Animal models of immune-mediated demyelinating polyneuropathies

Abstract

Immune-mediated demyelinating polyneuropathies (IMDPs) are rare disorders in which dysregulated adaptive immune responses cause peripheral nerve demyelinating inflammation and axonal injury in susceptible individuals. Despite significant advances in understanding IMDP pathogenesis guided by patient data and representative mammalian models, specific therapies are lacking. Significant knowledge gaps in IMDP pathogenesis still exist, e.g. precise antigen(s) and mechanisms that initially trigger immune system activation and identification of large population disease susceptibility factors. The initial directional cues for antigen-specific effector or autoreactive leukocyte trafficking into peripheral nerves are also unknown. An overview of current animal models, with emphasis on the experimental autoimmune neuritis and spontaneous autoimmune peripheral polyneuropathy models, is provided. Insights on the initial directional cues for peripheral nerve tissue specific autoimmunity using a novel Major Histocompatibility Complex class II conditional knockout mouse strain are also discussed, suggesting an essential research tool to study cell- and time-dependent adaptive immunity in autoimmune diseases.

Keywords: Acute inflammatory demyelinating polyradiculoneuropathy; animal models; blood-nerve barrier; chronic inflammatory demyelinating polyradiculoneuropathy; demyelinating neuritis; experimental autoimmune neuritis; major histocompatibility complex; mice; pathogenesis; spontaneous autoimmune peripheral polyneuropathy.

Figure 1.. Severe murine experimental autoimmune neuritis in adult female SJL mice.

Representative digital photomicrographs of axial cryostat 10 μm and glutaraldehyde-fixed, osmium tetroxide post-fixed, epoxy resin-embedded, toluidine blue-stained 1 μm sciatic nerve sections from sm-EAN-affected female SJL mice 2-3 days after disease onset (A-F) and at peak severity (G-L) are shown. Foci of Cd4+ T-cells (A, white arrows) associated with rare Cd19+ B-cells (B, white arrows) and more diffuse foci of F4/80+ macrophages (C, white arrows) are seen during the early stages of sm-EAN. Infiltrating leukocytes are associated with Schwann cell membranes (D, white arrows) with some focal reduction in axon density (E, white arrows). Foci of demyelination with axonal degeneration occur in association with endoneurial leukocyte infiltration (F, asterisk). At peak severity, there is further leukocyte infiltration with more diffuse foci of Cd4+ T-cells (G, white arrows), persistent Cd19+ B-cell infiltration (H, white arrows) and increased foci or near confluent infiltration of F4/80+ monocytes/ macrophages (I, white arrows). Leukocyte infiltration is associated with foci or near confluent loss of Schwann cell membranes (J, white arrows), consistent with demyelination, as well as reduction in axonal density (K, white arrows), consistent with axonal loss. Multiple foci or near confluent regional demyelination and axonal degeneration (L, asterisk) occurs with endoneurial leukocyte infiltration at peak severity. S100β: marker of myelinating Schwann cell membranes; NF-H: Neurofilament-heavy chain, marker of axon intermediate filaments, TB: toluidine blue. Scale bars: A, B, F, G, H, K, L = 100 μm and C, D, E, I, J = 50 μm.

Figure 2.. Severe murine experimental autoimmune neuritis in adult female H2-Aa^flox/flox; vWF-iCre/+ SJL mice.

Representative photomicrographs of axial cryostat 10 μm sciatic nerve sections from sm-EAN-affected adult female wildtype SJL mouse 3 days after disease-onset demonstrates increased MHC class II (H2-A) expression in peripheral nerve endoneurium (A) with some co-localization with Cd31+ endothelium (B and C, white arrows). Higher magnification images of an endoneurial microvessel from the same mouse show mononuclear cell diapedesis associated with H2-A membrane expression (D-G). Diffusely increased MHC class II (H2-A) expression is observed in the endoneurium at sm-EAN peak severity in an adult female Tamoxifen-treated H2-Aa^flox/flox; +/+ SJL mouse, associated with Cd45+ leukocyte infiltration (H-J, white arrows). In contrast, an adult female Tamoxifen-treated H2-Aa^flox/flox; vWF-iCre/+ SJL mice with microvascular endothelial cell specific MHC class II deletion after Cre-mediated recombination does not develop sm-EAN, with rare Cd45+ leukocytes observed outside the endoneurium (K, white arrows). Increased MHC class II expression is observed in the perineurium (p), as well as lymphoid and non-lymphoid organs (not shown), supporting the notion that microvascular endothelial cell MHC class II expression is necessary for tissue- or organ-specific autoimmunity.

Figure 3.. Hypothetical initial antigen-specific effector memory CD4+ Th1 T-cell trafficking at the blood-nerve barrier.

Guided by published data from human in vitro leukocyte trafficking assays across microvascular endothelial cells from other organs, and unpublished data from untreated adult AIDP patient single cell transcriptomics compared to healthy age- and sex-matched controls, recently activated antigen-specific effector memory CD4+ Th1 T-cells roll on the endoneurial endothelium luminal surface via interactions between P-Selectin Glycoprotein-1 interacting with P-Selectin, followed haptotaxis driven by chemokines constitutively expressed by endoneurial endothelial cells (e.g. CX3CL1) bound to specific glycosaminoglycan that attract leukocytes expressing the corresponding chemokine receptor (e.g. CX3CR1). Effector memory CD4+ T-cells then engage with endothelial cells expressing endogenous peripheral nerve-derived peptides on the MHC class II α-chain via its TCRαβ complex. If there is a mismatch, the effector memory CD4+ T-cell disengages from the peripheral nerve microvascular endothelium surface back into circulation. If there is a match, co-stimulation (e.g. via CD40L-CD40 or ICOS-ICOSL interactions) activates the antigen-specific effector CD4+ T-cell with consequential conformational change in leukocyte integrins (e.g. CD11b, CD49d) from inactive to active forms that facilitate firm adhesion to the endothelium by via binding to selective cell adhesion molecules (e.g. ICAM-1, VCAM-1). Effector memory CD4+ T-cells then develop protrusions and crawl to inter-endothelial cell junctions, and undergo diapedesis through homophilic interactions (e.g. CD99) with the endoneurial endothelial cell junctional membrane. Matrix metalloproteinases (e.g. MMP2, MMP9 [not shown]) are secreted from the leading protrusion to break down the endothelium basement membrane to permit effector memory CD4+ T-cell entry into the endoneurium.

Figure 4.. Spontaneous autoimmune peripheral polyneuropathy in adult female Cd86−/− NOD mice.

Representative photomicrographs of axial cryostat 10 μm and glutaraldehyde-fixed, osmium tetroxide post-fixed, epoxy resin-embedded, toluidine blue-stained 1 μm sciatic nerve sections from SAPP-affected adult female Cd86^−/− NOD mice at 22 weeks (early phase, 2 weeks after symptomatic onset; A-D) and 40 weeks (maximum severity plateau phase, 18 weeks after symptomatic onset, E-H) of age are shown. At 22 weeks of age, there is early infiltration of Cd11b+ (red) Cd45+ (green) leukocytes, consistent with monocytes (white arrows), as well as Cd11b− Cd45+ lymphocytes (l) into the endoneurium, as shown in A). Cd11b+ Cd45− cells, consistent with Schwann cells (sc) also observed. Foci of demyelination (asterisk, B) and reduction in axonal density associated mononuclear cell infiltration (white arrows, C) are seen during the early phases of SAPP, supported by epoxy resin-embedded sections showing mononuclear cellular infiltrates (white arrows, D) with normal appearing axons. At 40 weeks of age, there is more diffuse multi-focal infiltration of Cd11b+ CD45+ monocytes/ macrophages (white arrows) associated with Schwann cell (sc) proliferation and increased numbers of lymphocytes (l) in the endoneurium, as shown in E. This is associated with multi-focal demyelination (asterisk, F) and reduction in axonal density (white arrows, G). Mononuclear cell infiltration, associated with completely demyelinated axons (white arrows) and demyelinated axons surrounded by concentric layers of Schwann cell membranes (i.e. onion bulbs, yellow arrows), indicative of repeated demyelination-remyelination, is shown in H. Endoneurial microvessels (emv) with perivascular mononuclear cells and an axon undergoing Wallerian degeneration (Wd) are also shown. S100β: marker of myelinating Schwann cell membranes; NF-H: Neurofilament-heavy chain, marker of axon intermediate filaments, TB: toluidine blue. Scale bars: A-C = 100 μm and D-H = 50 μm.