HIV-Tat and morphine synergistically exacerbate neuroinflammation and DNA damage, yet the mechanisms linking the DNA damage response (DDR), chromatin remodeling, and innate immune activation remain poorly understood. Here, we investigated how HIV-Tat and morphine alter DNA repair dynamics and cGAS-STING-AIM2 inflammasome signaling in microglia and cortical tissue and evaluated the efficacy of dimethyl fumarate (DMF) in mitigating these effects. HMC3 microglial cells were exposed to HIV-Tat (50 ng/mL) and morphine (10 µM) for 24 h or 5 days, with H2A.X knockdown performed prior to treatment. In vivo, doxycycline (DOX)-induced Tat transgenic mice received escalating morphine doses for 4 days, followed by DMF (30 mg/kg, i.p.) for 7 days. Postmortem frontal cortical tissues from HIV-positive and matched HIV-negative donors were analyzed for validation. Behavioral testing, immunostaining, qPCR, and Western blotting assessed DDR markers, KAT5 acetylation, histone modifications, and innate immune mediators. HIV-Tat and morphine significantly enhanced DDR activation in vitro and in vivo, as indicated by increased ATM and γH2AX and elevated KAT5 acetylation. Large Organized Chromatin Lysine (K) domains (LOCK) displayed reduced H3K9me3 and increased H3K9ac, reflecting chromatin relaxation and disrupted epigenetic control over DDR and inflammation. DOX+Morphine treatment upregulated ERCC1 and ERCC8 but downregulated ERCC2, ERCC3, and ERCC6, demonstrating impaired nucleotide excision repair, and these changes were closely associated with cGAS-STING activation and AIM2/NLRP3 inflammasome induction. H2A.X knockdown or DMF treatment suppressed ATM phosphorylation, restored H3K9me3, normalized ERCC expression, and attenuated inflammasome activation. DMF also rescued HIV-Tat ± morphine-induced deficits in novel object recognition. Postmortem cortical tissue confirmed γH2AX accumulation, reduced ERCC2/ERCC3, and altered histone signatures consistent with experimental findings. Together, these results demonstrate that HIV-Tat and morphine cooperatively disrupt LOCK chromatin and DDR through the ATM/γH2AX axis, and that DMF reverses these molecular and functional abnormalities, highlighting its therapeutic potential in mitigating HIV- and opioid-induced neuroinflammation and cognitive decline.

This study examined anti-transferrin receptor (TfR) nanobody-functionalized bacterial outer membrane vesicles (OMVs) as an engineered platform for improving Alzheimer's disease (AD)-like neuronal injury. Escherichia coli-derived OMVs were characterized for morphology, size distribution, surface charge, protein features, nanobody loading, residual endotoxin levels, endotoxin-associated inflammatory responses, and repeated-exposure immune-safety readouts, and were evaluated in neuronal-related cells and APP/PS1 mice. Engineered OMVs showed receptor-associated uptake and preferential brain-delivery signals, increased HT-22 cell viability, reduced apoptosis, and elevated mitophagy-associated markers including LC3-II, PINK1, and Parkin. OMV treatment also reduced cytosolic mtDNA abundance, DNA/cGAS colocalization, phosphorylation of STING/TBK1/IRF3, and inflammatory-marker expression. PINK1 or Parkin knockdown partially weakened OMV-associated mitochondrial and inflammatory readouts, whereas pharmacological autophagy modulation supported a contribution of autophagy-related processes without establishing selective PINK1/Parkin-dependent mitophagy in vivo. In APP/PS1 mice, OMV administration improved Morris water maze and rotarod performance and alleviated neuroinflammatory and histological injury-related changes. Overall, anti-TfR nanobody-modified OMVs were associated with enhanced receptor-related delivery, mitophagy-linked phenotypes, reduced cytosolic mtDNA/cGAS-STING-related inflammatory signaling, and improved AD-like outcomes, supporting further delivery-system optimization and mechanistic validation.

Development of effective immune responses against any pathogen requires efficient activation of the innate immune system followed by induction of an appropriate adaptive immune response using specific signaling pathways tailored to each infection. The sole innate immune receptor implicated in the induction of adaptive immunity to retroviral infection is Toll-like receptor 7 (TLR7). We have found that germinal center responses, neutralizing antibody production, and clearance of murine leukemia virus (MLV) infection in BALB.J and C57BL/6N (B6N) mice occurs in the absence of TLR7 signaling. This suggests that a previously unknown alternative sensing pathway exists for the activation of protective immune responses upon retroviral infection. Genetic crosses indicate that the ability to control retroviral infection in the absence of TLR7 signaling is determined by the same recessive mechanism in both B6N and BALB.J mice, suggesting that TLR7-independent responses do not result from a gain-of-function of an alternative pattern recognition receptor. Additionally, we observed that TLR7-deficient BALB.J mice produce neutralizing antibodies against mouse mammary tumor virus (MMTV) infection, indicating that this alternative sensing pathway is active against retroviruses of multiple genera. Finally, we determined that the alternative sensing pathway is also independent of both MyD88 and STING signaling. The ability of mice of two genetic backgrounds to control retroviral infection in the absence of TLR7 signaling provides a valuable tool for the identification of an additional mechanism of retrovirus control. The dissection of this pathway has the potential to alter our understanding of the requirements for the stimulation of antigen-specific neutralizing immunity.

Cyclic GMP-AMP synthase (cGAS) is a cytosolic DNA sensor critical for host defense against microbial infections. To prevent aberrant activation, nuclear cGAS is restrained by nucleosome tethering or by barrier-to-autointegration factor-mediated restriction of its access to DNA. However, its physiological roles beyond enzymatic suppression in the nucleus remain poorly understood. Here, we present a genome-wide map of cGAS chromatin localization in human cells and unexpectedly find its binding beyond nucleosome tethering, where cGAS is recruited to chromatin by AP-1 transcription factors via a potentially previously uncharacterized transactivation domain. Integrated CUT&RUN, RNA-seq, and metabolomics analyses reveal that cGAS transcriptionally regulates cholesterol homeostasis. Mechanistically, cGAS promotes LGALS3 transcription, which inhibits ACAA2 activity to suppress fatty acid oxidation and acetyl-CoA production, thereby lowering intracellular cholesterol levels. Reduced cholesterol lowers plasma membrane phosphatidylinositol [3,4,5] triphosphate [PI(3,4,5)P3] levels and decreases mTOR complex 2 (mTORC2) activation. In contrast, cGAS depletion results in cholesterol accumulation and elevated PI(3,4,5)P3, driving mTORC2/Akt signaling, promoting xenografted tumor growth, and conferring resistance to DNA-damaging agents. In vivo, the elevated hepatic cholesterol levels observed in cGas-/- mice were largely reversed by inhibition of Akt but not STING, whereas cGAS deficiency had minimal effects on lung physiology. Collectively, our findings uncover an unexpected role for nuclear cGAS in orchestrating a transcriptional program that links lipid metabolism to cell proliferation, stress response, and cholesterol homeostasis in liver.

Cardiac aging is a major determinant of cardiovascular morbidity and mortality, characterized by progressive myocardial fibrosis, impaired contractile function, vascular dysfunction, and reduced regenerative capacity. Although mitochondrial dysfunction, cellular senescence, and chronic inflammation are recognized hallmarks of cardiac aging, the upstream molecular events that coordinate these processes remain incompletely understood. Emerging evidence suggests that endogenous repetitive genomic elements, particularly Long Interspersed Nuclear Element-1 (LINE-1), function as critical regulators of age-associated inflammatory responses. LINE-1 elements are normally maintained in a transcriptionally repressed state through DNA methylation and heterochromatin-associated mechanisms. However, aging-associated epigenetic erosion, loss of heterochromatin integrity, and nuclear structural instability promote LINE-1 derepression and accumulation of LINE-1-derived nucleic acids. These endogenous DNA species activate cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) signaling, triggering chronic type I interferon responses, NF-κB activation, and senescence-associated secretory phenotype (SASP) amplification. In the aging heart, persistent LINE-1/cGAS-STING activation may contribute to cardiomyocyte dysfunction, fibroblast activation, immune remodeling, and progressive myocardial fibrosis. Here, we propose LINE-1 reactivation as a molecular convergence mechanism that integrates several fundamental aging hallmarks, including epigenetic instability, genomic stress, and inflammaging. We discuss the mechanistic basis of LINE-1-driven cardiac aging and evaluate emerging therapeutic strategies targeting LINE-1 reverse transcription, STING signaling, and epigenetic restoration. This framework provides a new perspective for understanding cardiac aging and developing precision interventions for age-related cardiovascular disease.

During viral infection and tissue injury, efficient immune clearance of infected or damaged cells is crucial for host defense and homeostasis. Here, we identify mitochondrial RNA (mtRNA) as a broad-spectrum damage-associated molecular pattern (DAMP) that coordinates antiviral and damage-related immune clearance. Through an integrative approach combining in vitro cellular assays and multi-strain murine models, we demonstrate that viral infection and cellular stress promote POLRMT-dependent mtRNA synthesis and its release into the cytosol through BAX/mPTP-mediated mitochondrial pores. Once released, mtRNA activates the MAVS signaling pathway, triggering a robust type I interferon response that operates independently of the cGAS-STING axis. Notably, viruses exploit a negative feedback loop for immune evasion: type I interferon upregulates the exoribonuclease PNPT1, which degrades cytosolic mtRNA and thereby dampens the mtRNA-MAVS axis. Critically, pharmacological inhibition of PNPT1 with lanthanum chloride (LanC), combined with BH3 mimetics that relieve the BCL-2-mediated blockade of BAX/BAK pores, synergistically reactivates mtRNA release and restores antiviral immunity. This dual strategy demonstrates potent antiviral and anti-fibrotic efficacy in preclinical models without significant toxicity. Our findings establish the mtRNA-MAVS axis as a central, broadly applicable immune surveillance pathway and provide a mechanistic framework for developing therapies that overcome both viral immune evasion and the limitations of current STING-targeted agonists.

Synergistic therapeutic nanoplatforms are of considerable interest in tumor therapy, but their translational potential is often limited by complex multicomponent designs and unknown in vivo degradation behavior. Herein, we show a degradable, single-component manganese nanoparticles (Mn NPs) using liquid-phase exfoliation of commercial bulk manganese powder for cancer therapy. In the acidic tumor environment, Mn NPs react with H[+] to generate Mn[2+] and H2 and raise the local pH. The released Mn[2+] drives Fenton-like reactions to produce ROS, while H2 causes mitochondrial damage, which together lead to tumor cell death. In addition, Mn[2+] activates cGAS-STING signaling and initiates antitumor immunity. Moreover, Mn NPs inhibit tumor growth in CT26, 4T1, and B16F10 mouse models and show better effects than Mn[2+] alone. Transcriptomic analysis further reveals activation of multiple immune-related signaling pathways following Mn NP treatment. In a bilateral tumor model, unilateral injection of Mn NPs triggers a systemic antitumor immune response, leading to effective inhibition of distant, untreated tumor growth. Collectively, this study identifies Mn NPs as a simple, biodegradable, single-component nanoplatform that offers a promising new route for multimodal tumor therapy.

The molecular mechanism by which hypoxia-inducible factor 1-alpha (HIF-1α) regulates asthma pathogenesis through modulation of neutrophil extracellular trap (NET) formation remains poorly defined. Given that NETs act as core drivers of allergic asthma pathology by amplifying airway inflammation and inducing airway epithelial ferroptosis, this study aimed to elucidate the regulatory axis involving airway epithelial cell-derived exosomal miR-146a-3p. We proposed that exosomal miR-146a-3p directly targets HIF-1α to suppress the transcriptional activity of its downstream target gene ELANE, thereby blocking NETs formation and ultimately alleviating asthmatic airway inflammation. Using public transcriptomic data from asthma patients, we identified miR-146a-3p as significantly downregulated in asthmatic tissues. BEAS-2B cell-derived exosomes were isolated via differential centrifugation. Employing a PMA-induced neutrophil inflammation model and an HDM/NETs-challenged asthmatic mouse model, we systematically demonstrated that BEAS-2B cell-derived exosomes function as efficient delivery vehicles for miR-146a-3p, transporting it into neutrophils. By targeting and inhibiting HIF-1α nucleocytoplasmic translocation and its transcriptional activation capacity, exosomal miR-146a-3p significantly downregulated the expression of NE-key mediator of NET formation-thereby blocking aberrant NETs release. Moreover, we confirmed that NETs reduction modulates the cGAS-STING signaling pathway, reverses the aberrant expression patterns of the ferroptosis regulatory proteins GPX4, SLC7A11, and ACSL4, maintains cellular iron homeostasis, and subsequently alleviates airway epithelial ferroptosis, ultimately ameliorating asthmatic airway inflammation. Collectively, this study reveals a novel mechanism by which excessive NET accumulation exacerbates asthma pathology through cGAS-STING-mediated airway epithelial ferroptosis. Furthermore, it identifies BEAS-2B cell-derived exosomal miR-146a-3p as a key regulatory molecule and potential therapeutic target, offering a new theoretical framework and intervention strategy for understanding and treating allergic asthma.

Immunologically "cold" tumors remain poorly responsive to immunotherapy because hypoxia-limited oxidative therapy, insufficient immunogenic tumor-cell death, and dysfunctional T-cell responses restrict the transition from local tumor destruction to systemic antitumor immunity. Here, an ultrasound-responsive Ce(IV)-integrated biomimetic nanoplatform, TcEM@Ce-P4CO@M, is developed to address these barriers. Ce[4+]/Ce[3+] redox cycling promotes H2O2 decomposition and O2 generation, sustaining ultrasound-triggered reactive oxygen species production under hypoxic conditions. Concurrently, P4CO-derived carbon monoxide contributes to mitochondrial oxidative stress and lipid peroxidation. Endogenous iron-homeostasis remodeling and impairment of the SLC7A11/glutathione (GSH)/glutathione peroxidase 4 (GPX4) antioxidant defense further favor ferroptosis-associated tumor-cell injury. Mitochondrial dysfunction and DNA damage are accompanied by enhanced stimulator of interferon genes (STING)-associated signaling, while co-delivered MSA-2 further reinforces STING-mediated innate immune activation, collectively contributing to an immunogenic environment that supports dendritic-cell maturation. The engineered tumor-derived membrane promotes tumor-associated delivery and provides CD86-mediated costimulation and anti-TIM-3-mediated checkpoint modulation. In 4T1 tumor models, this coordinated cascade suppresses primary tumors, inhibits non-irradiated distant tumors, reduces experimental pulmonary metastatic burden, and remodels the tumor immune microenvironment. Collectively, TcEM@Ce-P4CO@M converts hypoxia-limited local sonodynamic injury into ferroptosis-linked innate-to-adaptive systemic antitumor immunity.

Adenosine deaminase 2 (ADA2) is a secreted enzyme recently shown to localize to lysosomes and bind DNA, yet its cellular function remains elusive. ADA2 is highly expressed by innate immune cells, particularly macrophages, which are implicated in the inflammatory pathogenesis of ADA2 deficiency (DADA2). Accordingly, we investigated its function in macrophages and identified ADA2 as a key regulator of mitochondrial quality control. Transcriptomic profiling of patient monocytes revealed upregulation of nuclear-encoded mitochondrial genes, alongside stress-response and apoptotic programs. Consistently, ADA2-deficient macrophages exhibited increased mitochondrial mass, hyperpolarization, elevated mitochondrial ROS and ATP production, structural abnormalities in cristae, and defective mitophagy. These defects promoted mitochondrial DNA leakage via VDAC, triggering cytosolic DNA sensing, cGAS-STING activation, and hyperactive JAK-STAT-mediated IFN-I responses. Restoration of ADA2 partially rescued mitochondrial integrity and dampened IFN hyperactivation in macrophages and zebrafish. Together, our findings identify ADA2 as a key regulator of mitochondrial quality control that prevents organelle dysfunction and limits inflammation driven by aberrant IFN signaling.

The cGAS-STING pathway constitutes an evolutionarily ancient antiviral defense mechanism, originating in bacteria. It is conserved in most animals, although with some differences. In insects, the model Drosophila melanogaster provides unique opportunities to decipher the function and regulation of this important pathway in hosts beyond mammals and shed light on its evolution. Here, we review recent advances in Drosophila that clarify the relationship between the cGAS-STING and IMD pathways, which both act through the NF-κB transcription factor Relish. We next consider the expansion of the cGLR family in Drosophila flies and the unexpected discovery of new cyclic dinucleotides (CDNs) besides cGAMP. We present the contribution of these CDNs in induction of systemic immunity and immune training in flies and discuss the conservation of these properties in other insects.

Snakebite envenoming is acknowledged as neglected tropical disease affecting populations of regions with limited access to healthcare systems. However, it had posed a major threat to human health since classical antiquity, when plant- and animal-based remedies were extensively used as antidotes to counteract bites by venomous snakes. Although the provision of safe and effective treatments by 2030 is among the strategic objectives of the World Health Organization, clinically approved therapies currently available remain animal-derived polyclonal antibodies, which suffer from significant limitations, including high production costs, limited cross-species efficacy, and risk of adverse immune reactions. In this review, we survey some historical antidotes used to treat snakebite envenoming in classical antiquity and early modern Europe such as theriac, mithridatum, Orvietan and Mattioli's scorpion oil. We then discuss natural compounds that, while found also as active ingredients in medical plants used for the preparation of the historical antidotes, over the last decades have inspired research efforts aimed at characterizing their evidence-based antivenom activity with the aim of offering potential alternatives to biologic-based therapies. Finally, we outline emerging therapeutic opportunities that, arising from drug repurposing efforts, may pave the way for the development of novel, broadly effective therapies against multiple venomous snake species.