Abstract
Purpose :
To determine the importance of the dystrophin-glycoprotein complex (DGC) in lens cytoarchitecture, mechanics and function using an ENU (N-ethyl-N-nitrosourea)-induced dystrophin-deficient mouse model (mdx3cv).
Methods :
Expression and distribution profiles of Dp71 in embryonic and adult mouse (C57BL/6) lenses were determined by immunoblot, immunofluorescence, and PCR analyses. Dp71 colocalization with different proteins was assessed using equatorial mouse lens sections. To identify extracellular ligands of the DGC complex, the lens fiber mass derived ECM-enriched fraction from adult mice was analyzed by mass spectrometry. Changes in lens stiffness of control and mdx3cv mice obtained from the Jackson laboratory (maintained on a C57BL/6 genetic background) were determined using a microstrain analyzer.
Results :
PCR and immunoblot analyses revealed that dystrophin isoforms- Dp427, Dp260, Dp140, and Dp40 are expressed at moderate levels relative to the high levels of Dp71 in P1 lenses. Dp71 was the predominant dystrophin isoform expressed in P21 and P90 mouse lenses. Dystrophin was found to be induced robustly during lens fiber cell differentiation. In equatorial sections, Dp71 staining presents as a large cluster at the center of the long arm with some localization to the vertices of the hexagonal fiber cells, colocalizing with b-dystroglycan and connexin-50 at the large clusters. Mdx3cv mouse lenses (P1 and older) were confirmed to be deficient in Dp71 (by >90%) but showed no changes in growth and transparency up to 6 months. However, the levels of NrCAM and connexin-50 were significantly decreased in the Dp71 deficient mouse (1 month-old) lens fiber cell membrane fraction relative to control lenses. Additionally, compressive stress/strain analyses of intact lenses (4 month-old) revealed a significant decrease in Young’s modulus in mdx3cv mice compared to wild-type lenses. Perlecan is found to be a major ECM protein in the mature lens fibers, a known ligand of α-dystroglycan. Age-dependent analyses of lens function in the mdx3cv mouse are currently in progress.
Conclusions :
Taken together, this ongoing study using mdx3cv mice reveals that Dp71 is required for lens membrane organization and maintenance of biomechanics.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.