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Background/Objectives: Genetic factors influence COVID-19 susceptibility and outcomes, including the development of pulmonary fibrosis (i.e. lung scarring). Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease and the most common cause of pulmonary fibrosis in the general population. Genome-wide association studies (GWAS) of COVID-19 and IPF revealed genes associated with both diseases, suggesting these share genetic risk factors. Here we performed a genetic overlap study between COVID-19 and IPF. Method(s): Summary statistics from an IPF 5-way meta-GWAS and from the COVID-19 Host Genetics initiative GWAS metaanalysis (v6) were used. We performed genetic correlation analyses and assessed individual genetic signals to identify those variants shared between both traits. We conducted colocalisation analyses to determine whether the same causal variant was driving both traits. Finally, the association of overlapping variants with gene expression was assessed and a phenome-wide association study was performed. Result(s): There was a positive genetic correlation between severe COVID-19 and IPF. We found four genetic loci with likely shared causal variants between both traits, including one novel risk locus at 7q22.1 that colocalised with decreased ZKSCAN1 and TRIM4 expression in blood. The other three loci colocalised with MUC5B, ATP11A and DPP9 expression. The locus associated with increased ATP11A expression was also associated with higher Hb1AC levels, a biomarker used in diabetes. Conclusion(s): Results suggest there are shared biological processes driving IPF and severe COVID-19 phenotypes.
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BACKGROUND: Genetic susceptibility to infectious diseases is partly due to the variation in the human genome, and COVID-19 is not the exception. This study aimed to identify whether risk alleles of known genes linked with emphysema (SERPINA1) and pulmonary fibrosis (MUC5B) are associated with severe COVID-19, and whether plasma mucin 5B differs according to patients' outcomes. MATERIALS AND METHODS: We included 1258 Mexican subjects diagnosed with COVID-19. We genotyped rs2892474 and rs17580 of the SERPINA1 gene and rs35705950 of MUC5B. Based on the rs35705950 genotypes, mucin 5B plasma protein levels were quantified. RESULTS: Homozygous for the risk alleles of the three polymorphisms were found in less than 5% of the study population, but no statistically significant difference in the genotype or allele association analysis. At the protein level, non-survivors carrying one or two copies of the risk allele rs35705950 in MUC5B (GT + TT) had lower levels of mucin 5B compared to the survivors (0.0 vs. 0.17 ng/mL, p = 0.0013). CONCLUSION: The polymorphisms rs28929474 and rs17580 of SERPINA1 and rs35705950 of MUC5B are not associated with the risk of severe COVID-19 in the Mexican population. COVID-19 survivor patients bearing one or two copies of the rs35705950 risk allele have higher plasma levels of mucin 5B.
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Rationale: Recent advancements in sequencing technologies have led to a substantial increase in the scale and resolution of transcriptomic data. Despite this progress, accessibility to this data, particularly among those who are coming from non-computational backgrounds is limited. To facilitate improved access and exploration of our single-cell RNA sequencing data, we generated several data sharing, mining and dissemination portals to accompany our idiopathic pulmonary fibrosis (IPF), chronic obstructive pulmonary disease (COPD), and lung endothelial cells (Lung EC) cell atlases. Descriptions and links of each website can be found here: https://medicine.yale.edu/lab/kaminski/research/atlas/. Methods: Each interactive data mining website is coded in the R language using the Shiny package and is hosted by Shinyapps.io. Percell expression data for each website is stored on a MySQL database hosted by Amazon Web Services (AWS). Time-associated website engagement statistics and gene query information is collected for each website using a combination of Google Analytics and a gene search table stored on our MySQL database. User exploration of available data is facilitated through several easy-touse visualization tools available on each website. Results: Website usage statistics since the publication of each website shows that 9,772 unique users from 56 countries and five continents have accessed at least one of the three websites. At the time of writing, 300,748 total queries have been made for 15,627 unique genes across the websites. The top five searched genes for the IPF Cell Atlas are CD14, ACE2, ACTA2, IL11 and MUC5B while for the COPD Cell Atlas they are FAM13A, MIRLET7BHG, HHIP, ISM1 and DDT. Finally, the top searched genes for the Lung Endothelial Cell Atlas are BMPR2, PECAM1, EDNRB, APLNR and PROX1. Of note, interaction with the IPF Cell Atlas increased dramatically at the start of the COVID-19 pandemic, with queries for the ACE2 gene, the putative binding receptor for the SARS-CoV-2 virus, increasing substantially at the pandemic's onset in the United States. Conclusions: Usage statistics, gene query information and feedback from users, both within academia and industry, have shown broad engagement with our websites by individuals across computational and non-computational backgrounds. We envision widespread adoption of web-based portals similar to ours will facilitate novel discoveries within these complex datasets and new scientific collaborations.
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Rationale There is a lack of knowledge of how CFTR-deficient airway epithelium intrinsically responds to SARS-CoV-2. Though prior work has demonstrated altered CF airway expression of viral entry factors, it is unknown whether these alterations are protective and whether they reflect host genetic variation or secondary response of chronic inflammation. We address this gap by infecting induced pluripotent stem cell (iPSC)-derived airways from CF patients and syngeneic CFTR-corrected controls with SARS-CoV-2 and assessing differential susceptibility to infection and inflammatory and anti-viral response. MethodsCF (F508del homozygous) and syngeneic CFTR-corrected (CRISPR-Cas9) iPSC- were differentiated into airway epithelium cultured at airliquid interface (ALI) by a directed differentiation protocol that generates a pure population of major and rare airway cell-types. After 21 days in ALI culture, the iPSC-airway were infected with either mock or SARS-CoV-2 (isolate USA-WA1/2020) with MOI of 4, and harvested at 0, 1, 3 days post infection (dpi) for RT-PCR and immune-stainingResultsBoth CF and CFTR-corrected iPSC-airway express viral entry factors of ACE2 and TMPRSS2, and are permissive to SARS-CoV-2 infection. CF iPSC-airway exhibited significantly increase in SARS-CoV-2 nucleocapsid protein (N) transcript at 1 dpi, accompanied by increases in IFN2, RSAD2, and CXCL10 at 3 dpi, compared to its CFTR-corrected counter-part. There are no baseline significant differences in ACE2, TMPRSS2, TP63, NGFR, MUC5B, MUC5AC, SCGB1A1, FOXJ1, FOXI1 expression between CF and CFTR-corrected iPSC-airway before SARS-CoV-2 infection. ConclusionsOur preliminary studies indicate increased early SARS-CoV-2 infection in CFTR-deficient epithelium with accompanied subsequent rise in anti-viral and inflammatory response compared to its genetically controlled CFTR-corrected counterpart. Future studies are aimed at assessing differential CF epithelial kinetics of SARS-CoV-2 viral entry and replication, morphological changes, global transcriptomic response, and how treatment with CFTRmodulator would alter the epithelial response. Ultimately, we aim to establish a reductionist, physiologically relevant model system that is coupled with gene-editing technology to study intrinsic CF epithelial response to SARS-CoV-2, which would generate insights to aid practice guidelines for CF patients, and open future directions to evaluate gene-specific mechanisms of airway response to pathogens. (Figure Presented).
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Introduction: Due to Covid-19 restrictions on collecting and processing sputum samples in real time in clinic, we designed a novel sputum home collection method with immediate freezing and delayed processing (“home”). A validation study was carried out to compare key sputum endpoints using the “home” vs “real time (RT)” collection and processing methods. Sputum soluble phase proteomics, mucins and RNA/DNA endpoints were measured and compared between the 2 methods to assess the validity of the “home” method. Methods: Spontaneous sputum samples were collected from N=10 healthy adult volunteers. Each sample was split evenly by weight and processed, half by the “home” method and half by the RT method. Home method samples were first aliquoted into 3 collection tubes (T) as follows: T1: 100-250mg for mucin analysis (refractive index, gel chromatography, and CsCl gradients);T2 and T3: equal weights each, T2 for proteomic analysis (MesoScale Discovery) and T3 for RNA/DNA analysis (Isohelix collection kit). Each was immediately frozen at -20 deg C (24-48hr), then at -80 deg C (2-4 weeks) without any processing. Thawed home T1 and T2 samples were processed by treating with 8M Urea (1:1) to deactivate SARS-CoV-2 if present. T1 was then stored at 2-4 deg C, and T2 was processed with 7x DPBS, centrifuged and recovered supernatants stored at -80 deg C. In contrast, the RT sputum was first treated with 8M Urea (1:1) soon after collection, and then processed for mucins and proteomics per the “home” method above. The remaining cell pellet from the RT processed sample was stored in Zymo research RNA/DNA shield (0.5ml) and, along with home T3 samples, extracted and analyzed for qualitative and quantitative yield, as well as for genes of interest. Paired T-Test analysis compared all sputum endpoints between the home and RT method. Results: There were no statistically significant differences (p<0.05) between the home and RT method for any mucin (MUC5B, MUC5AC, MUC5AC:MUC5B ratio, total mucin) or proteomic endpoint (IL-1a, IL-6, IL-8, TNFalpha, TIMP1, TIMP2, MMP-9, CRP, MPO). In addition, except for CRP and MUC5AC, correlation between sample pairings was strong (correlation coefficient R, range = 0.5-0.9) and statistically significant (p<0.05) for all sputum endpoints. RNA/DNA results are still pending. Conclusion: The sputum “home collection method with immediate freezing and delayed processing” does not result in significantly different proteomic and mucin measurements when compared to the same samples being processed in real time in an identical manner.