Fibronectin in the tear film.
OBJECTIVE
Fibronectin plays an important role in corneal healing and has been detected previously in the tear film. To investigate the levels of fibronectin in normal human tears, the authors measured and compared fibronectin concentration in open-eye, closed-eye, and reflex tear fluid. The origin of fibronectin in the tear film was investigated by comparing fibronectin concentration in sequentially collected reflex tear samples with the concentrations of total protein and albumin in the same samples.


METHODS
Open-eye and closed-eye tears were collected from 11 noncontact lens wearers. From 7 subjects, 20 uninterrupted reflex tear samples (10 microliters each) subsequently were collected, using the sneeze reflex method of stimulation, followed by an additional 10 nonstimulated tear samples (3 microliters each) immediately after cessation of stimulus. Enzyme-linked immunosorbent assays were used to determine fibronectin and albumin concentrations, and bicinchoninic acid protein assays were used to determine total protein concentration in each sample.


RESULTS
Fibronectin concentration in open-eye tears (19 +/- 24 eta g/ml, range 3 to 78 eta g/ml) was significantly different (P = 0.004) from that in closed-eyes tears (4127 +/- 3222 eta g/ml, range 1177 to 11384 eta g/ml). In the first 50 microliters of reflex tears, fibronectin concentrations were low (10 +/- 23 eta g/ml), but they increased significantly (P = 0.028) after 100 microliters of reflex tears had been collected (220 +/- 126 eta g/ml). There was a further marked transient increase (767 +/- 946 eta g/ml) after cessation of stimulus. Total protein concentration in the same samples decreased significantly during reflex tear collection compared to open-eye tears, and it increased gradually after cessation of stimulus. Albumin concentration in the same samples, analyzed for two subjects only, showed a pattern similar to that for fibronectin. Dilation of conjunctival blood vessels was noted in all subjects after reflex tear collection. Administration of a topical vasoconstrictor in two subjects eliminated the increase in fibronectin concentration during reflex tearing but did not affect total protein concentration. Under reducing conditions, the molecular mass of fibronectin in open-eye and reflex tears was 240 kDa, identical to commercially available purified plasma fibronectin, whereas fibronectin in closed-eye tears was degraded into small molecular mass fragments.


CONCLUSIONS
These findings suggest that fibronectin in tear fluid is derived from plasma and that the increase in concentration in closed-eye and reflex tear fluid is caused by leakage from dilated conjunctival blood vessels.
Fukuda
M
M
Cooperative Research Centre for Eye Research and Technology, University of New South Wales, Sydney, Australia.
Fullard
R J
RJ
Willcox
M D
MD
Baleriola-Lucas
C
C
Bestawros
F
F
Sweeney
D
D
Holden
B A
BA
D000328:Adult; D000418:Albumins / Q000032:analysis; D001767:Blinking; D002199:Capillary Permeability; D003228:Conjunctiva / Q000098:blood supply; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005136:Eye Proteins / Q000032:analysis; D005260:Female; D005353:Fibronectins / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D008297:Male; D013666:Tears / Q000737:chemistry*
D000418:Albumins; D005136:Eye Proteins; D005353:Fibronectins; C042434:tear proteins