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      Long-term stability and computational analysis of migration patterns of L- MYC immortalized neural stem cells in the brain

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          Abstract

          Background

          Preclinical studies indicate that neural stem cells (NSCs) can limit or reverse central nervous system (CNS) damage through delivery of therapeutic agents for cell regeneration. Clinical translation of cell-based therapies raises concerns about long-term stability, differentiation and fate, and absence of tumorigenicity of these cells, as well as manufacturing time required to produce therapeutic cells in quantities sufficient for clinical use. Allogeneic NSC lines are in growing demand due to challenges inherent in using autologous stem cells, including production costs that limit availability to patients.

          Methods/Principal findings

          We demonstrate the long-term stability of L- MYC immortalized human NSCs (LM-NSC008) cells in vivo, including engraftment, migration, and absence of tumorigenicity in mouse brains for up to nine months. We also examined the distributions of engrafted LM-NSC008 cells within brain, and present computational techniques to analyze NSC migration characteristics in relation to intrinsic brain structures.

          Conclusions/Significance

          This computational analysis of NSC distributions following implantation provides proof-of-concept for the development of computational models that can be used clinically to predict NSC migration paths in patients. Previously, models of preferential migration of malignant tumor cells along white matter tracts have been used to predict their final distributions. We suggest that quantitative measures of tissue orientation and white matter tracts determined from MR images can be used in a diffusion tensor imaging tractography-like approach to describe the most likely migration routes and final distributions of NSCs administered in a clinical setting. Such a model could be very useful in choosing the optimal anatomical locations for NSC administration to patients to achieve maximum therapeutic effects.

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          Most cited references32

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          Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system.

          Neurogenesis in the mammalian central nervous system is believed to end in the period just after birth; in the mouse striatum no new neurons are produced after the first few days after birth. In this study, cells isolated from the striatum of the adult mouse brain were induced to proliferate in vitro by epidermal growth factor. The proliferating cells initially expressed nestin, an intermediate filament found in neuroepithelial stem cells, and subsequently developed the morphology and antigenic properties of neurons and astrocytes. Newly generated cells with neuronal morphology were immunoreactive for gamma-aminobutyric acid and substance P, two neurotransmitters of the adult striatum in vivo. Thus, cells of the adult mouse striatum have the capacity to divide and differentiate into neurons and astrocytes.
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            Experimental investigation of collagen waviness and orientation in the arterial adventitia using confocal laser scanning microscopy.

            Mechanical properties of the adventitia are largely determined by the organization of collagen fibers. Measurements on the waviness and orientation of collagen, particularly at the zero-stress state, are necessary to relate the structural organization of collagen to the mechanical response of the adventitia. Using the fluorescence collagen marker CNA38-OG488 and confocal laser scanning microscopy, we imaged collagen fibers in the adventitia of rabbit common carotid arteries ex vivo. The arteries were cut open along their longitudinal axes to get the zero-stress state. We used semi-manual and automatic techniques to measure parameters related to the waviness and orientation of fibers. Our results showed that the straightness parameter (defined as the ratio between the distances of endpoints of a fiber to its length) was distributed with a beta distribution (mean value 0.72, variance 0.028) and did not depend on the mean angle orientation of fibers. Local angular density distributions revealed four axially symmetric families of fibers with mean directions of 0°, 90°, 43° and -43°, with respect to the axial direction of the artery, and corresponding circular standard deviations of 40°, 47°, 37° and 37°. The distribution of local orientations was shifted to the circumferential direction when measured in arteries at the zero-load state (intact), as compared to arteries at the zero-stress state (cut-open). Information on collagen fiber waviness and orientation, such as obtained in this study, could be used to develop structural models of the adventitia, providing better means for analyzing and understanding the mechanical properties of vascular wall.
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              The contribution of gliosis to diffusion tensor anisotropy and tractography following traumatic brain injury: validation in the rat using Fourier analysis of stained tissue sections.

              Diffusion tensor imaging is highly sensitive to the microstructural integrity of the brain and has uncovered significant abnormalities following traumatic brain injury not appreciated through other methods. It is hoped that this increased sensitivity will aid in the detection and prognostication in patients with traumatic injury. However, the pathological substrates of such changes are poorly understood. Specifically, decreases in fractional anisotropy derived from diffusion tensor imaging are consistent with axonal injury, myelin injury or both in white matter fibres. In contrast, in both humans and animal models, increases in fractional anisotropy have been suggested to reflect axonal regeneration and plasticity, but the direct histological evidence for such changes remains tenuous. We developed a method to quantify the anisotropy of stained histological sections using Fourier analysis, and applied the method to a rat controlled cortical impact model to identify the specific pathological features that give rise to the diffusion tensor imaging changes in subacute to chronic traumatic brain injury. A multiple linear regression was performed to relate the histological measurements to the measured diffusion tensor changes. The results show that anisotropy was significantly increased (P < 0.001) in the perilesioned cortex following injury. Cortical anisotropy was independently associated (standardized β = 0.62, P = 0.04) with the coherent organization of reactive astrocytes (i.e. gliosis) and was not attributed to axons. By comparison, a decrease in white matter anisotropy (P < 0.001) was significantly related to demyelination (β = 0.75, P = 0.0015) and to a lesser extent, axonal degeneration (β = -0.48, P = 0.043). Gliosis within the lesioned cortex also influenced diffusion tensor tractography, highlighting the fact that spurious tracts in the injured brain may not necessarily reflect continuous axons and may instead depict glial scarring. The current study demonstrates a novel method to relate pathology to diffusion tensor imaging findings, elucidates the underlying mechanisms of anisotropy changes following traumatic brain injury and significantly impacts the clinical interpretation of diffusion tensor imaging findings in the injured brain.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data curationRole: Formal analysisRole: InvestigationRole: Project administrationRole: SoftwareRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: Funding acquisitionRole: MethodologyRole: ValidationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Formal analysisRole: Funding acquisitionRole: ValidationRole: Visualization
                Role: Data curationRole: Funding acquisition
                Role: Funding acquisitionRole: Visualization
                Role: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: Project administrationRole: ResourcesRole: SupervisionRole: ValidationRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                2 August 2018
                2018
                : 13
                : 8
                : e0199967
                Affiliations
                [1 ] Department of Information Sciences, Division of Mathematical Oncology, Beckman Research Institute of City of Hope, Duarte, California, United States of America
                [2 ] Department of Developmental and Stem Cell Biology, Beckman Research Institute of City of Hope, Duarte, California, United States of America
                Universidad de Jaen, SPAIN
                Author notes

                Competing Interests: KSA is a shareholder, director, and officer of Therabiologics, Inc. No commercial funding was received for this manuscript and does not alter our adherence to PLOS ONE policies on sharing data and materials.

                Author information
                http://orcid.org/0000-0002-7094-626X
                Article
                PONE-D-18-07057
                10.1371/journal.pone.0199967
                6071994
                30071048
                498a8683-33c0-4926-94cc-c336e4e85947
                © 2018 Rockne et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 6 March 2018
                : 18 June 2018
                Page count
                Figures: 4, Tables: 0, Pages: 15
                Funding
                Funded by: funder-id http://dx.doi.org/10.13039/100000054, National Cancer Institute;
                Award ID: P30CA033572
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100001445, Alex's Lemonade Stand Foundation for Childhood Cancer;
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000902, Pediatric Cancer Research Foundation;
                Award Recipient :
                Funded by: Excellence Award at City of Hope
                Award Recipient :
                Research was supported by Alex’s Lemonade Stand Foundation, Pediatric Cancer Research Foundation, Excellence Award at City of Hope and the National Cancer Institute of the National Institutes of Health under grant number P30CA033572. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
                Categories
                Research Article
                Biology and Life Sciences
                Anatomy
                Nervous System
                Central Nervous System
                Medicine and Health Sciences
                Anatomy
                Nervous System
                Central Nervous System
                Medicine and Health Sciences
                Neurology
                Brain Damage
                Research and Analysis Methods
                Histochemistry and Cytochemistry Techniques
                Immunohistochemistry Techniques
                Research and Analysis Methods
                Immunologic Techniques
                Immunohistochemistry Techniques
                Biology and Life Sciences
                Anatomy
                Brain
                Corpus Callosum
                Medicine and Health Sciences
                Anatomy
                Brain
                Corpus Callosum
                Physical Sciences
                Physics
                Condensed Matter Physics
                Anisotropy
                Physical Sciences
                Materials Science
                Material Properties
                Anisotropy
                Research and Analysis Methods
                Specimen Preparation and Treatment
                Staining
                Cell Staining
                Biology and Life Sciences
                Cell Biology
                Cell Motility
                Cell Migration
                Biology and Life Sciences
                Developmental Biology
                Cell Migration
                Physical Sciences
                Mathematics
                Algebra
                Linear Algebra
                Eigenvectors
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                All relevant data are within the paper and its Supporting Information file.

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