Neuron Review

Reactive Gliosis and the Multicellular Response to CNS Damage and Disease

Joshua E. Burda1 and Michael V. Sofroniew1,* 1Department of Neurobiology and Brain Research Institute, University of California Los Angeles, Los Angeles, CA 90095-1763, USA *Correspondence: [email protected] http://dx.doi.org/10.1016/j.neuron.2013.12.034

The CNS is prone to heterogeneous insults of diverse etiologies that elicit multifaceted responses. Acute and focal injuries trigger wound repair with tissue replacement. Diffuse and chronic diseases provoke gradually escalating tissue changes. The responses to CNS insults involve complex interactions among cells of numerous lineages and functions, including CNS intrinsic neural cells, CNS intrinsic nonneural cells, and CNS extrinsic cells that enter from the circulation. The contributions of diverse nonneuronal cell types to outcome after acute injury, or to the progression of chronic disease, are of increasing interest as the push toward understanding and ameliorating CNS afflictions accelerates. In some cases, considerable information is available, in others, comparatively little, as examined and reviewed here.

Introduction enter from the circulation. The biology of cell types that partici- A major goal of contemporary neuroscience is to understand and pate in CNS responses to injury and disease models has gener- ameliorate a wide range of CNS disorders. Toward this end, ally been studied in isolation. There is increasing need to study there is increasing interest in cellular and molecular mechanisms interplay of different cells to understand mechanisms. This Re- of CNS responses to damage, disease, and repair. Neurons are view examines and reviews the multiple cell types involved in, the principal cells executing neural functions and have long and contributing to, different types of CNS insults. In some dominated investigations into mechanisms underlying CNS dis- cases, extensive information is available, in others, compara- orders. Nevertheless, mounting evidence indicates that treating tively little. all types of CNS disorders will require a deeper understanding of Terminology how multicellular responses to injury and disease are triggered, Various terms used in discussions of CNS injury and disease evolve, resolve (or not), and impact on neuronal function. can be subject to different interpretations. In this Review, we The ability to repair tissue damaged by injury is fundamental to will define and use certain specific terms as follows. ‘‘Reactive vertebrate biology and central to survival. Evolutionary pressure gliosis’’ will refer not only to microglia and astroglia, but also to is likely to have forged certain fundamental cellular and molecu- glial cells that have come to be known as NG2-positive oligoden- lar responses to damage that are common across different tis- drocyte progenitor cells (NG2-OPCs). Glial cells in healthy CNS sues. The wound or injury response in skin has long served as tissue will not be referred to as ‘‘resting’’ or ‘‘quiescent.’’ This a model system for dissecting mechanisms of tissue repair after is an antiquated concept. Glia are highly active in healthy CNS acute focal tissue damage and has provided insight into core and dynamically exert complex functions that play critical roles cellular and molecular interactions (Greaves et al., 2013; Gurtner in normal CNS functions (Barres, 2008; Sofroniew and Vinters, et al., 2008; Singer and Clark, 1999). In addition, organ-specific 2010). For example, astrocytes exhibit physiological activation features exist. Organ-intrinsic cells that are specialized in inflam- in the form of transient, ligand-evoked elevations in intracellular 2+ matory regulation and tissue repair are emerging as critical calcium ([Ca ]i) that represent a type of astrocyte excitability, elements in organ-specific responses to insults. Organ-specific which is under intense investigation as a potential means of features apply particularly in the CNS, where glial cells, which mediating dynamic astrocyte functions, including interactions maintain the cytoarchitecture and homeostatic regulation with synapses and regulation of blood flow (Attwell et al., 2010; without which neurons could not function normally in healthy tis- Halassa and Haydon, 2010; Tong et al., 2013; Verkhratsky sue, are also principal responders to CNS insults. Changes in et al., 1998). Microglia perform essential roles in synapse devel- glial cell function during responses to insults have the potential opment and turnover (Stephan et al., 2012; Stevens et al., 2007). to impact markedly on neuronal interactions and CNS functions. The term ‘‘activated’’ is often used in a binary all-or-none fashion CNS insults are caused by diverse etiologies that can elicit a to define glial cells that have responded to insults. We feel that wide range of responses. For example, acute and focal injuries use of the term in this manner is inaccurate in two ways. First, trigger wound repair with tissue replacement, whereas diffuse it does not recognize that glia are continually being ‘‘activated’’ and chronic diseases can trigger gradually escalating tissue in physiological contexts. Second, as discussed throughout changes. Analysis of similarities and differences in such re- this Review, glial cell responses to CNS insults are not binary sponses can provide valuable insights. Cellular responses to and are highly diverse and specifically regulated. To differentiate CNS insults involve complex interactions among cells of physiological activation of glial cells in healthy contexts from re- numerous lineages and functions, including CNS intrinsic neural sponses associated with injury or disease, we will use the term cells, CNS intrinsic nonneural cells, and CNS extrinsic cells that ‘‘reactive,’’ which is also meant denote a broad spectrum of

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 229 Neuron Review

Table 1. Diverse Cell Types in CNS Responses to Damage and progenitors and cells of different types that migrate to sites of Disease injury (Benner et al., 2013; Lagace, 2012; Meletis et al., 2008; CNS Intrinsic Cells Blood-Borne Nonneural Cells Ohab and Carmichael, 2008). CNS Intrinsic Nonneural Cells CNS Intrinsic Neural Cells Leukocytes Various nonneural-lineage cells intrinsic to CNS play critical roles Neurons Monocyte/macrophage in CNS damage and disease (Table 1). Microglia are well docu- Oligodendrocytes Neutrophils mented as highly sensitive early responders that stimulate and Astrocytes Eosinophils recruit other cells, as well phagocytose debris (Hanisch and NG2-OPC NK cells Kettenmann, 2007; Kreutzberg, 1996; Nimmerjahn et al., 2005; Neural stem/progenitor cells T cells Ransohoff and Perry, 2009). Fibroblast-related cells, including Ependyma B cells perivascular fibroblasts, meningeal fibroblasts, and pericytes, CNS Intrinsic Nonneural Cells Other Bone Marrow-Derived Cells contribute to tissue replacement by forming fibrotic scar tissue after severe damage (Go¨ ritz et al., 2011; Klapka and Mu¨ ller, Microglia Platelets (thrombocytes) 2006; Logan and Berry, 2002; Soderblom et al., 2013; Winkler Perivascular fibroblasts Fibrocytes et al., 2011). Endothelia and endothelial progenitors are promi- Pericytes Mesenchymal (bone marrow nent during tissue replacement after CNS injury (Loy et al., stromal) stem cells 2002; Oudega, 2012) and are of increasing interest in CNS repair Endothelia and progenitors in light of their emerging roles as trophic support cells in CNS development (Dugas et al., 2008) and their ability to produce and present laminin (Davis and Senger, 2005), a preferred growth potential responses of glial cells to CNS insults. Lastly, we will substrate for many migrating cells and axons. avoid use of the term ‘‘scar’’ on its own and will instead distin- Blood-Borne Nonneural Cells guish between ‘‘astrocyte scars’’ that form compact borders Blood-borne immune and inflammatory cells of different kinds around tissue lesions and ‘‘fibrotic scars’’ that are formed by (Table 1) play prominent roles in CNS responses to damage multiple nonneural cell types and extracellular matrix within and disease and have been studied and reviewed (Perry, 2010; lesion cores (as discussed in detail below). We will equate ‘‘glial Popovich and Longbrake, 2008). In addition to well-known roles scar’’ with ‘‘astrocyte scar.’’ in phagocytosis and removal of debris, there is also now increasing evidence that subtypes of leukocytes play active roles Multicellular Response to CNS Insults in tissue repair (Derecki et al., 2012; London et al., 2013; Popo- Before discussing the different types of responses to CNS dam- vich and Longbrake, 2008). Platelets aggregate rapidly after age and disease, it is useful briefly to introduce different cell damage for clot formation and haemostasis. Other blood-borne, types involved in these responses. For ease of consideration, bone marrow-derived cell types that home in to tissue injury, we have grouped cells according to lineages as (1) neural and including in CNS, include fibrocytes (Aldrich and Kielian, 2011; nonneural cells intrinsic to CNS and (2) blood-borne nonneural Reilkoff et al., 2011) and bone marrow-derived mesenchymal cells that derive primarily from bone marrow (Table 1). Some of stem cells (Askari et al., 2003; Bianco et al., 2001; Jaerve et al., these cell types have been studied extensively in CNS disorders, 2012), but their functions are not well understood. others comparatively little. Most often they have been studied in Extracellular Matrix isolation. In subsequent sections, we will endeavor to examine Extracellular matrix (ECM) generated by different cells plays crit- their interactions. ical roles in tissue repair and replacement after acute insults, as CNS Intrinsic Neural Cells well as in chronic tissue remodeling during chronic disease (Mid- The principal neural-lineage cells in the CNS, neurons, oligoden- wood et al., 2004). Molecules that modify ECM matrix, such as drocytes and astrocytes, have been studied and reviewed exten- metaloproteases (MMPs), are also important in this regard (Mid- sively as regards their individual responses to CNS injury and wood et al., 2004). In the CNS, ECM generated by different cell disease (Barres, 2008; Franklin and Ffrench-Constant, 2008; types responding to damage or disease can include a wide vari- Mattson, 2000; Sofroniew and Vinters, 2010). Less well studied ety of molecules such as laminin, collagens, and glycoproteins are glial cells that have come to be known as NG2-positive oligo- such as chondroitin or heparan sulfate proteoglycans (CSPGs dendrocyte progenitor cells (NG2-OPCs). Like other glia, NG2- or HSPGs) that are implicated both in supporting tissue repair OPC tile the CNS and respond to CNS insults (Nishiyama as well as in contributing to the failure of axonal regeneration et al., 2009). Features of the NG2-OPC response include migra- (Davis and Senger, 2005; Klapka and Mu¨ ller, 2006; Logan and tion toward injury and cell proliferation (Franklin and Ffrench- Berry, 2002; Silver and Miller, 2004). Constant, 2008; Hughes et al., 2013). Besides replacing lost oligodendrocytes (Sun et al., 2010), other roles of NG2-OPC in Response to Focal Insults CNS injury and disease await future study and elucidation. In Two principal types of acute focal insults in the CNS are focal addition, the adult CNS harbors neural stem cells (NSCs) of traumatic injury and ischemic stroke. In addition to being major different potencies that reside in the periependymal regions clinical problems, they have for decades served as prototypical along the ventricles and central canal in adult brain and spinal experimental models with which to study CNS mechanisms of cord (Garcia et al., 2004; Kriegstein and Alvarez-Buylla, 2009). response to damage and repair. In this section, we will examine These adult NSCs can also respond to CNS injury and generate basic features of CNS multicellular responses to acute focal

230 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

Figure 1. Phases and Time Course of Multicellular Responses to Acute Focal CNS Damage During periods of cell death (A), cell replacement (B), and tissue remodeling (C), numerous overlapping events occur (D) that involve interactions among CNS intrinsic neural cells, CNS intrinsic nonneural cells, and cells infiltrating from the circulation. injuries and how these change over time, using information from We will first examine CNS responses in a context in which the response to trauma or stroke as models. In this regard, it is note- acute insults remain uninfected and resolve. Below, we will worthy that acute focal trauma caused by contusion or crush re- also briefly consider more chronic focal CNS insults, including sults in severe vascular damage and therefore exhibits many focal infections with abscess formation, primary and secondary sequelae similar to stroke. We will make comparisons with the tumors, and chronic focal autoimmune lesions, such as multiple acute wound response in skin as a model system for cellular sclerosis plaques, which exhibit various features and cellular re- and molecular mechanisms of wound repair (Greaves et al., sponses similar to traumatic and ischemic injury. In order to pre- 2013; Gurtner et al., 2008; Shechter and Schwartz, 2013; Singer sent succinct summaries of cellular responses and interactions, and Clark, 1999). As in skin repair, CNS responses to acute focal discussion of information about underling molecular signals is damage can also be divided broadly into three overlapping but deferred to a later section below. It deserves mention that the distinct phases: (1) cell death and inflammation, (2) cell prolifer- time courses given for response phases and events are general- ation for tissue replacement, and (3) tissue remodeling (Figure 1). ized approximations (Figure 1), and there is much potential for

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 231 Neuron Review

Figure 2. Lesions Exhibit Tissue Compartments and Signaling Gradients (A and B) Focal damage gives rise to lesions with distinct tissue compartments in the form of nonneural lesion cores (LC), newly proliferated compact astrocyte scars (AS), and perilesion pe- rimeters (PLP) that exhibit diminishing gradients of reactive gliosis that transition gradually to healthy tissue (HT). AS abruptly demarcate the transition between nonneural LC and PLP with viable neural cells. Size of the lesion core and the location of AS, and this transition, are regulated by opposing molecular signals. Gradients of molecular signals emanating from LC impact on cells in PLP.

et al., 2005). Astrocytes, in contrast, remain in situ and do not migrate either to or away from injury sites but can swell osmotically and, depending on the severity of injury or ischemia, can die in the center of severe lesions or can overlap of events as well as variations in duration or timing in become reactive and hypertrophy and in some cases proliferate specific situations. (Bardehle et al., 2013; Zheng et al., 2010). Different aspects of Phase I: Cell Death and Inflammation this first phase of response occur in overlapping sequences dur- After focal CNS tissue damage from insults such as trauma or ing the first few days and then begin gradually diminishing ischemia that cause acute local cell death, the first phase of (Figure 1D), provided that the insult is a single acute event not response in the injury center or core includes both very rapid complicated by bony compression, infection, or other exacerba- events that occur over timescales of seconds to hours and tions that might prolong the damaging insult. As acute responses more gradually progressing events that develop over days (Fig- diminish, there is overlap with the onset of different types of cell ures 1A and 1D). Very rapid events include haemostasis with proliferation associated with the next phase of responses (Fig- coagulation cascade, platelet aggregation, and clot formation. ures 1B and 1D). These rapid events are followed by overlapping sequences of re- Phase II: Cell Proliferation and Tissue Replacement sponses of tissue intrinsic cells, recruitment of inflammatory and The second phase of response to acute CNS tissue damage oc- immune cells, subacute death of parenchymal cells, and initiation curs from about 2 to 10 days after the insult and is characterized of debris removal (Figures 1A and 1D). In healthy CNS tissue, large by the proliferation and local migration of cells that implement tis- and polar molecules in the circulation are excluded from diffusion sue repair and replacement. These cells include endothelial pro- into CNS parenchyma by the blood-brain barrier (BBB) (Abbott genitors, fibroblast-lineage cells, different types of inflammatory et al., 2006; Zlokovic, 2008). After focal insults with BBB damage, cells, and various types of glia and neural-lineage progenitor cells, blood-borne molecules normally excluded from the CNS influx including scar-forming astrocytes and their progenitors (Figures and signal to local cells, as discussed in more detail below in 1B and 1D). Some of these cellular proliferative phenomena, the section on molecular signaling. Platelets influx and rapidly such as proliferation of endothelia for neovascularization (Casella form aggregates for haemostasis and also signal to local cells. et al., 2002) or proliferation of certain fibroblast-lineage cells or in- Fibrin and collagen matrices begin to form over a period from flammatory cells, reflect classic wound responses (Gurtner et al., hours to days, which serve as a scaffold for neutrophil and then 2008; Singer and Clark, 1999), while others, such as proliferation macrophage and other leukocyte infiltration. Leukocytes then of scar-forming astrocytes, are specific and unique to CNS. This infiltrate heavily to monitor for pathogens, remove debris, and phase is also characterized throughout its duration by the provide molecular signals involved in wound repair over a variable absence of a BBB in the lesion core during the period in which number of days or longer depending on severity of tissue damage damaged blood vessels are replaced by new ones (Figure 1D). or presence of exacerbating factors (Figures 1A and 1D) (Perry, As a consequence, endogenous (as well as exogenous) proteins 2010; Popovich and Longbrake, 2008). Endogenous mesen- and other charged molecules can freely diffuse into the surround- chymal stem cells may also enter the lesion core, but their roles ing neural parenchyma (Bush et al., 1999). These molecules can are not well understood (Askari et al., 2003; Bianco et al., 2001; include serum proteins (e.g., thrombin, albumin) that signal to Jaerve et al., 2012). These basic cellular events reflect classic local cells, immunoglobulins, or pathogen-associated molecules, wound responses as found in other tissues (Greaves et al., as discussed below. This leaky BBB creates the potential for the 2013; Gurtner et al., 2008; Singer and Clark, 1999). lesion core to serve as source of circulating molecules that Certain CNS intrinsic cells also respond rapidly to CNS tissue generate gradients of molecular signaling that can extend for damage or BBB leak. Live imaging studies in CNS in vivo show considerable distances into neural tissue around the lesion until that microglia and NG2-OPC immediately migrate to sites of tis- the BBB is repaired (Figures 1B, 1D, 2A, and 2B). This signaling sue damage and BBB leak (Hughes et al., 2013; Nimmerjahn may contribute to the perimeter of tapering gradient of reactive

232 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

gliosis and other changes observed in tissue around mature Phase III: Tissue Remodeling lesions as discussed below. The third phase of response to acute tissue damage generally During the proliferative phase, cellular elements are generated begins toward the end of the first week after the insult and is that will in mature lesions form two of the three major tissue com- distinguished by tissue remodeling that includes events that partments of the mature lesion, (1) the central lesion core of non- are completed within weeks, such as BBB repair and scar orga- neural tissue and (2) the compact astrocyte scar that surrounds nization, as well as chronic events that can continue over many the lesion core (Figures 1B and 1D). Major cellular components months (Figures 1C and 1D). BBB repair or restoration along of the nonneural lesion core derive from expansion of CNS newly formed blood vessels in the lesion vicinity is generally intrinsic perivascular fibroblasts and pericytes (Go¨ ritz et al., completed within several weeks after acute uncomplicated in- 2011; Soderblom et al., 2013) and from proliferating endothelial sults and requires the presence of functional astrocytes (Bush progenitors. The compact astrocyte scar is formed primarily et al., 1999) and pericytes (Daneman et al., 2010; Winkler et al., from newly proliferated elongated astrocytes generated by local 2011). The lesion core of nonneural tissue becomes surrounded astroglial progenitors that gather around the edges of severely by a well-organized, interdigitating compact astrocyte scar by 2– damaged tissue containing inflammatory and fibroblast-lineage 3 weeks after acute insults not exacerbated by ongoing tissue cells (Figures 1B and 1D) (Faulkner et al., 2004; Sofroniew and damage (Figures 1C and 1D) (Wanner et al., 2013). These scar- Vinters, 2010; Wanner et al., 2013). It is noteworthy that during forming astrocytes appear actively to corral and surround inflam- this proliferative period, the location of compact astrocyte scar matory and fibroblast-lineage cells during tissue remodeling and borders is being determined. In mature lesions, these astrocyte scar formation (Wanner et al., 2013). This compact astrocyte scar borders will precisely demarcate and separate persisting scar forms a structural and functional border between nonneural areas of nonfunctional, nonneural lesion core tissue from imme- tissue in the central lesion core and the immediately adjacent diately surrounding and potentially functional neural tissue (Fig- and surrounding neural tissue that contains viable cells of all ures 1B, 2A, and 2B). Cellular and molecular mechanisms that three neural-lineage cell types, astrocytes, oligodendrocytes, underlie the determination of precisely where such astrocyte and neurons (Figures 1C and 2)(Wanner et al., 2013). This astro- scar borders are formed are incompletely understood but are cyte scar border serves as a protective barrier that restricts the likely to involve a complex interplay and balance of molecular migration of inflammatory cells from the nonneural lesion core signals that on the one hand foster phagocytosis and debris into surrounding viable neural tissue. Disruption of astrocyte clearance and signals that on the other hand foster protection scar formation in different kinds of transgenic loss-of-function and preservation of healthy self (Figure 2B), as discussed in models leads to increased lesion size, increased death of local more detail below in the section on molecular signals of cell neurons, increased demyelination, and decreased recovery of damage and death. Several decades of experimental studies function after traumatic or ischemic focal insults (Bush et al., on ischemic infarcts show that final lesion size can be influenced 1999; Faulkner et al., 2004; Herrmann et al., 2008; Li et al., by subacute metabolic events that continue for hours to days 2008; Wanner et al., 2013). After compact astrocyte scar forma- within penumbral zones around the lesions (Astrup et al., 1981; tion, the second period of tissue remodeling is long lasting, with Dirnagl et al., 1999; Lo, 2008). In addition, astrocyte intrinsic gradual remodeling of lesion core, astrocyte scar border, and signaling cascades have been identified that play critical roles surrounding perimeter regions (Figures 1D and 2). For example, (Herrmann et al., 2008; Okada et al., 2006; Wanner et al., with time there is gradual contraction of lesion cores and astro- 2013). A better understanding of the multicellular and molecular cyte scars. As transected axons continue to die back away from interactions that determine the location of astrocyte scar borders lesions, oligodendrocyte death can continue, leading to local tis- around areas of compromised tissue may lead to novel thera- sue responses and remodeling. There is long-term remodeling of peutic strategies for reducing lesion size (Figure 2B). These ECM in the lesion core, with modulation of collagen and glyco- events occur over a time frame of days after the insult protein components (Klapka and Mu¨ ller, 2006; Logan and Berry, (Figure 1D), during which intervention is a realistic possibility. 2002; Silver and Miller, 2004). In addition, there is gradual and Recent findings also indicate that during the proliferative chronically ongoing tissue remodeling in reactive perimeter phase after acute CNS damage in the forebrain, neural stem cells regions that surround mature lesions, as discussed next. in periventricular germinal zones give rise to considerable Mature Lesions Exhibit Three Tissue Compartments numbers of neural progenitors that migrate to the injury sites Astrocyte scar formation around central lesion cores is essen- in cortex or striatum (Carmichael, 2006; Kokaia et al., 2012; tially complete by 2–4 weeks after acute insults that are not Lagace, 2012; Lindvall and Kokaia, 2006). These cells form complicated by prolonged tissue damage, and the lesion can part of the mix of cells in perilesion perimeters (Figures 1B and at this time be considered mature and entering its chronic stage 1C). There are various lines of evidence suggesting that these (Figures 1 and 2). In mature lesions, it is useful to recognize three cells may contribute beneficially to tissue remodeling in perile- very distinct lesion compartments: (1) the central lesion core of sion tissue, but the nature of the contributions made by these nonneural tissue comprised of locally derived fibroblast-lineage cells remains nebulous. Under normal, unmanipulated circum- cells, blood vessels, infiltrating fibrocytes and inflammatory stances, very few newly generated neurons mature and survive, cells, and extracellular matrix; (2) the compact astrocyte scar and other potential contributions by these intriguing, migrating that immediately surrounds the lesion core and consists of progenitor cells, such as contributions to immune regulation, densely packed astrocytes with few if any neurons or oligoden- are still in the process of being defined (Kokaia et al., 2012; drocytes; and (3) the perilesion perimeter of viable neural tissue Lagace, 2012). that is adjacent to the compact astrocyte scar and contains all

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 233 Neuron Review

three types of neural-lineage cells (neurons, oligodendrocytes, prevented or attenuated in vivo, inflammation spreads, lesion and astrocytes) and exhibits a tapering reactive gliosis that grad- size increases, neuronal loss and demyelination are exacer- ually transitions to healthy tissue (Figure 2). All of these compart- bated, and functional recovery is diminished (Bush et al., 1999; ments will exhibit further tissue remodeling for weeks to months. Dro¨ gemu¨ ller et al., 2008; Faulkner et al., 2004; Wanner et al., Some of this remodeling has the potential to impact on functional 2013). Scar-forming astrocytes are also widely regarded as a, outcome and may provide targets for therapeutic interventions, if not the, major impediment to axon regeneration after CNS particularly in the perimeter of neural tissue that exhibits diffuse injury, based in part on in vitro studies showing astrocyte pro- reactive gliosis (Figures 1D and 2). It is useful to consider duction of certain proteoglycans that can inhibit elongation separately these compartments and their impact on functional (Silver et al., 1993). Nevertheless, other studies show that axon outcome. regeneration in vivo occurs along astrocyte bridges and that, in Lesion Core. Tissue in the mature lesion core contains few the absence of such bridges, axon regeneration does not occur or no neural-lineage cells and is nonfunctional in neurological and seems more to be impeded by confrontation with nonneural terms. Initially, the main cell types in mature lesion cores include lesion core tissue (Kawaja and Gage, 1991; Williams et al., 2013; fibroblast-lineage cells, endothelia, fibrocytes, and inflammatory Zukor et al., 2013). Transgenic loss-of-function studies conduct- cells (Figure 2A). With time and remodeling, and in the absence ed in vivo may help to clarify the precise nature and scale of of exacerbating damage or infection, inflammatory elements effects exerted by scar-forming astrocytes on axon regrowth will gradually withdraw, although some may persist for long after injury. times. Elements of lesion core will persist permanently as fibrotic Perilesion Perimeters. All mature lesion cores and astrocyte scar containing various nonneural cells and ECM. In some areas scars are surrounded by perimeters of viable neural tissue that of lesion core, cells and ECM will recede to leave fluid-filled cysts exhibits a gradient of tapering reactive gliosis that gradually tran- of quite variable size (Figure 2A) (Tuszynski and Steward, 2012). sitions to healthy tissue (Figure 2A) (Wanner et al., 2013). The Functionally, cells and ECM of lesion core serve as substrates tissue in these perimeter zones contains all neural-lineage cell for rapid wound repair and tissue replacement. The rapidity of types including neurons and oligodendrocytes, and these cells wound closure allowed by fibrotic replacement as compared are intermingled with reactive gliosis that can extend for consid- with parenchymal renewal has been suggested to afford advan- erable distances (Figure 2A). Reactive perimeter tissue begins tages in other tissues (Greaves et al., 2013; Gurtner et al., 2008; immediately adjacent to compact astrocyte scars, where the Singer and Clark, 1999). Nevertheless, this mechanism can elongated cell processes of scar-forming astrocytes overlap come with the cost of lost tissue functions. In CNS, lesion core with hypertrophic reactive astrocytes and reactive microglia tissue has since the 19th century been thought to have a negative that are intermingled with surviving viable neurons and other long-term functional impact as an impediment to axon regener- elements of neural tissue (Figure 2A). It is somewhat remarkable ation, confirmed by recent studies as well (Zukor et al., 2013). that viable neurons can be present within a few hundred micro- Molecular evaluations show that lesion core ECM contains colla- meters of compact astrocyte scars and the lesion core tissue gens and proteoglycans that poorly support or overtly inhibit filled with potentially cytotoxic inflammatory elements just regrowth of damaged axons (Hermanns et al., 2006; Kimura-Kur- beyond (Bush et al., 1999; Wanner et al., 2013). The reactive glio- oda et al., 2010; Klapka and Mu¨ ller, 2006; Yoshioka et al., 2010). sis in perimeters includes hypertrophic reactive astrocytes and Compact Astrocyte Scars (Glial Scars). Mature astrocyte scars microglia (Figure 2A). Both the cause and functions of perimeters consist of relatively narrow zones of newly generated astrocytes of tapering reactive gliosis are not clear. Regarding cause, it with elongated processes that intermingle and intertwine exten- is possible that gradients of signaling molecules diffusing out sively and that immediately abut and surround on all sides re- from the lesion core, either entering during the period of BBB gions of lesion core tissue (Figure 2A) (Sofroniew and Vinters, leak or produced there by infiltrating and proliferating cells, 2010; Wanner et al., 2013). These narrow zones of densely inter- may trigger such a gradient of gradually tapering responses twined elongated scar-forming astrocytes are generally not more among local glia (Figures 2A and 2B). From a functional perspec- than a millimeter across and are generally devoid of other neural- tive, the degree to which specific aspects of this reactive gliosis lineage cells (neurons or oligodendrocytes) (Wanner et al., are beneficial or harmful (or some mixture of both) remains to be 2013). These scar-forming astrocytes are newly proliferated in determined. response to the insult and the packing density of astrocyte cell Reactive perimeter areas around lesion cores in brain and spi- bodies in astrocyte scars is up to double that in healthy tissue nal cord are likely to exhibit long-term, and potentially intense, (Wanner et al., 2013). The densely packed elongated scar-form- tissue remodeling, with new synapse formation and the potential ing astrocytes transition seamlessly and rapidly to less densely for formation of new relay circuits (Bareyre et al., 2004; Carmi- packed and less densely intertwined hypertrophic reactive as- chael, 2006; Clarkson et al., 2010; Courtine et al., 2008; Li trocytes that are intermingled with viable neurons and other et al., 2010; Overman et al., 2012). Numerous factors have the elements of neural tissue in the surrounding perimeter of reactive potential to impact on this synapse and circuit remodeling, neural tissue discussed below (Figure 2A) (Wanner et al., 2013). including extracellular matrix molecules such as proteoglycans Transgenic loss-of-function studies show that newly proliferated in the perineuronal (Garcı´a-Alı´as et al., 2009). Reactive glia may compact astrocyte scars exert essential neuroprotective func- play important roles in this remodeling, but this topic is under- tions by restricting the spread of inflammatory cells away from studied. In the healthy CNS, astrocytes interact with neurons tissue lesions caused by trauma, stroke, and autoimmune on multiple spatial and temporal scales that can influence or inflammation. When astrocyte scar formation is experimentally modulate neural function in various ways (Halassa and Haydon,

234 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

2010). Both astrocytes and microglia have the potential to play separate nonneural tissue from perimeters of reactive gliosis in fundamental roles in the synapse remodeling in the perilesion tissue with all three neural cell types (Frohman et al., 2006; Luc- perimeter (Stephan et al., 2012; Stevens et al., 2007) and the chinetti et al., 1996) in manners similar to tissue damage caused impact of reactive gliosis on such remodeling is not yet under- by trauma or ischemia (Figures 1 and 2). Neuromyelitis optic stood. Such perilesion perimeter areas are likely to provide fertile (NMO) is an autoimmune inflammatory disease in which causal ground for investing new strategies for interventions that may autoantibodies are directed against aquaporin-4 on the astro- influence neural plasticity and relay circuit formation, which in cyte cell membrane, and NMO lesions are associated with com- combination with appropriate rehabilitative training (van den plement-mediated, lytic destruction of astrocytes (Lennon et al., Brand et al., 2012) have the potential to beneficially influence 2005; Popescu and Lucchinetti, 2012). There is remarkable functional outcome. congruence between clinical evidence from NMO and experi- Chronic Focal Insults mental loss-of-function studies demonstrating that transgeni- In addition to acute focal trauma and stroke, which often serve as cally mediated ablation or attenuation of scar-forming astrocytes models to study mechanisms of CNS damage as just discussed, results in severe exacerbation of autoimmune CNS inflammation other more chronic forms of damage warrant consideration. (Haroon et al., 2011; Voskuhl et al., 2009). Thus, both clinical and Important chronic insults include focal infections with abscess experimental evidence implicate critical roles for scar-forming formation, tumors, and autoimmune lesions. All of these condi- astrocytes in limiting the spread of autoimmune CNS inflamma- tions invoke reactive gliosis and multicellular responses that tion. These observations strongly suggest that dysfunction of have strong similarities with those that occur after acute focal CNS intrinsic cells such as astrocytes may contribute causally damage. This multicellular response plays critical roles in pro- to the onset or progression of CNS autoimmune conditions in gression, severity, and resolution (or not) of the insults. ways not generally considered by research focused only on Infection. Focal infections or abscesses can form when trying to identify causal mechanisms of the peripheral immune traumatic wounds become infected or can be seeded spontane- system. This notion may be further supported by suggestions ously, particularly in immune-compromised individuals. Such that a subgroup of patients with multiple sclerosis have dis- infections elicit reactive gliosis with astrocyte scar formation ease-related autoantibodies against the potassium channel similar to that after acute focal trauma (Sofroniew and Vinters, Kir4.1, which in the CNS is located on astrocyte membranes (Sri- 2010). The time course of lesion persistence and tissue remodel- vastava et al., 2012). It will be interesting to determine the extent ing is protracted and dependent on resolution of the infection. to which gain or loss of astrocyte functions contributes to the Transgenic loss-of-function studies show that astrocyte scar pathophysiology of CNS autoimmune conditions. formation is critical to focal containment of the infection. When astrocyte scars are disrupted, infection and inflammation spread Response to Diffuse Insults rapidly through adjacent neural tissue with devastating effects Diffuse insults to CNS tissue can be associated with neurode- (Dro¨ gemu¨ ller et al., 2008). generative diseases, certain seizure disorders, or mild traumatic Cancer. Both primary and metastatic tumors elicit reactive brain injury. Diffuse insults tend initially to be less intense than gliosis and multicellular responses that have similarities to other acute focal damage caused by trauma or stroke and may not forms of focal tissue damage. Interactions of tumor cells with initially cause overt tissue damage but instead accumulate grad- CNS glia and infiltrating inflammatory cells are complex and ually over chronic periods of time. As the tissue damage be- heavily dependent on tumor cell type. Noninvasive tumors are comes more severe, small individual lesions form, each of which surrounded by reactive gliosis and encapsulating scar similar invokes reactive gliosis and multicellular responses similar to to that seen around traumatic tissue damage. Aggressively inva- those caused by acute tissue damage with breakdown of the sive tumors are not encapsulated or surrounded by well-defined BBB, inflammation, and recruitment of leukocytes (Figures 1 astrocyte scars but evoke other forms of reactive gliosis and and 2) but on a smaller scale. Over time, the diffuse damage multicellular responses. Successful infiltration and spread of comes to resemble a diffuse collection of many small focal certain tumor cell types is thought to be associated with their lesions that are intermingled and can be dispersed over large ability to create an environment for growth, vascularization, areas (Figure 3A). Each of these small lesions gives rise to its and spread, which may include successfully neutralizing bar- own perilesion perimeter zones of tapering reactive gliosis, rier-forming functions of local CNS cells and co-opting programs with the consequence that overlapping zones of severe hyper- for creating proliferative niches (Louis, 2006; Silver et al., 2013; trophic reactive gliosis and multicellular responses can be Watkins and Sontheimer, 2012). distributed over large areas of tissue (Figure 3A). It is important Autoimmune Disease. Focal autoimmune lesions are inter- to realize that these many small lesions are interspersed among esting to consider from the perspective of being a response viable neurons and neural circuitry, which become enveloped by to CNS damage. Although CNS autoimmune disease is often these overlapping perilesion perimeters of reactive gliosis (Fig- viewed as diffuse, chronic, and caused primarily by dysfunctions ures 3A and 3B). As discussed above, there is now substantive of the peripheral immune system, individual autoimmune lesions evidence for the participation of astrocytes and microglia in bear many similarities to acute focal traumatic wounds. For normal neural circuit function, but the impact of reactive gliosis example, active multiple sclerosis plaques are filled with inflam- on such functions is not known. In the context of diffuse CNS matory cells and chronic plaques exhibit central lesion cores that insults, large areas of functioning neural tissue may be exposed are devoid of all neural-lineage cell types (no neurons, oligoden- to intense reactive gliosis that is likely to impact on synaptic in- drocytes, or astrocytes) surrounded by astrocyte scars that teractions and neural circuit functions (Figure 3B). A better

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 235 Neuron Review

Figure 3. Diffuse Tissue Damage Leads to Extended Regions of Reactive Gliosis (A) Small chronic insults can lead to diffuse tissue damage with formation of dispersed small lesion cores (LC), astrocyte scars (AS), and perilesion perimeters (PLP) that can overlap and coalesce into regions of contiguous reactive gliosis that can extend over large areas. (B) Reactive astrocytes and reactive microglia in PLP can impact on synapses and neuronal functions in various ways. understanding of such effects may open inroads to new thera- thought to contribute to clearing b-amyloid (Prokop et al., 2013; peutic strategies. Wyss-Coray et al., 2003), and attenuation of reactive astrogliosis Neurodegenerative Disease can increase b-amyloid load in a transgenic mouse model of AD Reactive gliosis and multicellular responses are triggered in (Kraft et al., 2013). In addition, glia may be affected by the dis- different and selective ways in different neurodegenerative dis- ease-causing molecular defects and become dysfunctional, eases. In some cases, the trigger can be accumulation of extra- further complicating efforts to understand the roles of reactive cellular toxins, such as b-amyloid in Alzheimer’s disease (AD) gliosis in the disease process. For example, in ALS, glial cell (Prokop et al., 2013; Zlokovic, 2011), which can accumulate dysfunction is causally implicated in mediating neuronal degen- and cause many small areas of focal tissue damage that trigger eration (Yamanaka et al., 2008), and ongoing reactive gliosis and gliosis with many similarities to other forms of diffuse insults inflammation are also thought to contribute to disease progres- (Figure 3A). In other cases, the trigger can be neuronal or synap- sion (Maragakis and Rothstein, 2006). Distinctions between tic damage or death secondary to neuronal intrinsic changes. disease-induced dysfunctions of glial cells and the reactive re- Some conditions, such as amyotrophic lateral sclerosis (ALS) sponses that are triggered in glial cells by cell and tissue damage or Huntington’s disease, cause intrinsic cellular changes in are sometimes blurred or equated but should not be. Under- both neurons and glia that can perturb cellular functions and in- standing these different phenomena and the ways in which teractions, which can serve as triggers (Boille´ e et al., 2006; Mar- they interact is likely to have important consequences for under- agakis and Rothstein, 2006). Neurodegenerative insults can also standing the mechanisms that underlie cellular pathophysiology lead to disturbance of the neurovascular unit, resulting in BBB and drive disease progression. It is important to remember that leak, which can serve as a further trigger for reactive gliosis although some aspects of glial reactivity may contribute to dis- and also signal to recruit blood-borne inflammatory cells (Zlo- ease progression, others are likely to be protective. Defining kovic, 2008, 2011). In many cases, reactive gliosis and multicel- the molecular basis of such differences may help to identify novel lular responses may not be apparent at onset or in early stages of therapeutic strategies. the neurodegenerative conditions but appear later and may then In the context of neurodegenerative disease, it is also inter- become involved in disease progression. Reactive gliosis trig- esting briefly to consider the potential for non-cell-autonomous gered in various ways, by accumulation of abnormal proteins, neuronal degeneration precipitated by dysfunction of glia or ischemic or traumatic cellular damage, or microbes, can also other nonneural cells. Loss- or gain-of-function genetic muta- lead to BBB leak via specific molecular signaling cascades as tions in microglia (Derecki et al., 2012) or astrocytes (Brenner discussed below, with resultant inflammation that may con- et al., 2001; Rothstein, 2009; Tao et al., 2011) have the potential tribute to disease progression. to cause non-cell-autonomous degeneration of different types of The contributions of glial cells and reactive gliosis to neurode- neurons in different human diseases and animal experimental generative conditions are complex and only beginning to be models. Such observations indicate that glial cell dysfunction understood. Glia not only become reactive in response to degen- may be the primary causal event in certain neurological condi- erative cues as just mentioned, but can also participate in trying tions. Such observations also raise the possibility that dysfunc- to combat them. For example, both microglia and astrocytes are tion of specific aspects of reactive gliosis during responses to

236 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

CNS insults may exacerbate damage or lead to worse tissue Molecular Signaling and Multicellular Interactions repair and worse outcome. The potential for genetic polymor- Molecular signaling among the different cell types that respond phisms to impact on glial cell functions and responses to CNS to CNS insults is complex, combinatorial, and densely inter- injury is an as yet untapped area that may reveal causal factors woven. Different cells have the capacity to produce and respond underlying differences in the responses of different individuals to similar sets of molecules, and there are hints that some mole- to seemingly similar types and severities of CNS insults. cules may coordinate multicellular responses. Many intercellular Epilepsy signaling molecules have been identified, but we are in the early Astrocytes and microglia can be involved in epilepsy in multiple stages of determining how these multiple signals regulate ways. Astrocytes play multiple critical roles in regulating synapse multicellular interactions and the temporal progression from activity and neuronal excitability and astrocytes may play critical one phase to another during responses to specific CNS insults. roles in the generation of seizure activity (Devinsky et al., 2013; Here we provide a general overview of molecular functional Jabs et al., 2008; Wetherington et al., 2008). In addition, reactive classes (Figure 4A) and a few representative examples of the gliosis of both microglia and astrocytes is a prominent feature many specific molecules that regulate or influence CNS cellular of chronically epileptic neural tissue, which can also exhibit responses to insults (Table 2). BBB breakdown and multicellular inflammation (Devinsky Cell Damage and Death et al., 2013; Jabs et al., 2008; Wetherington et al., 2008). With Cells that are dead, dying, or temporarily damaged but recover- increasing loss of neurons, tissue sclerosis sets in, which has able release or express molecules that signal damage. These similarities with severe diffuse astrogliosis due to other causes molecules fall into many categories including neurotransmitters, as discussed above, with many smaller tissue lesions sur- cytokines, chemokines, neuroimmune-regulators (NI-Regs), and rounded by a continuum of perilesion tissue with a mix of surviv- danger-associated molecular patterns (DAMPs) that stimulate ing neurons and increasingly severe hypertrophic gliosis reactive gliosis and other cellular responses including debris (Figure 3A). The impact of reactive gliosis and the multicellular clearance by immune cells (Figures 4B and 4C) (Table 2). Insults response to tissue damage on the surviving neurons in the peril- of different types and severities release different combinations esion perimeter regions is not well understood (Figure 3B) but of these molecules, which in turn trigger different responses. may contribute to reduced seizure thresholds. Selective experi- For example, mild cellular insults can subtly elevate extracellular mental induction of astrocyte reactivity is associated with a concentrations of glutamate or ATP that attract microglial cell reduction of inhibition in local hippocampal neurons (Ortinski processes without initially unleashing full-blown inflammation et al., 2010). More work in this area is warranted. (Figure 4B) (Davalos et al., 2005; Liu et al., 2009). ATP can Diffuse Traumatic Brain Injury also elicit calcium responses in astrocytes and cause con- Large focal lesions caused by severe focal traumatic brain injury nexin-dependent ATP release that increases extracellular ATP (TBI) (contusion or crush) have been discussed above. Neverthe- levels and amplify this ‘‘danger’’ signal (Figure 4B). More severe less, clinically relevant TBI is often diffuse and widely distributed. cell damage or death will release additional DAMPs and The cellular and molecular mechanisms of diffuse TBI that cause alarmins in the form of cytosolic and nuclear contents such as functional disturbances without large focal lesions are incom- K+, heat shock proteins such as ab-crystallin, S100 family cal- pletely understood. Diffuse TBI is generally characterized by cium-binding proteins, DNA-binding high mobility group box 1 small foci of vascular breakdown and tissue damage that can (HMGB1), as well of DNA and RNA. All of these molecules can be diffusely distributed over large areas of CNS (DeKosky serve as ‘‘danger’’ signals to alert innate immune cells to tissue et al., 2013; McIntosh et al., 1989). It is interesting to consider injury and promote clearance of ‘‘altered self’’ cellular debris via that each of these small areas of tissue or cellular damage may activation of pattern recognition receptors (PRRs) on phagocytic represent a small focal injury with compartments equivalent to innate immune cells (Figure 4C) (Chan et al., 2012; Griffiths et al., core, astrocyte scar, and perimeter, which may be spread over 2010; Kono and Rock, 2008). Molecules such as beta-amyloid large areas without an obvious single large focal region (Ab) produced during neurodegenerative disorders can also (Figure 3A). As discussed above, the interspersed viable neural act as alarmins (Figure 4C). A single type of alarmin may tissue in the many overlapping perilesion perimeter regions will bind multiple classes of PPRs. For example, HMBG1 elicits be exposed to intense hypertrophic reactive gliosis and inflam- proinflammatory signaling through activation of multiple TLRs, mation, which can impact on neural functions (Figure 3B). Spe- receptor for advanced glycation end products (RAGE), and cific cellular and molecular mechanisms of reactive gliosis and macrophage antigen complex-1 (MAC1) (Chan et al., 2012; multicellular responses triggered by mild or moderate diffuse Gao et al., 2011), and S100 activates multiple TLRs and RAGE TBI are only beginning to be characterized. Newly proliferated (Chan et al., 2012). reactive astrocytes exert protective functions essential for Healthy neurons and glia near CNS insults express neuroim- neuronal survival (Myer et al., 2006). Microglia can exert a broad mune regulatory molecules (NI-Regs) and self-defense proteins range of effects depending on specific molecular stimuli, which that signal ‘‘healthy self’’ and serve to confine potentially noxious either protect or exacerbate tissue loss (Hanisch and Ketten- proinflammatory signaling and prevent phagocytosis of viable mann, 2007). Understanding and manipulating the reactive glio- cells (Figure 4C) (Table 2)(Griffiths et al., 2010). These ‘‘healthy sis and multicellular sequelae triggered by mild to moderate TBI self’’ molecular signals include both membrane-bound and has enormous potential to identify new therapeutic targets in a soluble NI-Reg molecules. For example, sialic acids (SAs) in field of high incidence and importance. Emerging molecular membranes of healthy cells interact with SA receptors (SARs), mechanisms are discussed in the next section. including Siglecs, on innate immune cells and serve as ‘‘don’t

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 237 Neuron Review

Figure 4. Multimolecular Regulation and Graded Responses to Damage (A) Responses to CNS damage are regulated and influenced by diverse categories of molecular signals derived from many types of CNS intrinsic and extrinsic cells. (B) Low levels of ATP released by cells during mild CNS insults can act through (1) P2X receptors to induce rapid chemotaxis of microglial processes and (2) P2Y receptors on local astrocytes to evoke calcium signaling and connexin-dependent ATP release, which can increase extracellular ATP levels and thereby amplify this ‘‘danger’’ signal. (C) After severe injury, signaling molecules from ‘‘altered self’’ and ‘‘healthy self’’ vie to balance debris clearance and preservation of viable tissue. Severely damaged or dead cells release a number of hydrophobic intracellular molecules (left), or ‘‘alarmins,’’ which express damage-associated molecular patterns (DAMPs). These ‘‘danger’’ signals alert CNS innate immune cells to tissue injury and promote clearance of debris via activation of pattern recognition receptors on phagocytic immune cells (center). Viable CNS intrinsic cells at lesion borders (right) express membrane-bound and soluble neuroimmune regulatory molecules to prevent attack and phagocytosis by immune cells. eat me’’ signals (Figure 4C) (Varki and Gagneux, 2012). CD200 (Griffiths et al., 2010). Complement regulatory proteins (CRPs) and CD47 similarly act through receptors CD200R and SIRP-a can reduce complement activation (Figure 4C) (Griffiths on inflammatory cells to prevent phagocytic attack (Figure 4C) et al., 2010). Thrombomodulin (CD141) binds and sequesters

238 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

Table 2. Major Classes and a Few Specific Examples of Intercellular Signaling Molecules that Regulate Cellular Responses to CNS Damage Molecule Source Lesion Compartment Phase Targets and Effects Class: Protease and Related Thrombin S LC, AS I Clot formation; astrocyte proliferation MMP-9 M, O, S LC, AS, PLP I-III OPC proliferation, remyelination; BBB leak; neovascular remodeling Kallikreins A, M, N, S LC, AS, PLP I-III Proinflammatory; demyelination Serpins A, M, O LC, PLP I-III Inhibit deleterious protease Class: NTs ATP, Glutamate N, O, A LC, AS, PLP I Microglia chemotaxis; reactive astrogliosis Class: DAMPs Alarmins: HMGB1, Damaged Cells LC, PLP I.II ‘‘Altered self’’ signals; proinflammatory; increase phagocytosis S100s, DNA PAMPs: LPS Microbes LC, PLP I.II ‘‘Nonself invader’’ signals; proinflammatory; increase phagocytosis Class: Growth Factors, Morphogens FGF A, N, E, LC, AS, PLP I-III Fibrotic scar; ECM; neovascular remodeling; reactive astrogliosis VEGF E, P, F, A LC, AS I-III BBB permeability; neovascular remodeling PDGF-B E, A LS, PLP I-III Neovascular remodeling PDGF-A Remyelination; OPC Proliferation Endothelin, N, A, O LC, AS II, III Astrocyte proliferation, glial scar formation EGF, BMP Class: Cytokines, Chemokines TGFb A, M, L AS, PLP I-III Fibrotic scar formation; ECM production; astrocyte proliferation IL-1b, TNFa, INFg A, M, L LC, AS, I, II Proinflammatory regulation IL-6, CCL2 A, M, L LC, AS I, II Leukocyte instruction, astrocyte scar formation Class: NIRegs CD141 A, O, E LC, AS I-III Protection of healthy self; resolution of inflammation CD200, CD47 N LC, AS, PLP I, II Protection of healthy self Class: Neural Remodeling Neurotrophins, N, A PLP III Synapse remodeling BDNF, etc. Perineuronal net A, OPC PLP III Restrict terminal sprouting Thmbs, C1q A, M PLP III Synapse formation and pruning A, astrocyte; E, endothelia; F, fibroblast; L, leukocyte; M, microglia; N, neuron; NTs, neurotransmitters; O, oligodendrocyte; OPC, O progenitor cell; S, serum; Thmbs, thrombospondin.

HMGB1, thereby reducing alarmin bioavailability (Figure 4C) described, but also molecules entering via leaky BBB, molecules (Abeyama et al., 2005). released by infiltrating leukocytes, and molecules released Thus, specific receptor-mediated signals that indicate by local cells including reactive glia themselves (Figure 5A) ‘‘altered self’’ or ‘‘healthy self’’ can tailor the nature of the reactive (Sofroniew, 2009). In response to these many different and often gliosis and multicellular responses to the type and severity of combinatorial molecular signals, reactive glia alter their gene cellular damage or death (Figures 4B and 4C). It is important to expression, structure, and function in selective and specific note that the final level of cellular damage inflicted by insults of ways, depending on the type and severity of the insults and de- all kinds is determined in part by prolonged periods of different pending on the specific combinations of molecular signals that types of secondary events (Figure 1D), including a balancing are impinging on them. It is important to emphasize that glia act among innate immune mechanisms that regulate the clear- do not exist in simple all-or-none ‘‘quiescent’’ or ‘‘activated’’ ance of ‘‘altered-self’’ debris and ‘‘non-self’’ pathogens, while states and that there is no single program of ‘‘reactive gliosis’’ preserving ‘‘healthy self’’ (Figure 4C) (Griffiths et al., 2010). These that is triggered in an all-or-none fashion and is similar in all events will impact on final size, location of glial scar, and situations. Instead, reactive glia can exhibit a vast range of re- signaling to perlesion perimeters (Figure 2). sponses as determined by combinations of specific signaling Reactive Gliosis events. For example, exposure to PAMPs such as lipopolysac- Reactive gliosis is regulated by a large array of different extracel- charide (LPS) can markedly skew the transcriptome of reactive lular signals generated after CNS insults. This array includes astrocytes toward chemokines, cytotoxicity, and inflammation not only molecules released by damaged or dead cells as just (Hamby et al., 2012; Zamanian et al., 2012). It is also noteworthy

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 239 Neuron Review

Figure 5. Multicellular and Multimolecular Regulation of Reactive Gliosis and BBB (A) Reactive gliosis can be induced, regulated, and influenced by a wide variety of molecular signals that can derive from many types of CNS intrinsic and extrinsic sources. (B) BBB permeability can be regulated by spe- cific molecular signaling cascades involving in- teractions among different cell types that converge on endothelial tight junctions (Argaw et al., 2009, 2012; Seo et al., 2013).

late (1) local reactive astrocytes to release VEGF and (2) local NG2-OPCs to release MMP-9, which in turn influence tight junc- tions and barrier properties of local endo- thelial cells and allow entry of serum proteins (including IgGs and signaling proteins such as thrombin, albumin, pro- teases, etc.) and leukocytes into local CNS parenchyma (Figure 5B) (Argaw et al., 2009, 2012; Seo et al., 2013). Entering leukocytes have the potential to generate more IL-1b and sustain local BBB permeability, creating the potential for a feedforward loop under adverse cir- cumstances. Understanding the molecu- lar mechanisms whereby such signaling is downregulated and the BBB reseals may help to reveal rational therapeutic in- terventions for certain conditions. Repair of the BBB after an uncomplicated acute injury generally occurs within several weeks (Figure 1D). Astrocyte Scar Formation Formation of compact astrocyte scars is a specialized aspect of reactive astrogliosis that occurs in response to severe tissue damage and leukocyte infiltration and involves phases of cell proliferation and cell organization (Figure 1)(Sofroniew and Vinters, 2010; Wanner et al., 2013). Molecular signals that regulate astrocyte that different combinations of stimulatory factors can lead to proliferation signals after CNS damage can derive from serum synergistic changes in molecular expression that could not be proteins or local cells and include thrombin, endothelin, FGF2, predicted from individual effects (Hamby et al., 2012). ATP, bone morphogenic proteins (BMPs), sonic hedge hog Information about the effects mediated by reactive glia in (SHH), and others (Table 2)(Gadea et al., 2008; Neary et al., response to specific signals is also steadily increasing and it is 2003; Sahni et al., 2010; Shirakawa et al., 2010; Sirko et al., clear that many aspects of reactive gliosis are part of coordi- 2013). The location of astrocyte scar borders around lesions nated multicellular innate and adaptive immune responses to can be influenced by interactions among proliferating or newly CNS insults. It is interesting to consider that certain molecules proliferated scar-forming astroglia, fibroblast-linage cells, and are emerging as potential coordinators of multicellular re- inflammatory cells (Wanner et al., 2013) and can be modulated sponses. One interesting set of converging data implicates IL- by transgenic manipulation of astrocyte intrinsic signaling cas- 1b as a key trigger that stimulates different types of glial cells cades involving STAT3, SOCS3, or NFkB, which can increase to modulate BBB permeability to serum proteins and leukocytes or decrease lesion sizes (Brambilla et al., 2005; Herrmann during reactive gliosis in response to CNS insults (Figure 5B). et al., 2008; Okada et al., 2006; Wanner et al., 2013), providing Numerous DAMPs and PAMPs generated by CNS insults will targets for potential intervention. Signals for organization of stimulate IL-1b release from microglia, and this IL-1b will stimu- newly proliferated astrocytes into compact scars include the

240 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

IL-6 receptor-STAT3 signaling system, which critically regulates damage such as trauma or stroke, as well as effectors to mitigate astrocyte scar formation after trauma, infection, and autoim- functional deterioration during degenerative conditions. Under- mune inflammation (Dro¨ gemu¨ ller et al., 2008; Haroon et al., standing and manipulating the signaling mechanisms and 2011; Herrmann et al., 2008; Wanner et al., 2013). effects of reactive gliosis in perilesion perimeter tissue has the Neural Remodeling in Perilesion Perimeters potential to reveal novel therapeutic strategies (Gleichman and Reactive gliosis is not only important in responses to very local Carmichael, 2013). CNS damage, but also is likely to play important roles in the neu- Blood-Brain Barrier ral remodeling that continues for prolonged times in perilesion The blood-brain barrier (BBB) is generated and regulated by a perimeters after acute focal insults such as stroke and trauma complex multicellular neurovascular unit that involves critical (Figures 1 and 2), as well as in chronic diffuse CNS insults interactions among endothelia, pericytes, astrocytes, and other (Figures 3) including neurodegenerative conditions such as Alz- cells (Abbott et al., 2006; Zlokovic, 2008). Compromise of the heimer’s disease. Such perilesion perimeter regions can extend BBB can occur in various manners and to various degrees, for considerable distances away from large focal lesions (Figures ranging from specific molecular signaling events that open 1 and 2) or can cover large areas of diffuse tissue damage (Fig- BBB permeability during certain types of reactive gliosis as dis- ures 3). In such cases, the associated reactive gliosis has the po- cussed above (Figure 5B), to severe breakdown of the BBB tential to impact on substantial territories of functioning neural caused by traumatic or ischemic endothelial destruction result- tissue. There is mounting evidence that the diverse changes in ing in the need for neovascularization (Figure 1). In addition to molecular signaling associated with this diffuse reactive gliosis overt trauma and ischemia, considerable evidence implicates a (Table 2) will impact on the ongoing neural remodeling and central role for BBB dysfunction in aging and various neurode- changes in neural function. Astrocytes interact with neurons on generative disorders. For example, both transgenically induced multiple spatial and temporal scales that can influence or modu- and aging-related pericyte deficiencies result in BBB breakdown late neural function in various direct and indirect ways, including that precedes neuronal degeneration (Bell et al., 2010). Pericyte by modulation of the extracellular balance of ions, transmitters, deficiency is also apparent in postmortem amyotrophic lateral and water critical for neural function, as well as direct influences sclerosis tissue (Winkler et al., 2013) and BBB dysfunction pre- on synaptic activity directly via release of ‘‘gliotransmitters’’ cedes neurodegeneration in a murine model (Zhong et al., (Halassa and Haydon, 2010; Hamilton and Attwell, 2010; Henne- 2008). Polymorphisms in APOE isoforms are implicated in berger et al., 2010; Volterra and Meldolesi, 2005). Microglia are increased vulnerability to BBB breakdown in neurodegenerative also implicated in participating in the regulation of synaptic turn- disorders and traumatic injury (Bell et al., 2012). over (Stephan et al., 2012). Perineuronal net molecules such as Regardless of triggering events, BBB leaks lead immediately chondroitin sulfate proteoglycans generated by astrocytes and to CNS parenchymal entry of serum proteins such as thrombin, NG2-OPC influence synaptic plasticity (Table 2)(Wang and Faw- albumin, immunoglobulins, and proteases (Figure 5B) that can cett, 2012). The effects of reactive gliosis on such functions is have a wide range of direct and indirect effects. Thrombin can only beginning to be understood, but reactive astrogliosis has signal directly via protease-activated receptors on various CNS been reported to impact on neural functions in various ways, cells including microglia and astrocytes (Shigetomi et al., 2008; including (1) by downregulation of astrocyte glutamine synthase Suo et al., 2002). Immunoglobulins contribute to clearing of path- that is associated with reduced inhibitory synaptic currents in ogens but may contribute to autoimmune dysfunctions. Platelets local neurons (Ortinski et al., 2010), (2) increased expression of can enter and form aggregates for clotting and release PDGFs, xCT (Slc7a11), a cysteine-glutamate transporter associated important for neovascular remodeling as well as for signaling with increased glutamate signaling, seizures, and excitotoxicity to pericytes and NG2 cells. Metaloproteases act on multiple pro- (Buckingham et al., 2011; Jackman et al., 2010), and (3) changes teins and contribute to extracellular matrix and tissue remodeling in the expression of multiple GPCRs and G proteins and calcium (Table 2). Serum kallikrein proteases activate inflammation. Tis- signaling evoked by their ligands that have the potential to alter sue kallikreins, which can interact with serum proteases as part astrocyte-neuron interactions (Hamby et al., 2012). There is of larger proteolytic cascades, are implicated in oligodendrocyte also a growing body of evidence that in response to inflammatory pathology, demyelination, axonopathy, and neuronal degenera- mediators and cytokines, reactive astrocytes can modulate their tion (Burda et al., 2013; Radulovic et al., 2013). Serpins inhibit expression of multiple molecules such as transmitter and ion and balance protease effects (Table 2). transporters, neuromodulators such as nitric oxide and prosta- In response to signals associated with BBB leak, local glial glandins, growth factors, and synapse regulatory proteins, which cells produce cytokines and chemokines that attract leukocytes, can impact on synaptic and neuronal functions including com- which on arrival produce additional cytokines and chemokines. plex behaviors such as sickness behavior, pain, appetite, sleep, The result is a rich extracellular mixture of molecular signals and mood, as reviewed in more detail elsewhere (Sofroniew, that can instruct or influence multiple cell types in lesion cores 2013). Taken together, such findings provide evidence that and can also diffuse away into surrounding CNS parenchyma different molecular mediators and signaling mechanisms associ- and influence neural cells that may not have been involved in ated with reactive gliosis can modulate functions of reactive as- the initial insult (Figure 2). These molecular gradients may influ- trocytes and microglia in different ways that have the potential to ence or take part in shaping cellular events in the perilesion pe- impact on synaptic and other neuronal functions in perilesion rimeters such as the long-lasting gradient of reactive gliosis and perimeters. Such signaling mechanisms are likely to have effects the long-lasting neural remodeling that occurs in perimeter tis- on the efficacy of training and functional rehabilitation after acute sue. The mix of molecular signals generated in lesion cores

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 241 Neuron Review

changes over time as the injury response progresses through Impact of Peripheral Infections on Responses to CNS different phases and the BBB leak is repaired and different cell Insults types occupy this area and produce different effector molecules Many of the cell types responding to CNS damage are exqui- (Figure 1D; Table 2). Depending on the severity of the insult, sitely sensitive to molecular cues associated with microbial lesion cores exhibit a BBB leak for at least several days up to infection. This is not surprising because limiting infection spread 2 weeks throughout the initial period (Phase I) of damage is likely to have shaped the evolution of injury responses in all tis- response (Figure 1D). BBB repair requires the presence of func- sues including CNS. Various features of the response to infection tional astrocytes (Bush et al., 1999). Release of the protein such as production of cytotoxins and certain types of inflamma- HMGB1 by reactive astrocytes promotes endothelial cell neo- tion can damage host cells as well as microbes. Triggering such vascular remodeling, whereas blockade of HMGB1 release by responses in the absence of overt infection has the potential to reactive astrocytes prevents neovascular remodeling and cause or exacerbate tissue damage. Thus, certain responses worsens neurological deficits (Hayakawa et al., 2012). In com- that may be beneficial in microbial infection may be detrimental plex, chronic, or diffuse insults, BBB leaks may persist and if triggered during sterile (uninfected) tissue damage after have long-lasting effects on extended areas of neural tissue in trauma, stroke, degenerative disease, or autoimmune attack. perilesion perimeters (Figure 3), and late BBB disruption may In this regard, it is important to recognize that reactive glia, lead to secondary tissue injury (Zlokovic, 2008). Even when both microglia and astrocytes, that are responding to a sterile BBB leaks are repaired, lesion cores continue to contain collec- primary CNS insult can be influenced by peripheral infections tions of nonneural cells within the BBB that have the potential to that generate high levels of circulating cytokines and other continue producing high levels of many different signaling mole- inflammatory mediators such as LPS and other PAMPs. This cules for prolonged periods. Thus, lesion core tissue has the po- signaling can occur through blood-borne cytokines or LPS tential to generate diffusion gradients of many potent molecules released at the site of peripheral infection and entering through that spread into neighboring tissue parenchyma and form gradi- the leaky BBB in the injury core during phase I of the damage ents of molecular signals that influence host cells (Figure 2). It is response after focal insults or through BBB leaks triggered by also noteworthy that while the BBB is open, there is also the neurodegenerative or autoimmune processes (Figures 1A and potential for entry of various pathogen-associated molecular 5A). LPS and cytokines have powerful effects both on reactive patterns (PAMPs) (Table 2) that are generated by microbial microglia (Perry, 2010; Perry et al., 2007) and on reactive astro- infections in the periphery and that can substantively alter the cytes (Hamby et al., 2012; Zamanian et al., 2012) that can drive response of local cells to CNS damage, even when there is no transcriptome profiles and wound response toward proinflam- local infection, as discussed below. matory and potentially cytotoxic phenotypes over prolonged Infiltrating Leukocytes times in ways that would not normally be implemented in sterile Leukocytes that infiltrate from the circulation provide a major CNS insults and that can exacerbate tissue damage. Congruent source of molecular signaling during responses to CNS with such experimental observations is clinical epidemiological damage. These molecular signals include a diverse mix of evidence indicating that peripheral infections have a negative cytokines, chemokines, and growth factors (Table 2) that can impact on neurological outcome after spinal cord injury (Failli instruct and modify the functions of local reactive glia et al., 2012). (Figure 5A) and other intrinsic CNS cells as well as other local leukocytes. It is likely that the molecular signals produced by Common Features, Differences, and Building Models leukocytes, and their functional effects, will vary during different Given the large number of potential CNS insults and the consid- phases of response to acute or chronic insults (Figure 1) and erable amount of variation among the multicellular responses range from those mediating classical inflammatory responses they elicit, looking for common features and clear differences with cytotoxic and phagocytic activities, to those influencing may provide valuable insights regarding shared fundamental tissue repair and remodeling. Although temporal changes mechanisms. Understanding basic factors that drive responses during wound responses are not yet well studied, different to one type of insult may inform ideas regarding responses to functional roles for different leukocyte subtypes are gradually another. In this regard, it is interesting to consider that evolu- coming into focus. For example, in response to stimulation by tionary pressures shaping responses to CNS damage are likely different cytokines, monocytes generate macrophages with to have favored rapid responses to small CNS injuries that distinct functional phenotypes that produce different effector were not functionally incapacitating and that kept such injuries molecules (Mosser and Edwards, 2008). In response to INFg, small and uninfected or that kept small infections from classically activated macrophages generate cytotoxic antimi- spreading. One efficient means of doing so would be to isolate crobial molecules and phagocytose dead cells and debris small focal injuries or small infections with cellular barriers that (Figure 4C), whereas in response to stimulation with IL-4 or effectively wall off these lesions, allowing a robust inflammatory IL-10, macrophages take part in wound healing and anti-inflam- (and antimicrobial) responses in lesion cores, while preserving as matory activities (Mosser and Edwards, 2008). Similarly, much adjacent neural tissue as possible (Sofroniew, 2005). In T cells stimulated by different cytokines become polarized this Review, based on available information, we present a basic toward different activities, such as TH1, TReg, and TH17 cells, model of multicellular responses to different types of focal CNS which can exert very different effects ranging from cell killing tissue damage that is compatible with this notion (Figures 1, 2, to participation in tissue repair (MacIver et al., 2013; Mills, and 3). Certain common features in this model of cellular re- 2011; Walsh and Kipnis, 2011). sponses to CNS damage could be interpreted as serving the

242 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

Figure 6. Lesions as Targets for Therapeutic Strategies Nonneural lesion cores (LC) represent targets for biomaterial depots (A) or grafts of exogenous neural stem/progenitor cells (B), or mesenchymal stem cells (C) as potential therapeutic strategies. Molecules delivered by biomaterial depots (A) or certain cell grafts (C) have potential to support and attract axon growth (arrow a), modulate cells in perilesion perimeter (PLP) (arrow b), modulate cells in LC (arrow c), or modulate astrocyte scar (AS) (arrow d). New neurons generated by stem/ progenitor cell grafts can receive and send neural connects in PLP and beyond and may be able to form new, functional relay circuits (B). basic primitive function of preventing or limiting the spread of potential for identifying ways to manipulate these responses to focal infection. Viewing certain cellular responses to other CNS improve outcome. Manipulations under intense investigation insults in this light may provide insights regarding basic mecha- include not only traditional approaches such as development nisms and how to understand and eventually manipulate re- of drug candidates for parenteral pharmacological delivery, but sponses safely to improve outcomes. We of course recognize also lifestyle modifications such as reduced energy consumption that not all responses will be determined or influenced in this and implementation of exercise strategies intended to reduce manner. Nevertheless, the powerful and fundamental role of inflammatory responses, cellular degeneration, and functional response to infection is likely to be ancestral to all responses decline after stroke and traumatic injury (Arumugam et al., to CNS damage and therefore has the potential to influence all 2010; Mattson, 2012; Piao et al., 2013). In addition, novel inter- types of responses that evolved subsequently. This point is un- ventional strategies in the form of biomaterial implants as derscored by the growing recognition of the role of inflammatory molecular depots for local molecular delivery, as well as tissue mechanisms in different neurodegenerative diseases, and the regenerative approaches based on transplantation of progenitor potential for molecules released by peripheral infections to influ- or stem cells, are showing considerable promise for future appli- ence far-distant reactive gliosis as discussed above. It is impor- cation in neural repair and regeneration. The nonneural lesion tant to realize that the multicellular responses to diffuse CNS cores of focal lesions may represent particularly useful targets insults are likely to have been shaped by adapting already exist- for such approaches (Figure 6). For example, biomaterial im- ing responses to focal traumatic damage or focal infection. plants, particularly in the form of injectable hydrogels, have po- This notion is likely to apply not only to degenerative diseases, tential to serve both as scaffolds and depots that can influence but also to autoimmune-mediated damage, where the CNS tissue remodeling and cellular functions (Pakulska et al., 2012). response to such damage bears striking similarities to responses Large lesion cores of nonneural tissue after stroke or severe focal to other forms of CNS tissue damage. This discussion is meant to trauma may represent useful sites for placement of biomaterial highlight the usefulness of constructing models of how multiple depots that release gradients of molecules to influence locally cells interact and how multiple molecules drive these interac- surrounding neural tissue without inflicting additional damage tions during the responses to different types of CNS damage. (Figure 6A). For example, injectable hydrogel depots can provide The models we have presented here are compatible with basic sustained delivery of different kinds of bioactive molecules, evidence currently available but no doubt will require modifica- including growth factors or small hydrophobic molecules to influ- tion as information accrues. We hope that they are flexible ence gene expression and function of local cells over prolonged enough to accommodate modifications and can serve as useful subacute times, after which they are degraded without substan- frameworks. tive tissue damage (Pakulska et al., 2012; Song et al., 2012; Yang et al., 2009; Zhang et al., 2014). Such molecular delivery has the Manipulating CNS Wound Responses to Promote Repair potential to influence not only the ongoing repair in the subacute As the cell biology and molecular signaling that underlie re- period after injury such as scar formation, but also can influence sponses to CNS damage become defined, there is increasing cells in the large perimeter zone around CNS lesions that

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 243 Neuron Review

undergoes continual tissue remodeling for prolonged periods ACKNOWLEDGMENTS after tissue damage (Figure 6A). Delivery of molecules via depots placed into lesion cores have the potential to influence the neural The authors are supported by the NIH (NINDS), The Dr. Miriam and Sheldon G. Adelson Medical Foundation, and Wings for Life. plasticity and tissue remodeling in this perimeter region (Clark- son et al., 2010). REFERENCES Various cellular regenerative approaches may have potential for CNS repair (Bliss et al., 2007; Martino and Pluchino, 2006; Abbott, N.J., Ro¨ nnba¨ ck, L., and Hansson, E. (2006). Astrocyte-endothelial Snyder and Teng, 2012). Delivery of neural-lineage progenitor interactions at the blood-brain barrier. Nat. Rev. Neurosci. 7, 41–53. or stem cells into lesion cores (Figure 6B) can lead to the gener- Abematsu, M., Tsujimura, K., Yamano, M., Saito, M., Kohno, K., Kohyama, J., ation of local neurons that form afferent and efferent connections Namihira, M., Komiya, S., and Nakashima, K. (2010). Neurons derived from with host and have the potential to form functionally integrated transplanted neural stem cells restore disrupted neuronal circuitry in a mouse model of spinal cord injury. J. Clin. Invest. 120, 3255–3266. circuits (Abematsu et al., 2010; Lu et al., 2012). Stromal or mesenchymal stem cells derived from bone marrow or umbilical Abeyama, K., Stern, D.M., Ito, Y., Kawahara, K., Yoshimoto, Y., Tanaka, M., cord, delivered by peripheral (intravascular) or CNS injection, Uchimura, T., Ida, N., Yamazaki, Y., Yamada, S., et al. (2005). The N-terminal domain of thrombomodulin sequesters high-mobility group-B1 protein, a can home to injury sites (Figure 6C), where they can release novel antiinflammatory mechanism. J. Clin. Invest. 115, 1267–1274. growth factors, cytokines, and other factors that may influence Aldrich, A., and Kielian, T. (2011). Central nervous system fibrosis is associated the response to damage and repair process (Bliss et al., 2007; with fibrocyte-like infiltrates. Am. J. Pathol. 179, 2952–2962. Kokaia et al., 2012; Lemmens and Steinberg, 2013). Such grafting studies are being intensely pursued but are still in the Argaw, A.T., Gurfein, B.T., Zhang, Y., Zameer, A., and John, G.R. (2009). VEGF-mediated disruption of endothelial CLN-5 promotes blood-brain barrier early stages of development. Beneficial effects are reported on breakdown. Proc. Natl. Acad. Sci. USA 106, 1977–1982. cellular events such as immunomodulation, survival of CNS Argaw, A.T., Asp, L., Zhang, J., Navrazhina, K., Pham, T., Mariani, J.N., Ma- intrinsic cells including neurons, and growth of axons; however, hase, S., Dutta, D.J., Seto, J., Kramer, E.G., et al. (2012). Astrocyte-derived limited information is as yet available on overall functional VEGF-A drives blood-brain barrier disruption in CNS inflammatory disease. efficacy. J. Clin. Invest. 122, 2454–2468. Arumugam, T.V., Phillips, T.M., Cheng, A., Morrell, C.H., Mattson, M.P., and Concluding Remarks Wan, R. (2010). Age and energy intake interact to modify cell stress pathways and stroke outcome. Ann. Neurol. 67, 41–52. The study of cellular responses to CNS injury has progressed enormously in the last two decades, as have ideas about their Askari, A.T., Unzek, S., Popovic, Z.B., Goldman, C.K., Forudi, F., Kiedrowski, functions and mechanisms. It is now clear that reactive gliosis M., Rovner, A., Ellis, S.G., Thomas, J.D., DiCorleto, P.E., et al. (2003). Effect of stromal-cell-derived factor 1 on stem-cell homing and tissue regeneration in and multicellular responses to CNS damage are a complex ischaemic cardiomyopathy. Lancet 362, 697–703. mixture of events that balance clearance of dead cells, patho- Astrup, J., Siesjo¨ , B.K., and Symon, L. (1981). Thresholds in cerebral ischemia gens, and debris with maximal preservation of adjacent local tis- - the ischemic penumbra. Stroke 12, 723–725. sue. These events are regulated by a vast number of molecular signals that control specific cellular functions as dictated by spe- Attwell, D., Buchan, A.M., Charpak, S., Lauritzen, M., Macvicar, B.A., and Newman, E.A. (2010). Glial and neuronal control of brain blood flow. Nature cific situations. Reactive gliosis is not all-or-none or stereotypic; 468, 232–243. instead, it is highly variable and context specific. Loss-of- Bardehle, S., Kru¨ ger, M., Buggenthin, F., Schwausch, J., Ninkovic, J., Clevers, function studies show that reactive gliosis and multicellular re- H., Snippert, H.J., Theis, F.J., Meyer-Luehmann, M., Bechmann, I., et al. sponses to CNS damage exert essential beneficial functions (2013). Live imaging of astrocyte responses to acute injury reveals selective without which tissue damage (and function) would increase juxtavascular proliferation. Nat. Neurosci. 16, 580–586. and tissue repair would not occur. Nevertheless, over activity Bareyre, F.M., Kerschensteiner, M., Raineteau, O., Mettenleiter, T.C., Wein- of certain functions may in some cases exacerbate tissue dam- mann, O., and Schwab, M.E. (2004). The injured spinal cord spontaneously 7 age. Interventions to block such activities will need to target spe- forms a new intraspinal circuit in adult rats. Nat. Neurosci. , 269–277. cific molecular events rather than attempt to block the process of Barres, B.A. (2008). The mystery and magic of glia: a perspective on their roles gliosis per se. in health and disease. Neuron 60, 430–440. Understanding the ways in which dysfunction of reactive glio- Bell, R.D., Winkler, E.A., Sagare, A.P., Singh, I., LaRue, B., Deane, R., and Zlo- sis and multicellular responses to CNS damage can occur, for kovic, B.V. (2010). Pericytes control key neurovascular functions and neuronal phenotype in the adult brain and during brain aging. Neuron 68, 409–427. example, through genetic mutations or polymorphisms, has the potential to identify new mechanisms that may underlie Bell, R.D., Winkler, E.A., Singh, I., Sagare, A.P., Deane, R., Wu, Z., Holtzman, specific CNS disorders or that may underlie population vari- D.M., Betsholtz, C., Armulik, A., Sallstrom, J., et al. (2012). Apolipoprotein E controls cerebrovascular integrity via cyclophilin A. Nature 485, 512–516. ability in responses to similar insults such as stroke or trauma. There is a compelling need to deepen our understanding of Benner, E.J., Luciano, D., Jo, R., Abdi, K., Paez-Gonzalez, P., Sheng, H., Warner, D.S., Liu, C., Eroglu, C., and Kuo, C.T. (2013). Protective astrogenesis specific cellular interactions during responses to CNS insults, from the SVZ niche after injury is controlled by Notch modulator Thbs4. Nature and how these change over time and are governed by complex 497, 369–373. molecular signaling mechanisms. Such information is funda- Bianco, P., Riminucci, M., Gronthos, S., and Robey, P.G. (2001). Bone marrow mental to understanding most CNS disorders and for devel- stromal stem cells: nature, biology, and potential applications. Stem Cells 19, oping rationally targeted therapeutic strategies that can safely 180–192. manipulate or modify cellular responses to CNS insults in Bliss, T., Guzman, R., Daadi, M., and Steinberg, G.K. (2007). Cell transplanta- ways that improve outcome. tion therapy for stroke. Stroke Suppl. 38, 817–826.

244 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

Boille´ e, S., Vande Velde, C., and Cleveland, D.W. (2006). ALS: a disease of Failli, V., Kopp, M.A., Gericke, C., Martus, P., Klingbeil, S., Brommer, B., Lagi- motor neurons and their nonneuronal neighbors. Neuron 52, 39–59. nha, I., Chen, Y., DeVivo, M.J., Dirnagl, U., and Schwab, J.M. (2012). Func- tional neurological recovery after spinal cord injury is impaired in patients Brambilla, R., Bracchi-Ricard, V., Hu, W.H., Frydel, B., Bramwell, A., Karmally, with infections. Brain 135, 3238–3250. S., Green, E.J., and Bethea, J.R. (2005). Inhibition of astroglial nuclear factor kappaB reduces inflammation and improves functional recovery after spinal Faulkner, J.R., Herrmann, J.E., Woo, M.J., Tansey, K.E., Doan, N.B., and cord injury. J. Exp. Med. 202, 145–156. Sofroniew, M.V. (2004). Reactive astrocytes protect tissue and preserve func- tion after spinal cord injury. J. Neurosci. 24, 2143–2155. Brenner, M., Johnson, A.B., Boespflug-Tanguy, O., Rodriguez, D., Goldman, J.E., and Messing, A. (2001). Mutations in GFAP, encoding glial fibrillary acidic Franklin, R.J., and Ffrench-Constant, C. (2008). Remyelination in the CNS: protein, are associated with Alexander disease. Nat. Genet. 27, 117–120. from biology to therapy. Nat. Rev. Neurosci. 9, 839–855.

Buckingham, S.C., Campbell, S.L., Haas, B.R., Montana, V., Robel, S., Ogun- Frohman, E.M., Racke, M.K., and Raine, C.S. (2006). Multiple sclerosis—the rinu, T., and Sontheimer, H. (2011). Glutamate release by primary brain tumors plaque and its pathogenesis. N. Engl. J. Med. 354, 942–955. induces epileptic activity. Nat. Med. 17, 1269–1274. Gadea, A., Schinelli, S., and Gallo, V. (2008). Endothelin-1 regulates astrocyte Burda, J.E., Radulovic, M., Yoon, H., and Scarisbrick, I.A. (2013). Critical role proliferation and reactive gliosis via a JNK/c-Jun signaling pathway. for PAR1 in kallikrein 6-mediated oligodendrogliopathy. Glia 61, 1456–1470. J. Neurosci. 28, 2394–2408.

Bush, T.G., Puvanachandra, N., Horner, C.H., Polito, A., Ostenfeld, T., Svend- Gao, H.M., Zhou, H., Zhang, F., Wilson, B.C., Kam, W., and Hong, J.S. (2011). sen, C.N., Mucke, L., Johnson, M.H., and Sofroniew, M.V. (1999). Leukocyte HMGB1 acts on microglia Mac1 to mediate chronic neuroinflammation that infiltration, neuronal degeneration, and neurite outgrowth after ablation of drives progressive neurodegeneration. J. Neurosci. 31, 1081–1092. scar-forming, reactive astrocytes in adult transgenic mice. Neuron 23, 297–308. Garcia, A.D.R., Doan, N.B., Imura, T., Bush, T.G., and Sofroniew, M.V. (2004). GFAP-expressing progenitors are the principal source of constitutive neuro- 7 Carmichael, S.T. (2006). Cellular and molecular mechanisms of neural repair genesis in adult mouse forebrain. Nat. Neurosci. , 1233–1241. after stroke: making waves. Ann. Neurol. 59, 735–742. Garcı´a-Alı´as, G., Barkhuysen, S., Buckle, M., and Fawcett, J.W. (2009). Chon- Casella, G.T., Marcillo, A., Bunge, M.B., and Wood, P.M. (2002). New vascular droitinase ABC treatment opens a window of opportunity for task-specific 12 tissue rapidly replaces neural parenchyma and vessels destroyed by a contu- rehabilitation. Nat. Neurosci. , 1145–1151. sion injury to the rat spinal cord. Exp. Neurol. 173, 63–76. Gleichman, A.J., and Carmichael, S.T. (2013). Astrocytic therapies for neuronal Chan, J.K., Roth, J., Oppenheim, J.J., Tracey, K.J., Vogl, T., Feldmann, M., repair in stroke. Neurosci. Lett. Published online October 31, 2013. http://dx. Horwood, N., and Nanchahal, J. (2012). Alarmins: awaiting a clinical response. doi.org/10.1016/j.neulet.2013.10.055. J. Clin. Invest. 122, 2711–2719. Go¨ ritz, C., Dias, D.O., Tomilin, N., Barbacid, M., Shupliakov, O., and Frise´ n, J. 333 Clarkson, A.N., Huang, B.S., Macisaac, S.E., Mody, I., and Carmichael, S.T. (2011). A pericyte origin of spinal cord scar tissue. Science , 238–242. (2010). Reducing excessive GABA-mediated tonic inhibition promotes func- Greaves, N.S., Ashcroft, K.J., Baguneid, M., and Bayat, A. (2013). Current tional recovery after stroke. Nature 468, 305–309. understanding of molecular and cellular mechanisms in fibroplasia and angio- genesis during acute wound healing. J. Dermatol. Sci. 72, 206–217. Courtine, G., Song, B., Roy, R.R., Zhong, H., Herrmann, J.E., Ao, Y., Qi, J., Edgerton, V.R., and Sofroniew, M.V. (2008). Recovery of supraspinal control Griffiths, M.R., Gasque, P., and Neal, J.W. (2010). The regulation of the CNS of stepping via indirect propriospinal relay connections after spinal cord injury. 14 innate immune response is vital for the restoration of tissue homeostasis Nat. Med. , 69–74. (repair) after acute brain injury: a brief review. Int. J. Inflamm. 2010, 151097.

Daneman, R., Zhou, L., Kebede, A.A., and Barres, B.A. (2010). Pericytes are Gurtner, G.C., Werner, S., Barrandon, Y., and Longaker, M.T. (2008). Wound 468 required for blood-brain barrier integrity during embryogenesis. Nature , repair and regeneration. Nature 453, 314–321. 562–566. Halassa, M.M., and Haydon, P.G. (2010). Integrated brain circuits: astrocytic Davalos, D., Grutzendler, J., Yang, G., Kim, J.V., Zuo, Y., Jung, S., Littman, networks modulate neuronal activity and behavior. Annu. Rev. Physiol. 72, D.R., Dustin, M.L., and Gan, W.B. (2005). ATP mediates rapid microglial 335–355. response to local brain injury in vivo. Nat. Neurosci. 8, 752–758. Hamby, M.E., Coppola, G., Ao, Y., Geschwind, D.H., Khakh, B.S., and Sofro- Davis, G.E., and Senger, D.R. (2005). Endothelial extracellular matrix: niew, M.V. (2012). Inflammatory mediators alter the astrocyte transcriptome biosynthesis, remodeling, and functions during vascular morphogenesis and and calcium signaling elicited by multiple G-protein-coupled receptors. 97 neovessel stabilization. Circ. Res. , 1093–1107. J. Neurosci. 32, 14489–14510.

DeKosky, S.T., Blennow, K., Ikonomovic, M.D., and Gandy, S. (2013). Acute Hamilton, N.B., and Attwell, D. (2010). Do astrocytes really exocytose neuro- and chronic traumatic encephalopathies: pathogenesis and biomarkers. Nat. transmitters? Nat. Rev. Neurosci. 11, 227–238. Rev. Neurol. 9, 192–200. Hanisch, U.K., and Kettenmann, H. (2007). Microglia: active sensor and Derecki, N.C., Cronk, J.C., Lu, Z., Xu, E., Abbott, S.B., Guyenet, P.G., and versatile effector cells in the normal and pathologic brain. Nat. Neurosci. 10, Kipnis, J. (2012). Wild-type microglia arrest pathology in a mouse model of 1387–1394. Rett syndrome. Nature 484, 105–109. Haroon, F., Dro¨ gemu¨ ller, K., Ha¨ ndel, U., Brunn, A., Reinhold, D., Nishanth, G., Devinsky, O., Vezzani, A., Najjar, S., De Lanerolle, N.C., and Rogawski, M.A. Mueller, W., Trautwein, C., Ernst, M., Deckert, M., and Schlu¨ ter, D. (2011). (2013). Glia and epilepsy: excitability and inflammation. Trends Neurosci. 36, Gp130-dependent astrocytic survival is critical for the control of autoimmune 174–184. central nervous system inflammation. J. Immunol. 186, 6521–6531.

Dirnagl, U., Iadecola, C., and Moskowitz, M.A. (1999). Pathobiology of ischae- Hayakawa, K., Pham, L.D., Katusic, Z.S., Arai, K., and Lo, E.H. (2012). Astro- mic stroke: an integrated view. Trends Neurosci. 22, 391–397. cytic high-mobility group box 1 promotes endothelial progenitor cell-mediated neurovascular remodeling during stroke recovery. Proc. Natl. Acad. Sci. USA Dro¨ gemu¨ ller, K., Helmuth, U., Brunn, A., Sakowicz-Burkiewicz, M., Gutmann, 109, 7505–7510. D.H., Mueller, W., Deckert, M., and Schlu¨ ter, D. (2008). Astrocyte gp130 expression is critical for the control of Toxoplasma encephalitis. J. Immunol. Henneberger, C., Papouin, T., Oliet, S.H., and Rusakov, D.A. (2010). Long- 181, 2683–2693. term potentiation depends on release of D-serine from astrocytes. Nature 463, 232–236. Dugas, J.C., Mandemakers, W., Rogers, M., Ibrahim, A., Daneman, R., and Barres, B.A. (2008). A novel purification method for CNS projection neurons Hermanns, S., Klapka, N., Gasis, M., and Mu¨ ller, H.W. (2006). The collagenous leads to the identification of brain vascular cells as a source of trophic support wound healing scar in the injured central nervous system inhibits axonal regen- for corticospinal motor neurons. J. Neurosci. 28, 8294–8305. eration. Adv. Exp. Med. Biol. 557, 177–190.

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 245 Neuron Review

Herrmann, J.E., Imura, T., Song, B., Qi, J., Ao, Y., Nguyen, T.K., Korsak, R.A., London, A., Cohen, M., and Schwartz, M. (2013). Microglia and monocyte- Takeda, K., Akira, S., and Sofroniew, M.V. (2008). STAT3 is a critical regulator derived macrophages: functionally distinct populations that act in concert in of astrogliosis and scar formation after spinal cord injury. J. Neurosci. 28, CNS plasticity and repair. Front. Cell. Neurosci. 7, 34. 7231–7243. Louis, D.N. (2006). Molecular pathology of malignant gliomas. Annu. Rev. Hughes, E.G., Kang, S.H., Fukaya, M., and Bergles, D.E. (2013). Oligodendro- Pathol. 1, 97–117. cyte progenitors balance growth with self-repulsion to achieve homeostasis in the adult brain. Nat. Neurosci. 16, 668–676. Loy, D.N., Crawford, C.H., Darnall, J.B., Burke, D.A., Onifer, S.M., and Whitte- more, S.R. (2002). Temporal progression of angiogenesis and basal lamina Jabs, R., Seifert, G., and Steinha¨ user, C. (2008). Astrocytic function and its deposition after contusive spinal cord injury in the adult rat. J. Comp. Neurol. alteration in the epileptic brain. Epilepsia 49 (Suppl 2 ), 3–12. 445, 308–324.

Jackman, N.A., Uliasz, T.F., Hewett, J.A., and Hewett, S.J. (2010). Regulation Lu, P., Wang, Y., Graham, L., McHale, K., Gao, M., Wu, D., Brock, J., Blesch, of system x(c)(-)activity and expression in astrocytes by interleukin-1b: impli- A., Rosenzweig, E.S., Havton, L.A., et al. (2012). Long-distance growth and cations for hypoxic neuronal injury. Glia 58, 1806–1815. connectivity of neural stem cells after severe spinal cord injury. Cell 150, 1264–1273. Jaerve, A., Bosse, F., and Mu¨ ller, H.W. (2012). SDF-1/CXCL12: its role in spinal cord injury. Int. J. Biochem. Cell Biol. 44, 452–456. Lucchinetti, C.F., Bru¨ ck, W., Rodriguez, M., and Lassmann, H. (1996). Distinct patterns of multiple sclerosis pathology indicates heterogeneity on pathogen- Kawaja, M.D., and Gage, F.H. (1991). Reactive astrocytes are substrates for esis. Brain Pathol. 6, 259–274. the growth of adult CNS axons in the presence of elevated levels of nerve growth factor. Neuron 7, 1019–1030. MacIver, N.J., Michalek, R.D., and Rathmell, J.C. (2013). Metabolic regulation of T lymphocytes. Annu. Rev. Immunol. 31, 259–283. Kimura-Kuroda, J., Teng, X., Komuta, Y., Yoshioka, N., Sango, K., Kawamura, K., Raisman, G., and Kawano, H. (2010). An in vitro model of the inhibition of Maragakis, N.J., and Rothstein, J.D. (2006). Mechanisms of Disease: astro- axon growth in the lesion scar formed after central nervous system injury. cytes in neurodegenerative disease. Nat. Clin. Pract. Neurol. 2, 679–689. Mol. Cell. Neurosci. 43, 177–187. Martino, G., and Pluchino, S. (2006). The therapeutic potential of neural stem Klapka, N., and Mu¨ ller, H.W. (2006). Collagen matrix in spinal cord injury. cells. Nat. Rev. Neurosci. 7, 395–406. J. Neurotrauma 23, 422–435. Mattson, M.P. (2000). Apoptosis in neurodegenerative disorders. Nat. Rev. Kokaia, Z., Martino, G., Schwartz, M., and Lindvall, O. (2012). Cross-talk Mol. Cell Biol. 1, 120–129. between neural stem cells and immune cells: the key to better brain repair? 15 Nat. Neurosci. , 1078–1087. Mattson, M.P. (2012). Energy intake and exercise as determinants of brain health and vulnerability to injury and disease. Cell Metab. 16, 706–722. Kono, H., and Rock, K.L. (2008). How dying cells alert the immune system to danger. Nat. Rev. Immunol. 8, 279–289. McIntosh, T.K., Vink, R., Noble, L., Yamakami, I., Fernyak, S., Soares, H., and Faden, A.L. (1989). Traumatic brain injury in the rat: characterization of a lateral Kraft, A.W., Hu, X., Yoon, H., Yan, P., Xiao, Q., Wang, Y., Gil, S.C., Brown, J., fluid-percussion model. Neuroscience 28, 233–244. Wilhelmsson, U., Restivo, J.L., et al. (2013). Attenuating astrocyte activation accelerates plaque pathogenesis in APP/PS1 mice. FASEB J. 27, 187–198. Meletis, K., Barnabe´ -Heider, F., Carle´ n, M., Evergren, E., Tomilin, N., Shuplia- kov, O., and Frise´ n, J. (2008). Spinal cord injury reveals multilineage differen- Kreutzberg, G.W. (1996). Microglia: a sensor for pathological events in the tiation of ependymal cells. PLoS Biol. 6, e182. CNS. Trends Neurosci. 19, 312–318. Midwood, K.S., Williams, L.V., and Schwarzbauer, J.E. (2004). Tissue repair Kriegstein, A., and Alvarez-Buylla, A. (2009). The glial nature of embryonic and 36 adult neural stem cells. Annu. Rev. Neurosci. 32, 149–184. and the dynamics of the extracellular matrix. Int. J. Biochem. Cell Biol. , 1031–1037. Lagace, D.C. (2012). Does the endogenous neurogenic response alter behav- ioral recovery following stroke? Behav. Brain Res. 227, 426–432. Mills, K.H. (2011). TLR-dependent T cell activation in autoimmunity. Nat. Rev. Immunol. 11, 807–822. Lemmens, R., and Steinberg, G.K. (2013). Stem cell therapy for acute cerebral injury: what do we know and what will the future bring? Curr. Opin. Neurol. 26, Mosser, D.M., and Edwards, J.P. (2008). Exploring the full spectrum of macro- 8 617–625. phage activation. Nat. Rev. Immunol. , 958–969.

Lennon, V.A., Kryzer, T.J., Pittock, S.J., Verkman, A.S., and Hinson, S.R. Myer, D.J., Gurkoff, G.G., Lee, S.M., Hovda, D.A., and Sofroniew, M.V. (2006). (2005). IgG marker of optic-spinal multiple sclerosis binds to the aquaporin- Essential protective roles of reactive astrocytes in traumatic brain injury. Brain 129 4 water channel. J. Exp. Med. 202, 473–477. , 2761–2772.

Li, L., Lundkvist, A., Andersson, D., Wilhelmsson, U., Nagai, N., Pardo, A.C., Neary, J.T., Kang, Y., Willoughby, K.A., and Ellis, E.F. (2003). Activation of Nodin, C., Sta˚ hlberg, A., Aprico, K., Larsson, K., et al. (2008). Protective role extracellular signal-regulated kinase by stretch-induced injury in astrocytes of reactive astrocytes in brain ischemia. J. Cereb. Blood Flow Metab. 28, involves extracellular ATP and P2 purinergic receptors. J. Neurosci. 23, 468–481. 2348–2356.

Li, S., Overman, J.J., Katsman, D., Kozlov, S.V., Donnelly, C.J., Twiss, J.L., Nimmerjahn, A., Kirchhoff, F., and Helmchen, F. (2005). Resting microglial cells Giger, R.J., Coppola, G., Geschwind, D.H., and Carmichael, S.T. (2010). An are highly dynamic surveillants of brain parenchyma in vivo. Science 308, age-related sprouting transcriptome provides molecular control of axonal 1314–1318. sprouting after stroke. Nat. Neurosci. 13, 1496–1504. Nishiyama, A., Komitova, M., Suzuki, R., and Zhu, X. (2009). Polydendrocytes Lindvall, O., and Kokaia, Z. (2006). Stem cells for the treatment of neurological (NG2 cells): multifunctional cells with lineage plasticity. Nat. Rev. Neurosci. 10, disorders. Nature 441, 1094–1096. 9–22.

Liu, G.J., Nagarajah, R., Banati, R.B., and Bennett, M.R. (2009). Glutamate Ohab, J.J., and Carmichael, S.T. (2008). Poststroke neurogenesis: emerging induces directed chemotaxis of microglia. Eur. J. Neurosci. 29, 1108–1118. principles of migration and localization of immature neurons. Neuroscientist 14, 369–380. Lo, E.H. (2008). A new penumbra: transitioning from injury into repair after stroke. Nat. Med. 14, 497–500. Okada, S., Nakamura, M., Katoh, H., Miyao, T., Shimazaki, T., Ishii, K., Yamane, J., Yoshimura, A., Iwamoto, Y., Toyama, Y., and Okano, H. (2006). Logan, A., and Berry, M. (2002). Cellular and molecular determinants of glial Conditional ablation of Stat3 or Socs3 discloses a dual role for reactive astro- scar formation. Adv. Exp. Med. Biol. 513, 115–158. cytes after spinal cord injury. Nat. Med. 12, 829–834.

246 Neuron 81, January 22, 2014 ª2014 Elsevier Inc. Neuron Review

Ortinski, P.I., Dong, J., Mungenast, A., Yue, C., Takano, H., Watson, D.J., Hay- Silver, D.J., Siebzehnrubl, F.A., Schildts, M.J., Yachnis, A.T., Smith, G.M., don, P.G., and Coulter, D.A. (2010). Selective induction of astrocytic gliosis Smith, A.A., Scheffler, B., Reynolds, B.A., Silver, J., and Steindler, D.A. generates deficits in neuronal inhibition. Nat. Neurosci. 13, 584–591. (2013). Chondroitin sulfate proteoglycans potently inhibit invasion and serve as a central organizer of the brain tumor microenvironment. J. Neurosci. 33, Oudega, M. (2012). Molecular and cellular mechanisms underlying the role of 15603–15617. blood vessels in spinal cord injury and repair. Cell Tissue Res. 349, 269–288. Singer, A.J., and Clark, R.A. (1999). Cutaneous wound healing. N. Engl. J. Med. Overman, J.J., Clarkson, A.N., Wanner, I.B., Overman, W.T., Eckstein, I., 341, 738–746. Maguire, J.L., Dinov, I.D., Toga, A.W., and Carmichael, S.T. (2012). A role for ephrin-A5 in axonal sprouting, recovery, and activity-dependent plasticity after Sirko, S., Behrendt, G., Johansson, P.A., Tripathi, P., Costa, M., Bek, S., Hein- stroke. Proc. Natl. Acad. Sci. USA 109, E2230–E2239. rich, C., Tiedt, S., Colak, D., Dichgans, M., et al. (2013). Reactive glia in the injured brain acquire stem cell properties in response to sonic hedgehog. Pakulska, M.M., Ballios, B.G., and Shoichet, M.S. (2012). Injectable hydrogels [corrected]. Cell Stem Cell 12, 426–439. for central nervous system therapy. Biomed. Mater. 7, 024101. Snyder, E.Y., and Teng, Y.D. (2012). Stem cells and spinal cord repair. N. Engl. Perry, V.H. (2010). Contribution of systemic inflammation to chronic neurode- J. Med. 366, 1940–1942. generation. Acta Neuropathol. 120, 277–286. Soderblom, C., Luo, X., Blumenthal, E., Bray, E., Lyapichev, K., Ramos, J., Perry, V.H., Cunningham, C., and Holmes, C. (2007). Systemic infections and Krishnan, V., Lai-Hsu, C., Park, K.K., Tsoulfas, P., and Lee, J.K. (2013). Peri- inflammation affect chronic neurodegeneration. Nat. Rev. Immunol. 7, vascular fibroblasts form the fibrotic scar after contusive spinal cord injury. 161–167. J. Neurosci. 33, 13882–13887.

Piao, C.S., Stoica, B.A., Wu, J., Sabirzhanov, B., Zhao, Z., Cabatbat, R., Sofroniew, M.V. (2005). Reactive astrocytes in neural repair and protection. Loane, D.J., and Faden, A.I. (2013). Late exercise reduces neuroinflammation Neuroscientist 11, 400–407. and cognitive dysfunction after traumatic brain injury. Neurobiol. Dis. 54, 252–263. Sofroniew, M.V. (2009). Molecular dissection of reactive astrogliosis and glial scar formation. Trends Neurosci. 32, 638–647. Popescu, B.F., and Lucchinetti, C.F. (2012). Pathology of demyelinating diseases. Annu. Rev. Pathol. 7, 185–217. Sofroniew, M.V. (2013). Multiple roles for astrocytes as effectors of cytokines and inflammatory mediators. Neuroscientist. Published online October 8, Popovich, P.G., and Longbrake, E.E. (2008). Can the immune system be 2013. http://dx.doi.org/10.1177/1073858413504466. harnessed to repair the CNS? Nat. Rev. Neurosci. 9, 481–493. Sofroniew, M.V., and Vinters, H.V. (2010). Astrocytes: biology and pathology. Prokop, S., Miller, K.R., and Heppner, F.L. (2013). Microglia actions in Acta Neuropathol. 119, 7–35. Alzheimer’s disease. Acta Neuropathol. 126, 461–477. Song, B., Song, J., Zhang, S., Anderson, M.A., Ao, Y., Yang, C.Y., Deming, Radulovic, M., Yoon, H., Larson, N., Wu, J., Linbo, R., Burda, J.E., Diamandis, T.J., and Sofroniew, M.V. (2012). Sustained local delivery of bioactive nerve E.P., Blaber, S.I., Blaber, M., Fehlings, M.G., and Scarisbrick, I.A. (2013). growth factor in the central nervous system via tunable diblock copolypeptide Kallikrein cascades in traumatic spinal cord injury: in vitro evidence for roles hydrogel depots. Biomaterials 33, 9105–9116. in axonopathy and neuron degeneration. J. Neuropathol. Exp. Neurol. 72, 1072–1089. Srivastava, R., Aslam, M., Kalluri, S.R., Schirmer, L., Buck, D., Tackenberg, B., Rothhammer, V., Chan, A., Gold, R., Berthele, A., et al. (2012). Potassium Ransohoff, R.M., and Perry, V.H. (2009). Microglial physiology: unique stimuli, channel KIR4.1 as an immune target in multiple sclerosis. N. Engl. J. Med. specialized responses. Annu. Rev. Immunol. 27, 119–145. 367, 115–123.

Reilkoff, R.A., Bucala, R., and Herzog, E.L. (2011). Fibrocytes: emerging Stephan, A.H., Barres, B.A., and Stevens, B. (2012). The complement system: effector cells in chronic inflammation. Nat. Rev. Immunol. 11, 427–435. an unexpected role in synaptic pruning during development and disease. Annu. Rev. Neurosci. 35, 369–389. Rothstein, J.D. (2009). Current hypotheses for the underlying biology of amyo- trophic lateral sclerosis. Ann. Neurol. 65 (Suppl 1 ), S3–S9. Stevens, B., Allen, N.J., Vazquez, L.E., Howell, G.R., Christopherson, K.S., Nouri, N., Micheva, K.D., Mehalow, A.K., Huberman, A.D., Stafford, B., et al. Sahni, V., Mukhopadhyay, A., Tysseling, V., Hebert, A., Birch, D., Mcguire, (2007). The classical complement cascade mediates CNS synapse elimina- T.L., Stupp, S.I., and Kessler, J.A. (2010). BMPR1a and BMPR1b signaling tion. Cell 131, 1164–1178. exert opposing effects on gliosis after spinal cord injury. J. Neurosci. 30, 1839–1855. Sun, F., Lin, C.L., McTigue, D., Shan, X., Tovar, C.A., Bresnahan, J.C., and Beattie, M.S. (2010). Effects of axon degeneration on oligodendrocyte lineage Seo, J.H., Miyamoto, N., Hayakawa, K., Pham, L.D., Maki, T., Ayata, C., Kim, cells: dorsal rhizotomy evokes a repair response while axon degeneration K.W., Lo, E.H., and Arai, K. (2013). Oligodendrocyte precursors induce early rostral to spinal contusion induces both repair and apoptosis. Glia 58, 1304– blood-brain barrier opening after white matter injury. J. Clin. Invest. 123, 1319. 782–786. Suo, Z., Wu, M., Ameenuddin, S., Anderson, H.E., Zoloty, J.E., Citron, B.A., Shechter, R., and Schwartz, M. (2013). CNS sterile injury: just another wound Andrade-Gordon, P., and Festoff, B.W. (2002). Participation of protease-acti- healing? Trends Mol. Med. 19, 135–143. vated receptor-1 in thrombin-induced microglial activation. J. Neurochem. 80, 655–666. Shigetomi, E., Bowser, D.N., Sofroniew, M.V., and Khakh, B.S. (2008). Two forms of astrocyte calcium excitability have distinct effects on NMDA recep- Tao, J., Wu, H., Lin, Q., Wei, W., Lu, X.H., Cantle, J.P., Ao, Y., Olsen, R.W., tor-mediated slow inward currents in pyramidal neurons. J. Neurosci. 28, Yang, X.W., Mody, I., et al. (2011). Deletion of astroglial Dicer causes non- 6659–6663. cell-autonomous neuronal dysfunction and degeneration. J. Neurosci. 31, 8306–8319. Shirakawa, H., Sakimoto, S., Nakao, K., Sugishita, A., Konno, M., Iida, S., Kusano, A., Hashimoto, E., Nakagawa, T., and Kaneko, S. (2010). Transient Tong, X., Shigetomi, E., Looger, L.L., and Khakh, B.S. (2013). Genetically receptor potential canonical 3 (TRPC3) mediates thrombin-induced astrocyte encoded calcium indicators and astrocyte calcium microdomains. Neurosci- activation and upregulates its own expression in cortical astrocytes. entist 19, 274–291. J. Neurosci. 30, 13116–13129. Tuszynski, M.H., and Steward, O. (2012). Concepts and methods for the study Silver, J., and Miller, J.H. (2004). Regeneration beyond the glial scar. Nat. Rev. of axonal regeneration in the CNS. Neuron 74, 777–791. Neurosci. 5, 146–156. van den Brand, R., Heutschi, J., Barraud, Q., DiGiovanna, J., Bartholdi, K., Silver, J., Edwards, M.A., and Levitt, P. (1993). Immunocytochemical demon- Huerlimann, M., Friedli, L., Vollenweider, I., Moraud, E.M., Duis, S., et al. stration of early appearing astroglial structures that form boundaries and path- (2012). Restoring voluntary control of locomotion after paralyzing spinal cord ways along axon tracts in the fetal brain. J. Comp. Neurol. 328, 415–436. injury. Science 336, 1182–1185.

Neuron 81, January 22, 2014 ª2014 Elsevier Inc. 247 Neuron Review

Varki, A., and Gagneux, P. (2012). Multifarious roles of sialic acids in immunity. Yamanaka, K., Chun, S.J., Boillee, S., Fujimori-Tonou, N., Yamashita, H., Gut- Ann. N Y Acad. Sci. 1253, 16–36. mann, D.H., Takahashi, R., Misawa, H., and Cleveland, D.W. (2008). Astro- cytes as determinants of disease progression in inherited amyotrophic lateral Verkhratsky, A., Orkand, R.K., and Kettenmann, H. (1998). Glial calcium: sclerosis. Nat. Neurosci. 11, 251–253. homeostasis and signaling function. Physiol. Rev. 78, 99–141. Yang, C.Y., Song, B., Ao, Y., Nowak, A.P., Abelowitz, R.B., Korsak, R.A., Volterra, A., and Meldolesi, J. (2005). Astrocytes, from brain glue to communi- Havton, L.A., Deming, T.J., and Sofroniew, M.V. (2009). Biocompatibility of cation elements: the revolution continues. Nat. Rev. Neurosci. 6, 626–640. amphiphilic diblock copolypeptide hydrogels in the central nervous system. 30 Voskuhl, R.R., Peterson, R.S., Song, B., Ao, Y., Morales, L.B., Tiwari-Wood- Biomaterials , 2881–2898. ruff, S., and Sofroniew, M.V. (2009). Reactive astrocytes form scar-like peri- vascular barriers to leukocytes during adaptive immune inflammation of the Yoshioka, N., Hisanaga, S., and Kawano, H. (2010). Suppression of fibrotic CNS. J. Neurosci. 29, 11511–11522. scar formation promotes axonal regeneration without disturbing blood-brain barrier repair and withdrawal of leukocytes after traumatic brain injury. Walsh, J.T., and Kipnis, J. (2011). Regulatory T cells in CNS injury: the simple, J. Comp. Neurol. 518, 3867–3881. the complex and the confused. Trends Mol. Med. 17, 541–547. Zamanian, J.L., Xu, L., Foo, L.C., Nouri, N., Zhou, L., Giffard, R.G., and Barres, Wang, D., and Fawcett, J. (2012). The perineuronal net and the control of CNS B.A. (2012). Genomic analysis of reactive astrogliosis. J. Neurosci. 32, 6391– plasticity. Cell Tissue Res. 349, 147–160. 6410. Wanner, I.B., Anderson, M.A., Song, B., Levine, J., Fernandez, A., Gray- Thompson, Z., Ao, Y., and Sofroniew, M.V. (2013). Glial scar borders are Zhang, S., Anderson, M.A., Ao, Y., Khakh, B.S., Fan, J., Deming, T.J., and formed by newly proliferated, elongated astrocytes that interact to corral Sofroniew, M.V. (2014). Tunable diblock copolypeptide hydrogel depots for inflammatory and fibrotic cells via STAT3-dependent mechanisms after spinal local delivery of hydrophobic molecules in healthy and injured central nervous cord injury. J. Neurosci. 33, 12870–12886. system. Biomaterials 35, 1989–2000.

Watkins, S., and Sontheimer, H. (2012). Unique biology of gliomas: challenges Zheng, W., Watts, L.T., Holstein, D.M., Prajapati, S.I., Keller, C., Grass, E.H., 35 and opportunities. Trends Neurosci. , 546–556. Walter, C.A., and Lechleiter, J.D. (2010). Purinergic receptor stimulation reduces cytotoxic edema and brain infarcts in mouse induced by photothrom- Wetherington, J., Serrano, G., and Dingledine, R. (2008). Astrocytes in the bosis by energizing glial mitochondria. PLoS ONE 5, e14401. epileptic brain. Neuron 58, 168–178.

Williams, R.R., Henao, M., Pearse, D.D., and Bunge, M.B. (2013). Permissive Zhong, Z., Deane, R., Ali, Z., Parisi, M., Shapovalov, Y., O’Banion, M.K., Sto- Schwann Cell Graft/Spinal Cord Interfaces for Axon Regeneration. Cell Trans- janovic, K., Sagare, A., Boillee, S., Cleveland, D.W., and Zlokovic, B.V. (2008). plant. Published online October 22, 2013. http://dx.doi.org/10.3727/ ALS-causing SOD1 mutants generate vascular changes prior to motor neuron 096368913X674657. degeneration. Nat. Neurosci. 11, 420–422.

Winkler, E.A., Bell, R.D., and Zlokovic, B.V. (2011). Central nervous system Zlokovic, B.V. (2008). The blood-brain barrier in health and chronic neurode- pericytes in health and disease. Nat. Neurosci. 14, 1398–1405. generative disorders. Neuron 57, 178–201. Winkler, E.A., Sengillo, J.D., Sullivan, J.S., Henkel, J.S., Appel, S.H., and Zlokovic, B.V. (2011). Neurovascular pathways to neurodegeneration in Zlokovic, B.V. (2013). Blood-spinal cord barrier breakdown and pericyte 12 reductions in amyotrophic lateral sclerosis. Acta Neuropathol. 125, 111–120. Alzheimer’s disease and other disorders. Nat. Rev. Neurosci. , 723–738.

Wyss-Coray, T., Loike, J.D., Brionne, T.C., Lu, E., Anankov, R., Yan, F., Silver- Zukor, K., Belin, S., Wang, C., Keelan, N., Wang, X., and He, Z. (2013). Short stein, S.C., and Husemann, J. (2003). Adult mouse astrocytes degrade amy- hairpin RNA against PTEN enhances regenerative growth of corticospinal tract loid-beta in vitro and in situ. Nat. Med. 9, 453–457. axons after spinal cord injury. J. Neurosci. 33, 15350–15361.

248 Neuron 81, January 22, 2014 ª2014 Elsevier Inc.