Morphological comparison of astrocytes in the lamina cribrosa and glial lamina.

Purpose: Although the mechanisms underlying glaucomatous neurodegeneration are not yet well understood, cellular and small animal models suggest that LC astrocytes undergo early morphologic and functional changes, indicating their role as early responders to glaucomatous stress. These models, however, lack the LC found in larger animals and humans, leaving the morphology of LC astrocytes and their role in glaucoma initiation underexplored. In this work, we aimed to characterize the morphology of LC astrocytes and determine differences and similarities with astrocytes in the mouse glial lamina (GL), the analogous structure in a prominent glaucoma model.

Direct measurements of collagen fiber recruitment in the posterior pole of the eye.

Collagen is the main load-bearing component of the peripapillary sclera (PPS) and lamina cribrosa (LC) in the eye. Whilst it has been shown that uncrimping and recruitment of the PPS and LC collagen fibers underlies the macro-scale nonlinear stiffening of both tissues with increased intraocular pressure (IOP), the uncrimping and recruitment as a function of local stretch have not been directly measured. This knowledge is crucial to understanding their functions in bearing loads and maintaining tissue integrity.

Direct measurements of collagen fiber recruitment in the posterior pole of the eye.

Collagen is the main load-bearing component of the peripapillary sclera (PPS) and lamina cribrosa (LC) in the eye. Whilst it has been shown that uncrimping and recruitment of the PPS and LC collagen fibers underlies the macro-scale nonlinear stiffening of both tissues with increased intraocular pressure (IOP), the uncrimping and recruitment as a function of local stretch have not been directly measured. This knowledge is crucial for the development of constitutive models associating micro and macro scales.

Individual astrocyte morphology in the collagenous lamina cribrosa revealed by multicolor DiOlistic labeling.

Astrocytes in the lamina region of the optic nerve head play vital roles in supporting retinal ganglion cell axon health. In glaucoma, these astrocytes are implicated as early responders to stressors, undergoing characteristic changes in cell function as well as cell morphology. Much of what is currently known about individual lamina astrocyte morphology has been learned from rodent models which lack a defining feature of the human optic nerve head, the collagenous lamina cribrosa (LC).

In vivo Modulation of Intraocular and Intracranial Pressures Causes Nonlinear and Non-monotonic Deformations of the Lamina Cribrosa and Scleral Canal.

Purpose: To evaluate changes in monkey optic nerve head (ONH) morphology under acutely controlled intraocular pressure (IOP) and intracranial pressure (ICP).

Methods: Seven ONHs from six monkeys were imaged via optical coherence tomography while IOP and ICP were maintained at one of 16 conditions. These conditions were defined by 4 levels for each pressure: low, baseline, high and very high.

Tissue-engineered anterior segment eye cultures demonstrate hallmarks of conventional organ culture.

Background: Glaucoma is a blinding disease largely caused by dysregulation of outflow through the trabecular meshwork (TM), resulting in elevated intraocular pressure (IOP). We hypothesized that transplanting TM cells into a decellularized, tissue-engineered anterior segment eye culture could restore the outflow structure and function.

Methods: Porcine eyes were decellularized with freeze-thaw cycles and perfusion of surfactant.

Real-time imaging of optic nerve head collagen microstructure and biomechanics using instant polarized light microscopy.

Current tools lack the temporal or spatial resolution necessary to image many important aspects of the architecture and dynamics of the optic nerve head (ONH). We evaluated the potential of instant polarized light microscopy (IPOL) to overcome these limitations by leveraging the ability to capture collagen fiber orientation and density in a single image. Coronal sections through the ONH of fresh normal sheep eyes were imaged using IPOL while they were stretched using custom uniaxial or biaxial micro-stretch devices.

Lamina cribrosa vessel and collagen beam networks are distinct.

Our goal was to analyze the spatial interrelation between vascular and collagen networks in the lamina cribrosa (LC). Specifically, we quantified the percentages of collagen beams with/without vessels and of vessels inside/outside of collagen beams. To do this, the vasculature of six normal monkey eyes was labeled by perfusion post-mortem.

Interplay between intraocular and intracranial pressure effects on the optic nerve head in vivo.

Intracranial pressure (ICP) has been proposed to play an important role in the sensitivity to intraocular pressure (IOP) and susceptibility to glaucoma. However, the in vivo effects of simultaneous, controlled, acute variations in ICP and IOP have not been directly measured. We quantified the deformations of the anterior lamina cribrosa (ALC) and scleral canal at Bruch's membrane opening (BMO) under acute elevation of IOP and/or ICP.

So-Called Lamina Cribrosa Defects May Mitigate IOP-Induced Neural Tissue Insult.

Purpose: The prevailing theory about the function of lamina cribrosa (LC) connective tissues is that they provide structural support to adjacent neural tissues. Missing connective tissues would compromise this support and therefore are regarded as "LC defects", despite scarce actual evidence of their role. We examined how so-called LC defects alter IOP-related mechanical insult to the LC neural tissues.

Instant polarized light microscopy for imaging collagen microarchitecture and dynamics.

Collagen fibers are a primary load-bearing component of connective tissues and are therefore central to tissue biomechanics and pathophysiology. Understanding collagen architecture and behavior under dynamic loading requires a quantitative imaging technique with simultaneously high spatial and temporal resolutions. Suitable techniques are thus rare and often inaccessible.

Ripasudil in a Model of Pigmentary Glaucoma.

Purpose: To investigate the effects of Ripasudil (K-115), a Rho-kinase inhibitor, in a porcine model of pigmentary glaucoma.

Methods: In vitro trabecular meshwork (TM) cells and ex vivo perfused eyes were subjected to pigment dispersion followed by K-115 treatment (PK115). PK115 was compared to controls (C) and pigment (P).

Lamina Cribrosa Capillaries Straighten as Intraocular Pressure Increases.

Purpose: The purpose of this study was to visualize the lamina cribrosa (LC) capillaries and collagenous beams, measure capillary tortuosity (path length over straight end-to-end length), and determine if capillary tortuosity changes when intraocular pressure (IOP) increases.

Methods: Within 8 hours of sacrifice, 3 pig heads were cannulated via the external ophthalmic artery, perfused with PBS to remove blood, and then perfused with a fluorescent dye to label the capillaries. The posterior pole of each eye was mounted in a custom-made inflation chamber for control of IOP with simultaneous imaging.

Role of radially aligned scleral collagen fibers in optic nerve head biomechanics.

Collagen fibers organized circumferentially around the canal in the peripapillary sclera are thought to provide biomechanical support to the sensitive tissues within the optic nerve head (ONH). Recent studies have demonstrated the existence of a family of fibers in the innermost sclera organized radially from the scleral canal. Our goal was to determine the role of these radial fibers in the sensitivity of scleral canal biomechanics to acute increases in intraocular pressure (IOP).

Dose-dependent effects of netarsudil, a Rho-kinase inhibitor, on the distal outflow tract.

Purpose: To characterize the effects of netarsudil on the aqueous humor outflow tract distal to the trabecular meshwork (TM). We hypothesized that netarsudil increases outflow facility in eyes with and without circumferential ab interno trabeculectomy (AIT) that removes the TM.

Methods: Sixty-four porcine anterior segment cultures were randomly assigned to groups with (n = 32) and without circumferential AIT (n = 32).

Intraocular pressure reduction in a pigmentary glaucoma model by Goniotome Ab interno trabeculectomy.

Purpose: To investigate whether microsurgical excision of trabecular meshwork (TM) in an ex vivo pigmentary glaucoma model can normalize the hypertensive phenotype.

Methods: Eight eyes of a porcine pigmentary glaucoma model underwent 90° of microsurgical TM excision with an aspirating dual-blade (Goniotome (G)). 24 hours later, additional 90° of TM were removed.

Outflow facility and extent of angle closure in a porcine model.

Purpose: To establish the extent of anterior chamber angle circumference needed to maintain a physiological outflow facility (C). This could create a model to investigate focal outflow regulation.

Methods: Twenty anterior segments of porcine eyes were assigned to five groups, each with a different degree of cyanoacrylate-mediated angle closure: 90° (n = 4), 180° (n = 4), 270° (n = 4), 360° (n = 4), and four unoccluded control eyes.

Impact of pigment dispersion on trabecular meshwork cells.

Purpose: Dysfunction of the trabecular meshwork (TM) in pigmentary glaucoma contributes to increased aqueous humor outflow resistance and intraocular pressure. In this study, we investigated the effect of pigment dispersion on trabecular meshwork cells.

Methods: Porcine TM cells from ab interno trabeculectomy specimens were exposed to pigment dispersion, then, analyzed for changes in morphology, immunostaining, and ultrastructure.

RKI-1447, a Rho kinase inhibitor, causes ocular hypotension, actin stress fiber disruption, and increased phagocytosis.

Purpose: This study investigated the hypotensive effect of RKI-1447, a Rho kinase inhibitor, in a porcine ex vivo pigmentary glaucoma model.

Methods: Twenty-eight porcine anterior chambers were perfused with medium supplemented with 1.67 × 10 pigment particles/ml for 48 h before treatment with RKI-1447 (n = 16) or vehicle control (n = 12).

Structure-Function Changes of the Porcine Distal Outflow Tract in Response to Nitric Oxide.

Purpose: To correlate outflow function and outflow tract vessel diameter changes induced by nitric oxide (NO).

Methods: In a porcine anterior segment perfusion model, the effects of a nitric oxide donor (100 μM DETA-NO) on outflow facility were compared with controls (n = 8 per group) with trabecular meshwork (TM) and after circumferential ab interno trabeculectomy (AIT). Outflow structures were assessed with spectral-domain optical coherence tomography (SD-OCT) before and after NO, or an NO synthase inhibitor (100 μM L-NAME) and the vasoconstrictor, endothelin-1 (100 pg/mL ET-1).

High-Resolution, Three-Dimensional Reconstruction of the Outflow Tract Demonstrates Segmental Differences in Cleared Eyes.

Purpose: The rate of conventional aqueous humor outflow is the highest nasally. We hypothesized that this is reflected in regionally different outflow structures and analyzed the entire limbus by high-resolution, full-thickness ribbon-scanning confocal microscopy (RSCM).

Methods: We perfused pig eyes by anterior chamber cannulation with eight lectin-fluorophore conjugates, followed by optical clearance with benzyl alcohol benzyl benzoate (BABB).

Intraocular pressure elevation precedes a phagocytosis decline in a model of pigmentary glaucoma.

Outflow regulation and phagocytosis are key functions of the trabecular meshwork (TM), but it is not clear how the two are related in secondary open angle glaucomas characterized by an increased particle load. We hypothesized that diminished TM phagocytosis is not the primary cause of early ocular hypertension and recreated pigment dispersion in a porcine model. Sixteen porcine anterior chamber cultures received a continuous infusion of pigment granules (Pg), while 16 additional anterior chambers served as controls (C).

Outflow enhancement by three different ab interno trabeculectomy procedures in a porcine anterior segment model.

Purpose: To evaluate three different microincisional ab interno trabeculectomy procedures in a porcine eye perfusion model.

Methods: In perfused porcine anterior segments, 90° of trabecular meshwork (TM) was ablated using the Trabectome (T; n = 8), Goniotome (G; n = 8), or Kahook device (K; n = 8). After 24 h, additional 90° of TM was removed.