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Heparin potentiation of coagulation cascade inhibitors. Triggering of the coagulation cascade occurs through tissue factor and damage surface exposure. The inhibitory action of heparin is through potentiation of a range of inhibitors with interactions linking to the kinin and complement pathways. Coagulation factors: FXII, factor XII; red, activated factor; blue, inhibitor.
Source publication
Heparin has been used extensively as an antithrombotic and anticoagulant for close to 100 years. This anticoagulant activity is attributed to the pentasaccharide sequence which potentiates the inhibitory action of antithrombin, a major inhibitor of the coagulation cascade. More recently it has been determined that heparin also has antithrombotic ac...
Contexts in source publication
Context 1
... Heparin also possesses an antithrombotic effect, which can be considered an interaction with the cellular components of the coagulation system. The process of coagulation can be split into two steps: (1) primary, which involves cellular components, and (2) secondary, which involves the soluble clotting factors ( Versteeg et al., 2013) depicted in Fig. 4. The in vivo process of thrombosis can be described broadly as follows: surface damage exposes the endothelium and/or subendothelium to blood; thrombogenic cell surfaces activate platelets leading to their adherence; platelets localize activation of the coagulation cascade (Hoffman and Monroe, 2001); and activation of the coagulation ...
Context 2
... initiation of coagulation is typically through surface damage, due to trauma or injury, to the endothelial cell layer of the vasculature. The underlining subendothelium and extracellular matrix is highly thrombogenic, with fibroblasts expressing tissue factor ( Mandal et al., 2006), which affects the coagulation cascade (see Fig. 4) and the matrix itself containing collagen, a potent activator of platelets ( Roberts et al., 2004). Exposure of the subendothelium will lead to platelet adherence, first via transient interactions of platelet expressed glycoprotein Ia/IIa to collagen, which releases von Willebrand factor from platelets ( Peyvandi et al., 2011). The ...
Context 3
... different endogenous coagulation inhibitors-AT, HCII, TFPI, C1-esterase inhibitor, and protein C inhibitor (PCI)-act as a balance ensuring localization of the clotting response. Of these inhibitors the major inhibitor is the serine proteases inhibitor (serpin) AT, which targets many of the activated coagulation factors (Fig. 4). As described earlier, the interaction of heparin and AT requires a specific pentasaccharide sequence, whereas the other serpins require no highly defined sequence (Huntington, 2011). The nonserpin inhibitor, TFPI, is also potentiated by heparin (Ellery and Adams, 2014), and the interactions of heparin with the different coagulation ...
Context 4
... through actions on the clotting factors but also due to an action on platelets, which form thrombi ( Periayah et al., 2017). The anticoagulant and antithrombotic activities of heparin are not mutually inclusive as hemostasis is far more complex than the clotting cascade and involves cellular blood elements ( Versteeg et al., 2013) as shown in Fig. 4. Heparin is known to interact with a number of cell surface binding proteins involved in hemostasis as described previously ( Mulloy et al., 2016). Furthermore, heparin fractionated to possess no anticoagulant activity can still reduce thrombus formation in a thrombogenic challenge model ( Gray et al., 1994). Similarly, in an ...
Citations
... It is worth noting that anticoagulant treatment, which is the cornerstone of pharmacotherapy of acute PE, may affect complement activity (32)(33)(34)(35)(36). Growing evidence suggests that anticoagulants, including unfractionated heparin (UFH), low-molecular-weight heparins (LMWHs) and direct factor Xa inhibitors (e.g., rivaroxaban) can modulate complement activity through a variety of mechanisms, primarily by affecting the cross-talk between the blood coagulation and complement cascades (34)(35)(36). ...
... It is worth noting that anticoagulant treatment, which is the cornerstone of pharmacotherapy of acute PE, may affect complement activity (32)(33)(34)(35)(36). Growing evidence suggests that anticoagulants, including unfractionated heparin (UFH), low-molecular-weight heparins (LMWHs) and direct factor Xa inhibitors (e.g., rivaroxaban) can modulate complement activity through a variety of mechanisms, primarily by affecting the cross-talk between the blood coagulation and complement cascades (34)(35)(36). However, in our study, all laboratory tests, including measurements of complement components, were performed on admission, before the initiation of anticoagulant treatment, which limited the impact of this confounder on the study results. ...
Background
Little is known about the role of complement activation in acute pulmonary embolism (PE). We investigated whether complement activation is associated with the severity of acute PE, along with the associated prothrombotic state, systemic inflammation and neutrophil extracellular traps (NETs) formation.
Methods
We studied 109 normotensive, non-cancer PE patients (aged 58.1±15.0 years). On admission prior to initiation of anticoagulation, plasma soluble complement components, i.e., C3a and sC5b-9, were measured with enzyme-linked immunosorbent assay (ELISA), along with thrombin generation, fibrinolysis proteins (plasminogen, antiplasmin, plasminogen activator inhibitor-1), factor VIII (FVIII) activity, and fibrin clot properties, including clot permeability (Ks, a measure of clot density) and clot lysis time (CLT). Moreover, we determined inflammatory markers and citrullinated histone H3, a specific marker of NETs formation.
Results
Patients in the lower tertile of C3a (≤1.45 ng/mL, n=37) had lower simplified Pulmonary Embolism Severity Index (sPESI) values and were less likely to have right ventricular (RV) dysfunction compared to the remaining subjects. The former subgroup also had 13% lower FVIII activity, but not fibrinogen, interleukin-6, fibrinolysis proteins, and thrombin generation. Plasma C3a levels correlated inversely with Ks and positively with CLT indicating formation of denser and poorly lysable clots in subjects with elevated C3a. Despite a positive association between C3a and sC5b-9, the latter parameter was solely associated with higher FVIII, but not with other variables.
Conclusions
We showed that in acute PE enhanced complement activation characterizes patients with poorer short-term prognosis who display prothrombotic fibrin clot properties and elevated FVIII, which supports the involvement of complement proteins in acute thromboembolism.
... Furthermore, heparin's rapid onset of action and short half-life make it an essential therapeutic agent for the prevention and treatment of acute thrombotic events, especially in high-risk clinical scenarios such as myocardial infarction, stroke, and post-surgical settings [8, 9,10]. ...
Cardiovascular diseases (CVDs) are the major cause of global morbidity and mortality, representing a substantial burden on global healthcare. Thrombotic events serve as major determinants of patient outcomes. Heparin, an anticoagulant drug, is crucial in treating thrombotic disorders. Its clinical utility is limited due to the challenges related to targeted delivery. In this study, we employed computational material modeling techniques using DFT calculations to investigate the potential of black phosphorus as drug carrier for heparin drug delivery. Thermodynamic favorability of interaction between drug and carrier is investigated by adsorption energies calculations. Molecular orbital analysis reveals the electron transfer process and potential reactivity of drug, carrier and their complexes, important for understanding the drug delivery mechanism. The properties such as chemical potential, chemical hardness, chemical softness and global electrophilicity index are computed to study the stability of drug delivery complexes. Weak intermolecular interactions between heparin and black phosphorus are highlighted by non-covalent interaction (NCI) analysis. UV-vis analysis was employed to generate UV-vis spectra of drug delivery complexes. These spectra reveal the potential photochemical properties of complexes, crucial for triggering drug delivery mechanism. This study presents the valuable insights into the therapeutic potential of black phosphorus as drug carrier for heparin drug delivery to treat cardiovascular diseases such as thrombotic events.
... Anticoagulant coating in circulation tubes is an essential anticoagulation technique, with Bioline heparin coating (Maquet company, Sweden) being the most frequently utilized method in ECMO [5][6][7]. Heparin molecules enhance the activity of antithrombin, which in turn inhibits coagulation factors II, IX, X, XI, and XII, resulting in an anticoagulant effect [8,9]. However, inserting heparin tubes may cause postoperative hemorrhage, thrombocytopenia, hypertriglyceridemia, hyperkalemia, and even heparin-induced thrombocytopenia (HIT) [7,10]. ...
Existing anticoagulant coatings are designed to inhibit the initial attachment of coagulant factors, but they frequently suffer from undesirable coating defects that result in a declined physical barrier function. Heparin, serving as a commercialized pharmacological coating via the activation of antithrombin, frequently causes heparin-induced thrombocytopenia. Here, we developed an argatroban-polyphenol conjugate proteinaceous coating (named PDA-BSA-AG coating) that actively inhibits thrombin, in contrast to previously reported passive coatings and indirect pharmacological anticoagulants. Following clinical treatment standards, a magnetic levitation ven-tricular assist system was employed for the first time to accomplish extracorporeal circulation utilizing PDA-BSA-AG coated tubes. The results show that the clotting time of the PDA-BSA-AG coating was 68.9 % longer than that of clinical Bioline heparin coating. It is noteworthy that argatroban not only delivers anticoagulant effectiveness but also enhances patient safety and reduces the occurrence of complications. A novel combination of the primary specific proteins of platelets and the coagulation-inflammatory cascade reaction of the cytokines and complement systems explained the anticoagulation mechanism. Reliable biocompatibility (hemolysis, cytotoxicity, and antibacterial activity) was noted, along with impressive coating stability for up to 6 h of extracorporeal circulation. These results above suggested that the PDA-BSA-AG coating is promising for a wide range of blood-contacting devices.
... Next, we tested if the combination of rIxsS17 and 17 kDa heparin had synergistic anti-plasma clotting effect. As heparin is an approved blood clotting disorder therapeutic [56][57][58], it is interesting to note that pre-incubating plasma with the rIxsS17 and heparin mixture significantly delayed plasma clotting up to 532.9 seconds compared to clotting time for buffer control (64.5 seconds), rIxsS17 only (184 seconds), and heparin only (407 seconds) (Fig 7). It is also notable that plasma clotting was also delayed to 474 seconds when a reaction was assembled from plasma that was incubated separately with rIxsS17 and heparin. ...
... The observed stoichiometry inhibition of rIxsS17 against trypsin, rat trypsin IV, and factor Xa were high and not close to the ideal 1:1 serpin-to-target protease ratio. These findings could be explained by evidence some of the serpins such as blood clotting regulatory serpins antithrombin III and heparin II that require binding of glycosaminoglycans (GAGs) to enhance their inhibitory potency [58]. Similarly, we have shown that heparin binding potentiated functions of AAS19, the IxsS17 homolog in A. americanum [74]. ...
Lyme disease (LD) caused by Borrelia burgdorferi is among the most important human vector borne diseases for which there is no effective prevention method. Identification of tick saliva transmission factors of the LD agent is needed before the highly advocated tick antigen-based vaccine could be developed. We previously reported the highly conserved Ixodes scapularis ( Ixs ) tick saliva serpin (S) 17 ( Ixs S17) was highly secreted by B . burgdorferi infected nymphs. Here, we show that Ixs S17 promote tick feeding and enhances B . burgdorferi colonization of the host. We show that Ixs S17 is not part of a redundant system, and its functional domain reactive center loop (RCL) is 100% conserved in all tick species. Yeast expressed recombinant (r) Ixs S17 inhibits effector proteases of inflammation, blood clotting, and complement innate immune systems. Interestingly, differential precipitation analysis revealed novel functional insights that Ixs S17 interacts with both effector proteases and regulatory protease inhibitors. For instance, r Ixs S17 interacted with blood clotting proteases, fXII, fX, fXII, plasmin, and plasma kallikrein alongside blood clotting regulatory serpins (antithrombin III and heparin cofactor II). Similarly, r Ixs S17 interacted with both complement system serine proteases, C1s, C2, and factor I and the regulatory serpin, plasma protease C1 inhibitor. Consistently, we validated that r Ixs S17 dose dependently blocked deposition of the complement membrane attack complex via the lectin complement pathway and protected complement sensitive B . burgdorferi from complement-mediated killing. Likewise, co-inoculating C3H/HeN mice with r Ixs S17 and B . burgdorferi significantly enhanced colonization of mouse heart and skin organs in a reverse dose dependent manner. Taken together, our data suggests an important role for Ixs S17 in tick feeding and B . burgdorferi colonization of the host.
... The variability of sulfation patterns in HS across different individuals, tissues, and cell types may contribute to virus tropism, although this variability hinders a full comprehension of their mechanistic role (Kearns et al., 2022). Heparin, a long, linear, highly-sulfated polyanionic oligomer, is frequently used as a model for HS in experimental studies due to their structural similarity (Hogwood et al., 2023). Computational and experimental studies have demonstrated that heparin can act as a potent antiviral agent by competing with HSPGs for binding to spike (Clausen et al., 2020;Kim et al., 2022;Kim et al., 2020;Liu et al., 2021;Mycroft-West et al., 2020;Paiardi et al., 2022;Sun et al., 2023;Tree et al., 2021) and, intriguingly, that spike forms a ternary complex by simultaneously binding ACE2 and heparin or HS (Cecon et al., 2022;Clausen et al., 2020;Kim et al., 2023). ...
The SARS-CoV-2 spike glycoprotein mediates virus attachment to human host cells by binding angiotensin-converting enzyme 2 (ACE2) and heparan sulfate (HS) proteoglycans. To elucidate the structure, dynamics, and functional consequences of these interactions, we carried out microsecond-long all-atom molecular dynamics simulations, followed by random acceleration molecular dynamics simulations, of the fully glycosylated spike:ACE2 complex with and without heparin chains bound. We find that heparin, a model for HS, promotes structural and energetic stabilization of the active conformation of the spike receptor binding domain (RBD) and reorientation of ACE2 toward the N-terminal domain in the same spike subunit as the RBD. Spike and ACE2 N-glycans exert synergistic effects, promoting better packing, strengthening the protein:protein interaction, and prolonging the residence time of the complex. ACE2 and heparin binding trigger rearrangement of the S2’ functional site through allosteric interdomain communication. HS thus has a multifaceted role in facilitating SARS-CoV-2 infection.
... In recent years, enoxaparin has been one of the most used low-molecular-weight heparins in the daily clinical management of several conditions. From a biochemical point of view, enoxaparin is a low-molecular-weight heparin obtained from porcine intestinal mucosa, and it is able to inhibit the action of several proteases involved in the clotting cascade [2]. It is an acidic mucopolysaccharide formed of equal parts sulfated D-glucosamine and D-glucuronic acid with sulfamic bridges. ...
Low-molecular-weight heparins are a class of drugs derived from the enzymatic depolymerization of unfractionated heparin that includes enoxaparin. Several studies have been performed on enoxaparin in recent years, in particular for the prevention and treatment of venous thromboembolism and for the treatment of acute coronary syndrome. Furthermore, the use of enoxaparin has been extended to other clinical situations that require antithrombotic pharmacological prevention, such as hemodialysis and recurrent abortion. In this review, we report the main clinical experiences of using enoxaparin in the prevention of VTE in nonsurgical patients.
... The pchepII gene was artificially synthesized by the company GENEWIZ, Inc., located in Suzhou, China. Then, it was combined with the vector pET-30a ( +) to generate the recombinant plasmid pET30a-pchepII, which carries a C-terminal (His) 6 -tag. The recombinant plasmid was transformed into competent E. coli BL21 (DE3) cells by heat shock at 42°C for 1 min. ...
... As shown in Fig. 3, the intracellular target protein PCHepII was released into the supernatant after the cells were subjected to ultrasonication and centrifugation. The soluble fractions were then purified using a Ni 2+ affinity column based on the (His) 6 -tag. The SDS-PAGE results indicated that the purification procedure was highly effective, resulting in a single band, and the purity of purified PCHepII was calculated to be 100%. ...
Heparin (HP) and heparan sulfate (HS) are multifunctional polysaccharides widely used in clinical therapy. Heparinases (Hepases) are enzymes that specifically catalyse HP and HS degradation, and they are valuable tools for studying the structure and function of these polysaccharides and for preparing low molecular weight heparins. In this study, by searching the NCBI database, a novel enzyme named PCHepII was discovered in the genome of the marine bacterium Puteibacter caeruleilacuae. Heterologously expressed PCHepII in Escherichia coli (BL21) has high expression levels and good solubility, active in sodium phosphate buffer (pH 7.0) at 20°C. PCHepII exhibits an enzyme activity of 254 mU/mg towards HP and shows weak degradation capacity for HS. More importantly, PCHepII prefers to catalyse the high-sulfated regions of HP and HS rather than the low-sulfated regions. Although PCHepII functions primarily as an endolytic Hepase, it mainly generates disaccharide products during the degradation of HP substrates over time. Investigations reveal that PCHepII exhibits a preference for catalysing the degradation of small substrates, especially HP tetrasaccharides. The catalytic sites of PCHepII include the residues His¹⁹⁹, Tyr²⁵⁴, and His⁴⁰³, which play crucial roles in the catalytic process. The study and characterization of PCHepII can potentially benefit research and applications involving HP/HS, making it a promising enzyme.
... Mammalian heparin is a sulfated GAG that has been widely used in the clinic as an effective anticoagulant and antithrombotic molecule. This molecule has also been studied as a promising alternative for cancer treatment and tumor progression [39,40]. Nevertheless, the occurrence of molecules such as heparin is not restricted to mammals [41], and the search for analogous compounds derived from marine invertebrates, such as ascidians, with similar biological activities but possibly devoid of unwanted side effects, offers a promising alternative in this area [42]. ...
Ascidians are marine invertebrates that synthesize sulfated glycosaminoglycans (GAGs) within their viscera. Ascidian GAGs are considered analogues of mammalian GAGs and possess great potential as bioactive compounds, presenting antitumoral and anticoagulant activity. Due to its worldwide occurrence and, therefore, being a suitable organism for large-scale mariculture in many marine environments, our main objectives are to study Microcosmus exasperatus GAGs regarding composition, structure, and biological activity. We also aim to develop efficient protocols for sulfated polysaccharides extraction and purification for large-scale production and clinical applications. GAGs derived from M. exasperatus viscera were extracted by proteolytic digestion, purified by ion-exchange liquid chromatography, and characterized by agarose gel electrophoresis and enzymatic treatments. Anticoagulant activity was evaluated by APTT assays. Antitumoral activity was assessed in an in vitro model of tumor cell culture using MTT, clonogenic, and wound healing assays, respectively. Our results show that M. exasperatus presents three distinct polysaccharides; among them, two were identified: a dermatan sulfate and a fucosylated dermatan sulfate. Antitumoral activity was confirmed for the total polysaccharides (TP). While short-term incubation does not affect tumor cell viability at low concentrations, long-term TP incubation decreases LLC tumor cell growth/proliferation at different concentrations. In addition, TP decreased tumor cell migration at different concentrations. In conclusion, we state that M. exasperatus presents great potential as an alternative GAG source, producing compounds with antitumoral properties at low concentrations that do not possess anticoagulant activity and do not enhance other aspects of malignancy, such as tumor cell migration. Our perspectives are to apply these molecules in future preclinical studies for cancer treatment as antitumoral agents to be combined with current treatments to potentiate therapeutic efficacy.
... These disaccharides are, in summary, carbohydrates with a repeating and specific sequence called glycosaminoglycans. Glycosaminoglycans are classified according to their composition into six types: heparin, heparan sulfate, chondroitin sulfate, dermatan sulfate, keratan sulfate, hyaluronic acid or hyaluronan 3,4 (Figure 1). The diversity of the glycosaminoglycan chains confers a variety of functions upon proteoglycans including collagen alignment, mineralization, cell signaling, mechanical property modulation, immune regulation, etc. 5 (Table S1). ...
Proteoglycans are core proteins associated with carbohydrate/sugar moieties that are highly variable in disaccharide composition, which dictates their function. These carbohydrates are named glycosaminoglycans, and they can be attached to proteoglycans or found free in tissues or on cell surfaces. Glycosaminoglycans such as hyaluronan, chondroitin sulfate, dermatan sulfate, keratan sulfate, and heparin/heparan sulfate have multiple functions including involvement in inflammation, immunity and connective tissue structure, and integrity. Heparan sulfate is a highly sulfated polysaccharide that is abundant in the periodontium including alveolar bone. Recent evidence supports the contention that heparan sulfate is an important player in modulating interactions between damage associated molecular patterns and inflammatory receptors expressed by various cell types. The structure of heparan sulfate is reported to dictate its function, thus, the utilization of a homogenous and structurally defined heparan sulfate polysaccharide for modulation of cell function offers therapeutic potential. Recently, a chemoenzymatic approach was developed to allow production of many structurally defined heparan sulfate carbohydrates. These oligosaccharides have been studied in various pathological inflammatory conditions to better understand their function and their potential application in promoting tissue homeostasis. We have observed that specific size and sulfation patterns can modulate inflammation and promote tissue maintenance including an anabolic effect in alveolar bone. Thus, new evidence provides a strong impetus to explore heparan sulfate as a potential novel therapeutic agent to treat periodontitis, support alveolar bone maintenance, and promote bone formation.
... Certain sulfate groups on this pentasaccharide are the key functional groups contributing to its pharmacological action. UFH is parenterally used (IV or SC), has a rapid onset of action, and its half-life is relatively short [49][50][51]. ...
Venous thromboembolism (VTE), comprising pulmonary embolism (PE) and deep vein thrombosis (DVT), poses a significant risk during and after hospitalization, particularly for surgical patients. Among various patient groups, those undergoing major orthopedic surgeries are considered to have a higher susceptibility to PE and DVT. Major lower-extremity orthopedic procedures carry a higher risk of symptomatic VTE compared to most other surgeries, with an estimated incidence of 4%. The greatest risk period occurs within the first 7-14 days following surgery. Major bleeding is also more prevalent in these surgeries compared to others, with rates estimated between 2% and 4%. For patients undergoing major lower-extremity orthopedic surgery who have a low bleeding risk, it is recommended to use pharmacological thromboprophylaxis with or without mechanical devices. The choice of the initial agent depends on the specific surgery and patient comorbidities. First-line options include low-molecular-weight heparins (LMWHs), direct oral anticoagulants, and aspirin. Second-line options consist of unfractionated heparin (UFH), fondaparinux, and warfarin. For most patients undergoing knee or hip arthroplasty, the initial agents recommended for the early perioperative period are LMWHs (enoxaparin or dalteparin) or direct oral anticoagulants (rivaroxaban or apixaban). In the case of hip fracture surgery, LMWH is recommended as the preferred agent for the entire duration of prophylaxis. However, emerging factor XI(a) inhibitors, as revealed by a recent meta-analysis, have shown a substantial decrease in the occurrence of VTE and bleeding events among patients undergoing major orthopedic surgery. This discovery poses a challenge to the existing paradigm of anticoagulant therapy in this specific patient population and indicates that factor XI(a) inhibitors hold great promise as a potential strategy to be taken into serious consideration.
