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At least 613 records · Page 34

Orion Heat Shield Manufacturing Producibility Improvements for the EM-1 Flight Test Program

This paper describes how the Orion program is incorporating improvements in the heat shield design and manufacturing processes reducing programmatic risk and ensuring crew safety in support of NASA’s Exploration Missions. The Orion program successfully completed the EFT-1 flight test in 2014 and is currently developing the EM-1 spacecraft to meet the test objectives of an orbital mission to the moon and return to earth in 2019. Lessons learned from the EFT-1 manufacturing and flight test experience are being incorporated into a wide variety of vehicle systems and manufacturing processes to reduce risk to the Orion missions and flight crew. A critical contributor to crew safety is the heat shield that protects the crew capsule during re-entry through the earth’s atmosphere for return from deep space. The first flight test vehicle, EFT-1, was manufactured and tested in the Neil Armstrong Operations and Checkout (O&C) facility at KSC to demonstrate early risk reduction including the functionality of the Thermal Protection Systems (TPS) for capsule safe return to earth. The approach for the EFT-1 heat shield utilized a low risk approach using Apollo heritage design and manufacturing processes using an Avcoat TPS ablator with a honeycomb substrate to provide a one piece heat shield to meet the mission re-entry heating environments. The manufacturing processes used honeycomb cell injection guns which were redeveloped from the Apollo Lunar Program processes to build the EFT-1 heat shield. The completed heat shield was transported across the country by aircraft to the O&C at KSC for installation onto the capsule. The EFT-1 heat shield successfully performed its mission and experienced ~80% of the re-entry velocity (50% heating rate) for a lunar return for an Exploration Mission. The second flight test vehicle is the EM-1 mission which will have additional flight systems installed to fly to the moon and return. Heat shield design and producibility improvements have been incorporated in the EM-1 vehicle to meet deep space mission and programmatic requirements. The design continues to use the Avcoat material, but in a “block” configuration to enable improvements in the application processes as well as additional improvements in the carrier structure design and manufacturing operations. Incorporating flight test results and producibility improvements from EFT-1 for the heat shield system design and processes have improved the thermal protection capability, improved the producibility, and cost for EM-1 flight test.

Producibility↗

Shear stress reduces protease activated receptor-1 expression in human endothelial cells

Shear stress has been shown to regulate several genes involved in the thrombotic and proliferative functions of endothelial cells. Thrombin receptor (protease-activated receptor-1: PAR-1) increases at sites of vascular injury, which suggests an important role for PAR-1 in vascular diseases. However, the effect of shear stress on PAR-1 expression has not been previously studied. This work investigates effects of shear stress on PAR-1 gene expression in both human umbilical vein endothelial cells (HUVECs) and microvascular endothelial cells (HMECs). Cells were exposed to different shear stresses using a parallel plate flow system. Northern blot and flow cytometry analysis showed that shear stress down-regulated PAR-1 messenger RNA (mRNA) and protein levels in both HUVECs and HMECs but with different thresholds. Furthermore, shear-reduced PAR-1 mRNA was due to a decrease of transcription rate, not increased mRNA degradation. Postshear stress release of endothelin-1 in response to thrombin was reduced in HUVECs and HMECs. Moreover, inhibitors of potential signaling pathways applied during shear stress indicated mediation of the shear-decreased PAR-1 expression by protein kinases. In conclusion, shear stress exposure reduces PAR-1 gene expression in HMECs and HUVECs through a mechanism dependent in part on protein kinases, leading to altered endothelial cell functional responses to thrombin.

Non-NASA Center↗

Purification and Characterization of [NiFe]-Hydrogenase of Shewanella oneidensis MR-1

The γ-proteobacterium Shewanella oneidensis MR-1 possesses a periplasmic [NiFe]-hydrogenase (MR-1 [NiFe]-H2ase) that was implicated in both H2 production and oxidation as well as technetium [Tc(VII)] reduction. To characterize the roles of MR-1 [NiFe]-H2ase in these proposed reactions, the genes encoding both subunits of MR-1 [NiFe]-H2ase were cloned into a protein expression vector. The resulting plasmid was transformed into a MR-1 mutant deficient in H2 formation. Expression of MR-1 [NiFe]-H2ase in trans restored the mutant’s ability to produce H2 at 37% of that for wild type. Following expression, MR-1 [NiFe]-H2ase was purified to near homogeneity. The purified MR-1 [NiFe]-H2ase could couple H2 oxidation to reduction of Tc(VII) and methyl viologen directly. Change of the buffers used affected MR-1 [NiFe]-H2ase-mediated Tc(VII) but not methyl viologen reductions. Under the conditions tested, Tc(VII) reduction was complete in Tris buffer but not in HEPES buffer. The reduced Tc(IV) was soluble in Tris buffer but insoluble in HEPES buffer. Transmission electron microscopy analysis revealed that Tc(IV) precipitates formed in HEPES buffer were packed with crystallites. Although X-ray absorption near-edge spectroscopy measurements confirmed that the reduction products found in both buffers were Tc(IV), extended X-ray adsorption fine-structure measurements revealed that these products were very different. While the product in Tris buffer could not be determined, the Tc(IV) product in HEPES buffer was very similar to Tc(IV)O2•nH2O. These results shows for the first time that MR-1 [NiFe]-H2ase is a bidirectional enzyme that catalyzes both H2 formation and oxidation as well as Tc(VII) reduction directly by coupling H2 oxidation.

59 BASIC BIOLOGICAL SCIENCES↗

Transcription of the Tollip gene is elevated in intestinal epithelial cells through impaired O-GlcNAcylation-dependent nuclear translocation of the negative regulator Elf-1

Highlights: {yields} Transcriptional activation of the Tollitip gene is higher in IECs than in monocytes. {yields} Nt -194/-186 region acts as a cis-element and is recognized by Elf-1. {yields} Elf-1 suppresses Tollip gene transcription in monocytes but not in IECs. {yields} O-GlcNAc modification is necessary for nuclear translocation of Elf-1. {yields} O-GlcNAcylation-dependent nuclear translocation of Elf-1 is impaired in IECs. -- Abstract: Intestinal epithelial cells (IECs) must be tolerant of the large number of commensal bacteria inhabiting the intestinal tract to avoid excessive inflammatory reactions. Toll-interacting protein (Tollip), a negative regulator of Toll-like receptor signaling, is known to be expressed at high levels in IECs, and to thereby contribute to the hyporesponsiveness of IECs to commensals. In this study, we analyzed the underlying mechanisms for elevated transcription of the Tollip gene in IECs using a human IEC line, Caco-2, and a human monocyte line, THP-1, as a control. Elf-1 was identified as a transcription factor that negatively regulates Tollip gene expression. The transcription factor Elf-1 was localized in the nucleus by O-linked N-acetylglucosamine (O-GlcNAc) modification, whereas the unmodified form was detected only in the cytoplasm. Comparison of Caco-2 and THP-1 cells revealed that O-GlcNAc modification of Elf-1 was significantly lower in IECs than in monocytes. Collectively, the results indicate that insufficient O-GlcNAc modification prevents Elf-1-mediated transcriptional repression and thereby upregulates Tollip gene expression in IECs.

60 APPLIED LIFE SCIENCES↗

Green tea polyphenol epigallocatechin-3-gallate suppresses melanoma growth by inhibiting inflammasome and IL-1{beta} secretion

Highlights: Black-Right-Pointing-Pointer EGCG inhibits melanoma cell growth at physiological doses (0.1-1 {mu}M). Black-Right-Pointing-Pointer EGCG inhibits melanoma cell growth via inflammasomes and IL-1{beta} suppression. Black-Right-Pointing-Pointer Inflammasomes and IL-1{beta} could be potential targets for future melanoma therapeutics. -- Abstract: Epigallocatechin-3-gallate (EGCG), the major polyphenolic component of green tea, has been demonstrated to possess anti-inflammatory, antioxidant, anti-mutagenic and anti-carcinogenic properties. The anti-melanoma effect of EGCG has been previously suggested, but no clear mechanism of action has been established. In this study, we demonstrated that EGCG inhibits melanoma cell growth at physiological doses (0.1-1 {mu}M). In the search for mechanisms of EGCG-mediated melanoma cell suppression, we found that NF-{kappa}B was inhibited, and that reduced NF-{kappa}B activity was associated with decreased IL-1{beta} secretion from melanoma cells. Since inflammasomes are involved in IL-1{beta} secretion, we investigated whether IL-1{beta} suppression was mediated by inflammasomes, and found that EGCG treatment led to downregulation of the inflammasome component, NLRP1, and reduced caspase-1 activation. Furthermore, silencing the expression of NLRP1 abolished EGCG-induced inhibition of tumor cell proliferation both in vitro and in vivo, suggesting a key role of inflammasomes in EGCG efficacy. This paper provides a novel mechanism for EGCG-induced melanoma inhibition: inflammasome downregulation {yields} decreased IL-1{beta} secretion {yields} decreased NF-{kappa}B activities {yields} decreased cell growth. In addition, it suggests inflammasomes and IL-1{beta} could be potential targets for future melanoma therapeutics.

60 APPLIED LIFE SCIENCES↗

The PDZ protein TIP-1 facilitates cell migration and pulmonary metastasis of human invasive breast cancer cells in athymic mice

Highlights: Black-Right-Pointing-Pointer This study has revealed novel oncogenic functions of TIP-1 in human invasive breast cancer. Black-Right-Pointing-Pointer Elevated TIP-1 expression levels in human breast cancers correlate to the disease prognosis. Black-Right-Pointing-Pointer TIP-1 knockdown suppressed the cell migration and pulmonary metastasis of human breast cancer cells. Black-Right-Pointing-Pointer TIP-1 knockdown suppressed the expression and functionality of motility-related genes. -- Abstract: Tax-interacting protein 1 (TIP-1, also known as Tax1bp3) inhibited proliferation of colon cancer cells through antagonizing the transcriptional activity of beta-catenin. However, in this study, elevated TIP-1 expression levels were detected in human invasive breast cancers. Studies with two human invasive breast cancer cell lines indicated that RNAi-mediated TIP-1 knockdown suppressed the cell adhesion, proliferation, migration and invasion in vitro, and inhibited tumor growth in mammary fat pads and pulmonary metastasis in athymic mice. Biochemical studies showed that TIP-1 knockdown had moderate and differential effects on the beta-catenin-regulated gene expression, but remarkably down regulated the genes for cell adhesion and motility in breast cancer cells. The decreased expression of integrins and paxillin was accompanied with reduced cell adhesion and focal adhesion formation on fibronectin-coated surface. In conclusion, this study revealed a novel oncogenic function of TIP-1 suggesting that TIP-1 holds potential as a prognostic biomarker and a therapeutic target in the treatment of human invasive breast cancers.

60 APPLIED LIFE SCIENCES↗

Effects of matrix metalloproteinase-1 on the myogenic differentiation of bone marrow-derived mesenchymal stem cells in vitro

Highlights: Black-Right-Pointing-Pointer MMP-1 is a member of the zinc-dependent endopeptidase family. Black-Right-Pointing-Pointer MMP-1 has no cytotoxic effects on BMSCs. Black-Right-Pointing-Pointer MMP-1 can promote the myogenic differentiation of BMSCs. Black-Right-Pointing-Pointer MyoD and desmin were chosen as myogenic markers in this study. -- Abstract: Matrix metalloproteinase-1 (MMP-1) is a member of the family of zinc-dependent endopeptidases that are capable of degrading extracellular matrix (ECM) and certain non-matrix proteins. It has been shown that MMP-1 can enhance muscle regeneration by improving the differentiation and migration of myoblasts. However, it is still not known whether MMP-1 can promote the myogenesis of bone marrow-derived mesenchymal stem cells (BMSCs). To address this question, we isolated BMSCs from C57BL/6J mice and investigated the effects of MMP-1 on their proliferation and myogenic differentiation. Our results showed that MMP-1 treatment, which had no cytotoxic effects on BMSCs, increased the mRNA and protein levels of MyoD and desmin in a dose-dependent manner, indicating that MMP-1 promoted myogenic differentiation of BMSCs in vitro. These results suggest that BMSCs may have a therapeutic potential for treating muscular disorders.

60 APPLIED LIFE SCIENCES↗

EDF-1 downregulates the CaM/Cn/NFAT signaling pathway during adipogenesis

Highlights: ► EDF-1 participates early adipogenesis in 3T3F442A cells induced with Staurosporine/Dexamethasone. ► EDF-1 associates with CaM and Cn, most likely inactivating Cn. ► EDF-1/CaM complex seems to prevent NFATc1 activation by Cn. ► EDF-1 regulates the Cn/CaM/NFATc1 pathway during adipogenesis. ► EDF-1 may regulate the activation of Cn through a complex formation with CaM. - Abstract: The endothelial differentiation factor-1 (EDF-1) is a calmodulin binding protein that regulates calmodulin-dependent enzymes. In endothelial cells, this factor can form a protein complex with calmodulin. We analyzed the relationship between this factor and the members of calmodulin/calcineurin/nuclear factor of activated T-cells (NFAT) signaling pathway during adipogenesis of 3T3-F442A cells. We found that the expression of edf1 is upregulated during early adipogenesis, whereas that of calcineurin gene is lowered, suggesting that this pathway should be downregulated to allow for adipogenesis to occur. We also found that EDF-1 associates with calmodulin and calcineurin, most likely inactivating calcineurin. Our results showed that EDF-1 inactivates the calmodulin/calcineurin/NFAT pathway via sequestration of calmodulin, during early adipogenesis, and we propose a mechanism that negatively regulates the activation of calcineurin through a complex formation between EDF-1 and calmodulin. This finding raises the possibility that modulating this pathway might offer some alternatives to regulate adipose biology.

60 APPLIED LIFE SCIENCES↗

Impact of Mesenchymal Stem Cell secreted PAI-1 on colon cancer cell migration and proliferation

Highlights: •MSCs were directly co-cultured with colorectal cancer (CRC) cells on 3D scaffolds. •MSCs influence CRC protein/gene expression, proliferation and migration. •We report a significant functional role of MSC-secreted PAI-1 in colon cancer. -- Abstract: Mesenchymal Stem Cells are known to engraft and integrate into the architecture of colorectal tumours, with little known regarding their fate following engraftment. This study aimed to investigate mediators of Mesenchymal Stem Cell (MSC) and colon cancer cell (CCC) interactions. Mesenchymal Stem Cells and colon cancer cells (HT29 and HCT-116) were cultured individually or in co-culture on 3-dimensional scaffolds. Conditioned media containing all secreted factors was harvested at day 1, 3 and 7. Chemokine secretion and expression were analyzed by Chemi-array, ELISA (Macrophage migration inhibitory factor (MIF), plasminogen activator inhibitor type 1 (PAI-1)) and RQ-PCR. Colon cancer cell migration and proliferation in response to recombinant PAI-1, MSCs and MSCs + antibody to PAI-1 was analyzed using Transwell inserts and an MTS proliferation assay respectively. Chemi-array revealed secretion of a wide range of factors by each cell population, including PAI-1and MIF. ELISA analysis revealed Mesenchymal Stem Cells to secrete the highest levels of PAI-1 (MSC mean 10.6 ng/mL, CCC mean 1.01 ng/mL), while colon cancer cells were the principal source of MIF. MSC-secreted PAI-1 stimulated significant migration of both CCC lines, with an antibody to the chemokine shown to block this effect (67–88% blocking,). A cell-line dependant effect on CCC proliferation was shown for Mesenchymal Stem Cell-secreted PAI-1 with HCT-116 cells showing decreased proliferation at all concentrations, and HT29 cells showing increased proliferation in the presence of higher PAI-1 levels. This is the first study to identify PAI-1 as an important mediator of Mesenchymal Stem Cell/colon cancer cell interactions and highlights the significant functional impact of Mesenchymal Stem Cell-secreted PAI-1 on colon cancer cells.

60 APPLIED LIFE SCIENCES↗

Design, synthesis and evaluation of a potent substrate analog inhibitor identified by scanning Ala/Phe mutagenesis, mimicking substrate co-evolution, against multidrug-resistant HIV-1 protease

Highlights: •Inhibitors against MDR HIV-1 protease were designed, synthesized and evaluated. •Lead peptide (6a) showed potent inhibition (IC{sub 50}: 4.4 nM) of MDR HIV-1 protease. •(6a) Showed favorable binding isotherms against NL4-3 and MDR proteases. •(6a) Induced perturbations in the {sup 15}N-HSQC spectrum of MDR HIV-1 protease. •Molecular modeling suggested that (6a) may induce total flap closure inMDR protease. -- Abstract: Multidrug-resistant (MDR) clinical isolate-769, human immunodeficiency virus type-1 (HIV-1) protease (PDB ID: (1TW7)), was shown to exhibit wide-open flaps and an expanded active site cavity, causing loss of contacts with protease inhibitors. In the current study, the expanded active site cavity of MDR769 HIV-1 protease was screened with a series of peptide-inhibitors that were designed to mimic the natural substrate cleavage site, capsid/p2. Scanning Ala/Phe chemical mutagenesis approach was incorporated into the design of the peptide series to mimic the substrate co-evolution. Among the peptides synthesized and evaluated, a lead peptide (6a) with potent activity (IC{sub 50}: 4.4 nM) was identified against the MDR769 HIV-1 protease. Isothermal titration calorimetry data showed favorable binding profile for 6aagainst both wild type and MDR769 HIV-1 protease variants. Nuclear magnetic resonance spectrum of {sup 15}N-labeled MDR769 HIV-1 protease in complex with 6a showed some major perturbations in chemical shift, supporting the peptide induced conformational changes in protease. Modeling analysis revealed multiple contacts between 6a and MDR769 HIV-1 protease. The lead peptide-inhibitor, 6a, with high potency and good binding profile can be used as the basis for developing potent small molecule inhibitors against MDR variants of HIV.

60 APPLIED LIFE SCIENCES↗

Suppression of Wnt signaling by the miR-29 family is mediated by demethylation of WIF-1 in non-small-cell lung cancer

Highlights: •Dnmt3A and Dnmt3B are involved in the down-regulation of WIF-1 expression in non-small-cell lung cancer. •MiR-29 family members could restore WIF-1 expression through demethylation. •MiR-29s suppress Wnt/β-catenin signaling pathway and inhibit tumor growth. •The expression of miR-29a and miR-29b could be regulated partially in a positive feedback loop. -- Abstract: Wnt inhibitory factor-1 (WIF-1) silencing induced by promoter hypermethylation is a common mechanism of aberrant activation of the Wnt signaling pathway in non-small-cell lung cancer (NSCLC). However, the activity of regulators associated with the methylation of the WIF-1 gene remains unclear. Here, we investigated the role of three DNA methyltransferases (DNMT1, DNMT3A and DNMT3B) in the expression of WIF-1. The three DNMTs were up-regulated in NSCLC tumor tissues and suppression of DNMT3A and DNMT3B restored the expression of WIF-1 in NSCLC cells. The miR-29 family (miR-29a, -29b, and -29c), which negatively regulates DNMT3A and DNMT3B, was examined in association with the Wnt/β-catenin signaling pathway. A positive correlation between the expression of WIF-1 and that of MiR-29s was observed in NSCLC tissues. Methylation-specific PCR and Western blotting indicated that miR-29s positively regulate WIF-1 expression by inhibiting the methylation of its promoter. Furthermore, miR-29 overexpression downregulated β-catenin expression, inhibited cell proliferation and induced apoptosis. The expression of miR-29a and miR-29b was partially regulated by DNMT3A and DNMT3B in a positive feedback loop. Taken together, our findings show that miR-29s suppress the Wnt signaling pathway through demethylation of WIF-1 in NSCLC.

60 APPLIED LIFE SCIENCES↗

Assessment of DNA synthesis in Islet-1{sup +} cells in the adult murine heart

Highlights: • Islet-1 was expressed in the adult heart. • Islet-1-positive cells did not proliferate in the adult heart. • Sinoatrial node cells did not proliferate in the adult heart. - Abstract: Rationale: Islet-1 positive (Islet-1{sup +}) cardiac progenitor cells give rise to the right ventricle, atria and outflow tract during murine cardiac development. In the adult heart Islet-1 expression is limited to parasympathetic neurons, few cardiomyocytes, smooth muscle cells, within the proximal aorta and pulmonary artery and sinoatrial node cells. Its role in these cells is unknown. Here we tested the hypothesis that Islet-1{sup +} cells retain proliferative activity and may therefore play a role in regenerating specialized regions in the heart. Methods and results: DNA synthesis was analyzed by the incorporation of tritiated thymidine ({sup 3}H-thymidine) in Isl-1-nLacZ mice, a transgenic model with an insertion of a nuclear beta-galactosidase in the Islet-1 locus. Mice received daily injections of {sup 3}H-thymidine for 5 days. DNA synthesis was visualized throughout the heart by dipping autoradiography of cryosections. Colocalization of an nLacZ-signal and silver grains would indicate DNA synthesis in Islet-1{sup +} cells. Whereas Islet{sup −} non-myocyte nuclei were regularly marked by accumulation of silver grains, colocalization with nLacZ-signals was not detected in >25,000 cells analyzed. Conclusions: Islet-1{sup +} cells are quiescent in the adult heart, suggesting that, under normal conditions, even pacemaking cells do not proliferate at higher rates than normal cardiac myocytes.

60 APPLIED LIFE SCIENCES↗

Pancreatic adenocarcinoma upregulated factor (PAUF) confers resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNA receptor-mediated signaling

Pancreatic adenocarcinoma upregulated factor (PAUF), a novel oncogene, plays a crucial role in the development of pancreatic cancer, including its metastasis and proliferation. Therefore, PAUF-expressing pancreatic cancer cells could be important targets for oncolytic virus-mediated treatment. Panc-1 cells expressing PAUF (Panc-PAUF) showed relative resistance to parvovirus H-1 infection compared with Panc-1 cells expressing an empty vector (Panc-Vec). Of interest, expression of type I IFN-α receptor (IFNAR) was higher in Panc-PAUF cells than in Panc-Vec cells. Increased expression of IFNAR in turn increased the activation of Stat1 and Tyk2 in Panc-PAUF cells compared with that in Panc-Vec cells. Suppression of Tyk2 and Stat1, which are important downstream molecules for IFN-α signaling, sensitized pancreatic cancer cells to parvovirus H-1-mediated apoptosis. Further, constitutive suppression of PAUF sensitized Bxpc3 pancreatic cancer cells to parvovirus H-1 infection. Taken together, these results suggested that PAUF conferred resistance to pancreatic cancer cells against oncolytic parvovirus H-1 infection through IFNAR-mediated signaling. - Highlights: • PAUF confers resistance against oncolytic parvovirus H-1 infection. • PAUF enhances the expression of IFNAR in Panc-1 cells. • Increased activation of Tyk2 or Stat1 by PAUF provides resistance to parvovirus H-1-mediated apoptosis. • Constitutive inhibition of PAUF enhances parvovirus H-1-mediated oncolysis of Bxpc3 pancreatic cancer cells.

60 APPLIED LIFE SCIENCES↗

Androgen-androgen receptor system improves chronic inflammatory conditions by suppressing monocyte chemoattractant protein-1 gene expression in adipocytes via transcriptional regulation

Age-related decreases in sex hormones are closely related to chronic inflammation in obesity and metabolic diseases. Particularly, the molecular basis of androgen activity in regulating inflammation and controlling metabolism remains largely unknown. Obese adipocytes secrete monocyte chemoattractant protein-1 (MCP-1), a key chemokine that promotes the infiltration of monocytes/macrophages into adipose tissue, thereby leading to metabolic disorders. Here, we studied the role of androgen-androgen receptor (AR) action in regulating MCP-1 expression in adipose tissue. We observed the induction of Mcp-1 expression in 3T3-L1 adipocytes co-cultured with RAW264.7 macrophages. Additionally, Mcp-1 expression was upregulated by culturing in conditioned medium derived from inflammatory macrophages (M1-Mφ) containing tumor necrosis factor-alpha (TNF-α). We found that sex hormones downregulated TNF-α-induced Mcp-1 and interleukin (Il)-6 expression in 3T3-L1 adipocytes. Furthermore, luciferase-reporter analysis indicated that MCP-1 promoter activity was predominantly suppressed by dihydrotestosterone (DHT)-AR interactions through functional canonical nuclear factor-kappa B (NF-κB) sites, whereas non-canonical NF-κB site containing important flanking sequences exhibited minor contributions to DHT-AR transcriptional repression. These findings suggested that androgen-AR suppressed obesity-induced chronic inflammation in adipose tissue. - Highlights: • DHT, non-aromatizable androgen suppresses Mcp-1 expression in adipocytes. • Mcp-1 transcription was negatively regulated by DHT-AR action. • DHT-AR selectively regulates Mcp-1 transcription through distinct NF-κB sites.

60 APPLIED LIFE SCIENCES↗

PART-1 functions as a competitive endogenous RNA for promoting tumor progression by sponging miR-143 in colorectal cancer

LncRNAs were altered in several cancers and played a crucial role in various biological activities and progressions of different diseases, including proliferation, chemical resistance, and metastasis. In the present study, we revealed that prostrate androgen-regulated transcript-1 (PART-1) was highly expressed in colorectal cancer cells and tissues, and knockdown of PART-1 suppressed cell proliferation and metastasis, both in vitro and in vivo. In addition, PART-1 functioned as a ceRNA of DNMT3A, by sponging miR-143. Finally,PART-1 induced tumor progression by regulating DNMT3A. - Highlights: • Prostrate androgen-regulated transcript-1 (PART-1) was highly expressed in colorectal cancer cells and tissues. • Knockdown of PART-1 suppressed cell proliferation and metastasis, both in vitro and in vivo. • PART-1 functioned as a ceRNA of DNMT3A, by sponging miR-143. • PART-1 induced tumor progression by regulating DNMT3A.

60 APPLIED LIFE SCIENCES↗

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↗

Particulate matter increases beta-amyloid and activated glial cells in hippocampal tissues of transgenic Alzheimer's mouse: Involvement of PARP-1

Highlights: • Particulate matter (PM) activates PARP-1 in neuronal cells. • PM increases beta-amyloid in ex vivo hippocampus of 3xTg-AD mouse. • PM activates glial cells in ex vivo hippocampus of 3xTg-AD mouse. • Pharmacological inhibition of PARP-1 reverses PM-induced AD pathologies. • PARP-1 inhibition would be a promising approach counteracting PM's neurotoxicity. Exposure to air pollutants, such as particulate matter (PM), has been implicated in neurodegenerative disorders including Alzheimer's disease (AD). However, direct effects of PM on production of β-amyloid (Aβ), a key pathogenic molecule in AD, and its underlying mechanism are still elusive. Given PM's potential to induce oxidative stress in other tissues, we hypothesized that poly(ADP-ribose) polymerase (PARP-1) might be involved in PM-induced neurotoxicity. To address this, we used an ex vivo model of AD, the organotypic hippocampal slice tissue culture from old (12-14 months-of-age) triple transgenic 3xTg-AD mice. First, we observed that fine PM (aerodynamic diameter < 4 μm) can dose-dependently activate PARP-1 and decrease NAD{sup +} levels in Neuro2A cells. PARP-1 activation did occur under concentrations of PM which did not affect cell viability. Next, we observed that direct treatment of PM increased Aβ levels and activated glial cells in the ex vivo hippocampal tissues of 3xTg-AD mice. PM-induced glial activation was most prominent in CA1 region of the hippocampal tissue. Notably, we found that pharmacological inhibition of PARP-1 reversed both PM-induced Aβ increase and glial activation, arguing the possible involvement of PARP-1 in PM-induced AD pathogenesis. Our findings suggest that PARP-1 might be a potential molecular target, responsible for mediating negative effects of PM on the brain. Modulating PARP-1 activity could be a promising approach to prevent or alleviate PM-related environmental neurotoxicity which could initiate AD pathogenesis.

60 APPLIED LIFE SCIENCES↗

Amelioration of UVB-induced oxidative stress and inflammation in fat-1 transgenic mouse skin

ω-3 polyunsaturated fatty acids (PUFAs), which are abundant in fish oils, are known to scavenge lipid peroxyl radicals and potentiate host immune defence. As UVB-induced oxidative stress and inflammation have been implicated in apoptotic cell death, this study was aimed to investigate the anti-inflammatory, anti-oxidative, and anti-apoptotic capacity of fat-1 transgenic mice capable of converting ω-6 to ω-3 PUFAs. Wild-type (WT) C57BL/6 mice and fat-1 mice were maintained on the AIN-93 diet supplemented with 10% safflower oil rich in ω-6 PUFAs for 5 weeks. The ω-3/ω-6 PUFA ratio was significantly higher in the dorsal skin of fat-1 mice than that in the WT mice. Upon single exposure to UVB (5.0 kJ/m{sup 2}) radiation, fat-1 mice showed inflammatory as well as oxidative tissue damage and the expression of pro-inflammatory enzymes, cyclooxygenases-2 and inducible nitric oxide synthase in the skin to a much lesser extent than the WT mice. The protection of fat-1 mice from UVB-induced skin inflammation was associated with decreased phosphorylation of STAT3. Moreover, UVB-induced apoptosis was attenuated in fat-1 mouse skin. In comparison to WT animals, higher levels of Nrf2 and its target proteins, such as heme oxygenase-1, NAD(P)H:quinone oxidoreductase-1 and thioredoxin-1, were found in the skin of fat-1 mice.

60 APPLIED LIFE SCIENCES↗