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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 1,027 records · Page 57
All sourcesNASA NTRSDOE OSTI
DOE OSTI2017

Highly expressed lncRNA CCND2-AS1 promotes glioma cell proliferation through Wnt/β-catenin signaling

Glioma is the most common and aggressive primary brain tumor in adults. Long-non coding RNAs (lncRNAs) have been recently shown to play important roles in regulating numerous biological processes both in physiologic and pathologic condition. However, the role of lncRNAs in glioma remains largely unknown. In this study, we firstly found that lncRNA CCND2-AS2 is significantly up regulated in malignant glioma tissues and cell lines. Both loss- and gain-functions assays show that CCND2-AS1 promotes glioma cells proliferation and growth. In addition, we also revealed that highly expressed CCND2-AS1 could enhance Wnt/β-catenin signaling in glioma. Taken together, our findings revealed a novel lncRNA CCND2-AS1 promotes glioma cell proliferation through Wnt/β-catenin signaling and CCND2-AS1 might function as a potential novel therapeutic target for the treatment of glioma. - Highlights: • LncRNA CCND2-AS1 was significantly upregulated in glioma cancer tissues and cell lines. • Overexpression CCND2-AS1 confers a poor prognosis in glioma patients. • Knockdown of CCND2-AS1 inhibited glioma cell proliferation and induced cell cycle arrest. • Overexpression CCND2-AS1 promotes cell growth in Normal human astrocyte (NHA). • LncRNA CCND2-AS1 regulates the activation of Wnt/β-catenin signaling in glioma cells.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Retinal expression and localization of Mef2c support its important role in photoreceptor gene expression

Photoreceptor-specific gene expression is controlled by a hierarchical network of transcription factors, including the master regulators cone-rod homeobox (Crx) and neural retina leucine zipper (Nrl). Myocyte-enhancer factor 2c (Mef2c) is an ubiquitously expressed transcription factor with important functions in the cardiovascular system. Here, we performed a detailed analysis of Mef2c expression, localization and function in the retina to further elucidate its potential role for photoreceptor gene regulation. We showed that murine retinal Mef2c mRNA expression was high at birth and peaked at late postnatal developmental stages. Using immunohistochemistry and Western blot, Mef2c protein was detected in the outer nuclear layer of adult mouse and human retinas and localized to the nucleus of 661W photoreceptor-like cells. Mef2c knock-down in 661W cells reduced the expression of arrestin 3 (Arr3) and medium-wave-sensitive cone opsin (Opn1mw) but increased transcript levels of mitogen-activated protein kinase 15 (Mapk15) and phosphodiesterase 6h (Pde6h). In conclusion, Mef2c is highly expressed in the retina where it modulates photoreceptor-specific gene expression. - Highlights: • Mef2c is highly expressed in the mouse and human retina. • Mef2c localizes to mouse and human photoreceptor nuclei in situ. • Mef2c is present in the nucleus of 661W photoreceptor-like cells. • Mef2c knock-down in 661W cells impairs photoreceptor gene expression.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Berberine protects against ischemia/reperfusion injury after orthotopic liver transplantation via activating Sirt1/FoxO3α induced autophagy

The effects and mechanism of berberine (BBR) on hepatic injury after orthotopic liver transplantation (OLT) have not been well characterized. We examined the role of Sirt1/FoxO3α axis in the protective effect of BBR on ischemia/reperfusion injury after OLT. Adult male Wistar rats were randomly allocated into four groups: Sham, OLT, OLT with BBR pretreatment (BBR), OLT with BBR and Sirt1 inhibitor (EX527) pretreatment group (EX527). The liver function and oxidative stress level were measured by biochemical and histopathologic examinations. The formation of autophagosome was observed by transmission electron microscopy. The apoptotic rate was determined by TUNEL analysis and the apoptotic mRNA expression. The expression of Sirt1, FoxO3α, Beclin-1, LC3-II/LC3-I, p62 and the acetylation of FoxO3α were assayed by western blot assay and immunoprecipitation. Compared with the OLT group, BBR dramatically attenuated the histopathologic damage, restored the liver function, and decreased the oxidative stress level. Simultaneously, BBR significantly ameliorated apoptosis by decreasing the apoptotic rate and the expression of apoptotic mRNA in rats subjected to OLT. The level of Beclin-1 and LC3-II/LC3-I were upregulated with the inhibition of p62. The deacetylation of FoxO3α by Sirt1 was enhanced in the nuclear of liver after pretreated with BBR. However, the inhibition of Sirt1 by EX527 counteracted the protective effects of BBR. Thus, BBR preconditioning promotes liver transplant ischemia/reperfusion injury partly via activating Sirt1/FoxO3α mediated autophagy. - Highlights: • Berberine alleviated the histopathologic and functional damage of transplant liver. • Liver transplantation inhibited the acetylation and activation of FoxO3α which was responsible for motivating the autophagic machinery. • Berberine pretreatment reduced apoptotic cell death by enhancing Sirt1/FoxO3α induced autophagy.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Vaccination efficacy with marrow mesenchymal stem cell against cancer was enhanced under simulated microgravity

Stem cell vaccination can induce consistent and strong anti-tumor immunity against cancer in mice model. The antigenic similarity between tumors and embryos has been appreciated for many years and reflects the expression of embryonic gene products by cancer cells and/or cancer-initiating stem cells. Taking advantage of this similarity, we have tested a prophylactic lung cancer vaccine composed of allogeneic murine MSCs. Based on this conception, we first compared their tumor vaccines intervention effects of adult MSCs and MSCs under simulated microgravity (MSC/SMG). In this study, BALB/c mice were vaccinated with MSCs or MSC/SMG, compared with mice vaccinated with phosphate buffered saline (PBS) as negative controls. We then subcutaneously implanted the A549 human lung cancer cell line into vaccinated mice and monitored tumor growth potential in vivo. The smaller tumor size and less tumor weight were observed in mice vaccinated with MSCs or MSC/SMG, compared with that of the Control group. Particularly, it was much more significant in the group of MSC/SMG than that group of the MSCs. Vaccination with SMG treated MSCs inhibited proliferation and promoted apoptosis of tumor tissue. SMG/MSC vaccination induced bothTh1-mediated cytokine response; CD8-dependent cytotoxic response which reduced the proportion of Treg cells. Furthermore, SMG/MSC vaccination significantly increased MHC1 and HSPs proteins expression. In conclusion, we demonstrated the SMG could improve tumor-suppressive activity of MSC. The enhanced anti-tumor immune response of MSCs/SMG was strongly associated with the higher expression of MHC class I molecule on DCs, and the abundance of HSPs in the SMG treated MSCs may make antigens in the MSC more cross-presentable to the host DCs for generating protective antitumor activity. This study gains an insight into the mechanism of MSCs anti-tumor efficacy and gives a new strategy for cancer therapies in the future. - Highlights: • Vaccination with SMG treated MSCs inhibited tumor growth. • SMG/MSC vaccination induced Th1-mediated cytokine response, elicited CD8-dependent cytotoxic response and reduced the proportion of Treg cells. • SMG/MSC vaccination increased MHC1 and HSPs proteins expression.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Intradiencephalon injection of histamine inhibited the recovery of locomotor function of spinal cord injured zebrafish

Human spinal cord injury (SCI) usually causes irreversible disability beneath the injured site due to poor neural regeneration. On the contrary, zebrafish show significant regenerative ability after SCI, thus is usually worked as an animal model for studying neuroregeneration. Most of the previous SCI studies focused on the local site of SCI, the supraspinal-derived signals were rarely mentioned. Here we showed that intradiencephalon injection of histamine (HA) inhibited the locomotor recovery in adult zebrafish post-SCI. Immunofluorescence results showed that intradiencephalon HA administration increased the activated microglia 3 days post injury (dpi), promoted the proliferation of radial glial cells at 7 dpi and affected the morphology of radial glial cells at 11 dpi. Furthermore, quantitative real-time polymerase chain reaction (qRT-PCR) results showed that intradiencephalon HA administration also reduced the expression of neurotrophic factors including brain-derived neurotrophic factor (BDNF) and insulin-like growth factor1 (IGF-1) at the lesion site, however, had no effect on the expression of pro-inflammatory factors such as TNF-alpha and IL-1 beta. Hence, our data suggested that exogenous intradiencephalon HA retarded locomotor recovery in spinal cord injured zebrafish via modulating the repair microenvironment. - Highlights: • A novel model to study the effect of supraspinal signals on repair of SCI zebrafish. • Exogenous HA administration suppresses the locomotor function recovery after SCI. • Exogenous HA administration modulates the repair microenvironment after SCI.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Disrupting nNOS-PSD-95 coupling in the hippocampal dentate gyrus promotes extinction memory retrieval

Granule cells in the dentate gyrus regenerate constantly in adult hippocampus and then integrate into neural circuits in the hippocampus thereby providing the neural basis for learning and memory. Promoting the neurogenesis in the hippocampus facilitates learning and memory such as spatial learning, object identification, and extinction learning. The interaction between neuronal nitric oxide synthase (nNOS) and postsynaptic density protein-95 (PSD-95) is reported to negatively regulate neurogenesis in brain, so we hypothesized that disrupting this interaction might facilitate the neurogenesis in the dentate gyrus (DG) and thus enhance the extinction memory retrieval of fear learning. We found that uncoupling the nNOS-PSD-95 complex in remote contextual fear condition promoted both neuronal proliferation and survival in the DG, contributing to an enhanced retrieval of the extinction memory. Moreover, the nNOS-PSD-95 uncoupling-induced neurogenesis may be mediated by the extracellular signal-regulated kinase (ERK) as the phosphorylation level of ERK1/2 was increased after uncoupling. These findings suggest that the nNOS-PSD-95 complex may serve as a novel target for the treatment of post-traumatic stress disorder (PTSD). - Highlights: • nNOS-PSD-95 disruption promoted neurogenesis in the DG. • Delayed disruption of nNOS-PSD-95 still enhanced remote contextual fear extinction. • The enhancement of fear extinction relied on neuronal proliferation. • nNOS-PSD-95 disruption increased ERK1/2 phosphorylation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

The NAMPT/E2F2/SIRT1 axis promotes proliferation and inhibits p53-dependent apoptosis in human melanoma cells

Melanoma is the most common primary malignant neoplasm in adults, causing more deaths than any other skin cancer, necessitating the development of new target-based approaches. Current evidence suggests SIRT1, the mammalian nicotinamide adenine dinucleotide (NAD{sup +})-dependent protein deacetylase, and nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting NAD{sup +} biosynthetic enzyme, together comprise a novel systemic regulatory network to play a pivotal role in cell proliferation and apoptosis. Nevertheless, how the regulation of this cofactor interfaces with signal transduction network remains poorly understood in melanoma. Here, we report NAMPT is highly expressed in melanomaassociated with poor overall survival in patients. Pharmacological and genetic inhibition of NAMPT decreased NAD{sup +} levels and melanoma cell proliferation capacity, and NAMPT knockdown induced apoptosis through the activity of the tumor suppressor p53. Next, we demonstrate NAMPT regulates the transcription factor E2F family member 2 (E2F2) in the apoptosis process. Downstream, E2F2 control the mRNA and protein levels of SIRT1. Finally, we find NAMPT mediates the apoptosis resistance of melanoma cells through NAMPT-E2F2-SIRT1 axis, more than NAD{sup +}-driven transcriptional program. Accordingly, our results demonstrated that NAMPT is a prognostic marker in melanoma, and the identificationofNAMPT-E2F2-SIRT1 pathway establishes another link between NAMPT and apoptosis events in melanoma, with therapeutic implications for this deadly cancer. - Highlights: • The expression of NAMPT is remarkably high in melanoma tumors and associated with poor overall survival in patients. • NAMPT is required for E2F2 and SIRT1 expression for regulating p53 and apoptosis levels in human melanoma cells. • NAMPT/E2F2/SIRT1 is a novel regulatory pathway that governs melanoma cells proliferation and apoptosis resistance.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Copine1 regulates neural stem cell functions during brain development

Highlights: • Copine1 is highly expressed in neural lineage cells at early developmental stages. • Copine1 regulates the proliferation of embryonic neural stem cells (NSCs). • Copine1 regulates the differentiation of NSCs into neuronal and glial lineages. • mTOR signaling pathway is involved in Copine1-meidated neural development. Copine 1 (CPNE1) is a well-known phospholipid binding protein in plasma membrane of various cell types. In brain cells, CPNE1 is closely associated with AKT signaling pathway, which is important for neural stem cell (NSC) functions during brain development. Here, we investigated the role of CPNE1 in the regulation of brain NSC functions during brain development and determined its underlying mechanism. In this study, abundant expression of CPNE1 was observed in neural lineage cells including NSCs and immature neurons in human. With mouse brain tissues in various developmental stages, we found that CPNE1 expression was higher at early embryonic stages compared to postnatal and adult stages. To model developing brain in vitro, we used primary NSCs derived from mouse embryonic hippocampus. Our in vitro study shows decreased proliferation and multi-lineage differentiation potential in CPNE1 deficient NSCs. Finally, we found that the deficiency of CPNE1 downregulated mTOR signaling in embryonic NSCs. These data demonstrate that CPNE1 plays a key role in the regulation of NSC functions through the activation of AKT-mTOR signaling pathway during brain development.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Unexpected role of the IMD pathway in Drosophila gut defense against Staphylococcus aureus

Highlights: • Drosocrystallin (Dcy) is involved in resistance against oral infection with S. aureus in adult Drosophila. • The presence of S. aureus does not trigger reactive oxygen species (ROS) production in the gut. • The IMD pathway is required for the clearance of ingested S. aureus. In this study, fruit fly of the genus Drosophila is utilized as a suitable model animal to investigate the molecular mechanisms of innate immunity. To combat orally transmitted pathogenic Gram-negative bacteria, the Drosophila gut is armed with the peritrophic matrix, which is a physical barrier composed of chitin and glycoproteins: the Duox system that produces reactive oxygen species (ROS), which in turn sterilize infected microbes, and the IMD pathway that regulates the expression of antimicrobial peptides (AMPs), which in turn control ROS-resistant pathogens. However, little is known about the defense mechanisms against Gram-positive bacteria in the fly gut. Here, we show that the peritrophic matrix protects Drosophila against Gram-positive bacteria S. aureus. We also define the few roles of ROS in response to the infection and show that the IMD pathway is required for the clearance of ingested microbes, possibly independently from AMP expression. These findings provide a new aspect of the gut defense system of Drosophila, and helps to elucidate the processes of gut-microbe symbiosis and pathogenesis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

The Pleckstrin homology like domain family member, TDAG51, is temporally regulated during skeletal muscle regeneration

Highlights: • TDAG51 expression is low but detectable in undamaged skeletal muscle. • Muscle expression of TDAG51 increases significantly in response to damage and insulin-like growth factor 1 (IGF-1). • Mice lacking TDAG51 display reduced functional capacity and a delay in the early regenerative phases of muscle repair. • Despite early delays in regeneration, TDAG51null mice can compensate and normalize muscle repair by 10 days post injury. The capacity for skeletal muscle to repair from daily insults as well as larger injuries is a vital component to maintaining muscle health over our lifetime. Given the importance of skeletal muscle for our physical and metabolic well-being, identifying novel factors mediating the growth and repair of skeletal muscle will thus build our foundational knowledge and help lead to potential therapeutic avenues for muscle wasting disorders. To that end, we investigated the expression of T-cell death associated gene 51 (TDAG51) during skeletal muscle repair and studied the response of TDAG51 deficient (TDAG51{sup -/-}) mice to chemically-induced muscle damage. TDAG51 mRNA and protein expression within uninjured skeletal muscle is almost undetectable but, in response to chemically-induced muscle damage, protein levels increase by 5 days post-injury and remain elevated for up to 10 days of regeneration. To determine the impact of TDAG51 deletion on skeletal muscle form and function, we compared adult male TDAG51{sup -/-} mice with age-matched wild-type (WT) mice. Body and muscle mass were not different between the two groups, however, in situ muscle testing demonstrated a significant reduction in force production both before and after fatiguing contractions in TDAG51{sup -/-} mice. During the early phases of the regenerative process (5 days post-injury), TDAG51{sup -/-} muscles display a significantly larger area of degenerating muscle tissue concomitant with significantly less regenerating area compared to WT (as demonstrated by embryonic myosin heavy chain expression). Despite these early deficits in regeneration, TDAG51{sup -/-} muscles displayed no morphological deficits by 10 days post injury compared to WT mice. Taken together, the data presented herein demonstrate TDAG51 expression to be upregulated in damaged skeletal muscle and its absence attenuates the early phases of muscle regeneration.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Secreted glycoprotein BmApoD1 plays a critical role in anti-oxidation and anti-apoptosis in Bombyx mori

Highlights: • Three apolipoprotein D homologs were identified from the genome of lepidopteran insect. • N-linked glycans were essential to the secretion of BmApoD1. • BmApoD1 transcription was up-regulated after oxidant or starvation treatment in larvae. • BmApoD1 can protect cells from oxidative stress and reduce actinomycin D-induced cell apoptosis. Recent studies highlighted that apolipoprotein D (ApoD) and its homologs exert neuroprotective and antioxidant functions in mammals and Drosophila. Unlike mammals and Drosophila, lepidopteran insects possess three distinct ApoD homologs. However, few information on their functions in lepidopteran insects are available. In this study, we investigated the protective potential of a novel ApoD homolog, BmApoD1, in Bombyx mori. Quantitative PCR analyses demonstrated that BmApoD1 is extensively expressed at low levels during the larval stage but abundantly expressed in the testis during the pupal and adult stages. Tryptophan fluorescence titration demonstrated that recombinant BmApoD1 protein can bind retinoic acid and ergosterol. In addition, we provided evidence that N-linked glycans of BmApoD1 are essential to BmApoD1 secretion, and three residues, namely, Asp69, Asp104, and Asp196, are the glycosylation sites of BmApoD1. Furthermore, we showed that BmApoD1 is significantly up-regulated in the larvae after oxidant or starvation treatment. The recombinant BmApoD1 protein can protect cells from oxidative stress induced by H{sub 2}O{sub 2} and reduce actinomycin D-induced cell apoptosis. These observations, together with the transcriptional up-regulation of BmApoD1 in several tissues upon oxidative insult, identify BmApoD1 as a potent antioxidant. Our results demonstrate that BmApoD1 is critical for metabolic adaptation of B. mori to environmental challenges.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

GBP3 promotes glioma cell proliferation via SQSTM1/p62-ERK1/2 axis

Highlights: • GBP3, an interferon-inducible large GTPase, is highly elevated in human glioma tumors. • GBP3 promotes glioma cell growth in vitro and in vivo through the SQSTM1/p62-ERK1/2 pathway. • GBP3 expression predicts poor outcome of glioma patients. Guanylate binding proteins (GBPs) are interferon-inducible large GTPases and play a crucial role in cell-autonomous immunity. However, the biology function of GBPs in cancer remains elusive. GBP3 is specifically expressed in adult brain. Here we show that GBP3 is highly elevated in human glioma tumors and glioma cell lines. Overexpression of GBP3 dramatically increased glioma cell proliferation whereas silencing GBP3 by RNA interference produced opposite effects. We further showed that GBP3 expression was able to induce sequestosome-1(SQSTM1, also named p62) expression and activate extracellular signal-regulated kinase (ERK1/2). The SQSTM1-ERK1/2 signaling cascade was essential for GBP3-promoted cell growth because depletion of SQSTM1 markedly reduced the phosphorylated ERK1/2 levels and GBP3-mediated cell growth, and inhibition of mitogen-activated protein kinase/ERK kinase abolished GBP3-induced glioma cell proliferation. Consistently, GBP3 overexpression significantly promoted glioma tumor growth in vivo and its expression was inversely correlated with the survival rate of glioma patients. Taken together, these results for the first time suggest that GBP3 contributes to the proliferation of glioma cells via regulating SQSTM1-ERK1/2 pathway, and GBP3 might represent as a new potential therapeutic target against glioma.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Exosomes derived from human umbilical vein endothelial cells promote neural stem cell expansion while maintain their stemness in culture

The neural stem cell (NSC) niche in subventricular zone (SVZ) of adult mammalian brain contains dense vascular plexus, where endothelial cells (ECs) regulate NSCs by releasing plenty of angiocrine factors. However, the role of ECs-derived exosomes, a novel type of mediators of intercellular communications, in the regulation of NSCs remains unclear. In the current study, primary NSCs isolated from embryonic mouse brains form more neurospheres when cultured in the presence of human umbilical vein endothelial cells (HUVECs). The supportive role of ECs in the coculture was significantly attenuated when GW4869, a blocker of exosome formation, was included, suggesting that HUVECs-derived exosomes played a significant role in supporting NSCs. In order to investigate the role of ECs-derived exosomes on NSCs, we collected exosomes from HUVECs. We found that HUVECs-derived exosomes could significantly promote the formation of neurospheres by primary murine NSCs. EdU incorporation and TUNEL assays indicated that the proliferation of NSCs increased while apoptosis decreased when cultured in the presence of HUVECs-derived exosomes. NSCs incubated with the HUVECs-derived exosomes maintained their potential of multi-lineage differentiation potentials. The expression of stemness-related genes was up-regulated. These data suggested that ECs-derived exosomes could play an importantly role in NSC niche, and they might be used as a reagent for ex vivo NSC amplification for medical application.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Distinct roles of Rheb and Raptor in activating mTOR complex 1 for the self-renewal of hematopoietic stem cells

Highlights: • Rheb deficiency causes abnormalities in myeloid lineage cells. • Rheb deficiency impairs HSC-derived hematopoiesis post-transplantation. • Unlike Raptor, Rheb is dispensable for HSC self-renewal at steady-state. The mammalian target of rapamycin (mTOR) complex 1 (mTORC1) senses a cell's energy status and environmental levels of nutrients and growth factors. In response, mTORC1 mediates signaling that controls protein translation and cellular metabolism. Although mTORC1 plays a critical role in hematopoiesis, it remains unclear which upstream stimuli regulate mTORC1 activity in the context of hematopoietic stem cells (HSC) maintenance in vivo. In this study, we investigated the function of Rheb, a critical regulator of mTORC1 activity controlled by the PI3K-AKT-TSC axis, both in HSC maintenance in mice at steady-state and in HSC-derived hematopoiesis post-transplantation. In contrast to the severe hematopoietic dysfunction caused by Raptor deletion, which completely inactivates mTORC1, Rheb deficiency in adult mice did not show remarkable hematopoietic failure. Lack of Rheb caused abnormalities in myeloid cells but did not have impact on hematopoietic regeneration in mice subjected to injury by irradiation. As previously reported, Rheb deficiency resulted in defective HSC-derived hematopoiesis post-transplantation. However, while Raptor is essential for HSC competitiveness in vivo, Rheb is dispensable for HSC maintenance under physiological conditions, indicating that the PI3K-AKT-TSC pathway does not contribute to mTORC1 activity for sustaining HSC self-renewal activity at steady-state. Thus, the various regulatory elements that impinge upstream of mTORC1 activation pathways are differentially required for HSC homeostasis in vivo.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Knockdown of laminin α5 stimulates intestinal cell differentiation

Highlights: • Knockdown of laminin α5 accelerates early intestinal cell differentiation. • Laminin α5 depletion triggers precocious sucrase-isomaltase expression. • A transient increase of dipeptidylpeptidase IV, villin, CDX2 and HNF-1α is noted. • Dome formation and trans-epithelial resistance are also transiently enhanced. Interactions between cells and the extracellular matrix regulate a wide range of cell processes such as proliferation and differentiation. Laminins are major components of the basement membrane that actively participate in most biological functions via their interactions with a variety of specific cell receptors. The α5-containing laminins (LAMA5) are one of the three main types of laminins identified at the epithelial basal lamina in the adult intestine. The aim of the present study was to investigate the role of α5-containing laminins on intestinal cell proliferation and differentiation. Using an shRNA targeting approach, the effects of knocking down the expression of LAMA5 were investigated in the enterocytic-like Caco-2/15 cell line, a well-characterized model for intestinal cell differentiation. Surprisingly, the abolition of the laminin α5 chain resulted in a drastic increase in the differentiation marker sucrase-isomaltase which was correctly expressed at the apical pole of the cells as observed by indirect immunofluorescence. Transient increases of dipeptidylpeptidase IV, villin, CDX2, HNF-1α, HNF-4α and transepithelial resistance as well as an apparent redistribution of the junctional components ZO-1 and E-cadherin were also observed at early stages of differentiation but no specific effect was observed on cell proliferation as evaluated by BrdU incorporation. Taken together, these data suggest that α5-containing laminins repress intestinal differentiation in its early stages.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

The splicing factor transformer2 (tra2) functions in the Drosophila fat body to regulate lipid storage

Highlights: • Transformer2 (tra2) is a splicing protein whose metabolic functions are unknown. • Tra2 functions in the Drosophila fat body to modulate triglyceride levels. • Decreasing tra2 in the fat body promotes starvation resistance. • Tra2 alters CPT1 splicing in the fly fat body. Excess nutrients are stored as triglycerides mainly in the adipose tissue of an animal and these triglycerides are located in structures called lipid droplets. Previous genome-wide RNAi screens in Drosophila cells identified splicing factors as playing a role in lipid droplet formation. Our lab has recently identified the SR protein, 9G8, as an important factor in fat storage as decreasing its levels results in augmented triglyceride storage in the fat body. Previous in vitro studies have implicated 9G8 in the regulation of splicing of the sex determination gene doublesex (dsx) by binding to transformer (tra) and transformer2 (tra2); however, any function of these sex determination proteins in regulating metabolism is unknown. In this study, we have uncovered a role of tra2 to regulate fat storage in vivo. Inducing tra2{sup dsRNA} in the adult fat body resulted in an increase in triglyceride levels but had no effect on glycogen storage. Consistent with the triglyceride phenotype, tra2 knockdown flies lived longer under starvation conditions. In addition, this increase in triglycerides is due to more fat storage per cell and not an increase in the number of fat cells. Interestingly, the splicing of CPT1, an enzyme involved in the breakdown of lipids, was altered in flies with decreased tra2. The less-catalytically active isoform of CPT1 accumulated in tra2{sup dsRNA} flies suggesting a decrease in lipid breakdown, which is consistent with the increased triglyceride levels observed in these flies. Together, these results suggest a link between mRNA splicing, sex determination and lipid metabolism and may provide insight into the mechanisms underlying tissue-specific splicing and nutrient storage.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Trek2a regulates gnrh3 expression under control of melatonin receptor Mt1 and α2-adrenoceptor

Gonadotrophin-releasing hormone (GnRH) expression is associated with the two-pore domain potassium ion (K{sup +}) channel-related K{sup +} (TREK) channel trek2a expression and melatonin levels. We aimed to investigate correlation of trek2a expression with gnrh3 expression, and regulatory mechanisms of trek2a expression by the melatonin receptor Mt1 and α{sub 2}-adrenoceptor which are regulated by melatonin. trek2a specific siRNA, Mt1 antagonist luzindole and α{sub 2}-adrenoceptor antagonist prazosin were administered into the adult zebrafish brain and gene expressions were examined by real-time PCR. trek2a specific siRNA administration significantly reduced expression levels of trek2a, gnrh3 and mt1. Luzindole administration suppressed trek2a and gnrh3 expressions. Prazosin administration reduced trek2a and gnrh3 expressions. It is suggested that Trek2a regulates gnrh3 expression under the control of Mt1 and α{sub 2}-adrenoceptor.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

High expression of A-type lamin in the leading front is required for Drosophila thorax closure

Highlights: • Lamin C expression is high at the leading front of the closing tissue in pupal notum. • Lamin C is required for the coordinated movement of the leading front. • The closure defect by Lamin C knockdown correlates with insufficient F-actin formation. Tissue closure involves the coordinated unidirectional movement of a group of cells without loss of cell–cell contact. However, the molecular mechanisms controlling the tissue closure are not fully understood. Here, we demonstrate that Lamin C, the sole A-type lamin in Drosophila, contributes to the process of thorax closure in pupa. High expression of Lamin C was observed at the leading front of the migrating wing imaginal discs. Live imaging analysis revealed that knockdown of Lamin C in the thorax region affected the coordinated movement of the leading front, resulting in incomplete tissue fusion required for formation of the adult thorax. The closure defect due to knockdown of Lamin C correlated with insufficient accumulation of F-actin at the front. Our study indicates a link between A-type lamin and the cell migration behavior during tissue closure.

60 APPLIED LIFE SCIENCES↗