SARS-CoV-2 infection and persistence in the human body and brain at autopsy.

Coronavirus disease 2019 (COVID-19) is known to cause multi-organ dysfunction1-3 during acute infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with some patients experiencing prolonged symptoms, termed post-acute sequelae of SARS-CoV-2 (refs. 4,5). However, the burden of infection outside the respiratory tract and time to viral clearance are not well characterized, particularly in the brain3,6-14. Here we carried out complete autopsies on 44 patients who died with COVID-19, with extensive sampling of the central nervous system in 11 of these patients, to map and quantify the distribution, replication and cell-type specificity of SARS-CoV-2 across the human body, including the brain, from acute infection to more than seven months following symptom onset. We show that SARS-CoV-2 is widely distributed, predominantly among patients who died with severe COVID-19, and that virus replication is present in multiple respiratory and non-respiratory tissues, including the brain, early in infection. Further, we detected persistent SARS-CoV-2 RNA in multiple anatomic sites, including throughout the brain, as late as 230 days following symptom onset in one case. Despite extensive distribution of SARS-CoV-2 RNA throughout the body, we observed little evidence of inflammation or direct viral cytopathology outside the respiratory tract. Our data indicate that in some patients SARS-CoV-2 can cause systemic infection and persist in the body for months.

Genetically Modified Porcine-to-Human Cardiac Xenotransplantation.

A 57-year-old man with nonischemic cardiomyopathy who was dependent on venoarterial extracorporeal membrane oxygenation (ECMO) and was not a candidate for standard therapeutics, including a traditional allograft, received a heart from a genetically modified pig source animal that had 10 individual gene edits. Immunosuppression was based on CD40 blockade. The patient was weaned from ECMO, and the xenograft functioned normally without apparent rejection. Sudden diastolic thickening and failure of the xenograft occurred on day 49 after transplantation, and life support was withdrawn on day 60. On autopsy, the xenograft was found to be edematous, having nearly doubled in weight. Histologic examination revealed scattered myocyte necrosis, interstitial edema, and red-cell extravasation, without evidence of microvascular thrombosis - findings that were not consistent with typical rejection. Studies are under way to identify the mechanisms responsible for these changes. (Funded by the University of Maryland Medical Center and School of Medicine.).

Graft dysfunction in compassionate use of genetically engineered pig-to-human cardiac xenotransplantation: a case report.

BACKGROUND: A genetically engineered pig cardiac xenotransplantation was done on Jan 7, 2022, in a non-ambulatory male patient, aged 57 years, with end-stage heart failure, and on veno-arterial extracorporeal membrane oxygenation support, who was ineligible for an allograft. This report details our current understanding of factors important to the xenotransplantation outcome. METHODS: Physiological and biochemical parameters critical for the care of all heart transplant recipients were collected in extensive clinical monitoring in an intensive care unit. To ascertain the cause of xenograft dysfunction, we did extensive immunological and histopathological studies, including electron microscopy and quantification of porcine cytomegalovirus or porcine roseolovirus (PCMV/PRV) in the xenograft, recipient cells, and tissue by DNA PCR and RNA transcription. We performed intravenous immunoglobulin (IVIG) binding to donor cells and single-cell RNA sequencing of peripheral blood mononuclear cells. FINDINGS: After successful xenotransplantation, the graft functioned well on echocardiography and sustained cardiovascular and other organ systems functions until postoperative day 47 when diastolic heart failure occurred. At postoperative day 50, the endomyocardial biopsy revealed damaged capillaries with interstitial oedema, red cell extravasation, rare thrombotic microangiopathy, and complement deposition. Increased anti-pig xenoantibodies, mainly IgG, were detected after IVIG administration for hypogammaglobulinaemia and during the first plasma exchange. Endomyocardial biopsy on postoperative day 56 showed fibrotic changes consistent with progressive myocardial stiffness. Microbial cell-free DNA testing indicated increasing titres of PCMV/PRV cell-free DNA. Post-mortem single-cell RNA sequencing showed overlapping causes. INTERPRETATION: Hyperacute rejection was avoided. We identified potential mediators of the observed endothelial injury. First, widespread endothelial injury indicates antibody-mediated rejection. Second, IVIG bound strongly to donor endothelium, possibly causing immune activation. Finally, reactivation and replication of latent PCMV/PRV in the xenograft possibly initiated a damaging inflammatory response. The findings point to specific measures to improve xenotransplant outcomes in the future. FUNDING: The University of Maryland School of Medicine, and the University of Maryland Medical Center.

Histopathology and SARS-CoV-2 Cellular Localization in Eye Tissues of COVID-19 Autopsies.

Ophthalmic manifestations and tissue tropism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been reported in association with coronavirus disease 2019 (COVID-19), but the pathology and cellular localization of SARS-CoV-2 are not well characterized. The objective of this study was to evaluate macroscopic and microscopic changes and investigate cellular localization of SARS-CoV-2 across ocular tissues at autopsy. Ocular tissues were obtained from 25 patients with COVID-19 at autopsy. SARS-CoV-2 nucleocapsid gene RNA was previously quantified by droplet digital PCR from one eye. Herein, contralateral eyes from 21 patients were fixed in formalin and subject to histopathologic examination. Sections of the droplet digital PCR-positive eyes from four other patients were evaluated by in situ hybridization to determine the cellular localization of SARS-CoV-2 spike gene RNA. Histopathologic abnormalities, including cytoid bodies, vascular changes, and retinal edema, with minimal or no inflammation in ocular tissues were observed in all 21 cases evaluated. In situ hybridization localized SARS-CoV-2 RNA to neuronal cells of the retinal inner and outer layers, ganglion cells, corneal epithelia, scleral fibroblasts, and oligodendrocytes of the optic nerve. In conclusion, a range of common histopathologic alterations were identified within ocular tissue, and SARS-CoV-2 RNA was localized to multiple cell types. Further studies will be required to determine whether the alterations observed were caused by SARS-CoV-2 infection, the host immune response, and/or preexisting comorbidities.

Successful lung transplantation using an allograft from a COVID-19-recovered donor: a potential role for subgenomic RNA to guide organ utilization.

Although the risk of SARS-CoV-2 transmission through lung transplantation from acutely infected donors is high, the risks of virus transmission and long-term lung allograft outcomes are not as well described when using pulmonary organs from COVID-19-recovered donors. We describe successful lung transplantation for a COVID-19-related lung injury using lungs from a COVID-19-recovered donor who was retrospectively found to have detectable genomic SARS-CoV-2 RNA in the lung tissue by multiple highly sensitive assays. However, SARS-CoV-2 subgenomic RNA (sgRNA), a marker of viral replication, was not detectable in the donor respiratory tissues. One year after lung transplantation, the recipient has a good functional status, walking 1 mile several times per week without the need for supplemental oxygen and without any evidence of donor-derived SARS-CoV-2 transmission. Our findings highlight the limitations of current clinical laboratory diagnostic assays in detecting the persistence of SARS-CoV-2 RNA in the lung tissue. The persistence of SARS-CoV-2 RNA in the donor tissue did not appear to represent active viral replication via sgRNA testing and, most importantly, did not negatively impact the allograft outcome in the first year after lung transplantation. sgRNA is easily performed and may be a useful assay for assessing viral infectivity in organs from donors with a recent infection.

Functional recovery in a cohort of ECMO and non-ECMO acute respiratory distress syndrome survivors.

BACKGROUND: The mortality benefit of VV-ECMO in ARDS has been extensively studied, but the impact on long-term functional outcomes of survivors is poorly defined. We aimed to assess the association between ECMO and functional outcomes in a contemporaneous cohort of survivors of ARDS. METHODS: Multicenter retrospective cohort study of ARDS survivors who presented to follow-up clinic. The primary outcome was FVC% predicted. Univariate and multivariate regression models were used to evaluate the impact of ECMO on the primary outcome. RESULTS: This study enrolled 110 survivors of ARDS, 34 of whom were managed using ECMO. The ECMO cohort was younger (35 [28, 50] vs. 51 [44, 61] years old, p < 0.01), less likely to have COVID-19 (58% vs. 96%, p < 0.01), more severely ill based on the Sequential Organ Failure Assessment (SOFA) score (7 [5, 9] vs. 4 [3, 6], p < 0.01), dynamic lung compliance (15 mL/cmH20 [11, 20] vs. 27 mL/cmH20 [23, 35], p < 0.01), oxygenation index (26 [22, 33] vs. 9 [6, 11], p < 0.01), and their need for rescue modes of ventilation. ECMO patients had significantly longer lengths of hospitalization (46 [27, 62] vs. 16 [12, 31] days, p < 0.01) ICU stay (29 [19, 43] vs. 10 [5, 17] days, p < 0.01), and duration of mechanical ventilation (24 [14, 42] vs. 10 [7, 17] days, p < 0.01). Functional outcomes were similar in ECMO and non-ECMO patients. ECMO did not predict changes in lung function when adjusting for age, SOFA, COVID-19 status, or length of hospitalization. CONCLUSIONS: There were no significant differences in the FVC% predicted, or other markers of pulmonary, neurocognitive, or psychiatric functional recovery outcomes, when comparing a contemporaneous clinic-based cohort of survivors of ARDS managed with ECMO to those without ECMO.

Treatment of hyperammonemia using in-line renal replacement and hyperosmolar therapies within an extracorporeal membrane oxygenation circuit.

After orthotopic lung transplantation, hyperammonemia can be a rare complication secondary to infection by organisms that produce urease or inhibit the urea cycle. This can cause neurotoxicity, cerebral edema, and seizures. Ammonia is unique in that it has a large volume of distribution. However, it is also readily dialyzable given its small molecular weight. As such, removal of ammonia requires renal replacement modalities that can both rapidly remove ammonia from the plasma space and allow for continuous removal to prevent rebound accumulation from intracellular stores. Prevention of iatrogenic osmotic lowering in this setting is required to prevent worsening of cerebral edema. Herein, we describe use of sequential in-line renal replacement therapy using both intermittent hemodialysis and continuous venovenous hemofiltration within an extracorporeal membrane oxygenation circuit in conjunction with higher sodium dialysate and 7.5% hypertonic saline to achieve these treatment goals.

Veno-arterial extracorporeal membrane oxygenation without allogeneic blood transfusion: An observational cohort study.

INTRODUCTION: It remains unclear whether patients who will not accept allogeneic blood transfusion can be managed successfully with veno-arterial (V-A) extracorporeal membrane oxygenation (ECMO). The objective of our study was to determine what percentage of V-A ECMO patients were managed without allogeneic blood transfusion. METHODS: This was a retrospective, observational cohort study of patients with cardiogenic shock requiring V-A ECMO between January 2016 and January 2019. The primary outcome was avoidance of any allogeneic blood transfusion. RESULTS: Of the 206 patients included, 23 (11.2%) were managed without any allogeneic blood transfusion. Fourteen (60.9%) avoided allogeneic blood transfusion during their entire hospitalization. "No-transfusion" patients were younger, more commonly men, were less likely to have a prior diagnosis of hypertension or coronary artery disease, had higher baseline hemoglobin, had higher SAVE scores, and were less likely to have received aspirin before ECMO. No patients in the "no-transfusion" group had major bleeding compared to 35% of patients in the blood transfusion group (p < 0.001). In-hospital mortality was 17.4% for those who avoided blood transfusion and 41.5% for those who received blood transfusion (p = 0.04). ECMO duration was significantly shorter in patients who avoided blood transfusion compared to those who received blood transfusion (median 3.5 vs 7 days, p < 0.001). CONCLUSIONS: Select patients can be successfully managed on V-A ECMO without allogeneic blood transfusion. Jehovah's Witnesses and other patients with objections to allogeneic transfusion might be offered V-A ECMO if its anticipated duration is short (e.g. <7 days) and baseline hemoglobin concentration is high (e.g. ≥10 mg/dL).

In Vitro Comparison of Recombinant and Plasma-Derived von Willebrand Factor Concentrate for Treatment of Acquired von Willebrand Syndrome in Adult Extracorporeal Membrane Oxygenation Patients.

BACKGROUND: Coagulopathic bleeding is common during adult extracorporeal membrane oxygenation (ECMO), and acquired von Willebrand syndrome is a contributing factor. We compared ECMO patient blood samples that were treated in vitro with recombinant von Willebrand Factor concentrate and plasma-derived von Willebrand Factor concentrate. Our hypothesis was that recombinant von Willebrand Factor (vWF) would have greater efficacy in increasing vWF function. Secondarily, we hypothesized that recombinant vWF would have less impact on thrombin generation. METHODS: Thirty ECMO patients and 10 cardiac surgical controls were enrolled in the study. ECMO patient blood samples were treated in vitro with low- and high-dose recombinant vWFs and low- and high-dose plasma-derived vWFs. Whole blood ristocetin-induced platelet aggregation (RIPA), plasma ristocetin cofactor activity (RCo), and thrombin generation were compared between ECMO patient blood samples and control blood samples and between vWF-treated ECMO patient blood samples and nontreated samples. RESULTS: ECMO patient blood samples had severely reduced median RIPA compared to control samples 2 ohms (1-12 [25th-75th percentile]) vs 20 ohms (11-42) (P < .001). Treatment of ECMO patient blood samples with high-dose recombinant vWF significantly increased median RIPA to 10 ohms (2-15) (P < .001), while low-dose recombinant vWF and low- and high-dose plasma-derived vWFs did not significantly increase RIPA; 6 ohms (3-14), 4 ohms (1-13), and 6 ohms (2-10), respectively (P = .25, >.99, and >.99). Treatment with high-dose recombinant vWF and low- and high-dose plasma-derived vWFs significantly increased median plasma RCo to 4.7 international units (IU)/mL (3.7-5.9), 3.3 IU/mL (2.7-4.8), and 3.9 IU/mL (3.4-5.3), respectively, compared to controls 1.8 IU/mL (1.5-2.3) (all P < .001). Treatment with low- and high-dose plasma-derived vWFs significantly increased mean endogenous thrombin potential (6270.2 ± 2038.7 and 6313.1 ± 1913.3) compared to nontreated samples (5856.7 ± 1924.6) (P = .04 and .006), whereas treatment with low- and high-dose recombinant vWFs had no significant effect on mean endogenous thrombin potential (5776.1 ± 2087.3 and 5856.2 ± 1946.4) (P > .99 for both comparisons). CONCLUSIONS: In vitro treatment of ECMO patient blood samples with high-dose recombinant vWF was superior to low-dose recombinant vWF and plasma-derived vWF in terms of improving RIPA. In addition, recombinant vWF treatment did not increase endogenous thrombin potential, which may reduce overall thrombotic risk if it used to treat acquired von Willebrand syndrome in ECMO patients.

New-Onset Atrial Arrhythmias Are Independently Associated With In-Hospital Mortality in Veno-Venous Extracorporeal Membrane Oxygenation.

OBJECTIVE: To explore if atrial arrhythmias are associated with in-hospital mortality in veno-venous extracorporeal membrane oxygenation (VV-ECMO) patients. DESIGN: Retrospective observational cohort study. SETTING: Quaternary care academic medical center. PARTICIPANTS: Patients with respiratory failure requiring VV-ECMO for >24 hours between January 1, 2016, and January 1, 2019. INTERVENTIONS: None, observational study. MEASUREMENTS AND MAIN RESULTS: Two hundred nineteen VV-ECMO patients were included. Patients were stratified by absence or presence of clinically significant atrial arrhythmias during the VV-ECMO run. Atrial arrhythmias were defined as either atrial fibrillation or atrial flutter that occurred during VV-ECMO and required pharmacologic or electrical intervention. The primary outcome was in-hospital mortality. Secondary outcomes included a composite of thrombotic events, which included ischemic stroke and on-pump arterial thrombosis. Other objectives of this analysis included characterization of atrial arrhythmia incidence, risk factors, and management. A total of 67 patients (30.5%) experienced new-onset atrial arrhythmias post-ECMO cannulation. Age, male sex, and norepinephrine use were independently associated with atrial arrhythmia development. In-hospital mortality was significantly higher in the atrial arrhythmia group (38.8% v 19.1%; p = 0.003). In the multivariate logistic regression analysis, atrial arrhythmias during VV-ECMO were independently associated with increased odds of in-hospital mortality (odds ratio, 2.21; 95% confidence interval, 1.08-4.55; p = 0.03), after controlling for Respiratory Extracorporeal Membrane Oxygenation Survival Prediction score, acute renal failure, total norepinephrine dose, and total cannulation time. CONCLUSIONS: New-onset atrial arrhythmias are a frequent complication during VV-ECMO and are independently associated with excessive in-hospital mortality. Thus, their presence may serve as an important prognostic tool in this patient population.

Precannulation International Normalized Ratio is Independently Associated With Mortality in Veno-Arterial Extracorporeal Membrane Oxygenation.

OBJECTIVES: To explore whether precannulation international normalized ratio (INR) is associated with in-hospital mortality in venoarterial extracorporeal membrane oxygenation (VA-ECMO) patients. DESIGN: A retrospective, observational cohort study. SETTING: A quaternary care academic medical center. PARTICIPANTS: Patients with cardiogenic shock on VA-ECMO for >24 hours. INTERVENTIONS: None, observational study. MEASUREMENTS AND MAIN RESULTS: A total of 188 patients who were on VA-ECMO were included over three years. Patients were stratified into three groups based on their pre-ECMO INR: INR <1.5, INR 1.5 to 1.8, and INR >1.8. For all patients, demographics, comorbidities, and ECMO details were recorded. The study's primary outcome was in-hospital mortality and secondary outcomes included major bleeding, minor bleeding, allogeneic transfusion, ischemic stroke, intracranial hemorrhage, acute renal failure, acute liver failure, gastrointestinal bleeding, intensive care unit and hospital lengths of stay. A multivariate logistic regression was used to determine whether precannulation INR was associated independently with in-hospital mortality. In-hospital mortality differed significantly by INR group (51.6% INR >1.8 v 42.3% INR 1.5-1.8 v 24.3% INR <1.5; p = 0.004). In a multivariate logistic regression model, precannulation INR >1.8 was associated independently with an increased odds of mortality (odds ratio, 2.48; 95% confidence interval, 1.05-6.04) after controlling for sex, Survival after VA- ECMO score, and ECMO indication. An INR within 1.5 to 1.8 did not confer an increased mortality risk. CONCLUSIONS: An INR >1.8 before VA-ECMO cannulation is associated independently with in-hospital mortality. Precannulation INR should be considered by clinicians so that ECMO resources can be better allocated and risks of organ failure and intracranial hemorrhage can be better understood.

The Impact of Corticosteroids on Secondary Infection and Mortality in Critically Ill COVID-19 Patients.

BACKGROUND: Corticosteroids are part of the treatment guidelines for COVID-19 and have been shown to improve mortality. However, the impact corticosteroids have on the development of secondary infection in COVID-19 is unknown. We sought to define the rate of secondary infection in critically ill patients with COVID-19 and determine the effect of corticosteroid use on mortality in critically ill patients with COVID-19. STUDY DESIGN AND METHODS: One hundred and thirty-five critically ill patients with COVID-19 admitted to the Intensive Care Unit (ICU) at the University of Maryland Medical Center were included in this single-center retrospective analysis. Demographics, symptoms, culture data, use of COVID-19 directed therapies, and outcomes were abstracted from the medical record. The primary outcomes were secondary infection and mortality. Proportional hazards models were used to determine the time to secondary infection and the time to death. RESULTS: The proportion of patients with secondary infection was 63%. The likelihood of developing secondary infection was not significantly impacted by the administration of corticosteroids (HR 1.45, CI 0.75-2.82, P = 0.28). This remained consistent in sub-analysis looking at bloodstream, respiratory, and urine infections. Secondary infection had no significant impact on the likelihood of 28-day mortality (HR 0.66, CI 0.33-1.35, P = 0.256). Corticosteroid administration significantly reduced the likelihood of 28-day mortality (HR 0.27, CI 0.10-0.72, P = 0.01). CONCLUSION: Corticosteroids are an important and lifesaving pharmacotherapeutic option in critically ill patients with COVID-19, which have no impact on the likelihood of developing secondary infections.

Platelet factor-4 concentration in adult veno-arterial ECMO patients.

BACKGROUND: Heparin induced thrombocytopenia (HIT) is reported at a variable rate in extracorporeal membrane oxygenation (ECMO) patients. A critical factor impacting platelet factor-4 (PF4)-heparin antibody formation is plasma PF4 concentration. We hypothesized that PF4 concentration would be increased during veno-arterial (VA) ECMO. METHODS: Plasma PF4 concentration was measured during the first 5 ECMO days in 20 VA ECMO patients and 10 control plasma samples. PF4-heparin ratios were estimated using an assumed heparin concentration of 0.4 IU/mL. This correlates with an activated partial thromboplastin time of 60 to 80 seconds, which is the anticoagulation target in our center. RESULTS: Twenty VA ECMO patients were enrolled, 10 of which had pulmonary embolism. Median PF4 concentration was 0.03 µg/mL [0.01, 0.13] in control plasma. Median PF4 concentration was 0.21 µg/mL [0.12, 0.34] on ECMO day 1 or 2, 0.16 µg/mL [0.09, 0.25] on ECMO day 3, and 0.12 µg/mL [0.09, 0.22] on ECMO day 5. Estimated median PF4-heparin ratios were 0.04, 0.03, and 0.02 respectively. Two patients (10%) developed HIT that was confirmed by serotonin release assay. PF4 concentration did not differ significantly in these patients compared to non-HIT patients (p = 0.37). No patient had an estimated PF4-heparin ratio between 0.7 and 1.4, which is the reported optimal range for PF4-heparin antibody formation. CONCLUSION: Our data suggest that PF4 concentration is mildly elevated during VA ECMO compared to control plasma. Estimated PF4-heparin ratios were not optimal for HIT antibody formation. These data support epidemiologic studies where HIT incidence is low during VA ECMO.

Perioperative Applications of Therapeutic Plasma Exchange in Cardiac Surgery: A Narrative Review.

Therapeutic plasma exchange (TPE) has a number of applications in cardiac surgical patients and has been used increasingly in high-risk heart and lung transplant patients. In this narrative review, the authors describe TPE principles, complications, and specific indications for TPE, including thrombotic thrombocytopenic purpura, heparin-induced thrombocytopenia, induction of immunotolerance in heart and lung transplant patients, and treatment of antibody-mediated rejection in heart and lung transplant patients. The review is based on published literature and the authors' institutional experience with perioperative TPE in cardiac surgical patients.

High-efficiency, high-flux in-line hemofiltration using a high blood flow extracorporeal circuit.

The ability of current renal replacement therapy modalities to achieve rapid solute removal is limited by membrane surface area and blood flow rate. Extracorporeal membrane oxygenation offers high blood flow and hemodynamic support that may be harnessed to overcome limitations in traditional renal replacement therapy. Using an extracorporeal membrane oxygenation circuit, we describe a high blood flow, high-efficiency hemofiltration technique using in-line hemofilters (hemoconcentrators) and standard replacement fluid to enhance solute clearance. Using this approach and a total of 5 L of replacement volume per treatment, creatinine (Cr) clearances of 8.3 L/hour and 11.2 L/hour using one and two hemoconcentrators, respectively, were achieved. With use of a high blood flow rate of up to 5 L/min, this hemofiltration technique can potentially offer clearance of 30 times that of continuous renal replacement therapy and of 6 times that of hemodialysis which may expand the ability to remove substances traditionally not considered removable via existing extracorporeal therapies.