Vulnerability of corneal endothelial cells to mechanical trauma from indentation forces assessed using contact mechanics and fluorescence microscopy

Abstract

Corneal endothelial cell (CEC) loss occurs from tissue manipulation during anterior segment surgery and corneal transplantation as well as from contact with synthetic materials like intraocular lenses and tube shunts. While several studies have quantified CEC loss for specific surgical steps, the vulnerability of CECs to isolated, controllable and measurable mechanical forces has not been assessed previously. The purpose of this study was to develop an experimental testing platform where the susceptibility of CECs to controlled mechanical trauma could be measured. The corneal endothelial surfaces of freshly dissected porcine corneas were subjected to a range of indentation forces via a spherical stainless steel bead. A cell viability assay in combination with high-resolution fluorescence microscopy was used to visualize and quantify injured/dead CEC densities before and after mechanical loading. In specimens subjected to an indentation force of 9 mN, the mean ± SD peak contact pressure P was 18.64 ± 3.59 kPa (139.81 ± 26.93 mmHg) in the center of indentation and decreased radially outward. Injured/dead CEC densities were significantly greater (p ≤ 0.001) after mechanical indentation of 9 mN (167 ± 97 cells/mm) compared to before indentation (39 ± 52 cells/mm) and compared to the sham group (34 ± 31 cells/mm). In specimens subjected to "contact only" - defined as an applied indentation force of 0.65 mN - the peak contact pressure P was 7.31 ± 1.5 kPa (54.83 ± 11.25 mmHg). In regions where the contact pressures was below 78% of P (<5.7 kPa or 42.75 mmHg), injured/dead CEC densities were within the range of CEC loss observed in the sham group, suggesting negligible cell death. These findings indicate that CECs are highly susceptible to mechanical trauma via indentation, supporting the established "no-touch" policy for ophthalmological procedures. While CECs can potentially remain viable below contact pressures of 5.7 kPa (42.75 mmHg), this low threshold suggests that prevention of indentation-associated CEC loss may be challenging.

Unraveling the mechanobiology of cornea: From bench side to the clinic

“…In young adults, the endothelial cell density (ECD) within a healthy cornea is approximately 3,000–4,000 cells/mm 2 . However, an abnormal cornea loses its ability to pump sufficient water to maintain its function when ECD decreases below 750–500 cells/mm 2 ( Ramirez-Garcia et al, 2018 ).…”

“…Furthermore, mechanical compression interferes with the maintenance of normal stromal structures ( Figure 3C ). Ramirez-Garcia et al (2018) evaluated the response of corneal endothelial cells to the indentation forces in vitro and found that the damage/apoptosis of CEnCs increased and ECD decreased significantly when the contact pressure exceeded 5.7 kPa (42.75 mmHg). When acute ocular hypertension (∼82.6 mmHg for 2 h) was induced, the ECD was significantly decreased, and CEnCs became irregular and multiform with disrupted ZO-1 and F-actin ( Li et al, 2017 ).…”

The use of lens anterior capsule disc for corneal endothelium protection during femtosecond laser-assisted cataract surgery in eyes with low endothelial cell density

“…19 When cataracts are extracted, for example, lens fragments can collide with the endothelium. 20 New approaches, such as using newer platforms to minimize power, changing energy mode, using the softshell technique to protect the CECs, and pre-chopping the nucleus with manual methods or the femtosecond laser, can reduce trauma to CEC. 21 , 22 , 23 Chen et al.…”