1. AbstractHeparin cofactor II (HCII) inhibits thrombin rapidly in the presence of dermatan sulfate, heparan sulfate, or heparin. It is present in mouse and human plasma at micromolar concentrations. Although the physiologic function of HCII is unknown, it has been proposed to inhibit coagulation (e.g., in the placenta) or to participate in diverse processes such as inflammation, atherosclerosis, and wound repair. To investigate the physiologic function of HCII, we deleted ~2 kb of the mouse HCII gene by homologous recombination in 129/SvJ embryonic stem cells. The deletion included exon 1, which encodes the N-terminal half of the protein. Crosses of heterozygous HCII animals in a mixed 129/SvJ x C57Bl/6 genetic background produced 154 offspring at the expected Mendelian ratio (25.3% HCII+/+, 50.6% HCII+/-, 24.0% HCII-/-). Thus, homozygous HCII deficiency is not lethal in utero. At 11-12 weeks of age, the homozygous HCII-deficient animals were grossly indistinguishable from their wild-type and heterozygous littermates in weight, appearance, and survival. Biochemical assays of plasma confirmed the absence of HCII activity (i.e., dermatan sulfate-dependent thrombin inhibition) in the HCII-/-animals (0.00±0.03 µM, mean±SD) in comparison with HCII+/- (0.16±0.02 µM) and HCII+/+ (0.36±0.05 µM) animals. Antithrombin activity (i.e., heparin-dependent factor Xa inhibition) was similar in all 3 genotypes (1.90-2.03 µM). Crosses of homozygous HCII-/- mice produced litters of similar size (6.8±3.0 pups/litter, n=11 litters) to those obtained from heterozygous matings (6.7±3.1 pups/litter, n=38 litters). Thus, homozygous HCII deficiency in both parents is compatible with normal gestation in a mixed genetic background. Backcrosses are being performed to obtain HCII-deficient mice in pure 129/SvJ and C57Bl/6 strains, and experiments to test the effects of HCII deficiency in various models of pathology are underway. In summary, HCII-deficient mice are viable and fertile. These mice will provide a tool to elucidate the physiologic function of HCII.