In Vivo Clearance of Human Protein S in a Mouse Model

Abstract

Objective-To explore the effect of the Heerlen polymorphism and C4b-binding protein (C4BP) on protein S catabolism in vitro and in vivo. Methods and Results-Radiolabeled protein S was efficiently bound and intracellularly degraded by THP-1 macrophages, and both processes were strongly reduced in the presence of the protein S-carrier protein C4BP. To test whether C4BP displays a similar protective effect in vivo, survival experiments were performed in mice. In the absence of C4BP, radiolabeled human protein S disappeared in a biphasic manner (mean residence time [MRT] 2 hours). However, the presence of C4BP resulted in a 4-fold prolonged survival of protein S (MRT 8 hours; PϽ0.0001). We also applied this experimental model to recombinant protein S-Heerlen, a naturally occurring variant that contains a Ser460Pro substitution. These clearance experiments revealed a strongly decreased survival of recombinant protein S-S460P (MRT 0.6 hours; Pϭ0.021), which could be compensated partially by C4BP (MRT 1.4 hours; Pϭ0.012 compared with protein S-S460P). Conclusion-Protein S-S460P has a reduced survival in vivo, which may explain the low levels of free protein S in individuals carrying this polymorphism. Furthermore, C4BP prevents premature clearance of protein S and uses this ability to compensate the increased clearance of protein S-S460P. (Arterioscler Thromb Vasc Biol. 2005;25:2209-2215.)Key Words: protein S Ⅲ C4b-binding Ⅲ clearance P rotein S circulates in plasma at a concentration of 0.3 mol/L, although this value may vary between individuals. 1 Only 30% to 40% of protein S circulates in a free form, whereas the remaining part is in complex with C4b-binding protein (C4BP). 2 Various isoforms of C4BP exist, and only those that contain the ␤-chain (ie, C4BP␣␤) bind protein S. 3 Because the C4BP␣␤ plasma concentration is lower than that of protein S, the molar excess of protein S over C4BP␣␤ determines the amount of free protein S.The function of protein S relates to various physiological processes, including the clearance of apoptotic cells and neuronal protection during ischemic/hypoxic injury. 4 -6 However, protein S is originally known from its role in the anticoagulant process, 7,8 in which it functions as a cofactor for activated protein C (APC). 9,10 In addition, protein S may downregulate the coagulation pathway independently of APC. 11,12 The notion that deficiencies in protein S are associated with an increased thrombotic tendency 13,14 demonstrates that appropriate plasma levels of protein S are needed to maintain the hemostatic balance.Two categories of protein S deficiency can be distinguished: 15 (1) qualitative defects (type II deficiency), and (2) quantitative defects (type I or III deficiency). Type I deficiency refers to a parallel reduction of total and free protein S. Type III deficiency is characterized by low levels of free protein S but normal levels of total protein S. However, both types of quantitative deficiencies often occur within the same family, 1 and more detailed analysis revealed tha...

A Case of a Pediatric Patient With Protein S Heerlen Polymorphism and Deep Venous Thrombosis

“…6 This may be explained by increased clearance of free PS with 3-fold reduced in vivo survival from 2 to 0.6 hours. 7 In addition, PS Heerlen has been found to slightly increase thrombin generation in the presence of APC compared with wild type PS. 4 The association between PS Heerlen and VTE is controversial and is currently described as a variant of uncertain significance by the American College of Medical Genetics and Genomics.…”

“…However, it has since been shown that the Heerlen polymorphism is associated with a reduced level of free PS in heterozygotes and very low levels in homozygotes 6. This may be explained by increased clearance of free PS with 3-fold reduced in vivo survival from 2 to 0.6 hours 7. In addition, PS Heerlen has been found to slightly increase thrombin generation in the presence of APC compared with wild type PS 4…”

Thrombophilia Screening: Not So Straightforward

“…148,149 PS Heerlen has a reduced half-life in vivo, which has been linked to the mild and marked FPS reductions in heterozygotes and homozygotes, respectively. 150,151 Heeb et al reported that the lack of clear evidence (at the time) for increased VTE risk could be due to increased direct anticoagulant activity of PS Heerlen in complex with C4BP, 152 whereas Suchon et al hypothesized that since the mutation is located in the TFPI binding domain, impaired TFPI function could contribute to thrombosis risk with PS Heerlen. 148 PS Tokushima (p.Lys196Glu) is prevalent in the Japanese population, causing a type II deficiency with an increased risk for DVT of 4-to 8-fold in heterozygotes.…”

Unfolded Von Willebrand Factor Binds Protein S and Reduces Anticoagulant Activity

“…7 The remaining ~60% of plasma PS is bound to the β-chain of complement component C4b-binding protein (C4BP-β). 4 C4BP stabilizes PS, preventing its clearance, 8 and blocks most anticoagulant activity. 9 Acquired PS deficiency is a common complication of severe viral infections, including human immunodeficiency virus, 10,11 varicella, 12 dengue, 13 and severe acute respiratory syndromecoronavirus-2 (SARS-CoV-2), [14][15][16][17][18][19] all of which are associated with an increased risk of thrombosis.…”