CD4 T cell epitope abundance in ferritin core potentiates responses to hemagglutinin nanoparticle vaccines.

Nanoparticle vaccines based on H. pylori ferritin are increasingly used as a vaccine platform for many pathogens, including RSV, influenza, and SARS-CoV-2. They have been found to elicit enhanced, long-lived B cell responses. The basis for improved efficacy of ferritin nanoparticle vaccines remains unresolved, including whether recruitment of CD4 T cells specific for the ferritin component of these vaccines contributes to cognate help in the B cell response. Using influenza HA-ferritin nanoparticles as a prototype, we have performed an unbiased assessment of the CD4 T cell epitope composition of the ferritin particles relative to that contributed by influenza HA using mouse models that express distinct constellations of MHC class II molecules. The role that these CD4 T cells play in the B cell responses was assessed by quantifying follicular helper cells (TFH), germinal center (GC) B cells, and antibody secreting cells. When mice were immunized with equimolar quantities of soluble HA-trimers and HA-Fe nanoparticles, HA-nanoparticle immunized mice had an increased overall abundance of TFH that were found to be largely ferritin-specific. HA-nanoparticle immunized mice had an increased abundance of HA-specific isotype-switched GC B cells and HA-specific antibody secreting cells (ASCs) relative to mice immunized with soluble HA-trimers. Further, there was a strong, positive correlation between CD4 TFH abundance and GC B cell abundance. Thus, availability of helper CD4 T cell epitopes may be a key additional mechanism that underlies the enhanced immunogenicity of ferritin-based HA-Fe-nanoparticle vaccines.

Development of a Mouse Model to Explore CD4 T Cell Specificity, Phenotype, and Recruitment to the Lung after Influenza B Infection.

The adaptive T cell response to influenza B virus is understudied, relative to influenza A virus, for which there has been considerable attention and progress for many decades. Here, we have developed and utilized the C57BL/6 mouse model of intranasal infection with influenza B (B/Brisbane/60/2008) virus and, using an iterative peptide discovery strategy, have identified a series of robustly elicited individual CD4 T cell peptide specificities. The CD4 T cell repertoire encompassed at least eleven major epitopes distributed across hemagglutinin, nucleoprotein, neuraminidase, and non-structural protein 1 and are readily detected in the draining lymph node, spleen, and lung. Within the lung, the CD4 T cells are localized to both lung vasculature and tissue but are highly enriched in the lung tissue after infection. When studied by flow cytometry and MHC class II: peptide tetramers, CD4 T cells express prototypical markers of tissue residency including CD69, CD103, and high surface levels of CD11a. Collectively, our studies will enable more sophisticated analyses of influenza B virus infection, where the fate and function of the influenza B-specific CD4 T cells elicited by infection and vaccination can be studied as well as the impact of anti-viral reagents and candidate vaccines on the abundance, functionality, and localization of the elicited CD4 T cells.

Potentiating Lung Mucosal Immunity Through Intranasal Vaccination.

Yearly administration of influenza vaccines is our best available tool for controlling influenza virus spread. However, both practical and immunological factors sometimes result in sub-optimal vaccine efficacy. The call for improved, or even universal, influenza vaccines within the field has led to development of pre-clinical and clinical vaccine candidates that aim to address limitations of current influenza vaccine approaches. Here, we consider the route of immunization as a critical factor in eliciting tissue resident memory (Trm) populations that are not a target of current licensed intramuscular vaccines. Intranasal vaccination has the potential to boost tissue resident B and T cell populations that reside within specific niches of the upper and lower respiratory tract. Within these niches, Trm cells are poised to respond rapidly to pathogen re-encounter by nature of their anatomic localization and their ability to rapidly deliver anti-pathogen effector functions. Unique features of mucosal immunity in the upper and lower respiratory tracts suggest that antigen localized to these regions is required for the elicitation of protective B and T cell immunity at these sites and will need to be considered as an important attribute of a rationally designed intranasal vaccine. Finally, we discuss outstanding questions and areas of future inquiry in the field of lung mucosal immunity.

Intranasal Nanoparticle Vaccination Elicits a Persistent, Polyfunctional CD4 T Cell Response in the Murine Lung Specific for a Highly Conserved Influenza Virus Antigen That Is Sufficient To Mediate Protection from Influenza Virus Challenge.

Lung-localized CD4 T cells play a critical role in the control of influenza virus infection and can provide broadly protective immunity. However, current influenza vaccination strategies primarily target influenza hemagglutinin (HA) and are administered peripherally to induce neutralizing antibodies. We have used an intranasal vaccination strategy targeting the highly conserved influenza nucleoprotein (NP) to elicit broadly protective lung-localized CD4 T cell responses. The vaccine platform consists of a self-assembling nanolipoprotein particle (NLP) linked to NP with an adjuvant. We have evaluated the functionality, in vivo localization, and persistence of the T cells elicited. Our study revealed that intranasal vaccination elicits a polyfunctional subset of lung-localized CD4 T cells that persist long term. A subset of these lung CD4 T cells localize to the airway, where they can act as early responders following encounter with cognate antigen. Polyfunctional CD4 T cells isolated from airway and lung tissue produce significantly more effector cytokines IFN-γ and TNF-α, as well as cytotoxic functionality. When adoptively transferred to naive recipients, CD4 T cells from NLP:NP-immunized lung were sufficient to mediate 100% survival from lethal challenge with H1N1 influenza virus. IMPORTANCE Exploiting new, more efficacious strategies to potentiate influenza virus-specific immune responses is important, particularly for at-risk populations. We have demonstrated the promise of direct intranasal protein vaccination to establish long-lived immunity in the lung with CD4 T cells that possess features and positioning in the lung that are associated with both immediate and long-term immunity, as well as demonstrating direct protective potential.

A Brief Intervention to Support Implementation of Telerehabilitation by Community Rehabilitation Services During COVID-19: A Feasibility Study.

The coronavirus disease 2019 (COVID-19) pandemic has necessitated adoption of telerehabilitation in services where face-to-face consultations were previously standard. We aimed to understand barriers to implementing a telerehabilitation clinical service and design a behavior support strategy for clinicians to implement telerehabilitation. A hybrid implementation study design included pre- and post-intervention questionnaires, identification of key barriers to implementation using the theoretical domains framework, and development of a targeted intervention. Thirty-one clinicians completed baseline questionnaires identifying key barriers to the implementation of telerehabilitation. Barriers were associated with behavior domains of knowledge, environment, social influences, and beliefs. A 6-week brief intervention focused on remote clinician support, and education was well received but achieved little change in perceived barriers to implementation. The brief intervention to support implementation of telerehabilitation during COVID-19 achieved clinical practice change, but barriers remain. Longer follow-up may determine the sustainability of a brief implementation strategy, but needs to consider pandemic-related stressors.

Effect of age on the failure properties of human meniscus: High-speed strain mapping of tissue tears.

The knee meniscus is a soft fibrous tissue with a high incidence of injury in older populations. The objective of this study was to determine the effect of age on the failure behavior of human knee meniscus when applying uniaxial tensile loads parallel or perpendicular to the primary circumferential fiber orientation. Two age groups were tested: under 40 and over 65 years old. We paired high-speed video with digital image correlation to quantify for the first time the planar strains occurring in the tear region at precise time points, including at ultimate tensile stress, when the tissue begins losing load-bearing capacity. On average, older meniscus specimens loaded parallel to the fiber axis had approximately one-third less ultimate tensile strain and absorbed 60% less energy to failure within the tear region than younger specimens (p < 0.05). Older specimens also had significantly reduced strength and material toughness when loaded perpendicular to the fibers (p < 0.05). These age-related changes indicate a loss of collagen fiber extensibility and weakening of the non-fibrous matrix with age. In addition, we found that when loaded perpendicular to the circumferential fibers, tears propagated near the planes of maximum tensile stress and strain. Whereas when loaded parallel to the circumferential fibers, tears propagated oblique to the loading axis, closer to the planes of maximum shear stress and strain. Our experimental results can assist the selection of valid failure criteria for meniscus, and provide insight into the effect of age on the failure mechanisms of soft fibrous tissue.

Print-A-Punch: A 3D printed device to cut dumbbell-shaped specimens from soft tissue for tensile testing.

The failure behavior and mechanical properties of soft tissue can be characterized by conducting uniaxial tensile tests on small sectioned specimens, called test coupons. An ideal coupon geometry for tensile testing is a dumbbell shape (dog-bone), yet the cost and time required to fabricate custom steel punches to cut dumbbell-shaped coupons has hindered their universal application in biomechanics research. In this study, we developed an economical and reliable cutting device that can extract dumbbell-shaped coupons from soft biological tissue. The novel device, called Print-A-Punch, uses three-dimensional (3D) printed components in combination with standard fasteners and replaceable flexible razors. We identified design factors that influence the dimensional accuracy and symmetry of elastomer coupons extracted using this cutting device, and demonstrated its use on bovine meniscus. Advantages of this 3D printed device include a fast fabrication time, low material cost, good accuracy, replaceable blades, and an ability to scale coupon dimensions for specific tissues and experiments. By reducing the cost and time to cut accurate dumbbell-shaped coupons, this technology can facilitate the broad adoption of standard test methods that improve the quality and reproducibility of tensile tests in soft biological tissue. Researchers can freely download a set of STL files from this study to build their own Print-A-Punch device (https://boisestate.edu/coen-ntm/technology/print-a-punch).

Imprinting and Editing of the Human CD4 T Cell Response to Influenza Virus.

Immunity to influenza is unique among pathogens, in that immune memory is established both via intermittent lung localized infections with highly variable influenza virus strains and by intramuscular vaccinations with inactivated protein-based vaccines. Studies in the past decades have suggested that the B cell responses to influenza infection and vaccination are highly biased by an individual's early history of influenza infection. This reactivity likely reflects both the competitive advantage that memory B cells have in an immune response and the relatively limited diversity of epitopes in influenza hemagglutinin that are recognized by B cells. In contrast, CD4 T cells recognize a wide array of epitopes, with specificities that are heavily influenced by the diversity of influenza antigens available, and a multiplicity of functions that are determined by both priming events and subsequent confrontations with antigens. Here, we consider the events that prime and remodel the influenza-specific CD4 T cell response in humans that have highly diverse immune histories and how the CD4 repertoire may be edited in terms of functional potential and viral epitope specificity. We discuss the consequences that imprinting and remodeling may have on the potential of different human hosts to rapidly respond with protective cellular immunity to infection. Finally, these issues are discussed in the context of future avenues of investigation and vaccine strategies.

Mucosal Mesenchymal Cells: Secondary Barrier and Peripheral Educator for the Gut Immune System.

Stromal connective tissue contains mesenchymal cells, including fibroblasts and myofibroblasts, which line the tissue structure. However, it has been identified that the function of mesenchymal cells is not just structural-they also play critical roles in the creation and regulation of intestinal homeostasis. Thus, mucosal mesenchymal cells instruct intestinal immune cell education (or peripheral immune education) and epithelial cell differentiation thereby shaping the local environment of the mucosal immune system. Malfunction of the mesenchymal cell-mediated instruction system (e.g., fibrosis) leads to pathological conditions such as intestinal stricture.

Outbreak of Escherichia coli O157:H7 in a child care center in Cook County, Illinois, with prolonged shedding and household transmission.

During the investigation of an outbreak of Escherichia coli O157:H7 in a child care center, illness logs were reviewed and parents interviewed to identify classroom and household exposures. Costs incurred by the center and the public health laboratory were estimated from self-administered questionnaires. We conclude that household transmission played a role in this outbreak and estimate the cost of investigation and intervention at over $6,000 per case.