title: Neurotrophic and neuroprotective potential of human limbus-derived mesenchymal stromal cells.
abstract: OBJECTIVE
  The purpose of this study was to examine neurotrophic and neuroprotective effects of limbus stroma-derived mesenchymal stromal cells (L-MSCs) on cortical neurons in vitro and in vivo.
  
  
  METHODS
  Cultured L-MSCs were characterized by flow cytometry and immunofluorescence through the use of specific MSC marker antibodies. Conditioned media were collected from normoxia- and hypoxia-treated L-MSCs to assess neurotrophic effects. Neuroprotective potentials were evaluated through the use of in vitro hypoxic cortical neuron culture and in vivo rat focal cerebral ischemia models. Neuronal morphology was confirmed by immunofluorescence with the use of anti-MAP2 antibody. Post-ischemic infarct volume and motor behavior were assayed by means of triphenyltetrazolium chloride staining and open-field testing, respectively. Human growth antibody arrays and enzyme-linked immunoassays were used to analyze trophic/growth factors contained in conditioned media.
  
  
  RESULTS
  Isolated human L-MSCs highly expressed CD29, CD90 and CD105 but not CD34 and CD45. Mesenchymal lineage cell surface expression pattern and differentiation capacity were identical to MSCs derived form human bone marrow and adipose tissue. The L-MSC normoxic and hypoxic conditioned media both promoted neurite outgrowth in cultured cortical neurons. Hypoxic conditioned medium showed superior neurotrophic function and neuroprotective potential with reduced ischemic brain injury and improved functional recovery in rat focal cerebral ischemia models. Human growth factor arrays and enzyme-linked immunoassays measurements showed neuroprotective and growth-associated cytokines (vascular endothelial growth factor [VEGF], VEGFR3, brain-derived neurotrophic factor, insulin-like growth factor -2 and hepatocyte growth factor) contained in conditioned media. Hypoxic exposure caused VEGF and brain-derived neurotrophic factor upregulation, possibly contributing to neurotrophic and neuroprotective effects.
  
  
  CONCLUSIONS
  L-MSCs can secrete various neurotrophic factors stimulating neurite outgrowth and protecting neurons against brain ischemic injury through paracrine mechanism.
authors.affiliation: Department of Ophthalmology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan; Graduate Institute of Aerospace and Undersea Medicine, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Chang-Min
authors.initials: CM
authors.lastname: Liang
authors.affiliation: Department of Biology and Anatomy, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Shao-Ju
authors.initials: SJ
authors.lastname: Weng
authors.affiliation: Department of Neurosurgery, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Tung-Han
authors.initials: TH
authors.lastname: Tsai
authors.affiliation: Department of Pharmacy Practice, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: I-Hsun
authors.initials: IH
authors.lastname: Li
authors.affiliation: Institute of Medicine, Chung Shan Medical University, Taichung, Taiwan.
authors.forename: Pin-Hui
authors.initials: PH
authors.lastname: Lu
authors.affiliation: Department of Biology and Anatomy, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Kuo-Hsing
authors.initials: KH
authors.lastname: Ma
authors.affiliation: Department of Ophthalmology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Ming-Cheng
authors.initials: MC
authors.lastname: Tai
authors.affiliation: Department of Ophthalmology, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Jiann-Torng
authors.initials: JT
authors.lastname: Chen
authors.affiliation: Department of Nuclear Medicine, Tri-Service General Hospital, National Defense Medical Center, Taipei, Taiwan.
authors.forename: Cheng-Yi
authors.initials: CY
authors.lastname: Cheng
authors.affiliation: Department of Biology and Anatomy, National Defense Medical Center, Taipei, Taiwan. Electronic address: anatoman2001@yahoo.com.tw.
authors.forename: Yuahn-Sieh
authors.initials: YS
authors.lastname: Huang
mesh_terms: D000818:Animals; D002545:Brain Ischemia / Q000628:therapy*; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D017077:Culture Media, Conditioned / Q000494:pharmacology; D006801:Humans; D016850:Limbus Corneae / Q000166:cytology*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism*; D009414:Nerve Growth Factors / Q000378:metabolism / Q000494:pharmacology; D055495:Neurogenesis; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D018696:Neuroprotective Agents / Q000378:metabolism / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
chemical_list: D017077:Culture Media, Conditioned; D009414:Nerve Growth Factors; D018696:Neuroprotective Agents; D042461:Vascular Endothelial Growth Factor A
keywords: mesenchymal stromal cells; neuroprotection; paracrine