Visualization of Intravital Immune Cell Dynamics After Conjunctival Surgery Using Multiphoton Microscopy

Area of Science:

• Ophthalmology
• Immunology
• Microscopy

Background:

• Subconjunctival tissue plays a critical role in ocular wound healing.
• Understanding immune cell dynamics is crucial for developing effective treatments for ocular injuries.

Purpose of the Study:

• To visualize intravital immune cell dynamics in subconjunctival tissue during wound healing using multiphoton microscopy.
• To analyze changes in immune cell number and velocity following conjunctival injury.

Main Methods:

• LysM-enhanced green fluorescent protein (eGFP) mice were used to track immune cells.
• Conjunctival injury was induced using a nylon suture.
• Multiphoton microscopy captured 4D images of subconjunctival tissue over time.
• Imaris software analyzed cell number and velocity.

Main Results:

• Multiphoton microscopy successfully visualized immune cells and blood vessels in vivo.
• The number of LysM-eGFP-positive cells increased in a time-dependent manner post-injury.
• Immune cell velocity significantly increased after injury and remained elevated for 72 hours.

Conclusions:

• This study demonstrates the utility of multiphoton microscopy for visualizing intravital immune cell dynamics in the subconjunctival space.
• This technique provides a valuable tool for studying wound healing mechanisms after ocular injuries, including those from glaucoma surgery.