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This prospective observational study including 245 patients hospitalized for COVID‑19 demonstrates that the degree of hypercoagulability, hypofibrinolysis, and endothelial dysfunction is associated with the requirement for more advanced respiratory support. These abnormalities are significantly attenuated within 28 days since the admission, but intergroup differences persist in the most severely ill patients. This may suggest that endotheliopathy plays a more important role in the pathogenesis of COVID‑19 than abnormalities of coagulation and fibrinolysis.

Introduction

The COVID‑19 pandemic was declared by the World Health Organization in 2020 and led to millions of hospitalizations and deaths worldwide.¹ Over time, the growing body of evidence on the disease mechanisms led to the development of effective treatments and vaccines.^2-4 It is now well established that COVID‑19 pathophysiology is a complex interplay between inflammation, hemostatic disturbances, and immunological abnormalities.⁵ Clinically, the disease usually presents with respiratory symptoms, frequently leading to respiratory failure and requirement for oxygen therapy, mechanical ventilation, and even extracorporeal oxygenation.^6,7

One of the characteristic features of COVID‑19 is a high prevalence of thromboembolic complications, which encouraged the international societies to recommend a routine use of thromboprophylaxis in the patients requiring hospitalization for SARS‑CoV‑2 infection.^8,9 At the same time, basic studies revealed hypercoagulability, hypofibrinolysis, and evidence of endothelial dysfunction in COVID‑19 patients.^10-13 The endothelial damage is believed to be a crucial contributor to respiratory failure in the course of COVID‑19. It is responsible for intrapulmonary hyperperfusion and impairment of hypoxic vasoconstriction, thus leading to alveolar damage, edema, hemorrhage, and intra‑alveolar fibrin deposition in a manner similar to non–COVID‑19 acute respiratory distress syndrome.⁵ The degree of these abnormalities seems to be related to the severity of the disease.^14,15 Less is known about the temporal changes in hemostatic activity and their patterns in the patients with respiratory failure of a different degree.^16-18

We conducted this prospective, observational study in the patients hospitalized for COVID‑19 to evaluate their hemostatic activity depending on the severity of respiratory failure. Additionally, we aimed to assess the direction and magnitude of changes in coagulation, fibrinolysis, and endothelial function during a short‑term observation among COVID‑19 survivors.

Patients and methods

Results

Temporal variability of hemostatic activity

The 28‑day follow‑up evaluation of hemostatic activity was available for 164 patients (66.9%). We revealed a significant decrease in all coagulation, fibrinolysis, and endothelial dysfunction markers, except for plasminogen activity (123.75% vs 121.6%; P = 0.94). Particularly large differences were observed for fibrinogen (4.5 vs 3.2 g/l; P <⁠0.001), PAP (1537.6 vs 741.5 ng/ml; P <⁠0.001), PAI‑1:Ag (34.1 vs 22.8 ng/ml; P <⁠0.001), and vWF:Ag (300.1% vs 178.2%; P <⁠0.001). This analysis is summarized in Table 3 and visualized in Figures 1 and 2. Additional analyses revealed that similar trends were evident in all subgroups (Supplementary material, Table S1).

Table 3. Comparison of hemostatic markers and antiphospholipid antibody levels between the admission and 28‑day follow‑up

Factor	Admission	28‑day follow‑up	P valuea
Data are presented as median (interquartile range). a P value for the Wilcoxon test Abbreviations: see Table 2
Fibrinogen, g/l	4.5 (3.8–5.2)	3.2 (2.8–3.6)	<⁠0.001
Factor VIII, %	137.35 (108.83–171.42)	121.1 (88.77–161)	<⁠0.001
TAT, µg/l	7.74 (5.42–13.12)	4.68 (3.6–6.44)	<⁠0.001
CLT, min	112 (99.25–132)	99 (88–109)	<⁠0.001
D‑Dimer, ng/ml	834.5 (584.25–1473.5)	643.5 (377.75–1083.25)	<⁠0.001
α2-Antiplasmin, %	109.95 (104.5–115.73)	105.2 (100–108.8)	<⁠0.001
Plasminogen, %	123.75 (111.57–134.52)	121.6 (112.98–133.38)	0.94
t‑PA:Ag, ng/ml	12.84 (9.02–16.94)	8.1 (6.08–9.85)	<⁠0.001
PAI‑1:Ag, ng/ml	34.06 (23.73–43.06)	22.76 (14.49–33.36)	<⁠0.001
PAP, ng/ml	1537.58 (1342.16–1991.57)	741.45 (596.15–1086.99)	<⁠0.001
vWF:Ag, %	300.1 (240.33–365)	178.2 (130.57–232.85)	<⁠0.001
ADAMTS‑13, IU/ml	0.87 (0.8–0.92)	0.88 (0.82–0.93)	0.003
vWF:Ag/ADAMTS‑13 ratio	3.56 (2.91–4.21)	2.02 (1.46–2.74)	<⁠0.001
aCL IgG antibodies, GPL	5.86 (4.26–9.12)	12.55 (8.33–20.9)	<⁠0.001
aCL IgM antibodies, MPL	14.60 (11.54–20.23)	17.91 (12.96–29.03)	<⁠0.001
aβ2GP1 IgG antibodies, SGU	2.96 (2.77–3.38)	2.59 (2.33–3.32)	<⁠0.001
aβ2GP1 IgM antibodies, SMU	2.47 (1.75–4.4)	2.69 (2.04–4.64)	0.06
aPS/PT IgG	7.65 (6.67–10.02)	8.66 (7.19–10.47)	<⁠0.001
aPS/PT IgM	14.58 (10.80–23.31)	15.31 (10.92–24.01)	0.76

Discussion

In this prospective observational study including 245 patients hospitalized for COVID‑19, we confirmed a tendency for hypercoagulability, hypofibrinolysis, and endothelial dysfunction at a degree associated with severity of respiratory failure. In all studied groups, the markers of coagulation and fibrinolysis decreased within 28 days since initial hospital admission but endothelial dysfunction remained most pronounced in a subgroup of patients with the most severe COVID‑19.

The analysis of coagulation markers on admission revealed increased levels of fibrinogen and TAT suggesting marked hypercoagulability. It was accompanied by a significant fibrinolysis shutdown reflected by prolonged CLT. We believe that increased concentrations of potent fibrinolysis inhibitors (α2AP and PAI‑1) outweighed elevated levels of tPA and shifted the balance toward hypofibrinolysis.^23,24 Finally, increased vWF:Ag, PAI‑1, and particularly high vWF:Ag/ADAMTS‑13 ratio suggest significant endothelial dysfunction and significant deviations in the ADAMTS‑13–von Willebrand factor axis.^15,25-27 Our results corroborate previous reports in terms of observed abnormalities and confirm their association with severity of COVID‑19. The majority of available studies divided patients according to their survival status or requirement for intensive care unit (ICU) admission.^14,28 Our analysis broadens the current knowledge on this issue by using a wider panel of hemostatic markers and by focusing on patients treated primarily in hospital wards and stratified according to the modality of respiratory support. Similar approach was adopted previously by Rauch et al,²⁹ who reported analogous results but used more common coagulation tests without assessment of fibrinolysis. The hemostatic abnormalities described in patients with COVID‑19 made appropriate thromboprophylaxis one of the cornerstones of COVID‑19 management. There are also some promising reports showing a potential benefit of fibrinolytic therapy in the patients with severe COVID‑19.^30-32

The requirement for postdischarge thromboprophylaxis remains one of the most widely discussed topics regarding the management of COVID‑19 patients. The American Society of Hematology guideline panel discourages it, while some experts suggest considering extended thromboprophylaxis in the patients with a high risk for thrombotic complications and without a high risk for bleeding.^33,34 On the one hand, the risk of thromboembolic events in patients with COVID‑19 is increased up to 4 months after the diagnosis, in comparison with COVID‑19–negative controls.³⁵ On the other hand, a meta‑analysis including nearly 19 000 COVID‑19 survivors revealed that the absolute incidence of postdischarge venous thromboembolism (VTE) in patients hospitalized for COVID‑19 was low and accounted for 1.8%.³⁶ The currently available literature exploring the impact of COVID‑19 on hemostasis after discharge from the hospital pertained to ICU survivors. Serial measurements using viscoelastic assays showed that hypercoagulability and hypofibrinolysis persisted over 6 weeks since admission, but were no longer present in 3‑month and 6‑month follow‑ups.^17,18,37 Our results indicate that the severity of all initially observed abnormalities decreased significantly within 28 days since the admission to the hospital, and such a trend was observed in all subgroups. Another important observation is the disappearance of intergroup differences found upon admission in all assessed markers of coagulation and fibrinolysis. This may suggest that hypercoagulability and hypofibrinolysis are typical for acute phase of the disease and subside relatively quickly. Conversely, the degree of endothelial dysfunction remains the highest in the advanced respiratory support group. This observation, together with the results of Fogarty et al,³⁸ may indicate a potentially crucial role of endothelial dysfunction in the development of long COVID. Unfortunately, the lack of a healthy control group in our study does not allow for drawing conclusions about normalization of hemostatic parameters in the follow‑up. Further studies are required to precisely describe the association of prolonged endotheliopathy with long‑term sequelae of COVID‑19.

The clinical significance of antiphospholipid antibodies in COVID‑19 has been discussed since the first months of the pandemic. The transient presence of aPL antibodies is not an exceptional feature of COVID‑19, and had previously been observed in other viral infections.³⁹ In previous studies including COVID‑19 patients, Lupus anticoagulant was found in 5% to 90% of ICU patients and in 2% to 66% of regular ward patients. The prevalence of other aPL antibodies was characterized by similar variability.⁴⁰ However, to date there has been no robust evidence on their pathogenic role in thrombotic events during COVID‑19, and data on the association between the level of aPL antibodies and severity of the disease are conflicting.^41-44 Our study shows some differences between the groups in aPS/PT and aβ₂GP1 IgM antibodies levels. Their magnitude and variable directions suggest that the relation between antiphospholipid antibodies and severity of respiratory failure is rather weak and probably clinically insignificant. This corroborates the results of a meta‑analysis by Taha et al,⁴⁵ which showed a lack of association between aPL positivity and requirement for invasive mechanical ventilation. Temporal changes in antiphospholipid antibodies profile have not been studied extensively but available studies suggest that the majority of patients with initially positive antibodies have normal levels of antiphospholipid antibodies after 1 to 6 months.^46,47 Our study shows that aCL and aPS/PT levels increase, while aβ₂GP1 levels decrease within 28 days since the initial admission. Importantly, 1 in 4 patients had at least double‑positive antibodies on the 28th day. Unfortunately, we did not gather long‑term data on antibody levels, which would definitely offer some valuable insight into the dynamics of the antibody clearance. This information is crucial in terms of establishing an optimal approach to clinical follow‑up in the patients with COVID‑19 in whom antiphospholipid antibodies were found in the acute phase of the disease. The current recommendations suggest retesting antiphospholipid antibodies–positive patients 12 weeks after the diagnosis, however, this recommendation is based on low‑quality data and further research in this area is warranted.⁴⁸

The main strength of this paper is a precise description of the disease course with particular regard to respiratory failure and wide range of tests of hemostatic activity. We are aware of several weaknesses of our study. First, a relatively small study sample as well as relatively low mortality and VTE incidence hindered a precise assessment of associations between these outcomes and hemostatic activity. Second, despite our best efforts, low‑molecular‑weight heparin was probably administered in some of the enrolled patients before blood samples were collected, which could affect the results. Third, even though we actively endorsed patients to attend the follow‑up visit, the proportion of patients missing in the 28‑day follow‑up was relatively high. Finally, some might argue that different stratification of patients according to the severity of respiratory failure would be more appropriate, particularly with regard to separation of the patients requiring intubation and hospitalization in the ICU. However, due to a very low number of such patients, after careful discussion among the experts, we decided to include the individuals requiring invasive mechanical ventilation into the advanced respiratory support group.

Summing up, this prospective observational study confirmed that the patients hospitalized for COVID‑19 are characterized by hypercoagulability, hypofibrinolysis, and endothelial dysfunction and these abnormalities are related to the severity of respiratory failure. Interestingly, the majority of hemostatic derangements subsided, at least partially, within a month since admission, regardless of the disease course, with an exception of endothelial damage, which was still present at that time. This may suggest that evaluation of hemostasis on admission may be useful in the prediction of the disease severity, and that the observed abnormalities in coagulation and fibrinolysis are limited to the acute phase of the disease. Further studies are required to determine whether persistent endothelial dysfunction and presence of antibodies are associated with long‑term thromboembolic complications.