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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 1,693 records · Page 94
All sourcesNASA NTRSDOE OSTI
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↗
DOE OSTI2018

Role of acid responsive genes in the susceptibility of Escherichia coli to ciclopirox

Highlights: • Ciclopirox antibacterial activity was analysed by genome-wide expression profiling. • Ciclopirox affects expression of glutamate-dependent acid resistance pathway genes. • Down-regulation of hns transcription likely increases ciclopirox activity. • Susceptibility to ciclopirox accompanies increased cell size and decreased motility. • The findings identify probable pathways for ciclopirox action against E. coli. Antibiotic resistance poses a huge threat to the effective treatment of bacterial infections. To circumvent the limitations in developing new antibiotics, researchers are attempting to repurpose pre-developed drugs that are known to be safe. Ciclopirox, an off-patent antifungal agent, inhibits the growth of Gram-negative bacteria, and genes involved in galactose metabolism and lipopolysaccharide (LPS) biosynthesis are plausible antibacterial targets for ciclopirox, since their expression levels partially increase susceptibility at restrictive concentrations. In the present study, to identify new target genes involved in the susceptibility of Escherichia coli to ciclopirox, genome-wide mRNA profiling was performed following ciclopirox addition at sublethal concentrations, and glutamate-dependent acid resistance (GDAR) genes were differentially regulated. Additional susceptibility testing, growth analyses and viability assays of GDAR regulatory genes revealed that down-regulation of evgS or hns strongly enhanced susceptibility to ciclopirox. Further microscopy and phenotypic analyses revealed that down-regulation of these genes increased cell size and decreased motility. Our findings could help to maximise the efficacy of ciclopirox against hard-to-treat Gram-negative pathogens.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

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↗
DOE OSTI2018

Biophysical characterization of actin bundles generated by the Chlamydia trachomatis Tarp effector

Highlights: • The F-actin binding domain of Tarp is sufficient to bundle filamentous actin. • Actin bundles formed by Tarp are more flexible than that of α-actinin and fascin. • Tarp mediated actin bundling is independent of actin nucleation. Chlamydia trachomatis entry into host cells is mediated by pathogen-directed remodeling of the actin cytoskeleton. The chlamydial type III secreted effector, translocated actin recruiting phosphoprotein (Tarp), has been implicated in the recruitment of actin to the site of internalization. Tarp harbors G-actin binding and proline rich domains required for Tarp-mediated actin nucleation as well as unique F-actin binding domains implicated in the formation of actin bundles. Little is known about the mechanical properties of actin bundles generated by Tarp or the mechanism by which Tarp mediates actin bundle formation. In order to characterize the actin bundles and elucidate the role of different Tarp domains in the bundling process, purified Tarp effectors and Tarp truncation mutants were analyzed using Total Internal Reflection Fluorescence (TIRF) microscopy. Our data indicate that Tarp mediated actin bundling is independent of actin nucleation and the F-actin binding domains are sufficient to bundle actin filaments. Additionally, Tarp-mediated actin bundles demonstrate distinct bending stiffness compared to those crosslinked by the well characterized actin bundling proteins fascin and alpha-actinin, suggesting Tarp may employ a novel actin bundling strategy. The capacity of the Tarp effector to generate novel actin bundles likely contributes to chlamydia's efficient mechanism of entry into human cells.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Long noncoding RNA HAGLROS regulates cell apoptosis and autophagy in lipopolysaccharides-induced WI-38 cells via modulating miR-100/NF-κB axis

Highlights: • LncRNA HAGLROS was higher expressed in patients with pneumonia. • LPS caused WI-38 cell injury and increased HAGLROS levels. • Downregulation of HAGLROS alleviated LPS-induced WI-38 cell injury by miR-100. • NFΚB3 was verified as a functional target of miR-100. • Knockdown of HAGLROS inhibited the activation of PI3K/AKT/NF-κB pathway. Pneumonia is a lower respiratory disease caused by pathogens or other factors. This study aimed to explore the roles and mechanism of long noncoding RNA HAGLROS in lipopolysaccharides (LPS)-induced inflammatory injury in pneumonia. The HAGLROS expression in serum of patients with acute stage pneumonia was detected. To induce pulmonary injury, WI-38 human lung fibroblasts were stimulated with lipopolysaccharides (LPS). The HAGLROS expressions in LPS-treated WI-38 cells and the effects of HAGLROS knockdown on the viability, apoptosis, and autophagy of LPS-induced cells were detected. Moreover, the regulatory relationship between HAGLROS and miR-100 was explored as well as the functional targets of miR-100 were identified. Furthermore, the regulatory relationship between miR-100 and PI3K/AKT/NF-κB pathway was elucidated. LncRNA HAGLROS was higher expressed in serum of patients with acute stage pneumonia compared with that in serum of healthy control. LPS caused WI-38 cell injury and increased HAGLROS levels. Downregulation of HAGLROS alleviated LPS-induced cell injury via increasing cell viability, and inhibiting apoptosis and autophagy. Moreover, there was a negative correlation between HAGLROS and miR-100, and the effects of HAGLROS downregulation on LPS-induced apoptosis and autophagy in WI-38 cells were by regulation of miR-100. Furthermore, NFΚB3 was verified as a functional target of miR-100 and effects of miR-100 inhibition on LPS-induced WI-38 cell injury were alleviated by knockdown of NFΚB3. Besides, Knockdown of HAGLROS inhibited the activation of PI3K/AKT/NF-κB pathway. Our findings reveal that downregulation of HAGLROS may alleviate LPS-induced inflammatory injury in WI-38 cells via modulating miR-100/NF-κB axis. HAGLROS/miR-100/NF-κB axis may provide a new strategy for treating acute stage of pneumonia.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Matrix stiffness regulate apoptotic cell death in HIV-HCV co-infected hepatocytes: Importance for liver fibrosis progression

Highlights: • HIV-HCV co-infection increases HIV/HCV RNAs levels and apoptosis in hepatocytes. • Fibrotic matrix stiffness enhances HIV/HCV RNAs and apoptosis in hepatocytes. • Apoptotic infected hepatocytes induce pro-fibrotic changes in hepatic stellate cells. HIV-HCV co-infection causes rapid progression of liver fibrosis. These outcomes to liver cirrhosis can be improved, but not stopped by specific antiviral therapies. Due to high significance of HIV-HCV interactions for morbidity and mortality in co-infected patients, our attention was attracted to the multi-component pathogenesis of fibrosis progression as the transition to end-stage liver disease development. In this study, we hypothesize that increased matrix stiffness enhances apoptosis in HCV-HIV-co-infected hepatocytes and that capturing of apoptotic bodies (AB) derived from these infected hepatocytes by hepatic stellate cells (HSC) drives the fibrosis progression. As the source of viruses, JFH1 (HCV genotype 2a) and HIV-1{sub ADA} (either purified or containing in infected macrophage supernatants) were chosen. Using Huh7.5-CYP (RLW) cells and primary human hepatocytes mono-infected with HCV and HIV or co-infected, we have shown that both HCV and HIV RNA levels were increased in co-infected cells, which was accompanied by hepatocyte apoptosis. This apoptosis was attenuated by azidothymidine treatment. The levels of both infections and apoptosis were more prominent in primary hepatocytes cultured on substrates mimicking fibrotic stiffness (24 kPa-stiff) compared to substrates mimicking healthy liver (2.4 kPa-soft). The engulfment of AB from pathogen-exposed hepatocytes activated pro-fibrotic mRNAs in HSC. Overall, the increased matrix stiffness is not only a consequence of liver inflammation/fibrosis, but the condition that further accelerates liver fibrosis development. This is attributed to the switching of HSC to pro-fibrotic phenotype by capturing of excessive amounts of apoptotic HCV- and HIV-infected hepatocytes.

60 APPLIED LIFE SCIENCES↗