Influence of molecular shape, conformability, net surface charge, and tissue interaction on transscleral macromolecular diffusion

Abstract

Purpose: To study the influence of molecular shape, conformability, net surface charge and tissue interaction on transscleral diffusion.Methods: Unfixed, porcine sclera was clamped in an Ussing chamber. Fluorophore labelled, neutral, albumin, dextran, or ficoll were placed in one hemi-chamber and the rate of transscleral diffusion was measured over 24 hours using a spectrophotometer. Experiments were repeated using dextrans and ficoll with positive, or negative, net surface charges.Fluorescence recovery after photobleaching (FRAP) was undertaken to compare transscleral diffusion with diffusion through a solution.All molecules were 70 kDa.Results: Using FRAP, mean ± SD diffusion coefficient (D) was highest for albumin, followed by ficoll, then dextran (p = 0.0005). Positive dextrans diffused fastest, followed by negative, then neutral dextrans (p = 0.0005). Neutral ficoll diffused the fastest, followed by positive then negative ficoll (p = 0.0008). For the neutral molecules, transscleral D was highest for albumin, followed by dextran, then ficoll (p < 0.0001). D was highest for negative ficoll, followed by neutral, then positive ficoll (p < 0.0001). By contrast, D was highest for positive dextran, followed by neutral, then negative dextran (p = 0.0021).Conclusions: Diffusion in free solution does not predict transscleral diffusion and the molecular-tissue interaction is important. Molecular size, shape, and charge may all markedly influence transscleral diffusion, as may conformability to a lesser degree, but all need to be considered when selecting or designing drugs for transscleral delivery.Drugs directed against vascular endothelial growth factor (VEGF) have proven efficacy in some of the most common retinal diseases. The ANCHOR and MARINA studies demonstrated efficacy in wet age-related macular degeneration (AMD), 1,2 and studies such as CRUISE, BRAVO 3,4 and several from the DRCnet Group have shown favourable results in retinal vein occlusion and diabetic macular oedema. Whilst the data in these studies have led to improved clinical care, the need for regular intravitreal injections represents a burdensome regimen of treatment. A less invasive mode of delivery would have potential advantages in terms of reduced cost, discomfort, and complication rate.Systemic administration is impeded by the blood-aqueous and blood-retinal-barriers. High drug concentrations may be required to overcome these barriers to diffusion, with the attendant risk of systemic side effects. 5 A topical route would have obvious advantages. Delivery to the retina is however difficult for several reasons, such as lacrimation, low corneal permeability to large molecules, counter directional intraocular convection, and importantly the long diffusional distance. 53 Blood flow in the conjunctiva, episclera and choroid can also reduce drug concentration. 33,50 Notwithstanding these potential difficulties, transscleral delivery has a number of potential advantages. 6,7,8,9 The sclera has a large and accessible surface area, and a high d...

A spectrophotometer-based diffusivity assay reveals that diffusion hindrance of small molecules in extracellular matrix gels used in 3D cultures is dominated by viscous effects

“…Despite all these variability sources, it is remarkable the good agreement between our macroscopic diffusivity data and that obtained with FRAP and other microscopic assays. Accordingly, our work expands a recent study that used a Nanodrop spectrophotometer to assess the apparent diffusivity in scleral tissue [36], and altogether support that spectrophotometer-based assays provide a suitable and costeffective alternative to assess the apparent diffusivity in ECM gels used in 3D cultures with reasonable accuracy.…”

Permeation of Proteins, Oligonucleotide and Dextrans Across Ocular Tissues: Experimental Studies and a Literature Update

“…Recently, Wen et al compared proteins and dextrans and found that dextrans had a higher normalized permeability across the sclera (permeation coefficient normalized by the aqueous diffusion coefficient) compared with proteins of similar MW, probably because of higher structure flexibility, analogously to reports of diffusion of proteins and polysaccharides across other porous membranes. On the contrary, Srikantha et al reported slower diffusion of neutral dextran and neutral Ficoll compared with that of albumin.…”

“…The analysis of the data in Table does not permit to draw conclusions of the effect of the charge on the permeability of macromolecules, also because differences in the charge often follow differences in the shape and conformation. In this regard, Srikantha et al studied macromolecules diffusion across the sclera using neutral, cationic and anionic Ficoll and dextrans with MW of 70 kDa. Their results indicate that a charge have a different impact on the permeability, depending on the structure of the molecules, suggesting that a charge can have an indirect effect because of a change in the molecular conformation.…”

Chitosan-coated PLGA nanoparticles of bevacizumab as novel drug delivery to target retina: optimization, characterization, and in vitro toxicity evaluation

“…Drug injected into SC space has direct access into intraocular tissue through transscleral route and fast clearance via conjunctival blood and lymphatic flow Urtti 2006, Amrite et al 2008). Larger accessible surface area of sclera and a high degree of hydration make it permeable to hydrophilic substances (Srikantha et al 2012). Scleral matrix is having proteoglycans which contributes to negative charge on the sclera and may bind to positively charged molecules (Ranta et al 2010).…”