-
Journal of Thrombosis and Haemostasis :... Feb 2010Protein S and tissue factor pathway inhibitor (TFPI) act together in down-regulating coagulation.
BACKGROUND
Protein S and tissue factor pathway inhibitor (TFPI) act together in down-regulating coagulation.
OBJECTIVE
To investigate the TFPI/protein S system in hereditary and acquired protein S deficiency.
METHODS
Plasma antigen levels of protein S and full-length TFPI were determined in heterozygous type I protein S-deficient individuals (n=35), patients on oral anticoagulant treatment (OAT) (n=29), oral contraceptive (OC) users (n=10) and matched controls. Thrombin generation was determined using calibrated automated thrombography.
RESULTS
Full-length TFPI levels were lower in type I protein S-deficient individuals (76.8+/-33.8%) than in age- and sex-matched controls (128.0+/-59.4%, P<0.001). Among protein S-deficient individuals with thrombosis, those on OAT had not only lower total protein S levels (25.7+/-8.2% vs. 54.7+/-8.2%, P<0.001), but also lower full-length TFPI levels (52.6+/-15.0% vs. 75.4+/-22.9%, P=0.009) than those not on OAT. Similarly, OC users had lower protein S (73.8+/-11.5% vs. 87.9+/-10.8%, P=0.005) and full-length TFPI levels (73.7+/-27.7% vs. 106.4+/-29.2%, P=0.007) than non-users. When triggered with tissue factor, plasma from protein S-deficient individuals generated 3-5-fold more thrombin than control plasma. The difference was only partially corrected by normalization of the protein S level, full correction requiring additional normalization of the TFPI level. Protein S-immunodepletion experiments indicated that free protein S and full-length TFPI form a complex in plasma, and the protein S/TFPI interaction was confirmed by surface plasmon resonance analysis.
CONCLUSIONS
Full-length TFPI binds to protein S in plasma and is reduced in genetic and acquired protein S deficiency. The concomitant TFPI deficiency substantially contributes to the hypercoagulable state associated with protein S deficiency.
Topics: Administration, Oral; Adult; Aged; Anticoagulants; Blood Coagulation; Case-Control Studies; Contraceptives, Oral; Down-Regulation; Female; Genetic Predisposition to Disease; Heterozygote; Humans; Lipoproteins; Male; Middle Aged; Protein Binding; Protein S; Protein S Deficiency; Risk Factors; Thrombin; Thromboplastin; Time Factors; Young Adult
PubMed: 20002538
DOI: 10.1111/j.1538-7836.2009.03712.x -
Journal of Thrombosis and Haemostasis :... Aug 2016Essentials Epithelial cell apoptosis is critical in the pathogenesis of idiopathic pulmonary fibrosis. Protein S, a circulating anticoagulant, inhibited apoptosis of...
UNLABELLED
Essentials Epithelial cell apoptosis is critical in the pathogenesis of idiopathic pulmonary fibrosis. Protein S, a circulating anticoagulant, inhibited apoptosis of lung epithelial cells. Overexpression of protein S in lung cells reduced bleomycin-induced pulmonary fibrosis. Intranasal therapy with exogenous protein S ameliorated bleomycin-induced pulmonary fibrosis.
SUMMARY
Background Pulmonary fibrosis is the terminal stage of interstitial lung diseases, some of them being incurable and of unknown etiology. Apoptosis plays a critical role in lung fibrogenesis. Protein S is a plasma anticoagulant with potent antiapoptotic activity. The role of protein S in pulmonary fibrosis is unknown. Objectives To evaluate the clinical relevance of protein S and its protective role in pulmonary fibrosis. Methods and Results The circulating level of protein S was measured in patients with pulmonary fibrosis and controls by the use of enzyme immunoassays. Pulmonary fibrosis was induced with bleomycin in transgenic mice overexpressing human protein S and wild-type mice, and exogenous protein S or vehicle was administered to wild-type mice; fibrosis was then compared in both models. Patients with pulmonary fibrosis had reduced circulating levels of protein S as compared with controls. Inflammatory changes, the levels of profibrotic cytokines, fibrosis score, hydroxyproline content in the lungs and oxygen desaturation were significantly reduced in protein S-transgenic mice as compared with wild-type mice. Wild-type mice treated with exogenous protein S showed significant decreases in the levels of inflammatory and profibrotic markers and fibrosis in the lungs as compared with untreated control mice. After bleomycin infusion, mice overexpressing human protein S showed significantly low caspase-3 activity, enhanced expression of antiapoptotic molecules and enhanced Akt and Axl kinase phosphorylation as compared with wild-type counterparts. Protein S also inhibited apoptosis of alveolar epithelial cells in vitro. Conclusions These observations suggest clinical relevance and a protective role of protein S in pulmonary fibrosis.
Topics: A549 Cells; Aged; Animals; Apoptosis; Bleomycin; Blood Proteins; Bronchoalveolar Lavage Fluid; Caspase 3; Epithelial Cells; Female; Fibrosis; Gene Expression Profiling; Humans; Idiopathic Pulmonary Fibrosis; Immunoenzyme Techniques; Inflammation; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Middle Aged; Phosphorylation; Protein S
PubMed: 27172994
DOI: 10.1111/jth.13362 -
Blood Jan 2009Human protein S is an anticoagulation protein. However, it is unknown whether protein S could regulate the expression and function of macrophage scavenger receptor A...
Human protein S is an anticoagulation protein. However, it is unknown whether protein S could regulate the expression and function of macrophage scavenger receptor A (SR-A) in macrophages. Human THP-1 monocytes and peripheral blood monocytes were differentiated into macrophages and then treated with physiological concentrations of human protein S. We found that protein S significantly reduced acetylated low-density lipoprotein (AcLDL) uptake and binding by macrophages and decreased the intracellular cholesteryl ester content. Protein S suppressed the expression of the SR-A at both mRNA and protein levels. Protein S reduced the SR-A promoter activity primarily through inhibition in the binding of transcription factors to the AP-1 promoter element in macrophages. Furthermore, human protein S could bind and induce phosphorylation of Mer receptor tyrosine kinase (Mer RTK). Soluble Mer protein or tyrosine kinase inhibitor herbimycin A effectively blocked the effects of protein S on AcLDL uptake. Immunohistochemical analysis revealed that the level of protein S was substantially increased in human atherosclerotic arteries. Thus, human protein S can inhibit the expression and activity of SR-A through Mer RTK in macrophages, suggesting that human protein S is a modulator for macrophage functions in uptaking of modified lipoproteins.
Topics: Atherosclerosis; Cholesterol Esters; Down-Regulation; Gene Expression; Humans; Lipoproteins, LDL; Macrophages; Oncogene Proteins; Protein Binding; Protein S; Proto-Oncogene Proteins; Receptor Protein-Tyrosine Kinases; Scavenger Receptors, Class A; Transcription, Genetic; U937 Cells; c-Mer Tyrosine Kinase; Axl Receptor Tyrosine Kinase
PubMed: 18922854
DOI: 10.1182/blood-2008-05-158048 -
Journal of Thrombosis and Haemostasis :... Oct 2004Plasma protein S normally circulates free (40%) or complexed with C4b-binding protein (PS-C4BP); only free protein S is a cofactor for activated protein C during factor... (Comparative Study)
Comparative Study
BACKGROUND
Plasma protein S normally circulates free (40%) or complexed with C4b-binding protein (PS-C4BP); only free protein S is a cofactor for activated protein C during factor (F) Va inactivation. Protein S-Heerlen lacks a carbohydrate group, leading to low plasma free protein S levels, but normal levels of PS-C4BP.
OBJECTIVES
Because protein S-Heerlen is not associated with thrombosis, we investigated whether PS-C4BP is directly anticoagulant in plasma and whether PS-Heerlen-C4BP has enhanced direct anticoagulant activity.
METHODS
An assay for protein S direct activity was applied to Heerlen-heterozygous plasmas. Free and complexed protein S were repeatedly isolated from normal and Heerlen-heterozygous plasmas and tested for direct anticoagulant activity in prothrombinase assays and in plasma.
RESULTS
Heerlen-heterozygous plasmas were deficient in free and total protein S antigen but had normal to high protein S direct anticoagulant activity. Purified Heerlen-heterozygous PS-C4BP was 7-fold more potent than normal PS-C4BP in inhibiting full prothrombinase activity, and 22-fold more potent in inhibiting prothrombin activation in the absence of FVa; it also specifically prolonged plasma clotting times 14-fold more than normal PS-C4BP. Heerlen-heterozygous PS-C4BP did not compete for limiting phospholipids any better than normal PS-C4BP. However, ligand blots and surface plasmon resonance studies showed that Heerlen-heterozygous PS-C4BP bound more avidly to FXa than did normal PS-C4BP (apparent Kd = 4.3 nm vs. 82 nm).
CONCLUSIONS
Plasma-derived PS-C4BP has direct anticoagulant activity in plasma and in purified systems. Enhanced direct activity of PS-Heerlen-C4BP may compensate for low free protein S levels and low cofactor activity in individuals with protein S-Heerlen.
Topics: Adult; Anticoagulants; Blood Coagulation Tests; Blood Platelets; Complement C4b-Binding Protein; Factor Xa; Family Health; Female; Heterozygote; Histocompatibility Antigens; Humans; Male; Phospholipids; Protein Binding; Protein S; Prothrombin; Thromboplastin
PubMed: 15456488
DOI: 10.1111/j.1538-7836.2004.00901.x -
The Journal of Clinical Investigation Oct 2009Protein S (ProS) is a blood anticoagulant encoded by the Pros1 gene, and ProS deficiencies are associated with venous thrombosis, stroke, and autoimmunity. These...
Protein S (ProS) is a blood anticoagulant encoded by the Pros1 gene, and ProS deficiencies are associated with venous thrombosis, stroke, and autoimmunity. These associations notwithstanding, the relative risk that reduced ProS expression confers in different disease settings has been difficult to assess without an animal model. We have now described a mouse model of ProS deficiency and shown that all Pros1-/- mice die in utero,from a fulminant coagulopathy and associated hemorrhages. Although ProS is known to act as a cofactor for activated Protein C (aPC), plasma from Pros1+/- heterozygous mice exhibited accelerated thrombin generation independent of aPC, and Pros1 mutants displayed defects in vessel development and function not seen in mice lacking protein C. Similar vascular defects appeared in mice in which Pros1 was conditionally deleted in vascular smooth muscle cells. Mutants in which Pros1 was deleted specifically in hepatocytes, which are thought to be the major source of ProS in the blood, were viable as adults and displayed less-severe coagulopathy without vascular dysgenesis. Finally, analysis of mutants in which Pros1 was deleted in endothelial cells indicated that these cells make a substantial contribution to circulating ProS. These results demonstrate that ProS is a pleiotropic anticoagulant with aPC-independent activities and highlight new roles for ProS in vascular development and homeostasis.
Topics: Animals; Blood Circulation; Blood Coagulation; Blood Coagulation Disorders; Blood Vessels; Brain; Embryo Loss; Endothelial Cells; Gene Targeting; Hemorrhage; Hepatocytes; Heterozygote; Homeostasis; Mice; Mice, Knockout; Protein C; Protein S; Protein S Deficiency; Spinal Cord; Thrombin
PubMed: 19729839
DOI: 10.1172/JCI39325 -
The Biochemical Journal Mar 1983Vitamin K-dependent protein S exists in two forms in human plasma, namely as the free protein and in complex with C4b-binding protein [Dahlbäck & Stenflo (1981) Proc....
Vitamin K-dependent protein S exists in two forms in human plasma, namely as the free protein and in complex with C4b-binding protein [Dahlbäck & Stenflo (1981) Proc. Natl. Acad. Sci. U.S.A. 78, 2512-2516]. Now reported is a simple purification procedure for human protein S that includes barium citrate adsorption, DEAE-Sephacel chromatography and chromatography on Blue Sepharose. The yield was approx. 30% relative to the concentration of free protein S in plasma, which was found to be approx. 10 mg/l. Purified protein S migrated as a single-chain band on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis under non-reducing conditions and as a doublet of Mr approx. 85 000 and 75 000 on reduction. A third band of Mr 16 000 was observed after electrophoresis of 125I-labelled protein S and radioautography of reduced samples. This band appears to be disulphide-linked to the 75 000-Mr chain before reduction. Thrombin converted the 85 000-Mr chain of protein S into a 75 000-Mr chain and an 8000-Mr fragment, the latter again being detectable only by radioautography of reduced samples. The 16 000-Mr fragment was not observed, suggesting its degradation by thrombin. Under non-reducing conditions, no change in apparent molecular weight of thrombin-treated protein S was observed, indicating disulphide linkage of the fragments. Thrombin also affected the mobility of protein S on agarose-gel electrophoresis in the presence of Ca2+, suggesting a decreased affinity to Ca2+ of the cleaved form of protein S as compared with the undegraded molecule. After activation of the complement system in human serum, protein S was found to be a constituent part of the complex formed by C4b-binding protein and component C4b.
Topics: Blood Coagulation; Chromatography, Ion Exchange; Complement Activation; Electrophoresis, Polyacrylamide Gel; Glycoproteins; Humans; Immunoelectrophoresis, Two-Dimensional; Protein S; Thrombin
PubMed: 6223624
DOI: 10.1042/bj2090837 -
Molecular Medicine Reports Dec 2021Label‑free quantitative mass spectrometry was used to analyze the differences in the granulation tissue protein expression profiles of patients with diabetic foot...
Label‑free quantitative mass spectrometry was used to analyze the differences in the granulation tissue protein expression profiles of patients with diabetic foot ulcers (DFUs) before and after negative‑pressure wound therapy (NPWT) to understand how NPWT promotes the healing of diabetic foot wounds. A total of three patients with DFUs hospitalized for Wagner grade 3 were enrolled. The patients received NPWT for one week. The granulation tissue samples of the patients prior to and following NPWT for one week were collected. The protein expression profiles were analyzed with label‑free quantitative mass spectrometry and the differentially expressed proteins (DEPs) in the DFU patients prior to and following NPWT for one week were identified. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were conducted to annotate the DEPs and DEP‑associated signaling pathways. Western blotting and ELISA were performed to validate the results. By comparing the differences in the protein profiles of granulation tissue samples prior to and following NPWT for one week, 36 proteins with significant differences were identified (P<0.05); 33 of these proteins were upregulated and three proteins were downregulated. NPWT altered proteins mainly associated with antioxidation and detoxification, the cytoskeleton, regulation of the inflammatory response, complement and coagulation cascades and lipid metabolism. The functional validation of the DEPs demonstrated that the levels of cathepsin S in peripheral blood and granulation tissue were significantly lower than those prior to NPWT (P<0.05), while the levels of protein S isoform 1, inter α‑trypsin inhibitor heavy chain H4 and peroxiredoxin‑2 in peripheral blood and granulation tissue were significantly higher than those prior to NPWT (P<0.05). The present study identified multiple novel proteins altered by NPWT and laid a foundation for further studies investigating the mechanism of action of NPWT.
Topics: Aged; Cathepsins; Diabetic Foot; Female; Foot Ulcer; Granulation Tissue; Humans; Male; Mass Spectrometry; Middle Aged; Negative-Pressure Wound Therapy; Peroxiredoxins; Protein S; Proteinase Inhibitory Proteins, Secretory; Proteome; Proteomics; Signal Transduction; Wound Healing
PubMed: 34608502
DOI: 10.3892/mmr.2021.12474 -
Blood Feb 2009Conflicting data have been reported on the risk for venous thrombosis in subjects with low free protein S levels. We performed a post-hoc analysis in a single-center... (Clinical Trial)
Clinical Trial
Conflicting data have been reported on the risk for venous thrombosis in subjects with low free protein S levels. We performed a post-hoc analysis in a single-center retrospective thrombophilic family cohort, to define the optimal free protein S level that can identify subjects at risk for venous thrombosis. Relatives (1143) were analyzed. Relatives with venous thrombosis (mean age 39 years) had lower free protein S levels than relatives without venous thrombosis (P < .001), which was most pronounced in the lowest quartile. Only relatives with free protein S levels less than the 5th percentile (< 41 IU/dL) or less than the 2.5th percentile (< 33 IU/dL) were at higher risk of first venous thrombosis compared with the upper quartile (> 91 IU/dL); annual incidence 1.20% (95% confidence interval [CI], 0.72-1.87) and 1.81% (95% CI, 1.01-2.99), respectively; adjusted hazard ratios 5.6, (95% CI, 2.7-11.5) and 11.3 (95% CI, 5.4-23.6). Recurrence rates were 12.12% (95 CI, 5.23-23.88) and 12.73% (95% CI, 5.12-26.22) per year; adjusted hazard ratios were 3.0 (95% CI, 1.03-8.5) and 3.4 (95% CI, 1.1-10.3). In conclusion, free protein S level can identify young subjects at risk for venous thrombosis in thrombophilic families, although the cutoff level lies far below the normal range in healthy volunteers.
Topics: Adolescent; Cohort Studies; Family; Humans; Incidence; Protein S; Retrospective Studies; Risk Factors; Venous Thrombosis
PubMed: 18945960
DOI: 10.1182/blood-2008-08-174128 -
Journal of Thrombosis and Haemostasis :... May 2017Essentials Protein S is a cofactor of activated protein C (APC) and tissue factor pathway inhibitor (TFPI). There are no assays to quantify separate APC and TFPI...
UNLABELLED
Essentials Protein S is a cofactor of activated protein C (APC) and tissue factor pathway inhibitor (TFPI). There are no assays to quantify separate APC and TFPI cofactor activities of protein S in plasma. We developed assays to measure the APC- and TFPI-cofactor activities of protein S in plasma. The assays were sensitive to protein S deficiency, and not affected by the Factor V Leiden mutation.
SUMMARY
Background Protein S plays an important role in the down-regulation of coagulation as cofactor for activated protein C (APC) and tissue factor pathway inhibitor (TFPI). Aim To develop functional assays to quantify the APC- and TFPI-cofactor activities of protein S in plasma. Methods APC- and TFPI-cofactor activities of protein S in plasma were measured using calibrated automated thrombography in protein S-depleted plasma supplemented with a small amount of sample plasma either in the presence of anti-TFPI antibodies and APC (APC-cofactor activity) or at excess full-length TFPI without APC (TFPI-cofactor activity). Total and free protein S levels in plasma were measured by ELISAs. Results Average APC-cofactor activities of protein S were 113%, 108% and 89% in plasma from normal individuals (n = 15), FV Leiden heterozygotes (n = 14) and FV Leiden homozygotes (n = 7), respectively, whereas the average APC-cofactor activity of protein S in plasma from heterozygous protein S-deficient individuals (n = 21) was significantly lower (55%). Similar trends were observed for the TFPI-cofactor activity of protein S, with averages of 109%, 115% and 124% in plasma from individuals with normal protein S levels and different FV Leiden genotypes, and 64% in plasma from protein S-deficient patients. APC-cofactor activities of protein S correlated significantly with free and total protein S antigen levels, whereas TFPI-cofactor activities correlated less with protein S antigen levels. Conclusion We have developed functional protein S assays that measure both the APC- and TFPI-cofactor activities of protein S in plasma, which are hardly if at all affected by the FV Leiden mutation.
Topics: Activated Protein C Resistance; Blood Coagulation; Blood Coagulation Tests; Case-Control Studies; Enzyme-Linked Immunosorbent Assay; Factor V; Humans; Lipoproteins; Point Mutation; Predictive Value of Tests; Protein C; Protein S; Protein S Deficiency; Thrombin
PubMed: 28211163
DOI: 10.1111/jth.13657 -
International Journal of Molecular... Mar 2019Acute lung injury is a fatal disease characterized by inflammatory cell infiltration, alveolar-capillary barrier disruption, protein-rich edema, and impairment of gas...
Acute lung injury is a fatal disease characterized by inflammatory cell infiltration, alveolar-capillary barrier disruption, protein-rich edema, and impairment of gas exchange. Protein S is a vitamin K-dependent glycoprotein that exerts anticoagulant, immunomodulatory, anti-inflammatory, anti-apoptotic, and neuroprotective effects. The aim of this study was to evaluate whether human protein S inhibits cell apoptosis in acute lung injury. Acute lung injury in human protein S transgenic and wild-type mice was induced by intratracheal instillation of lipopolysaccharide. The effect of human protein S on apoptosis of lung tissue cells was evaluated by Western blotting. Inflammatory cell infiltration, alveolar wall thickening, myeloperoxidase activity, and the expression of inflammatory cytokines were reduced in human protein S transgenic mice compared to the wild-type mice after lipopolysaccharide instillation. Apoptotic cells and caspase-3 activity were reduced while phosphorylation of extracellular signal-regulated kinase was enhanced in the lung tissue from human protein S transgenic mice compared to wild-type mice after lipopolysaccharide instillation. The results of this study suggest that human protein S is protective in lipopolysaccharide-induced acute lung injury by inhibiting apoptosis of lung cells.
Topics: Acute Lung Injury; Animals; Apoptosis; Humans; Lipopolysaccharides; MAP Kinase Signaling System; Male; Mice; Mice, Inbred C57BL; Protein S
PubMed: 30832349
DOI: 10.3390/ijms20051082