Hemostatic Activity Research Articles

Factor VIII Mimetic Rescue of Hemophilia B Causing Factor IX Variants

Background The intrinsic Xase enzyme complex is responsible for the rate-limiting step of sustained coagulation. It is composed of the protease, activated coagulation factor IX (FIXa), and the cofactor, FVIIIa, bound on phospholipid membranes; deficiencies in the activity of the intrinsic Xase result in hemophilia B (HB) or hemophilia A (HB), respectively. The FVIII-mimetic emicizumab has been widely adopted for prophylaxis for HA patients, in part because of the convenience of subcutaneous administration. Prophylaxis for HB patients, however, still requires frequent intravenous FIX administration. Within the Xase, complex, FVIIIa binding allosterically activates FIXa and enhances FX substrate binding, though the molecular details of the FVIIIa/FIXa interaction remain incompletely understood. We hypothesize that the hemostatic activity of select HB-causing mutations due to dysfunctional FVIIIa/FIXa interactions may be rescuable by FVIII-mimetics such as emicizumab. Aims To determine the spectrum of HB-causing missense FIX variants that have rescuable procoagulant activity with FVIII-mimetics and clarify the interaction between FIXa and FVIIIa at an amino acid level. Methods Recombinant FIX variants were expressed from transiently transfected HEK293 cells. Procoagulant activity of recombinant FIX variant and patient samples were assayed by either aPTT assay, one-stage FIX activity, thrombin generation assay (TGA), and rotation thromboelastography (ROTEM). Results The ability of the FVIII-mimetic emicizumab to rescue the procoagulant activity of HB-causing FIX variants was screened using a modified one-stage FIX activity assay. To account for the high clotting activity of emicizumab in aPTT-based assays, the FIX activity of the HB-causing variants was compared against the activity of wild-type (WT) FIX in the presence of an equal concentration of emicizumab. FVIII-mimetic rescuable HB-causing variants were selected if their activity relative-to-FIX-WT increased at least twofold with the addition of emicizumab. About 20% of FIX variants evaluated to-date appear to be rescuable with emicizumab, including K5E, L6S, D47E, D332Y, R333Q, R338P, P368T, E387G, E387K, and I397T. To further define the procoagulant rescue of these FIX variants, the amount of thrombin generated by these FIX variants in HB plasma was measured with and without emicizumab and compared to FIX-WT with the same emicizumab concentrations. Representative thrombograms are shown in the Figure. The addition of emicizumab increased the peak thrombin generation of the rescuable HB-causing variants 2-6 fold relative-to-FIX-WT. In contrast, emicizumab had no effect on the amount of thrombin generated by the catalytically inert FIX-S365A (S195A in chymotrypsin numbering). We also obtained samples from two moderate HB patients with the I397T missense variant; these patients have between 1-3% normal FIX activity and a moderate bleeding phenotype. We observed that emicizumab could normalized the prolonged aPTT of their plasma and improved the ROTEM clot time of their whole blood in a dose-dependent manner. Conclusions The procoagulant activity of select HB-causing missense FIX variants can be rescued with therapeutics amounts of emicizumab. These variants likely have a dysfunctional interaction with FVIIIa cofactor, which is circumvented in the presence of a FVIII-mimetic. Importantly, such rescuable variants are identified throughout the FIXa protein, highlighting the range of amino acids involved in the FIXa/FVIIIa interaction. Identified rescuable variants likely has dysfunctional FVIIIa-binding (K5E, L6S) and/or disturbed allosteric activation (E387K, I397T). FVIII mimetics may be an alternative prophylactic hemostatic therapy for HB patients with such rescuable variants. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Quantitative Estimation of the In Vivo Equivalent Factor VIII Activity of NXT007, an Emicizumab-Based, Next-Generation, Activated Factor VIII-Mimetic Bispecific Antibody, from Nonclinical Data

INTRODUCTION: NXT007 is a bispecific antibody (BsAb) that bridges activated factor IX (FIXa) and factor X (FX), thereby mimicking the cofactor function of activated factor VIII, and was discovered by engineering the first-in-class drug of emicizumab. Nonclinical investigations suggested the potential of NXT007 to provide a supra-emicizumab and even non-hemophilic level of coagulation activity to patients with hemophilia A [1,2]. Meanwhile, quantitative characterization and comparison of the in vivo hemostatic activity of NXT007 with factor VIII (FVIII) and emicizumab remained to be addressed. This research aimed to quantitatively estimate the in vivo hemostatic activity of NXT007 in terms of relative potency to FVIII ("equivalent FVIII activity") by leveraging and analyzing previously published nonclinical data with model-based approaches. METHODS: Simulation of factor IX (FIX)-BsAb-FX ternary complex formation in human plasma using a binding kinetic model was employed for predicting the in vivo equivalent FVIII activity of NXT007, as previously investigated to associate the ternary complex concentration with in vivo equivalent FVIII activity in a non-human primate model of acquired hemophilia A for emicizumab [3]. Dissociation constants against human FIX and FX, which were determined by a surface plasmon resonance analysis in buffer solution, were used for simulating the equilibrium binding of the BsAbs with these antigens as functions of their plasma concentrations. The conversion factor from ternary complex concentration to in vivo equivalent FVIII activity for NXT007 was assumed to be equivalent to that for emicizumab, given their comparable effects for increasing the turnover rate of FIXa-catalyzed FX activation on top of their similar affinities to FIX/FIXa. To support the validity of this assumption, in vitro equivalent FVIII activities of NXT007 and emicizumab were estimated and compared. Peak height data from in vitro activated factor XI (FXIa)- or tissue factor (TF)-triggered thrombin generation (TG) assays, in which FVIII-deficient human plasma was spiked with vehicle, FVIII (10.0 to 150 IU/dL), NXT007 (0.0300 to 3000 μg/mL), or emicizumab (0.0300 to 3000 μg/mL), were used for developing pharmacodynamic models to describe the concentration-response relationships of the three active drugs. The developed models were then transformed to derive the corresponding in vitro equivalent FVIII activities of NXT007 and emicizumab as functions of their plasma concentrations. RESULTS: The simulated relationships between plasma concentration and equivalent FVIII activity by the ternary complex, FXIa-triggered TG, and TF-triggered TG approaches for NXT007 and emicizumab were all bell-shaped, peaking at 109, 63.7, and 127 IU/dL with 53.1, 130, and 82.8 μg/mL, respectively, for NXT007 and at 28.2, 19.0, and 48.2 IU/dL with 265, 452, and 476 μg/mL, respectively, for emicizumab (Figure). With higher peak equivalent FVIII activities achieved with lower plasma concentrations for NXT007 than emicizumab, the interrelationship among the simulated relationships between plasma concentration and equivalent FVIII activity by the three approaches appeared consistent between NXT007 and emicizumab, as characterized by the same order of peak equivalent FVIII activity, i.e., TF-triggered TG > ternary complex > FXIa-triggered TG. This consistency may suggest comparable coagulation activities per ternary complex between NXT007 and emicizumab, supporting the validity of assuming an equivalent conversion factor from ternary complex concentration to in vivo equivalent FVIII activity for NXT007 and emicizumab. By the ternary complex approach, the minimum plasma concentrations of NXT007 to achieve in vivo equivalent FVIII activities of 15 (anticipated therapeutic level with emicizumab), 40 (threshold for hemophilia A), and 100 (typical normal) IU/dL were predicted to be 2.50, 7.26, and 30.2 μg/mL, respectively (Figure). CONCLUSIONS: Model-based approaches supported application of the ternary complex approach for predicting the in vivo equivalent FVIII activity of NXT007. Findings from this research provided a basis for designing a phase I/II clinical study of NXT007 that is currently ongoing (NXTAGE; JapicCTI-194919). REFERENCES: 1. Res Pract Thromb Haemost 2020;4 Suppl 1:579. 2. Blood 2020;136 Suppl 1:19. 3. Thromb Haemost 2017;117:1348-57. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Isolation and identification of hemostatic steroidal glycosides from Ypsilandra thibetica

The phytoconstituents of the fraction with hemostatic activity of the 70% aqueous ethanol extract of Ypsilandra thibetica Franch. were investigated. As a result, fourteen previously unreported spirostanol saponins, ypsilandrosides Z1-Z14, and nine known analogues were isolated and characterized by MS, NMR, and chemical methods. Among them, ypsilandrosides Z1-Z4 (1–4) have a rare 12-O-β-d-glucopyranosyl group, while ypsilandrosides Z5-Z8 (5–8) possess a rare double bond between C-4 and C-5, and a hydroxyl or carbonyl located at the C-6. All isolates were further tested for their hemostatic activity. The results suggested that five spirostanol tetraglycosides show favorable inducing platelet aggregation activities. Among them, ypsilandroside G (16) displayed significant inducing platelet aggregation activity with an EC50 value of 57.17 μM. Furthermore, the preliminary structure–activity relationship of these spirostanol glycosides’ hemostatic activity was discussed.

Study of specific properties of Polygonum Aviculare L. dry extract as a hemostatic agent

Polygonum aviculare L., has been used in traditional medicine for many years. Hemostatic activity of the Polygonum aviculare L. dry extract (PADE) on the duration and amount of bleeding was studied in the current research. Moreover, the effect of the PADE was investigated on platelet count and capillary permeability. The results showed that the PADE administration increased platelet count by 212%, decreased retraction time by 50% and increased adhesive capacity by 225% and significantly reduced capillary permeability in the experimental animals. According to the results of this experiment, the Polygonum aviculare L. dry extract can be recommended to use in medical practice as a hemostatic agent in various hemorrhages.

Assembly of Clay Nanotubes on Cotton Fibers Mediated by Biopolymer for Robust and High-Performance Hemostatic Dressing.

Uncontrollable bleeding from military conflicts, accidents, and surgical procedures is a major life-threatening factor. Rapid, safe, and convenient hemostasis is critical to the survival of bleeding patients in prehospital care. However, the peel-off of hemostats such as kaolinite sheets from the cotton fibers often poses a risk of distal thrombosis. Here, an efficient clay hemostat of halloysite nanotubes is tightly bound onto commercial cotton fibers, which is capillary mediated by biopolymer alginate with Ca2+ crosslinking. The robust clay nanotube dressing materials maintain high procoagulant activity after harsh water treatment, and only a few residuals of halloysite exist in the wound area. Compared with commercial hemostat QuikClot Combat gauze, halloysite-alginate-cotton composite dressing exhibits hemostatic properties both in vivo and in vitro with high safety. The hemostatic mechanism of the dressing is attributed to activating platelets, locally concentrating clotting components in the nanoclay, halloysite coagulation factors, and alginate cross-linked with Ca2+ . This work inspires robust self-assembly of clay nanotubes on textile fibers and offers a hemostatic material with balanced high hemostatic activity, minimal ingredient loss, and biocompatibility. The robust dressing based on halloysite tightly bounded cotton shows great potential for military, medical, and civil bleeding control with low health risks.

Hemostatic Cryogels Based on Oxidized Pullulan/Dopamine with Potential Use as Wound Dressings.

The impetus for research into hydrogels based on selectively oxidized polysaccharides has been stimulated by the diversity of potential biomedical applications. Towards the development of a hemostatic wound dressing in this study, we creatively combined the (hemi)acetal and Schiff base bonds to prepare a series of multifunctional cryogels based on dialdehyde pullulan and dopamine. The designed structures were verified by NMR and FTIR spectroscopy. Network parameters and dynamic sorption studies were correlated with environmental scanning microscopy results, thus confirming the successful integration of the two components and the opportunities for finely tuning the structure-properties balance. The viscoelastic parameters (storage and loss moduli, complex and apparent viscosities, zero shear viscosity, yield stress) and the structural recovery capacity after applying a large deformation were determined and discussed. The mechanical stability and hemostatic activity suggest that the optimal combination of selectively oxidized pullulan and dopamine can be a promising toolkit for wound management.

A topical platelet-independent multilevel clotting initiator for intraoperative hemostasis

Quick and stable clotting is essential in topically controlling intraoperative hemorrhage for patients with chemotherapy-induced thrombocytopenia. Here, we proposed and validated a kind of vesicle originated from gene-engineering cell membrane to manage uncontrolled bleeding with double clotting-triggering strategy. The vesicle constitutively exhibited truncated tissue factor (tTF) protein and phosphatidylserine (PS), thus function as a fibrin growth stimulus and erythrocyte attractant initiator in vitro and in vivo. Erythrocytes assembled in blood clot protected formed fibrin from fibrinolysis while the grown fibrin network enhanced erythrocyte aggregation in situ. Notably, these double clotting-triggering processes worked synergistically to constitute a positive feedback effect to quickly and potently form fibrinolysis-resisted erythrocyte-modified fibrin structures and achieve more stable clotting than the commonly used thrombin and gelatin coagulant. Furthermore, FDA-approved biocompatible Pluronic F127 hydrogel was rationally introduced to assist vesicles depositing onto the wound, thereby maximizing the hemostatic activity and minimizing the postoperative adhesion in a hemorrhage model with cyclophosphamide-induced myelosuppression. This multi-level coagulant method reported here sheds new light on controlling bleeding when patients with myelosuppression undergo surgery.

Silk Fibroin/Tannin/ZnO Nanocomposite Hydrogel with Hemostatic Activities.

The inevitable bleeding and infections caused by disasters and accidents are the main causes of death owing to extrinsic trauma. Hemostatic agents are often used to quickly suppress bleeding and infection, and they can solve this problem in a short time. Silk fibroin (SF) has poor processibility in water, owing to incomplete solubility therein. In this study, aiming to overcome this disadvantage, a modified silk fibroin (SF-BGE), easily soluble in water, was prepared by introducing butyl glycidyl ether (BGE) into its side chain. Subsequently, a small amount of tannic acid (TA) was introduced to prepare an SF-BGE /TA solution, and ZnO nanoparticles (NPs) were added to the solution to form the coordination bonds between the ZnO and TA, leading to an SF-based nanocomposite hydrogel. A structural characterization of the SF-BGE, SF-BGE/TA, SF-BGE/TA/ZnO, and the coordination bonds between ZnO/TA was observed by attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), and the phase change was observed by rheological measurements. The pore formation of the SF-BGE/TA/ZnO hydrogel and dispersibility of ZnO were verified through energy-dispersive X-ray spectroscopy (EDS) and scanning electron microscopy (SEM). The cytocompatible and hemostatic performances of the SF-BGE/TA/ZnO NPs composite hydrogels were evaluated, and the hydrogels showed superior hemostatic and cytocompatible activities. Therefore, the SF-based nanocomposite hydrogel is considered as a promising material for hemostasis.

Euphorbia species latex: A comprehensive review on phytochemistry and biological activities.

The genus Euphorbia includes about 2,000 species commonly widespread in both temperate and tropical zones that contain poisonous milky juice fluid or latex. Many species have been used in traditional and complementary medicine for the treatment of various health issues such as dropsy, paralysis, deafness, wounds, warts on the skin, and amaurosis. The medicinal applications of these species have been attributed to the presence of various compounds, and most studies on Euphorbia species have focused on their latex. In this review, we summarize the current state of knowledge on chemical composition and biological activities of the latex from various species of the genus Euphorbia. Our aim was to explore the applications of latex extracts in the medical field and to evaluate their ethnopharmacological potential. The databases employed for data collection, are obtained through Web of Science, PubMed, Google Scholar, Science Direct and Scopus, from 1983 to 2022. The bibliographic data indicate that terpenoids are the most common secondary metabolites in the latex. Furthermore, the latex has interesting biological properties and pharmacological functions, including antibacterial, antioxidant, free radical scavenger, cytotoxic, tumor, anti-inflammatory, healing, hemostatic, anti-angiogenic, insecticidal, genotoxic, and mutagenic activities. However, the role of other components in the latex, such as phenolic compounds, alkaloids, saponins, and flavonoids, remains unknown, which limits the application of the latex. Future studies are required to optimize the therapeutic use of latex extracts.

Screening for the extracorporeal coagulation activity quality markers (Q-markers) of dried and stir fried ginger

Ginger is a common condiment that is widely used as Chinese medicine in China and Southeast Asia. Dried ginger and stir fried ginger are two common processed products of ginger, with distinct clinical uses. The aim of this study was to identify the chemical components (quality markers, Q-mark) responsible for the differences in in vitro hemostatic activity between dried and stir fried ginger and to provide a basis for the selection between the two types of ginger in clinical application. In this study, methanolic extracts of dried and stir fried ginger were characterized using UPLC-Q/TOF-MS and then evaluated for in vitro coagulation activity. Spectral effect correlation analysis was used to identify quality markers, while molecular docking simulation was used to evaluate the binding energy between potential active compounds and target proteins. A total of 49 chemical constituents of the ginger extracts were identified using UPLC-Q/TOF-MS, 27 of which were significantly different between the two extracts. A fingerprint of 18 batches of dried and stir fried ginger established that zingiberone, 6-gingerol, 8-gingerol, 6-shogaol, 10-gingerol, 8-shogaol, and 10-shogaol were common constituents of the two extracts. Results of coagulation assays revealed that dried ginger had anti-coagulation effects, while stir fried ginger had hemostatic effect. Zingiberone, 6-shogaol and 10-shogaol were identified as the active components responsible for the hemostatic effect of Stir fried ginger through multivariate statistical analysis. In addition, molecular docking simulations suggested that these three components bound to Src proteins on platelets. Consequently, 6-gingerol, zingiberone, 6-shogaol and 10-shogaol were selected as quality markers to distinguish between dried and stir fried ginger. These results provide scientific evidence for the establishment of a quality evaluation system for the integrity and specificity of dried and stir fried ginger.

CHITOSAN ADDUCT WITH TRANEXAMIC ACID AND ITS HAEMOSTATIC EFFECT

Chitosan is a cationic polymer capable of binding acidic drugs. In addition, it has haemostatic and antimicrobial activity. Chitosan derivatives withanti-fibrinolytic properties may present increased effectiveness, especially when the added substance forms an adduct with chitosan. The aim of this work was to study the haemostatic action of the chitosan–tranexamic acid complex. Two chitosan solutions (molecularweight of 250 and 625 kDa at pH 5.7 and 6.2, and after tranexamic acid had been added to chitosan solutions) werestudied. Haemostatic evaluation was performed on white outbred mice. The time to complete cessation of bleeding from the tail was determined. Chitosan 625 kDa at pH 6.2 had the best haemostatic properties. Adding tranexamic acid to the chitosan solution reduced the bleeding time. This phenomenon was more pronounced for chitosan 625 kDa. Compared with control animals, this chitosan reduced bleeding arrest time by 30% and the chitosan–tranexamic acid adduct reduced the bleeding arrest time by 75%.

Development of cellulosic-based hemostatic dressing with antibacterial activity

A cotton-based hemostatic dressing featuring antibacterial properties was developed with the potential of being used in traffic accidents to control hemorrhage. Cotton gauze was oxidized initially in an acidic medium and then coated by PVA nanofibers and/or PVA nanofibers loaded with Ciprofloxacin. Fabricated dressings were characterized by FTIR analysis and SEM images. The FTIR spectrum showed the existence of carboxyl groups on the oxidized cotton gauze's surface. The carboxyl groups content was estimated to be 17.3 ± 0.3 for the oxidized sample with a mixture of nitric acid and phosphoric acid for 24 h (OCF-Mixed acid24). Moreover, the effect of the exposure duration of cotton gauze in the acidic medium on the blood coagulation activity was assessed. It was observed that the OCF-Mixed acid24 sample exhibited an agreeable hemostatic activity (BCIs = 10). The antibacterial activity against E. coli and S. aureus bacteria was also captured for the coated cotton gauze by the PVA nanofibers loaded with Ciprofloxacin.

Combination of natural polyanions and polycations based on interfacial complexation for multi-functionalization of wound dressings.

Multi-functionalization of wound dressings with natural polymers is meaningful and remains a challenge. The combination of natural polyanions and polycations appears to be a promising strategy. Still, its performances based on current layer-by-layer self-assembly or homogeneous complexation are mutable and limited. Herein, Ca2+-incorporated carboxymethyl cellulose (Ca/Na-CMC) and hydroxypropyltrimethyl ammonium chloride chitosan (HACC) are adopted as the model polyanion and polycation, respectively, to develop multi-functionalized dressings based on interfacial complexation. The dressings exhibit a multilayer structure composed of a polyanion layer (Ca/Na-CMC) for hemostasis and promotion of cell proliferation, a formed polyelectrolyte complex (PEC) layer for structural stability, and a polycation layer (HACC) for antibiosis. Compared to the dressing based on homogeneous complexation, the multilayer dressings show stronger moisture penetrability (around 1,150 g/m2/24 h), higher hemostatic activity, and higher antibacterial rate (up to 100%) and promoted effect on cell proliferation. An in vivo evaluation using a rat full-thickness skin defect model reveals that the multilayer dressings can accelerate wound healing in 2 weeks. Overall, owing to interfacial complexation resulting in separate layers, the performances of polyanions and polycations after combination are more predictable, and their biological functions can be effectively preserved. These findings not only support the extensive application of multilayer dressings but also offer an alternative strategy for multi-functionalizing wound dressings with natural polyanions and polycations.

Synthesis of 3,3-Dimethyl- and 3,3-Diethyl-Substituted 2-(3,4-Dihydroisoquinolin-1-yl)Carboxylic Acid Amide Hydrochlorides and Their Anti-Arrhythmic, Hemostatic, Anthelmintic, and Larvicidal Activity

Synthesis of 3,3-Dimethyl- and 3,3-Diethyl-Substituted 2-(3,4-Dihydroisoquinolin-1-yl)Carboxylic Acid Amide Hydrochlorides and Their Anti-Arrhythmic, Hemostatic, Anthelmintic, and Larvicidal Activity

The Effect on Hemostasis of Gelatin-Graphene Oxide Aerogels Loaded with Grape Skin Proanthocyanidins: In Vitro and In Vivo Evaluation.

Using in vitro and in vivo models, this study investigated the hemostatic potential to control bleeding of both unloaded gelatin-graphene oxide aerogels and the same loaded with proanthocyanidins (PAs) from Vitis vinifera grape skin extract. Our results showed that the physicochemical and mechanical properties of the aerogels were not affected by PA inclusion. In vitro studies showed that PA-loaded aerogels increased the surface charge, blood absorption capacity and cell viability compared to unloaded ones. These results are relevant for hemostasis, since a greater accumulation of blood cells on the aerogel surface favors aerogel–blood cell interactions. Although PAs alone were not able to promote hemostasis through extrinsic and intrinsic pathways, their incorporation into aerogels did not affect the in vitro hemostatic activity of these composites. In vivo studies demonstrated that both aerogels had significantly increased hemostatic performance compared to SpongostanTM and gauze sponge, and no noticeable effects of PA alone on the in vivo hemostatic performance of aerogels were observed; this may have been related to its poor diffusion from the aerogel matrix. Thus, PAs have a positive effect on hemostasis when incorporated into aerogels, although further studies should be conducted to elucidate the role of this extract in the different stages of hemostasis.

Thromboelastography and thrombin generation assessments for pediatric severe hemophilia A patients are highly variable and not predictive of clinical phenotypes

BackgroundSevere hemophilia A (SHA) patients vary in severity of bleeding, arthropathy, and requirements for replacement factor VIII (FVIII). Baseline hemostatic activity assays using calibrated automated thrombography (CAT) and thromboelastography (TEG) may offer insights into the physiological basis of clinical heterogeneity. ObjectivesUse CAT and TEG to measure baseline hemostatic activity in a cohort of 30 pediatric SHA patients with available clinical data. Determine effect of contact activation inhibition with corn trypsin inhibitor (CTI). Assess heterogeneity among patients for baseline hemostatic activity and examine correlations between assay results and clinical parameters including FVIII dosing regimen, von Willebrand factor level, and Pettersson arthropathy score. MethodsSHA blood after FVIII washout was subjected to TEG, and platelet‐rich (PRP) and platelet‐poor plasma was used for CAT assays. Varying concentrations of tissue factor (TF) were used. Statistical analysis examined relationships between assay results, and clinical parameters. ResultsCTI treatment was required to obtain TEG and CAT results representative of baseline hemostatic activity. Weak activity was observed in assays with low TF concentrations (0.5–2 pM), and most but not all samples approached normal activity levels at high TF concentrations (10–20 pM). A significant positive correlation was observed between results of TEG and CAT‐PRP assays. Correlations were not detected between hemostatic assay results and clinical parameters. ConclusionsIn vitro hemostatic assay results of samples containing platelets showed concordance. Assay results were not predictive of FVIII requirements or correlated with other clinical parameters. SHA patient heterogeneity is influenced by factors other than baseline hemostatic activity.

Tunicate inspired gelatin-based tough hydrogel wound dressing containing twisted phthalazinone with adhesive, self-healing and antibacterial properties

As a hydrolytic product of collagen, gelatin is a polypeptide of biological origin. Gelatin hydrogels emerge as promising material candidates for traditional dressings due to good biocompatibility and the ability to keep wounds moist. However, it is difficult to simultaneously achieve gelatin hydrogel with robust mechanical property for long-term usage, reliable tissue adhesion, self-healing and antibacterial properties. Herein, we propose a simply synthesized strategy of a multifunctional gelatin hydrogel dressing, which is constructed by conjugating a newly synthesized 2-(4′-aldehydephenyl)-4-(2′,3′,4′-trihydroxyphenyl)-2,3-phthalazine-1(2H)-one (THPZB) to gelatin with Schiff base and chelating with Fe3+ ions (termed G/THPZB/Fe hydrogel). The twisted structure of phthalazinone in THPZB leads to entanglement of gelatin molecular chains, which resolves the stiffness-toughness conflict of the hydrogel. Furthermore, the strong tissue adhesion and fast self-healing capability mainly originate from the hydrogen bonding of the pyrogallol in THPZB. In vitro study shows that the hydrogels possess good biocompatibility with L929 cells, hemostatic and antibacterial activity. In the rat model of skin infection, the hydrogel dressing not only have no adverse effects on vital organs, but also can effectively promote wound healing of bacterial infection. Considering that it has multiple functions, G/THPZB/Fe hydrogel can be used as a promising wound dressing for biomedical applications.

Injectable Self-Healing Adhesive Chitosan Hydrogel with Antioxidative, Antibacterial, and Hemostatic Activities for Rapid Hemostasis and Skin Wound Healing.

Engineered wound dressing materials with excellent injectability, self-healing ability, tissue-adhesiveness, especially the ones possessing potential therapeutic effects have great practical significance in healthcare. Herein, an injectable quaternary ammonium chitosan (QCS)/tannic acid (TA) hydrogel based on QCS and TA was designed and fabricated by facile mixing of the two ingredients under physiological conditions. In this system, hydrogels were mainly cross-linked by dynamic ionic bonds and hydrogen bonds between QCS and TA, which endows the hydrogel with excellent injectable, self-healing, and adhesive properties. Benefitting from the inherent antioxidative, antibacterial, and hemostatic abilities of TA and QCS, this hydrogel showed superior reactive oxygen species scavenging activity, broad-spectrum antibacterial ability, as well as rapid hemostatic capability. Moreover, the QCS/TA2.5 hydrogel (containing 2.5% TA) exhibited excellent biocompatibility. The in vivo experiments also showed that QCS/TA2.5 hydrogel dressing not only rapidly stopped the bleeding of arterial and deep incompressible wounds in mouse tail amputation, femoral artery hemorrhage, and liver incision models but also significantly accelerated wound healing in a full-thickness skin wound model. For the great potentials listed above, this multifunctional QCS/TA2.5 hydrogel offers a promising network as a dressing material for both rapid hemostasis and skin wound repair.

Mussel-inspired polysaccharide-based sponges for hemostasis and bacteria infected wound healing

Effective bleeding control and wound protecting from infection play critical roles in the tissue healing process. However, local hemostats are not involved in the whole healing processes to promote the final healing efficiency. Here, a multi-functional mussel-inspired polysaccharide-based sponge with hemostatic, antibacterial and adhesive properties was fabricated via cryopolymerization of oxidized dextran (OD), carboxymethyl chitosan (CC) and polydopamine nanoparticles (PDA-NPs), followed by lyophilization. Combining with the adsorbed thrombin, the sponges yielded a considerably lower amount of blood than the commercially available hemostatic dressings. Benefiting from the high photo-thermal transition efficiency of PDA-NPs, the sponges exhibited excellent antibacterial activity to both gram positive and negative bacteria. Owing to the rapid hemostatic activity and effective infection resistance, the sponges illustrated the significantly acceleratory wound healing efficiency compared with the control group. The thrombin-loaded OD/CC-PDA polysaccharide-based sponge has great potential for future clinical use as wound dressing.

Optimization and evaluation of a two-stage chromogenic assay procedure for measurement of emicizumab plasma levels.

Emicizumab mimics the hemostatic activity of activated factor VIII (FVIIIa) within the tenase complex. Despite functional similarities between FVIIIa and emicizumab, conventional laboratory methods designed for monitoring of FVIII activity are inappropriate for the measurement of emicizumab. At present, a modified one stage (FVIII) assay (mOSA) is mainly used for emicizumab monitoring. Two-stage chromogenic FVIII assays based on human factors can be used, although limited performance due to lack of corresponding optimization might be observed. Furthermore, the presence of FVIII or anticoagulants in the patient sample may falsify assay results. To address these issues, we optimized and evaluated a two-stage chromogenic assay (emi-tenase) for measurement of emicizumab in plasma samples. Heat inactivation of samples was established to abolish the influence of endogenous or substituted FVIII. The lower limit of quantification (LLoQ) was found to be 2 μg/ml in a manual assay format and 9.5 μg/ml on an automated coagulation analyzer. Intra- and inter-assay coefficients of variation (CV) did not exceed 20%. Analysis of 17 patient plasma samples with severe haemophilia A under emicizumab treatment showed good correlation of results between the emi-tenase assay and the mOSA (Cohens Kappa coefficient = 0.9). Taken together, the emi-tenase assay allows specific measurement of emicizumab plasma levels over a broad concentration range (10 μg/ml to 100 μg/ml). The assay can be applied on an automated coagulation analyzer, demonstrating its applicability within a routine laboratory setting.