Fibrillin-1 mutant mouse captures defining features of human primary open glaucoma including anomalous aqueous humor TGF beta-2.

Primary open angle glaucoma (POAG) features an optic neuropathy, elevated aqueous humor (AH) TGFβ2, and major risk factors of central corneal thickness (CCT), increasing age and intraocular pressure (IOP). We examined Tight skin (Tsk) mice to see if mutation of fibrillin-1, a repository for latent TGFβ, is associated with characteristics of human POAG. We measured: CCT by ocular coherence tomography (OCT); IOP; retinal ganglion cell (RGC) and optic nerve axon counts by microscopic techniques; visual electrophysiologic scotopic threshold responses (STR) and pattern electroretinogram (PERG); and AH TGFβ2 levels and activity by ELISA and MINK epithelial cell-based assays respectively.

Organogenesis and distribution of the ocular lymphatic vessels in the anterior eye.

Glaucoma surgeries, such as trabeculectomy, are performed to lower intraocular pressure to reduce risk of vision loss. These surgeries create a new passage in the eye that reroutes the aqueous humor outflow to the subconjunctival space, where the fluid is presumably absorbed by the conjunctival lymphatics. Here, we characterized the development and function of the ocular lymphatics using transgenic lymphatic reporter mice and rats.

Consensus recommendations for trabecular meshwork cell isolation, characterization and culture.

Cultured trabecular meshwork (TM) cells are a valuable model system to study the cellular mechanisms involved in the regulation of conventional outflow resistance and thus intraocular pressure; and their dysfunction resulting in ocular hypertension. In this review, we describe the standard procedures used for the isolation of TM cells from several animal species including humans, and the methods used to validate their identity. Having a set of standard practices for TM cells will increase the scientific rigor when used as a model, and enable other researchers to replicate and build upon previous findings.

Total Outflow Facility in Live C57BL/6 Mice of Different Age.

Purpose: To characterize total outflow facility across the live adult mouse lifespan as a reference for mouse glaucoma studies and the common C57BL/6 background strain.

Methods: Microperfusion was performed by single-needle cannulation and feedback-controlled coupling of pressure and flow to maintain a constant pressure in the anterior chambers of live C57BL/6NCrl mice aged 3-4 months ( = 17), 6-9 months ( = 10), and 23-27 months ( = 12). This mouse age range represented an equivalent human age range of young adult to elderly.

Reversible Venting Stitch for Fenestrating Valve-less Glaucoma Shunts.

Purpose: The purpose of this is to describe a venting stitch modification for valveless glaucoma aqueous shunts and characterize early postoperative intraocular pressure (IOP) and glaucoma medication use following the modification.

Materials And Methods: Retrospective chart review of 61 sequential patients undergoing Baerveldt glaucoma implant (BGI)-350 implantation at the Doheny Eye Institute. Twenty-four patients received a glaucoma shunt with venting stitch modification (modified BGI) and 37 patients received an unmodified shunt (BGI-only).

Pilot Study of Lamina Cribrosa Intensity Measurements in Glaucoma Using Swept-Source Optical Coherence Tomography.

Purpose: To compare the lamina cribrosa (LC) intensity in glaucoma-suspect eyes and eyes with mild to moderate glaucoma using swept-source optical coherence tomography.

Methods: Optic disc volume scans were collected using swept-source optical coherence tomography in 19 clinically defined glaucoma-suspect eyes and 29 eyes with mild to moderate glaucoma. LC intensity was measured using Image J software, and the resultant values were normalized using the retinal pigment epithelium and vitreous signal.

Aqueous Angiography with Fluorescein and Indocyanine Green in Bovine Eyes.

Purpose: We characterize aqueous angiography as a real-time aqueous humor outflow imaging (AHO) modality in cow eyes with two tracers of different molecular characteristics.

Methods: Cow enucleated eyes ( = 31) were obtained and perfused with balanced salt solution via a Lewicky AC maintainer through a 1-mm side-port. Fluorescein (2.

Aqueous Angiography-Mediated Guidance of Trabecular Bypass Improves Angiographic Outflow in Human Enucleated Eyes.

Purpose: To assess the ability of trabecular micro-bypass stents to improve aqueous humor outflow (AHO) in regions initially devoid of AHO as assessed by aqueous angiography.

Methods: Enucleated human eyes (14 total from 7 males and 3 females [ages 52-84]) were obtained from an eye bank within 48 hours of death. Eyes were oriented by inferior oblique insertion, and aqueous angiography was performed with indocyanine green (ICG; 0.

Neuropathic arthropathy of the elbow treated with double-plate arthrodesis and resection site bone graft.

Neuropathic arthropathy of the elbow is a rare condition, which is disabling and difficult to treat. Initial treatment is conservative and arthrodesis is rarely indicated. We describe an unusual case of progressive unilateral elbow swelling in a 37-year-old female domestic helper.

Optimizing two-photon multiple fluorophore imaging of the human trabecular meshwork.

Purpose: Advances in two-photon (2P) deep tissue imaging provide powerful options for simultaneously viewing multiple fluorophores within tissues. We determined imaging parameters for optimally visualizing three fluorophores in the human trabecular meshwork (TM) to simultaneously detect broad-spectrum autofluorescence and multiple fluorophores through a limited number of emission filters.

Methods: 2P imaging of viable human postmortem TM was conducted to detect Hoechst 33342-labeled nuclei, Alexa-568-conjugated phalloidin labeling of filamentous actin, and autofluorescence of the structural extracellular matrix (ECM).

Dose- and time-dependent effects of actomyosin inhibition on live mouse outflow resistance and aqueous drainage tissues.

Actomyosin contractility modulates outflow resistance of the aqueous drainage tissues and intraocular pressure, a key pathogenic factor of glaucoma. We established methodology to reliably analyze the effect of latrunculin-B (Lat-B)-induced actin depolymerization on outflow physiology in live mice. A voltage-controlled microperfusion system for delivering drugs and simultaneously analyzing outflow resistance was tested in live C57BL/6 mice.

Tissue-based multiphoton analysis of actomyosin and structural responses in human trabecular meshwork.

The contractile trabecular meshwork (TM) modulates aqueous humor outflow resistance and intraocular pressure. The primary goal was to visualize and quantify human TM contractile state by analyzing actin polymerization (F-actin) by 2-photon excitation fluorescence imaging (TPEF) in situ. A secondary goal was to ascertain if structural extracellular matrix (ECM) configuration changed with contractility.

Aqueous Angiography: Real-Time and Physiologic Aqueous Humor Outflow Imaging.

Purpose: Trabecular meshwork (TM) bypass surgeries attempt to enhance aqueous humor outflow (AHO) to lower intraocular pressure (IOP). While TM bypass results are promising, inconsistent success is seen. One hypothesis for this variability rests upon segmental (non-360 degrees uniform) AHO.

Endoscopic Cyclophotocoagulation and Pars Plana Ablation (ECP-plus) to Treat Refractory Glaucoma.

Purpose: To report clinical outcomes after pars plana endoscopic cyclophotocoagulation of the ciliary processes and pars plana (ECP-plus), a novel treatment for refractory glaucoma.

Design: Retrospective, noncomparative interventional case series.

Setting: multicenter tertiary referral academic and clinical practice.

Feedback-controlled constant-pressure anterior chamber perfusion in live mice.

Purpose: To describe live mouse, anterior chamber constant-pressure perfusion by an approach using feedback-controlled coupling of pressure and flow to maintain a preset pressure.

Methods: We established a microperfusion system that maintains a constant preset pressure in the anterior chamber of live mice by automatically regulating the microsyringe pump flow rate with a computer-controlled voltage feedback loop. Perfusion was by single-needle cannulation.

Tissue-based imaging model of human trabecular meshwork.

We have developed a tissue-based model of the human trabecular meshwork (TM) using viable postmortem corneoscleral donor tissue. Two-photon microscopy is used to optically section and image deep in the tissue to analyze cells and extracellular matrix (ECM) within the original three-dimensional (3D) environment of the TM. Multimodal techniques, including autofluorescence (AF), second harmonic generation (SHG), intravital dye fluorescence, and epifluorescence, are combined to provide unique views of the tissue at the cellular and subcellular level.

Contractile markers distinguish structures of the mouse aqueous drainage tract.

Purpose: Structures of the aqueous humor drainage tract are contractile, although the tract is not entirely composed of muscle. We characterized the mouse aqueous drainage tract by immunolabeling contractile markers and determined whether profiling these markers within the tract distinguished its key structures of the trabecular meshwork (TM) and ciliary muscle (CM).

Methods: Enucleated eyes from pigmented C57BL/6 (n=8 mice) and albino BALB/c (n=6 mice) mice were processed for cryo- and formalin-fixed paraffin-embedded sectioning.

Sources of structural autofluorescence in the human trabecular meshwork.

Purpose: In situ 2-photon excitation fluorescence (TPEF) of the human trabecular meshwork (TM) reveals beams of heterogeneous autofluorescence (AF) comprising high intensity fluorescent fibers (AF-high) on a background of lower intensity fluorescence (AF-low). To determine the sources of this AF heterogeneity, we imaged human TM to characterize AF, second harmonic generation (SHG) for collagen, and eosin-labeled fluorescence identifying elastin.

Methods: Corneoscleral rims retained after corneal transplantation were incubated with and without eosin, and imaged by TPEF.

Analyzing live cellularity in the human trabecular meshwork.

Purpose: To directly visualize the live cellularity of the intact human trabecular meshwork (TM) and quantitatively analyze tissue viability in situ.

Methods: Human donor corneoscleral rims were sectioned immediately before intravital dye incubation to label nuclei (Hoechst 33342 & propidium iodide [PI]); cytosol (CellTracker Red CMTPX, calcein AM); and membranes (octadecyl rhodamine B chloride [R18]), followed by 2-photon microscopy. Viability was assessed by counting cells in tissue colabeled with PI and Calcein AM.

Minimally invasive helical plating for shaft of humerus fractures: technique and outcome.

Introduction: The humerus is subjected to substantial amount of torsional stress. Conventional plating may not address this sufficiently and may lead to fixation failure or non-union. A helical plate may offer the solution.

Two-photon immunofluorescence characterization of the trabecular meshwork in situ.

Purpose: To develop an in situ model to study biological responses and glaucoma pathology in the human trabecular meshwork (TM). Characteristic TM cell- and glaucoma-associated markers were localized in situ in relation to the tissue's autofluorescent structural extracellular matrix (ECM) by two-photon excitation fluorescence optical sectioning (TPEF).

Methods: Human donor corneoscleral (CS) tissue containing the intact aqueous drainage tract was incubated with dexamethasone (Dex) or TGF-β1, and immunostained for epifluorescence (EF) microscopy with antibodies to myocilin and alpha smooth muscle (α-SMA).