Thrombotic Events and Anticoagulation-Related Bleeding Complications in Critically Ill Patients with Coronavirus Disease 2019.

Background: Thrombosis (T) is common in coronavirus disease 2019 (COVID-19) patients, and d-dimer concentrations correlate with outcomes. Controversy exists with regards to anticoagulation (AC) for patients. We implemented a full-heparinization AC protocol from the onset of the pandemic and hypothesized that a safety signal would be undetectable. Patients and Methods: Prospective evaluation of 111 patients with COVID-19 critical illness hospitalized from March to June 2020. All patients received therapeutic heparinoid-based AC from admission. Incidences of T, bleeding (B), or both (BT) were noted. The primary outcome was mortality. Kruskal-Wallis test and logistic regression were performed. Results are expressed as n (%), median (interquartile range) and odds ratios with 95% confidence intervals. Alpha was set at 0.05. Results: Thirty-two patients (28%) had T, 23 (20%) had B, and 14 (12%) had BT; 42 (40%) patients were unaffected. Two logistic regression models (outcome = mortality) evaluated BT as T, or BT as B. For BT as T, neither T, B, nor male gender predicted mortality; similarly, for BT as B, neither T, B, nor male gender predicted mortality. Factors associated with higher odds of death included higher Acute Physiology and Chronic Health Evaluation (APACHE) II score (odds ratio [OR], 1.06; 95% confidence interval [CI], 1.00-1.13; p = 0.0045), higher d-dimer concentration (OR, 1.00; 95% CI, 1.00-1.01; p = 0.043), and higher activated partial thromboplastin time (aPTT; OR, 1.09; 95% CI, 1.02-1.16; p = 0.010). Conclusions: Neither T nor B predicted mortality in this prospective cohort of anticoagulated patients with COVID-19 critical illness. These data support continued full-dose heparinoid prophylaxis.

Rotational thromboelastometry in patients with acute respiratory distress syndrome owing to coronavirus disease 2019: Is there a viscoelastic fingerprint and a role for predicting thrombosis?

OBJECTIVES: We evaluated rotational thromboelastometry tracings in 44 critically ill coronavirus disease 2019 patients, to determine whether there is a viscoelastic fingerprint and to test the hypothesis that the diagnosis and prediction of venous thromboembolism would be enhanced by the addition of rotational thromboelastometry testing. RESULTS: Rotational thromboelastometry values reflected an increase in clot strength for the EXTEM, INTEM, and FIBTEM assays beyond the reference range. No hyperfibrinolysis was noted. Fibrinolysis shutdown was present but did not correlate with thrombosis; 32% (14/44) of patients experienced a thrombotic episode. For every 1 mm increase of FIBTEM maximum clot formation, the odds of developing thrombosis increased 20% (95% confidence interval, 0-40%, P = .043), whereas for every 1,000 ng/mL increase in D-dimer, the odds of thrombosis increased by 70% (95% confidence interval, 20%-150%, P = .004), after adjustment for age and sex (AUC 0.96, 95% confidence interval, 0.90-1.00). There was a slight but significant improvement in model performance after adding FIBTEM maximum clot formation and EXTEM clot formation time to D-dimer in a multivariable model (P = .04). CONCLUSIONS: D-dimer concentrations were more predictive of thrombosis in our patient population than any other parameter. Rotational thromboelastometry confirmed the hypercoagulable state of coronavirus disease 2019 intensive care unit patients. FIBTEM maximum clot formation and EXTEM clot formation time increased the predictability for thrombosis compared with only using D-dimer. Rotational thromboelastometry analysis is most useful in augmenting the information provided by the D-dimer concentration for venous thromboembolism risk assessment when the D-dimer concentration is between 1,625 and 6,900 ng/dL, but the enhancement is modest. Fibrinolysis shutdown did not correlate with thrombosis.

Probative Value of the D-Dimer Assay for Diagnosis of Deep Venous Thrombosis in the Coronavirus Disease 2019 Syndrome.

OBJECTIVES: To describe the predictive utility of the D-dimer assay among patients with the coronavirus disease 2019 syndrome for unprovoked lower extremity deep venous thrombosis. DESIGN: Prospective observational study with retrospective data analysis. SETTING: Academic medical center surgical ICU. PATIENTS: Seventy-two intubated patients with critical illness from coronavirus disease 2019. INTERVENTIONS: Therapeutic anticoagulation after imaging diagnosis of the first three deep venous thrombosis cases was confirmed; therapeutic anticoagulation as prophylaxis thereafter to all subsequent ICU admissions. MEASUREMENTS AND MAIN RESULTS: Seventy-two patients with severe coronavirus disease 2019 were screened for deep venous thrombosis after ICU admission with 102 duplex ultrasound examinations, with 12 cases (16.7%) of lower extremity deep venous thrombosis identified. There were no differences between groups with respect to age, renal function, or biomarkers except for D-dimer (median, 12,858 ng/mL [interquartile range, 3,176-30,770 ng/mL] for lower extremity deep venous thrombosis vs 2,087 ng/mL [interquartile range, 638-3,735 ng/mL] for no evidence of deep venous thrombosis; p < 0.0001). Clinical screening tools (Wells score and Dutch Primary Care Rule) had no utility. The C-statistic for D-dimer concentration was 0.874 ± 0.065. At the model-predicted cutoff value of 3,000 ng/mL, sensitivity was 100%, specificity was 51.1%, positive predictive value was 21.8%, and negative predictive value was 100%. CONCLUSIONS: Lower extremity deep venous thrombosis is prevalent in coronavirus disease 2019 disease and can be present on ICU admission. Screening has been recommended in the context of the pro-inflammatory, hypercoagulable background milieu. D-dimer concentrations are elevated in nearly all coronavirus disease 2019 patients, and the test appears reliable for screening for lower extremity deep venous thrombosis at or above a concentration of 3,000 ng/mL (more than 13-fold above the normal range). Full anticoagulation is indicated if the diagnosis is confirmed, and therapeutic anticoagulation should be considered for prophylaxis, as all coronavirus disease 2019 patients are at increased risk.

CD161 defines a transcriptional and functional phenotype across distinct human T cell lineages.

The C-type lectin CD161 is expressed by a large proportion of human T lymphocytes of all lineages, including a population known as mucosal-associated invariant T (MAIT) cells. To understand whether different T cell subsets expressing CD161 have similar properties, we examined these populations in parallel using mass cytometry and mRNA microarray approaches. The analysis identified a conserved CD161++/MAIT cell transcriptional signature enriched in CD161+CD8+ T cells, which can be extended to CD161+ CD4+ and CD161+TCRγδ+ T cells. Furthermore, this led to the identification of a shared innate-like, TCR-independent response to interleukin (IL)-12 plus IL-18 by different CD161-expressing T cell populations. This response was independent of regulation by CD161, which acted as a costimulatory molecule in the context of T cell receptor stimulation. Expression of CD161 hence identifies a transcriptional and functional phenotype, shared across human T lymphocytes and independent of both T cell receptor (TCR) expression and cell lineage.

Downregulation of bone morphogenetic protein 4 expression in coronary arterial endothelial cells: role of shear stress and the cAMP/protein kinase A pathway.

OBJECTIVE: Bone morphogenetic protein 4 (BMP-4) is a transforming growth factor beta family member cytokine that exerts proinflammatory effects on the endothelium and is likely to play a role in atherogenesis. Recent studies suggested that atheroprotective levels of shear stress control endothelial BMP-4 expression; however, the underlying mechanisms remained unknown. METHODS AND RESULTS: We found that shear stress downregulated BMP-4 expression in human and rat coronary arterial endothelial cells (CAECs) as well as in cultured mesenteric arterioles, although it had no effect on the expression of BMP-2, a related cytokine. In human coronary arterial endothelial cells, 8-bromo-cAMP, the adenylate cyclase activator forskolin, or a cAMP-dependent protein kinase (PKA) activator effectively decreased BMP-4 expression, mimicking the effects of shear stress. Indeed, shear stress induced the nuclear translocation of PKA-c, and inhibition of PKA attenuated the effects of shear stress and forskolin on BMP-4 expression. RNA decay assay and BMP-4 promoter-driven luciferase reporter gene assay showed that cAMP regulates BMP-4 expression at the transcriptional level. CONCLUSIONS: Laminar shear stress and the cAMP/PKA pathway are important negative regulators of BMP-4 expression in the vascular endothelium. Because BMP-4 elicits endothelial activation and dysfunction, hypertension, and vascular calcification, inhibition of BMP-4 expression by shear stress and the cAMP/PKA pathway is likely to exert antiatherogenic and vasculoprotective effects.

Bone morphogenetic protein-2 induces proinflammatory endothelial phenotype.

The transforming growth factor-beta superfamily member bone morphogenetic protein-2 (BMP-2) is up-regulated in atherosclerotic arteries; however, its effects on the endothelium are not well characterized. Using microdissected coronary arterial endothelial cells (CAECs) and cultured primary CAECs, we demonstrated endothelial mRNA expression of BMP-2 and BMP-4. The proinflammatory cytokine tumor necrosis factor-alpha and H2O2 significantly increased endothelial expression of BMP-2 but not BMP-4. In organ culture, BMP-2 substantially decreased relaxation of rat carotid arteries to acetylcholine and increased production of reactive oxygen species, events inhibited by pharmacologically blocking protein kinase C (PKC) or NAD(P)H oxidase. BMP-2 activated nuclear factor-kappaB in CAECs, and BMP-2 and BMP-4 substantially increased adhesion of monocytic THP-1 cells, which was reduced by pharmacologically inhibiting p42/44 MAP kinase pathway (also by siRNA down-regulating ERK-1/2) or PKC. Incubation of rat carotid arteries with BMP-2 ex vivo also increased adhesion of mononuclear cells to the endothelium, requiring p42/44 MAP kinase and PKC. Western blotting showed that in CAECs and carotid arteries BMP-2 elicited phosphorylation of p42/44 MAP kinase, which was reduced by blocking MAP kinase kinase and PKC. Collectively, expression of BMP-2 is regulated by proinflammatory stimuli, and increased levels of BMP-2 induce endothelial dysfunction, oxidative stress, and endothelial activation. Thus, the proinflammatory effects of BMP-2 may play a role in vascular pathophysiology.

Regulation of bone morphogenetic protein-2 expression in endothelial cells: role of nuclear factor-kappaB activation by tumor necrosis factor-alpha, H2O2, and high intravascular pressure.

BACKGROUND: Recent studies suggest that bone morphogenetic protein-2 (BMP-2), a transforming growth factor-beta superfamily member cytokine, plays an important role both in vascular development and pathophysiological processes, including endothelial activation that is likely to contribute to the development of coronary atherosclerosis, yet the factors that regulate arterial expression of BMP-2 are completely unknown. We tested the hypothesis that BMP-2 expression in endothelial cells is governed by an H2O2 and nuclear factor (NF)-kappaBeta-dependent pathway that can be activated by both proinflammatory and mechanical stimuli. METHODS AND RESULTS: The proinflammatory cytokine tumor necrosis factor (TNF)-alpha induced NF-kappaBeta activation and elicited significant increases in BMP-2 mRNA and protein in primary coronary arterial endothelial cells and human umbilical vein endothelial cells that were prevented by NF-kappaBeta inhibitors (pyrrolidine dithiocarbamate and SN-50), silencing of p65 (siRNA), or catalase. Administration of H2O2 also elicited NF-kappaBeta activation and BMP-2 induction. In organ culture, exposure of rat arteries to high pressure (160 mm Hg) elicited H2O2 production, nuclear translocation of NF-kappaBeta, and upregulation of BMP-2 expression. Although high pressure upregulated TNF-alpha, it appears that it directly regulates BMP-2 expression, because upregulation of BMP-2 was also observed in vessels of TNF-alpha knockout mice. CONCLUSIONS: Vascular BMP-2 expression can be regulated by H2O2-mediated activation of NF-kappaBeta both by inflammatory stimuli and by high intravascular pressure.