Dynamic angiopoietin-2 assessment predicts survival and chronic course in hospitalized patients with COVID-19

Abstract

This study examined the association between dynamic angiopoietin-2 assessment and COVID-19 short- and long-term clinical course. We included consecutive hospitalized patients from 1 February to 31 May 2020 with laboratory-confirmed COVID-19 from 2 Italian tertiary referral centers (derivation cohort, n = 187 patients; validation cohort, n = 62 patients). Serum biomarker levels were measured by sandwich enzyme-linked immunosorbent assay. Lung tissue from 9 patients was stained for angiopoietin-2, Tie2, CD68, and CD34. Cox model was used to identify risk factors for mortality and nonresolving pulmonary condition. Area under the receiver operating characteristic curve (AUROC) was used to assess the accuracy of 3- and 10-day angiopoietin-2 for in-hospital mortality and nonresolving pulmonary condition, respectively. Three-day angiopoietin-2 increase of at least twofold from baseline was significantly associated with in-hospital mortality by multivariate analysis (hazard ratio [HR], 6.69; 95% confidence interval [CI], 1.85-24.19; P = .004) with AUROC = 0.845 (95% CI, 0.725-0.940). Ten-day angiopoietin-2 of at least twofold from baseline was instead significantly associated with nonresolving pulmonary condition by multivariate analysis (HR, 5.33; 95% CI, 1.34-11.77; P ≤ .0001) with AUROC = 0.969 (95% CI, 0.919-1.000). Patients with persistent elevation of 10-day angiopoietin-2 levels showed severe reticular interstitial thickening and fibrous changes on follow-up computed tomography scans. Angiopoietin-2 and Tie2 were diffusely colocalized in small-vessel endothelia and alveolar new vessels and macrophages. Angiopoietin-2 course is strongly associated with COVID-19 in-hospital mortality and nonresolving pulmonary condition. Angiopoietin-2 may be an early and useful predictor of COVID-19 clinical course, and it could be a relevant part of disease pathogenesis. Angiopoietin-2 blockade may be a COVID-19 treatment option.

Graphical abstract

Figure 1.

Relationship between angiopoietin-2 and IL-6 and in-hospital mortality in the derivation and validation cohorts in patients with COVID-19. Day 1 (blue columns) and day 3 (red columns) angiopoietin-2 levels according to survivors and nonsurvivors in the derivation (A) and validation (B) cohorts. Day 1 (blue columns) and day 3 (red columns) IL-6 levels according to survivors and nonsurvivors in the derivation (C) and validation (D) cohorts. In-hospital mortality according to 3-day course of angiopoietin-2 in the derivation (E) and validation (F) cohorts. In-hospital mortality according to 3-day course of IL-6 in the derivation (G) and validation (H) cohorts. Red line, increase of at least twofold from baseline; blue line, no increase.

Figure 2.

ROC curves of angiopoietin-2 and IL-6 courses for in-hospital mortality and nonresolving pulmonary condition in patients with COVID-19. Three-day course for in-hospital mortality of angiopoietin-2 in the (A) derivation and (B) validation cohorts. Three-day course for in-hospital mortality of IL-6 in the (C) derivation and (D) validation cohorts. Ten-day course for nonresolving pulmonary condition of angiopoietin-2 in the (E) derivation and (F) validation cohorts. Ten-day course for nonresolving pulmonary condition of IL-6 in the (G) derivation and (H) validation cohorts.

Figure 3.

Outcomes according to 10-day angiopoietin-2 and IL-6 course. Length of hospitalization according to 10-day angiopoietin-2 (A) and IL-6 (B) courses. PaO₂/FiO₂ ratio levels at 1 month according to 10-day angopoietin-2 (C) and IL-6 (D) courses. Cumulative incidence of nonresolving pulmonary condition according to 10-day angiopoietin-2 course in derivation (E) and validation (F) cohorts. Cumulative incidence of nonresolving pulmonary condition according to 10-day IL-6 course in derivation (G) and validation (H) cohorts. Red line, increase of at least twofold from baseline; blue line, no increase.

Figure 4.

Immunohistochemical analysis of autoptic lung tissue from a patient with COVID-19. (A) Immunostaining for angiopoietin-2 (20×): immunohistochemistry shows immunoreactivity in the endothelia, pneumocytes, and macrophages. Scale bar, 6.7 μm. (B) Immunostaining for Tie2 (20×): the same field as in Figure 1. Tie2 showed the same immunoreactivity observed for angiopoietin-2 immunoreaction. (C) Hematoxylin and eosin staining (20×): diffuse alveolar damage in COVID-19, interstitial inflammation, and dilated capillaries in the alveolar wall. The alveolar spaces show fibrin and macrophages. (D-F) The brown DAB signal for angiopoietin-2 shown in panel A was converted into a blue color and that for Tie2 shown in panel B was converted into a red color. The images in panels D and E were merged, resulting in a violet color, where the angiopoietin-2 and Tie2 colocalize. (G-I) Higher magnification (63×) of the inset in panel A highlights angiopoietin2 and tie2 immunoreactivity both in endothelia (red arrowheads) and in pneumocytes (yellow arrows). Scale bar, 1.7 μm. (J-L) The brown DAB signal for angiopoietin-2 shown in panel G was converted into a blue color and that for Tie2 shown in panel H was converted into a red color. The images in panels J and K were merged, resulting in a violet color, where the angiopoietin-2 and Tie2 colocalize.