Functional, structural, and molecular identification of lymphatic outflow from subconjunctival blebs

Goichi Akiyama, Sindhu Saraswathy, Thania Bogarin, Xiaojing Pan, Ernesto Barron, Tina T. Wong, Mika K. Kaneko, Yukinari Kato, Young Hong, Alex S. Huang

Research output: Contribution to journalArticle

Abstract

The purpose of this study is to evaluate outflow pathways from subconjunctival blebs and to identify their identity. Post-mortem porcine (n = 20), human (n = 1), and bovine (n = 1) eyes were acquired, and tracers (fluorescein, indocyanine green, or fixable/fluorescent dextrans) were injected into the subconjunctival space to create raised blebs where outflow pathways were visualized qualitatively and quantitatively. Rodents with fluorescent reporter transgenes were imaged for structural comparison. Concurrent optical coherence tomography (OCT) was obtained to study the structural nature of these pathways. Using fixable/fluorescent dextrans, tracers were trapped to the bleb outflow pathway lumen walls for histological visualization and molecular identification using immunofluorescence against lymphatic and blood vessel markers. Bleb outflow pathways could be observed using all tracers in all species. Quantitative analysis showed that the nasal quadrant had more bleb-related outflow pathways compared to the temporal quadrant (nasal: 1.9±0.3 pathways vs. temporal: 0.7±0.2 pathways; p = 0.003). However, not all blebs resulted in an outflow pathway (0-pathways = 18.2%; 1-pathway = 36.4%; 2-pathways = 38.6%; and 3-pathways = 6.8%). Outflow signal was validated as true luminal pathways using optical coherence tomography and histology. Bicuspid valves were identified in the direction of flow in porcine eyes. Immunofluorescence of labeled pathways demonstrated a lymphatic (Prox-1 and podoplanin) but not a blood vessel (CD31) identity. Therefore, subconjunctival bleb outflow occurs in discrete luminal pathways. They are lymphatic as assessed by structural identification of valves and molecular identification of lymphatic markers. Better understanding of lymphatic outflow may lead to improved eye care for glaucoma surgery and ocular drug delivery.

Original languageEnglish
Article number108049
JournalExperimental Eye Research
Volume196
DOIs
Publication statusPublished - 2020 Jul

Keywords

  • Aqueous outflow
  • Blebs
  • Conjunctiva
  • Glaucoma surgery
  • Lymphatics

ASJC Scopus subject areas

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience

Fingerprint Dive into the research topics of 'Functional, structural, and molecular identification of lymphatic outflow from subconjunctival blebs'. Together they form a unique fingerprint.

  • Cite this

    Akiyama, G., Saraswathy, S., Bogarin, T., Pan, X., Barron, E., Wong, T. T., Kaneko, M. K., Kato, Y., Hong, Y., & Huang, A. S. (2020). Functional, structural, and molecular identification of lymphatic outflow from subconjunctival blebs. Experimental Eye Research, 196, [108049]. https://doi.org/10.1016/j.exer.2020.108049