Astrocytic tight junctions control inflammatory CNS lesion pathogenesis
Sam Horng, … , Candice Chapouly, Gareth R. John
Sam Horng, … , Candice Chapouly, Gareth R. John
Published July 24, 2017
Citation Information: J Clin Invest. 2017;127(8):3136-3151. https://doi.org/10.1172/JCI91301.
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Research Article Immunology Neuroscience

Astrocytic tight junctions control inflammatory CNS lesion pathogenesis

Abstract

Lesions and neurologic disability in inflammatory CNS diseases such as multiple sclerosis (MS) result from the translocation of leukocytes and humoral factors from the vasculature, first across the endothelial blood-brain barrier (BBB) and then across the astrocytic glia limitans (GL). Factors secreted by reactive astrocytes open the BBB by disrupting endothelial tight junctions (TJs), but the mechanisms that control access across the GL are unknown. Here, we report that in inflammatory lesions, a second barrier composed of reactive astrocyte TJs of claudin 1 (CLDN1), CLDN4, and junctional adhesion molecule A (JAM-A) subunits is induced at the GL. In a human coculture model, CLDN4-deficient astrocytes were unable to control lymphocyte segregation. In models of CNS inflammation and MS, mice with astrocyte-specific Cldn4 deletion displayed exacerbated leukocyte and humoral infiltration, neuropathology, motor disability, and mortality. These findings identify a second inducible barrier to CNS entry at the GL. This barrier may be therapeutically targetable in inflammatory CNS disease.

Authors

Sam Horng, Anthony Therattil, Sarah Moyon, Alexandra Gordon, Karla Kim, Azeb Tadesse Argaw, Yuko Hara, John N. Mariani, Setsu Sawai, Per Flodby, Edward D. Crandall, Zea Borok, Michael V. Sofroniew, Candice Chapouly, Gareth R. John

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Figure 2

CLDN4 and JAM-A are expressed within TJ strands of reactive astrocytes in vivo.

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CLDN4 and JAM-A are expressed within TJ strands of reactive astrocytes i...
(A) Immunostaining within an AdIL-1 lesion at 7 dpi for GFAP (green), the basement membrane marker pan-laminin (blue), and the astrocytic TJ protein CLDN4 (red) demonstrates CLDN4 expression at reactive astrocytic endfeet surrounding the vasculature (arrows). Scale bar: 10 μm. See also Supplemental Figure 1C. (B) Transmission electron microscopy of astrocytic endfeet within cortical AdIL-1 and AdCtrl injection sites demonstrates TJs in AdIL-1 lesions but not in controls. Immunogold staining shows colocalization of CLDN4 and JAM-A to the TJ structures (arrows point to gold particles). A, astrocyte; EC, endothelial cell. Original magnification, ×10,000. (C) Immunostaining within an EAE lesion at 21 days for GFAP (blue), pan-laminin (red), and CLDN4 (green) demonstrates the structural organization of the reactive GL. This cross section shows basement membranes of the endothelial BBB (EBM) and astrocytic GL (ABM), demarcated by pan-laminin staining and differentiated by astrocytic endfeet, stained by GFAP and CLDN4. Leukocytes, identified in gray based on morphologic features and DAPI nuclear staining, are seen within the endothelial lumen (LUM) and PVS. White arrows highlight colocalization of CLDN4 and GFAP; pink arrows indicate areas of irregular CLDN4 staining, possibly reflecting irregularities of expression in the plane of staining or degradation in proximity to leukocytes. Scale bars: 10 μm. See also Supplemental Figure 1, D and E. (D) Schematic of the endothelial BBB and astrocytic GL in health and inflammatory disease. Under healthy conditions, endothelial cells express TJ proteins CLDN5 and occludin (OCLN), which reinforce a closed BBB. In response to inflammation, CLDN5 and OCLN are downregulated, opening the BBB. In turn, astrocytes of the GL upregulate TJ proteins CLDN1, CLDN4, and JAM-A, closing the GL and restricting incoming leukocytes to the PVS (blue). Data are representative of findings from at least 3 (A–C) biological replicates.