A Large Proportion of the Neonatal Iron Pool Is Acquired from the Gestational Diet in a Murine Model.

BACKGROUND: Iron is crucial for growth and development, but excess iron is harmful. Neonatal mice have elevated concentrations of circulating iron, but the source of this iron is unclear. This lack of understanding makes it difficult to optimize early life iron balance. OBJECTIVES: Identify the origins of neonatal tissue-specific iron pools using dietary manipulation and cross-fostering murine models. METHODS: To determine whether tissue-specific neonatal iron was primarily acquired during gestation or after birth, pups born to iron-sufficient or iron-deficient dams were cross-fostered, and tissues were harvested at postnatal days 3-5 to measure iron content. A separate set of female mice were fed a diet enriched with the stable iron isotope 57 (57Fe) for 4 generations to replace naturally abundant liver iron isotope 56 (56Fe) stores with 57Fe. To quantify the proportions of neonatal iron acquired during gestation, pups born to dams with 56Fe or 57Fe stores were cross-fostered, and tissues were harvested at postnatal day 3-5 to determine 56Fe:57Fe ratios by inductively coupled plasma mass spectrometry. Finally, to quantify the proportion of neonatal iron acquired from the maternal diet, female mice with 56Fe or 57Fe stores switched diets upon mating, and pup tissues were harvested on P0 to determine 56Fe:57Fe ratios by inductively coupled plasma mass spectrometry. RESULTS: Perinatal iron deficiency resulted in smaller pups, and gestational iron deficiency resulted in lower neonatal serum and liver iron. Cross-fostering between dams with 56Fe and 57Fe stores demonstrated that ≤70% of neonatal serum, liver, and brain iron were acquired during gestation. Dietary manipulation experiments using dams with 56Fe and 57Fe stores showed that over half of neonatal serum, liver, and brain iron were from the dam's gestational diet rather than preconception iron stores. CONCLUSIONS: This study provides quantitative values for the sources of neonatal iron, which may inform approaches to optimize neonatal iron status.

Neutrophil-Derived Tumor Necrosis Factor Drives Fungal Acute Lung Injury in Chronic Granulomatous Disease.

Chronic granulomatous disease (CGD) results from deficiency of nicotinamide adenine dinucleotide phosphate(NADPH) oxidase and impaired reactive oxygen species (ROS) generation. This leads to impaired killing of Aspergillus and, independently, a pathologic hyperinflammatory response to the organism. We hypothesized that neutrophil-derived ROS inhibit the inflammatory response to Aspergillus and that acute lung injury in CGD is due to failure of this regulation. Mice with gp91phox deficiency, the most common CGD mutation, had more severe lung injury, increased neutrophilinfiltration, and increased lung tumor necrosis factor (TNF) after Aspergillus challenge compared with wild-types. Neutrophils were surprisingly the predominant source of TNF in gp91phox-deficient lungs. TNF neutralization inhibited neutrophil recruitment in gp91phox-deficient mice and protected from lung injury. We propose that, in normal hosts, Aspergillus stimulates TNF-dependent neutrophil recruitment to the lungs and neutrophil-derived ROS limit inflammation. In CGD, in contrast, recruited neutrophils are the dominant source of TNF, promoting further neutrophil recruitment in a pathologic positive-feedback cycle, resulting in progressive lung injury.

The Role of Iron in the Susceptibility of Neonatal Mice to Escherichia coli K1 Sepsis.

Sepsis from Escherichia coli expressing the K1 antigen is a leading cause of death in neonates. In a murine model, E. coli K1 grew rapidly in the peritoneal cavity of neonatal mice, causing fatal disease. In contrast, adult mice cleared the infection. Neonatal mice mounted a rapid and equivalent antimicrobial immune response compared to adult mice. Interestingly, peritoneal fluid from neonatal mice contained significantly more total iron than that of adult mice, which was sufficient to support enhanced E. coli growth. Transient iron overload in adult mice infected with E. coli resulted in 100% mortality. Maternal diet-induced mild iron deficiency decreased offspring peritoneal iron, decreased bacterial growth, and conferred protection against sepsis. Taken together, neonatal susceptibility to E. coli K1 sepsis is enhanced by a localized excess of peritoneal iron that allows for unchecked bacterial growth. Targeting this excess iron may provide a new therapeutic target in human patients.

TLR Activation and Allergic Disease: Early Life Microbiome and Treatment.

PURPOSE OF REVIEW: Allergy and asthma are growing problems in the developed world. The accelerated increase of these diseases may be related to microbiome modification that leads to aberrant activation of Toll-like receptors (TLRs). Current research supports the concept that changes in microbial communities in early life impact TLR activation, resulting in an altered risk for the development of asthma and allergies. RECENT FINDINGS: Prenatal and early childhood events that generate microbiome modification are closely related with TLR activation. Early childhood exposure to a rich array of TLR agonists, particularly lipopolysaccharide, strongly predicts protection against allergic disease later in life even when other lifestyle factors are accounted for. Genetic deletion of TLR signaling components in mice results in reduced function of tolerogenic cell populations in the gut. In contrast, weak TLR signaling can promote allergic sensitization later in life. This review summarizes the role of TLR signaling in microbiome-mediated protection against allergy.

Preclinical proof of concept for VivoVec, a lentiviral-based platform for in vivo CAR T-cell engineering.

BACKGROUND: Chimeric antigen receptor (CAR) T-cell therapies have demonstrated transformational outcomes in the treatment of B-cell malignancies, but their widespread use is hindered by technical and logistical challenges associated with ex vivo cell manufacturing. To overcome these challenges, we developed VivoVec, a lentiviral vector-based platform for in vivo engineering of T cells. UB-VV100, a VivoVec clinical candidate for the treatment of B-cell malignancies, displays an anti-CD3 single-chain variable fragment (scFv) on the surface and delivers a genetic payload that encodes a second-generation CD19-targeted CAR along with a rapamycin-activated cytokine receptor (RACR) system designed to overcome the need for lymphodepleting chemotherapy in supporting successful CAR T-cell expansion and persistence. In the presence of exogenous rapamycin, non-transduced immune cells are suppressed, while the RACR system in transduced cells converts rapamycin binding to an interleukin (IL)-2/IL-15 signal to promote proliferation. METHODS: UB-VV100 was administered to peripheral blood mononuclear cells (PBMCs) from healthy donors and from patients with B-cell malignancy without additional stimulation. Cultures were assessed for CAR T-cell transduction and function. Biodistribution was evaluated in CD34-humanized mice and in canines. In vivo efficacy was evaluated against normal B cells in CD34-humanized mice and against systemic tumor xenografts in PBMC-humanized mice. RESULTS: In vitro, administration of UB-VV100 resulted in dose-dependent and anti-CD3 scFv-dependent T-cell activation and CAR T-cell transduction. The resulting CAR T cells exhibited selective expansion in rapamycin and antigen-dependent activity against malignant B-cell targets. In humanized mouse and canine studies, UB-VV100 demonstrated a favorable biodistribution profile, with transduction events limited to the immune compartment after intranodal or intraperitoneal administration. Administration of UB-VV100 to humanized mice engrafted with B-cell tumors resulted in CAR T-cell transduction, expansion, and elimination of systemic malignancy. CONCLUSIONS: These findings demonstrate that UB-VV100 generates functional CAR T cells in vivo, which could expand patient access to CAR T technology in both hematological and solid tumors without the need for ex vivo cell manufacturing.

Hepcidin-mediated iron sequestration protects against bacterial dissemination during pneumonia.

Gram-negative pneumonia is a dangerous illness, and bacterial dissemination to the bloodstream during the infection is strongly associated with death. Antibiotic resistance among the causative pathogens has resulted in diminishing treatment options against this infection. Hepcidin is the master regulator of extracellular iron availability in vertebrates, but its role in the context of host defense is undefined. We hypothesized that hepcidin-mediated depletion of extracellular iron during Gram-negative pneumonia protects the host by limiting dissemination of bacteria to the bloodstream. During experimental pneumonia, hepcidin was induced in the liver in an IL-6-dependent manner and mediated a rapid decline in plasma iron. In contrast, hepcidin-deficient mice developed a paradoxical increase in plasma iron during infection associated with profound susceptibility to bacteremia. Incubation of bacteria with iron-supplemented plasma enhanced bacterial growth in vitro, and systemic administration of iron to WT mice similarly promoted increased susceptibility to bloodstream infection. Finally, treatment with a hepcidin analogue restored hypoferremia in hepcidin-deficient hosts, mediated bacterial control, and improved outcomes. These data show hepcidin induction during pneumonia to be essential to preventing bacterial dissemination by limiting extracellular iron availability. Hepcidin agonists may represent an effective therapy for Gram-negative infections in patients with impaired hepcidin production or signaling.

A computational model of invasive aspergillosis in the lung and the role of iron.

BACKGROUND: Invasive aspergillosis is a severe infection of immunocompromised hosts, caused by the inhalation of the spores of the ubiquitous environmental molds of the Aspergillus genus. The innate immune response in this infection entails a series of complex and inter-related interactions between multiple recruited and resident cell populations with each other and with the fungal cell; in particular, iron is critical for fungal growth. RESULTS: A computational model of invasive aspergillosis is presented here; the model can be used as a rational hypothesis-generating tool to investigate host responses to this infection. Using a combination of laboratory data and published literature, an in silico model of a section of lung tissue was generated that includes an alveolar duct, adjacent capillaries, and surrounding lung parenchyma. The three-dimensional agent-based model integrates temporal events in fungal cells, epithelial cells, monocytes, and neutrophils after inhalation of spores with cellular dynamics at the tissue level, comprising part of the innate immune response. Iron levels in the blood and tissue play a key role in the fungus' ability to grow, and the model includes iron recruitment and consumption by the different types of cells included. Parameter sensitivity analysis suggests the model is robust with respect to unvalidated parameters, and thus is a viable tool for an in silico investigation of invasive aspergillosis. CONCLUSIONS: Using laboratory data from a mouse model of invasive aspergillosis in the context of transient neutropenia as validation, the model predicted qualitatively similar time course changes in fungal burden, monocyte and neutrophil populations, and tissue iron levels. This model lays the groundwork for a multi-scale dynamic mathematical model of the immune response to Aspergillus species.