated AKT expression, indicating cross-talk between the AKT and HO-1 pathways. These protective effects may contribute to the cardiac function improvement by PLCA. These findings provide new insight into therapeutic approaches using a modified natural compound against oxidative stress from myocardial injuries. PMID:26845693

  15. Exclusive inhibition of PI3K/Akt/mTOR signaling is not sufficient to prevent PDGF-mediated effects on glycolysis and proliferation in colorectal cancer.

    PubMed

    Moench, Romana; Grimmig, Tanja; Kannen, Vinicius; Tripathi, Sudipta; Faber, Marc; Moll, Eva-Maria; Chandraker, Anil; Lissner, Reinhard; Germer, Christoph-Thomas; Waaga-Gasser, Ana Maria; Gasser, Martin

    2016-10-18

    Platelet-derived growth factor (PDGF) and signaling via its receptors plays a crucial role in tumor cell proliferation and thus may represent an attractive target besides VEGF/EGFR-based antibody therapies. In this study we analyzed the influence of PDGF in colorectal cancer. PDGF was expressed intensively in early and even more intensively in late stage primary CRCs. Like VEGF, PDGF enhanced human colon cancer proliferation, and increased oxidative glycolytic activity, and activated HIF1α and c-Myc in vitro. PDGF activated the PI3K/Akt/mTOR pathway while leaving MAPK signaling untouched. Further dissection showed that inhibition of Akt strongly impeded cancer cell growth while inhibition of PI3K did not. MAPK analysis suggested an inhibitory crosstalk between both pathways, thus explaining the different effects of the Akt and PI3K inhibitors on cancer cell proliferation. PDGF stimulates colon cancer cell proliferation, and prevents inhibitor induced apoptosis, resulting in tumor growth. Therefore inhibition of PDGF signaling seems to be a promising target in colorectal cancer therapy. However, due to the multifaceted nature of the intracellular PDGF signaling, careful intervention strategies are needed when looking into specific signaling pathways like PI3K/Akt/mTOR and MAPK.

  16. Exclusive inhibition of PI3K/Akt/mTOR signaling is not sufficient to prevent PDGF-mediated effects on glycolysis and proliferation in colorectal cancer

    PubMed Central

    Moench, Romana; Grimmig, Tanja; Kannen, Vinicius; Tripathi, Sudipta; Faber, Marc; Moll, Eva-Maria; Chandraker, Anil; Lissner, Reinhard; Germer, Christoph-Thomas

    2016-01-01

    Platelet-derived growth factor (PDGF) and signaling via its receptors plays a crucial role in tumor cell proliferation and thus may represent an attractive target besides VEGF/EGFR-based antibody therapies. In this study we analyzed the influence of PDGF in colorectal cancer. PDGF was expressed intensively in early and even more intensively in late stage primary CRCs. Like VEGF, PDGF enhanced human colon cancer proliferation, and increased oxidative glycolytic activity, and activated HIF1α and c-Myc in vitro. PDGF activated the PI3K/Akt/mTOR pathway while leaving MAPK signaling untouched. Further dissection showed that inhibition of Akt strongly impeded cancer cell growth while inhibition of PI3K did not. MAPK analysis suggested an inhibitory crosstalk between both pathways, thus explaining the different effects of the Akt and PI3K inhibitors on cancer cell proliferation. PDGF stimulates colon cancer cell proliferation, and prevents inhibitor induced apoptosis, resulting in tumor growth. Therefore inhibition of PDGF signaling seems to be a promising target in colorectal cancer therapy. However, due to the multifaceted nature of the intracellular PDGF signaling, careful intervention strategies are needed when looking into specific signaling pathways like PI3K/Akt/mTOR and MAPK. PMID:27626684

  17. Molecular characterization of the Akt-TOR signaling pathway in rainbow trout: potential role in muscle growth/degradation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Akt-TOR signaling pathway plays a key role in cellular metabolism and muscle growth. Hormone, nutrition and stress factors affect the Akt-TOR pathway by regulating gene transcription, protein synthesis and degradation. In addition, we previously showed that energetic demands elevate during vit...

  18. Influence of zinc deficiency on AKT-MDM2-P53 signaling axes in normal and malignant human prostate cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    With prostate being the highest zinc-accumulating tissue before the onset of cancer, the effects of physiologic levels of zinc on Akt-Mdm2-p53 and Akt-p21 signaling axes in human normal prostate epithelial cells (PrEC) and malignant prostate LNCaP cells were examined. Cells were cultured for 6 d in...

  19. Oncogenic activation of the PI3K/Akt pathway promotes cellular glucose uptake by downregulating the expression of thioredoxin-interacting protein.

    PubMed

    Hong, Shin Yee; Yu, Fa-Xing; Luo, Yan; Hagen, Thilo

    2016-05-01

    Oncogenic activation of the PI3K/Akt pathway is known to play an important role to promote glucose metabolism in cancer cells. However, the molecular mechanism through which the PI3K/Akt signalling pathway promotes glucose utilisation in cancer cells is still not well understood. It has recently been shown that the oncogenic activation of the PI3K/Akt/mTOR signalling in lung adenocarcinoma is important in promoting the localisation of glucose transporter 1 (GLUT1) at the plasma membrane. We thus hypothesised that the effect of constitutive activation of the PI3K/AKT signalling on glucose metabolism is mediated by thioredoxin interacting protein (TXNIP), a known regulator of the GLUT1 plasma membrane localisation. Consistent with pr