Molecular Neurobiology

, Volume 55, Issue 8, pp 7025–7037 | Cite as

Disruption of De Novo Serine Synthesis in Müller Cells Induced Mitochondrial Dysfunction and Aggravated Oxidative Damage

  • Ting Zhang
  • Mark C. Gillies
  • Michele C. Madigan
  • Weiyong Shen
  • Jianhai Du
  • Ulrike Grünert
  • Fanfan Zhou
  • Michelle Yam
  • Ling Zhu


De novo serine synthesis plays important roles in normal mitochondrial function and cellular anti-oxidative capacity. It is reported to be mainly activated in glial cells of the central nervous system, but its role in retinal Müller glia remains unclear. In this study, we inhibited de novo serine synthesis using CBR-5884, a specific inhibitor of phosphoglycerate dehydrogenase (PHGDH, a rate limiting enzyme in de novo serine metabolism) in MIO-M1 cells (immortalized human Müller cells) and huPMCs (human primary Müller cells) under mild oxidative stress. Alamar blue and LDH (lactate dehydrogenase) assays showed significantly reduced metabolic activities and increased cellular damage of Müller cells, when exposed to CBR-5884 accompanied by mild oxidative stress; however, CBR-5884 alone had little effect. The increased cellular damage was partially reversed by supplementation with exogenous serine/glycine. HSP72 (an oxidative stress marker) and reactive oxygen species (ROS) levels were significantly increased; glutathione and NADPH/NADP+ levels were pronouncedly reduced under PHGDH inhibition accompanied by oxidative stress. JC-1 staining and Seahorse respiration experiments showed that inhibition of de novo serine synthesis in Müller cells can also increase mitochondrial stress and decrease mitochondrial ATP production. qPCR and Western blot demonstrated an increased expression of HSP60 (a key mitochondrial stress-related gene), and this was further validated in human retinal explants. Our study suggests that de novo serine synthesis is important for Müller cell survival, particularly when they are exposed to mild oxidative stress, possibly by maintaining mitochondrial function and generating glutathione and NADPH to counteract ROS.


Müller cells De novo serine synthesis Mitochondrial dysfunction Oxidative stress Phosphoglycerate dehydrogenase Glutathione 



This study is supported by a grant from the Lowy Medical Research Institute. Professor Mark C. Gillies is a Sydney Medical School Fellow and is supported by a NHMRC Practitioner Fellowship. This paper formed the foundation of NHMRC project grant APP1145121.

Compliance with Ethical Standards

Conflict of Interest

The authors declare that they have no conflict of interest.

Supplementary material

12035_2017_840_MOESM1_ESM.pdf (212 kb)
ESM 1 (PDF 211 kb)


  1. 1.
    Lin MT, Beal MF (2006) Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases. Nature 443(7113):787–795. CrossRefPubMedGoogle Scholar
  2. 2.
    Cui H, Kong Y, Zhang H (2012) Oxidative stress, mitochondrial dysfunction, and aging. J Signal Transduction 2012:646354. CrossRefGoogle Scholar
  3. 3.
    Martinez-Reyes I, Chandel NS (2014) Mitochondrial one-carbon metabolism maintains redox balance during hypoxia. Cancer Discov 4(12):1371–1373. CrossRefPubMedPubMedCentralGoogle Scholar
  4. 4.
    Yang M, Vousden KH (2016) Serine and one-carbon metabolism in cancer. Nat Rev Cancer 16(10):650–662. CrossRefPubMedGoogle Scholar
  5. 5.
    Pacold ME, Brimacombe KR, Chan SH, Rohde JM, Lewis CA, Swier LJ, Possemato R, Chen WW et al (2016) A PHGDH inhibitor reveals coordination of serine synthesis and one-carbon unit fate. Nat Chem Biol 12(6):452–458. CrossRefPubMedPubMedCentralGoogle Scholar
  6. 6.
    Locasale JW (2013) Serine, glycine and one-carbon units: cancer metabolism in full circle. Nat Rev Cancer 13(8):572–583. CrossRefPubMedPubMedCentralGoogle Scholar
  7. 7.
    Mukherjee B, Mukherjee JR, Chatterjee M (1994) Lipid peroxidation, glutathione levels and changes in glutathione-related enzyme activities in streptozotocin-induced diabetic rats. Immunol Cell Biol 72(2):109–114. CrossRefPubMedGoogle Scholar
  8. 8.
    Owen JB, Butterfield DA (2010) Measurement of oxidized/reduced glutathione ratio. Methods Mol Biol 648:269–277. CrossRefPubMedGoogle Scholar
  9. 9.
    Tabatabaie L, Klomp LW, Berger R, de Koning TJ (2010) L-serine synthesis in the central nervous system: a review on serine deficiency disorders. Mol Genet Metab 99(3):256–262. CrossRefPubMedGoogle Scholar
  10. 10.
    Furuya S (2008) An essential role for de novo biosynthesis of L-serine in CNS development. Asia Pac J Clin Nutr 17(Suppl 1):312–315PubMedGoogle Scholar
  11. 11.
    Furuya S, Tabata T, Mitoma J, Yamada K, Yamasaki M, Makino A, Yamamoto T, Watanabe M et al (2000) L-serine and glycine serve as major astroglia-derived trophic factors for cerebellar Purkinje neurons. Proc Natl Acad Sci U S A 97(21):11528–11533. CrossRefPubMedPubMedCentralGoogle Scholar
  12. 12.
    Sagara JI, Miura K, Bannai S (1993) Maintenance of neuronal glutathione by glial cells. J Neurochem 61(5):1672–1676. CrossRefPubMedGoogle Scholar
  13. 13.
    Bringmann A, Reichenbach A (2001) Role of Muller cells in retinal degenerations. Front Biosci 6:E72–E92CrossRefPubMedGoogle Scholar
  14. 14.
    Hurley JB, Lindsay KJ, Du J (2015) Glucose, lactate, and shuttling of metabolites in vertebrate retinas. J Neurosci Res 93(7):1079–1092. CrossRefPubMedPubMedCentralGoogle Scholar
  15. 15.
    Bringmann A, Iandiev I, Pannicke T, Wurm A, Hollborn M, Wiedemann P, Osborne NN, Reichenbach A (2009) Cellular signaling and factors involved in Muller cell gliosis: neuroprotective and detrimental effects. Prog Retin Eye Res 28(6):423–451. CrossRefPubMedGoogle Scholar
  16. 16.
    Poitry S, Poitry-Yamate C, Ueberfeld J, MacLeish PR, Tsacopoulos M (2000) Mechanisms of glutamate metabolic signaling in retinal glial (Muller) cells. J Neurosci 20(5):1809–1821CrossRefPubMedGoogle Scholar
  17. 17.
    Fletcher EL, Phipps JA, Ward MM, Puthussery T, Wilkinson-Berka JL (2007) Neuronal and glial cell abnormality as predictors of progression of diabetic retinopathy. Curr Pharm Des 13(26):2699–2712. CrossRefPubMedGoogle Scholar
  18. 18.
    Wu KH, Madigan MC, Billson FA, Penfold PL (2003) Differential expression of GFAP in early v late AMD: a quantitative analysis. Br J Ophthalmol 87(9):1159–1166CrossRefPubMedPubMedCentralGoogle Scholar
  19. 19.
    Powner MB, Gillies MC, Tretiach M, Scott A, Guymer RH, Hageman GS, Fruttiger M (2010) Perifoveal muller cell depletion in a case of macular telangiectasia type 2. Ophthalmology 117(12):2407–2416. CrossRefPubMedPubMedCentralGoogle Scholar
  20. 20.
    Powner MB, Gillies MC, Zhu M, Vevis K, Hunyor AP, Fruttiger M (2013) Loss of Muller's cells and photoreceptors in macular telangiectasia type 2. Ophthalmology 120(11):2344–2352. CrossRefPubMedGoogle Scholar
  21. 21.
    Scerri TS, Quaglieri A, Cai C, Zernant J, Matsunami N, Baird L, Scheppke L, Bonelli R et al (2017) Genome-wide analyses identify common variants associated with macular telangiectasia type 2. Nat Genet 49(4):559–567. CrossRefPubMedGoogle Scholar
  22. 22.
    Zhu L, Shen W, Lyons B, Wang Y, Zhou F, Gillies MC (2015) Dysregulation of inter-photoreceptor retinoid-binding protein (IRBP) after induced Muller cell disruption. J Neurochem 133(6):909–918. CrossRefPubMedGoogle Scholar
  23. 23.
    Zhu L, Shen W, Zhu M, Coorey NJ, Nguyen AP, Barthelmes D, Gillies MC (2013) Anti-retinal antibodies in patients with macular telangiectasia type 2. Invest Ophthalmol Vis Sci 54(8):5675–5683. CrossRefPubMedGoogle Scholar
  24. 24.
    Mehrmohamadi M, Liu X, Shestov AA, Locasale JW (2014) Characterization of the usage of the serine metabolic network in human cancer. Cell Rep 9(4):1507–1519. CrossRefPubMedPubMedCentralGoogle Scholar
  25. 25.
    Cossarizza A, Baccarani-Contri M, Kalashnikova G, Franceschi C (1993) A new method for the cytofluorimetric analysis of mitochondrial membrane potential using the J-aggregate forming lipophilic cation 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolcarbocyanine iodide (JC-1). Biochem Biophys Res Commun 197(1):40–45. CrossRefPubMedGoogle Scholar
  26. 26.
    Wu M, Neilson A, Swift AL, Moran R, Tamagnine J, Parslow D, Armistead S, Lemire K et al (2007) Multiparameter metabolic analysis reveals a close link between attenuated mitochondrial bioenergetic function and enhanced glycolysis dependency in human tumor cells. Am J Physiol Cell Physiol 292(1):C125–C136. CrossRefPubMedGoogle Scholar
  27. 27.
    Samanta D, Park Y, Andrabi SA, Shelton LM, Gilkes DM, Semenza GL (2016) PHGDH expression is required for mitochondrial redox homeostasis, breast cancer stem cell maintenance, and lung metastasis. Cancer Res 76(15):4430–4442. CrossRefPubMedGoogle Scholar
  28. 28.
    Ng SK, Wood JP, Chidlow G, Han G, Kittipassorn T, Peet DJ, Casson RJ (2015) Cancer-like metabolism of the mammalian retina. Clin Exp Ophthalmol 43(4):367–376. CrossRefPubMedGoogle Scholar
  29. 29.
    Mullarky E, Lucki NC, Beheshti Zavareh R, Anglin JL, Gomes AP, Nicolay BN, Wong JC, Christen S et al (2016) Identification of a small molecule inhibitor of 3-phosphoglycerate dehydrogenase to target serine biosynthesis in cancers. Proc Natl Acad Sci U S A 113(7):1778–1783. CrossRefPubMedPubMedCentralGoogle Scholar
  30. 30.
    Luo J (2011) Cancer’s sweet tooth for serine. Breast Cancer Res: BCR 13(6):317. CrossRefPubMedGoogle Scholar
  31. 31.
    Zoppi CC, Macedo DV (2008) Overreaching-induced oxidative stress, enhanced HSP72 expression, antioxidant and oxidative enzymes downregulation. Scand J Med Sci Sports 18(1):67–76. CrossRefPubMedGoogle Scholar
  32. 32.
    Mayer MP, Bukau B (2005) Hsp70 chaperones: cellular functions and molecular mechanism. Cell Mol Life Sci: CMLS 62(6):670–684. CrossRefPubMedGoogle Scholar
  33. 33.
    Park HS, Cho SG, Kim CK, Hwang HS, Noh KT, Kim MS, Huh SH, Kim MJ et al (2002) Heat shock protein hsp72 is a negative regulator of apoptosis signal-regulating kinase 1. Mol Cell Biol 22(22):7721–7730. CrossRefPubMedPubMedCentralGoogle Scholar
  34. 34.
    Nover L (1991) Heat shock response. Crc PressGoogle Scholar
  35. 35.
    Dorrell MI, Aguilar E, Jacobson R, Yanes O, Gariano R, Heckenlively J, Banin E, Ramirez GA et al (2009) Antioxidant or neurotrophic factor treatment preserves function in a mouse model of neovascularization-associated oxidative stress. J Clin Invest 119(3):611–623. CrossRefPubMedPubMedCentralGoogle Scholar
  36. 36.
    Yu DY, Cringle SJ (2001) Oxygen distribution and consumption within the retina in vascularised and avascular retinas and in animal models of retinal disease. Prog Retin Eye Res 20(2):175–208CrossRefPubMedGoogle Scholar
  37. 37.
    Limb GA, Salt TE, Munro PM, Moss SE, Khaw PT (2002) In vitro characterization of a spontaneously immortalized human Muller cell line (MIO-M1). Invest Ophthalmol Vis Sci 43(3):864–869PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2017

Authors and Affiliations

  1. 1.State Key Laboratory of Biotherapy and Cancer Center, Collaborative Innovation Center for Biotherapy, West China HospitalSichuan UniversityChengduPeople’s Republic of China
  2. 2.Save Sight InstituteThe University of SydneySydneyAustralia
  3. 3.School of Optometry and Vision SciencesUniversity of New South WalesSydneyAustralia
  4. 4.West Virginia University Health Sciences CenterMorgantownUSA
  5. 5.Faculty of PharmacyThe University of SydneySydneyAustralia

Personalised recommendations