Abstract
Purpose :
Available treatments for retinal tissue degeneration resulting in vision loss are unreliable, invasive, and do not address the root cause of disease. Cell transplantation has been demonstrated as a feasible approach for a lifelong treatment to repair and halt further retinal degeneration, however, results have been mostly ineffective and exhibit poor survival and integration. Thermogelling copolymers can be optimized as cell delivery vehicles to enhance cell viability and engraftment following sub-retinal administration.
Methods :
N-isopropylacrylamide, acrylamide, acrylic acid N-hydroxysuccinimide, and (r)-α-acryloyloxy-β,β-dimethyl-γ-butyrolactone (pNAND) copolymers were synthesized using an established method in different ratios (A,B,C). Bovine fibronectin (FN) and/or phosphate buffered saline (PBS) were used to modify the polymers and reduce the prevalence of reactive groups. The temperature at which the polymer transitions to a gel (lower critical solution temperature) was assessed using differential scanning colourimetry [Fig. 1]. The viability and proliferation rates of human and rat retinal pigment epithelial cells (ARPE-19) cultured in 2-dimensional (2D) and 3-dimensional (3D) copolymer gel constructs were assessed with an MTT assay, and fluorescent cell imaging [Fig. 2]. Differentiation of retinal pigment epithelial (RPE) cell lines using established methods is currently underway, to be followed with immunocytochemistry methods for detection of mature RPE expression markers ZO-1, CRALBP, and RPE65.
Results :
Synthesized pNAND copolymers and FN-pNAND copolymers gel at physiological temperatures [Fig. 1]. Viability and proliferation assays of ARPE-19 cells demonstrated that pNAND and pNAND-FN prepared in 20% w/v PBS and DMEM provided viable environments for 2D cell survival and proliferation, and 3D cell survival in vitro [Fig. 2]. Preliminary results indicate that RPE cell lines cultured in the presence of pNAND can differentiate into mature retinal epithelium and will be confirmed with immunocytochemistry.
Conclusions :
Thermogelling pNAND copolymers have the potential to function as an injectable cell delivery scaffold to improve the survival and proliferation of transplanted cells in the retina. Further investigations are necessary to evaluate efficacy in vivo.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.