Hypocretinergic and cholinergic contributions to sleep-wake disturbances in a mouse model of traumatic brain injury.

Disorders of sleep and wakefulness occur in the majority of individuals who have experienced traumatic brain injury (TBI), with increased sleep need and excessive daytime sleepiness often reported. Behavioral and pharmacological therapies have limited efficacy, in part, because the etiology of post-TBI sleep disturbances is not well understood. Severity of injuries resulting from head trauma in humans is highly variable, and as a consequence so are their sequelae. Here, we use a controlled laboratory model to investigate the effects of TBI on sleep-wake behavior and on candidate neurotransmitter systems as potential mediators. We focus on hypocretin and melanin-concentrating hormone (MCH), hypothalamic neuropeptides important for regulating sleep and wakefulness, and two potential downstream effectors of hypocretin actions, histamine and acetylcholine. Adult male C57BL/6 mice (n=6-10/group) were implanted with EEG recording electrodes and baseline recordings were obtained. After baseline recordings, controlled cortical impact was used to induce mild or moderate TBI. EEG recordings were obtained from the same animals at 7 and 15 days post-surgery. Separate groups of animals (n=6-8/group) were used to determine effects of TBI on the numbers of hypocretin and MCH-producing neurons in the hypothalamus, histaminergic neurons in the tuberomammillary nucleus, and cholinergic neurons in the basal forebrain. At 15 days post-TBI, wakefulness was decreased and NREM sleep was increased during the dark period in moderately injured animals. There were no differences between groups in REM sleep time, nor were there differences between groups in sleep during the light period. TBI effects on hypocretin and cholinergic neurons were such that more severe injury resulted in fewer cells. Numbers of MCH neurons and histaminergic neurons were not altered under the conditions of this study. Thus, we conclude that moderate TBI in mice reduces wakefulness and increases NREM sleep during the dark period, effects that may be mediated by hypocretin-producing neurons and/or downstream cholinergic effectors in the basal forebrain.

Time-dependent effects of CX3CR1 in a mouse model of mild traumatic brain injury.

BACKGROUND: Neuroinflammation is an important secondary mechanism that is a key mediator of the long-term consequences of neuronal injury that occur in traumatic brain injury (TBI). Microglia are highly plastic cells with dual roles in neuronal injury and recovery. Recent studies suggest that the chemokine fractalkine (CX3CL1, FKN) mediates neural/microglial interactions via its sole receptor CX3CR1. CX3CL1/CX3CR1 signaling modulates microglia activation, and depending upon the type and time of injury, either protects or exacerbates neurological diseases. METHODS: In this study, mice deficient in CX3CR1 were subjected to mild controlled cortical impact injury (CCI), a model of TBI. We evaluated the effects of genetic deletion of CX3CR1 on histopathology, cell death/survival, microglia activation, and cognitive function for 30 days post-injury. RESULTS: During the acute post-injury period (24 h-15 days), motor deficits, cell death, and neuronal cell loss were more profound in injured wild-type than in CX3CR1(-/-) mice. In contrast, during the chronic period of 30 days post-TBI, injured CX3CR1(-/-) mice exhibited greater cognitive dysfunction and increased neuronal death than wild-type mice. The protective and deleterious effects of CX3CR1 were associated with changes in microglia phenotypes; during the acute phase CX3CR1(-/-) mice showed a predominant anti-inflammatory M2 microglial response, with increased expression of Ym1, CD206, and TGFß. In contrast, increased M1 phenotypic microglia markers, Marco, and CD68 were predominant at 30 days post-TBI. CONCLUSION: Collectively, these novel data demonstrate a time-dependent role for CX3CL1/CX3CR1 signaling after TBI and suggest that the acute and chronic responses to mild TBI are modulated in part by distinct microglia phenotypes.

The role of microglia in adult hippocampal neurogenesis.

Our view of microglia has dramatically changed in the last decade. From cells being "silent" in the healthy brain, microglia have emerged to be actively involved in several brain physiological functions including adult hippocampal neurogenesis, and cognitive and behavioral function. In light of recent discoveries revealing a role of microglia as important effectors of neuronal circuit reorganization, considerable attention has been focused on how microglia and hippocampal neurogenesis could be an interdependent phenomenon. In this review the role of microglia in the adult hippocampal neurogenesis under physiological condition is discussed.

Fractalkine overexpression suppresses tau pathology in a mouse model of tauopathy.

Alzheimer's disease is characterized by amyloid plaques, neurofibrillary tangles, glial activation, and neurodegeneration. In mouse models, inflammatory activation of microglia accelerates tau pathology. The chemokine fractalkine serves as an endogenous neuronal modulator to quell microglial activation. Experiments with fractalkine receptor null mice suggest that fractalkine signaling diminishes tau pathology, but exacerbates amyloid pathology. Consistent with this outcome, we report here that soluble fractalkine overexpression using adeno-associated viral vectors significantly reduced tau pathology in the rTg4510 mouse model of tau deposition. Furthermore, this treatment reduced microglial activation and appeared to prevent neurodegeneration normally found in this model. However, in contrast to studies with fractalkine receptor null mice, parallel studies in an APP/PS1 model found no effect of increased fractalkine signaling on amyloid deposition. These data argue that agonism at fractalkine receptors might be an excellent target for therapeutic intervention in tauopathies, including those associated with amyloid deposition.

A single administration of human umbilical cord blood T cells produces long-lasting effects in the aging hippocampus.

Neurogenesis occurs throughout life but significantly decreases with age. Human umbilical cord blood mononuclear cells (HUCB MNCs) have been shown to increase the proliferation of neural stem cells (NSCs) in the dentate gyrus (DG) of the hippocampus and the subgranular zone of aging rats (Bachstetter et al., BMC Neurosci 9:22, 2008), but it is unclear which fraction or combination of the HUCB MNCs are responsible for neurogenesis. To address this issue, we examined the ability of HUCB MNCs, CD4+, CD8+, CD3+, CD14+, and CD133+ subpopulations to increase proliferation of NSCs both in vitro and in vivo. NSCs were first grown in conditioned media generated from HUCB cultures, and survival and proliferation of NSC were determined with the fluorescein diacetate/propidium iodide and 5-bromo-2'-deoxyuridine incorporation assays, respectively. In a second study, we injected HUCB cells intravenously in young and aged Fisher 344 rats and examined proliferation in the DG at 1 week (study 2.1) and 2 weeks (study 2.2) postinjection. The effects of the HUCB MNC fractions on dendritic spine density and microglial activation were also assessed. HUCB T cells (CD3+, CD4+, and CD8+ cells) induced proliferation of NSCs (p < 0.001) and increased cell survival. In vivo, HUCB-derived CD4+ cells increased NSC proliferation at both 1 and 2 weeks while also enhancing the density of dendritic spines at 1 week and decreasing inflammation at 2 weeks postinjection. Collectively, these data indicate that a single injection of HUCB-derived T cells induces long-lasting effects and may therefore have tremendous potential to improve aging neurogenesis.

The soluble isoform of CX3CL1 is necessary for neuroprotection in a mouse model of Parkinson's disease.

The chemokine CX3CL1/fractalkine is expressed by neurons as a transmembrane-anchored protein that can be cleaved to yield a soluble isoform. However, the roles for these two types of endogenous CX3CL1 in neurodegenerative pathophysiology remain elusive. As such, it has been difficult to delineate the function of the two isoforms of CX3CL1, as both are natively present in the brain. In this study we examined each isoform's ability to regulate neuroinflammation in a mouse model of Parkinson's disease initiated by the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We were able to delineate the function of both CX3CL1 isoforms by using adeno-associated virus-mediated gene therapy to selectively express synthetic variants of CX3CL1 that remain either permanently soluble or membrane bound. In the present study we injected each CX3CL1 variant or a GFP-expressing vector directly into the substantia nigra of CX3CL1(-/-) mice. Our results show that only the soluble isoform of CX3CL1 is sufficient for neuroprotection after exposure to MPTP. Specifically, we show that the soluble CX3CL1 isoform reduces impairment of motor coordination, decreases dopaminergic neuron loss, and ameliorates microglial activation and proinflammatory cytokine release resulting from MPTP exposure. Furthermore, we show that the membrane-bound isoform provides no neuroprotective capability to MPTP-induced pathologies, exhibiting similar motor coordination impairment, dopaminergic neuron loss, and inflammatory phenotypes as MPTP-treated CX3CL1(-/-) mice, which received the GFP-expressing control vector. Our results reveal that the neuroprotective capacity of CX3CL1 resides solely upon the soluble isoform in an MPTP-induced model of Parkinson's disease.

CX3CR1 deficiency leads to impairment of hippocampal cognitive function and synaptic plasticity.

The protective/neurotoxic role of fractalkine (CX3CL1) and its receptor CX3C chemokine receptor 1 (CX3CR1) signaling in neurodegenerative disease is an intricate and highly debated research topic and it is becoming even more complicated as new studies reveal discordant results. It appears that the CX3CL1/CX3CR1 axis plays a direct role in neurodegeneration and/or neuroprotection depending on the CNS insult. However, all the above studies focused on the role of CX3CL1/CX3CR1 signaling in pathological conditions, ignoring the relevance of CX3CL1/CX3CR1 signaling under physiological conditions. No approach to date has been taken to decipher the significance of defects in CX3CL1/CX3CR1 signaling in physiological condition. In the present study we used CX3CR1⁻/⁻, CX3CR1⁺/⁻, and wild-type mice to investigate the physiological role of CX3CR1 receptor in cognition and synaptic plasticity. Our results demonstrate for the first time that mice lacking the CX3CR1 receptor show contextual fear conditioning and Morris water maze deficits. CX3CR1 deficiency also affects motor learning. Importantly, mice lacking the receptor have a significant impairment in long-term potentiation (LTP). Infusion with IL-1ß receptor antagonist significantly reversed the deficit in cognitive function and impairment in LTP. Our results reveal that under physiological conditions, disruption in CX3CL1 signaling will lead to impairment in cognitive function and synaptic plasticity via increased action of IL-1ß.

CX3CL1 reduces neurotoxicity and microglial activation in a rat model of Parkinson's disease.

BACKGROUND: Parkinson's disease is characterized by a progressive loss of dopaminergic neurons in the substantia nigra. The cause of the neurodegeneration is unknown. Neuroinflammation has been clearly shown in Parkinson's disease and may be involved in the progressive nature of the disease. Microglia are capable of producing neuronal damage through the production of bioactive molecules such as cytokines, as well as reactive oxygen species (ROS), and nitric oxide (NO). The inflammatory response in the brain is tightly regulated at multiple levels. One form of immune regulation occurs via neurons. Fractalkine (CX3CL1), produced by neurons, suppresses the activation of microglia. CX3CL1 is constitutively expressed. It is not known if addition of exogenous CX3CL1 beyond otherwise physiologically normal levels could decrease microglia activation and thereby minimize the secondary neurodegeneration following a neurotoxic insult. METHODS: The intrastriatal 6-hydroxydopamine (6-OHDA) rat model of Parkinson disease, was used to test the hypothesis that exogenous CX3CL1 could be neuroprotective. Treatment with recombinant CX3CL1 was delivered to the striatum by an osmotic minipump for 28 days beginning 7 days after the initial insult. Unbiased stereological methods were used to quantify the lesion size in the striatum, the amount of neuronal loss in the substantia nigra, and the amount of microglia activation. RESULTS: As hypothesized, CX3CL1 was able to suppress this microglia activation. The reduced microglia activation was found to be neuroprotective as the CX3CL1 treated rats had a smaller lesion volume in the striatum and importantly significantly fewer neurons were lost in the CX3CL1 treated rats. CONCLUSION: These findings demonstrated that CX3CL1 plays a neuroprotective role in 6-OHDA-induced dopaminergic lesion and it might be an effective therapeutic target for many neurodegenerative diseases, including Parkinson disease and Alzheimer disease, where inflammation plays an important role.

CD45 deficiency drives amyloid-ß peptide oligomers and neuronal loss in Alzheimer's disease mice.

Converging lines of evidence indicate dysregulation of the key immunoregulatory molecule CD45 (also known as leukocyte common antigen) in Alzheimer's disease (AD). We report that transgenic mice overproducing amyloid-ß peptide (Aß) but deficient in CD45 (PSAPP/CD45(-/-) mice) faithfully recapitulate AD neuropathology. Specifically, we find increased abundance of cerebral intracellular and extracellular soluble oligomeric and insoluble Aß, decreased plasma soluble Aß, increased abundance of microglial neurotoxic cytokines tumor necrosis factor-α and interleukin-1ß, and neuronal loss in PSAPP/CD45(-/-) mice compared with CD45-sufficient PSAPP littermates (bearing mutant human amyloid precursor protein and mutant human presenilin-1 transgenes). After CD45 ablation, in vitro and in vivo studies demonstrate an anti-Aß phagocytic but proinflammatory microglial phenotype. This form of microglial activation occurs with elevated Aß oligomers and neural injury and loss as determined by decreased ratio of anti-apoptotic Bcl-xL to proapoptotic Bax, increased activated caspase-3, mitochondrial dysfunction, and loss of cortical neurons in PSAPP/CD45(-/-) mice. These data show that deficiency in CD45 activity leads to brain accumulation of neurotoxic Aß oligomers and validate CD45-mediated microglial clearance of oligomeric Aß as a novel AD therapeutic target.

Fractalkine and CX 3 CR1 regulate hippocampal neurogenesis in adult and aged rats.

Microglia have neuroprotective capacities, yet chronic activation can promote neurotoxic inflammation. Neuronal fractalkine (FKN), acting on CX(3)CR1, has been shown to suppress excessive microglia activation. We found that disruption in FKN/CX(3)CR1 signaling in young adult rodents decreased survival and proliferation of neural progenitor cells through IL-1ß. Aged rats were found to have decreased levels of hippocampal FKN protein; moreover, interruption of CX(3)CR1 function in these animals did not affect neurogenesis. The age-related loss of FKN could be restored by exogenous FKN reversing the age-related decrease in hippocampal neurogenesis. There were no measureable changes in young animals by the addition of exogenous FKN. The results suggest that FKN/CX(3)CR1 signaling has a regulatory role in modulating hippocampal neurogenesis via mechanisms that involve indirect modification of the niche environment. As elevated neuroinflammation is associated with many age-related neurodegenerative diseases, enhancing FKN/CX(3)CR1 interactions could provide an alternative therapeutic approach to slow age-related neurodegeneration.

NT-020, a natural therapeutic approach to optimize spatial memory performance and increase neural progenitor cell proliferation and decrease inflammation in the aged rat.

The process of aging is linked to oxidative stress, microglial activation, and proinflammatory factors, which are known to decrease cell proliferation and limit neuroplasticity. These factors may lead the transition from normal aging to more severe cognitive dysfunction associated with neurodegenerative diseases. We have shown that natural compounds such as polyphenols from blueberry and green tea and amino acids like carnosine are high in antioxidant and antiinflammatory activity that decreases the damaging effects of reactive oxygen species (ROS), in the blood, brain, and other tissues of the body. Furthermore, we have shown that the combination of these nutrients (called NT-020) creates a synergistic effect that promotes the proliferation of stem cells in vitro and in vivo. In the current study, we examined the effects of NT-020 on neurogenesis and performance on a Morris water maze (MWM). Aged (20-month-old) male Fischer 344 rats were treated with 135.0 mg/kg per day (n = 13) of NT-020. Young (3-month-old) (n = 10) and aged (20-month-old) (n = 13) control male Fischer 344 rats were treated with water by oral gavage. All groups were treated for a period of 4 weeks. Although there was no difference in performance in the MWM when comparing all aged rats, when the data for aged impaired rats were compared, there was a significant difference between groups on the last day of training with the treatment group performing better than controls. Using the cell cycle-regulating protein (Ki67), doublecortin (DCX), and OX6 antibody markers, cell proliferation, neurogenesis, and microglial activation were estimated in the dentate gyrus (DG) of young and aged animals. Cell proliferation was also examined in the subventricular zone (SVZ). A decreased number of OX6 MHC II-positive cells, increased neurogenesis, and increased number of proliferating cells were found in rats treated with NT-020 in comparison with aged control rats. In sum, NT-020 may promote health, proliferation, and maintenance of neurons in the age animals and exert antiinflammatory actions that promote function in the aged stem cell niche.

Spirulina promotes stem cell genesis and protects against LPS induced declines in neural stem cell proliferation.

Adult stem cells are present in many tissues including, skin, muscle, adipose, bone marrow, and in the brain. Neuroinflammation has been shown to be a potent negative regulator of stem cell and progenitor cell proliferation in the neurogenic regions of the brain. Recently we demonstrated that decreasing a key neuroinflammatory cytokine IL-1beta in the hippocampus of aged rats reversed the age-related cognitive decline and increased neurogenesis in the age rats. We also have found that nutraceuticals have the potential to reduce neuroinflammation, and decrease oxidative stress. The objectives of this study were to determine if spirulina could protect the proliferative potential of hippocampal neural progenitor cells from an acute systemic inflammatory insult of lipopolysaccharide (LPS). To this end, young rats were fed for 30 days a control diet or a diet supplemented with 0.1% spirulina. On day 28 the rats were given a single i.p. injection of LPS (1 mg/kg). The following day the rats were injected with BrdU (50 mg/kg b.i.d. i.p.) and were sacrificed 24 hours after the first injection of BrdU. Quantification of the BrdU positive cells in the subgranular zone of the dentate gyrus demonstrated a decrease in proliferation of the stem/progenitor cells in the hippocampus as a result of the LPS insult. Furthermore, the diet supplemented with spirulina was able to negate the LPS induced decrease in stem/progenitor cell proliferation. In a second set of studies we examined the effects of spirulina either alone or in combination with a proprietary formulation (NT-020) of blueberry, green tea, vitamin D3 and carnosine on the function of bone marrow and CD34+ cells in vitro. Spirulina had small effects on its own and more than additive effects in combination with NT-020 to promote mitochondrial respiration and/or proliferation of these cells in culture. When examined on neural stem cells in culture spirulina increased proliferation at baseline and protected against the negative influence of TNFalpha to reduce neural stem cell proliferation. These results support the hypothesis that a diet enriched with spirulina and other nutraceuticals may help protect the stem/progenitor cells from insults.

Neuroimmunomodulation and Aging.

Inflammation is by definition a protective phase of the immune response. The very first goal of inflammation is destroying and phagocytosing infected or damaged cells to avoid the spread of the pathogen or of the damage to neighboring, healthy, cells. However, we now know that during many chronic neurological disorders, inflammation and degeneration always coexist at certain time points. For example, inflammation comes first in multiple sclerosis, but degeneration follows, while in Alzheimer's or Parkinson's disease degeneration starts and inflammation is secondary. Either way these are the two pathological detectable problems. The central nervous system (CNS) has long been viewed as exempt from the effects of the immune system. The brain has physical barriers for protection, and it is now clear that cells in the nervous system respond to inflammation and injury in unique ways. In recent years, researchers have presented evidence supporting the idea that in the CNS there is an ongoing protective inflammatory mechanism, which involves macrophage, monocytes, T cells, regulatory T-cells, effector T cells and many others; these, in turn, promote repair mechanisms in the brain not only during inflammatory, and degenerative disorders but also in healthy people. This "repair mechanism" can be considered as an intrinsic part of the physiological activities of the brain. It is now well known that the microenvironment of the brain is a crucial player in determining the relative contribution of the two different outcomes. Failure of molecular and cellular mechanisms sustaining the "brain-repair programme" might be, at least in part, a cause of neurological disorders. Today, the neurotoxic and neuroprotective roles of the innate immune reactions in aging, brain injury, ischemia, autoimmune and neurodegenerative disorders of the CNS are widely investigated and highly debated research topics. Nevertheless, several issues remain to be elucidated, notably the earlier cellular events that initiate dysregulation of brain inflammatory pathways. If these inflammatory processes could be identified and harnessed, then cognitive function may be protected during aging and age-related neurodegenerative diseases through early interventions directed against the negative consequences of inflammation. This commentary highlights the major issues/opinions presented by experts on the involvement of the brain immune system in aging and age-related diseases in a special edition of the journal Aging and Disease.

Neuron-Microglia Dialogue and Hippocampal Neurogenesis in the Aged Brain.

Age-related changes in innate immune function and glial-neuronal communication are early and critical events in brain aging and neurodegenerative disease, and lead to a chronic increase in oxidative stress and inflammation, which initiates neuronal dysfunction and reduced synaptic plasticity, and ultimately disruption in learning and memory in the aged brain. Several lines of evidence suggest a correlation between adult neurogenesis and learning. It has been proposed that a decline in hippocampal neurogenesis contributes to a physiologic decline in brain function. Recently, new and important insights relating to the production of new neurons affecting hippocampal-dependent memory ability have been provided. A multitude of factors have been shown to regulate the production of new neurons in the adult hippocampus, many of which change as a result of aging. Yet, the potential importance of neurogenesis in some affective and cognitive behaviors, as well as endogenous tissue repair mechanisms, makes further investigation of neurogenic regulators warranted. We have recent evidence that key regulators of communication between neurons and microglia are disrupted in the aged brain and may be one of the factors that precedes and initiates the observed increase in chronic inflammatory state. In this review the role of dysfunction in these neuronal-glial communication regulators underlying age-related impairments in cognition and hippocampal neurogenesis will be discussed. An understanding of these mechanisms will lead to the development of preventive or protective therapies.

Increased neuronal proliferation in the dentate gyrus of aged rats following neural stem cell implantation.

It is now well accepted that the brain is able to generate newborn neurons from a population of resident multipotential neural stem cells (NSCs) located in two discrete regions of the brain. The capacity for neurogenesis appears to diminish over the lifespan of an organism. Methods to potentiate the proliferation of new neuronal or glial cells within the central nervous system from resident NSCs could have therapeutic potential following an insult, such as stroke, or to replace lost cells as a result of a neurodegenerative disease. We implanted cells from a human NSC cell line, CTX0E03, originally derived from fetal cortical tissue directly into the ventricles of aged rats. CTX0E03 cells have angiogenic properties via secretion of growth factors, so we investigated if the implanted cells would stimulate proliferation of NSCs within the subgranular zone (SGZ) of the dentate gyrus. Bromodeoxyuridine staining demonstrated significantly increased proliferation in the SGZ. Absence of double labeling for human nuclear antigen suggested that the increased proliferation was from endogenous neural progenitor cells. The acute treatment also led to an increased number of immature neurons as demonstrated by immunohistochemical staining for the immature neuronal marker doublecortin. The data suggest that implants of exogenous NSCs may promote regeneration in aging organisms through stimulation of endogenous neurogenesis.

Role of TNFα Induced Inflammation in Delay Eyeblink Conditioning in Young and Aged Rats.

Tumor necrosis factor alpha (TNF-α) is a multifunctional proinflammatory cytokine, which is a critical inflammatory mediator involved in aging and neurodegenerative diseases of aging. Previous work has shown that diets enriched with antioxidants reduce levels of the cytokine TNF-α and improve classical eyeblink conditioning performance. Therefore we tested the hypothesis that the proinflamatory cytokine TNF-α may be a critical factor that modulates classical conditioning behavior. If increased levels of endogenous cerebellar TNF-α negatively affect performance on the eyeblink conditioning task in aged rats, then exogenous administration of TNF-α in young rats should result in an impaired acquisition and/or retention of eyeblink conditioning memory. On the other hand, the reduction or blockage of the age-related increase in cerebellar TNF-α levels in aged rats should result in an improvement in memory. Young (3 month old) F344 rats were pretreated with an intracerebellar injection of recombinant rat (rr)TNF-α or denatured (rr)TNF-α prior to eyeblink conditioning coupled to microdialysis. The results showed that young rats treated with rrTNF-α have a decreased rate of learning compared to the control group. Norepinephrine which has been shown to play a critical role in cerebellar learning tasks presented a shift on training day one of young rats resembling that observed in aged rats. In a second experiment aged (22 month old) F344 rats were pretreated with intracerebellar microinjection of anti-rat TNF-α three times a week for 4 weeks prior to eyeblink conditioning training couple to microdialysis. Aged rats showed a better performance in the conditioned responses when compared to controls. The release of norepinephrine in this group reached basal levels sooner than the control group but not as early as the young rats. The results of these experiments demonstrate a critical correlation between TNF-α and the rate of learning and the pattern of NE release during eyeblink conditioning.

Human umbilical cord blood treatment in a mouse model of ALS: optimization of cell dose.

BACKGROUND: Amyotrophic Lateral Sclerosis (ALS) is a multicausal disease characterized by motor neuron degeneration in the spinal cord and brain. Cell therapy may be a promising new treatment for this devastating disorder. We recently showed that a single low dose (10(6) cells) of mononuclear human umbilical cord blood (MNC hUCB) cells administered intravenously to G93A mice delayed symptom progression and modestly prolonged lifespan. The aim of this pre-clinical translation study is to optimize the dose of MNC hUCB cells to retard disease progression in G93A mice. Three different doses of MNC hUCB cells, 10x10(6), 25x10(6) and 50x10(6), were administered intravenously into pre-symptomatic G93A mice. Motor function tests and various assays to determine cell effects were performed on these mice. METHODOLOGY/PRINCIPAL FINDINGS: Our results showed that a cell dose of 25x10(6) cells significantly increased lifespan of mice by 20-25% and delayed disease progression by 15%. The most beneficial effect on decreasing pro-inflammatory cytokines in the brain and spinal cord was found in this group of mice. Human Th2 cytokines were found in plasma of mice receiving 25x10(6) cells, although prevalent human Th1 cytokines were indicated in mice with 50x10(6) cells. High response of splenic cells to mitogen (PHA) was indicated in mice receiving 25x10(6) (mainly) and 10x10(6) cells. Significantly increased lymphocytes and decreased neutrophils in the peripheral blood were found only in animals receiving 25x10(6) cells. Stable reduction in microglia density in both cervical and lumbar spinal cords was also noted in mice administered with 25x10(6) cells. CONCLUSIONS/SIGNIFICANCE: These results demonstrate that treatment for ALS with an appropriate dose of MNC hUCB cells may provide a neuroprotective effect for motor neurons through active involvement of these cells in modulating the host immune inflammatory system response.

Peripheral injection of human umbilical cord blood stimulates neurogenesis in the aged rat brain.

BACKGROUND: Neurogenesis continues to occur throughout life but dramatically decreases with increasing age. This decrease is mostly related to a decline in proliferative activity as a result of an impoverishment of the microenvironment of the aged brain, including a reduction in trophic factors and increased inflammation. RESULTS: We determined that human umbilical cord blood mononuclear cells (UCBMC) given peripherally, by an intravenous injection, could rejuvenate the proliferative activity of the aged neural stem/progenitor cells. This increase in proliferation lasted for at least 15 days after the delivery of the UCBMC. Along with the increase in proliferation following UCBMC treatment, an increase in neurogenesis was also found in the aged animals. The increase in neurogenesis as a result of UCBMC treatment seemed to be due to a decrease in inflammation, as a decrease in the number of activated microglia was found and this decrease correlated with the increase in neurogenesis. CONCLUSION: The results demonstrate that a single intravenous injection of UCBMC in aged rats can significantly improve the microenvironment of the aged hippocampus and rejuvenate the aged neural stem/progenitor cells. Our results raise the possibility of a peripherally administered cell therapy as an effective approach to improve the microenvironment of the aged brain.

Blockade of caspase-1 increases neurogenesis in the aged hippocampus.

Adult hippocampal neurogenesis dramatically decreases with increasing age, and it has been proposed that this decline contributes to age-related memory deficits. Central inflammation contributes significantly to the decrease in neurogenesis associated with ageing. Interleukin-1beta is a proinflammatory cytokine initially synthesized as an inactive precursor that is cleaved by caspase-1 to generate the biologically active mature form. Whether IL-1beta affects neurogenesis in the aged hippocampus is unknown. Here we analysed cells positive for 5-bromo-2-deoxyuridine (BrdU; 50 mg/kg) in animals in which cleavage of IL-1beta was inhibited by the caspase-1 inhibitor Ac-YVAD-CMK (10 pmol). Aged (22 months) and young (4 months) rats received Ac-YVAD-CMK for 28 days intracerebroventricularly through a brain infusion cannula connected to an osmotic minipump. Starting on day 14, animals received a daily injection of BrdU for five consecutive days. Unbiased stereology analyses performed 10 days after the last injection of BrdU revealed that the total number of newborn cells generated over a 5-day period was higher in young rats than in aged rats. In addition, there was a 53% increase in the number of BrdU-labelled cells of the aged Ac-YVAD-CMK-treated rats compared to aged controls. Immunofluorescence studies were performed to identify the cellular phenotype of BrdU-labelled cells. The increase in BrdU-positive cells was not due to a change in the proportion of cells expressing neuronal or glial phenotypes in the subgranular zone. These findings demonstrate that the intracerebroventricular administration of Ac-YVAD-CMK reversed the decrease in hippocampal neurogenesis associated with ageing.

Interleukin-1beta and caspase-1: players in the regulation of age-related cognitive dysfunction.

Scientific research on the unprecedented and growing number of older adults in the United States and other industrialized countries has focused much attention on the health consequences of aging. Over the last few decades, inflammation in the brain and its implication in the progression of aging and age-related cognitive dysfunction has been an area of increasing importance to neuroscientists and is now considered as one of the most interesting and promising topics for aging research. One of the critical aspects of inflammatory processes is that the activation of one upstream inflammatory molecule initiates a cascade of self-sustaining inflammatory events which leads to the activation of a number of different downstream functions. Recently, a great deal of attention has been given to the interplay between inflammatory and apoptotic processes and the regulation of these processes by the caspases. The caspase family of proteases can be divided into proapoptotic and pro-inflammatory members. The present review summarizes recent observations of the interactions between the inflammatory cytokine interleuldn-1 (IL-1) beta and the inflammatory/apoptotic caspase-1 and their involvement in age-related impairments in cognition. A comprehensive understanding of these mechanisms could potentially lead to the development of preventive or protective therapies that reduce or inhibit the cognitive decline associated with aging and age-related neurodegenerative disease.