McDonald, JT;Enguita, FJ;Taylor, D;Griffin, RJ;Priebe, W;Emmett, MR;Sajadi, MM;Harris, AD;Clement, J;Dybas, JM;Aykin-Burns, N;Guarnieri, JW;Singh, LN;Grabham, P;Baylin, SB;Yousey, A;Pearson, AN;Corry, PM;Saravia-Butler, A;Aunins, TR;Sharma, S;Nagpal, P;Meydan, C;Foox, J;Mozsary, C;Cerqueira, B;Zaksas, V;Singh, U;Wurtele, ES;Costes, SV;Davanzo, GG;Galeano, D;Paccanaro, A;Meinig, SL;Hagan, RS;Bowman, NM;UNC COVID-19 Pathobiology Consortium, ;Wolfgang, MC;Altinok, S;Sapoval, N;Treangen, TJ;Moraes-Vieira, PM;Vanderburg, C;Wallace, DC;Schisler, JC;Mason, CE;Chatterjee, A;Meller, R;Beheshti, A;
PMID: 34624208 | DOI: 10.1016/j.celrep.2021.109839
MicroRNAs (miRNAs) are small non-coding RNAs involved in post-transcriptional gene regulation that have a major impact on many diseases and provide an exciting avenue toward antiviral therapeutics. From patient transcriptomic data, we determined that a circulating miRNA, miR-2392, is directly involved with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) machinery during host infection. Specifically, we show that miR-2392 is key in driving downstream suppression of mitochondrial gene expression, increasing inflammation, glycolysis, and hypoxia, as well as promoting many symptoms associated with coronavirus disease 2019 (COVID-19) infection. We demonstrate that miR-2392 is present in the blood and urine of patients positive for COVID-19 but is not present in patients negative for COVID-19. These findings indicate the potential for developing a minimally invasive COVID-19 detection method. Lastly, using in vitro human and in vivo hamster models, we design a miRNA-based antiviral therapeutic that targets miR-2392, significantly reduces SARS-CoV-2 viability in hamsters, and may potentially inhibit a COVID-19 disease state in humans.
Evidence for residual SARS-CoV-2 in glioblastoma tissue of a convalescent patient
Lei, J;Liu, Y;Xie, T;Yao, G;Wang, G;Diao, B;Song, J;
PMID: 33994523 | DOI: 10.1097/WNR.0000000000001654
Since coronavirus disease 2019 (COVID-19) swept all over the world, several studies have shown the susceptibility of a patient with cancer to COVID-19. In this case, the removed glioblastoma multiforme (GBM)-adjacent (GBM-A), GBM-peritumor and GBM-central (GBM-C) tissues from a convalescent patient of COVID-19, who also suffered from glioblastoma meanwhile, together with GBM-A and GBM tissues from a patient without COVID-19 history as negative controls, were used for RNA ISH, electron microscopy observing and immunohistochemical staining of ACE2 and the virus antigen (N protein). The results of RNA ISH, electron microscopy observing showed that SARS-CoV-2 directly infects some cells within human GBM tissues and SARS-CoV-2 in GBM-C tissue still exists even when it is cleared elsewhere. Immunohistochemical staining of ACE2 and N protein showed that the expressions of ACE2 are significantly higher in specimens, including GBM-C tissue from COVID-19 patient than other types of tissue. The unique phenomenon suggests that the surgical protection level should be upgraded even if the patient is in a convalescent period and the pharyngeal swab tests show negative results. Furthermore, more attention should be paid to confirm whether the shelter-like phenomenon happens in other malignancies due to the similar microenvironment and high expression of ACE2 in some malignancies.
Kidney International (2016).
Madan B, Patel MB, Zhang J, Bunte RM, Rudemiller NP, Griffiths R, Virshup DM, Crowley SD.
PMID: - | DOI: 10.1016/j.kint.2016.01.017
Activated Wnt signaling is critical in the pathogenesis of renal fibrosis, a final common pathway for most forms of chronic kidney disease. Therapeutic intervention by inhibition of individual Wnts or downstream Wnt/β-catenin signaling has been proposed, but these approaches do not interrupt the functions of all Wnts nor block non-canonical Wnt signaling pathways. Alternatively, an orally bioavailable small molecule, Wnt-C59, blocks the catalytic activity of the Wnt-acyl transferase porcupine, and thereby prevents secretion of all Wnt isoforms. We found that inhibiting porcupine dramatically attenuates kidney fibrosis in the murine unilateral ureteral obstruction model. Wnt-C59 treatment similarly blunts collagen mRNA expression in the obstructed kidney. Consistent with its actions to broadly arrest Wnt signaling, porcupine inhibition reduces expression of Wnt target genes and bolsters nuclear exclusion of β-catenin in the kidney following ureteral obstruction. Importantly, prevention of Wnt secretion by Wnt-C59 blunts expression of inflammatory cytokines in the obstructed kidney that otherwise provoke a positive feedback loop of Wnt expression in collagen-producing fibroblasts and epithelial cells. Thus, therapeutic targeting of porcupine abrogates kidney fibrosis not only by overcoming the redundancy of individual Wnt isoforms but also by preventing upstream cytokine-induced Wnt generation. These findings reveal a novel therapeutic maneuver to protect the kidney from fibrosis by interrupting a pathogenic crosstalk loop between locally generated inflammatory cytokines and the Wnt/β-catenin signaling pathway.
Journal of neuropathology and experimental neurology
Normandin, E;Valizadeh, N;Rudmann, EA;Uddin, R;Dobbins, ST;MacInnis, BL;Padera, RF;Siddle, KJ;Lemieux, JE;Sabeti, PC;Mukerji, SS;Solomon, IH;
PMID: 36847705 | DOI: 10.1093/jnen/nlad015
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is continually evolving resulting in variants with increased transmissibility, more severe disease, reduced effectiveness of treatments or vaccines, or diagnostic detection failure. The SARS-CoV-2 Delta variant (B.1.617.2 and AY lineages) was the dominant circulating strain in the United States from July to mid-December 2021, followed by the Omicron variant (B.1.1.529 and BA lineages). Coronavirus disease 2019 (COVID-19) has been associated with neurological sequelae including loss of taste/smell, headache, encephalopathy, and stroke, yet little is known about the impact of viral strain on neuropathogenesis. Detailed postmortem brain evaluations were performed for 22 patients from Massachusetts, including 12 who died following infection with Delta variant and 5 with Omicron variant, compared to 5 patients who died earlier in the pandemic. Diffuse hypoxic injury, occasional microinfarcts and hemorrhage, perivascular fibrinogen, and rare lymphocytes were observed across the 3 groups. SARS-CoV-2 protein and RNA were not detected in any brain samples by immunohistochemistry, in situ hybridization, or real-time quantitative PCR. These results, although preliminary, demonstrate that, among a subset of severely ill patients, similar neuropathological features are present in Delta, Omicron, and non-Delta/non-Omicron variant patients, suggesting that SARS-CoV-2 variants are likely to affect the brain by common neuropathogenic mechanisms.
Gynecologic and obstetric investigation
Shen, WB;Turan, S;Wang, B;Cojocaru, L;Harman, C;Logue, J;Reece, EA;Frieman, MB;Yang, P;
PMID: 35526532 | DOI: 10.1159/000524905
Studies indicate a very low rate of SARS-CoV-2 detection in the placenta or occasionally a low rate of vertical transmission in COVID-19 pregnancy. SARS-CoV-2 Delta variant has become a dominant strain over the world and possesses higher infectivity due to mutations in its spike receptor-binding motif.To determine whether SARS-CoV-2 Delta variant has increased potential for placenta infection and vertical transmission, we analyzed SARS-CoV-2 infection in the placenta, umbilical cord, and fetal membrane from a case that unvaccinated mother and her neonate were COVID-19 positive. A 35-year-old primigravida with COVID-19 underwent an emergent cesarean delivery due to placental abruption in the setting of premature rupture of membranes. The neonate tested positive for SARS-CoV-2 within the first 24 hours, and then again on days of life 2, 6, 13, and 21. The placenta exhibited intervillositis, increased fibrin deposition, and syncytiotrophoblast necrosis. Sequencing of viral RNA from fixed placental tissue revealed SAR-CoV-2 B.1.167.2 (Delta) variant. Both spike protein and viral RNA were abundantly present in syncytiotrophoblasts, cytotrophoblasts, umbilical cord vascular endothelium, and fetal membranes.We report with strong probability the first SARS-CoV-2 Delta variant transplacental transmission. Placental cells exhibited extensive apoptosis, senescence, and ferroptosis after SARS-CoV-2 Delta infection.S. Karger AG, Basel.
Iwanaga, N;Cooper, L;Rong, L;Maness, NJ;Beddingfield, B;Qin, Z;Crabtree, J;Tripp, RA;Yang, H;Blair, R;Jangra, S;García-Sastre, A;Schotsaert, M;Sruti, C;Robinson, JE;Srivastava, A;Rabito, F;Qin, X;Kolls, JK;
PMID: 34957381 | DOI: 10.1016/j.isci.2021.103670
SARS-CoV-2, the etiologic agent of COVID-19, uses ACE2 as a cell entry receptor. Soluble ACE2 has been shown to have neutralizing antiviral activity but has a short half-life and no active transport mechanism from the circulation into the alveolar spaces of the lung. To overcome this, we constructed an ACE2-human IgG1 fusion protein with mutations in the catalytic domain of ACE2. A mutation in the catalytic domain of ACE2, MDR504, significantly increased binding to SARS-CoV-2 spike protein, as well as to a spike variant, in vitro with more potent viral neutralization in plaque assays. Parental administration of the protein showed stable serum concentrations with excellent bioavailability in the epithelial lining fluid of the lung, and ameliorated lung SARS-CoV-2 infection in vivo. These data support that the MDR504 hACE2-Fc is an excellent candidate for treatment or prophylaxis of COVID-19 and potentially emerging variants.
Intravenous, Intratracheal, and Intranasal Inoculation of Swine with SARS-CoV-2
Buckley, A;Falkenberg, S;Martins, M;Laverack, M;Palmer, MV;Lager, K;Diel, DG;
PMID: 34452371 | DOI: 10.3390/v13081506
Since the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the susceptibility of animals and their potential to act as reservoirs or intermediate hosts for the virus has been of significant interest. Pigs are susceptible to multiple coronaviruses and have been used as an animal model for other human infectious diseases. Research groups have experimentally challenged swine with human SARS-CoV-2 isolates with results suggesting limited to no viral replication. For this study, a SARS-CoV-2 isolate obtained from a tiger which is identical to human SARS-CoV-2 isolates detected in New York City and contains the D614G S mutation was utilized for inoculation. Pigs were challenged via intravenous, intratracheal, or intranasal routes of inoculation (n = 4/route). No pigs developed clinical signs, but at least one pig in each group had one or more PCR positive nasal/oral swabs or rectal swabs after inoculation. All pigs in the intravenous group developed a transient neutralizing antibody titer, but only three other challenged pigs developed titers greater than 1:8. No gross or histologic changes were observed in tissue samples collected at necropsy. In addition, no PCR positive samples were positive by virus isolation. Inoculated animals were unable to transmit virus to naïve contact animals. The data from this experiment as well as from other laboratories supports that swine are not likely to play a role in the epidemiology and spread of SARS-CoV-2.
The Skin as a critical window in unveiling the pathophysiologic principles of COVID-19
Magro, C;Nuovo, G;Mulvey, J;Laurence, J;Harp, J;Neil Crowson, A;
| DOI: 10.1016/j.clindermatol.2021.07.001
The severe acute respiratory distress syndrome-associated coronavirus-2 (SARS-CoV-2), the etiologic agent of Coronavirus disease 2019 (COVID-19), is a single-stranded RNA virus whose sequence is known. COVID-19 is associated with a heterogeneous clinical phenotype ranging from asymptomatic to fatal disease. It appears that access to nasopharyngeal respiratory epithelia expressing angiotensin-converting enzyme (ACE) 2, the receptor for SARS CoV-2, is followed by viral replication in the pulmonary alveolar septal capillary bed. We have shown in prior studies that incomplete viral particles, termed pseudovirions, dock to deep subcutaneous and other vascular beds potentially contributing to the prothrombotic state and systemic complement activation that characterizes severe and critical COVID-19. A variety of skin rashes have been described in the setting of SARS-CoV-2 infection and more recently, following COVID-19 vaccination. The vaccines deliver a laboratory synthesized mRNA that encodes a protein that is identical to the spike glycoprotein of SARS-COV-2 allowing the production of immunogenic spike glycoprotein that will then elicit T cell and B cell adaptive immune responses. In this paper we review an array of cutaneous manifestations of COVID-19 that provide an opportunity to study critical pathophysiologic mechanisms that underlie all clinical facets of COVID-19 ranging from asymptomatic/mild to severe and critical COVID-19. We classify cutaneous COVID-19 according to underlying pathophysiologic principles. In this regard we propose two main pathways: 1) complement mediated thrombotic vascular injury syndromes deploying the alternative and mannan binding lectin pathways in the setting of severe and critical COVID-19 and 2) the robust T cell and type I interferon driven inflammatory and humoral driven immune complex mediated vasculitic cutaneous reactions seen with mild and moderate COVID-19. Novel data on cutaneous vaccine reactions are presented that manifest a clinical and morphologic parallel with similar eruptions seen in patients suffering from mild and moderate COVID-19 and in most cases represent systemic eczematoid hypersensitivity reactions to a putative vaccine based antigen. Finally, we show for the first time the localization of human synthesized spike glycoprotein following the COVID-19 vaccine to the cutaneous and subcutaneous vasculature confirming the ability of SARS CoV-2 spike glycoprotein to bind endothelium in the absence of intact virus.
The American journal of pathology
Nida Sen, H;Vannella, KM;Wang, Y;Chung, JY;Kodati, S;Ramelli, SC;Lee, JW;Perez, P;Stein, SR;Grazioli, A;Dickey, JM;Ylaya, K;Singh, M;Yinda, KC;Platt, A;Ramos-Benitez, MJ;Zerbe, C;Munster, VJ;de Wit, E;Warner, BM;Herr, DL;Rabin, J;Saharia, KK;NIH COVID-19 Autopsy Consortium, ;Kleiner, DE;Hewitt, SM;Chan, CC;Chertow, DS;
PMID: 36963628 | DOI: 10.1016/j.ajpath.2023.02.016
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. For this current study, contralateral eyes from 21 patients were fixed in formalin and subject to histopathologic examination. From the other four patients, sections of the droplet digital PCR-positive eyes 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, within ocular tissues, a range of common histopathologic alterations were identified, 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.
Knott, D;Fell, R;Potter, JA;Yuille, S;Salguero, FJ;Graham, VA;Hewson, R;Howat, D;Dowall, SD;
PMID: 36992434 | DOI: 10.3390/v15030725
The emergence of severe acute respiratory syndrome coronavirus (SARS-CoV-2) and its expansion to a worldwide pandemic resulted in efforts to assess and develop interventions to reduce the disease burden. Despite the introduction of vaccine programmes against SARS-CoV-2, global incidence levels in early 2022 remained high, demonstrating a need for the development of physiologically relevant models, which are essential for the identification of alternative antiviral strategies. The hamster model of SARS-CoV-2 infection has been widely adopted due to similarities with humans in terms of host cell entry mechanism (via ACE2), and aspects of symptomology and virus shedding. We have previously described a natural transmission hamster model that better represents the natural course of infection. In the present study, we have conducted further testing of the model using the first-in-class antiviral Neumifil, which has previously shown promise against SARS-CoV-2 after a direct intranasal challenge. Neumifil is an intranasally delivered carbohydrate-binding module (CBM) which reduces the binding of viruses to their cellular receptor. By targeting the host cell, Neumifil has the potential to provide broad protection against multiple pathogens and variants. This study demonstrates that using a combination of a prophylactic and therapeutic delivery of Neumifil significantly reduces the severity of clinical signs in animals infected via a natural route of transmission and indicates a reduction of viral loads in the upper respiratory tract. Further refinements of the model are required in order to ensure the adequate transmission of the virus. However, our results provide additional data to the evidence base of Neumifil efficacy against respiratory virus infection and demonstrate that the transmission model is a potentially valuable tool for testing antiviral compounds against SARS-CoV-2.
Proc Natl Acad Sci U S A.
Xing L, Anbarchian T, Tsai JM, Plant GW, Nusse R.
PMID: 29891676 | DOI: 10.1073/pnas.1803297115
In the adult mouse spinal cord, the ependymal cell population that surrounds the central canal is thought to be a promising source of quiescent stem cells to treat spinal cord injury. Relatively little is known about the cellular origin of ependymal cells during spinal cord development, or the molecular mechanisms that regulate ependymal cells during adult homeostasis. Using genetic lineage tracing based on the Wnt target gene Axin2, we have characterized Wnt-responsive cells during spinal cord development. Our results revealed that Wnt-responsive progenitor cells are restricted to the dorsal midline throughout spinal cord development, which gives rise to dorsal ependymal cells in a spatially restricted pattern. This is contrary to previous reports that suggested an exclusively ventral origin of ependymal cells, suggesting that ependymal cells may retain positional identities in relation to their neural progenitors. Our results further demonstrated that in the postnatal and adult spinal cord, all ependymal cells express the Wnt/β-catenin signaling target gene Axin2, as well as Wnt ligands. Genetic elimination of β-catenin or inhibition of Wnt secretion in Axin2-expressing ependymal cells in vivo both resulted in impaired proliferation, indicating that Wnt/β-catenin signaling promotes ependymal cell proliferation. These results demonstrate the continued importance of Wnt/β-catenin signaling for both ependymal cell formation and regulation. By uncovering the molecular signals underlying the formation and regulation of spinal cord ependymal cells, our findings thus enable further targeting and manipulation of this promising source of quiescent stem cells for therapeutic interventions.
Maeda M, Takeshima H, Iida N, Hattori N, Yamashita S, Moro H, Yasukawa Y, Nishiyama K, Hashimoto T, Sekine S, Ishii G, Ochiai A, Fukagawa T, Katai H, Sakai Y, Ushijima T.
PMID: 31085554 | DOI: 10.1136/gutjnl-2018-317645
Abstract
OBJECTIVE:
Cancer-associated fibroblasts (CAFs), a major component of cancer stroma, can confer aggressive properties to cancer cells by secreting multiple factors. Their phenotypes are stably maintained, but the mechanisms are not fully understood. We aimed to show the critical role of epigenetic changes in CAFs in maintaining their tumour-promoting capacity and to show the validity of the epigenomic approach in identifying therapeutic targets from CAFs to starve cancer cells.
DESIGN:
Twelve pairs of primary gastric CAFs and their corresponding non-CAFs (NCAFs) were established from surgical specimens. Genome-wide DNA methylation and H3K27me3 analyses were conducted by BeadArray 450K and ChIP-on-Chip, respectively. Functions of potential a therapeutic target were analysed by inhibiting it, and prognostic impact was assessed in a database.
RESULTS:
CAFs had diverse and distinct DNA methylation and H3K27me3 patterns compared with NCAFs. Loss of H3K27me3, but not DNA methylation, in CAFs was enriched for genes involved in stem cell niche, cell growth, tissue development and stromal-epithelial interactions, such as WNT5A, GREM1, NOG and IGF2. Among these, we revealed that WNT5A, which had been considered to be derived from cancer cells, was highly expressed in cancer stromal fibroblasts, and was associated with poor prognosis. Inhibition of secreted WNT5A from CAFs suppressed cancer cell growth and migration.
CONCLUSIONS:
H3K27me3 plays a crucial role in defining tumour-promoting capacities of CAFs, and multiple stem cell niche factors were secreted from CAFs due to loss of H3K27me3. The validity of the epigenetic approach to uncover therapeutic targets for cancer-starving therapy was demonstrated.
