Cellular and Molecular Mechanisms of Glial Scarring and Progressive Cavitation:In VivoandIn VitroAnalysis of Inflammation-Induced Secondary Injury after CNS Trauma

Abstract

Post-traumatic cystic cavitation, in which the size and severity of a CNS injury progress from a small area of direct trauma to a greatly enlarged secondary injury surrounded by glial scar tissue, is a poorly understood complication of damage to the brain and spinal cord. Using minimally invasive techniques to avoid primary physical injury, this study demonstrates in vivo that inflammatory processes alone initiate a cascade of secondary tissue damage, progressive cavitation, and glial scarring in the CNS. An in vitro model allowed us to test the hypothesis that specific molecules that stimulate macrophage inflammatory activation are an important step in initiating secondary neuropathology. Time-lapse video analyses of inflammationinduced cavitation in our in vitro model revealed that this process occurs primarily via a previously undescribed cellular mechanism involving dramatic astrocyte morphological changes and rapid migration. The physical process of cavitation leads to astrocyte abandonment of neuronal processes, neurite stretching, and secondary injury. The macrophage mannose receptor and the complement receptor type 3 ␤2-integrin are implicated in the cascade that induces cavity and scar formation. We also demonstrate that anti-inflammatory agents modulating transcription via the nuclear hormone receptor peroxisome proliferator-activated receptor-␥ may be therapeutic in preventing progressive cavitation by limiting inflammation and subsequent secondary damage after CNS injury. Injury to the adult mammalian C NS leads to a complex series of cellular and molecular events, as cells respond to trauma and attempt to repair damaged regions of the brain or spinal cord (for review, see Fitch and Silver, 1999a). Unlike the successf ul healing responses in the peripheral nervous system, adult C NS injury leads to permanent disability, because most severed axons fail to regenerate (Ramon y C ajal, 1928;Guth, 1975;Reier et al., 1983). A phenomenon that adds to the complexity of regenerative failure is the process of progressive cavitation in which, after days to weeks, a C NS injury can expand in size leading to a scarencapsulated cavity many times the size of the initial wound (Balentine, 1978). Although various hypotheses suggest that this secondary process of cavitation is related to ischemia (Balentine, 1978), hemorrhage (Ducker et al., 1971;Wallace et al., 1987), lysozyme activity (Kao et al., 1977), pulsatile hydrodynamics (Williams et al., 1981), or macrophage infiltration and inflammation (Blight, 1991a(Blight, , 1994Szczepanik et al., 1996;Fitch and Silver, 1997a;Z hang et al., 1997), the underlying causes of progressive axon damage and the cellular mechanisms that lead to cyst formation are poorly understood. Insights into this process will provide direction for therapeutic intervention designed to minimize secondary damage and lead to enhanced function after a debilitating injury.In this study we have used both in vivo and in vitro models to test our hypothesis that post-traumatic inflammation can lead to ...

CNS injury, glial scars, and inflammation: Inhibitory extracellular matrices and regeneration failure

“…Some have suggested that degenerating axons and their dying terminals may be a trigger for such extracellular matrix production by astrocytes (see Massey, et al, in this issue). Evidence also suggests that the inflammatory infiltration and activation associated with CNS injury is associated with production of inhibitory molecules, as CSPGs are associated with breakdown of the blood brain barrier and infiltrating macrophages present within a lesion site (Fitch, et al, 1999;Fitch and Silver, 1997a), a phenomenon that is also seen with activation of oligodendrocyte precursor cells after injury (Rhodes, et al, 2006). Pro-inflammatory molecules that activate microglial cells and macrophages can trigger astrocyte gliosis and upregulation of proteoglycans (Fitch, et al, 1999), upregulation of NG2 proteoglycan by oligodendrocyte precursor cells (Rhodes, et al, 2006), and cytokines associated with inflammation can also influence astrocyte extracellular matrix production (DiProspero et al, 1997).…”

“…Microglial cells from the CNS and activated macrophages from the periphery both respond to trauma in the brain and spinal cord, and this inflammatory response may contribute to secondary tissue damage after the primary insult (Blight, 1994;Fitch, et al, 1999). This cascade of secondary damage, progressive cavitation, and glial scarring was demonstrated in an in vivo model of inflammation using microinjection of zymosan, a non-toxic, specific phagocytic activator of the macrophage mannose receptor and the beta-glucan site of the CR3 integrin receptor (Fitch, et al, 1999). The resulting inflammation leads to significant axotomy and increases in astrocyte cavity size, while control injections of latex microspheres do not induce this damaging inflammatory cascade.…”

“…Astrocytes are important for neurotransmitter regulation (Schousboe and Westergaarde, 1995), ion homeostasis (Walz, 1989), blood brain barrier maintenance (Wolburg and Risau, 1995), and the production of extracellular matrix molecules destined for the basal lamina and perineuronal net (Ard et al, 1993;Bernstein et al, 1985;Canning et al, 1996;Grierson et al, 1990;Massey et al, 2006;McKeon et al, 1991;SmithThomas et al, 1994;Tom et al, 2004a). After injury, they are also the major cell type responsible for walling off areas of damage to protect the fragile brain tissue from further erosion (Fitch et al, 1999;Myer et al, 2006). The formation of a mechanically obstructive glial scar composed of astrocytes and connective tissue elements was at one time thought to explain the failure of regeneration within the CNS after injury (Windle and Chambers, 1950).…”

Setting the stage for functional repair of spinal cord injuries: a cast of thousands

“…[11][12][13][14] The nontraumatic injection of the micgroglia/macrophage activator zymosan into the brain and spinal cord results in cavitation, demyelination, and axonal injury. 15,16 The molecules released by mature phagocytes, including superoxide, hydrogen peroxide, hypochlorous acid, as well as neurotoxic chemokines and cytokines may be partially responsible for axonal damage and retraction, and neuronal death. 17 The second-generation tetracycline derivative, minocycline, has been shown to be an antiinflammatory agent.…”

Progress in Spinal Cord Research - A refined strategy for the International Spinal Research Trust

“…They further showed in a culture model that progressive cavitation could be prevented by treatment with anti-in¯ammatory agents which inhibited macrophage in¯ammatory gene transcription. 44 Further evidence for the detrimental role of neutrophils and macrophages is that reactive oxygen species are produced during the process of phagocytosis by these cells. 45 Thus, neutrophil in¯ux in a SCI has been associated with oxidative damage of surrounding healthy tissue.…”