Implantation of stimulated homologous macrophages results in partial recovery of paraplegic rats.

Postinjury recovery in most tissues requires an effective dialog with macrophages; however, in the mammalian central nervous system, this dialog may be restricted (possibly due to its immune-privileged status), which probably contributes to its regeneration failure. We circumvented this by implanting macrophages, pre-exposed ex vivo to peripheral nerve segments, into transected rat spinal cord. This stimulated tissue repair and partial recovery of motor function, manifested behaviorally by movement of hind limbs, plantar placement of the paws and weight support, and electrophysiologically by cortically evoked hind-limb muscle response. We substantiated these findings immunohistochemically by demonstrating continuity of labeled nerve fibers across the transected site, and by tracing descending fibers distally to it by anterograde labeling. In recovered rats, retransection of the cord above the primary transection site led to loss of recovery, indicating the involvement of long descending spinal tracts. Injection of macrophages into the site of injury is relatively non-invasive and, as the cells are autologous, it may be developed into a clinical therapy.

The remedy may lie in ourselves: prospects for immune cell therapy in central nervous system protection and repair.

The irreversible loss of function after axonal injury in the central nervous system (CNS) is a result of the lack of neurogenesis, poor regeneration, and the spread of damage caused by toxicity emanating from the degenerating axons to uninjured neurons in the vicinity. Now, 100 years after Ramon y Cajal's discovery that CNS neurons--unlike neurons of the peripheral nervous system--fail to regenerate, it has become evident that (a) CNS tissue is indeed capable of regenerating, at.least in part, provided that it acquires the appropriate conditions for growth support, and (b) that the spread of damage can be stopped and the postinjury rescue of neurons thus achieved, if ways are found to neutralize the mediators of toxicity, either by inhibiting their action or by increasing tissue resistance to them. In most physiological systems the processes of tissue maintenance and repair depend on the active assistance of immune cells. In the CNS, however, communication with the immune system is restricted. The accumulated evidence from our previous studies suggests that the poor posttraumatic repair and maintenance in the CNS is due at least in part to this restriction. Key factors in the recovery of injured tissues, but missing or deficient in the CNS, are the processes of recruitment and activation of immune cells. We therefore propose the development of immune cell therapies in which the injured CNS is exogenously provided with an adequate number of appropriately activated immune cells (macrophages for regrowth and autoimmune T cells for maintenance), controlled in such a way as to derive maximal benefit with minimal risk of disease. It is expected that these self-adjusting cells will communicate with the damaged tissue, monitor tissue needs, and control the dynamic course of CNS healing.

Potential repair of rat spinal cord injuries using stimulated homologous macrophages.

The failure of the adult mammalian central nervous system (CNS) to regenerate after injury has long been viewed as a unique phenomenon resulting from the specific nature of this system. The finding that some CNS axons could be induced to regrow if provided with a permissive environment suggested that this failure is a result, at least in part, of the nature of the postinjury neuronal environment. It was further shown that the involvement of inflammatory cells, particularly macrophages, in postinjury processes in the CNS is limited. We have suggested that, to achieve recovery after injury, the adult mammalian CNS may require the assistance of the same postinjury factors as those involved in the recovery of spontaneously regenerating systems but that its accessibility to such assistance is restricted. Accordingly, we proposed that it might be possible to circumvent the restriction, allowing regeneration to occur. We showed that the implantation of autologous macrophages, which had been prestimulated by exposure to a regenerative (sciatic) nerve, into completely transected spinal cords of adult rats led to partial motor recovery. This treatment intervenes in the postinjury process by simulating in the axotomized CNS the events that occur naturally in spontaneously regenerating systems.

Link between optic nerve regrowth failure and macrophage stimulation in mammals.

The adult mammalian central nervous system (CNS) fails to regenerate its axons following injury. A comparison between its postinjury response and that of axons of nervous systems capable of regeneration reveals major differences with respect to inflammation. In regenerative systems, a large number of macrophages rapidly invade the injured site during the first few hours and days after the injury. Following their activation/differentiation through interaction with the host tissue, they play a central role in tissue healing through phagocytosis of cell debris and communication with cellular and molecular elements of the damaged tissue. Relative to the peripheral nervous system (PNS), macrophage recruitment in the adult mammalian CNS is delayed and is restricted in amount and activity. It was recently proposed that in injured mammalian CNS tissue, implantation of macrophages stimulated by prior co-culture with segments of peripheral (sciatic) nerves can compensate, at least in part, of the restricted postinjury inflammatory reaction. In the present study, this experimental paradigm is further explored and shows that there is no conflict between the systemic use of anti-inflammatory compounds and treatment with stimulated macrophages to promote regrowth of neuronal tissue.

Restricted inflammatory reaction in the CNS: a key impediment to axonal regeneration?

Axons in the central nervous system (CNS) of adult mammals do not regenerate after injury. Mammalian CNS differs in this respect from other mammalian tissues, including the peripheral nervous system (PNS), and from the CNS of lower vertebrates. In most parts of the body, including the nervous system, injury triggers an inflammatory reaction involving macrophages. This reaction is needed for tissue healing; when it is delayed or insufficient, healing is incomplete. The CNS, although needing an efficient inflammatory reaction resembling that in the periphery for tissue healing, appears to have lost the ability to supply it. We suggest that restricted CNS recruitment and activation of macrophages are linked to regeneration failure and might reflect the immune privilege that characterizes the mammalian CNS. As macrophages play a critical role in tissue restoration, and because their recruitment and activation are among the most upstream of the events leading to tissue healing, overcoming the deficiencies in these steps might trigger a self-repair processing leading to recovery after CNS injury.

Peripheral nerve-stimulated macrophages simulate a peripheral nerve-like regenerative response in rat transected optic nerve.

We have previously demonstrated that the failure of the mammalian central nervous system (CNS) to regenerate following axonal injury is related to its immunosuppressive nature, which restricts the ability of both recruited blood-borne monocytes and CNS-resident microglia to support a process of repair. In this study we show that transected optic nerve transplanted with macrophages stimulated by spontaneously regenerating nerve tissue, e.g., segments of peripheral nerve (sciatic nerve), exhibit axonal regrowth at least as far as the optic chiasma. Axonal regrowth was confirmed by double retrograde labeling of the injured optic axons, visualized in their cell bodies. Transplanted macrophages exposed to segments of CNS (optic) nerve were significantly less effective in inducing regrowth. Immunocytochemical analysis showed that the induced regrowth was correlated with a wide distribution of macrophages within the transplanted-transected nerves. It was also correlated with an enhanced clearance of myelin, known to be inhibitory for regrowth and poorly eliminated after injury in the CNS. These results suggest that healing of the injured mammalian CNS, like healing of any other injured tissue, requires the partnership of the immune system, which is normally restricted, but that the restriction can be circumvented by transplantation of peripheral nerve-stimulated macrophages.

Differential effects of central and peripheral nerves on macrophages and microglia.

The poor ability of injured central nervous system (CNS) axons to regenerate has been correlated, at least partially, with a limited and suppressed postinjury inflammatory response. A key cell type in the inflammatory process is the macrophage, which can respond in various ways, depending on the conditions of stimulation. The aim of this study is to compare the activities of macrophages or microglia when encountering CNS and peripheral nervous systems (PNS), on the assumption that nerve-related differences in the inflammatory response may have implications for tissue repair and thus for nerve regeneration. Phagocytic activity of macrophages or of isolated brain-derived microglia was enhanced upon their exposure to sciatic (PNS) nerve segments, but inhibited by exposure to optic (CNS) nerve segments. Similarly, nitric oxide production by macrophages or microglia was induced by sciatic nerve segments but not by optic nerve segments. The previously demonstrated presence of a resident inhibitory activity in CNS nerve, could account, at least in part, for the inhibited phagocytic activity of blood-borne macrophages in CNS nerve as well as of microglia resident in the brain. It seems that the CNS microglia are reversibly immunosuppressed by the CNS environment, at least with respect to the activities examined here. It also appears from this study that the weak induction of early healing-related activities of macrophages/microglia in the environment of CNS might explain the subsequent failure of this environment to acquire growth-supportive properties in temporal and spatial synchrony with the needs of regrowing axons.

Transplantation of activated macrophages overcomes central nervous system regrowth failure.

Macrophages have long been known to play a key role in the healing processes of tissues that regenerate after injury; however, the nature of their involvement in healing of the injured central nervous system (CNS) is still a subject of controversy. Here we show that the absence of regrowth in transected rat optic nerve (which, like all other CNS nerves in mammals, cannot regenerate after injury) can be overcome by local transplantation of macrophages preincubated ex vivo with segments of a nerve (e.g., sciatic nerve) that can regenerate after injury. The observed effect of the transplanted macrophages was found to be an outcome of their stimulated activity, as indicated by phagocytosis. Thus, macrophage phagocytic activity was stimulated by their preincubation with sciatic nerve segments but inhibited by their preincubation with optic nerve segments. We conclude that the inability of nerves of the mammalian CNS to regenerate is related to the failure of their macrophages recruited after injury to acquire growth-supportive activity. We attribute this failure to the presence of a CNS resident macrophage inhibitory activity, which may be the biochemical basis underlying the immune privilege of the CNS. The transplantation of suitably activated macrophages into injured nerves may overcome multiple malfunctioning aspects of the CNS response to trauma, and thus may be developed into a novel, practical, and multipotent therapy for CNS injuries.