Thrombin-Activatable Fibrinolysis Inhibitor (TAFI) Regulates Activated Complement C5a in Vivo.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2973-2973
Author(s):  
Toshihiko Nishimura ◽  
Timothy Myles ◽  
Mariko Nagashima ◽  
John Morser ◽  
Ronald G. Pearl ◽  
...  
Keyword(s):  
Low Dose ◽  
Inflammatory Responses ◽  
Pulmonary Inflammation ◽  
Saline Control ◽  
High Dose ◽  
Thrombin Activatable Fibrinolysis Inhibitor ◽  
Cell Counts ◽  
Fibrinolysis Inhibitor ◽  
Anti Inflammatory ◽  
Deficient Mice

Abstract The latent plasma carboxypeptidase thrombin-activatable fibrinolysis inhibitor (TAFI) is activated by thrombin/thrombomodulin on the endothelial cell surface, and functions in dampening fibrinolysis. We recently showed that TAFIa efficiently inactivates bradykinin, C5a, and thrombin-cleaved osteopontin, suggesting that it may have a broad anti-inflammatory role. In this study, we examined the pulmonary inflammatory responses in a C5a-induced alveolitis model in wild-type (WT) and TAFI deficient mice. TAFI deficient mice had been backcrossed into a pure C57/B6 background. C5a was instilled into the trachea of anesthetized mice, which were then allowed to recover. Six hours later, bronchiole alveolar lavage (BAL) was performed and the extent of pulmonary inflammation determined. C5a caused a dose-dependent increase in cell counts (WBC/ul) in the BAL fluids in WT mice (saline control, 5.3±4.1 (mean±SD); C5a at 0.05 mg/ml, 28.7±8.5, C5a at 0.1 mg/ml, 60.4±23.8; n=6 for saline and low-dose C5a, n=12 for high dose C5a; p < 0.0005 and < 0.0001 for low-dose and high-dose C5a compared to saline control respectively). Significantly increased BAL cell counts in the TAFI deficient mice in response to C5a stimulation were noted (saline control, 3.3±3.9; low-dose C5a 70±11.2; high-dose C5a 159.6±58.7; n=6 for saline and low-dose C5a, n=12 for high dose C5a; p < 0.00001 and < 5E-0.6 respectively), representing ~2.4–2.6 fold increase in BAL cell counts in the TAFI deficient mice. Determinations of the wet/dried lung weight ratios, an index of pulmonary edema, showed similarly significant results (WT mice: 4.1±0.14, 4.25±0.1, 4.52±0.15 and TAFI-deficient mice: 4.27±0.41, 5.3±0.53, 5.77±0.62 for saline control, low- and high-dose C5a respectively). Our data indicate that in the absence of TAFI, the chemotactic and inflammatory effects of C5a in pulmonary inflammation were enhanced. We are in the process of testing whether E229K thrombin, which selectively activates protein C and TAFI, will ameliorate the C5a alveolitis response in WT mice. Our results support our thesis that the function of TAFIa is not restricted to fibrinolysis. It has a broad anti-inflammatory role and may serve as a homeostatic counterbalance to thrombin’s inflammatory functions.

scholarly journals Thrombin-activatable procarboxypeptidase B regulates activated complement C5a in vivo

Blood ◽  
2006 ◽  
Vol 109 (5) ◽  
pp. 1992-1997 ◽  
Author(s):  
Toshihiko Nishimura ◽  
Timothy Myles ◽  
Adrian M. Piliposky ◽  
Peter N. Kao ◽  
Gerald J. Berry ◽  
...  
Keyword(s):  
Beneficial Effect ◽  
Inflammatory Mediators ◽  
Protein C ◽  
Inflammatory Responses ◽  
Cell Counts ◽  
Fibrinolysis Inhibitor ◽  
Complement C5a ◽  
Homeostatic Response ◽  
Deficient Mice

Abstract Plasma procarboxypeptidase B (proCPB) is activated by the endothelial thrombin-prothrombomodulin complex. Activated (CPB) functions as a fibrinolysis inhibitor, but it may play a broader role by inactivating inflammatory mediators. To test this hypothesis, C5a-induced alveolitis was studied in wild-type (WT) and proCPB-deficient mice (proCPB−/−). C5a-induced alveolitis, as measured by cell counts and total protein contents in bronchoalveolar lavage fluids, was markedly enhanced in the proCPB−/− mice. E229K thrombin, a thrombin mutant with minimal clotting activity but retaining its ability to activate protein C and proCPB, attenuated C5a-induced alveolitis in WT but not in proCPB−/− mice, indicating that its beneficial effect is mediated primarily by its activation of proCPB. Lung tissue histology confirmed these cellular inflammatory responses. Delayed administration of E229K thrombin after the C5a instillation was ineffective in reducing alveolitis in WT mice, suggesting that the beneficial effect of E229K thrombin is due to the direct inhibition of C5a by CPB. Our studies show that thrombin-activatable proCPB, in addition to its role in fibrinolysis, has intrinsic anti-inflammatory functions. Its activation, along with protein C, by the endothelial thrombin-TM complex represents a homeostatic response to counteract the inflammatory mediators generated at the site of vascular injury.

Abstract WP282: DS-1040 an Inhibitor of the Activated Thrombin Activatable Fibrinolysis Inhibitor Improves Behavior in Embolized Rabbits

Stroke ◽  
2016 ◽  
Vol 47 (suppl_1) ◽  
Author(s):  
Paul A Lapchak ◽  
Paul D Boitano ◽  
Kengo Noguchi
Keyword(s):  
Plasma Concentrations ◽  
Inhibitory Effect ◽  
Neurological Deficits ◽  
Saline Control ◽  
Control Group ◽  
High Dose ◽  
Low Doses ◽  
Thrombin Activatable Fibrinolysis Inhibitor ◽  
Treatment Groups ◽  
Fibrinolysis Inhibitor

Introduction: DS-1040 is an inhibitor of the activated thrombin-activatable fibrinolysis inhibitor (TAFIa) being developed for the treatment of acute ischemic stroke (AIS). We evaluated the effect of DS-1040 infusiontreatment on rabbit TAFIa activity; plasma concentrations and we also determined if DS-1040 treatment affected behavioral outcome following embolization using the standard rabbit small clot embolic stroke model (RSCEM). Safety was assessed by measuring the effect of DS-1040 on intracerebral hemorrhage (ICH) and mortality. Methods: The inhibitory effect of DS-1040 on TAFIa was evaluated using hippuryl-Arg, a substrate of carboxypeptidase. The study was conducted randomized and blinded per RIGOR guidelines. Male New Zealand white rabbits (2-2.5kg) were embolized by injecting a suspension of small blood clots into the brain via an indwelling carotid catheter. Saline (control), tPA (3.3 mg/kg, i.v., 10% bolus/90% infusion for 0.5 h), high and low doses of DS-1040 (approx. 940 mg/per rabbit and 470 mg/per rabbit, respectively) were given 1 h following embolization as an i.v. bolus followed by a s.c. infusion for 47 h. Behavioral analysis was conducted 48 h following embolization, resulting in the calculation of an effective stroke dose (P50) or clot amount (mg) that produces neurological deficits in 50% of the rabbits. ICH was also assessed in all treatment groups. Results: DS-1040 inhibited rabbit TAFIa with an IC50 value of 11.7 ng/mL. Using the dosing regimen described above, the mean plasma concentration of DS-1040 at 48 h was 1838 ng/mL (high dose) and 591 ng/mL (low dose). Rabbits treated with high and low doses of DS-1040 had P50 values of 1.61 ± 0.23 mg (p<0.05) and 0.96 ± 0.32 mg, respectively; only the high dose significantly increased (p<0.05) the group P50 by 206% compared to control group P50 (0.78 ± 0.39 mg). The tPA group had a P50 value of 2.29 ± 0.46 mg (p<0.05). Neither the hemorrhage rate nor mortality rate were significantly affected by DS-1040 treatment. Conclusions: DS-1040 significantly improved behavioral function in small clot embolized rabbits, suggesting that DS-1040 should be further pursued as a treatment for stroke victims.

The neonatal Fc receptor (FcRn) is not required for IVIg or anti-CD44 monoclonal antibody–mediated amelioration of murine immune thrombocytopenia

Blood ◽  
2011 ◽  
Vol 118 (24) ◽  
pp. 6403-6406 ◽  
Author(s):  
Andrew R. Crow ◽  
Sara J. Suppa ◽  
Xi Chen ◽  
Patrick J. Mott ◽  
Alan H. Lazarus
Keyword(s):  
Immune Thrombocytopenia ◽  
Low Dose ◽  
Fc Receptor ◽  
High Dose ◽  
Neonatal Fc Receptor ◽  
Antiplatelet Antibody ◽  
Antibody Treatment ◽  
Acute Itp ◽  
Deficient Mice ◽  
Cd44 Antibody

Abstract To definitively determine whether the neonatal Fc receptor (FcRn) is required for the acute amelioration of immune thrombocytopenia (ITP) by IVIg, we used FcRn-deficient mice in a murine ITP model. Mice injected with antiplatelet antibody in the presence or absence of IVIg displayed no difference in platelet-associated IgG between FcRn deficient versus C57BL/6 mice. FcRn-deficient mice treated with high-dose (2 g/kg) IVIg or a low–dose (2 mg/kg) of an IVIg-mimetic CD44 antibody were, however, protected from thrombocytopenia to an equivalent extent as wild-type mice. To verify and substantiate the results found with FcRn-deficient mice, we used β2-microglobulin–deficient mice (which do not express functional FcRn) and found that IVIg or CD44 antibody also protected them from thrombocytopenia. These data suggest that for both high-dose IVIg as well as low-dose CD44 antibody treatment in an acute ITP model, FcRn expression is neither necessary nor required.

scholarly journals CD8+ T Cells Mediate Recovery andImmunopathology in West Nile VirusEncephalitis

2003 ◽  
Vol 77 (24) ◽  
pp. 13323-13334 ◽  
Author(s):  
Yang Wang ◽  
Mario Lobigs ◽  
Eva Lee ◽  
Arno Müllbacher
Keyword(s):  
T Cells ◽  
Dose Group ◽  
Low Dose ◽  
Neuronal Degeneration ◽  
High Dose ◽  
West Nile ◽  
Activation Markers ◽  
High Doses ◽  
The Brain ◽  
Deficient Mice

ABSTRACT C57BL/6J mice infected intravenously with the Sarafend strain of West Nile virus (WNV) develop a characteristic central nervous system (CNS) disease, including an acute inflammatory reaction. Dose response studies indicate two distinct kinetics of mortality. At high doses of infection (108 PFU), direct infection of the brain occurred within 24 h, resulting in 100% mortality with a 6-day mean survival time (MST), and there was minimal destruction of neural tissue. A low dose (103 PFU) of infection resulted in 27% mortality (MST, 11 days), and virus could be detected in the CNS 7 days postinfection (p.i.). Virus was present in the hypogastric lymph nodes and spleens at days 4 to 7 p.i. Histology of the brains revealed neuronal degeneration and inflammation within leptomeninges and brain parenchyma. Inflammatory cell infiltration was detectable in brains from day 4 p.i. onward in the high-dose group and from day 7 p.i. in the low-dose group, with the severity of infiltration increasing over time. The cellular infiltrates in brain consisted predominantly of CD8+, but not CD4+, T cells. CD8+ T cells in the brain and the spleen expressed the activation markers CD69 early and expressed CD25 at later time points. CD8+ T-cell-deficient mice infected with 103 PFU of WNV showed increased mortalities but prolonged MST and early infection of the CNS compared to wild-type mice. Using high doses of virus in CD8-deficient mice leads to increased survival. These results provide evidence that CD8+ T cells are involved in both recovery and immunopathology in WNV infection.

scholarly journals Toll-Like Receptor Signaling in Airborne Burkholderia thailandensis Infection

2009 ◽  
Vol 77 (12) ◽  
pp. 5612-5622 ◽  
Author(s):  
T. Eoin West ◽  
Thomas R. Hawn ◽  
Shawn J. Skerrett
Keyword(s):  
Low Dose ◽  
Inflammatory Responses ◽  
High Dose ◽  
Tlr Signaling ◽  
Necrosis Factor Alpha ◽  
Airborne Infection ◽  
Essential Components ◽  
High Doses

ABSTRACT Melioidosis is a tropical disease endemic in southeast Asia and northern Australia caused by the gram-negative soil saprophyte Burkholderia pseudomallei. Although infection is often systemic, the lung is frequently involved. B. thailandensis is a closely related organism that at high doses causes lethal pneumonia in mice. We examined the role of Toll-like receptors (TLRs), essential components of innate immunity, in vitro and in vivo during murine B. thailandensis pneumonia. TLR2, TLR4, and TLR5 mediate NF-κB activation by B. thailandensis in transfected HEK293 or CHO cells. In macrophages, TLR4 and the adaptor molecule MyD88, but not TLR2 or TLR5, are required for tumor necrosis factor alpha production induced by B. thailandensis. In low-dose airborne infection, TLR4 is needed for early, but not late, bacterial containment, and MyD88 is essential for control of infection and host survival. TLR2 and TLR5 are not necessary to contain low-dose infection. In high-dose airborne infection, TLR2 deficiency confers a slight survival advantage. Lung and systemic inflammatory responses are induced by low-dose inhaled B. thailandensis independently of individual TLRs or MyD88. These findings suggest that redundancy in TLR signaling or other MyD88-dependent pathways may be important in pneumonic B. thailandensis infection but that MyD88-independent mechanisms of inflammation are also activated. TLR signaling in B. thailandensis infection is substantially comparable to signaling induced by virulent B. pseudomallei. These studies provide additional insights into the host-pathogen interaction in pneumonic Burkholderia infection.

scholarly journals Dose-dependent roles of aspirin and other non-steroidal anti-inflammatory drugs in abnormal bone remodeling and skeletal regeneration

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Yong Xie ◽  
Meng Pan ◽  
Yanpan Gao ◽  
Licheng Zhang ◽  
Wei Ge ◽  
...  
Keyword(s):  
Bone Formation ◽  
Bone Remodeling ◽  
Low Dose ◽  
High Dose ◽  
Dose Aspirin ◽  
Low Dose Aspirin ◽  
Inflammatory Drugs ◽  
Anti Inflammatory ◽  
Dose Dependent ◽  
High Dose Aspirin

AbstractThe failure of remodeling process that constantly regenerates effete, aged bone is highly associated with bone nonunion and degenerative bone diseases. Numerous studies have demonstrated that aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs) activate cytokines and mediators on osteoclasts, osteoblasts and their constituent progenitor cells located around the remodeling area. These cells contribute to a complex metabolic scenario, resulting in degradative or synthetic functions for bone mineral tissues. The spatiotemporal effects of aspirin and NSAIDs in the bone remodeling are controversial according the specific therapeutic doses used for different clinical conditions. Herein, we review in vitro, in vivo, and clinical studies on the dose-dependent roles of aspirin and NSAIDs in bone remodeling. Our results show that low-dose aspirin (< 100 μg/mL), which is widely recommended for prevention of thrombosis, is very likely to be benefit for maintaining bone mass and qualities by activation of osteoblastic bone formation and inhibition of osteoclast activities via cyclooxygenase-independent manner. While, the roles of high-dose aspirin (150–300 μg/mL) and other NSAIDs in bone self-regeneration and fracture-healing process are difficult to elucidate owing to their dual effects on osteoclast activity and bone formation of osteoblast. In conclusion, this study highlighted the potential clinical applications of low-dose aspirin in abnormal bone remodeling as well as the risks of high-dose aspirin and other NSAIDs for relieving pain and anti-inflammation in fractures and orthopedic operations.

scholarly journals The Endogenous Estradiol Metabolite 2-Methoxyestradiol Reduces Atherosclerotic Lesion Formation in Female Apolipoprotein E-Deficient Mice

Endocrinology ◽  
2007 ◽  
Vol 148 (9) ◽  
pp. 4128-4132 ◽  
Author(s):  
Johan Bourghardt ◽  
Göran Bergström ◽  
Alexandra Krettek ◽  
Sara Sjöberg ◽  
Jan Borén ◽  
...  
Keyword(s):  
Apolipoprotein E ◽  
Low Dose ◽  
Hormone Replacement ◽  
Atherosclerotic Lesion ◽  
Total Serum ◽  
High Dose ◽  
Lesion Formation ◽  
Cholesterol Levels ◽  
Deficient Mice ◽  
Atherosclerotic Lesion Formation

Estradiol, the major endogenous estrogen, reduces experimental atherosclerosis and metabolizes to 2-methoxyestradiol in vascular cells. Currently undergoing evaluation in clinical cancer trials, 2-methoxyestradiol potently inhibits cell proliferation independently of the classical estrogen receptors. This study examined whether 2-methoxyestradiol affects atherosclerosis development in female mice. Apolipoprotein E-deficient mice, a well-established mouse model of atherosclerosis, were ovariectomized and treated through slow-release pellets with placebo, 17β-estradiol (6 μg/d), or 2-methoxyestradiol [6.66 μg/d (low-dose) or 66.6 μg/d (high-dose)]. After 90 d, body weight gain decreased and uterine weight increased in the high-dose but not low-dose 2-methoxyestradiol group. En face analysis showed that the fractional area of the aorta covered by atherosclerotic lesions decreased in the high-dose 2-methoxyestradiol (52%) but not in the low-dose 2-methoxyestradiol group. Total serum cholesterol levels decreased in the high- and low-dose 2-methoxyestradiol groups (19%, P &lt; 0.05 and 21%, P = 0.062, respectively). Estradiol treatment reduced the fractional atherosclerotic lesion area (85%) and decreased cholesterol levels (42%). In conclusion, our study shows for the first time that 2-methoxyestradiol reduces atherosclerotic lesion formation in vivo. The antiatherogenic activity of an estradiol metabolite lacking estrogen receptor activating capacity may argue that trials on cardiovascular effects of hormone replacement therapy should use estradiol rather than other estrogens. Future research should define the role of 2-methoxyestradiol as a mediator of the antiatherosclerotic actions of estradiol. Furthermore, evaluation of the effects of 2-methoxyestradiol on cardiovascular disease endpoints in ongoing clinical trials is of great interest.

Comparative study of high-dose Xuezhikang and low-dose Xuezhikang plus ezetimibe on pro- and anti-inflammatory markers

2013 ◽  
Vol 8 (1) ◽  
pp. 151-157 ◽  
Author(s):  
Jun Liu ◽  
Song-Hui Luo ◽  
Ping Qing ◽  
Long-Hui Di ◽  
Xiang-Dong You ◽  
...  
Keyword(s):  
Comparative Study ◽  
Inflammatory Markers ◽  
Low Dose ◽  
High Dose ◽  
Anti Inflammatory

scholarly journals Possible Divergent Local and Peripheral Immunological Effects of Low-Dose Mesencure, an Enhanced Mesenchymal Cell Therapy, May Contribute to Its Success in Treating Severe Covid-19 Patients

Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 2172-2172
Author(s):  
Tomer Bronshtein ◽  
Dror Ben David ◽  
Atara Novak ◽  
Vered Kivity ◽  
Shai Meretzki ◽  
...  
Keyword(s):  
T Cells ◽  
Low Dose ◽  
De Novo ◽  
Lung Edema ◽  
High Dose ◽  
Local Inflammation ◽  
Blood Leukocytes ◽  
Gene Encoding ◽  
Leukocyte Counts ◽  
Anti Inflammatory

Abstract Mesenchymal stromal cells (MSC) are widely investigated for treating ARDS in Covid-19. Nonetheless, these efforts are overshadowed by studies predating the pandemic that mostly failed to show MSC efficacy in ARDS and recent disappointments with repurposed MSC products. Relying on years of MSC-related experience, Bonus BioGroup developed MesenCure: An enhanced allogeneic MSC therapy for Covid-19, professionalized by a unique combination of culture conditions and optimized in ARDS-relevant models. MesenCure is currently evaluated in a Phase II study in severe Covid-19 patients and administered (IV) in three doses (1.5M cells/kg, d1, d3, d5). A Phase I/II study on ten severe patients demonstrated a significant improvement in ARDS-related parameters following MesenCure treatment. Patients were discharged within one day (median) following treatment, requiring no respiratory support. Speedy recovery from local inflammation was observed in these patients, demonstrated by a rapid reduction in diffuse lung pneumonia, from 55% of the lung area to 15% within 5-6 days from the first dose (p&lt;0.01, Fig. A-C). A corresponding drop in CRP was detected (p&lt;0.01), which returned to normal. A multivariate regression analysis revealed that the reduction in CRP was mainly associated with the number of doses administered and not and the time elapsed since the first dose. MesenCure efficacy may be attributable to the cells' de novo expression of the gene encoding for the IL-6 receptor, making them more responsive to inflammation than non-professionalized naïve MSC (NA-MSC); as well as &gt;8-fold upregulation of the EDIL3 gene, encoding for an endogenous inhibitor of immune infiltration. A corresponding immunosuppressive effect of MesenCure MSCs was demonstrated in vitro, showing their ability to suppress T cells activation twice more effectively than NA-MSC. In this study, MesenCure inhibited the proliferation of primary CD4 T cells in a concentration-depended manner following non-specific activation. Over 98% inhibition was achieved in co-culture of 1:10 MSC-to-PBMC with an IC 50 of 6k MSC/200k PBMCs (r 2=1.00) compared to 12k NA-MSC/200k PBMCs (r 2=0.95). Comparable results were also obtained for CD8 T cells. Similarly, MesenCure inhibited ROS production by primary neutrophils remarkably fast and by up to 80% within less than 40 minutes following their activation (IC 50 = 19k MSC/200k neutrophils, r 2=1.00). In addition to local immunosuppressive outcomes, a significant increase in blood leukocytes was observed in patients treated with MesenCure (p&lt;0.05, Fig. D-F). Further analysis suggested that the increase in total WBCs and neutrophils was associated with the number of MesenCure doses administered (p&lt;0.05, Fig. G-H). In contrast, the increase in lymphocytes was time-dependent (R=0.72, Fig. I). The seemingly exclusively localized anti-inflammatory effects seen in severe patients treated with MesenCure were also observed in animal (murine) studies. An in vivo study in an acute lung injury model demonstrated a dose-dependent localized reduction in leukocyte counts in the lung fluids of animals treated with MesenCure (IV) using two dose levels. Relative to untreated animals, MesenCure reduced lung leukocyte counts by 35%-43% in animals treated with the low dose and by 62%-67% following high-dose MesenCure treatment (p&lt;0.05). The leukocytes' clearance from the lungs was accompanied by a 41%-57% reduction in lung edema (p&lt;0.05) following MesenCure treatment. Notably, NA-MSC did not achieve the same effect. Similar to our clinical findings, a significant increase was measured in neutrophil counts in animals treated with low-dose MesenCure (p&lt;0.05), which decreased dramatically (p&lt; 0.01) in animals treated with a four-times higher dose. MesenCure is administered at a much lower dose compared to other MSC products administered at up to 10M cells/kg. Considering the increase in blood leukocytes measured in patients treated with low-dose MesenCure and comparable preclinical findings, our data suggest that low-dose MesenCure could elicit a potent local anti-inflammatory effect without suppressing, and even enhancing, peripheral immunity that is needed to fight the virus. Further research is inevitably required into the mechanism behind this phenomenon. However, our results indicate MesenCure's potential in relieving local inflammation while giving the patient a fighting chance against viremia. Figure 1 Figure 1. Disclosures Bronshtein: Bonus BioGroup: Current Employment. Ben David: Bonus BioGroup: Current Employment. Novak: Bonus BioGroup: Current Employment. Kivity: Bonus BioGroup: Current Employment. Meretzki: Bonus BioGroup: Current Employment. Rozen: Bonus BioGroup: Consultancy.

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