We previously showed correction of sickle cell anemia (SCA) in mice utilizing a lentiviral vector (LV) expressing human γ-globin. Herein, we made a G16D mutation in the γ-globin gene to generate the G16D mutation (GbGM) LV to increase fetal hemoglobin formation. We also generated an insulated version of this LV, GbGMI, inserting a 36-bp insulator from the Foamy virus in the long terminal repeats of the LV. Preclinical batches of GbGM and GbGMI LV showed both were highly efficacious in correcting SCA in mice, with sustained gene transfer in primary transplanted SCA mice and high hematopoietic stem cell (HSC) transduction in colony-forming unit-spleen in secondary transplanted mice. CRISPR-mediated targeting of the proviruses into the LMO2 proto-oncogene showed remarkably reduced LMO2 activation by both insulated and uninsulated LV, compared to the SFFV γ-RV vector targeted to the same locus. We therefore used the GbGM LV to perform preclinical human CD34+ gene transfer. We assessed gene transfer and engraftment of human HSCs in two immunocompromised mouse models: persistent stable GbGM-transduced cell engraftment was comparable to that of untransduced cells with no detrimental effects on hematopoiesis up to 20 weeks post transplant. These robust preclinical studies in mouse and human HSCs allowed its translation into a clinical trial.
© 2025 The Authors.

Dendritic cells (DCs) play a crucial role in orchestrating immune responses by bridging innate and adaptive immunity. In vitro generation of DCs from mouse and human tissues such as bone marrow and peripheral blood monocytes, has been widely used to study their immunological functions. In chicken, DCs have mainly been derived from bone marrow cell cultures, with limited characterization from blood monocytes.
The present study takes advantage of newly available chicken immunological tools to further characterize chicken monocyte-derived dendritic cells (MoDCs), focusing on their phenotype, and functions, including antigen capture and T-cell stimulation, and response to live Newcastle disease virus (NDV) stimulation.
Adherent chicken PBMCs were cultured with recombinant chicken granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-4 (IL-4), for 5 days, leading to the upregulation of putative CD11c and MHCII, markers of DC differentiation. Subsequent stimulation with lipopolysaccharide (LPS) or 24 h triggered phenotypic maturation of MoDCs, characterized by the increased surface expression of MHCII and co-stimulatory molecules CD80 and CD40, and elevated IL-12p40 secretion. This maturation reduced endocytic capacity but enhanced the allogenic stimulatory activity of the chicken MoDCs. Upon NDV stimulation for 6 h, MoDCs upregulated antiviral pathways, including retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs), melanoma differentiation-associated protein 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2), alongside increased production of type I interferons (IFNs), and the pro-inflammatory cytokines tumor necrosis factor-α (TNF-α), IL-1β, and IL-6. However, these responses were downregulated after 24 hours.
These findings provide a comprehensive characterization of chicken MoDCs and suggest their potential as a model for studying host-pathogen interactions.
Copyright © 2025 Ngantcha Tatchou, Milcamps, Oldenhove, Lambrecht and Ingrao.

Immune responses in the intestine are intricately balanced to prevent pathogen entry without inducing immunopathology. The nervous system is well established to interface with the immune system to fine-tune immunity in various organ systems including the gastrointestinal tract. Specialized sensory neurons can detect bacteria, bacterial products, and the resulting inflammation, to coordinate the immune response in the gastrointestinal tract. These sensory neurons release peptide neurotransmitters such as Substance P (SP), to induce both neuronal signaling and localized responses in nonneuronal cells. With this in mind, we assessed the immunoregulatory roles of SP receptor signaling during enteric bacterial infection with the noninvasive pathogen Citrobacter rodentium. Pharmacological antagonism of the SP receptor significantly reduced bacterial burden and prevented colonic crypt hyperplasia. Mice with SP receptor signaling blockade had significantly reduced inflammation and recruitment of T cells in the colon. Reduced colonic T cell recruitment is due to reduced expression of adhesion molecules on colonic endothelial cells in SP receptor antagonist-treated mice. Using SP receptor T cell conditional knockout mice, we further confirmed SP receptor signaling enhanced select aspects of T cell responses. Our data demonstrate that SP receptor signaling can significantly reduce inflammation and prevent host-maladaptive responses without impinging upon host protection.

Vagus nerve stimulation (VNS) has been shown to limit immune cell activity across several pathologies ranging from sepsis to auto-immune diseases. While stimulation of vagal efferent neurons has been previously demonstrated to reduce maladaptive host responses during endotoxemia, only selective stimulation of vagal afferent neurons was able to inhibit TLR7-induced macrophage activation and neutrophil recruitment in the lung. These anti-inflammatory actions are facilitated by systemic increases in epinephrine, as VNS significantly increased epinephrine in the serum and bronchoalveolar lavage fluid, and inhibition of epinephrine production eliminated the protection afforded by VNS. Selective afferent VNS induced activation in the nucleus tractus solitarius and the rostral ventrolateral medulla. Inhibition of neuronal activity in this brain region that controls peripheral sympathetic nervous system activity rendered VNS ineffective. Activation of the β 2 -adrenergic receptor (β 2 AR) is critical for innate immune cell suppression, as the anti-inflammatory effects of VNS were eliminated in β 2 AR-knock out mice, and with pharmacological inhibition of the β 2 AR. Analysis of the immune cells responding to R848 critically identified that plasmacytoid dendritic cells were refractive to inhibition by VNS, and this corresponded to lack of β 2 AR expression. These findings demonstrate a novel neuro-immune circuit elicited by VNS that can control acute lung inflammation. Summary We have identified a novel neuro-immune circuit activated by afferent vagus nerve stimulation to reduce acute lung inflammation. This effect was dependent on vagal-induced adrenal gland-derived epinephrine release that initiates anti-inflammatory β2-adrenergic receptor signaling in innate immune cells within the lung.

In acute myeloid leukemia (AML), malignant cells surviving chemotherapy rely on high mRNA translation and their microenvironmental metabolic support to drive relapse. However, the role of translational reprogramming in the niche is unclear. Here, we found that relapsing AML cells increase translation in their bone marrow (BM) niches, where BM mesenchymal stromal cells (BMSCs) become a source of eIF4A-cap-dependent translation machinery that is transferred to AML cells via extracellular vesicles (EVs) to meet their translational demands. In two independent models of highly chemo-resistant AML driven by MLL-AF9 or FLT3-ITD (internal tandem duplication) and nucleophosmin (NPMc) mutations, protein synthesis levels increase in refractory AML dependent on nestin+ BMSCs. Inhibiting cap-dependent translation in BMSCs abolishes their chemoprotective ability, while EVs from BMSCs carrying eIF4A boost AML cell translation and survival. Consequently, eIF4A inhibition synergizes with conventional chemotherapy. Together, these results suggest that AML cells rely on BMSCs to maintain an oncogenic translational program required for relapse.
Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.