improved-methodology-for-liquid-delivery-to-mouse-lung-intubation

Improved Methodology for Liquid Delivery to the Mouse Lung: Intubation using a Consumer Otoscope

Consistent delivery of liquids, including potential therapies, to the lungs is critical for modeling pulmonary disease and pre-clinical therapeutic ...

Tulane University School of Medicine

Gene Therapy for Occupational Lung Disease: Steering Macrophages in the Right Direction?

Single-cell RNA-seq has been used to characterize human COVID-19. To determine if preclinical models successfully mimic the cell-intrinsic and -extrinsic effects of severe disease, we conducted a meta-analysis of single-cell data across five model species. To assess whether dissemination of viral RNA in lung cells tracks pathology and results in cell-intrinsic and -extrinsic transcriptomic changes in COVID-19. We conducted a meta-analysis by analyzing six publicly available, scRNA-seq data sets. We used dual mapping (host and virus) and differential gene expression analyses to compare viral and viral cell populations. We conducted a principal component analysis to identify successful models of human COVID-19. We found expression of viral RNA in many non-epithelial cell types. Fibroblasts, macrophages, and endothelial cells exhibit clear evidence of viral-intrinsic and -extrinsic effects on host gene expression. Using viral RNA expression, we found that K18-hACE2 mice most closely modeled severe human COVID-19, followed by hamsters. Ferrets and macaques are poor models of human disease due to the low presence of viral RNA. Moreover, we found that increased transcripts of certain key inflammatory genes such as , and  are not restricted to virally infected cells, suggesting these genes are regulated in a paracrine or autocrine fashion. These data affirm widespread dissemination of viral RNA in the lung, which may be key in the pathogenesis of severe COVID-19 and demonstrate ferrets and Rhesus macaques are poor models of human COVID-19. IMPORTANCE We conducted a high-resolution meta-analysis of scRNA-seq data from humans and five animal models of COVID-19. This study reports viral RNA dissemination in several cell types in human data as well as in some of the pre-clinical models. Using this metric, the K18-hACE2 mouse model, followed by the hamster model, most closely resembled human COVID-19. We observed clear evidence of viral-intrinsic effects within cells (e.g.,  expression) as well as viral-extrinsic cytokine modulation (e.g., ). We observed proinflammatory chemokine expression in cells devoid of viral RNA expression, suggesting autocrine/paracrine interferon regulation. This report serves as a resource-synthesizing data from COVID-19 humans and animal models and suggesting improvements for relevant pre-clinical models that may aid future diagnostic and therapeutic development projects.

Cell-intrinsic and -extrinsic effects of SARS-CoV-2 RNA on pathogenesis: single-cell meta-analysis.

Lung tissue resident memory (TRM) cells are thought to play crucial roles in lung host defense. We have recently shown that immunization with the adjuvant LTA1 (derived from the A1 domain of E. coli heat labile toxin) admixed with OmpX from K. pneumoniae can elicit antigen specific lung Th17 TRM cells that provide serotype independent immunity to members of the Enterobacteriaceae family. However, the upstream requirements to generate these cells are unclear. Single-cell RNA-seq showed that vaccine-elicited Th17 TRM cells expressed high levels of IL-1R1, suggesting that IL-1 family members may be critical to generate these cells. Using a combination of genetic and antibody neutralization approaches, we show that Th17 TRM cells can be generated independent of caspase-1 but are compromised when IL-1α is neutralized. Moreover IL-1α could serve as a molecular adjuvant to generate lung Th17 TRM cells independent of LTA1. Taken together, these data suggest that IL-1α plays a major role in vaccine-mediated lung Th17 TRM generation.

Vaccine-elicited IL-1R signaling results in Th17 TRM-mediated immunity.

Although the last decade has seen tremendous progress in drugs that treat cystic fibrosis (CF) due to mutations that lead to protein misfolding, there are approximately 8%-10% of subjects with mutations that result in no significant CFTR protein expression demonstrating the need for gene editing or gene replacement with inhaled mRNA or vector-based approaches. A limitation for vector-based approaches is the formation of neutralizing humoral responses. Given that αCD20 has been used to manage post-transplant lymphoproliferative disease in CF subjects with lung transplants, we studied the ability of αCD20 to module both T and B cell responses in the lung to one of the most immunogenic vectors, E1-deleted adenovirus serotype 5. We found that αCD20 significantly blocked luminal antibody responses and efficiently permitted re-dosing. αCD20 had more limited impact on the T cell compartment, but reduced tissue resident memory T cell responses in bronchoalveolar lavage fluid. Taken together, these pre-clinical studies suggest that αCD20 could be re-purposed for lung gene therapy protocols to permit re-dosing.

Article	Year
Gene Therapy for Occupational Lung Disease: Steering Macrophages in the Right Direction? American journal of respiratory cell and molecular biology\| PubMed ID: 36315436	2023
Cell-intrinsic and -extrinsic effects of SARS-CoV-2 RNA on pathogenesis: single-cell meta-analysis. mSphere\| PubMed ID: 37737611	2023
Vaccine-elicited IL-1R signaling results in Th17 TRM-mediated immunity. Communications biology\| PubMed ID: 38594380	2024
Evaluation of anti-vector immune responses to adenovirus-mediated lung gene therapy and modulation by αCD20. Molecular therapy. Methods & clinical development\| PubMed ID: 39070292	2024
Running Helps: Medical Students and Attending Physicians are on the Same Track. Southern medical journal\| PubMed ID: 39486457	2024

T. Parks Remcho

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