scholarly journals A Quantitative Systems Pharmacology (QSP) Model to Compare the Non-Clinical Biodistribution and Efficacy between Recombinant Factor IX (rIX) Therapies

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 4240-4240
Author(s):  
Sabine Pestel ◽  
Douglas Chung ◽  
Alireza Rezvani-Sharif ◽  
Ineke Muir ◽  
Sivarmurthy Krupa ◽  
...  
Keyword(s):  
Blood Loss ◽  
Significant Role ◽  
Bleeding Time ◽  
Total Blood Loss ◽  
Publicly Traded ◽  
Total Blood ◽  
The Past ◽  
Binding Partners ◽  
Pharmacologically Active

Abstract Background and Aims: Replacement FIX therapy (rIX) is an effective treatment for hemophilia B even with undetectable levels in the blood 1. However, the mechanistic reason for hemostasis with low plasma levels is not well understood. There is growing evidence that FIX interactions with one or multiple binding partners (BP), may play a significant role in the exposure and hemostatic efficacy of rIX 2,3. The aim of this study is to explore this hypothesis by comparing the plasma PK, tissue biodistribution, and in vivo endpoints of different rIX variants using a mouse QSP model. Method:  in vitro and in vivo FIX-KO mice studies and mathematical models were used to build a QSP model consisting of 8 tissue compartments , with each tissue divided into vascular, endothelial and interstitial spaces 4,5. The model simulates endogenous mouse IgG (mIgG), mouse serum albumin (MSA), and rIX dynamics including key clearance and distribution mechanisms. Competition for the endothelial FcRn receptor between Fc, albumin, mIgG, and MSA is explicitly modeled 6,7,8. The model was calibrated using mouse studies of radiolabeled rIX-Fc (Alprolix®), rIX-WT (BeneFIX®), and rIX-FP (Idelvion®). Tail-clip experiments following administration of rIX-Fc, rIX-WT, and rIX-FP were used to correlate the predicted exposures with the observed effects on bleeding time and total blood loss. Results: Preliminary simulations proved that having at least one BP best explains the rapid distribution of rIX-Fc and rIX-WT into the tissues, and the long plasma T 1/2 of rIX-Fc and rIX-FP. Visual predictive checks of the full PBPK model showed good agreement with the PK in the tissues. The best fit was achieved using a specific arrangement of four distinct binding partners: Shared BP (SBP) between all compounds (e.g. N-terminal binder) located within the vasculature with estimated K D of 470/600/4100 nM, for rIX-WT/rIX-FP/rIX-Fc, respectively. BP binding specific to rIX-WT (e.g. C-terminal binder) located in the interstitium of the tissue (varying densities) with estimated K D of 23 nM BP binding only for rIX-FP (e.g. albumin binder) located in both; the vasculature and interstitium of the tissue with estimated K D 20/0.05 μM (vascular/interstitial) BP binding only for rIX-Fc (e.g. Fc binder) located in the interstitium of tissue (varying densities) with estimated K D 3 μM The high degree of extravasation of rIX-Fc (and rIX-WT to a lesser degree) results in rapid distribution and sequestration in the tissues. The limited extravasation of rIX-FP and its high affinity to the SBP, results in increased recovery and a greater pool of bound rIX available in the tissue vasculature. Additionally, strong inverse correlation between the bound rIX in the vasculature and bleeding time/total blood loss suggests that the vascular pool plays a more significant role in FIX pharmacology, as compared to the pool in the extravascular space. Conclusion: The mouse QSP model demonstrated that the plasma and tissue biodistribution of rIX-Fc, rIX-FP, and rIX-WT cannot be explained without a BP, and that it is plausible to assume that different binding partners, both intra- and extravascular, for different rFIX variants exist. The correlation between the levels of bound rIX and the coagulation endpoints suggests that the vascular bound rIX may be the pharmacologically active pool or reservoir for haemostasis. The extravasation and sequestration of rIX-WT and rIX-Fc into the tissues may explain the decreased vascular exposure, and hence, the reduced efficacy (increased bleeding time/total blood loss) at later time points. Although the exact identity of the BP's remains to be further elucidated, the model estimates of their affinity, density and location provide guidance for further experimental investigations. Expansion of the QSP model with additional data and coagulation kinetics will further our understanding of the role of BPs in rIX pharmacology. References 1Srivastava A et al (2013) Haemophilia 19(1), e1-47 2Feng D et al (2013) JTH, Vol. 11 (12), 2176-2178 3Cheung WF et al (1996) PNAS USA, 93(20), 11068-11073 4Li L et al (2014) AAPS Journal 16(5), 1097-1109 5Shah DK & Betts AM (2012) J Pharmacokinet Pharmacodyn 39(1), 67-86 6Chia J et al (2018) J Biol Chem 293(17), 6363-6373 7Andersen JT et al (2010) J Biol Chem 285(7), 4826-4836 8Andersen JT et al (2013) J Biol Chem 288(33), 24277-24285 Disclosures Pestel: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Rezvani-Sharif: CSL Behring Ltd: Current Employment, Current equity holder in publicly-traded company. Muir: CSL Behring Ltd: Current Employment, Current holder of stock options in a privately-held company. Krupa: CSL Behring LLC: Current Employment, Current equity holder in publicly-traded company. Brechmann: CSL Behring Innovation GmbH, Ended employment in the past 24 months: Bayer Ag (Bayer Pharmaceuticals),: Current Employment, Ended employment in the past 24 months, Patents & Royalties: Bayer. Verhagen: CSL Behring Ltd: Current Employment, Current equity holder in publicly-traded company. Dower: CSL Behring Ltd: Current Employment, Current equity holder in publicly-traded company. Herzog: CSL Behring GmbH: Current Employment, Current equity holder in publicly-traded company.

Detection of Carriers of Hemophilia by Hemorrhagometry

1975 ◽  
Author(s):  
J. Jesdinsky-Buscher ◽  
A. H. Sutor ◽  
H. Niederhoff
Keyword(s):  
Blood Loss ◽  
Cold Tolerance ◽  
Room Temperature ◽  
Bleeding Time ◽  
Hemophilia A ◽  
Tolerance Test ◽  
Normal Range ◽  
Total Blood ◽  
Abnormal Bleeding

Hemorrhagometry, a new in-vivo method, measures bleeding time, intensity and total blood loss from a small standardized wound (Sutor et al.: Blut 22, 27, 1971). In patients with hemophilia these values are within the normal range when the test is performed at room temperature (24° C). However, when the wound is cooled to 17° C (cold tolerance test), bleeding time is abnormally long in hemophiliacs. Therefore, we investigated this test in carriers of hemophilia. The cold tolerance test was performed in 16 ‘proven’ and 6 ‘probable’ carriers (criteria according to Nilsson). 14 proven and 4 probable carriers showed abnormal bleeding times of 15 min and more (2 proven carriers of hemophilia B did not differ markedly from 20 carriers of hemophilia A). When hemorrhagometry was performed at room temperature, the carriers could not be distinguished from normal control persons.Of 15 sisters and aunts of hemophiliacs 9 had abnormal cold tolerance test findings, in fair agreement with the probability of .5 to be expected theoretically. Thus the hemorrhagometry cold tolerance test is helpful in detecting carriers of hemophilia.

scholarly journals Pharmacokinetics and Pharmacodynamics of Recombinant FIX Variants in the FIX Knockout Mouse Tail Clip Bleeding Model

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3180-3180
Author(s):  
Sabine Pestel ◽  
Hendrik Peil ◽  
Steffi Knoll Machado ◽  
Philipp Claar ◽  
Elmar Raquet ◽  
...  
Keyword(s):  
Blood Loss ◽  
Plasma Levels ◽  
Half Life ◽  
Bleeding Time ◽  
Isotonic Saline ◽  
Publicly Traded ◽  
Total Blood ◽  
Time Points ◽  
Hemostatic Activity ◽  
Tail Clip

Abstract Introduction: The recessive X-linked bleeding disorder Hemophilia B is caused by a mutation in the coagulation factor (F) IX gene leading to partial or total loss of its function. Preventive treatment with replacement long-acting FIX is an attractive option for patients to reduce administration frequency and prevent bleeding. New recombinant FIX therapeutics like the albumin-fused FIX (rFIX-FP) or the Fc-fused FIX (rFIX-Fc) enable longer half-life in circulation and thus less frequent administration, as compared to non-fused FIX (rFIX). Studies in FIX knockout (KO) mice were conducted to characterize the effect of the modifications on the pharmacokinetic (PK) and pharmacodynamic (PD) properties of the different recombinant FIX products. Methods: Pharmacokinetics: Recombinant FIXs were administered intravenously at doses of 25 nmol/kg (corresponding to ~175-400 IU/kg FIX clotting activity) to FIX KO mice. Blood samples were collected starting at 5 min, and up to 336 h. FIX plasma levels were measured with an ELISA-based assay with anti-human FIX paired antibodies. PK was evaluated by non-compartmental analysis. Biodistribution: 3H-labeled recombinant FIXs were administered intravenously at doses of 200 IU/kg to FIX KO mice. Plasma levels and organ distribution were quantified starting at 15 min, and up to 240 h. Pharmacodynamics: FIX KO mice were treated intravenously with 50 IU/kg FIX clotting activity (nominal or labeled potency) of different rFIX products at 24, 72, 120 168 and 336 h prior to determination of bleeding time and total blood loss in a tail clip bleeding model. Immediately upon lesion, the tail tip was submerged in isotonic saline (0.9 %), kept at the mice physiological body temperature. Time to hemostasis is quantified as the time until bleeding stops for a minimum of 2 min. The volume of total blood loss was calculated by measuring the hemoglobin present in the isotonic saline solution at the end of the experiment. Results: Distinct PK profiles were observed for the three FIX molecules, where rFIX and rFIX-Fc exhibit an initial rapid distribution phase from plasma, while rFIX-FP showed a monophasic elimination profile up to 120 h post administration (p.a.). In the terminal phase, rFIX levels were quantifiable for up to 48 h p.a., while both; rFIX-FP and rFIX-Fc were measurable in plasma up to 240 h p.a. In line with this, the overall exposure AUC 0-inf is ranked in the following order: rFIX-FP > rFIX-Fc > rFIX. In the biodistribution study, a similar plasma PK profile was determined. Given the sensitivity of the radioactive method, an exposure plateau was observed for rFIX-Fc, and at lower levels for rFIX, whereas rFIX-FP continued to exhibit monophasic plasma clearance behavior. rFIX-FP exposure in the extravascular space (EVS) was lower than for the other FIX products. This is in line with volumes of distribution (Vss and Vz) which were highest for rFIX-Fc (AUC ranking rFIX-Fc > rFIX > rFIX-FP). FIX hemostatic efficacy in tail clip model was comparable for all 3 FIXs at the early time points but diverged at later time points post dosing. The blood loss and bleeding time measurements returned to baseline within 168 h for rFIX and rFIX-Fc, while the rFIX-FP group maintained robust hemostatic activity for up to 336 h. In contrast to lowest tissue exposure of rFIX-FP, the plasma AUC for rFIX-FP was highest, compared to FIX-Fc or FIX. In line, efficacy over time was also highest for rFIX-FP, suggesting that tissue exposure might not be the main driver for hemostatic activity. Conclusion: Different FIX products exhibit divergent PK and PD behaviors. rFIX-FP plasma PK profile suggests somewhat lower tissue distribution in comparison to rFIX-Fc and rFIX, which was confirmed in the tissue biodistribution study. Despite its limited extravasation into tissue, rFIX-FP exhibits superior and prolonged hemostatic activity in the FIX KO mouse tail clip model. rFIX and rFIX-Fc show comparable tissue biodistribution behavior, with robust extravasation into the EVS. Despite having the longest half-life and overall (plasma and tissue) exposure in the mouse, rFIX-Fc lost hemostatic activity in the tail clip model significantly faster than rFIX-FP. As a result, hemostatic efficacy was highest for the FIX-FP, the product with the lowest distribution volumes. The results therefore suggest that EVS is not the main determining factor for FIX efficacy in vivo. Disclosures Pestel: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Peil: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Knoll Machado: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Claar: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Raquet: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Ponnuswamy: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Mischnik: CSL Behring Innovation GmbH: Current Employment, Current equity holder in publicly-traded company. Herzog: CSL Behring GmbH: Current Employment, Current equity holder in publicly-traded company.

scholarly journals Less blood loss in supercapsular percutaneously assisted versus posterolateral total hip arthroplasty

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Yong Hu ◽  
Min-Cong Wang ◽  
Tao Wang ◽  
Yue Meng ◽  
Xiao-Min Chao ◽  
...  
Keyword(s):  
Total Hip Arthroplasty ◽  
Blood Loss ◽  
Hip Arthroplasty ◽  
Total Blood Loss ◽  
Transfusion Rate ◽  
Operative Blood Loss ◽  
Total Blood ◽  
Radiographic Findings ◽  
Total Hip ◽  
Affected Side

Abstract Background Although excellent clinical outcomes of supercapsular percutaneously assisted total hip arthroplasty (SuperPath) have been reported, the peri-operative blood loss has rarely been reported. The current study determined the blood loss during SuperPath and compared the blood loss with conventional posterolateral total hip arthroplasty (PLTH). Methods This retrospective study enrolled patients who underwent unilateral primary THA between January 2017 and December 2019. The demographic data, diagnoses, affected side, radiographic findings, hemoglobin concentration, hematocrit, operative time, transfusion requirements, and intra-operative blood loss were recorded. The peri-operative blood loss was calculated using the OSTHEO formula. Blood loss on the 1st, 3rd, and 5th post-operative days was calculated. Hidden blood loss (HBL) was determined by subtracting the intra-operative blood loss from the total blood loss. Results Two hundred sixty-three patients were included in the study, 85 of whom were in the SuperPath group and 178 in the posterolateral total hip arthroplasty (PLTH) group. Patient demographics, diagnoses, affected side, operative times, and pre-operative hemoglobin concentrations did not differ significantly between the two groups (all P > 0.05). Compared to the PLTH group, the SuperPath group had less blood loss, including intra-operative blood loss, 1st, 3rd, and 5th post-operative days blood loss, and HBL (all P < 0.05). Total blood loss and HBL was 790.07 ± 233.37 and 560.67 ± 195.54 mL for the SuperPath group, respectively, and 1141.26 ± 482.52 and 783.45 ± 379.24 mL for the PLTH group. PLTH led to a greater reduction in the post-operative hematocrit than SuperPath (P < 0.001). A much lower transfusion rate (P = 0.028) and transfusion volume (P = 0.019) was also noted in the SuperPath group. Conclusion SuperPath resulted in less perioperative blood loss and a lower transfusion rate than conventional PLTH.

scholarly journals 3D-printed models for periacetabular osteotomy surgical planning

2021 ◽  
Author(s):  
B Keegan Markhardt ◽  
Matthew A Beilfuss ◽  
Scott J Hetzel ◽  
David C Goodspeed ◽  
Andrea M Spiker
Keyword(s):  
Radiation Dose ◽  
Blood Loss ◽  
Surgical Planning ◽  
Preoperative Planning ◽  
Periacetabular Osteotomy ◽  
Total Blood Loss ◽  
Preoperative Imaging ◽  
Total Blood ◽  
Patient Age ◽  
3D Printed

Abstract The purpose of this study was to determine the feasibility and clinical benefits of using 3D-printed hemipelvis models for periacetabular osteotomy preoperative planning in the treatment of hip dysplasia. This retrospective study included 28 consecutive cases in 26 patients, with two bilateral cases, who underwent periacetabular osteotomy between January 2017 and February 2020 and had routine radiographs, CT and MR imaging. Of these, 14 cases [mean patient age 30.7 (SD 8.4) years, 11 female] had routine preoperative imaging, and 14 cases [mean patient age 28.0 (SD 8.7) years, 13 female] had routine preoperative imaging and creation of a full-scale 3D-printed hemipelvis model from the CT data. The expected surgical cuts were performed on the 3D-printed models. All patients underwent Bernese periacetabular osteotomy. Operative times, including time to achieve proper acetabular position and total periacetabular osteotomy time, fluoroscopy radiation dose and estimated total blood loss were compiled. ANOVA compared outcome variables between the two patient groups, controlling for possible confounders. On average, patients who had additional preoperative planning using the 3D-printed model had a 5.5-min reduction in time to achieve proper acetabular position and a 14.5-min reduction in total periacetabular osteotomy time; however, these changes were not statistically significant (P = 0.526 and 0.151, respectively). No significant difference was identified in fluoroscopy radiation dose or total blood loss. Detailed surgical planning for periacetabular osteotomy using 3D-printed models is feasible using widely available and affordable technology and shows promise to improve surgical efficiency.

Evaluating the Role of Anesthesia on Intraoperative Blood Loss and Visibility during Endoscopic Sinus Surgery: A Meta-analysis

2021 ◽  
pp. 194589242198915
Author(s):  
David C. Moffatt ◽  
Robert A. McQuitty ◽  
Alex E. Wright ◽  
Tawanda S. Kamucheka ◽  
Ali L. Haider ◽  
...  
Keyword(s):  
Heart Rate ◽  
Blood Loss ◽  
Endoscopic Sinus Surgery ◽  
Average Rate ◽  
Meta Analysis ◽  
Sinus Surgery ◽  
Total Blood Loss ◽  
Total Blood ◽  
Intraoperative Bleeding ◽  
Significant Difference

Background Previous studies and meta analyses have led to incongruent and incomplete results respectively when total intravenous anesthesia (TIVA) and inhalational anesthesia (IA) are compared in endoscopic sinus surgeries in regards to intraoperative bleeding and visibility. Objective To perform a more comprehensive meta-analysis on randomized controlled trial (RCTs) comparing TIVA with IA in endoscopic sinus surgery to evaluate their effects on intraoperative bleeding and visibility. Methods A systematic review and meta-analysis of studies comparing TIVA and IA in endoscopic sinus surgery for chronic rhinosinusitis was completed in May 2020. Utilizing databases, articles were systematically screened for analysis and 19 studies met our inclusion criteria. The primary outcome included intraoperative visibility scores combining Boezaart, Wormald and Visual Analogue Scale (VAS). Secondary outcomes included rate of blood loss (mL/kg/min), estimated total blood loss (mL), Boezaart, Wormald scores, VAS, heart rate, and mean arterial pressure (MAP). Results 19 RCTs with 1,010 patients were analyzed. TIVA had a significantly lower intraoperative bleeding score indicating better endoscopic visibility (Boezaart, VAS, and Wormald) than IA (−0.514, p = 0.020). IA had a significantly higher average rate of blood loss than TIVA by 0.563 mL/kg/min (p = 0.016). Estimated total blood loss was significantly lower in TIVA than IA (−0.853 mL, p = 0.002). There were no significant differences between TIVA and IA in the mean heart rate (−0.225, p = 0.63) and MAP values (−0.126, p = 0.634). The subgroup analyses revealed no significant difference between TIVA and IA when remifentanil was not utilized and whenever desflurane was the IA agent. Conclusion TIVA seemed to have superior intraoperative visibility scores and blood loss during endoscopic sinus surgery when compared to IA. However, the results are not consistent when stratifying the results based on the use of remifentanil and different inhaled anesthetics. Therefore, the conclusion cannot be made that one approach is superior to the other.

Improved hemostasis with superactive analogs of factor VIIa in a mouse model of hemophilia A

Blood ◽  
2003 ◽  
Vol 102 (10) ◽  
pp. 3615-3620 ◽  
Author(s):  
Mikael Tranholm ◽  
Kim Kristensen ◽  
Annemarie T. Kristensen ◽  
Charles Pyke ◽  
Rasmus Røjkjær ◽  
...  
Keyword(s):  
Blood Loss ◽  
Bleeding Time ◽  
Hemophilia A ◽  
Recombinant Factor Viia ◽  
Factor Viia ◽  
Intrinsic Activity ◽  
Hemostatic Effect ◽  
Therapeutic Doses ◽  
Hemostatic Effects

AbstractIt is currently debated whether the mechanism of action of therapeutic doses of recombinant factor VIIa (rFVIIa, Novo-Seven) relies on the tissue factor (TF)-independent activity of the enzyme. The present study was conducted to investigate the in vivo hemostatic effects of rFVIIa and 3 analogs thereof with superior intrinsic activity (FVIIaIIa, K337A-FVIIaIia, and M298Q-FVIIa) in mice with antibody-induced hemophilia A. A highly significant dose response was observed for the bleeding time and blood loss for each of the rFVIIa variants. The bleeding time and blood loss were normalized after administration of 10 mg/kg rFVIIa, 3 mg/kg K337A-FVIIaIia, and 3 mg/kg M298Q-FVIIa, indicating a potency of these FVIIa analogs 3-4 times above that of rFVIIa in FVIII-depleted mice. The different in vivo potencies of the various forms of FVIIa could not be explained by the pharmacokinetics. Histopathological evaluation of kidneys revealed no signs of treatment-related pathological changes even after treatment with the superactive variants. The fact that FVIIa analogs with enhanced intrinsic activity are more efficacious in the murine hemophilia A model strongly suggests that the TF-independent procoagulant activity of FVIIa contributes to its clinical hemostatic effect. (Blood. 2003; 102:3615-3620)

scholarly journals Uterine Balloon Tamponade in Combination with Topical Administration of Tranexamic Acid for Management of Postpartum Hemorrhage

2015 ◽  
Vol 2015 ◽  
pp. 1-4 ◽  
Author(s):  
Masato Kinugasa ◽  
Hanako Tamai ◽  
Mayu Miyake ◽  
Takashi Shimizu
Keyword(s):  
Blood Loss ◽  
Tranexamic Acid ◽  
Postpartum Hemorrhage ◽  
Balloon Catheter ◽  
Stable Condition ◽  
Total Blood Loss ◽  
Total Blood ◽  
Balloon Tamponade ◽  
First Case ◽  
Uterine Balloon Tamponade

While uterine balloon tamponade is an effective modality for control of postpartum hemorrhage, the reported success rates have ranged from the level of 60% to the level of 80%. In unsuccessful cases, more invasive interventions are needed, including hysterectomy as a last resort. We developed a modified tamponade method and applied it to two cases of refractory postpartum hemorrhage after vaginal delivery. The first case was accompanied by uterine myoma and low-lying placenta. After an induced delivery, the patient had excessive hemorrhage due to uterine atony. Despite oxytocin infusion and bimanual uterine compression, the total blood loss was estimated at 2,800 mL or more. The second case was diagnosed as placental abruption complicated by fetal death and severe disseminated intravascular coagulation, subsequently. A profuse hemorrhage continued despite administration of uterotonics, fluid, and blood transfusion. The total blood loss was more than 5,000 mL. In each case, an intrauterine balloon catheter was wrapped in gauze impregnated with tranexamic acid, inserted into the uterus, and inflated sufficiently with sterile water. In this way, mechanical compression by a balloon and a topical antifibrinolytic agent were combined together. This method brought complete hemostasis and no further treatments were needed. Both the women left hospital in stable condition.

scholarly journals 3-h drain clamping is not effective to reduce total blood loss after primary total knowledge

2020 ◽  
Vol 32 (1) ◽  
Author(s):  
Dojoon Park ◽  
Youn Ho Choi ◽  
Kwang Hyun Cho ◽  
Hae Seok Koh
Keyword(s):  
Blood Loss ◽  
Total Blood Loss ◽  
Total Blood ◽  
Drain Clamping

o-Raffinose cross-linked hemoglobin improves the hemostatic defect associated with anemia and thrombocytopenia in rabbits

Blood ◽  
2000 ◽  
Vol 96 (10) ◽  
pp. 3630-3636 ◽  
Author(s):  
David H. Lee ◽  
Leslie Bardossy ◽  
Nichole Peterson ◽  
Morris A. Blajchman
Keyword(s):  
Blood Loss ◽  
Bleeding Time ◽  
Platelet Reactivity ◽  
Inhibitory Effect ◽  
Clinical Settings ◽  
No Donor ◽  
Hemostatic Effect ◽  
Circulating Levels

Abstract Several different preparations of cross-linked hemoglobin (CLHb) are being evaluated for their efficacy and safety as red cell substitutes in a variety of preclinical and clinical settings. Because CLHb is known to sequester nitric oxide (NO) and inhibit NO-mediated processes, we hypothesized that CLHb would have a hemostatic effect by enhancing platelet reactivity, inducing vasoconstriction, or both. Infusion of o-raffinose CLHb shortened the prolonged microvascular bleeding time and decreased blood loss from ear incisions in rabbits rendered anemic and thrombocytopenic. Moreover, this hemostatic effect persisted for at least 24 hours after infusion. Phenylephrine induced a degree of vasoconstriction similar to that induced by CLHb but did not shorten the bleeding time or decrease blood loss, suggesting that vasoconstriction alone cannot account for the hemostatic effect of CLHb. There was no evidence of CLHb-induced activation of coagulation in vivo, since infusion of CLHb did not increase circulating levels of thrombin-antithrombin complex. In vitro, CLHb abolished the inhibitory effect of the NO donor 3-morpholinosydnonimine on platelet aggregation and enhanced the aggregation of stimulated but not resting platelets. This potentiating effect was not attenuated by the addition of superoxide dismutase or catalase. To evaluate the potential arterial thrombogenicity of CLHb, a model of carotid artery thrombosis was developed in rabbits without thrombocytopenia or anemia. Compared with albumin infusion, CLHb infusion shortened the time to complete carotid occlusion. These data suggest that CLHb may shift the thromboregulatory balance toward clot formation, resulting in decreased bleeding in anemic and thrombocytopenic rabbits and possibly increasing arterial thrombogenicity in normal rabbits.

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