Immunodominant fragments of myelin basic protein initiate T cell-dependent pain

Huaqing Liu 1 2 Sergey A Shiryaev 0 Andrei V Chernov 0 Youngsoon Kim 1 2 Igor Shubayev 1 Albert G Remacle 0 Svetlana Baranovskaya 3 Vladislav S Golubkov 0 Alex Y Strongin 0 Veronica I Shubayev 1 2 0 Sanford-Burnham Medical Research Institute , La Jolla, CA , USA 1 VA San Diego Healthcare System , La Jolla, CA , USA 2 Department of Anesthesiology, University of California , San Diego, 9500 Gilman Dr., Mail Box 0629, La Jolla, CA 92093-0629 , USA 3 Agilent Technologies , La Jolla, CA , USA - Immunodominant fragments of myelin basic protein initiate T cell-dependent pain Background: The myelin sheath provides electrical insulation of mechanosensory A-afferent fibers. Myelin-degrading matrix metalloproteinases (MMPs) damage the myelin sheath. The resulting electrical instability of A-fibers is believed to activate the nociceptive circuitry in A-fibers and initiate pain from innocuous tactile stimulation (mechanical allodynia). The precise molecular mechanisms, responsible for the development of this neuropathic pain state after nerve injury (for example, chronic constriction injury, CCI), are not well understood. Methods and results: Using mass spectrometry of the whole sciatic nerve proteome followed by bioinformatics analyses, we determined that the pathways, which are classified as the Infectious Disease and T-helper cell signaling, are readily activated in the nerves post-CCI. Inhibition of MMP-9/MMP-2 suppressed CCI-induced mechanical allodynia and concomitant TNF- and IL-17A expression in nerves. MMP-9 proteolysis of myelin basic protein (MBP) generated the MBP84-104 and MBP68-86 digest peptides, which are prominent immunogenic epitopes. In agreement, the endogenous MBP69-86 epitope co-localized with MHCII and MMP-9 in Schwann cells and along the nodes of Ranvier. Administration of either the MBP84-104 or MBP68-86 peptides into the nave nerve rapidly produced robust mechanical allodynia with a concomitant increase in T cells and MHCII-reactive cell populations at the injection site. As shown by the genome-wide expression profiling, a single intraneural MBP84-104 injection stimulated the inflammatory, immune cell trafficking, and antigen presentation pathways in the injected nave nerves and the associated spinal cords. Both MBP84-104-induced mechanical allodynia and characteristic pathway activation were remarkably less prominent in the T cell-deficient athymic nude rats. Conclusions: These data implicate MBP as a novel mediator of pain. Furthermore, the action of MMPs expressed within 1 day post-injury is critical to the generation of tactile allodynia, neuroinflammation, and the immunodominant MBP digest peptides in nerve. These MBP peptides initiate mechanical allodynia in both a T cell-dependent and -independent manner. In the course of Wallerian degeneration, the repeated exposure of the cryptic MBP epitopes, which are normally sheltered from immunosurveillance, may induce the MBP-specific T cell clones and a self-sustaining immune reaction, which may together contribute to the transition of acute pain into a chronic neuropathic pain state. Background Pain is typically mediated by small unmyelinated C-nociceptive and thinly myelinated A-afferents. Non-nociceptive, large-diameter myelinated A-afferents transmit touch and vibration sense. However, following damage to the peripheral nervous system (PNS), A-afferents join nociceptive circuitry [1]. Our mechanistic understanding * Correspondence: 1Department of Anesthesiology, University of California, San Diego, 9500 Gilman Dr., Mail Box 0629, La Jolla, CA 92093-0629, USA 2VA San Diego Healthcare System, La Jolla, CA, USA Full list of author information is available at the end of the article of why the damaged A-afferents interpret an innocuous, low-threshold tactile stimulus as painful (that is, mechanical allodynia) remains exceedingly limited. Growing evidence supports a model in which the damage to the electrically insulating myelin sheath and the resulting loss of electrical stability in A-afferents contribute to the development of mechanical allodynia [2-6]. Mechanical allodynia and other forms of neuropathic pain (NP, that is, pain arising as a direct consequence of a lesion or disease affecting the somatosensory nervous system [7]) have features of a neuroimmune disorder [8]. T lymphocyte infiltration into both the damaged nerve [9-11] and the spinal cord at a corresponding segment [12-14] has been implicated in NP. Following chronic constriction injury (CCI), T cell-deficient athymic nude rats exhibit a diminished ability to develop NP, which is reversed with adoptive transfer of T-helper (Th)1 cells [9]. Interleukin (IL)-17, expressed by certain Th cells, is essential to mediating mechanical, but not thermal, hypersensitivity [15]. Major histocompatibility complex (MHC) II, produced by antigen-presenting cells to capture and present antigens to T cells, is required for the development of NP [16,17]. An increase in MHCII is observed in the nerve and the corresponding DRG [18], spinal cord [14,16], and brainstem [19] after a peripheral nerve lesion. However, the antigens, which are involved in the recruitment and the homing of activated T cells in the pathogenesis of NP, remain unknown. Myelin basic protein (MBP) is the component of the compact myelin that is believed to participate in the maintenance of the major dense line and interactions of the myelin sheath with the cytosolic surfaces [20-23]. In the PNS, MBP comprises 5% to 15% of total myelin protein and is considered to be non-essential [22]. However, Th1-mediated autoimmune peripheral neuropathies in humans and the relevant experimental models induced by immunization of animals using immunodominant MBP and other myelin peptides are often painful [24]. It has been demonstrated that the autoimmune response to immunodominant MBP peptides assists in myelin clearance and regeneration after peripheral nerve injury [25]. Certain digest fragments of MBP and its splice variant (Golli-MBP) expressed in immune cells [26] are generated by matrix metalloproteinase (MMP) proteolysis and exhibit key T cell epitopes [27,28]. MMPs are a family of zinc-endopeptidases comprising collagenases, gelatinases, matrilysins, stromelysins, and membrane-type MMPs [29]. After peripheral nerve injury, gelatinases B (MMP-9) and A (MMP-2) degrade the bloodnerve barrier, release the pro-inflammatory cytokines, control immune cell infiltration and cell survival along the injured neural axis. These two MMPs are believed to consecutively initiate and maintain NP [5,30-34]. Having proposed that MMPs promote mechanical hypersensitivity via the proteolysis of myelin [5], we herein aimed to determine the specific mechanisms involved. Our present experimental evidence suggests that MMP-mediated fragmentation of MBP as a consequence of Wallerian degeneration exposes cryptic MBP epitopes, which are normally sheltered from immunosurveillance. These exposed immunodominant MBP peptide (...truncated)