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1.
Lab Chip ; 21(19): 3667-3674, 2021 09 28.
Artigo em Inglês | MEDLINE | ID: mdl-34476426

RESUMO

Point-of-care diagnostics of platelet and coagulation function present demanding challenges. Current clinical diagnostics often use centrifuged plasmas or platelets and frozen plasma standards, recombinant protein standards, or even venoms. Almost all commercialized tests of blood do not recreate the in vivo hemodynamics where platelets accumulate to high densities and thrombin is generated from a procoagulant surface. Despite numerous drugs that target platelets, insufficient coagulation, or excess coagulation, POC blood testing is essentially limited to viscoelastic methods that provide a clotting time, clot strength, and clot lysis, while used mostly in trauma centers with specialized capabilities. Microfluidics now allows small volumes of whole blood (<1 mL) to be tested under venous or arterial shear rates with multi-color readouts to follow platelet function, thrombin generation, fibrin production, and clot stability. Injection molded chips containing pre-patterned fibrillar collagen and lipidated tissue factor can be stored dry for 6 months at 4C, thus allowing rapid blood testing on single-use disposable chips. Using only a small imaging microscope and micropump, these microfluidic devices can detect platelet inhibitors, direct oral anticoagulants (DOACs) and their reversal agents. POC microfluidics are ideal for neonatal surgical applications that involve small blood samples, rapid DOAC testing in stroke or bleeding or emergency surgery situations with patients presenting high risk cofactors for either bleeding or thrombosis.


Assuntos
Microfluídica , Trombose , Coagulação Sanguínea , Plaquetas , Fibrina , Humanos , Recém-Nascido , Sistemas Automatizados de Assistência Junto ao Leito , Trombina , Trombose/diagnóstico
2.
Clin Lab ; 67(9)2021 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-34542980

RESUMO

BACKGROUND: D-dimer is a molecular marker of fibrin degradation and fibrinolytic system activation and is an effective indicator for early diagnosis of thrombotic diseases, monitoring of thrombolytic therapy, and efficacy evaluation; therefore, the accuracy of D-dimer test results is of great significance to the diagnosis and treatment of diseases in clinical practice. METHODS: This paper reports two cases of pseudoelevation of plasma D-dimer levels by different test systems. RESULTS: In case 1, the abnormal increases in the results of the ACL TOP 700 analyzer were considered to be the pseudoelevation caused by interferences, and the abnormal increases in the STA-R Max results were considered to be the pseudoelevation caused by interferences in case 2. CONCLUSIONS: Laboratories should be equipped with two different brands of test systems and reagents to identify suspicious test results in a timely manner and avoid adverse events.


Assuntos
Produtos de Degradação da Fibrina e do Fibrinogênio , Trombose , Fibrina , Humanos , Terapia Trombolítica
3.
Mater Sci Eng C Mater Biol Appl ; 128: 112304, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34474855

RESUMO

Delayed wound healing in heavily irradiated areas is a serious clinical complication that makes widespread therapeutic use of radiation difficult. Efficient treatment strategies are urgently required for addressing radiation-induced wound failure. Herein, we applied liquid-type nonthermal atmospheric plasma (LTP) to a silk-fibrin (SF) composite gel to investigate whether controlled release of LTP from SF hydrogel not only induced favorable cellular events in an irradiated wound bed but also modulated the SF hydrogel microstructure itself, eventually facilitating the development of a regenerative microenvironment. Scanning electron microscopy and Fourier-transform infrared spectroscopy revealed that LTP modulated the microstructures and chemical bindings of the SF gel. Improved cell viability, morphology, and extracellular matrix depositions by the LTP-treated SF hydrogel were identified with wound-healing assays and immunofluorescence staining. An irradiated random-pattern skin-flap animal model was established in six-week-old C57/BL6 mice. Full-thickness skin was flapped from the dorsum and SF hydrogel was placed underneath the raised skin flap. Postoperative histological analysis of the irradiated random-pattern skin-flap mice model suggested that LTP-treated SF hydrogel much improved wound regeneration and the inflammatory response compared to the SF hydrogel- and sham-treated groups. These results support that LTP-treated SF hydrogel significantly enhanced irradiated wound healing. Cellular and tissue reactions to released LTP from the SF hydrogel were favorable for the regenerative process of the wound; furthermore, mechanochemical properties of the SF gel were improved by LTP.


Assuntos
Fibroínas , Seda , Animais , Fibrina , Hidrogéis , Camundongos , Cicatrização
4.
Mater Sci Eng C Mater Biol Appl ; 128: 112352, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34474900

RESUMO

Human plasma-derived bilayered skin substitutes have been successfully used by our group in different skin tissue engineering applications. However, several issues associated with their poor mechanical properties were observed, and they often resulted in rapid contraction and degradation. In this sense, hydrogels composed of plasma-derived fibrin and thiolated-hyaluronic acid (HA-SH, 0.05-0.2% w/v) crosslinked with poly(ethylene glycol) diacrylate (PEGDA, 2:1, 6:1, 10:1 and 14:1 mol of thiol to moles of acrylate) were developed to reduce the shrinking rates and enhance the mechanical properties of the plasma-derived matrices. Plasma/HA-SH-PEGDA hydrogels showed a decrease in the contraction behaviour ranging from 5% to 25% and an increase in Young's modulus. Furthermore, the results showed that a minimal amount of the added HA-SH was able to escape the plasma/HA-SH-PEGDA hydrogels after incubation in PBS. The results showed that the increase in rigidity of the matrices as well as the absence of adhesion cellular moieties in the second network of HA-SH/PEGDA, resulted in a decrease in contraction in the presence of the encapsulated primary human fibroblasts (hFBs), which may have been related to an overall decrease in proliferation of hFBs found for all hydrogels after 7 days with respect to the plasma control. The metabolic activity of hFB returned to the control levels at 14 days except for the 2:1 PEGDA crosslinking ratio. The metabolic activity of primary human keratinocytes (hKCs) seeded on the hydrogels showed a decrease when high amounts of HA-SH and PEGDA crosslinker were incorporated. Organotypic skins formed in vitro after 21 days with plasma/HA-SH-PEGDA hydrogels with an HA content of 0.05% w/v and a 2:1 crosslinking ratio were up to three times thicker than the plasma controls, evidencing a reduction in contraction, while they also showed better and more homogeneous keratin 10 (K10) expression in the supra-basal layer of the epidermis. Furthermore, filaggrin expression showed the formation of an enhanced stratum corneum for the constructs containing HA. These promising results indicate the potential of using these biomimetic hydrogels as in vitro skin models for pharmaceutical products and cosmetics and future work will elucidate their potential functionality for clinical treatment.


Assuntos
Hidrogéis , Pele Artificial , Epiderme , Fibrina , Humanos , Ácido Hialurônico , Engenharia Tecidual
5.
Cell Mol Life Sci ; 78(17-18): 6251-6264, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34398251

RESUMO

Ischemic stroke is one of the most serious diseases today, and only a minority of patients are provided with effective clinical treatment. Importantly, leukocytes have gradually been discovered to play vital roles in stroke thrombosis, including promoting the activation of thrombin and the adhesion and aggregation of platelets. However, they have not received enough attention in the field of acute ischemic stroke. It is possible that we could not only prevent stroke-related thrombosis by inhibiting leukocyte activation, but also target leukocyte components to dissolve thrombi in the cerebral artery. In this review, we expound the mechanisms by which leukocytes are activated and participate in the formation of stroke thrombus, then describe the histopathology of leukocytes in thrombi of stroke patients and the influence of leukocyte composition on vascular recanalization effects and patient prognosis. Finally, we discuss the relevant antithrombotic strategies targeting leukocytes.


Assuntos
AVC Isquêmico/patologia , Leucócitos/metabolismo , Trombose/patologia , Armadilhas Extracelulares/efeitos dos fármacos , Armadilhas Extracelulares/metabolismo , Fibrina/metabolismo , Humanos , Sistema Imunitário/metabolismo , AVC Isquêmico/complicações , Ativação Plaquetária , Prognóstico , Trombose/tratamento farmacológico , Trombose/etiologia , Ativador de Plasminogênio Tecidual/farmacologia , Ativador de Plasminogênio Tecidual/uso terapêutico
6.
Dokl Biochem Biophys ; 499(1): 242-246, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34426920

RESUMO

Using the methods of dynamic and elastic light scattering and confocal laser scanning microscopy, the damage in the spatial fibrin structure during peroxide- and hypochlorite-induced oxidation of fibrinogen was studied. Peroxide had a weak effect on the structural organization of fibrin, whereas hypochlorite caused the formation of abnormal fibrin with reduced individual fiber diameter and decreased porosity. Measurements of the size distributions of the native and oxidized fibrinogen revealed a decrease in the hydrodynamic size of the oxidized fibrinogen molecule with an increase in the concentration of oxidizers. These results indicate that the hydrophobicity of fibrinogen surface increased and its colloidal stability decreased. The possible role of oxidative sites in the assembly of structurally abnormal fibrin is analyzed.


Assuntos
Fibrina/química , Fibrinogênio/metabolismo , Ácido Hipocloroso/farmacologia , Peróxidos/farmacologia , Fibrina/metabolismo , Oxirredução/efeitos dos fármacos
7.
J Immunol ; 207(6): 1641-1651, 2021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34380648

RESUMO

Thrombin activation of C5 connects thrombosis to inflammation. Complement research in whole blood ex vivo necessitates anticoagulation, which potentially interferes with the inflammatory modulation by thrombin. We challenged the concept of thrombin as an activator of native C5 by analyzing complement activation and C5 cleavage in human whole blood anticoagulated with Gly-Pro-Arg-Pro (GPRP), a peptide targeting fibrin polymerization downstream of thrombin, allowing complete endogenous thrombin generation. GPRP dose-dependently inhibited coagulation but allowed for platelet activation in accordance with thrombin generation. Spontaneous and bacterial-induced complement activation by Escherichia coli and Staphylococcus aureus, analyzed at the level of C3 and C5, were similar in blood anticoagulated with GPRP and the thrombin inhibitor lepirudin. In the GPRP model, endogenous thrombin, even at supra-physiologic concentrations, did not cleave native C5, despite efficiently cleaving commercially sourced purified C5 protein, both in buffer and when added to C5-deficient serum. In normal serum, only exogenously added, commercially sourced C5 was cleaved, whereas the native plasma C5 remained intact. Crucially, affinity-purified C5, eluted under mild conditions using an MgCl2 solution, was not cleaved by thrombin. Acidification of plasma to pH ≤ 6.8 by hydrochloric or lactic acid induced a C5 antigenic change, nonreversible by pH neutralization, that permitted cleavage by thrombin. Circular dichroism on purified C5 confirmed the structural change during acidification. Thus, we propose that pH-induced conformational change allows thrombin-mediated cleavage of C5 and that, contrary to previous reports, thrombin does not cleave plasma C5 in its native form, suggesting that thrombin cleavage of C5 may be restricted to certain pathophysiological conditions.


Assuntos
Complemento C5 , Trombina , Coagulação Sanguínea , Ativação do Complemento , Fibrina , Humanos
8.
Gen Dent ; 69(5): 14-19, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34424206

RESUMO

The use of autogenous materials to promote tissue regeneration has guided the direction of modern dentistry, and platelet-rich fibrin (PRF) is a promising biomaterial for tissue engineering. This in vitro immunohistochemical study aimed to analyze the presence of factors of endothelial growth and cell differentiation in PRF membranes by using the CD31 (endothelial cells) and CD163 (monocytes) markers. Five men and 5 women, aged between 25 and 60 years and without systemic health problems, were enrolled in the study. Blood samples were collected, submitted to a centrifugation protocol, and fixed in 4% formaldehyde, and then immunohistochemical analysis was performed. The histologic analysis of the slides showed that the fibrin clot was formed by a dense fiber network and cells trapped in its structure. One sample was excluded from the markers testing due to poor quality. All 9 of the valid samples were positive for the CD31 and CD163 markers, with reactivity ranging from 5% to 30% and 10% to 40% of cells, respectively. The immunohistochemical analysis showed the presence of CD31 and CD163 in the PRF membranes, indicating the potential for vascular neoformation and the significant presence of monocytes, which play an important role in tissue remodeling via their differentiation into macrophages.


Assuntos
Fibrina Rica em Plaquetas , Adulto , Plaquetas , Centrifugação , Células Endoteliais , Feminino , Fibrina , Humanos , Masculino , Pessoa de Meia-Idade
9.
Front Cell Infect Microbiol ; 11: 708739, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34277477

RESUMO

Leptospirosis is a neglected zoonosis, caused by pathogenic spirochetes bacteria of the genus Leptospira. The molecular mechanisms of leptospirosis infection are complex, and it is becoming clear that leptospires express several functionally redundant proteins to invade, disseminate, and escape the host's immune response. Here, we describe a novel leptospiral protein encoded by the gene LIC13086 as an outer membrane protein. The recombinant protein LIC13086 can interact with the extracellular matrix component laminin and bind plasminogen, thus possibly participating during the adhesion process and dissemination. Also, by interacting with fibrinogen and plasma fibronectin, the protein LIC13086 probably has an inhibitory effect in the fibrin clot formation during the infection process. The newly characterized protein can also bind molecules of the complement system and the regulator C4BP and, thus, might have a role in the evasion mechanism of Leptospira. Taken together, our results suggest that the protein LIC13086 may have a multifunctional role in leptospiral pathogenesis, participating in host invasion, dissemination, and immune evasion processes.


Assuntos
Leptospira interrogans , Leptospira , Leptospirose , Fibrina/metabolismo , Humanos , Leptospira interrogans/genética , Leptospira interrogans/metabolismo , Plasminogênio/metabolismo , Ligação Proteica
10.
Arterioscler Thromb Vasc Biol ; 41(9): 2370-2383, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34261330

RESUMO

Thrombosis is a major complication of cardiovascular disease, leading to myocardial infarction, acute ischemic stroke, or venous thromboembolism. Thrombosis occurs when a thrombus forms inside blood vessels disrupting blood flow. Developments in thrombectomy to remove thrombi from vessels have provided new opportunities to study thrombus composition which may help to understand mechanisms of disease and underpin improvements in treatments. We aimed to review thrombus compositions, roles of components in thrombus formation and stability, and methods to investigate thrombi. Also, we summarize studies on thrombus structure obtained from cardiovascular patients and animal models. Thrombi are composed of fibrin, red blood cells, platelets, leukocytes, and neutrophil extracellular traps. These components have been analyzed by several techniques, including scanning electron microscopy, laser scanning confocal microscopy, histochemistry, and immunohistochemistry; however, each technique has advantages and limitations. Thrombi are heterogenous in composition, but overall, thrombi obtained from myocardial infarction are composed of mainly fibrin and other components, including platelets, red blood cells, leukocytes, and cholesterol crystals. Thrombi from patients with acute ischemic stroke are characterized by red blood cell- and platelet-rich regions. Thrombi from patients with venous thromboembolism contain mainly red blood cells and fibrin with some platelets and leukocytes. Thrombus composition from patients with myocardial infarction is influenced by ischemic time. Animal thrombosis models are crucial to gain further mechanistic information about thrombosis and thrombus structure, with thrombi being similar in composition compared with those from patients. Further studies on thrombus composition and function are key to improve treatment and clinical outcome of thrombosis.


Assuntos
Coagulação Sanguínea , Plaquetas/metabolismo , Eritrócitos/metabolismo , Fibrina/metabolismo , Trombose/metabolismo , Animais , Plaquetas/patologia , Colesterol/metabolismo , Modelos Animais de Doenças , Eritrócitos/patologia , Humanos , Leucócitos/metabolismo , Trombectomia , Trombose/patologia , Trombose/terapia
11.
Biomed Res Int ; 2021: 9996071, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34307687

RESUMO

Background: Platelet concentrates like leucocyte- and platelet-rich fibrin (L-PRF) have been widely evaluated in different oral surgical procedures to promote the healing process. However, liquid L-PRF products such as liquid fibrinogen have been poorly explored, especially in the biomimetic functionalization of dental implants. The aim of this in vitro study is to evaluate the interaction between 5 different dental implant surfaces and liquid fibrinogen. Methods: Five commercially available dental implants with different surfaces (Osseospeed™, TiUnite™, SLActive®, Ossean®, and Plenum®) were immersed for 60 minutes in liquid fibrinogen obtained from healthy donors. After this period, the implants were removed and fixed for scanning electron microscopy (SEM). Results: All dental implants were covered by a fibrin mesh. However, noticeable noncontact areas were observed for the Osseospeed™, TiUnite™, and SLActive® surfaces. On the other hand, Ossean® and Plenum® surfaces showed a dense and uniform layer of fibrin covering almost the entire implant surface. The Osseospeed™, TiUnite™, and SLActive® surfaces presented with lower blood cell numbers inside the fibrin mesh compared with the others. Moreover, at higher magnification, thicker fibrin fibers were observed in contact with Ossean® and Plenum® surfaces. The Plenum ®surface showed the thickest fibers which also inserted and interconnect to the microroughness. Conclusion: The initial contact between an implant surface and the fibrin network differs significantly among different implant brands. Further studies are necessary to explore the clinical impact of these observations in the osseointegration process of dental implants.


Assuntos
Fibrinogênio/metabolismo , Implantes Dentários , Fibrina/metabolismo , Humanos , Microscopia Eletrônica de Varredura , Projetos Piloto
12.
Medicine (Baltimore) ; 100(27): e26603, 2021 Jul 09.
Artigo em Inglês | MEDLINE | ID: mdl-34232212

RESUMO

RATIONALE: The amount of aqueous humor that is constantly produced by the ciliary processes and the equal amount that flows out through the trabecular meshwork via the drainage angle or through the uveoscleral route is very small (2-3 µg/min each), representing approximately 1% of the content of the anterior chamber; therefore, it is challenging to visualize its flow. PATIENT CONCERNS: A 69-year-old man who had high intraocular pressure (IOP) (>20 mm Hg) with the maximum glaucoma eyedrop dose and presented with severe visual field loss (Humphrey Field Analyzer 30-2: -26.32 dB) had been implanted with a 350-mm2 Baerveldt tube of the aqueous chamber type for refractory open-angle glaucoma. The IOP ultimately decreased (<15 mm Hg) with no need for glaucoma eyedrops. DIAGNOSES: After the procedures, a fibrin membrane repeatedly formed on the anterior surface of the intraocular lens. INTERVENTIONS: This issue was resolved by two rounds of neodymium-doped yttrium aluminum granet (Nd:YAG) laser surgery and prescription steroidal eyedrops. OUTCOMES: During the laser surgery, an unusual and unintended fibrin flap appeared at the opening of the Baerveldt tube; this flap moved synchronously with the heartbeat, as verified by checking the pulse at the radial artery of the wrist. The fibrin flap mimicked the behavior of a cardiac valve, and the aqueous humor and stray fibrin particles mimicked the blood in the chambers of the heart. Although the Baerveldt tube itself is an artificial instrument that is not present in normal human eyes, we hypothesize that our observation shows the fundamental mechanism of aqueous humor drainage. LESSONS: This novel, vividly descriptive observation highlights the important role of the heartbeat as a drainage pump in aqueous humor flow dynamics and IOP homeostasis, which are treatment targets for glaucoma, the leading cause of blindness.


Assuntos
Fibrina , Implantes para Drenagem de Glaucoma , Glaucoma de Ângulo Aberto/cirurgia , Frequência Cardíaca/fisiologia , Pressão Intraocular/fisiologia , Retalhos Cirúrgicos , Trabeculectomia/métodos , Idoso , Humor Aquoso/metabolismo , Glaucoma de Ângulo Aberto/fisiopatologia , Humanos , Terapia a Laser/métodos , Lasers de Estado Sólido/uso terapêutico , Masculino , Acuidade Visual
13.
Int J Mol Sci ; 22(13)2021 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-34203139

RESUMO

Fibrinogen is one of the key molecular players in haemostasis. Thrombin-mediated release of fibrinopeptides from fibrinogen converts this soluble protein into a network of fibrin fibres that form a building block for blood clots. Thrombin-activated factor XIII further crosslinks the fibrin fibres and incorporates antifibrinolytic proteins into the network, thus stabilising the clot. The conversion of fibrinogen to fibrin also exposes binding sites for fibrinolytic proteins to limit clot formation and avoid unwanted extension of the fibrin fibres. Altered clot structure and/or incorporation of antifibrinolytic proteins into fibrin networks disturbs the delicate equilibrium between clot formation and lysis, resulting in either unstable clots (predisposing to bleeding events) or persistent clots that are resistant to lysis (increasing risk of thrombosis). In this review, we discuss the factors responsible for alterations in fibrin(ogen) that can modulate clot stability, in turn predisposing to abnormal haemostasis. We also explore the mechanistic pathways that may allow the use of fibrinogen as a potential therapeutic target to treat vascular thrombosis or bleeding disorders. Better understanding of fibrinogen function will help to devise future effective and safe therapies to modulate thrombosis and bleeding risk, while maintaining the fine balance between clot formation and lysis.


Assuntos
Fator XIIIa/metabolismo , Fibrina/metabolismo , Fibrinogênio/metabolismo , Trombose/metabolismo , Animais , Fator XIIIa/genética , Fibrina/genética , Fibrinogênio/genética , Fibrinólise/genética , Fibrinólise/fisiologia , Humanos , Trombose/genética
14.
Transfusion ; 61 Suppl 1: S68-S79, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34269433

RESUMO

Although it is well established that transfusion of platelets in cases of severe bleeding reduces mortality, the availability of platelets is hampered by harsh restrictions on shelf life due to elevated risks of microbial contamination and functional losses with room temperature-stored platelets (RTP) kept at 22°C. In contrast, many recent studies have shown that 4°C cold-stored platelets (CSP) are able to overcome these shortcomings leading to the recent Food and Drug Administration licensure for 14-day stored CSP when conventional platelets are unavailable. This work expands the evidence supporting superiority of CSP function by assaying the less explored platelet-mediated clot retraction of RTP and CSP in either autologous plasma (AP) or platelet additive solution (PAS) for up to 21 days. The results demonstrate that CSP have better preservation of contractile function, exhibiting retraction for up to 21 days in both AP and PAS and forming highly ordered fibrin scaffolds similar to those of fresh platelets. In contrast, RTP stored in AP showed impaired contractile function by Day 5 with no retraction after 10 days, whereas PAS-stored RTP retained contractile function for up to 21 days. Collectively, these findings support extended storage of CSP and suggest that storage in PAS can mitigate functional losses in RTP.


Assuntos
Plaquetas/citologia , Preservação de Sangue/métodos , Coagulação Sanguínea , Plaquetas/metabolismo , Fibrina/metabolismo , Humanos , Testes de Função Plaquetária , Refrigeração , Temperatura
15.
Acta Biomater ; 131: 355-369, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34233219

RESUMO

Fibrin is the major determinant of the mechanical stability and integrity of blood clots and thrombi. To explore the rupture of blood clots, emulating thrombus breakage, we stretched fibrin gels with single-edge cracks of varying size. Ultrastructural alterations of the fibrin network correlated with three regimes of stress vs. strain profiles: the weakly non-linear regime due to alignment of fibrin fibers; linear regime owing to further alignment and stretching of fibers; and the rupture regime for large deformations reaching the critical strain and stress, at which irreversible breakage of fibers ahead of the crack tip occurs. To interpret the stress-strain curves, we developed a new Fluctuating Spring model, which maps the fibrin alignment at the characteristic strain, network stretching with the Young modulus, and simultaneous cooperative rupture of coupled fibrin fibers into a theoretical framework to obtain the closed-form expressions for the strain-dependent stress profiles. Cracks render network rupture stochastic, and the free energy change for fiber deformation and rupture decreases with the crack length, making network rupture more spontaneous. By contrast, mechanical cooperativity due to the presence of inter-fiber contacts strengthens fibrin networks. The results obtained provide a fundamental understanding of blood clot breakage that underlies thrombotic embolization. STATEMENT OF SIGNIFICANCE: Fibrin, a naturally occurring biomaterial, is the major determinant of mechanical stability and integrity of blood clots and obstructive thrombi. We tested mechanically fibrin gels with single-edge cracks and followed ultrastructural alterations of the fibrin network. Rupture of fibrin gel involves initial alignment and elastic stretching of fibers followed by their eventual rupture for deformations reaching the critical level. To interpret the stress-strain curves, we developed Fluctuating Spring model, which showed that cracks render rupture of fibrin networks more spontaneous; yet, coupled fibrin fibers reinforce cracked fibrin networks. The results obtained provide fundamental understanding of blood clot breakage that underlies thrombotic embolization. Fluctuating Spring model can be applied to other protein networks with cracks and to interpret the stress-strain profiles.


Assuntos
Fibrina , Trombose , Fenômenos Biomecânicos , Módulo de Elasticidade , Humanos , Termodinâmica
16.
Biosci Rep ; 41(8)2021 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-34328172

RESUMO

Severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2)-induced infection, the cause of coronavirus disease 2019 (COVID-19), is characterized by unprecedented clinical pathologies. One of the most important pathologies, is hypercoagulation and microclots in the lungs of patients. Here we study the effect of isolated SARS-CoV-2 spike protein S1 subunit as potential inflammagen sui generis. Using scanning electron and fluorescence microscopy as well as mass spectrometry, we investigate the potential of this inflammagen to interact with platelets and fibrin(ogen) directly to cause blood hypercoagulation. Using platelet-poor plasma (PPP), we show that spike protein may interfere with blood flow. Mass spectrometry also showed that when spike protein S1 is added to healthy PPP, it results in structural changes to ß and γ fibrin(ogen), complement 3, and prothrombin. These proteins were substantially resistant to trypsinization, in the presence of spike protein S1. Here we suggest that, in part, the presence of spike protein in circulation may contribute to the hypercoagulation in COVID-19 positive patients and may cause substantial impairment of fibrinolysis. Such lytic impairment may result in the persistent large microclots we have noted here and previously in plasma samples of COVID-19 patients. This observation may have important clinical relevance in the treatment of hypercoagulability in COVID-19 patients.


Assuntos
COVID-19/patologia , Fibrina/metabolismo , Fibrinólise/fisiologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Trombose/patologia , Adulto , Idoso , Amiloide/metabolismo , Plaquetas/metabolismo , Complemento C3/metabolismo , Feminino , Fibrinogênio/metabolismo , Humanos , Pulmão/patologia , Masculino , Técnicas Analíticas Microfluídicas , Pessoa de Meia-Idade , Protrombina/metabolismo , SARS-CoV-2/metabolismo , Trombose/virologia , Tripsina/metabolismo
17.
Int J Mol Sci ; 22(13)2021 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-34201667

RESUMO

Human plasma-derived bilayered skin substitutes were successfully used by our group to produce human-based in vitro skin models for toxicity, cosmetic, and pharmaceutical testing. However, mechanical weakness, which causes the plasma-derived fibrin matrices to contract significantly, led us to attempt to improve their stability. In this work, we studied whether an increase in fibrin concentration from 1.2 to 2.4 mg/mL (which is the useful fibrinogen concentration range that can be obtained from plasma) improves the matrix and, hence, the performance of the in vitro skin cultures. The results show that this increase in fibrin concentration indeed affected the mechanical properties by doubling the elastic moduli and the maximum load. A structural analysis indicated a decreased porosity for the 2.4 mg/mL hydrogels, which can help explain this mechanical behavior. The contraction was clearly reduced for the 2.4 mg/mL matrices, which also allowed for the growth and proliferation of primary fibroblasts and keratinocytes, although at a somewhat reduced rate compared to the 1.2 mg/mL gels. Finally, both concentrations of fibrin gave rise to organotypic skin cultures with a fully differentiated epidermis, although their lifespans were longer (25-35%) in cultures with more concentrated matrices, which improves their usefulness. These systems will allow the generation of much better in vitro skin models for the testing of drugs, cosmetics and chemicals, or even to "personalized" skin for the diagnosis or determination of the most effective treatment possible.


Assuntos
Diferenciação Celular , Derme/citologia , Epiderme/fisiologia , Fibrina/metabolismo , Hidrogéis/metabolismo , Queratinócitos/citologia , Tecidos Suporte/química , Proliferação de Células , Células Cultivadas , Derme/metabolismo , Fibroblastos/citologia , Fibroblastos/metabolismo , Humanos , Hidrogéis/química , Queratinócitos/metabolismo , Pele/citologia , Pele/metabolismo , Engenharia Tecidual
18.
Pediatr Dev Pathol ; 24(5): 450-454, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34082613

RESUMO

An emerging complication of COVID-19 (SARS-CoV-2) infection is reported. A 23-year-old patient presented with high temperature and reduced fetal movements at 25 + 5/40 weeks of gestation. RT-PCR proved maternal COVID-19 infection. Ultrasound examination confirmed intrauterine death. Placenta histology showed necrosis of the villous trophoblast, associated with Chronic Histiocytic Intervillositis (CHI) and Massive Perivillous Fibrin Deposition (MPFD) with up to 90% - of the intervillous spaces being involved. Immunohistochemistry showed CD68 positive histiocytes in the intervillous spaces and the villous trophoblast was positive for the COVID-19 spike protein. RNA scope signal was indicative of the presence of the viral genome and active viral replication in the villous trophoblastic cells, respectively. MPFD is a gradually developing end-stage disease with various etiology, including autoimmune and alloimmune maternal response to antigens expressed at the feto-maternal interface and frequently accompanies chronic alloimmune villitis or histiocytic intervillositis. Covid-19 infection is associated with similar pattern of histological changes of the placenta leading to placental insufficiency and fetal death. This case report supports maternal- fetal vertical transmission of SARS-CoV-2 virus leading to placental insufficiency and fetal demise. MPFD and CHI appear to be the typical placental histology for SARS-CoV-2 virus infection associated fetal demise.


Assuntos
COVID-19/virologia , Vilosidades Coriônicas/virologia , Fibrina/metabolismo , Complicações Infecciosas na Gravidez/virologia , SARS-CoV-2/patogenicidade , Adulto , Vilosidades Coriônicas/patologia , Feminino , Morte Fetal/etiologia , Histiócitos/virologia , Humanos , Placenta/patologia , Placenta/virologia , Gravidez , Complicações Infecciosas na Gravidez/patologia , RNA Viral
19.
J Stroke Cerebrovasc Dis ; 30(8): 105755, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34116489

RESUMO

OBJECTIVE: The aim of the present study was to determine whether there is a correlation between thrombotic pathology and prognosis of endovascular treatment (EVT) for acute ischemic stroke (AIS). METHODS: Thrombi were taken from 58 patients with cerebral ischemic thrombosis who were consecutively selected for EVT for AIS. The collected thrombi then underwent hematoxylin-eosin staining for pathological examinations to determine the red blood cell (RBC) ratio and fibrin/platelet components. The patients were divided into the following three groups according to their proportions of RBCs in thrombi: RBC-rich group (RBC ratio ≥ 70%), mixed group (RBC ratio at 31-69%), and fibrin/platelet-rich group (RBC ratio ≤ 30%). Prognosis was classified into good (0-2 points on modified Rankin scale [mRS] at postoperative 90 days) and poor (3-6 points on mRS at postoperative 90 days). Correlational analysis was performed between thrombotic pathology and prognosis of EVT for AIS. RESULTS: Among all patients, the distributions were as follows: 18.96% (11/58) patients in the RBC-rich group, 63.79% (37/58) patients in the mixed group, and 17.24% (10/58) patients in the fibrin/platelet-rich group. In addition, 43.10% (25/58) of the patients had good prognosis and 56.90% (33/58) had poor prognosis.There was no statistically significant difference between the good prognosis and the poor prognosis in the RBC-rich group, the mixed group, and the fibrin/platelet-rich group (P=0.713, 0.829, 0.748).Multivariate logistic regression analysis to explored the association between RBC-rich group and good prognosis while adjusting for other baseline prognostic factors (age, ASPECTS, NIHSS score, and PRT and intravenous alteplase-bridging therapy). Compared to the fibrin/platelet-rich group, the odds ratio(OR) of achieving good prognosis was 0.60 (P = 0.592) for the mixed group and OR = 0.74 (P = 0.793) for the RBC-rich group.Notably, age was found to be negatively associated with good prognosis (OR = 0.91, P = 0.013). The ASPECTS score was found to be positively associated with good prognosis (OR = 2.01, P = 0.002). Alteplase bridging was associated with a marginally significant positive association with good prognosis (OR = 4.23, P = 0.083). CONCLUSIONS: No correlation was found between thrombotic pathology and prognosis of EVT for AIS. Good prognosis after endovascular treatment was associated with low age, high ASPECTS at admission, and alteplase bridging.


Assuntos
Plaquetas/patologia , Procedimentos Endovasculares , Eritrócitos/patologia , Trombose Intracraniana/terapia , AVC Isquêmico/terapia , Fatores Etários , Idoso , Plaquetas/química , Avaliação da Deficiência , Procedimentos Endovasculares/efeitos adversos , Procedimentos Endovasculares/instrumentação , Feminino , Fibrina/análise , Fibrinolíticos/uso terapêutico , Humanos , Trombose Intracraniana/sangue , Trombose Intracraniana/patologia , AVC Isquêmico/sangue , AVC Isquêmico/patologia , Masculino , Pessoa de Meia-Idade , Medição de Risco , Fatores de Risco , Stents , Terapia Trombolítica , Ativador de Plasminogênio Tecidual/uso terapêutico , Resultado do Tratamento
20.
Pol Arch Intern Med ; 131(7-8): 666-672, 2021 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-34100569

RESUMO

INTRODUCTION: Patients with granulomatosis with polyangiitis (GPA) show increased tendency toward thromboembolic phenomena in exacerbation of their disease. OBJECTIVES: The aim of the study was to evaluate thrombin generation potential and fibrinolytic plasma activity in patients with GPA, both in exacerbation and in remission. PATIENTS AND METHODS: We included 38 patients with GPA: 18 with exacerbated GPA and 20 in remission. The control group included 39 healthy participants matched for age and sex. Plasma thrombogenic potential was assessed using calibrated automated thrombography. Plasma fibrinolytic potential was assessed using clot lysis time (CLT). We also measured levels of inflammatory markers, thrombomodulin, and fibrinolysis proteins in all participants. RESULTS: In the whole group of patients with GPA, endogenous thrombin potential was higher by about 25% (P <0.001), while CLT was lower by about 20% (P = 0.02) when compared with controls. The endogenous thrombin potential was higher, the CLT lower, and the levels of thrombomodulin and inflammation markers (C­reactive protein, fibrinogen, factor VIII) higher both in patients with exacerbation and in remission than in the control group; no such differences were noted when comparing those with exacerbation and those in remission, however. The only parameter that differentiated patients with GPA exacerbation from those in remission was the D­dimer level (median [interquartile range], 1151 [597.2-2468.7] ng/ml vs 340.4 [255.1-500.7] ng/ml; P <0.001), a marker of lysis of intravascularly formed fibrin. CONCLUSIONS: Patients with GPA show an increased prothrombotic state, regardless of the disease phase. This is probably related to ongoing low-grade inflammation and endothelial injury. Large clinical studies are required to address the need for, and appropriate type of, antithrombotic prophylaxis during the course of GPA.


Assuntos
Granulomatose com Poliangiite , Fibrina , Tempo de Lise do Coágulo de Fibrina , Fibrinólise , Granulomatose com Poliangiite/complicações , Humanos , Trombina
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