Daniel Brotchie

Publications (2)7.62 Total impact

• Article: Cross-linked actin networks (CLANs) in the trabecular meshwork of the normal and glaucomatous human eye in situ.

  Mary-Jo Hoare, Ian Grierson, Daniel Brotchie, Natalie Pollock, Kathy Cracknell, Abbot F Clark
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  ABSTRACT: A percentage of trabecular meshwork (TM) cells in tissue and organ culture have been shown to contain cross-linked actin networks (CLANs) when exposed to dexamethasone, as have TM cultures derived from glaucomatous individuals. The purpose of this study was to determine whether CLANs exist within TM cells in situ in tissue unmanipulated by culturing, thereby eliminating the possibility that CLANs are artifacts of culture conditions, and to determine their numbers and dimensions in normal and glaucoma TM cells. Twelve human donor eyes (five normal and seven with glaucoma) provided the TM tissue. Each eye was dissected, and the TM tissue was exposed either by microdissection (qualitative studies) or cryo-sectioning (quantitative analysis). The actin cytoskeleton was visualized using a high-affinity probe and viewed using confocal microscopy. Qualitative examination of the microdissected tissue revealed that CLANs and CLAN-like structures were a common finding in the TM cells in every specimen, irrespective of whether they were from normal or glaucomatous eyes. CLAN size and phenotype were variable, with the same variations occurring in both normal and glaucomatous eyes. Quantitative analysis showed that there were more CLANs in glaucoma TM specimens than normal TM specimens, but this difference was not statistically significant. The mean number of CLANs/TM cell in our glaucoma tissue was estimated to be 1.03, while in the elderly normal controls it was 0.67. This study showed for the first time that CLANs exist in cells of TM tissues from both normal and glaucomatous eyes that have not been manipulated by either tissue or organ culture procedures. It also provides quantitative data on CLAN prevalence in organized TM tissue, which indicates that CLANs are far more common than predicted (even from tissue culture) and there may be one in every cell in the glaucomatous TM in situ.
  Investigative ophthalmology & visual science 11/2008; 50(3):1255-63. · 3.43 Impact Factor
• Article: Dexamethasone alters F-actin architecture and promotes cross-linked actin network formation in human trabecular meshwork tissue.

  Abbot F Clark, Daniel Brotchie, A Thomas Read, Peggy Hellberg, Sherry English-Wright, Iok-Hou Pang, C Ross Ethier, Ian Grierson
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  ABSTRACT: Elevated intraocular pressure is an important risk factor for the development of glaucoma, a leading cause of irreversible blindness. This ocular hypertension is due to increased hydrodynamic resistance to the drainage of aqueous humor through specialized outflow tissues, including the trabecular meshwork (TM) and the endothelial lining of Schlemm's canal. We know that glucocorticoid therapy can cause increased outflow resistance and glaucoma in susceptible individuals, that the cytoskeleton helps regulate aqueous outflow resistance, and that glucocorticoid treatment alters the actin cytoskeleton of cultured TM cells. Our purpose was to characterize the actin cytoskeleton of cells in outflow pathway tissues in situ, to characterize changes in the cytoskeleton due to dexamethasone treatment in situ, and to compare these with changes observed in cell culture. Human ocular anterior segments were perfused with or without 10(-7) M dexamethasone, and F-actin architecture was investigated by confocal laser scanning microscopy. We found that outflow pathway cells contained stress fibers, peripheral actin staining, and occasional actin "tangles." Dexamethasone treatment caused elevated IOP in several eyes and increased overall actin staining, with more actin tangles and the formation of cross-linked actin networks (CLANs). The actin architecture in TM tissues was remarkably similar to that seen in cultured TM cells. Although CLANs have been reported previously in cultured cells, this is the first report of CLANs in tissue. These cytoskeletal changes may be associated with increased aqueous humor outflow resistance after ocular glucocorticoid treatment.
  Cell Motility and the Cytoskeleton 03/2005; 60(2):83-95. · 4.19 Impact Factor