Acta Neuropathologica

, Volume 126, Issue 2, pp 207–218 | Cite as

mTOR-dependent abnormalities in autophagy characterize human malformations of cortical development: evidence from focal cortical dysplasia and tuberous sclerosis

  • Shireena A. Yasin
  • Abu M. Ali
  • Mathew Tata
  • Simon R. Picker
  • Glenn W. Anderson
  • Elizabeth Latimer-Bowman
  • Sarah L. Nicholson
  • William Harkness
  • J. Helen Cross
  • Simon M. L. Paine
  • Thomas S. JacquesEmail author
Original Paper


Focal cortical dysplasia (FCD) is a localized malformation of cortical development and is the commonest cause of severe childhood epilepsy in surgical practice. Children with FCD are severely disabled by their epilepsy, presenting with frequent seizures early in life. The commonest form of FCD in children is characterized by the presence of an abnormal population of cells, known as balloon cells. Similar pathological changes are seen in the cortical malformations that characterize patients with tuberous sclerosis complex (TSC). However, the cellular and molecular mechanisms that underlie the malformations of FCD and TSC are not well understood. We provide evidence for a defect in autophagy in FCD and TSC. We have found that balloon cells contain vacuoles that include components of the autophagy pathway. Specifically, we show that balloon cells contain prominent lysosomes by electron microscopy, immunohistochemistry for LAMP1 and LAMP2, LysoTracker labelling and enzyme histochemistry for acid phosphatase. Furthermore, we found that balloon cells contain components of the ATG pathway and that there is cytoplasmic accumulation of the regulator of autophagy, DOR. Most importantly we found that there is abnormal accumulation of the autophagy cargo protein, p62. We show that this defect in autophagy can be, in part, reversed in vitro by inhibition of the mammalian target of rapamycin (mTOR) suggesting that abnormal activation of mTOR may contribute directly to a defect in autophagy in FCD and TSC.


Autophagy Epilepsy Balloon cells Focal cortical dysplasia Tuberous sclerosis 



TSJ was in receipt of funding from the Great Ormond Street Hospital Children’s Charity and holds a HEFCE Clinical Senior Lecturer Award. This report is independent research supported by the National Institute for Health Research Great Ormond Street Hospital Biomedical Research Centre. The views expressed in this publication are those of the author(s) and not necessarily those of the NHS, the National Institute for Health Research or the Department of Health. We are grateful to Prof. Zorzano, Institute of Research in Biomedicine, Barcelona for the DOR antibody.

Supplementary material

Supplementary video 1 LysoTracker™ labelling of cultured balloon cells showed that they contained abundant lysosomes, which were dynamic in culture. Lysosomes were particularly concentrated close to the nucleus and were dynamic in the periphery of cells (MP4 152 kb)

401_2013_1135_MOESM2_ESM.tiff (650 kb)
Supplementary figure 2 Balloon cells show down-regulation of pS6 and p62 in response to rapamycin (a–f). Immunofluorescence for pS6 (a and d) and p62 (b and e) in control media (upper row) and rapamycin (lower row, 500 nM) (Scale bar = 30 μm). Balloon cells are indicated by arrows in the rapamycin condition (TIFF 650 kb)
401_2013_1135_MOESM3_ESM.doc (27 kb)
Supplementary table 3 The table summarizes the clinical features of the cohort of patients used in the study (DOC 27 kb)


  1. 1.
    Baek S-H, Kim E-K, Goudreau JL, Lookingland KJ, Kim SW, Yu S-W (2009) Insulin withdrawal-induced cell death in adult hippocampal neural stem cells as a model of autophagic cell death. Autophagy 5(2):277–279PubMedCrossRefGoogle Scholar
  2. 2.
    Baumgartner BG, Orpinell M, Duran J et al (2007) Identification of a novel modulator of thyroid hormone receptor-mediated action. PLoS ONE 2(11):e1183PubMedCrossRefGoogle Scholar
  3. 3.
    Baybis M, Yu J, Lee A, Golden JA, Weiner H, Mckhann G, Aronica E, Crino PB (2004) mTOR cascade activation distinguishes tubers from focal cortical dysplasia. Ann Neurol 56(4):478–487PubMedCrossRefGoogle Scholar
  4. 4.
    Becker AJ, Urbach H, Scheffler B et al (2002) Focal cortical dysplasia of Taylor’s balloon cell type: mutational analysis of the TSC1 gene indicates a pathogenic relationship to tuberous sclerosis. Ann Neurol 52(1):29–37PubMedCrossRefGoogle Scholar
  5. 5.
    Berg AT, Mathern GW, Bronen RA, Fulbright RK, DiMario F, Testa FM, Levy SR (2009) Frequency, prognosis and surgical treatment of structural abnormalities seen with magnetic resonance imaging in childhood epilepsy. Brain 132(Pt 10):2785–2797PubMedCrossRefGoogle Scholar
  6. 6.
    Bjorkoy G (2005) p62/SQSTM1 forms protein aggregates degraded by autophagy and has a protective effect on huntingtin-induced cell death. J Cell Biol 171(4):603–614PubMedCrossRefGoogle Scholar
  7. 7.
    Blümcke I, Thom M, Aronica E et al (2011) The clinicopathologic spectrum of focal cortical dysplasias: a consensus classification proposed by an ad hoc Task Force of the ILAE Diagnostic Methods Commission. Epilepsia 52(1):158–174PubMedCrossRefGoogle Scholar
  8. 8.
    Chen J, Tsai V, Parker WE, Aronica E, Baybis M, Crino PB (2012) Detection of human papillomavirus in human focal cortical dysplasia type IIB. Ann Neurol 72(6):881–892PubMedCrossRefGoogle Scholar
  9. 9.
    Choo AY, Yoon S-O, Kim SG, Roux PP, Blenis J (2008) Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci USA 105(45):17414–17419PubMedCrossRefGoogle Scholar
  10. 10.
    Crino PB, Trojanowski JQ, Eberwine J (1997) Internexin, MAP1B, and nestin in cortical dysplasia as markers of developmental maturity. Acta Neuropathol 93(6):619–627PubMedCrossRefGoogle Scholar
  11. 11.
    Crino PB (2011) mTOR: a pathogenic signaling pathway in developmental brain malformations. Trends Mol Med 17(12):734–742PubMedCrossRefGoogle Scholar
  12. 12.
    Crino PB (2013) Evolving neurobiology of tuberous sclerosis complex. Acta Neuropathol 125(3):317–332PubMedCrossRefGoogle Scholar
  13. 13.
    Glick D, Barth S, Macleod KF (2010) Autophagy: cellular and molecular mechanisms. J Pathol 221(1):3–12PubMedCrossRefGoogle Scholar
  14. 14.
    Grajkowska W, Kotulska K, Matyja E, Larysz-Brysz M, Mandera M, Roszkowski M, Domańska-Pakieła D, Lewik-Kowalik J, Jozwiak S (2008) Expression of tuberin and hamartin in tuberous sclerosis complex-associated and sporadic cortical dysplasia of Taylor’s balloon cell type. Folia Neuropathol 46(1):43–48PubMedGoogle Scholar
  15. 15.
    Hall MN (2008) mTOR-what does it do? TPS 40(10 Suppl):S5–S8Google Scholar
  16. 16.
    Hosokawa N, Hara T, Kaizuka T et al (2009) Nutrient-dependent mTORC1 association with the ULK1-Atg13-FIP200 complex required for autophagy. Mol Biol Cell 20(7):1981–1991PubMedCrossRefGoogle Scholar
  17. 17.
    Ichimura Y, Komatsu M (2010) Selective degradation of p62 by autophagy. Semin Immunopathol 32(4):431–436PubMedCrossRefGoogle Scholar
  18. 18.
    Joint Epilepsy Council (2011) Epilepsy prevalence, incidence and other statistics. Available via
  19. 19.
    Jozwiak J, Jozwiak S, Wlodarski P (2008) Possible mechanisms of disease development in tuberous sclerosis. Lancet Oncol 9(1):73–79PubMedCrossRefGoogle Scholar
  20. 20.
    Jung CH, Jun CB, Ro S-H, Kim Y-M, Otto NM, Cao J, Kundu M, Kim D-H (2009) ULK-Atg13-FIP200 complexes mediate mTOR signaling to the autophagy machinery. Mol Biol Cell 20(7):1992–2003PubMedCrossRefGoogle Scholar
  21. 21.
    Kim J, Kundu M, Viollet B, Guan K-L (2011) AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1. Nat Cell Biol 13(2):132–141PubMedCrossRefGoogle Scholar
  22. 22.
    Kurtz Z, Tookey P, Ross E (1998) Epilepsy in young people: 23 year follow up of the British national child development study. BMJ 316(7128):339–342PubMedCrossRefGoogle Scholar
  23. 23.
    Lamparello P, Baybis M, Pollard J, Hol EM, Eisenstat DD, Aronica E, Crino PB (2007) Developmental lineage of cell types in cortical dysplasia with balloon cells. Brain 130(Pt 9):2267–2276PubMedCrossRefGoogle Scholar
  24. 24.
    Ljungberg MC, Bhattacharjee MB, Lu Y, Armstrong DL, Yoshor D, Swann JW, Sheldon M, D’arcangelo G (2006) Activation of mammalian target of rapamycin in cytomegalic neurons of human cortical dysplasia. Ann Neurol 60(4):420–429PubMedCrossRefGoogle Scholar
  25. 25.
    Lugnier C, Majores M, Fassunke J, Pernhorst K, Niehusmann P, Simon M, Nellist M, Schoch S, Becker A (2009) Hamartin variants that are frequent in focal dysplasias and cortical tubers have reduced tuberin binding and aberrant subcellular distribution in vitro. J Neuropathol Exp Neurol 68(10):1136–1146PubMedCrossRefGoogle Scholar
  26. 26.
    Ma J-F, Huang Y, Chen S-D, Halliday G (2010) Immunohistochemical evidence for macroautophagy in neurones and endothelial cells in Alzheimer’s disease. Neuropathol Appl Neurobiol 36(4):312–319PubMedCrossRefGoogle Scholar
  27. 27.
    Mauvezin C, Orpinell M, Francis VA, Mansilla F, Duran J, Ribas V, Palacín M, Boya P, Teleman AA, Zorzano A (2010) The nuclear cofactor DOR regulates autophagy in mammalian and Drosophila cells. EMBO Rep 11(1):37–44PubMedCrossRefGoogle Scholar
  28. 28.
    Mauvezin C, Sancho A, Ivanova S, Palacín M, Zorzano A (2012) DOR undergoes nucleo-cytoplasmic shuttling, which involves passage through the nucleolus. FEBS Lett 586(19):3179–3186PubMedCrossRefGoogle Scholar
  29. 29.
    McMahon J, Huang X, Yang J, Komatsu M, Yue Z, Qian J, Zhu X, Huang Y (2012) Impaired autophagy in neurons after disinhibition of mammalian target of rapamycin and its contribution to epileptogenesis. J Neurosci 32(45):15704–15714PubMedCrossRefGoogle Scholar
  30. 30.
    Miyahara H, Natsumeda M, Shiga A et al (2013) Suppressed expression of autophagosomal protein LC3 in cortical tubers of tuberous sclerosis complex. Brain Pathol 23(3):254–262PubMedCrossRefGoogle Scholar
  31. 31.
    Miyata H, Chiang ACY, Vinters HV (2004) Insulin signaling pathways in cortical dysplasia and TSC-tubers: tissue microarray analysis. Ann Neurol 56(4):510–519PubMedCrossRefGoogle Scholar
  32. 32.
    Mizuguchi M, Ikeda K, Takashima S (2000) Simultaneous loss of hamartin and tuberin from the cerebrum, kidney and heart with tuberous sclerosis. Acta Neuropathol 99(5):503–510PubMedCrossRefGoogle Scholar
  33. 33.
    Mizuguchi M, Kato M, Yamanouchi H, Ikeda K, Takashima S (1996) Loss of tuberin from cerebral tissues with tuberous sclerosis and astrocytoma. Ann Neurol 40(6):941–944PubMedCrossRefGoogle Scholar
  34. 34.
    Schick V, Majores M, Engels G, Hartmann W, Elger CE, Schramm J, Schoch S, Becker AJ (2007) Differential Pi3K-pathway activation in cortical tubers and focal cortical dysplasias with balloon cells. Brain Pathol 17(2):165–173PubMedCrossRefGoogle Scholar
  35. 35.
    Schick V, Majores M, Koch A, Elger CE, Schramm J, Urbach H, Becker AJ (2007) Alterations of phosphatidylinositol 3-kinase pathway components in epilepsy-associated glioneuronal lesions. Epilepsia 48(Suppl 5):65–73PubMedCrossRefGoogle Scholar
  36. 36.
    Sebire N, Malone M, Ashworth M, Jacques TS (2010) Diagnostic Pediatric Surgical Pathology. Elsevier, LondonGoogle Scholar
  37. 37.
    Urbach H, Scheffler B, Heinrichsmeier T, von Oertzen J, Kral T, Wellmer J, Schramm J, Wiestler OD, Blümcke I (2002) Focal cortical dysplasia of Taylor’s balloon cell type: a clinicopathological entity with characteristic neuroimaging and histopathological features, and favorable postsurgical outcome. Epilepsia 43(1):33–40PubMedCrossRefGoogle Scholar
  38. 38.
    Vázquez P, Arroba AI, Cecconi F, de la Rosa EJ, Boya P, de Pablo F (2012) Atg5 and Ambra1 differentially modulate neurogenesis in neural stem cells. Autophagy 8((2):187–199PubMedCrossRefGoogle Scholar
  39. 39.
    Yasin SA, Latak K, Becherini F et al (2010) Balloon cells in human cortical dysplasia and tuberous sclerosis: isolation of a pathological progenitor-like cell. Acta Neuropathol 120(1):85–96PubMedCrossRefGoogle Scholar
  40. 40.
    Ying Z, Gonzalez-Martinez J, Tilelli C, Bingaman W, Najm I (2005) Expression of neural stem cell surface marker CD133 in balloon cells of human focal cortical dysplasia. Epilepsia 46(11):1716–1723PubMedCrossRefGoogle Scholar
  41. 41.
    Yu L, Mcphee CK, Zheng L et al (2010) Termination of autophagy and reformation of lysosomes regulated by mTOR. Nature 465(7300):942–946PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2013

Authors and Affiliations

  • Shireena A. Yasin
    • 1
    • 3
  • Abu M. Ali
    • 1
  • Mathew Tata
    • 1
  • Simon R. Picker
    • 1
    • 3
    • 6
  • Glenn W. Anderson
    • 3
  • Elizabeth Latimer-Bowman
    • 3
  • Sarah L. Nicholson
    • 3
  • William Harkness
    • 2
    • 4
  • J. Helen Cross
    • 2
    • 5
  • Simon M. L. Paine
    • 1
    • 3
  • Thomas S. Jacques
    • 1
    • 3
    Email author
  1. 1.Neural Development Unit, Birth Defects Research CentreUCL Institute of Child HealthLondonUK
  2. 2.Neurosciences UnitUCL Institute of Child HealthLondonUK
  3. 3.Department of HistopathologyGreat Ormond Street Hospital for Children NHS Foundation TrustLondonUK
  4. 4.Department of NeurosurgeryGreat Ormond Street Hospital for Children NHS Foundation TrustLondonUK
  5. 5.Department of NeurologyGreat Ormond Street Hospital for Children NHS Foundation TrustLondonUK
  6. 6.MRC National Institute for Medical ResearchLondonUK

Personalised recommendations