Jun Song

University of Kentucky, Lexington, KY, United States

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Publications (15)83.59 Total impact

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    ABSTRACT: The precise involvement of the PI3K/mTOR and RAS/MEK pathways in carcinoid tumors is not well defined. The purpose of our study was to evaluate the role these pathways play in carcinoid cell proliferation, apoptosis, and secretion and to determine the effects of combined treatment on carcinoid tumor inhibition. The human neuroendocrine cell lines BON (pancreatic carcinoid), NCI-H727 (lung carcinoid), and QGP-1 (somatostatinoma) were treated with either the pan-PI3K inhibitor, BKM120, or the dual PI3K-mTOR inhibitor, BEZ235, alone or in combination with the MEK inhibitor, PD0325901; proliferation, apoptosis, and protein expression were assessed. Peptide secretion was evaluated in BON and QGP-1 cells. The anti-proliferative effect of BEZ235, alone or combined with PD0325901, was then tested in vivo. Both BKM120 and BEZ235 decreased proliferation and increased apoptosis; combination with PD0325901 significantly enhanced the antineoplastic effects of either treatment alone. In contrast, neurotensin (NT) peptide secretion was markedly stimulated with BKM120 treatment, but not BEZ235. The combination of BEZ235 + PD0325901 significantly inhibited the growth of BON xenografts without systemic toxicity. Both BKM120 and BEZ235 effectively inhibited NET cell proliferation and stimulated apoptosis. However, inhibition of the PI3K pathway alone with BKM120 significantly stimulated NT peptide secretion; this did not occur with the dual inhibition of both PI3K and mTOR using BEZ235 suggesting that this would be a more effective treatment regimen for NETs. Moreover, the combination of BEZ235 and the MEK inhibitor PD0325901 was a safe and more effective therapy in vivo compared with single agents alone.
    Clinical Cancer Research 01/2014; · 7.84 Impact Factor
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    ABSTRACT: Neurotensin (NT), an intestinal peptide secreted from N cells in the small bowel, regulates a variety of physiological functions of the gastrointestinal tract, including secretion, gut motility, and intestinal growth. The class IA phosphatidylinositol 3-kinase (PI3K) family, which comprised of p110 catalytic (α, β and δ) and p85 regulatory subunits, has been implicated in the regulation of hormone secretion from endocrine cells. However, the underlying mechanisms remain poorly understood. In particular, the role of PI3K in intestinal peptide secretion is not known. Here, we show that PI3K catalytic subunit, p110α, negatively regulates NT secretion in vitro and in vivo. We demonstrate that inhibition of p110α, but not p110β, induces NT release in BON, a human endocrine cell line, which expresses NT mRNA and produces NT peptide in a manner analogous to N cells, and QGP-1, a pancreatic endocrine cell line that produces NT peptide. In contrast, overexpression of p110α decreases NT secretion. Consistently, p110α-inhibition increases plasma NT levels in mice. To further delineate the mechanisms contributing to this effect, we demonstrate that inhibition of p110α increases NT granule trafficking by up-regulating α-tubulin acetylation; NT secretion is prevented by overexpression of HDAC6, an α-tubulin deacetylase. Moreover, ras-related protein Rab27A (a small G protein) and kinase D-interacting substrate of 220 kDa (Kidins220), which are associated with NT granules, play a negative and positive role, respectively, in p110α-inhibition-induced NT secretion. Our findings identify the critical role and novel mechanisms for the PI3K signaling pathway in the control of intestinal hormone granule transport and release.
    Molecular Endocrinology 06/2012; 26(8):1380-93. · 4.75 Impact Factor
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    ABSTRACT: The mammalian target of rapamycin (mTOR) signaling exists in two complexes: mTORC1 and mTORC2. Neurotensin (NT), an intestinal hormone secreted by enteroendocrine (N) cells in the small bowel, has important physiological effects in the gastrointestinal tract. The human endocrine cell line BON abundantly expresses the NT gene and synthesizes and secretes NT in a manner analogous to that of N cells. Here, we demonstrate that the inhibition of mTORC1 by rapamycin (mTORC1 inhibitor), torin1 (both mTORC1 and mTORC2 inhibitor) or short hairpin RNA-mediated knockdown of mTOR, regulatory associated protein of mTOR (RAPTOR), and p70 S6 kinase (p70S6K) increased basal NT release via upregulating NT gene expression in BON cells. c-Jun activity was increased by rapamycin or torin1 or p70S6K knockdown. c-Jun overexpression dramatically increased NT promoter activity, which was blocked by PD98059, an mitogen-activated protein kinase kinase (MEK) inhibitor. Furthermore, overexpression of MEK1 or extracellular signal-regulated kinase 1 (ERK1) increased c-Jun expression and NT promoter activity. More importantly, PD98059 blocked rapamycin- or torin1-enhanced NT secretion. Consistently, rapamycin and torin1 also increased NT gene expression in Hep3B cells, a human hepatoma cell line that, similar to BON, expresses high levels of NT. Phosphorylation of c-Jun and ERK1/2 was also increased by rapamycin and torin1 in Hep3B cells. Finally, we showed activation of mTOR in BON cells treated with amino acids, high glucose, or serum and, concurrently, the attenuation of ERK1/2 and c-Jun phosphorylation and NT secretion. Together, mTORC1, as a nutrient sensor, negatively regulates NT secretion via the MEK/ERK/c-Jun signaling pathway. Our results identify a physiological link between mTORC1 and MEK/ERK signaling in controlling intestinal hormone gene expression and secretion.
    AJP Cell Physiology 04/2011; 301(1):C213-26. · 3.71 Impact Factor
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    ABSTRACT: Reactive oxygen species (ROS) are thought to contribute to the pathogenesis of necrotizing enterocolitis (NEC). Mitochondria as a major source of intracellular ROS and apoptotic signaling during oxidative stress in NEC have not been investigated. We sought to determine: (1) the effects of oxidative stress on intestinal mitochondrial apoptotic signaling, and (2) the role of growth factors in this process. We used Swiss-Webster mice pups, and rat intestinal epithelial (RIE)-1, mitochondrial DNA-depleted RIE-1 cell line (RIE-1-ρ°) and human fetal intestinal epithelial cells (FHs74 Int) for our studies. H(2)O(2) induced apoptosis and ROS production. ROS-mediated activation of apoptotic signaling was significantly attenuated with mitochondrial silencing in RIE-1-ρ° cells. Growth factors, especially IGF-1, attenuated this response to H(2)O(2) in intestinal epithelial cells. Our findings suggest that mitochondria are a major source of intestinal apoptotic signaling during oxidative stress, and modulating mitochondrial apoptotic responses may help ameliorate the effects of NEC.
    Pediatric Surgery International 03/2011; 27(8):871-7. · 1.22 Impact Factor
  • Gastroenterology 01/2011; 140(5). · 12.82 Impact Factor
  • Gastroenterology 01/2011; 140(5). · 12.82 Impact Factor
  • Advanced Materials Research 01/2010;
  • Jun Song, Jing Li, Mark Evers
    Gastroenterology 01/2010; 138(5). · 12.82 Impact Factor
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    ABSTRACT: Oxidative stress and inflammation may contribute to the disruption of the protective gut barrier through various mechanisms; mitochondrial dysfunction resulting from inflammatory and oxidative injury may potentially be a significant source of apoptosis during necrotizing enterocolitis (NEC). Tumor necrosis factor (TNF)-alpha is thought to generate reactive oxygen species (ROS) and activate the apoptosis signal-regulating kinase 1 (ASK1)-c-Jun N-terminal kinase (JNK)/p38 pathway. Hence, the focus of our study was to examine the effects of TNF-alpha/ROS on mitochondrial function, ASK1-JNK/p38 cascade activation in intestinal epithelial cells during NEC. We found (a) abundant tissue TNF-alpha and ASK1 expression throughout all layers of the intestine in neonates with NEC, suggesting that TNF-alpha/ASK1 may be a potential source (indicators) of intestinal injury in neonates with NEC; (b) TNF-alpha-induced rapid and transient activation of JNK/p38 apoptotic signaling in all cell lines suggests that this may be an important molecular characteristic of NEC; (c) TNF-alpha-induced rapid and transient ROS production in RIE-1 cells indicates that mitochondria are the predominant source of ROS, demonstrated by significantly attenuated response in mitochondrial DNA-depleted (RIE-1-rho) intestinal epithelial cells; (d) further studies with mitochondria-targeted antioxidant PBN supported our hypothesis that effective mitochondrial ROS trapping is protective against TNF-alpha/ROS-induced intestinal epithelial cell injury; (e) TNF-alpha induces significant mitochondrial dysfunction in intestinal epithelial cells, resulting in increased production of mtROS, drop in mitochondrial membrane potential (MMP) and decreased oxygen consumption; (f) although the significance of mitochondrial autophagy in NEC has not been unequivocally shown, our studies provide a strong preliminary indication that TNF-alpha/ROS-induced mitochondrial autophagy may play a role in NEC, and this process is a late phenomenon. Paraffin-embedded intestinal sections from neonates with NEC and non-inflammatory condition of the gastrointestinal tract undergoing bowel resections were analyzed for TNF-alpha and ASK1 expression. Rat (RIE-1) and mitochondrial DNA-depleted (RIE-1-rho) intestinal epithelial cells were used to determine the effects of TNF-alpha on mitochondrial function. Our findings suggest that TNF-alpha induces significant mitochondrial dysfunction and activation of mitochondrial apoptotic responses, leading to intestinal epithelial cell apoptosis during NEC. Therapies directed against mitochondria/ROS may provide important therapeutic options, as well as ameliorate intestinal epithelial cell apoptosis during NEC.
    Oxidative Medicine and Cellular Longevity 01/2009; 2(5):297-306.
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    ABSTRACT: Previously, we demonstrated that protein kinase D (PKD) plays a protective role during H(2)O(2)-induced intestinal cell death. Here, we sought to determine whether this effect is mediated by nuclear factor-kappaB (NF-kappaB) and mitogen-activated protein kinases (MAPKs). Treatment with H(2)O(2) activated NF-kappaB in RIE-1 cells; H(2)O(2) also induced the translocation of NF-kappaB p65 as well as phosphorylation of IkappaB-alpha. PKD1 siRNA inhibited H(2)O(2)-induced activation, translocation of NF-kappaB, and phosphorylation of IkappaB-alpha. We also found that overexpression of wild type PKD1 attenuated H(2)O(2)-induced phosphorylation of p38 MAPK and its upstream activator, MAPK kinase (MKK) 3/6, whereas the phosphorylation was increased by PKD1 siRNA or kinase-dead PKD1. Phosphorylation of neither extracellular signal-regulated kinases (ERK) 1/2 nor c-Jun N-terminal kinases (JNK) was altered by PKD1 plasmids or siRNA. Our findings suggest that PKD protects intestinal cells through up-regulation of NF-kappaB and down-regulation of p38 MAPK.
    Biochemical and Biophysical Research Communications 01/2009; 378(3):610-4. · 2.28 Impact Factor
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    ABSTRACT: We recently demonstrated that protein kinase D (PKD) exerts a protective function during oxidative stress-induced intestinal epithelial cell injury; however, the exact role of DAG kinase (DGK)zeta, an isoform expressed in intestine, during this process is unknown. We sought to determine the role of DGK during oxidative stress-induced intestinal cell injury and whether DGK acts as an upstream regulator of PKD. Inhibition of DGK with R59022 compound or DGKzeta siRNA transfection decreased H2O2-induced RIE-1 cell apoptosis as measured by DNA fragmentation and increased PKD phosphorylation. Overexpression of kinase-dead DGKzeta also significantly increased PKD phosphorylation. Additionally, endogenous nuclear DGKzeta rapidly translocated to the cytoplasm following H2O2 treatment. Our findings demonstrate that DGK is involved in the regulation of oxidative stress-induced intestinal cell injury. PKD activation is induced by DGKzeta, suggesting DGK is an upstream regulator of oxidative stress-induced activation of the PKD signaling pathway in intestinal epithelial cells.
    Biochemical and Biophysical Research Communications 11/2008; 375(2):200-4. · 2.28 Impact Factor
  • Gastroenterology 01/2008; 134(4). · 12.82 Impact Factor
  • Journal of the American College of Surgeons 09/2007; 205(3):S51. · 4.50 Impact Factor
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    ABSTRACT: Reactive oxygen species (ROS) are involved in the pathogenesis of necrotizing enterocolitis (NEC) in premature infants. We have recently found that activation of multiple cellular signaling transduction pathways occurs during ROS-induced intestinal cell apoptosis; the phosphatidylinositol 3-kinase (PI3-K) pathway plays an anti-apoptotic role during this process. Insulin-like growth factor (IGF)-1 activates PI3-K pathway to promote cell survival; however, the effects of IGF-1 treatment during gut injury are not clearly defined. The purpose of this study was to determine whether IGF-1 protects intestinal cells from ROS-induced apoptosis. Rat intestinal epithelial (RIE)-1 cells were treated with either IGF-1 (100 nm), hydrogen peroxide (H2O2; 500 microm), or combination. Western blotting was performed to assess phosphorylation of Akt, a downstream effector of PI3-K. Cell Death Detection ELISA, DCHF, and JC-1 assays were performed to demonstrate protective effects of IGF-1. Wortmannin, an inhibitor of PI3-K, was used to show PI3-K-dependent mechanism of action for IGF-1. H2O2 treatment resulted in increased intestinal epithelial cell apoptosis with intracellular ROS generation and mitochondrial membrane depolarization; IGF-1 pre-treatment attenuated this response without affecting ROS production. H2O2-induced phosphorylation of Akt was further increased with IGF-1 treatment; wortmannin abolished these effects in RIE-1 cells. PI3-K pathway is activated during ROS-induced intestinal epithelial cell injury; IGF-1 exerted an anti-apoptotic effect during this response by PI3-K activation. A better understanding of the exact role of IGF-1-mediated activation of PI3-K may allow us to facilitate the development of novel therapy against NEC.
    Journal of Surgical Research 12/2006; 136(1):31-7. · 2.02 Impact Factor
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    ABSTRACT: Protein kinase D (PKD) is a novel protein serine kinase that has recently been implicated in diverse cellular functions, including apoptosis and cell proliferation. The purpose of our present study was 1) to define the activation of PKD in intestinal epithelial cells treated with H2O2, an agent that induces oxidative stress, and 2) to delineate the upstream signaling mechanisms mediating the activation of PKD. We found that the activation of PKD is induced by H2O2 in both a dose- and time-dependent fashion. PKD phosphorylation was attenuated by rottlerin, a selective PKC-delta inhibitor, and by small interfering RNA (siRNA) directed against PKC-delta, suggesting the regulation of PKD activity by upstream PKC-delta. Activation of PKD was also blocked by a Rho kinase (ROK)-specific inhibitor, Y-27632, as well as by C3, a Rho protein inhibitor, demonstrating that the Rho/ROK pathway also mediates PKD activity in intestinal cells. In addition, H2O2-induced PKC-delta phosphorylation was inhibited by C3 treatment, further suggesting that PKC-delta is downstream of Rho/ROK. Interestingly, H2O2-induced intestinal cell apoptosis was enhanced by PKD siRNA. Together, these results clearly demonstrate that oxidative stress induces PKD activation in intestinal epithelial cells and that this activation is regulated by upstream PKC-delta and Rho/ROK pathways. Importantly, our findings suggest that PKD activation protects intestinal epithelial cells from oxidative stress-induced apoptosis. These findings have potential clinical implications for intestinal injury associated with oxidative stress (e.g., necrotizing enterocolitis in infants).
    AJP Cell Physiology 07/2006; 290(6):C1469-76. · 3.71 Impact Factor

Publication Stats

127 Citations
83.59 Total Impact Points

Institutions

  • 2011–2012
    • University of Kentucky
      • Department of Surgery
      Lexington, KY, United States
  • 2006–2011
    • University of Texas Medical Branch at Galveston
      • Department of Surgery
      Galveston, TX, United States