To replicate the human findings and validate the clinical utility of an existing murine model, we investigated the effect of a murine
Pitx2 +/− mutation on corneal thickness.
27 This null mutation is comparable to R69H, which exhibits ∼90% reduction in transactivation compared with wild-type
PITX2 as well as reduced DNA binding.
31 As histologic measurement of murine CCT is associated with appreciable inaccuracy,
8 the human optical coherence tomogram (Stratus OCT, software version 2.0; Carl Zeiss Meditec, Dublin, CA) was adapted for murine use. This involved calibrating OCT measurements of the thickness of uniform plastic films (
n = 9) against those provided by scanning electron microscopy (SEM; Electron Microscope Model 1430; LEO Electron Microscopy, Ltd., Cambridge, UK). For SEM analysis, portions of film were coated with gold (DESK-II cold sputter-etch; Denton Vacuum, Cherry Hill, NJ) before SEM measurements were recorded. For OCT, films were mounted vertically on a precision stand permitting
x,
y, and
z-axis movement. The stand consisted of a rotatable holder on which the sample was mounted and attached via a series of stainless steel rods and a further clamp, to the side bar of the OCT (model numbers: ASC, PR, CR0.5 and MPR; Siskiyou Inc., Grants Pass, OR). The smallest thickness measurement obtained from three scans of each film was used for analysis. The correlation between the OCT and SEM measurements was subsequently determined.