Neurotrophic cross-talk between the nervous and immune systems: Relevance for repair strategies in multiple sclerosis?

Journal of the Neurological Sciences 265 (2008) 93 – 96 www.elsevier.com/locate/jns

Neurotrophic cross-talk between the nervous and immune systems: Relevance for repair strategies in multiple sclerosis?☆ Reinhard Hohlfeld ⁎ Institute of Clinical Neuroimmunology, Campus Grosshadern, Ludwig-Maximilians University, Marchioninistr. 15, D-81366 Munich, Germany Department of Neuroimmunology, Max Planck-Institute of Neurobiology, Am Klopferspitz 18A, D-82152 Martinsried, Germany Received 9 February 2007; accepted 16 March 2007 Available online 24 April 2007

Abstract Autoreactive T and B cells are regular components of the healthy immune system. It has been proposed that some of these cells might have a protective function. Recent studies support this notion by demonstrating that a) myelin-autoreactive T cells show neuroprotective effects in vivo, and b) activated antigen-specific human T cells and other immune cells produce bioactive brain-derived neurotrophic factor (BDNF) and other neurotrophic factors in vitro. Furthermore, neurotrophic factors are expressed in different types of inflammatory cells in brain lesions of patients with acute disseminated leukoencephalopathy or multiple sclerosis. It seems plausible that the immune cell-mediated import of neurotrophic factors into the central nervous system has functional consequences, with obvious implications for the therapy of multiple sclerosis and other neuroimmunological diseases. © 2007 Elsevier B.V. All rights reserved. Keywords: Neuroprotection; Immunotherapy; Autoimmunity; Neurotrophic factors; Brain-derived neurotrophic factor (BDNF); Glial cell line-derived neurotrophic factor (GDNF)

1. Introduction The immune system and the nervous system are interconnected at different levels. For example, when immune cells attack the nervous system, neuroimmunological diseases arise. Multiple sclerosis and its animal models provide the paradigm for such a deleterious interaction between cells of the immune and nervous system. In experimental autoimmune encephalomyelitis (EAE) it was demonstrated that myelinreactive T cells may have – seemingly paradoxical – neuroprotective (side-) effects [3,4]. So far, the precise mechanisms of these unexpected neuroprotective effects of T cells and other immune cells have remained unknown. A different line of research has revealed that unexpectedly, T cells and other cells of the immune system are capable of ☆

This article was adapted and partly reprinted from previous publications [1,2], by permission of the publishers. ⁎ Tel.: +49 89 7095 4780; fax: +49 89 7095 4782. E-mail address: [email protected]. 0022-510X/$ - see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.jns.2007.03.012

producing neurotrophic factors (reviewed in [1]). Together, the two lines of investigation suggest that the observed neuroprotective effects of immune cells may at least partially be mediated by local secretion of neurotrophic factors. This concept has obvious implications for the therapy of neuroimmunological diseases, especially multiple sclerosis. 2. Evidence for neuroprotective functions of T cells and other immune cells Initial studies showed that myelin-reactive T cells can protect neurons from secondary degeneration after a partial crush injury of the optic nerve [3,4]. In these pioneering experiments, T cells specific for myelin basic protein (MBP), ovalbumin (OVA), or a heatshock protein (hsp) peptide were activated with their respective antigens in vitro, and then injected intraperitoneally into rats immediately after unilateral optic nerve injury. Seven days after injury, the optic nerves were analyzed immunohistochemically for the presence of T cells. Small numbers of T cells were observed in the intact

94

R. Hohlfeld / Journal of the Neurological Sciences 265 (2008) 93–96

(uninjured) optic nerves of rats injected with anti-MBP T cells. Greater numbers of T cells, however, were observed in the crushed optic nerves of the rats injected with T cells specific for MBP, hsp peptide, or OVA [3,4]. The degree of primary and secondary damages to the optic nerve axons and their attached retinal ganglion cells was measured by injecting a neurotracer distal to the site of the optic nerve lesion immediately after the injury, and again after two weeks. The percentage of labeled retinal ganglion cells (reflecting viable axons) was significantly greater in the retinas of the rats injected with anti-MBP T cells than in the retinas of rats injected with anti-OVA or anti-hsp peptide T cells. Thus, although all three T-cell lines accumulated at the site of injury, only the MBP-specific, autoimmune T cells had a substantial effect in limiting the extent of secondary degeneration. The neuroprotective effect was confirmed by electrophysiological studies [3,4]. Taken together, these results suggest that T-cell autoimmunity can mediate a significant neuroprotection after CNS injury. The authors speculate that after injury, “cryptic” epitopes might become available and might be recognized by endogenous nonencephalitogenic (benign) T cells. After local stimulation, these protective autoreactive T cells could exert their neuroprotective effect. The findings further substantiate the idea that “natural autoimmunity” can have benign consequences and may even function as a protective mechanism [5]. Subsequently, other investigators who used essentially the same experimental system were unable to reproduce some of the initially reported findings [6]. These discrepancies are presumably explained by the complexity of the experimental system(s). The group led by Dr Michal Schwartz has meanwhile extended and corroborated their earlier observations. For example, they showed that in immune deficient animals (severe combined immunodeficiency (SCID) and nude mice) hippocampal neurogenesis was impaired, but could be restored by transfer of CNS-reactive T cells [7]. 3. Role of neurotrophins in the immune system In view of their crucial functions in the nervous system, it was initially surprising to discover that some neurotrophins and their receptors act in the immune system [1]. NGF was the first neurotrophin shown to be expressed by immune cells (T- and Blymphocytes, macrophages, and mast cells). B-lymphocytes also express the two NGF receptors, p75NTR and TrkA, and can therefore react to NGF stimulation. Indeed, important functions of B cells such as proliferation, immunoglobulin production, and cell survival are regulated by NGF. Different functions of macrophages, e.g., antigen presentation and migration, are also influenced by NGF. More recently, we and others have described the expression of BDNF, another member of the NGF neurotrophin family, in immune cells [1,8,9]. It turned out that BDNF, which was previously thought to be primarily expressed in the nervous system, can be produced in vitro by essentially all major types of immune cells, including CD4+ and CD8+ T-lymphocytes, B-

lymphocytes, and monocytes. In addition, BDNF production was demonstrated in infiltrating immune cells in situ in experimental models of CNS damage and airway inflammation [1]. Comparison of mBDNF mRNA splice variants in the immune system and CNS revealed that whereas all splice variants are expressed in the CNS, only mBDNF 3 mRNA was detectable in immune cells (primary and secondary lymphoid organs, purified T cells and macrophages) [10]. After activation, only mBDNF 3 mRNA expression was upregulated in T cells. Therefore, mBDNF mRNA seems to be differentially regulated in the CNS and the immune system, opening perspectives for selective therapeutic manipulation [10]. As there are also reports indicating the presence of NT 3 and NT 4/5 in immune cells, one may conclude that most, if not all, members of the NGF neurotrophin family can be produced within the immune system. It is conceivable that immune cells can also be the target of auto- or paracrine neurotrophin actions, because they express neurotrophin receptors. It therefore appears likely that neurotrophins can mediate bi-directional cross-talk between the nervous and immune systems [1]. We also investigated the expression and function of glial cell line-derived neurotrophic factor (GDNF) family ligands (GFL) and their receptors on human immune cells [11]. The study focused on the related molecules GDNF and neurturin (NTN) and their receptors. GDNF and NTN signaling is mediated by a two-component receptor: a signal-transducing component, RET, which is shared by both ligands, and a ligand-specific binding component, GFRα-1 (higher GDNF affinity) or GFRα-2 (higher NTN affinity). We found that human T cells, B cells, and monocytes produce NTN but not GDNF, as seen by RT-PCR and immunocytochemistry. RET was expressed by B cells, T cells, and monocytes. Exons 2–5 of RET encoding the cadherin-like domains 1–3 in the extracellular part and exons 16–19 encoding a section of the second tyrosine kinase domain were transcribed in CD4+ T cells, CD8+ T cells, B cells, and monocytes. Different splice variants encoding the C-terminal intracellular part (exons 19– 21) of RET were detected. The ligand-binding receptors GFRα-1 and GFRα-2 were transcribed in all immune cell subsets. Quantitative PCR showed that GFRα-2 is by far the dominant ligand-binding chain in T cells, B cells, and monocytes. Addition of GDNF or NTN to activated PBMCs reduced the amount of detectable TNF protein without altering its transcription [11]. Together, these data demonstrate that different immune cell subsets express the GDNF family ligand NTN and different isoforms of the GFL receptors RET and GFRα. Notably, NTN and GDNF modulate TNF turnover in immune cells at the post-transcriptional level [11]. 4. Neurotrophin expression in multiple sclerosis and other neurological diseases BDNF was the first member of the neurotrophins demonstrated to be expressed in inflammatory brain lesions of MS patients [1,8,12]. In line with findings of in vitro studies, BDNF expression in MS is not restricted to neuronal cell

R. Hohlfeld / Journal of the Neurological Sciences 265 (2008) 93–96

populations. Conspicuous BDNF immunoreactivity was also demonstrated in infiltrating immune cells, especially T cells and macrophages, as well as in neurons and reactive astrocytes [1]. In particular, a larger proportion of immune cells expresses BDNF in actively demyelinating areas of MS lesions than in areas without ongoing myelin breakdown. Outside MS lesions, neurons are the major source of BDNF. It is well established that BDNF can be anterogradely transported and released by neurons. This process is upregulated after axonal injury and transection. Its common occurrence in MS lesions suggests that neuronal BDNF might also contribute to endogenous neurotrophic support in MS lesions. Neurons are considered to be the major targets for neurotrophic interactions in the CNS. In particular, the fulllength isoforms of TrkB (receptor for BDNF and NT 4/5) and TrkC (receptor for NT 3) generally occur on neuronal cells. Likewise full-length TrkB (gp145TrkB) is prominently expressed in neurons in the immediate vicinity of MS plaques [1,12]. Single neurons with pronounced gp145TrkB immunoreactivity are also observed close to MS lesions, suggesting that TrkB is upregulated in a proportion of damaged neurons [12]. An important feature of the endogenous expression of neurotrophins in MS lesions is that neurotrophins seem to be present at the actively demyelinating edge of the lesion early in its development. They are released just adjacent to axons, which may not be directly attacked by inflammatory cells, but are still at high risk of bystander damage. Thus, it is possible that a “penumbra” of subtotal damage exists around the MS plaque. A delicate balance between destructive and protective factors may prevail in this area. Neuroprotective strategies in MS could, therefore, focus on rescuing these axons, e.g., by reinforcing endogenous neurotrophic support in the periplaque area. Fewer endogenous neurotrophins are present in older, chronic MS plaques than in the early stages of lesion development. This may be one reason for the ongoing axonal degeneration in these plaques in the chronic progressive, “neurodegenerative” stage of the disease. A reasonable strategy for slowing axonal degeneration in this late phase of MS, which is notoriously resistant to other forms of therapy, would be to provide exogenous (therapeutic) neurotrophic support. Neurotrophic support by infiltrating immune cells most likely occurs in other conditions as well. Studies in experimental models of ischaemic, traumatic, or degenerative CNS disorders describe the production of BDNF and other members of the NGF neurotrophin family by infiltrating immune cells. It is noteworthy that activated microglia can serve as an additional source of neurotrophins. Since a reduction of autocrine trophic support is thought to be involved in the pathogenesis of neurodegeneration, e.g., in Parkinson's and Alzheimer's disease, one may speculate that immune cell-derived neurotrophic support represents a compensatory mechanism in such diseases. As a note of caution in interpreting the above findings it should be noted that neurotrophins have many diverse

95

functions in the CNS. These molecules are initially synthesized as precursors (proneurotrophins). Proneurotrophins are cleaved to produce mature proteins, which promote neuronal survival and enhance synaptic plasticity by activating Trk receptor tyrosine kinases. In contrast, proneurotrophins seem to interact preferentially with the p75 neurotrophin receptor (p75NTR). Interestingly, proneurotrophins often have biological effects that oppose those of mature neurotrophins. For these reasons neurotrophin activity is said to have a ‘yin and yang’ character [13]. Whether the overall effects of the neurotrophins “imported” into the CNS by immune cell are beneficial or detrimental cannot be decided on the basis of histological studies alone. Functional experiments in animal model to address this important issue are currently underway (Professor Ralf Gold, personal communication). 5. Implications for MS therapy In the past, neuroprotective therapies were mostly explored in neurodegenerative disorders like Parkinson's and Alzheimer's disease, and in ischaemic stroke. More recently, however, neuroprotection has been proclaimed an important goal for multiple sclerosis (MS) therapy. The basis for widening the scope of neuroprotection is evidence that neuronal and axonal injuries are key features of MS lesions [14–16]. In contrast with degenerative and ischaemic CNS injury however, neurodegeneration in MS appears to be caused by an inflammatory, presumably autoimmune, process. The challenge for neuroprotection in MS is therefore even greater than in degenerative and ischaemic disorders, because MS requires the combination of neuroprotective and immunomodulatory therapies. The precise mechanisms of axonal and neuronal damages in MS have not been identified [17]. According to one extreme view, MS is primarily a neurodegenerative disease. But the majority of evidence supports the more traditional view, that MS is an inflammatory rather than a degenerative disease. There are two broad categories of immunologically mediated neuronal injury: antigen-specific and antigen-nonspecific. In the first, T cells and antibodies (which belong to the adaptive, antigen-specific immune system) attack neurons and axons directly. In the second, inflammatory mediators of the innate, antigen-non-specific immune system damage neurons and axons indirectly [17]. The theoretical scenarios of antigen-specific and antigen-non-specific neuronal injury are complex, but the real-life situation could be even more intricate in view of growing evidence suggesting that the inflammatory process itself has a (neuro)protective component. This poses a therapeutic dilemma, because immunosuppressive therapies likely impair both the destructive and protective effects of inflammation. The neuroprotective strategies developed for degenerative and ischaemic disorders cannot be simply transferred to MS. There are however, options and candidate agents for neuroprotective treatment of MS [18,19]. First, it is

96

R. Hohlfeld / Journal of the Neurological Sciences 265 (2008) 93–96

conceivable that some established immunomodulatory treatments possess a neuroprotective component. It should be noted however, that any effects of immunomodulatory treatments on MRI features like brain atrophy, black holes, N-acetyl-aspartate (NAA) levels, etc., do not prove a genuinely “neurotrophic” effect, because these MRI changes could be secondary to anti-inflammatory and immunomodulatory actions of the drug in question. What are the prospects for neuroprotection in MS? So far, experience with neuroprotective therapies in stroke and neurodegenerative diseases has been disappointing [20]. Numerous sodium and calcium ion channel antagonists, Nmethyl-D-aspartate-receptor antagonists, gamma-aminobutyric acid-A receptor modulators, NO-pathway modulators, freeradical scavengers and anti-apoptotic agents, which showed promising results in animal stroke models, were ineffective in human trials [20]. This dampens any over-enthusiastic expectations for MS, where the situation is complicated by the need to combine neuroprotective and immunomodulatory agents while preserving the endogenous neuroprotective potential of inflammation. 6. Outlook In principle the treatment of multiple sclerosis has two major objectives, namely a) suppression of the inflammatory process, and b) restoration and protection of glial and neuronal functions. The potential neuroprotective function of inflammatory cells is relevant to both these treatment goals. For example, it must be assumed that many of the nonselective immunosuppressive treatments that have been used for the treatment of multiple sclerosis eliminate the neuroprotective (benign) autoimmune cells along with the autoaggressive offenders. This may be one of the reasons why treatment with conventional immunosuppressive agents often fails to show a convincing clinical benefit. Undebatably, there is a convincing rationale for immunosuppressive treatment in situations when the noxious effects of inflammation prevail. However, immunosuppressive therapy is likely to fail when the beneficial effects of the inflammatory reaction outweigh its negative consequences. In multiple sclerosis it is unfortunately unclear whether there is a stage of the disease when the inflammatory reaction is more beneficial than harmful. In any case, it is important to learn how to preserve or even enhance the proposed neuroprotective function of “benign” inflammatory cells during immunomodulatory intervention. Acknowledgement The Institute of Clinical Neuroimmunology is supported by the Hermann and Lilly Schilling Foundation.

References [1] Kerschensteiner M, Stadelmann C, Dechant G, Wekerle H, Hohlfeld R. Neurotrophic cross-talk between the nervous and immune systems: implications for neurological diseases. Ann Neurol 2003;53(3):292–304. [2] Hohlfeld R, Kerschensteiner M, Stadelmann C, Lassmann H, Wekerle H. The neuroprotective effect of inflammation: implications for the therapy of multiple sclerosis. Ernst Schering Res Found Workshop 2005;53:23–38. [3] Moalem G, Leibowitz-Amit R, Yoles E, Mor F, Cohen IR, Schwartz M. Autoimmune T cells protect neurons from secondary degeneration after central nervous system axotomy. Nat Med 1999;5(1):49–55. [4] Schwartz M, Moalem G, Leibowitz-Amit R, Cohen IR. Innate and adaptive immune responses can be beneficial for CNS repair. Trends Neurosci 1999;22(7):295–9. [5] Cohen IR. The cognitive paradigm and the immunological homunculus. Immunol Today 1992;13:490–4. [6] Jones TB, Ankeny DP, Guan Z, McGaughy V, Fisher LC, Basso DM, et al. Passive or active immunization with myelin basic protein impairs neurological function and exacerbates neuropathology after spinal cord injury in rats. J Neurosci 2004;24(15):3752–61. [7] Ziv Y, Ron N, Butovsky O, Landa G, Sudai E, Greenberg N, et al. Immune cells contribute to the maintenance of neurogenesis and spatial learning abilities in adulthood. Nat Neurosci 2006;9(2):268–75. [8] Kerschensteiner M, Gallmeier E, Behrens L, Klinkert WEF, Kolbeck R, Hoppe E, et al. Activated human T cells, B cells and monocytes produce brain-derived neurotrophic factor (BDNF) in vitro and in brain lesions: a neuroprotective role of inflammation? J Exp Med 1999;189:865–70. [9] Besser M, Wank R. Clonally restricted production of the neurotrophins brain-derived neurotrophic factor and neurotrophin-3 mRNA by human immune cells and Th1/Th2-polarized expression of their receptors. J Immunol 1999;162:6303–6. [10] Kruse N, Cetin S, Chan A, Gold R, Luhder F. Differential expression of BDNF mRNA splice variants in mouse brain and immune cells. J Neuroimmunol 2007;182(1–2):13–21. [11] Vargas-Leal V, Bruno R, Derfuss T, Krumbholz M, Hohlfeld R, Meinl E. Expression and function of glial cell line-derived neurotrophic factor family ligands and their receptors on human immune cells. J Immunol 2005;175(4):2301–8. [12] Stadelmann C, Kerschensteiner M, Misgeld T, Brück W, Hohlfeld R, Lassmann H. BDNF and gp145trkB in multiple sclerosis brain lesions: neuroprotective interactions between immune and neuronal cells? Brain 2002;125:75–85. [13] Lu B, Pang PT, Woo NH. The yin and yang of neurotrophin action. Nat Rev Neurosci 2005;6(8):603–14. [14] Trapp BD, Peterson J, Ransohoff RM, Rudick R, Mørk S, Bø L. Axonal transection in the lesion of multiple sclerosis. N Engl J Med 1998;338(5):278–85. [15] Ferguson B, Matyszak MK, Esiri MM, Perry VH. Axonal damage in acute multiple sclerosis lesions. Brain 1997;120:393–9. [16] Kornek B, Lassmann H. Axonal pathology in multiple sclerosis. A historical note. Brain Pathol 1999;9:651–6. [17] Hauser SL, Oksenberg JR. The neurobiology of multiple sclerosis: genes, inflammation, and neurodegeneration. Neuron 2006;52(1):61–76. [18] Kapoor R. Neuroprotection in multiple sclerosis: therapeutic strategies and clinical trial design. Curr Opin Neurol 2006;19(3):255–9. [19] Dhib-Jalbut S, Arnold DL, Cleveland DW, Fisher M, Friedlander RM, Mouradian MM, et al. Neurodegeneration and neuroprotection in multiple sclerosis and other neurodegenerative diseases. J Neuroimmunol 2006;176(1–2):198–215. [20] Muir KW, Teal PA. Why have neuro-protectants failed?: lessons learned from stroke trials. J Neurol 2005;252(9):1011–20.