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The effects of harvesting media on biological characteristics and repair potential of neural stem cells after traumatic brain injury.

ABSTRACT: Various solutions are utilized widely for the isolation, harvesting, sorting, testing and transplantation of neural stem cells (NSCs), whereas the effects of harvesting media on the biological characteristics and repair potential of NSCs remain unclear. To examine some of these effects, NSCs were isolated from cortex of E14.5 mice and exposed to the conventional harvesting media [0.9% saline (Saline), phosphate-buffered saline (PBS) or artificial cerebrospinal fluid (ACSF)] or the proliferation culture medium (PCM) for different durations at 4°C. Treated NSCs were grafted by in situ injection into the lesion sites of traumatic brain injury (TBI) mice. In vitro, harvesting media-exposed NSCs displayed time-dependent reduction of viability and proliferation. S phase entry decreased in harvesting media-exposed cells, which was associated with upregulation of p53 protein and downregulation of cyclin E1 protein. Moreover, harvesting media exposure induced the necrosis and apoptosis of NSCs. The levels of Fas-L, cleaved caspase 3 and 8 were increased, which suggests that the death receptor signaling pathway is involved in the apoptosis of NSCs. In addition, exposure to Saline did not facilitate the neuronal differentiation of NSCs, suggesting that Saline exposure may be disadvantageous for neurogenesis. In vivo, NSC-mediated functional recovery in harvesting media-exposed NSC groups was notably attenuated in comparison with the PCM-exposed NSC group. In conclusion, harvesting media exposure modulates the biological characteristics and repair potential of NSCs after TBI. Our results suggest that insight of the effects of harvesting media exposure on NSCs is critical for developing strategies to assure the successful long-term engraftment of NSCs.

SUBMITTER: Liu S

PROVIDER: S-EPMC4172630 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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The effects of harvesting media on biological characteristics and repair potential of neural stem cells after traumatic brain injury.

Liu Shengliang S Li Zhuying Z Fu Jin J Sun Liang L Xu Fengyan F Harada Toshihide T Lou Yu Y Chu Ming M Sun Qi Q Xu Kun K Zhang Rui R Jin Lianhong L Xiao Hui H Wu Shuliang S

PloS one 20140923 9

Various solutions are utilized widely for the isolation, harvesting, sorting, testing and transplantation of neural stem cells (NSCs), whereas the effects of harvesting media on the biological characteristics and repair potential of NSCs remain unclear. To examine some of these effects, NSCs were isolated from cortex of E14.5 mice and exposed to the conventional harvesting media [0.9% saline (Saline), phosphate-buffered saline (PBS) or artificial cerebrospinal fluid (ACSF)] or the proliferation ...[more]

PMID: 25247595

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Project description:Traumatic brain injury (TBI) is a major public health concern and remains a leading cause of disability and socio-economic burden. To date, there is no proven therapy that promotes brain repair following an injury to the brain. In this study, we explored the role of an isoform of adenosine kinase expressed in the cell nucleus (ADK-L) as a potential regulator of neural stem cell proliferation in the brain. The rationale for this hypothesis is based on coordinated expression changes of ADK-L during foetal and postnatal murine and human brain development indicating a role in the regulation of cell proliferation and plasticity in the brain. We first tested whether the genetic disruption of ADK-L would increase neural stem cell proliferation after TBI. Three days after TBI, modelled by a controlled cortical impact, transgenic mice, which lack ADK-L (ADKΔneuron) in the dentate gyrus (DG) showed a significant increase in neural stem cell proliferation as evidenced by significant increases in doublecortin and Ki67-positive cells, whereas animals with transgenic overexpression of ADK-L in dorsal forebrain neurons (ADK-Ltg) showed an opposite effect of attenuated neural stem cell proliferation. Next, we translated those findings into a pharmacological approach to augment neural stem cell proliferation in the injured brain. Wild-type C57BL/6 mice were treated with the small molecule adenosine kinase inhibitor 5-iodotubercidin for 3 days after the induction of TBI. We demonstrate significantly enhanced neural stem cell proliferation in the DG of 5-iodotubercidin-treated mice compared to vehicle-treated injured animals. To rule out the possibility that blockade of ADK-L has any effects in non-injured animals, we quantified baseline neural stem cell proliferation in ADKΔneuron mice, which was not altered, whereas baseline neural stem cell proliferation in ADK-Ltg mice was enhanced. Together these findings demonstrate a novel function of ADK-L involved in the regulation of neural stem cell proliferation after TBI.

Project description:Parkinson's Disease (PD) is a progressive neurodegenerative disorder with no cure. Clinical presentation is characterized by postural instability, resting tremors, and gait problems that result from progressive loss of A9 dopaminergic neurons in the substantia nigra pars compacta. Traumatic brain injury (TBI) has been implicated as a risk factor for several neurodegenerative diseases, but the strongest evidence is linked to development of PD. Mild TBI (mTBI), is the most common and is defined by minimal, if any, loss of consciousness and the absence of significant observable damage to the brain tissue. mTBI is responsible for a 56% higher risk of developing PD in U.S. Veterans and the risk increases with severity of injury. While the mounting evidence from human studies suggests a link between TBI and PD, fundamental questions as to whether TBI nucleates PD pathology or accelerates PD pathology in vulnerable populations remains unanswered. Several promising lines of research point to inflammation, metabolic dysregulation, and protein accumulation as potential mechanisms through which TBI can initiate or accelerate PD. Amyloid precursor protein (APP), alpha synuclein (?-syn), hyper-phosphorylated Tau, and TAR DNA-binding protein 43 (TDP-43), are some of the most frequently reported proteins upregulated following a TBI and are also closely linked to PD. Recently, upregulation of Leucine Rich Repeat Kinase 2 (LRRK2), has been found in the brain of mice following a TBI. Subset of Rab proteins were identified as biological substrates of LRRK2, a protein also extensively linked to late onset PD. Inhibition of LRRK2 was found to be neuroprotective in PD and TBI models. The goal of this review is to survey current literature concerning the mechanistic overlap between TBI and PD with a particular focus on inflammation, metabolic dysregulation, and aforementioned proteins. This review will also cover the application of rodent TBI models to further our understanding of the relationship between TBI and PD.

Project description:International Stem Cell Corporation human parthenogenetic neural stem cells (ISC-hpNSC) have potential therapeutic value for patients suffering from traumatic brain injury (TBI). Here, we demonstrate the behavioral and histological effects of transplanting ISC-hpNSC intracerebrally in an animal model of TBI. Methods: Sprague-Dawley rats underwent a moderate controlled cortical impact TBI surgery. Transplantation occurred at 72 h post-TBI with functional readouts of behavioral and histological deficits conducted during the subsequent 3-month period after TBI. We characterized locomotor, neurological, and cognitive performance at baseline (before TBI), then on days 0, 1, 7, 14, 30, 60, and 90 (locomotor and neurological), and on days 28-30, 58-60, and 88-90 (cognitive) after TBI. Following completion of behavioral testing at 3 months post-TBI, animals were euthanized by transcardial perfusion and brains harvested to histologically characterize the extent of brain damage. Neuronal survival was revealed by Nissl staining, and stem cell engraftment and host tissue repair mechanisms such as the anti-inflammatory response in peri-TBI lesion areas were examined by immunohistochemical analyses. Results: We observed that TBI groups given high and moderate doses of ISC-hpNSC had an improved swing bias on an elevated body swing test for motor function, increased scores on forelimb akinesia and paw grasp neurological tests, and committed significantly fewer errors on a radial arm water maze test for cognition. Furthermore, histological analyses indicated that high and moderate doses of stem cells increased the expression of phenotypic markers related to the neural lineage and myelination and decreased reactive gliosis and inflammation in the brain, increased neuronal survival in the peri-impact area of the cortex, and decreased inflammation in the spleen at 90 days post-TBI. Conclusion: These results provide evidence that high and moderate doses of ISC-hpNSC ameliorate TBI-associated histological alterations and motor, neurological, and cognitive deficits.

Project description:BackgroundTraumatic Brain Injury (TBI) is a major cause of disability and mortality, to which there is currently no comprehensive treatment. Blood Brain Barrier (BBB) dysfunction is well documented in human TBI patients, yet the molecular mechanisms that underlie this neurovascular unit (NVU) pathology remains unclear. The apolipoprotein-E (apoE) protein has been implicated in controlling BBB integrity in an isoform dependent manner, via suppression of Cyclophilin A (CypA)-Matrix metallopeptidase-9 (MMP-9) signaling cascades, however the contribution of this pathway in TBI-induced BBB permeability is not fully investigated.MethodsWe exposed C57Bl/6 mice to controlled cortical impact and assessed NVU and BBB permeability responses up to 21 days post-injury. We pharmacologically probed the role of the CypA-MMP-9 pathway in BBB permeability after TBI using Cyclosporin A (CsA, 20 mg/kg). Finally, as the apoE4 protein is known to be functionally deficient compared to the apoE3 protein, we used humanized APOE mice as a clinically relevant model to study the role of apoE on BBB injury and repair after TBI.ResultsIn C57Bl/6 mice there was an inverse relationship between soluble apoE and BBB permeability, such that damaged BBB stabilizes as apoE levels increase in the days following TBI. TBI mice displayed acute pericyte loss, increased MMP-9 production and activity, and reduced tight-junction expression. Treatment with the CypA antagonist CsA in C57Bl/6 mice attenuates MMP-9 responses and enhances BBB repair after injury, demonstrating that MMP-9 plays an important role in the timing of spontaneous BBB repair after TBI. We also show that apoe mRNA is present in both astrocytes and pericytes after TBI. We report that APOE3 and APOE4 mice have similar acute BBB responses to TBI, but APOE3 mice display faster spontaneous BBB repair than APOE4 mice. Isolated microvessel analysis reveals delayed pericyte repopulation, augmented and sustained MMP-9 expression at the NVU, and impaired stabilization of Zonula Occludens-1, Occludin and Claudin-5 expression at tight junctions in APOE4 mice after TBI compared to APOE3 mice.ConclusionsThese data confirm apoE as an important modulator of spontaneous BBB stabilization following TBI, and highlights the APOE4 allele as a risk factor for poor outcome after TBI.

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The effects of harvesting media on biological characteristics and repair potential of neural stem cells after traumatic brain injury.

Publications

The effects of harvesting media on biological characteristics and repair potential of neural stem cells after traumatic brain injury.

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