is a leading cause of bacteremia, further complicated by the emergence of antibiotic-resistant strains such as methicillin-resistant
(MRSA). A better understanding of host defense mechanisms is ...needed for the development of host-directed therapies as an alternative approach to antibiotics. The levels of IL-1, IL-17, and TNF-α cytokines in circulation have been associated with predictive outcomes in patients with
bacteremia. However, their causative role in survival and the cell types involved in these responses during bacteremia is not entirely clear. Using a mouse model of
bacteremia, we demonstrated that IL-17A/F and TNF-α had no significant impact on survival, whereas IL-1R signaling was critical for survival during
bacteremia. Furthermore, we identified that T cells, but not neutrophils, monocytes/macrophages, or endothelial cells were the crucial cell type for IL-1R-mediated survival against
bacteremia. Finally, we determined that the expression of IL-1R on γδ T cell, but not CD4
or CD8
T cells was responsible for survival against the
bacteremia. Taken together, we uncovered a role for IL-1R, but not IL-17A/F and TNF-α in protection against
bacteremia. Importantly, γδ T cell-intrinsic expression of IL-1R was crucial for survival, but not on other immune cells or endothelial cells. These findings reveal potential cellular and immunological targets for host-directed therapies for improved outcomes against
bacteremia.
IgE induced by type 2 immune responses in atopic dermatitis is implicated in the progression of atopic dermatitis to other allergic diseases, including food allergies, allergic rhinitis, and asthma. ...However, the keratinocyte-derived signals that promote IgE and ensuing allergic diseases remain unclear. Herein, in a mouse model of atopic dermatitis-like skin inflammation induced by epicutaneous Staphylococcus aureus exposure, keratinocyte release of IL‑36α along with IL-4 triggered B cell IgE class-switching, plasma cell differentiation, and increased serum IgE levels-all of which were abrogated in IL-36R-deficient mice or anti-IL‑36R-blocking antibody-treated mice. Moreover, skin allergen sensitization during S. aureus epicutaneous exposure-induced IL-36 responses was required for the development of allergen-specific lung inflammation. In translating these findings, elevated IL‑36 cytokines in human atopic dermatitis skin and in IL‑36 receptor antagonist-deficiency patients coincided with increased serum IgE levels. Collectively, keratinocyte-initiated IL‑36 responses represent a key mechanism and potential therapeutic target against allergic diseases.
Mutations in the leucine-rich repeat kinase 2 gene (LRRK2) cause late-onset Parkinson's disease indistinguishable from idiopathic disease. The mechanisms whereby missense alterations in the LRRK2 ...gene initiate neurodegeneration remain unknown. Here, we demonstrate that seven of 10 suspected familial-linked mutations result in increased kinase activity. Functional and disease-associated mutations in conserved residues reveal the critical link between intrinsic guanosine triphosphatase (GTPase) activity and downstream kinase activity. LRRK2 kinase activity requires GTPase activity, whereas GTPase activity functions independently of kinase activity. Both LRRK2 kinase and GTPase activity are required for neurotoxicity and potentiate peroxide-induced cell death, although LRRK2 does not function as a canonical MAP-kinase-kinase-kinase. These results suggest a link between LRRK2 kinase activity and pathogenic mechanisms relating to neurodegeneration, further supporting a gain-of-function role for LRRK2 mutations.
The connection between aging‐related immune dysfunction and the lung manifestations of aging is poorly understood. A detailed characterization of the aging IL10‐deficient murine lung, a model of ...accelerated aging and frailty, reconciles features of both immunosenescence and lung aging in a coherent model. Airspace enlargement developed in the middle‐aged (12 months old) and aged (20–22 months old) IL10‐deficient lung punctuated by an expansion of macrophages and alveolar cell apoptosis. Compared to wild‐type (WT) controls, the IL10‐deficient lungs from young (4‐month‐old) mice showed increased oxidative stress which was enhanced in both genotypes by aging. Active caspase 3 staining was increased in the alveolar epithelial cells of aged WT and mutant lungs but was greater in the IL10‐deficient milieu. Lung macrophages were increased in the aged IL10‐deficient lungs with exuberant expression of MMP12. IL10 treatment of naïve and M2‐polarized bone marrow‐derived WT macrophages reduced MMP12 expression. Conditioned media studies demonstrated the secretome of aged mutant macrophages harbors reduced AECII prosurvival factors, specifically keratinocyte growth factor (KGF) and hepatocyte growth factor (HGF), promotes cell death, and reduces survival of primary alveolar epithelial cells. Compared to WT controls, aged IL10‐deficient mice have increased parenchymal lymphoid collections comprised of a reduced number of apoptotic cells and B cells. We establish that IL10 is a key modulator of airspace homeostasis and lymphoid morphogenesis in the aging lung enabling macrophage‐mediated alveolar epithelial cell survival and B‐cell survival within tertiary lymphoid structures.
The loss of IL10 causes airspace enlargement and expansion of lymphoid aggregates in the aged murine lung. Enhanced alveolar epithelial cell death from reduced macrophage‐secreted prosurvival factors and increased MMP12 production contributes to the airspace phenotype. The expansion of altered lymphoid aggregates in the aged mutant lung features reduced cell apoptosis and reduced B‐cell content. IL10 is a key modulator of airspace homeostasis and lymphoid morphogenesis in the lung.
Post-surgical implant-associated spinal infection is a devastating complication commonly caused by
Biofilm formation is thought to reduce penetration of antibiotics and immune cells, contributing to ...chronic and difficult-to-treat infections. A rabbit model of a posterior-approach spinal surgery was created, in which bilateral titanium pedicle screws were interconnected by a plate at the level of lumbar vertebra L6 and inoculated with a methicillin-resistant
(MRSA) bioluminescent strain.
whole-animal bioluminescence imaging (BLI) and
bacterial cultures demonstrated a peak in bacterial burden by day 14, when wound dehiscence occurred. Structures suggestive of biofilm, visualized by scanning electron microscopy, were evident up to 56 days following infection. Infection-induced inflammation and bone remodeling were also monitored using
F-fluorodeoxyglucose (
F-FDG) positron emission tomography (PET) and computed tomography (CT). PET imaging signals were noted in the soft tissue and bone surrounding the implanted materials. CT imaging demonstrated marked bone remodeling and a decrease in dense bone at the infection sites. This rabbit model of implant-associated spinal infection provides a valuable preclinical
approach to investigate the pathogenesis of implant-associated spinal infections and to evaluate novel therapeutics.
is a major human pathogen causing serious implant–associated infections. Combination treatment with rifampin (10 to 15 mg/kg per day), which has dose-dependent activity, is recommended to treat
...orthopedic implant–associated infections. Rifampin, however, has limited bone penetration. Here, dynamic
C-rifampin positron emission tomography (PET) performed in prospectively enrolled patients with confirmed
bone infection (
= 3) or without orthopedic infection (
= 12) demonstrated bone/plasma area under the concentration-time curve ratio of 0.14 (interquartile range, 0.09 to 0.19), exposures lower than previously thought. PET-based pharmacokinetic modeling predicted rifampin concentration-time profiles in bone and facilitated studies in a mouse model of
orthopedic implant infection. Administration of high-dose rifampin (human equipotent to 35 mg/kg per day) substantially increased bone concentrations (2 mg/liter versus <0.2 mg/liter with standard dosing) in mice and achieved higher bacterial killing and biofilm disruption. Treatment for 4 weeks with high-dose rifampin and vancomycin was noninferior to the recommended 6-week treatment of standard-dose rifampin with vancomycin in mice (risk difference, −6.7% favoring high-dose rifampin regimen). High-dose rifampin treatment ameliorated antimicrobial resistance (0% versus 38%;
= 0.04) and mitigated adverse bone remodeling (
< 0.01). Last, whole-genome sequencing demonstrated that administration of high-dose rifampin in mice reduced selection of bacterial mutations conferring rifampin resistance (
) and mutations in genes potentially linked to persistence. These data suggest that administration of high-dose rifampin is necessary to achieve optimal bone concentrations, which could shorten and improve treatments for
orthopedic implant infections.
Bacterial infection of spinal instrumentation is a significant challenge in spinal fusion surgery. Although the intraoperative local application of powdered vancomycin is common practice for ...mitigating infection, the antimicrobial effects of this route of administration are short-lived. Therefore, novel antibiotic-loaded bone grafts as well as a reliable animal model to permit the testing of such therapies are needed to improve the efficacy of infection reduction practices in spinal fusion surgery.
This study aims to establish a clinically relevant rat model of spinal implant-associated infection to permit the evaluation of antimicrobial bone graft materials used in spinal fusion.
Rodent study of chronic spinal implant-associated infection.
Instrumentation anchored in and spanning the vertebral bodies of L4 and L5 was inoculated with bioluminescent methicillin-resistant Staphylococcus aureus bacteria (MRSA). Infection was monitored using an in vivo imaging system (IVIS) for 8 weeks. Spines were harvested and evaluated histologically, and colony-forming units (CFUs) were quantified in harvested implants and spinal tissue.
Postsurgical analysis of bacterial infection in vivo demonstrated stratification between MRSA and phosphate-buffered saline (PBS) control groups during the first 4 weeks of the 8-week infection period, indicating the successful establishment of acute infection. Over the 8-week chronic infection period, groups inoculated with 1 × 105 MRSA CFU and 1 × 106 MRSA CFU demonstrated significantly higher bioluminescence than groups inoculated with PBS control (p = 0.009 and p = 0.041 respectively). Histological examination at 8 weeks postimplantation revealed the presence of abscesses localized to implant placement in all MRSA inoculation groups, with the most pervasive abscess formation in samples inoculated with 1 × 105 MRSA CFU and 1 × 106 MRSA CFU. Quantification of CFU plated from harvested spinal tissue at 8 weeks post-implantation revealed the 1 × 105 MRSA CFU inoculation group as the only group with a significantly greater average CFU count compared to PBS control (p = 0.017). Further, CFU quantification from harvested spinal tissue was greater than CFU quantification from harvested implants across all inoculation groups.
Our model demonstrated that the inoculation dosage of 1 × 105 MRSA CFU exhibited the most robust chronic infection within instrumented vertebral bodies. This dosage had the greatest difference in bioluminescence signal from control (p < 0.01), the lowest mortality (0% compared to 50% for samples inoculated with 1 × 106 MRSA CFU), and a significantly higher amount of CFUs from harvested spine samples than CFUs from control harvested spine samples. Further, histological analysis confirmed the reliability of this novel rodent model of implanted-associated infection to establish infection and biofilm formation of MRSA for all inoculation groups.
This model is intended to simulate the infection of instrumentation used in spinal fusion surgeries concerning implant locality and material. This model may evaluate potential antimicrobial and osteogenic biomaterials and investigate the relationship between implant-associated infection and failed fusion.
Orthopedic implant‐associated infection (OIAI) is a major complication that leads to implant failure. In preclinical models of Staphylococcus aureus OIAI, osteomyelitis and septic arthritis, ...interleukin‐1α (IL‐1α), IL‐1β, and tumor necrosis factor (TNF) are induced, but whether they have interactive or distinctive roles in host defense are unclear. Herein, a S. aureus OIAI model was performed in mice deficient in IL‐1α, IL‐1β, or TNF. Mice deficient in IL‐1β or TNF (to a lesser extent) but not IL‐1α had increased bacterial burden at the site of the OIAI throughout the 28‐day experiment. IL‐1β and TNF had a combined and critical role in host defense as mice deficient in both IL‐1R and TNF (IL‐1R/TNF‐deficient mice) had a 40% mortality rate, which was associated with markedly increased bacterial burden at the site of the OIAI infection. Finally, IL‐1α‐ and IL‐1β‐deficient mice had impaired neutrophil recruitment whereas IL‐1β‐, TNF‐, and IL‐1R/TNF‐deficient mice all had impaired recruitment of both neutrophils and monocytes. Therefore, IL‐1β and TNF contributed to host defense against S. aureus OIAI and neutrophil recruitment was primarily mediated by IL‐1β and monocyte recruitment was mediated by both IL‐1β and TNF.
Phosphodiesterase 4 (PDE4) is highly expressed in keratinocytes and immune cells and promotes pro‐inflammatory responses upon activation. The activity of PDE4 has been attributed to various ...inflammatory conditions, leading to the development and approval of PDE4 inhibitors as host‐directed therapeutics in humans. For example, the topical PDE4 inhibitor, crisaborole, is approved for the treatment of mild‐to‐moderate atopic dermatitis and has shown efficacy in patients with psoriasis. However, the role of crisaborole in regulating the immunopathogenesis of inflammatory skin diseases and infection is not entirely known. Therefore, we evaluated the effects of crisaborole in multiple mouse models, including psoriasis‐like dermatitis, AD‐like skin inflammation with and without filaggrin mutations, and Staphylococcus aureus skin infection. We discovered that crisaborole dampens myeloid cells and itch in the skin during psoriasis‐like dermatitis. Furthermore, crisaborole was effective in reducing skin inflammation in the context of filaggrin deficiency. Importantly, crisaborole reduced S. aureus skin colonization during AD‐like skin inflammation. However, crisaborole was not efficacious in treating S. aureus skin infections, even as adjunctive therapy to antibiotics. Taken together, we found that crisaborole reduced itch during psoriasis‐like dermatitis and decreased S. aureus skin colonization upon AD‐like skin inflammation, which act as additional mechanisms by which crisaborole dampens the immunopathogenesis in mouse models of inflammatory skin diseases. Further examination is warranted to translate these preclinical findings to human disease.
In vivo bioluminescence imaging has been used to monitor Staphylococcus aureus infections in preclinical models by employing bacterial reporter strains possessing a modified lux operon from ...Photorhabdus luminescens. However, the relatively short emission wavelength of lux (peak 490 nm) has limited tissue penetration. To overcome this limitation, the gene for the click beetle (Pyrophorus plagiophtalamus) red luciferase (luc) (with a longer >600 emission wavelength), was introduced singly and in combination with the lux operon into a methicillin-resistant S. aureus strain. After administration of the substrate D-luciferin, the luc bioluminescent signal was substantially greater than the lux signal in vitro. The luc signal had enhanced tissue penetration and improved anatomical co-registration with infected internal organs compared with the lux signal in a mouse model of S. aureus bacteremia with a sensitivity of approximately 3 × 10
CFU from the kidneys. Finally, in an in vivo mixed bacterial wound infection mouse model, S. aureus luc signals could be spectrally unmixed from Pseudomonas aeruginosa lux signals to noninvasively monitor the bacterial burden of both strains. Therefore, the S. aureus luc reporter may provide a technological advance for monitoring invasive organ dissemination during S. aureus bacteremia and for studying bacterial dynamics during mixed infections.
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