A Noninvasive Clinical Method to Measure in Vivo Mechanical Strains of the Lamina Cribrosa by OCT.

Objective: To measure mechanical strain of the lamina cribrosa (LC) after intraocular pressure (IOP) change produced 1 week after a change in glaucoma medication. Design: Cohort study. Participants: Adult glaucoma patients (23 eyes, 15 patients) prescribed a change in IOP-lowering medication. Intervention: Noninvasive OCT imaging of the eye. Main Outcome Measures: Deformation calculated by digital volume correlation of OCT scans of the LC before and after IOP lowering by medication. Results: Among 23 eyes, 17 eyes of 12 persons had IOP lowering ≥ 3 mmHg (reduced IOP group) with tensile anterior-posterior Ezz strain = 1.0% ± 1.1% (P = 0.003) and compressive radial strain (Err) = -0.3% ± 0.5% (P = 0.012; random effects models accounting inclusion of both eyes in some persons). Maximum in-plane principal (tensile) strain and maximum shear strain in the reduced-IOP group were as follows: Emax = 1.7% ± 1.0% and Γmax = 1.4% ± 0.7%, respectively (both P < 0.0001 vs. zero). Reduced-IOP group strains Emax and Γmax were significantly larger with greater % IOP decrease (P < 0.0001 and P < 0.0001, respectively). The compliances of the Ezz, Emax, and Γmax strain responses, defined as strain normalized by the IOP decrease, were larger with more abnormal perimetric mean deviation or visual field index values (all P ≤ 0.02). Strains were unrelated to age (all P ≥ 0.088). In reduced-IOP eyes, mean LC anterior border posterior movement was only 2.05 µm posteriorly (P = 0.052) and not related to % IOP change (P = 0.94, random effects models). Only Err was significantly related to anterior lamina depth change, becoming more negative with greater posterior LC border change (P = 0.015). Conclusions: Lamina cribrosa mechanical strains can be effectively measured by changes in eye drop medication using OCT and are related to degree of visual function loss in glaucoma. Financial Disclosures: Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Long-term Remodeling Response in the Lamina Cribrosa Years after Intraocular Pressure Lowering by Suturelysis after Trabeculectomy.

OBJECTIVE: To measure the remodeling of the lamina cribrosa (LC) years after intraocular pressure (IOP) lowering by suturelysis. DESIGN: Cohort study. PARTICIPANTS: Glaucoma patients were imaged 20 minutes after laser suturelysis after trabeculectomy surgery and at their follow-up appointment 1 to 4 years later (16 image pairs; 15 persons). INTERVENTION: Noninvasive OCT imaging of the eye. MAIN OUTCOME MEASURES: Deformation calculated by correlating OCT scans of the LC immediately after IOP lowering by suturelysis and those acquired years later (defined as remodeling strain). RESULTS: The LC anterior border moved 60.9 ± 54.6 µm into the eye (P = 0.0007), and the LC exhibited regions of large local stretch in the anterior-posterior direction on long-term, maintained IOP lowering, resulting in a mean anterior-posterior remodeling strain of 14.0% ± 21.3% (P = 0.02). This strain and the LC border movement was 14 times and 124 times larger, respectively, than the direct response to IOP lowering by suturelysis. A larger anterior LC border movement was associated with greater mean anterior-posterior remodeling strain (P = 0.004). A thinner retinal nerve fiber layer at suturelysis was also associated with greater mean anterior-posterior remodeling strain at follow-up (P = 0.05). Worsening visual field indexes during follow-up were associated with a greater mean circumferential remodeling strain (P = 0.02), due to regions of large local circumferential stretch of the LC. Eyes with a more compliant LC torsional shear strain response at lysis were associated with worse mean deviation at follow-up (P = 0.03). CONCLUSIONS: Strains and LC border position changes measured years after IOP lowering are far larger than the immediate response to IOP lowering and indicate dramatic remodeling of the LC anatomical structure caused by IOP lowering and glaucoma progression. The remodeling strains indicate substantial local stretch in the anterior-posterior direction and are associated with movement of the LC anterior border into the eye. Eyes with greater direct strain response to IOP lowering, greater glaucoma damage at suturelysis, and greater worsening of visual field at follow-up experienced greater remodeling. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03267849. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

IOP and glaucoma damage: The essential role of optic nerve head and retinal mechanosensors.

There are many unanswered questions on the relation of intraocular pressure to glaucoma development and progression. IOP itself cannot be distilled to a single, unifying value, because IOP level varies over time, differs depending on ocular location, and can be affected by method of measurement. Ultimately, IOP level creates mechanical strain that affects axonal function at the optic nerve head which causes local extracellular matrix remodeling and retinal ganglion cell death - hallmarks of glaucoma and the cause of glaucomatous vision loss. Extracellular tissue strain at the ONH and lamina cribrosa is regionally variable and differs in magnitude and location between healthy and glaucomatous eyes. The ultimate targets of IOP-induced tissue strain in glaucoma are retinal ganglion cell axons at the optic nerve head and the cells that support axonal function (astrocytes, the neurovascular unit, microglia, and fibroblasts). These cells sense tissue strain through a series of signals that originate at the cell membrane and alter cytoskeletal organization, migration, differentiation, gene transcription, and proliferation. The proteins that translate mechanical stimuli into molecular signals act as band-pass filters - sensing some stimuli while ignoring others - and cellular responses to stimuli can differ based on cell type and differentiation state. Therefore, to fully understand the IOP signals that are relevant to glaucoma, it is necessary to understand the ultimate cellular targets of IOP-induced mechanical stimuli and their ability to sense, ignore, and translate these signals into cellular actions.

Comparison of the Biomechanics of the Mouse Astrocytic Lamina Cribrosa Between Glaucoma and Optic Nerve Crush Models.

Purpose: The strain response of the mouse astrocytic lamina (AL) to an ex vivo mechanical test was compared between two protocols: eyes that underwent sustained intraocular pressure (IOP) increase and eyes after optic nerve crush. Methods: Chronic IOP elevation was induced by microbead injection or the optic nerve was crushed in mice with widespread green fluorescence. After 3 days or 6 weeks, eyes were inflation tested by a published method of two-photon fluorescence to image the AL. Digital volume correlation was used to calculate strains. Optic nerve axon damage was also evaluated. Results: In the central AL but not the peripheral AL, four strains were greater in eyes at the 3-day glaucoma time point than control (P from 0.029 to 0.049, n = 8 eyes per group). Also, at this time point, five strains were greater in the central AL compared to the peripheral AL (P from 0.041 to 0.00003). At the 6-week glaucoma time point, the strains averaged across the specimen, in the central AL, and the peripheral AL were indistinguishable from the respective controls. Strains were not significantly different between controls and eyes 3 days or 6 weeks after crush (n = 8 and 16). Conclusions: We found alterations in the ex vivo mechanical behavior in eyes from mice with experimental glaucoma but not in those with crushed optic nerves. The results of this study demonstrate that significant axon injury does not directly affect mechanical behavior of the astrocytic lamina.

Computational study of the mechanical behavior of the astrocyte network and axonal compartments in the mouse optic nerve head.

Glaucoma is a blinding disease characterized by the degeneration of the retinal ganglion cell (RGC) axons at the optic nerve head (ONH). A major risk factor for glaucoma is the intraocular pressure (IOP). However, it is currently impossible to measure the IOP-induced mechanical response of the axons of the ONH. The objective of this study was to develop a computational modeling method to estimate the IOP-induced strains and stresses in the axonal compartments in the mouse astrocytic lamina (AL) of the ONH, and to investigate the effect of the structural features on the mechanical behavior. We developed experimentally informed finite element (FE) models of six mouse ALs to investigate the effect of structure on the strain responses of the astrocyte network and axonal compartments to pressure elevation. The specimen-specific geometries of the FE models were reconstructed from confocal fluorescent images of cryosections of the mouse AL acquired in a previous study that measured the structural features of the astrocytic processes and axonal compartments. The displacement fields obtained from digital volume correlation in prior inflation tests of the mouse AL were used to determine the displacement boundary conditions of the FE models. We then applied Gaussian process regression to analyze the effects of the structural features on the strain outcomes simulated for the axonal compartments. The axonal compartments experienced, on average, 6 times higher maximum principal strain but 1800 times lower maximum principal stress compared to those experienced by the astrocyte processes. The strains experienced by the axonal compartments were most sensitive to variations in the area of the axonal compartments. Larger axonal compartments that were more vertically aligned, closer to the AL center, and with lower local actin area fraction had higher strains. Understanding the factors affecting the deformation in the axonal compartments will provide insights into mechanisms of glaucomatous axonal damage.

A noninvasive clinical method to measure in vivo mechanical strains of the lamina cribrosa by optical coherence tomography.

Objective: To measure mechanical strain of the lamina cribrosa (LC) after intraocular pressure (IOP) change produced one week after a change in glaucoma medication. Design: Cohort study. Participants: Adult glaucoma patients (23 eyes, 15 patients) prescribed a change in IOP-lowering medication. Intervention: Non-invasive optical coherence tomography (OCT) imaging of the eye. Main Outcomes: Deformation calculated by digital volume correlation of OCT scans of the LC before and after IOP lowering by medication. Results: Among 23 eyes, 17 eyes of 12 persons had IOP lowering ≥ 3 mmHg (reduced IOP group) with tensile anterior-posterior E zz strain = 1.0% ± 1.1% (p = 0.003) and compressive radial strain ( E rr ) = -0.3% ± 0.5% (p=0.012; random effects models accounting inclusion of both eyes in some persons). Maximum in-plane principal (tensile) strain and maximum shear strain in the reduced IOP group were: E max = 1.7% ± 1.0% and Γ max = 1.4% ± 0.7%, respectively (both p<0.0001 versus zero). Reduced IOP group strains E max and Γ max were significantly larger with greater %IOP decrease (<0.0001, <0.0001). The compliance of the E zz , E max , and Γ max strain response, defined as strain normalized by the IOP decrease, were larger with more abnormal perimetric mean deviation or visual field index values (all p≥0.02). Strains were unrelated to age (all p≥0.088). In reduced IOP eyes, mean LC anterior border posterior movement was only 2.05 µm posteriorly (p=0.052) and not related to % IOP change (p=0.94, random effects models). Only E rr was significantly related to ALD change, becoming more negative with greater posterior LC border change (p=0.015). Conclusion: LC mechanical strains can be effectively measured by changes in eye drop medication using OCT and are related to degree of visual function loss in glaucoma. Trial Registration: ClinicalTrials.gov Identifier: NCT03267849.

The Curvature, Collagen Network Structure, and Their Relationship to the Pressure-Induced Strain Response of the Human Lamina Cribrosa in Normal and Glaucoma Eyes.

The lamina cribrosa (LC) is a connective tissue in the optic nerve head (ONH). The objective of this study was to measure the curvature and collagen microstructure of the human LC, compare the effects of glaucoma and glaucoma optic nerve damage, and investigate the relationship between the structure and pressure-induced strain response of the LC in glaucoma eyes. Previously, the posterior scleral cups of 10 normal eyes and 16 diagnosed glaucoma eyes were subjected to inflation testing with second harmonic generation (SHG) imaging of the LC and digital volume correlation (DVC) to calculate the strain field. In this study, we applied a custom microstructural analysis algorithm to the maximum intensity projection of SHG images to measure features of the LC beam and pore network. We also estimated the LC curvatures from the anterior surface of the DVC-correlated LC volume. Results showed that the LC in glaucoma eyes had larger curvatures p≤0.03), a smaller average pore area (p = 0.001), greater beam tortuosity (p < 0.0001), and more isotropic beam structure (p = 0.01) than in normal eyes. The difference measured between glaucoma and normal eyes may indicate remodeling of the LC with glaucoma or baseline differences that contribute to the development of glaucomatous axonal damage.

Acute downregulation of emerin alters actomyosin cytoskeleton connectivity and function.

Fetal lung fibroblasts contribute dynamic infrastructure for the developing lung. These cells undergo dynamic mechanical transitions, including cyclic stretch and spreading, which are integral to lung growth in utero. We investigated the role of the nuclear envelope protein emerin in cellular responses to these dynamic mechanical transitions. In contrast to control cells, which briskly realigned their nuclei, actin cytoskeleton, and extracellular matrices in response to cyclic stretch, fibroblasts that were acutely downregulated for emerin showed incomplete reorientation of both nuclei and actin cytoskeleton. Emerin-downregulated fibroblasts were also aberrantly circular in contrast to the spindle-shaped controls and exhibited an altered pattern of filamentous actin organization that was disconnected from the nucleus. Emerin knockdown was also associated with reduced myosin light chain phosphorylation during cell spreading. Interestingly, emerin-downregulated fibroblasts also demonstrated reduced fibronectin fibrillogenesis and production. These findings indicate that nuclear-cytoskeletal coupling serves a role in the dynamic regulation of cytoskeletal structure and function and may also impact the transmission of traction force to the extracellular matrix microenvironment.

The Strain Response to Intraocular Pressure Decrease in the Lamina Cribrosa of Patients with Glaucoma.

OBJECTIVE: To measure biomechanical strains in the lamina cribrosa (LC) of living human eyes with intraocular pressure (IOP) lowering. DESIGN: Cohort study. PARTICIPANTS: Patients with glaucoma underwent imaging before and after laser suturelysis after trabeculectomy surgery (29 image pairs; 26 persons). INTERVENTION: Noninvasive imaging of the eye. MAIN OUTCOME MEASURES: Strains in optic nerve head tissue and changes in depths of the anterior border of the LC. RESULTS: Intraocular pressure decreases caused the LC to expand in thickness in the anterior-posterior strain (Ezz = 0.94 ± 1.2%; P = 0.00020) and contract in radius in the radial strain (Err = - 0.19 ± 0.33%; P = 0.0043). The mean LC depth did not significantly change with IOP lowering (1.33 ± 6.26 µm; P = 0.26). A larger IOP decrease produced a larger, more tensile Ezz (P < 0.0001), greater maximum principal strain (Emax; P < 0.0001), and greater maximum shear strain (Γmax; P < 0.0001). The average LC depth change was associated with the Γmax and radial-circumferential shear strain (Erθ; P < 0.02) but was not significantly related to tensile or compressive strains. An analysis by clock hour showed that in temporal clock hours 3 to 6, a more anterior LC movement was associated with a more positive Emax, and in clock hours 3, 5, and 6, it was associated with a more positive Γmax. At 10 o'clock, a more posterior LC movement was related to a more positive Emax (P < 0.004). Greater compliance (strain/ΔIOP) of Emax (P = 0.044), Γmax (P = 0.052), and Erθ (P = 0.018) was associated with a thinner retinal nerve fiber layer. Greater compliance of Emax (P = 0.041), Γmax (P = 0.021), Erθ (P = 0.024), and in-plane shear strain (Erz; P = 0.0069) was associated with more negative mean deviations. Greater compliance of Γmax (P = 0.055), Erθ (P = 0.040), and Erz (P = 0.015) was associated with lower visual field indices. CONCLUSIONS: With IOP lowering, the LC moves either into or out of the eye but, on average, expands in thickness and contracts in radius. Shear strains are nearly as substantial as in-plane strains. Biomechanical strains are more compliant in eyes with greater glaucoma damage. This work was registered at ClinicalTrials.gov as NCT03267849.

Mechanical strain in the mouse astrocytic lamina increases after exposure to recombinant trypsin.

The responses of astrocytes in the optic nerve head (ONH) to mechanical and biochemical stimuli are important to understanding the degeneration of retinal ganglion cell axons in glaucoma. The ONH in glaucoma is vulnerable to stress produced by the intraocular pressure (IOP). Notably, after three days of elevated IOP in a mouse model, the junctions between the astrocytic processes and the peripapillary sclera were altered and the structural compliance of the ONH increased. In order to simulate this aspect of glaucomatous remodeling, explanted mouse eyes were treated with TrypLE, a recombinant trypsin enzyme. Treatment with TrypLE caused the periphery of the astrocytic lamina to contract radially by 0.044 ± 0.038. Transmission electron microscopy showed that TrypLE caused a separation of the end-feet of the astrocyte processes from the basement membrane at the junction with the sclera. Inflation testing after treatment with TrypLE caused an increased strain response in the astrocytic lamina compared to the strain response before treatment. The greatest increase was in the radial Green-Lagrange strain, Err = 0.028 ± 0.009, which increased by 340%. The alterations in the microstructure and in the strain response of the astrocytic lamina reported in mouse experimental glaucoma were partially reproduced by experimental treatment of mouse eyes with TrypLE. The results herein suggest that separation of junctions between the astrocyte processes and the sclera may be instrumental in increasing the structural compliance of the ONH after a period of elevated IOP. STATEMENT OF SIGNIFICANCE: Astrocytes of the optic nerve of the eye spread out from edge to edge across the optic nerve in a region referred to as the astrocytic lamina. In an experimental model of glaucoma caused by elevated eye-pressure, there is disruption of the connections between astrocytes and the edge of the astrocytic lamina. We caused a similar event in the lamina by incubating explanted mouse eyes with an enzyme. Disruption of the astrocyte connections to the edge of their tissue caused the tissue to stretch more when we increased the eye-pressure, compared to the control tissue. This work is the first on the tissue of the optic nerve to demonstrate the importance of cell connections in preventing the over-stretching of the astrocytic lamina.

Untethered unidirectionally crawling gels driven by asymmetry in contact forces.

Reversible thermoresponsive hydrogels, which swell and shrink (deswell) in the temperature range of 30° to 60°C, provide an attractive material class for operating untethered soft robots in human physiological and ambient conditions. Crawling has been demonstrated previously with thermoresponsive hydrogels but required patterned or constrained gels or substrates to break symmetry for unidirectional motion. Here, we demonstrate a locomotion mechanism for unidirectionally crawling gels driven by spontaneous asymmetries in contact forces during swelling and deswelling of segmented active thermoresponsive poly(N-isopropylacrylamide) (pNIPAM) and passive polyacrylamide (pAAM) bilayers with suspended linkers. Actuation studies demonstrate the consistent unidirectional movement of these gel crawlers across multiple thermal cycles on flat, unpatterned substrates. We explain the mechanism using finite element simulations and by varying experimental parameters such as the linker stiffness and the number of bilayer segments. We elucidate design criteria and validate experiments using image analysis and finite element models. We anticipate that this mechanism could potentially be applied to other shape-changing locomotors.

Quantitative Microstructural Analysis of Cellular and Tissue Remodeling in Human Glaucoma Optic Nerve Head.

Purpose: To measure quantitatively changes in lamina cribrosa (LC) cell and connective tissue structure in human glaucoma eyes. Methods: We studied 27 glaucoma and 19 age-matched non-glaucoma postmortem eyes. In 25 eyes, LC cross-sections were examined by confocal and multiphoton microscopy to quantify structures identified by anti-glial fibrillary acidic protein (GFAP), phalloidin-labeled F-actin, nuclear 4',6-diamidino-2-phenylindole (DAPI), and by second harmonic generation imaging of LC beams. Additional light and transmission electron microscopy were performed in 21 eyes to confirm features of LC remodeling, including immunolabeling by anti-SOX9 and anti-collagen IV. All glaucoma eyes had detailed clinical histories of open-angle glaucoma status, and degree of axon loss was quantified in retrolaminar optic nerve cross-sections. Results: Within LC pores, the proportionate area of both GFAP and F-actin processes was significantly lower in glaucoma eyes than in controls (P = 0.01). Nuclei were rounder (lower median aspect ratio) in glaucoma specimens (P = 0.02). In models assessing degree of glaucoma damage, F-actin process width was significantly wider in glaucoma eyes with more damage (P = 0.024), average LC beam width decreased with worse glaucoma damage (P = 0.042), and nuclear count per square millimeter rose with worse damage (P = 0.019). The greater cell count in LC pores represented 92.3% astrocytes by SOX9 labeling. The results are consistent with replacement of axons in LC pores by basement membrane labeled by anti-collagen IV and in-migrating astrocytes. Conclusions: Alteration in LC structure in glaucoma involves migration of astrocytes into axonal bundles, change in astrocyte orientation and processes, production of basement membrane material, and thinning of connective tissue beams.

Synergistic Energy Absorption Mechanisms of Architected Liquid Crystal Elastomers.

A unique rate-dependent energy absorption behavior of liquid crystal elastomer (LCE)-based architected materials is reported. The architected materials consist of repeating unit cells of bistable tilted LCE beams sandwiched between stiff supports. The viscoelastic behavior of the LCE causes the energy absorption to increase with strain rate according to a power-law relationship, which can be modulated by changing the degree of mesogen alignment and the loading direction relative to the director. For a strain rate of 600 s-1 , the unit cell exhibits up to a 5 MJ m-3 energy absorption density, which is two orders of magnitude higher than the same structure fabricated from poly(dimethylsiloxane) elastomer and is comparable to the dissipation from irreversible plastic deformation exhibited by denser metals. For a multilayered structure of unit cells, nonuniform buckling of the different layers produces additional viscoelastic dissipation. This synergistic interaction between viscoelastic dissipation and snap-through buckling causes the energy absorption density to increase with the number of layers. The sequence of cell collapse can be controlled by grading the beam thickness to further promote viscous dissipation and enhance the energy absorption density. It is envisioned that the study can contribute to the development of lightweight extreme energy-absorbing metamaterials.

Regional Differences and Physiologic Behaviors in Peripapillary Scleral Fibroblasts.

Purpose: The purpose of this study was to describe the cellular architecture of normal human peripapillary sclera (PPS) and evaluate surface topography's role in fibroblast behavior. Methods: PPS cryosections from nonglaucomatous eyes were labelled for nuclei, fibrillar actin (FA), and alpha smooth muscle actin (αSMA) and imaged. Collagen fibrils were imaged using second harmonic generation. Nuclear density and aspect ratio of the internal PPS (iPPS), outer PPS (oPPS), and peripheral sclera were determined. FA and αSMA fibril alignment with collagen extracellular matrix (ECM) was determined. PPS fibroblasts were cultured on smooth or patterned membranes under mechanical strain and in the presence of TGFß1 and 2. Results: The iPPS (7.1 ± 2.0 × 10-4, P < 0.0001) and oPPS (5.3 ± 1.4 × 10-4, P = 0.0013) had greater nuclei density (nuclei/µm2) than peripheral sclera (2.5 ± 0.8 × 10-4). The iPPS (2.0 ± 0.3, P = 0.002) but not oPPS (2.4 ± 0.4, P = 0.45) nuclei had smaller aspect ratios than peripheral (2.7 ± 0.5) nuclei. FA was present throughout the scleral stroma and was more aligned with oPPS collagen (9.6 ± 1.9 degrees) than in the peripheral sclera (15.9 ± 3.9 degrees, P =0.002). The αSMA fibers in the peripheral sclera were less aligned with collagen fibrils (26.4 ± 4.8 degrees) than were FA (15.9 ± 3.9 degrees, P = 0.0002). PPS fibroblasts cultured on smooth membranes shifted to an orientation perpendicular to the direction of cyclic uniaxial strain (1 Hz, 5% strain, 42.2 ± 7.1 degrees versus 62.0 ± 8.5 degrees, P < 0.0001), whereas aligned fibroblasts on patterned membranes were resistant to strain-induced reorientation (5.9 ± 1.4 degrees versus 10 ± 3.3 degrees, P = 0.21). Resistance to re-orientation was reduced by TGFß treatment (10 ± 3.3 degrees without TGFß1 compared to 23.1 ± 4.5 degrees with TGFß1, P < 0.0001). Conclusions: Regions of the posterior sclera differ in cellular density and nuclear morphology. Topography alters the cellular response to mechanical strain.

Estimations of the effective Young's modulus of specimens prepared by fused filament fabrication.

The elastic response of homogeneous isotropic materials is most commonly represented by their Young's modulus (E), but geometric variability associated with additive manufacturing results in materials that are neither homogeneous nor isotropic. Here we investigated methods to estimate the effective elastic modulus (Eeff) of samples fabricated by fused filament fabrication. We conducted finite element analysis (FEA) on printed samples based on material properties and CT-scanned geometries. The analysis revealed how the layer structure of a specimen altered the internal stress distribution and the resulting Eeff. We also investigated different empirical methods to estimate Eeff as guides. We envision the findings from our study can provide guidelines for modulus estimation of as-printed specimens, with the potential of applying to other extrusion-based additive manufacturing technologies.

Biomechanics of the optic nerve head and peripapillary sclera in a mouse model of glaucoma.

The deformation of the mouse astrocytic lamina (AL) and adjacent peripapillary sclera (PPS) was measured in response to elevated intraocular pressure. We subjected explanted mouse eyes to inflation testing, comparing control eyes to those 3 days and 6 weeks after induction of ocular hypertension (OHT) via ocular microbead injection. Laser scanning microscopy was used with second harmonic generation to image the collagenous PPS and two-photon fluorescence to image transgenic fluorescent astrocytes in the AL. Digital volume correlation was applied to calculate strains in the PPS and AL. The specimen-averaged strains were biaxial in the AL and PPS, with greater strain overall in the x- than y-direction in the AL and greater strain in the θ- than the r-direction in the PPS. Strains increased after 3-day OHT, with greater strain overall in the 3-day AL than control AL, and greater circumferential strain in the 3-day PPS than control PPS. In the 6-week OHT eyes, AL and PPS strains were similar overall to controls. This experimental glaucoma model demonstrated a dynamic change in the mechanical behaviour of the AL and PPS over time at the site of neuronal injury and remodelling in glaucoma.

Pressure-Induced Changes in Astrocyte GFAP, Actin, and Nuclear Morphology in Mouse Optic Nerve.

Purpose: To conduct quantitative analysis of astrocytic glial fibrillary acidic protein (GFAP), actin and nuclei distribution in mouse optic nerve (ON) and investigate changes in the measured features after 3 days of ocular hypertension (OHT). Method: Serial cross-sections of 3-day microbead-induced OHT and control ONs were fluorescently labelled and imaged using confocal microscope. Eighteen structural features were measured from the acquired images, including GFAP coverage, actin area fraction, process thickness, and aspect ratio of cell nucleus. The measured features were analyzed for variations with axial locations along ON and radial zones transverse to ON, as well as for the correlations with degree of intraocular pressure (IOP) change. Results: The most significant changes in structural features after 3-day OHT occurred in the unmyelinated ON region (R1), and the changes were greater with greater IOP elevation. Although the GFAP, actin, axonal, and ON areas all increased in 3-day OHT ONs in R1 (P ≤ 0.004 for all), the area fraction of GFAP actually decreased (P = 0.02), the actin area fraction was stable and individual axon compartments were unchanged in size. Within R1, the number of nuclear clusters increased (P < 0.001), but the mean size of nuclear clusters was smaller (P = 0.02) and the clusters became rounder (P < 0.001). In all cross-sections of control ONs, astrocytic processes were thickest in the rim zone compared with the central and peripheral zones (P ≤ 0.002 for both), whereas the overall process width in R1 decreased after 3 days of OHT (P < 0.001). Conclusions: The changes in structure elucidated IOP-generated alterations that underlie astrocyte mechanotranslational responses relevant to glaucoma.

The effect of alignment on the rate-dependent behavior of a main-chain liquid crystal elastomer.

This study investigated the effect of alignment on the rate-dependent behavior of a main-chain liquid crystal elastomer (LCE). Polydomain nematic LCE networks were synthesized from a thiol-acrylate Michael addition reaction in the isotropic state. The polydomain networks were stretched to different strain levels to induce alignment then crosslinked in a second stage photopolymerization process. The LCE networks were subjected to dynamic mechanical tests to measure the temperature-dependent storage modulus and uniaxial tension load-unload tests to measure the rate-dependence of the Young's modulus, mechanical dissipation, and characteristics of the soft stress response. Three-dimensional (3D) digital image correlation (DIC) was used to characterize the effect of domain/mesogen relaxation on the strain fields. All LCE networks exhibited a highly rate-dependent stress response with significant inelastic strains after unloading. The Young's modulus of the loading curve and hysteresis of the load-unload curves showed a power-law dependence on the strain rate. The Young's modulus increased with alignment and larger anisotropy and a smaller power-law exponent was measured for the Young's modulus and hysteresis for the highly aligned monodomains. The polydomain and pre-stretched networks loaded perpendicular to the alignment direction exhibited a soft stress response that featured a rate-dependent peak stress, strain-softening, and strain-stiffening. The 3D-DIC strain fields for the polydomain network and programmed networks stretched in the perpendicular direction were highly heterogeneous, showing regions of alternating higher and lower strains. The strain variations increased initially with loading, peaked during the strain softening part of the stress response, then decreased during the strain stiffening part of the stress response. Greater variability was measured for lower strain rates. These observations suggest that local domain/mesogen relaxation led to the development of the heterogeneous strain patterns and strain softening in stress response. These findings improved understanding of the kinetics of mesogen relaxation and its contributions to the rate-dependent stress response and mechanical dissipation.

The Effects of Glaucoma on the Pressure-Induced Strain Response of the Human Lamina Cribrosa.

Purpose: To measure the ex vivo pressure-induced strain response of the human optic nerve head and analyze for variations with glaucoma diagnosis and optic nerve axon damage. Methods: The posterior sclera of 16 eyes from 8 diagnosed glaucoma donors and 10 eyes from 6 donors with no history of glaucoma were inflation tested between 5 and 45 mm Hg. The optic nerve from each donor was examined for degree of axon loss. The posterior volume of the lamina cribrosa (LC) was imaged with second harmonic generation and analyzed using volume correlation to calculate LC strains between 5 and 10 and 5 and 45 mm Hg. Results: Eye length and LC area were larger in eyes diagnosed with glaucoma (P ≤ 0.03). Nasal-temporal EXX and circumferential Eθθ strains were lower in the LC of diagnosed glaucoma eyes at 10 mm Hg (P ≤ 0.05) and 45 mm Hg (P ≤ 0.07). EXX was smaller in the LC of glaucoma eyes with <25% axon loss compared with undamaged normal eyes (P = 0.01, 45 mm Hg). In general, the strains were larger in the peripheral than central LC. The ratio of the maximum principal strain Emax in the peripheral to central LC was larger in glaucoma eyes with >25% axon loss than in glaucoma eyes with milder damage (P = 0.004, 10 mm Hg). Conclusions: The stiffness of the LC pressure-strain response was greater in diagnosed glaucoma eyes and varied with glaucomatous axon damage. Lower LC strains in glaucoma eyes with milder damage may represent baseline biomechanical behavior that contributes to axon loss, whereas greater LC strain and altered radial LC strain variation in glaucoma eyes with more severe damage may be caused by glaucoma-related remodeling.