Baseline patient characteristics at study inclusion as well as a description of the further clinical course are summarized in Table 1. Organ failure was not restricted to the lungs, and multiorgan dysfunction was common both at inclusion and during the further clinical course. Surprisingly, global markers of endothelial injury such as Angpt-1 (angiopoietin-1) (control: 29 [26.2–30.9] ng/ml vs. COVID-19: 27.8 [23.4–36.2] ng/ml; P = 0.79) and Angpt-2 (control: 0.655 [0.336–1.113] ng/ml vs. COVID-19: 0.434 [0.035–1.338] ng/ml; P = 0.6) were unchanged in patients with COVID-19. In contrast, marked increases in the soluble form of the sTie2 (Tie2 receptor) (Figure 1A) and in syndecan-1 (Figure 1B)—indicating pathological shedding of transmembrane proteins involved in glycocalyx structure and processing—were observed. The key eGC sheddase Hpa-1 (heparanase-1) and its enzymatic activity were both not significantly increased (data not shown). To the contrary, the Hpa-1 counterpart, the protective Hpa-2 (heparanase-2), was pertinently reduced in all patients with COVID-19 (Figure 1C). Driven by this acquired Hpa-2 deficiency, the Hpa-1:Hpa-2 ratio was higher in patients with COVID-19 (P = 0.012; data not shown). Together, this indicates that critically ill patients with COVID-19 suffer from an acquired Hpa-2 deficiency that can contribute to the degradation of the eGC, maybe even before classical endothelial activation and injury. Next, eGC structure was analyzed in humans, employing sublingual sidestream darkfield (SDF) imaging. We quantified the size of the individual patients’ eGC using an indirect surrogate termed the perfused boundary region and found a decrease of perfused boundary region, indicating reduced eGC thickness in patients with COVID-19. To demonstrate that the deficiency of Hpa-2 is mechanistically involved in the degradation of the eGC, we used a microfluidic chamber with cultured endothelial cells (ECs) under flow that synthesize an intact and stable eGC under in vitro conditions. After stimulation with COVID-19 or control serum, the eGC was visualized by confocal microscopy followed by computerized three-dimensional reconstruction. Its thickness was then quantified by analyzing the heparan sulfate (HS)-positive area. We found that stimulation with COVID-19 was sufficient to severely damage the eGC (Figure 1E). The HS-positive area was reduced by 34% (control: 6.1 ± 0.9% vs. COVID-19: 4 ± 0.4% P < 0.001). Consistent with our observation in patients, we found that the transcription of Hpa-2 in COVID-19–stimulated ECs was significantly reduced after 6 hours (0.63 ± 0.02 relative expression to control; P = 0.003). Of note, transgenic overexpression of Hpa-2 in a lentivirus-transduced EC line was sufficient to reverse this phenotype, as the HS area in COVID-19 serum–treated lenti-control cells was 1.9 ± 0.6% but was 4.2 ± 1.2% in lenti-Hpa-2–overexpressing cells (P < 0.001). In other words, if ECs overexpress Hpa-2, the serum of patients with COVID-19 cannot degrade the eGC anymore.