title: Bioluminescence imaging of stroke-induced endogenous neural stem cell response.
abstract: Brain injury following stroke affects neurogenesis in the adult mammalian brain. However, a complete understanding of the origin and fate of the endogenous neural stem cells (eNSCs) in vivo is missing. Tools and technology that allow non-invasive imaging and tracking of eNSCs in living animals will help to overcome this hurdle. In this study, we aimed to monitor eNSCs in a photothrombotic (PT) stroke model using in vivo bioluminescence imaging (BLI). In a first strategy, inducible transgenic mice expressing firefly luciferase (Fluc) in the eNSCs were generated. In animals that received stroke, an increased BLI signal originating from the infarct region was observed. However, due to histological limitations, the identity and exact origin of cells contributing to the increased BLI signal could not be revealed. To overcome this limitation, we developed an alternative strategy employing stereotactic injection of conditional lentiviral vectors (Cre-Flex LVs) encoding Fluc and eGFP in the subventricular zone (SVZ) of Nestin-Cre transgenic mice, thereby specifically labeling the eNSCs. Upon induction of stroke, increased eNSC proliferation resulted in a significant increase in BLI signal between 2days and 2weeks after stroke, decreasing after 3months. Additionally, the BLI signal relocalized from the SVZ towards the infarct region during the 2weeks following stroke. Histological analysis at 90days post stroke showed that in the peri-infarct area, 36% of labeled eNSC progeny differentiated into astrocytes, while 21% differentiated into mature neurons. In conclusion, we developed and validated a novel imaging technique that unequivocally demonstrates that nestin(+) eNSCs originating from the SVZ respond to stroke injury by increased proliferation, migration towards the infarct region and differentiation into both astrocytes and neurons. In addition, this new approach allows non-invasive and specific monitoring of eNSCs over time, opening perspectives for preclinical evaluation of candidate stroke therapeutics.
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium; KU Leuven, Molecular Small Animal Imaging Center, MOSAIC, KU Leuven, 3000 Leuven, Flanders, Belgium; Division of Nuclear Medicine, University Hospital and KU Leuven, 3000 Leuven, Flanders, Belgium.
authors.forename: Caroline
authors.initials: C
authors.lastname: Vandeputte
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium.
authors.forename: Veerle
authors.initials: V
authors.lastname: Reumers
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium.
authors.forename: Sarah-Ann
authors.initials: SA
authors.lastname: Aelvoet
authors.affiliation: KU Leuven, Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, 3000 Leuven, Flanders, Belgium.
authors.forename: Irina
authors.initials: I
authors.lastname: Thiry
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium.
authors.forename: Sylvie
authors.initials: S
authors.lastname: De Swaef
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium; KU Leuven, Leuven Viral Vector Core, 3000 Leuven, Flanders, Belgium.
authors.forename: Chris
authors.initials: C
authors.lastname: Van den Haute
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium.
authors.forename: Jesus
authors.initials: J
authors.lastname: Pascual-Brazo
authors.affiliation: In-vivo-NMR Laboratory, Max Planck Institute for Neurological Research, 50931 Cologne, Germany.
authors.forename: Tracy D
authors.initials: TD
authors.lastname: Farr
authors.affiliation: KU Leuven, Molecular Small Animal Imaging Center, MOSAIC, KU Leuven, 3000 Leuven, Flanders, Belgium; KU Leuven, Biomedical MRI, Department of Imaging and Pathology, 3000 Leuven, Flanders, Belgium.
authors.forename: Greetje
authors.initials: G
authors.lastname: Vande Velde
authors.affiliation: In-vivo-NMR Laboratory, Max Planck Institute for Neurological Research, 50931 Cologne, Germany.
authors.forename: Mathias
authors.initials: M
authors.lastname: Hoehn
authors.affiliation: KU Leuven, Molecular Small Animal Imaging Center, MOSAIC, KU Leuven, 3000 Leuven, Flanders, Belgium; KU Leuven, Biomedical MRI, Department of Imaging and Pathology, 3000 Leuven, Flanders, Belgium.
authors.forename: Uwe
authors.initials: U
authors.lastname: Himmelreich
authors.affiliation: KU Leuven, Molecular Small Animal Imaging Center, MOSAIC, KU Leuven, 3000 Leuven, Flanders, Belgium; Division of Nuclear Medicine, University Hospital and KU Leuven, 3000 Leuven, Flanders, Belgium.
authors.forename: Koen
authors.initials: K
authors.lastname: Van Laere
authors.affiliation: KU Leuven, Molecular Small Animal Imaging Center, MOSAIC, KU Leuven, 3000 Leuven, Flanders, Belgium; KU Leuven, Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, 3000 Leuven, Flanders, Belgium.
authors.forename: Zeger
authors.initials: Z
authors.lastname: Debyser
authors.affiliation: KU Leuven, Laboratory for Molecular Virology and Gene Therapy, Department of Pharmaceutical and Pharmacological Sciences, 3000 Leuven, Flanders, Belgium; KU Leuven, Leuven Viral Vector Core, 3000 Leuven, Flanders, Belgium. Electronic address: rik.gijsbers@med.kuleuven.be.
authors.forename: Rik
authors.initials: R
authors.lastname: Gijsbers
authors.affiliation: KU Leuven, Laboratory for Neurobiology and Gene Therapy, Department of Neurosciences, 3000 Leuven, Flanders, Belgium; KU Leuven, Molecular Small Animal Imaging Center, MOSAIC, KU Leuven, 3000 Leuven, Flanders, Belgium. Electronic address: veerle.baekelandt@med.kuleuven.be.
authors.forename: Veerle
authors.initials: V
authors.lastname: Baekelandt
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000473:pathology / Q000502:physiology; D001921:Brain / Q000473:pathology / Q000503:physiopathology*; D002465:Cell Movement / Q000502:physiology; D018450:Disease Progression; D005500:Follow-Up Studies; D008163:Luminescent Measurements / Q000379:methods*; D008822:Mice, Transgenic; D058953:Neural Stem Cells / Q000473:pathology / Q000502:physiology*; D055495:Neurogenesis; D009474:Neurons / Q000473:pathology / Q000502:physiology; D061848:Optical Imaging / Q000379:methods*; D020521:Stroke / Q000473:pathology / Q000503:physiopathology*; D013997:Time Factors
keywords: Bioluminescence imaging; Cre-Flex lentiviral vector; Endogenous neural stem cells; Nestin-Cre mice; Stroke