June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Muc4 Knockout Disrupts the Ocular Surface Mucosal Barrier in Mice
Author Affiliations & Notes
  • Rafael Martínez-Carrasco
    New England Eye Center and Department of Ophthalmology, Tufts Medical Center, Boston, Massachusetts, United States
  • Satyanarayan Rachagani
    Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska, United States
  • Surinder Batra
    Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, Nebraska, United States
  • Pablo Argueso
    New England Eye Center and Department of Ophthalmology, Tufts Medical Center, Boston, Massachusetts, United States
  • M.Elizabeth Fini
    New England Eye Center and Department of Ophthalmology, Tufts Medical Center, Boston, Massachusetts, United States
  • Footnotes
    Commercial Relationships   Rafael Martínez-Carrasco None; Satyanarayan Rachagani None; Surinder Batra Sanguine Diagnostics and Therapeutics, Inc., Code O (Owner); Pablo Argueso None; M.Elizabeth Fini None
  • Footnotes
    Support  R01EY026479
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 3257 – A0292. doi:
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    • Get Citation

      Rafael Martínez-Carrasco, Satyanarayan Rachagani, Surinder Batra, Pablo Argueso, M.Elizabeth Fini; Muc4 Knockout Disrupts the Ocular Surface Mucosal Barrier in Mice. Invest. Ophthalmol. Vis. Sci. 2022;63(7):3257 – A0292.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : The membrane-associated mucins (MAMs) MUC1, MUC4, MUC16, MUC20, MUC21 and MUC22 project from the apical layer of the ocular surface mucosal epithelia and into the tear film (PMID: 33775913; PMID: 31493487). They are thought to provide a barrier against pathogens and other noxious agents, contribute to hydration, and participate in signal transduction and immune suppression. We investigated the specific role of Muc4 at the ocular surface by characterizing Muc4 knockout (KO) mice.

Methods : Muc4 KO mice were compared to their wildtype (WT) littermates. The ocular surface was imaged using the rodent micron IV with slit-lamp attachment, using white or cobalt blue light after staining with fluorescein (detects epithelial cell damage) or rose bengal (detects mucosal barrier disruption). Staining was scored from 0 to 9 using a scoring system modified from van Bijserveld’s. Smoothness was scored from 0-4 considering the alteration of a ring light reflection. The ocular surface was also imaged by scanning electron microscopy (SEM). Gene expression was analyzed by qPCR.

Results : Gross examination revealed no conjunctivitis, blepharitis or other inflammatory disease in KO mice. Fluorescein staining revealed no evidence of epithelial cell damage. In contrast, rose bengal staining was distinctly higher in KO mice than in WT littermates (n=20; P < 0.01), with a punctate pattern indicative of mucosal barrier disruption. SEM revealed more surface epithelial cells with a lower density of microplicae in KO mice (n=8; P < 0.05). Pax6 expression was decreased, while K10 expression was increased (P < 0.05, P < 0.01; respectively), indicating keratinization. In contrast, no increase was detected in expression of cornified envelope genes Sprr1b and Sprr2h. Corneal smoothness was reduced in KO mice (n=18; P < 0.05), suggesting tear film disruption. No compensatory changes due to Muc4 KO were observed in expression of Muc1 or Muc16, or the secreted mucin Muc5ac (n=11).

Conclusions : Cell culture studies have provided evidence that deficiency of a single MAM is sufficient to disrupt the mucosal epithelial barrier at the ocular surface and reduce surface microplicae (PMID: 24968021). This is the first in vivo validation of this finding. We also provide the first evidence that loss of Muc4 results in keratinization, but not cornification of the ocular surface, and leads to tear film disruption.

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

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