Mild respiratory COVID can cause multi-lineage neural cell and myelin dysregulation.

COVID survivors frequently experience lingering neurological symptoms that resemble cancer-therapy-related cognitive impairment, a syndrome for which white matter microglial reactivity and consequent neural dysregulation is central. Here, we explored the neurobiological effects of respiratory SARS-CoV-2 infection and found white-matter-selective microglial reactivity in mice and humans. Following mild respiratory COVID in mice, persistently impaired hippocampal neurogenesis, decreased oligodendrocytes, and myelin loss were evident together with elevated CSF cytokines/chemokines including CCL11. Systemic CCL11 administration specifically caused hippocampal microglial reactivity and impaired neurogenesis. Concordantly, humans with lasting cognitive symptoms post-COVID exhibit elevated CCL11 levels. Compared with SARS-CoV-2, mild respiratory influenza in mice caused similar patterns of white-matter-selective microglial reactivity, oligodendrocyte loss, impaired neurogenesis, and elevated CCL11 at early time points, but after influenza, only elevated CCL11 and hippocampal pathology persisted. These findings illustrate similar neuropathophysiology after cancer therapy and respiratory SARS-CoV-2 infection which may contribute to cognitive impairment following even mild COVID.

Inflammation Improves Glucose Homeostasis through IKKß-XBP1s Interaction.

It is widely believed that inflammation associated with obesity has an important role in the development of type 2 diabetes. IκB kinase beta (IKKß) is a crucial kinase that responds to inflammatory stimuli such as tumor necrosis factor α (TNF-α) by initiating a variety of intracellular signaling cascades and is considered to be a key element in the inflammation-mediated development of insulin resistance. We show here, contrary to expectation, that IKKß-mediated inflammation is a positive regulator of hepatic glucose homeostasis. IKKß phosphorylates the spliced form of X-Box Binding Protein 1 (XBP1s) and increases the activity of XBP1s. We have used three experimental approaches to enhance the IKKß activity in the liver of obese mice and observed increased XBP1s activity, reduced ER stress, and a significant improvement in insulin sensitivity and consequently in glucose homeostasis. Our results reveal a beneficial role of IKKß-mediated hepatic inflammation in glucose homeostasis.

Treatment of obesity with celastrol.

Despite all modern advances in medicine, an effective drug treatment of obesity has not been found yet. Discovery of leptin two decades ago created hopes for treatment of obesity. However, development of leptin resistance has been a big obstacle, mitigating a leptin-centric treatment of obesity. Here, by using in silico drug-screening methods, we discovered that Celastrol, a pentacyclic triterpene extracted from the roots of Tripterygium Wilfordi (thunder god vine) plant, is a powerful anti-obesity agent. Celastrol suppresses food intake, blocks reduction of energy expenditure, and leads to up to 45% weight loss in hyperleptinemic diet-induced obese (DIO) mice by increasing leptin sensitivity, but it is ineffective in leptin-deficient (ob/ob) and leptin receptor-deficient (db/db) mouse models. These results indicate that Celastrol is a leptin sensitizer and a promising agent for the pharmacological treatment of obesity.

Distinguishing features of long COVID identified through immune profiling.

Post-acute infection syndromes may develop after acute viral disease1. Infection with SARS-CoV-2 can result in the development of a post-acute infection syndrome known as long COVID. Individuals with long COVID frequently report unremitting fatigue, post-exertional malaise, and a variety of cognitive and autonomic dysfunctions2-4. However, the biological processes that are associated with the development and persistence of these symptoms are unclear. Here 275 individuals with or without long COVID were enrolled in a cross-sectional study that included multidimensional immune phenotyping and unbiased machine learning methods to identify biological features associated with long COVID. Marked differences were noted in circulating myeloid and lymphocyte populations relative to the matched controls, as well as evidence of exaggerated humoral responses directed against SARS-CoV-2 among participants with long COVID. Furthermore, higher antibody responses directed against non-SARS-CoV-2 viral pathogens were observed among individuals with long COVID, particularly Epstein-Barr virus. Levels of soluble immune mediators and hormones varied among groups, with cortisol levels being lower among participants with long COVID. Integration of immune phenotyping data into unbiased machine learning models identified the key features that are most strongly associated with long COVID status. Collectively, these findings may help to guide future studies into the pathobiology of long COVID and help with developing relevant biomarkers.

Impact of circulating SARS-CoV-2 variants on mRNA vaccine-induced immunity.

The emergence of SARS-CoV-2 variants with mutations in major neutralizing antibody-binding sites can affect humoral immunity induced by infection or vaccination1-6. Here we analysed the development of anti-SARS-CoV-2 antibody and T cell responses in individuals who were previously infected (recovered) or uninfected (naive) and received mRNA vaccines to SARS-CoV-2. While individuals who were previously infected sustained higher antibody titres than individuals who were uninfected post-vaccination, the latter reached comparable levels of neutralization responses to the ancestral strain after the second vaccine dose. T cell activation markers measured upon spike or nucleocapsid peptide in vitro stimulation showed a progressive increase after vaccination. Comprehensive analysis of plasma neutralization using 16 authentic isolates of distinct locally circulating SARS-CoV-2 variants revealed a range of reduction in the neutralization capacity associated with specific mutations in the spike gene: lineages with E484K and N501Y/T (for example, B.1.351 and P.1) had the greatest reduction, followed by lineages with L452R (for example, B.1.617.2). While both groups retained neutralization capacity against all variants, plasma from individuals who were previously infected and vaccinated displayed overall better neutralization capacity than plasma from individuals who were uninfected and also received two vaccine doses, pointing to vaccine boosters as a relevant future strategy to alleviate the effect of emerging variants on antibody neutralizing activity.

Intranasal neomycin evokes broad-spectrum antiviral immunity in the upper respiratory tract.

Respiratory virus infections in humans cause a broad-spectrum of diseases that result in substantial morbidity and mortality annually worldwide. To reduce the global burden of respiratory viral diseases, preventative and therapeutic interventions that are accessible and effective are urgently needed, especially in countries that are disproportionately affected. Repurposing generic medicine has the potential to bring new treatments for infectious diseases to patients efficiently and equitably. In this study, we found that intranasal delivery of neomycin, a generic aminoglycoside antibiotic, induces the expression of interferon-stimulated genes (ISGs) in the nasal mucosa that is independent of the commensal microbiota. Prophylactic or therapeutic administration of neomycin provided significant protection against upper respiratory infection and lethal disease in a mouse model of COVID-19. Furthermore, neomycin treatment protected Mx1 congenic mice from upper and lower respiratory infections with a highly virulent strain of influenza A virus. In Syrian hamsters, neomycin treatment potently mitigated contact transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). In healthy humans, intranasal application of neomycin-containing Neosporin ointment was well tolerated and effective at inducing ISG expression in the nose in a subset of participants. These findings suggest that neomycin has the potential to be harnessed as a host-directed antiviral strategy for the prevention and treatment of respiratory viral infections.

LRRC15 inhibits SARS-CoV-2 cellular entry in trans.

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection is mediated by the entry receptor angiotensin-converting enzyme 2 (ACE2). Although attachment factors and coreceptors facilitating entry are extensively studied, cellular entry factors inhibiting viral entry are largely unknown. Using a surfaceome CRISPR activation screen, we identified human LRRC15 as an inhibitory attachment factor for SARS-CoV-2 entry. LRRC15 directly binds to the receptor-binding domain (RBD) of spike protein with a moderate affinity and inhibits spike-mediated entry. Analysis of human lung single-cell RNA sequencing dataset reveals that expression of LRRC15 is primarily detected in fibroblasts and particularly enriched in pathological fibroblasts in COVID-19 patients. ACE2 and LRRC15 are not coexpressed in the same cell types in the lung. Strikingly, expression of LRRC15 in ACE2-negative cells blocks spike-mediated viral entry in ACE2+ cell in trans, suggesting a protective role of LRRC15 in a physiological context. Therefore, LRRC15 represents an inhibitory attachment factor for SARS-CoV-2 that regulates viral entry in trans.

Comparative single-cell transcriptomic profile of hybrid immunity induced by adenovirus vector-based COVID-19 vaccines.

In this study, antibody response and a single-cell RNA-seq analysis were conducted on peripheral blood mononuclear cells from five different groups: naïve subjects vaccinated with AZD1222 (AZ) or Ad5-nCoV (Cso), individuals previously infected and later vaccinated (hybrid) with AZD1222 (AZ-hb) or Ad5-nCoV (Cso-hb), and those who were infected and had recovered from COVID-19 (Inf). The results showed that AZ induced more robust neutralizing antibody responses than Cso. The single-cell RNA data revealed a high frequency of memory B cells in the Cso and Cso-hb. In contrast, AZ and AZ-hb groups exhibited the highest proportion of activated naïve B cells expressing CXCR4. Transcriptomic analysis of CD4+ and CD8+ T cells demonstrated a heterogeneous response following vaccination, hybrid immunity, or natural infection. However, a single dose of Ad5-nCoV was sufficient to strongly activate CD4+ T cells (naïve and memory) expressing ANX1 and FOS, similar to the hybrid response observed with AZ. An interesting finding was the robust activation of a subset of CD8+ T cells expressing GZMB, GZMH, and IFNG genes in the Cso-hb group. Our findings suggest that both vaccines effectively stimulated the cellular immune response; however, the Ad5-nCoV induced a more robust CD8+ T-cell response in previously infected individuals.

Resident training in interventional pathology: Ultrasound-guided fine-needle aspiration and rapid on site evaluation-5 years of teaching experience in a single university hospital.

INTRODUCTION: Despite the established role of the interventional pathologist, their diagnostic performance is difficult to establish. At least in Spain training of pathology residents in ultrasound-guided interventional procedures for specimen collection is limited or absent in most institutions. We present our teaching experience in the instruction of ultrasound-guided fine-needle aspiration (FNA) to pathology residents in a tertiary-level hospital. MATERIALS AND METHODS: The training of pathology residents who rotated through the interventional unit of the pathology department and the application of ultrasound-guided FNA and rapid on-site evaluation (U-ROSE) was documented over 5 years. The training period was broken down into learning phases and included the number of ultrasound-guided FNA performed, anatomical location, and their diagnostic performance, among other aspects. RESULTS: Nineteen (19) pathology residents were trained in U-ROSE, and performed a total of 4003 procedures, with a mean of 211 per resident. In 53% of cases only one pass was required for an adequated sample. The specimen was diagnostic in more than 97% of cases. The most frequently sampled anatomical sites were the thyroid gland (n = 2347), followed by lymph node (n = 667), soft tissues (n = 663) and salivary glands (n = 322). CONCLUSION: The results support the training programme followed by pathology residents in learning U-ROSE, which is essential to lay the foundations for the future interventional pathologist.

How to Estimate Kidney Growth in Patients with Autosomal Dominant Polycystic Kidney Disease.

Autosomal dominant polycystic kidney disease (ADPKD) is a disease characterized by a progressive kidney growth due to the development of cysts that lead to gradual destruction of the surrounding parenchyma. In the first stage, the estimated GFR will remain stable despite the reduction of the renal parenchyma because of an increase in glomerular hyperfiltration. The total kidney volume (TKV) measured with computed tomography or magnetic resonance imaging is related to the future GFR decline. Thus, TKV has become an early marker to be analyzed in all patients with ADPKD. In addition, in recent years, it has been pointed out that kidney growth rate estimated with a single TKV measurement can be a clear prognostic marker for future glomerular filtration decline. However, there is no consensus on how to measure kidney volume growth in ADPKD, so each author has used different models that, not having the same meaning, have been handled as if they produced similar values. This may lead to erroneous estimates of kidney growth rate with the consequent prognostic error. The Mayo Clinic classification is now the most widely accepted prognostic model in clinical practice to predict patients who will deteriorate faster and to decide what patients should be treated with tolvaptan. However, some aspects of this model have not been discussed in depth. Our aim in this review was to present the models that can be used to estimate kidney volume growth rate in ADPKD, to facilitate their applicability in daily clinical practice.

Underdiagnosis of Primary Aldosteronism: A Review of Screening and Detection.

A clinical condition may be missed due to its higher-than-recognized prevalence or inadequate diagnostic screening. Both factors apply to primary aldosteronism, which is woefully underdiagnosed as a cause of hypertension and end-organ damage. Screening tests should be strongly considered for diseases that pose significant morbidity or mortality if left untreated, that have a high prevalence, and that have treatments that lead to improvement or cure. In this review we present the evidence for each of these points. We outline studies that estimate the prevalence of primary aldosteronism in different at-risk populations and how its recognition has changed over time. We also evaluate myriad studies of screening rates for primary aldosteronism and what factors do and do not influence current screening practices. We discuss the ideal conditions for screening, measuring the aldosterone to renin ratio in different populations that use plasma renin activity or direct renin concentration, and the steps for diagnostic workup of primary aldosteronism. Finally, we conclude with potential strategies to implement higher rates of screening and diagnosis of this common, consequential, and treatable disease.

Real-time public health communication of local SARS-CoV-2 genomic epidemiology.

Genomic epidemiology can provide a unique, real-time understanding of SARS-CoV-2 transmission patterns. Yet the potential for genomic analyses to guide local policy and community-based behavioral decisions is limited because they are often oriented towards specially trained scientists and conducted on a national or global scale. Here, we propose a new paradigm: Phylogenetic analyses performed on a local level (municipal, county, or state), with results communicated in a clear, timely, and actionable manner to strengthen public health responses. We believe that presenting results rapidly, and tailored to a non-expert audience, can serve as a template for effective public health response to COVID-19 and other emerging viral diseases.

Longitudinal Immune Profiling of a Severe Acute Respiratory Syndrome Coronavirus 2 Reinfection in a Solid Organ Transplant Recipient.

BACKGROUND: The underlying immunologic deficiencies enabling severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) reinfection are currently unknown. We describe deep longitudinal immune profiling of a transplant recipient hospitalized twice for coronavirus disease 2019 (COVID-19). METHODS: A 66-year-old male renal transplant recipient was hospitalized with COVID-19 March 2020 then readmitted to the hospital with COVID-19 233 days after initial diagnosis. Virologic and immunologic investigations were performed on samples from the primary and secondary infections. RESULTS: Whole viral genome sequencing and phylogenetic analysis revealed that viruses causing both infections were caused by distinct genetic lineages without evidence of immune escape mutations. Longitudinal comparison of cellular and humoral responses during primary SARS-CoV-2 infection revealed that this patient responded to the primary infection with low neutralization titer anti-SARS-CoV-2 antibodies that were likely present at the time of reinfection. CONCLUSIONS: The development of neutralizing antibodies and humoral memory responses in this patient failed to confer protection against reinfection, suggesting that they were below a neutralizing titer threshold or that additional factors may be required for efficient prevention of SARS-CoV-2 reinfection. Development of poorly neutralizing antibodies may have been due to profound and relatively specific reduction in naive CD4 T-cell pools. Seropositivity alone may not be a perfect correlate of protection in immunocompromised patients.

Exploring structural signatures of the lanthipeptide prochlorosin 2.8 using tandem mass spectrometry and trapped ion mobility-mass spectrometry.

Lanthipeptides are a family of ribosomally synthesized and post-translationally modified peptides (RiPPs) characterized by intramolecular thioether cross-links formed between a dehydrated serine/threonine (dSer/dThr) and a cysteine residue. Prochlorosin 2.8 (Pcn2.8) is a class II lanthipeptide that exhibits a non-overlapping thioether ring pattern, for which no biological activity has been reported yet. The variant Pcn2.8[16RGD] has been shown to bind tightly to the αvß3 integrin receptor. In the present work, tandem mass spectrometry, using collision-induced dissociation (CID) and electron capture dissociation (ECD), and trapped ion mobility spectrometry-mass spectrometry (TIMS-MS) were used to investigate structural signatures for the non-overlapping thioether ring pattern of Pcn2.8. CID experiments on Pcn2.8 yielded bi and yj fragments between the thioether cross-links, evidencing the presence of a non-overlapping thioether ring pattern. ECD experiments of Pcn2.8 showed a significant increase of hydrogen migration events near the residues involved in the thioether rings with a more pronounced effect at the dehydrated residues as compared to the cysteine residues. The high-resolution mobility analysis, aided by site-directed mutagenesis ([P8A], [P11A], [P12A], [P8A/P11A], [P8A/P12A], [P11A/P12A], and [P8A/P11A/P12A] variants), demonstrated that Pcn2.8 adopts cis/trans-conformations at Pro8, Pro11, and Pro12 residues. These observations were complementary to recent NMR findings, for which only the Pro8 residue was evidenced to adopt cis/trans-orientations. This study highlights the analytical power of the TIMS-MS/MS workflow for the structural characterization of lanthipeptides and could be a useful tool in our understanding of the biologically important structural elements that drive the thioether cyclization process.

Cutaneous Polymer-Coating Embolism After Endovascular Procedures: Report of Two Cases and a Literature Review.

ABSTRACT: Different hydrophilic and hydrophobic polymers are used as lubricious coatings to reduce vascular traumas in minimally invasive percutaneous procedures. Although they are usually very safe, there is still a risk of serious complications in patients undergoing such procedures, mostly derived from the devices' coating detachment and systemic embolization. The lungs are the most common organ involved, followed by the central nervous system. Yet, cutaneous embolization is unusual, and only 19 cases are available in the literature. Most commonly, they present as asymptomatic retiform purpura on the lower legs, which tends to involve spontaneously. Correct clinical diagnosis is not suspected in most cases, being cholesterol emboly or vasculitis the preferred options. Time interval since surgical procedure and appearance of lesions vary widely but they generally start in the first few days. Histopathological identification of the embolus as bluish, amorphous intraluminal material in dermal vessels is diagnostic, but vasculitic signs are not present. We report 2 cases of skin lesions as the main manifestation of polymer embolization after endovascular surgical procedures. In both cases, biopsy allowed identification of embolized foreign material and lesions resolved without specific treatment.

Multimodal, in Situ Imaging of Ex Vivo Human Skin Reveals Decrease of Cholesterol Sulfate in the Neoepithelium during Acute Wound Healing.

Skin repair is a significant aspect of human health. While the makeup of healthy stratum corneum and epidermis is generally understood, the mobilization of molecular components during skin repair remains largely unknown. In the present work, we utilize multimodal, in situ, mass spectrometry, and immunofluorescence imaging for the characterization of newly formed epidermis, following an initial acute wound for the first 96 h of epithelization. In particular, TOF-SIMS and confirmatory MALDI FT-ICR MS (/MS) analysis permitted the mapping of several lipid classes, including phospholipids, neutral lipids, cholesterol, ceramides, and free fatty acids. Endogenous lipid species were localized in discrete epidermal skin layers, including the stratum corneum (SC), stratum granulosum (SG), stratum basale (SB), and dermis. Experiments revealed that healthy re-epithelializing skin is characterized by diminished cholesterol sulfate signal along the stratum corneum toward the migrating epithelial tongue. The spatial distribution and relative abundances of cholesterol sulfate are reported and correlated with the healing time. The multimodal imaging approach enabled in situ high-confidence chemical mapping based on accurate mass and fragmentation pattern of molecular components. The use of postanalysis immunofluorescence imaging from the same tissue confirmed the localization of endogenous lipid species and Filaggrin and Cav-1 proteins at high spatial resolution (approximately a few microns).

Endoscopic stenting for gastroduodenal outlet obstruction of a malignant origin, real life experience in a single center.

AIM: to evaluate the safety and effectiveness of self-expandable metal stent placement for malignant gastric outlet obstruction (GOO). METHODS: a retrospective, analytic cohort study at a single, tertiary-care center. RESULTS: thirty-six patients that underwent stent placement for GOO of malignant origin were identified during the study period. Technical success was achieved in 36 (100 %) patients and clinical success was achieved in 31 patients (86.1 %). Before the procedure, 17 (54.8 %) patients had a gastric outlet obstruction score (GOOSS) of 0, which is a complete inability of oral intake. Twenty-three patients were alive 30 days after the procedure, two (8.6 %) patients had a GOOSS of 1, ten (43.3 %) had a GOOSS of 2 and eleven (47.9 %) had a GOOSS of 3. Abdominal pain was present in all 31 patients before the procedure and only seven (22.6 %) patients continued with abdominal pain 24 hours after the procedure. During follow-up, ten (30.3 %) patients developed complications related to the stents and none of them was fatal. Additional therapy due to partial occlusion of the stent was necessary in three patients. The stents functional duration had a median of 72 days (IQR 25-75 15-105 days) and was closely related to overall survival. CONCLUSION: palliative stenting for gastroduodenal obstruction is a safe, feasible and effective therapy to treat patients with malignant gastric outlet obstruction.

Biochemical Basis of E. coli Topoisomerase I Relaxation Activity Reduction by Nonenzymatic Lysine Acetylation.

The relaxation activity of E. coli topoisomerase I is required for regulation of global and local DNA supercoiling. The in vivo topoisomerase I enzyme activity is sensitive to lysine acetylation⁻deacetylation and can affect DNA supercoiling and growth as a result. Nonenzymatic lysine acetylation by acetyl phosphate has been shown to reduce the relaxation activity of E. coli topoisomerase I. In this work, the biochemical consequence of topoisomerase I modification by acetyl phosphate with enzymatic assays was studied. Results showed that noncovalent binding to DNA and DNA cleavage by the enzyme were reduced as a result of the acetylation, with greater effect on DNA cleavage. Four lysine acetylation sites were identified using bottom-up proteomics: Lys13, Lys45, Lys346, and Lys488. The Lys13 residue modified by acetyl phosphate has not been reported previously as a lysine acetylation site for E. coli topoisomerase I. We discuss the potential biochemical consequence of lysine acetylation at this strictly conserved lysine and other lysine residues on the enzyme based on available genetic and structural information.

XBP1s Is an Anti-lipogenic Protein.

Endoplasmic reticulum (ER) stress has been shown to contribute to various metabolic diseases, including non-alcoholic fatty liver disease and type 2 diabetes. Reduction of ER stress by treatment with chemical chaperones or overexpression of ER chaperone proteins alleviates hepatic steatosis. Nonetheless, X-box binding protein 1s (XBP1s), a key transcription factor that reduces ER stress, has been proposed as a lipogenic transcription factor. In this report, we document that XBP1s leads to suppression of lipogenic gene expression and reduction of hepatic triglyceride and diacylglycerol content in livers of diet-induced obese and genetically obese and insulin-resistant ob/ob mice. Furthermore, we also show that PKCϵ activity, which correlates with fatty liver and which causes insulin resistance, was significantly reduced in diet-induced obese mice. Finally, we have shown that XBP1s reduces the hepatic fatty acid synthesis rate and enhances macrolipophagy, an initiating step in lipolysis. Our results reveal that XBP1s reduces hepatic lipogenic gene expression and improves hepatosteatosis in mouse models of obesity and insulin resistance, which leads us to conclude that XBP1s has anti-lipogenic properties in the liver.