Suppression of fibrin(ogen)-driven pathologies through controlled knockdown by lipid nanoparticle delivery of siRNA

Blood ◽  
2021 ◽  
Author(s):  
Lih Jiin Juang ◽  
Woosuk Steve Hur ◽  
Lakmali Munasinghage Silva ◽  
Amy W Strilchuk ◽  
Brenton Francisco ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Metastatic Potential ◽  
Plasma Fibrinogen ◽  
Fibrinogen Concentration ◽  
Nanoparticle Delivery ◽  
Lipid Nanoparticle ◽  
Cytokine Levels ◽  
Endotoxemia Model ◽  
Peritonitis Model ◽  
Deficient Mice

Fibrinogen plays a pathologic role in multiple diseases. It contributes to thrombosis and modifies inflammatory and immune responses, supported by studies in mice expressing fibrinogen variants with altered function or with a germline fibrinogen deficiency. However, therapeutic strategies to safely and effectively tailor plasma fibrinogen concentration are lacking. Here, we developed a strategy to tune fibrinogen expression by administering lipid nanoparticle (LNP)-encapsulated siRNA targeting the fibrinogen α chain (siFga). Three distinct LNP-siFga reagents reduced both hepatic Fga mRNA and fibrinogen levels in platelets and plasma, with plasma levels decreased to 42%, 16% and 4% of normal within one-week of administration. Using the most potent siFga, circulating fibrinogen was controllably decreased to 32%, 14%, and 5% of baseline with a 0.5, 1, and 2 mg/kg dose, respectively. Whole blood from mice treated with siFga formed clots with significantly decreased clot strength ex vivo, but siFga treatment did not compromise hemostasis following saphenous vein puncture or tail transection. In an endotoxemia model, siFga suppressed the acute phase response and decreased plasma fibrinogen, D-dimer, and proinflammatory cytokine levels. In a sterile peritonitis model, siFga restored normal macrophage migration in plasminogen-deficient mice. Finally, treatment of mice with siFga decreased the metastatic potential of tumour cells in a manner comparable to that observed in fibrinogen-deficient mice. The results indicate that siFga causes robust and controllable depletion of fibrinogen and provide the proof-of-concept that this strategy can modulate the pleiotropic effects of fibrinogen in relevant disease models.

Some Factors Influencing Metabolism and Distribution of Fibrinogen in Man and Rabbits

1977 ◽  
Vol 37 (02) ◽  
pp. 243-252
Author(s):  
Yi-Hsiang Chen ◽  
E. B Reeve
Keyword(s):  
Plasma Fibrinogen ◽  
Synthesis Rate ◽  
Fibrinogen Concentration ◽  
Fractional Catabolic Rate ◽  
System Parameters ◽  
Catabolic Rate ◽  
Simultaneous Decrease ◽  
Normal Males ◽  
Healthy Females ◽  
Made In

SummaryTo shed some light on the homeostatic regulation of plasma fibrinogen, metabolic studies were made in healthy females, and in normal, thyroidectomized, and thyroxine-treated rabbits. In females, compared with normal males, plasma fibrinogen concentration, plasma and interstitial fibrinogen decreased consequent to an increased fractional catabolic rate and a normal fibrinogen synthesis rate. The interstitial/plasma fibrinogen ratio remained unchanged. In normal rabbits, with increasing body weight fractional catabolic rate and catabolic rate decreased, while fibrinogen concentration and plasma fibrinogen remained constant owing to a simultaneous decrease in fibrinogen synthesis. In addition, fractional transcapillary transfer rate and transcapillary flux also decreased resulting in a shrinkage of interstitial fibrinogen. Thyroidectomy and thyroxine-injection markedly altered fibrinogen metabolism: thyroid hormone accelerated fibrinogen catabolism but also stimulated synthesis. The net result was an increase in plasma fibrinogen and fibrinogen concentration. The interstitial/plasma fibrinogen ratio decreased in thyroxine-treated, and increased in thyroidectomized animals. This study defines the variations of the fibrinogen system parameters in these physiologic and pathologic conditions, and illustrates some patterns of alterations in fibrinogen metabolism.

Factors Regulating Plasma Protein Synthesis IV. Influence of Fragments D and E on Plasma Fibrinogen Concentration

1974 ◽  
Vol 31 (03) ◽  
pp. 395-402 ◽  
Author(s):  
V Bocci ◽  
T Conti ◽  
M Muscettola ◽  
A Pacini ◽  
G. P Pessina
Keyword(s):  
Protein Synthesis ◽  
Plasma Protein ◽  
Plasma Fibrinogen ◽  
Fibrinogen Concentration

SummaryRabbit fibrinogen digest products (FDP) have been separated by Pevikon block electrophoresis yielding fragments D, E and other unidentified FDP.The fragments were injected into rabbits. Surprisingly, as little as 4.3 mg of fragment D elicited a significant increase in plasma fibrinogen concentration 24 hr after injection. The stimulating activity of fragment D is at least 10-fold higher than that of fragment E.

scholarly journals Decellularised Human Umbilical Artery as a Vascular Graft Elicits Minimal Pro-Inflammatory Host Response Ex Vivo and In Vivo

2021 ◽  
Vol 22 (15) ◽  
pp. 7981
Author(s):  
Alexander Høgsted Ahlmann ◽  
Shu Fang ◽  
Sussi Bagge Mortensen ◽  
Line Weis Andersen ◽  
Pernille Gejl Pedersen ◽  
...  
Keyword(s):  
Vascular Graft ◽  
Umbilical Artery ◽  
Ex Vivo ◽  
White Blood Cells ◽  
Small Diameter ◽  
M1 Macrophages ◽  
Host Immune Responses ◽  
Cytokine Levels ◽  
Anti Inflammatory Cytokine

Small diameter (<6 mm) vessel grafts still pose a challenge for scientists worldwide. Decellularised umbilical artery (dUA) remains promising as small diameter tissue engineered vascular graft (TEVG), yet their immunogenicity remains unknown. Herein, we evaluated the host immune responses, with a focus on the innate part, towards human dUA implantation in mice, and confirmed our findings in an ex vivo allogeneic human setup. Overall, we did not observe any differences in the number of circulating white blood cells nor the number of monocytes among three groups of mice (1) dUA patch; (2) Sham; and (3) Mock throughout the study (day −7 to 28). Likewise, we found no difference in systemic inflammatory and anti-inflammatory cytokine levels between groups. However, a massive local remodelling response with M2 macrophages were observed in the dUA at day 28, whereas M1 macrophages were less frequent. Moreover, human monocytes from allogeneic individuals were differentiated into macrophages and exposed to lyophilised dUA to maximize an eventual M1 response. Yet, dUA did not elicit any immediate M1 response as determined by the absence of CCR7 and CXCL10. Together this suggests that human dUA elicits a minimal pro-inflammatory response further supporting its use as a TEVG in an allogeneic setup.

scholarly journals Impaired platelet responses to thrombin and collagen in AKT-1–deficient mice

Blood ◽  
2004 ◽  
Vol 104 (6) ◽  
pp. 1703-1710 ◽  
Author(s):  
Juhua Chen ◽  
Sarmishtha De ◽  
Derek S. Damron ◽  
William S. Chen ◽  
Nissim Hay ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Ex Vivo ◽  
Dense Granules ◽  
Fibrinogen Binding ◽  
Downstream Effectors ◽  
Low Concentrations ◽  
Phosphoinositide 3 Kinase ◽  
Deficient Mice ◽  
Granule Release

Abstract We investigated the role of Akt-1, one of the major downstream effectors of phosphoinositide 3-kinase (PI3K), in platelet function using mice in which the gene for Akt-1 had been inactivated. Using ex vivo techniques, we showed that Akt-1-deficient mice exhibited impaired platelet aggregation and spreading in response to various agonists. These differences were most apparent in platelets activated with low concentrations of thrombin. Although Akt-1 is not the predominant Akt isoform in mouse platelets, its absence diminished the amount of total phospho-Akt and inhibited increases in intracellular Ca2+ concentration in response to thrombin. Moreover, thrombin-induced platelet α-granule release as well as release of adenosine triphosphate from dense granules was also defective in Akt-1-null platelets. Although the absence of Akt-1 did not influence expression of the major platelet receptors for thrombin and collagen, fibrinogen binding in response to these agonists was significantly reduced. As a consequence of impaired αIIbβ3 activation and platelet aggregation, Akt-1 null mice showed significantly longer bleeding times than wild-type mice. (Blood. 2004;104:1703-1710)

scholarly journals Upregulation of Cardiac IL-10 and Downregulation of IFN-γin Balb/c IL-4−/−in Acute Chagasic Myocarditis due to Colombian Strain ofTrypanosoma cruzi

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Marcos Vinicius da Silva ◽  
Vera Lúcia de Almeida ◽  
Wendyson Duarte de Oliveira ◽  
Natália Carasek Matos Cascudo ◽  
Pollyana Guimarães de Oliveira ◽  
...  
Keyword(s):  
Acute Phase ◽  
Experimental Infection ◽  
Cardiac Tissue ◽  
Histopathological Analysis ◽  
Nest Density ◽  
Cytokine Levels ◽  
Splenic Cells ◽  
Immune System Regulation ◽  
Deficient Mice

Inflammatory response in Chagas disease is related to parasite and host factors. However, immune system regulation has not been fully elucidated. Thus, this study is aimed at evaluating IL-4 influence on acute phase ofTrypanosoma cruziexperimental infection through dosage of cytokine levels in cardiac homogenate of infected Balb/c WT and Balb/c IL-4−/−as well as its histopathological repercussions. For such purpose, mice were divided into two groups: an infected group with 100 forms of the Colombian strain and an uninfected group. After 21 days of infection, animals were euthanized and the blood, spleen, and heart were collected. The spleen was used to culture splenic cells in 48 h. Subsequently, cytokines TNF-α, IL-12p70, IL-10, IFN-γ, and IL-17 were measured in the blood, culture supernatant, and heart apex by ELISA. The base of the heart was used for histopathological analysis. From these analysis, infected Balb/c IL-4−/−mice showed milder inflammatory infiltrate compared to Balb/c WT, but without changes in nest density and collagen deposition. IL-4 absence culminated in lower cardiac tissue IFN-γproduction, although it did not affect TNF-αexpression in situ. It also decreased TNF-αsystemic production and increased IL-10, both systemically andin situ. In addition, IL-4 absence did not influence IL-17 expression. Splenocytes of IL-4-deficient mice produced higher amounts of IFN-γ, TNF-α, and IL-17 and lower amounts of IL-10. Thus, IL-4 absence in acute phase of experimental infection withT. cruziColombian strain reduces myocarditis due to lower IFN-γproduction and greater IL-10 productionin situand this pattern is not influenced by splenocyte general repertoire.

scholarly journals Overlapping Roles for Interleukin-36 Cytokines in Protective Host Defense against MurineLegionella pneumophilaPneumonia

2018 ◽  
Vol 87 (1) ◽  
Author(s):  
Yuta Nanjo ◽  
Michael W. Newstead ◽  
Tetsuji Aoyagi ◽  
Xianying Zeng ◽  
Kazuhisa Takahashi ◽  
...  
Keyword(s):  
Host Defense ◽  
Legionella Pneumophila ◽  
Ex Vivo ◽  
Inflammatory Cells ◽  
Bacterial Clearance ◽  
Content Type ◽  
Cytokines And Chemokines ◽  
M1 Polarization ◽  
Life Threatening ◽  
Deficient Mice

ABSTRACTLegionella pneumophilacauses life-threatening pneumonia culminating in acute lung injury. Innate and adaptive cytokines play an important role in host defense againstL. pneumophilainfection. Interleukin-36 (IL-36) cytokines are recently described members of the larger IL-1 cytokine family known to exert potent inflammatory effects. In this study, we elucidated the role for IL-36 cytokines in experimental pneumonia caused byL. pneumophila. Intratracheal (i.t.) administration ofL. pneumophilainduced the upregulation of both IL-36α and IL-36γ mRNA and protein production in the lung. Compared to the findings forL. pneumophila-infected wild-type (WT) mice, the i.t. administration ofL. pneumophilato IL-36 receptor-deficient (IL-36R−/−) mice resulted in increased mortality, a delay in lung bacterial clearance, increasedL. pneumophiladissemination to extrapulmonary organs, and impaired glucose homeostasis. Impaired lung bacterial clearance in IL-36R−/−mice was associated with a significantly reduced accumulation of inflammatory cells and the decreased production of proinflammatory cytokines and chemokines.Ex vivo, reduced expression of costimulatory molecules and impaired M1 polarization were observed in alveolar macrophages isolated from infected IL-36R−/−mice compared to macrophages from WT mice. WhileL. pneumophila-induced mortality in IL-36α- or IL-36γ-deficient mice was not different from that in WT animals, antibody-mediated neutralization of IL-36γ in IL-36α−/−mice resulted in mortality similar to that observed in IL-36R−/−mice, indicating redundant and overlapping roles for these cytokines in experimental murineL. pneumophilapneumonia.

Effects of smoking and abstention from smoking on fibrinogen synthesis in humans

2001 ◽  
Vol 100 (4) ◽  
pp. 459-465 ◽  
Author(s):  
Kirsty A. HUNTER ◽  
Peter J. GARLICK ◽  
Iain BROOM ◽  
Susan E. ANDERSON ◽  
Margaret A. McNURLAN
Keyword(s):  
Risk Factors ◽  
Significant Risk ◽  
Plasma Fibrinogen ◽  
Fibrinogen Concentration ◽  
Primary Role ◽  
Absolute Rate ◽  
Albumin Synthesis ◽  
Plasma Albumin ◽  
Concomitant Reduction ◽  
Metabolic Mechanism

Cigarette smoking and hyperfibrinogenaemia are both significant risk factors for the development of cardiovascular disease. Two studies are described here which aimed to establish the metabolic mechanism responsible for the raised plasma fibrinogen concentration observed in smokers. Chronic smokers had a significantly elevated absolute rate of fibrinogen synthesis (ASR) compared with non-smokers (22.7±1.3 mg/kg per day versus 16.0±1.3 mg/kg per day; means±S.E.M., P < 0.01), with plasma levels of fibrinogen significantly correlated with fibrinogen synthesis (r = 0.65, P = 0.04). Unlike fibrinogen, plasma albumin concentrations were lower in smokers than in non-smokers (45±0.4 versus 47±0.7 g/l, P < 0.05), but there was no difference in rates of albumin synthesis between the two groups. Two weeks cessation from smoking by previously chronic smokers was associated with a rapid and marked fall in plasma fibrinogen concentration (from 3.06±0.11 g/l to 2.49±0.14 g/l, P < 0.001), and a significant reduction in ASR (a 33% reduction, from 24.1±1.7 to 16.1±1.0 mg/kg per day, P < 0.001). These studies suggest a primary role for increased synthesis in producing the hyperfibrinogenaemia associated with smoking. Moreover, abstention from smoking for a period of only 2 weeks induces a significant decrease in the rate of fibrinogen synthesis by the liver, with a concomitant reduction in the plasma fibrinogen concentration.

Fibrinogen measurement in cardiac surgery with cardiopulmonary bypass: Analysis of repeatability and agreement of Clauss method within and between six different laboratories

2014 ◽  
Vol 112 (07) ◽  
pp. 109-117 ◽  
Author(s):  
Ekaterina Baryshnikova ◽  
Armando Tripodi ◽  
Christoph J. Schlimp ◽  
Herbert Schöchl ◽  
Janne Cadamuro ◽  
...  
Keyword(s):  
Cardiac Surgery ◽  
Cardiopulmonary Bypass ◽  
Plasma Fibrinogen ◽  
Detection Methods ◽  
Fibrinogen Concentration ◽  
Plasma Samples ◽  
Limits Of Agreement ◽  
Study Objective ◽  
Standard Operating ◽  
Mean Differences

SummaryPlasma fibrinogen concentration is important for coagulopathy assessment, and is most commonly measured using the Clauss method. Several factors, including device type and reagent, have been shown to affect results. The study objective was to evaluate performance and repeatability of the Clauss method and to assess differences between measurements performed during and after cardiopulmonary bypass (CPB), by testing plasma samples from patients undergoing cardiac surgery with CPB. Samples were collected from 30 patients before surgery, approximately 20 minutes before weaning from CPB, and 5 minutes after CPB and protamine. Fibrinogen concentration was determined using the Clauss method at six quality-controlled specialised laboratories, according to accredited standard operating procedures. Regarding within-centre agreement for Clauss measurement, mean differences between duplicate measurements were between 0.00 g/l and 0.15 g/l, with intervals for 95% limits of agreement for mean Bland-Altman differences up to 1.3 g/l. Regarding between-centre agreement, some mean differences between pairs of centres were above 0.5 g/l. Differences of up to ∼2 g/l were observed with individual samples. Increased variability was observed between centres, with inter-class correlation values below 0.5 suggesting only fair agreement. There were no significant differences in fibrinogen concentration before weaning from CPB and after CPB for most centres and methods. In conclusion, considerable differences exist between Clauss-based plasma fibrinogen measured using different detection methods. Nevertheless, the similarity between measurements shortly before weaning from CPB and after CPB within centres suggests that on-pump measurements could provide an early estimation of fibrinogen deficit after CPB and thus guidance for haemostatic therapy.

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