During early brain development, the nervous system evolves as cells connect to form a unique neural network, with communication between cell populations vital for neurological balance. This study investigates how the loss of PD-1 in myeloid cells disrupts nervous system development. Specific ablation of PD-1 affects myeloid cell proliferation and classification. As astrogenesis begins, astrocyte proliferation ceases, continuous astrocyte proliferation is observed. Immunofluorescence staining revealed high expression of astrocyte-related genes in PD-1f/f; LysM-Cre mice, which also exhibited more extroverted behaviour. Additionally, the absence of PD-1 enhances CXCL1 expression through the NF-κB pathway, promoting astrocyte proliferation by interacting with CXCR2. These findings underscore PD-1's regulatory role in myeloid cells and its implications for the myeloid-brain axis.
Benign prostatic hyperplasia (BPH) is a common condition in older men, with its prevalence increasing as age advances. Chronic inflammation orchestrates oxidative stress to exacerbate BPH. YAP1, which regulates organ size, cellular homeostasis, and tissue fibrosis, can be activated by ROCK1. Given the urgent clinical need for more effective therapies, this study explored whether targeting the ROCK1/YAP1 axis could mitigate BPH progression. Here, rats received in situ adeno-associated virus (AAV) injection to induce prostate-specific YAP1 overexpression. An inflammation-associated experimental autoimmune prostatitis (EAP) model was established by prostate antigen immunisation, followed by treatment with ROCK1 inhibitor fasudil and YAP1 inhibitor verteporfin. Cell models were treated with specific inhibitors to confirm the critical role of YAP1 in modulating mitochondrial function. As a result, YAP1 overexpression was sufficient to induce a pathological BPH phenotype. Specifically, YAP1 activated the inflammatory cascade to provoke an immune response, disrupted proliferation/apoptosis balance to induce tissue hyperplasia, triggered epithelial-mesenchymal transition (EMT) and reactive stroma to drive fibrosis, and promoted NOX4/ROS generation and antioxidant depletion to cause oxidative stress. The inflammation-induced experimental autoimmune prostatitis (EAP) model also presented analogous BPH lesions, which were significantly alleviated when treated with ROCK1 inhibitor fasudil and YAP1 inhibitor verteporfin. Mechanistically, YAP1 activation under inflammatory conditions suppressed SIRT1 expression, thereby exacerbating oxidative stress through the disruption of DRP1/MFN2-mediated mitochondrial dynamics. Overall, inflammation-driven activation of the ROCK1/YAP1 axis aggravates oxidative stress, promoting BPH hyperplasia and fibrosis by impairing SIRT1-regulated mitochondrial dynamics. These findings provide a preclinical rationale for developing ROCK1 or YAP1 inhibitors as targeted therapies for BPH patients with chronic inflammation.
Melanoma is the most serious type of skin cancer. About half of all melanomas have activating BRAF mutations. Targeted therapy for malignant melanoma with BRAF inhibitor (BRAFi) or its combination with MEK inhibitor (MEKi) improves the clinical outcomes of patients, but resistance develops invariably. The underlying mechanisms remain incompletely understood. Here, we show that caveolae number is increased in both BRAFi and BRAFi + MEKi-resistant melanoma cells, and the expression of the critical caveolae component PTRF is significantly upregulated in drug-resistant melanoma cell lines and tumour tissues. Knockdown of PTRF in drug-resistant cells reduces proliferation with increased apoptosis, whereas ectopic expression of PTRF confers resistance on parental cells to BRAFi or BRAFi + MEKi. On the contrary, the knockdown of PTRF in parental cells reduces their ability to acquire drug resistance induced by BRAFi treatment. Interestingly, we find that the expression of EGFR is increased along with PTRF and caveolin-1 in drug-resistant cells and in PTRF transduced parental cells, whereas knockdown of PTRF results in down-regulation of EGFR expression and attenuates drug resistance of parental cells induced by PTRF expression. Together, these results suggest that PTRF contributes to therapy resistance through upregulating EGFR in melanoma cells.
Micropeptides are endogenous peptides translated from alternative open reading frames (alt-ORFs) within coding or non-coding genes. Emerging evidence suggests that some micropeptides play critical roles in both physiological and pathological processes. Multiple myeloma (MM), a haematological malignancy, remains incurable due to frequent relapses and a limited understanding of its underlying mechanisms. In this study, we sought to investigate the function and molecular mechanism of a novel micropeptide in MM pathogenesis. We identified a novel micropeptide, altH19, encoded by the lncRNA H19, which is highly expressed in patients of MM. Functional assays revealed that altH19 promotes myeloma cell proliferation and colony formation significantly. Furthermore, altH19 induces multipolar mitosis by upregulating the expression of Aurora B, Centrin 2 and phosphorylated histone H3. Flow cytometry analyses confirmed that overexpression of altH19 enhances DNA replication and accelerates the transition from early to mid-late stages of the DNA replication process. Conversely, knockout of altH19 reverses these effects. Mechanistically, altH19 directly interacts with phosphorylated CDK2 at threonine 160, thereby enhancing CDK2 T160 phosphorylation and activating the downstream E2F1 target RB phosphorylation. Notably, altH19 was able to restore phosphorylation levels of CDK2 and RB that were otherwise suppressed by the CDK2-selective inhibitor Seliciclib. In summary, we identify altH19 as a novel lncRNA-derived micropeptide with a pivotal role in myeloma progression, highlighting the therapeutic potential of targeting the altH19-CDK2-RB axis in MM treatment.
Acute monocytic leukaemia, a subtype of acute myeloid leukaemia (AML), is a highly aggressive malignancy characterised by a poor prognosis, primarily due to the ability of leukaemic cells to evade immune surveillance. In this study, we demonstrate that homoharringtonine (HHT), an FDA-approved therapeutic agent for chronic myeloid leukaemia (CML), inhibits this immune evasion by targeting the FTO/m6A/LILRB4 signalling pathway in monocytic AML. Utilising RNA sequencing (RNA-seq) and various functional assays, we reveal that HHT treatment significantly reduces LILRB4 expression at both the RNA and protein levels, suggesting that the effects of HHT on LILRB4 are distinct from its well-established role as a protein synthesis inhibitor. Mechanistically, HHT treatment markedly increases global levels of RNA m6A in THP-1 cells by promoting the degradation of FTO, which subsequently diminishes the expression of its downstream targets, MLL1 and LILRB4. Furthermore, in vitro and in vivo analyses employing monocytic AML cell lines, mouse-derived AML xenograft models, and patient samples collectively support the conclusion that HHT suppresses immune evasion in monocytic AML by reducing LILRB4 expression. Importantly, the downregulation of LILRB4 resulting from HHT treatment enhances the susceptibility of THP-1 cells to CD8+ T cell cytotoxicity, accompanied by increased markers of immune activation. Overall, our findings position HHT as a promising clinical agent for enhancing CD8+ T cell-based cancer immunotherapy by mitigating immune evasion in monocytic AML.
Mitochondrial stress-induced mitophagy plays a critical role to maintain cellular homeostasis; however, in cancer cells, this process may also contribute to drug resistance. Our previous work identified CDK12 as a critical regulator of prostate cancer (PCa) cell survival under sustained enzalutamide exposure, though the precise mechanism remains to be elucidated. In this study, we hypothesize that CDK12 plays a key role in mitophagy regulation under mitochondrial stress, potentially modulating PCa cell resistance to enzalutamide, the first-line clinical medication in PCa therapy. Utilising multiple in vitro PCa cell models, we demonstrate that both CDK12 knockdown and pharmacological inhibition with THZ531 impaired mitophagy following treatment with enzalutamide and mitophagy inducer CCCP. Mechanistically, our finding reveal that CDK12 inhibition disrupts FOXO3-induced BNIP3 transcription, thereby preventing receptor-mediated mitophagy and sensitising PCa cells to enzalutamide. This study identifies the CDK12-FOXO3-BNIP3 pathway as a novel regulatory mechanism governing mitophagy under mitochondrial stress. Importantly, these results underscore CDK12's role in preserving mitochondrial function and promoting PCa cell survival during enzalutamide treatment. These findings highlight the therapeutic potential of targeting the CDK12-BNIP3-mitophagy axis in combination with antiandrogen therapies, offering a promising strategy to overcome drug resistance in PCa and improve clinical outcomes.
Understanding the structural and functional organisation of brain networks is a fundamental objective in neuroscience, with three-dimensional (3D) reconstruction of single-neuron morphology serving as a critical foundation. The Golgi staining method, which enables random neuronal labeling and provides high-contrast signals in both optical and X-ray microscopy, remains a valuable tool for morphological analysis. However, its widespread application in large-scale neuronal reconstructions is hindered by signal discontinuities in neuronal branches, high-density labeling, and complex background interference. While automated reconstruction methods perform well in sparsely labelled and morphologically simple neuronal populations, their effectiveness is limited in Golgi-stained samples. Here we develop a semi-automated single-neuron reconstruction method for Golgi-stained mouse brain neurons (SNR-Golgi). By integrating three key technical modules—background denoising, single-neuron extraction, and branch repair—SNR-Golgi significantly enhances the accuracy and completeness of neuronal reconstruction. In fluorescence micro-optical sectioning tomography (fMOST) datasets, SNR-Golgi demonstrated superior performance in neuronal reconstruction within the mouse somatosensory cortex, achieving a 30% increase in reconstructed branch count, a 76% improvement in total branch length, and a 3.7-fold increase in axonal length. Additionally, in synchrotron-based X-ray imaging datasets, SNR-Golgi enabled submicron-resolution 3D reconstruction of single neurons. These results demonstrate that SNR-Golgi effectively addresses the complexity of Golgi-stained samples and provides robust technical support for the structural analysis of brain neurons across various imaging modalities.
Traumatic brain injury (TBI) represents a global health burden, often resulting in persistent neurological deficits due to impaired hippocampal neurogenesis. Nevertheless, the temporal progression of post-TBI neurogenesis and its molecular mechanisms remain elusive. To investigate the mechanism of impaired hippocampal neurogenesis and neurological deficits following TBI. Single-cell RNA sequencing (scRNA-seq) was employed to explore the mechanism of abnormal hippocampal neurogenesis after TBI in mice. Antagonists and conditional gene knockout (CKO) strategies were applied to dissect the molecular function of target genes. Here, we found that neural stem cells (NSCs) were hyperactivated as observed in Nestin-GFP reporter mice in hippocampus during the early phases of TBI, followed by progressive depletion of the NSC pool, impaired neurogenesis, and the onset of progressive cognitive dysfunction. ScRNA-seq transcriptomic analysis revealed sustained upregulation of Rho-associated coiled-coil protein kinase 1 (ROCK1) in hippocampal NSCs post-TBI. Pharmacological inhibition of ROCK1 or ROCK1 CKO rescued chronic neurogenic deficits and improved cognitive functions in TBI mice. Mechanistically, ROCK1 dysregulation impaired neurogenesis via aberrant AKT hyperphosphorylation, establishing a unidirectional ROCK1-AKT signalling axis in adult hippocampal neurogenesis. Our findings position ROCK1 as a pivotal regulator of the post-TBI NSC pool hyperactivation and aberrant neurogenesis and propose targeted kinase inhibition strategies as a potential therapy to mitigate abnormal neurogenesis in TBI patients.
Amelogenesis imperfecta type 1G (AI1G), also known as Enamel-Renal-Gingival Syndrome (ERGS), is an autosomal recessive disorder caused by variants in FAM20A, encoding a Golgi apparatus protein crucial for protein processing and secretion. AI1G presents with enamel defects, nephrocalcinosis and gingival overgrowth. Building upon our previous findings demonstrating the impact of FAM20A insufficiency on deciduous dental pulp cells, this study investigated the molecular mechanisms underlying gingival fibromatosis in AI1G. RNA sequencing of gingival fibroblasts from an AI1G patient revealed widespread differential gene expression (DEG). Gene Ontology (GO) analysis demonstrated enrichment of DEGs in biological processes related to cell adhesion, differentiation, proliferation (including positive regulation and cell division), cell cycle regulation, apoptosis and signal transduction. Pathway analysis (Reactome and KEGG) further highlighted the dysregulation of signalling pathways, including Wnt, TGF-β, cell cycle, DNA replication, Rho GTPase signalling and extracellular matrix organisation. Functional assays confirmed these findings, revealing delayed initial attachment and spreading, impaired osteogenic differentiation (evidenced by reduced mineralization and downregulation of DLX5, OCN, RUNX2 and OPN), enhanced cell cycle progression and proliferation (increased colony size and proliferation rates, along with a shift from G0/G1 to G2/M phase) and suppressed apoptosis in FAM20A-insufficient fibroblasts. These results suggest that FAM20A plays a critical role in regulating fundamental processes in gingival fibroblasts, and its insufficiency contributes to the gingival fibromatosis phenotype observed in AI1G through the disruption of cell adhesion, differentiation, proliferation and apoptosis. This study proposes novel insights into the pathogenesis of AI1G and highlights potential therapeutic targets for this complex disorder.
Breast cancer remains the most prevalent malignancy among women, and radiotherapy plays a pivotal role in reducing local recurrence and improving prognosis. However, the emergence of radioresistance in a subset of patients significantly compromises treatment efficacy, underscoring the need for a deeper understanding of the underlying molecular mechanisms. In recent years, non-coding RNAs (ncRNAs) have emerged as key regulators of gene expression and have garnered increasing attention for their roles in mediating radioresistance in breast cancer. This review systematically summarises the major molecular mechanisms by which ncRNAs contribute to breast cancer radioresistance, including cell cycle regulation, DNA damage repair, programmed cell death (e.g., apoptosis, autophagy and ferroptosis), oxidative stress response, tumour microenvironment remodelling and maintenance of cancer stem cell properties. On the translational front, RNA-based therapeutic approaches—including antisense oligonucleotides (ASOs), small interfering RNAs (siRNAs), miRNA mimics and CRISPR/Cas9—offer promising avenues for radiosensitisation, yet face substantial clinical hurdles. These include immune activation, poor delivery specificity, intracellular trafficking barriers and limited stability. Advances in chemical modifications and nanoparticle-based delivery systems—such as redox-responsive nanocarriers—have shown potential in enhancing the efficacy and safety of ncRNA-targeted therapies. Despite encouraging progress, clinical translation remains constrained by a lack of methodological standardisation, insufficient high-quality clinical data, limited biomarker reliability, suboptimal target selection and unresolved safety concerns. Future efforts should prioritise optimisation of delivery platforms, validation of multi-ncRNA biomarker panels in large, multicentre cohorts and integration of multi-omics data to reconstruct comprehensive regulatory networks, ultimately accelerating the clinical deployment of ncRNA-based radiosensitisation strategies.
The malignant transformation of cancer cells is underpinned by the dysregulation of essential cellular processes, including genome stability maintenance, DNA repair, transcriptional control and signal transduction. These processes are not randomly distributed but are spatiotemporally coordinated through dynamic molecular assemblies. Recent advances have highlighted the pivotal role of biomolecular condensates, membraneless compartments formed via liquid–liquid phase separation (LLPS), in compartmentalising and regulating these key functions. LLPS enables the concentration and organisation of proteins and nucleic acids, creating distinct biochemical environments that facilitate cellular decision-making. Importantly, aberrant phase separation has been increasingly implicated in the acquisition of cancer hallmarks, such as sustained proliferative signalling, resistance to cell death and immune evasion. In this review, we summarise the physicochemical principles of LLPS, examine its emerging roles in oncogenic transformation and discuss the therapeutic potential of targeting phase separation in cancer. Our findings highlight LLPS as a novel and versatile regulatory layer in tumour biology and an emerging frontier in precision oncology.
Pulmonary vascular endothelial cells (VECs) are essential for the normal function of the lung, through maintaining vascular barrier integrity, regulating blood flow, and participating in inflammatory responses to safeguard oxygen exchange and physiological homeostasis. The occurrence and development of various pulmonary diseases all take the injury of pulmonary VECs as an important pathological hub, which directly affects the therapeutic effect and prognosis recovery of patients. The injury mechanisms of pulmonary VECs present multi-dimensional network characteristics, involving inflammation and oxidative stress, genetic factors, cellular senescence, metabolic abnormalities, and immune dysregulation. Due to the unique physiological structure of the lungs, traditional drugs often encounter significant challenges in clinical application such as insufficient targeting, low bioavailability, and systemic side effects. In order to overcome the existing treatment bottlenecks, it is crucial to implement an in-depth analysis of the molecular mechanism of pulmonary VECs injury. This review systematically explores the mechanisms of pulmonary VECs injury, evaluates novel therapeutic strategies targeting pulmonary VECs' dysfunction, and discusses the challenges and future prospects of clinical translation. The goal is to shift pulmonary diseases treatment from symptom management to precise molecular intervention.