Intranasal soluble ACE2 improves survival and prevents brain SARS-CoV-2 infection.

A soluble ACE2 protein bioengineered for long duration of action and high affinity to SARS-CoV-2 was administered either intranasally (IN) or intraperitoneally (IP) to SARS-CoV-2-inoculated k18hACE2 mice. This decoy protein (ACE2 618-DDC-ABD) was given either IN or IP, pre- and post-inoculation, or IN, IP, or IN + IP but only post-inoculation. Survival by day 5 was 0% in untreated mice, 40% in the IP-pre, and 90% in the IN-pre group. In the IN-pre group, brain histopathology was essentially normal and lung histopathology significantly improved. Consistent with this, brain SARS-CoV-2 titers were undetectable and lung titers reduced in the IN-pre group. When ACE2 618-DDC-ABD was administered only post-inoculation, survival was 30% in the IN + IP, 20% in the IN, and 20% in the IP group. We conclude that ACE2 618-DDC-ABD results in markedly improved survival and provides organ protection when given intranasally as compared with when given either systemically or after viral inoculation, and that lowering brain titers is a critical determinant of survival and organ protection.

Cytomorphology of renal angiomyolipoma: performance and accuracy of touch preparation of core needle biopsy.

INTRODUCTION: The recognition of renal angiomyolipoma (AML) can be challenging based on cytology preparations such as touch preparation (TP) of core needle biopsy (CNB) and fine needle aspiration. This study evaluated the cytologic features and performance of TP of CNB during rapid onsite evaluation (ROSE) of renal AML. MATERIALS AND METHODS: A pathology database search was performed between 2000 and 2021 for renal CNB specimens with ROSE using TP that were primarily favored AML on preliminary impression and/or confirmed AML on CNB or subsequent resection. RESULTS: Twenty confirmed AML were identified (90% female, median age 65.5 years). Sixteen (80%) were deemed adequate for diagnosis at the time of ROSE, and 9 of 16 (56%) had available onsite impression: AML was favored in 4 of 9 cases (44%). Examination of TP slides revealed spindle/epithelioid cells in 20 (100%), adipose tissue in 14 (70%), and blood vessels in 3 (15%). All AML cases were subsequently confirmed by immunohistochemistry. Additionally, 3 other cases with ROSE favoring AML revealed to be "renal parenchyma with fibrosis," clear cell papillary renal cell tumor and clear cell renal cell carcinoma. CONCLUSIONS: Onsite evaluation of TP ensures adequate material for diagnosis in most renal AML. Spindle/epithelioid cells were the most common component seen on TP, followed by adipose tissue. Blood vessels were rarely seen. While the recognition of AML at ROSE can be challenging, proper evaluation is important in obtaining adequate diagnostic tissue. Correlation with CNB and utilization of immunohistochemistry are crucial for arriving at the diagnosis.

Video-Based Education Improves Sampling (Grossing) Confidence in Pathology Trainees.

CONTEXT.­: Pathology resident education has a steep learning curve. Specimen sampling (grossing) is a procedural task, and procedural fields add video materials to their curricula to familiarize trainees with procedure(s), reduce errors, and improve patient care. Our team applied this strategy to develop original in-house sampling videos for our program. OBJECTIVES.­: To evaluate the effect of in-house sampling videos on resident sampling confidence. DESIGN.­: Sampling videos covering all major organ systems (AMOS) were created for our postgraduate year 1 (PGY1) trainees. Videos were hosted on a Northwestern cloud server for on-demand access. Trainees completed 3 surveys (0, 6, 12 months) evaluating sampling confidence comparing those who used in-house videos as an educational supplement with those who did not use the videos. RESULTS.­: Sampling confidence significantly improved at 6 and 12 months (P < .001) across AMOS and PGY levels. When compared with those who did not use in-house sampling videos, trainees who supplemented their education with in-house sampling videos had significantly higher confidence ratings across AMOS and PGY levels at the start of the study (P < .001) and at 6 months (P = .004). Sampling confidence significantly improved for PGY1 trainees at 6 and 12 months (P < .001); for PGY2 and PGY3 trainees, confidence significantly improved at 6 months (P < .001). When evaluated by organ-specific analyses, sampling and teaching confidence improved across all organ systems and, except for the gastrointestinal system, reached significance at 12 months for all PGY levels. CONCLUSIONS.­: Sampling videos, when used as a supplement to the existing curriculum, significantly improved trainee confidence.

Superiority of intranasal over systemic administration of bioengineered soluble ACE2 for survival and brain protection against SARS-CoV-2 infection.

The present study was designed to investigate the effects of a soluble ACE2 protein termed ACE2 618-DDC-ABD, bioengineered to have long duration of action and high binding affinity to SARS-CoV-2, when administered either intranasally (IN) or intraperitoneally (IP) and before or after SARS-CoV-2 inoculation. K18hACE2 mice permissive for SARS-CoV-2 infection were inoculated with 2Ã-10 4 PFU wildtype SARS-CoV-2. In one protocol, ACE2 618-DDC-ABD was given either IN or IP, pre- and post-viral inoculation. In a second protocol, ACE2 618-DDC-ABD was given either IN, IP or IN+IP but only post-viral inoculation. In addition, A549 and Vero E6 cells were used to test neutralization of SARS-CoV-2 variants by ACE2 618-DDC-ABD at different concentrations. Survival by day 5 was 0% in infected untreated mice, and 40% in mice from the ACE2 618-DDC-ABD IP-pre treated group. By contrast, in the IN-pre group survival was 90%, histopathology of brain and kidney was essentially normal and markedly improved in the lungs. When ACE2 618-DDC-ABD was administered only post viral inoculation, survival was 30% in the IN+IP group, 20% in the IN and 0% in the IP group. Brain SARS-CoV-2 titers were high in all groups except for the IN-pre group where titers were undetectable in all mice. In cells permissive for SARS-CoV-2 infection, ACE2 618-DDC-ABD neutralized wildtype SARS-CoV-2 at high concentrations, whereas much lower concentrations neutralized omicron BA. 1. We conclude that ACE2 618-DDC-ABD provides much better survival and organ protection when administered intranasally than when given systemically or after viral inoculation and that lowering brain titers is a critical determinant of survival and organ protection.

Spherical nucleic acids as an infectious disease vaccine platform.

SignificanceUsing SARS-CoV-2 as a relevant case study for infectious disease, we investigate the structure-function relationships that dictate antiviral spherical nucleic acid (SNA) vaccine efficacy. We show that the SNA architecture can be rapidly employed to target COVID-19 through incorporation of the receptor-binding domain, and that the resulting vaccine potently activates human cells in vitro and mice in vivo. Furthermore, when challenged with a lethal viral infection, only mice treated with the SNA vaccine survived. Taken together, this work underscores the importance of rational vaccine design for infectious disease to yield vaccines that elicit more potent immune responses to effectively fight disease.

A Novel Soluble ACE2 Protein Provides Lung and Kidney Protection in Mice Susceptible to Lethal SARS-CoV-2 Infection.

BACKGROUND: Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) uses full-length angiotensin converting enzyme 2 (ACE2) as a main receptor to enter target cells. The goal of this study was to demonstrate the preclinical efficacy of a novel soluble ACE2 protein with increased duration of action and binding capacity in a lethal mouse model of COVID-19. METHODS: A human soluble ACE2 variant fused with an albumin binding domain (ABD) was linked via a dimerization motif hinge-like 4-cysteine dodecapeptide (DDC) to improve binding capacity to SARS-CoV-2. This novel soluble ACE2 protein (ACE2-1-618-DDC-ABD) was then administered intranasally and intraperitoneally to mice before intranasal inoculation of SARS-CoV-2 and then for two additional days post viral inoculation. RESULTS: Untreated animals became severely ill, and all had to be humanely euthanized by day 6 or 7 and had pulmonary alveolar hemorrhage with mononuclear infiltrates. In contrast, all but one mouse infected with a lethal dose of SARS-CoV-2 that received ACE2-1-618-DDC-ABD survived. In the animals inoculated with SARS-CoV-2 that were untreated, viral titers were high in the lungs and brain, but viral titers were absent in the kidneys. Some untreated animals, however, had variable degrees of kidney proximal tubular injury as shown by attenuation of the proximal tubular brush border and increased NGAL and TUNEL staining. Viral titers in the lung and brain were reduced or nondetectable in mice that received ACE2-1-618-DDC-ABD, and the animals developed only moderate disease as assessed by a near-normal clinical score, minimal weight loss, and improved lung and kidney injury. CONCLUSIONS: This study demonstrates the preclinical efficacy of a novel soluble ACE2 protein, termed ACE2-1-618-DDC-ABD, in a lethal mouse model of SARS-CoV-2 infection that develops severe lung injury and variable degrees of moderate kidney proximal tubular injury.

Cytology-histology correlation of myoepithelial tumors harboring EWSR1-POU5F1 fusions: A report of two cases.

Myoepithelial tumors (MET) constitute a group of neoplasms with a variety of morphologic, immunophenotypic, and molecular features. Approximately half of MET of soft tissue harbor EWSR1 gene rearrangements with a subset showing EWSR1-POU5F1 fusions and demonstrating distinctive tendency towards aggressive behavior in children. Histologically, EWSR1-POU5F1-positive MET typically show clear-cell morphology with malignant features including marked pleomorphism and atypical mitotic figures. The cytomorphology of these tumors has not been well characterized. Reported here are the cytomorphologic features of two cases of EWSR1-POUF1-positive MET with histology correlation.

Nondestructive versus advanced post-transplant lymphoproliferative disorder in a single-center pediatric kidney transplant population.

BACKGROUND: Pediatric kidney transplant recipients are at risk for the development of post-transplant lymphoproliferative disorders (PTLD), a group of potentially devastating diseases that present on a spectrum of severity ranging from nondestructive PTLD to more histologically destructive lesions. Currently, there is inadequate evidence to guide evaluation and management of nondestructive PTLD. METHODS: This is a single-center case series of pediatric kidney transplant recipients between January 1, 2008 and December 31, 2019, who were diagnosed with PTLD. The aim was to describe clinical characteristics, presentation, and management of nondestructive versus advanced PTLD. RESULTS: Eighteen patients were diagnosed with nondestructive PTLD and seven with more advanced PTLD histopathology. The majority (66.7%) of nondestructive PTLD patients (n = 16) presented with tonsillar hypertrophy and/or snoring and were managed conservatively, with minimal reduction in tacrolimus dose and no further evaluation. No patient progressed to more advanced PTLD. Advanced PTLD patients (n = 7) were more likely to present with fever, elevated creatinine, a new mass of gastrointestinal symptoms. They received workup with imaging and oncology consultation, and were managed with chemotherapy. CONCLUSIONS: Patients with nondestructive PTLD often present with symptoms of sleep-disordered breathing and can be managed conservatively with excellent clinical outcomes. More study is needed to guide care of this under-researched population.

Comprehensive evaluation of bronchoalveolar lavage from patients with severe COVID-19 and correlation with clinical outcomes.

Information on bronchoalveolar lavage (BAL) in patients with COVID-19 is limited, and clinical correlation has not been reported. This study investigated the key features of BAL fluids from COVID-19 patients and assessed their clinical significance. A total of 320 BAL samples from 83 COVID-19 patients and 70 non-COVID-19 patients (27 patients with other respiratory viral infections) were evaluated, including cell count/differential, morphology, flow cytometric immunophenotyping, and immunohistochemistry. The findings were correlated with clinical outcomes. Compared to non-COVID-19 patients, BAL from COVID-19 patients was characterized by significant lymphocytosis (p < 0.001), in contrast to peripheral blood lymphopenia commonly observed in COVID-19 patients and the presence of atypical lymphocytes with plasmacytoid/plasmablastic features (p < 0.001). Flow cytometry and immunohistochemistry demonstrated that BAL lymphocytes, including plasmacytoid and plasmablastic cells, were composed predominantly of T cells with a mixture of CD4+ and CD8+ cells. Both populations had increased expression of T-cell activation markers, suggesting important roles of helper and cytotoxic T-cells in the immune response to SARS-CoV-2 infection in the lung. More importantly, BAL lymphocytosis was significantly associated with longer hospital stay (p < 0.05) and longer requirement for mechanical ventilation (p < 0.05), whereas the median atypical (activated) lymphocyte count was associated with shorter hospital stay (p < 0.05), shorter time on mechanical ventilation (p < 0.05) and improved survival. Our results indicate that BAL cellular analysis and morphologic findings provide additional important information for diagnostic and prognostic work-up, and potential new therapeutic strategies for patients with severe COVID-19.

A novel soluble ACE2 protein totally protects from lethal disease caused by SARS-CoV-2 infection.

Severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) uses full-length angiotensin converting enzyme 2 (ACE2), which is membrane bound, as its initial cell contact receptor preceding viral entry. Here we report a human soluble ACE2 variant fused with a 5kD albumin binding domain (ABD) and bridged via a dimerization motif hinge-like 4-cysteine dodecapeptide, which we term ACE2 1-618-DDC-ABD. This protein is enzymatically active, has increased duration of action in vivo conferred by the ABD-tag, and displays 20-30-fold higher binding affinity to the SARS-CoV-2 receptor binding domain than its des-DDC monomeric form (ACE2 1-618-ABD) due to DDC-linked dimerization. ACE2 1-618-DDC-ABD was administered for 3 consecutive days to transgenic k18-hACE2 mice, a model that develops lethal SARS-CoV-2 infection, to evaluate the preclinical preventative/ therapeutic value for COVID-19. Mice treated with ACE2 1-618-DDC-ABD developed a mild to moderate disease for the first few days assessed by a clinical score and modest weight loss. The untreated control animals, by contrast, became severely ill and had to be sacrificed by day 6/7 and lung histology revealed extensive pulmonary alveolar hemorrhage and mononuclear infiltrates. At 6 days, mortality was totally prevented in the treated group, lung histopathology was improved and viral titers markedly reduced. This demonstrates for the first time in vivo the preventative/ therapeutic potential of a novel soluble ACE2 protein in a preclinical animal model.

Experimental coevolution: rapid local adaptation by parasites depends on host mating system.

Host-parasite interactions can drive rapid, reciprocal genetic changes (coevolution), provided both hosts and parasites have high heritabilities for resistance/infectivity. Similarly, the host's mating system should also affect the rate of coevolutionary change in host-parasite interactions. Using experimental coevolution, we determined the effect of obligate outcrossing verses partial self-fertilization (mixed mating) on the rate of evolutionary change in a nematode host (Caenorhabditis elegans) and its bacterial parasite (Serratia marcescens). Bacterial populations were derived from a common ancestor. We measured the effects of host mating system on host adaptation to the parasite. We then determined the extent of parasite adaptation to their local host populations. Obligately outcrossing hosts exhibited more rapid adaptation to parasites than did mixed mating hosts. In addition, most of the parasites became adapted to infecting their local hosts, but parasites from obligately outcrossing hosts showed a greater level of local adaptation. These results suggest that host populations evolved along separate trajectories and that outcrossing host populations diverged further than partially selfing populations. Finally, parasites tracking outcrossing host populations diverged further than parasites tracking the partially selfing host populations. These results show that the evolutionary trajectories of both hosts and parasites can be shaped by the host's mating system.

Temporal dynamics of outcrossing and host mortality rates in host-pathogen experimental coevolution.

Cross-fertilization is predicted to facilitate the short-term response and the long-term persistence of host populations engaged in antagonistic coevolutionary interactions. Consistent with this idea, our previous work has shown that coevolving bacterial pathogens (Serratia marcescens) can drive obligately selfing hosts (Caenorhabditis elegans) to extinction, whereas the obligately outcrossing and partially outcrossing populations persisted. We focused the present study on the partially outcrossing (mixed mating) and obligately outcrossing hosts, and analyzed the changes in the host resistance/avoidance (and pathogen infectivity) over time. We found that host mortality rates increased in the mixed mating populations over the first 10 generations of coevolution when outcrossing rates were initially low. However, mortality rates decreased after elevated outcrossing rates evolved during the experiment. In contrast, host mortality rates decreased in the obligately outcrossing populations during the first 10 generations of coevolution, and remained low throughout the experiment. Therefore, predominant selfing reduced the ability of the hosts to respond to coevolving pathogens compared to outcrossing hosts. Thus, we found that host-pathogen coevolution can generate rapid evolutionary change, and that host mating system can influence the outcome of coevolution at a fine temporal scale.

Running with the Red Queen: host-parasite coevolution selects for biparental sex.

Most organisms reproduce through outcrossing, even though it comes with substantial costs. The Red Queen hypothesis proposes that selection from coevolving pathogens facilitates the persistence of outcrossing despite these costs. We used experimental coevolution to test the Red Queen hypothesis and found that coevolution with a bacterial pathogen (Serratia marcescens) resulted in significantly more outcrossing in mixed mating experimental populations of the nematode Caenorhabditis elegans. Furthermore, we found that coevolution with the pathogen rapidly drove obligately selfing populations to extinction, whereas outcrossing populations persisted through reciprocal coevolution. Thus, consistent with the Red Queen hypothesis, coevolving pathogens can select for biparental sex.