Similar hemostatic responses to hypovolemia induced by hemorrhage and lower body negative pressure reveal a hyperfibrinolytic subset of non-human primates

Abstract

Background: To study central hypovolemia in humans, lower body negative pressure (LBNP) is a recognized alternative to blood removal (HEM). While LBNP mimics the cardiovascular responses of HEM in baboons, similarities in hemostatic responses to LBNP and HEM remain unknown in this species.

Methods: Thirteen anesthetized baboons were exposed to progressive hypovolemia by HEM and, four weeks later, by LBNP. Hemostatic activity was evaluated by plasma markers, thromboelastography (TEG), flow cytometry, and platelet aggregometry at baseline (BL), during and after hypovolemia.

Results: BL values were indistinguishable for most parameters although platelet count, maximal clot strength (MA), protein C, thrombin anti-thrombin complex (TAT), thrombin activatable fibrinolysis inhibitor (TAFI) activity significantly differed between HEM and LBNP. Central hypovolemia induced by either method activated coagulation; TEG R-time decreased and MA increased during and after hypovolemia compared to BL. Platelets displayed activation by flow cytometry; platelet count and functional aggregometry were unchanged. TAFI activity and protein, Factors V and VIII, vWF, Proteins C and S all demonstrated hemodilution during HEM and hemoconcentration during LBNP, whereas tissue plasminogen activator (tPA), plasmin/anti-plasmin complex, and plasminogen activator inhibitor-1 did not. Fibrinolysis (TEG LY30) was unchanged by either method; however, at BL, fibrinolysis varied greatly. Post-hoc analysis separated baboons into low-lysis (LY30 <2%) or high-lysis (LY30 >2%) whose fibrinolytic state matched at both HEM and LBNP BL. In high-lysis, BL tPA and LY30 correlated strongly (r = 0.95; P<0.001), but this was absent in low-lysis. In low-lysis, BL TAFI activity and tPA correlated (r = 0.88; P<0.050), but this was absent in high-lysis.

Conclusions: Central hypovolemia induced by either LBNP or HEM resulted in activation of coagulation; thus, LBNP is an adjunct to study hemorrhage-induced pro-coagulation in baboons. Furthermore, this study revealed a subset of baboons with baseline hyperfibrinolysis, which was strongly coupled to tPA and uncoupled from TAFI activity.

Fig 1. Swarm plot of selected data with baseline (BL) values for blood removal (HEM) and lower body negative pressure (LBNP) trials.

LBNP BL values were obtained 4 weeks after HEM BL. P-values of significantly different parameters between trials as determined by one-way RMANOVA are shown above graph. Mean is presented as a gray bar.

Fig 2. Red blood cell (RBC) and platelet (PLT) counts during HEM and LBNP in anesthetized baboons.

Mean±SE, n = 13; * different from baseline (P<0.05); † different from blood removal (P<0.05); ‡ different between trials.

Fig 3. Thromboelastography parameters during HEM and LBNP in anesthetized baboons.

Reaction time until initial fibrin formation (R-time), rate of clot formation (α-Angle); maximal amplitude reflecting clot strength (MA) and clot lysis at 30 min after MA (LY30). Mean ±SE, n = 13; * different from baseline (P<0.05); and † different from HEM (P<0.05).

Fig 4. Plasma pro-coagulation factors VIII and V, fibrinogen, and von Willebrand factor antigen (vWF:Ag) during HEM and LBNP in anesthetized baboons.

Mean ±SE, n = 11–13; * different from baseline (P<0.05); and † different from HEM (P<0.05).

Fig 5. Plasma anti-coagulant protein C activity, protein S activity, and thrombin-antithrombin III complex (TAT) during HEM and LBNP in anesthetized baboons.

Mean ±SE, n = 11–13; * different from baseline (P<0.05); and † different from blood removal (P<0.05).

Fig 6. Fibrinolysis indicators tissue-type plasminogen activator (tPA), D-dimer, plasmin-α₂-antiplasmin complex (PAP), plasminogen activator inhibitor (PAI-1), thrombin activatable fibrinolysis inhibitor (TAFI) protein and activity during HEM and LBNP in anesthetized baboons.

Mean ±SE, n = 12–13; * different from baseline (P<0.05); and † different from blood removal (P<0.05).

Fig 7. Post hoc analyses of fibrinolysis data.

A. Swarm data plot shows TEG LY30 data for all subjects for HEM (dark circles) or LBNP (open circles) at each time point. B. Swarm data plot for TEG LY30 separates HEM and LBNP data sets into subjects whose LY30_BL <2< LY30_BL. Low-lysis designated with circles; high-lysis designated with squares; HEM are closed symbols; LBNP are open symbols. C. Swarm data plot for TEG LY30 separated into groups whose HEM and LBNP BL matched (either both low-lysis or both high-lysis BL matched; low-lysis (LY30_BL<2) subjects designated with grey circles; BL matched high-lysis (LY30_BL>2) subjects designated with grey squares.

Fig 8. Post Hoc analysis of fibrinolytic indicators grouped low-lysis (LY30_BL <2%) or high-lysis (LY30_BL >2%) baboons whose fibrinolytic state matched at both HEM and LBNP BL.

Swarm plots show all data points for fibrinolytic indicators LY30, D-dimer, tPA, PAI-1, TAFI-activity, TAFI-protein and PAP as well as the indicator of coagulation, TAT during HEM and LBNP treatment. Solid circles are BL low lysis; open circles are BL high lysis.

Fig 9. Post hoc analysis of correlations of LY30 to tPA; tPA to TAFI-activity; TAFI-protein to TAFI-activity; and tPA to PAI-1.

Graphs show all data points at individual time points BL and MAX hypovolemia for low-lysis and high-lysis groups. Solid circles are LY30_BL <2%; open circles are LY30_BL >2%.