Replacing Cells in Multiple Sclerosis ⁎ Ian D

Journal of the Neurological Sciences 265 (2008) 89–92 www.elsevier.com/locate/jns

Replacing cells in multiple sclerosis ⁎ Ian D. Duncan

Department of Medical Sciences, School of Veterinary Medicine, University of Wisconsin-Madison, 2015 Linden Drive, Madison, WI 53706, USA Received 1 March 2007; accepted 2 April 2007 Available online 22 June 2007

Abstract

Cell transplantation is emerging as a major potential therapeutic approach in the treatment of otherwise untreatable neurodegenerative diseases. In multiple sclerosis (MS), a major direction of current research is to devise strategies that will remyelinate axons and protect them against subsequent ongoing degeneration. Ongoing loss of axons will lead to chronic disability. Oligodendrocytes and their progenitors are lost during multiple relapses in the course of MS and either needs to be replaced from an exogenous source or the remaining progenitors stimulated to differentiate and remyelinate. The successful isolation and purification of human oligodendrocytes from neural or embryonic stem cells offer hope that a source of sufficient cells for translational application might be achievable in the future. Focal repair of strategic lesions followed by more disseminated delivery of exogenous cells will be the short and long-term goals. © 2007 Elsevier B.V. All rights reserved.

Keywords: Stem cells; Multiple sclerosis; Remyelination; Demyelinated axons

1. Introduction cells in a wide range of animal models. To treat MS lesions, human cells will likely be used initially, and the isolation and Novel ideas are emerging on the treatment of multiple purification of large numbers of human oligodendrocytes are sclerosis (MS) that are aimed at promoting remyelination and likely to be the key requirement for the translation of the axonal protection, thus addressing different targets from the experimental studies into clinical practice. While advances current disease modifying drugs which are designed to modify are being made in isolating human cells, other complemen- the immune response and lessen inflammation, but which do tary advances in our knowledge of MS are being reported. not address tissue repair or neuroprotection. These new strat- Much more is being discovered on the pathology of MS and egies would involve the replacement of lost oligodendrocytes of imaging lesions that will provide the necessary back- or oligodendrocyte precursor cells (OPCs) from an exogenous ground information needed prior to transplanting cells. source. Replacement of oligodendrocytes could also be pro- In this brief review on the topic of cell replacement in MS moted, however, by the “rescue” of OPCs that remain in, or lesions, I will consider some of these issues and in particular, near lesions, but appear to be unable to differentiate and suc- discuss the replacement of cells in a single, strategic lesion in cessfully remyelinate chronically demyelinated axons [1]. the CNS. The long-term goal, however, will be to bring about The impetus to explore cell therapy in MS has come from the diffuse dissemination of exogenous cells to repair multiple many years of experimental work on the transplantation of areas of demyelination throughout the brain and spinal cord. myelin-forming cells into the CNS [2–5], and from clinical and experimental experience of neural stem cell therapies in 2. Choice of lesions other neurodegenerative diseases [6,7]. There is now a large body of evidence that shows that glial cell transplantation There are two major decisions to be made here that remain can result in extensive myelin formation by the transplanted unresolved. The first choice is whether to target acute or chronic areas of the demyelination and secondly at which site in the ⁎ Tel.: +1 608 263 9829; fax: +1 608 265 8008. CNS could clinically significant lesions be surgically E-mail address: [email protected]. approached. The choice between acute or chronic lesions is

0022-510X/$ - see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.jns.2007.04.043 90 I.D. Duncan / Journal of the Neurological Sciences 265 (2008) 89–92 the more complex. In early relapsing–remitting disease, seen in Fig. 1, there are many astrocytes, likely some cells of spontaneous remyelination is profuse [8,9].Indeed,in the oligodendrocyte lineage, most likely OPCs, microglia experiment models of demyelination, especially those caused and perhaps some residual lymphocytes and macrophages. It by myelinotoxic chemicals, complete endogenous remyelina- appears that OPCs within such a chronic plaque differentiate tion can occur [10]. In MS, remyelination has been described in poorly or if they do, are unable to normally ensheathe and the presence of inflammatory cell infiltrate, suggesting that remyelinate axons [14,15]. Attempts to remyelinate such OPCs are not inhibited by certain aspects of the immune/ areas by transplantation of exogenous cells will not need to inflammatory response, and may actually require this for remove these cells. Experimental studies have shown that remyelination to occur. Evidence of the latter comes from a myelination or re-myelination occurs in the presence of number of experiments. It has been shown that rejection of endogenous cells of the oligodendrocyte lineage, although it transplanted cells can result in extensive remyelination by has been suggested host OPCs may inhibit exogenous cells neighboring endogenous OPCs [11], and that in chronic areas from remyelinating lesions. An animal model in which of demyelination, extensive remyelination by transplanted cells chronic areas of demyelination resemble those of on MS only occurs when focal areas of inflammation are induced [12]. plaque would be extremely helpful here to test this question. Likewise, focal inflammation can promote the myelinating Such a model could also be used to study the means to pro- capacity of cells transplanted into areas containing normally mote endogenous repair. Ideally, such lesions would consist non-myelinated axons [13].ThedilemmainMS,however,is of chronically demyelinated (over 4–6 months) axons in a that acute lesions are likely to spontaneously remyelinate at gliotic milieu in which inactive OPCs are preserved. In our least early in disease, and implanting cells into such lesion may studies on the transplantation of glia into the shaking pup impede this natural process. Therefore repair of plaques with mutant, we were able to show that OPCs transplanted into persistent demyelination associated with fixed and chronic the mature mutant CNS in which there is a marked gliosis, neurological deficits is the most pressing need for cellular were able to myelinate a large number of axons at the site [4]. therapy. For therapy of such lesions to be useful, however, there Thus astrocytes and the gliosis seen in MS plaques may not must be sufficient axons in the plaque to remyelinate. prevent transplanted cells from myelinating axons, although Single, strategic lesions that might be targeted for focal repair in severely gliotic lesions this may restrict migration of are in the spinal cord, brain stem, cerebellar peduncles or optic OPCs and ensheathment of axons. Further testing of this nerve. Lesions in the cervical spinal cord that cause either all or question in appropriate models is desirable. the major clinical dysfunction might be the most promising target. Much of the experimental work on focal implantation of 4. Cells to be used for replacement cells has been carried out in the spinal cord. However, the caution here is that any transplantation of cells should not cause This remains the greatest impediment to the translational significant damage to intact myelinated axons in adjacent long- application of cellular therapy in the repair of focal lesions. tracts. The optic nerve is an attractive target as focal lesions in a The choices of cell type ranges from cells of the single nerve could be viewed as a low risk target as there is no oligodendrocyte lineage, to Schwann cells and olfactory risk to adjacent CNS tissue. However, the optic nerve has ensheathing cells. We have focused our efforts on oligoden- proven to be a difficult anatomic structure to penetrate and drocytes and OPCs. Unlike rodent or other animal tissue transplant cells successfully into because of the thick, tough dura oligodendrocytes derived, it has proven difficult to isolate and astrocytic compartmentation that characterize the nerve. large numbers of OPCs from human tissue. The in vitro The brain stem is clearly a complex site in which to transplant conditions required to maintain human OPCs and promote cells because of the critical nuclei throughout the medulla. Both their division are different from other species [16]. We have it and the cerebellar peduncles would require stereotactic ap- shown, as have others, that OPCs can be isolated from proaches, but neither should be excluded from consideration. neurospheres, grown from the striata of the human fetal brain [16]. We have also shown that OPCs can be differentiated 3. Cells within the plaque from human embryonic stem cells (hESCs) [17]. However, in both cases, the number of cells was insufficient for If a single, chronic MS plaque is to be targeted for cell translational purposes. However, other investigators have replacement, it is important to consider the milieu into which shown more promising data. In the case of fetal or even the cells might be transplanted. The chronic MS plaque is not adult brain, it is possible to isolate relatively large numbers acellular (Fig. 1). Within areas of chronic demyelination as of OPCs by the use of progenitor cell-selective reporter gene

Fig. 1. A) A chronic MS plaque, stained for myelin. Note that there are many cells throughout the plaque but no evidence of perivascular accumulation of T cells. The identity of cells within the plaque is possible through appropriate immunolabeling (modified from Prineas and Wright. 1978, with permission). B) To replace cells of the oligodendrocyte lineage, lost over time in the plaque, persistent OPCs, either in the plaque or closely adjacent, might be recruited. It is likely that such cells will need to proliferate before they ensheathe and remyelinate axons. To recruit dormant, endogenous OPCs a number of approaches might be used, which are noted. As more becomes known about the molecular differences between myelination and remyelination, so the ability to control the appropriate temporal and spatial expression of transcription factors in OPCs might be critical for successful endogenous repair. C) Exogenous repair by the injection of OPCs may result in complete repair of large lesions if sufficient numbers of cells are delivered, or divide appropriately on implantation and differentiate into myelinating cells. I.D. Duncan / Journal of the Neurological Sciences 265 (2008) 89–92 91 92 I.D. Duncan / Journal of the Neurological Sciences 265 (2008) 89–92 expression [18]. Cells are transduced with a fluorescent References reporter gene such as green fluorescent protein under the control of the myelin gene CNP. Fluorescent positive cells [1] Dubois-Dalcq M, ffrench-Constant C, Franklin RJM. Enhancing central can then be isolated and purified by fluorescent activated cell nervous system remyelination in multiple sclerosis. Neuron 2005;48 (1):9–12. sorting (FACS) [18]. FACS sorting using antibodies against [2] Duncan ID, Milward EA. Glial cell transplants: experimental therapies surface antigens of human fetal or adult progenitor cells can of myelin diseases. Brain Pathol 1995;5:301–10. also be used to isolate populations of cells for transplantation [3] Blakemore WF, Crang AJ, Franklin RJM. Transplantation of glial cells. [19]. Likewise, with human embryonic stem cells, Keirstead In: Ransom BR, Kettenmann H, editors. Neuroglial Cells. Cambridge: – and colleagues have successfully differentiated ESCs to Oxford University Press; 1995. p. 869 82. [4] Archer DR, Cuddon PA, Lipsitz D, Duncan ID. Myelination of the OPCs with a resultant 90% purity [20]. It will be important to canine central nervous system by glial cell transplantation: a model for test the remyelinating capacities of OPCs derived as cited repair of human myelin disease. Nature Med 1997;3:54–9. above, in models that mimic both acute and chronic MS, [5] Duncan ID, Grever WE, Zhang S-C. Repair of myelin disease: strategies most likely in experimental allergic encephalomyelitis. and progress in animal models. Mol Med Today 1997;3:554–61. [6] Björklund A, Lindvall O. Cell replacement therapies for central nervous system disorders. Nat Neurosci 2000;3(6):537–44. 5. Analyses of the outcome of cell replacement [7] Martino G, Pluchino S. The therapeutic potential of neural stem cells. Nat Rev Neurosci 2006;7(5):395–406. An important component of this new therapy will be to [8] Patrikios P, Stadelmann C, Kutzelnigg A, Rauschka H, Schmidbauer devise outcome measures that can critically evaluate the success M, Laursen H, et al. Remyelination is extensive in a subset of multiple of the cell replacement strategy. The goal of focal repair, how- sclerosis patients. Brain 2006. [9] Prineas JW, McDonald WI. Demyelinating diseases. In: Graham DI, ever, is to improve or restore function; hence detailed neuro- Lantos PL, editors. Greenfield's Neuropathology; 1997. p. 813–81. logical evaluation must be the primary method of judging [10] Blakemore WF, Crang AJ, Evans RJ. The effect of chemical injury on success. However, imaging of the lesion or lesions will also be oligodendrocytes. In: Cuzner ML, Kelly RE, editors. Viruses and important for two reasons. Firstly, it may be possible to follow Demyelinating Diseases. London: Academic Press; 1983. p. 167–90. cells after implantation if they are labeled with superparamag- [11] Blakemore WF, Crang AJ, Franklin RJM, Tang K, Ryder S. Glial cell – transplants that are subsequently rejected can be used to influence netic iron particles (SPIO) prior to transplantation [21 23]. regeneration of glial cell environments in the CNS. GLIA 1995;13:79–91. Secondly, the MRI appearance of the plaque and whether it has [12] Foote AK, Blakemore WF. Inflammation stimulates remyelination in the features of remyelination (although more needs to be known areas of chronic demyelination. Brain 2005;128:528–39. about the MRI characteristics of remyelination) will be impor- [13] Setzu A, Lathia JD, Zhao C, Wells K, Rao MS, ffrench-Constant C, et al. tant to evaluate. It would seem likely that progress in imaging, Inflammation stimulates myelination by transplanted oligodendrocyte precursor cells. GLIA 2006;54(4):297–303. using contemporary imaging techniques such as diffusion [14] Chang A, Nishiyama A, Peterson J, Prineas J, Trapp BD. NG2-positive tensor, will provide the prerequisite surrogate markers of axon oligodendrocyte progenitor cells in adult human brain and multiple survival and CNS remyelination within the next few years. sclerosis lesions. J Neurosci 2000;20:6404–12. [15] Chang A, Tourtellotte WW, Rudick R, Trapp BD. Premyelinating 6. Conclusion oligodendrocytes in chronic lesions of multiple sclerosis. N Engl J Med 2002;346(3):165–73. [16] Zhang S-C, Ge B, Duncan ID. Tracing human oligodendroglial Remyelination of demyelinated axons in MS is a rational development in vitro. J Neurosci Res 2000;59:421–9. and important strategy that will help to restore function to [17] Zhang S-C, Wernig M, Duncan ID, Brüstle O, Thomson JA. In vitro axons and protect them against ongoing loss. While current and differentiation of transplantable neural precursors from human embryonic – future medical therapies may prove successful in limiting the stem cells. Nat Biotechnol 2001;19:1129 33. [18] Goldman S. Stem and progenitor cell-based therapy of the human early loss of axons, the greatest challenge appears to be central nervous system. Nat Biotechnol 2005;23(7):862–71. remyelinating chronically demyelinated axons in non-repairing [19] Windrem MS, Nunes MC, Rashbaum WK, Schwartz TH, Goodman RA, lesions. It may not be possible, however, to devise experimen- McKhann G, et al. Fetal and adult human oligodendrocyte progenitor cell tal models that represent such a disease stage, allowing the isolates myelinate the congenitally dysmyelinated brain. Nat Med – testing of transplant-induced repair. Thus transplanting cells 2004;10(1):93 7. “ ” [20] Nistor GI, Totoiu MO, Haque N, Carpenter MK, Keirstead HS. Human into chronic, focal MS lesions may be the final experiment. embryonic stem cells differentiate into oligodendrocytes in high purity However, such a clinical trial in MS patients may be the only and myelinate after spinal cord transplantation. GLIA 2005;49:385–96. way to determine whether exogenous cells are capable of [21] Bulte JWM, Zhang S-C, van Gelderen P, Herynek V,Jordan EK, Duncan surviving, differentiating and myelinating axons, in plaques ID, et al. Neurotransplantation of magnetically labeled oligodendrocyte where endogenous OPCs are present, but incapable of repair. progenitors: MR tracking of cell migration and myelination. Proc Natl Acad Sci 1999. [22] Bulte JWM, Douglas T, Witwer B, Zhang S-C, Strable E, Lewis BK, et al. Acknowledgements Magnetodendrimers allow endosomal magnetic labeling and in vivo tracking of stem cells. Nat Biotechnol 2001;19(12):1141–7. I would like to thank the many colleagues and collaborators [23] Franklin RJM, Blaschuk KL, Bearchell MC, Prestoz LLC, Setzu A, Brindle in my lab who have contributed to our cell transplantation KM, et al. Magnetic resonance imaging of transplanted oligodendrocyte precursors in the rat brain. NeuroReport 1999;10:3961–5. studies over the years. Our current work is supported by the NMSS Grant #TR 3761.