The online version of this article (https://doi.org/10.1186/s12974-017-1034-z ) contains supplementary material, which is available to authorized users.
Microglia are major players in the pathogenesis of multiple sclerosis (MS) and may play a dual role in disease progression. The activation status of microglia in vivo is highly dynamic and occurs as a continuum, with the pro-inflammatory and anti-inflammatory phenotypes on either end of this spectrum. Little is known about in vivo dynamics of microglia phenotypes in MS due to the lack of diagnostic tools. Positron emission tomography (PET) imaging is a powerful non-invasive technique that allows real-time imaging of microglia activation phenotypes in the central nervous system, depending on the availability of selective PET tracers. Our objective is to investigate and characterize the expression of the purinergic receptors P2Y12R and P2X7R as potential targets for PET tracer development and subsequent PET imaging in order to evaluate the dynamics of microglia status in vivo.
We used immunohistochemical analysis to explore the expression of P2Y12R and P2X7R in experimental autoimmune encephalomyelitis (EAE) post-mortem tissues and different stages of well-characterized MS lesions. We evaluated by quantitative real-time polymerase chain reaction the expression of P2Y12R and P2X7R in human polarized microglia, and we performed autoradiography binding assay with radiolabeled P2Y12R and P2X7R antagonists using MS and rat EAE tissues.
Here, we demonstrate that P2X7R is associated with a pro-inflammatory phenotype of human microglia in vitro, and is highly expressed in microglia in MS lesions as well as during the peak of EAE. In contrast, P2Y12R was associated with an anti-inflammatory phenotype in human microglia in vitro and was expressed at lower levels in active inflammatory MS lesions compared to normal-appearing white matter (NAWM) and similarly in EAE, while its expression increased in the remission phase of EAE. Binding of radiolabeled tracers specific for P2Y12R and P2X7R on ex vivo tissues validated the value of these receptors as PET imaging targets for microglia phenotypes in vivo.
Our results suggest that P2Y12R and P2X7R are excellent targets for PET imaging to discriminate distinct microglia phenotypes in MS. Ultimately, this may provide insight into the role of microglia in disease progression and monitor novel treatment strategies to alter microglia phenotype.
Peferoen LAN, Vogel DYS, Ummenthum K, Breur M, Heijnen, Priscilla DAM, Gerritsen WH, et al. Activation status of human microglia is dependent on lesion formation stage and remyelination in multiple sclerosis. J Neuropathol Exp Neurol. 2015;74:48–63. https://doi.org/10.1097/NEN.0000000000000149 . CrossRefPubMed
Parkhurst CN, Gan W. Microglia dynamics and function in the CNS. Curr Opin Neurobiol. 2010;20:595–600. https://doi.org/10.1016/j.conb.2010.07.002. CrossRefPubMedPubMedCentral
Kotter MR, Li W, Zhao C, Franklin RJM. Myelin impairs CNS remyelination by inhibiting oligodendrocyte precursor cell differentiation. J Neurosci. 2006;26:328–32. https://doi.org/10.1523/JNEUROSCI.2615-05.2006 . CrossRefPubMed
Butovsky O, Ziv Y, Schwartz A, Landa G, Talpalar AE, Pluchino S, et al. Microglia activated by IL-4 or IFN-gamma differentially induce neurogenesis and oligodendrogenesis from adult stem/progenitor cells. Mol Cell Neurosci. 2006;31:149–60. https://doi.org/10.1016/j.mcn.2005.10.006 . CrossRefPubMed
Yu Y, Ugawa S, Ueda T, Ishida Y, Inoue K, Kyaw Nyunt A, et al. Cellular localization of P2X7 receptor mRNA in the rat brain. Brain. 2008;1194:45–55. https://doi.org/10.1016/j.brainres.2007.11.064 . CrossRef
Clark AK, Staniland AA, Marchand F, Kaan TKY, McMahon SB, Malcangio M. P2X7-dependent release of interleukin-1 beta and nociception in the spinal cord following lipopolysaccharide. J Neurosci. 2010;30:573–82. https://doi.org/10.1523/JNEUROSCI.3295-09.2010 . CrossRefPubMedPubMedCentral
Monif M, Burnstock G, Williams DA. Microglia: proliferation and activation driven by the P2X7 receptor. Int J Biochem Cell Biol. 2010;42:1753–6. https://doi.org/10.1016/j.biocel.2010.06.021 . CrossRefPubMed
Janssen B, Vugts DJ, Funke U, Perry S, Kruijer, Dolle F, Bormans G, et al. Synthesis of the first carbon-11 labelled P2Y12 receptor antagonist for PET imaging of microglial activation in neuroinflammation. J Label Compd Radiopharm. 2015;58:S1–S74. https://doi.org/10.1002/jlcr.3302_1 . CrossRef
de Vries HE, JJA H, Honing H, de Lavalette CR, SMA v d P, Hooijberg E, et al. Signal-regulatory protein -CD47 interactions are required for the transmigration of monocytes across cerebral endothelium. J Immunol. 2002;168:5832–9. https://doi.org/10.4049/jimmunol.168.11.5832 . CrossRefPubMed
Garcia-Vallejo JJ, van Dijk W, van Het Hof B, van Die I, Engelse MA, Hinsbergh V, Victor WM, Gringhuis SI. Activation of human endothelial cells by tumor necrosis factor-alpha results in profound changes in the expression of glycosylation-related genes. J Cell Physiol. 2006;206:203–10. https://doi.org/10.1002/jcp.20458 . CrossRefPubMed
Matute C, Torre I, Perez-Cerda F, Perez-Samartin A, Alberdi E, Etxebarria E, et al. P2X(7) receptor blockade prevents ATP excitotoxicity in oligodendrocytes and ameliorates experimental autoimmune encephalomyelitis. J Neurosci. 2007;27:9525–33. https://doi.org/10.1523/JNEUROSCI.0579-07.2007 . CrossRefPubMed
Yiangou Y, Facer P, Durrenberger P, Chessell IP, Naylor A, Bountra C, et al. COX-2, CB2 and P2X7-immunoreactivities are increased in activated microglial cells/macrophages of multiple sclerosis and amyotrophic lateral sclerosis spinal cord. BMC Neurol. 2006;6:12. https://doi.org/10.1186/1471-2377-6-12. CrossRefPubMedPubMedCentral
Ruuls SR, Sedgwick JD. Cytokine-directed therapies in multiple sclerosis and experimental autoimmune encephalomyelitis. Immunol Cell Biol. 1998;76:65–73. https://doi.org/10.1046/j.1440-1711.1998.00715.x . CrossRefPubMed
Moore CS, Ase AR, Kinsara A, Rao VTS, Michell-Robinson M, Leong SY, et al. P2Y12 expression and function in alternatively activated human microglia. Neurol Neuroimmunol Neuroinflamm. 2015;2:e80. https://doi.org/10.1212/NXI.0000000000000080. CrossRefPubMedPubMedCentral
Ory D, Celen S, Gijsbers R, van den Haute C, Postnov A, Koole M, et al. Preclinical evaluation of a P2X7 receptor-selective radiotracer: PET studies in a rat model with local overexpression of the human P2X7 receptor and in nonhuman primates. J Nucl Med. 2016;57:1436–41. https://doi.org/10.2967/jnumed.115.169995 . CrossRefPubMed
- Purinergic receptors P2Y12R and P2X7R: potential targets for PET imaging of microglia phenotypes in multiple sclerosis
Susanne M. A. van der Pol
B. van Het Hof
Jack van Horssen
Albert D. Windhorst
Helga E. de Vries
- BioMed Central