A study was undertaken to review international literature pertaining to people’s behavior in and around floodwater. The review focused on people’s voluntary entry of floodwater. From the literature, five predominant reasons for entering floodwater were identified, including undertaking a recreational activity; attempting to reach a destination; retrieving property, livestock, or pets; undertaking employment duties; and rescuing or assisting with evacuation. Two primary influences on entering floodwater were found, namely risk perception (i.e., being unaware of or underestimating the risk from flooding) and social influences (i.e., being influenced by others). Demographics and environmental and temporal factors also played a part in decision-making about whether to enter floodwater or not. Emergency managers should take account of such factors when devising future public education strategies. Further research, including comparisons with current theoretical models, could help identify additional influences on decision-making for floodwater entry.

Traditionally, in vitro stem cell systems have used oxygen tensions that are far removed from the in vivo situation. This is particularly true for the central nervous system, where oxygen (O-2) levels range from 8% at the pia to 0.5% in the midbrain, whereas cells are usually cultured in a 20% O-2 environment. Cell transplantation strategies therefore typically introduce a stress challenge at the time of transplantation as the cells are switched from 20% to 3% O-2 (the average in adult organs). We have modeled the oxygen stress that occurs during transplantation, demonstrating that in vitro transfer of neonatal rat cortical neural precursor cells (NPCs) from a 20% to a 3% O-2 environment results in significant cell death, whereas maintenance at 3% O-2 is protective. This survival benefit translates to the in vivo environment, where culture of NPCs at 3% rather than 20% O-2 approximately doubles survival in the immediate post-transplantation phase. Furthermore, NPC fate is affected by culture at low, physiological O-2 tensions (3%), with particularly marked effects on the oligodendrocyte lineage, both in vitro and in vivo. We propose that careful consideration of physiological oxygen environments, and particularly changes in oxygen tension, has relevance for the practical approaches to cellular therapies.

Background More than half of patients with multiple sclerosis have progressive disease characterised by accumulating disability. The absence of treatments for progressive multiple sclerosis represents a major unmet clinical need. On the basis of evidence that mesenchymal stem cells have a beneficial effect in acute and chronic animal models of multiple sclerosis, we aimed to assess the safety and efficacy of these cells as a potential neuroprotective treatment for secondary progressive multiple sclerosis.Methods Patients with secondary progressive multiple sclerosis involving the visual pathways (expanded disability status score 5.5-6.5) were recruited from the East Anglia and north London regions of the UK. Participants received intravenous infusion of autologous bone-marrow-derived mesenchymal stem cells in this open-label study. Our primary objective was to assess feasibility and safety; we compared adverse events from up to 20 months before treatment until up to 10 months after the infusion. As a secondary objective, we chose efficacy outcomes to assess the anterior visual pathway as a model of wider disease. Masked endpoint analyses was used for electrophysiological and selected imaging outcomes. We used piecewise linear mixed models to assess the change in gradients over time at the point of intervention. This trial is registered with ClinicalTrials.gov, number NCT00395200.Findings We isolated, expanded, characterised, and administered mesenchymal stem cells in ten patients. The mean dose was 1.6x10(6) cells per kg bodyweight (range 1.1-2-0). One patient developed a transient rash shortly after treatment; two patients had self-limiting bacterial infections 3-4 weeks after treatment. We did not identify any serious adverse events. We noted improvement after treatment in visual acuity (difference in monthly rates of change) -0.02 logMAR units, 95% CI -0.03 to -0.01; p=0.003) and visual evoked response latency (-1.33 ms, 2-44 to). 21; p=0.020), with an increase in optic nerve area (difference in monthly rates of change 0.13 mm(2),0.04 to 0.22; p=0.006). We did not identify any significant effects on colour vision, visual fields, macular volume, retinal nerve fibre layer thickness, or optic nerve magnetisation transfer ratio.Interpretation Autologous mesenchymal stem cells were safely given to patients with secondary progressive multiple sclerosis in our study. The evidence of structural, functional, and physiological improvement after treatment in some visual endpoints is suggestive of neuroprotection.

This protocol has been designed to generate neural precursor cells (NPCs) from human embryonic stem cells (hESCs) using a physiological oxygen (O2) level of 3% (previously termed hypoxia) and chemically defined conditions. The first stage involves suspension culture of hESC colonies at 3% O2, where they acquire a neuroepithelial identity over a period of 2 weeks. This timescale is comparable to that observed at 20% O2, but survival is enhanced. Sequential application of retinoic acid and purmorphamine (PM), from day 14 to day 28, directs differentiation toward spinal motor neurons. Alternatively, addition of fibroblast growth factor-8 and PM generates midbrain dopaminergic neurons. OLIG2 (encoding oligodendrocyte lineage transcription factor 2) induction in motor neuron precursors is twofold greater than that at 20% O2, whereas EN1 (encoding engrailed homeobox 1) expression is enhanced fivefold. NPCs (at 3% O2) can be differentiated into all three neural lineages, and such cultures can be maintained long term in the absence of neurotrophins. The ability to generate defined cell types at 3% O2 should represent a significant advancement for in vitro disease modeling and potentially for cell-based therapies.

BACKGROUND:No treatments are currently available that slow, stop, or reverse disease progression in established multiple sclerosis (MS). The Mesenchymal Stem Cells in Multiple Sclerosis (MSCIMS) trial tests the safety and feasibility of treatment with a candidate cell-based therapy, and will inform the wider challenge of designing early phase clinical trials to evaluate putative neuroprotective therapies in progressive MS. Illustrated by the MSCIMS trial protocol, we describe a novel methodology based on detailed assessment of the anterior visual pathway as a model of wider disease processes--the "sentinel lesion approach". METHODS/DESIGN:MSCIMS is a phase IIA study of autologous mesenchymal stem cells (MSCs) in secondary progressive MS. A pre-test : post-test design is used with healthy controls providing normative data for inter-session variability. Complementary eligibility criteria and outcomes are used to select participants with disease affecting the anterior visual pathway. RESULTS:Ten participants with MS and eight healthy controls were recruited between October 2008 and March 2009. Mesenchymal stem cells were successfully isolated, expanded and characterised in vitro for all participants in the treatment arm. CONCLUSIONS:In addition to determining the safety and feasibility of the intervention and informing design of future studies to address efficacy, MSCIMS adopts a novel strategy for testing neuroprotective agents in MS--the sentinel lesion approach--serving as proof of principle for its future wider applicability. TRIAL REGISTRATION:ClinicalTrials.gov (NCT00395200).

In vitro stem cell systems traditionally employ oxygen levels that are far removed from the in vivo situation. This study investigates whether an ambient environment containing a physiological oxygen level of 3% (normoxia) enables the generation of neural precursor cells (NPCs) from human embryonic stem cells (hESCs) and whether the resultant NPCs can undergo regional specification and functional maturation. We report robust and efficient neural conversion at 3% O 2 , demonstration of tri-lineage potential of resultant NPCs and the subsequent electrophysiological maturation of neurons. We also show that NPCs derived under 3% O 2 can be differentiated long term in the absence of neurotrophins and can be readily specified into both spinal motor neurons and midbrain dopaminergic neurons. Finally, modelling the oxygen stress that occurs during transplantation, we demonstrate that in vitro transfer of NPCs from a 20 to 3% O 2 environment results in significant cell death, while maintenance in 3% O 2 is protective. Together these findings support 3% O 2 as a physiologically relevant system to study stem cell-derived neuronal differentiation and function as well as to model neuronal injury.

Several diseases of the brain including Alzheimer's disease and Pick's disease are characterized by the presence of intracellular filamentous aggregates made of hyperphosphorylated microtubule-associated protein tau. Tau is involved in microtubule assembly, stabilization and axonal transport. Mutations in the Tau gene cause frontotemporal dementia and parkinsonism linked to chromosome 17, establishing that dysfunction of tau protein is sufficient to cause neurodegeneration and dementia. Several lines of transgenic mice have been produced that express human mutant tau. We have produced a mouse line expressing human P301S tau under the control of the murine Thy1 promoter. These mice exhibit the essential features of tauopathies, including neurodegeneration and abundant hyperphosphorylated tau deposits. We aimed to determine whether neural precursor cell transplantation could prevent tau-related nerve cell death in P301S tau transgenic mice. Cell-death was identified in specific areas of the cerebral cortex by immunoistochemistry with anti-NeuN antibody, cresyl violet staining and cell counting. Mouse neural precursor cells were transplanted to one hemisphere of 2-month-old P301S tau mice and the effect of transplantation on nerve cell death was examined 1 and 3 months later. RT-PCR and immunohistochemistry were used to identify mRNA and protein changes in both transplanted cells and the cortical area with neuronal cell death. We show that the transgenic human mutant P301S tau mouse line exhibits an age-related, progressive frontal cortical neuron loss, especially in the superficial layers, along with increased astrogliosis and that focal transplantation of neural precursor cells that differentiate into glia or direct astrocyte implantation results in neuroprotection. Neural precursor cell implantation, resulting in glial cell differentiation, leads to the rescue of cortical neurons in P301S tau transgenic mice. This indicates that early cell transplantation can rescue neurons from tau-related nerve cell death; it suggests a beneficial role for glial cell-based repair in neurodegenerative diseases.

Tau protein in a hyperphosphorylated state makes up the intracellular inclusions of several neurodegenerative diseases, including Alzheimer's disease and cases of frontotemporal dementia. Mutations in Tau cause familial forms of frontotemporal dementia, establishing that dysfunction of tau protein is sufficient to cause neurodegeneration and dementia. Transgenic mice expressing human mutant tau in neurons exhibit the essential features of tauopathies, including neurodegeneration and abundant filaments composed of hyperphosphorylated tau. Here we show that a previously described mouse line transgenic for human P301S tau exhibits an age-related, layer-specific loss of superficial cortical neurons, similar to what has been observed in human frontotemporal dementias. We also show that focal neural precursor cell implantation, resulting in glial cell differentiation, leads to the sustained rescue of cortical neurons. Together with evidence indicating that astrocyte transplantation may be neuroprotective, our findings suggest a beneficial role for glial cell-based repair in neurodegenerative diseases.

Treatment of early relapsing-remitting multiple sclerosis with the lymphocyte-depleting humanized monoclonal antibody alemtuzumab (Campath [registered trade mark]) significantly reduced the risk of relapse and accumulation of disability compared with interferon β-1a in a phase 2 trial [Coles et al., (Alemtuzumab vs. interferon β-1a in early multiple sclerosis. N Engl J Med 2008; 359: 1786-801)]. Patients treated with alemtuzumab experienced an improvement in disability at 6 months that was sustained for at least 3 years. In contrast, those treated with interferon β-1a steadily accumulated disability. Here, by post hoc subgroup analyses of the CAMMS223 trial, we show that among participants with no clinical disease activity immediately before treatment, or any clinical or radiological disease activity on-trial, disability improved after alemtuzumab but not following interferon β-1a. This suggests that disability improvement after alemtuzumab is not solely attributable to its anti-inflammatory effect. So we hypothesized that lymphocytes, reconstituting after alemtuzumab, permit or promote brain repair. Here we show that after alemtuzumab, and only when specifically stimulated with myelin basic protein, peripheral blood mononuclear cell cultures produced increased concentrations of brain-derived neurotrophic factor, platelet-derived growth factor and ciliary neurotrophic factor. Analysis by reverse transcriptase polymerase chain reaction of cell separations showed that the increased production of ciliary neurotrophic factor and brain-derived neurotrophic factor after alemtuzumab is attributable to increased production by T cells. Media from these post-alemtuzumab peripheral blood mononuclear cell cultures promoted survival of rat neurones and increased axonal length in vitro, effects that were partially reversed by neutralizing antibodies against brain-derived nerve growth factor and ciliary neurotrophic factor. This conditioned media also enhanced oligodendrocyte precursor cell survival, maturation and myelination. Taken together, the clinical analyses and laboratory findings support the interpretation that improvement in disability after alemtuzumab may result, in part, from neuroprotection associated with increased lymphocytic delivery of neurotrophins to the central nervous system.

Following an injury to the dorsal roots primary sensory afferents fail to regenerate past the hostile dorsal root entry zone (DREZ), the interface between the peripheral and central nervous system. Neural progenitor cells have previously been utilised as a cellular replacement therapy in a variety of CNS injury models. Here we show for the first time that NPCs are capable of promoting neurite outgrowth from adult sensory neurons in vitro and ex vivo cryo-cultures. The effectiveness of NPCs as a potential means of promoting regeneration of primary afferents across the DREZ was assessed following rhizotomy at the cervical level in the adult rat. Adult rats were subjected to rhizotomy of the dorsal roots between C(5)-T(1) which were then reanastamosed. In conjunction with the rhizotomy NPCs were delivered at the DREZ. NPCs survived transplantation and were observed to differentiate predominantly into glia. Regeneration of the dorsal root fibers was assessed with immunhistochemical analysis of the large and small diameter peptidergic and non-peptidergic afferents. Although afferents appeared near to the DREZ there was little regeneration beyond the DREZ. Furthermore, no significant improvement was observed in behavioural tasks.

Perinatal white matter injury, or periventricular leukomalacia (PVL), is the most common cause of brain injury in premature infants and is the leading cause of cerebral palsy. Despite increasing numbers of surviving extreme premature infants and associated long-term neurological morbidity, our understanding and treatment of PVL remains incomplete. Inflammation- or ischemia/hypoxia-based rodent models, although immensely valuable, are largely restricted to reproducing short-term features of up to 3 weeks after injury. Given the long-term sequelae of PVL, there is a need for subchronic models that will enable testing of putative neuroprotective therapies. Here, we report long term characterization of a neonatal inflammation-induced rat model of PVL. We show bilateral ventriculomegaly, inflammation, reactive astrogliosis, injury to pre-oligodendrocytes, and neuronal loss 8 weeks after injury. We demonstrate neuroprotective effects of oligodendrocyte precursor cell transplantation. Our findings present a subchronic model of PVL and highlight the tissue protective effects of oligodendrocyte precursor cell transplants that demonstrate the potential of cell-based therapy for PVL.

The adult bone marrow contains a population of multipotent mesenchymal stromal cells (MSCs), defined by plastic adherence, expression of stromal cell surface markers, and differentiation into mesenchymal lineages. There has been much interest in the possible therapeutic use of MSCs in the treatment of demyelinating diseases of the central nervous system. One therapeutic possibility is that these cells may be able to remyelinate when directly injected into the demyelinated spinal cord. Here we examine the effects of direct transplantation of green fluorescent protein (GFP)-labeled MSCs into a model of focal spinal cord demyelination induced by ethidium bromide. We demonstrate that direct intralesional injection of undifferentiated MSCs does not lead to remyelination. Furthermore, we report that transplanted MSCs migrate into areas of normal tissue, deposit collagen, and are associated with axonal damage. These findings support the need for further experimental evaluation of the safety and efficacy of direct parenchymal injection of MSCs into demyelinated lesions and highlight an important issue regarding potential clinical consequences of culture heterogeneity of MSCs between centers.

Human embryonic stem cells (hESCs) are a potential source of defined tissue for cell-based therapies in regenerative neurology. In order for this potential to be realized, there is a need for the evaluation of the behaviour of human embryonic stem cell-derived neural stem cells (hES-NSCs) both in the normal and the injured CNS. Using normal tissue and two experimental models, we examined the response of clinically compatible hES-NSCs to physiological and pathological signals. We demonstrate that the phenotypic potential of a multipotent population of hES-NSCs is influenced by these cues both in vitro and in vivo. hES-NSCs display a temporal profile of neurogenic and gliogenic differentiation, with the generation of mature neurons and glia over 4 weeks in vitro, and 20 weeks in the uninjured rodent brain. However, transplantation into the pathological CNS accelerates maturation and polarizes hES-NSC differentiation potential. This study highlights the role of environmental signals in determining both lineage commitment and temporal maturation of human neural stem cells. Controlled manipulation of environmental signals appropriate to the pathological specificity of the targeted disease will be necessary in the design of therapeutic stem cell-based strategies.

UNLABELLED:Endogenous repair after injury in the adult CNS is limited by a number of factors including cellular loss, inflammation, cavitation and glial scarring. Spinal cord neural progenitor cells (SCNPCs) may provide a valuable cellular source for promoting repair following spinal cord injury. SCNPCs are multipotent, can be expanded in vitro, have the capacity to differentiate into CNS cell lineages and are capable of long-term survival following transplantation. AIMS & METHOD:To determine the extent to which SCNPCs may contribute to spinal cord repair SCNPCs isolated from rat fetal spinal cord were expanded ex vivo and transplanted into the adult rat spinal cord after a dorsal column crush lesion. RESULTS:The survival and distribution of transplanted cells were examined at 24 h, 1, 2 and 6 weeks after injury. Transplanted cells were identified at all time points, located mainly at the lesion perimeter, indicating good post-transplant cell survival. Furthermore, SCNPCs maintained their ability to differentiate in vivo, with approximately 40% differentiating into cells with a glial morphology, whilst 8% displayed a neural morphology. Transplanted animals were also assessed on a number of behavioral tasks measuring sensorimotor and proprioceptive function to determine the extent to which SCNPC transplants might attenuate lesion-induced functional deficits. SCNPCs failed to promote significant functional recovery, with a small improvement observed in only one of the four tasks employed, primarily related to improvements in sensory function. Tracing of the corticospinal tract and ascending dorsal column pathway revealed no regeneration of the axons beyond the lesion site. CONCLUSIONS:These data indicate that, although transplanted SCNPCs show good survival in the spinal cord injury environment, combination with other treatment strategies is likely to be required for these cells to fully exert their therapeutic potential.

Oligodendrocyte precursor cells (OPCs) are widely regarded as the best characterized cell population in the mammalian CNS and until recently were believed to be a lineage-restricted precursor terminally differentiating to postmitotic oligodendrocytes. Recent evidence has suggested that OPCs may have in vitro and in vivo neuronal potential. In this report we examine the differentiation potential of cortical OPC populations following transplantation into the neurogenic environment of the intact neonatal and adult hippocampus. Donor OPCs were minimally manipulated and not subjected to long-term ex vivo manipulation such as expansion or treatment with mitogens. Minimally manipulated OPCs did not exhibit any intrinsic neuronal potential in vitro prior to transplantation. Following transplantation of GFP-OPCs into intact neonatal and adult hippocampus, cells were able to survive and integrate for at least 14 weeks but did not exhibit neuronal differentiation. Induction of a focal neurotoxic lesion also did not result in neuronal differentiation of graft-derived OPCs. These findings show that unselected and unmanipulated populations of cortical OPCs remain as precursor cells, commit to the oligodendrocyte lineage and fail to respond to the extrinsic cues of a neurogenic or injured environment.

The myelin surrounding axons is highly organized, with ion channels and adaptor molecules densely clustered at nodes of Ranvier. Loss of myelin decreases impulse conduction and ultimately leads to degeneration of axons. Demyelinated axons also have disrupted organization, with loss of K+ channels

By culturing neural progenitor cells in the presence of retinoid receptor agonists, we have defined the components of the retinoid-signalling pathway that are important for the birth and maintenance of neuronal cells. We provide evidence that depending on the order and combination of retinoid receptors activated, different neuronal cells are obtained. Astrocytes and oligodendrocytes are predominantly formed in the presence of activated retinoic acid receptor (RAR) α, whereas motoneurons are formed when RARβ is activated. We have looked at the regulation of two transcription factors islet-1/2 which are involved in neuronal development. We find that activated RARβ up-regulates islet-1 expression, whereas activation of RARα can either act in combination with RARβ signalling to maintain islet-1 expression or induce islet-2 expression in the absence of activated RARβ. RARγ cannot directly regulate islet-1/2 but can down-regulate RARβ expression, which results in loss of islet-1 expression. We finally show that activated RARα is one of the final steps required for a mature motoneuron phenotype.

Neurodegenerative disorders and traumatic brain injury result in the loss of specific neuronal populations. Stem cells are self-renewing, multi- or pluripotent cells capable of differentiating into a wide range of cell types, properties which make stem cells a potentially invaluable source of transplantable cells. Recent experimental studies have indicated that several stem cell populations have the ability to replace lost neurons and to repair the damaged nervous system following transplantation. This review evaluates the potential of various stem cell populations in the treatment of human neurodegenerative conditions and traumatic brain injury.