Abstract
The positive role of platelet gel (PG) in tissue regeneration is well known, however, other characteristics of PG still remain to be determined. We investigated cellular and molecular changes in cultured human retinal pigment epithelial (hRPE) cells when treated with different concentrations of PG named PG1, PG2, and PG3. hRPE cells were isolated from donor eyes of two newborn children, within 24 hours after their death. The cells were treated with three concentrations of PG for 7 days: 3 × 104/ml (PG1), 6 × 104/ml (PG2), and 9 × 104/ml (PG3). Fetal bovine serum was used as a control. Immunocytochemistry was performed with anti-RPE65 (H-85), anti-Cytokeratin 8/18 (NCL-5D3), and anti-PAX6 antibody. We used MTT assay to determine cell viability. Gene expressions of PAX6, MMP2, RPE65, ACTA2, MKI67, MMP9, and KDR were analyzed using real-time PCR. A significant increase in viability was observed for PG3-treated cells compared to control (p = 0.044) and compared to PG1 group (p = 0.027), on day 7. Cellular elongation together with dendritiform extensions were observed in PG-treated cells on days 1 and 3, while epithelioid morphology was observed on day 7. All cells were immunoreactive for RPE65, cytokeratin 8/18, and PAX6. No significant change was observed in the expression of MKI67 and PAX6, but the expressions of MMP2, MMP9, ACTA2, and KDR were significantly higher in PG2-treated cells compared to controls (p < 0.05). Our results indicate that increased concentration of PG and extended exposure time have positive effects on viability of hRPE cells. PG may be useful for hRPE cell encapsulation in retinal cell replacement therapy.
Highlights
The retinal pigment epithelium (RPE) is a layer of tightly connected hexagonal cells, which supports the structural and functional integrity of the photoreceptors and exerts a critical role in the visual process [1]
All platelet gel (PG)‐treated cells were immunoreactive for cytokeratin 8/18 (Figure 1E), RPE65 (Figure 1I), and PAX6 (Figure 1M), and this was not significantly different from the corresponding controls (Figure 1G, K, and O, respectively)
Our study demonstrated for the first time that increased concentrations of PG and extended exposure time have positive effects on the viability of human retinal pigment epithelial (hRPE) cells
Summary
The retinal pigment epithelium (RPE) is a layer of tightly connected hexagonal cells, which supports the structural and functional integrity of the photoreceptors and exerts a critical role in the visual process [1]. Given the viability of RPE cells in culture media, absence of synaptic connections between the cells and photoreceptors, and easy imaging of the outer retina, novel cell replacement therapies based on these cells have recently emerged for the treat‐ ment of AMD [4,5]. In this regard, all attempts are applied to increase the viability and improve the quality of the cells in a culture, to achieve the best results [6]
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