homeRNA self-blood collection enables high-frequency temporal profiling of pre-symptomatic host immune kinetics to respiratory viral infection: a prospective cohort study.

Background: Early host immunity to acute respiratory infections (ARIs) is heterogenous, dynamic, and critical to an individual's infection outcome. Due to limitations in sampling frequency/timepoints, kinetics of early immune dynamics in natural human infections remain poorly understood. In this nationwide prospective cohort study, we leveraged a self-blood collection tool (homeRNA) to profile detailed kinetics of the pre-symptomatic to convalescence host immunity to contemporaneous respiratory pathogens. Methods: We enrolled non-symptomatic adults with recent exposure to ARIs who subsequently tested negative (exposed-uninfected) or positive for respiratory pathogens. Participants self-collected blood and nasal swabs daily for seven consecutive days followed by weekly blood collection for up to seven additional weeks. Symptom burden was assessed during each collection. Nasal swabs were tested for SARS-CoV-2 and common respiratory pathogens. 92 longitudinal blood samples spanning the pre-shedding to post-acute phase of eight SARS-CoV-2-infected participants and 40 interval-matched samples from four exposed-uninfected participants were subjected to high-frequency longitudinal profiling of 773 host immune genes. Findings: Between June 2021 - April 2022, 68 participants across 26 U.S. states completed the study and self-collected a total of 691 and 466 longitudinal blood and nasal swab samples along with 688 symptom surveys. SARS-CoV-2 was detected in 17 out of 22 individuals with study-confirmed respiratory infection. With rapid dissemination of home self-collection kits, two and four COVID-19+ participants started collection prior to viral shedding and symptom onset, respectively, enabling us to profile detailed expression kinetics of the earliest blood transcriptional response to contemporaneous variants of concern. In pre-shedding samples, we observed transient but robust expression of T-cell response signatures, transcription factor complexes, prostaglandin biosynthesis genes, pyrogenic cytokines, and cytotoxic granule genes. This is followed by a rapid induction of many interferon-stimulated genes (ISGs), concurrent to onset of viral shedding and increase in nasal viral load. Finally, we observed increased expression of host defense peptides (HDPs) in exposed-uninfected individuals over the 4-week observational window. Interpretation: We demonstrated that unsupervised self-collection and stabilization of capillary blood can be applied to natural infection studies to characterize detailed early host immune kinetics at a temporal resolution comparable to that of human challenge studies. The remote (decentralized) study framework enables conduct of large-scale population-wide longitudinal mechanistic studies. Expression of cytotoxic/T-cell signatures in pre-shedding samples preceding expansion of innate ISGs suggests a potential role for T-cell mediated pathogen control during early infection. Elevated expression of HDPs in exposed-uninfected individuals warrants further validation studies to assess their potential role in protective immunity during pathogen exposure. Funding: This study was funded by R35GM128648 to ABT for in-lab developments of homeRNA, Packard Fellowship from the David and Lucile Packard Foundation to ABT, and R01AI153087 to AW.

Longitudinal home self-collection of capillary blood using homeRNA correlates interferon and innate viral defense pathways with SARS-CoV-2 viral clearance.

Blood transcriptional profiling is a powerful tool to evaluate immune responses to infection; however, blood collection via traditional phlebotomy remains a barrier to precise characterization of the immune response in dynamic infections (e.g., respiratory viruses). Here we present an at-home self-collection methodology, homeRNA, to study the host transcriptional response during acute SARS-CoV-2 infections. This method uniquely enables high frequency measurement of the host immune kinetics in non-hospitalized adults during the acute and most dynamic stage of their infection. COVID-19+ and healthy participants self-collected blood every other day for two weeks with daily nasal swabs and symptom surveys to track viral load kinetics and symptom burden, respectively. While healthy uninfected participants showed remarkably stable immune kinetics with no significant dynamic genes, COVID-19+ participants, on the contrary, depicted a robust response with over 418 dynamic genes associated with interferon and innate viral defense pathways. When stratified by vaccination status, we detected distinct response signatures between unvaccinated and breakthrough (vaccinated) infection subgroups; unvaccinated individuals portrayed a response repertoire characterized by higher innate antiviral responses, interferon signaling, and cytotoxic lymphocyte responses while breakthrough infections portrayed lower levels of interferon signaling and enhanced early cell-mediated response. Leveraging cross-platform longitudinal sampling (nasal swabs and blood), we observed that IFI27, a key viral response gene, tracked closely with SARS-CoV-2 viral clearance in individual participants. Taken together, these results demonstrate that at-home sampling can capture key host antiviral responses and facilitate frequent longitudinal sampling to detect transient host immune kinetics during dynamic immune states.

Freestanding hydrogel lumens for modeling blood vessels and vasodilation.

Lumen structures exist throughout the human body, and the vessels of the circulatory system are essential for carrying nutrients and oxygen and regulating inflammation. Vasodilation, the widening of the blood vessel lumen, is important to the immune response as it increases blood flow to a site of inflammation, raises local temperature, and enables optimal immune system function. A common method for studying vasodilation uses excised vessels from animals; major drawbacks include heterogeneity in vessel shape and size, time-consuming procedures, sacrificing animals, and differences between animal and human biology. We have developed a simple, user-friendly in vitro method to form freestanding cell-laden hydrogel rings from collagen and quantitatively measure the effects of vasodilators on ring size. The hydrogel rings are composed of collagen I and can be laden with human vascular smooth muscle cells, a major cellular and structural component of blood vessels, or lined with endothelial cells in the lumen. The methods presented include a 3D printed device (which is amenable to future fabrication by injection molding) and commercially available components (e.g., Teflon tubing or a syringe) to form hydrogel rings between 2.6-4.6 mm outer diameter and 0.79-1.0 mm inner diameter. Here we demonstrate a significant difference in ring area in the presence of a known vasodilator, fasudil (p < 0.0001). Our method is easy to implement and provides a foundation for a medium-throughput solution to generating vessel model structures for future investigations of the fundamental mechanisms of vasodilation (e.g., studying uncharacterized endogenous molecules that may have vasoactivity) and testing vasoactive drugs.