Search Results (609)

Tamarix chinensis Lour. (T. chinensis) is a traditional herb with functions for releasing the exterior to promote the eruption of rashes, among other ailments. However, these function-related pharmacological effects, such as anti-complement, anti-oxidative, and anti-inflammatory, remain unclear. This study aims to reveal the aforementioned effects and the molecular mechanisms of the ethanol extract of T. chinensis (TCE). Our results demonstrated that TCE inhibited classical- and lectin-mediated complement activation, reduced intracellular ROS via NADPH oxidase inhibition, and directly scavenged DPPH radicals and superoxide anions. By using lipopolysaccharide (LPS)-stimulated macrophages, along with LPS-induced acute lung injury (ALI) and endotoxemia mice, the anti-inflammatory activity and the underlying molecular mechanisms of TCE were deeply investigated. In LPS-activated macrophages, it suppressed iNOS, CCL2, IL-6 and IL-1β transcriptionally and translationally. Mechanistically, TCE inhibited NF-κB signaling by blocking IκBα phosphorylation and p65 nuclear translocation, as well as suppresses AP-1 signaling by reducing ERK and JNK phosphorylation. In vivo, TCE lowered serum multiple pro-inflammatory cytokines of endotoxemic mice and alleviated lung injury of ALI mice. Collectively, our results demonstrated that TCE possesses anti-complement and anti-oxidative activities and exerts anti-inflammatory effects through inhibiting NF-κB and AP-1 signaling. These findings provide scientific evidence for supporting the traditional use of T. chinensis. Full article

Background/Objectives: Gouty arthritis (GA) is an inflammatory disease caused by increased purine metabolism. The limitations of current anti-GA therapies remain a major challenge. Astilbin, the main flavonoid in Smilax glabra Roxb., was found to exert potential anti-GA effects in our previous study. Methods: In this study, a mouse model of monosodium urate (MSU)-induced arthritis and an inflammatory model using mouse bone marrow-derived macrophages (BMDMs) were established. Results: Our data showed that astilbin reduced MSU-induced joint swelling and inflammatory infiltration in mice, restored lipopolysaccharide (LPS)/MSU-induced reductions in cell viability, and inhibited the expression levels of inflammatory factors IL-1β, IL-6 and TNF-α. Further studies showed that astilbin significantly reduced MSU-induced increases in NLRP3 and P-p65 protein levels, as well as the expression of ASC, P-IKKα, P-IκBα, and cleaved-caspase-1. Conclusions: This study suggests that astilbin may be a promising natural product for the treatment of GA by inhibiting the activation of the NLRP3 inflammasome and NF-κB signaling pathway. Full article

Boiogito (BOT), a traditional Kampo herbal medicine, has been reported to exhibit anti-inflammatory and anti-obesity properties. However, its potential role in protecting adipocyte function under inflammatory conditions at different stages of adipocyte development remains unclear. This study investigated the effects of BOT on adipogenesis and tumor necrosis factor-α (TNF-α)-induced inflammatory responses in differentiating and mature 3T3-L1 adipocytes. In this study, 3T3-L1 preadipocytes were induced to differentiate and exposed to TNF-α in the presence or absence of BOT during differentiation or after full adipocyte maturation. Lipid accumulation was assessed by Oil Red O staining, while adipokine secretion and inflammatory cytokine production were evaluated by ELISA. The expression of adipogenic markers and inflammatory signaling molecules was analyzed using quantitative PCR and Western blotting. TNF-α significantly inhibited the expression of adipogenesis-related factors at the transcriptional level in adipocytes, reduced lipid accumulation and adiponectin expression, and enhanced inflammatory cytokine production. BOT treatment dose-dependently attenuated these effects, restoring adipogenic capacity and suppressing inflammatory responses in both differentiating and mature adipocytes. Mechanistically, BOT reduced TNF-α-induced activation of the NF-κB pathway, as evidenced by decreased phosphorylation of NF-κB p65 and IκB. These findings demonstrate that BOT preserves adipocyte function and mitigates inflammation-associated adipocyte dysfunction throughout adipocyte development. The protective effects of BOT may contribute to the regulation of obesity-associated metabolic inflammation, partly through modulation of NF-κB signaling. Full article

Acute liver failure (ALF) is characterized by a rapid decline in liver function, leading to metabolic and organ failure. This study employed liver metabolomics, Nuclear Factor kappa-B (NF-κB) signaling pathway analysis, and inflammatory factor profiling to investigate the therapeutic mechanisms of xanthoxylin in ALF. Xanthoxylin administration led to increased antioxidant levels and reduced markers of inflammation and tissue damage. Xanthoxylin downregulated the messenger RNA (mRNA) expression of Nitric Oxide Synthase (NOS), Interleukin-1β (IL-1β), Interleukin-6 (IL-6), Tumor Necrosis Factor-α (TNF-α), NF-κB, Inhibitor of NF-κB α (IκBα), and Toll-like receptor 4 (TLR4), and inhibited the protein expression of p-p38 and p-p65 while upregulating B-cell CLL/Lymphoma 2 (Bcl-2) and B-cell Lymphoma-x (Bcl-xl). Metabolomic analysis identified 41 differentially expressed metabolites, 20 of which showed strong correlations with pharmacodynamic parameters. These 20 candidate metabolite signatures are involved in amino acid and carboxylic acid metabolic pathways, with potential links to glycolysis and the tricarboxylic acid (TCA) cycle. Together, these findings suggest that xanthoxylin exerts therapeutic effects against ALF by modulating the IκBα/NF-κB signaling pathway and related metabolic pathways, providing a scientific basis for understanding its multi-target mechanism. Full article

Objectives: The rhizome of Iris tectorum Maxim. (Chuan She Gan), commonly used as a substitute for She Gan in Sichuan and other regions, is traditionally applied for inflammation-related disorders. This study aimed to evaluate the anti-inflammatory activity of Chuan She Gan ethanolic extract (CSG) and elucidate its molecular mechanism. Methods: CSG was prepared by 85% ethanol extraction and analyzed by HPLC to identify representative constituents. In LPS-stimulated RAW264.7 macrophages, nitrite accumulation and iNOS activity, cytokine production and inflammatory gene expression were evaluated using Griess assays, ELISA, and qRT-PCR, respectively. NF-κB and AP-1/MAPK signaling were determined using luciferase reporter assays and Wwestern blotting. Serum inflammatory cytokine levels of endotoxemic mice were measured by ELISA. Results: Four characteristic isoflavones/glycosides in CSG were identified, including tectoridin, iridin, tectorigenin, and irigenin. In LPS-activated RAW264.7 macrophages, CSG not only dose-dependently suppressed supernatant NO by inhibiting iNOS activity and downregulating iNOS expression, but also reduced IL-6, MCP-1, and intracellular pro-IL-1β at the protein and mRNA levels. Mechanistic analyses indicated that CSG attenuated NF-κB activation by reducing IκBα phosphorylation and limiting p65 nuclear accumulation, while AP-1/MAPK signaling remained largely unchanged. In endotoxemic mice, a single oral gavage of CSG (50–200 mg/kg) significantly lowered serum IL-6, MCP-1, and TNF-α levels. Conclusions: CSG showed anti-inflammatory activity in LPS-stimulated macrophages and endotoxemic mice. In LPS-stimulated macrophages, CSG suppressed inflammatory mediator production primarily through the inhibition of NF-κB signaling. In endotoxemic mice, CSG reduced the serum levels of pro-inflammatory cytokines. These findings provide pharmacological basis for the traditional use of Chuan She Gan. Full article

Objective: The present study investigates the effect of 1,8-cineole on improving lipid metabolism disorder by regulating oxidative stress and inflammation via the TLR4/MyD88/NF-κB pathway. 1,8-Cineole is a monoterpene compound widely found in the essential oils of many plants and has been reported to possess anti-inflammatory and antioxidant activities. However, its role in regulating lipid metabolism disorders remains unclear. This study aimed to investigate the effects of 1,8-cineole on lipid metabolism and explore the potential mechanisms related to oxidative stress and inflammation. Methods: Cell viability was assessed by MTT assay to determine the optimal PA concentration for inducing lipid accumulation and the non-cytotoxic range of 1,8-cineole in HepG2 and AML-12 cells. Lipid droplets were visualized by Oil Red O staining, while triglyceride (TG) and total cholesterol (TC) levels were quantified using enzymatic kits. Oxidative stress markers (ROS by DCFH-DA fluorescence; MDA by TBA method; CAT activity by ammonium molybdate method) and inflammatory cytokines (TNF-α, IL-6, IL-1β, IL-18 by ELISA) were measured. Western blotting analyzed key proteins in the TLR4/MyD88/NF-κB pathway (TLR4, MyD88, p-P65, p-IκBα). Pathway-specific inhibitors were employed for mechanistic validation. Results: 1,8-Cineole (up to 1000 μg/mL) showed no cytotoxicity. It significantly attenuated PA-induced lipid droplet accumulation, reduced TG and TC levels (p < 0.05), and ameliorated oxidative stress by decreasing ROS and MDA while enhancing CAT activity in AML-12 cells (p < 0.01). Furthermore, 1,8-cineole suppressed pro-inflammatory cytokine release (TNF-α, IL-6, and IL-1β; p < 0.01), whereas no significant effect was observed on IL-18 levels. Downregulated TLR4/MyD88/NF-κB pathway activation. Inhibition of TLR4 or NF-κB mirrored these protective effects. Conclusions: 1,8-Cineole alleviates PA-induced lipid metabolism disorders, oxidative stress, and inflammation in hepatocytes, likely through suppression of the TLR4/MyD88/NF-κB signaling pathway. Full article

This study aimed to evaluate the protective effects of sodium humate (HNa) alone and in combination with low-dose cefixime (CFM) in mice infected with enterotoxigenic Escherichia coli (ETEC). An ETEC infection mouse model was established to compare the effects of individual or combined interventions on physiological parameters, intestinal morphology, barrier function, levels of specific intestinal bacterial groups, cell proliferation/apoptosis, and inflammatory pathways. The results showed that the HNa + CFM combination significantly promoted body weight recovery, ameliorated damage to jejunal villus structure and ultrastructure, and increased the mRNA expression of mucins (MUC1/2/3) and tight junction proteins (ZO-1, Occludin, Claudin-1) compared to the ETEC group. Concurrently, the combined treatment significantly reduced fecal E. coli counts and increased the abundance of Lactobacillus and Bifidobacterium, promoted epithelial repair by upregulating proliferation-related genes (EGFR, PCNA, TGF-β1), and decreased the Bax/Bcl-2 ratio. Furthermore, the combined intervention significantly reduced serum LPS levels and consequently suppressed ETEC-induced activation of the TLR4/MyD88/NF-κB pathway, as evidenced by reduced protein expression of TLR4 and MyD88, decreased phosphorylation of IκBα and p65, and diminished nuclear accumulation of NF-κB p65, leading to downregulation of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) and elevation of IL-10. In conclusion, the combined application of HNa and low-dose CFM showed additional protective benefits against ETEC infection. These effects were associated with multi-targeted repair of the intestinal barrier, modulation of measured bacterial levels, and suppression of excessive inflammatory responses. This strategy offers a potential approach for the clinical management of bacterial enteritis and reducing antibiotic dependence. Full article

Background: Triple-negative breast cancer (TNBC) remains a formidable clinical challenge due to the scarcity of targeted therapies and profound metabolic heterogeneity. Although Artemisian B, a dimeric sesquiterpene lactone derived from Artemisia argyi, exhibits potent antiproliferative activity, its comprehensive metabolic footprint and the translation of these perturbations into downstream signaling regulation remain poorly characterized. Methods: To address this gap, we employed an integrative analytical framework combining untargeted metabolomics, topology-guided metabolite–gene network mapping, and parallel experimental validation in MDA-MB-231 cells. This workflow systematically profiled cellular phenotypes, global metabolic reprogramming, and key signaling nodes, enabling the prioritization of high-confidence mechanistic links between metabolic alterations and signal transduction. Results: Artemisian B dose-dependently suppressed TNBC cell viability (IC₅₀ = 12.12 μM) and triggered mitochondrial apoptosis, characterized by Bax upregulation, Bcl-2 downregulation, and caspase-9/3 activation. Untargeted metabolomics identified 129 significantly altered metabolites, reflecting extensive dysregulation across lipid peroxidation, bioenergetics, and nucleotide metabolism. Topological analysis of the metabolite–gene network identified the NF-κB pathway as a highly interconnected hub within this perturbed landscape. Parallel experimental validation corroborated this prediction, demonstrating that Artemisian B consistently suppressed the phosphorylation of IKKα/β, IκBα, and p65, while markedly attenuating p65 nuclear translocation. Conclusions: Artemisian B induces TNBC apoptosis through extensive metabolic reprogramming coupled with concurrent inhibition of NF-κB signaling. By seamlessly integrating untargeted metabolomics with network topology, our framework not only successfully bridges metabolic perturbations with signaling outcomes but also establishes a versatile, dual-perspective strategy applicable to both biochemical reaction networks and signal transduction pathways. This approach provides a robust predictive paradigm for decoding the multi-target pharmacological mechanisms of natural products. Full article

Background/Objectives: Salvia miltiorrhiza is a medicinal plant rich in phenolic acids and tanshinones, compounds that have been linked to anti-inflammatory and neuroprotective activities. ‘Hongdan’ is a Korean cultivar characterized by relatively high levels of salvianolic acid B and tanshinone IIA, but its anti-inflammatory activity in microglial cells has not yet been examined. Methods: Nitrite production and the mRNA expression of inflammatory mediators (iNOS and COX-2) and pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) were examined. In addition, activation of MAPK (ERK1/2, JNK, and p38) signaling pathway and expression of the NF-κB regulatory protein IκB-α were analyzed. Results: The Hongdan extract inhibited nitrite production and reduced the expression of iNOS and COX-2 in LPS-stimulated BV2 microglial cells. In addition, the expression of IL-1β and IL-6 was markedly reduced, whereas TNF-α was significantly suppressed only at the highest concentration tested. Furthermore, phosphorylation of ERK1/2, JNK, and p38 was significantly inhibited, while IκB-α degradation was not altered. Conclusions: These findings demonstrate that the Hongdan extract effectively suppresses LPS-induced inflammatory responses through inhibition of MAPK signaling pathways and may serve as a promising natural therapeutic candidate for neuroinflammatory disorders. Full article

Colorectal cancer (CRC) and non-Hodgkin lymphoma (NHL) are among the most prevalent cancer types globally by incidence and mortality. Both types are influenced differentially by chronic inflammation. Central to this inflammation are inflammatory genes that are meticulously regulated by nuclear factor kappa B (NF-κB) and tumor necrosis factor-α (TNF-α). NF-κB is negatively regulated by IκBα (encoded by NFKBIA), while TNF-α’s actions can be modulated by ghrelin (encoded by GHRL). We investigated four single nucleotide polymorphisms (SNPs) in NFKB1 (rs4648068), NFKBIA (rs2233406), TNF-α (rs1800629), and GHRL (rs1629816) as biomarkers for CRC and NHL risk in a cohort of Kuwaiti individuals. DNA samples from patients and controls were collected and genotyped for all SNPs, and their association with CRC or NHL risk was assessed. While rs4648068 showed a modest association with increased CRC risk, it had no significant impact on NHL risk. Conversely, rs2233406 increased NHL risk without affecting CRC risk. Interestingly, while rs1800629 showed a protective effect against NHL, it showed an increased risk for CRC. Finally, rs1629816 was associated with greater NHL but not CRC risk. Our findings suggests that variations of these inflammatory genes may be useful indicators for predicting cancer risk but might have unpredictable effects on cancer susceptibility, depending on the cancer type. Full article

Ulcerative colitis (UC) is a form of inflammatory bowel disease (IBD), which is marked by severe abdominal pain, weight loss, perianal bleeding, and diarrhea. This study successfully isolated and purified four low-molecular-weight fucoidan oligosaccharides through acid hydrolysis and Bio Gel P10 gel filtration. The molecular weights were 2.9 × 10⁴–1.36 × 10⁵ Da, 182–1012 Da, 161–939 Da and 161–939 Da, respectively. A mouse model of colitis was induced using Dextran Sulfate Sodium (DSS). The results indicated that fucoidan and fucoidan oligosaccharides could ameliorate murine ulcerative colitis, with the oligosaccharides (200 mg/kg/d) demonstrating superior therapeutic effects. This superiority was likely attributed to the lower molecular weight and higher content of total sugars and fucose. The primary mechanisms involved the modulation of gene and protein expression levels associated with the Toll-like receptor 4, Myeloid differentiation primary response 88, nuclear factor kappa-light-chain-enhancer of activated B cells, p65, and Inhibitor of kappa light polypeptide gene enhancer in B cells, alpha (TLR4, MYD88, NF-κB p65, and IκB-α) signaling pathways, which reduce the production of inflammatory cytokines such as tumor necrosis factor-alpha, Interleukin-1 beta and Interleukin-6 (TNF-α, IL-1β, and IL-6). Additionally, these oligosaccharides alleviated oxidative stress, enhanced the levels of intestinal barrier proteins (Claudin family member 4 and Zonula occludens protein 1), regulated the abundance and diversity of the gut microbiota, and increased the levels of short-chain fatty acids (SCFAs) in the intestine. It is worth emphasizing that this study can only demonstrate that fucoidan oligosaccharides have a mitigating effect on intestinal inflammation in mice. Further research is needed in the future to investigate the structure–activity relationship of fucoidan oligosaccharides and their impact on human intestinal microbiota, in order to further elucidate their anti-inflammatory mechanisms. Full article

Artemether, a derivative of the natural compound artemisinin, is increasingly recognized for its multi-target anti-inflammatory and antioxidant properties. This review systematically elucidates the molecular mechanisms underlying these effects, focusing on artemether’s dual modulation of the MAPK/NF-κB and Nrf2 signaling pathways. We detail how artemether concurrently inhibits the MAPK/NF-κB axis—suppressing IKKβ phosphorylation and IκBα degradation to block NF-κB nuclear translocation—and downregulates p38/contextually modulates ERK phosphorylation. This leads to a significant reduction in key inflammatory mediators, including TNF-α, IL-6, and COX-2. Simultaneously, artemether activates the Nrf2 antioxidant pathway, upregulating HO-1 expression and enhancing the activity of SOD and GSH-Px, which effectively scavenges free radicals and reduces markers of oxidative damage such as MDA and 8-OHdG. The core therapeutic synergy arises from artemether’s disruption of the ROS-NF-κB positive feedback loop, which inhibits neutrophil infiltration and lipid peroxidation, thereby ameliorating tissue injury in experimental models of arthritis and neurodegenerative diseases. Compared to conventional NSAIDs and glucocorticoids, artemether exhibits a favorable safety profile, particularly regarding gastrointestinal effects, and demonstrates unique immunomodulatory potential. Future research directions should prioritize the development of nano-targeted delivery systems and the elucidation of pathway crosstalk at the single-cell level to advance the clinical translation of artemether for chronic inflammatory diseases. Full article

Earthworm-derived bioactive compounds are emerging as promising pharmaceutical agents; however, the immunomodulatory effects of Eisenia fetida proteins at different developmental stages remain unclear. This study evaluated, for the first time, the stage-dependent immunomodulatory activity of E. fetida protein extracts in RAW 264.7 macrophages. Soluble proteins isolated from juvenile, mature, and senescent worms were lyophilized and tested for their effects on cell viability, phagocytic activity, nitric oxide (NO), reactive oxygen species (ROS), and inflammatory gene expression. Amino acid profiling and Western blot analysis were additionally performed to investigate biochemical composition and signaling mechanisms. Mature-stage extracts exhibited the highest protein yield, minimal cytotoxicity, enhanced macrophage phagocytosis, and significant suppression of LPS-induced NO, ROS, and proinflammatory cytokines. In contrast, juvenile-stage extracts showed moderate immunomodulatory activity, whereas senescent-stage extracts induced oxidative stress and inflammatory responses. Western blot analysis demonstrated that mature-stage proteins strongly inhibited phosphorylation of NF-κB and MAPK signaling proteins, including p65, IκBα, p38, ERK1/2, and JNK, while senescent-stage extracts maintained elevated pathway activation. Amino acid analysis further revealed enriched immunologically relevant amino acids in mature-stage extracts. These findings demonstrate that developmental stage strongly influences the biological activity of E. fetida proteins and highlight mature-stage extracts as promising natural immunomodulatory agents. Full article

Objective: In this study, Dehydroepiandrosterone (DHEA-induced Polycystic Ovary Syndrome (PCOS) mice were used as models to evaluate the improvement effect of Vitexin (Vit) on ovarian fibrosis and explore the mechanism of action of the NR4A1/NLRP3 signaling pathway. Method: Sixty 4-week-old female ICR mice of the same batch number were selected and their systems were divided into 6 groups (n = 10): normal (Control, Ctrl) group, model (Polycystic Ovary Syndrome, PCOS) group, treatment (Vitexin, The Vit group, normal NR4A1 gene silencing group (Ctrl NR4A1^-/-), NR4A1 gene silencing model group (PCOS NR4A1^-/-), and NR4A1 gene silencing treatment group (Vit NR4A1^-/-). Silencing gene modeling was performed by tail vein injection of adeno-associated virus (serotype AAV-8), and the mouse genotypes were detected by qRT-PCR technology 14 days after injection. After the genotype was determined, the PCOS group and the PCOS NR4A1^-/- group were administered dehydroepandrosterone (6 mg/100 g/d) by gavage for 28 consecutive days for modeling, while the Vit group and the Vit NR4A1^-/- group were treated with dehydroepandrosterone + vitexin (10 mg/kg/d) by gavage for 28 consecutive days. All mice were raised with pure water and regular maintenance food. After 4 weeks of drug intervention, the mice were euthanized and samples were collected. The pathological changes in ovarian tissue were observed by H&E staining, and the degree of ovarian tissue fibrosis was observed by Masson staining. The levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GSH-Px), malondialdehyde (MDA), total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in mouse serum were detected by biochemical kits. The levels of inflammatory factors (IL-1β, IL-6, IL-18, TNF-α) in mouse serum were determined by enzyme-linked immunosorbent assay. Real-time fluorescence quantitative PCR (qRT-PCR) was used to detect oxidative kinase (Gsta4, Prdx3, Mgst1, Gpx3, Gsr), inflammatory factors (Nlrp3, Caspase-1, Asc, Il-1β, Il-18, Tnf-α) and fibrotic pathway-related genes (Tgf-β1, Smad3, Collagen1, CTGF, α-SMA, Mmp-13, and β-catenin) in ovarian tissues. The levels of inflammatory factors (NLRP3, Caspase-1, ASC, IL-1β, IL-18, TNF-α, IκBα) and fibrosis in mice were determined by Western blot method, and statistical description and analysis were performed using SPSS software. Result: In the wild-type genotype group, compared with the PCOS group, Vit treatment could effectively regulate the metabolic abnormalities of PCOS mice, including inhibiting excessive weight gain, restoring normal glucose tolerance, and reducing body fat content. After Vit treatment, the levels of MDA, TC, TG, LDL, IL-1β, IL-6, IL-18 and TNF-α in the serum of PCOS mice were significantly reduced, while the levels of SOD and HDL in the serum of PCOS mice were increased. The staining results indicated that Vit treatment could significantly inhibit the process of ovarian fibrosis in PCOS mice. The results of WB and PCR demonstrated that after Vit gavage treatment in mice, inflammatory and fibrotic factors such as Nlrp3, Caspase-1, Asc, Il-1β, Il-18, Tgf-β1, Smad3, Collagen1, CTGF, and α-SMA in ovarian tissues could be significantly down-regulated, and the fibrotic level of ovarian tissues could be reduced. Among the same measurement indicators, the silenced NR4A1 group showed a certain degree of increase compared with the wild genotype group, but there was no significant difference. Conclusions: Vit intervention can restore the sex hormone levels and follicular development in ovarian tissues of PCOS mice, regulate reproductive endocrine disorders and abnormal lipid metabolism levels, and regulate the expression of Collagen I, a-SMA and CTGF in the ovaries by inhibiting the NR4A1/NLRP3 signaling pathway, thereby improving the ovarian fibrosis level of PCOS mice. It is suggested that it may play a key role in the treatment of PCOS and the prevention and delay of its long-term complications. Full article

During donation after circulatory death (DCD), circulating levels of mitochondrial damage-associated molecular patterns (mtDAMPs) may increase, thereby exposing donor hearts to mtDAMPs prior to procurement and during machine perfusion. Mitochondrial DNA (mtDNA) is a pro-inflammatory mtDAMP that may stimulate several intracellular cascades including that of toll-like receptor 9 (TLR9). We administered mtDNA or ODN2088 (TLR9 antagonist) to hearts at reperfusion onset using an isolated rat heart model of DCD transplantation to investigate their effects. Four experimental groups were compared: (1) no ischemia; (2) ischemia; (3) ischemia + mtDNA; (4) ischemia + ODN2088. During reperfusion, cardiac power in ischemic hearts was significantly reduced compared to non-ischemic hearts (p < 0.01), and was further decreased with mtDNA (p < 0.05), but remained unchanged with ODN2088. Reduced ventricular recovery in mtDNA-treated hearts likely resulted from lower recovery of oxidative metabolism, demonstrated by reduced oxygen efficiency (p < 0.05) and a strong tendency for increased cytochrome c release (p < 0.06),indicating mitochondrial dysfunction and disruption, respectively. ODN2088 phosphorylated IκBα (NF-κB inhibitor alpha) and appeared to decrease cardiomyocyte death compared to ischemic hearts. Given the detrimental effects of circulating mtDNA on cardiac functional and metabolic recovery, circulating mtDAMPs, and particularly mtDNA, are of clinical relevance as potential therapeutic targets for optimizing graft quality and post-transplant outcomes. Full article