Neural stem cell therapy

The rise of precursor cell biology has brought new life to neural transplantation and the consideration of cellular replacement strategies to treat diseases of the brain. The idea of

“making new neurons” is appealing for neurodegenerative diseases, or selective neuronal loss associated with chronic neurological or psychiatric disorders. One goal of neural precursor biology is to learn from this regionally limited, constitutive neurogenesis how to manipulate neural precursors towards therapeutically useful neuronal or glial population. Elucidation of the relevant molecular control of neurogenesis, morphological maturation of newly generated neural cell types and their functional insertion into the adult brain might result in the development of cell replacements paradigm either by transplantation of cells or by recruitment of endogenous cells.

Some studies reported a surprisingly broad differentiation potential of neural stem cells into cell types of other organs or even other germ layers, but these observations remain a source of debates and controversies (Anderson et al., 2001). In fact, some reports have raised concerns that the broader potential of neural stem cells could derive from phenomenon like cell transformation, transdifferentiation or fusion, all of which can affect the use of neural stem cells for therapy. For example, cell transformation may be associated with aberrant cell growth, and with the risk of tumor formation upon grafting. Thus, neuronal replacement therapies based on manipulation of endogenous precursors may be an attractive aim in the future. However, many questions must be answered before neuronal replacement therapies using endogenous precursors become reality. The multiple signals that are responsible for endogenous precursor division, migration, differentiation, axon extension, circuit integration and survival will need to be elucidated in order for such therapies to be developed efficiently.

These challenges also exist for neuronal replacement strategies based upon transplantation of precursors, because donor cells, whatever may be their source, must interact with an extremely complex and intricate mature CNS environment in order to functionally integrate into the brain.

Altogether, NSCs might be considered as candidate cells to develop cell-based therapies for neurological disorders. While ES can be efficiently expanded in vitro and are capable to give rise to all cell types of the body, these cells might form tumors upon transplantation. A mulitpotent cell type for the development of cell based therapies is NSC that is considered not to form tumours (Winkler et al., 1998; Englund et al., 2002).

Futhermore, the use of human ES cells is limited because of ethical concerns. Similarly, the use of fetal-derived primary cells for cell-based therapies is associated with ethical and also logistic problems. Efficient cell replacement strategies require widespread integration, long-term survival and differentiation of grafted cells and, in particular, cell types that are capable to functionally integrate into the host tissue. Manipulation of NSCs prior to transplantation might be one strategy to achieve this aims. Indeed, several studies have demonstrated that neural stem/precursor cells might be “optimized” for cell based therapies by genetic manipulations. For instance, overexpression of PSA in neural precursor cells has been demonstrated to play an instructive role on the choice of a cells migration pathway depending on the environment (Franceschini et al., 2004). Furthermore, forced expression of the transcription factor Nurr1 has been shown to direct the differentiation of NSCs into dopaminergic neurons (Andersson et al., 2007; Li et al., 2007; Shim et al., 2007). Another example is the directed differentitation of neural precursor cells into actively myelinating oligodendrocytes by overexpression of the transcription factor olig2 (Copray et al., 2006).

Neural precursor cells derived from L1-transfected embryonic stem cells showed decreased cell proliferation in vitro, enhanced neuronal differentiation in vitro and in vivo, and decreased astrocytic differentiation in vivo without influencing cell death. L1 overexpression also resulted in an increased yield of GABAergic neurons and enhanced migration of embryonic stem cell-derived neural precursor cells into the excitotoxicly lesioned striatum.

Mice grafted with L1-transfected cells showed recovery in rotation behavior 1 and 4 weeks, but not 8 weeks, after transplantation compared with mice that had received nontransfected cells, thus demonstrating for the first time that neural precursor cells with a transgenic expression of a recognition molecule are capable to improve functional recovery during the initial phase in a syngeneic transplantation paradigm (Bernreuther et al., 2006).

In another study, precursor cells derived from L1-transfected embryonic stem cells and injected into the lesion site after spinal cord injury migrated rostrally and caudally from the lesion. Anterogradely labeled corticospinal tract axons showed interdigitation with L1-positive donor cells and extended into the lesion site 1 month after transplantation and, in some cases, extended beyond the lesion site (Chen et al., 2005).

The fate and behaviour of stem cells evidently depends on environmental cues and cell intrinsic properties that probably interact with each other. Which are the environmental factors that influence the migration and differentiation of endogenous and transplanted stem cells and which mechanisms are employed? Understanding which cellular characteristics make a NSC and which events guide them to different neural cell fates could also open up strategies in the therapeutic use of stem cells which is engaging the endogenous pool of NSCs in the CNS. The aim of the study was reveal L1 role in controlling or influencing NSC fate decisions and behaviour following transplantation into a diseased adult brain region.

2. Rationale and aims of the study

NSCs are among the candidate cell type to develop cell-based therapies for a variety of neurological disorders. Strategies dependent on efficient cell replacement, widespread integration, long-term survival and directed differentiation of grafted cells into particular neutal cell types. During the development of the nervous system, L1 has ben implicated in migration of postmitotic neurons, outgrowth, pathfinding and fasciculation of axons, growth cone morphology and myelination. In addition, L1 has been implicated in axonal regeneration, neuronal cell survival, and proliferation and fate decision of neuronal precursor.

The aim of the present study was to generate stably L1-transfected neural precursor population, and to analyze the impact of ectopic L1 expression on the functional properties of these cells in vitro and after transplantation in vivo. The specific aim included:

1. To generate stably L1-transfected neural precursor population.

2. To analyze the effect of ectopic L1 expression on fate decision of NSCs in vivo.

3. To study the migrating capacity, fate decision and the survival of L1-positive NSCs after transplantation into a mouse model for Huntingtons disease.

3. Materials and methods