Vet Immunol Immunopathol.
McGill JL, Sacco RE.
PMID: 26923879 | DOI: 10.1016/j.vetimm.2016.02.012
γδ T cells are a subset of nonconventional T cells that play a critical role in bridging the innate and adaptive arms of the immune system. γδ T cells are particularly abundant in ruminant species and may constitute up to 60% of the circulating lymphocyte pool in young cattle. The frequency of circulating γδ T cells is highest in neonatal calves and declines as the animal ages, suggesting these cells may be particularly important in the immune system of the very young. Bovine respiratory syncytial virus (BRSV) is a significant cause of respiratory infection in calves, and is most severe in animals under one year of age. BRSV is also a significant factor in the development of bovine respiratory disease complex (BRDC), the leading cause of morbidity and mortality in feedlot cattle. Human respiratory syncytial virus (RSV) is closely related to BRSV and a leading cause of lower respiratory tract infection in infants and children worldwide. BRSV infection in calves shares striking similarities with RSV infection in human infants. To date, there have been few studies defining the role of γδ T cells in the immune response to BRSV or RSV infection in animals or humans, respectively. However, emerging evidence suggests that γδ T cells may play a critical role in the early recognition of infection and in shaping the development of the adaptive immune response through inflammatory chemokine and cytokine production. Further, while it is clear that γδ T cells accumulate in the lungs during BRSV and RSV infection, their role in protection vs. immunopathology remains unclear. This review will summarize what is currently known about the role of γδ T cells in the immune response to BRSV and BRDC in cattle, and where appropriate, draw parallels to the role of γδ T cells in the human response to RSV infection.
Good, PI;Li, L;Hurst, HA;Serrano-Herrera, IM;Xu, K;Rao, M;Bateman, DA;Al-Awqati, Q;D'Agati, VD;Costantini, F;Lin, F;
PMID: 36626229 | DOI: 10.1172/jci.insight.161316
Preterm birth results in low nephron endowment and increased risk of acute kidney injury (AKI) and chronic kidney disease (CKD). To understand the pathogenesis of AKI and CKD in preterm humans, we generated novel mouse models with a 30-70% reduction in nephron number by inhibiting or deleting Ret tyrosine kinase in the developing ureteric bud. These mice developed glomerular and tubular hypertrophy followed by the transition to CKD, recapitulating the renal pathological changes seen in humans born preterm. We injected neonatal mice with gentamicin, a ubiquitous nephrotoxic exposure in preterm infants, and detected more severe proximal tubular injury in mice with low nephron number compared to controls with normal nephron number. Mice with low nephron number have reduced proliferative repair with more rapid development of CKD. Furthermore, mice had more profound inflammation with highly elevated levels of MCP-1 and CXCL10, produced in part by damaged proximal tubules. Our study directly links low nephron endowment with postnatal renal hypertrophy, which in this model is maladaptive and results in CKD. Underdeveloped kidneys are more susceptible to gentamicin-induced AKI, suggesting that AKI in the setting of low nephron number is more severe and further increases the risk of CKD in this vulnerable population.
Lee, D;Helal, Z;Kim, J;Hunt, A;Barbieri, A;Tocco, N;Frasca, S;Kerr, K;Hyeon, J;Chung, D;Risatti, G;
| DOI: 10.3390/v13112141
We report the first detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in a 3-month-old dog in Connecticut that died suddenly and was submitted to the state veterinary diagnostic laboratory for postmortem examination. Viral RNA was detected in multiple organs of the dog by reverse transcription real time-PCR (RT-qPCR). Negative and positive sense strands of viral RNA were visualized by in situ hybridization using RNAscope technology. Complete genome sequencing and phylogenetic analysis of the hCoV-19/USA/CT-CVMDL-Dog-1/2021 (CT_Dog/2021) virus were conducted to identify the origin and lineage of the virus. The CT_Dog/2021 virus belonged to the GH/B1.2. genetic lineage and was genetically similar to SARS-CoV-2 identified in humans in the U.S. during the winter of 2020-2021. However, it was not related to other SARS-CoV-2 variants identified from companion animals in the U.S. It contained both the D614G in spike and P323L in nsp12 substitutions, which have become the dominant mutations in the United States. The continued sporadic detections of SARS-CoV-2 in companion animals warrant public health concerns about the zoonotic potential of SARS-CoV-2 and enhance our collective understanding of the epidemiology of the virus.
Arteriosclerosis, thrombosis, and vascular biology
Owsiany, KM;Deaton, RA;Soohoo, KG;Tram Nguyen, A;Owens, GK;
PMID: 35735018 | DOI: 10.1161/ATVBAHA.122.317882
Smooth muscle cells (SMCs) in atherosclerotic plaque take on multiple nonclassical phenotypes that may affect plaque stability and, therefore, the likelihood of myocardial infarction or stroke. However, the mechanisms by which these cells affect stability are only beginning to be explored.In this study, we investigated the contribution of inflammatory MCP1 (monocyte chemoattractant protein 1) produced by both classical Myh11 (myosin heavy chain 11)+ SMCs and SMCs that have transitioned through an Lgals3 (galectin 3)+ state in atherosclerosis using smooth muscle lineage tracing mice that label all Myh11+ cells and a dual lineage tracing system that targets Lgals3-transitioned SMC only.We show that loss of MCP1 in all Myh11+ smooth muscle results in a paradoxical increase in plaque size and macrophage content, driven by a baseline systemic monocytosis early in atherosclerosis pathogenesis. In contrast, knockout of MCP1 in Lgals3-transitioned SMCs using a complex dual lineage tracing system resulted in lesions with an increased Acta2 (actin alpha 2, smooth muscle)+ fibrous cap and decreased investment of Lgals3-transitioned SMCs, consistent with increased plaque stability. Finally, using flow cytometry and single-cell RNA sequencing, we show that MCP1 produced by Lgals3-transitioned SMCs influences multiple populations of inflammatory cells in late-stage plaques.MCP1 produced by classical SMCs influences monocyte levels beginning early in disease and was atheroprotective, while MCP1 produced by the Lgals3-transitioned subset of SMCs exacerbated plaque pathogenesis in late-stage disease. Results are the first to determine the function of Lgals3-transitioned inflammatory SMCs in atherosclerosis and highlight the need for caution when considering therapeutic interventions involving MCP1.
Journal of clinical pathology
Humphries, MP;Bingham, V;Abdullah Sidi, F;Craig, S;Lara, B;El-Daly, H;O'Doherty, N;Maxwell, P;Lewis, C;McQuaid, S;Lyness, J;James, J;Snead, DRJ;Salto-Tellez, M;
PMID: 36717223 | DOI: 10.1136/jcp-2022-208525
Interrogation of immune response in autopsy material from patients with SARS-CoV-2 is potentially significant. We aim to describe a validated protocol for the exploration of the molecular physiopathology of SARS-CoV-2 pulmonary disease using multiplex immunofluorescence (mIF).The application of validated assays for the detection of SARS-CoV-2 in tissues, originally developed in our laboratory in the context of oncology, was used to map the topography and complexity of the adaptive immune response at protein and mRNA levels.SARS-CoV-2 is detectable in situ by protein or mRNA, with a sensitivity that could be in part related to disease stage. In formalin-fixed, paraffin-embedded pneumonia material, multiplex immunofluorescent panels are robust, reliable and quantifiable and can detect topographic variations in inflammation related to pathological processes.Clinical autopsies have relevance in understanding diseases of unknown/complex pathophysiology. In particular, autopsy materials are suitable for the detection of SARS-CoV-2 and for the topographic description of the complex tissue-based immune response using mIF.
Choudhary, S;Kanevsky, I;Yildiz, S;Sellers, RS;Swanson, KA;Franks, T;Rathnasinghe, R;Munoz-Moreno, R;Jangra, S;Gonzalez, O;Meade, P;Coskran, T;Qian, J;Lanz, TA;Johnson, JG;Tierney, CA;Smith, JD;Tompkins, K;Illenberger, A;Corts, P;Ciolino, T;Dormitzer, PR;Dick, EJ;Shivanna, V;Hall-Ursone, S;Cole, J;Kaushal, D;Fontenot, JA;Martinez-Romero, C;McMahon, M;Krammer, F;Schotsaert, M;García-Sastre, A;
PMID: 35128980 | DOI: 10.1177/01926233211072767
Coronavirus disease 2019 (COVID-19) in humans has a wide range of presentations, ranging from asymptomatic or mild symptoms to severe illness. Suitable animal models mimicking varying degrees of clinical disease manifestations could expedite development of therapeutics and vaccines for COVID-19. Here we demonstrate that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection resulted in subclinical disease in rhesus macaques with mild pneumonia and clinical disease in Syrian hamsters with severe pneumonia. SARS-CoV-2 infection was confirmed by formalin-fixed, paraffin-embedded (FFPE) polymerase chain reaction (PCR), immunohistochemistry, or in situ hybridization. Replicating virus in the lungs was identified using in situ hybridization or virus plaque forming assays. Viral encephalitis, reported in some COVID-19 patients, was identified in one macaque and was confirmed with immunohistochemistry. There was no evidence of encephalitis in hamsters. Severity and distribution of lung inflammation were substantially more in hamsters compared with macaques and exhibited vascular changes and virus-induced cytopathic changes as seen in COVID-19 patients. Neither the hamster nor macaque models demonstrated evidence for multisystemic inflammatory syndrome (MIS). Data presented here demonstrate that macaques may be appropriate for mechanistic studies of mild asymptomatic COVID-19 pneumonia and COVID-19-associated encephalitis, whereas Syrian hamsters may be more suited to study severe COVID-19 pneumonia.
Diffuse trophoblast damage is the hallmark of SARS-CoV-2-associated fetal demise
Modern pathology : an official journal of the United States and Canadian Academy of Pathology, Inc
Garrido-Pontnou, M;Navarro, A;Camacho, J;Crispi, F;Alguacil-Guillén, M;Moreno-Baró, A;Hernandez-Losa, J;Sesé, M;Ramón Y Cajal, S;Garcia Ruíz, I;Serrano, B;Garcia-Aguilar, P;Suy, A;Ferreres, JC;Nadal, A;
PMID: 34006935 | DOI: 10.1038/s41379-021-00827-5
Placental pathology in SARS-CoV-2-infected pregnancies seems rather unspecific. However, the identification of the placental lesions due to SARS-CoV-2 infection would be a significant advance in order to improve the management of these pregnancies and to identify the mechanisms involved in a possible vertical transmission. The pathological findings in placentas delivered from 198 SARS-CoV-2-positive pregnant women were investigated for the presence of lesions associated with placental SARS-CoV-2 infection. SARS-CoV-2 infection was investigated in placental tissues through immunohistochemistry, and positive cases were further confirmed by in situ hybridization. SARS-CoV-2 infection was also investigated by RT-PCR in 33 cases, including all the immunohistochemically positive cases. Nine cases were SARS-CoV-2-positive by immunohistochemistry, in situ hybridization, and RT-PCR. These placentas showed lesions characterized by villous trophoblast necrosis with intervillous space collapse and variable amounts of mixed intervillous inflammatory infiltrate and perivillous fibrinoid deposition. Such lesions ranged from focal to massively widespread in five cases, resulting in intrauterine fetal death. Two of the stillborn fetuses showed some evidence of SARS-CoV-2 positivity. The remaining 189 placentas did not show similar lesions. The strong association between trophoblastic damage and placenta SARS-CoV-2 infection suggests that this lesion is a specific marker of SARS-CoV-2 infection in placenta. Diffuse trophoblastic damage, massively affecting chorionic villous tissue, can result in fetal death associated with COVID-19 disease.
Glycated ACE2 receptor in diabetes: open door for SARS-COV-2 entry in cardiomyocyte
Cardiovascular diabetology
D'Onofrio, N;Scisciola, L;Sardu, C;Trotta, MC;De Feo, M;Maiello, C;Mascolo, P;De Micco, F;Turriziani, F;Municinò, E;Monetti, P;Lombardi, A;Napolitano, MG;Marino, FZ;Ronchi, A;Grimaldi, V;Hermenean, A;Rizzo, MR;Barbieri, M;Franco, R;Campobasso, CP;Napoli, C;Municinò, M;Paolisso, G;Balestrieri, ML;Marfella, R;
PMID: 33962629 | DOI: 10.1186/s12933-021-01286-7
About 50% of hospitalized coronavirus disease 2019 (COVID-19) patients with diabetes mellitus (DM) developed myocardial damage. The mechanisms of direct SARS-CoV-2 cardiomyocyte infection include viral invasion via ACE2-Spike glycoprotein-binding. In DM patients, the impact of glycation of ACE2 on cardiomyocyte invasion by SARS-CoV-2 can be of high importance. To evaluate the presence of SARS-CoV-2 in cardiomyocytes from heart autopsy of DM cases compared to Non-DM; to investigate the role of DM in SARS-COV-2 entry in cardiomyocytes. We evaluated consecutive autopsy cases, deceased for COVID-19, from Italy between Apr 30, 2020 and Jan 18, 2021. We evaluated SARS-CoV-2 in cardiomyocytes, expression of ACE2 (total and glycosylated form), and transmembrane protease serine protease-2 (TMPRSS2) protein. In order to study the role of diabetes on cardiomyocyte alterations, independently of COVID-19, we investigated ACE2, glycosylated ACE2, and TMPRSS2 proteins in cardiomyocytes from DM and Non-DM explanted-hearts. Finally, to investigate the effects of DM on ACE2 protein modification, an in vitro glycation study of recombinant human ACE2 (hACE2) was performed to evaluate the effects on binding to SARS-CoV-2 Spike protein. The authors included cardiac tissue from 97 autopsies. DM was diagnosed in 37 patients (38%). Fourth-seven out of 97 autopsies (48%) had SARS-CoV-2 RNA in cardiomyocytes. Thirty out of 37 DM autopsy cases (81%) and 17 out of 60 Non-DM autopsy cases (28%) had SARS-CoV-2 RNA in cardiomyocytes. Total ACE2, glycosylated ACE2, and TMPRSS2 protein expressions were higher in cardiomyocytes from autopsied and explanted hearts of DM than Non-DM. In vitro exposure of monomeric hACE2 to 120 mM glucose for 12 days led to non-enzymatic glycation of four lysine residues in the neck domain affecting the protein oligomerization. The upregulation of ACE2 expression (total and glycosylated forms) in DM cardiomyocytes, along with non-enzymatic glycation, could increase the susceptibility to COVID-19 infection in DM patients by favouring the cellular entry of SARS-CoV2.
Bräuninger, H;Stoffers, B;Fitzek, ADE;Meißner, K;Aleshcheva, G;Schweizer, M;Weimann, J;Rotter, B;Warnke, S;Edler, C;Braun, F;Roedl, K;Scherschel, K;Escher, F;Kluge, S;Huber, TB;Ondruschka, B;Schultheiss, HP;Kirchhof, P;Blankenberg, S;Püschel, K;Westermann, D;Lindner, D;
PMID: 34647998 | DOI: 10.1093/cvr/cvab322
Cardiac involvement in COVID-19 is associated with adverse outcome. However, it is unclear whether cell specific consequences are associated with cardiac SARS-CoV-2 infection. Therefore, we investigated heart tissue utilizing in situ hybridization, immunohistochemistry and RNA-sequencing in consecutive autopsy cases to quantify virus load and characterize cardiac involvement in COVID-19.In this study, 95 SARS-CoV-2-positive autopsy cases were included. A relevant SARS-CoV-2 virus load in the cardiac tissue was detected in 41/95 deceased (43%). MACE-RNA-sequencing was performed to identify molecular pathomechanisms caused by the infection of the heart. A signature matrix was generated based on the single-cell dataset "Heart Cell Atlas" and used for digital cytometry on the MACE-RNA-sequencing data. Thus, immune cell fractions were estimated and revealed no difference in immune cell numbers in cases with and without cardiac infection. This result was confirmed by quantitative immunohistological diagnosis.MACE-RNA-sequencing revealed 19 differentially expressed genes (DEGs) with a q-value <0.05 (e.g. up: IFI44L, IFT3, TRIM25; down: NPPB, MB, MYPN). The upregulated DEGs were linked to interferon pathways and originate predominantly from endothelial cells. In contrast, the downregulated DEGs originate predominately from cardiomyocytes. Immunofluorescent staining showed viral protein in cells positive for the endothelial marker ICAM1 but rarely in cardiomyocytes. The GO term analysis revealed that downregulated GO terms were linked to cardiomyocyte structure, whereas upregulated GO terms were linked to anti-virus immune response.This study reveals, that cardiac infection induced transcriptomic alterations mainly linked to immune response and destruction of cardiomyocytes. While endothelial cells are primarily targeted by the virus, we suggest cardiomyocyte-destruction by paracrine effects. Increased pro-inflammatory gene expression was detected in SARS-CoV-2-infected cardiac tissue but no increased SARS-CoV-2 associated immune cell infiltration was observed.Cardiac injury can be documented in COVID-19, regardless the direct cardiac virus infection and is known to be associated with outcome. However, the direct virus infection of the myocardium leads to transcriptomic alterations and might therefore additionally contribute to pathophysiological processes in COVID-19. Therefore, consequences of cardiac virus infection need to be investigated in future studies, since they might also contribute to long-term effects in case of survival.
Tissue factor upregulation is associated with SARS-CoV-2 in the lungs of COVID-19 patients
Journal of thrombosis and haemostasis : JTH
Subrahmanian, S;Borczuk, A;Salvatore, S;Fung, KM;Merrill, JT;Laurence, J;Ahamed, J;
PMID: 34236752 | DOI: 10.1111/jth.15451
A substantial proportion of patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) develop severe/critical coronavirus disease 2019 (COVID-19) characterized by acute respiratory distress syndrome (ARDS) with thrombosis.We tested the hypothesis that SARS-CoV-2--induced upregulation of tissue factor (TF) expression may be responsible for thrombus formation in COVID-19.We compared autopsy lung tissues from 11 patients with COVID-19--associated ARDS with samples from 6 patients with ARDS from other causes (non-COVID-19 ARDS) and 11 normal control lungs.Dual RNA in situ hybridization for SARS-CoV-2 and TF identified sporadic clustered SARS-CoV-2 with prominent co-localization of SARS-CoV-2 and TF RNA. TF expression was 2-fold higher in COVID-19 than in non-COVID-19 ARDS lungs (P = .017) and correlated with the intensity of SARS-CoV-2 staining (R2 = .36, P = .04). By immunofluorescence, TF protein expression was 2.1-fold higher in COVID-19 versus non-COVID-19 ARDS lungs (P = .0048) and 11-fold (P < .001) higher than control lungs. Fibrin thrombi and thrombi positive for platelet factor 4 (PF4) were found in close proximity to regions expressing TF in COVID-19 ARDS lung, and correlated with TF expression (fibrin, R2 = .52, P < .001; PF4, R2 = .59, P < .001).These data suggest that upregulation of TF expression is associated with thrombus formation in COVID-19 lungs and could be a key therapeutic target. Correlation of TF expression with SARS-CoV-2 in lungs of COVID-19 patients also raises the possibility of direct TF induction by the virus.
SARS-CoV-2 crosses the blood-brain barrier accompanied with basement membrane disruption without tight junctions alteration
Signal transduction and targeted therapy
Zhang, L;Zhou, L;Bao, L;Liu, J;Zhu, H;Lv, Q;Liu, R;Chen, W;Tong, W;Wei, Q;Xu, Y;Deng, W;Gao, H;Xue, J;Song, Z;Yu, P;Han, Y;Zhang, Y;Sun, X;Yu, X;Qin, C;
PMID: 34489403 | DOI: 10.1038/s41392-021-00719-9
SARS-CoV-2 has been reported to show a capacity for invading the brains of humans and model animals. However, it remains unclear whether and how SARS-CoV-2 crosses the blood-brain barrier (BBB). Herein, SARS-CoV-2 RNA was occasionally detected in the vascular wall and perivascular space, as well as in brain microvascular endothelial cells (BMECs) in the infected K18-hACE2 transgenic mice. Moreover, the permeability of the infected vessel was increased. Furthermore, disintegrity of BBB was discovered in the infected hamsters by administration of Evans blue. Interestingly, the expression of claudin5, ZO-1, occludin and the ultrastructure of tight junctions (TJs) showed unchanged, whereas, the basement membrane was disrupted in the infected animals. Using an in vitro BBB model that comprises primary BMECs with astrocytes, SARS-CoV-2 was found to infect and cross through the BMECs. Consistent with in vivo experiments, the expression of MMP9 was increased and collagen IV was decreased while the markers for TJs were not altered in the SARS-CoV-2-infected BMECs. Besides, inflammatory responses including vasculitis, glial activation, and upregulated inflammatory factors occurred after SARS-CoV-2 infection. Overall, our results provide evidence supporting that SARS-CoV-2 can cross the BBB in a transcellular pathway accompanied with basement membrane disrupted without obvious alteration of TJs.
Fluids and barriers of the CNS
Errede, M;Annese, T;Petrosino, V;Longo, G;Girolamo, F;de Trizio, I;d'Amati, A;Uccelli, A;Kerlero de Rosbo, N;Virgintino, D;
PMID: 36042496 | DOI: 10.1186/s12987-022-00365-5
In myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), several areas of demyelination are detectable in mouse cerebral cortex, where neuroinflammation events are associated with scarce inflammatory infiltrates and blood-brain barrier (BBB) impairment. In this condition, the administration of mesenchymal stem cells (MSCs) controls neuroinflammation, attenuating astrogliosis and promoting the acquisition of stem cell traits by astrocytes. To contribute to the understanding of the mechanisms involved in the pathogenesis of EAE in gray matter and in the reverting effects of MSC treatment, the neocortex of EAE-affected mice was investigated by analyzing the cellular source(s) of chemokine CCL2, a molecule involved in immune cell recruitment and BBB-microvessel leakage.The study was carried out by immunohistochemistry (IHC) and dual RNAscope IHC/in situ hybridization methods, using astrocyte, NG2-glia, macrophage/microglia, and microglia elective markers combined with CCL2.The results showed that in EAE-affected mice, hypertrophic microglia are the primary source of CCL2, surround the cortex neurons and the damaged BBB microvessels. In EAE-affected mice treated with MSCs, microgliosis appeared diminished very soon (6 h) after treatment, an observation that was long-lasting (tested after 10 days). This was associated with a reduced CCL2 expression and with apparently preserved/restored BBB features. In conclusion, the hallmark of EAE in the mouse neocortex is a condition of microgliosis characterized by high levels of CCL2 expression.This finding supports relevant pathogenetic and clinical aspects of the human disease, while the demonstrated early control of neuroinflammation and BBB permeability exerted by treatment with MSCs may have important therapeutic implications.
