Molecular Pathology Demonstration of SARS-CoV-2 in Cytotrophoblast from Placental Tissue with Chronic Histiocytic Intervillositis, Trophoblast Necrosis and COVID-19

Journal of developmental biology

2021 Aug 25

Schwartz, DA;Bugatti, M;Santoro, A;Facchetti, F;
PMID: 34449643 | DOI: 10.3390/jdb9030033

A subset of placentas from pregnant women having the SARS-CoV-2 infection have been found to be infected with the coronavirus using molecular pathology methods including immunohistochemistry and RNA in situ hybridization. These infected placentas can demonstrate several unusual findings which occur together-chronic histiocytic intervillositis, trophoblast necrosis and positive staining of the syncytiotrophoblast for SARS-CoV-2. They frequently also have increased fibrin deposition, which can be massive in some cases. Syncytiotrophoblast is the most frequent fetal-derived cell type to be positive for SARS-CoV-2. It has recently been shown that in a small number of infected placentas, villous stromal macrophages, termed Hofbauer cells, and villous capillary endothelial cells can also stain positive for SARS-CoV-2. This report describes a placenta from a pregnant woman with SARS-CoV-2 that had chronic histiocytic intervillositis, trophoblast necrosis, increased fibrin deposition and positive staining of the syncytiotrophoblast for SARS-CoV-2. In addition, molecular pathology testing including RNAscope and immunohistochemistry for SARS-CoV-2 and double-staining immunohistochemistry using antibodies to E-cadherin and GATA3 revealed that cytotrophoblast cells stained intensely for SARS-CoV-2. All of the cytotrophoblast cells that demonstrated positive staining for SARS-CoV-2 were in direct physical contact with overlying syncytiotrophoblast that also stained positive for the virus. The pattern of cytotrophoblast staining for SARS-CoV-2 was patchy, and there were chorionic villi having diffuse positive staining of the syncytiotrophoblast for SARS-CoV-2, but without staining of cytotrophoblast. This first detailed description of cytotrophoblast involvement by SARS-CoV-2 adds another fetal cell type from infected placentas that demonstrate viral staining.

The neurons that restore walking after paralysis

Nature

2022 Nov 01

Kathe, C;Skinnider, MA;Hutson, TH;Regazzi, N;Gautier, M;Demesmaeker, R;Komi, S;Ceto, S;James, ND;Cho, N;Baud, L;Galan, K;Matson, KJE;Rowald, A;Kim, K;Wang, R;Minassian, K;Prior, JO;Asboth, L;Barraud, Q;Lacour, SP;Levine, AJ;Wagner, F;Bloch, J;Squair, JW;Courtine, G;
PMID: 36352232 | DOI: 10.1038/s41586-022-05385-7

A spinal cord injury interrupts pathways from the brain and brainstem that project to the lumbar spinal cord, leading to paralysis. Here we show that spatiotemporal epidural electrical stimulation (EES) of the lumbar spinal cord1-3 applied during neurorehabilitation4,5 (EESREHAB) restored walking in nine individuals with chronic spinal cord injury. This recovery involved a reduction in neuronal activity in the lumbar spinal cord of humans during walking. We hypothesized that this unexpected reduction reflects activity-dependent selection of specific neuronal subpopulations that become essential for a patient to walk after spinal cord injury. To identify these putative neurons, we modelled the technological and therapeutic features underlying EESREHAB in mice. We applied single-nucleus RNA sequencing6-9 and spatial transcriptomics10,11 to the spinal cords of these mice to chart a spatially resolved molecular atlas of recovery from paralysis. We then employed cell type12,13 and spatial prioritization to identify the neurons involved in the recovery of walking. A single population of excitatory interneurons nested within intermediate laminae emerged. Although these neurons are not required for walking before spinal cord injury, we demonstrate that they are essential for the recovery of walking with EES following spinal cord injury. Augmenting the activity of these neurons phenocopied the recovery of walking enabled by EESREHAB, whereas ablating them prevented the recovery of walking that occurs spontaneously after moderate spinal cord injury. We thus identified a recovery-organizing neuronal subpopulation that is necessary and sufficient to regain walking after paralysis. Moreover, our methodology establishes a framework for using molecular cartography to identify the neurons that produce complex behaviours.

A bacterial extracellular vesicle-based intranasal vaccine against SARS-CoV-2 protects against disease and elicits neutralizing antibodies to wild-type and Delta variants

bioRxiv : the preprint server for biology

2022 Feb 01

Jiang, L;Driedonks, TAP;Jong, WSP;Dhakal, S;van den Berg van Saparoea, HB;Sitaras, I;Zhou, R;Caputo, C;Littlefield, K;Lowman, M;Chen, M;Lima, G;Gololobova, O;Smith, B;Mahairaki, V;Richardson, MR;Mulka, KR;Lane, AP;Klein, SL;Pekosz, A;Brayton, CF;Mankowski, JL;Luirink, J;Villano, JS;Witwer, KW;
PMID: 35132418 | DOI: 10.1101/2021.06.28.450181

Several vaccines have been introduced to combat the coronavirus infectious disease-2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Current SARS-CoV-2 vaccines include mRNA-containing lipid nanoparticles or adenoviral vectors that encode the SARS-CoV-2 Spike (S) protein of SARS-CoV-2, inactivated virus, or protein subunits. Despite growing success in worldwide vaccination efforts, additional capabilities may be needed in the future to address issues such as stability and storage requirements, need for vaccine boosters, desirability of different routes of administration, and emergence of SARS-CoV-2 variants such as the Delta variant. Here, we present a novel, well-characterized SARS-CoV-2 vaccine candidate based on extracellular vesicles (EVs) of Salmonella typhimurium that are decorated with the mammalian cell culture-derived Spike receptor-binding domain (RBD). RBD-conjugated outer membrane vesicles (RBD-OMVs) were used to immunize the golden Syrian hamster ( Mesocricetus auratus ) model of COVID-19. Intranasal immunization resulted in high titers of blood anti-RBD IgG as well as detectable mucosal responses. Neutralizing antibody activity against wild-type and Delta variants was evident in all vaccinated subjects. Upon challenge with live virus, hamsters immunized with RBD-OMV, but not animals immunized with unconjugated OMVs or a vehicle control, avoided body mass loss, had lower virus titers in bronchoalveolar lavage fluid, and experienced less severe lung pathology. Our results emphasize the value and versatility of OMV-based vaccine approaches.

Neurovascular injury with complement activation and inflammation in COVID-19

Brain : a journal of neurology

2022 Jul 29

Lee, MH;Perl, DP;Steiner, J;Pasternack, N;Li, W;Maric, D;Safavi, F;Horkayne-Szakaly, I;Jones, R;Stram, MN;Moncur, JT;Hefti, M;Folkerth, RD;Nath, A;
PMID: 35788639 | DOI: 10.1093/brain/awac151

The underlying mechanisms by which severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) leads to acute and long-term neurological manifestations remains obscure. We aimed to characterize the neuropathological changes in patients with coronavirus disease 2019 and determine the underlying pathophysiological mechanisms. In this autopsy study of the brain, we characterized the vascular pathology, the neuroinflammatory changes and cellular and humoral immune responses by immunohistochemistry. All patients died during the first wave of the pandemic from March to July 2020. All patients were adults who died after a short duration of the infection, some had died suddenly with minimal respiratory involvement. Infection with SARS-CoV-2 was confirmed on ante-mortem or post-mortem testing. Descriptive analysis of the pathological changes and quantitative analyses of the infiltrates and vascular changes were performed. All patients had multifocal vascular damage as determined by leakage of serum proteins into the brain parenchyma. This was accompanied by widespread endothelial cell activation. Platelet aggregates and microthrombi were found adherent to the endothelial cells along vascular lumina. Immune complexes with activation of the classical complement pathway were found on the endothelial cells and platelets. Perivascular infiltrates consisted of predominantly macrophages and some CD8+ T cells. Only rare CD4+ T cells and CD20+ B cells were present. Astrogliosis was also prominent in the perivascular regions. Microglial nodules were predominant in the hindbrain, which were associated with focal neuronal loss and neuronophagia. Antibody-mediated cytotoxicity directed against the endothelial cells is the most likely initiating event that leads to vascular leakage, platelet aggregation, neuroinflammation and neuronal injury. Therapeutic modalities directed against immune complexes should be considered.

SARS-CoV2 infects pancreatic beta cells in vivo and induces cellular and subcellular disruptions that reflect beta cell dysfunction

Research square

2021 Jul 20

Millette, K;Cuala, J;Wang, P;Marks, C;Woo, V;Hayun, M;Kang, H;Martin, M;Dhawan, S;Chao, L;Fraser, S;Junge, J;Lewis, M;Georgia, S;
PMID: 34312617 | DOI: 10.21203/rs.3.rs-592374/v1

Increasing evidence of new-onset diabetes during the COVID19 pandemic indicates that the SARS-CoV2 virus may drive beta-cell dysfunction leading to diabetes, but it is unclear if it is a primary or secondary effect. Here, we present evidence of SARS-CoV-2 infection of pancreatic beta cells in vivo using a robust and reproducible non-human primates model of mild to moderate COVID19 pathogenesis. Pancreas from SARS-CoV-2 infected subjects were positive for the SARS-CoV2 spike protein by immunohistochemistry and structures indicative of viral replication were evident by electron microscopy. Total beta cell area was decreased in SARS-CoV-2-infected pancreas, attributable to beta cell atrophy. Beta cell granularity was decreased. These histologic phenotypes persisted beyond the duration of the clinical disease course. Detailed electron microscopy of SARS-CoV-2 infected beta-cells revealed ultrastructural hallmarks of beta cell stress that are seen in islets of patients with Type 2 diabetes, including disrupted mitochondria and dilated endoplasmic reticulum. To assess the metabolic status of beta cells from SARS-CoV-2-infected subjects, we used fluorescence life-time imaging to measure the ratio of free and bound NADH as a surrogate of glycolytic and oxidative metabolism. We report an increase in free NADH levels, suggesting that beta cells from SARS-CoV-2-infected subjects adopt a more glycolytic metabolic profile. Taken together, we conclude that SARS-CoV-2 infection induces beta cell stress that may compromise beta-cell function beyond the duration of the disease course. This raises the possibility that the beta cell stress and injury may have clinical implications of the long-term future health of patients that have recovered from COVID19.

COVID-19-associated cardiac pathology at post-mortem evaluation: A Collaborative systematic Review

Clinical microbiology and infection : the official publication of the European Society of Clinical Microbiology and Infectious Diseases

2022 Mar 23

Almamlouk, R;Kashour, T;Obeidat, S;Bois, MC;Maleszewski, JJ;Omrani, OA;Tleyjeh, R;Berbari, E;Chakhachiro, Z;Zein-Sabatto, B;Gerberi, D;Tleyjeh, IM;Cardiac Autopsy in COVID-19 Study Group, ;
PMID: 35339672 | DOI: 10.1016/j.cmi.2022.03.021

Many post-mortem studies addressing the cardiovascular effects of COVID-19 provided valuable information but were limited by their small sample size.The aim of this systematic review is to better understand the various aspects of the cardiovascular complications of COVID-19 by pooling data from a large number of autopsy studies.We searched online databases Ovid EBM Reviews, Ovid Embase, Ovid Medline, Scopus and Web of Science for the concepts of autopsy or histopathology combined with COVID-19 published between database inception to February 2021. We also searched for unpublished manuscripts using the medRxiv services operated by Cold Spring Harbor Laboratory.Articles were considered eligible for inclusion if they reported human post-mortem cardiovascular findings among individuals with confirmed SARS-CoV-2 infection.Studies were individually assessed for risk of selection bias, detection bias and reporting bias.Median prevalence of different autopsy findings with associated interquartile ranges.This review cohort contained 50 studies including 548 hearts. The median age of the deceased was 69 years. The most prevalent acute cardiovascular findings were myocardial necrosis (median=100.0%, IQR 20-100%, number of studies=9, number of patients=64) and myocardial edema (median=55.5%, IQR 19.5-92.5%, number of studies=4, number of patients=46). Median reported prevalence's of extensive, focal active and multifocal myocarditis were all 0.0%. The most prevalent chronic changes were myocyte hypertrophy (median=69.0%, IQR 46.8-92.1%) and fibrosis (median=35.0%, IQR 35.0-90.5%). SARS-CoV-2 was detected in the myocardium with median prevalence of 60.8% (IQR 40.4-95.6%).Our systematic review confirmed the high prevalence of acute and chronic cardiac pathologies in COVID-19, SARS-CoV-2 cardiac tropism, and the low prevalence of myocarditis in COVID-19 disease.

Fluoxetine plus lithium for treatment of mental health impairment in Long Covid

Qeios

2022 Sep 21

Fessel, J;
| DOI: 10.32388/cf8mip

Mental disability is a serious and often disabling symptom of Long Covid, for which currently there is no recommendable pharmacotherapy for those patients whose response to psychotherapy is suboptimal. Treatment could be formulated by using drugs that address the brain cell-types that have been demonstrated as dominantly affected in Long Covid. Those cell-types are astrocytes, oligodendrocytes, endothelial cells/pericytes, and microglia. Lithium and fluoxetine each address all of those four cell-types. They should be administered in combination for both depth of benefit and reduction of dosages. Low dosage of each is likely to be well-tolerated and to cause neither adverse events (AE) nor serious adverse events (SAE).

Non-Productive Infection of Glial Cells with SARS-CoV-2 in Hamster Organotypic Cerebellar Slice Cultures

Viruses

2022 Jun 03

Lamoureux, L;Sajesh, B;Slota, JA;Medina, SJ;Mayor, M;Frost, KL;Warner, B;Manguiat, K;Wood, H;Kobasa, D;Booth, SA;
PMID: 35746689 | DOI: 10.3390/v14061218

The numerous neurological syndromes associated with COVID-19 implicate an effect of viral pathogenesis on neuronal function, yet reports of direct SARS-CoV-2 infection in the brain are conflicting. We used a well-established organotypic brain slice culture to determine the permissivity of hamster brain tissues to SARS-CoV-2 infection. We found levels of live virus waned after inoculation and observed no evidence of cell-to-cell spread, indicating that SARS-CoV-2 infection was non-productive. Nonetheless, we identified a small number of infected cells with glial phenotypes; however, no evidence of viral infection or replication was observed in neurons. Our data corroborate several clinical studies that have assessed patients with COVID-19 and their association with neurological involvement.

SARS-CoV-2 infection in hamsters and humans results in lasting and unique systemic perturbations post recovery

Science Translational Medicine

2022 Jun 07

Frere, J;Serafini, R;Pryce, K;Zazhytska, M;Oishi, K;Golynker, I;Panis, M;Zimering, J;Horiuchi, S;Hoagland, D;Møller, R;Ruiz, A;Kodra, A;Overdevest, J;Canoll, P;Borczuk, A;Chandar, V;Bram, Y;Schwartz, R;Lomvardas, S;Zachariou, V;tenOever, B;
| DOI: 10.1126/scitranslmed.abq3059

The host response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can result in prolonged pathologies collectively referred to as post-acute sequalae of COVID-19 (PASC) or long COVID. To better understand the mechanism underlying long COVID biology, we compared the short- and long-term systemic responses in the golden hamster following either SARS-CoV-2 or influenza A virus (IAV) infection. Results demonstrated that SARS-CoV-2 exceeded IAV in its capacity to cause permanent injury to the lung and kidney and uniquely impacted the olfactory bulb (OB) and epithelium (OE). Despite a lack of detectable infectious virus, the OB and OE demonstrated myeloid and T cell activation, proinflammatory cytokine production, and an interferon response that correlated with behavioral changes extending a month post viral clearance. These sustained transcriptional changes could also be corroborated from tissue isolated from individuals who recovered from COVID-19. These data highlight a molecular mechanism for persistent COVID-19 symptomology and provide a small animal model to explore future therapeutics.

The Omicron Sub-Variant BA.4 Displays a Remarkable Lack of Clinical Signs in a Golden Syrian Hamster Model of SARS-CoV-2 Infection

Viruses

2023 May 10

Davies, ER;Ryan, KA;Bewley, KR;Coombes, NS;Salguero, FJ;Carnell, OT;Biddlecombe, S;Charlton, M;Challis, A;Cross, ES;Handley, A;Ngabo, D;Weldon, TM;Hall, Y;Funnell, SGP;
PMID: 37243219 | DOI: 10.3390/v15051133

The ongoing emergence of SARS-CoV-2 virus variants remains a source of concern because it is accompanied by the potential for increased virulence as well as evasion of immunity. Here we show that, although having an almost identical spike gene sequence as another Omicron variant (BA.5.2.1), a BA.4 isolate lacked all the typical disease characteristics of other isolates seen in the Golden Syrian hamster model despite replicating almost as effectively. Animals infected with BA.4 had similar viral shedding profiles to those seen with BA.5.2.1 (up to day 6 post-infection), but they all failed to lose weight or present with any other significant clinical signs. We hypothesize that this lack of detectable signs of disease during infection with BA.4 was due to a small (nine nucleotide) deletion (∆686-694) in the viral genome (ORF1ab) responsible for the production of non-structural protein 1, which resulted in the loss of three amino acids (aa 141-143).

A Potential Novel Treatment for Chronic Cough in Long COVID Patients: Clearance of Epipharyngeal Residual SARS-CoV-2 Spike RNA by Epipharyngeal Abrasive Therapy

Cureus

2023 Jan 01

Nishi, K;Yoshimoto, S;Tanaka, T;Kimura, S;Shinchi, Y;Yamano, T;
PMID: 36618501 | DOI: 10.7759/cureus.33421

A major target of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the epipharyngeal mucosa. Epipharyngeal abrasive therapy (EAT) is a Japanese treatment for chronic epipharyngitis. EAT is a treatment for chronic epipharyngitis in Japan that involves applying zinc chloride as an anti-inflammatory agent to the epipharyngeal mucosa. Here, we present a case of a 21-year-old man with chronic coughing that persisted for four months after a diagnosis of mild coronavirus disease 2019 (COVID-19), who was treated by EAT. We diagnosed chronic epipharyngitis as the cause of the chronic cough after the SARS-CoV-2 infection. SARS-CoV-2 spike RNA had persisted in the epipharyngeal mucosa of this Long COVID patient. EAT was performed once a week for three months, which eliminated residual SARS-CoV-2 RNA and reduced epipharyngeal inflammation. Moreover, a reduction in the expression of proinflammatory cytokines was found by histopathological examination. We speculate that the virus was excreted with the drainage induced by EAT, which stopped the secretion of proinflammatory cytokines. This case study suggests that EAT is a useful treatment for chronic epipharyngitis involving long COVID.

Intranasal delivery of a rationally attenuated SARS-CoV-2 is immunogenic and protective in Syrian hamsters

Nature communications

2022 Nov 10

Liu, S;Stauft, CB;Selvaraj, P;Chandrasekaran, P;D'Agnillo, F;Chou, CK;Wu, WW;Lien, CZ;Meseda, CA;Pedro, CL;Starost, MF;Weir, JP;Wang, TT;
PMID: 36357440 | DOI: 10.1038/s41467-022-34571-4

Few live attenuated severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines are in pre-clinical or clinical development. We seek to attenuate SARS-CoV-2 (isolate WA1/2020) by removing the polybasic insert within the spike protein and the open reading frames (ORFs) 6-8, and by introducing mutations that abolish non-structural protein 1 (Nsp1)-mediated toxicity. The derived virus (WA1-ΔPRRA-ΔORF6-8-Nsp1K164A/H165A) replicates to 100- to 1000-fold-lower titers than the ancestral virus and induces little lung pathology in both K18-human ACE2 (hACE2) transgenic mice and Syrian hamsters. Immunofluorescence and transcriptomic analyses of infected hamsters confirm that three-pronged genetic modifications attenuate the proinflammatory pathways more than the removal of the polybasic cleavage site alone. Finally, intranasal administration of just 100 PFU of the WA1-ΔPRRA-ΔORF6-8-Nsp1K164A/H165A elicits robust antibody responses in Syrian hamsters and protects against SARS-CoV-2-induced weight loss and pneumonia. As a proof-of-concept study, we demonstrate that live but sufficiently attenuated SARS-CoV-2 vaccines may be attainable by rational design.

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	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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