Abstract
Purpose: :
Neutrophil migration within the inflamed corneal stroma involves contact with extracellular matrix molecules and keratocytes. Cell surface integrin receptors have been implicated in these interactions. While members of β1-, β2-, and β3-integrin families are expressed on extravascular neutrophils, it is reported that in vitro migration in 3-D collagen matrices can be integrin-independent. The purpose of the present study was to use, for the first time, in vivo confocal microscopy for time-lapse recordings to quantify the effect of blocking integrin function on neutrophil motility following corneal epithelial abrasion.
Methods: :
Anesthetized adult C57BL/6 mice received corneal epithelial abrasions using an Alger brush. Immediately after wounding, blocking antibody (anti-β1-, β2-, or β3-integrin) or IgG control was applied to the cornea. Eight hours later, the injured mice were imaged with an HRTIII-RCM for 10 minute at 1 frame per second. Image stabilization, cell tracking and movement analysis were accomplished with a custom MatLab program.
Results: :
Control IgG antibody had no effect on neutrophil motility within the corneal stroma. Neutrophils moved with an average cell speed of 6.93±1.01(SD) µm/minute. Average straight-line index (SI) of the neutrophil population was 0.60±0.07, where a value of 1 indicates a perfectly straight path. While anti-β2- or β3- integrin antibodies had no effect on cell speed or SI, β1-integrin antibody resulted in a 28 % reduction in speed (p<0.05) and a 33% reduction in SI (p<0.05). The resultant vector for average neutrophil movement within a given cornea was toward the wound in 65.5% of the corneas, parallel to the wound in 24.1% and away from the wound in 10.4%, with no difference among experimental groups.
Conclusions: :
β1-integrin, but not β2- or β3-integrin, appears to play a role in neutrophil trafficking within the corneal stroma. Our results showing that β1-integrin blockade only partially reduces neutrophil migration velocity and SI (~30%, respectively), suggest neutrophils are capable of migration through the physically confined environment of the corneal stroma without the benefit of integrin adhesion.
Keywords: cornea: basic science • inflammation • extracellular matrix