There is interest in therapeutic approaches to improve stroke injuries beyond the current 4.5 hour window of opportunity available for tPA treatment. We are interested in determining if neural progenitor cells can provide protection one week out from a stroke injury, and if the location of the cells relative to the core or penumbra of the injury site is important. Methods: We used a unilateral permanent middle cerebral artery ligation (MCAL) stroke injury model in C57B6 mice (male 3 months old) that generates a modest cortical lesion with limited subcortical injury. A single 3 ul Stereotactic injection was made either in the stroke core, or outside the injury site penumbra under the ipsilateral hippocampus. We injected 100,000 neural progenitor cells (iNPs) differentiated from induced pluripotent stem cells derived from human fibroblasts. Previous work showed that injection in a matrix enhanced iNP survival, therefore, the cells were injected either in a hydrogel matrix (n=4) or in matrigel (n=5) one week post-MCAL. Hydrogel (n=5) and Matrigel (n=4) only control groups were also used. Mice were sacrificed three weeks post-MCAL (two weeks post-transplant) and evaluated for lesion size as a percent of cortex. Results: Mice that received stem cells placed into the core of the lesion, regardless of the specific matrix, had significantly smaller cortical lesions (p=0.0021) than those who did not receive stem cells (hydrogel alone control 36.0% mean cortical lesion (10.7% SD); Hydrogel + iNP cells 15.6% (3.1%); Matrigel alone control 30.4% (10.0%), matrigel + iNP cells 19.4% (8.6%). There was no difference between the controls: saline only, matrigel only, or hydrogel only. Using the same approach a second set of mice received 100,000 iNP cells outside of the lesion site, below the ipsilateral hippocampus (matrigel + iNPs (n=9), matrigel alone (n=9), or saline (n=8). In contrast there was no reduction in lesion size. Conclusion: This suggests that iNPs in the region of greatest injury can provide protection even one week following stroke, but that these effects are spatially limited. Future studies will focus on aged male and female populations.

INTRODUCTION:Due to the publicity about stem cell transplantation for the treatment of cerebral palsy, many families seek information on treatment, and many travel overseas for cell transplantation. Even so, there is little scientific confirmation of benefit, and therefore existing knowledge in the field must be summarized. AREAS COVERED:This paper addresses the clinical protocols examining the problem, types of stem cells available for transplant, experimental models used to test the benefit of the cells, possible mechanisms of action, potential complications of cell treatment and what is needed in the field to help accelerate cell-based therapies. EXPERT OPINION:While stem cells may be beneficial in acute injuries of the CNS the biology of stem cells is not well enough understood in chronic injuries or disorders such as cerebral palsy. More work is required at the basic level of stem cell biology, in the development of animal models, and finally in well-conceived clinical trials.

Autophagy is a highly regulated, catabolic process, through which macromolecules such as damaged organelles and proteins are degraded and recycled to maintain cellular homeostasis.Various studies have shown the role of autophagy activation in the brain cells such as astrocytes, microglia, neurons and capillary endothelial cells upon an ischaemic insult.The underlying mechanism and the role of autophagy in ischaemic stroke, however, are yet to be fully elucidated.Recent studies have suggested that insufficient or excessive autophagy results in nerve damage and cell death whereas mild/moderate autophagy has a neuroprotective effect.It has been proposed that autophagy may be a therapeutic target in stroke treatment; however, there is a lot of debate as to whether induction or diminution of autophagy plays a role in neuronal survival after cerebral ischaemia and indicate that it has a dual role depending on the time of induction of autophagy.This review has summarized the role of autophagy in ischaemic stroke and explored effects of pharmacological autophagy modulators in ischaemic stroke treatment.Further studies are needed for translating the potential therapeutic approach in stroke treatment aiming at characterizing the timing, amount and specificity of the autophagy modulation.

To determine the potential for using stem cells in the treatment of ischemic injuries of the central nervous sys-tem, clinically relevant experiments were performed administering human adult adherent bone marrow derived stem cells into a rat model of ischemic stroke. Variables such as the requirement for immunosuppression, route of cell administration, window for therapeutic benefit and optimal cell dosage have all been examined in a series of relevant translational experiments in animals undergoing middle cerebral artery ligation stroke injury. Animals were tested for improvements in locomotor and neurological function at time points as late as 6 months post-cell transplantation and demonstrated sustained statistically significant benefit from a single dose of cells. Following sacrifice, immunohistochemistry was performed on tissues to determine stem cell engraftment and fate, as well as neuroprotection of endogenous tissue at the sites of ischemic injury. The results indicate locomotor and neu-rological improvement correlates with improved neuroprotection and limited retention of the transplanted stem cells, with limited neuronal fate. The observed benefit occurs in a cell dosage dependent fashion suggesting a pharmacological role for the cells when administered intravenously in these injury models.

Once hypoxic-ischemic ( HI) injury ensues in the human neonate at birth, the resulting brain damage lasts throughout the individual's lifetime, as no ameliorative treatments are currently available. We have recently shown that intracerebral transplantation of multipotent adult progenitor cells (MAPCs) results in behavioral improvement and reduction in ischemic cell loss in neonatal rat HI-injury model. In an attempt to advance this cellular therapy to the clinic, we explored the more practical and less invasive intravenous administration of MAPCs. Seven-day-old Sprague-Dawley rats were initially subjected to unilateral HI injury, then 7 days later received intracerebral or intravenous injections of allogeneic rat MAPCs. On post-transplantation days 7 and 14, the animals that received MAPCs via the intracerebral or intravenous route exhibited improved motor and neurologic scores compared with those that received vehicle infusion alone. Immunohistochemical evaluations at day 14 after transplantation revealed that both intracerebrally and intravenously transplanted MAPCs were detected in the ischemic hippocampal area. The degree of hippocampal cell preservation was almost the same in the two treatment groups and greater than that in the vehicle group. These results show that intravenous delivery of MAPCs is a feasible and efficacious cell therapy with potential for clinical use.

Adult stem cell therapy has been proposed for brain injury in young children. While there have been no clinical trials in the US, the therapy is widely advertised and anecdotally reported in multiple internet sources, leading families to seek the treatment in uncontrolled circumstances. The purpose of this review is to present a discussion of the various types of stem cell preparations, with emphasis on adult stem cells, the scientific basis of their development, and the available experimental evidence for their utility in childhood brain injury. We will also provide background information on the biologic events occurring in injured immature brain, as they relate to the transplantation of stem cells. We will then review our own data from neonatal rodent studies with experimental hypoxic-ischemic brain injury. We have shown that early intracerebral administration promotes improved behavioral outcome in the animals, the formation of new neurons, and the preservation of intrinsic cells. New experiments demonstrate the equality of intracerebral and intravenous transplantation in acute neonatal hypoxic-ischemic injury in rodent. We will speculate on the possible clinical uses of adult stem cells. Our current impression is that the cells have the greatest potential for success when administered soon after an injury. What needs to be done to further the field? The different types of cell preparations should be tested against each other in experimental situations. A suitable model of chronic brain injury should be utilized for evaluating the benefit of the cells for this purpose. Long term safety of the cells should be confirmed in animal models. Finally, multicenter clinical trials should be conducted in highly controlled protocols.

L'invention concerne le traitement de plusieurs lesions, troubles, dysfonctionnements, maladies, et analogues, du cerveau a l'aide de MAPC, en particulier, dans des modes de realisation, l'invention concerne le traitement desdits etats resultant d'une hypoxie, notamment ceux causes par l'hypoxie systemique et ceux causes par une irrigation sanguine insuffisante. Dans d'autres modes de realisation particuliers, l'invention concerne, par exemple, le traitement d'une lesion cerebrale ischemique hypoxique a l'aide de MPAC, chez les enfants par exemple, ainsi que le traitement d'infarctus corticaux et d'accidents cerebrovasculaires a l'aide de MAPC chez les adultes, par exemple.

Children born with hypoxic-ischemic (HI) brain injury account for a significant number of live births wherein no clinical treatment is available. Limited clinical trials of stem cell therapy have been initiated in a number of neurological disorders, but the preclinical evidence of a cell-based therapy for neonatal HI injury remains in its infancy. One major postulated mechanism underlying therapeutic benefits of stem cell therapy involves stimulation of endogenous neurogenesis via transplantation of exogenous stem cells. To this end, transplantation has targeted neurogenic sites, such as the hippocampus, for brain protection and repair. The hippocampus has been shown to secrete growth factors, especially during the postnatal period, suggesting that this brain region presents as highly conducive microenvironment for cell survival. Based on its neurogenic and neurotrophic factor-secreting features, the hippocampus stands as an appealing target for stem cell therapy. Here, we investigated the efficacy of intrahippocampal transplantation of multipotent progenitor cells (MPCs), which are pluripotent progenitor cells with the ability to differentiate into a neuronal lineage. Seven-day-old Sprague-Dawley rats were initially subjected to unilateral HI injury, which involved permanent ligation of the right common carotid artery and subsequent exposure to hypoxic environment. At day 7 after HI injury, animals received stereotaxic hippocampal injections of vehicle or cryopreserved MPCs (thawed just prior to transplantation) derived either from Sprague-Dawley rats (syngeneic) or Fisher rats (allogeneic). All animals were treated with daily immunosuppression throughout the survival period. Behavioral tests were conducted on posttransplantation days 7 and 14 using the elevated body swing test and the rotarod to reveal general and coordinated motor functions. MPC transplanted animals exhibited reduced motor asymmetry and longer time spent on the rotarod than those that received the vehicle infusion. Both syngeneic and allogeneic MPC transplanted injured animals did not significantly differ in their behavioral improvements at both test periods. Immunohistochemical evaluations of graft survival after behavioral testing at day 14 posttransplantation revealed that syngeneic and allogeneic transplanted MPCs survived in the hippocampal region. These results demonstrate for the first time that transplantation of MPCs ameliorated motor deficits associated with HI injury. In view of comparable behavioral recovery produced by syngeneic and allogeneic MPC grafts, allogeneic transplantation poses as a feasible and efficacious cell replacement strategy with direct clinical application. An equally major finding is the observation lending support to the hippocampus as an excellent target brain region for stem cell therapy in treating HI injury.

Background Stromal cell-derived factor 1 (SDF-1 or CXCL12) is chemotaxic for CXCR4 expressing bone marrow-derived cells. It functions in brain embryonic development and in response to ischemic injury in helping guide neuroblast migration and vasculogenesis. In experimental adult stroke models SDF-1 is expressed perivascularly in the injured region up to 30 days after the injury, suggesting it could be a therapeutic target for tissue repair strategies. We hypothesized that SDF-1 would be expressed in similar temporal and spatial patterns following hypoxic-ischemic (HI) injury in neonatal brain. Results Twenty-five 7-day-old C57BL/J mice underwent HI injury. SDF-1 expression was up regulated up to 7 days after the injury but not at the later time points. The chief sites of SDF-1 up regulation were astrocytes, their foot processes along blood vessels and endothelial cells. Conclusion The localization of SDF-1 along blood vessels in the HI injury zone suggests that these perivascular areas are where chemotaxic signaling for cellular recruitment originates and that reactive astrocytes are major mediators of this process. The associated endothelium is likely to be the site for vascular attachment and diapedesis of CXCR4 receptor expressing cells to enter the injured tissue. Here we show that, relative to adults, neonates have a significantly smaller window of opportunity for SDF-1 based vascular chemotaxic recruitment of bone marrow-derived cells. Therefore, without modification, following neonatal HI injury there is only a narrow period of time for endogenous SDF-1 mediated chemotaxis and recruitment of reparative cells, including exogenously administered stem/progenitor cells.

n the past 5 years, accumulating evidence has demonstrated plasticity of bone marrowderived cells.Bone marrow-derived cells display the capacity to change their fate, differentiating into hepatocytes, endothelial cells, muscle cells, and cardiomyocytes, and even neurons. 1The findings that bone marrow cells differentiate into neurons in vitro and in vivo challenge previous assumptions that tissue-specific stem cells give rise only to cells of their organ of origin and do not cross lineages.

The chemokine stromal-derived factor-1 (SDF-1, also known as CXCL12) and its receptor CXCR4 have been implicated in homing of stem cells to the bone marrow and the homing of bone marrow-derived cells to sites of injury. Bone marrow cells infiltrate brain and give rise to long-term resident cells following injury. Therefore, SDF-1 and CXCR4 expression patterns in 40 mice were examined relative to the homing of bone marrow-derived cells to sites of ischemic injury using a stroke model. Mice received bone marrow transplants from green fluorescent protein (GFP) transgenic donors and later underwent a temporary middle cerebral artery suture occlusion (MCAo). SDF-1 was associated with blood vessels and cellular profiles by 24 hours through at least 30 days post-MCAo. SDF-1 expression was principally localized to the ischemic penumbra. The majority of SDF-1 expression was associated with reactive astrocytes; much of this was perivascular. GFP+ cells were associated with SDF-1-positive vessels and were also found in the neuropil of regions with increased SDF-1 immunoreactivity. Most vessel-associated GFP+ cells resemble pericytes or perivascular microglia and the majority of the GFP+ cells in the parenchyma displayed characteristics of activated microglial cells. These findings suggest SDF-1 is important in the homing of bone marrow-derived cells, especially monocytes, to areas of ischemic injury.

Cerebral palsy is a group of brain diseases which produce chronic motor disability in children. The causes are quite varied and range from abnormalities of brain development to birth-related injuries to postnatal brain injuries. Due to the increased survival of very premature infants, the incidence of cerebral palsy may be increasing. While premature infants and term infants who have suffered neonatal hypoxic-ischaemic (HI) injury represent only a minority of the total cerebral palsy population, this group demonstrates easily identifiable clinical findings, and much of their injury is to oligodendrocytes and the cerebral white matter. While the use of stem cell therapy is promising, there are no controlled trials in humans with cerebral palsy and only a few trials in patients with other neurologic disorders. However, studies in animals with experimentally induced strokes or traumatic injuries have indicated that benefit is possible. The potential to do these transplants via injection into the vasculature rather than directly into the brain increases the likelihood of timely human studies. As a result, variables appropriate to human experiments with intravascular injection of cells, such as cell type, timing of the transplant and effect on function, need to be systematically performed in animal models with HI injury, with the hope of rapidly translating these experiments to human trials.