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Sample records for 3-kinase pi3k signalling

  1. Icaritin requires Phosphatidylinositol 3 kinase (PI3K)/Akt signaling to counteract skeletal muscle atrophy following mechanical unloading

    PubMed Central

    ZHANG, Zong-Kang; LI, Jie; LIU, Jin; GUO, Baosheng; LEUNG, Albert; ZHANG, Ge; ZHANG, Bao-Ting

    2016-01-01

    Counteracting muscle atrophy induced by mechanical unloading/inactivity is of great clinical need and challenge. A therapeutic agent that could counteract muscle atrophy following mechanical unloading in safety is desired. This study showed that natural product Icaritin (ICT) could increase the phosphorylation level of Phosphatidylinositol 3 kinase (PI3K) at p110 catalytic subunit and promote PI3K/Akt signaling markers in C2C12 cells. This study further showed that the high dose ICT treatment could significantly attenuate the decreases in the phosphorylation level of PI3K at p110 catalytic subunit and its downstream markers related to protein synthesis, and inhibit the increases in protein degradation markers at mRNA and protein levels in rat soleus muscle following 28-day hindlimb unloading. In addition, the decreases in soleus muscle mass, muscle fiber cross-sectional area, twitch force, specific force, contraction time and half relaxation time could be significantly attenuated by the high dose ICT treatment. The low dose ICT treatment could moderately attenuate the above changes induced by unloading. Wortmannin, a specific inhibitor of PI3K at p110 catalytic subunit, could abolish the above effects of ICT in vitro and in vivo, indicating that PI3K/Akt signaling could be required by ICT to counteract skeletal muscle atrophy following mechanical unloading. PMID:26831566

  2. 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.

  3. Fine particulate matter leads to reproductive impairment in male rats by overexpressing phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway.

    PubMed

    Cao, Xi-Ning; Yan, Chao; Liu, Dong-Yao; Peng, Jin-Pu; Chen, Jin-Jun; Zhou, Yue; Long, Chun-Lan; He, Da-Wei; Lin, Tao; Shen, Lian-Ju; Wei, Guang-Hui

    2015-09-17

    Maintenance of male reproductive function depends on normal sperm generation during which process Sertoli cells play a vital role. Studies found that fine particulate matter (PM) causes decreased male sperm quality, mechanism of which unestablished. We aim to investigate the definite mechanism of PM impairment on male reproduction. Male Sprague-Dawley rats were daily exposed to normal saline (NS) or PM2.5 with the doses of 9 mg/kg.b.w and 24 mg/kg.b.w. via intratracheal instillation for seven weeks. Reproductive function was tested by mating test and semen analysis after last exposure. Testes were collected to assess changes in histomorphology, and biomarkers including connexin 43 (Cx43), superoxide dismutase (SOD), phosphatidylinositol 3-kinase (PI3K) and phosphorylated protein kinase B (p-Akt). Male rats exposed to PM2.5 showed noticeable decreased fertility, significantly reduced sperm count, increased sperm abnormality rate and severe testicular damage in histomorphology. After PM2.5 exposure, the levels of Cx43 was significantly downregulated, and SOD was upregulated and downregulated significantly with different dose, respectively. Protein expression of PI3K and p-Akt dramatically enhanced, and the later one being located in Sertoli cells, the upward or declining trend was in dose dependent. PM2.5 exposure leads to oxidative stress impairment via PI3K/Akt signaling pathway on male reproduction in rats.

  4. Berberine Induced Apoptosis of Human Osteosarcoma Cells by Inhibiting Phosphoinositide 3 Kinase/Protein Kinase B (PI3K/Akt) Signal Pathway Activation

    PubMed Central

    2016-01-01

    Background: Osteosarcoma is a malignant tumor with high mortality but effective therapy has not yet been developed. Berberine, an isoquinoline alkaloid component in several Chinese herbs including Huanglian, has been shown to induce growth inhibition and the apoptosis of certain cancer cells. The aim of this study was to determine the role of berberine on human osteosarcoma cell lines U2OS and its potential mechanism. Methods: The proliferation effect of U20S was exanimed by 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di- phenytetrazoliumromide (MTT) and the percentage of apoptotic cells were determined by flow cytometric analysis. The expression of PI3K, p-Akt, Bax, Bcl-2, cleavage-PARP and Caspase3 were detected by Western blott. Results: Berberine treatment caused dose-dependent inhibiting proliferation and inducing apoptosis of U20S cell. Mechanistically, berberine inhibits PI3K/AKT activation that, in turn, results in up-regulating the expression of Bax, and PARP and down-regulating the expression of Bcl-2 and caspase3. In all, berberine can suppress the proliferation and induce the apoptosis of U2OS cell through inhibiting the PI3K/Akt signaling pathway activation. Conclusion: Berberine can suppress the proliferation and induce the apoptosis of U2OS cell through inhibiting the PI3K/Akt signaling pathway activation. PMID:27398330

  5. PI3K signaling supports amphetamine-induced dopamine efflux.

    PubMed

    Lute, Brandon J; Khoshbouei, Habibeh; Saunders, Christine; Sen, Namita; Lin, Richard Z; Javitch, Jonathan A; Galli, Aurelio

    2008-08-01

    The dopamine (DA) transporter (DAT) is a major molecular target of the psychostimulant amphetamine (AMPH). AMPH, as a result of its ability to reverse DAT-mediated inward transport of DA, induces DA efflux thereby increasing extracellular DA levels. This increase is thought to underlie the behavioral effects of AMPH. We have demonstrated previously that insulin, through phosphatidylinositol 3-kinase (PI3K) signaling, regulates DA clearance by fine-tuning DAT plasma membrane expression. PI3K signaling may represent a novel mechanism for regulating DA efflux evoked by AMPH, since only active DAT at the plasma membrane can efflux DA. Here, we show in both a heterologous expression system and DA neurons that inhibition of PI3K decreases DAT cell surface expression and, as a consequence, AMPH-induced DA efflux.

  6. E6 variants of human papillomavirus 18 differentially modulate the protein kinase B/phosphatidylinositol 3-kinase (akt/PI3K) signaling pathway

    SciTech Connect

    Contreras-Paredes, Adriana

    2009-01-05

    Intra-type genome variations of high risk Human papillomavirus (HPV) have been associated with a differential threat for cervical cancer development. In this work, the effect of HPV18 E6 isolates in Akt/PKB and Mitogen-associated protein kinase (MAPKs) signaling pathways and its implication in cell proliferation were analyzed. E6 from HPV types 16 and 18 are able to bind and promote degradation of Human disc large (hDlg). Our results show that E6 variants differentially modulate hDlg degradation, rebounding in levels of activated PTEN and PKB. HPV18 E6 variants are also able to upregulate phospho-PI3K protein, strongly correlating with activated MAPKs and cell proliferation. Data was supported by the effect of E6 silencing in HPV18-containing HeLa cells, as well as hDlg silencing in the tested cells. Results suggest that HPV18 intra-type variations may derive in differential abilities to activate cell-signaling pathways such as Akt/PKB and MAPKs, directly involved in cell survival and proliferation.

  7. PI3K/Akt signalling pathway and cancer.

    PubMed

    Fresno Vara, Juan Angel; Casado, Enrique; de Castro, Javier; Cejas, Paloma; Belda-Iniesta, Cristóbal; González-Barón, Manuel

    2004-04-01

    Phosphatidylinositol-3 kinases, PI3Ks, constitute a lipid kinase family characterized by their ability to phosphorylate inositol ring 3'-OH group in inositol phospholipids to generate the second messenger phosphatidylinositol-3,4,5-trisphosphate (PI-3,4,5-P(3)). RPTK activation results in PI(3,4,5)P(3) and PI(3,4)P(2) production by PI3K at the inner side of the plasma membrane. Akt interacts with these phospholipids, causing its translocation to the inner membrane, where it is phosphorylated and activated by PDK1 and PDK2. Activated Akt modulates the function of numerous substrates involved in the regulation of cell survival, cell cycle progression and cellular growth. In recent years, it has been shown that PI3K/Akt signalling pathway components are frequently altered in human cancers. Cancer treatment by chemotherapy and gamma-irradiation kills target cells primarily by the induction of apoptosis. However, the development of resistance to therapy is an important clinical problem. Failure to activate the apoptotic programme represents an important mode of drug resistance in tumor cells. Survival signals induced by several receptors are mediated mainly by PI3K/Akt, hence this pathway may decisively contribute to the resistant phenotype. Many of the signalling pathways involved in cellular transformation have been elucidated and efforts are underway to develop treatment strategies that target these specific signalling molecules or their downstream effectors. The PI3K/Akt pathway is involved in many of the mechanisms targeted by these new drugs, thus a better understanding of this crossroad can help to fully exploit the potential benefits of these new agents. PMID:15023437

  8. PI3K Signaling in B and T Lymphocytes: New Developments and Therapeutic Advances

    PubMed Central

    So, Lomon; Fruman, David A.

    2012-01-01

    Synopsis Activation of phosphoinositide 3-kinase (PI3K) is a shared response to engagement of diverse types of transmembrane receptors. Depending on the cell type and stimulus, PI3K activation can promote different fates including proliferation, survival, migration and differentiation. The diverse roles of PI3K signaling are well illustrated by studies of lymphocytes, the cells that mediate adaptive immunity. Genetic and pharmacological experiments have shown that PI3K activation regulates many steps in the development, activation and differentiation of both B and T cells. These findings have prompted the development of PI3K inhibitors for the treatment of autoimmunity and inflammatory diseases. However, PI3K activation has both positive and negative roles in immune system activation. Consequently, while PI3K suppression can attenuate immune responses it can also enhance inflammation, disrupt peripheral tolerance and promote autoimmunity. An exciting discovery is that a selective inhibitor of the p110δ catalytic isoform of PI3K, CAL-101, achieves impressive clinical efficacy in certain B cell malignancies. A model is emerging in which p110δ inhibition disrupts signals from the lymphoid microenvironment, leading to release of leukemia and lymphoma cells from their protective niche. These encouraging findings have given further momentum to PI3K drug development efforts in both cancer and immune diseases. PMID:22364281

  9. Reactivation of AKT signaling following treatment of cancer cells with PI3K inhibitors attenuates their antitumor effects

    SciTech Connect

    Dufour, Marc; Dormond-Meuwly, Anne; Pythoud, Catherine; Demartines, Nicolas; Dormond, Olivier

    2013-08-16

    Highlights: •PI3K inhibitors inhibit AKT only transiently. •Re-activation of AKT limits the anti-cancer effect of PI3K inhibitors. •The results suggest to combine PI3K and AKT inhibitors in cancer therapy. -- Abstract: Targeting the phosphatidylinositol-3-kinase (PI3K) is a promising approach in cancer therapy. In particular, PI3K blockade leads to the inhibition of AKT, a major downstream effector responsible for the oncogenic activity of PI3K. However, we report here that small molecule inhibitors of PI3K only transiently block AKT signaling. Indeed, treatment of cancer cells with PI3K inhibitors results in a rapid inhibition of AKT phosphorylation and signaling which is followed by the reactivation of AKT signaling after 48 h as observed by Western blot. Reactivation of AKT signaling occurs despite effective inhibition of PI3K activity by PI3K inhibitors. In addition, wortmannin, a broad range PI3K inhibitor, did not block AKT reactivation suggesting that AKT signals independently of PI3K. In a therapeutical perspective, combining AKT and PI3K inhibitors exhibit stronger anti-proliferative and pro-apoptotic effects compared to AKT or PI3K inhibitors alone. Similarly, in a tumor xenograft mouse model, concomitant PI3K and AKT blockade results in stronger anti-cancer activity compared with either blockade alone. This study shows that PI3K inhibitors only transiently inhibit AKT which limits their antitumor activities. It also provides the proof of concept to combine PI3K inhibitors with AKT inhibitors in cancer therapy.

  10. Nuclear PI3K signaling in cell growth and tumorigenesis

    PubMed Central

    Davis, William J.; Lehmann, Peter Z.; Li, Weimin

    2015-01-01

    The PI3K/Akt signaling pathway is a major driving force in a variety of cellular functions. Dysregulation of this pathway has been implicated in many human diseases including cancer. While the activity of the cytoplasmic PI3K/Akt pathway has been extensively studied, the functions of these molecules and their effector proteins within the nucleus are poorly understood. Harboring key cellular processes such as DNA replication and repair as well as nascent messenger RNA transcription, the nucleus provides a unique compartmental environment for protein–protein and protein–DNA/RNA interactions required for cell survival, growth, and proliferation. Here we summarize recent advances made toward elucidating the nuclear PI3K/Akt signaling cascade and its key components within the nucleus as they pertain to cell growth and tumorigenesis. This review covers the spatial and temporal localization of the major nuclear kinases having PI3K activities and the counteracting phosphatases as well as the role of nuclear PI3K/Akt signaling in mRNA processing and exportation, DNA replication and repair, ribosome biogenesis, cell survival, and tumorigenesis. PMID:25918701

  11. Fangchinoline targets PI3K and suppresses PI3K/AKT signaling pathway in SGC7901 cells

    PubMed Central

    TIAN, FENG; DING, DING; LI, DANDAN

    2015-01-01

    Fangchinoline, an important compound in Stephania tetrandra S. Moore, as a novel antitumor agent, has been implicated in several types of cancers cells except gastric cancer. To investigate whether fangchinoline affects gastric cancer cells, we detected the signaling pathway by which fangchinoline plays a role in different human gastric cancer cells lines. We found that fangchinoline effectively suppressed proliferation and invasion of SGC7901 cell lines, but not MKN45 cell lines by inhibiting the expression of PI3K and its downstream pathway. All of the Akt/MMP2/MMP9 pathway, Akt/Bad pathway, and Akt/Gsk3β/CDK2 pathway could be inhibited by fangchinoline through inhibition of PI3K. Taken together, these results suggest that fangchinoline targets PI3K in tumor cells that express PI3K abundantly and inhibits the growth and invasive ability of the tumor cells. PMID:25872479

  12. Fangchinoline targets PI3K and suppresses PI3K/AKT signaling pathway in SGC7901 cells.

    PubMed

    Tian, Feng; Ding, Ding; Li, Dandan

    2015-01-01

    Fangchinoline, an important compound in Stephania tetrandra S. Moore, as a novel antitumor agent, has been implicated in several types of cancers cells except gastric cancer. To investigate whether fangchinoline affects gastric cancer cells, we detected the signaling pathway by which fangchinoline plays a role in different human gastric cancer cells lines. We found that fangchinoline effectively suppressed proliferation and invasion of SGC7901 cell lines, but not MKN45 cell lines by inhibiting the expression of PI3K and its downstream pathway. All of the Akt/MMP2/MMP9 pathway, Akt/Bad pathway, and Akt/Gsk3β/CDK2 pathway could be inhibited by fangchinoline through inhibition of PI3K. Taken together, these results suggest that fangchinoline targets PI3K in tumor cells that express PI3K abundantly and inhibits the growth and invasive ability of the tumor cells. PMID:25872479

  13. The Role of PI3K/Akt/mTOR Signaling in Gastric Carcinoma

    PubMed Central

    Matsuoka, Tasuku; Yashiro, Masakazu

    2014-01-01

    The phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway is one of the key signaling pathways induced by various receptor-tyrosine kinases. Accumulating evidence shows that this pathway is an important promoter of cell growth, metabolism, survival, metastasis, and resistance to chemotherapy. Genetic alterations in the PI3K/Akt/mTOR pathway in gastric carcinoma have often been demonstrated. Many kinds of molecular targeting therapies are currently undergoing clinical testing in patients with solid tumors. However, with the exception of the ErbB2-targeting antibody, targeting agents, including PI3K/Akt/mTOR inhibitors, have not been approved for treatment of patients with gastric carcinoma. This review summarizes the current knowledge on PI3K/Akt/mTOR signaling in the pathogenesis of gastric carcinoma and the possible therapeutic targets for gastric carcinoma. Improved knowledge of the PI3K/Akt/mTOR pathway in gastric carcinoma will be useful in understanding the mechanisms of tumor development and for identifying ideal targets of anticancer therapy for gastric carcinoma. PMID:25003395

  14. Following the trail of lipids: Signals initiated by PI3K function at multiple cellular membranes.

    PubMed

    Naguib, Adam

    2016-05-17

    Phosphatidylinositol 3,4,5-trisphosphate [PtdIns(3,4,5)P3] is the signaling currency of the phosphoinositide 3-kinase (PI3K)/AKT pathway; transduction through this axis depends on this signaling lipid. Formation of PtdIns(3,4,5)P3 is dictated not only by PI3K activation but also by the localization and access of PI3K to its substrate PtdIns(4,5)P2 (phosphatidylinositol 4,5-bisphosphate). PI3K/AKT-mediated signaling is antagonized by PtdIns(3,4,5)P3 dephosphorylation. Although previously typically considered an event associated with the plasma membrane, it is now appreciated that the formation and metabolism of PtdIns(3,4,5)P3 occur on multiple membranes with distinct kinetics. Modulated activity of phosphatidylinositol lipid kinases and phosphatases contributes to intricately orchestrated lipid gradients that define the signaling status of the pathway at multiple sites within the cell.

  15. Thyroid hormone inhibits the proliferation of piglet Sertoli cell via PI3K signaling pathway.

    PubMed

    Sun, Yan; Yang, WeiRong; Luo, HongLin; Wang, XianZhong; Chen, ZhongQiong; Zhang, JiaoJiao; Wang, Yi; Li, XiaoMin

    2015-01-01

    Accumulating researches show that thyroid hormone (TH) inhibits Sertoli cells (SCs) proliferation and stimulates their functional maturation in prepubertal rat testis, confirming that TH plays a key role in testicular development. However, the mechanism under the T3 regulation of piglet SC proliferation remains unclear. In the present study, in order to investigate the possible mechanism of T3 on the suppression of SC proliferation, the expression pattern of TRα1 and cell cycle-related molecules, effect of T3 on SC proliferation, and the role of phosphoinositide 3-kinase (PI3K)/Akt signaling pathway on the T3-mediated SC proliferation in piglet testis were explored. Our results demonstrated that TRα1 was expressed in all tested stages of SCs and decreased along with the ages. T3 inhibited the proliferation of SCs in a time- and dose-dependent manner, and T3 treatment downregulated the expressions of cell cycling molecules, such as cyclinA2, cyclinD1, cyclinE1, PCNA, and Skp2, but upregulated the p27 expression in SCs. Most importantly, the suppressive effects of T3 on SC proliferation seemed dependent on the inhibition of PI3K/Akt signaling pathway, and pre-stimulation of PI3K could enhance such suppressive effects. Together, our findings demonstrate that TH inhibits the proliferation of piglet SCs via the suppression of PI3K/Akt signaling pathway.

  16. Shiga toxin type-2 (Stx2) induces glutamate release via phosphoinositide 3-kinase (PI3K) pathway in murine neurons

    PubMed Central

    Obata, Fumiko; Hippler, Lauren M.; Saha, Progyaparamita; Jandhyala, Dakshina M.; Latinovic, Olga S.

    2015-01-01

    Shiga toxin-producing Escherichia coli (STEC) can cause central nervous system (CNS) damage resulting in paralysis, seizures, and coma. The key STEC virulence factors associated with systemic illness resulting in CNS impairment are Shiga toxins (Stx). While neurons express the Stx receptor globotriaosylceramide (Gb3) in vivo, direct toxicity to neurons by Stx has not been studied. We used murine neonatal neuron cultures to study the interaction of Shiga toxin type 2 (Stx2) with cell surface expressed Gb3. Single molecule imaging three dimensional STochastic Optical Reconstruction Microscopy—Total Internal Reflection Fluorescence (3D STORM-TIRF) allowed visualization and quantification of Stx2-Gb3 interactions. Furthermore, we demonstrate that Stx2 increases neuronal cytosolic Ca2+, and NMDA-receptor inhibition blocks Stx2-induced Ca2+ influx, suggesting that Stx2-mediates glutamate release. Phosphoinositide 3-kinase (PI3K)-specific inhibition by Wortmannin reduces Stx2-induced intracellular Ca2+ indicating that the PI3K signaling pathway may be involved in Stx2-associated glutamate release, and that these pathways may contribute to CNS impairment associated with STEC infection. PMID:26236186

  17. Role of phosphoinositide 3-kinase IA (PI3K-IA) activation in cardioprotection induced by ouabain preconditioning.

    PubMed

    Duan, Qiming; Madan, Namrata D; Wu, Jian; Kalisz, Jennifer; Doshi, Krunal Y; Haldar, Saptarsi M; Liu, Lijun; Pierre, Sandrine V

    2015-03-01

    Acute myocardial infarction, the clinical manifestation of ischemia-reperfusion (IR) injury, is a leading cause of death worldwide. Like ischemic preconditioning (IPC) induced by brief episodes of ischemia and reperfusion, ouabain preconditioning (OPC) mediated by Na/K-ATPase signaling protects the heart against IR injury. Class I PI3K activation is required for IPC, but its role in OPC has not been investigated. While PI3K-IB is critical to IPC, studies have suggested that ouabain signaling is PI3K-IA-specific. Hence, a pharmacological approach was used to test the hypothesis that OPC and IPC rely on distinct PI3K-I isoforms. In Langendorff-perfused mouse hearts, OPC was initiated by 4 min of ouabain 10 μM and IPC was triggered by 4 cycles of 5 min ischemia and reperfusion prior to 40 min of global ischemia and 30 min of reperfusion. Without affecting PI3K-IB, ouabain doubled PI3K-IA activity and Akt phosphorylation at Ser(473). IPC and OPC significantly preserved cardiac contractile function and tissue viability as evidenced by left ventricular developed pressure and end-diastolic pressure recovery, reduced lactate dehydrogenase release, and decreased infarct size. OPC protection was blunted by the PI3K-IA inhibitor PI-103, but not by the PI3K-IB inhibitor AS-604850. In contrast, IPC-mediated protection was not affected by PI-103 but was blocked by AS-604850, suggesting that PI3K-IA activation is required for OPC while PI3K-IB activation is needed for IPC. Mechanistically, PI3K-IA activity is required for ouabain-induced Akt activation but not PKCε translocation. However, in contrast to PKCε translocation which is critical to protection, Akt activity was not required for OPC. Further studies shall reveal the identity of the downstream targets of this new PI3K IA-dependent branch of OPC. These findings may be of clinical relevance in patients at risk for myocardial infarction with underlying diseases and/or medication that could differentially affect the

  18. Activity-dependent survival of developing neocortical neurons depends on PI3K signalling.

    PubMed

    Wagner-Golbs, Antje; Luhmann, Heiko J

    2012-02-01

    Spontaneous electrical network activity plays a major role in the control of cell survival in the developing brain. Several intracellular pathways are implicated in transducing electrical activity into gene expression dependent and independent survival signals. These include activation of phosphatidylinositol 3-kinase (PI3K) and its downstream effector Akt, activation of Ras and subsequently MAPK/extracellular signal-regulated kinase (MEK) and extracellular signal-regulated kinase and signalling via calcium/calmodulin-dependent protein kinase (CaMK). In the present study, we analyzed the role of these pathways for the control of neuronal survival in different extracellular potassium concentrations ([K(+) ](ex) ). Organotypic neocortical slice cultures prepared from newborn mice were kept in 5.3, 8.0 and 25.0mM [K(+) ](ex) and treated with specific inhibitors of PI3K, MEK1, CaMKK and a broad spectrum CaMK inhibitor. After 6h of incubation, slices were immunostained for activated caspase 3 (a-caspase 3) and the number of apoptotic cells was quantified by computer based analysis. We found that in 5.3 and 8.0mM [K(+) ](ex) only PI3K was important for neuronal survival. When [K(+) ](ex) was raised to 25.0mM, a concentration above the depolarization block, we found no influence of PI3K on neuronal survival. Our data demonstrate that only the PI3K pathway, and not the MEK1, CaMKK or CaMKs pathway, plays a central role in the regulation of activity-dependent neuronal survival in the developing cerebral cortex.

  19. Coordinate activation of Shh and PI3K signaling in PTEN-deficient glioblastoma: new therapeutic opportunities.

    PubMed

    Filbin, Mariella Gruber; Dabral, Sukriti K; Pazyra-Murphy, Maria F; Ramkissoon, Shakti; Kung, Andrew L; Pak, Ekaterina; Chung, Jarom; Theisen, Matthew A; Sun, Yanping; Franchetti, Yoko; Sun, Yu; Shulman, David S; Redjal, Navid; Tabak, Barbara; Beroukhim, Rameen; Wang, Qi; Zhao, Jean; Dorsch, Marion; Buonamici, Silvia; Ligon, Keith L; Kelleher, Joseph F; Segal, Rosalind A

    2013-11-01

    In glioblastoma, phosphatidylinositol 3-kinase (PI3K) signaling is frequently activated by loss of the tumor suppressor phosphatase and tensin homolog (PTEN). However, it is not known whether inhibiting PI3K represents a selective and effective approach for treatment. We interrogated large databases and found that sonic hedgehog (SHH) signaling is activated in PTEN-deficient glioblastoma. We demonstrate that the SHH and PI3K pathways synergize to promote tumor growth and viability in human PTEN-deficient glioblastomas. A combination of PI3K and SHH signaling inhibitors not only suppressed the activation of both pathways but also abrogated S6 kinase (S6K) signaling. Accordingly, targeting both pathways simultaneously resulted in mitotic catastrophe and tumor apoptosis and markedly reduced the growth of PTEN-deficient glioblastomas in vitro and in vivo. The drugs tested here appear to be safe in humans; therefore, this combination may provide a new targeted treatment for glioblastoma. PMID:24076665

  20. Sensing and Integration of Erk and PI3K Signals by Myc

    PubMed Central

    Lee, Tae; Yao, Guang; Nevins, Joseph; You, Lingchong

    2008-01-01

    The transcription factor Myc plays a central role in regulating cell-fate decisions, including proliferation, growth, and apoptosis. To maintain a normal cell physiology, it is critical that the control of Myc dynamics is precisely orchestrated. Recent studies suggest that such control of Myc can be achieved at the post-translational level via protein stability modulation. Myc is regulated by two Ras effector pathways: the extracellular signal-regulated kinase (Erk) and phosphatidylinositol 3-kinase (PI3K) pathways. To gain quantitative insight into Myc dynamics, we have developed a mathematical model to analyze post-translational regulation of Myc via sequential phosphorylation by Erk and PI3K. Our results suggest that Myc integrates Erk and PI3K signals to result in various cellular responses by differential stability control of Myc protein isoforms. Such signal integration confers a flexible dynamic range for the system output, governed by stability change. In addition, signal integration may require saturation of the input signals, leading to sensitive signal integration to the temporal features of the input signals, insensitive response to their amplitudes, and resistance to input fluctuations. We further propose that these characteristics of the protein stability control module in Myc may be commonly utilized in various cell types and classes of proteins. PMID:18463697

  1. Targeting the phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway: an emerging treatment strategy for squamous cell lung carcinoma.

    PubMed

    Beck, Joseph Thaddeus; Ismail, Amen; Tolomeo, Christina

    2014-09-01

    Squamous cell lung carcinoma accounts for approximately 30% of all non-small cell lung cancers (NSCLCs). Despite progress in the understanding of the biology of cancer, cytotoxic chemotherapy remains the standard of care for patients with squamous cell lung carcinoma, but the prognosis is generally poor. The phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) pathway is one of the most commonly activated signaling pathways in cancer, leading to cell proliferation, survival, and differentiation. It has therefore become a major focus of clinical research. Various alterations in the PI3K/AKT/mTOR pathway have been identified in squamous cell lung carcinoma and a number of agents targeting these alterations are in clinical development for use as single agents and in combination with other targeted and conventional treatments. These include pan-PI3K inhibitors, isoform-specific PI3K inhibitors, AKT inhibitors, mTOR inhibitors, and dual PI3K/mTOR inhibitors. These agents have demonstrated antitumor activity in preclinical models of NSCLC and preliminary clinical evidence is also available for some agents. This review will discuss the role of the PI3K/AKT/mTOR pathway in cancer and how the discovery of genetic alterations in this pathway in patients with squamous cell lung carcinoma can inform the development of targeted therapies for this disease. An overview of ongoing clinical trials investigating PI3K/AKT/mTOR pathway inhibitors in squamous cell lung carcinoma will also be included.

  2. A Switch of G Protein-Coupled Receptor Binding Preference from Phosphoinositide 3-Kinase (PI3K)–p85 to Filamin A Negatively Controls the PI3K Pathway

    PubMed Central

    Najib, Souad; Saint-Laurent, Nathalie; Estève, Jean-Pierre; Schulz, Stefan; Boutet-Robinet, Elisa; Fourmy, Daniel; Lättig, Jens; Mollereau, Catherine; Pyronnet, Stéphane; Susini, Christiane

    2012-01-01

    Frequent oncogenic alterations occur in the phosphoinositide 3-kinase (PI3K) pathway, urging identification of novel negative controls. We previously reported an original mechanism for restraining PI3K activity, controlled by the somatostatin G protein-coupled receptor (GPCR) sst2 and involving a ligand-regulated interaction between sst2 with the PI3K regulatory p85 subunit. We here identify the scaffolding protein filamin A (FLNA) as a critical player regulating the dynamic of this complex. A preexisting sst2-p85 complex, which was shown to account for a significant basal PI3K activity in the absence of ligand, is disrupted upon sst2 activation. FLNA was here identified as a competitor of p85 for direct binding to two juxtaposed sites on sst2. Switching of GPCR binding preference from p85 toward FLNA is determined by changes in the tyrosine phosphorylation of p85- and FLNA-binding sites on sst2 upon activation. It results in the disruption of the sst2-p85 complex and the subsequent inhibition of PI3K. Knocking down FLNA expression, or abrogating FLNA recruitment to sst2, reversed the inhibition of PI3K and of tumor growth induced by sst2. Importantly, we report that this FLNA inhibitory control on PI3K can be generalized to another GPCR, the mu opioid receptor, thereby providing an unprecedented mechanism underlying GPCR-negative control on PI3K. PMID:22203038

  3. Dual PI3K/mTOR inhibition is required to effectively impair microenvironment survival signals in mantle cell lymphoma

    PubMed Central

    Rosich, Laia; Montraveta, Arnau; Xargay-Torrent, Sílvia; López-Guerra, Mónica; Roldán, Jocabed; Aymerich, Marta; Salaverria, Itziar; Beà, Sílvia; Campo, Elías; Pérez-Galán, Patricia; Roué, Gaël; Colomer, Dolors

    2014-01-01

    Phosphatidylinositol-3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway activation contributes to mantle cell lymphoma (MCL) pathogenesis and drug resistance. Antitumor activity has been observed with mTOR inhibitors. However, they have shown limited clinical efficacy in relation to drug activation of feedback loops. Selective PI3K inhibition or dual PI3K/mTOR catalytic inhibition are different therapeutic approaches developed to achieve effective pathway blockage. Here, we have performed a comparative analysis of the mTOR inhibitor everolimus, the pan-PI3K inhibitor NVP-BKM120 and the dual PI3K/mTOR inhibitor NVP-BEZ235 in primary MCL cells. We found NVP-BEZ235 to be more powerful than everolimus or NVP-BKM120 in PI3K/Akt/mTOR signaling inhibition, indicating that targeting the PI3K/Akt/mTOR pathway at multiple levels is likely to be a more effective strategy for the treatment of MCL than single inhibition of these kinases. Among the three drugs, NVP-BEZ235 induced the highest change in gene expression profile. Functional validation demonstrated that NVP-BEZ235 inhibited angiogenesis, migration and tumor invasiveness in MCL cells. NVP-BEZ235 was the only drug able to block IL4 and IL6/STAT3 signaling which compromise the therapeutic effect of chemotherapy in MCL. Our findings support the use of the dual PI3K/mTOR inhibitor NVP-BEZ235 as a promising approach to interfere with the microenvironment-related processes in MCL. PMID:25216518

  4. TDRG1 functions in testicular seminoma are dependent on the PI3K/Akt/mTOR signaling pathway

    PubMed Central

    Wang, Yong; Gan, Yu; Tan, Zhengyu; Zhou, Jun; Kitazawa, Riko; Jiang, Xianzhen; Tang, Yuxin; Yang, Jianfu

    2016-01-01

    Human testis development-related gene 1 (TDRG1) is a recently identified gene that is expressed exclusively in the testes and promotes the development of testicular germ cell tumors. In this study, the role of TDRG1 in the development of testicular seminoma, which is the most common testicular germ cell tumor, was further investigated. Based on polymerase chain reaction, Western blotting, and immunohistochemistry tests, both gene and protein expression levels of TDRG1 were significantly upregulated in testicular seminoma tissues compared with normal testicular tissues. Additionally, the levels of phosphoinositide-3 kinase (PI3K)/p110 and Akt phosphorylation were dramatically upregulated in testicular seminoma tissues. Accordingly, in our cell experiment, seminoma TCam-2 cells were subjected to different treatments: the TDRG1 knockout, TDRG1 overexpression, PI3K inhibition (LY294002 administration), or PI3K activation (insulin-like growth factor-1 administration). Cell proliferation, the proliferation index, apoptosis rate, cell adhesive capacity, and cell invasion capability were assessed. Cells with both TDRG1 knockout and PI3K inhibition exhibited decreased cell proliferation, proliferation indexes, cell adhesion capacity, and cell invasion capability and increased apoptosis rates. Most of these effects were reversed by TDRG1 overexpression or PI3K activation, indicating that both TDRG1- and PI3K-mediated signaling promote proliferation and invasion of testicular seminoma cells. The knockout of TDRG1 significantly decreased the phosphorylation levels of PI3K/p85, PI3K/p110, Akt, and mammalian target of rapamycin (mTOR; Ser2448). Except for PI3K/p110, TDRG1 overexpression had the opposite effects on phosphorylation levels. Phosphorylated mTOR at Ser2481 and Thr2446 was not affected by TDRG1 or PI3K in our tests. Thus, these results indicate that TDRG1 promotes the development and migration of seminoma cells via the regulation of the PI3K/Akt/mTOR signaling pathway

  5. PI3K-GSK3 signalling regulates mammalian axon regeneration by inducing the expression of Smad1

    NASA Astrophysics Data System (ADS)

    Saijilafu; Hur, Eun-Mi; Liu, Chang-Mei; Jiao, Zhongxian; Xu, Wen-Lin; Zhou, Feng-Quan

    2013-10-01

    In contrast to neurons in the central nervous system, mature neurons in the mammalian peripheral nervous system (PNS) can regenerate axons after injury, in part, by enhancing intrinsic growth competence. However, the signalling pathways that enhance the growth potential and induce spontaneous axon regeneration remain poorly understood. Here we reveal that phosphatidylinositol 3-kinase (PI3K) signalling is activated in response to peripheral axotomy and that PI3K pathway is required for sensory axon regeneration. Moreover, we show that glycogen synthase kinase 3 (GSK3), rather than mammalian target of rapamycin, mediates PI3K-dependent augmentation of the growth potential in the PNS. Furthermore, we show that PI3K-GSK3 signal is conveyed by the induction of a transcription factor Smad1 and that acute depletion of Smad1 in adult mice prevents axon regeneration in vivo. Together, these results suggest PI3K-GSK3-Smad1 signalling as a central module for promoting sensory axon regeneration in the mammalian nervous system.

  6. 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. PMID:27494022

  7. Integrin β1 mediates vaccinia virus entry through activation of PI3K/Akt signaling.

    PubMed

    Izmailyan, Roza; Hsao, Jye-Chian; Chung, Che-Sheng; Chen, Chein-Hung; Hsu, Paul Wei-Che; Liao, Chung-Lin; Chang, Wen

    2012-06-01

    Vaccinia virus has a broad range of infectivity in many cell lines and animals. Although it is known that the vaccinia mature virus binds to cell surface glycosaminoglycans and extracellular matrix proteins, whether additional cellular receptors are required for virus entry remains unclear. Our previous studies showed that the vaccinia mature virus enters through lipid rafts, suggesting the involvement of raft-associated cellular proteins. Here we demonstrate that one lipid raft-associated protein, integrin β1, is important for vaccinia mature virus entry into HeLa cells. Vaccinia virus associates with integrin β1 in lipid rafts on the cell surface, and the knockdown of integrin β1 in HeLa cells reduces vaccinia mature virus entry. Additionally, vaccinia mature virus infection is reduced in a mouse cell line, GD25, that is deficient in integrin β1 expression. Vaccinia mature virus infection triggers the activation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling, and the treatment of cells with inhibitors to block P13K activation reduces virus entry in an integrin β1-dependent manner, suggesting that integrin β1-mediates PI3K/Akt activation induced by vaccinia virus and that this signaling pathway is essential for virus endocytosis. The inhibition of integrin β1-mediated cell adhesion results in a reduction of vaccinia virus entry and the disruption of focal adhesion and PI3K/Akt activation. In summary, our results show that the binding of vaccinia mature virus to cells mimics the outside-in activation process of integrin functions to facilitate vaccinia virus entry into HeLa cells.

  8. Phosphatidylinositol-3-kinase (PI3K) is activated by influenza virus vRNA via the pathogen pattern receptor Rig-I to promote efficient type I interferon production.

    PubMed

    Hrincius, Eike R; Dierkes, Rüdiger; Anhlan, Darisuren; Wixler, Viktor; Ludwig, Stephan; Ehrhardt, Christina

    2011-12-01

    The phosphatidylinositol-3-kinase (PI3K) was identified to be activated upon influenza A virus (IAV) infection. An early and transient induction of PI3K signalling is caused by viral attachment to cells and promotes virus entry. In later phases of infection the kinase is activated by the viral NS1 protein to prevent premature apoptosis. Besides these virus supporting functions, it was suggested that PI3K signalling is involved in dsRNA and IAV induced antiviral responses by enhancing the activity of interferon regulatory factor-3 (IRF-3). However, molecular mechanisms of activation remained obscure. Here we show that accumulation of vRNA in cells infected with influenza A or B viruses results in PI3K activation. Furthermore, expression of the RNA receptors Rig-I and MDA5 was increased upon stimulation with virion extracted vRNA or IAV infection. Using siRNA approaches, Rig-I was identified as pathogen receptor necessary for influenza virus vRNA sensing and subsequent PI3K activation in a TRIM25 and MAVS signalling dependent manner. Rig-I induced PI3K signalling was further shown to be essential for complete IRF-3 activation and consequently induction of the type I interferon response. These data identify PI3K as factor that is activated as part of the Rig-I mediated anti-pathogen response to enhance expression of type I interferons.

  9. Will targeting PI3K/Akt/mTOR signaling work in hematopoietic malignancies?

    PubMed Central

    Gao, Yanan; Yuan, Chase Y.

    2016-01-01

    The constitutive activation of phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) signaling pathway has been demonstrated to be critical in clinical cancer patients as well as in laboratory cancer models including hematological malignancies. Great efforts have been made to develop inhibitors targeting this pathway in hematological malignancies but so far the efficacies of these inhibitors were not as good as expected. By analyzing existing literatures and datasets available, we found that mutations of genes in the pathway only constitute a very small subset of hematological malignancies. Deep understanding of the function of gene, the pathway and/or its regulators, and the cellular response to inhibitors, may help us design better drugs targeting the hematological malignancies. PMID:27583254

  10. Follistatin could promote the proliferation of duck primary myoblasts by activating PI3K/Akt/mTOR signalling

    PubMed Central

    Li, Xinxin; Liu, Hehe; Wang, Haohan; Sun, Lingli; Ding, Fang; Sun, Wenqiang; Han, Chunchun; Wang, Jiwen

    2014-01-01

    FST (follistatin) is essential for skeletal muscle development, but the intracellular signalling networks that regulate FST-induced effects are not well defined. We sought to investigate whether FST promotes the proliferation of myoblasts through the PI3K (phosphoinositide 3-kinase)/Akt (protein kinase B)/mTOR (mammalian target of rapamycin) signalling. In the present study, we transfected the pEGFP-duFST plasmid and added PI3K and mTOR inhibitors to the medium of duck primary myoblasts. Then, we analysed the cellular phenotypic changes that occurred and analysed the expression of target genes. The results showed that FST promoted myoblast proliferation, induced the mRNA expression of PI3K, Akt, mTOR, 70-kDa ribosomal protein S6K (S6 kinase) and the protein expression of phospho-Akt (Thr308), mTOR, phospho-mTOR (serine 2448), phospho-S6K (Ser417), inhibited the mRNA expression of FoxO1, MuRF1 (muscle RING finger-1) and the protein expression of phospho-FoxO1 (Ser256). Moreover, we found that the overexpression of FST could alleviate the inhibitory effect of myoblast proliferation caused by the addition of LY294002, a PI3K inhibitor. Additionally, the overexpression of duck FST also relieved the inhibition of myoblast proliferation caused by the addition of rapamycin (an mTOR inhibitor) through PI3K/Akt/mTOR signalling. In light of the present results, we hypothesize that duck FST could promote myoblast proliferation, which is dependent on PI3K/Akt/mTOR signalling. PMID:25200144

  11. Closing escape routes: inhibition of IL-8 signaling enhances the anti-tumor efficacy of PI3K inhibitors.

    PubMed

    Juvekar, Ashish; Wulf, Gerburg M

    2013-04-08

    The phosphoinositide 3-kinase (PI3K) pathway serves as a relay where signals that emanate from the cell membrane are received and converted into intracellular signals that promote proliferation and survival. Inhibitors of PI3K hold promise for the treatment of breast cancer because activation of this pathway is highly prevalent. However, as is increasingly observed with inhibitors of cell signaling, there appear to be mechanisms of primary and secondary resistance. Britschgi and colleagues report that compensatory activation of the IL-8 signaling axis is a mechanism of primary resistance to PI3K inhibitors in some triple-negative breast cancers. In a set of experiments that carefully emulate the clinical scenario in a mouse model, they show that simultaneous inhibition of Janus kinase 2 enhances the efficacy of PI3K/mammalian target of rapamycin inhibition. Their paper lends further support to the concept that successful design of treatments with signal transduction inhibitors must anticipate potential escape routes - and include agents to simultaneously block them.

  12. PTEN regulates angiogenesis through PI3K/Akt/VEGF signaling pathway in human pancreatic cancer cells.

    PubMed

    Ma, Jiachi; Sawai, Hirozumi; Ochi, Nobuo; Matsuo, Yoichi; Xu, Donghui; Yasuda, Akira; Takahashi, Hiroki; Wakasugi, Takehiro; Takeyama, Hiromitsu

    2009-11-01

    Phosphoinositide 3-kinase (PI3K) pathway exerts its effects through Akt, its downstream target molecule, and thereby regulates various cell functions including cell proliferation, cell transformation, apoptosis, tumor growth, and angiogenesis. Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) has been implicated in regulating cell survival signaling through the PI3K/Akt pathway. However, the mechanism by PI3K/PTEN signaling regulates angiogenesis and tumor growth in vivo remains to be elucidated. Vascular endothelial growth factor (VEGF) plays a pivotal role in tumor angiogenesis. The effect of PTEN on VEGF-mediated signal in pancreatic cancer is unknown. This study aimed to determine the effect of PTEN on both the expression of VEGF and angiogenesis. Toward that end, we used the siRNA knockdown method to specifically define the role of PTEN in the expression of VEGF and angiogenesis. We found that siRNA-mediated inhibition of PTEN gene expression in pancreatic cancer cells increase their VEGF secretion, up-modulated the proliferation, and migration of co-cultured vascular endothelial cell and enhanced tubule formation by HUVEC. In addition, PTEN modulated VEGF-mediated signaling and affected tumor angiogenesis through PI3K/Akt/VEGF/eNOS pathway.

  13. Eupatilin induces human renal cancer cell apoptosis via ROS-mediated MAPK and PI3K/AKT signaling pathways

    PubMed Central

    Zhong, Wei-Feng; Wang, Xiao-Hong; Pan, Bin; Li, Feng; Kuang, Lu; Su, Ze-Xuan

    2016-01-01

    Phosphatidylinositol 3-kinase (PI3K)/AKT and mitogen activated protein kinase (MAPK) signaling cascades have significant roles in cell proliferation, survival, angiogenesis and metastasis of tumor cells. Eupatilin, one of the major compounds present in Artemisia species, has been demonstrated to have antitumor properties. However, the effect of eupatilin in renal cell carcinoma (RCC) remains to be elucidated. Therefore, the present study investigated the biological effects and mechanisms of eupatilin in RCC cell apoptosis. The results of the present study demonstrated that eupatilin significantly induced cell apoptosis and enhanced the production of reactive oxygen species (ROS) in 786-O cells. In addition, eupatilin induced phosphorylation of p38α (Thr180/Tyr182), extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase 1/2 (Thr183/Tyr185), and decreased the phosphorylation of PI3K and AKT in 786-O cells in a concentration-dependent manner. Furthermore, the ROS inhibitor N-acetyl-L-cysteine was able to rescue the MAPK activation and PI3K/AKT inhibition induced by eupatilin. Taken together, the results of the present study provide evidence that inhibition of eupatilin induces apoptosis in human RCC via ROS-mediated activation of the MAPK signaling pathway and inhibition of the PI3K/AKT signaling pathway. Thus, eupatilin may serve as a potential therapeutic agent for the treatment of human RCC. PMID:27698876

  14. Eupatilin induces human renal cancer cell apoptosis via ROS-mediated MAPK and PI3K/AKT signaling pathways

    PubMed Central

    Zhong, Wei-Feng; Wang, Xiao-Hong; Pan, Bin; Li, Feng; Kuang, Lu; Su, Ze-Xuan

    2016-01-01

    Phosphatidylinositol 3-kinase (PI3K)/AKT and mitogen activated protein kinase (MAPK) signaling cascades have significant roles in cell proliferation, survival, angiogenesis and metastasis of tumor cells. Eupatilin, one of the major compounds present in Artemisia species, has been demonstrated to have antitumor properties. However, the effect of eupatilin in renal cell carcinoma (RCC) remains to be elucidated. Therefore, the present study investigated the biological effects and mechanisms of eupatilin in RCC cell apoptosis. The results of the present study demonstrated that eupatilin significantly induced cell apoptosis and enhanced the production of reactive oxygen species (ROS) in 786-O cells. In addition, eupatilin induced phosphorylation of p38α (Thr180/Tyr182), extracellular signal-regulated kinase 1/2 and c-Jun N-terminal kinase 1/2 (Thr183/Tyr185), and decreased the phosphorylation of PI3K and AKT in 786-O cells in a concentration-dependent manner. Furthermore, the ROS inhibitor N-acetyl-L-cysteine was able to rescue the MAPK activation and PI3K/AKT inhibition induced by eupatilin. Taken together, the results of the present study provide evidence that inhibition of eupatilin induces apoptosis in human RCC via ROS-mediated activation of the MAPK signaling pathway and inhibition of the PI3K/AKT signaling pathway. Thus, eupatilin may serve as a potential therapeutic agent for the treatment of human RCC.

  15. Phospshoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) dual inhibitors: discovery and structure-activity relationships of a series of quinoline and quinoxaline derivatives.

    PubMed

    Nishimura, Nobuko; Siegmund, Aaron; Liu, Longbin; Yang, Kevin; Bryan, Marian C; Andrews, Kristin L; Bo, Yunxin; Booker, Shon K; Caenepeel, Sean; Freeman, Daniel; Liao, Hongyu; McCarter, John; Mullady, Erin L; San Miguel, Tisha; Subramanian, Raju; Tamayo, Nuria; Wang, Ling; Whittington, Douglas A; Zalameda, Leeanne; Zhang, Nancy; Hughes, Paul E; Norman, Mark H

    2011-07-14

    The phosphoinositide 3-kinase (PI3K) family catalyzes the ATP-dependent phosphorylation of the 3'-hydroxyl group of phosphatidylinositols and plays an important role in cell growth and survival. There is abundant evidence demonstrating that PI3K signaling is dysregulated in many human cancers, suggesting that therapeutics targeting the PI3K pathway may have utility for the treatment of cancer. Our efforts to identify potent, efficacious, and orally available PI3K/mammalian target of rapamycin (mTOR) dual inhibitors resulted in the discovery of a series of substituted quinolines and quinoxalines derivatives. In this report, we describe the structure-activity relationships, selectivity, and pharmacokinetic data of this series and illustrate the in vivo pharmacodynamic and efficacy data for a representative compound.

  16. PI3K-C2γ is a Rab5 effector selectively controlling endosomal Akt2 activation downstream of insulin signalling

    PubMed Central

    Braccini, Laura; Ciraolo, Elisa; Campa, Carlo C.; Perino, Alessia; Longo, Dario L.; Tibolla, Gianpaolo; Pregnolato, Marco; Cao, Yanyan; Tassone, Beatrice; Damilano, Federico; Laffargue, Muriel; Calautti, Enzo; Falasca, Marco; Norata, Giuseppe D.; Backer, Jonathan M.; Hirsch, Emilio

    2015-01-01

    In the liver, insulin-mediated activation of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway is at the core of metabolic control. Multiple PI3K and Akt isoenzymes are found in hepatocytes and whether isoform-selective interplays exist is currently unclear. Here we report that insulin signalling triggers the association of the liver-specific class II PI3K isoform γ (PI3K-C2γ) with Rab5-GTP, and its recruitment to Rab5-positive early endosomes. In these vesicles, PI3K-C2γ produces a phosphatidylinositol-3,4-bisphosphate pool specifically required for delayed and sustained endosomal Akt2 stimulation. Accordingly, loss of PI3K-C2γ does not affect insulin-dependent Akt1 activation as well as S6K and FoxO1-3 phosphorylation, but selectively reduces Akt2 activation, which specifically inhibits glycogen synthase activity. As a consequence, PI3K-C2γ-deficient mice display severely reduced liver accumulation of glycogen and develop hyperlipidemia, adiposity as well as insulin resistance with age or after consumption of a high-fat diet. Our data indicate PI3K-C2γ supports an isoenzyme-specific forking of insulin-mediated signal transduction to an endosomal pool of Akt2, required for glucose homeostasis. PMID:26100075

  17. Aged black garlic extract inhibits HT29 colon cancer cell growth via the PI3K/Akt signaling pathway.

    PubMed

    Dong, Menghua; Yang, Guiqing; Liu, Hanchen; Liu, Xiaoxu; Lin, Sixiang; Sun, Dongning; Wang, Yishan

    2014-03-01

    Accumulating evidence indicates that aged black garlic extract (ABGE) may prove beneficial in preventing or inhibiting oncogenesis; however, the underlying mechanisms have not been fully elucidated. The present study aimed to investigate the effects of ABGE on the proliferation and apoptosis of HT29 colon cancer cells. Our results demonstrated that ABGE inhibited HT29 cell growth via the induction of apoptosis and cell cycle arrest. We further investigated the phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) signal transduction pathway and the molecular mechanisms underlying the ABGE-induced inhibition of HT29 cell proliferation. We observed that ABGE may regulate the function of the PI3K/Akt pathway through upregulating PTEN and downregulating Akt and p-Akt expression, as well as suppressing its downstream target, 70-kDa ribosomal protein S6 kinase 1, at the mRNA and protein levels. In conclusion, these findings suggest that the PI3K/Akt signal transduction pathway is crucial for the development of colon cancer. ABGE inhibited the growth and induced apoptosis in HT29 cells through the inhibition of the PI3K/Akt pathway, suggesting that ABGE may be effective in the prevention and treatment of colon cancer in humans. PMID:24649105

  18. 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

  19. 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.

  20. Quantitative phosphoproteomic analysis of the PI3K-regulated signaling network.

    PubMed

    Gnad, Florian; Wallin, Jeffrey; Edgar, Kyle; Doll, Sophia; Arnott, David; Robillard, Liliane; Kirkpatrick, Donald S; Stokes, Matthew P; Vijapurkar, Ulka; Hatzivassiliou, Georgia; Friedman, Lori S; Belvin, Marcia

    2016-07-01

    The PI3K pathway is commonly activated in cancer. Only a few studies have attempted to explore the spectrum of phosphorylation signaling downstream of the PI3K cascade. Such insight, however, is imperative to understand the mechanisms responsible for oncogenic phenotypes. By applying MS-based phosphoproteomics, we mapped 2509 phosphorylation sites on 1096 proteins, and quantified their responses to activation or inhibition of PIK3CA using isogenic knock-in derivatives and a series of targeted inhibitors. We uncovered phosphorylation changes in a wide variety of proteins involved in cell growth and proliferation, many of which have not been previously associated with PI3K signaling. A significant update of the posttranslational modification database PHOSIDA (http://www.phosida.com) allows efficient use of the data. All MS data have been deposited in the ProteomeXchange with identifier PXD003899 (http://proteomecentral.proteomexchange.org/dataset/PXD003899). PMID:27282143

  1. Inhibition of PI3K Signalling Selectively Affects Medulloblastoma Cancer Stem Cells.

    PubMed

    Frasson, Chiara; Rampazzo, Elena; Accordi, Benedetta; Beggio, Giacomo; Pistollato, Francesca; Basso, Giuseppe; Persano, Luca

    2015-01-01

    Medulloblastoma is the most common malignant brain tumor of childhood. Although survival has slowly increased in the past years, the prognosis of these patients remains unfavourable. In this context, it has been recently shown that the intracellular signaling pathways activated during embryonic cerebellar development are deregulated in MDB. One of the most important is PI3K/AKT/mTOR, implicated in cell proliferation, survival, growth, and protein synthesis. Moreover, a fraction of MDB cells has been shown to posses stemlike features, to express typical neuronal precursor markers (Nestin and CD133), and to be maintained by the hypoxic cerebellar microenvironment. This subpopulation of MDB cells is considered to be responsible for treatment resistance and recurrence. In this study, we evaluated the effects of PI3K/AKT pathway inhibition on primary cultures of MDB and particularly on the cancer stem cell (CSC) population (CD133(+)). PI3K inhibition was able to counteract MDB cell growth and to promote differentiation of stemlike MDB cells. Moreover, PI3K/AKT pathway suppression induced dramatic cell death through activation of the mitochondrial proapoptotic cascade. Finally, analysis on the stem cells fraction revealed that the MDB CSC population is more sensitive to PI3K targeting compared to the whole cancerous population and its nonstem cell counterpart.

  2. PREX2 promotes the proliferation, invasion and migration of pancreatic cancer cells by modulating the PI3K signaling pathway

    PubMed Central

    Yang, Jianyi; Gong, Xuejun; Ouyang, Lu; He, Wen; Xiao, Rou; Tan, Li

    2016-01-01

    Phosphatidylinositol-3,4,5-trisphosphate-dependent Rac exchanger factor 2 (PREX2) is a novel regulator of the small guanosine triphosphatase Rac, and has been observed to be implicated in human cancer by inhibiting the activity of phosphatase and tensin homolog (PTEN), thus upregulating the activity of the phosphoinositide 3-kinase (PI3K) signaling pathway. However, the exact role of PREX2 in pancreatic cancer has not been reported to date. In the present study, the expression levels of PREX2 were observed to be frequently increased in pancreatic cancer specimens compared with those in their matched adjacent normal tissues. In addition, PREX2 expression was also frequently upregulated in several pancreatic cancer cell lines, including AsPC-1, BxPC-3, PANC-1 and CFAPC-1, compared with that in the normal pancreatic epithelial cell line HPC-Y5. Overexpression of PREX2 significantly promoted the proliferation, invasion and migration of pancreatic cancer PANC-1 cells, while small interfering RNA-induced knockdown of PREX2 expression significantly inhibited the proliferation, invasion and migration of these cells. Investigation of the molecular mechanism revealed that the overexpression of PREX2 upregulated the phosphorylation levels of PTEN, indicating that the activity of PTEN was reduced, which further increased the phosphorylation levels of AKT, which indicated that the activity of the PI3K signaling pathway was upregulated. By contrast, knockdown of PREX2 upregulated the activity of PTEN and inhibited the activity of the PI3K signaling pathway. In conclusion, the present study demonstrated that PREX2 regulates the proliferation, invasion and migration of pancreatic cancer cells, probably at least via modulation of the activity of PTEN and the PI3K signaling pathway. PMID:27446408

  3. Modulatory effects of yerba maté (Ilex paraguariensis) on the PI3K-AKT signaling pathway.

    PubMed

    Arçari, Demétrius Paiva; Santos, Juliana Carvalho; Gambero, Alessandra; Ferraz, Lucio Fábio Caldas; Ribeiro, Marcelo Lima

    2013-10-01

    The aim of this study was to evaluate the effects of yerba maté (YM) extract on the phosphatidylinositol 3-kinase (PI3K)-AKT signaling pathway in vivo. The mice were introduced to either standard- or high-fat diet (HFD). After 8 weeks on an HFD, mice were randomly assigned to one of the two treatment conditions, water or yerba maté extract at 1.0 g/kg. After treatment, glucose blood level and hepatic insulin response were evaluated. Liver tissue was examined to determine the mRNA levels using the PI3K-AKT PCR array. The nuclear translocation of forkhead box O1 (FOXO1) was determined by an electrophoretic mobility-shift assay. Our data demonstrated that yerba maté extract significantly decreased the final body weight, glucose blood levels, and insulin resistance of mice. Molecular analysis demonstrated that an HFD downregulated Akt2, Irs1, Irs2, Pi3kca, Pi3kcg, and Pdk1; after yerba maté treatment, the levels of those genes returned to baseline. In addition, an HFD upregulated Pepck and G6pc and increased FOXO1 nuclear translocation. The intervention downregulated these genes by decreasing FOXO1 nuclear translocation. The results obtained demonstrate for the first time the specific action of yerba maté on the PI3K-AKT pathway, which contributed to the observed improvement in hepatic insulin signaling.

  4. Leishmania promastigotes activate PI3K/Akt signalling to confer host cell resistance to apoptosis.

    PubMed

    Ruhland, Aaron; Leal, Nicole; Kima, Peter E

    2007-01-01

    Previous reports have shown that cells infected with promastigotes of some Leishmania species are resistant to the induction of apoptosis. This would suggest that either parasites elaborate factors that block signalling from apoptosis inducers or that parasites engage endogenous host signalling pathways that block apoptosis. To investigate the latter scenario, we determined whether Leishmania infection results in the activation of signalling pathways that have been shown to mediate resistance to apoptosis in other infection models. First, we showed that infection with the promastigote form of Leishmania major, Leishmania pifanoi and Leishmania amazonensis activates signalling through p38 mitogen-activated protein kinase (MAPK), NFkappaB and PI3K/Akt. Then we found that inhibition of signalling through the PI3K/Akt pathway with LY294002 and Akt IV inhibitor reversed resistance of infected bone marrow-derived macrophages and RAW 264.7 macrophages to potent inducers of apoptosis. Moreover, reduction of Akt levels with small interfering RNAs to Akt resulted in the inability of infected macrophages to resist apoptosis. Further evidence of the role of PI3K/Akt signalling in the promotion of cell survival by infected cells was obtained with the finding that Bad, which is a substrate of Akt, becomes phosphorylated during the course of infection. In contrast to the observations with PI3K/Akt signalling, inhibition of p38 MAPK signalling with SB202190 or NFkappaB signalling with wedelolactone had limited effect on parasite-induced resistance to apoptosis. We conclude that Leishmania promastigotes engage PI3K/Akt signalling, which confers to the infected cell, the capacity to resist death from activators of apoptosis.

  5. PTEN Contributes to Profound PI3K/Akt Signaling Pathway Deregulation in Dystrophin-Deficient Dog Muscle

    PubMed Central

    Feron, Marie; Guevel, Laetitia; Rouger, Karl; Dubreil, Laurence; Arnaud, Marie-Claire; Ledevin, Mireille; Megeney, Lynn A.; Cherel, Yan; Sakanyan, Vehary

    2009-01-01

    Duchenne muscular dystrophy is the most common and severe form of muscular dystrophy, and although the genetic basis of this disease is well defined, the overall mechanisms that define its pathogenesis remain obscure. Alterations in individual signaling pathways have been described, but little information is available regarding their putative implications in Duchenne muscular dystrophy pathogenesis. Here, we studied the status of various major signaling pathways in the Golden Retriever muscular dystrophy dog that specifically reproduces the full spectrum of human pathology. Using antibody arrays, we found that Akt1, glycogen synthase kinase-3β (GSK3β), 70-kDa ribosomal protein S6 kinase (p70S6K), extracellular signal-regulated kinases 1/2, and p38δ and p38γ kinases all exhibited decreased phosphorylation in muscle from a 4-month-old animal with Golden Retriever muscular dystrophy, revealing a deep alteration of the phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen-activated protein kinase pathways. Immunohistochemistry analysis revealed the presence of muscle fibers exhibiting a cytosolic accumulation of Akt1, GSK3β, and phosphatidylinositol-3,4,5-trisphosphate 3-phosphatase (PTEN), an enzyme counteracting PI3K-mediated Akt activation. Enzymatic assays established that these alterations in phosphorylation and expression levels were associated with decreased Akt and increased GSK3β and PTEN activities. PTEN/GSK3β-positive fibers were also observed in muscle sections from 3- and 36-month-old animals, indicating long-term PI3K/Akt pathway alteration. Collectively, our data suggest that increased PTEN expression and activity play a central role in PI3K/Akt/GSK3β and p70S6K pathway modulation, which could exacerbate the consequences of dystrophin deficiency. PMID:19264909

  6. Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes.

    PubMed

    Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

    2014-11-01

    In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes.

  7. Targeting the PI3K/Akt signaling pathway in gastric carcinoma: A reality for personalized medicine?

    PubMed Central

    Singh, Shikha Satendra; Yap, Wei Ney; Arfuso, Frank; Kar, Shreya; Wang, Chao; Cai, Wanpei; Dharmarajan, Arunasalam M; Sethi, Gautam; Kumar, Alan Prem

    2015-01-01

    Frequent activation of phosphatidylinositol-3 kinases (PI3K)/Akt/mTOR signaling pathway in gastric cancer (GC) is gaining immense popularity with identification of mutations and/or amplifications of PIK3CA gene or loss of function of PTEN, a tumor suppressor protein, to name a few; both playing a crucial role in regulating this pathway. These aberrations result in dysregulation of this pathway eventually leading to gastric oncogenesis, hence, there is a need for targeted therapy for more effective anticancer treatment. Several inhibitors are currently in either preclinical or clinical stages for treatment of solid tumors like GC. With so many inhibitors under development, further studies on predictive biomarkers are needed to measure the specificity of any therapeutic intervention. Herein, we review the common dysregulation of PI3K/Akt/mTOR pathway in GC and the various types of single or dual pathway inhibitors under development that might have a superior role in GC treatment. We also summarize the recent developments in identification of predictive biomarkers and propose use of predictive biomarkers to facilitate more personalized cancer therapy with effective PI3K/Akt/mTOR pathway inhibition. PMID:26604635

  8. Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes.

    PubMed

    Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

    2014-11-01

    In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes. PMID:25489416

  9. GLCCI1 is a novel component associated with the PI3K signaling pathway in podocyte foot processes

    PubMed Central

    Kim, Sang-Hoon; Kim, Hyun-Jung; Kim, Chan-Wha

    2016-01-01

    Podocyte foot processes are interdigitated to form the slit diaphragm and are crucial for the glomerular filtration barrier. Glucocorticoid-induced transcript 1 (GLCCI1) is transcriptionally regulated, but its signaling pathway in podocytes is unknown. The main objective of this study was to investigate the regulation of podocyte foot process proteins and to investigate the role of GLCCI1 in the phosphoinositide 3-kinase (PI3K) pathway using high glucose-induced podocytes and streptozotocin-induced diabetic rats. In podocytes and rat kidneys, GLCCI1 was found to be highly specific for the glomerulus and podocyte foot processes similar to other podocyte-specific proteins (nephrin, podocin, synatopodin and podocalyxin) based on reverse transcription-PCR, western blotting, immunofluorescence and immunoelectron microscopy analyses. In addition, the decrease in the GLCCI1 expression level under hyperglycemic conditions was restored by treatment with a PI3K inhibitor (wortmannin). Immunofluorescence analysis confirmed that GLCCI1 colocalized with nephrin and synaptopodin both in vivo and in vitro. Finally, immunoelectron microscopy data from streptozotocin-induced diabetic rats showed that GLCCI1 also localized in podocyte foot processes. Hence, GLCCI1 is a component of podocyte foot processes, and its expression appears to be regulated via the PI3K pathway. PMID:27174202

  10. Arctigenin Increases Hemeoxygenase-1 Gene Expression by Modulating PI3K/AKT Signaling Pathway in Rat Primary Astrocytes

    PubMed Central

    Jeong, Yeon-Hui; Park, Jin-Sun; Kim, Dong-Hyun; Kim, Hee-Sun

    2014-01-01

    In the present study, we found that the natural compound arctigenin inhibited hydrogen peroxide-induced reactive oxygen species (ROS) production in rat primary astrocytes. Since hemeoxygenase-1 (HO-1) plays a critical role as an antioxidant defense factor in the brain, we examined the effect of arctigenin on HO-1 expression in rat primary astrocytes. We found that arctigenin increased HO-1 mRNA and protein levels. Arctigenin also increases the nuclear translocation and DNA binding of Nrf2/c-Jun to the antioxidant response element (ARE) on HO-1 promoter. In addition, arctigenin increased ARE-mediated transcriptional activities in rat primary astrocytes. Further mechanistic studies revealed that arctigenin increased the phosphorylation of AKT, a downstream substrate of phosphatidylinositol 3-kinase (PI3K). Treatment of cells with a PI3K-specific inhibitor, LY294002, suppressed the HO-1 expression, Nrf2 DNA binding and ARE-mediated transcriptional activities in arctigenin-treated astrocyte cells. The results collectively suggest that PI3K/AKT signaling pathway is at least partly involved in HO-1 expression by arctigenin via modulation of Nrf2/ARE axis in rat primary astrocytes. PMID:25489416

  11. Oocyte-specific deletion of Pten in mice reveals a stage-specific function of PTEN/PI3K signaling in oocytes in controlling follicular activation.

    PubMed

    Jagarlamudi, Krishna; Liu, Lian; Adhikari, Deepak; Reddy, Pradeep; Idahl, Annika; Ottander, Ulrika; Lundin, Eva; Liu, Kui

    2009-07-09

    Immature ovarian primordial follicles are essential for maintenance of the reproductive lifespan of female mammals. Recently, it was found that overactivation of the phosphatidylinositol 3-kinase (PI3K) signaling in oocytes of primordial follicles by an oocyte-specific deletion of Pten (phosphatase and tensin homolog deleted on chromosome ten), the gene encoding PI3K negative regulator PTEN, results in premature activation of the entire pool of primordial follicles, indicating that activation of the PI3K pathway in oocytes is important for control of follicular activation. To investigate whether PI3K signaling in oocytes of primary and further developed follicles also plays a role at later stages in follicular development and ovulation, we conditionally deleted the Pten gene from oocytes of primary and further developed follicles by using transgenic mice expressing zona pellucida 3 (Zp3) promoter-mediated Cre recombinase. Our results show that Pten was efficiently deleted from oocytes of primary and further developed follicles, as indicated by the elevated phosphorylation of the major PI3K downstream component Akt. However, follicular development was not altered and oocyte maturation was also normal, which led to normal fertility with unaltered litter size in the mutant mice. Our data indicate that properly controlled PTEN/PI3K-Akt signaling in oocytes is essential for control of the development of primordial follicles whereas overactivation of PI3K signaling in oocytes does not appear to affect the development of growing follicles. This suggests that there is a stage-specific function of PTEN/PI3K signaling in mouse oocytes that controls follicular activation.

  12. P-REX1 creates a positive feedback loop to activate growth factor receptor, PI3K/AKT, and MEK/ERK signaling in breast cancer

    PubMed Central

    Dillon, Lloye M.; Bean, Jennifer R.; Yang, Wei; Shee, Kevin; Symonds, Lynn K.; Balko, Justin M.; McDonald, W. Hayes; Liu, Shuying; Gonzalez-Angulo, Ana M.; Mills, Gordon B.; Arteaga, Carlos L.; Miller, Todd W.

    2014-01-01

    Phosphatidylinositol 3-kinase (PI3K) promotes cancer cell survival, migration, growth, and proliferation by generating phosphatidylinositol 3,4,5-trisphosphate (PIP3) in the inner leaflet of the plasma membrane. PIP3 recruits pleckstrin homology (PH) domain-containing proteins to the membrane to activate oncogenic signaling cascades. Anti-cancer therapeutics targeting the PI3K/AKT/mTOR pathway are in clinical development. In a mass spectrometric screen to identify PIP3-regulated proteins in breast cancer cells, levels of the Rac activator PIP3-dependent Rac exchange factor 1 (P-REX1) increased in response to PI3K inhibition, and decreased upon loss of the PI3K antagonist PTEN. P-REX1 mRNA and protein levels were positively correlated with ER expression, and inversely correlated with PI3K pathway activation in breast tumors as assessed by gene expression and phosphoproteomic analyses. P-REX1 increased activation of Rac1, PI3K/AKT, and MEK/ERK signaling in a PTEN-independent manner, and promoted cell and tumor viability. Loss of P-REX1 or inhibition of Rac suppressed PI3K/AKT and MEK/ERK, and decreased viability. P-REX1 also promoted insulin-like growth factor-1 receptor (IGF-1R) activation, suggesting that P-REX1 provides positive feedback to activators upstream of PI3K. In support of a model where PIP3-driven P-REX1 promotes both PI3K/AKT and MEK/ERK signaling, high levels of P-REX1 mRNA (but not phospho-AKT or a transcriptomic signature of PI3K activation) were predictive of sensitivity to PI3K inhibitors among breast cancer cell lines. P-REX1 expression was highest in ER+ breast tumors compared to many other cancer subtypes, suggesting that neutralizing the P-REX1/Rac axis may provide a novel therapeutic approach to selectively abrogate oncogenic signaling in breast cancer cells. PMID:25284585

  13. Activation of PI3K/Akt signaling has a dominant negative effect on IL-12 production by macrophages infected with Leishmania amazonensis promastigotes

    PubMed Central

    Ruhland, Aaron; Kima, Peter E.

    2009-01-01

    Infection of macrophages with Leishmania parasites does not result in the production of IL-12. In addition, infection with Leishmania suppresses IL-12 production elicited by otherwise potent activators of IL-12. We provide evidence that engagement of phosphatidyl inositol-3 kinase (PI3K) signaling during Leishmania amazonensis infection leads to the prevention of IL-12 p70 production at the level of transcription of its p40 subunit in bone marrow derived macrophages (BMDMϕ). Inhibition of PI3K signaling with specific inhibitors of PI3K or the downstream kinase Akt, reverses the IL-12 blockade. Although the MAP kinase ERK (p44 and p42) was transiently activated by infection with L. amazonensis, inhibition of MEK, the kinase upstream of ERK, with PD98059, did not reverse the blockade of IL-12. Furthermore, inhibition of the other MAP kinases JNK and p38 as well as treatment of cells with pertussis toxin that blocks G protein mediated signaling, did not reverse the prevention of IL-12 production by Leishmania infection. Interestingly, activation of PI3K/Akt signaling had differential effects on ERK and p38 activation. Taken together we propose that infection of BMDMϕ with Leishmania promastigotes activates both positive and negative signaling pathways that control IL-12 production. PI3K signaling activated by the infection is the negative signaling pathway that prevents IL-12 production. PMID:19186178

  14. Activation of PI3K/Akt signaling has a dominant negative effect on IL-12 production by macrophages infected with Leishmania amazonensis promastigotes.

    PubMed

    Ruhland, Aaron; Kima, Peter E

    2009-05-01

    Infection of macrophages with Leishmania parasites does not result in the production of IL-12. In addition, infection with Leishmania suppresses IL-12 production elicited by otherwise potent activators of IL-12. We provide evidence that engagement of phosphatidyl inositol-3 kinase (PI3K) signaling during Leishmania amazonensis infection leads to the prevention of IL-12 p70 production at the level of transcription of its p40 subunit in bone marrow derived macrophages (BMDMPhi). Inhibition of PI3K signaling with specific inhibitors of PI3K or the downstream kinase Akt, reverses the IL-12 blockade. Although the MAP kinase ERK (p44 and p42) was transiently activated by infection with L. amazonensis, inhibition of MEK, the kinase upstream of ERK, with PD98059, did not reverse the blockade of IL-12. Furthermore, inhibition of the other MAP kinases JNK and p38 as well as treatment of cells with pertussis toxin that blocks G protein mediated signaling, did not reverse the prevention of IL-12 production by Leishmania infection. Interestingly, activation of PI3K/Akt signaling had differential effects on ERK and p38 activation. Taken together we propose that infection of BMDMPhi with Leishmania promastigotes activates both positive and negative signaling pathways that control IL-12 production. PI3K signaling activated by the infection is the negative signaling pathway that prevents IL-12 production.

  15. Imbalanced PTEN and PI3K Signaling Impairs Class Switch Recombination.

    PubMed

    Chen, Zhangguo; Getahun, Andrew; Chen, Xiaomi; Dollin, Yonatan; Cambier, John C; Wang, Jing H

    2015-12-01

    Class switch recombination (CSR) generates isotype-switched Abs with distinct effector functions. B cells express phosphatase and tensin homolog (PTEN) and multiple isoforms of class IA PI3K catalytic subunits, including p110α and p110δ, whose roles in CSR remain unknown or controversial. In this article, we demonstrate a direct effect of PTEN on CSR signaling by acute deletion of Pten specifically in mature B cells, thereby excluding the developmental impact of Pten deletion. We show that mature B cell-specific PTEN overexpression enhances CSR. More importantly, we establish a critical role for p110α in CSR. Furthermore, we identify a cooperative role for p110α and p110δ in suppressing CSR. Mechanistically, dysregulation of p110α or PTEN inversely affects activation-induced deaminase expression via modulating AKT activity. Thus, our study reveals that a signaling balance between PTEN and PI3K isoforms is essential to maintain normal CSR.

  16. 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

  17. Involvement of the PI3K and ERK signaling pathways in largemouth bass virus-induced apoptosis and viral replication.

    PubMed

    Huang, Xiaohong; Wang, Wei; Huang, Youhua; Xu, Liwen; Qin, Qiwei

    2014-12-01

    Increased reports demonstrated that largemouth Bass, Micropterus salmoides in natural and artificial environments were always suffered from an emerging iridovirus disease, largemouth Bass virus (LMBV). However, the underlying mechanism of LMBV pathogenesis remained largely unknown. Here, we investigated the cell signaling events involved in virus induced cell death and viral replication in vitro. We found that LMBV infection in epithelioma papulosum cyprini (EPC) cells induced typical apoptosis, evidenced by the appearance of apoptotic bodies, cytochrome c release, mitochondrial membrane permeabilization (MMP) destruction and reactive oxygen species (ROS) generation. Two initiators of apoptosis, caspase-8 and caspase-9, and the executioner of apoptosis, caspase-3, were all significantly activated with the infection time, suggested that not only mitochondrion-mediated, but also death receptor-mediated apoptosis were involved in LMBV infection. Reporter gene assay showed that the promoter activity of transcription factors including p53, NF-κB, AP-1 and cAMP response element-binding protein (CREB) were decreased during LMBV infection. After treatment with different signaling pathway inhibitors, virus production were significantly suppressed by the inhibition of phosphatidylinositol 3-kinase (PI3K) pathway and extracellular-signal-regulated kinases (ERK) signaling pathway. Furthermore, LMBV infection induced apoptosis was enhanced by PI3K inhibitor LY294002, but decreased by addition of ERK inhibitor UO126. Therefore, we speculated that apoptosis was sophisticatedly regulated by a series of cell signaling events for efficient virus propagation. Taken together, our results provided new insights into the molecular mechanism of ranavirus infection. PMID:25260912

  18. Involvement of the PI3K and ERK signaling pathways in largemouth bass virus-induced apoptosis and viral replication.

    PubMed

    Huang, Xiaohong; Wang, Wei; Huang, Youhua; Xu, Liwen; Qin, Qiwei

    2014-12-01

    Increased reports demonstrated that largemouth Bass, Micropterus salmoides in natural and artificial environments were always suffered from an emerging iridovirus disease, largemouth Bass virus (LMBV). However, the underlying mechanism of LMBV pathogenesis remained largely unknown. Here, we investigated the cell signaling events involved in virus induced cell death and viral replication in vitro. We found that LMBV infection in epithelioma papulosum cyprini (EPC) cells induced typical apoptosis, evidenced by the appearance of apoptotic bodies, cytochrome c release, mitochondrial membrane permeabilization (MMP) destruction and reactive oxygen species (ROS) generation. Two initiators of apoptosis, caspase-8 and caspase-9, and the executioner of apoptosis, caspase-3, were all significantly activated with the infection time, suggested that not only mitochondrion-mediated, but also death receptor-mediated apoptosis were involved in LMBV infection. Reporter gene assay showed that the promoter activity of transcription factors including p53, NF-κB, AP-1 and cAMP response element-binding protein (CREB) were decreased during LMBV infection. After treatment with different signaling pathway inhibitors, virus production were significantly suppressed by the inhibition of phosphatidylinositol 3-kinase (PI3K) pathway and extracellular-signal-regulated kinases (ERK) signaling pathway. Furthermore, LMBV infection induced apoptosis was enhanced by PI3K inhibitor LY294002, but decreased by addition of ERK inhibitor UO126. Therefore, we speculated that apoptosis was sophisticatedly regulated by a series of cell signaling events for efficient virus propagation. Taken together, our results provided new insights into the molecular mechanism of ranavirus infection.

  19. Too much of a good thing: immunodeficiency due to hyperactive PI3K signaling.

    PubMed

    Walsh, Craig M; Fruman, David A

    2014-09-01

    Primary immune deficiency diseases arise due to heritable defects that often involve signaling molecules required for immune cell function. Typically, these genetic defects cause loss of gene function, resulting in primary immune deficiencies such as severe combined immune deficiency (SCID) and X-linked agammaglobulinemia (XLA); however, gain-of-function mutations may also promote immune deficiency. In this issue of the JCI, Deau et al. establish that gain-of-function mutations in PIK3R1, which encodes the p85α regulatory subunit of class IA PI3Ks, lead to immunodeficiency. These observations are consistent with previous reports that hyperactivating mutations in PIK3CD, which encodes the p110δ catalytic subunit, are capable of promoting immune deficiency. Mutations that reduce PI3K activity also result in defective lymphocyte development and function; therefore, these findings support the notion that too little or too much PI3K activity leads to immunodeficiency.

  20. TEIF associated centrosome activity is regulated by EGF/PI3K/Akt signaling.

    PubMed

    Zhao, Jing; Zou, Yongxin; Liu, Haijing; Wang, Huali; Zhang, Hong; Hou, Wei; Li, Xin; Jia, Xinying; Zhang, Jing; Hou, Lin; Zhang, Bo

    2014-09-01

    Centrosome amplification, which is a characteristic of cancer cells, has been understood as a driving force of genetic instability in the development of cancer. In previous work, we demonstrated that TEIF (transcriptional element-interacting factor) distributes in the centrosomes and regulates centrosome status under both physiologic and pathologic conditions. Here we identify TEIF as a downstream effector in EGF/PI3K/Akt signaling. The addition of EGF or transfection of active Akt stimulates centrosome TEIF distribution, resulting in an increase of centrosome splitting and amplification, while inhibitors of either PI3K or Akt attenuate these changes in TEIF and the associated centrosome status. A consensus motif for Akt phosphorylation (RHRVLT) proved to be involved in centrosomal TEIF localization, and the 469-threonine of this motif may be phosphorylated by Akt both in vitro and in vivo. Elimination of this phosphorylated site on TEIF caused reduced centrosome distribution and centrosome splitting or amplification. Moreover, TEIF closely co-localized with C-NAP1 at the proximal ends of centrioles, and centriolar loading of TEIF stimulated by EGF/Akt could displace C-NAP1, resulting in centrosome splitting. These findings reveal linkage of the EGF/PI3K/Akt signaling pathway to regulation of centrosome status which may act as an oncogenic pathway and induce genetic instability in carcinogenesis. PMID:24769208

  1. 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.

  2. Low-Dose Endothelial Monocyte-Activating Polypeptide-II Increases Blood-Tumor Barrier Permeability by Activating the RhoA/ROCK/PI3K Signaling Pathway.

    PubMed

    Li, Zhen; Liu, Xiao-Bai; Liu, Yun-Hui; Xue, Yi-Xue; Liu, Jing; Teng, Hao; Xi, Zhuo; Yao, Yi-Long

    2016-06-01

    Previous studies have demonstrated that low-dose endothelial monocyte-activating polypeptide-II (EMAP-II) can increase blood-tumor barrier (BTB) permeability via both paracellular and transcellular pathways. In addition, we revealed that the RhoA/Rho kinase (ROCK) signaling pathway is involved in EMAP-II-induced BTB opening. This study further investigated the exact mechanisms by which the RhoA/ROCK signaling pathway affects EMAP-II-induced BTB hyperpermeability. In an in vitro BTB model, low-dose EMAP-II significantly activated phosphatidylinositol-3-kinase (PI3K) in rat brain microvascular endothelial cells (RBMECs) at 0.75 h. Pretreatment with RhoA inhibitor C3 exoenzyme or ROCK inhibitor Y-27632 completely blocked EMAP-II-induced activation of PI3K. PKC-α/β inhibitor GÖ6976 pretreatment caused no change in EMAP-II-induced activation of PI3K. Besides, pretreatment with LY294002, a specific inhibitor of PI3K, did not affect EMAP-II-induced activation of PKC-α/β. Furthermore, LY294002 pretreatment significantly diminished EMAP-II-induced changes in BTB permeability, phosphorylation of myosin light chain and cofilin, expression and distribution of tight junction-associated protein ZO-1, and actin cytoskeleton arrangement in RBMECs. In summary, this study demonstrates that low-dose EMAP-II can increase BTB permeability by activating the RhoA/ROCK/PI3K signaling pathway.

  3. Ramentaceone, a Naphthoquinone Derived from Drosera sp., Induces Apoptosis by Suppressing PI3K/Akt Signaling in Breast Cancer Cells

    PubMed Central

    Kawiak, Anna; Lojkowska, Ewa

    2016-01-01

    The phosphoinositide 3-kinase (PI3K) signaling pathway plays an important role in processes critical for breast cancer progression and its upregulation confers increased resistance of cancer cells to chemotherapy and radiation. The present study aimed at determining the activity of ramentaceone, a constituent of species in the plant genera Drosera, toward breast cancer cells and defining the involvement of PI3K/Akt inhibition in ramentaceone-mediated cell death induction. The results showed that ramentaceone exhibited high antiproliferative activity toward breast cancer cells, in particular HER2-overexpressing breast cancer cells. The mode of cell death induced by ramentaceone was through apoptosis as determined by cytometric analysis of caspase activity and Annexin V staining. Apoptosis induction was found to be mediated by inhibition of PI3K/Akt signaling and through targeting its downstream anti-apoptotic effectors. Ramentaceone inhibited PI3-kinase activity, reduced the expression of the PI3K protein and inhibited the phosphorylation of the Akt protein in breast cancer cells. The expression of the anti-apoptotic Bcl-2 protein was decreased and the levels of the pro-apoptotic proteins, Bax and Bak, were elevated. Moreover, inhibition of PI3K and silencing of Akt expression increased the sensitivity of cells to ramentaceone-induced apoptosis. In conclusion, our results indicate that ramentaceone induces apoptosis in breast cancer cells through PI3K/Akt signaling inhibition. These findings suggest further investigation of ramentaceone as a potential therapeutic agent in breast cancer therapy, in particular HER2-positive breast cancer. PMID:26840401

  4. Optimal targeting of HER2-PI3K signaling in breast cancer: mechanistic insights and clinical implications.

    PubMed

    Rexer, Brent N; Arteaga, Carlos L

    2013-07-01

    The combination of a PI3K inhibitor with trastuzumab has been shown to be effective at overcoming trastuzumab resistance in models of HER2(+) breast cancer by inhibiting HER2-PI3K-FOXO-survivin signaling. In this review the potential clinical implications of these findings are discussed.

  5. 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

  6. Ursolic Acid Increases Glucose Uptake through the PI3K Signaling Pathway in Adipocytes

    PubMed Central

    He, Yonghan; Li, Wen; Li, Ying; Zhang, Shuocheng; Wang, Yanwen; Sun, Changhao

    2014-01-01

    Background Ursolic acid (UA), a triterpenoid compound, is reported to have a glucose-lowering effect. However, the mechanisms are not fully understood. Adipose tissue is one of peripheral tissues that collectively control the circulating glucose levels. Objective The objective of the present study was to determine the effect and further the mechanism of action of UA in adipocytes. Methods and Results The 3T3-L1 preadipocytes were induced to differentiate and treated with different concentrations of UA. NBD-fluorescent glucose was used as the tracer to measure glucose uptake and Western blotting used to determine the expression and activity of proteins involved in glucose transport. It was found that 2.5, 5 and 10 µM of UA promoted glucose uptake in a dose-dependent manner (17%, 29% and 35%, respectively). 10 µM UA-induced glucose uptake with insulin stimulation was completely blocked by the phosphatidylinositol (PI) 3-kinase (PI3K) inhibitor wortmannin (1 µM), but not by SB203580 (10 µM), the inhibitor of mitogen-activated protein kinase (MAPK), or compound C (2.5 µM), the inhibitor of AMP-activated kinase (AMPK) inhibitor. Furthmore, the downstream protein activities of the PI3K pathway, phosphoinositide-dependent kinase (PDK) and phosphoinositide-dependent serine/threoninekinase (AKT) were increased by 10 µM of UA in the presence of insulin. Interestingly, the activity of AS160 and protein kinase C (PKC) and the expression of glucose transporter 4 (GLUT4) were stimulated by 10 µM of UA under either the basal or insulin-stimulated status. Moreover, the translocation of GLUT4 from cytoplasm to cell membrane was increased by UA but decreased when the PI3K inhibitor was applied. Conclusions Our results suggest that UA stimulates glucose uptake in 3T3-L1 adipocytes through the PI3K pathway, providing important information regarding the mechanism of action of UA for its anti-diabetic effect. PMID:25329874

  7. Vitamin E succinate induces apoptosis via the PI3K/AKT signaling pathways in EC109 esophageal cancer cells

    PubMed Central

    Yang, Peng; Zhao, Jiaying; Hou, Liying; Yang, Lei; Wu, Kun; Zhang, Linyou

    2016-01-01

    Esophageal cancer is the fourth most common gastrointestinal cancer, it generally has a poor prognosis and novel strategies are required for prevention and treatment. Vitamin E succinate (VES) is a potential chemical agent for cancer prevention and therapy as it exerts anti-tumor effects in a variety of cancers. However, the role of VES in tumorigenesis and progression of cancer remains to be elucidated. The present study aimed to determine the effects of VES in regulating the survival and apoptosis of human esophageal cancer cells. EC109 human esophageal cancer cells were used to investigate the anti-proliferative effects of VES. The MTT and Annexin V-fluorescein isothiocyanate/propidium iodide assays demonstrated that VES inhibited cell proliferation and induced apoptosis in esophageal cancer cells. Furthermore, VES downregulated constitutively active basal levels of phosphorylated (p)-serine-threonine kinase AKT (AKT) and p-mammalian target of rapamycin (mTOR), and decreased the phosphorylation of AKT substrates Bcl-2-associated death receptor and caspase-9, in addition to mTOR effectors, ribosomal protein S6 kinase β1 and eIF4E-binding protein 1. Phosphoinositide-3-kinase (PI3K) inhibitor, LY294002 suppressed p-AKT and p-mTOR, indicating PI3K is a common upstream mediator. The apoptosis induced by VES was increased by inhibition of AKT or mTOR with their respective inhibitor in esophageal cancer cells. The results of the present study suggested that VES targeted the PI3K/AKT signaling pathways and induced apoptosis in esophageal cancer cells. Furthermore, the current study suggests that VES may be useful in a combinational therapeutic strategy employing an mTOR inhibitor. PMID:27357907

  8. Crosstalk Between MAPK/ERK and PI3K/AKT Signal Pathways During Brain Ischemia/Reperfusion

    PubMed Central

    Zhou, Jing; Du, Ting; Li, Baoman; Rong, Yan; Verkhratsky, Alexei

    2015-01-01

    The epidermal growth factor receptor (EGFR) is linked to the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) and Raf/mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK1/2) signaling pathways. During brain ischemia/reperfusion, EGFR could be transactivated, which stimulates these intracellular signaling cascades that either protect cells or potentiate cell injury. In the present study, we investigated the activation of EGFR, PI3K/AKT, and Raf/MAPK/ERK1/2 during ischemia or reperfusion of the brain using the middle cerebral artery occlusion model. We found that EGFR was phosphorylated and transactivated during both ischemia and reperfusion periods. During ischemia, the activity of PI3K/AKT pathway was significantly increased, as judged from the strong phosphorylation of AKT; this activation was suppressed by the inhibitors of EGFR and Zn-dependent metalloproteinase. Ischemia, however, did not induce ERK1/2 phosphorylation, which was dependent on reperfusion. Coimmunoprecipitation of Son of sevenless 1 (SOS1) with EGFR showed increased association between the receptor and SOS1 in ischemia, indicating the inhibitory node downstream of SOS1. The inhibitory phosphorylation site of Raf-1 at Ser259, but not its stimulatory phosphorylation site at Ser338, was phosphorylated during ischemia. Furthermore, ischemia prompted the interaction between Raf-1 and AKT, while both the inhibitors of PI3K and AKT not only abolished AKT phosphorylation but also restored ERK1/2 phosphorylation. All these findings suggest that Raf/MAPK/ERK1/2 signal pathway is inhibited by AKT via direct phosphorylation and inhibition at Raf-1 node during ischemia. During reperfusion, we observed a significant increase of ERK1/2 phosphorylation but no change in AKT phosphorylation. Inhibitors of reactive oxygen species and phosphatase and tensin homolog restored AKT phosphorylation but abolished ERK1/2 phosphorylation, suggesting that the reactive oxygen species

  9. Optimization of the phenylurea moiety in a phosphoinositide 3-kinase (PI3K) inhibitor to improve water solubility and the PK profile by introducing a solubilizing group and ortho substituents.

    PubMed

    Kawada, Hatsuo; Ebiike, Hirosato; Tsukazaki, Masao; Yamamoto, Shun; Koyama, Kohei; Nakamura, Mitsuaki; Morikami, Kenji; Yoshinari, Kiyoshi; Yoshida, Miyuki; Ogawa, Kotaro; Shimma, Nobuo; Tsukuda, Takuo; Ohwada, Jun

    2016-07-01

    Phosphoinositide 3-kinase (PI3K) is a promising anti-cancer target, because various mutations and amplifications are observed in human tumors isolated from cancer patients. Our dihydropyrrolopyrimidine derivative with a phenylurea moiety showed strong PI3K enzyme inhibitory activity, but its pharmacokinetic property was poor because of lack of solubility. Herein, we report how we improved the solubility of our PI3K inhibitors by introducing a solubilizing group and ortho substituents to break molecular planarity. PMID:27189888

  10. Activation of PI3K signaling prevents aminoglycoside-induced hair cell death in the murine cochlea.

    PubMed

    Jadali, Azadeh; Kwan, Kelvin Y

    2016-01-01

    Loss of sensory hair cells of the inner ear due to aminoglycoside exposure is a major cause of hearing loss. Using an immortalized multipotent otic progenitor (iMOP) cell line, specific signaling pathways that promote otic cell survival were identified. Of the signaling pathways identified, the PI3K pathway emerged as a strong candidate for promoting hair cell survival. In aging animals, components for active PI3K signaling are present but decrease in hair cells. In this study, we determined whether activated PI3K signaling in hair cells promotes survival. To activate PI3K signaling in hair cells, we used a small molecule inhibitor of PTEN or genetically ablated PTEN using a conditional knockout animal. Hair cell survival was challenged by addition of gentamicin to cochlear cultures. Hair cells with activated PI3K signaling were more resistant to aminoglycoside-induced hair cell death. These results indicate that increased PI3K signaling in hair cells promote survival and the PI3K signaling pathway is a target for preventing aminoglycoside-induced hearing loss.

  11. Activation of PI3K signaling prevents aminoglycoside-induced hair cell death in the murine cochlea

    PubMed Central

    Jadali, Azadeh

    2016-01-01

    ABSTRACT Loss of sensory hair cells of the inner ear due to aminoglycoside exposure is a major cause of hearing loss. Using an immortalized multipotent otic progenitor (iMOP) cell line, specific signaling pathways that promote otic cell survival were identified. Of the signaling pathways identified, the PI3K pathway emerged as a strong candidate for promoting hair cell survival. In aging animals, components for active PI3K signaling are present but decrease in hair cells. In this study, we determined whether activated PI3K signaling in hair cells promotes survival. To activate PI3K signaling in hair cells, we used a small molecule inhibitor of PTEN or genetically ablated PTEN using a conditional knockout animal. Hair cell survival was challenged by addition of gentamicin to cochlear cultures. Hair cells with activated PI3K signaling were more resistant to aminoglycoside-induced hair cell death. These results indicate that increased PI3K signaling in hair cells promote survival and the PI3K signaling pathway is a target for preventing aminoglycoside-induced hearing loss. PMID:27142333

  12. Dimethyl Cardamonin Exhibits Anti-inflammatory Effects via Interfering with the PI3K-PDK1-PKCα Signaling Pathway

    PubMed Central

    Yu, Wan-Guo; He, Hao; Yao, Jing-Yun; Zhu, Yi-Xiang; Lu, Yan-Hua

    2015-01-01

    Consumption of herbal tea [flower buds of Cleistocalyx operculatus (Roxb.) Merr. et Perry (Myrtaceae)] is associated with health beneficial effects against multiple diseases including diabetes, asthma, and inflammatory bowel disease. Emerging evidences have reported that High mobility group box 1 (HMGB1) is considered as a key “late” proinflammatory factor by its unique secretion pattern in aforementioned diseases. Dimethyl cardamonin (2′,4′-dihydroxy-6′-methoxy-3′,5′-dimethylchalcone, DMC) is a major ingredient of C. operculatus flower buds. In this study, the anti-inflammatory effects of DMC and its underlying molecular mechanisms were investigated on lipopolysaccharide (LPS)-induced macrophages. DMC notably suppressed the mRNA expressions of TNF-α, IL-1β, IL-6, and HMGB1, and also markedly decreased their productions in a time- and dose-dependent manner. Intriguingly, DMC could notably reduce LPS-stimulated HMGB1 secretion and its nucleo-cytoplasmic translocation. Furthermore, DMC dose-dependently inhibited the activation of phosphatidylinositol 3-kinase (PI3K), phosphoinositide-dependent kinase 1 (PDK1), and protein kinase C alpha (PKCα). All these data demonstrated that DMC had anti-inflammatory effects through reducing both early (TNF-α, IL-1β, and IL-6) and late (HMGB1) cytokines expressions via interfering with the PI3K-PDK1-PKCα signaling pathway. PMID:26535080

  13. PI3K-AKT signaling is a downstream effector of retinoid prevention of murine basal cell carcinogenesis.

    PubMed

    So, Po-Lin; Wang, Grace Y; Wang, Kevin; Chuang, Mindy; Chiueh, Venice Calinisan; Kenny, Paraic A; Epstein, Ervin H

    2014-04-01

    Basal cell carcinoma (BCC) is the most common human cancer. We have demonstrated previously that topical application of the retinoid prodrug tazarotene profoundly inhibits murine BCC carcinogenesis via retinoic acid receptor γ-mediated regulation of tumor cell transcription. Because topical retinoids can cause adverse cutaneous effects and because tumors can develop resistance to retinoids, we have investigated mechanisms downstream of tazarotene's antitumor effect in this model. Specifically we have used (i) global expression profiling to identify and (ii) functional cell-based assays to validate the phosphoinositide 3-kinase (PI3K)/AKT/mTOR pathway as a downstream target pathway of tazarotene's action. Crucially, we have demonstrated that pharmacologic inhibition of this downstream pathway profoundly reduces murine BCC cell proliferation and tumorigenesis both in vitro and in vivo. These data identify PI3K/AKT/mTOR signaling as a highly attractive target for BCC chemoprevention and indicate more generally that this pathway may be, in some contexts, an important mediator of retinoid anticancer effects.

  14. Cell type-specific dependency on the PI3K/Akt signaling pathway for the endogenous Epo and VEGF induction by baicalein in neurons versus astrocytes.

    PubMed

    Sun, Yu-Yo; Lin, Shang-Hsuan; Lin, Hung-Cheng; Hung, Chia-Chi; Wang, Chen-Yu; Lin, Yen-Chu; Hung, Kuo-Sheng; Lien, Cheng-Chang; Kuan, Chia-Yi; Lee, Yi-Hsuan

    2013-01-01

    The neuroprotective effect of baicalein is generally attributed to inhibition of 12/15-lipoxygenase (12/15-LOX) and suppression of oxidative stress, but recent studies showed that baicalein also activates hypoxia-inducible factor-α (HIF1α) through inhibition of prolyl hydrolase 2 (PHD2) and activation of the phosphatidylinositide-3 kinase (PI3K)/Akt signaling pathway. Yet, the significance and regulation of prosurvival cytokines erythropoietin (Epo) and vascular endothelial growth factor (VEGF), two transcriptional targets of HIF1α, in baicalein-mediated neuroprotection in neurons and astrocytes remains unknown. Here we investigated the causal relationship between the PI3K/Akt signaling pathway and Epo/VEGF expression in baicalein-mediated neuroprotection in primary rat cortical neurons and astrocytes. Our results show that baicalein induced Epo and VEGF expression in a HIF1α- and PI3K/Akt-dependent manner in neurons. Baicalein also protected neurons against excitotoxicity in a PI3K- and Epo/VEGF-dependent manner without affecting neuronal excitability. In contrast, at least a 10-fold higher concentration of baicalein was needed to induce Epo/VEGF production and PI3K/Akt activity in astrocytes for protection of neurons. Moreover, only baicalein-induced astrocytic VEGF, but not Epo expression requires HIF1α, while PI3K/Akt signaling had little role in baicalein-induced astrocytic Epo/VEGF expression. These results suggest distinct mechanisms of baicalein-mediated Epo/VEGF production in neurons and astrocytes for neuroprotection, and provide new insights into the mechanisms and potential of baicalein in treating brain injury in vivo. PMID:23904909

  15. γ-Enolase C-terminal peptide promotes cell survival and neurite outgrowth by activation of the PI3K/Akt and MAPK/ERK signalling pathways.

    PubMed

    Hafner, Anja; Obermajer, Nataša; Kos, Janko

    2012-04-15

    γ-Enolase, a glycolytic enzyme, is expressed specifically in neurons. It exerts neurotrophic activity and has been suggested to regulate growth, differentiation, survival and regeneration of neurons. In the present study, we investigated the involvement of γ-enolase in PI3K (phosphoinositide 3-kinase)/Akt and MAPK (mitogen-activated protein kinase)/ERK (extracellular-signal-regulated kinase) signalling, the two pathways triggered predominantly by neurotrophic factors. Whereas the PI3K/Akt pathway, rather than the MAPK/ERK pathway, is involved in γ-enolase-enhanced cell survival, γ-enolase-stimulated neurite outgrowth requires both pathways, i.e. the activation of both PI3K and ERK1/2, leading to subsequent expression of the growth-cone-specific protein GAP-43 (growth-associated protein of 43 kDa). MEK (MAPK/ERK kinase) and PI3K inhibition blocked or attenuated the neurite outgrowth associated with dynamic remodelling of the actin-based cytoskeleton. We show that γ-enolase-mediated PI3K activation regulates RhoA kinase, a key regulator of actin cytoskeleton organization. Moreover, the inhibition of RhoA downstream effector ROCK (Rho-associated kinase) results in enhanced γ-enolase-induced neurite outgrowth, accompanied by actin polymerization and its redistribution to growth cones. Our results show that γ-enolase controls neuronal survival, differentiation and neurite regeneration by activating the PI3K/Akt and MAPK/ERK signalling pathways, resulting in downstream regulation of the molecular and cellular processes of cytoskeleton reorganization and cell remodelling, activation of transcriptional factors and regulation of the cell cycle.

  16. Involvement of phosphoinositide 3-kinase class IA (PI3K 110α) and NADPH oxidase 1 (NOX1) in regulation of vascular differentiation induced by vascular endothelial growth factor (VEGF) in mouse embryonic stem cells.

    PubMed

    Bekhite, Mohamed M; Müller, Veronika; Tröger, Sebastian H; Müller, Jörg P; Figulla, Hans-Reiner; Sauer, Heinrich; Wartenberg, Maria

    2016-04-01

    The impact of reactive oxygen species and phosphoinositide 3-kinase (PI3K) in differentiating embryonic stem (ES) cells is largely unknown. Here, we show that the silencing of the PI3K catalytic subunit p110α and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 1 (NOX1) by short hairpin RNA or pharmacological inhibition of NOX and ras-related C3 botulinum toxin substrate 1 (Rac1) abolishes superoxide production by vascular endothelial growth factor (VEGF) in mouse ES cells and in ES-cell-derived fetal liver kinase-1(+) (Flk-1(+)) vascular progenitor cells, whereas the mitochondrial complex I inhibitor rotenone does not have an effect. Silencing p110α or inhibiting Rac1 arrests vasculogenesis at initial stages in embryoid bodies, even under VEGF treatment, as indicated by platelet endothelial cell adhesion molecule-1 (PECAM-1)-positive areas and branching points. In the absence of p110α, tube-like structure formation on matrigel and cell migration of Flk-1(+) cells in scratch migration assays are totally impaired. Silencing NOX1 causes a reduction in PECAM-1-positive areas, branching points, cell migration and tube length upon VEGF treatment, despite the expression of vascular differentiation markers. Interestingly, silencing p110α but not NOX1 inhibits the activation of Rac1, Ras homologue gene family member A (RhoA) and Akt leading to the abrogation of VEGF-induced lamellipodia structure formation. Thus, our data demonstrate that the PI3K p110α-Akt/Rac1 and NOX1 signalling pathways play a pivotal role in VEGF-induced vascular differentiation and cell migration. Rac1, RhoA and Akt phosphorylation occur downstream of PI3K and upstream of NOX1 underscoring a role of PI3K p110α in the regulation of cell polarity and migration. PMID:26553657

  17. The role of the PI3K/Akt/mTOR signalling pathway in human cancers induced by infection with human papillomaviruses.

    PubMed

    Zhang, Lifang; Wu, Jianhong; Ling, Ming Tat; Zhao, Liang; Zhao, Kong-Nan

    2015-01-01

    Infection with Human papillomaviruses (HPVs) leads to the development of a wide-range of cancers, accounting for 5% of all human cancers. A prominent example is cervical cancer, one of the leading causes of cancer death in women worldwide. It has been well established that tumor development and progression induced by HPV infection is driven by the sustained expression of two oncogenes E6 and E7. The expression of E6 and E7 not only inhibits the tumor suppressors p53 and Rb, but also alters additional signalling pathways that may be equally important for transformation. Among these pathways, the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signalling cascade plays a very important role in HPV-induced carcinogenesis by acting through multiple cellular and molecular events. In this review, we summarize the frequent amplification of PI3K/Akt/mTOR signals in HPV-induced cancers and discuss how HPV oncogenes E6/E7/E5 activate the PI3K/Akt/mTOR signalling pathway to modulate tumor initiation and progression and affect patient outcome. Improvement of our understanding of the mechanism by which the PI3K/Akt/mTOR signalling pathway contributes to the immortalization and carcinogenesis of HPV-transduced cells will assist in devising novel strategies for preventing and treating HPV-induced cancers. PMID:26022660

  18. The role of the PI3K/Akt/mTOR signalling pathway in human cancers induced by infection with human papillomaviruses.

    PubMed

    Zhang, Lifang; Wu, Jianhong; Ling, Ming Tat; Zhao, Liang; Zhao, Kong-Nan

    2015-04-17

    Infection with Human papillomaviruses (HPVs) leads to the development of a wide-range of cancers, accounting for 5% of all human cancers. A prominent example is cervical cancer, one of the leading causes of cancer death in women worldwide. It has been well established that tumor development and progression induced by HPV infection is driven by the sustained expression of two oncogenes E6 and E7. The expression of E6 and E7 not only inhibits the tumor suppressors p53 and Rb, but also alters additional signalling pathways that may be equally important for transformation. Among these pathways, the phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signalling cascade plays a very important role in HPV-induced carcinogenesis by acting through multiple cellular and molecular events. In this review, we summarize the frequent amplification of PI3K/Akt/mTOR signals in HPV-induced cancers and discuss how HPV oncogenes E6/E7/E5 activate the PI3K/Akt/mTOR signalling pathway to modulate tumor initiation and progression and affect patient outcome. Improvement of our understanding of the mechanism by which the PI3K/Akt/mTOR signalling pathway contributes to the immortalization and carcinogenesis of HPV-transduced cells will assist in devising novel strategies for preventing and treating HPV-induced cancers.

  19. Biphasic activation of PI3K/Akt and MAPK/Erk1/2 signaling pathways in bovine herpesvirus type 1 infection of MDBK cells.

    PubMed

    Zhu, Liqian; Ding, Xiuyan; Zhu, Xiaofang; Meng, Songshu; Wang, Jianye; Zhou, Hong; Duan, Qiangde; Tao, Jie; Schifferli, Dieter M; Zhu, Guoqiang

    2011-04-14

    Many viruses have been known to control key cellular signaling pathways to facilitate the virus infection. The possible involvement of signaling pathways in bovine herpesvirus type 1 (BoHV-1) infection is unknown. This study indicated that infection of MDBK cells with BoHV-1 induced an early-stage transient and a late-stage sustained activation of both phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen activated protein kinases/extracellular signal-regulated kinase 1/2 (MAPK/Erk1/2) signaling pathways. Analysis with the stimulation of UV-irradiated virus indicated that the virus binding and/or entry process was enough to trigger the early phase activations, while the late phase activations were viral protein expression dependent. Biphasic activation of both pathways was suppressed by the selective inhibitor, Ly294002 for PI3K and U0126 for MAPK kinase (MEK1/2), respectively. Furthermore, treatment of MDBK cells with Ly294002 caused a 1.5-log reduction in virus titer, while U0126 had little effect on the virus production. In addition, the inhibition effect of Ly294002 mainly occurred at the post-entry stage of the virus replication cycle. This revealed for the first time that BoHV-1 actively induced both PI3K/Akt and MAPK/Erk1/2 signaling pathways, and the activation of PI3K was important for fully efficient replication, especially for the post-entry stage.

  20. Ketamine affects the neurogenesis of rat fetal neural stem progenitor cells via the PI3K/Akt-p27 signaling pathway

    PubMed Central

    Dong, Chaoxuan; Rovnaghi, Cynthia R.; Anand, KJS

    2014-01-01

    Ketamine is widely used as an anesthetic, analgesic, or sedative in pediatric patients. We reported that ketamine alters the normal neurogenesis of rat fetal neural stem progenitor cells (NSPCs) in the developing brain, but the underlying mechanisms remain unknown. The PI3K-PKB/Akt (Phosphatidylinositide 3-kinases/protein kinase B) signaling pathway plays many important roles in cell survival, apoptosis, and proliferation. We hypothesized that PI3K-PKB/Akt signaling may be involved in ketamine-altered neurogenesis of cultured NSPCs in vitro. NSPCs were isolated from Sprague-Dawley rat fetuses on gestational day 17. BrdU (bromodeoxyuridine) incorporation, Ki67 staining, and differentiation tests were utilized to identify primary cultured NSPCs. Immunofluorescent staining was used to detect Akt expression, whereas, Western blots measured phosphorylated Akt and p27 expression in NSPCs exposed to different treatments. We report that cultured NSPCs had properties of neurogenesis: proliferation and neural differentiation. PKB/Akt was expressed in cultured rat fetal cortical NSPCs. Ketamine inhibited the phosphorylation of Akt and further enhanced p27 expression in cultured NSPCs. All ketamine-induced PI3K/Akt signaling changes could be recovered by NMDA (N-Methyl-D-aspartate) receptor agonist, NMDA. These data suggest that inhibition of PI3K/Akt-p27 signaling may be involved in ketamine-induced neurotoxicity in the developing brain, whereas excitatory NMDA receptor activation may reverse these effects. PMID:25231110

  1. p87 and p101 subunits are distinct regulators determining class IB phosphoinositide 3-kinase (PI3K) specificity.

    PubMed

    Shymanets, Aliaksei; Prajwal; Bucher, Kirsten; Beer-Hammer, Sandra; Harteneck, Christian; Nürnberg, Bernd

    2013-10-25

    Class IB phosphoinositide 3-kinase γ (PI3Kγ) comprises a single catalytic p110γ subunit, which binds to two non-catalytic subunits, p87 or p101, and controls a plethora of fundamental cellular responses. The non-catalytic subunits are assumed to be redundant adaptors for Gβγ enabling G-protein-coupled receptor-mediated regulation of PI3Kγ. Growing experimental data provide contradictory evidence. To elucidate the roles of the non-catalytic subunits in determining the specificity of PI3Kγ, we tested the impact of p87 and p101 in heterodimeric p87-p110γ and p101-p110γ complexes on the modulation of PI3Kγ activity in vitro and in living cells. RT-PCR, biochemical, and imaging data provide four lines of evidence: (i) specific expression patterns of p87 and p101, (ii) up-regulation of p101, providing the basis to consider p87 as a protein forming a constitutively and p101 as a protein forming an inducibly expressed PI3Kγ, (iii) differences in basal and stimulated enzymatic activities, and (iv) differences in complex stability, all indicating apparent diversity within class IB PI3Kγ. In conclusion, expression and activities of PI3Kγ are modified differently by p87 and p101 in vitro and in living cells, arguing for specific regulatory roles of the non-catalytic subunits in the differentiation of PI3Kγ signaling pathways. PMID:24014027

  2. PI3K signaling of autophagy is required for starvation tolerance and virulenceof Cryptococcus neoformans

    PubMed Central

    Hu, Guowu; Hacham, Moshe; Waterman, Scott R.; Panepinto, John; Shin, Soowan; Liu, Xiaoguang; Gibbons, Jack; Valyi-Nagy, Tibor; Obara, Keisuke; Jaffe, H. Ari; Ohsumi, Yoshinori; Williamson, Peter R.

    2008-01-01

    Autophagy is a process by which cells recycle cytoplasm and defective organelles during stress situations such as nutrient starvation. It can also be used by host cells as an immune defense mechanism to eliminate infectious pathogens. Here we describe the use of autophagy as a survival mechanism and virulence-associated trait by the human fungal pathogen Cryptococcus neoformans. We report that a mutant form of C. neoformans lacking the Vps34 PI3K (vps34Δ), which is known to be involved in autophagy in ascomycete yeast, was defective in the formation of autophagy-related 8–labeled (Atg8-labeled) vesicles and showed a dramatic attenuation in virulence in mouse models of infection. In addition, autophagic vesicles were observed in WT but not vps34Δ cells after phagocytosis by a murine macrophage cell line, and Atg8 expression was exhibited in WT C. neoformans during human infection of brain. To dissect the contribution of defective autophagy in vps34Δ C. neoformans during pathogenesis, a strain of C. neoformans in which Atg8 expression was knocked down by RNA interference was constructed and these fungi also demonstrated markedly attenuated virulence in a mouse model of infection. These results demonstrated PI3K signaling and autophagy as a virulence-associated trait and survival mechanism during infection with a fungal pathogen. Moreover, the data show that molecular dissection of such pathogen stress-response pathways may identify new approaches for chemotherapeutic interventions. PMID:18259613

  3. 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

  4. Flavonoids Extraction from Propolis Attenuates Pathological Cardiac Hypertrophy through PI3K/AKT Signaling Pathway

    PubMed Central

    Sun, Guang-wei; Qiu, Zhi-dong; Wang, Wei-nan; Sui, Xin

    2016-01-01

    Propolis, a traditional medicine, has been widely used for a thousand years as an anti-inflammatory and antioxidant drug. The flavonoid fraction is the main active component of propolis, which possesses a wide range of biological activities, including activities related to heart disease. However, the role of the flavonoids extraction from propolis (FP) in heart disease remains unknown. This study shows that FP could attenuate ISO-induced pathological cardiac hypertrophy (PCH) and heart failure in mice. The effect of the two fetal cardiac genes, atrial natriuretic factor (ANF) and β-myosin heavy chain (β-MHC), on PCH was reversed by FP. Echocardiography analysis revealed cardiac ventricular dilation and contractile dysfunction in ISO-treated mice. This finding is consistent with the increased heart weight and cardiac ANF protein levels, massive replacement fibrosis, and myocardial apoptosis. However, pretreatment of mice with FP could attenuate cardiac dysfunction and hypertrophy in vivo. Furthermore, the cardiac protection of FP was suppressed by the pan-PI3K inhibitor wortmannin. FP is a novel cardioprotective agent that can attenuate adverse cardiac dysfunction, hypertrophy, and associated disorder, such as fibrosis. The effects may be closely correlated with PI3K/AKT signaling. FP may be clinically used to inhibit PCH progression and heart failure. PMID:27213000

  5. Tripeptide SQL Inhibits Platelet Aggregation and Thrombus Formation by Affecting PI3K/Akt Signaling.

    PubMed

    Su, Xing-li; Su, Wen; He, Zhi-long; Ming, Xin; Kong, Yi

    2015-09-01

    Centipede has been prescribed for the treatment of cardiovascular diseases in Asian countries for several hundred years. Previously, a new antiplatelet tripeptide SQL (H-Ser-Gln-Leu-OH) was isolated and characterized from centipede. In this study, we investigated its antithrombotic activities in vivo and underlying mechanism. It was found that SQL inhibited platelet aggregation induced by adenosine diphosphate, thrombin, epinephrine, and collagen and attenuated thrombus formation in both the ferric chloride-induced arterial thrombosis model and arteriovenous shunt thrombosis model in rats. It did not prolong the bleeding time in mice even at the dose of 10 mg/kg that showed potent antithrombosis effects. Molecular docking revealed that SQL binds PI3Kβ with the binding free energy of -24.341 kcal/mol, which is close to that of cocrystallized ligand (-24.220 kcal/mol). Additionally, SQL displayed inhibition on the late (180 seconds) but did not influence the early (60 seconds) Akt Ser473 phosphorylation in the immunoblot assay. These results suggest that SQL inhibits thrombus formation in vivo and that SQL inhibits PI3K-mediated signaling or even the PI3K itself in platelets. This study may help elucidate the mechanism for centipede treating cardiovascular diseases. PMID:25923322

  6. Tripeptide SQL Inhibits Platelet Aggregation and Thrombus Formation by Affecting PI3K/Akt Signaling.

    PubMed

    Su, Xing-li; Su, Wen; He, Zhi-long; Ming, Xin; Kong, Yi

    2015-09-01

    Centipede has been prescribed for the treatment of cardiovascular diseases in Asian countries for several hundred years. Previously, a new antiplatelet tripeptide SQL (H-Ser-Gln-Leu-OH) was isolated and characterized from centipede. In this study, we investigated its antithrombotic activities in vivo and underlying mechanism. It was found that SQL inhibited platelet aggregation induced by adenosine diphosphate, thrombin, epinephrine, and collagen and attenuated thrombus formation in both the ferric chloride-induced arterial thrombosis model and arteriovenous shunt thrombosis model in rats. It did not prolong the bleeding time in mice even at the dose of 10 mg/kg that showed potent antithrombosis effects. Molecular docking revealed that SQL binds PI3Kβ with the binding free energy of -24.341 kcal/mol, which is close to that of cocrystallized ligand (-24.220 kcal/mol). Additionally, SQL displayed inhibition on the late (180 seconds) but did not influence the early (60 seconds) Akt Ser473 phosphorylation in the immunoblot assay. These results suggest that SQL inhibits thrombus formation in vivo and that SQL inhibits PI3K-mediated signaling or even the PI3K itself in platelets. This study may help elucidate the mechanism for centipede treating cardiovascular diseases.

  7. PI3K/Akt1 signalling specifies foregut precursors by generating regionalized extra-cellular matrix

    PubMed Central

    Villegas, S Nahuel; Rothová, Michaela; Barrios-Llerena, Martin E; Pulina, Maria; Hadjantonakis, Anna-Katerina; Le Bihan, Thierry; Astrof, Sophie; Brickman, Joshua M

    2013-01-01

    During embryonic development signalling pathways act repeatedly in different contexts to pattern the emerging germ layers. Understanding how these different responses are regulated is a central question for developmental biology. In this study, we used mouse embryonic stem cell (mESC) differentiation to uncover a new mechanism for PI3K signalling that is required for endoderm specification. We found that PI3K signalling promotes the transition from naïve endoderm precursors into committed anterior endoderm. PI3K promoted commitment via an atypical activity that delimited epithelial-to-mesenchymal transition (EMT). Akt1 transduced this activity via modifications to the extracellular matrix (ECM) and appropriate ECM could itself induce anterior endodermal identity in the absence of PI3K signalling. PI3K/Akt1-modified ECM contained low levels of Fibronectin (Fn1) and we found that Fn1 dose was key to specifying anterior endodermal identity in vivo and in vitro. Thus, localized PI3K activity affects ECM composition and ECM in turn patterns the endoderm. DOI: http://dx.doi.org/10.7554/eLife.00806.001 PMID:24368729

  8. Different inhibition of Gβγ-stimulated class IB phosphoinositide 3-kinase (PI3K) variants by a monoclonal antibody

    PubMed Central

    Shymanets, Aliaksei; Prajwal; Vadas, Oscar; Czupalla, Cornelia; LoPiccolo, Jaclyn; Brenowitz, Michael; Ghigo, Alessandra; Hirsch, Emilio; Krause, Eberhard; Wetzker, Reinhard; Williams, Roger L.; Harteneck, Christian; Nürnberg, Bernd

    2015-01-01

    Class IB phosphoinositide 3-kinases (PI3Kγ) are second-messenger-generating enzymes downstream of signalling cascades triggered by G-protein-coupled-receptors (GPCRs). PI3Kγ variants have one catalytic p110γ subunit that can form two different heterodimers by binding to one of a pair of non-catalytic subunits, p87 or p101. Growing experimental data argue for a different regulation of p87-p110γ and p101-p110γ allowing integration into distinct signalling pathways. Pharmacological tools enabling distinct modulation of the two variants are missing. The ability of an anti-p110γ monoclonal antibody (mAb(A)p110γ) to block PI3Kγ enzymatic activity attracted us to characterize this tool in detail using purified proteins. In order to get insight into the antibody-p110γ-interface, hydrogen-deuterium exchange coupled to mass spectrometry measurements were performed demonstrating binding of the monoclonal antibody to the C2 domain in p110γ, which was accompanied by conformational changes in the helical domain harbouring the Gβγ-binding site. We then studied the modulation of phospholipid vesicles association of PI3Kγ by the antibody. p87-p110γ showed a significantly reduced Gβγ-mediated phospholipid recruitment as compared with p101-p110γ. Concomitantly, in the presence of mAb(A)p110γ Gβγ did not bind to p87-p110γ. These data correlated with the ability of the antibody to block Gβγ-stimulated lipid kinase activity of p87-p110γ 30 times more potently than p101-p110γ. Our data argue for differential regulatory functions of the non-catalytic subunits and a specific Gβγ-dependent regulation of p101 in PI3Kγ activation. In this scenario, we consider the antibody as a valuable tool to dissect the distinct roles of the two PI3Kγ variants downstream of GPCRs. PMID:26173259

  9. Receptor for advanced glycation end products inhibits proliferation in osteoblast through suppression of Wnt, PI3K and ERK signaling

    SciTech Connect

    Li, Guofeng; Xu, Jingren; Li, Zengchun

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer RAGE overexpression suppresses cell proliferation in MC3T3-E1 cells. Black-Right-Pointing-Pointer RAGE overexpression decreases Wnt/{beta}-catenin signaling. Black-Right-Pointing-Pointer RAGE overexpression decreases ERK and PI3K signaling. Black-Right-Pointing-Pointer Inhibition of Wnt signaling abolishes PI3K signaling restored by RAGE blockade. Black-Right-Pointing-Pointer Inhibition of Wnt signaling abolishes ERK signaling restored by RAGE blockade. -- Abstract: Expression of receptor for advanced glycation end products (RAGE) plays a crucial role in bone metabolism. However, the role of RAGE in the control of osteoblast proliferation is not yet evaluated. In the present study, we demonstrate that RAGE overexpression inhibits osteoblast proliferation in vitro. The negative regulation of RAGE on cell proliferation results from suppression of Wnt, PI3K and ERK signaling, and is restored by RAGE neutralizing antibody. Prevention of Wnt signaling using Sfrp1 or DKK1 rescues RAGE-decreased PI3K and ERK signaling and cell proliferation, indicating that the altered cell growth in RAGE overexpressing cells is in part secondary to alterations in Wnt signaling. Consistently, RAGE overexpression inhibits the expression of Wnt targets cyclin D1 and c-myc, which is partially reversed by RAGE blockade. Overall, these results suggest that RAGE inhibits osteoblast proliferation via suppression of Wnt, PI3K and ERK signaling, which provides novel mechanisms by which RAGE regulates osteoblast growth.

  10. Major vault protein supports glioblastoma survival and migration by upregulating the EGFR/PI3K signalling axis

    PubMed Central

    Lötsch, Daniela; Steiner, Elisabeth; Holzmann, Klaus; Spiegl-Kreinecker, Sabine; Pirker, Christine; Hlavaty, Juraj; Petznek, Helga; Hegedus, Balazs; Garay, Tamas; Mohr, Thomas; Sommergruber, Wolfgang; Grusch, Michael; Berger, Walter

    2013-01-01

    Despite their ubiquitous expression and high conservation during evolution, precise cellular functions of vault ribonucleoparticles, mainly built of multiple major vault protein (MVP) copies, are still enigmatic. With regard to cancer, vaults were shown to be upregulated during drug resistance development as well as malignant transformation and progression. Such in a previous study we demonstrated that human astrocytic brain tumours including glioblastoma are generally high in vault levels while MVP expression in normal brain is comparably low. However a direct contribution to the malignant phenotype in general and that of glioblastoma in particular has not been established so far. Thus we address the questions whether MVP itself has a pro-tumorigenic function in glioblastoma. Based on a large tissue collection, we re-confirm strong MVP expression in gliomas as compared to healthy brain. Further, the impact of MVP on human glioblastoma aggressiveness was analysed by using gene transfection, siRNA knock-down and dominant-negative genetic approaches. Our results demonstrate that MVP/vaults significantly support migratory and invasive competence as well as starvation resistance of glioma cells in vitro and in vivo. The enhanced aggressiveness was based on MVP-mediated stabilization of the epidermal growth factor receptor (EGFR)/phosphatidyl-inositol-3-kinase (PI3K) signalling axis. Consequently, MVP overexpression resulted in enhanced growth and brain invasion in human glioblastoma xenograft models. Our study demonstrates, for the first time, that vaults have a tumour-promoting potential by stabilizing EGFR/PI3K-mediated migration and survival pathways in human glioblastoma. PMID:24243798

  11. Silica nanoparticles induce autophagy and endothelial dysfunction via the PI3K/Akt/mTOR signaling pathway.

    PubMed

    Duan, Junchao; Yu, Yongbo; Yu, Yang; Li, Yang; Wang, Ji; Geng, Weijia; Jiang, Lizhen; Li, Qiuling; Zhou, Xianqing; Sun, Zhiwei

    2014-01-01

    Although nanoparticles have a great potential for biomedical applications, there is still a lack of a correlative safety evaluation on the cardiovascular system. This study is aimed to clarify the biological behavior and influence of silica nanoparticles (Nano-SiO2) on endothelial cell function. The results showed that the Nano-SiO2 were internalized into endothelial cells in a dose-dependent manner. Monodansylcadaverine staining, autophagic ultrastructural observation, and LC3-I/LC3-II conversion were employed to verify autophagy activation induced by Nano-SiO2, and the whole autophagic process was also observed in endothelial cells. In addition, the level of nitric oxide (NO), the activities of NO synthase (NOS) and endothelial (e)NOS were significantly decreased in a dose-dependent way, while the activity of inducible (i)NOS was markedly increased. The expression of C-reactive protein, as well as the production of proinflammatory cytokines (tumor necrosis factor α, interleukin [IL]-1β, and IL-6) were significantly elevated. Moreover, Nano-SiO2 had an inhibitory effect on the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway. Our findings demonstrated that Nano-SiO2 could disturb the NO/NOS system, induce inflammatory response, activate autophagy, and eventually lead to endothelial dysfunction via the PI3K/Akt/mTOR pathway. This indicates that exposure to Nano-SiO2 is a potential risk factor for cardiovascular diseases.

  12. Proliferation, survival and metabolism: the role of PI3K/AKT/mTOR signalling in pluripotency and cell fate determination.

    PubMed

    Yu, Jason S L; Cui, Wei

    2016-09-01

    Phosphatidylinositide 3 kinases (PI3Ks) and their downstream mediators AKT and mammalian target of rapamycin (mTOR) constitute the core components of the PI3K/AKT/mTOR signalling cascade, regulating cell proliferation, survival and metabolism. Although these functions are well-defined in the context of tumorigenesis, recent studies - in particular those using pluripotent stem cells - have highlighted the importance of this pathway to development and cellular differentiation. Here, we review the recent in vitro and in vivo evidence for the role PI3K/AKT/mTOR signalling plays in the control of pluripotency and differentiation, with a particular focus on the molecular mechanisms underlying these functions. PMID:27578176

  13. Better Understanding of Phosphoinositide 3-Kinase (PI3K) Pathways in Vasculature: Towards Precision Therapy Targeting Angiogenesis and Tumor Blood Supply.

    PubMed

    Tsvetkov, D; Shymanets, A; Huang, Yu; Bucher, K; Piekorz, R; Hirsch, E; Beer-Hammer, S; Harteneck, C; Gollasch, M; Nürnberg, B

    2016-07-01

    The intracellular PI3K-AKT-mTOR pathway is involved in regulation of numerous important cell processes including cell growth, differentiation, and metabolism. The PI3Kα isoform has received particular attention as a novel molecular target in gene therapy, since this isoform plays critical roles in tumor progression and tumor blood flow and angiogenesis. However, the role of PI3Kα and other class I isoforms, i.e. PI3Kβ, γ, δ, in the regulation of vascular tone and regional blood flow are largely unknown. We used novel isoform-specific PI3K inhibitors and mice deficient in both PI3Kγ and PI3Kδ (Pik3cg(-/-)/Pik3cd(-/-)) to define the putative contribution of PI3K isoform(s) to arterial vasoconstriction. Wire myography was used to measure isometric contractions of isolated murine mesenteric arterial rings. Phenylephrine-dependent contractions were inhibited by the pan PI3K inhibitors wortmannin (100 nM) and LY294002 (10 µM). These vasoconstrictions were also inhibited by the PI3Kα isoform inhibitors A66 (10 µM) and PI-103 (1 µM), but not by the PI3Kβ isoform inhibitor TGX 221 (100 nM). Pik3cg(-/-)/Pik3cd(-/-)-arteries showed normal vasoconstriction. We conclude that PI3Kα is an important downstream element in vasoconstrictor GPCR signaling, which contributes to arterial vasocontraction via α1-adrenergic receptors. Our results highlight a regulatory role of PI3Kα in the cardiovascular system, which widens the spectrum of gene therapy approaches targeting PI3Kα in cancer cells and tumor angiogenesis and regional blood flow. PMID:27449615

  14. 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.

  15. 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

  16. Psoralidin inhibits proliferation and enhances apoptosis of human esophageal carcinoma cells via NF-κB and PI3K/Akt signaling pathways

    PubMed Central

    Jin, Zhiliang; Yan, Wei; Jin, Hui; Ge, Changzheng; Xu, Yanhua

    2016-01-01

    Esophageal cancer is the most common gastrointestinal cancer. Psoralidin exhibits antioxidant, anti-apoptotic, anti-inflammatory and antitumor effects, which result in the inhibition of cancer formation. The present study aimed to investigate the effect of psoralidin on esophageal carcinoma proliferation and growth, and to elucidate its underlying mechanism of action. The effect of psoralidin on cell proliferation was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Using an annexin V-fluorescein isothiocyanate/propidium iodide apoptosis detection kit and 4′,6-diamidino-2-phenylindole staining assay, the present study demonstrated that psoralidin significantly enhanced apoptosis of human esophageal carcinoma Eca9706 cells. In addition, caspase-3 activity was analyzed with a caspase-3 colorimetric assay kit, while nuclear factor (NF)-κB activity and protein phosphatidylinositol 3-kinase (PI3K)/Akt expression were measured with an NF-κB enzyme-linked immunosorbent assay kit and western blot analysis, respectively. Eca9706 cells were treated with a PI3K agonist in order to investigate the mechanism of action of psoralidin. It was observed that psoralidin was able to decrease the proliferation and promote the cellular apoptosis of Eca9706 cells in a dose-dependent manner. Furthermore, psoralidin was also able to inhibit the caspase-3 activity of Eca9706 cells in a dose-dependent manner. In addition, psoralidin inhibited NF-κB activity and reduced PI3K and Akt protein expression in Eca9706 cells. Notably, the PI3K agonist was able to reverse the effect of psoralidin on Eca9706 cells. The results of the present study demonstrated that psoralidin was able to inhibit proliferation and enhance apoptosis of human esophageal carcinoma cells via the NF-κB and PI3K/Akt signaling pathways. PMID:27446379

  17. Low-dose testosterone alleviates vascular damage caused by castration in male rats in puberty via modulation of the PI3K/AKT signaling pathway

    PubMed Central

    Zhao, Jing; Liu, Ge-Li; Wei, Ying; Jiang, Li-Hong; Bao, Peng-Li; Yang, Qing-Yan

    2016-01-01

    The aim of the present study was to investigate the effect of testosterone on glucolipid metabolism and vascular injury in male rats, and examine the underlying molecular mechanisms. A total of 40 male Sprague-Dawley rats were divided into a control group (n=10), high-fat-diet + castration group (n=10), high-fat-diet + castration + low dose testosterone group (n=10), and high-fat-diet + castration + high dose testosterone group (n=10). Hematoxylin and eosin staining was performed to evaluate the morphology of the thoracic aortic tissues. Immunohistochemical staining was used to detect biomarkers of the phosphoinositide 3-kinase (PI3K) signaling pathway. The mRNA and protein expression levels of PI3K, AKT, insulin receptor substrate-1 (IRS-1), glucose transporter type 4 (GLUT-4), nuclear factor (NF)-κB and tumor necrosis factor (TNF)-α in the aortas were determined using quantitative polymerase chain reaction and Western blot analyses, respectively. Apoptosis in the aortic tissues was detected using a TUNEL assay. Castration induced apoptosis in the animals fed a high-fat-diet, whereas low dose testosterone replacement ameliorated the apoptosis in the aorta. However, the levels of apoptosis was more severe following high-dose testosterone treatment. Low-dose testosterone induced upregulation in the levels of IRS-1, AKT, GLUT-4 protein, NF-κB, TNF-α and PI3K, compared with those in the animals fed a high-fat diet following castration. A high dose of testosterone resulted in a significant decrease in the levels of IRS-1, AKT, GLUT-4, NF-κB, TNF-α and PI3K. Compared with the rats in the high-fat diet + castration group, a low dose of testosterone induced upregulation in the mRNA levels of IRS-1, AKT and GLUT-4, and downregulation of the mRNA levels of NF-κB, TNF-α and PI3K. A high dose of testosterone resulted in a significant decrease in the levels of IRS-1, AKT and GLUT-4, and marked increases in the mRNA levels of NF-κB, TNF-α and PI3K, compared with the

  18. Gardenamide A Protects RGC-5 Cells from H₂O₂-Induced Oxidative Stress Insults by Activating PI3K/Akt/eNOS Signaling Pathway.

    PubMed

    Wang, Rikang; Peng, Lizhi; Zhao, Jiaqiang; Zhang, Laitao; Guo, Cuiping; Zheng, Wenhua; Chen, Heru

    2015-01-01

    Gardenamide A (GA) protects the rat retinal ganglion (RGC-5) cells against cell apoptosis induced by H₂O₂. The protective effect of GA was completely abrogated by the specific phosphoinositide 3-kinase (PI3K) inhibitor LY294002, and the specific protein kinase B (Akt) inhibitor Akt VIII respectively, indicating that the protective mechanism of GA is mediated by the PI3K/Akt signaling pathway. The specific extracellular signal-regulated kinase (ERK1/2) inhibitor PD98059 could not block the neuroprotection of GA. GA attenuated the levels of reactive oxygen species (ROS) and malondialdehyde (MDA) induced by H₂O₂. Western blotting showed that GA promoted the phosphorylation of ERK1/2, Akt and endothelial nitric oxide synthase (eNOS), respectively, and effectively reversed the H₂O₂-inhibited phosphorylation of these three proteins. LY294002 completely inhibited the GA-activated phosphorylation of Akt, while only partially inhibiting eNOS. This evidence implies that eNOS may be activated directly by GA. PD98059 attenuated only partially the GA-induced phosphorylation of ERK1/2 with/without the presence of H₂O₂, indicating that GA may activate ERK1/2 directly. All these results put together confirm that GA protects RGC-5 cells from H₂O₂ insults via the activation of PI3K/Akt/eNOS signaling pathway. Whether the ERK1/2 signaling pathway is involved requires further investigations.

  19. PTEN/PI3K/Akt/VEGF signaling and the cross talk to KRIT1, CCM2, and PDCD10 proteins in cerebral cavernous malformations.

    PubMed

    Kar, Souvik; Samii, Amir; Bertalanffy, Helmut

    2015-04-01

    Cerebral cavernous malformations (CCM) are common vascular malformation of the brain and are associated with abnormal angiogenesis. Although the exact etiology and the underlying molecular mechanism are still under investigation, recent advances in the identification of the mutations in three genes and their interactions with different signaling pathways have shed light on our understanding of CCM pathogenesis. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is known to play a major role in angiogenesis. Studies have shown that the phosphatase and tensin homologue deleted on chromosome ten (PTEN), a tumor suppressor, is an antagonist regulator of the PI3K/Akt pathway and mediates angiogenesis by activating vascular endothelial growth factor (VEGF) expression. Here, we provide an update literature review on the current knowledge of the PTEN/PI3K/Akt/VEGF signaling in angiogenesis, more importantly in CCM pathogenesis. In addition to reviewing the current literatures, this article will also focus on the structural domain of the three CCM proteins and their interacting partners. Understanding the biology of these proteins with respect to their signaling counterpart will help to guide future research towards new therapeutic targets applicable for CCM treatment.

  20. Myrcia bella Leaf Extract Presents Hypoglycemic Activity via PI3k/Akt Insulin Signaling Pathway

    PubMed Central

    Vareda, Priscilla Maria Ponce; Saldanha, Luiz Leonardo; Camaforte, Nathalia Aparecida de Paula; Violato, Natalia Moretti; Dokkedal, Anne Lígia; Bosqueiro, José Roberto

    2014-01-01

    Species of Myrcia are used by indigenous people and in traditional communities in Brazil for the treatment of Diabetes mellitus. We investigated the hypoglycemic effect of the extract of leaves of Myrcia bella in diabetic mice. The chemical fingerprinting of the 70% EtOH extract characterized as main constituents flavonoid aglycones, flavonoid-O-glycosides, and acylated flavonoid-O-glycosides derivatives of quercetin and myricetin. Mice were treated with saline or extract of M. bella (300 or 600 mg/Kg b.w.) for 14 days. Body weight and water and food intake were measured every day. Fasting blood glucose was measured weekly. At the end of the treatment, blood insulin, triglycerides, total cholesterol, and protein were measured. Glycogen content and expression of proteins of the insulin signaling pathway were measured in liver. The treatment with 600 mg/Kg reduced the fasting blood glucose in diabetic mice of the 7th day as water and food intake and increased hepatic glycogen. Total cholesterol and triglycerides were reduced in diabetic treated mice. The treatment increased the expression of IRS-1, PI3-K, and AKT in the livers of diabetic treated mice. The results indicate that the extract of the leaves of Myrcia bella has hypoglycemic properties and possibly acts to regulate glucose uptake by the liver. PMID:24872834

  1. Hedgehog signaling is a novel therapeutic target in tamoxifen-resistant breast cancer aberrantly activated by PI3K/AKT pathway.

    PubMed

    Ramaswamy, Bhuvaneswari; Lu, Yuanzhi; Teng, Kun-yu; Nuovo, Gerard; Li, Xiaobai; Shapiro, Charles L; Majumder, Sarmila

    2012-10-01

    Endocrine resistance is a major challenge in the management of estrogen receptor (ER)-positive breast cancers. Although multiple mechanisms leading to endocrine resistance have been proposed, the poor outcome of patients developing resistance to endocrine therapy warrants additional studies. Here we show that noncanonical Hedgehog (Hh) signaling is an alternative growth promoting mechanism that is activated in tamoxifen-resistant tumors. Importantly, phosphoinositide 3-kinase inhibitor/protein kinase B (PI3K/AKT) pathway plays a key role in regulating Hh signaling by protecting key components of this pathway from proteasomal degradation. The levels of Hh-signaling molecules SMO and GLI1 and the targets were significantly elevated in tamoxifen-resistant MCF-7 cells and T47D cells. Serial passage of the resistant cells in mice resulted in aggressive tumors that metastasized to distant organs with concurrent increases in Hh marker expression and epithelial mesenchymal transition. RNAi-mediated depletion of SMO or GLI1 in the resistant cells resulted in reduced proliferation, clonogenic survival and delayed G(1)-S transition. Notably, treatment of resistant cells with PI3K inhibitors decreased SMO and GLI1 protein levels and activity that was rescued upon blocking GSK3β and proteasomal degradation. Furthermore, treatment of tamoxifen-resistant xenografts with anti-Hh compound GDC-0449 blocked tumor growth in mice. Importantly, high GLI1 expression correlated inversely with disease-free and overall survival in a cohort of 315 patients with breast cancer. In summary, our results describe a signaling event linking PI3K/AKT pathway with Hh signaling that promotes tamoxifen resistance. Targeting Hh pathway alone or in combination with PI3K/AKT pathway could therefore be a novel therapeutic option in treating endocrine-resistant breast cancer.

  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. Girdin regulates the migration and invasion of glioma cells via the PI3K-Akt signaling pathway

    PubMed Central

    NI, WEIMIN; FANG, YAN; TONG, LEI; TONG, ZHAOXUE; YI, FUXIN; QIU, JIANWU; WANG, RUI; TONG, XIAOJIE

    2015-01-01

    Girdin, an actin-binding protein, is associated with cell migration and is expressed at high levels in glioma cells. However, the association between girdin and the development of glioma remains to be elucidated. In the present study, short-hairpin RNA technology was used to silence the gene expression of girdin. The effects of girdin silencing on glioma cell proliferation, migration and invasion were then assessed using a cell viability assay, wound-healing assay, transwell invasion assay, reverse transcription-quantitative polymerase chain reaction, western blot analysis and gelatin zymography. The results suggested that girdin silencing inhibited the proliferation, migration and invasion of glioma cells. In addition, the expression levels and activity of matrix metalloproteinase (MMP)-2 and MMP-9 were also affected by girdin silencing. Further mechanistic investigation indicated that girdin may regulate glioma cell migration and invasion through the phosphatidylinositol-3-kinase/protein kinase B (PI3K-Akt) signaling pathway. Therefore, the results of the present study provide a theoretical foundation for the development of anticancer drugs. PMID:26151295

  4. The PI3K pathway balances self-renewal and differentiation of nephron progenitor cells through β-catenin signaling.

    PubMed

    Lindström, Nils Olof; Carragher, Neil Oliver; Hohenstein, Peter

    2015-04-14

    Nephron progenitor cells differentiate to form nephrons during embryonic kidney development. In contrast, self-renewal maintains progenitor numbers and premature depletion leads to impaired kidney function. Here we analyze the PI3K pathway as a point of convergence for the multiple pathways that are known to control self-renewal in the kidney. We demonstrate that a reduction in PI3K signaling triggers premature differentiation of the progenitors and activates a differentiation program that precedes the mesenchymal-to-epithelial transition through ectopic activation of the β-catenin pathway. Therefore, the combined output of PI3K and other pathways fine-tunes the balance between self-renewal and differentiation in nephron progenitors.

  5. Jolkinolide B induces apoptosis in MDA-MB-231 cells through inhibition of the PI3K/Akt signaling pathway.

    PubMed

    Lin, Yu; Cui, Hongxia; Xu, Huiyu; Yue, Liling; Xu, Hao; Jiang, Liyan; Liu, Jicheng

    2012-06-01

    The phosphoinositol-3-kinase (PI3K)/Akt signal transduction pathway is critically important for tumor cell growth, proliferation and apoptosis. Apoptosis activation has been reported to be a good target in cancer therapies. In this study, we have found that jolkinolide B (JB), a diterpenoid from the traditional Chinese medicinal herb Euphorbia fischeriana Steud, strongly inhibited the expression of the PI3K p85 subunit and the phosphorylation of Akt. Furthermore, we evaluated the effects of JB on the proliferation and apoptosis of MDA-MB-231 human breast cancer cells. Our results show significant induction of apoptosis in MDA-MB-231 cells incubated with JB. This effect was enhanced by combination with LY294002. In addition, treatment with JB could induce downregulation of the Bcl-2/Bax ratio, and subsequent promotion of mitochondrial release of cytochrome c and activation of caspase-3. Taken together, JB-induced apoptosis of MDA-MB-231 cells occurs through the mitochondrial pathway. Further, the PI3K/Akt signaling cascade plays a role in the induction of apoptosis in JB-treated cells. These observations suggest that JB may have therapeutic applications in the treatment of cancer.

  6. beta-Sitosterol induces G2/M arrest, endoreduplication, and apoptosis through the Bcl-2 and PI3K/Akt signaling pathways.

    PubMed

    Moon, Dong-Oh; Kim, Mun-Ock; Choi, Yung Hyun; Kim, Gi-Young

    2008-06-18

    beta-Sitosterol (SITO) is a potentially valuable candidate for cancer chemotherapy, however the cellular and molecular mechanisms responsible for its anti-cancer activity are unknown. Therefore, we attempted to elucidate the mechanisms responsible for SITO-induced anti-proliferation in human leukemia cells. Treatment with SITO increased caspase-3 activation and DNA fragmentation in U937 and HL60 cells. This effect was associated with significant G2/M arrest and endoreduplication. We also demonstrated that SITO treatment significantly increases levels of polymeric alpha-tubulin and promoted microtubule polymerization. We next elucidated that ectopic expression of Bcl-2 accelerates endoreduplication in U937 cells. Furthermore, the specific Bcl-2 inhibitor, HA14-1, prevented endoreduplication through G2 phase arrest. Interestingly, SITO treatment did not significantly promote endoreduplication or decrease cell viability in Bcl-2 null K562 cells. SITO treatment also induced a gradual increase of phosphatidyl-inositol 3-kinase (PI3K) and Akt phosphorylation. Treatment with the selective PI3K/Akt inhibitor LY29004 completely blocked endoreduplication and apoptosis in the presence of SITO. In addition, treatment with SITO-induced phosphorylation of extracellular signal-regulated protein kinase (ERK), however significance of ERK activation in the execution of apoptosis and endoreduplication is unknown. These results suggest that SITO induces endoreduplication by promoting spindle microtubule dynamics through the Bcl-2 and PI3K/Akt signaling pathways.

  7. Sanguinarine Induces Apoptosis of Human Oral Squamous Cell Carcinoma KB Cells via Inactivation of the PI3K/Akt Signaling Pathway.

    PubMed

    Lee, Tae Kyung; Park, Cheol; Jeong, Soon-Jeong; Jeong, Moon-Jin; Kim, Gi-Young; Kim, Wun-Jae; Choi, Yung Hyun

    2016-08-01

    Preclinical Research Sanguinarine, an alkaloid isolated from the root of Sanguinaria canadensis and other plants of the Papaveraceae family, selectively induces apoptotic cell death in a variety of human cancer cells, but its mechanism of action requires further elaboration. The present study investigated the pro-apoptotic effects of sanguinarine in human oral squamous cell carcinoma KB cells. Sanguinarine treatment increased DR5/TRAILR2 (death receptor 5/TRAIL receptor 2) expression and enhanced the activation of caspase-8 and cleavage of its substrate, Bid. Sanguinarine also induced the mitochondrial translocation of pro-apoptotic Bax, mitochondrial dysfunction, cytochrome c release to the cytosol, and activation of caspase-9 and -3. However, a pan-caspase inhibitor, z-VAD-fmk, reversed the growth inhibition and apoptosis induced by sanguinarine. Sanguinarine also suppressed the phosphorylation of phosphoinositide 3-kinase (PI3K) and Akt in KB cells, while co-treatment of cells with sanguinarine and a PI3K inhibitor revealed synergistic apoptotic effects. However, pharmacological inhibition of AMP-activated protein kinase and mitogen-activated protein kinases did not reduce or enhance sanguinarine-induced growth inhibition and apoptosis. Collectively, these findings indicate that the pro-apoptotic effects of sanguinarine in KB cells may be regulated by a caspase-dependent cascade via activation of both intrinsic and extrinsic signaling pathways and inactivation of PI3K/Akt signaling. Drug Dev Res 77 : 227-240, 2016.   © 2016 Wiley Periodicals, Inc.

  8. Anger Emotional Stress Influences VEGF/VEGFR2 and Its Induced PI3K/AKT/mTOR Signaling Pathway

    PubMed Central

    Sun, Peng; Wei, Sheng; Wei, Xia; Wang, Jieqiong; Zhang, Yuanyuan; Qiao, Mingqi; Wu, Jibiao

    2016-01-01

    Objective. We discuss the influence of anger emotional stress upon VEGF/VEGFR2 and its induced PI3K/AKT/mTOR signal pathway. Methods. We created a rat model of induced anger (anger-out and anger-in) emotional response using social isolation and resident-intruder paradigms and assessed changes in hippocampus' VEGF content, neuroplasticity, and the PI3K/AKT/mTOR signaling pathway. Results. The resident-intruder method successfully generated anger-out and anger-in models that differed significantly in composite aggression score, aggression incubation, open field behavior, sucrose preference, and weight gain. Anger emotional stress decreased synaptic connections and VEGFR2 expression. Anger emotional stress led to abnormal expression of VEGF/VEGFR2 mRNA and protein and disorderly expression of key factors in the PI3K/AKT/mTOR signal pathway. Fluoxetine administration ameliorated behavioral abnormalities and damage to hippocampal neurons caused by anger emotional stress, as well as abnormal expression of some proteins in VEGF/VEGFR2 and its induced PI3K/AKT/mTOR signal pathway. Conclusion. This research provides a detailed classification of anger emotion and verifies its influence upon VEGF and the VEGF-induced signaling pathway, thus providing circumstantial evidence of mechanisms by which anger emotion damages neurogenesis. As VEGFR2 can promote neurogenesis and vasculogenesis in the hippocampus and frontal lobe, these results suggest that anger emotional stress can result in decreased neurogenesis. PMID:27057362

  9. Anger Emotional Stress Influences VEGF/VEGFR2 and Its Induced PI3K/AKT/mTOR Signaling Pathway.

    PubMed

    Sun, Peng; Wei, Sheng; Wei, Xia; Wang, Jieqiong; Zhang, Yuanyuan; Qiao, Mingqi; Wu, Jibiao

    2016-01-01

    Objective. We discuss the influence of anger emotional stress upon VEGF/VEGFR2 and its induced PI3K/AKT/mTOR signal pathway. Methods. We created a rat model of induced anger (anger-out and anger-in) emotional response using social isolation and resident-intruder paradigms and assessed changes in hippocampus' VEGF content, neuroplasticity, and the PI3K/AKT/mTOR signaling pathway. Results. The resident-intruder method successfully generated anger-out and anger-in models that differed significantly in composite aggression score, aggression incubation, open field behavior, sucrose preference, and weight gain. Anger emotional stress decreased synaptic connections and VEGFR2 expression. Anger emotional stress led to abnormal expression of VEGF/VEGFR2 mRNA and protein and disorderly expression of key factors in the PI3K/AKT/mTOR signal pathway. Fluoxetine administration ameliorated behavioral abnormalities and damage to hippocampal neurons caused by anger emotional stress, as well as abnormal expression of some proteins in VEGF/VEGFR2 and its induced PI3K/AKT/mTOR signal pathway. Conclusion. This research provides a detailed classification of anger emotion and verifies its influence upon VEGF and the VEGF-induced signaling pathway, thus providing circumstantial evidence of mechanisms by which anger emotion damages neurogenesis. As VEGFR2 can promote neurogenesis and vasculogenesis in the hippocampus and frontal lobe, these results suggest that anger emotional stress can result in decreased neurogenesis. PMID:27057362

  10. 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

  11. Sedanolide induces autophagy through the PI3K, p53 and NF-κB signaling pathways in human liver cancer cells.

    PubMed

    Hsieh, Shu-Ling; Chen, Chi-Tsai; Wang, Jyh-Jye; Kuo, Yu-Hao; Li, Chien-Chun; Hsieh, Lan-Chi; Wu, Chih-Chung

    2015-12-01

    Sedanolide (SN), a phthalide-like compound from celery seed oil, possesses antioxidant effects. However, the effect of SN on cell death in human liver cancer cells has yet to be determined. In this study, cell viability determination, monodansylcadaverine (MDC) fluorescent staining and immunoblot analysis were performed to determine autophagy induction and autophagy-induced protein expression changes via molecular examination after human liver cancer (J5) cells were treated with SN. Our studies demonstrate that SN suppressed J5 cell viability by inducing autophagy. Phosphoinositide 3-kinase (PI3K)-I, mammalian target of rapamycin (mTOR) and Akt protein levels decreased, whereas PI3K-III, LC3-II and Beclin-1 protein levels increased following SN treatment in J5 cells. In addition, SN treatment upregulated nuclear p53 and damage-regulated autophagy modulator (DRAM) and downregulated cytosolic p53 and Tp53-induced glycolysis and apoptosis regulator (TIGAR) expression in J5 cells. Furthermore, the cytosolic phosphorylation of inhibitor of kappa B (IκB) and nuclear p65 and the DNA-binding activity of NF-κB increased after SN treatment. These results suggest that SN induces J5 cell autophagy by regulating PI3K, p53 and NF-κB autophagy-associated signaling pathways in J5 cells. PMID:26500073

  12. Discovery of a Selective Phosphoinositide-3-Kinase (PI3K)-γ Inhibitor (IPI-549) as an Immuno-Oncology Clinical Candidate.

    PubMed

    Evans, Catherine A; Liu, Tao; Lescarbeau, André; Nair, Somarajan J; Grenier, Louis; Pradeilles, Johan A; Glenadel, Quentin; Tibbitts, Thomas; Rowley, Ann M; DiNitto, Jonathan P; Brophy, Erin E; O'Hearn, Erin L; Ali, Janid A; Winkler, David G; Goldstein, Stanley I; O'Hearn, Patrick; Martin, Christian M; Hoyt, Jennifer G; Soglia, John R; Cheung, Culver; Pink, Melissa M; Proctor, Jennifer L; Palombella, Vito J; Tremblay, Martin R; Castro, Alfredo C

    2016-09-01

    Optimization of isoquinolinone PI3K inhibitors led to the discovery of a potent inhibitor of PI3K-γ (26 or IPI-549) with >100-fold selectivity over other lipid and protein kinases. IPI-549 demonstrates favorable pharmacokinetic properties and robust inhibition of PI3K-γ mediated neutrophil migration in vivo and is currently in Phase 1 clinical evaluation in subjects with advanced solid tumors. PMID:27660692

  13. The PI3K signaling-mediated nitric oxide contributes to cardiovascular effects of angiotensin-(1-7) in the nucleus tractus solitarii of rats.

    PubMed

    Wu, Zhao-Tang; Ren, Chang-Zhen; Yang, Ya-Hong; Zhang, Ru-Wen; Sun, Jia-Cen; Wang, Yang-Kai; Su, Ding-Feng; Wang, Wei-Zhong

    2016-01-30

    Angiotensin-1-7 [Ang-(1-7)], acting via the Mas receptor in the central nervous system, is involved in the regulation of cardiovascular activity. Nitric oxide (NO) is implicated as an important modulator in the nucleus tractus solitarii (NTS), a key region involved in control of cardiovascular activity. The aim of the present study was to determine the role of phosphatidylinositol 3-kinase (PI3K) signaling in mediating the effect of Ang-(1-7) on NO generation in the NTS. In Sprague-Dawley rats, acute injection of Ang-(1-7) into the NTS significantly increased NO generation and neuronal/endothelial NO synthase (n/eNOS) activity, which were abolished by the selective Mas receptor antagonist d-Alanine-[Ang-(1-7)] (A-779), the PI3K inhibitor LY294002, or the Akt inhibitor triciribine (TCN). Western blotting analysis further demonstrated that Ang-(1-7) significantly increased levels of Akt/NOS phosphorylation in the NTS, and Ang-(1-7)-induced e/nNOS phosphorylation was antagonized by LY294002 or TCN. Furthermore, gene knockdown of PI3K by lentivirus containing small hairpin RNA in the NTS prevented the Ang-(1-7)-induced increases in NOS/Akt phosphorylation and NO production. The physiological (in vivo) experiments showed that pretreatment with the NOS inhibitor l-NAME, LY294002, or TCN abolished the decreases in blood pressure, heart rate, and renal sympathetic nerve activity induced by Ang-(1-7) injected into the NTS. Our findings suggest that nitric oxide release meditated by the Mas-PI3K-NOS signaling pathway is involved in the cardiovascular effects of Ang-(1-7) in the NTS.

  14. Bone marrow-derived mesenchymal stem cells enhance autophagy via PI3K/AKT signalling to reduce the severity of ischaemia/reperfusion-induced lung injury

    PubMed Central

    Li, Jing; Zhou, Jian; Zhang, Dan; Song, Yuanlin; She, Jun; Bai, Chunxue

    2015-01-01

    Autophagy, a type II programmed cell death, is essential for cell survival under stress, e.g. lung injury, and bone marrow-derived mesenchymal stem cells (BM-MSCs) have great potential for cell therapy. However, the mechanisms underlying the BM-MSC activation of autophagy to provide a therapeutic effect in ischaemia/reperfusion-induced lung injury (IRI) remain unclear. Thus, we investigate the activation of autophagy in IRI following transplantation with BM-MSCs. Seventy mice were pre-treated with BM-MSCs before they underwent lung IRI surgery in vivo. Human pulmonary micro-vascular endothelial cells (HPMVECs) were pre-conditioned with BM-MSCs by oxygen-glucose deprivation/reoxygenation (OGD) in vitro. Expression markers for autophagy and the phosphoinositide 3-kinase/protein kinase B (PI3K/Akt) signalling pathway were analysed. In IRI-treated mice, administration of BM-MSCs significantly attenuated lung injury and inflammation, and increased the level of autophagy. In OGD-treated HPMVECs, co-culture with BM-MSCs attenuated endothelial permeability by decreasing the level of cell death and enhanced autophagic activation. Moreover, administration of BM-MSCs decreased the level of PI3K class I and p-Akt while the expression of PI3K class III was increased. Finally, BM-MSCs-induced autophagic activity was prevented using the inhibitor LY294002. Administration of BM-MSCs attenuated lung injury by improving the autophagy level via the PI3K/Akt signalling pathway. These findings provide further understanding of the mechanisms related to BM-MSCs and will help to develop new cell-based therapeutic strategies in lung injury. PMID:26177266

  15. PI3K-Akt-mTOR signal inhibition affects expression of genes related to endoplasmic reticulum stress.

    PubMed

    Song, Q; Han, C C; Xiong, X P; He, F; Gan, W; Wei, S H; Liu, H H; Li, L; Xu, H Y

    2016-01-01

    PI3K-Akt-mTOR signaling pathway is associated with endoplasmic reticulum (ER) stress. However, it is not clear how this signaling pathway affects the ER stress. The present study aimed to determine whether the PI3K-Akt-mTOR signaling pathway regulates tunicamycin (TM)-induced increases in mRNA levels of genes involved in the ER stress, to help elucidate the mechanism by which this pathway affects the ER stress in primary goose hepatocytes. Primary hepatocytes were isolated from geese and cultured in vitro. After 12 h in a serum-free medium, the hepatocytes were incubated for 24 h in a medium with either no addition (control) or with supplementation of TM or TM together with PI3K-Akt-mTOR signaling pathway inhibitors (LY294002, rapamycin, NVP-BEZ235). Thereafter, the expression levels of genes involved in the ER stress (BIP, EIF2a, ATF6, and XBP1) were assessed. The results indicated that the mRNA level of BIP was up-regulated in 0.2, 2, and 20 μM TM treatment group (P < 0.05), whereas the mRNA levels of EIF2a, ATF6, and XBP1 were up-regulated in the 2 μM TM treatment group (P < 0.05). However, the TM mediated induction of mRNA levels of genes involved in the ER stress (BIP, EIF2a, ATF6, and XBP1) was down-regulated after the treatment with PI3K-Akt-mTOR pathway inhibitors (LY294002, NVP-BEZ235, and rapamycin). Therefore, our results strongly suggest that the PI3K-Akt-mTOR signaling pathway might be involved in the down-regulation of the TM-induced ER stress in primary goose hepatocytes. PMID:27525855

  16. Predicting the structures of complexes between phosphoinositide 3-kinase (PI3K) and romidepsin-related compounds for the drug design of PI3K/histone deacetylase dual inhibitors using computational docking and the ligand-based drug design approach.

    PubMed

    Oda, Akifumi; Saijo, Ken; Ishioka, Chikashi; Narita, Koichi; Katoh, Tadashi; Watanabe, Yurie; Fukuyoshi, Shuichi; Takahashi, Ohgi

    2014-11-01

    Predictions of the three-dimensional (3D) structures of the complexes between phosphoinositide 3-kinase (PI3K) and two inhibitors were conducted using computational docking and the ligand-based drug design approach. The obtained structures were refined by structural optimizations and molecular dynamics (MD) simulations. The ligands were located deep inside the ligand binding pocket of the p110α subunit of PI3K, and the hydrogen bond formations and hydrophobic effects of the surrounding amino acids were predicted. Although rough structures were obtained for the PI3K-inhibitor complexes before the MD simulations, the refinement of the structures by these simulations clarified the hydrogen bonding patterns of the complexes.

  17. Loss of NDRG2 expression activates PI3K-AKT signalling via PTEN phosphorylation in ATLL and other cancers

    PubMed Central

    Nakahata, Shingo; Ichikawa, Tomonaga; Maneesaay, Phudit; Saito, Yusuke; Nagai, Kentaro; Tamura, Tomohiro; Manachai, Nawin; Yamakawa, Norio; Hamasaki, Makoto; Kitabayashi, Issay; Arai, Yasuhito; Kanai, Yae; Taki, Tomohiko; Abe, Takaya; Kiyonari, Hiroshi; Shimoda, Kazuya; Ohshima, Koichi; Horii, Akira; Shima, Hiroshi; Taniwaki, Masafumi; Yamaguchi, Ryoji; Morishita, Kazuhiro

    2014-01-01

    Constitutive phosphatidylinositol 3-kinase (PI3K)-AKT activation has a causal role in adult T-cell leukaemia-lymphoma (ATLL) and other cancers. ATLL cells do not harbour genetic alterations in PTEN and PI3KCA but express high levels of PTEN that is highly phosphorylated at its C-terminal tail. Here we report a mechanism for the N-myc downstream-regulated gene 2 (NDRG2)-dependent regulation of PTEN phosphatase activity via the dephosphorylation of PTEN at the Ser380, Thr382 and Thr383 cluster within the C-terminal tail. We show that NDRG2 is a PTEN-binding protein that recruits protein phosphatase 2A (PP2A) to PTEN. The expression of NDRG2 is frequently downregulated in ATLL, resulting in enhanced phosphorylation of PTEN at the Ser380/Thr382/Thr383 cluster and enhanced activation of the PI3K-AKT pathway. Given the high incidence of T-cell lymphoma and other cancers in NDRG2-deficient mice, PI3K-AKT activation via enhanced PTEN phosphorylation may be critical for the development of cancer. PMID:24569712

  18. Cyclic mechanical strain maintains Nanog expression through PI3K/Akt signaling in mouse embryonic stem cells

    SciTech Connect

    Horiuchi, Rie; Akimoto, Takayuki; Hong, Zhang; Ushida, Takashi

    2012-08-15

    Mechanical strain has been reported to affect the proliferation/differentiation of many cell types; however, the effects of mechanotransduction on self-renewal as well as pluripotency of embryonic stem (ES) cells remains unknown. To investigate the effects of mechanical strain on mouse ES cell fate, we examined the expression of Nanog, which is an essential regulator of self-renewal and pluripotency as well as Nanog-associated intracellular signaling during uniaxial cyclic mechanical strain. The mouse ES cell line, CCE was plated onto elastic membranes, and we applied 10% strain at 0.17 Hz. The expression of Nanog was reduced during ES cell differentiation in response to the withdrawal of leukemia inhibitory factor (LIF); however, two days of cyclic mechanical strain attenuated this reduction of Nanog expression. On the other hand, the cyclic mechanical strain promoted PI3K-Akt signaling, which is reported as an upstream of Nanog transcription. The cyclic mechanical strain-induced Akt phosphorylation was blunted by the PI3K inhibitor wortmannin. Furthermore, cytochalasin D, an inhibitor of actin polymerization, also inhibited the mechanical strain-induced increase in phospho-Akt. These findings imply that mechanical force plays a role in regulating Nanog expression in ES cells through the actin cytoskeleton-PI3K-Akt signaling. -- Highlights: Black-Right-Pointing-Pointer The expression of Nanog, which is an essential regulator of 'stemness' was reduced during embryonic stem (ES) cell differentiation. Black-Right-Pointing-Pointer Cyclic mechanical strain attenuated the reduction of Nanog expression. Black-Right-Pointing-Pointer Cyclic mechanical strain promoted PI3K-Akt signaling and mechanical strain-induced Akt phosphorylation was blunted by the PI3K inhibitor and an inhibitor of actin polymerization.

  19. Cinnamaldehyde affects the biological behavior of human colorectal cancer cells and induces apoptosis via inhibition of the PI3K/Akt signaling pathway.

    PubMed

    Li, Jiepin; Teng, Yuhao; Liu, Shenlin; Wang, Zifan; Chen, Yan; Zhang, Yingying; Xi, Songyang; Xu, Song; Wang, Ruiping; Zou, Xi

    2016-03-01

    Cinnamaldehyde (CA) is a bioactive compound isolated from the stem bark of Cinnamomum cassia, that has been identified as an antiproliferative substance with pro-apoptotic effects on various cancer cell lines in vitro. In the present study, the effects of CA on human colon cancer cells were investigated at both the molecular and cellular levels. Three types of colorectal cancer cells at various stages of differentiation and invasive ability (SW480, HCT116 and LoVo) were treated with CA at final concentrations of 20, 40 and 80 µg/ml for 24 h. Compared with the control group, the proliferation inhibition rate of the human colorectal cancer cells following treatment with CA increased in a dose- and time-dependent manner. The invasion and adhesion abilities of the cells were significantly inhibited as indicated by Transwell and cell-matrix adhesion assays. Meanwhile, CA also upregulated the expression of E-cadherin and downregulated the expression of matrix metalloproteinase-2 (MMP-2) and MMP-9. CA also elevated the apoptotic rate. The levels of pro-apoptotic genes were upregulated while the levels of apoptosis inhibitory genes were decreased which further confirmed the pro-apoptotic effect of CA. In order to explore the mechanism of CA-induced apoptosis, insulin-like growth factor-1 (IGF-1) and PI3K inhibitor (LY294002) were used to regulate the phosphoinositide 3-kinase (PI3K)/AKT pathway. The transcription activity of PI3K/AKT was markedly inhibited by CA, as well as IGF-1 which functions as an anti-apoptotic factor. In conclusion, CA has the potential to be developed as a new antitumor drug. The mechanisms of action involve the regulation of expression of genes involved in apoptosis, invasion and adhesion via inhibition of the PI3K/Akt signaling pathway. PMID:26677144

  20. 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.

  1. Cinnamaldehyde affects the biological behavior of human colorectal cancer cells and induces apoptosis via inhibition of the PI3K/Akt signaling pathway.

    PubMed

    Li, Jiepin; Teng, Yuhao; Liu, Shenlin; Wang, Zifan; Chen, Yan; Zhang, Yingying; Xi, Songyang; Xu, Song; Wang, Ruiping; Zou, Xi

    2016-03-01

    Cinnamaldehyde (CA) is a bioactive compound isolated from the stem bark of Cinnamomum cassia, that has been identified as an antiproliferative substance with pro-apoptotic effects on various cancer cell lines in vitro. In the present study, the effects of CA on human colon cancer cells were investigated at both the molecular and cellular levels. Three types of colorectal cancer cells at various stages of differentiation and invasive ability (SW480, HCT116 and LoVo) were treated with CA at final concentrations of 20, 40 and 80 µg/ml for 24 h. Compared with the control group, the proliferation inhibition rate of the human colorectal cancer cells following treatment with CA increased in a dose- and time-dependent manner. The invasion and adhesion abilities of the cells were significantly inhibited as indicated by Transwell and cell-matrix adhesion assays. Meanwhile, CA also upregulated the expression of E-cadherin and downregulated the expression of matrix metalloproteinase-2 (MMP-2) and MMP-9. CA also elevated the apoptotic rate. The levels of pro-apoptotic genes were upregulated while the levels of apoptosis inhibitory genes were decreased which further confirmed the pro-apoptotic effect of CA. In order to explore the mechanism of CA-induced apoptosis, insulin-like growth factor-1 (IGF-1) and PI3K inhibitor (LY294002) were used to regulate the phosphoinositide 3-kinase (PI3K)/AKT pathway. The transcription activity of PI3K/AKT was markedly inhibited by CA, as well as IGF-1 which functions as an anti-apoptotic factor. In conclusion, CA has the potential to be developed as a new antitumor drug. The mechanisms of action involve the regulation of expression of genes involved in apoptosis, invasion and adhesion via inhibition of the PI3K/Akt signaling pathway.

  2. Progesterone is neuroprotective against ischemic brain injury through its effects on the PI3K/Akt signaling pathway

    PubMed Central

    Ishrat, Tauheed; Sayeed, Iqbal; Atif, Fahim; Hua, Fang; Stein, Donald G.

    2012-01-01

    We tested the hypothesis that the phosphatidylinositol-3 kinase (PI3K/Akt) pathway mediates some of the neuroprotective effects of progesterone (PROG) after ischemic stroke. We examined whether PROG acting through the PI3K/Akt pathway could affect the expression of vascular endothelial growth factor (VEGF) and brain-derived neurotrophic factor (BDNF). Rats underwent permanent focal cerebral ischemia (pMCAO) by electro-coagulation and received intraperitoneal injections of PROG (8mg/kg) or vehicle at 1h post-occlusion and subcutaneous injections at 6, 24, and 48h. PAkt/Akt levels, apoptosis and apoptosis-related proteins (pBAD, BAD, caspase-3, and cleaved caspase-3) were analyzed by TUNEL assays, Western blotting and immunohistochemistry at 24h post-pMCAO. VEGF and BDNF were analyzed at 24, 72h and 14 days post-pMCAO with Western blots. Following pMCAO, PROG treatment significantly (p<0.05) reduced ischemic lesion size and edema. Treatment with PROG significantly (p<0.05) decreased VEGF at 24 and 72h but increased VEGF expression 14d after injury. The treatment also increased BDNF, and attenuated apoptosis by increasing Akt phosphorylation compared to vehicle-alone. The selective PI3K inhibitor Wortmannin compromised PROG-induced neuroprotective effects and reduced the elevation of pAkt levels in the ischemic penumbra. Our findings lead us to suggest that the PI3K/Akt pathway can play a role in mediating the neuroprotective effects of PROG after stroke by altering the expression of trophic factors in the brain. PMID:22450229

  3. Krüppel-like factor 14 increases insulin sensitivity through activation of PI3K/Akt signal pathway.

    PubMed

    Yang, Min; Ren, Yan; Lin, Zhimin; Tang, Chenchen; Jia, Yanjun; Lai, Yerui; Zhou, Tingting; Wu, Shaobo; Liu, Hua; Yang, Gangyi; Li, Ling

    2015-11-01

    Genome-wide association studies (GWAS) have shown that Krüppel-like factor 14 (KLF14) is associated with type 2 diabetes mellitus (T2DM). However, no report has demonstrated a relationship between KLF14 and glucose metabolism. The aim of this study was to determine whether KLF14 is associated with glucose metabolism and insulin signaling in vitro. The mRNA and protein expressions of KLF14 were determined by Real-time PCR and Western blotting. Glucose uptake was assessed by 2-[(3)H]-deoxyglucose (2-DG) uptake. Western blotting was used to identify the activation of insulin signaling proteins. KLF14 mRNA and protein in fat and muscle were significantly decreased in HFD-fed mice, db/db mice and T2DM patients. Overexpression of KLF14 enhanced insulin-stimulated glucose uptake and the activation of Akt kinase in Hepa1-6 cells. The phosphorylation of insulin receptor (InsR), insulin receptor substrate-1(IRS-1), glycogen synthase kinase-3β (GSK-3β) and Akt also elevated significantly by up-regulation of KLF14. KLF14 overexpression in Hepa1-6 cells prevented the inhibition of glucose uptake and Akt phosphorylation induced by high glucose and/or high insulin, or T2DM serum. However, KLF14's ability to increase glucose uptake and Akt activation was significantly attenuated by LY294002, a PI3-kinase inhibitor. These data suggested that KLF14 could increase insulin sensitivity probably through the PI3K/Akt pathway. PMID:26226221

  4. Interactions between Ras1, dMyc, and dPI3K signaling in the developing Drosophila wing

    PubMed Central

    Prober, David A.; Edgar, Bruce A.

    2002-01-01

    The Ras GTPase links extracellular signals to intracellular mechanisms that control cell growth, the cell cycle, and cell identity. An activated form of Drosophila Ras (RasV12) promotes these processes in the developing wing, but the effector pathways involved are unclear. Here, we present evidence indicating that RasV12 promotes cell growth and G1/S progression by increasing dMyc protein levels and activating dPI3K signaling, and that it does so via separate effector pathways. We also show that endogenous Ras is required to maintain normal levels of dMyc, but not dPI3K signaling during wing development. Finally, we show that induction of dMyc and regulation of cell identity are separable effects of Raf/MAPK signaling. These results suggest that Ras may only affect PI3K signaling when mutationally activated, such as in RasV12-transformed cells, and provide a basis for understanding the synergy between Ras and other growth-promoting oncogenes in cancer. PMID:12208851

  5. The Role of EGFR/PI3K/Akt/cyclinD1 Signaling Pathway in Acquired Middle Ear Cholesteatoma

    PubMed Central

    Liu, Wei; Ren, Hongmiao; Ren, Jihao; Yin, Tuanfang; Hu, Bing; Xie, Shumin; Dai, Yinghuan; Wu, Weijing; Xiao, Zian; Yang, Xinming; Xie, Dinghua

    2013-01-01

    Cholesteatoma is a benign keratinizing and hyper proliferative squamous epithelial lesion of the temporal bone. Epidermal growth factor (EGF) is one of the most important cytokines which has been shown to play a critical role in cholesteatoma. In this investigation, we studied the effects of EGF on the proliferation of keratinocytes and EGF-mediated signaling pathways underlying the pathogenesis of cholesteatoma. We examined the expressions of phosphorylated EGF receptor (p-EGFR), phosphorylated Akt (p-Akt), cyclinD1, and proliferating cell nuclear antigen (PCNA) in 40 cholesteatoma samples and 20 samples of normal external auditory canal (EAC) epithelium by immunohistochemical method. Furthermore, in vitro studies were performed to investigate EGF-induced downstream signaling pathways in primary external auditory canal keratinocytes (EACKs). The expressions of p-EGFR, p-Akt, cyclinD1, and PCNA in cholesteatoma epithelium were significantly increased when compared with those of control subjects. We also demonstrated that EGF led to the activation of the EGFR/PI3K/Akt/cyclinD1 signaling pathway, which played a critical role in EGF-induced cell proliferation and cell cycle progression of EACKs. Both EGFR inhibitor AG1478 and PI3K inhibitor wortmannin inhibited the EGF-induced EGFR/PI3K/Akt/cyclinD1 signaling pathway concomitantly with inhibition of cell proliferation and cell cycle progression of EACKs. Taken together, our data suggest that the EGFR/PI3K/Akt/cyclinD1 signaling pathway is active in cholesteatoma and may play a crucial role in cholesteatoma epithelial hyper-proliferation. This study will facilitate the development of potential therapeutic targets for intratympanic drug therapy for cholesteatoma. PMID:24311896

  6. The PI3K/mTOR inhibitor PF-04691502 induces apoptosis and inhibits microenvironmental signaling in CLL and the Eµ-TCL1 mouse model.

    PubMed

    Blunt, Matthew D; Carter, Matthew J; Larrayoz, Marta; Smith, Lindsay D; Aguilar-Hernandez, Maria; Cox, Kerry L; Tipton, Thomas; Reynolds, Mark; Murphy, Sarah; Lemm, Elizabeth; Dias, Samantha; Duncombe, Andrew; Strefford, Jonathan C; Johnson, Peter W M; Forconi, Francesco; Stevenson, Freda K; Packham, Graham; Cragg, Mark S; Steele, Andrew J

    2015-06-25

    Current treatment strategies for chronic lymphocytic leukemia (CLL) involve a combination of conventional chemotherapeutics, monoclonal antibodies, and targeted signaling inhibitors. However, CLL remains largely incurable, with drug resistance and treatment relapse a common occurrence, leading to the search for novel treatments. Mechanistic target of rapamycin (mTOR)-specific inhibitors have been previously assessed but their efficacy is limited due to a positive feedback loop via mTOR complex 2 (mTORC2), resulting in activation of prosurvival signaling. In this study, we show that the dual phosphatidylinositol 3-kinase (PI3K)/mTOR inhibitor PF-04691502 does not induce an mTORC2 positive feedback loop similar to other PI3K inhibitors but does induce substantial antitumor effects. PF-04691502 significantly reduced survival coincident with the induction of Noxa and Puma, independently of immunoglobulin heavy chain variable region mutational status, CD38, and ZAP-70 expression. PF-04691502 inhibited both anti-immunoglobulin M-induced signaling and overcame stroma-induced survival signals and migratory stimuli from CXCL12. Equivalent in vitro activity was seen in the Eμ-TCL1 murine model of CLL. In vivo, PF-04691502 treatment of tumor-bearing animals resulted in a transient lymphocytosis, followed by a clear reduction in tumor in the blood, bone marrow, spleen, and lymph nodes. These data indicate that PF-04691502 or other dual PI3K/mTOR inhibitors in development may prove efficacious for the treatment of CLL, increasing our armamentarium to successfully manage this disease.

  7. 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.

  8. Class III PI3K regulates organismal glucose homeostasis by providing negative feedback on hepatic insulin signalling

    PubMed Central

    Nemazanyy, Ivan; Montagnac, Guillaume; Russell, Ryan C.; Morzyglod, Lucille; Burnol, Anne-Françoise; Guan, Kun-Liang; Pende, Mario; Panasyuk, Ganna

    2015-01-01

    Defective hepatic insulin receptor (IR) signalling is a pathogenic manifestation of metabolic disorders including obesity and diabetes. The endo/lysosomal trafficking system may coordinate insulin action and nutrient homeostasis by endocytosis of IR and the autophagic control of intracellular nutrient levels. Here we show that class III PI3K—a master regulator of endocytosis, endosomal sorting and autophagy—provides negative feedback on hepatic insulin signalling. The ultraviolet radiation resistance-associated gene protein (UVRAG)-associated class III PI3K complex interacts with IR and is stimulated by insulin treatment. Acute and chronic depletion of hepatic Vps15, the regulatory subunit of class III PI3K, increases insulin sensitivity and Akt signalling, an effect that requires functional IR. This is reflected by FoxO1-dependent transcriptional defects and blunted gluconeogenesis in Vps15 mutant cells. On depletion of Vps15, the metabolic syndrome in genetic and diet-induced models of insulin resistance and diabetes is alleviated. Thus, feedback regulation of IR trafficking and function by class III PI3K may be a therapeutic target in metabolic conditions of insulin resistance. PMID:26387534

  9. Intestinal trefoil factor activates the PI3K/Akt signaling pathway to protect gastric mucosal epithelium from damage.

    PubMed

    Sun, Zhaorui; Liu, Hongmei; Yang, Zhizhou; Shao, Danbing; Zhang, Wei; Ren, Yi; Sun, Baodi; Lin, Jinfeng; Xu, Min; Nie, Shinan

    2014-09-01

    Intestinal trefoil factor (ITF, also named as trefoil factor 3, TFF3) is a member of the TFF-domain peptide family, which plays an essential role in the regulation of cell survival, cell migration and maintains mucosal epithelial integrity in the gastrointestinal tract. However, the underlying mechanisms and associated molecules remain unclear. The aim of this study was to explore the protective effects of ITF on gastric mucosal epithelium injury and its possible molecular mechanisms of action. In the present study, we show that ITF was able to promote the proliferation and migration of GES-1 cells via a mechanism that involves the PI3K/Akt signaling pathway. Western blot results indicated that ITF induced a dose- and time-dependent increase in the Akt signaling pathway. ITF also plays an essential role in the restitution of GES-1 cell damage induced by lipopolysaccharide (LPS). LPS induced the apoptosis of GES-1 cells, decreased cell viability significantly (P<0.01) and led to epithelial tight junction damage, which is attenuated via ITF treatment. The protective effect of ITF on the integrity of GES-1 was abrogated by inhibition of the PI3K/Akt pathway. Taken together, our results demonstrate that ITF promotes the proliferation and migration of gastric mucosal epithelial cells and preserves gastric mucosal epithelial integrity after damage is mediated by activation of the PI3K/Akt signaling pathway. This study suggested that the PI3K/Akt pathway could act as a key intracellular pathway in the gastric mucosal epithelium that may serve as a therapeutic target to preserve epithelial integrity during injury.

  10. Wogonin inhibits LPS-induced tumor angiogenesis via suppressing PI3K/Akt/NF-κB signaling.

    PubMed

    Zhao, Kai; Song, Xiuming; Huang, Yujie; Yao, Jing; Zhou, Mi; Li, Zhiyu; You, Qidong; Guo, Qinglong; Lu, Na

    2014-08-15

    Wogonin has been shown to have anti-angiogenesis and anti-tumor effects. However, whether wogonin inhibits LPS-induced tumor angiogenesis is not well known. In this study, we investigated the effect of wogonin on inhibiting LPS-induced tumor angiogenesis and further probed the underlying mechanisms. ELISA results revealed that wogonin could suppress LPS-induced VEGF secretion from tumor cells. Transwell assay, tube formation assay, rat aortic ring assay and CAM model were used to evaluate the effect of wogonin on angiogenesis induced by MCF-7 cell (treated with LPS) in vitro and in vivo. The inhibitory effect of wogonin on angiogenesis in LPS-treated MCF-7 cells was then confirmed by the above in vitro and in vivo assays. The study of the molecular mechanism showed that wogonin could suppress PI3K/Akt signaling activation. Moreover, wogonin inhibited nuclear translocation of NF-κB and its binding to DNA. The result of real-time PCR and luciferase reporter assay suggested that VEGF expression was down-regulated by wogonin primarily at the transcriptional level. IGF-1 and p65 expression plasmid were used to activate PI3K/Akt and NF-κB pathways, and to observe the effect of wogonin on the simualtion of PI3K/Akt/NF-κB signaling. Taken together, the result suggested that wogonin was a potent inhibitor of tumor angiogenesis and provided a new insight into the mechanisms of wogonin against cancer.

  11. Chinese herbal medicine formula tao hong si wu decoction protects against cerebral ischemia-reperfusion injury via PI3K/Akt and the Nrf2 signaling pathway.

    PubMed

    Li, Li; Yang, Na; Nin, Ling; Zhao, Zhilong; Chen, Lu; Yu, Jie; Jiang, Zhuyun; Zhong, Zhendong; Zeng, Daiwen; Qi, Hongyi; Xu, Xiaoyu

    2015-01-01

    The Chinese herbal medicine formula Tao Hong Si Wu decoction (THSWD) is traditionally used for the prevention and treatment of ischemic stroke. Transcription factor NF-E2-related factor 2 (Nrf2) regulates a battery of phase II enzymes and is known as the major mechanism of cellular defense against oxidative stress. The present study aimed to explore the potential effect of THSWD on the Nrf2 signaling pathway and the consequent effect during cerebral ischemia-reperfusion (I/R) injury. We found that THSWD reduced infarct volume and improved neurological function in a rat stroke model induced by middle cerebral artery occlusion (MCAO). Additionally, heme oxygenase 1 (HO-1), a key endogenous antioxidant enzyme regulated by Nrf2, was significantly further induced by THSWD in this in vivo model. In neuronal-like PC12 cells, THSWD remarkably up-regulated HO-1 expression and promoted Nrf2 nuclear translocation. Furthermore, phosphatidylinositol 3-kinase (PI3K)/Akt kinase was found to be involved in the upstream of Nrf2 regulation. In an in vitro oxygen-glucose deprivation/reperfusion (OGD-Rep) model, THSWD treatment significantly reduced cell death induced by OGD-Rep insult. Importantly, the protective action was attenuated while PI3K activity was inhibited by a specific inhibitor, LY294002, and the Nrf2 signaling pathway was blocked by antioxidant response element (ARE) decoy oligonucleotides. Collectively, these results demonstrated that THSWD exhibited notable neuroprotective properties in vitro and in vivo and activation of PI3K/Akt and the Nrf2 signaling pathway may be, at least in part, responsible for the protection. This study provides a better understanding of the molecular mechanism underlying the traditional use of the Chinese herbal medicine formula THSWD.

  12. Inhibition of PI3K-Akt Signaling Blocks Exercise-Mediated Enhancement of Adult Neurogenesis and Synaptic Plasticity in the Dentate Gyrus

    PubMed Central

    Bruel-Jungerman, Elodie; Veyrac, Alexandra; Dufour, Franck; Horwood, Jennifer; Laroche, Serge; Davis, Sabrina

    2009-01-01

    Background Physical exercise has been shown to increase adult neurogenesis in the dentate gyrus and enhances synaptic plasticity. The antiapoptotic kinase, Akt has also been shown to be phosphorylated following voluntary exercise; however, it remains unknown whether the PI3K-Akt signaling pathway is involved in exercise-induced neurogenesis and the associated facilitation of synaptic plasticity in the dentate gyrus. Methodology/Principal Findings To gain insight into the potential role of this signaling pathway in exercise-induced neurogenesis and LTP in the dentate gyrus rats were infused with the PI3K inhibitor, LY294002 or vehicle control solution (icv) via osmotic minipumps and exercised in a running wheel for 10 days. Newborn cells in the dentate gyrus were date-labelled with BrdU on the last 3 days of exercise. Then, they were either returned to the home cage for 2 weeks to assess exercise-induced LTP and neurogenesis in the dentate gyrus, or were killed on the last day of exercise to assess proliferation and activation of the PI3K-Akt cascade using western blotting. Conclusions/Significance Exercise increases cell proliferation and promotes survival of adult-born neurons in the dentate gyrus. Immediately after exercise, we found that Akt and three downstream targets, BAD, GSK3β and FOXO1 were activated. LY294002 blocked exercise-induced phosphorylation of Akt and downstream target proteins. This had no effect on exercise-induced cell proliferation, but it abolished most of the beneficial effect of exercise on the survival of newly generated dentate gyrus neurons and prevented exercise-induced increase in dentate gyrus LTP. These results suggest that activation of the PI3 kinase-Akt signaling pathway plays a significant role via an antiapoptotic function in promoting survival of newly formed granule cells generated during exercise and the associated increase in synaptic plasticity in the dentate gyrus. PMID:19936256

  13. The Molecular Interaction of CAR and JAML Recruits the Central Cell Signal Transducer PI3K

    SciTech Connect

    Verdino, Petra; Witherden, Deborah A.; Havran, Wendy L.; Wilson, Ian A.

    2010-11-15

    Coxsackie and adenovirus receptor (CAR) is the primary cellular receptor for group B coxsackieviruses and most adenovirus serotypes and plays a crucial role in adenoviral gene therapy. Recent discovery of the interaction between junctional adhesion molecule-like protein (JAML) and CAR uncovered important functional roles in immunity, inflammation, and tissue homeostasis. Crystal structures of JAML ectodomain (2.2 angstroms) and its complex with CAR (2.8 angstroms) reveal an unusual immunoglobulin-domain assembly for JAML and a charged interface that confers high specificity. Biochemical and mutagenesis studies illustrate how CAR-mediated clustering of JAML recruits phosphoinositide 3-kinase (P13K) to a JAML intracellular sequence motif as delineated for the {alpha}{beta} T cell costimulatory receptor CD28. Thus, CAR and JAML are cell signaling receptors of the immune system with implications for asthma, cancer, and chronic nonhealing wounds.

  14. Knockdown of TACC3 inhibits trophoblast cell migration and invasion through the PI3K/Akt signaling pathway.

    PubMed

    Zhu, Xiaojun; Cao, Qianqian; Li, Xia; Wang, Zhengping

    2016-10-01

    The insufficient invasion of trophoblasts is known to be correlated with the development of preeclampsia. Transforming acidic coiled‑coil protein 3 (TACC3), a member of the TACC domain family, is important in the regulation of cell differentiation, migration and invasion. However, the role of TACC3 in trophoblast function during placental development remains to be fully elucidated. The present study aimed to determine the expression and function of TACC3 in human placenta and to examine the underlying mechanisms. TACC3 expression was analyzed in preeclamptic placentas using reverse transcription‑quantitative polymerase chain reaction and western blotting. Cell proliferation was determined by the MTT assay, and cell migration and invasion were measured using Transwell assays. The expression levels of TACC3, matrix metalloproteinase (MMP)‑2, MMP‑9, tissue inhibitor of metalloproteinase (TIMP)‑1, TIMP‑2, phosphoinositide 3‑kinase (PI3K), phosphorylated (p)‑PI3K, AKT and p‑AKT were detected by western blotting. The results showed that the expression of TACC3 was downregulated in preeclamptic placentas. The knockdown of TACC3 significantly inhibited HTR8/SVneo cell proliferation, migration and invasion, and inhibited the expression of matrix metalloproteinases. In addition, the knockdown of TACC3 significantly reduced the levels of p‑PI3K and Akt in the HTR8/SVneo cells. Taken together, the results of the present study demonstrated that the knockdown of TACC3 inhibited the migration and invasion of HTR8/SVneo cells through suppression of the PI3K/Akt signaling pathway. Therefore, TACC3 may serve as a novel potential target for treating preeclampsia. PMID:27572091

  15. FAK mediates a compensatory survival signal parallel to PI3K-AKT in PTEN-null T-ALL cells.

    PubMed

    You, Dewen; Xin, Junping; Volk, Andrew; Wei, Wei; Schmidt, Rachel; Scurti, Gina; Nand, Sucha; Breuer, Eun-Kyoung; Kuo, Paul C; Breslin, Peter; Kini, Ameet R; Nishimura, Michael I; Zeleznik-Le, Nancy J; Zhang, Jiwang

    2015-03-31

    Mutations and inactivation of phosphatase and tensin homolog deleted from chromosome 10 (PTEN) are observed in 15%-25% of cases of human T cell acute lymphoblastic leukemia (T-ALL). Pten deletion induces myeloproliferative disorders (MPDs), acute myeloid leukemia (AML), and/or T-ALL in mice. Previous studies attributed Pten-loss-related hematopoietic defects and leukemogenesis to excessive activation of phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR signaling. Although inhibition of this signal dramatically suppresses the growth of PTEN-null T-ALL cells in vitro, treatment with inhibitors of this pathway does not cause a complete remission in vivo. Here, we report that focal adhesion kinase (Fak), a protein substrate of Pten, also contributes to T-ALL development in Pten-null mice. Inactivation of the FAK signaling pathway by either genetic or pharmacologic methods significantly sensitizes both murine and human PTEN-null T-ALL cells to PI3K/AKT/mTOR inhibition when cultured in vitro on feeder layer cells or a matrix and in vivo. PMID:25801032

  16. PI3K regulates endocytosis after insulin secretion by mediating signaling crosstalk between Arf6 and Rab27a.

    PubMed

    Yamaoka, Mami; Ando, Tomomi; Terabayashi, Takeshi; Okamoto, Mitsuhiro; Takei, Masahiro; Nishioka, Tomoki; Kaibuchi, Kozo; Matsunaga, Kohichi; Ishizaki, Ray; Izumi, Tetsuro; Niki, Ichiro; Ishizaki, Toshimasa; Kimura, Toshihide

    2016-02-01

    In secretory cells, endocytosis is coupled to exocytosis to enable proper secretion. Although endocytosis is crucial to maintain cellular homeostasis before and after secretion, knowledge about secretagogue-induced endocytosis in secretory cells is still limited. Here, we searched for proteins that interacted with the Rab27a GTPase-activating protein (GAP) EPI64 (also known as TBC1D10A) and identified the Arf6 guanine-nucleotide-exchange factor (GEF) ARNO (also known as CYTH2) in pancreatic β-cells. We found that the insulin secretagogue glucose promotes phosphatidylinositol (3,4,5)-trisphosphate (PIP3) generation through phosphoinositide 3-kinase (PI3K), thereby recruiting ARNO to the intracellular side of the plasma membrane. Peripheral ARNO promotes clathrin assembly through its GEF activity for Arf6 and regulates the early stage of endocytosis. We also found that peripheral ARNO recruits EPI64 to the same area and that the interaction requires glucose-induced endocytosis in pancreatic β-cells. Given that GTP- and GDP-bound Rab27a regulate exocytosis and the late stage of endocytosis, our results indicate that the glucose-induced activation of PI3K plays a pivotal role in exocytosis-endocytosis coupling, and that ARNO and EPI64 regulate endocytosis at distinct stages.

  17. Thymoquinone attenuates liver fibrosis via PI3K and TLR4 signaling pathways in activated hepatic stellate cells.

    PubMed

    Bai, Ting; Lian, Li-Hua; Wu, Yan-Ling; Wan, Ying; Nan, Ji-Xing

    2013-02-01

    Thymoquinone (TQ) is the major active compound derived from the medicinal Nigella sativa. In the present study, we investigated the anti-fibrotic mechanism of TQ in lipopolysaccharide (LPS)-activated rat hepatic stellate cells line, T-HSC/Cl-6. T-HSC/Cl-6 cells were treated with TQ (3.125, 6.25 and 12.5μM) prior to LPS (1μg/ml). Our data demonstrated that TQ effectively decreased activated T-HSC/Cl-6 cell viability. TQ significantly attenuated the expression of CD14 and Toll-like receptor 4 (TLR4). TQ also significantly inhibited phosphatidylinositol 3-kinase (PI3K) and serine/threonine kinase-protein kinase B (Akt) phosphorylation. The expression of α-SMA and collagen-I were significantly decreased by TQ. Furthermore, TQ decreased X linked inhibitor of apoptosis (XIAP) and cellular FLIP (c-FLIPL) expression, which are related with the regulation of apoptosis. Furthermore, TQ significantly increased the survival against LPS challenge in d-galactosamine (d-GlaN)-sensitized mice, and decreased the levels of serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST), which were in line with in vitro results. Our data demonstrated that TQ attenuates liver fibrosis partially via blocking TLR4 expression and PI3K phosphorylation on the activated HSCs. Therefore, TQ may be a potential candidate for the therapy of hepatic fibrosis.

  18. Catalase expression in MCF-7 breast cancer cells is mainly controlled by PI3K/Akt/mTor signaling pathway.

    PubMed

    Glorieux, Christophe; Auquier, Julien; Dejeans, Nicolas; Sid, Brice; Demoulin, Jean-Baptiste; Bertrand, Luc; Verrax, Julien; Calderon, Pedro Buc

    2014-05-15

    Catalase is an antioxidant enzyme that catalyzes mainly the transformation of hydrogen peroxide into water and oxygen. Although catalase is frequently down-regulated in tumors the underlying mechanism remains unclear. Few transcription factors have been reported to directly bind the human catalase promoter. Among them FoxO3a has been proposed as a positive regulator of catalase expression. Therefore, we decided to study the role of the transcription factor FoxO3a and the phosphatidylinositol-3 kinase (PI3K) signaling pathway, which regulates FoxO3a, in the expression of catalase. To this end, we developed an experimental model of mammary breast MCF-7 cancer cells that acquire resistance to oxidative stress, the so-called Resox cells, in which catalase is overexpressed as compared with MCF-7 parental cell line. In Resox cells, Akt expression is decreased but its phosphorylation is enhanced when compared with MCF-7 cells. A similar profile is observed for FoxO3a, with less total protein but more phosphorylated FoxO3a in Resox cells, correlating with its higher Akt activity. The modulation of FoxO3a expression by knockdown and overexpression strategies did not affect catalase expression, neither in MCF-7 nor in Resox cells. Inhibition of PI3K and mTOR by LY295002 and rapamycin, respectively, decreases the phosphorylation of downstream targets (i.e. GSK3β and p70S6K) and leads to an increase of catalase expression only in MCF-7 but not in Resox cells. In conclusion, FoxO3a does not appear to play a critical role in the regulation of catalase expression in both cancer cells. Only MCF-7 cells are sensitive and dependent on PI3K/Akt/mTOR signaling.

  19. 2,2',4,4'-Tetrabromodiphenyl ether promotes human neuroblastoma SH-SY5Y cells migration via the GPER/PI3K/Akt signal pathway.

    PubMed

    Tian, P-C; Wang, H-L; Chen, G-H; Luo, Q; Chen, Z; Wang, Y; Liu, Y-F

    2016-02-01

    Neuroblastoma is the predominant tumor of early childhood. 2,2',4,4'-Tetrabromodiphenyl ether (BDE-47) has the highest concentration among all polybrominated diphenyl ether (PBDE) congeners in human body, particularly for children. Considering that accumulating evidences showed developmental neurotoxicity of PBDE, there is an urgent need to investigate the effects of BDE-47 on the development of neuroblastoma. This study revealed that BDE-47 had limited effects on the cytotoxicity while significantly increased the in vitro migration and invasion of human neuroblastoma SH-SY5Y cells. This was further confirmed by the results that BDE-47 treatment significantly downregulated the expression of E-cadherin and zona occludin-1 and upregulated the expression of matrix metalloproteinase-9 (MMP-9). Silencing of MMP-9 by specific small interfering RNA significantly abolished the BDE-47-induced migration and invasion of SH-SY5Y cells. Further, the signals G protein-coupled estrogen receptor 1 (GPER)/phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/protein kinase B (Akt) mediated the BDE-47-induced upregulation of MMP-9 and in vitro migration of SH-SY5Y cells since G15 (GPER inhibitor) and LY 294002 (PI3K/Akt inhibitor) significantly abolished the effects of BDE-47. Our results revealed that BDE-47 significantly triggered the metastasis of human neuroblastoma SH-SY5Y cells via upregulation of MMP-9 by the GPER/PI3K/Akt signal pathway. This study revealed for the first time that BDE-47 can promote the migration of SH-SY5Y cells. It also provided a better understanding about the metastasis of human neuroblastoma induced by environmental endocrine disruptors.

  20. Heat Stress-Induced PI3K/mTORC2-Dependent AKT Signaling Is a Central Mediator of Hepatocellular Carcinoma Survival to Thermal Ablation Induced Heat Stress.

    PubMed

    Thompson, Scott M; Callstrom, Matthew R; Jondal, Danielle E; Butters, Kim A; Knudsen, Bruce E; Anderson, Jill L; Lien, Karen R; Sutor, Shari L; Lee, Ju-Seog; Thorgeirsson, Snorri S; Grande, Joseph P; Roberts, Lewis R; Woodrum, David A

    2016-01-01

    Thermal ablative therapies are important treatment options in the multidisciplinary care of patients with hepatocellular carcinoma (HCC), but lesions larger than 2-3 cm are plagued with high local recurrence rates and overall survival of these patients remains poor. Currently no adjuvant therapies exist to prevent local HCC recurrence in patients undergoing thermal ablation. The molecular mechanisms mediating HCC resistance to thermal ablation induced heat stress and local recurrence remain unclear. Here we demonstrate that the HCC cells with a poor prognostic hepatic stem cell subtype (Subtype HS) are more resistant to heat stress than HCC cells with a better prognostic hepatocyte subtype (Subtype HC). Moreover, sublethal heat stress rapidly induces phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) dependent-protein kinase B (AKT) survival signaling in HCC cells in vitro and at the tumor ablation margin in vivo. Conversely, inhibition of PI3K/mTOR complex 2 (mTORC2)-dependent AKT phosphorylation or direct inhibition of AKT function both enhance HCC cell killing and decrease HCC cell survival to sublethal heat stress in both poor and better prognostic HCC subtypes while mTOR complex 1 (mTORC1)-inhibition has no impact. Finally, we showed that AKT isoforms 1, 2 and 3 are differentially upregulated in primary human HCCs and that overexpression of AKT correlates with worse tumor biology and pathologic features (AKT3) and prognosis (AKT1). Together these findings define a novel molecular mechanism whereby heat stress induces PI3K/mTORC2-dependent AKT survival signaling in HCC cells and provide a mechanistic rationale for adjuvant AKT inhibition in combination with thermal ablation as a strategy to enhance HCC cell killing and prevent local recurrence, particularly at the ablation margin. PMID:27611696

  1. Apoptosis Induction of Human Prostate Carcinoma DU145 Cells by Diallyl Disulfide via Modulation of JNK and PI3K/AKT Signaling Pathways

    PubMed Central

    Shin, Dong Yeok; Kim, Gi-Young; Lee, Jun Hyuk; Choi, Byung Tae; Yoo, Young Hyun; Choi, Yung Hyun

    2012-01-01

    Diallyl disulfide (DADS), a sulfur compound derived from garlic, has various biological properties, such as anticancer, antiangiogenic and anti-inflammatory effects. However, the mechanisms of action underlying the compound’s anticancer activity have not been fully elucidated. In this study, the apoptotic effects of DADS were investigated in DU145 human prostate carcinoma cells. Our results showed that DADS markedly inhibited the growth of the DU145 cells by induction of apoptosis. Apoptosis was accompanied by modulation of Bcl-2 and inhibitor of apoptosis protein (IAP) family proteins, depolarization of the mitochondrial membrane potential (MMP, ΔΨm) and proteolytic activation of caspases. We also found that the expression of death-receptor 4 (DR4) and Fas ligand (FasL) proteins was increased and that the level of intact Bid proteins was down-regulated by DADS. Moreover, treatment with DADS induced phosphorylation of mitogen-activated protein kinases (MAPKs), including extracellular-signal regulating kinase (ERK), p38 MAPK and c-Jun N-terminal kinase (JNK). A specific JNK inhibitor, SP600125, significantly blocked DADS-induced-apoptosis, whereas inhibitors of the ERK (PD98059) and p38 MAPK (SB203580) had no effect. The induction of apoptosis was also accompanied by inactivation of phosphatidylinositol 3-kinase (PI3K)/Akt and the PI3K inhibitor LY29004 significantly increased DADS-induced cell death. These findings provide evidence demonstrating that the proapoptotic effect of DADS is mediated through the activation of JNK and the inhibition of the PI3K/Akt signaling pathway in DU145 cells. PMID:23203057

  2. Heat Stress-Induced PI3K/mTORC2-Dependent AKT Signaling Is a Central Mediator of Hepatocellular Carcinoma Survival to Thermal Ablation Induced Heat Stress.

    PubMed

    Thompson, Scott M; Callstrom, Matthew R; Jondal, Danielle E; Butters, Kim A; Knudsen, Bruce E; Anderson, Jill L; Lien, Karen R; Sutor, Shari L; Lee, Ju-Seog; Thorgeirsson, Snorri S; Grande, Joseph P; Roberts, Lewis R; Woodrum, David A

    2016-01-01

    Thermal ablative therapies are important treatment options in the multidisciplinary care of patients with hepatocellular carcinoma (HCC), but lesions larger than 2-3 cm are plagued with high local recurrence rates and overall survival of these patients remains poor. Currently no adjuvant therapies exist to prevent local HCC recurrence in patients undergoing thermal ablation. The molecular mechanisms mediating HCC resistance to thermal ablation induced heat stress and local recurrence remain unclear. Here we demonstrate that the HCC cells with a poor prognostic hepatic stem cell subtype (Subtype HS) are more resistant to heat stress than HCC cells with a better prognostic hepatocyte subtype (Subtype HC). Moreover, sublethal heat stress rapidly induces phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) dependent-protein kinase B (AKT) survival signaling in HCC cells in vitro and at the tumor ablation margin in vivo. Conversely, inhibition of PI3K/mTOR complex 2 (mTORC2)-dependent AKT phosphorylation or direct inhibition of AKT function both enhance HCC cell killing and decrease HCC cell survival to sublethal heat stress in both poor and better prognostic HCC subtypes while mTOR complex 1 (mTORC1)-inhibition has no impact. Finally, we showed that AKT isoforms 1, 2 and 3 are differentially upregulated in primary human HCCs and that overexpression of AKT correlates with worse tumor biology and pathologic features (AKT3) and prognosis (AKT1). Together these findings define a novel molecular mechanism whereby heat stress induces PI3K/mTORC2-dependent AKT survival signaling in HCC cells and provide a mechanistic rationale for adjuvant AKT inhibition in combination with thermal ablation as a strategy to enhance HCC cell killing and prevent local recurrence, particularly at the ablation margin.

  3. Heat Stress-Induced PI3K/mTORC2-Dependent AKT Signaling Is a Central Mediator of Hepatocellular Carcinoma Survival to Thermal Ablation Induced Heat Stress

    PubMed Central

    Thompson, Scott M.; Callstrom, Matthew R.; Jondal, Danielle E.; Butters, Kim A.; Knudsen, Bruce E.; Anderson, Jill L.; Lien, Karen R.; Sutor, Shari L.; Lee, Ju-Seog; Thorgeirsson, Snorri S.; Grande, Joseph P.; Roberts, Lewis R.; Woodrum, David A.

    2016-01-01

    Thermal ablative therapies are important treatment options in the multidisciplinary care of patients with hepatocellular carcinoma (HCC), but lesions larger than 2–3 cm are plagued with high local recurrence rates and overall survival of these patients remains poor. Currently no adjuvant therapies exist to prevent local HCC recurrence in patients undergoing thermal ablation. The molecular mechanisms mediating HCC resistance to thermal ablation induced heat stress and local recurrence remain unclear. Here we demonstrate that the HCC cells with a poor prognostic hepatic stem cell subtype (Subtype HS) are more resistant to heat stress than HCC cells with a better prognostic hepatocyte subtype (Subtype HC). Moreover, sublethal heat stress rapidly induces phosphoinositide 3-kinase (PI3K)/mammalian target of rapamycin (mTOR) dependent-protein kinase B (AKT) survival signaling in HCC cells in vitro and at the tumor ablation margin in vivo. Conversely, inhibition of PI3K/mTOR complex 2 (mTORC2)-dependent AKT phosphorylation or direct inhibition of AKT function both enhance HCC cell killing and decrease HCC cell survival to sublethal heat stress in both poor and better prognostic HCC subtypes while mTOR complex 1 (mTORC1)-inhibition has no impact. Finally, we showed that AKT isoforms 1, 2 and 3 are differentially upregulated in primary human HCCs and that overexpression of AKT correlates with worse tumor biology and pathologic features (AKT3) and prognosis (AKT1). Together these findings define a novel molecular mechanism whereby heat stress induces PI3K/mTORC2-dependent AKT survival signaling in HCC cells and provide a mechanistic rationale for adjuvant AKT inhibition in combination with thermal ablation as a strategy to enhance HCC cell killing and prevent local recurrence, particularly at the ablation margin. PMID:27611696

  4. 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. PMID:24390959

  5. Inhibition of the PI3K/Akt/mTOR signaling pathway in diffuse large B-cell lymphoma: current knowledge and clinical significance.

    PubMed

    Majchrzak, Agata; Witkowska, Magdalena; Smolewski, Piotr

    2014-09-11

    Diffuse large B-cell lymphoma (DLBCL) is one of the most common non-Hodgkin lymphomas in adults. The disease is very heterogeneous in its presentation, that is DLBCL patients may differ from each other not only in regard to histology of tissue infiltration, clinical course or response to treatment, but also in respect to diversity in gene expression profiling. A growing body of knowledge on the biology of DLBCL, including abnormalities in intracellular signaling, has allowed the development of new treatment strategies, specifically directed against lymphoma cells. The phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR) signaling pathway plays an important role in controlling proliferation and survival of tumor cells in various types of malignancies, including DLBCL, and therefore it may be a promising target for therapeutic intervention. Currently, novel anticancer drugs are undergoing assessment in different phases of clinical trials in aggressive lymphomas, with promising outcomes. In this review we present a state of art review on various classes of small molecule inhibitors selectively involving PI3K/Akt/mTOR pathway and their clinical potential in this disease.

  6. Effect of Compound Chuanxiong Capsule on Inflammatory Reaction and PI3K/Akt/NF-κB Signaling Pathway in Atherosclerosis.

    PubMed

    Kang, Qunfu; Liu, Weihong; Liu, Hongxu; Zhou, Mingxue

    2015-01-01

    Compound Chuanxiong Capsule (CCC), a Chinese herbal compound, can exhibit antiatherosclerotic effect; however, its mechanism is still unclear. This study is designed to study the mechanism of CCC on atherosclerosis in the ApoE-knockout (ApoE(-/-)) mice fed with a high-fat diet. After 6 weeks of high-fat feeding, 40 ApoE(-/-) mice were randomized (n = 10) and treated with lipitor, high-dose or low-dose CCC, or distilled water (ApoE(-/-) group) for 7 weeks. The blood lipids in serum and the plaque areas of the mice were measured and the mRNA expressions of phosphatidylinositol-3-kinases (PI3K), Akt, nuclear factor-kappa B (NF-κB), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) of the aortae were determined. The data showed that CCC can significantly decrease the levels of blood lipids, atherosclerosis index, and plaque areas and increase collagen proportion in plaques as compared with the untreated mice (p < 0.05, p < 0.01). In addition, CCC can significantly reduce the mRNA expressions of PI3K, Akt, NF-κB, IL-6, and TNF-α in the mice fed with a high-fat diet (p < 0.001). Thus, we concluded that CCC can inhibit inflammatory reaction in the ApoE(-/-) mice fed with a high-fat diet. This mechanism may be attributed to regulating PI3K/Akt/NF-κB signaling pathway. PMID:26539229

  7. Effect of Compound Chuanxiong Capsule on Inflammatory Reaction and PI3K/Akt/NF-κB Signaling Pathway in Atherosclerosis

    PubMed Central

    Kang, Qunfu; Liu, Weihong; Liu, Hongxu; Zhou, Mingxue

    2015-01-01

    Compound Chuanxiong Capsule (CCC), a Chinese herbal compound, can exhibit antiatherosclerotic effect; however, its mechanism is still unclear. This study is designed to study the mechanism of CCC on atherosclerosis in the ApoE-knockout (ApoE−/−) mice fed with a high-fat diet. After 6 weeks of high-fat feeding, 40 ApoE−/− mice were randomized (n = 10) and treated with lipitor, high-dose or low-dose CCC, or distilled water (ApoE−/− group) for 7 weeks. The blood lipids in serum and the plaque areas of the mice were measured and the mRNA expressions of phosphatidylinositol-3-kinases (PI3K), Akt, nuclear factor-kappa B (NF-κB), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) of the aortae were determined. The data showed that CCC can significantly decrease the levels of blood lipids, atherosclerosis index, and plaque areas and increase collagen proportion in plaques as compared with the untreated mice (p < 0.05, p < 0.01). In addition, CCC can significantly reduce the mRNA expressions of PI3K, Akt, NF-κB, IL-6, and TNF-α in the mice fed with a high-fat diet (p < 0.001). Thus, we concluded that CCC can inhibit inflammatory reaction in the ApoE−/− mice fed with a high-fat diet. This mechanism may be attributed to regulating PI3K/Akt/NF-κB signaling pathway. PMID:26539229

  8. Nerve Regeneration Potential of Protocatechuic Acid in RSC96 Schwann Cells by Induction of Cellular Proliferation and Migration through IGF-IR-PI3K-Akt Signaling.

    PubMed

    Ju, Da-Tong; Liao, Hung-En; Shibu, Marthandam Asokan; Ho, Tsung-Jung; Padma, Viswanadha Vijaya; Tsai, Fuu-Jen; Chung, Li-Chin; Day, Cecilia Hsuan; Lin, Chien-Chung; Huang, Chih-Yang

    2015-12-31

    Peripheral nerve injuries, caused by accidental trauma, acute compression or surgery, often result in temporary or life-long neuronal dysfunctions and inflict great economic or social burdens on the patients. Nerve cell proliferation is an essential process to restore injured nerves of adults. Schwann cells play a crucial role in endogenous repair of peripheral nerves due to their ability to proliferate, migrate and provide trophic support to axons via expression of various neurotrophic factors, such as the nerve growth factor (NGF), especially after nerve injury. Protocatechuic acid (PCA) is a dihydroxybenzoic acid, a type of phenolic acid, isolated from the kernels of Alpinia oxyphylla Miq (AOF), a traditional Chinese herbal medicine the fruits of which are widely used as a tonic, aphrodisiac, anti-salivation and anti-diarrheatic. This study investigated the molecular mechanisms by which PCA induces Schwann cell proliferation by activating IGF-IR-PI3K-Akt pathway. Treatment with PCA induces phosphorylation of the insulin-like growth factor-I (IGF-I)-mediated phosphatidylinositol 3 kinase/serine - threonine kinase (PI3K/Akt) pathway, and activates expression of cell nuclear antigen (PCNA) in a dose-dependent manner. Cell cycle analysis after 18 h of treatment showed that proliferation of the RSC96 cells was enhanced by PCA treatment. The PCA induced proliferation was accompanied by modulation in the expressions of cell cycle proteins cyclin D1, cyclin E and cyclin A. Knockdown of PI3K using small interfering RNA (siRNA) and inhibition of IGF-IR receptor resulted in the reduction in cell survival proteins. The results collectively showed that PCA treatment promoted cell proliferation and cell survival via IGF-I signaling. PMID:26717920

  9. Macrophage SR-BI mediates efferocytosis via Src/PI3K/Rac1 signaling and reduces atherosclerotic lesion necrosis[S

    PubMed Central

    Tao, Huan; Yancey, Patricia G.; Babaev, Vladimir R.; Blakemore, John L.; Zhang, Youmin; Ding, Lei; Fazio, Sergio; Linton, MacRae F.

    2015-01-01

    Macrophage apoptosis and efferocytosis are key determinants of atherosclerotic plaque inflammation and necrosis. Bone marrow transplantation studies in ApoE- and LDLR-deficient mice revealed that hematopoietic scavenger receptor class B type I (SR-BI) deficiency results in severely defective efferocytosis in mouse atherosclerotic lesions, resulting in a 17-fold higher ratio of free to macrophage-associated dead cells in lesions containing SR-BI−/− cells, 5-fold more necrosis, 65.2% less lesional collagen content, nearly 7-fold higher dead cell accumulation, and 2-fold larger lesion area. Hematopoietic SR-BI deletion elicited a maladaptive inflammatory response [higher interleukin (IL)-1β, IL-6, and TNF-α lower IL-10 and transforming growth factor β]. Efferocytosis of apoptotic thymocytes was reduced by 64% in SR-BI−/− versus WT macrophages, both in vitro and in vivo. In response to apoptotic cells, macrophage SR-BI bound with phosphatidylserine and induced Src phosphorylation and cell membrane recruitment, which led to downstream activation of phosphoinositide 3-kinase (PI3K) and Ras-related C3 botulinum toxin substrate 1 (Rac1) for engulfment and clearance of apoptotic cells, as inhibition of Src decreased PI3K, Rac1-GTP, and efferocytosis in WT cells. Pharmacological inhibition of Rac1 reduced macrophage efferocytosis in a SR-BI-dependent fashion, and activation of Rac1 corrected the defective efferocytosis in SR-BI−/− macrophages. Thus, deficiency of macrophage SR-BI promotes defective efferocytosis signaling via the Src/PI3K/Rac1 pathway, resulting in increased plaque size, necrosis, and inflammation. PMID:26059978

  10. The pyrrolidinoindoline alkaloid Psm2 inhibits platelet aggregation and thrombus formation by affecting PI3K/Akt signaling

    PubMed Central

    Su, Xing-li; Su, Wen; Wang, Ying; Wang, Yue-hu; Ming, Xin; Kong, Yi

    2016-01-01

    Aim: Psm2, one of the pyrrolidinoindoline alkaloids isolated from whole Selaginella moellendorffii plants, has shown a potent antiplatelet activity. In this study, we further evaluated the antiplatelet effects of Psm2, and elucidated the underlying mechanisms. Methods: Human platelet aggregation in vitro and rat platelet aggregation ex vivo were investigated. Agonist-induced platelet aggregation was measured using a light transmission aggregometer. The antithrombotic effects of Psm2 were evaluated in arteriovenous shunt thrombosis model in rats. To elucidate the mechanisms underlying the antiplatelet activity of Psm2, ELISAs, Western blotting and molecular docking were performed. The bleeding risk of Psm2 administration was assessed in a mouse tail cutting model, and the cytotoxicity of Psm2 was measured with MTT assay in EA.hy926 cells. Results: Psm2 dose-dependently inhibited human platelet aggregation induced by ADP, U4619, thrombin and collagen with IC50 values of 0.64, 0.37, 0.35 and 0.87 mg/mL, respectively. Psm2 (1, 3, 10 mg/kg) administered to rats significantly inhibited platelet aggregation ex vivo induced by ADP. Psm2 (1, 3, 10 mg/mL, iv) administered to rats with the A–V shunt dose-dependently decreased the thrombus formation. Psm2 inhibited platelet adhesion to fibrinogen and collagen with IC50 values of 84.5 and 96.5 mg/mL, respectively, but did not affect the binding of fibrinogen to GPIIb/IIIa. Furthermore, Psm2 inhibited AktSer473 phosphorylation, but did not affect MAPK signaling and Src kinase activation. Molecular docking showed that Psm2 bound to phosphatidylinositol 3-kinase β (PI3Kβ) with a binding free energy of −13.265 kcal/mol. In addition, Psm2 did not cause toxicity in EA.hy926 cells and produced only slight bleeding in a mouse tail cutting model. Conclusion: Psm2 inhibits platelet aggregation and thrombus formation by affecting PI3K/Akt signaling. Psm2 may be a lead compound or drug candidate that could be developed for the

  11. Effects of dexmedetomidine postconditioning on myocardial ischemia and the role of the PI3K/Akt-dependent signaling pathway in reperfusion injury

    PubMed Central

    CHENG, XIANG YANG; GU, XIAO YU; GAO, QIN; ZONG, QIAO FENG; LI, XIAO HONG; ZHANG, YE

    2016-01-01

    The present study aimed to determine whether post-ischemic treatment with dexmedetomidine (DEX) protected the heart against acute myocardial ischemia/reperfusion (I/R)-induced injury in rats. The phosphatidylinositol-3 kinase/protein kinase B(PI3K/Akt)-dependent signaling pathway was also investigated. Male Sprague Dawley rats (n=64) were subjected to ligation of the left anterior descending artery (LAD), which produced ischemia for 25 min, followed by reperfusion. Following LAD ligation, rats were treated with DEX (5, 10 and 20 µg/kg) or underwent post-ischemic conditioning, which included three cycles of ischemic insult. In order to determine the role of the PI3K/Akt signaling pathway, wortmannin (Wort), a PI3K inhibitor, was used to treat a group of rats that had also been treated with DEX (20 µg/kg). Post-reperfusion, lactate dehydrogenase (LDH), cardiac troponin I (cTnI), creatine kinase isoenzymes (CK-MB), superoxide dismutase (SOD) and malondialdehyde (MDA) serum levels were measured using an ultraviolet spectrophotometer. The protein expression levels of phosphorylated (p)-Akt, Ser9-p-glycogen synthase kinase-3β (p-GSK-3β) and cleaved caspase-3 were detected in heart tissue by western blotting. The mRNA expression levels of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were detected using reverse transcription-polymerase chain reaction. At the end of the experiment, the hearts were removed and perfused in an isolated perfusion heart apparatus with Evans blue (1%) in order to determine the non-ischemic areas. The risk and infarct areas of the heart were not dyed. As expected, I/R induced myocardial infarction, as determined by the increased serum levels of cTnI, CK-MB and MDA, and the decreased levels of SOD. Post-ischemic treatment with DEX increased the expression levels of p-Akt and p-GSK-3β, whereas caspase-3 expression was reduced following DEX treatment compared with in the I/R group. Compared with the I/R group, the ratio of Bcl

  12. 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

  13. Salidroside Improves Behavioral and Histological Outcomes and Reduces Apoptosis via PI3K/Akt Signaling after Experimental Traumatic Brain Injury

    PubMed Central

    Chen, Szu-Fu; Tsai, Hsin-Ju; Hung, Tai-Ho; Chen, Chien-Cheng; Lee, Chao Yu; Wu, Chun-Hu; Wang, Pei-Yi; Liao, Nien-Chieh

    2012-01-01

    Background Traumatic brain injury (TBI) induces a complex sequence of apopototic cascades that contribute to secondary tissue damage. The aim of this study was to investigate the effects of salidroside, a phenolic glycoside with potent anti-apoptotic properties, on behavioral and histological outcomes, brain edema, and apoptosis following experimental TBI and the possible involvement of the phosphoinositide 3-kinase/protein kinase B (PI3K)/Akt signaling pathway. Methodology/Principal Findings Mice subjected to controlled cortical impact injury received intraperitoneal salidroside (20, or 50 mg/kg) or vehicle injection 10 min after injury. Behavioral studies, histology analysis and brain water content assessment were performed. Levels of PI3K/Akt signaling-related molecules, apoptosis-related proteins, cytochrome C (CytoC), and Smac/DIABLO were also analyzed. LY294002, a PI3K inhibitor, was administered to examine the mechanism of protection. The protective effect of salidroside was also investigated in primary cultured neurons subjected to stretch injury. Treatment with 20 mg/kg salidroside_significantly improved functional recovery and reduced brain tissue damage up to post-injury day 28. Salidroside_also significantly reduced neuronal death, apoptosis, and brain edema at day 1. These changes were associated with significant decreases in cleaved caspase-3, CytoC, and Smac/DIABLO at days 1 and 3. Salidroside increased phosphorylation of Akt on Ser473 and the mitochondrial Bcl-2/Bax ratio at day 1, and enhanced phosphorylation of Akt on Thr308 at day 3. This beneficial effect was abolished by pre-injection of LY294002. Moreover, delayed administration of salidroside at 3 or 6 h post-injury reduced neuronal damage at day 1. Salidroside treatment also decreased neuronal vulnerability to stretch-induced injury in vitro. Conclusions/Significance Post-injury salidroside improved long-term behavioral and histological outcomes and reduced brain edema and apoptosis

  14. Downregulation of PI3K/Akt/mTOR signaling pathway in curcumin-induced autophagy in APP/PS1 double transgenic mice.

    PubMed

    Wang, Chen; Zhang, Xiong; Teng, Zhipeng; Zhang, Tong; Li, Yu

    2014-10-01

    Autophagy is a lysosomal degradation pathway, which is essential for cell survival, proliferation, differentiation and homeostasis. It is well known that beta-amyloid (Aβ) aggregation is one of key characteristics for Alzheimer's disease (AD), which triggers a complex pathological cascade, leading to neurodegeneration. Recent studies have shown that Aβ peptide is generated from amyloid β precursor protein (APP) during autophagic turnover of APP-rich organelles by autophagy. Aβ generation during normal autophagy is subsequently degraded by lysosomes. Curcumin, a nature plant extraction, has been reported to inhibit the generation and deposition of Aβ; however, the underlying mechanisms are not fully understood yet. In the present study, we reported that curcumin treatment not only attenuated cognitive impairment detected by Morris water maze test, but also inhibited the generation of Aβ investigated by immunohistochemistry in APP/PS1 double transgenic AD mice. Moreover, curcumin induced autophagy in the mice, evidenced by LC3 immunofluorescence analysis and western blot assays on LC3. Furthermore, we found that curcumin significantly decreased the expression of Phosphatidylinositol 3-Kinase (PI3K), phosphorylated Akt and rapamycin (mTOR) at protein levels, respectively. Taken together, our data suggests that curcumin inhibits Aβ generation and induces of autophagy by downregulating PI3K/Akt/mTOR signaling pathway, and further shows a neuroprotective effect. Meanwhile curcumin might be a candidate neuroprotective agent for AD patients treatment by inducing autophagy.

  15. Interleukin-21 promotes osteoclastogenesis in RAW264.7 cells through the PI3K/AKT signaling pathway independently of RANKL.

    PubMed

    Xing, Rui; Zhang, Yingjian; Li, Changhong; Sun, Lin; Yang, Lin; Zhao, Jinxia; Liu, Xiangyuan

    2016-10-01

    Cytokines play a key role in the bone destruction of rheumatoid arthritis (RA). Interleukin-21 (IL-21) promotes osteoclastogenesis in RA in a receptor activator of nuclear factor-κB ligand (RANKL)-dependent way. Whether IL-21 is capable of promoting osteoclastogenesis directly in the absence of RANKL remains unknown. In the present study, we examined the osteoclastogenic activity of IL-21 in RAW264.7 cells in the absence of RANKL. We found that IL-21 enhanced osteoclastogenesis and this was demonstrated by increased numbers of tartrate-resistant acid phosphatase (TRAP)-positive stained, multinucleated cells compared with the negative control. Western blot analysis and immunocytochemistry showed the positive expression of calcitonin receptor (CTR) in the IL-21 group. RT-PCR and RT-qPCR also verified the increased mRNA expression of CTR and cathepsin K in the IL-21 group compared with the negative control. The scanning electronic microscope images showed a few resorption pits on the bone slices cultured with IL-21. The phosphoinositide 3-kinase (PI3K)/AKT pathway inhibitor LY294002 significantly suppressed IL-21-induced osteoclastogenesis. Taken together, these findings suggest that IL-21 has direct osteoclastogenic potential independently of RANKL. IL-21 may promote osteoclastogenesis through the PI3K/AKT signaling pathway. Therapy targeting IL-21 may be of value in preventing bone erosions in patients with RA. PMID:27599586

  16. Interleukin-21 promotes osteoclastogenesis in RAW264.7 cells through the PI3K/AKT signaling pathway independently of RANKL

    PubMed Central

    Xing, Rui; Zhang, Yingjian; Li, Changhong; Sun, Lin; Yang, Lin; Zhao, Jinxia; Liu, Xiangyuan

    2016-01-01

    Cytokines play a key role in the bone destruction of rheumatoid arthritis (RA). Interleukin-21 (IL-21) promotes osteoclastogenesis in RA in a receptor activator of nuclear factor-κB ligand (RANKL)-dependent way. Whether IL-21 is capable of promoting osteoclastogenesis directly in the absence of RANKL remains unknown. In the present study, we examined the osteoclastogenic activity of IL-21 in RAW264.7 cells in the absence of RANKL. We found that IL-21 enhanced osteoclastogenesis and this was demonstrated by increased numbers of tartrate-resistant acid phosphatase (TRAP)-positive stained, multinucleated cells compared with the negative control. Western blot analysis and immunocytochemistry showed the positive expression of calcitonin receptor (CTR) in the IL-21 group. RT-PCR and RT-qPCR also verified the increased mRNA expression of CTR and cathepsin K in the IL-21 group compared with the negative control. The scanning electronic microscope images showed a few resorption pits on the bone slices cultured with IL-21. The phosphoinositide 3-kinase (PI3K)/AKT pathway inhibitor LY294002 significantly suppressed IL-21-induced osteoclastogenesis. Taken together, these findings suggest that IL-21 has direct osteoclastogenic potential independently of RANKL. IL-21 may promote osteoclastogenesis through the PI3K/AKT signaling pathway. Therapy targeting IL-21 may be of value in preventing bone erosions in patients with RA. PMID:27599586

  17. Hypoglycemic effect of D-chiro-inositol in type 2 diabetes mellitus rats through the PI3K/Akt signaling pathway.

    PubMed

    Gao, Yun-Feng; Zhang, Meng-Na; Wang, Tian-Xin; Wu, Tian-Chen; Ai, Ru-Dan; Zhang, Ze-Sheng

    2016-09-15

    In this investigation, a model of type 2 diabetes mellitus (T2DM) was used on Sprague-Dawley (SD) rats to clarify more details of the mechanism in the therapy of T2DM. D-chiro-inositol (DCI) was administrated to the diabetic rats as two doses [30, 60 mg/(kg·body weight·day)]. The biochemical indices revealed that DCI had a positive effect on hypoglycemic activity and promoted the glycogen synthesis. The rats in DCI high-dosage group had a blood glucose reduction rate of 21.5% after 5 weeks of treatment, and had insulin content in serum about 15.3 ± 2.37 mIU/L which was significantly decreased than diabetes control group. Real-time polymerase chain reaction (RT-PCR) results revealed that DCI gave a positive regulation on glycogen synthase (GS) and protein glucose transporter-4 (Glut4). Western blotting suggested that DCI could up-regulated the expression of the phosphatidylinositol-3-kinase (PI3K) p85, PI3Kp110, GS as well as the phosphorylation of protein kinase B (Akt) both in the liver and the skeletal muscle. The results also revealed that DCI enhanced the Glut4 expression on skeletal muscle. Above all, DCI played a positive role in regulating insulin-mediated glucose uptake through the PI3K/Akt signaling pathway in T2DM rats.

  18. MicroRNA-214 acts as a potential oncogene in breast cancer by targeting the PTEN-PI3K/Akt signaling pathway.

    PubMed

    Wang, Fang; Li, Lin; Chen, Zhuo; Zhu, Mingzhi; Gu, Yuanting

    2016-05-01

    Breast cancer ranks as the leading cause of cancer-related mortality in females worldwide. It has been proven that microRNAs (miRNAs or miRs), a type of non‑coding RNA, are involved in tumorigenesis. An increasing number of studies has confirmed the critical role of miR‑214 in certain types of cancer. Nevertheless, the biological function of miR‑214, as well as its underlying mechanisms of action in breast cancer remain largely unknown. In the present study, the expression of miR‑214 was found to be upregulated in four human breast cancer cell lines in contrast to its expression level in the non‑malignant breast epithelial cell line, MCF‑10A. Moreover, the overexpression of miR‑214 markedly increased cell viability and abrogated the apoptosis triggered by serum starvation, indicating that miR‑214 plays a pivotal role in breast cancer cell growth. Further analysis suggested that the upregulation of miR‑214 markedly induced the activation of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway, which largely accounted for the protective effects of miR‑124 on cancer cell growth. This was further confimed by pre‑treatment with the PI3K/Akt inhibitor, LY294002, which markedly attenuated the miR‑214‑induced increase in cell viability and resistance to apoptosis. Furthermore, the expression of phosphatase and tensin homolog (PTEN) was decreased following transfection wtih miR‑214 mimics and PTEN was confirmed as the direct target of miR‑214 by bioinformatics analysis and a dual‑firefly luciferase reporter assay. Importantly, the introduction of PTEN cDNA lacking the 3' untranslated region (3'UTR) significantly inhibited the miR‑214‑induced activation of the PI3K/Akt signaling pathway, and abrogated the protetive effects of miR‑214 on cell survival and resistance to apoptosis. Taken together, these findings suggest that miR‑214 possesses oncogenic activity and that its effects are mediated through the promotion of cell growth

  19. Recent Syntheses of PI3K/Akt/mTOR Signaling Pathway Inhibitors

    PubMed Central

    2013-01-01

    This review focuses on the syntheses of PI3K/Akt/mTOR inhibitors that have been reported outside of the patent literature in the last 5 years but is largely centered on synthetic work reported in 2011 and 2012. While focused on syntheses of inhibitors, some information on in vitro and in vivo testing of compounds is also included. Many of these reported compounds are reversible, competitive adenosine triphosphate (ATP) binding inhibitors, so given the structural similarities of many of these compounds to the adenine core, this review presents recent work on inhibitors based on where the synthetic chemistry was started, i.e. inhibitor syntheses which started with purines/pyrimidines are followed by inhibitor syntheses which began with pyridines, pyrazines, azoles, and triazines then moves to inhibitors which bear no structural resemblance to adenine: liphagal, wortmannin and quercetin analogs. The review then finishes with a short section on recent syntheses of phosphotidyl inositol (PI) analogs since competitive PI binding inhibitors represent an alternative to the competitive ATP binding inhibitors which have received the most attention. PMID:23735831

  20. Layered double hydroxide nanoparticles promote self-renewal of mouse embryonic stem cells through the PI3K signaling pathway

    NASA Astrophysics Data System (ADS)

    Wu, Youjun; Zhu, Rongrong; Zhou, Yang; Zhang, Jun; Wang, Wenrui; Sun, Xiaoyu; Wu, Xianzheng; Cheng, Liming; Zhang, Jing; Wang, Shilong

    2015-06-01

    Embryonic stem cells (ESCs) hold great potential for regenerative medicine due to their two unique characteristics: self-renewal and pluripotency. Several groups of nanoparticles have shown promising applications in directing the stem cell fate. Herein, we investigated the cellular effects of layered double hydroxide nanoparticles (LDH NPs) on mouse ESCs (mESCs) and the associated molecular mechanisms. Mg-Al-LDH NPs with an average diameter of ~100 nm were prepared by hydrothermal methods. To determine the influences of LDH NPs on mESCs, cellular cytotoxicity, self-renewal, differentiation potential, and the possible signaling pathways were explored. Evaluation of cell viability, lactate dehydrogenase release, ROS generation and apoptosis demonstrated the low cytotoxicity of LDH NPs. The alkaline phosphatase activity and the expression of pluripotency genes in mESCs were examined, which indicated that exposure to LDH NPs could support self-renewal and inhibit spontaneous differentiation of mESCs under feeder-free culture conditions. The self-renewal promotion was further proved to be independent of the leukemia inhibitory factor (LIF). Furthermore, cells treated with LDH NPs maintained the potential to differentiate into all three germ layers both in vitro and in vivo through formation of embryoid bodies and teratomas. In addition, we observed that LDH NPs initiated the activation of the PI3K/Akt pathway, while treatment with the PI3K inhibitor LY294002 could block the effects of LDH NPs on mESCs. The results confirmed that the promotion of self-renewal by LDH NPs was associated with activation of the PI3K/Akt signaling pathway. Altogether, our studies identified a new role of LDH NPs in maintaining self-renewal of mouse ES cells which could potentially be applied in stem cell research.Embryonic stem cells (ESCs) hold great potential for regenerative medicine due to their two unique characteristics: self-renewal and pluripotency. Several groups of nanoparticles

  1. PI3K inhibitors for cancer therapy: what has been achieved so far?

    PubMed

    Wu, Peng; Liu, Tao; Hu, Yongzhou

    2009-01-01

    PI3K is a large duel lipid and protein kinase that catalyzes phosphorylation of the 3-hydroxyl position of phosphatidylinositides (PIs) and plays a crucial role in the cellular signaling network. Inhibition of the phosphatidylinositol 3-kinase (PI3K) signaling pathway is a newly identified strategy for the discovery and development of certain therapeutic agents. Among the various subtypes of PI3K, class IA PI3Kalpha has gained increasing attention as a promising drug target for the treatment of cancer due to its frequent mutations and amplifications in various human cancers. Here, we discuss the insights gained so far relevant to the development of PI3K inhibitors for the treatment of human cancers. Emphasis is on the structure-activity relationship of PI3K inhibitors which bear the most significant PI3Kalpha inhibitory activities. We also highlight PI3K inhibitors that are currently under clinical trials for cancers. PMID:19275602

  2. Phosphoinositide 3-kinase mediated signaling in lobster olfactory receptor neurons.

    PubMed

    Corey, Elizabeth A; Bobkov, Yuriy; Pezier, Adeline; Ache, Barry W

    2010-04-01

    In vertebrates and some invertebrates, odorant molecules bind to G protein-coupled receptors on olfactory receptor neurons (ORNs) to initiate signal transduction. Phosphoinositide 3-kinase (PI3K) activity has been implicated physiologically in olfactory signal transduction, suggesting a potential role for a G protein-coupled receptor-activated class I PI3K. Using isoform-specific antibodies, we identified a protein in the olfactory signal transduction compartment of lobster ORNs that is antigenically similar to mammalian PI3Kgamma and cloned a gene for a PI3K with amino acid homology with PI3Kbeta. The lobster olfactory PI3K co-immunoprecipitates with the G protein alpha and beta subunits, and an odorant-evoked increase in phosphatidylinositol (3,4,5)-trisphosphate can be detected in the signal transduction compartment of the ORNs. PI3Kgamma and beta isoform-specific inhibitors reduce the odorant-evoked output of lobster ORNs in vivo. Collectively, these findings provide evidence that PI3K is indeed activated by odorant receptors in lobster ORNs and further support the potential involvement of G protein activated PI3K signaling in olfactory transduction.

  3. Vitamin E succinate induces apoptosis via the PI3K/AKT signaling pathways in EC109 esophageal cancer cells.

    PubMed

    Yang, Peng; Zhao, Jiaying; Hou, Liying; Yang, Lei; Wu, Kun; Zhang, Linyou

    2016-08-01

    Esophageal cancer is the fourth most common gastrointestinal cancer, it generally has a poor prognosis and novel strategies are required for prevention and treatment. Vitamin E succinate (VES) is a potential chemical agent for cancer prevention and therapy as it exerts anti‑tumor effects in a variety of cancers. However, the role of VES in tumorigenesis and progression of cancer remains to be elucidated. The present study aimed to determine the effects of VES in regulating the survival and apoptosis of human esophageal cancer cells. EC109 human esophageal cancer cells were used to investigate the anti‑proliferative effects of VES. The MTT and Annexin V‑fluorescein isothiocyanate/propidium iodide assays demonstrated that VES inhibited cell proliferation and induced apoptosis in esophageal cancer cells. Furthermore, VES downregulated constitutively active basal levels of phosphorylated (p)‑serine‑threonine kinase AKT (AKT) and p‑mammalian target of rapamycin (mTOR), and decreased the phosphorylation of AKT substrates Bcl‑2‑associated death receptor and caspase‑9, in addition to mTOR effectors, ribosomal protein S6 kinase β1 and eIF4E‑binding protein 1. Phosphoinositide‑3‑kinase (PI3K) inhibitor, LY294002 suppressed p‑AKT and p‑mTOR, indicating PI3K is a common upstream mediator. The apoptosis induced by VES was increased by inhibition of AKT or mTOR with their respective inhibitor in esophageal cancer cells. The results of the present study suggested that VES targeted the PI3K/AKT signaling pathways and induced apoptosis in esophageal cancer cells. Furthermore, the current study suggests that VES may be useful in a combinational therapeutic strategy employing an mTOR inhibitor.

  4. Wogonin inhibits H2O2-induced angiogenesis via suppressing PI3K/Akt/NF-κB signaling pathway.

    PubMed

    Zhou, Mi; Song, Xiuming; Huang, Yujie; Wei, Libin; Li, Zhiyu; You, Qidong; Guo, Qinglong; Lu, Na

    2014-03-01

    Wogonin, a natural monoflavonoid extracted from Scutellariae radix, has been reported for its ability of inhibiting tumor angiogenesis. In this study, we assessed the effect of wogonin on angiogenesis induced by low level of H2O2 (10 μM) in human umbilical vein endothelial cells (HUVECs). Wogonin suppressed H2O2-induced migration and tube formation of HUVECs as well as microvessel sprouting from rat aortic rings in vitro. Meanwhile, wogonin suppressed vessel growth in chicken chorioallantoic membrane (CAM) model in vivo. Mechanistic studies showed that wogonin suppressed H2O2-activated PI3K/Akt pathway and reduced the expression of vascular endothelial growth factor (VEGF) up-regulated by H2O2 in both protein and mRNA levels. In addition, wogonin also inhibited nuclear translocation of NF-κB, and decreased the binding ability of NF-κB with exogenous consensus DNA oligonucleotide. Then we further investigated the effect of wogonin on over-activated PI3K/Akt pathway by insulin-like growth factor-1 (IGF-1) and H2O2. We found that wogonin suppressed phosphorylation of Akt, up-regulation of VEGF and angiogenesis in vitro which was further induced by IGF-1 and H2O2. Moreover, in NF-κB overexpressed HUVECs, wogonin could also reduce the expression of VEGF and inhibited the migration and tube formation. Taken together, these results suggested that wogonin was potential in inhibiting H2O2-induced angiogenesis in vitro and in vivo via suppressing PI3K/Akt pathway and NF-κB signaling.

  5. 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.

  6. Hesperetin Inhibits Vascular Formation by Suppressing of the PI3K/AKT, ERK, and p38 MAPK Signaling Pathways.

    PubMed

    Kim, Gi Dae

    2014-12-01

    Hesperetin has been shown to possess a potential anti-angiogenic effect, including vascular formation by endothelial cells. However, the mechanisms underlying the potential anti-angiogenic activity of hesperetin are not fully understood. In the present study, we evaluated whether hesperetin has anti-angiogenic effects in human umbilical vascular endothelial cells (HUVECs). HUVECs were treated with 50 ng/mL vascular endothelial growth factor (VEGF) to induce proliferation as well as vascular formation, followed by treatment with several doses of hesperetin (25, 50, and 100 μM) for 24 h. Cell proliferation and vascular formation were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and tube formation assay, respectively. In addition, cell signaling related to cell proliferation and vascular formation was analyzed by western blot. Furthermore, a mouse aorta ring assay was performed to confirm the effect of hesperetin on vascular formation. Hesperetin treatment did not cause differences in HUVECs proliferation. However, hesperetin significantly inhibited VEGF-induced cell migration and tube formation of HUVECs (P<0.05). Moreover, hesperetin suppressed the expression of ERK, p38 MAPK, and PI3K/AKT in the VEGF-induced HUVECs. In an ex vivo model, hesperetin also suppressed microvessel sprouting of mouse aortic rings. Taken together, the findings suggest that hesperetin inhibited vascular formation by endothelial cells via the inhibition of the PI3K/AKT, ERK and p38 MAPK signaling.

  7. Phosphoinositide 3-Kinase (PI3K) Subunit p110δ Is Essential for Trophoblast Cell Differentiation and Placental Development in Mouse

    PubMed Central

    Hu, Xiwen; Li, Jiangchao; Zhang, Qianqian; Zheng, Lingyun; Wang, Guang; Zhang, Xiaohan; Zhang, Jingli; Gu, Quliang; Ye, Yuxiang; Guo, Sun-Wei; Yang, Xuesong; Wang, Lijing

    2016-01-01

    Maternal PI3K p110δ has been implicated in smaller litter sizes in mice, but its underlying mechanism remains unclear. The placenta is an indispensable chimeric organ that supports mammalian embryonic development. Using a mouse model of genetic inactivation of PI3K p110δ (p110δD910A/D910A), we show that fetuses carried by p110δD910A/D910A females were growth retarded and showed increased mortality in utero mainly during placentation. The placentas in p110δD910A/D910A females were anomalously anemic, exhibited thinner spongiotrophoblast layer and looser labyrinth zone, which indicate defective placental vasculogenesis. In addition, p110δ was detected in primary trophoblast giant cells (P-TGC) at early placentation. Maternal PI3K p110δ inactivation affected normal TGCs generation and expansion, impeded the branching of chorioallantoic placenta but enhanced the expression of matrix metalloproteinases (MMP-2, MMP-12). Poor vasculature support for the developing fetoplacental unit resulted in fetal death or gross growth retardation. These data, taken together, provide the first in vivo evidence that p110δ may play an important role in placental vascularization through manipulating trophoblast giant cell. PMID:27306493

  8. PI3K-AKT signaling is a downstream effector of retinoid prevention of murine basal cell carcinogenesis

    PubMed Central

    So, Po-Lin; Wang, Grace Y.; Wang, Kevin; Chuang, Mindy; Calinisan Chiueh, Venice; Kenny, Paraic A.; Epstein, Ervin H.

    2014-01-01

    Basal cell carcinoma (BCC) is the most common human cancer. We have demonstrated previously that topical application of the retinoid prodrug tazarotene profoundly inhibits murine BCC carcinogenesis via RARγ-mediated regulation of tumor cell transcription. Since topical retinoids can cause adverse cutaneous effects and since tumors can develop resistance to retinoids, we have investigated mechanisms downstream of tazarotene’s anti-tumor effect in this model. Specifically we have used (i) global expression profiling to identify and (ii) functional cell-based assays to validate the PI3K/AKT/mTOR pathway as a downstream target pathway of tazarotene’s action. Crucially, we have demonstrated that pharmacologic inhibition of this downstream pathway profoundly reduces murine BCC cell proliferation and tumorigenesis both in vitro and in vivo. These data identify PI3K/AKT/mTOR signaling as a highly attractive target for BCC chemoprevention and indicate more generally that this pathway may be, in some contexts, an important mediator of retinoid anti-cancer effects. PMID:24449057

  9. Tetramethylpyrazine attenuates TNF-α-induced iNOS expression in human endothelial cells: Involvement of Syk-mediated activation of PI3K-IKK-IκB signaling pathways

    SciTech Connect

    Zheng, Zhen; Li, Zhiliang; Chen, Song; Pan, Jieyi; Ma, Xiaochun

    2013-08-15

    Endothelial cells produce nitric oxide (NO) by activation of constitutive nitric oxide synthase (NOS) and transcription of inducible NO synthase (iNOS). We explored the effect of tetramethylpyrazine (TMP), a compound derived from chuanxiong, on tumor necrosis factor (TNF)-α-induced iNOS in human umbilical vein endothelial cells (HUVECs) and explored the signal pathways involved by using RT-PCR and Western blot. TMP suppressed TNF-α-induced expression of iNOS by inhibiting IκB kinase (IKK) phosphorylation, IκB degradation and nuclear factor κB (NF-κB) nuclear translocation, which were required for NO gene transcription. Exposure to wortmannin abrogated IKK/IκB/NF-κB-mediated iNOS expression, suggesting activation of such a signal pathway might be phosphoinositide-3-kinase (PI3K) dependent. Spleen tyrosine kinase (Syk) inhibitor piceatannol significantly inhibited NO production. Furthermore, piceatannol obviously suppressed TNF-α-induced IκB phosphorylation and the downstream NF-κB activation, suggesting that Syk is an upstream key regulator in the activation of PI3K/IKK/IκB-mediated signaling. TMP significantly inhibited TNF-α-induced phosphorylation of Syk and PI3K. Our data indicate that TMP might repress iNOS expression, at least in part, through its inhibitory effect of Syk-mediated PI3K phosphorylation in TNF-α-stimulated HUVECs. -- Highlights: •TMP suppressed TNF-α-induced expression of iNOS by inhibiting IKK/IκB/NF-κB pathway. •PI3K inhibitor wortmannin abrogated IKK/IκB/NF-κB-mediated iNOS expression. •Syk inhibitor piceatannol repressed PI3K/IKK/IκB mediated NO production. •Syk is an upstream regulator in the activation of PI3K/IKK/IκB-mediated signaling. •TMP might repress iNOS expression through Syk-mediated PI3K pathway.

  10. Insulin-like growth factor-I stimulates H{sub 4}II rat hepatoma cell proliferation: Dominant role of PI-3'K/Akt signaling

    SciTech Connect

    Alexia, Catherine; Fourmatgeat, Pascal; Delautier, Daniele; Groyer, Andre . E-mail: groyer@bichat.inserm.fr

    2006-04-15

    Although hepatocytes are the primary source of endocrine IGF-I and -II in mammals, their autocrine/paracrine role in the dysregulation of proliferation and apoptosis during hepatocarcinogenesis and in hepatocarcinomas (HCC) remains to be elucidated. Indeed, IGF-II and type-I IGF receptors are overexpressed in HCC cells, and IGF-I is synthesized in adjacent non-tumoral liver tissue. In the present study, we have investigated the effects of type-I IGF receptor signaling on H{sub 4}II rat hepatoma cell proliferation, as estimated by {sup 3}H-thymidine incorporation into DNA. IGF-I stimulated the rate of DNA synthesis of serum-deprived H{sub 4}II cells, stimulation being maximal 3 h after the onset of IGF-I treatment and remaining elevated until at least 6 h. The IGF-I-induced increase in DNA replication was abolished by LY294002 and only partially inhibited by PD98059, suggesting that phosphoinositol-3' kinase (PI-3'K) and to a lesser extent MEK/Erk signaling were involved. Furthermore, the 3- to 19-fold activation of the Erks in the presence of LY294002 suggested a down-regulation of the MEK/Erk cascade by PI-3'K signaling. Finally, the effect of IGF-I on DNA replication was almost completely abolished in clones of H{sub 4}II cells expressing a dominant-negative form of Akt but was unaltered by rapamycin treatment of wild-type H{sub 4}II cells. Altogether, these data support the notion that the stimulation of H{sub 4}II rat hepatoma cell proliferation by IGF-I is especially dependent on Akt activation but independent on the Akt/mTOR signal0009i.

  11. Allicin inhibits oxidative stress-induced mitochondrial dysfunction and apoptosis by promoting PI3K/AKT and CREB/ERK signaling in osteoblast cells

    PubMed Central

    DING, GUOLIANG; ZHAO, JIANQUAN; JIANG, DIANMING

    2016-01-01

    Osteoporosis is a disease of the skeleton that is characterized by the loss of bone mass and degeneration of bone microstructure, resulting in an increased risk of fracture. Oxidative stress, which is known to promote oxidative damage to mitochondrial function and also cell apoptosis, has been recently indicated to be implicated in osteoporosis. However, there are few agents that counteract oxidative stress in osteoporosis. In the present study, the protective effects of allicin against the oxidative stress-induced mitochondrial dysfunction and apoptosis were investigated in murine osteoblast-like MC3T3-E1 cells. The results demonstrated that allicin counteracted the reduction of cell viability and induction of apoptosis caused by hydrogen peroxide (H2O2) exposure. The inhibition of apoptosis by allicin was confirmed by the inhibition of H2O2-induced cytochrome c release and caspase-3 activation. Moreover, the inhibition of apoptosis by allicin was identified to be associated with the counteraction of H2O2-induced mitochondrial dysfunction. In addition, allicin was demonstrated to be able to significantly ameliorate the repressed phosphoinositide 3-kinase (PI3K)/AKT and cyclic adenosine monophosphate response element-binding protein (CREB)/extracellular-signal-regulated kinase (ERK) signaling pathways by H2O2, which may also be associated with the anti-oxidative stress effects of allicin. In conclusion, allicin protects osteoblasts from H2O2-induced oxidative stress and apoptosis in MC3T3-E1 cells by improving mitochondrial function and the activation of PI3K/AKT and CREB/ERK signaling. The present study implies a promising role of allicin in oxidative stress-associated osteoporosis. PMID:27284348

  12. {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.

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

    PubMed

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

    2016-10-01

    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

  14. Obese and lean porcine difference of FoxO1 and its regulation through C/EBPβ and PI3K/GSK3β signaling pathway.

    PubMed

    Pang, W J; Wei, N; Wang, Y; Xiong, Y; Chen, F F; Wu, W J; Zhao, C Z; Sun, S D; Yang, G S

    2014-05-01

    Forkhead box O 1 (FoxO1) is an important transcription factor implicated in adipogenesis. In this study, we detected the breed differences in FoxO1 between Bamei pigs (an obese breed) and Large White pigs (a lean breed). Compared with Large White pigs, the BW of Bamei pigs was lower (P < 0.01), but back fat thickness, fat percent, and intramuscular fat content were greater (P < 0.01). The levels of FoxO1 mRNA and protein were lower (P < 0.01) in subcutaneous adipose tissue (SAT) of Bamei pigs at 180 d, adipocytes and stromal-vascular fraction extracted from SAT of Bamei pigs at 1 d compared with Large White pigs. Knockdown of FoxO1 increased triglyceride content (P < 0.01) and upregulated the levels of adipocyte fatty-acid binding protein, PPARγ, and CCAAT enhancer-binding protein α (C/EBPα) at 6 d after porcine preadipocytes were induced. Furthermore, the transcriptional regulation of FoxO1 through C/EBPβ during early porcine preadipocyte differentiation and the effect of insulin on phosphoinositide 3 kinase (PI3K)/glycogen synthase kinase 3β (GSK3β) signal pathway by FoxO1 were examined. The results indicated that FoxO1 inhibited transcription activity of C/EBPβ, whereas C/EBPβ did not affect transcription activity of FoxO1. At 6 and 12 h of early differentiation, knockdown of FoxO1 triggered the transcription activity of C/EBPβ. In addition, FoxO1 protein interacted with C/EBPβ protein in porcine adipocytes at 12 h after induction. Under treatment with 100 nM insulin, knockdown or overexpression of FoxO1 mediated PI3K/GSK3β signaling via upregulating or downregulating the levels of GSK3β and its phosphorylation in adipocytes. Taken together, there is low, but detectable, expression of FoxO1 in SAT of obese pigs and FoxO1 inhibited adipogenesis through C/EBPβ and PI3K/GSK3β signaling pathway. These findings provide useful information to further the understanding of the function of FoxO1 in porcine adipogenesis. PMID:24663213

  15. Eukaryotic elongation factor-1α 2 knockdown inhibits hepatocarcinogenesis by suppressing PI3K/Akt/NF-κB signaling

    PubMed Central

    Qiu, Fu-Nan; Huang, Yi; Chen, Dun-Yan; Li, Feng; Wu, Yan-An; Wu, Wen-Bing; Huang, Xiao-Li

    2016-01-01

    AIM: To assess the impact of eukaryotic elongation factor 1 alpha 2 (eEF1A2) on hepatocellular carcinoma (HCC) cell proliferation, apoptosis, migration and invasion, and determine the underlying mechanisms. METHODS: eEF1A2 levels were detected in 62 HCC tissue samples and paired pericarcinomatous specimens, and the human HCC cell lines SK-HEP-1, HepG2 and BEF-7402, by real-time PCR and immunohistochemistry. Experimental groups included eEF1A2 silencing in BEL-7402 cells with lentivirus eEF1A2-shRNA (KD group) and eEF1A2 overexpression in SK-HEP-1 cells with eEF1A2 plasmid (OE group). Non-transfected cells (control group) and lentivirus-based empty vector transfected cells (NC group) were considered control groups. Cell proliferation (MTT and colony formation assays), apoptosis (Annexin V-APC assay), cell cycle (DNA ploidy assay), and migration and invasion (Transwell assays) were assessed. Protein levels of PI3K/Akt/NF-κB signaling effectors were evaluated by Western blot. RESULTS: eEF1A2 mRNA and protein levels were significantly higher in HCC cancer tissue samples than in paired pericarcinomatous and normal specimens. SK-HEP-1 cells showed lower eEF1A2 mRNA levels; HepG2 and BEL-7402 cells showed higher eEF1A2 mRNA levels, with BEL-7402 cells displaying the highest amount. Efficient eEF1A2 silencing resulted in reduced cell proliferation, migration and invasion, increased apoptosis, and induced cell cycle arrest. The PI3K/Akt/NF-κB signaling pathway was notably inhibited. Inversely, eEF1A2 overexpression resulted in promoted cell proliferation, migration and invasion. CONCLUSION: eEF1A2, highly expressed in HCC, is a potential oncogene. Its silencing significantly decreases HCC tumorigenesis, likely by inhibiting PI3K/Akt/NF-κB signaling. PMID:27122673

  16. Monomeric C-reactive protein and Notch-3 co-operatively increase angiogenesis through PI3K signalling pathway.

    PubMed

    Boras, Emhamed; Slevin, Mark; Alexander, M Yvonne; Aljohi, Ali; Gilmore, William; Ashworth, Jason; Krupinski, Jerzy; Potempa, Lawrence A; Al Abdulkareem, Ibrahim; Elobeid, Adila; Matou-Nasri, Sabine

    2014-10-01

    C-reactive protein (CRP) is the most acute-phase reactant serum protein of inflammation and a strong predictor of cardiovascular disease. Its expression is associated with atherosclerotic plaque instability and the formation of immature micro-vessels. We have previously shown that CRP upregulates endothelial-derived Notch-3, a key receptor involved in vascular development, remodelling and maturation. In this study, we investigated the links between the bioactive monomeric CRP (mCRP) and Notch-3 signalling in angiogenesis. We used in vitro (cell counting, wound-healing and tubulogenesis assays) and in vivo (chorioallantoic membrane) angiogenic assays and Western blotting to study the angiogenic signalling pathways induced by mCRP and Notch-3 activator chimera protein (Notch-3/Fc). Our results showed an additive effect on angiogenesis of mCRP stimulatory effect combined with Notch-3/Fc promoting bovine aortic endothelial cell (BAEC) proliferation, migration, tube formation in Matrigel(TM) with up-regulation of phospho-Akt expression. The pharmacological blockade of PI3K/Akt survival pathway by LY294002 fully inhibited in vitro and in vivo angiogenesis induced by mCRP/Notch-3/Fc combination while blocking Notch signalling by gamma-secretase inhibitor (DAPT) partially inhibited mCRP/Notch-3/Fc-induced angiogenesis. Using a BAEC vascular smooth muscle cell co-culture sprouting angiogenesis assay and transmission electron microscopy, we showed that activation of both mCRP and Notch-3 signalling induced the formation of thicker sprouts which were shown later by Western blotting to be associated with an up-regulation of N-cadherin expression and a down-regulation of VE-cadherin expression. Thus, mCRP combined with Notch-3 activator promote angiogenesis through the PI3K/Akt pathway and their therapeutic combination has potential to promote and stabilize vessel formation whilst reducing the risk of haemorrhage from unstable plaques. PMID:24972386

  17. Curcumin inhibited HGF-induced EMT and angiogenesis through regulating c-Met dependent PI3K/Akt/mTOR signaling pathways in lung cancer.

    PubMed

    Jiao, Demin; Wang, Jian; Lu, Wei; Tang, Xiali; Chen, Jun; Mou, Hao; Chen, Qing-Yong

    2016-01-01

    The epithelial-mesenchymal transition (EMT) and angiogenesis have emerged as two pivotal events in cancer progression. Curcumin has been extensively studied in preclinical models and clinical trials of cancer prevention due to its favorable toxicity profile. However, the possible involvement of curcumin in the EMT and angiogenesis in lung cancer remains unclear. This study found that curcumin inhibited hepatocyte growth factor (HGF)-induced migration and EMT-related morphological changes in A549 and PC-9 cells. Moreover, pretreatment with curcumin blocked HGF-induced c-Met phosphorylation and downstream activation of Akt, mTOR, and S6. These effects mimicked that of c-Met inhibitor SU11274 or PI3 kinase inhibitor LY294002 or mTOR inhibitor rapamycin treatment. c-Met gene overexpression analysis further demonstrated that curcumin suppressed lung cancer cell EMT by inhibiting c-Met/Akt/mTOR signaling pathways. In human umbilical vein endothelial cells (HUVECs), we found that curcumin also significantly inhibited PI3K/Akt/mTOR signaling and induced apoptosis and reduced migration and tube formation of HGF-treated HUVEC. Finally, in the experimental mouse model, we showed that curcumin inhibited HGF-stimulated tumor growth and induced an increase in E-cadherin expression and a decrease in vimentin, CD34, and vascular endothelial growth factor (VEGF) expression. Collectively, these findings indicated that curcumin could inhibit HGF-promoted EMT and angiogenesis by targeting c-Met and blocking PI3K/Akt/mTOR pathways. PMID:27525306

  18. Curcumin inhibited HGF-induced EMT and angiogenesis through regulating c-Met dependent PI3K/Akt/mTOR signaling pathways in lung cancer

    PubMed Central

    Jiao, Demin; Wang, Jian; Lu, Wei; Tang, Xiali; Chen, Jun; Mou, Hao; Chen, Qing-yong

    2016-01-01

    The epithelial-mesenchymal transition (EMT) and angiogenesis have emerged as two pivotal events in cancer progression. Curcumin has been extensively studied in preclinical models and clinical trials of cancer prevention due to its favorable toxicity profile. However, the possible involvement of curcumin in the EMT and angiogenesis in lung cancer remains unclear. This study found that curcumin inhibited hepatocyte growth factor (HGF)-induced migration and EMT-related morphological changes in A549 and PC-9 cells. Moreover, pretreatment with curcumin blocked HGF-induced c-Met phosphorylation and downstream activation of Akt, mTOR, and S6. These effects mimicked that of c-Met inhibitor SU11274 or PI3 kinase inhibitor LY294002 or mTOR inhibitor rapamycin treatment. c-Met gene overexpression analysis further demonstrated that curcumin suppressed lung cancer cell EMT by inhibiting c-Met/Akt/mTOR signaling pathways. In human umbilical vein endothelial cells (HUVECs), we found that curcumin also significantly inhibited PI3K/Akt/mTOR signaling and induced apoptosis and reduced migration and tube formation of HGF-treated HUVEC. Finally, in the experimental mouse model, we showed that curcumin inhibited HGF-stimulated tumor growth and induced an increase in E-cadherin expression and a decrease in vimentin, CD34, and vascular endothelial growth factor (VEGF) expression. Collectively, these findings indicated that curcumin could inhibit HGF-promoted EMT and angiogenesis by targeting c-Met and blocking PI3K/Akt/mTOR pathways. PMID:27525306

  19. Lycium barbarum Polysaccharides Protect against Trimethyltin Chloride-Induced Apoptosis via Sonic Hedgehog and PI3K/Akt Signaling Pathways in Mouse Neuro-2a Cells

    PubMed Central

    Zhao, Wanyun; Pan, Xiaoqi; Li, Tao; Zhang, Changchun; Shi, Nian

    2016-01-01

    Trimethyltin chloride (TMT) is a classic neurotoxicant that can cause severe neurodegenerative diseases. Some signaling pathways involving cell death play pivotal roles in the central nervous system. In this study, the role of Sonic Hedgehog (Shh) and PI3K/Akt pathways in TMT-induced apoptosis and protective effect of Lycium barbarum polysaccharides (LBP) on mouse neuro-2a (N2a) cells were investigated. Results showed that TMT treatment significantly enhanced apoptosis, upregulated proapoptotic Bax, downregulated antiapoptotic Bcl-2 expression, and increased caspase-3 activity in a dose-dependent manner in N2a cells. TMT induced oxidative stress in cells, performing reactive oxygen species (ROS) and malondialdehyde (MDA) excessive generation, and superoxide dismutase (SOD) activity reduction. TMT significantly decreased phosphorylated glycogen synthase kinase-3β (GSK-3β) and inhibited Shh and PI3K/Akt pathways. However, the addition of LBP upregulated GSK-3β phosphorylation, activated Shh and PI3K/Akt pathways, and eventually reduced apoptosis and oxidative stress caused by TMT. The interaction between Shh and PI3K/Akt pathways was clarified by specific PI3K inhibitor LY294002 or Shh inhibitor GDC-0449. Moreover, LY294002 and GDC-0449 pretreatment both induced phosphorylated GSK-3β downregulation and significantly promoted apoptosis induced by TMT. These results suggest that LBP could reduce TMT-induced N2a cells apoptosis by regulating GSK-3β phosphorylation, Shh, and PI3K/Akt signaling pathways. PMID:27143997

  20. Activation of transient receptor potential vanilloid 4 induces apoptosis in hippocampus through downregulating PI3K/Akt and upregulating p38 MAPK signaling pathways

    PubMed Central

    Jie, P; Hong, Z; Tian, Y; Li, Y; Lin, L; Zhou, L; Du, Y; Chen, L; Chen, L

    2015-01-01

    Transient receptor potential vanilloid 4 (TRPV4) is a calcium-permeable cation channel that is sensitive to cell swelling, arachidonic acid and its metabolites, epoxyeicosatrienoic acids, which are associated with cerebral ischemia. The activation of TRPV4 induces cytotoxicity in many types of cells, accompanied by an increase in the intracellular free calcium concentration. TRPV4 activation modulates the mitogen-activated protein kinase (MAPK) and phosphatidyl inositol 3 kinase (PI3K)/ protein kinase B (Akt) signaling pathways that regulate cell death and survival. Herein, we examined TRPV4-induced neuronal apoptosis by intracerebroventricular (ICV) injection of a TRPV4 agonist (GSK1016790A) and assessed its involvement in cerebral ischemic injury. ICV injection of GSK1016790A dose-dependently induced apoptosis in the mouse hippocampi (GSK-injected mice). The protein level of phosphorylated p38 MAPK (p-p38 MAPK) was markedly increased and that of phosphorylated c-Jun N-terminal protein kinase (p-JNK) was virtually unchanged. TRPV4 activation also decreased Bcl-2/Bax protein ratio and increased the cleaved caspase-3 protein level, and these effects were blocked by a PI3K agonist and a p38 MAPK antagonist, but were unaffected by a JNK antagonist. ICV injection of the TRPV4 antagonist HC-067047 reduced brain infarction after reperfusion for 48 h in mice with middle cerebral artery occlusion (MCAO). In addition, HC-067047 treatment attenuated the decrease in the phosphorylated Akt protein level and the increase in p-p38 MAPK protein level at 48 h after MCAO, while the increase in p-JNK protein level remained unchanged. Finally, the decreased Bcl-2/Bax protein ratio and the increased cleaved caspase-3 protein level at 48 h after MCAO were markedly attenuated by HC-067047. We conclude that activation of TRPV4 induces apoptosis by downregulating PI3K/Akt and upregulating p38 MAPK signaling pathways, which is involved in cerebral ischemic injury. PMID:26043075

  1. Involvement of adipokines, AMPK, PI3K and the PPAR signaling pathways in ovarian follicle development and cancer.

    PubMed

    Dupont, Joëlle; Reverchon, Maxime; Cloix, Lucie; Froment, Pascal; Ramé, Christelle

    2012-01-01

    The physiological mechanisms that control energy balance are reciprocally linked to those that control reproduction, and together, these mechanisms optimize reproductive success under fluctuating metabolic conditions. Adipose tissue plays an important role in this regulation. Indeed, it releases a variety of factors, termed adipokines that regulate energy metabolism, but also reproductive functions. This article summarizes the function and regulation of some better-characterized adipokines (leptin, adiponectin, resistin, visfatin, chemerin and apelin) involved in ovarian follicle development. The follicle appears to use various "nutrient sensing" mechanisms that may form the link between nutrient status and folliculogenesis. This review examines evidence for the presence of pathways that may sense nutrient flux from within the follicle including the PI3K/Akt pathway, adenosine monophosphate-activated kinase (AMPK), and peroxisome proliferator-activated receptors (PPARs). It also reviews current information on the role of these adipokines and signalling pathways in ovarian cancers. PMID:23417417

  2. Control of Cardiac Repolarization by Phosphoinositide 3-kinase Signaling to Ion Channels

    PubMed Central

    Ballou, Lisa M.; Lin, Richard Z.; Cohen, Ira S.

    2014-01-01

    Upregulation of phosphoinositide 3-kinase (PI3K) signaling is a common alteration in human cancer, and numerous drugs that target this pathway have been developed for cancer treatment. However, recent studies have implicated inhibition of the PI3K signaling pathway as the cause of a drug-induced long QT syndrome in which alterations in several ion currents contribute to arrhythmogenic drug activity. Surprisingly, some drugs that were thought to induce long QT syndrome by direct block of the rapid delayed rectifier (IKr) also appear to inhibit PI3K signaling, an effect that may contribute to their arrhythmogenicity. The importance of PI3K in regulating cardiac repolarization is underscored by evidence that QT interval prolongation in diabetes also may result from changes in multiple currents due to decreased insulin activation of PI3K in the heart. How PI3K signaling regulates ion channels to control the cardiac action potential is poorly understood. Hence, this review summarizes what is known about the impact of PI3K and its downstream effectors including Akt on sodium, potassium and calcium currents in cardiac myocytes. We also refer to some studies in non-cardiac cells that provide insight into potential mechanisms of ion channel regulation by this signaling pathway in the heart. Drug development and safety could be improved with a better understanding of the mechanisms by which PI3K regulates cardiac ion channels and the extent to which PI3K inhibition contributes to arrhythmogenic susceptibility. PMID:25552692

  3. Sat-Nav for T cells: Role of PI3K isoforms and lipid phosphatases in migration of T lymphocytes.

    PubMed

    Ward, Stephen G; Westwick, John; Harris, Stephanie

    2011-07-01

    Phosphoinositide 3-kinase (PI3K)-dependent signaling has been placed at the heart of conserved biochemical mechanisms that facilitate cell migration of leukocytes in response to a range of chemoattractant stimuli. This review assesses the evidence for and against PI3K-dependent mechanisms of T lymphocyte migration and whether pharmacological targeting of PI3K isoforms is likely to offer potential benefit for T cell mediated pathologies. PMID:21333676

  4. Multiple cytoskeletal pathways and PI3K signaling mediate CDC-42-induced neuronal protrusion in C. elegans.

    PubMed

    Alan, Jamie K; Struckhoff, Eric C; Lundquist, Erik A

    2013-01-01

    Rho GTPases are key regulators of cellular protrusion and are involved in many developmental events including axon guidance during nervous system development. Rho GTPase pathways display functional redundancy in developmental events, including axon guidance. Therefore, their roles can often be masked when using simple loss-of-function genetic approaches. As a complement to loss-of-function genetics, we constructed a constitutively activated CDC-42(G12V) expressed in C. elegans neurons. CDC-42(G12V) drove the formation of ectopic lamellipodial and filopodial protrusions in the PDE neurons, which resembled protrusions normally found on migrating growth cones of axons. We then used a candidate gene approach to identify molecules that mediate CDC-42(G12V)-induced ectopic protrusions by determining if loss of function of the genes could suppress CDC-42(G12V). Using this approach, we identified 3 cytoskeletal pathways previously implicated in axon guidance, the Arp2/3 complex, UNC-115/abLIM, and UNC-43/Ena. We also identified the Nck-interacting kinase MIG-15/NIK and p21-activated kinases (PAKs), also implicated in axon guidance. Finally, PI3K signaling was required, specifically the Rictor/mTORC2 branch but not the mTORC1 branch that has been implicated in other aspects of PI3K signaling including stress and aging. Our results indicate that multiple pathways can mediate CDC-42-induced neuronal protrusions that might be relevant to growth cone protrusions during axon pathfinding. Each of these pathways involves Rac GTPases, which might serve to integrate the pathways and coordinate the multiple CDC-42 pathways. These pathways might be relevant to developmental events such as axon pathfinding as well as disease states such as metastatic melanoma.

  5. Role of TLR4-Mediated PI3K/AKT/GSK-3β Signaling Pathway in Apoptosis of Rat Hepatocytes

    PubMed Central

    Zhang, Xian; Jiang, Daorong; Jiang, Wei; Zhao, Min; Gan, Jianhe

    2015-01-01

    We investigated the mechanism of the Toll-like receptor 4- (TLR4-) mediated PI3K/AKT/GSK-3β signaling pathway in rat hepatocytes apoptosis induced by LPS. The cultured rat hepatocytes were treated with LPS alone or first pretreated with TLR4 inhibitor, AKT inhibitor, and GSK-3β inhibitor, respectively, and then stimulated with the same dose of LPS. Cell viability, cell apoptotic rate, and apoptosis morphology were assessed; the level of P-AKTSer473, P-GSK-3βSer9, and active Caspase-3 and the ratio of Bax/Bcl-2 were evaluated. The results indicated that cell viability decreased, while cell apoptotic rate increased with time after LPS stimulation. The expression of P-AKTSer473 and P-GSK-3βSer9 in the LPS group decreased compared with the control, while the level of active Caspase-3 and the ratio of Bax/Bcl-2 were significantly increased. These effects were attenuated by pretreatment with CLI-095. In addition, the apoptotic ratio decreased after pretreatment with LiCl but increased following pretreatment with LY294002. The expression of P-AKTSer473 further decreased following pretreatment with LY294002 and the expression of P-GSK-3βSer9 increased following pretreatment with LiCl. Moreover, pretreatment with CLI-095 weakened LPS-induced nuclear translocation of GSK-3β. Our findings suggest that the TLR4-mediated PI3K/AKT/GSK-3β signaling pathway is present in rat hepatocytes and participates in apoptosis of BRL-3A cells. PMID:26770978

  6. RLIP76 Regulates PI3K/Akt Signaling and Chemo-Radiotherapy Resistance in Pancreatic Cancer

    PubMed Central

    Leake, Kathryn; Singhal, Jyotsana; Nagaprashantha, Lokesh Dalasanur; Awasthi, Sanjay; Singhal, Sharad S.

    2012-01-01

    Purpose Pancreatic cancer is an aggressive malignancy with characteristic metastatic course of disease and resistance to conventional chemo-radiotherapy. RLIP76 is a multi-functional cell membrane protein that functions as a major mercapturic acid pathway transporter as well as key regulator of receptor-ligand complexes. In this regard, we investigated the significance of targeting RLIP76 on PI3K/Akt pathway and mechanisms regulating response to chemo-radiotherapy. Research Design and Methods Cell survival was assessed by MTT and colony forming assays. Cellular levels of proteins and phosphorylation was determined by Western blot analyses. The impact on apoptosis was determined by TUNEL assay. The anti-cancer effects of RLIP76 targeted interventions in vivo were determined using mice xenograft model of the pancreatic cancer. The regulation of doxorubicin transport and radiation sensitivity were determined by transport studies and colony forming assays, respectively. Results Our current studies reveal an encompassing model for the role of RLIP76 in regulating the levels of fundamental proteins like PI3K, Akt, E-cadherin, CDK4, Bcl2 and PCNA which are of specific importance in the signal transduction from critical upstream signaling cascades that determine the proliferation, apoptosis and differentiation of pancreatic cancer cells. RLIP76 depletion also caused marked and sustained regression of established human BxPC-3 pancreatic cancer tumors in nude mouse xenograft model. RLIP76 turned out to be a major regulator of drug transport along with contributing to the radiation resistance in pancreatic cancer. Conclusions/Significance RLIP76 represents a mechanistically significant target for developing effective interventions in aggressive and refractory pancreatic cancers. PMID:22509328

  7. Silencing of VEGF inhibits human osteosarcoma angiogenesis and promotes cell apoptosis via VEGF/PI3K/AKT signaling pathway

    PubMed Central

    Peng, Ningning; Gao, Shuming; Guo, Xu; Wang, Guangya; Cheng, Cai; Li, Min; Liu, Kehun

    2016-01-01

    Background: Osteosarcoma is a kind of highly malignant tumor and the growth and metastasis is closely related to angiogenesis. Vascular endothelial growth factor (VEGF) is an important angiogenesis-promoting factor. In the current study, we investigated the effects of suppressed VEGF on osteosarcoma and its molecular mechanism provided for a basis by targeting angiogenesis. Material/Methods: We established bearing human osteosarcoma Wistar rats model by subcutaneous inoculation of human SaOS-2 cells and the adenovirus vector Ad-VEGF-siRNA was constructed for further study. We assessed the efficiency of VEGF silencing and its influence on SaOS-2 cells. The expression of mRNA and protein were detected by RT-PCR and western blotting, respectively. Intratumoral microvessel density (MVD), VEGF and CD31 were evaluated by immunohistochemistry. We detected the cell apoptotic rates by flow cytometry. Results: Our results indicated that Ad-VEGF-siRNA could effectively suppressed the expression of VEGF expression, inhibited the proliferation capability and promoted apoptosis of SaOS-2 cells in vitro. Silencing of VEGF expression also suppress osteosarcoma tumor growth and reduce osteosarcoma angiogenesis in the Wistar rats model in vivo. Furthermore, We found that phosphoinositide 3-kinase (PI3K) and protein kinase B (AKT) activation were considerably reduced while inhibition VEGF expression in SaOS-2 cells. Conclusion: Our data demonstrated that VEGF silencing could suppress cells proliferation, promote cells apoptosis and reduce osteosarcoma angiogenesis through inactivation of VEGF/PI3K/AKT signaling pathway. PMID:27158386

  8. Neuritogenic Monoglyceride Derived from the Constituent of a Marine Fish for Activating the PI3K/ERK/CREB Signalling Pathways in PC12 Cells

    PubMed Central

    Yang, Wei; Luo, Yan; Tang, Ruiqi; Zhang, Hui; Ye, Ying; Xiang, Lan; Qi, Jianhua

    2013-01-01

    A neuritogenic monoglyceride, 1-O-(myristoyl) glycerol (MG), was isolated from the head of Ilisha elongate using a PC12 cell bioassay system, and its chemical structure was elucidated using spectroscopic methods. MG significantly induced 42% of the neurite outgrowth of PC12 cells at a concentration of 10 μM. To study the structure-activity relationships of MG, a series of monoglycerides was designed and synthesised. Bioassay results indicated that the alkyl chain length plays a key role in the neuritogenic activity of the monoglycerides. The groups that link the propane-1,2-diol and alkyl chain were also investigated. An ester linkage, rather than an amido one, was found to be optimal for neuritogenic activity. Therefore, 1-O-(stearoyl) glycerol (SG), which induces 57% of the neurite outgrowth of PC12 cells at 10 μM, was determined to be a lead compound for neuritogenic activity. We then investigated the mechanism of action of neurite outgrowth induced by SG on PC12 cells using protein specific inhibitors and Western blot analysis. The mitogen-activated kinase/ERK kinase (MEK) inhibitor U0126 and the phosphatidylinositol-3 kinase (PI3K) inhibitor LY294002 significantly decreased neurite outgrowth. At the same time, SG increased phosphorylation of CREB in protein level. Thus, SG-induced neuritogenic activity depends on the activation of the extracellular-regulated protein kinase (ERK), cAMP responsive element-binding protein (CREB) and PI3K signalling pathways in PC12 cells. PMID:24351811

  9. Effects of D-Pinitol on Insulin Resistance through the PI3K/Akt Signaling Pathway in Type 2 Diabetes Mellitus Rats.

    PubMed

    Gao, Yunfeng; Zhang, Mengna; Wu, Tianchen; Xu, Mengying; Cai, Haonan; Zhang, Zesheng

    2015-07-01

    D-pinitol, a compound isolated from Pinaceae and Leguminosae plants, has been reported to possess insulin-like properties. Although the hypoglycemic activity of D-pinitol was recognized in recent years, the molecular mechanism of D-pinitol in the treatment of diabetes mellitus remains unclear. In this investigation, a model of type 2 diabetes mellitus (T2DM) with insulin resistance was established by feeding a high-fat diet (HFD) and injecting streptozocin (STZ) to Sprague-Dawley (SD) rats, targeting the exploration of more details of the mechanism in the therapy of T2DM. D-pinitol was administrated to the diabetic rats as two doses [30, 60 mg/(kg·body weight·day)]. The level of fasting blood glucose (FBG) was decreased 12.63% in the high-dosage group, and the ability of oral glucose tolerance was improved in D-pinitol-treated groups. The biochemical indices revealed that D-pinitol had a positive effect on hypoglycemic activity. Western boltting suggested that D-pinitol could promote the expression of the phosphatidylinositol-3-kinase (PI3K) p85, PI3Kp110, as well as the downstream target protein kinase B/Akt (at Ser473). Besides, D-pinitol inhibited the expression of glycogen synthesis kinase-3β (GSK-3β) protein and regulated the expression of glycogen synthesis (GS) protein and then accelerated the glycogen synthesis. Above all, D-pinitol played a positive role in regulating insulin-mediated glucose uptake in the liver through translocation and activation of the PI3K/Akt signaling pathway in T2DM rats.

  10. The Regulation of Lipid Deposition by Insulin in Goose Liver Cells Is Mediated by the PI3K-AKT-mTOR Signaling Pathway

    PubMed Central

    Han, Chunchun; Wei, Shouhai; He, Fang; Liu, Dandan; Wan, Huofu; Liu, Hehe; Li, Liang; Xu, Hongyong; Du, Xiaohui; Xu, Feng

    2015-01-01

    Background We previously showed that the fatty liver formations observed in overfed geese are accompanied by the activation of the PI3K-Akt-mTOR pathway and an increase in plasma insulin concentrations. Recent studies have suggested a crucial role for the PI3K-Akt-mTOR pathway in regulating lipid metabolism; therefore, we hypothesized that insulin affects goose hepatocellular lipid metabolism through the PI3K-Akt-mTOR signaling pathway. Methods Goose primary hepatocytes were isolated and treated with serum-free media supplemented with PI3K-Akt-mTOR pathway inhibitors (LY294002, rapamycin, and NVP-BEZ235, respectively) and 50 or 150 nmol/L insulin. Results Insulin induced strong effects on lipid accumulation as well as the mRNA and protein levels of genes involved in lipogenesis, fatty acid oxidation, and VLDL-TG assembly and secretion in primary goose hepatocytes. The stimulatory effect of insulin on lipogenesis was significantly decreased by treatment with PI3K-Akt-mTOR inhibitors. These inhibitors also rescued the insulin-induced down-regulation of fatty acid oxidation and VLDL-TG assembly and secretion. Conclusion These findings suggest that the stimulatory effect of insulin on lipid deposition is mediated by PI3K-Akt-mTOR regulation of lipogenesis, fatty acid oxidation, and VLDL-TG assembly and secretion in goose hepatocytes. PMID:25945932

  11. Puquitinib mesylate (XC-302) induces autophagy via inhibiting the PI3K/AKT/mTOR signaling pathway in nasopharyngeal cancer cells.

    PubMed

    Wang, Ke-Feng; Yang, Hang; Jiang, Wen-Qi; Li, Su; Cai, Yu-Chen

    2015-12-01

    There are numerous studies that demonstrate the anti-neoplastic activity of phosphatidylinositol 3-kinase (PI3K) inhibitors and the mechanisms of inducing autophagy in cancer cells. The new anticancer drug puquitinib mesylate (XC-302) is a molecular-targeted drug, which suppresses the activity of PI3K directly. However, it remains unclear whether XC‑302 can develop an antitumor effect by inducing autophagy in nasopharyngeal cancer cells. The MTT assay was used to study the anti-proliferative effects of XC-302. Subsequently, autophagy was determined by monodansylcadaverine (MDC) staining, punctate localization of green fluorescent protein (GFP)-light chain 3 (LC3), LC3 protein blotting and electron microscopy. The expression levels of beclin 1, p62, protein kinase B (AKT), phospho (p)‑AKT, mechanistic target of rapamycin (mTOR) and p‑mTOR in XC-302‑induced autophagy were detected. Autophagy inhibition was assayed by 3-methyladenine (3‑MA) or small interfering RNA (siRNA) silencing of beclin 1. XC-302 inhibited the viability of CNE‑2 in a dose-dependent manner and the IC50 of 72 h was 5.2 µmol/l. After cells were exposed to XC-302 for 24 h, MDC-labeled autophagolysosomes were evident in CNE-2 cells by fluorescence microscope. Autophagosomes and autolysosomes were identified by transmission electron microscopy. Following transfection with GFP‑LC3, XC-302 induced a significant accumulation of GFP‑LC3, as monitored by a confocal microscope, which was reduced by 3-MA. XC-302 induced the formation of LC3‑II, increased beclin 1 levels and decreased the expression of p62. Additionally, the expression levels of p‑AKT and p‑mTOR were reduced with the elevation of XC-302. Knockdown of beclin 1 with siRNA or co-treatment with 3-MA enhanced significantly the survival of CNE-2 and promoted the ability of clone formation. XC-302 also induced apoptosis in CNE-2, and when autophagy was inhibited by 3-MA, the apoptosis rate was decreased. The present data

  12. Aberrant Cytoplasm Localization and Protein Stability of SIRT1 is Regulated by PI3K/IGF-1R Signaling in Human Cancer Cells

    PubMed Central

    Byles, Vanessa; Chmilewski, Laura K.; Wang, Joyce; Zhu, Lijia; Forman, Lora W.; Faller, Douglas V.; Dai, Yan

    2010-01-01

    SIRT1, an NAD-dependent histone/protein deacetylase, has classically been thought of as a nuclear protein. In this study, we demonstrate that SIRT1 is mainly localized in the nucleus of normal cells, but is predominantly localized in the cytoplasm of the cancer / transformed cells we tested. We found this predominant cytoplasmic localization of SIRT1 is regulated by elevated mitotic activity and PI3K/IGF-1R signaling in cancer cells. We show that aberrant cytoplasmic localization of SIRT1 is due to increased protein stability and is regulated by PI3K/IGF-1R signaling. In addition, we determined that SIRT1 is required for PI3K-mediated cancer cell growth. Our study represents the first identification that aberrant cytoplasm localization is one of the specific alternations to SIRT1 that occur in cancer cells, and PI3K/IGF-1R signaling plays an important role in the regulation of cytoplasmic SIRT1 stability. Our findings suggest that the over-expressed cytoplasmic SIRT1 in cancer cells may greatly contribute to its cancer-specific function by working downstream of the PI3K/IGF-1R signaling pathway. PMID:20941378

  13. PI3K in cancer: divergent roles of isoforms, modes of activation, and therapeutic targeting

    PubMed Central

    Thorpe, Lauren M.; Yuzugullu, Haluk; Zhao, Jean J.

    2015-01-01

    Preface Phosphatidylinositol 3-Kinases (PI3Ks) are critical coordinators of intracellular signaling in response to extracellular stimuli. Hyperactivation of PI3K signaling cascades is one of the most common events in human cancers. In this Review, we discuss recent advances in our knowledge of the roles of distinct PI3K isoforms in normal and oncogenic signaling, the different ways in which PI3K can be upregulated, and the current state and future potential of targeting this pathway in the clinic. PMID:25533673

  14. PI3K/AKT signaling is essential for communication between tissue infiltrating mast cells, macrophages, and epithelial cells in colitis-induced cancer

    PubMed Central

    Khan, Mohammad W.; Keshavarzian, Ali; Gounaris, Elias; Melson, Joshua E.; Cheon, Eric; Blatner, Nichole R.; Chen, Zongmin E.; Tsai, Fu-Nien; Lee, Goo; Ryu, Hyunji; Barrett, Terrence A.; Bentrem, David; Beckhove, Philipp; Khazaie, Khashayarsha

    2013-01-01

    Purpose To understand signaling pathways that shape inflamed tissue and predispose to cancer is critical for effective prevention and therapy of chronic inflammatory diseases. We have explored PI3K activity in human inflammatory bowel diseases (IBD) and mouse colitis models. Experimental Design We performed immunostaining of phosphorylated AKT (pAKT) and unbiased high throughput image acquisition and quantitative analysis of samples of non-inflamed normal colon, colitis, dysplasia, and colorectal cancer (CRC). Mechanistic insights were gained from ex vivo studies of cell interactions, the Piroxicam / IL-10−/− mouse model of progressive colitis, and use of the PI3K inhibitor LY294002. Results Progressive increase in densities of pAKT-positive tumor-associated macrophages (TAMs) and increase in densities of mast cells (MCs) in the colonic submucosa were noted with colitis and progression to dysplasia and cancer. MCs recruited macrophages in ex vivo migration assays, and both MCs and TAMs promoted invasion of cancer cells. Pre-treatment of MCs with LY294002 blocked recruitment of TAMs. LY294002 inhibited MC and TAM-mediated tumor invasion, and in mice, blocked stromal PI3K, colitis, and cancer. Conclusion The PI3K / AKT pathway is active in cells infiltrating inflamed human colon tissue. This pathway sustains the recruitment of inflammatory cells through a positive feed back loop. The PI3K / AKT pathway is essential for tumor invasion and the malignant features of the Piroxicam / IL-10−/− mouse model. LY294002 targets the PI3K pathway and hinders progressive colitis. These findings indicate that colitis and progression to cancer are dependent on stromal PI3K and sensitive to treatment with LY294002. PMID:23487439

  15. Pharmacological inhibition of mTORC1 prevents over-activation of the primordial follicle pool in response to elevated PI3K signaling.

    PubMed

    Adhikari, Deepak; Risal, Sanjiv; Liu, Kui; Shen, Yan

    2013-01-01

    The majority of ovarian primordial follicles must be preserved in a quiescent state to allow for the regular production of gametes over the female reproductive lifespan. However, the molecular mechanism that maintains the long quiescence of primordial follicles is poorly understood. Under certain pathological conditions, the entire pool of primordial follicles matures simultaneously leading to an accelerated loss of primordial follicles and to premature ovarian failure (POF). We have previously shown that loss of Pten (phosphatase and tensin homolog deleted on chromosome ten) in mouse oocytes leads to premature activation of the entire pool of primordial follicles, subsequent follicular depletion in early adulthood, and the onset of POF. Lack of PTEN leads to increased phosphatidylinositol 3-kinase (PI3K)-Akt and mammalian target of rapamycin complex 1 (mTORC1) signaling in the oocytes. To study the functional and pathological roles of elevated mTORC1 signaling in the oocytes, we treated the Pten-mutant mice with the specific mTORC1 inhibitor rapamycin. When administered to Pten-deficient mice prior to the activation of the primordial follicles, rapamycin effectively prevented global follicular activation and preserved the ovarian reserve. These results provide a rationale for exploring the possible use of rapamycin as a drug for the preservation of the primordial follicle pool, and the possible prevention of POF.

  16. Alpha-chaconine-reduced metastasis involves a PI3K/Akt signaling pathway with downregulation of NF-kappaB in human lung adenocarcinoma A549 cells.

    PubMed

    Shih, Yuan-Wei; Chen, Pin-Shern; Wu, Cheng-Hsun; Jeng, Ya-Fang; Wang, Chau-Jong

    2007-12-26

    Alpha-chaconine, isolated from Solanum tuberosum Linn., is a naturally occurring steroidal glycoalkaloid in potato sprouts. Some reports demonstrated that alpha-chaconine had various anticarcinogenic properties. The aim of this study is to investigate the inhibitory effect of alpha-chaconine on lung adenocarcinoma cell metastasis in vitro. We chose the highly metastatic A549 cells, which were treated with various concentrations of alpha-chaconine to clarify the potential of inhibiting A549 cells invasion and migration. Data showed that alpha-chaconine inhibited A549 cell invasion/migration according to wound healing assay and Boyden chamber assay. Our results also showed that alpha-chaconine could inhibit phosphorylation of c-Jun N-terminal kinase (JNK) and Akt, whereas it did not affected phosphorylation of extracellular signal regulating kinase (ERK) and p38. In addition, alpha-chaconine significantly decreased the nuclear level of nuclear factor kappa B (NF-kappaB) and the binding ability of NF-kappaB. These results suggested that alpha-chaconine inhibited A549 cell metastasis by a reduction of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) activities involving suppression of phosphoinositide 3-kinase/Akt/NF-kappaB (PI3K/Akt/NF-kappaB) signaling pathway. Inhibiting metastasis by alpha-chaconine might offer a pivotal mechanism for its effective chemotherapeutic action.

  17. Curcumin Promotes Cell Cycle Arrest and Inhibits Survival of Human Renal Cancer Cells by Negative Modulation of the PI3K/AKT Signaling Pathway.

    PubMed

    Zhang, Hao; Xu, Weili; Li, Baolin; Zhang, Kai; Wu, Yudong; Xu, Haidong; Wang, Junyong; Zhang, Jun; Fan, Rui; Wei, Jinxing

    2015-12-01

    Curcumin possesses anti-cancer effects. In the current study, we tested the effect of curcumin on cell proliferation, viability, apoptosis, cell cycle phases, and activation of the PI3K/Akt pathway in the renal cell carcinoma (RCC) cell line RCC-949. We observed that cell proliferation and viability were markedly inhibited by curcumin, while cell apoptosis was promoted. The latter effect was associated with increased expression of Bcl-2 and diminished expression of Bax (both: mRNA and protein). The cells treated with curcumin increasingly went into cell cycle arrest, which was likely mediated by diminished expression of cyclin B1, as seen in curcumin-treated cells. In addition, curcumin decreased activation of the PI3K/AKT signaling pathway. In conclusion, our results demonstrate that curcumin exerts anti-cancer effects by negative modulation of the PI3K/AKT signaling pathway and may represent a promising new drug to treat RCC. PMID:27259310

  18. Opisthorchis viverrini infection activates the PI3K/ AKT/PTEN and Wnt/β-catenin signaling pathways in a Cholangiocarcinogenesis model.

    PubMed

    Yothaisong, Supak; Thanee, Malinee; Namwat, Nisana; Yongvanit, Puangrat; Boonmars, Thidarut; Puapairoj, Anucha; Loilome, Watcharin

    2014-01-01

    Opisthorchis viverrini (Ov) infection is the major etiological factor for cholangiocarcinoma (CCA), especially in northeast Thailand. We have previously reported significant involvement of PI3K/AKT/PTEN and Wnt/β- catenin in human CCA tissues. The present study, therefore, examined the expression and activation of PI3K/ AKT/PTEN and Wnt/β-catenin signaling components during Ov-induced cholangiocarcinogenesis in a hamster animal model. Hamsters were divided into two groups; non-treated and Ov plus NDMA treated. The results of immunohistochemical staining showed an upregulation of PI3K/AKT signaling as determined by elevated expression of the p85α-regulatory and p110α-catalytic subunits of PI3K as well as increased expression and activation of AKT during cholangiocarcinogenesis. Interestingly, the staining intensity of activated AKT (p-AKT) increased in the apical regions of the bile ducts and strong staining was detected where the liver fluke resides. Moreover, PTEN, a negative regulator of PI3K/AKT, was suppressed by decreased expression and increased phosphorylation during cholangiocarcinogenesis. We also detected upregulation of Wnt/β-catenin signaling as determined by increased positive staining of Wnt3, Wnt3a, Wnt5a, Wnt7b and β-catenin, corresponded with the period of cholangiocarcinogenesis. Furthermore, nuclear staining of β-catenin was observed in CCA tissues. Our results suggest the liver fluke infection causes chronic inflammatory conditions which lead to upregulation of the PI3K/AKT and Wnt/β-catenin signaling pathways which may drive CCA carcinogenesis. These results provide useful information for drug development, prevention and treatment of CCA. PMID:25556493

  19. Colon Cancer Tumorigenesis Initiated by the H1047R Mutant PI3K

    PubMed Central

    Yueh, Alexander E.; Payne, Susan N.; Leystra, Alyssa A.; Van De Hey, Dana R.; Foley, Tyler M.; Pasch, Cheri A.; Clipson, Linda; Matkowskyj, Kristina A.; Deming, Dustin A.

    2016-01-01

    The phosphoinositide 3-kinase (PI3K) signaling pathway is critical for multiple important cellular functions, and is one of the most commonly altered pathways in human cancers. We previously developed a mouse model in which colon cancers were initiated by a dominant active PI3K p110-p85 fusion protein. In that model, well-differentiated mucinous adenocarcinomas developed within the colon and initiated through a non-canonical mechanism that is not dependent on WNT signaling. To assess the potential relevance of PI3K mutations in human cancers, we sought to determine if one of the common mutations in the human disease could also initiate similar colon cancers. Mice were generated expressing the Pik3caH1047R mutation, the analog of one of three human hotspot mutations in this gene. Mice expressing a constitutively active PI3K, as a result of this mutation, develop invasive adenocarcinomas strikingly similar to invasive adenocarcinomas found in human colon cancers. These tumors form without a polypoid intermediary and also lack nuclear CTNNB1 (β-catenin), indicating a non-canonical mechanism of tumor initiation mediated by the PI3K pathway. These cancers are sensitive to dual PI3K/mTOR inhibition indicating dependence on the PI3K pathway. The tumor tissue remaining after treatment demonstrated reduction in cellular proliferation and inhibition of PI3K signaling. PMID:26863299

  20. PI3K is negatively regulated by PIK3IP1, a novel p110 interacting protein

    SciTech Connect

    Zhu, Zhenqi; He, Xin; Johnson, Carla; Stoops, John; Eaker, Amanda E.; Stoffer, David S.; Bell, Aaron; Zarnegar, Reza; DeFrances, Marie C. . E-mail: defrancesmc@upmc.edu

    2007-06-22

    Signaling initiated by Class Ia phosphatidylinositol-3-kinases (PI3Ks) is essential for cell proliferation and survival. We discovered a novel protein we call PI3K interacting protein 1 (PIK3IP1) that shares homology with the p85 regulatory PI3K subunit. Using a variety of in vitro and cell based assays, we demonstrate that PIK3IP1 directly binds to the p110 catalytic subunit and down modulates PI3K activity. Our studies suggest that PIK3IP1 is a new type of PI3K regulator.

  1. PIK3CA mutations can initiate pancreatic tumorigenesis and are targetable with PI3K inhibitors

    PubMed Central

    Payne, S N; Maher, M E; Tran, N H; Van De Hey, D R; Foley, T M; Yueh, A E; Leystra, A A; Pasch, C A; Jeffrey, J J; Clipson, L; Matkowskyj, K A; Deming, D A

    2015-01-01

    Aberrations in the phosphoinositide 3-kinase (PI3K) signaling pathway have a key role in the pathogenesis of numerous cancers by altering cell growth, metabolism, proliferation and apoptosis. Interest in targeting the PI3K signaling cascade continues, as new agents are being clinically evaluated. PIK3CA mutations result in a constitutively active PI3K and are present in a subset of pancreatic cancers. Here we examine mutant PIK3CA-mediated pancreatic tumorigenesis and the response of PIK3CA mutant pancreatic cancers to dual PI3K/mammalian target of rapamycin (mTOR) inhibition. Two murine models were generated expressing a constitutively active PI3K within the pancreas. An increase in acinar-to-ductal metaplasia and pancreatic intraepithelial neoplasms (PanINs) was identified. In one model these lesions were detected as early as 10 days of age. Invasive pancreatic ductal adenocarcinoma developed in these mice as early as 20 days of age. These cancers were highly sensitive to treatment with dual PI3K/mTOR inhibition. In the second model, PanINs and invasive cancer develop with a greater latency owing to a lesser degree of PI3K pathway activation in this murine model. In addition to PI3K pathway activation, increased ERK1/2 signaling is common in human pancreatic cancers. Phosphorylation of ERK1/2 was also investigated in these models. Phosphorylation of ERK1/2 is demonstrated in the pre-neoplastic lesions and invasive cancers. This activation of ERK1/2 is diminished with dual PI3K/mTOR inhibition. In summary, PIK3CA mutations can initiate pancreatic tumorigenesis and these cancers are particularly sensitive to dual PI3K/mTOR inhibition. Future studies of PI3K pathway inhibitors for patients with PIK3CA mutant pancreatic cancers are warranted. PMID:26436951

  2. Src kinase and Syk activation initiate PI3K signaling by a chimeric latent membrane protein 1 in Epstein-Barr virus (EBV)+ B cell lymphomas.

    PubMed

    Hatton, Olivia; Lambert, Stacie L; Krams, Sheri M; Martinez, Olivia M

    2012-01-01

    The B lymphotrophic γ-herpesvirus EBV is associated with a variety of lymphoid- and epithelial-derived malignancies, including B cell lymphomas in immunocompromised and immunosuppressed individuals. The primary oncogene of EBV, latent membrane protein 1 (LMP1), activates the PI3K/Akt pathway to induce the autocrine growth factor, IL-10, in EBV-infected B cells, but the mechanisms underlying PI3K activation remain incompletely understood. Using small molecule inhibition and siRNA strategies in human B cell lines expressing a chimeric, signaling-inducible LMP1 protein, nerve growth factor receptor (NGFR)-LMP1, we show that NGFR-LMP1 utilizes Syk to activate PI3K/Akt signaling and induce IL-10 production. NGFR-LMP1 signaling induces phosphorylation of BLNK, a marker of Syk activation. Whereas Src kinases are often required for Syk activation, we show here that PI3K/Akt activation and autocrine IL-10 production by NGFR-LMP1 involves the Src family kinase Fyn. Finally, we demonstrate that NGFR-LMP1 induces phosphorylation of c-Cbl in a Syk- and Fyn-dependent fashion. Our results indicate that the EBV protein LMP1, which lacks the canonical ITAM required for Syk activation, can nevertheless activate Syk, and the Src kinase Fyn, resulting in downstream c-Cbl and PI3K/Akt activation. Fyn, Syk, and PI3K/Akt antagonists thus may present potential new therapeutic strategies that target the oncogene LMP1 for treatment of EBV+ B cell lymphomas.

  3. Id-1 promotes osteosarcoma cell growth and inhibits cell apoptosis via PI3K/AKT signaling pathway.

    PubMed

    Hao, Liang; Liao, Qi; Tang, Qiang; Deng, Huan; Chen, Lu

    2016-02-12

    Accumulating evidence reveals that Id-1 is upregulated and functions as a potential tumor promoter in several human cancer types. However, the role of Id-1 in osteosarcoma (OS) is unknown. In present study, we found that Id-1 expression was elevated in OS tissues than adjacent normal bone tissues. More importantly, we demonstrated that overexpression of Id-1 is significantly correlated with tumor progression and poor survival in OS patients. Furthermore, increased expression of Id-1 was observed in OS cell lines and ectopic expression of Id-1 significantly enhanced in vitro cell proliferation and promoted in vivo tumor growth, whereas knockdown of Id-1 suppressed OS cells growth. Moreover, our experimental data revealed that Id-1 promotes cell proliferation by facilitating cell cycle progression and inhibits cell apoptosis. Mechanistically, the effects of Id-1 in OS cells is at least partly through activation of PI3K/Akt signaling pathway. Therefore, we identified a tumorigenic role of Id-1 in OS and suggested a potential therapeutic target for OS patients.

  4. Involvement of the PI3K/Akt signal pathway in the hypoglycemic effects of tea polysaccharides on diabetic mice.

    PubMed

    Li, Shuqin; Chen, Haixia; Wang, Jia; Wang, Xiuming; Hu, Bo; Lv, Funing

    2015-11-01

    In order to investigate the hypoglycemic effects and potential mechanism of tea polysaccharides (TPS) on diabetes, type 2 diabetes mellitus (T2DM) mice were established and treated with TPS. Results showed that TPS treatment could result in the increase of body weight and the decrease of blood glucose in the diabetic mice. The contents of total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-c) were decreased significantly (p<0.05) while the contents of triglyceride (TG) and (high-density lipoprotein cholesterol) HDL-c were almost restored to the normal levels. TPS possessed the efficacy of insulin resistance alleviation and exerted obvious cytoprotective action. The enzyme activities of superoxide dismutase (SOD) and glutathione peroxidase (GPX) were notably improved in both liver and kidney tissues (p<0.05) after the treatment of TPS. Furthermore, the expressions of PI3Kp85/p-Akt/GLUT4 were upregulated by TPS, which indicated the involvement of the PI3K/Akt signal pathway in the hypoglycemic mechanism of TPS. Results suggested that TPS could ameliorate the T2MD and might be a promising candidate functional food or medicine for T2DM treatment.

  5. α-Lipoic acid inhibits sevoflurane-induced neuronal apoptosis through PI3K/Akt signalling pathway.

    PubMed

    Ma, Rong; Wang, Xiang; Peng, Peipei; Xiong, Jingwei; Dong, Hongquan; Wang, Lixia; Ding, Zhengnian

    2016-01-01

    Sevoflurane is a widely used anaesthetic agent, including in anaesthesia of children and infants. Recent studies indicated that the general anaesthesia might cause the cell apoptosis in the brain. This issue raises the concerns about the neuronal toxicity induced by the application of anaesthetic agents, especially in the infants and young children. In this study, we used Morris water maze, western blotting and immunohistochemistry to elucidate the role of α-lipoic acid in the inhibition of neuronal apoptosis. We found that sevoflurane led to the long-term cognitive impairment in the young rats. This adverse effect may be caused by the neuronal death in the hippocampal region, mediated through PI3K/Akt signalling pathway. We also showed that α-lipoic acid offset the effect of sevoflurane on the neuronal apoptosis and cognitive dysfunction. This study elucidated the potential clinical role of α-lipoic acid, providing a promising way in the prevention and treatment of long-term cognitive impairment induced by sevoflurane general anesthesia.

  6. E-Cadherin-Dependent Stimulation of Traction Force at Focal Adhesions via the Src and PI3K Signaling Pathways

    PubMed Central

    Jasaitis, Audrius; Estevez, Maruxa; Heysch, Julie; Ladoux, Benoit; Dufour, Sylvie

    2012-01-01

    The interplay between cadherin- and integrin-dependent signals controls cell behavior, but the precise mechanisms that regulate the strength of adhesion to the extracellular matrix remains poorly understood. We deposited cells expressing a defined repertoire of cadherins and integrins on fibronectin (FN)-coated polyacrylamide gels (FN-PAG) and on FN-coated pillars used as a micro-force sensor array (μFSA), and analyzed the functional relationship between these adhesion receptors to determine how it regulates cell traction force. We found that cadherin-mediated adhesion stimulated cell spreading on FN-PAG, and this was modulated by the substrate stiffness. We compared S180 cells with cells stably expressing different cadherins on μFSA and found that traction forces were stronger in cells expressing cadherins than in parental cells. E-cadherin-mediated contact and mechanical coupling between cells are required for this increase in cell-FN traction force, which was not observed in isolated cells, and required Src and PI3K activities. Traction forces were stronger in cells expressing type I cadherins than in cells expressing type II cadherins, which correlates with our previous observation of a higher intercellular adhesion strength developed by type I compared with type II cadherins. Our results reveal one of the mechanisms whereby molecular cross talk between cadherins and integrins upregulates traction forces at cell-FN adhesion sites, and thus provide additional insight into the molecular control of cell behavior. PMID:22853894

  7. Activation of Robo1 signaling of breast cancer cells by Slit2 from stromal fibroblast restrains tumorigenesis via blocking PI3K/Akt/β-catenin pathway.

    PubMed

    Chang, Po-Hao; Hwang-Verslues, Wendy W; Chang, Yi-Cheng; Chen, Chun-Chin; Hsiao, Michael; Jeng, Yung-Ming; Chang, King-Jen; Lee, Eva Y-H P; Shew, Jin-Yuh; Lee, Wen-Hwa

    2012-09-15

    Tumor microenvironment plays a critical role in regulating tumor progression by secreting factors that mediate cancer cell growth. Stromal fibroblasts can promote tumor growth through paracrine factors; however, restraint of malignant carcinoma progression by the microenvironment also has been observed. The mechanisms that underlie this paradox remain unknown. Here, we report that the tumorigenic potential of breast cancer cells is determined by an interaction between the Robo1 receptor and its ligand Slit2, which is secreted by stromal fibroblasts. The presence of an active Slit2/Robo1 signal blocks the translocation of β-catenin into nucleus, leading to downregulation of c-myc and cyclin D1 via the phosphoinositide 3-kinase (PI3K)/Akt pathway. Clinically, high Robo1 expression in the breast cancer cells correlates with increased survival in patients with breast cancer, and low Slit2 expression in the stromal fibroblasts is associated with lymph node metastasis. Together, our findings explain how a specific tumor microenvironment can restrain a given type of cancer cell from progression and show that both stromal fibroblasts and tumor cell heterogeneity affect breast cancer outcomes.

  8. miR-16 targets fibroblast growth factor 2 to inhibit NPC cell proliferation and invasion via PI3K/AKT and MAPK signaling pathways

    PubMed Central

    Li, Yingqin; Tang, Xinran; Wen, Xin; Yang, Xiaojing; Zhang, Jian; Wang, Yaqin; Ma, Jun; Liu, Na

    2016-01-01

    Dysregulation of miRNAs has been shown to contribute to the carcinogenesis and progression of nasopharyngeal carcinoma (NPC). Our previous microarray data showed that miR-16 expression is significantly decreased in archived NPC tissues. Here, we confirmed that miR-16 was reduced in NPC cell lines and freshly-frozen samples. Ectopic expression of miR-16 suppressed NPC cell proliferation, migration, and invasion in vitro and inhibited tumor growth and metastatic colonization in the lung in vivo. Furthermore, fibroblast growth factor 2 (FGF2) was identified as a direct target of miR-16, and both phosphoinositide-3- kinase/AKT (PI3K/AKT) and mitogen-activated protein kinase (MAPK) signaling pathways were repressed after miR-16 overexpression. In addition, the restoration of FGF2 reversed the suppressive effects of miR-16. Together, these results indicated that miR-16 suppresses NPC carcinogenesis and progression by targeting FGF2, thereby representing a potential target for miRNA-based therapy for NPC in the future. PMID:26655091

  9. PI3K-C2α: One enzyme for two products coupling vesicle trafficking and signal transduction.

    PubMed

    Campa, Carlo C; Franco, Irene; Hirsch, Emilio

    2015-06-22

    The spatial restriction of phosphorylated phosphoinositides generated downstream activated membrane receptors is critical for proper cell response to environmental cues. The α isoform of class II PI3Ks, PI3K-C2α, has emerged as a modulator of receptor localization, acting both in the control of receptor endocytosis and resensitization. This unexpectedly versatile enzyme was found to differentially produce two distinct 3-phosphorylated phosphoinositides and to selectively control distinct steps of vesicular traffic such as endocytosis and recycling. This review focuses on the latest discoveries regarding PI3K-C2α function in vesicle trafficking and its impact on cell biology and mammalian embryonic development.

  10. Involvement of PI3K/Akt/FoxO3a and PKA/CREB Signaling Pathways in the Protective Effect of Fluoxetine Against Corticosterone-Induced Cytotoxicity in PC12 Cells.

    PubMed

    Zeng, Bingqing; Li, Yiwen; Niu, Bo; Wang, Xinyi; Cheng, Yufang; Zhou, Zhongzhen; You, Tingting; Liu, Yonggang; Wang, Haitao; Xu, Jiangping

    2016-08-01

    The selective serotonin reuptake inhibitor fluoxetine is neuroprotective in several brain injury models. It is commonly used to treat major depressive disorder and related conditions, but its mechanism of action remains incompletely understood. Activation of the phosphatidylinositol-3-kinase/protein kinase B/forkhead box O3a (PI3K/Akt/FoxO3a) and protein kinase A/cAMP-response element binding protein (PKA/CREB) signaling pathways has been strongly implicated in the pathogenesis of depression and might be the downstream target of fluoxetine. Here, we used PC12 cells exposed to corticosterone (CORT) to study the neuroprotective effects of fluoxetine and the involvement of the PI3K/Akt/FoxO3a and PKA/CREB signaling pathways. Our results show that CORT reduced PC12 cells viability by 70 %, and that fluoxetine showed a concentration-dependent neuroprotective effect. Neuroprotective effects of fluoxetine were abolished by inhibition of PI3K, Akt, and PKA using LY294002, KRX-0401, and H89, respectively. Treatment of PC12 cells with fluoxetine resulted in increased phosphorylation of Akt, FoxO3a, and CREB. Fluoxetine also dose-dependently rescued the phosphorylation levels of Akt, FoxO3a, and CREB, following administration of CORT (from 99 to 110, 56 to 170, 80 to 170 %, respectively). In addition, inhibition of PKA and PI3K/Akt resulted in decreased levels of p-CREB, p-Akt, and p-FoxO3a in the presence of fluoxetine. Furthermore, fluoxetine reversed CORT-induced upregulation of p53-upregulated modulator of apoptosis (Puma) and Bcl-2-interacting mediator of cell death (Bim) via the PI3K/Akt/FoxO3a signaling pathway. H89 treatment reversed the effect of fluoxetine on the mRNA level of brain-derived neurotrophic factor, which was decreased in the presence of CORT. Our data indicate that fluoxetine elicited neuroprotection toward CORT-induced cell death that involves dual regulation from PI3K/Akt/FoxO3a and PKA/CREB pathways. PMID:27412469

  11. Regulation of H-Ras-driven MAPK signaling, transformation and tumorigenesis, but not PI3K signaling and tumor progression, by plasma membrane microdomains.

    PubMed

    Michael, J V; Wurtzel, J G T; Goldfinger, L E

    2016-05-30

    In this study, we assessed the contributions of plasma membrane (PM) microdomain targeting to the functions of H-Ras and R-Ras. These paralogs have identical effector-binding regions, but variant C-terminal targeting domains (tDs) which are responsible for lateral microdomain distribution: activated H-Ras targets to lipid ordered/disordered (Lo/Ld) domain borders, and R-Ras to Lo domains (rafts). We hypothesized that PM distribution regulates Ras-effector interactions and downstream signaling. We used tD swap mutants, and assessed effects on signal transduction, cell proliferation, transformation and tumorigenesis. R-Ras harboring the H-Ras tD (R-Ras-tH) interacted with Raf, and induced Raf and ERK phosphorylation similar to H-Ras. R-Ras-tH stimulated proliferation and transformation in vitro, and these effects were blocked by both MEK and PI3K inhibition. Conversely, the R-Ras tD suppressed H-Ras-mediated Raf activation and ERK phosphorylation, proliferation and transformation. Thus, Ras access to Raf at the PM is sufficient for MAPK activation and is a principal component of Ras mitogenesis and transformation. Fusion of the R-Ras extended N-terminal domain to H-Ras had no effect on proliferation, but inhibited transformation and tumor progression, indicating that the R-Ras N-terminus also contributes negative regulation to these Ras functions. PI3K activation was tD independent; however, H-Ras was a stronger activator of PI3K than R-Ras, with either tD. PI3K inhibition nearly ablated transformation by R-Ras-tH, H-Ras and H-Ras-tR, whereas MEK inhibition had a modest effect on Ras-tH-driven transformation but no effect on H-Ras-tR transformation. R-Ras-tH supported tumor initiation, but not tumor progression. While H-Ras-tR-induced transformation was reduced relative to H-Ras, tumor progression was robust and similar to H-Ras. H-Ras tumor growth was moderately suppressed by MEK inhibition, which had no effect on H-Ras-tR tumor growth. In contrast, PI3K inhibition

  12. Anti-Fibrotic Actions of Interleukin-10 against Hypertrophic Scarring by Activation of PI3K/AKT and STAT3 Signaling Pathways in Scar-Forming Fibroblasts

    PubMed Central

    Cai, Weixia; Bai, Xiaozhi; Fang, Xiaobing; Hu, Xiaolong; Wang, Yaojun; Wang, Hongtao; Zheng, Zhao; Su, Linlin; Hu, Dahai; Zhu, Xiongxiang

    2014-01-01

    Background The hypertrophic scar (HS) is a serious fibrotic skin condition and a major clinical problem. Interleukin-10 (IL-10) has been identified as a prospective scar-improving compound based on preclinical trials. Our previous work showed that IL-10 has anti-fibrotic effects in transforming growth factor (TGF)-β1-stimulated fibroblasts, as well as potential therapeutic benefits for the prevention and reduction of scar formation. However, relatively little is known about the mechanisms underlying IL-10-mediated anti-fibrotic and scar-improvement actions. Objective To explore the expression of the IL-10 receptor in human HS tissue and primary HS fibroblasts (HSFs), and the molecular mechanisms contributing to the anti-fibrotic and scar-improvement capabilities of IL-10. Methods Expression of the IL-10 receptor was assessed in HS tissue and HSFs by immunohistochemistry, immunofluorescence microscopy, and polymerase chain reaction analysis. Primary HSFs were treated with IL-10, a specific phosphatidylinositol 3 kinase (PI3K) inhibitor (LY294002) or a function-blocking antibody against the IL-10 receptor (IL-10RB). Next, Western blot analysis was used to evaluate changes in the phosphorylation status of AKT and signal transducers and activators of transcription (STAT) 3, as well as the expression levels of fibrosis-related proteins. Results HS tissue and primary HSFs were characterized by expression of the IL-10 receptor and by high expression of fibrotic markers relative to normal controls. Primary HSFs expressed the IL-10 receptor, while IL-10 induced AKT and STAT3 phosphorylation in these cells. In addition, LY294002 blocked AKT and STAT phosphorylation, and also up-regulated expression levels of type I and type III collagen (Col 1 and Col 3) and alpha-smooth muscle actin (α-SMA) in IL-10-treated cells. Similarly, IL-10RB reduced STAT3/AKT phosphorylation and blocked the IL-10-mediated mitigation of fibrosis in HSFs. Conclusion IL-10 apparently inhibits

  13. Estrogen receptor beta growth-inhibitory effects are repressed through activation of MAPK and PI3K signalling in mammary epithelial and breast cancer cells.

    PubMed

    Cotrim, C Z; Fabris, V; Doria, M L; Lindberg, K; Gustafsson, J-Å; Amado, F; Lanari, C; Helguero, L A

    2013-05-01

    Two thirds of breast cancers express estrogen receptors (ER). ER alpha (ERα) mediates breast cancer cell proliferation, and expression of ERα is the standard choice to indicate adjuvant endocrine therapy. ERbeta (ERβ) inhibits growth in vitro; its effects in vivo have been incompletely investigated and its role in breast cancer and potential as alternative target in endocrine therapy needs further study. In this work, mammary epithelial (EpH4 and HC11) and breast cancer (MC4-L2) cells with endogenous ERα and ERβ expression and T47-D human breast cancer cells with recombinant ERβ (T47-DERβ) were used to explore effects exerted in vitro and in vivo by the ERβ agonists 2,3-bis (4-hydroxy-phenyl)-propionitrile (DPN) and 7-bromo-2-(4-hydroxyphenyl)-1,3-benzoxazol-5-ol (WAY). In vivo, ERβ agonists induced mammary gland hyperplasia and MC4-L2 tumour growth to a similar extent as the ERα agonist 4,4',4''-(4-propyl-(1H)-pyrazole-1,3,5-triyl) trisphenol (PPT) or 17β-estradiol (E2) and correlated with higher number of mitotic and lower number of apoptotic features. In vitro, in MC4-L2, EpH4 or HC11 cells incubated under basal conditions, ERβ agonists induced apoptosis measured as upregulation of p53 and apoptosis-inducible factor protein levels and increased caspase 3 activity, whereas PPT and E2 stimulated proliferation. However, when extracellular signal-regulated kinase 1 and 2 (ERK ½) were activated by co-incubation with basement membrane extract or epidermal growth factor, induction of apoptosis by ERβ agonists was repressed and DPN induced proliferation in a similar way as E2 or PPT. In a context of active ERK ½, phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/RAC-alpha serine/threonine-protein kinase (AKT) signalling was necessary to allow proliferation stimulated by ER agonists. Inhibition of MEK ½ with UO126 completely restored ERβ growth-inhibitory effects, whereas inhibition of PI3K by LY294002 inhibited ERβ-induced proliferation. These

  14. Suppressing the formation of lipid raft-associated Rac1/PI3K/Akt signaling complexes by curcumin inhibits SDF-1α-induced invasion of human esophageal carcinoma cells.

    PubMed

    Lin, Meng-Liang; Lu, Yao-Cheng; Chen, Hung-Yi; Lee, Chuan-Chun; Chung, Jing-Gung; Chen, Shih-Shun

    2014-05-01

    Stromal cell-derived factor-1α (SDF-1α) is a ligand for C-X-C chemokine receptor type 4 (CXCR4), which contributes to the metastasis of cancer cells by promoting cell migration. Here, we show that the SDF-1α/CXCR4 axis can significantly increase invasion of esophageal carcinoma (EC) cells. We accomplished this by examining the effects of CXCR4 knockdown as well as treatment with a CXCR4-neutralizing antibody and the CXCR4-specific inhibitor AMD3100. Curcumin suppressed SDF-1α-induced cell invasion and matrix metalloproteinase-2 (MMP-2) promoter activity, cell surface localization of CXCR4 at lipid rafts, and lipid raft-associated ras-related C3 botulinum toxin substrate 1 (Rac1)/phosphatidylinositol 3-kinase (PI3K) p85α/Akt signaling. Curcumin inhibited SDF-1α-induced cell invasion by suppressing the Rac1-PI3K signaling complex at lipid rafts but did not abrogate lipid raft formation. We further demonstrate that the attenuation of lipid raft-associated Rac1 activity by curcumin was critical for the inhibition of SDF-1α-induced PI3K/Akt/NF-κB activation, cell surface localization of CXCR4 at lipid rafts, MMP-2 promoter activity, and cell invasion. Collectively, our results indicate that curcumin inhibits SDF-1α-induced EC cell invasion by suppressing the formation of the lipid raft-associated Rac1-PI3K-Akt signaling complex, the localization of CXCR4 with lipid rafts at the cell surface, and MMP-2 promoter activity, likely through the inhibition of Rac1 activity.

  15. Anti-tumor effects of progesterone in human glioblastoma multiforme: role of PI3K/Akt/mTOR signaling.

    PubMed

    Atif, Fahim; Yousuf, Seema; Stein, Donald G

    2015-02-01

    Glioblastoma multiforme (GBM) is an aggressive primary brain tumor with a mean patient survival of 13-15 months despite surgical resection, radiation therapy and standard-of-care chemotherapy. We investigated the chemotherapeutic effects of the hormone progesterone (P4) on the growth of human GBM in four genetically different cell lines (U87MG, U87dEGFR, U118MG, LN-229) in vitro and in a U87MG subcutaneous xenograft mouse model. At high concentrations (20, 40, and 80 μM), P4 significantly (P<0.05) decreased tumor cell viability in all cell lines except LN-229. This effect was not blocked by the P4 receptor antagonist RU468. Conversely, at low physiological concentrations (0.1, 1, and 5 μM) P4 showed a proliferative effect in all cell lines which was blocked by RU486. In nude mice, P4 (100 and 200 mg/kg) inhibited tumor growth significantly (P<0.05) over 5 weeks of treatment and extended survival time of tumor-bearing mice by 60% without signs of systemic toxicity. P4 suppressed tumor vascularization as indicated by the expression of CD31, vascular endothelial growth factor and matrix metalloproteinase-9. Apoptosis in tumor tissue was detected by the expression of cleaved caspase-3, BCl-2, BAD and p53 proteins and confirmed by TUNEL assay. P4 treatment also suppressed PI3K/Akt/mTOR signaling, which regulates tumor growth, as demonstrated by the suppression of proliferating cell nuclear antigen. Our data can be interpreted to suggest that P4 suppresses the growth of human GBM cells both in vitro and in vivo and enhances survival time in mice without any demonstrable side effects. This article is part of a Special Issue entitled 'Sex steroids and brain disorders'.

  16. Hepatocyte growth factor (HGF) upregulates heparanase expression via the PI3K/Akt/NF-κB signaling pathway for gastric cancer metastasis.

    PubMed

    Hao, Ning-Bo; Tang, Bo; Wang, Guo-Zheng; Xie, Rui; Hu, Chang-Jiang; Wang, Su-Min; Wu, Yu-Yun; Liu, En; Xie, Xia; Yang, Shi-Ming

    2015-05-28

    Heparanase (HPA) is an endoglucuronidase that can promote the shedding of associated cytokines in several types of tumors. However, little is known about what controls the expression of HPA or its role in gastric cancer. In this study, we report for the first time that HGF regulates HPA expression to promote gastric cancer metastasis. In this study, HGF and HPA were found to be significantly expressed in 58 gastric cancer patients. High expression of both HGF and HPA was positively associated with TNM stage, invasion depth and poor prognosis. In MKN74 cells, exogenous HGF significantly increased HPA expression at both the mRNA and protein levels. Further study revealed that HGF first activated PI3K/Akt signaling. NF-κB signaling was activated downstream of PI3K/Akt and promoted HPA expression. However, when c-met, PI3K/Akt or NF-κB signal inhibitors were used, HPA expression was significantly decreased. All of these results indicate that HGF regulates HPA expression by PI3K/Akt and downstream NF-κB signaling. Using bioinformatics and the ChIP assay, p65 was observed to bind to the HPA promoter. Furthermore, HGF significantly induced tumor cell migration, whereas treatment with an NF-κB inhibitor decreased migration. Moreover, when HPA was overexpressed in MKN74 cells, migration was significantly enhanced, and the HGF concentration was increased. However, when HPA was down-regulated in MKN45 cells, migration and HGF levels decreased. Together, these results demonstrate that HGF/c-met can activate PI3K/Akt and downstream NF-κB signaling to promote HPA expression and subsequent tumor metastasis.

  17. PI3K-GLUT4 Signal Pathway Associated with Effects of EX-B3 Electroacupuncture on Hyperglycemia and Insulin Resistance of T2DM Rats

    PubMed Central

    2016-01-01

    Objectives. To explore electroacupuncture's (EA's) effects on fasting blood glucose (FBG) and insulin resistance of type 2 diabetic mellitus (T2DM) model rats and give a possible explanation for the effects. Method. It takes high fat diet and intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) for model preparation. Model rats were randomly divided into T2DM Model group, EA weiwanxiashu (EX-B3) group, and sham EA group (n = 12/group). EA (2 Hz continuous wave, 2 mA, 20 min/day, 6 days/week, 4 weeks) was applied as intervention. FBG, area under curve (AUC) of oral glucose tolerance test (OGTT), insulin resistance index (HOMA-IR), pancreatic B cell function index (HOMA-B), skeletal muscle phosphorylated phosphatidylinositol-3-kinase (PI3K), glucose transporter 4 (GLUT4), and membrane GLUT4 protein expression were measured. Results. EA weiwanxiashu (EX-B3) can greatly upregulate model rat's significantly reduced skeletal muscle PI3K (Y607) and membrane GLUT4 protein expression (P < 0.01), effectively reducing model rats' FBG and AUC of OGTT (P < 0.01). The effects are far superior to sham EA group. Conclusion. EA weiwanxiashu (EX-B3) can upregulate skeletal muscle phosphorylated PI3K protein expression, to stimulate membrane translocation of GLUT4 and thereby increase skeletal muscle glucose intake to treat T2DM.

  18. PI3K-GLUT4 Signal Pathway Associated with Effects of EX-B3 Electroacupuncture on Hyperglycemia and Insulin Resistance of T2DM Rats.

    PubMed

    Cao, Bing-Yan; Li, Rui; Tian, Huan-Huan; Ma, Yan-Jia; Hu, Xiao-Gang; Jia, Ning; Wang, Yue-Ying

    2016-01-01

    Objectives. To explore electroacupuncture's (EA's) effects on fasting blood glucose (FBG) and insulin resistance of type 2 diabetic mellitus (T2DM) model rats and give a possible explanation for the effects. Method. It takes high fat diet and intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) for model preparation. Model rats were randomly divided into T2DM Model group, EA weiwanxiashu (EX-B3) group, and sham EA group (n = 12/group). EA (2 Hz continuous wave, 2 mA, 20 min/day, 6 days/week, 4 weeks) was applied as intervention. FBG, area under curve (AUC) of oral glucose tolerance test (OGTT), insulin resistance index (HOMA-IR), pancreatic B cell function index (HOMA-B), skeletal muscle phosphorylated phosphatidylinositol-3-kinase (PI3K), glucose transporter 4 (GLUT4), and membrane GLUT4 protein expression were measured. Results. EA weiwanxiashu (EX-B3) can greatly upregulate model rat's significantly reduced skeletal muscle PI3K (Y607) and membrane GLUT4 protein expression (P < 0.01), effectively reducing model rats' FBG and AUC of OGTT (P < 0.01). The effects are far superior to sham EA group. Conclusion. EA weiwanxiashu (EX-B3) can upregulate skeletal muscle phosphorylated PI3K protein expression, to stimulate membrane translocation of GLUT4 and thereby increase skeletal muscle glucose intake to treat T2DM. PMID:27656242

  19. PI3K-GLUT4 Signal Pathway Associated with Effects of EX-B3 Electroacupuncture on Hyperglycemia and Insulin Resistance of T2DM Rats

    PubMed Central

    2016-01-01

    Objectives. To explore electroacupuncture's (EA's) effects on fasting blood glucose (FBG) and insulin resistance of type 2 diabetic mellitus (T2DM) model rats and give a possible explanation for the effects. Method. It takes high fat diet and intraperitoneal injection of streptozotocin (STZ, 30 mg/kg) for model preparation. Model rats were randomly divided into T2DM Model group, EA weiwanxiashu (EX-B3) group, and sham EA group (n = 12/group). EA (2 Hz continuous wave, 2 mA, 20 min/day, 6 days/week, 4 weeks) was applied as intervention. FBG, area under curve (AUC) of oral glucose tolerance test (OGTT), insulin resistance index (HOMA-IR), pancreatic B cell function index (HOMA-B), skeletal muscle phosphorylated phosphatidylinositol-3-kinase (PI3K), glucose transporter 4 (GLUT4), and membrane GLUT4 protein expression were measured. Results. EA weiwanxiashu (EX-B3) can greatly upregulate model rat's significantly reduced skeletal muscle PI3K (Y607) and membrane GLUT4 protein expression (P < 0.01), effectively reducing model rats' FBG and AUC of OGTT (P < 0.01). The effects are far superior to sham EA group. Conclusion. EA weiwanxiashu (EX-B3) can upregulate skeletal muscle phosphorylated PI3K protein expression, to stimulate membrane translocation of GLUT4 and thereby increase skeletal muscle glucose intake to treat T2DM. PMID:27656242

  20. TGF-β1 induces human aortic vascular smooth muscle cell phenotype switch through PI3K/AKT/ID2 signaling

    PubMed Central

    Zhu, Shui-Bo; Zhu, Jian; Zhou, Zi-Zi; Xi, Er-Ping; Wang, Rong-Ping; Zhang, Yu

    2015-01-01

    The vascular smooth muscle cell (VSMC) phenotypic switch is considered to be the key pathophysiological change in various cardiovascular diseases, such as aortic dissection, atherosclerosis, and hypertension. The results in this study showed that TGF-β1 promotes the proliferation, migration and morphological changes of VSMC.TGF-β1 promoted the expressions of PI3K, P-PI3K, AKT, P-AKT, ID2, and OPN protein and suppressed the expressions of α-SMA and SM22α protein; the opposite results were observed for TGF-β1 inhibitor group, AKT inhibitor group and Combined inhibitors group. After the stimulation of TGF-β1 signaling, the mRNA levels of PI3K, AKT, ID2, and OPN were the highest, while the mRNA levels of α-SMA and SM22α were the lowest; the opposite results were found in the same groups above. These results suggested the PI3K/AKT/ID2 signaling pathway is involved in TGF-β1-mediated human aortic VSMC phenotypic switching, that is from a contractile to synthetic phenotype, and Combined inhibitors was more effective in inhibiting the phenotypic switch than a single inhibitor. The Combined inhibitors experiments may provide new avenues for the prevention and treatment of thoracic aortic dissection (TAD) that are based on the pathological effects of phenotypic switching. PMID:26885273

  1. GD2 ganglioside specific antibody treatment downregulates PI3K/Akt/mTOR signaling network in human neuroblastoma cell lines.

    PubMed

    Durbas, Małgorzata; Horwacik, Irena; Boratyn, Elżbieta; Kamycka, Elżbieta; Rokita, Hanna

    2015-09-01

    Mechanisms leading to inhibitory effects of an anti-GD2 ganglioside (GD2) 14G2a mouse monoclonal antibody (mAb) and PI3K/Akt/mTOR pathway inhibitors on human neuroblastoma cell survival were studied in vitro. We have recently shown on IMR-32, CHP‑134, and LA-N-1 neuroblastoma cells that targeting GD2 with the mAb decreases cell viability of the cell lines. In this study we used cytotoxicity assays, proteomic arrays and immunoblotting to evaluate the response of the three cell lines to the anti‑GD2 14G2a mAb and specific PI3K/Akt/mTOR pathway inhibitors. We show here that the mAb modulates intracellular signal transduction through changes in several kinases and their substrates phosphorylation. More detailed analysis of the PI3K/Akt/mTOR pathway showed significant decrease in activity of Akt, mTOR, p70 S6 and 4E-BP1 proteins and transient increase in PTEN (a suppressor of the pathway), leading to inhibition of the signaling network responsible for stimulation of translation and proliferation. Additionally, combining the GD2-specific 14G2a mAb with an Akt inhibitor (perifosine), dual mTOR/PI3K inhibitors (BEZ-235 and SAR245409), and a pan-PI3K inhibitor (LY294002) was shown to enhance cytotoxic effects against IMR-32, CHP‑134 and LA-N-1 cells. Our study extends knowledge on mechanisms of action of the 14G2a mAb on the neuroblastoma cells. Also, it stresses the need for further delineation of molecular signal orchestration aimed at more reasonable selection of drugs to target key cellular pathways in quest for better cure for neuroblastoma patients. PMID:26134970

  2. Basic fibroblast growth factor promotes melanocyte migration via activating PI3K/Akt-Rac1-FAK-JNK and ERK signaling pathways.

    PubMed

    Shi, Hongxue; Lin, Beibei; Huang, Yan; Wu, Jiang; Zhang, Hongyu; Lin, Cai; Wang, Zhouguang; Zhu, Jingjing; Zhao, Yingzhen; Fu, Xiaobing; Lou, Zhencai; Li, Xiaokun; Xiao, Jian

    2016-09-01

    Vitiligo is a depigmentation disorder characterized by loss of functional melanocytes of the skin epidermis. The pathogenesis of vitiligo remains elusive. The purpose of this study is to investigate the effects of basic fibroblast growth factor (bFGF) on melanocyte migration, including its biochemical mechanism using transwell assay in vitro. We found that melanocyte treated with bFGF showed a significant increase in migration and cytoskeletal rearrangement. These changes were associated with increased activation of PI3K/Akt, Rac1, FAK, JNK, and ERK. Likewise, reduction of PI3K/Akt, Rac1, FAK, JNK, and ERK activity using selective inhibitors or siRNA was associated with impediment of bFGF-induced melanocyte migration. In addition, activity of Rac1, FAK, and JNK was reduced in cells in which PI3K/Akt was inhibited, activity of FAK and JNK was reduced in cells in which the Rac1 was inhibited, and activity of JNK was reduced in cells in which the FAK was inhibited. Collectively, these data demonstrate that bFGF facilitated melanocyte migration via PI3K/Akt-Rac1-FAK-JNK and ERK signaling pathways. © 2016 IUBMB Life, 68(9):735-747, 2016. PMID:27350596

  3. The pan-PI3K inhibitor GDC-0941 activates canonical WNT signaling to confer resistance in TNBC cells: resistance reversal with WNT inhibitor.

    PubMed

    Tzeng, Huey-En; Yang, Lixin; Chen, Kemin; Wang, Yafan; Liu, Yun-Ru; Pan, Shiow-Lin; Gaur, Shikha; Hu, Shuya; Yen, Yun

    2015-05-10

    The pan-PI3K inhibitors are one treatment option for triple-negative breast cancer (TNBC). However, this treatment is ineffective for unknown reasons. Here, we report that aberrant expression of wingless-type MMTV integration site family (WNT) and activated WNT signals, which crosstalk with the PI3K-AKT-mTOR signaling pathway through GSK3β, plays the most critical role in resistance to pan-PI3K inhibitors in TNBC cells. GDC-0941 is a pan-PI3K inhibitor that activates the WNT/beta-catenin pathway in TNBC cells through stimulation of WNT secretion. GDC-0941-triggered WNT/beta-catenin pathway activation was observed in MDA-MB-231 and HCC1937 cells, which are TNBC cell lines showing aberrant WNT/beta-catenin activation, and not in SKBR3 and MCF7 cells. This observation is further investigated in vivo. GDC-0941 exhibited minimal tumor inhibition in MDA-MB-231 cells, but it significantly suppressed tumor growth in HER-positive SK-BR3 cells. In vivo mechanism study revealed the activation of WNT/beta-catenin pathway by GDC-0941. A synergistic effect was observed when combined treatment with GDC-0941 and the WNT inhibitor LGK974 at low concentrations in MDA-MB-231 cells. These findings indicated that WNT pathway activation conferred resistance in TNBC cells treated with GDC-0941. This resistance may be further circumvented through combined treatment with pan-PI3K and WNT inhibitors. Future clinical trials of these two inhibitors are warranted.

  4. The pan-PI3K inhibitor GDC-0941 activates canonical WNT signaling to confer resistance in TNBC cells: resistance reversal with WNT inhibitor

    PubMed Central

    Tzeng, Huey-En; Yang, Lixin; Chen, Kemin; Wang, Yafan; Liu, Yun-Ru; Pan, Shiow-Lin; Gaur, Shikha; Hu, Shuya; Yen, Yun

    2015-01-01

    The pan-PI3K inhibitors are one treatment option for triple-negative breast cancer (TNBC). However, this treatment is ineffective for unknown reasons. Here, we report that aberrant expression of wingless-type MMTV integration site family (WNT) and activated WNT signals, which crosstalk with the PI3K-AKT-mTOR signaling pathway through GSK3β, plays the most critical role in resistance to pan-PI3K inhibitors in TNBC cells. GDC-0941 is a pan-PI3K inhibitor that activates the WNT/beta-catenin pathway in TNBC cells through stimulation of WNT secretion. GDC-0941-triggered WNT/beta-catenin pathway activation was observed in MDA-MB-231 and HCC1937 cells, which are TNBC cell lines showing aberrant WNT/beta-catenin activation, and not in SKBR3 and MCF7 cells. This observation is further investigated in vivo. GDC-0941 exhibited minimal tumor inhibition in MDA-MB-231 cells, but it significantly suppressed tumor growth in HER-positive SK-BR3 cells. In vivo mechanism study revealed the activation of WNT/beta-catenin pathway by GDC-0941. A synergistic effect was observed when combined treatment with GDC-0941 and the WNT inhibitor LGK974 at low concentrations in MDA-MB-231 cells. These findings indicated that WNT pathway activation conferred resistance in TNBC cells treated with GDC-0941. This resistance may be further circumvented through combined treatment with pan-PI3K and WNT inhibitors. Future clinical trials of these two inhibitors are warranted. PMID:25857298

  5. Downregulation of uPAR inhibits migration, invasion, proliferation, FAK/PI3K/Akt signaling and induces senescence in papillary thyroid carcinoma cells.

    PubMed

    Nowicki, Theodore S; Zhao, Hong; Darzynkiewicz, Zbigniew; Moscatello, Augustine; Shin, Edward; Schantz, Stimson; Tiwari, Raj K; Geliebter, Jan

    2011-01-01

    Papillary thyroid carcinoma (PTC) is the most common endocrine and thyroid malignancy.  The urokinase plasminogen activator receptor (uPAR) plays an important role in cancer pathogenesis, including breakdown of the extracellular matrix, invasion, and metastasis.  Additionally, there is increasing evidence that uPAR also promotes tumorigenesis via the modulation of multiple signaling pathways.  BRAFV600E, the most common initial genetic mutation in PTC, leads to ERK1/2 hyperphosphorylation, which has been shown in numerous cancers to induce uPAR.  Treatment of the BRAFV600E-positive PTC cell line, BCPAP, with the MEK/ERK inhibitor U0126 reduced uPAR RNA levels by 90%.  siRNA-mediated down-regulation of uPAR in BCPAP cells resulted in greatly decreased activity in the focal adhesion kinase (FAK)/phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway.  This phenomenon was concurrent with drastically reduced proliferation rates and decreased clonigenic survival, as well as demonstrated senescence-associated nuclear morphology and induction of b-galactosidase activity. uPAR-knockdown BCPAP cells also displayed greatly reduced migration and invasion rates, as well as a complete loss of the cells' ability to augment their invasiveness following plasminogen supplementation. Taken together, these data provide new evidence of a novel role for uPAR induction (as a consequence of constitutive ERK1/2 activation) as a central component in PTC pathogenesis, and highlight the potential of uPAR as a therapeutic target. PMID:21191179

  6. Wortmannin, PI3K/Akt signaling pathway inhibitor, attenuates thyroid injury associated with severe acute pancreatitis in rats.

    PubMed

    Abliz, Ablikim; Deng, Wenhong; Sun, Rongze; Guo, Wenyi; Zhao, Liang; Wang, Weixing

    2015-01-01

    Increasing evidences suggest that PI3K/AKT pathway plays an important role in the pathogenesis of inflammatory diseases such as acute pancreatitis. However, the exact effect of PI3K/AKT on thyroid injury associated with acute pancreatitis has not been investigated. This study aimed to investigate the protective effects of wortmannin, PI3K/AKT inhibitor, on thyroid injury in a rat model of severe acute pancreatitis (SAP). Sixty male SD rats were randomly divided into four groups: sham operating group (SO), SAP group, wortmannin treatment (WOR) group and drug control (WOR-CON) group. Serum amylase (AMY), lipase (LIP) and thyroid hormone levels were evaluated. The morphological change of thyroid tissue was analyzed under the light and transmission electron microscopy. AKT, P38MAPK and NF-κB expression in the thyroid tissue was evaluated by immunohistochemical staining. Oxidative stress and inflammatory cytokines were detected. Results showed that wortmannin attenuated the following: (1) serum AMY, LIP and thyroid hormone (2) pancreatic and thyroid pathological injuries (3) thyroid MDA, (4) thyroid ultrastructural change, (5) serum TNF-α, IL-6 and IL-1β (6) AKT, MAPKP38 and NF-κB expression in thyroid tissues. These results suggested that wortmannin attenuates thyroid injury in SAP rats, presumably because of its role on prevent ROS generation and inhibits the activation of P38MAPK, NF-κB pathway. Our findings provide new therapeutic targets for thyroid injury associated with SAP. PMID:26823696

  7. Wortmannin, PI3K/Akt signaling pathway inhibitor, attenuates thyroid injury associated with severe acute pancreatitis in rats.

    PubMed

    Abliz, Ablikim; Deng, Wenhong; Sun, Rongze; Guo, Wenyi; Zhao, Liang; Wang, Weixing

    2015-01-01

    Increasing evidences suggest that PI3K/AKT pathway plays an important role in the pathogenesis of inflammatory diseases such as acute pancreatitis. However, the exact effect of PI3K/AKT on thyroid injury associated with acute pancreatitis has not been investigated. This study aimed to investigate the protective effects of wortmannin, PI3K/AKT inhibitor, on thyroid injury in a rat model of severe acute pancreatitis (SAP). Sixty male SD rats were randomly divided into four groups: sham operating group (SO), SAP group, wortmannin treatment (WOR) group and drug control (WOR-CON) group. Serum amylase (AMY), lipase (LIP) and thyroid hormone levels were evaluated. The morphological change of thyroid tissue was analyzed under the light and transmission electron microscopy. AKT, P38MAPK and NF-κB expression in the thyroid tissue was evaluated by immunohistochemical staining. Oxidative stress and inflammatory cytokines were detected. Results showed that wortmannin attenuated the following: (1) serum AMY, LIP and thyroid hormone (2) pancreatic and thyroid pathological injuries (3) thyroid MDA, (4) thyroid ultrastructural change, (5) serum TNF-α, IL-6 and IL-1β (6) AKT, MAPKP38 and NF-κB expression in thyroid tissues. These results suggested that wortmannin attenuates thyroid injury in SAP rats, presumably because of its role on prevent ROS generation and inhibits the activation of P38MAPK, NF-κB pathway. Our findings provide new therapeutic targets for thyroid injury associated with SAP.

  8. The roles of PI3K/Akt signaling pathway in regulating MC3T3-E1 preosteoblast proliferation and differentiation on SLA and SLActive titanium surfaces.

    PubMed

    Gu, Ying-Xin; Du, Juan; Si, Mi-Si; Mo, Jia-Ji; Qiao, Shi-Chong; Lai, Hong-Chang

    2013-03-01

    Chemical modification to produce a hydrophilic microrough titanium (Ti) implant surface has been shown to increase osseointegration compared with microrough topography alone. This study aimed to investigate the roles of PI3K/Akt signaling pathway in regulating proliferation and differentiation of osteoblasts in response to surface microroughness and hydrophilicity. Ti disks were manufactured to present different surface morphologies: a smooth pretreatment surface (PT), a rough hydrophobic surface that was sand-blasted, large-grit, acid-etched (SLA), and an SLA surface with the same roughness that was chemically modified to possess high wettability/hydrophilicity (SLActive/modSLA). MC3T3-E1 cells were cultured on these substrates with or without LY294002, a PI3K inhibitor, and their behaviors, including cell viability (MTT colorimetric assay), alkaline phosphatase (ALP) activity, and osteogenic genes expression of osteopontin (OPN) and osteocalcin (OCN) were measured. Western blot was applied to detect the expression of PI3K/Akt signal pathway proteins. The results showed that a decrease in osteoblast proliferation associated with the Ti surfaces (SLActive > SLA > PT) correlated with an increase in activity of the osteogenic differentiation markers ALP. The peak of ALP activity appeared earlier at 7 days for the SLActive surfaces compared with the SLA and PT surfaces. Osteoblast proliferation, as well as the level of p-Akt, was significantly inhibited by LY294002 in all three Ti surfaces. The top value of ALP activity was increased with the inhibition of PI3K/Akt signaling pathway while the time of the peak appeared was not advanced. The expression levels of OPN and OCN were upregulated by the effect of surface roughness and hydrophilicity, which were further enhanced by LY294002. In conclusion, osteogenic responses to SLActive surface were moderately better than the SLA surface and protein expression studies indicated that PI3K/Akt signaling activation may be

  9. Supercritical Fluid Extract of Spent Coffee Grounds Attenuates Melanogenesis through Downregulation of the PKA, PI3K/Akt, and MAPK Signaling Pathways

    PubMed Central

    Huang, Huey-Chun; Wei, Chien-Mei; Siao, Jen-Hung; Tsai, Tsang-Chi; Ko, Wang-Ping; Chang, Kuei-Jen; Hii, Choon-Hoon; Chang, Tsong-Min

    2016-01-01

    The mode of action of spent coffee grounds supercritical fluid CO2 extract (SFE) in melanogenesis has never been reported. In the study, the spent coffee grounds were extracted by the supercritical fluid CO2 extraction method; the chemical constituents of the SFE were investigated by gas chromatography-mass spectrometry (GC-MS). The effects of the SFE and its major fatty acid components on melanogenesis were evaluated by mushroom tyrosinase activity assay and determination of intracellular tyrosinase activity and melanin content. The expression level of melanogenesis-related proteins was analyzed by western blotting assay. The results revealed that the SFE of spent coffee grounds (1–10 mg/mL) and its major fatty acids such as linoleic acid and oleic acid (6.25–50 μM) effectively suppressed melanogenesis in the B16F10 murine melanoma cells. Furthermore, the SFE decreased the expression of melanocortin 1 receptor (MC1R), microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2 (TRP-2). The SFE also decreased the protein expression levels of p-JNK, p-p38, p-ERK, and p-CREB. Our results revealed that the SFE of spent coffee grounds attenuated melanogenesis in B16F10 cells by downregulation of protein kinase A (PKA), phosphatidylinositol-3-kinase (PI3K/Akt), and mitogen-activated protein kinases (MAPK) signaling pathways, which may be due to linoleic acid and oleic acid. PMID:27375763

  10. Supercritical Fluid Extract of Spent Coffee Grounds Attenuates Melanogenesis through Downregulation of the PKA, PI3K/Akt, and MAPK Signaling Pathways.

    PubMed

    Huang, Huey-Chun; Wei, Chien-Mei; Siao, Jen-Hung; Tsai, Tsang-Chi; Ko, Wang-Ping; Chang, Kuei-Jen; Hii, Choon-Hoon; Chang, Tsong-Min

    2016-01-01

    The mode of action of spent coffee grounds supercritical fluid CO2 extract (SFE) in melanogenesis has never been reported. In the study, the spent coffee grounds were extracted by the supercritical fluid CO2 extraction method; the chemical constituents of the SFE were investigated by gas chromatography-mass spectrometry (GC-MS). The effects of the SFE and its major fatty acid components on melanogenesis were evaluated by mushroom tyrosinase activity assay and determination of intracellular tyrosinase activity and melanin content. The expression level of melanogenesis-related proteins was analyzed by western blotting assay. The results revealed that the SFE of spent coffee grounds (1-10 mg/mL) and its major fatty acids such as linoleic acid and oleic acid (6.25-50 μM) effectively suppressed melanogenesis in the B16F10 murine melanoma cells. Furthermore, the SFE decreased the expression of melanocortin 1 receptor (MC1R), microphthalmia-associated transcription factor (MITF), tyrosinase, tyrosinase-related protein-1 (TRP-1), and tyrosinase-related protein-2 (TRP-2). The SFE also decreased the protein expression levels of p-JNK, p-p38, p-ERK, and p-CREB. Our results revealed that the SFE of spent coffee grounds attenuated melanogenesis in B16F10 cells by downregulation of protein kinase A (PKA), phosphatidylinositol-3-kinase (PI3K/Akt), and mitogen-activated protein kinases (MAPK) signaling pathways, which may be due to linoleic acid and oleic acid. PMID:27375763

  11. Abrogating endocrine resistance by targeting ERα and PI3K in breast cancer

    PubMed Central

    Fox, Emily M.; Arteaga, Carlos L.; Miller, Todd W.

    2012-01-01

    Antiestrogen therapies targeting estrogen receptor α (ER) signaling are a mainstay for patients with ER+ breast cancer. While many cancers exhibit resistance to antiestrogen therapies, a large body of clinical and experimental evidence indicates that hyperactivation of the phosphatidylinositol 3-kinase (PI3K) pathway promotes antiestrogen resistance. In addition, continued ligand-independent ER signaling in the setting of estrogen deprivation may contribute to resistance to endocrine therapy. PI3K activates several proteins which promote cell cycle progression and survival. In ER+ breast cancer cells, PI3K promotes ligand-dependent and -independent ER transcriptional activity. Models of antiestrogen-resistant breast cancer often remain sensitive to estrogen stimulation and PI3K inhibition, suggesting that clinical trials with combinations of drugs targeting both the PI3K and ER pathways are warranted. Herein, we review recent findings on the roles of PI3K and ER in antiestrogen resistance, and clinical trials testing drug combinations which target both pathways. We also discuss the need for clinical investigation of ER downregulators in combination with PI3K inhibitors. PMID:23087906

  12. MiR-126 regulates proliferation and invasion in the bladder cancer BLS cell line by targeting the PIK3R2-mediated PI3K/Akt signaling pathway

    PubMed Central

    Xiao, Jun; Lin, Huan-Yi; Zhu, Yuan-Yuan; Zhu, Yu-Ping; Chen, Ling-Wu

    2016-01-01

    Objective To assess whether microRNA-126 (miR-126) targets phosphatidylinositol 3-kinase regulatory subunit beta (PIK3R2) and to determine the potential roles of miR-126 in regulating proliferation and invasion via the PIK3R2-mediated phosphatidylinositol 3 kinase (PI3K)-protein kinase B (Akt) signaling pathway in the human bladder BLS cell line. Materials and methods A recombinant lentivirus (Lv) vector expressing miR-216 (Lv-miR-126) was successfully constructed, and Lv-miR-126 and Lv vector were transfected into the BLS cell line. A direct regulatory relationship between miR-126 and the PIK3R2 gene was demonstrated by luciferase reporter assays. To determine whether PIK3R2 directly participates in the miR-126-induced effects in BLS cells, anti-miR-126 and a PIK3R2 small interfering RNA (siRNA) were transfected into the BLS cells. Quantitative real-time polymerase chain reaction was used to measure miR-126 and PIK3R2 expressions. 5-Ethynyl-2′-deoxyuridine and colony formation assays to assess cell proliferation, flow cytometry for cell apoptosis and cell cycle analysis, Transwell assays for cell migration and invasion, and Western blots for PIK3R2, PI3K, phosphorylated PI3K (p-PI3K), Akt, and phosphorylated Akt (p-Akt) protein expressions were performed. Results Lv-miR-126 significantly enhanced the relative expression of miR-126 in the BLS cells after infection (P<0.0001). MiR-126 overexpression inhibited the proliferation, cloning, migration, and invasion of BLS cells, promoted cell apoptosis, and induced S phase arrest (all P<0.05). PIK3R2, p-PI3K, and p-Akt protein expressions were significantly decreased in the BLS cells infected with Lv-miR-126. Luciferase assays showed that miR-126 significantly inhibited the PIK3R2 3′ untranslated region (3′UTR) luciferase reporter activity (P<0.05). The anti-miR-126 + PIK3R2 siRNA group had significantly decreased PIK3R2, p-PI3K, and p-Akt expressions compared with those of anti-miR-126 alone, as well as

  13. Targeted Regulation of PI3K/Akt/mTOR/NF-κB Signaling by Indole Compounds and their Derivatives: Mechanistic Details and Biological Implications for Cancer Therapy

    PubMed Central

    Ahmad, Aamir; Biersack, Bernhard; Li, Yiwei; Kong, Dejuan; Bao, Bin; Schobert, Rainer; Padhye, Subhash B.; Sarkar, Fazlul H.

    2014-01-01

    Indole compounds, found in cruciferous vegetables, are potent anti-cancer agents. Studies with indole-3-carbinol (I3C) and its dimeric product, 3,3’-diindolylmethane (DIM) suggest that these compounds have the ability to deregulate multiple cellular signaling pathways, including PI3K/Akt/mTOR signaling pathway. These natural compounds are also effective modulators of downstream transcription factor NF-κB signaling which might help explain their ability to inhibit invasion and angiogenesis, and the reversal of epithelial-to-mesenchymal transition (EMT) phenotype and drug resistance. Signaling through PI3K/Akt/mTOR and NF-κB pathway is increasingly being realized to play important role in EMT through the regulation of novel miRNAs which further validates the importance of this signaling network and its regulations by indole compounds. Here we will review the available literature on the modulation of PI3K/Akt/mTOR/NF-κB signaling by both parental I3C and DIM, as well as their analogs/derivatives, in an attempt to catalog their anticancer activity. PMID:23272910

  14. Mouse model of human ovarian endometrioid adenocarcinoma based on somatic defects in the Wnt/beta-catenin and PI3K/Pten signaling pathways.

    PubMed

    Wu, Rong; Hendrix-Lucas, Neali; Kuick, Rork; Zhai, Yali; Schwartz, Donald R; Akyol, Aytekin; Hanash, Samir; Misek, David E; Katabuchi, Hidetaka; Williams, Bart O; Fearon, Eric R; Cho, Kathleen R

    2007-04-01

    One histologic subtype of ovarian carcinoma, ovarian endometrioid adenocarcinoma (OEA), frequently harbors mutations that constitutively activate Wnt/beta-catenin-dependent signaling. We now show that defects in the PI3K/Pten and Wnt/beta-catenin signaling pathways often occur together in a subset of human OEAs, suggesting their cooperation during OEA pathogenesis. Deregulation of these two pathways in the murine ovarian surface epithelium by conditional inactivation of the Pten and Apc tumor suppressor genes results in the formation of adenocarcinomas morphologically similar to human OEAs with 100% penetrance, short latency, and rapid progression to metastatic disease in upwards of 75% of mice. The biological behavior and gene expression patterns of the murine cancers resemble those of human OEAs with defects in the Wnt/beta-catenin and PI3K/Pten pathways.

  15. Pseudomonas aeruginosa pyocyanin activates NRF2-ARE-mediated transcriptional response via the ROS-EGFR-PI3K-AKT/MEK-ERK MAP kinase signaling in pulmonary epithelial cells.

    PubMed

    Xu, Ying; Duan, Chaohui; Kuang, Zhizhou; Hao, Yonghua; Jeffries, Jayme L; Lau, Gee W

    2013-01-01

    The redox-active pyocyanin (PCN) secreted by the respiratory pathogen Pseudomonas aeruginosa generates reactive oxygen species (ROS) and causes oxidative stress to pulmonary epithelial cells. Nuclear factor (erythroid-derived 2)-like 2 (NRF2) confers protection against ROS-mediated cell death by inducing the expression of detoxifying enzymes and proteins via its binding to the cis-acting antioxidant response element (ARE). However, a clear relationship between NRF2 and PCN-mediated oxidative stress has not been established experimentally. In this study, we investigated the induction of NRF2-ARE response by PCN in the pulmonary epithelial cells. We analyzed the effect of PCN on NRF2 expression and nuclear translocation in cultured human airway epithelial cells, and in a mouse model of chronic PCN exposure. NRF2-dependent transcription of antioxidative enzymes was also assessed. Furthermore, we used inhibitors to examine the involvement of EGFR and its downstream signaling components that mediate NRF2-ARE-activation in response to PCN. PCN enhances the nuclear NRF2 accumulation and activates the transcription of ARE-mediated antioxidant genes. Furthermore, PCN activates NRF2 by inducing the EGFR-phosphoinositide-3-kinase (PI3K) signaling pathway and its main downstream effectors, AKT and MEK1/2-ERK1/2 MAP kinases. Inhibition of the EGFR-PI3K signaling markedly attenuates PCN-stimulated NRF2 accumulation in the nucleus. We demonstrate for the first time that PCN-mediated oxidative stress activates the EGFR-PI3K-AKT/MEK1/2-ERK1/2 MAP kinase signaling pathway, leading to nuclear NRF2 translocation and ARE responsiveness in pulmonary epithelial cells.

  16. Induction of Multidrug Resistance of Acute Myeloid Leukemia Cells by Cocultured Stromal Cells via Upregulation of the PI3K/Akt Signaling Pathway.

    PubMed

    Chen, Ping; Jin, Qing; Fu, Qiang; You, Peidong; Jiang, Xi; Yuan, Qin; Huang, Huifang

    2016-01-01

    This study aimed to investigate the role of the PI3K/Akt signaling pathway in multidrug resistance of acute myeloid leukemia (AML) cells induced by cocultured stromal cells. Human AML cell lines HL-60 and U937 were adhesion cocultured with human bone marrow stromal cell line HS-5 cells. Such coculturing induced HL-60 and U937 cells resistant to chemotherapeutic drugs including daunorubicin (DNR), homoharringtonine (HHT), and cytosine arabinoside (Ara-C). The coculturing-induced resistance of AML cells to DNR, HHT, and Ara-C can be partially reversed by inhibition of the PI3K/Akt signaling pathway. Clinically, AML patients with a low level of PTEN and a high level of CCND1 had high relapse rates within 1 year, and newly diagnosed AML patients with extramedullary infiltration had a low level of PTEN. This study confirms the involvement of the PI3K/Akt signaling pathway in multidrug resistance in AML cells induced by stroma and suggests that the expression of PTEN and CCND1 may be a prognostic indicator for AML. PMID:27656831

  17. Jolkinolide B induces apoptosis in MCF-7 cells through inhibition of the PI3K/Akt/mTOR signaling pathway.

    PubMed

    Xu, Hui-Yu; Chen, Zhi-Wei; Hou, Jin-Cai; Du, Feng-Xia; Liu, Ji-Cheng

    2013-01-01

    The aim of this study was to explore the molecular mechanisms of jolkinolide B (JB), which is extracted from the root of Euphorbia fischeriana Steud. In this study, we found that JB, a diterpenoid from the traditional Chinese medicinal herb, strongly inhibited the PI3K/Akt/mTOR signaling pathway. Furthermore, we evaluated the effects of JB on the proliferation and apoptosis of MCF-7 human breast cancer cells. Our results showed significant induction of apoptosis in MCF-7 cells incubated with JB. The viability of the MCF-7 cells was assessed by MTT assay. Flow cytometry was used to detect apoptosis and cell cycle analysis. Transmission electron microscopy (TEM) analysis was used to observe cell morphology. MCF-7 cells were subcutaneously inoculated into nude mice to study the in vivo antitumor effects of JB. The growth of MCF-7 cells was inhibited and arrested in the S phase by JB. The data showed significantly decreased tumor volume and weight in nude mice inoculated with MCF-7 cells. In addition, treatment with JB was able to induce downregulation of cyclinD1, cyclinE, mTOR, p-PI3K and p-Akt, and upregulation of PTEN and p-eIF4E. Collectively, JB-induced apoptosis of MCF-7 cells occurs through the PI3K/Akt/mTOR signaling pathway. Furthermore, the PI3K/Akt signaling cascade plays a role in the induction of apoptosis in JB-treated cells. These observations suggest that JB may have therapeutic applications in the treatment of cancer.

  18. Selective Inhibition of PI3K/Akt/mTOR Signaling Pathway Regulates Autophagy of Macrophage and Vulnerability of Atherosclerotic Plaque

    PubMed Central

    Zhai, Chungang; Cheng, Jing; Mujahid, Haroon; Wang, Hefeng; Kong, Jing; Yin, Yue; Li, Jifu; Zhang, Yun; Ji, Xiaoping; Chen, Wenqiang

    2014-01-01

    Macrophage infiltration contributes to the instability of atherosclerotic plaques. In the present study, we investigated whether selective inhibition of PI3K/Akt/mTOR signaling pathway can enhance the stability of atherosclerotic plaques by activation of macrophage autophagy. In vitro study, selective inhibitors or siRNA of PI3K/Akt/mTOR pathways were used to treat the rabbit's peritoneal primary macrophage cells. Inflammation related cytokines secreted by macrophages were measured. Ultrastructure changes of macrophages were examined by transmission electron microscope. mRNA or protein expression levels of autophagy related gene Beclin 1, protein 1 light chain 3 II dots (LC3-II) or Atg5-Atg12 conjugation were assayed by quantitative RT-PCR or Western blot. In vivo study, vulnerable plaque models were established in 40 New Zealand White rabbits and then drugs or siRNA were given for 8 weeks to inhibit the PI3K/Akt/mTOR signaling pathway. Intravascular ultrasound (IVUS) was performed to observe the plaque imaging. The ultrastructure of the abdominal aortic atherosclerosis lesions were analyzed with histopathology. RT-PCR or Western blot methods were used to measure the expression levels of corresponding autophagy related molecules. We found that macrophage autophagy was induced in the presence of Akt inhibitor, mTOR inhibitor and mTOR-siRNA in vitro study, while PI3K inhibitor had the opposite role. In vivo study, we found that macrophage autophagy increased significantly and the rabbits had lower plaque rupture incidence, lower plaque burden and decreased vulnerability index in the inhibitors or siRNA treated groups. We made a conclusion that selective inhibition of the Akt/mTOR signal pathway can reduce macrophages and stabilize the vulnerable atherosclerotic plaques by promoting macrophage autophagy. PMID:24599185

  19. Minoxidil sulfate induced the increase in blood-brain tumor barrier permeability through ROS/RhoA/PI3K/PKB signaling pathway.

    PubMed

    Gu, Yan-ting; Xue, Yi-xue; Wang, Yan-feng; Wang, Jin-hui; Chen, Xia; ShangGuan, Qian-ru; Lian, Yan; Zhong, Lei; Meng, Ying-nan

    2013-12-01

    Adenosine 5'-triphosphate-sensitive potassium channel (KATP channel) activator, minoxidil sulfate (MS), can selectively increase the permeability of the blood-tumor barrier (BTB); however, the mechanism by which this occurs is still under investigation. Using a rat brain glioma (C6) model, we first examined the expression levels of occludin and claudin-5 at different time points after intracarotid infusion of MS (30 μg/kg/min) by western blotting. Compared to MS treatment for 0 min group, the protein expression levels of occludin and claudin-5 in brain tumor tissue of rats showed no changes within 1 h and began to decrease significantly after 2 h of MS infusion. Based on these findings, we then used an in vitro BTB model and selective inhibitors of diverse signaling pathways to investigate whether reactive oxygen species (ROS)/RhoA/PI3K/PKB pathway play a key role in the process of the increase of BTB permeability induced by MS. The inhibitor of ROS or RhoA or PI3K or PKB significantly attenuated the expression of tight junction (TJ) protein and the increase of the BTB permeability after 2 h of MS treatment. In addition, the significant increases in RhoA activity and PKB phosphorylation after MS administration were observed, which were partly inhibited by N-2-mercaptopropionyl glycine (MPG) or C3 exoenzyme or LY294002 pretreatment. The present study indicates that the activation of signaling cascades involving ROS/RhoA/PI3K/PKB in BTB was required for the increase of BTB permeability induced by MS. Taken together, all of these results suggested that MS might increase BTB permeability in a time-dependent manner by down-regulating TJ protein expression and this effect could be related to ROS/RhoA/PI3K/PKB signal pathway. PMID:23973310

  20. Nephroprotective Effects of Polydatin against Ischemia/Reperfusion Injury: A Role for the PI3K/Akt Signal Pathway

    PubMed Central

    Liu, Hong-Bao; Meng, Qiu-Hong; Huang, Chen; Wang, Jian-Bo; Liu, Xiao-Wei

    2015-01-01

    Oxidative stress and inflammation are involved in the pathogenesis in renal ischemia/reperfusion (I/R) injury. It has been demonstrated that polydatin processed the antioxidative, anti-inflammatory, and nephroprotective properties. However, whether it has beneficial effects and the possible mechanisms on renal I/R injury remain unclear. In our present study I/R models were simulated both in vitro and in vivo. Compared with vehicle control, the administration of polydatin significantly improved the renal function, accelerated the mitogenic response and reduced cell apoptosis in renal I/R injury models, strongly suppressed the I/R-induced upregulation of the expression of tumor necrosis factor-α, interleukin-1β, cyclooxygenase-2, inducible nitric oxide synthase, prostaglandin E-2, and nitric oxide levels, and dramatically decreased contents of malondialdehyde, but it increased the activity of superoxide dismutase, glutathione transferase, glutathione peroxidase and catalase, and the level of glutathione. Further investigation showed that polydatin upregulated the phosphorylation of Akt in kidneys of I/R injury dose-dependently. However, all beneficial effects of polydatin mentioned above were counteracted when we inhibited PI3K/Akt pathway with its specific inhibitor, wortmannin. Taken together, the present findings provide the first evidence demonstrating that PD exhibited prominent nephroprotective effects against renal I/R injury by antioxidative stress and inflammation through PI3-K/Akt-dependent molecular mechanisms. PMID:26576221

  1. PI3K/AKT signaling modulates transcriptional expression of EWS/FLI1 through specificity protein 1

    PubMed Central

    Giorgi, Chiara; Boro, Aleksandar; Rechfeld, Florian; Lopez-Garcia, Laura A.; Gierisch, Maria E.; Schäfer, Beat W.; Niggli, Felix K.

    2015-01-01

    Ewing sarcoma (ES) is the second most frequent bone cancer in childhood and is characterized by the presence of the balanced translocation t(11;22)(q24;q12) in more than 85% of cases, generating a dysregulated transcription factor EWS/FLI1. This fusion protein is an essential oncogenic component of ES development which is necessary for tumor cell maintenance and represents an attractive therapeutic target. To search for modulators of EWS/FLI1 activity we screened a library of 153 targeted compounds and identified inhibitors of the PI3K pathway to directly modulate EWS/FLI1 transcription. Surprisingly, treatment of four different ES cell lines with BEZ235 resulted in down regulation of EWS/FLI1 mRNA and protein by ∼50% with subsequent modulation of target gene expression. Analysis of the EWS/FLI1 promoter region (−2239/+67) using various deletion constructs identified two 14bp minimal elements as being important for EWS/FLI1 transcription. We identified SP1 as modulator of EWS/FLI1 gene expression and demonstrated direct binding to one of these regions in the EWS/FLI1 promoter by EMSA and ChIP experiments. These results provide the first insights on the transcriptional regulation of EWS/FLI1, an area that has not been investigated so far, and offer an additional molecular explanation for the known sensitivity of ES cell lines to PI3K inhibition. PMID:26336820

  2. A2B adenosine receptor contributes to penile erection via PI3K/AKT signaling cascade-mediated eNOS activation

    PubMed Central

    Wen, Jiaming; Grenz, Almut; Zhang, Yujin; Dai, Yingbo; Kellems, Rodney E.; Blackburn, Michael R.; Eltzschig, Holger K.; Xia, Yang

    2011-01-01

    Normal penile erection is under the control of multiple factors and signaling pathways. Although adenosine signaling is implicated in normal and abnormal penile erection, the exact role and the underlying mechanism for adenosine signaling in penile physiology remain elusive. Here we report that shear stress leads to increased adenosine release from endothelial cells. Subsequently, we determined that ecto-5′-nucleotidase (CD73) is a key enzyme required for the production of elevated adenosine from ATP released by shear-stressed endothelial cells. Mechanistically, we demonstrate that shear stress-mediated elevated adenosine functions through the adenosine A2B receptor (A2BR) to activate the PI3K/AKT signaling cascade and subsequent increased endothelial nitric oxide synthase (eNOS) phosphorylation. These in vitro studies led us to discover further that adenosine was induced during sustained penile erection and contributes to PI3K/AKT activation and subsequent eNOS phosphorylation via A2BR signaling in intact animal. Finally, we demonstrate that lowering adenosine in wild-type mice or genetic deletion of A2BR in mutant mice significantly attenuated PI3K/AKT activation, eNOS phosphorylation, and subsequent impaired penile erection featured with the reduction of ratio of maximal intracavernosal pressure to systemic arterial pressure from 0.49 ± 0.03 to 0.41 ± 0.05 and 0.38 ± 0.04, respectively (both P<0.05). Overall, using biochemical, cellular, genetic, and physiological approaches, our findings reveal that adenosine is a novel molecule signaling via A2BR activation, contributing to penile erection via PI3K/AKT-dependent eNOS activation. These studies suggest that this signaling pathway may be a novel therapeutic target for erectile disorders.—Wen, J., Grenz, A., Zhang, Y., Dai, Y., Kellems, R. E., Blackburn, M. R., Eltzschig, H. K., Xia, Y. A2B adenosine receptor contributes to penile erection via PI3K/AKT signaling cascade-mediated eNOS activation. PMID

  3. Glucagon-like peptide-1 protects cardiomyocytes from advanced oxidation protein product-induced apoptosis via the PI3K/Akt/Bad signaling pathway.

    PubMed

    Zhang, Hua; Xiong, Zhouyi; Wang, Jiao; Zhang, Shuangshuang; Lei, Lei; Yang, Li; Zhang, Zhen

    2016-02-01

    Cardiomyocyte apoptosis is a major event in the pathogenesis of diabetic cardiomyopathy. Currently, no single effective treatment for diabetic cardiomyopathy exists. The present study investigated whether advanced oxidative protein products (AOPPs) have a detrimental role in the survival of cardiomyocytes and if glucagon-like peptide-1 (GLP-1) exerts a cardioprotective effect under these circumstances. The present study also aimed to determine the underlying mechanisms. H9c2 cells were exposed to increasing concentrations of AOPPs in the presence or absence of GLP-1, and the viability and apoptotic rate were detected using a cell counting kit-8 assay and flow cytometry, respectively. In addition, a phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) inhibitor, LY294002, was employed to illustrate the mechanism of the antiapoptotic effect of GLP-1. The expression levels of the apoptotic-associated proteins, Akt, B-cell lymphoma (Bcl)-2, Bcl-2-associated death promoter (Bad), Bcl-2-associated X protein (Bax) and caspase-3 were measured by western blotting. It was revealed that GLP-1 significantly attenuated AOPP-induced cell toxicity and apoptosis. AOPPs inactivated the phosphorylation of Akt, reduced the phosphorylation of Bad, decreased the expression of Bcl-2, increased the expression of Bax and the activation of caspase-3 in H9c2 cells. GLP-1 reversed the above changes induced by AOPPs and the protective effects of GLP-1 were abolished by the PI3K inhibitor, LY294002. In conclusion, the present data suggested that GLP-1 protected cardiomyocytes against AOPP-induced apoptosis, predominantly via the PI3K/Akt/Bad pathway. These results provided a conceivable mechanism for the development of diabetic cardiomyopathy and rendered a novel application of GLP-1 exerting favorable cardiac effects for the treatment of diabetic cardiomyopathy.

  4. Glucagon-like peptide-1 protects cardiomyocytes from advanced oxidation protein product-induced apoptosis via the PI3K/Akt/Bad signaling pathway

    PubMed Central

    ZHANG, HUA; XIONG, ZHOUYI; WANG, JIAO; ZHANG, SHUANGSHUANG; LEI, LEI; YANG, LI; ZHANG, ZHEN

    2016-01-01

    Cardiomyocyte apoptosis is a major event in the pathogenesis of diabetic cardiomyopathy. Currently, no single effective treatment for diabetic cardiomyopathy exists. The present study investigated whether advanced oxidative protein products (AOPPs) have a detrimental role in the survival of cardiomyocytes and if glucagon-like peptide-1 (GLP-1) exerts a cardioprotective effect under these circumstances. The present study also aimed to determine the underlying mechanisms. H9c2 cells were exposed to increasing concentrations of AOPPs in the presence or absence of GLP-1, and the viability and apoptotic rate were detected using a cell counting kit-8 assay and flow cytometry, respectively. In addition, a phosphatidylino-sitol-4,5-bisphosphate 3-kinase (PI3K) inhibitor, LY294002, was employed to illustrate the mechanism of the antiapoptotic effect of GLP-1. The expression levels of the apoptotic-associated proteins, Akt, B-cell lymphoma (Bcl)-2, Bcl-2-associated death promoter (Bad), Bcl-2-associated X protein (Bax) and caspase-3 were measured by western blotting. It was revealed that GLP-1 significantly attenuated AOPP-induced cell toxicity and apoptosis. AOPPs inactivated the phosphorylation of Akt, reduced the phosphorylation of Bad, decreased the expression of Bcl-2, increased the expression of Bax and the activation of caspase-3 in H9c2 cells. GLP-1 reversed the above changes induced by AOPPs and the protective effects of GLP-1 were abolished by the PI3K inhibitor, LY294002. In conclusion, the present data suggested that GLP-1 protected cardiomyocytes against AOPP-induced apoptosis, predominantly via the PI3K/Akt/Bad pathway. These results provided a conceivable mechanism for the development of diabetic cardiomyopathy and rendered a novel application of GLP-1 exerting favorable cardiac effects for the treatment of diabetic cardiomyopathy. PMID:26717963

  5. Gene expression profiling of LPS-stimulated murine macrophages and role of the NF-kappaB and PI3K/mTOR signaling pathways.

    PubMed

    Dos Santos, S; Delattre, A-I; De Longueville, F; Bult, H; Raes, M

    2007-01-01

    Lipopolysaccharide (LPS), a major component of the outer membrane of Gram-negative bacteria, activates a broad spectrum of signaling pathways in immune cells. In this article, RAW264.7 cells have been stimulated for 4 h with 1 microg/mL of LPS in the presence or not of specific inhibitors of the NF-kappaB pathway (BAY 11-7082) and the PI3K pathway (LY294002). Gene expression profiles were characterized using the DNA microarray "Dual Chip Mouse Inflammation." This array monitors the expression of 233 genes encoding proteins playing a role in inflammation. Both signaling pathways exert an important role in the response to LPS, but they are not completely overlapping. For example, genes encoding the PAF receptor, PAI-1, PlA2 (group V), IL-13 receptor (alpha2), and GTP cyclohydrolase 1, were upregulated after LPS treatment, but this upregulation was counteracted by LY294002. The same was observed for BAY 11-7082: genes encoding the kit ligand, TLR2, or TNFRSF5 were mainly under the control of NF-kappaB. NF-kappaB plays an important role in the macrophage response to LPS, but we have also shown that the PI3K pathway partially contributes to it. Further experiments with the specific inhibitor of mTOR (rapamycin) will provide more information on the specific contribution of the PI3K/mTOR pathway in the inflammatory response in LPS-stimulated macrophages.

  6. Momordin Ic couples apoptosis with autophagy in human hepatoblastoma cancer cells by reactive oxygen species (ROS)-mediated PI3K/Akt and MAPK signaling pathways.

    PubMed

    Mi, Yashi; Xiao, Chunxia; Du, Qingwei; Wu, Wanqiang; Qi, Guoyuan; Liu, Xuebo

    2016-01-01

    Momordin Ic is a principal saponin constituent of Fructus Kochiae, which acts as an edible and pharmaceutical product more than 2000 years in China. Our previous research found momordin Ic induced apoptosis by PI3K/Akt and MAPK signaling pathways in HepG2 cells. While the role of autophagy in momordin Ic induced cell death has not been discussed, and the connection between the apoptosis and autophagy is not clear yet. In this work, we reported momordin Ic promoted the formation of autophagic vacuole and expression of Beclin 1 and LC-3 in a dose- and time-dependent manner. Compared with momordin Ic treatment alone, the autophagy inhibitor 3-methyladenine (3-MA) also can inhibit apoptosis, while autophagy activator rapamycin (RAP) has the opposite effect, and the apoptosis inhibitor ZVAD-fmk also inhibited autophagy induced by momordin Ic. Momordin Ic simultaneously induces autophagy and apoptosis by suppressing the ROS-mediated PI3K/Akt and activating the ROS-related JNK and P38 pathways. Additionally, momordin Ic induces apoptosis by suppressing PI3K/Akt-dependent NF-κB pathways and promotes autophagy by ROS-mediated Erk signaling pathway. Those results suggest that momordin Ic has great potential as a nutritional preventive strategy in cancer therapy. PMID:26593748

  7. Fra-1 is upregulated in gastric cancer tissues and affects the PI3K/Akt and p53 signaling pathway in gastric cancer.

    PubMed

    He, Junyu; Zhu, Guangchao; Gao, Lu; Chen, Pan; Long, Yuehua; Liao, Shan; Yi, Hong; Yi, Wei; Pei, Zhen; Wu, Minghua; Li, Xiaoling; Xiang, Juanjuan; Peng, Shuping; Ma, Jian; Zhou, Ming; Xiong, Wei; Zeng, Zhaoyang; Xiang, Bo; Tang, Ke; Cao, Li; Li, Guiyuan; Zhou, Yanhong

    2015-11-01

    Gastric cancer is an aggressive disease that continues to have a daunting impact on global health. Fra-1 (FOSL1) plays important roles in oncogenesis in various malignancies. We investigated the expression of Fra-1 in gastric cancer (GC) tissues by qPCR, immunohistochemistry (IHC) and western blot technologies. The results showed that Fra-1 was overexpressed in gastric cancer tissues compared with the adjacent non‑cancerous tissues. To explore the possible mechanism of Fra-1 in GC, we elucidated the effect of Fra-1 in the apoptosis and cell cycle of gastric cancer cells, AGS, and found that a considerable decrease in apoptotic cells and increase of S phase rate were observed for AGS cells with Fra-1 overexpession. We identified and confirmed that Fra-1 affected the expression level of CTTN and EZR in vitro through LC-MS/MS analyses and western blot technology. Furthermore, we found that Fra-1 was correlated with dysregulation PI3K/Akt and p53 signaling pathway in gastric cancer tissues in vitro. Moreover, we found that Fra-1 overexpression affected the expression of PI3K, Akt, MDM2 and p53 in vivo. In summary, our results suggest that Fra-1 is upregulated in gastric cancer tissues and plays its function by affecting the PI3K/Akt and p53 signaling pathway in gastric cancer.

  8. Investigation of PI3K/PKB/mTOR/S6K1 signaling pathway in relationship of type 2 diabetes and Alzheimer’s disease

    PubMed Central

    Ma, Yunqing; Wu, Dongke; Zhang, Wei; Liu, Jiankun; Chen, Siping; Hua, Binghong

    2015-01-01

    The aim of this study was to investigate the roles of PI3K/PKB/mTOR/S6K1 signaling pathway in the risk-increasing mechanisms of type 2 diabetes mellitus (T2DM) towards the Alzheimer’s disease (AD). Based on the high-sugar high-fat diet, the single intraperitoneal injection of streptozotocin was performed to induce the T2DM rat model; the immunohistochemistry and RT-PCR technique were then performed to detect the expression levels of mTOR, PI3K, PKB, S6K1 and phosphorylated Tau protein in the hippocampal tissues of each group. The related metabolic indicators of the T2DM group and the T2DM + AD group were significantly higher than the normal control group and the AD group (P<0.01); the Morris water maze test of the AD group and the learning and memory of the T2DM + AD group were than significantly decreased than the T2DM group (P<0.01); the T2DM + AD group exhibited significantly increased expression levels of mTOR, S6K1 and Tau protein in the hippocampal tissues than the AD group and the T2DM group (P<0.05), and while the expression levels of PI3K and PKB were decreased (P<0.05). Among the possible mechanisms through which T2DM increased the risk of AD, the dystransduction of insulin signaling pathway (PI3K/PKB/mTOR/S6K1) was the important cause of hyperphosphorylation of Tau protein, thus it prompted the AD occurrence. PMID:26770471

  9. 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.

  10. Interleukin-6-mediated functional upregulation of TRPV1 receptors in dorsal root ganglion neurons through the activation of JAK/PI3K signaling pathway: roles in the development of bone cancer pain in a rat model.

    PubMed

    Fang, Dong; Kong, Ling-Yu; Cai, Jie; Li, Song; Liu, Xiao-Dan; Han, Ji-Sheng; Xing, Guo-Gang

    2015-06-01

    Primary and metastatic cancers that affect bone are frequently associated with severe and intractable pain. The mechanisms underlying the pathogenesis of bone cancer pain still remain largely unknown. Previously, we have reported that sensitization of primary sensory dorsal root ganglion (DRG) neurons contributes to the pathogenesis of bone cancer pain in rats. In addition, numerous preclinical and clinical studies have revealed the pathological roles of interleukin-6 (IL-6) in inflammatory and neuropathic hyperalgesia. In this study, we investigated the role and the underlying mechanisms of IL-6 in the development of bone cancer pain using in vitro and in vivo approaches. We first demonstrated that elevated IL-6 in DRG neurons plays a vital role in the development of nociceptor sensitization and bone cancer-induced pain in a rat model through IL-6/soluble IL-6 receptor (sIL-6R) trans-signaling. Moreover, we revealed that functional upregulation of transient receptor potential vanilloid channel type 1 (TRPV1) in DRG neurons through the activation of Janus kinase (JAK)/phosphatidylinositol 3-kinase (PI3K) signaling pathway contributes to the effects of IL-6 on the pathogenesis of bone cancer pain. Therefore, suppression of functional upregulation of TRPV1 in DRG neurons by the inhibition of JAK/PI3K pathway, either before surgery or after surgery, reduces the hyperexcitability of DRG neurons and pain hyperalgesia in bone cancer rats. We here disclose a novel intracellular pathway, the IL-6/JAK/PI3K/TRPV1 signaling cascade, which may underlie the development of peripheral sensitization and bone cancer-induced pain.

  11. Antiviral Potential of ERK/MAPK and PI3K/AKT/mTOR Signaling Modulation for Middle East Respiratory Syndrome Coronavirus Infection as Identified by Temporal Kinome Analysis

    PubMed Central

    Ork, Britini; Hart, Brit J.; Holbrook, Michael R.; Frieman, Matthew B.; Traynor, Dawn; Johnson, Reed F.; Dyall, Julie; Olinger, Gene G.; Hensley, Lisa E.

    2014-01-01

    Middle East respiratory syndrome coronavirus (MERS-CoV) is a lineage C betacoronavirus, and infections with this virus can result in acute respiratory syndrome with renal failure. Globally, MERS-CoV has been responsible for 877 laboratory-confirmed infections, including 317 deaths, since September 2012. As there is a paucity of information regarding the molecular pathogenesis associated with this virus or the identities of novel antiviral drug targets, we performed temporal kinome analysis on human hepatocytes infected with the Erasmus isolate of MERS-CoV with peptide kinome arrays. bioinformatics analysis of our kinome data, including pathway overrepresentation analysis (ORA) and functional network analysis, suggested that extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) and phosphoinositol 3-kinase (PI3K)/serine-threonine kinase (AKT)/mammalian target of rapamycin (mTOR) signaling responses were specifically modulated in response to MERS-CoV infection in vitro throughout the course of infection. The overrepresentation of specific intermediates within these pathways determined by pathway and functional network analysis of our kinome data correlated with similar patterns of phosphorylation determined through Western blot array analysis. In addition, analysis of the effects of specific kinase inhibitors on MERS-CoV infection in tissue culture models confirmed these cellular response observations. Further, we have demonstrated that a subset of licensed kinase inhibitors targeting the ERK/MAPK and PI3K/AKT/mTOR pathways significantly inhibited MERS-CoV replication in vitro whether they were added before or after viral infection. Taken together, our data suggest that ERK/MAPK and PI3K/AKT/mTOR signaling responses play important roles in MERS-CoV infection and may represent novel drug targets for therapeutic intervention strategies. PMID:25487801

  12. PI3K/Akt signaling pathway triggers P2X7 receptor expression as a pro-survival factor of neuroblastoma cells under limiting growth conditions

    PubMed Central

    Gómez-Villafuertes, Rosa; García-Huerta, Paula; Díaz-Hernández, Juan Ignacio; Miras-Portugal, Mª Teresa

    2015-01-01

    The expression of purinergic P2X7 receptor (P2X7R) in neuroblastoma cells is associated to accelerated growth rate, angiogenesis, metastasis and poor prognosis. Noticeably, P2X7R allows the survival of neuroblastoma cells under restrictive conditions, including serum and glucose deprivation. Previously we identified specificity protein 1 (Sp1) as the main factor involved in the transcriptional regulation of P2rx7 gene, reporting that serum withdrawal triggers the expression of P2X7R in Neuro-2a (N2a) neuroblastoma cell line. Here we demonstrate that PI3K/Akt pathway is crucial for the upregulation of P2X7R expression in serum-deprived neuroblastoma cells, circumstance that facilitates cell proliferation in the absence of trophic support. The effect exerted by PI3K/Akt is independent of both mTOR and GSK3, but requires the activation of EGF receptor (EGFR). Nuclear levels of Sp1 are strongly reduced by inhibition of PI3K/Akt pathway, and blockade of Sp1-dependent transcription with mithramycin A prevents upregulation of P2rx7 gene expression following serum withdrawal. Furthermore, atypical PKCζ plays a key role in the regulation of P2X7R expression by preventing phosphorylation and, consequently, activation of Akt. Altogether, these data indicate that activation of EGFR enhanced the expression of P2X7R in neuroblastoma cells lacking trophic support, being PI3K/Akt/PKCζ signaling pathway and Sp1 mediating this pro-survival outcome. PMID:26687764

  13. Quercetin Protects against Okadaic Acid-Induced Injury via MAPK and PI3K/Akt/GSK3β Signaling Pathways in HT22 Hippocampal Neurons.

    PubMed

    Jiang, Wei; Luo, Tao; Li, Sheng; Zhou, Yue; Shen, Xiu-Yin; He, Feng; Xu, Jie; Wang, Hua-Qiao

    2016-01-01

    Increasing evidence shows that oxidative stress and the hyperphosphorylation of tau protein play essential roles in the progression of Alzheimer's disease (AD). Quercetin is a major flavonoid that has anti-oxidant, anti-cancer and anti-inflammatory properties. We investigated the neuroprotective effects of quercetin to HT22 cells (a cell line from mouse hippocampal neurons). We found that Okadaic acid (OA) induced the hyperphosphorylation of tau protein at Ser199, Ser396, Thr205, and Thr231 and produced oxidative stress to the HT22 cells. The oxidative stress suppressed the cell viability and decreased the levels of lactate dehydrogenase (LDH), superoxide dismutase (SOD), mitochondria membrane potential (MMP) and Glutathione peroxidase (GSH-Px). It up-regulated malondialdehyde (MDA) production and intracellular reactive oxygen species (ROS). In addition, phosphoinositide 3 kinase/protein kinase B/Glycogen synthase kinase3β (PI3K/Akt/GSK3β) and mitogen activated protein kinase (MAPK) were also involved in this process. We found that pre-treatment with quercetin can inhibited OA-induced the hyperphosphorylation of tau protein and oxidative stress. Moreover, pre-treatment with quercetin not only inhibited OA-induced apoptosis via the reduction of Bax, and up-regulation of cleaved caspase 3, but also via the inhibition of PI3K/Akt/GSK3β, MAPKs and activation of NF-κB p65. Our findings suggest the therapeutic potential of quercetin to treat AD. PMID:27050422

  14. Nobiletin, a citrus flavonoid, suppresses invasion and migration involving FAK/PI3K/Akt and small GTPase signals in human gastric adenocarcinoma AGS cells.

    PubMed

    Lee, Yi-Chieh; Cheng, Tsan-Hwang; Lee, Jung-Shin; Chen, Jiun-Hwan; Liao, Yi-Chen; Fong, Yao; Wu, Cheng-Hsun; Shih, Yuan-Wei

    2011-01-01

    Nobiletin, a compound isolated from citrus fruits, is a polymethoxylated flavone derivative shown to have anti-inflammatory, antitumor, and neuroprotective properties. This study has investigated that nobiletin exerted inhibitory effects on the cell adhesion, invasion, and migration abilities of a highly metastatic AGS cells under non-cytotoxic concentrations. Data also showed nobiletin could inhibit the activation of focal adhesion kinase (FAK) and phosphoinositide-3-kinase/Akt (PI3K/Akt) involved in the downregulation of the enzyme activities, protein expressions, messenger RNA levels of matrix metalloproteinase-2 (MMP-2), and matrix metalloproteinase-2 (MMP-9). Also, our data revealed that nobiletin inhibited FAK/PI3K/Akt with concurrent reduction in the protein expressions of Ras, c-Raf, Rac-1, Cdc42, and RhoA by western blotting, whereas the protein level of RhoB increased progressively. Otherwise, nobiletin-treated AGS cells showed tremendously decreased in the phosphorylation and degradation of inhibitor of kappaBα (IκBα), the nuclear level of NF-κB, and the binding ability of NF-κB to NF-κB response element. Furthermore, nobiletin significantly decreased the levels of phospho-Akt and MMP-2/9 in Akt1-cDNA-transfected cells concomitantly with a marked reduction in cell invasion and migration. These results suggest that nobiletin can reduce invasion and migration of AGS cells, and such a characteristic may be of great value in the development of a potential cancer therapy.

  15. Quercetin Protects against Okadaic Acid-Induced Injury via MAPK and PI3K/Akt/GSK3β Signaling Pathways in HT22 Hippocampal Neurons

    PubMed Central

    Li, Sheng; Zhou, Yue; Shen, Xiu-Yin; He, Feng; Xu, Jie; Wang, Hua-Qiao

    2016-01-01

    Increasing evidence shows that oxidative stress and the hyperphosphorylation of tau protein play essential roles in the progression of Alzheimer’s disease (AD). Quercetin is a major flavonoid that has anti-oxidant, anti-cancer and anti-inflammatory properties. We investigated the neuroprotective effects of quercetin to HT22 cells (a cell line from mouse hippocampal neurons). We found that Okadaic acid (OA) induced the hyperphosphorylation of tau protein at Ser199, Ser396, Thr205, and Thr231 and produced oxidative stress to the HT22 cells. The oxidative stress suppressed the cell viability and decreased the levels of lactate dehydrogenase (LDH), superoxide dismutase (SOD), mitochondria membrane potential (MMP) and Glutathione peroxidase (GSH-Px). It up-regulated malondialdehyde (MDA) production and intracellular reactive oxygen species (ROS). In addition, phosphoinositide 3 kinase/protein kinase B/Glycogen synthase kinase3β (PI3K/Akt/GSK3β) and mitogen activated protein kinase (MAPK) were also involved in this process. We found that pre-treatment with quercetin can inhibited OA-induced the hyperphosphorylation of tau protein and oxidative stress. Moreover, pre-treatment with quercetin not only inhibited OA-induced apoptosis via the reduction of Bax, and up-regulation of cleaved caspase 3, but also via the inhibition of PI3K/Akt/GSK3β, MAPKs and activation of NF-κB p65. Our findings suggest the therapeutic potential of quercetin to treat AD. PMID:27050422

  16. 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.

  17. Furano-1,2-Naphthoquinone Inhibits Src and PI3K/Akt Signaling Pathways in Ca9-22 Human Oral Squamous Carcinoma Cells.

    PubMed

    Lin, Kuei-Li; Chien, Ching-Ming; Tseng, Chih-Hua; Chen, Yeh-Long; Chang, Long-Sen; Lin, Shinne-Ren

    2014-05-01

    Furano-1,2-naphthoquinone (FNQ), a biologically active component ofAvicennia marina, has been demonstrated to display anticancer activity. FNQ exerted cytotoxicity with the G2/M cell cycle arrest and apoptosis in Ca9-22 cells. FNQ-induced G2/M arrest was correlated with a marked decrease in the expression levels of cyclin A and cyclin B, and their activating partner cyclin-dependent kinases (CDK) 1 and 2 with concomitant induction of p27. FNQ-induced apoptosis was accompanied by Bax and Bad upregulation, and the downregulation of Bcl-2, Bcl-XL, Mcl-1, and X-linked inhibitor of apoptosis (XIAP), resulting in cytochrome C release and sequential activation of caspase-9 and caspase-3. Mechanistic studies showed that FNQ suppressed Src phosphorylation, PI3K, and Akt activation in Ca9-22 cells. Moreover, the Src inhibitor PP2 reduced the phosphorylation of Src and activation of PI3K/Akt, which was comparable with FNQ treatment. The combined treatment of FNQ with PP2 enhanced the cell cycle arrest and apoptosis and also led to the downregulation of Bcl-XL, Mcl-1, XIAP, cyclin A, cyclin B, CDK1, and CDK2 and upregulation of p27, Bax, and Bad. These findings suggest that FNQ-mediated cytotoxicity of Ca9-22 cells is related with the G2/M cell cycle arrest and apoptosis via inactivation of Src and PI3K/Akt-mediated signaling pathways. PMID:22505597

  18. CCL5 promotes VEGF-dependent angiogenesis by down-regulating miR-200b through PI3K/Akt signaling pathway in human chondrosarcoma cells

    PubMed Central

    Liu, Guan-Ting; Chen, Hsien-Te; Tsou, Hsi-Kai; Tan, Tzu-Wei; Fong, Yi-Chin; Chen, Po-Chen; Yang, Wei-Hung; Wang, Shih-Wei; Chen, Jui-Chieh; Tang, Chih-Hsin

    2014-01-01

    Chondrosarcoma is the second most common primary malignant bone cancer, with potential for local invasion and distant metastasis. Chemokine CCL5 (formerly RANTES) of the CC-chemokine family plays a crucial role in metastasis. Angiogenesis is essential for the cancer metastasis. However, correlation of CCL5 with vascular endothelial growth factor (VEGF) expression and angiogenesis in human chondrosarcoma is still unknown. CCL5-mediated VEGF expression was assessed by qPCR, ELISA, and Western blotting. CCL5-induced angiogenesis was examined by migration and tube formation in endothelial progenitor cells in vitro. CCL5 increased VEGF expression and also promoted chondrosarcoma conditional medium-mediated angiogenesis in vitro and in vivo. Stimulation of chondrosarcoma with CCL5 augmented PI3K and Akt phosphorylation, while PI3K and Akt inhibitor or siRNA abolished CCL5-induced VEGF expression and angiogenesis. We also demonstrated CCL5 inhibiting miR-200b expression and miR-200b mimic reversing the CCL5-enhanced VEGF expression and angiogenesis. Moreover, in chondrosarcoma patients showed the positive correlation between CCL5 and VEGF; negative correlation between CCL5 and miR-200b. Taken together, results demonstrate CCL5 promoting VEGF-dependent angiogenesis in human chondrosarcoma cells by down-regulating miR-200b through PI3K/Akt signaling pathway. PMID:25301739

  19. 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

  20. Resveratrol induces cell cycle arrest in human gastric cancer MGC803 cells via the PTEN-regulated PI3K/Akt signaling pathway.

    PubMed

    Jing, Xiaoping; Cheng, Weiwei; Wang, Shiying; Li, Pin; He, Li

    2016-01-01

    Resveratrol is a polyphenolic compound that is extracted from Polygonum cuspidatum and is used in traditional Chinese medicine. Previous data have shown that resveratrol inhibits the growth of human gastric cancer. MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] and trypan blue assays showed that resveratrol significantly decreased the survival rate of MGC803 cells in a concentration- and time-dependent manner. Our flow cytometric analysis showed that resveratrol treatment arrested the cells at the G0/G1 phase of the cell cycle. Furthermore, western blotting demonstrated that resveratrol decreased the protein expression of phospho-glycogen synthase kinase 3β (p-GSK3β), cyclin D1, phospho-phosphatase and tensin homologue (p-PTEN), phospho-phosphatidylinositol 3'-OH kinase (p-PI3K), and phospho-protein kinase B (p-PKB/Akt). We also found that resveratrol inhibited the progression of the cell cycle in MGC803 cells by repressing p-PI3K and p-Akt expression. Meanwhile, resveratrol did not decrease the phosphorylation level of Akt when the PTEN gene expression was knocked down by an siRNA in the MGC803 cells. Taken together, these results suggest that resveratrol induced cell cycle arrest in human gastric cancer MGC803 cells by regulating the PTEN/PI3K/Akt signaling pathway.

  1. Resveratrol induces cell cycle arrest in human gastric cancer MGC803 cells via the PTEN-regulated PI3K/Akt signaling pathway.

    PubMed

    Jing, Xiaoping; Cheng, Weiwei; Wang, Shiying; Li, Pin; He, Li

    2016-01-01

    Resveratrol is a polyphenolic compound that is extracted from Polygonum cuspidatum and is used in traditional Chinese medicine. Previous data have shown that resveratrol inhibits the growth of human gastric cancer. MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide] and trypan blue assays showed that resveratrol significantly decreased the survival rate of MGC803 cells in a concentration- and time-dependent manner. Our flow cytometric analysis showed that resveratrol treatment arrested the cells at the G0/G1 phase of the cell cycle. Furthermore, western blotting demonstrated that resveratrol decreased the protein expression of phospho-glycogen synthase kinase 3β (p-GSK3β), cyclin D1, phospho-phosphatase and tensin homologue (p-PTEN), phospho-phosphatidylinositol 3'-OH kinase (p-PI3K), and phospho-protein kinase B (p-PKB/Akt). We also found that resveratrol inhibited the progression of the cell cycle in MGC803 cells by repressing p-PI3K and p-Akt expression. Meanwhile, resveratrol did not decrease the phosphorylation level of Akt when the PTEN gene expression was knocked down by an siRNA in the MGC803 cells. Taken together, these results suggest that resveratrol induced cell cycle arrest in human gastric cancer MGC803 cells by regulating the PTEN/PI3K/Akt signaling pathway. PMID:26530632

  2. α-Lipoic Acids Promote the Protein Synthesis of C2C12 Myotubes by the TLR2/PI3K Signaling Pathway.

    PubMed

    Jing, Yuanyuan; Cai, Xingcai; Xu, Yaqiong; Zhu, Canjun; Wang, Lina; Wang, Songbo; Zhu, Xiaotong; Gao, Ping; Zhang, Yongliang; Jiang, Qingyan; Shu, Gang

    2016-03-01

    Skeletal muscle protein turnover is regulated by endocrine hormones, nutrients, and inflammation. α-Lipoic acid (ALA) plays an important role in energy homeostasis. Therefore, the aim of this study was to investigate the effects of ALA on protein synthesis in skeletal muscles and reveal the underlying mechanism. ALA (25 μM) significantly increased the protein synthesis and phosphorylation of Akt, mTOR, and S6 in C2C12 myotubes with attenuated phosphorylation of AMPK, Ikkα/β, and eIF2α. Intraperitoneal injection of 50 mg/kg ALA also produced the same results in mouse gastrocnemius. Both the PI3K (LY294002) and mTOR (rapamycin) inhibitors abolished the effects of ALA on protein synthesis in the C2C12 myotubes. However, AICAR (AMPK agonist) failed to block the activation of mTOR and S6 by ALA. ALA increased TLR2 and MyD88 mRNA expression in the C2C12 myotubes. TLR2 knockdown by siRNA almost eliminated the effects of ALA on protein synthesis and the Akt/mTOR pathway in the C2C12 myotubes. Immunoprecipitation data showed that ALA enhanced the p85 subunit of PI3K binding to MyD88. These findings indicate that ALA induces protein synthesis and the PI3K/Akt signaling pathway by TLR2.

  3. Cannabinoid receptor agonist WIN55,212-2 and fatty acid amide hydrolase inhibitor URB597 may protect against cognitive impairment in rats of chronic cerebral hypoperfusion via PI3K/AKT signaling.

    PubMed

    Su, Shao-Hua; Wang, Yue-Qing; Wu, Yi-Fang; Wang, Da-Peng; Lin, Qi; Hai, Jian

    2016-10-15

    The present study further investigated the protective effects of cannabinoid receptor agonist WIN55,212-2 (WIN) and fatty acid amide hydrolase (FAAH) inhibitor URB597 (URB) on chronic cerebral hypoperfusion (CCH)-induced cognitive impairment in rats. Spatial learning and memory were assessed with the Morris water maze and by measuring Long-term potentiation. The expression of microtubule-associated protein-2 (MAP)-2, growth-associated protein-43 (GAP)-43, synaptophysin, cannabinoid receptor 1 (CB1), brain-derived neurotrophic factor (BDNF), FAAH, N-acylphosphatidylethanolamine phospholipase D(NAPE-PLD) and monoacyl glycerol lipase (MGL) as well as phosphoinositide 3-kinase (PI3K)/AKT signaling pathway molecules and downstream targets including AKT, phosphorylated (p-)AKT, cyclic AMP response element- binding protein (CREB), p-CREB, Bcl-2-associated death protein (BAD), p-BAD, glycogen synthase kinase (GSK)-3β, p-GSK-3β, forkhead box protein (FOXO) 3A and p-FOXO3A was determined by western blotting. WIN and URB treatment improved learning and memory performance, effects that were abolished by co-administration of the PI3K/AKT inhibitor LY294002. Moreover, WIN and URB reversed the decreases in MAP-2 and synaptophysin expression resulting from CCH, and stimulated BDNF and CB1 expression as well as CREB, FOXO3A, GSK-3β, and BAD phosphorylation, confirming that WIN and URB mediate neuroprotection by preventing neuronal apoptosis and improving cognition via PI3K/AKT signaling. These findings suggest that WIN and URB are promising agents for therapeutic management of CCH. PMID:27424778

  4. 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

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

    PubMed

    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

  6. 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

    2014-09-01

    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.

  7. Gedunin abrogates aldose reductase, PI3K/Akt/mToR, and NF-κB signaling pathways to inhibit angiogenesis in a hamster model of oral carcinogenesis.

    PubMed

    Kishore T, Kranthi Kiran; Ganugula, Raghu; Gade, Deepak Reddy; Reddy, Geereddy Bhanuprakash; Nagini, Siddavaram

    2016-02-01

    Aberrant activation of oncogenic signaling pathways plays a central role in tumor development and progression. The aim of this present study was to investigate the chemopreventive effects of the neem limonoid gedunin in the hamster model of oral cancer based on its ability to modulate aldose reductase (AR), phosphatidyl inositol-3-kinase (PI3K)/Akt, and nuclear factor kappa B (NF-κB) pathways to block angiogenesis. Administration of gedunin suppressed the development of HBP carcinomas by inhibiting PI3K/Akt and NF-κB pathways through the inactivation of Akt and inhibitory kappa B kinase (IKK), respectively. Immunoblot and molecular docking interactions revealed that inhibition of these signaling pathways may be mediated via inactivation of AR by gedunin. Gedunin blocked angiogenesis by downregulating the expression of miR-21 and the pro-angiogenic factors vascular endothelial growth factor and hypoxia inducible factor-1 alpha (HIF-1α). In conclusion, the results of the present study provide compelling evidence that gedunin prevents progression of hamster buccal pouch (HBP) carcinomas via inhibition of the kinases Akt, IKK, and AR, and the oncogenic transcription factors NF-κB and HIF-1α to block angiogenesis. PMID:26342697

  8. Osthole shows the potential to overcome P-glycoprotein‑mediated multidrug resistance in human myelogenous leukemia K562/ADM cells by inhibiting the PI3K/Akt signaling pathway.

    PubMed

    Wang, Hong; Jia, Xiu-Hong; Chen, Jie-Ru; Wang, Jian-Yong; Li, You-Jie

    2016-06-01

    P-glycoprotein (P-gp)-mediated multidrug resistance (MDR) has been reported to play a pivotal role in tumor chemotherapy failure. Study after study has illustrated that the phosphoinositide 3-kinase (PI3K)/Akt signaling cascade is involved in the MDR phenotype and is correlated with P-gp expression in many human malignancies. In the present study, osthole, an O-methylated coumarin, exhibited potent reversal capability of MDR in myelogenous leukemia K562/ADM cells. Simultaneously, the uptake and efflux of Rhodamine-123 (Rh-123) and the accumulation of doxorubicin assays combined with flow cytometric analysis suggested that osthole could increase intracellular drug accumulation. Furthermore, osthole decreased the expression of multidrug resistance gene 1 (MDR1) at both the mRNA and protein levels. Further experiments elucidated that osthole could suppress P-gp expression by inhibiting the PI3K/Akt signaling pathway which might be the main mechanism accounting for the reversal potential of osthole in the MDR in K562/ADM cells. In conclusion, osthole combats MDR and could be a promising candidate for the development of novel MDR reversal modulators. PMID:27109742

  9. RhoC promotes human melanoma invasion in a PI3K/Akt-dependent pathway.

    PubMed

    Ruth, Mariah C; Xu, Yisheng; Maxwell, Ian H; Ahn, Natalie G; Norris, David A; Shellman, Yiqun G

    2006-04-01

    Overexpression of the small GTPase, RhoC, in various human cancers has been correlated with high metastatic ability and poor prognosis. Rho-kinase (ROCK) is an important effector of Rho GTPases. The oncogenic serine/threonine kinase Akt (also known as PKB) is a downstream effector of phosphatidylinositol-3 kinase (PI3K). Akt activation contributes to the neoplastic phenotype by promoting cell cycle progression, increasing antiapoptotic functions, and enhancing tumor cell invasion. Rho signaling via ROCK has been previously shown either to activate or to downregulate PI3K/Akt. Using a human radial growth phase melanoma cell line, WM35, we have established stable transfectants that overexpress RhoC (called WM35RhoC). We found that overexpression of RhoC increased phosphorylated-Akt (Ser473/474/472, pAkt) expression and promoted cell invasion. Inhibition of RhoC with C3 transferase downregulated pAkt expression and decreased cell invasion in these cells. In addition, inhibition of PI3K, Akt, or ROCK partially decreased invasion. Further, inhibition of PI3K but not ROCK decreased the pAkt level. These results suggest that RhoC promotes invasion in part via activation of a PI3K/Akt pathway, in a manner independent of ROCK signaling. We propose that RhoC promotes melanoma progression via separate mechanisms that regulate the PI3K/Akt pathway and the ROCK signaling pathway.

  10. Isoorientin induces apoptosis through mitochondrial dysfunction and inhibition of PI3K/Akt signaling pathway in HepG2 cancer cells

    SciTech Connect

    Yuan, Li; Wang, Jing; Xiao, Haifang; Xiao, Chunxia; Wang, Yutang; Liu, Xuebo

    2012-11-15

    Isoorientin (ISO) is a flavonoid compound that can be extracted from several plant species, such as Phyllostachys pubescens, Patrinia, and Drosophyllum lusitanicum; however, its biological activity remains poorly understood. The present study investigated the effects and putative mechanism of apoptosis induced by ISO in human hepatoblastoma cancer (HepG2) cells. The results showed that ISO induced cell death in a dose-dependent manner in HepG2 cells, but no toxicity in human liver cells (HL-7702) and buffalo rat liver cells (BRL-3A) treated with ISO at the indicated concentrations. ISO-induced cell death included apoptosis which characterized by the appearance of nuclear shrinkage, the cleavage of poly (ADP-ribose) polymerase (PARP) and DNA fragmentation. ISO significantly (p < 0.01) increased the Bax/Bcl-2 ratio, disrupted the mitochondrial membrane potential (MMP), increased the release of cytochrome c, activated caspase-3, and enhanced intracellular levels of reactive oxygen species (ROS) and nitric oxide (NO). In addition, ISO effectively inhibited the phosphorylation of Akt and increased FoxO4 expression. The PI3K/Akt inhibitor LY294002 enhanced the apoptosis-inducing effect of ISO. However, LY294002 markedly quenched ROS and NO generation and diminished the protein expression of heme peroxidase enzyme (HO-1) and inducible nitric oxide synthase (iNOS). Furthermore, the addition of a ROS inhibitor (N-acetyl cysteine, NAC) or iNOS inhibitor (N-[3-(aminomethyl) benzyl] acetamidine, dihydrochloride, 1400W) significantly diminished the apoptosis induced by ISO and also blocked the phosphorylation of Akt. These results demonstrated for the first time that ISO induces apoptosis in HepG2 cells and indicate that this apoptosis might be mediated through mitochondrial dysfunction and PI3K/Akt signaling pathway, and has no toxicity in normal liver cells, suggesting that ISO may have good potential as a therapeutic and chemopreventive agent for liver cancer. Highlights:

  11. Ras-related C3 Botulinum Toxin Substrate (Rac) and Src Family Kinases (SFK) Are Proximal and Essential for Phosphatidylinositol 3-Kinase (PI3K) Activation in Natural Killer (NK) Cell-mediated Direct Cytotoxicity against Cryptococcus neoformans.

    PubMed

    Xiang, Richard F; Stack, Danuta; Huston, Shaunna M; Li, Shu Shun; Ogbomo, Henry; Kyei, Stephen K; Mody, Christopher H

    2016-03-25

    The activity of Rac in leukocytes is essential for immunity. However, its role in NK cell-mediated anti-microbial signaling remains unclear. In this study, we investigated the role of Rac in NK cell mediated anti-cryptococcal killing. We found thatCryptococcus neoformansindependently activates both Rac and SFK pathways in NK cells, and unlike in tumor killing,Cryptococcusinitiated a novel Rac → PI3K → Erk cytotoxicity cascade. Remarkably, Rac was not required for conjugate formation, despite its essential role in NK cytotoxicity againstC. neoformans Taken together, our data show that, unlike observations with tumor cells, NK cells use a novel Rac cytotoxicity pathway in conjunction with SFK, to killC. neoformans. PMID:26867574

  12. Ras-related C3 Botulinum Toxin Substrate (Rac) and Src Family Kinases (SFK) Are Proximal and Essential for Phosphatidylinositol 3-Kinase (PI3K) Activation in Natural Killer (NK) Cell-mediated Direct Cytotoxicity against Cryptococcus neoformans.

    PubMed

    Xiang, Richard F; Stack, Danuta; Huston, Shaunna M; Li, Shu Shun; Ogbomo, Henry; Kyei, Stephen K; Mody, Christopher H

    2016-03-25

    The activity of Rac in leukocytes is essential for immunity. However, its role in NK cell-mediated anti-microbial signaling remains unclear. In this study, we investigated the role of Rac in NK cell mediated anti-cryptococcal killing. We found thatCryptococcus neoformansindependently activates both Rac and SFK pathways in NK cells, and unlike in tumor killing,Cryptococcusinitiated a novel Rac → PI3K → Erk cytotoxicity cascade. Remarkably, Rac was not required for conjugate formation, despite its essential role in NK cytotoxicity againstC. neoformans Taken together, our data show that, unlike observations with tumor cells, NK cells use a novel Rac cytotoxicity pathway in conjunction with SFK, to killC. neoformans.

  13. Nerve growth factor (NGF) regulates activity of nuclear factor of activated T-cells (NFAT) in neurons via the phosphatidylinositol 3-kinase (PI3K)-Akt-glycogen synthase kinase 3β (GSK3β) pathway.

    PubMed

    Kim, Man-Su; Shutov, Leonid P; Gnanasekaran, Aswini; Lin, Zhihong; Rysted, Jacob E; Ulrich, Jason D; Usachev, Yuriy M

    2014-11-01

    The Ca(2+)/calcineurin-dependent transcription factor nuclear factor of activated T-cells (NFAT) plays an important role in regulating many neuronal functions, including excitability, axonal growth, synaptogenesis, and neuronal survival. NFAT can be activated by action potential firing or depolarization that leads to Ca(2+)/calcineurin-dependent dephosphorylation of NFAT and its translocation to the nucleus. Recent data suggest that NFAT and NFAT-dependent functions in neurons can also be potently regulated by NGF and other neurotrophins. However, the mechanisms of NFAT regulation by neurotrophins are not well understood. Here, we show that in dorsal root ganglion sensory neurons, NGF markedly facilitates NFAT-mediated gene expression induced by mild depolarization. The effects of NGF were not associated with changes in [Ca(2+)]i and were independent of phospholipase C activity. Instead, the facilitatory effect of NGF depended on activation of the PI3K/Akt pathway downstream of the TrkA receptor and on inhibition of glycogen synthase kinase 3β (GSK3β), a protein kinase known to phosphorylate NFAT and promote its nuclear export. Knockdown or knockout of NFATc3 eliminated this facilitatory effect. Simultaneous monitoring of EGFP-NFATc3 nuclear translocation and [Ca(2+)]i changes in dorsal root ganglion neurons indicated that NGF slowed the rate of NFATc3 nuclear export but did not affect its nuclear import rate. Collectively, our data suggest that NGF facilitates depolarization-induced NFAT activation by stimulating PI3K/Akt signaling, inactivating GSK3β, and thereby slowing NFATc3 export from the nucleus. We propose that NFAT serves as an integrator of neurotrophin action and depolarization-driven calcium signaling to regulate neuronal gene expression.

  14. Medroxyprogestogen enhances apoptosis of SKOV-3 cells via inhibition of the PI3K/Akt signaling pathway

    PubMed Central

    Li, Yan; Jiang, Yi; Wan, Yicong; Zhang, Lin; Tang, Weiwei; Ma, Jingjing; Wu, Shan; Cheng, Wenjun

    2013-01-01

    We sought to assess the effect of progestin on the apoptosis of epithelial ovarian cancer cell line SKOV-3 and via regulation of phosphorylation signaling in. Epithelial ovarian cancer cell line SKOV-3 was treated with medroxyprogestogen, phosphatidylinositol 3-kinase inhibitor LY294002 and vehicle control. Akt, phospho-Akt, Bcl-2 and phospho-Bad proteins were examined by immunoblotting assays. Medroxyprogestogen-induced apoptosis was assessed by MTT assays and Annexin V apoptosis assay. We found no significant difference in Akt and Bad expression in both the medroxyprogestogen groups and the control group. The levels of phospho-Akt, Bcl-2 and phospho-Bad were decreased in all the medroxyprogestogen groups and significantly decreased in the high dose mitogen-activated protein (MAP) group (10 µmol/L). Viability of SKOV-3 was reduced and apparent apoptosis of SKOV-3 cells was observed with increased doses of MAP. The findings suggest that medroxyprogestogen can induce SKOV-3 cell apoptosis by inhibiting Akt phosphorylation. PMID:23554793

  15. Dryofragin inhibits the migration and invasion of human osteosarcoma U2OS cells by suppressing MMP-2/9 and elevating TIMP-1/2 through PI3K/AKT and p38 MAPK signaling pathways.

    PubMed

    Su, Yan; Wan, Daqian; Song, Wenqi

    2016-08-01

    Dryofragin, a phloroglucinol derivative extracted from Dryopteris fragrans (L.) Schott, was found to inhibit proliferation and induce apoptosis of tumor cells. However, the mechanism involved in the suppression of cancer cell metastasis by dryofragin remains unclear. Our study investigated the mechanisms for the antitumor properties of dryofragin on the migration and invasion of human osteosarcoma U2OS cells. Dryofragin suppressed the migration and invasive ability of U2OS cells, and it decreased the expression of MMP-2 and MMP-9 and elevated the expression of TIMP-1 and TIMP-2. Western blotting assays indicated that dryofragin was effective in suppressing the phosphorylation of phosphatidylinositide-3 kinase (PI3K), Akt, and p38 MAPK. These results suggest that dryofragin inhibited U2OS cell migration and invasion by reducing the expression of MMP-2 and MMP-9 and elevating the expression of TIMP-1 and TIMP-2 through the PI3K/AKT and p38 MAPK signaling pathways. Above all, we conclude that dryofragin represents an anti-invasive agent and may potentially be applicable in osteosarcoma therapy. PMID:27243922

  16. Drosophila Spidey/Kar Regulates Oenocyte Growth via PI3-Kinase Signaling

    PubMed Central

    Cinnamon, Einat; Sawala, Annick; Tittiger, Claus; Paroush, Ze'ev

    2016-01-01

    Cell growth and proliferation depend upon many different aspects of lipid metabolism. One key signaling pathway that is utilized in many different anabolic contexts involves Phosphatidylinositide 3-kinase (PI3K) and its membrane lipid products, the Phosphatidylinositol (3,4,5)-trisphosphates. It remains unclear, however, which other branches of lipid metabolism interact with the PI3K signaling pathway. Here, we focus on specialized fat metabolizing cells in Drosophila called larval oenocytes. In the presence of dietary nutrients, oenocytes undergo PI3K-dependent cell growth and contain very few lipid droplets. In contrast, during starvation, oenocytes decrease PI3K signaling, shut down cell growth and accumulate abundant lipid droplets. We now show that PI3K in larval oenocytes, but not in fat body cells, functions to suppress lipid droplet accumulation. Several enzymes of fatty acid, triglyceride and hydrocarbon metabolism are required in oenocytes primarily for lipid droplet induction rather than for cell growth. In contrast, a very long chain fatty-acyl-CoA reductase (FarO) and a putative lipid dehydrogenase/reductase (Spidey, also known as Kar) not only promote lipid droplet induction but also inhibit oenocyte growth. In the case of Spidey/Kar, we show that the growth suppression mechanism involves inhibition of the PI3K signaling pathway upstream of Akt activity. Together, the findings in this study show how Spidey/Kar and FarO regulate the balance between the cell growth and lipid storage of larval oenocytes. PMID:27500738

  17. Drosophila Spidey/Kar Regulates Oenocyte Growth via PI3-Kinase Signaling.

    PubMed

    Cinnamon, Einat; Makki, Rami; Sawala, Annick; Wickenberg, Leah P; Blomquist, Gary J; Tittiger, Claus; Paroush, Ze'ev; Gould, Alex P

    2016-08-01

    Cell growth and proliferation depend upon many different aspects of lipid metabolism. One key signaling pathway that is utilized in many different anabolic contexts involves Phosphatidylinositide 3-kinase (PI3K) and its membrane lipid products, the Phosphatidylinositol (3,4,5)-trisphosphates. It remains unclear, however, which other branches of lipid metabolism interact with the PI3K signaling pathway. Here, we focus on specialized fat metabolizing cells in Drosophila called larval oenocytes. In the presence of dietary nutrients, oenocytes undergo PI3K-dependent cell growth and contain very few lipid droplets. In contrast, during starvation, oenocytes decrease PI3K signaling, shut down cell growth and accumulate abundant lipid droplets. We now show that PI3K in larval oenocytes, but not in fat body cells, functions to suppress lipid droplet accumulation. Several enzymes of fatty acid, triglyceride and hydrocarbon metabolism are required in oenocytes primarily for lipid droplet induction rather than for cell growth. In contrast, a very long chain fatty-acyl-CoA reductase (FarO) and a putative lipid dehydrogenase/reductase (Spidey, also known as Kar) not only promote lipid droplet induction but also inhibit oenocyte growth. In the case of Spidey/Kar, we show that the growth suppression mechanism involves inhibition of the PI3K signaling pathway upstream of Akt activity. Together, the findings in this study show how Spidey/Kar and FarO regulate the balance between the cell growth and lipid storage of larval oenocytes. PMID:27500738

  18. CHRNA7 inhibits cell invasion and metastasis of LoVo human colorectal cancer cells through PI3K/Akt signaling.

    PubMed

    Xiang, Tao; Yu, Feng; Fei, Rushan; Qian, Jing; Chen, Wenbin

    2016-02-01

    The α7 neuronal nicotinic receptor gene (CHRNA7) is widely expressed in both the brain and periphery whereas its encoding protein of α7 neuronal acetylcholine receptor (α7nAChR) belongs to the nicotinic acetylcholine receptor family. Considerable evidence suggests that α7nAChR plays an important role in chronic inflammatory and neuropathic pain signaling and thus has been proposed as a potential target for treating cognitive deficits in patients with schizophrenia, attention deficit hyperactivity disorder (ADHD) and Alzheimer's disease. The aim of the present study was to determine the role of endogenous α7nAChR signaling in human colorectal cancer growth and metastasis. pLVX‑CHRNA7 encoding the full length of CHRNA7 was constructed and transfected into LoVo human colorectal cancer cells. Cell proliferation was measured by Cell Counting Kit‑8 (CCK‑8), and cell migration and invasion were detected by Transwell chamber assays. Expression and activity of metastasis‑related metalloproteinases (MMPs) were analyzed by western blotting and gelatin zymography, respectively. Activation of metastasis-related signaling molecules was detected by western blotting. LY294002 was used to specifically block the phosphatidylinositol 3‑kinase/v‑akt murine thymoma viral oncogene homologue (PI3K/Akt) pathway. We showed that concomitantly with an increase in α7nAChR expression after transfection, LoVo cells presented reduced abilities for migration and invasion, which was accompanied by reduced expression levels of MMP‑1 and MMP‑9 as well as activation of the PI3K/Akt signaling pathway. The application of LY294002 restored the migration and invasion abilities of the LoVo cells bearing CHRNA7. Collectively, we conclude that overexpression of CHRNA7 negatively controls colorectal cancer LoVo cell invasion and metastasis via PI3K/Akt pathway activation and may serve as either a diagnostic marker or a therapeutic target for colorectal cancer metastasis.

  19. Hypoactivity Affects IGF-1 Level and PI3K/AKT Signaling Pathway in Cerebral Structures Implied in Motor Control

    PubMed Central

    Mysoet, Julien; Canu, Marie-Hélène; Cieniewski-Bernard, Caroline; Bastide, Bruno; Dupont, Erwan

    2014-01-01

    A chronic reduction in neuromuscular activity through prolonged body immobilization in human alters motor task performance through a combination of peripheral and central factors. Studies performed in a rat model of sensorimotor restriction have shown functional and biochemical changes in sensorimotor cortex. However, the underlying mechanisms are still unclear. Interest was turned towards a possible implication of Insulin-like Growth Factor 1 (IGF-1), a growth factor known to mediate neuronal excitability and synaptic plasticity by inducing phosphorylation cascades which include the PI3K–AKT pathway. In order to better understand the influence of IGF-1 in cortical plasticity in rats submitted to a sensorimotor restriction, we analyzed the effect of hindlimb unloading on IGF-1 and its main molecular pathway in structures implied in motor control (sensorimotor cortex, striatum, cerebellum). IGF-1 level was determined by ELISA, and phosphorylation of its receptor and proteins of the PI3K–AKT pathway by immunoblot. In the sensorimotor cortex, our results indicate that HU induces a decrease in IGF-1 level; this alteration is associated to a decrease in activation of PI3K-AKT pathway. The same effect was observed in the striatum, although to a lower extent. No variation was noticed in the cerebellum. These results suggest that IGF-1 might contribute to cortical and striatal plasticity induced by a chronic sensorimotor restriction. PMID:25226394

  20. 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

  1. Camptothecin inhibits platelet-derived growth factor-BB-induced proliferation of rat aortic vascular smooth muscle cells through inhibition of PI3K/Akt signaling pathway

    SciTech Connect

    Park, Eun-Seok; Kang, Shin-il; Yoo, Kyu-dong; Lee, Mi-Yea; Yoo, Hwan-Soo; Hong, Jin-Tae; Shin, Hwa-Sup; Kim, Bokyung; Yun, Yeo-Pyo

    2013-04-15

    The abnormal proliferation of vascular smooth muscle cells (VSMCs) in arterial wall is a major cause of vascular disorders such as atherosclerosis and restenosis after angioplasty. In this study, we investigated not only the inhibitory effects of camptothecin (CPT) on PDGF-BB-induced VSMC proliferation, but also its molecular mechanism of this inhibition. CPT significantly inhibited proliferation with IC50 value of 0.58 μM and the DNA synthesis of PDGF-BB-stimulated VSMCs in a dose-dependent manner (0.5–2 μM ) without any cytotoxicity. CPT induced the cell cycle arrest at G0/G1 phase. Also, CPT decreased the expressions of G0/G1-specific regulatory proteins including cyclin-dependent kinase (CDK)2, cyclin D1 and PCNA in PDGF-BB-stimulated VSMCs. Pre-incubation of VSMCs with CPT significantly inhibited PDGF-BB-induced Akt activation, whereas CPT did not affect PDGF-receptor beta phosphorylation, extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and phospholipase C (PLC)-γ1 phosphorylation in PDGF-BB signaling pathway. Our data showed that CPT pre-treatment inhibited VSMC proliferation, and that the inhibitory effect of CPT was enhanced by LY294002, a PI3K inhibitor, on PDGF-BB-induced VSMC proliferation. In addition, inhibiting the PI3K/Akt pathway by LY294002 significantly enhanced the suppression of PCNA expression and Akt activation by CPT. These results suggest that the anti-proliferative activity of CPT is mediated in part by downregulating the PI3K/Akt signaling pathway. - Highlights: ► CPT inhibits proliferation of PDGF-BB-induced VSMC without cytotoxicity. ► CPT arrests the cell cycle in G0/G1 phase by downregulation of cyclin D1 and CDK2. ► CPT significantly attenuates Akt phosphorylation in PDGF-BB signaling pathway. ► LY294002 enhanced the inhibitory effect of CPT on VSMC proliferation. ► Thus, CPT is mediated by downregulating the PI3K/Akt signaling pathway.

  2. Disruption of the PI3K/AKT/mTOR signaling cascade and induction of apoptosis in HL-60 cells by an essential oil from Monarda citriodora.

    PubMed

    Pathania, Anup Singh; Guru, Santosh Kumar; Verma, M K; Sharma, Chetna; Abdullah, Sheikh Tasduq; Malik, Fayaz; Chandra, Suresh; Katoch, Meenu; Bhushan, Shashi

    2013-12-01

    We have isolated an essential oil from Monarda citriodora (MC) and characterized its 22 chemical constituents with thymol (82%), carvacrol (4.82%), β-myrcene (3.45%), terpinen-4-ol (2.78%) and p-cymene (1.53%) representing the major constituents. We have reported for the first time the chemotherapeutic potential of MC in human promyelocytic leukemia HL-60 cells by means of apoptosis and disruption of the PI3K/AKT/mTOR signaling cascade. MC and its major constituent, thymol, inhibit the cell proliferation in different types of cancer cell lines like HL-60, MCF-7, PC-3, A-549 and MDAMB-231. MC was found to be more cytotoxic than thymol in HL-60 cells with an IC50 value of 22 μg/ml versus 45 μg/ml for thymol. Both MC and thymol induce apoptosis in HL-60 cells, which is evident by Hoechst staining, cell cycle analysis and immuno-expression of Bcl-xL, caspase-3,-8,-9 and PARP-1 cleavage. Both induce apoptosis by extrinsic and intrinsic apoptotic pathways that were confirmed by enhanced expression of death receptors (TNF-R1, Fas), caspase-9, loss of mitochondrial membrane potential and regression of Bcl-2/Bax ratio. Interestingly, both MC and thymol inhibit the downstream and upstream signaling of PI3K/AKT/mTOR pathway. The degree of apoptosis induction and disruption of the PI3K signaling cascade by MC was significantly higher when compared to thymol. PMID:23994707

  3. Disruption of the PI3K/AKT/mTOR signaling cascade and induction of apoptosis in HL-60 cells by an essential oil from Monarda citriodora.

    PubMed

    Pathania, Anup Singh; Guru, Santosh Kumar; Verma, M K; Sharma, Chetna; Abdullah, Sheikh Tasduq; Malik, Fayaz; Chandra, Suresh; Katoch, Meenu; Bhushan, Shashi

    2013-12-01

    We have isolated an essential oil from Monarda citriodora (MC) and characterized its 22 chemical constituents with thymol (82%), carvacrol (4.82%), β-myrcene (3.45%), terpinen-4-ol (2.78%) and p-cymene (1.53%) representing the major constituents. We have reported for the first time the chemotherapeutic potential of MC in human promyelocytic leukemia HL-60 cells by means of apoptosis and disruption of the PI3K/AKT/mTOR signaling cascade. MC and its major constituent, thymol, inhibit the cell proliferation in different types of cancer cell lines like HL-60, MCF-7, PC-3, A-549 and MDAMB-231. MC was found to be more cytotoxic than thymol in HL-60 cells with an IC50 value of 22 μg/ml versus 45 μg/ml for thymol. Both MC and thymol induce apoptosis in HL-60 cells, which is evident by Hoechst staining, cell cycle analysis and immuno-expression of Bcl-xL, caspase-3,-8,-9 and PARP-1 cleavage. Both induce apoptosis by extrinsic and intrinsic apoptotic pathways that were confirmed by enhanced expression of death receptors (TNF-R1, Fas), caspase-9, loss of mitochondrial membrane potential and regression of Bcl-2/Bax ratio. Interestingly, both MC and thymol inhibit the downstream and upstream signaling of PI3K/AKT/mTOR pathway. The degree of apoptosis induction and disruption of the PI3K signaling cascade by MC was significantly higher when compared to thymol.

  4. Netrin-1 induces the migration of Schwann cells via p38 MAPK and PI3K-Akt signaling pathway mediated by the UNC5B receptor

    SciTech Connect

    Lv, Jianwei; Sun, Xiaolei; Ma, Jianxiong; Ma, Xinlong; Zhang, Yang; Li, Fengbo; Li, Yanjun; Zhao, Zhihu

    2015-08-14

    Schwann cells (SCs) play an essentially supportive role in the regeneration of injured peripheral nerve system (PNS). As Netrin-1 is crucial for the normal development of nervous system (NS) and can direct the process of damaged PNS regeneration, our study was designed to determine the role of Netrin-1 in RSC96 Schwann cells (an immortalized rat Schwann cell line) proliferation and migration. Our studies demonstrated that Netrin-1 had no effect on RSC96 cells proliferation, while significantly promoted RSC96 cells migration. The Netrin-1-induced RSC96 cells migration was significantly attenuated by inhibition of p38 and PI3K through pretreatment with SB203580 and LY294002 respectively, but not inhibition of MEK1/2 and JNK by U0126-EtOH and SP600125 individually. Treatment with Netrin-1 enhanced the phosphorylation of p38 and Akt. QRT-PCR indicated that Netrin-1 and only its receptors Unc5a, Unc5b and Neogenin were expressed in RSC96 cells, among which Unc5b expressed the most. And UNC5B protein was significantly increased after stimulated by Netrin-1. In conclusion, we show here that Netrin-1-enhanced SCs migration is mediated by activating p38 MAPK and PI3K-Akt signal cascades via receptor UNC5B, which suggests that Netrin-1 could serve as a new therapeutic strategy and has potential application value for PNS regeneration. - Highlights: • Netrin-1 attracts RSC96 Schwann cells migration in a dose dependent manner. • Netrin-1 induced Schwann cells migration is p38 and PI3K-Akt signaling dependent. • UNC5B may be dominant receptor mediating Netrin-1′ effect on RSC96 cells motility. • Netrin-1 may promote peripheral nerve repair by enhancing Schwann cells motility.

  5. Methylophiopogonanone A suppresses ischemia/reperfusion-induced myocardial apoptosis in mice via activating PI3K/Akt/eNOS signaling pathway

    PubMed Central

    He, Fei; Xu, Bang-long; Chen, Cai; Jia, Hong-jing; Wu, Ji-xiong; Wang, Xiao-chen; Sheng, Jian-long; Huang, Li; Cheng, Jing

    2016-01-01

    Aim: The dried tuber root of Ophiopogon japonicus has been used in the traditional Chinese medicine for treatment of myocardial ischemia and thrombosis. In this study we investigated the effects of methylophiopogonanone A (MO-A), a major homoisoflavonoid in Ophiopogon japonicus, on myocardial ischemia/reperfusion (I/R) injury. Methods: Mice were pretreated with MO-A (10 mg·kg-1·d-1, po) for 2 weeks and then subjected to transient occlusion of the left anterior descending coronary artery. Cardiac function was evaluated, and the infarct size and apoptosis index were assessed. The mechanisms underlying the cardio-protection of MO-A were analyzed in H9C2 rat cardiomyocytes subjected to hypoxia/reoxygenation (H/R). The cell viability and apoptosis were evaluated; apoptotic and relevant signaling proteins were analyzed. NO levels in the culture medium were assessed. Results: In I/R mice, pretreatment with MO-A significantly reduced the infarct size (by 60.7%) and myocardial apoptosis (by 56.8%), and improved cardiac function. In H9C2 cells subjected to H/R, pretreatment with MO-A (10 μmol/L) significantly decreased apoptosis and cleaved caspase-3 expression, elevated the Bcl-2/Bax ratio and restored NO production. Furthermore, pretreatment with MO-A markedly increased the activation of PI3K/Akt/eNOS pathway in H9C2 cells subjected to H/R, and the protective effects of MO-A were abolished in the presence of the PI3K inhibitor wortmannin (100 nmol/L). Conclusion: MO-A attenuates I/R-induced myocardial apoptosis in mice via activating the PI3K/Akt/eNOS signaling pathway. PMID:27063216

  6. miR-21 Reduces Hydrogen Peroxide-Induced Apoptosis in c-kit+ Cardiac Stem Cells In Vitro through PTEN/PI3K/Akt Signaling

    PubMed Central

    Wang, Yan; Long, Xianping; Zhao, Ranzun; Wang, Zhenglong; Liu, Zhijiang

    2016-01-01

    The low survival rate of cardiac stem cells (CSCs) in the infarcted myocardium hampers cell therapy for ischemic cardiomyopathy. MicroRNA-21 (miR-21) and one of its target proteins, PTEN, contribute to the survival and proliferation of many cell types, but their prosurvival effects in c-kit+ CSC remain unclear. Thus, we hypothesized that miR-21 reduces hydrogen peroxide- (H2O2-) induced apoptosis in c-kit+ CSC and estimated the contribution of PTEN/PI3K/Akt signaling to this oxidative circumstance. miR-21 mimics efficiently reduced H2O2-induced apoptosis in c-kit+ CSC, as evidenced by the downregulation of the proapoptosis proteins caspase-3 and Bax and upregulation of the antiapoptotic Bcl-2. In addition, the gain of function of miR-21 in c-kit+ CSC downregulated the protein level of PTEN although its mRNA level changed slightly; in the meantime, miR-21 overexpression also increased phospho-Akt (p-Akt). The antiapoptotic effects of miR-21 were comparable with Phen (bpV), the selective inhibitor of PTEN, while miR-21 inhibitor or PI3K's inhibitor LY294002 efficiently attenuated the antiapoptotic effect of miR-21. Taken together, these results indicate that the anti-H2O2-induced apoptosis effect of miR-21 in c-kit+ CSC is contributed by PTEN/PI3K/Akt signaling. miR-21 could be a potential molecule to facilitate the c-kit+ CSC therapy in ischemic myocardium. PMID:27803763

  7. Differential regulation of protrusion and polarity by PI(3)K during neutrophil motility in live zebrafish

    PubMed Central

    Yoo, Sa Kan; Deng, Qing; Cavnar, Peter J.; Wu, Yi I.; Hahn, Klaus M.; Huttenlocher, Anna

    2010-01-01

    Summary Cell polarity is crucial for directed migration. Here we show that phosphoinositide 3-kinase (PI(3)K) mediates neutrophil migration in vivo by differentially regulating cell protrusion and polarity. The dynamics of PI(3)K products PI(3,4,5)P3-PI(3,4)P2 during neutrophil migration were visualized in living zebrafish, revealing that PI(3)K activation at the leading edge is critical for neutrophil motility in intact tissues. A genetically encoded photoactivatable Rac was used to demonstrate that localized activation of Rac is sufficient to direct migration with precise temporal and spatial control in vivo. Similar stimulation of PI(3)K-inhibited cells did not direct migration. Localized Rac activation rescued membrane protrusion but not anteroposterior polarization of F-actin dynamics of PI(3)K-inhibited cells. Uncoupling Rac-mediated protrusion and polarization suggests a paradigm of two-tiered PI(3)K-mediated regulation of cell motility. This work provides new insight into how cell signaling at the front and back of the cell is coordinated during polarized cell migration in intact tissues within a multicellular organism. PMID:20159593

  8. Calcium Signaling Involvement in Cadmium-Induced Astrocyte Cytotoxicity and Cell Death Through Activation of MAPK and PI3K/Akt Signaling Pathways.

    PubMed

    Jiang, Jiao Hua; Ge, Guo; Gao, Kai; Pang, Ying; Chai, Rui Chao; Jia, Xi Hua; Kong, Jin Ge; Yu, Albert Cheung-Hoi

    2015-09-01

    Cadmium (Cd), a highly ubiquitous toxic heavy metal, can contaminate the environment, including agricultural soil, water and air, via industrial runoff and other sources of pollution. Cd accumulated in the body via direct exposure or through the food chain results in neurodegeneration and many other diseases. Previous studies on its toxicity in the central nervous system (CNS) focused mainly on neurons. To obtain a more comprehensive understanding of Cd toxicity for the CNS, we investigated how astrocytes respond to acute and chronic Cd exposure and its toxic molecular mechanisms. When primary cultures of cerebral cortical astrocytes incubated with 1-300 μM CdCl2, morphological changes, LDH release and cell death were observed in a time and dose-dependent manner. Further studies demonstrated that acute and chronic Cd treatment phosphorylated JNK, p38 and Akt to different degrees, while ERK1/2 was only phosphorylated under low doses of Cd (10 μM) exposure. Inhibition of JNK and PI3K/Akt, but not of p38, could partially protect astrocyte from cytotoxicity in chronic and acute Cd exposure. Moreover, Cd also induced a strong calcium signal, while BAPTA, a specific intracellular calcium (Ca(2+)) chelator, prevented Cd-induced intracellular increase of calcium levels in astrocytes; inhibited the Cd-induced activation of ERK1/2, JNK, p38 and Akt; and also significantly reduced astrocyte cell death. All of these results suggested that the Cd-Ca(2+)-MAPK and PI3K/Akt signaling pathways were involved in Cd-induced toxicity in astrocytes. This toxicity involvement indicates that these pathways may be exploited as a target for the prevention of Cd-induced neurodegenerative diseases. PMID:26248512

  9. First-in-human Phase I study of Pictilisib (GDC-0941), a potent pan-class I phosphatidylinositol-3-kinase (PI3K) inhibitor, in patients with advanced solid tumors

    PubMed Central

    Baird, Richard; Kristeleit, Rebecca; Shah, Krunal; Moreno, Victor; Clarke, Paul A.; Raynaud, Florence I.; Levy, Gallia; Ware, Joseph A; Mazina, Kathryn; Lin, Ray; Wu, Jenny; Fredrickson, Jill; Spoerke, Jill M; Lackner, Mark R; Yan, Yibing; Friedman, Lori S.; Kaye, Stan B.; Derynck, Mika K.; Workman, Paul; de Bono, Johann S.

    2014-01-01

    Purpose This first-in-human dose-escalation trial evaluated the safety, tolerability, maximal tolerated dose (MTD), dose limiting toxicities (DLTs), pharmacokinetics, pharmacodynamics and preliminary clinical activity of pictilisib (GDC-0941), an oral, potent and selective inhibitor of the Class I phosphatidylinositol-3-kinases (PI3K). Patients and Methods Sixty patients with solid tumors received pictilisib at 14 dose levels from 15 to 450mg once-daily, initially on days 1-21 every 28 days and later, utilizing continuous dosing for selected dose levels. Pharmacodynamic studies incorporated 18F-FDG-PET, and assessment of phosphorylated AKT and S6 ribosomal protein in platelet-rich plasma and tumor tissue. Results Pictilisib was well-tolerated. The most common toxicities were grade 1-2 nausea, rash and fatigue while the DLT was grade 3 maculopapular rash (450mg, 2 of 3 patients; 330mg, 1 of 7 patients). The pharmacokinetic profile was dose-proportional and supported once-daily dosing. Levels of phosphorylated serine-473 AKT were suppressed >90% in platelet rich plasma at 3 hours post-dose at the MTD and in tumor at pictilisib doses associated with AUC >20uM.hr. Significant increase in plasma insulin and glucose levels, and >25% decrease in 18F-FDG uptake by PET in 7 of 32 evaluable patients confirmed target modulation. A patient with V600E BRAF mutant melanoma and another with platinum-refractory epithelial ovarian cancer exhibiting PTEN loss and PIK3CA amplification demonstrated partial response by RECIST and GCIG-CA125 criteria, respectively. Conclusion Pictilisib was safely administered with a dose-proportional pharmacokinetic profile, on-target pharmacodynamic activity at dose levels ≥100mg and signs of antitumor activity. The recommended Phase II dose was continuous dosing at 330mg once-daily. PMID:25370471

  10. Do airborne biogenic chemicals interact with the PI3K/Akt/mTOR cell signalling pathway to benefit human health and wellbeing in rural and coastal environments?

    PubMed

    Moore, Michael N

    2015-07-01

    Living and taking recreation in rural and coastal environments promote health and wellbeing, although the causal factors involved are unclear. It has been proposed that such environments provide a counter to the stresses of everyday living, leading to enhanced mental and physical health. Living in natural environments will result in airborne exposure to a wide range of biogenic chemicals through inhalation and ingestion of airborne microbiota and particles. The "biogenics" hypothesis formulated here is that regular exposure to low concentrations of mixtures of natural compounds and toxins in natural environments confers pleiotropic health benefits by inhibiting the activities of interconnected cell signalling systems, particularly PI3K/Akt/mTORC1. When overactive, Akt and mTOR (mTORC1) can lead to many pathological processes including cancers, diabetes, inflammation, immunosuppression, and neurodegenerative diseases. There is a substantial body of evidence that many natural products (i.e., from bacteria, algae, fungi and higher plants) inhibit the activities of these protein kinases. Other mTOR-related interconnected metabolic control "switches" (e.g., PTEN & NF-κB), autophagy and other cytoprotective processes are also affected by natural products. The "biogenics" hypothesis formulated here is that regular intermittent exposure to a mixture of airborne biogenic compounds in natural environments confers pleiotropic health benefits by inhibiting activities of the highly interconnected PI3K/Akt/mTORC1 system. It is proposed that future experimental exposures to biogenic aerosols in animal models coupled with epidemiology, should target the activities of the various kinases in the PI3K/Akt/mTORC1 systems and related physiological processes for selected urban, rural and coastal populations in order to test this hypothesis.

  11. Twist promotes reprogramming of glucose metabolism in breast cancer cells through PI3K/AKT and p53 signaling pathways.

    PubMed

    Yang, Li; Hou, Yixuan; Yuan, Jie; Tang, Shifu; Zhang, Hailong; Zhu, Qing; Du, Yan-e; Zhou, Mingli; Wen, Siyang; Xu, Liyun; Tang, Xi; Cui, Xiaojiang; Liu, Manran

    2015-09-22

    Twist, a key regulator of epithelial-mesenchymal transition (EMT), plays an important role in the development of a tumorigenic phenotype. Energy metabolism reprogramming (EMR), a newly discovered hallmark of cancer cells, potentiates cancer cell proliferation, survival, and invasion. Currently little is known about the effects of Twist on tumor EMR. In this study, we found that glucose consumption and lactate production were increased and mitochondrial mass was decreased in Twist-overexpressing MCF10A mammary epithelial cells compared with vector-expressing MCF10A cells. Moreover, these Twist-induced phenotypic changes were augmented by hypoxia. The expression of some glucose metabolism-related genes such as PKM2, LDHA, and G6PD was also found to be upregulated. Mechanistically, activated β1-integrin/FAK/PI3K/AKT/mTOR and suppressed P53 signaling were responsible for the observed EMR. Knockdown of Twist reversed the effects of Twist on EMR in Twist-overexpressing MCF10A cells and Twist-positive breast cancer cells. Furthermore, blockage of the β1-integrin/FAK/PI3K/AKT/mTOR pathway by siRNA or specific chemical inhibitors, or rescue of p53 activation can partially reverse the switch of glucose metabolism and inhibit the migration of Twist-overexpressing MCF10A cells and Twist-positive breast cancer cells. Thus, our data suggest that Twist promotes reprogramming of glucose metabolism in MCF10A-Twist cells and Twist-positive breast cancer cells via activation of the β1-integrin/FAK/PI3K/AKT/mTOR pathway and inhibition of the p53 pathway. Our study provides new insight into EMR. PMID:26342198

  12. Twist promotes reprogramming of glucose metabolism in breast cancer cells through PI3K/AKT and p53 signaling pathways.

    PubMed

    Yang, Li; Hou, Yixuan; Yuan, Jie; Tang, Shifu; Zhang, Hailong; Zhu, Qing; Du, Yan-e; Zhou, Mingli; Wen, Siyang; Xu, Liyun; Tang, Xi; Cui, Xiaojiang; Liu, Manran

    2015-09-22

    Twist, a key regulator of epithelial-mesenchymal transition (EMT), plays an important role in the development of a tumorigenic phenotype. Energy metabolism reprogramming (EMR), a newly discovered hallmark of cancer cells, potentiates cancer cell proliferation, survival, and invasion. Currently little is known about the effects of Twist on tumor EMR. In this study, we found that glucose consumption and lactate production were increased and mitochondrial mass was decreased in Twist-overexpressing MCF10A mammary epithelial cells compared with vector-expressing MCF10A cells. Moreover, these Twist-induced phenotypic changes were augmented by hypoxia. The expression of some glucose metabolism-related genes such as PKM2, LDHA, and G6PD was also found to be upregulated. Mechanistically, activated β1-integrin/FAK/PI3K/AKT/mTOR and suppressed P53 signaling were responsible for the observed EMR. Knockdown of Twist reversed the effects of Twist on EMR in Twist-overexpressing MCF10A cells and Twist-positive breast cancer cells. Furthermore, blockage of the β1-integrin/FAK/PI3K/AKT/mTOR pathway by siRNA or specific chemical inhibitors, or rescue of p53 activation can partially reverse the switch of glucose metabolism and inhibit the migration of Twist-overexpressing MCF10A cells and Twist-positive breast cancer cells. Thus, our data suggest that Twist promotes reprogramming of glucose metabolism in MCF10A-Twist cells and Twist-positive breast cancer cells via activation of the β1-integrin/FAK/PI3K/AKT/mTOR pathway and inhibition of the p53 pathway. Our study provides new insight into EMR.

  13. Twist promotes reprogramming of glucose metabolism in breast cancer cells through PI3K/AKT and p53 signaling pathways

    PubMed Central

    Yuan, Jie; Tang, Shifu; Zhang, Hailong; Zhu, Qing; Du, Yan-e; Zhou, Mingli; Wen, Siyang; Xu, Liyun; Tang, Xi; Cui, Xiaojiang; Liu, Manran

    2015-01-01

    Twist, a key regulator of epithelial-mesenchymal transition (EMT), plays an important role in the development of a tumorigenic phenotype. Energy metabolism reprogramming (EMR), a newly discovered hallmark of cancer cells, potentiates cancer cell proliferation, survival, and invasion. Currently little is known about the effects of Twist on tumor EMR. In this study, we found that glucose consumption and lactate production were increased and mitochondrial mass was decreased in Twist-overexpressing MCF10A mammary epithelial cells compared with vector-expressing MCF10A cells. Moreover, these Twist-induced phenotypic changes were augmented by hypoxia. The expression of some glucose metabolism-related genes such as PKM2, LDHA, and G6PD was also found to be upregulated. Mechanistically, activated β1-integrin/FAK/PI3K/AKT/mTOR and suppressed P53 signaling were responsible for the observed EMR. Knockdown of Twist reversed the effects of Twist on EMR in Twist-overexpressing MCF10A cells and Twist-positive breast cancer cells. Furthermore, blockage of the β1-integrin/FAK/PI3K/AKT/mTOR pathway by siRNA or specific chemical inhibitors, or rescue of p53 activation can partially reverse the switch of glucose metabolism and inhibit the migration of Twist-overexpressing MCF10A cells and Twist-positive breast cancer cells. Thus, our data suggest that Twist promotes reprogramming of glucose metabolism in MCF10A-Twist cells and Twist-positive breast cancer cells via activation of the β1-integrin/FAK/PI3K/AKT/mTOR pathway and inhibition of the p53 pathway. Our study provides new insight into EMR. PMID:26342198

  14. 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. PMID:26162710

  15. Andrographolide promotes vincristine-induced SK-NEP-1 tumor cell death via PI3K-AKT-p53 signaling pathway

    PubMed Central

    Zhang, Mingsheng; Xue, Enda; Shao, Wei

    2016-01-01

    Background Nephroblastoma (Wilms’ tumor [WT]) is the most common malignant renal cancer in children. Although the outcome of WT has significantly improved as a result of the combination of surgery, chemotherapy, and radiotherapy; in some cases WT results in severe complications. Thus, novel strategies that would decrease treatment burden are required. The aim of the current study was to investigate the synergistic antitumor effect of andrographolide (AND) in combination with vincristine (VCR) on WT cells. Methods Cell Counting Kit-8 assay was used to investigate the synergistic antiproliferation effect of AND and/or VCR on SK-NEP-1 cells in vitro. Meanwhile, SK-NEP-1 xenografts were used to detect the antitumor effect in vivo. Apoptosis and autophagy were then detected by Annexin V, monodansylcadaverine staining. Finally, the underlying signaling transduction was determined with Western blotting. Results The combination of AND with VCR significantly suppressed SK-NEP-1 cell proliferation in vitro and inhibited xenograft tumor growth in vivo, compared with AND or VCR treatment alone. In addition, the synergistic antitumor effect of AND on the cells was due to an increased apoptosis, not autophagy. Moreover, PI3K-AKT-p53 signaling pathway was involved in the process of combination treatment, which was confirmed when a selective AKT activator was applied. Conclusion The combination of AND with VCR has a strong synergistic antitumor effect on WT via PI3K-AKT-p53 signaling pathway, thereby representing a potential treatment for WT in the near future. PMID:27729773

  16. 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.

  17. Co-targeting the PI3K/mTOR and JAK2 signalling pathways produces synergistic activity against myeloproliferative neoplasms

    PubMed Central

    Bartalucci, Niccolò; Tozzi, Lorenzo; Bogani, Costanza; Martinelli, Serena; Rotunno, Giada; Villeval, Jean-Luc; Vannucchi, Alessandro M

    2013-01-01

    Aberrant JAK2 signalling plays a central role in myeloproliferative neoplasms (MPN). JAK2 inhibitors have proven to be clinically efficacious, however, they are not mutation-specific and competent enough to suppress neoplastic clonal haematopoiesis. We hypothesized that, by simultaneously targeting multiple activated signalling pathways, MPN could be more effectively treated. To this end we investigated the efficacy of BEZ235, a dual PI3K/mTOR inhibitor, alone and in combination with the JAK1/JAK2 inhibitor ruxolitinib, in different preclinical models of MPN. Single-agent BEZ235 inhibited the proliferation and induced cell cycle arrest and apoptosis of mouse and human JAK2V617F mutated cell lines at concentrations significantly lower than those required to inhibit the wild-type counterpart, and preferentially prevented colony formation from JAK2V617F knock-in mice and patients' progenitor cells compared with normal ones. Co-treatment of BEZ235 and ruxolitinib produced significant synergism in all these in-vitro models. Co-treatment was also more effective than single drugs in reducing the extent of disease and prolonging survival of immunodeficient mice injected with JAK2V617F-mutated Ba/F3-EPOR cells and in reducing spleen size, decreasing reticulocyte count and improving spleen histopathology in conditional JAK2V617F knock-in mice. In conclusion, combined inhibition of PI3K/mTOR and JAK2 signalling may represent a novel therapeutic strategy in MPN. PMID:24237791

  18. miRs-134 and -370 function as tumor suppressors in colorectal cancer by independently suppressing EGFR and PI3K signalling

    PubMed Central

    El-Daly, Sherien M.; Abba, Mohammed L.; Patil, Nitin; Allgayer, Heike

    2016-01-01

    Growth factor receptor signalling plays a central and critical role in colorectal cancer. Most importantly, the EGFR signalling cascade involving PI3K/AKT/mTOR and Raf/MEK/ERK pathways are particularly relevant, since they are commonly activated in several cancer entities, including colorectal cancer. In this study, we show that miRs-134 and -370 are both capable of regulating these pathways by targeting EGFR and PIK3CA. In three different colorectal cancer cell lines (DLD1, HCT-116 and RKO), suppression of EGFR and PIK3CA through the enhanced expression of miR-134 or -370 led to a suppression of the key molecules of the PI3K/AKT/mTOR pathway. Furthermore, overexpression of miR-134 or -370 resulted in a significant reduction of cell proliferation, colony formation, migration, invasion and in-vivo tumor growth and metastasis. Concurrent experiments with small interfering RNAs targeting the prime targets show that our selected miRNAs exert a greater functional influence and affect more downstream molecules than is seen with silencing of the individual proteins. Taken together, these data indicate that miRs-134 and -370 are potential tumour suppressor miRNAs and could play a fundamental role in suppressing colorectal cancer tumorigenesis through their ability to co-ordinately regulate EGFR signalling cascade by independently targeting EGFR and PIK3CA. PMID:27095166

  19. 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.

  20. Anti-angiogenic activity of macrolactin A and its succinyl derivative is mediated through inhibition of class I PI3K activity and its signaling.

    PubMed

    Kang, Youra; Regmi, Sushil Chandra; Kim, Mi Yeong; Banskota, Suhrid; Gautam, Jaya; Kim, Dong Hee; Kim, Jung-Ae

    2015-02-01

    In the current study, macrolactin compounds, macrolactin A (MA) and 7-O-succinyl macrolactin A (SMA), were investigated for their anti-angiogenic activities and action mechanism. MA and SMA inhibited in vitro and in vivo angiogenesis induced by three different classes of pro-angiogenic factors, VEGF, IL-8, and TNF-α. SMA exhibited stronger anti-angiogenic activity than MA, and such anti-angiogenic activity of SMA was consistently observed in MDA-MB-231 human breast cancer cell-inoculated CAM assay showing dose-dependent suppression of tumor growth and tumor-induced angiogenesis. In an in vitro PI3K competitive activity assay, SMA induced concentration-dependent inhibition of class I PI3K isoforms, p110α, p110β, p110δ, and p110γ. In addition, non-receptor tyrosine kinase c-Src, which is involved in the activation of PI3K heterodimer, was suppressed by MA and SMA. Correspondingly, MA and SMA significantly inhibited the stimulus-induced phosphorylation of Akt, mTOR, p70S6K, and ribosomal S6 in human umbilical vein endothelial cells (HUVECs). At the same time, the stimulus-induced production of reactive oxygen species (ROS) and activation of NF-κB were significantly suppressed by MA and SMA. Moreover, the macrolactins suppressed NF-κB-regulated HSP90 protein expression, which stabilizes phosphorylated Akt and NADPH oxidase. Suppression of NF-κB in macrolactin-treated HUVECs with concurrent inhibition of rS6 indicates that MAs effectively block angiogenesis through down-regulation of genes related to angiogenesis at both transcriptional and translational levels. Taken together, the results demonstrate that anti-angiogenic effect of MA and SMA is mediated through inhibition of PI3K/Akt and NADPH oxidase-derived ROS/NF-κB signaling pathways. These results further indicate that MA and SMA may be applicable for treatment of various diseases associated with angiogenesis.

  1. Serine 1179 phosphorylation of endothelial nitric oxide synthase caused by 2,4,6-trinitrotoluene through PI3K/Akt signaling in endothelial cells

    SciTech Connect

    Sun Yang; Sumi, Daigo; Kumagai, Yoshito . E-mail: yk-em-tu@md.tsukuba.ac.jp

    2006-07-01

    Although 2,4,6-trinitrotoluene (TNT) has been found to uncouple nitric oxide synthase (NOS), thereby leading to reactive oxygen species (ROS), cellular response against TNT still remains unclear. Exposure of bovine aortic endothelial cells (BAECs) to TNT (100 {mu}M) resulted in serine 1179 phosphorylation of endothelial NOS (eNOS). With specific inhibitors (wortmannin and LY294002), we found that PI3K/Akt signaling participated in the eNOS phosphorylation caused by TNT, whereas the ERK pathway did not. ROS were generated following exposure of BAECs to TNT. However, TNT-mediated phosphorylation of either eNOS or Akt was drastically blocked by NAC and PEG-CAT. Interestingly, pretreatment with apocynin, a specific inhibitor for NADPH oxidase, diminished the phosphorylation of eNOS and Akt. These results suggest that TNT affects NADPH oxidase, thereby generating hydrogen peroxide, which is capable of activating PI3K/Akt signaling associated with eNOS Ser 1179 phosphorylation.

  2. Induction of metastatic potential by TrkB via activation of IL6/JAK2/STAT3 and PI3K/AKT signaling in breast cancer.

    PubMed

    Kim, Min Soo; Lee, Won Sung; Jeong, Joon; Kim, Seong-Jin; Jin, Wook

    2015-11-24

    In metastatic breast cancers, the acquisition of metastatic ability, which leads to clinically incurable disease and poor survival, has been associated with acquisition of epithelial-mesenchymal transition (EMT) program and self-renewing trait (CSCs) via activation of PI3K/AKT and IL6/JAK2/STAT3 signaling pathways. We found that TrkB is a key regulator of PI3K/AKT and JAK/STAT signal pathway-mediated tumor metastasis and EMT program. Here, we demonstrated that TrkB activates AKT by directly binding to c-Src, leading to increased proliferation. Also, TrkB increases Twist-1 and Twist-2 expression through activation of JAK2/STAT3 by inducing c-Src-JAK2 complex formation. Furthermore, TrkB in the absence of c-Src binds directly to JAK2 and inhibits SOCS3-mediated JAK2 degradation, resulting in increased total JAK2 and STAT3 levels, which subsequently leads to JAK2/STAT3 activation and Twist-1 upregulation. Additionally, activation of the JAK2/STAT3 pathway via induction of IL-6 secretion by TrkB enables induction of activation of the EMT program via induction of STAT3 nuclear translocation. These observations suggest that TrkB is a promising target for future intervention strategies to prevent tumor metastasis, EMT program and self-renewing trait in breast cancer. PMID:26515594

  3. PI3K/Akt and Stat3 signaling regulated by PTEN control of the cancer stem cell population, proliferation and senescence in a glioblastoma cell line.

    PubMed

    Moon, Seok-Ho; Kim, Dae-Kwan; Cha, Young; Jeon, Iksoo; Song, Jihwan; Park, Kyung-Soon

    2013-03-01

    Malignant gliomas are the most common primary brain tumor in adults. A number of genes have been implicated in glioblastoma including mutation and deletion of PTEN. PTEN is a regulator of PI3K-mediated Akt signaling pathways and has been recognized as a therapeutic target in glioblastoma. To achieve potent therapeutic inhibition of the PI3K-Akt pathway in glioblastoma, it is essential to understand the interplay between the regulators of its activation. Here, ectopic expression of PTEN in the U-87MG human glioblastoma-astrocytoma cell line is shown to result in the depletion of glioblastoma stem cells (GSCs) and to cause growth retardation and senescence. These effects are likely to be associated with PTEN-mediated cooperative perturbation of Akt and Stat3 signals. Using an in vivo rat model of glioblastoma, we showed that PTEN-overexpressing U-87MG cells failed to induce tumor formation, while untreated U-87MG cells did so. Furthermore, cells expressing the phosphorylated form of Stat3 were completely absent from the brain of rats implanted with PTEN-overexpressing U-87MG cells. Based on these results, PTEN appears to function as a crucial inhibitor of GSCs and as an inducer of senescence, suggesting that functional enhancement of the PTEN pathway will be useful to provide a therapeutic strategy for targeting glioblastoma. PMID:23314408

  4. Thrombin Promotes Matrix Metalloproteinase-13 Expression through the PKCδ/c-Src/EGFR/PI3K/Akt/AP-1 Signaling Pathway in Human Chondrocytes

    PubMed Central

    Huang, Chun-Yin; Lin, Hsiu-Jung; Chen, Hsin-Shui; Cheng, Shi-Yann; Hsu, Horng-Chaung; Tang, Chih-Hsin

    2013-01-01

    Thrombin is a key mediator of fibrin deposition, angiogenesis, and proinflammatory processes. Abnormalities in these processes are primary features of rheumatoid arthritis and osteoarthritis. Matrix metalloproteinase-13 (MMP-13) may contribute to the breakdown of articular cartilage during arthritis. However, the role of thrombin in MMP-13 production in chondrocytes is unknown. In this study, we investigated the intracellular signaling pathways involved in thrombin-induced MMP-13 expression in human chondrocytes. We found that stimulation with thrombin led to increased secretion of MMP-13 in cultured human chondrocytes. Further, this thrombin-induced MMP-13 production was reduced after transfection with siRNAs against protease activated receptors 1 and 3 (PAR1 and PAR3), but not with PAR4 siRNA. Treatment with specific inhibitors for PKCδ, c-Src, EGFR, PI3K, Akt, or AP-1 or with the corresponding siRNAs against these signaling proteins also abolished the thrombin-mediated increase in MMP-13 production in chondrocytes. Our results provide evidence that thrombin acts through the PAR1/PAR3 receptors and activates PKCδ and c-Src, resulting in EGFR transactivation and activation of PI3K, Akt, and finally AP-1 on the MMP-13 promoter, thereby contributing to cartilage destruction during arthritis. PMID:24385683

  5. A PI3K p110β–Rac signalling loop mediates Pten-loss-induced perturbation of haematopoiesis and leukaemogenesis

    PubMed Central

    Yuzugullu, Haluk; Baitsch, Lukas; Von, Thanh; Steiner, Allison; Tong, Haoxuan; Ni, Jing; Clayton, Linda K.; Bronson, Roderick; Roberts, Thomas M.; Gritsman, Kira; Zhao, Jean J.

    2015-01-01

    The tumour suppressor PTEN, which antagonizes PI3K signalling, is frequently inactivated in haematologic malignancies. In mice, deletion of PTEN in haematopoietic stem cells (HSCs) causes perturbed haematopoiesis, myeloproliferative neoplasia (MPN) and leukaemia. Although the roles of the PI3K isoforms have been studied in PTEN-deficient tumours, their individual roles in PTEN-deficient HSCs are unknown. Here we show that when we delete PTEN in HSCs using the Mx1–Cre system, p110β ablation prevents MPN, improves HSC function and suppresses leukaemia initiation. Pharmacologic inhibition of p110β in PTEN-deficient mice recapitulates these genetic findings, but suggests involvement of both Akt-dependent and -independent pathways. Further investigation reveals that a p110β–Rac signalling loop plays a critical role in PTEN-deficient HSCs. Together, these data suggest that myeloid neoplasia driven by PTEN loss is dependent on p110β via p110β–Rac-positive-feedback loop, and that disruption of this loop may offer a new and effective therapeutic strategy for PTEN-deficient leukaemia. PMID:26442967

  6. FGF2 regulates proliferation, migration, and invasion of ECA109 cells through PI3K/Akt signalling pathway in vitro.

    PubMed

    Shi, Hui; Xu, Jingjing; Zhao, Rui; Wu, Huiqun; Gu, Luo; Chen, Yijiang

    2016-05-01

    Esophageal cancer is one of the most common malignant cancers that arise from esophagus tissues. Fibroblast growth factor 2 (FGF2) has been implicated in multiple biological functions and was considered as an oncogenic factor in tumorigenesis. However, the effects of FGF2 in esophageal carcinoma are yet to be fully elucidated. To better understand the function of FGF2 in esophageal cancer, we used the esophageal cancer cell line ECA109 as a cell model and downregulated FGF2 expression using RNAi; the results showed that insufficient expression of FGF2 inhibited cells proliferation, migration, and invasion of ECA109 cells. Meanwhile, the proliferation, migration, and invasion abilities were stimulated after treatment of exogenous FGF2. In addition, a PI3K/Akt signalling pathway inhibitor (LY294002) alleviated the tumorigenic effects of FGF2. These findings implied that the oncogenic effects of FGF2 was mediated, at least in part, through the PI3K/Akt signalling pathway and FGF2 may be a potential therapeutic target to constrain the tumorigenesis of esophageal cancer. PMID:26833879

  7. Discovery of a Novel Series of Thienopyrimidine as Highly Potent and Selective PI3K Inhibitors

    PubMed Central

    2015-01-01

    Inhibition of the phosphoinositide 3-kinase (PI3K)/AKT/mammalian target of rapamycin (mTOR) signaling pathway provides a promising new approach for cancer therapy. Through a rational design, a novel series of thienopyrimidine was discovered as highly potent and selective PI3K inhibitors. These thienopyrimidine derivatives were demonstrated to bear nanomolar PI3Kα inhibitory potency with over 100-fold selectivity against mTOR kinase. The lead compounds 6g and 6k showed good developability profiles in cell-based proliferation and ADME assays. In this communication, their design, synthesis, structure–activity relationship, selectivity, and some developability properties are described. PMID:25893045

  8. The role of PI3K/Akt in human herpesvirus infection: From the bench to the bedside.

    PubMed

    Liu, XueQiao; Cohen, Jeffrey I

    2015-05-01

    The phosphatidylinositol-3-kinase (PI3K)-Akt signaling pathway regulates several key cellular functions including protein synthesis, cell growth, glucose metabolism, and inflammation. Many viruses have evolved mechanisms to manipulate this signaling pathway to ensure successful virus replication. The human herpesviruses undergo both latent and lytic infection, but differ in cell tropism, growth kinetics, and disease manifestations. Herpesviruses express multiple proteins that target the PI3K/Akt cell signaling pathway during the course of their life cycle to facilitate viral infection, replication, latency, and reactivation. Rare human genetic disorders with mutations in either the catalytic or regulatory subunit of PI3K that result in constitutive activation of the protein predispose to severe herpesvirus infections as well as to virus-associated malignancies. Inhibiting the PI3K/Akt pathway or its downstream proteins using drugs already approved for other diseases can block herpesvirus lytic infection and may reduce malignancies associated with latent herpesvirus infections. PMID:25798530

  9. The Role of PI3K/Akt in Human Herpesvirus Infection: from the Bench to the Bedside

    PubMed Central

    Liu, XueQiao; Cohen, Jeffrey I.

    2015-01-01

    The phosphatidylinositol-3-kinase (PI3K)-Akt signaling pathway regulates several key cellular functions including protein synthesis, cell growth, glucose metabolism, and inflammation. Many viruses have evolved mechanisms to manipulate this signaling pathway to ensure successful virus replication. The human herpesviruses undergo both latent and lytic infection, but differ in cell tropism, growth kinetics, and disease manifestations. Herpesviruses express multiple proteins that target the PI3K/Akt cell signaling pathway during the course of their life cycle to facilitate viral infection, replication, latency, and reactivation. Rare human genetic disorders with mutations in either the catalytic or regulatory subunit of PI3K that result in constitutive activation of the protein predispose to severe herpesvirus infections as well as to virus-associated malignancies. Inhibiting the PI3K/Akt pathway or its downstream proteins using drugs already approved for other diseases can block herpesvirus lytic infection and may reduce malignancies associated with latent herpesvirus infections. PMID:25798530

  10. Trisubstituted-Imidazoles Induce Apoptosis in Human Breast Cancer Cells by Targeting the Oncogenic PI3K/Akt/mTOR Signaling Pathway

    PubMed Central

    Mervin, Lewis; Mohan, Surender; Paricharak, Shardul; Baday, Sefer; Li, Feng; Shanmugam, Muthu K.; Chinnathambi, Arunachalam; Zayed, M. E.; Alharbi, Sulaiman Ali; Bender, Andreas; Sethi, Gautam; Basappa; Rangappa, Kanchugarakoppal S.

    2016-01-01

    Overactivation of PI3K/Akt/mTOR is linked with carcinogenesis and serves a potential molecular therapeutic target in treatment of various cancers. Herein, we report the synthesis of trisubstituted-imidazoles and identified 2-chloro-3-(4, 5-diphenyl-1H-imidazol-2-yl) pyridine (CIP) as lead cytotoxic agent. Naïve Base classifier model of in silico target prediction revealed that CIP targets RAC-beta serine/threonine-protein kinase which comprises the Akt. Furthermore, CIP downregulated the phosphorylation of Akt, PDK and mTOR proteins and decreased expression of cyclin D1, Bcl-2, survivin, VEGF, procaspase-3 and increased cleavage of PARP. In addition, CIP significantly downregulated the CXCL12 induced motility of breast cancer cells and molecular docking calculations revealed that all compounds bind to Akt2 kinase with high docking scores compared to the library of previously reported Akt2 inhibitors. In summary, we report the synthesis and biological evaluation of imidazoles that induce apoptosis in breast cancer cells by negatively regulating PI3K/Akt/mTOR signaling pathway. PMID:27097161

  11. STYK1 promotes epithelial-mesenchymal transition and tumor metastasis in human hepatocellular carcinoma through MEK/ERK and PI3K/AKT signaling

    PubMed Central

    Wang, Zhaowen; Qu, Lei; Deng, Biao; Sun, Xing; Wu, Shaohan; Liao, Jianhua; Fan, Junwei; Peng, Zhihai

    2016-01-01

    Serine/threonine/tyrosine kinase 1 (STYK1) is known to be involved in tumor progression. However, its molecular role and mechanism in hepatocellular carcinoma (HCC) remains unknown. We evaluated the effect of STYK1 expression in HCC tissues and investigated the underlying mechanisms associated with progression. HCC tissues expressed greater levels of STYK1 than paired non-tumor tissues. Patients with HCC expressing low levels of STYK1 showed both, greater disease-free (p < 0.0001) and overall (p = 0.0004) survival than those expressing high levels of STYK1. Decreased expression of STYK1 was significantly associated with decreased cell proliferation, reduced migratory capability, and reduced invasive capability. Overexpression of STYK1 was significantly associated with increased cell proliferation, migratory capability, and invasive capability in vitro, as well as increased volume of tumor, weight of tumor, and number of pulmonary metastases in vivo. Furthermore, STYK1’s mechanism of promoting cancer cell mobility and epithelial-mesenchymal transition (EMT) was found to be via the MEK/ERK and PI3K/AKT pathways, resulting in increased expression of mesenchymal protein markers: snail, fibronectin, and vimentin, and decreased E-cadherin expression. Our results suggest that STYK1 acts as an oncogene by inducing cell invasion and EMT via the MEK/ERK and PI3K/AKT signaling pathways and it therefore may be a potential therapeutic target in HCC. PMID:27628214

  12. Extracellular matrix secreted by senescent fibroblasts induced by UVB promotes cell proliferation in HaCaT cells through PI3K/AKT and ERK signaling pathways.

    PubMed

    Kang, Jian; Chen, Wenqi; Xia, Jiping; Li, Yanhua; Yang, Bo; Chen, Bin; Sun, Weiling; Song, Xiuzu; Xiang, Wenzhong; Wang, Xiaoyong; Wang, Fei; Bi, Zhigang; Wan, Yinsheng

    2008-06-01

    Chronic exposure to solar ultraviolet radiation (UV) induces photoaging, and ultimately photocarcinogenesis. Senescent human skin fibroblasts (HSFs) in UVB stress-induced premature senescence (UVB-SIPS) share a similar extracellular matrix (ECM) phenotype with other types of senescent fibroblast. ECM from senescent fibroblasts induced by a variety of stresses has been shown to promote preneoplastic and neoplastic epithelial cell growth, a potential mechanism in carcinogenesis. We undertook this study to explore whether the extracellular matrices from UVB-induced senescent fibroblasts have any effect on the proliferation of HaCaT cells. The results showed that ECM secreted from HSFs in UVB-SIPS has 13.15 and 29.27% more stimulatory effect on proliferation than ECM secreted from presenescent HSFs and non-ECM, respectively. ECM from fibroblasts in UVB-SIPS activates FAK, ERK, and AKT in HaCaT cells. ERK and PI3K/AKT inhibitors inhibit ECM-induced ERK, AKT activation and cell proliferation. Cytochalasin D, a destructive agent of the cytoskeleton, inhibits ECM-induced FAK activation and cell proliferation in HaCaT cells. Collectively, we conclude that ECM secreted from HSFs in UVB-SIPS promotes cell proliferation via ERK and PI3K/AKT pathways and modulation of FAK and cytoskeletal proteins in HaCaT cells. Pharmacological manipulation of those signaling components may lead to the prevention and treatment of skin cancer induced by chronic solar exposure.

  13. GDC-0152 induces apoptosis through down-regulation of IAPs in human leukemia cells and inhibition of PI3K/Akt signaling pathway.

    PubMed

    Hu, Rong; Li, Jia; Liu, Zhuogang; Miao, Miao; Yao, Kun

    2015-02-01

    The inhibitor of apoptosis proteins (IAPs) is closely related to leukemia apoptosis. The present study was undertaken to determine the molecular mechanisms by which GDC-0152, an IAP inhibitor, induces apoptosis in human leukemia cells (K562 and HL60 cells). GDC-0152 inhibited the proliferation of K562 and HL60 cells in a dose- and time-dependent manner, which was largely attributed to intrinsic apoptosis. GDC-0152 down-regulated the IAPs including X-linked inhibitor of apoptosis protein (XIAP), cellular inhibitor of apoptosis protein-1 (cIAP1), and cellular inhibitor of apoptosis protein-2 (cIAP2) expression and induced the activation of caspase-9 and caspase-3. GDC-0152-induced cell proliferation inhibition in K562 cells was prevented by pan-caspase inhibitor. GDC-0152 also inhibited PI3K and Akt expression in K562 and HL60 cells. Taken together, these findings suggest that GDC-0152 results in human leukemia apoptosis through caspase-dependent mechanisms involving down-regulation of IAPs and inhibition of PI3K/Akt signaling.

  14. Overexpression of osteopontin induces angiogenesis of endothelial progenitor cells via the avβ3/PI3K/AKT/eNOS/NO signaling pathway in glioma cells.

    PubMed

    Wang, Yingyi; Yan, Wei; Lu, Xiaoming; Qian, Chunfa; Zhang, Junxia; Li, Ping; Shi, Lei; Zhao, Peng; Fu, Zhen; Pu, Peiyu; Kang, Chunshen; Jiang, Tao; Liu, Ning; You, Yongping

    2011-08-01

    Angiogenesis, a hallmark of tumor growth, is regulated by various angiogenic factors. Recent studies have shown that osteopontin (OPN) is a secreted, integrin-binding protein that contributes to glioma progression. However, its effect on the angiogenesis of gliomas is not fully understood. To elucidate the role of OPN in the process of glioma angiogenesis, endothelial progenitor cells (EPCs) were treated with conditioned media of human glioma SHG44 cells overexpressing OPN. Here, we identified that OPN secreted by glioma cells accelerated EPCs angiogenesis in vitro, including proliferation, migration, and tube formation. OPN also induced the activation of AKT and endothelial nitric oxide synthase (eNOS) and increased NO production without affecting the expression of VEGF, VEGFR-1, or VEGFR-2. Moreover, the avβ3 antibody, the PI3-K inhibitor LY294002 and the eNOS inhibitor NMA suppressed the OPN-mediated increase in NO production and angiogenesis in EPCs. Taken together, these results demonstrate that OPN directly stimulates angiogenesis via the avβ3/PI3-K/AKT/eNOS/NO signaling pathway and may play an important role in tumorigenesis by enhancing angiogenesis in gliomas.

  15. The CLC-2 Chloride Channel Modulates ECM Synthesis, Differentiation, and Migration of Human Conjunctival Fibroblasts via the PI3K/Akt Signaling Pathway

    PubMed Central

    Sun, Lixia; Dong, Yaru; Zhao, Jing; Yin, Yuan; Zheng, Yajuan

    2016-01-01

    Recent evidence suggests that chloride channels are critical for cell proliferation, migration, and differentiation. We examined the effects of transforming growth factor (TGF)-β1 on chloride channel expression and associations with human conjunctival fibroblast (HConF) biology. To investigate the potential role of chloride channel (CLC)-2 in migration, transition to myofibroblasts and extracellular matrix (ECM) synthesis of HconF, a small interfering RNA (siRNA) approach was applied. TGF-β1-induced migration and transition of fibroblasts to myofibroblasts characterized by α-smooth muscle actin (α-SMA) expression, supported by increased endogenous expression of CLC-2 protein and mRNA transcripts. ECM (collagen I and fibronectin) synthesis in HConF was enhanced by TGF-β1. CLC-2 siRNA treatment reduced TGF-β1-induced cell migration, transition of fibroblasts to myofibroblasts, and ECM synthesis of HConF. CLC-2 siRNA treatment in the presence of TGF-β1 inhibited phosphorylation of PI3K and Akt in HConF. These findings demonstrate that CLC-2 chloride channels are important for TGF-β1-induced migration, differentiation, and ECM synthesis via PI3K/Akt signaling in HConF. PMID:27294913

  16. 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

  17. Reactive Oxygen Species via Redox Signaling to PI3K/AKT Pathway Contribute to the Malignant Growth of 4-Hydroxy Estradiol-Transformed Mammary Epithelial Cells

    PubMed Central

    Okoh, Victor O.; Felty, Quentin; Parkash, Jai; Poppiti, Robert; Roy, Deodutta

    2013-01-01

    The purpose of this study was to investigate the effects of 17-β-estradiol (E2)-induced reactive oxygen species (ROS) on the induction of mammary tumorigenesis. We found that ROS-induced by repeated exposures to 4-hydroxy-estradiol (4-OH-E2), a predominant catechol metabolite of E2, caused transformation of normal human mammary epithelial MCF-10A cells with malignant growth in nude mice. This was evident from inhibition of estrogen-induced breast tumor formation in the xenograft model by both overexpression of catalase as well as by co-treatment with Ebselen. To understand how 4-OH-E2 induces this malignant phenotype through ROS, we investigated the effects of 4-OH-E2 on redox-sensitive signal transduction pathways. During the malignant transformation process we observed that 4-OH-E2 treatment increased AKT phosphorylation through PI3K activation. The PI3K-mediated phosphorylation of AKT in 4-OH-E2-treated cells was inhibited by ROS modifiers as well as by silencing of AKT expression. RNA interference of AKT markedly inhibited 4-OH-E2-induced in vitro tumor formation. The expression of cell cycle genes, cdc2, PRC1 and PCNA and one of transcription factors that control the expression of these genes – nuclear respiratory factor-1 (NRF-1) was significantly up-regulated during the 4-OH-E2-mediated malignant transformation process. The increased expression of these genes was inhibited by ROS modifiers as well as by silencing of AKT expression. These results indicate that 4-OH-E2-induced cell transformation may be mediated, in part, through redox-sensitive AKT signal transduction pathways by up-regulating the expression of cell cycle genes cdc2, PRC1 and PCNA, and the transcription factor – NRF-1. In summary, our study has demonstrated that: (i) 4-OH-E2 is one of the main estrogen metabolites that induce mammary tumorigenesis and (ii) ROS-mediated signaling leading to the activation of PI3K/AKT pathway plays an important role in the generation of 4-OH-E2-induced

  18. Diosgenin and 5-Methoxypsoralen Ameliorate Insulin Resistance through ER-α/PI3K/Akt-Signaling Pathways in HepG2 Cells.

    PubMed

    Fang, Ke; Dong, Hui; Jiang, Shujun; Li, Fen; Wang, Dingkun; Yang, Desen; Gong, Jing; Huang, Wenya; Lu, Fuer

    2016-01-01

    To determine the effects and the underlying mechanism of diosgenin (DSG) and 5-methoxypsoralen (5-MOP), two main active components in the classical Chinese prescription Hu-Lu-Ba-Wan (HLBW), on insulin resistance, HepG2 cells were incubated in medium containing insulin. Treatments with DSG, 5-MOP, and their combination were performed, respectively. The result showed that the incubation of HepG2 cells with high concentration insulin markedly decreased glucose consumption and glycogen synthesis. However, treatment with DSG, 5-MOP, or their combination significantly reversed the condition and increased the phosphorylated expression of estrogen receptor-α (ERα), sarcoma (Src), Akt/protein kinase B, glycogen synthase kinase-3β (GSK-3β), and the p85 regulatory subunit of phosphatidylinositol 3-kinase p85 (PI3Kp85). At the transcriptional level, expression of the genes mentioned above also increased except for the negative regulation of GSK-3β mRNA. The increased expression of glucose transport-4 (GLUT-4) was meanwhile observed through immunofluorescence. Nevertheless, the synergistic effect of DSG and 5-MOP on improving glycometabolism was not obvious in the present study. These results suggested that DSG and 5-MOP may improve insulin resistance through an ER-mediated PI3K/Akt activation pathway which may be a new strategy for type 2 diabetes mellitus, especially for women in an estrogen-deficient condition. PMID:27656241

  19. Diosgenin and 5-Methoxypsoralen Ameliorate Insulin Resistance through ER-α/PI3K/Akt-Signaling Pathways in HepG2 Cells

    PubMed Central

    Dong, Hui; Jiang, Shujun; Li, Fen; Wang, Dingkun; Yang, Desen; Gong, Jing; Huang, Wenya

    2016-01-01

    To determine the effects and the underlying mechanism of diosgenin (DSG) and 5-methoxypsoralen (5-MOP), two main active components in the classical Chinese prescription Hu-Lu-Ba-Wan (HLBW), on insulin resistance, HepG2 cells were incubated in medium containing insulin. Treatments with DSG, 5-MOP, and their combination were performed, respectively. The result showed that the incubation of HepG2 cells with high concentration insulin markedly decreased glucose consumption and glycogen synthesis. However, treatment with DSG, 5-MOP, or their combination significantly reversed the condition and increased the phosphorylated expression of estrogen receptor-α (ERα), sarcoma (Src), Akt/protein kinase B, glycogen synthase kinase-3β (GSK-3β), and the p85 regulatory subunit of phosphatidylinositol 3-kinase p85 (PI3Kp85). At the transcriptional level, expression of the genes mentioned above also increased except for the negative regulation of GSK-3β mRNA. The increased expression of glucose transport-4 (GLUT-4) was meanwhile observed through immunofluorescence. Nevertheless, the synergistic effect of DSG and 5-MOP on improving glycometabolism was not obvious in the present study. These results suggested that DSG and 5-MOP may improve insulin resistance through an ER-mediated PI3K/Akt activation pathway which may be a new strategy for type 2 diabetes mellitus, especially for women in an estrogen-deficient condition.

  20. Diosgenin and 5-Methoxypsoralen Ameliorate Insulin Resistance through ER-α/PI3K/Akt-Signaling Pathways in HepG2 Cells

    PubMed Central

    Dong, Hui; Jiang, Shujun; Li, Fen; Wang, Dingkun; Yang, Desen; Gong, Jing; Huang, Wenya

    2016-01-01

    To determine the effects and the underlying mechanism of diosgenin (DSG) and 5-methoxypsoralen (5-MOP), two main active components in the classical Chinese prescription Hu-Lu-Ba-Wan (HLBW), on insulin resistance, HepG2 cells were incubated in medium containing insulin. Treatments with DSG, 5-MOP, and their combination were performed, respectively. The result showed that the incubation of HepG2 cells with high concentration insulin markedly decreased glucose consumption and glycogen synthesis. However, treatment with DSG, 5-MOP, or their combination significantly reversed the condition and increased the phosphorylated expression of estrogen receptor-α (ERα), sarcoma (Src), Akt/protein kinase B, glycogen synthase kinase-3β (GSK-3β), and the p85 regulatory subunit of phosphatidylinositol 3-kinase p85 (PI3Kp85). At the transcriptional level, expression of the genes mentioned above also increased except for the negative regulation of GSK-3β mRNA. The increased expression of glucose transport-4 (GLUT-4) was meanwhile observed through immunofluorescence. Nevertheless, the synergistic effect of DSG and 5-MOP on improving glycometabolism was not obvious in the present study. These results suggested that DSG and 5-MOP may improve insulin resistance through an ER-mediated PI3K/Akt activation pathway which may be a new strategy for type 2 diabetes mellitus, especially for women in an estrogen-deficient condition. PMID:27656241

  1. Involvement of HDAC1 and the PI3K/PKC signaling pathways in NF-{kappa}B activation by the HDAC inhibitor apicidin

    SciTech Connect

    Kim, Yong Kee . E-mail: yksnbk@kwandong.ac.kr; Seo, Dong-Wan; Kang, Dong-Won; Lee, Hoi Young; Han, Jeung-Whan; Kim, Su-Nam . E-mail: snkim@kist.re.kr

    2006-09-08

    Histone deacetylase (HDAC) inhibitors are appreciated as one of promising anticancer drugs, but they exert differential responses depending on the cell type. We recently reported the critical role of NF-{kappa}B as a modulator in determining cell fate for apoptosis in response to an HDAC inhibitor. In this study, we investigate a possible signaling pathway required for NF-{kappa}B activation in response to the HDAC inhibitor apicidin. Treatment of HeLa cells with apicidin leads to an increase in transcriptional activity of NF-{kappa}B and the expression of its target genes, IL-8 and TNF-{alpha}. TNF-{alpha} expression by apicidin is induced at earlier time points than NF-{kappa}B activation or IL-8 expression. In addition, our data show that the early expression of TNF-{alpha} does not lead to activation of NF-{kappa}B, because disruption of TNF-{alpha} activity by a neutralizing antibody does not affect nuclear translocation of NF-{kappa}B, I{kappa}B{alpha} degradation or reporter gene activation by apicidin. However, this activation of NF-{kappa}B requires the PI3K and PKC signaling pathways, but not ERK or JNK. Furthermore, apicidin activation of NF-{kappa}B seems to result from HDAC1 inhibition, as evidenced by the observation that overexpression of HDAC1, but not HDAC2, 3 or 4, dramatically inhibits NF-{kappa}B reporter gene activity. Collectively, our results suggest that activation of NF-{kappa}B signaling by apicidin requires both the PI3K/PKC signaling pathways and HDAC1, and functions as a critical modulator in determining the cellular effect of apicidin.

  2. Anti-inflammatory effects of egg white combined with chalcanthite in lipopolysaccharide-stimulated BV2 microglia through the inhibition of NF-κB, MAPK and PI3K/Akt signaling pathways.

    PubMed

    Choi, Eun A; Park, Hye Young; Yoo, Hwa-Seung; Choi, Yung Hyun

    2013-01-01

    Egg white-chalcanthite (EWCC) is a mixture of egg white and chalcanthite prepared by roasting chalcanthite (which is a natural mineral mainly composed of CuSO4•5H2O) to the point of dehydration, pulverizing the dehydrated chalcanthite and then mixing the pulverized chalcanthite to react with egg white to trigger a reaction. When egg white-chalcanthite is prepared in this manner, the toxicity of chalcanthite is neutralized by the egg white, so that the toxicity is reduced or removed and the pharmaceutical properties are increased. However, the cellular and molecular mechanisms underlying the pharmacological activity of EWCC remain poorly understood. In this study, we investigated the inhibitory effects of EWCC on the production of lipopolysaccharide (LPS)-induced pro-inflammatory mediators in BV2 microglia. Our data indicated that the EWCC treatment significantly inhibited the excessive production of nitric oxide and prostaglandin E2 in LPS-stimulated BV2 microglia in a concentration-dependent manner without causing cytotoxicity. It also attenuated the expression of inducible nitric oxide synthase, cyclooxygenase-2 and pro-inflammatory cytokines, including interleukin-1β and tumor necrosis factor-α. Moreover, EWCC exhibited anti-inflammatory properties by the suppression of nuclear factor‑κB (NF-κB) activation by blocking IκB-α degradation, downregulation of extracellular signal-regulated kinase, c-Jun N-terminal kinase, p38 mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K)/Akt pathways. Our results indicate that the inhibitory effects of EWCC on LPS-stimulated inflammatory mediator production in BV2 microglia are associated with the suppression of the NF-κB, MAPK and PI3K/Akt signaling pathways. These findings suggest that EWCC may offer a substantial therapeutic potential for the treatment of neurodegenerative diseases that are accompanied by microglial activation. PMID:23128312

  3. Targeting PI3 kinase/AKT/mTOR signaling in cancer.

    PubMed

    Sheppard, Karen; Kinross, Kathryn M; Solomon, Benjamin; Pearson, Richard B; Phillips, Wayne A

    2012-01-01

    The phosphatidylinositol 3 kinase (PI3K) pathway is one of the major pathways modulating cell growth, proliferation, metabolism, survival, and angiogenesis. Hyperactivation of this pathway is one of the most frequent occurrences in human cancer and is thus an obvious target for treatment of this disease. Currently there are 26 novel compounds targeting the PI3K pathway being assessed in more than 150 cancer-related clinical trials. Although this pathway is involved in many vital biologic functions, data emanating from these clinical trials indicate that these drugs are well tolerated. This review outlines the interaction of the PI3K pathway with other signaling cascades, highlights mechanisms involved in hyperactivation, discusses current therapeutics in cancer-related clinical trials that target this pathway, and, based on preclinical data, discusses possible leads on patient selection and combinational therapy, including targeting multiple components of the associated signaling network.

  4. Cancer Associated Fibroblast-Derived Hepatocyte Growth Factor Inhibits the Paclitaxel-Induced Apoptosis of Lung Cancer A549 Cells by Up-Regulating the PI3K/Akt and GRP78 Signaling on a Microfluidic Platform.

    PubMed

    Ying, Li; Zhu, Ziwei; Xu, Zhiyun; He, Tianrui; Li, Encheng; Guo, Zhe; Liu, Fen; Jiang, Chunmeng; Wang, Qi

    2015-01-01

    Tumor stroma and growth factors provide a survival environment to tumor cells and can modulate their chemoresistance by dysregulating several signal pathways. In this study, we fabricated a three-dimensional (3D) microfluidic chip using polydimethylsiloxane (PDMS) to investigate the impact of hepatocyte growth factor (HGF) from cancer-associated fibroblasts (CAF) on the Met/PI3K/AKT activation, glucose regulatory protein (GRP78) expression and the paclitaxel-induced A549 cell apoptosis. With a concentration gradient generator, the assembled chip was able to reconstruct a tumor microenvironment in vitro. We found high levels of HGF in the supernatants of CAF and the CAF matrix from the supernatants of activated HFL1 fibroblasts or HGF enhanced the levels of Met, PI3K and AKT phosphorylation and GRP78 expression in A549 cells cultured in a 3D cell chamber, which was abrogated by anti-HGF. Inhibition of Met attenuated the CAF matrix-enhanced PI3K/AKT phosphorylation and GRP78 expression while inhibition of PI3K reduced GRP78 expression, but not Met phosphorylation in A549 cells. Inhibition of GRP78 failed to modulate the CAF matrix-enhanced Met/PI3K/AKT phosphorylation in A549 cells. Furthermore, inhibition of PI3K or GRP78 enhanced spontaneous and paclitaxel-induced A549 cell apoptosis. Moreover, treatment with the CAF matrix inhibited spontaneous and medium or high dose of paclitaxel-induced A549 cell apoptosis. Inhibition of PI3K or GRP78 attenuated the CAF matrix-mediated inhibition on paclitaxel-induced A549 cell apoptosis. Our data indicated that HGF in the CAF matrix activated the Met/PI3K/AKT and up-regulated GRP78 expression, promoting chemoresistance to paclitaxel-mediated apoptosis in A549 cells. Our findings suggest that the microfluidic system may represent an ideal platform for signaling research and drug screening. PMID:26115510

  5. Cancer Associated Fibroblast-Derived Hepatocyte Growth Factor Inhibits the Paclitaxel-Induced Apoptosis of Lung Cancer A549 Cells by Up-Regulating the PI3K/Akt and GRP78 Signaling on a Microfluidic Platform

    PubMed Central

    Xu, Zhiyun; He, Tianrui; Li, Encheng; Guo, Zhe; Liu, Fen; Jiang, Chunmeng; Wang, Qi

    2015-01-01

    Tumor stroma and growth factors provide a survival environment to tumor cells and can modulate their chemoresistance by dysregulating several signal pathways. In this study, we fabricated a three-dimensional (3D) microfluidic chip using polydimethylsiloxane (PDMS) to investigate the impact of hepatocyte growth factor (HGF) from cancer-associated fibroblasts (CAF) on the Met/PI3K/AKT activation, glucose regulatory protein (GRP78) expression and the paclitaxel-induced A549 cell apoptosis. With a concentration gradient generator, the assembled chip was able to reconstruct a tumor microenvironment in vitro. We found high levels of HGF in the supernatants of CAF and the CAF matrix from the supernatants of activated HFL1 fibroblasts or HGF enhanced the levels of Met, PI3K and AKT phosphorylation and GRP78 expression in A549 cells cultured in a 3D cell chamber, which was abrogated by anti-HGF. Inhibition of Met attenuated the CAF matrix-enhanced PI3K/AKT phosphorylation and GRP78 expression while inhibition of PI3K reduced GRP78 expression, but not Met phosphorylation in A549 cells. Inhibition of GRP78 failed to modulate the CAF matrix-enhanced Met/PI3K/AKT phosphorylation in A549 cells. Furthermore, inhibition of PI3K or GRP78 enhanced spontaneous and paclitaxel-induced A549 cell apoptosis. Moreover, treatment with the CAF matrix inhibited spontaneous and medium or high dose of paclitaxel-induced A549 cell apoptosis. Inhibition of PI3K or GRP78 attenuated the CAF matrix-mediated inhibition on paclitaxel-induced A549 cell apoptosis. Our data indicated that HGF in the CAF matrix activated the Met/PI3K/AKT and up-regulated GRP78 expression, promoting chemoresistance to paclitaxel-mediated apoptosis in A549 cells. Our findings suggest that the microfluidic system may represent an ideal platform for signaling research and drug screening. PMID:26115510

  6. 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.

  7. 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

  8. 6″-Debromohamacanthin A, a Bis (Indole) Alkaloid, Inhibits Angiogenesis by Targeting the VEGFR2-Mediated PI3K/AKT/mTOR Signaling Pathways

    PubMed Central

    Kim, Gi Dae; Cheong, Oug Jae; Bae, Song Yi; Shin, Jongheon; Lee, Sang Kook

    2013-01-01

    Hamacanthins, bis (indole) alkaloids, are found in a few marine sponges, including Spongosorites sp. Hamacanthins have been shown to possess cytotoxic, antibacterial and antifungal activities. However, the precise mechanism for the biological activities of hamacanthins has not yet been elucidated. In the present study, the anti-angiogenic effects of 6″-debromohamacanthin A (DBHA), an active component of isolated hamacanthins, were evaluated in cultured human umbilical vascular endothelial cells (HUVEC) and endothelial-like cells differentiated from mouse embryonic stem (mES) cells. DBHA significantly inhibited vascular endothelial growth factor (VEGF)-induced cell proliferation, migration and tube formation in the HUVEC. DBHA also suppressed the capillary-like structure formation and the expression of platelet endothelial cell adhesion molecule (PECAM), an endothelial biomarker, in mES cell-derived endothelial-like cells. To further understand the precise molecular mechanism of action, VEGF-mediated signaling pathways were analyzed in HUVEC cells and mES cell-derived endothelial-like cells. DBHA suppressed the VEGF-induced expression of MAPKs (p38, ERK and SAPK/JNK) and the PI3K/AKT/mTOR signaling pathway. In addition, DBHA inhibited microvessel sprouting in mES/EB-derived embryoid bodies. In an ex vivo model, DBHA also suppressed the microvessel sprouting of mouse aortic rings. The findings suggest for the first time that DBHA inhibits angiogenesis by targeting the vascular endothelial growth factor receptor 2 (VEGFR2)-mediated PI3K/AKT/mTOR signaling pathway in endothelial cells. PMID:23549281

  9. Eupatilin inhibits EGF-induced JB6 cell transformation by targeting PI3K.

    PubMed

    Li, Feng; Tao, Ya; Qiao, Yan; Li, Ke; Jiang, Yanan; Cao, Chang; Ren, Shuxin; Chang, Xiaobin; Wang, Xiaona; Wang, Yanhong; Xie, Yifei; Dong, Ziming; Zhao, Jimin; Liu, Kangdong

    2016-09-01

    Phosphatidylinositol 3-kinases (PI3Ks) are lipid kinases that play fundamental roles in regulation of multiple signaling pathways, including cell proliferation, survival and cell cycle. Increasing evidence has shown that abnormal activation of PI3K pathway contributes to tumorigenesis and progression of various malignant tumors. Therefore, it is an attractive target of chemoprevention and chemotherapy. Eupatilin, a natural flavone compound extracted from Artemisia vulgaris, has antitumor and anti-inflammation efficacy. However, the direct target(s) of eupatilin in cancer chemoprevention are still elusive. In the present study, we reported eupatilin suppressed JB6 cell proliferation and its EGF-induced colony formation. Eupatilin attenuated phosphorylation of PI3K downstream signaling molecules. Downregulation of cyclin D1 expression and arresting in G1 phase were induced through eupatilin treatment. Furthermore, we found it could bind to the p110α, a catalytic subunit of PI3K, by computational docking methods. Pull down assay outcomes also verified the binding of eupatilin with PI3K. Taken together, our results suggest that epatilin is a potential chemopreventive agent in inhibition of skin cell transformation by targeting PI3K. PMID:27573489

  10. The CB1 cannabinoid receptor signals striatal neuroprotection via a PI3K/Akt/mTORC1/BDNF pathway

    PubMed Central

    Blázquez, C; Chiarlone, A; Bellocchio, L; Resel, E; Pruunsild, P; García-Rincón, D; Sendtner, M; Timmusk, T; Lutz, B; Galve-Roperh, I; Guzmán, M

    2015-01-01

    The CB1 cannabinoid receptor, the main molecular target of endocannabinoids and cannabis active components, is the most abundant G protein-coupled receptor in the mammalian brain. In particular, the CB1 receptor is highly expressed in the basal ganglia, mostly on terminals of medium-sized spiny neurons, where it plays a key neuromodulatory function. The CB1 receptor also confers neuroprotection in various experimental models of striatal damage. However, the assessment of the physiological relevance and therapeutic potential of the CB1 receptor in basal ganglia-related diseases is hampered, at least in part, by the lack of knowledge of the precise mechanism of CB1 receptor neuroprotective activity. Here, by using an array of pharmacological, genetic and pharmacogenetic (designer receptor exclusively activated by designer drug) approaches, we show that (1) CB1 receptor engagement protects striatal cells from excitotoxic death via the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin complex 1 pathway, which, in turn, (2) induces brain-derived neurotrophic factor (BDNF) expression through the selective activation of BDNF gene promoter IV, an effect that is mediated by multiple transcription factors. To assess the possible functional impact of the CB1/BDNF axis in a neurodegenerative-disease context in vivo, we conducted experiments in the R6/2 mouse, a well-established model of Huntington's disease, in which the CB1 receptor and BDNF are known to be severely downregulated in the dorsolateral striatum. Adeno-associated viral vector-enforced re-expression of the CB1 receptor in the dorsolateral striatum of R6/2 mice allowed the re-expression of BDNF and the concerted rescue of the neuropathological deficits in these animals. Collectively, these findings unravel a molecular link between CB1 receptor activation and BDNF expression, and support the relevance of the CB1/BDNF axis in promoting striatal neuron survival. PMID:25698444

  11. The CB₁ cannabinoid receptor signals striatal neuroprotection via a PI3K/Akt/mTORC1/BDNF pathway.

    PubMed

    Blázquez, C; Chiarlone, A; Bellocchio, L; Resel, E; Pruunsild, P; García-Rincón, D; Sendtner, M; Timmusk, T; Lutz, B; Galve-Roperh, I; Guzmán, M

    2015-10-01

    The CB1 cannabinoid receptor, the main molecular target of endocannabinoids and cannabis active components, is the most abundant G protein-coupled receptor in the mammalian brain. In particular, the CB1 receptor is highly expressed in the basal ganglia, mostly on terminals of medium-sized spiny neurons, where it plays a key neuromodulatory function. The CB1 receptor also confers neuroprotection in various experimental models of striatal damage. However, the assessment of the physiological relevance and therapeutic potential of the CB1 receptor in basal ganglia-related diseases is hampered, at least in part, by the lack of knowledge of the precise mechanism of CB1 receptor neuroprotective activity. Here, by using an array of pharmacological, genetic and pharmacogenetic (designer receptor exclusively activated by designer drug) approaches, we show that (1) CB1 receptor engagement protects striatal cells from excitotoxic death via the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin complex 1 pathway, which, in turn, (2) induces brain-derived neurotrophic factor (BDNF) expression through the selective activation of BDNF gene promoter IV, an effect that is mediated by multiple transcription factors. To assess the possible functional impact of the CB1/BDNF axis in a neurodegenerative-disease context in vivo, we conducted experiments in the R6/2 mouse, a well-established model of Huntington's disease, in which the CB1 receptor and BDNF are known to be severely downregulated in the dorsolateral striatum. Adeno-associated viral vector-enforced re-expression of the CB1 receptor in the dorsolateral striatum of R6/2 mice allowed the re-expression of BDNF and the concerted rescue of the neuropathological deficits in these animals. Collectively, these findings unravel a molecular link between CB1 receptor activation and BDNF expression, and support the relevance of the CB1/BDNF axis in promoting striatal neuron survival.

  12. Carvacrol Alleviates Prostate Cancer Cell Proliferation, Migration, and Invasion through Regulation of PI3K/Akt and MAPK Signaling Pathways

    PubMed Central

    Luo, Yun; Wu, Jie-Ying; Lu, Min-Hua; Shi, Zhi

    2016-01-01

    TRPM7 is a potential therapeutic target for treatment of prostate cancer. In this study, we investigated the effects of nonselective TRPM7 inhibitor carvacrol on cell proliferation, migration, and invasion of prostate cancer PC-3 and DU145 cells. Our results showed that carvacrol blocked TRPM7-like currents in PC-3 and DU145 cells and reduced their proliferation, migration, and invasion. Moreover, carvacrol treatment significantly decreased MMP-2, p-Akt, and p-ERK1/2 protein expression and inhibited F-actin reorganization. Furthermore, consistently, TRPM7 knockdown reduced prostate cancer cell proliferation, migration, and invasion as well. Our study suggests that carvacrol may have therapeutic potential for the treatment of prostate cancer through its inhibition of TRPM7 channels and suppression of PI3K/Akt and MAPK signaling pathways. PMID:27803760

  13. Anti-diabetic effect of citrus pectin in diabetic rats and potential mechanism via PI3K/Akt signaling pathway.

    PubMed

    Liu, Yanlong; Dong, Man; Yang, Ziyu; Pan, Siyi

    2016-08-01

    This study was performed to investigate the anti-diabetic effect of citrus pectin in type 2 diabetic rats and its potential mechanism of action. The results showed that fasting blood glucose levels were significantly decreased after 4 weeks of citrus pectin administration. Citrus pectin improved glucose tolerance, hepatic glycogen content and blood lipid levels (TG, TC, LDL-c and HDL-c) in diabetic rats. Citrus pectin also significantly reduced insulin resistance, which played an important role in the resulting anti-diabetic effect. Moreover, after the pectin treatment, phosphorylated Akt expression was upregulated and GSK3β expression was downregulated, indicating that the potential anti-diabetic mechanism of citrus pectin might occur through regulation of the PI3K/Akt signaling pathway. Together, these results suggested that citrus pectin could ameliorate type 2 diabetes and potentially be used as an adjuvant treatment.

  14. Suppression of Nkx3.2 by phosphatidylinositol-3-kinase signaling regulates cartilage development by modulating chondrocyte hypertrophy

    PubMed Central

    Kim, Jeong-Ah; Im, Suhjean; Cantley, Lewis C.; Kim, Dae-Won

    2016-01-01

    Phosphatidylinositol-3-kinase (PI3K) is a key regulator of diverse biological processes including cell proliferation, migration, survival, and differentiation. While a role of PI3K in chondrocyte differentiation has been suggested, its precise mechanisms of action are poorly understood. Here we show that PI3K signaling can down-regulate Nkx3.2 at both mRNA and protein levels in various chondrocyte cultures in vitro. In addition, we have intriguingly found that p85β, not p85α, is specifically employed as a regulatory subunit for PI3K-mediated Nkx3.2 suppression. Furthermore, we found that regulation of Nkx3.2 by PI3K requires Rac1–PAK1, but not Akt, signaling downstream of PI3K. Finally, using embryonic limb bud cultures, ex vivo long bone cultures, and p85β knockout mice, we demonstrated that PI3K-mediated suppression of Nkx3.2 in chondrocytes plays a role in the control of cartilage hypertrophy during skeletal development in vertebrates. PMID:26363466

  15. A Genome-Wide RNAi Screen Identifies FOXO4 as a Metastasis-Suppressor through Counteracting PI3K/AKT Signal Pathway in Prostate Cancer

    PubMed Central

    Su, Bing; Gao, Lingqiu; Baranowski, Catherine; Gillard, Bryan; Wang, Jianmin; Ransom, Ryan; Ko, Hyun-Kyung; Gelman, Irwin H.

    2014-01-01

    Activation of the PI3K/AKT signal pathway is a known driving force for the progression to castration-recurrent prostate cancer (CR-CaP), which constitutes the major lethal phenotype of CaP. Here, we identify using a genomic shRNA screen the PI3K/AKT-inactivating downstream target, FOXO4, as a potential CaP metastasis suppressor. FOXO4 protein levels inversely correlate with the invasive potential of a panel of human CaP cell lines, with decreased mRNA levels correlating with increased incidence of clinical metastasis. Knockdown (KD) of FOXO4 in human LNCaP cells causes increased invasion in vitro and lymph node (LN) metastasis in vivo without affecting indices of proliferation or apoptosis. Increased Matrigel invasiveness was found by KD of FOXO1 but not FOXO3. Comparison of differentially expressed genes affected by FOXO4-KD in LNCaP cells in culture, in primary tumors and in LN metastases identified a panel of upregulated genes, including PIP, CAMK2N1, PLA2G16 and PGC, which, if knocked down by siRNA, could decrease the increased invasiveness associated with FOXO4 deficiency. Although only some of these genes encode FOXO promoter binding sites, they are all RUNX2-inducible, and RUNX2 binding to the PIP promoter is increased in FOXO4-KD cells. Indeed, the forced expression of FOXO4 reversed the increased invasiveness of LNCaP/shFOXO4 cells; the forced expression of FOXO4 did not alter RUNX2 protein levels, yet it decreased RUNX2 binding to the PIP promoter, resulting in PIP downregulation. Finally, there was a correlation between FOXO4, but not FOXO1 or FOXO3, downregulation and decreased metastasis-free survival in human CaP patients. Our data strongly suggest that increased PI3K/AKT-mediated metastatic invasiveness in CaP is associated with FOXO4 loss, and that mechanisms to induce FOXO4 re-expression might suppress CaP metastatic aggressiveness. PMID:24983969

  16. Discovery of GSK2126458, a Highly Potent Inhibitor of PI3K and the Mammalian Target of Rapamycin

    SciTech Connect

    Knight, Steven D.; Adams, Nicholas D.; Burgess, Joelle L.; Chaudhari, Amita M.; Darcy, Michael G.; Donatelli, Carla A.; Luengo, Juan I.; Newlander, Ken A.; Parrish, Cynthia A.; Ridgers, Lance H.; Sarpong, Martha A.; Schmidt, Stanley J.; Aller, Glenn S.Van; Carson, Jeffrey D.; Diamond, Melody A.; Elkins, Patricia A.; Gardiner, Christine M.; Garver, Eric; Gilbert, Seth A.; Gontarek, Richard R.; Jackson, Jeffrey R.; Kershner, Kevin L.; Luo, Lusong; Raha, Kaushik; Sherk, Christian S.; Sung, Chiu-Mei; Sutton, David; Tummino, Peter J.; Wegrzyn, Ronald J.; Auger, Kurt R.; Dhanak, Dashyant

    2010-09-30

    Phosphoinositide 3-kinase {alpha} (PI3K{alpha}) is a critical regulator of cell growth and transformation, and its signaling pathway is the most commonly mutated pathway in human cancers. The mammalian target of rapamycin (mTOR), a class IV PI3K protein kinase, is also a central regulator of cell growth, and mTOR inhibitors are believed to augment the antiproliferative efficacy of PI3K/AKT pathway inhibition. 2,4-Difluoro-N-{l_brace}2-(methyloxy)-5-[4-(4-pyridazinyl)-6-quinolinyl]-3-pyridinyl{r_brace}benzenesulfonamide (GSK2126458, 1) has been identified as a highly potent, orally bioavailable inhibitor of PI3K{alpha} and mTOR with in vivo activity in both pharmacodynamic and tumor growth efficacy models. Compound 1 is currently being evaluated in human clinical trials for the treatment of cancer.

  17. Hepatocyte growth factor promotes proliferation, invasion, and metastasis of myeloid leukemia cells through PI3K-AKT and MAPK/ERK signaling pathway

    PubMed Central

    Guo, Jiang-Rui; Li, Wei; Wu, Yong; Wu, Lin-Qing; Li, Xin; Guo, Ya-Fei; Zheng, Xiao-Hui; Lian, Xiao-Lan; Huang, Hui-Fang; Chen, Yuan-Zhong

    2016-01-01

    This study aims to investigate effects of HGF expression on biological behaviors of Kasumi-1 and HL60. Expression of HGF and c-Met gene were detected using qRT-PCR. Short hairpin RNA (shRNA) was used to reduce HGF expression. Silencing effect of shRNA was verified by qRT-PCR and western blot. Cell reproductive capacity, cell clonality and cell cycle (apoptosis) were detected by CCK-8, clone formation, flow cytometry (FCM), respectively. Cell adhesion, cell invasion ability and cell proliferation were also examined. Changes of PI3K-AKT, MAPK/ERK signaling factors were detected by western blot. HGF and c-Met expression in first-vist AML group was significantly higher than in AML-relief and normal control group. HGF shRNA can inhibit cell proliferation, inhibit cloning ability. Compared with control group, apoptosis ratios of Kasumi-1 and HL60 cell in interference groups were significantly higher. After shRNA interference, the number of adherent cells and transmembrane cells were significantly decreased compared with control group. Meanwhile, shRNA also down-regulated Bad, Bcl-XL, Bcl-2, CDK1, Cyclin B, MMP2, MMP9, and up-regulated cleaved caspase9, cleaved caspase3, cleaved PARP, Bax, and P21. Moreover, phosphorylated c-Met, AKT, Erk, and mTOR were also reduced. In conclusion, HGF and c-Met gene highly expressed among first-visit AML patients, but decreased after relief treatment. HGF may promote proliferation, invasion, and metastasis of AML cells through PI3K-AKT and MAPK/ERK signaling pathway. Therefore, proliferation and invasion ability of AML cell can be inhibited by down-regulating HGF gene to retardate cell in G2/M stage. PMID:27725846

  18. Expression of human telomerase reverse transcriptase mediates the senescence of mesenchymal stem cells through the PI3K/AKT signaling pathway.

    PubMed

    Zhao, Qiang; Wang, Xue-Yao; Yu, Xiao-Xia; Zhai, Ying-Xian; He, Xu; Wu, Shan; Shi, Ying-Ai

    2015-09-01

    Multipotent mesenchymal stem cells (MSCs) are widely used as seed cells in studies of tissue engineering and regenerative medicine; however, their clinical application is limited due to replicative senescence. It has been demonstrated that telomerase expression extends the lifespan and maintains the bone-forming ability of MSCs; however, the detailed role and the underlying molecular mechanisms in MSCs remain largely unknown. In the present study, we found that senescence was associated with human telomerase reverse transcriptase (hTERT) expression, and telomere length and telomerase activity. We established a short interfering RNA (siRNA) targeting hTERT and a gene expression vector carrying hTERT and transfected these into the MSCs to investigate the detailed role and the underlying molecular mechanisms of action of hTERT in MSCs. We found that the downregulation of hTERT by siRNA markedly decreased telomere length and telomerase activity in the MSCs, whereas the overexpression of hTERT increased telomere length and telomerase activity in the MSCs. The downregulation of hTERT inhibited cell proliferation and promoted the senescence and apoptosis of MSCs, whereas the upregulation of hTERT increased cell proliferation and decreased the senescence and apoptosis of MSCs. Of note, we also found that the activation of the PI3K/AKT signaling pathway was mediated by hTERT and that blocking this pathway using LY294002 inhibited hTERT expression, induced senescence and decreased the proliferation of MSCs. These findings reveal a previously unknown regulatory mechanism of hTERT, indicating that hTERT mediates the senescence of MSCs through the PI3K/AKT signaling pathway. PMID:26178664

  19. Helicobacter pylori VacA-induced Inhibition of GSK3 through the PI3K/Akt Signaling Pathway*S⃞

    PubMed Central

    Nakayama, Masaaki; Hisatsune, Junzo; Yamasaki, Eiki; Isomoto, Hajime; Kurazono, Hisao; Hatakeyama, Masanori; Azuma, Takeshi; Yamaoka, Yoshio; Yahiro, Kinnosuke; Moss, Joel; Hirayama, Toshiya

    2009-01-01

    Helicobacter pylori VacA toxin contributes to the pathogenesis and severity of gastric injury. We found that incubation of AZ-521 cells with VacA resulted in phosphorylation of protein kinase B (Akt) and glycogen synthase kinase-3β (GSK3β) through a PI3K-dependent pathway. Following phosphorylation and inhibition of GSK3β,β-catenin was released from a GSK3β/β-catenin complex, with subsequent nuclear translocation. Methyl-β-cyclodextrin (MCD) and phosphatidylinositol-specific phospholipase C (PI-PLC), but not 5-nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB) and bafilomycin A1, inhibited VacA-induced phosphorylation of Akt, indicating that it does not require VacA internalization and is independent of vacuolation. VacA treatment of AZ-521 cells transfected with TOPtkLuciferase reporter plasmid or control FOPtkLucifease reporter plasmid resulted in activation of TOPtkLuciferase, but not FOPtkLucifease. In addition, VacA transactivated the β-catenin-dependent cyclin D1 promoter in a luciferase reporter assay. Infection of AZ-521 cells by a vacA mutant strain of H. pylori failed to induce phosphorylation of Akt and GSK3β, or release of β-catenin from a GSK3β/β-catenin complex. Taken together, these results support the conclusion that VacA activates the PI3K/Akt signaling pathway, resulting in phosphorylation and inhibition of GSK3β, and subsequent translocation ofβ-catenin to the nucleus, consistent with effects of VacA on β-catenin-regulated transcriptional activity. These data introduce the possibility that Wnt-dependent signaling might play a role in the pathogenesis of H. pylori infection, including the development of gastric cancer. PMID:18996844

  20. PI3K/Akt Signaling Pathway Modulates Influenza Virus Induced Mouse Alveolar Macrophage Polarization to M1/M2b

    PubMed Central

    Zhao, Xiangfeng; Dai, Jianping; Xiao, Xuejun; Wu, Liqi; Zeng, Jun; Sheng, Jiangtao; Su, Jinghua; Chen, Xiaoxuan; Wang, Gefei; Li, Kangsheng

    2014-01-01

    Macrophages polarized to M1 (pro-inflammation) or M2 (anti-inflammation) phenotypes in response to environmental signals. In this study, we examined the polarization of alveolar macrophage (AM), following induction by different influenza virus strains (ST169 (H1N1), ST602 (H3N2) and HKG9 (H9N2)). Macrophages from other tissues or cell line exert alternative responding pattern, and AM is necessary for investigating the respiratory system. AM polarized toward the M1 phenotype after 4 hours of infection by all three virus strains, and AM to presented M2b phenotype after 8 hours induction, and immunosuppressive phenotype after 24 hours of induction. Protein expression assay showed similar results as the gene expression analysis for phenotype verification. The ELISA assay showed that TNF-α secretion was up-regulated after 4 and 8 hours of infection by influenza viruses, and it returned to basal levels after 24 hours of infection. IL-10 expression was elevated after 8 and 24 hours of infection. Immunofluorescence showed that iNOS expression was up-regulated but not Arg1 expression. Influenza virus notably increased phospho-Akt but not phospho-Erk1/2 or phospho-p38, and the AM polarization pattern have been changed by LY294002 (PI3K inhibitor). In conclusion, our results demonstrate the dynamic polarization of AM induced by influenza viruses, and suggested that PI3K/Akt signaling pathway modulates AM polarization to M1/M2b. PMID:25105760

  1. PIK3CAH1047R and Her2 initiated mammary tumors escape PI3K dependency by compensatory activation of MEK-ERK signaling

    PubMed Central

    Cheng, Hailing; Liu, Pixu; Ohlson, Carolynn; Xu, Erbo; Symonds, Lynn; Isabella, Adam; Muller, William J.; Lin, Nancy U.; Krop, Ian E.; Roberts, Thomas M.; Winer, Eric P.; Arteaga, Carlos L.; Zhao, Jean J.

    2015-01-01

    Human breast cancers that have HER2 amplification/overexpression frequently carry PIK3CA mutations, and are often associated with a worse prognosis. However, the role of PIK3CA mutations in the initiation and maintenance of these breast cancers remains elusive. In the present study, we generated a compound mouse model that genetically mimics HER2 positive breast cancer with coexisting PIK3CAH1047R. Induction of PIK3CAH1047R expression in mouse mammary glands with constitutive expression of activated Her2/Neu resulted in accelerated mammary tumorigenesis with enhanced metastatic potential. Interestingly, inducible expression of mutant PIK3CA resulted in a robust activation of PI3K/AKT signaling but attenuation of Her2/Her3 signaling, and this can be reversed by deinduction of PIK3CAH1047R expression. Strikingly, while these Her2+ PIK3CAH1047R initiated primary mammary tumors are refractory to HER2-targeted therapy, all tumors responded to inactivation of the oncogenic PIK3CAH1047R, a situation closely mimicking the use of a highly effective inhibitor specifically targeting the mutant PIK3CA/p110a. Notably, these tumors eventually resumed growth, and a fraction of them escaped PI3K dependence by compensatory ERK activation, which can be blocked by combined inhibition of Her2 and MEK. Together, these results suggest that PIK3CA-specific inhibition as a monotherapy followed by combination therapy targeting MAPK and HER2 in a timely manner may be an effective treatment approach against HER2 positive cancers with coexisting PIK3CA-activating mutations. PMID:26640141

  2. Phosphatidylinositol 3-kinase signaling determines kidney size

    PubMed Central

    Chen, Jian-Kang; Nagai, Kojiro; Chen, Jianchun; Plieth, David; Hino, Masayo; Xu, Jinxian; Sha, Feng; Ikizler, T. Alp; Quarles, C. Chad; Threadgill, David W.; Neilson, Eric G.; Harris, Raymond C.

    2015-01-01

    Kidney size adaptively increases as mammals grow and in response to the loss of 1 kidney. It is not clear how kidneys size themselves or if the processes that adapt kidney mass to lean body mass also mediate renal hypertrophy following unilateral nephrectomy (UNX). Here, we demonstrated that mice harboring a proximal tubule–specific deletion of Pten (PtenptKO) have greatly enlarged kidneys as the result of persistent activation of the class I PI3K/mTORC2/AKT pathway and an increase of the antiproliferative signals p21Cip1/WAF and p27Kip1. Administration of rapamycin to PtenptKO mice diminished hypertrophy. Proximal tubule–specific deletion of Egfr in PtenptKO mice also attenuated class I PI3K/mTORC2/AKT signaling and reduced the size of enlarged kidneys. In PtenptKO mice, UNX further increased mTORC1 activation and hypertrophy in the remaining kidney; however, mTORC2-dependent AKT phosphorylation did not increase further in the remaining kidney of PtenptKO mice, nor was it induced in the remaining kidney of WT mice. After UNX, renal blood flow and amino acid delivery to the remaining kidney rose abruptly, followed by increased amino acid content and activation of a class III PI3K/mTORC1/S6K1 pathway. Thus, our findings demonstrate context-dependent roles for EGFR-modulated class I PI3K/mTORC2/AKT signaling in the normal adaptation of kidney size and PTEN-independent, nutrient-dependent class III PI3K/mTORC1/S6K1 signaling in the compensatory enlargement of the remaining kidney following UNX. PMID:25985273

  3. Apelin-13 promotes cardiomyocyte hypertrophy via PI3K-Akt-ERK1/2-p70S6K and PI3K-induced autophagy.

    PubMed

    Xie, Feng; Liu, Wei; Feng, Fen; Li, Xin; He, Lu; Lv, Deguan; Qin, Xuping; Li, Lifang; Li, Lanfang; Chen, Linxi

    2015-12-01

    Apelin is highly expressed in rat left ventricular hypertrophy Sprague Dawley rat models, and it plays a crucial role in the cardiovascular system. The aim this study was to clarify whether apelin-13 promotes hypertrophy in H9c2 rat cardiomyocytes and to investigate its underlying mechanism. The cardiomyocyte hypertrophy was observed by measuring the diameter, volume, and protein content of H9c2 cells. The activation of autophagy was evaluated by observing the morphology of autophagosomes by transmission electron microscopy, observing the subcellular localization of LC3 by light microscopy, and detecting the membrane-associated form of LC3 by western blot analysis. The phosphatidylinositol 3-kinase (PI3K) signaling pathway was identified and the proteins expression was detected using western blot analysis. The results revealed that apelin-13 increased the diameter, volume, and protein content of H9c2 cells and promoted the phosphorylation of PI3K, Akt, ERK1/2, and p70S6K. Apelin-13 activated the PI3K-Akt-ERK1/2-p70S6K pathway. PI3K inhibitor LY294002, Akt inhibitor 1701-1, ERK1/2 inhibitor PD98059 attenuated the increase of the cell diameter, volume, protein content induced by apelin-13. Apelin-13 increased the autophagosomes and up-regulated the expressions of beclin 1 and LC3-II/I both transiently and stably. The autophagy inhibitor 3MA ameliorated the increase of cell diameter, volume, and protein content that were induced by apelin-13. These results suggested that apelin-13 promotes H9c2 rat cardiomyocyte hypertrophy via PI3K-Akt-ERK1/2-p70S6K and PI3K-induced autophagy. PMID:26607438

  4. Reversal of the glycolytic phenotype of primary effusion lymphoma cells by combined targeting of cellular metabolism and PI3K/Akt/ mTOR signaling

    PubMed Central

    Bertacchini, Jessika; Frasson, Chiara; Bosco, Raffaella; Accordi, Benedetta; Basso, Giuseppe; Bonora, Massimo; Calabrò, Maria Luisa; Mattiolo, Adriana; Sgarbi, Gianluca; Baracca, Alessandra; Pinton, Paolo; Riva, Giovanni; Rampazzo, Enrico; Petrizza, Luca; Prodi, Luca; Milani, Daniela; Luppi, Mario; Potenza, Leonardo; De Pol, Anto; Cocco, Lucio; Capitani, Silvano; Marmiroli, Sandra

    2016-01-01

    PEL is a B-cell non-Hodgkin lymphoma, occurring predominantly as a lymphomatous effusion in body cavities, characterized by aggressive clinical course, with no standard therapy. Based on previous reports that PEL cells display a Warburg phenotype, we hypothesized that the highly hypoxic environment in which they grow in vivo makes them more reliant on glycolysis, and more vulnerable to drugs targeting this pathway. We established here that indeed PEL cells in hypoxia are more sensitive to glycolysis inhibition. Furthermore, since PI3K/Akt/mTOR has been proposed as a drug target in PEL, we ascertained that pathway-specific inhibitors, namely the dual PI3K and mTOR inhibitor, PF-04691502, and the Akt inhibitor, Akti 1/2, display improved cytotoxicity to PEL cells in hypoxic conditions. Unexpectedly, we found that these drugs reduce lactate production/extracellular acidification rate, and, in combination with the glycolysis inhibitor 2-deoxyglucose (2-DG), they shift PEL cells metabolism from aerobic glycolysis towards oxidative respiration. Moreover, the associations possess strong synergistic cytotoxicity towards PEL cells, and thus may reduce adverse reaction in vivo, while displaying very low toxicity to normal lymphocytes. Finally, we showed that the association of 2-DG and PF-04691502 maintains its cytotoxic and proapoptotic effect also in PEL cells co-cultured with human primary mesothelial cells, a condition known to mimic the in vivo environment and to exert a protective and pro-survival action. All together, these results provide a compelling rationale for the clinical development of new therapies for the treatment of PEL, based on combined targeting of glycolytic metabolism and constitutively activated signaling pathways. PMID:26575168

  5. PRL-3 promotes the peritoneal metastasis of gastric cancer through the PI3K/Akt signaling pathway by regulating PTEN.

    PubMed

    Xiong, Jianbo; Li, Zhengrong; Zhang, Yang; Li, Daojiang; Zhang, Guoyang; Luo, Xianshi; Jie, Zhigang; Liu, Yi; Cao, Yi; Le, Zhibiao; Tan, Shengxing; Zou, Wenyu; Gong, Peitao; Qiu, Lingyu; Li, Yuanyuan; Wang, Huan; Chen, Heping

    2016-10-01

    Peritoneal metastasis is the most frequent cause of death in patients with advanced gastric carcinoma (GC). The phosphatase of regenerating liver-3 (PRL-3) is recognized as an oncogene and plays an important role in GC peritoneal metastasis. However, the mechanism of how PRL-3 regulates GC invasion and metastasis is unknown. In the present study, we found that PRL-3 presented with high expression in GC with peritoneal metastasis, but phosphatase and tensin homologue (PTEN) was weakly expressed. The p-PTEN/PTEN ratio was also higher in GC with peritoneal metastasis than that in the normal gastric tissues. We also found the same phenomenon when comparing the gastric mucosa cell line with the GC cell lines. After constructing a wild-type and a mutant-type plasmid without enzyme activity and transfecting them into GC SGC7901 cells, we showed that only PRL-3 had enzyme activity to downregulate PTEN and cause PTEN phosphorylation. The results also showed that PRL-3 increased the expression levels of MMP-2/MMP-9 and promoted the migration and invasion of the SGC7901 cells. Knockdown of PRL-3 decreased the expression levels of MMP-2/MMP-9 significantly, which further inhibited the migration and invasion of the GC cells. PRL-3 also increased the expression ratio of p-Akt/Akt, which indicated that PRL-3 may mediate the PI3K/Akt pathway to promote GC metastasis. When we transfected the PTEN siRNA plasmid into the PRL-3 stable low expression GC cells, the expression of p-Akt, MMP-2 and MMP-9 was reversed. In conclusion, our results provide a bridge between PRL-3 and PTEN; PRL-3 decreased the expression of PTEN as well as increased the level of PTEN phosphorylation and inactivated it, consequently activating the PI3K/Akt signaling pathway, and upregulating MMP-2/MMP-9 expression to promote GC cell peritoneal metastasis.

  6. PRL-3 promotes the peritoneal metastasis of gastric cancer through the PI3K/Akt signaling pathway by regulating PTEN

    PubMed Central

    Xiong, Jianbo; Li, Zhengrong; Zhang, Yang; Li, Daojiang; Zhang, Guoyang; Luo, Xianshi; Jie, Zhigang; Liu, Yi; Cao, Yi; Le, Zhibiao; Tan, Shengxing; Zou, Wenyu; Gong, Peitao; Qiu, Lingyu; Li, Yuanyuan; Wang, Huan; Chen, Heping

    2016-01-01

    Peritoneal metastasis is the most frequent cause of death in patients with advanced gastric carcinoma (GC). The phosphatase of regenerating liver-3 (PRL-3) is recognized as an oncogene and plays an important role in GC peritoneal metastasis. However, the mechanism of how PRL-3 regulates GC invasion and metastasis is unknown. In the present study, we found that PRL-3 presented with high expression in GC with peritoneal metastasis, but phosphatase and tensin homologue (PTEN) was weakly expressed. The p-PTEN/PTEN ratio was also higher in GC with peritoneal metastasis than that in the normal gastric tissues. We also found the same phenomenon when comparing the gastric mucosa cell line with the GC cell lines. After constructing a wild-type and a mutant-type plasmid without enzyme activity and transfecting them into GC SGC7901 cells, we showed that only PRL-3 had enzyme activity to downregulate PTEN and cause PTEN phosphorylation. The results also showed that PRL-3 increased the expression levels of MMP-2/MMP-9 and promoted the migration and invasion of the SGC7901 cells. Knockdown of PRL-3 decreased the expression levels of MMP-2/MMP-9 significantly, which further inhibited the migration and invasion of the GC cells. PRL-3 also increased the expression ratio of p-Akt/Akt, which indicated that PRL-3 may mediate the PI3K/Akt pathway to promote GC metastasis. When we transfected the PTEN siRNA plasmid into the PRL-3 stable low expression GC cells, the expression of p-Akt, MMP-2 and MMP-9 was reversed. In conclusion, our results provide a bridge between PRL-3 and PTEN; PRL-3 decreased the expression of PTEN as well as increased the level of PTEN phosphorylation and inactivated it, consequently activating the PI3K/Akt signaling pathway, and upregulating MMP-2/MMP-9 expression to promote GC cell peritoneal metastasis. PMID:27572739

  7. 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.

  8. Reversal of the glycolytic phenotype of primary effusion lymphoma cells by combined targeting of cellular metabolism and PI3K/Akt/ mTOR signaling.

    PubMed

    Mediani, Laura; Gibellini, Federica; Bertacchini, Jessika; Frasson, Chiara; Bosco, Raffaella; Accordi, Benedetta; Basso, Giuseppe; Bonora, Massimo; Calabrò, Maria Luisa; Mattiolo, Adriana; Sgarbi, Gianluca; Baracca, Alessandra; Pinton, Paolo; Riva, Giovanni; Rampazzo, Enrico; Petrizza, Luca; Prodi, Luca; Milani, Daniela; Luppi, Mario; Potenza, Leonardo; De Pol, Anto; Cocco, Lucio; Capitani, Silvano; Marmiroli, Sandra

    2016-02-01

    PEL is a B-cell non-Hodgkin lymphoma, occurring predominantly as a lymphomatous effusion in body cavities, characterized by aggressive clinical course, with no standard therapy. Based on previous reports that PEL cells display a Warburg phenotype, we hypothesized that the highly hypoxic environment in which they grow in vivo makes them more reliant on glycolysis, and more vulnerable to drugs targeting this pathway. We established here that indeed PEL cells in hypoxia are more sensitive to glycolysis inhibition. Furthermore, since PI3K/Akt/mTOR has been proposed as a drug target in PEL, we ascertained that pathway-specific inhibitors, namely the dual PI3K and mTOR inhibitor, PF-04691502, and the Akt inhibitor, Akti 1/2, display improved cytotoxicity to PEL cells in hypoxic conditions. Unexpectedly, we found that these drugs reduce lactate production/extracellular acidification rate, and, in combination with the glycolysis inhibitor 2-deoxyglucose (2-DG), they shift PEL cells metabolism from aerobic glycolysis towards oxidative respiration. Moreover, the associations possess strong synergistic cytotoxicity towards PEL cells, and thus may reduce adverse reaction in vivo, while displaying very low toxicity to normal lymphocytes. Finally, we showed that the association of 2-DG and PF-04691502 maintains its cytotoxic and proapoptotic effect also in PEL cells co-cultured with human primary mesothelial cells, a condition known to mimic the in vivo environment and to exert a protective and pro-survival action. All together, these results provide a compelling rationale for the clinical development of new therapies for the treatment of PEL, based on combined targeting of glycolytic metabolism and constitutively activated signaling pathways. PMID:26575168

  9. Insulin-like growth factor-1 (IGF-1) promotes myoblast proliferation and skeletal muscle growth of embryonic chickens via the PI3K/Akt signalling pathway.

    PubMed

    Yu, Minli; Wang, Huan; Xu, Yali; Yu, Debing; Li, Dongfeng; Liu, Xiuhong; Du, Wenxing

    2015-08-01

    During embryonic development, IGF-1 fulfils crucial roles in skeletal myogenesis. However, the involvement of IGF-1-induced myoblast proliferation in muscle growth is still unclear. In the present study, we have characterised the role of IGF-1 in myoblast proliferation both in vitro and in vivo and have revealed novel details of how exogenous IGF-1 influences myogenic genes in chicken embryos. The results show that IGF-1 significantly induces the proliferation of cultured myoblasts in a dose-dependent manner. Additionally, the IGF-1 treatment significantly promoted myoblasts entering a new cell cycle and increasing the mRNA expression levels of cell cycle-dependent genes. However, these effects were inhibited by the PI3K inhibitor LY294002 and the Akt inhibitor KP372-1. These data indicated that the pro-proliferative effect of IGF-1 was mediated in response to the PI3K/Akt signalling pathway. Moreover, we also showed that exogenous IGF-1 stimulated myoblast proliferation in vivo. IGF-1 administration obviously promoted the incorporation of BrdU and remarkably increased the number of PAX7-positive cells in the skeletal muscle of chicken embryos. Administration of IGF-1 also significantly induced the upregulation of myogenic factors gene, the enhancement of c-Myc and the inhibition of myostatin (Mstn) expression. These findings demonstrate that IGF-1 has strong activity as a promoter of myoblast expansion and muscle fiber formation during early myogenesis. Therefore, this study offers insight into the mechanisms responsible for IGF-1-mediated stimulation of embryonic skeletal muscle development, which could have important implications for the improvement of chicken meat production.

  10. PI3K-Akt pathway: its functions and alterations in human cancer.

    PubMed

    Osaki, M; Oshimura, M; Ito, H

    2004-11-01

    Phosphatidylinositol-3-kinase (PI3K) is a lipid kinase and generates phosphatidylinositol-3,4,5-trisphosphate (PI(3, 4, 5)P3). PI(3, 4, 5)P3 is a second messenger essential for the translocation of Akt to the plasma membrane where it is phosphorylated and activated by phosphoinositide-dependent kinase (PDK) 1 and PDK2. Activation of Akt plays a pivotal role in fundamental cellular functions such as cell proliferation and survival by phosphorylating a variety of substrates. In recent years, it has been reported that alterations to the PI3K-Akt signaling pathway are frequent in human cancer. Constitutive activation of the PI3K-Akt pathway occurs due to amplification of the PIK3C gene encoding PI3K or the Akt gene, or as a result of mutations in components of the pathway, for example PTEN (phosphatase and tensin homologue deleted on chromosome 10), which inhibit the activation of Akt. Several small molecules designed to specifically target PI3K-Akt have been developed, and induced cell cycle arrest or apoptosis in human cancer cells in vitro and in vivo . Moreover, the combination of an inhibitor with various cytotoxic agents enhances the anti-tumor efficacy. Therefore, specific inhibition of the activation of Akt may be a valid approach to treating human malignancies and overcoming the resistance of cancer cells to radiation or chemotherapy. PMID:15505410

  11. EMBRYONIC TESTIS CORD FORMATION AND MESONEPHRIC CELL MIGRATION REQUIRES THE PHOSPHOTIDYLINOSITOL 3-KINASE (PI3K) SIGNALING PATHWAY. (R827405)

    EPA Science Inventory

    The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

  12. Runx2 activates PI3K/Akt signaling via mTORC2 regulation in invasive breast cancer cells

    PubMed Central

    2014-01-01

    Introduction The Runt-related transcription factor Runx2 is critical for skeletal development but is also aberrantly expressed in breast cancers, and promotes cell growth and invasion. A de-regulated serine/threonine kinase Akt signaling pathway is implicated in mammary carcinogenesis and cell survival; however, the mechanisms underlying Runx2 role in survival of invasive breast cancer cells are still unclear. Methods The phenotypic analysis of Runx2 function in cell survival was performed by gene silencing and flow cytometric analysis in highly invasive MDA-MB-231 and SUM-159-PT mammary epithelial cell lines. The expression analysis of Runx2 and pAkt (serine 473) proteins in metastatic breast cancer specimens was performed by immunohistochemistry. The mRNA and protein levels of kinases and phosphatases functional in Akt signaling were determined by real-time PCR and Western blotting, while DNA-protein interaction was studied by chromatin immunoprecipitation assays. Results The high Runx2 levels in invasive mammary epithelial cell lines promoted cell survival in Akt phosphorylation (pAkt-serine 473) dependent manner. The analysis of kinases and phosphatases associated with pAkt regulation revealed that Runx2 promotes pAkt levels via mammalian target of rapamycin complex-2 (mTORC2). The recruitment of Runx2 on mTOR promoter coupled with Runx2-dependent expression of mTORC2 component Rictor defined Runx2 function in pAkt-mediated survival of invasive breast cancer cells. Conclusions Our results identified a novel mechanism of Runx2 regulatory crosstalk in Akt signaling that could have important consequences in targeting invasive breast cancer-associated cell survival. PMID:24479521

  13. BMX acts downstream of PI3K to promote colorectal cancer cell survival and pathway inhibition sensitizes to the BH3 mimetic ABT-737.

    PubMed

    Potter, Danielle S; Kelly, Paul; Denneny, Olive; Juvin, Veronique; Stephens, Len R; Dive, Caroline; Morrow, Christopher J

    2014-02-01

    Evasion of apoptosis is a hallmark of cancer, and reversing this process by inhibition of survival signaling pathways is a potential therapeutic strategy. Phosphoinositide 3-kinase (PI3K) signaling can promote cell survival and is upregulated in solid tumor types, including colorectal cancer (CRC), although these effects are context dependent. The role of PI3K in tumorigenesis combined with their amenability to specific inhibition makes them attractive drug targets. However, we observed that inhibition of PI3K in HCT116, DLD-1, and SW620 CRC cells did not induce apoptotic cell death. Moreover, these cells were relatively resistant to the Bcl-2 homology domain 3 (BH3) mimetic ABT-737, which directly targets the Bcl-2 family of apoptosis regulators. To test the hypothesis that PI3K inhibition lowers the apoptotic threshold without causing apoptosis per se, PI3K inhibitors were combined with ABT-737. PI3K inhibition enhanced ABT-737-induced apoptosis by 2.3- to 4.5-fold and reduced expression levels of MCL-1, the resistance biomarker for ABT-737. PI3K inhibition enhanced ABT-737-induced apoptosis a further 1.4- to 2.4-fold in CRC cells with small interfering RNA-depleted MCL-1, indicative of additional sensitizing mechanisms. The observation that ABT-737-induced apoptosis was unaffected by inhibition of PI3K downstream effectors AKT and mTOR, implicated a novel PI3K-dependant pathway. To elucidate this, an RNA interference (RNAi) screen of potential downstream effectors of PI3K signaling was conducted, which demonstrated that knockdown of the TEC kinase BMX sensitized to ABT-737. This suggests that BMX is an antiapoptotic downstream effector of PI3K, independent of AKT. PMID:24709422

  14. Rab31 and APPL2 enhance FcγR-mediated phagocytosis through PI3K/Akt signaling in macrophages.

    PubMed

    Yeo, Jeremy C; Wall, Adam A; Luo, Lin; Stow, Jennifer L

    2015-03-01

    Membrane remodeling in the early stages of phagocytosis enables the engulfment of particles or pathogens and receptor signaling to activate innate immune responses. Members of the Rab GTPase family and their disparate effectors are recruited sequentially to regulate steps throughout phagocytosis. Rab31 (Rab22b) is known for regulating post-Golgi trafficking, and here we show in macrophages that Rab31-GTP is additionally and specifically recruited to early-stage phagosomes. At phagocytic cups, Rab31 is first recruited during the phosphoinositide transition from PI(4,5)P2 to PI(3,4,5)P3, and it persists on PI(3)P-enriched phagosomes. During early phagocytosis, we find that Rab31 recruits the signaling adaptor APPL2. siRNA depletion of either Rab31 or APPL2 reduces FcγR-mediated phagocytosis. Mechanistically, this corresponds with a delay in the transition to PI(3,4,5)P3 and phagocytic cup closure. APPL2 depletion also reduced PI3K/Akt signaling and enhanced p38 signaling from FcγR. We thus conclude that Rab31/APPL2 is required for key roles in phagocytosis and prosurvival responses of macrophages. Of interest, in terms of localization and function, this Rab31/APPL2 complex is distinct from the Rab5/APPL1 complex, which is also involved in phagocytosis and signaling.

  15. 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

  16. Ammonia-induced Na,K-ATPase/ouabain-mediated EGF receptor transactivation, MAPK/ERK and PI3K/AKT signaling and ROS formation cause astrocyte swelling.

    PubMed

    Dai, Hongliang; Song, Dan; Xu, Junnan; Li, Baoman; Hertz, Leif; Peng, Liang

    2013-11-01

    Ammonia toxicity is clinically important and biologically poorly understood. We reported previously that 3mM ammonia chloride (ammonia), a relevant concentration for hepatic encephalopathy studies, increases production of endogenous ouabain and activity of Na,K-ATPase in astrocytes. In addition, ammonia-induced upregulation of gene expression of α2 isoform of Na,K-ATPase in astrocytes could be inhibited by AG1478, an inhibitor of the EGF receptor (EGFR), and by PP1, an inhibitor of Src, but not by GM6001, an inhibitor of metalloproteinase and shedding of growth factor, suggesting the involvement of endogenous ouabain-induced EGF receptor transactivation. In the present cell culture study, we investigated ammonia effects on phosphorylation of EGF receptor and its intracellular signal pathway towards MAPK/ERK1/2 and PI3K/AKT; interaction between EGF receptor, α1, and α2 isoforms of Na,K-ATPase, Src, ERK1/2, AKT and caveolin-1; and relevance of these signal pathways for ammonia-induced cell swelling, leading to brain edema, an often fatal complication of ammonia toxicity. We found that (i) ammonia increases EGF receptor phosphorylation at EGFR(845) and EGFR(1068); (ii) ammonia-induced ERK1/2 and AKT phosphorylation depends on the activity of EGF receptor and Src, but not on metalloproteinase; (iii) AKT phosphorylation occurs upstream of ERK1/2 phosphorylation; (iv) ammonia stimulates association between the α1 Na,K-ATPase isoform, Src, EGF receptor, ERK1/2, AKT and caveolin-1; (v) ammonia-induced ROS production might occur later than EGFR transactivation; (vi) both ammonia induced ERK phosphorylation and ROS production can be abolished by canrenone, an inhibitor of ouabain, and (vii) ammonia-induced cell swelling depends on signaling via the Na,K-ATPase/ouabain/Src/EGF receptor/PI3K-AKT/ERK1/2, but in response to 3mM ammonia it does not appear until after 12h. Based on literature data it is suggested that the delayed appearance of the ammonia-induced swelling at

  17. PfIRR Interacts with HrIGF-I and Activates the MAP-kinase and PI3-kinase Signaling Pathways to Regulate Glycogen Metabolism in Pinctada fucata

    PubMed Central

    Shi, Yu; He, Mao-xian

    2016-01-01

    The insulin-induced mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathways are major intracellular signaling modules and conserved among eukaryotes that are known to regulate diverse cellular processes. However, they have not been investigated in the mollusk species Pinctada fucata. Here, we demonstrate that insulin-related peptide receptor of P. fucata (pfIRR) interacts with human recombinant insulin-like growth factor I (hrIGF-I), and stimulates the MAPK and PI3K signaling pathways in P. fucata oocytes. We also show that inhibition of pfIRR by the inhibitor PQ401 significantly attenuates the basal and hrIGF-I-induced phosphorylation of MAPK and PI3K/Akt at amino acid residues threonine 308 and serine 473. Furthermore, our experiments show that there is cross-talk between the MAPK and PI3K/Akt pathways, in which MAPK kinase positively regulates the PI3K pathway, and PI3K positively regulates the MAPK cascade. Intramuscular injection of hrIGF-I stimulates the PI3K and MAPK pathways to increase the expression of pfirr, protein phosphatase 1, glucokinase, and the phosphorylation of glycogen synthase, decreases the mRNA expression of glycogen synthase kinase-3 beta, decreases glucose levels in hemocytes, and increases glycogen levels in digestive glands. These results suggest that the MAPK and PI3K pathways in P. fucata transmit the hrIGF-I signal to regulate glycogen metabolism. PMID:26911653

  18. PfIRR Interacts with HrIGF-I and Activates the MAP-kinase and PI3-kinase Signaling Pathways to Regulate Glycogen Metabolism in Pinctada fucata.

    PubMed

    Shi, Yu; He, Mao-xian

    2016-01-01

    The insulin-induced mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathways are major intracellular signaling modules and conserved among eukaryotes that are known to regulate diverse cellular processes. However, they have not been investigated in the mollusk species Pinctada fucata. Here, we demonstrate that insulin-related peptide receptor of P. fucata (pfIRR) interacts with human recombinant insulin-like growth factor I (hrIGF-I), and stimulates the MAPK and PI3K signaling pathways in P. fucata oocytes. We also show that inhibition of pfIRR by the inhibitor PQ401 significantly attenuates the basal and hrIGF-I-induced phosphorylation of MAPK and PI3K/Akt at amino acid residues threonine 308 and serine 473. Furthermore, our experiments show that there is cross-talk between the MAPK and PI3K/Akt pathways, in which MAPK kinase positively regulates the PI3K pathway, and PI3K positively regulates the MAPK cascade. Intramuscular injection of hrIGF-I stimulates the PI3K and MAPK pathways to increase the expression of pfirr, protein phosphatase 1, glucokinase, and the phosphorylation of glycogen synthase, decreases the mRNA expression of glycogen synthase kinase-3 beta, decreases glucose levels in hemocytes, and increases glycogen levels in digestive glands. These results suggest that the MAPK and PI3K pathways in P. fucata transmit the hrIGF-I signal to regulate glycogen metabolism.

  19. Xyloketal B suppresses glioblastoma cell proliferation and migration in vitro through inhibiting TRPM7-regulated PI3K/Akt and MEK/ERK signaling pathways.

    PubMed

    Chen, Wen-Liang; Turlova, Ekaterina; Sun, Christopher L F; Kim, Ji-Sun; Huang, Sammen; Zhong, Xiao; Guan, Yong-Yuan; Wang, Guan-Lei; Rutka, James T; Feng, Zhong-Ping; Sun, Hong-Shuo

    2015-04-01

    Glioblastoma, the most common and aggressive type of brain tumors, has devastatingly proliferative and invasive characteristics. The need for finding a novel and specific drug target is urgent as the current approaches have limited therapeutic effects in treating glioblastoma. Xyloketal B is a marine compound obtained from mangrove fungus Xylaria sp. (No. 2508) from the South China Sea, and has displayed antioxidant activity and protective effects on endothelial and neuronal oxidative injuries. In this study, we used a glioblastoma U251 cell line to (1) explore the effects of xyloketal B on cell viability, proliferation, and migration; and (2) investigate the underlying molecular mechanisms and signaling pathways. MTT assay, colony formation, wound healing, western blot, and patch clamp techniques were employed. We found that xyloketal B reduced cell viability, proliferation, and migration of U251 cells. In addition, xyloketal B decreased p-Akt and p-ERK1/2 protein expressions. Furthermore, xyloketal B blocked TRPM7 currents in HEK-293 cells overexpressing TRPM7. These effects were confirmed by using a TRPM7 inhibitor, carvacrol, in a parallel experiment. Our findings indicate that TRPM7-regulated PI3K/Akt and MEK/ERK signaling is involved in anti-proliferation and migration effects of xyloketal B on U251 cells, providing in vitro evidence for the marine compound xyloketal B to be a potential drug for treating glioblastoma. PMID:25913706

  20. Xyloketal B Suppresses Glioblastoma Cell Proliferation and Migration in Vitro through Inhibiting TRPM7-Regulated PI3K/Akt and MEK/ERK Signaling Pathways

    PubMed Central

    Chen, Wen-Liang; Turlova, Ekaterina; Sun, Christopher L. F.; Kim, Ji-Sun; Huang, Sammen; Zhong, Xiao; Guan, Yong-Yuan; Wang, Guan-Lei; Rutka, James T.; Feng, Zhong-Ping; Sun, Hong-Shuo

    2015-01-01

    Glioblastoma, the most common and aggressive type of brain tumors, has devastatingly proliferative and invasive characteristics. The need for finding a novel and specific drug target is urgent as the current approaches have limited therapeutic effects in treating glioblastoma. Xyloketal B is a marine compound obtained from mangrove fungus Xylaria sp. (No. 2508) from the South China Sea, and has displayed antioxidant activity and protective effects on endothelial and neuronal oxidative injuries. In this study, we used a glioblastoma U251 cell line to (1) explore the effects of xyloketal B on cell viability, proliferation, and migration; and (2) investigate the underlying molecular mechanisms and signaling pathways. MTT assay, colony formation, wound healing, western blot, and patch clamp techniques were employed. We found that xyloketal B reduced cell viability, proliferation, and migration of U251 cells. In addition, xyloketal B decreased p-Akt and p-ERK1/2 protein expressions. Furthermore, xyloketal B blocked TRPM7 currents in HEK-293 cells overexpressing TRPM7. These effects were confirmed by using a TRPM7 inhibitor, carvacrol, in a parallel experiment. Our findings indicate that TRPM7-regulated PI3K/Akt and MEK/ERK signaling is involved in anti-proliferation and migration effects of xyloketal B on U251 cells, providing in vitro evidence for the marine compound xyloketal B to be a potential drug for treating glioblastoma. PMID:25913706

  1. Gadolinium promoted proliferation in mouse embryo fibroblast NIH3T3 cells through Rac and PI3K/Akt signaling pathways.

    PubMed

    Shen, Liming; Yang, Aochu; Yao, Pengwei; Sun, Xiaohong; Chen, Cheng; Mo, Cuiping; Shi, Lei; Chen, Youjiao; Liu, Qiong

    2014-08-01

    Nephrogenic systemic fibrosis (NSF) is a fibrosing disorder disease developed in patients with underlying renal insufficiency following exposure to gadolinium-based contrast agents (GBCAs). Previous studies have demonstrated that GdCl3 can promote NIH3T3 fibroblast cell proliferation, which provide a new clue to the role of GBCAs in the development of NSF. In the present study, we further clarify the molecular mechanism of Gd-promoted proliferation. The results showed that intervention with the Rac inhibitor NSC23766 abrogated Gd-promoted proliferation. The levels of active Rac1 significantly increased in Gd-treated cells detected by pull-down assays. In addition, the phosphorylation of Akt was significantly elevated in the treatment group, which was blocked by NSC23766. NSC23766 also reduced the migration of NIH3T3 cells enhanced by Gd. Moreover, the F-actin cytoskeleton was strengthened and the mitotic cell numbers was significantly increased after exposure to Gd. These results suggest that Rac and PI3K/Akt signaling pathways, as well as integrin-mediated signal pathway may play important roles in Gd-induced cell proliferation. In addition, under serum-free condition, Gd could decrease ROS accumulation and increase NIH3T3 cell survival.

  2. Xyloketal B suppresses glioblastoma cell proliferation and migration in vitro through inhibiting TRPM7-regulated PI3K/Akt and MEK/ERK signaling pathways.

    PubMed

    Chen, Wen-Liang; Turlova, Ekaterina; Sun, Christopher L F; Kim, Ji-Sun; Huang, Sammen; Zhong, Xiao; Guan, Yong-Yuan; Wang, Guan-Lei; Rutka, James T; Feng, Zhong-Ping; Sun, Hong-Shuo

    2015-04-01

    Glioblastoma, the most common and aggressive type of brain tumors, has devastatingly proliferative and invasive characteristics. The need for finding a novel and specific drug target is urgent as the current approaches have limited therapeutic effects in treating glioblastoma. Xyloketal B is a marine compound obtained from mangrove fungus Xylaria sp. (No. 2508) from the South China Sea, and has displayed antioxidant activity and protective effects on endothelial and neuronal oxidative injuries. In this study, we used a glioblastoma U251 cell line to (1) explore the effects of xyloketal B on cell viability, proliferation, and migration; and (2) investigate the underlying molecular mechanisms and signaling pathways. MTT assay, colony formation, wound healing, western blot, and patch clamp techniques were employed. We found that xyloketal B reduced cell viability, proliferation, and migration of U251 cells. In addition, xyloketal B decreased p-Akt and p-ERK1/2 protein expressions. Furthermore, xyloketal B blocked TRPM7 currents in HEK-293 cells overexpressing TRPM7. These effects were confirmed by using a TRPM7 inhibitor, carvacrol, in a parallel experiment. Our findings indicate that TRPM7-regulated PI3K/Akt and MEK/ERK signaling is involved in anti-proliferation and migration effects of xyloketal B on U251 cells, providing in vitro evidence for the marine compound xyloketal B to be a potential drug for treating glioblastoma.

  3. PI-3K Inhibitors Preferentially Target CD15+ Cancer Stem Cell Population in SHH Driven Medulloblastoma

    PubMed Central

    Singh, Alok R.; Joshi, Shweta; Zulcic, Muamera; Alcaraz, Michael; Garlich, Joseph R.; Morales, Guillermo A.; Cho, Yoon J.; Bao, Lei; Levy, Michael L.; Newbury, Robert; Malicki, Denise; Messer, Karen; Crawford, John; Durden, Donald L.

    2016-01-01

    Sonic hedgehog (SHH) medulloblastoma (MB) subtype is driven by a proliferative CD15+ tumor propagating cell (TPC), also considered in the literature as a putative cancer stem cell (CSC). Despite considerable research, much of the biology of this TPC remains unknown. We report evidence that phosphatase and tensin homolog (PTEN) and phosphoinositide 3-kinase (PI-3K) play a crucial role in the propagation, survival and potential response to therapy in this CD15+ CSC/TPC-driven malignant disease. Using the ND2-SmoA1 transgenic mouse model for MB, mouse genetics and patient-derived xenografts (PDXs), we demonstrate that the CD15+TPCs are 1) obligately required for SmoA1Tg-driven tumorigenicity 2) regulated by PTEN and PI-3K signaling 3) selectively sensitive to the cytotoxic effects of pan PI-3K inhibitors in vitro and in vivo but resistant to chemotherapy 4) in the SmoA1Tg mouse model are genomically similar to the SHH human MB subgroup. The results provide the first evidence that PTEN plays a role in MB TPC signaling and biology and that PI-3K inhibitors target and suppress the survival and proliferation of cells within the mouse and human CD15+ cancer stem cell compartment. In contrast, CD15+ TPCs are resistant to cisplatinum, temozolomide and the SHH inhibitor, NVP-LDE-225, agents currently used in treatment of medulloblastoma. These studies validate the therapeutic efficacy of pan PI-3K inhibitors in the treatment of CD15+ TPC dependent medulloblastoma and suggest a sequential combination of PI-3K inhibitors and chemotherapy will have augmented efficacy in the treatment of this disease. PMID:26938241

  4. PI-3K Inhibitors Preferentially Target CD15+ Cancer Stem Cell Population in SHH Driven Medulloblastoma.

    PubMed

    Singh, Alok R; Joshi, Shweta; Zulcic, Muamera; Alcaraz, Michael; Garlich, Joseph R; Morales, Guillermo A; Cho, Yoon J; Bao, Lei; Levy, Michael L; Newbury, Robert; Malicki, Denise; Messer, Karen; Crawford, John; Durden, Donald L

    2016-01-01

    Sonic hedgehog (SHH) medulloblastoma (MB) subtype is driven by a proliferative CD15+ tumor propagating cell (TPC), also considered in the literature as a putative cancer stem cell (CSC). Despite considerable research, much of the biology of this TPC remains unknown. We report evidence that phosphatase and tensin homolog (PTEN) and phosphoinositide 3-kinase (PI-3K) play a crucial role in the propagation, survival and potential response to therapy in this CD15+ CSC/TPC-driven malignant disease. Using the ND2-SmoA1 transgenic mouse model for MB, mouse genetics and patient-derived xenografts (PDXs), we demonstrate that the CD15+TPCs are 1) obligately required for SmoA1Tg-driven tumorigenicity 2) regulated by PTEN and PI-3K signaling 3) selectively sensitive to the cytotoxic effects of pan PI-3K inhibitors in vitro and in vivo but resistant to chemotherapy 4) in the SmoA1Tg mouse model are genomically similar to the SHH human MB subgroup. The results provide the first evidence that PTEN plays a role in MB TPC signaling and biology and that PI-3K inhibitors target and suppress the survival and proliferation of cells within the mouse and human CD15+ cancer stem cell compartment. In contrast, CD15+ TPCs are resistant to cisplatinum, temozolomide and the SHH inhibitor, NVP-LDE-225, agents currently used in treatment of medulloblastoma. These studies validate the therapeutic efficacy of pan PI-3K inhibitors in the treatment of CD15+ TPC dependent medulloblastoma and suggest a sequential combination of PI-3K inhibitors and chemotherapy will have augmented efficacy in the treatment of this disease. PMID:26938241

  5. miR-564 acts as a dual inhibitor of PI3K and MAPK signaling networks and inhibits proliferation and invasion in breast cancer

    NASA Astrophysics Data System (ADS)

    Mutlu, Merve; Saatci, Özge; Ansari, Suhail A.; Yurdusev, Emre; Shehwana, Huma; Konu, Özlen; Raza, Umar; Şahin, Özgür

    2016-09-01

    Dysregulation of PI3K and MAPK pathways promotes uncontrolled cell proliferation, apoptotic inhibition and metastasis. Individual targeting of these pathways using kinase inhibitors has largely been insufficient due to the existence of cross-talks between these parallel cascades. MicroRNAs are small non-coding RNAs targeting several genes simultaneously and controlling cancer-related processes. To identify miRNAs repressing both PI3K and MAPK pathways in breast cancer, we re-analyzed our previous miRNA mimic screen data with reverse phase protein array (RPPA) output, and identified miR-564 inhibiting both PI3K and MAPK pathways causing markedly decreased cell proliferation through G1 arrest. Moreover, ectopic expression of miR-564 blocks epithelial-mesenchymal transition (EMT) and reduces migration and invasion of aggressive breast cancer cells. Mechanistically, miR-564 directly targets a network of genes comprising AKT2, GNA12, GYS1 and SRF, thereby facilitating simultaneous repression of PI3K and MAPK pathways. Notably, combinatorial knockdown of these target genes using a cocktail of siRNAs mimics the phenotypes exerted upon miR-564 expression. Importantly, high miR-564 expression or low expression of target genes in combination is significantly correlated with better distant relapse-free survival of patients. Overall, miR-564 is a potential dual inhibitor of PI3K and MAPK pathways, and may be an attractive target and prognostic marker for breast cancer.

  6. miR-564 acts as a dual inhibitor of PI3K and MAPK signaling networks and inhibits proliferation and invasion in breast cancer.

    PubMed

    Mutlu, Merve; Saatci, Özge; Ansari, Suhail A; Yurdusev, Emre; Shehwana, Huma; Konu, Özlen; Raza, Umar; Şahin, Özgür

    2016-01-01

    Dysregulation of PI3K and MAPK pathways promotes uncontrolled cell proliferation, apoptotic inhibition and metastasis. Individual targeting of these pathways using kinase inhibitors has largely been insufficient due to the existence of cross-talks between these parallel cascades. MicroRNAs are small non-coding RNAs targeting several genes simultaneously and controlling cancer-related processes. To identify miRNAs repressing both PI3K and MAPK pathways in breast cancer, we re-analyzed our previous miRNA mimic screen data with reverse phase protein array (RPPA) output, and identified miR-564 inhibiting both PI3K and MAPK pathways causing markedly decreased cell proliferation through G1 arrest. Moreover, ectopic expression of miR-564 blocks epithelial-mesenchymal transition (EMT) and reduces migration and invasion of aggressive breast cancer cells. Mechanistically, miR-564 directly targets a network of genes comprising AKT2, GNA12, GYS1 and SRF, thereby facilitating simultaneous repression of PI3K and MAPK pathways. Notably, combinatorial knockdown of these target genes using a cocktail of siRNAs mimics the phenotypes exerted upon miR-564 expression. Importantly, high miR-564 expression or low expression of target genes in combination is significantly correlated with better distant relapse-free survival of patients. Overall, miR-564 is a potential dual inhibitor of PI3K and MAPK pathways, and may be an attractive target and prognostic marker for breast cancer. PMID:27600857

  7. miR-564 acts as a dual inhibitor of PI3K and MAPK signaling networks and inhibits proliferation and invasion in breast cancer

    PubMed Central

    Mutlu, Merve; Saatci, Özge; Ansari, Suhail A.; Yurdusev, Emre; Shehwana, Huma; Konu, Özlen; Raza, Umar; Şahin, Özgür

    2016-01-01

    Dysregulation of PI3K and MAPK pathways promotes uncontrolled cell proliferation, apoptotic inhibition and metastasis. Individual targeting of these pathways using kinase inhibitors has largely been insufficient due to the existence of cross-talks between these parallel cascades. MicroRNAs are small non-coding RNAs targeting several genes simultaneously and controlling cancer-related processes. To identify miRNAs repressing both PI3K and MAPK pathways in breast cancer, we re-analyzed our previous miRNA mimic screen data with reverse phase protein array (RPPA) output, and identified miR-564 inhibiting both PI3K and MAPK pathways causing markedly decreased cell proliferation through G1 arrest. Moreover, ectopic expression of miR-564 blocks epithelial-mesenchymal transition (EMT) and reduces migration and invasion of aggressive breast cancer cells. Mechanistically, miR-564 directly targets a network of genes comprising AKT2, GNA12, GYS1 and SRF, thereby facilitating simultaneous repression of PI3K and MAPK pathways. Notably, combinatorial knockdown of these target genes using a cocktail of siRNAs mimics the phenotypes exerted upon miR-564 expression. Importantly, high miR-564 expression or low expression of target genes in combination is significantly correlated with better distant relapse-free survival of patients. Overall, miR-564 is a potential dual inhibitor of PI3K and MAPK pathways, and may be an attractive target and prognostic marker for breast cancer. PMID:27600857

  8. PI3K signalling in GnRH actions on dispersed goldfish pituitary cells: relationship with PKC-mediated LH and GH release and regulation of long-term effects on secretion and total cellular hormone availability.

    PubMed

    Pemberton, Joshua G; Orr, Michael E; Stafford, James L; Chang, John P

    2014-09-01

    Goldfish pituitary cells are exposed to two GnRHs, salmon (s)GnRH and chicken (c)GnRH-II. Phosphoinositide 3-kinase (PI3K) and protein kinase C (PKC) both participate in acute sGnRH- and cGnRH-II-stimulated LH and GH release. Using goldfish pituitary cells, we examined the relationship between PI3K and PKC in acute LH and GH secretion, and PI3K involvement in chronic hormone release and total LH and GH availability. The PI3K inhibitor LY294002 did not affect PKC agonists-induced LH or GH release, and PKC agonists did not alter PI3K p85 phosphorylation, suggesting PKC activation is not upstream of PI3K in acute hormone release. In 2, 6, 12 and 24h treatments, LY294002 did not affect LH release but stimulated total LH availability at 6h. sGnRH stimulatory actions on LH release and total availability at 12 and 24h, and cGnRH-II effects on these parameters at 6h were inhibited by LY294002. LY294002 enhanced basal GH release at 2 and 6h, but reduced total GH at 12 and 24h. Increased GH release was seen following 6, 12 and 24h of sGnRH, and 2, 6 and 24h of cGnRH-II treatment but total GH availability was only elevated by 24h cGnRH-II treatment. Whereas LY294002 inhibited GH release responses to sGnRH at 12h and cGnRH-II at 6h, it attenuated cGnRH-II-elicited, but not sGnRH-induced, effects on total GH. These results indicate that PI3K differentially modulates long-term basal and GnRH-stimulated hormone release, and total hormone availability, in a time-, cell-type-, and GnRH isoform-selective manner.

  9. Ganoderma tsugae Extract Inhibits Growth of HER2-Overexpressing Cancer Cells via Modulation of HER2/PI3K/Akt Signaling Pathway

    PubMed Central

    Kuo, Han-Peng; Hsu, Shih-Chung; Li, Jhy-Wei; Tseng, Hsiu-Hsueh; Chuang, Tzu-Chao; Liu, Jah-Yao; Chen, Shih-Jung; Su, Muh-Hwan; Cheng, Yung-Chi; Chou, Wei-Yuan; Kao, Ming-Ching

    2013-01-01

    Ganoderma, also known as Lingzhi or Reishi, has been used for medicinal purposes in Asian countries for centuries. It is a medicinal fungus with a variety of biological properties including immunomodulatory and antitumor activities. In this study, we investigated the molecular mechanisms by which Ganoderma tsugae (GT), one of the most common species of Ganoderma, inhibits the proliferation of HER2-overexpressing cancer cells. Here, we show that a quality assured extract of GT (GTE) inhibited the growth of HER2-overexpressing cancer cells in vitro and in vivo and enhanced the growth-inhibitory effect of antitumor drugs (e.g., taxol and cisplatin) in these cells. We also demonstrate that GTE induced cell cycle arrest by interfering with the HER2/PI3K/Akt signaling pathway. Furthermore, GTE curtailed the expression of the HER2 protein by modulating the transcriptional activity of the HER2 gene and the stability/degradation of the HER2 protein. In conclusion, this study suggests that GTE may be a useful adjuvant therapeutic agent in the treatment of cancer cells that highly express HER2. PMID:23662119

  10. 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

  11. An Integrative Analysis of the InR/PI3K/Akt Network Identifies the Dynamic Response to Insulin Signaling.

    PubMed

    Vinayagam, Arunachalam; Kulkarni, Meghana M; Sopko, Richelle; Sun, Xiaoyun; Hu, Yanhui; Nand, Ankita; Villalta, Christians; Moghimi, Ahmadali; Yang, Xuemei; Mohr, Stephanie E; Hong, Pengyu; Asara, John M; Perrimon, Norbert

    2016-09-13

    Insulin regulates an essential conserved signaling pathway affecting growth, proliferation, and metabolism. To expand our understanding of the insulin pathway, we combine biochemical, genetic, and computational approaches to build a comprehensive Drosophila InR/PI3K/Akt network. First, we map the dynamic protein-protein interaction network surrounding the insulin core pathway using bait-prey interactions connecting 566 proteins. Combining RNAi screening and phospho-specific antibodies, we find that 47% of interacting proteins affect pathway activity, and, using quantitative phosphoproteomics, we demonstrate that ∼10% of interacting proteins are regulated by insulin stimulation at the level of phosphorylation. Next, we integrate these orthogonal datasets to characterize the structure and dynamics of the insulin network at the level of protein complexes and validate our method by identifying regulatory roles for the Protein Phosphatase 2A (PP2A) and Reptin-Pontin chromatin-remodeling complexes as negative and positive regulators of ribosome biogenesis, respectively. Altogether, our study represents a comprehensive resource for the study of the evolutionary conserved insulin network. PMID:27626673

  12. The Apoptotic Effect of Ursolic Acid on SK-Hep-1 Cells is Regulated by the PI3K/Akt, p38 and JNK MAPK Signaling Pathways.

    PubMed

    Chuang, Wan-Ling; Lin, Ping-Yi; Lin, Hui-Chuan; Chen, Yao-Li

    2016-01-01

    Ursolic acid (UA) is a pentacyclic triterpene acid that is present in a wide variety of medicinal herbs and edible plants. This study investigated the effect of UA on apoptosis and proliferation of hepatocellular carcinoma SK-Hep-1 cells. After treatment of SK-Hep-1 cells with different concentrations of UA, we observed that cell viability was reduced in a dose- and time-dependent manner. Furthermore, there was a dose-dependent increase in the percentage of cells in the sub-G1 and G2/M phases, with cells treated with 60 μM showing the highest percentages of cells in those phases. UA-induced chromatin condensation of nuclei was observed by using DAPI staining. The western blot results revealed that exposure to UA was associated with decreased expression of the anti-apoptotic proteins Mcl-1, Bcl-xL, Bcl-2, and TCTP and increased expression of apoptosis-related proteins TNF-α, Fas, FADD, Bax, cleaved caspase-3, caspase-8, caspase-9, and PARP. Immunocytochemistry staining showed that treatment with UA resulted in increased expression of caspase-3. Moreover, exposure to UA resulted in the inhibition of the PI3K/Akt and p38 MAPK signaling pathways. These findings suggest that UA inhibits the proliferation of SK-Hep-1 cells and induces apoptosis.

  13. Naoxintong Protects Primary Neurons from Oxygen-Glucose Deprivation/Reoxygenation Induced Injury through PI3K-Akt Signaling Pathway

    PubMed Central

    Zhao, Pei; Zhu, Jinqiang; Yan, Chen; Li, Lin; Zhang, Han; Zhang, Meng; Gao, Xiumei

    2016-01-01

    Naoxintong capsule (NXT), developed from Buyang Huanwu Decoction, has shown the neuroprotective effects in cerebrovascular diseases, but the neuroprotection mechanisms of NXT on ischemia/reperfusion injured neurons have not yet been well known. In this study, we established the oxygen-glucose deprivation/reoxygenation (OGD/R) induced neurons injury model and treat the neurons with cerebrospinal fluid containing NXT (BNC) to investigate the effects of NXT on OGD/R induced neurons injury and potential mechanisms. BNC improved neuron viability and decreased apoptotic rate induced by OGD/R. BNC attenuated OGD/R induced cytosolic and mitochondrial Ca2+ overload, ROS generation, intracellular NO levels and nNOS mRNA increase, and cytochrome-c release when compared with OGD/R group. BNC significantly inhibited both mPTP opening and ΔΨm depolarization. BNC increased Bcl-2 expression and decreased Bax expression, upregulated the Bcl-2/Bax ratio, downregulated caspase-3 mRNA and caspase-9 mRNA expression, and decreased cleaved caspase-3 expression and caspase-3 activity. BNC increased phosphorylation of Akt following OGD/R, while LY294002 attenuated BNC induced increase of phosphorylated Akt expression. Our study demonstrated that NXT protected primary neurons from OGD/R induced injury by inhibiting calcium overload and ROS generation, protecting mitochondria, and inhibiting mitochondrial apoptotic pathway which was mediated partially by PI3K-Akt signaling pathway activation. PMID:26949405

  14. NVP-BKM120, a novel PI3K inhibitor, shows synergism with a STAT3 inhibitor in human gastric cancer cells harboring KRAS mutations

    PubMed Central

    PARK, EUNJU; PARK, JINAH; HAN, SAE-WON; IM, SEOCK-AH; KIM, TAE-YOU; OH, DO-YOUN; BANG, YUNG-JUE

    2012-01-01

    Aberrations of Phosphoinositide 3-kinase (PI3K)/AKT signaling are frequently observed in many types of cancer, promoting its emergence as a promising target for cancer treatment. PI3K can become activated by various pathways, one of which includes RAS. RAS can not only directly activate the PI3K/AKT pathway via binding to p110 of PI3K, but also regulates mTOR via ERK or RSK independently of the PI3K/AKT pathway. Thus, actively mutated RAS can constitutively activate PI3K signaling. Additionally, in RAS tumorigenic transformation, signal transducer and activator of transcription 3 (STAT3) has been known also to be required. In this study, we examined the efficacy of NVP-BKM120, a pan-class I PI3K inhibitor in human gastric cancer cells and hypothesized that the combined inhibition of PI3K and STAT3 would be synergistic in KRAS mutant gastric cancer cells. NVP-BKM120 demonstrated anti-proliferative activity in 11 human gastric cancer cell lines by decreasing mTOR downstream signaling. But NVP-BKM120 treatment increased p-AKT by subsequent abrogation of feedback inhibition by stabilizing insulin receptor substrate-1. In KRAS mutant gastric cancer cells, either p-ERK or p-STAT3 was also increased upon treatment of NVP-BKM120. The synergistic efficacy study demonstrated that dual PI3K and STAT3 blockade showed a synergism in cells harboring mutated KRAS by inducing apoptosis. The synergistic effect was not seen in KRAS wild-type cells. Together, these findings suggest for the first time that the dual inhibition of PI3K and STAT3 signaling may be an effective therapeutic strategy for KRAS mutant gastric cancer patients. PMID:22159814

  15. Polyphenols Isolated from Allium cepa L. Induces Apoptosis by Induction of p53 and Suppression of Bcl-2 through Inhibiting PI3K/Akt Signaling Pathway in AGS Human Cancer Cells

    PubMed Central

    Lee, Won Sup; Yi, Sang Mi; Yun, Jeong Won; Jung, Ji Hyun; Kim, Dong Hoon; Kim, Hye Jung; Chang, Seong-Hwan; Kim, GonSup; Ryu, Chung Ho; Shin, Sung Chul; Hong, Soon Chan; Choi, Yung Hyun; Jung, Jin-Myung

    2014-01-01

    Background: The extract of Allium cepa Linn is commonly used as adjuvant food for cancer therapy. We assumed that it includes a potential source of anti-cancer properties. Methods: We investigated anti-cancer effects of polyphenols extracted from lyophilized A. cepa Linn (PEAL) in AGS human cancer cells. Results: PEAL inhibited cell growth in a dose-dependent manner. It was related to caspase-dependent apoptosis. We confirmed this finding with annexin V staining. PEAL up-regulated p53 expression, and subsequent Bax induction, down regulated Bcl-2 protein, anti-apoptotic protein. In addition, PEAL suppressed Akt activity and PEAL-induced apoptosis were significantly accentuated with Akt inhibitor (LY294002). Conclusions: Our data suggested that PEAL induce caspase-dependent apoptosis through mitochondrial pathway by up-regulating p53 protein, and subsequent Bax protein as well as by modulating Bcl-2 protein, and that PEAL induces caspase-dependent apoptosis at least in part through the inhibition of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. This study provides evidence that PEAL might be useful for the treatment of cancer. PMID:25337568

  16. Epigallocatechin gallate-induced modulation of FoxO signaling in mammalian cells and C. elegans: FoxO stimulation is masked via PI3K/Akt activation by hydrogen peroxide formed in cell culture.

    PubMed

    Bartholome, André; Kampkötter, Andreas; Tanner, Stephan; Sies, Helmut; Klotz, Lars-Oliver

    2010-09-01

    The green tea flavonoid epigallocatechin gallate (EGCG) is demonstrated in this study to modulate FoxO transcription factors in human skin fibroblasts in culture. EGCG at 1 microM stimulated FoxO transcription factor nuclear accumulation and DNA binding activity. This effect was masked at higher EGCG concentrations (100 microM) by EGCG-derived hydrogen peroxide generated in cell culture media that stimulates phosphoinositide-3'-kinase (PI3K)/Akt signaling to attenuate FoxO activity, involving FoxO phosphorylation, nuclear exclusion and attenuation of DNA binding activity. Like low concentrations of EGCG, harmine, an inhibitor of the FoxO kinase DYRK1a, stimulated FoxO nuclear accumulation and DNA binding activity. Exposure of Caenorhabditis elegans worms to EGCG caused nuclear accumulation of the FoxO ortholog, DAF-16, and enhanced expression of the DAF-16 target gene, sod-3. In line with the role of FoxO/DAF-16 in the control of life span, C. elegans mean and maximum life span were enhanced by 20% and 13%, respectively, by EGCG. PMID:20513639

  17. Distinct roles of class IA PI3K isoforms in primary and immortalised macrophages.

    PubMed

    Papakonstanti, Evangelia A; Zwaenepoel, Olivier; Bilancio, Antonio; Burns, Emily; Nock, Gemma E; Houseman, Benjamin; Shokat, Kevan; Ridley, Anne J; Vanhaesebroeck, Bart

    2008-12-15

    The class IA isoforms of phosphoinositide 3-kinase (p110alpha, p110beta and p110delta) often have non-redundant functions in a given cell type. However, for reasons that are unclear, the role of a specific PI3K isoform can vary between cell types. Here, we compare the relative contributions of PI3K isoforms in primary and immortalised macrophages. In primary macrophages stimulated with the tyrosine kinase ligand colony-stimulating factor 1 (CSF1), all class IA PI3K isoforms participate in the regulation of Rac1, whereas p110delta selectively controls the activities of Akt, RhoA and PTEN, in addition to controlling proliferation and chemotaxis. The prominent role of p110delta in these cells correlates with it being the main PI3K isoform that is recruited to the activated CSF1 receptor (CSF1R). In immortalised BAC1.2F5 macrophages, however, the CSF1R also engages p110alpha, which takes up a more prominent role in CSF1R signalling, in processes including Akt phosphorylation and regulation of DNA synthesis. Cell migration, however, remains dependent mainly on p110delta. In other immortalised macrophage cell lines, such as IC-21 and J774.2, p110alpha also becomes more prominently involved in CSF1-induced Akt phosphorylation, at the expense of p110delta.These data show that PI3K isoforms can be differentially regulated in distinct cellular contexts, with the dominant role of the p110delta isoform in Akt phosphorylation and proliferation being lost upon cell immortalisation. These findings suggest that p110delta-selective PI3K inhibitors may be more effective in inflammation than in cancer. PMID:19033389

  18. Effects of PI3K inhibition and low docosahexaenoic acid on cognition and behavior.

    PubMed

    Bandaru, Sathyajit S; Lin, Kristen; Roming, Stephanie L; Vellipuram, Ramana; Harney, Jacob P

    2010-06-01

    Alterations in two components of the brain's insulin signaling pathway, docosahexaenoic acid (DHA) content and phosphoinositide 3-kinase (PI3K) activity, have been implicated in the insulin resistance that is central to type II diabetes mellitus (DM). A 2- to 3-fold increased risk of developing Alzheimer's disease (AD) in patients with type II DM suggests a potential link between cognition and insulin action. The current study was designed to examine the impact of DHA dietary content and PI3K activity on learning, memory, depression, and anxiety in rodents. Mice were divided into the following groups: (1) control diet and vehicle injection (control PI3K), (2) control diet and wortmannin injection (PI3K inhibition), (3) low DHA diet and vehicle, and (4) low DHA diet and wortmannin. Each group was assessed for effects on activity, cognition, depression, and anxiety. Concentrations of glucose and insulin in plasma were quantified to confirm insulin resistance. Results showed significant increases in depression, anxiety, plasma insulin and glucose, and significant decreases in activity in wortmannin-treated mice regardless of diet. The control diet/wortmannin-treated group showed a significant decrease in memory compared to all other groups. The low DHA diet/wortmannin-treated group had slightly improved memory and lower levels of depression compared to the control diet/wortmannin-treated group. Results of the present study suggest that inhibition of PI3K decreases activity and memory while increasing insulin resistance, depression, and anxiety. In addition, these results suggest a possible compensatory role of low DHA in decreasing the effects of dysfunctional PI3K in AD associated cognitive decline and depression. PMID:19914265

  19. Alisertib induces cell cycle arrest and autophagy and suppresses epithelial-to-mesenchymal transition involving PI3K/Akt/mTOR and sirtuin 1-mediated signaling pathways in human pancreatic cancer cells

    PubMed Central

    Wang, Feng; Li, Hai; Yan, Xiao-Gang; Zhou, Zhi-Wei; Yi, Zhi-Gang; He, Zhi-Xu; Pan, Shu-Ting; Yang, Yin-Xue; Wang, Zuo-Zheng; Zhang, Xueji; Yang, Tianxing; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    Pancreatic cancer is the most aggressive cancer worldwide with poor response to current therapeutics. Alisertib (ALS), a potent and selective Aurora kinase A inhibitor, exhibits potent anticancer effects in preclinical and clinical studies; however, the effect and underlying mechanism of ALS in the pancreatic cancer treatment remain elusive. This study aimed to examine the effects of ALS on cell growth, autophagy, and epithelial-to-mesenchymal transition (EMT) and to delineate the possible molecular mechanisms in human pancreatic cancer PANC-1 and BxPC-3 cells. The results showed that ALS exerted potent cell growth inhibitory, pro-autophagic, and EMT-suppressing effects in PANC-1 and BxPC-3 cells. ALS remarkably arrested PANC-1 and BxPC-3 cells in G2/M phase via regulating the expression of cyclin-dependent kinases 1 and 2, cyclin B1, cyclin D1, p21 Waf1/Cip1, p27 Kip1, and p53. ALS concentration-dependently induced autophagy in PANC-1 and BxPC-3 cells, which may be attributed to the inhibition of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR), p38 mitogen-activated protein kinase (p38 MAPK), and extracellular signal-regulated kinases 1 and 2 (Erk1/2) but activation of 5′-AMP-dependent kinase signaling pathways. ALS significantly inhibited EMT in PANC-1 and BxPC-3 cells with an increase in the expression of E-cadherin and a decrease in N-cadherin. In addition, ALS suppressed the expression of sirtuin 1 (Sirt1) and pre-B cell colony-enhancing factor/visfatin in both cell lines with a rise in the level of acetylated p53. These findings show that ALS induces cell cycle arrest and promotes autophagic cell death but inhibits EMT in pancreatic cancer cells with the involvement of PI3K/Akt/mTOR, p38 MAPK, Erk1/2, and Sirt1-mediated signaling pathways. Taken together, ALS may represent a promising anticancer drug for pancreatic cancer treatment. More studies are warranted to investigate other molecular targets and

  20. Alisertib induces cell cycle arrest and autophagy and suppresses epithelial-to-mesenchymal transition involving PI3K/Akt/mTOR and sirtuin 1-mediated signaling pathways in human pancreatic cancer cells.

    PubMed

    Wang, Feng; Li, Hai; Yan, Xiao-Gang; Zhou, Zhi-Wei; Yi, Zhi-Gang; He, Zhi-Xu; Pan, Shu-Ting; Yang, Yin-Xue; Wang, Zuo-Zheng; Zhang, Xueji; Yang, Tianxing; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    Pancreatic cancer is the most aggressive cancer worldwide with poor response to current therapeutics. Alisertib (ALS), a potent and selective Aurora kinase A inhibitor, exhibits potent anticancer effects in preclinical and clinical studies; however, the effect and underlying mechanism of ALS in the pancreatic cancer treatment remain elusive. This study aimed to examine the effects of ALS on cell growth, autophagy, and epithelial-to-mesenchymal transition (EMT) and to delineate the possible molecular mechanisms in human pancreatic cancer PANC-1 and BxPC-3 cells. The results showed that ALS exerted potent cell growth inhibitory, pro-autophagic, and EMT-suppressing effects in PANC-1 and BxPC-3 cells. ALS remarkably arrested PANC-1 and BxPC-3 cells in G2/M phase via regulating the expression of cyclin-dependent kinases 1 and 2, cyclin B1, cyclin D1, p21 Waf1/Cip1, p27 Kip1, and p53. ALS concentration-dependently induced autophagy in PANC-1 and BxPC-3 cells, which may be attributed to the inhibition of phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt)/mammalian target of rapamycin (mTOR), p38 mitogen-activated protein kinase (p38 MAPK), and extracellular signal-regulated kinases 1 and 2 (Erk1/2) but activation of 5'-AMP-dependent kinase signaling pathways. ALS significantly inhibited EMT in PANC-1 and BxPC-3 cells with an increase in the expression of E-cadherin and a decrease in N-cadherin. In addition, ALS suppressed the expression of sirtuin 1 (Sirt1) and pre-B cell colony-enhancing factor/visfatin in both cell lines with a rise in the level of acetylated p53. These findings show that ALS induces cell cycle arrest and promotes autophagic cell death but inhibits EMT in pancreatic cancer cells with the involvement of PI3K/Akt/mTOR, p38 MAPK, Erk1/2, and Sirt1-mediated signaling pathways. Taken together, ALS may represent a promising anticancer drug for pancreatic cancer treatment. More studies are warranted to investigate other molecular targets and

  1. 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

    In children, neuroblastomas are the most common and deadly solid tumor. Our previous study showed that honokiol, a small-molecule polyphenol, can traverse the blood-brain barrier and kill neuroblastoma cells. In this study, we further investigated the mechanisms of honokiol-induced insults to neuroblastoma cells. Treatment of neuroblastoma neuro-2a cells with honokiol elevated the levels of microtubule-associated protein light chain 3 (LC3)-II and induced cell autophagy in time- and concentration-dependent manners. Interestingly, pretreatment with 3-methyladenine (3-MA), an inhibitor of autophagy, led to the simultaneous attenuation of honokiol-induced cell autophagy and apoptosis but did not influence cell necrosis. As to the mechanisms, exposure of neuro-2a cells to honokiol time-dependently decreased the amount of phosphatidylinositol 3-kinase (PI3K). Sequentially, honokiol downregulated phosphorylation of protein kinase B (Akt) and mammalian target of rapamycin (mTOR) in neuro-2a cells. Furthermore, honokiol elevated the levels of glucose-regulated protein (GpR)78, an endoplasmic reticular stress (ERS)-associated protein, and amounts of intracellular reactive oxygen species (ROS). In contrast, reducing production of intracellular ROS using N-acetylcysteine, a scavenger of ROS, concurrently suppressed honokiol-induced cellular autophagy. Consequently, honokiol stimulated phosphorylation of extracellular signal-regulated kinase (ERK)1/2. However, pretreatment of neuro-2a cells with PD98059, an inhibitor of ERK1/2, lowered honokiol-induced autophagy. The effects of honokiol on inducing autophagy and apoptosis of neuroblastoma cells were further confirmed using mouse neuroblastoma NB41A3 cells as our experimental model. Fascinatingly, treatment of neuroblastoma neuro-2a and NB41A3 cells with honokiol for 12 h did not affect cell autophagy or apoptosis but caused significant suppression of cell migration. Taken together, this study showed that honokiol can induce

  2. 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

    In children, neuroblastomas are the most common and deadly solid tumor. Our previous study showed that honokiol, a small-molecule polyphenol, can traverse the blood-brain barrier and kill neuroblastoma cells. In this study, we further investigated the mechanisms of honokiol-induced insults to neuroblastoma cells. Treatment of neuroblastoma neuro-2a cells with honokiol elevated the levels of microtubule-associated protein light chain 3 (LC3)-II and induced cell autophagy in time- and concentration-dependent manners. Interestingly, pretreatment with 3-methyladenine (3-MA), an inhibitor of autophagy, led to the simultaneous attenuation of honokiol-induced cell autophagy and apoptosis but did not influence cell necrosis. As to the mechanisms, exposure of neuro-2a cells to honokiol time-dependently decreased the amount of phosphatidylinositol 3-kinase (PI3K). Sequentially, honokiol downregulated phosphorylation of protein kinase B (Akt) and mammalian target of rapamycin (mTOR) in neuro-2a cells. Furthermore, honokiol elevated the levels of glucose-regulated protein (GpR)78, an endoplasmic reticular stress (ERS)-associated protein, and amounts of intracellular reactive oxygen species (ROS). In contrast, reducing production of intracellular ROS using N-acetylcysteine, a scavenger of ROS, concurrently suppressed honokiol-induced cellular autophagy. Consequently, honokiol stimulated phosphorylation of extracellular signal-regulated kinase (ERK)1/2. However, pretreatment of neuro-2a cells with PD98059, an inhibitor of ERK1/2, lowered honokiol-induced autophagy. The effects of honokiol on inducing autophagy and apoptosis of neuroblastoma cells were further confirmed using mouse neuroblastoma NB41A3 cells as our experimental model. Fascinatingly, treatment of neuroblastoma neuro-2a and NB41A3 cells with honokiol for 12 h did not affect cell autophagy or apoptosis but caused significant suppression of cell migration. Taken together, this study showed that honokiol can induce

  3. Growth-factor dependent expression of the translationally controlled tumour protein TCTP is regulated through the PI3-K/Akt/mTORC1 signalling pathway.

    PubMed

    Bommer, Ulrich-Axel; Iadevaia, Valentina; Chen, Jiezhong; Knoch, Bianca; Engel, Martin; Proud, Christopher G

    2015-08-01

    Translationally controlled tumour protein TCTP (gene symbol: TPT1) is a highly-conserved, cyto-protective protein implicated in many physiological and disease processes, in particular cancer, where it is associated with poor patient outcomes. To understand the mechanisms underlying the accumulation of high TCTP levels in cancer cells, we studied the signalling pathways that control translation of TCTP mRNA, which contains a 5'-terminal oligopyrimidine tract (5'-TOP). In HT29 colon cancer cells and in HeLa cells, serum increases the expression of TCTP two- and four-fold, respectively, and this is inhibited by rapamycin or mTOR kinase inhibitors. Polysome profiling and mRNA quantification indicate that these effects occur at the level of mRNA translation. Blocking this pathway upstream of mTOR complex 1 (mTORC1) by inhibiting Akt also prevented increases in TCTP levels in both HeLa and HT29 colon cancer cells, whereas knockout of TSC2, a negative regulator of mTORC1, led to derepression of TCTP synthesis under serum starvation. Overexpression of eIF4E enhanced the polysomal association of the TCTP mRNA, although it did not protect its translation from inhibition by rapamycin. Conversely, expression of a constitutively-active mutant of the eIF4E inhibitor 4E-BP1, which is normally inactivated by mTORC1, inhibited TCTP mRNA translation in HEK293 cells. Our results demonstrate that TCTP mRNA translation is regulated by signalling through the PI3-K/Akt/mTORC1 pathway. This explains why TCTP levels are frequently increased in cancers, since mTORC1 signalling is hyperactive in ~80% of tumours.

  4. IGF1R Derived PI3K/AKT Signaling Maintains Growth in a Subset of Human T-Cell Acute Lymphoblastic Leukemias

    PubMed Central

    Gusscott, Samuel; Jenkins, Catherine E.; Lam, Sonya H.; Giambra, Vincenzo; Pollak, Michael; Weng, Andrew P.

    2016-01-01

    Insulin-like growth factor 1 receptor (IGF1R) is a prevalent signaling pathway in human cancer that supports cell growth/survival and thus contributes to aggressive biological behavior. Much work has gone into development of IGF1R inhibitors; however, candidate agents including small molecule tyrosine kinase inhibitors and blocking antibodies have yet to fulfill their promise clinically. Understanding cellular features that define sensitivity versus resistance are important for effective patient selection and anticipation of outgrowth of a resistant clone. We previously identified an important role for IGF signaling in T-cell acute lymphoblastic leukemia (T-ALL) relying primarily upon genetically defined mouse models. We present here an assessment of IGF1R dependence in human T-ALL using a broad panel of 27 established cell lines that capture a spectrum of the genetic variation that might be encountered in clinical practice. We observed that a subset of cell lines are sensitive to IGF1R inhibition and are characterized by high levels of surface IGF1R expression and PTEN positivity. Interestingly, lentiviral expression or knock-down of PTEN in PTEN-negative/positive cell lines, respectively, had limited effects on their response to IGF1R inhibition, suggesting that PTEN contributes to, but does not define IGF dependence. Additionally, we characterize downstream PI3K/AKT signaling as dominant over RAS/RAF/MEK/ERK in mediating growth and/or survival in this context. Finally, we demonstrate that IGF and interleukin-7 (IL-7) fulfill non-overlapping roles in supporting T-ALL growth. These findings are significant in that they reveal cellular features and downstream mechanisms that may determine the response of an individual patient’s tumor to IGF1R inhibitor therapy. PMID:27532210

  5. IGF1R Derived PI3K/AKT Signaling Maintains Growth in a Subset of Human T-Cell Acute Lymphoblastic Leukemias.

    PubMed

    Gusscott, Samuel; Jenkins, Catherine E; Lam, Sonya H; Giambra, Vincenzo; Pollak, Michael; Weng, Andrew P

    2016-01-01

    Insulin-like growth factor 1 receptor (IGF1R) is a prevalent signaling pathway in human cancer that supports cell growth/survival and thus contributes to aggressive biological behavior. Much work has gone into development of IGF1R inhibitors; however, candidate agents including small molecule tyrosine kinase inhibitors and blocking antibodies have yet to fulfill their promise clinically. Understanding cellular features that define sensitivity versus resistance are important for effective patient selection and anticipation of outgrowth of a resistant clone. We previously identified an important role for IGF signaling in T-cell acute lymphoblastic leukemia (T-ALL) relying primarily upon genetically defined mouse models. We present here an assessment of IGF1R dependence in human T-ALL using a broad panel of 27 established cell lines that capture a spectrum of the genetic variation that might be encountered in clinical practice. We observed that a subset of cell lines are sensitive to IGF1R inhibition and are characterized by high levels of surface IGF1R expression and PTEN positivity. Interestingly, lentiviral expression or knock-down of PTEN in PTEN-negative/positive cell lines, respectively, had limited effects on their response to IGF1R inhibition, suggesting that PTEN contributes to, but does not define IGF dependence. Additionally, we characterize downstream PI3K/AKT signaling as dominant over RAS/RAF/MEK/ERK in mediating growth and/or survival in this context. Finally, we demonstrate that IGF and interleukin-7 (IL-7) fulfill non-overlapping roles in supporting T-ALL growth. These findings are significant in that they reveal cellular features and downstream mechanisms that may determine the response of an individual patient's tumor to IGF1R inhibitor therapy. PMID:27532210

  6. The phosphoinositide 3-kinase signaling pathway in normal and malignant B cells: activation mechanisms, regulation and impact on cellular functions.

    PubMed

    Pauls, Samantha D; Lafarge, Sandrine T; Landego, Ivan; Zhang, Tingting; Marshall, Aaron J

    2012-01-01

    The phosphoinositide 3-kinase (PI3K) pathway is a central signal transduction axis controlling normal B cell homeostasis and activation in humoral immunity. The p110δ PI3K catalytic subunit has emerged as a critical mediator of multiple B cell functions. The activity of this pathway is regulated at multiple levels, with inositol phosphatases PTEN and SHIP both playing critical roles. When deregulated, the PI3K pathway can contribute to B cell malignancies and autoantibody production. This review summarizes current knowledge on key mechanisms that activate and regulate the PI3K pathway and influence normal B cell functional responses including the development of B cell subsets, antigen presentation, immunoglobulin isotype switch, germinal center responses, and maintenance of B cell anergy. We also discuss PI3K pathway alterations reported in select B cell malignancies and highlight studies indicating the functional significance of this pathway in malignant B cell survival and growth within tissue microenvironments. Finally, we comment on early clinical trial results, which support PI3K inhibition as a promising treatment of chronic lymphocytic leukemia.

  7. Notoginsenoside Ft1 Promotes Fibroblast Proliferation via PI3K/Akt/mTOR Signaling Pathway and Benefits Wound Healing in Genetically Diabetic Mice.

    PubMed

    Zhang, Eryun; Gao, Bo; Yang, Li; Wu, Xiaojun; Wang, Zhengtao

    2016-02-01

    Wound healing requires the essential participation of fibroblasts, which is impaired in diabetic foot ulcers (DFU). Notoginsenoside Ft1 (Ft1), a saponin from Panax notoginseng, can enhance platelet aggregation by activating signaling network mediated through P2Y12 and induce proliferation, migration, and tube formation in cultured human umbilical vein endothelial cells. However, whether it can accelerate fibroblast proliferation and benefit wound healing, especially DFU, has not been elucidated. In the present study on human dermal fibroblast HDF-a, Ft1 increased cell proliferation and collagen production via PI3K/Akt/mTOR signaling pathway. On the excisional wound splinting model established on db/db diabetic mouse, topical application of Ft1 significantly shortened the wound closure time by 5.1 days in contrast with phosphate-buffered saline (PBS) treatment (15.8 versus 20.9 days). Meanwhile, Ft1 increased the rate of re-epithelialization and the amount of granulation tissue at day 7 and day 14. The molecule also enhanced mRNA expressions of COL1A1, COL3A1, transforming growth factor (TGF)-β1 and TGF-β3 and fibronectin, the genes that contributed to collagen expression, fibroblast proliferation, and consequent scar formation. Moreover, Ft1 facilitated the neovascularization accompanied with elevated vascular endothelial growth factor, platelet-derived growth factor, and fibroblast growth factor at either mRNA or protein levels and alleviated the inflammation of infiltrated monocytes indicated by reduced tumor necrosis factor-α and interleukin-6 mRNA expressions in the diabetic wounds. Altogether, these results indicated that Ft1 might accelerate diabetic wound healing by orchestrating multiple processes, including promoting fibroblast proliferation, enhancing angiogenesis, and attenuating inflammatory response, which provided a great potential application of it in clinics for patients with DFU. PMID:26567319

  8. Simultaneous Inhibition of EGFR and PI3K Enhances Radiosensitivity in Human Breast Cancer

    SciTech Connect

    Li Ping; Zhang Qing; Torossian, Artour; Li Zhaobin; Xu Wencai; Lu Bo; Fu Shen

    2012-07-01

    Purpose: Mutations in the epidermal growth factor receptor (EGFR)/phosphoinositide 3-kinase (PI3K)/Akt signaling transduction pathway are common in cancer. This pathway is imperative to the radiosensitivity of cancer cells. We aimed to investigate the radiosensitizing effects of the simultaneous inhibition of EGFR and PI3K in breast cancer cells. Methods and Materials: MCF-7 cell lines with low expression of EGFR and wild-type PTEN and MDA-MB-468 cell lines with high expression of EGFR and mutant PTEN were used. The radiosensitizing effects by the inhibition of EGFR with AG1478 and/or PI3K with Ly294002 were determined by colony formation assay, Western blot was used to investigate the effects on downstream signaling. Flow cytometry was used for apoptosis and cell cycle analysis. Mice-bearing xenografts of MDA-MB-468 breast cancer cells were also used to observe the radiosensitizing effect. Results: Simultaneous inhibition of EGFR and PI3K greatly enhanced radiosensitizing effect in MDA-MB-468 in terms of apoptosis and mitotic death, either inhibition of EGFR or PI3K alone could enhance radiosensitivity with a dose-modifying factor (DMF{sub SF2}) of 1.311 and 1.437, radiosensitizing effect was further enhanced by simultaneous inhibition of EGFR and PI3K with a DMF{sub SF2} at 2.698. DNA flow cytometric analysis indicated that dual inhibition combined with irradiation significantly induced G0/G1 phase arrest in MDA-MB-468 cells. The expression of phosphor-Akt and phosphor-Erk1/2 (induced by irradiation and PI3K inhibitor) were fully attenuated by simultaneous treatment with both inhibitors in combination with irradiation. In addition, dual inhibition combined with irradiation induced dramatic tumor growth delay in MDA-MB-468 xenografts. Conclusions: Our study indicated that simultaneous inhibition of EGFR and PI3K could further sensitize the cancer cells to irradiation compared to the single inhibitor with irradiation in vitro and in vivo. The approach may have

  9. Leader cells regulate collective cell migration via Rac activation in the downstream signaling of integrin β1 and PI3K.

    PubMed

    Yamaguchi, Naoya; Mizutani, Takeomi; Kawabata, Kazushige; Haga, Hisashi

    2015-01-07

    Collective cell migration plays a crucial role in several biological processes, such as embryonic development, wound healing, and cancer metastasis. Here, we focused on collectively migrating Madin-Darby Canine Kidney (MDCK) epithelial cells that follow a leader cell on a collagen gel to clarify the mechanism of collective cell migration. First, we removed a leader cell from the migrating collective with a micromanipulator. This then caused disruption of the cohesive migration of cells that followed in movement, called "follower" cells, which showed the importance of leader cells. Next, we observed localization of active Rac, integrin β1, and PI3K. These molecules were clearly localized in the leading edge of leader cells, but not in follower cells. Live cell imaging using active Rac and active PI3K indicators was performed to elucidate the relationship between Rac, integrin β1, and PI3K. Finally, we demonstrated that the inhibition of these molecules resulted in the disruption of collective migration. Our findings not only demonstrated the significance of a leader cell in collective cell migration, but also showed that Rac, integrin β1, and PI3K are upregulated in leader cells and drive collective cell migration.

  10. 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.

  11. 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

  12. Isoquercitrin Inhibits Hydrogen Peroxide-Induced Apoptosis of EA.hy926 Cells via the PI3K/Akt/GSK3β Signaling Pathway.

    PubMed

    Zhu, Meixia; Li, Jiankuan; Wang, Ke; Hao, Xuliang; Ge, Rui; Li, Qingshan

    2016-01-01

    Oxidative stress plays a critical role in endothelial injury and the pathogenesis of diverse cardiovascular diseases, including atherosclerosis. Isoquercitrin (quercetin-3-glucoside), a flavonoid distributed widely in plants, exhibits many biological activities, including anti-allergic, anti-viral, anti-inflammatory, and anti-oxidative effects. In the present study, the inhibitory effect of isoquercitrin on H2O2-induced apoptosis of EA.hy926 cells was evaluated. MTT assays showed that isoquercitrin significantly inhibited H2O2-induced loss of viability in EA.hy926 cells. Hoechst33342/PI and Annexin V-FITC/PI fluorescent double staining indicated that isoquercitrin inhibited H2O2-induced apoptosis of EA.hy926 cells. Western blotting demonstrated that isoquercitrin prevented H2O2-induced increases in cleaved caspase-9 and cleaved caspase-3 expression, while increasing expression of anti-apoptotic protein Mcl-1. Additionally, isoquercitrin significantly increased the expression of p-Akt and p-GSK3β in a dose-dependent manner in EA.hy926 cells. LY294002, a PI3K/Akt inhibitor, inhibited isoquercitrin-induced GSK3β phosphorylation and increase of Mcl-1 expression, which indicated that regulation of isoquercitrin on Mcl-1 expression was likely related to the modulation of Akt activation. These results demonstrated that the anti-apoptotic effect of isoquercitrin on H2O2-induced EA.hy926 cells was likely associated with the regulation of isoquercitrin on Akt/GSK3β signaling pathway and that isoquercitrin could be used clinically to interfere with the progression of endothelial injury-associated cardiovascular disease. PMID:27007368

  13. Activation of the PI3K/Akt signal transduction pathway and increased levels of insulin receptor in protein repair-deficient mice.

    PubMed

    Farrar, Christine; Houser, Carolyn R; Clarke, Steven

    2005-02-01

    Protein L-isoaspartate (D-aspartate) O-methyltransferase is an enzyme that catalyses the repair of isoaspartyl damage in proteins. Mice lacking this enzyme (Pcmt1-/- mice) have a progressive increase in brain size compared with wild-type mice (Pcmt1+/+ mice), a phenotype that can be associated with alterations in the PI3K/Akt signal transduction pathway. Here we show that components of this pathway, including Akt, GSK3beta and PDK-1, are more highly phosphorylated in the brains of Pcmt1-/- mice, particularly in cells of the hippocampus, in comparison with Pcmt1+/+ mice. Examination of upstream elements of this pathway in the hippocampus revealed that Pcmt1-/- mice have increased activation of insulin-like growth factor-I (IGF-I) receptor and/or insulin receptor. Western blot analysis revealed an approximate 200% increase in insulin receptor protein levels and an approximate 50% increase in IGF-I receptor protein levels in the hippocampus of Pcmt1-/- mice. Higher levels of the insulin receptor protein were also found in other regions of the adult brain and in whole tissue extracts of brain, liver, heart and testes of both juvenile and adult Pcmt1-/- mice. There were no significant differences in plasma insulin levels for adult Pcmt1-/- mice during glucose tolerance tests. However, they did show higher peak levels of blood glucose, suggesting a mild impairment in glucose tolerance. We propose that Pcmt1-/- mice have altered regulation of the insulin pathway, possibly as a compensatory response to altered glucose uptake or metabolism or as an adaptive response to a general accumulation of isoaspartyl protein damage in the brain and other tissues.

  14. Fine-Tuning of PI3K/AKT Signalling by the Tumour Suppressor PTEN Is Required for Maintenance of Flight Muscle Function and Mitochondrial Integrity in Ageing Adult Drosophila melanogaster

    PubMed Central

    Mensah, Lawrence B.; Davison, Claire; Fan, Shih-Jung; Morris, John F.; Goberdhan, Deborah C. I.; Wilson, Clive

    2015-01-01

    Insulin/insulin-like growth factor signalling (IIS), acting primarily through the PI3-kinase (PI3K)/AKT kinase signalling cassette, plays key evolutionarily conserved regulatory roles in nutrient homeostasis, growth, ageing and longevity. The dysfunction of this pathway has been linked to several age-related human diseases including cancer, Type 2 diabetes and neurodegenerative disorders. However, it remains unclear whether minor defects in IIS can independently induce the age-dependent functional decline in cells that accompany some of these diseases or whether IIS alters the sensi