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Objectives: Angiotensin-converting enzyme 2 (ACE2) represents the primary receptor for SARS-CoV-2 to enter endothelial cells. Here we investigated circulating ACE2 activity to predict the severity and mortality of COVID-19. Methods: Serum ACE2 activity was measured in COVID-19 (110 critically ill and 66 severely ill subjects at hospital admission and 106 follow-up samples) and in 32 non-COVID-19 severe sepsis patients. Associations between ACE2, inflammation-dependent biomarkers, pre-existing comorbidities, and clinical outcomes were studied. Results: Initial ACE2 activity was significantly higher in critically ill COVID-19 patients (54.4 [36.7-90.8] mU/L) than in severe COVID-19 (34.5 [25.2-48.7] mU/L; P

Following an intraventricular hemorrhage (IVH), red blood cell lysis and hemoglobin (Hb) oxidation with the release of heme can cause sterile neuroinflammation. In this study, we measured Hb derivates and cellular adhesion molecules ICAM-1 and VCAM-1 with cell-free miRNAs in cerebrospinal fluid (CSF) samples obtained from Grade-III and Grade-IV preterm IVH infants (IVH-III and IVH-IV, respectively) at multiple time points between days 0–60 after the onset of IVH. Furthermore, human choroid plexus epithelial cells (HCPEpiCs) were incubated with IVH and non-IVH CSF (10 v/v %) for 24 h in vitro to investigate the IVH-induced inflammatory response that was investigated via: (i) HMOX1, IL8, VCAM1, and ICAM1 mRNAs as well as miR-155, miR-223, and miR-181b levels by RT-qPCR, (ii) nuclear translocation of the NF-κB p65 subunit by fluorescence microscopy, and (iii) reactive oxygen species (ROS) measurement. We found a time-dependent alteration of heme, IL-8, and adhesion molecules which revealed a prolonged elevation in IVH-IV vs. IVH-III with higher miR-155 and miR-181b expression at days 41–60. Exposure of HCPEpiCs to IVH CSF samples induced HMOX1, IL8, and ICAM1 mRNA levels along with increased ROS production via the NF-κB pathway activation but without cell death, as confirmed by the cell viability assay. Additionally, the enhanced intracellular miR-155 level was accompanied by lower miR-223 and miR-181b expression in HCPEpiCs after CSF treatment. Overall, choroid plexus epithelial cells exhibit an abnormal cell phenotype after interaction with pro-inflammatory CSF of IVH origin which may contribute to the development of later clinical complications in preterm IVH.

Highlights • Upregulated expression of ACE2 was recently reported in bronchoalveolar lavage fluid samples from COVID-19 patients. • We have first analyzed serum ACE2 activity in COVID-19 that increased about 40-fold over the normal range in the presence of 2-3-fold higher levels of endothelium biomarkers. • Soluble E-selectin followed the clinical status of our patient similarly to ferritin and IL-6 levels. • Based on the distinct time course of circulating ACE2, its dramatic rise may act as an endogenous nonspecific protective mechanism against SARS-CoV-2 infection that preceded the recovery of the patient., Endothelial cells express surface angiotensin converting enzyme 2 (ACE2), the main receptor for SARS-CoV-2 that promotes the infection of endothelial cells showing activation and damage. Bronchoalveolar lavage fluid from COVID-19 subjects showed a critical imbalance in the renin-angiotensin-aldosterone system with upregulated expression of ACE2. Recently, intravenous recombinant ACE2 was reported as an effective therapy in severe COVID-19 by blocking the viral entry to target cells. Here we present a case of a critically ill COVID-19 patient with acute respiratory distress syndrome where circulating ACE2 was first measured to monitor disease prognosis. ACE2 activity increased about 40-fold over the normal range and showed a distinct time course compared to 2-3-fold higher levels of endothelium biomarkers. Although the level of soluble E-selectin followed the clinical status of our patient similarly to ferritin and IL-6 levels, the dramatic rise in serum ACE2 activity may act as an endogenous nonspecific protective mechanism against SARS-CoV-2 infection that preceded the recovery of our patient.

Decreased human epididymis protein 4 (HE4) plasma levels were reported in cystic fibrosis (CF) patients under CFTR potentiator ivacaftor therapy, which inversely correlated with lung function improvement. In this study, we investigated whether HE4 expression was affected via modulation of CFTR function in CF bronchial epithelial (CFBE) cells in vitro. HE4 protein levels were measured in the supernatants of CFBE 41o− cells expressing F508del-CFTR or wild-type CFTR (wt-CFTR) after administration of lumacaftor/ivacaftor or tezacaftor/ivacaftor, while HE4 expression in CFBE 41o− cells were also analyzed following application of adenylate cyclase activators Forskolin/IBMX or CFTRinh172. The effect of all of these compounds on CFTR function was monitored by the whole-cell patch-clamp technique. Induced HE4 expression was studied with interleukin-6 (IL-6) in F508del-CFTR CFBE 41o− cells under TNF-α stimulation for 1 h up to 1 week in duration. In parallel, plasma HE4 was determined in CF subjects homozygous for p.Phe508del-CFTR mutation receiving lumacaftor/ivacaftor (Orkambi®) therapy. NF-κB-mediated signaling was observed via the nuclear translocation of p65 subunit by fluorescence microscopy together with the analysis of IL-6 expression by an immunoassay. In addition, HE4 expression was examined after NF-κB pathway inhibitor BAY 11-7082 treatment with or without CFTR modulators. CFTR modulators partially restored the activity of F508del-CFTR and reduced HE4 concentration was found in F508del-CFTR CFBE 41o− cells that was close to what we observed in CFBE 41o− cells with wt-CFTR. These data were in agreement with decreased plasma HE4 concentrations in CF patients treated with Orkambi®. Furthermore, CFTR inhibitor induced elevated HE4 levels, while CFTR activator Forskolin/IBMX downregulated HE4 in the cell cultures and these effects were more pronounced in the presence of CFTR modulators. Higher activation level of baseline and TNF-α stimulated NF-κB pathway was detected in F508del-CFTR vs. wt-CFTR CFBE 41o− cells that was substantially reduced by CFTR modulators based on lower p65 nuclear positivity and IL-6 levels. Finally, HE4 expression was upregulated by TNF-α with elevated IL-6, and both protein levels were suppressed by combined administration of NF-κB pathway inhibitor and CFTR modulators in CFBE 41o− cells. In conclusion, CFTR dysfunction contributes to abnormal HE4 expression via NF-κB in CF.

Intraventricular hemorrhage (IVH) is a frequent complication of prematurity that is associated with high neonatal mortality and morbidity. IVH is accompanied by red blood cell (RBC) lysis, hemoglobin (Hb) oxidation, and sterile inflammation. Here we investigated whether extracellular Hb, metHb, ferrylHb, and heme contribute to the inflammatory response after IVH. We collected cerebrospinal fluid (CSF) (n = 20) from premature infants with grade III IVH at different time points after the onset of IVH. Levels of Hb, metHb, total heme, and free heme were the highest in CSF samples obtained between days 0 and 20 after the onset of IVH and were mostly non-detectable in CSF collected between days 41 and 60 of post-IVH. Besides Hb monomers, we detected cross-linked Hb dimers and tetramers in post-IVH CSF samples obtained in days 0–20 and 21–40, but only Hb tetramers were present in CSF samples obtained after 41–60 days. Vascular cell adhesion molecule-1 (VCAM-1) and interleukin-8 (IL-8) levels were higher in CSF samples obtained between days 0 and 20 than in CSF collected between days 41 and 60 of post-IVH. Concentrations of VCAM-1, intercellular adhesion molecule-1 (ICAM-1), and IL-8 strongly correlated with total heme levels in CSF. Applying the identified heme sources on human brain microvascular endothelial cells revealed that Hb oxidation products and free heme contribute to the inflammatory response. We concluded that RBC lysis, Hb oxidation, and heme release are important components of the inflammatory response in IVH. Pharmacological interventions targeting cell-free Hb, Hb oxidation products, and free heme could have potential to limit the neuroinflammatory response following IVH.

Although the clinical outcomes of cystic fibrosis (CF) have been markedly improved through the recent implementation of novel CF transmembrane conductance regulator (CFTR) modulator drugs, robust and reliable biomarkers are still demanded for the early detection of CF lung disease progression, monitoring treatment efficacy and predicting life-threatening clinical complications. Thus, there is an unmet need to identify and validate novel, ideally blood based biomarkers with strong correlations to the severity of CF lung disease, which represents a major contribution to overall CF morbidity and mortality. In this review, we aim to summarize the utility of thus far studied blood-, sputum- and bronchoalveolar lavage (BAL)-based biomarkers to evaluate inflammatory conditions in the lung and to follow treatment efficacy in CF. Measurements of sweat chloride concentrations and the spirometric parameter FEV1 are currently utilized to monitor CFTR function and the effect of various CF therapies. Nonetheless, both have inherent pitfalls and limitations, thus routinely analyzed biomarkers in blood, sputum or BAL samples are required as surrogates for lung disorders. Recent discovery of new protein (e.g. HE4) and RNA-based biomarkers, such as microRNAs may offer a higher efficacy, which in aggregate may be valuable to evaluate disease prognosis and to substantiate CF drug efficacy.

Sepsis is a life-threating condition with dysregulated systemic host response to microbial pathogens leading to disproportionate inflammatory response and multi-organ failure. Various biomarkers are available for the diagnosis and prognosis of sepsis; however, these laboratory parameters may show limitations in these severe clinical conditions. MicroRNAs (miRNA) are single-stranded non-coding RNAs with the function of post-transcriptional gene silencing. They normally control numerous intracellular events, such as signaling cascade downstream of Toll-like receptors (TLRs) to avoid excessive inflammation after infection. In contrast, abnormal miRNA expression contributes to the development of sepsis correlating with its clinical features and outcomes. Based on recent clinical studies altered levels of circulating miRNAs can act as potential diagnostic and prognostic biomarkers in sepsis. In this review, we summarized the available data about TLR-mediated inflammatory signaling with its intracellular response in immune cells and platelets upon sepsis, which are, at least in part, under the regulation of miRNAs. Furthermore, the role of circulating miRNAs is also described as potential laboratory biomarkers in sepsis.