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Background/Objectives: The broad spectrum of clinical manifestations from SARS-COV-2 infection and observed risk factors for severe disease highlight the importance of understanding molecular mechanisms underlying SARS-CoV-2 associated disease pathogenesis. Research studies have identified a large number of host proteins that play roles in viral entry, innate immune response, or immune signalling during infection. The ability to interrogate subsets of these genes simultaneously within SARSCOV-2 infected samples is critical to understanding how their expression contribute to phenotypic variability of the disease caused by the pathogen. Method(s): 30 Nasopharyngeal swab were obtained and included SARS-CoV-2 infected and control samples. RNA was extracted, reverse transcribed and loaded onto flexible TaqMan array panels designed specifically for targeting the most cited genes related to SARS-COV-2 entry and restriction factors as well as cytokines, chemokines, and growth factors involved in the pathogenesis. Result(s): Our data indicated that not only were the levels of several of these host factors differentially modulated between the two study groups, but also that SARS-CoV-2 infected subjects presented with greater frequency of several important inflammatory cytokines and chemokines such as CCL2, CCL3, IFNG, entry receptors such as ACE2, TMRPS11A, and host restriction factors including LY6E and ZBP1. Conclusion(s): TaqMan array plates provide a fast, midthroughput solution to determine the levels of several virus and host-associated factors in various cell types and add to our understanding of how the pathogenesis may vary depending on gender, age, infection site etc.
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This review highlights ten important advances in the neuromuscular disease field that were reported in 2021. As with prior updates in this article series, the overarching topics include (i) advances in understanding of fundamental neuromuscular biology; (ii) new / emerging diseases; (iii) advances in understanding of disease etiology and pathogenesis; (iii) diagnostic advances; and (iv) therapeutic advances. Within this general framework, the individual disease entities that are discussed in more detail include neuromuscular complications of COVID-19 (another look at the topic first covered in the 2021 review), autosomal recessive myopathy caused by MLIP mutations, autosomal recessive neuromuscular disease caused by VWA1 mutations, Leber's hereditary optic neuropathy, myopathies with autophagic defects, tRNA synthetase-associated Charcot-Marie-Tooth disease, systemic sclerosis-associated myopathy, humoral immune endoneurial microvasculopathy, and late-onset Pompe disease. In addition, the review highlights a few other advances (including new insights into mechanisms of muscle and nerve regeneration and the use of gene expression profiling to better characterize different subtypes of immune-mediated myopathies) that will be of significant interest for clinicians and researchers who specialize in neuromuscular disease.
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The COVID-19 pandemic is a global outbreak of coronavirus, an infectious disease caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). One in five adults who have had COVID-19 in the past was still experiencing any one of the symptoms of long COVID like headache, brain fog, fatigue, and shortness of breath. Up to 30% of individuals with mild to severe infection show diverse neurological symptoms, including dementias. Hence, it is very much important to characterize the neurotropism and neurovirulence of the SARS-CoV-2 virus. This helps us understand the mechanisms involved in initiating inflammation in the brain, further leading to the development of earlyonset Alzheimer's disease and related dementias (ADRDs). In our brain gene expression analysis, we found that severe COVID-19 patients showed increased expression of innate immune response genes and genes that are implicated in AD pathogenesis. To study the infection-induced ADRDs, we used a mouse-adapted strain of the SARS-CoV-2 (MA10) virus to infect mice of different age groups (3, 6, and 20 Months). In this study, we found that aged mice showed evidence of viral neurotropism, prolonged viral infection, increased expression of tau aggregator FKBP51, interferoninducible gene Ifi204, and complement genes like C4 and C5AR1. Brain histopathology also showed the AD signature including tau-phosphorylation, tau-oligomerization, and alpha-synuclein expression in aged MA10-infected mice. The results from gene expression profiling of SARS-CoV-2 infected and AD brains and studies with MA10 aged mice show that COVID-19 infection increases the risk of AD in the aged population. Furthermore, this study helps us to understand the crucial molecular markers that are regulated during COVID infection that could act as major players in developing ADRDs. Future studies will be involved in understanding the molecular mechanisms of ADRD in response to COVID infection and developing novel therapies targeting AD.
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The novel SARS-CoV-2 virus known to cause the COVID-19 outbreak has resulted in a global healthcare crisis that has persisted the past 3 years. Thus, understanding the mechanisms underlying this disease are vital at this time. While there are issues of research infrastructure to handle the virus and because of the refractoriness of rodents to this disease, the availability of these tools is still limited. The cytokine storm and fatality presented in patients with severe COVID-19 can be mimicked with Staphylococcal enterotoxin B (SEB)-induced Acute Respiratory Distress Syndrome (ARDS). Within ~7 days, the survival rate drops to 0% for C3H/HeJ mice exposed to a dual dose of SEB. In this study, we administered cannabidiol (CBD) intraperitoneally for 3 days pre- and post-SEB dosing and found that the clinical outcomes improved significantly. Initial evaluation of scRNASeq data from lungs comparing naive to SEB-induced ARDS mice illustrated an increase in infiltrating immune cells, and a loss in pulmonary epithelial cells in the latter group. When evaluating the effect of CBD treatment on SEB-induced ARDS, we were able to demonstrate that CBD reduced the macrophage population. To characterize the mechanism by which CBD treatment ameliorated the inflammatory response, we found that CBD treated mice had significant reduction in infiltrating immune cells and alveolar thickening. This same histology and infiltration is presented in ARDS. MicroRNA expression analysis showed a significant increase in the expression mmu-miR-298-5p and mmu-miR- 566 with CBD treatment. Ingenuity Pathway Analysis (IPA) indicated that the dysregulated miRNAs were also implicated in pathways associated with macrophage activation, respiratory disease and inflammation, interferon stimulated genes, as well as genes which have been upregulated in the disease state of this model. These targets include but are not limited to Cebpb, Efhd2, Stat3, Socs3, Cxcl5, Gbp2, and Birc3. This finding offers insights for the development of preventive and therapeutic strategies in the treatment of ARDS, including that induced in COVID-19. Supported by NIH grants P01AT003961, P20GM103641, R01ES003961, R01AI129788, R01AI123947, R01AI160896 to MN and PSN and K99GM147910 to KW.Copyright © 2023 The American Society for Biochemistry and Molecular Biology, Inc.
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Respiratory viral infections are important cause of morbidity and mortality in early life. The relative influence of host and viral factors possibly contribute to the disease pathogenesis. Predisposing conditions like prematurity, Low birth weight and congenital heart diseases etc. have been incriminated in the disease progression. The development of cough, wheezing, and tachypnea, usually peaking on days 4 to 5, go parallel with host cytokine responses and viral load. Various host cytokines, chemokines and molecules involved in the immune response against RSV infection might be responsible for the outcome of the disease process. Nasopharyngeal aspirates (NPAs) from children (n = 349) between 2013-2017 were subjected for IL-17A, IFN-gamma, TNF-alpha, IL-10, IL-6 levels by CBA and MMP-9 and TIMP-1 levels by ELISA. The viral load in RSV positive samples and cytokine levels were correlated with the WHO criteria for acute lower respiratory tract illness (ALRTI). RSV viral load, Pro-inflammatory cytokine (TNF-alpha) levels in severe ALRTI patients were significantly higher than the ALRTI patients [p<0.001]. Whereas Th17 cytokine (IL-17) was found to be significantly higher (p<0.05) in ALRTI patients than severe patients. MMP-9 is secreted in higher levels in severe ALRTI patients (n = 77) in comparison to Acute LRTI patients (n = 35) with an increase of thirty seven fold (p<0.001). Thus, the study highlights the role of TNF -alpha, IL-17 and Th2 cytokine biasness in the pathogenesis of RSV disease with the possible contribution of higher MMP-9/TIMP-1 ratio as a bad prognostic marker towards disease severity. To study the gene expression of autophagy and mTOR signalling pathways in RSV infected children with ALRTI. Nasopharyngeal aspirate (NPA) samples (n = 145) from children suffering from ALRTI were subjected for detection of RSV (Oct 2019 to March 2020). Semi-quantitative gene expression analysis for 5 representative genes each of mTOR signalling and autophagy pathway were performed in respiratory tract epithelial cells using 25 RSV positive cases and 10 healthy controls subjects. Autophagy gene expression analysis revealed significant upregulation in NPC1 and ATG3 autophagy genes. mTOR, AKT1 and TSC1 genes of mTOR pathway were significantly down-regulated in RSV positive patients except RICTOR gene which was significantly upregulated. Thus, survival of RSV within autophagosome might have been facilitated by upregulation of autophagy and downregulation of mTOR signalling genes. To assess the impact of SARS-CoV2 pandemic on RSV, samples were collected from children with ALRTIs admitted to emergency, PICU and indoor admissions during pre-pandemic period (October 2019 to February 2020;n = 166) and during COVID-19 Pandemic (July 2021 to July 2022;n = 189, SARS-CoV2 negative). These NP swabs were analyzed for pdm InfA H1N1, InfA H3N2, Inf B, RSV, hMPV, hBoV, hRV, PIV-2 and PIV-3 by PCR. Higher proportion of children with ALRTIs have had virus/es isolated during pre-pandemic period than during pandemic period (p<0.001). During pre-pandemic period, significantly higher proportion of children had RSV positivity (p<0.001);and significantly lower positivity for hRV (p<0.05), hMPV (p<0.05), and hBoV (p <= 0.005). The occurrence of COVID-19 pandemic has significantly impacted the frequency and pattern of detection of RSV among hospitalized children with LRTIs. RSV Fusion protein plays a critical role in the entry of the virus into the host cell by initiating the fusion of host and viral membranes. It happens to be a target of neutralizing antibodies paving the way as a vaccine candidate. Hence effort was made to introduce point mutation in hRSV fusion protein which can confer stability in its prefusion form. In-silico a stable structure of RSV fusion protein was generated making it a potential vaccine candidate. The timely diagnosis of RSV infection in this population is important for initiating therapy and instituting appropriate infection prevention measures. Serological testing is not widely used for the diagnosis of RSV. C ll Cultures including shell vial culture were used for RSV diagnosis. However, culture approaches lack sensitivity, often quite significantly, compared to nucleic acid amplification assays for the diagnosis of RSV infections. Molecular multiplex assays now offer increased sensitivity for a more accurate diagnosis. However issues with the use of these types of commercial panel assays include the requirement for substantial training, quality systems, and infrastructure to maintain and run these assays and many a times identification of viruses where the true pathogenic potential of those multiple viruses are debatable. Studies are available with laboratory- developed nucleic acid amplification test systems for the detection of RSVA and RSVB in clinical specimens either by PCRbased technologies or RT-LAMP. Gene targets of laboratory-developed molecular assays point towards M gene and the N gene in RSVA and -B with the benefits of flexibility to modify assays when targets are under evolutionary pressure to change, as well as a perceived initial low cost to carry out testing.
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Introduction: Malignancies are a major complication of heart transplant (HT). Noninvasive surveillance after HT using gene expression (GEP) profiling and donor derived cell free DNA (dd-cfDNA) are noninferior to biopsy and are widely utilized. The interpretation of % dd-cfDNA, is not well understood in malignancies with a conceptual increase in the recipient fraction. The effect of chemotherapy on GEP in the setting of post-HT surveillance has not been described to the best of our knowledge. Hypothesis: Induction of chemotherapy will cause global transcriptional reduction in GEP. Method(s): GEP was performed with AlloMap (AM, CareDx), which evaluates expression levels of 11 mononuclear cell genes, involved in lymphocyte activation, T-cell priming, cell migration, hematopoietic proliferation, steroid sensitivity, and platelet activation. Scores range from 0-40, higher scores have a stronger correlation with rejection. At our center a total of 995 draws were analyzed from 2019-2022. In parallel dd-cfDNA, which informs about graft injury was analyzed using AlloSure (AS, CareDx). Case Events: A 71-year-old male HT recipient for nonischemic cardiomyopathy and no rejection history was diagnosed with metastatic gastric adenocarcinoma at 16 months post-HT. Following diagnosis, mycophenolic acid was stopped, prednisone 5 mg was started, and tacrolimus trough goal was gradually lowered to 4-6 given infectious complications. Palliative chemotherapy with folinic acid, fluorouracil (5-FU), oxaliplatin (FOLFOX) was initiated at 18 months post-HT with planned dose reduction of oxaliplatin and holding of 5-FU bolus to reduce risk of myelosuppression given comorbidities. Oxaliplatin was stopped at 18 months post HT. Due to COVID he last received 5-FU at 33 months post-HT. Graft function remained stable and DSA negative. At 36 months post-HT, he developed a bowel obstruction without surgical options for interventions and expired shortly thereafter. Result(s): With initiation of prednisone and following chemotherapy there was a drastic decrease in AM scores (Fig. A). Steroid therapy led to an 18% decline in AM scores, the greatest decrease occurred with chemotherapy, with 67% decline from the mean when compared to all center patients (Fig B). Dd-cfDNA levels remained stable during the course aside from one early elevation. Conclusion(s): To the best of our knowledge this is the first published case on the effect of chemotherapy on GEP profiling in the setting of post-HT surveillance. This case advises caution when interpreting GEP in the setting of chemotherapy showing great reduction in GEP scores. While dd-cfDNA levels remained relatively stable after malignancy diagnosis and treatment initiation further studies will need to inform on the use of both GEP and dd-cfDNA in these patients.Copyright © 2022
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Patients with severe COVID-19-associated pneumonia are at risk to develop pulmonary fibrosis. To study the underlying mechanisms, we aim to develop advanced cell culture models that reliably reflect COVID-19-related profibrotic microenvironment. To identify key cellular players, we performed pilot immunohistochemistry analysis on lung tissue from COVID-19 patients with fibrosis collected during autopsy. Results revealed diffuse alveolar damage with macrophage infiltration, and myofibroblast accumulation with enriched collagen deposition surrounding the damaged alveoli. To mimic SARS-CoV-2 infection in alveoli, we infected human primary type II alveolar epithelial cells (AEC2) and found enhanced signaling of profibrotic cytokine transforming growth factor beta (TGFbeta) in some donors. To recreate the early fibrotic niche, an alveolar-macrophage-fibroblast (AMF) tri-culture model was established. After infecting AEC2 with SARS-CoV-2 in this AMF model, gene expression analysis provided evidence for fibroblast-to-myofibroblast transition. Furthermore, we found that overexpression of SARS-CoV-2 papain-like protease (PLpro) can promote TGFbeta signaling in HEK293T and A549 cells. After infecting AEC2 with SARS-CoV-2 PLpro lentivirus in the AMF model, we found signs of epithelial-to-mesenchymal transition and fibroblast-to myofibroblast transition. In future studies, we will use a detailed analysis of COVID-19-associated lung fibrosis with other types of lung fibrosis, to further refine COVID-19-related fibrosis models, including lung-on-chip models.
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Objective To explore the genomic changes of human olfactory neuroepithelial cells after the novel coronavirus (SARS-COV-2) infecting the human body, and establish a protein-protein interaction (PPI) network of differentially expressed genes (DEGs), in order to understand the impact of SARS-COV-2 infection on human olfactory neuroepithelial cells, and provide reference for the prevention and treatment of new coronavirus pneumonia. Methods The public dataset GSE151973 was analyzed by NetworkAnalyst. DEGs were selected by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis. PPI network, DEGs-microRNA regulatory network, transcription factor-DEGs regulatory network, environmental chemicals-DEGs regulatory network, and drug-DEGs regulatory network were created and visualized by using Cytoscape 3.7.2. Results After SAR-COV-2 invading human olfactory neuroepithelial cells, part of the gene expression profile was significantly up-regulated or down-regulated. A total of 568 DEGs were found, including 550 up-regulated genes (96.8%) and 18 down-regulated genes (3.2%). DEGs were mainly involved in biological processes such as endothelial development and angiogenesis of the olfactory epithelium, and the expression of molecular functions such as the binding of the N-terminal myristylation domain. PPI network suggested that RTP1 and RTP2 were core proteins. MAZ was the most influential transcription factor. Hsa-mir-26b-5p had the most obvious interaction with DEGs regulation. Environmental chemical valproic acid and drug ethanol had the most influence on the regulation of DEG. Conclusion The gene expression of olfactory neuroepithelial cells is significantly up-regulated or down-regulated after infection with SAR-COV-2. SARS-CoV-2 may inhibit the proliferation and differentiation of muscle satellite cells by inhibiting the function of PAX7. RTP1 and RTP2 may resist SARS-CoV-2 by promoting the ability of olfactory receptors to coat the membrane and enhancing the ability of olfactory receptors to respond to odorant ligands. MAZ may regulate DEGs by affecting cell growth and proliferation. Micro RNA, environmental chemicals and drugs also play an important role in the anti-SAR-COV-2 infection process of human olfactory neuroepithelial cells.Copyright © 2022 Editorial Department of Journal of Shanghai Second Medical University. All rights reserved.
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Noninvasive heart transplant rejection surveillance using gene expression profiling (GEP) to monitor immune activation is widely used among heart transplant programs. With the new development of donor-derived cell-free DNA (dd-cfDNA) assays, more programs are transitioning to a predominantly noninvasive rejection surveillance protocol with a reduced frequency of endomyocardial biopsies. As a result, many practical questions arise that potentially delay implementation of these valuable new tools. The purpose of this review is to provide practical guidance for clinicians transitioning toward a less invasive acute rejection monitoring protocol after heart transplantation, and to answer 10 common questions about the GEP and dd-cfDNA assays. Evidence supporting GEP and dd-cfDNA testing is reviewed, as well as guidance on test interpretation and future directions.
Subject(s)
Cell-Free Nucleic Acids , Heart Failure , Heart Transplantation , Humans , Graft Rejection/diagnosis , Postoperative Complications , Biopsy , Cell-Free Nucleic Acids/genetics , Tissue DonorsABSTRACT
Objective To explore the genomic changes of human olfactory neuroepithelial cells after the novel coronavirus (SARS-COV-2) infecting the human body, and establish a protein-protein interaction (PPI) network of differentially expressed genes (DEGs), in order to understand the impact of SARS-COV-2 infection on human olfactory neuroepithelial cells, and provide reference for the prevention and treatment of new coronavirus pneumonia. Methods The public dataset GSE151973 was analyzed by NetworkAnalyst. DEGs were selected by conducting Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) signal pathway analysis. PPI network, DEGs-microRNA regulatory network, transcription factor-DEGs regulatory network, environmental chemicals-DEGs regulatory network, and drug-DEGs regulatory network were created and visualized by using Cytoscape 3.7.2. Results After SAR-COV-2 invading human olfactory neuroepithelial cells, part of the gene expression profile was significantly up-regulated or down-regulated. A total of 568 DEGs were found, including 550 up-regulated genes (96.8%) and 18 down-regulated genes (3.2%). DEGs were mainly involved in biological processes such as endothelial development and angiogenesis of the olfactory epithelium, and the expression of molecular functions such as the binding of the N-terminal myristylation domain. PPI network suggested that RTP1 and RTP2 were core proteins. MAZ was the most influential transcription factor. Hsa-mir-26b-5p had the most obvious interaction with DEGs regulation. Environmental chemical valproic acid and drug ethanol had the most influence on the regulation of DEG. Conclusion The gene expression of olfactory neuroepithelial cells is significantly up-regulated or down-regulated after infection with SAR-COV-2. SARS-CoV-2 may inhibit the proliferation and differentiation of muscle satellite cells by inhibiting the function of PAX7. RTP1 and RTP2 may resist SARS-CoV-2 by promoting the ability of olfactory receptors to coat the membrane and enhancing the ability of olfactory receptors to respond to odorant ligands. MAZ may regulate DEGs by affecting cell growth and proliferation. Micro RNA, environmental chemicals and drugs also play an important role in the anti-SAR-COV-2 infection process of human olfactory neuroepithelial cells.
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Background: Primary Sjogren's syndrome (pSS) is a chronic, systemic, inflammatory autoimmune disease in which existing studies have found the presence of pSS-specific antibodies anti-SSA/ Ro in acute infection with COVID-19.1 The emergence of this phenomenon makes us aware that in the context of the long-term epidemic of COVID-19, it is necessary to further study the molecular mechanisms of the high susceptibility of pSS patients to COVID-19. Method(s): The gene expression profiles of 8 COVID-19 datasets and 5 pSS datasets were downloaded from the Gene Expression Omnibus (GEO) database. The differentially expressed genes (DEGs) between COVID-19 and PSS were identified using the limma software package and Weighted Gene Co-expression Network Analysis (WGCNA). A Venn diagram was used to discover common upregulated DEGs. To explore the possible pathogenesis of both diseases, common signaling pathways were explored by enriching DEGs using Gene Ontology (GO) and the Kyoto Gene and Genome Encyclopedia (KEGG) pathway. Protein-protein interactions (PPIs) were established to identify hub genes and key modules. The analysis of key gene expression characteristics by The Connectivity Map was used to predict potentially effective drugs. Finally, the CIBERSORT method was used to comprehensively evaluate the immune infiltrates of patients with COVID-19 and PSS to study the mechanisms that may have a common immune response or immune cell infiltration. Result(s): A total of 82 upregulated DEGs were identified in both COVID-19 and PSS (Figure 1 A-E). Functional enrichment analysis illustrated the important role of enhanced signaling pathways in response to virus defense and interferon-alpha in both diseases (Figure 1F).Three key modules including 25 hub genes were identified (Figure 1G). The correlation analysis of immune cell infiltration showed the expression of B cells memory resting decreased and NK cells resting increased significantly in the two diseases (Figure 1H, I). Finally, estradiol in drug prediction outcomes has been shown to reduce susceptibility to COVID-19 and its severity through its involvement in regulating immune cells, while the most common manifestation of dry eye in pSS patients is strongly associated with low estrogen. Conclusion(s): High defense response to virus and response to interferon-alpha in pSS patients might be a crucial susceptible factor for COVID-19 and predictive drugs such as estradiol, suggested by susceptibility genes common to COVID-19 and pSS, may help in the clinical treatment of both diseases.
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AIMS: Lung tissue from COVID-19 patients shares similar histomorphological features with chronic lung allograft disease, also suggesting activation of autoimmune-related pathways in COVID-19. To more clearly understand the underlying spectrum of pathophysiology in COVID-19 pneumonia, we analysed mRNA expression of autoimmune-related genes in post-mortem lung tissue from COVID-19 patients. METHODS AND RESULTS: Formalin-fixed, paraffin-embedded lung tissue samples of 18 COVID-19 patients and eight influenza patients were used for targeted gene expression profiling using NanoString technology. Multiplex immunofluorescence for tryptase and chymase was applied for validation. Genes related to mast cells were significantly increased in COVID-19. This finding was strengthened by multiplex immunofluorescence also showing a significant increase of tryptase- and chymase-positive cells in COVID-19. Furthermore, receptors for advanced glycation end-products (RAGE) and pro-platelet basic protein (PPBP) were up-regulated in COVID-19 compared to influenza. Genes associated with Type I interferon signalling showed a significant correlation to detected SARS-CoV2 pathway-related genes. The comparison of lung tissue samples from both groups based on the presence of histomorphological features indicative of acute respiratory distress syndrome did not result in finding any specific gene or pathways. CONCLUSION: Two separate means of measuring show a significant increase of mast cells in SARS-CoV-2-infected lung tissue compared to influenza. Additionally, several genes involved in fibrosis and thrombosis, among which are RAGE and PPBP, are up-regulated in COVID-19. As mast cells are able to induce thrombosis and fibrosis, they may play an important role in the pathogenesis of COVID-19.
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Objective: Single-cell RNA-sequencing (scRNA-seq) and spatial transcriptomics have identified novel cell subtypes and microenvironments which compartmentalize diverse functions at the maternal-fetal interface. We aimed to combine these high-resolution technologies with a rigorous classification of transcription alterations associated with diabetes subtypes in pregnancy. We hypothesized characteristic transcriptome profiles in specific cell populations would be linked to these classifications. Study Design: We clinically validated gestational diabetes mellitus type 1 (GDMA1), GDMA2, and type 2 diabetes (T2DM) classes within a cohort of placentae and compared them to healthy controls by bulk RNA-seq (N=53). We then integrated our non-diabetic term placentae spatial transcriptomics data (N=12) with 273,944 publicly available transcriptomes from term placenta scRNA-seq or single-nuclei RNA-seq (snRNA-seq) datasets (accessions phs001886, GSE173193, EGAS00001002449) with control, GDM, or SARS-CoV-2 positive subjects to create a placental transcriptomic catalog. Result(s): In bulk, we identified 104 significantly differentially expressed transcripts (-2< log2fold-change >2,p< 0.05) in our GDMA1 samples, 102 with GDMA2, and 121 with T2DM (Fig. 1a). Comparisons revealed 88 transcripts uniquely marking GDMA1, 68 for GDMA2, and 85 for T2DM, while FGA and CYP1A1 perturbations were shared across diabetes classes (Fig. 1b). We then compared these bulk GDM subtype markers with the 5,211 significantly differentially expressed transcripts associated with 22 cell-type clusters in our term placenta atlas (Fig. 2), and 20 unique GDM bulk markers aligned with extravillous trophoblast, stromal, endometrial epithelia, endothelial, NK, and dendritic single-cell placental markers. Conclusion(s): Together, these results detail the gene expression profiles and the cell types in the maternal-fetal milieu of pregnancies affected by diabetes. Consistent with their distinct clinical outcomes, GDM and T2DM have unique cellular transcriptomes and would thus be targets for new therapeutics. [Formula presented] [Formula presented] Copyright © 2022
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INTRODUCTION: Transcriptome-derived sepsis subphenotypes, termed 'adaptive', 'inflammopathic' and 'coagulopathic', have been reliably identified in sepsis cohorts, however plasma proteomics in these groups have not been well characterized. We hypothesized that inflammatory and vascular injury markers would be elevated in the inflammopathic and coagulopathic groups compared to the adaptive group. METHOD(S): We prospectively enrolled and obtained blood from 130 inpatients with COVID19-related sepsis. Severity was classified by NIH ordinal scale. Gene expression analysis was performed by Nanostring nCounter (Inflammatix). Inflammatory proteins interleukin (IL)-6, IL8, IL10, IL1RA, IL1RL1, and IFNg and vascular markers ANGPT2, sICAM, vWF, ADAMTS13, and protein C were measured with OLINK proximity extension assay. Clinical variables were compared by chi-square and protein levels were compared using ANOVA with Bonferroni adjustment. RESULT(S): The transcriptomic classifier identified 32% (41) inflammopathic, 50% (65) adaptive and 18% (24) coagulopathic subjects. The inflammopathic group had more patients requiring mechanical ventilation (39% vs 9% vs 21%;p < 0.001) and higher 90-day mortality (32% vs 8% vs 13%, p = 0.016). Inflammatory cytokines IL8 and IL10 were significantly higher in inflammopathic compared to adaptive (p=0.038 and p=0.017 respectively), but not compared to coagulopathic (p>0.99 and p=0.24, respectively). Both the inflammopathic and coagulopathic groups expressed higher IL1RL1 and interferon-gamma compared to adaptive (IL1RL1;p< 0.001, p=0.002, IFNg;p=0.007, p=0.001). Plasma IL6 and IL1RA did not differ between groups, nor did many vascular proteins. The inflammopathic group expressed higher sICAM (p=0.049 vs adaptive) and lower ADAMTS13 compared to the adaptive group, and the coagulopathic group did not differ in its vascular protein expression. CONCLUSION(S): Transcriptomic subphenotypes are present in COVID-19 sepsis at similar proportions to non-COVID-19 sepsis. Inflammopathic subjects manifested higher severity of illness at admission, higher expression of inflammatory proteins and higher mortality. Markers of vascular injury did not distinguish the coagulopathic group. Integrating RNA and protein expression may offer new insights to host immune dysregulation during COVID sepsis.
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Methods: Here, we sought to investigate the effects of TLR7 pathway activation on mouse behaviour 24 hours post-activation. Female CD1 mice received an intraperitoneal injection of the synthetic TLR7 agonist, R848, or an equivalent volume of saline and were subjected to the Open Field and Forced Swim Test 24 hours later (n=10/group). Brain and liver tissues were then collected for downstream gene expression analysis. Result(s): Independent T-tests confirmed that systemic R848 challenge induced a strong peripheral and central inflammatory response, as indicated by a 250-fold increase in hepatic SAA-2 mRNA expression (p<0.0001) and a 75-fold increase in CXCL10 mRNA expression in the prefrontal cortex (p<0.01), relative to controls. These changes in inflammatory markers were accompanied by evidence of sickness behaviour - in particular, a decrease in exploratory rearing (p<0.01). Conclusion(s): This demonstrates that R848 can be used to create a model reflective of viral-like illness and provides a useful tool for investigating the behavioural effects of TLR7-mediated inflammation. To further these results, we aim to explore the metabolic consequences of LPS and R848 challenge and relate these to inflammatory and behavioural changes. This will accompany our data on the metabolic signatures of SARS-CoV-2-infected cells and animals, and of long-COVID patients. Copyright © 2022
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The palatine tonsils (hereinafter referred to as "tonsils") serve as a reservoir for viral infections and play roles in the immune system's first line of defense. The aims of this study were to establish tonsil epithelial cell-derived organoids and examine their feasibility as an ex vivo model for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. The tonsil organoids successfully recapitulated the key characteristics of the tonsil epithelium, including cellular composition, histologic properties, and biomarker distribution. Notably, the basal layer cells of the organoids express molecules essential for SARS-CoV-2 entry, such as angiotensin-converting enzyme 2 (ACE2), transmembrane serine protease 2 (TMPRSS2) and furin, being susceptible to the viral infection. Changes in the gene expression profile in tonsil organoids revealed that 395 genes associated with oncostatin M signaling and lipid metabolism were highly upregulated within 72 h after SARS-CoV-2 infection. Notably, remdesivir suppressed the viral RNA copy number in organoid culture supernatants and intracellular viral protein levels in a dosedependent manner. Here, we suggest that tonsil epithelial organoids could provide a preclinical and translational research platform for investigating SARS-CoV-2 infectivity and transmissibility or for evaluating antiviral candidates.
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Background: SARS-CoV- 2 tropism for the ACE2 receptor, along with the multifaceted inflammatory reaction, is likely to drive the generalized hypercoagulable state seen in patients with COVID-19. Aim(s): We aimed identify microRNAs (miRNAs) play role in ACE2-related thrombosis in coronavirus infection and validated the expressions of those miRNAs in 79 hospitalized COVID-19 patients and 32 healthy volunteers and monitored miRNAs patterns during the acute phase of COVID-19, as well as the prognostic potential biomarker. Method(s): Using the original bioinformatic workflow and network medicine approaches we reanalyzed four coronavirus-related expression datasets and performed co-expression analysis focused on thrombosis and ACE2 related genes. Blood RNA;quality RNA was assessed: Fluorometric assay;RT-PCR for miRNAs expression measurement. Result(s): We identified EGFR, HSP90AA1, APP, TP53, PTEN, UBC, FN1, ELAVL1 and CALM1 as regulatory genes which could play a pivotal role in COVID-19 related thrombosis. We also found miR-16- 5p, miR-27a- 3p, Let-7b- 5p and miR-155- 5p as regulators in coagulation and thrombosis process. We observed in separate cohort of COVID-19 patients and healthy controls that (i) expression of miR-16- 5p, miR-27a- 3p and miR-155- 5p increased during observation, compared to the baseline measurement;(ii) a low baseline miR-16- 5p expression presents predictive utility in assessment of the hospital length of stay or death in follow-up as a composite endpoint (AUC:0.810, 95% CI, 0.71-0.91, p < 0.0001);(iii) low baseline expression of miR-16- 5p and T2DM are independent predictors of increased length of stay or death according to a multivariate analysis (OR: 9.417;95% CI, 2.647-33.506;p = 0.0005 and OR: 6.257;95% CI, 1.049-37.316;p = 0.044). Conclusion(s): This study enabled us to better characterize changes in gene expression and signaling pathways related to COVID-19 thrombosis. In this study we identified, characterized and validated miRNAs which could serve as novel, thrombosis-related biomarkers of the COVID-19, can be used for early stratification of patients and prediction of severity of infection development in an individual. (Figure Presented).
ABSTRACT
Purpose: Preliminary studies suggest that kidney transplant recipients (KTRs) show diminished humoral responses to SARS-CoV-2 vaccination. Although reports of allograft rejection after SARS-CoV-2 vaccination have been rare, there is no recommended framework for monitoring for potential vaccine-related allograft injury. Here, we describe an approach for longitudinal assessment of immunogenicity and safety of SARS-COV-2 vaccination in KTRs. Method(s): KTRs eligible for SARS-CoV-2 vaccination were identified through medical records, beginning March 12, 2021. Baseline and weekly blood samples were collected for SARS-CoV-2 spike protein antibody titers, dd-cfDNA and gene expression profiling (GEP) for 12 weeks. Donor specific antibody (DSA) testing was performed at baseline, 2 weeks after completion of vaccine doses and at week 12. Antibody response was defined as a 10-fold increase in total binding IgG titers. Result(s): 49 KTRs were identified for analysis. Patient demographics are shown in Table 1. Ten patients (20.4%) demonstrated a spike antibody response post- vaccination. Of responders, 80% (n=8) had a history of COVID-19. The odds ratio for the association of a history of COVID-19 with vaccine response was 18.3 (95% CI 3.2, 105.0, p=0.0005). Median dd-cfDNA levels did not differ between pre- and postvaccination (0.23% versus 0.21% respectively). There was no significant difference between pre- and post-vaccination GEP scores (9.85 versus 10.4 respectively). No patients developed clinically significant DSA, eGFR decline or allograft rejection following vaccination. Conclusion(s): Quantitative antibody responses were strongly associated with a diagnosis of prior SARS-CoV-2 infection. Stability of eGFR, dd-cfDNA, GEP profiles and lack of allosensitization reinforce the safety profile of SARS-CoV-2 vaccination in KTRs. Further studies are needed to better understand immunogenicity in SARSCoV- 2 naive individuals, including whether cellular responses are protective in the absence of humoral responses.
ABSTRACT
Background: Approximately 70%-85% of patients who undergo sentinel lymph node biopsy (SLNb) show no nodal metastasis in the sentinel node (SN). The clinicopathological and gene expression profile (CP-GEP) model (Merlin Assay) was developed and validated to identify patients that may forgo the SLNb surgery due to their low risk for for nodal metastasis This study was initiated during the first wave of Covid-19 pandemic to allow for surgical triage on SLNb and evaluate the implementation of the Merlin assay in clinical practice. Methods: This study was conducted in four designated melanoma centers in the Netherlands. Patients (age > 18y) with newly diagnosed melanoma of the skin, eligible to undergo SLNb were screened for study inclusion. Main exclusion criteria was prior history of primary melanoma (> T1b) in the past 5 years. After enrollment, tissue sections of the primary melanoma were centrally reviewed at the Erasmus MC Cancer Institute to determine Breslow thickness at primary diagnosis. FFPE tumor tissue was dispatched for molecular analysis of eight target genes known to play a role in cancer development. In combination with age, Breslow thickness, and GEP outcome, risk of having nodal metastasis was calculated. Results were binary presented as 'CP-GEP low risk' and 'CPGEP high risk'. SLNb status was used as gold standard for comparison. Results: A total of 177 patients were analyzed using the CP-GEP model. Median age was 64 years (IQR 52-73) Median Breslow thickness was 1.4mm (IQR 1.0-2.4). Of all patients 28.2% was diagnosed with T1, 40.7% with T2 and 20.9% with T3 melanoma. Corresponding positivity rate was 7%, 14% and 29% respectively. A total of 24 out of 177 patients had a positive SLNb. Median turn-around time from inclusion to CP-GEP result was 15 days. Overall 37.1.% of patients had a CP-GEP low risk profile. The CP-GEP model had a NPV of 94.6%. Conclusions: This is the first prospective multicenter implementation study for the Merlin assay. Results are in line with previous validation studies. The CP-GEP model could accurately identify patients at low risk for SN metastasis. Implementation in clinical practice is feasible based on current turn-around time. In the future, using the Merlin assay to deselect patients for SLNB may allow for a reduction of surgery in patients with melanoma.
ABSTRACT
: Lung cancer is the most commonly occurring cancer in men worldwide. To search for new biological markers of this pathology, the transcriptome of the blood mononuclear cells from patients and healthy donors (residents of Kemerovo oblast, Russia) was studied using SurePrint G3 Human Gene Expression microarray technology. A total of 288 differentially expressed genes were identified, including 108 up-regulated genes and 180 down-regulated genes. Functional enrichment analysis using the WebGestalt resource and different databases (Gene Ontology, KEGG, and Reactome) indicated changes in the expression profiles of genes involved in the processes of immune response, protein synthesis, cell cycle control, and apoptosis. Analysis of protein–protein interactions using the STRING algorithm made it possible to identify functional clusters of gene products with different expression levels.