Mechanical Strain Induces Distinct Human Scleral Fibroblast Lineages: Differential Roles in Cell Proliferation, Apoptosis, Migration and Differentiation

Purpose: The purpose of this study was to explore the effect of mechanical strain on human scleral fibroblasts (HSFs) and compare cell behaviors of HSFs from distinct regions. Methods: Primary HSFs were cultivated using a digestive protocol. Cells were seeded on collagen I-coated Bioflex plates, and a FX-5000 tension system was used to perform biaxial mechanical strain in vitro. We applied 10%, 0.5-Hz mechanical strain. Cell behaviors of peripapillary and periphery HSFs were compared after the strain. Edu imaging, Cell Counting Kit-8 assay, and cell cycle flow cytometry were conducted to analyze cell proliferation ability. For cell apoptosis, flow cytometry of Annexin V/propidium iodide, caspase 3 activity, and Western blot were performed. Immunofluorescence, real-time PCR, and Western blot were used to investigate cell differentiation. A migration assay was also performed. Results: Under the mechanical strain of 10%, 0.5 Hz for 24 hours, the proliferation ability and cell apoptosis of peripapillary HSFs did not have a significant change. The expression of alpha-smooth muscle actin (α-SMA) slightly decreased. However, increased cell proliferation, attenuated cell apoptosis and more expression of α-SMA were shown in the periphery HSFs under the same condition. The migration rate was also increased for periphery HSFs, whereas it kept almost the same for peripapillary HSFs under 10%, 0.5-Hz strain for 8 hours. Conclusions: Mechanical strain affected the cell behaviors of HSFs. The different performance of cells from distinct regions may suggest familial linages of HSFs, probably induced by mechanical strain.