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  • Loss of MPC1 reprograms retinal metabolism to impair visual function PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Grenell, A., Wang, Y., Yam, M., Swarup, A., Dilan, T. L., Hauer, A., Linton, J. D., Philp, N. J., Gregor, E., Zhu, S., Shi, Q., Murphy, J., Guan, T., Lohner, D., Kolandaivelu, S., Ramamurthy, V., Goldberg, A. X., Hurley, J. B., Du, J. 2019; 116 (9): 3530–35

    Abstract

    Glucose metabolism in vertebrate retinas is dominated by aerobic glycolysis (the "Warburg Effect"), which allows only a small fraction of glucose-derived pyruvate to enter mitochondria. Here, we report evidence that the small fraction of pyruvate in photoreceptors that does get oxidized by their mitochondria is required for visual function, photoreceptor structure and viability, normal neuron-glial interaction, and homeostasis of retinal metabolism. The mitochondrial pyruvate carrier (MPC) links glycolysis and mitochondrial metabolism. Retina-specific deletion of MPC1 results in progressive retinal degeneration and decline of visual function in both rod and cone photoreceptors. Using targeted-metabolomics and 13C tracers, we found that MPC1 is required for cytosolic reducing power maintenance, glutamine/glutamate metabolism, and flexibility in fuel utilization.

    View details for PubMedID 30808746

  • Modulating GLUT1 expression in retinal pigment epithelium decreases glucose levels in the retina: impact on photoreceptors and Muller glial cells AMERICAN JOURNAL OF PHYSIOLOGY-CELL PHYSIOLOGY Swarup, A., Samuels, I. S., Bell, B. A., Han, J. S., Du, J., Massenzio, E., Abel, E., Boesze-Battaglia, K., Peachey, N. S., Philp, N. J. 2019; 316 (1): C121–C133

    Abstract

    The retina is one of the most metabolically active tissues in the body and utilizes glucose to produce energy and intermediates required for daily renewal of photoreceptor cell outer segments. Glucose transporter 1 (GLUT1) facilitates glucose transport across outer blood retinal barrier (BRB) formed by the retinal pigment epithelium (RPE) and the inner BRB formed by the endothelium. We used conditional knockout mice to study the impact of reducing glucose transport across the RPE on photoreceptor and Müller glial cells. Transgenic mice expressing Cre recombinase under control of the Bestrophin1 ( Best1) promoter were bred with Glut1flox/flox mice to generate Tg-Best1-Cre:Glut1flox/flox mice ( RPEΔGlut1). The RPEΔGlut1 mice displayed a mosaic pattern of Cre expression within the RPE that allowed us to analyze mice with ~50% ( RPEΔGlut1m) recombination and mice with >70% ( RPEΔGlut1h) recombination separately. Deletion of GLUT1 from the RPE did not affect its carrier or barrier functions, indicating that the RPE utilizes other substrates to support its metabolic needs thereby sparing glucose for the outer retina. RPEΔGlut1m mice had normal retinal morphology, function, and no cell death; however, where GLUT1 was absent from a span of RPE greater than 100 µm, there was shortening of the photoreceptor cell outer segments. RPEΔGlut1h mice showed outer segment shortening, cell death of photoreceptors, and activation of Müller glial cells. The severe phenotype seen in RPEΔGlut1h mice indicates that glucose transport via the GLUT1 transporter in the RPE is required to meet the anabolic and catabolic requirements of photoreceptors and maintain Müller glial cells in a quiescent state.

    View details for PubMedID 30462537

    View details for PubMedCentralID PMC6383144

  • Modulation of GLUT1 expression in the RPE impacts outer segment renewal and results in photoreceptor degeneration Swarup, A., Samuels, I. S., Soto, J., Abel, E., Peachey, N., Philp, N. J. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2018
  • Deletion of GLUT1 in mouse lens epithelium leads to cataract formation EXPERIMENTAL EYE RESEARCH Swarup, A., Bell, B. A., Du, J., Han, J. S., Soto, J., Abel, E., Bravo-Nuevo, A., FitzGerald, P. G., Peachey, N. S., Philp, N. J. 2018; 172: 45–53

    Abstract

    The primary energy substrate of the lens is glucose and uptake of glucose from the aqueous humor is dependent on glucose transporters. GLUT1, the facilitated glucose transporter encoded by Slc2a1 is expressed in the epithelium of bovine, human and rat lenses. In the current study, we examined the expression of GLUT1 in the mouse lens and determined its role in maintaining lens transparency by studying effects of postnatal deletion of Slc2a1. In situ hybridization and immunofluorescence labeling were used to determine the expression and subcellular distribution of GLUT1 in the lens. Slc2a1 was knocked out of the lens epithelium by crossing transgenic mice expressing Cre recombinase under control of the GFAP promoter with Slc2a1loxP/loxP mice to generate Slc2a1loxP/loxP;GFAP-Cre+/0 (LensΔGlut1) mice. LensΔGlut1 mice developed visible lens opacities by around 3 months of age, which corresponded temporally with the total loss of detectable GLUT1 expression in the lens. Spectral domain optical coherence tomography (SD-OCT) imaging was used to monitor the formation of cataracts over time. SD-OCT imaging revealed that small nuclear cataracts were first apparent in the lenses of LensΔGlut1 mice beginning at about 2.7 months of age. Longitudinal SD-OCT imaging of LensΔGlut1 mice revealed disruption of mature secondary fiber cells after 3 months of age. Histological sections of eyes from LensΔGlut1 mice confirmed the disruption of the secondary fiber cells. The structural changes were most pronounced in fiber cells that had lost their organelles. In contrast, the histology of the lens epithelium in these mice appeared normal. Lactate and ATP were measured in lenses from LensΔGlut1 and control mice at 2 and 3 months of age. At 2 months of age, when GLUT1 was still detectable in the lens epithelium, albeit at low levels, the amount of lactate and ATP were not significantly different from controls. However, in lenses isolated from 3-month-old LensΔGlut1 mice, when GLUT1 was no longer detectable, levels of lactate and ATP were 50% lower than controls. Our findings demonstrate that in vivo, the transparency of mature lens fiber cells was dependent on glycolysis for ATP and the loss of GLUT1 transporters led to cataract formation. In contrast, lens epithelium and cortical fiber cells have mitochondria and could utilize other substrates to support their anabolic and catabolic needs.

    View details for PubMedID 29604281

  • Loss of GLUT1 in the retinal pigment epithelium does not diminish its function, differentiation or polarity Samuels, I. S., Swarup, A., Beight, C., Han, J. S., Soto, J., Abel, E., Peachey, N. S., Philp, N. J. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2017
  • Microtubule-associated protein 1 light chain 3 (LC3) isoforms in RPE: expression and function Dhingra, A., Reyes-Reveles, J., Alexander, D., Sharp, R. C., Swarup, A., Kim, H., Sparrow, J. R., Boesze-Battaglia, K. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2017
  • Deletion of GLUT1 in mouse lens epithelium leads to cataract formation Swarup, A., Bravo-Nuevo, A., Soto, J., Abel, E., Bell, B. A., Peachey, N., FitzGerald, P. G., Philp, N. J. ASSOC RESEARCH VISION OPHTHALMOLOGY INC. 2017
  • Inhibition of miR-21 rescues liver regeneration after partial hepatectomy in ethanol-fed rats AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY Juskeviciute, E., Dippold, R. P., Antony, A. N., Swarup, A., Vadigepalli, R., Hoek, J. B. 2016; 311 (5): G794–G806

    Abstract

    Liver regeneration is a clinically significant tissue repair process that is suppressed by chronic alcohol intake through poorly understood mechanisms. Recently, microRNA-21 (miR-21) has been suggested to serve as a crucial microRNA (miRNA) regulator driving hepatocyte proliferation after partial hepatectomy (PHx) in mice. However, we reported recently that miR-21 is significantly upregulated in ethanol-fed rats 24 h after PHx, despite inhibition of cell proliferation, suggesting a more complex role for this miRNA. Here, we investigate how inhibition of miR-21 in vivo affects the early phase of liver regeneration in ethanol-fed rats. Chronically ethanol-fed rats and pair-fed control animals were treated with AM21, a mixed locked nucleic acid-DNA analog antisense to miR-21 that inhibited miR-21 in vivo to undetectable levels. Liver regeneration after PHx was followed by cell proliferation marker and gene expression analysis, miRNA profiling, and cell signaling pathway analysis. Although liver regeneration was not significantly impaired by AM21 in chow-fed rats, AM21 treatment in ethanol-fed animals completely restored regeneration and enhanced PHx-induced hepatocyte proliferation to levels comparable to those of untreated or chow-fed animals. In addition, a marked deposition of α-smooth muscle actin, a marker of stellate cell activation, which was evident in ethanol-treated animals after PHx, was effectively suppressed by AM21 treatment. Gene expression analysis further indicated that suppression of stellate cell-specific profibrogenic profiles and the Notch signaling contributed to AM21-mediated rescue from deficient hepatocyte proliferation in ethanol-fed animals. Our results indicate that the impact of miR-21 balances proproliferative effects with antiproliferative profibrogenic actions in regulating distinctive regenerative responses in normal vs. disease conditions.

    View details for PubMedID 27634014

    View details for PubMedCentralID PMC5130549

  • Adiponectin fine-tuning of liver regeneration dynamics revealed through cellular network modelling JOURNAL OF PHYSIOLOGY-LONDON Correnti, J. M., Cook, D., Aksamitiene, E., Swarup, A., Ogunnaike, B., Vadigepalli, R., Hoek, J. B. 2015; 593 (2): 365–83

    Abstract

    Following partial hepatectomy, the liver initiates a regenerative programme involving hepatocyte priming and replication driven by the coordinated actions of cytokine and growth factors. We investigated the mechanisms underlying adiponectin's (Adn) regulation of liver regeneration through modulation of these mediators. Adn(-/-) mice showed delayed onset of hepatocyte replication, but accelerated cell cycle progression relative to wild-type mice, suggesting Adn has multiple effects fine-tuning the kinetics of liver regeneration. We developed a computational model describing the molecular and physiological kinetics of liver regeneration in Adn(-/-) mice. We employed this computational model to evaluate the underlying regulatory mechanisms. Our analysis predicted that Adn is required for an efficient early cytokine response to partial hepatectomy, but is inhibitory to later growth factor actions. Consistent with this prediction, Adn knockout reduced hepatocyte responses to interleukin-6 during the priming phase, but enhanced growth factor levels through peak hepatocyte replication. By contrast, supraphysiological concentrations of Adn resulting from rosiglitazone treatment suppressed regeneration by reducing growth factor levels during S phase, consistent with computational predictions. Together, these results revealed that Adn fine-tunes the progression of liver regeneration through dynamically modulating molecular mediator networks and cellular interactions within the liver.

    View details for PubMedID 25630259

    View details for PubMedCentralID PMC4303383

  • Adiponectin fine-tuning of liver regeneration dynamics revealed through cellular network modeling. The Journal of physiology Correnti, J. M., Cook, D., Aksamitiene, E., Swarup, A., Ogunnaike, B., Vadigepalli, R., Hoek, J. B. 2014

    Abstract

    Following partial hepatectomy, the liver initiates a regenerative program involving hepatocyte priming and replication driven by coordinated cytokine and growth factor actions. We investigated the mechanisms underlying Adiponectin's (Adn) regulation of liver regeneration through modulation of these mediators. Adn-/- mice showed delayed onset of hepatocyte replication, but accelerated cell cycle progression relative to wild-type mice, suggesting Adn has multiple effects fine-tuning the kinetics of liver regeneration. We developed a computational model describing the molecular and physiological kinetics of liver regeneration in Adn-/- mice. We employed this computational model to evaluate the underlying regulatory mechanisms. Our analysis predicted that Adn is required for an efficient early cytokine response to partial hepatectomy, but is inhibitory to later growth factor actions. Consistent with this prediction, Adn knockout reduced hepatocyte responses to IL-6 during the priming phase, but enhanced growth factor levels through peak hepatocyte replication. By contrast, supraphysiological concentrations of Adn resulting from rosiglitazone treatment suppressed regeneration by reducing growth factor levels during S phase, consistent with computational predictions. Together, these results revealed that Adn fine-tunes the progression of liver regeneration through dynamically modulating molecular mediator networks and cellular interactions within the liver. This article is protected by copyright. All rights reserved.

    View details for PubMedID 25384784

  • Metabolic network reconstruction, growth characterization and C-13-metabolic flux analysis of the extremophile Thermus thermophilus HB8 METABOLIC ENGINEERING Swarup, A., Lu, J., DeWoody, K. C., Antoniewicz, M. R. 2014; 24: 173–80

    Abstract

    Thermus thermophilus is an extremely thermophilic bacterium with significant biotechnological potential. In this work, we have characterized aerobic growth characteristics of T. thermophilus HB8 at temperatures between 50 and 85°C, constructed a metabolic network model of its central carbon metabolism and validated the model using (13)C-metabolic flux analysis ((13)C-MFA). First, cells were grown in batch cultures in custom constructed mini-bioreactors at different temperatures to determine optimal growth conditions. The optimal temperature for T. thermophilus grown on defined medium with glucose was 81°C. The maximum growth rate was 0.25h(-1). Between 50 and 81°C the growth rate increased by 7-fold and the temperature dependence was described well by an Arrhenius model with an activation energy of 47kJ/mol. Next, we performed a (13)C-labeling experiment with [1,2-(13)C] glucose as the tracer and calculated intracellular metabolic fluxes using (13)C-MFA. The results provided support for the constructed network model and highlighted several interesting characteristics of T. thermophilus metabolism. We found that T. thermophilus largely uses glycolysis and TCA cycle to produce biosynthetic precursors, ATP and reducing equivalents needed for cells growth. Consistent with its proposed metabolic network model, we did not detect any oxidative pentose phosphate pathway flux or Entner-Doudoroff pathway activity. The biomass precursors erythrose-4-phosphate and ribose-5-phosphate were produced via the non-oxidative pentose phosphate pathway, and largely via transketolase, with little contribution from transaldolase. The high biomass yield on glucose that was measured experimentally was also confirmed independently by (13)C-MFA. The results presented here provide a solid foundation for future studies of T. thermophilus and its metabolic engineering applications.

    View details for PubMedID 24909362

  • Pharmacological ceramide reduction alleviates alcohol-induced steatosis and hepatomegaly in adiponectin knockout mice AMERICAN JOURNAL OF PHYSIOLOGY-GASTROINTESTINAL AND LIVER PHYSIOLOGY Correnti, J. M., Juskeviciute, E., Swarup, A., Hoek, J. B. 2014; 306 (11): G959–G973

    Abstract

    Hepatosteatosis, the ectopic accumulation of lipid in the liver, is one of the earliest clinical signs of alcoholic liver disease (ALD). Alcohol-dependent deregulation of liver ceramide levels as well as inhibition of AMP-activated protein kinase (AMPK) and peroxisome proliferator-activated receptor α (PPAR-α) activity are thought to contribute to hepatosteatosis development. Adiponectin can regulate lipid handling in the liver and has been shown to reduce ceramide levels and activate AMPK and PPAR-α. However, the mechanisms by which adiponectin prevents alcoholic hepatosteatosis remain incompletely characterized. To address this question, we assessed ALD progression in wild-type (WT) and adiponectin knockout (KO) mice fed an ethanol-containing liquid diet or isocaloric control diet. Adiponectin KO mice relative to WT had increased alcohol-induced hepatosteatosis and hepatomegaly, similar modest increases in serum alanine aminotransferase, and reduced liver TNF. Restoring circulating adiponectin levels using recombinant adiponectin ameliorated alcohol-induced hepatosteatosis and hepatomegaly in adiponectin KO mice. Alcohol-fed WT and adiponectin KO animals had equivalent reductions in AMPK protein and PPAR-α DNA binding activity compared with control-fed animals. No difference in P-AMPK/AMPK ratio was detected, suggesting that alcohol-dependent deregulation of AMPK and PPAR-α in the absence of adiponectin are not primary causes of the observed increase in hepatosteatosis in these animals. By contrast, alcohol treatment increased liver ceramide levels in adiponectin KO but not WT mice. Importantly, pharmacological inhibition of de novo ceramide synthesis in adiponectin KO mice abrogated alcohol-mediated increases in liver ceramides, steatosis, and hepatomegaly. These data suggest that adiponectin reduces alcohol-induced steatosis and hepatomegaly through regulation of liver ceramides, but its absence does not exacerbate alcohol-induced liver damage.

    View details for PubMedID 24742988

    View details for PubMedCentralID PMC4042116

  • miR-21 inhibition overcomes ethanol suppression of rat liver regeneration Juskeviciute, E., Dippold, R. P., Swarup, A., Hoek, J. B. FEDERATION AMER SOC EXP BIOL. 2013
  • Substrate Oxidation Control of Respiratory Rates in Primary Hepatocytes Anil Noronha Antony Antony, A., Moffat, C., Swarup, A., Seifert, E., Hoek, J. B. CELL PRESS. 2013: 302A
  • Thermus thermophilus: A model thermophilic organism for biofuels production Lu, J., Swarup, A., DeWoody, K., Antoniewicz, M. R. AMER CHEMICAL SOC. 2012
  • Metabolic network reconstruction and C-13-metabolic flux analysis for the extremophile Thermus thermophilus HB8 Antoniewicz, M. R., Swarup, A., Dewoody, K. AMER CHEMICAL SOC. 2011