In Situ Autofluorescence Visualization of Human Trabecular Meshwork Structure

Abstract Background: Elevated intraocular pressure (IOP) is a major risk factor for the development and progression of primary open angle glaucoma and is due to trabecular meshwork (TM) damage. Here, we investigate the role of an endogenous Toll-like receptor 4 (TLR4) ligand, FN-EDA, in the development of glaucoma utilizing a transgenic mouse strain (B6.EDA +/+ ) that constitutively expresses only FN containing the EDA isoform. Methods: Eyes from C57BL6/J (wild-type), B6.EDA +/+ (constitutively active EDA), B6.EDA -/- (EDA null) mice were processed for electron microscopy and consecutive images of the entire length of the TM and Schlemm’s canal (SC) from anterior to posterior were collected and montaged into a single image. ECM accumulation, basement membrane length, and size and number of giant vacuoles were quantified by ImageJ analysis. Tlr4 and Iba1 expression in the TM and ONH cells was conducted using RNAscope in situ hybridization and immunohistochemistry protocols. IOP was measured using a rebound tonometer, ON damage assessed by PPD stain, and RGC loss quantified in RBPMS labeled retina flat mounts. Results: Ultrastructure analyses show the TM of B6.EDA +/+ mice have significantly increased accumulation of ECM between TM beams with few empty spaces compared to C57BL/6J mice (P<0.05). SC basement membrane is thicker and more continuous in B6.EDA +/+ mice compared to C57BL/6J. No significant structural differences are detected in the TM of EDA null mice. Tlr4 and Iba1 expression is increased in the TM of B6.EDA +/+ mice compared to C57BL/6J eyes (p<0.05). IOP is significantly higher in B6.EDA +/+ mice compared to C57BL/6J eyes (p<0.001), and significant ON damage (p<0.001) and RGC loss (p<0.05) detected at one year of age. Tlr4 mRNA is expressed in mouse ONH cells, and is present in ganglion cell axons, microglia, and astrocytes. There is a significant increase in the area occupied by Iba-1 positive microglia cells in the ONH of B6.EDA +/+ mice compared to C57BL/6J control eyes (p<0.01). Conclusions: B6.EDA +/+ mice have increased ECM accumulation in the TM, elevated IOP, enhanced proinflammatory changes in the ONH, loss of RGCs, and ONH damage. These data suggest B6.EDA +/+ mice recapitulate many aspects of glaucomatous damage.

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Functional Implications of Cross-Linked Actin Networks in Trabecular Meshwork Cells

Cellular Physiology and Biochemistry ◽

10.1159/000487170 ◽

2018 ◽

Vol 45 (2) ◽

pp. 783-794 ◽

Cited By ~ 6

Author(s):

Laura Duffy ◽

Steven O’Reilly

Keyword(s):

Trabecular Meshwork ◽

Open Angle Glaucoma ◽

Collagen Gel ◽

Mechanical Stretch ◽

Actin Networks ◽

Trabecular Meshwork Cells ◽

Microscopy Technique ◽

Collagen Gel Contraction

Background/Aims: The Trabecular meshwork (TM) is the tissue responsible for outflow resistance and therefore intraocular pressure. TM cells contain a contractile apparatus that is composed of actin stress fibres which run parallel to the axis of the cell and are responsible for facilitating contraction. Cross-Linked Actin Networks (CLANs) are polygonal arrangements of actin that form a geodesic network found predominantly in TM cells both in situ and in vitro. The aim of this work is to determine the functional significance of CLANs in TM cells and to assess the effect of mechanical stretch stimulation on the induction (or not) of CLANs. Methods: We used collagen gel contraction models to demonstrate functional impairment of cells when induced to express CLANs in situ. Cyclic mechanical stretch was used to stimulate cells and measure CLANs Results: CLANs inhibited contraction and cyclic mechanical stretch induced CLANs. Furthermore, we also demonstrated that using shape alone we could predict the appearance of CLANs using a simple light microscopy technique. Conclusion: Taken together we have now shown, for the first time, a functional deficit In TM cells with CLANs Furthermore that shape alone can predict the appearance of CLAN containing cells. CLANs can now be linked to a functional effect and may underlie the appearance of CLANs with the pathology of primary open angle glaucoma (POAG).

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Rendez vous with Saturn's rings

International Astronomical Union Colloquium ◽

10.1017/s0252921100101885 ◽

1984 ◽

Vol 75 ◽

pp. 743-759 ◽

Cited By ~ 2

Author(s):

Kerry T. Nock

Keyword(s):

Solar Energy ◽

Electric Propulsion ◽

Power Source ◽

Propulsion System ◽

Low Thrust ◽

Ring Plane ◽

The Earth ◽

Total Impulse ◽

Saturn's Rings

ABSTRACTA mission to rendezvous with the rings of Saturn is studied with regard to science rationale and instrumentation and engineering feasibility and design. Future detailedin situexploration of the rings of Saturn will require spacecraft systems with enormous propulsive capability. NASA is currently studying the critical technologies for just such a system, called Nuclear Electric Propulsion (NEP). Electric propulsion is the only technology which can effectively provide the required total impulse for this demanding mission. Furthermore, the power source must be nuclear because the solar energy reaching Saturn is only 1% of that at the Earth. An important aspect of this mission is the ability of the low thrust propulsion system to continuously boost the spacecraft above the ring plane as it spirals in toward Saturn, thus enabling scientific measurements of ring particles from only a few kilometers.

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