1. Distinct dose-dependent effects of plasmin and TPA on coagulation and hemorrhage
by Daphne Stewart, Mansze Kong, Valery Novokhatny, Gary Jesmok, and Victor J. Marder
Blood
Volume 101(8):
April 15, 2003
©2003 by American Society of Hematology

2. Schematic representation of the experimental design
Schematic representation of the experimental design.Diagram shows the bleeding times (arrows) and blood samples (*) relative to the 50- to 60-minute infusion of plasmin or TPA. Infusions into rabbits (4 kg body weight) were performed in a randomized, blinde...
Schematic representation of the experimental design.Diagram shows the bleeding times (arrows) and blood samples (*) relative to the 50- to 60-minute infusion of plasmin or TPA. Infusions into rabbits (4 kg body weight) were performed in a randomized, blinded manner, using coded vials containing plasmin (16, 24, or 32 mg) or TPA (1, 2, or 4 mg) in a total volume of 10 mL. Data were pooled by group for statistical analysis, without knowledge of treatment. Control infusions of 10 mL saline to 5 animals were performed in an open-label manner at the end of the study.
Daphne Stewart et al. Blood 2003;101:
©2003 by American Society of Hematology

3. Schematic representation of bleeding
Schematic representation of bleeding.Bleeding is diagramed before, during, and after infusion of plasmin (left) or TPA (right) to groups of 5 rabbits.
Schematic representation of bleeding.Bleeding is diagramed before, during, and after infusion of plasmin (left) or TPA (right) to groups of 5 rabbits. PBTs were performed on cohorts of 5 animals at −30 minutes, −10 minutes, and 0 minutes before infusion and at 10, 30, 60, 70, 90, 120, and 180 minutes after the start of infusion. The vertical shaded rectangle represents the infusion period from 0 to 55 to 60 minutes. The heavy lines indicate when bleeding occurred, either the primary bleeding times shown at the extreme left margin of each line, or renewed bleeding at these same sites over the 180-minute observation period. Intervals of stable hemostasis are indicated by the light horizontal lines.
Daphne Stewart et al. Blood 2003;101:
©2003 by American Society of Hematology

4. Total bleeding induced by TPA or plasmin
Total bleeding induced by TPA or plasmin.Results shown for 5 animals that received 0.25, 0.5, or 1 mg/kg TPA (left panel) or 4, 6, or 8 mg/kg of plasmin (right panel).
Total bleeding induced by TPA or plasmin.Results shown for 5 animals that received 0.25, 0.5, or 1 mg/kg TPA (left panel) or 4, 6, or 8 mg/kg of plasmin (right panel). Total bleeding is calculated for individual animals as the sum of bleeding detected from all 10 puncture sites (Figure 2).
Daphne Stewart et al. Blood 2003;101:
©2003 by American Society of Hematology

5. Total bleeding induced by TPA or plasmin
Total bleeding induced by TPA or plasmin.Total bleeding induced by TPA (1 mg/kg) or plasmin (8 mg/kg) relative to the time during which agent was infused.
Total bleeding induced by TPA or plasmin.Total bleeding induced by TPA (1 mg/kg) or plasmin (8 mg/kg) relative to the time during which agent was infused. Results are expressed as mean of 5 animals for either treatment group. The preinfusion ear puncture bleeding times were performed at −30, −10, and 0 minutes before starting TPA or plasmin. During the 55- to 60-minute infusions, ear punctures were induced at 10 and 30 minutes; postinfusion bleeding times were performed at 60, 70, 90, and 120 minutes. TPA (░) caused increased bleeding from the sites induced before or during infusion, most evident at the 10-minute site, whereas plasmin (▪) caused bleeding from midpoint of infusion and at all subsequent ear puncture sites. Results for bleeding at 180 minutes are not shown because experiments were terminated just after these observations.
Daphne Stewart et al. Blood 2003;101:
©2003 by American Society of Hematology

6. Concentrations of plasma antiplasmin, factor VIII, and fibrinogen
Concentrations of plasma antiplasmin, factor VIII, and fibrinogen.Plasma antiplasmin (top), factor VIII (middle), and fibrinogen concentrations (bottom) in animals exposed to TPA (open symbols) or plasmin (solid symbols) over the course of the 180-minute ob...
Concentrations of plasma antiplasmin, factor VIII, and fibrinogen.Plasma antiplasmin (top), factor VIII (middle), and fibrinogen concentrations (bottom) in animals exposed to TPA (open symbols) or plasmin (solid symbols) over the course of the 180-minute observation. Mean results for each cohort of 5 animals expressed as percent of pretreatment value at −30 minutes, normalized to 100%. Data are corrected for results obtained with saline alone, which was associated with a 5% decrease in all results during the 60-minute infusion and with a 10% decrease in values of samples obtained from 60 to 180 minutes. Results for TPA were not different at dosages of 0.25 mg/kg (○), 0.5 mg/kg (■), or 1.0 mg/kg (⋄). Plasmin showed a dose-dependent decrease in all 3 assays on exposure to increasing dosages of 4 mg/kg (♦), 6 mg/kg (▪), and 8 mg/kg (●). Antiplasmin activity was fully depleted with plasmin at 6 mg/kg, a dose that decreased fibrinogen and factor VIII to 20% to 30% of initial activity. Only at a dose of 8 mg/kg did plasmin cause depletion of fibrinogen and factor VIII to levels at or below the lower limit of quantification.
Daphne Stewart et al. Blood 2003;101:
©2003 by American Society of Hematology

7. Fibrinogen concentrations compared with total bleeding
Fibrinogen concentrations compared with total bleeding.Nadir fibrinogen concentrations (top) are compared with total bleeding (bottom) in animals receiving TPA or plasmin.
Fibrinogen concentrations compared with total bleeding.Nadir fibrinogen concentrations (top) are compared with total bleeding (bottom) in animals receiving TPA or plasmin. Control values for nadir fibrinogen concentrations, at 0 mg/kg of plasmin or TPA, were determined in plasma samples obtained before infusion (at −30 and 0 minutes). Control values for total bleeding were determined in the 5 rabbits that received saline (mean of 50 determinations, 2.25 ± 0.4 minutes). Decrease in plasma fibrinogen concentration with plasmin was dose-dependent and more profound than with TPA, which did not alter the fibrinogen concentration significantly from the pretreatment level. Nevertheless, TPA induced bleeding in a stepwise manner at increasing dosages, unrelated to the plasma fibrinogen concentration. Plasmin did not induce excessive bleeding at 4 or 6 mg/kg. At a plasmin dose of 8 mg/kg, the nadir fibrinogen concentration dropped below the lower limit of quantification, in association with the appearance of excessive bleeding.
Daphne Stewart et al. Blood 2003;101:
©2003 by American Society of Hematology