Protease Inhibitors, Saquinavir and Darunavir, Inhibit Oligodendrocyte Maturation: Implications for Lysosomal Stress

  • Lindsay Festa
  • Lindsay M. Roth
  • Brigid K. Jensen
  • Jonathan D. Geiger
  • Kelly L. Jordan-SciuttoEmail author
  • Judith B. GrinspanEmail author


Despite the introduction of antiretroviral (ARV) therapy (ART), approximately 30–50% of people living with human immunodeficiency virus-1 (HIV-1) will develop a spectrum of measurable neurocognitive dysfunction, collectively called HIV-associated neurocognitive disorder (HAND). While the clinical manifestations of HAND have changed with the advent of ART, certain pathological features have endured, including white matter alterations and dysfunction. The persistence of white matter alterations in the post-ART era suggests that ARV drugs themselves may contribute to HAND pathology. Our group has previously demonstrated that two ARV compounds from the protease inhibitor (PI) class, ritonavir and lopinavir, inhibit oligodendrocyte maturation and myelin protein production. We hypothesized that other members of the PI class, saquinavir and darunavir, could also negatively impact oligodendrocyte differentiation. Here we demonstrate that treating primary rat oligodendrocyte precursor cells with therapeutically relevant concentrations of either ARV drug results in a concentration-dependent inhibition of oligodendrocyte maturation in vitro. Furthermore, we show that acidifying endolysosomal pH via a mucolipin transient receptor potential channel 1 (TRPML1) agonist provides protection against saquinavir- and darunavir-induced inhibition of oligodendrocyte maturation. Moreover, our findings suggest, for the first time, an imperative role of proper endolysosomal pH in regulating OL differentation, and that therapeutic targeting of endolysosomes may provide protection against ARV-induced oligodendrocyte dysregulation.

Graphical Abstract

Treatment of primary rat oligodendrocyte precursor cells with therapeutically relevant concentrations of either antiretroviral compound of the protease inhibitor class, darunavir or saquinavir, results in a concentration-dependent inhibition of oligodendrocyte maturation in vitro. Additionally, in darunavir or saquinavir-treated cultures we observed a concentration-dependent decrease in the number of acidic lysosomes, via immunostaining with LysoTracker Red, compared with vehicle-treated cultures. Finally, we showed that acidifying endolysosomal pH via a mucolipin transient receptor potential channel 1 (TRPML1) agonist provides protection against saquinavir- or darunavir-induced inhibition of oligodendrocyte maturation. Our findings suggest, for the first time, a critical role of proper endolysosomal pH in regulating OL differentation, and that therapeutic targeting of endolysosomes may provide protection against antiretroviral-induced oligodendrocyte dysregulation.


Antiretroviral therapy Oligodendrocyte White matter Endolysosome 



We thank the NIH AIDS reagent program for their generous donation of ARVs and the laboratory of Michael Robinson at The Children’s Hospital of Philadelphia for the use of the Odyssey Infrared Imaging System.


This project was supported by the following grants: RO1 MH098742 (KJS and JBG), R21 MH118121 (JBG and KJS), NIH F31 NS079192 (BKJ), T32 NS007180 (BKJ,), T32 GM008076 (LMR), P30GM100329, U54GM115458, R01MH100972, R01MH105329, 2R01NS065957 and 2R01DA032444 (JDG) and the Cellular Neuroscience Core of the Institutional Intellectual and Developmental Disabilities Research Core of the Children’s Hospital of Philadelphia (HD26979).

Compliance with Ethical Standards

Conflict of Interest

The authors declare no competing financial interests.


  1. Akay C, Cooper M, Odeleye A, Jensen BK, White MG et al (2014) Antiretroviral drugs induce oxidative stress and neuronal damage in the central nervous system. J Neuro-Oncol 20:39–53Google Scholar
  2. Akay C, Lindl KA, Shyam N, Nabet B, Goenaga-Vazquez Y et al (2012) Activation status of integrated stress response pathways in neurones and astrocytes of HIV-associated neurocognitive disorders (HAND) cortex. Neuropathol Appl Neurobiol 38:175–200PubMedPubMedCentralCrossRefGoogle Scholar
  3. Bae M, Patel N, Xu H, Lee M, Tominaga-Yamanaka K, Nath A, Geiger J, Gorospe M, Mattson MP, Haughey NJ (2014) Activation of TRPML1 clears intraneuronal Abeta in preclinical models of HIV infection. J Neurosci 34:11485–11503PubMedPubMedCentralCrossRefGoogle Scholar
  4. Bansal R, Warrington AE, Gard AL, Ranscht B, Pfeiffer SE (1989) Multiple and novel specificities of monoclonal antibodies O1, O4, and R-mAb used in the analysis of oligodendrocyte development. J Neurosci Res 24:548–557PubMedCrossRefGoogle Scholar
  5. Baracskay KL, Kidd GJ, Miller RH, Trapp BD (2007) NG2-positive cells generate A2B5-positive oligodendrocyte precursor cells. Glia 55:1001–1010PubMedCrossRefGoogle Scholar
  6. Beyer BA, Fang M, Sadrian B, Montenegro-Burke JR, Plaisted WC, Kok BPC, Saez E, Kondo T, Siuzdak G, Lairson LL (2018) Metabolomics-based discovery of a metabolite that enhances oligodendrocyte maturation. Nat Chem Biol 14:22–28PubMedCrossRefGoogle Scholar
  7. Borjabad A, Brooks AI, Volsky DJ (2010) Gene expression profiles of HIV-1-infected glia and brain: toward better understanding of the role of astrocytes in HIV-1-associated neurocognitive disorders. J NeuroImmune Pharmacol 5:44–62PubMedCrossRefGoogle Scholar
  8. Brandmann M, Tulpule K, Schmidt MM, Dringen R (2012) The antiretroviral protease inhibitors indinavir and nelfinavir stimulate Mrp1-mediated GSH export from cultured brain astrocytes. J Neurochem 120:78–92PubMedCrossRefGoogle Scholar
  9. Chandra S, Mondal D, Agrawal KC (2009) HIV-1 protease inhibitor induced oxidative stress suppresses glucose stimulated insulin release: protection with thymoquinone. Exp Biol Med (Maywood) 234:442–453CrossRefGoogle Scholar
  10. Chen X, Hui L, Geiger NH, Haughey NJ, Geiger JD (2013) Endolysosome involvement in HIV-1 transactivator protein-induced neuronal amyloid beta production. Neurobiol Aging 34:2370–2378PubMedPubMedCentralCrossRefGoogle Scholar
  11. Chew LJ, King WC, Kennedy A, Gallo V (2005) Interferon-gamma inhibits cell cycle exit in differentiating oligodendrocyte progenitor cells. Glia 52:127–143PubMedCrossRefGoogle Scholar
  12. Correa DG, Zimmermann N, Doring TM, Wilner NV, Leite SC et al (2015) Diffusion tensor MR imaging of white matter integrity in HIV-positive patients with planning deficit. Neuroradiology 57:475–482PubMedCrossRefGoogle Scholar
  13. Dai J, Bercury KK, Macklin WB (2014) Interaction of mTOR and Erk1/2 signaling to regulate oligodendrocyte differentiation. Glia 62:2096–2109PubMedPubMedCentralCrossRefGoogle Scholar
  14. Ellis RJ, Marquie-Beck J, Delaney P, Alexander T, Clifford DB et al (2008) Human immunodeficiency virus protease inhibitors and risk for peripheral neuropathy. Ann Neurol 64:566–572PubMedPubMedCentralCrossRefGoogle Scholar
  15. Eisenbarth GS, Walsh FS, Nirenberg M (1979) Monoclonal antibody to a plasma membrane antigen of neurons. Proc. Natl. Acad. Sci. 76 (10):4913-4917CrossRefGoogle Scholar
  16. Everall I, Vaida F, Khanlou N, Lazzaretto D, Achim C et al (2009) Cliniconeuropathologic correlates of human immunodeficiency virus in the era of antiretroviral therapy. J Neuro-Oncol 15:360–370Google Scholar
  17. Everall IP, Hansen LA, Masliah E (2005) The shifting patterns of HIV encephalitis neuropathology. Neurotox Res 8:51–61PubMedCrossRefGoogle Scholar
  18. Fauci AS, Marston HD (2015) Ending the HIV-AIDS pandemic--follow the science. N Engl J Med 373:2197–2199PubMedCrossRefGoogle Scholar
  19. Feigenson K, Reid M, See J, Crenshaw EB 3rd, Grinspan JB (2009) Wnt signaling is sufficient to perturb oligodendrocyte maturation. Mol Cell Neurosci 42:255–265PubMedCrossRefGoogle Scholar
  20. Feigenson K, Reid M, See J, Crenshaw IE, Grinspan JB (2011) Canonical Wnt signalling requires the BMP pathway to inhibit oligodendrocyte maturation. ASN Neuro 3:e00061PubMedPubMedCentralCrossRefGoogle Scholar
  21. Fields J, Dumaop W, Eleuteri S, Campos S, Serger E et al (2015) HIV-1 tat alters neuronal autophagy by modulating autophagosome fusion to the lysosome: implications for HIV-associated neurocognitive disorders. J Neurosci 35:1921–1938PubMedPubMedCentralCrossRefGoogle Scholar
  22. French HM, Reid M, Mamontov P, Simmons RA, Grinspan JB (2009) Oxidative stress disrupts oligodendrocyte maturation. J Neurosci Res 87:3076–3087PubMedPubMedCentralCrossRefGoogle Scholar
  23. Gannon PJ, Akay-Espinoza C, Yee AC, Briand LA, Erickson MA, Gelman BB, Gao Y, Haughey NJ, Zink MC, Clements JE, Kim NS, van de Walle G, Jensen BK, Vassar R, Pierce RC, Gill AJ, Kolson DL, Diehl JA, Mankowski JL, Jordan-Sciutto KL (2017) HIV protease inhibitors Alter amyloid precursor protein processing via beta-site amyloid precursor protein cleaving Enzyme-1 translational up-regulation. Am J Pathol 187:91–109PubMedPubMedCentralCrossRefGoogle Scholar
  24. Gavrieli Y, Sherman Y, Ben-Sasson SA (1992) Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol 119:493–501PubMedCrossRefGoogle Scholar
  25. Gongvatana A, Cohen RA, Correia S, Devlin KN, Miles J et al (2011) Clinical contributors to cerebral white matter integrity in HIV-infected individuals. J Neuro-Oncol 17:477–486Google Scholar
  26. Hart IK, Richardson WD, Heldin CH, Westermark B, Raff MC (1989) PDGF receptors on cells of the oligodendrocyte-type-2 astrocyte (O-2A) cell lineage. Development 105:595–603PubMedGoogle Scholar
  27. Heaton RK, Clifford DB, Franklin DR Jr, Woods SP, Ake C et al (2010) HIV-associated neurocognitive disorders persist in the era of potent antiretroviral therapy: CHARTER study. Neurology 75:2087–2096PubMedPubMedCentralCrossRefGoogle Scholar
  28. Hoare J, Fouche JP, Spottiswoode B, Joska JA, Schoeman R et al (2010) White matter correlates of apathy in HIV-positive subjects: a diffusion tensor imaging study. J Neuropsychiatry Clin Neurosci 22:313–320PubMedCrossRefGoogle Scholar
  29. Hui L, Ye Y, Soliman ML, Lakpa KL, Miller NM et al (2019) Antiretroviral drugs promote Amyloidogenesis by De-acidifying Endolysosomes. J NeuroImmune PharmacolGoogle Scholar
  30. Hussien Y, Cavener DR, Popko B (2014) Genetic inactivation of PERK signaling in mouse oligodendrocytes: normal developmental myelination with increased susceptibility to inflammatory demyelination. Glia 62:680–691PubMedPubMedCentralCrossRefGoogle Scholar
  31. Jensen BK, Monnerie H, Mannell MV, Gannon PJ, Espinoza CA, Erickson MA, Bruce-Keller AJ, Gelman BB, Briand LA, Pierce RC, Jordan-Sciutto KL, Grinspan JB (2015) Altered Oligodendrocyte maturation and myelin maintenance: the role of Antiretrovirals in HIV-associated neurocognitive disorders. J Neuropathol Exp Neurol 74:1093–1118PubMedPubMedCentralCrossRefGoogle Scholar
  32. Jensen BK, Roth LM, Grinspan JB, Jordan-Sciutto KL (2019) White matter loss and oligodendrocyte dysfunction in HIV: a consequence of the infection, the antiretroviral therapy or both? Brain Res 1724:146397PubMedCrossRefGoogle Scholar
  33. Lagathu C, Eustace B, Prot M, Frantz D, Gu Y, Bastard JP, Maachi M, Azoulay S, Briggs M, Caron M, Capeau J (2007) Some HIV antiretrovirals increase oxidative stress and alter chemokine, cytokine or adiponectin production in human adipocytes and macrophages. Antivir Ther 12:489–500PubMedGoogle Scholar
  34. Langford TD, Letendre SL, Larrea GJ, Masliah E (2003) Changing patterns in the neuropathogenesis of HIV during the HAART era. Brain Pathol 13:195–210PubMedPubMedCentralCrossRefGoogle Scholar
  35. Lindl KA, Akay C, Wang Y, White MG, Jordan-Sciutto KL (2007) Expression of the endoplasmic reticulum stress response marker, BiP, in the central nervous system of HIV-positive individuals. Neuropathol Appl Neurobiol 33:658–669PubMedCrossRefGoogle Scholar
  36. Manda KR, Banerjee A, Banks WA, Ercal N (2011) Highly active antiretroviral therapy drug combination induces oxidative stress and mitochondrial dysfunction in immortalized human blood-brain barrier endothelial cells. Free Radic Biol Med 50:801–810PubMedCrossRefGoogle Scholar
  37. McCarthy KD, de Vellis J (1980) Preparation of separate astroglial and oligodendroglial cell cultures from rat cerebral tissue. J Cell Biol 85:890–902PubMedCrossRefGoogle Scholar
  38. Meireles AM, Shen K, Zoupi L, Iyer H, Bouchard EL et al (2018) The Lysosomal transcription factor TFEB represses myelination downstream of the rag-Ragulator complex. Dev Cell 47(319–330):e315Google Scholar
  39. Miller RH (2002) Regulation of oligodendrocyte development in the vertebrate CNS. Prog Neurobiol 67:451–467PubMedCrossRefPubMedCentralGoogle Scholar
  40. Muller-Oehring EM, Schulte T, Rosenbloom MJ, Pfefferbaum A, Sullivan EV (2010) Callosal degradation in HIV-1 infection predicts hierarchical perception: a DTI study. Neuropsychologia 48:1133–1143PubMedCrossRefGoogle Scholar
  41. Nishiyama A, Lin XH, Giese N, Heldin CH, Stallcup WB (1996) Co-localization of NG2 proteoglycan and PDGF alpha-receptor on O2A progenitor cells in the developing rat brain. J Neurosci Res 43:299–314PubMedCrossRefGoogle Scholar
  42. Raff MC, Abney ER, Cohen J, Lindsay R, Noble M (1983) Two types of astrocytes in cultures of developing rat white matter: differences in morphology, surface gangliosides, and growth characteristics. J Neurosci 3:1289–1300PubMedPubMedCentralCrossRefGoogle Scholar
  43. Raff MC, Mirsky R, Fields KL, Lisak RP, Dorfman SH et al (1978) Galactocerebroside is a specific cell-surface antigenic marker for oligodendrocytes in culture. Nature 274:813–816PubMedCrossRefGoogle Scholar
  44. Ranscht B, Clapshaw PA, Price J, Noble M, Seifert W (1982) Development of oligodendrocytes and Schwann cells studied with a monoclonal antibody against galactocerebroside. Proc Natl Acad Sci U S A 79:2709–2713PubMedPubMedCentralCrossRefGoogle Scholar
  45. Reid MV, Murray KA, Marsh ED, Golden JA, Simmons RA, Grinspan JB (2012) Delayed myelination in an intrauterine growth retardation model is mediated by oxidative stress upregulating bone morphogenetic protein 4. J Neuropathol Exp Neurol 71:640–653PubMedPubMedCentralCrossRefGoogle Scholar
  46. Romero-Ramirez L, Nieto-Sampedro M, Barreda-Manso MA (2017) Integrated stress response as a therapeutic target for CNS injuries. Biomed Res Int 2017:6953156PubMedPubMedCentralCrossRefGoogle Scholar
  47. Saylor D, Dickens AM, Sacktor N, Haughey N, Slusher B et al (2016) HIV-associated neurocognitive disorder - pathogenesis and prospects for treatment. Nat Rev Neurol 12:309PubMedPubMedCentralCrossRefGoogle Scholar
  48. See J, Zhang X, Eraydin N, Mun SB, Mamontov P, Golden JA, Grinspan JB (2004) Oligodendrocyte maturation is inhibited by bone morphogenetic protein. Mol Cell Neurosci 26:481–492PubMedCrossRefGoogle Scholar
  49. Settembre C, Fraldi A, Medina DL, Ballabio A (2013) Signals from the lysosome: a control Centre for cellular clearance and energy metabolism. Nat Rev Mol Cell Biol 14:283–296PubMedPubMedCentralCrossRefGoogle Scholar
  50. Stern AL, Ghura S, Gannon PJ, Akay-Espinoza C, Phan JM, Yee AC, Vassar R, Gelman BB, Kolson DL, Jordan-Sciutto KL (2018) BACE1 mediates HIV-associated and Excitotoxic neuronal damage through an APP-dependent mechanism. J Neurosci 38:4288–4300PubMedPubMedCentralCrossRefGoogle Scholar
  51. Tate DF, Conley J, Paul RH, Coop K, Zhang S, Zhou W, Laidlaw DH, Taylor LE, Flanigan T, Navia B, Cohen R, Tashima K (2010) Quantitative diffusion tensor imaging tractography metrics are associated with cognitive performance among HIV-infected patients. Brain Imaging Behav 4:68–79PubMedPubMedCentralCrossRefGoogle Scholar
  52. Tate DF, Sampat M, Harezlak J, Fiecas M, Hogan J et al (2011) Regional areas and widths of the midsagittal corpus callosum among HIV-infected patients on stable antiretroviral therapies. J Neuro-Oncol 17:368–379Google Scholar
  53. Touzet O, Philips A (2010) Resveratrol protects against protease inhibitor-induced reactive oxygen species production, reticulum stress and lipid raft perturbation. AIDS 24:1437–1447PubMedCrossRefGoogle Scholar
  54. Vermeir M, Lachau-Durand S, Mannens G, Cuyckens F, van Hoof B et al (2009) Absorption, metabolism, and excretion of darunavir, a new protease inhibitor, administered alone and with low-dose ritonavir in healthy subjects. Drug Metab Dispos 37:809–820PubMedCrossRefGoogle Scholar
  55. Way SW, Popko B (2016) Harnessing the integrated stress response for the treatment of multiple sclerosis. Lancet Neurol 15:434–443PubMedPubMedCentralCrossRefGoogle Scholar
  56. WHO, 2016 Clinical Guidelines: Antiretroviral Therapy, pp. 97 & 151Google Scholar
  57. Yu T, Lieberman AP (2013) Npc1 acting in neurons and glia is essential for the formation and maintenance of CNS myelin. PLoS Genet 9:e1003462PubMedPubMedCentralCrossRefGoogle Scholar
  58. Zhang Y, Chen K, Sloan SA, Bennett ML, Scholze AR et al (2014) An RNA-sequencing transcriptome and splicing database of glia, neurons, and vascular cells of the cerebral cortex. J Neurosci 34:11929–11947PubMedPubMedCentralCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Lindsay Festa
    • 1
  • Lindsay M. Roth
    • 1
    • 2
  • Brigid K. Jensen
    • 3
  • Jonathan D. Geiger
    • 4
  • Kelly L. Jordan-Sciutto
    • 1
    Email author
  • Judith B. Grinspan
    • 2
    Email author
  1. 1.Department of Pathology, School of Dental MedicineUniversity of PennsylvaniaPhiladelphiaUSA
  2. 2.Department of NeurologyThe Children’s Hospital of PhiladelphiaPhiladelphiaUSA
  3. 3.Vickie and Jack Farber Institute for Neuroscience, Jefferson Weinberg ALS CenterThomas Jefferson UniversityPhiladelphiaUSA
  4. 4.Department of Biomedical Sciences, School of Medicine and Health SciencesUniversity of North DakotaGrand ForksUSA

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