α2-antiplasmin gene deficiency in mice is associated with enhanced fibrinolytic potential without overt bleeding

alpha(2)-antiplasmin (alpha(2)-AP) is the main physiologic plasmin inhibitor in mammalian plasma. Inactivation of the murine alpha(2)-AP gene was achieved by replacing, through homologous recombination in embryonic stem cells, a 7-kb genomic sequence encoding the entire murine protein (exon 2 through part of exon 10, including the stop codon) with the neomycin resistance expression cassette. Germline transmission of the mutated allele was confirmed by Southern blot analysis. Mendelian inheritance of the inactivated alpha(2)-AP allele was observed, and homozygous deficient (alpha(2)-AP(-/-)) mice displayed normal fertility, viability, and development. Reverse transcription-polymerase chain reaction confirmed the absence of alpha(2)-AP mRNA in kidney and liver from alpha(2)-AP(-/-) mice, in contrast to wild-type (alpha(2)-AP(+/+)) mice. Immunologic and functional alpha(2)-AP levels were undetectable in plasma of alpha(2)-AP(-/-) mice, and were about half of wild-type in heterozygous littermates (alpha(2)-AP(+/-)). Other hemostasis parameters, including plasminogen activator inhibitor-1, plasminogen, fibrinogen, hemoglobin, hematocrit, and blood cell counts were comparable for alpha(2)-Ap(+/+), alpha(2)-AP(+/-), and alpha(2)-AP(-/-) mice. After amputation of tail or toe tips, bleeding stopped spontaneously in alpha(2)-AP(+/+), as well as In alpha(2)-AP(+/-) and alpha(2)-AP(-/-) mice. Spontaneous lysis after 4 hours of intravenously injected I-125-fibrin-labeled plasma clots was significantly higher in alpha(2)-AP(-/-) than in alpha(2)-AP(+/+) mice when injecting clots prepared from alpha(2)-AP(+/+) plasma (78% +/- 5% v 46% +/- 9%; mean +/- SEM, n = 6 to 7; P = .02) or from alpha(2)-AP(-/-) plasma (81% +/- 5% v 46% +/- 5%; mean +/- SEM, n = 5; P = .008), Four to 8 hours after endotoxin injection, fibrin deposition in the kidneys was significantly reduced In alpha(2)-AP(-/-) mice, as compared with alpha(2)-AP(+/+) mice (P less than or equal to .005), Thus, alpha(2)-AP(-/-) mice develop and reproduce normally; they have an enhanced endogenous fibrinolytic capacity without overt bleeding. (C) 1999 by The American Society of Hematology.