Biology Direct最新文献

Background: Iron overload and ferroptosis are associated with intestinal ischemia and reperfusion (II/R)-induced acute lung injury (ALI). However, the mechanisms underlying the regulation of iron homeostasis remain unclear. Nrf2 regulates cellular iron homeostasis; nonetheless, its impact on ALI pathology and its underlying mechanism of action requires further investigation. Ubiquitin ligase E3B (UBE3B) plays a critical role in the proteasome pathway, which is essential for protein turnover and ubiquitin-mediated signaling. A recent study found that UBE3B plays a role in oxidative stress; nevertheless, it remains unknown whether its role is related to Nrf2. Furthermore, the exact role of UBE3B in ALI and its underlying mechanism remain largely uncharacterized.

Methods and results: In the present study, immunohistochemical analysis of UBE3B expression in type II alveolar epithelial cells (AECII) was conducted, and its expression was found to be decreased in II/R-ALI. Western blot analysis indicated that UBE3B hypoactivation may aggravate oxidative stress, thereby promoting ALI. Moreover, UBE3B was involved in iron metabolism dysfunction and ferroptosis. UBE3B deficiency enhanced the process of nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy and increased ferrous ion content, whereas overexpression of UBE3B reversed the harmful effects of Nrf2 knockdown on AECII, which may promote AECII ferroptosis.

Conclusions: This study highlights the role of the Nrf2/UBE3B/NCOA4 axis in AECII ferroptosis and II/R-ALI pathogenesis, suggesting that Nrf2 activation may be a promising strategy for ALI treatment.

Doxorubicin (DOX) is a potent chemotherapeutic widely used against various cancers, but its clinical application is limited by DOX-induced cardiotoxicity (DIC). This study explored the cardioprotective potential of extracellular vesicle-enriched secretome derived from adipose stem cells (EVS_ASC) in mitigating DOX-induced apoptosis in cardiomyocytes. Adipose-derived stem cells were cultured, and their conditioned medium and extraceullular vesicles were isolated and characterized according to the Minimal Information for Studies of Extracellular Vesicles 2023 guidelines. HL-1 cardiomyocytes were pretreated with EVS_ASC before exposure to 1 µM DOX. Cell viability was assessed via the cell counting kit-8 assay, while apoptosis markers and survival mediators were evaluated through Western blotting. RNA sequencing identified differentially expressed genes, including clusterin (Clu), which was further quantified using an enzyme-linked immunosorbent assay. The functional role of clusterin was validated through siRNA-mediated knockdown. EVS_ASC significantly improved cell viability in DOX-exposed cardiomyocytes and reduced the cleaved caspase-3 to procaspase-3 ratio. Clusterin expression was highest in EVS_ASC-treated cells, and its knockdown markedly increased caspase-3 cleavage, confirming its pivotal role in cardioprotection. Moreover, EVS_ASC enhanced the phosphorylation of AKT, Bcl2-associated agonist of cell death, and glycogen synthase kinase-3β, implicating PI3K/AKT pathway activation in clusterin upregulation and anti-apoptotic effects. These findings demonstrate that EVS_ASC mitigates DOX-induced apoptosis in cardiomyocytes through clusterin upregulation and PI3K/AKT pathway activation. Clusterin is identified as a potential biomarker for evaluating EVS_ASC efficacy. While EVS_ASC shows promise as a cardioprotective strategy against DIC, further studies are needed to optimize its therapeutic safety by addressing potential oncogenic risks.

Background: Idiopathic pulmonary fibrosis (IPF) is a chronic progressive pulmonary disease characterized by alveolar structural destruction and fibrosis. In recent years, efferocytosis has been recognized as playing a crucial role in the occurrence and progression of IPF. This study aimed to identify and regulate key efferocytosis-related genes to elucidate their potential roles and clinical significance in IPF.

Methods: IPF-related datasets (GSE32537) were obtained from the Gene Expression Omnibus (GEO) database. Differential gene expression analysis and weighted gene coexpression network analysis (WGCNA) were applied to identify key genes associated with IPF, intersecting them with efferocytosis-related genes (ERGs) to obtain IPF-ERGs. Protein‒protein interaction (PPI) network construction and enrichment analysis were performed to elucidate the potential functions of these genes in IPF. Seven machine learning algorithms were employed to screen for hub genes with high diagnostic value. The GSE70866 dataset was used for validation, and a nomogram was constructed. Additionally, the CIBERSORT algorithm was used to analyze immune infiltration levels, and transcriptomic validation of the hub genes was conducted in animal experiments.

Results: A total of 21 IPF-ERGs were identified, and machine learning further identified TLR2, ATG7, SPHK1, and ICAM1 as hub genes, which were significantly upregulated in the IPF group. Immune infiltration analysis revealed a significant increase in the infiltration levels of immune cell subsets, including memory B cells, CD8 + T cells, and resting dendritic cells, in the IPF group. Further clinical correlation analysis revealed a strong association between the expression levels of the hub genes and pulmonary function. A nomogram was constructed on the basis of the hub genes and validated for its potential clinical application. Consensus clustering classified IPF patients into two subtypes: C1, which was primarily by metabolic pathway activation, and C2, which was enriched in inflammatory and immune pathways. Transcriptomic analysis of animal experiments also confirmed the upregulation of hub gene expression in IPF.

Conclusion: This study identified TLR2, ATG7, SPHK1, and ICAM1 as four key hub genes, revealing their potential diagnostic value and biological functions in IPF. These genes may serve as potential diagnostic biomarkers and therapeutic targets, providing new insights for precision treatment.

Clinical trial number: Not applicable.

Background: Oxidative stress plays a crucial role in the development of diabetic foot ulcers (DFU). However, its underlying mechanisms are not fully understood. The purpose of this study was to use bioinformatics and preliminary validation methods to preliminarily reveal the oxidative stress landscape in DFU.

Methods: Based on the single-cell and bulk RNA sequencing data of DFU, we conducted differential genes screening, machine learning, PPI network construction, immune infiltration analysis, drug prediction, TF-mRNA-miRNA network, cell-cell interaction, pseudotime trajectory analysis, external cohort validation, and in vitro experiments to develop the oxidative stress landscape in DFU.

Results: Bulk RNA-seq analysis identified 63 oxidative stress-related genes of DFU (DORGs), and the top 59 genes were screened out for key nodes with close functional associations. Functional enrichment analysis showed significant involvement in oxidative stress response. Drug prediction highlighted Thymoquinone and Erlotinib as potential therapeutic candidates. Machine learning algorithms (SVM-RFE, LASSO and RF) identified BCL2 and FOXP2 as candidate hub DORGs for DFU diagnosis. Immune cell infiltration analysis indicated a significant presence of naive B cells and CD8 T cells in DFU. The analysis of single-cell RNA sequencing identified a total of 31,787 cells across 10 distinct clusters, with a notably lower proportion of fibroblasts in DFU group than that in the control group. The expression patterns of BCL2 and FOXP2 across the different groups were consistent with findings from bulk RNA sequencing analysis. Notably, fibroblasts derived from DFU patients exhibited the highest oxidative stress scores. Intercellular signaling analysis indicated that fibroblasts serve as crucial communication cells, primarily engaged in COLLAGEN signaling network. Additionally, fibroblasts are categorized into five distinct clusters. Among these, COL6A5+ fibroblasts constitute the predominant cluster in DFU and exhibit low differentiation potential. Furthermore, in vitro experiments successfully established a DFU oxidative stress model of fibroblasts, revealing reduced migration ability in the absence of cell death. Both in vitro findings and external data corroborated the decreased expression levels of BCL2andFOXP2in DFU.

Conclusion: The oxidative stress-related genes BCL2 and FOXP2 could serve as diagnostic markers for DFU. Furthermore, we identified the novel pathogenic mechanism associated with oxidative stress in DFU fibroblasts. This study may offer new insights for the diagnosis and treatment of DFU.

Background: Cancer-associated fibroblasts (CAFs) have been reported to play a significant role in the development and metastasis of various tumors; however, research on their role in promoting prostate cancer (PCa) metastasis under castration conditions remains unclear.

Methods: In this study, we utilized quantitative reverse transcription polymerase chain reaction (qRT-PCR) to detect the expression differences of microRNA-196b-5p (miR-196b-5p) in the exosomes secreted by CAFs before and after castration. We further characterized the transcriptional regulatory landscape through RNA sequencing combined with bioinformatics databases. In vitro and in vivo experiments were conducted to determine the role of miR-196b-5p in promoting tumor migration and metastasis. The dual-luciferase reporter assay, RT-PCR analysis, and Western blot analysis confirmed that miR-196b-5p targets HOXC8 in prostate cancer. Additionally, transwell assays and Western blot analysis were performed to elucidate the role and specific mechanisms of HOXC8 in tumor metastasis.

Results: By analyzing the expression differences of miRNAs in the exosomes secreted by CAFs before and after castration, along with relevant data from databases, we found that miR-196b-5p is highly secreted by CAFs after castration. miR-196b-5p promotes the migration and metastasis of prostate cancer cells. Subsequently, through RNA sequencing analysis and experimental validation, we determined that miR-196b-5p targets HOXC8. This interaction activates the NF-κB pathway, leading to the upregulation of epithelial-mesenchymal transition (EMT)-related protein expression, thereby driving the metastasis of prostate cancer.

Conclusions: Our study elucidates a specific mechanism by which CAF-derived exosomes promote prostate cancer metastasis via miR-196b-5p regulation, contributing to the identification of therapeutic targets for managing tumor metastasis following castration.

Background: Non-small-cell lung cancer (NSCLC) exhibits variable outcomes and remains a leading cause of cancer-related mortality, despite advances in immunotherapy. This study aimed to develop a prognostic model using real-world data (RWD) to stratify patients by survival outcomes and evaluate the benefit of immunotherapy across risk groups.

Methods: A retrospective cohort of 270 patients with NSCLC (2015-2024) treated with chemotherapy alone (54%) or chemoimmunotherapy (46%) was analyzed. Clinical, laboratory (neutrophil-to-lymphocyte ratio [NLR], platelet-to-lymphocyte ratio [PLR], monocyte-to-lymphocyte ratio [MLR]), and histopathological data were collected. Multivariate Cox regression identified prognostic factors for overall survival (OS) and validated them via bootstrapping.

Results: The cohort (median age, 65; 78% male) had a median OS of 11.2 months and a median progression-free survival (PFS) of 7.7 months. The final prognostic model incorporated histology (adenocarcinoma vs. large cell/squamous cell carcinoma/rare subtypes: HR = 1.6-2.03), recurrence state (HR = 0.51), and NLR (HR = 1.13). Patients were stratified into low- (median OS = 14.6 months) and high-risk (median OS = 9.6 months; p < 0.001) groups. Immunotherapy significantly increased PFS in low-risk patients (12.2 vs. 7.1 months, p = 0.002) and showed an increasing trend in OS (16.9 vs. 11.3 months, p = 0.12). High-risk patients derived no OS/PFS benefit (p ≥ 0.56).

Conclusion: This RWD-derived prognostic model effectively stratifies NSCLC patients into distinct risk groups. Immunotherapy-chemotherapy provided meaningful PFS improvement in low-risk patients but minimal benefit in high-risk subgroups, underscoring the need for tailored therapeutic strategies.

The prognosis for patients with head and neck cancer (HNC) is usually poor, highlighting the need for new therapeutic strategies. To this end, this study aims to evaluate the antitumor efficacy of a combined treatment with low doses of different molecular targeted drugs, i.e. Y15, a FAK inhibitor, Afatinib (AFA) an ErbB inhibitor and TP-0903, an Axl inhibitor, on HNC. Human cell lines from salivary gland, tongue and pharynx HNC, cultured in 2D and 3D (spheroids) conditions, were used to evaluate the antitumor effects of Y15, AFA and TP-0903, alone or in combination. Cell survival, death and migration were evaluated. Western blotting and immunofluorescence analysis were performed to investigate the expression and activation of proteins involved in signal transduction and epithelial to mesenchymal transition. The combined treatment with low doses of Y15, AFA and TP-0903, was more effective than the individual and dual drug treatments in reducing survival, increasing cell death and reducing migration of HNC cells. The three inhibitors in combination had a synergistic effect in reducing survival of HNC cell lines in both 2D and 3D conditions. Moreover, as compared to the individual inhibitors and their pairwise combinations, the triple drug combination was the only able to simultaneously downregulate Axl, FAK, and N-cadherin while upregulating E-cadherin expression levels. The results reported herein provide compelling preliminary evidence supporting the combined use of Y15, AFA and TP-0903 as a novel therapeutic strategy for HNCs.