Structure and function of mouse lens suture examined by 2-photon fluorescence microscopic imaging

Abstract We used two-photon fluorescence microscopy (2PFM) to examine lens fiber and suture architecture, as well as permeability, in wild-type (WT) lenses and in lenses from klotho-like protein homology (KLPH) knockout (KLPH-KO) mice. KLPH is a type I membrane glycoprotein encoded by the Lctl gene; KLPH-KO mice develop lens suture cataracts. Lens sutures have been hypothesized to serve as pathways for transporting ions, nutrients, and other factors as part of the lens microcirculation required to maintain homeostasis and transparency. Three-dimensional (3D) in vivo imaging revealed typical “Y” and “double-Y” anterior suture geometries, along with depth-dependent variation, in WT lenses. In contrast, KLPH-KO lenses exhibited markedly heterogeneous suture morphologies and misalignment between the anterior and posterior suture planes. Quantification of pattern variability using the mean structural similarity index (SSIM) of individual z-stacks relative to the stack mean projection revealed a significant increase in pattern randomization in KLPH-KO lenses ( p < 0.05). Both WT and KLPH-KO lenses displayed voids near sutures and enlarged vacuoles distributed throughout the lens in vivo. Notably, KLPH-KO lenses exhibited irregular, enlarged central voids containing ~ 2–5 μm amorphous structures, consistent with subcellular remnants and/or membrane-associated aggregates. In conclusion, this study provides novel morphological markers for characterizing suture cataracts and associated fiber pathology. It further demonstrates that the dye-impermeable lens suture represents a stabilized interface formed by elongated fiber ends, supporting lens integrity and maintaining the organization of lens fibers.