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Abstract Background Acute myeloid leukemia (AML) is characterized by the abnormal proliferation of myeloid precursor cells and presents significant challenges in treatment due to its heterogeneity. Recently, the NLRP3 inflammasome has emerged as a potential contributor to AML pathogenesis, although its precise mechanisms remain poorly understood. Methods Public genome datasets were utilized to evaluate the expression of NLRP3 inflammasome-related genes (IL-1β, IL-18, ASC, and NLRP3) in AML patients compared to healthy individuals. CRISPR/Cas9 technology was employed to generate NLRP3-deficient MOLM-13 AML cells, followed by comprehensive characterization using real-time PCR, western blotting, FACS analysis, and transmission electron and immunofluorescence microscopy. Proteomic analyses were conducted to identify NLRP3-dependent alterations in protein levels, with a focus on the eIF2 kinase PERK-mediated signaling pathways. Additionally, in vivo studies were performed using a leukemic mouse model to elucidate the pathogenic role of NLRP3 in AML. Results Elevated expression of NLRP3 was significantly associated with diminished overall survival in AML patients. Genetic deletion, pharmacological inhibition and silencing by RNA interference of NLRP3 led to decreased AML cell survival through the induction of apoptosis. Proteomic analyses uncovered NLRP3-dependent alterations in protein translation, characterized by enhanced eIF2α phosphorylation in NLRP3-deficient AML cells. Moreover, inhibition of PERK-mediated eIF2α phosphorylation reduced apoptosis by downregulating pro-apoptotic Bcl-2 family members. In vivo studies demonstrated reduced leukemic burden in mice engrafted with NLRP3 knockout AML cells, as evidenced by alleviated leukemic symptoms. Conclusion Our findings elucidate the involvement of the NLRP3/PERK/eIF2 axis as a novel driver of AML cell survival. Targeting NLRP3-induced signaling pathways, particularly through the PERK/eIF2 axis, presents a promising therapeutic strategy for AML intervention. These insights into the role of the NLRP3 inflammasome offer potential avenues for improving the prognosis and treatment outcomes of AML patients.

Sophisticated immune evasion strategies enable Helicobacter pylori (H. pylori) to colonize the gastric mucosa of approximately half of the world’s population. Persistent infection and the resulting chronic inflammation are a major cause of gastric cancer. To understand the intricate interplay between H. pylori and host immunity, spatial profiling was used to monitor immune cells in H. pylori infected gastric tissue. Dendritic cell (DC) and T cell phenotypes were further investigated in gastric organoid/immune cell co-cultures and mechanistic insights were acquired by proteomics of human DCs. Here, we show that ADP-heptose, a bacterial metabolite originally reported to act as a bona fide PAMP, reduces H. pylori-induced DC maturation and subsequent T cell responses. Mechanistically, we report that H. pylori uptake and subsequent DC activation by an ADP-heptose deficient H. pylori strain depends on TLR2. Moreover, ADP-heptose attenuates full-fledged activation of primary human DCs in the context of H. pylori infection by impairing type I IFN signaling. This study reveals that ADP-heptose mitigates host immunity during H. pylori infection.

Introduction & ObjectiveAllergic sensitization is an essential step in the development of allergic airway inflammation to birch pollen (BP); however, this process remains to be fully elucidated. Recent scientific advances have highlighted the importance of the allergen context. In this regard, microbial patterns (PAMPs) present on BP have attracted increasing interest. As these PAMPs are recognized by specialized pattern recognition receptors (PRRs), this study aims at investigating the roles of intracellular PRRs and the inflammasome regulator NLRP3.MethodsWe established a physiologically relevant intranasal and adjuvant-free sensitization procedure to study BP-induced systemic and local lung inflammation.ResultsStrikingly, BP-sensitized Nlrp3-deficient mice showed significantly lower IgE levels, Th2-associated cytokines, cell infiltration into the lung, mucin production and epithelial thickening than their wild-type counterparts, which appears to be independent of inflammasome formation. Intriguingly, bone-marrow chimera revealed that expression of NLRP3 in the hematopoietic system is required to trigger an allergic response.ConclusionOverall, this study identifies NLRP3 as an important driver of BP-induced allergic immune responses.

Helicobacter pylori is a widespread Gram-negative pathogen involved in a variety of gastrointestinal diseases, including gastritis, ulceration, mucosa-associated lymphoid tissue (MALT) lymphoma and gastric cancer. Immune responses aimed at eradication of H. pylori often prove futile, and paradoxically play a crucial role in the degeneration of epithelial integrity and disease progression. We have previously shown that H. pylori infection of primary human monocytes increases their potential to respond to subsequent bacterial stimuli – a process that may be involved in the generation of exaggerated, yet ineffective immune responses directed against the pathogen. In this study, we show that H. pylori-induced monocyte priming is not a common feature of Gram-negative bacteria, as Acinetobacter lwoffii induces tolerance to subsequent Escherichia coli lipopolysaccharide (LPS) challenge. Although the increased reactivity of H. pylori-infected monocytes seems to be specific to H. pylori, it appears to be independent of its virulence factors Cag pathogenicity island (CagPAI), cytotoxin associated gene A (CagA), vacuolating toxin A (VacA) and γ-glutamyl transferase (γ-GT). Utilizing whole-cell proteomics complemented with biochemical signaling studies, we show that H. pylori infection of monocytes induces a unique proteomic signature compared to other pro-inflammatory priming stimuli, namely LPS and the pathobiont A. lwoffii. Contrary to these tolerance-inducing stimuli, H. pylori priming leads to accumulation of NF-кB proteins, including p65/RelA, and thus to the acquisition of a monocyte phenotype more responsive to subsequent LPS challenge. The plasticity of pro-inflammatory responses based on abundance and availability of intracellular signaling molecules may be a heretofore underappreciated form of regulating innate immune memory as well as a novel facet of the pathobiology induced by H. pylori.

Infection with Helicobacter pylori (H. pylori) affects almost half of the world’s population and is a major cause of stomach cancer. Although immune cells react strongly to this gastric bacterium, H. pylori is still one of the rare pathogens that can evade elimination by the host and cause chronic inflammation. In the present study, we characterized the inflammatory response of primary human monocytes to repeated H. pylori infection and their responsiveness to an ensuing bacterial stimulus. We show that, although repeated stimulations with H. pylori do not result in an enhanced response, H. pylori-primed monocytes are hyper-responsive to an Escherichia coli-lipopolysaccharide (LPS) stimulation that takes place shortly after infection. This hyper-responsiveness to bacterial stimuli is observed upon infection with viable H. pylori only, while heat-killed H. pylori fails to boost both cytokine secretion and STAT activation in response to LPS. When the secondary challenge occurs several days after the primary infection with live bacteria, H. pylori-infected monocytes lose their hyper-responsiveness. The observation that H. pylori makes primary human monocytes more susceptible to subsequent/overlapping stimuli provides an important basis to better understand how H. pylori can maintain chronic inflammation and thus contribute to gastric cancer progression.

Silica nanoparticles (SiNPs) are generally regarded as safe and may represent an attractive carrier platform for nanomedical applications when loaded with biopharmaceuticals. Surface functionalization by different chemistries may help to optimize protein loading and may further impact uptake into the targeted tissues or cells, however, it may also alter the immunologic profile of the carrier system. In order to circumvent side effects, novel carrier candidates need to be tested thoroughly, early in their development stage within the pharmaceutical innovation pipeline, for their potential to activate or modify the immune response. Previous studies have identified surface functionalization by different chemistries as providing a plethora of modifications for optimizing efficacy of biopharmaceutical (nano)carrier platforms while maintaining an acceptable safety profile. In this study, we synthesized SiNPs and chemically functionalized them to obtain different surface characteristics to allow their application as a carrier system for allergen-specific immunotherapy. In the present study, crude natural allergen extracts are used in combination with alum instead of well-defined active pharmaceutical ingredients (APIs), such as recombinant allergen, loaded onto (nano)carrier systems with immunologically inert and stable properties in suspension. This study was motivated by the hypothesis that comparing different charge states could allow tailoring of the binding capacity of the particulate carrier system, and hence the optimization of biopharmaceutical uptake while maintaining an acceptable safety profile, which was investigated by determining the maturation of human antigen-presenting cells (APCs). The functionalized nanoparticles were characterized for primary and hydrodynamic size, polydispersity index, zeta potential, endotoxin contamination. As potential candidates for allergen-specific immunotherapy, the differently functionalized SiNPs were non-covalently coupled with a highly purified, endotoxin-free recombinant preparation of the major birch pollen allergen Bet v 1 that functioned for further immunological testing. Binding efficiencies of allergen to SiNPs was controlled to determine uptake of API. For efficacy and safety assessment, we employed human monocyte-derived dendritic cells as model for APCs to detect possible differences in the particles’ APC maturation potential. Functionalization of SiNP did not affect the viability of APCs, however, the amount of API physisorbed onto the nanocarrier system, which induced enhanced uptake, mainly by macropinocytosis. We found slight differences in the maturation state of APCs for the differently functionalized SiNP–API conjugates qualifying surface functionalization as an effective instrument for optimizing the immune response towards SiNPs. This study further suggests that surface-functionalized SiNPs could be a suitable, immunologically inert vehicle for the efficient delivery of biopharmaceutical products, as evidenced here for allergen-specific immunotherapy.

Introduction & Objective: Allergic sensitization is an essential step in the development of allergic airway inflammation to birch pollen (BP); however, this process remains to be fully elucidated. Recent scientific advances have highlighted the importance of the allergen context. In this regard, microbial patterns (PAMPs) present on BP have attracted increasing interest. As these PAMPs are recognized by specialized pattern recognition receptors (PRRs), this study aims at investigating the roles of intracellular PRRs and the inflammasome regulator NLRP3. Methods: We established a physiologically relevant intranasal and adjuvant-free sensitization procedure to study BP-induced systemic and local lung inflammation. Results: Strikingly, BP-sensitized Nlrp3-deficient mice showed significantly lower IgE levels, Th2-associated cytokines, cell infiltration into the lung, mucin production and epithelial thickening than their wild-type counterparts, which appears to be independent of inflammasome formation. Intriguingly, bonemarrow chimera revealed that expression of NLRP3 in the hematopoietic system is required to trigger an allergic response. Conclusion: Overall, this study identifies NLRP3 as an important driver of BP-induced allergic immune responses. [ABSTRACT FROM AUTHOR]

Helicobacter pylori is a widespread Gram-negative pathogen involved in a variety of gastrointestinal diseases, including gastritis, ulceration, mucosa-associated lymphoid tissue (MALT) lymphoma and gastric cancer. Immune responses aimed at eradication of H. pylori often prove futile, and paradoxically play a crucial role in the degeneration of epithelial integrity and disease progression. We have previously shown that H. pylori infection of primary human monocytes increases their potential to respond to subsequent bacterial stimuli - a process that may be involved in the generation of exaggerated, yet ineffective immune responses directed against the pathogen. In this study, we show that H. pylori-induced monocyte priming is not a common feature of Gram-negative bacteria, as Acinetobacter lwoffii induces tolerance to subsequent Escherichia coli lipopolysaccharide (LPS) challenge. Although the increased reactivity of H. pylori-infected monocytes seems to be specific to H. pylori, it appears to be independent of its virulence factors Cag pathogenicity island (CagPAI), cytotoxin associated gene A (CagA), vacuolating toxin A (VacA) and γ-glutamyl transferase (γ-GT). Utilizing whole-cell proteomics complemented with biochemical signaling studies, we show that H. pylori infection of monocytes induces a unique proteomic signature compared to other pro-inflammatory priming stimuli, namely LPS and the pathobiont A. lwoffii. Contrary to these toleranceinducing stimuli, H. pylori priming leads to accumulation of NF-κB proteins, including p65/RelA, and thus to the acquisition of a monocyte phenotype more responsive to subsequent LPS challenge. The plasticity of pro-inflammatory responses based on abundance and availability of intracellular signaling molecules may be a heretofore underappreciated form of regulating innate immune memory as well as a novel facet of the pathobiology induced by H. pylori. [ABSTRACT FROM AUTHOR]