Anticoagulant and Thrombotic Activities of dHG-5 and its Containing Oliogosaccharides
Anticoagulant activity. The anticoagulant activities of dHG-5 and its oligosaccharides were evaluated as their effects on APTT, PT and TT of human coagulation control plasma.
dHG-5 had small EC2.0×. For dHG-5 containing oligosaccharides, oHG-5 had the largest EC2.0×, the EC2.0× of oligosaccharides with dp 8~14 decreased significantly with the extending of their chain length, while the EC2.0× of oligosaccharides with dp 17~29 had no large change, and the EC2.0× of oHG-17 approximated to that of dHG-5 (Figure 5B). dHG-5 and its oligosaccharides had no obvious effects on plasma PT and TT (Table S2). The relationship between Mw (x ) and EC2.0× of dHG-5 containing oligosaccharides (y ) was fitted well with the power function: y = 1.33 × 1011 × x-2.92 (R2 = 0.999) (Figure 5B). Further calculation showed that the anticoagulant potency of dHG-5 (100 U, used as standard) was close to the weighted average sum of that of its containing oligosaccharides (Table 1)
Nagase H. reported that dFG had the anti-f.IIa activity in the presence of heparin cofactor II (HCII) besides anti-iXase activity (Nagase, Enjyoji et al. , 1995), our group also confirmed this (Wu, Wenet al. , 2015). To evaluate the effects of AT-III and HCII on the anticoagulant activities of dHG-5 and its containing oligosaccharides, AT-III- or HCII-deficient plasmas were used for assays. Compared with AT-III-deficient plasma, oHG-5, oHG-17, oHG-29 and dHG-5 didn’t increase the APTT of AT-III-complementary plasma. And the APTT-prolonging activities of oHG-8, oHG-17, oHG-29 and dHG-5 in HCII-deficient plasma approached to those in coagulation control plasma (Figure 5C-D).
Antithrombotic activity. The antithrombotic activities of dHG-5 and its containing oligosaccharides were evaluated using the rat deep venous thrombosis model.
The results showed that after subcutaneously injecting, dHG-5 dose-dependently inhibit the venous thrombosis induced by stasis and thromboplastin, with the ED50 of 2.38 mg/kg (Figure 6A). To further compare the antithrombotic activities of dHG-5 containing oligosaccharides, their antithrombotic activities were evaluated at the equivalent anticoagulant doses. The result showed that, at the equivalent anticoagulant doses, the thrombosis inhibition effects of these oligosaccharides were not significantly different among treated groups, and closed to 50% except oHG-5 and -29 (Figure 6B). Further calculation showed that the antithrombotic potency of dHG-5 (100 U, used as standard) was close to the weighted average sum of that of its containing oligosaccharides (Table 1).