title: A magnetic bead-based method for mouse trabecular meshwork cell isolation.
abstract: OBJECTIVE
  Mice have been used widely for glaucoma research. However, due to the small size of the mouse eye, it is difficult to dissect mouse trabecular meshwork (MTM) tissues and establish MTM cell strains. To circumvent this problem, we took advantage of the phagocytic property of trabecular meshwork (TM) cells, and developed a novel magnetic bead-based method that enables us to isolate pure MTM cells.
  
  
  METHODS
  After anesthesia, up to 2 μL of fluorescent or magnetic microbeads were injected intracamerally into the mouse eyes. To study the distribution and localization of the beads, mice were sacrificed 1 to 7 days after injection, and eyes were enucleated for fluorescent or transmission electron microscopy (TEM) study, respectively. To isolate MTM cells, anterior segments injected with magnetic beads were dissected from 10 to 15 sterilized mouse eyes 7 days after injection. The tissues were digested with collagenase A and purified by using a magnetic field as well as repeated washing.
  
  
  RESULTS
  TEM studies showed that the magnetic beads were located in the mouse TM, but not in corneal or scleral fibroblast cells. Cultured MTM cells were similar morphologically to human TM cells. MTM cells expressed TM markers, including collagen IV, laminin, and α-smooth muscle actin. Also, MTM cells treated with 100 nM dexamethasone showed increased formation of cross-linked actin networks and induction of myocilin expression.
  
  
  CONCLUSIONS
  The magnetic bead-based method is efficient for isolating MTM cells with minimal microdissection techniques required. It will be a useful approach for isolating TM cells from small animals for glaucoma research.
authors: Department of Cell Biology and Anatomy, North Texas Eye Research Institute, University of North Texas Health Science Center, Fort Worth, Texas 76107, USA. weiming.mao@unthsc.edu
authors: Weiming
authors: W
authors: Mao
authors: Yang
authors: Y
authors: Liu
authors: Robert J
authors: RJ
authors: Wordinger
authors: Abbot F
authors: AF
authors: Clark
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D000869:Anterior Eye Segment / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D015415:Biomarkers / Q000378:metabolism; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D024141:Collagen Type IV / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism; D004195:Disease Models, Animal; D005136:Eye Proteins / Q000378:metabolism; D005260:Female; D005453:Fluorescence; D005901:Glaucoma / Q000378:metabolism / Q000473:pathology*; D006023:Glycoproteins / Q000378:metabolism; D018189:Immunomagnetic Separation / Q000379:methods*; D007797:Laminin / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D046529:Microscopy, Electron, Transmission; D014129:Trabecular Meshwork / Q000166:cytology* / Q000378:metabolism / Q000648:ultrastructure; D000099179:Myocilin
chemical_list: D000199:Actins; D015415:Biomarkers; D024141:Collagen Type IV; D003598:Cytoskeletal Proteins; D005136:Eye Proteins; D006023:Glycoproteins; D007797:Laminin; D000099179:Myocilin
keywords: cell isolation; glaucoma; magnetic beads; mouse models; trabecular meshwork
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