Transscleral diffusion of ethacrynic acid and sodium fluorescein.
OBJECTIVE
One of the current limitations in developing novel glaucoma drugs that target the trabecular meshwork (TM) is the induced corneal toxicity from eyedrop formulations. To avoid the corneal toxicity, an alternative approach would be to deliver TM drugs through the sclera. To this end, we quantified ex vivo diffusion coefficient of a potential TM drug, ethacrynic acid (ECA), and investigated mechanisms of ECA transport in the sclera.


METHODS
An Ussing-type diffusion apparatus was built to measure the apparent diffusion coefficient of ECA in fresh porcine sclera at 4 degrees C. To understand mechanisms of ECA transport, we quantified the transscleral transport of a fluorescent tracer, sodium fluorescein (NaF), that has a similar molecular weight but is more hydrophilic compared to ECA. Furthermore, we developed a mathematical model to simulate the transport processes and used it to analyze the experimental data. The model was also used to investigate the dependence of diffusion coefficients on volume fraction of viable cells and the binding of NaF and ECA to scleral tissues.


RESULTS
The diffusion coefficients of ECA and NaF in the sclera were 48.5+/-15.1 x 10-7 cm(2)/s (n=9) and 5.23+/-1.93 x 10(-7) cm(2)/s (n=8), respectively. Both diffusion coefficients were insensitive to cell shrinkage caused by ECA during the diffusion experiments and cell damage caused by the storage of tissues ex vivo before the experiments. Binding of ECA to scleral tissues could not be detected. The apparent maximum binding capacity and the apparent equilibrium dissociation constant for NaF were 80+/-5 mM and 2.5+/-0.5 mM (n=3), respectively.


CONCLUSIONS
These data demonstrated that ECA diffusion was minimally hindered by structures in the sclera, presumably due to the lack of cells and binding sites for ECA in the sclera.
Lin
Cheng-Wen
CW
Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA. cl28@duke.edu <cl28@duke.edu>
Wang
Yong
Y
Challa
Pratap
P
Epstein
David L
DL
Yuan
Fan
F
D000818:Animals; D001692:Biological Transport; D002470:Cell Survival; D002829:Choroid / Q000378:metabolism; D004058:Diffusion; D004798:Enzymes; D004976:Ethacrynic Acid / Q000378:metabolism / Q000493:pharmacokinetics*; D019793:Fluorescein / Q000378:metabolism / Q000493:pharmacokinetics*; D005456:Fluorescent Dyes / Q000378:metabolism / Q000493:pharmacokinetics*; D006652:Histological Techniques; D066298:In Vitro Techniques; D008954:Models, Biological; D009994:Osmolar Concentration; D012590:Sclera / Q000166:cytology / Q000378:metabolism* / Q000502:physiology; D013552:Swine; D013997:Time Factors; D014018:Tissue Distribution
D004798:Enzymes; D005456:Fluorescent Dyes; D004976:Ethacrynic Acid; D019793:Fluorescein