Astrocytic CX43 hemichannels and gap junctions play a crucial role in development of chronic neuropathic pain following spinal cord injury

Chronic neuropathic pain is a frequent consequence of spinal cord injury (SCI). Yet despite recent advances, upstream releasing mechanisms and effective therapeutic options remain elusive. Previous studies have demonstrated that SCI results in excessive ATP release to the peritraumatic regions and that purinergic signaling, among glial cells, likely plays an essential role in facilitating inflammatory responses and nociceptive sensitization. We sought to assess the role of connexin 43 (Cx43) as a mediator of CNS inflammation and chronic pain. To determine the extent of Cx43 involvement in chronic pain, a weight‐drop SCI was performed on transgenic mice with Cx43/Cx30 deletions. SCI induced robust and persistent neuropathic pain including heat hyperalgesia and mechanical allodynia in wild‐type control mice, which developed after 4 weeks and was maintained after 8 weeks. Notably, SCI‐induced heat hyperalgesia and mechanical allodynia were prevented in transgenic mice with Cx43/Cx30 deletions, but fully developed in transgenic mice with only Cx30 deletion. SCI‐induced gliosis, detected as upregulation of glial fibrillary acidic protein in the spinal cord astrocytes at different stages of the injury, was also reduced in the knockout mice with Cx43/Cx30 deletions, when compared with littermate controls. In comparison, a standard regimen of post‐SCI treatment of minocycline attenuated neuropathic pain to a significantly lesser degree than Cx43 deletion. These findings suggest Cx43 is critically linked to the development of central neuropathic pain following acute SCI. Since Cx43/Cx30 is expressed by astrocytes, these findings also support an important role of astrocytes in the development of chronic pain. © 2012 Wiley Periodicals, Inc.     

Inhibition of Astrocyte Connexin 43 Channels Facilitates the Differentiation of Oligodendrocyte Precursor Cells Under Hypoxic Conditions In Vitro

Oligodendrocyte precursor cells (OPCs) proliferation and differentiation are essential for remyelination after white matter injury. Astrocytes could promote oligodendrogenesis after white matter damage whereas the underlying mechanisms are unknown. In this study, the role of astrocytic connexin43 (Cx43) hemichannels involved in OPC proliferation and differentiation in chronic hypoxia was evaluated. In an astrocyte-OPC co-culture chronic hypoxia model, OPCs became proliferative but failed to mature into oligodendrocytes. Application of astrocytic Cx43 blockers attenuated astrocyte activation, suppressed Cx43 hemichannel uptake activity and glutamate release induced by hypoxia, as well as improved OPC differentiation. Moreover, AMPA but not NMDA glutamate receptor antagonist rescued OPC differentiation in hypoxia. In conclusion, these findings suggested that astrocytic Cx43 hemichannel inhibition could potentially improve OPC maturation by attenuating AMPAR-mediated glutamate signaling. Astrocytic Cx43 hemichannels could serve as a potential therapeutic target for remyelination after chronic hypoxia.     

Blockade of interleukin-6 receptor suppresses reactive astrogliosis and ameliorates functional recovery in experimental spinal cord injury

Endogenous neural stem/progenitor cells (NSPCs) have recently been shown to differentiate exclusively into astrocytes, the cells that are involved in glial scar formation after spinal cord injury (SCI). The microenvironment of the spinal cord, especially the inflammatory cytokines that dramatically increase in the acute phase at the injury site, is considered to be an important cause of inhibitory mechanism of neuronal differentiation following SCI. Interleukin-6 (IL-6), which has been demonstrated to induce NSPCs to undergo astrocytic differentiation selectively through the JAK/STAT pathway in vitro, has also been demonstrated to play a critical role as a proinflammatory cytokine and to be associated with secondary tissue damage in SCI. In this study, we assessed the efficacy of rat anti-mouse IL-6 receptor monoclonal antibody (MR16-1) in the treatment of acute SCI in mice. Immediately after contusive SCI with a modified NYU impactor, mice were intraperitoneally injected with a single dose of MR16-1 (100 microg/g body weight), the lesions were assessed histologically, and the functional recovery was evaluated. MR16-1 not only suppressed the astrocytic diffentiation-promoting effect of IL-6 signaling in vitro but inhibited the development of astrogliosis after SCI in vivo. MR16-1 also decreased the number of invading inflammatory cells and the severity of connective tissue scar formation. In addition, we observed significant functional recovery in the mice treated with MR16-1 compared with control mice. These findings suggest that neutralization of IL-6 signaling in the acute phase of SCI represents an attractive option for the treatment of SCI.     

Tumor necrosis factor-mediated downregulation of spinal astrocytic connexin43 leads to increased glutamatergic neurotransmission and neuropathic pain in mice

Spinal cord astrocytes are critical in the maintenance of neuropathic pain. Connexin 43 (Cx43) expressed on spinal dorsal horn astrocytes modulates synaptic neurotransmission, but its role in nociceptive transduction has yet to be fully elaborated. In mice, Cx43 is mainly expressed in astrocytes, not neurons or microglia, in the spinal dorsal horn. Hind paw mechanical hypersensitivity was observed beginning 3 days after partial sciatic nerve ligation (PSNL), but a persistent downregulation of astrocytic Cx43 in ipsilateral lumbar spinal dorsal horn was not observed until 7 days post-PSNL, suggesting that Cx43 downregulation mediates the maintenance and not the initiation of nerve injury-induced hypersensitivity. Downregulation of Cx43 expression by intrathecal treatment with Cx43 siRNA also induced mechanical hypersensitivity. Conversely, restoring Cx43 by an adenovirus vector expressing Cx43 (Ad-Cx43) ameliorated PSNL-induced mechanical hypersensitivity. The sensitized state following PSNL is likely maintained by dysfunctional glutamatergic neurotransmission, as Cx43 siRNA-induced mechanical hypersensitivity was attenuated with intrathecal treatment of glutamate receptor antagonists MK801 and CNQX, but not neurokinin-1 receptor antagonist CP96345 or the Ca²⁺ channel subunit α₂δ₁ blocker gabapentin. The source of this dysfunctional glutamatergic neurotransmission is likely decreased clearance of glutamate from the synapse rather than increased glutamate release into the synapse. Astrocytic expression of glutamate transporter GLT-1, but not GLAST, and activity of glutamate transport were markedly decreased in mice intrathecally injected with Cx43-targeting siRNA but not non-targeting siRNA. Glutamate release from spinal synaptosomes prepared from mice treated with either Cx43-targeting siRNA or non-targeting siRNA was unchanged. Intrathecal injection of Ad-Cx43 in PSNL mice restored astrocytic GLT-1 expression. The cytokine tumor necrosis factor (TNF) has been implicated in the induction of central sensitization, particularly through its actions on astrocytes, in the spinal cord following peripheral injury. Intrathecal injection of TNF in naïve mice induced the downregulation of both Cx43 and GLT-1 in spinal dorsal horn, as well as hind paw mechanical hypersensitivity, as observed in PSNL mice. Conversely, intrathecal treatment of PSNL mice with the TNF inhibitor etanercept prevented not only mechanical hypersensitivity but also the downregulation of Cx43 and GLT-1 expression in astrocytes. The current findings indicate that spinal astrocytic Cx43 are essential for the maintenance of neuropathic pain following peripheral nerve injury and suggest modulation of Cx43 as a novel target for developing analgesics for neuropathic pain.     

Blocking connexin43 expression reduces inflammation and improves functional recovery after spinal cord injury

After traumatic CNS injury, a cascade of secondary events expands the initial lesion. The gap-junction protein connexin43 (Cx43), which is transiently up-regulated, has been implicated in the spread of 'bystander' damage. We have used an antisense oligodeoxynucleotide (asODN) to suppress Cx43 up-regulation in two rat models of spinal cord injury. Within 24 h of compression injury, rats treated with Cx43-asODN scored higher than sense-ODN and vehicle-treated controls on behavioural tests of locomotion. Their spinal cords showed less swelling and tissue disruption, less up-regulation of astrocytic GFAP, and less extravasation of fluorescently-labelled bovine serum albumin and neutrophils. The locomotor improvement was sustained over at least 4 weeks. Following partial spinal cord transection, Cx43-asODN treatment reduced GFAP immunoreactivity, neutrophil recruitment, and the activity of OX42(+) microglia in and around the lesion site. Cx43 has many potential roles in the pathophysiology of CNS injury and may be a valuable target for therapeutic intervention.     

Connexin43 and astrocytic gap junctions in the rat spinal cord after acute compression injury

To examine the possible role of interastrocytic gap junctions in the maintenance of tissue homeostasis after spinal cord damage, we initiated studies of the astrocytic gap junctional protein connexin43 (Cx43) in relation to temporal and spatial parameters of neuronal loss, reactive gliosis, and white matter survival in a rat model of traumatic spinal cord injury (SCI). Cx43 immunolocalization in normal and compression-injured spinal cord was compared by using two different sequence-specific anti-Cx43 antibodies that have previously exhibited different immunorecognition properties at lesion sites in brain. At 1- and 3-day survival times, gray matter areas with mild to moderate neuronal depletion exhibited a loss of immunolabeling with one of the two antibodies. At the lesion epicenter, these areas consisted of a zone that separated normal staining distal to the lesion from intensified labeling seen with both antibodies immediately adjacent to the lesion. Loss of immunoreactivity with only one of the two antibodies suggested masking of the corresponding Cx43 epitope. By 7 days post-SCI, Cx43 labeling was absent with both antibodies in all regions extending up to 1 mm from the lesion site. Reactive astrocytes displaying glial fibrillary acidic protein (GFAP) appeared by 1 day and were prominent by 3 days post-SCI. Their distribution in white and gray matter corresponded closely to that of Cx43 staining at 1 day, but less so at 3 days when GFAP-positive profiles were present at sites where Cx43 labeling was absent. By 7 days post-SCI, Cx43 again co-localized with GFAP-positive cells in the surviving subpial rim, and with astrocytic processes on radially oriented vascular profiles investing the central borders of the lesion. The results indicate that alterations in Cx43 cellular localization and Cx43 molecular modifications reflected by epitope masking, which were previously correlated with gap junction remodeling following excitotoxin-induced lesions in brain, are not responses limited to exogenously applied excitotoxins; they also occur in damaged spinal cord and are evoked by endogenous mechanisms after traumatic SCI. The GFAP/Cx43 co-localization results suggest that during their transformation to a reactive state, spinal cord astrocytes undergo a transitional phase marked by altered Cx43 localization or expression. J. Comp. Neurol. 382:199-214, 1997. © 1997 Wiley-Liss, Inc.     

Signaling pathways of ATP-induced PGE2 release in spinal cord astrocytes are EGFR transactivation-dependent

Traumatic spinal cord injury is characterized by an immediate, irreversible loss of tissue at the lesion site, as well as a secondary expansion of tissue damage over time. Although secondary injury should, in principle, be preventable, no effective treatment options currently exist for patients with acute spinal cord injury (SCI). Excessive release of ATP by the traumatized tissue, triggers the rapid release of arachidonic acid (AA) and prostaglandin E2 (PGE2), and has beenimplicated in acute and chronic neuropathic pain and inflammation. But the intracellular pathways between ATP and PGE2 remain largely unknown. We have explored the signaling events involved in this synthesis by primarily culturing spinal cord astrocytes: (1) we determined significant PGE2 production increased by ATP is mainly via Subtype 1 of P2 purinoceptors (P2Y1) but not P2Y2; (2) we found that ATP strongly increased the level of intracellular Ca(2+) via P2Y1 receptor; (3) we indicated that ATP stimulates the definitely release of AA and PGE2 which involved the transactivation of epidermal growth factor (EGF) receptor, the phosphorylation of extracellular-regulated protein kinases 1 and 2 (ERK(1/2) ) and the activation of cytosolic phospholipase A(2) (cPLA(2) ); (4) we examined ATP could increase the phosphorylation of Akt via P2Y1 receptor which also depend on the transactivation of EGFR, but the activation of Akt has no effect on the downstream of cPLA(2) phosphorylation. ATP induced by SCI could mobilize the release of AA and PGE2. And inhibition of PGE2 release reduces behavioral signs of pain after SCI and peripheral nerve injury.     

Current therapeutic strategies for inflammation following traumatic spinal cord injury

Damage from spinal cord injury occurs in two phases-the trauma of the initial mechanical insult and a secondary injury to nervous tissue spared by the primary insult.Apart from damage sustained as a result of direct trauma to the spinal cord,the post-traumatic inflammatory response contributes significantly to functional motor deficits exacerbated by the secondary injury.Attenuating the detrimental aspects of the inflammatory response is a promising strategy to potentially ameliorate the secondary injury,and promote significant functional recovery.This review details how the inflammatory component of secondary injury to the spinal cord can be treated currently and in the foreseeable future.     

Expression and detrimental role of hematopoietic prostaglandin D synthase in spinal cord contusion injury

Prostaglandin D(2) (PGD(2) ) is a potent inflammatory mediator, which is implicated in both the initiation and resolution of inflammation in peripheral non-neural tissues. Its role in the central nervous system has not been fully elucidated. Spinal cord injury (SCI) is associated with an acute inflammatory response, which contributes to secondary tissue damage that worsens functional loss. We show here, with the use of hematopoietic prostaglandin D synthase (HPGDS) deficient mice and a HPGDS selective inhibitor (HQL-79), that PGD(2) plays a detrimental role after SCI. We also show that HPGDS is expressed in macrophages in the injured mouse spinal cord and contributes to the increase in PGD(2) in the contused spinal cord. HPGDS(-/-) mice also show reduced secondary tissue damage and reduced expression of the proinflammatory chemokine CXCL10 as well as an increase in IL-6 and TGFβ-1 expression in the injured spinal cord. This was accompanied by a reduction in the expression of the microglia/macrophage activation marker Mac-2 and an increase in the antioxidant metallothionein III. Importantly, HPGDS deficient mice exhibit significantly better locomotor recovery after spinal cord contusion injury than wild-type (Wt) mice. In addition, systemically administered HPGDS inhibitor (HQL-79) also enhanced locomotor recovery after SCI in Wt mice. These data suggest that PGD(2) generated via HPGDS has detrimental effects after SCI and that blocking the activity of this enzyme can be beneficial.     

Downregulation of spinal astrocytic connexin43 leads to upregulation of interleukin‐6 and cyclooxygenase‐2 and mechanical hypersensitivity in mice

Connexin43 (Cx43), involved in intercellular signaling, is expressed in spinal dorsal horn astrocytes and crucial in the maintenance of neuropathic pain. Downregulation of spinal astrocytic Cx43 in mice enhances glutamatergic neurotransmission by decreasing glutamate transporter GLT‐1 expression, resulting in cutaneous hypersensitivity. Decreased expression of astrocytic Cx43 could lead to altered expression of other nociceptive molecules. Transfection of Cx43‐targeting siRNA in cultured spinal astrocytes increased expression of the pronociceptive cytokine interleukin‐6 (IL‐6) and the prostaglandin synthesizing enzyme cyclooxygenase‐2 (COX‐2). Increased expression of IL‐6 and COX‐2 was due to decreased Cx43 expression rather than due to diminished Cx43 channel function. In mice, downregulation of spinal Cx43 expression by intrathecal treatment with Cx43‐targeting siRNA increased IL‐6 and COX‐2 expression and induced hind paw mechanical hypersensitivity. Cx43 siRNA‐induced mechanical hypersensitivity was attenuated by intrathecal treatment with anti‐IL‐6 neutralizing antibody and intraperitoneal treatment of selective COX‐2 inhibitor celecoxib, demonstrating that these molecules play a role in nociceptive processing following Cx43 downregulation. Restoring spinal Cx43 by intrathecal injection of an adenovirus vector expressing Cx43 in mice with a partial sciatic nerve ligation reduced spinal IL‐6 and COX‐2 expression. Suppression of glycogen synthase kinase‐3β (GSK‐3β), a serine/threonine protein kinase, prevented upregulation of IL‐6 and COX‐2 expression induced by Cx43 downregulation in both cultured astrocytes and in mouse spinal dorsal horn. Inhibition of spinal GSK‐3β also ameliorated Cx43 siRNA‐induced mechanical hypersensitivity. The current findings indicate that downregulation of spinal astrocytic Cx43 leads to changes in spinal expression of pronociceptive molecules underlying the maintenance of pain following nerve injury.     

Role of connexin 32 hemichannels in the release of ATP from peripheral nerves

Extracellular purines elicit strong signals in the nervous system. Adenosine‐5′‐triphosphate (ATP) does not spontaneously cross the plasma membrane, and nervous cells secrete ATP by exocytosis or through plasma membrane proteins such as connexin hemichannels. Using a combination of imaging, luminescence and electrophysiological techniques, we explored the possibility that Connexin 32 (Cx32), expressed in Schwann cells (SCs) myelinating the peripheral nervous system could be an important source of ATP in peripheral nerves. We triggered the release of ATP in vivo from mice sciatic nerves by electrical stimulation and from cultured SCs by high extracellular potassium concentration‐evoked depolarization. No ATP was detected in the extracellular media after treatment of the sciatic nerve with Octanol or Carbenoxolone, and ATP release was significantly inhibited after silencing Cx32 from SCs cultures. We investigated the permeability of Cx32 to ATP by expressing Cx32 hemichannels in Xenopus laevis oocytes. We found that ATP release is coupled to the inward tail current generated after the activation of Cx32 hemichannels by depolarization pulses, and it is sensitive to low extracellular calcium concentrations. Moreover, we found altered ATP release in mutated Cx32 hemichannels related to the X‐linked form of Charcot‐Marie‐Tooth disease, suggesting that purinergic‐mediated signaling in peripheral nerves could underlie the physiopathology of this neuropathy. GLIA 2013;61:1976–1989     

Lipocalin 2 plays an immunomodulatory role and has detrimental effects after spinal cord injury

Lipocalin 2 (Lcn2) plays an important role in defense against bacterial infection by interfering with bacterial iron acquisition. Although Lcn2 is expressed in a number of aseptic inflammatory conditions, its role in these conditions remains unclear. We examined the expression and role of Lcn2 after spinal cord injury (SCI) in adult mice by using a contusion injury model. Lcn2 expression at the protein level is rapidly increased 12-fold at 1 d after SCI and decreases gradually thereafter, being three times as high as control levels at 21 d after injury. Lcn2 expression is strongly induced after contusion injury in astrocytes, neurons, and neutrophils. The Lcn2 receptor (Lcn2R), which has been shown to influence cell survival, is also expressed after SCI in the same cell types. Lcn2-deficient (Lcn2?/?) mice showed significantly better locomotor recovery after spinal cord contusion injury than wild-type (Lcn2?/?) mice. Histological assessments indicate improved neuronal and tissue survival and greater sparing of myelin in Lcn2?/? mice after contusion injury. Flow cytometry showed a decrease in neutrophil influx and a small increase in the monocyte population in Lcn2?/? injured spinal cords. This change was accompanied by a reduction in the expression of several pro-inflammatory chemokines and cytokines as well as inducible nitric oxide synthase early after SCI in Lcn2?/? mice compared with wild-type animals. Our results, therefore, suggest a role for Lcn2 in regulating inflammation in the injured spinal cord and that lack of Lcn2 reduces secondary damage and improves locomotor recovery after spinal cord contusion injury.     

Neuroprotective effects of nitidine against traumatic CNS injury via inhibiting microglia activation

Glial cell response to injury has been well documented in the pathogenesis after traumatic brain injury (TBI) and spinal cord injury (SCI). Although microglia, the resident macrophages in the central nervous system (CNS), are responsible for clearing debris and toxic substances, excessive activation of these cells will lead to exacerbated secondary damage by releasing a variety of inflammatory and cytotoxic mediators and ultimately influence the subsequent repair after CNS injury. In fact, inhibition of microgliosis represents a therapeutic strategy for CNS trauma. We here showed that nitidine, a benzophenanthridine alkaloid, restricted reactive microgliosis and promoted CNS repair after traumatic injury. Nitidine was shown to prevent cultured microglia from LPS-induced reactive activation by regulation of ERK and NF-κB signaling pathway. Furthermore, the nitidine-mediated inhibition of microgliosis was also shown in injured brain and spinal cord, which significantly increased neuronal survival and decreased neural tissue damage after injury. Importantly, behavioral analysis revealed that nitidine-treated mice with SCI had improved functional recovery as assessed by Basso Mouse Scale and swimming test. Together, these findings indicated that nitidine increased CNS tissue sparing and improved functional recovery by attenuating reactive microgliosis, suggestive of the potential therapeutic benefit for CNS injury.     

Severity-dependent expression of pro-inflammatory cytokines in traumatic spinal cord injury in the rat.

The post-traumatic inflammatory response in acute spinal cord contusion injury was studied in the rat. Mild and severe spinal cord injury (SCI) was produced by dropping a 10 g weight from 3 and 12 cm at the T12 vertebral level. Increased immunoreactivity of TNF-alpha in mild and severe SCI was detected in neurons at 1 h post-injury, and in neurons and microglia at 6 h post-injury, with a less significant increase in mild SCI. Expression was short-lived and declined sharply by 1 d post-injury. RT-PCR showed an early significant up-regulation of IL-1 beta, IL-6 and TNF-alpha mRNAs, maximal at 6 h post-injury with return to control levels by 24 h post-injury, the changes being less statistically significantly in mild SCI. Western blot showed early transient increases of IL-1 beta, IL-6 and TNF-alpha proteins in severe SCI but not mild SCI. Immunocytochemical, western blotting and RT-PCR analyses suggest that endogenous cells (neurons and microglia) in the spinal cord, not blood-borne leucocytes, contribute to IL-1 beta, IL-6 and TNF-alpha production in the post-traumatic inflammatory response and that their up-regulation is greater in severe than mild SCI.     

Decreased connexin 43 in astrocytes inhibits the neuroinflammatory reaction in an acute mouse model of neonatal sepsis

Neonatal sepsis is common in neonatal intensive care units,often complicated by injury to the immature brain. Previous studies have shown that the expression of the gap junction protein connexin 43(Cx43) in the brain decreases when stimulated by neuro-infl ammatory drugs such as lipopolysaccharide(LPS). Here we showed that partial deletion of Cx43 in astrocytes resulted in weakened inflammatory responses. The up-regulation of pro-inflammatory cytokines was significantly reduced in mice with partial deletion of Cx43 in astrocytes compared with wild-type littermates after systemic LPS injection. Moreover,microglial activation was inhibited in mice with partial deletion of Cx43. These results showed that Cx43 in astrocytes plays a critical role in neuro-infl ammatory responses. This work provides a potential therapeutic target for inhibiting neuroinfl ammatory responses in neonatal sepsis.     

Pretreatment with NMDA receptor antagonist MK801 improves neurophysiological outcome after an acute spinal cord injury

Abstract

The post-traumatic release of excitatory amino acids (EAA) and their actions on N-methyl-D-aspartate (NMDA) receptors plays a major role in the spinal cord secondary injury process. The neuronal damage caused by the release of EAA may be reduced by NMDA-receptor channel blockers. To investigate the involvement of NMDA receptors in spinal cord injury (SCI), we pretreated animals with the noncompetitive NMDA antagonist MK801 (1.0 mg kg~before a compressive acute SCI. Pretreated animals with MK801 significantly (p = 0.038) improved the recovery of function as measured by evoked potential activities. Morphologically, specimens from rats treated with MK801 were characterized by milder and more localized hemorrhage in the gray matter. Immunohistochemical staining for glial fibrillary acidic protein (GFAP) and neurofilament (NF) histochemistry showed leakage of these antigens in traumatized cord while characteristic staining of astrocytes and neurons and their processes was observed in morphologically preserved tissue. The loss of NF immunoreactivity was reduced by MK801 treatment. [Neurol Res 1996; 18: 509-515]     

Intercellular communication in spinal cord astrocytes: fine tuning between gap junctions and P2 nucleotide receptors in calcium wave propagation.

Electrophysiological properties of gap junction channels and mechanisms involved in the propagation of intercellular calcium waves were studied in cultured spinal cord astrocytes from sibling wild-type (WT) and connexin43 (Cx43) knock-out (KO) mice. Comparison of the strength of coupling between pairs of WT and Cx43 KO spinal cord astrocytes indicates that two-thirds of total coupling is attributable to channels formed by Cx43, with other connexins contributing the remaining one-third of junctional conductance. Although such a difference in junctional conductance was expected to result in the reduced diffusion of signaling molecules through the Cx43 KO spinal cord syncytium, intercellular calcium waves were found to propagate with the same velocity and amplitude and to the same number of cells as between WT astrocytes. Measurements of calcium wave propagation in the presence of purinoceptor blockers indicate that calcium waves in Cx43 KO spinal cord astrocytes are mediated primarily by extracellular diffusion of ATP; measurements of responses to purinoceptor agonists revealed that the functional P2Y receptor subtype is shifted in the Cx43 KO astrocytes, with a markedly potentiated response to ATP and UTP. Thus, the reduction in gap junctional communication in Cx43 KO astrocytes leads to an increase in autocrine communication, which is a consequence of a functional switch in the P2Y nucleotide receptor subtype. Intercellular communication via calcium waves therefore is sustained in Cx43 null mice by a finely tuned interaction between gap junction-dependent and independent mechanisms.     

Inhibition of monocyte/macrophage migration to a spinal cord injury site by an antibody to the integrin alphaD: a potential new anti-inflammatory treatment

The inflammatory response that ensues during the initial 48 to 72 h after spinal cord injury causes considerable secondary damage to neurons and glia. Infiltration of proinflammatory-activated neutrophils and monocytes/macrophages into the cord contributes to spinal cord injury-associated secondary damage. beta2 integrins play an essential role in leukocyte trafficking and activation and arbitrate cell-cell interactions during inflammation. The beta2 integrin, alphaDbeta2, is expressed on monocytes/macrophages and neutrophils and binds to vascular adhesion molecule-1 (VCAM-1). The increased expression of VCAM-1 during central nervous system (CNS) inflammation likely contributes to leukocyte extravasation into the CNS. Accordingly, blocking the interaction between alphaDbeta2 and VCAM-1 may attenuate the inflammatory response at the SCI site. We investigated whether the administration of monoclonal antibodies (mAbs) specific for the rat alphaD subunit would reduce the inflammatory response after a spinal cord transection injury in rats. At a 1 mg/kg dose two of three anti-alphaD mAbs caused a significant ( approximately 65%) reduction in the number of macrophages at the injury site and one anti-alphaD mAb led to a approximately 43% reduction in the number of neutrophils at the SCI site. Thus, our results support the concept that the alphaDbeta2 integrins play an important role in the trafficking of leukocytes to a site of central nervous system inflammation. This study also offers preliminary evidence that anti-alphaD mAbs can reduce the extravasation of macrophages and, to a lesser extent, neutrophils, to the SCI site.     

Cell Cycle Activation and Spinal Cord Injury

Traumatic spinal cord injury (SCI) evokes a complex cascade of events with initial mechanical damage leading to secondary injury processes that contribute to further tissue loss and functional impairment. Growing evidence suggests that the cell cycle is activated following SCI. Up-regulation of cell cycle proteins after injury appears to contribute not only to apoptotic cell death of postmitotic cells, including neurons and oligodendrocytes, but also to post-traumatic gliosis and microglial activation. Inhibition of key cell cycle regulatory pathways reduces injury-induced cell death, as well as microglial and astroglial proliferation both in vitro and in vivo. Treatment with cell cycle inhibitors in rodent SCI models prevents neuronal cell death and reduces inflammation, as well as the surrounding glial scar, resulting in markedly reduced lesion volumes and improved motor recovery. Here we review the effects of SCI on cell cycle pathways, as well as the therapeutic potential and mechanism of action of cell cycle inhibitors for this disorder.