Dynamic equilibrium of skeletal muscle macrophage ontogeny in the diaphragm during homeostasis, injury, and recovery.

The diaphragm is a unique skeletal muscle due to its continuous activation pattern during the act of breathing. The ontogeny of macrophages, pivotal cells for skeletal muscle maintenance and regeneration, is primarily based on two distinct origins: postnatal bone marrow-derived monocytes and prenatal embryonic progenitors. Here we employed chimeric mice to study the dynamics of these two macrophage populations under different conditions. Traditional chimeric mice generated through whole body irradiation showed virtually complete elimination of the original tissue-resident macrophage pool. We then developed a novel method which employs lead shielding to protect the diaphragm tissue niche from irradiation. This allowed us to determine that up to almost half of tissue-resident macrophages in the diaphragm can be maintained independently from bone marrow-derived monocytes under steady-state conditions. These findings were confirmed by long-term (5 months) parabiosis experiments. Acute diaphragm injury shifted the macrophage balance toward an overwhelming predominance of bone marrow (monocyte)-derived macrophages. However, there was a remarkable reversion to the pre-injury ontological landscape after diaphragm muscle recovery. This diaphragm shielding method permits analysis of the dynamics of macrophage origin and corresponding function under different physiological and pathological conditions. It may be especially useful for studying diseases which are characterized by acute or chronic injury of the diaphragm and accompanying inflammation.

Virus-Induced Acute Respiratory Distress Syndrome Causes Cardiomyopathy Through Eliciting Inflammatory Responses in the Heart.

BACKGROUND: Viral infections can cause acute respiratory distress syndrome (ARDS), systemic inflammation, and secondary cardiovascular complications. Lung macrophage subsets change during ARDS, but the role of heart macrophages in cardiac injury during viral ARDS remains unknown. Here we investigate how immune signals typical for viral ARDS affect cardiac macrophage subsets, cardiovascular health, and systemic inflammation. METHODS: We assessed cardiac macrophage subsets using immunofluorescence histology of autopsy specimens from 21 patients with COVID-19 with SARS-CoV-2-associated ARDS and 33 patients who died from other causes. In mice, we compared cardiac immune cell dynamics after SARS-CoV-2 infection with ARDS induced by intratracheal instillation of Toll-like receptor ligands and an ACE2 (angiotensin-converting enzyme 2) inhibitor. RESULTS: In humans, SARS-CoV-2 increased total cardiac macrophage counts and led to a higher proportion of CCR2+ (C-C chemokine receptor type 2 positive) macrophages. In mice, SARS-CoV-2 and virus-free lung injury triggered profound remodeling of cardiac resident macrophages, recapitulating the clinical expansion of CCR2+ macrophages. Treating mice exposed to virus-like ARDS with a tumor necrosis factor α-neutralizing antibody reduced cardiac monocytes and inflammatory MHCIIlo CCR2+ macrophages while also preserving cardiac function. Virus-like ARDS elevated mortality in mice with pre-existing heart failure. CONCLUSIONS: Our data suggest that viral ARDS promotes cardiac inflammation by expanding the CCR2+ macrophage subset, and the associated cardiac phenotypes in mice can be elicited by activating the host immune system even without viral presence in the heart.

Navigating the blurred path of mixed neuroimmune signaling.

Evolution has created complex mechanisms to sense environmental danger and protect tissues, with the nervous and immune systems playing pivotal roles. These systems work together, coordinating local and systemic reflexes to restore homeostasis in response to tissue injury and infection. By sharing receptors and ligands, they influence the pathogenesis of various diseases. Recently, a less-explored aspect of neuroimmune communication has emerged: the release of neuropeptides from immune cells and cytokines/chemokines from sensory neurons. This article reviews evidence of this unique neuroimmune interplay and its impact on the development of allergy, inflammation, itch, and pain. We highlight the effects of this neuroimmune signaling on vital processes such as host defense, tissue repair, and inflammation resolution, providing avenues for exploration of the underlying mechanisms and therapeutic potential of this signaling.

BCG immunization induces CX3CR1hi effector memory T cells to provide cross-protection via IFN-γ-mediated trained immunity.

After a century of using the Bacillus Calmette-Guérin (BCG) vaccine, our understanding of its ability to provide protection against homologous (Mycobacterium tuberculosis) or heterologous (for example, influenza virus) infections remains limited. Here we show that systemic (intravenous) BCG vaccination provides significant protection against subsequent influenza A virus infection in mice. We further demonstrate that the BCG-mediated cross-protection against influenza A virus is largely due to the enrichment of conventional CD4+ effector CX3CR1hi memory αß T cells in the circulation and lung parenchyma. Importantly, pulmonary CX3CR1hi T cells limit early viral infection in an antigen-independent manner via potent interferon-γ production, which subsequently enhances long-term antimicrobial activity of alveolar macrophages. These results offer insight into the unknown mechanism by which BCG has persistently displayed broad protection against non-tuberculosis infections via cross-talk between adaptive and innate memory responses.

Lung Epithelial Cells from Obese Patients Have Impaired Control of SARS-CoV-2 Infection.

Obesity is known to increase the complications of the COVID-19 coronavirus disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). However, the exact mechanisms of SARS-CoV-2 infection in obese patients have not been clearly elucidated. This study aims to better understand the effect of obesity on the course of SARS-CoV-2 infection and identify candidate molecular pathways involved in the progression of the disease, using an in vitro live infection model and RNA sequencing. Results from this study revealed the enhancement of viral load and replication in bronchial epithelial cells (NHBE) from obese subjects at 24 h of infection (MOI = 0.5) as compared to non-obese subjects. Transcriptomic profiling via RNA-Seq highlighted the enrichment of lipid metabolism-related pathways along with LPIN2, an inflammasome regulator, as a unique differentially expressed gene (DEG) in infected bronchial epithelial cells from obese subjects. Such findings correlated with altered cytokine and angiotensin-converting enzyme-2 (ACE2) expression during infection of bronchial cells. These findings provide a novel insight on the molecular interplay between obesity and SARS-CoV-2 infection. In conclusion, this study demonstrates the increased SARS-CoV-2 infection of bronchial epithelial cells from obese subjects and highlights the impaired immunity which may explain the increased severity among obese COVID-19 patients.

BCG vaccination provides protection against IAV but not SARS-CoV-2.

Since the vast majority of species solely rely on innate immunity for host defense, it stands to reason that a critical evolutionary trait like immunological memory evolved in this primitive branch of our immune system. There is ample evidence that vaccines such as bacillus Calmette-Guérin (BCG) induce protective innate immune memory responses (trained immunity) against heterologous pathogens. Here we show that while BCG vaccination significantly reduces morbidity and mortality against influenza A virus (IAV), it fails to provide protection against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). In contrast to IAV, SARS-CoV-2 infection leads to unique pulmonary vasculature damage facilitating viral dissemination to other organs, including the bone marrow (BM), a central site for BCG-mediated trained immunity. Finally, monocytes from BCG-vaccinated individuals mount an efficient cytokine response to IAV infection, while this response is minimal following SARS-CoV-2. Collectively, our data suggest that the protective capacity of BCG vaccination is contingent on viral pathogenesis and tissue tropism.

TLR4 is a regulator of trained immunity in a murine model of Duchenne muscular dystrophy.

Dysregulation of the balance between pro-inflammatory and anti-inflammatory macrophages has a key function in the pathogenesis of Duchenne muscular dystrophy (DMD), a fatal genetic disease. We postulate that an evolutionarily ancient protective mechanism against infection, known as trained immunity, drives pathological inflammation in DMD. Here we show that bone marrow-derived macrophages from a murine model of DMD (mdx) exhibit cardinal features of trained immunity, consisting of transcriptional hyperresponsiveness associated with metabolic and epigenetic remodeling. The hyperresponsive phenotype is transmissible by bone marrow transplantation to previously healthy mice and persists for up to 11 weeks post-transplant. Mechanistically, training is induced by muscle extract in vitro. The functional and epigenetic changes in bone marrow-derived macrophages from dystrophic mice are TLR4-dependent. Adoptive transfer experiments further support the TLR4-dependence of trained macrophages homing to damaged muscles from the bone marrow. Collectively, this suggests that a TLR4-regulated, memory-like capacity of innate immunity induced at the level of the bone marrow promotes dysregulated inflammation in DMD.

Human alveolar macrophage metabolism is compromised during Mycobacterium tuberculosis infection.

Pulmonary macrophages have two distinct ontogenies: long-lived embryonically-seeded alveolar macrophages (AM) and bone marrow-derived macrophages (BMDM). Here, we show that after infection with a virulent strain of Mycobacterium tuberculosis (H37Rv), primary murine AM exhibit a unique transcriptomic signature characterized by metabolic reprogramming distinct from conventional BMDM. In contrast to BMDM, AM failed to shift from oxidative phosphorylation (OXPHOS) to glycolysis and consequently were unable to control infection with an avirulent strain (H37Ra). Importantly, healthy human AM infected with H37Ra equally demonstrated diminished energetics, recapitulating our observation in the murine model system. However, the results from seahorse showed that the shift towards glycolysis in both AM and BMDM was inhibited by H37Rv. We further demonstrated that pharmacological (e.g. metformin or the iron chelator desferrioxamine) reprogramming of AM towards glycolysis reduced necrosis and enhanced AM capacity to control H37Rv growth. Together, our results indicate that the unique bioenergetics of AM renders these cells a perfect target for Mtb survival and that metabolic reprogramming may be a viable host targeted therapy against TB.

M. tuberculosis Reprograms Hematopoietic Stem Cells to Limit Myelopoiesis and Impair Trained Immunity.

A greater understanding of hematopoietic stem cell (HSC) regulation is required for dissecting protective versus detrimental immunity to pathogens that cause chronic infections such as Mycobacterium tuberculosis (Mtb). We have shown that systemic administration of Bacille Calmette-Guérin (BCG) or ß-glucan reprograms HSCs in the bone marrow (BM) via a type II interferon (IFN-II) or interleukin-1 (IL1) response, respectively, which confers protective trained immunity against Mtb. Here, we demonstrate that, unlike BCG or ß-glucan, Mtb reprograms HSCs via an IFN-I response that suppresses myelopoiesis and impairs development of protective trained immunity to Mtb. Mechanistically, IFN-I signaling dysregulates iron metabolism, depolarizes mitochondrial membrane potential, and induces cell death specifically in myeloid progenitors. Additionally, activation of the IFN-I/iron axis in HSCs impairs trained immunity to Mtb infection. These results identify an unanticipated immune evasion strategy of Mtb in the BM that controls the magnitude and intrinsic anti-microbial capacity of innate immunity to infection.

ß-Glucan Induces Protective Trained Immunity against Mycobacterium tuberculosis Infection: A Key Role for IL-1.

ß-glucan is a potent inducer of epigenetic and functional reprogramming of innate immune cells, a process called "trained immunity," resulting in an enhanced host response against secondary infections. We investigate whether ß-glucan exposure confers protection against pulmonary Mycobacterium tuberculosis (Mtb) infection. ß-glucan induces trained immunity via histone modifications at gene promoters in human monocytes, which is accompanied by the enhanced production of proinflammatory cytokines upon secondary Mtb challenge and inhibition of Mtb growth. Mice treated with ß-glucan are significantly protected against pulmonary Mtb infection, which is associated with the expansion of hematopoietic stem and progenitor cells in the bone marrow and increased myelopoiesis. The protective signature of ß-glucan is mediated via IL-1 signaling, as ß-glucan shows no protection in mice lacking a functional IL-1 receptor (IL1R-/-). The administration of ß-glucan may be used as a novel strategy in the treatment of mycobacterial infections and possibly as an adjuvant to improve anti-tuberculosis vaccines.

NK cell recruitment limits tissue damage during an enteric helminth infection.

Parasitic helminths cause significant damage as they migrate through host tissues to complete their life cycle. While chronic helminth infections are characterized by a well-described Type 2 immune response, the early, tissue-invasive stages are not well understood. Here we investigate the immune pathways activated during the early stages of Heligmosomoides polygyrus bakeri (Hpb), a natural parasitic roundworm of mice. In contrast to the Type 2 immune response present at later stages of infection, a robust Type 1 immune signature including IFNg production was dominant at the time of parasite invasion and granuloma formation. This early response was associated with an accumulation of activated Natural Killer (NK) cells, with no increase of other innate lymphoid cell populations. Parabiosis and confocal microscopy studies indicated that NK cells were recruited from circulation to the small intestine, where they surrounded parasitic larvae. NK cell recruitment required IFNγ receptor signaling, but was independent of CXCR3 expression. The depletion of tissue-infiltrating NK cells altered neither worm burden nor parasite fitness, but increased vascular injury, suggesting a role for NK cells in mediating tissue protection. Together, these data identify an unexpected role for NK cells in promoting disease tolerance during the invasive stage of an enteric helminth infection.

BCG Educates Hematopoietic Stem Cells to Generate Protective Innate Immunity against Tuberculosis.

The dogma that adaptive immunity is the only arm of the immune response with memory capacity has been recently challenged by several studies demonstrating evidence for memory-like innate immune training. However, the underlying mechanisms and location for generating such innate memory responses in vivo remain unknown. Here, we show that access of Bacillus Calmette-Guérin (BCG) to the bone marrow (BM) changes the transcriptional landscape of hematopoietic stem cells (HSCs) and multipotent progenitors (MPPs), leading to local cell expansion and enhanced myelopoiesis at the expense of lymphopoiesis. Importantly, BCG-educated HSCs generate epigenetically modified macrophages that provide significantly better protection against virulent M. tuberculosis infection than naïve macrophages. By using parabiotic and chimeric mice, as well as adoptive transfer approaches, we demonstrate that training of the monocyte/macrophage lineage via BCG-induced HSC reprogramming is sustainable in vivo. Our results indicate that targeting the HSC compartment provides a novel approach for vaccine development.

Beyond Killing Mycobacterium tuberculosis: Disease Tolerance.

Host defense strategies against infectious diseases are comprised of both host resistance and disease tolerance. Resistance is the ability of the host to prevent invasion or to eliminate the pathogen, while disease tolerance is defined by limiting the collateral tissue damage caused by the pathogen and/or the immune response without exerting direct effects on pathogen growth. Our incomplete understanding of host immunity against tuberculosis (TB) is predominately rooted in our bias toward investigating host resistance. Thus, we must refocus our efforts to understand the entire spectrum of immunity against M. tuberculosis to control TB.

Unravelling the networks dictating host resistance versus tolerance during pulmonary infections.

The appearance of single cell microorganisms on earth dates back to more than 3.5 billion years ago, ultimately leading to the development of multicellular organisms approximately 3 billion years later. The evolutionary burst of species diversity and the "struggle for existence", as proposed by Darwin, generated a complex host defense system. Host survival during infection in vital organs, such as the lung, requires a delicate balance between host defense, which is essential for the detection and elimination of pathogens and host tolerance, which is critical for minimizing collateral tissue damage. Whereas the cellular and molecular mechanisms of host defense against many invading pathogens have been extensively studied, our understanding of host tolerance as a key mechanism in maintaining host fitness is extremely limited. This may also explain why current therapeutic and preventive approaches targeting only host defense mechanisms have failed to provide full protection against severe infectious diseases, including pulmonary influenza virus and Mycobacterium tuberculosis infections. In this review, we aim to outline various host strategies of resistance and tolerance for effective protection against acute or chronic pulmonary infections.