Investigation of fibrillin-1 and LOXL1 in ocular health and disease

Abstract

Purpose : Lysyl oxidase-like 1 (LOXL1) and fibrillin-1 (FBN1) are abundant proteins in exfoliation material (XFM), a hallmark of exfoliation syndrome (XFS). LOXL1 is associated with XFS and many mutations in FBN1 cause Marfan syndrome (MFS). LOXL1 and fibrillin-1 also co-exist in normal lens zonules. Zonule weakness is a common ocular feature shared by XFS and MFS. Here we sought to uncover their involvement in ocular health and disease.

Methods : Loxl1^-/- and Fbn1^C1039G/+ mice on a 129 background were gifts from Drs. Tiansen Li and Hal Dietz and were used to create a double mutant (dbm) line, Fbn1^C1039G/+/Loxl1^-/-. Survival and systemic phenotypes were monitored and anterior segment morphology was measured by SD-OCT. Immunohistochemistry (IHC), transmission electron microscopy (TEM), atomic force microscopy (AFM) and biochemistry approaches were employed.

Results : Fbn1^C1039G/+ mice survive more than 1 year, consistent with being a mild MFS mouse model. Likewise, Loxl1^-/- mice survive beyond 1 year of age. However, dbm mice do not survive beyond 3 months of age due to aorta dilation. Dbm mice also exhibited more pelvic floor organ prolapse. SD-OCT demonstrated thin central corneal thickness (CCT) and increased anterior chamber depth (ACD) in both Fbn1^C1039G/+ and Loxl1^-/- mice. Thinning of CCT and deepening of ACD were more pronounced in dbm mice with normal axial length, indicating compromised zonules, which was confirmed by IHC for MAGP-1. Compared to wt, Loxl1^-/- mice had fewer and abnormally formed elastic fibers, and enlarged and less defined collagen fibrils in their peripapillary sclera (PPS) revealed by TEM. Interestingly, by AFM we observed lower Young’s modulus of cornea in all 3 lines compared to wt. Conversely, Loxl1^-/- mice demonstrated higher Young’s modulus of PPS than wt did. To investigate such differential biomechanical effects due to LOXL1 absence in the cornea and sclera, we used acetic acid collagen extraction which revealed differential collagen solubility and post-translational modification. Lastly, we also observed the differential influence of aging on collagen crosslinking in cornea and sclera.

Conclusions : The phenotypic and mechanistic investigations of our mouse models demonstrated their potential as an invaluable resource for further understanding and treatment of ocular and systemic manifestations of connective tissue disease.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.