Search Results (5,327)

Background/Objectives: Triple-negative breast cancer (TNBC) is frequently characterized by notably elevated Ki-67 expression, a hallmark of uncontrolled rapid cell-cycle progression. However, the underlying mechanisms remain unclear, leading to limited therapeutic options. Methods: In this study, hub gene was identified through integrated bioinformatic analysis of public datasets (TCGA-BRCA and METABRIC). Subsequent functional validation was performed both in vitro and in vivo using siRNA-mediated knockdown and small-molecule inhibitors. Phenotypic effects—including cell viability, cell cycle distribution, DNA synthesis, and clonogenic survival—were comprehensively assessed using MTT assays, flow cytometry, EdU, and colony formation assays. Protein-level changes were confirmed by Western blotting and immunohistochemistry (IHC). To dissect the transcriptional regulation of the key hub gene TTK, we first predicted potential upstream transcription factors using the JASPAR database; binding specificity was then validated through in silico motif analysis, luciferase reporter assays, and chromatin immunoprecipitation followed by quantitative PCR (ChIP-qPCR). Results: The mitotic kinase TTK is significantly overexpressed in TNBC compared with non-TNBC breast cancers. Notably, TTK overexpression exhibited a strong positive correlation with elevated Ki-67 indices and reduced overall survival in TNBC patients. Functional validation demonstrated that pharmacological or genetic inhibition of TTK effectively induced G2/M cell-cycle arrest and potently suppressed TNBC proliferation in both in vitro cell cultures and in vivo xenograft models. Mechanistically, TTK overexpression stems from enhanced transcriptional initiation driven by the transcription factor NFYA binding to the CCAAT box in the TTK promoter—an interaction newly identified here. Concurrently, TTK blockade disrupted spindle assembly checkpoint (SAC) signaling via BUB1B/MAD1L1 downregulation, triggering mitotic arrest and catastrophe. Conclusions: Collectively, these findings establish TTK as a key cell-cycle regulator driving TNBC proliferation. More importantly, targeting mitotic control through TTK inhibition represents an efficient strategy to impede the aberrantly fast cell cycle progression in TNBC. Full article

Endocrine therapy is the mainstay for estrogen receptor (ER) α-positive breast cancer (BC), yet many patients display acquired resistance. We then screened natural compounds using human ERα-positive BC cells and identified perillyl alcohol (POH), a monoterpene from perilla, that reduces ERα protein levels. Chemoproteome analysis using POH-immobilized nanomagnetic beads revealed adenine nucleotide translocase 2 (ANT2), a mitochondrial inner membrane protein, as a direct target of POH. Molecular dynamics (MD) simulations predicted POH binding to the central pore of ANT2, which functions in ATP transport. ANT2 depletion reduced ERα levels, and public datasets indicate that high ANT2 expression correlates with poor prognosis in ERα-positive BC. POH also inhibited the growth of Tamoxifen- and Fulvestrant-resistant BC cells. RNA sequencing showed that fatty acid elongation-related genes were upregulated in Fulvestrant-resistant cells but downregulated by ANT2 depletion. Both ANT2 depletion and POH treatment led to the accumulation of intracellular lipid droplets in Fulvestrant-resistant cells, consistent with impaired fatty acid elongation. Finally, in silico screening using MD simulations identified venetoclax and nystatin as potential ANT2 pore binders. Both compounds reduced ERα levels in ERα-positive BC cells and increased lipid droplet formation in Fulvestrant-resistant cells. These findings highlight ANT2 as a druggable target against endocrine-resistant BC. Full article

Pulmonary arterial hypertension (PAH) is a severe and progressive cardiopulmonary disorder with limited treatment options. Camellia petelotii (Merr.) Sealy (CP) contains multiple flavonoids and other phytochemicals, but its active compounds and molecular mechanisms in PAH remain unclear. Active compounds of CP were screened by comprehensive literature mining and absorption, distribution, metabolism, and excretion (ADME) evaluation. PAH-related hub targets were identified from transcriptomic data using weighted gene co-expression network analysis (WGCNA), machine learning, and external validation. Functional enrichment, immune infiltration, and single-cell RNA-sequencing analyses were performed to characterize their biological roles and cellular localization. Molecular docking and molecular dynamics simulations assessed compound–target interactions. The effects of CP were further evaluated in hypoxia-induced rat pulmonary artery smooth muscle cells (RPASMCs). Five core bioactive compounds were identified, among which luteolin and quercetin were prioritized for further analysis. HSP90AA1 and ROCK2 were screened as hub targets. Bioinformatic analyses suggested that these targets were mainly associated with the “Lipid and atherosclerosis” pathway, metabolic reprogramming, and modulation of the immune microenvironment. Single-cell analysis showed broad expression of HSP90AA1 and enrichment of ROCK2 in fibroblasts and endothelial cells. Molecular docking and molecular dynamics simulations supported stable binding of luteolin to HSP90AA1. In vitro, CP extract inhibited hypoxia-induced hyperproliferation of RPASMCs and reduced HSP90AA1 protein expression. HSP90AA1 may represent a candidate molecular mediator of CP in PAH, and CP inhibited hypoxia-induced RPASMC proliferation in association with downregulation of HSP90AA1. Full article

Background: Midkine (MDK), a secreted heparin-binding growth factor, is involved in tumor progression and metastasis. While serum MDK is widely recognized as a potential prognostic biomarker for colorectal cancer (CRC), its specific functional role and underlying mechanisms in CRC development are not fully understood. Methods: The four publicly available CRC microarray datasets—GSE41258, GSE44076, GSE81558, and GSE117606—along with TCGA-COAD and TCGA-READ datasets and their associated clinical data were obtained. MDK expression was measured at both the mRNA and protein levels using quantitative real-time PCR (qRT-PCR) and Western blotting. To investigate its oncogenic functions, a comprehensive set of assays was performed: transwell and wound healing assays for invasion and migration; CCK-8 and colony formation assays for proliferation; and tail vein/spleen injection models combined with xenograft models to study metastasis and tumor growth in vivo. To uncover underlying mechanisms, Western blotting was used to examine the involvement of epithelial–mesenchymal transition (EMT) and the PI3K/AKT signaling pathway. Results: MDK is significantly overexpressed in CRC tissues and cells compared to normal tissues and cells. Notably, patients with high MDK levels show poorer overall survival (OS). Overexpression of MDK increases CRC invasion, migration, proliferation, and metastasis both in vivo and in vitro, while its knockdown reverses these effects. Mechanistically, MDK activates the PI3K/AKT pathway, leading to increased AP2A1 expression and promotion of EMT in CRC. Conclusions: MDK promotes invasion, migration, proliferation, metastasis, and EMT in CRC cells through the PI3K/AKT pathway by inducing AP2A1 expression, which could serve as a diagnostic marker. The PI3K inhibitor LY294002 significantly reduces AP2A1 levels and inhibits MDK-induced malignant behaviors. Targeting MDK-related signaling pathways may offer new strategies for CRC treatment. Full article

The tomato leaf miner, Phthorimaea (Tuta) absoluta, Meyrick 1917 is recognized as a highly destructive pest, causing significant economic losses to crops in both greenhouse and open field environments across four continents: Asia, Africa, Europe, and South America. High genetic homogeneity among populations from various regions and countries indicates significant gene flow between P. absoluta populations, suggesting a lack of geographical barriers to dispersion. Furthermore, P. absoluta has developed resistance to insecticides due to target-site mutations or metabolic resistance, which enable the insect to withstand lethal doses of insecticides. To control this insect pest, the plant-mediated RNA interference (RNAi) is most promising host-induced gene silencing technique, utilized the plant’s machinery to express double-stranded (dsRNA), which triggers the RNAi pathway in P. absoluta. Due to thermal tolerance, the P. absoluta has increased its area of invasion by 600 km per year over 9 years. Female P. absoluta releases pheromones that are recognized by males with a sophisticated olfactory circuit on their antenna. Pheromone binding proteins (PBPs) play a crucial role in mate recognition and attraction, and their expression peaks during courtship, specifically around 6:00 a.m. Given its potential to significantly alter the insect genome, clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated protein 9 (Cas9) offer a revolutionary strategy to control P. absoluta. Furthermore, this pest has developed remarkable adaptations to survive on unfavorable hosts by secreting specific proteins from its salivary glands that detoxify plant defenses. Insecticide resistance is likely the cause of field control failures of P. absoluta. Biological control, sex pheromone traps, and cultural control are the most promising approaches to address insecticide resistance resulting from these failures. Therefore, the implementation of integrated control programs and appropriate resistance management strategies is necessary to keep P. absoluta infestations under economic damage thresholds. Full article

Salt stress is a major abiotic factor that significantly limits crop yields worldwide. Late embryogenesis abundant (LEA) proteins, which are widely present across diverse organisms, play critical and multifaceted roles in plant responses to abiotic stress. However, only a few salt tolerance-related HvLEA genes have been identified in barley. In this study, we characterized 107 HvLEA proteins in barley, which were classified into eight groups and found to be distributed across all seven chromosomes. RNA-Seq analysis of root and leaf tissues from the cultivar “Golden Promise” at 12, 48, and 120 h after salt stress treatment identified 69 differentially expressed HvLEA genes across both tissues. Among these, 41 HvLEA genes were commonly differentially expressed in leaves and roots. Six genes (HvDHN2, HvDHN5, HvDHN10, HvLEA1.1, HvLEA1.6, and HvSMP2) were extremely up-regulated after salt stress in both roots and leaves, with log2FC values exceeding 10, indicating their potential key roles in salt stress response. qPCR validation of selected genes confirmed expression trends consistent with the RNA-Seq data. Database predictions and co-expression network analysis suggested that, in addition to potential protein interactions within the same family, these genes may interact with partners such as cysteine-rich receptor kinases, zinc finger proteins, calcium-binding EF-hand family proteins, NAC domain-containing proteins, and glycosyltransferases. This study identified key HvLEA genes involved in salt stress response and provided valuable genetic resources for improving barley tolerance through molecular breeding. Full article

Background/Objectives: Bone-targeted drug delivery systems hold great promise for treating skeletal diseases, yet the optimal strategy for functionalizing lipid nanoparticles (LNPs) with bone-homing ligands remains insufficiently explored. Herein, we compared two alendronate sodium (Alen) modification approaches (pre-conjugation and post-conjugation) for constructing bone-targeted LNPs capable of delivering mRNA to skeletal tissues. Methods: LNPs were fabricated via microfluidic mixing, and the 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-polyethylene glycol-alendronate conjugate (DSPE-PEG-Alen) required for the pre-conjugation method was synthesized. The bone-targeting ability of LNPs prepared by the two Alen modification strategies was evaluated using an in vitro hydroxyapatite (HAP) binding assay. Furthermore, the physicochemical properties, bone-targeting performance, mRNA delivery efficiency, and biosafety of the LNPs prepared by the post-conjugation method were assessed through cellular uptake, in vivo imaging, and other methods. Results: Hydroxyapatite binding assays revealed that the post-conjugation strategy afforded significantly superior bone affinity compared to the pre-conjugation approach. In addition, ex vivo bone fragment binding experiments further confirmed that the bone-targeting LNPs prepared by the post-conjugation method exhibited stronger bone-binding capability compared to unmodified LNPs. The optimized Alen-LNPs demonstrated efficient cellular uptake and functional mRNA translation in bone marrow mesenchymal stem cells with negligible cytotoxicity. In vivo studies in mice confirmed the preferential accumulation of Alen-LNPs in bone tissues, with successful green fluorescent protein (GFP) mRNA translation detected in bone tissue sections. Histopathological analysis confirmed the biosafety of the formulation. Conclusions: This study establishes the post-conjugation strategy as the superior approach for Alen functionalization of LNPs, providing a robust and reproducible platform for bone-targeted mRNA therapeutics. Full article

The MADS-box family is a multifunctional family of transcription factors characterized by the presence of a unique MADS domain, which plays an important part in regulating essential biological processes, including metabolic synthesis and the stress response. In this review, we analyze the structural features and classification of MADS-box proteins, then summarize the functions of the MADS-box family in the stress response. The MADS-box family can directly regulate downstream functional genes by binding to the CArG-box in the promoters of target genes, thereby influencing growth, development, and stress responses. Also, MADS-box transcription factors can form protein complexes with both MADS-box proteins and other types of transcription factors and chromatin regulatory proteins to modulate the chromatin state or transcriptional activation. Furthermore, they can regulate plant physiological responses by facilitating the synthesis of essential signaling molecules, including hormones and non-coding RNA. Finally, we discuss the potential of the MADS-box family in crop molecular breeding, offering a novel approach for developing high-yield and stress-resistant cultivars for solving global food security and climate change challenges. Full article

Background: The immunologically cold nature and immunosuppressive tumor microenvironment (TME) of intrahepatic cholangiocarcinoma (ICC) contribute to its poor prognosis. This study aims to identify novel biomarkers related to prognosis and TME in ICC. Methods: We first identified the high expression of m6A reader insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) in ICC through bioinformatics screening. Subsequently, a retrospective study was conducted on 224 ICC patients who had undergone radical resection. The expression levels of IGF2BP2 and programmed death ligand 1 (PD-L1) were detected in a tissue microarray (TMA) using immunohistochemistry (IHC). The co-localization of IGF2BP2, PD-L1, programmed cell death protein 1 (PD-1), and CD8⁺T cells was evaluated by multiple immunofluorescence techniques. Results: IHC confirmed a significant upregulation of IGF2BP2 in tumor tissues compared with normal bile duct epithelia (p < 0.05). IGF2BP2 expression was positively correlated with PD-L1 expression (TPS R = 0.215, p = 0.016; CPS R = 0.295, p = 0.008). High IGF2BP2 expression was associated with increased PD-L1/PD-1 positivity and reduced CD8⁺T cell infiltration. Kaplan–Meier analysis revealed significantly worse 3-year overall survival (OS: 20.56% vs. 29.91%, p = 0.0291) and recurrence-free survival (RFS: 9.72% vs. 18.56%, p = 0.0372) in the IGF2BP2-high group. Multivariate analysis identified IGF2BP2 as an independent risk factor for both OS (HR = 1.683, p = 0.044) and RFS (HR = 1.946, p = 0.042). Conclusions: IGF2BP2, as a potential biomarker and independent prognostic factor for ICC, is associated with increased PD-L1 expression. Full article

Breast cancer (BRCA) is the most common cancer worldwide, with an incidence exceeding that of lung cancer. Protein lactylation, a newly identified post-translational modification involving the binding of lactic acid to lysine residues, plays an important role in BRCA. However, its role in BRCA progression remains largely unexplored. This study aims to identify and characterize the lactylation-related genes involved in BRCA biology. Transcriptomic and clinical data of BRCA and normal breast tissues were obtained from TCGA and GEO. Lactylation-related genes were curated from literature and intersected with BRCA datasets to identify candidates. A prognostic risk model was constructed using LASSO and Cox regression. Functional enrichment was performed using KEGG, GSVA, and GSEA. Immune correlations were evaluated by ESTIMATE, CIBERSORT. Single-cell RNA-seq data were integrated to assess gene expression heterogeneity across tumor and immune compartments. In vitro, MDA-MB-231 cells were treated with sodium L-lactate and lactylation-inducing agents, and gene expression was validated by Western blot and RT-qPCR, while EdU and wound healing assays evaluated proliferation and migration. We identified six hub genes associated with the immune microenvironment. Notably, S100A4 is significantly underexpressed, suggesting their potential regulatory roles in BRCA. Further analysis demonstrated that lactylation-related genes are closely linked to immune regulation in BRCA, indicating a possible crosstalk between metabolic modification and tumor immunity. Additionally, we found that lactylation significantly influences gene expression patterns and immune infiltration in BRCA. Importantly, lactic acid ions were shown to upregulate lactylation levels in BRCA cells, underscoring the functional impact of metabolic signals on post-translational modifications in tumorigenesis. Our findings indicate a potential mechanism wherein lactylation affects BRCA progression via lactic acid-driven regulation of the immune microenvironment; they also highlight the possible involvement of S100A4 in this process and offer new insights that could contribute to the diagnosis and treatment of BRCA. Full article

Fish oil (FO) has been shown to confer beneficial effects on hepatic diseases in both humans and animals. This study aimed to investigate whether dietary fish oil (FO) supplementation alleviates deoxynivalenol (DON)-induced liver injury by modulating the ferroptosis signaling pathway in weaned piglets. Twenty-four weaned piglets were allocated to a 2 × 2 factorial design, with the main factors consisting of dietary treatment (5% corn oil or 5% FO supplementation) and DON exposure (basal diet or diet contaminated with 4 mg/kg DON). After 21 days of dietary treatment, piglets were euthanized for collection of blood and liver samples. Dietary FO significantly attenuated DON-induced hepatic structural damage and inflammatory infiltration. Specifically, FO supplementation reduced the activities of aspartate transaminase (AST) and alkaline phosphatase (ALP), as well as the AST/alanine aminotransferase (ALT) ratio following DON exposure. Dietary FO also decreased malondialdehyde (MDA) concentrations in both the liver and serum, lowered hepatic 4-hydroxynonenal (4-HNE) level and Fe²⁺ content, and increased hepatic glutathione (GSH) content. Moreover, dietary FO ameliorated ultrastructural liver damage induced by DON. Furthermore, DON significantly downregulated the mRNA levels of multiple genes associated with iron metabolism and ferroptosis, including heat shock protein beta-1 (HSPB1), acyl-CoA synthetase long chain family member 4 (ACSL4), and arachidonate 15-lipoxygenase (ALOX15), and upregulated the mRNA levels of transferrin (TF), ferritin heavy chain (FTH), solute carrier family 7 member 11 (SLC7A11), and transferrin receptor 1 (TFR1). Dietary FO counteracted these alterations by decreasing the mRNA of SLC7A11, TFR1, FTH, and TF after DON exposure. Finally, FO significantly decreased the protein expression of SLC7A11, iron-responsive element-binding protein 2 (IREB2), and FHT1 and increased the GPX4 protein expression following DON exposure. These findings suggest that FO may ameliorate DON-induced liver injury in weaned piglets, possibly through suppressing the ferroptosis signaling pathway. Full article

This study systematically elucidated the developmental characteristics and molecular regulatory mechanisms of the testis during the critical period of sexual maturation in Kazakh horses by combining histological observation of one- and two-year-old testicular tissues with transcriptomic sequencing. In the testes of one-year-old horses, no obvious lumen was observed, and the interior is mainly comprising supporting cells and spermatogonia on the basement membrane; in contrast, in the testes of two-year-old horses, the tubular lumen was complete with spermatogonia, spermatocytes, and spermatozoa, indicating that spermatogenic function had approached maturity. Transcriptome profiling identified 979 differentially expressed genes (DEGs), with 209 up-regulated genes, including CYP11A1 and CATSPER2, and 770 down-regulated genes, including CD9. Gene Ontology (GO) annotation indicated primary enrichment of DEGs in biological processes related to multicellular organism development, cell membrane composition, and ion binding. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis showed significant enrichment of DEGs in the calcium signaling pathway, cell adhesion molecules, and neuroactive ligand–receptor interaction, among other key pathways. Protein–protein interaction (PPI) network analysis further highlighted core genes, including TNF, CATSPER2, and CDH13. Validation by RT-qPCR confirmed the reliability of the RNA-Seq data. Our findings reveal the dynamics of testicular development in Kazakh horses through histological and molecular analyses, thereby providing a theoretical framework and candidate genes to further elucidate regulatory mechanisms and guide genetic improvement in reproductive traits. Full article

Extracellular vesicles (EVs) are increasingly explored as vehicles for intercellular communication and the delivery of functional molecules, such as RNA. Recent studies have identified transcript-intrinsic features that influence EV-RNA sorting, including sequence length/complexity and coding probability. However, predicting the enrichment of coding transcripts into EVs remains exploratory. Using the workflow we previously described for the characterization of CD81 fusion constructs, we measured the vesicular distribution of recombinant transcripts transiently expressed in HEK293T cells, yielding protein cargo that mirrored the intracellular abundance. We included the CD81Δ and E7 constructs to obtain insights into the potential correlation between transcript length and EV distribution. We observed that EV-RNA levels do not scale proportionally with intracellular abundance, unlike the corresponding protein cargo. We consider cumulative RNA structure, sub-cellular dynamics, post-transcriptional modifications, and RNA-binding protein interactions as necessary factors that may dictate mRNA recruitment into EVs independent of transcript length, possibly inspiring new trajectories to maximize EV-RNA loading strategies. Full article

Sterol Carrier Protein X (SCP-x) is an evolutionarily conserved lipid transport protein that plays important roles in sterol metabolism. In insects, cholesterol is an essential component of cellular membranes and the precursor of ecdysteroids, yet insects cannot synthesize cholesterol de novo and must obtain it from dietary sources. However, the functional role of SCP-x in cholesterol absorption and transport in insects remains poorly understood. In this study, the SCP-x gene from the migratory locust Locusta migratoria was identified and characterized using transcriptomic data from the midgut and fat body. The full-length LmSCP-x encodes a 404-amino-acid protein containing both the 3-oxoacyl-CoA thiolase domain and the sterol carrier protein-2 domain. Expression analysis revealed that LmSCP-x is predominantly expressed in the midgut and fat body, and subcellular localization experiments showed that the protein is mainly distributed in the cytoplasm. RNA interference-mediated knockdown of LmSCP-x significantly reduced cholesterol levels in the fat body and delayed nymphal development. Structural prediction using AlphaFold 3 further revealed a conserved three-dimensional structure of the SCP-2 domain, and molecular docking identified key amino acid residues involved in cholesterol binding, which were subsequently validated by bio-layer interferometry assays. Together, these results demonstrate that LmSCP-x plays a crucial role in cholesterol transport in L. migratoria and provide new insights into sterol metabolism in insects, offering potential targets for the development of novel pest management strategies. Full article

Gallbladder cancer (GBC) is an aggressive malignancy with limited treatment options and poor clinical outcomes. Identifying novel molecular targets is critical for improving therapeutic strategies. Sorcin (SRI), a calcium-binding protein implicated in tumor progression, has not been comprehensively investigated in GBC. SRI expression was analyzed by immunohistochemistry (IHC) in a large cohort of gallstone disease (GSD) controls (n = 85) and GBC tissues (n = 85). Functional assays, including cell proliferation, wound healing, transwell invasion, and Western blot analyses of epithelial–mesenchymal transition (EMT) markers, were performed in the NOZ GBC cell line following siRNA-mediated SRI knockdown. IHC revealed that 67% of GBC cases exhibited positive staining whereas all the GSD cases exhibited negative staining of SRI, demonstrating a significant upregulation of SRI in GBC (p < 0.001). SRI knockdown resulted in reduced proliferative capacity and markedly impaired migration and invasion. Further, SRI knockdown decreased vimentin levels, indicating suppression of EMT. SRI is significantly overexpressed in GBC and promotes key oncogenic traits, including proliferation, migration, invasion, and EMT. These findings highlight SRI as a potential therapeutic target in GBC. Further validation in animal models may facilitate translation into clinical applications. Full article