Argueta, LB;Lacko, LA;Bram, Y;Tada, T;Carrau, L;Rendeiro, AF;Zhang, T;Uhl, S;Lubor, BC;Chandar, V;Gil, C;Zhang, W;Dodson, BJ;Bastiaans, J;Prabhu, M;Houghton, S;Redmond, D;Salvatore, CM;Yang, YJ;Elemento, O;Baergen, RN;tenOever, BR;Landau, NR;Chen, S;Schwartz, RE;Stuhlmann, H;
PMID: 35434541 | DOI: 10.1016/j.isci.2022.104223
The effect of SARS-CoV-2 infection on placental function is not well understood. Analysis of placentas from women who tested positive at delivery showed SARS-CoV-2 genomic and subgenomic RNA in 22 out of 52 placentas. Placentas from two mothers with symptomatic COVID-19 whose pregnancies resulted in adverse outcomes for the fetuses contained high levels of viral Alpha variant RNA. The RNA was localized to the trophoblasts that cover the fetal chorionic villi in direct contact with maternal blood. The intervillous spaces and villi were infiltrated with maternal macrophages and T cells. Transcriptome analysis showed an increased expression of chemokines and pathways associated with viral infection and inflammation. Infection of placental cultures with live SARS-CoV-2 and spike protein-pseudotyped lentivirus showed infection of syncytiotrophoblast and, in rare cases, endothelial cells mediated by ACE2 and Neuropilin-1. Viruses with Alpha, Beta, and Delta variant spikes infected the placental cultures at significantly greater levels.
Nature biomedical engineering
Wang, P;Jin, L;Zhang, M;Wu, Y;Duan, Z;Guo, Y;Wang, C;Guo, Y;Chen, W;Liao, Z;Wang, Y;Lai, R;Lee, LP;Qin, J;
PMID: 37349391 | DOI: 10.1038/s41551-023-01054-w
In some patients, COVID-19 can trigger neurological symptoms with unclear pathogenesis. Here we describe a microphysiological system integrating alveolus and blood-brain barrier (BBB) tissue chips that recapitulates neuropathogenesis associated with infection by SARS-CoV-2. Direct exposure of the BBB chip to SARS-CoV-2 caused mild changes to the BBB, and infusion of medium from the infected alveolus chip led to more severe injuries on the BBB chip, including endothelial dysfunction, pericyte detachment and neuroinflammation. Transcriptomic analyses indicated downregulated expression of the actin cytoskeleton in brain endothelium and upregulated expression of inflammatory genes in glial cells. We also observed early cerebral microvascular damage following lung infection with a low viral load in the brains of transgenic mice expressing human angiotensin-converting enzyme 2. Our findings suggest that systemic inflammation is probably contributing to neuropathogenesis following SARS-CoV-2 infection, and that direct viral neural invasion might not be a prerequisite for this neuropathogenesis. Lung-brain microphysiological systems should aid the further understanding of the systemic effects and neurological complications of viral infection.
Handley, A;Ryan, KA;Davies, ER;Bewley, KR;Carnell, OT;Challis, A;Coombes, NS;Fotheringham, SA;Gooch, KE;Charlton, M;Harris, DJ;Kennard, C;Ngabo, D;Weldon, TM;Salguero, FJ;Funnell, SGP;Hall, Y;
PMID: 36992457 | DOI: 10.3390/v15030748
The golden Syrian hamster (Mesocricetus auratus) is now commonly used in preclinical research for the study of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and the assessment of vaccines, drugs and therapeutics. Here, we show that hamsters inoculated via the intranasal route with the same infectious virus dose of prototypical SARS-CoV-2 administered in a different volume present with different clinical signs, weight loss and viral shedding, with a reduced volume resulting in reduced severity of disease similar to that obtained by a 500-fold reduction in the challenge dose. The tissue burden of the virus and the severity of pulmonary pathology were also significantly affected by different challenge inoculum volumes. These findings suggest that a direct comparison between the severity of SARS-CoV-2 variants or studies assessing the efficacy of treatments determined by hamster studies cannot be made unless both the challenge dose and inoculation volume are matched when using the intranasal route. Additionally, analysis of sub-genomic and total genomic RNA PCR data demonstrated no link between sub-genomic and live viral titres and that sub-genomic analyses do not provide any information beyond that provided by more sensitive total genomic PCR.
Compagnone, M;Pinto, E;Salvatori, E;Lione, L;Conforti, A;Marchese, S;Ravà, M;Ryan, K;Hall, Y;Rayner, E;Salguero, FJ;Paterson, J;Iannacone, M;De Francesco, R;Aurisicchio, L;Palombo, F;
PMID: 35893826 | DOI: 10.3390/vaccines10081178
The COVID-19 pandemic is entering a new era with the approval of many SARS-CoV-2 vaccines. In spite of the restoration of an almost normal way of life thanks to the immune protection elicited by these innovative vaccines, we are still facing high viral circulation, with a significant number of deaths. To further explore alternative vaccination platforms, we developed COVID-eVax-a genetic vaccine based on plasmid DNA encoding the RBD domain of the SARS-CoV-2 spike protein. Here, we describe the correlation between immune responses and the evolution of viral infection in ferrets infected with the live virus. We demonstrate COVID-eVax immunogenicity as means of antibody response and, above all, a significant T-cell response, thus proving the critical role of T-cell immunity, in addition to the neutralizing antibody activity, in controlling viral spread.
Fell, R;Potter, JA;Yuille, S;Salguero, FJ;Watson, R;Ngabo, D;Gooch, K;Hewson, R;Howat, D;Dowall, S;
PMID: 35632718 | DOI: 10.3390/v14050976
The rapid global spread of severe acute respiratory coronavirus 2 (SARS-CoV-2) has resulted in an urgent effort to find efficacious therapeutics. Broad-spectrum therapies which could be used for other respiratory pathogens confer advantages, as do those based on targeting host cells that are not prone to the development of resistance by the pathogen. We tested an intranasally delivered carbohydrate-binding module (CBM) therapy, termed Neumifil, which is based on a CBM that has previously been shown to offer protection against the influenza virus through the binding of sialic acid receptors. Using the recognised hamster model of SARS-CoV-2 infection, we demonstrate that Neumifil significantly reduces clinical disease severity and pathological changes in the nasal cavity. Furthermore, we demonstrate Neumifil binding to the human angiotensin-converting enzyme 2 (ACE2) receptor and spike protein of SARS-CoV-2. This is the first report describing the testing of this type of broad-spectrum antiviral therapy in vivo and provides evidence for the advancement of Neumifil in further preclinical and clinical studies.
Carossino, M;Kenney, D;O'Connell, AK;Montanaro, P;Tseng, AE;Gertje, HP;Grosz, KA;Ericsson, M;Huber, BR;Kurnick, SA;Subramaniam, S;Kirkland, TA;Walker, JR;Francis, KP;Klose, AD;Paragas, N;Bosmann, M;Saeed, M;Balasuriya, UBR;Douam, F;Crossland, NA;
PMID: 35336942 | DOI: 10.3390/v14030535
Animal models recapitulating COVID-19 are critical to enhance our understanding of SARS-CoV-2 pathogenesis. Intranasally inoculated transgenic mice expressing human angiotensin-converting enzyme 2 under the cytokeratin 18 promoter (K18-hACE2) represent a lethal model of SARS-CoV-2 infection. We evaluated the clinical and virological dynamics of SARS-CoV-2 using two intranasal doses (104 and 106 PFUs), with a detailed spatiotemporal pathologic analysis of the 106 dose cohort. Despite generally mild-to-moderate pneumonia, clinical decline resulting in euthanasia or death was commonly associated with hypothermia and viral neurodissemination independent of inoculation dose. Neuroinvasion was first observed at 4 days post-infection, initially restricted to the olfactory bulb suggesting axonal transport via the olfactory neuroepithelium as the earliest portal of entry. Absence of viremia suggests neuroinvasion occurs independently of transport across the blood-brain barrier. SARS-CoV-2 tropism was neither restricted to ACE2-expressing cells (e.g., AT1 pneumocytes), nor inclusive of some ACE2-positive cell lineages (e.g., bronchiolar epithelium and brain vasculature). Absence of detectable ACE2 protein expression in neurons but overexpression in neuroepithelium suggest this as the most likely portal of neuroinvasion, with subsequent ACE2 independent lethal neurodissemination. A paucity of epidemiological data and contradicting evidence for neuroinvasion and neurodissemination in humans call into question the translational relevance of this model.
International journal of infectious diseases : IJID : official publication of the International Society for Infectious Diseases
Märkl, B;Dintner, S;Schaller, T;Sipos, E;Kling, E;Miller, S;Farfan, F;Grochowski, P;Reitsam, N;Waidhauser, J;Hirschbühl, K;Spring, O;Fuchs, A;Wibmer, T;Boor, P;Beer, M;Wylezich, C;
PMID: 36584746 | DOI: 10.1016/j.ijid.2022.12.029
Omicron lineages BA.1/2 are considered to cause mild clinical courses. Nevertheless, fatal cases after those infections are recognized but little is known about risk factors.Twenty-three full and three partial autopsies in deceased with known Omicron BA.1/2 infections have been consecutively performed. The investigations included histology, blood analyses and molecular virus detection.COVID-19-associated diffuse alveolar damage (DAD) was found in only eight cases (31%). This rate is significantly lower compared to previous studies, including non-Omicron variants, where rates between 69% and 92% were observed. Neither vaccination nor known risk factors were significantly associated with a direct cause of death by COVID-19. Only those patients who were admitted to the clinic due to COVID-19 but not for other reasons had a significant association with a direct COVID-19 caused death (P > 0.001).).DAD still occurred in the Omicron BA.1/BA.2 era but at considerably lower frequency than seen with previous variants of concern. None of the known risk factors discriminated the cases with COVID-19-caused death from those that died due to a different disease. Therefore, the host's genomics might play a key role in this regard. Further studies should elucidate the existence of such a genomic risk factor.
Ali Marandi Ghoddousi, R;Magalong, VM;Kamitakahara, AK;Levitt, P;
PMID: 36313803 | DOI: 10.1016/j.crmeth.2022.100316
Spatial gene expression, achieved classically through in situ hybridization, is a fundamental tool for topographic phenotyping of cell types in the nervous system. Newly developed techniques allow for visualization of multiple mRNAs at single-cell resolution and greatly expand the ability to link gene expression to tissue topography, yet there are challenges in efficient quantification and analysis of these high-dimensional datasets. We have therefore developed the single-cell automated multiplex pipeline for RNA (SCAMPR), facilitating rapid and accurate segmentation of neuronal cell bodies using a dual immunohistochemistry-RNAscope protocol and quantification of low- and high-abundance mRNA signals using open-source image processing and automated segmentation tools. Proof of principle using SCAMPR focused on spatial mapping of gene expression by peripheral (vagal nodose) and central (visual cortex) neurons. The analytical effectiveness of SCAMPR is demonstrated by identifying the impact of early life stress on gene expression in vagal neuron subtypes.
Biering, SB;Sarnik, SA;Wang, E;Zengel, JR;Leist, SR;Schäfer, A;Sathyan, V;Hawkins, P;Okuda, K;Tau, C;Jangid, AR;Duffy, CV;Wei, J;Gilmore, RC;Alfajaro, MM;Strine, MS;Nguyenla, X;Van Dis, E;Catamura, C;Yamashiro, LH;Belk, JA;Begeman, A;Stark, JC;Shon, DJ;Fox, DM;Ezzatpour, S;Huang, E;Olegario, N;Rustagi, A;Volmer, AS;Livraghi-Butrico, A;Wehri, E;Behringer, RR;Cheon, DJ;Schaletzky, J;Aguilar, HC;Puschnik, AS;Button, B;Pinsky, BA;Blish, CA;Baric, RS;O'Neal, WK;Bertozzi, CR;Wilen, CB;Boucher, RC;Carette, JE;Stanley, SA;Harris, E;Konermann, S;Hsu, PD;
PMID: 35879412 | DOI: 10.1038/s41588-022-01131-x
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a range of symptoms in infected individuals, from mild respiratory illness to acute respiratory distress syndrome. A systematic understanding of host factors influencing viral infection is critical to elucidate SARS-CoV-2-host interactions and the progression of Coronavirus disease 2019 (COVID-19). Here, we conducted genome-wide CRISPR knockout and activation screens in human lung epithelial cells with endogenous expression of the SARS-CoV-2 entry factors ACE2 and TMPRSS2. We uncovered proviral and antiviral factors across highly interconnected host pathways, including clathrin transport, inflammatory signaling, cell-cycle regulation, and transcriptional and epigenetic regulation. We further identified mucins, a family of high molecular weight glycoproteins, as a prominent viral restriction network that inhibits SARS-CoV-2 infection in vitro and in murine models. These mucins also inhibit infection of diverse respiratory viruses. This functional landscape of SARS-CoV-2 host factors provides a physiologically relevant starting point for new host-directed therapeutics and highlights airway mucins as a host defense mechanism.
Findlay-Wilson, S;Easterbrook, L;Smith, S;Pope, N;Humphries, G;Schuhmann, H;Ngabo, D;Rayner, E;Otter, AD;Coleman, T;Hicks, B;Graham, VA;Halkerston, R;Apostolakis, K;Taylor, S;Fotheringham, S;Horton, A;Tree, JA;Wand, M;Hewson, R;Dowall, SD;
PMID: 35533779 | DOI: 10.1016/j.antiviral.2022.105332
Antibodies against SARS-CoV-2 are important to generate protective immunity, with convalescent plasma one of the first therapies approved. An alternative source of polyclonal antibodies suitable for upscaling would be more amendable to regulatory approval and widespread use. In this study, sheep were immunised with SARS-CoV-2 whole spike protein or one of the subunit proteins: S1 and S2. Once substantial antibody titres were generated, plasma was collected and samples pooled for each antigen. Non-specific antibodies were removed via affinity-purification to yield candidate products for testing in a hamster model of SARS-CoV-2 infection. Affinity-purified polyclonal antibodies to whole spike, S1 and S2 proteins were evaluated for in vitro for neutralising activity against SARS-CoV-2 Wuhan-like virus (Australia/VIC01/2020) and a recent variant of concern, B.1.1.529 BA.1 (Omicron), antibody-binding, complement fixation and phagocytosis assays were also performed. All antibody preparations demonstrated an effect against SARS-CoV-2 disease in the hamster model of challenge, with those raised against the S2 subunit providing the most promise. A rapid, cost-effective therapy for COVID-19 was developed which provides a source of highly active immunoglobulin specific to SARS-CoV-2 with multi-functional activity.Crown
Lyra E Silva, NM;Barros-Aragão, FGQ;De Felice, FG;Ferreira, ST;
PMID: 35257690 | DOI: 10.1016/j.neuropharm.2022.109023
Acute neurological alterations have been associated with SARS-CoV-2 infection. Additionally, it is becoming clear that coronavirus disease 2019 (COVID-19) survivors may experience long-term neurological abnormalities, including cognitive deficits and mood alterations. The mechanisms underlying acute and long-term impacts of COVID-19 in the brain are being actively investigated. Due to the heterogeneous manifestations of neurological outcomes, it is possible that different mechanisms operate following SARS-CoV-2 infection, which may include direct brain infection by SARS-CoV-2, mechanisms resulting from hyperinflammatory systemic disease, or a combination of both. Inflammation is a core feature of COVID-19, and both central and systemic inflammation are known to lead to acute and persistent neurological alterations in other diseases. Here, we review evidence indicating that COVID-19 is associated with neuroinflammation, along with blood-brain barrier dysfunction. Similar neuroinflammatory signatures have been associated with Alzheimer's disease and major depressive disorder. Current evidence demonstrates that patients with pre-existing cognitive and neuropsychiatric deficits show worse outcomes upon infection by SARS-CoV-2 and, conversely, COVID-19 survivors may be at increased risk of developing dementia and mood disorders. Considering the high prevalence of COVID-19 patients that recovered from infection in the world and the alarming projections for the prevalence of dementia and depression, investigation of possible molecular similarities between those diseases may shed light on mechanisms leading to long-term neurological abnormalities in COVID-19 survivors.
Smith, KD;Prince, DK;Henriksen, K;Nicosia, RF;Alpers, CE;Akilesh, S;
PMID: 35227689 | DOI: 10.1016/j.kint.2022.01.033
Collapsing glomerulopathy is a histologically distinct variant of focal and segmental glomerulosclerosis that presents with heavy proteinuria and portends a poor prognosis. Collapsing glomerulopathy can be triggered by viral infections such as HIV or SARS-CoV-2. Transcriptional profiling of collapsing glomerulopathy lesions is difficult since only a few glomeruli may exhibit this histology within a kidney biopsy and the mechanisms driving this heterogeneity are unknown. Therefore, we used recently developed digital spatial profiling (DSP) technology which permits quantification of mRNA at the level of individual glomeruli. Using DSP, we profiled 1,852 transcripts in glomeruli isolated from formalin fixed paraffin embedded sections from HIV or SARS-CoV-2 infected patients with biopsy-confirmed collapsing glomerulopathy and used normal biopsy sections as controls. Even though glomeruli with collapsing features appeared histologically similar across both groups of patients by light microscopyhe increased resolution of DSP uncovered intra- and inter-patient heterogeneity in glomerular transcriptional profiles that were missed in early laser capture microdissection studies of pooled glomeruli. Focused validation using immunohistochemistry and RNA in situ hybridization showed good concordance with DSP results. Thus, DSP represents a powerful method to dissect transcriptional programs of pathologically discernible kidney lesions.
