Tiao-Shen-Zhi-Ai Formula (TSZAF) is a compound prescription of traditional Chinese medicine used clinically for the treatment of ovarian cancer. In this study, we selected three main active ingredients from TSZAF and combined them into a new TSZAF monomer combination (TSZAF mc) to investigate its effects and mechanisms on inhibiting ovarian cancer proliferation and inducing apoptosis. The effects of TSZAF mc on proliferative activity and apoptosis in ovarian cancer HEY and SKOV3.IP1 cells were assessed in vitro using CCK-8 assay, colony formation assay, and apoptosis assay. Micromethods, flow cytometry, and immunofluorescence were employed to evaluate the impact of TSZAF mc on aerobic glycolytic metabolites and mitochondrial membrane potential. The mRNA and protein expression of key glycolytic genes were detected by quantitative real-time PCR (RT-PCR) and Western blot (WB). An ovarian cancer subcutaneous tumor model was established in NOD-SCID mice using SKOV3.IP1 cells. TSZAF mc was administered via continuous intraperitoneal injection, and its antitumor efficacy in vivo was assessed through anatomical observation, hematoxylin and eosin (H&E) staining, and immunohistochemistry (IHC). Further RT-PCR, WB, and IHC were performed to validate the expression of key upstream glycolytic genes at mRNA and protein levels. Drug affinity responsive target stability (DARTS) and cellular thermal shift assay (CETSA) were used to confirm binding targets. Molecular docking was performed to predict the binding interactions between the monomers and AKT. Finally, the regulatory relationships within signaling pathways were elucidated based on functional assays. TSZAF mc effectively inhibited ovarian cancer proliferation and induced apoptosis. It reduced lactate and ATP production, downregulated mitochondrial membrane potential, and decreased the mRNA and protein expression of key glycolytic genes, including HK2, PKM2, PFKM, GLUT1, and LDHA. In vivo, TSZAF mc suppressed ovarian cancer growth. Moreover, TSZAF mc downregulated the expression of phosphorylated AKT (p-AKT) and phosphorylated FOXO3A (p-FOXO3A) at both protein and tissue levels. CETSA and DARTS demonstrated that TSZAF mc binds AKT. The AKT activator SC79 reversed the inhibitory effects of TSZAF mc on ovarian cancer proliferation and the downregulation of glycolytic proteins. TSZAF mc inhibits ovarian cancer progression by regulating the AKT/ FOXO3A-mediated glycolysis pathway, which may represent one of the mechanisms underlying the clinical efficacy of TSZAF in ovarian cancer treatment.

BackgroundProstate cancer (PCa) is the most common male reproductive malignancy, with traditional Chinese medicine (TCM) offering potential complementary treatments. Aidi injection, used for liver, lung, and colorectal cancers, shows promise for PCa, but its mechanisms are unclear. This study aims to uncover the molecular pathways through which Aidi injection affects PCa using network pharmacology and experiments.MethodsThe study used network pharmacology to identify active components of Aidi injection and their targets from various databases. These targets were cross-referenced with PCa-related targets to uncover therapeutic possibilities. Protein-protein interaction (PPI) and KEGG pathway analyses highlighted key molecular pathways affected by Aidi injection. Molecular docking examined ingredient-target binding. In vitro tests evaluated effects on PCa cell proliferation, apoptosis, and IL-6/STAT3 signaling, while in vivo studies assessed tumor growth and immune response in mice.ResultsThe study identified 58 active components and 804 PCa-related target genes, with 49 overlapping therapeutic targets. Key compounds such as quercetin, kaempferol, isorhamnetin, and cantharidin were highlighted. Core targets included ESR1, IL-6, STAT3, HSP90AA1, EGFR, and VEGFA. KEGG analysis revealed the IL-6/STAT3 and EGFR pathways as significant in PCa treatment. In vitro, Aidi injection inhibited PCa cell proliferation, induced apoptosis, and suppressed IL-6/STAT3 activation. In vivo, Aidi injection reduced tumor growth and enhanced immune response in mice.ConclusionsAidi injection influences the IL-6/STAT3 and EGFR pathways, suppressing PCa cell growth, inducing apoptosis, and reducing migration and invasion. These findings offer valuable insights into the therapeutic potential of Aidi injection for prostate cancer.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12575-025-00299-w.

Membrane-bound vesicles called apoptotic bodies (ApoBDs) formed during apoptosis are being recognized as non-invasive biomarkers for neurological diseases like dementia and ischemic stroke. Centrifugation-based isolation of these vesicles from blood was suggested in a recent paper by Serrano-Heras et al. (2020), which asserted their use for clinical diagnosis. This initial study provides a foundational methodology for ApoBD diagnostics. Albeit, centrifugation parameters employed present technical challenges that require optimization to ensure the purity and reproducibility of ApoBD preparations for clinical utility. Due to the use of improper centrifugation speeds (160×g and 700×g) not consistent with accepted norms for the separation of platelets (ideally 2,000-5,000×g), running the risk of being seriously contaminated by platelets and platelet-derived EVs (P-EVs), which make up 70-90% of EVs in circulation. Such contamination can render functional studies problematic, cloud proteomic and size-distribution analysis, and lower the specificity of ApoBDs enumeration on the basis of Annexin V/PI labeling. Even more extreme centrifugation has little chance to eliminate platelets completely, according to comparative literature data. Increased centrifugal pressure, density gradient separation, size-exclusion chromatography (SEC), and confirmation with platelet-specific markers are among some procedural changes recommended to improve sample purity in attempts to solve such problems. Even as the Serrano-Heras et al. study pinpoints the potential of ApoBDs as biomarkers, constraints to the procedure under consideration pose the threat of bias in future studies. Proper isolation of ApoBDs requires multimodal confirmation, optimized platelet removal protocols and clinically feasible methods like SEC are essential to enhance ApoBDs isolation for neurological disease diagnostics. Rectification of such issues enhance significantly the practicality of ApoBDs in clinical practice for the detection of neurological illnesses.

With its increasing prevalence, diabetes continues to be a major global health concern. Insulin resistance and elevated blood glucose levels are the results of type 1 diabetes (T1D), an autoimmune disease that destroys pancreatic β cells. The underlying immune-mediated harm cannot be prevented or reversed despite a variety of therapy approaches. Recent advancements in immunotherapy are positioned as a viable option for treating T1D. A genetically modified T cell that recognizes and destroys disease-specific targets is used in chimeric antigen receptor (CAR) T-cell therapy. Its targeting of the CD19 and BCMA antigens led to a breakthrough in hematologic malignancies. The present knowledge of CAR-T treatment in T1D is expanded upon by this review. However, there are still a number of important obstacles that limit clinical application, including safety, effectiveness, and technological problems. The main achievements of CAR-T cell treatment for T1D will be discussed in this paper, along with the present challenges and potential future paths for customized CAR-T cell therapies. By combining existing knowledge with uncharted territory, this review aims to stimulate more research and innovation in the field, perhaps leading to a paradigm shift in the treatment of diabetes.

Western blot is one of the most routinely conducted biochemical assays due to its technical ease and relatively low cost. The use of antibody is at the center of Western blot assay, providing great sensitivity and specificity. However, challenges can be posed when using a rare antibody stock. There have been efforts to improve the Western blot procedure to minimize the use of antibody, but these methods require specialized devices. In this study, we hypothesized that the conventional large pool of antibody is not essential for detection and attempted applying only a small volume of antibody. We used sheet protector (SP), a common stationery material that protects office documents, to effectively distribute the antibody solution over the nitrocellulose (NC) membrane. This way, 20-150 µL antibody solution was sufficient for mini-sized membrane, which was adjustable depending on the size of the membrane. We confirmed that the sensitivity and specificity of this SP strategy was comparable to conventional (CV) method. The SP strategy brought a few additional advantages including: (1) antibody incubation without agitation (2), incubation at room temperature, and (3) faster detection on the order of minutes. Finally, we examined 15-min incubation SP protocol using time-series apoptosis samples. We propose that SP strategy is a universally accessible approach using a common consumable, greatly enhancing the efficiency of antibody consumption and incubation time in Western blot assays.

ObjectiveGastric cancer (GCa) is a common malignancy where the epithelial-to-mesenchymal transition (EMT) pathway and lncRNAs are key to proliferation and metastasis. This study aimed to identify lncRNAs regulating the EMT pathway as therapeutic and diagnostic targets in GCa.MethodsWe utilized the TCGA-STAD dataset and differential expression gene (DEG) analysis was performed to identify overlapping genes between DEGs and the EMT pathway. The LASSO feature reduction algorithm and ROC curve analysis were applied to prioritize candidate biomarkers. Additionally, immunological and immunotherapy analyses were conducted to evaluate the significance of TIMP1 in GCa. Pearson correlation analysis was performed between TIMP1 and all differentially expressed lncRNAs, and the most strongly correlated candidates were selected. Experimental validation of the results was carried out using RT-qPCR with 25 cancerous and 25 adjacent normal tissue samples in GCa.ResultsDEG analysis identified 2,926 DEGs, among which 87 were associated with the EMT pathway. The LASSO algorithm reduced this list to 11 genes, and ROC curve analysis identified TIMP1 as a candidate biomarker in GCa. Higher TIMP1 expression was associated with improved response rates. Correlation analysis revealed that LINC01615 was most strongly co-regulated with TIMP1 in GCa. RT-qPCR confirmed that both TIMP1 and LINC01615 were significantly upregulated in tumor tissues compared to adjacent normal tissues, highlighting their potential as diagnostic biomarkers.ConclusionA strong correlation between TIMP1 and LINC01615 suggests their role in promoting GCa proliferation, invasion, and metastasis. TIMP1/LINC01615 axis regulates the EMT pathway and is a potential biomarker and therapeutic target in GCa.Supplementary InformationThe online version contains supplementary material available at 10.1186/s12575-025-00297-y.

Tumor recurrence driven by mitochondrial hypermetabolism remains a critical challenge in cancer therapy, as aberrant energy metabolism fuels therapeutic resistance and disease progression. We aimed to develop a multifunctional nanoplatform combining mitochondrial metabolism inhibition, photothermal therapy, and controlled chemotherapy to overcome tumor recurrence mechanisms. Biodegradable polydopamine nanoparticles (PDA-DOX-CO NPs) were engineered via molecular self-assembly, co-loading doxorubicin (DOX) and a carbon monoxide (CO) prodrug. The PDA-DOX-CO NPs demonstrated three synergistic therapeutic effects: (1) Photothermal ablation (48.38 °C tumor hyperthermia), (2) CO-mediated mitochondrial suppression, and (3) Spatiotemporally controlled DOX release. In HCT-116 tumor models, PDA-DOX-CO NPs with NIR irradiation induced 60% tumor complete ablation. Histopathological analysis confirmed significant apoptosis induction and mitochondrial morphology alterations in treated tumors. This “metabolic blockade + energy depletion + precision delivery” paradigm provides a synergistic solution to tumor recurrence, demonstrating enhanced therapeutic efficacy and biosafety through mitochondrial-targeted multimodal action.