A Histomorphometric and Computational Investigation of the Stabilizing Role of Pectinate Ligaments in the Aqueous Outflow Pathway.

Murine models are commonly used to study glaucoma, the leading cause of irreversible blindness. Glaucoma is associated with elevated intraocular pressure (IOP), which is regulated by the tissues of the aqueous outflow pathway. In particular, pectinate ligaments (PLs) connect the iris and trabecular meshwork (TM) at the anterior chamber angle, with an unknown role in maintenance of the biomechanical stability of the aqueous outflow pathway, thus motivating this study. We conducted histomorphometric analysis and optical coherence tomography-based finite element (FE) modeling on three cohorts of C57BL/6 mice: 'young' (2-6 months), 'middle-aged' (11-16 months), and 'elderly' (25-32 months). We evaluated the age-specific morphology of the outflow pathway tissues. Further, because of the known pressure-dependent Schlemm's canal (SC) narrowing, we assessed the dependence of the SC lumen area to varying IOPs in age-specific FE models over a physiological range of TM/PL stiffness values. We found age-dependent changes in morphology of outflow tissues; notably, the PLs were more developed in older mice compared to younger ones. In addition, FE modeling demonstrated that murine SC patency is highly dependent on the presence of PLs, and that increased IOP caused SC collapse only with sufficiently low TM/PL stiffness values. Moreover, the elderly model showed more susceptibility to SC collapse compared to the younger models. In conclusion, our study elucidated the previously unexplored role of PLs in the aqueous outflow pathway, indicating their function in supporting TM and SC under elevated IOP.

Individual-Specific Modeling of Rat Optic Nerve Head Biomechanics in Glaucoma.

Glaucoma is the second leading cause of blindness worldwide and is characterized by the death of retinal ganglion cells (RGCs), the cells that send vision information to the brain. Their axons exit the eye at the optic nerve head (ONH), the main site of damage in glaucoma. The importance of biomechanics in glaucoma is indicated by the fact that elevated intraocular pressure (IOP) is a causative risk factor for the disease. However, exactly how biomechanical insult leads to RGC death is not understood. Although rat models are widely used to study glaucoma, their ONH biomechanics have not been characterized in depth. Therefore, we aimed to do so through finite element (FE) modeling. Utilizing our previously described method, we constructed and analyzed ONH models with individual-specific geometry in which the sclera was modeled as a matrix reinforced with collagen fibers. We developed eight sets of scleral material parameters based on results from our previous inverse FE study and used them to simulate the effects of elevated IOP in eight model variants of each of seven rat ONHs. Within the optic nerve, highest strains were seen inferiorly, a pattern that was consistent across model geometries and model variants. In addition, changing the collagen fiber direction to be circumferential within the peripapillary sclera resulted in more pronounced decreases in strain than changing scleral stiffness. The results from this study can be used to interpret data from rat glaucoma studies to learn more about how biomechanics affects RGC pathogenesis in glaucoma.

A Methodology for Individual-Specific Modeling of Rat Optic Nerve Head Biomechanics in Glaucoma.

Glaucoma is the leading cause of irreversible blindness and involves the death of retinal ganglion cells (RGCs). Although biomechanics likely contributes to axonal injury within the optic nerve head (ONH), leading to RGC death, the pathways by which this occurs are not well understood. While rat models of glaucoma are well-suited for mechanistic studies, the anatomy of the rat ONH is different from the human, and the resulting differences in biomechanics have not been characterized. The aim of this study is to describe a methodology for building individual-specific finite element (FE) models of rat ONHs. This method was used to build three rat ONH FE models and compute the biomechanical environment within these ONHs. Initial results show that rat ONH strains are larger and more asymmetric than those seen in human ONH modeling studies. This method provides a framework for building additional models of normotensive and glaucomatous rat ONHs. Comparing model strain patterns with patterns of cellular response seen in studies using rat glaucoma models will help us to learn more about the link between biomechanics and glaucomatous cell death, which in turn may drive the development of novel therapies for glaucoma.

Biomechanics of the posterior eye: a critical role in health and disease.

The posterior eye is a complex biomechanical structure. Delicate neural and vascular tissues of the retina, choroid, and optic nerve head that are critical for visual function are subjected to mechanical loading from intraocular pressure, intraocular and extraorbital muscles, and external forces on the eye. The surrounding sclera serves to counteract excessive deformation from these forces and thus to create a stable biomechanical environment for the ocular tissues. Additionally, the eye is a dynamic structure with connective tissue remodeling occurring as a result of aging and pathologies such as glaucoma and myopia. The material properties of these tissues and the distribution of stresses and strains in the posterior eye is an area of active research, relying on a combination of computational modeling, imaging, and biomechanical measurement approaches. Investigators are recognizing the increasing importance of the role of the collagen microstructure in these material properties and are undertaking microstructural measurements to drive microstructurally-informed models of ocular biomechanics. Here, we review notable findings and the consensus understanding on the biomechanics and microstructure of the posterior eye. Results from computational and numerical modeling studies and mechanical testing of ocular tissue are discussed. We conclude with some speculation as to future trends in this field.

Biaxial mechanical testing of human sclera.

The biomechanical environment of the optic nerve head (ONH), of interest in glaucoma, is strongly affected by the biomechanical properties of sclera. However, there is a paucity of information about the variation of scleral mechanical properties within eyes and between individuals. We thus used biaxial testing to measure scleral stiffness in human eyes. Ten eyes from 5 human donors (age 55.4+/-3.5 years; mean+/-SD) were obtained within 24h of death. Square scleral samples (6mm on a side) were cut from each ocular quadrant 3-9 mm from the ONH centre and were mechanically tested using a biaxial extensional tissue tester (BioTester 5000, CellScale Biomaterials Testing, Waterloo). Stress-strain data in the latitudinal (toward the poles) and longitudinal (circumferential) directions, here referred to as directions 1 and 2, were fit to the four-parameter Fung constitutive equation W=c(e(Q)-1), where Q=c(1)E(11)(2)+c(2)E(22)(2)+2c(3)E(11)E(22) and W, c's and E(ij) are the strain energy function, material parameters and Green strains, respectively. Fitted material parameters were compared between samples. The parameter c(3) ranged from 10(-7) to 10(-8), but did not contribute significantly to the accuracy of the fitting and was thus fixed at 10(-7). The products cc(1) and cc(2), measures of stiffness in the 1 and 2 directions, were 2.9+/-2.0 and 2.8+/-1.9 MPa, respectively, and were not significantly different (two-sided t-test; p=0.795). The level of anisotropy (ratio of stiffness in orthogonal directions) was 1.065+/-0.33. No statistically significant correlations between sample thickness and stiffness were found (correlation coefficients=-0.026 and -0.058 in directions 1 and 2, respectively). Human sclera showed heterogeneous, near-isotropic, nonlinear mechanical properties over the scale of our samples.

Modeling aqueous humor collection from the human eye.

Glaucoma is a common cause of blindness. Studies of this disease can involve collection of aqueous humor (AH) fluid from eyes of patients undergoing surgery, which involves aspirating a small fluid volume from the anterior region of the eye through a fine-bore needle. Unfortunately, the composition of the AH is spatially non-uniform in the eye, and thus the composition of the aspirated fluid is uncertain. Our goal was to numerically simulate the AH aspiration process to determine where the aspirated fluid was being collected from and thus gain insight into the composition of the collected fluid. A 3D computational model of the anterior region of the human eye was created and the Navier-Stokes equations were numerically solved during the aspiration process for a set of typical (baseline) conditions: 40 microl aspirated volume and needle placement in the central anterior chamber. We also ran variations of this baseline simulation. The main finding was that the aspirated fluid comes from a very localized region around the needle tip, so that for typical conditions, almost no aspirated fluid is withdrawn from the angle region of the anterior chamber. This is important because the AH in this angle region is protein-rich and directly interacts with the tissues that control fluid drainage from the eye. Recommendations for standardizing aspiration conditions are given.

Targeted gene transfer to Schlemm's canal by retroperfusion.

The goal of the present study was to specifically modify protein expression in the resistance-generating region of the conventional outflow pathway, namely the inner wall of Schlemm's canal (SC) and the juxtacanalicular region of the trabecular meshwork, in perfused human anterior segments. Anterior segments from human cadaveric eyes were prepared for organ culture using standard techniques and were perfused at constant flow while recording pressure. After reaching a stable outflow facility within physiological limits, forward perfusion was stopped and a fluid-tight fence encircling the limbus was installed and filled with media containing an adenovirus encoding the lacZ reporter gene (either 2 x 10(6) or 6 x 10(6)PFU/ml). With the limbus submerged, pressure inside the chamber was lowered to -1 mmHg to facilitate reverse perfusion of virus into SC ("retroperfusion"). After 30-60 min at zero pressure (with some mixing), forward perfusion was restarted and continued for 5-7 days, after which anterior segments were fixed and processed for visualization of lacZ activity. Retroperfusion of nine anterior segments with adenovirus encoding a reporter gene did not appreciably alter baseline outflow facility (0.27+/-0.05 versus 0.29+/-0.08 microl/min per mmHg post-retroperfusion). Gross examination of outflow tissues showed focal distribution of lacZ activity around the circumference of SC, presumably near collector channels. In segments that were sequentially tilted during retroperfusion, the distribution of lacZ activity appeared more uniform. Sagittal histological sections showed lacZ activity in all portions of the conventional drainage tract, particularly cells in the resistance-generating region. Taken together, the results demonstrate that candidate protein expression by cells in the resistance-generating region of the conventional drainage pathway can be specifically modified by retroperfusion of adenovirus and examined for effects on outflow facility.

Twenty-fold difference in hemodynamic wall shear stress between murine and human aortas.

Endothelial cells regulate vascular tone and mural remodelling in a shear-dependent manner that is commonly assumed to keep wall shear stress constant across arteries and species. Allometric arguments show that aortic flow velocity is constant across species, a deduction that is consistent with much experimental data, but the same arguments also show that the shear stress experienced by aortic endothelium will depend inversely on body mass to the 3/8th power, and hence will be 20-fold higher in mice than in men. This conclusion is robust and has important implications for the study of shear-dependent vascular biology and pathology.