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M.J. Kelley, K. Keller, J.R. Samples, T.S. Acott; Immunocharacterization of the Trabecular Meshwork Insert Region: The Search for a TM Stem Cell . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1871.
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© ARVO (1962-2015); The Authors (2016-present)
Previous work showed that trabecular meshwork (TM) cell division, which is increased in response to laser trabeculoplasty (LTP), occurs primarily in the anterior non–filtering portion of the TM, where it inserts into the cornea just below Schwalbe’s line. This is the "insert" region. After LTP, insert cells divide and migrate to repopulate the area of the LTP burns, suggesting that they are TM stem cells. To determine if insert cells can be distinguished from other TM cells, we subjected human eyes to LTP and investigated the expression of several potential cellular markers.
Potential biomarkers were either selected from previous work suggesting their possible value, such as Ankyrin G, Human Milk Fat Globule protein (HMFG or Breast Antigen 46), and YKL–40 (chitinase–3–like protein); or because they were elevated in TM microarray studies, such as CD44 and CD105. Standard clinical LTP treatment was applied to human anterior segments in stationary organ culture. Paraffin or cryostat sections were used to evaluate the potential markers by immunofluorescence and confocal microscopy. Parallel studies were conducted using TM cells in culture.
Positive immunostaining was observed for all of these antigens, but with different cellular locations and patterns. HMFG, a cell surface protein, and ankyrin G, a cellular protein, were localized only to the insert cells, while YKL–40(chitinase–3–like protein), an extracellular protein, and CD44 were found in both the insert and juxtacanalicular regions. CD44 appeared extracellularly as well as cellularly. CD105, which may label proliferating endothelial cells, was observed in endothelial cells lining Schlemm’s canal, as well as in a few cells in the insert region. Cell culture studies showed extensive staining with YKL–40, which was largely extracellular. However, in HMFG culture studies, only a very small percentage of cells showed a positive response.
Several biomarkers show possibilities for identifying or enriching putative TM stem cells. All of the potential biomarkers were found in the insert region, but Ankyrin G and HMFG appeared localized to insert cells alone, and may be useful for isolation of the cells. Biomarker use with insert cells may facilitate exploitation of their therapeutic potential.
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