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Sample records for cell preserving brain

  1. Histone deacetylase inhibitors are neuroprotective and preserve NGF-mediated cell survival following traumatic brain injury

    PubMed Central

    Lu, Jie; Frerich, Jason M.; Turtzo, L. Christine; Li, Siqi; Chiang, Jeffrey; Yang, Chunzhang; Wang, Xiaoping; Zhang, Chao; Wu, Chenxi; Sun, Zhongchan; Niu, Gang; Zhuang, Zhengping; Brady, Roscoe O.; Chen, Xiaoyuan

    2013-01-01

    Acute traumatic brain injury (TBI) is associated with long-term cognitive and behavioral dysfunction. In vivo studies have shown histone deacetylase inhibitors (HDACis) to be neuroprotective following TBI in rodent models. HDACis are intriguing candidates because they are capable of provoking widespread genetic changes and modulation of protein function. By using known HDACis and a unique small-molecule pan-HDACi (LB-205), we investigated the effects and mechanisms associated with HDACi-induced neuroprotection following CNS injury in an astrocyte scratch assay in vitro and a rat TBI model in vivo. We demonstrate the preservation of sufficient expression of nerve growth factor (NGF) and activation of the neurotrophic tyrosine kinase receptor type 1 (TrkA) pathway following HDACi treatment to be crucial in stimulating the survival of CNS cells after TBI. HDACi treatment up-regulated the expression of NGF, phospho-TrkA, phospho-protein kinase B (p-AKT), NF-κB, and B-cell lymphoma 2 (Bcl-2) cell survival factors while down-regulating the expression of p75 neurotrophin receptor (NTR), phospho-JNK, and Bcl-2–associated X protein apoptosis factors. HDACi treatment also increased the expression of the stem cell biomarker nestin, and decreased the expression of reactive astrocyte biomarker GFAP within damaged tissue following TBI. These findings provide further insight into the mechanisms by which HDACi treatment after TBI is neuroprotective and support the continued study of HDACis following acute TBI. PMID:23754423

  2. Scaling brain size, keeping timing: evolutionary preservation of brain rhythms.

    PubMed

    Buzsáki, György; Logothetis, Nikos; Singer, Wolf

    2013-10-30

    Despite the several-thousand-fold increase of brain volume during the course of mammalian evolution, the hierarchy of brain oscillations remains remarkably preserved, allowing for multiple-time-scale communication within and across neuronal networks at approximately the same speed, irrespective of brain size. Deployment of large-diameter axons of long-range neurons could be a key factor in the preserved time management in growing brains. We discuss the consequences of such preserved network constellation in mental disease, drug discovery, and interventional therapies.

  3. Scaling Brain Size, Keeping Timing: Evolutionary Preservation of Brain Rhythms

    PubMed Central

    Buzsáki, György; Logothetis, Nikos; Singer, Wolf

    2014-01-01

    Despite the several-thousand-fold increase of brain volume during the course of mammalian evolution, the hierarchy of brain oscillations remains remarkably preserved, allowing for multiple-time-scale communication within and across neuronal networks at approximately the same speed, irrespective of brain size. Deployment of large-diameter axons of long-range neurons could be a key factor in the preserved time management in growing brains. We discuss the consequences of such preserved network constellation in mental disease, drug discovery, and interventional therapies. PMID:24183025

  4. Preservational Pathways of Corresponding Brains of a Cambrian Euarthropod.

    PubMed

    Ma, Xiaoya; Edgecombe, Gregory D; Hou, Xianguang; Goral, Tomasz; Strausfeld, Nicholas J

    2015-11-16

    The record of arthropod body fossils is traceable back to the "Cambrian explosion," marked by the appearance of most major animal phyla. Exceptional preservation provides crucial evidence for panarthropod early radiation. However, due to limited representation in the fossil record of internal anatomy, particularly the CNS, studies usually rely on exoskeletal and appendicular morphology. Recent studiesshow that despite extreme morphological disparities, euarthropod CNS evolution appears to have been remarkably conservative. This conclusion is supported by descriptions from Cambrian panarthropods of neural structures that contribute to understanding early evolution of nervous systems and resolving controversies about segmental homologies. However, the rarity of fossilized CNSs, even when exoskeletons and appendages show high levels of integrity, brought into question data reproducibility because all but one of the aforementioned studies were based on single specimens. Foremost among objections is the lack of taphonomic explanation for exceptional preservation of a tissue that some see as too prone to decay to be fossilized. Here we describe newly discovered specimens of the Chengjiang euarthropod Fuxianhuia protensa with fossilized brains revealing matching profiles, allowing rigorous testing of the reproducibility of cerebral structures. Their geochemical analyses provide crucial insights of taphonomic pathways for brain preservation, ranging from uniform carbon compressions to complete pyritization, revealing that neural tissue was initially preserved as carbonaceous film and subsequently pyritized. This mode of preservation is consistent with the taphonomic pathways of gross anatomy, indicating that no special mode is required for fossilization of labile neural tissue.

  5. Interaction of ARC and Daxx: A Novel Endogenous Target to Preserve Motor Function and Cell Loss after Focal Brain Ischemia in Mice

    PubMed Central

    Donath, Stefan; An, Junfeng; Lee, Sabrina Lin Lin; Gertz, Karen; Datwyler, Anna Lena; Harms, Ulrike; Müller, Susanne; Farr, Tracy Deanne; Füchtemeier, Martina; Lättig-Tünnemann, Gisela; Lips, Janet; Foddis, Marco; Mosch, Larissa; Bernard, René; Grittner, Ulrike; Balkaya, Mustafa; Kronenberg, Golo; Dirnagl, Ulrich; Endres, Matthias

    2016-01-01

    The aim of this study was to explore the signaling and neuroprotective effect of transactivator of transcription (TAT) protein transduction of the apoptosis repressor with CARD (ARC) in in vitro and in vivo models of cerebral ischemia in mice. In mice, transient focal cerebral ischemia reduced endogenous ARC protein in neurons in the ischemic striatum at early reperfusion time points, and in primary neuronal cultures, RNA interference resulted in greater neuronal susceptibility to oxygen glucose deprivation (OGD). TAT.ARC protein delivery led to a dose-dependent better survival after OGD. Infarct sizes 72 h after 60 min middle cerebral artery occlusion (MCAo) were on average 30 ± 8% (mean ± SD; p = 0.005; T2-weighted MRI) smaller in TAT.ARC-treated mice (1 μg intraventricularly during MCAo) compared with controls. TAT.ARC-treated mice showed better performance in the pole test compared with TAT.β-Gal-treated controls. Importantly, post-stroke treatment (3 h after MCAo) was still effective in affording reduced lesion volume by 20 ± 7% (mean ± SD; p < 0.05) and better functional outcome compared with controls. Delayed treatment in mice subjected to 30 min MCAo led to sustained neuroprotection and functional behavior benefits for at least 28 d. Functionally, TAT.ARC treatment inhibited DAXX–ASK1–JNK signaling in the ischemic brain. ARC interacts with DAXX in a CARD-dependent manner to block DAXX trafficking and ASK1–JNK activation. Our work identifies for the first time ARC–DAXX binding to block ASK1–JNK activation as an ARC-specific endogenous mechanism that interferes with neuronal cell death and ischemic brain injury. Delayed delivery of TAT.ARC may present a promising target for stroke therapy. SIGNIFICANCE STATEMENT Up to now, the only successful pharmacological target of human ischemic stroke is thrombolysis. Neuroprotective pharmacological strategies are needed to accompany therapies aiming to achieve reperfusion. We describe that apoptosis

  6. Intracranial mechanisms for preserving brain blood flow in health and disease.

    PubMed

    McBryde, F D; Malpas, S C; Paton, J F R

    2017-01-01

    The brain is an exceptionally energetically demanding organ with little metabolic reserve, and multiple systems operate to protect and preserve the brain blood supply. But how does the brain sense its own perfusion? In this review, we discuss how the brain may harness the cardiovascular system to counter threats to cerebral perfusion sensed via intracranial pressure (ICP), cerebral oxygenation and ischaemia. Since the work of Cushing over 100 years ago, the existence of brain baroreceptors capable of eliciting increases in sympathetic outflow and blood pressure has been hypothesized. In the clinic, this response has generally been thought to occur only in extremis, to perfuse the severely ischaemic brain as cerebral autoregulation fails. We review evidence that pressor responses may also occur with smaller, physiologically relevant increases in ICP. The incoming brain oxygen supply is closely monitored by the carotid chemoreceptors; however, hypoxia and other markers of ischaemia are also sensed intrinsically by astrocytes or other support cells within brain tissue itself and elicit reactive hyperaemia. Recent studies suggest that astrocytic oxygen signalling within the brainstem may directly affect sympathetic nerve activity and blood pressure. We speculate that local cerebral oxygen tension is a major determinant of the mean level of arterial pressure and discuss recent evidence that this may be the case. We conclude that intrinsic intra- and extra-cranial mechanisms sense and integrate information about hypoxia/ischaemia and ICP and play a major role in determining the long-term level of sympathetic outflow and arterial pressure, to optimize cerebral perfusion.

  7. Roles of PINK1, mTORC2, and mitochondria in preserving brain tumor-forming stem cells in a noncanonical Notch signaling pathway.

    PubMed

    Lee, Kyu-Sun; Wu, Zhihao; Song, Yan; Mitra, Siddhartha S; Feroze, Abdullah H; Cheshier, Samuel H; Lu, Bingwei

    2013-12-15

    The self-renewal versus differentiation choice of Drosophila and mammalian neural stem cells (NSCs) requires Notch (N) signaling. How N regulates NSC behavior is not well understood. Here we show that canonical N signaling cooperates with a noncanonical N signaling pathway to mediate N-directed NSC regulation. In the noncanonical pathway, N interacts with PTEN-induced kinase 1 (PINK1) to influence mitochondrial function, activating mechanistic target of rapamycin complex 2 (mTORC2)/AKT signaling. Importantly, attenuating noncanonical N signaling preferentially impaired the maintenance of Drosophila and human cancer stem cell-like tumor-forming cells. Our results emphasize the importance of mitochondria to N and NSC biology, with important implications for diseases associated with aberrant N signaling.

  8. Preservation of frozen yeast cells by trehalose.

    PubMed

    Diniz-Mendes, L; Bernardes, E; de Araujo, P S; Panek, A D; Paschoalin, V M

    1999-12-05

    Two different methods commonly used to preserve intact yeast cells-freezing and freeze-drying-were compared. Different yeast cells submitted to these treatments were stored for 28 days and cell viability assessed during this period. Intact yeast cells showed to be less tolerant to freeze-drying than to freezing. The rate of survival for both treatments could be enhanced by exogenous trehalose (10%) added during freezing and freeze-drying treatments or by a combination of two procedures: a pre-exposure of cells to 40 degrees C for 60 min and addition of trehalose. A maximum survival level of 71.5 +/- 6.3% after freezing could be achieved at the end of a storage period of 28 days, whereas only 25.0 +/- 1.4% showed the ability to tolerate freeze-drying treatment, if both low-temperature treatments were preceded by a heat exposure and addition of trehalose to yeast cells. Increased survival ability was also obtained when the pre-exposure treatment of yeast cells was performed at 10 degrees C for 3 h and trehalose was added: these treatments enhanced cell survival following freezing from 20.5 +/- 7. 7% to 60.0 +/- 3.5%. Although both mild cold and heat shock treatments could enhance cell tolerance to low temperature, only the heat treatment was able to increase the accumulation of intracellular trehalose whereas, during cold shock exposure, the intracellular amount of trehalose remained unaltered. Intracellular trehalose levels seemed not to be the only factor contributing to cell tolerance against freezing and freeze-drying treatments; however, the protection that this sugar confers to cells can be exerted only if it is to be found on both sides of the plasma membrane.

  9. Preserving cell shape under environmental stress.

    PubMed

    Cook, Boaz; Hardy, Robert W; McConnaughey, William B; Zuker, Charles S

    2008-03-20

    Maintaining cell shape and tone is crucial for the function and survival of cells and tissues. Mechanotransduction relies on the transformation of minuscule mechanical forces into high-fidelity electrical responses. When mechanoreceptors are stimulated, mechanically sensitive cation channels open and produce an inward transduction current that depolarizes the cell. For this process to operate effectively, the transduction machinery has to retain integrity and remain unfailingly independent of environmental changes. This is particularly challenging for poikilothermic organisms, where changes in temperature in the environment may impact the function of mechanoreceptor neurons. Thus, we wondered how insects whose habitat might quickly vary over several tens of degrees of temperature manage to maintain highly effective mechanical senses. We screened for Drosophila mutants with defective mechanical responses at elevated ambient temperatures, and identified a gene, spam, whose role is to protect the mechanosensory organ from massive cellular deformation caused by heat-induced osmotic imbalance. Here we show that Spam protein forms an extracellular shield that guards mechanosensory neurons from environmental insult. Remarkably, heterologously expressed Spam protein also endowed other cells with superb defence against physically and chemically induced deformation. We studied the mechanical impact of Spam coating and show that spam-coated cells are up to ten times stiffer than uncoated controls. Together, these results help explain how poikilothermic organisms preserve the architecture of critical cells during environmental stress, and illustrate an elegant and simple solution to such challenge.

  10. Reorganization and preservation of motor control of the brain in spinal cord injury: a systematic review.

    PubMed

    Kokotilo, Kristen J; Eng, Janice J; Curt, Armin

    2009-11-01

    Reorganization of brain function in people with CNS damage has been identified as one of the fundamental mechanisms involved in the recovery of sensorimotor function. Spinal cord injury (SCI) brain mapping studies during motor tasks aim for assessing the reorganization and preservation of brain networks involved in motor control. Revealing the activation of cortical and subcortical brain areas in people with SCI can indicate principal patterns of brain reorganization when the neurotrauma is distal to the brain. This review assessed brain activation after SCI in terms of intensity, volume, and somatotopic localization, as well as preservation of activation during attempted and/or imagined movements. Twenty-five studies meeting the inclusion criteria could be identified in Medline (1980 to January 2008). Relevant characteristics of studies (level of lesion, time after injury, motor task) and mapping techniques varied widely. Changes in brain activation were found in both cortical and subcortical areas of individuals with SCI. In addition, several studies described a shift in the region of brain activation. These patterns appeared to be dynamic and influenced by the level, completeness, and time after injury, as well as extent of clinical recovery. In addition, several aspects of reorganization of brain function following SCI resembled those reported in stroke. This review demonstrates that brain networks involved in different demands of motor control remain responsive even in chronic paralysis. These findings imply that therapeutic strategies aimed at restoring spinal cord function, even in people with chronic SCI, can build on preserved competent brain control.

  11. Pediatric brain tumor cell lines.

    PubMed

    Xu, Jingying; Margol, Ashley; Asgharzadeh, Shahab; Erdreich-Epstein, Anat

    2015-02-01

    Pediatric brain tumors as a group, including medulloblastomas, gliomas, and atypical teratoid rhabdoid tumors (ATRT) are the most common solid tumors in children and the leading cause of death from childhood cancer. Brain tumor-derived cell lines are critical for studying the biology of pediatric brain tumors and can be useful for initial screening of new therapies. Use of appropriate brain tumor cell lines for experiments is important, as results may differ depending on tumor properties, and can thus affect the conclusions and applicability of the model. Despite reports in the literature of over 60 pediatric brain tumor cell lines, the majority of published papers utilize only a small number of these cell lines. Here we list the approximately 60 currently-published pediatric brain tumor cell lines and summarize some of their central features as a resource for scientists seeking pediatric brain tumor cell lines for their research.

  12. P7C3 neuroprotective chemicals block axonal degeneration and preserve function after traumatic brain injury.

    PubMed

    Yin, Terry C; Britt, Jeremiah K; De Jesús-Cortés, Héctor; Lu, Yuan; Genova, Rachel M; Khan, Michael Z; Voorhees, Jaymie R; Shao, Jianqiang; Katzman, Aaron C; Huntington, Paula J; Wassink, Cassie; McDaniel, Latisha; Newell, Elizabeth A; Dutca, Laura M; Naidoo, Jacinth; Cui, Huxing; Bassuk, Alexander G; Harper, Matthew M; McKnight, Steven L; Ready, Joseph M; Pieper, Andrew A

    2014-09-25

    The P7C3 class of neuroprotective aminopropyl carbazoles has been shown to block neuronal cell death in models of neurodegeneration. We now show that P7C3 molecules additionally preserve axonal integrity after injury, before neuronal cell death occurs, in a rodent model of blast-mediated traumatic brain injury (TBI). This protective quality may be linked to the ability of P7C3 molecules to activate nicotinamide phosphoribosyltransferase, the rate-limiting enzyme in nicotinamide adenine dinucleotide salvage. Initiation of daily treatment with our recently reported lead agent, P7C3-S243, 1 day after blast-mediated TBI blocks axonal degeneration and preserves normal synaptic activity, learning and memory, and motor coordination in mice. We additionally report persistent neurologic deficits and acquisition of an anxiety-like phenotype in untreated animals 8 months after blast exposure. Optimized variants of P7C3 thus offer hope for identifying neuroprotective agents for conditions involving axonal damage, neuronal cell death, or both, such as occurs in TBI.

  13. P7C3 Neuroprotective Chemicals Block Axonal Degeneration and Preserve Function after Traumatic Brain Injury

    PubMed Central

    Yin, Terry C.; Britt, Jeremiah K.; De Jesús-Cortés, Héctor; Lu, Yuan; Genova, Rachel M.; Khan, Michael Z.; Voorhees, Jaymie R.; Shao, Jianqiang; Katzman, Aaron C.; Huntington, Paula J.; Wassink, Cassie; McDaniel, Latisha; Newell, Elizabeth A.; Dutca, Laura M.; Naidoo, Jacinth; Cui, Huxing; Bassuk, Alexander G.; Harper, Matthew M.; McKnight, Steven L.; Ready, Joseph M.; Pieper, Andrew A.

    2014-01-01

    SUMMARY The P7C3 class of neuroprotective aminopropyl carbazoles has been shown to block neuronal cell death in models of neurodegeneration. We now show that P7C3 molecules additionally preserve axonal integrity after injury, before neuronal cell death occurs, in a rodent model of blast-mediated traumatic brain injury (TBI). This protective quality may be linked to the ability of P7C3 molecules to activate nicotinamide phosphoribosyltransferase, the rate-limiting enzyme in nicotinamide adenine dinucleotide salvage. Initiation of daily treatment with our recently reported lead agent, P7C3-S243, one day after blast-mediated TBI blocks axonal degeneration and preserves normal synaptic activity, learning and memory, and motor coordination in mice. We additionally report persistent neurologic deficits and acquisition of an anxiety-like phenotype in untreated animals eight months after blast exposure. Optimized variants of P7C3 thus offer hope for identifying neuroprotective agents for conditions involving axonal damage, neuronal cell death, or both, such as occurs in TBI. PMID:25220467

  14. X-linked inhibitor of apoptosis inhibits apoptosis and preserves the blood-brain barrier after experimental subarachnoid hemorrhage.

    PubMed

    Gao, Cheng; Yu, Hongwei; Yan, Cong; Zhao, Wenyang; Liu, Yao; Zhang, Dongdong; Li, Jingwei; Liu, Nan

    2017-03-22

    Early brain injury following subarachnoid hemorrhage (SAH) strongly determines the prognosis of patients suffering from an aneurysm rupture, and apoptosis is associated with early brain injury after SAH. This study was designed to explore the role of X-linked inhibitor of apoptosis (XIAP) in early brain injury following SAH. The expression of XIAP was detected using western blotting and real-time RT-PCR in an autologous blood injection model of SAH. We also studied the role of XIAP in early brain injury and detected apoptosis-related proteins. The results showed that XIAP was significantly up-regulated in the cortex and hippocampus and that XIAP was mainly expressed in neuronal cells following SAH. The inhibition of endogenous XIAP aggravated blood-brain barrier disruption, neurological deficits and brain edema. Recombinant XIAP preserved the blood-brain barrier, improved the neurological scores and ameliorated brain edema. Recombinant XIAP treatment also decreased the expression of cleaved caspase-3, caspase-8 and caspase-9, whereas there was no effect on the expression of p53, apoptosis-inducing factor or cytochrome c. These results show that XIAP acts as an endogenous neuroprotective and anti-apoptotic agent following SAH. The effects of XIAP on early brain injury was associated with the inhibition of the caspase-dependent apoptosis pathway.

  15. X-linked inhibitor of apoptosis inhibits apoptosis and preserves the blood-brain barrier after experimental subarachnoid hemorrhage

    PubMed Central

    Gao, Cheng; Yu, Hongwei; Yan, Cong; Zhao, Wenyang; Liu, Yao; Zhang, Dongdong; Li, Jingwei; Liu, Nan

    2017-01-01

    Early brain injury following subarachnoid hemorrhage (SAH) strongly determines the prognosis of patients suffering from an aneurysm rupture, and apoptosis is associated with early brain injury after SAH. This study was designed to explore the role of X-linked inhibitor of apoptosis (XIAP) in early brain injury following SAH. The expression of XIAP was detected using western blotting and real-time RT-PCR in an autologous blood injection model of SAH. We also studied the role of XIAP in early brain injury and detected apoptosis-related proteins. The results showed that XIAP was significantly up-regulated in the cortex and hippocampus and that XIAP was mainly expressed in neuronal cells following SAH. The inhibition of endogenous XIAP aggravated blood-brain barrier disruption, neurological deficits and brain edema. Recombinant XIAP preserved the blood-brain barrier, improved the neurological scores and ameliorated brain edema. Recombinant XIAP treatment also decreased the expression of cleaved caspase-3, caspase-8 and caspase-9, whereas there was no effect on the expression of p53, apoptosis-inducing factor or cytochrome c. These results show that XIAP acts as an endogenous neuroprotective and anti-apoptotic agent following SAH. The effects of XIAP on early brain injury was associated with the inhibition of the caspase-dependent apoptosis pathway. PMID:28327595

  16. Reorganization and Preservation of Motor Control of the Brain in Spinal Cord Injury: A Systematic Review

    PubMed Central

    Kokotilo, Kristen J; Eng, Janice J; Curt, Armin

    2011-01-01

    Reorganization of brain function in people with CNS damage has been identified as one of the fundamental mechanisms involved in the recovery of sensori-motor function. Spinal cord injury (SCI) brain mapping studies during motor tasks aim for assessing the reorganization and preservation of brain networks involved in motor control. Revealing the activation of cortical and sub-cortical brain areas in people with SCI can indicate principal patterns of brain reorganization when the neurotrauma is distal to the brain. This review assessed brain activation after SCI in terms of intensity, volume, and somatotopic localization, as well as preservation of activation during attempted and/or imagined movements. Twenty-five studies meeting the inclusion criteria could be identified in MEDLINE (1980 to January 2008). Relevant characteristics of studies (level of lesion, time after injury, motor task) and mapping techniques varied widely. Changes in brain activation were found in both cortical and subcortical areas of individuals with SCI. In addition, several studies described a shift in the region of brain activation. These patterns appeared to be dynamic and influenced by the level, completeness and time after injury, as well as extent of clinical recovery. In addition, several aspects of reorganization of brain function following SCI resembled those reported in stroke. This review demonstrates that brain networks involved in different demands of motor control remain responsive even in chronic paralysis. These findings imply that therapeutic strategies aiming for restoring spinal cord function even in people with chronic SCI can build on a preserved competent brain control. PMID:19604097

  17. Preservation of cell structures in permafrost: a model for exobiology.

    PubMed

    Soina, V S; Vorobiova, E A; Zvyagintsev, D G; Gilichinsky, D A

    1995-03-01

    The present report is the first contribution toward a comprehensive fine-structural study of microbial cells from permafrost. Prokaryotes with a variety of cell wall types demonstrate high stability of cell structure after long-term cryopreservation in frozen soils and sediments of the Arctic. The surface capsular layers that were a salient feature of the cells both in situ and on nutrient media may be an adaptation to low temperature. To the extent that permafrost regions on Earth approximate Martian conditions, preservation of cell structure there can serve as the basis for predictions about preservation in Martian permafrost sediments.

  18. Preserved brains from the Spanish Civil War mass grave (1936) at La Pedraja1, Burgos, Spain.

    PubMed

    Serrulla, Fernando; Herrasti, Lourdes; Navarro, Carmen; Cascallana, Jose Luis; Bermejo, Ana Maria; Marquez-Grant, Nicholas; Etxeberria, Francisco

    2016-12-01

    During the excavation of the Spanish Civil War mass grave at La Pedraja (Burgos, Spain), 104 individuals were found interred within it, 45 of which displayed brains that were preserved but dehydrated and reduced in size. This exceptional finding has resulted in the formation of a multidisciplinary team, with the aim of obtaining as much information as possible and to primarily understand the taphonomic phenomena that has led to the preservation of these brains. The following types of analyses were undertaken on three of these brains: macroscopy, histology, radiology, chemical-toxicology, genetics, chemical analysis of the soil and 3D modelling for stereolithography. The historical context was considered, plus all archaeological and other forensic data provided by the investigation of the mass grave. The results of the analyses on these morphologically identifiable human brains confirmed the presence of nerve structures, fatty acids, and in one case ante-mortem evidence for an intracranial haemorrhage. The fatty acid profile corresponds to the process of saponification. Therefore, the interpretation is that the preservation of these brains at the mass grave of La Pedraja was due to the saponification process, which was influenced by the manner and cause of death, the chemical composition of the brain, the physicochemical properties of the soil and the meteorological conditions at the time.

  19. Peroxide detoxification by brain cells.

    PubMed

    Dringen, Ralf; Pawlowski, Petra G; Hirrlinger, Johannes

    Peroxides are generated continuously in cells that consume oxygen. Among the different peroxides, hydrogen peroxide is the molecule that is formed in highest quantities. In addition, organic hydroperoxides are synthesized as products of cellular metabolism. Generation and disposal of peroxides is a very important process in the human brain, because cells of this organ consume 20% of the oxygen used by the body. To prevent cellular accumulation of peroxides and damage generated by peroxide-derived radicals, brain cells contain efficient antioxidative defense mechanisms that dispose of peroxides and protect against oxidative damage. Cultured brain cells have been used frequently to investigate peroxide metabolism of neural cells. Efficient disposal of exogenous hydrogen peroxide was found for cultured astrocytes, oligodendrocytes, microglial cells, and neurons. Comparison of specific peroxide clearance rates revealed that cultured oligodendrocytes dispose of the peroxide quicker than the other neural cell cultures. Both catalase and the glutathione system contribute to the clearance of hydrogen peroxide by brain cells. For efficient glutathione-dependent reduction of peroxides, neural cells contain glutathione in high concentration and have substantial activity of glutathione peroxidase, glutathione reductase, and enzymes that supply the NADPH required for the glutathione reductase reaction. This article gives an overview on the mechanisms involved in peroxide detoxification in brain cells and on the capacity of the different types of neural cells to dispose of peroxides.

  20. Preserved pontine glucose metabolism in Alzheimer disease: A reference region for functional brain image (PET) analysis

    SciTech Connect

    Minoshima, Satoshi; Frey, K.A.; Foster, N.L.; Kuhl, D.W.

    1995-07-01

    Our goal was to examine regional preservation of energy metabolism in Alzheimer disease (AD) and to evaluate effects of PET data normalization to reference regions. Regional metabolic rates in the pons, thalamus, putamen, sensorimotor cortex, visual cortex, and cerebellum (reference regions) were determined stereotaxically and examined in 37 patients with probable AD and 22 normal controls based on quantitative {sup 18}FDG-PET measurements. Following normalization of metabolic rates of the parietotemporal association cortex and whole brain to each reference region, distinctions of the two groups were assessed. The pons showed the best preservation of glucose metabolism in AD. Other reference regions showed relatively preserved metabolism compared with the parietotemporal association cortex and whole brain, but had significant metabolic reduction. Data normalization to the pons not only enhanced statistical significance of metabolic reduction in the parietotemporal association cortex, but also preserved the presence of global cerebral metabolic reduction indicated in analysis of the quantitative data. Energy metabolism in the pons in probable AD is well preserved. The pons is a reliable reference for data normalization and will enhance diagnostic accuracy and efficiency of quantitative and nonquantitative functional brain imaging. 39 refs., 2 figs., 3 tabs.

  1. PRMT7 Preserves Satellite Cell Regenerative Capacity.

    PubMed

    Blanc, Roméo Sébastien; Vogel, Gillian; Chen, Taiping; Crist, Colin; Richard, Stéphane

    2016-02-16

    Regeneration of skeletal muscle requires the continued presence of quiescent muscle stem cells (satellite cells), which become activated in response to injury. Here, we report that whole-body protein arginine methyltransferase PRMT7(-/-) adult mice and mice conditionally lacking PRMT7 in satellite cells using Pax7-CreERT2 both display a significant reduction in satellite cell function, leading to defects in regenerative capacity upon muscle injury. We show that PRMT7 is preferentially expressed in activated satellite cells and, interestingly, PRMT7-deficient satellite cells undergo cell-cycle arrest and premature cellular senescence. These defects underlie poor satellite cell stem cell capacity to regenerate muscle and self-renew after injury. PRMT7-deficient satellite cells express elevated levels of the CDK inhibitor p21CIP1 and low levels of its repressor, DNMT3b. Restoration of DNMT3b in PRMT7-deficient cells rescues PRMT7-mediated senescence. Our findings define PRMT7 as a regulator of the DNMT3b/p21 axis required to maintain muscle stem cell regenerative capacity.

  2. Intravenous administration of xenogenic adipose-derived mesenchymal stem cells (ADMSC) and ADMSC-derived exosomes markedly reduced brain infarct volume and preserved neurological function in rat after acute ischemic stroke

    PubMed Central

    Wallace, Christopher Glenn; Yuen, Chun-Man; Kao, Gour-Shenq; Chen, Yi-Ling; Shao, Pei-Lin; Chen, Yung-Lung; Chai, Han-Tan; Lin, Kun-Chen; Liu, Chu-Feng; Chang, Hsueh-Wen; Lee, Mel S.; Yip, Hon-Kan

    2016-01-01

    We tested the hypothesis that combined xenogenic (from mini-pig) adipose-derived mesenchymal stem cell (ADMSC) and ADMSC-derived exosome therapy could reduce brain-infarct zone (BIZ) and enhance neurological recovery in rat after acute ischemic stroke (AIS) induced by 50-min left middle cerebral artery occlusion. Adult-male Sprague-Dawley rats (n = 60) were divided equally into group 1 (sham-control), group 2 (AIS), group 3 [AIS-ADMSC (1.2×106 cells)], group 4 [AIS-exosome (100μg)], and group 5 (AIS-exosome-ADMSC). All therapies were provided intravenously at 3h after AIS procedure. BIZ determined by histopathology (by day-60) and brain MRI (by day-28) were highest in group 2, lowest in group 1, higher in groups 3 and 4 than in group 5, but they showed no difference between groups 3 and 4 (all p < 0.0001). By day-28, sensorimotor functional results exhibited an opposite pattern to BIZ among the five groups (p < 0.005). Protein expressions of inflammatory (inducible nitric oxide synthase/tumor necrosis factor-α/nuclear factor-κB/interleukin-1β/matrix metalloproteinase-9/plasminogen activator inhibitor-1/RANTES), oxidative-stress (NOX-1/NOX-2/oxidized protein), apoptotic (caspase-3/ Poly-ADP-ribose polymerase), and fibrotic (Smad3/transforming growth factor-β) biomarkers, and cellular expressions of brain-damaged (γ-H2AX+/ XRCC1-CD90+/p53BP1-CD90+), inflammatory (CD11+/CD68+/glial fibrillary acid protein+) and brain-edema (aquaporin-4+) markers showed a similar pattern of BIZ among the groups (all n < 0.0001). In conclusion, xenogenic ADMSC/ADMSC-derived exosome therapy was safe and offered the additional benefit of reducing BIZ and improving neurological function in rat AIS. PMID:27793019

  3. Clinical guide to fertility preservation in hematopoietic cell transplant recipients.

    PubMed

    Joshi, S; Savani, B N; Chow, E J; Gilleece, M H; Halter, J; Jacobsohn, D A; Pidala, J; Quinn, G P; Cahn, J-Y; Jakubowski, A A; Kamani, N R; Lazarus, H M; Rizzo, J D; Schouten, H C; Socie, G; Stratton, P; Sorror, M L; Warwick, A B; Wingard, J R; Loren, A W; Majhail, N S

    2014-04-01

    With broadening indications, more options for hematopoietic cell transplantation (HCT) and improvement in survival, the number of long-term HCT survivors is expected to increase steadily. Infertility is a frequent problem that long-term HCT survivors and their partners face and it can negatively impact on the quality of life. The most optimal time to address fertility issues is before the onset of therapy for the underlying disease; however, fertility preservation should also be addressed before HCT in all children and patients of reproductive age, with referral to a reproductive specialist for patients interested in fertility preservation. In vitro fertilization (IVF) and embryo cryopreservation, oocyte cryopreservation and ovarian tissue banking are acceptable methods for fertility preservation in adult women/pubertal females. Sperm banking is the preferred method for adult men/pubertal males. Frequent barriers to fertility preservation in HCT recipients may include the perception of lack of time to preserve fertility given an urgency to move ahead with transplant, lack of patient-physician discussion because of several factors (for example, time constraints, lack of knowledge), inadequate access to reproductive specialists, and costs and lack of insurance coverage for fertility preservation. There is a need to raise awareness in the medical community about fertility preservation in HCT recipients.

  4. Searching for the philosopher's stone: promising links between meditation and brain preservation.

    PubMed

    Luders, Eileen; Cherbuin, Nicolas

    2016-06-01

    In the context of an aging population and increased prevalence of dementia and other neurodegenerative diseases, developing strategies to decrease the negative effects of aging is imperative. The scientific study of meditation as a potential tool to downregulate processes implicated in brain aging is an emerging field, and a growing body of research suggests that mindfulness practices are beneficial for cerebral resilience. Adding further evidence to this notion, an increasing number of imaging studies report effects of meditation on brain structure that are consistent with our understanding of neuroprotection. Here, we review the published findings in this field of research addressing the question of whether meditation diminishes age-related brain degeneration. Altogether, although analyses are still sparse and based on cross-sectional data, study outcomes suggest that meditation might be beneficial for brain preservation-both with respect to gray and white matter-possibly by slowing down the natural (age-related) decrease of brain tissue. Nevertheless, it should also be recognized that, until robust longitudinal data become available, there is no evidence for causation between meditation and brain preservation. This review includes a comprehensive commentary on limitations of the existing research and concludes with implications and directions for future studies.

  5. Purine and pyrimidine nucleosides preserve human astrocytoma cell adenylate energy charge under ischemic conditions.

    PubMed

    Balestri, Francesco; Giannecchini, Michela; Sgarrella, Francesco; Carta, Maria Caterina; Tozzi, Maria Grazia; Camici, Marcella

    2007-02-01

    The brain depends on both glycolysis and mitochondrial oxidative phosphorylation for maintenance of ATP pools. Astrocytes play an integral role in brain functions providing trophic supports and energy substrates for neurons. In this paper, we report that human astrocytoma cells (ADF) undergoing ischemic conditions may use both purine and pyrimidine nucleosides as energy source to slow down cellular damage. The cells are subjected to metabolic stress conditions by exclusion of glucose and incubation with oligomycin (an inhibitor of oxidative phosphorylation). This treatment brings about a depletion of the ATP pool, with a concomitant increase in the AMP levels, which results in a significant decrease of the adenylate energy charge. The presence of purine nucleosides in the culture medium preserves the adenylate energy charge, and improves cell viability. Besides purine nucleosides, also pyrimidine nucleosides, such as uridine and, to a lesser extent, cytidine, are able to preserve the ATP pool. The determination of lactate in the incubation medium indicates that nucleosides can preserve the ATP pool through anaerobic glycolysis, thus pointing to a relevant role of the phosphorolytic cleavage of the N-glycosidic bond of nucleosides which generates, without energy expense, the phosphorylated pentose, which through the pentose phosphate pathway and glycolysis can be converted to energetic intermediates also in the absence of oxygen. In fact, ADF cells possess both purine nucleoside phosphorylase and uridine phosphorylase activities.

  6. Caloric restriction increases ketone bodies metabolism and preserves blood flow in aging brain

    PubMed Central

    Lin, Ai-Ling; Zhang, Wei; Gao, Xiaoli; Watts, Lora

    2015-01-01

    Caloric restriction (CR) has been shown to increase the life span and health span of a broad range of species. However, CR effects on in vivo brain functions are far from explored. In this study, we used multimetric neuroimaging methods to characterize the CR-induced changes of brain metabolic and vascular functions in aging rats. We found that old rats (24 months of age) with CR diet had reduced glucose uptake and lactate concentration, but increased ketone bodies level, compared with the age-matched and young (5 months of age) controls. The shifted metabolism was associated with preserved vascular function: old CR rats also had maintained cerebral blood flow relative to the age-matched controls. When investigating the metabolites in mitochondrial tricarboxylic acid cycle, we found that citrate and α-ketoglutarate were preserved in the old CR rats. We suggest that CR is neuroprotective; ketone bodies, cerebral blood flow, and α-ketoglutarate may play important roles in preserving brain physiology in aging. PMID:25896951

  7. Preservation of mitochondrial functional integrity in mitochondria isolated from small cryopreserved mouse brain areas.

    PubMed

    Valenti, Daniela; de Bari, Lidia; De Filippis, Bianca; Ricceri, Laura; Vacca, Rosa Anna

    2014-01-01

    Studies of mitochondrial bioenergetics in brain pathophysiology are often precluded by the need to isolate mitochondria immediately after tissue dissection from a large number of brain biopsies for comparative studies. Here we present a procedure of cryopreservation of small brain areas from which mitochondrial enriched fractions (crude mitochondria) with high oxidative phosphorylation efficiency can be isolated. Small mouse brain areas were frozen and stored in a solution containing glycerol as cryoprotectant. Crude mitochondria were isolated by differential centrifugation from both cryopreserved and freshly explanted brain samples and were compared with respect to their ability to generate membrane potential and produce ATP. Intactness of outer and inner mitochondrial membranes was verified by polarographic ascorbate and cytochrome c tests and spectrophotometric assay of citrate synthase activity. Preservation of structural integrity and oxidative phosphorylation efficiency was successfully obtained in crude mitochondria isolated from different areas of cryopreserved mouse brain samples. Long-term cryopreservation of small brain areas from which intact and phosphorylating mitochondria can be isolated for the study of mitochondrial bioenergetics will significantly expand the study of mitochondrial defects in neurological pathologies, allowing large comparative studies and favoring interlaboratory and interdisciplinary analyses.

  8. Preserving human cells for regenerative, reproductive, and transfusion medicine

    PubMed Central

    Asghar, Waseem; Assal, Rami El; Shafiee, Hadi; Anchan, Raymond M.; Demirci, Utkan

    2014-01-01

    Cell cryopreservation enables maintaining cellular life at sub-zero temperatures by slowing down biochemical processes. Various cell types are routinely cryopreserved in modern reproductive, regenerative, and transfusion medicine. Current cell cryopreservation methods involve freezing (slow/rapid) or vitrifying cells in the presence of a cryoprotective agent (CPA). Although these methods are clinically utilized, cryo-injury due to ice crystals, osmotic shock, and CPA toxicity cause loss of cell viability and function. Recent approaches using minimum volume vitrification provide alternatives to the conventional cryopreservation methods. Minimum volume vitrification provides ultra-high cooling and rewarming rates that enable preserving cells without ice crystal formation. Herein, we review recent advances in cell cryopreservation technology and provide examples of techniques that are utilized in oocyte, stem cell, and red blood cell cryopreservation. PMID:24995723

  9. The preservation of living cells with biocompatible microparticles

    NASA Astrophysics Data System (ADS)

    Yang, Jing; Zhu, Yingnan; Xu, Tong; Pan, Chao; Cai, Nana; Huang, He; Zhang, Lei

    2016-07-01

    Biomedical applications of living cells have rapidly expanded in many fields such as toxic detection, drug screening, and regenerative medicine, etc. Efficient methods to support cell survival and maintain activity in vitro have become increasingly important. However, traditional cryopreservation for living cell-based applications is limited by several problems. Here, we report that magnetic hydrogel microparticles can physically assemble into a 3D environment for efficient cell preservation in physiological conditions, avoiding any chemical reactions that would damage the cells. Two representative cell lines (loosely and firmly adherent) were tested to evaluate the versatility of this method. The results showed that cell longevity was significantly extended to at least 15 days, while the control cell samples without microparticles quickly died within 3 days. Moreover, after preservation, cells can be easily retrieved by applying a magnet to separate the magnetic particles. This strategy can also inhibit cell over-proliferation while avoiding the use of temperature extremes or toxic cryoprotectants that are essential in cryopreservation.

  10. An enkephalin degrading aminopeptidase of human brain preserved during the vertebrate phylogeny.

    PubMed

    de Souza, A N; Bruno, J A; Carvalho, K M

    1991-01-01

    1. A soluble human brain aminopeptidase which hydrolyses the Tyr-Gly bond of Met-enkephalin and Leu-enkephalin was identified in the brains of the following vertebrates: mammals (Callithrix jacchus and Rattus norvegicus), bird (Gallus domesticus), reptile (Tupinambis teguixin), amphibia (Bufo paracnemis), fish (Sarotherdon niloticus) and elasmobranchy (Galeocerdo cuvieri). 2. The properties of this enzyme are: molecular weight near 100,000 Da, isoelectric point near 4.9, optimum pH near 7.5, activation by dithiothreitol, strong inhibition by Cu2+, Zn2+, Ni2+, puromycin and bacitracin, hydrolysis of enkephalins and basic and neutral aminoacid-beta-naphythylamide substrates. 3. The results indicate the preservation of this human brain aminopeptidase during the course of vertebrate phylogeny.

  11. PCR amplification of microsatellites from single cells of Karenia brevis preserved in Lugol's iodine solution.

    PubMed

    Henrichs, D W; Renshaw, M A; Santamaria, C A; Richardson, B; Gold, J R; Campbell, L

    2008-01-01

    A simple and effective protocol is described for multiplex polymerase chain reaction (PCR) amplification of single cells of Karenia brevis. The protocol requires minimum processing, avoids additions that might dilute target DNA template, and can be used on cells preserved in Lugol's iodine preservative. Destaining of Lugol's-preserved cells with sodium thiosulfate allowed successful amplification of single-copy, nuclear-encoded microsatellites in single cells of K. brevis that have been preserved for up to 6 years.

  12. Apicobasal polarity of brain endothelial cells

    PubMed Central

    Worzfeld, Thomas

    2015-01-01

    Normal brain homeostasis depends on the integrity of the blood–brain barrier that controls the access of nutrients, humoral factors, and immune cells to the CNS. The blood–brain barrier is composed mainly of brain endothelial cells. Forming the interface between two compartments, they are highly polarized. Apical/luminal and basolateral/abluminal membranes differ in their lipid and (glyco-)protein composition, allowing brain endothelial cells to secrete or transport soluble factors in a polarized manner and to maintain blood flow. Here, we summarize the basic concepts of apicobasal cell polarity in brain endothelial cells. To address potential molecular mechanisms underlying apicobasal polarity in brain endothelial cells, we draw on investigations in epithelial cells and discuss how polarity may go awry in neurological diseases. PMID:26661193

  13. Ebselen Preserves Tissue-Engineered Cell Sheets and their Stem Cells in Hypothermic Conditions

    PubMed Central

    Katori, Ryosuke; Hayashi, Ryuhei; Kobayashi, Yuki; Kobayashi, Eiji; Nishida, Kohji

    2016-01-01

    Clinical trials have been performed using autologous tissue-engineered epithelial cell sheets for corneal regenerative medicine. To improve stem cell-based therapy for convenient clinical practice, new techniques are required for preserving reconstructed tissues and their stem/progenitor cells until they are ready for use. In the present study, we screened potential preservative agents and developed a novel medium for preserving the cell sheets and their stem/progenitor cells; the effects were evaluated with a luciferase-based viability assay. Nrf2 activators, specifically ebselen, could maintain high ATP levels during preservation. Ebselen also showed a strong influence on maintenance of the viability, morphology, and stem cell function of the cell sheets preserved under hypothermia by protecting them from reactive oxygen species-induced damage. Furthermore, ebselen drastically improved the preservation performance of human cornea tissues and their stem cells. Therefore, ebselen shows good potential as a useful preservation agent in regenerative medicine as well as in cornea transplantation. PMID:27966584

  14. [Stem cells of mammalian brain: biology of the stem cells in vivo and in vitro].

    PubMed

    Viktorov, I V

    2001-01-01

    Stem cells are totipotent cells of the blastocyst (embryonal stem cells) and multipotent germinative cells of ento-, ecto-, and mesoderm that give rise to all tissues during embryogenesis. The stem cells have high proliferation activity and an unlimited capacity for self-production by symmetrical mitosis. Asymmetrical mitosis of the stem cells generates daughter cells ("progenitor cells") with unlimited proliferation potential. During differentiation, the progenitor cells give rise to definitive somatic cells. The stem and progenitor cells are preserved in most tissues of adult organism and provide for the constant replacement of the cells after their physiological death and damage. At the end of last century, stem cells were found in the brain of the adult mouse and rat and later in the brain of other mammals including humans. The subependymal zone of the lateral ventricles is considered the site of stem cells localization; however, there are indications of stem cells origination from ependyma while the subependymal zone serves as a collector of the progenitor cells where these cells divide. The problem of the localization of stem cells in a mature brain has not yet been resolved and is actively discussed. The stem and progenitor cells, as well as neuro- and gliogenesis, are most explored in the hippocampus and olfactory bulb. The progenitor cells migrate to the olfactory bulb from the subependymal zone of the lateral ventricles via a rostral migratory stream formed by the astrocytes, and then they differentiate to neural and glial cells. In the hippocampus, the neurons are formed in the subgranular zone of dentate gyrus. The discovery of stem and progenitor cells in the mature brain and their subsequent investigation point to an ongoing neuro- and gliogenesis in all periventricular sections of the brain and spinal cord during the whole animal or human lifespan. These processes proved to be related to the functional condition of CNS, and the de novo formed neural

  15. Specimen Sample Preservation for Cell and Tissue Cultures

    NASA Technical Reports Server (NTRS)

    Meeker, Gabrielle; Ronzana, Karolyn; Schibner, Karen; Evans, Robert

    1996-01-01

    The era of the International Space Station with its longer duration missions will pose unique challenges to microgravity life sciences research. The Space Station Biological Research Project (SSBRP) is responsible for addressing these challenges and defining the science requirements necessary to conduct life science research on-board the International Space Station. Space Station will support a wide range of cell and tissue culture experiments for durations of 1 to 30 days. Space Shuttle flights to bring experimental samples back to Earth for analyses will only occur every 90 days. Therefore, samples may have to be retained for periods up to 60 days. This presents a new challenge in fresh specimen sample storage for cell biology. Fresh specimen samples are defined as samples that are preserved by means other than fixation and cryopreservation. The challenge of long-term storage of fresh specimen samples includes the need to suspend or inhibit proliferation and metabolism pending return to Earth-based laboratories. With this challenge being unique to space research, there have not been any ground based studies performed to address this issue. It was decided hy SSBRP that experiment support studies to address the following issues were needed: Fixative Solution Management; Media Storage Conditions; Fresh Specimen Sample Storage of Mammalian Cell/Tissue Cultures; Fresh Specimen Sample Storage of Plant Cell/Tissue Cultures; Fresh Specimen Sample Storage of Aquatic Cell/Tissue Cultures; and Fresh Specimen Sample Storage of Microbial Cell/Tissue Cultures. The objective of these studies was to derive a set of conditions and recommendations that can be used in a long duration microgravity environment such as Space Station that will permit extended storage of cell and tissue culture specimens in a state consistent with zero or minimal growth, while at the same time maintaining their stability and viability.

  16. Structural and functional brain rewiring clarifies preserved interhemispheric transfer in humans born without the corpus callosum

    PubMed Central

    Tovar-Moll, Fernanda; Monteiro, Myriam; Andrade, Juliana; Bramati, Ivanei E.; Vianna-Barbosa, Rodrigo; Marins, Theo; Rodrigues, Erika; Dantas, Natalia; Behrens, Timothy E. J.; de Oliveira-Souza, Ricardo; Moll, Jorge; Lent, Roberto

    2014-01-01

    Why do humans born without the corpus callosum, the major interhemispheric commissure, lack the disconnection syndrome classically described in callosotomized patients? This paradox was discovered by Nobel laureate Roger Sperry in 1968, and has remained unsolved since then. To tackle the hypothesis that alternative neural pathways could explain this puzzle, we investigated patients with callosal dysgenesis using structural and functional neuroimaging, as well as neuropsychological assessments. We identified two anomalous white-matter tracts by deterministic and probabilistic tractography, and provide supporting resting-state functional neuroimaging and neuropsychological evidence for their functional role in preserved interhemispheric transfer of complex tactile information, such as object recognition. These compensatory pathways connect the homotopic posterior parietal cortical areas (Brodmann areas 39 and surroundings) via the posterior and anterior commissures. We propose that anomalous brain circuitry of callosal dysgenesis is determined by long-distance plasticity, a set of hardware changes occurring in the developing brain after pathological interference. So far unknown, these pathological changes somehow divert growing axons away from the dorsal midline, creating alternative tracts through the ventral forebrain and the dorsal midbrain midline, with partial compensatory effects to the interhemispheric transfer of cortical function. PMID:24821757

  17. Neural stem cell transplantation in mouse brain.

    PubMed

    Lee, Jean-Pyo; McKercher, Scott; Muller, Franz-Josef; Snyder, Evan Y

    2008-01-01

    Neural stem cells (NSCs) are the most primordial, least committed cells of the nervous system, and transplantation of these multipotent cells holds the promise of regenerative therapy for many central nervous system (CNS) diseases. This unit describes methods for NSC transplantation into neonatal mouse pups, embryonic mouse brain, and adult mouse brain. A description of options for detection of labeled donor cells in engrafted mouse brain is provided along with an example protocol for detecting lacZ-expressing cells in situ. Also included is a protocol for preparing NSCs for transplantation.

  18. 2-Methoxystypandrone ameliorates brain function through preserving BBB integrity and promoting neurogenesis in mice with acute ischemic stroke.

    PubMed

    Chern, Chang-Ming; Wang, Yea-Hwey; Liou, Kuo-Tong; Hou, Yu-Chang; Chen, Chien-Chih; Shen, Yuh-Chiang

    2014-02-01

    2-Methoxystypandrone (2-MS), a naphthoquinone, has been shown to display an immunomodulatory effect in a cellular model. To explore whether 2-MS could protect mice against cerebral ischemic/reperfusion (I/R)-induced brain injury, we evaluated 2-MS's protective effects on an acute ischemic stroke by inducing a middle cerebral artery occlusion/reperfusion (MCAO) injury in murine model. Treatment of mice that have undergone I/R injury with 2-MS (10-100 μg/kg, i.v.) at 2 h after MCAO enhanced survival rate and ameliorated neurological deficits, brain infarction, neural dysfunction and massive oxidative stress, due to an enormous production of free radicals and breakdown of blood-brain barrier (BBB) by I/R injury; this primarily occurred with extensive infiltration of CD11b-positive inflammatory cells and upexpression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 and p65 nuclear factor-kappa B (NF-κB). All of these pathological changes were diminished by 2-MS; 2-MS also intensively limited cortical infarction and promoted upexpression of neurodevelopmental genes near peri-infarct cortex and endogenous neurogenesis near subgranular zone of hippocampal dentate gyrus and the subventricular zone, most possibly by inactivation of GSK3β which in turn upregulating β-catenin, Bcl-2 adam11 and adamts20. We conclude that 2-MS blocks inflammatory responses by impairing NF-κB signaling to limit the inflammation and oxidative stress for preservation of BBB integrity; 2-MS also concomitantly promotes neurodevelopmental protein expression and endogenous neurogenesis through inactivation of GSK3β to enhance β-catenin signaling for upexpression of neuroprotective genes and proteins.

  19. Early Shifts of Brain Metabolism by Caloric Restriction Preserve White Matter Integrity and Long-Term Memory in Aging Mice.

    PubMed

    Guo, Janet; Bakshi, Vikas; Lin, Ai-Ling

    2015-01-01

    Preservation of brain integrity with age is highly associated with lifespan determination. Caloric restriction (CR) has been shown to increase longevity and healthspan in various species; however, its effects on preserving living brain functions in aging remain largely unexplored. In the study, we used multimodal, non-invasive neuroimaging (PET/MRI/MRS) to determine in vivo brain glucose metabolism, energy metabolites, and white matter structural integrity in young and old mice fed with either control or 40% CR diet. In addition, we determined the animals' memory and learning ability with behavioral assessments. Blood glucose, blood ketone bodies, and body weight were also measured. We found distinct patterns between normal aging and CR aging on brain functions - normal aging showed reductions in brain glucose metabolism, white matter integrity, and long-term memory, resembling human brain aging. CR aging, in contrast, displayed an early shift from glucose to ketone bodies metabolism, which was associated with preservations of brain energy production, white matter integrity, and long-term memory in aging mice. Among all the mice, we found a positive correlation between blood glucose level and body weight, but an inverse association between blood glucose level and lifespan. Our findings suggest that CR could slow down brain aging, in part due to the early shift of energy metabolism caused by lower caloric intake, and we were able to identify the age-dependent effects of CR non-invasively using neuroimaging. These results provide a rationale for CR-induced sustenance of brain health with extended longevity.

  20. Early Shifts of Brain Metabolism by Caloric Restriction Preserve White Matter Integrity and Long-Term Memory in Aging Mice

    PubMed Central

    Guo, Janet; Bakshi, Vikas; Lin, Ai-Ling

    2015-01-01

    Preservation of brain integrity with age is highly associated with lifespan determination. Caloric restriction (CR) has been shown to increase longevity and healthspan in various species; however, its effects on preserving living brain functions in aging remain largely unexplored. In the study, we used multimodal, non-invasive neuroimaging (PET/MRI/MRS) to determine in vivo brain glucose metabolism, energy metabolites, and white matter structural integrity in young and old mice fed with either control or 40% CR diet. In addition, we determined the animals’ memory and learning ability with behavioral assessments. Blood glucose, blood ketone bodies, and body weight were also measured. We found distinct patterns between normal aging and CR aging on brain functions – normal aging showed reductions in brain glucose metabolism, white matter integrity, and long-term memory, resembling human brain aging. CR aging, in contrast, displayed an early shift from glucose to ketone bodies metabolism, which was associated with preservations of brain energy production, white matter integrity, and long-term memory in aging mice. Among all the mice, we found a positive correlation between blood glucose level and body weight, but an inverse association between blood glucose level and lifespan. Our findings suggest that CR could slow down brain aging, in part due to the early shift of energy metabolism caused by lower caloric intake, and we were able to identify the age-dependent effects of CR non-invasively using neuroimaging. These results provide a rationale for CR-induced sustenance of brain health with extended longevity. PMID:26617514

  1. Melatonin Preserves Blood-Brain Barrier Integrity and Permeability via Matrix Metalloproteinase-9 Inhibition

    PubMed Central

    Alluri, Himakarnika; Wilson, Rickesha L.; Anasooya Shaji, Chinchusha; Wiggins-Dohlvik, Katie; Patel, Savan; Liu, Yang; Peng, Xu; Beeram, Madhava R.; Davis, Matthew L.; Huang, Jason H.; Tharakan, Binu

    2016-01-01

    Microvascular hyperpermeability that occurs at the level of the blood-brain barrier (BBB) often leads to vasogenic brain edema and elevated intracranial pressure following traumatic brain injury (TBI). At a cellular level, tight junction proteins (TJPs) between neighboring endothelial cells maintain the integrity of the BBB via TJ associated proteins particularly, zonula occludens-1 (ZO-1) that binds to the transmembrane TJPs and actin cytoskeleton intracellularly. The pro-inflammatory cytokine, interleukin-1β (IL-1β) as well as the proteolytic enzymes, matrix metalloproteinase-9 (MMP-9) are key mediators of trauma-associated brain edema. Recent studies indicate that melatonin a pineal hormone directly binds to MMP-9 and also might act as its endogenous inhibitor. We hypothesized that melatonin treatment will provide protection against TBI-induced BBB hyperpermeability via MMP-9 inhibition. Rat brain microvascular endothelial cells grown as monolayers were used as an in vitro model of the BBB and a mouse model of TBI using a controlled cortical impactor was used for all in vivo studies. IL-1β (10 ng/mL; 2 hours)-induced endothelial monolayer hyperpermeability was significantly attenuated by melatonin (10 μg/mL; 1 hour), GM6001 (broad spectrum MMP inhibitor; 10 μM; 1 hour), MMP-9 inhibitor-1 (MMP-9 specific inhibitor; 5 nM; 1 hour) or MMP-9 siRNA transfection (48 hours) in vitro. Melatonin and MMP-9 inhibitor-1 pretreatment attenuated IL-1β-induced MMP-9 activity, loss of ZO-1 junctional integrity and f-actin stress fiber formation. IL-1β treatment neither affected ZO-1 protein or mRNA expression or cell viability. Acute melatonin treatment attenuated BBB hyperpermeability in a mouse controlled cortical impact model of TBI in vivo. In conclusion, one of the protective effects of melatonin against BBB hyperpermeability occurs due to enhanced BBB integrity via MMP-9 inhibition. In addition, acute melatonin treatment provides protection against BBB

  2. The PPARalpha Agonist Fenofibrate Preserves Hippocampal Neurogenesis and Inhibits Microglial Activation After Whole-Brain Irradiation

    SciTech Connect

    Ramanan, Sriram; Kooshki, Mitra; Zhao Weiling; Hsu, F.-C.; Riddle, David R.; Robbins, Mike E.

    2009-11-01

    Purpose: Whole-brain irradiation (WBI) leads to cognitive impairment months to years after radiation. Numerous studies suggest that decreased hippocampal neurogenesis and microglial activation are involved in the pathogenesis of WBI-induced brain injury. The goal of this study was to investigate whether administration of the peroxisomal proliferator-activated receptor (PPAR) alpha agonist fenofibrate would prevent the detrimental effect of WBI on hippocampal neurogenesis. Methods and Materials: For this study, 129S1/SvImJ wild-type and PPARalpha knockout mice that were fed either regular or 0.2% wt/wt fenofibrate-containing chow received either sham irradiation or WBI (10-Gy single dose of {sup 137}Cs gamma-rays). Mice were injected intraperitoneally with bromodeoxyuridine to label the surviving cells at 1 month after WBI, and the newborn neurons were counted at 2 months after WBI by use of bromodeoxyuridine/neuronal nuclei double immunofluorescence. Proliferation in the subgranular zone and microglial activation were measured at 1 week and 2 months after WBI by use of Ki-67 and CD68 immunohistochemistry, respectively. Results: Whole-brain irradiation led to a significant decrease in the number of newborn hippocampal neurons 2 months after it was performed. Fenofibrate prevented this decrease by promoting the survival of newborn cells in the dentate gyrus. In addition, fenofibrate treatment was associated with decreased microglial activation in the dentate gyrus after WBI. The neuroprotective effects of fenofibrate were abolished in the knockout mice, indicating a PPARalpha-dependent mechanism or mechanisms. Conclusions: These data highlight a novel role for PPARalpha ligands in improving neurogenesis after WBI and offer the promise of improving the quality of life for brain cancer patients receiving radiotherapy.

  3. New Nerve Cells for the Adult Brain.

    ERIC Educational Resources Information Center

    Kempermann, Gerd; Gage, Fred H.

    1999-01-01

    Contrary to dogma, the human brain does produce new nerve cells in adulthood. The mature human brain spawns neurons routinely in the hippocampus, an area important to memory and learning. This research can make it possible to ease any number of disorders involving neurological damage and death. (CCM)

  4. Analysis of the brain mural cell transcriptome

    PubMed Central

    He, Liqun; Vanlandewijck, Michael; Raschperger, Elisabeth; Andaloussi Mäe, Maarja; Jung, Bongnam; Lebouvier, Thibaud; Ando, Koji; Hofmann, Jennifer; Keller, Annika; Betsholtz, Christer

    2016-01-01

    Pericytes, the mural cells of blood microvessels, regulate microvascular development and function and have been implicated in many brain diseases. However, due to a paucity of defining markers, pericyte identification and functional characterization remain ambiguous and data interpretation problematic. In mice carrying two transgenic reporters, Pdgfrb-eGFP and NG2-DsRed, we found that double-positive cells were vascular mural cells, while the single reporters marked additional, but non-overlapping, neuroglial cells. Double-positive cells were isolated by fluorescence-activated cell sorting (FACS) and analyzed by RNA sequencing. To reveal defining patterns of mural cell transcripts, we compared the RNA sequencing data with data from four previously published studies. The meta-analysis provided a conservative catalogue of 260 brain mural cell-enriched gene transcripts. We validated pericyte-specific expression of two novel markers, vitronectin (Vtn) and interferon-induced transmembrane protein 1 (Ifitm1), using fluorescent in situ hybridization and immunohistochemistry. We further analyzed signaling pathways and interaction networks of the pericyte-enriched genes in silico. This work provides novel insight into the molecular composition of brain mural cells. The reported gene catalogue facilitates identification of brain pericytes by providing numerous new candidate marker genes and is a rich source for new hypotheses for future studies of brain mural cell physiology and pathophysiology. PMID:27725773

  5. Key issues relating to the genetic stability and preservation of cells and cell banks.

    PubMed

    Simione, F P

    1992-01-01

    The long term maintenance of genetically stable cells is important for ensuring reproducible results and continuity in the advance of microbiology, cell biology and biotechnology. As actively growing cultures, cells are constantly at risk of changing, and the necessity for subculturing living materials increases the chances for genetic change and contamination. Many techniques are available for stabilizing living cells; the method employed must be compatible with the intended use of the culture. The most commonly utilized means of preserving living cells are by freezing to cryogenic temperatures, and freeze-drying. Master stocks are usually maintained at liquid nitrogen or comparable temperatures, while working stocks can be frozen or freeze-dried, and maintained at more economical and easily managed temperatures where possible. However, low temperature techniques may cause damage that can result in genetic change, or potential selection when only a small portion of the population survives. Therefore, a good preservation program must include a comprehensive cell characterization regimen that is applied both before and after preserving the cells to ensure that changes are detected when they do occur. Assurance of long term stability necessitates well designed safekeeping and security measures that include minimizing specimen handling through well designed inventory systems, validation and monitoring of storage temperatures, provisions for backup inventory, and training of personnel. Cell banking also requires good cataloguing and data management practices to avoid duplication and misidentification, and to ensure proper tracking of specimens and ease of access.

  6. Histological Architecture Underlying Brain-Immune Cell-Cell Interactions and the Cerebral Response to Systemic Inflammation.

    PubMed

    Shimada, Atsuyoshi; Hasegawa-Ishii, Sanae

    2017-01-01

    Although the brain is now known to actively interact with the immune system under non-inflammatory conditions, the site of cell-cell interactions between brain parenchymal cells and immune cells has been an open question until recently. Studies by our and other groups have indicated that brain structures such as the leptomeninges, choroid plexus stroma and epithelium, attachments of choroid plexus, vascular endothelial cells, cells of the perivascular space, circumventricular organs, and astrocytic endfeet construct the histological architecture that provides a location for intercellular interactions between bone marrow-derived myeloid lineage cells and brain parenchymal cells under non-inflammatory conditions. This architecture also functions as the interface between the brain and the immune system, through which systemic inflammation-induced molecular events can be relayed to the brain parenchyma at early stages of systemic inflammation during which the blood-brain barrier is relatively preserved. Although brain microglia are well known to be activated by systemic inflammation, the mechanism by which systemic inflammatory challenge and microglial activation are connected has not been well documented. Perturbed brain-immune interaction underlies a wide variety of neurological and psychiatric disorders including ischemic brain injury, status epilepticus, repeated social defeat, and neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. Proinflammatory status associated with cytokine imbalance is involved in autism spectrum disorders, schizophrenia, and depression. In this article, we propose a mechanism connecting systemic inflammation, brain-immune interface cells, and brain parenchymal cells and discuss the relevance of basic studies of the mechanism to neurological disorders with a special emphasis on sepsis-associated encephalopathy and preterm brain injury.

  7. Preserving GABAergic interneurons in acute brain slices of mice using the N-methyl-D-glucamine-based artificial cerebrospinal fluid method.

    PubMed

    Pan, Geng; Li, Yue; Geng, Hong-Yan; Yang, Jian-Ming; Li, Ke-Xin; Li, Xiao-Ming

    2015-04-01

    Defects in the function and development of GABAergic interneurons have been linked to psychiatric disorders, so preservation of these interneurons in brain slices is important for successful electrophysiological recording in various ex vivo methods. However, it is difficult to maintain the activity and morphology of neurons in slices from mice of >30 days old. Here we evaluated the N-methyl-D-glucamine (NMDG)-based artificial cerebrospinal fluid (aCSF) method for the preservation of interneurons in slices from mice of up to ∼6 months old and discussed the steps that may affect their quality during slicing. We found that the NMDG-aCSF method rescued more cells than sucrose-aCSF and successfully preserved different types of interneurons including parvalbumin- and somatostatin-positive interneurons. In addition, both the chemical and electrical synaptic signaling of interneurons were maintained. These results demonstrate that the NMDG-aCSF method is suitable for the preservation of interneurons, especially in studies of gap junctions.

  8. Youthful Brains in Older Adults: Preserved Neuroanatomy in the Default Mode and Salience Networks Contributes to Youthful Memory in Superaging

    PubMed Central

    Sun, Felicia W.; Stepanovic, Michael R.; Andreano, Joseph

    2016-01-01

    Decline in cognitive skills, especially in memory, is often viewed as part of “normal” aging. Yet some individuals “age better” than others. Building on prior research showing that cortical thickness in one brain region, the anterior midcingulate cortex, is preserved in older adults with memory performance abilities equal to or better than those of people 20–30 years younger (i.e., “superagers”), we examined the structural integrity of two large-scale intrinsic brain networks in superaging: the default mode network, typically engaged during memory encoding and retrieval tasks, and the salience network, typically engaged during attention, motivation, and executive function tasks. We predicted that superagers would have preserved cortical thickness in critical nodes in these networks. We defined superagers (60–80 years old) based on their performance compared to young adults (18–32 years old) on the California Verbal Learning Test Long Delay Free Recall test. We found regions within the networks of interest where the cerebral cortex of superagers was thicker than that of typical older adults, and where superagers were anatomically indistinguishable from young adults; hippocampal volume was also preserved in superagers. Within the full group of older adults, thickness of a number of regions, including the anterior temporal cortex, rostral medial prefrontal cortex, and anterior midcingulate cortex, correlated with memory performance, as did the volume of the hippocampus. These results indicate older adults with youthful memory abilities have youthful brain regions in key paralimbic and limbic nodes of the default mode and salience networks that support attentional, executive, and mnemonic processes subserving memory function. SIGNIFICANCE STATEMENT Memory performance typically declines with age, as does cortical structural integrity, yet some older adults maintain youthful memory. We tested the hypothesis that superagers (older individuals with

  9. Should deciduous teeth be preserved in adult patients? How about stem cells? Is it reasonable to preserve them?

    PubMed Central

    Consolaro, Alberto

    2016-01-01

    Abstract When seeking orthodontic treatment, many adolescents and adult patients present with deciduous teeth. Naturally, deciduous teeth will inevitably undergo exfoliation at the expected time or at a later time. Apoptosis is the biological trigger of root resorption. In adult patients, deciduous teeth should not be preserved, as they promote: infraocclusion, traumatic occlusion, occlusal trauma, diastemata and size as well as morphology discrepancy malocclusion. Orthodontic movement speeds root resorption up, and so do restoring or recontouring deciduous teeth in order to establish esthetics and function. Deciduous teeth cells are dying as a result of apoptosis, and their regeneration potential, which allows them to act as stem cells, is limited. On the contrary, adult teeth cells have a greater proliferative potential. All kinds of stem cell therapies are laboratory investigative non authorized trials. PMID:27275612

  10. Should deciduous teeth be preserved in adult patients? How about stem cells? Is it reasonable to preserve them?

    PubMed

    Consolaro, Alberto

    2016-01-01

    When seeking orthodontic treatment, many adolescents and adult patients present with deciduous teeth. Naturally, deciduous teeth will inevitably undergo exfoliation at the expected time or at a later time. Apoptosis is the biological trigger of root resorption. In adult patients, deciduous teeth should not be preserved, as they promote: infraocclusion, traumatic occlusion, occlusal trauma, diastemata and size as well as morphology discrepancy malocclusion. Orthodontic movement speeds root resorption up, and so do restoring or recontouring deciduous teeth in order to establish esthetics and function. Deciduous teeth cells are dying as a result of apoptosis, and their regeneration potential, which allows them to act as stem cells, is limited. On the contrary, adult teeth cells have a greater proliferative potential. All kinds of stem cell therapies are laboratory investigative non authorized trials.

  11. Chronic alcoholism in rats induces a compensatory response, preserving brain thiamine diphosphate, but the brain 2-oxo acid dehydrogenases are inactivated despite unchanged coenzyme levels.

    PubMed

    Parkhomenko, Yulia M; Kudryavtsev, Pavel A; Pylypchuk, Svetlana Yu; Chekhivska, Lilia I; Stepanenko, Svetlana P; Sergiichuk, Andrej A; Bunik, Victoria I

    2011-06-01

    Thiamine-dependent changes in alcoholic brain were studied using a rat model. Brain thiamine and its mono- and diphosphates were not reduced after 20 weeks of alcohol exposure. However, alcoholism increased both synaptosomal thiamine uptake and thiamine diphosphate synthesis in brain, pointing to mechanisms preserving thiamine diphosphate in the alcoholic brain. In spite of the unchanged level of the coenzyme thiamine diphosphate, activities of the mitochondrial 2-oxoglutarate and pyruvate dehydrogenase complexes decreased in alcoholic brain. The inactivation of pyruvate dehydrogenase complex was caused by its increased phosphorylation. The inactivation of 2-oxoglutarate dehydrogenase complex (OGDHC) correlated with a decrease in free thiols resulting from an elevation of reactive oxygen species. Abstinence from alcohol following exposure to alcohol reactivated OGDHC along with restoration of the free thiol content. However, restoration of enzyme activity occurred before normalization of reactive oxygen species levels. Hence, the redox status of cellular thiols mediates the action of oxidative stress on OGDHC in alcoholic brain. As a result, upon chronic alcohol consumption, physiological mechanisms to counteract the thiamine deficiency and silence pyruvate dehydrogenase are activated in rat brain, whereas OGDHC is inactivated due to impaired antioxidant ability.

  12. Bio-inspired cryo-ink preserves red blood cell phenotype and function during nanoliter vitrification.

    PubMed

    El Assal, Rami; Guven, Sinan; Gurkan, Umut Atakan; Gozen, Irep; Shafiee, Hadi; Dalbeyler, Sedef; Abdalla, Noor; Thomas, Gawain; Fuld, Wendy; Illigens, Ben M W; Estanislau, Jessica; Khoory, Joseph; Kaufman, Richard; Zylberberg, Claudia; Lindeman, Neal; Wen, Qi; Ghiran, Ionita; Demirci, Utkan

    2014-09-03

    Current red-blood-cell cryopreservation methods utilize bulk volumes, causing cryo-injury of cells, which results in irreversible disruption of cell morphology, mechanics, and function. An innovative approach to preserve human red-blood-cell morphology, mechanics, and function following vitrification in nanoliter volumes is developed using a novel cryo-ink integrated with a bioprinting approach.

  13. Bio-inspired Cryo-ink Preserves Red Blood Cell Phenotype and Function during Nanoliter Vitrification

    PubMed Central

    Assal, Rami El; Guven, Sinan; Gurkan, Umut Atakan; Gozen, Irep; Shafiee, Hadi; Dalbeyber, Sedef; Abdalla, Noor; Thomas, Gawain; Fuld, Wendy; Illigens, Ben M.W.; Estanislau, Jessica; Khoory, Joseph; Kaufman, Richard; Zylberberg, Claudia; Lindeman, Neal; Wen, Qi; Ghiran, Ionita; Demirci, Utkan

    2014-01-01

    Current red blood cell cryopreservation methods utilize bulk volumes, causing cryo-injury of cells, which results in irreversible disruption of cell morphology, mechanics, and function. An innovative approach to preserve human red blood cell morphology, mechanics, and function following vitrification in nanoliter volumes is developed using a novel cryo-ink integrated with a bio-printing approach. PMID:25047246

  14. Topology preserving non-rigid image registration using time-varying elasticity model for MRI brain volumes.

    PubMed

    Ahmad, Sahar; Khan, Muhammad Faisal

    2015-12-01

    In this paper, we present a new non-rigid image registration method that imposes a topology preservation constraint on the deformation. We propose to incorporate the time varying elasticity model into the deformable image matching procedure and constrain the Jacobian determinant of the transformation over the entire image domain. The motion of elastic bodies is governed by a hyperbolic partial differential equation, generally termed as elastodynamics wave equation, which we propose to use as a deformation model. We carried out clinical image registration experiments on 3D magnetic resonance brain scans from IBSR database. The results of the proposed registration approach in terms of Kappa index and relative overlap computed over the subcortical structures were compared against the existing topology preserving non-rigid image registration methods and non topology preserving variant of our proposed registration scheme. The Jacobian determinant maps obtained with our proposed registration method were qualitatively and quantitatively analyzed. The results demonstrated that the proposed scheme provides good registration accuracy with smooth transformations, thereby guaranteeing the preservation of topology.

  15. Endothelial Progenitor Cells Physiology and Metabolic Plasticity in Brain Angiogenesis and Blood-Brain Barrier Modeling

    PubMed Central

    Malinovskaya, Natalia A.; Komleva, Yulia K.; Salmin, Vladimir V.; Morgun, Andrey V.; Shuvaev, Anton N.; Panina, Yulia A.; Boitsova, Elizaveta B.; Salmina, Alla B.

    2016-01-01

    Currently, there is a considerable interest to the assessment of blood-brain barrier (BBB) development as a part of cerebral angiogenesis developmental program. Embryonic and adult angiogenesis in the brain is governed by the coordinated activity of endothelial progenitor cells, brain microvascular endothelial cells, and non-endothelial cells contributing to the establishment of the BBB (pericytes, astrocytes, neurons). Metabolic and functional plasticity of endothelial progenitor cells controls their timely recruitment, precise homing to the brain microvessels, and efficient support of brain angiogenesis. Deciphering endothelial progenitor cells physiology would provide novel engineering approaches to establish adequate microfluidically-supported BBB models and brain microphysiological systems for translational studies. PMID:27990124

  16. Oscar Wilde and the brain cell.

    PubMed

    Cohn, Elisha

    2013-01-01

    This chapter considers Oscar Wilde's interest in the brain cell as an aesthetic object. Offering an account of Wilde's career that analyzes his early interest in physiology and philosophy, this chapter argues that Wilde's uniquely aesthetic take on the brain suggests that he rejects an account of the self as autonomous or self-determining. For many late Victorians brain science threatened both the freedom of human action and the legitimacy of beauty because it had the potential to invalidate conscious experience. But writers whose work Wilde knew, like John Ruskin, W. K. Clifford, and John Tyndall, avoided the despair of materialism by using aesthetic terms in their own discussions of life's invisible materials. Wilde's art collaborates with the contemporary sciences. His depictions of the cell direct the senses to a new field of being that emphasizes the molecular life all humans have in common, in which individual responsibility and activity matter less than the necessity of beauty.

  17. Supercooling as a viable non-freezing cell preservation method of rat hepatocytes.

    PubMed

    Usta, O Berk; Kim, Yeonhee; Ozer, Sinan; Bruinsma, Bote G; Lee, Jungwoo; Demir, Esin; Berendsen, Tim A; Puts, Catheleyne F; Izamis, Maria-Louisa; Uygun, Korkut; Uygun, Basak E; Yarmush, Martin L

    2013-01-01

    Supercooling preservation holds the potential to drastically extend the preservation time of organs, tissues and engineered tissue products, and fragile cell types that do not lend themselves well to cryopreservation or vitrification. Here, we investigate the effects of supercooling preservation (SCP at -4(o)C) on primary rat hepatocytes stored in cryovials and compare its success (high viability and good functional characteristics) to that of static cold storage (CS at +4(o)C) and cryopreservation. We consider two prominent preservation solutions a) Hypothermosol (HTS-FRS) and b) University of Wisconsin solution (UW) and a range of preservation temperatures (-4 to -10 (o)C). We find that there exists an optimum temperature (-4(o)C) for SCP of rat hepatocytes which yields the highest viability; at this temperature HTS-FRS significantly outperforms UW solution in terms of viability and functional characteristics (secretions and enzymatic activity in suspension and plate culture). With the HTS-FRS solution we show that the cells can be stored for up to a week with high viability (~56%); moreover we also show that the preservation can be performed in large batches (50 million cells) with equal or better viability and no loss of functionality as compared to smaller batches (1.5 million cells) performed in cryovials.

  18. P2X7 Receptor Suppression Preserves Blood-Brain Barrier through Inhibiting RhoA Activation after Experimental Intracerebral Hemorrhage in Rats.

    PubMed

    Zhao, Hengli; Zhang, Xuan; Dai, Zhiqiang; Feng, Yang; Li, Qiang; Zhang, John H; Liu, Xin; Chen, Yujie; Feng, Hua

    2016-03-16

    Blockading P2X7 receptor(P2X7R) provides neuroprotection toward various neurological disorders, including stroke, traumatic brain injury, and subarachnoid hemorrhage. However, whether and how P2X7 receptor suppression protects blood-brain barrier(BBB) after intracerebral hemorrhage(ICH) remains unexplored. In present study, intrastriatal autologous-blood injection was used to mimic ICH in rats. Selective P2X7R inhibitor A438079, P2X7R agonist BzATP, and P2X7R siRNA were administrated to evaluate the effects of P2X7R suppression. Selective RhoA inhibitor C3 transferase was administered to clarify the involvement of RhoA. Post-assessments, including neurological deficits, Fluoro-Jade C staining, brain edema, Evans blue extravasation and fluorescence, western blot, RhoA activity assay and immunohistochemistry were performed. Then the key results were verified in collagenase induced ICH model. We found that endogenous P2X7R increased at 3 hrs after ICH with peak at 24 hrs, then returned to normal at 72 hrs after ICH. Enhanced immunoreactivity was observed on the neurovascular structure around hematoma at 24 hrs after ICH, along with perivascular astrocytes and endothelial cells. Both A438079 and P2X7R siRNA alleviated neurological deficits, brain edema, and BBB disruption after ICH, in association with RhoA activation and down-regulated endothelial junction proteins. However, BzATP abolished those effects. In addition, C3 transferase reduced brain injury and increased endothelial junction proteins' expression after ICH. These data indicated P2X7R suppression could preserve BBB integrity after ICH through inhibiting RhoA activation.

  19. Alginate-Encapsulation for the Improved Hypothermic Preservation of Human Adipose-Derived Stem Cells.

    PubMed

    Swioklo, Stephen; Constantinescu, Andrei; Connon, Che J

    2016-03-01

    Despite considerable progress within the cell therapy industry, unmet bioprocessing and logistical challenges associated with the storage and distribution of cells between sites of manufacture and the clinic exist. We examined whether hypothermic (4°C-23°C) preservation of human adipose-derived stem cells could be improved through their encapsulation in 1.2% calcium alginate. Alginate encapsulation improved the recovery of viable cells after 72 hours of storage. Viable cell recovery was highly temperature-dependent, with an optimum temperature of 15°C. At this temperature, alginate encapsulation preserved the ability for recovered cells to attach to tissue culture plastic on rewarming, further increasing its effect on total cell recovery. On attachment, the cells were phenotypically normal, displayed normal growth kinetics, and maintained their capacity for trilineage differentiation. The number of cells encapsulated (up to 2 × 10(6) cells per milliliter) did not affect viable cell recovery nor did storage of encapsulated cells in a xeno-free, serum-free,current Good Manufacturing Practice-grade medium. We present a simple, low-cost system capable of enhancing the preservation of human adipose-derived stem cells stored at hypothermic temperatures, while maintaining their normal function. The storage of cells in this manner has great potential for extending the time windows for quality assurance and efficacy testing, distribution between the sites of manufacture and the clinic, and reducing the wastage associated with the limited shelf life of cells stored in their liquid state.

  20. A Lesion-Proof Brain? Multidimensional Sensorimotor, Cognitive, and Socio-Affective Preservation Despite Extensive Damage in a Stroke Patient

    PubMed Central

    García, Adolfo M.; Sedeño, Lucas; Herrera Murcia, Eduar; Couto, Blas; Ibáñez, Agustín

    2017-01-01

    In this study, we report an unusual case of mutidimensional sensorimotor, cognitive, and socio-affective preservation in an adult with extensive, acquired bilateral brain damage. At age 43, patient CG sustained a cerebral hemorrhage and a few months later, she suffered a second (ischemic) stroke. As a result, she exhibited extensive damage of the right hemisphere (including frontal, temporal, parietal, and occipital regions), left Sylvian and striatal areas, bilateral portions of the insula and the amygdala, and the splenium. However, against all probability, she was unimpaired across a host of cognitive domains, including executive functions, attention, memory, language, sensory perception (e.g., taste recognition and intensity discrimination), emotional processing (e.g., experiencing of positive and negative emotions), and social cognition skills (prosody recognition, theory of mind, facial emotion recognition, and emotional evaluation). Her functional integrity was further confirmed through neurological examination and contextualized observation of her performance in real-life tasks. In sum, CG's case resists straightforward classifications, as the extent and distribution of her lesions would typically produce pervasive, multidimensional deficits. We discuss the rarity of this patient against the backdrop of other reports of atypical cognitive preservation, expound the limitations of several potential accounts, and highlight the challenges that the case poses for current theories of brain organization and resilience. PMID:28119603

  1. Spermatogonial stem cells as a therapeutic alternative for fertility preservation of prepubertal boys

    PubMed Central

    Galuppo, Andrea Giannotti

    2015-01-01

    ABSTRACT Spermatogonial stem cells, which exist in the testicles since birth, are progenitors cells of male gametes. These cells are critical for the process of spermatogenesis, and not able to produce mature sperm cells before puberty due to their dependency of hormonal stimuli. This characteristic of the reproductive system limits the preservation of fertility only to males who are able to produce an ejaculate. This fact puts some light on the increase in survival rates of childhood cancer over the past decades because of improvements in the diagnosis and effective treatment in pediatric cancer patients. Therefore, we highlight one of the most important challenges concerning male fertility preservation that is the toxic effect of cancer therapy on reproductive function, especially the spermatogenesis. Currently, the experimental alternative for fertility preservation of prepubertal boys is the testicular tissue cryopreservationfor, for future isolation and spermatogonial stem cells transplantation, in order to restore the spermatogenesis. We present a brief review on isolation, characterization and culture conditions for the in vitro proliferation of spermatogonial stem cells, as well as the future perspectives as an alternative for fertility preservation in prepubertal boys. The possibility of restoring male fertility constitutes a research tool with an huge potential in basic and applied science. The development of these techniques may be a hope for the future of fertility preservation in cases that no other options exist, e.g, pediatric cancer patients. PMID:26761559

  2. Ektacytometry: Instrumentation and Applications in Red Blood Cell Preservation Studies.

    DTIC Science & Technology

    1982-05-10

    that it was necessary to maintain an isotonic environment, since the red blood cells respond differently to shear stress under hypertonic or...the red blood cells respond differently to shear stress under hypertonic or hypotonic conditions. The most unexpected observation was an absence of...required to compare samples analyzed the same day. 2. Treatment of Human Red Blood Cells With Hypotonic and Hypertonic Solutions and Glutaraldehyde

  3. Treatment of Severe Adult Traumatic Brain Injury Using Bone Marrow Mononuclear Cells.

    PubMed

    Cox, Charles S; Hetz, Robert A; Liao, George P; Aertker, Benjamin M; Ewing-Cobbs, Linda; Juranek, Jenifer; Savitz, Sean I; Jackson, Margaret L; Romanowska-Pawliczek, Anna M; Triolo, Fabio; Dash, Pramod K; Pedroza, Claudia; Lee, Dean A; Worth, Laura; Aisiku, Imoigele P; Choi, Huimahn A; Holcomb, John B; Kitagawa, Ryan S

    2017-04-01

    Preclinical studies using bone marrow derived cells to treat traumatic brain injury have demonstrated efficacy in terms of blood-brain barrier preservation, neurogenesis, and functional outcomes. Phase 1 clinical trials using bone marrow mononuclear cells infused intravenously in children with severe traumatic brain injury demonstrated safety and potentially a central nervous system structural preservation treatment effect. This study sought to confirm the safety, logistic feasibility, and potential treatment effect size of structural preservation/inflammatory biomarker mitigation in adults to guide Phase 2 clinical trial design. Adults with severe traumatic brain injury (Glasgow Coma Scale 5-8) and without signs of irreversible brain injury were evaluated for entry into the trial. A dose escalation format was performed in 25 patients: 5 controls, followed 5 patients in each dosing cohort (6, 9, 12 ×10(6) cells/kg body weight), then 5 more controls. Bone marrow harvest, cell processing to isolate the mononuclear fraction, and re-infusion occurred within 48 hours after injury. Patients were monitored for harvest-related hemodynamic changes, infusional toxicity, and adverse events. Outcome measures included magnetic resonance imaging-based measurements of supratentorial and corpus callosal volumes as well as diffusion tensor imaging-based measurements of fractional anisotropy and mean diffusivity of the corpus callosum and the corticospinal tract at the level of the brainstem at 1 month and 6 months postinjury. Functional and neurocognitive outcomes were measured and correlated with imaging data. Inflammatory cytokine arrays were measured in the plasma pretreatment, posttreatment, and at 1 and 6 month follow-up. There were no serious adverse events. There was a mild pulmonary toxicity of the highest dose that was not clinically significant. Despite the treatment group having greater injury severity, there was structural preservation of critical regions of interest

  4. Distribution of Brain Metastases in Relation to the Hippocampus: Implications for Neurocognitive Functional Preservation

    SciTech Connect

    Ghia, Amol; Tome, Wolfgang A.; Thomas, Sayana; Cannon, George; Khuntia, Deepak; Kuo, John S.; Mehta, Minesh P. . E-mail: mehta@humonc.wisc.edu

    2007-07-15

    Purpose: With the advent of intensity-modulated radiotherapy, the ability to limit the radiation dose to normal tissue offers an avenue to limit side effects. This study attempted to delineate the distribution of brain metastases with relation to the hippocampus for the purpose of exploring the viability of tomotherapy-guided hippocampal sparing therapy potentially to reduce neurocognitive deficits from radiation. Methods and Materials: The pre-radiotherapy T1-weighted, postcontrast axial MR images of 100 patients who received whole brain radiotherapy, stereotactic radiosurgery, or a radiosurgical boost following whole brain radiotherapy between 2002 and 2006 were examined. We contoured brain metastases as well as hippocampi with 5-, 10-, and 15-mm expansion envelopes. Results: Of the 272 identified metastases, 3.3% (n = 9) were within 5 mm of the hippocampus, and 86.4% of metastases were greater than 15 mm from the hippocampus (n = 235). The most common location for metastatic disease was the frontal lobe (31.6%, n = 86). This was followed by the cerebellum (24.3%, n = 66), parietal lobe (16.9%, n = 46), temporal lobe (12.9%, n = 35), occipital lobe (7.7%, n = 21), deep brain nuclei (4.0%, n = 11), and brainstem (2.6%, n = 7). Conclusions: Of the 100 patients, 8 had metastases within 5 mm of the hippocampus. Hence, a 5-mm margin around the hippocampus for conformal avoidance whole brain radiotherapy represents an acceptable risk, especially because these patients in the absence of any other intracranial disease could be salvaged using stereotactic radiosurgery. Moreover, we developed a hippocampal sparing tomotherapy plan as proof of principle to verify the feasibility of this therapy in the setting of brain metastases.

  5. Language in the aging brain: the network dynamics of cognitive decline and preservation.

    PubMed

    Shafto, Meredith A; Tyler, Lorraine K

    2014-10-31

    Language is a crucial and complex lifelong faculty, underpinned by dynamic interactions within and between specialized brain networks. Whereas normal aging impairs specific aspects of language production, most core language processes are robust to brain aging. We review recent behavioral and neuroimaging evidence showing that language systems remain largely stable across the life span and that both younger and older adults depend on dynamic neural responses to linguistic demands. Although some aspects of network dynamics change with age, there is no consistent evidence that core language processes are underpinned by different neural networks in younger and older adults.

  6. Acid fibroblast growth factor preserves blood-brain barrier integrity by activating the PI3K-Akt-Rac1 pathway and inhibiting RhoA following traumatic brain injury

    PubMed Central

    Wu, Fenzan; Chen, Zaifeng; Tang, Chonghui; Zhang, Jinjing; Cheng, Li; Zuo, Hongxia; Zhang, Hongyu; Chen, Daqing; Xiang, Liping; Xiao, Jian; Li, Xiaokun; Xu, Xinlong; Wei, Xiaojie

    2017-01-01

    The blood-brain barrier (BBB) plays important roles in the recovery of traumatic brain injury (TBI) which is a major factor contributing to cerebral edema. Acid fibroblast growth factor (aFGF) contributes to maintain vascular integrity and restores nerve function. However, whether aFGF protects BBB following TBI remains unknown. The purpose of this study was to determine whether exogenous aFGF preserves BBB integrity by activating the PI3K-Akt-Rac1 pathway and inhibiting RhoA after TBI. BBB permeability was assessed using evans blue dye and fluorescein isothiocyanate dextran fluorescence. Neurofunctional tests, such as the garcia test, were conducted in a blinded fashion, and protein expression was evaluated via western blotting and immunofluorescence staining. Our results showed that aFGF improved neurofunctional deficits, preserved BBB integrity, and up-regulated tight junction proteins and adherens junction proteins 24 h after experimental TBI. However, the PI3K/Akt inhibitor LY294002 reversed the protective effects of aFGF on neurofunctional deficits and junction protein expression and significantly suppressed p-Akt and GTP-Rac1 activity. Furthermore, aFGF administration significantly decreased GTP-RhoA expression in the treated group compared with the vehicle group, while PI3K/Akt inhibition increased GTP-RhoA expression. Similar results were observed in vitro, as aFGF exerted protective effects on endothelial cell integrity by up-regulating junction proteins and PI3K-Akt-Rac1 pathway and down-regulating RhoA expression under oxygen-glucose deprivation/reoxygenation (OGD) conditions. These data suggest that exogenous aFGF reduces RhoA activity in part by activating the PI3K-Akt-Rac1 signaling pathway, thus improving neurofunctional deficits and preserving BBB integrity after TBI. PMID:28386321

  7. A calorie-restricted diet decreases brain iron accumulation and preserves motor performance in old rhesus monkeys

    PubMed Central

    Kastman, EK; Willette, AA; Coe, CL; Bendlin, BB; Kosmatka, KJ; McLaren, DG; Xu, G; Canu, E; Field, AS; Alexander, AL; Voytko, ML; Beasley, TM; Colman, RJ; Weindruch, R; Johnson, SC

    2010-01-01

    Caloric restriction (CR) reduces the pathological effects of aging and extends the lifespan in many species, including nonhuman primates, although the effect on the brain is less well characterized. We used two common indicators of aging, motor performance speed and brain iron deposition measured in vivo using MRI, to determine the potential effect of CR on elderly rhesus macaques eating restricted (n = 24, 13 males, 11 females) and standard diets (n= 17, 8 males, 9 females). Both the CR and control monkeys showed age-related increases in iron concentrations in globus pallidus (GP) and substantia nigra (SN), although the CR group had significantly less iron deposition in the GP, SN, red nucleus and temporal cortex. A Diet x Age interaction revealed that CR modified age-related brain changes, evidenced as attenuation in the rate of iron accumulation in basal ganglia and parietal, temporal, and perirhinal cortex. Additionally, control monkeys had significantly slower fine motor performance on the Movement Assessment Panel, which was negatively correlated with iron accumulation in left SN and parietal lobe, although CR animals did not show this relationship. Our observations suggest that the CR induced benefit of reduced iron deposition and preserved motor function may indicate neural protection similar to effects described previously in aging rodent and primate species. PMID:20534842

  8. Imaging studies in congenital anophthalmia reveal preservation of brain architecture in 'visual' cortex.

    PubMed

    Bridge, Holly; Cowey, Alan; Ragge, Nicola; Watkins, Kate

    2009-12-01

    The functional specialization of the human brain means that many regions are dedicated to processing a single sensory modality. When a modality is absent, as in congenital total blindness, 'visual' regions can be reliably activated by non-visual stimuli. The connections underlying this functional adaptation, however, remain elusive. In this study, using structural and diffusion-weighted magnetic resonance imaging, we investigated the structural differences in the brains of six bilaterally anophthalmic subjects compared with sighted subjects. Surprisingly, the gross structural differences in the brains were small, even in the occipital lobe where only a small region of the primary visual cortex showed a bilateral reduction in grey matter volume in the anophthalmic subjects compared with controls. Regions of increased cortical thickness were apparent on the banks of the Calcarine sulcus, but not in the fundus. Subcortically, the white matter volume around the optic tract and internal capsule in anophthalmic subjects showed a large decrease, yet the optic radiation volume did not differ significantly. However, the white matter integrity, as measured with fractional anisotropy showed an extensive reduction throughout the brain in the anophthalmic subjects, with the greatest difference in the optic radiations. In apparent contradiction to the latter finding, the connectivity between the lateral geniculate nucleus and primary visual cortex measured with diffusion tractography did not differ between the two populations. However, these findings can be reconciled by a demonstration that at least some of the reduction in fractional anisotropy in the optic radiation is due to an increase in the strength of fibres crossing the radiations. In summary, the major changes in the 'visual' brain in anophthalmic subjects may be subcortical, although the evidence of decreased fractional anisotropy and increased crossing fibres could indicate considerable re-organization.

  9. Suppressor cell function is preserved in pemphigus and pemphigoid

    SciTech Connect

    King, A.J.; Schwartz, S.A.; Lopatin, D.; Voorhees, J.J.; Diaz, L.A.

    1982-09-01

    Human peripheral blood lymphocytes (PBL) are activated to become suppressor T cells (S-T-C) by incubation with Concanavalin-A (Con-A). This has become the standard method for evaluation of suppressor function in patients. S-T-C function has been found to be impaired in several autoimmune diseases, including systemic lupus erythematosus (SLE). Using this assay, we have investigated suppressor-cell function in 2 autoimmune disorders, bullous pemphigoid (BP) and pemphigus vulgaris (PV), studying 6 patients from each group. Three patients with active SLE (positive controls), and 11 normal donors (negative controls) were also included. None of these patients had received systemic therapy with the exception of 2 patients with PV who were treated with gold in the past. PBL from these patients were incubated with and without 40 micrograms/ml Con-A for 72 hr to generate suppressor cells. Both groups of PBL were then irradiated wih 1500 r cobalt. Co-cultures were set up in sextuplicate using normal PBL as responders. Responder PBL were stimulated with 0.5, 1.0, and 2.0 micrograms/ml of phytohemagglutin (PHA) and 5.0, 10.0, and 20.0 micrograms/ml of Con-A. Cultures were pulsed on day 3 with /sup 3/H-thymidine and harvested on day 4. Data were analyzed using Student's t-test. S-T-C function was found to be significantly impaired in SLE vs normal (p . 0.0316). No statistically significant difference was seen in BP (p . 0.5883) and PV (p . 0.0921) as compared with normals. A defect in suppressor cell function may still be present in patients with PV and BP for the defect may be antigen-specific and therefore remain undetected by the Con-A suppressor assay.

  10. New Developments in Red Blood Cell Preservation Using Liquid and Freezing Procedures.

    DTIC Science & Technology

    1982-04-02

    phosphate ion exchange resin.30 ൪ Solutions of bicarbonate, adenine, glucose and mannitol (BAGM), saline, adenine and RED BLOOD CELL PRESERVATION 10...glucose (SAO), and adenine, glucose, sodium chloride and mannitol (ADSOL), have been used to maintain or increase the red cell organic phosphate...compounds, ATP which influences posttransfusion survival, and 2,3 DPG which influences red cell oxygen transport function. Studies have shown that mannitol

  11. Reorganization of syntactic processing following left-hemisphere brain damage: does right-hemisphere activity preserve function?

    PubMed

    Tyler, Lorraine K; Wright, Paul; Randall, Billi; Marslen-Wilson, William D; Stamatakis, Emmanuel A

    2010-11-01

    The extent to which the human brain shows evidence of functional plasticity across the lifespan has been addressed in the context of pathological brain changes and, more recently, of the changes that take place during healthy ageing. Here we examine the potential for plasticity by asking whether a strongly left-lateralized system can successfully reorganize to the right-hemisphere following left-hemisphere brain damage. To do this, we focus on syntax, a key linguistic function considered to be strongly left-lateralized, combining measures of tissue integrity, neural activation and behavioural performance. In a functional neuroimaging study participants heard spoken sentences that differentially loaded on syntactic and semantic information. While healthy controls activated a left-hemisphere network of correlated activity including Brodmann areas 45/47 and posterior middle temporal gyrus during syntactic processing, patients activated Brodmann areas 45/47 bilaterally and right middle temporal gyrus. However, voxel-based morphometry analyses showed that only tissue integrity in left Brodmann areas 45/47 was correlated with activity and performance; poor tissue integrity in left Brodmann area 45 was associated with reduced functional activity and increased syntactic deficits. Activity in the right-hemisphere was not correlated with damage in the left-hemisphere or with performance. Reduced neural integrity in the left-hemisphere through brain damage or healthy ageing results in increased right-hemisphere activation in homologous regions to those left-hemisphere regions typically involved in the young. However, these regions do not support the same linguistic functions as those in the left-hemisphere and only indirectly contribute to preserved syntactic capacity. This establishes the unique role of the left hemisphere in syntax, a core component in human language.

  12. Hypothermia and amiloride preserve energetics in a neonatal brain slice model.

    PubMed

    Robertson, Nicola J; Bhakoo, Kishore; Puri, Basant K; Edwards, A David; Cox, I Jane

    2005-08-01

    A period of secondary energy failure consisting of a decline in phosphocreatine/inorganic phosphate (PCr/Pi), a rise in brain lactate, and alkaline intracellular pH (pH(i)) has been described in infants with neonatal encephalopathy. Strategies that ameliorate this energy failure may be neuroprotective. We hypothesized that a neonatal rat brain slice model undergoes a progressive decline in energetics, which can be ameliorated with hypothermia or amiloride. Interleaved phosphorus ((31)P) and proton ((1)H) magnetic resonance (MR) spectra were obtained from 350 microm neonatal rat brain slices over 8 h in a bicarbonate buffer at 37 degrees C and at 32 degrees C in 7- and 14-d models. (31)P MR spectra were obtained with amiloride in a bicarbonate-free buffer at 37 degrees C in the 14-d model. Findings were similar in 7- and 14-d models. In the 14-d model, there was a Pi doublet structure corresponding to alkaline pH(i) values of 7.50 +/- 0.02 and 7.21 +/- 0.04. Compared with the stabilized baseline of 100, at 5 h PCr/Pi was 65 +/- 6.3 and lactate/NAA was 187 +/- 3 at 37 degrees C, but PCr/Pi and lactate/NAA were not significantly different from baseline at 32 degrees C. Nucleotide triphosphate (NTP)/phosphomonoester (PME) was 0.93 +/- 0.23 at 37 degrees C and 1.81 +/- 0.21 at 32 degrees C at 5 h. With amiloride exposure in the 14-d model, baseline pH(i) values were 7.25 +/- 0.09 and 6.98 +/- 0.02 and NTP/PME was 1.81 +/- 0.05; these parameters were not significantly different at 5 h. Our interpretation of these findings is that the brain slice model underwent secondary energy failure, which was delayed with hypothermia or amiloride.

  13. A new system for cutting brain tissue preserving vessels: water jet cutting.

    PubMed

    Terzis, A J; Nowak, G; Rentzsch, O; Arnold, H; Diebold, J; Baretton, G

    1989-01-01

    The water jet cutting system allows transaction and dissection of biological structures with little bleeding. Structures of higher tissue rigidity remain unchanged while softer tissues are mechanically dissected. In brain tissue, all vessels larger than 20 microns are left intact after the passage of the jet stream with a pressure of up to 5 bar, and therefore vessels can be isolated selectively from the surrounding tissue. Oedema is present adjacent to the cut and no increase of temperature occurs.

  14. Association of fish oil supplement use with preservation of brain volume and cognitive function

    PubMed Central

    Daiello, Lori A.; Gongvatana, Assawin; Dunsiger, Shira; Cohen, Ronald A.; Ott, Brian R.

    2016-01-01

    Objective The aim of this study was to investigate whether the use of fish oil supplements (FOSs) is associated with concomitant reduction in cognitive decline and brain atrophy in older adults. Methods We conducted a retrospective cohort study to examine the relationship between FOS use during the Alzheimer’s Disease Neuroimaging Initiative and indicators of cognitive decline. Older adults (229 cognitively normal individuals, 397 patients with mild cognitive impairment, and 193 patients with Alzheimer’s disease) were assessed with neuropsychological tests and brain magnetic resonance imaging every 6 months. Primary outcomes included (1) global cognitive status and (2) cerebral cortex gray matter and hippocampus and ventricular volumes. Results FOS use during follow-up was associated with significantly lower mean cognitive subscale of the Alzheimer’s Disease Assessment Scale and higher Mini-Mental State Examination scores among those with normal cognition. Associations between FOS use and the outcomes were observed only in APOE ε4–negative participants. FOS use during the study was also associated with less atrophy in one or more brain regions of interest. PMID:24954371

  15. Breast Cancer Cells May Change When They Spread to Brain

    MedlinePlus

    ... Cancer Cells May Change When They Spread to Brain: Study Finding might lead to better treatment, researchers ... HealthDay News) -- When breast cancer spreads to the brain, important molecular changes may occur in the cancer, ...

  16. Heat shock treatment protects osmotic stress–induced dysfunction of the blood-brain barrier through preservation of tight junction proteins

    PubMed Central

    Lu, Tzong-Shi; Chen, Hsiang-Wen; Huang, Maw-Hsiung; Wang, Shu-Jung; Yang, Rei-Cheng

    2004-01-01

    The blood-brain barrier (BBB) is a specialized structure in the central nervous system (CNS), which participates in maintenance of a state of cerebrospinal fluid homeostasis. The endothelial cells of the cerebral capillaries and the tight junctions between them form the basis of the BBB. Research has shown that destruction of the BBB is associated with diseases of the CNS. However, there is little research on how the BBB might be protected. In this study, we used a high osmotic solution (1.6 M d-mannitol) to open the BBB of rats and Evans blue dye as a macromolecular marker. The effect of heat shock treatment was evaluated. The results show that increased synthesis of heat shock protein 72 (Hsp72) was induced in the heated group only. BBB permeability was significantly less in the heat shock–treated group after hyperosmotic shock. The major tight junction proteins, occludin and zonula occludens (ZO)-1, were significantly decreased after d-mannitol treatment in the nonheated group, whereas they were preserved in the heated group. The coimmunoprecipitation studies demonstrated that Hsp72 could be detected in the precipitates of brain extract interacting with anti–ZO-1 antibodies as well as those interacting with anti–occludin antibodies in the heated group. We conclude that the integrity of tight junctions could be maintained by previous heat shock treatment, which might be associated with the increased production of Hsp72. PMID:15633295

  17. Brain repair: cell therapy in stroke

    PubMed Central

    Kalladka, Dheeraj; Muir, Keith W

    2014-01-01

    Stroke affects one in every six people worldwide, and is the leading cause of adult disability. Some spontaneous recovery is usual but of limited extent, and the mechanisms of late recovery are not completely understood. Endogenous neurogenesis in humans is thought to contribute to repair, but its extent is unknown. Exogenous cell therapy is promising as a means of augmenting brain repair, with evidence in animal stroke models of cell migration, survival, and differentiation, enhanced endogenous angiogenesis and neurogenesis, immunomodulation, and the secretion of trophic factors by stem cells from a variety of sources, but the potential mechanisms of action are incompletely understood. In the animal models of stroke, both mesenchymal stem cells (MSCs) and neural stem cells (NSCs) improve functional recovery, and MSCs reduce the infarct volume when administered acutely, but the heterogeneity in the choice of assessment scales, publication bias, and the possible confounding effects of immunosuppressants make the comparison of effects across cell types difficult. The use of adult-derived cells avoids the ethical issues around embryonic cells but may have more restricted differentiation potential. The use of autologous cells avoids rejection risk, but the sources are restricted, and culture expansion may be necessary, delaying treatment. Allogeneic cells offer controlled cell numbers and immediate availability, which may have advantages for acute treatment. Early clinical trials of both NSCs and MSCs are ongoing, and clinical safety data are emerging from limited numbers of selected patients. Ongoing research to identify prognostic imaging markers may help to improve patient selection, and the novel imaging techniques may identify biomarkers of recovery and the mechanism of action for cell therapies. PMID:24627643

  18. Human brain glial cells synthesize thrombospondin.

    PubMed Central

    Asch, A S; Leung, L L; Shapiro, J; Nachman, R L

    1986-01-01

    Thrombospondin, a 450-kDa multinodular glycoprotein with lectin-type activity, is found in human platelets, endothelial cells, fibroblasts, smooth muscle cells, monocytes, and granular pneumocytes. Thrombospondin interacts with heparin, fibrinogen, fibronectin, collagen, histidine-rich glycoprotein, and plasminogen. Recently, thrombospondin synthesis by smooth muscle cells has been reported to be augmented by platelet-derived growth factor. We present evidence that thrombospondin is present within and synthesized by astrocytic neuroglial cells. Heparin-Sepharose affinity chromatography of material derived from a human brain homogenate yielded a protein that, when reduced, had an apparent size of 180 kDa and comigrated with reduced platelet thrombospondin on NaDodSO4/PAGE. Immunoblot analysis with monospecific anti-thrombospondin confirmed the presence of immunoreactive thrombospondin. Indirect immunofluorescence of cultured human glial cells indicated the presence of thrombospondin. Metabolic labeling of glial cell cultures with [35S]methionine followed by immunoprecipitation with monospecific anti-thrombospondin revealed synthesis of a 180-kDa polypeptide that comigrated with platelet thrombospondin on NaDodSO4/PAGE. Cultured human glial cells were incubated for 48 hr in serum-free medium with purified platelet-derived growth factor at concentrations up to 50 ng/ml. Aliquots taken at intervals were analyzed by a quantitative double-antibody ELISA. The growth factor stimulated the release of thrombospondin into the culture medium by as much as 10-fold over control cultures. The presence of thrombospondin within glial cells of the central nervous system and the augmentation of its synthesis by platelet-derived growth factor suggest that thrombospondin may play an important role in regulating cell-cell and cell-matrix interactions during periods of cell division and growth. Images PMID:2939460

  19. Immune interventions to preserve β cell function in type 1 diabetes.

    PubMed

    Ehlers, Mario R

    2016-01-01

    Type 1 diabetes (T1D) is a chronic autoimmune disease that leads to destruction of pancreatic β cells, lifelong dependence on insulin, and increased morbidity and mortality from diabetes-related complications. Preservation of residual β cells at diagnosis is a major goal because higher levels of endogenous insulin secretion are associated with better short- and long-term outcomes. For the past 3 decades, a variety of immune interventions have been evaluated in the setting of new-onset T1D, including nonspecific immunosuppression, pathway-specific immune modulation, antigen-specific therapies, and cellular therapies. To date, no single intervention has produced durable remission off therapy in most treated patients, but the field has gained valuable insights into disease mechanisms and potential immunologic correlates of success. In particular, T-cell–directed therapies, including therapies that lead to partial depletion or modulation of effector T cells and preservation or augmentation of regulatory T cells, have shown the most success and will likely form the backbone of future approaches. The next phase will see evaluation of rational combinations, comprising one or more of the following: an effector T-depleting or -modulating drug, a cytokine-based tolerogenic (regulatory T-cells–promoting) agent, and an antigen-specific component. The long term goal is to reestablish immunologic tolerance to β cells, thereby preserving residual β cells early after diagnosis or enabling restoration of β-cell mass from autologous stem cells or induced neogenesis in patients with established T1D.

  20. Stem Cell-Mediated Regeneration of the Adult Brain

    PubMed Central

    Jessberger, Sebastian

    2016-01-01

    Acute or chronic injury of the adult mammalian brain is often associated with persistent functional deficits as its potential for regeneration and capacity to rebuild lost neural structures is limited. However, the discovery that neural stem cells (NSCs) persist throughout life in discrete regions of the brain, novel approaches to induce the formation of neuronal and glial cells, and recently developed strategies to generate tissue for exogenous cell replacement strategies opened novel perspectives how to regenerate the adult brain. Here, we will review recently developed approaches for brain repair and discuss future perspectives that may eventually allow for developing novel treatment strategies in acute and chronic brain injury. PMID:27781019

  1. Communication of brain network core connections altered in behavioral variant frontotemporal dementia but possibly preserved in early-onset Alzheimer's disease

    NASA Astrophysics Data System (ADS)

    Daianu, Madelaine; Jahanshad, Neda; Mendez, Mario F.; Bartzokis, George; Jimenez, Elvira E.; Thompson, Paul M.

    2015-03-01

    Diffusion imaging and brain connectivity analyses can assess white matter deterioration in the brain, revealing the underlying patterns of how brain structure declines. Fiber tractography methods can infer neural pathways and connectivity patterns, yielding sensitive mathematical metrics of network integrity. Here, we analyzed 1.5-Tesla wholebrain diffusion-weighted images from 64 participants - 15 patients with behavioral variant frontotemporal dementia (bvFTD), 19 with early-onset Alzheimer's disease (EOAD), and 30 healthy elderly controls. Using whole-brain tractography, we reconstructed structural brain connectivity networks to map connections between cortical regions. We evaluated the brain's networks focusing on the most highly central and connected regions, also known as hubs, in each diagnostic group - specifically the "high-cost" structural backbone used in global and regional communication. The high-cost backbone of the brain, predicted by fiber density and minimally short pathways between brain regions, accounted for 81-92% of the overall brain communication metric in all diagnostic groups. Furthermore, we found that the set of pathways interconnecting high-cost and high-capacity regions of the brain's communication network are globally and regionally altered in bvFTD, compared to healthy participants; however, the overall organization of the high-cost and high-capacity networks were relatively preserved in EOAD participants, relative to controls. Disruption of the major central hubs that transfer information between brain regions may impair neural communication and functional integrity in characteristic ways typical of each subtype of dementia.

  2. Preserving cortico-striatal function: deep brain stimulation in Huntington's disease.

    PubMed

    Nagel, Sean J; Machado, Andre G; Gale, John T; Lobel, Darlene A; Pandya, Mayur

    2015-01-01

    Huntington's disease (HD) is an incurable neurodegenerative disease characterized by the triad of chorea, cognitive dysfunction and psychiatric disturbances. Since the discovery of the HD gene, the pathogenesis has been outlined, but to date a cure has not been found. Disease modifying therapies are needed desperately to improve function, alleviate suffering, and provide hope for symptomatic patients. Deep brain stimulation (DBS), a proven therapy for managing the symptoms of some neurodegenerative movement disorders, including Parkinson's disease, has been reported as a palliative treatment in select cases of HD with debilitating chorea with variable success. New insights into the mechanism of action of DBS suggest it may have the potential to circumvent other manifestations of HD including cognitive deterioration. Furthermore, because DBS is already widely used, reversible, and has a risk profile that is relatively low, new studies can be initiated. In this article we contend that new clinical trials be considered to test the effects of DBS for HD.

  3. Intranasal Delivery of A Novel Amnion Cell Secretome Prevents Neuronal Damage and Preserves Function In A Mouse Multiple Sclerosis Model

    PubMed Central

    Khan, Reas S.; Dine, Kimberly; Bauman, Bailey; Lorentsen, Michael; Lin, Lisa; Brown, Helayna; Hanson, Leah R.; Svitak, Aleta L.; Wessel, Howard; Brown, Larry; Shindler, Kenneth S.

    2017-01-01

    The ability of a novel intranasally delivered amnion cell derived biologic to suppress inflammation, prevent neuronal damage and preserve neurologic function in the experimental autoimmune encephalomyelitis animal model of multiple sclerosis was assessed. Currently, there are no existing optic nerve treatment methods for disease or trauma that result in permanent vision loss. Demyelinating optic nerve inflammation, termed optic neuritis, induces permanent visual dysfunction due to retinal ganglion cell damage in multiple sclerosis and experimental autoimmune encephalomyelitis. ST266, the biological secretome of Amnion-derived Multipotent Progenitor cells, contains multiple anti-inflammatory cytokines and growth factors. Intranasally administered ST266 accumulated in rodent eyes and optic nerves, attenuated visual dysfunction, and prevented retinal ganglion cell loss in experimental optic neuritis, with reduced inflammation and demyelination. Additionally, ST266 reduced retinal ganglion cell death in vitro. Neuroprotective effects involved oxidative stress reduction, SIRT1-mediated mitochondrial function promotion, and pAKT signaling. Intranasal delivery of neuroprotective ST266 is a potential novel, noninvasive therapeutic modality for the eyes, optic nerves and brain. The unique combination of biologic molecules in ST266 provides an innovative approach with broad implications for suppressing inflammation in autoimmune diseases, and for preventing neuronal damage in acute neuronal injury and chronic neurodegenerative diseases such as multiple sclerosis. PMID:28139754

  4. Adult human brain cell culture for neuroscience research.

    PubMed

    Gibbons, Hannah M; Dragunow, Mike

    2010-06-01

    Studies of the brain have progressed enormously through the use of in vivo and in vitro non-human models. However, it is unlikely such studies alone will unravel the complexities of the human brain and so far no neuroprotective treatment developed in animals has worked in humans. In this review we discuss the use of adult human brain cell culture methods in brain research to unravel the biology of the normal and diseased human brain. The advantages of using adult human brain cells as tools to study human brain function from both historical and future perspectives are discussed. In particular, studies using dissociated cultures of adult human microglia, astrocytes, oligodendrocytes and neurons are described and the applications of these types of study are evaluated. Alternative sources of human brain cells such as adult neural stem cells, induced pluripotent stem cells and slice cultures of adult human brain tissue are also reviewed. These adult human brain cell culture methods could benefit basic research and more importantly, facilitate the translation of basic neuroscience research to the clinic for the treatment of brain disorders.

  5. A survey of human brain transcriptome diversity at the single cell level.

    PubMed

    Darmanis, Spyros; Sloan, Steven A; Zhang, Ye; Enge, Martin; Caneda, Christine; Shuer, Lawrence M; Hayden Gephart, Melanie G; Barres, Ben A; Quake, Stephen R

    2015-06-09

    The human brain is a tissue of vast complexity in terms of the cell types it comprises. Conventional approaches to classifying cell types in the human brain at single cell resolution have been limited to exploring relatively few markers and therefore have provided a limited molecular characterization of any given cell type. We used single cell RNA sequencing on 466 cells to capture the cellular complexity of the adult and fetal human brain at a whole transcriptome level. Healthy adult temporal lobe tissue was obtained during surgical procedures where otherwise normal tissue was removed to gain access to deeper hippocampal pathology in patients with medical refractory seizures. We were able to classify individual cells into all of the major neuronal, glial, and vascular cell types in the brain. We were able to divide neurons into individual communities and show that these communities preserve the categorization of interneuron subtypes that is typically observed with the use of classic interneuron markers. We then used single cell RNA sequencing on fetal human cortical neurons to identify genes that are differentially expressed between fetal and adult neurons and those genes that display an expression gradient that reflects the transition between replicating and quiescent fetal neuronal populations. Finally, we observed the expression of major histocompatibility complex type I genes in a subset of adult neurons, but not fetal neurons. The work presented here demonstrates the applicability of single cell RNA sequencing on the study of the adult human brain and constitutes a first step toward a comprehensive cellular atlas of the human brain.

  6. Brain mesenchymal stem cells: The other stem cells of the brain?

    PubMed

    Appaix, Florence; Nissou, Marie-France; van der Sanden, Boudewijn; Dreyfus, Matthieu; Berger, François; Issartel, Jean-Paul; Wion, Didier

    2014-04-26

    Multipotent mesenchymal stromal cells (MSC), have the potential to differentiate into cells of the mesenchymal lineage and have non-progenitor functions including immunomodulation. The demonstration that MSCs are perivascular cells found in almost all adult tissues raises fascinating perspectives on their role in tissue maintenance and repair. However, some controversies about the physiological role of the perivascular MSCs residing outside the bone marrow and on their therapeutic potential in regenerative medicine exist. In brain, perivascular MSCs like pericytes and adventitial cells, could constitute another stem cell population distinct to the neural stem cell pool. The demonstration of the neuronal potential of MSCs requires stringent criteria including morphological changes, the demonstration of neural biomarkers expression, electrophysiological recordings, and the absence of cell fusion. The recent finding that brain cancer stem cells can transdifferentiate into pericytes is another facet of the plasticity of these cells. It suggests that the perversion of the stem cell potential of pericytes might play an even unsuspected role in cancer formation and tumor progression.

  7. Immune Interventions to Preserve Beta Cell Function in Type 1 Diabetes

    PubMed Central

    Ehlers, Mario R.

    2015-01-01

    Type 1 diabetes (T1D) is a chronic autoimmune disease that leads to destruction of pancreatic beta cells, lifelong dependence on insulin, and increased morbidity and mortality from diabetes-related complications. Preservation of residual beta cells at diagnosis is a major goal because higher levels of endogenous insulin secretion are associated with better short- and long-term outcomes. Over the past 3 decades, a variety of immune interventions have been evaluated in the setting of new-onset T1D, including nonspecific immunosuppression, pathway-specific immune modulation, antigen-specific therapies, and cellular therapies. To date, no single intervention has produced durable remission off-therapy in the majority of treated patients, but the field has gained valuable insights into disease mechanisms and potential immunologic correlates of success. In particular, T cell-directed therapies, including therapies that lead to partial depletion or modulation of effector T (Teff) cells and preservation or augmentation of regulatory T (Treg) cells, have shown the most success and will likely form the backbone of future approaches. The next phase will see evaluation of rational combinations, comprising one or more of the following: a Teff-depleting or modulating drug, a cytokine-based tolerogenic (Treg-promoting) agent, and an antigen-specific component. The long-term goal is to reestablish immunologic tolerance to beta cells, thereby preserving residual beta cells early after diagnosis or enabling restoration of beta cell mass from autologous stem cells or induced neogenesis in patients with established T1D. PMID:26225763

  8. Stem cells in cardiovascular regeneration: from preservation of endogenous repair to future cardiovascular therapies.

    PubMed

    Templin, Christian; Kränkel, Nicolle; Lüscher, Thomas F; Landmesser, Ulf

    2011-10-01

    Cardiovascular disease remains the leading cause of morbidity and mortality in the developed countries. This review summarizes current pre-clinical and clinical evidence for the potential role and mechanisms of action of stem and progenitor cells in vascular and cardiac repair and regeneration. Apart from cell transplantation strategies, approaches to maintain stem cell niche function and targeting mobilization/recruitment of specific stem/progenitor cell populations may aid in preserving vascular and cardiac function. Moreover, with the use of patient-derived induced pluripotent stem cells, the field of regenerative medicine is entering a new era. Potential applications of induced pluripotent stem cells and direct reprogrammed cells as well as recent developments in tissue engineering are discussed.

  9. Preservation of Ocular Epithelial Limbal Stem Cells: The New Frontier in Regenerative Medicine.

    PubMed

    Lužnik, Zala; Bertolin, Marina; Breda, Claudia; Ferrari, Barbara; Barbaro, Vanessa; Schollmayer, Petra; Ferrari, Stefano

    2016-01-01

    Significant advances have been made in the field of ocular regenerative medicine. Promising stem cell-based therapeutic strategies have been translated into the clinical practice over the last few decades. These new stem cell-based therapies offer the possibility of permanently restoring corneal epithelium in patients with severe disabling and blinding ocular surface disease. The European Union has already classified stem cell-based therapies as "medicinal products". Therefore, manipulation is strictly regulated according to the defined conditions of good manufacturing practice, with the production of stem cell therapeutics at only accredited production sites authorized by the national regulatory agencies. In this regard, as first medical products are licensed for commercial use in Europe enabling a more widespread access to a stem cell-based therapy, the need for safe, validated and reproducible techniques for ex vivo cultured tissue preservation and distribution are coming to the forefront of research. However, these provide various new challenges for biobanking industry such as the retention of viability, good functionality of stem cells and sterility issues. This chapter provides an overview of the current advances in the field of corneal/limbal epithelial stem cell culture preservation techniques using either hypothermic storage or cryopreservation methods, that were used in different culturing steps (from stem cell isolation to the ex vivo epithelial graft preparation), with the reported impact on the post-thawing product recovery.

  10. Donor satellite cell engraftment is significantly augmented when the host niche is preserved and endogenous satellite cells are incapacitated.

    PubMed

    Boldrin, Luisa; Neal, Alice; Zammit, Peter S; Muntoni, Francesco; Morgan, Jennifer E

    2012-09-01

    Stem cell transplantation is already in clinical practice for certain genetic diseases and is a promising therapy for dystrophic muscle. We used the mdx mouse model of Duchenne muscular dystrophy to investigate the effect of the host satellite cell niche on the contribution of donor muscle stem cells (satellite cells) to muscle regeneration. We found that incapacitation of the host satellite cells and preservation of the muscle niche promote donor satellite cell contribution to muscle regeneration and functional reconstitution of the satellite cell compartment. But, if the host niche is not promptly refilled, or is filled by competent host satellite cells, it becomes nonfunctional and donor engraftment is negligible. Application of this regimen to aged host muscles also promotes efficient regeneration from aged donor satellite cells. In contrast, if the niche is destroyed, yet host satellite cells remain proliferation-competent, donor-derived engraftment is trivial. Thus preservation of the satellite cell niche, concomitant with functional impairment of the majority of satellite cells within dystrophic human muscles, may improve the efficiency of stem cell therapy.

  11. Interleaved programming of subthalamic deep brain stimulation to avoid adverse effects and preserve motor benefit in Parkinson's disease.

    PubMed

    Ramirez-Zamora, Adolfo; Kahn, Max; Campbell, Joannalee; DeLaCruz, Priscilla; Pilitsis, Julie G

    2015-03-01

    Subthalamic nucleus (STN) deep brain stimulation (DBS) is the most common surgical treatment for managing motor complications in Parkinson's disease (PD). Ultimately, outcomes depend on a variety of factors including lead location, access and expertize in programming and PD medical management. Nevertheless, achieving ideal programming settings can be difficult in certain patients, leading to suboptimal control of symptoms and stimulation-induced side effects, notably dysarthria and dyskinesia. Interleaved stimulation (ILS) is a newer programming technique that attempts to optimize the stimulation field, improving control of symptoms while minimizing stimulation-induced adverse effects. A retrospective chart review was performed on PD patients receiving STN DBS over the past 12 months. Clinical and demographic data were collected from patients identified as having received ILS. The rationale and clinical efficacy of ILS was analyzed. Nine patients received ILS due to incomplete PD symptom control or stimulation-induced side effects after attempting multiple programming options. Appropriate lead location was confirmed with postoperative MRI except in one case. Following ILS, patients reported improvement in symptoms and resolution of side effects, while preserving adequate control in Parkinsonism with a mean improvement in UPDRS-MOTOR scores of 51.2 %. ILS continues to emerge as a safe and effective programming strategy for maximizing symptom control in PD while diminishing stimulation-induced side effects.

  12. In Vitro Effects of Preserved and Unpreserved Anti-Allergic Drugs on Human Corneal Epithelial Cells

    PubMed Central

    Calvo, Patricia; Ropero, Inés; Pintor, Jesús

    2014-01-01

    Abstract Purpose: Treatment with topical eye drops for long-standing ocular diseases like allergy can induce detrimental side effects. The purpose of this study was to investigate in vitro cytotoxicity of commercially preserved and unpreserved anti-allergic eye drops on the viability and barrier function of monolayer and stratified human corneal-limbal epithelial cells. Methods: Cells were treated with unpreserved ketotifen solution, benzalkonium chloride (BAC)-containing anti-allergic drugs (ketotifen, olopatadine, levocabastine) as well as BAC alone. 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used to determine cell viability. Effects of compounds on barrier function were analyzed measuring transepithelial electrical resistance (TEER) to determine paracellular permeability and rose bengal assays to evaluate transcellular barrier formation. Results: The BAC-preserved anti-allergic formulations and BAC alone significantly reduced cell viability, monolayer cultures being more sensitive to damage by these solutions. Unpreserved ketotifen induced the least diminution in cell viability. The extent of decrease of cell viability was clearly dependent of BAC presence, but it was also affected by the different types of drugs when the concentration of BAC was low and the short time of exposure. Treatment with BAC-containing anti-allergic drugs and BAC alone resulted in increased paracellular permeability and loss of transcellular barrier function as indicated by TEER measurement and rose bengal assays. Conclusions: The presence of the preservative BAC in anti-allergic eye drop formulations contributes importantly to the cytotoxic effects induced by these compounds. Stratified cell cultures seem to be a more relevant model for toxicity evaluation induced on the ocular surface epithelia than monolayer cultures. PMID:25100331

  13. The aged brain: genesis and fate of residual progenitor cells in the subventricular zone

    PubMed Central

    Capilla-Gonzalez, Vivian; Herranz-Pérez, Vicente; García-Verdugo, Jose Manuel

    2015-01-01

    Neural stem cells (NSCs) persist in the adult mammalian brain through life. The subventricular zone (SVZ) is the largest source of stem cells in the nervous system, and continuously generates new neuronal and glial cells involved in brain regeneration. During aging, the germinal potential of the SVZ suffers a widespread decline, but the causes of this turn down are not fully understood. This review provides a compilation of the current knowledge about the age-related changes in the NSC population, as well as the fate of the newly generated cells in the aged brain. It is known that the neurogenic capacity is clearly disrupted during aging, while the production of oligodendroglial cells is not compromised. Interestingly, the human brain seems to primarily preserve the ability to produce new oligodendrocytes instead of neurons, which could be related to the development of neurological disorders. Further studies in this matter are required to improve our understanding and the current strategies for fighting neurological diseases associated with senescence. PMID:26441536

  14. FACS purification of immunolabeled cell types from adult rat brain.

    PubMed

    Guez-Barber, Danielle; Fanous, Sanya; Harvey, Brandon K; Zhang, Yongqing; Lehrmann, Elin; Becker, Kevin G; Picciotto, Marina R; Hope, Bruce T

    2012-01-15

    Molecular analysis of brain tissue is greatly complicated by having many different classes of neurons and glia interspersed throughout the brain. Fluorescence-activated cell sorting (FACS) has been used to purify selected cell types from brain tissue. However, its use has been limited to brain tissue from embryos or transgenic mice with promoter-driven reporter genes. To overcome these limitations, we developed a FACS procedure for dissociating intact cell bodies from adult wild-type rat brains and sorting them using commercially available antibodies against intracellular and extracellular proteins. As an example, we isolated neurons using a NeuN antibody and confirmed their identity using microarray and real time PCR of mRNA from the sorted cells. Our FACS procedure allows rapid, high-throughput, quantitative assays of molecular alterations in identified cell types with widespread applications in neuroscience.

  15. An epigenetic gateway to brain tumor cell identity.

    PubMed

    Mack, Stephen C; Hubert, Christopher G; Miller, Tyler E; Taylor, Michael D; Rich, Jeremy N

    2016-01-01

    Precise targeting of genetic lesions alone has been insufficient to extend brain tumor patient survival. Brain cancer cells are diverse in their genetic, metabolic and microenvironmental compositions, accounting for their phenotypic heterogeneity and disparate responses to therapy. These factors converge at the level of the epigenome, representing a unified node that can be disrupted by pharmacologic inhibition. Aberrant epigenomes define many childhood and adult brain cancers, as demonstrated by widespread changes to DNA methylation patterns, redistribution of histone marks and disruption of chromatin structure. In this Review, we describe the convergence of genetic, metabolic and microenvironmental factors on mechanisms of epigenetic deregulation in brain cancer. We discuss how aberrant epigenetic pathways identified in brain tumors affect cell identity, cell state and neoplastic transformation, as well as addressing the potential to exploit these alterations as new therapeutic strategies for the treatment of brain cancer.

  16. Immunoelectron Microscopic Studies Indicate the Existence of a Cell Shape Preserving Cytoskeleton in Prokaryotes

    NASA Astrophysics Data System (ADS)

    Mayer, F.; Vogt, B.; Poc, C.

    1998-06-01

    , Ralstonia eutropha, Thermoanaerobacterium thermosulfurigenes, T. thermosaccharolyticum, and Methanococcus jannaschii. Substantial label also in the cytoplasm was observed in Bacillus sp., Methanococcus voltae, and Methanobacterium thermoautotrophicum. Only very minor amounts of label were found in the nucleoid region of the cells. Whole-mount immunogold studies, combined with negative staining, revealed the existence of an intracellular network of fibrils which could be labeled by anti-actin antibodies. This network is assumed to be located below the cytoplasmic membrane all around the cytoplasm. It appears to have properties that would allow its function as a cytoskeletonlike structure preserving cell shape.

  17. Nucleic Acid Encoding A Lectin-Derived Progenitor Cell Preservation Factor

    DOEpatents

    Colucci, M. Gabriella; Chrispeels, Maarten J.; Moore, Jeffrey G.

    2001-10-30

    The invention relates to an isolated nucleic acid molecule that encodes a protein that is effective to preserve progenitor cells, such as hematopoietic progenitor cells. The nucleic acid comprises a sequence defined by SEQ ID NO:1, a homolog thereof, or a fragment thereof. The encoded protein has an amino acid sequence that comprises a sequence defined by SEQ ID NO:2, a homolog thereof, or a fragment thereof that contains an amino acid sequence TNNVLQVT. Methods of using the encoded protein for preserving progenitor cells in vitro, ex vivo, and in vivo are also described. The invention, therefore, include methods such as myeloablation therapies for cancer treatment wherein myeloid reconstitution is facilitated by means of the specified protein. Other therapeutic utilities are also enabled through the invention, for example, expanding progenitor cell populations ex vivo to increase chances of engraftation, improving conditions for transporting and storing progenitor cells, and facilitating gene therapy to treat and cure a broad range of life-threatening hematologic diseases.

  18. Blood cell mRNAs and microRNAs: optimized protocols for extraction and preservation.

    PubMed

    Eikmans, Michael; Rekers, Niels V; Anholts, Jacqueline D H; Heidt, Sebastiaan; Claas, Frans H J

    2013-03-14

    Assessing messenger RNA (mRNA) and microRNA levels in peripheral blood cells may complement conventional parameters in clinical practice. Working with small, precious samples requires optimal RNA yields and minimal RNA degradation. Several procedures for RNA extraction and complementary DNA (cDNA) synthesis were compared for their efficiency. The effect on RNA quality of freeze-thawing peripheral blood cells and storage in preserving reagents was investigated. In terms of RNA yield and convenience, quality quantitative polymerase chain reaction signals per nanogram of total RNA and using NucleoSpin and mirVana columns is preferable. The SuperScript III protocol results in the highest cDNA yields. During conventional procedures of storing peripheral blood cells at -180°C and thawing them thereafter, RNA integrity is maintained. TRIzol preserves RNA in cells stored at -20°C. Detection of mRNA levels significantly decreases in degraded RNA samples, whereas microRNA molecules remain relatively stable. When standardized to reference targets, mRNA transcripts and microRNAs can be reliably quantified in moderately degraded (quality index 4-7) and severely degraded (quality index <4) RNA samples, respectively. We describe a strategy for obtaining high-quality and quantity RNA from fresh and stored cells from blood. The results serve as a guideline for sensitive mRNA and microRNA expression assessment in clinical material.

  19. UW is superior to Celsior and HTK in the protection of human liver endothelial cells against preservation injury.

    PubMed

    Janssen, Hermann; Janssen, Petra H E; Broelsch, Christoph E

    2004-12-01

    Celsior solution (CS), a new preservation solution in thoracic organ transplantation, was evaluated for its efficacy in cold preservation of human liver endothelial cells (HLEC) and was compared to University of Wisconsin solution (UW) and histidine-tryptophan-ketoglutarate solution (HTK, Custodiol). HLEC cultures were preserved at 4 degrees C in CS, UW, and HTK, for 2, 6, 12, 24, and 48 hours, with 6 hours of reperfusion. Levels of lactate dehydrogenase (LDH), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and adenosine 5'-triphosphate (ATP) were measured after each interval of ischemia and the respective phase of reperfusion. Preservation injury of HLEC as measured by LDH release, intracellular ATP level, and MTT reduction were overall significantly (P preservation. Furthermore, cold preservation remains the main cause of preservation injury of HLEC regardless of the preservation solution used. Additionally, the maintenance of a high intracellular ATP level of HLEC after ischemia and reperfusion, as shown by UW, could be taken as a beneficial effect, particularly in long-term ischemia. In conclusion, our cell culture model reveals the order of efficacy to protect HLEC against preservation injury as: UW > CS > HTK.

  20. Cryopreservation of testicular tissue or testicular cell suspensions: a pivotal step in fertility preservation

    PubMed Central

    Onofre, J.; Baert, Y.; Faes, K.; Goossens, E.

    2016-01-01

    BACKGROUND Germ cell depletion caused by chemical or physical toxicity, disease or genetic predisposition can occur at any age. Although semen cryopreservation is the first reflex for preserving male fertility, this cannot help out prepubertal boys. Yet, these boys do have spermatogonial stem cells (SSCs) that able to produce sperm at the start of puberty, which allows them to safeguard their fertility through testicular tissue (TT) cryopreservation. SSC transplantation (SSCT), TT grafting and recent advances in in vitro spermatogenesis have opened new possibilities to restore fertility in humans. However, these techniques are still at a research stage and their efficiency depends on the amount of SSCs available for fertility restoration. Therefore, maintaining the number of SSCs is a critical step in human fertility preservation. Standardizing a successful cryopreservation method for TT and testicular cell suspensions (TCSs) is most important before any clinical application of fertility restoration could be successful. OBJECTIVE AND RATIONALE This review gives an overview of existing cryopreservation protocols used in different animal models and humans. Cell recovery, cell viability, tissue integrity and functional assays are taken into account. Additionally, biosafety and current perspectives in male fertility preservation are discussed. SEARCH METHODS An extensive PubMED and MEDline database search was conducted. Relevant studies linked to the topic were identified by the search terms: cryopreservation, male fertility preservation, (immature)testicular tissue, testicular cell suspension, spermatogonial stem cell, gonadotoxicity, radiotherapy and chemotherapy. OUTCOMES The feasibility of fertility restoration techniques using frozen-thawed TT and TCS has been proven in animal models. Efficient protocols for cryopreserving human TT exist and are currently applied in the clinic. For TCSs, the highest post-thaw viability reported after vitrification is 55.6 ± 23

  1. Uptake and metabolism of iron oxide nanoparticles in brain cells.

    PubMed

    Petters, Charlotte; Irrsack, Ellen; Koch, Michael; Dringen, Ralf

    2014-09-01

    Magnetic iron oxide nanoparticles (IONPs) are used for various applications in biomedicine, for example as contrast agents in magnetic resonance imaging, for cell tracking and for anti-tumor treatment. However, IONPs are also known for their toxic effects on cells and tissues which are at least in part caused by iron-mediated radical formation and oxidative stress. The potential toxicity of IONPs is especially important concerning the use of IONPs for neurobiological applications as alterations in brain iron homeostasis are strongly connected with human neurodegenerative diseases. Since IONPs are able to enter the brain, potential adverse consequences of an exposure of brain cells to IONPs have to be considered. This article describes the pathways that allow IONPs to enter the brain and summarizes the current knowledge on the uptake, the metabolism and the toxicity of IONPs for the different types of brain cells in vitro and in vivo.

  2. Application of cell sheet technology to bone marrow stromal cell transplantation for rat brain infarct.

    PubMed

    Ito, Masaki; Shichinohe, Hideo; Houkin, Kiyohiro; Kuroda, Satoshi

    2017-02-01

    Bone marrow stromal cells (BMSC) transplantation enhances functional recovery after cerebral infarct, but the optimal delivery route is undetermined. This study was aimed to assess whether a novel cell-sheet technology non-invasively serves therapeutic benefits to ischemic stroke. First, the monolayered cell sheet was engineered by culturing rat BMSCs on a temperature-responsive dish. The cell sheet was analysed histologically and then transplanted onto the ipsilateral neocortex of rats subjected to permanent middle cerebral artery occlusion at 7 days after the insult. Their behaviours and histology were compared with those in the animals treated with direct injection of BMSCs or vehicle over 4 weeks post-transplantation. The cell sheet was 27.9 ± 8.0 μm thick and was composed of 9.8 ± 2.4 × 10(5) cells. Cell sheet transplantation significantly improved motor function when compared with the vehicle-injected animals. Histological analysis revealed that the BMSCs were densely distributed to the neocortex adjacent to the cerebral infarct and expressed neuronal phenotype in the cell sheet-transplanted animals. These findings were almost equal to those for the animals treated with direct BMSC injection. The attachment of the BMSC sheet to the brain surface did not induce reactive astrocytes in the adjacent neocortex, although direct injection of BMSCs profoundly induced reactive astrocytes around the injection site. These findings suggest that the BMSCs in cell sheets preserve their biological capacity of migration and neural differentiation. Cell-sheet technology may enhance functional recovery after ischaemic stroke, using a less invasive method. Copyright © 2014 John Wiley & Sons, Ltd.

  3. Rescuing Perishable Neuroanatomical Information from a Threatened Biodiversity Hotspot: Remote Field Methods for Brain Tissue Preservation Validated by Cytoarchitectonic Analysis, Immunohistochemistry, and X-Ray Microcomputed Tomography

    PubMed Central

    Hughes, Daniel F.; Walker, Ellen M.; Gignac, Paul M.; Martinez, Anais; Negishi, Kenichiro; Lieb, Carl S.; Greenbaum, Eli

    2016-01-01

    Biodiversity hotspots, which harbor more endemic species than elsewhere on Earth, are increasingly threatened. There is a need to accelerate collection efforts in these regions before threatened or endangered species become extinct. The diverse geographical, ecological, genetic, morphological, and behavioral data generated from the on-site collection of an individual specimen are useful for many scientific purposes. However, traditional methods for specimen preparation in the field do not permit researchers to retrieve neuroanatomical data, disregarding potentially useful data for increasing our understanding of brain diversity. These data have helped clarify brain evolution, deciphered relationships between structure and function, and revealed constraints and selective pressures that provide context about the evolution of complex behavior. Here, we report our field-testing of two commonly used laboratory-based techniques for brain preservation while on a collecting expedition in the Congo Basin and Albertine Rift, two poorly known regions associated with the Eastern Afromontane biodiversity hotspot. First, we found that transcardial perfusion fixation and long-term brain storage, conducted in remote field conditions with no access to cold storage laboratory equipment, had no observable impact on cytoarchitectural features of lizard brain tissue when compared to lizard brain tissue processed under laboratory conditions. Second, field-perfused brain tissue subjected to prolonged post-fixation remained readily compatible with subsequent immunohistochemical detection of neural antigens, with immunostaining that was comparable to that of laboratory-perfused brain tissue. Third, immersion-fixation of lizard brains, prepared under identical environmental conditions, was readily compatible with subsequent iodine-enhanced X-ray microcomputed tomography, which facilitated the non-destructive imaging of the intact brain within its skull. In summary, we have validated

  4. Rescuing Perishable Neuroanatomical Information from a Threatened Biodiversity Hotspot: Remote Field Methods for Brain Tissue Preservation Validated by Cytoarchitectonic Analysis, Immunohistochemistry, and X-Ray Microcomputed Tomography.

    PubMed

    Hughes, Daniel F; Walker, Ellen M; Gignac, Paul M; Martinez, Anais; Negishi, Kenichiro; Lieb, Carl S; Greenbaum, Eli; Khan, Arshad M

    2016-01-01

    Biodiversity hotspots, which harbor more endemic species than elsewhere on Earth, are increasingly threatened. There is a need to accelerate collection efforts in these regions before threatened or endangered species become extinct. The diverse geographical, ecological, genetic, morphological, and behavioral data generated from the on-site collection of an individual specimen are useful for many scientific purposes. However, traditional methods for specimen preparation in the field do not permit researchers to retrieve neuroanatomical data, disregarding potentially useful data for increasing our understanding of brain diversity. These data have helped clarify brain evolution, deciphered relationships between structure and function, and revealed constraints and selective pressures that provide context about the evolution of complex behavior. Here, we report our field-testing of two commonly used laboratory-based techniques for brain preservation while on a collecting expedition in the Congo Basin and Albertine Rift, two poorly known regions associated with the Eastern Afromontane biodiversity hotspot. First, we found that transcardial perfusion fixation and long-term brain storage, conducted in remote field conditions with no access to cold storage laboratory equipment, had no observable impact on cytoarchitectural features of lizard brain tissue when compared to lizard brain tissue processed under laboratory conditions. Second, field-perfused brain tissue subjected to prolonged post-fixation remained readily compatible with subsequent immunohistochemical detection of neural antigens, with immunostaining that was comparable to that of laboratory-perfused brain tissue. Third, immersion-fixation of lizard brains, prepared under identical environmental conditions, was readily compatible with subsequent iodine-enhanced X-ray microcomputed tomography, which facilitated the non-destructive imaging of the intact brain within its skull. In summary, we have validated

  5. Preserving and Using Germplasm and Dissociated Embryonic Cells for Conserving Caribbean and Pacific Coral

    PubMed Central

    Hagedorn, Mary; Carter, Virginia; Martorana, Kelly; Paresa, Malia K.; Acker, Jason; Baums, Iliana B.; Borneman, Eric; Brittsan, Michael; Byers, Michael; Henley, Michael; Laterveer, Michael; Leong, Jo-Ann; McCarthy, Megan; Meyers, Stuart; Nelson, Brian D.; Petersen, Dirk; Tiersch, Terrence; Uribe, Rafael Cuevas; Woods, Erik; Wildt, David

    2012-01-01

    Coral reefs are experiencing unprecedented degradation due to human activities, and protecting specific reef habitats may not stop this decline, because the most serious threats are global (i.e., climate change), not local. However, ex situ preservation practices can provide safeguards for coral reef conservation. Specifically, modern advances in cryobiology and genome banking could secure existing species and genetic diversity until genotypes can be introduced into rehabilitated habitats. We assessed the feasibility of recovering viable sperm and embryonic cells post-thaw from two coral species, Acropora palmata and Fungia scutaria that have diffferent evolutionary histories, ecological niches and reproductive strategies. In vitro fertilization (IVF) of conspecific eggs using fresh (control) spermatozoa revealed high levels of fertilization (>90% in A. palmata; >84% in F. scutaria; P>0.05) that were unaffected by tested sperm concentrations. A solution of 10% dimethyl sulfoxide (DMSO) at cooling rates of 20 to 30°C/min most successfully cryopreserved both A. palmata and F. scutaria spermatozoa and allowed producing developing larvae in vitro. IVF success under these conditions was 65% in A. palmata and 53% in F. scutaria on particular nights; however, on subsequent nights, the same process resulted in little or no IVF success. Thus, the window for optimal freezing of high quality spermatozoa was short (∼5 h for one night each spawning cycle). Additionally, cryopreserved F. scutaria embryonic cells had∼50% post-thaw viability as measured by intact membranes. Thus, despite some differences between species, coral spermatozoa and embryonic cells are viable after low temperature (−196°C) storage, preservation and thawing. Based on these results, we have begun systematically banking coral spermatozoa and embryonic cells on a large-scale as a support approach for preserving existing bio- and genetic diversity found in reef systems. PMID:22413020

  6. Activated Brain Endothelial Cells Cross-Present Malaria Antigen.

    PubMed

    Howland, Shanshan W; Poh, Chek Meng; Rénia, Laurent

    2015-06-01

    In the murine model of cerebral malaria caused by P. berghei ANKA (PbA), parasite-specific CD8+ T cells directly induce pathology and have long been hypothesized to kill brain endothelial cells that have internalized PbA antigen. We previously reported that brain microvessel fragments from infected mice cross-present PbA epitopes, using reporter cells transduced with epitope-specific T cell receptors. Here, we confirm that endothelial cells are the population responsible for cross-presentation in vivo, not pericytes or microglia. PbA antigen cross-presentation by primary brain endothelial cells in vitro confers susceptibility to killing by CD8+ T cells from infected mice. IFNγ stimulation is required for brain endothelial cross-presentation in vivo and in vitro, which occurs by a proteasome- and TAP-dependent mechanism. Parasite strains that do not induce cerebral malaria were phagocytosed and cross-presented less efficiently than PbA in vitro. The main source of antigen appears to be free merozoites, which were avidly phagocytosed. A human brain endothelial cell line also phagocytosed P. falciparum merozoites. Besides being the first demonstration of cross-presentation by brain endothelial cells, our results suggest that interfering with merozoite phagocytosis or antigen processing may be effective strategies for cerebral malaria intervention.

  7. Cell-based therapy for traumatic brain injury.

    PubMed

    Gennai, S; Monsel, A; Hao, Q; Liu, J; Gudapati, V; Barbier, E L; Lee, J W

    2015-08-01

    Traumatic brain injury is a major economic burden to hospitals in terms of emergency department visits, hospitalizations, and utilization of intensive care units. Current guidelines for the management of severe traumatic brain injuries are primarily supportive, with an emphasis on surveillance (i.e. intracranial pressure) and preventive measures to reduce morbidity and mortality. There are no direct effective therapies available. Over the last fifteen years, pre-clinical studies in regenerative medicine utilizing cell-based therapy have generated enthusiasm as a possible treatment option for traumatic brain injury. In these studies, stem cells and progenitor cells were shown to migrate into the injured brain and proliferate, exerting protective effects through possible cell replacement, gene and protein transfer, and release of anti-inflammatory and growth factors. In this work, we reviewed the pathophysiological mechanisms of traumatic brain injury, the biological rationale for using stem cells and progenitor cells, and the results of clinical trials using cell-based therapy for traumatic brain injury. Although the benefits of cell-based therapy have been clearly demonstrated in pre-clinical studies, some questions remain regarding the biological mechanisms of repair and safety, dose, route and timing of cell delivery, which ultimately will determine its optimal clinical use.

  8. Electroporation of Brain Endothelial Cells on Chip toward Permeabilizing the Blood-Brain Barrier

    PubMed Central

    Bonakdar, Mohammad; Wasson, Elisa M.; Lee, Yong W.; Davalos, Rafael V.

    2016-01-01

    The blood-brain barrier, mainly composed of brain microvascular endothelial cells, poses an obstacle to drug delivery to the brain. Controlled permeabilization of the constituent brain endothelial cells can result in overcoming this barrier and increasing transcellular transport across it. Electroporation is a biophysical phenomenon that has shown potential in permeabilizing and overcoming this barrier. In this study we developed a microengineered in vitro model to characterize the permeabilization of adhered brain endothelial cells to large molecules in response to applied pulsed electric fields. We found the distribution of affected cells by reversible and irreversible electroporation, and quantified the uptaken amount of naturally impermeable molecules into the cells as a result of applied pulse magnitude and number of pulses. We achieved 81 ± 1.7% (N = 6) electroporated cells with 17 ± 8% (N = 5) cell death using an electric-field magnitude of ∼580 V/cm and 10 pulses. Our results provide the proper range for applied electric-field intensity and number of pulses for safe permeabilization without significantly compromising cell viability. Our results demonstrate that it is possible to permeabilize the endothelial cells of the BBB in a controlled manner, therefore lending to the feasibility of using pulsed electric fields to increase drug transport across the BBB through the transcellular pathway. PMID:26789772

  9. Electroporation of Brain Endothelial Cells on Chip toward Permeabilizing the Blood-Brain Barrier.

    PubMed

    Bonakdar, Mohammad; Wasson, Elisa M; Lee, Yong W; Davalos, Rafael V

    2016-01-19

    The blood-brain barrier, mainly composed of brain microvascular endothelial cells, poses an obstacle to drug delivery to the brain. Controlled permeabilization of the constituent brain endothelial cells can result in overcoming this barrier and increasing transcellular transport across it. Electroporation is a biophysical phenomenon that has shown potential in permeabilizing and overcoming this barrier. In this study we developed a microengineered in vitro model to characterize the permeabilization of adhered brain endothelial cells to large molecules in response to applied pulsed electric fields. We found the distribution of affected cells by reversible and irreversible electroporation, and quantified the uptaken amount of naturally impermeable molecules into the cells as a result of applied pulse magnitude and number of pulses. We achieved 81 ± 1.7% (N = 6) electroporated cells with 17 ± 8% (N = 5) cell death using an electric-field magnitude of ∼580 V/cm and 10 pulses. Our results provide the proper range for applied electric-field intensity and number of pulses for safe permeabilization without significantly compromising cell viability. Our results demonstrate that it is possible to permeabilize the endothelial cells of the BBB in a controlled manner, therefore lending to the feasibility of using pulsed electric fields to increase drug transport across the BBB through the transcellular pathway.

  10. Impact of stress and mast cells on brain metastases.

    PubMed

    Theoharides, Theoharis C; Rozniecki, Jacek J; Sahagian, Gary; Jocobson, Stanley; Kempuraj, Duraisamy; Conti, Pio; Kalogeromitros, Dimitris

    2008-12-15

    Metastases continue to be the chief cause of morbidity and mortality for many tumors, including brain metastases of lung and mammary adenocarcinoma. Stress appears to increase metastases, but the mechanism is not understood. Recent evidence suggests that local inflammation is conducive for cancer growth and a unique immune cell, the mast cell, accumulates in the stroma surrounding tumors and is critically located at the blood-brain-barrier (BBB). Mast cells express receptors for and can be stimulated by corticotropin-releasing hormone (CRH), secreted under stress, to release mediators such as histamine, IL-8, tryptase and vascular endothelial growth factor (VEGF), which disrupt the BBB permitting metastases. Stress and mast cells could serve as new targets for drug development to prevent brain metastases, especially since CRH receptor antagonists and brain mast cell inhibitors have recently been developed.

  11. Germ cell transplantation in felids: a potential approach to preserving endangered species.

    PubMed

    Silva, Robson C; Costa, Guilherme M J; Lacerda, Samyra M S N; Batlouni, Sérgio R; Soares, Jaqueline M; Avelar, Gleide F; Böttger, Karin B; Silva, Silvério F; Nogueira, Maria S; Andrade, Leonardo M; França, Luiz R

    2012-01-01

    With the exception of the domestic cat, all members of the family Felidae are considered either endangered or threatened. Although not yet used for this purpose, spermatogonial stem cell (SSC) transplantation has a high potential to preserve the genetic stock of endangered species. However, this technique has not previously been established in felids. Therefore, we developed the necessary procedures to perform syngeneic and xenogeneic SSC transplants (eg, germ cell [GC] depletion in the recipient domestic cats, enrichment and labeling of donor cell suspension, and the transplantation method) in order to investigate the feasibility of the domestic cat as a recipient for the preservation and propagation of male germ plasm from wild felids. In comparison with busulfan treatment, local x-ray fractionated radiation was a more effective approach to depleting endogenous spermatogenesis. The results of both syngeneic and xenogeneic transplants revealed that SSCs were able to successfully colonize and differentiate in the recipient testis, generating elongated spermatids several weeks posttransplantation. Specifically, ocelot spermatozoa were observed in the cat epididymis 13 weeks following transplantation. As donor GCs from domestic cats and ocelots were able to develop and form mature GCs in the recipient environment seminiferous tubules, these findings indicate that the domestic cat is a suitable recipient for SSC transplantation. Moreover, as modern cats descended from a medium-size cat that existed approximately 10 to 11 million years ago, these results strongly suggest that the domestic cat could be potentially used as a recipient for generating and propagating the genome of wild felids.

  12. Bone marrow–derived stem cells preserve cone vision in retinitis pigmentosa

    PubMed Central

    Smith, Lois E.H.

    2004-01-01

    Retinitis pigmentosa is a heritable group of blinding diseases resulting from loss of photoreceptors, primarily rods and secondarily cones, that mediate central vision. Loss of retinal vasculature is a presumed metabolic consequence of photoreceptor degeneration. A new study shows that autologous bone marrow–derived lineage-negative hematopoietic stem cells, which incorporate into the degenerating blood vessels in two murine models of retinitis pigmentosa, rd1 and rd10, prevent cone loss. The use of autologous bone marrow might avoid problems with rejection while preserving central cone vision in a wide variety of genetically disparate retinal degenerative diseases. PMID:15372096

  13. Presence of Germ Cells in Disorders of Sex Development: Implications for Fertility Potential and Preservation

    PubMed Central

    Finlayson, Courtney; Fritsch, Michael K.; Johnson, Emilie K.; Rosoklija, Ilina; Gosiengfiao, Yasmin; Yerkes, Elizabeth; Madonna, Mary Beth; Woodruff, Teresa K.; Cheng, Earl

    2017-01-01

    Purpose We sought to determine the presence of germ cells in the gonads of patients with disorders of sex development to establish whether preservation of germ cells for future fertility potential is possible. We hypothesized that germ cells are present but vary by age and diagnosis. Materials and Methods We reviewed histology from patients with disorders of sex development who underwent gonadectomy/biopsy from 2002 to 2014 at a single institution for pathological classification of the gonad, composition of gonadal stroma and germ cell presence. Results A total of 44 patients were identified and germ cells were present in 68%. The presence and average number of germ cells per mm2 were analyzed by gonad type and diagnosis. By gonad type all ovotestes, most testes, ovaries and dysgenetic testes, and 15% of streak gonads had germ cells present. By diagnosis germ cells were present in all patients with complete androgen insensitivity syndrome, Denys-Drash syndrome, SRY mutation, mixed gonadal dysgenesis, ovotesticular conditions and StAR (steroid acute regulatory protein) deficiency, in some patients with persistent müllerian duct syndrome, XO/XY Turner syndrome and disorders of sex development not otherwise specified, and in none with complete or partial gonadal dysgenesis. Germ cells were present in the gonads of 88% of patients 0 to 3 years old, 50% of those 4 to 11 years old and 43% of those older than 12 years. Conclusions Germ cells were present in the majority of our cohort and the presence decreased with age. This novel, fertility driven evaluation of germ cell quantity in a variety of disorders of sex development suggests that fertility potential may be greater than previously thought. Further studies must be done to evaluate a larger population and examine germ cell quality to determine the viability of these germ cells. PMID:27840018

  14. Preservation of diffusion tensor properties during spatial normalization by use of tensor imaging and fibre tracking on a normal brain database.

    PubMed

    Müller, H-P; Unrath, A; Ludolph, A C; Kassubek, J

    2007-03-21

    White matter connectivity in the human brain can be mapped by diffusion tensor magnetic resonance imaging (DTI). After reconstruction, the diffusion tensors, the diffusion amplitude and the diffusion direction can be displayed on a morphological background. Consequently, diffusion tensor fibre tracking can be applied as a non-invasive in vivo technique for the delineation and quantification of specific white matter pathways. The aim of this study was to show that normalization to the Montreal Neurological Institute (MNI) stereotaxic standard space preserves specific diffusion features. Therefore, techniques for tensor imaging and fibre tracking were applied to the normalized brains as well as to the group averaged brain data. A normalization step of individual data was included by registration to a scanner- and sequence-specific DTI template data set which was created from a normal database transformed to MNI space. The algorithms were tested and validated for a group of 13 healthy controls.

  15. Expansion of Multipotent Stem Cells from the Adult Human Brain

    PubMed Central

    Murrell, Wayne; Palmero, Emily; Bianco, John; Stangeland, Biljana; Joel, Mrinal; Paulson, Linda; Thiede, Bernd; Grieg, Zanina; Ramsnes, Ingunn; Skjellegrind, Håvard K.; Nygård, Ståle; Brandal, Petter; Sandberg, Cecilie; Vik-Mo, Einar; Palmero, Sheryl; Langmoen, Iver A.

    2013-01-01

    The discovery of stem cells in the adult human brain has revealed new possible scenarios for treatment of the sick or injured brain. Both clinical use of and preclinical research on human adult neural stem cells have, however, been seriously hampered by the fact that it has been impossible to passage these cells more than a very few times and with little expansion of cell numbers. Having explored a number of alternative culturing conditions we here present an efficient method for the establishment and propagation of human brain stem cells from whatever brain tissue samples we have tried. We describe virtually unlimited expansion of an authentic stem cell phenotype. Pluripotency proteins Sox2 and Oct4 are expressed without artificial induction. For the first time multipotency of adult human brain-derived stem cells is demonstrated beyond tissue boundaries. We characterize these cells in detail in vitro including microarray and proteomic approaches. Whilst clarification of these cells’ behavior is ongoing, results so far portend well for the future repair of tissues by transplantation of an adult patient’s own-derived stem cells. PMID:23967194

  16. Brain mast cell relationship to neurovasculature during development.

    PubMed

    Khalil, Mona; Ronda, Jocelyn; Weintraub, Michael; Jain, Kim; Silver, Rae; Silverman, Ann-Judith

    2007-09-26

    Mast cells, derived from the hematopoietic stem cell, are present in the brain from birth. During development, mast cells occur in two locations, namely the pia and the brain parenchyma. The current hypothesis regarding their origin states that brain mast cells (or their precursors) enter the pia and access the thalamus by traveling along the abluminal wall of penetrating blood vessels. The population in the pia reaches a maximum at postnatal (PN) day 11, and declines rapidly thereafter. Chromatin fragmentation suggests that this cell loss is due to apoptosis. In contrast, the thalamic population expands from PN8 to reach adult levels at PN30. Stereological analysis demonstrates that mast cells home to blood vessels. More than 96% of mast cells are inside the blood-brain barrier, with ~90% contacting the blood vessel wall or its extracellular matrix. Mast cells express alpha4 integrins -- a potential mechanism for adhesion to the vascular wall. Despite the steady increase in the volume of microvasculature, at all ages studied, mast cells are preferentially located on large diameter vessels (>16 microm; possibly arteries), and contact only those maturing blood vessels that are ensheathed by astroglial processes. Mast cells not only home to large vessels but also maintain a preferential position at branch points, sites of vessel growth. This observation presents the possibility that mast cells participate in and/or regulate vasculature growth or differentiation. The biochemical and molecular signals that induce mast cell homing in the CNS is an area of active investigation.

  17. NMR imaging of cell phone radiation absorption in brain tissue

    PubMed Central

    Gultekin, David H.; Moeller, Lothar

    2013-01-01

    A method is described for measuring absorbed electromagnetic energy radiated from cell phone antennae into ex vivo brain tissue. NMR images the 3D thermal dynamics inside ex vivo bovine brain tissue and equivalent gel under exposure to power and irradiation time-varying radio frequency (RF) fields. The absorbed RF energy in brain tissue converts into Joule heat and affects the nuclear magnetic shielding and the Larmor precession. The resultant temperature increase is measured by the resonance frequency shift of hydrogen protons in brain tissue. This proposed application of NMR thermometry offers sufficient spatial and temporal resolution to characterize the hot spots from absorbed cell phone radiation in aqueous media and biological tissues. Specific absorption rate measurements averaged over 1 mg and 10 s in the brain tissue cover the total absorption volume. Reference measurements with fiber optic temperature sensors confirm the accuracy of the NMR thermometry. PMID:23248293

  18. NMR imaging of cell phone radiation absorption in brain tissue.

    PubMed

    Gultekin, David H; Moeller, Lothar

    2013-01-02

    A method is described for measuring absorbed electromagnetic energy radiated from cell phone antennae into ex vivo brain tissue. NMR images the 3D thermal dynamics inside ex vivo bovine brain tissue and equivalent gel under exposure to power and irradiation time-varying radio frequency (RF) fields. The absorbed RF energy in brain tissue converts into Joule heat and affects the nuclear magnetic shielding and the Larmor precession. The resultant temperature increase is measured by the resonance frequency shift of hydrogen protons in brain tissue. This proposed application of NMR thermometry offers sufficient spatial and temporal resolution to characterize the hot spots from absorbed cell phone radiation in aqueous media and biological tissues. Specific absorption rate measurements averaged over 1 mg and 10 s in the brain tissue cover the total absorption volume. Reference measurements with fiber optic temperature sensors confirm the accuracy of the NMR thermometry.

  19. Salvage surgery for locoregional recurrences of advanced pharyngolaryngeal squamous cell carcinoma after organ preservation failure.

    PubMed

    López Delgado, I; Riestra Ayora, J; Arenas Brítez, O; García López, I; Martínez Guirado, T; Scola Yurrita, B

    2014-12-01

    Organ preservation treatment for advanced head and neck squamous cell carcinoma is associated with poor outcomes due to locoregional recurrences. Salvage surgery is the main therapeutic option for some of these patients. The aim of this study was to analyse the results of salvage surgery for advanced pharyngolaryngeal squamous cell carcinoma previously treated with radiochemotherapy. We performed a retrospective study on 38 patients (36 men, 2 women). The median age at diagnosis was 60 years with a mean follow-up period of 49.8 months. Recurrences were diagnosed at a mean of 395 days after finalising organ preservation treatment. Patients went under different salvage surgeries, including 22 total laryngectomies, 6 partial laryngectomies (3 transoral laser surgeries and 3 opened surgeries), 8 functional neck dissections and 2 tongue base surgeries. Nineteen patients had no postoperative complications after a mean hospital stay of 2 weeks. However, 5 patients died of significant recurrent bleedings. There were 4 salivary fistulas that responded to conservative management, while 7 patients had important pharyngostomas that required reconstruction with either regional or free flaps. The mean hospital stay was of 61.60 days for all patients. Five-year overall survival from diagnosis, overall survival after salvage surgery and survival after salvage surgery were 44.20, 37.90 and 45.70%, respectively. In summary, we conclude that salvage surgery is an optimal treatment for pharyngolaryngeal and regional recurrences and provides improvement in locoregional control and survival, despite the severe complications.

  20. Investigations on the change of texture of plant cells due to preservative treatments by digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Vora, Priyanka; Anand, Arun

    2014-10-01

    Texture change is observed in preserved fruits and vegetables. Responsible factors for texture change during preservative treatments are cell morphology, cell wall structure, cell turger, water content and some biochemical components, and also the environmental conditions. Digital Holographic microscopy (DHM) is a quantitative phase contrast imaging technique, which provides three dimensional optical thickness profiles of transparent specimen. Using DHM the morphology of plant cells preserved by refrigeration or stored in vinegar or in sodium chloride can be obtained. This information about the spatio-temporal evolution of optical volume and thickness can be an important tool in area of food processing. Also from the three dimensional images, the texture of the cell can be retrieved and can be investigated under varying conditions.

  1. Preparation and preservation of viable Akkermansia muciniphila cells for therapeutic interventions.

    PubMed

    Ouwerkerk, J P; Aalvink, S; Belzer, C; De Vos, W M

    2017-01-24

    The anaerobic gut bacterium Akkermansia muciniphila is a well-characterised member of the mucosal microbiota and has shown to be a gut symbiont in human. A. muciniphila has been negatively associated with obesity and its associated metabolic disorders in various human cohorts while treatment with A. muciniphila cells reversed highfat diet-induced obesity and its associated metabolic disorders in mouse models. Therefore, administration of A. muciniphila has been suggested as a possible new therapeutic treatment for these omnipresent diseases. Here we describe a potentially scalable workflow for the preparation and preservation of high numbers of viable cells of A. muciniphila obtained from 1 l laboratory scale growth under strict anaerobic conditions for therapeutic interventions. This resulted in viable A. muciniphila cells with high yields and very high stability, with up to 97.9±4.5% survival for a time period of 1 year at -80 °C in glycerol-amended medium. Moreover, various quality assessment and control procedures were developed to ensure the use of viable cells of A. muciniphila. Several microscopic, culturing, and molecular approaches were applied to monitor the presence, abundance and recovery of A. muciniphila before, during, and after its administration to high-fat treated mice. We show that viable A. muciniphila cells can be recovered from caecal and colon content (up to 1×10(10) cells/g), testifying for the efficiency of the described workflow.

  2. Preservation of epithelial progenitor cells from collagenase-digested oral mucosa during ex vivo cultivation

    PubMed Central

    Hsueh, Yi-Jen; Huang, Shiang-Fu; Lai, Jui-Yang; Ma, Shih-Chieh; Chen, Hung-Chi; Wu, Sung-En; Wang, Tze-Kai; Sun, Chi-Chin; Ma, Kevin Sheng-Kai; Chen, Jan-Kan; Lai, Chyong-Huey; Ma, David Hui-Kang

    2016-01-01

    To avoid xenogeneic infection, we report a novel protocol for producing animal-derived component-free oral mucosal epithelial cells (OMECs) sheet for transplantation, in which collagenase was used to replace dispase II/trypsin-EDTA for digesting oral mucosal tissue, and human platelet-derived PLTMax to replace fetal bovine serum. The resulting epithelial aggregates were expanded on de-epithelialized amniotic membranes without 3T3 feeder cells, and serum-free EpiLife was used to reduce contamination by submucosal mesenchymal cells. The OMEC sheets thus generated showed similar positive keratin 3/76-positive and keratin 8-negative staining patterns compared with those generated by the original protocol. Colony formation efficiency assay, BrdU label retention assay, and p63 and p75NTR immunostaining results indicated that higher proliferative potentials and more progenitor cells were preserved by the modified protocol. TaqMan array analysis revealed that the transcription of integrin-linked kinase (ILK) was up-regulated along with an increase in β-catenin signaling and its downstream cell cycle modulators, cyclin D1 and p27KIP1. Furthermore, ILK silencing led to the inhibition of nuclear β-catenin accumulation, suppressed p63 expression, and reduced the expression of cyclin D1 and p27KIP1; these observations suggest that ILK/β-catenin pathway may be involved in cell proliferation regulation during the ex vivo expansion of OMECs for transplantation purposes. PMID:27824126

  3. Clear cell carcinoma of the uterine cervix: clinical characteristics and feasibility of fertility-preserving treatment

    PubMed Central

    Jiang, Xiang; Jin, Ying; Li, Yan; Huang, Hui-Fang; Wu, Ming; Shen, Keng; Pan, Ling-Ya

    2014-01-01

    The objective of this retrospective study was to analyze the clinical characteristics and prognosis of clear cell adenocarcinoma (CCA) in the post-diethylstilbestrol (DES) era and to evaluate the feasibility of fertility-preserving treatment. The records of 32 patients with CCAs who were treated at Peking Union Medical College Hospital from August 1986 to June 2012 were retrospectively reviewed. Three of the patients had undergone fertility-preserving treatment. The incidence of CCA among cervical adenocarcinomas was 15.2%. The median age was 38 years: 11 patients (34.4%) were diagnosed before 30 years of age and two (6.3%) after 70 years of age. Ten patients (31.2%) were nulliparous. No patient had been exposed to DES. Twenty-nine patients (90.6%) presented with obvious symptoms, and the cervix appeared abnormal in 26 patients (81.3%). Cervical Papanicolaou (Pap) tests were abnormal in all four patients in whom they were performed (three had high-grade squamous intraepithelial lesions and one had atypical squamous cells of undetermined significance). The distribution by stage was 56.3% stage I, 34.4% stage II, 6.3% stage III, and 3.1% stage IV. Treatments mainly included surgery for patients with stage I to IIA CCA and radiochemotherapy for patients with advanced CCA. The overall 5-year progression-free survival was 72.2%. Patients with stage I to IIA CCA had better 5-year progression-free survival than did patients with stage IIB to IV CCA (81.5% versus 40.0%, P=0.003). The three patients who had undergone fertility-preserving treatment had no recurrences. CCA may also affect adolescents and children without prior DES exposure, who are often misdiagnosed as having functional uterine bleeding. Radiotherapy appears to be effective for local control but to have no effect on distant recurrences. In our study, the prognosis of patients with early-stage CCA, including those who had undergone fertility-preserving treatment, was not inferior to that of patients with

  4. Cell fusion in the brain: two cells forward, one cell back.

    PubMed

    Kemp, Kevin; Wilkins, Alastair; Scolding, Neil

    2014-11-01

    Adult stem cell populations, notably those which reside in the bone marrow, have been shown to contribute to several neuronal cell types in the rodent and human brain. The observation that circulating bone marrow cells can migrate into the central nervous system and fuse with, in particular, cerebellar Purkinje cells has suggested, at least in part, a potential mechanism behind this process. Experimentally, the incidence of cell fusion in the brain is enhanced with age, radiation exposure, inflammation, chemotherapeutic drugs and even selective damage to the neurons themselves. The presence of cell fusion, shown by detection of increased bi-nucleated neurons, has also been described in a variety of human central nervous system diseases, including both multiple sclerosis and Alzheimer's disease. Accumulating evidence is therefore raising new questions into the biological significance of cell fusion, with the possibility that it represents an important means of cell-mediated neuroprotection or rescue of highly complex neurons that cannot be replaced in adult life. Here, we discuss the evidence behind this phenomenon in the rodent and human brain, with a focus on the subsequent research investigating the physiological mechanisms of cell fusion underlying this process. We also highlight how these studies offer new insights into endogenous neuronal repair, opening new exciting avenues for potential therapeutic interventions against neurodegeneration and brain injury.

  5. Rhinovirus Load Is High despite Preserved Interferon-β Response in Cystic Fibrosis Bronchial Epithelial Cells

    PubMed Central

    Cammisano, Maria; Chen, He; Singh, Sareen; Kooi, Cora; Leigh, Richard; Beaudoin, Trevor; Rousseau, Simon; Lands, Larry C.

    2015-01-01

    Lung disease in cystic fibrosis (CF) is often exacerbated following acute upper respiratory tract infections caused by the human rhinovirus (HRV). Pathophysiology of these exacerbations is presently unclear and may involve deficient innate antiviral or exaggerated inflammatory responses in CF airway epithelial cells. Furthermore, responses of CF cells to HRV may be adversely affected by pre-exposure to virulence factors of Pseudomonas (P.) aeruginosa, the microorganism that frequently colonizes CF airways. Here we examined production of antiviral cytokine interferon-β and inflammatory chemokine interleukin-8, expression of the interferon-responsive antiviral gene 2’-5’-oligoadenylate synthetase 1 (OAS1), and intracellular virus RNA load in primary CF (delF508 CFTR) and healthy airway epithelial cells following inoculation with HRV16. Parallel cells were exposed to virulence factors of P. aeruginosa prior to and during HRV16 inoculation. CF cells exhibited production of interferon-β and interleukin-8, and expression of OAS1 at levels comparable to those in healthy cells, yet significantly higher HRV16 RNA load during early hours post-inoculation with HRV16. In line with this, HRV16 RNA load was higher in the CFBE41o- dF cell line overexpessing delF508 CFTR, compared with the isogenic control CFBE41o- WT (wild-type CFTR). Pre-exposure to virulence factors of P. aeruginosa did not affect OAS1 expression or HRV16 RNA load, but potentiated interleukin-8 production. In conclusion, CF cells demonstrate elevated HRV RNA load despite preserved interferon-β and OAS1 responses. High HRV load in CF airway epithelial cells appears to be due to deficiencies manifesting early during HRV infection, and may not be related to interferon-β. PMID:26599098

  6. High-Throughput Single-Cell Manipulation in Brain Tissue

    PubMed Central

    Steinmeyer, Joseph D.; Yanik, Mehmet Fatih

    2012-01-01

    The complexity of neurons and neuronal circuits in brain tissue requires the genetic manipulation, labeling, and tracking of single cells. However, current methods for manipulating cells in brain tissue are limited to either bulk techniques, lacking single-cell accuracy, or manual methods that provide single-cell accuracy but at significantly lower throughputs and repeatability. Here, we demonstrate high-throughput, efficient, reliable, and combinatorial delivery of multiple genetic vectors and reagents into targeted cells within the same tissue sample with single-cell accuracy. Our system automatically loads nanoliter-scale volumes of reagents into a micropipette from multiwell plates, targets and transfects single cells in brain tissues using a robust electroporation technique, and finally preps the micropipette by automated cleaning for repeating the transfection cycle. We demonstrate multi-colored labeling of adjacent cells, both in organotypic and acute slices, and transfection of plasmids encoding different protein isoforms into neurons within the same brain tissue for analysis of their effects on linear dendritic spine density. Our platform could also be used to rapidly deliver, both ex vivo and in vivo, a variety of genetic vectors, including optogenetic and cell-type specific agents, as well as fast-acting reagents such as labeling dyes, calcium sensors, and voltage sensors to manipulate and track neuronal circuit activity at single-cell resolution. PMID:22536416

  7. Brain tissue banking for stem cells for our future

    PubMed Central

    Palmero, Emily; Palmero, Sheryl; Murrell, Wayne

    2016-01-01

    In our lab we study neurogenesis and the development of brain tumors. We work towards treatment strategies for glioblastoma and towards using autologous neural stem cells for tissue regeneration strategies for brain damage and neurodegenerative disorders. It has been our policy to try to establish living cell cultures from all human biopsy material that we obtain. We hypothesized that small pieces of brain tissue could be cryopreserved and that live neural stem cells could be recovered at a later time. DMSO has been shown to possess a remarkable ability to diffuse through cell membranes and pass into cell interiors. Its chemical properties prevent the formation of damaging ice crystals thus allowing cell storage at or below −180 C. We report here a protocol for successful freezing of small pieces of tissue derived from human brain and human brain tumours. Virtually all specimens could be successfully revived. Assays of phenotype and behaviour show that the cell cultures derived were equivalent to those cultures previously derived from fresh tissue. PMID:27991551

  8. Stem cells for brain repair in neonatal hypoxia-ischemia.

    PubMed

    Chicha, L; Smith, T; Guzman, R

    2014-01-01

    Neonatal hypoxic-ischemic insults are a significant cause of pediatric encephalopathy, developmental delays, and spastic cerebral palsy. Although the developing brain's plasticity allows for remarkable self-repair, severe disruption of normal myelination and cortical development upon neonatal brain injury are likely to generate life-persisting sensory-motor and cognitive deficits in the growing child. Currently, no treatments are available that can address the long-term consequences. Thus, regenerative medicine appears as a promising avenue to help restore normal developmental processes in affected infants. Stem cell therapy has proven effective in promoting functional recovery in animal models of neonatal hypoxic-ischemic injury and therefore represents a hopeful therapy for this unmet medical condition. Neural stem cells derived from pluripotent stem cells or fetal tissues as well as umbilical cord blood and mesenchymal stem cells have all shown initial success in improving functional outcomes. However, much still remains to be understood about how those stem cells can safely be administered to infants and what their repair mechanisms in the brain are. In this review, we discuss updated research into pathophysiological mechanisms of neonatal brain injury, the types of stem cell therapies currently being tested in this context, and the potential mechanisms through which exogenous stem cells might interact with and influence the developing brain.

  9. The topographical properties of silica nanoparticle film preserve the osteoblast-like cell characteristics in vitro

    NASA Astrophysics Data System (ADS)

    Shim, Wooyoung; Lee, Seung Yun; Kim, Hyo-Sop; Kim, Jae-Ho

    2016-07-01

    The Transplantation of osteoblasts, along with an artificial implant, is experimentally considered as a therapeutics for degenerative bone diseases. However, osteoblasts have several limitations for application of transplantation in therapeutics, including a low-efficiency for bone mineralization and easy loss of characteristics in in vitro culture condition. In this study, we fabricated silica nano-particle (SNP) films using particles of different sizes to culture osteoblast-like cells for analysis the effect of topography on cellular behavior and characteristics. The physical parameters of films, such as intervals, height and roughness, were proportionally increased depending on the SNP diameter. When osteoblast-like cells were cultured on the various SNP films, the cell attachment rate on SNP-300 and SNP-700 was significantly decreased when it compared to tissue culture polystyrene (TCPS) group. In addition, the genes responsible for cell adhesion showed differential expression profiles in SNP films. The expression and activity of alkaline phosphatase were elevated in SNP-300 and SNP-700, and the extra-cellular matrix and osteoblast marker showed increased gene expression in these SNP films when compared to TCPS group. In the present study, we demonstrate that the topographical property of a nano-scale structure preserves the characteristics of osteoblast-like cells, and regulates the cellular behavior.

  10. Training stem cells for treatment of malignant brain tumors.

    PubMed

    Li, Shengwen Calvin; Kabeer, Mustafa H; Vu, Long T; Keschrumrus, Vic; Yin, Hong Zhen; Dethlefs, Brent A; Zhong, Jiang F; Weiss, John H; Loudon, William G

    2014-09-26

    The treatment of malignant brain tumors remains a challenge. Stem cell technology has been applied in the treatment of brain tumors largely because of the ability of some stem cells to infiltrate into regions within the brain where tumor cells migrate as shown in preclinical studies. However, not all of these efforts can translate in the effective treatment that improves the quality of life for patients. Here, we perform a literature review to identify the problems in the field. Given the lack of efficacy of most stem cell-based agents used in the treatment of malignant brain tumors, we found that stem cell distribution (i.e., only a fraction of stem cells applied capable of targeting tumors) are among the limiting factors. We provide guidelines for potential improvements in stem cell distribution. Specifically, we use an engineered tissue graft platform that replicates the in vivo microenvironment, and provide our data to validate that this culture platform is viable for producing stem cells that have better stem cell distribution than with the Petri dish culture system.

  11. Could Cord Blood Cell Therapy Reduce Preterm Brain Injury?

    PubMed Central

    Li, Jingang; McDonald, Courtney A.; Fahey, Michael C.; Jenkin, Graham; Miller, Suzanne L.

    2014-01-01

    Major advances in neonatal care have led to significant improvements in survival rates for preterm infants, but this occurs at a cost, with a strong causal link between preterm birth and neurological deficits, including cerebral palsy (CP). Indeed, in high-income countries, up to 50% of children with CP were born preterm. The pathways that link preterm birth and brain injury are complex and multifactorial, but it is clear that preterm birth is strongly associated with damage to the white matter of the developing brain. Nearly 90% of preterm infants who later develop spastic CP have evidence of periventricular white matter injury. There are currently no treatments targeted at protecting the immature preterm brain. Umbilical cord blood (UCB) contains a diverse mix of stem and progenitor cells, and is a particularly promising source of cells for clinical applications, due to ethical and practical advantages over other potential therapeutic cell types. Recent studies have documented the potential benefits of UCB cells in reducing brain injury, particularly in rodent models of term neonatal hypoxia–ischemia. These studies indicate that UCB cells act via anti-inflammatory and immuno-modulatory effects, and release neurotrophic growth factors to support the damaged and surrounding brain tissue. The etiology of brain injury in preterm-born infants is less well understood than in term infants, but likely results from episodes of hypoperfusion, hypoxia–ischemia, and/or inflammation over a developmental period of white matter vulnerability. This review will explore current knowledge about the neuroprotective actions of UCB cells and their potential to ameliorate preterm brain injury through neonatal cell administration. We will also discuss the characteristics of UCB-derived from preterm and term infants for use in clinical applications. PMID:25346720

  12. Your brain under the microscope: the promise of stem cells.

    PubMed

    Marchetto, Maria C; Gage, Fred H

    2014-01-01

    Until recently, scientists primarily worked with two kinds of stem cells from animals and humans: embryonic stem cells and non-embryonic "somatic" or "adult" stem cells. Scientists are just now beginning to improve their understanding of a third kind: induced pluripotent stem cells. Our authors describe how they were discovered, what they are, and why a growing number of researchers and clinicians believe that they may be one of the keys in helping address various brain disorders.

  13. Mast Cell - Glia Dialogue in Chronic Pain and Neuropathic Pain: Blood-Brain Barrier Implications.

    PubMed

    Skaper, Stephen D

    2016-01-01

    Mast cells and microglia, working singly and in partnership, produce proinflammatory agents which play key roles in a wide array of nervous system disorders. Such neuroinflammatory settings may compromise integrity of both the blood-nerve barrier and blood-brain barrier (BBB) and blood-spinal cord barrier. While both belong to the innate immune system mast cells are far more ubiquitous, are resident in peripheral nerves and the central nervous system, and can influence blood-nerve barrier characteristics. Mast cells, being near the perivasculature especially within the dura, on the brain side of the BBB, are strategically located to play havoc with the BBB. Mast cells and glia are endowed with homeostatic mechanisms/molecules which come into play following tissue damage. These include the N-acylethanolamine family, especially N-palmitoylethanolamine, which is posited to be a key player in maintaining cellular homeostasis against external stressors provoking, for example, inflammation. This review is intended as an overview covering the pathobiology of neuroinflammation in the context of mast cells and microglia, their role in BBB integrity, and therapeutic perspectives in targeting these cells to preserve BBB function.

  14. Preservation of cranial nerves during removal of the brain for an enhanced student experience in neuroanatomy classes.

    PubMed

    Long, Jennifer; Roberts, David J H; Pickering, James D

    2014-01-01

    Neuroanatomy teaching at the University of Leeds includes the examination of isolated brains by students working in small groups. This requires the prosected brains to exhibit all 12 pairs of cranial nerves. Traditional methods of removing the brain from the skull involve elevating the frontal lobes and cutting each cranial nerve as the brain is reflected posteriorly. This can leave a substantial length of each nerve attached to the skull base rather than to the removed brain. We have found a posterior approach more successful. In this study, five adult heads were disarticulated at the level of the thyroid cartilage and placed, prone, in a head stand. A wedge of bone from the occipital region was removed before the cerebellum and brainstem were elevated to visualize the cranial nerves associated with the medulla oblongata, cerebellopontine angle and mesencephalic-pontine junction prior to cutting them as close to the skull as possible. Five brains were successfully removed from the skull, each having a full complement of cranial nerves of good length attached to them. This approach significantly increases the length and number of cranial nerves remaining attached to the brain, which supports student education. For integration into head and neck dissection courses, careful consideration will be required to ensure the necks are suitably dissected and to decide whether the cranial nerves are best left attached to the skull base or brain.

  15. Recellularization of Well-Preserved Acellular Kidney Scaffold Using Embryonic Stem Cells

    PubMed Central

    Bonandrini, Barbara; Figliuzzi, Marina; Papadimou, Evangelia; Morigi, Marina; Perico, Norberto; Casiraghi, Federica; Sangalli, Fabio; Conti, Sara; Benigni, Ariela; Remuzzi, Giuseppe

    2014-01-01

    For chronic kidney diseases, there is little chance that the vast majority of world's population will have access to renal replacement therapy with dialysis or transplantation. Tissue engineering would help to address this shortcoming by regeneration of damaged kidney using naturally occurring scaffolds seeded with precursor renal cells. The aims of the present study were to optimize the production of three-dimensional (3D) rat whole-kidney scaffolds by shortening the duration of organ decellularization process using detergents that avoid nonionic compounds, to investigate integrity of extracellular matrix (ECM) structure and to enhance the efficacy of scaffold cellularization using physiological perfusion method. Intact rat kidneys were successfully decellularized after 17 h perfusion with sodium dodecyl sulfate. The whole-kidney scaffolds preserved the 3D architecture of blood vessels, glomeruli, and tubuli as shown by transmission and scanning electron microscopy. Micro-computerized tomography (micro-CT) scan confirmed integrity, patency, and connection of the vascular network. Collagen IV, laminin, and fibronectin staining of decellularized scaffolds were similar to those of native kidney tissues. After infusion of whole-kidney scaffolds with murine embryonic stem (mES) cells through the renal artery, and pressure-controlled perfusion with recirculating cell medium for 24 and 72 h, seeded cells were almost completely retained into the organ and uniformly distributed in the vascular network and glomerular capillaries without major signs of apoptosis. Occasionally, mES cells reached peritubular capillary and tubular compartment. We observed the loss of cell pluripotency and the start of differentiation toward meso-endodermal lineage. Our findings indicate that, with the proposed optimized protocol, rat kidneys can be efficiently decellularized to produce renal ECM scaffolds in a relatively short time, and rapid recellularization of vascular structures and

  16. Effects of benzalkonium chloride- or polyquad-preserved fixed combination glaucoma medications on human trabecular meshwork cells

    PubMed Central

    Ammar, David A.

    2011-01-01

    Purpose We investigated the potential short and long-term effects in cultured human trabecular meshwork (TM) cells of various topical glaucoma formulations containing different preservatives. Methods We tested the fixed combination medications 0.004% travoprost plus 0.5% timolol preserved with either 0.015% benzalkonium chloride (BAK; DuoTrav®), or with 0.001% polyquad (PQ; DuoTrav® BAK-free); and 0.005% latanoprost plus 0.5% timolol preserved with 0.020% BAK (Xalacom®). Also tested was a range of BAK concentrations (0.001%–0.020%) in balanced salt solution (BSS). Cells were treated for 25 min at 37 °C with solutions diluted 1:10 and 1:100 to mimic the reduced penetration of topical preparations to the anterior chamber. The percentage of live cells was determined immediately after treatment through the uptake of the fluorescent vital dye calcein-AM. To determine any long-term effects, we assayed release of matrix metalloproteinase 9 (MMP-9) and apoptosis 24 h after treatments. Results BAK demonstrated a dose-dependent reduction in TM cell viability, ranging from 71±5% live cells at 0.001% BAK (diluted 1:10) to 33±3% live cells at 0.020% BAK (diluted 1:10). Travoprost (0.004%) plus 0.5% timolol preserved with 0.015% BAK had statistically fewer live TM cells (79±7%) than the same preparation preserved with 0.001% polyquad® (PQ; 93±1%; p<0.001). Latanoprost plus timolol preserved with 0.020% BAK (29±9% live cells) was similar to the 0.020% BAK (33±3%) treatment. However, travoprost plus timolol preserved in 0.015% BAK had significantly more live cells (83±12%) than the 1:10 dilution of 0.015% BAK (49±10%). We also found 0.020% BAK (diluted 1:100) resulted in elevated levels of extracellular MMP-9 at 24 h. Conclusions These results demonstrate that the substitution of the preservative BAK from topical ophthalmic drugs results in greater in vitro viability of TM cells. Travoprost with timolol, but not latanoprost with timolol, countered some of the toxic

  17. Brain and atrial natriuretic peptides bind to common receptors in brain capillary endothelial cells.

    PubMed

    Gelfand, R A; Frank, H J; Levin, E; Pedram, A

    1991-08-01

    The recent discovery of brain natriuretic peptides (BNP) that stimulates natriuresis, diuresis, and vascular smooth muscle relaxation in a manner similar to that of atrial natriuretic peptide (ANP) suggests the possibility that these endocrine hormones function via some common mechanism. Indirect evidence from several laboratories suggests that BNP and ANP may bind to the same receptors. We examined whether ANP and BNP bind to a common set of receptors in cultured bovine brain capillary endothelial cells and in bovine aortic endothelial cells. Scatchard plot analysis of binding data shows a similar dissociation constant (KD) of approximately 0.3 nM and a maximal binding capacity (Bmax) of 50 fmol/mg protein for both natriuretic peptides in brain capillary cells and 0.6 nM and 80 fmol/mg protein, respectively, in the aortic endothelial cells. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the affinity cross-linked receptor-ligand complex shows a strongly labeled 65-kDa receptor and a 125-kDa band that is likely to be a receptor of the guanylate cyclase type. ANP and BNP cross compete equally for binding to the two receptors identified on the gels. ANP and BNP also stimulate guanosine 3', 5'-cyclic monophosphate production in these cells, consistent with the presence of a functional guanylate cyclase-linked B receptor. We conclude that ANP and BNP share common receptors in brain capillary and aortic endothelial cells.

  18. Ischaemia-related cell damage in extracorporeal preserved tissue - new findings with a novel perfusion model.

    PubMed

    Taeger, Christian D; Müller-Seubert, Wibke; Horch, Raymund E; Präbst, Konstantin; Münch, Frank; Geppert, Carol I; Birkholz, Torsten; Dragu, Adrian

    2014-05-01

    Tissue undergoing free transfer in transplant or reconstructive surgery always is at high risk of ischaemia-related cell damage. This study aims at assessing different procedures using an extracorporeal perfusion and oxygenation system to investigate the expression of hypoxia inducible factor (HIF)-1-α as marker for hypoxia and of the pro-apoptotic protein Caspase-3 in skeletal muscle to elucidate potential improvements in tissue conservation. Twenty-four porcine rectus abdominis muscles were assigned to five different groups and examined after they had been extracorporeally preserved for 60 min. time. Group I was left untreated (control), group II was perfused with a cardioplegic solution, group III was flushed with 10 ml of a cardioplegic solution and then left untreated. Group IV and V were perfused and oxygenated with either an isotone crystalloid solution or a cardioplegic solution. Among others, immunohistochemistry (Caspase-3 and HIF-1-α) of muscle samples was performed. Furthermore, oxygen partial pressure in the perfusate at the arterial and venous branch was measured. Expression of Caspase-3 after 60 min. was reduced in all groups compared to the control group. Furthermore, all groups (except group III) expressed less HIF-1-α than the control group. Oxygenation leads to higher oxygen levels at the venous branch compared to groups without oxygenation. Using an extracorporeal perfusion and oxygenation system cell damage could be reduced as indicated by stabilized expressions of Caspase-3 and HIF-1-α for 60 min. of tissue preservation. Complete depletion of oxygen at the venous branch can be prevented by oxygenation of the perfusate with ambient air.

  19. Mesenchymal Stem Cells in the Treatment of Traumatic Brain Injury

    PubMed Central

    Hasan, Anwarul; Deeb, George; Rahal, Rahaf; Atwi, Khairallah; Mondello, Stefania; Marei, Hany Elsayed; Gali, Amr; Sleiman, Eliana

    2017-01-01

    Traumatic brain injury (TBI) is characterized by a disruption in the normal function of the brain due to an injury following a trauma, which can potentially cause severe physical, cognitive, and emotional impairment. The primary insult to the brain initiates secondary injury cascades consisting of multiple complex biochemical responses of the brain that significantly influence the overall severity of the brain damage and clinical sequelae. The use of mesenchymal stem cells (MSCs) offers huge potential for application in the treatment of TBI. MSCs have immunosuppressive properties that reduce inflammation in injured tissue. As such, they could be used to modulate the secondary mechanisms of injury and halt the progression of the secondary insult in the brain after injury. Particularly, MSCs are capable of secreting growth factors that facilitate the regrowth of neurons in the brain. The relative abundance of harvest sources of MSCs also makes them particularly appealing. Recently, numerous studies have investigated the effects of infusion of MSCs into animal models of TBI. The results have shown significant improvement in the motor function of the damaged brain tissues. In this review, we summarize the recent advances in the application of MSCs in the treatment of TBI. The review starts with a brief introduction of the pathophysiology of TBI, followed by the biology of MSCs, and the application of MSCs in TBI treatment. The challenges associated with the application of MSCs in the treatment of TBI and strategies to address those challenges in the future have also been discussed. PMID:28265255

  20. Mesenchymal Stem Cells in the Treatment of Traumatic Brain Injury.

    PubMed

    Hasan, Anwarul; Deeb, George; Rahal, Rahaf; Atwi, Khairallah; Mondello, Stefania; Marei, Hany Elsayed; Gali, Amr; Sleiman, Eliana

    2017-01-01

    Traumatic brain injury (TBI) is characterized by a disruption in the normal function of the brain due to an injury following a trauma, which can potentially cause severe physical, cognitive, and emotional impairment. The primary insult to the brain initiates secondary injury cascades consisting of multiple complex biochemical responses of the brain that significantly influence the overall severity of the brain damage and clinical sequelae. The use of mesenchymal stem cells (MSCs) offers huge potential for application in the treatment of TBI. MSCs have immunosuppressive properties that reduce inflammation in injured tissue. As such, they could be used to modulate the secondary mechanisms of injury and halt the progression of the secondary insult in the brain after injury. Particularly, MSCs are capable of secreting growth factors that facilitate the regrowth of neurons in the brain. The relative abundance of harvest sources of MSCs also makes them particularly appealing. Recently, numerous studies have investigated the effects of infusion of MSCs into animal models of TBI. The results have shown significant improvement in the motor function of the damaged brain tissues. In this review, we summarize the recent advances in the application of MSCs in the treatment of TBI. The review starts with a brief introduction of the pathophysiology of TBI, followed by the biology of MSCs, and the application of MSCs in TBI treatment. The challenges associated with the application of MSCs in the treatment of TBI and strategies to address those challenges in the future have also been discussed.

  1. Silica bioreplication preserves three-dimensional spheroid structures of human pluripotent stem cells and HepG2 cells

    PubMed Central

    Lou, Yan-Ru; Kanninen, Liisa; Kaehr, Bryan; Townson, Jason L.; Niklander, Johanna; Harjumäki, Riina; Jeffrey Brinker, C.; Yliperttula, Marjo

    2015-01-01

    Three-dimensional (3D) cell cultures produce more in vivo-like multicellular structures such as spheroids that cannot be obtained in two-dimensional (2D) cell cultures. Thus, they are increasingly employed as models for cancer and drug research, as well as tissue engineering. It has proven challenging to stabilize spheroid architectures for detailed morphological examination. Here we overcome this issue using a silica bioreplication (SBR) process employed on spheroids formed from human pluripotent stem cells (hPSCs) and hepatocellular carcinoma HepG2 cells cultured in the nanofibrillar cellulose (NFC) hydrogel. The cells in the spheroids are more round and tightly interacting with each other than those in 2D cultures, and they develop microvilli-like structures on the cell membranes as seen in 2D cultures. Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins. These findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures. PMID:26323570

  2. Silica bioreplication preserves three-dimensional spheroid structures of human pluripotent stem cells and HepG2 cells

    SciTech Connect

    Lou, Yan-Ru; Kanninen, Liisa; Kaehr, Bryan; Townson, Jason L.; Niklander, Johanna; Harjumäki, Riina; Jeffrey Brinker, C.; Yliperttula, Marjo

    2015-09-01

    Three-dimensional (3D) cell cultures produce more in vivo-like multicellular structures such as spheroids that cannot be obtained in two-dimensional (2D) cell cultures. Thus, they are increasingly employed as models for cancer and drug research, as well as tissue engineering. It has proven challenging to stabilize spheroid architectures for detailed morphological examination. Here we overcome this issue using a silica bioreplication (SBR) process employed on spheroids formed from human pluripotent stem cells (hPSCs) and hepatocellular carcinoma HepG2 cells cultured in the nanofibrillar cellulose (NFC) hydrogel. The cells in the spheroids are more round and tightly interacting with each other than those in 2D cultures, and they develop microvilli-like structures on the cell membranes as seen in 2D cultures. Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins. In conclusion, these findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.

  3. Silica bioreplication preserves three-dimensional spheroid structures of human pluripotent stem cells and HepG2 cells

    DOE PAGES

    Lou, Yan-Ru; Kanninen, Liisa; Kaehr, Bryan; ...

    2015-09-01

    Three-dimensional (3D) cell cultures produce more in vivo-like multicellular structures such as spheroids that cannot be obtained in two-dimensional (2D) cell cultures. Thus, they are increasingly employed as models for cancer and drug research, as well as tissue engineering. It has proven challenging to stabilize spheroid architectures for detailed morphological examination. Here we overcome this issue using a silica bioreplication (SBR) process employed on spheroids formed from human pluripotent stem cells (hPSCs) and hepatocellular carcinoma HepG2 cells cultured in the nanofibrillar cellulose (NFC) hydrogel. The cells in the spheroids are more round and tightly interacting with each other than thosemore » in 2D cultures, and they develop microvilli-like structures on the cell membranes as seen in 2D cultures. Furthermore, SBR preserves extracellular matrix-like materials and cellular proteins. In conclusion, these findings provide the first evidence of intact hPSC spheroid architectures and similar fine structures to 2D-cultured cells, providing a pathway to enable our understanding of morphogenesis in 3D cultures.« less

  4. Expression of arginine decarboxylase in brain regions and neuronal cells

    PubMed Central

    Iyo, Abiye H.; Zhu, Meng-Yang; Ordway, Gregory A.; Regunathan, Soundar

    2010-01-01

    After our initial report of a mammalian gene for arginine decarboxylase, an enzyme for the synthesis of agmatine from arginine, we have determined the regional expression of ADC in rat. We have analyzed the expression of ADC in rat brain regions by activity, protein and mRNA levels, and the regulation of expression in neuronal cells by RNA interference. In rat brain, ADC was widely expressed in major brain regions, with a substantial amount in hypothalamus, followed by cortex, and with least amounts in locus coeruleus and medulla. ADC mRNA was detected in primary astrocytes and C6 glioma cells. While no ADC message was detected in fresh neurons (3 days old), significant message appeared in differentiated neurons (3 weeks old). PC12 cells, treated with nerve growth factor, had higher ADC mRNA compared with naive cells. The siRNA mixture directed towards the N-terminal regions of ADC cDNA down-regulated the levels of mRNA and protein in cultured neurons/C6 glioma cells and these cells produced lower agmatine. Thus, this study demonstrates that ADC message is expressed in rat brain regions, that it is regulated in neuronal cells and that the down-regulation of ADC activity by specific siRNA leads to lower agmatine production. PMID:16445852

  5. Fluorescence-activated sorting of fixed nuclei: a general method for studying nuclei from specific cell populations that preserves post-translational modifications.

    PubMed

    Marion-Poll, Lucile; Montalban, Enrica; Munier, Annie; Hervé, Denis; Girault, Jean-Antoine

    2014-04-01

    Long-lasting brain alterations that underlie learning and memory are triggered by synaptic activity. How activity can exert long-lasting effects on neurons is a major question in neuroscience. Signalling pathways from cytoplasm to nucleus and the resulting changes in transcription and epigenetic modifications are particularly relevant in this context. However, a major difficulty in their study comes from the cellular heterogeneity of brain tissue. A promising approach is to directly purify identified nuclei. Using mouse striatum we have developed a rapid and efficient method for isolating cell type-specific nuclei from fixed adult brain (fluorescence-activated sorting of fixed nuclei; FAST-FIN). Animals are quickly perfused with a formaldehyde fixative that stops enzymatic reactions and maintains the tissue in the state it was at the time of death, including nuclear localisation of soluble proteins such as GFP and differences in nuclear size between cell types. Tissue is subsequently dissociated with a Dounce homogeniser and nuclei prepared by centrifugation in an iodixanol density gradient. The purified fixed nuclei can then be immunostained with specific antibodies and analysed or sorted by flow cytometry. Simple criteria allow distinction of neurons and non-neuronal cells. Immunolabelling and transgenic mice that express fluorescent proteins can be used to identify specific cell populations, and the nuclei from these populations can be efficiently isolated, even rare cell types such as parvalbumin-expressing interneurons. FAST-FIN allows the preservation and study of dynamic and labile post-translational protein modifications. It should be applicable to other tissues and species, and allow study of DNA and its modifications.

  6. Preservation of perisomatic inhibitory input of granule cells in the epileptic human dentate gyrus.

    PubMed

    Wittner, L; Maglóczky, Z; Borhegyi, Z; Halász, P; Tóth, S; Eross, L; Szabó, Z; Freund, T F

    2001-01-01

    Temporal lobe epilepsy is known to be associated with hyperactivity that is likely to be generated or amplified in the hippocampal formation. The majority of granule cells, the principal cells of the dentate gyrus, are found to be resistant to damage in epilepsy, and may serve as generators of seizures if their inhibition is impaired. Therefore, the parvalbumin-containing subset of interneurons, known to provide the most powerful inhibitory input to granule cell somata and axon initial segments, were examined in human control and epileptic dentate gyrus. A strong reduction in the number of parvalbumin-containing cells was found in the epileptic samples especially in the hilar region, although in some patches of the granule cell layer parvalbumin-positive terminals that form vertical clusters characteristic of axo-axonic cells were more numerous than in controls. Analysis of the postsynaptic target elements of parvalbumin-positive axon terminals showed that they form symmetric synapses with somata, dendrites, axon initial segments and spines as in the control, but the ratio of axon initial segment synapses was increased in the epileptic tissue (control: 15.9%, epileptic: 31.3%). Furthermore, the synaptic coverage of granule cell axon initial segments increased more than three times (control: 0.52, epileptic: 2.10 microm synaptic length/100 microm axon initial segment membrane) in the epileptic samples, whereas the amount of somatic symmetric synapses did not change significantly. Although the number of parvalbumin-positive interneurons is decreased, the perisomatic inhibitory input of dentate granule cells is preserved in temporal lobe epilepsy. Basket and axo-axonic cell terminals - whether positive or negative for parvalbumin - are present, moreover, the axon collaterals targeting axon initial segments sprout in the epileptic dentate gyrus. We suggest that perisomatic inhibitory interneurons survive in epilepsy, but their somadendritic compartment and partly the

  7. Impulsive pressurization of neuronal cells for traumatic brain injury study.

    PubMed

    Nienaber, Matthew; Lee, Jeong Soon; Feng, Ruqiang; Lim, Jung Yul

    2011-10-12

    A novel impulsive cell pressurization experiment has been developed using a Kolsky bar device to investigate blast-induced traumatic brain injury (TBI). We demonstrate in this video article how blast TBI-relevant impulsive pressurization is applied to the neuronal cells in vitro. This is achieved by using well-controlled pressure pulse created by a specialized Kolsky bar device, with complete pressure history within the cell pressurization chamber recorded. Pressurized neuronal cells are inspected immediately after pressurization, or further incubated to examine the long-term effects of impulsive pressurization on neurite/axonal outgrowth, neuronal gene expression, apoptosis, etc. We observed that impulsive pressurization at about 2 MPa induces distinct neurite loss relative to unpressurized cells. Our technique provides a novel method to investigate the molecular/cellular mechanisms of blast TBI, via impulsive pressurization of brain cells at well-controlled pressure magnitude and duration.

  8. Thyroid hormones are essential to preserve non-proliferative cells of adult neurogenesis of the dentate gyrus.

    PubMed

    Sánchez-Huerta, K; García-Martínez, Y; Vergara, P; Segovia, J; Pacheco-Rosado, J

    2016-10-01

    Thyroid hormones (THs) regulate adult hippocampal neurogenesis, a process that involves both cell populations that dynamically switch between pools of proliferative and quiescent cells, and cells that definitely leave the cell cycle to maturate into granular neurons. This investigation was carried out to determine the role of THs on the mitotic activity of specific proliferative cell populations and the preservation of non-proliferative cells participating in the neurogenic process of the dentate gyrus (DG) of the hippocampus. Hypothyroidism was induced in male adult Wistar rats with methimazole for 28days. We quantified the total number of proliferative cells (BrdU+), proliferative type 1 (BrdU+/GFAP+), and 2b and 3 (BrdU+/DCX+) cells. Early non-proliferative cells (BrdU-/DCX+ cells lacking dendritic process), postmitotic neuroblasts (Tuj 1+ cells lacking dendritic process), and immature granular neurons (IGN; DCX+ or Tuj 1+ and the presence of dendritic processes into granular or molecular layer) were also included. The evidence showed that the proliferation of Type 1, 2b and 3 cells is not modified by hypothyroidism. In contrast, hypothyroidism reduced the number of early non-proliferative cells and also leads to a decrement in the number of IGN. Our results show that proliferative cells of the DG are not sensitive to thyroid perturbations. However, THs are essential to preserve cell populations that leave the cell cycle in the DG of the hippocampus.

  9. Connecting Malfunctioning Glial Cells and Brain Degenerative Disorders.

    PubMed

    Kaminsky, Natalie; Bihari, Ofer; Kanner, Sivan; Barzilai, Ari

    2016-06-01

    The DNA damage response (DDR) is a complex biological system activated by different types of DNA damage. Mutations in certain components of the DDR machinery can lead to genomic instability disorders that culminate in tissue degeneration, premature aging, and various types of cancers. Intriguingly, malfunctioning DDR plays a role in the etiology of late onset brain degenerative disorders such as Parkinson's, Alzheimer's, and Huntington's diseases. For many years, brain degenerative disorders were thought to result from aberrant neural death. Here we discuss the evidence that supports our novel hypothesis that brain degenerative diseases involve dysfunction of glial cells (astrocytes, microglia, and oligodendrocytes). Impairment in the functionality of glial cells results in pathological neuro-glial interactions that, in turn, generate a "hostile" environment that impairs the functionality of neuronal cells. These events can lead to systematic neural demise on a scale that appears to be proportional to the severity of the neurological deficit.

  10. Neural stem cell-based gene therapy for brain tumors.

    PubMed

    Kim, Seung U

    2011-03-01

    Advances in gene-based medicine since 1990s have ushered in new therapeutic strategy of gene therapy for inborn error genetic diseases and cancer. Malignant brain tumors such as glioblastoma multiforme and medulloblastoma remain virtually untreatable and lethal. Currently available treatment for brain tumors including radical surgical resection followed by radiation and chemotherapy, have substantially improved the survival rate in patients suffering from these brain tumors; however, it remains incurable in large proportion of patients. Therefore, there is substantial need for effective, low-toxicity therapies for patients with malignant brain tumors, and gene therapy targeting brain tumors should fulfill this requirement. Gene therapy for brain tumors includes many therapeutic strategies and these strategies can be grouped in two major categories: molecular and immunologic. The widely used molecular gene therapy approach is suicide gene therapy based on the conversion of non-toxic prodrugs into active anticancer agents via introduction of enzymes and genetic immunotherapy involves the gene transfer of immune-stimulating cytokines including IL-4, IL-12 and TRAIL. For both molecular and immune gene therapy, neural stem cells (NSCs) can be used as delivery vehicle of therapeutic genes. NSCs possess an inherent tumor tropism that supports their use as a reliable delivery vehicle to target therapeutic gene products to primary brain tumors and metastatic cancers throughout the brain. Significance of the NSC-based gene therapy for brain tumor is that it is possible to exploit the tumor-tropic property of NSCs to mediate effective, tumor-selective therapy for primary and metastatic cancers in the brain and outside, for which no tolerated curative treatments are currently available.

  11. Endotoxin-induced lung alveolar cell injury causes brain cell damage

    PubMed Central

    Rodríguez-González, Raquel; Ramos-Nuez, Ángela; Martín-Barrasa, José Luis; López-Aguilar, Josefina; Baluja, Aurora; Álvarez, Julián; Rocco, Patricia RM; Pelosi, Paolo

    2015-01-01

    Sepsis is the most common cause of acute respiratory distress syndrome, a severe lung inflammatory disorder with an elevated morbidity and mortality. Sepsis and acute respiratory distress syndrome involve the release of inflammatory mediators to the systemic circulation, propagating the cellular and molecular response and affecting distal organs, including the brain. Since it has been reported that sepsis and acute respiratory distress syndrome contribute to brain dysfunction, we investigated the brain-lung crosstalk using a combined experimental in vitro airway epithelial and brain cell injury model. Conditioned medium collected from an in vitro lipopolysaccharide-induced airway epithelial cell injury model using human A549 alveolar cells was subsequently added at increasing concentrations (no conditioned, 2%, 5%, 10%, 15%, 25%, and 50%) to a rat mixed brain cell culture containing both astrocytes and neurons. Samples from culture media and cells from mixed brain cultures were collected before treatment, and at 6 and 24 h for analysis. Conditioned medium at 15% significantly increased apoptosis in brain cell cultures 24 h after treatment, whereas 25% and 50% significantly increased both necrosis and apoptosis. Levels of brain damage markers S100 calcium binding protein B and neuron-specific enolase, interleukin-6, macrophage inflammatory protein-2, as well as matrix metalloproteinase-9 increased significantly after treating brain cells with ≥2% conditioned medium. Our findings demonstrated that human epithelial pulmonary cells stimulated with bacterial lipopolysaccharide release inflammatory mediators that are able to induce a translational clinically relevant and harmful response in brain cells. These results support a brain-lung crosstalk during sepsis and sepsis-induced acute respiratory distress syndrome. PMID:25135986

  12. The Effect of Underwater Blast on Aggregating Brain Cell Cultures.

    PubMed

    Sawyer, Thomas W; Lee, Julian J; Villanueva, Mercy; Wang, Yushan; Nelson, Peggy; Song, Yanfeng; Fan, Chengyang; Barnes, Julia; McLaws, Lori

    2017-01-15

    Although the deleterious effects of primary blast on gas-filled organs are well accepted, the effect of blast-induced shock waves on the brain is less clear because of factors that complicate the interpretation of clinical and experimental data. Brain cell aggregate cultures are comprised of multiple differentiated brain cell types and were used to examine the effects of underwater blast. Suspensions of these cultures encased in dialysis tubing were exposed to explosive-generated underwater blasts of low (∼300 kPa), medium (∼2,700 kPa), or high (∼14,000 kPa) intensities and harvested at 1-28 days post-exposure. No changes in gross morphology were noted immediately or weeks after blast wave exposure, and no increases in either apoptotic (caspase-3) or necrotic (lactate dehydrogenase) cell death were observed. Changes in neuronal (neurofilament H, acetylcholinesterase, and choline acetyltransferase) and glial (glial fibrillary acidic protein, glutamine synthetase) endpoints did not occur. However, significant time- and pressure-related increases in Akt (protein kinase B) phosphorylation were noted, as well as declines in vascular endothelial growth factor levels, implicating pathways involved in cellular survival mechanisms. The free-floating nature of the aggregates during blast wave exposure, coupled with their highly hydrolyzed dialysis tubing containment, results in minimized boundary effects, thus enabling accurate assessment of brain cell response to a simplified shock-induced stress wave. This work shows that, at its simplest, blast-induced shock waves produce subtle changes in brain tissue. This study has mechanistic implications for the study of primary blast-induced traumatic brain injury and supports the thesis that underwater blast may cause subtle changes in the brains of submerged individuals.

  13. Radial glia cells in the developing human brain.

    PubMed

    Howard, Brian M; Zhicheng Mo; Filipovic, Radmila; Moore, Anna R; Antic, Srdjan D; Zecevic, Nada

    2008-10-01

    Human radial glia (RG) share many of the features described in rodents, but also have a number of characteristics unique to the human brain. Results obtained from different mammalian species including human and non-human primates reveal differences in the involvement of RG in neurogenesis and oligodendrogenesis and in the timing of the initial expression of typical RG immunomarkers. A common problem in studying the human brain is that experimental procedures using modern molecular and genetic methods, such as in vivo transduction with retroviruses or creation of knockout or transgenic mutants, are not possible. Nevertheless, abundant and valuable information about the development of the human brain has been revealed using postmortem human material. Additionally, a combination and spectrum of in vitro techniques are used to gain knowledge about normal developmental processes in the human brain, including better understanding of RG as progenitor cells. Molecular and functional characterization of multipotent progenitors, such as RG, is important for future cell replacement therapies in neurological and psychiatric disorders, which are often resistant to conventional treatments. The protracted time of development and larger size of the human brain could provide insight into processes that may go unnoticed in the much smaller rodent cortex, which develops over a much shorter period. With that in mind, we summarize results on the role of RG in the human fetal brain.

  14. Nuclear magnetic resonance studies of biological systems: Applications to liver preservation and metabolism in cultured pituitary tumor cells

    SciTech Connect

    Fralix, T.A.

    1989-01-01

    This study centers on applications of both {sup 31}P and {sup 13}C nuclear magnetic resonance spectroscopy to two different biological systems. The first application utilizes {sup 31}P NMR to study mobile phospholipids in the MMQ cell line, a pituitary tumor cell line. These measurements characterize membrane phospholipids thought to be part of a RNA-proteolipid complex unique to cellular transformation. The second application utilizes both {sup 31}P and {sup 13}C spectroscopy to study liver preservation and transplantation an a rat model. In this work, several questions were addressed: (1) to what extent do successful preservation solutions slow ATP breakdown (2) can clinically successful preservation conditions ameliorate total nucleotide breakdown (3) to what extent is energy reconstitution following cold storage correlated with transport success and (4) can any spectroscopic parameter be used as a diagnostic indicator of tissue viability

  15. Brain mesenchymal stem cells: physiology and pathological implications.

    PubMed

    Pombero, Ana; Garcia-Lopez, Raquel; Martinez, Salvador

    2016-06-01

    Mesenchymal stem cells (MSCs) are defined as progenitor cells that give rise to a number of unique, differentiated mesenchymal cell types. This concept has progressively evolved towards an all-encompassing concept including multipotent perivascular cells of almost any tissue. In central nervous system, pericytes are involved in blood-brain barrier, and angiogenesis and vascular tone regulation. They form the neurovascular unit (NVU) together with endothelial cells, astrocytes and neurons. This functional structure provides an optimal microenvironment for neural proliferation in the adult brain. Neurovascular niche include both diffusible signals and direct contact with endothelial and pericytes, which are a source of diffusible neurotrophic signals that affect neural precursors. Therefore, MSCs/pericyte properties such as differentiation capability, as well as immunoregulatory and paracrine effects make them a potential resource in regenerative medicine.

  16. Nitric oxide consumption through lipid peroxidation in brain cell suspensions and homogenates.

    PubMed

    Keynes, Robert G; Griffiths, Charmaine H; Hall, Catherine; Garthwaite, John

    2005-05-01

    Mechanisms which inactivate NO (nitric oxide) are probably important in governing the physiological and pathological effects of this ubiquitous signalling molecule. Cells isolated from the cerebellum, a brain region rich in the NO signalling pathway, consume NO avidly. This property was preserved in brain homogenates and required both particulate and supernatant fractions. A purified fraction of the particulate component was rich in phospholipids, and NO consumption was inhibited by procedures that inhibited lipid peroxidation, namely a transition metal chelator, the vitamin E analogue Trolox and ascorbate oxidase. The requirement for the supernatant was accounted for by its content of ascorbate which catalyses metal-dependent lipid peroxidation. The NO-degrading activity of the homogenate was mimicked by a representative mixture of brain lipids together with ascorbate and, under these conditions, the lipids underwent peroxidation. In a suspension of cerebellar cells, there was a continuous low level of lipid peroxidation, and consumption of NO by the cells was decreased by approx. 50% by lipid-peroxidation inhibitors. Lipid peroxidation was also abolished when NO was supplied at a continuously low rate (approximately 100 nM/min), which explains why NO consumption by this process is saturable. Part of the activity remaining after the inhibition of lipid peroxidation was accounted for by contaminating red blood cells, but there was also another component whose activity was greatly enhanced when the cells were maintained under air-equilibrated conditions. A similar NO-consuming process was present in cerebellar glial cells grown in tissue culture but not in blood platelets or leucocytes, suggesting a specialized mechanism.

  17. SGLT2 Deletion Improves Glucose Homeostasis and Preserves Pancreatic β-Cell Function

    PubMed Central

    Jurczak, Michael J.; Lee, Hui-Young; Birkenfeld, Andreas L.; Jornayvaz, Francois R.; Frederick, David W.; Pongratz, Rebecca L.; Zhao, Xiaoxian; Moeckel, Gilbert W.; Samuel, Varman T.; Whaley, Jean M.; Shulman, Gerald I.; Kibbey, Richard G.

    2011-01-01

    OBJECTIVE Inhibition of the Na+-glucose cotransporter type 2 (SGLT2) is currently being pursued as an insulin-independent treatment for diabetes; however, the behavioral and metabolic consequences of SGLT2 deletion are unknown. Here, we used a SGLT2 knockout mouse to investigate the effect of increased renal glucose excretion on glucose homeostasis, insulin sensitivity, and pancreatic β-cell function. RESEARCH DESIGN AND METHODS SGLT2 knockout mice were fed regular chow or a high-fat diet (HFD) for 4 weeks, or backcrossed onto the db/db background. The analysis used metabolic cages, glucose tolerance tests, euglycemic and hyperglycemic clamps, as well as isolated islet and perifusion studies. RESULTS SGLT2 deletion resulted in a threefold increase in urine output and a 500-fold increase in glucosuria, as well as compensatory increases in feeding, drinking, and activity. SGLT2 knockout mice were protected from HFD-induced hyperglycemia and glucose intolerance and had reduced plasma insulin concentrations compared with controls. On the db/db background, SGLT2 deletion prevented fasting hyperglycemia, and plasma insulin levels were also dramatically improved. Strikingly, prevention of hyperglycemia by SGLT2 knockout in db/db mice preserved pancreatic β-cell function in vivo, which was associated with a 60% increase in β-cell mass and reduced incidence of β-cell death. CONCLUSIONS Prevention of renal glucose reabsorption by SGLT2 deletion reduced HFD- and obesity-associated hyperglycemia, improved glucose intolerance, and increased glucose-stimulated insulin secretion in vivo. Taken together, these data support SGLT2 inhibition as a viable insulin-independent treatment of type 2 diabetes. PMID:21357472

  18. Environmentally Friendly Carbon-Preserving Recovery of Noble Metals From Supported Fuel Cell Catalysts.

    PubMed

    Latsuzbaia, R; Negro, E; Koper, G J M

    2015-06-08

    The dissolution of noble-metal catalysts under mild and carbon-preserving conditions offers the possibility of in situ regeneration of the catalyst nanoparticles in fuel cells or other applications. Here, we report on the complete dissolution of the fuel cell catalyst, platinum nanoparticles, under very mild conditions at room temperature in 0.1 M HClO4 and 0.1 M HCl by electrochemical potential cycling between 0.5-1.1 V at a scan rate of 50 mV s(-1) . Dissolution rates as high as 22.5 μg cm(-2) per cycle were achieved, which ensured a relatively short dissolution timescale of 3-5 h for a Pt loading of 0.35 mg cm(-2) on carbon. The influence of chloride ions and oxygen in the electrolyte on the dissolution was investigated, and a dissolution mechanism is proposed on the basis of the experimental observations and available literature results. During the dissolution process, the corrosion of the carbon support was minimal, as observed by X-ray photoelectron spectroscopy (XPS).

  19. Transport systems of serine at the brain barriers and in brain parenchymal cells.

    PubMed

    Kasai, Yasuyuki; Tachikawa, Masanori; Hirose, Shirou; Akanuma, Shin-ichi; Hosoya, Ken-ichi

    2011-07-01

    D-Serine is a co-agonist for NMDA-type glutamate receptors. Although D-serine levels in CSF and interstitial fluid (ISF) affect CNS function, the regulatory system remains to be fully understood. Therefore, the purpose of this study was to investigate d-serine transport across the blood-brain barrier (BBB) and blood-CSF barrier (BCSFB) and in brain parenchymal cells. D-Serine microinjected into the cerebrum was not eliminated, suggesting a negligible contribution of D-serine efflux transport at the BBB. In contrast, D-serine was taken up from the circulating blood across the BBB via a carrier-mediated process. D-Serine elimination clearance from CSF was fourfold greater than that of d-mannitol, which is considered to reflect CSF bulk flow. The characteristics of D-serine uptake by isolated choroid plexus were consistent with those of Na(+)-independent alanine-serine-cysteine transporter 1 (asc-1). Uptake of D-serine by brain slices appeared to occur predominantly via asc-1 and Na(+)-dependent alanine-serine-cysteine transporter 2. These findings suggest that the regulatory system of D-serine levels in ISF and CSF involves (i) asc-1 at the BCSFB, acting as a major pathway of D-serine elimination from the CSF, (ii) blood-to-brain and blood-to-CSF influx transport of D-serine across the BBB and BCSFB, and (iii) concentrative uptake of D-serine by brain parenchymal cells.

  20. Spatial organization and correlations of cell nuclei in brain tumors.

    PubMed

    Jiao, Yang; Berman, Hal; Kiehl, Tim-Rasmus; Torquato, Salvatore

    2011-01-01

    Accepting the hypothesis that cancers are self-organizing, opportunistic systems, it is crucial to understand the collective behavior of cancer cells in their tumorous heterogeneous environment. In the present paper, we ask the following basic question: Is this self-organization of tumor evolution reflected in the manner in which malignant cells are spatially distributed in their heterogeneous environment? We employ a variety of nontrivial statistical microstructural descriptors that arise in the theory of heterogeneous media to characterize the spatial distributions of the nuclei of both benign brain white matter cells and brain glioma cells as obtained from histological images. These descriptors, which include the pair correlation function, structure factor and various nearest neighbor functions, quantify how pairs of cell nuclei are correlated in space in various ways. We map the centroids of the cell nuclei into point distributions to show that while commonly used local spatial statistics (e.g., cell areas and number of neighboring cells) cannot clearly distinguish spatial correlations in distributions of normal and abnormal cell nuclei, their salient structural features are captured very well by the aforementioned microstructural descriptors. We show that the tumorous cells pack more densely than normal cells and exhibit stronger effective repulsions between any pair of cells. Moreover, we demonstrate that brain gliomas are organized in a collective way rather than randomly on intermediate and large length scales. The existence of nontrivial spatial correlations between the abnormal cells strongly supports the view that cancer is not an unorganized collection of malignant cells but rather a complex emergent integrated system.

  1. Taurine treatment preserves brain and liver mitochondrial function in a rat model of fulminant hepatic failure and hyperammonemia.

    PubMed

    Jamshidzadeh, Akram; Heidari, Reza; Abasvali, Mozhgan; Zarei, Mehdi; Ommati, Mohammad Mehdi; Abdoli, Narges; Khodaei, Forouzan; Yeganeh, Yasaman; Jafari, Faezeh; Zarei, Azita; Latifpour, Zahra; Mardani, Elnaz; Azarpira, Negar; Asadi, Behnam; Najibi, Asma

    2017-02-01

    Ammonia-induced mitochondrial dysfunction and energy crisis is known as a critical consequence of hepatic encephalopathy (HE). Hence, mitochondria are potential targets of therapy in HE. The current investigation was designed to evaluate the role of taurine treatment on the brain and liver mitochondrial function in a rat model of hepatic encephalopathy and hyperammonemia. The animals received thioacetamide (400mg/kg, i.p, for three consecutive days at 24-h intervals) as a model of acute liver failure and hyperammonemia. Several biochemical parameters were investigated in the serum, while the animals' cognitive function and locomotor activity were monitored. Mitochondria was isolated from the rats' brain and liver and several indices were assessed in isolated mitochondria. Liver failure led to cognitive dysfunction and impairment in locomotor activity in the rats. Plasma and brain ammonia was high and serum markers of liver injury were drastically elevated in the thioacetamide-treated group. An assessment of brain and liver mitochondrial function in the thioacetamide-treated animals revealed an inhibition of succinate dehydrogenase activity (SDA), collapsed mitochondrial membrane potential, mitochondrial swelling, and increased reactive oxygen species (ROS). Furthermore, a significant decrease in mitochondrial ATP was detected in the brain and liver mitochondria isolated from thioacetamide-treated animals. Taurine treatment (250, 500, and 1000mg/kg) decreased mitochondrial swelling, ROS, and LPO. Moreover, the administration of this amino acid restored brain and liver mitochondrial ATP. These data suggest taurine to be a potential protective agent with therapeutic capability against hepatic encephalopathy and hyperammonemia-induced mitochondrial dysfunction and energy crisis.

  2. Growth hormone (GH), brain development and neural stem cells.

    PubMed

    Waters, M J; Blackmore, D G

    2011-12-01

    A range of observations support a role for GH in development and function of the brain. These include altered brain structure in GH receptor null mice, and impaired cognition in GH deficient rodents and in a subgroup of GH receptor defective patients (Laron dwarfs). GH has been shown to alter neurogenesis, myelin synthesis and dendritic branching, and both the GH receptor and GH itself are expressed widely in the brain. We have found a population of neural stem cells which are activated by GH infusion, and which give rise to neurons in mice. These stem cells are activated by voluntary exercise in a GH-dependent manner. Given the findings that local synthesis of GH occurs in the hippocampus in response to a memory task, and that GH replacement improves memory and cognition in rodents and humans, these new observations warrant a reappraisal of the clinical importance of GH replacement in GH deficient states.

  3. Culture and Isolation of Brain Tumor Initiating Cells.

    PubMed

    Vora, Parvez; Venugopal, Chitra; McFarlane, Nicole; Singh, Sheila K

    2015-08-03

    Brain tumors are typically composed of heterogeneous cells that exhibit distinct phenotypic characteristics and proliferative potentials. Only a relatively small fraction of cells in the tumor with stem cell properties, termed brain tumor initiating cells (BTICs), possess an ability to differentiate along multiple lineages, self-renew, and initiate tumors in vivo. This unit describes protocols for the culture and isolation BTICs. We applied culture conditions and assays originally used for normal neural stem cells (NSCs) in vitro to a variety of brain tumors. Using fluorescence-activated cell sorting for the neural precursor cell surface marker CD133/CD15, BTICs can be isolated and studied prospectively. Isolation of BTICs from GBM bulk tumor will enable examination of dissimilar morphologies, self-renewal capacities, tumorigenicity, and therapeutic sensitivities. As cancer is also considered a disease of unregulated self-renewal and differentiation, an understanding of BTICs is fundamental to understanding tumor growth. Ultimately, it will lead to novel drug discovery approaches that strategically target the functionally relevant BTIC population.

  4. Brain microvascular endothelium induced-annexin A1 secretion contributes to small cell lung cancer brain metastasis.

    PubMed

    Liu, Yi; Liu, Yong-Shuo; Wu, Peng-Fei; Li, Qiang; Dai, Wu-Min; Yuan, Shuai; Xu, Zhi-Hua; Liu, Ting-Ting; Miao, Zi-Wei; Fang, Wen-Gang; Chen, Yu-Hua; Li, Bo

    2015-09-01

    Small cell lung cancer is the most aggressive histologic subtype of lung cancer, with a strong predilection for metastasizing to brain early. However, the cellular and molecular basis is poorly known. Here, we provided evidence to reveal the role of annexin A1 in small cell lung cancer metastasis to brain. Firstly, the elevated annexin A1 serum levels in small cell lung cancer patients were associated with brain metastasis. The levels of annexin A1 were also upregulated in NCI-H446 cells, a small cell lung cancer cell line, upon migration into the mice brain. More interestingly, annexin A1 was secreted by NCI-H446 cells in a time-dependent manner when co-culturing with human brain microvascular endothelial cells, which was identified with the detections of annexin A1 in the co-cultured cellular supernatants by ELISA and western blot. Further results showed that blockage of annexin A1 in the co-cultured cellular supernatants using a neutralized antibody significantly inhibited NCI-H446 cells adhesion to brain endothelium and its transendothelial migration. Conversely, the addition of Ac2-26, an annexin A1 mimic peptide, enhanced these effects. Furthermore, knockdown of annexin A1 in NCI-H446 cells prevented its transendothelial migration in vitro and metastasis to mice brain in vivo. Our data showed that small cell lung cancer cell in brain microvasculature microenvironment could express much more annexin A1 and release it outside, which facilitated small cell lung cancer cell to gain malignant properties of entry into brain. These findings provided a potential target for the management of SCLC brain metastasis.

  5. A novel Gfer-Drp1 link in preserving mitochondrial dynamics and function in pluripotent stem cells.

    PubMed

    Todd, Lance R; Gomathinayagam, Rohini; Sankar, Uma

    2010-08-01

    Mitochondria, the dynamic energy powerhouses of the cell, have vital roles in a multitude of cellular processes including differentiation and cell survival. Tight regulation of mitochondrial dynamics, integrity and function is indispensible for preservation of homeostasis in all cells, including pluripotent stem cells. The ability to proliferate and self-renew indefinitely bestows the pluripotent embryonic stem cells (ESCs) with immense curative potential. Mechanisms that preserve mitochondrial well-being, and therefore maintain "stemness," are vital in realizing the full potential of ESCs in therapeutic regenerative medicine. However, virtually nothing is known regarding the regulation of mitochondrial dynamics and function and the relationship thereof to overall cell fate and function in pluripotent ESCs or other somatic stem cells. Using loss- and gain-of-function approaches, we show that growth factor erv1-like (Gfer) plays an essential pro-survival role in the maintenance of murine ESC pluripotency by preserving the structural and functional integrity of their mitochondria, through modulation of the key mitochondrial fission factor Drp1.

  6. Use of refrigeration as a practical means to preserve viability of in vitro-cultured IDE8 tick cells.

    PubMed

    Bastos, Camila V; das Vasconcelos, Maria Mercês C; Ribeiro, Múcio Flávio B; Passos, Lygia M Friche

    2006-01-01

    In vitro cultivation of the IDE8 cell line, derived from embryonic Ixodes scapularis ticks, constitutes an important system for the study of tick-borne pathogens, as these cells support growth of rickettsial species which are not normally transmitted by this tick. However, since cryopreservation of IDE8 cells is not always successful, there is a need to develop alternative ways to preserve these cells. In the present study, a suspension of IDE8 cells in culture medium was kept under refrigeration at 4 degrees C for up to 60 days. Every 15 days, the suspension was mixed and aliquots were re-cultured in 2-ml tubes, under standardized conditions. In addition, three techniques for cryopreservation, using two different cryoprotectants (DMSO and glycerol), were evaluated. Medium changes were carried out every week and subculturing every 2 weeks. The development of cultures and their respective subcultures, after returning to standard culture temperature, was evaluated by percentage viability and by cellular morphology evaluated in Giemsa-stained cytocentrifuge smears. All cultures and subcultures appeared healthy, showing growth rates comparable to cultures that had not been kept under refrigeration. The results demonstrated that storage under refrigeration at 4 degrees C is an efficient method for preservation of IDE8 cells for up to 60 days and that refrigeration may be preferable to cryopreservation for short-term preservation of IDE8 cells.

  7. Increased osmolarity and cell clustering preserve canine notochordal cell phenotype in culture.

    PubMed

    Spillekom, Sandra; Smolders, Lucas A; Grinwis, Guy C M; Arkesteijn, Irene T M; Ito, Keita; Meij, Björn P; Tryfonidou, Marianna A

    2014-08-01

    Degeneration of the intervertebral disc (IVD) is associated with a loss of notochordal cells (NCs) from the nucleus pulposus (NP) and their replacement by chondrocyte-like cells. NCs are known to maintain extracellular matrix quality and stimulate the chondrocyte-like NP cells, making NCs attractive for designing new tissue engineering approaches for IVD regeneration. However, optimal conditions, such as osmolarity and other characteristics of the culture media, for long-term culture of NCs are not known. The purpose of this study was to investigate the effects of different culture media and osmolarity on the physiology of NCs in vitro. NC clusters isolated from canine IVDs were suspended in alginate beads and cultured at 37°C under normoxic conditions for 28 days. Three different culture conditions were investigated; (1) Dulbecco's modified Eagle's medium (DMEM)/F12 (300 mOsm/L), (2) α-MEM (300 mOsm/L), and (3) α-MEM adjusted to 400 mOsm/L to mimic a hyperosmolar environment. NC morphology, expression of genes related to NC markers, matrix production and remodeling, and DNA- and glycosaminoglycan (GAG) analyses were performed on 1, 7, 14, and 28 days in culture. Large, vesicle-containing cells organized in clusters, characterized as NCs, remained present during 28 days for all culture conditions. However, the proportion of the NC clusters decreased over time, whereas the proportion of spindle-shaped cells increased. Gene expression profiling at 7, 14, and 28 days in culture compared to day 1 indicated a initial loss of NC phenotype followed by some recovery of brachyury and aggrecan gene expression after 28 days of culture supporting a potential recovery of NC phenotype. NCs cultured in α-MEM adjusted to 400 mOsm/L showed the highest gene expression of brachyury, cytokeratin 18, and aggrecan, the highest GAG production, and the lowest collagen 1α1 gene expression. In conclusion, NCs cultured in alginate in native cell clusters, partially retained their

  8. Cell Delivery System for Traumatic Brain Injury

    DTIC Science & Technology

    2008-03-21

    from collagen sponges using the dish test method (Figure 16B). The advantage of the dish test over the diffusion cells test is that samples can be...composite mat with collagen fibers and some chitosan fibers as well as globules but were not able to test for cell response to these matrices...dimensional collagen scaffold. MSCs cultured in monolayer and on a three- dimensional collagen sponge were treated with retinoic acid (RA) for up to

  9. Two sexually dimorphic cell groups in the human brain.

    PubMed

    Allen, L S; Hines, M; Shryne, J E; Gorski, R A

    1989-02-01

    A quantitative analysis of the volume of 4 cell groups in the preoptic-anterior hypothalamic area (PO-AHA) and of the supraoptic nucleus (SON) of the human brain was performed in 22 age-matched male and female individuals. We suggest the term Interstitial Nuclei of the Anterior Hypothalamus (INAH 1-4) to identify these 4 previously undescribed cell groups in the PO-AHA. While 2 INAH and the SON were not sexually dimorphic, gender-related differences were found in the other 2 cell groups. One nucleus (INAH-3) was 2.8 times larger in the male brain than in the female brain irrespective of age. The other cell group (INAH-2) was twice as large in the male brain, but also appeared to be related in women to circulating steroid hormone levels. Since the PO-AHA influences gonadotropin secretion, maternal behavior, and sexual behavior in several mammalian species, these results suggest that functional sex differences in the hypothalamus may be related to sex differences in neural structure.

  10. Alterations of natural killer cells in traumatic brain injury.

    PubMed

    Kong, Xiao-Dong; Bai, Sheng; Chen, Xin; Wei, Hui-Jie; Jin, Wei-Na; Li, Min-Shu; Yan, Yaping; Shi, Fu-Dong

    2014-12-01

    To investigate the relationship between natural killer (NK) cells and traumatic brain injury (TBI), we tracked an established phenotype of circulating NK cells at several time points in patients with different grades of TBI. In serial peripheral blood samples, NK cells were prospectively measured by flow cytometry of CD3(-) CD56(+) lymphocytes. Compared to healthy controls, TBI patients had reductions in both the percentage and the absolute number of NK cells. Furthermore, the magnitude of NK cell reduction correlated with the degree of TBI severity at several time points. That is, NK cell population size was independently associated with lower Glasgow Coma Scale scores. In addition, at some time points, a positive correlation was found between the NK cell counts and Glasgow Outcome Scale scores. Our results indicate that TBI induces a reduction in the number of NK cells, and the magnitude of the reduction appears to parallel the severity of TBI.

  11. High-resolution single-cell imaging for functional studies in the whole brain and spinal cord and thick tissue blocks using light-emitting diode illumination

    PubMed Central

    Safronov, Boris V.; Pinto, Vitor; Derkach, Victor A.

    2009-01-01

    Functional studies of neuronal networks require recordings from visually identified neurons in their natural environment, preservation of which may demand experimenting with a tissue of a significant depth or the entire brain. Here we describe a new technique of single-cell imaging and visually controlled patch-clamp recordings in both brain slices of unlimited thickness and the whole brain or spinal cord preparations with a cut upper surface. It utilizes an upright microscope and ultra bright light-emitting diodes (LEDs) as a source of oblique illumination. This technique provided high quality images of superficial cells regardless of slice thickness or the presence of opaque structures, like metal plate or bone, below the tissue, when conventional differential interference contrast (DIC) optics became powerless. The technique opens broad possibilities for a single-cell imaging and visually guided recordings from intact neuronal networks in the entire brain or spinal cord. PMID:17586052

  12. Purification of endothelial cells from rodent brain by immunopanning.

    PubMed

    Zhou, Lu; Sohet, Fabien; Daneman, Richard

    2014-01-01

    This protocol describes the use of immunopanning to purify endothelial cells from the rodent brain. Immunopanning permits the prospective isolation of endothelial cells from nervous tissue by relying on the binding of the endothelial cells to an anti-CD31 antibody adhered to a Petri dish. The cells are viable at the end of this gentle procedure, and they can be analyzed acutely for gene expression or cultured alone or in coculture with other central nervous system (CNS) cell types, including CNS pericytes and CNS astrocytes. This procedure can be used to isolate endothelial cells from either rat or mouse. We have suggested specific antibodies that work for each species. Note that endothelial cells from rats and mice have different morphologies; in general, rat CNS endothelial cells are longer and thinner than mouse CNS endothelial cells. This procedure can also be used to purify endothelial cells from different regions of the CNS, including brain and optic nerve. Dissociation procedures must be optimized for each tissue.

  13. Electrophysiological Properties of Subventricular Zone Cells in Adult Mouse Brain

    PubMed Central

    Lai, Bin; Mao, Xiao Ou; Xie, Lin; Chang, Su-Youne; Xiong, Zhi-Gang; Jin, Kunlin; Greenberg, David A.

    2010-01-01

    The subventricular zone (SVZ) is a principal site of adult neurogenesis and appears to participate in the brain’s response to injury. Thus, measures that enhance SVZ neurogenesis may have a role in treatment of neurological disease. To better characterize SVZ cells and identify potential targets for therapeutic intervention, we studied electrophysiological properties of SVZ cells in adult mouse brain slices using patch-clamp techniques. Electrophysiology was correlated with immunohistochemical phenotype by injecting cells with lucifer yellow and by studying transgenic mice carrying green fluorescent protein under control of the doublecortin (DCX) or glial fibrillary acidic protein (GFAP) promoter. We identified five types of cells in the adult mouse SVZ: type 1 cells, with 4-aminopyridine (4-AP)/tetraethylammonium (TEA)-sensitive and CdCl2-sensitive inward currents; type 2 cells, with Ca2+-sensitive K+ and both 4-AP/TEA-sensitive and -insensitive currents; type 3 cells, with 4-AP/TEA-sensitive and -insensitive and small Na+ currents; type 4 cells, with slowly activating, large linear outward current and sustained outward current without fast-inactivating component; and type 5 cells, with a large outward rectifying current with a fast inactivating component. Type 2 and 3 cells expressed DCX, types 4 and 5 cells expressed GFAP, and type 1 cells expressed neither. We propose that SVZ neurogenesis involves a progression of electrophysiological cell phenotypes from types 4 and 5 cells (astrocytes) to type 1 cells (neuronal progenitors) to types 2 and 3 cells (nascent neurons), and that drugs acting on. ion channels expressed during neurogenesis might promote therapeutic neurogenesis in the injured brain. PMID:20434436

  14. Brain morphology in children with nevoid basal cell carcinoma syndrome.

    PubMed

    Shiohama, Tadashi; Fujii, Katsunori; Miyashita, Toshiyuki; Mizuochi, Hiromi; Uchikawa, Hideki; Shimojo, Naoki

    2017-04-01

    Brain morphology is tightly regulated by diverse signaling pathways. Hedgehog signaling is a candidate pathway considered responsible for regulating brain morphology. Nevoid basal cell carcinoma syndrome (NBCCS), caused by a PTCH1 mutation in the hedgehog signaling pathway, occasionally exhibits macrocephaly and medulloblastoma. Although cerebellar enlargement occurs in ptch1 heterozygous-deficient mice, its impact on human brain development remains unknown. We investigated the brain morphological characteristics of children with NBCCS. We evaluated brain T1-weighted images from nine children with NBCCS and 15 age-matched normal control (NC) children (mean [standard deviation], 12.2 [2.8] vs. 11.6 [2.3] years old). The diameters of the cerebrum, corpus callosum, and brain stem and the cerebellar volume were compared using two-tailed t-tests with Welch's correction. The transverse diameters (150.4 [9.9] vs. 136.0 [5.5] mm, P = 0.002) and longitudinal diameters (165.4 [8.0] vs. 151.3 [8.7] mm, P = 0.0007) of the cerebrum, cross-sectional area of the cerebellar vermis (18.7 [2.6] vs. 11.8 [1.7] cm(2) , P = 0.0001), and total volume of the cerebellar hemispheres (185.1 [13.0] vs. 131.9 [10.4] cm(3) , P = 0.0001) were significantly larger in the children with NBCCS than in NC children. Thinning of the corpus callosum and ventricular enlargement were also confirmed in children with NBCCS. We demonstrate that, on examination of the brain morphology, an increase in the size of the cerebrum, cerebellum, and cerebral ventricles is revealed in children with NBCCS compared to NC children. This suggests that constitutively active hedgehog signaling affects human brain morphology and the PI3K/AKT and RAS/MAPK pathways.

  15. Computational neuroanatomy: mapping cell-type densities in the mouse brain, simulations from the Allen Brain Atlas

    NASA Astrophysics Data System (ADS)

    Grange, Pascal

    2015-09-01

    The Allen Brain Atlas of the adult mouse (ABA) consists of digitized expression profiles of thousands of genes in the mouse brain, co-registered to a common three-dimensional template (the Allen Reference Atlas).This brain-wide, genome-wide data set has triggered a renaissance in neuroanatomy. Its voxelized version (with cubic voxels of side 200 microns) is available for desktop computation in MATLAB. On the other hand, brain cells exhibit a great phenotypic diversity (in terms of size, shape and electrophysiological activity), which has inspired the names of some well-studied cell types, such as granule cells and medium spiny neurons. However, no exhaustive taxonomy of brain cell is available. A genetic classification of brain cells is being undertaken, and some cell types have been chraracterized by their transcriptome profiles. However, given a cell type characterized by its transcriptome, it is not clear where else in the brain similar cells can be found. The ABA can been used to solve this region-specificity problem in a data-driven way: rewriting the brain-wide expression profiles of all genes in the atlas as a sum of cell-type-specific transcriptome profiles is equivalent to solving a quadratic optimization problem at each voxel in the brain. However, the estimated brain-wide densities of 64 cell types published recently were based on one series of co-registered coronal in situ hybridization (ISH) images per gene, whereas the online ABA contains several image series per gene, including sagittal ones. In the presented work, we simulate the variability of cell-type densities in a Monte Carlo way by repeatedly drawing a random image series for each gene and solving the optimization problem. This yields error bars on the region-specificity of cell types.

  16. Cold preservation of endothelial cells in sucrose-based solution (SbS) and University of Wisconsin (UW) solutions: comparison of normoxic or hypoxic storage.

    PubMed

    Hawkins, M; Sales, K M; Dijk, S; Fuller, B

    2005-01-01

    Cold preservation of endothelial cells was studied, comparing primary endothelial cells (human umbilical vein endothelial cells - HUVECs) and a continuously growing cell line (ECV304 cells). Viability at the end of 24h cold preservation was measured by dye exclusion, whilst metabolism was assessed by Alamar blue conversion. Two preservation solutions were studied (UW solution) and sucrose-based (SbS) in both cell types. The response was similar in both cell types to preservation under normoxic conditions (with percentage dye exclusion maintained at about 80 percent in both preservation solutions) whereas under hypoxic conditions ECV304 were more sensitive to preservation in UW solution (dye exclusion reduced to 43.5+/-1.4 percent versus 73.6+/-14 percent (P<0.01). Metabolism assessed by Alamar blue conversion after cold preservation and rewarming was similar in both ECV304 and HUVECs after storage under normoxic conditions in UW solution, but in both cell types, metabolism was higher in SbS (P<0.05 and p<0.01) than in UW solution. Under hypoxic conditions, both cell types showed similar recovery of metabolism after storage in either UW or SbS. If the cells (in this case ECV304 under aerobic conditions) were stored for 24h and then allowed to rewarm in either of the respective preservation solutions (UW or SbS for 1h) before the Alamar blue test, metabolism was higher (p less than 0.01) in those exposed to SbS. UW solution and SbS provide similar protection for endothelial cells under hypoxic conditions, but SbS has some advantages under normoxic storage or if the cells experience variable temperatures in the presence of residual preservation solution at the end of cold preservation period.

  17. Small caliber arterial endothelial cells calcium signals elicited by PAR2 are preserved from endothelial dysfunction

    PubMed Central

    Hennessey, John C; Stuyvers, Bruno D; McGuire, John J

    2015-01-01

    Endothelial cell (EC)-dependent vasodilation by proteinase-activated receptor 2 (PAR2) is preserved in small caliber arteries in disease states where vasodilation by muscarinic receptors is decreased. In this study, we identified and characterized the PAR2-mediated intracellular calcium (Ca2+)-release mechanisms in EC from small caliber arteries in healthy and diseased states. Mesenteric arterial EC were isolated from PAR2 wild-type (WT) and null mice, after saline (controls) or angiotensin II (AngII) infusion, for imaging intracellular calcium and characterizing the calcium-release system by immunofluorescence. EC Ca2+ signals comprised two forms of Ca2+-release events that had distinct spatial-temporal properties and occurred near either the plasmalemma (peripheral) or center of EC. In healthy EC, PAR2-dependent increases in the densities and firing rates of both forms of Ca2+-release were abolished by inositol 1,4,5- trisphosphate receptor (IP3R) inhibitor, but partially reduced by transient potential vanilloid channels inhibitor ruthenium red (RR). Acetylcholine (ACh)-induced less overall Ca2+-release than PAR2 activation, but enhanced selectively the incidence of central events. PAR2-dependent Ca2+-activity, inhibitors sensitivities, IP3R, small- and intermediate-conductance Ca2+-activated potassium channels expressions were unchanged in EC from AngII WT. However, the same cells exhibited decreases in ACh-induced Ca2+-release, RR sensitivity, and endothelial nitric oxide synthase expression, indicating AngII-induced dysfunction was differentiated by receptor, Ca2+-release, and downstream targets of EC activation. We conclude that PAR2 and muscarinic receptors selectively elicit two elementary Ca2+ signals in single EC. PAR2-selective IP3R-dependent peripheral Ca2+-release mechanisms are identical between healthy and diseased states. Further study of PAR2-selective Ca2+-release for eliciting pathological and/or normal EC functions is warranted. PMID:25729579

  18. Doublecortin in Oligodendrocyte Precursor Cells in the Adult Mouse Brain

    PubMed Central

    Boulanger, Jenna J.; Messier, Claude

    2017-01-01

    Key Points Oligodendrocyte precursor cells express doublecortin, a microtubule-associated protein.Oligodendrocyte precursor cells express doublecortin, but at a lower level of expression than in neuronal precursor.Doublecortin is not associated with a potential immature neuronal phenotype in Oligodendrocyte precursor cells. Oligodendrocyte precursor cells (OPC) are glial cells that differentiate into myelinating oligodendrocytes during embryogenesis and early stages of post-natal life. OPCs continue to divide throughout adulthood and some eventually differentiate into oligodendrocytes in response to demyelinating lesions. There is growing evidence that OPCs are also involved in activity-driven de novo myelination of previously unmyelinated axons and myelin remodeling in adulthood. Considering these roles in the adult brain, OPCs are likely mobile cells that can migrate on some distances before they differentiate into myelinating oligodendrocytes. A number of studies have noted that OPCs express doublecortin (DCX), a microtubule-associated protein expressed in neural precursor cells and in migrating immature neurons. Here we describe the distribution of DCX in OPCs. We found that almost all OPCs express DCX, but the level of expression appears to be much lower than what is found in neural precursor. We found that DCX is downregulated when OPCs start expressing mature oligodendrocyte markers and is absent in myelinating oligodendrocytes. DCX does not appear to signal an immature neuronal phenotype in OPCs in the adult mouse brain. Rather, it could be involved either in cell migration, or as a marker of an immature oligodendroglial cell phenotype.

  19. Radiopotentiation of human brain tumor cells by sodium phenylacetate.

    PubMed

    Ozawa, T; Lu, R M; Hu, L J; Lamborn, K R; Prados, M D; Deen, D F

    1999-08-03

    Phenylacetate (PA) inhibits the growth of tumor cells in vitro and in vivo and shows promise as a relatively nontoxic agent for cancer treatment. A recent report shows that prolonged exposure of cells to low concentrations of PA can enhance the radiation response of brain tumor cells in vitro, opening up the possibility of using this drug to improve the radiation therapy of brain tumor patients. We investigated the cytotoxicity produced by sodium phenylacetate (NaPA) alone and in combination with X-rays in SF-767 human glioblastoma cells and in two medulloblastoma cell lines, Masden and Daoy. Exposure of all three cell lines to relatively low concentrations of NaPA for up to 5 days did not enhance the subsequent cell killing produced by X-irradiation. However, enhanced cell killing was achieved by exposing either oxic or hypoxic cells to relatively high drug concentrations ( > 50-70 mM) for 1 h immediately before X-irradiation. Because central nervous system toxicity can occur in humans at serum concentrations of approximately 6 mM PA, translation of these results into clinical trials will likely require local drug-delivery strategies to achieve drug concentrations that can enhance the radiation response. The safety of such an approach with this drug has not been demonstrated.

  20. Internally and externally generated emotions in people with acquired brain injury: preservation of emotional experience after right hemisphere lesions.

    PubMed

    Salas Riquelme, Christian E; Radovic, Darinka; Castro, Osvaldo; Turnbull, Oliver H

    2015-01-01

    The study of emotional changes after brain injury has contributed enormously to the understanding of the neural basis of emotion. However, little attention has been placed on the methods used to elicit emotional responses in people with brain damage. Of particular interest are subjects with right hemisphere [RH] cortical lesions, who have been described as presenting impairment in emotional processing. In this article, an internal and external mood induction procedure [MIP] was used to trigger positive and negative emotions, in a sample of 10 participants with RH damage, and 15 healthy controls. Emotional experience was registered by using a self-report questionnaire. As observed in previous studies, internal and external MIPs were equally effective in eliciting the target emotion, but the internal procedure generated higher levels of intensity. Remarkably, participants with RH lesions were equally able to experience both positive and negative affect. The results are discussed in relation to the role of the RH in the capacity to experience negative emotions.

  1. Retrovirus-mediated transduction of a cytosine deaminase gene preserves the stemness of mesenchymal stem cells.

    PubMed

    Park, Jin Sung; Chang, Da-Young; Kim, Ji-Hoi; Jung, Jin Hwa; Park, JoonSeong; Kim, Se-Hyuk; Lee, Young-Don; Kim, Sung-Soo; Suh-Kim, Haeyoung

    2013-02-22

    Human mesenchymal stem cells (MSCs) have emerged as attractive cellular vehicles to deliver therapeutic genes for ex-vivo therapy of diverse diseases; this is, in part, because they have the capability to migrate into tumor or lesion sites. Previously, we showed that MSCs could be utilized to deliver a bacterial cytosine deaminase (CD) suicide gene to brain tumors. Here we assessed whether transduction with a retroviral vector encoding CD gene altered the stem cell property of MSCs. MSCs were transduced at passage 1 and cultivated up to passage 11. We found that proliferation and differentiation potentials, chromosomal stability and surface antigenicity of MSCs were not altered by retroviral transduction. The results indicate that retroviral vectors can be safely utilized for delivery of suicide genes to MSCs for ex-vivo therapy. We also found that a single retroviral transduction was sufficient for sustainable expression up to passage 10. The persistent expression of the transduced gene indicates that transduced MSCs provide a tractable and manageable approach for potential use in allogeneic transplantation.

  2. Cell proliferation and cell death are disturbed during prenatal and postnatal brain development after uranium exposure.

    PubMed

    Legrand, M; Elie, C; Stefani, J; N Florès; Culeux, C; Delissen, O; Ibanez, C; Lestaevel, P; Eriksson, P; Dinocourt, C

    2016-01-01

    The developing brain is more susceptible to neurotoxic compounds than adult brain. It is also well known that disturbances during brain development cause neurological disorders in adulthood. The brain is known to be a target organ of uranium (U) exposure and previous studies have noted that internal U contamination of adult rats induces behavioral disorders as well as affects neurochemistry and neurophysiological properties. In this study, we investigated whether depleted uranium (DU) exposure affects neurogenesis during prenatal and postnatal brain development. We examined the structural morphology of the brain, cell death and finally cell proliferation in animals exposed to DU during gestation and lactation compared to control animals. Our results showed that DU decreases cell death in the cortical neuroepithelium of gestational day (GD) 13 embryos exposed at 40mg/L and 120mg/L and of GD18 fetuses exposed at 120mg/L without modification of the number of apoptotic cells. Cell proliferation analysis showed an increase of BrdU labeling in the dentate neuroepithelium of fetuses from GD18 at 120mg/L. Postnatally, cell death is increased in the dentate gyrus of postnatal day (PND) 0 and PND5 exposed pups at 120mg/L and is associated with an increase of apoptotic cell number only at PND5. Finally, a decrease in dividing cells is observed in the dentate gyrus of PND21 rats developmentally exposed to 120mg/L DU, but not at PND0 and PND5. These results show that DU exposure during brain development causes opposite effects on cell proliferation and cell death processes between prenatal and postnatal development mainly at the highest dose. Although these modifications do not have a major impact in brain morphology, they could affect the next steps of neurogenesis and thus might disrupt the fine organization of the neuronal network.

  3. Tomographic brain imaging with nucleolar detail and automatic cell counting

    NASA Astrophysics Data System (ADS)

    Hieber, Simone E.; Bikis, Christos; Khimchenko, Anna; Schweighauser, Gabriel; Hench, Jürgen; Chicherova, Natalia; Schulz, Georg; Müller, Bert

    2016-09-01

    Brain tissue evaluation is essential for gaining in-depth insight into its diseases and disorders. Imaging the human brain in three dimensions has always been a challenge on the cell level. In vivo methods lack spatial resolution, and optical microscopy has a limited penetration depth. Herein, we show that hard X-ray phase tomography can visualise a volume of up to 43 mm3 of human post mortem or biopsy brain samples, by demonstrating the method on the cerebellum. We automatically identified 5,000 Purkinje cells with an error of less than 5% at their layer and determined the local surface density to 165 cells per mm2 on average. Moreover, we highlight that three-dimensional data allows for the segmentation of sub-cellular structures, including dendritic tree and Purkinje cell nucleoli, without dedicated staining. The method suggests that automatic cell feature quantification of human tissues is feasible in phase tomograms obtained with isotropic resolution in a label-free manner.

  4. Tomographic brain imaging with nucleolar detail and automatic cell counting

    PubMed Central

    Hieber, Simone E.; Bikis, Christos; Khimchenko, Anna; Schweighauser, Gabriel; Hench, Jürgen; Chicherova, Natalia; Schulz, Georg; Müller, Bert

    2016-01-01

    Brain tissue evaluation is essential for gaining in-depth insight into its diseases and disorders. Imaging the human brain in three dimensions has always been a challenge on the cell level. In vivo methods lack spatial resolution, and optical microscopy has a limited penetration depth. Herein, we show that hard X-ray phase tomography can visualise a volume of up to 43 mm3 of human post mortem or biopsy brain samples, by demonstrating the method on the cerebellum. We automatically identified 5,000 Purkinje cells with an error of less than 5% at their layer and determined the local surface density to 165 cells per mm2 on average. Moreover, we highlight that three-dimensional data allows for the segmentation of sub-cellular structures, including dendritic tree and Purkinje cell nucleoli, without dedicated staining. The method suggests that automatic cell feature quantification of human tissues is feasible in phase tomograms obtained with isotropic resolution in a label-free manner. PMID:27581254

  5. Preservation of Bacillus firmus strain 37 and optimization of cyclodextrin biosynthesis by cells immobilized on loofa sponge.

    PubMed

    Pazzetto, Rúbia; Ferreira, Sabrina Barbosa de Souza; Santos, Elder James Silva; Moriwaki, Cristiane; Guedes, Teresinha Aparecida; Matioli, Graciette

    2012-08-08

    The preservation of Bacillus firmus strain 37 cells by lyophilization was evaluated and response surface methodology (RSM) was used to optimize the β-cyclodextrin (β-CD) production by cells immobilized on loofa sponge. Interactions were studied with the variables temperature, pH and dextrin concentration using a central composite design (CCD). Immobilization time influence on β-CD production was also investigated. B. firmus strain 37 cells remained viable after one year of storage, showing that the lyophilization is a suitable method for preservation of the microorganism. From the three-dimensional diagrams and contour plots, the best conditions for β-CD production were determined: temperature 60 °C, pH 8, and 18% dextrin. Considering that the amount of dextrin was high, a new assay was carried out, in which dextrin concentrations of 10, 15, and 18% were tested and the temperature of 60 °C and pH 8 were maintained. The results achieved showed very small differences and therefore, for economic reasons, the use of 10% dextrin is suggested. Increasing the immobilization time of cells immobilized on synthetic sponge the β-CD production decreased and did not change for cells immobilized on loofa sponge. The results of this research are important for microorganism preservation and essential in the optimization of the biosynthesis of CD.

  6. The stem cell secretome and its role in brain repair.

    PubMed

    Drago, Denise; Cossetti, Chiara; Iraci, Nunzio; Gaude, Edoardo; Musco, Giovanna; Bachi, Angela; Pluchino, Stefano

    2013-12-01

    Compelling evidence exists that non-haematopoietic stem cells, including mesenchymal (MSCs) and neural/progenitor stem cells (NPCs), exert a substantial beneficial and therapeutic effect after transplantation in experimental central nervous system (CNS) disease models through the secretion of immune modulatory or neurotrophic paracrine factors. This paracrine hypothesis has inspired an alternative outlook on the use of stem cells in regenerative neurology. In this paradigm, significant repair of the injured brain may be achieved by injecting the biologics secreted by stem cells (secretome), rather than implanting stem cells themselves for direct cell replacement. The stem cell secretome (SCS) includes cytokines, chemokines and growth factors, and has gained increasing attention in recent years because of its multiple implications for the repair, restoration or regeneration of injured tissues. Thanks to recent improvements in SCS profiling and manipulation, investigators are now inspired to harness the SCS as a novel alternative therapeutic option that might ensure more efficient outcomes than current stem cell-based therapies for CNS repair. This review discusses the most recent identification of MSC- and NPC-secreted factors, including those that are trafficked within extracellular membrane vesicles (EVs), and reflects on their potential effects on brain repair. It also examines some of the most convincing advances in molecular profiling that have enabled mapping of the SCS.

  7. The stem cell secretome and its role in brain repair

    PubMed Central

    Drago, Denise; Cossetti, Chiara; Iraci, Nunzio; Gaude, Edoardo; Musco, Giovanna; Bachi, Angela; Pluchino, Stefano

    2014-01-01

    Compelling evidence exists that non-haematopoietic stem cells, including mesenchymal (MSCs) and neural/progenitor stem cells (NPCs), exert a substantial beneficial and therapeutic effect after transplantation in experimental central nervous system (CNS) disease models through the secretion of immune modulatory or neurotrophic paracrine factors. This paracrine hypothesis has inspired an alternative outlook on the use of stem cells in regenerative neurology. In this paradigm, significant repair of the injured brain may be achieved by injecting the biologics secreted by stem cells (secretome), rather than implanting stem cells themselves for direct cell replacement. The stem cell secretome (SCS) includes cytokines, chemokines and growth factors, and has gained increasing attention in recent years because of its multiple implications for the repair, restoration or regeneration of injured tissues. Thanks to recent improvements in SCS profiling and manipulation, investigators are now inspired to harness the SCS as a novel alternative therapeutic option that might ensure more efficient outcomes than current stem cell-based therapies for CNS repair. This review discusses the most recent identification of MSC- and NPC-secreted factors, including those that are trafficked within extracellular membrane vesicles (EVs), and reflects on their potential effects on brain repair. It also examines some of the most convincing advances in molecular profiling that have enabled mapping of the SCS. PMID:23827856

  8. Uptake and utilization of CDP-choline in primary brain cell cultures from fetal brain.

    PubMed

    Vecchini, A; Binaglia, L; Floridi, A; Palmerini, C A; Procellati, G

    1983-03-01

    The utilization of double-labeled CDP-choline by cultured brain cells has been studied. CDP-choline is demonstrated to be rapidly hydrolysed into CMP and choline phosphate. The fragments, or their hydrolysis products, penetrate into the cells and are utilized for lipid synthesis. At short times after the isotope administration a rapid labeling of phosphatidylcholine was detected, when cells were incubated with CDP-choline. The same was not seen when cells were incubated with labeled choline. From these observations it can be inferred that either CDP- choline can penetrate the cell membrane or that some mechanism involving CDP-choline and leading to phospholipid synthesis can work at the external surface of the plasma membranes.

  9. Targeting brain microvascular endothelial cells: a therapeutic approach to neuroprotection against stroke

    PubMed Central

    Yu, Qi-jin; Tao, Hong; Wang, Xin; Li, Ming-chang

    2015-01-01

    Brain microvascular endothelial cells form the interface between nervous tissue and circulating blood, and regulate central nervous system homeostasis. Brain microvascular endothelial cells differ from peripheral endothelial cells with regards expression of specific ion transporters and receptors, and contain fewer fenestrations and pinocytotic vesicles. Brain microvascular endothelial cells also synthesize several factors that influence blood vessel function. This review describes the morphological characteristics and functions of brain microvascular endothelial cells, and summarizes current knowledge regarding changes in brain microvascular endothelial cells during stroke progression and therapies. Future studies should focus on identifying mechanisms underlying such changes and developing possible neuroprotective therapeutic interventions. PMID:26807131

  10. Methamphetamine disrupts blood-brain barrier function by induction of oxidative stress in brain endothelial cells.

    PubMed

    Ramirez, Servio H; Potula, Raghava; Fan, Shongshan; Eidem, Tess; Papugani, Anil; Reichenbach, Nancy; Dykstra, Holly; Weksler, Babette B; Romero, Ignacio A; Couraud, Pierre O; Persidsky, Yuri

    2009-12-01

    Methamphetamine (METH), a potent stimulant with strong euphoric properties, has a high abuse liability and long-lasting neurotoxic effects. Recent studies in animal models have indicated that METH can induce impairment of the blood-brain barrier (BBB), thus suggesting that some of the neurotoxic effects resulting from METH abuse could be the outcome of barrier disruption. In this study, we provide evidence that METH alters BBB function through direct effects on endothelial cells and explore possible underlying mechanisms leading to endothelial injury. We report that METH increases BBB permeability in vivo, and exposure of primary human brain microvascular endothelial cells (BMVEC) to METH diminishes the tightness of BMVEC monolayers in a dose- and time-dependent manner by decreasing the expression of cell membrane-associated tight junction (TJ) proteins. These changes were accompanied by the enhanced production of reactive oxygen species, increased monocyte migration across METH-treated endothelial monolayers, and activation of myosin light chain kinase (MLCK) in BMVEC. Antioxidant treatment attenuated or completely reversed all tested aspects of METH-induced BBB dysfunction. Our data suggest that BBB injury is caused by METH-mediated oxidative stress, which activates MLCK and negatively affects the TJ complex. These observations provide a basis for antioxidant protection against brain endothelial injury caused by METH exposure.

  11. Transcranial amelioration of inflammation and cell death after brain injury

    NASA Astrophysics Data System (ADS)

    Roth, Theodore L.; Nayak, Debasis; Atanasijevic, Tatjana; Koretsky, Alan P.; Latour, Lawrence L.; McGavern, Dorian B.

    2014-01-01

    Traumatic brain injury (TBI) is increasingly appreciated to be highly prevalent and deleterious to neurological function. At present, no effective treatment options are available, and little is known about the complex cellular response to TBI during its acute phase. To gain insights into TBI pathogenesis, we developed a novel murine closed-skull brain injury model that mirrors some pathological features associated with mild TBI in humans and used long-term intravital microscopy to study the dynamics of the injury response from its inception. Here we demonstrate that acute brain injury induces vascular damage, meningeal cell death, and the generation of reactive oxygen species (ROS) that ultimately breach the glial limitans and promote spread of the injury into the parenchyma. In response, the brain elicits a neuroprotective, purinergic-receptor-dependent inflammatory response characterized by meningeal neutrophil swarming and microglial reconstitution of the damaged glial limitans. We also show that the skull bone is permeable to small-molecular-weight compounds, and use this delivery route to modulate inflammation and therapeutically ameliorate brain injury through transcranial administration of the ROS scavenger, glutathione. Our results shed light on the acute cellular response to TBI and provide a means to locally deliver therapeutic compounds to the site of injury.

  12. Derivation of blood-brain barrier endothelial cells from human pluripotent stem cells.

    PubMed

    Lippmann, Ethan S; Azarin, Samira M; Kay, Jennifer E; Nessler, Randy A; Wilson, Hannah K; Al-Ahmad, Abraham; Palecek, Sean P; Shusta, Eric V

    2012-08-01

    The blood-brain barrier (BBB) is crucial to the health of the brain and is often compromised in neurological disease. Moreover, because of its barrier properties, this endothelial interface restricts uptake of neurotherapeutics. Thus, a renewable source of human BBB endothelium could spur brain research and pharmaceutical development. Here we show that endothelial cells derived from human pluripotent stem cells (hPSCs) acquire BBB properties when co-differentiated with neural cells that provide relevant cues, including those involved in Wnt/β-catenin signaling. The resulting endothelial cells have many BBB attributes, including well-organized tight junctions, appropriate expression of nutrient transporters and polarized efflux transporter activity. Notably, they respond to astrocytes, acquiring substantial barrier properties as measured by transendothelial electrical resistance (1,450 ± 140 Ω cm2), and they possess molecular permeability that correlates well with in vivo rodent blood-brain transfer coefficients.

  13. Preservation of beta cell function after pancreatic islet autotransplantation: University of Chicago experience.

    PubMed

    Savari, Omid; Golab, Karolina; Wang, Ling-Jia; Schenck, Lindsay; Grose, Randall; Tibudan, Martin; Ramachandran, Sabarinathan; Chon, W James; Posner, Mitchell C; Millis, J Michael; Matthews, Jeffrey B; Gelrud, Andres; Witkowski, Piotr

    2015-04-01

    The aim of the study was to assess the rate of insulin independence in patients after total pancreatectomy (TP) and islet autotransplantation in our center. TP followed by islet autotransplantation was performed in 10 patients. Severe unrelenting pain associated with chronic pancreatitis was the major indication for surgery. Islets were isolated using the modified Ricordi method and infused through the portal vein. Exogenous insulin therapy was implemented for at least two months posttransplant to support islet engraftment and was subsequently weaned off, if possible. Median follow-up was 26 months (range, 2 to 60 months). Median islet yield was 158,860 islet equivalents (IEQ) (range, 40,203 to 330,472 IEQ) with an average islet yield of 2,478 IEQ/g (range, 685 to 6,002 IEQ/g) of processed pancreas. One patient developed transient partial portal vein thrombosis, which resolved without sequela. Five (50%) patients are currently off insulin with excellent glucose control and HbA1c below 6. Patients who achieved and maintained insulin independence were transplanted with significantly more islets (median, 202,291 IEQ; range, 145,000 to 330,474 IEQ) than patients who required insulin support (64,348 IEQ; range, 40,203 to 260,476 IEQ; P < 0.05). Patient body mass index and time of chronic pancreatitis prior transplant procedure did not correlate with the outcome. The remaining five patients, who require insulin support, had present C-peptide in blood and experience good glucose control without incidence of severe hypoglycemic episodes. Islet autotransplantation efficiently preserved beta cell function in selected patients with chronic pancreatitis and the outcome correlated with transplanted islet mass.

  14. Melatonin Treatment Improves Mesenchymal Stem Cells Therapy by Preserving Stemness during Long-term In Vitro Expansion

    PubMed Central

    Shuai, Yi; Liao, Li; Su, Xiaoxia; Yu, Yang; Shao, Bingyi; Jing, Huan; Zhang, Xinjing; Deng, Zhihong; Jin, Yan

    2016-01-01

    Mesenchymal stem cells (MSCs) are promising candidates for tissue regeneration and disease treatment. However, long-term in vitro passaging leads to stemness loss of MSCs, resulting in failure of MSCs therapy. Here, we report a melatonin-based strategy to improve cell therapy of in vitro cultured MSCs. Among four small molecules with anti-aging and stem cell-protection properties (rapamycin, resveratrol, quercetin and melatonin), colony forming, proliferation, and osteogenic differentiation assay showed that melatonin was the most efficient to preserve self-renewal and differentiation properties of rat bone marrow MSCs (BMMSCs) after long-term passaging. Functional assays confirmed melatonin treatment did not affect the colony forming, proliferation and osteogenic differentiation of BMMSCs cultured for 1 or 4 passages, but largely prevented the decline of self-renew and differentiation capacity of BMMSCs cultured for 15 passages in vitro. Furthermore, heterotopic osteogenesis assay, critical size calvarial defects repair assay, osteoporosis treatment and experimental colitis therapy assay strongly certified that melatonin preserved the therapeutic effect of long-term passaged BMMSCs on bone regeneration and immunotherapy in vivo. Mechanistically, melatonin functioned by activating antioxidant defense system, inhibiting the pathway of cell senescence, and preserving the expression of gene governing the stemness. Taken together, our findings showed that melatonin treatment efficiently prevented the dysfunction and therapeutic failure of BMMSCs after long-term passaging, providing a practical strategy to improve the application of BMMSCs in tissue engineering and cytotherapy. PMID:27570559

  15. Stereotaxic implantation of dispersed cell suspensions into brain. A systematic appraisal of cell placement and survival

    SciTech Connect

    Plunkett, R.J.; Weber, R.J.; Oldfield, E.H.

    1988-08-01

    The application of several recent advances in cell biology, brain implantation, and cell-mediated tumor immunotherapy requires successful and reproducible placement of viable cell suspensions into brain. Stereotaxic implantation is being used to inject cytotoxic lymphocytes into gliomas and to replace dopaminergic cells in parkinsonian models. Systematic assessment of the factors that influence success in implantation of cell suspensions into solid tissues is needed. A model was developed for investigation of stereotaxic implantation using radiolabeled rat lymphokine-activated killer (LAK) cells. Anesthetized rats received microliter injections of cell suspension into the right caudate nucleus. The injection volume, cell concentration, infusion rate, and needle size were varied systematically. The animals were sacrificed 1 hour after injection; the brain was removed and sectioned, and the radioactivity was counted. Three aliquots of the suspension were injected into counting tubes for control analysis. Recovery of radioactivity was expressed as the percent of mean counts per minute (cpm) in the right frontal lobe/mean cpm in the three control tubes. To assess the viability of implanted cells, the right frontal region was mechanically dissociated in media and centrifuged, and the pellet and supernatant were counted. By using small needles and slow infusion of volumes of 10 microliters or less, 85% to 90% of the radioactivity was recovered in the caudate nucleus. At least half of the implanted cells were viable. Consistent, accurate implantation of dispersed cells into brain over a range of volumes, cell concentrations, infusion rates, and needle sizes was achieved.

  16. Preserving elemental content in adherent mammalian cells for analysis by synchrotron‐based x‐ray fluorescence microscopy

    PubMed Central

    JIN, QIAOLING; PAUNESKU, TATJANA; LAI, BARRY; GLEBER, SOPHIE‐CHARLOTTE; CHEN, SI; FINNEY, LYDIA; VINE, DAVID; VOGT, STEFAN; WOLOSCHAK, GAYLE

    2016-01-01

    Summary Trace metals play important roles in biological function, and x‐ray fluorescence microscopy (XFM) provides a way to quantitatively image their distribution within cells. The faithfulness of these measurements is dependent on proper sample preparation. Using mouse embryonic fibroblast NIH/3T3 cells as an example, we compare various approaches to the preparation of adherent mammalian cells for XFM imaging under ambient temperature. Direct side‐by‐side comparison shows that plunge‐freezing‐based cryoimmobilization provides more faithful preservation than conventional chemical fixation for most biologically important elements including P, S, Cl, K, Fe, Cu, Zn and possibly Ca in adherent mammalian cells. Although cells rinsed with fresh media had a great deal of extracellular background signal for Cl and Ca, this approach maintained cells at the best possible physiological status before rapid freezing and it does not interfere with XFM analysis of other elements. If chemical fixation has to be chosen, the combination of 3% paraformaldehyde and 1.5 % glutaraldehyde preserves S, Fe, Cu and Zn better than either fixative alone. When chemically fixed cells were subjected to a variety of dehydration processes, air drying was proved to be more suitable than other drying methods such as graded ethanol dehydration and freeze drying. This first detailed comparison for x‐ray fluorescence microscopy shows how detailed quantitative conclusions can be affected by the choice of cell preparation method. PMID:27580164

  17. The organ preservation and enhancement of donation success ratio effect of extracorporeal membrane oxygenation in circulatory unstable brain death donor.

    PubMed

    Fan, Xiaoli; Chen, Zhiquan; Nasralla, David; Zeng, Xianpeng; Yang, Jing; Ye, Shaojun; Zhang, Yi; Peng, Guizhu; Wang, Yanfeng; Ye, Qifa

    2016-10-01

    Between 2010 and 2013, we recorded 66 cases of failed organ donation after brain death (DBD) due to the excessive use of the vasoactive drugs resulting in impaired hepatic and/or renal function. To investigate the effect of extracorporeal membrane oxygenation (ECMO) in donor management, ECMO was used to provide support for DBD donors with circulatory and/or respiratory failure from 2013 to 2015. A retrospective cohort study between circulatory non-stable DBD with vasoactive drugs (DBD-drug) and circulatory non-stable DBD with ECMO (DBD-ECMO) was designed to compare the transplant outcomes. A total of 19 brain death donors were supported by ECMO. The incidence rate of post-transplant liver primary non-function (PNF) was 10% (two of 20) in DBD-drug group and zero in DBD-ECMO group. Kidney function indicators, including creatinine clearance and urine production, were significantly better in DBD-ECMO group, as well as the kidney delayed graft function (DGF) rate was found to be decreased by the use of ECMO in our study. Donation success rate increased steadily from 47.8% in 2011 to 84.6% in 2014 after the ECMO intervention. The use of ECMO in assisting circulatory and respiratory function of DBD can reduce liver and kidney injury from vasoactive drugs, thereby improving organ quality and reducing the organ discard rates.

  18. Internally and externally generated emotions in people with acquired brain injury: preservation of emotional experience after right hemisphere lesions

    PubMed Central

    Salas Riquelme, Christian E.; Radovic, Darinka; Castro, Osvaldo; Turnbull, Oliver H.

    2015-01-01

    The study of emotional changes after brain injury has contributed enormously to the understanding of the neural basis of emotion. However, little attention has been placed on the methods used to elicit emotional responses in people with brain damage. Of particular interest are subjects with right hemisphere [RH] cortical lesions, who have been described as presenting impairment in emotional processing. In this article, an internal and external mood induction procedure [MIP] was used to trigger positive and negative emotions, in a sample of 10 participants with RH damage, and 15 healthy controls. Emotional experience was registered by using a self-report questionnaire. As observed in previous studies, internal and external MIPs were equally effective in eliciting the target emotion, but the internal procedure generated higher levels of intensity. Remarkably, participants with RH lesions were equally able to experience both positive and negative affect. The results are discussed in relation to the role of the RH in the capacity to experience negative emotions. PMID:25762951

  19. A silk platform that enables electrophysiology and targeted drug delivery in brain astroglial cells

    PubMed Central

    Benfenati, Valentina; Toffanin, Stefano; Capelli, Raffaella; Camassa, Laura M. A.; Ferroni, Stefano; Kaplan, David L.; Omenetto, Fiorenzo G.; Muccini, Michele; Zamboni, Roberto

    2010-01-01

    Astroglial cell survival and ion channel activity are relevant molecular targets for the mechanistic study of neural cell interactions with biomaterials and/or electronic interfaces. Astrogliosis is the most typical reaction to in-vivo brain implants and needs to be avoided by developing biomaterials that preserve astroglial cell physiological function. This cellular phenomenon is characterized by a proliferative state and altered expression of astroglial potassium (K+) channels. Silk is a natural polymer with potential for new biomedical applications due to its ability to support in vitro growth and differentiation of many cell types. We report on silk interactions with cultured neocortical astroglial cells. Astrocytes survival is similar when plated on silk-coated glass and on poly-D-lysine (PDL), a well-known polyionic substrate used to promote astroglial cell adhesion to glass surfaces. Comparative analyses of whole-cell and single-cell patch-clamp experiments reveal that silk- and PDL-coated cells display depolarized resting membrane potentials (∼ -40 mV), very high input resistance, and low specific conductance, with values similar to those of undifferentiated glial cells. Analysis of K+ channel conductance reveals that silk-astrocytes express large outwardly delayed rectifying K+ current (KDR). The magnitude of KDR in PDL- and silk-coated astrocytes is similar, indicating that silk does not alter the resting K+ current. We also demonstrate that guanosine-(GUO) embedded silk enables the direct modulation of astroglial K+ conductance in vitro. Astrocytes plated on GUO-embedded silk are more hyperpolarized and express inward rectifying K+ conductance (Kir). The K+ inward current increase and this is paralleled by upregulation and membrane-polarization of Kir4.1 protein signal. Collectively these results indicate that silk is a suitable biomaterial platform for the in vitro studies of astroglial ion channel responses and related physiology. PMID:20688390

  20. What cell biologists should know about the National Institutes of Health BRAIN Initiative.

    PubMed

    Insel, Thomas R; Koroshetz, Walter

    2015-12-15

    The BRAIN (Brain Research through Advancing Innovative Neurotechnologies) Initiative is an ambitious project to develop innovative tools for a deeper understanding of how the brain functions in health and disease. Early programs in the National Institutes of Health BRAIN Initiative focus on tools for next-generation imaging and recording, studies of cell diversity and cell census, and integrative approaches to circuit function. In all of these efforts, cell biologists can play a leading role.

  1. Preservation of Essential Odor-Guided Behaviors and Odor-Based Reversal Learning after Targeting Adult Brain Serotonin Synthesis

    PubMed Central

    Carlson, Kaitlin S.

    2016-01-01

    Abstract The neurotransmitter serotonin (5-HT) is considered a powerful modulator of sensory system organization and function in a wide range of animals. The olfactory system is innervated by midbrain 5-HT neurons into both its primary and secondary odor-processing stages. Facilitated by this circuitry, 5-HT and its receptors modulate olfactory system function, including odor information input to the olfactory bulb. It is unknown, however, whether the olfactory system requires 5-HT for even its most basic behavioral functions. To address this question, we established a conditional genetic approach to specifically target adult brain tryptophan hydroxylase 2 (Tph2), encoding the rate-limiting enzyme in brain 5-HT synthesis, and nearly eliminate 5-HT from the mouse forebrain. Using this novel model, we investigated the behavior of 5-HT–depleted mice during performance in an olfactory go/no-go task. Surprisingly, the near elimination of 5-HT from the forebrain, including the olfactory bulbs, had no detectable effect on the ability of mice to perform the odor-based task. Tph2-targeted mice not only were able to learn the task, but also had levels of odor acuity similar to those of control mice when performing coarse odor discrimination. Both groups of mice spent similar amounts of time sampling odors during decision-making. Furthermore, odor reversal learning was identical between 5-HT–depleted and control mice. These results suggest that 5-HT neurotransmission is not necessary for the most essential aspects of olfaction, including odor learning, discrimination, and certain forms of cognitive flexibility. PMID:27896310

  2. Tight regulation of ubiquitin-mediated DNA damage response by USP3 preserves the functional integrity of hematopoietic stem cells

    PubMed Central

    Lancini, Cesare; van den Berk, Paul C.M.; Vissers, Joseph H.A.; Gargiulo, Gaetano; Song, Ji-Ying; Hulsman, Danielle; Serresi, Michela; Tanger, Ellen; Blom, Marleen; Vens, Conchita; van Lohuizen, Maarten; Jacobs, Heinz

    2014-01-01

    Histone ubiquitination at DNA breaks is required for activation of the DNA damage response (DDR) and DNA repair. How the dynamic removal of this modification by deubiquitinating enzymes (DUBs) impacts genome maintenance in vivo is largely unknown. To address this question, we generated mice deficient for Ub-specific protease 3 (USP3; Usp3Δ/Δ), a histone H2A DUB which negatively regulates ubiquitin-dependent DDR signaling. Notably, USP3 deletion increased the levels of histone ubiquitination in adult tissues, reduced the hematopoietic stem cell (HSC) reserves over time, and shortened animal life span. Mechanistically, our data show that USP3 is important in HSC homeostasis, preserving HSC self-renewal, and repopulation potential in vivo and proliferation in vitro. A defective DDR and unresolved spontaneous DNA damage contribute to cell cycle restriction of Usp3Δ/Δ HSCs. Beyond the hematopoietic system, Usp3Δ/Δ animals spontaneously developed tumors, and primary Usp3Δ/Δ cells failed to preserve chromosomal integrity. These findings broadly support the regulation of chromatin ubiquitination as a key pathway in preserving tissue function through modulation of the response to genotoxic stress. PMID:25113974

  3. Brain self-protection: the role of endogenous neural progenitor cells in adult brain after cerebral cortical ischemia.

    PubMed

    Li, Bin; Piao, Chun-Shu; Liu, Xiao-Yun; Guo, Wen-Ping; Xue, Yue-Qiang; Duan, Wei-Ming; Gonzalez-Toledo, Maria E; Zhao, Li-Ru

    2010-04-23

    Convincing evidence has shown that brain ischemia causes the proliferation of neural stem cells/neural progenitor cells (NSCs/NPCs) in both the subventricular zone (SVZ) and the subgranular zone (SGZ) of adult brain. The role of brain ischemia-induced NSC/NPC proliferation, however, has remained unclear. Here we have determined whether brain ischemia-induced amplification of the NSCs/NPCs in adult brain is required for brain self-protection. The approach of intracerebroventricular (ICV) infusion of cytosine arabinoside (Ara-C), an inhibitor for cell proliferation, for the first 7days after brain ischemia was used to block ischemia-induced NSC/NPC proliferation. We observed that ICV infusion of Ara-C caused a complete blockade of NSC/NPC proliferation in the SVZ and a dramatic reduction of NSC/NPC proliferation in the SGZ. Additionally, as a result of the inhibition of ischemia-induced NSC/NPC pool amplification, the number of neurons in the hippocampal CA1 and CA3 was significantly reduced, the infarction size was significantly enlarged, and neurological deficits were significantly worsened after focal brain ischemia. We also found that an NSC/NPC-conditioned medium showed neuroprotective effects in vitro and that adult NSC/NPC-released brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) are required for NSC/NPC-conditioned medium-induced neuroprotection. These data suggest that NSC/NPC-generated trophic factors are neuroprotective and that brain ischemia-triggered NSC/NPC proliferation is crucial for brain protection. This study provides insights into the contribution of endogenous NSCs/NPCs to brain self-protection in adult brain after ischemia injury.

  4. Acetaminophen protects brain endothelial cells against oxidative stress.

    PubMed

    Tripathy, Debjani; Grammas, Paula

    2009-05-01

    Increasing evidence suggests that acetaminophen has unappreciated anti-oxidant and anti-inflammatory properties. Drugs that affect oxidant and inflammatory stress in the brain are of interest because both processes are thought to contribute to the pathogenesis of neurodegenerative disease. The objective of this study is to determine whether acetaminophen affects the response of brain endothelial cells to oxidative stress. Cultured brain endothelial cells are pre-treated with acetaminophen and then exposed to the superoxide-generating compound menadione (25 microM). Cell survival, inflammatory protein expression, and anti-oxidant enzyme activity are measured. Menadione causes a significant (p<0.001) increase in endothelial cell death as well as an increase in RNA and protein levels of tumor necrosis factor alpha, interleukin-1, macrophage inflammatory protein alpha, and RANTES. Menadione also evokes a significant (p<0.001) increase in the activity of the anti-oxidant enzyme superoxide dismutase (SOD). Pre-treatment of endothelial cell cultures with acetaminophen (25-100 microM) increases endothelial cell survival and inhibits menadione-induced expression of inflammatory proteins and SOD activity. In addition, we document, for the first time, that acetaminophen increases expression of the anti-apoptotic protein Bcl2. Suppressing Bcl2 with siRNA blocks the pro-survival effect of acetaminophen. These data show that acetaminophen has anti-oxidant and anti-inflammatory effects on the cerebrovasculature and suggest a heretofore unappreciated therapeutic potential for this drug in neurodegenerative diseases such as Alzheimer's disease that are characterized by oxidant and inflammatory stress.

  5. Bone marrow-derived stem cell therapy for metastatic brain cancers.

    PubMed

    Kaneko, Yuji; Tajiri, Naoki; Staples, Meaghan; Reyes, Stephanny; Lozano, Diego; Sanberg, Paul R; Freeman, Thomas B; van Loveren, Harry; Kim, Seung U; Borlongan, Cesar V

    2015-01-01

    We propose that stem cell therapy may be a potent treatment for metastatic melanoma in the brain. Here we discuss the key role of a leaky blood-brain barrier (BBB) that accompanies the development of brain metastases. We review the need to characterize the immunological and inflammatory responses associated with tumor-derived BBB damage in order to reveal the contribution of this brain pathological alteration to the formation and growth of brain metastatic cancers. Next, we discuss the potential repair of the BBB and attenuation of brain metastasis through transplantation of bone marrow-derived mesenchymal stem cells with the endothelial progenitor cell phenotype. In particular, we review the need for evaluation of the efficacy of stem cell therapy in repairing a disrupted BBB in an effort to reduce neuroinflammation, eventually attenuating brain metastatic cancers. The demonstration of BBB repair through augmented angiogenesis and vasculogenesis will be critical to establishing the potential of stem cell therapy for the treatment/prevention of metastatic brain tumors. The overarching hypothesis we advanced here is that BBB breakdown is closely associated with brain metastatic cancers of melanoma, exacerbating the inflammatory response of the brain during metastasis, and ultimately worsening the outcome of metastatic brain cancers. Abrogating this leaky BBB-mediated inflammation via stem cell therapy represents a paradigm-shifting approach to treating brain cancer. This review article discusses the pros and cons of cell therapy for melanoma brain metastases.

  6. Stem Cells for Ischemic Brain Injury: A Critical Review

    PubMed Central

    Burns, Terry C.; Verfaillie, Catherine M.; Low, Walter C.

    2014-01-01

    No effective therapy is currently available to promote recovery following ischemic stroke. Stem cells have been proposed as a potential source of new cells to replace those lost due to central nervous system injury, as well as a source of trophic molecules to minimize damage and promote recovery. We undertook a detailed review of data from recent basic science and preclinical studies to investigate the potential application of endogenous and exogenous stem cell therapies for treatment of cerebral ischemia. To date, spontaneous endogenous neurogenesis has been observed in response to ischemic injury, and can be enhanced via infusion of appropriate cytokines. Exogenous stem cells from multiple sources can generate neural cells that survive and form synaptic connections after transplantation in the stroke-injured brain. Stem cells from multiple sources cells also exhibit neuroprotective properties that may ameliorate stroke deficits. In many cases, functional benefits observed are likely independent of neural differentiation, though exact mechanisms remain poorly understood. Future studies of neuroregeneration will require the demonstration of function in endogenously born neurons following focal ischemia. Further, methods are currently lacking to definitively demonstrate the therapeutic effect of newly introduced neural cells. Increased plasticity following stroke may facilitate the functional integration of new neurons, but the loss of appropriate guidance cues and supporting architecture in the infarct cavity will likely impede the restoration of lost circuitry. As such careful investigation of the mechanisms underlying trophic benefits will be essential. Evidence to date suggest that continued development of stem cell therapies may ultimately lead to viable treatment options for ischemic brain injury. PMID:19399885

  7. The effects of preservation procedures on amniotic membrane's ability to serve as a substrate for cultivation of endothelial cells.

    PubMed

    Niknejad, Hassan; Deihim, Tina; Solati-Hashjin, Mehran; Peirovi, Habibollah

    2011-12-01

    Amniotic membrane (AM) has been used as a scaffold for the ex vivo expansion of different types of cells and a cell delivery matrix in regenerative medicine. Since the preservation procedures can influence the AM properties for experimental and clinical purposes, this study was established to investigate the feasibility of using the AM after different preservation methods to serve as substrates for endothelial cell expansion ex vivo. The effects of cryopreservation and lyophilization were evaluated on mechanical and histological characteristics of the AM, and the results were compared with the fresh AM. The ECM components of the basement membrane were well conserved in all groups. Although lyophilization resulted in more histological changes and lower level of physical variables including thickness, F(max), elongation at break and suture retention than the fresh and cryopreserved AM, endothelial cells grown on the lyophilized AM were better attached to the basement membrane. Cytotoxicity assay by MTT showed that the lyophilized AM is a compatible substrate for endothelial cells cultivation. The findings of this study suggest that the lyophilized AM is a suitable matrix for cultivation of endothelial cells due to this fact that lyophilization led to exposure of basement membrane of the AM.

  8. The role of maintenance proteins in the preservation of epithelial cell identity during mammary gland remodeling and breast cancer initiation.

    PubMed

    Coradini, Danila; Oriana, Saro

    2014-02-01

    During normal postnatal mammary gland development and adult remodeling related to the menstrual cycle, pregnancy, and lactation, ovarian hormones and peptide growth factors contribute to the delineation of a definite epithelial cell identity. This identity is maintained during cell replication in a heritable but DNA-independent manner. The preservation of cell identity is fundamental, especially when cells must undergo changes in response to intrinsic and extrinsic signals. The maintenance proteins, which are required for cell identity preservation, act epigenetically by regulating gene expression through DNA methylation, histone modification, and chromatin remodeling. Among the maintenance proteins, the Trithorax (TrxG) and Polycomb (PcG) group proteins are the best characterized. In this review, we summarize the structures and activities of the TrxG and PcG complexes and describe their pivotal roles in nuclear estrogen receptor activity. In addition, we provide evidence that perturbations in these epigenetic regulators are involved in disrupting epithelial cell identity, mammary gland remodeling, and breast cancer initiation.

  9. Brain microvascular endothelial cell transplantation ameliorates ischemic white matter damage.

    PubMed

    Puentes, Sandra; Kurachi, Masashi; Shibasaki, Koji; Naruse, Masae; Yoshimoto, Yuhei; Mikuni, Masahiko; Imai, Hideaki; Ishizaki, Yasuki

    2012-08-21

    Ischemic insults affecting the internal capsule result in sensory-motor disabilities which adversely affect the patient's life. Cerebral endothelial cells have been reported to exert a protective effect against brain damage, so the transplantation of healthy endothelial cells might have a beneficial effect on the outcome of ischemic brain damage. In this study, endothelin-1 (ET-1) was injected into the rat internal capsule to induce lacunar infarction. Seven days after ET-1 injection, microvascular endothelial cells (MVECs) were transplanted into the internal capsule. Meningeal cells or 0.2% bovine serum albumin-Hank's balanced salt solution were injected as controls. Two weeks later, the footprint test and histochemical analysis were performed. We found that MVEC transplantation improved the behavioral outcome based on recovery of hind-limb rotation angle (P<0.01) and induced remyelination (P<0.01) compared with the control groups. Also the inflammatory response was repressed by MVEC transplantation, judging from fewer ED-1-positive activated microglial cells in the MVEC-transplanted group than in the other groups. Elucidation of the mechanisms by which MVECs ameliorate ischemic damage of the white matter may provide important information for the development of effective therapies for white matter ischemia.

  10. Nonlinear Dynamic Theory of Acute Cell Injuries and Brain Ischemia

    NASA Astrophysics Data System (ADS)

    Taha, Doaa; Anggraini, Fika; Degracia, Donald; Huang, Zhi-Feng

    2015-03-01

    Cerebral ischemia in the form of stroke and cardiac arrest brain damage affect over 1 million people per year in the USA alone. In spite of close to 200 clinical trials and decades of research, there are no treatments to stop post-ischemic neuron death. We have argued that a major weakness of current brain ischemia research is lack of a deductive theoretical framework of acute cell injury to guide empirical studies. A previously published autonomous model based on the concept of nonlinear dynamic network was shown to capture important facets of cell injury, linking the concept of therapeutic to bistable dynamics. Here we present an improved, non-autonomous formulation of the nonlinear dynamic model of cell injury that allows multiple acute injuries over time, thereby allowing simulations of both therapeutic treatment and preconditioning. Our results are connected to the experimental data of gene expression and proteomics of neuron cells. Importantly, this new model may be construed as a novel approach to pharmacodynamics of acute cell injury. The model makes explicit that any pro-survival therapy is always a form of sub-lethal injury. This insight is expected to widely influence treatment of acute injury conditions that have defied successful treatment to date. This work is supported by NIH NINDS (NS081347) and Wayne State University President's Research Enhancement Award.

  11. Amantadine preserves dopamine level and attenuates depression-like behavior induced by traumatic brain injury in rats.

    PubMed

    Tan, Liang; Ge, Hongfei; Tang, Jun; Fu, Chuhua; Duanmu, Wangsheng; Chen, Yujie; Hu, Rong; Sui, Jianfeng; Liu, Xin; Feng, Hua

    2015-02-15

    Traumatic brain injury (TBI) often results in multiple neuropsychiatric sequelae, including cognitive, emotional, and behavioral problems. Among them, depression is a common psychiatric symptom, and links to poorer recovery. Amantadine, as an antiparkinsonian, increases dopamine release, and blocks dopamine reuptake, but has recently received attention for its effectiveness as an antidepressant. In the present study, we first induced a post-TBI depression rat model to probe the efficacy of amantadine therapy in reducing post-TBI depression. The DA concentration in the striatum of the injured rats, as well as the degeneration and apoptosis of dopaminergic neurons in the substantia nigra (SN), were checked along with the depression-like behavior. The results showed that amantadine therapy could significantly ameliorate the depression-like behavior, improving the DA level in the striatum and decreasing the degeneration and apoptosis of dopaminergic neurons in the SN. The results indicated that the anti-depression effect may result from the increase of extracellular DA concentration in the striatum and/or the indirect neuroprotection on the dopaminergic neurons in the SN. We conclude that DA plays a critical role in post-TBI depression, and that amantadine shows its potential value in anti-depression treatment for TBI.

  12. The dance of the perivascular and endothelial cells: mechanisms of brain response to immune signaling.

    PubMed

    Saper, Clifford B

    2010-01-14

    The mechanisms underlying the brain response to systemic inflammation remain unclear. In this issue of Neuron, Serrats and colleagues demonstrate that two cell types that produce prostaglandins that act on the brain, perivascular and endothelial cells, have an unexpectedly complex interaction in regulating the timing and types of brain responses that occur.

  13. Anticancer and antioxidant properties of terpinolene in rat brain cells.

    PubMed

    Aydin, Elanur; Türkez, Hasan; Taşdemir, Sener

    2013-09-01

    Terpinolene (TPO) is a natural monoterpene present in essential oils of many aromatic plant species. Although various biological activities of TPO have been demonstrated, its neurotoxicity has never been explored. In this in vitro study we investigated TPO's antiproliferative and/or cytotoxic properties using the 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) test, genotoxic damage potential using the single-cell gel electrophoresis (SCGE), and oxidative effects through total antioxidant capacity (TAC) and total oxidative stress (TOS) in cultured primary rat neurons and N2a neuroblastoma cells. Dose-dependent effects of TPO (at 10 mg L(-1), 25 mg L(-1), 50 mg L(-1), 100 mg L(-1), 200 mg L(-1), and 400 mg L(-1)) were tested in both cell types. Significant (P<0.05) decrease in cell proliferation were observed in cultured primary rat neurons starting with the dose of 100 mg L(-1) and in N2a neuroblastoma cells starting with 50 mg L(-1). TPO was not genotoxic in either cell type. In addition, TPO treatment at 10 mg L(-1), 25 mg L(-1), and 50 mg L(-1) increased TAC in primary rat neurons, but not in N2a cells. However, at concentrations above 50 mg L(-1) it increased TOS in both cell types. Our findings clearly demonstrate that TPO is a potent antiproliferative agent for brain tumour cells and may have potential as an anticancer agent, which needs to be further studied.

  14. Neutral amino acid transport across brain microvessel endothelial cell monolayers

    SciTech Connect

    Audus, K.L.; Borchardt, R.T.

    1986-03-01

    Brain microvessel endothelial cells (BMEC) which form the blood-brain barrier (BBB) possess an amino acid carrier specific for large neutral amino acids (LNAA). The carrier is important for facilitating the delivery of nutrient LNAA's and centrally acting drugs that are LNAA's, to the brain. Bovine BMEC's were isolated and grown up to complete monolayers on regenerated cellulose-membranes in primary culture. To study the transendothelial transport of leucine, the monolayers were placed in a side-by-side diffusion cell, and transport across the monolayers followed with (/sup 3/H)-leucine. The transendothelial transport of leucine in this in vitro model was determined to be bidirectional, and time-, temperature-, and concentration-dependent. The transport of leucine was saturable and the apparent K/sub m/ and V/sub max/, 0.18 mM and 6.3 nmol/mg/min, respectively. Other LNAA's, including the centrally acting drugs, ..cap alpha..-methyldopa, L-DOPA, ..cap alpha..-methyl-tyrosine, and baclofen, inhibited leucine transport. The leucine carrier was also found to be stereospecific and not sensitive to inhibitors of active transport. These results are consistent with previous in vitro and in vivo studies. Primary cultures of BMEC's appear to be a potentially important tool for investigating at the cellular level, the transport mechanisms of the BBB.

  15. Scanning electron microscopic analysis of endothelial cell coverage and quality in large vessels from multi-organ donors: effects of preservation on endothelial cell integrity.

    PubMed

    van Leeuwen, E B; Molema, G; van Luyn, M J; de Jong, K P; Dijk, F; Slooff, M J; Ruiters, M H; van der Meer, J

    2000-06-01

    Endothelial cell integrity (coverage and quality) of large donor vessels is important because these vessels are used for vascular reconstructions in solid-organ transplantation. Disruption of the endothelial cell monolayer will initiate blood coagulation and may lead to thrombosis of large vessels, often resulting in the loss of the transplanted organ. Iliac arteries and veins, removed from 10 heart-beating multi-organ donors at the end of the donor procedure, were analyzed using scanning electron microscopy at three different time points of preservation. Endothelial cell coverage and quality were determined immediately after removal from the donor, after 10 h (time of transplantation) and 7 d storage in 'University of Wisconsin' cold preservation solution (UW). Endothelial cell coverage decreased during the preservation of arteries, but was maintained in veins. Storage of the veins for 7 d in plastic bags showed a decreased endothelial cell coverage compared to storage in glass vials. Early removal of the blood vessels and proper storage, free floating and in clean UW, may improve maintenance of the endothelial cell integrity. These findings may be important in order to reduce the risk of thrombosis and, consequently, organ failure after transplantation. Furthermore, vessels with maintained endothelial cell integrity after 7 d may be used for in vitro research.

  16. Intermittent Hypoxia and Stem Cell Implants Preserve Breathing Capacity in a Rodent Model of Amyotrophic Lateral Sclerosis

    PubMed Central

    Nichols, Nicole L.; Gowing, Genevieve; Satriotomo, Irawan; Nashold, Lisa J.; Dale, Erica A.; Suzuki, Masatoshi; Avalos, Pablo; Mulcrone, Patrick L.; McHugh, Jacalyn

    2013-01-01

    Rationale: Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease causing paralysis and death from respiratory failure. Strategies to preserve and/or restore respiratory function are critical for successful treatment. Although breathing capacity is maintained until late in disease progression in rodent models of familial ALS (SOD1G93A rats and mice), reduced numbers of phrenic motor neurons and decreased phrenic nerve activity are observed. Decreased phrenic motor output suggests imminent respiratory failure. Objectives: To preserve or restore phrenic nerve activity in SOD1G93A rats at disease end stage. Methods: SOD1G93A rats were injected with human neural progenitor cells (hNPCs) bracketing the phrenic motor nucleus before disease onset, or exposed to acute intermittent hypoxia (AIH) at disease end stage. Measurements and Main Results: The capacity to generate phrenic motor output in anesthetized rats at disease end stage was: (1) transiently restored by a single presentation of AIH; and (2) preserved ipsilateral to hNPC transplants made before disease onset. hNPC transplants improved ipsilateral phrenic motor neuron survival. Conclusions: AIH-induced respiratory plasticity and stem cell therapy have complementary translational potential to treat breathing deficits in patients with ALS. PMID:23220913

  17. Serpins promote cancer cell survival and vascular co-option in brain metastasis.

    PubMed

    Valiente, Manuel; Obenauf, Anna C; Jin, Xin; Chen, Qing; Zhang, Xiang H-F; Lee, Derek J; Chaft, Jamie E; Kris, Mark G; Huse, Jason T; Brogi, Edi; Massagué, Joan

    2014-02-27

    Brain metastasis is an ominous complication of cancer, yet most cancer cells that infiltrate the brain die of unknown causes. Here, we identify plasmin from the reactive brain stroma as a defense against metastatic invasion, and plasminogen activator (PA) inhibitory serpins in cancer cells as a shield against this defense. Plasmin suppresses brain metastasis in two ways: by converting membrane-bound astrocytic FasL into a paracrine death signal for cancer cells, and by inactivating the axon pathfinding molecule L1CAM, which metastatic cells express for spreading along brain capillaries and for metastatic outgrowth. Brain metastatic cells from lung cancer and breast cancer express high levels of anti-PA serpins, including neuroserpin and serpin B2, to prevent plasmin generation and its metastasis-suppressive effects. By protecting cancer cells from death signals and fostering vascular co-option, anti-PA serpins provide a unifying mechanism for the initiation of brain metastasis in lung and breast cancers.

  18. Transient oxygen-glucose deprivation sensitizes brain capillary endothelial cells to rtPA at 4h of reoxygenation.

    PubMed

    Kuntz, Mélanie; Mysiorek, Caroline; Pétrault, Olivier; Boucau, Marie-Christine; Aijjou, Rachid; Uzbekov, Rustem; Bérézowski, Vincent

    2014-01-01

    Thrombolysis treatment of acute ischemic stroke is limited by the pro-edematous and hemorrhagic effects exerted by reperfusion, which disrupts the blood-brain barrier (BBB) capillary endothelium in the infarct core. Most studies of the ischemic BBB overlook the complexity of the penumbral area, where the affected brain cells are still viable following deprivation. Our present objective was to examine in vitro the kinetic impact of reoxygenation on the integrity of ischemic BBB cells after oxygen-glucose deprivation. Through the use of a co-culture of brain capillary endothelial cells and glial cells, we first showed that the transendothelial permeability increase induced by deprivation can occur with both preserved cell viability and interendothelial tight junction network. The subtle and heterogeneous alteration of the tight junctions was observable only through electron microscopy. A complete permeability recovery was then found after reoxygenation, when Vimentin and Actin networks were reordered. However, still sparse ultrastructural alterations of tight junctions suggested an acquired vulnerability. Endothelial cells were then exposed to recombinant tissue-type plasminogen activator (rtPA) to define a temporal profile for the toxic effect of this thrombolytic on transendothelial permeability. Interestingly, the reoxygenated BBB broke down with aggravated tight junction disruption when exposed to rtPA only at 4h after reoxygenation. Moreover, this breakdown was enhanced by 50% when ischemic glial cells were present during the first hours of reoxygenation. Our results suggest that post-stroke reoxygenation enables retrieval of the barrier function of brain capillary endothelium when in a non-necrotic environment, but may sensitize it to rtPA at the 4-hour time point, when both endothelial breakdown mechanisms and glial secretions could be identified and targeted in a therapeutical perspective.

  19. Mesenchymal Stem Cells Preserve Working Memory in the 3xTg-AD Mouse Model of Alzheimer's Disease.

    PubMed

    Ruzicka, Jiri; Kulijewicz-Nawrot, Magdalena; Rodrigez-Arellano, Jose Julio; Jendelova, Pavla; Sykova, Eva

    2016-01-25

    The transplantation of stem cells may have a therapeutic effect on the pathogenesis and progression of neurodegenerative disorders. In the present study, we transplanted human mesenchymal stem cells (MSCs) into the lateral ventricle of a triple transgenic mouse model of Alzheimer's disease (3xTg-AD) at the age of eight months. We evaluated spatial reference and working memory after MSC treatment and the possible underlying mechanisms, such as the influence of transplanted MSCs on neurogenesis in the subventricular zone (SVZ) and the expression levels of a 56 kDa oligomer of amyloid β (Aβ*56), glutamine synthetase (GS) and glutamate transporters (Glutamate aspartate transporter (GLAST) and Glutamate transporter-1 (GLT-1)) in the entorhinal and prefrontal cortices and the hippocampus. At 14 months of age we observed the preservation of working memory in MSC-treated 3xTg-AD mice, suggesting that such preservation might be due to the protective effect of MSCs on GS levels and the considerable downregulation of Aβ*56 levels in the entorhinal cortex. These changes were observed six months after transplantation, accompanied by clusters of proliferating cells in the SVZ. Since the grafted cells did not survive for the whole experimental period, it is likely that the observed effects could have been transiently more pronounced at earlier time points than at six months after cell application.

  20. Induction of Brain Microvascular Endothelial Cell Urokinase Expression by Cryptococcus neoformans Facilitates Blood-Brain Barrier Invasion

    PubMed Central

    Stie, Jamal; Fox, Deborah

    2012-01-01

    The invasive ability of the blood-borne fungal pathogen Cryptococcus neoformans can be enhanced through interactions with host plasma components, such as plasminogen. Previously we showed by in vitro studies that plasminogen coats the surface of C. neoformans and is converted to the active serine protease, plasmin, by host plasminogen activators. Viable, but not formaldehyde- or sodium azide-killed, cryptococcal strains undergo brain microvascular endothelial cell-dependent plasminogen-to-plasmin activation, which results in enhanced, plasmin-dependent cryptococcal invasion of primary bovine brain microvascular endothelial cells and fungal ability to degrade plasmin substrates. In the present work, brain microvascular endothelial cells cultured with viable, but not killed, cryptococcal strains led to significant increases in both urokinase mRNA transcription and cell-associated urokinase protein expression. Soluble urokinase was also detected in conditioned medium from brain microvascular endothelial cells cultured with viable, but not killed, C. neoformans. Exposure of plasminogen pre-coated viable C. neoformans to conditioned medium from strain-matched brain microvascular endothelial cell-fungal co-cultures resulted in plasminogen-to-plasmin activation and plasmin-dependent cryptococcal invasion. siRNA-mediated silencing of urokinase gene expression or the use of specific inhibitors of urokinase activity abrogated both plasminogen-to-plasmin activation on C. neoformans and cryptococcal-brain microvascular endothelial cell invasion. Our results suggest that pathogen exploitation of the host urokinase-plasmin(ogen) system may contribute to C. neoformans virulence during invasive cryptococcosis. PMID:23145170

  1. Melanoma Cells Homing to the Brain: An In Vitro Model

    PubMed Central

    Rizzo, A.; Vasco, C.; Girgenti, V.; Fugnanesi, V.; Calatozzolo, C.; Canazza, A.; Salmaggi, A.; Rivoltini, L.; Morbin, M.; Ciusani, E.

    2015-01-01

    We developed an in vitro contact through-feet blood brain barrier (BBB) model built using type IV collagen, rat astrocytes, and human umbilical vein endothelial cells (HUVECs) cocultured through Transwell porous polycarbonate membrane. The contact between astrocytes and HUVECs was demonstrated by electron microscopy: astrocytes endfeet pass through the 8.0 μm pores inducing HUVECs to assume a cerebral phenotype. Using this model we evaluated transmigration of melanoma cells from two different patients (M1 and M2) selected among seven melanoma primary cultures. M2 cells showed a statistically significant higher capability to pass across the in vitro BBB model, compared to M1. Expression of adhesion molecules was evaluated by flow cytometry: a statistically significant increased expression of MCAM, αvβ3, and CD49b was detected in M1. PCR array data showed that M2 had a higher expression of several matrix metalloproteinase proteins (MMPs) compared to M1. Specifically, data suggest that MMP2 and MMP9 could be directly involved in BBB permeability and that brain invasion by melanoma cells could be related to the overexpression of many MMPs. Future studies will be necessary to deepen the mechanisms of central nervous system invasion. PMID:25692137

  2. Preservation of high glycolytic phenotype by establishing new acute lymphoblastic leukemia cell lines at physiologic oxygen concentration

    SciTech Connect

    Sheard, Michael A.; Ghent, Matthew V.; Cabral, Daniel J.; Lee, Joanne C.; Khankaldyyan, Vazgen; Ji, Lingyun; Wu, Samuel Q.; Kang, Min H.; and others

    2015-05-15

    Cancer cells typically exhibit increased glycolysis and decreased mitochondrial oxidative phosphorylation, and they continue to exhibit some elevation in glycolysis even under aerobic conditions. However, it is unclear whether cancer cell lines employ a high level of glycolysis comparable to that of the original cancers from which they were derived, even if their culture conditions are changed to physiologically relevant oxygen concentrations. From three childhood acute lymphoblastic leukemia (ALL) patients we established three new pairs of cell lines in both atmospheric (20%) and physiologic (bone marrow level, 5%) oxygen concentrations. Cell lines established in 20% oxygen exhibited lower proliferation, survival, expression of glycolysis genes, glucose consumption, and lactate production. Interestingly, the effects of oxygen concentration used during cell line initiation were only partially reversible when established cell cultures were switched from one oxygen concentration to another for eight weeks. These observations indicate that ALL cell lines established at atmospheric oxygen concentration can exhibit relatively low levels of glycolysis and these levels are semi-permanent, suggesting that physiologic oxygen concentrations may be needed from the time of cell line initiation to preserve the high level of glycolysis commonly exhibited by leukemias in vivo. - Highlights: • Establishing new ALL cell lines in 5% oxygen resulted in higher glycolytic expression and function. • Establishing new ALL cell lines in 5% oxygen resulted in higher proliferation and lower cell death. • The divergent metabolic phenotypes selected in 5% and 20% oxygen are semi-permanent.

  3. Neural stem cells sustain natural killer cells that dictate recovery from brain inflammation

    PubMed Central

    Liu, Qiang; Sanai, Nader; Jin, Wei-Na; La Cava, Antonio; Van Kaer, Luc; Shi, Fu-Dong

    2017-01-01

    Recovery from organ-specific autoimmune diseases largely relies on the mobilization of endogenous repair mechanisms and local factors that control them. Natural killer (NK) cells are swiftly mobilized to organs targeted by autoimmunity and typically undergo numerical contraction when inflammation wanes. We report the unexpected finding that NK cells are retained in the brain subventricular zone (SVZ) during the chronic phase of multiple sclerosis in humans and its animal model in mice. These NK cells were found preferentially in close proximity to SVZ neural stem cells (NSCs) that produce interleukin-15 and sustain functionally competent NK cells. Moreover, NK cells limited the reparative capacity of NSCs following brain inflammation. These findings reveal that reciprocal interactions between NSCs and NK cells regulate neurorepair. PMID:26752157

  4. Isolation of primary murine brain microvascular endothelial cells.

    PubMed

    Ruck, Tobias; Bittner, Stefan; Epping, Lisa; Herrmann, Alexander M; Meuth, Sven G

    2014-11-14

    The blood-brain-barrier is ultrastructurally assembled by a monolayer of brain microvascular endothelial cells (BMEC) interconnected by a junctional complex of tight and adherens junctions. Together with other cell-types such as astrocytes or pericytes, they form the neurovascular unit (NVU), which specifically regulates the interchange of fluids, molecules and cells between the peripheral blood and the CNS. Through this complex and dynamic system BMECs are involved in various processes maintaining the homeostasis of the CNS. A dysfunction of the BBB is observed as an essential step in the pathogenesis of many severe CNS diseases. However, specific and targeted therapies are very limited, as the underlying mechanisms are still far from being understood. Animal and in vitro models have been extensively used to gain in-depth understanding of complex physiological and pathophysiological processes. By reduction and simplification it is possible to focus the investigation on the subject of interest and to exclude a variety of confounding factors. However, comparability and transferability are also reduced in model systems, which have to be taken into account for evaluation. The most common animal models are based on mice, among other reasons, mainly due to the constantly increasing possibilities of methodology. In vitro studies of isolated murine BMECs might enable an in-depth analysis of their properties and of the blood-brain-barrier under physiological and pathophysiological conditions. Further insights into the complex mechanisms at the BBB potentially provide the basis for new therapeutic strategies. This protocol describes a method to isolate primary murine microvascular endothelial cells by a sequence of physical and chemical purification steps. Special considerations for purity and cultivation of MBMECs as well as quality control, potential applications and limitations are discussed.

  5. A Child's Brain. Part II. The Human Brain: How Every Single Cell is Organized for Action.

    ERIC Educational Resources Information Center

    Sylwester, Robert

    1982-01-01

    The second in a series of three articles concerning children's brain development focuses on the organization of the brain. Aspects of the brain's vertical, neocortex, and temporal organization are discussed and references for further reading are provided. (CJ)

  6. Brain Invasion by Mouse Hepatitis Virus Depends on Impairment of Tight Junctions and Beta Interferon Production in Brain Microvascular Endothelial Cells

    PubMed Central

    Bleau, Christian; Filliol, Aveline; Samson, Michel

    2015-01-01

    ABSTRACT Coronaviruses (CoVs) have shown neuroinvasive properties in humans and animals secondary to replication in peripheral organs, but the mechanism of neuroinvasion is unknown. The major aim of our work was to evaluate the ability of CoVs to enter the central nervous system (CNS) through the blood-brain barrier (BBB). Using the highly hepatotropic mouse hepatitis virus type 3 (MHV3), its attenuated variant, 51.6-MHV3, which shows low tropism for endothelial cells, and the weakly hepatotropic MHV-A59 strain from the murine coronavirus group, we investigated the virus-induced dysfunctions of BBB in vivo and in brain microvascular endothelial cells (BMECs) in vitro. We report here a MHV strain-specific ability to cross the BBB during acute infection according to their virulence for liver. Brain invasion was observed only in MHV3-infected mice and correlated with enhanced BBB permeability associated with decreased expression of zona occludens protein 1 (ZO-1), VE-cadherin, and occludin, but not claudin-5, in the brain or in cultured BMECs. BBB breakdown in MHV3 infection was not related to production of barrier-dysregulating inflammatory cytokines or chemokines by infected BMECs but rather to a downregulation of barrier protective beta interferon (IFN-β) production. Our findings highlight the importance of IFN-β production by infected BMECs in preserving BBB function and preventing access of blood-borne infectious viruses to the brain. IMPORTANCE Coronaviruses (CoVs) infect several mammals, including humans, and are associated with respiratory, gastrointestinal, and/or neurological diseases. There is some evidence that suggest that human respiratory CoVs may show neuroinvasive properties. Indeed, the severe acute respiratory syndrome coronavirus (SARS-CoV), causing severe acute respiratory syndrome, and the CoVs OC43 and 229E were found in the brains of SARS patients and multiple sclerosis patients, respectively. These findings suggest that hematogenously spread

  7. Neural Stem Cell Delivery of Therapeutic Antibodies to Treat Breast Cancer Brain Metastases

    DTIC Science & Technology

    2010-10-01

    cell numbers, and brain metastatic growth was monitored by non-invasive bioluminescence imaging . Figure 5 shows that, surprisingly, the ALDH1 negative...the brains of SCID mice and lesion growth followed by non- invasive bioluminescence imaging over time (x-axis in the graph denotes days after...mouse brain. Histological analysis of brain lesions 60 days after implanting 500 tumor cells of each subpopulation. Each image represents one mouse

  8. Characterization of brain cell nuclei with decondensed chromatin.

    PubMed

    Yu, Ping; McKinney, Elizabeth C; Kandasamy, Muthugapatti M; Albert, Alexandria L; Meagher, Richard B

    2015-07-01

    Although multipotent cell types have enlarged nuclei with decondensed chromatin, this property has not been exploited to enhance the characterization of neural progenitor cell (NPC) populations in the brain. We found that mouse brain cell nuclei that expressed exceptionally high levels of the pan neuronal marker NeuN/FOX3 (NeuN-High) had decondensed chromatin relative to most NeuN-Low or NeuN-Neg (negative) nuclei. Purified NeuN-High nuclei expressed significantly higher levels of transcripts encoding markers of neurogenesis, neuroplasticity, and learning and memory (ARC, BDNF, ERG1, HOMER1, NFL/NEF1, SYT1), subunits of chromatin modifying machinery (SIRT1, HDAC1, HDAC2, HDAC11, KAT2B, KAT3A, KAT3B, KAT5, DMNT1, DNMT3A, Gadd45a, Gadd45b) and markers of NPC and cell cycle activity (BRN2, FOXG1, KLF4, c-MYC, OCT4, PCNA, SHH, SOX2) relative to neuronal NeuN-Low or to mostly non-neuronal NeuN-Neg nuclei. NeuN-High nuclei expressed higher levels of HDAC1, 2, 4, and 5 proteins. The cortex, hippocampus, hypothalamus, thalamus, and nucleus accumbens contained high percentages of large decondensed NeuN-High nuclei, while the cerebellum, and pons contained very few. NeuN-High nuclei have the properties consistent with their being derived from extremely active neurons with elevated rates of chromatin modification and/or NPC-like cells with multilineage developmental potential. The further analysis of decondensed neural cell nuclei should provide novel insights into neurobiology and neurodegenerative disease.

  9. Preservation of perceptual integration improves temporal stability of bimanual coordination in the elderly: an evidence of age-related brain plasticity.

    PubMed

    Blais, Mélody; Martin, Elodie; Albaret, Jean-Michel; Tallet, Jessica

    2014-12-15

    Despite the apparent age-related decline in perceptual-motor performance, recent studies suggest that the elderly people can improve their reaction time when relevant sensory information are available. However, little is known about which sensory information may improve motor behaviour itself. Using a synchronization task, the present study investigates how visual and/or auditory stimulations could increase accuracy and stability of three bimanual coordination modes produced by elderly and young adults. Neurophysiological activations are recorded with ElectroEncephaloGraphy (EEG) to explore neural mechanisms underlying behavioural effects. Results reveal that the elderly stabilize all coordination modes when auditory or audio-visual stimulations are available, compared to visual stimulation alone. This suggests that auditory stimulations are sufficient to improve temporal stability of rhythmic coordination, even more in the elderly. This behavioural effect is primarily associated with increased attentional and sensorimotor-related neural activations in the elderly but similar perceptual-related activations in elderly and young adults. This suggests that, despite a degradation of attentional and sensorimotor neural processes, perceptual integration of auditory stimulations is preserved in the elderly. These results suggest that perceptual-related brain plasticity is, at least partially, conserved in normal aging.

  10. Evaluation of zinc salt based fixatives for preserving antigenic determinants for immunohistochemical demonstration of murine immune system cell markers.

    PubMed

    Hicks, D J; Johnson, L; Mitchell, S M; Gough, J; Cooley, W A; La Ragione, R M; Spencer, Y I; Wangoo, A

    2006-01-01

    Conventional aldehyde based fixatives produce good morphological preservation. However, owing to their cross-linking mechanism of action, epitope loss may occur during fixation compromising the tissue for subsequent immunohistochemical (IHC) analysis. IHC is an important tool for characterizing antigen, cytokine and cytomorphological markers. The increasing use of mouse models for study of pathogenesis has highlighted the need to investigate alternative fixatives. In the study reported here, tissue samples from RIII mice with immune mediated lesions, Mycobacterium bovis infected mice, and uninfected control mice were fixed in either zinc salt fixative or buffered formalin, then tested for IHC using a panel of antibodies (CD3, CD4, CD8, CD45, CD54, F4/80, Interferon-gamma and MIP2). Zinc salt fixation preserved processing-sensitive murine cell markers (CD4, CD8 and CD54) and improved the intensity of immunolabeling for CD45, F4/80 and CD3. Buffered formalin failed to preserve any of the processing-sensitive murine epitopes for demonstration by subsequent IHC.

  11. Asymptotic-Preserving Particle-In-Cell methods for the Vlasov-Maxwell system in the quasi-neutral limit

    NASA Astrophysics Data System (ADS)

    Degond, P.; Deluzet, F.; Doyen, D.

    2017-02-01

    In this article, we design Asymptotic-Preserving Particle-In-Cell methods for the Vlasov-Maxwell system in the quasi-neutral limit, this limit being characterized by a Debye length negligible compared to the space scale of the problem. These methods are consistent discretizations of the Vlasov-Maxwell system which, in the quasi-neutral limit, remain stable and are consistent with a quasi-neutral model (in this quasi-neutral model, the electric field is computed by means of a generalized Ohm law). The derivation of Asymptotic-Preserving methods is not straightforward since the quasi-neutral model is a singular limit of the Vlasov-Maxwell model. The key step is a reformulation of the Vlasov-Maxwell system which unifies the two models in a single set of equations with a smooth transition from one to another. As demonstrated in various and demanding numerical simulations, the Asymptotic-Preserving methods are able to treat efficiently both quasi-neutral plasmas and non-neutral plasmas, making them particularly well suited for complex problems involving dense plasmas with localized non-neutral regions.

  12. Plumbagin alters telomere dynamics, induces DNA damage and cell death in human brain tumour cells.

    PubMed

    Khaw, Aik Kia; Sameni, Safoura; Venkatesan, Shriram; Kalthur, Guruprasad; Hande, M Prakash

    2015-11-01

    Natural plant products may possess much potential in palliative therapy and supportive strategies of current cancer treatments with lesser cytotoxicity to normal cells compared to conventional chemotherapy. In the current study, anti-cancer properties of plumbagin, a plant-derived naphthoquinone, on brain cancer cells were determined. Plumbagin treatment resulted in the induction of DNA damage, cell cycle arrest and apoptosis, followed by suppression of the colony forming ability of the brain tumour cells. These effects were substantiated by upregulation of PTEN, TNFRSF1A and downregulation of E2F1 genes, along with a drop in MDM2, cyclin B1, survivin and BCL2 protein expression. Plumbagin induced elevated levels of caspase-3/7 activity as well. For the first time, we show here that plumbagin inhibits telomerase in brain tumour cells and results in telomere shortening following chronic long-term treatment. This observation implies considerable cytotoxicity of plumbagin towards cancer cells with higher telomerase activity. Collectively, our findings suggest plumbagin as a potential chemotherapeutic phytochemical in brain tumour treatment modalities.

  13. A glucose fuel cell for implantable brain-machine interfaces.

    PubMed

    Rapoport, Benjamin I; Kedzierski, Jakub T; Sarpeshkar, Rahul

    2012-01-01

    We have developed an implantable fuel cell that generates power through glucose oxidation, producing 3.4 μW cm(-2) steady-state power and up to 180 μW cm(-2) peak power. The fuel cell is manufactured using a novel approach, employing semiconductor fabrication techniques, and is therefore well suited for manufacture together with integrated circuits on a single silicon wafer. Thus, it can help enable implantable microelectronic systems with long-lifetime power sources that harvest energy from their surrounds. The fuel reactions are mediated by robust, solid state catalysts. Glucose is oxidized at the nanostructured surface of an activated platinum anode. Oxygen is reduced to water at the surface of a self-assembled network of single-walled carbon nanotubes, embedded in a Nafion film that forms the cathode and is exposed to the biological environment. The catalytic electrodes are separated by a Nafion membrane. The availability of fuel cell reactants, oxygen and glucose, only as a mixture in the physiologic environment, has traditionally posed a design challenge: Net current production requires oxidation and reduction to occur separately and selectively at the anode and cathode, respectively, to prevent electrochemical short circuits. Our fuel cell is configured in a half-open geometry that shields the anode while exposing the cathode, resulting in an oxygen gradient that strongly favors oxygen reduction at the cathode. Glucose reaches the shielded anode by diffusing through the nanotube mesh, which does not catalyze glucose oxidation, and the Nafion layers, which are permeable to small neutral and cationic species. We demonstrate computationally that the natural recirculation of cerebrospinal fluid around the human brain theoretically permits glucose energy harvesting at a rate on the order of at least 1 mW with no adverse physiologic effects. Low-power brain-machine interfaces can thus potentially benefit from having their implanted units powered or recharged by

  14. Immune cell trafficking from the brain maintains CNS immune tolerance.

    PubMed

    Mohammad, Mohammad G; Tsai, Vicky W W; Ruitenberg, Marc J; Hassanpour, Masoud; Li, Hui; Hart, Prue H; Breit, Samuel N; Sawchenko, Paul E; Brown, David A

    2014-03-01

    In the CNS, no pathway dedicated to immune surveillance has been characterized for preventing the anti-CNS immune responses that develop in autoimmune neuroinflammatory disease. Here, we identified a pathway for immune cells to traffic from the brain that is associated with the rostral migratory stream (RMS), which is a forebrain source of newly generated neurons. Evaluation of fluorescently labeled leukocyte migration in mice revealed that DCs travel via the RMS from the CNS to the cervical LNs (CxLNs), where they present antigen to T cells. Pharmacologic interruption of immune cell traffic with the mononuclear cell-sequestering drug fingolimod influenced anti-CNS T cell responses in the CxLNs and modulated experimental autoimmune encephalomyelitis (EAE) severity in a mouse model of multiple sclerosis (MS). Fingolimod treatment also induced EAE in a disease-resistant transgenic mouse strain by altering DC-mediated Treg functions in CxLNs and disrupting CNS immune tolerance. These data describe an immune cell pathway that originates in the CNS and is capable of dampening anti-CNS immune responses in the periphery. Furthermore, these data provide insight into how fingolimod treatment might exacerbate CNS neuroinflammation in some cases and suggest that focal therapeutic interventions, outside the CNS have the potential to selectively modify anti-CNS immunity.

  15. Methamphetamine is not Toxic but Disrupts the Cell Cycle of Blood-Brain Barrier Endothelial Cells.

    PubMed

    Fisher, D; Gamieldien, K; Mafunda, P S

    2015-07-01

    The cytotoxic effects of methamphetamine (MA) are well established to be caused via induced oxidative stress which in turn compromises the core function of the blood-brain barrier (BBB) by reducing its ability to regulate the homeostatic environment of the brain. While most studies were conducted over a period of 24-48 h, this study investigated the mechanisms by which chronic exposure of MA adversely affect the endothelial cells of BBB over an extended period of 96 h. MA induced significant depression of cell numbers at 96 h. This result was supported by flow cytometric data on the cell cycle which showed that brain endothelial cells (bEnd5) at 96 h were significantly suppressed in the S-phase of the cell cycle. In contrast, at 24-72 h control cell numbers for G1, S and G2-M phases were similar to MA-exposed cells. MA (0-1,000 µM) did not, however, statistically affect the viability and cytotoxicity of the bEnd5 cells, and the profile of ATP production and DNA synthesis (BrdU) across 96 h did not provide a rationale for the suppression of cell division. Our study reports for the first time that chronic exposure to MA results in long-term disruption of the cell cycle phases which eventuates in the attenuation of brain capillary endothelial cell growth after 96 h, compounding and contributing to the already well-known adverse short-term permeability effects of MA exposure on the BBB.

  16. Alkamides from Echinacea angustifolia Interact with P-glycoprotein of primary brain capillary endothelial cells isolated from porcine brain blood vessels.

    PubMed

    Mahringer, Anne; Ardjomand-Woelkart, Karin; Bauer, Rudolf; Fricker, Gert; Efferth, Thomas

    2013-03-01

    The blood-brain barrier prevents the passage of toxic compounds from blood circulation into brain tissue. Unfortunately, drugs for the treatment of neurodegenerative diseases, brain tumors, and other diseases also do not cross the blood-brain barrier. In the present investigation, we used isolated porcine brain capillary endothelial cells and a flow cytometric calcein-AM assay to analyze inhibition of P-glycoprotein, a major constituent of the blood-brain barrier. We tested 8 alkamides isolated from Echinacea angustifolia and found that four of them inhibited P-glycoprotein-mediated calcein transport in porcine brain capillary endothelial cells.

  17. Noninvasive photodetachment of stem cells on tunable conductive polymer nano thin films: selective harvesting and preserved differentiation capacity.

    PubMed

    You, Jungmok; Heo, June Seok; Kim, Jeonghun; Park, Teahoon; Kim, Byeonggwan; Kim, Han-Soo; Choi, Youjeong; Kim, Hyun Ok; Kim, Eunkyoung

    2013-05-28

    Viable mesenchymal stem cells (MSCs) were efficiently and selectively harvested by near-infrared (NIR) light using the photothermal effect of a conductive polymer nano thin film. The poly(3,4-ethylenedioxy thiophene) (PEDOT)-coated cell culture surfaces were prepared via a simple and fast solution-casting polymerization (SCP) technique. The absorption of PEDOT thin films in the NIR region was effectively triggered cell harvesting upon exposure to an NIR source. By controlling the NIR absorption of the PEDOT film through electrochemical doping or growing PEDOT with different thin film thickness from 70 to 300 nm, the proliferation and harvesting of MSCs on the PEDOT surface were controlled quantitatively. This light-induced cell detachment method based on PEDOT films provides the temporal and spatial control of cell harvesting, as well as cell patterning. The harvested stem cells were found to be alive and well proliferated despite the use of temperature increase by NIR. More importantly, the harvested MSCs by this method preserved their intrinsic characteristics as well as multilineage differentiation capacities. This PEDOT surfaces could be used for repetitive culture and detachment of MSCs or for efficient selection or depletion of a specific subset from heterogeneous population during culture of various tissue-derived cells because there were no photodegradation and photobreakage in the PEDOT films by NIR exposure.

  18. Purkinje cell vulnerability to mild traumatic brain injury.

    PubMed

    Fukuda, K; Aihara, N; Sagar, S M; Sharp, F R; Pitts, L H; Honkaniemi, J; Noble, L J

    1996-05-01

    In this study we examined the cerebellar response to mild traumatic brain injury by assessing microglial activation and Purkinje cell loss. Activated microglia were identified using the antibodies OX-42 and ED-1 as well as isolectin B4. The anti-Purkinje cell antibody PEP-19 was used to evaluate Purkinje cell loss after injury. The mechanism of cell injury was examined using a monoclonal antibody to the inducible 72-kDa heat shock protein. A monoclonal antibody to the N-terminal sequence of Fos was used as a marker for neuronal activation. There was progressive activation of microglia in the cerebellar vermis within a few days after forebrain injury. In coronal sections the processes of activated microglia were oriented in "stripes" perpendicular to the cortical surface. In sagittal sections the activated microglia were in irregularly shaped clusters or in a fan-like distribution that radiated from the Purkinje cell layer toward the cortical surface. There was a significant loss of Purkinje cells 7 days postinjury as compared to the control group. There was no evidence of induction of heat shock protein in the cerebellum. In addition, there was no evidence of induction of c-Fos protein in either the cerebellar cortex or inferior olivary nuclei within the first 3 h after injury. These studies demonstrate that a fluid percussive impact to the forebrain results in cerebellar damage. The close anatomical association between activated microglia and Purkinje cells suggests that Purkinje cell injury is the cause of the microglial activation. The mechanism of Purkinje cell death, however, remains unclear.

  19. Developmental and cell type-specific expression of thyroid hormone transporters in the mouse brain and in primary brain cells.

    PubMed

    Braun, Doreen; Kinne, Anita; Bräuer, Anja U; Sapin, Remy; Klein, Marc O; Köhrle, Josef; Wirth, Eva K; Schweizer, Ulrich

    2011-03-01

    Cellular thyroid hormone uptake and efflux are mediated by transmembrane transport proteins. One of these, monocarboxylate transporter 8 (MCT8) is mutated in Allan-Herndon-Dudley syndrome, a severe mental retardation associated with abnormal thyroid hormone constellations. Since mice deficient in Mct8 exhibit a milder neurological phenotype than patients, we hypothesized that alternative thyroid hormone transporters may compensate in murine brain cells for the lack of Mct8. Using qPCR, Western Blot, and immunocytochemistry, we investigated the expression of three different thyroid hormone transporters, i.e., Mct8 and L-type amino acid transporters Lat1 and Lat2, in mouse brain. All three thyroid hormone transporters are expressed from corticogenesis and peak around birth. Primary cultures of neurons and astrocytes express Mct8, Lat1, and Lat2. Microglia specifically expresses Mct10 and Slco4a1 in addition to high levels of Lat2 mRNA and protein. As in vivo, a brain microvascular endothelial cell line expressed Mct8 and Lat1. 158N, an oligodendroglial cell line expressed Mct8 protein, consistent with delayed myelination in MCT8-deficient patients. Functional T(3)- and T(4)-transport assays into primary astrocytes showed K(M) values of 4.2 and 3.7 μM for T(3) and T(4). Pharmacological inhibition of L-type amino acid transporters by BCH and genetic inactivation of Lat2 reduced astrocytic T(3) uptake to the same extent. BSP, a broad spectrum inhibitor, including Mct8, reduced T(3) uptake further suggesting the cooperative activity of several T(3) transporters in astrocytes.

  20. Ultrasound fails to induce proliferation of human brain and mouse endothelial cell lines

    NASA Astrophysics Data System (ADS)

    Rodemer, Claus; Jenne, Jürgen; Fatar, Marc; Hennerici, Michael G.; Meairs, Stephen

    2012-11-01

    Both in vitro and in vivo studies suggest that ultrasound (US) is capable of inducing angiogenesis. There is no information, however, on whether ultrasound can induce proliferation of brain endothelial cells. We therefore explored the angiogenic potential of ultrasound on a novel immortalised human brain endothelial cell line (hCMEC/D3) and on mouse brain microvascular endothelial cells (bEND3). Ultrasound failed to enhance cell proliferation in both cell lines at all acoustic pressures studied. Endothelial cell damage occurred at 0.24 MPa with significantly slower proliferation. Cells growing in Opticell{trade mark, serif} dishes did not show damage or reduced proliferation at these pressures.

  1. Human Blood-Brain Barrier Endothelial Cells Derived from Pluripotent Stem Cells

    PubMed Central

    Lippmann, Ethan S.; Azarin, Samira M.; Kay, Jennifer E.; Nessler, Randy A.; Wilson, Hannah K.; Al-Ahmad, Abraham; Palecek, Sean P.; Shusta, Eric V.

    2012-01-01

    The blood-brain barrier (BBB) plays an important role in brain health and is often compromised in disease. Moreover, as a result of its significant barrier properties, this endothelial interface restricts neurotherapeutic uptake. Thus, a renewable source of human BBB endothelium could prove enabling for brain research and pharmaceutical development. Herein, we demonstrate that endothelial cells generated from human pluripotent stem cells (hPSCs) can be specified to possess many BBB attributes, including well-organized tight junctions, expression of nutrient transporters, and polarized efflux transporter activity. Importantly, hPSC-derived BBB endothelial cells respond to astrocytic cues yielding impressive barrier properties as measured by transendothelial electrical resistance (1450±140 Ωxcm2) and molecular permeability that correlates well with in vivo brain uptake. In addition, specification of hPSC-derived BBB endothelial cells occurs in concert with neural cell co-differentiation via Wnt/β-catenin signaling, consistent with previous transgenic studies. This study represents the first example of organ-specific endothelial differentiation from hPSCs. PMID:22729031

  2. Nootropic agents stimulate neurogenesis. Brain Cells, Inc.: WO2007104035.

    PubMed

    Taupin, Philippe

    2009-05-01

    The application is in the field of adult neurogenesis, neural stem cells and cellular therapy. It aims to characterize the activity of nootropic agents on adult neurogenesis in vitro. Nootropic agents are substances improving cognitive and mental abilities. AMPA (alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate) and nootropic agents were assessed for the potential to differentiate human neural progenitor and stem cells into neuronal cells in vitro. They were also tested for their behavioural activity on the novel object recognition task. AMPA, piracetam, FK-960 and SGS-111 induce and stimulate neuronal differentiation of human-derived neural progenitor and stem cells. SGS-111 increases the number of visits to the novel object. The neurogenic activity of piracetam and SGS-111 is mediated through AMPA receptor. The neurogenic activity of SGS-111 may contribute and play a role in its nootropic activity. These results suggest that nootropic agents may elicit some of their effects through their neurogenic activity. The application claims the use of nootropic agents for their neurogenic activity and for the treatment of neurological diseases, disorders and injuries, by stimulating or increasing the generation of neuronal cells in the adult brain.

  3. Cell cavities increase tortuosity in brain extracellular space.

    PubMed

    Tao, A; Tao, L; Nicholson, C

    2005-06-21

    Brain extracellular space (ECS) forms hindered pathways for molecular diffusion in chemical signaling and drug delivery. Hindrance is quantified by the tortuosity lambda; the tortuosity obtained from simulations using uniformly spaced convex cells is significantly lower than that measured experimentally. To attempt to account for the difference in results, this study employed a variety of ECS models based on an array of cubic cells containing open rectangular cavities that provided the ECS with dead-space microdomains. Monte Carlo simulations demonstrated that, in such ECS models, lambda can equal or exceed the typical experimental value of about 1.6. The simulations further revealed that lambda is relatively independent of cavity shape and the number of cavities per cell. It mainly depends on the total ECS volume fraction alpha, the cavity volume fraction alpha(c), and whether the cavity is located at the center of a cell face or formed at the junction of multiple cells. To describe the results from the different ECS models, an expression was obtained that related lambda to alpha, alpha(c), and an empirical exit factor beta that correlated with the ease with which a molecule could leave a cavity and its vicinity.

  4. The Mouse Blood-Brain Barrier Transcriptome: A New Resource for Understanding the Development and Function of Brain Endothelial Cells

    PubMed Central

    Daneman, Richard; Zhou, Lu; Agalliu, Dritan; Cahoy, John D.; Kaushal, Amit; Barres, Ben A.

    2010-01-01

    The blood-brain barrier (BBB) maintains brain homeostasis and limits the entry of toxins and pathogens into the brain. Despite its importance, little is known about the molecular mechanisms regulating the development and function of this crucial barrier. In this study we have developed methods to highly purify and gene profile endothelial cells from different tissues, and by comparing the transcriptional profile of brain endothelial cells with those purified from the liver and lung, we have generated a comprehensive resource of transcripts that are enriched in the BBB forming endothelial cells of the brain. Through this comparison we have identified novel tight junction proteins, transporters, metabolic enzymes, signaling components, and unknown transcripts whose expression is enriched in central nervous system (CNS) endothelial cells. This analysis has identified that RXRalpha signaling cascade is specifically enriched at the BBB, implicating this pathway in regulating this vital barrier. This dataset provides a resource for understanding CNS endothelial cells and their interaction with neural and hematogenous cells. PMID:21060791

  5. CYP24 inhibition preserves 1α,25-dihydroxyvitamin D3 anti-proliferative signaling in lung cancer cells

    PubMed Central

    Zhang, Qiuhong; Kanterewicz, Beatriz; Buch, Shama; Petkovich, Martin; Parise, Robert; Beumer, Jan; Lin, Yan; Diergaarde, Brenda; Hershberger, Pamela A.

    2012-01-01

    Human lung tumors aberrantly express the 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3)-catabolizing enzyme, CYP24. We hypothesized that CYP24 reduces 1,25(OH)2D3-mediated transcription and allows lung cancer cells to escape its growth-inhibitory action. To test this, H292 lung cancer cells and the CYP24-selective inhibitor CTA091 were utilized. In H292 cells, CTA091 reduces 1,25(OH)2D3 catabolism, significantly increases 1,25(OH)2D3-mediated growth inhibition, and increases 1,25(OH)2D3 effects on induced and repressed genesin gene expression profiling studies. Pathway mapping of repressed genes uncovered cell cycle as a predominant 1,25(OH)2D3 target. In H292 cells, 1,25(OH)2D3 significantly decreases cyclin E2 levels and induces G0/G1 arrest. A broader set of cyclins is down-regulated when 1,25(OH)2D3 is combined with CTA091, and cell cycle arrest further increases. Effects of CTA091 on 1,25(OH)2D3 signaling are vitamin D receptor-dependent. These data provide evidence that CYP24 limits 1,25(OH)2D3 anti-proliferative signaling in cancer cells, and suggest that CTA091 may be beneficial in preserving 1,25(OH)2D3 action in lung cancer. PMID:22386975

  6. No attenuation of ischemic and reperfusion injury in Kupffer cell-depleted, cold-preserved rat livers.

    PubMed

    Reinders, M E; van Wagensveld, B A; van Gulik, T M; Corssmit, N P; Frederiks, W M; Chamuleau, R A; van Rooijen, N; Obertop, H

    1997-02-15

    Activated Kupffer cells (KC) have been implicated in the damage sustained by preserved liver grafts during ischemia and reperfusion. The aim of this study was to compare ischemia/reperfusion injury in preserved, KC-depleted rat livers and preserved control livers, with special regard to sinusoidal endothelial cell (SEC) injury. Wistar rats were injected with liposome-encapsulated dichloromethylene diphosphonate, 48 hr before hepatectomy, to eliminate KC, or were withheld this pretreatment (controls). Livers were flushed with cold University of Wisconsin solution and after 0, 8, 16, or 24 hr of storage at 4 degrees C, were reperfused in a recirculation system with 200 ml of oxygenated Krebs-Henseleit solution at 37 degrees C for 90 min. Damage to SEC was measured by the uptake of hyaluronic acid (HA) from the perfusate and release of purine nucleoside phosphorylase (PNP). Perfusate samples were, furthermore, analyzed for aspartate aminotransferase (AST) and tumor necrosis factor-alpha. Carbon particles were infused in the perfusate to determine the phagocytotic capacity of KC. Biopsies were taken for histological examination and sections were stained with ED2 monoclonal antibodies to confirm the absence of KC. After 90 min of reperfusion, immediately after cold flush (t0), the uptake of HA was 72.2+/-2.3% and 69.3+/-1.3% in KC-depleted livers and in control livers, respectively (n.s.). After 8 hr of storage, HA uptake was 21.6+/-4.5% and 34.6+/-8.0%, respectively (n.s.). After 16 and 24 hr of storage and reperfusion, no uptake of HA was found in either KC-depleted or control livers, indicating abolished SEC function. PNP activities in the perfusate were higher in control livers (after 8 and 24 hr of storage), presumably due to release from damaged KC. No difference was found in AST and no tumor necrosis factor-alpha was measured in the perfusates of normal and KC-depleted livers. Electron microscopic studies showed that after 8 and 24 hr of storage and reperfusion

  7. Robust methods for purification of histones from cultured mammalian cells with the preservation of their native modifications.

    PubMed

    Rodriguez-Collazo, Pedro; Leuba, Sanford H; Zlatanova, Jordanka

    2009-06-01

    Post-translational modifications (PTMs) of histones play a role in modifying chromatin structure for DNA-templated processes in the eukaryotic nucleus, such as transcription, replication, recombination and repair; thus, histone PTMs are considered major players in the epigenetic control of these processes. Linking specific histone PTMs to gene expression is an arduous task requiring large amounts of highly purified and natively modified histones to be analyzed by various techniques. We have developed robust and complementary procedures, which use strong protein denaturing conditions and yield highly purified core and linker histones from unsynchronized proliferating, M-phase arrested and butyrate-treated cells, fully preserving their native PTMs without using enzyme inhibitors. Cell hypotonic swelling and lysis, nuclei isolation/washing and chromatin solubilization under mild conditions are bypassed to avoid compromising the integrity of histone native PTMs. As controls for our procedures, we tested the most widely used conventional methodologies and demonstrated that they indeed lead to drastic histone dephosphorylation. Additionally, we have developed methods for preserving acid-labile histone modifications by performing non-acid extractions to obtain highly purified H3 and H4. Importantly, isolation of histones H3, H4 and H2A/H2B is achieved without the use of HPLC. Functional supercoiling assays reveal that both hyper- and hypo-phosphorylated histones can be efficiently assembled into polynucleosomes. Notably, the preservation of fully phosphorylated mitotic histones and their assembly into polynucleosomes should open new avenues to investigate an important but overlooked question: the impact of mitotic phosphorylation in chromatin structure and function.

  8. Cediranib Maleate and Whole Brain Radiation Therapy in Patients With Brain Metastases From Non-Small Cell Lung Cancer

    ClinicalTrials.gov

    2013-03-07

    Male Breast Cancer; Stage IV Breast Cancer; Stage IV Melanoma; Stage IV Non-small Cell Lung Cancer; Stage IV Renal Cell Cancer; Stage IVA Colon Cancer; Stage IVA Rectal Cancer; Stage IVB Colon Cancer; Stage IVB Rectal Cancer; Tumors Metastatic to Brain

  9. Transplantation of multipotent Isl1+ cardiac progenitor cells preserves infarcted heart function in mice

    PubMed Central

    Li, Yunpeng; Tian, Shuo; Lei, Ienglam; Liu, Liu; Ma, Peter; Wang, Zhong

    2017-01-01

    Cell-based cardiac therapy is a promising therapeutic strategy to restore heart function after myocardial infarction (MI). However, the cell type selection and ensuing effects remain controversial. Here, we intramyocardially injected Isl1+ cardiac progenitor cells (CPCs) derived from EGFP/luciferase double-tagged mouse embryonic stem (dt-mES) cells with vehicle (fibrin gel) or phosphate-buffered saline (PBS) into the infarcted area in nude mice to assess the contribution of CPCs to the recovery of cardiac function post-MI. Our results showed that Isl1+ CPCs differentiated normally into three cardiac lineages (cardiomyocytes (CMs), endothelial cells and smooth muscle cells) both on cell culture plates and in fibrin gel. Cell retention was significantly increased when the transplanted cells were injected with vehicle. Importantly, 28 days after injection, CPCs were observed to differentiate into CMs within the infarcted area. Moreover, numerous CD31+ endothelial cells derived from endogenous revascularization and differentiation of the injected CPCs were detected. SMMHC-, Ki67- and CX-43-positive cells were identified in the injected CPC population, further demonstrating the proliferation, differentiation and integration of the transplanted CPCs in host cells. Furthermore, animal hearts injected with CPCs showed increased angiogenesis, decreased infarct size, and improved heart function. In conclusion, our studies showed that Isl1+ CPCs, when combined with a suitable vehicle, can produce notable therapeutic effects in the infarcted heart, suggesting that CPCs might be an ideal cell source for cardiac therapy. PMID:28386378

  10. Transplantation of multipotent Isl1+ cardiac progenitor cells preserves infarcted heart function in mice.

    PubMed

    Li, Yunpeng; Tian, Shuo; Lei, Ienglam; Liu, Liu; Ma, Peter; Wang, Zhong

    2017-01-01

    Cell-based cardiac therapy is a promising therapeutic strategy to restore heart function after myocardial infarction (MI). However, the cell type selection and ensuing effects remain controversial. Here, we intramyocardially injected Isl1+ cardiac progenitor cells (CPCs) derived from EGFP/luciferase double-tagged mouse embryonic stem (dt-mES) cells with vehicle (fibrin gel) or phosphate-buffered saline (PBS) into the infarcted area in nude mice to assess the contribution of CPCs to the recovery of cardiac function post-MI. Our results showed that Isl1+ CPCs differentiated normally into three cardiac lineages (cardiomyocytes (CMs), endothelial cells and smooth muscle cells) both on cell culture plates and in fibrin gel. Cell retention was significantly increased when the transplanted cells were injected with vehicle. Importantly, 28 days after injection, CPCs were observed to differentiate into CMs within the infarcted area. Moreover, numerous CD31+ endothelial cells derived from endogenous revascularization and differentiation of the injected CPCs were detected. SMMHC-, Ki67- and CX-43-positive cells were identified in the injected CPC population, further demonstrating the proliferation, differentiation and integration of the transplanted CPCs in host cells. Furthermore, animal hearts injected with CPCs showed increased angiogenesis, decreased infarct size, and improved heart function. In conclusion, our studies showed that Isl1+ CPCs, when combined with a suitable vehicle, can produce notable therapeutic effects in the infarcted heart, suggesting that CPCs might be an ideal cell source for cardiac therapy.

  11. Therapeutic Approaches for Preserving or Restoring Pancreatic β-Cell Function and Mass

    PubMed Central

    Jung, Kyong Yeun; Kim, Kyoung Min

    2014-01-01

    The goal for the treatment of patients with diabetes has today shifted from merely reducing glucose concentrations to preventing the natural decline in β-cell function and delay the progression of disease. Pancreatic β-cell dysfunction and decreased β-cell mass are crucial in the development of diabetes. The β-cell defects are the main pathogenesis in patients with type 1 diabetes and are associated with type 2 diabetes as the disease progresses. Recent studies suggest that human pancreatic β-cells have a capacity for increased proliferation according to increased demands for insulin. In humans, β-cell mass has been shown to increase in patients showing insulin-resistance states such as obesity or in pregnancy. This capacity might be useful for identifying new therapeutic strategies to reestablish a functional β-cell mass. In this context, therapeutic approaches designed to increase β-cell mass might prove a significant way to manage diabetes and prevent its progression. This review describes the various β-cell defects that appear in patients with diabetes and outline the mechanisms of β-cell failure. We also review common methods for assessing β-cell function and mass and methodological limitations in vivo. Finally, we discuss the current therapeutic approaches to improve β-cell function and increase β-cell mass. PMID:25541605

  12. Brain and muscle Arnt-like 1 promotes skeletal muscle regeneration through satellite cell expansion

    SciTech Connect

    Chatterjee, Somik; Yin, Hongshan; Nam, Deokhwa; Li, Yong; Ma, Ke

    2015-02-01

    Circadian clock is an evolutionarily conserved timing mechanism governing diverse biological processes and the skeletal muscle possesses intrinsic functional clocks. Interestingly, although the essential clock transcription activator, Brain and muscle Arnt-like 1 (Bmal1), participates in maintenance of muscle mass, little is known regarding its role in muscle growth and repair. In this report, we investigate the in vivo function of Bmal1 in skeletal muscle regeneration using two muscle injury models. Bmal1 is highly up-regulated by cardiotoxin injury, and its genetic ablation significantly impairs regeneration with markedly suppressed new myofiber formation and attenuated myogenic induction. A similarly defective regenerative response is observed in Bmal1-null mice as compared to wild-type controls upon freeze injury. Lack of satellite cell expansion accounts for the regeneration defect, as Bmal1{sup −/−} mice display significantly lower satellite cell number with nearly abolished induction of the satellite cell marker, Pax7. Furthermore, satellite cell-derived primary myoblasts devoid of Bmal1 display reduced growth and proliferation ex vivo. Collectively, our results demonstrate, for the first time, that Bmal1 is an integral component of the pro-myogenic response that is required for muscle repair. This mechanism may underlie its role in preserving adult muscle mass and could be targeted therapeutically to prevent muscle-wasting diseases. - Highlights: • Bmal1 is highly inducible by muscle injury and myogenic stimuli. • Genetic ablation of Bmal1 significantly impairs muscle regeneration. • Bmal1 promotes satellite cell expansion during muscle regeneration. • Bmal1-deficient primary myoblasts display attenuated growth and proliferation.

  13. Preservation Microfilming

    ERIC Educational Resources Information Center

    Sajor, Ladd Z.

    1972-01-01

    Microfilming preserves the library's holdings while creating space for new acquisitions without the need for new library construction and physical expansion. In addition, microfilming protects rare originals from excessive handling, preserves material with permanent research value and makes possible economic demand" reprinting via positive…

  14. Evidence for Inflammatory Cell Involvement in Brain Arteriovenous Malformations

    PubMed Central

    Chen, Yongmei; Zhu, Wei; Bollen, Andrew W.; Lawton, Michael T.; Barbaro, Nicholas M.; Dowd, Christopher F.; Hashimoto, Tomoki; Yang, Guo-Yuan; Young, William L.

    2008-01-01

    Objective Brain arteriovenous malformations (AVM) have high MMP-9, IL-6 and MPO expression, and polymorphic variations in inflammatory genes are associated with increased risk of hemorrhage. In this study, we characterized the presence of inflammatory cells in AVM lesional tissues. Methods Immunohistochemistry was used to identify and localize neutrophils (MPO as marker), macrophages/microglia (CD68 as marker), T lymphocytes (CD3 as marker), and B lymphocytes (CD20 as marker). Endothelial cell (EC) marker CD31 was used as an index to assess vascular mass (EC mass). Surgical specimens from 20 unruptured, non-embolized AVMs were examined; seven cortical samples from temporal lobectomy were used as controls. Positive signals for inflammatory cell markers were counted and analyzed by normalizing to the area of the tissue section and the amount of endothelial cells (cells/mm2/EC mass pixels). Levels of MPO and MMP-9 were determined by ELISA. Results Neutrophils and macrophages are all frequently identified in the vascular wall of AVM tissues. In contrast, T and B lymphocytes are rarely observed in AVM tissues. AVM tissues displayed more neutrophil and macrophage/microglia markers than epilepsy control tissues (MPO: 434 ± 333 vs 5 ± 4, P=0.0001; CD68: 454 ± 404 vs 4 ± 2, P=0.0001; cells/mm2/EC mass pixels). In ex vivo studies, neutrophil quantity, MPO, and MMP-9 levels were all co-linear(R2=0.98–0.99). Conclusion Our study demonstrates that inflammatory cells are present in AVM tissues. Taken together with prior genetic and cytokine studies, these data are consistent with a novel view that inflammation is associated with AVM disease progression and rupture. PMID:18825001

  15. Cranial grafting of stem cell-derived microvesicles improves cognition and reduces neuropathology in the irradiated brain

    PubMed Central

    Baulch, Janet E.; Acharya, Munjal M.; Allen, Barrett D.; Ru, Ning; Chmielewski, Nicole N.; Martirosian, Vahan; Giedzinski, Erich; Syage, Amber; Park, Audrey L.; Benke, Sarah N.; Parihar, Vipan K.; Limoli, Charles L.

    2016-01-01

    Cancer survivors face a variety of challenges as they cope with disease recurrence and a myriad of normal tissue complications brought on by radio- and chemotherapeutic treatment regimens. For patients subjected to cranial irradiation for the control of CNS malignancy, progressive and debilitating cognitive dysfunction remains a pressing unmet medical need. Although this problem has been recognized for decades, few if any satisfactory long-term solutions exist to resolve this serious unintended side effect of radiotherapy. Past work from our laboratory has demonstrated the neurocognitive benefits of human neural stem cell (hNSC) grafting in the irradiated brain, where intrahippocampal transplantation of hNSC ameliorated radiation-induced cognitive deficits. Using a similar strategy, we now provide, to our knowledge, the first evidence that cranial grafting of microvesicles secreted from hNSC affords similar neuroprotective phenotypes after head-only irradiation. Cortical- and hippocampal-based deficits found 1 mo after irradiation were completely resolved in animals cranially grafted with microvesicles. Microvesicle treatment was found to attenuate neuroinflammation and preserve host neuronal morphology in distinct regions of the brain. These data suggest that the neuroprotective properties of microvesicles act through a trophic support mechanism that reduces inflammation and preserves the structural integrity of the irradiated microenvironment. PMID:27044087

  16. Cranial grafting of stem cell-derived microvesicles improves cognition and reduces neuropathology in the irradiated brain.

    PubMed

    Baulch, Janet E; Acharya, Munjal M; Allen, Barrett D; Ru, Ning; Chmielewski, Nicole N; Martirosian, Vahan; Giedzinski, Erich; Syage, Amber; Park, Audrey L; Benke, Sarah N; Parihar, Vipan K; Limoli, Charles L

    2016-04-26

    Cancer survivors face a variety of challenges as they cope with disease recurrence and a myriad of normal tissue complications brought on by radio- and chemotherapeutic treatment regimens. For patients subjected to cranial irradiation for the control of CNS malignancy, progressive and debilitating cognitive dysfunction remains a pressing unmet medical need. Although this problem has been recognized for decades, few if any satisfactory long-term solutions exist to resolve this serious unintended side effect of radiotherapy. Past work from our laboratory has demonstrated the neurocognitive benefits of human neural stem cell (hNSC) grafting in the irradiated brain, where intrahippocampal transplantation of hNSC ameliorated radiation-induced cognitive deficits. Using a similar strategy, we now provide, to our knowledge, the first evidence that cranial grafting of microvesicles secreted from hNSC affords similar neuroprotective phenotypes after head-only irradiation. Cortical- and hippocampal-based deficits found 1 mo after irradiation were completely resolved in animals cranially grafted with microvesicles. Microvesicle treatment was found to attenuate neuroinflammation and preserve host neuronal morphology in distinct regions of the brain. These data suggest that the neuroprotective properties of microvesicles act through a trophic support mechanism that reduces inflammation and preserves the structural integrity of the irradiated microenvironment.

  17. Tissue Engineered, Hydrogel-Based Endothelial Progenitor Cell Therapy Robustly Revascularizes Ischemic Myocardium and Preserves Ventricular Function

    PubMed Central

    Atluri, Pavan; Miller, Jordan S; Emery, Robert J; Hung, George; Trubelja, Alen; Cohen, Jeffrey E; Lloyd, Kelsey; Han, Jason; Gaffey, Ann C; MacArthur, John W; Chen, Christopher S; Woo, Y Joseph

    2014-01-01

    Objective Cell based angiogenic therapy for ischemic heart failure has had limited clinical impact, likely related to very low cell retention (<1%) and dispersion. We developed a novel, tissue engineered, hydrogel based cell delivery strategy to overcome these limitations and provide prolonged regional retention of myocardial endothelial progenitor cells (EPC) at high cell dosage. Methods EPCs were isolated from Wistar Rats and encapsulated in fibrin gels. In vitro viability was quantified using a fluorescent live-dead stain of transgenic eGFP+ EPCs. EPC-laden constructs were implanted onto ischemic rat myocardium in a model of acute myocardial infarction (LAD ligation) for 4 weeks. Intramyocardial cell injection (IC, 2×106 EPCs), empty fibrin, and isolated LAD ligation groups served as controls. Hemodynamics were quantified using echocardiography, Doppler flow analysis, and intraventricular pressure-volume analysis. Vasculogenesis and ventricular geometry were quantified. EPC migration was analyzed by utilizing EPCs from transgenic eGFP+ rodents. Results EPCs demonstrated an overall 88.7% viability for all matrix and cell conditions investigated after 48 hours. Histologic assessment of 1-wk implants demonstrated significant migration of transgenic eGFP+ EPCs from the fibrin matrix to the infarcted myocardium as compared to IC (28±12.3 vs. 2.4±2.1cells/hpf, p=0.0001). We also observed a marked increase in vasculogenesis at the implant site. Significant improvements in ventricular hemodynamics and geometry were present following EPC-hydrogel therapy as compared to control. Conclusion We present a tissue engineered hydrogel-based EPC mediated therapy to enhance cell delivery, cell retention, vasculogenesis, and preservation of myocardial structure and function. PMID:25129603

  18. Palmitoylethanolamide relieves pain and preserves pancreatic islet cells in a murine model of diabetes.

    PubMed

    Donvito, Giulia; Bettoni, Isabella; Comelli, Francesca; Colombo, Anita; Costa, Barbara

    2015-01-01

    We previously demonstrated that the intraperitoneal administration of palmitoylethanolamide (PEA) in mice with chronic constriction injury of the sciatic nerve evoked a relief of both thermal hyperalgesia and mechanical allodynia in neuropathic mice. Since diabetic neuropathy is one of the most common long-term complications of diabetes, we explored the ability of PEA to also relief this kind of chronic pain, employing the well established streptozotocin-induced animal model of type 1 diabetes. Our findings demonstrated that PEA relieves mechanical allodynia, counteracts nerve growth factor deficit, improves insulin level, preserves Langerhans islet morphology reducing the development of insulitis in diabetic mice. These results suggest that PEA could be effective in type 1-diabetic patients not only as pain reliever but also in controlling the development of pathology.

  19. Preserved Ex Vivo Inflammatory Status in Decidual Cells from Women with Preterm Labor and Subclinical Intrauterine Infection

    PubMed Central

    Castro-Leyva, Violeta; Espejel-Nuñez, Aurora; Barroso, Gerardo; Zaga-Clavellina, Veronica; Flores-Pliego, Arturo; Morales-Mendez, Iyari; Giono-Cerezo, Silvia; Walsh, Scott W.; Estrada-Gutierrez, Guadalupe

    2012-01-01

    Objective To compare the inflammatory response preserved ex vivo by decidual cells isolated from women who experienced preterm labor with and without subclinical intrauterine infection. Methods Fetal membranes were obtained after cesarean section from 35 women who delivered before 37 weeks of gestation following spontaneous preterm labor, with no clinical evidence of intrauterine infection. Decidua was microbiologically tested and cultured. Concentrations of anti-inflammatory cytokines (IL-2, IL-4, IL-10), pro-inflammatory cytokines (IL-6, IL-8, IL-1β and TNF-α), and matrix metalloproteinases (MMP-1, MMP-2, MMP-3, MMP-7, MMP-8, MMP-9) were measured in the supernatants using Bio-Plex, and prostaglandin E2 (PGE2) was measured by enzyme immunoassay. Results Subclinical infection was confirmed in 10 women (28.5%). Microorganisms isolated were Ureaplasma urealyticum (4), group B streptococci (3), Gardnerella vaginalis (1), and Escherichia coli (2). We found a significant increase of pro-inflammatory cytokines and a significant decrease of anti-inflammatory cytokines in supernatants from decidual cells obtained from women with preterm labor and subclinical intrauterine infection compared to women without infection. Secretion of MMP-1, MMP-8, MMP-9 and PGE2 was significantly higher in infected women. Secretion of IL-8 by decidual cells from infected women persisted upon repeated in vitro culture passages. Conclusions Almost 30% of idiopathic preterm labor cases were associated with subclinical intrauterine infection, and decidual cells isolated from these cases preserved an ex vivo inflammatory status after in vivo bacterial exposure. PMID:22928002

  20. Brain Tumors

    MedlinePlus

    A brain tumor is a growth of abnormal cells in the tissues of the brain. Brain tumors can be benign, with no cancer cells, ... cancer cells that grow quickly. Some are primary brain tumors, which start in the brain. Others are ...

  1. Identification of Vulnerable Cell Types in Major Brain Disorders Using Single Cell Transcriptomes and Expression Weighted Cell Type Enrichment.

    PubMed

    Skene, Nathan G; Grant, Seth G N

    2016-01-01

    The cell types that trigger the primary pathology in many brain diseases remain largely unknown. One route to understanding the primary pathological cell type for a particular disease is to identify the cells expressing susceptibility genes. Although this is straightforward for monogenic conditions where the causative mutation may alter expression of a cell type specific marker, methods are required for the common polygenic disorders. We developed the Expression Weighted Cell Type Enrichment (EWCE) method that uses single cell transcriptomes to generate the probability distribution associated with a gene list having an average level of expression within a cell type. Following validation, we applied EWCE to human genetic data from cases of epilepsy, Schizophrenia, Autism, Intellectual Disability, Alzheimer's disease, Multiple Sclerosis and anxiety disorders. Genetic susceptibility primarily affected microglia in Alzheimer's and Multiple Sclerosis; was shared between interneurons and pyramidal neurons in Autism and Schizophrenia; while intellectual disabilities and epilepsy were attributable to a range of cell-types, with the strongest enrichment in interneurons. We hypothesized that the primary cell type pathology could trigger secondary changes in other cell types and these could be detected by applying EWCE to transcriptome data from diseased tissue. In Autism, Schizophrenia and Alzheimer's disease we find evidence of pathological changes in all of the major brain cell types. These findings give novel insight into the cellular origins and progression in common brain disorders. The methods can be applied to any tissue and disorder and have applications in validating mouse models.

  2. Liraglutide Enhances the Efficacy of Human Mesenchymal Stem Cells in Preserving Islet β-cell Function in Severe Non-obese Diabetic Mice

    PubMed Central

    Li, Li-rong; Lu, Jing; Jia, Xiao-lei; Hui, Hui; Zhang, Jie; Liu, Ying; Cui, Wei-juan; Xu, Qian-yue; Zhu, Da-long

    2016-01-01

    Glucagon-like peptide 1 (GLP-1) can promote islet β cell replication and function, and mesenchymal stem cells (MSCs) can inhibit T cell autoimmunity. The aim of this study was to test the dynamic distribution of infused human MSCs and the therapeutic effect of combined MSCs and liraglutide, a long-acting GLP-1 analog, on preserving β cell function in severe nonobese diabetic (NOD) mice. We found that infused MSCs accumulated in the pancreas at 4 wks post infusion, which was not affected by liraglutide treatment. Liraglutide significantly enhanced the function of MSCs to preserve islet β cells by reducing glucose levels 30 min post glucose challenge and increasing the content and secretion of insulin by islet β cells in severely diabetic mice. Infusion with MSCs significantly reduced insulitis scores but increased the frequency of splenic Tregs, accompanied by a reduction in plasma IFN-γ and TNF-α levels and an elevation of plasma IL-10 and transforming growth factor-β1 (TGF-β1) levels in NOD mice. Although liraglutide mitigated MSC-mediated changes in the frequency of Tregs and the level of plasma IL-10, it significantly increased the plasma TGF-β1 levels in severely diabetic mice. Therefore, our findings suggest that liraglutide could enhance the therapeutic efficacy of MSCs in the treatment of severe type 1 diabetes. PMID:27878211

  3. Cytotoxicity of prostaglandin analog eye drops preserved with benzalkonium chloride in multiple corneoconjunctival cell lines

    PubMed Central

    Ayaki, Masahiko; Iwasawa, Atsuo

    2010-01-01

    Purpose: This study evaluated the cytotoxicity of five prostaglandin analog ophthalmic solutions on four ocular surface cell lines, ie, Chang (human conjunctiva), SIRC (rabbit cornea), RC-1 (rabbit cornea), and BCE C/D-1b (bovine cornea). Methods: Cell viability was measured by neutral red and MTT assays in cells treated for 10, 30, or 60 minutes with various doses of prostaglandins (undiluted, and 2- and 10-fold dilutions). The number of cell lines with viability ≥50% in the presence of selected dilution of the drug (CVS50) was used for comparison. In addition, 24 cell viability comparisons (four cell lines, two assays, and three exposure times) were made between latanoprost (Xalatan®) and each other solution at each dose. A comparison between the newly introduced tafluprost (Tapros®) with 0.01% benzalkonium chloride was also made. Results: The order of cell viability determined by CVS50 was Travatan Z® (travoprost with the SofZia system) > Tapros ≥ Travatan® (travoprost) = Xalatan > Rescula® (unoproston). This was consistent with the results of direct comparisons between Xalatan and the other drugs. There was no clear difference in cell viability between Tapros and benzalkonium chloride. Conclusions: Use of two assays, multiple cell lines, and various dilutions and exposure times provided a unique evaluation of cytotoxicity among ophthalmic solutions. CVS50 was useful for comparison of the cell viability of the solutions. PMID:20823934

  4. Embryo production and possible species preservation by nuclear transfer of somatic cells isolated from bovine semen.

    PubMed

    Liu, Jie; Westhusin, Mark; Long, Charles; Johnson, Gregory; Burghardt, Robert; Kraemer, Duane

    2010-12-01

    Somatic cells in semen are a potential source of nuclei for nuclear transfer to produce genetically identical animals; this is especially important when an animal has died and the only viable genetic material available is frozen semen. Usefulness of somatic cells obtained from fresh (cultured) and frozen (isolated, not cultured) bovine semen for nuclear transfer was evaluated. Twelve ejaculates were collected from nine bulls representing three breeds: Charolais, Brahman, and crossbred Rodeo bull. All samples were processed immediately and cell growth was obtained from seven of the twelve ejaculates (58.3%). Cells from three bulls (with the best growth rates) were evaluated by optical microscopy and used in cloning experiments. In culture, these cells exhibited classic epithelial morphology and expressed cytokeratin and vimentin, indicating they were of epithelial origin. When cells from the three bulls were used as donor cells, 15.9% (18/113), 34.5% (29/84), and 14.4% (13/90) of the fused embryos developed into blastocysts, respectively. Of the blastocyst stage embryos, 38.9% (7/18), 72.4% (21/29), and 61.5% (8/13) hatched, respectively. Somatic cells isolated (not cultured) from frozen bovine semen were also used in the cloning experiments. Although cleavage occurred, no compact morulae or blastocysts were obtained. In conclusion, epithelial cell growth was obtained from fresh bovine ejaculates with relatively high efficiency. Somatic cells from semen can be used as nucleus donors to produce cloned blastocyst-stage embryos.

  5. Preservation of the 3D Phenotype Upon Dispersal of Cultured Cell Spheroids Into Monolayer Cultures.

    PubMed

    Koshkin, Vasilij; Ailles, Laurie E; Liu, Geoffrey; Krylov, Sergey N

    2017-01-01

    In functional cytometric studies, cultured cells are exposed to effectors (e.g., drugs), and the heterogeneity of cell responses are studied using cytometry techniques (e.g., image cytometry). Such studies are difficult to perform on 3D cell cultures. A solution is to disperse 3D clusters and transfer the cells to the 2D state before applying effectors and using cytometry. This approach requires that the lifetime of the 3D phenotype be longer than the duration of the experiment. Here we studied the dynamics of phenotype transformation from 3D to 2D and searched for means of slowing this transformation down in dispersed spheroids of MCF7 cells. We found three functional biomarkers of the 3D phenotype in MCF7 cell spheroids that are absent in the 2D cell culture: (i) the presence of a subpopulation with an elevated drug-expelling capacity; (ii) the presence of a subpopulation with an elevated cytoprotective capacity; and (iii) the accumulation of cells in the G1 phase of the cell cycle. Monitoring these biomarkers in cells transferred from the 3D state to the 2D state revealed their gradual extinction. We found that the combined application of an elevated cell density and thiol-containing medium supplements increased the lifetime of the 3D phenotype by several fold to as long as 96 h. Our results suggest that extending the lifetime of the 3D phenotype in the cells transferred from the 3D state to the 2D state can facilitate detailed functional cytometric studies, such as measurements of population heterogeneity of cytotoxicity, chemosensitivity, and radiosensitivity. J. Cell. Biochem. 118: 154-162, 2017. © 2016 Wiley Periodicals, Inc.

  6. Internalization of targeted quantum dots by brain capillary endothelial cells in vivo.

    PubMed

    Paris-Robidas, Sarah; Brouard, Danny; Emond, Vincent; Parent, Martin; Calon, Frédéric

    2016-04-01

    Receptors located on brain capillary endothelial cells forming the blood-brain barrier are the target of most brain drug delivery approaches. Yet, direct subcellular evidence of vectorized transport of nanoformulations into the brain is lacking. To resolve this question, quantum dots were conjugated to monoclonal antibodies (Ri7) targeting the murine transferrin receptor. Specific transferrin receptor-mediated endocytosis of Ri7-quantum dots was first confirmed in N2A and bEnd5 cells. After intravenous injection in mice, Ri7-quantum dots exhibited a fourfold higher volume of distribution in brain tissues, compared to controls. Immunofluorescence analysis showed that Ri7-quantum dots were sequestered throughout the cerebral vasculature 30 min, 1 h, and 4 h post injection, with a decline of signal intensity after 24 h. Transmission electron microscopic studies confirmed that Ri7-quantum dots were massively internalized by brain capillary endothelial cells, averaging 37 ± 4 Ri7-quantum dots/cell 1 h after injection. Most quantum dots within brain capillary endothelial cells were observed in small vesicles (58%), with a smaller proportion detected in tubular structures or in multivesicular bodies. Parenchymal penetration of Ri7-quantum dots was extremely low and comparable to control IgG. Our results show that systemically administered Ri7-quantum dots complexes undergo extensive endocytosis by brain capillary endothelial cells and open the door for novel therapeutic approaches based on brain endothelial cell drug delivery.

  7. Generation recruitment and death of brain cells throughout the life cycle of Sorex shrews (Lipotyphla).

    PubMed

    Bartkowska, Katarzyna; Djavadian, Rouzanna L; Taylor, Jan R E; Turlejski, Kris

    2008-04-01

    Young shrews of the genus Sorex that are born in early summer reduce their body size before wintering, including a reduction of brain weight of 10-30%. In the spring they mature sexually, double their body weight and regain about half of the loss in brain weight. To investigate the mechanisms of brain weight oscillations we studied the rate of cell death and generation in the brain during the whole life cycle of the common shrew (Sorex araneus) and pygmy shrew (S. minutus). After weaning, shrews generate new brain cells in only two mammalian neurogenic zones and approximately 80% of these develop into neurones. The increase of the shrew brain weight in the spring did not depend on recruitment of new cells. Moreover, adult Sorex shrews did not generate new cells in the dentate gyri. Injections of 5-HT1A receptor agonists in the adult shrews induced neurogenesis in their dentate gyri, showing the presence of dormant progenitor cells. Generation of new neurones in the subventricular zone of the lateral ventricles and their recruitment to olfactory bulbs continued throughout life. TUNEL labelling showed that the rate of cell death in all brain structures, including the proliferation zones and olfactory bulb, was very low throughout life. We conclude that neither cell death nor recruitment significantly contributes to seasonal oscillations and the net loss of brain weight in the Sorex shrews. With the exception of dentate gyrus and olfactory bulb, cellular populations of brain structures are stable throughout the life cycle of these shrews.

  8. Targeted delivery of neural stem cells to the brain using MRI-guided focused ultrasound to disrupt the blood-brain barrier.

    PubMed

    Burgess, Alison; Ayala-Grosso, Carlos A; Ganguly, Milan; Jordão, Jessica F; Aubert, Isabelle; Hynynen, Kullervo

    2011-01-01

    Stem cell therapy is a promising strategy to treat neurodegenerative diseases, traumatic brain injury, and stroke. For stem cells to progress towards clinical use, the risks associated with invasive intracranial surgery used to deliver the cells to the brain, needs to be reduced. Here, we show that MRI-guided focused ultrasound (MRIgFUS) is a novel method for non-invasive delivery of stem cells from the blood to the brain by opening the blood brain barrier (BBB) in specific brain regions. We used MRI guidance to target the ultrasound beam thereby delivering the iron-labeled, green fluorescent protein (GFP)-expressing neural stem cells specifically to the striatum and the hippocampus of the rat brain. Detection of cellular iron using MRI established that the cells crossed the BBB to enter the brain. After sacrifice, 24 hours later, immunohistochemical analysis confirmed the presence of GFP-positive cells in the targeted brain regions. We determined that the neural stem cells expressed common stem cell markers (nestin and polysialic acid) suggesting they survived after transplantation with MRIgFUS. Furthermore, delivered stem cells expressed doublecortin in vivo indicating the stem cells were capable of differentiating into neurons. Together, we demonstrate that transient opening of the BBB with MRIgFUS is sufficient for transplantation of stem cells from the blood to targeted brain structures. These results suggest that MRIgFUS may be an effective alternative to invasive intracranial surgery for stem cell transplantation.

  9. Aldehyde dehydrogenase 1 positive glioblastoma cells show brain tumor stem cell capacity.

    PubMed

    Rasper, Michael; Schäfer, Andrea; Piontek, Guido; Teufel, Julian; Brockhoff, Gero; Ringel, Florian; Heindl, Stefan; Zimmer, Claus; Schlegel, Jürgen

    2010-10-01

    Glioblastoma (GBM) is the most aggressive primary brain tumor and is resistant to all therapeutic regimens. Relapse occurs regularly and might be caused by a poorly characterized tumor stem cell (TSC) subpopulation escaping therapy. We suggest aldehyde dehydrogenase 1 (ALDH1) as a novel stem cell marker in human GBM. Using the neurosphere formation assay as a functional method to identify brain TSCs, we show that high protein levels of ALDH1 facilitate neurosphere formation in established GBM cell lines. Even single ALDH1 positive cells give rise to colonies and neurospheres. Consequently, the inhibition of ALDH1 in vitro decreases both the number of neurospheres and their size. Cell lines without expression of ALDH1 do not form tumor spheroids under the same culturing conditions. High levels of ALDH1 seem to keep tumor cells in an undifferentiated, stem cell-like state indicated by the low expression of beta-III-tubulin. In contrast, ALDH1 inhibition induces premature cellular differentiation and reduces clonogenic capacity. Primary cell cultures obtained from fresh tumor samples approve the established GBM cell line results.

  10. Preserving Mafa expression in diabetic islet β-cells improves glycemic control in vivo.

    PubMed

    Matsuoka, Taka-aki; Kaneto, Hideaki; Kawashima, Satoshi; Miyatsuka, Takeshi; Tochino, Yoshihiro; Yoshikawa, Atsushi; Imagawa, Akihisa; Miyazaki, Jun-ichi; Gannon, Maureen; Stein, Roland; Shimomura, Iichiro

    2015-03-20

    The murine Mafa transcription factor is a key regulator of postnatal islet β-cell activity, affecting insulin transcription, insulin secretion, and β-cell mass. Human MAFA expression is also markedly decreased in islet β-cells of type 2 diabetes mellitus (T2DM) patients. Moreover, levels are profoundly reduced in db/db islet β-cells, a mouse model of T2DM. To examine the significance of this key islet β-cell-enriched protein to glycemic control under diabetic conditions, we generated transgenic mice that conditionally and specifically produced Mafa in db/db islet β-cells. Sustained expression of Mafa resulted in significantly lower plasma glucose levels, higher plasma insulin, and augmented islet β-cell mass. In addition, there was increased expression of insulin, Slc2a2, and newly identified Mafa-regulated genes involved in reducing β-cell stress, like Gsta1 and Gckr. Importantly, the levels of human GSTA1 were also compromised in T2DM islets. Collectively, these results illustrate how consequential the reduction in Mafa activity is to islet β-cell function under pathophysiological conditions.

  11. Hydrogen gas treatment prolongs replicative lifespan of bone marrow multipotential stromal cells in vitro while preserving differentiation and paracrine potentials.

    PubMed

    Kawasaki, Haruhisa; Guan, Jianjun; Tamama, Kenichi

    2010-07-02

    Cell therapy with bone marrow multipotential stromal cells/mesenchymal stem cells (MSCs) represents a promising approach in the field of regenerative medicine. Low frequency of MSCs in adult bone marrow necessitates ex vivo expansion of MSCs after harvest; however, such a manipulation causes cellular senescence with loss of differentiation, proliferative, and therapeutic potentials of MSCs. Hydrogen molecules have been shown to exert organ protective effects through selective reduction of hydroxyl radicals. As oxidative stress is one of the key insults promoting cell senescence in vivo as well as in vitro, we hypothesized that hydrogen molecules prevent senescent process during MSC expansion. Addition of 3% hydrogen gas enhanced preservation of colony forming early progenitor cells within MSC preparation and prolonged the in vitro replicative lifespan of MSCs without losing differentiation potentials and paracrine capabilities. Interestingly, 3% hydrogen gas treatment did not decrease hydroxyl radical, protein carbonyl, and 8-hydroxydeoxyguanosine, suggesting that scavenging hydroxyl radical might not be responsible for these effects of hydrogen gas in this study.

  12. Transient inhibition of ROR-γt therapeutically limits intestinal inflammation by reducing TH17 cells and preserving ILC3

    PubMed Central

    Withers, David R.; Hepworth, Matthew R.; Wang, Xinxin; Mackley, Emma C.; Halford, Emily E.; Dutton, Emma E.; Marriott, Clare L.; Brucklacher-Waldert, Verena; Veldhoen, Marc; Kelsen, Judith; Baldassano, Robert N.; Sonnenberg, Gregory F.

    2016-01-01

    RAR-related orphan receptor γt (ROR-γt) directs differentiation of pro-inflammatory T helper 17 (TH17) cells and is a potential therapeutic target in chronic autoimmune and inflammatory diseases1–3. However, ROR-γt-dependent group 3 innate lymphoid cells (ILC3s) provide essential immunity and tissue protection in the intestine4–11, suggesting that targeting ROR-γt could also result in impaired host defense to infection or enhanced tissue damage. Here, we demonstrate that transient chemical inhibition of ROR-γt in mice selectively reduces cytokine production from TH17 cells but not ILC3s in the context of intestinal infection with Citrobacter rodentium, resulting in preserved innate immunity. Transient genetic deletion of ROR-γt in mature ILC3s also did not impair cytokine responses in the steady state or during infection. Finally, pharmacologic inhibition of ROR-γt provided therapeutic benefit in mouse models of intestinal inflammation, and reduced the frequencies of TH17 cells but not ILC3s isolated from primary intestinal samples of individuals with inflammatory bowel disease (IBD). Collectively, these results reveal differential requirements for ROR-γt in the maintenance of TH17 cell versus ILC3 responses, and suggest that transient inhibition of ROR-γt is a safe and effective therapeutic approach during intestinal inflammation. PMID:26878233

  13. Hydrogen gas treatment prolongs replicative lifespan of bone marrow multipotential stromal cells in vitro while preserving differentiation and paracrine potentials

    SciTech Connect

    Kawasaki, Haruhisa; Guan, Jianjun; Tamama, Kenichi

    2010-07-02

    Cell therapy with bone marrow multipotential stromal cells/mesenchymal stem cells (MSCs) represents a promising approach in the field of regenerative medicine. Low frequency of MSCs in adult bone marrow necessitates ex vivo expansion of MSCs after harvest; however, such a manipulation causes cellular senescence with loss of differentiation, proliferative, and therapeutic potentials of MSCs. Hydrogen molecules have been shown to exert organ protective effects through selective reduction of hydroxyl radicals. As oxidative stress is one of the key insults promoting cell senescence in vivo as well as in vitro, we hypothesized that hydrogen molecules prevent senescent process during MSC expansion. Addition of 3% hydrogen gas enhanced preservation of colony forming early progenitor cells within MSC preparation and prolonged the in vitro replicative lifespan of MSCs without losing differentiation potentials and paracrine capabilities. Interestingly, 3% hydrogen gas treatment did not decrease hydroxyl radical, protein carbonyl, and 8-hydroxydeoxyguanosine, suggesting that scavenging hydroxyl radical might not be responsible for these effects of hydrogen gas in this study.

  14. Preservation Environments

    NASA Technical Reports Server (NTRS)

    Moore, Reagan W.

    2004-01-01

    The long-term preservation of digital entities requires mechanisms to manage the authenticity of massive data collections that are written to archival storage systems. Preservation environments impose authenticity constraints and manage the evolution of the storage system technology by building infrastructure independent solutions. This seeming paradox, the need for large archives, while avoiding dependence upon vendor specific solutions, is resolved through use of data grid technology. Data grids provide the storage repository abstractions that make it possible to migrate collections between vendor specific products, while ensuring the authenticity of the archived data. Data grids provide the software infrastructure that interfaces vendor-specific storage archives to preservation environments.

  15. Behavior of neural stem cells in the Alzheimer brain.

    PubMed

    Waldau, B; Shetty, A K

    2008-08-01

    Alzheimer's disease (AD) is characterized by the deposition of beta-amyloid peptides (Abeta) and a progressive loss of neurons leading to dementia. Because hippocampal neurogenesis is linked to functions such as learning, memory and mood, there has been great interest in examining the effects of AD on hippocampal neurogenesis. This article reviews the pertinent studies and tries to unite them in one possible disease model. Early in the disease, oligomeric Abeta may transiently promote the generation of immature neurons from neural stem cells (NSCs). However, reduced concentrations of multiple neurotrophic factors and higher levels of fibroblast growth factor-2 seem to induce a developmental arrest of newly generated neurons. Furthermore, fibrillary Abeta and down-regulation of oligodendrocyte-lineage transcription factor-2 (OLIG2) may cause the death of these nonfunctional neurons. Therefore, altering the brain microenvironment for fostering apt maturation of graft-derived neurons may be critical for improving the efficacy of NSC transplantation therapy for AD.

  16. Cellular microenvironment modulates the galvanotaxis of brain tumor initiating cells.

    PubMed

    Huang, Yu-Ja; Hoffmann, Gwendolyn; Wheeler, Benjamin; Schiapparelli, Paula; Quinones-Hinojosa, Alfredo; Searson, Peter

    2016-02-22

    Galvanotaxis is a complex process that represents the collective outcome of various contributing mechanisms, including asymmetric ion influxes, preferential activation of voltage-gated channels, and electrophoretic redistribution of membrane components. While a large number of studies have focused on various up- and downstream signaling pathways, little is known about how the surrounding microenvironment may interact and contribute to the directional response. Using a customized galvanotaxis chip capable of carrying out experiments in both two- and three-dimensional microenvironments, we show that cell-extracellular matrix (ECM) interactions modulate the galvanotaxis of brain tumor initiating cells (BTICs). Five different BTICs across three different glioblastoma subtypes were examined and shown to all migrate toward the anode in the presence of a direct-current electric field (dcEF) when cultured on a poly-L-ornithine/laminin coated surface, while the fetal-derived neural progenitor cells (fNPCs) migrated toward the cathode. Interestingly, when embedded in a 3D ECM composed of hyaluronic acid and collagen, BTICs exhibited opposite directional response and migrated toward the cathode. Pharmacological inhibition against a panel of key molecules involved in galvanotaxis further revealed the mechanistic differences between 2- and 3D galvanotaxis in BTICs. Both myosin II and phosphoinositide 3-kinase (PI3K) were found to hold strikingly different roles in different microenvironments.

  17. Analysis of Spine Motility of Newborn Granule Cells in Acute Brain Slices.

    PubMed

    Tashiro, Ayumu; Zhao, Chunmei; Suh, Hoonkyo; Gage, Fred H

    2015-10-01

    In this protocol, acute brain slices are prepared from mice in which newborn granule cells have been labeled using retroviral vector technology. Using a live-cell imaging stage and confocal microscopy coupled to imaging software, dendritic spines are analyzed.

  18. Urine Preservative

    NASA Technical Reports Server (NTRS)

    Smith, Scott M. (Inventor); Nillen, Jeannie (Inventor)

    2001-01-01

    Disclosed is CPG, a combination of a chlorhexidine salt (such as chlorhexidine digluconate, chlorhexidine diacetate, or chlorhexidine dichloride) and n-propyl gallate that can be used at ambient temperatures as a urine preservative.

  19. Tight junction protein expression and barrier properties of immortalized mouse brain microvessel endothelial cells.

    PubMed

    Brown, Rachel C; Morris, Andrew P; O'Neil, Roger G

    2007-01-26

    Understanding the molecular and biochemical mechanisms regulating the blood-brain barrier is aided by in vitro model systems. Many studies have used primary cultures of brain microvessel endothelial cells for this purpose. However, primary cultures limit the generation of material for molecular and biochemical assays since cells grow slowly, are prone to contamination by other neurovascular unit cells, and lose blood-brain barrier characteristics when passaged. To address these issues, immortalized cell lines have been generated. In these studies, we assessed the suitability of the immortalized mouse brain endothelial cell line, bEnd3, as a blood-brain barrier model. RT-PCR and immunofluorescence indicated expression of multiple tight junction proteins. bEnd3 cells formed barriers to radiolabeled sucrose, and responded like primary cultures to disrupting stimuli. Exposing cells to serum-free media on their basolateral side significantly decreased paracellular permeability; astrocyte-conditioned media did not enhance barrier properties. The serum-free media-induced decrease in permeability was correlated with an increase in claudin-5 and zonula occludens-1 immunofluorescence at cell-cell contracts. We conclude that bEnd3 cells are an attractive candidate as a model of the blood-brain barrier due to their rapid growth, maintenance of blood-brain barrier characteristics over repeated passages, formation of functional barriers and amenability to numerous molecular interventions.

  20. Suppression of Brain Mast Cells Degranulation Inhibits Microglial Activation and Central Nervous System Inflammation.

    PubMed

    Dong, Hongquan; Zhang, Xiang; Wang, Yiming; Zhou, Xiqiao; Qian, Yanning; Zhang, Shu

    2017-03-01

    Brain inflammation has a critical role in the pathophysiology of brain diseases. Microglia, the resident immune cells in the brain, play an important role in brain inflammation, while brain mast cells are the "first responder" in the injury rather than microglia. Functional aspects of mast cell-microglia interactions remain poorly understood. Our results demonstrated that site-directed injection of the "mast cell degranulator" compound 48/80 (C48/80) in the hypothalamus induced mast cell degranulation, microglial activation, and inflammatory factor production, which initiated the acute brain inflammatory response. "Mast cell stabilizer" disodium cromoglycate (cromolyn) inhibited this effect, including decrease of inflammatory cytokines, reduced microglial activation, inhibition of MAPK and AKT pathways, and repression of protein expression of histamine receptor 1 (H1R), histamine receptor 4 (H4R), protease-activated receptor 2 (PAR2), and toll-like receptor 4 (TLR4) in microglia. We also demonstrated that C48/80 had no effect on microglial activation in mast cell-deficient Kit(W-sh/W-sh) mice. These results implicate that activated brain mast cells trigger microglial activation and stabilization of mast cell inhibits microglial activation-induced central nervous system (CNS) inflammation. Interactions between mast cells and microglia could constitute a new and unique therapeutic target for CNS immune inflammation-related diseases.

  1. Intracerebral transplants of primary muscle cells: a potential 'platform' for transgene expression in the brain

    NASA Technical Reports Server (NTRS)

    Jiao, S.; Schultz, E.; Wolff, J. A.

    1992-01-01

    After the transplantation of rat primary muscle cells into the caudate or cortex of recipient rats, the muscle cells were able to persist for at least 6 months. Muscle cells transfected with expression plasmids prior to transplantation were able to express reporter genes in the brains for at least 2 months. These results suggest that muscle cells might be a useful 'platform' for transgene expression in the brain.

  2. Single cell transcriptomics reveals specific RNA editing signatures in the human brain.

    PubMed

    Picardi, Ernesto; Horner, David Stephen; Pesole, Graziano

    2017-03-03

    While RNA editing by A-to-I deamination is a requisite for neuronal function in humans, it is under investigated in single cells. Here we fill this gap by analysing RNA editing profiles of single cells from the brain cortex of living human subjects. We show that RNA editing levels per cell are bimodally distributed and distinguish between major brain cell types thus providing new insights into neuronal dynamics.

  3. Intravenous transplantation of bone marrow mesenchymal stem cells promotes neural regeneration after traumatic brain injury.

    PubMed

    Anbari, Fatemeh; Khalili, Mohammad Ali; Bahrami, Ahmad Reza; Khoradmehr, Arezoo; Sadeghian, Fatemeh; Fesahat, Farzaneh; Nabi, Ali

    2014-05-01

    To investigate the supplement of lost nerve cells in rats with traumatic brain injury by intravenous administration of allogenic bone marrow mesenchymal stem cells, this study established a Wistar rat model of traumatic brain injury by weight drop impact acceleration method and administered 3 × 10(6) rat bone marrow mesenchymal stem cells via the lateral tail vein. At 14 days after cell transplantation, bone marrow mesenchymal stem cells differentiated into neurons and astrocytes in injured rat cerebral cortex and rat neurological function was improved significantly. These findings suggest that intravenously administered bone marrow mesenchymal stem cells can promote nerve cell regeneration in injured cerebral cortex, which supplement the lost nerve cells.

  4. Intravenous transplantation of bone marrow mesenchymal stem cells promotes neural regeneration after traumatic brain injury

    PubMed Central

    Anbari, Fatemeh; Khalili, Mohammad Ali; Bahrami, Ahmad Reza; Khoradmehr, Arezoo; Sadeghian, Fatemeh; Fesahat, Farzaneh; Nabi, Ali

    2014-01-01

    To investigate the supplement of lost nerve cells in rats with traumatic brain injury by intravenous administration of allogenic bone marrow mesenchymal stem cells, this study established a Wistar rat model of traumatic brain injury by weight drop impact acceleration method and administered 3 × 106 rat bone marrow mesenchymal stem cells via the lateral tail vein. At 14 days after cell transplantation, bone marrow mesenchymal stem cells differentiated into neurons and astrocytes in injured rat cerebral cortex and rat neurological function was improved significantly. These findings suggest that intravenously administered bone marrow mesenchymal stem cells can promote nerve cell regeneration in injured cerebral cortex, which supplement the lost nerve cells. PMID:25206912

  5. SOX9 Maintains Reserve Stem Cells and Preserves Radioresistance in Mouse Small Intestine

    PubMed Central

    Roche, Kyle C.; Gracz, Adam D.; Liu, Xiao Fu; Newton, Victoria; Akiyama, Haruhiko; Magness, Scott T.

    2015-01-01

    BACKGROUND & AIMS Reserve intestinal stem cells (rISCs) are quiescent/slowly cycling under homeostatic conditions, allowing for their identification with label-retention assays. rISCs mediate epithelial regeneration after tissue damage by converting to actively proliferating stem cells (aISCs) that self renew and demonstrate multipotency, which are defining properties of stem cells. Little is known about the genetic mechanisms that regulate the production and maintenance of rISCs. High expression levels of the transcription factor Sox9 (Sox9high) are associated with rISCs. This study investigates the role of SOX9 in regulating the rISC state. METHODS We used fluorescence-activated cell sorting to isolate cells defined as aISCs (Lgr5high) and rISCs (Sox9high) from Lgr5EGFP and Sox9EGFP reporter mice. Expression of additional markers associated with active and reserve ISCs were assessed in Lgr5high and Sox9high populations by single-cell gene expression analyses. We used label-retention assays to identify whether Sox9high cells were label-retatining cells (LRCs). Lineage-tracing experiments were performed in Sox9-CreERT2 mice to measure the stem cell capacities and radioresistance of Sox9-expressing cells. Conditional SOX9 knockout mice and inducible-conditional SOX9 knockout mice were used to determine whether SOX9 was required to maintain LRCs and rISC function. RESULTS Lgr5high and a subset of crypt-based Sox9high cells co-express markers of aISC and rISC (Lgr5. Bmi1. Lrig1, and Hopx). LRCs express high levels of Sox9 and are lost in SOX9-knockout mice. SOX9 is required for epithelial regeneration after high-dose irradiation. Crypts from SOX9-knockout mice have increased sensitivity to radiation, compared with control mice, which could not be attributed to impaired cell-cycle arrest or DNA repair. CONCLUSIONS SOX9 limits proliferation in LRCs and imparts radiation resistance to rISCs in mice. PMID:26170137

  6. Intracellular pathways regulating ciliary beating of rat brain ependymal cells

    PubMed Central

    Nguyen, Thien; Chin, Wei-Chun; O’Brien, Jennifer A; Verdugo, Pedro; Berger, Albert J

    2001-01-01

    The mammalian brain ventricles are lined with ciliated ependymal cells. As yet little is known about the mechanisms by which neurotransmitters regulate cilia beat frequency (CBF). Application of 5-HT to ependymal cells in cultured rat brainstem slices caused CBF to increase. 5-HT had an EC50 of 30 μM and at 100 μM attained a near-maximal CBF increase of 52.7 ± 4.1 % (mean ± s.d.) (n= 8). Bathing slices in Ca2+-free solution markedly reduced the 5-HT-mediated increase in CBF. Fluorescence measurements revealed that 5-HT caused a marked transient elevation in cytosolic Ca2+ ([Ca2+]c) that then slowly decreased to a plateau level. Analysis showed that the [Ca2+]c transient was due to release of Ca2+ from inositol 1,4,5-trisphosphate (IP3)-sensitive stores; the plateau was probably due to extracellular Ca2+ influx through Ca2+ release-activated Ca2+ (CRAC) channels. Application of ATP caused a sustained decrease in CBF. ATP had an EC50 of about 50 μM and 100 μM ATP resulted in a maximal 57.5 ± 6.5 % (n= 12) decrease in CBF. The ATP-induced decrease in CBF was unaffected by lowering extracellular [Ca2+], and no changes in [Ca2+]c were observed. Exposure of ependymal cells to forskolin caused a decrease in CBF. Ciliated ependymal cells loaded with caged cAMP exhibited a 54.3 ± 7.5 % (n= 9) decrease in CBF following uncaging. These results suggest that ATP reduces CBF by a Ca2+-independent cAMP-mediated pathway. Application of 5-HT and adenosine-5′-O-3-thiotriphosphate (ATP-γ-S) to acutely isolated ciliated ependymal cells resulted in CBF responses similar to those of ependymal cells in cultured slices suggesting that these neurotransmitters act directly on these cells. The opposite response of ciliated ependymal cells to 5-HT and ATP provides a novel mechanism for their active involvement in central nervous system signalling. PMID:11179397

  7. Capsule independent uptake of the fungal pathogen Cryptococcus neoformans into brain microvascular endothelial cells.

    PubMed

    Sabiiti, Wilber; May, Robin C

    2012-01-01

    Cryptococcosis is a life-threatening fungal disease with a high rate of mortality among HIV/AIDS patients across the world. The ability to penetrate the blood-brain barrier (BBB) is central to the pathogenesis of cryptococcosis, but the way in which this occurs remains unclear. Here we use both mouse and human brain derived endothelial cells (bEnd3 and hCMEC/D3) to accurately quantify fungal uptake and survival within brain endothelial cells. Our data indicate that the adherence and internalisation of cryptococci by brain microvascular endothelial cells is an infrequent event involving small numbers of cryptococcal yeast cells. Interestingly, this process requires neither active signalling from the fungus nor the presence of the fungal capsule. Thus entry into brain microvascular endothelial cells is most likely a passive event that occurs following 'trapping' within capillary beds of the BBB.

  8. Growth Factor erv1-like Modulates Drp1 to Preserve Mitochondrial Dynamics and Function in Mouse Embryonic Stem Cells

    PubMed Central

    Todd, Lance R.; Damin, Matthew N.; Gomathinayagam, Rohini; Horn, Sarah R.; Means, Anthony R.

    2010-01-01

    The relationship of mitochondrial dynamics and function to pluripotency are rather poorly understood aspects of stem cell biology. Here we show that growth factor erv1-like (Gfer) is involved in preserving mouse embryonic stem cell (ESC) mitochondrial morphology and function. Knockdown (KD) of Gfer in ESCs leads to decreased pluripotency marker expression, embryoid body (EB) formation, cell survival, and loss of mitochondrial function. Mitochondria in Gfer-KD ESCs undergo excessive fragmentation and mitophagy, whereas those in ESCs overexpressing Gfer appear elongated. Levels of the mitochondrial fission GTPase dynamin-related protein 1 (Drp1) are highly elevated in Gfer-KD ESCs and decreased in Gfer-overexpressing cells. Treatment with a specific inhibitor of Drp1 rescues mitochondrial function and apoptosis, whereas expression of Drp1-dominant negative resulted in the restoration of pluripotency marker expression in Gfer-KD ESCs. Altogether, our data reveal a novel prosurvival role for Gfer in maintaining mitochondrial fission–fusion dynamics in pluripotent ESCs. PMID:20147447

  9. Growth factor erv1-like modulates Drp1 to preserve mitochondrial dynamics and function in mouse embryonic stem cells.

    PubMed

    Todd, Lance R; Damin, Matthew N; Gomathinayagam, Rohini; Horn, Sarah R; Means, Anthony R; Sankar, Uma

    2010-04-01

    The relationship of mitochondrial dynamics and function to pluripotency are rather poorly understood aspects of stem cell biology. Here we show that growth factor erv1-like (Gfer) is involved in preserving mouse embryonic stem cell (ESC) mitochondrial morphology and function. Knockdown (KD) of Gfer in ESCs leads to decreased pluripotency marker expression, embryoid body (EB) formation, cell survival, and loss of mitochondrial function. Mitochondria in Gfer-KD ESCs undergo excessive fragmentation and mitophagy, whereas those in ESCs overexpressing Gfer appear elongated. Levels of the mitochondrial fission GTPase dynamin-related protein 1 (Drp1) are highly elevated in Gfer-KD ESCs and decreased in Gfer-overexpressing cells. Treatment with a specific inhibitor of Drp1 rescues mitochondrial function and apoptosis, whereas expression of Drp1-dominant negative resulted in the restoration of pluripotency marker expression in Gfer-KD ESCs. Altogether, our data reveal a novel prosurvival role for Gfer in maintaining mitochondrial fission-fusion dynamics in pluripotent ESCs.

  10. Cytotoxic effects of aflatoxin B1 on human brain microvascular endothelial cells of the blood-brain barrier.

    PubMed

    Qureshi, Humaira; Hamid, Saeed S; Ali, Syed Shayan; Anwar, Javeria; Siddiqui, Anwar Ali; Khan, Naveed Ahmed

    2015-05-01

    Aflatoxins are mycotoxins produced by Aspergillus spp. Although AFB1 is implicated as a carcinogen in hepatocellular carcinoma, brain autopsies in affected areas have revealed its presence in 81% of cases. Given its haematogenous spread, here we determined the cytotoxic effects of AFB1 on primary human brain microvascular endothelial cells (HBMEC), which constitute the blood-brain barrier, human umbilical vein endothelial cells (HUVEC) as well as immortalized epithelial cells of human hepatocellular carcinoma (Huh7). The cell types were exposed to AFB1 (3-32 nM) for 24 h and release of lactate dehydrogenase was measured as cell cytotoxicity marker. Furthermore, DNA was collected from both cell types and DNA adduct formation was determined by immunoblot using anti-AFB1-DNA adduct antibody. At 32 nM, AFB1 killed >85% HBMEC, while controls showed minimal effects (P < .05). Similar concentrations of AFB1 showed 22% cell death of HUVEC, while the same concentration did not kill Huh7. At low concentrations, in other words, 3.2 nM, AFB1 produced DNA adduct formation in HBMEC, while high concentration (32 nM) did not form DNA adducts. For HUVEC, 16 nM and 32 nM exhibited DNA adduct formation. For Huh7, 3.2 nM did not form DNA adducts, while 32 nM exhibited DNA adduct formation. For the first time, we report that AFB1 affected the viability of primary endothelial cells but not immortalized Huh7 cells. Cytotoxicity of brain endothelial cells suggests extra-hepatic complications post-AFB1 exposure.

  11. [Molecular cytogenetic methods for studying interphase chromosomes in human brain cells].

    PubMed

    Iurov, I Iu; Vorsanova, S G; Solov'ev, I V; Iurov, Iu B

    2010-09-01

    One of the main genetic factors determining the functional activity of the genome in somatic cells, including brain nerve cells, is the spatial organization of chromosomes in the interphase nucleus. For a long time, no studies of human brain cells were carried out until high-resolution methods of molecular cytogenetics were developed to analyze interphase chromosomes in nondividing somatic cells. The purpose of the present work was to assess the potential of high-resolution methods of interphase molecular cytogenetics for studying chromosomes and the nuclear organization in postmitotic brain cells. A high efficiency was shown by such methods as multiprobe and quantitative fluorescence in situ hybridization (Multiprobe FISH and QFISH), ImmunoMFISH (analysis of the chromosome organization in different types of brain cells), and interphase chromosome-specific multicolor banding (ICS-MCB). These approaches allowed studying the nuclear organization depending on the gene composition and types of repetitive DNA of specific chromosome regions in certain types of brain cells (in neurons and glial cells, in particular). The present work demonstrates a high potential of interphase molecular cytogenetics for studying the structural and functional organizations of the cell nucleus in highly differentiated nerve cells. Analysis of interphase chromosomes of brain cells in the normal and pathological states can be considered as a promising line of research in modern molecular cytogenetics and cell neurobiology, i. e., molecular neurocytogenetics.

  12. [Stem Cells in the Brain of Mammals and Human: Fundamental and Applied Aspects].

    PubMed

    Aleksandrova, M A; Marey, M V

    2015-01-01

    Brain stem cells represent an extremely intriguing phenomenon. The aim of our review is to present an integrity vision of their role in the brain of mammals and humans, and their clinical perspectives. Over last two decades, investigations of biology of the neural stem cells produced significant changes in general knowledge about the processes of development and functioning of the brain. Researches on the cellular and molecular mechanisms of NSC differentiation and behavior led to new understanding of their involvement in learning and memory. In the regenerative medicine, original therapeutic approaches to neurodegenerative brain diseases have been elaborated due to fundamental achievements in this field. They are based on specific regenerative potential of neural stem cells and progenitor cells, which possess the ability to replace dead cells and express crucially significant biologically active factors that are missing in the pathological brain. For the needs of cell substitution therapy in the neural diseases, adequate methods of maintaining stem cells in culture and their differentiation into different types of neurons and glial cells, have been developed currently. The success of modern cellular technologies has significantly expanded the range of cells used for cell therapy. The near future may bring new perspective and distinct progress in brain cell therapy due to optimizing the cells types most promising for medical needs.

  13. Gain of glucose-independent growth upon metastasis of breast cancer cells to the brain.

    PubMed

    Chen, Jinyu; Lee, Ho-Jeong; Wu, Xuefeng; Huo, Lei; Kim, Sun-Jin; Xu, Lei; Wang, Yan; He, Junqing; Bollu, Lakshmi R; Gao, Guang; Su, Fei; Briggs, James; Liu, Xiaojing; Melman, Tamar; Asara, John M; Fidler, Isaiah J; Cantley, Lewis C; Locasale, Jason W; Weihua, Zhang

    2015-02-01

    Breast cancer brain metastasis is resistant to therapy and a particularly poor prognostic feature in patient survival. Altered metabolism is a common feature of cancer cells, but little is known as to what metabolic changes benefit breast cancer brain metastases. We found that brain metastatic breast cancer cells evolved the ability to survive and proliferate independent of glucose due to enhanced gluconeogenesis and oxidations of glutamine and branched chain amino acids, which together sustain the nonoxidative pentose pathway for purine synthesis. Silencing expression of fructose-1,6-bisphosphatases (FBP) in brain metastatic cells reduced their viability and improved the survival of metastasis-bearing immunocompetent hosts. Clinically, we showed that brain metastases from human breast cancer patients expressed higher levels of FBP and glycogen than the corresponding primary tumors. Together, our findings identify a critical metabolic condition required to sustain brain metastasis and suggest that targeting gluconeogenesis may help eradicate this deadly feature in advanced breast cancer patients.

  14. Changing numbers of neuronal and non-neuronal cells underlie postnatal brain growth in the rat

    PubMed Central

    Bandeira, Fabiana; Lent, Roberto; Herculano-Houzel, Suzana

    2009-01-01

    The rat brain increases >6× in mass from birth to adulthood, presumably through the addition of glial cells and increasing neuronal size, without the addition of neurons. To test this hypothesis, here we investigate quantitatively the postnatal changes in the total number of neuronal and non-neuronal cells in the developing rat brain, and examine how these changes correlate with brain growth. Total numbers of cells were determined with the isotropic fractionator in the brains of 53 Wistar rats, from birth to young adulthood. We find that at birth, >90% of the cells in the rat brain are neurons. Following a dormant period of ≈3 days after birth, the net number of neurons in the cerebral cortex, hippocampus, and remaining tissue (excluding cerebellum and olfactory bulb) doubles during the first week, then is reduced by 70% during the second postnatal week, concurrently with net gliogenesis. A second round of net addition of 6 million neurons is observed in the cerebral cortex over the following 2 weeks. During the first postnatal week, brain growth relates mainly to increased numbers of neurons of larger average size. In the second and third weeks, it correlates with increased numbers of non-neuronal cells that are smaller in size than the preexisting neurons. Postnatal rat brain development is thus characterized by dramatic changes in the cellular composition of the brain, whose growth is governed by different combinations of cell addition and loss, and changes in average cell size during the first months after birth. PMID:19666520

  15. Calpain 2 Is Required for the Invasion of Glioblastoma Cells in the Zebrafish Brain Microenvironment

    PubMed Central

    Lal, Sangeet; La Du, Jane; Tanguay, Robert L.; Greenwood, Jeffrey A.

    2012-01-01

    Glioblastoma is an aggressive primary brain tumor with a 5-year survival rate of less than 5%. The ability of glioblastoma cells to invade surrounding brain tissue presents the primary challenge for the success of focal therapeutic approaches. We previously reported that the calcium-activated protease calpain 2 is critical for glioblastoma cell invasion in vitro. Here, we show that expression of calpain 2 is required for the dispersal of glioblastoma cells in a living brain microenvironment. Knockdown of calpain 2 resulted in a 2.9-fold decrease in the invasion of human glioblastoma cells in zebrafish brain. Control cells diffusely migrated up to 450 μm from the site of injection, whereas knockdown cells remained confined in clusters. The invasion study was repeated in organotypic mouse brain tissues, and calpain 2 knockdown cells demonstrated a 2.3-fold lower area of dispersal compared with control cells. In zebrafish brain, glioblastoma cells appeared to migrate in part along the blood vessels of the host. Furthermore, angiogenesis was detected in 27% of zebrafish injected with control cells, whereas only 12.5% of fish receiving knockdown cells showed the formation of new vessels, suggesting a role for calpain 2 in tumor cell angiogenesis. Consistent with the progression of glioblastoma in humans, transplanted tumor cells were not observed to metastasize outside the brain of zebrafish. This study demonstrates that calpain 2 expression is required for the dispersal of glioblastoma cells within the dynamic microenvironment of the brain, identifying zebrafish as a valuable orthotopic system for studying glioblastoma cell invasion. PMID:22183788

  16. Closely related T-memory stem cells correlate with in vivo expansion of CAR.CD19-T cells and are preserved by IL-7 and IL-15.

    PubMed

    Xu, Yang; Zhang, Ming; Ramos, Carlos A; Durett, April; Liu, Enli; Dakhova, Olga; Liu, Hao; Creighton, Chad J; Gee, Adrian P; Heslop, Helen E; Rooney, Cliona M; Savoldo, Barbara; Dotti, Gianpietro

    2014-06-12

    Adoptive transfer of T lymphocytes expressing a CD19-specific chimeric antigen receptor (CAR.CD19) induces complete tumor regression in patients with lymphoid malignancies. Although in vivo persistence of CAR-T cells correlates with clinical responses, it remains unknown whether specific cell subsets within the CAR-T-cell product correlate with their subsequent in vivo expansion and persistence. We analyzed 14 patients with B-cell malignancies infused with autologous CAR.CD19-redirected T cells expanded ex vivo using IL-2, and found that their in vivo expansion only correlated with the frequency within the infused product of a CD8(+)CD45RA(+)CCR7(+) subset, whose phenotype is closest to "T-memory stem cells." Preclinical models showed that increasing the frequency of CD8(+)CD45RA(+)CCR7(+) CAR-T cells in the infused line by culturing the cells with IL-7 and IL-15 produced greater antitumor activity of CAR-T cells mediated by increased resistance to cell death, following repetitive encounters with the antigen, while preserving their migration to secondary lymphoid organs. This trial was registered at www.clinicaltrials.gov as #NCT00586391 and #NCT00709033.

  17. Limb Preservation in Recurrent Giant Cell Tumour of Distal End of Radius with Fibular Graft Fracture: Role of Ulnocarpal Arthrodesis.

    PubMed

    Kumar, Narinder

    2015-01-01

    Giant cell tumors of distal radius are locally aggressive tumors with a high rate of recurrence. Though surgery remains the mainstay of treatment, reconstruction remains a challenge in cases of recurrence. Recurrences of GCT in autogenous fibular grafts have been rarely reported and pathological fractures through such grafts are even rarer. Ulnocarpal arthrodesis has never been described as a limb preservation procedure in such a recurrent lesion in distal radius with pathological fracture through a well incorporated fibular graft. A case of pathological fracture in a well incorporated autogenous non-vascularized fibular bone graft in recurrent GCT of distal radius and its successful management with ulnocarpal arthrodesis is reported. In such a scenario where other reconstructive options like allograft or prosthetic reconstructions are not likely to succeed, ulnocarpal arthrodesis may be considered as a salvage procedure.

  18. Brain Cancer Stem Cells in Adults and Children: Cell Biology and Therapeutic Implications.

    PubMed

    Abou-Antoun, Tamara J; Hale, James S; Lathia, Justin D; Dombrowski, Stephen M

    2017-04-03

    Brain tumors represent some of the most malignant cancers in both children and adults. Current treatment options target the majority of tumor cells but do not adequately target self-renewing cancer stem cells (CSCs). CSCs have been reported to resist the most aggressive radiation and chemotherapies, and give rise to recurrent, treatment-resistant secondary malignancies. With advancing technologies, we now have a better understanding of the genetic, epigenetic and molecular signatures and microenvironmental influences which are useful in distinguishing between distinctly different tumor subtypes. As a result, efforts are now underway to identify and target CSCs within various tumor subtypes based on this foundation. This review discusses progress in CSC biology as it relates to targeted therapies which may be uniquely different between pediatric and adult brain tumors. Studies to date suggest that pediatric brain tumors may benefit more from genetic and epigenetic targeted therapies, while combination treatments aimed specifically at multiple molecular pathways may be more effective in treating adult brain tumors which seem to have a greater propensity towards microenvironmental interactions. Ultimately, CSC targeting approaches in combination with current clinical therapies have the potential to be more effective owing to their ability to compromise CSCs maintenance and the mechanisms which underlie their highly aggressive and deadly nature.

  19. Bone Marrow–Derived Mesenchymal Stem Cells Enhance Bacterial Clearance and Preserve Bioprosthetic Integrity in a Model of Mesh Infection

    PubMed Central

    Criman, Erik T.; Kurata, Wendy E.; Matsumoto, Karen W.; Aubin, Harry T.; Campbell, Carmen E.

    2016-01-01

    Background: The reported incidence of mesh infection in contaminated operative fields is as high as 30% regardless of the material used. Recently, mesenchymal stem cells (MSCs) have been shown to possess favorable immunomodulatory properties and improve tissue incorporation when seeded onto bioprosthetics. The aim of this study was to evaluate whether seeding noncrosslinked bovine pericardium (Veritas Collagen Matrix) with allogeneic bone marrow–derived MSCs improves infection resistance in vivo after inoculation with Escherichia coli (E. coli). Methods: Rat bone marrow–derived MSCs at passage 3 were seeded onto bovine pericardium and cultured for 7 days before implantation. Additional rats (n = 24) were implanted subcutaneously with MSC-seeded or unseeded mesh and inoculated with 7 × 105 colony-forming units of E. coli or saline before wound closure (group 1, unseeded mesh/saline; group 2, unseeded mesh/E. coli; group 3, MSC-seeded mesh/E. coli; 8 rats per group). Meshes were explanted at 4 weeks and underwent microbiologic and histologic analyses. Results: MSC-seeded meshes inoculated with E. coli demonstrated superior bacterial clearance and preservation of mesh integrity compared with E. coli–inoculated unseeded meshes (87.5% versus 0% clearance; p = 0.001). Complete mesh degradation concurrent with abscess formation was observed in 100% of rats in the unseeded/E. coli group, which is in contrast to 12.5% of rats in the MSC-seeded/E. coli group. Histologic evaluation determined that remodeling characteristics of E. coli–inoculated MSC-seeded meshes were similar to those of uninfected meshes 4 weeks after implantation. Conclusions: Augmenting a bioprosthetic material with stem cells seems to markedly enhance resistance to bacterial infection in vivo and preserve mesh integrity. PMID:27482490

  20. A bioengineered niche preserves the quiescence of muscle stem cells and enhances their therapeutic efficacy

    PubMed Central

    Quarta, Marco; Brett, Jamie O.; DiMarco, Rebecca; De Morree, Antoine; Boutet, Stephane C.; Chacon, Robert; Gibbons, Michael C.; Garcia, Victor A.; Su, James; Shrager, Joseph B.; Heilshorn, Sarah; Rando, Thomas A.

    2016-01-01

    A promising therapeutic strategy for diverse genetic disorders involves transplantation of autologous stem cells that have been genetically corrected ex vivo. A major challenge in such approaches is a loss of stem cell potency during stem cell culture. Here we describe a system for maintaining muscle stem cells (MuSCs) in vitro in a potent, quiescent state. Using a machine learning method, we identified a molecular signature of quiescence and used it to screen for factors that could maintain mouse MuSC quiescence, thus defining a quiescence medium (QM). We also designed artificial muscle fibers (AMFs) that mimic the native myofiber of the MuSC niche. Mouse MuSCs maintained in QM on AMFs showed enhanced potential for engraftment, tissue regeneration and self-renewal after transplantation in mice. An artificial niche adapted to human MuSCs showed similarly prolonged quiescence in vitro and enhanced potency in vivo. Our approach for maintaining quiescence may be applicable to stem cells from a range of other tissues. PMID:27240197

  1. Renal cell carcinoma in a transplanted kidney: successful organ-preserving procedure.

    PubMed

    Höppner, W; Grosse, K; Dreikorn, K

    1996-01-01

    We report a case of a de novo renal cell carcinoma in a transplanted kidney, which was detected 3 years after the transplantation. The tumor was excised under hypothermia and perfusion. Immunosuppression was not stopped and the function remained excellent. Close-mesh follow-up of 45 months showed no evidence or recurrence or metastasis.

  2. Human iPS cell-derived astrocyte transplants preserve respiratory function after spinal cord injury

    PubMed Central

    Li, Ke; Javed, Elham; Scura, Daniel; Hala, Tamara J.; Seetharam, Suneil; Falnikar, Aditi; Richard, Jean-Philippe; Chorath, Ashley; Maragakis, Nicholas J.; Wright, Megan C.; Lepore, Angelo C.

    2015-01-01

    Transplantation-based replacement of lost and/or dysfunctional astrocytes is a promising therapy for spinal cord injury (SCI) that has not been extensively explored, despite the integral roles played by astrocytes in the central nervous system (CNS). Induced pluripotent stem (iPS) cells are a clinically-relevant source of pluripotent cells that both avoid ethical issues of embryonic stem cells and allow for homogeneous derivation of mature cell types in large quantities, potentially in an autologous fashion. Despite their promise, the iPS cell field is in its infancy with respect to evaluating in vivo graft integration and therapeutic efficacy in SCI models. Astrocytes express the major glutamate transporter, GLT1, which is responsible for the vast majority of glutamate uptake in spinal cord. Following SCI, compromised GLT1 expression/function can increase susceptibility to excitotoxicity. We therefore evaluated intraspinal transplantation of human iPS cell-derived astrocytes (hIPSAs) following cervical contusion SCI as a novel strategy for reconstituting GLT1 expression and for protecting diaphragmatic respiratory neural circuitry. Transplant-derived cells showed robust long-term survival post-injection and efficiently differentiated into astrocytes in injured spinal cord of both immunesuppressed mice and rats. However, the majority of transplant-derived astrocytes did not express high levels of GLT1, particularly at early times post-injection. To enhance their ability to modulate extracellular glutamate levels, we engineered hIPSAs with lentivirus to constitutively express GLT1. Overexpression significantly increased GLT1 protein and functional GLT1-mediated glutamate uptake levels in hIPSAs both in vitro and in vivo post-transplantation. Compared to human fibroblast control and unmodified hIPSA transplantation, GLT1-overexpressing hIPSAs reduced (1) lesion size within the injured cervical spinal cord, (2) morphological denervation by respiratory phrenic motor

  3. Human neural stem cells can target and deliver therapeutic genes to breast cancer brain metastases.

    PubMed

    Joo, Kyeung Min; Park, In H; Shin, Ji Y; Jin, Juyoun; Kang, Bong Gu; Kim, Mi Hyun; Lee, Se Jeong; Jo, Mi-young; Kim, Seung U; Nam, Do-Hyun

    2009-03-01

    The tumor-tropic properties of neural stem cells (NSCs) led to the development of a novel strategy for delivering therapeutic genes to tumors in the brain. To apply this strategy to the treatment of brain metastases, we made a human NSC line expressing cytosine deaminase (F3.CD), which converts 5-fluorocytosine (5-FC) into 5-fluorouracil, an anticancer agent. In vitro, the F3.CD cells significantly inhibited the growth of tumor cell lines in the presence of the prodrug 5-FC. In vivo, MDA-MB-435 human breast cancer cells were implanted into the brain of immune-deficient mouse stereotactically, and F3.CD cells were injected into the contralateral hemisphere followed by systemic 5-FC administration. The F3.CD cells migrated selectively into the brain metastases located in the opposite hemisphere and resulted in significantly reduced volumes. The F3.CD and 5-FC treatment also decreased both tumor volume and number of tumor mass significantly, when immune-deficient mouse had MDA-MB-435 cells injected into the internal carotid artery and F3.CD cells were transplanted into the contralateral brain hemisphere stereotactically. Taken together, brain transplantation of human NSCs, encoding the suicide enzyme CD, combined with systemic administration of the prodrug 5-FC, is an effective treatment regimen for brain metastases of tumors.

  4. The SOX2-interactome in brain cancer cells identifies the requirement of MSI2 and USP9X for the growth of brain tumor cells.

    PubMed

    Cox, Jesse L; Wilder, Phillip J; Gilmore, Joshua M; Wuebben, Erin L; Washburn, Michael P; Rizzino, Angie

    2013-01-01

    Medulloblastomas and glioblastomas, the most common primary brain tumors in children and adults, respectively, are extremely difficult to treat. Efforts to identify novel proteins essential for the growth of these tumors may help to further our understanding of the biology of these tumors, as well as, identify targets for future therapies. The recent identification of multiple transcription factor-centric protein interaction landscapes in embryonic stem cells has identified numerous understudied proteins that are essential for the self-renewal of these stem cells. To identify novel proteins essential for the fate of brain tumor cells, we examined the protein interaction network of the transcription factor, SOX2, in medulloblastoma cells. For this purpose, Multidimensional Protein Identification Technology (MudPIT) identified >280 SOX2-associated proteins in the medulloblastoma cell line DAOY. To begin to understand the roles of SOX2-associated proteins in brain cancer, we focused on two SOX2-associated proteins, Musashi 2 (MSI2) and Ubiquitin Specific Protease 9x (USP9X). Recent studies have implicated MSI2, a putative RNA binding protein, and USP9X, a deubiquitinating enzyme, in several cancers, but not brain tumors. We demonstrate that knockdown of MSI2 significantly reduces the growth of DAOY cells as well as U87 and U118 glioblastoma cells. We also demonstrate that the knockdown of USP9X in DAOY, U87 and U118 brain tumor cells strongly reduces their growth. Together, our studies identify a large set of SOX2-associated proteins in DAOY medulloblastoma cells and identify two proteins, MSI2 and USP9X, that warrant further investigation to determine whether they are potential therapeutic targets for brain cancer.

  5. Predators inhibit brain cell proliferation in natural populations of electric fish, Brachyhypopomus occidentalis.

    PubMed

    Dunlap, Kent D; Tran, Alex; Ragazzi, Michael A; Krahe, Rüdiger; Salazar, Vielka L

    2016-02-10

    Compared with laboratory environments, complex natural environments promote brain cell proliferation and neurogenesis. Predators are one important feature of many natural environments, but, in the laboratory, predatory stimuli tend to inhibit brain cell proliferation. Often, laboratory predatory stimuli also elevate plasma glucocorticoids, which can then reduce brain cell proliferation. However, it is unknown how natural predators affect cell proliferation or whether glucocorticoids mediate the neurogenic response to natural predators. We examined brain cell proliferation in six populations of the electric fish, Brachyhypopomus occidentalis, exposed to three forms of predator stimuli: (i) natural variation in the density of predatory catfish; (ii) tail injury, presumably from predation attempts; and (iii) the acute stress of capture. Populations with higher predation pressure had lower density of proliferating (PCNA+) cells, and fish with injured tails had lower proliferating cell density than those with intact tails. However, plasma cortisol did not vary at the population level according to predation pressure or at the individual level according to tail injury. Capture stress significantly increased cortisol, but only marginally decreased cell proliferation. Thus, it appears that the presence of natural predators inhibits brain cell proliferation, but not via mechanisms that depend on changes in basal cortisol levels. This study is the first demonstration of predator-induced alteration of brain cell proliferation in a free-living vertebrate.

  6. Typical and atypical stem cells in the brain, vitamin C effect and neuropathology.

    PubMed

    Nualart, Francisco; Salazar, Katterine; Oyarce, Karina; Cisternas, Pedro; Jara, Nery; Silva-Álvarez, Carmen; Pastor, Patricia; Martínez, Fernando; García, Andrea; García-Robles, María de los Ángeles; Tapia, Juan Carlos

    2012-01-01

    Stem cells are considered a valuable cellular resource for tissue replacement therapies in most brain disorders. Stem cells have the ability to self-replicate and differentiate into numerous cell types, including neurons, oligodendrocytes and astrocytes. As a result, stem cells have been considered the "holy grail" of modern medical neuroscience. Despite their tremendous therapeutic potential, little is known about the mechanisms that regulate their differentiation. In this review, we analyze stem cells in embryonic and adult brains, and illustrate the differentiation pathways that give origin to most brain cells. We also evaluate the emergent role of the well known anti-oxidant, vitamin C, in stem cell differentiation. We believe that a complete understanding of all molecular players, including vitamin C, in stem cell differentiation will positively impact on the use of stem cell transplantation for neurodegenerative diseases.

  7. From germ cell preservation to regenerative medicine: an exciting research career in biotechnology.

    PubMed

    Wilmut, Ian

    2014-02-01

    Collection, manipulation, assessment, and storage of mammalian gametes, embryos, and stem cells are providing important opportunities in agriculture, research, and medicine. Semen and embryo freezing in livestock are used in breeding schemes, especially in cattle and for international trade, with no risk of spreading disease. In human medicine, they are used in treatment of infertility. Usually, knowledge gained in one species is applicable in the others. In one exception, some ruminant embryos cultured according to protocols used in human in vitro fertilization become unusually large offspring. This is due to disturbances in expression of imprinted genes. The nuclear transfer procedure developed at the Roslin Institute is being used to make genetic modifications in livestock to either direct production of biomedical proteins, create animal models of human disease, or enhance animal health and productivity. Human pluripotent cells are being used in Edinburgh to identify drugs to treat degenerative diseases.

  8. Chibby functions to preserve normal ciliary morphology through the regulation of intraflagellar transport in airway ciliated cells.

    PubMed

    Siller, Saul S; Burke, Michael C; Li, Feng-Qian; Takemaru, Ken-Ichi

    2015-01-01

    Airway cilia provide the coordinated motive force for mucociliary transport, which prevents the accumulation of mucus, debris, pollutants, and bacteria in our respiratory tracts. As airway cilia are constantly exposed to the environment and, hence, are an integral component of the pathogenesis of several congenital and chronic pulmonary disorders, it is necessary to understand the molecular mechanisms that control ciliated cell differentiation and ciliogenesis. We have previously reported that loss of the basal body protein Chibby (Cby) results in chronic upper airway infection in mice due to a significant reduction in the number of airway cilia. In the present work, we demonstrate that Cby is required for normal ciliary structure and proper distribution of proteins involved in the bidirectional intraflagellar transport (IFT) system, which consists of 2 distinct sub-complexes, IFT-A and IFT-B, and is essential for ciliary biogenesis and maintenance. In fully differentiated ciliated cells, abnormal paddle-like cilia with dilated ciliary tips are observed in Cby-/- airways and primary cultures of mouse tracheal epithelial cells (MTECs). In addition, IFT88, an IFT-B sub-complex protein, robustly accumulates within the dilated tips of both multicilia in Cby-/- MTECs and primary cilia in Cby-/- mouse embryonic fibroblasts (MEFs). Furthermore, we show that only IFT-B components, including IFT20 and IFT57, but not IFT-A and Bardet-Biedl syndrome (BBS) proteins, amass with IFT88 in these distended tips in Cby-/- ciliated cells. Taken together, our findings suggest that Cby plays a role in the proper distribution of IFT particles to preserve normal ciliary morphology in airway ciliated cells.

  9. Single cell PCR amplification of diatoms using fresh and preserved samples

    PubMed Central

    Hamilton, Paul B.; Lefebvre, Keely E.; Bull, Roger D.

    2015-01-01

    Single cell Chelex® DNA extraction and nested PCR amplification were used to examine partial gene sequences from natural diatom populations for taxonomic and phylogenetic studies at and above the level of species. DNA was extracted from cells that were either fresh collected or stored in RNAlater. Extractions from Lugol's fixation were also attempted with limited success. Three partial gene sequences (rbcL, 18S, and psbA) were recovered using existing and new primers with a nested or double nested PCR approach with amplification and success rates between 70 and 96%. An rbcL consensus tree grouped morphologically similar specimens and was consistent across the two primary sample treatments: fresh and RNAlater. This tool will greatly enhance the number of microscopic diatom taxa (and potentially other microbes) available for barcoding and phylogenetic studies. The near-term increase in sequence data for diatoms generated via routine single cell extractions and PCR will act as a multiproxy validation of longer-term next generation genomics. PMID:26528252

  10. MicroRNA-21 preserves the fibrotic mechanical memory of mesenchymal stem cells

    NASA Astrophysics Data System (ADS)

    Li, Chen Xi; Talele, Nilesh P.; Boo, Stellar; Koehler, Anne; Knee-Walden, Ericka; Balestrini, Jenna L.; Speight, Pam; Kapus, Andras; Hinz, Boris

    2016-10-01

    Expansion on stiff culture substrates activates pro-fibrotic cell programs that are retained by mechanical memory. Here, we show that priming on physiologically soft silicone substrates suppresses fibrogenesis and desensitizes mesenchymal stem cells (MSCs) against subsequent mechanical activation in vitro and in vivo, and identify the microRNA miR-21 as a long-term memory keeper of the fibrogenic program in MSCs. During stiff priming, miR-21 levels were gradually increased by continued regulation through the acutely mechanosensitive myocardin-related transcription factor-A (MRTF-A/MLK-1) and remained high over 2 weeks after removal of the mechanical stimulus. Knocking down miR-21 once by the end of the stiff-priming period was sufficient to erase the mechanical memory and sensitize MSCs to subsequent exposure to soft substrates. Soft priming and erasing mechanical memory following cell culture expansion protects MSCs from fibrogenesis in the host wound environment and increases the chances for success of MSC therapy in tissue-repair applications.

  11. Nanoparticle accumulation and transcytosis in brain endothelial cell layers

    NASA Astrophysics Data System (ADS)

    Ye, Dong; Raghnaill, Michelle Nic; Bramini, Mattia; Mahon, Eugene; Åberg, Christoffer; Salvati, Anna; Dawson, Kenneth A.

    2013-10-01

    The blood-brain barrier (BBB) is a selective barrier, which controls and limits access to the central nervous system (CNS). The selectivity of the BBB relies on specialized characteristics of the endothelial cells that line the microvasculature, including the expression of intercellular tight junctions, which limit paracellular permeability. Several reports suggest that nanoparticles have a unique capacity to cross the BBB. However, direct evidence of nanoparticle transcytosis is difficult to obtain, and we found that typical transport studies present several limitations when applied to nanoparticles. In order to investigate the capacity of nanoparticles to access and transport across the BBB, several different nanomaterials, including silica, titania and albumin- or transferrin-conjugated gold nanoparticles of different sizes, were exposed to a human in vitro BBB model of endothelial hCMEC/D3 cells. Extensive transmission electron microscopy imaging was applied in order to describe nanoparticle endocytosis and typical intracellular localisation, as well as to look for evidence of eventual transcytosis. Our results show that all of the nanoparticles were internalised, to different extents, by the BBB model and accumulated along the endo-lysosomal pathway. Rare events suggestive of nanoparticle transcytosis were also observed for several of the tested materials.The blood-brain barrier (BBB) is a selective barrier, which controls and limits access to the central nervous system (CNS). The selectivity of the BBB relies on specialized characteristics of the endothelial cells that line the microvasculature, including the expression of intercellular tight junctions, which limit paracellular permeability. Several reports suggest that nanoparticles have a unique capacity to cross the BBB. However, direct evidence of nanoparticle transcytosis is difficult to obtain, and we found that typical transport studies present several limitations when applied to nanoparticles. In

  12. Neural stem cells harvested from live brains by antibody-conjugated magnetic nanoparticles.

    PubMed

    Lui, C N P; Tsui, Y P; Ho, A S L; Shum, D K Y; Chan, Y S; Wu, C T; Li, H W; Tsang, S C Edman; Yung, K K L

    2013-11-18

    It stems from the magnetism: The extraction of stem/progenitor cells from the brain of live animals is possible using antibodies conjugated to magnetic nanoparticles (Ab-MNPs). The Ab-MNPs are introduced to a rat's brain with a superfine micro-syringe. The stem cells attach to the Ab-MNPs and are magnetically isolated and removed. They can develop into neurospheres and differentiate into different types of cells outside the subject body. The rat remains alive and healthy.

  13. Beta4 tubulin identifies a primitive cell source for oligodendrocytes in the mammalian brain.

    PubMed

    Wu, Chuanshen; Chang, Ansi; Smith, Maria C; Won, Roy; Yin, Xinghua; Staugaitis, Susan M; Agamanolis, Dimitri; Kidd, Grahame J; Miller, Robert H; Trapp, Bruce D

    2009-06-17

    We have identified a novel population of cells in the subventricular zone (SVZ) of the mammalian brain that expresses beta4 tubulin (betaT4) and has properties of primitive neuroectodermal cells. betaT4 cells are scattered throughout the SVZ of the lateral ventricles in adult human brain and are significantly increased in the SVZs bordering demyelinated white matter in multiple sclerosis brains. In human fetal brain, betaT4 cell densities peak during the latter stages of gliogenesis, which occurs in the SVZ of the lateral ventricles. betaT4 cells represent <2% of the cells present in neurospheres generated from postnatal rat brain but >95% of cells in neurospheres treated with the anti-mitotic agent Ara C. betaT4 cells produce oligodendrocytes, neurons, and astrocytes in vitro. We compared the myelinating potential of betaT4-positive cells with A2B5-positive oligodendrocyte progenitor cells after transplantation (25,000 cells) into postnatal day 3 (P3) myelin-deficient rat brains. At P20, the progeny of betaT4 cells myelinated up to 4 mm of the external capsule, which significantly exceeded that of transplanted A2B5-positive progenitor cells. Such extensive and rapid mature CNS cell generation by a relatively small number of transplanted cells provides in vivo support for the therapeutic potential of betaT4 cells. We propose that betaT4 cells are an endogenous cell source that can be recruited to promote neural repair in the adult telencephalon.

  14. Immortalized endothelial cell lines for in vitro blood-brain barrier models: A systematic review.

    PubMed

    Rahman, Nurul Adhwa; Rasil, Alifah Nur'ain Haji Mat; Meyding-Lamade, Uta; Craemer, Eva Maria; Diah, Suwarni; Tuah, Ani Afiqah; Muharram, Siti Hanna

    2016-07-01

    Endothelial cells play the most important role in construction of the blood-brain barrier. Many studies have opted to use commercially available, easily transfected or immortalized endothelial cell lines as in vitro blood-brain barrier models. Numerous endothelial cell lines are available, but we do not currently have strong evidence for which cell lines are optimal for establishment of such models. This review aimed to investigate the application of immortalized endothelial cell lines as in vitro blood-brain barrier models. The databases used for this review were PubMed, OVID MEDLINE, ProQuest, ScienceDirect, and SpringerLink. A narrative systematic review was conducted and identified 155 studies. As a result, 36 immortalized endothelial cell lines of human, mouse, rat, porcine and bovine origins were found for the establishment of in vitro blood-brain barrier and brain endothelium models. This review provides a summary of immortalized endothelial cell lines as a guideline for future studies and improvements in the establishment of in vitro blood-brain barrier models. It is important to establish a good and reproducible model that has the potential for multiple applications, in particular a model of such a complex compartment such as the blood-brain barrier.

  15. Systemic Autoimmunity in TAM Triple Knockout Mice Causes Inflammatory Brain Damage and Cell Death

    PubMed Central

    Li, Qiutang; Lu, Qingjun; Lu, Huayi; Tian, Shifu; Lu, Qingxian

    2013-01-01

    The Tyro3, Axl and Mertk (TAM) triply knockout (TKO) mice exhibit systemic autoimmune diseases, with characteristics of increased proinflammatory cytokine production, autoantibody deposition and autoreactive lymphocyte infiltration into a variety of tissues. Here we show that TKO mice produce high level of serum TNF-α and specific autoantibodies deposited onto brain blood vessels. The brain-blood barrier (BBB) in mutant brains exhibited increased permeability for Evans blue and fluorescent-dextran, suggesting a breakdown of the BBB in the mutant brains. Impaired BBB integrity facilitated autoreactive T cells infiltrating into all regions of the mutant brains. Brain autoimmune disorder caused accumulation of the ubiquitin-reactive aggregates in the mutant hippocampus, and early formation of autofluorescent lipofuscins in the neurons throughout the entire brains. Chronic neuroinflammation caused damage of the hippocampal mossy fibers and neuronal apoptotic death. This study shows that chronic systemic inflammation and autoimmune disorders in the TKO mice cause neuronal damage and death. PMID:23840307

  16. Isolation and Flow Cytometric Analysis of Immune Cells from the Ischemic Mouse Brain

    PubMed Central

    Boltze, Johannes; Wagner, Daniel-Christoph; Weise, Gesa

    2016-01-01

    Ischemic stroke initiates a robust inflammatory response that starts in the intravascular compartment and involves rapid activation of brain resident cells. A key mechanism of this inflammatory response is the migration of circulating immune cells to the ischemic brain facilitated by chemokine release and increased endothelial adhesion molecule expression. Brain-invading leukocytes are well-known contributing to early-stage secondary ischemic injury, but their significance for the termination of inflammation and later brain repair has only recently been noticed. Here, a simple protocol for the efficient isolation of immune cells from the ischemic mouse brain is provided. After transcardial perfusion, brain hemispheres are dissected and mechanically dissociated. Enzymatic digestion with Liberase is followed by density gradient (such as Percoll) centrifugation to remove myelin and cell debris. One major advantage of this protocol is the single-layer density gradient procedure which does not require time-consuming preparation of gradients and can be reliably performed. The approach yields highly reproducible cell counts per brain hemisphere and allows for measuring several flow cytometry panels in one biological replicate. Phenotypic characterization and quantification of brain-invading leukocytes after experimental stroke may contribute to a better understanding of their multifaceted roles in ischemic injury and repair. PMID:26967380

  17. The glia doctrine: addressing the role of glial cells in healthy brain ageing.

    PubMed

    Nagelhus, Erlend A; Amiry-Moghaddam, Mahmood; Bergersen, Linda H; Bjaalie, Jan G; Eriksson, Jens; Gundersen, Vidar; Leergaard, Trygve B; Morth, J Preben; Storm-Mathisen, Jon; Torp, Reidun; Walhovd, Kristine B; Tønjum, Tone

    2013-10-01

    Glial cells in their plurality pervade the human brain and impact on brain structure and function. A principal component of the emerging glial doctrine is the hypothesis that astrocytes, the most abundant type of glial cells, trigger major molecular processes leading to brain ageing. Astrocyte biology has been examined using molecular, biochemical and structural methods, as well as 3D brain imaging in live animals and humans. Exosomes are extracelluar membrane vesicles that facilitate communication between glia, and have significant potential for biomarker discovery and drug delivery. Polymorphisms in DNA repair genes may indirectly influence the structure and function of membrane proteins expressed in glial cells and predispose specific cell subgroups to degeneration. Physical exercise may reduce or retard age-related brain deterioration by a mechanism involving neuro-glial processes. It is most likely that additional information about the distribution, structure and function of glial cells will yield novel insight into human brain ageing. Systematic studies of glia and their functions are expected to eventually lead to earlier detection of ageing-related brain dysfunction and to interventions that could delay, reduce or prevent brain dysfunction.

  18. Endogenous mesenchymal stromal cells in bone marrow are required to preserve muscle function in mdx mice.

    PubMed

    Fujita, Ryo; Tamai, Katsuto; Aikawa, Eriko; Nimura, Keisuke; Ishino, Saki; Kikuchi, Yasushi; Kaneda, Yasufumi

    2015-03-01

    The physiological role of "endogenous" bone marrow (BM) mesenchymal stromal cells (MSCs) in tissue regeneration is poorly understood. Here, we show the significant contribution of unique endogenous BM-MSC populations to muscle regeneration in Duchenne muscular dystrophy (DMD) mice (mdx). Transplantation of BM cells (BMCs) from 10-week-old mdx into 3-4-week-old mdx mice increased inflammation and fibrosis and reduced muscle function compared with mdx mice that received BMCs from 10-week-old wild-type mice, suggesting that the alteration of BMC populations in mdx mice affects the progression of muscle pathology. Two distinct MSC populations in BM, that is, hematopoietic lineage (Lin)(-) /ckit(-) /CD106(+) /CD44(+) and Lin(-) /ckit(-) /CD106(+) /CD44(-) cells, were significantly reduced in 10-week-old mdx mice in disease progression. The results of a whole-transcriptome analysis indicated that these two MSC populations have distinct gene expression profiles, indicating that the Lin(-) /ckit(-) /CD106(+) /CD44(+) and Lin(-) /ckit(-) /CD106(+) /CD44(-) MSC populations are proliferative- and dormant-state populations in BM, respectively. BM-derived Lin(-) /CD106(+) /CD44(+) MSCs abundantly migrated to damaged muscles and highly expressed tumor necrosis factor-alpha-stimulated gene/protein-6 (TSG-6), an anti-inflammatory protein, in damaged muscles. We also demonstrated that TSG-6 stimulated myoblast proliferation. The injection of Lin(-) /ckit(-) /CD106(+) /CD44(+) MSCs into the muscle of mdx mice successfully ameliorated muscle dysfunction by decreasing inflammation and enhancing muscle regeneration through TSG-6-mediated activities. Thus, we propose a novel function of the unique endogenous BM-MSC population, which countered muscle pathology progression in a DMD model.

  19. Bone marrow stromal cell transplantation preserves gammaaminobutyric acid receptor function in the injured spinal cord.

    PubMed

    Yano, Shunsuke; Kuroda, Satoshi; Shichinohe, Hideo; Seki, Toshitaka; Ohnishi, Takako; Tamagami, Hiroshi; Hida, Kazutoshi; Iwasaki, Yoshinobu

    2006-11-01

    A surprising shortage of information surrounds the mechanisms by which bone marrow stromal cells (BMSC) restore lost neurologic functions when transplanted into the damaged central nervous system. In the present study, we sought to elucidate whether BMSCs express the neuron-specific gamma-aminobutyric acid (GABA) receptor when transplanted into injured spinal cord. To examine this, we harvested and cultured rat femoral BMSCs. We then subjected Sprague-Dawley rats to thoracic spinal cord injury (SCI) with a pneumatic impact device. Fluorescence-labeled BMSCs (n = 7) were transplanted stereotactically or the vehicle in which these cells were cultured (n = 4) was introduced stereotactically into the rostral site of SCI at 7 days after injury. We evaluated GABA receptor function by measuring the binding potential for 125I-iomazenil (125I-IMZ) through in vitro autoradiography at 4 weeks after BMSC transplantation and simultaneously examined the fate of the transplanted BMSCs by immunocytochemistry. We found that the transplanted BMSC migrated toward the core of the injury and were densely distributed in the marginal region at 4 weeks after transplantation. BMSC transplantation significantly increased the binding potential for 125I-IMZ (p = 0.0376) and increased the number of GABA receptor-positive cells (p = 0.0077) in the marginal region of the injury site. Some of the transplanted BMSCs were positive for microtubule-associated protein-2 and the alpha1 subunit of GABA(A) receptor in the region of injury. These findings suggest that BMSCs have the potential to support the survival of neurons in the marginal region of SCI and can partly differentiate into neurons, regenerating spinal cord tissue at the site of injury.

  20. Schwann Cell-Mediated Preservation of Vision in Retinal Degenerative Diseases via the Reduction of Oxidative Stress: A Possible Mechanism

    PubMed Central

    MAHMOUDZADEH, Raziyeh; HEIDARI-KESHEL, Saeed; LASHAY, Alireza

    2016-01-01

    After injury to the central nervous system (CNS), regeneration is often inadequate, except in the case of remyelination. This remyelination capacity of the CNS is a good example of a stem/precursor cell-mediated renewal process. Schwann cells have been found to act as remyelinating agents in the peripheral nervous system (PNS), but several studies have highlighted their potential role in remyelination in the CNS too. Schwann cells are able to protect and support retinal cells by secreting growth factors such as brain-derived neurotrophic factor, glial cell line-derived neurotrophic factor, and basic fibroblast growth factor. Retinal degenerative diseases can be highly debilitating, and they are a major concern in countries with an ageing populations. One of the leading causes of permanent loss of vision in the West is a retinal degenerative disease known as age-related macular degeneration (AMD). In the United States, nearly 1.75 million people over the age of 40 have advanced AMD, and it is estimated that this number will increase to approximately 3 million people by 2020. One of the most common pathways involved in the initiation and development of retinal diseases is the oxidative stress pathway. In patients with diabetes, Schwann cells have been shown to be able to secrete large amounts of antioxidant enzymes that protect the PNS from the oxidative stress that results from fluctuations in blood glucose levels. This antioxidant ability may be involved in the mechanism by which Schwann cells are able to promote reconstruction in the CNS, especially in individuals with retinal injuries and degenerative diseases. PMID:28293647

  1. DAPK2 regulates oxidative stress in cancer cells by preserving mitochondrial function

    PubMed Central

    Schlegel, C R; Georgiou, M L; Misterek, M B; Stöcker, S; Chater, E R; Munro, C E; Pardo, O E; Seckl, M J; Costa-Pereira, A P

    2015-01-01

    Death-associated protein kinase (DAPK) 2 is a serine/threonine kinase that belongs to the DAPK family. Although it shows significant structural differences from DAPK1, the founding member of this protein family, DAPK2 is also thought to be a putative tumour suppressor. Like DAPK1, it has been implicated in programmed cell death, the regulation of autophagy and diverse developmental processes. In contrast to DAPK1, however, few mechanistic studies have been carried out on DAPK2 and the majority of these have made use of tagged DAPK2, which almost invariably leads to overexpression of the protein. As a consequence, physiological roles of this kinase are still poorly understood. Using two genetically distinct cancer cell lines as models, we have identified a new role for DAPK2 in the regulation of mitochondrial integrity. RNA interference-mediated depletion of DAPK2 leads to fundamental metabolic changes, including significantly decreased rate of oxidative phosphorylation in combination with overall destabilised mitochondrial membrane potential. This phenotype is further corroborated by an increase in the production of mitochondrial superoxide anions and increased oxidative stress. This then leads to the activation of classical stress-activated kinases such as ERK, JNK and p38, which is observed on DAPK2 genetic ablation. Interestingly, the generation of oxidative stress is further enhanced on overexpression of a kinase-dead DAPK2 mutant indicating that it is the kinase domain of DAPK2 that is important to maintain mitochondrial integrity and, by inference, for cellular metabolism. PMID:25741596

  2. Hepatic glucose sensing is required to preserve β cell glucose competence.

    PubMed

    Seyer, Pascal; Vallois, David; Poitry-Yamate, Carole; Schütz, Frédéric; Metref, Salima; Tarussio, David; Maechler, Pierre; Staels, Bart; Lanz, Bernard; Grueter, Rolf; Decaris, Julie; Turner, Scott; da Costa, Anabela; Preitner, Frédéric; Minehira, Kaori; Foretz, Marc; Thorens, Bernard

    2013-04-01

    Liver glucose metabolism plays a central role in glucose homeostasis and may also regulate feeding and energy expenditure. Here we assessed the impact of glucose transporter 2 (Glut2) gene inactivation in adult mouse liver (LG2KO mice). Loss of Glut2 suppressed hepatic glucose uptake but not glucose output. In the fasted state, expression of carbohydrate-responsive element-binding protein (ChREBP) and its glycolytic and lipogenic target genes was abnormally elevated. Feeding, energy expenditure, and insulin sensitivity were identical in LG2KO and control mice. Glucose tolerance was initially normal after Glut2 inactivation, but LG2KO mice exhibited progressive impairment of glucose-stimulated insulin secretion even though β cell mass and insulin content remained normal. Liver transcript profiling revealed a coordinated downregulation of cholesterol biosynthesis genes in LG2KO mice that was associated with reduced hepatic cholesterol in fasted mice and reduced bile acids (BAs) in feces, with a similar trend in plasma. We showed that chronic BAs or farnesoid X receptor (FXR) agonist treatment of primary islets increases glucose-stimulated insulin secretion, an effect not seen in islets from Fxr(-/-) mice. Collectively, our data show that glucose sensing by the liver controls β cell glucose competence and suggest BAs as a potential mechanistic link.

  3. Behavior of neural stem cells in the Alzheimer brain

    PubMed Central

    Waldau, B.; Shetty, A. K.

    2013-01-01

    Alzheimer’s disease (AD) is characterized by the deposition of β-amyloid peptides (Aβ) and a progressive loss of neurons leading to dementia. Because hippocampal neurogenesis is linked to functions such as learning, memory and mood, there has been great interest in examining the effects of AD on hippocampal neurogenesis. This article reviews the pertinent studies and tries to unite them in one possible disease model. Early in the disease, oligomeric Aβ may transiently promote the generation of immature neurons from neural stem cells (NSCs). However, reduced concentrations of multiple neurotrophic factors and higher levels of fibroblast growth factor-2 seem to induce a developmental arrest of newly generated neurons. Furthermore, fibrillary Aβ and down-regulation of oligodendrocyte-lineage transcription factor-2 (OLIG2) may cause the death of these nonfunctional neurons. Therefore, altering the brain microenvironment for fostering apt maturation of graft-derived neurons may be critical for improving the efficacy of NSC transplantation therapy for AD. PMID:18500448

  4. Brain vascular pericytes following ischemia have multipotential stem cell activity to differentiate into neural and vascular lineage cells.

    PubMed

    Nakagomi, Takayuki; Kubo, Shuji; Nakano-Doi, Akiko; Sakuma, Rika; Lu, Shan; Narita, Aya; Kawahara, Maiko; Taguchi, Akihiko; Matsuyama, Tomohiro

    2015-06-01

    Brain vascular pericytes (PCs) are a key component of the blood-brain barrier (BBB)/neurovascular unit, along with neural and endothelial cells. Besides their crucial role in maintaining the BBB, increasing evidence shows that PCs have multipotential stem cell activity. However, their multipotency has not been considered in the pathological brain, such as after an ischemic stroke. Here, we examined whether brain vascular PCs following ischemia (iPCs) have multipotential stem cell activity and differentiate into neural and vascular lineage cells to reconstruct the BBB/neurovascular unit. Using PCs extracted from ischemic regions (iPCs) from mouse brains and human brain PCs cultured under oxygen/glucose deprivation, we show that PCs developed stemness presumably through reprogramming. The iPCs revealed a complex phenotype of angioblasts, in addition to their original mesenchymal properties, and multidifferentiated into cells from both a neural and vascular lineage. These data indicate that under ischemic/hypoxic conditions, PCs can acquire multipotential stem cell activity and can differentiate into major components of the BBB/neurovascular unit. Thus, these findings support the novel concept that iPCs can contribute to both neurogenesis and vasculogenesis at the site of brain injuries.

  5. Digitizing Preservation.

    ERIC Educational Resources Information Center

    Conway, Paul

    1994-01-01

    Discussion of digital imaging technology focuses on its potential use for preservation of library materials. Topics addressed include converting microfilm to digital; the high cost of conversion from paper or microfilm; quality; indexing; database management issues; incompatibility among imaging systems; longevity; cooperative pilot projects; and…

  6. Preservation Matters

    ERIC Educational Resources Information Center

    Noriega, Chon A.

    2005-01-01

    One must undertake multi-institutional efforts that include universities, archives, museums, libraries and community-based arts organizations and the artists to preserve Latino art history. Arts infrastructure can be strengthened by various Chicano Studies Research Center projects that are concerned with archive building and scholarship, and with…

  7. Neighborhood Preservation.

    ERIC Educational Resources Information Center

    Benin, Shirley

    1984-01-01

    Because of concern about the preservation of the historic character of Stamford (Connecticut), children in a pilot program at an elementary school learned about neighborhood history, sketched houses, researched houses which had been torn down and drew and constructed replicas of them, and learned about renovation and period interior design. (IS)

  8. Microglial cell dysregulation in brain aging and neurodegeneration.

    PubMed

    von Bernhardi, Rommy; Eugenín-von Bernhardi, Laura; Eugenín, Jaime

    2015-01-01

    Aging is the main risk factor for neurodegenerative diseases. In aging, microglia undergoes phenotypic changes compatible with their activation. Glial activation can lead to neuroinflammation, which is increasingly accepted as part of the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD). We hypothesize that in aging, aberrant microglia activation leads to a deleterious environment and neurodegeneration. In aged mice, microglia exhibit an increased expression of cytokines and an exacerbated inflammatory response to pathological changes. Whereas LPS increases nitric oxide (NO) secretion in microglia from young mice, induction of reactive oxygen species (ROS) predominates in older mice. Furthermore, there is accumulation of DNA oxidative damage in mitochondria of microglia during aging, and also an increased intracellular ROS production. Increased ROS activates the redox-sensitive nuclear factor kappa B, which promotes more neuroinflammation, and can be translated in functional deficits, such as cognitive impairment. Mitochondria-derived ROS and cathepsin B, are also necessary for the microglial cell production of interleukin-1β, a key inflammatory cytokine. Interestingly, whereas the regulatory cytokine TGFβ1 is also increased in the aged brain, neuroinflammation persists. Assessing this apparent contradiction, we have reported that TGFβ1 induction and activation of Smad3 signaling after inflammatory stimulation are reduced in adult mice. Other protective functions, such as phagocytosis, although observed in aged animals, become not inducible by inflammatory stimuli and TGFβ1. Here, we discuss data suggesting that mitochondrial and endolysosomal dysfunction could at least partially mediate age-associated microglial cell changes, and, together with the impairment of the TGFβ1-Smad3 pathway, could result in the reduction of protective activation and the facilitation of cytotoxic activation of microglia, resulting in the promotion of

  9. Microglial cell dysregulation in brain aging and neurodegeneration

    PubMed Central

    von Bernhardi, Rommy; Eugenín-von Bernhardi, Laura; Eugenín, Jaime

    2015-01-01

    Aging is the main risk factor for neurodegenerative diseases. In aging, microglia undergoes phenotypic changes compatible with their activation. Glial activation can lead to neuroinflammation, which is increasingly accepted as part of the pathogenesis of neurodegenerative diseases, including Alzheimer’s disease (AD). We hypothesize that in aging, aberrant microglia activation leads to a deleterious environment and neurodegeneration. In aged mice, microglia exhibit an increased expression of cytokines and an exacerbated inflammatory response to pathological changes. Whereas LPS increases nitric oxide (NO) secretion in microglia from young mice, induction of reactive oxygen species (ROS) predominates in older mice. Furthermore, there is accumulation of DNA oxidative damage in mitochondria of microglia during aging, and also an increased intracellular ROS production. Increased ROS activates the redox-sensitive nuclear factor kappa B, which promotes more neuroinflammation, and can be translated in functional deficits, such as cognitive impairment. Mitochondria-derived ROS and cathepsin B, are also necessary for the microglial cell production of interleukin-1β, a key inflammatory cytokine. Interestingly, whereas the regulatory cytokine TGFβ1 is also increased in the aged brain, neuroinflammation persists. Assessing this apparent contradiction, we have reported that TGFβ1 induction and activation of Smad3 signaling after inflammatory stimulation are reduced in adult mice. Other protective functions, such as phagocytosis, although observed in aged animals, become not inducible by inflammatory stimuli and TGFβ1. Here, we discuss data suggesting that mitochondrial and endolysosomal dysfunction could at least partially mediate age-associated microglial cell changes, and, together with the impairment of the TGFβ1-Smad3 pathway, could result in the reduction of protective activation and the facilitation of cytotoxic activation of microglia, resulting in the promotion of

  10. Hyaluronan preserves the proliferation and differentiation potentials of long-term cultured murine adipose-derived stromal cells

    SciTech Connect

    Chen, P.-Y.; Huang, Lynn L.H. . E-mail: lynn@mail.ncku.edu.tw; Hsieh, H.-J. . E-mail: hjhsieh@ntu.edu.tw

    2007-08-17

    For long-term culture, murine adipose-derived stromal cells (mADSCs) at latter passages demonstrated a marked decline in proliferative activity, exhibited senescent morphology and reduced differentiation potentials, particularly osteogenesis. To extend the lifespan of mADSCs, two culture conditions containing hyaluronan (HA) was compared in our study, one as a culture medium supplement (SHA), and the other where HA was pre-coated on culture surface (CHA). mADSCs cultivated with SHA exhibited a prolonged lifespan, reduced cellular senescence, and enhanced osteogenic potential compared to regular culture condition (control). Upon CHA treatment, mADSCs tended to form cell aggregates with gradual growth profiles, while their differentiation activities remained similar to SHA groups. After transferring mADSCs from CHA to control surface, they were shown to have an extended lifespan and an increase of osteogenic potential. Our results suggested that HA can be useful for preserving the proliferation and differentiation potentials of long-term cultured mADSCs.

  11. Impaired culture generated cytotoxicity with preservation of spontaneous natural killer-cell activity in cartilage-hair hypoplasia

    SciTech Connect

    Pierce, G.F.; Brovall, C.; Schacter, B.Z.; Polmar, S.H.

    1983-06-01

    Recent studies of cartilage-hair hypoplasia (CHH), a form of short-limbed dwarfism, have shown that all affected individuals have a cellular proliferation defect that results in a cellular immunodeficiency. However, only a minority of CHH individuals suffer from severe, life-threatening infections. For this reason, relevant immune defense mechanisms that may be responsible for maintaining intact host defenses in the majority of CHH individuals were studied. Spontaneous and allogeneic culture-induced (mixed lymphocyte response-MLR) specific and nonspecific (NK-like) cytotoxic mechanisms were analyzed and correlated with lymphocyte subpopulations present in CHH and normal individuals. Spontaneous natural-killer (NK) activity was present at or above normal levels, but culture-induced specific cytotoxicity and NK-like cytotoxicity as well as NK-like activity by T cell lines were significantly reduced in CHH individuals. The generation of radiation-resistant cytotoxicity, which normally occurs during allogeneic MLR, was markedly diminished in CHH, and was correlated with the decreased proliferation observed in CHH cultures. Preservation of spontaneous NK activity and loss of all forms of culture-induced cytotoxicity was associated with an increase in the proportion of lymphocytes bearing a thymic independent NK phenotype, and a significant decrease in thymic derived cytolytic T cell sub-populations in CHH individuals. Therefore, an intact cellular cytotoxic effector mechanism has been identified in CHH (i.e., NK activity).

  12. Proregenerative Microenvironment Triggered by Donor Mesenchymal Stem Cells Preserves Renal Function and Structure in Mice with Severe Diabetes Mellitus.

    PubMed

    Ezquer, Fernando; Giraud-Billoud, Maximiliano; Carpio, Daniel; Cabezas, Fabián; Conget, Paulette; Ezquer, Marcelo

    2015-01-01

    The aim of our work was to evaluate, in an animal model of severe diabetes mellitus, the effect of mesenchymal stem cells (MSCs) administration on diabetic nephropathy (DN) progression. After diabetes induction, one group of mice received the vehicle (DM) and other group received a single dose of MSCs (DM + MSCs). DM + MSCs mice showed a significant improvement in functional parameters of the kidney compared with untreated mice. While DM mice presented marked histopathological changes characteristics of advanced stages of DN (fibrosis, glomerulosclerosis, glomerular basement membrane thickening, capillary occlusion, decreased podocyte density, and effacement of foot processes), DM + MSCs mice showed only slight tubular dilatation. The renoprotection was not associated with an improvement in diabetic condition and very low number of donor cells was found in the kidney of DM + MSCs mice, suggesting that renoprotection could be mediated by paracrine effects. Indeed, DM + MSC mice presented increased renal proliferation index, decreased renal apoptotic index and the restoration of proregenerative factors, and anti-inflammatory cytokines levels. Moreover, macrophage infiltration and oxidative stress damage were also reduced in DM + MSCs mice. Our data demonstrate that MSC administration triggers a proregenerative microenvironment in DN kidney, which allows the preservation of the renal function even if diabetes was uncorrected.

  13. Proregenerative Microenvironment Triggered by Donor Mesenchymal Stem Cells Preserves Renal Function and Structure in Mice with Severe Diabetes Mellitus

    PubMed Central

    Ezquer, Fernando; Giraud-Billoud, Maximiliano; Carpio, Daniel; Cabezas, Fabián; Conget, Paulette; Ezquer, Marcelo

    2015-01-01

    The aim of our work was to evaluate, in an animal model of severe diabetes mellitus, the effect of mesenchymal stem cells (MSCs) administration on diabetic nephropathy (DN) progression. After diabetes induction, one group of mice received the vehicle (DM) and other group received a single dose of MSCs (DM + MSCs). DM + MSCs mice showed a significant improvement in functional parameters of the kidney compared with untreated mice. While DM mice presented marked histopathological changes characteristics of advanced stages of DN (fibrosis, glomerulosclerosis, glomerular basement membrane thickening, capillary occlusion, decreased podocyte density, and effacement of foot processes), DM + MSCs mice showed only slight tubular dilatation. The renoprotection was not associated with an improvement in diabetic condition and very low number of donor cells was found in the kidney of DM + MSCs mice, suggesting that renoprotection could be mediated by paracrine effects. Indeed, DM + MSC mice presented increased renal proliferation index, decreased renal apoptotic index and the restoration of proregenerative factors, and anti-inflammatory cytokines levels. Moreover, macrophage infiltration and oxidative stress damage were also reduced in DM + MSCs mice. Our data demonstrate that MSC administration triggers a proregenerative microenvironment in DN kidney, which allows the preservation of the renal function even if diabetes was uncorrected. PMID:26167475

  14. A unified and preserved Dirichlet boundary treatment for the cell-centered finite volume discrete Boltzmann method

    NASA Astrophysics Data System (ADS)

    Chen, Leitao; Schaefer, Laura A.

    2015-02-01

    A new boundary treatment is proposed for the finite volume discrete Boltzmann method (FVDBM) that can be used for accurate simulations of curved boundaries and complicated flow conditions. First, a brief review of different boundary treatments for the general Boltzmann simulations is made in order to primarily explain what type of boundary treatment will be developed in this paper for the cell-centered FVDBM. After that, the new boundary treatment along with the cell-centered FVDBM model is developed in detail. Next, the proposed boundary treatment is applied to a series of numerical tests with a detailed discussion of its qualitative and quantitative properties. From the results, it can be concluded that the new boundary treatment is at least first-order accurate for a variety of Dirichlet boundary conditions (BCs). It can handle both the velocity and density Dirichlet BCs in a unified way and further realize some BCs that the conventional lattice Boltzmann model fails to simulate. In addition, such a boundary treatment can incorporate different lattice models without changing its framework, and it can preserve the Dirichlet BCs up to machine accuracy in different situations.

  15. Brain tumor initiating cells adapt to restricted nutrition through preferential glucose uptake.

    PubMed

    Flavahan, William A; Wu, Qiulian; Hitomi, Masahiro; Rahim, Nasiha; Kim, Youngmi; Sloan, Andrew E; Weil, Robert J; Nakano, Ichiro; Sarkaria, Jann N; Stringer, Brett W; Day, Bryan W; Li, Meizhang; Lathia, Justin D; Rich, Jeremy N; Hjelmeland, Anita B

    2013-10-01

    Like all cancers, brain tumors require a continuous source of energy and molecular resources for new cell production. In normal brain, glucose is an essential neuronal fuel, but the blood-brain barrier limits its delivery. We now report that nutrient restriction contributes to tumor progression by enriching for brain tumor initiating cells (BTICs) owing to preferential BTIC survival and to adaptation of non-BTICs through acquisition of BTIC features. BTICs outcompete for glucose uptake by co-opting the high affinity neuronal glucose transporter, type 3 (Glut3, SLC2A3). BTICs preferentially express Glut3, and targeting Glut3 inhibits BTIC growth and tumorigenic potential. Glut3, but not Glut1, correlates with poor survival in brain tumors and other cancers; thus, tumor initiating cells may extract nutrients with high affinity. As altered metabolism represents a cancer hallmark, metabolic reprogramming may maintain the tumor hierarchy and portend poor prognosis.

  16. Tissue-engineered skin preserving the potential of epithelial cells to differentiate into hair after grafting.

    PubMed

    Larouche, Danielle; Cuffley, Kristine; Paquet, Claudie; Germain, Lucie

    2011-03-01

    The aim of this study was to evaluate whether tissue-engineered skin produced in vitro was able to sustain growth of hair follicles in vitro and after grafting. Different tissues were designed. Dissociated newborn mouse keratinocytes or newborn mouse hair buds (HBs) were added onto dermal constructs consisting of a tissue-engineered cell-derived matrix elaborated from either newborn mouse or adult human fibroblasts cultured with ascorbic acid. After 7-21 days of maturation at the air-liquid interface, no hair was noticed in vitro. Epidermal differentiation was observed in all tissue-engineered skin. However, human fibroblast-derived tissue-engineered dermis (hD) promoted a thicker epidermis than mouse fibroblast-derived tissue-engineered dermis (mD). In association with mD, HBs developed epithelial cyst-like inclusions presenting outer root sheath-like attributes. In contrast, epidermoid cyst-like inclusions lined by a stratified squamous epithelium were present in tissues composed of HBs and hD. After grafting, pilo-sebaceous units formed and hair grew in skin elaborated from HBs cultured 10-26 days submerged in culture medium in association with mD. However, the number of normal hair follicles decreased with longer culture time. This hair-forming capacity after grafting was not observed in tissues composed of hD overlaid with HBs. These results demonstrate that epithelial stem cells can be kept in vitro in a permissive tissue-engineered dermal environment without losing their potential to induce hair growth after grafting.

  17. Brain Ischemia Suppresses Immunity in the Periphery and Brain via Different Neurogenic Innervations.

    PubMed

    Liu, Qiang; Jin, Wei-Na; Liu, Yaou; Shi, Kaibin; Sun, Haoran; Zhang, Fang; Zhang, Chao; Gonzales, Rayna J; Sheth, Kevin N; La Cava, Antonio; Shi, Fu-Dong

    2017-03-21

    Brain ischemia inhibits immune function systemically, with resulting infectious complications. Whether in stroke different immune alterations occur in brain and periphery and whether analogous mechanisms operate in these compartments remains unclear. Here we show that in patients with ischemic stroke and in mice subjected to middle cerebral artery occlusion, natural killer (NK) cells display remarkably distinct temporal and transcriptome profiles in the brain as compared to the periphery. The activation of catecholaminergic and hypothalamic-pituitary-adrenal axis leads to splenic atrophy and contraction of NK cell numbers in the periphery through a modulated expression of SOCS3, whereas cholinergic innervation-mediated suppression of NK cell responses in the brain involves RUNX3. Importantly, pharmacological or genetic ablation of innervation preserved NK cell function and restrained post-stroke infection. Thus, brain ischemia compromises NK cell-mediated immune defenses through mechanisms that differ in the brain versus the periphery, and targeted inhibition of neurogenic innervation limits post-stroke infection.

  18. Carcinoma cells misuse the host tissue damage response to invade the brain.

    PubMed

    Chuang, Han-Ning; van Rossum, Denise; Sieger, Dirk; Siam, Laila; Klemm, Florian; Bleckmann, Annalen; Bayerlová, Michaela; Farhat, Katja; Scheffel, Jörg; Schulz, Matthias; Dehghani, Faramarz; Stadelmann, Christine; Hanisch, Uwe-Karsten; Binder, Claudia; Pukrop, Tobias

    2013-08-01

    The metastatic colonization of the brain by carcinoma cells is still barely understood, in particular when considering interactions with the host tissue. The colonization comes with a substantial destruction of the surrounding host tissue. This leads to activation of damage responses by resident innate immune cells to protect, repair, and organize the wound healing, but may distract from tumoricidal actions. We recently demonstrated that microglia, innate immune cells of the CNS, assist carcinoma cell invasion. Here we report that this is a fatal side effect of a physiological damage response of the brain tissue. In a brain slice coculture model, contact with both benign and malignant epithelial cells induced a response by microglia and astrocytes comparable to that seen at the interface of human cerebral metastases. While the glial damage response intended to protect the brain from intrusion of benign epithelial cells by inducing apoptosis, it proved ineffective against various malignant cell types. They did not undergo apoptosis and actually exploited the local tissue reaction to invade instead. Gene expression and functional analyses revealed that the C-X-C chemokine receptor type 4 (CXCR4) and WNT signaling were involved in this process. Furthermore, CXCR4-regulated microglia were recruited to sites of brain injury in a zebrafish model and CXCR4 was expressed in human stroke patients, suggesting a conserved role in damage responses to various types of brain injuries. Together, our findings point to a detrimental misuse of the glial damage response program by carcinoma cells resistant to glia-induced apoptosis.

  19. Brain Region and Cell Type Transcripts for Informative Diagnostics

    DTIC Science & Technology

    2010-09-01

    cerebral cortex (CTX) region that is a sheet of neural tissue that is outermost to the cerebrum of the mammalian brain and takes a key role in...the cerebrum of the mammalian brain and plays a role in memory, attention, perceptual awareness, thought, language, and consciousness. The MDRN

  20. Proteolytic Mechanisms of cell Death Following Traumatic Brain Injury

    DTIC Science & Technology

    2000-10-01

    produces early and sustained increases in calpain and caspae -3 activity that vary depending upon the brain regions studies. (2) TBI can produce...different brain regions. Changes may be differentially localized in subcellular fractions. (4) Increased magnitudes and duration of calpain and caspae -3

  1. 3D BrainCV: simultaneous visualization and analysis of cells and capillaries in a whole mouse brain with one-micron voxel resolution.

    PubMed

    Wu, Jingpeng; He, Yong; Yang, Zhongqin; Guo, Congdi; Luo, Qingming; Zhou, Wei; Chen, Shangbin; Li, Anan; Xiong, Benyi; Jiang, Tao; Gong, Hui

    2014-02-15

    Systematic cellular and vascular configurations are essential for understanding fundamental brain anatomy and metabolism. We demonstrated a 3D brainwide cellular and vascular (called 3D BrainCV) visualization and quantitative protocol for a whole mouse brain. We developed a modified Nissl staining method that quickly labeled the cells and blood vessels simultaneously in an entire mouse brain. Terabytes 3D datasets of the whole mouse brains, with unprecedented details of both individual cells and blood vessels, including capillaries, were simultaneously imaged at 1-μm voxel resolution using micro-optical sectioning tomography (MOST). For quantitative analysis, we proposed an automatic image-processing pipeline to perform brainwide vectorization and analysis of cells and blood vessels. Six representative brain regions from the cortex to the deep, including FrA, M1, PMBSF, V1, striatum, and amygdala, and six parameters, including cell number density, vascular length density, fractional vascular volume, distance from the cells to the nearest microvessel, microvascular length density, and fractional microvascular volume, had been quantitatively analyzed. The results showed that the proximity of cells to blood vessels was linearly correlated with vascular length density, rather than the cell number density. The 3D BrainCV made overall snapshots of the detailed picture of the whole brain architecture, which could be beneficial for the state comparison of the developing and diseased brain.

  2. Physiological Correlates of Intellectual Function in Children with Sickle Cell Disease: Hypoxaemia, Hyperaemia and Brain Infarction

    ERIC Educational Resources Information Center

    Hogan, Alexandra M.; Pit-ten Cate, Ineke M.; Vargha-Khadem, Faraneh; Prengler, Mara; Kirkham, Fenella J.

    2006-01-01

    Lowered intelligence relative to controls is evident by mid-childhood in children with sickle cell disease. There is consensus that brain infarct contributes to this deficit, but the subtle lowering of IQ in children with normal MRI scans might be accounted for by chronic systemic complications leading to insufficient oxygen delivery to the brain.…

  3. How Stem Cells Speak with Host Immune Cells in Inflammatory Brain Diseases

    PubMed Central

    Pluchino, Stefano; Cossetti, Chiara

    2014-01-01

    Advances in stem cell biology have raised great expectations that diseases and injuries of the central nervous system (CNS) may be ameliorated by the development of non-hematopoietic stem cell medicines. Yet, the application of adult stem cells as CNS therapeutics is challenging and the interpretation of some of the outcomes ambiguous. In fact, the initial idea that stem cell transplants work only via structural cell replacement has been challenged by the observation of consistent cellular signaling between the graft and the host. Cellular signaling is the foundation of coordinated actions and flexible responses, and arises via networks of exchanging and interacting molecules that transmit patterns of information between cells. Sustained stem cell graft-to-host communication leads to remarkable trophic effects on endogenous brain cells and beneficial modulatory actions on innate and adaptive immune responses in vivo, ultimately promoting the healing of the injured CNS. Among a number of adult stem cell types, mesenchymal stem cells (MSCs) and neural stem/precursor cells (NPCs) are being extensively investigated for their ability to signal to the immune system upon transplantation in experimental CNS diseases. Here, we focus on the main cellular signaling pathways that grafted MSCs and NPCs use to establish a therapeutically relevant cross talk with host immune cells, while examining the role of inflammation in regulating some of the bidirectionality of these communications. We propose that the identification of the players involved in stem cell signaling might contribute to the development of innovative, high clinical impact therapeutics for inflammatory CNS diseases. PMID:23633288

  4. Hemoglobin Function in Stored Blood. IX. A Modified Preservative with Optimal pH to Maintain Red Cell 2,3-DPG (Function) and ATP (Viability).

    DTIC Science & Technology

    differences at the 95 percent level were based on the paired ’t’ test. In addition, osmotic fragility and methylene blue uptake were determined to...optimal for maintaining hemoglobin function and red cell viability. In addition, methylene blue uptake seems to correlate fairly well with maintenance of ATP in the two lowest pH preservatives. (Author)

  5. Rat umbilical cord blood cells attenuate hypoxic–ischemic brain injury in neonatal rats

    PubMed Central

    Nakanishi, Keiko; Sato, Yoshiaki; Mizutani, Yuka; Ito, Miharu; Hirakawa, Akihiro; Higashi, Yujiro

    2017-01-01

    Increasing evidence has suggested that human umbilical cord blood cells (hUCBC) have a favorable effect on hypoxic–ischemic (HI) brain injury. However, the efficacy of using hUCBCs to treat this injury has been variable and the underlying mechanism remains elusive. Here, we investigated its effectiveness using stereological analysis in an allogeneic system to examine whether intraperitoneal injection of cells derived from UCBCs of green fluorescent protein (GFP)-transgenic rats could ameliorate brain injury in neonatal rats. Three weeks after the HI event, the estimated residual brain volume was larger and motor function improved more in the cell-injected rats than in the control (PBS-treated) rats. The GFP-positive cells were hardly detectable in the brain (0.0057% of injected cells) 9 days after injection. Although 60% of GFP-positive cells in the brain were Iba1-positive, none of these were positive for NeuroD or DCX. While the number of proliferating cells increased in the hippocampus, that of activated microglia/macrophages decreased and a proportion of M2 microglia/macrophages increased in the ipsilateral hemisphere of cell-injected rats. These results suggest that intraperitoneal injection of cells derived from UCBCs could ameliorate HI injury, possibly through an endogenous response and not by supplying differentiated neurons derived from the injected stem cells. PMID:28281676

  6. BDNF Expression in Larval and Adult Zebrafish Brain: Distribution and Cell Identification

    PubMed Central

    Cacialli, Pietro; Gueguen, Marie-Madeleine; Coumailleau, Pascal; D’Angelo, Livia; Kah, Olivier; Lucini, Carla; Pellegrini, Elisabeth

    2016-01-01

    Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, has emerged as an active mediator in many essential functions in the central nervous system of mammals. BDNF plays significant roles in neurogenesis, neuronal maturation and/or synaptic plasticity and is involved in cognitive functions such as learning and memory. Despite the vast literature present in mammals, studies devoted to BDNF in the brain of other animal models are scarse. Zebrafish is a teleost fish widely known for developmental genetic studies and is emerging as model for translational neuroscience research. In addition, its brain shows many sites of adult neurogenesis allowing higher regenerative properties after traumatic injuries. To add further knowledge on neurotrophic factors in vertebrate brain models, we decided to determine the distribution of bdnf mRNAs in the larval and adult zebrafish brain and to characterize the phenotype of cells expressing bdnf mRNAs by means of double staining studies. Our results showed that bdnf mRNAs were widely expressed in the brain of 7 days old larvae and throughout the whole brain of mature female and male zebrafish. In adults, bdnf mRNAs were mainly observed in the dorsal telencephalon, preoptic area, dorsal thalamus, posterior tuberculum, hypothalamus, synencephalon, optic tectum and medulla oblongata. By combining immunohistochemistry with in situ hybridization, we showed that bdnf mRNAs were never expressed by radial glial cells or proliferating cells. By contrast, bdnf transcripts were expressed in cells with neuronal phenotype in all brain regions investigated. Our results provide the first demonstration that the brain of zebrafish expresses bdnf mRNAs in neurons and open new fields of research on the role of the BDNF factor in brain mechanisms in normal and brain repairs situations. PMID:27336917

  7. Galectin-1 suppresses methamphetamine induced neuroinflammation in human brain microvascular endothelial cells: Neuroprotective role in maintaining blood brain barrier integrity.

    PubMed

    Parikh, Neil U; Aalinkeel, R; Reynolds, J L; Nair, B B; Sykes, D E; Mammen, M J; Schwartz, S A; Mahajan, S D

    2015-10-22

    Methamphetamine (Meth) abuse can lead to the breakdown of the blood-brain barrier (BBB) integrity leading to compromised CNS function. The role of Galectins in the angiogenesis process in tumor-associated endothelial cells (EC) is well established; however no data are available on the expression of Galectins in normal human brain microvascular endothelial cells and their potential role in maintaining BBB integrity. We evaluated the basal gene/protein expression levels of Galectin-1, -3 and -9 in normal primary human brain microvascular endothelial cells (BMVEC) that constitute the BBB and examined whether Meth altered Galectin expression in these cells, and if Galectin-1 treatment impacted the integrity of an in-vitro BBB. Our results showed that BMVEC expressed significantly higher levels of Galectin-1 as compared to Galectin-3 and -9. Meth treatment increased Galectin-1 expression in BMVEC. Meth induced decrease in TJ proteins ZO-1, Claudin-3 and adhesion molecule ICAM-1 was reversed by Galectin-1. Our data suggests that Galectin-1 is involved in BBB remodeling and can increase levels of TJ proteins ZO-1 and Claudin-3 and adhesion molecule ICAM-1 which helps maintain BBB tightness thus playing a neuroprotective role. Galectin-1 is thus an important regulator of immune balance from neurodegeneration to neuroprotection, which makes it an important therapeutic agent/target in the treatment of drug addiction and other neurological conditions.

  8. Brain transplantation of genetically engineered human neural stem cells globally corrects brain lesions in the mucopolysaccharidosis type VII mouse.

    PubMed

    Meng, Xing-Li; Shen, Jin-Song; Ohashi, Toya; Maeda, Hiroshi; Kim, Seung Up; Eto, Yoshikatsu

    2003-10-15

    In the present study, we investigated the feasibility of using human neural stem cells (NSCs) in the treatment of diffuse central nervous system (CNS) alterations in a murine model of mucopolysaccharidosis VII (MPS VII), a lysosomal storage disease caused by a genetic defect in the beta-glucuronidase gene. An immortalized NSC line derived from human fetal telencephalon was genetically engineered to overexpress beta-glucuronidase and transplanted into the cerebral ventricles of neonatal MPS VII mouse. Transplanted human NSCs were found to integrate and migrate in the host brain and to produce large amount of beta-glucuronidase. Brain contents of the substrates of beta-glucuronidase were reduced to nearly normal levels, and widespread clearing of lysosomal storage was observed in the MPS VII mouse brain at 25 days posttransplantation. The number of engrafted cells decreased markedly after the transplantation, and it appears that the major cause of the cell death was not the immune response of the host but apoptotic cell death of grafted human NSCs. Results showed that human NSCs would serve as a useful gene transfer vehicle for the treatment of diffuse CNS lesions in human lysosomal storage diseases and are potentially applicable in the treatment of patients suffering from neurological disorders.

  9. Human Brain Microvascular Endothelial Cells Derived from the BC1 iPS Cell Line Exhibit a Blood-Brain Barrier Phenotype.

    PubMed

    Katt, Moriah E; Xu, Zinnia S; Gerecht, Sharon; Searson, Peter C

    2016-01-01

    The endothelial cells that form capillaries in the brain are highly specialized, with tight junctions that minimize paracellular transport and an array of broad-spectrum efflux pumps that make drug delivery to the brain extremely challenging. One of the major limitations in blood-brain barrier research and the development of drugs to treat central nervous system diseases is the lack of appropriate cell lines. Recent reports indicate that the derivation of human brain microvascular endothelial cells (hBMECs) from human induced pluripotent stem cells (iPSCs) may provide a solution to this problem. Here we demonstrate the derivation of hBMECs extended to two new human iPSC lines: BC1 and GFP-labeled BC1. These hBMECs highly express adherens and tight junction proteins VE-cadherin, ZO-1, occludin, and claudin-5. The addition of retinoic acid upregulates VE-cadherin expression, and results in a significant increase in transendothelial electrical resistance to physiological values. The permeabilities of tacrine, rhodamine 123, and Lucifer yellow are similar to values obtained for MDCK cells. The efflux ratio for rhodamine 123 across hBMECs is in the range 2-4 indicating polarization of efflux transporters. Using the rod assay to assess cell organization in small vessels and capillaries, we show that hBMECs resist elongation with decreasing diameter but show progressive axial alignment. The derivation of hBMECs with a blood-brain barrier phenotype from the BC1 cell line highlights that the protocol is robust. The expression of GFP in hBMECs derived from the BC1-GFP cell line provides an important new resource for BBB research.

  10. Human Brain Microvascular Endothelial Cells Derived from the BC1 iPS Cell Line Exhibit a Blood-Brain Barrier Phenotype

    PubMed Central

    Gerecht, Sharon; Searson, Peter C.

    2016-01-01

    The endothelial cells that form capillaries in the brain are highly specialized, with tight junctions that minimize paracellular transport and an array of broad-spectrum efflux pumps that make drug delivery to the brain extremely challenging. One of the major limitations in blood-brain barrier research and the development of drugs to treat central nervous system diseases is the lack of appropriate cell lines. Recent reports indicate that the derivation of human brain microvascular endothelial cells (hBMECs) from human induced pluripotent stem cells (iPSCs) may provide a solution to this problem. Here we demonstrate the derivation of hBMECs extended to two new human iPSC lines: BC1 and GFP-labeled BC1. These hBMECs highly express adherens and tight junction proteins VE-cadherin, ZO-1, occludin, and claudin-5. The addition of retinoic acid upregulates VE-cadherin expression, and results in a significant increase in transendothelial electrical resistance to physiological values. The permeabilities of tacrine, rhodamine 123, and Lucifer yellow are similar to values obtained for MDCK cells. The efflux ratio for rhodamine 123 across hBMECs is in the range 2–4 indicating polarization of efflux transporters. Using the rod assay to assess cell organization in small vessels and capillaries, we show that hBMECs resist elongation with decreasing diameter but show progressive axial alignment. The derivation of hBMECs with a blood-brain barrier phenotype from the BC1 cell line highlights that the protocol is robust. The expression of GFP in hBMECs derived from the BC1-GFP cell line provides an important new resource for BBB research. PMID:27070801

  11. Neurothelin: an inducible cell surface glycoprotein of blood-brain barrier-specific endothelial cells and distinct neurons

    PubMed Central

    1990-01-01

    The blood-brain barrier is characterized by still poorly understood barrier and transport functions performed by specialized endothelial cells. Hybridoma technology has been used to identify a protein termed neurothelin that is specific for these endothelial cells. Neurothelin is defined by the species-specific mouse mAb 1W5 raised against lentil- lectin-binding proteins of neural tissue from embryonic chick. In the posthatch chick, neurothelin expression is found on endothelial cells within the brain but not on those of the systemic vascular system. Injection of the monoclonal antibody in vivo leads to labeling of brain capillaries, indicating that the corresponding antigen is expressed on the luminal surface of brain endothelial cells. Transplantation of embryonic mouse brain onto the chick chorioallantoic membrane results in rodent brain vascularization by the avian vascular system. Subsequently, normally mAb 1W5-negative endothelial cells, originating from blood vessels of the chick chorioallantoic membrane, are induced to express neurothelin when they are in contact with mouse neural tissue. In contrast to differentiated brain neurons that do not express neurothelin, neurons of the nonvascularized chick retina synthesize neurothelin. However, neurothelin is not found on retinal ganglion cell axons terminating on 1W5-negative brain cells. 1W5 immunoreactivity was also found in the pigment epithelium that forms the blood-eye barrier. Putting epithelial cells into culture results in concentration of neurothelin at cell-cell contact sites, leaving other cell surface areas devoid of antigen. Therefore, the distribution of neurothelin appears to be regulated by cell-cell interactions. In Western blot analysis, neurothelin was identified as a protein with a molecular mass of approximately 43 kD. The protein bears at least one intramolecular disulfide bridge and sulfated glucuronic acid as well as alpha-D- substituted mannose/glucose moieties. The exclusive

  12. No increase in brain cancer rates during period of expanding cell phone use

    Cancer.gov

    In a new examination of United States cancer incidence data, investigators at the National Cancer Institute (NCI) reported that incidence trends have remained roughly constant for glioma, the main type of brain cancer hypothesized to be related to cell ph

  13. Local apoptosis modulates early mammalian brain development through the elimination of morphogen-producing cells.

    PubMed

    Nonomura, Keiko; Yamaguchi, Yoshifumi; Hamachi, Misato; Koike, Masato; Uchiyama, Yasuo; Nakazato, Kenichi; Mochizuki, Atsushi; Sakaue-Sawano, Asako; Miyawaki, Atsushi; Yoshida, Hiroki; Kuida, Keisuke; Miura, Masayuki

    2013-12-23

    Apoptotic cells are observed in the early developing brain. Apoptosis deficiency is proposed to cause brain overgrowth, but here we show that brain malformations in apoptosis-deficient mutants are due to insufficient brain ventricle expansion as a result of uncompleted cranial neural tube closure. Apoptosis eliminates Fgf8-expressing cells in the anterior neural ridge (ANR), which acts as an organizing center of the forebrain by producing FGF8 morphogen. Deficiency of apoptosis leads to the accumulation of undead and nonproliferative cells in the ventral part of the ANR. The undead cells in apoptosis-deficient mutants express Fgf8 continuously, which perturbs gene expression in the ventral forebrain. Thus, apoptosis within a specific subdomain of the ANR is required for correct temporal elimination of an FGF8-producing region within a limited developmental time window, thereby ensuring proper forebrain development.

  14. Nose-to-brain delivery of macromolecules mediated by cell-penetrating peptides.

    PubMed

    Lin, Tingting; Liu, Ergang; He, Huining; Shin, Meong Cheol; Moon, Cheol; Yang, Victor C; Huang, Yongzhuo

    2016-07-01

    Brain delivery of macromolecular therapeutics (e.g., proteins) remains an unsolved problem because of the formidable blood-brain barrier (BBB). Although a direct pathway of nose-to-brain transfer provides an answer to circumventing the BBB and has already been intensively investigated for brain delivery of small drugs, new challenges arise for intranasal delivery of proteins because of their larger size and hydrophilicity. In order to overcome the barriers and take advantage of available pathways (e.g., epithelial tight junctions, uptake by olfactory neurons, transport into brain tissues, and intra-brain diffusion), a low molecular weight protamine (LMWP) cell-penetrating peptide was utilized to facilitate nose-to-brain transport. Cell-penetrating peptides (CPP) have been widely used to mediate macromolecular delivery through many kinds of biobarriers. Our results show that conjugates of LMWP-proteins are able to effectively penetrate into the brain after intranasal administration. The CPP-based intranasal method highlights a promising solution for protein therapy of brain diseases.

  15. Olfactory cells via nasal biopsy reflect the developing brain in gene expression profiles: utility and limitation of the surrogate tissues in research for brain disorders.

    PubMed

    Horiuchi, Yasue; Kano, Shin-Ichi; Ishizuka, Koko; Cascella, Nicola G; Ishii, Seiji; Talbot, C Conover; Jaffe, Andrew E; Okano, Hideyuki; Pevsner, Jonathan; Colantuoni, Carlo; Sawa, Akira

    2013-12-01

    Human olfactory cells obtained by rapid nasal biopsy have been suggested to be a good surrogate system to address brain disease-associated molecular changes. Nonetheless, whether use of this experimental strategy is justified remains unclear. Here we compared expression profiles of olfactory cells systematically with those from the brain tissues and other cells. Principal component analysis indicated that the expression profiles of olfactory cells are very different from those of blood cells, but are closer to those of stem cells, in particular mesenchymal stem cells, that can be differentiated into the cells of the central nervous system.

  16. Olfactory cells via nasal biopsy reflect the developing brain in gene expression profiles: utility and limitation of the surrogate tissues in research for brain disorders

    PubMed Central

    Horiuchi, Yasue; Kano, Shin-ichi; Ishizuka, Koko; Cascella, Nicola G.; Ishii, Seiji; Talbot, C. Conover; Jaffe, Andrew E.; Okano, Hideyuki; Pevsner, Jonathan; Colantuoni, Carlo; Sawa, Akira

    2014-01-01

    Human olfactory cells obtained by rapid nasal biopsy have been suggested to be a good surrogate system to address brain disease-associated molecular changes. Nonetheless, whether use of this experimental strategy is justified remains unclear. Here we compared expression profiles of olfactory cells systematically with those from the brain tissues and other cells. Principal component analysis indicated that the expression profiles of olfactory cells are very different from those of blood cells, but are closer to those of stem cells, in particular mesenchymal stem cells, that can be differentiated into the cells of the central nervous system. PMID:24120685

  17. Autofluorescent cells in rat brain can be convincing impostors in green fluorescent reporter studies.

    PubMed

    Spitzer, Nadja; Sammons, Gregory S; Price, Elmer M

    2011-04-15

    Cell transplant and gene therapies are promising approaches to many disorders of the nervous system. In studies involving cell transplants to the brain or nervous system, expression of green fluorescent protein (GFP) is commonly used to label cells, allowing their identification and histological assessment even after long post-operative survival times. Techniques employing viral tracing or reporter genes also commonly use GFP to label cells. Here, we document the presence of a subpopulation of green autofluorescent cells in the cortex and hippocampus of formaldehyde fixed, cryosectioned rat brains aged 3-9 months. Using standard microscopic fluorescence imaging techniques, we acquired clear images of green autofluorescent cells, complete with extensive processes, which appear to be well integrated into the host tissue. Treatment of brain sections with sodium borohydride followed by cupric sulfate in ammonium acetate buffer reduced background and cellular autofluorescence throughout sections but, especially in hippocampus, did not eliminate considerable green fluorescence in a subset of neurons. This autofluorescence was weak and would therefore pose a problem only when cells weakly express GFP or when few labeled cells survive. We suggest that investigators be aware of the potential for false positives, especially if the cells expressing GFP are expected to migrate widely from the transplant site. Parallel sections from naïve brains should regularly be processed and imaged alongside experimental brain sections, and anti-GFP immunohistochemistry should be performed to ensure that true GFP+ signals are imaged instead of endogenous autofluorescent neurons.

  18. Pericyte abundance affects sucrose permeability in cultures of rat brain microvascular endothelial cells.

    PubMed

    Parkinson, Fiona E; Hacking, Cindy

    2005-07-05

    The blood-brain barrier is a physical and metabolic barrier that restricts diffusion of blood-borne substances into brain. In vitro models of the blood-brain barrier are used to characterize this structure, examine mechanisms of damage and repair and measure permeability of test substances. The core component of in vitro models of the blood-brain barrier is brain microvascular endothelial cells. We cultured rat brain microvascular endothelial cells (RBMEC) from isolated rat cortex microvessels. After 2-14 days in vitro (DIV), immunohistochemistry of these cells showed strong labeling for zona occludens 1 (ZO-1), a tight junction protein expressed in endothelial cells. Pericytes were also present in these cultures, as determined by expression of alpha-actin. The present study was performed to test different cell isolation methods and to compare the resulting cell cultures for abundance of pericytes and for blood-brain barrier function, as assessed by 14C-sucrose flux. Two purification strategies were used. First, microvessels were preabsorbed onto uncoated plastic for 4 h, then unattached microvessels were transferred to coated culture ware. Second, microvessels were incubated with an antibody to platelet-endothelial cell adhesion molecule 1 (PECAM-1; CD31) precoupled to magnetic beads, and a magnetic separation procedure was performed. Our results indicate that immunopurification, but not preadsorption, was an effective method to purify microvessels and reduce pericyte abundance in the resulting cultures. This purification significantly reduced 14C-sucrose fluxes across cell monolayers. These data indicate that pericytes can interfere with the development of blood-brain barrier properties in in vitro models that utilize primary cultures of RBMECs.

  19. Ectopic cell cycle proteins predict the sites of neuronal cell death in Alzheimer's disease brain.

    PubMed

    Busser, J; Geldmacher, D S; Herrup, K

    1998-04-15

    Alzheimer's disease (AD) is a major dementing illness characterized by regional concentrations of senile plaques, neurofibrillary tangles, and extensive neuronal cell death. Although cell and synaptic loss is most directly linked to the severity of symptoms, the mechanisms leading to the neuronal death remain unclear. Based on evidence linking neuronal death during development to unexpected reappearance of cell cycle events, we investigated the brains of 12 neuropathologically verified cases of Alzheimer's disease and eight age-matched, disease-free controls for the presence of cell cycle proteins. Aberrant expression of cyclin D, cdk4, proliferating cell nuclear antigen, and cyclin B1 were identified in the hippocampus, subiculum, locus coeruleus, and dorsal raphe nuclei, but not inferotemporal cortex or cerebellum of AD cases. With only one exception, control subjects showed no significant expression of cell cycle markers in any of the six regions. We propose that disregulation of various components of the cell cycle is a significant contributor to regionally specific neuronal death in AD.

  20. Concise review: Patient-derived olfactory stem cells: new models for brain diseases.

    PubMed

    Mackay-Sim, Alan

    2012-11-01

    Traditional models of brain diseases have had limited success in driving candidate drugs into successful clinical translation. This has resulted in large international pharmaceutical companies moving out of neuroscience research. Cells are not brains, obviously, but new patient-derived stem models have the potential to elucidate cell biological aspects of brain diseases that are not present in worm, fly, or rodent models, the work horses of disease investigations and drug discovery. Neural stem cells are present in the olfactory mucosa, the organ of smell in the nose. Patient-derived olfactory mucosa has demonstrated disease-associated differences in a variety of brain diseases and recently olfactory mucosa stem cells have been generated from patients with schizophrenia, Parkinson's disease, and familial dysautonomia. By comparison with cells from healthy controls, patient-derived olfactory mucosa stem cells show disease-specific alterations in gene expression and cell functions including: a shorter cell cycle and faster proliferation in schizophrenia, oxidative stress in Parkinson's disease, and altered cell migration in familial dysautonomia. Olfactory stem cell cultures thus reveal patient-control differences, even in complex genetic diseases such as schizophrenia and Parkinson's disease, indicating that multiple genes of small effect can converge on shared cell signaling pathways to present as a disease-specific cellular phenotype. Olfactory mucosa stem cells can be maintained in homogeneous cultures that allow robust and repeatable multiwell assays suitable for screening libraries of drug candidate molecules.

  1. The Pluripotency of Neural Crest Cells and Their Role in Brain Development.

    PubMed

    Le Douarin, Nicole M; Dupin, Elisabeth

    2016-01-01

    The neural crest (NC) is, in the Chordate phylum, an innovation of vertebrates, which exhibits several original characteristics: its component cells are pluripotent and give rise to both ectodermal and mesodermal cell types. Moreover, during the early stages of neurogenesis, the NC cells exert a paracrine stimulating effect on the development of the preotic brain.

  2. Tridermal tumorigenesis of induced pluripotent stem cells transplanted in ischemic brain.

    PubMed

    Kawai, Hiromi; Yamashita, Toru; Ohta, Yasuyuki; Deguchi, Kentaro; Nagotani, Shoko; Zhang, Xuemei; Ikeda, Yoshio; Matsuura, Tohru; Abe, Koji

    2010-08-01

    Stroke is a major neurologic disorder. Induced pluripotent stem (iPS) cells can be produced from basically any part of patients, with high reproduction ability and pluripotency to differentiate into various types of cells, suggesting that iPS cells can provide a hopeful therapy for cell transplantation. However, transplantation of iPS cells into ischemic brain has not been reported. In this study, we showed that the iPS cells fate in a mouse model of transient middle cerebral artery occlusion (MCAO). Undifferentiated iPS cells (5 x 10(5)) were transplanted into ipsilateral striatum and cortex at 24 h after 30 mins of transient MCAO. Behavioral and histologic analyses were performed at 28 day after the cell transplantation. To our surprise, the transplanted iPS cells expanded and formed much larger tumors in mice postischemic brain than in sham-operated brain. The clinical recovery of the MCAO+iPS group was delayed as compared with the MCAO+PBS (phosphate-buffered saline) group. iPS cells formed tridermal teratoma, but could supply a great number of Dcx-positive neuroblasts and a few mature neurons in the ischemic lesion. iPS cells have a promising potential to provide neural cells after ischemic brain injury, if tumorigenesis is properly controlled.

  3. Lung endothelial cells strengthen, but brain endothelial cells weaken barrier properties of a human alveolar epithelium cell culture model.

    PubMed

    Neuhaus, Winfried; Samwer, Fabian; Kunzmann, Steffen; Muellenbach, Ralf M; Wirth, Michael; Speer, Christian P; Roewer, Norbert; Förster, Carola Y

    2012-11-01

    The blood-air barrier in the lung consists of the alveolar epithelium, the underlying capillary endothelium, their basement membranes and the interstitial space between the cell layers. Little is known about the interactions between the alveolar and the blood compartment. The aim of the present study was to gain first insights into the possible interplay between these two neighbored cell layers. We established an in vitro Transwell model of the alveolar epithelium based on human cell line H441 and investigated the influence of conditioned medium obtained from human lung endothelial cell line HPMEC-ST1.6R on the barrier properties of the H441 layers. As control for tissue specificity H441 layers were exposed to conditioned medium from human brain endothelial cell line hCMEC/D3. Addition of dexamethasone was necessary to obtain stable H441 cell layers. Moreover, dexamethasone increased expression of cell type I markers (caveolin-1, RAGE) and cell type II marker SP-B, whereas decreased the transepithelial electrical resistance (TEER) in a concentration dependent manner. Soluble factors obtained from the lung endothelial cell line increased the barrier significantly proven by TEER values and fluorescein permeability on the functional level and by the differential expression of tight junctional proteins on the molecular level. In contrast to this, soluble factors derived from brain endothelial cells weakened the barrier significantly. In conclusion, soluble factors from lung endothelial cells can strengthen the alveolar epithelium barrier in vitro, which suggests communication between endothelial and epithelial cells regulating the integrity of the blood-air barrier.

  4. Cell-free DNA as a marker for prediction of brain damage in traumatic brain injury in rats.

    PubMed

    Ohayon, Sharon; Boyko, Matthew; Saad, Amit; Douvdevani, Amos; Gruenbaum, Benjamin F; Melamed, Israel; Shapira, Yoram; Teichberg, Vivian I; Zlotnik, Alexander

    2012-01-20

    Traumatic brain injury (TBI) is a major cause of morbidity and mortality, and early predictors of neurological outcomes are of great clinical importance. Cell free DNA (CFD), a biomarker used for the diagnosis and monitoring of several diseases, has been implicated as a possible prognostic indicator after TBI. The purpose of this study was to determine the pattern and timing of CFD levels after TBI, and whether a relationship exists between the level of CFD and brain edema and neurological outcomes. Thirty-nine Sprague-Dawley rats were randomly assigned to two groups: rats in group 1 (sham group) were anesthetized and had a scalp incision without TBI, and rats in group 2 were anesthetized and had a scalp incision with TBI, which was induced by using a weight drop model that causes diffuse brain injury. A neurological severity score (NSS) was assessed at 1, 24, and 48 h after TBI. CFD was measured via blood samples drawn at t=0 (baseline), 12, 24, 48, 72, and 120 h after TBI. At 48 h after TBI, brain edema was determined in a subgroup of 11 rats by calculating the difference between rats' wet and dry brain weight. The significance of comparisons between and within groups (CFD levels, brain water content, and NSS) were determined using the Kruskal-Wallis, Mann-Whitney and Student t test. The correlation between CFD levels and the NSS, as well as between CFD levels and the extent of brain edema, was calculated using the Spearman and Pearson tests, respectively. Compared with baseline levels, the CFD levels in rats subjected to TBI were significantly increased at 24 and 48 h after TBI (p<0.01 and p<0.05, respectively). A positive correlation was demonstrated between CFD levels 24 h following TBI and the extent of brain edema (r=0.63, p<0.05), as well as between CFD levels and the NSS (r=0.79, p<0.005). In this study, we demonstrated an increase in CFD levels after TBI, as well as a correlation between CFD levels and brain edema and NSS. CFD levels may provide a

  5. Plant cytoplasm preserved by lightning.

    PubMed

    Wang, X

    2004-10-01

    Usually only an organism with hard parts may be preserved in the fossil record. Cytoplasm, which is a physiologically active part of a plant, is rarely seen in the fossil record. Two Cretaceous plant fossils older than 100 million years with exceptional preservation of cytoplasm are reported here. Some cytoplasm is well preserved with subcellular details while other cytoplasm is highly hydrolyzed in the cortex of the same fossil even though both of preservations may be less than 2 microm away. The unique preservation pattern, sharp contrast of preservation in adjacent cells and the exceptional preservation of cytoplasm in the cortex suggest that lightning should play an important role in the preservation of cytoplasm and that cytoplasmic membranes may be more stable than the cell contents. Interpreting the preservation needs knowledge scattering in several formerly unrelated fields of science, including geophysics, botany, biophysics, cytology and microwave fixation technology. This new interpretation of fossilization will shed new light on preservation of cytoplasm and promote cytoplasm fossils from a position of rarity to a position of common research objects available for biological research. The importance of the identification of cytoplasm in fossil lies not in itself but in how much it influences the future research in paleobotany.

  6. Intramyocardial Delivery of Mesenchymal Stem Cell-Seeded Hydrogel Preserves Cardiac Function and Attenuates Ventricular Remodeling after Myocardial Infarction

    PubMed Central

    Mathieu, Eva; Lamirault, Guillaume; Toquet, Claire; Lhommet, Pierre; Rederstorff, Emilie; Sourice, Sophie; Biteau, Kevin; Hulin, Philippe; Forest, Virginie; Weiss, Pierre

    2012-01-01

    Background To improve the efficacy of bone marrow-derived mesenchymal stem cell (MSC) therapy targeted to infarcted myocardium, we investigated whether a self-setting silanized hydroxypropyl methylcellulose (Si-HPMC) hydrogel seeded with MSC (MSC+hydrogel) could preserve cardiac function and attenuate left ventricular (LV) remodeling during an 8-week follow-up study in a rat model of myocardial infarction (MI). Methodology/Principal Finding Si-HPMC hydrogel alone, MSC alone or MSC+hydrogel were injected into the myocardium immediately after coronary artery ligation in female Lewis rats. Animals in the MSC+hydrogel group showed an increase in cardiac function up to 28 days after MI and a mid-term prevention of cardiac function alteration at day 56. Histological analyses indicated that the injection of MSC+hydrogel induced a decrease in MI size and an increase in scar thickness and ultimately limited the transmural extent of MI. These findings show that intramyocardial injection of MSC+hydrogel induced short-term recovery of ventricular function and mid-term attenuation of remodeling after MI. Conclusion/Significance These beneficial effects may be related to the specific scaffolding properties of the Si-HPMC hydrogel that may provide the ability to support MSC injection and engraftment within myocardium. PMID:23284842

  7. Frequency of brain metastasis in adenocarcinoma and large cell carcinoma of the lung: correlation with survival

    SciTech Connect

    Komaki, R.; Cox, J.D.; Stark, R.

    1983-10-01

    From January 1970 through December 1981, 469 patients with histologically or cytologically proven adenocarcinoma (AC) (349) and large cell carcinoma (LC) (120) of the lung were seen at the Department of Radiation Oncology, Medical College of Wisconsin Affiliated Hospitals. One quarter (126/469) of these patients had brain metastasis: 48 patients presented with brain metastasis and 78 patients subsequently developed brain metastasis. Brain was the dominant site of metastasis in 82 patients who received only cranial + thoracic irradiation; 37 patients (17 simultaneous, 20 metachronous) also required irradiation of other sites of metastasis. All 17 patients with LC, and 47/61 (77%) with AC who developed metachronous brain metastasis did so within one year. The cumulative probability of brain metastasis increased with survival to the levels predicted by autopsy studies. Therapeutic brain irradiation may result in long-term survival in patients with single organ brain metastasis. Since patients with AC and LC so frequently develop brain metastasis and the brain may be the only site of metastasis, prophylactic cranial irradiation may significantly reduce morbidity and mortality from these diseases.

  8. ALA-PDT of glioma cell micro-clusters in BD-IX rat brain

    NASA Astrophysics Data System (ADS)

    Madsen, Steen J.; Angell-Petersen, Even; Spetalen, Signe; Carper, Stephen W.; Ziegler, Sarah A.; Hirschberg, Henry

    2006-02-01

    A significant contributory factor to the poor prognosis of patients with glioblastoma multiforme is the inability of conventional treatments to eradicate infiltrating glioma cells. A syngeneic rat brain tumor model is used to investigate the effects of aminolevulinic acid (ALA)-mediated photodynamic therapy (PDT) on small clusters of tumor cells sequestered in normal brain. The intrinsic sensitivity of rat glioma cells to PDT was investigated by exposing ALA-incubated cells to a range of radiant exposures and irradiances using 635 nm light. Biodistribution studies were undertaken on tumor-bearing animals in order to determine the tumor selectivity of the photosensitizer following systemic administration (i.p.) of ALA. Effects of ALA-PDT on normal brain and gross tumor were evaluated using histopathology. Effects of PDT on isolated glioma cells in normal brain were investigated by treating animals 48 h after tumor cell implantation: a time when the micro-clusters of cells are protected by an intact blood-brain-barrier (BBB). Rat glioma cells in monolayer are susceptible to ALA-PDT - lower irradiances are more effective than higher ones. Fluorescence microscopy of frozen tissue sections showed that photosensitizer is produced with better than 200:1 tumor-to-normal tissue selectivity following i.p. ALA administration. ALA-PDT resulted in significant damage to both gross tumor and normal brain, however, the treatment failed to prolong survival of animals with newly implanted glioma cells compared to non-treated controls if the drug was delivered either i.p. or directly into the brain. In contrast, animals inoculated with tumor cells pre-incubated in vitro with ALA showed a significant survival advantage in response to PDT.

  9. Calcium-dependent neuroepithelial contractions expel damaged cells from the developing brain

    PubMed Central

    Herrgen, Leah; Voss, Oliver P.; Akerman, Colin J.

    2016-01-01

    Summary Both developing and adult organisms need efficient strategies for wound repair. In adult mammals, wounding triggers an inflammatory response that can exacerbate tissue injury and lead to scarring. In contrast, embryonic wounds heal quickly and with minimal inflammation, but how this is achieved remains incompletely understood. Using in vivo imaging in the developing brain of Xenopus laevis, we show that ATP release from damaged cells and subsequent activation of purinergic receptors induce long-range calcium waves in neural progenitor cells. Cytoskeletal reorganization, and activation of the actomyosin contractile machinery in a Rho kinase-dependent manner, then lead to rapid and pronounced apical-basal contractions of the neuroepithelium. These contractions drive the expulsion of damaged cells into the brain ventricle within seconds. Successful cell expulsion prevents the death of nearby cells and an exacerbation of the injury. Cell expulsion through neuroepithelial contraction represents a novel mechanism for rapid wound healing in the developing brain. PMID:25468753

  10. Cre Fused with RVG Peptide Mediates Targeted Genome Editing in Mouse Brain Cells In Vivo

    PubMed Central

    Zou, Zhiyuan; Sun, Zhaolin; Li, Pan; Feng, Tao; Wu, Sen

    2016-01-01

    Cell penetrating peptides (CPPs) are short peptides that can pass through cell membranes. CPPs can facilitate the cellular entry of proteins, macromolecules, nanoparticles and drugs. RVG peptide (RVG hereinafter) is a 29-amino-acid CPP derived from a rabies virus glycoprotein that can cross the blood-brain barrier (BBB) and enter brain cells. However, whether RVG can be used for genome editing in the brain has not been reported. In this work, we combined RVG with Cre recombinase for bacterial expression. The purified RVG-Cre protein cut plasmids in vitro and traversed cell membranes in cultured Neuro2a cells. By tail vein-injecting RVG-Cre into Cre reporter mouse lines mTmG and Rosa26lacZ, we demonstrated that RVG-Cre could target brain cells and achieve targeted somatic genome editing in adult mice. This direct delivery of the gene-editing enzyme protein into mouse brains with RVG is much safer than plasmid- or viral-based methods, holding promise for further applications in the treatment of various brain diseases. PMID:27983648

  11. GAD1 Upregulation Programs Aggressive Features of Cancer Cell Metabolism in the Brain Metastatic Microenvironment.

    PubMed

    Schnepp, Patricia M; Lee, Dennis D; Guldner, Ian H; O'Tighearnaigh, Treasa K; Howe, Erin N; Palakurthi, Bhavana; Eckert, Kaitlyn E; Toni, Tiffany A; Ashfeld, Brandon L; Zhang, Siyuan

    2017-04-11

    The impact of altered amino acid metabolism on cancer progression is not fully understood. We hypothesized that a metabolic transcriptome shift during metastatic evolution is crucial for brain metastasis. Here we report a powerful impact in this setting caused by epigenetic upregulation of glutamate decarboxylase 1 (GAD1), a regulator of the GABA neurotransmitter metabolic pathway. In cell-based culture and brain metastasis models, we found that downegulation of the DNA methyltransferase DNMT1 induced by the brain microenvironment-derived clusterin resulted in decreased GAD1 promoter methylation and subsequent upregulation of GAD1 expression in brain metastatic tumor cells. In a system to dynamically visualize cellular metabolic responses mediated by GAD1, we monitored the cytosolic NADH:NAD+ equilibrium in tumor cells. Reducing GAD1 in metastatic cells by primary glia cell co-culture abolished the capacity of metastatic cells to utilize extracellular glutamine, leading to cytosolic accumulation of NADH and increased oxidative status. Similarly, genetic or pharmacological disruption of the GABA metabolic pathway decreased the incidence of brain metastasis in vivo. Taken together, our results show how epigenetic changes in GAD1 expression alter local glutamate metabolism in the brain metastatic microenvironment, contributing to a metabolic adaption that facilitates metastasis outgrowth in that setting.

  12. Differential Activation of Infiltrating Monocyte-Derived Cells After Mild and Severe Traumatic Brain Injury.

    PubMed

    Trahanas, Diane M; Cuda, Carla M; Perlman, Harris; Schwulst, Steven J

    2015-03-01

    Microglia are the resident innate immune cells of the brain. Although embryologically and functionally distinct, they are morphologically similar to peripheral monocyte-derived cells, resulting in a poor ability to discriminate between the two cell types. The purpose of this study was to develop a rapid and reliable method to simultaneously characterize, quantify, and discriminate between whole populations of myeloid cells from the brain in a murine model of traumatic brain injury. Male C57BL/6 mice underwent traumatic brain injury (n = 16) or sham injury (n = 14). Brains were harvested at 24 h after injury. Multiparameter flow cytometry and sequential gating analysis were performed, allowing for discrimination between microglia and infiltrating leukocytes as well as for the characterization and quantification of individual subtypes within the infiltrating population. The proportion of infiltrating leukocytes within the brain increased with the severity of injury, and the predominant cell types within the infiltrating population were monocyte derived (P = 0.01). In addition, the severity of injury altered the overall makeup of the infiltrating monocyte-derived cells. In conclusion, we describe a flow cytometry-based technique for gross discrimination between infiltrating leukocytes and microglia as well as the ability to simultaneously characterize and quantify individual myeloid subtypes and their maturation states within these populations.

  13. Cell and tissue kinetics of the subependymal layer in mouse brain following heavy charged particle irradiation

    SciTech Connect

    Manley, N.B.; Fabrikant, J.I.; Alpen, E.L.

    1988-12-01

    The following studies investigate the cellular response and cell population kinetics of the subependymal layer in the mouse brain exposed to heavy charged particle irradiation. Partial brain irradiation with helium and neon ions was confined to one cortex of the brain. Both the irradiated and the unirradiated contralateral cortex showed similar disturbances of the cell and tissue kinetics in the subependymal layers. The irradiated hemisphere exhibited histological damage, whereas the unirradiated side appeared normal histologically. This study concerns the cell population and cell cycle kinetics of the subependymal layer in the mouse brain, and the effects of charged particle irradiations on this cell population. Quantitative high resolution autoradiography was used to study the kinetic parameters in this cell layer. This study should help in understanding the effects of these high-energy heavy ions on normal mammalian brain tissue. The response of the mammalian brain exposure to charged particle ionizing radiation may be extremely variable. It varies from minimal physiological changes to overt tissue necrosis depending on a number of factors such as: the administered dose, dose-rate, the volume of the irradiated tissue, and the biological end-point being examined.

  14. Evidence of endothelial progenitor cells in the human brain and spinal cord arteriovenous malformations

    PubMed Central

    Gao, Peng; Chen, Yongmei; Lawton, Michael T.; Barbaro, Nicholas M.; Yang, Guo-Yuan; Su, Hua; Ling, Feng; Young, William L.

    2010-01-01

    Objective Brain and spinal cord arteriovenous malformations (AVMs) are characterized by aberrant angiogenesis and vascular remodeling. Endothelial progenitor cells (EPCs) can be recruited by stromal cell-derived factor-1 (SDF-1), and participate in vascular remodeling in both physiological and pathological settings. We hypothesized that there was increased EPC levels in the brain and spinal cord AVM nidus. Methods Microsurgical specimens without endovascular embolization and radiosurgery from the brain (n=12) and spinal cord (n=5) AVMs were examined. Hemangioblastoma, meningioma, cerebral cortex obtained from epilepsy surgery, and the basilar artery (BA) from the autopsy were chosen for control comparisons. EPCs were identified as cells that were double-positive for the stem cell marker CD133 and the endothelial cell marker VEGFR-2 (vascular endothelial growth factor receptor-2 or KDR). In addition, SDF-1 was characterized by immunohistochemistry. Results Both brain and spinal AVM tissues displayed more CD133, SDF-1, and CD68-positive signals than epilepsy and basilar artery control tissues. The level of EPCs was increased in the brain and spinal cord AVM nidus, mainly at the edge of the vessel wall. The expression of SDF-1 was co-localized with CD31-positive and α-smooth muscle cells, and was predominantly found within the vessel wall. Conclusion Our data demonstrate that EPCs are present in the nidus of the brain and spinal cord AVMs, which may mediate pathological vascular remodeling and impact the clinical course of AVMs. PMID:20881566

  15. Characterization of atrial natriuretic peptide receptors in brain microvessel endothelial cells

    NASA Technical Reports Server (NTRS)

    Whitson, Peggy A.; Huls, M. H.; Sams, Clarence F.

    1989-01-01

    In view of the suggestions by Chabrier et al. (1987) and Steardo and Nathanson (1987) that atrial natriuretic peptide (ANP) may play a role in the fluid homeostasis of the brain, the ANP receptors in primary cultures of bovine brain microvessel endothelian cells were quantitated and characterized. Results of partition binding studies and the effect of cGMP additions indicated the presence of at least two types of ANP receptors, with the majority of the receptors being the nonguanylate cyclase coupled receptors. The presence of at least two ANP receptor types suggests an active role for ANP in regulating brain endothelial cell function.

  16. Using fluorescence activated cell sorting to examine cell-type-specific gene expression in rat brain tissue.

    PubMed

    Schwarz, Jaclyn M

    2015-05-28

    The brain is comprised of four primary cell types including neurons, astrocytes, microglia and oligodendrocytes. Though they are not the most abundant cell type in the brain, neurons are the most widely studied of these cell types given their direct role in impacting behaviors. Other cell types in the brain also impact neuronal function and behavior via the signaling molecules they produce. Neuroscientists must understand the interactions between the cell types in the brain to better understand how these interactions impact neural function and disease. To date, the most common method of analyzing protein or gene expression utilizes the homogenization of whole tissue samples, usually with blood, and without regard for cell type. This approach is an informative approach for examining general changes in gene or protein expression that may influence neural function and behavior; however, this method of analysis does not lend itself to a greater understanding of cell-type-specific gene expression and the effect of cell-to-cell communication on neural function. Analysis of behavioral epigenetics has been an area of growing focus which examines how modifications of the deoxyribonucleic acid (DNA) structure impact long-term gene expression and behavior; however, this information may only be relevant if analyzed in a cell-type-specific manner given the differential lineage and thus epigenetic markers that may be present on certain genes of individual neural cell types. The Fluorescence Activated Cell Sorting (FACS) technique described below provides a simple and effective way to isolate individual neural cells for the subsequent analysis of gene expression, protein expression, or epigenetic modifications of DNA. This technique can also be modified to isolate more specific neural cell types in the brain for subsequent cell-type-specific analysis.

  17. Antibody response to a T-cell-independent antigen is preserved after splenic artery embolization for trauma.

    PubMed

    Olthof, D C; Lammers, A J J; van Leeuwen, E M M; Hoekstra, J B L; ten Berge, I J M; Goslings, J C

    2014-11-01

    Splenic artery embolization (SAE) is increasingly being used as a nonoperative management strategy for patients with blunt splenic injury following trauma. The aim of this study was to assess the splenic function of patients who were embolized. A clinical study was performed, with splenic function assessed by examining the antibody response to polysaccharide antigens (pneumococcal 23-valent polysaccharide vaccine), B-cell subsets, and the presence of Howell-Jolly bodies (HJB). The data were compared to those obtained from splenectomized patients and healthy controls (HC) who had been included in a previously conducted study. A total of 30 patients were studied: 5 who had proximal SAE, 7 who had distal SAE, 8 who had a splenectomy, and 10 HC. The median vaccine-specific antibody response of the SAE patients (fold increase, 3.97) did not differ significantly from that of the HC (5.29; P = 0.90); however, the median response of the splenectomized patients (2.30) did differ (P = 0.003). In 2 of the proximally embolized patients and none of the distally embolized patients, the ratio of the IgG antibody level postvaccination compared to that prevaccination was <2. There were no significant differences in the absolute numbers of lymphocytes or B-cell subsets between the SAE patients and the HC. HJB were not observed in the SAE patients. The splenic immune function of embolized patients was preserved, and therefore routine vaccination appears not to be indicated. Although the median antibody responses did not differ between the patients who underwent proximal SAE and those who underwent distal SAE, 2 of the 5 proximally embolized patients had insufficient responses to vaccination, whereas none of the distally embolized patients exhibited an insufficient response. Further research should be done to confirm this finding.

  18. Testosterone Production is Better Preserved After 16 than 20 Gray Irradiation Treatment Against Testicular Carcinoma In Situ Cells

    SciTech Connect

    Bang, Anne K.; Petersen, Jorgen H.; Petersen, Peter M.; Andersson, Anna-Maria; Daugaard, Gedske; Jorgensen, Niels

    2009-11-01

    Purpose: To study the effect of 16 Gy radiotherapy (RT) vs. 20 Gy RT on Leydig cell function in men treated with radiotherapy against carcinoma in situ (CIS) of the testis. Methods and Materials: Fifty-one men who were treated between 1985 and 2005 were included. Fourteen men had been treated with 20 Gy and 37 with 16 Gy RT. Measurements of sex hormone-binding globulin and basic and stimulated testosterone, as well as luteinizing hormone levels were performed. Results: The follow-up periods for the patients treated without additional chemotherapy were for the 20 Gy and 16 Gy group mean/median/min-max: 9.0/10.0/1.0-20.3 years and 4.0/3.1/0.4-14.1 years, respectively. During the follow-up period, men treated with 16 Gy RT had stable testosterone levels (-1.1%/year, p = 0.4), whereas men treated with 20 Gy had an annual decrease of 2.4% (p = 0.008). For the latter group, the testosterone decrease was most pronounced in the first 5 years, leveling off during the following 5 years. Additionally, more men treated with 20 Gy needed androgen substitution treatment. Our study showed an increased luteinizing hormone level for the men treated with 16 Gy, although this was not significant (p = 0.5). We anticipated a similar increase in the patients treated with 20 Gy but instead observed a decrease (-3.1%, p = 0.01). Conclusion: RT at 16 and 20 Gy seem to affect Leydig cell function differently, with 16 Gy RT better preserving testosterone levels and thus being preferred from an endocrinological point of view.

  19. Brain microvascular endothelial cell association and distribution of a 5 nm ceria engineered nanomaterial

    PubMed Central

    Dan, Mo; Tseng, Michael T; Wu, Peng; Unrine, Jason M; Grulke, Eric A; Yokel, Robert A

    2012-01-01

    Purpose: Ceria engineered nanomaterials (ENMs) have current commercial applications and both neuroprotective and toxic effects. Our hypothesis is that ceria ENMs can associate with brain capillary cells and/or cross the blood–brain barrier. Methods: An aqueous dispersion of ∼5 nm ceria ENM was synthesized and characterized in house. Its uptake space in the Sprague Dawley rat brain was determined using the in situ brain perfusion technique at 15 and 20 mL/minute flow rates; 30, 100, and 500 μg/mL ceria perfused for 120 seconds at 20 mL/minute; and 30 μg/mL perfused for 20, 60, and 120 seconds at 20 mL/minute. The capillary depletion method and light and electron microscopy were used to determine its capillary cell and brain parenchymal association and localization. Results: The vascular space was not significantly affected by brain perfusion flow rate or ENM, demonstrating that this ceria ENM did not influence blood–brain barrier integrity. Cerium concentrations, determined by inductively coupled plasma mass spectrometry, were significantly higher in the choroid plexus than in eight brain regions in the 100 and 500 μg/mL ceria perfusion groups. Ceria uptake into the eight brain regions was similar after 120-second perfusion of 30, 100, and 500 μg ceria/mL. Ceria uptake space significantly increased in the eight brain regions and choroid plexus after 60 versus 20 seconds, and it was similar after 60 and 120 seconds. The capillary depletion method showed 99.4% ± 1.1% of the ceria ENM associated with the capillary fraction. Electron microscopy showed the ceria ENM located on the endothelial cell luminal surface. Conclusion: Ceria ENM association with brain capillary endothelial cells saturated between 20 and 60 seconds and ceria ENM brain uptake was not diffusion-mediated. During the 120-second ceria ENM perfusion, ceria ENM predominately associated with the surface of the brain capillary cells, providing the opportunity for its cell uptake or redistribution

  20. NFL-lipid nanocapsules for brain neural stem cell targeting in vitro and in vivo.

    PubMed

    Carradori, Dario; Saulnier, Patrick; Préat, Véronique; des Rieux, Anne; Eyer, Joel

    2016-09-28

    The replacement of injured neurons by the selective stimulation of neural stem cells in situ represents a potential therapeutic strategy for the treatment of neurodegenerative diseases. The peptide NFL-TBS.40-63 showed specific interactions towards neural stem cells of the subventricular zone. The aim of our work was to produce a NFL-based drug delivery system able to target neural stem cells through the selective affinity between the peptide and these cells. NFL-TBS.40-63 (NFL) was adsorbed on lipid nanocapsules (LNC) whom targeting efficiency was evaluated on neural stem cells from the subventricular zone (brain) and from the central canal (spinal cord). NFL-LNC were incubated with primary neural stem cells in vitro or injected in vivo in adult rat brain (right lateral ventricle) or spinal cord (T10). NFL-LNC interactions with neural stem cells were different depending on the origin of the cells. NFL-LNC showed a preferential uptake by neural stem cells from the brain, while they did not interact with neural stem cells from the spinal cord. The results obtained in vivo correlate with the results observed in vitro, demonstrating that NFL-LNC represent a promising therapeutic strategy to selectively deliver bioactive molecules to brain neural stem cells.

  1. Dexamethasone suppresses JMJD3 gene activation via a putative negative glucocorticoid response element and maintains integrity of tight junctions in brain microvascular endothelial cells.

    PubMed

    Na, Wonho; Shin, Jee Y; Lee, Jee Y; Jeong, Sangyun; Kim, Won-Sun; Yune, Tae Y; Ju, Bong-Gun

    2017-01-01

    The blood-brain barrier (BBB) exhibits a highly selective permeability to support the homeostasis of the central nervous system (CNS). The tight junctions in the BBB microvascular endothelial cells seal the paracellular space to prevent diffusion. Thus, disruption of tight junctions results in harmful effects in CNS diseases and injuries. It has recently been demonstrated that glucocorticoids have beneficial effects on maintaining tight junctions in both in vitro cell and in vivo animal models. In the present study, we found that dexamethasone suppresses the expression of JMJD3, a histone H3K27 demethylase, via the recruitment of glucocorticoid receptor α (GRα) and nuclear receptor co-repressor (N-CoR) to the negative glucocorticoid response element (nGRE) in the upstream region of JMJD3 gene in brain microvascular endothelial cells subjected to TNFα treatment. The decreased JMJD3 gene expression resulted in the suppression of MMP-2, MMP-3, and MMP-9 gene activation. Dexamethasone also activated the expression of the claudin 5 and occludin genes. Collectively, dexamethasone attenuated the disruption of the tight junctions in the brain microvascular endothelial cells subjected to TNFα treatment. Therefore, glucocorticoids may help to preserve the integrity of the tight junctions in the BBB via transcriptional and post-translational regulation following CNS diseases and injuries.

  2. Vitamin B-complex initiates growth and development of human embryonic brain cells in vitro.

    PubMed

    Danielyan, K E; Abramyan, R A; Galoyan, A A; Kevorkian, G A

    2011-09-01

    We studied a combined effect of subcomponents of vitamin B complex on the growth, development, and death of human embryonic brain-derived cells (E90) cultured using a modified method of Matson. Cell death was detected by trypan blue staining. According to our results, vitamin B-complex in low-doses promote the development, maturation, and enlargement of human embryonic brain cells, on the one hand, and increases the percent of cell death, which attests to accelerated maturation and metabolism, on the other.

  3. Magnetic particle spectroscopy allows precise quantification of nanoparticles after passage through human brain microvascular endothelial cells

    NASA Astrophysics Data System (ADS)

    Gräfe, C.; Slabu, I.; Wiekhorst, F.; Bergemann, C.; von Eggeling, F.; Hochhaus, A.; Trahms, L.; Clement, J. H.

    2016-06-01

    Crossing the blood-brain barrier is an urgent requirement for the treatment of brain disorders. Superparamagnetic iron oxide nanoparticles (SPIONs) are a promising tool as carriers for therapeutics because of their physical properties, biocompatibility, and their biodegradability. In order to investigate the interaction of nanoparticles with endothelial cell layers in detail, in vitro systems are of great importance. Human brain microvascular endothelial cells are a well-suited blood-brain barrier model. Apart from generating optimal conditions for the barrier-forming cell units, the accurate detection and quantification of SPIONs is a major challenge. For that purpose we use magnetic particle spectroscopy to sensitively and directly quantify the SPION-specific iron content. We could show that SPION concentration depends on incubation time, nanoparticle concentration and location. This model system allows for further investigations on particle uptake and transport at cellular barriers with regard to parameters including particles’ shape, material, size, and coating.

  4. PACAP38/PAC1 Signaling Induces Bone Marrow-Derived Cells Homing to Ischemic Brain

    PubMed Central

    Lin, Chen-Huan; Chiu, Lian; Lee, Hsu-Tung; Chiang, Chun-Wei; Liu, Shih-Ping; Hsu, Yung-Hsiang; Lin, Shinn-Zong; Hsu, Chung Y; Hsieh, Chia-Hung; Shyu, Woei-Cherng

    2015-01-01

    Understanding stem cell homing, which is governed by environmental signals from the surrounding niche, is important for developing effective stem cell-based repair strategies. The molecular mechanism by which the brain under ischemic stress recruits bone marrow-derived cells (BMDCs) to the vascular niche remains poorly characterized. Here we report that hypoxia-inducible factor-1α (HIF-1α) activation upregulates pituitary adenylate cyclase-activating peptide 38 (PACAP38), which in turn activates PACAP type 1 receptor (PAC1) under hypoxia in vitro and cerebral ischemia in vivo. BMDCs homing to endothelial cells in the ischemic brain are mediated by HIF-1α activation of the PACAP38-PAC1 signaling cascade followed by upregulation of cellular prion protein and α6-integrin to enhance the ability of BMDCs to bind laminin in the vascular niche. Exogenous PACAP38 confers a similar effect in facilitating BMDCs homing into the ischemic brain, resulting in reduction of ischemic brain injury. These findings suggest a novel HIF-1α-activated PACAP38-PAC1 signaling process in initiating BMDCs homing into the ischemic brain for reducing brain injury and enhancing functional recovery after ischemic stroke. Stem Cells 2015;33:1153–1172 PMID:25523790

  5. NK cells promote neutrophil recruitment in the brain during sepsis-induced neuroinflammation

    PubMed Central

    He, Hao; Geng, Tingting; Chen, Piyun; Wang, Meixiang; Hu, Jingxia; Kang, Li; Song, Wengang; Tang, Hua

    2016-01-01

    Sepsis could affect the central nervous system and thus induces neuroinflammation, which subsequently leads to brain damage or dysfunction. However, the mechanisms of generation of neuroinflammation during sepsis remain poorly understood. By administration of lipopolysaccharides (LPS) in mice to mimic sepsis, we found that shortly after opening the blood–brain barrier, conventional CD11b+CD27+ NK subset migrated into the brain followed by subsequent neutrophil infiltration. Interestingly, depletion of NK cells prior to LPS treatment severely impaired neutrophil recruitment in the inflamed brain. By in vivo recruitment assay, we found that brain-infiltrated NK cells displayed chemotactic activity to neutrophils, which depended on the higher expression of chemokines such as CXCL2. Moreover, microglia were also responsible for neutrophil recruitment, and their chemotactic activity was significantly impaired by ablation of NK cells. Furthermore, depletion of NK cells could significantly ameliorate depression-like behavior in LPS-treated mice. These data indicated a NK cell-regulated neutrophil recruitment in the blamed brain, which also could be seen on another sepsis model, cecal ligation and puncture. So, our findings revealed an important scenario in the generation of sepsis-induced neuroinflammation. PMID:27270556

  6. A Triple Culture Model of the Blood-Brain Barrier Using Porcine Brain Endothelial cells, Astrocytes and Pericytes.

    PubMed

    Thomsen, Louiza Bohn; Burkhart, Annette; Moos, Torben

    2015-01-01

    In vitro blood-brain barrier (BBB) models based on primary brain endothelial cells (BECs) cultured as monoculture or in co-culture with primary astrocytes and pericytes are useful for studying many properties of the BBB. The BECs retain their expression of tight junction proteins and efflux transporters leading to high trans-endothelial electric resistance (TEER) and low passive paracellular permeability. The BECs, astrocytes and pericytes are often isolated from small rodents. Larger species as cows and pigs however, reveal a higher yield, are readily available and have a closer resemblance to humans, which make them favorable high-throughput sources for cellular isolation. The aim of the present study has been to determine if the preferable combination of purely porcine cells isolated from the 6 months old domestic pigs, i.e. porcine brain endothelial cells (PBECs) in co-culture with porcine astrocytes and pericytes, would compare with PBECs co-cultured with astrocytes and pericytes isolated from newborn rats with respect to TEER value and low passive permeability. The astrocytes and pericytes were grown both as contact and non-contact co-cultures as well as in triple culture to examine their effects on the PBECs for barrier formation as revealed by TEER, passive permeability, and expression patterns of tight junction proteins, efflux transporters and the transferrin receptor. This syngenic porcine in vitro BBB model is comparable to triple cultures using PBECs, rat astrocytes and rat pericytes with respect to TEER formation, low passive permeability, and expression of hallmark proteins signifying the brain endothelium (tight junction proteins claudin 5 and occludin, the efflux transporters P-glycoprotein (PgP) and breast cancer related protein (BCRP), and the transferrin receptor).

  7. Coculture system with an organotypic brain slice and 3D spheroid of carcinoma cells.

    PubMed

    Chuang, Han-Ning; Lohaus, Raphaela; Hanisch, Uwe-Karsten; Binder, Claudia; Dehghani, Faramarz; Pukrop, Tobias

    2013-10-09

    Patients with cerebral metastasis of carcinomas have a poor prognosis. However, the process at the metastatic site has barely been investigated, in particular the role of the resident (stromal) cells. Studies in primary carcinomas demonstrate the influence of the microenvironment on metastasis, even on prognosis(1,2). Especially the tumor associated macrophages (TAM) support migration, invasion and proliferation(3). Interestingly, the major target sites of metastasis possess tissue-specific macrophages, such as Kupffer cells in the liver or microglia in the CNS. Moreover, the metastatic sites also possess other tissue-specific cells, like astrocytes. Recently, astrocytes were demonstrated to foster proliferation and persistence of cancer cells(4,5). Therefore, functions of these tissue-specific cell types seem to be very important in the process of brain metastasis(6,7). Despite these observations, however, up to now there is no suitable in vivo/in vitro model available to directly visualize glial reactions during cerebral metastasis formation, in particular by bright field microscopy. Recent in vivo live imaging of carcinoma cells demonstrated their cerebral colonization behavior(8). However, this method is very laborious, costly and technically complex. In addition, these kinds of animal experiments are restricted to small series and come with a substantial stress for the animals (by implantation of the glass plate, injection of tumor cells, repetitive anaesthesia and long-term fixation). Furthermore, in vivo imaging is thus far limited to the visualization of the carcinoma cells, whereas interactions with resident cells have not yet been illustrated. Finally, investigations of human carcinoma cells within immunocompetent animals are impossible(8). For these reasons, we established a coculture system consisting of an organotypic mouse brain slice and epithelial cells embedded in matrigel (3D cell sphere). The 3D carcinoma cell spheres were placed directly next to

  8. Cellular immortality in brain tumours: an integration of the cancer stem cell paradigm.

    PubMed

    Rahman, Ruman; Heath, Rachel; Grundy, Richard

    2009-04-01

    Brain tumours are a diverse group of neoplasms that continue to present a formidable challenge in our attempt to achieve curable intervention. Our conceptual framework of human brain cancer has been redrawn in the current decade. There is a gathering acceptance that brain tumour formation is a phenotypic outcome of dysregulated neurogenesis, with tumours viewed as abnormally differentiated neural tissue. In relation, there is accumulating evidence that brain tumours, similar to leukaemia and many solid tumours, are organized as a developmental hierarchy which is maintained by a small fraction of cells endowed with many shared properties of tissue stem cells. Proof that neurogenesis persists throughout adult life, compliments this concept. Although the cancer cell of origin is unclear, the proliferative zones that harbour stem cells in the embryonic, post-natal and adult brain are attractive candidates within which tumour-initiation may ensue. Dysregulated, unlimited proliferation and an ability to bypass senescence are acquired capabilities of cancerous cells. These abilities in part require the establishment of a telomere maintenance mechanism for counteracting the shortening of chromosomal termini. A strategy based upon the synthesis of telomeric repeat sequences by the ribonucleoprotein telomerase, is prevalent in approximately 90% of human tumours studied, including the majority of brain tumours. This review will provide a developmental perspective with respect to normal (neurogenesis) and aberrant (tumourigenesis) cellular turnover, differentiation and function. Within this context our current knowledge of brain tumour telomere/telomerase biology will be discussed with respect to both its developmental and therapeutic relevance to the hierarchical model of brain tumourigenesis presented by the cancer stem cell paradigm.

  9. The effects of preserved red blood cells on the severe adverse events observed in patients infused with hemoglobin based oxygen carriers.

    PubMed

    Valeri, C Robert; Ragno, Gina

    2008-01-01

    The severe adverse events observed in patients who received hemoglobin based oxygen carriers (HBOCs) were associated with the Ringer's D.L lactate resuscitative solution administered and to the excipient used in the HBOCs containing Ringer's D,L lactate and the length of storage of the preserved RBC administered to the patient at the time that the HBOCs were infused. This paper reports the quality of the red blood cells preserved in the liquid state at 4 degrees C and that of previously frozen RBCs stored at 4 degrees C with regard to their survival, function and safety. Severe adverse events have been observed related to the length of storage of the liquid preserved RBC stored at 4 degrees C prior to transfusion. The current methods to preserve RBC in the liquid state in additive solutions at 4 degrees C maintain their survival and function for only 2 weeks. The freezing of red blood cells with 40% W/V glycerol and storage at -80 degrees C allows for storage at -80 degrees C for 10 years and following thawing, deglycerolization and storage at 4 degrees C in the additive solution (AS-3, Nutricel) for 2 weeks with acceptable 24 hour posttransfusion survival, less than 1% hemolysis, and moderately impaired oxygen transport function with no associated adverse events. Frozen deglycerolized RBCs are leukoreduced and contain less than 5% of residual plasma and non-plasma substances. Frozen deglycerolized RBCs are the ideal RBC product to transfuse patients receiving HBOCs.

  10. West Nile virus infection modulates human brain microvascular endothelial cells tight junction proteins and cell adhesion molecules: Transmigration across the in vitro blood-brain barrier

    PubMed Central

    Verma, Saguna; Lo, Yeung; Chapagain, Moti; Lum, Stephanie; Kumar, Mukesh; Gurjav, Ulziijargal; Luo, Haiyan; Nakatsuka, Austin; Nerurkar, Vivek R.

    2009-01-01

    Neurological complications such as inflammation, failure of the blood-brain barrier (BBB), and neuronal death contribute to the mortality and morbidity associated with WNV-induced meningitis. Compromised BBB indicates the ability of the virus to gain entry into the CNS via the BBB, however, the underlying mechanisms, and the specific cell types associated with WNV-CNS trafficking are not well understood. Brain microvascular endothelial cells, main component of the BBB, represent a barrier to virus dissemination into the CNS and could play key role in WNV spread via hematogenous route. To investigate WNV entry into the CNS, we infected primary human brain microvascular endothelial (HBMVE) cells with the neurovirulent strain of WNV (NY99) and examined WNV replication kinetics together with the changes in the expressions of key tight junction proteins (TJP) and cell adhesion molecules (CAM). WNV infection of HBMVE cells was productive as analyzed by plaque assay and qRT-PCR, and did not induce cytopathic effect. Increased mRNA and protein expressions of TJP (claudin-1) and CAM (vascular cell adhesion molecule and E-selectin) were observed at days 2 and 3 after infection, respectively, which coincided with the peak in WNV replication. Further, using an in vitro BBB model comprised of HBMVE cells, we demonstrate that cell-free WNV can cross the BBB, without compromising the BBB integrity. These data suggest that infection of HBMVE cells can facilitate entry of cell-free virus into the CNS without disturbing the BBB, and increased CAM may assist in the trafficking of WNV-infected immune cells into the CNS, via ‘Trojan horse’ mechanism, thereby contributing to WNV dissemination in the CNS and associated pathology. PMID:19135695

  11. Wharton's Jelly Mesenchymal Stem Cells Protect the Immature Brain in Rats and Modulate Cell Fate.

    PubMed

    Mueller, Martin; Oppliger, Byron; Joerger-Messerli, Marianne; Reinhart, Ursula; Barnea, Eytan; Paidas, Michael; Kramer, Boris W; Surbek, Daniel V; Schoeberlein, Andreina

    2017-02-15

    The development of a mammalian brain is a complex and long-lasting process. Not surprisingly, preterm birth is the leading cause of death in newborns and children. Advances in perinatal care reduced mortality, but morbidity still represents a major burden. New therapeutic approaches are thus desperately needed. Given that mesenchymal stem/stromal cells (MSCs) emerged as a promising candidate for cell therapy, we transplanted MSCs derived from the Wharton's Jelly (WJ-MSCs) to reduce the burden of immature brain injury in a murine animal model. WJ-MSCs transplantation resulted in protective activity characterized by reduced myelin loss and astroglial activation. WJ-MSCs improved locomotor behavior as well. To address the underlying mechanisms, we tested the key regulators of responses to DNA-damaging agents, such as cyclic AMP-dependent protein kinase/calcium-dependent protein kinase (PKA/PKC), cyclin-dependent kinase (CDK), ataxia-telangiectasia-mutated/ATM- and Rad3-related (ATM/ATR) substrates, protein kinase B (Akt), and 14-3-3 binding protein partners. We characterized WJ-MSCs using a specific profiler polymerase chain reaction array. We provide evidence that WJ-MSCs target pivotal regulators of the cell fate such as CDK/14-3-3/Akt signaling. We identified leukemia inhibitory factor as a potential candidate of WJ-MSCs' induced modifications as well. We hypothesize that WJ-MSCs may exert adaptive responses depending on the type of injury they are facing, making them prominent candidates for cell therapy in perinatal injuries.

  12. Preservation of Antigen-Specific Functions of αβ T Cells and B Cells Removed from Hematopoietic Stem Cell Transplants Suggests Their Use As an Alternative Cell Source for Advanced Manipulation and Adoptive Immunotherapy

    PubMed Central

    Li Pira, Giuseppina; Di Cecca, Stefano; Biagini, Simone; Girolami, Elia; Cicchetti, Elisabetta; Bertaina, Valentina; Quintarelli, Concetta; Caruana, Ignazio; Lucarelli, Barbarella; Merli, Pietro; Pagliara, Daria; Brescia, Letizia Pomponia; Bertaina, Alice; Montanari, Mauro; Locatelli, Franco

    2017-01-01

    Hematopoietic stem cell transplantation is standard therapy for numerous hematological diseases. The use of haploidentical donors, sharing half of the HLA alleles with the recipient, has facilitated the use of this procedure as patients can rely on availability of a haploidentical donor within their family. Since HLA disparity increases the risk of graft-versus-host disease, T-cell depletion has been used to remove alloreactive lymphocytes from the graft. Selective removal of αβ T cells, which encompass the alloreactive repertoire, combined with removal of B cells to prevent EBV-related lymphoproliferative disease, proved safe and effective in clinical studies. Depleted αβ T cells and B cells are generally discarded as by-products. Considering the possible use of donor T cells for donor lymphocyte infusions or for generation of pathogen-specific T cells as mediators of graft-versus-infection effect, we tested whether cells in the discarded fractions were functionally intact. Response to alloantigens and to viral antigens comparable to that of unmanipulated cells indicated a functional integrity of αβ T cells, in spite of the manipulation used for their depletion. Furthermore, B cells proved to be efficient antigen-presenting cells, indicating that antigen uptake, processing, and presentation were fully preserved. Therefore, we propose that separated αβ T lymphocytes could be employed for obtaining pathogen-specific T cells, applying available methods for positive selection, which eventually leads to indirect allodepletion. In addition, these functional T cells could undergo additional manipulation, such as direct allodepletion or genetic modification. PMID:28386262

  13. Analysis of complement and plasma cells in the brain of patients with anti-NMDAR encephalitis

    PubMed Central

    Martinez-Hernandez, E.; Horvath, J.; Shiloh-Malawsky, Y.; Sangha, N.; Martinez-Lage, M.

    2011-01-01

    Objectives: Most patients with anti-NMDA receptor (NMDAR) encephalitis have intrathecal synthesis of antibodies, which cause a decrease of cell surface and synaptic NMDAR. Antibodies are immunoglobulin G (IgG)1 and IgG3 subtypes and can potentially activate complement. We examined whether complement immunoreactivity and antibody-secreting cells (plasma cells/plasmablasts) are present in the brain of these patients. Methods: Cultured rat hippocampal neurons were used in an immunocytochemical assay to test whether patients' antibodies can fix complement. Using the same reagents (antibodies to C9neo, C5b-9, C3), complement immunoreactivity was determined in the brain of 5 patients, the teratoma of 21 patients, and appropriate control tissues. A set of markers for B (CD20), T (CD3, CD4, CD8) and antibody-secreting cells (plasma cells/plasmablasts, CD138) were used to examine the brain inflammatory infiltrates. Results: Patients' antibodies were able to bind complement in vitro, but deposits of complement were not detected in patients' brain. Parallel experiments with teratomas showed that in contrast to the brain, the neural tissue of the tumors contained complement. Analysis of the inflammatory infiltrates in brain samples from autopsy or biopsy performed 3–4 weeks after symptom presentation demonstrated numerous antibody-secreting cells (CD138+) in perivascular, interstitial, and Virchow-Robin spaces, and B and T cells predominantly located in perivascular regions. Conclusions: Complement-mediated mechanisms do not appear to play a substantial pathogenic role in anti-NMDAR encephalitis. In contrast, there are copious infiltrates of antibody-secreting cells (plasma cells/plasmablasts) in the CNS of these patients. The demonstration of these cells provides an explanation for the intrathecal synthesis of antibodies and has implications for treatment. PMID:21795662

  14. Tumourigenicity and Immunogenicity of Induced Neural Stem Cell Grafts Versus Induced Pluripotent Stem Cell Grafts in Syngeneic Mouse Brain

    PubMed Central

    Gao, Mou; Yao, Hui; Dong, Qin; Zhang, Hongtian; Yang, Zhijun; Yang, Yang; Zhu, Jianwei; Xu, Minhui; Xu, Ruxiang

    2016-01-01

    Along with the development of stem cell-based therapies for central nervous system (CNS) disease, the safety of stem cell grafts in the CNS, such as induced pluripotent stem cells (iPSCs) and induced neural stem cells (iNSCs), should be of primary concern. To provide scientific basis for evaluating the safety of these stem cells, we determined their tumourigenicity and immunogenicity in syngeneic mouse brain. Both iPSCs and embryonic stem cells (ESCs) were able to form tumours in the mouse brain, leading to tissue destruction along with immune cell infiltration. In contrast, no evidence of tumour formation, brain injury or immune rejection was observed with iNSCs, neural stem cells (NSCs) or mesenchymal stem cells (MSCs). With the help of gene ontology (GO) enrichment analysis, we detected significantly elevated levels of chemokines in the brain tissue and serum of mice that developed tumours after ESC or iPSC transplantation. Moreover, we also investigated the interactions between chemokines and NF-κB signalling and found that NF-κB activation was positively correlated with the constantly rising levels of chemokines, and vice versa. In short, iNSC grafts, which lacked any resulting tumourigenicity or immunogenicity, are safer than iPSC grafts. PMID:27417157

  15. Salvinorin A Pretreatment Preserves Cerebrovascular Autoregulation After Brain Hypoxic/Ischemic Injury via Extracellular Signal-Regulated Kinase / Mitogen-Activated Protein Kinase in Piglets

    PubMed Central

    Su, Diansan; Riley, John; Armstead, William M.; Liu, Renyu

    2012-01-01

    Background Cerebral hypoxia/ischemia during infant congenital heart surgery is not uncommon, and may induce devastating neurologic disabilities persistent over the lifespan. Hypoxia/ischemia-induced cerebrovascular dysfunction is thought to be an important contributor to neurological damage. No pharmacological agents have been found to prevent this. Mitogen activated protein kinase (MAPK), including extracellular signal regulated kinase (ERK), c-Jun-N-terminal kinase (JNK) and p38, is thought to contribute to ischemic preconditioning. We investigated whether pretreatment with salvinorin A, the only natural non-opioid kappa receptor agonist, could preserve autoregulation of the pial artery via MAPK. Methods The response of the pial artery to hypotension and hypercapnia was monitored in piglets equipped with a closed cranial window before and after hypoxia and ischemia in the presence or absence of U0126, an inhibitor for the protein kinase upstream of ERK, sp600125, an inhibitor of c-JNK or sb203580, an inhibitor of p38. Salvinorin A (10 μg/kg IV) was administered 30 minutes before hypoxia/ischemia in salvinorin-treated animals. Cerebrospinal fluid samples were collected before and 30 minutes after salvinorin A administration for the measurement of MAPK. Data (n=5) were analyzed by repeated-measures analysis of variance. Results Pial artery dilation to hypercapnia and hypotension was blunted after hypoxia/ischemia, but preserved well by pretreatment with salvinorin A. U0126, but not sp600125 or sb203580, abolished the preservative effects of salvinorin A on cerebral vascular autoregulation to hypotension and hypercapnia. The ratio of pERK/ERK in cerebrospinal fluid increased significantly in salvinorin-treated animals, which was inhibited by U0126. Conclusions Salvinorin A pretreatment preserves autoregulation of the pial artery to hypotension and hypercapnia after hypoxia/ischemia via ERK in a piglet model. PMID:22075021

  16. Inflammatory infiltrates and natural killer cell presence in human brain tumors.

    PubMed

    Stevens, A; Klöter, I; Roggendorf, W

    1988-02-15

    Immunohistochemical analysis of subpopulations of inflammatory cells in 81 primary and secondary human brain tumors was done. Natural killer (NK) cells, representing non-major histocompatibility complex-restricted, spontaneous cytotoxicity and monocytic cells are virtually absent in infiltrates of gliomas and account only for a minor percentage of inflammatory cells in brain metastases of carcinoma and in craniopharyngeomas. Infiltrates in gliomas consist almost exclusively of T-cells of the suppressor/cytotoxic type whereas infiltrates in carcinoma metastases and craniopharyngeomas contain considerable numbers of T-helper/inducer cells and B-cells. From this the authors conclude (1) that NK cells do not play a major role in tumor rejection, and (2) that the kind of inflammatory reaction does not depend upon the tumor site but more likely on the tumor type. No correlation between tumor differentiation and infiltrate composition is evident.

  17. Selection of brain metastasis-initiating breast cancer cells determined by growth on hard agar.

    PubMed

    Guo, Lixia; Fan, Dominic; Zhang, Fahao; Price, Janet E; Lee, Ju-Seog; Marchetti, Dario; Fidler, Isaiah J; Langley, Robert R

    2011-05-01

    An approach that facilitates rapid isolation and characterization of tumor cells with enhanced metastatic potential is highly desirable. Here, we demonstrate that plating GI-101A human breast cancer cells on hard (0.9%) agar selects for the subpopulation of metastasis-initiating cells. The agar-selected cells, designated GI-AGR, were homogeneous for CD44(+) and CD133(+) and five times more invasive than the parental GI-101A cells. Moreover, mice injected with GI-AGR cells had significantly more experimental brain metastases and shorter overall survival than did mice injected with GI-101A cells. Comparative gene expression analysis revealed that GI-AGR cells were markedly distinct from the parental cells but shared an overlapping pattern of gene expression with the GI-101A subline GI-BRN, which was generated by repeated in vivo recycling of GI-101A cells in an experimental brain metastasis model. Data mining on 216 genes shared between GI-AGR and GI-BRN breast cancer cells suggested that the molecular phenotype of these cells is consistent with that of cancer stem cells and the aggressive basal subtype of breast cancer. Collectively, these results demonstrate that analysis of cell growth in a hard agar assay is a powerful tool for selecting metastasis-initiating cells in a heterogeneous population of breast cancer cells, and that such selected cells have properties similar to those of tumor cells that are selected based on their potential to form metastases in mice.

  18. MIF Maintains the Tumorigenic Capacity of Brain Tumor-Initiating Cells by Directly Inhibiting p53.

    PubMed

    Fukaya, Raita; Ohta, Shigeki; Yaguchi, Tomonori; Matsuzaki, Yumi; Sugihara, Eiji; Okano, Hideyuki; Saya, Hideyuki; Kawakami, Yutaka; Kawase, Takeshi; Yoshida, Kazunari; Toda, Masahiro

    2016-05-01

    Tumor-initiating cells thought to drive brain cancer are embedded in a complex heterogeneous histology. In this study, we isolated primary cells from 21 human brain tumor specimens to establish cell lines with high tumorigenic potential and to identify the molecules enabling this capability. The morphology, sphere-forming ability upon expansion, and differentiation potential of all cell lines were indistinguishable in vitro However, testing for tumorigenicity revealed two distinct cell types, brain tumor-initiating cells (BTIC) and non-BTIC. We found that macrophage migration inhibitory factor (MIF) was highly expressed in BTIC compared with non-BTIC. MIF bound directly to both wild-type and mutant p53 but regulated p53-dependent cell growth by different mechanisms, depending on glioma cell line and p53 status. MIF physically interacted with wild-type p53 in the nucleus and inhibited its transcription-dependent functions. In contrast, MIF bound to mutant p53 in the cytoplasm and abrogated transcription-independent induction of apoptosis. Furthermore, MIF knockdown inhibited BTIC-induced tumor formation in a mouse xenograft model, leading to increased overall survival. Collectively, our findings suggest that MIF regulates BTIC function through direct, intracellular inhibition of p53, shedding light on the molecular mechanisms underlying the tumorigenicity of certain malignant brain cells. Cancer Res; 76(9); 2813-23. ©2016 AACR.

  19. DRP1 inhibition rescues retinal ganglion cells and their axons by preserving mitochondrial integrity in a mouse model of glaucoma

    PubMed Central

    Kim, K-Y; Perkins, G A; Shim, M S; Bushong, E; Alcasid, N; Ju, S; Ellisman, M H; Weinreb, R N; Ju, W-K

    2015-01-01

    Glaucoma is the leading cause of irreversible blindness and is characterized by slow and progressive degeneration of the optic nerve head axons and retinal ganglion cell (RGC), leading to loss of visual function. Although oxidative stress and/or alteration of mitochondrial (mt) dynamics induced by elevated intraocular pressure (IOP) are associated with this neurodegenerative disease, the mechanisms that regulate mt dysfunction-mediated glaucomatous neurodegeneration are poorly understood. Using a mouse model of glaucoma, DBA/2J (D2), which spontaneously develops elevated IOP, as well as an in vitro RGC culture system, we show here that oxidative stress, as evidenced by increasing superoxide dismutase 2 (SOD2) and mt transcription factor A (Tfam) protein expression, triggers mt fission and loss by increasing dynamin-related protein 1 (DRP1) in the retina of glaucomatous D2 mice as well as in cultured RGCs exposed to elevated hydrostatic pressure in vitro. DRP1 inhibition by overexpressing DRP1 K38A mutant blocks mt fission and triggers a subsequent reduction of oxidative stress, as evidenced by decreasing SOD2 and Tfam protein expression. DRP1 inhibition promotes RGC survival by increasing phosphorylation of Bad at serine 112 in the retina and preserves RGC axons by maintaining mt integrity in the glial lamina of glaucomatous D2 mice. These findings demonstrate an important vicious cycle involved in glaucomatous neurodegeneration that starts with elevated IOP producing oxidative stress; the oxidative stress then leads to mt fission and a specific form of mt dysfunction that generates further oxidative stress, thus perpetuating the cycle. Our findings suggest that DRP1 is a potential therapeutic target for ameliorating oxidative stress-mediated mt fission and dysfunction in RGC and its axons during glaucomatous neurodegeneration. Thus, DRP1 inhibition may provide a new therapeutic strategy for protecting both RGCs and their axons in glaucoma and other optic

  20. Brain tumor - primary - adults

    MedlinePlus

    ... Vestibular schwannoma (acoustic neuroma) - adults; Meningioma - adults; Cancer - brain tumor (adults) ... Primary brain tumors include any tumor that starts in the brain. Primary brain tumors can start from brain cells, ...

  1. Benzyl isothiocyanate alters the gene expression with cell cycle regulation and cell death in human brain glioblastoma GBM 8401 cells.

    PubMed

    Tang, Nou-Ying; Chueh, Fu-Shin; Yu, Chien-Chih; Liao, Ching-Lung; Lin, Jen-Jyh; Hsia, Te-Chun; Wu, King-Chuen; Liu, Hsin-Chung; Lu, Kung-Wen; Chung, Jing-Gung

    2016-04-01

    Glioblastoma multiforme (GBM) is a highly malignant devastating brain tumor in adults. Benzyl isothiocyanate (BITC) is one of the isothiocyanates that have been shown to induce human cancer cell apoptosis and cell cycle arrest. Herein, the effect of BITC on cell viability and apoptotic cell death and the genetic levels of human brain glioblastoma GBM 8401 cells in vitro were investigated. We found that BITC induced cell morphological changes, decreased cell viability and the induction of cell apoptosis in GBM 8401 cells was time-dependent. cDNA microarray was used to examine the effects of BITC on GBM 8401 cells and we found that numerous genes associated with cell death and cell cycle regulation in GBM 8401 cells were altered after BITC treatment. The results show that expression of 317 genes was upregulated, and two genes were associated with DNA damage, the DNA-damage-inducible transcript 3 (DDIT3) was increased 3.66-fold and the growth arrest and DNA-damage-inducible α (GADD45A) was increased 2.34-fold. We also found that expression of 182 genes was downregulated and two genes were associated with receptor for cell responses to stimuli, the EGF containing fibulin-like extracellular matrix protein 1 (EFEMP1) was inhibited 2.01-fold and the TNF receptor-associated protein 1 (TRAP1) was inhibited 2.08-fold. BITC inhibited seven mitochondria ribosomal genes, the mitochondrial ribosomal protein; tumor protein D52 (MRPS28) was inhibited 2.06-fold, the mitochondria ribosomal protein S2 (MRPS2) decreased 2.07-fold, the mitochondria ribosomal protein L23 (MRPL23) decreased 2.08-fold, the mitochondria ribosomal protein S2 (MRPS2) decreased 2.07-fold, the mitochondria ribosomal protein S12 (MRPS12) decreased 2.08-fold, the mitochondria ribosomal protein L12 (MRPL12) decreased 2.25-fold and the mitochondria ribosomal protein S34 (MRPS34) was decreased 2.30-fold in GBM 8401 cells. These changes of gene expression can provide the effects of BITC on the genetic level and are

  2. [Determination of the electrophoretic mobility of peripheral red blood cells from healthy individuals, by using ethylene-2-amine-4-chloroacetic acid as a preservative].

    PubMed

    Iastrubinetskaia, O I; Popova, O V; Sarycheva, T G; Tentsova, I A; Kozinets, G I

    2009-01-01

    It is now known that the electric charge of a surface cellular membrane is a rather characteristic value for each type of a cell. Cellular electrophoresis may be used to study the electrokinetic properties of red blood cells in each specific case, thereby obtaining indirect information on the state of a surface membrane as the electrophoretic mobility (EPM) rate is directly proportional to the value of a cellular electric charge. The study was undertaken to compare the EPM rates of peripheral red blood cells from donors, by employing two different preservatives: EDTCHA and sodium citrate. No statistically significant differences were found in the groups under study and these parameters were comparable.

  3. The impairment of hepatocytes and sinusoidal endothelial cells during cold preservation in rat fatty liver induced by alcohol and the beneficial effect of hepatocyte growth factor.

    PubMed

    Takeda, Yoshihisa; Arii, Shigeki; Kaido, Toshimi; Imamura, Masayuki

    2003-04-01

    A fatty liver resulting from alcohol intake is often unattractive for grafting. In this study, we investigated the impairment of hepatocytes and sinusoidal endothelial cells (SECs) during cold preservation of alcohol-induced fatty liver and examined the efficacy of human recombinant hepatocyte growth factor (hrHGF). Rats were fed an alcohol diet. We performed histological examinations of the hepatocytes and observed the ultrastructural alteration of the SECs. Additionally, we measured hepatic transaminase and peroxidative lipids for hepatocellular injury and the hyaluronic acid uptake rate (HUR) to determine SEC injury. We added hrHGF to University of Wisconsin (UW) solution to assess the protective effect of the agent. Numerous fatty deposits were observed in ethanol-induced fatty livers. These grew with the duration of cold storage. Hepatic transaminases of the effluents increased during cold preservation in the livers of alcohol-treated rats. Additionally, peroxidative lipids in the effluents increased during cold preservation in the livers of alcohol-treated rats, whereas they were undetectable in non-alcohol-treated rat livers. The sinusoidal endothelium had severely deteriorated in the livers of alcohol-treated rats. Further, the HUR decreased with ethanol treatment and/or cold preservation. The addition of hrHGF suppressed the increase of hepatic transaminase in the effluent of cold-preserved alcohol-treated livers. Peroxidative lipids in the same effluents were undetectable. In fatty livers, both hepatocytes and SECs received severe damage during cold preservation. Furthermore, we demonstrated that hepatocellular injury was significantly inhibited by hrHGF.

  4. Delivery of Small Interfering RNA to Inhibit Vascular Endothelial Growth Factor in Zebrafish Using Natural Brain Endothelia Cell-Secreted Exosome Nanovesicles for the Treatment of Brain Cancer.

    PubMed

    Yang, Tianzhi; Fogarty, Brittany; LaForge, Bret; Aziz, Salma; Pham, Thuy; Lai, Leanne; Bai, Shuhua

    2017-03-01

    Although small interfering RNA (siRNA) holds great therapeutic promise, its delivery to the disease site remains a paramount obstacle. In this study, we tested whether brain endothelial cell-derived exosomes could deliver siRNA across the blood-brain barrier (BBB) in zebrafish. Natural exosomes were isolated from brain endothelial bEND.3 cell culture media and vascular endothelial growth factor (VEGF) siRNA was loaded in exosomes with the assistance of a transfection reagent. While fluorescence-activated cell flow cytometry and immunocytochemistry staining studies indicated that wild-type exosomes significantly increased the uptake of fluorescence-labeled siRNA in the autologous brain endothelial cells, decreased fluorescence intensity was observed in the cells treated with the tetraspanin CD63 antibody-blocked exosome-delivered formulation (p < 0.05). In the transport study, exosomes also enhanced the permeability of rhodamine 123 in a co-cultured monolayer of brain endothelial bEND.3 cell and astrocyte. Inhibition at the expression of VEGF RNA and protein levels was observed in glioblastoma-astrocytoma U-87 MG cells treated with exosome-delivered siRNAs. Imaging results showed that exosome delivered more siRNAs across the BBB in Tg(fli1:GFP) zebrafish. In a xenotransplanted brain tumor model, exosome-delivered VEGF siRNAs decreased the fluorescence intensity of labeled cancer cells in the brain of zebrafish. Brain endothelial cell-derived exosomes could be potentially used as a natural carrier for the brain delivery of exogenous siRNA.

  5. Involvement of the cellular prion protein in the migration of brain microvascular endothelial cells.

    PubMed

    Watanabe, Takuya; Yasutaka, Yuki; Nishioku, Tsuyoshi; Kusakabe, Sae; Futagami, Koujiro; Yamauchi, Atsushi; Kataoka, Yasufumi

    2011-06-01

    The conversion of cellular prion protein (PrP(C)) to its protease-resistant isoform is involved in the pathogenesis of prion disease. Although PrP(C) is a ubiquitous glycoprotein that is present in various cell types, the physiological role of PrP(C) remains obscure. The present study aimed to determine whether PrP(C) mediates migration of brain microvascular endothelial cells. Small interfering RNAs (siRNAs) targeting PrP(C) were transfected into a mouse brain microvascular endothelial cell line (bEND.3 cells). siPrP1, selected among three siRNAs, reduced mRNA and protein levels of PrP(C) in bEND.3 cells. Cellular migration was evaluated with a scratch-wound assay. siPrP1 suppressed migration without significantly affecting cellular proliferation. This study provides the first evidence that PrP(C) may be necessary for brain microvascular endothelial cells to migrate into damaged regions in the brain. This function of PrP(C) in the brain endothelium may be a mechanism by which the neurovascular unit recovers from an injury such as an ischemic insult.

  6. Control of adult neurogenesis by programmed cell death in the mammalian brain.

    PubMed

    Ryu, Jae Ryun; Hong, Caroline Jeeyeon; Kim, Joo Yeon; Kim, Eun-Kyoung; Sun, Woong; Yu, Seong-Woon

    2016-04-21

    The presence of neural stem cells (NSCs) and the production of new neurons in the adult brain have received great attention from scientists and the public because of implications to brain plasticity and their potential use for treating currently incurable brain diseases. Adult neurogenesis is controlled at multiple levels, including proliferation, differentiation, migration, and programmed cell death (PCD). Among these, PCD is the last and most prominent process for regulating the final number of mature neurons integrated into neural circuits. PCD can be classified into apoptosis, necrosis, and autophagic cell death and emerging evidence suggests that all three may be important modes of cell death in neural stem/progenitor cells. However, the molecular mechanisms that regulate PCD and thereby impact the intricate balance between self-renewal, proliferation, and differentiation during adult neurogenesis are not well understood. In this comprehensive review, we focus on the extent, mechanism, and biological significance of PCD for the control of adult neurogenesis in the mammalian brain. The role of intrinsic and extrinsic factors in the regulation of PCD at the molecular and systems levels is also discussed. Adult neurogenesis is a dynamic process, and the signals for differentiation, proliferation, and death of neural progenitor/stem cells are closely interrelated. A better understanding of how adult neurogenesis is influenced by PCD will help lead to important insights relevant to brain health and diseases.

  7. MANF silencing, immunity induction or autophagy trigger an unusual cell type in metamorphosing Drosophila brain.

    PubMed

    Stratoulias, Vassilis; Heino, Tapio I

    2015-05-01

    Glia are abundant cells in the brain of animals ranging from flies to humans. They perform conserved functions not only in neural development and wiring, but also in brain homeostasis. Here we show that by manipulating gene expression in glia, a previously unidentified cell type appears in the Drosophila brain during metamorphosis. More specifically, this cell type appears in three contexts: (1) after the induction of either immunity, or (2) autophagy, or (3) by silencing of neurotrophic factor DmMANF in glial cells. We call these cells MANF immunoreactive Cells (MiCs). MiCs are migratory based on their shape, appearance in brain areas where no cell bodies exist and the nuclear localization of dSTAT. They are labeled with a unique set of molecular markers including the conserved neurotrophic factor DmMANF and the transcription factor Zfh1. They possess the nuclearly localized protein Relish, which is the hallmark of immune response activation. They also express the conserved engulfment receptor Draper, therefore indicating that they are potentially phagocytic. Surprisingly, they do not express any of the common glial and neuronal markers. In addition, ultrastructural studies show that MiCs are extremely rich in lysosomes. Our findings reveal critical molecular and functional components of an unusual cell type in the Drosophila brain. We suggest that MiCs resemble macrophages/hemocytes and vertebrate microglia based on their appearance in the brain upon genetically challenged conditions and the expression of molecular markers. Interestingly, macrophages/hemocytes or microglia-like cells have not been reported in the fly nervous system before.

  8. Histamine from brain resident MAST cells promotes wakefulness and modulates behavioral states.

    PubMed

    Chikahisa, Sachiko; Kodama, Tohru; Soya, Atsushi; Sagawa, Yohei; Ishimaru, Yuji; Séi, Hiroyoshi; Nishino, Seiji

    2013-01-01

    Mast cell activation and degranulation can result in the release of various chemical mediators, such as histamine and cytokines, which significantly affect sleep. Mast cells also exist in the central nervous system (CNS). Since up to 50% of histamine contents in the brain are from brain mast cells, mediators from brain mast cells may significantly influence sleep and other behaviors. In this study, we examined potential involvement of brain mast cells in sleep/wake regulations, focusing especially on the histaminergic system, using mast cell deficient (W/W(v)) mice. No significant difference was found in the basal amount of sleep/wake between W/W(v) mice and their wild-type littermates (WT), although W/W(v) mice showed increased EEG delta power and attenuated rebound response after sleep deprivation. Intracerebroventricular injection of compound 48/80, a histamine releaser from mast cells, significantly increased histamine levels in the ventricular region and enhanced wakefulness in WT mice, while it had no effect in W/W(v) mice. Injection of H1 antagonists (triprolidine and mepyramine) significantly increased the amounts of slow-wave sleep in WT mice, but not in W/W(v) mice. Most strikingly, the food-seeking behavior observed in WT mice during food deprivation was completely abolished in W/W(v) mice. W/W(v) mice also exhibited higher anxiety and depression levels compared to WT mice. Our findings suggest that histamine released from brain mast cells is wake-promoting, and emphasizes the physiological and pharmacological importance of brain mast cells in the regulation of sleep and fundamental neurobehavior.

  9. Emerging Insights for Better Delivery of Chemicals and Stem Cells to the Brain.

    PubMed

    Baig, Abdul Mannan

    2017-04-03

    The human central nervous system and its vascularity has evolved a complex yet useful barricade, called the blood-brain barrier. The understanding of this barrier and the transport proteins that allow a selective access across it remains fundamental in designing chemicals and molecules that could cross it and prove therapeutically beneficial. After the recent findings of the brain-lymphatic link and the movement of chemical molecules from the central nervous system into general circulation, the model of the blood-brain barrier needs to be revisited. Here the author intends to discuss the components of the blood-brain barrier, debate the novel routes of administration and delivery systems for the drugs and embryonic stem cells that could cross or bypass the blood-brain barrier in order to reach the central nervous system.

  10. Transplantation of marrow stromal cells restores cerebral blood flow and reduces cerebral atrophy in rats with traumatic brain injury: in vivo MRI study.

    PubMed

    Li, Lian; Jiang, Quan; Qu, Chang Sheng; Ding, Guang Liang; Li, Qing Jiang; Wang, Shi Yang; Lee, Ji Hyun; Lu, Mei; Mahmood, Asim; Chopp, Michael

    2011-04-01

    Cell therapy promotes brain remodeling and improves functional recovery after various central nervous system disorders, including traumatic brain injury (TBI). We tested the hypothesis that treatment of TBI with intravenous administration of human marrow stromal cells (hMSCs) provides therapeutic benefit in modifying hemodynamic and structural abnormalities, which are detectable by in vivo MRI. hMSCs were labeled with superparamagnetic iron oxide (SPIO) nanoparticles. Male Wistar rats (300-350 g, n=18) subjected to controlled cortical impact TBI were intravenously injected with 1 mL of saline (n=9) or hMSCs in suspension (n=9, approximately 3 × 10(6) SPIO-labeled hMSCs) 5 days post-TBI. In vivo MRI measurements consisting of cerebral blood flow (CBF), T2-weighted imaging, and 3D gradient echo imaging were performed for all animals 2 days post-TBI and weekly for 6 weeks. Functional outcome was evaluated with modified neurological severity score and Morris water maze test. Cell engraftment was detected in vivo by 3D MRI and confirmed by double staining. Ventricle and lesion volumetric alterations were measured using T2 maps, and hemodynamic abnormality was tracked by MRI CBF measurements. Our data demonstrate that treatment with hMSCs following TBI diminishes hemodynamic abnormalities by early restoration and preservation of CBF in the brain regions adjacent to and remote from the impact site, and reduces generalized cerebral atrophy, all of which may contribute to the observed improvement of functional outcome.

  11. Brain Tumor Cells in Circulation are Enriched for Mesenchymal Gene Expression

    PubMed Central

    Sullivan, James P.; Nahed, Brian V.; Madden, Marissa W.; Oliveira, Samantha M.; Springer, Simeon; Bhere, Deepak; Chi, Andrew S.; Wakimoto, Hiroaki; Rothenberg, S. Michael; Sequist, Lecia V.; Kapur, Ravi; Shah, Khalid; Iafrate, A. John; Curry, William T.; Loeffler, Jay S.; Batchelor, Tracy T.; Louis, David N.; Toner, Mehmet; Maheswaran, Shyamala; Haber, Daniel A.

    2014-01-01

    Glioblastoma (GBM) is a highly aggressive brain cancer characterized by local invasion and angiogenic recruitment, yet metastatic dissemination is extremely rare. Here, we adapted a microfluidic device to deplete hematopoietic cells from blood specimens of patients with GBM, uncovering evidence of circulating brain tumor cells (CTCs). Staining and scoring criteria for GBM CTCs were first established using orthotopic patient-derived xenografts (PDX), and then applied clinically: CTCs were identified in at least one blood specimen from 13/33 patients (39%; 26/87 samples). Single GBM CTCs isolated from both patients and mouse PDX models demonstrated enrichment for mesenchymal over neural differentiation markers, compared with primary GBMs. Within primary GBMs, RNA-in-situ hybridization identifies a subpopulation of highly migratory mesenchymal tumor cells, and in a rare patient with disseminated GBM, systemic lesions were exclusively mesenchymal. Thus, a mesenchymal subset of GBM cells invades into the vasculature, and may proliferate outside the brain. PMID:25139148

  12. Taip2 is a novel cell death-related gene expressed in the brain during development

    SciTech Connect

    Yamada, Kazumi; Akiyama, Nobutake; Yamada, Shuichi; Tanaka, Hiromitsu; Saito, Saburo; Hiraoka, Masahiro; Kizaka-Kondoh, Shinae

    2008-05-02

    TAIP2 was isolated as one of the homologous genes of TAIP3 (TGF-{beta}-up-regulated apoptosis-inducing-protein chromosome 3). The transcript of the mouse counterpart of TAIP2, designated mTaip2, was detected in several tissue specimens from embryos to adults, while mTaip2 was dominantly expressed in the embryonic brain. The overexpression of the full-length mTaip2 induced cell death in various cell lines. An analysis of mTaip2 deletion mutants revealed that the N-terminal half of mTaip2, but not the C-terminal half, had nuclear localization and cell death-inducing activities. The results indicate that mTaip2 is a novel cell death-related gene dominantly expressed in the embryonic brain, thus suggesting that mTaip2 may play a role in development of the brain.

  13. Control of the blood-brain barrier function in cancer cell metastasis.

    PubMed

    Blecharz, Kinga G; Colla, Ruben; Rohde, Veit; Vajkoczy, Peter

    2015-10-01

    Cerebral metastases are the most common brain neoplasms seen clinically in the adults and comprise more than half of all brain tumours. Actual treatment options for brain metastases that include surgical resection, radiotherapy and chemotherapy are rarely curative, although palliative treatment improves survival and life quality of patients carrying brain-metastatic tumours. Chemotherapy in particular has also shown limited or no activity in brain metastasis of most tumour types. Many chemotherapeutic agents used systemically do not cross the blood-brain barrier (BBB), whereas others may transiently weaken the BBB and allow extravasation of tumour cells from the circulation into the brain parenchyma. Increasing evidence points out that the interaction between the BBB and tumour cells plays a key role for implantation and growth of brain metastases in the central nervous system. The BBB, as the tightest endothelial barrier, prevents both early detection and treatment by creating a privileged microenvironment. Therefore, as observed in several in vivo studies, precise targetting the BBB by a specific transient opening of the structure making it permeable for therapeutic compounds, might potentially help to overcome this difficult clinical problem. Moreover, a better understanding of the molecular features of the BBB, its interrelation with metastatic tumour cells and the elucidation of cellular mechanisms responsible for establishing cerebral metastasis must be clearly outlined in order to promote treatment modalities that particularly involve chemotherapy. This in turn would substantially expand the survival and quality of life of patients with brain metastasis, and potentially increase the remission rate. Therefore, the focus of this review is to summarise the current knowledge on the role and function of the BBB in cancer metastasis.

  14. Neural stem cell-based dual suicide gene delivery for metastatic brain tumors.

    PubMed

    Wang, C; Natsume, A; Lee, H J; Motomura, K; Nishimira, Y; Ohno, M; Ito, M; Kinjo, S; Momota, H; Iwami, K; Ohka, F; Wakabayashi, T; Kim, S U

    2012-11-01

    In our previous works, we demonstrated that human neural stem cells (NSCs) transduced with the cytosine deaminase (CD) gene showed remarkable 'bystander killer effect' on glioma and medulloblastoma cells after administration of the prodrug 5-fluorocytosine (5-FC). In addition, herpes simplex virus thymidine kinase (TK) is a widely studied enzyme used for suicide gene strategies, for which the prodrug is ganciclovir (GCV). To apply this strategy to brain metastasis treatment, we established here a human NSC line (F3.CD-TK) expressing the dual suicide genes CD and TK. We examined whether F3.CD-TK cells intensified the antitumor effect on lung cancer brain metastases. In vitro studies showed that F3.CD-TK cells exerted a marked bystander effect on human lung cancer cells after treatment with 5-FC and GCV. In a novel experimental brain metastases model, intravenously administered F3 cells migrated near lung cancer metastatic lesions, which were induced by the injection of lung cancer cells via the intracarotid artery. More importantly, F3.CD-TK cells in the presence of prodrugs 5-FC and GCV decreased tumor size and considerably prolonged animal survival. The results of the present study indicate that the dual suicide gene-engineered, NSC-based treatment strategy might offer a new promising therapeutic modality for brain metastases.

  15. Selective capture of endothelial and perivascular cells from brain microvessels using laser capture microdissection.

    PubMed

    Kinnecom, Katie; Pachter, Joel S

    2005-12-01

    Laser capture microdissection (LCM) of the major cell types comprising brain microvessels offers a powerful technology to explore the molecular basis of the blood-brain barrier in health and disease. However, the ability to selectively retrieve endothelial or perivascular cells, without cross-contamination from the other, has proven difficult. Additionally, histochemical methods previously described for use with LCM have not allowed for identification of all the different size branches of the microvascular tree. Here, we describe a double immunostaining method, combining bright-field and fluorescence microscopy, and using an extensive dehydration with xylene, to clearly identify and spatially resolve endothelial from perivascular cells within all size microvascular branches in frozen brain sections. LCM of these sections, coupled with RNA analysis by reverse-transcription polymerase chain reaction, revealed that captured endothelial cells show endothelial markers but no detectable markers for astrocytes or smooth muscle cells/pericytes. Conversely, captured astrocytes or smooth muscle cells/pericytes demonstrate their respective markers, but not those of endothelial cells. This approach has applicability to microarray analysis, thereby enabling global gene profiling of the different cell types along the entirety of the brain microvascular tree.

  16. Ribosome Profiling Reveals a Cell-Type-Specific Translational Landscape in Brain Tumors

    PubMed Central

    Gonzalez, Christian; Sims, Jennifer S.; Hornstein, Nicholas; Mela, Angeliki; Garcia, Franklin; Lei, Liang; Gass, David A.; Amendolara, Benjamin; Bruce, Jeffrey N.

    2014-01-01

    Glioma growth is driven by signaling that ultimately regulates protein synthesis. Gliomas are also complex at the cellular level and involve multiple cell types, including transformed and reactive cells in the brain tumor microenvironment. The distinct functions of the various cell types likely lead to different requirements and regulatory paradigms for protein synthesis. Proneural gliomas can arise from transformation of glial progenitors that are driven to proliferate via mitogenic signaling that affects translation. To investigate translational regulation in this system, we developed a RiboTag glioma mouse model that enables cell-type-specific, genome-wide ribosome profiling of tumor tissue. Infecting glial progenitors with Cre-recombinant retrovirus simultaneously activates expression of tagged ribosomes and delivers a tumor-initiating mutation. Remarkably, we find that although genes specific to transformed cells are highly translated, their translation efficiencies are low compared with normal brain. Ribosome positioning reveals sequence-dependent regulation of ribosomal activity in 5′-leaders upstream of annotated start codons, leading to differential translation in glioma compared with normal brain. Additionally, although transformed cells express a proneural signature, untransformed tumor-associated cells, including reactive astrocytes and microglia, express a mesenchymal signature. Finally, we observe the same phenomena in human disease by combining ribosome profiling of human proneural tumor and non-neoplastic brain tissue with computational deconvolution to assess cell-type-specific translational regulation. PMID:25122893

  17. Neural stem cells improve neuronal survival in cultured postmortem brain tissue from aged and Alzheimer patients

    PubMed Central

    Wu, L; Sluiter, A A; Guo, Ho-fu; Balesar, R A; Swaab, D F; Zhou, Jiang-Ning; Verwer, R W H

    2008-01-01

    Abstract Neurodegenerative diseases are progressive and incurable and are becoming ever more prevalent. To study whether neural stem cell can reactivate or rescue functions of impaired neurons in the human aging and neurodegenerating brain, we co-cultured postmortem slices from Alzheimer patients and control participants with rat embryonic day 14 (E14) neural stem cells. Viability staining based on the exclusion of ethidium bromide by intact plasma membranes showed that there were strikingly more viable cells and fewer dead cells in slices co-cultured with neural stem cells than in untreated slices. The presence of Alzheimer pathology in the brain slices did not influence this effect, although the slices from Alzheimer patients, in general, contained fewer viable cells. Co-culturing with rat E14 fibroblasts did not improve the viability of neurons in the human brain slices. Since the human slices and neural stem cells were separated by a membrane during co-culturing our data show for the first time that neural stem cells release diffusible factors that may improve the survival of aged and degenerating neurons in human brains. PMID:18088384

  18. Neuro-immune Dysfunction During Brain Aging: New Insights in Microglial Cell Regulation

    PubMed Central

    Matt, Stephanie M.; Johnson, Rodney W.

    2015-01-01

    Microglia, the resident immune cells of the brain, are at the center of communication between the central nervous system and immune system. While these brain-immune interactions are balanced in healthy adulthood, the ability to maintain homeostasis during aging is impaired. Microglia develop a loss of integrated regulatory networks including aberrant signaling from other brain cells, immune sensors, and epigenetic modifiers. The low-grade chronic neuroinflammation associated with this dysfunctional activity likely contributes to cognitive deficits and susceptibility to age-related pathologies. A better understanding of the underlying mechanisms responsible for neuro-immune dysregulation with age is crucial for providing targeted therapeutic strategies to support brain repair and healthy aging. PMID:26595306

  19. Neurodevelopment. Live imaging of adult neural stem cell behavior in the intact and injured zebrafish brain.

    PubMed

    Barbosa, Joana S; Sanchez-Gonzalez, Rosario; Di Giaimo, Rossella; Baumgart, Emily Violette; Theis, Fabian J; Götz, Magdalena; Ninkovic, Jovica

    2015-05-15

    Adult neural stem cells are the source for restoring injured brain tissue. We used repetitive imaging to follow single stem cells in the intact and injured adult zebrafish telencephalon in vivo and found that neurons are generated by both direct conversions of stem cells into postmitotic neurons and via intermediate progenitors amplifying the neuronal output. We observed an imbalance of direct conversion consuming the stem cells and asymmetric and symmetric self-renewing divisions, leading to depletion of stem cells over time. After brain injury, neuronal progenitors are recruited to the injury site. These progenitors are generated by symmetric divisions that deplete the pool of stem cells, a mode of neurogenesis absent in the intact telencephalon. Our analysis revealed changes in the behavior of stem cells underlying generation of additional neurons during regeneration.

  20. Environmental complexity, seasonality and brain cell proliferation in a weakly electric fish, Brachyhypopomus gauderio

    PubMed Central

    Dunlap, Kent D.; Silva, Ana C.; Chung, Michael

    2011-01-01

    Environmental complexity and season both influence brain cell proliferation in adult vertebrates, but their relative importance and interaction have not been directly assessed. We examined brain cell proliferation during both the breeding and non-breeding seasons in adult male electric fish, Brachyhypopomus gauderio, exposed to three environments that differed in complexity: (1) a complex natural habitat in northern Uruguay, (2) an enriched captive environment where fish were housed socially and (3) a simple laboratory setting where fish were isolated. We injected fish with BrdU 2.5 h before sacrifice to label newborn cells. We examined the hindbrain and midbrain and quantified the density of BrdU+ cells in whole transverse sections, proliferative zones and two brain nuclei in the electrocommunication circuitry (the pacemaker nucleus and the electrosensory lateral line lobe). Season had the largest effect on cell proliferation, with fish during the breeding season having three to seven times more BrdU+ cells than those during the non-breeding season. Although the effect was smaller, fish from a natural environment had greater rates of cell proliferation than fish in social or isolated captive environments. For most brain regions, fish in social and isolated captive environments had equivalent levels of cell proliferation. However, for brain regions in the electrocommunication circuitry, group-housed fish had more cell proliferation than isolated fish, but only during the breeding season (season × environment interaction). The regionally and seasonally specific effect of social environment on cell proliferation suggests that addition of new cells to these nuclei may contribute to seasonal changes in electrocommunication behavior. PMID:21307066

  1. Cooling treatment transiently increases the permeability of brain capillary endothelial cells through translocation of claudin-5.

    PubMed

    Inamura, Akinori; Adachi, Yasuhiro; Inoue, Takao; He, Yeting; Tokuda, Nobuko; Nawata, Takashi; Shirao, Satoshi; Nomura, Sadahiro; Fujii, Masami; Ikeda, Eiji; Owada, Yuji; Suzuki, Michiyasu

    2013-08-01

    The blood-brain-barrier (BBB) is formed by different cell types, of which brain microvascular endothelial cells are major structural constituents. The goal of this study was to examine the effects of cooling on the permeability of the BBB with reference to tight junction formation of brain microendothelial cells. The sensorimotor cortex above the dura mater in adult male Wistar rats was focally cooled to a temperature of 5 °C for 1 h, then immunostaining for immunoglobulin G (IgG) was performed to evaluate the permeability of the BBB. Permeability produced by cooling was also evaluated in cultured murine brain endothelial cells (bEnd3) based on measurement of trans-epithelial electric resistance (TEER). Immunocytochemistry and Western blotting of proteins associated with tight junctions in bEnd3 were performed to determine protein distribution before and after cooling. After focal cooling of the rat brain cortex, diffuse immunostaining for IgG was observed primarily around the small vasculature and in the extracellular spaces of parenchyma of the cortex. In cultured bEnd3, TEER significantly decreased during cooling (15 °C) and recovered to normal levels after rewarming to 37 °C. Immunocytochemistry and Western blotting showed that claudin-5, a critical regulatory protein for tight junctions, was translocated from the membrane to the cytoplasm after cooling in cultured bEnd3 cells. These results suggest that focal brain cooling may open the BBB transiently through an effect on tight junctions of brain microendothelial cells, and that therapeutically this approach may allow control of BBB function and drug delivery through the BBB.

  2. 'Stealth' nanoparticles evade neural immune cells but also evade major brain cell populations: Implications for PEG-based neurotherapeutics.

    PubMed

    Jenkins, Stuart I; Weinberg, Daniel; Al-Shakli, Arwa F; Fernandes, Alinda R; Yiu, Humphrey H P; Telling, Neil D; Roach, Paul; Chari, Divya M

    2016-02-28

    Surface engineering to control cell behavior is of high interest across the chemical engineering, drug delivery and biomaterial communities. Defined chemical strategies are necessary to tailor nanoscale protein interactions/adsorption, enabling control of cell behaviors for development of novel therapeutic strategies. Nanoparticle-based therapies benefit from such strategies but particle targeting to sites of neurological injury remains challenging due to circulatory immune clearance. As a strategy to overcome this barrier, the use of stealth coatings can reduce immune clearance and prolong circulatory times, thereby enhancing therapeutic capacity. Polyethylene glycol (PEG) is the most widely-used stealth coating and facilitates particle accumulation in the brain. However, once within the brain, the mode of handling of PEGylated particles by the resident immune cells of the brain itself (the 'microglia') is unknown. This is a critical question as it is well established that microglia avidly sequester nanoparticles, limiting their bioavailability and posing a major translational barrier. If PEGylation can be proved to promote evasion of microglia, then this information will be of high value in developing tailored nanoparticle-based therapies for neurological applications. Here, we have conducted the first comparative study of uptake of PEGylated particles by all the major (immune and non-immune) brain cell types. We prove for the first time that PEGylated nanoparticles evade major brain cell populations - a phenomenon which will enhance extracellular bioavailability. We demonstrate changes in protein coronas around these particles within biological media, and discuss how surface chemistry presentation may affect this process and subsequent cellular interactions.

  3. Mechanisms of modulation of brain microvascular endothelial cells function by thrombin.

    PubMed

    Brailoiu, Eugen; Shipsky, Megan M; Yan, Guang; Abood, Mary E; Brailoiu, G Cristina

    2017-02-15

    Brain microvascular endothelial cells are a critical component of the blood-brain barrier. They form a tight monolayer which is essential for maintaining the brain homeostasis. Blood-derived proteases such as thrombin may enter the brain during pathological conditions like trauma, stroke, and inflammation and further disrupts the permeability of the blood-brain barrier, via incompletely characterized mechanisms. We examined the underlying mechanisms evoked by thrombin in rat brain microvascular endothelial cells (RBMVEC). Our results indicate that thrombin, acting on protease-activated receptor 1 (PAR1) increases cytosolic Ca(2+) concentration in RBMVEC via Ca(2+) release from endoplasmic reticulum through inositol 1,4,5-trisphosphate receptors and Ca(2+) influx from extracellular space. Thrombin increases nitric oxide production; the effect is abolished by inhibition of the nitric oxide synthase or by antagonism of PAR1 receptors. In addition, thrombin increases mitochondrial and cytosolic reactive oxygen species production via PAR1-dependent mechanisms. Immunocytochemistry studies indicate that thrombin increases F-actin stress fibers, and disrupts the tight junctions. Thrombin increased the RBMVEC permeability assessed by a fluorescent flux assay. Taken together, our results indicate multiple mechanisms by which thrombin modulates the function of RBMVEC and may contribute to the blood-brain barrier dysfunction.

  4. Brain delivery of insulin boosted by intranasal coadministration with cell-penetrating peptides.

    PubMed

    Kamei, Noriyasu; Takeda-Morishita, Mariko

    2015-01-10

    Intranasal administration is considered as an alternative route to enable effective drug delivery to the central nervous system (CNS) by bypassing the blood-brain barrier. Several reports have proved that macromolecules can be transferred directly from the nasal cavity to the brain. However, strategies to enhance the delivery of macromolecules from the nasal cavity to CNS are needed because of their low delivery efficiencies via this route in general. We hypothesized that the delivery of biopharmaceuticals to the brain parenchyma can be facilitated by increasing the uptake of drugs by the nasal epithelium including supporting and neuronal cells to maximize the potentiality of the intranasal pathway. To test this hypothesis, the CNS-related model peptide insulin was intranasally coadministered with the cell-penetrating peptide (CPP) penetratin to mice. As a result, insulin coadministered with l- or d-penetratin reached the distal regions of the brain from the nasal cavity, including the cerebral cortex, cerebellum, and brain stem. In particular, d-penetratin could intranasally deliver insulin to the brain with a reduced risk of systemic insulin exposure. Thus, the results obtained in this study suggested that CPPs are potential tools for the brain delivery of peptide- and protein-based pharmaceuticals via intranasal administration.

  5. Early Activation of Primary Brain Microvascular Endothelial Cells by Nipah Virus Glycoprotein-Containing Particles

    PubMed Central

    Freitag, Tanja C.

    2015-01-01

    Nipah virus (NiV) is a highly pathogenic paramyxovirus that causes pronounced infection of brain endothelia and central nervous system (CNS) inflammation. Using primary porcine brain microvascular endothelial cells, we showed that upregulation of E-selectin precedes cytokine induction and is induced not only by infectious NiV but also by NiV-glycoprotein-containing virus-like particles. This demonstrates that very early events in NiV brain endothelial infection do not depend on NiV replication but can be triggered by the NiV glycoproteins alone. PMID:26676791

  6. Molecular Mechanisms Mediating Involvement of Glial Cells in Brain Plastic Remodeling in Epilepsy.

    PubMed

    Khaspekov, L G; Frumkina, L E

    2017-03-01

    In this review we summarize published data on the involvement of glial cells in molecular mechanisms underlying brain plastic reorganization in epilepsy. The role of astrocytes as glial elements in pathological plasticity in epilepsy is discussed. Data on the involvement of aquaporin-4 in epileptogenic plastic changes and on participation of microglia and extracellular matrix in dysregulation of synaptic transmission and plastic remodeling in epileptic brain tissue are reviewed.

  7. Loss of proteostasis induced by amyloid beta peptide in brain endothelial cells.

    PubMed

    Fonseca, Ana Catarina; Oliveira, Catarina R; Pereira, Cláudia F; Cardoso, Sandra M

    2014-06-01

    Abnormal accumulation of amyloid-β (Aβ) peptide in the brain is a pathological hallmark of Alzheimer's disease (AD). In addition to neurotoxic effects, Aβ also damages brain endothelial cells (ECs) and may thus contribute to the degeneration of cerebral vasculature, which has been proposed as an early pathogenic event in the course of AD and is able to trigger and/or potentiate the neurodegenerative process and cognitive decline. However, the mechanisms underlying Aβ-induced endothelial dysfunction are not completely understood. Here we hypothesized that Aβ impairs protein quality control mechanisms both in the secretory pathway and in the cytosol in brain ECs, leading cells to death. In rat brain RBE4 cells, we demonstrated that Aβ1-40 induces the failure of the ER stress-adaptive unfolded protein response (UPR), deregulates the ubiquitin-proteasome system (UPS) decreasing overall proteasome activity with accumulation of ubiquitinated proteins and impairs the autophagic protein degradation pathway due to failure in the autophagic flux, which culminates in cell demise. In conclusion, Aβ deregulates proteostasis in brain ECs and, as a consequence, these cells die by apoptosis.

  8. Contribution of mast cells to injury mechanisms in a mouse model of pediatric traumatic brain injury.

    PubMed

    Moretti, Raffaella; Chhor, Vibol; Bettati, Donatella; Banino, Elena; De Lucia, Silvana; Le Charpentier, Tifenn; Lebon, Sophie; Schwendimann, Leslie; Pansiot, Julien; Rasika, Sowmyalakshmi; Degos, Vincent; Titomanlio, Luigi; Gressens, Pierre; Fleiss, Bobbi

    2016-12-01

    The cognitive and behavioral deficits caused by traumatic brain injury (TBI) to the immature brain are more severe and persistent than injuries to the adult brain. Understanding this developmental sensitivity is critical because children under 4 years of age of sustain TBI more frequently than any other age group. One of the first events after TBI is the infiltration and degranulation of mast cells (MCs) in the brain, releasing a range of immunomodulatory substances; inhibition of these cells is neuroprotective in other types of neonatal brain injury. This study investigates for the first time the role of MCs in mediating injury in a P7 mouse model of pediatric contusion-induced TBI. We show that various neural cell types express histamine receptors and that histamine exacerbates excitotoxic cell death in primary cultured neurons. Cromoglycate, an inhibitor of MC degranulation, altered the inflammatory phenotype of microglia activated by TBI, reversing several changes but accentuating others, when administered before TBI. However, without regard to the time of cromoglycate administration, inhibiting MC degranulation did not affect cell loss, as evaluated by ventricular dilatation or cleaved caspase-3 labeling, or the density of activated microglia, neurons, or myelin. In double-heterozygous cKit mutant mice lacking MCs, this overall lack of effect was confirmed. These results suggest that the role of MCs in this model of pediatric TBI is restricted to subtle effects and that they are unlikely to be viable neurotherapeutic targets. © 2016 Wiley Periodicals, Inc.

  9. Multiple sclerosis: Brain-infiltrating CD8+ T cells persist as clonal expansions in the cerebrospinal fluid and blood

    PubMed Central

    Skulina, Christian; Schmidt, Stephan; Dornmair, Klaus; Babbe, Holger; Roers, Axel; Rajewsky, Klaus; Wekerle, Hartmut; Hohlfeld, Reinhard; Goebels, Norbert

    2004-01-01

    We surveyed the T cell receptor repertoire in three separate compartments (brain, cerebrospinal fluid, and blood) of two multiple sclerosis patients who initially had diagnostic brain biopsies to clarify their unusual clinical presentation but were subsequently confirmed to have typical multiple sclerosis. One of the brain biopsy specimens had been previously investigated by microdissection and single-cell PCR to determine the clonal composition of brain-infiltrating T cells at the single-cell level. Using complementarity-determining region 3 spectratyping, we identified several identical, expanded CD8+ (but not CD4+) T cell clones in all three compartments. Some of the expanded CD8+ T cells also occurred in sorted CD38+ blood cells, suggesting that they were activated. Strikingly, some of the brain-infiltrating CD8+ T cell clones persisted for >5 years in the cerebrospinal fluid and/or blood and may thus contribute to the progression of the disease. PMID:14983026

  10. Uptake and transport of superparamagnetic iron oxide nanoparticles through human brain capillary endothelial cells.

    PubMed

    Thomsen, L B; Linemann, T; Pondman, K M; Lichota, J; Kim, K S; Pieters, R J; Visser, G M; Moos, T

    2013-10-16

    The blood-brain barrier (BBB) formed by brain capillary endothelial cells (BCECs) constitutes a firm physical, chemical, and immunological barrier, making the brain accessible to only a few percent of potential drugs intended for treatment inside the central nervous system. With the purpose of overcoming the restraints of the BBB by allowing the transport of drugs, siRNA, or DNA into the brain, a novel approach is to use superparamagnetic iron oxide nanoparticles (SPIONs) as drug carriers. The aim of this study was to investigate the ability of fluorescent SPIONs to pass through human brain microvascular endothelial cells facilitated by an external magnet. The ability of SPIONs to penetrate the barrier was shown to be significantly stronger in the presence of an external magnetic force in an in vitro BBB model. Hence, particles added to the luminal side of the in vitro BBB model were found in astrocytes cocultured at a remote distance on the abluminal side, indicating that particles were transported through the barrier and taken up by astrocytes. Addition of the SPIONs to the culture medium did not negatively affect the viability of the endothelial cells. The magnetic force-mediated dragging of SPIONs through BCECs may denote a novel mechanism for the delivery of drugs to the brain.

  11. Gain of glucose-independent growth upon metastasis of breast cancer cells to the brain

    PubMed Central

    Chen, Jinyu; Lee, Ho-Jeong; Wu, Xuefeng; Huo, Lei; Kim, Sun-Jin; Xu, Lei; Wang, Yan; He, Junqing; Bollu, Lakshmi Reddy; Gao, Guang; Su, Fei; Briggs, James; Liu, Xiaojing; Melman, Tamar; Asara, John M.; Fidler, Isaiah J.; Cantley, Lewis C.; Locasale, Jason W.; Weihua, Zhang

    2014-01-01

    Breast cancer brain metastasis is resistant to therapy and a particularly poor prognostic feature in patient survival. Altered metabolism is a common feature of cancer cells but little is known as to what metabolic changes benefit breast cancer brain metastases. We found that brain-metastatic breast cancer cells evolved the ability to survive and proliferate independent of glucose due to enhanced gluconeogenesis and oxidations of glutamine and branched chain amino acids, which together sustain the non-oxidative pentose pathway for purine synthesis. Silencing expression of fructose-1,6-bisphosphatases (FBPs) in brain metastatic cells reduced their viability and improved the survival of metastasis-bearing immunocompetent hosts. Clinically, we showed that brain metastases from human breast cancer patients expressed higher levels of FBP and glycogen than the corresponding primary tumors. Together, our findings identify a critical metabolic condition required to sustain brain metastasis, and suggest that targeting gluconeogenesis may help eradicate this deadly feature in advanced breast cancer patients. PMID:25511375

  12. Oxidative Stress Induced by Cigarette Smoke Extracts in Human Brain Cells (T98G) and Human Brain Microvascular Endothelial Cells (HBMEC) in Mono- and Co-Culture.

    PubMed

    Kim, Ju-Hyeong; Cho, Myung-Haing; Choi, Kyung-Chul; Lee, Kyuhong; Kim, Kwang-Sik; Shim, Soon-Mi

    2015-01-01

    The objective of the current study was to examine oxidative stress induced by cigarette smoke extract (CSE) or cigarette smoke condensate (CSC) in human brain cells (T98G) and human brain microvascular endothelial cells (HBMEC) in mono- and co-culture systems. Cell viability of T98G cells exposed to CSC (0.05-4 mg/ml) was significantly decreased compared to CSE (0.025-20%). There were no marked differences between quantities of reactive oxygen species (ROS) generation by either CSE (2, 4, and 10%) or CSC (0.2, 0.4, and 0.8 mg/ml) treatment compared to control. However, a significant effect was noted in ROS generation following CSC incubation at 4mg/ml. Cellular integrity of HBMEC decreased to 74 and 64% within 120 h of exposure at the IC50 value of CSE and CSC, respectively. This study suggests that chronic exposure to cigarette smoking might initiate damage to the blood-brain barrier.

  13. Prion Protein Deficiency Causes Diverse Proteome Shifts in Cell Models That Escape Detection in Brain Tissue

    PubMed Central

    Mehrabian, Mohadeseh; Brethour, Dylan; Williams, Declan; Wang, Hansen; Arnould, Hélène; Schneider, Benoit; Schmitt-Ulms, Gerold

    2016-01-01

    A popular method for studying the function of a given protein is to generate and characterize a suitable model deficient for its expression. For the prion protein (PrP), best known for its role in several invariably fatal neurodegenerative diseases, a natural choice, therefore, would be to undertake such studies with brain samples. We recently documented the surprising observation that PrP deficiency caused a loss or enhancement of NCAM1 polysialylation, dependent on the cell model used. To identify possible causes for this disparity, we set out to systematically investigate the consequence of PrP deficiency on the global proteome in brain tissue and in four distinct cell models. Here we report that PrP deficiency causes robust but surprisingly divergent changes to the global proteomes of cell models but has no discernible impact on the global brain proteome. Amongst >1,500 proteins whose levels were compared in wild-type and PrP-deficient models, members of the MARCKS protein family exhibited pronounced, yet cell model-dependent changes to their steady-state levels. Follow-up experiments revealed that PrP collaborates with members of the MARCKS protein family in its control of NCAM1 polysialylation. We conclude that the physiological function of PrP may be masked in analyses of complex brain samples but its cell-type specific influence on a lipid raft-based NCAM1-related cell biology comes to the fore in investigations of specific cell types. PMID:27327609

  14. Measuring antigen presentation in mouse brain endothelial cells ex vivo and in vitro.

    PubMed

    Howland, Shanshan W; Gun, Sin Yee; Claser, Carla; Poh, Chek Meng; Rénia, Laurent

    2015-12-01

    We have recently demonstrated that brain endothelial cells cross-present parasite antigen during mouse experimental cerebral malaria (ECM). Here we describe a 2-d protocol to detect cross-presentation by isolating the brain microvessels and incubating them with a reporter cell line that expresses lacZ upon detection of the relevant peptide-major histocompatibility complex. After X-gal staining, a typical positive result consists of hundreds of blue spots, compared with fewer than 20 spots from a naive brain. The assay is generalizable to other disease contexts by using reporter cells that express appropriate specific T cell receptors. Also described is the protocol for culturing endothelial cells from brain microvessels isolated from naive mice. After 7-10 d, an in vitro cross-presentation assay can be performed by adding interferon-γ, antigen (e.g., Plasmodium berghei-infected red blood cells) and reporter cells in sequence over 3 d. This is useful for comparing different antigen forms or for probing the effects of various interventions.

  15. Influence of curvature on the morphology of brain microvascular endothelial cells

    NASA Astrophysics Data System (ADS)

    Ye, Mao; Yang, Zhen; Wong, Andrew; Searson, Peter; Searson Group Team

    2013-03-01

    There are hundreds or thousands of endothelial cells around the perimeter of a single artery or vein, and hence an individual cell experiences little curvature. In contrast, a single endothelial cell may wrap around itself to form the lumen of a brain capillary. Curvature plays a key role in many biological, chemical and physical processes, however, its role in dictating the morphology and polarization of brain capillary endothelial cells has not been investigated. We hypothesize that curvature and shear flow play a key role in determining the structure and function of the blood-brain barrier (BBB). We have developed the ``rod'' assay to study the influence of curvature on the morphology of confluent monolayers of endothelial cells. In this assay cells are plated onto glass rods pulled down to the desired diameter in the range from 5 - 500 μm and coated with collagen. We show that curvature has a significant influence on the morphology of endothelial cells and may have an important role in blood-brain barrier function.

  16. Characterization of multiciliated ependymal cells that emerge in the neurogenic niche of the aged zebrafish brain.

    PubMed

    Ogino, Takashi; Sawada, Masato; Takase, Hiroshi; Nakai, Chiemi; Herranz-Pérez, Vicente; Cebrián-Silla, Arantxa; Kaneko, Naoko; García-Verdugo, José Manuel; Sawamoto, Kazunobu

    2016-10-15

    In mammals, ventricular walls of the developing brain maintain a neurogenic niche, in which radial glial cells act as neural stem cells (NSCs) and generate new neurons in the embryo. In the adult brain, the neurogenic niche is maintained in the ventricular-subventricular zone (V-SVZ) of the lateral wall of lateral ventricles and the hippocampal dentate gyrus. In the neonatal V-SVZ, radial glial cells transform into astrocytic postnatal NSCs and multiciliated ependymal cells. On the other hand, in zebrafish, radial glial cells continue to cover the surface of the adult telencephalic ventricle and maintain a higher neurogenic potential in the adult brain. However, the cell composition of the neurogenic niche of the aged zebrafish brain has not been investigated. Here we show that multiciliated ependymal cells emerge in the neurogenic niche of the aged zebrafish telencephalon. These multiciliated cells appear predominantly in the dorsal part of the ventral telencephalic ventricular zone, which also contains clusters of migrating new neurons. Scanning electron microscopy and live imaging analyses indicated that these multiple cilia beat coordinately and generate constant fluid flow within the ventral telencephalic ventricle. Analysis of the cell composition by transmission electron microscopy revealed that the neurogenic niche in the aged zebrafish contains different types of cells, with ultrastructures similar to those of ependymal cells, transit-amplifying cells, and migrating new neurons in postnatal mice. These data suggest that the transformation capacity of radial glial cells is conserved but that its timing is different between fish and mice. J. Comp. Neurol. 524:2982-2992, 2016. © 2016 Wiley Periodicals, Inc.

  17. Dynamics of PDGFRβ expression in different cell types after brain injury.

    PubMed

    Kyyriäinen, Jenni; Ekolle Ndode-Ekane, Xavier; Pitkänen, Asla

    2017-02-01

    Platelet-derived growth factor receptor β (PDGFRβ) is upregulated after brain injury and its depletion results in the blood-brain barrier (BBB) damage. We investigated the time-window and localization of PDGFRβ expression in mice with intrahippocampal kainic acid-induced status epilepticus (SE) and in rats with lateral fluid-percussion-induced traumatic brain injury (TBI). Tissue immunohistochemistry was evaluated at several time-points after SE and TBI. The distribution of PDGFRβ was analyzed, and its cell type-specific expression was verified with double/triple-labeling of astrocytes (GFAP), NG2 cells, and endothelial cells (RECA-1). In normal mouse hippocampus, we found evenly distributed PDGFRβ+ parenchymal cells. In double-labeling, all NG2+ and 40%-60% GFAP+ cells were PDGFRβ+. After SE, PDGFRβ+ cells clustered in the ipsilateral hilus (178% of that in controls at fourth day, 225% at seventh day, P < 0.05) and in CA3 (201% at seventh day, P < 0.05), but the total number of PDGFRβ+ cells was not altered. As in controls, PDGFRβ-immunoreactivity was detected in parenchymal NG2+ and GFAP+ cells. We also observed PDGFRβ+ structural pericytes, detached reactive pericytes, and endothelial cells. After TBI, PDGFRβ+ cells clustered in the perilesional cortex and thalamus, particularly during the first post-injury week. PDGFRβ immunopositivity was observed in NG2+ and GFAP+ cells, structural pericytes, detached reactive pericytes, and endothelial cells. In some animals, PDGFRβ vascular staining was observed around the cortical glial scar for up to 3 months. Our data revealed an acute accumulation of PDGFRβ+ BBB-related cells in degenerating brain areas, which can be long lasting, suggesting an active role for PDGFRβ-signaling in blood vessel and post-injury tissue recovery. GLIA 2017;65:322-341.

  18. Zika virus cell tropism in the developing human brain and inhibition by azithromycin

    PubMed Central

    Retallack, Hanna; Di Lullo, Elizabeth; Arias, Carolina; Knopp, Kristeene A.; Laurie, Matthew T.; Sandoval-Espinosa, Carmen; Mancia Leon, Walter R.; Krencik, Robert; Ullian, Erik M.; Spatazza, Julien; Pollen, Alex A.; Mandel-Brehm, Caleigh; Nowakowski, Tomasz J.; Kriegstein, Arnold R.; DeRisi, Joseph L.

    2016-01-01

    The rapid spread of Zika virus (ZIKV) and its association with abnormal brain development constitute a global health emergency. Congenital ZIKV infection produces a range of mild to severe pathologies, including microcephaly. To understand the pathophysiology of ZIKV infection, we used models of the developing brain that faithfully recapitulate the tissue architecture in early to midgestation. We identify the brain cell populations that are most susceptible to ZIKV infection in primary human tissue, provide evidence for a mechanism of viral entry, and show that a commonly used antibiotic protects cultured brain cells by reducing viral proliferation. In the brain, ZIKV preferentially infected neural stem cells, astrocytes, oligodendrocyte precursor cells, and microglia, whereas neurons were less susceptible to infection. These findings suggest mechanisms for microcephaly and other pathologic features of infants with congenital ZIKV infection that are not explained by neural stem cell infection alone, such as calcifications in the cortical plate. Furthermore, we find that blocking the glia-enriched putative viral entry receptor AXL reduced ZIKV infection of astrocytes in vitro, and genetic knockdown of AXL in a glial cell line nearly abolished infection. Finally, we evaluate 2,177 compounds, focusing on drugs safe in pregnancy. We show that the macrolide antibiotic azithromycin reduced viral proliferation and virus-induced cytopathic effects in glial cell lines and human astrocytes. Our characterization of infection in the developing human brain clarifies the pathogenesis of congenital ZIKV infection and provides the basis for investigating possible therapeutic strategies to safely alleviate or prevent the most severe consequences of the epidemic. PMID:27911847

  19. A simple method for isolating and culturing the rat brain microvascular endothelial cells.

    PubMed

    Liu, Yang; Xue, Qiang; Tang, Qing; Hou, Min; Qi, Hongyi; Chen, Gang; Chen, Weihai; Zhang, Jifen; Chen, Yi; Xu, Xiaoyu

    2013-11-01

    Brain microvascular endothelial cells (BMECs), a main component of the blood-brain barrier, play a critical role in the pathogenesis of many brain diseases. The primary culture of BMECs has been used in various models for studying cerebrovascular diseases in vitro. However, there are still several problems existing in the isolation and cultivation of primary rat BMECs, such as low yield, contamination with other cell types, and requirement of a large number of animals and expensive growth factor. In this study, we describe a simple, economical (without any growth factor) and repeatable method to obtain endothelial cells with high purity (>99%) and yield (about 2.2×10(7) per rat) from cerebral cortexes of neonatal rat, mainly from gray matter. In vitro examinations determined that the isolated cells expressed typical phenotypic markers of differentiated brain endothelium such as multiple drug resistant protein, von Willebrand factor, platelet endothelial cell adhesion molecule 1 (PECAM-1/CD31), and intercellular adhesion molecule (ICAM). These cells also possessed morphological and ultra-structural characteristics that were observed by phase contrast microscope and electric microscope. Then GFAP and α-SMA were used, respectively, to identify astrocyte and pericyte which were potential to contaminate primary culturing of BMECs. And specific reaction of endothelial cells to external stimulation was tested by culture with TNF-α for 24h. All these results of our experiments supply that our protocol provides an effective and reliable method to obtain high purity and yield of rat BMECs and offers a useful tool for studying cellular physiology, cerebrovascular diseases, brain tumors, blood-brain barrier and neurovascular units, etc.

  20. Brain metastatic cancer cells release microRNA-181c-containing extracellular vesicles capable of destructing blood-brain barrier.

    PubMed

    Tominaga, Naoomi; Kosaka, Nobuyoshi; Ono, Makiko; Katsuda, Takeshi; Yoshioka, Yusuke; Tamura, Kenji; Lötvall, Jan; Nakagama, Hitoshi; Ochiya, Takahiro

    2015-04-01

    Brain metastasis is an important cause of mortality in breast cancer patients. A key event during brain metastasis is the migration of cancer cells through blood-brain barrier (BBB). However, the molecular mechanism behind the passage through this natural barrier remains unclear. Here we show that cancer-derived extracellular vesicles (EVs), mediators of cell-cell communication via delivery of proteins and microRNAs (miRNAs), trigger the breakdown of BBB. Importantly, miR-181c promotes the destruction of BBB through the abnormal localization of actin via the downregulation of its target gene, PDPK1. PDPK1 degradation by miR-181c leads to the downregulation of phosphorylated cofilin and the resultant activated cofilin-induced modulation of actin dynamics. Furthermore, we demonstrate that systemic injection of brain metastatic cancer cell-derived EVs promoted brain metastasis of breast cancer cell lines and are preferentially incorporated into the brain in vivo. Taken together, these results indicate a novel mechanism of brain metastasis mediated by EVs that triggers the destruction of BBB.

  1. Improved methodology for identification of protists and microalgae from plankton samples preserved in Lugol's iodine solution: combining microscopic analysis with single-cell PCR.

    PubMed

    Auinger, Barbara M; Pfandl, Karin; Boenigk, Jens

    2008-04-01

    Here we introduce a method for quantitative analysis of planktonic protists and microalgae from preserved field samples combining morphological and small-subunit (SSU) rRNA gene sequence analysis. We linked a microscopic screening with PCR of single cells using field samples preserved with Lugol's iodine solution. Cells possessing a rigid cell wall were incubated with Viscozyme and subsequently with proteinase K for cell disruption; this was unnecessary for fragile cells. The addition of sodium thiosulfate to the PCR tube considerably decreased the inhibiting effect of the fixative (iodine) on the PCR and thus allowed for successful single-cell PCR even of long DNA fragments (up to as many as 3,000 base pairs). We further applied the protocol to investigate the dominant SSU rRNA genotypes in distinct flagellate morphospecies originating from different samples. We hypothesized that despite the morphological similarity, protist morphospecies in different habitats or sampled during different seasons are represented by different genotypes. Our results indicate species-specific differences: the two species Ochromonas sp. and Dinobryon divergens were represented by several different genotypes each, and for the latter species, the dominating genotype differed with habitat. In contrast, Dinobryon pediforme, Dinobryon bavaricum, and Synura sphagnicola were exclusively represented by a single genotype each, and the respective genotype was the same in different samples. In summary, our results highlight the significance of molecular variation within protist morphospecies.

  2. Planar Organization of Multiciliated Ependymal (E1) Cells in the Brain Ventricular Epithelium.

    PubMed

    Ohata, Shinya; Alvarez-Buylla, Arturo

    2016-08-01

    Cerebrospinal fluid (CSF) continuously flows through the cerebral ventricles, a process essential for brain homeostasis. Multiciliated ependymal (E1) cells line the walls of the ventricles and contribute importantly to CSF flow through ciliary beating. Key to this function is the rotational and translational planar cell polarity (PCP) of E1 cells. Defects in the PCP of E1 cells can result in abnormal CSF accumulation and hydrocephalus. Here, we integrate recent data on the roles of early CSF flow in the embryonic ventricles, PCP regulators (e.g., Vangl2 and Dishevelled), and cytoskeletal networks in the establishment, refinement, and maintenance of E1 cells' PCP. The planar organization mechanisms of E1 cells could explain how CSF flow contributes to brain function and may help in the diagnosis and prevention of hydrocephalus.

  3. Effect of glutamate analogues on brain tumor cell lines.

    PubMed

    Campbell, G L; Bartel, R; Freidman, H S; Bigner, D D

    1985-10-01

    Glutamate analogues have been used in many different experimental approaches in neurobiology. A small number of these analogues have been classified as gliotoxic. We have examined the effect of seven glutamate analogues (five gliotoxic and two neurotoxic) on the growth and viability of four human glioma cell lines, one human medulloblastoma cell line, and one human sarcoma cell line. Aminoadipic acid and homocysteic acid predominantly affected the growth of two glioma cell lines in the presence of 4 mM glutamine. Phosphonobutyric acid predominantly affected the other two glioma cell lines and the medulloblastoma cell line in the presence of 4 mM glutamine. In medium containing no glutamine, all three analogues had marked effects on all the cell lines except the sarcoma cell line. These effects were dose dependent. We postulate that these results can in part be explained on the basis of metabolic compartmentalization.

  4. BK K+ channel blockade inhibits radiation-induced migration/brain infiltration of glioblastoma cells

    PubMed Central

    Klumpp, Lukas; Haehl, Erik; Schilbach, Karin; Lukowski, Robert; Kühnle, Matthias; Bernhardt, Günther; Buschauer, Armin; Zips, Daniel; Ruth, Peter; Huber, Stephan M.

    2016-01-01

    Infiltration of the brain by glioblastoma cells reportedly requires Ca2+ signals and BK K+ channels that program and drive glioblastoma cell migration, respectively. Ionizing radiation (IR) has been shown to induce expression of the chemokine SDF-1, to alter the Ca2+ signaling, and to stimulate cell migration of glioblastoma cells. Here, we quantified fractionated IR-induced migration/brain infiltration of human glioblastoma cells in vitro and in an orthotopic mouse model and analyzed the role of SDF-1/CXCR4 signaling and BK channels. To this end, the radiation-induced migratory phenotypes of human T98G and far-red fluorescent U-87MG-Katushka glioblastoma cells were characterized by mRNA and protein expression, fura-2 Ca2+ imaging, BK patch-clamp recording and transfilter migration assay. In addition, U-87MG-Katushka cells were grown to solid glioblastomas in the right hemispheres of immunocompromised mice, fractionated irradiated (6 MV photons) with 5 × 0 or 5 × 2 Gy, and SDF-1, CXCR4, and BK protein expression by the tumor as well as glioblastoma brain infiltration was analyzed in dependence on BK channel targeting by systemic paxilline application concomitant to IR. As a result, IR stimulated SDF-1 signaling and induced migration of glioblastoma cells in vitro and in vivo. Importantly, paxilline blocked IR-induced migration in vivo. Collectively, our data demonstrate that fractionated IR of glioblastoma stimulates and BK K+ channel targeting mitigates migration and brain infiltration of glioblastoma cells in vivo. This suggests that BK channel targeting might represent a novel approach to overcome radiation-induced spreading of malignant brain tumors during radiotherapy. PMID:26893360

  5. Trophic factors and cell therapy to stimulate brain repair after ischaemic stroke

    PubMed Central

    Gutiérrez-Fernández, María; Fuentes, Blanca; Rodríguez-Frutos, Berta; Ramos-Cejudo, Jaime; Vallejo-Cremades, María Teresa; Díez-Tejedor, Exuperio

    2012-01-01

    Brain repair involves a compendium of natural mechanisms that are activated following stroke. From a therapeutic viewpoint, reparative therapies that encourage cerebral plasticity are needed. In the last years, it has been demonstrated that modulatory treatments for brain repair such as trophic factor- and stem cell-based therapies can promote neurogenesis, gliogenesis, oligodendrogenesis, synaptogenesis and angiogenesis, all of which having a beneficial impact on infarct volume, cell death and, finally, and most importantly, on the functional recovery. However, even when promising results have been obtained in a wide range of experimental animal models and conditions these preliminary results have not yet demonstrated their clinical efficacy. Here, we focus on brain repair modulatory treatments for ischaemic stroke, that use trophic factors, drugs with trophic effects and stem cell therapy. Important and still unanswered questions for translational research ranging from experimental animal models to recent and ongoing clinical trials are reviewed here. PMID:22452968

  6. Joining microelectronics and microionics: Nerve cells and brain tissue on semiconductor chips

    NASA Astrophysics Data System (ADS)

    Fromherz, Peter

    2008-09-01

    The direct electrical interfacing of semiconductor chips with individual nerve cells and with brain tissue is considered. At first, the structure of the cell-chip contact is described and then the electrical coupling is characterized between ion channels, the electrical elements of nerve cells, and transistors and capacitors of silicon chips. On that basis, the signal transmission between microelectronics and microionics is implemented in both directions. Simple hybrid systems are assembled with neuron pairs and with small neuronal networks. Finally, the interfacing with capacitors and transistors is extended to brain tissue on silicon. The application of CMOS chips with capacitively coupled recording sites allows an imaging of neuronal activity with high spatiotemporal resolution. Goal of the work is an integration of neuronal network dynamics and digital electronics on a microscopic level for applications in brain research, medical prosthetics and information technology.

  7. Effects of Cell Phone Radiofrequency Signal Exposure on Brain Glucos Metabolism

    SciTech Connect

    Volkow, N.D.; Wang, G.; Volkow, N.D.; Tomasi, D.; Wang, G.-J.; Vaska, P.; Fowler, J.S.; Telang, F.; Alexoff, D.; Logan, J.; Wong, C.

    2011-03-01

    The dramatic increase in use of cellular telephones has generated concern about possible negative effects of radiofrequency signals delivered to the brain. However, whether acute cell phone exposure affects the human brain is unclear. To evaluate if acute cell phone exposure affects brain glucose metabolism, a marker of brain activity. Randomized crossover study conducted between January 1 and December 31, 2009, at a single US laboratory among 47 healthy participants recruited from the community. Cell phones were placed on the left and right ears and positron emission tomography with ({sup 18}F)fluorodeoxyglucose injection was used to measure brain glucose metabolism twice, once with the right cell phone activated (sound muted) for 50 minutes ('on' condition) and once with both cell phones deactivated ('off' condition). Statistical parametric mapping was used to compare metabolism between on and off conditions using paired t tests, and Pearson linear correlations were used to verify the association of metabolism and estimated amplitude of radiofrequency-modulated electromagnetic waves emitted by the cell phone. Clusters with at least 1000 voxels (volume >8 cm{sup 3}) and P < .05 (corrected for multiple comparisons) were considered significant. Brain glucose metabolism computed as absolute metabolism ({micro}mol/100 g per minute) and as normalized metabolism (region/whole brain). Whole-brain metabolism did not differ between on and off conditions. In contrast, metabolism in the region closest to the antenna (orbitofrontal cortex and temporal pole) was significantly higher for on than off conditions (35.7 vs 33.3 {micro}mol/100 g per minute; mean difference, 2.4 [95% confidence interval, 0.67-4.2]; P = .004). The increases were significantly correlated with the estimated electromagnetic field amplitudes both for absolute metabolism (R = 0.95, P < .001) and normalized metabolism (R = 0.89; P < .001). In healthy participants and compared with no exposure, 50-minute

  8. Live-cell imaging to detect phosphatidylserine externalization in brain endothelial cells exposed to ionizing radiation: implications for the treatment of brain arteriovenous malformations.

    PubMed

    Zhao, Zhenjun; Johnson, Michael S; Chen, Biyi; Grace, Michael; Ukath, Jaysree; Lee, Vivienne S; McRobb, Lucinda S; Sedger, Lisa M; Stoodley, Marcus A

    2016-06-01

    OBJECT Stereotactic radiosurgery (SRS) is an established intervention for brain arteriovenous malformations (AVMs). The processes of AVM vessel occlusion after SRS are poorly understood. To improve SRS efficacy, it is important to understand the cellular response of blood vessels to radiation. The molecular changes on the surface of AVM endothelial cells after irradiation may also be used for vascular targeting. This study investigates radiation-induced externalization of phosphatidylserine (PS) on endothelial cells using live-cell imaging. METHODS An immortalized cell line generated from mouse brain endothelium, bEnd.3 cells, was cultured and irradiated at different radiation doses using a linear accelerator. PS externalization in the cells was subsequently visualized using polarity-sensitive indicator of viability and apoptosis (pSIVA)-IANBD, a polarity-sensitive probe. Live-cell imaging was used to monitor PS externalization in real time. The effects of radiation on the cell cycle of bEnd.3 cells were also examined by flow cytometry. RESULTS Ionizing radiation effects are dose dependent. Reduction in the cell proliferation rate was observed after exp