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Res Vet Sci. 2017 Apr;111:1-8. doi: 10.1016/j.rvsc.2016.11.004. Epub 2016 Nov 8.

Development of a plasminogen activator inhibitor (PAI-1) assay and comparison of plasma PAI-1 activity in hyperlipidemic/dyslipidemic dogs with either hyperadrenocorticism or diabetes mellitus, and healthy dogs.

Author information

1
Department of Population Medicine and Diagnostic Sciences, College of Veterinary Medicine, Cornell University, 602 Tower Rd, Ithaca, NY 14853, United States. Electronic address: cjw228@cornell.edu.
2
Veterinary Specialists and Emergency Services, 825 White Spruce Blvd, Rochester, NY 14623, United States. Electronic address: kochm@att.net.
3
Department of Population Medicine and Diagnostic Sciences, College of Veterinary Medicine, Cornell University, 602 Tower Rd, Ithaca, NY 14853, United States. Electronic address: eb58@cornell.edu.

Abstract

Thrombosis is a serious complication of many canine diseases and may be related to decreased fibrinolytic potential. Plasminogen activator inhibitor-1 (PAI-1) is the key regulator of fibrinolysis with increased levels demonstrated in states of pro-thrombosis and abnormal lipid metabolism. Our objective was to develop and validate a canine PAI-1 activity assay and test whether dogs with hyperadrenocorticism or diabetes mellitus that are hyperlipidemic/dyslipidemic have increased plasma PAI-1 activity. Functionally active PAI-1 in the plasma sample was incubated with recombinant tissue plasminogen activator (tPA), allowing the formation of a 1:1 stoichiometric inactive complex. Residual unbound tPA was then reacted with excess plasminogen in the presence of a colorimetric plasmin substrate. Plasmin production is quantified by computing the area under the curve of time (x) vs optical density (y) plot and converted to tPA IU/mL by comparison to a calibration curve of tPA standards. PAI-1 activity was determined by calculating the proportion of exogeneous tPA suppressed by PAI-1 in plasma. Assay verification included assessment of linearity, specificity, precision, sensitivity, and stability. PAI-1 activity was increased in hyperlipidemic compared to healthy dogs, but there was no significant difference between dogs with hyperadrenocorticism and diabetes mellitus. A near significant decrease in activity was detected in thawed plasma stored for 20h at 4°C. Our successfully validated assay offers a new tool for investigating fibrinolysis in dogs. Investigation of PAI-1 activity in dogs with other diseases associated with an increased risk of thrombosis would be valuable. Future studies of PAI-1 activity should consider its lability.

KEYWORDS:

Cholesterol; Endocrinopathy; Fibrinolysis; Thrombosis; Triglycerides

PMID:
28266313
DOI:
10.1016/j.rvsc.2016.11.004
[Indexed for MEDLINE]

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