Journal of Inherited Metabolic Disease

, Volume 36, Issue 3, pp 479–490 | Cite as

Expression of the Nrf2-system at the blood-CSF barrier is modulated by neonatal inflammation and hypoxia-ischemia

  • Barbara D’Angelo
  • C. Joakim Ek
  • Mats Sandberg
  • Carina Mallard
Original Article


Transcription factor NF-E2-related factor-2 (Nrf2) is a key regulator of endogenous anti-oxidant systems shown to play a neuroprotective role in the adult by preserving blood–brain barrier function. The choroid plexus, site for the blood-CSF barrier, has been suggested to be particularly important in maintaining brain barrier function in development. We investigated the expression of Nrf2- and detoxification-system genes in choroid plexus following systemic LPS injections, unilateral cerebral hypoxia-ischemia (HI) as well as the combination of LPS and HI (LPS/HI). Plexuses were collected at different time points after LPS, HI and LPS/HI in 9-day old mice. mRNA levels of Nrf2 and many of its target genes were analyzed by quantitative PCR. Cell death was analyzed by caspase-3 immunostaining and TUNEL. LPS caused down-regulation of the Nrf2-system genes while HI increased expression at earlier time points. LPS exposure prior to HI prevented many of the HI-induced gene increases. None of the insults resulted in any apparent cell death to choroidal epithelium. These data imply that the function of the inducible anti-oxidant system in the choroid plexus is down-regulated by inflammation, even if choroid cells are not structurally damaged. Further, LPS prevented the endogenous antioxidant response following HI, suggesting the possibility that the choroid plexus may be at risk if LPS is united with an insult that increases oxidative stress such as hypoxia-ischemia.


  1. Beiswanger CM, Diegmann MH, Novak RF et al (1995) Developmental changes in the cellular distribution of glutathione and glutathione S-transferases in the murine nervous system. Neurotoxicol 16:425–440Google Scholar
  2. Borst P, Evers R, Kool M, Wijnholds J (2000) A family of drug transporters: the multidrug resistance-associated proteins. J Natl Cancer Inst 92:1295–1302PubMedCrossRefGoogle Scholar
  3. Calkins MJ, Vargas MR, Johnson DA, Johnson JA (2010) Astrocyte-specific overexpression of Nrf2 protects striatal neurons from mitochondrial complex II inhibition. Toxicol Sci 115:557–568PubMedCrossRefGoogle Scholar
  4. Chen PC, Vargas MR, Pani AK et al (2009) Nrf2-mediated neuroprotection in the MPTP mouse model of Parkinson’s disease: Critical role for the astrocyte. Proc Natl Acad Sci USA 106:2933–2938PubMedCrossRefGoogle Scholar
  5. Correa F, Ljunggren E, Mallard C, Nilsson M, Weber SG, Sandberg M (2011) The Nrf2-inducible antioxidant defense in astrocytes can be both up- and down-regulated by activated microglia: Involvement of p38 MAPK. Glia 59:785–799PubMedCrossRefGoogle Scholar
  6. Ek CJ, Habgood MD, Dziegielewska KM, Saunders NR (2003) Structural characteristics and barrier properties of the choroid plexuses in developing brain of the opossum (Monodelphis Domestica). J Comp Neurol 460:451–464PubMedCrossRefGoogle Scholar
  7. Ek CJ, Wong A, Liddelow SA, Johansson PA, Dziegielewska KM, Saunders NR (2010) Efflux mechanisms at the developing brain barriers: ABC-transporters in the fetal and postnatal rat. Toxicol Lett 197:51–59PubMedCrossRefGoogle Scholar
  8. Ek CJ, Dziegielewska KM, Habgood MD, Saunders NR (2012) Barriers in the developing brain and Neurotoxicology. Neurotoxicol 33:586–604CrossRefGoogle Scholar
  9. Eklind S, Mallard C, Leverin AL et al (2001) Bacterial endotoxin sensitizes the immature brain to hypoxic–ischaemic injury. Eur J Neurosci 13(6):1101–1106PubMedCrossRefGoogle Scholar
  10. Ferriero DM (2001) Oxidant mechanisms in neonatal hypoxia-ischemia. Dev Neurosci 23:198–202PubMedCrossRefGoogle Scholar
  11. Garbuzova-Davis S, Louis MK, Haller EM, Derasari HM, Rawls AE, Sanberg PR (2011) Blood–brain barrier impairment in an animal model of MPS III B. PLoS One 6:e16601. doi:10.1371/journal.pone.0016601 PubMedCrossRefGoogle Scholar
  12. Gazzin S, Strazielle N, Schmitt C et al (2008) Differential expression of the multidrug resistance-related proteins ABCb1 and ABCc1 between blood–brain interfaces. J Comp Neurol 510:497–507PubMedCrossRefGoogle Scholar
  13. Ghersi-Egea JF, Strazielle N, Murat A, Jouvet A, Buenerd A, Belin MF (2006) Brain protection at the blood-cerebrospinal fluid interface involves a glutathione-dependent metabolic barrier mechanism. J Cereb Blood Flow Metab 26:1165–1175PubMedGoogle Scholar
  14. Gulcan H, Ozturk IC, Arslan S (2005) Alterations in antioxidant enzyme activities in cerebrospinal fluid related with severity of hypoxic ischemic encephalopathy in newborns. Biol Neonate 88:87–91PubMedCrossRefGoogle Scholar
  15. Hedtjärn M, Leverin AL, Eriksson K, Blomgren K, Mallard C, Hagberg H (2002) Interleukin-18 involvement in hypoxic-ischemic brain injury. J Neurosci 22:5910–5919PubMedGoogle Scholar
  16. Huber JD, Campos CR, Mark KS, Davis TP (2006) Alterations in blood–brain barrier ICAM-1 expression and brain microglial activation after lambda-carrageenan-induced inflammatory pain. Am J Physiol Heart Circ Physiol 290:H732–740PubMedCrossRefGoogle Scholar
  17. Innamorato NG, Rojo AI, Garcia-Yague AJ, Yamamoto M, de Ceballos ML, Cuadrado A (2008) The transcription factor Nrf2 is a therapeutic target against brain inflammation. J Immunol 181:680–689PubMedGoogle Scholar
  18. Itoh S, Yanagishita T, Aoki S et al (1999) Generation of free radicals and the damage done to the sarcoplasmic reticulum during reperfusion injury following brief ischemia in the canine heart. Jpn Circ J 63:373–378PubMedCrossRefGoogle Scholar
  19. Johanson CE (1995) Ventricles and cerebrospinal fluid. In: Conn PM (ed) Neuroscience in medicine. J.B Lippincott Company, Philadelphia, pp 171–196Google Scholar
  20. Johanson CE, Jones HC, Stopa EG, Ayala C, Duncan JA, McMillan PN (2002) Enhanced expression of the NA-K-2 Cl cotransporter at different regions of the blood-CSF barrier in the perinatal H-Tx rat. Eur J Pediatr Surg 12:S47–49PubMedGoogle Scholar
  21. Johansson PA, Dziegielewska KM, Ek CJ et al (2006) Blood-CSF barrier function in the rat embryo. Eur J Neurosci 24:65–76PubMedCrossRefGoogle Scholar
  22. Johnson DA, Andrews GK, Xu W, Johnson JA (2002) Activation of the antioxidant response element in primary cortical neuronal cultures derived from transgenic reporter mice. J Neurochem 81:1233–1241PubMedCrossRefGoogle Scholar
  23. Kauffmann HM, Pfannschmidt S, Zoller H et al (2002) Influence of redox-active compounds and PXR-activators on human MRP1 and MRP2 gene expression. Toxicology 171:137–146PubMedCrossRefGoogle Scholar
  24. Klaassen CD, Slitt A (2005) Regulation of hepatic transporters by xenobiotic receptors. Curr Drug Metab 6:309–328PubMedCrossRefGoogle Scholar
  25. Ko K, Yang H, Noureddin M et al (2008) Changes in S-adenosylmethionine and GSH homeostasis during endotoxemia in mice. Lab Invest 88:1121–1129PubMedCrossRefGoogle Scholar
  26. Kwak MK, Wakabayashi N, Itoh K, Motohashi H, Yamamoto M, Kensler TW (2003) Modulation of gene expression by cancer chemopreventive dithiolethiones through the Keap1–Nrf2 pathway. Identification of novel gene clusters for cell survival. J Biol Chem 278:8135–8145PubMedCrossRefGoogle Scholar
  27. Laflamme N, Rivest S (2001) Toll-like receptor 4: the missing link of the cerebral innate immune response triggered by circulating gram-negative bacterial cell wall components. FASEB J 15:155–63PubMedCrossRefGoogle Scholar
  28. Lee JM, Shih AY, Murphy TH, Johnson JA (2003) NF-E2-related factor-2 mediates neuroprotection against mitochondrial complex I inhibitors and increased concentrations of intracellular calcium in primary cortical neurons. J Biol Chem 278:37948–37956PubMedCrossRefGoogle Scholar
  29. Liddelow SA, Temple S, Møllgård K et al (2012) Molecular characterisation of transport mechanisms at the developing mouse blood-CSF interface: a transcriptome approach. PLoS One 7:e33554. doi:10.1371/journal.pone.0033554 PubMedCrossRefGoogle Scholar
  30. Maeda S, Nakatsuka I, Hayashi Y, Higuchi H, Shimada M, Miyawaki T (2008) Heme oxygenase-1 induction in the brain during lipopolysaccharide-induced acute inflammation. Neuropsychiatr Dis Treat 4:663–667PubMedCrossRefGoogle Scholar
  31. Marques F, Sousa JC, Coppola G et al (2009) Kinetic profile of the transcriptome changes induced in the choroid plexus by peripheral inflammation. J Cereb Blood Flow Metab 29:921–932PubMedCrossRefGoogle Scholar
  32. Meijerman I, Beijnen JH, Schellens JH (2008) Combined action and regulation of phase II enzymes and multidrug resistance proteins in multidrug resistance in cancer. Cancer Treat Rev 34:505–520PubMedCrossRefGoogle Scholar
  33. Nagai N, Thimmulappa RK, Cano M et al (2009) Nrf2 is a critical modulator of the innate immune response in a model of uveitis. Free Radic Biol Med 47:300–306PubMedCrossRefGoogle Scholar
  34. Ogihara T, Hirano K, Ogihara H et al (2003) Non-protein-bound transition metals and hydroxyl radical generation in cerebrospinal fluid of newborn infants with hypoxic ischemic encephalopathy. Pediatr Res 53:594–599PubMedCrossRefGoogle Scholar
  35. Ping Z, Liu W, Kang Z et al (2010) Sulforaphane protects brains against hypoxic-ischemic injury through induction of Nrf2-dependent phase 2 enzyme. Brain Res 1343:178–185PubMedCrossRefGoogle Scholar
  36. Rothstein RP, Levison SW (2002) Damage to the choroid plexus, ependyma and subependyma as a consequence of perinatal hypoxia/ischemia. Dev Neurosci 24:426–436PubMedCrossRefGoogle Scholar
  37. Saha A, Sarkar C, Singh SP et al (2012) The blood–brain barrier is disrupted in a mouse model of infantile neuronal ceroid lipofuscinosis: amelioration by resveratrol. Hum Mol Genet 21:2233–2244PubMedCrossRefGoogle Scholar
  38. Sävman K, Nilsson UA, Blennow M, Kjellmer I, Whitelaw A (2001) Non-protein-bound iron is elevated in cerebrospinal fluid from preterm infants with posthemorrhagic ventricular dilatation. Pediatr Res 49:208–212PubMedCrossRefGoogle Scholar
  39. Schipper HM, Song W, Zukor H, Hascalovici JR, Zeligman D (2009) Heme oxygenase-1 and neurodegeneration: expanding frontiers of engagement. J Neurochem 110:469–485PubMedCrossRefGoogle Scholar
  40. Senjo M, Ishibashi T, Terashima T, Inoue Y (1986) Successive appearance of glutathione S-transferase-positive cells in developing rat brain: choroid plexus, pia mater, ventricular zone and astrocytes. Neurosci Lett 66:131–134PubMedCrossRefGoogle Scholar
  41. Shih AY, Imbeault S, Barakauskas V, Erb H, Jiang L, Li P, Murphy TH (2005) Induction of the Nrf2-driven antioxidant response confers neuroprotection during mitochondrial stress in vivo. J Biol Chem 280:22925–22936PubMedCrossRefGoogle Scholar
  42. Strazielle N, Ghersi-Egea JF (2000) Choroid plexus in the central nervous system: biology and physiopathology. J Neuropathol Exp Neurol 59:561–574PubMedGoogle Scholar
  43. Strazielle N, Khuth ST, Ghersi-Egea JF (2004) Detoxification systems, passive and specific transport for drugs at the blood-CSF barrier in normal and pathological situations. Adv Drug Deliv Rev 56:1717–1740PubMedCrossRefGoogle Scholar
  44. Sun Y, Liou B, Ran H et al (2010) Neuronopathic Gaucher disease in the mouse: viable combined selective saposin C deficiency and mutant glucocerebrosidase (V394L) mice with glucosylsphingosine and glucosylceramide accumulation and progressive neurological deficits. Hum Mol Genet 19:1088–1097PubMedCrossRefGoogle Scholar
  45. Svedin P, Hagberg H, Sävman K, Zhu C, Mallard C (2007) Matrix metalloproteinase-9 gene knock-out protects the immature brain after cerebral hypoxia-ischemia. J Neurosci 27:1511–1518PubMedCrossRefGoogle Scholar
  46. Tirona RG, Kim RB (2005) Nuclear receptors and drug disposition gene regulation. J Pharm Sci 94:1169–1186PubMedCrossRefGoogle Scholar
  47. Vargas MR, Johnson DA, Sirkis DW, Messing A, Johnson JA (2008) Nrf2 activation in astrocytes protects against neurodegeneration in mouse models of familial amyotrophic lateral sclerosis. J Neurosci 28:13574–13581PubMedCrossRefGoogle Scholar
  48. Wang X, Svedin P, Nie C et al (2007) N-acetylcysteine reduces lipopolysaccharide-sensitized hypoxic-ischemic brain injury. Ann Neurol 61:263–271PubMedCrossRefGoogle Scholar
  49. Wang X, Stridh L, Li W et al (2009) Lipopolysaccharide sensitizes neonatal hypoxic-ischemic brain injury in a MyD88-dependent manner. J Immunol 183:7471–7477PubMedCrossRefGoogle Scholar
  50. Wasserman WW, Fahl WE (1997) Functional antioxidant responsive elements. Proc Natl Acad Sci USA 94:5361–5266PubMedCrossRefGoogle Scholar
  51. Wiesel P, Foster LC, Pellacani A et al (2000) Thioredoxin facilitates the induction of heme oxygenase-1 in response to inflammatory mediators. J Biol Chem 275:24840–24846PubMedCrossRefGoogle Scholar
  52. Wu YP, Proia RL (2004) Deletion of macrophage-inflammatory protein 1 alpha retards neurodegeneration in Sandhoff disease mice. Proc Natl Acad Sci USA 101:8425–8430PubMedCrossRefGoogle Scholar
  53. Xiang J, Alesi GN, Zhou N, Keep RF (2012) Protective effects of isothiocyanates on blood-CSF barrier disruption induced by oxidative stress. Am J Physiol Regul Integr Comp Physiol 303:R1–7. doi:10.1152/ajpregu.00518.2011 PubMedCrossRefGoogle Scholar
  54. Zhao J, Moore AN, Redell JB, Dash PK (2007) Enhancing expression of Nrf2-driven genes protects the blood brain barrier after brain injury. J Neurosci 27:10240–10248PubMedCrossRefGoogle Scholar

Copyright information

© SSIEM and Springer Science+Business Media Dordrecht 2012

Authors and Affiliations

  • Barbara D’Angelo
    • 1
  • C. Joakim Ek
    • 1
  • Mats Sandberg
    • 2
  • Carina Mallard
    • 1
  1. 1.Department of Neuroscience and Physiology, Sahlgrenska AcademyUniversity of GothenburgGothenburgSweden
  2. 2.Institute of Biomedicine, Sahlgrenska AcademyUniversity of GothenburgGothenburgSweden

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