Background: Indoleamine-2,3-dioxygenase 1 (IDO1) is responsible for tumor immune escape by regulating T cell-associated immune responses and promoting the activation of immunosuppressive. Given the vital role of IDO1 in immune response, further investigation on the regulation of IDO1 in tumors is needed.Methods: Herein, we used ELISA kit to detect the interferon-gamma (IFN-γ), Tryptophan (Trp), and kynurenic acid (Kyn) levels; western blot, Flow cytometry, and immunofluorescence assays detected the expression of the proteins; Molecular docking assay, SPR assay and Cellular Thermal Shift Assay (CETSA) were used to detect the interaction between IDO1 and Abrine; nano live label-free system was used to detect the phagocytosis activity; tumor xenografts animal experiments were used to explore the anti-tumor effect of Abrine; flow cytometry detected the immune cells changes.Results: The important immune and inflammatory response cytokine interferon-gamma (IFN-γ) up-regulated the IDO1 expression in cancer cells through the methylation of 6-methyladenosine (m6A) m6A modification of RNA, metabolism of Trp into Kyn, and JAK1/STAT1 signaling pathway, which could be inhibited by IDO1 inhibitor Abrine. CD47 is IFN-γ-stimulated genes (ISGs) and prevents the phagocytosis of macrophages, leading to the cancer immune escape, and this effect could be inhibited by Abrine both in vivo and in vitro. The PD-1/PD-L1 axis is an important immune checkpoint in regulating immune response, overexpression of PD-1 or PD-L1 promotes immune suppression, while in this study Abrine could inhibit the expression of PD-L1 in cancer cells or tumor tissue. The combination treatment of Abrine and anti-PD-1 antibody has a synergistic effect on suppressing the tumor growth through up-regulating CD4⁺ or CD8⁺ T cells, down-regulating the Foxp3⁺ Treg cells, and inhibiting the expression of IDO1, CD47, and PD-L1.Conclusion: Overall, this study reveals that Abrine as an IDO1 inhibitor has an inhibition effect on immune escape and has a synergistic effect with the anti-PD-1 antibody on the treatment of HCC.

Background: Tanshinone I (TI) is a primary component of Salvia miltiorrhiza Bunge (Danshen), which confers a favorable role in a variety of pharmacological activities including cardiovascular protection. However, the exact mechanism of the cardiovascular protection activity of TI remains to be illustrated. In this study, the cardiovascular protective effect and its mechanism of TI were investigated. Methods: In this study, tert-butyl hydroperoxide (t-BHP)-stimulated H9c2 cells model was employed to investigate the protective effect in vitro. The cell viability was determined by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay and lactate dehydrogenase (LDH) kit. The reactive-oxygen-species (ROS) level and mitochondrial membrane potential (MMP) were investigated by the flow cytometry and JC-1 assay, respectively. While in vivo experiment, the cardiovascular protective effect of TI was determined by using myocardial ischemia–reperfusion (MI/R) model including hematoxylin–eosin (H&E) staining assay and determination of superoxide dismutase (SOD) and malondialdehyde (MDA). Tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) release were detected by Enzyme-linked immunosorbent assay (ELISA). Receptor interacting protein kinase 1 (RIP1), receptor interacting protein kinase 3 (RIP3), receptor interacting protein kinase 3 (MLKL), protein kinase B (Akt), Nuclear factor erythroid 2 related factor 2 (Nrf2), Heme oxygenase-1 (HO-1) and NAD(P)H: quinone oxidoreductase-1 (NQO-1) were determined by western blotting. Results: Our data demonstrated that TI pretreatment attenuated t-BHP and MI/R injury-induced necroptosis by inhibiting the expression of p-RIP1, p-RIP3, and p-MLKL. TI activated the Akt/Nrf2 pathway to promote the expression of antioxidant-related proteins such as phosphorylation of Akt, nuclear factor erythroid 2 related factor 2 (Nrf2), quinone oxidoreductase-1 (NQO-1) and heme oxygenase-1 (HO-1) expression in t-BHP-stimulated H9c2 cells. TI relieved oxidative stress by mitigating ROS generation and reversing MMP loss. In vivo experiment, TI made electrocardiograph (ECG) recovery better and lessened the degree of myocardial tissue damage. The counts of white blood cell (WBC), neutrophil (Neu), lymphocyte (Lym), and the release of TNF-α and IL-6 were reversed by TI treatment. SOD level was increased, while MDA level was decreased by TI treatment. Conclusion: Collectively, our findings indicated that TI exerted cardiovascular protective activities in vitro and in vivo through suppressing RIP1/RIP3/MLKL and activating Akt/Nrf2 signaling pathways, which could be developed into a cardiovascular protective agent.

Background: Pneumonia refers to the inflammation of the terminal airway, alveoli and pulmonary interstitium, which can be caused by pathogenic microorganisms, physical and chemical factors, immune damage, and drugs. Anemoside B4, the major ingredient ofPulsatilla chinensis(Bunge) Regel, exhibited anti-inflammatory activity. However, the therapeutic effect of anemoside B4 on pneumonia has not been unraveled. This study aims to investigate that anemoside B4 attenuates the inflammatory responses inKlebsiella pneumonia(KP)- and influenza virus FM1 (FM1)-induced pneumonia mice model.Methods: The network pharmacology and molecular docking assays were employed to predict the targets of anemoside B4’s treatment of pneumonia. Two models (bacterial KP-infected mice and virus FM1-infected mice) were employed in our study. BALB/c mice were divided into six groups: control, model group (KP-induced pneumonia or FM1-induced pneumonia), anemoside B4 (B4)-treated group (2.5, 5, 10 mg/kg), and positive drug group (ribavirin or ceftriaxone sodium injection). Blood samples were collected for hematology analysis. The effects of B4 on inflammation-associated mediators were investigated by Enzyme-linked immunosorbent assay (ELISA) and hematoxylin and eosin staining (HE) staining. Proteins expression was quantified by western blotting.Results: The network results indicated that many pro-inflammatory cytokines such as tumor necrosis factor α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6) participated in anemoside B4’s anti-inflammatory activity. The counts of neutrophil (NEU) and white blood cell (WBC), the level of myeloperoxidase (MPO), and the release of pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 increased by KP or FM1 infection, which were reversed by anemoside B4. In addition, anemoside B4 significantly suppressed the FM1-induced expression of toll-like receptor 4 (TLR4), myeloid differential protein-88 (MyD88), and myeloid differentiation protein-2 (MD-2), which were further validated by molecular docking data that anemoside B4 bound to bioactive sites of TLR4. Therefore, anemoside B4 exhibited a significant therapeutic effect on pneumonia via the TLR4/MyD88 pathway.Conclusion: Our findings demonstrated that anemoside B4 attenuates pneumonia via the TLR4/Myd88 signaling pathway, suggesting that anemoside B4 is a promising therapeutic candidate for bacterial-infected or viral-infected pneumonia.

Lung cancer is the primary cause of cancer-related mortality worldwide owing to its strong metastatic ability. EGFR-TKI (Gefitinib) has demonstrated efficacy in metastatic lung cancer therapy, but most patients ultimately develop resistance to Gefitinib, leading to a poor prognosis. Pedunculoside (PE), a triterpene saponin extracted from Ilex rotunda Thunb., has shown anti-inflammatory, lipid-lowering and anti-tumor effects. Nevertheless, the therapeutic effect and potential mechanisms of PE on NSCLC treatment are unclear. To investigate the inhibitory effect and prospective mechanisms of PE on NSCLC metastases and Gefitinib-resistant NSCLC. In vitro, A549/GR cells were established by Gefitinib persistent induction of A549 cells with a low dose and shock with a high dose. The cell migratory ability was measured using wound healing and Transwell assays. Additionally, EMT-related Markers or ROS production were assessed by RT-qPCR, immunofluorescence, Western blotting, and flow cytometry assays in A549/GR and TGF-β1-induced A549 cells. In vivo, B16-F10 cells were intravenously injected into mice, and the effect of PE on tumor metastases were determined using hematoxylin-eosin staining, Caliper IVIS Lumina, DCFH₂-DA staining, and western blotting assays. PE reversed TGF-β1-induced EMT by downregulating EMT-related protein expression through MAPK and Nrf2 pathways, decreasing ROS production, and inhibiting cell migration and invasion ability. Moreover, PE treatment enabled A549/GR cells to retrieve the sensitivity to Gefitinib and mitigate the biological characteristics of EMT. PE also significantly inhibited lung metastasis in mice by reversing EMT proteins expression, decreasing ROS production, and inhibiting MAPK and Nrf2 pathways. Collectively, this research presents a novel finding that PE can reverse NSCLC metastasis and improve Gefitinib sensitivity in Gefitinib-resistant NSCLC through the MAPK and Nrf2 pathways, subsequently suppressing lung metastasis in B16-F10 lung metastatic mice model. Our findings indicate that PE is a potential agent for inhibiting metastasis and improving Gefitinib resistance in NSCLC.

Acute gouty arthritis is a metabolic disorder caused by monosodium urate (MSU) accumulation, leading to NLRP3 inflammasome activation and joint inflammation. Anemoside B4 (B4), a pentacyclic triterpenoid saponin, exerts significant anti-inflammatory effects. However, the precise molecular mechanisms underlying its therapeutic action, particularly its targeting of key components in NLRP3 inflammasome activation, remain unclear. The aim of this study was to elucidate the therapeutic mechanisms and target of B4 in treating MSU-induced macrophage pyroptosis and acute gouty arthritis, focusing specifically, on its interaction with NEK7, a critical regulator of NLRP3 inflammasome activation. Comprehensive in vitro and in vivo methods were employed to examine the effects and mechanisms of B4. In vitro analyses included Western blot, co-immunoprecipitation (Co-IP), and immunofluorescence assays to assess NLRP3 inflammasome components and NEK7-NLRP3 interactions. The binding of B4 to NEK7 was evaluated using molecular docking, surface plasmon resonance (SPR), cellular thermal shift assay (CETSA), drug affinity responsive target stability (DARTS), NEK7 gene silencing, and site-specific amino acid mutation experiments. In vivo, MSU-induced acute gouty arthritis mouse models and NEK7 knockdown mouse models were used to demonstrate the therapeutic effects and specificity of B4. This study provides the first evidence that B4 significantly inhibits MSU-induced inflammation and pyroptosis in macrophages by directly targeting NEK7 and disrupting the NEK7-NLRP3 complex, thereby reducing NLRP3 inflammasome activation. Additionally, B4 effectively suppressed MSU-induced ROS production, mitochondrial damage, and NF-κB activation. In vivo, B4 alleviated symptoms of acute gouty arthritis, reduced NLRP3 expression, and demonstrated specificity for NEK7 in NEK7 knockdown mouse models. This study highlights B4 as an effective inhibitor of NLRP3 inflammasome activation by directly targeting NEK7, thereby mitigating inflammation and pyroptosis in acute gouty arthritis. These findings position B4 as a prospective therapeutic candidate for the management of acute gouty arthritis, providing insights into its molecular targets and mechanisms.

Lung cancer persists as the foremost malignancy in terms of both incidence and mortality on a global scale. Paraptosis, a unique type of regulated cell death, differs from apoptosis in its reliance on protein synthesis and its morphological hallmarks, characterized by cytoplasmic vacuole formation and distension of the endoplasmic reticulum (ER) or mitochondrial swelling. Flemiphilippinin A (Flp-A), a bioactive compound isolated from Flemingia prostrata, exhibits potent antitumor activity against lung cancer. However, the precise molecular mechanism underlying Flp-A's anti-lung cancer effects-as well as its direct cellular targets-remain unclear, leaving a critical gap in current research. To reveal the therapeutic role and molecular mechanism of Flp-A in lung carcinoma and, for the initial time, evaluate its effects on gefitinib resistance in lung carcinoma cells. The efficacy and mechanism of Flp-A were probed through in vitro and in vivo experiments. In vitro assays included the MTT assay, plate cloning assay, EdU proliferation assay, cell morphology analysis, Western blotting, transcriptome sequencing, JC-1 staining, ROS detection, Seahorse energy metabolism analysis, immunofluorescence, molecular docking, and CETSA to evaluate the therapeutic effects and mechanistic action of Flp-A on lung carcinoma. In vivo experiments were conducted using a mouse xenograft model of lung cancer to assess the antitumor efficacy, systemic toxicity, and underlying mechanisms of Flp-A. This study illustrated the therapeutic potential of Flp-A against lung carcinoma through in vitro and in vivo experiments. Our findings revealed that Flp-A triggers paraptosis in lung The evidence points to the possibility that Flp-A can enhance the specificity of drug-resistant lung carcinoma cells to gefitinib. cells by targeting c-Myc and hyperactivating endoplasmic reticulum (ER) stress. Furthermore, excessive ER stress created a glutamine-deficient environment, which promoted CHOP accumulation in mitochondria. This led to mitochondrial membrane potential disruption, increased ROS production, and inhibition of mitochondrial oxidative phosphorylation. Additionally, we observed that low concentrations of Flp-A combined with gefitinib triggered excessive ER stress and activated paraptosis in gefitinib-resistant lung cancer cells. The evidence points to the possibility that Flp-A can enhance the sensitivity of drug-resistant lung carcinoma cells to gefitinib. This research demonstrates that Flp-A exerts potent anti-tumor effects against lung cancer by targeting c-Myc to induce excessive ER stress and mediating mitochondrial dysfunction through CHOP, ultimately triggering paraptosis. Furthermore, Flp-A increases sensitivity of lung carcinoma cells to gefitinib through paraptosis induction. Our work provides the first evidence for Flp-A's anti-lung cancer mechanism via the paraptosis pathway and confirms its safety profile. Additionally, we reveal Flp-A's capacity to bypass gefitinib resistance in lung carcinoma. These findings position Flp-A as a encouraging novel medical agent for lung cancer treatment, particularly for addressing drug resistance.