IMMUNOCOMPETENCE IN THE CNS
The CNS was long considered an immunologically privileged site, a place wherein the immune system performed few functions. Over the past decade, this view has altered dramatically. The CNS is actively involved in immunological phenomena that are physiological and pathological. Work in our laboratory has focused on the mechanisms by which inflammation develops in the CNS in response to peripheral nerve and root injuries. Toward this end, we have utilized rodent models of neuro- pathic and radicular pain previously discussed in this review. In our dissection of these immunologically distinct systems, we have investigated topics relative to cell migration and trafficking, adhesion molecule expression, antigen presentation, and cytokine production. The recent discovery that the CNS broadly responds similarly, albeit not exactly, in an immunologic fashion to autoimmunity, trauma, brain abscess, and peripheral nerve or root injury that leads to chronic pain is highly innovative.
For the first time, the mechanisms of CNS cellular glial and neuronal activation and leukocyte trafficking in CNS reactions following peripheral nerve and nerve root injury are being directly compared in the same laboratory.
6.4.1 POTENTIAL ROLE OF IMMUNOCOMPETENT GLIA
As alluded to earlier in this review, cytokines may play an important role in the development of central sensitization through their interaction with glia. Glial cells (microglia, astrocytes, and oligodendrocytes) constitute over 70% of the total cell population in the brain and spinal cord. Once thought of as merely a physical support system for neurons, glial cells have recently come under intense scrutiny as key neuromodulatory, neurotrophic, and neuroimmune elements in the CNS. Microglia, cells of monocytic origin, are the macrophages of the brain and as such perform a vast number of immune-related duties.89 Pathological stimuli provoke a graded transformation of microglia from a highly ramified resting surveillance state ulti- mately to a phagocytic macrophage. Microglial activation involves a stereotypic pattern of cellular responses, such as proliferation, increased expression of immuno- molecules, recruitment to the site of injury, and functional changes that include the release of cytotoxic and/or inflammatory mediators. The initial signal for microglial activation is not well understood; however, following injury, neuronal depolarization combined with extracellular ion changes may be major stimuli.90 Alternatively, neuronal signals such as nitric oxide (NO) or proinflammatory cytokines may provide the stimuli for this activation.6,91 During autoimmune inflammation of the nervous system, microglia both release and respond to several cytokines including TNF, IL-1β, and IL-6, all of which are instrumental in astrocyte activation, induction of cell adhesion molecule expression, and recruitment of T-cells into the lesion. In addition to the synthesis of inflammatory cytokines, microglia act as cytotoxic effector cells by releasing harmful substances including proteases, reactive oxygen intermediates, and NO.
Glial changes in response to injury include proliferation and hypertrophy of astrocytic and microglia cells and overexpression of GFAP. GFAP increases in the spinal cord following different nerve injuries such as chronic constriction injury, nerve crush, and axotomy.92–94 Following a peripheral nerve freeze lesion, immuno- reactive GFAP expression increases at 14 d with a second major increase at 42 d, consistent with peaks in autotomy behavior and mechanical allodynia, respectively.36 Glia are, indeed, intimately involved in the neuroimmune network following periph- eral nerve injury. However, the concept that all glia are producing the same delete- rious proteins and that ameliorating their function will have a beneficial outcome for persistent pain appears nạve. Using immunocytochemistry, we have demon- strated that only a subpopulation of astrocytes co-localize with specific cytokines and that a subpopulation of microglia express major histocompatibility complex class II (MHC II) and CD4+ expression.
6.4.2 MAJOR HISTOCOMPATIBILITY COMPLEX
The MHC is a region of highly polymorphic genes whose products are expressed on the surfaces of a variety of cells. MHC genes play a central role in immune responses to protein antigens. The principal function of MHC molecules is to bind fragments of foreign proteins, thereby forming complexes that can be recognized by T-lymphocytes. T-lymphocytes do not recognize antigens in free or soluble form but instead only recognize portions of protein antigens (i.e., peptides) complexed to MHC. In the periphery, CD8 expression by T-cells is strongly correlated with MHC I, and CD4 T-helper cells are strongly correlated with MHC II. Having defined the role of MHC in the periphery, its role in the CNS is difficult to discern. In normal CNS, both MHC I and MHC II expression is minimal compared to other tissues.89,95,96 This low expression of MHC and reduced immune surveillance are believed to contribute to the immune-privileged status of non-renewable CNS neurons.
However, microglia and astrocytes clearly express MHC II after neuronal and axonal degeneration.97–99 The significance of this expression is still unclear and may involve functions other than antigen presentation. It has been recognized that a subpopulation of microglia become immunocompetent in response to infection or injury. One way in which this occurs is through the expression of MHC II. Cytokines are key modulators of MHC I and II genes in a wide variety of cells. Of relevance to the discussion of cytokines and pain processing, glia do not normally express MHC II but its expression on glial cells can be induced by cytokines such as TNF.100 The mediators that act to alter the expression of the MHC II antigens are tissue specific. This has implications for possible selective immunomodulation of MHC II in tissues in which it is overexpressed without affecting MHC II expression in other tissues. We have preliminary data showing MHC II expression on cells with glial morphology after peripheral nerve injury (Fig. 6.5). These data further support the notion that central neuroinflammatory processes involving immune recognition play a role in peripheral nerve injury.
Of particular clinical interest, specific alleles for MHC II have been associated with a variety of autoimmune diseases like multiple sclerosis, rheumatoid arthritis, and systemic lupus erythematosus.100 A genetic component of chronic pain has
recently emerged to help explain why all individuals with a similar injury or disease may not experience chronic pain.101 To further support a genetic predisposition to chronic pain, a statistically significant increase of the MHC II antigen DQ1 was observed in patients with Complex Regional Pain Syndrome I as compared to control frequencies.102 Similarly, we have put forth a provocative theory that specific poly- morphisms in membrane glycoproteins such as MHC II may render an individual more susceptible to persistent pain after a root or peripheral nerve injury.103 6.4.3 CELLULAR ADHESION MOLECULES
In addition to the expression of MHC II on glial cells, upregulation of cellular adhesion molecules (CAM) has been observed in ischemia and multiple sclerosis.104–106 We have preliminary data demonstrating increased intracellular adhesion molecule (ICAM) and platelet endothelial cellular adhesion molecule (PECAM) expression in the spinal cord following peripheral nerve injury (Fig. 6.6). CAMs are pivotal FIGURE 6.5 MHC II expression in L5 spinal cord on day 7 post L5 spinal nerve transection.
MHC II expression was observed in the ipsilateral (A) but not contralateral dorsal horn (B).
The slightly ramified appearance of the MHC II immunoreactivity supports a glial source of expression.
FIGURE 6.6 At day 3 post L5 spinal nerve transection an upregulation of both PECAM (A) and ICAM (B) immunoreactivity can be observed in the dorsal horn of the L5 spinal cord.
Adhesion molecule staining appeared associated more with the vasculature than with glial cells.
in the capture, rolling, adhesion, and migration of leukocytes into an area of infection. CAMs are cell-surface macromolecules that also control cell–cell inter- actions during the development of the nervous system by regulating such processes as neuronal adhesion and migration, neurite outgrowth, synaptogenesis, and intra- cellular signaling.107 The cellular adhesion molecules are classified into four major families: the integrins, the immunoglobulin superfamily, cadherins, and selectins.
ICAM and PECAM belong to the immunoglobulin superfamily.108 TNF has been found to regulate ICAM in an autocrine manner when the CNS is exposed to an immunologic challenge.109 TNF and IL-1β have been found to induce ICAM-1 expression on astrocytes.55 In addition, ICAM expression is an important facilitator for astrocytes to function as antigen-presenting cells in intracerebral immune responses.99 This means that proinflammatory cytokine expression is imperative for conversion of latent glial cells into reactive/activated immunocompetent cells.