LPS-induced lung tissue-resident trained innate immunity provides differential protection against pneumococci and SARS-CoV-2.

Recent evidence indicates that tissue-resident innate immune memory and trained innate immunity (TII) can be induced centrally in myeloid cells within the bone marrow and locally in tissue-resident macrophages in respiratory mucosal tissues. However, it remains unclear whether acute exposure to airborne microbial components like lipopolysaccharide (LPS) induces lasting innate immune memory in airway macrophages and TII capable of protection against heterologous pathogens. Using a murine model, we demonstrate that acute LPS exposure leads to dynamic changes in the immune phenotype of airway macrophages that persist long after the acute inflammatory response has subsided. The original airway-resident alveolar macrophage pool remains stable in size despite these changes and the earlier transient acute inflammatory responses, including monocytic recruitment in the lung. We further demonstrate that the induction of innate immune memory in airway macrophages is accompanied by TII capable of robust protection against acute pneumococcal infection, whereas it provides minimal protection against acute SARS-CoV-2 infection.

Cannabis smoke suppresses antiviral immune responses to influenza A in mice.

Rationale: Despite its increasingly widespread use, little is known about the impact of cannabis smoking on the response to viral infections like influenza A virus (IAV). Many assume that cannabis smoking will disrupt antiviral responses in a manner similar to cigarette smoking; however, since cannabinoids exhibit anti-inflammatory effects, cannabis smoke exposure may impact viral infection in distinct ways. Methods: Male and female BALB/c mice were exposed daily to cannabis smoke and concurrently intranasally instilled with IAV. Viral burden, inflammatory mediator levels (multiplex ELISA), lung immune cells populations (flow cytometry) and gene expression patterns (RNA sequencing) were assessed in the lungs. Plasma IAV-specific antibodies were measured via ELISA. Results: We found that cannabis smoke exposure increased pulmonary viral burden while decreasing total leukocytes, including macrophages, monocytes and dendritic cell populations in the lungs. Furthermore, infection-induced upregulation of certain inflammatory mediators (interferon-γ and C-C motif chemokine ligand 5) was blunted by cannabis smoke exposure, which in females was linked to the transcriptional downregulation of pathways involved in innate and adaptive immune responses. Finally, plasma levels of IAV-specific IgM and IgG1 were significantly decreased in cannabis smoke-exposed, infected mice compared to infected controls, only in female mice. Conclusions: Overall, cannabis smoke exposure disrupted host-defence processes, leading to increased viral burden and dampened inflammatory signalling. These results suggest that cannabis smoking is detrimental to the maintenance of pulmonary homeostasis during viral infection and highlight the need for data regarding the impact on immune competency in humans.

Viral-vectored respiratory mucosal vaccine strategies.

Increasing global concerns of pandemic respiratory viruses highlight the importance of developing optimal vaccination strategies that encompass vaccine platform, delivery route, and regimens. The decades-long effort to develop vaccines to combat respiratory infections such as influenza, respiratory syncytial virus, and tuberculosis has met with challenges, including the inability of systemically administered vaccines to induce respiratory mucosal (RM) immunity. In this regard, ample preclinical and available clinical studies have demonstrated the superiority of RM vaccination to induce RM immunity over parenteral route of vaccination. A great stride has been made in developing vaccines for RM delivery against respiratory pathogens, including M. tuberculosis and SARS-CoV-2. In particular, inhaled aerosol delivery of adenoviral-vectored vaccines has shown significant promise.

Adenoviral-vectored next-generation respiratory mucosal vaccines against COVID-19.

The world is in need of next-generation COVID-19 vaccines. Although first-generation injectable COVID-19 vaccines continue to be critical tools in controlling the current global health crisis, continuous emergence of SARS-CoV-2 variants of concern has eroded the efficacy of these vaccines, leading to staggering breakthrough infections and posing threats to poor vaccine responders. This is partly because the humoral and T-cell responses generated following intramuscular injection of spike-centric monovalent vaccines are mostly confined to the periphery, failing to either access or be maintained at the portal of infection, the respiratory mucosa (RM). In contrast, respiratory mucosal-delivered vaccine can induce immunity encompassing humoral, cellular, and trained innate immunity positioned at the respiratory mucosa that may act quickly to prevent the establishment of an infection. Viral vectors, especially adenoviruses, represent the most promising platform for RM delivery that can be designed to express both structural and nonstructural antigens of SARS-CoV-2. Boosting RM immunity via the respiratory route using multivalent adenoviral-vectored vaccines would be a viable next-generation vaccine strategy.

Subcutaneous BCG vaccination protects against streptococcal pneumonia via regulating innate immune responses in the lung.

Bacillus Calmette-Guérin (BCG) still remains the only licensed vaccine for TB and has been shown to provide nonspecific protection against unrelated pathogens. This has been attributed to the ability of BCG to modulate the innate immune system, known as trained innate immunity (TII). Trained innate immunity is associated with innate immune cells being in a hyperresponsive state leading to enhanced host defense against heterologous infections. Both epidemiological evidence and prospective studies demonstrate cutaneous BCG vaccine-induced TII provides enhanced innate protection against heterologous pathogens. Regardless of the extensive progress made thus far, the effect of cutaneous BCG vaccination against heterologous respiratory bacterial infections and the underlying mechanisms still remain unknown. Here, we show that s.c. BCG vaccine-induced TII provides enhanced heterologous innate protection against pulmonary Streptococcus pneumoniae infection. We further demonstrate that this enhanced innate protection is mediated by enhanced neutrophilia in the lung and is independent of centrally trained circulating monocytes. New insight from this study will help design novel effective vaccination strategies against unrelated respiratory bacterial pathogens.

Intranasal multivalent adenoviral-vectored vaccine protects against replicating and dormant M.tb in conventional and humanized mice.

Viral-vectored vaccines are highly amenable for respiratory mucosal delivery as a means of inducing much-needed mucosal immunity at the point of pathogen entry. Unfortunately, current monovalent viral-vectored tuberculosis (TB) vaccine candidates have failed to demonstrate satisfactory clinical protective efficacy. As such, there is a need to develop next-generation viral-vectored TB vaccine strategies which incorporate both vaccine antigen design and delivery route. In this study, we have developed a trivalent chimpanzee adenoviral-vectored vaccine to provide protective immunity against pulmonary TB through targeting antigens linked to the three different growth phases (acute/chronic/dormancy) of Mycobacterium tuberculosis (M.tb) by expressing an acute replication-associated antigen, Ag85A, a chronically expressed virulence-associated antigen, TB10.4, and a dormancy/resuscitation-associated antigen, RpfB. Single-dose respiratory mucosal immunization with our trivalent vaccine induced robust, sustained tissue-resident multifunctional CD4+ and CD8+ T-cell responses within the lung tissues and airways, which were further quantitatively and qualitatively improved following boosting of subcutaneously BCG-primed hosts. Prophylactic and therapeutic immunization with this multivalent trivalent vaccine in conventional BALB/c mice provided significant protection against not only actively replicating M.tb bacilli but also dormant, non-replicating persisters. Importantly, when used as a booster, it also provided marked protection in the highly susceptible C3HeB/FeJ mice, and a single respiratory mucosal inoculation was capable of significant protection in a humanized mouse model. Our findings indicate the great potential of this next-generation TB vaccine strategy and support its further clinical development for both prophylactic and therapeutic applications.

Corrigendum: Differential biodistribution of adenoviral-vectored vaccine following intranasal and endotracheal deliveries leads to different immune outcomes.

[This corrects the article DOI: 10.3389/fimmu.2022.860399.].

Parenteral BCG vaccine induces lung-resident memory macrophages and trained immunity via the gut-lung axis.

Aside from centrally induced trained immunity in the bone marrow (BM) and peripheral blood by parenteral vaccination or infection, evidence indicates that mucosal-resident innate immune memory can develop via a local inflammatory pathway following mucosal exposure. However, whether mucosal-resident innate memory results from integrating distally generated immunological signals following parenteral vaccination/infection is unclear. Here we show that subcutaneous Bacillus Calmette-Guérin (BCG) vaccination can induce memory alveolar macrophages (AMs) and trained immunity in the lung. Although parenteral BCG vaccination trains BM progenitors and circulating monocytes, induction of memory AMs is independent of circulating monocytes. Rather, parenteral BCG vaccination, via mycobacterial dissemination, causes a time-dependent alteration in the intestinal microbiome, barrier function and microbial metabolites, and subsequent changes in circulating and lung metabolites, leading to the induction of memory macrophages and trained immunity in the lung. These data identify an intestinal microbiota-mediated pathway for innate immune memory development at distal mucosal tissues and have implications for the development of next-generation vaccine strategies against respiratory pathogens.

Comparing Current and Next-Generation Humanized Mouse Models for Advancing HIV and HIV/Mtb Co-Infection Studies.

In people living with HIV, Mycobacterium tuberculosis (Mtb) is the major cause of death. Due to the increased morbidity/mortality in co-infection, further research is urgently required. A limiting factor to research in HIV and HIV/Mtb co-infection is the lack of accessible in vivo models. Next-generation humanized mice expressing HLA transgenes report improved human immune reconstitution and functionality, which may better recapitulate human disease. This study compares well-established huNRG mice and next-generation HLA I/II-transgenic (huDRAG-A2) mice for immune reconstitution, disease course, and pathology in HIV and TB. HuDRAG-A2 mice have improved engraftment of key immune cell types involved in HIV and TB disease. Upon intravaginal HIV-1 infection, both models developed significant HIV target cell depletion in the blood and tissues. Upon intranasal Mtb infection, both models sustained high bacterial load within the lungs and tissue dissemination. Some huDRAG-A2 granulomas appeared more classically organized, characterized by focal central necrosis, multinucleated giant cells, and foamy macrophages surrounded by a halo of CD4+ T cells. HIV/Mtb co-infection in huNRG mice trended towards worsened TB pathology and showed potential for modeling co-infection. Both huNRG and huDRAG-A2 mice are viable options for investigating HIV and TB, but the huDRAG-A2 model may offer advantages.

Protocol for isolation and characterization of lung tissue resident memory T cells and airway trained innate immunity after intranasal vaccination in mice.

Vaccination route dictates the quality and localization of immune responses within tissues. Intranasal vaccination seeds tissue-resident adaptive immunity, alongside trained innate responses within the lung/airways, critical for superior protection against SARS-CoV-2. This protocol encompasses intranasal vaccination in mice, step-by-step bronchoalveolar lavage for both cellular and acellular airway components, lung mononuclear cell isolation, and detailed flow cytometric characterization of lung tissue-resident memory T cell responses, and airway macrophage-trained innate immunity. For complete details on the use and execution of this protocol, please refer to Afkhami et al. (2022).

Differential Biodistribution of Adenoviral-Vectored Vaccine Following Intranasal and Endotracheal Deliveries Leads to Different Immune Outcomes.

Infectious diseases of the respiratory tract are one of the top causes of global morbidity and mortality with lower respiratory tract infections being the fourth leading cause of death. The respiratory mucosal (RM) route of vaccine delivery represents a promising strategy against respiratory infections. Although both intranasal and inhaled aerosol methods have been established for human application, there is a considerable knowledge gap in the relationship of vaccine biodistribution to immune efficacy in the lung. Here, by using a murine model and an adenovirus-vectored model vaccine, we have compared the intranasal and endotracheal delivery methods in their biodistribution, immunogenicity and protective efficacy. We find that compared to intranasal delivery, the deepened and widened biodistribution in the lung following endotracheal delivery is associated with much improved vaccine-mediated immunogenicity and protection against the target pathogen. Our findings thus support further development of inhaled aerosol delivery of vaccines over intranasal delivery for human application.

Respiratory mucosal delivery of next-generation COVID-19 vaccine provides robust protection against both ancestral and variant strains of SARS-CoV-2.

The emerging SARS-CoV-2 variants of concern (VOCs) threaten the effectiveness of current COVID-19 vaccines administered intramuscularly and designed to only target the spike protein. There is a pressing need to develop next-generation vaccine strategies for broader and long-lasting protection. Using adenoviral vectors (Ad) of human and chimpanzee origin, we evaluated Ad-vectored trivalent COVID-19 vaccines expressing spike-1, nucleocapsid, and RdRp antigens in murine models. We show that single-dose intranasal immunization, particularly with chimpanzee Ad-vectored vaccine, is superior to intramuscular immunization in induction of the tripartite protective immunity consisting of local and systemic antibody responses, mucosal tissue-resident memory T cells and mucosal trained innate immunity. We further show that intranasal immunization provides protection against both the ancestral SARS-CoV-2 and two VOC, B.1.1.7 and B.1.351. Our findings indicate that respiratory mucosal delivery of Ad-vectored multivalent vaccine represents an effective next-generation COVID-19 vaccine strategy to induce all-around mucosal immunity against current and future VOC.

Aerosol delivery, but not intramuscular injection, of adenovirus-vectored tuberculosis vaccine induces respiratory-mucosal immunity in humans.

BackgroundAdenovirus-vectored (Ad-vectored) vaccines are typically administered via i.m. injection to humans and are incapable of inducing respiratory mucosal immunity. However, aerosol delivery of Ad-vectored vaccines remains poorly characterized, and its ability to induce mucosal immunity in humans is unknown. This phase Ib trial evaluated the safety and immunogenicity of human serotype-5 Ad-vectored tuberculosis (TB) vaccine (AdHu5Ag85A) delivered to humans via inhaled aerosol or i.m. injection.MethodsThirty-one healthy, previously BCG-vaccinated adults were enrolled. AdHu5Ag85A was administered by single-dose aerosol using Aeroneb Solo Nebulizer or by i.m. injection. The study consisted of the low-dose (LD) aerosol, high-dose (HD) aerosol, and i.m. groups. The adverse events were assessed at various times after vaccination. Immunogenicity data were collected from the peripheral blood and bronchoalveolar lavage samples at baseline, as well as at select time points after vaccination.ResultsThe nebulized aerosol droplets were < 5.39 µm in size. Both LD and HD of AdHu5Ag85A administered by aerosol inhalation and i.m. injection were safe and well tolerated. Both aerosol doses, particularly LD, but not i.m., vaccination markedly induced airway tissue-resident memory CD4+ and CD8+ T cells of polyfunctionality. While as expected, i.m. vaccination induced Ag85A-specific T cell responses in the blood, the LD aerosol vaccination also elicited such T cells in the blood. Furthermore, the LD aerosol vaccination induced persisting transcriptional changes in alveolar macrophages.ConclusionInhaled aerosol delivery of Ad-vectored vaccine is a safe and superior way to elicit respiratory mucosal immunity. This study warrants further development of aerosol vaccine strategies against respiratory pathogens, including TB and COVID-19.Trial registrationClinicalTrial.gov, NCT02337270.FundingThe Canadian Institutes for Health Research (CIHR) and the Natural Sciences and Engineering Research Council of Canada funded this work.

Cigarette smoke augments CSF3 expression in neutrophils to compromise alveolar-capillary barrier function during influenza infection.

BACKGROUND: Cigarette smokers are at increased risk of acquiring influenza, developing severe disease and requiring hospitalisation/intensive care unit admission following infection. However, immune mechanisms underlying this predisposition are incompletely understood, and therapeutic strategies for influenza are limited. METHODS: We used a mouse model of concurrent cigarette smoke exposure and H1N1 influenza infection, colony-stimulating factor (CSF)3 supplementation/receptor (CSF3R) blockade and single-cell RNA sequencing (scRNAseq) to investigate this relationship. RESULTS: Cigarette smoke exposure exacerbated features of viral pneumonia such as oedema, hypoxaemia and pulmonary neutrophilia. Smoke-exposed infected mice demonstrated an increase in viral (v)RNA, but not replication-competent viral particles, relative to infection-only controls. Interstitial rather than airspace neutrophilia positively predicted morbidity in smoke-exposed infected mice. Screening of pulmonary cytokines using a novel dysregulation score identified an exacerbated expression of CSF3 and interleukin-6 in the context of smoke exposure and influenza. Recombinant (r)CSF3 supplementation during influenza aggravated morbidity, hypothermia and oedema, while anti-CSF3R treatment of smoke-exposed infected mice improved alveolar-capillary barrier function. scRNAseq delineated a shift in the distribution of Csf3 + cells towards neutrophils in the context of cigarette smoke and influenza. However, although smoke-exposed lungs were enriched for infected, highly activated neutrophils, gene signatures of these cells largely reflected an exacerbated form of typical influenza with select unique regulatory features. CONCLUSION: This work provides novel insight into the mechanisms by which cigarette smoke exacerbates influenza infection, unveiling potential therapeutic targets (e.g. excess vRNA accumulation, oedematous CSF3R signalling) for use in this context, and potential limitations for clinical rCSF3 therapy during viral infectious disease.

Experimental and natural evidence of SARS-CoV-2-infection-induced activation of type I interferon responses.

Type I interferons (IFNs) are our first line of defense against virus infection. Recent studies have suggested the ability of SARS-CoV-2 proteins to inhibit IFN responses. Emerging data also suggest that timing and extent of IFN production is associated with manifestation of COVID-19 severity. In spite of progress in understanding how SARS-CoV-2 activates antiviral responses, mechanistic studies into wild-type SARS-CoV-2-mediated induction and inhibition of human type I IFN responses are scarce. Here we demonstrate that SARS-CoV-2 infection induces a type I IFN response in vitro and in moderate cases of COVID-19. In vitro stimulation of type I IFN expression and signaling in human airway epithelial cells is associated with activation of canonical transcriptions factors, and SARS-CoV-2 is unable to inhibit exogenous induction of these responses. Furthermore, we show that physiological levels of IFNα detected in patients with moderate COVID-19 is sufficient to suppress SARS-CoV-2 replication in human airway cells.

COVID-19: Current knowledge in clinical features, immunological responses, and vaccine development.

The COVID-19 pandemic has unfolded to be the most challenging global health crisis in a century. In 11 months since its first emergence, according to WHO, the causative infectious agent SARS-CoV-2 has infected more than 100 million people and claimed more than 2.15 million lives worldwide. Moreover, the world has raced to understand the virus and natural immunity and to develop vaccines. Thus, within a short 11 months a number of highly promising COVID-19 vaccines were developed at an unprecedented speed and are now being deployed via emergency use authorization for immunization. Although a considerable number of review contributions are being published, all of them attempt to capture only a specific aspect of COVID-19 or its therapeutic approaches based on ever-expanding information. Here, we provide a comprehensive overview to conceptually thread together the latest information on global epidemiology and mitigation strategies, clinical features, viral pathogenesis and immune responses, and the current state of vaccine development.

Advancing Immunotherapeutic Vaccine Strategies Against Pulmonary Tuberculosis.

Chemotherapeutic intervention remains the primary strategy in treating and controlling tuberculosis (TB). However, a complex interplay between therapeutic and patient-related factors leads to poor treatment adherence. This in turn continues to give rise to unacceptably high rates of disease relapse and the growing emergence of drug-resistant forms of TB. As such, there is considerable interest in strategies that simultaneously improve treatment outcome and shorten chemotherapy duration. Therapeutic vaccines represent one such approach which aims to accomplish this through boosting and/or priming novel anti-TB immune responses to accelerate disease resolution, shorten treatment duration, and enhance treatment success rates. Numerous therapeutic vaccine candidates are currently undergoing pre-clinical and clinical assessment, showing varying degrees of efficacy. By dissecting the underlying mechanisms/correlates of their successes and/or shortcomings, strategies can be identified to improve existing and future vaccine candidates. This mini-review will discuss the current understanding of therapeutic TB vaccine candidates, and discuss major strategies that can be implemented in advancing their development.

Airway Macrophages Mediate Mucosal Vaccine-Induced Trained Innate Immunity against Mycobacterium tuberculosis in Early Stages of Infection.

Mycobacterium tuberculosis, the causative agent of pulmonary tuberculosis (TB), is responsible for millions of infections and deaths annually. Decades of TB vaccine development have focused on adaptive T cell immunity, whereas the importance of innate immune contributions toward vaccine efficacy has only recently been recognized. Airway macrophages (AwM) are the predominant host cell during early pulmonary M. tuberculosis infection and, therefore, represent attractive targets for vaccine-mediated immunity. We have demonstrated that respiratory mucosal immunization with a viral-vectored vaccine imprints AwM, conferring enhanced protection against heterologous bacterial challenge. However, it is unknown if innate immune memory also protects against M. tuberculosis In this study, by using a murine model, we detail whether respiratory mucosal TB vaccination profoundly alters the airway innate immune landscape associated with AwM prior to M. tuberculosis exposure and whether such AwM play a critical role in host defense against M. tuberculosis infection. Our study reveals an important role of AwM in innate immune protection in early stages of M. tuberculosis infection in the lung.

Immunological considerations for COVID-19 vaccine strategies.

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the most formidable challenge to humanity in a century. It is widely believed that prepandemic normalcy will never return until a safe and effective vaccine strategy becomes available and a global vaccination programme is implemented successfully. Here, we discuss the immunological principles that need to be taken into consideration in the development of COVID-19 vaccine strategies. On the basis of these principles, we examine the current COVID-19 vaccine candidates, their strengths and potential shortfalls, and make inferences about their chances of success. Finally, we discuss the scientific and practical challenges that will be faced in the process of developing a successful vaccine and the ways in which COVID-19 vaccine strategies may evolve over the next few years.