Ehsan Vaghefi

Visualizing ocular lens fluid dynamics using MRI: manipulation of steady state water content and water fluxes

Studies using various MRI techniques have shown that a water-protein concentration gradient exists in the ocular lens. Because this concentration is higher in the core relative to the lens periphery, a gradient in refractive index is established in the lens. To investigate how the water-protein concentration profile is maintained, bovine lenses were incubated in different solutions, and changes in water-protein concentration ratio monitored using proton density weighted (PD-weighted) imaging in the absence and presence of heavy water (D(2)O). Lenses incubated in artificial aqueous humor (AAH) maintained the steady state water-protein concentration gradient, but incubating lenses in high extracellular potassium (KCl-AAH) or low temperature (Low T-AAH) caused a collapse of the gradient due to a rise in water content in the core of the lens. To visualize water fluxes, lenses were incubated in D(2)O, which acts as a contrast agent. Incubation in KCl-AAH and low T-AAH dramatically slowed the movement of D(2)O into the core but did not affect the movement of D(2)O into the outer cortex. D(2)O seemed to preferentially enter the lens cortex at the anterior and posterior poles before moving circumferentially toward the equatorial regions. This directionality of D(2)O influx into the lens cortex was abolished by incubating lenses in high KCl-AAH or low T-AAH, and resulted in homogenous influx of D(2)O into the outer cortex. Taken together, our results show that the water-protein concentration ratio is actively maintained in the core of the lens and that water fluxes preferentially enter the lens at the poles.

Visualization of transverse diffusion paths across fiber cells of the ocular lens by small animal MRI

The sense of vision requires that light penetrate through the ocular lens. Experiments, performed and published by many research groups, have suggested that the lens, which has no blood vessels, relies on internally directed ion and water fluxes for its circulation, survival and transparency. We investigated the internal diffusive pathways of the lens in order to better understand the constraints that may be operating on directional lens fluxes. Small animal magnetic resonance imaging, including T2-weighted and diffusion tensor imaging, was used to measure tissue properties and diffusivity throughout cultured bovine lenses. A range of concentric regions of signal intensity was distinguished inside the lens, by both T2-weighted signal and mean diffusivity. Diffusivity mapping of the lens revealed novel anisotropic polar and equatorial zones of pronounced diffusivity directed transverse to the fiber cells. In contrast, an inner zone including the lens nucleus showed isotropic and weak diffusivity. Our results lend support to models of internally directed lens micro-circulation, by placing non-structural diffusive constraints on global patterns of fluid circulation.

Magnetic resonance and confocal imaging of solute penetration into the lens reveals a zone of restricted extracellular space diffusion

It has been proposed that in the absence of blood supply, the ocular lens operates an internal microcirculation system that delivers nutrients to internalized fiber cells faster and more efficiently than would occur by passive diffusion alone. To visualize the extracellular space solute fluxes potentially generated by this system, bovine lenses were organ cultured in artificial aqueous humor (AAH) for 4 h in the presence or absence of two gadolinium-based contrast agents, ionic Gd(3+), or a chelated form of Gd(3+), Gd-diethylenetriamine penta-acetic acid (Gd-DTPA; mol mass = 590 Da). Contrast reagent penetration into the lens core was monitored in real time using inversion recovery-spin echo (IR-SE) magnetic resonance imaging (MRI), while steady-state accumulation of [Gd-DTPA](-2) was also determined by calculating T1 values. After incubation, lenses were fixed and cryosectioned, and sections were labeled with the membrane marker wheat germ agglutinin (WGA). Sections were imaged by confocal microscopy using standard and reflectance imaging modalities to visualize the fluorescent WGA label and gadolinium reagents, respectively. Real-time IR-SE MRI showed rapid penetration of Gd(3+) into the outer cortex of the lens and a subsequent bloom of signal in the core. These two areas of signal were separated by an area in the inner cortex that limited entry of Gd(3+). Similar results were obtained for Gd-DTPA, but the penetration of the larger negatively charged molecule into the core could only be detected by calculating T1 values. The presence of Gd-DTPA in the extracellular space of the outer cortex and core, but its apparent absence from the inner cortex was confirmed using reflectance imaging of equatorial sections. In axial sections, Gd-DTPA was associated with the sutures, suggesting these structures provide a pathway from the surface, across the inner cortex barrier to the lens core. Our studies have revealed inner and outer boundaries of a zone within which a narrowing of the extracellular space restricts solute diffusion and acts to direct fluxes into the lens core via the sutures.

The effect of flavonoids on visual function in patients with glaucoma or ocular hypertension: a systematic review and meta-analysis

BackgroundGlaucoma is a leading cause of irreversible blindness worldwide. A major symptom of this pathology is the loss to the visual field in a peripheral to central pattern. Flavonoids are polyphenol compounds sourced from plants, commonly found in green tea, red wine and cocoa, and they have neuroprotective and antioxidant characteristics proposed to be advantageous within the context of glaucoma. Currently, the literature presents conflicting evidence regarding the effect of flavonoids on patients with glaucoma and ocular hypertension; hence a systematic review and meta-analysis was conducted.MethodDatabases included in our literature search were EMBASE (1980-present), MEDLINE Ovid, Alternative and Complementary Medicine Database (AMED), Cumulative Index to Nursing and Allied Health Literature (CINAHL) and Cochrane Central Register of Controlled Trials (CENTRAL). Meta-analysis was performed using RevMan 5 (Review Manager) 5 software, version 5.3 (The Nordic Cochrane Centre, The Cochrane Collaboration, Copenhagen). The primary outcomes were visual field mean deviation (MD) and intraocular pressure (IOP). Secondary outcomes were ocular blood flow and blood pressure (BP).ConclusionMeta-analyses showed that flavonoids have a promising role in improving visual function in patients with glaucoma and ocular hypertension (OHT), and appear to play a part in both improving and slowing the progression of visual field loss.

The physiological optics of the lens.

ABSTRACT The optical properties of the ocular lens are important to overall vision quality. As a transparent biological tissue, the lens contributes to the overall and dynamic focussing power of the eye, and corrects for optical errors introduced by the cornea. The optical properties of the lens change throughout life. Alterations to the refractive properties and transparency of the lens result in presbyopia and cataract, respectively. However, it is not well understood how changes to lens cellular structure and function initiate these changes in refraction and transparency. Here, we attempt to bridge this knowledge gap by reviewing how the optical properties of the lens are first established, and then maintained at the cellular level throughout the lifetime of an individual. Central to this understanding is the fact that the lens has a microcirculation system that generates a flux of ions and water that circulates through the lens. By supporting ionic and metabolic homeostasis in the lens, the system actively maintains lens transparency, and by regulating the steady state water content of the lens, controls the two key parameters, lens geometry and the gradient of refractive index, which determine the refractive properties of the lens. Thus, water transport is emerging as the critical parameter that links the transparency and refractive properties of the lens at the cellular level, and highlights the need to study how age‐related changes in water transport result in presbyopia and cataract, the leading causes of refractive error and blindness in the world today. HIGHLIGHTSThe lens operates a microcirculation system that maintains lens homeostasis.Spatial differences in cellular physiology generates the lens microcirculation.Lens physiology is linked to the active maintenance of its optical properties.Failure of the microcirculation can explain the onset of presbyopia and cataract.

Characterization of the Effects of Hyperbaric Oxygen on the Biochemical and Optical Properties of the Bovine Lens

PURPOSE To assess the morphologic, biochemical, and optical properties of bovine lenses treated with hyperbaric oxygen. METHODS Lenses were exposed to hyperbaric nitrogen (HBN) or hyperbaric oxygen (HBO) for 5 or 15 hours, lens transparency was assessed using bright field microscopy and lens morphology was visualized using confocal microscopy. Lenses were dissected into the outer cortex, inner cortex, and core, and glutathione (GSH) and malondialdehyde (MDA) measured. Gel electrophoresis and Western blotting were used to detect high molecular weight aggregates (HMW) and glutathione mixed protein disulfides (PSSG). T2-weighted MRI was used to measure lens geometry and map the water/protein ratio to allow gradient refractive index (GRIN) profiles to be calculated. Optical modeling software calculated the change in lens optical power, and an anatomically correct model of the light pathway of the bovine eye was used to determine the effects of HBN and HBO on focal length and overall image quality. RESULTS Lenses were transparent and lens morphology similar between HBN- and HBO-treated lenses. At 5- and 15-hour HBO exposure, GSH and GSSG were depleted and MDA increased in the core. Glutathione mixed protein disulfides were detected in the outer and inner cortex only with no appearance of HMW. Optical changes were detectable only with 15-hour HBO treatment with a decrease in the refractive index of the core, slightly reduced lens thickness, and an increase in optimal focal length, consistent with a hyperopic shift. CONCLUSIONS This system may serve as a model to study changes that occur with advanced aging rather than nuclear cataract formation per se.

Active Maintenance of the Gradient of Refractive Index Is Required to Sustain the Optical Properties of the Lens.

PURPOSE To determine whether the cellular physiology of the lens actively maintains the optical properties of the lens and whether inhibition of lens transport affects overall visual quality. METHODS One lens from a pair of bovine lenses was cultured in artificial aqueous humor (AAH), while the other was cultured in either AAH-High-K+ or AAH + 0.1 mM ouabain for 4 hours. Lens pairs or whole enucleated eyes were then imaged in 4.7 Tesla (T) high-field small animal magnet. Lens surface curvatures, T1 measurements of water content, and T2 measurements of water/protein ratios were extracted from cultured lenses, while the geometrical parameters that define the optical pathway were obtained from whole eyes. Gradients of refractive index (GRIN), calculated from T2 measurements, and the extracted geometric parameters were inputted into optical models of the isolated lens and the whole bovine eye. RESULTS Inhibiting circulating fluxes by inhibiting the Na/K-ATPase with ouabain or depolarization of the lens potential by High K+ caused changes to lens water content, the water/protein ratio (GRIN) and surface geometry that manifested as an increase in optical power and a decrease in negative spherical aberration in cultured lenses. Changes to optical properties of the lens resulted in a myopic shift that impaired vision quality in the optical model of the bovine eye. CONCLUSIONS The cellular physiology of the lens actively maintains its optical properties and inhibiting the Na/K/ATPase induces a myopic shift in vision similar to that observed clinically in patients who go on to develop cataract.

A computer model of lens structure and function predicts experimental changes to steady state properties and circulating currents

BackgroundIn a previous study (Vaghefi et al. 2012) we described a 3D computer model that used finite element modeling to capture the structure and function of the ocular lens. This model accurately predicted the steady state properties of the lens including the circulating ionic and fluid fluxes that are believed to underpin the lens internal microcirculation system. In the absence of a blood supply, this system brings nutrients to the core of the lens and removes waste products faster than would be achieved by passive diffusion alone. Here we test the predictive properties of our model by investigating whether it can accurately mimic the experimentally measured changes to lens steady-state properties induced by either depolarising the lens potential or reducing Na+ pump rate.MethodsTo mimic experimental manipulations reported in the literature, the boundary conditions of the model were progressively altered and the model resolved for each new set of conditions. Depolarisation of lens potential was implemented by increasing the extracellular [K+], while inhibition of the Na+ pump was stimulated by utilising the inherent temperature sensitivity of the pump and changing the temperature at which the model was solved.ResultsOur model correctly predicted that increasing extracellular [K+] depolarizes the lens potential, reducing and then reversing the magnitude of net current densities around the lens. While lowering the temperature reduced Na+ pump activity and caused a reduction in circulating current, it had a minimal effect on the lens potential, a result consistent with published experimental data.ConclusionWe have shown that our model is capable of accurately simulating the effects of two known experimental manipulations on lens steady-state properties. Our results suggest that the model will be a valuable predictive tool to support ongoing studies of lens structure and function.

Development of a 3D finite element model of lens microcirculation

BackgroundIt has been proposed that in the absence of a blood supply, the ocular lens operates an internal microcirculation system. This system delivers nutrients, removes waste products and maintains ionic homeostasis in the lens. The microcirculation is generated by spatial differences in membrane transport properties; and previously has been modelled by an equivalent electrical circuit and solved analytically. While effective, this approach did not fully account for all the anatomical and functional complexities of the lens. To encapsulate these complexities we have created a 3D finite element computer model of the lens.MethodsInitially, we created an anatomically-correct representative mesh of the lens. We then implemented the Stokes and advective Nernst-Plank equations, in order to model the water and ion fluxes respectively. Next we complemented the model with experimentally-measured surface ionic concentrations as boundary conditions and solved it.ResultsOur model calculated the standing ionic concentrations and electrical potential gradients in the lens. Furthermore, it generated vector maps of intra- and extracellular space ion and water fluxes that are proposed to circulate throughout the lens. These fields have only been measured on the surface of the lens and our calculations are the first 3D representation of their direction and magnitude in the lens.ConclusionValues for steady state standing fields for concentration and electrical potential plus ionic and fluid fluxes calculated by our model exhibited broad agreement with observed experimental values. Our model of lens function represents a platform to integrate new experimental data as they emerge and assist us to understand how the integrated structure and function of the lens contributes to the maintenance of its transparency.

3D Finite Element Modeling of Avascular Circulation in the Ocular Lens

The ability to see is dependent on the actions of several structures in and around the eyeball. By looking at an object, light rays are reflected from the object to the cornea. Light rays are refracted and focused by the cornea, lens, and “vitreous”. The lens function is to ensure that the light rays come to a sharp focus point on the retina.