Sepsis is a life-threatening syndrome arising from a dysregulated host response to infection, characterized by a biphasic immune trajectory in which an early hyperinflammatory phase transitions into a prolonged state of immunosuppression. This late phase is dominated by massive apoptotic depletion of lymphocytes, functional exhaustion of monocytes and macrophages, and an inability to mount effective responses against secondary infections, collectively defining immune paralysis. Indeed, most sepsis-related deaths occur during this phase of immune paralysis. Throughout the sepsis response, various pro- and anti-inflammatory mediators are released, particularly cytokines. Among the factors released during sepsis, Lipocalin-2 (Lcn2), also known as neutrophil gelatinase-associated lipocalin (NGAL), is a secreted innate immune glycoprotein induced during infection as a bacteriostatic mediator that sequesters iron-bound bacterial siderophores, thereby depriving pathogens of a nutrient essential for growth and dissemination. While this early protective function of Lcn2 is well established, accumulating experimental and clinical evidence points to a second, temporally distinct role during the immunosuppressive phase. In murine models of polymicrobial sepsis, Lcn2 expression at both the mRNA and protein levels is uniquely sustained throughout the late phase, in sharp contrast to other inflammatory mediators that rapidly resolve. This persistent expression may drive iron-dependent metabolic stress, activation of intrinsic apoptotic pathways in lymphocytes, and stabilization of macrophage tolerance. In human patients, circulating NGAL correlates with inflammatory cytokines, endothelial activation, and disease severity, and persistently elevated levels associate with adverse outcomes. We propose that Lcn2 may act as a phase-dependent regulator of the septic immune response, serving a protective and antimicrobial role during early infection while potentially contributing to immune dysfunction when its expression is sustained. This dual behavior positions Lcn2 as both a clinically informative biomarker and a candidate target for therapeutic intervention in sepsis.
Background Acinetobacter baumannii pneumonia (ABP) presents a clinical challenge due to drug resistance and immune evasion. The specific mechanism by which the natural immunomodulator lentinan (LNT) exerts anti-infective effects remains unclear. This study aimed to investigate whether LNT functions through the PINK1/Parkin-mediated mitophagy pathway. Methods An immunosuppressed mouse model of ABP was established, and the mice were treated with LNT alone or in combination with the mitophagy inhibitor mdivi-1. Therapeutic efficacy was evaluated through survival analysis. The bacterial load in lung tissue was determined using the plate counting method, and lung histopathology was analyzed via H&E staining. Flow cytometry was used to analyze the recruitment of immune cells (neutrophils, macrophages, dendritic cells) in bronchoalveolar lavage fluid (BALF). Inflammatory cytokine (TNF-α, IL-6, IL-1β) levels and myeloperoxidase (MPO) activity were measured using assay kits. The expression of key mitophagy-related proteins (PINK1, Parkin, Beclin-1, LC3, p62) in lung tissue was assessed by Western blot. Results LNT treatment significantly improved the survival rate of ABP mice, reduced bacterial load in lung tissue, and alleviated inflammatory pathological damage. LNT promoted the recruitment of neutrophils, macrophages, and dendritic cells to infection sites, while downregulating the expression of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-1β. Mechanistic studies revealed that LNT upregulated the expression of PINK1, Parkin, and Beclin-1 proteins, increased the LC3-II/I ratio, and reduced p62 accumulation. These protective effects were reversed by mdivi-1 treatment. Conclusion LNT exerts a protective effect in ABP by activating the PINK1/Parkin-mediated mitophagy pathway, thereby promoting effective immune cell recruitment and regulating inflammatory responses. This study provides a new theoretical basis for the clinical treatment of drug-resistant bacterial infections.
Fibroblasts are considered structural cells that maintain tissue homeostasis and facilitate repair. They also participate in innate immunity, although their pattern recognition capabilities remain incompletely characterized. Here, we systematically assessed the innate immune responses of commercially available primary human dermal fibroblasts from a male and a female donor. Fibroblasts were stimulated with a panel of microbe-associated molecular patterns (MAMPs) targeting various pattern recognition receptors (PRRs), including Toll-like receptors (TLRs), NOD-like receptors (NODs), Alpha kinase 1 (ALPK1) and STING. We quantified innate immune activation by measuring the nuclear translocation of NF-κB via high-content microscopy, and cytokines and chemokines secretion by ELISA. Baseline PRRs expression was determined using quantitative PCR. Only a restricted subset of agonists induced robust NF-κB activation with 30-70% positive cells, specifically E. coli LPS (TLR4), Poly I:C (TLR3/RIG-I), and unexpectedly ADP heptose (ALPK1). Dose-response analysis revealed surprisingly high sensitivity to LPS, as low as 1 ng/mL. Apart from IL-6 and RANTES, which were produced exclusively following Poly I:C stimulation, the secretion of pro-inflammatory cytokines IL-1β, and TNF, as well as the anti-inflammatory cytokines IL-10 and IFN-β remained undetectable compared to that of the chemokines IL-8 and MCP-1. Consistent with this limited reactivity, qPCR of PRRs revealed the basal gene expression of TLR4 co-receptors MD2, CD14, and ALPK1, while most other receptors were expressed at very low or undetectable levels. Notably, NOD1 was highly expressed despite the absence of cell activation with several NOD1 agonists. In conclusion, primary human dermal fibroblasts exhibit a highly selective yet sensitive innate immune response, that is largely restricted to chemokine production. This unexpected dissociation between chemokine and cytokine responses suggests that fibroblasts function as sentinel cells in early skin defence, capable of detecting key microbial patterns at low concentrations, and orchestrating local immune surveillance. Further investigation into interindividual variability and context-dependent activation is required.
Influenza is a highly contagious virus with most individuals showing recovery within a week after symptom onset. However, children, the elderly, and those with chronic health conditions are at high risk of developing serious complications. Extensive immune cell infiltration of the lung and massive production of pro-inflammatory cytokines can result in lung tissue damage and disruption, resulting in hypoxia and death. During the 2024-2025 influenza season in the USA, influenza caused an estimated 27,000-130,000 deaths. Our previous studies have shown that agents known to drive polarization of macrophages into the M2a phenotype mitigated inflammatory responses to influenza H1N1 A/PR/8/34 (PR8). Since IRS2 dampens the IL-4-induced differentiation to the M2a subtype, we sought to determine the impact of IRS2 on PR8 infection. Contrary to expectations, IRS2 -/- mice exhibited enhanced susceptibility to PR8 when compared to IRS2 +/+ mice, which was associated with a significant drop in blood oxygen saturation. Additionally, PR8 infection induced significantly greater inflammation in the lungs of IRS2 -/- mice than IRS2 +/+ mice. Conversely, viral replication was not significantly different. Significantly increased levels of the inflammatory mediator HMGB1 and airway epithelial cell denuding were observed in the lungs of IRS2 -/- mice, with a concomitant decrease in M2a gene expression. Mice with myeloid-specific knockout of IRS2 showed no enhanced sensitivity to PR8, supporting the hypothesis that the major impact of IRS2-deficiency on host response to PR8 infection is not myeloid cell-intrinsic. Overall, these results suggest that in the absence of IRS2, airway epithelial cells are especially sensitive to influenza-induced damage through dysregulated inflammation, resulting in increased susceptibility during infection.
Background and Objectives Trained immunity (TI) refers to a non-specific, long-lasting protective immune response that occurs following initial stimulation of the immune system and thought to be largely mediated by functional reprogramming of myeloid cells. TI has been demonstrated in BCG-vaccinated infants and can be induced in human cells and adult mice via agonists of pattern recognition receptors (PRR), such as β-glucan or MDP (a muropeptide that activates NOD2). However, its induction in neonates remains poorly understood. Previously, we demonstrated that the synthetic TLR2-NOD2 dual agonist CL429 enhances antimicrobial functions in adult mice and protects against subsequent infection with Leptospira interrogans , a zoonotic pathogen. We also demonstrated the immediate protective benefits of NOD2 stimulation in neonates against Cryptosporidium, a zoonotic pathogen that affects young animals in livestock herds. However, whether NOD2 agonists can induce TI in neonates and protect them at adulthood is unknown. Methods and Results Here, we investigated whether exposure of neonatal mice to PRR agonists (CL429, MDP or β-glucan), administered intraperitoneally at one week interval at 7/14 or 14/21 days of age, could enhance inflammatory cytokines production after ex vivo restimulation and confer long-term protection into adulthood. Surprisingly, none of the treatments enhanced ex vivo cytokine responses in adulthood after restimulation, nor did they confer protection against experimental leptospirosis. Instead, MDP-treated neonates exhibited 50% mortality following adult infection, revealing an unexpected detrimental effect. Conclusion These findings demonstrate that these PRR agonists fail to induce protective TI against Leptospira when administered to neonatal mice, challenging assumptions derived from adult models. Furthermore, our study reveals the risks of administering immunostimulants during the early stages of life, and highlights unanticipated and potentially harmful, PRR- and age-specific mechanisms of immune system modulation.
