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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 1,747 records · Page 97
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DOE OSTI2016

The Mycobacterium tuberculosis desaturase DesA1 (Rv0824c) is a Ca{sup 2+} binding protein

The hallmark feature of Mycobacterium tuberculosis (M.tb) the causative agent of human tuberculosis, is its complex lipid rich cell wall comprised primarily of mycolic acids, long chain fatty acids that play a key role in structural stability and permeability of the cell wall. In addition, they are involved in inhibiting phagosome-lysosome fusion and aid in granuloma formation during the pathogenic process. M.tb DesA1 is an essential acyl-acyl carrier protein desaturase predicted to catalyze the introduction of position specific double bonds during the biosynthesis of mycolic acids. This protein is one among three annotated desaturases (DesA1-3) in the M.tb genome but is unique in containing a βγ-crystallin Greek key signature motif, a well-characterized fold known to mediate Ca{sup 2+} binding in both prokaryotic and eukaryotic organisms. Using Isothermal Titration Calorimetry and {sup 45}CaCl{sub 2} overlay, we demonstrate that Ca{sup 2+} binds to DesA1. Spectroscopic measurements suggested that this binding induces changes in protein conformation but does not lead to significant alterations in the secondary structure of the protein, a feature common to several βγ-crystallins. An M. smegmatis strain over-expressing M.tb desA1 showed a Ca{sup 2+} dependent variation in surface phenotype, revealing a functional role for Ca{sup 2+}in DesA1 activity. This study represents the first identification of a Ca{sup 2+} binding βγ-crystallin in M.tb, emphasizing the implicit role of Ca{sup 2+} in the pathogenesis of M.tb. - Highlights: • Mycobacterium tuberculosis DesA1 is an essential acyl-ACP desaturase. • DesA1 was identified to contain a βγ-crystallin Greek key signature motif. • Ca{sup 2+} binds to DesA1 with an affinity of 53 μM and induces changes in its conformation. • M. smegmatis overexpressing M.tb DesA1 shows a Ca{sup 2+} dependent phenotype. • Targetting the Ca{sup 2+} dependent function of DesA1 could be of therapeutic value.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Imbalance of mitochondrial dynamics in Drosophila models of amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is the most common neurodegenerative disease, characterized by progressive and selective loss of motor neurons in the brain and spinal cord. DNA/RNA-binding proteins such as TDP-43, FUS, and TAF15 have been linked with the sporadic and familial forms of ALS. However, the exact pathogenic mechanism of ALS is still unknown. Recently, we found that ALS-causing genes such as TDP-43, FUS, and TAF15 genetically interact with mitochondrial dynamics regulatory genes. In this study, we show that mitochondrial fission was highly enhanced in muscles and motor neurons of TDP-43, FUS, and TAF15-induced fly models of ALS. Furthermore, the mitochondrial fission defects were rescued by co-expression of mitochondrial dynamics regulatory genes such as Marf, Opa1, and the dominant negative mutant form of Drp1. Moreover, we found that the expression level of Marf was decreased in ALS-induced flies. These results indicate that the imbalance of mitochondrial dynamics caused by instability of Marf is linked to the pathogenesis of TDP-43, FUS, and TAF15-associated ALS. - Highlights: • Mitochondrial fission is highly enhanced in TDP-43, FUS, and TAF15-induced fly models of ALS. • Excessive mitochondrial fragmentation in fly models of ALS is restored by mitochondrial dynamics regulatory genes. • Level of Marf protein is decreased in TDP-43, FUS, and TAF15-mediated ALS. • Imbalance of mitochondrial dynamics caused by Marf instability is linked to the pathogenesis of ALS.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Negative regulation of NOD1 mediated angiogenesis by PPARγ-regulated miR-125a

Infection with pathogens activates the endothelial cell and its sustained activation may result in impaired endothelial function. Endothelial dysfunction contributes to the pathologic angiogenesis that is characteristic of infection-induced inflammatory pathway activation. Nucleotide-binding oligomerization domain-containing protein 1 (NOD1) is a protein receptor which recognizes bacterial molecules and stimulates an immune reaction in various cells; however, the underlying molecular mechanisms in the regulation of inflammation-triggered angiogenesis are not fully understood. Here we report that peroxisome proliferator-activated receptor gamma (PPARγ)-mediated miR-125a serves as an important regulator of NOD1 agonist-mediated angiogenesis in endothelial cells by directly targeting NOD1. Treatment of human umbilical vein endothelial cells with natural PPARγ ligand, 15-Deoxy-Delta12,14-prostaglandin J2, led to inhibition of NOD1 expression; contrarily, protein levels of NOD1 were significantly increased by PPARγ knockdown. We report that PPARγ regulation of NOD1 expression is a novel microRNA-mediated regulation in endothelial cells. MiR-125a expression was markedly decreased in human umbilical vein endothelial cells subjected to PPARγ knockdown while 15-Deoxy-Delta12,14-prostaglandin J2 treatment increased the level of miR-125a. In addition, NOD1 is closely regulated by miR-125a, which directly targets the 3′ untranslated region of NOD1. Moreover, both overexpression of miR-125a and PPARγ activation led to inhibition of NOD1 agonist-induced tube formation in endothelial cells. Finally, NOD1 agonist increased the formation of cranial and subintestinal vessel plexus in zebrafish, and this effect was abrogated by concurrent PPARγ activation. Overall, these findings identify a PPARγ-miR-125a-NOD1 signaling axis in endothelial cells that is critical in the regulation of inflammation-mediated angiogenesis. - Highlights: • Expression of NOD1 is regulated by PPARγ signaling in HUVECs. • PPARγ-regulated NOD1 expression is mediated at least in part by of miR-125a. • NOD1 is a novel target gene of miR-125. • miR-125a inhibits inflammation induced angiogenesis in HUVECs. • PPARγ activation inhibits inflammation mediated angiogenesis in vitro and in vivo.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Antimicrobial activity and mechanism of PDC213, an endogenous peptide from human milk

Human milk has always been considered an ideal source of elemental nutrients to both preterm and full term infants in order to optimally develop the infant's tissues and organs. Recently, hundreds of endogenous milk peptides were identified in human milk. These peptides exhibited angiotensin-converting enzyme inhibition, immunomodulation, or antimicrobial activity. Here, we report the antimicrobial activity and mechanism of a novel type of human antimicrobial peptide (AMP), termed PDC213 (peptide derived from β-Casein 213-226 aa). PDC213 is an endogenous peptide and is present at higher levels in preterm milk than in full term milk. The inhibitory concentration curve and disk diffusion tests showed that PDC213 had obvious antimicrobial against S. aureus and Y. enterocolitica, the common nosocomial pathogens in neonatal intensive care units (NICUs). Fluorescent dye methods, electron microscopy experiments and DNA-binding activity assays further indicated that PDC213 can permeabilize bacterial membranes and cell walls rather than bind intracellular DNA to kill bacteria. Together, our results suggest that PDC213 is a novel type of AMP that warrants further investigation. - Highlights: • PDC213 is an endogenous peptide presenting higher levels in preterm milk. • PDC213 showed obvious antimicrobial against S. aereus and Y. enterocolitica. • PDC213 can permeabilize bacterial membranes and cell walls to kill bacterias. • PDC213 is a novel type of antimicrobial peptides worthy further investigation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Osteopontin attenuates acute gastrointestinal graft-versus-host disease by preventing apoptosis of intestinal epithelial cells

Background and aims: Acute graft-versus-host disease (GVHD) is a major complication after allogeneic hematopoietic stem cell transplantation, which often targets gastrointestinal (GI) tract. Osteopontin (OPN) plays an important physiological role in the efficient development of Th1 immune responses and cell survival by inhibiting apoptosis. The role of OPN in acute GI-GVHD is poorly understood. In the present study, we investigated the role of OPN in donor T cells in the pathogenicity of acute GI-GVHD. Methods: OPN knockout (KO) mice and C57BL/6 (B6) mice were used as donors, and (C57BL/6 × DBA/2) F1 (BDF1) mice were used as allograft recipients. Mice with acute GI-GVHD were divided into three groups: the control group (BDF1→BDF1), B6 group (B6→BDF1), and OPN-KO group (OPN-KO→BDF1). Bone marrow cells and spleen cells from donors were transplanted to lethally irradiated recipients. Clinical GVHD scores were assessed daily. Recipients were euthanized on day 7 after transplantation, and colons and small intestines were collected for various analyses. Results: The clinical GVHD score in the OPN-KO group was significantly increased compared with the B6 and control groups. We observed a difference in the severity of colonic GVHD between the OPN-KO group and B6 group, but not small intestinal-GVHD between these groups. Interferon-γ, Tumor necrosis factor-α, Interleukin-17A, and Interleukin-18 gene expression in the OPN-KO group was differed between the colon and small intestine. Flow cytometric analysis revealed that the fluorescence intensity of splenic and colonic CD8 T cells expressing Fas Ligand was increased in the OPN-KO group compared with the B6 group. Conclusion: We demonstrated that the importance of OPN in T cells in the onset of acute GI-GVHD involves regulating apoptosis of the intestinal cell via the Fas-Fas Ligand pathway. - Highlights: • A lack of osteopontin in donor cells exacerbated clinical gastrointestinal GVHD. • Donor cells lacking osteopontin affected intestinal inflammation of GVHD. • Donor cells lacking osteopontin increased apoptotic epithelial cells in GVHD. • Osteopontin plays an anti-inflammatory role in acute gastrointestinal GVHD.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

The changes in glucose metabolism and cell proliferation in the kidneys of polycystic kidney disease mini-pig models

The pathogenic mechanism of polycystic kidney disease (PKD) is unclear. Abnormal glucose metabolism is maybe involved in hyper-proliferation of renal cyst epithelial cells. Mini-pigs are more similar to humans than rodents and therefore, are an ideal large animal model. In this study, for the first time, we systematically investigated the changes in glucose metabolism and cell proliferation signaling pathways in the kidney tissues of chronic progressive PKD mini-pig models created by knock-outing PKD1gene. The results showed that in the kidneys of PKD mini-pigs, the glycolysis is increased and the expressions of key oxidative phosphorylation enzymes Complexes I and IV significantly decreased. The activities of mitochondrial respiration chain Complexes I and IV significantly decreased; the phosphorylation level of key metabolism-modulating molecule AMP-activated protein kinase (AMPK) significantly decreased; and the mammalian target of rapamycin (mTOR) and extracellular signal-regulated kinase (ERK) signaling pathway are activated obviously. This study showed that in the kidneys of PKD mini-pigs, the level of glycolysis significantly increased, oxidative phosphorylation significantly decreased, and cell proliferation signaling pathways significantly activated, suggesting that metabolic changes in PKD may result in the occurrence and development of PKD through the activation of proliferation signaling pathways. - Highlights: • Mini-pigs are more similar to humans than rodents and are an ideal animal model. • The study was first performed in chronic model created by knock-outing PKD1gene. • We investigated the changes in glucose metabolism and cell proliferation. • The glycolysis was significantly increased in the kidneys of PKD mini-pigs. • The mTOR and ERK signaling pathway are activated obviously in PKD mini-pigs.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Effect of calcium carbonate particle shape on phagocytosis and pro-inflammatory response in differentiated THP-1 macrophages

Phagocytosis is a physiological process used by immune cells such as macrophages to actively ingest and destroy foreign pathogens and particles. It is the cellular process that leads to the failure of drug delivery carriers because the drug carriers are cleared by immune cells before reaching their target. Therefore, clarifying the mechanism of particle phagocytosis would have a significant implication for both fundamental understanding and biomedical engineering. As far as we know, the effect of particle shape on biological response has not been fully investigated. In the present study, we investigated the particle shape-dependent cellular uptake and biological response of differentiated THP-1 macrophages by using calcium carbonate (CaCO{sub 3})-based particles as a model. Transmission electron microscopy analysis revealed that the high uptake of needle-shaped CaCO{sub 3} particles by THP-1 macrophages because of their high phagocytic activity. In addition, the THP-1 macrophages exposed to needle-shaped CaCO{sub 3} accumulated a large amount of calcium in the intracellular matrix. The enhanced release of interleukin-8 (IL-8) and tumor necrosis factor-alpha (TNF-α) by the THP-1 macrophages suggested that the needle-shaped CaCO{sub 3} particles trigger a pro-inflammatory response. In contrast, no pro-inflammatory response was induced in undifferentiated THP-1 monocytes exposed to either needle- or cuboidal-shaped CaCO{sub 3} particles, probably because of their low phagocytic activity. We also found that phosphate-coated particles efficiently repressed cellular uptake and the resulting pro-inflammatory response in both THP-1 macrophages and primary peritoneal macrophages. Our results indicate that the pro-inflammatory response of macrophages upon exposure to CaCO{sub 3} particles is shape- and surface property-dependent, and is mediated by the intracellular accumulation of calcium ions released from phagocytosed CaCO{sub 3} particles. - Highlights: • Needle-shaped CaCO{sub 3} particles are phagocytized by macrophages. • Needle-shaped CaCO{sub 3} particles induce intracellular accumulation of calcium ions. • Needle-shaped CaCO{sub 3} particles induce pro-inflammatory responses. • Surface-coating with phosphate decreases adverse effects on macrophages.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Atorvastatin prevents endothelial dysfunction in high glucose condition through Skp2-mediated degradation of FOXO1 and ICAM-1

Highlights: • The molecular mechanism of statin in diabetes to reduce the endothelial cell (EC) dysfunction and inflammation is proposed. • In high-glucose exposure, increased FOXO1 and ICAM1 leads to EC dysfunction and increased monocyte adhesion. • Atorvastatin facilitates ubiquitin-mediated degradation of FOXO1 and ICAM1 through Skp2 in EC exposed to high-glucose. The 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor atorvastatin has been reported to exert vasculo-protective action in diabetes. We investigated the vasculo-protective mechanism of atorvastatin by evaluating its effect on two major pathogenic molecules, FOXO1 and ICAM1, mediated by S-phase kinase-associated protein 2 (Skp2) in diabetic endothelial dysfunction.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

A 2 bp deletion in the mitochondrial ATP 6 gene responsible for the NARP (neuropathy, ataxia, and retinitis pigmentosa) syndrome

Highlights: • A 2 bp deletion of the ATP6 gene causes a truncated a subunit. • The truncated a subunit is responsible for a decrease of complex V activity. • A 2 bp deletion of the ATP6 gene causes a NARP syndrome. Mitochondrial (mt) DNA-associated NARP (neurogenic muscle weakness, ataxia, and retinitis pigmentosa) syndrome is due to mutation in the MT-ATP6 gene. We report the case of a 18-year-old man who presented with deafness, a myoclonic epilepsy, muscle weakness since the age of 10 and further developed a retinitis pigmentosa and ataxia. The whole mtDNA analysis by next-generation sequencing revealed the presence of the 2 bp microdeletion m.9127–9128 del AT in the ATP6 gene at 82% heteroplasmy in muscle and to a lower load in blood (10–20%) and fibroblasts (50%). Using the patient's fibroblasts, we demonstrated a 60% reduction of the oligomycin-sensitive ATPase hydrolytic activity, a 40% decrease in the ATP synthesis and determination of the mitochondrial membrane potential using the fluorescent probe tetramethylrhodamine, ethyl ester indicated a significant reduction in oligomycin sensitivity. In conclusion, we demonstrated that this novel AT deletion in the ATP6 gene is pathogenic and responsible for the NARP syndrome.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Ankyrin repeats as a dimerization module

Highlights: • We determined crystal structure of Legionella effector AnkC. • The structure displays seven ankyrin repeats with unique structural features. • AnkC dimerizes using outer surface of loops between ankyrin repeats. • This is the first example of ankyrin repeats-mediated dimerization. Legionella pneumophila is a pathogen, causing severe pneumonia in humans called Legionnaires' disease. AnkC (LegA12) is a poorly characterized 495-residue effector protein conserved in multiple Legionella species. Here, we report the crystal structure of a C-terminally truncated AnkC (2–384) at 3.2 Å resolution. The structure shows seven ankyrin repeats (ARs) with unique structural features. AnkC forms a dimer along the outer surface of loops between ARs. The dimer exists both in the crystal form and in solution, as shown by analytical ultracentrifugation. This is the first example of ARs as a dimerization module as opposed to solely a protein interaction domain. In addition, a novel α-helix insert between AR3-AR4 is positioned across the surface opposite the ankyrin groove. Sequence conservation suggests that the ankyrin groove of AnkC is a functional site that interacts with binding targets. This ankyrin domain structure is an important step towards a functional characterization of AnkC.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Parkinson's disease-associated mutant LRRK2 phosphorylates Rab7L1 and modifies trans-Golgi morphology

Highlights: • Rab7L1 is identified as a bona fide substrate of LRRK2. • Ser72 is a major in-cell phosphorylation site of Rab7L1. • Parkinson's-linked mutations in LRRK2 increase the Ser72 phosphorylation of Rab7L1. • Rab7L1 phosphorylation at Ser72 modifies the trans-Golgi morphology. Mutations in leucine-rich repeat kinase 2 (LRRK2) are the major genetic cause of autosomal-dominantly inherited Parkinson's disease. LRRK2 is implicated in the regulation of intracellular trafficking, neurite outgrowth and PD risk in connection with Rab7L1, a putative interactor of LRRK2. Recently, a subset of Rab GTPases have been reported as substrates of LRRK2. Here we examine the kinase activity of LRRK2 on Rab7L1 in situ in cells. Phos-tag analyses and metabolic labeling assays revealed that LRRK2 readily phosphorylates Golgi-localized wild-type Rab7L1 but not mutant forms that are distributed in the cytoplasm. In vitro assays demonstrated direct phosphorylation of Rab7L1 by LRRK2. Subsequent screening using Rab7L1 mutants harboring alanine-substitution for every single Ser/Thr residue revealed that Ser72 is a major phosphorylation site, which was confirmed by using a phospho-Ser72-specific antibody. Moreover, LRRK2 pathogenic Parkinson mutants altogether markedly enhanced the phosphorylation at Ser72. The modulation of Ser72 phosphorylation in Rab7L1 resulted in an alteration of the morphology and distribution of the trans-Golgi network. These data collectively support the involvement of Rab7L1 phosphorylation in the LRRK2-mediated cellular and pathogenetic mechanisms.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

The interaction of soybean reticulon homology domain protein (GmRHP) with Soybean mosaic virus encoded P3 contributes to the viral infection

Highlights: • GmRHP interacted with SMV-P3 in a screening of soybean cDNAs using mYTHS. • The interaction between GmRHP and SMV-P3 was confirmed using BiFC. • GmRHP's expression promoted TuMV replication and spread in N. benthamiana. • The interaction between SMV-P3 and GmRHP may contribute to the potyvirus infection. Soybean mosaic virus (SMV), a member of the Potyvirus genus, is a prevalent and devastating viral pathogen in soybean-growing regions worldwide. Potyvirus replication occurs in the 6K2-induced viral replication complex at endoplasmic reticulum exit sites. Potyvirus-encoded P3 is also associated with the endoplasmic reticulum and is as an essential component of the viral replication complex, playing a key role in viral replication. This study provides evidence that the soybean (Glycine max) reticulon homology domain protein (designated as GmRHP) interacts with SMV-P3 by using a two-hybrid yeast system to screen a soybean cDNA library. A bimolecular fluorescence complementation assay further confirmed the interaction, which occurred on the cytomembrane, endoplasmic reticulum and cytoskeleton in Nicotiana benthamiana cells. The transient expression of GmRHP can promote the coupling of Turnip mosaic virus replication and cell-to-cell movement in N. benthamiana. The interaction between the membrane protein SMV-P3 and GmRHP may contribute to the potyvirus infection, and GmRHP may be an essential host factor for P3's involvement in potyvirus replication.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Moesin and myosin IIA modulate phagolysosomal biogenesis in macrophages

Highlights: • SHP-1 interacts with moesin and myosin IIA during phagocytosis. • Moesin and myosin IIA are required for the biogenesis of phagolysosomes. • Moesin and myosin IIA are necessary for full bactericidal activity of macrophages. Biogenesis of phagolysosomes is central to the elimination of pathogens by macrophages. We previously showed that Src homology region 2 domain-containing phosphatase 1 (SHP-1) participates in the regulation of phagosome maturation. Through proteomics, we identified moesin and the non-muscle myosin-IIA as proteins interacting with SHP-1 during phagocytosis. Silencing of either moesin or myosin IIA with small interfering RNA inhibited phagosomal acidification and recruitment of LAMP-1. Moreover, the intraphagosomal oxidative burst was impaired in the absence of either SHP-1 or myosin IIA but not moesin. Finally, absence of either SHP-1, moesin, or myosin IIA ablated the capacity of macrophages to clear bacterial infection. Collectively, these results implicate both moesin and myosin IIA in the regulation of phagolysosome biogenesis and in host defense against infections.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Novel and rapid osteoporosis model established in zebrafish using high iron stress

Highlights: • Establish a rapid and robust zebrafish model of osteoporosis using high iron stress. • High iron-overloaded results in severe defects in skeletal development. • High iron stress leads significant downregulation of osteoblast-specific mRNAs. • This osteoporosis phenotype is effectively repaired by a known drug Alendronate (AL). • AL could rescue osteoporosis larval fish through facilitating osteoblast differentiation. Osteoporosis is a global public health concern and, it can result from numerous pathogenic mechanisms, many of which are closely related with age, nutritional disorders, endocrine imbalance, or adverse drug side effects presented by glucocorticoids, heparin, and anti-epileptics. Given its wide range etiologies, it is crucial to establish an animal model of osteoporosis for use in screening potential drugs quickly and effectively. Previous research has reported that an accumulation of elevated iron in the body is an independent risk factor for osteoporosis. As such, we sought to use both zebrafish larvae and adults to model an osteoporosis phenotype using high iron stress (FAC, ferric ammonium citrate). Skeletal staining results suggested that iron-overload caused a significant decrease in bone calcification as well as severe developmental cartilage defects. In addition, osteoblast and cartilage-specific mRNA expression levels were downregulated after exposure to a high-iron environment. Most importantly, we demonstrated in both larval and adult fish that high iron-induced osteogenic defects were significantly rescued using alendronate (AL), a drug known to be effective against to human osteoporosis. Even more, the repair effect of AL was achieved by facilitating osteoblast differentiation and targeting Bmp signaling. Taken together, our findings propose an rapid and effective osteoporosis model, which could be used widely for future osteoporosis drug screening.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

CaMKII-mediated phosphorylation of RyR2 plays a crucial role in aberrant Ca2+ release as an arrhythmogenic substrate in cardiac troponin T-related familial hypertrophic cardiomyopathy

Highlights: • We assessed RyR2 phosphorylation in FHC-related TnT-mutated transgenic mice. • The SpF was higher in TG cardiomyocytes than in non-TG cardiomyocytes. • RyR2 was susceptible to CaMKII-mediated phosphorylation due to the mutation. • Increase in diastolic [Ca{sup 2+}]{sub i} caused aberrant Ca{sup 2+} release and arrhythmia. Cardiac Troponin T (TnT) mutation-linked familial hypertrophic cardiomyopathy (FHC) is known to cause sudden cardiac death at a young age. Here, we investigated the role of the Ca{sup 2+} release channel of the cardiac sarcoplasmic reticulum (SR), ryanodine receptor (RyR2), in the pathogenic mechanism of lethal arrhythmia in FHC-related TnT-mutated transgenic mice (TG; TnT-delta160E).

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Effect of tamoxifen on the sphingolipid biosynthetic pathway in the different intraerythrocytic stages of the apicomplexa Plasmodium falciparum

Highlights: • The different intraerythrocytic stages of P. falciparum were treated with tamoxifen. • Analysis of labelled sphingolipids was performed by TLC, HPLC and UV-MALDI-TOF m. s. • Tamoxifen inhibits GCS and SM synthase although in a stage dependent way. • The accumulation of PI indicates an inhibitory action on GPI synthesis. • The differential effect on SLs with DHcer or Cer indicate they do not interconvert. Parasites of the genus Plasmodium responsible for Malaria are obligate intracellular pathogens residing in mammalian red blood cells, hepatocytes, or mosquito midgut epithelial cells. Regarding that detailed knowledge on the sphingolipid biosynthetic pathway of the apicomplexan protozoan parasites is scarce, different stages of Plasmodium falciparum were treated with tamoxifen in order to evaluate the effects of this drug on the glycosphingolipid biosynthesis. Thin layer chromatography, High performance reverse phase chromatography and UV-MALDI-TOF mass spectrometry were the tools used for the analysis. In the ring forms, the increase of NBD-phosphatidyl inositol biosynthesis was notorious but differences at NBD-GlcCer levels were undetectable. In trophozoite forms, an abrupt decrease of NBD-acylated GlcDHCer and NBD-GlcDHCer in addition to an increase of NBD-PC biosynthesis was observed. On the contrary, in schizonts, tamoxifen seems not to be producing substantial changes in lipid biosynthesis. Our findings indicate that in this parasite, tamoxifen is exerting an inhibitory action on Glucosylceramidesynthase and sphingomyelin synthase levels. Moreover, regarding that Plasmodium does not biosynthesize inositolphosphoceramides, the accumulation of phosphatidylinositol should indicate an inhibitory action on glycosylinositol phospholipid synthesis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Cyclosporin A inhibits adipogenic differentiation and regulates immunomodulatory functions of murine mesenchymal stem cells

Highlights: • CsA decreased BM-MSCs proliferation and increased cells apoptosis. • CsA inhibited BM-MSCs adipogenic differentiation. • CsA had no effect on the phenotype and osteogenic differentiation of BM-MSCs. • CsA inhibited BM-MSCs secreting IL-6 and promoted the expression of PD-L2. Aplastic anemia (AA) is generally considered as an immune-mediated bone marrow failure syndrome. Several studies show that bone marrow mesenchymal stem cells (BM-MSCs), as key cellular components of the bone marrow microenvironment, are also involved in the pathogenic mechanism of AA. Cyclosporin A (CsA) is a classic immunosuppressive drug for AA, and it specifically inhibits mammalian T cells by preventing activation of transcription factors involved in cytokine gene expression. However, little is known about the effect of CsA on the BM-MSCs. In this study, murine BM-MSCs were stimulated in the presence of CsA. Further, we found that CsA could inhibit murine BM-MSC proliferation and promote BM-MSC apoptosis, what's more CsA could inhibit adipogenic differentiation. Our study also showed that CsA could inhibit interleukin-6 expression in BM-MSCs, while promoting programmed death-ligand 2 expression. In conclusion, our results proposed that CsA may exert an effect on regulating the bone marrow environment by influencing BM-MSCs, which have a beneficial effect on treating AA.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

HMGB1 mediates HAdV-7 infection-induced pulmonary inflammation in mice

Highlights: • HMGB1 was associated with HAdV-7-induced lung inflammation and pathology. • HMGB1 promoted HAdV-7 replication and bound to TLR-4, TLR-9, and RAGE receptors. • Receptor engagement by HMGB1 activated NF-κB, inducing inflammatory mediators. • Thus, HMGB1 could serve as a therapeutic target in HAdV-7 infection. Human adenovirus (HAdV) is a common respiratory pathogen in children, with no safe and effective treatment currently available. HAdV type 7 (HAdV-7), in particular, causes severe pediatric pneumonia with a high incidence of sequelae and mortality. Clinical data and animal experiments suggest that HAdV-7-induced pneumonia promotes cell necrosis, releasing a large number of inflammatory mediators. In recent years, the high mobility group box-1 (HMGB1) protein, released by necrotic cells, has been shown to play important roles in several viral infections. Here, we show that HMGB1 levels gradually increased in the media supernatants of HAdV-7 infected A549 cells, starting at 12 h post-infection. In vivo, HMGB1 levels in BALF and mRNA levels in lung tissues significantly increased after 3 days of HAdV-7 infection. Among the HMGB1 receptor genes, TLR-4 and TLR-9 expression increased, and so did the receptor for advanced glycation end-products (RAGE). Interestingly, NF-κB levels also increased concomitantly. Conversely, when HMGB1 was blocked, the pathological scores from lung tissues, inflammatory mediator levels, and viral copy number all were reduced significantly; in addition, HMGB1-related signaling pathway molecules, namely TLR-4, TLR-9, RAGE, and NF-κB were also reduced. We conclude that HMGB1 promotes HAdV-7 replication and signals through TLR-4, TLR-9, and RAGE receptors to activate NF-κB, stimulating the release of inflammatory mediators and contributing to adenoviral pathology. Thus, HMGB1 could be used as a therapeutic target in HAdV-7 infection.

60 APPLIED LIFE SCIENCES↗