Abstract
Purpose :
The pathology of glaucoma is characterized by loss of retinal ganglion cells (RGCs). Since loss of these specialized neurons is irreversible, it is urgent to develop treatments that protect these cells. For development of neuroprotective drugs, an in vitro glaucoma model would be very useful. The purpose of this study is to develop such a model by creating a device in which RGCs can be cultured and exposed to conditions that mimic those of elevated intra-ocular pressure (IOP), the main risk factor of glaucoma.
Methods :
Elevated IOP leads to increased hydrostatic pressure and deformation in eye tissues. Moreover, IOP fluctuates considerably in vivo; deformation is therefore dynamic. To mimic these conditions, we developed a cell culture device, based on Sinha et al. (Lab Chip 2015; 15 429-439). The device is made of polydimethylsiloxane (PDMS), contains 100 culture units, and is compatible with standard well plate holders. A culture unit consists of a plateau surrounded by a trench, all covered by an elastic membrane on which the cells are grown. The device can simultaneously apply hydrostatic pressure and strain. Pressure is applied by elevating a reservoir of culture medium, while strain is applied dynamically with a vacuum pump creating a pressure drop in the trench, which leads to stretching of the membrane and the cells. We used the neuronal PC-12 cell line to establish model biocompatibility.
Results :
Our device can screen a range of hydrostatic pressures (0-90 mmHg). By designing different shapes of the plateau we created different strain conditions (uni-axial, isotropic) assisted by computational modeling. Actual strains were measured using tracking of beads embedded in the elastic membrane. Strains measured 0 to 20% depending on the magnitude of the applied pressure drop and the dimensions of the trench in the particular culture unit. First experiments with PC-12 cells indicate that cyclic (1 Hz) strain (10%) applied for 48 hours under 70mmHg resulted in a reduced cell density compared to control.
Conclusions :
We constructed a medium-throughput cell culture device that can simultaneously apply pressure and strain to cultured cells. This glaucoma-on-a-chip can help to identify the molecular mechanisms of IOP-induced RGC death and design and test neuroprotective drugs. In addition, it can serve to characterize sensitivity of patient cells to mechanical stress.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.