Anat Cell Biol.  2022 Jun;55(2):179-189. 10.5115/acb.21.202.

A study on the effect of JNJ-10397049 on proliferation and differentiation of neural precursor cells

• Karami N
• Azari H
• Rahimi M
• Aligholi H
• Kalantari T

• KMID: 2531205
• DOI: http://doi.org/10.5115/acb.21.202

Abstract

The orexin 2 receptor plays a central role in maintaining sleep and wakefulness. Recently, it has been shown that sleep and wakefulness orchestrate the proliferation and differentiation of oligodendrocytes. Here, we explored the role of a selective orexin 2 receptor antagonist (JNJ-10397049) in proliferation and differentiation of neural progenitor cells (NPCs). We evaluated the proliferation potential of NPCs after exposure to different concentrations of JNJ-10397049 by using 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide and neurosphere assays. Moreover, the expression of differentiation markers was assessed by immunocytochemistry and real-time polymerase chain reaction. JNJ-10397049 significantly increased the proliferation of NPCs at lower concentrations. In addition, orexin 2 receptor antagonist facilitated progression of differentiation of NPCs towards oligodendroglial lineage by considerable expression of Olig2 and 2’,3’-cyclicnucleotide 3’-phosphodiesterase as well as decreased expression of nestin marker. The results open a new avenue for future investigations in which the production of more oligodendrocytes from NPCs is needed.

Keyword

Orexins; Orexin receptors; JNJ 10397049; Oligodendroglia; Neural stem cells

Figure

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  Fig. 1 Characterization of neurospheres established embryonic 14.5 days old mice brain. Phase contrast images of suspended neurospheres on day 5 and day 7 (A) of primary neurosphere culture (passage 0) at 4× magnification. (B) Nestin-labeled neural precursor cells (NPCs; green) on day 7 of culture (passage 0) with 7-aminoactinomycin D (7AAD)-counterstained nuclei (red) at 4× magnification. (C) HCRTR2 expression in the NPCs after passages 2 and 4. *P<0.001. Scale bar=100 μm.

• 

  Fig. 2 The results of 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) assay after exposure of the neural precursor cells (NPCs) to different concentration of Orexin 2 receptor inhibitor. (A) Graph representing obtained absorbance at 490 nm from an MTT assay of NPCs treated with varying concentrations (μM) of JNJ-10397049. Final concentration of dimethyl sulfoxide was 0.05% and was included in control wells. (B) Log(half maximal effective concentration, EC50) of JNJ-10397049 from log-normalized data was calculated using Graph Pad Prism 7. Data are presented as mean±standard error of the mean. *P<0.001 compared to control, n=3.

• 

  Fig. 3 Neurosphere assay after exposure of the neural precursor cells (NPCs) to different concentration of orexin 2 receptor inhibitor. (A) Phase contrast images of NPCs on day 7 of cell culture treated with varying concentrations of JNJ-10397049 at 4× magnification. The quantitative analysis of the number (B) and the diameter of neurospheres (C). Data are represented as mean±standard error of the mean. *P<0.001, n=3. Scale bar=100 μm.

• 

  Fig. 4 Changing the fate of neural precursor cells (NPCs) after exposure to orexin 2 receptor inhibitor. The expression of oligodendroglial marker Olig2 (green; A) and NPCs marker nestin (green; B) were evaluated in NPCs after treatment with 0 or 20 µm of JNJ-10397049. The nuclei were stained with 7-aminoactinomycin D (7AAD) and are shown in red. (C) Representative phase contrast images taken from the differentiation culture on day 7 at 10× magnification. The black arrowheads specify morphologically suggested mature oligodendrocytes. The white arrowhead in image bottom panel of C specifies cells likely to be of neuronal lineage. (D, E) Quantification of the percentage of Olig2 and nestin-positive cells per area on day 7 of culture. *P<0.001. Scale bar=100 μm, n=3.

• 

  Fig. 5 Expression of oligodendrocyte-related genes in neural precursor cells (NPCs) after exposure to orexin 2 receptor inhibitor. The relative expression of platelet derived growth factor a (PDGFa) and 2’,3’-cyclic-nucleotide 3’-phosphodiesterase (CNPase) in NPCs were evaluated after treatment of the cells with 0 or 20 µm of JNJ-10397049. *P<0.05, n=4.

• 

  Fig. 6 Orexin interplay with other neurotransmitters involved in vigilance state and expressed biomarkers of oligodendroglial lineage throughout differentiation from neural precursor cells (NPCs). (A) Orexin (Hcrt) is one of the main players of vigilance state and acts via regulating several neurotransmitters involved in this state. Vigilance state is mainted by cholinergic (Ach) inputs from brain stem (LDT, laterodorsal tegmentum; PRP, pontine reticular formation; PPT, pendonuclopontine tegmentum) to thalamus and subsequently activates the cortex. Cholinergic inputs from the basal forebrain (BF) are also involved. Other factors involved in maintaining cortical activation in wakefulness are serotonergic (5HT) dorsal raphe nucleus (DRN), noradrenergic (NA) (LC, corpus coeruleus), dopaminergic (DA) (vPAG, ventral periaqueductal), histaminergic (His) (TMN, tuberomammillary nucleus), and hypocretinergic (Hcrt) inputs [49]. (B) Expression of prominent biomarkers of oligodendrogenesis at different stages of differentiation of NPCs to myelinating oligodendrocytes. Nestin expression is pronounced in NPCs and oligodendrocyte precursor cells (OPCs) and is not detectable in immature oligodendrocytes. Platelet derived growth factor alpha (PDGFa) is present in OPCs and immature oligodendrocytes. Olig2 and Sox10 are expressed throughout almost all stages of oligodendrocyte differentiation (OPC, immature OL, mature and myelinating OL). 2’,3’-cyclic-nucleotide 3’-phosphodiesterase (CNPase) expression is noticeable in immature OLs and gradually increases in mature and myelinating OLs [54, 55].

Reference

References

1. Gage FH. 2000; Mammalian neural stem cells. Science. 287:1433–8. DOI: 10.1126/science.287.5457.1433. PMID: 10688783.

2. Ming GL, Song H. 2011; Adult neurogenesis in the mammalian brain: significant answers and significant questions. Neuron. 70:687–702. DOI: 10.1016/j.neuron.2011.05.001. PMID: 21609825. PMCID: PMC3106107.

3. Takano M, Kawabata S, Shibata S, Yasuda A, Nori S, Tsuji O, Nagoshi N, Iwanami A, Ebise H, Horiuchi K, Okano H, Nakamura M. 2017; Enhanced functional recovery from spinal cord injury in aged mice after stem cell transplantation through HGF induction. Stem Cell Reports. 8:509–18. DOI: 10.1016/j.stemcr.2017.01.013. PMID: 28216143. PMCID: PMC5355635.

4. Desai RA, Davies AL, Tachrount M, Kasti M, Laulund F, Golay X, Smith KJ. 2016; Cause and prevention of demyelination in a model multiple sclerosis lesion. Ann Neurol. 79:591–604. DOI: 10.1002/ana.24607. PMID: 26814844. PMCID: PMC4949637.

5. Haider L, Zrzavy T, Hametner S, Höftberger R, Bagnato F, Grabner G, Trattnig S, Pfeifenbring S, Brück W, Lassmann H. 2016; The topograpy of demyelination and neurodegeneration in the multiple sclerosis brain. Brain. 139(Pt 3):807–15. DOI: 10.1093/brain/awv398. PMID: 26912645. PMCID: PMC4766379.

6. Cohen-Adad J, El Mendili MM, Lehéricy S, Pradat PF, Blancho S, Rossignol S, Benali H. 2011; Demyelination and degeneration in the injured human spinal cord detected with diffusion and magnetization transfer MRI. Neuroimage. 55:1024–33. DOI: 10.1016/j.neuroimage.2010.11.089. PMID: 21232610.

7. Bouhrara M, Reiter DA, Bergeron CM, Zukley LM, Ferrucci L, Resnick SM, Spencer RG. 2018; Evidence of demyelination in mild cognitive impairment and dementia using a direct and specific magnetic resonance imaging measure of myelin content. Alzheimers Dement. 14:998–1004. DOI: 10.1016/j.jalz.2018.03.007. PMID: 29679574. PMCID: PMC6097903.

8. Mitew S, Kirkcaldie MT, Halliday GM, Shepherd CE, Vickers JC, Dickson TC. 2010; Focal demyelination in Alzheimer's disease and transgenic mouse models. Acta Neuropathol. 119:567–77. DOI: 10.1007/s00401-010-0657-2. PMID: 20198482.

9. Franklin RJ, Ffrench-Constant C. 2008; Remyelination in the CNS: from biology to therapy. Nat Rev Neurosci. 9:839–55. DOI: 10.1038/nrn2480. PMID: 18931697.

10. Yue T, Xian K, Hurlock E, Xin M, Kernie SG, Parada LF, Lu QR. 2006; A critical role for dorsal progenitors in cortical myelination. J Neurosci. 26:1275–80. DOI: 10.1523/JNEUROSCI.4717-05.2006. PMID: 16436615. PMCID: PMC6674567.

11. Lubetzki C, Zalc B, Williams A, Stadelmann C, Stankoff B. 2020; Remyelination in multiple sclerosis: from basic science to clinical translation. Lancet Neurol. 19:678–88. DOI: 10.1016/S1474-4422(20)30140-X. PMID: 32702337.

12. Melchor GS, Khan T, Reger JF, Huang JK. 2019; Remyelination pharmacotherapy investigations highlight diverse mechanisms underlying multiple sclerosis progression. ACS Pharmacol Transl Sci. 2:372–86. DOI: 10.1021/acsptsci.9b00068. PMID: 32259071. PMCID: PMC7088971.

13. Chen Y, Zhen W, Guo T, Zhao Y, Liu A, Rubio JP, Krull D, Richardson JC, Lu H, Wang R. 2017; Histamine receptor 3 negatively regulates oligodendrocyte differentiation and remyelination. PLoS One. 12:e0189380. DOI: 10.1371/journal.pone.0189380. PMID: 29253893. PMCID: PMC5734789.

14. Fan LW, Bhatt A, Tien LT, Zheng B, Simpson KL, Lin RC, Cai Z, Kumar P, Pang Y. 2015; Exposure to serotonin adversely affects oligodendrocyte development and myelination in vitro. J Neurochem. 133:532–43. DOI: 10.1111/jnc.12988. PMID: 25382136. PMCID: PMC4400220.

15. Ghareghani M, Sadeghi H, Zibara K, Danaei N, Azari H, Ghanbari A. 2017; Melatonin increases oligodendrocyte differentiation in cultured neural stem cells. Cell Mol Neurobiol. 37:1319–24. DOI: 10.1007/s10571-016-0450-4. PMID: 27987059.

16. Olivier P, Fontaine RH, Loron G, Van Steenwinckel J, Biran V, Massonneau V, Kaindl A, Dalous J, Charriaut-Marlangue C, Aigrot MS, Pansiot J, Verney C, Gressens P, Baud O. 2009; Melatonin promotes oligodendroglial maturation of injured white matter in neonatal rats. PLoS One. 4:e7128. DOI: 10.1371/journal.pone.0007128. PMID: 19771167. PMCID: PMC2742165.

17. Sotthibundhu A, Phansuwan-Pujito P, Govitrapong P. 2010; Melatonin increases proliferation of cultured neural stem cells obtained from adult mouse subventricular zone. J Pineal Res. 49:291–300. DOI: 10.1111/j.1600-079X.2010.00794.x. PMID: 20663047.

18. Sakurai T, Mieda M. 2011; Connectomics of orexin-producing neurons: interface of systems of emotion, energy homeostasis and arousal. Trends Pharmacol Sci. 32:451–62. DOI: 10.1016/j.tips.2011.03.007. PMID: 21565412.

19. Yin J, Babaoglu K, Brautigam CA, Clark L, Shao Z, Scheuermann TH, Harrell CM, Gotter AL, Roecker AJ, Winrow CJ, Renger JJ, Coleman PJ, Rosenbaum DM. 2016; Structure and ligand-binding mechanism of the human OX1 and OX2 orexin receptors. Nat Struct Mol Biol. 23:293–9. DOI: 10.1038/nsmb.3183. PMID: 26950369.

20. Ng MC. 2017; Orexin and epilepsy: potential role of REM sleep. Sleep. 40:zsw061. DOI: 10.1093/sleep/zsw061. PMID: 28364414.

21. Soya S, Takahashi TM, McHugh TJ, Maejima T, Herlitze S, Abe M, Sakimura K, Sakurai T. 2017; Orexin modulates behavioral fear expression through the locus coeruleus. Nat Commun. 8:1606. DOI: 10.1038/s41467-017-01782-z. PMID: 29151577. PMCID: PMC5694764. PMID: b6b7b0ef9da64f028089ee1720edac0e.

22. Liblau RS, Vassalli A, Seifinejad A, Tafti M. 2015; Hypocretin (orexin) biology and the pathophysiology of narcolepsy with cataplexy. Lancet Neurol. 14:318–28. DOI: 10.1016/S1474-4422(14)70218-2. PMID: 25728441.

23. Liguori C. 2017; Orexin and Alzheimer's disease. Curr Top Behav Neurosci. 33:305–22. DOI: 10.1007/7854_2016_50. PMID: 28012089.

24. Liu RJ, van den Pol AN, Aghajanian GK. 2002; Hypocretins (orexins) regulate serotonin neurons in the dorsal raphe nucleus by excitatory direct and inhibitory indirect actions. J Neurosci. 22:9453–64. DOI: 10.1523/JNEUROSCI.22-21-09453.2002. PMID: 12417670. PMCID: PMC6758063.

25. Lyons DJ, Hellysaz A, Ammari R, Broberger C. 2017; Hypocretin/orexin peptides excite rat neuroendocrine dopamine neurons through orexin 2 receptor-mediated activation of a mixed cation current. Sci Rep. 7:41535. DOI: 10.1038/srep41535. PMID: 28145492. PMCID: PMC5286397.

26. Tao R, Ma Z, McKenna JT, Thakkar MM, Winston S, Strecker RE, McCarley RW. 2006; Differential effect of orexins (hypocretins) on serotonin release in the dorsal and median raphe nuclei of freely behaving rats. Neuroscience. 141:1101–5. DOI: 10.1016/j.neuroscience.2006.05.027. PMID: 16820265.

27. Yamada N, Katsuura G, Tatsuno I, Asaki T, Kawahara S, Ebihara K, Saito Y, Nakao K. 2008; Orexin decreases mRNA expressions of NMDA and AMPA receptor subunits in rat primary neuron cultures. Peptides. 29:1582–7. DOI: 10.1016/j.peptides.2008.05.002. PMID: 18573570.

28. Yamanaka A, Tsujino N, Funahashi H, Honda K, Guan JL, Wang QP, Tominaga M, Goto K, Shioda S, Sakurai T. 2002; Orexins activate histaminergic neurons via the orexin 2 receptor. Biochem Biophys Res Commun. 290:1237–45. DOI: 10.1006/bbrc.2001.6318. PMID: 11811995.

29. Yin J, Mobarec JC, Kolb P, Rosenbaum DM. 2015; Crystal structure of the human OX2 orexin receptor bound to the insomnia drug suvorexant. Nature. 519:247–50. DOI: 10.1038/nature14035. PMID: 25533960.

30. Azari H, Sharififar S, Rahman M, Ansari S, Reynolds BA. 2011; Establishing embryonic mouse neural stem cell culture using the neurosphere assay. J Vis Exp. (47):2457. DOI: 10.3791/2457. PMID: 21248704. PMCID: PMC3182648.

31. Lassmann H, van Horssen J. 2011; The molecular basis of neurodegeneration in multiple sclerosis. FEBS Lett. 585:3715–23. DOI: 10.1016/j.febslet.2011.08.004. PMID: 21854776.

32. Geurts JJ, Bö L, Roosendaal SD, Hazes T, Daniëls R, Barkhof F, Witter MP, Huitinga I, van der Valk P. 2007; Extensive hippocampal demyelination in multiple sclerosis. J Neuropathol Exp Neurol. 66:819–27. DOI: 10.1097/nen.0b013e3181461f54. PMID: 17805012.

33. Kim LJ, Martinez D, Fiori CZ, Baronio D, Kretzmann NA, Barros HM. 2015; Hypomyelination, memory impairment, and blood-brain barrier permeability in a model of sleep apnea. Brain Res. 1597:28–36. DOI: 10.1016/j.brainres.2014.11.052. PMID: 25482664.

34. Pillai JA, Leverenz JB. 2017; Sleep and neurodegeneration: a critical appraisal. Chest. 151:1375–86. DOI: 10.1016/j.chest.2017.01.002. PMID: 28087304.

35. Mothe AJ, Tam RY, Zahir T, Tator CH, Shoichet MS. 2013; Repair of the injured spinal cord by transplantation of neural stem cells in a hyaluronan-based hydrogel. Biomaterials. 34:3775–83. DOI: 10.1016/j.biomaterials.2013.02.002. PMID: 23465486.

36. Wolswijk G. 1998; Chronic stage multiple sclerosis lesions contain a relatively quiescent population of oligodendrocyte precursor cells. J Neurosci. 18:601–9. DOI: 10.1523/JNEUROSCI.18-02-00601.1998. PMID: 9425002. PMCID: PMC6792542.

37. Yousefifard M, Rahimi-Movaghar V, Nasirinezhad F, Baikpour M, Safari S, Saadat S, Moghadas Jafari A, Asady H, Razavi Tousi SM, Hosseini M. 2016; Neural stem/progenitor cell transplantation for spinal cord injury treatment; a systematic review and meta-analysis. Neuroscience. 322:377–97. DOI: 10.1016/j.neuroscience.2016.02.034. PMID: 26917272.

38. Sakurai T, Amemiya A, Ishii M, Matsuzaki I, Chemelli RM, Tanaka H, Williams SC, Richardson JA, Kozlowski GP, Wilson S, Arch JR, Buckingham RE, Haynes AC, Carr SA, Annan RS, McNulty DE, Liu WS, Terrett JA, Elshourbagy NA, Bergsma DJ, Yanagisawa M. 1998; Orexins and orexin receptors: a family of hypothalamic neuropeptides and G protein-coupled receptors that regulate feeding behavior. Cell. 92:573–85. DOI: 10.1016/S0092-8674(00)80949-6. PMID: 9527442.

39. Recourt K, de Boer P, Zuiker R, Luthringer R, Kent J, van der Ark P, Van Hove I, van Gerven J, Jacobs G, van Nueten L, Drevets W. 2019; The selective orexin-2 antagonist seltorexant (JNJ-42847922/MIN-202) shows antidepressant and sleep-promoting effects in patients with major depressive disorder. Transl Psychiatry. 9:216. Erratum in: Transl Psychiatry 2019;9:240. DOI: 10.1038/s41398-019-0585-4. PMID: 31578318. PMCID: PMC6775147.

40. Zammit G, Dauvilliers Y, Pain S, Sebök Kinter D, Mansour Y, Kunz D. 2020; Daridorexant, a new dual orexin receptor antagonist, in elderly subjects with insomnia disorder. Neurology. 94:e2222–32. DOI: 10.1212/WNL.0000000000009475. PMID: 32341187.

41. Elam HB, Perez SM, Donegan JJ, Lodge DJ. 2021; Orexin receptor antagonists reverse aberrant dopamine neuron activity and related behaviors in a rodent model of stress-induced psychosis. Transl Psychiatry. 11:114. DOI: 10.1038/s41398-021-01235-8. PMID: 33558469. PMCID: PMC7870676. PMID: 98cbb80306624c4e97396c531721d1e7.

42. Brooks S, Jacobs GE, de Boer P, Kent JM, Van Nueten L, van Amerongen G, Zuiker R, Kezic I, Luthringer R, van der Ark P, van Gerven JM, Drevets W. 2019; The selective orexin-2 receptor antagonist seltorexant improves sleep: an exploratory double-blind, placebo controlled, crossover study in antidepressant-treated major depressive disorder patients with persistent insomnia. J Psychopharmacol. 33:202–9. DOI: 10.1177/0269881118822258. PMID: 30644312.

43. Dubey AK, Handu SS, Mediratta PK. 2015; Suvorexant: the first orexin receptor antagonist to treat insomnia. J Pharmacol Pharmacother. 6:118–21. DOI: 10.4103/0976-500X.155496. PMID: 25969666. PMCID: PMC4419247.

44. Kishi T, Matsunaga S, Iwata N. 2015; Suvorexant for primary insomnia: a systematic review and meta-analysis of randomized placebo-controlled trials. PLoS One. 10:e0136910. DOI: 10.1371/journal.pone.0136910. PMID: 26317363. PMCID: PMC4552781.

45. Scammell TE, Winrow CJ. 2011; Orexin receptors: pharmacology and therapeutic opportunities. Annu Rev Pharmacol Toxicol. 51:243–66. DOI: 10.1146/annurev-pharmtox-010510-100528. PMID: 21034217. PMCID: PMC3058259.

46. Roth T, Black J, Cluydts R, Charef P, Cavallaro M, Kramer F, Zammit G, Walsh J. 2017; Dual orexin receptor antagonist, almorexant, in elderly patients with primary insomnia: a randomized, controlled study. Sleep. 40:zsw034. DOI: 10.1093/sleep/zsw034. PMCID: PMC5806584. PMID: 28364509.

47. Braley TJ, Kratz AL, Kaplish N, Chervin RD. 2016; Sleep and cognitive function in multiple sclerosis. Sleep. 39:1525–33. DOI: 10.5665/sleep.6012. PMID: 27166237. PMCID: PMC4945311.

48. Bellesi M, Pfister-Genskow M, Maret S, Keles S, Tononi G, Cirelli C. 2013; Effects of sleep and wake on oligodendrocytes and their precursors. J Neurosci. 33:14288–300. DOI: 10.1523/JNEUROSCI.5102-12.2013. PMID: 24005282. PMCID: PMC3874087.

49. Andretic R, Franken P, Tafti M. 2008; Genetics of sleep. Annu Rev Genet. 42:361–88. DOI: 10.1146/annurev.genet.42.110807.091541. PMID: 18983259.

50. Bellesi M, Haswell JD, de Vivo L, Marshall W, Roseboom PH, Tononi G, Cirelli C. 2018; Myelin modifications after chronic sleep loss in adolescent mice. Sleep. 41:zsy034. DOI: 10.1093/sleep/zsy034. PMID: 29741724. PMCID: PMC5946929.

51. Becquet L, Abad C, Leclercq M, Miel C, Jean L, Riou G, Couvineau A, Boyer O, Tan YV. 2019; Systemic administration of orexin A ameliorates established experimental autoimmune encephalomyelitis by diminishing neuroinflammation. J Neuroinflammation. 16:64. DOI: 10.1186/s12974-019-1447-y. PMID: 30894198. PMCID: PMC6425555. PMID: 69e7f6833b604226b92795a7fa86ab95.

52. Fatemi I, Shamsizadeh A, Ayoobi F, Taghipour Z, Sanati MH, Roohbakhsh A, Motevalian M. 2016; Role of orexin-A in experimental autoimmune encephalomyelitis. J Neuroimmunol. 291:101–9. DOI: 10.1016/j.jneuroim.2016.01.001. PMID: 26857503.

53. Gencer M, Akbayır E, Şen M, Arsoy E, Yılmaz V, Bulut N, Tüzün E, Türkoğlu R. 2019; Serum orexin-A levels are associated with disease progression and motor impairment in multiple sclerosis. Neurol Sci. 40:1067–70. DOI: 10.1007/s10072-019-3708-z. PMID: 30645749.

54. Long KLP, Breton JM, Barraza MK, Perloff OS, Kaufer D. 2021; Hormonal regulation of oligodendrogenesis I: effects across the lifespan. Biomolecules. 11:283. DOI: 10.3390/biom11020283. PMID: 33672939. PMCID: PMC7918364. PMID: 394ed4eb50074feab425dcbd3ffb6324.

55. Armada-Moreira A, Ribeiro FF, Sebastião AM, Xapelli S. 2015; Neuroinflammatory modulators of oligodendrogenesis. Neuroimmunol Neuroinflammation. 2:263–73. DOI: 10.4103/2347-8659.167311.

56. Li H, He Y, Richardson WD, Casaccia P. 2009; Two-tier transcriptional control of oligodendrocyte differentiation. Curr Opin Neurobiol. 19:479–85. DOI: 10.1016/j.conb.2009.08.004. PMID: 19740649. PMCID: PMC2826212.

57. Copray S, Balasubramaniyan V, Levenga J, de Bruijn J, Liem R, Boddeke E. 2006; Olig2 overexpression induces the in vitro differentiation of neural stem cells into mature oligodendrocytes. Stem Cells. 24:1001–10. DOI: 10.1634/stemcells.2005-0239. PMID: 16253982.

58. Mei F, Wang H, Liu S, Niu J, Wang L, He Y, Etxeberria A, Chan JR, Xiao L. 2013; Stage-specific deletion of Olig2 conveys opposing functions on differentiation and maturation of oligodendrocytes. J Neurosci. 33:8454–62. DOI: 10.1523/JNEUROSCI.2453-12.2013. PMID: 23658182. PMCID: PMC3865513.

59. Raasakka A, Myllykoski M, Laulumaa S, Lehtimäki M, Härtlein M, Moulin M, Kursula I, Kursula P. 2015; Determinants of ligand binding and catalytic activity in the myelin enzyme 2',3'-cyclic nucleotide 3'-phosphodiesterase. Sci Rep. 5:16520. DOI: 10.1038/srep16520. PMID: 26563764. PMCID: PMC4643303.

60. Rösener M, Muraro PA, Riethmüller A, Kalbus M, Sappler G, Thompson RJ, Lichtenfels R, Sommer N, McFarland HF, Martin R. 1997; 2',3'-cyclic nucleotide 3'-phosphodiesterase: a novel candidate autoantigen in demyelinating diseases. J Neuroimmunol. 75:28–34. DOI: 10.1016/S0165-5728(96)00230-5. PMID: 9143234.

61. Scherer SS, Braun PE, Grinspan J, Collarini E, Wang DY, Kamholz J. 1994; Differential regulation of the 2',3'-cyclic nucleotide 3'-phosphodiesterase gene during oligodendrocyte development. Neuron. 12:1363–75. DOI: 10.1016/0896-6273(94)90451-0. PMID: 8011341.

62. Raasakka A, Kursula P. 2014; The myelin membrane-associated enzyme 2',3'-cyclic nucleotide 3'-phosphodiesterase: on a highway to structure and function. Neurosci Bull. 30:956–66. DOI: 10.1007/s12264-013-1437-5. PMID: 24807122. PMCID: PMC5562554.