Urinary single-cell sequencing captures kidney injury and repair processes in human acute kidney injury.

Acute kidney injury (AKI) is a major health issue, the outcome of which depends primarily on damage and reparative processes of tubular epithelial cells. Mechanisms underlying AKI remain incompletely understood, specific therapies are lacking and monitoring the course of AKI in clinical routine is confined to measuring urine output and plasma levels of filtration markers. Here we demonstrate feasibility and potential of a novel approach to assess the cellular and molecular dynamics of AKI by establishing a robust urine-to-single cell RNA sequencing (scRNAseq) pipeline for excreted kidney cells via flow cytometry sorting. We analyzed 42,608 single cell transcriptomes of 40 urine samples from 32 patients with AKI and compared our data with reference material from human AKI post-mortem biopsies and published mouse data. We demonstrate that tubular epithelial cells transcriptomes mirror kidney pathology and reflect distinct injury and repair processes, including oxidative stress, inflammation, and tissue rearrangement. We also describe an AKI-specific abundant urinary excretion of adaptive progenitor-like cells. Thus, single cell transcriptomics of kidney cells excreted in urine provides noninvasive, unprecedented insight into cellular processes underlying AKI, thereby opening novel opportunities for target identification, AKI sub-categorization, and monitoring of natural disease course and interventions.

Assessment of Renal Transplant Perfusion by Contrast-Enhanced Ultrasound after Switch from Calcineurin Inhibitor to Belatacept: A Pilot Study.

Calcineurin inhibitors (CNIs) have improved short-term kidney allograft survival but are nephrotoxic and vasoconstrictive. Vasoconstriction is potentially reversible after switching from CNIs to belatacept. The kidney allograft shows optimal requirements for dynamic perfusion imaging using contrast-enhanced ultrasound (CEUS). We performed standardized CEUS in patients after switching from CNIs to belatacept for clinical indication to study the suitability of CEUS, in order to assess the effects of CNI cessation on kidney allograft perfusion. Eleven kidney transplant patients were enrolled from February 2020 until November 2020. Demographic, clinical, and laboratory parameters, as well as perfusion imaging, were assessed at baseline and 6 months after switching immunosuppression. Quantification of perfusion imaging on CEUS was performed using a post-processing software tool on uncompressed DICOM cine loops. After CNI cessation, estimated glomerular filtration rate increased by 4.8 mL/min/1.73 m2 (16%). Despite good quality of fit and comparable regions of interest in baseline and follow-up CEUS examinations, quantification of perfusion imaging showed a slightly improved cortical perfusion without reaching statistical significance after CNI cessation. This is the first study that systematically investigates the suitability of CEUS to detect changes of microvascular perfusion in kidney transplant recipients in vivo. No significant differences could be detected in perfusion measurements before and after CNI cessation.

Efficacy and complications of regional citrate anticoagulation during continuous renal replacement therapy in critically ill patients with COVID-19.

BACKGROUND: We compared filter survival and citrate-induced complications during continuous renal replacement therapy (CRRT) with regional citrate anticoagulation (RCA) in COVID-19 and Non-COVID-19 patients. METHODS: In this retrospective study we included all consecutive adult patients (n = 97) treated with RCA-CRRT. Efficacy and complications of RCA-CRRT were compared between COVID-19 and Non-COVID-19 patients. RESULTS: Mean filter run-time was significantly higher in COVID-19 patients compared to Non-COVID-19 patients (68.4 (95%CI 67.0-69.9) vs. 65.2 (95%CI 63.2-67.2) hours, respectively; log-rank 0.014). COVID-19 patients showed significantly higher activated partial thromboplastin time (aPTT) throughout the CRRT due to intensified systemic anticoagulation compared to Non-COVID-19 patients (54 (IQR 45-61) vs. 47 (IQR 41-58) seconds, respectively; p < 0.001). A significantly higher incidence of metabolic alkalosis, hypercalcemia and hypernatremia, consistent with reduced filter patency and citrate overload, was observed in COVID-19 patients compared to Non-COVID-19 patients (19.1% vs. 12.7%, respectively; p = 0.04). These metabolic disarrangements were resistant to per-protocol adjustments and disappeared after replacement of the CRRT-filter. CONCLUSIONS: RCA-CRRT in COVID-19 patients with intensified systemic anticoagulation provides an adequate filter lifespan. However, close monitoring of the acid-base balance appears warranted, as these patients tend to develop reduced filter patency leading to a higher incidence of citrate overload and metabolic disturbances. TRIAL REGISTRATION (LOCAL AUTHORITY): EA1/285/20 (Ethikkommission der Charité - Universitätsmedizin Berlin); date of registration 08.10.2020.

Macrophage activation syndrome in a patient with adult-onset Still's disease following first COVID-19 vaccination with BNT162b2.

BACKGROUND: Adult-onset Still's disease (AOSD) is an autoinflammatory multi-systemic syndrome. Macrophage activation syndrome (MAS) is a potentially life-threatening complication of AOSD with a mortality rate of 10-20%. Especially viral infection is thought to be a common trigger for development of MAS. On the other hand, the occurrence of MAS following vaccinations is extremely rare and has been described in a few cases after measles or influenza vaccinations and more recently after ChAdOx1 nCoV-19 (COVID-19 viral vector vaccine, Oxford-AZ). CASE PRESENTATION: We report the case of a twenty-year-old female with adult-onset Still's disease (AOSD), who developed a MAS six days after receiving her first COVID-19 vaccine dose of BNT162b2 (mRNA vaccine, BioNTech/Pfizer) with ferritin levels of 136,680 µg/l (ref.: 13-150 µg/l). CONCLUSIONS: To the best of our knowledge, this is the first case report of development of MAS in a patient with preexisting AOSD after vaccination in general, and SARS-CoV-2 vaccination in particular. The new mRNA vaccines have generally shown a reassuring safety profile, but it has been shown that nucleic acids in general, including mRNA can act as pathogen-associated molecular patterns that activate toll-like receptors with extensive production of pro-inflammatory cytokines and further activation of immune cells. Proving an interferon 1 response in our patient directly after vaccination, we think that in this particular case the vaccination might have acted as trigger for the development of MAS. Even if it remains difficult to establish causality in the case of rare adverse events, especially in patients with autoimmune or autoinflammatory conditions, these complications are important to monitor and register, but do not at all diminish the overwhelming positive benefit-risk ratio of licensed COVID-19 vaccines.

Point-of-care lung ultrasound in COVID-19 patients: inter- and intra-observer agreement in a prospective observational study.

With an urgent need for bedside imaging of coronavirus disease 2019 (COVID-19), this study's main goal was to assess inter- and intraobserver agreement in lung ultrasound (LUS) of COVID-19 patients. In this single-center study we prospectively acquired and evaluated 100 recorded ten-second cine-loops in confirmed COVID-19 intensive care unit (ICU) patients. All loops were rated by ten observers with different subspeciality backgrounds for four times by each observer (400 loops overall) in a random sequence using a web-based rating tool. We analyzed inter- and intraobserver variability for specific pathologies and a semiquantitative LUS score. Interobserver agreement for both, identification of specific pathologies and assignment of LUS scores was fair to moderate (e.g., LUS score 1 Fleiss' κ = 0.27; subpleural consolidations Fleiss' κ = 0.59). Intraobserver agreement was mostly moderate to substantial with generally higher agreement for more distinct findings (e.g., lowest LUS score 0 vs. highest LUS score 3 (median Fleiss' κ = 0.71 vs. 0.79) or air bronchograms (median Fleiss' κ = 0.72)). Intraobserver consistency was relatively low for intermediate LUS scores (e.g. LUS Score 1 median Fleiss' κ = 0.52). We therefore conclude that more distinct LUS findings (e.g., air bronchograms, subpleural consolidations) may be more suitable for disease monitoring, especially with more than one investigator and that training material used for LUS in point-of-care ultrasound (POCUS) should pay refined attention to areas such as B-line quantification and differentiation of intermediate LUS scores.

Measuring myocardial extracellular volume of the right ventricle in patients with congenital heart disease.

The right ventricle´s (RV) characteristics-thin walls and trabeculation-make it challenging to evaluate extracellular volume (ECV). We aimed to assess the feasibility of RV ECV measurements in congenital heart disease (CHD), and to introduce a novel ECV analysis tool. Patients (n = 39) and healthy controls (n = 17) underwent cardiovascular magnetic resonance T1 mapping in midventricular short axis (SAX) and transverse orientation (TRANS). Regions of interest (ROIs) were evaluated with regard to image quality and maximum RV wall thickness per ROI in pixels. ECV from plane ROIs was compared with values obtained with a custom-made tool that derives the mean T1 values from a "line of interest" (LOI) centered in the RV wall. In CHD, average image quality was good (no artifacts in the RV, good contrast between blood/myocardium), and RV wall thickness was 1-2 pixels. RV ECV was not quantifiable in 4/39 patients due to insufficient contrast or wall thickness < 1 pixel. RV myocardium tended to be more clearly delineated in SAX than TRANS. ECV from ROIs and corresponding LOIs correlated strongly in both directions (SAX/TRANS: r = 0.97/0.87, p < 0.001, respectively). In conclusion, RV ECV can be assessed if image quality allows sufficient distinction between myocardium and blood, and RV wall thickness per ROI is ≥ 1 pixel. T1 maps in SAX are recommended for RV ECV analysis. LOI application simplifies RV ECV measurements.

Cardiac T1 mapping in congenital heart disease: bolus vs. infusion protocols for measurements of myocardial extracellular volume fraction.

Myocardial extracellular volume fraction (ECV) reflecting diffuse myocardial fibrosis can be measured with T1 mapping cardiovascular magnetic resonance (CMR) before and after the application of a gadolinium-based extracellular contrast agent. The equilibrium between blood and myocardium contrast concentration required for ECV measurements can be obtained with a primed contrast infusion (equilibrium contrast-CMR). We hypothesized that equilibrium can also be achieved with a single contrast bolus to accurately measure diffuse myocardial fibrosis in patients with congenital heart disease (CHD). Healthy controls (n = 17; median age 24.0 years) and patients with CHD (n = 19; 25.0 years) were prospectively enrolled. Using modified Look-Locker inversion recovery T1 mapping before, 15 min after bolus injection, and during constant infusion of gadolinium-DOTA, T1 values were obtained for blood pool and myocardium of the left ventricle (LV), the interventricular septum (IVS), and the right ventricle (RV) in a single midventricular plane in short axis or in transverse orientation. ECV of LV, IVS and RV by bolus-only and bolus-infusion correlated significantly in CHD patients (r = 0.94, 0.95, and 0.74; p < 0.01, respectively) and healthy controls (r = 0.96, 0.89, and 0.64; p < 0.05, respectively). Bland-Altman plots revealed no significant bias between the techniques for any of the analyzed regions. ECV of LV and RV myocardium measured by bolus-only T1 mapping agrees well with bolus-infusion measurements in patients with CHD. The use of a bolus-only approach facilitates the integration of ECV measurements into existing CMR imaging protocols, allowing for assessment of diffuse myocardial fibrosis in CHD in clinical routine.

Alterations in creatine metabolism observed in experimental autoimmune myocarditis using ex vivo proton magic angle spinning MRS.

Experimental autoimmune myocarditis (EAM) in rodents is an accepted model of myocarditis and dilated cardiomyopathy (DCM). Altered metabolism is thought to play an important role in the pathogenesis of DCM and heart failure (HF). Study of the metabolism may provide new diagnostic information and insights into the mechanisms of myocarditis and HF. Proton MRS ((1)H-MRS) has not yet been used to study the changes occurring in myocarditis and subsequent HF. We aimed to explore the changes in creatine metabolism using this model and compare them with the findings in healthy animals. Myocardial function of male young Lewis rats with EAM was quantified by performing left ventricular ejection fraction (LVEF) analysis in short-axis cine images throughout the whole heart. Inflammatory cellular infiltrate was assessed by immunohistochemistry. Myocardial tissue was analyzed using ex vivo proton magic angle spinning MRS ((1)H-MAS-MRS). Myocarditis was confirmed histologically by the presence of an inflammatory cellular infiltrate and CD68 positive staining. A significant increase in the metabolic ratio of Tau/tCr (taurine/total creatine) obtained by (1)H-MAS-MRS was observed in myocarditis compared with healthy controls (21 d acute EAM, 4.38 (±0.23); 21 d control, 2.84 (±0.08); 35 d chronic EAM, 4.47 (±0.83); 35 d control, 2.59 (±0.38); P < 0.001). LVEF was reduced in diseased animals (EAM, 55.2% (±11.3%); control, 72.6% (±3.8%); P < 0.01) and correlated with Tau/tCr ratio (R = 0.937, P < 0.001). Metabolic alterations occur acutely with the development of myocarditis. Myocardial Tau/tCr ratio as detected by (1)H-MRS correlates with LVEF and is able to differentiate between healthy myocardium and myocardium from rats with EAM.