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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 1,369 records · Page 76
All sourcesNASA NTRSDOE OSTI
DOE OSTI2016

GSK621 activates AMPK signaling to inhibit LPS-induced TNFα production

LPS stimulation in macrophages/monocytes induces TNFα production. We here tested the potential effect of GSK621, a novel AMP-activated protein kinase (AMPK) activator, against the process. In RAW264.7 macrophages, murine bone marrow-derived macrophages (BMDMs), and chronic obstructive pulmonary disease (COPD) patients' monocytes, GSK621 significantly inhibited LPS-induced TNFα protein secretion and mRNA synthesis. Inhibition of AMPK, through AMPKα shRNA knockdown or dominant negative mutation (T172A), almost abolished GSK621's suppression on TNFα in RAW264.7 cells. Reversely, forced-expression of a constitutively-active AMPKα (T172D) mimicked GSK621 actions and reduced LPS-induced TNFα production. Molecularly, GSK621 suppressed LPS-induced reactive oxygen species (ROS) production and nuclear factor kappa B (NFκB) activation. In vivo, GSK621 oral administration inhibited LPS-induced TNFα production and endotoxin shock in mice. In summary, GSK621 activates AMPK signaling to inhibit LPS-induced TNFα production in macrophages/monocytes. - Highlights: • GSK621 inhibits LPS-induced TNFα production/expression in RAW264.7 cells and BMDMs. • GSK621 inhibits LPS-induced TNFα production/expression in COPD patients' PBMCs. • GSK621's inhibition on TNFα production by LPS requires AMPK activation. • GSK621 inhibits LPS-induced ROS production and NFκB activation, dependent on AMPK. • GSK621 oral administration inhibits LPS-induced TNFα production and endotoxin shock in mice.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Autophagy prevention sensitizes AKTi-1/2-induced anti-hepatocellular carcinoma cell activity in vitro and in vivo

Molecule-targeted therapy has become the research focus for hepatocellular carcinoma (HCC). Persistent PI3K-AKT activation is often detected in HCC, representing a valuable oncotarget for treatment. Here, we tested the anti-HCC activity by a potent AKT inhibitor: AKT inhibitor 1/2 (AKTi-1/2). In both established (HepG2 and Huh-7) and primary human HCC cells, treatment with AKTi-1/2 inhibited cell survival and proliferation, but induced cell apoptosis. AKTi-1/2 blocked AKT-mTOR activation, yet simultaneously provoked cytoprotective autophagy in HCC cells. The latter was evidenced by ATG-5 and Beclin-1 upregulation, p62 downregulation as well as LC3B-GFP puncta formation. Autophagy inhibition, via pharmacological inhibitors (3-methyladenine, ammonium chloride, and bafilomycin A1) or Beclin-1 siRNA knockdown, significantly potentiated AKTi-1/2-induced HepG2 cell death and apoptosis. In nude mice, AKTi-1/2 intraperitoneal injection inhibited HepG2 tumor growth. Significantly, its anti-tumor activity in vivo was further sensitized when combined with Beclin-1 shRNA knockdown in HepG2 tumors. Together, these results demonstrate that autophagy activation serves as a main resistance factor of AKTi-1/2 in HCC cells. Autophagy prevention therefore sensitizes AKTi-1/2-induced anti-HCC activity in vitro and in vivo. - Highlights: • AKTi-1/2 inhibits human HCC cells in vitro. • Autophagy inhibitors sensitize AKTi-1/2-induced HCC cell death and apoptosis. • Beclin-1 siRNA potentiates AKTi-1/2-induced HepG2 cell death and apoptosis. • Beclin-1 knockdown augments AKTi-1/2-induced anti-HepG2 tumor activity in vivo.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

p53R2 regulates thioredoxin reductase activity through interaction with TrxR2

Ribonucleotide reductase small subunit p53R2 is a member of the ribonucleotide reductase family that supplies dNTPs for nuclear and mitochondrial DNA replication and repair. Here, we have identified a mitochondrial thioredoxin reductase 2 (TrxR2) as a novel p53R2-binding protein. We demonstrated a direct interaction between the two, and observed that p53R2 stimulated the enzymatic activity of TrxR in vitro. Moreover, TrxR2 activity was significantly lower in p53R2 knockdown cells, and increased when p53R2 was overexpressed, effects that were independent of p53. Furthermore, p53R2 knockdown suppressed UV-induced TrxR activity. These findings suggest that p53R2 acts as a positive regulator of TrxR2 activity in mitochondria both under normal physiological conditions and during the cellular response to DNA damage. - Highlights: • P53R2 binds to thioredoxin reductase 2 (TrxR2) in vivo and in vitro. • P53R2 enhances TrxR activity. • P53R2 regulates TrxR activity in a p53-independent manner. • P53R2 acts as a positive regulator of TrxR2 activity under both normal and DNA damage conditions.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Activation of EVI1 transcription by the LEF1/β-catenin complex with p53-alteration in myeloid blast crisis of chronic myeloid leukemia

The presence of a BCR-ABL1 fusion gene is necessary for the pathogenesis of chronic myeloid leukemia (CML) through t(9;22)(q34;q11) translocation. Imatinib, an ABL tyrosine kinase inhibitor, is dramatically effective in CML patients; however, 30% of CML patients will need further treatment due to progression of CML to blast crisis (BC). Aberrant high expression of ecotropic viral integration site 1 (EVI1) is frequently observed in CML during myeloid-BC as a potent driver with a CML stem cell signature; however, the precise molecular mechanism of EVI1 transcriptional regulation during CML progression is poorly defined. Here, we demonstrate the transcriptional activity of EVI1 is dependent on activation of lymphoid enhancer-binding factor 1 (LEF1)/β-catenin complex by BCR-ABL with loss of p53 function during CML-BC. The activation of β-catenin is partly dependent on BCR-ABL expression through enhanced GSK3β phosphorylation, and EVI1 expression is directly enhanced by the LEF1/β-catenin complex bound to the EVI1 promoter region. Moreover, the loss of p53 expression is inversely correlated with high expression of EVI1 in CML leukemia cells with an aggressive phase of CML, and a portion of the activation mechanism of EVI1 expression is dependent on β-catenin activation through GSK3β phosphorylation by loss of p53. Therefore, we found that the EVI1 activation in CML-BC is dependent on LEF1/β-catenin activation by BCR-ABL expression with loss of p53 function, representing a novel selective therapeutic approach targeting myeloid blast crisis progression. - Highlights: • Transcriptional regulation of EVI1 in CML-BC is proposed. • EVI1 transcription is directly regulated by LEF1/β-catenin complex in CML-BC. • Loss of p53 function as a key regulator for β-catenin-EVI1 in CML myeloid-BC. • The LEF1/β-catenin binding site on the EVI1 promoter is a new target for CML-BC.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Activation of Nrf2 attenuates carbonyl stress induced by methylglyoxal in human neuroblastoma cells: Increase in GSH levels is a critical event for the detoxification mechanism

The present study focused on the methylglyoxal (MG) detoxification mechanism in neuroblastoma cells. The involvement of nuclear factor erythroid 2-related factor 2 (Nrf2)/Kelch-like ECH-associated protein 1 (Keap1) pathway as a defense response against the formation of MG-modified proteins, which is well-known evidence of carbonyl stress, was also examined. We found that MG treatment resulted in accumulation of modified proteins bearing the structure of advanced glycation end products (AGEs) derived from MG in SH-SY5Y cells. This accumulation was suppressed by activation of the Nrf2 pathway prior to MG exposure via pre-treatment with an Nrf2 activator, carnosic acid and CDDO-Im, confirming the involvement of the Nrf2 pathway in MG detoxification. Although pre-treatment with the Nrf2 activator did not affect mRNA levels of GLO1, AKR1B1, and AKR7A2, the expressions of GCL and xCT mRNA, involved in GSH synthesis, were induced prior to increase in GSH levels. Furthermore, we demonstrated that a GSH synthesis inhibitor eliminated the MG detoxification effect derived from pretreatment with the Nrf2 activator. These results indicated that increase in GSH levels, induced by pre-treatment with carnosic acid, promoted the formation of the GLO1 substrate, hemithioacetal, thereby accelerating MG metabolism via the glyoxalase system and suppressing its toxicity. It was, therefore, determined that promotion of GSH synthesis via the Nrf2/Keap1pathway is important in the MG detoxification mechanism against neuronal MG-induced carbonyl stress, and Nrf2 activators contribute to reduction in the accumulation and toxic expression of carbonyl proteins. - Highlights: • Nrf2 activator prevents methylglyoxal-induced cytotoxicity and protein modifications. • MG treatment resulted in accumulation of modified proteins bearing the structure of CEL and MG-H1. • The promotion of GSH synthesis by activation of Nrf2 is a key response to MG detoxification.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Tyrosine kinase activity of EphA2 promotes its S897 phosphorylation and glioblastoma cell proliferation

Highlights: • Overexpression of EphA2 induces its S897 phosphorylation without growth factor stimulation. • Tyrosine kinase of EphA2 promotes EphA2-induced S897 phosphorylation. • Overexpression of EphA2 induces ERK activation through its tyrosine kinase activity. • ERK activation is required for EphA2-induced S897 phosphorylation. • Tyrosine kinase of EphA2 promotes glioblastoma cell proliferation. EphA2, a member of the Eph family of receptor tyrosine kinases, has been reported to promote tumor malignancy through phosphorylation of serine 897 (S897). Here, we found that overexpression of wild-type EphA2 induced S897 phosphorylation through ERK activation without growth factors or cytokines and promoted glioblastoma cell proliferation. However, overexpression of a kinase-inactive mutant of EphA2 failed to induce ERK activation, S897 phosphorylation, and promotion of glioblastoma cell proliferation. These data suggest that when overexpressed, EphA2 induces ERK activation through its tyrosine kinase activity, leading to S897 phosphorylation and promotion of glioblastoma cell proliferation. Our findings provide a new insight into how EphA2 mediates glioblastoma progression.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Rg1 inhibits high glucose-induced mesenchymal activation and fibrosis via regulating miR-2113/RP11-982M15.8/Zeb1 pathway

Highlights: • High glucose conditions up-regulate MMP-2, MMP-9 and down-regulate TIMP-2, and promote mesenchymal activation in Müller cells. • Inhibition of miR-2113 and over-expression of RP11-982M15.8 significantly restored the HG induced mesenchymal activasion. • Taken together, Rg1 inhibited HG-induced mesenchymal activation and fibrosis via regulating miR-2113/RP11-982M15.8/Zeb1 pathway. Recent study has showed that Ginsenoside Rg1, the mian active compound of Panax ginseng, could ameliorate oxidative stress and myocardial apoptosis in diabetes mellitus. However, the roles and mechanisms of Rg1 in proliferative diabetic retinopathy (PDR) are still unclear. In the present study, we aimed to investigate the effects of Rg1 on mesenchymal activation of high-glucose (HG) cultured müller cells. High glucose conditions up-regulate MMP-2, MMP-9 and down-regulate TIMP-2, and promote mesenchymal activation in Müller cells. And Rg1 inhibits the HG-induced mesenchymal activation and HG-increased MMP-2 and MMP-9 and HG-decreased TIMP-2 in Müller cells. HG up-regulates Zeb1 and lncRNA RP11-982M15.8, and down-regulates miR-2113, and Rg1 inhibits these effects of HG. Both inhibition of miR-2113 and over-expression of RP11-982M15.8 significantly restored the HG induced mesenchymal activasion. Taken together, our findings suggested that Rg1 inhibited HG-induced mesenchymal activation and fibrosis via regulating miR-2113/RP11-982M15.8/Zeb1 pathway.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Different molecular mechanisms of HTLV-1 and HIV LTR activation by TPA

HTLV-1 and HIV-1 are retroviruses involved in different human diseases. However, following infection, these viruses inter into a latent state. Tax and Tat are regarded as trans-activators of HTLV-1 and HIV-1 respectively. As it known, during the latent state the infected cells contain low Tax and Tat protein levels, so the activation of these viruses must be independent of these proteins. Here we focus on exploring the mechanism of activation of these viruses by 12-O-tetradecanoylphorbol-13-acetate (TPA), which is a potent activator of protein kinase C (PKC) and considered as a stress-inducing agent. Our results showed that short exposure to TPA considerably stimulated only the HIV-1 LTR expression, while long exposure stimulated only the HTLV-1 LTR and that their activation is agonized or antagonized by PKC respectively. It was found that TPA induced interaction between the transcriptional factors Sp1 and P53 producing Sp1-p53 complex which strongly interacted with c-Jun only after short exposure to TPA. In addition, TPA treatment highly induced the expression of CREB which attached to the Sp1-p53 complex mainly after a long exposure to TPA. A strong binding of sp1, p53 and CREB proteins with HTLV-1 LTR and strong binding of NF-κB with HIV-1 LTR were observed after long (24 h) and short (6 h) exposures to TPA respectively by Chip assay. These results support the possibility that sp1, p53 and CREB are involved in the TPA induced HTLV-1 LTR expression while TPA activation of HIV-1 LTR seems to be dependent on PKC activity through the NF-κB pathway.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

DUSP14 knockout accelerates cardiac ischemia reperfusion (IR) injury through activating NF-κB and MAPKs signaling pathways modulated by ROS generation

Highlights: • DUSP14 expression is decreased in the infarcted area following myocardial I/R. • DUSP14 knockout aggregates cardiac function following myocardial I/R. • DUSP14 ablation intensifies inflammation in murine hearts with IR injury. • DUSP14 deficiency exacerbates MAPKs pathway and oxidative stress in murine hearts. Inflammation and oxidative stress are significantly involved in the progression of a variety of diseases, including myocardial ischemia/reperfusion (IR). In the present study, we hypothesized a protective role of dual-specificity phosphatase 14 (DUSP14) in myocardial IR, as well as the underlying molecular mechanism. The results indicated that DUSP14 was down-regulated following cardiac IR injury. Subsequently, the wild type (WT) and DUSP14-knockout (KO) mice were included to further reveal the potential role of DUSP14 in cardiac IR injury progression. DUSP14-KO mice exhibited increased infarction area and elevated apoptosis, as evidenced by the increased TUNEL-positive cells in ischemia heart following reperfusion compared to WT mice. Further, DUSP14-KO significantly aggregated cardiac dysfunction of mice after IR injury. Cardiac IR injury to DUSP14-KO mice led to markedly increased expression of pro-inflammatory cytokines and activated nuclear factor-κB (NF-κB) pathway in the heart in comparison to WT mice. Meanwhile, mitogen-activated protein kinases (MAPKs), including p38, ERK1/2 and JNK, were significantly activated by DUSO14-KO in mice after IR injury. Compared to WT mice, DUSP14-KO mice showed markedly increased oxidative stress markers in cardiac tissues, including malondialdehyde (MDA), NADPH oxidase-4 (NOX4) and p47, while decreased activities or expressions of anti-oxidants, such as glutathione (GSH), glutathione peroxidase (GPx), glutathion reductases (GR), superoxide dismutase (SOD) and hemeoxygenase-1 (HO-1). DUSP14-knockdown (KD) in primary cardiomyocytes using its specific siRNA sequence elevated hypoxia and reoxygenation (HR)-induced activation of NF-κB and MAPKs signaling pathways, and reactive oxygen species (ROS) generation. Intriguingly, pre-treatment of ROS scavenger, N-acetylcysteine (NAC), markedly abolished DUSP14-KD-augmented NF-κB and MAPKs activation in HR-stimulated primary cardiomyocytes. Together, the results above indicated that DUSP14 might be served as a positive regulator to attenuate cardiac IR injury. Suppressing DUSP14 exacerbated cardiac injury through activating NF-κB and MAPKs signaling pathways regulated by ROS production. Thus, DUSP14 could be a valuable target for developing treatments for myocardial IR injury.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Keratinocyte growth factor protects endometrial cells from oxygen glucose deprivation/re-oxygenation via activating Nrf2 signaling

Highlights: • Keratinocyte growth factor (KGF) receptor (KGFR) is functionally expressed in endometrial cells. • KGF protects endometrial cells from oxygen glucose deprivation/re-oxygenation (OGDR). • KGF inhibits OGDR-induced programmed necrosis in endometrial cells. • KGF activates Nrf2 signaling to protect endometrial cells from OGDR. • Akt-mTOR activation mediates KGF-induced Nrf2 activation and endometrial cell protection. Oxygen and glucose deprivation (OGD)-re-oxygenation (OGDR) exposure to endometrial cells mimics ischemia-reperfusion injury. The present study tests the potential effect of keratinocyte growth factor (KGF) on the process. We show that KGF receptor KGFR is expressed in human endometrial T-HESC cells and primary murine endometrial cells. KGF pre-treatment protected endometrial cells from OGDR, inhibiting cell viability reduction and cell death. KGF attenuated OGDR-induced programmed necrosis in endometrial cells. Significantly, KGF activated Nrf2 signaling, causing Nrf2 Ser-40 phosphorylation, protein stabilization, nuclear translocation to promote anti-oxidant gene (HO1, NOQ1 and GCLC) expression. Nrf2 silencing (by targeted shRNAs) or CRISPR/Cas9 knockout almost abolished KGF-induced endometrial cell protection against OGDR. Furthermore, KGF activated Akt-mTOR signaling in endometrial cells. Whereas Akt-mTOR inhibitors (LY294002, AZD2014 and RAD001) abolished KGF-induced Nrf2 activation and anti-OGDR cytoprotection. Together, KGF protects endometrial cells from OGDR via activating Akt-mTOR-Nrf2 signaling.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

The orphan G protein-coupled receptor 25 (GPR25) is activated by Apelin and Apela in non-mammalian vertebrates

Highlights: • The orphan receptor GPR25 was cloned from zebrafish, pigeons and spotted gars. • GPR25 is activated by Apelin and Apela in non-mammalian vertebrates. • Like Apelin receptor, GPR25 is coupled to Gi-cAMP signaling pathway. • GPR25 mRNA is widely expressed in zebrafish/spotted gar/human tissues. G protein-coupled receptor 25 (GPR25) is an orphan G protein-coupled receptor in vertebrates, that has been implicated to be associated with autoimmune diseases and regulate blood pressure in humans. However, the endogenous ligand of GPR25 remains unknown in vertebrates. Here, we reported that in non-mammalian vertebrates (zebrafish, spotted gars, and pigeons), GPR25 could be activated by Apelin and Apela peptides, which are also the two endogenous ligands of vertebrate Apelin receptor (APLNR). Using the pGL3-CRE-luciferase reporter assay and confocal microscopy, we first demonstrated that like APLNR, zebrafish GPR25 expressing in HEK293 cells could be effectively activated by zebrafish Apelin and Apela peptides, leading to the inhibition of forskolin-stimulated cAMP production and receptor internalization. Like zebrafish GPR25, pigeon and spotted gar GPR25 could also be activated by Apelin and Apela, and their activation could inhibit forskolin-induced cAMP accumulation. Interestingly, unlike zebrafish (/spotted gar/pigeon) GPR25, human GPR25 could not be activated by Apelin and Apela under the same experimental conditions. RNA-seq analysis further revealed that GPR25 is expressed in a variety of tissues, including the testes and intestine of zebrafish/spotted gars/humans, implying the potential roles of GPR25 signaling in many physiological processes in vertebrates. Taken together, our data not only provides the first proof that the orphan receptor GPR25 possesses two potential ligands ‘Apelin and Apela’ and its activation decreases intracellular cAMP levels in non-mammalian vertebrates, but also facilitates to unravel the physiological roles of GPR25 signaling in vertebrates.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

STAT1 activation represses IL-22 gene expression and psoriasis pathogenesis

Highlights: • STAT1 binds to the IL-22 promoter and directly antagonizes STAT3-mediated IL-22 induction. • The balance between activated STAT3 and STAT1 determines IL-22 promoter activities. • Reduced STAT1 expression results in elevated STAT3 activation and IL-22 production. • STAT1 activation represses skin inflammation in an imiquimod-induced murine psoriasis model. IL-22 plays an important role in tissue repair and inflammatory responses, and is implicated in the pathogenesis of psoriasis, ulcerative colitis, as well as liver and pancreas damage. The molecular mechanisms of its regulation have been actively studied. Here, we show that the differential regulation of IL-22 expression in CD4{sup +} T cells by IL-6 and IL-27 was detected rapidly after stimulation. Chromatin immunoprecipitation (ChIP) and luciferase reporter assays demonstrated that both STAT1 and STAT3 directly bind to the STAT responsive elements (SRE) of the IL-22 promoter, and the balance between activated STAT3 and STAT1 determines IL-22 promoter activities. We further show that the heterozygous mutation of the STAT1 gene results in elevated levels of IL-22 production and induces much severer skin inflammation in an imiquimod (IMQ)-induced murine psoriasis model. Together, our results reveal a novel regulatory mechanism of IL-22 expression by STAT1 through directly antagonizing STAT3, and the importance of the balance between STAT3 and STAT1 in IL-22 regulation and psoriasis pathogenesis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

CAR-mediated repression of Cdkn1a(p21) is accompanied by the Akt activation

Highlights: • CAR-mediated liver growth is accompanied by the decrease in PTEN. • CAR-mediated liver growth is accompanied by the Akt activation. • CAR-mediated Akt activation is accompanied by the decrease in Foxo1. • CAR activation decreased the level of p21. It was shown that CAR participates in the regulation of many cell processes. Thus, the activation of CAR causes a proliferating effect in the liver, which provides grounds to consider CAR as a therapeutic target when having a partial resection of this organ. Even though a lot of work has been done on the function of CAR in regulating hepatocyte proliferation, very little has been done on its complex mediating mechanism. This study, therefore, showed that the liver growth resulting from CAR activation leads to the decline in the level of PTEN protein and subsequent Akt activation in mouse liver. The increase of Akt activation produced by CAR agonist was accompanied by a decrease in the level of Foxo1, which was correlated with decreased expression of Foxo1 target genes, including Cdkn1a(p21). Moreover, the study also demonstrated that there exists a negative regulatory impact of CAR on the relationship between Foxo1 and targeted Cdkn1a(p21) promoter. Therefore, the study results revealed an essential function of CAR-Akt-Foxo1 signalling pathway in controlling hepatocyte proliferation by repressing the cell cycle regulator Cdkn1a (p21).

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Dihydromyricetin inhibits caerulin-induced TRAF3-p38 signaling activation and acute pancreatitis response

Highlights: • Dihydromyricetin (DHM) inhibits caerulin-induced cytokine production in BMDMs. • DHM inhibits caerulin-induced TRAF3-MKK3-p38 cascade activation in BMDMs. • DHM attenuates the cytotoxicity of caerulin-activated BMDMs to pancreatic acinar cells. • DHM inhibits pancreatic and systemic inflammation in caerulin-induced AP mice. Acute pancreatitis (AP) is a common inflammatory disease in gastrointestinal tract. Our previous study has shown that caerulin induces TNF receptor-associated factor 3 (TRAF3)-p38 signaling activation and pro-inflammatory response in macrophages, causing damage to co-cultured pancreatic acinar cells. Dihydromyricetin (DHM) is a flavonoid extracted from Ampelopsis grossedentata, which has displayed anti-inflammation and anti-oxidant functions. Our results here show that DHM potently inhibited caerulin-induced expression and productions of multiple pro-inflammatory cytokines (IL-1β, TNF-α and IL-17) in murine bone marrow–derived macrophages (BMDMs). DHM significantly inhibited caerulin-induced TRAF3 protein stabilization, TRAF3-mitogen-activated protein kinase kinase 3 (MKK3) association and following MKK3-p38 activation in BMDMs. Significantly, DHM was ineffective against caerulin in TRAF3-silenced BMDMs. Importantly, DHM supplement attenuated the cytotoxicity of caerulin-activated BMDMs to co-cultured pancreatic acinar cells, resulting in significantly decreased acinar cell death and apoptosis. In vivo, DHM co-administration largely attenuated pancreatic and systemic inflammation in caerulin-injected AP mice. Together, DHM inhibits caerulin-induced TRAF3-p38 signaling activation and AP response. DHM could be further studied as a potential anti-AP agent.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Pull down assay for GTP-bound form of Sar1a reveals its activation during morphological differentiation

Highlights: • The C-terminal region of Sec23B is available for a Sar1a pull down assay. • Neuronal differentiation correlates to an increase in GTP-bound Sar1a. • Oligodendrocyte differentiation correlates to an increase in GTP-bound Sar1a. • PREB binding to nucleotide-free Sar1a also increases during differentiation. The intracellular molecular transport system is a basic and general cellular mechanism that is regulated by an array of signaling molecules. Sar1 small GTPases are molecules that play a key role in controlling vehicle transport between the endoplasmic reticulum (ER) and Golgi bodies. Like other small GTPases, the activities of Sar1a depend on their guanine-nucleotide-binding states, which are regulated by guanine-nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). Despite the well-known function of mammalian Sar1 in the intracellular transport system, little is known about when and how Sar1 is activated during cell morphological changes. Here we show that the C-terminal, but not the N-terminal, regions of Sec23A and Sec23B, the effector proteins of Sar1a, specifically bind to the active, GTP-bound form of Sar1a. An affinity precipitation (pull-down) assay using a recombinant C-terminal region of Sec23B reveals that Sar1a is activated following differentiation in neuronal cell lines. In neuronal N1E-115 cells, GTP-bound Sar1a is increased when cells elongate neuronal processes. Similar results are observed in morphological differentiation in oligodendroglial FBD-102b cells. Additionally, prolactin regulatory element binding (PREB), the GEF for Sar1 (Sar1 activator), increases the binding ability to the nucleotide-free form of Sar1a when morphological differentiation occurs. Nucleotide-free small GTPases preferentially interact with the cognate, active GEFs. These results provide evidence that using previously unreported pull down assays reveals that Sar1 and PREB are upregulated following the induction of morphological differentiation, suggesting the potential role of signaling through Sar1a during morphological differentiation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Resveratrol-induced p53 activation is associated with autophagy in mouse embryonic stem cells

Highlights: • Resveratrol-induced p53 activation does not cause G1/S cell cycle checkpoint and differentiation in mESCs. • The level of resveratrol-induced p53 activity is regulated by SIRT1 deacetylation in mESCs. • Resveratrol-activated p53 stimulates autophagy in a transcription-dependent manner in mESCs. Resveratrol is a natural polyphenol with several therapeutic effects, in particular, inducing p53-dependent cell cycle arrest and/or apoptosis in tumor cells. Resveratrol-induced p53 activation may trigger differentiation and apoptosis in embryonic stem cells (ESCs). We show that resveratrol activates p53 that is negatively regulated by SIRT1 deacetylation on Lys379 and positively by AMPK phosphorylation on Ser15 in mouse ESCs (mESCs). Surprisingly, the resveratrol-activated p53 is not associated with either G1/S cell cycle checkpoint or apoptosis in mESCs. Instead, it stimulates autophagy in a transcriptional-dependent manner involving up-regulation of dram1 gene expression. This study demonstrates a novel mechanism of resveratrol-dependent p53 activation in mESCs.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Sanguisorba officinalis L. extracts activate Wnt/β-catenin pathway, and subsequently control adipo-osteogenic differentiation

Highlights: • Identification of herbal extracts as an activator of Wnt/β-catenin pathway. • Activation of Wnt/β-catenin pathway by Sanguisorba officinalis L. extracts (SOE). • Inhibitory effects of SOE on adipogenesis via Wnt/β-catenin pathway activation. • Stimulating effects of SOE on osteogenesis via Wnt/β-catenin pathway activation. • The effectiveness of SOE for accelerating mice calvaroal bone formation ex vivo. Wnt/β-catenin pathway plays an important role in adipogenesis and osteogenesis. To search for novel activators of the Wnt/β-catenin pathway, we screened plant extracts and found that Sangusorba officinalis L. extracts (SOE) could increase β-catenin expression level and nuclear accumulation in 3T3-L1 and MC3T3-E1 cells. It was confirmed that SOE could effectively control adipo-osteogenic differentiation. SOE inhibited adipocyte differentiation of 3T3-L1 cells, markedly decreased intracellular lipid accumulation, and decreased expression levels of key adipogenic transcription factors including PPARγ, C/EBPα, and SREBP-1c. SOE enhanced ALP activity and terminal osteoblast differentiation which was confirmed by Alizarin Red S staining of MC3T3-E1 cells. In ex vivo culture, SOE significantly increased the thickness of calvarial bone in mice. Taken together, our results indicate that SOE could be used as a potential therapeutic agent for dual-treatment of anti-obesity and anti-osteoporosis via activation of Wnt/β-catenin pathway.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Prevention of acute liver injury by suppressing plasma kallikrein-dependent activation of latent TGF-β

Highlights: • TGF-β contributes to the pathogenesis of acute liver injury (ALI). • PLK-dependent TGF-β activation occurs in/around macrophages in ALI mice. • R58 latency associated protein-degradation products serve as an indicator for ALI. • Inhibiting PLK suppresses apoptosis and improves survival rate of ALI mice. Acute liver injury (ALI) is highly lethal acute liver failure caused by different etiologies. Transforming growth factor β (TGF-β) is a multifunctional cytokine and a well-recognized inducer of apoptotic and necrotic cell death in hepatocytes. Latent TGF-β is activated partly through proteolytic cleavage by a serine protease plasma kallikrein (PLK) between the R58 and L59 residues of its propeptide region. Recently, we developed a specific monoclonal antibody to detect the N-terminal side LAP degradation products ending at residue R58 (R58 LAP-DPs) that reflect PLK-dependent TGF-β activation. This study aimed to explore the potential roles of PLK-dependent TGF-β activation in the pathogenesis of ALI. We established a mouse ALI model via the injection of anti-Fas antibodies (Jo2) and observed increases in the TGF-β1 mRNA level, Smad3 phosphorylation, TUNEL-positive apoptotic hepatocytes and R58-positive cells in the liver tissues of Jo2-treated mice. The R58 LAP-DPs were observed in/around F4/80-positive macrophages, while macrophage depletion with clodronate liposomes partly alleviated the Jo2-induced liver injury. Blocking PLK-dependent TGF-β activation using either the serine proteinase inhibitor FOY305 or the selective PLK inhibitor PKSI-527 or blocking the TGF-β receptor-mediated signaling pathway using SB431542 significantly prevented Jo2-induced hepatic apoptosis and mortality. Furthermore, similar phenomena were observed in the mouse model of ALI with the administration of acetaminophen (APAP). In summary, R58 LAP-DPs reflecting PLK-dependent TGF-β activation may serve as a biomarker for ALI, and targeting PLK-dependent TGF-β activation has potential as a therapeutic strategy for ALI.

60 APPLIED LIFE SCIENCES↗