Dysregulation of Protein S in COVID-19.

Coronavirus Disease 2019 (COVID-19) has been widely associated with increased thrombotic risk, with many different proposed mechanisms. One such mechanism is acquired deficiency of protein S (PS), a plasma protein that regulates coagulation and inflammatory processes, including complement activation and efferocytosis. Acquired PS deficiency is common in patients with severe viral infections and has been reported in multiple studies of COVID-19. This deficiency may be caused by consumption, degradation, or clearance of the protein, by decreased synthesis, or by binding of PS to other plasma proteins, which block its anticoagulant activity. Here, we review the functions of PS, the evidence of acquired PS deficiency in COVID-19 patients, the potential mechanisms of PS deficiency, and the evidence that those mechanisms may be occurring in COVID-19.

Decreased protein C activity, lower ADAMTS13 antigen and free protein S levels accompanied by unchanged thrombin generation potential in hospitalized COVID-19 patients.

INTRODUCTION: COVID-19 is associated with an increased thromboembolic risk. However, the mechanisms triggering clot formation in those patients remain unknown. PATIENTS AND METHODS: In 118 adult Caucasian severe but non-critically ill COVID-19 patients (median age 58 years; 73 % men) and 46 controls, we analyzed in vitro plasma thrombin generation profile (calibrated automated thrombogram [CAT assay]) and investigated thrombophilia-related factors, such as protein C and antithrombin activity, free protein S level, presence of antiphospholipid antibodies and factor V Leiden R506Q and prothrombin G20210A mutations. We also measured circulating von Willebrand factor (vWF) antigen and a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13 (ADAMTS13) antigen and activity. In patients, blood samples were collected on admission to the hospital before starting any therapy, including heparin. Finally, we examined the relationship between observed alterations and disease follow-up, such as thromboembolic complications. RESULTS: COVID-19 patients showed 17 % lower protein C activity, 22 % decreased free protein S levels, and a higher prevalence of positive results for IgM anticardiolipin antibodies. They also had 151 % increased vWF, and 27 % decreased ADAMTS13 antigens compared with controls (p < 0.001, all). On the contrary, thrombin generation potential was similar to controls. In the follow-up, pulmonary embolism (PE) occurred in thirteen (11 %) patients. They were characterized by a 55 % elevated D-dimer (p = 0.04) and 2.7-fold higher troponin I (p = 0.002) during hospitalization and 29 % shorter time to thrombin peak in CAT assay (p = 0.009) compared to patients without PE. CONCLUSIONS: In COVID-19, we documented prothrombotic abnormalities of peripheral blood. PE was characterized by more dynamic thrombin generation growth in CAT assay performed on admittance to the hospital.

Protein S: function, regulation, and clinical perspectives.

PURPOSE OF REVIEW: Protein S (PS) is an essential natural anticoagulant. PS deficiency is a major contributor to acquired hypercoagulability. Acquired hypercoagulability causes myocardial infarction, stroke, and deep vein thrombosis in millions of individuals. Yet, despite its importance in hemostasis, PS is the least understood anticoagulant. Even after 40 years since PS was first described, we are still uncovering information about how PS functions. The purpose of this review is to highlight recent findings that advance our understanding of the functions of PS and explain hypercoagulability caused by severe PS deficiency. RECENT FINDINGS: PS has long been described as a cofactor for Activated Protein C (APC) and Tissue Factor Pathway Inhibitor (TFPI). However, a recent report describes direct inhibition of Factor IXa (FIXa) by PS, an activity of PS that had been completely overlooked. Thrombophilia is becoming a more frequently reported disorder. Hereditary PS deficiency is an anticoagulant deficiency that results eventually in thrombophilia. In addition, PS deficiency is a predisposing factor for venous thromboembolism (VTE), but an effect of PS deficiency in arterial thrombosis, such as arterial ischemic stroke, is uncertain. Plasma PS concentration decreases in pregnant women. Inherited thrombophilias are important etiologies for recurrent pregnancy loss, and anticoagulation therapy is of benefit to women with recurrent pregnancy loss who had documented only PS deficiency.Hypoxia is a risk factor for VTE, and hypoxia downregulates plasma PS level. Importantly, COVID-19 can lead to hypoxemia because of lung damage from IL6-driven inflammatory responses to the viral infection. Because hypoxia decreases the abundance of the key anticoagulant PS, we surmise that the IL6-induced cytokine explosion combined with hypoxemia causes a drop in PS level that exacerbates the thrombotic risk in COVID-19 patients. SUMMARY: This review is intended to advance understanding of the anticoagulant function of an important plasma protein, PS. Despite 40+ years of research, we have not had a complete description of PS biology as it pertains to control of blood coagulation. However, the picture of PS function has become sharper with the recent discovery of FIXa inhibition by PS. Hemostasis mediated by PS now includes regulation of FIXa activity alongside the cofactor activities of PS in the TFPI/APC pathways. In addition, the direct inhibition of FIXa by PS suggests that PS, particularly a small derivative of PS, could be used to treat individuals with PS deficiencies or abnormalities that cause thrombotic complications.

Vitamin K metabolism as the potential missing link between lung damage and thromboembolism in Coronavirus disease 2019.

Coronavirus disease 2019 (Covid-19), caused by severe acute respiratory syndrome coronavirus (SARS-CoV)-2, exerts far-reaching effects on public health and socio-economic welfare. The majority of infected individuals have mild to moderate symptoms, but a significant proportion develops respiratory failure due to pneumonia. Thrombosis is another frequent manifestation of Covid-19 that contributes to poor outcomes. Vitamin K plays a crucial role in the activation of both pro- and anticlotting factors in the liver and the activation of extrahepatically synthesised protein S which seems to be important in local thrombosis prevention. However, the role of vitamin K extends beyond coagulation. Matrix Gla protein (MGP) is a vitamin K-dependent inhibitor of soft tissue calcification and elastic fibre degradation. Severe extrahepatic vitamin K insufficiency was recently demonstrated in Covid-19 patients, with high inactive MGP levels correlating with elastic fibre degradation rates. This suggests that insufficient vitamin K-dependent MGP activation leaves elastic fibres unprotected against SARS-CoV-2-induced proteolysis. In contrast to MGP, Covid-19 patients have normal levels of activated factor II, in line with previous observations that vitamin K is preferentially transported to the liver for activation of procoagulant factors. We therefore expect that vitamin K-dependent endothelial protein S activation is also compromised, which would be compatible with enhanced thrombogenicity. Taking these data together, we propose a mechanism of pneumonia-induced vitamin K depletion, leading to a decrease in activated MGP and protein S, aggravating pulmonary damage and coagulopathy, respectively. Intervention trials should be conducted to assess whether vitamin K administration plays a role in the prevention and treatment of severe Covid-19.

Hemostatic alterations linked to body fat distribution, fatty liver, and insulin resistance.

OBJECTIVE: Obesity, in particular visceral obesity, and insulin resistance emerged as major risk factors for severe coronavirus disease 2019 (COVID-19), which is strongly associated with hemostatic alterations. Because obesity and insulin resistance predispose to thrombotic diseases, we investigated the relationship between hemostatic alterations and body fat distribution in participants at risk for type 2 diabetes. SUBJECTS: Body fat distribution (visceral and subcutaneous abdominal adipose tissue) and liver fat content of 150 participants - with impaired glucose tolerance and/or impaired fasting glucose - were determined using magnetic resonance imaging and spectroscopy. Participants underwent precise metabolic characterization and major hemostasis parameters were analyzed. RESULTS: Procoagulant factors (FII, FVII, FVIII, and FIX) and anticoagulant proteins (antithrombin, protein C, and protein S) were significantly associated with body fat distribution. In patients with fatty liver, fibrinogen (298 mg/dl vs. 264 mg/dl, p = 0.0182), FVII (99% vs. 90%, p = 0.0049), FVIII (114% vs. 90%, p = 0.0098), protein C (124% vs. 111%, p = 0.0006), and protein S (109% vs. 89%, p < 0.0001) were higher than in controls. In contrast, antithrombin (97% vs. 102%, p = 0.0025) was higher in control patients. In multivariate analyses controlling for insulin sensitivity, body fat compartments, and genotype variants (PNPLA3I148MM/MI/TM6SF2E167kK/kE), only protein C and protein S remained significantly increased in fatty liver. CONCLUSIONS: Body fat distribution is significantly associated with alterations of procoagulant and anticoagulant parameters. Liver fat plays a key role in the regulation of protein C and protein S, suggesting a potential counteracting mechanism to the prothrombotic state in subjects with prediabetes and fatty liver.

Anticoagulant protein S in COVID-19: low activity, and associated with outcome.

Introduction. COVID-19 disease was associated with both thrombo-embolic events and in-situ thrombi formation in small vessels. Antiphospholipidic antibodies were found in some studies.Aim. Assessment of protein S activity in patients with COVID-19 as a cause of this prothrombotic state, and of the association of protein S activity with worse outcome.Methods. All patients admitted for COVID-19 disease in a university hospital between 15th of May and 15th of July 2020 were prospectively enrolled into this cohort study. Patients treated with antivitamin K anticoagulants and with liver disease were excluded. All patients had protein S activity determined at admission. The main outcome was survival, while secondary outcomes were clinical severity and lung damage.Results. 91 patients were included, of which 21 (23.3%) died. Protein S activity was decreased in 65% of the patients. Death was associated with lower activity of protein S (median 42% vs. 58%, p < 0.001), and the association remained after adjustment for age, inflammation markers and ALAT. There was a dose-response relationship between protein S activity and clinical severity (Kendall_tau coefficient = -0.320, p < 0.001; Jonckheere-Terpstra for trend: p < 0.001) or pulmonary damage on CT scan (Kendall_tau coefficient = -0.290, p < 0.001; Jonckheere-Terpstra for trend: p < 0.001). High neutrophil count was also independently associated with death (p = 0.002).Conclusion. Protein S activity was lower in COVID-19 patients, and its level was associated with survival and disease severity, suggesting that it may have a role in the thrombotic manifestations of the disease.

Role of Vitamin K-Dependent Factors Protein S and GAS6 and TAM Receptors in SARS-CoV-2 Infection and COVID-19-Associated Immunothrombosis.

The vitamin K-dependent factors protein S (PROS1) and growth-arrest-specific gene 6 (GAS6) and their tyrosine kinase receptors TYRO3, AXL, and MERTK, the TAM subfamily of receptor tyrosine kinases (RTK), are key regulators of inflammation and vascular response to damage. TAM signaling, which has largely studied in the immune system and in cancer, has been involved in coagulation-related pathologies. Because of these established biological functions, the GAS6-PROS1/TAM system is postulated to play an important role in SARS-CoV-2 infection and progression complications. The participation of the TAM system in vascular function and pathology has been previously reported. However, in the context of COVID-19, the role of TAMs could provide new clues in virus-host interplay with important consequences in the way that we understand this pathology. From the viral mimicry used by SARS-CoV-2 to infect cells, to the immunothrombosis that is associated with respiratory failure in COVID-19 patients, TAM signaling seems to be involved at different stages of the disease. TAM targeting is becoming an interesting biomedical strategy, which is useful for COVID-19 treatment now, but also for other viral and inflammatory diseases in the future.

Coagulation profile of COVID-19 patients admitted to the ICU: An exploratory study.

BACKGROUND: Coagulation abnormalities in COVID-19 patients have not been addressed in depth. OBJECTIVE: To perform a longitudinal evaluation of coagulation profile of patients admitted to the ICU with COVID-19. METHODS: Conventional coagulation tests, rotational thromboelastometry (ROTEM), platelet function, fibrinolysis, antithrombin, protein C and S were measured at days 0, 1, 3, 7 and 14. Based on median total maximum SOFA score, patients were divided in two groups: SOFA ≤ 10 and SOFA > 10. RESULTS: Thirty patients were studied. Some conventional coagulation tests, as aPTT, PT and INR remained unchanged during the study period, while alterations on others coagulation laboratory tests were detected. Fibrinogen levels were increased in both groups. ROTEM maximum clot firmness increased in both groups from Day 0 to Day 14. Moreover, ROTEM-FIBTEM maximum clot firmness was high in both groups, with a slight decrease from day 0 to day 14 in group SOFA ≤ 10 and a slight increase during the same period in group SOFA > 10. Fibrinolysis was low and decreased over time in all groups, with the most pronounced decrease observed in INTEM maximum lysis in group SOFA > 10. Also, D-dimer plasma levels were higher than normal reference range in both groups and free protein S plasma levels were low in both groups at baseline and increased over time, Finally, patients in group SOFA > 10 had lower plasminogen levels and Protein C ​​than patients with SOFA <10, which may represent less fibrinolysis activity during a state of hypercoagulability. CONCLUSION: COVID-19 patients have a pronounced hypercoagulability state, characterized by impaired endogenous anticoagulation and decreased fibrinolysis. The magnitude of coagulation abnormalities seems to correlate with the severity of organ dysfunction. The hypercoagulability state of COVID-19 patients was not only detected by ROTEM but it much more complex, where changes were observed on the fibrinolytic and endogenous anticoagulation system.

TAM receptors: A phosphatidylserine receptor family and its implications in viral infections.

Phosphatidylserine (PS) is an anionic phospholipid that is usually localized in the inner leaflets of the plasma membrane. However, the enzyme scramblase catalyzes the externalization of PS on the outer leaflet of the plasma membrane during apoptosis or cellular stress. This event prompts the recognition of PS displaying cells by phagocytes leading to "apoptotic clearance." Multiple PS receptors (PSRs) mediate this process including members from the TAM (Tyro3, Axl, Mertk) receptor Tyrosine kinases (RTKs) by interacting with PS via bridging proteins like Gas6 and ProS1. Ironically, this network (PS/TAM) that evolved for boosting cellular health through clearance of apoptotic and necrotic cells, has been manoeuvred by pathogens and tumor cells using "apoptotic mimicry." Enveloped viruses, responsible for most of the lethal epidemics and pandemics including the current SARS-CoV2 outbreak, have employed apoptotic mimicry to their advantage. In the current chapter, we summarize the existing knowledge regarding the involvement of PS/Gas6, ProS1/TAM in facilitating infectivity in a diverse set of cell lines, animals as well as organoids. This network executes a largely proviral role in facilitating infection as seen with Zika, Ebola, Influenza and Dengue viruses. However, this response varies with strains and the cells infected, and in some cases, this same signaling displays an antiviral function. We also report multiple studies that have used neutralizing antibodies and small molecule inhibitors in successfully reducing viral replication and ameliorating pathogenicity. Knowledge about this unique signaling pathway and measures that can be taken to inhibit it is most valuable now given how enveloped viruses lead to plagues on the entire globe.

Red blood cell exchange for SARS-CoV-2: A Gemini of therapeutic opportunities.

As of now, therapeutic strategies for the novel coronavirus (SARS-CoV-2) are limited and much focus has been placed on social distancing techniques to "flatten the curve". Initial treatment efforts including ventilation and hydroxychloroquine garnered significant controversy and today, SARS-CoV-2 outbreaks are still occurring throughout the world. Needless to say, new therapeutic strategies are needed to combat this unprecedented pandemic. Nature Reviews Immunology recently published an article hypothesizing the pathogenesis of TAM (Tyro3, Axl, and Mer) receptor signaling in COVID-19. In it they expressed that hypercoagulation and immune hyper-reaction could occur secondary to decreased Protein S (PROS1). And hypoxia has been recently discovered to significantly decrease expression of PROS1. Regarding the cause of hypoxia in COVID-19; NIH funded research utilizing state-of-the-art topologies has recently demonstrated significant metabolomic, proteomic, and lipidomic structural aberrations in hemoglobin (Hb) secondary to infection with SARS-CoV-2. In this setting, Hb may be incapacitated and unable to respond to environmental variations, compromising RBCs and oxygen delivery to tissues. The use of red blood cell exchange would target hypoxia at its source; representing a Gemini of therapeutic opportunities.

COVID-19: a probable role of the anticoagulant Protein S in managing COVID-19-associated coagulopathy.

The COVID-19 pandemic has caused monumental mortality, and there are still no adequate therapies. Most severely ill COVID-19 patients manifest a hyperactivated immune response, instigated by interleukin 6 (IL6) that triggers a so called "cytokine storm" and coagulopathy. Hypoxia is also associated with COVID-19. So far overlooked is the fact that both IL6 and hypoxia depress the abundance of a key anticoagulant, Protein S. We speculate that the IL6-driven cytokine explosion plus hypoxemia causes a severe drop in Protein S level that exacerbates the thrombotic risk in COVID-19 patients. Here we highlight a mechanism by which the IL6-hypoxia curse causes a deadly hypercoagulable state in COVID-19 patients, and we suggest a path to therapy.

COVID-19 coagulopathy: An in-depth analysis of the coagulation system.

BACKGROUND: Abnormal coagulation parameters have been reported in COVID-19-infected patients. Although the underlying mechanism of COVID-19 coagulopathy remains unknown, it has been suggested to be a form of disseminated intravascular coagulation (DIC). OBJECTIVES: The aim of our study was to analyze the coagulation parameters of patients with COVID-19, determine whether coagulation factors consumption occurs and identify potential prognostic biomarkers of the disease. PATIENTS/METHODS: Blood samples from hospitalized patients with COVID-19 pneumonia were collected. We performed basic coagulation tests and quantification of coagulation factors and physiological inhibitor proteins. Laboratory data were compared with clinical data and outcomes. RESULTS: The study involved 206 patients (63.6% male). D-dimer was particularly elevated (median 450 ng/mL; IQR 222.5-957.3). Free protein S levels were below the normal range (median 56.6%; IQR: 43.6-68.9), and factor VIII showed an increasing trend (median 173.4%; IQR: 144.1-214.9). However, all coagulation factors were within normal limits. We found no correlation between abnormal coagulation parameters and thrombosis, except for higher D-dimer (HR 1.99; 95% CI 1.3-3.1; P = .002). CONCLUSIONS: COVID-19 is associated with coagulopathy that correlates with poor prognosis. However, we did not demonstrate a consumption of coagulation factors, as seen in DIC.