Abstract
Purpose :
The corneal endothelium is responsible for corneal clarity; however, it poses a challenge when damaged due to its lack of regenerative potential and reducing cell population as we age. This study implements computer modeling to investigate the feasibility of strategically guiding injected nanoparticle-loaded donor human endothelial cells (HCEC) to the back of the cornea using an external magnetic field. Repopulating the corneal endothelium by injection of HCEC into the anterior chamber may serve as an alternative to surgical corneal transplants.
Methods :
A computer model (Matlab) incorporating the effects of aqueous flow, stokes law, gravity, and magnetic field strength was used to investigate the necessary magnetic field strength and nanoparticle cell loading density to feasibly induce a controlled cellular movement across the anterior aqueous chamber. HCEC were cultured in human endothelial serum free media containing 10 ng/ml FGF-2, and loaded with super paramagnetic iron oxide nanoparticles (SPIONP), based on the results suggested by the computer model. Cell lineage and viability were evaluated using FACS analysis and CyQuant assays. Intracellular iron content was evaluated using Elzone particle analysis and Prussian blue staining. PCR analysis was used to evaluate expression of such HCEC markers as CD200, ACTA alpha, ATPA1, GPC4 and Zo-1 before and after loading. Lastly, SPIONP loaded HCECs were evaluated for inner cornea attachment.
Results :
The computer model indicated that SPIONP-loaded HCEC’s can be directed in a controlled manner to desired locations of the endothelium using a broad range of magnetic forces. HCEC endothelial lineage was confirmed by simultaneous expression of CD200 and glycoproteins. PCR analysis noted a significant difference in ATPA1 by day 3. SPIONP internalization was demonstrated by Prussian blue staining and Elzone particle analysis. Lastly, HCEC maintained similar viability ratios as unloaded control cells, and demonstrated the in vitro ability to attach to the endothelium layer of explant corneas.
Conclusions :
HCEC readily incorporate SPIONPs. Computer modeling helps to minimize SPIONP exposure and facilitates directed cellular movement by indicating appropriate loading values. Future studies will incorporate such parameters as wound size, wound location and curvature of the cornea to further personalize this potential therapeutic treatment specific for the individual.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.