Abstract
Purpose :
The earliest hallmarks of DR is the loss of retinal pericytes through apoptosis. Human retinal pericytes (HRP) are contractile cells that share a common basement membrane with retinal endothelial cells (REC) and provide support for REC. HRP are most prominent in retinal capillaries in comparison with capillaries in other tissues in the body. HRP envelop the REC, and with their contractile ability, aid in maintaining tubular integrity and control the hydrostatic pressure of the microcirculation. The loss of HRP leads to development of advanced-stage DR pathology including angiogenesis. However, the mechanism by which interactions between HRP and REC leads to either cell proliferation or cell survival remains unclear. In this present study, HRP and REC in coculture were examined in order to evaluate effects of their interactions in terms of cell growth inhibitions.
Methods :
In a well of 24-well plate, a transwell insert holding 40,000 cells grown on a PET membrane in the upper chamber (500ul Complete Cell Media) and 60,000 cells grown onto the well (lower chamber, 1ml CCM) fashioned a two-cell type (non-contact) co-culture system of HRP and REC. In contrast, a one-cell type mono-culture system consists of 60,000 cells (either HRP or REC) grown in a well of a 24 well plate with a transwell insert and PET membrane only. Cell culture media in both chambers were changed every 24hrs. The number of viable cells were determined using a trypan blue dye exclusion method (% viability: 70-94). Growth curves for HRP and EC in the mono- and co-culture system were obtained using the number of viable cells recorded at 0, 24, 48 and 72hrs. Means were derived from triplicated observations.
Results :
Results from mono-culture system show that HRP had a slightly shorter population doubling time (DT) compared to REC (37hr for HRP; 48hr for REC). When HRP were co-cultured with REC, the DT of HRP in the transwell increased significantly from that observed in the monoculture system (DT of 37hr to 119hr), while the DT of REC remained unchanged (48hr). When REC were cocultured with HRP, the number of REC (from the transwell) diminished from the initially seeded 40,000 cells, while the growth rate of the HRP remains similar (DT: 34hr).
Conclusions :
Results suggest that HRP and EC exhibited a significant decrease of cell growth in a co-culture system compared to a monoculture system. Additional experiments will be conducted to further substantiate this conclusion.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.