By: Orest Kayder
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Mechanical trauma to the spinal cord leads to the physical disruption of the neuro-glial interactions and the vasculature surrounding the tissue. A multidimensional set of interrelated pathophysiological responses is then triggered which include oxidative stress, excitotoxicity and reactive astrogliosis. The former is an imbalance between the production and degradation of reactive oxygen species resulting in lipid peroxidation, malfunction of enzymatic proteins, and DNA fragmentation(1). The changes in the morphology and molecular expression of astrocytes are commonly referred to as reactive astrogliosis. Furthermore, the abnormal proliferation of the astrocytes leads to the formation of glial scar which impedes the regrowth of injured axons(2). Inflammation of the area contiguous to the site of the injury is primarily due to the activation of the microglia. Additionally, the disruption of the blood-brain barrier with the successive break down, release chemokines and other inflammatory mediators favouring the recruitment of inflammatory cells including neutrophils, lymphocytes and macrophages(3). Macrophages and neutrophils produce proteolytic enzymes, matrix metalloproteinases, reactive oxygen and nitrogen species contributing to degradation of extracellular matrix and oxidative stress(4). Moreover, the accumulation of the pro-inflammatory cytokines induces other intracellular mechanisms leading to demyelination, both neural and glial apoptosis resulting in a neurological deficit(5). Due to the space limitations which prevent comprehensive assessment of all the topics, only principal mechanisms underlying inflammatory processes would be unravelled.


Inflammation


Inflammation plays a fundamental role in regulating the pathogenesis of spinal cord injury. Understanding the molecular framework following SCI can provide solid ground for the development of new therapeutic strategies, whose effectiveness in human clinical trials has remained intangible (6,7). Ultimately, the focus of the neurowiki is to provide the analysis of the spatio-temporal expression of cells invading the site of the injury, as well as the cytokines and interleukins associated with posttraumatic inflammation.



Neutrophils


Following a spinal cord trauma, neutrophils are the first cells to initiate an inflammatory response, and they are recruited to the site of the injury by chemotaxis with the aid of cytokine-derived neutrophil-chemoattractant (CINC) (8) (fig 1).
Temporal_histopathological_changes_SCI.jpg
Fig. 1. Illustrations showing the histopathological changes at 8 h (B), 1 day (C), 3 days (D), and 5 days (E) after spinal cord contusion compared with control group (A). Amount of haemorrhage (B), necrotic components (C), inflammatory cells (D), and cyst formation (E) appeared in the contused spinal cord (area between arrows). Scale bar 0.5 mm applies to A-E.(figure taken from Yan, X., et al. (2010) Proteomic profiling of proteins in rat spinal cord injury induced by contusion injury. Neurochem Internat, 56, 971-983.) (click to enlarge)
Neutrophils are major exogeneous sources of reactive oxygen and nitrogen species, contributing to oxidative stress (9). n addition, neutrophils play a central role in the generation of matrix metalloproteinases whose function is to break down the extracellular matrix. The proinflammatory effects associated with the zinc-dependent endopepetidases, in particular, gelatinase B (MMP-9), are primarily due to disruption of vascular endothelium contributing to increased permeability of spinal cord barrier, posttraumatic edema and haemorrhage (10). Given recent findings, the functional recovery following SCI has been improved in the models with the early pharmacological inhibition using valproic acid of MMP-2 and MMP-9 (11, 12). On the contrary, it has been established that MMP-9 regulate the cell proliferation and maturation of multipotent NG2+ progenitor cells into oligodendrocytes improving myelin neuropathology in the hemisected spinal cord (13).


Administration of methothrexate, leads to reduction of early neutrophil infiltration and the concominant lipid peroxidation, displaying neuroprotective properties associated with the protection of axons by minimizing the ultrastructural changes in the SCI (14). In terms of the secretory activity, the neutrophils have been shown to be the major producers of interleukin (IL)-4, an anti-inflammatory cytokine, highly expressed in rats at 24hrs after contusive SCI (15). The neutralization of IL-4 with an antibody leads to macrophage activation, suggesting a possible mechanism of how neutrophils regulate the number of other infiltrating cells thus confining the size of the cavity in the post-traumatic spinal cord (15).


Microglia



Microglia are derived from myeloid progenitors in the yolk sac and are the resident macrophages of the central nervous system (CNS) which supervise the extracellular parenchyma, and exist in a quiescent state in the healthy CNS (16). The invasion of microglia occurs primarily at 3 to 7 days postinjury in Sprague-Dawley and Lewis rats induced by displacement of the cord at T8 level (17). They display a functional behaviour typical for macrophages such as phagocytosis associated with the clearance of debris due to necrosis, and produce various factors and proteases which are cytotoxic to the neuronal tissue (18). Further studies have suggested that microglia become activated following toll-like receptor (TLR) engagement and express proinflammatory cytokines such as tumour necrosis factor alpha (TNFα) and interleukins (IL)-6, IL-1 (19). The main consequence associated with TNFα, which is released and increase 6-12hrs post-injury with peak at 4 days (20), is the potentiation of excitotoxic effects of glutamatergic afferents by pathological activation of non-NMDA receptors (19). This view is supported by observation that IL-10, which blocks the injury induced increase in the TNFα (21), and a tumor necrosis factor alpha antagonist (etanercept) reduce apoptosis of neurons and oligodendroglia improving hindlimb activity and facilitating myelin regeneration (22).


Lymphocytes



The physical insult to the spinal cord, results in introduction of infection to the site of the injury inducing the lymphocytic infiltration. Lymphocytes are the type of white blood cells which mediate innate, natural killer (NK) cells, and adaptive immune response, B and T cells, in the vertebrates. Since lymphocytes are not residents of CNS, the degree of infiltration is closely associated with the vascular permeability (23). Recent mice models suggest that functional recovery following SCI is improved in mice lacking B cells (24).

One of the major regulators of the immune system which works through activation of T cells is IL-1 (25). Interleukin-1 is a proinflammatory cytokine which mediates a diverse range of effects and its expression at the protein level has been detected to rise in microglia cells 5 hours following SCI in humans (26). The significance of IL-1 has been clearly demonstrated with IL-1 receptor antagonist proteins which lead to decreased inflammation, aggravation in neurofilament proteins and brain-derived neurotrophic factor expression, ultimately enhancing neuroglia survival following SCI (27). Although, the exact role that T cells play in remyelination following spinal cord injury has not been established yet, the evidence suggests that T cells take part in demyelination processes associated with multiple sclerosis (28), so potentially could be involved in the same process in the pathogenesis of SCI.


Macrophages


Hematopoiesis_(human)_diagram.png
Fig. 2. Diagram showing the development of different blood cells from haematopoietic stem cell to mature cells (figure adopted from Robbins et al., Pathologic Basis of disease. 7th ed. Chapter 13, Red blood cell and bleeding disorders, page 621: figure 13-1.)

Macrophages are cells produced by differentiation of monocytes, which arises from monoblast precursors in the bone marrow (fig. 2) and are dependent on the circulating levels of granulocyte-macrophage colony stimulating factor (GM-CSF) (29). Macrophages are multidimensional cells which mediate an array of cellular processes as removal of dead cell material during chronic inflammation, regulation of the immune response through the release of a type of cytokines (monokines) which attract the neutrophils by chemotaxis. One of the key regulators associated with role of macrophages in inflammation is migration inhibitory factor (MIF). Although it could be produced by other cells including microglia, neurons and astrocytes (30), MIF is a multipotent pro-inflammatory cytokine and a glucocorticoid-induced immunomodulator (30). The levels of MIF increase post-injury (31) and the attenuation of expression reduces the neuronal death and promotes functional recovery (32).

Recent in vivo studies provide compelling evidence that there are two different phenotypes of macrophages (M1 and M2), the activation of which is dependent on different intracellular cascades, referred to as macrophage polarization, resulting in cells with either pro- or anti-inflammatory capabilities. The abolishment of M1 macrophages with temporal blockade of IL-6 signalling attenuates inflammatory activity and promotes development of alternatively activated M2 cells contributing to functional recovery (33). In addition, the inhibition of IL-6 signalling pathway shifts the emphasis in terms of the major pro-inflammatory cell to microglia, which has been shown to have greater phagocytic ability against myelin debris following SCI (34).


Blood - spinal cord barrier alterations



One of the critical components which determines the degree of an inflammatory response following a spinal cord injury is the permeability of blood-spinal cord barrier (BSCB). During the early post-injury stages, the permeability is defined primarily due to the physical disruption of the BSCB by the mechanical forces which induced the SCI. Alternatively, with the progression of time the degree of microvascular permeability becomes more dependent on the cytokines and other molecular mediators associated with the intraparenchymal inflammation (35). In addition to regulation the activity and migration patterns of inflammatory and immune cells, cytokines and chemokines modify the expression profile of cyclooxygenase (COX) enzymes (36). The aggravated levels of COX enzymes produce excessive amounts of prostanoids leading to alterations of vascular permeability causing edema, and infiltration of CNS non-resident leukocytes (37). As noted earlier, the matrix metalloproineases (9), TNFα (20), IL-1 (26), and other vasoactive agents as histamines (38), kinins (39) and elastase (7) may favour diapedesis. Ultimately, the loss of the integrity within BSCB compromises the functionality of neuroglia with the subsequent loss of neurological function.


Conclusive remarks



Despite a great number of experimental data the exact mechanism and effect associated with the inflammatory process following SCI has not been elucidated. One of the key factors is the cross-talk between inflammatory mediators, which do influence the intraparenchymal processes. The other caveat is the variation in the outcomes due to genetic differences of various models. The same problem translates onto the clinical practice, when diverse outcomes are established due to the genetic variations amid patients. Better understanding of the neurobiological processes associated with the acute spinal cord injury will permit the development of better therapeutic strategies.





References
  1. Jia, Z., Zhu, H., Wang, X., Misra, H., and Li, Y. (2011) Oxidative stress in spinal cord injury and antioxidant-based intervention. Spinal Cord, 10: 1038-1045.
  2. Sofroniew, M. V. (2009) Molecular dissection of reactive astrogliosis and glial scar formation. Trends Neurosci., 32 (12): 638-647.
  3. Pineau, S., Sun, L., Bastien, D. and Lacroix, S. (2010) Astrocytes initiate inflammation in the injured mouse spinal cord by promoting the entry of neutrophils and inflammatory monocytes in an IL-1 receptor/MyD88-dependent fashion. Brain, Behavior, and Immunity., 24: 540-553.
  4. Noble, L. J., Donovan, F., and Igarashi, T. (2002) Matrix metalloproteinases limit functional recovery after spinal cord injury by modulation of early vascular event. J Neurosci, 22: 7526-7535.
  5. Liu, H., and Shubayev, V. I. (2011) Matrix metalloproteinase-9 controls proliferation of NG2+ progenitor cells immediately after spinal cord injury. Exp Neurol, 231 (2): 236-246.
  6. Fehlings, M.G., and Baptiste, D. C. (2005) Current status of clinical trials of acute spinal cord injury. Injury, 36 (2): B113-B122.
  7. Hawryluk, G. W., Rowland, J., Kwon, B. K., and Fehlings, M. G. (2008) Protection and repair of the injured spinal cord: a review of completed, ongoing, and planned clinical trials for acute spinal cord injury. Neurosurg Focus, 25: E14.
  8. Tonai, T., Shiba, K., Taketani, Y., Ohmoto, Y., Murata, K., Muraguchi, M., Ohsaki, H., Takeda, E., and Nishisho, T. (2001) A neutrophil elastase inhibitor (ONO-5046) reduces neurologic damage after spinal cord injury in rats. J Neurochem, 78 (5): 1064-1072.
  9. Bao, F., Omana, V., Brown, A., and Weaver, L. C. (2012) The systemic inflammatory response after spinal cord injury in the rat is decreased by α4β1 integrin blockade. J Neurotr , epub ahead of print.
  10. Noble, L. J., Donovan, F., and Igarashi, T. (2002) Matrix metalloproteinases limit functional recovery after spinal cord injury by modulation of early vascular event. J Neurosci, 22: 7526-7535.
  11. Zhang, H., Chang, M., Hansen, C. N., Basso, D. M., and Noble-Haesslein, L. J. (2011) Role of matrix metalloproteinases and therapeutic benefits of the their inhibition in spinal cord injury. Neurotherap, 8 (2): 206-220.
  12. Lee, J.Y, Kim, H. S., Choi, H. Y., Oh, T. H., Ju, B. G., and Yune, T. Y. (2012) Valproic acid attenuates blood-spinal cord barrier disruption by inhibition matrix metalloprotease-9 activity and improves functional recovery after spinal cord injury. J Neurochem, 10: 1471-4159.
  13. Liu, H., and Shubayev, V. I. (2011). Matrix metalloproteinase-9 controls proliferation of NG2+ progenitor cells immediately after spinal cord injury. Exp Neurol, 231 (2): 236-246.
  14. Sanli, A. M., Serbes, G., Sargon, M. F., Caliskan, M., Kilinc, K., Bulut, H., and Sekerci, Z. (2012) Methothrexate attenuates early neutrophil infiltration and the associated lipid peroxidation in the injured spinal cord but does not induce neurotoxicity in the uninjured spinal cord in rats. Acta Neurochir, (epub ahead of print).
  15. Lee, S. I., Jeong, S. R., Kang, Y. M., Han, D. H., Jin, B. K., Namgung, U., and Kim, B. G. (2010) Endogeneous expression of inteleukin-4 regulates macrophage activation and confines cavity formation after traumatic spinal cord injury. J Neurosci Res, 88 (11): 2409-2419.
  16. Ginhoux, F., Greter, M., Leboeuf, M., Nandi, S., See, P., Gokhan, S., Mehler, M. F., Conway, S. J., and Merad, M. (2010) Fate mapping analysis reveals that adult microglia derive from primitive macrophages. Science, 330 (6005): 841-845.
  17. Popovich, P. G., Wei, P., and Stokes, B. T. (1997) Cellular inflammatory response after spinal cord injury in Sprague-Dawley and Lewis rats. J Comp Neurol, 377: 443-464.
  18. Pan J. Z., Ni, L., Sodhi, A., Aquanno, A., Young, W., and Hart, R. P. (2002) Cytokine activity contributes to induction of inflammatory cytokine mRNAs in spinal cord following contusion. J Neurosci Res, 68: 315-322.
  19. Kigerl, K. A., Lai, W., Rivest, S., Hart, R. P., Satoskar, A. R., and Popovich, P. G. (2007) Toll-like receptor (TLR)-2 and TLR-4 regulate inflammation, gliosis, and myelin sparing after spinal cord injury. J Neurochem, 102 (1): 37-50.
  20. Sobani, Z. A., Quadri, S. A., and Enam, S. A. (2010) Stem cells for spinal cord regeneration: Current status. Surg Neurol Int, 1: 93.
  21. Bethea, J. R., Nagashima, H., Acosta, M. C., Briceno, C., Gomez, F., Marcillo, A. E., Loor, K., Green, J., and Dietrich, W. D. (1999) Systematically administered interleukin-10 reduces tumor necrosis factor-alpha production and significantly improves functional recovery following traumatic spinal cord injury in rats. J Neurotraum, 16: 851-863.
  22. Chen, K. B., Uchida, K., Nakajima, H., Yayama, T., Hirai, T., Watanabe, S., Guerrero, A. R., Kobayashi, S., Ma, W. Y., Liu, S. Y., and Baba, H. (2011) Tumor necrosis factor-α reduces apoptosis of neurons and oligodendroglia in rat spinal cord injury. Spine, 36 (17): 1350-1358.
  23. Vanegas, H., and Schaible, H. G. (2001) Prostaglandins and cyclooxygenases in the spinal cord. Prog Neurobiol, 64: 327-363.
  24. Ankeny, D. P., Guan, Z., and Popovich, P. G. (2009) B cells produce pathogenic antibodies and impair recovery after spinal cord injury in mice. J Clin Invest, 119 (10): 2990-2999.
  25. Allan, S. M., Tyrrell, P. J., and Rothwell, N. J. (2005) Interleukin-1 and neuronal injury. Nat Rev Immunol, 5 (8): 629-640.
  26. Yang, L., Blumbergs, P. C., Jones, N. R., Manavis, J., Sarvestani, G. T., and Ghabriel, M. N. (2004) Early expression and cellular localization of proinflammatory cytokines inteleukin-1, interleukin-6, and tumor necrosis factor-alpha in human traumatic spinal cord injury. Spine, 29 (9): 966-971.
  27. Zong, S., Zeng, G., Wei, B., Xiong, C., and Zhao, Y. (2012) Beneficial effect of interleukin-1 receptor antagonist protein on spinal cord injury recovery in the rat. Inflamm, 35 (2): 520-526.
  28. Weber, M. S., and Hemmer, B. (2010) Cooperation of B cells and T cells in the pathogenesis of multiple sclerosis. Results Probl Cell Differ, 51: 115-126.
  29. Ha, Y., Kim, Y. S., Cho, J. M., Yoon, S. H., Park, S. R., Yoon, D. H., Kim, E. Y., and Park, H. C. (2005) Role of granulocyte-macrophage colony-stimulating factor in preventing apoptosis and improving functional outcome in experimental spinal cord contusion injury. J Neurosurg Spine, 2 (1): 55-61.
  30. Calandra, T., and Bucala, R. (1995) Macrophage migration inhibitory factor: a counter-regulator of glucocorticoid action and critical mediator of septic shock. J Inflamm, 37: 39-51.
  31. Koda, M., Nishio, Y., Hashimoto, M., Kamada, T., Koshizuka, S., Yoshinaga, K., Onodera, S., Nishihira, J., Moriya, H., and Yamazaki, M. (2004) Up-regulation of macrophage-inhibitory factor expression after compression-induced spinal cord injury in rats. Acta Neuropathol, 108: 31-36.
  32. Nishio, Y., Koda, M., Hashimoto, M., Kamada, T., Koshizuka, S., Yoshinaga, K., Onodera, S., Nishihira, J., Okawa, A., and Yamazaki, M. (2009) Deletion of macrophage migration inhibitory factor attenuates neuronal death and promotes functional recovery after compression-induced spinal cord injury in mice. Acta Neuropathol, 117 (3): 321-328.
  33. Guerrero, A. R., Uchida, K., Nakajima, H., Watanabe, S., Nakamura, M., Johnson, W. E., and Baba, H. (2012) Blockade of interleukin-6 signaling inhibits the classic pathway and promotes an alternative pathway of macrophage activation after spinal cord injury in mice. J Neuroinfl, 9: 40.
  34. Sinescu, C., Popa, F., Grigorean, V. T., Onose, G., Sandu, A. M., Popescu, M., Burnei, G., Strambu, V., and Popa, C. (2010) Molecular basis of vascular events following spinal cord injury. J Med Life, 3 (3): 254-261.
  35. Mukaino, M., Nakamura, M., Yamada, O., Okada, S., Morikawa, S., Renault-Mihara, F., Iwanami, A., Ikegami, T., Ohsugi, Y., Tsuji, O., Katoh, H., Matsuzaki, Y., Toyama, Y., Liu, M., and Okano, H. (2010) Anti-IL-6-receptor antibody promotes repair of spinal cord injury be inducing microglia-dominant inflammation. Exp Neurol, 224: 403-414.
  36. Tonai, T., Taketani, Y., and Ueda, N. (1999) Possible involvement of interleukin-1 in cyclooxygenase-2 induction after spinal cord injury in rats. J Neurochem, 72: 302-309.
  37. Schwab, J. M., Brechtel, K., and Nguyen, T. D. (2000) Persistent accumulation of cyclooxygenase-1 (COX-1) expressing microglia/macrophages and upregulation by endothelium following spinal cord injury. J Neuroimm, 111: 122-130.
  38. Sharma, H. S., Vannemreddy, P., Patnaik, R., Patnaik, S., and Mohanty, S. (2006) Histamine receptors influence blood-spinal cord barrier permeability, edema formation, and spinal cord blood flow following trauma to the rat spinal cord. Acta Neurochir Suppl, 96: 316-321.
  39. Pan, W., Kastin, A. J., Gera, L., and Stewart, J. M. (2001) Bradykinin antagonist decreases early disruption of the blood-spinal cord barrier after spinal cord injury in mice. Neurosci Lett, 307 (1): 25-28.