This multi-source bibliometric and translational mapping study provides a panoramic synthesis of how research on microglia-mediated spinal pain signaling has evolved from foundational mechanistic studies to clinically oriented innovations. The aim is to identify developmental trajectories, mechanistic hotspots, and translational opportunities, thereby offering strategic insight into guiding the future direction of neuropathic pain research. We analyzed 1313 original research papers from the Web of Science Core Collection (WoSCC; 2005-2024) using CiteSpace and VOSviewer to construct collaboration networks, journal co-citation graphs, and keyword-driven mechanism clustering. To add a translational medicine dimension, we conducted a targeted PubMed search ("microglia AND spinal cord AND (translational OR therapeutic OR drug targets)"), retrieving 692 additional records, enabling cross-database overlay to link mechanistic themes with specific therapeutic targets. The scientometric model indicates that spinal pain research has shifted from primarily descriptive work to more detailed regulatory models. Key themes include glial cell activation, oxidative stress, mitochondrial dysfunction, and changes in microglia state. Research on heat shock protein pathways and sex-related microglial responses is also increasing. Some core terms have remained frequent over the years, such as "neuroinflammation" and "activated protein kinases". In contrast, the explosive emergence of brain-derived neurotrophic factor (BDNF) and spinal cord stimulation (2020-2021; burst intensity = 2.56) indicates a growing interest in synaptic and circuit control and neuromodulation-based approaches. In the PubMed subset, 33.6% of studies directly focused on treatment development, with gene therapy, intrathecal administration, and microenvironment remediation also appearing more frequently. When we combine data from WoSCC and PubMed over the past 20 years, we can see a significant shift in the explanation of spinal pain in this field. Early research often described the problem as "glial cell activation-cytokine release." Recent research, however, focuses on specific pathways, particularly microglial state regulation, oxidative stress-autophagy connections, and kinase signaling. This shift in treatment approaches is also reflected in translational studies. Many studies no longer rely primarily on systemic drugs but instead focus on targeted strategies such as intrathecal administration, gene or cell therapy, extracellular vesicles, and neuromodulation. These trends make polarization-related molecular nodes ideal candidate targets for precision analgesia. However, bibliometric results are dependent on database coverage, keyword processing, and clustering settings. Some "hotspots" may reflect changes in terminology or citation habits rather than true mechanistic importance. The rise of neuromodulation keywords may also reflect broader clinical applications; microglial mechanisms are plausible, but contributions from other circuit-level mechanisms may also play a role. These results indicate that the field is moving beyond a purely inflammatory perspective toward systemic intervention models. Currently, there is a greater focus on microglial homeostasis and M2-like anti-inflammatory/immune repair processes, as well as sex and metabolic factors that may influence responses. This research direction supports immune repair and more personalized analgesia. Simultaneously, stronger mechanistic arguments require cell state-specific measurements rather than broad phenotypic labels.
Postbiotics, defined as preparations of inanimate microorganisms and/or their components, have attracted increasing attention because of their stability and functional benefits. Although lactic acid bacteria (LAB) modulate immune responses and oxidative stress, their direct effects on host mitochondrial homeostasis remain poorly understood. In this study, we investigated whether postbiotics derived from Ligilactobacillus salivarius enhance mitochondrial robustness in porcine intestinal macrophages (IPIMs). To establish a model of disrupted mitochondrial redox homeostasis, mitochondrial dysfunction was induced using the mitochondrial complex III inhibitor Antimycin A (AMA). High-dose AMA induced mitochondrial reactive oxygen species (mtROS) accumulation and suppressed the expression of antioxidant genes, including SOD2. Pre-stimulation with heat-killed L. salivarius suppressed AMA-induced mtROS production in selected representative strains, with the most effective strains enhancing SOD2 expression. These mtROS-suppressive strains selectively activated Toll like receptor (TLR)1/2 and TLR2/6 signaling, whereas non-protective strains failed to induce TLR responsiveness. Pharmacological inhibition of TLR2 abolished mtROS suppression and SOD2 induction, confirming TLR2 dependency. Furthermore, Seahorse extracellular flux analysis revealed that postbiotics enhanced mitochondrial respiration, including maximal respiration and spare respiratory capacity, via TLR2 signaling. Collectively, our findings demonstrate that postbiotics function as immunometabolic modulators that reinforce mitochondrial redox homeostasis and respiratory capacity via TLR2 signaling, highlighting a novel mechanism by which microbial components contribute to intestinal immune cell homeostasis.
Autism spectrum disorder (ASD) encompasses a group of neurodevelopmental disorders influenced by genetic and environmental factors, although the molecular mechanisms underlying their interactions remain unclear. We previously reported dysregulated cytokine signalling in chromosome 15q-duplication syndrome (Dup(15q)), a common syndromic form of ASD. Dup(15q) induced pluripotent stem cell (iPSC)-derived neurons exhibit an amplified Signal Transducer and Activator of Transcription-3 (STAT3) response to Interleukin-6 (IL-6), a cytokine often upregulated in ASD. To identify candidate genes within the 15q region that may modify cytokine signalling, we investigated Cytoplasmic FMRP-Interacting Protein 1 (CYFIP1), as CYFIP1 dysregulation has been linked to altered expression of genes involved in immunoregulatory pathways. CYFIP1 was overexpressed using a plasmid vector in human HEK-293 and SH-SY5Y neuroblastoma cells. Following stimulation with IL-6 or Interferon (IFN)-γ, a variety of biochemical assays (qRT-PCR, Western blotting and dual-luciferase reporter assays) and neurite tracing experiments were performed to assess the effects of increased CYFIP1 expression on IL-6/STAT3 and IFN-γ/STAT1 signaling responses. CYFIP1-overexpression HEK-293 cells display reduced STAT3 and STAT1 expression, but enhanced IL-6-induced/IFN-γ-induced STAT3/STAT1 transcriptional activity. Furthermore, CYFIP1-overexpression in SH-SY5Y cells was associated with reduced basal neurite outgrowth and altered IL-6-associated neurite outgrowth. These findings suggest that CYFIP1-overexpression modifies cytokine-responsive transcriptional pathways in vitro and provides novel insight into how CYFIP1 dysregulation may contribute to dysregulated cytokine signaling in ASD, advancing our understanding of the molecular mechanisms underlying neuroinflammatory processes in this disorder.
The major histocompatibility complex class II (MHC-II) pathway is central to adaptive immunity and immune tolerance, and its age-related dysregulation is increasingly linked to chronic neuroinflammation. The HLA-DRB1*15:01 allele, the strongest genetic risk factor for multiple sclerosis, has been implicated in shaping pathogenic CD4+ T-cell responses and broader neuroimmune vulnerability, yet how this allele modulates age- and sex-dependent neuroimmune processes within the central nervous system (CNS) remains poorly defined. We investigated the impact of HLA-DRB1*15:01 expression using a humanized mouse model (HLA mice) and wild-type (WT) controls. Male and female mice were analyzed at 6, 9, and 15 months of age, with endocrine stratification in females. Behavioral testing, flow cytometry, immunofluorescence, and multiplex cytokine analyses were used to assess cognitive performance, glial immune-associated changes and oxidative stress, astrocyte-microglia IL-3/IL-3R signaling, endothelial activation, selective immune cell accumulation at CNS borders, tissue organization, and hippocampal cytokine profiles. HLA mice developed age- and sex-dependent cognitive impairment, most pronounced in aged females. HLA-DRB1*15:01 expression promoted progressive microglial immune-associated changes, characterized by increased CD14 and CD68 expression, elevated mitochondrial oxidative stress, altered astrocyte phenotypes, and enhanced IL-3/IL-3R signaling. Hippocampal axonal and myelin organization was disrupted in aged HLA mice and was spatially associated with increased microglial presence. HLA mice also exhibited selective immune remodeling, including increased accumulation of CD4+ T cells and NK1.1+CD3+ natural killer T (NKT) cells, particularly in females, accompanied by endothelial activation marked by elevated ICAM-1 and E-selectin expression. Hippocampal cytokine profiling revealed selective sex-biased alterations, without broad induction of classical inflammatory cytokines. Together, these findings demonstrate that HLA-DRB1*15:01 drives a coordinated, age- and sex-dependent neuroinflammatory program linking behavioral dysfunction, glial immune-associated changes and oxidative stress, selective immune cell recruitment, endothelial activation, tissue remodeling, and targeted cytokine imbalance. This integrated phenotype provides mechanistic insight into how this major MS risk allele confers vulnerability to chronic neuroinflammation during aging, with heightened impact in females, independent of reproductive cycling stage.
Although fear conditioning has elucidated cue-evoked acute fear responses, the mechanisms by which stress experiences induce generalized internal states linked to anxiety or phobia are poorly understood. Here, we report that robust stress induces a persistent behavioral change characterized by avoidance of a confined space, claustrophobia-like behavior (CLB) in Drosophila. Unlike aversive memory formation, the development of CLB does not require dopamine receptors. Our neuronal screening determined that neuropeptide signaling via Allatostatin-A inactivates the downstream neurons via its receptor AstA-R1, causally inducing CLB. Moreover, gene expression profiling of individual fly heads revealed that innate immune response activation in the blood-brain barrier is involved in CLB. Our data demonstrate that stress-induced persistent behavioral change would not be related to a canonical mechanism of aversive memory formation, rather involves neuropeptidergic signaling and the blood-brain barrier, providing the mechanism determining internal states which persistently change into a phobia-like mode.
DNA N6-methyladenine (6 mA) has recently been recognized as a novel epigenetic modification, with ALKBH4 identified as a specific demethylase. However, the role of ALKBH4 and DNA 6 mA in the pathogenesis of ossification of the posterior longitudinal ligament (OPLL) remains unclear. Tissue samples from OPLL patients and normal posterior longitudinal ligaments were collected, and ligament fibroblastic cells (LFCs) were isolated from OPLL tissues using primary culture. The expression and functional relevance of ALKBH4 in OPLL were investigated through reverse transcription quantitative polymerase chain reaction and western blot analyses. Osteogenic potential was further assessed using alizarin red S staining and alkaline phosphatase activity assays. Methylation status was evaluated by enzyme-linked immunosorbent assay and chromatin immunoprecipitation. Furthermore, the regulatory role of ALKBH4 in LFC ossification was investigated through the bone morphogenetic protein 2 (BMP2) and Wnt/β-catenin pathways. ALKBH4 expression was significantly elevated in both OPLL tissues and LFCs, whereas global 6 mA levels and BMP2-associated 6 mA were reduced. Overexpression of ALKBH4 promoted the ossification of LFCs, while its knockdown suppressed osteogenesis. ALKBH4-mediated DNA demethylation at the 6 mA site facilitated Yin Yang 1 (YY1) binding to the BMP2 promoter, enhancing BMP2 transcription and driving ossification. Silencing of BMP2, Wnt/β-catenin, or YY1 attenuated the pro-ossification effects of ALKBH4. Furthermore, the mutation that affects the action of ALKBH4 at the 6 mA site fails to induce the osteogenic effect of LFC. These findings demonstrate that ALKBH4 regulates OPLL progression by modulating BMP2 and Wnt/β-catenin signaling and by promoting BMP2 transcription through site-specific demethylation in human OPLL tissues and primary LFCs. ALKBH4 may therefore represent a promising therapeutic target for the management of OPLL.
In transfected cells, adhesion G protein-coupled receptors (GPCRs) are activated by tethered agonists that are embedded in their canonical autoproteolytic GAIN domain. It is unknown, however, whether a tethered agonist-dependent activation mechanism generally mediates the physiological functions of adhesion GPCRs. Here, we show that G protein signaling by BAI3 (Adgrb3), a brain-specific adhesion GPCR, is essential for its functions in controlling axon and dendrite growth and promoting synapse formation. Moreover, our signal transduction assays confirm that constitutive exposure of BAI3's tethered agonist massively stimulates (~5-fold) its GPCR activity. However, the constitutive exposure of BAI3's tethered agonist, produced by deletion of its extracellular domains, blocked instead of activating BAI3's functions in regulating axonal and dendritic growth and promoting synapse formation. Moreover, inactivating mutations of BAI3's tethered agonist or deletion of BAI3's constituent GAIN domain did not detectably impair BAI3's physiological functions. Thus, the GPCR activity of BAI3 is functionally required, whereas tethered agonist-mediated stimulation of its GPCR activity is not.
MDM2 is transcriptionally activated by the ST-MYCL-Tip60 complex in virus-positive Merkel cell carcinoma (MCC). MDM2 suppresses p53 and is a rational therapeutic target. MDM2 inhibitors face an intrinsic limitation: p53 activation induces MDM2 transcription, creating a feedback loop that blunts inhibitor efficacy. We demonstrate that MDM2 degraders KTX-049 and KT-253 overcome this limitation by collapsing the p53/MDM2 negative feedback loop. KTX-049 was >100-fold more potent than the MDM2 inhibitor DS-3032 across WT p53 MCC cell lines, and this superior potency was quantitatively supported by mechanistic mathematical modeling. In vivo, KT-253 produced deep and durable tumor regressions, including complete responses in patient-derived xenograft models. Acquired resistance was strongly associated with acquisition of TP53 mutations, confirming on-target pathway pressure. These findings establish feedback architecture as a critical determinant of therapeutic response and position MDM2 degradation as a qualitatively distinct strategy that produces more durable pathway engagement than MDM2 inhibition, providing a preclinical rationale for prioritizing MDM2 degraders in WT TP53 MCC.
Hepatocellular carcinoma (HCC) is a major health issue, but treatment options are limited. This study investigated the role of CCR4-NOT transcription complex subunit 9 (CNOT9) in the pathogenesis of HCC and its potential as a therapeutic target. Bioinformatics analysis was performed on RNA-seq data from the TCGA and GTEx databases to assess CNOT9 expression and prognostic significance. CNOT9 expression was validated in clinical samples and cell lines using qRT-PCR and Western blot. CNOT9 was knocked down in HCC cell lines, and its effects on proliferation, apoptosis, and cell cycle were investigated using functional assays (CCK-8, EdU, colony formation, and flow cytometry). The underlying molecular mechanisms were explored via RNA-seq and Western blot analysis of the AKT pathway and cell cycle regulators. Xenograft mouse models were used to confirm the oncogenic role of CNOT9 in vivo. CNOT9 mRNA and protein expression were upregulated in HCC patients and associated with poor prognosis. CNOT9 induces abnormal proliferation of HCC cells and G2/M phase cell cycle progression. Knocking down CNOT9 reduces cell proliferation, increases apoptosis, and causes cell arrest at the G2 phase. CNOT9 knockdown activates PTEN to inhibit the AKT pathway and suppresses the expression of cell cycle-related proteins p53, p21, CCNE1 and CDK2. CNOT9-deficient tumors exhibited reduced growth in mice, supporting its pro-oncogenic role. This study first elucidates the molecular mechanism by which CNOT9 drives HCC progression through post-transcriptional regulation of the PTEN/AKT/p53 axis, providing a theoretical basis for precision treatment strategies targeting CNOT9 or the PTEN/AKT/p53 pathway.
Domestication has profoundly transformed human production and lifestyles. The Qingtian rice-fish co-culture system is the first globally important agricultural heritage system (GIAHS). PF-carp are a key species in the Qingtian rice-fish system and have been domesticated in rice paddies for more than a millennium, yet the mechanisms of their tolerance to high temperature conditions remain unresolved. In this study, 28℃ as the control group (C0), and two heat stress groups were established at 38℃ for 0 h (H0) and 24 h (H24). Brain tissues were sampled for physiological index measurements and transcriptomic analysis. Physiological analyses showed that the activities of SOD, CAT, and GSH-Px increased initially and then declined, whereas MDA levels exhibited a continuous increase. Transcriptome profiling identified 9,825 differentially expressed genes (DEGs). KEGG enrichment analysis of DEGs indicated that immune responses and metabolic regulation were consistently involved throughout the thermal adaptation process. During acute warming phase, pathways such as protein processing in the endoplasmic reticulum, FoxO signaling pathway and glycerophospholipid metabolism were enriched. Under prolonged high temperature exposure, cytokine-cytokine receptor interaction, PPAR signaling pathway and TGF-β signaling pathway were prominently enriched. Within these pathways, genes including grp94, hsp70, bip, nef, il-1r, tlr8, cctgα6, ccr4, cxcr3, and il-10 were significantly up-regulated (p < 0.05). These results indicate that PF-carp exhibit coordinated brain physiological and transcriptomic responses to high temperature exposure, involving protein quality control, immune signaling, and metabolic regulation. Collectively, our findings provide new insights into the mechanisms by which PF-carp adapt to thermal exposure and provide theoretical support for the breeding of heat tolerant fish.
To explore whether pyroptosis, an inflammatory type of programmed cell death, participates in the initiation and progression of myopia, and to further elucidate its regulatory role in scleral remodeling. Scleral tissues from form-deprivation myopia (FDM) mouse models were subjected to transcriptome sequencing to screen inflammatory and cell death-related molecular characteristics. Differentially expressed gene analysis and pathway enrichment analysis were adopted to identify pyroptosis-related signaling pathways. Meanwhile, human scleral fibroblasts were treated with complement component 3a (C3a) to construct an in vitro inflammatory cell model. Western blot, immunofluorescence staining, lactate dehydrogenase (LDH) release assay and transmission electron microscopy were applied to detect extracellular matrix (ECM) alterations and the expression levels of core pyroptosis markers including NOD-like receptor family pyrin domain-containing protein 3 (NLRP3), caspase-1 and N-terminal gasdermin D (GSDMD-N). Transcriptomic results revealed that inflammatory response, immune regulation, and pyroptosis-related pathways were significantly enriched in myopic scleral tissues, accompanied by synchronous activation of inflammasome signaling. In vitro inflammatory intervention downregulated the expression of type I collagen and upregulated matrix metalloproteinase-2 (MMP-2), suggesting aggravated ECM degradation. The levels of interleukin-1β (IL-1β), interleukin-18 (IL-18), cell membrane permeability, as well as NLRP3, caspase-1, and GSDMD-N were obviously increased in activated fibroblasts. Immunofluorescence and ultrastructural observations further confirmed gasdermin protein-mediated cell membrane damage and typical pyroptotic morphological changes. In vivo animal experiments and in vitro cellular studies collectively verify that the activation of inflammasome-caspase-1-GSDMD signaling axis is involved in myopia-related scleral remodeling. Pyroptosis acts as a key mechanistic bridge linking inflammatory response and scleral structural weakening, which offers novel molecular targets for the intervention and suppression of myopia progression.
To investigate the anti-fibrotic effects of Isofraxidin (IF) on liver fibrosis and its underlying mechanisms. C57BL/6J mice (bred by The Jackson Laboratory) were induced with liver fibrosis using CCl4 and treated with different doses of IF (20 mg/kg and 40 mg/kg) via gavage. Colchicine served as the positive control. Liver fibrosis markers (α-smooth muscle actin,α-SMA; Fibronectin, FN; Collagen type I, Col-I) were assessed by histological staining, immunohistochemistry, and western blotting. Network pharmacology and molecular docking were adopted to conjecture underlying targets and signaling pathways of IF, followed by experimental validation. IF at various doses improved liver function and exhibited dose-dependent anti-fibrotic effects. Network pharmacology identified Protein kinase Kinase B (PKB, Akt), Proto-oncogene tyrosine-protein kinase (Src), Epidermal growth factor receptor (EGFR), Heat Shock Protein 90 Alpha Family Class A Member 1 (HSP90AA1), Glycogen Synthase Kinase 3 beta (GSK3β), and Monoamine Oxidase B (MAOB) as the core targets. Molecular docking showed good binding affinity between IF and these targets. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses indicated that the Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway is a major pathway involved in IF's anti-fibrotic effects. Experimental validation confirmed that IF inhibited the expression of inflammatory factors (Tumor Necrosis Factor-alpha, TNF-α; Interleukin-1 beta, IL-1β; Interleukin-10, IL-10) and the phosphorylation of NF-κB and Inhibitor of Nuclear Factor Kappa B Alpha (IκBα). IF effectively alleviated CCl4-induced liver fibrosis in mice by modulating key targets (AkT, Src, EGFR, HSP90AA1, GSK3B, and MAOB) and the NF-κB signaling pathway.
Visceral hypersensitivity and impaired intestinal barrier function are key features in the pathophysiology of irritable bowel syndrome (IBS). Activation of central orexin signaling has been shown to ameliorate these gastrointestinal disturbances via vagal pathways mediated by brain orexin and central muscarinic receptors. Given that several studies suggest that L-carnitine, a naturally occurring amino acid, can activate orexin signaling, we hypothesized that L-carnitine attenuates these gastrointestinal abnormalities in a rat model of IBS. Visceral pain thresholds to colonic balloon distension were evaluated by electromyographic recordings of abdominal muscle contractions. Colonic permeability was assessed by measuring Evans blue uptake in lipopolysaccharide (LPS) and corticotropin-releasing factor (CRF) -induced IBS models in male Sprague-Dawley rats. Intragastric administration of L-carnitine (3-200 mg kg-1 day-1 for 3 days) dose-dependently prevented LPS-induced visceral hypersensitivity and colonic hyperpermeability, and attenuated these changes induced by CRF. Intracisternal administration of SB-334867, an orexin 1 receptor antagonist, abolished the effects of L-carnitine in the LPS model. Moreover, atropine, sulpiride, a dopamine D2 receptor antagonist, and NG-nitro-L-arginine methyl ester, a nitric oxide synthesis inhibitor, but not scopolamine butylbromide, a peripheral muscarinic receptor antagonist, blocked the effects of L-carnitine. Additionally, compound C, an AMPK inhibitor, and GW9662, a PPAR-γ antagonist, also abolished L-carnitine's effects. L-Carnitine prevents visceral hypersensitivity and colonic hyperpermeability in IBS models via mechanisms involving brain orexin, as well as central muscarinic, dopamine D2, nitric oxide, AMPK, and PPAR-γ signaling. These findings suggest that L-carnitine may represent a promising therapeutic option for IBS.
This study aimed to explore the dental applications of AG73, a laminin-derived adhesive peptide, by examining its effects on human dental pulp cells (hDPCs), particularly in cell adhesion and mineralization. It also sought to identify the key functional residue of AG73 and the signaling pathways involved in its pro-mineralization activity. We compared AG73's pro-mineralization activity with peptides derived from LAMA5 using alizarin red staining. Immunofluorescence staining assessed its cell adhesion properties. Alanine-scanning mutagenesis was performed to identify key residues for AG73's adhesion activity. Finally, we used p38 and JNK MAPK inhibitors to determine which signaling pathways were involved in AG73-induced mineralization. AG73 exhibited superior pro-mineralization activity compared to other peptides derived from LAMA5 in hDPCs. Immunofluorescence staining confirmed AG73's strong cell adhesive ability within 20 minutes. Alanine-scanning identified isoleucine at the tenth position as crucial for adhesion. AG73 was the most effective peptide in inducing mineralization, with no mutated versions outperforming it. p38 signaling was found to play a positive role in AG73-induced mineralization, while JNK inhibition enhanced mineralization, suggesting JNK inhibitors as potential inducers. AG73 has strong pro-mineralization activity and significant cell adhesive properties in hDPCs. The isoleucine at the tenth amino acid position was identified as a critical residue for AG73's adhesive ability. Additionally, p38 was found to be involved in AG73-regulated mineralization, while JNK inhibition promoted the process. This is the first study to explore AG73's effects in hDPCs and provides a foundation for its potential use as a dental-pulp regenerative material.
Although epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have substantially improved the clinical outcomes of patients with EGFR-mutated non-small cell lung cancer (NSCLC), most patients ultimately develop acquired resistance. MET amplification is a common resistance mechanism to EGFR-TKIs; however, its effects on receptor dimerization and the resulting dependence on EGFR or MET signaling have not been fully elucidated. We established a classical lung cancer cell line model with acquired resistance to osimertinib, a third-generation EGFR-TKI, through exposure to the drug (HCC827OR). Activation profiles of phospho-receptor tyrosine kinases and genomic alterations were investigated, and drug sensitivity to osimertinib and the MET-TKI savolitinib was evaluated. EGFR and MET signaling status and dimerization/interaction of EGFR, MET, and ERBB3 were analyzed. In addition, exogenous MET overexpression was introduced into parental HCC827 cells to assess its effects on drug sensitivity and receptor dimerization. HCC827OR cells exhibited MET amplification and showed sensitivity to savolitinib monotherapy. ERBB3 phosphorylation correlated with the efficacy of the TKIs. Consistently, we found reduced interaction between EGFR and ERBB3 and increased interaction between MET and ERBB3 in HCC827OR cells. MET homodimers and MET-EGFR interaction increased while EGFR homodimers decreased in those cells. Exogenous MET overexpression in the parental cells partially recapitulated the drug-resistant phenotype, showing partial sensitivity to savolitinib, increased MET homodimers, and decreased EGFR homodimers. MET amplification alters the balance of EGFR/MET/ERBB3 dimerization, leading to a shift in signaling dependence from EGFR to MET. These findings provide insight into therapeutic strategies for EGFR-mutated NSCLC.
Pancreatic ductal adenocarcinoma (PDAC) is the most frequent type of pancreatic cancer with a poor prognosis and resistance to conventional radio- and chemotherapy. Radiation can induce pro-inflammatory signaling by activating the type 1 interferon (IFN-I) response, which can be enhanced by targeting negative regulators of the IFN-I pathway, such as ADP-ribosyltransferase PARP7. Here, we show that PARP7 inhibitors (PARP7i) enhance radiation-induced cell death and promote STING- and NF-κB-dependent immunogenic signaling in PDAC cells, leading to inflammatory gene expression and cytokine release. These effects were most pronounced in BxPC-3 cells, which exhibit higher baseline expression of PARP7, AHR, and interferon response genes. PARP7i potentiated the immunogenic effects of hypofractionated radiation by inducing a STING-dependent IFN-I response, leading to immunogenic cell death and activation of monocytes and NK cells. Carbon ion irradiation elicited stronger immunogenicity than X-rays when combined with PARP7i. KRAS-mutated PANC-1 cells showed a higher expression of enzymes that convert ATP to immunosuppressive adenosine, which was enhanced by radiation. This may explain why PARP7i were more effective as monotherapy in PANC-1 cells, promoting NK cell activation. These findings support further evaluation of PARP7i in PDAC in combination with radiotherapy or as monotherapy, depending on the immunosuppressive effects of radiation.
Erectile dysfunction (ED) secondary to cavernous nerve injury (CNI), such as that occurring after radical prostatectomy, remains a significant clinical challenge, particularly among patients who do not respond to phosphodiesterase Type 5 inhibitors (PDE5i). Extracellular vesicles (EVs) derived from umbilical cord blood (UCB) have recently emerged as a promising cell-free therapeutic modality owing to their potent regenerative and anti-inflammatory properties. This study aimed to investigate the therapeutic efficacy of UCB-derived EVs (UCB-EVs) in restoring erectile function in a bilateral CNI mouse model. UCB-EVs were isolated and characterized by transmission electron microscopy, nanoparticle tracking analysis, and western blotting. Their neuroregenerative effects were assessed in vitro using Neuro-2a (N2a) cell migration assays and ex vivo by evaluating neurite outgrowth in major pelvic ganglia (MPG) and dorsal root ganglia (DRG). In vivo, fluorescently labeled UCB-EVs were tracked following intracavernous administration, and erectile function was quantified using maximal ΔICP and baseline-corrected total ΔICP during cavernous nerve stimulation. Histological and biochemical analyses were conducted to assess neurovascular integrity, apoptosis, proliferation, oxidative stress, and activation of PI3K/AKT signaling in penile tissues. UCB-EVs significantly enhanced N2a cell migration in vitro and promoted neurite sprouting in MPG and DRG explants ex vivo. In CNI mice, intracavernous administration of UCB-EVs resulted in dose-dependent recovery of erectile function, as demonstrated by significant increases in maximal ΔICP and baseline-corrected total ΔICP during cavernous nerve stimulation. Biodistribution analysis demonstrated preferential localization of UCB-EVs to the penile neurovascular bundle. Histological and molecular evaluations revealed preservation of endothelial and neuronal populations, suppression of apoptosis, enhanced cellular proliferation, attenuation of reactive oxygen species (ROS) accumulation, and activation of PI3K/AKT signaling pathways in UCB-EV-treated mice. UCB-EVs promote neurovascular regeneration, mitigate oxidative stress, and improve erectile function following CNI. These findings suggest that UCB-EVs represent a promising regenerative strategy for postprostatectomy ED, particularly in patients refractory to conventional pharmacotherapy.
Chronic graft-versus-host disease (cGVHD) is a major complication of allogeneic hematopoietic stem cell transplantation, with cutaneous fibrosis representing a debilitating and treatment-resistant manifestation. We identified a hypoxia-driven inflammatory and fibrotic program in cGVHD skin mediated by a hypoxia-inducible factor 1α-phosphatidylinositol 3-kinase-interleukin-13 (HIF-1α-PI3Kδ-IL-13) signaling axis. Spatial transcriptomics, single-cell RNA sequencing, and multiplex immunofluorescence revealed stabilization of HIF-1α in epidermal cells in hypoxic regions. HIF-1α stabilization was accompanied by IL-13 production from PI3Kδ-activated T cells and macrophage-epidermal cross-talk, which together promoted fibrosis and the formation of tertiary lymphoid structure (TLS)-like aggregates. In human epidermal cell lines and induced pluripotent stem cell-derived skin organoids, IL-13 directly induced HIF-1α expression in keratinocytes under normoxic conditions. HIF-1α expression was further amplified in hypoxic conditions, driving profibrotic signaling. Pharmacologic inhibition of HIF-1α, PI3Kδ, or IL-13 reduced tissue hypoxia, disrupted TLS-like aggregates, and ameliorated disease in murine cGVHD models. These findings identify a targetable circuit linking hypoxia, type 2 inflammation, and immune-stromal dysfunction in cGVHD and provide a translational framework for treating fibrotic cGVHD and related immune-mediated fibrosing diseases.
To integrate evidence from human, animal, and in vitro studies to elucidate the molecular mechanisms underlying phagocytic aberrations in asthmatic macrophages and to construct a cross-hierarchical mechanistic framework. This systematic review followed the PRISMA 2020 statement. PubMed and Web of Science Core Collection were searched from inception to March 6, 2026. We included original human, animal, and cellular studies on asthma that investigated macrophage phagocytic function and reported related molecular mechanisms. Two independent reviewers performed screening, data extraction, and quality assessment. Due to substantial heterogeneity across studies, a narrative synthesis approach was used to integrate the evidence. A total of 37 studies, published between 1982 and 2025, were ultimately included. Overall, macrophage phagocytic function in asthma is characterized by aberrations dependent on clinical phenotype, phagocytic substrate, and the microenvironment. Based on the included studies, the relevant mechanisms were categorized into eight categories: dysregulation of phagocytic receptor signaling; defects at various stages of efferocytosis; immunometabolic reprogramming; aberrant signal transduction; regulation by immunomodulatory factors; acquired alterations in cellular function; circadian clock regulation; and other unclassified mechanisms. Phagocytic aberrations in macrophages in asthma represent a complex process driven by a multi-layered, interconnected molecular network. These aberrations contribute to the persistence of chronic airway inflammation, increased susceptibility to infection, elevated risk of acute exacerbations, and the development of severe or refractory asthma. Systematic integration of these mechanisms enhances the understanding of innate immune dysfunction in asthma and provides a theoretical basis for developing therapeutic strategies targeting macrophage phagocytic function. https://www.crd.york.ac.uk/prospero/, identifier CRD420261332569.
The tumor microenvironment (TME) limits durable antitumor immunity by impairing CD8+ T cell responses. Memory like CD8+ T cells are important for long-term immune control but are often restricted in the TME. Dendritic cells (DCs) are key regulators of T cell fate. Previous studies have shown that SHP1 in DCs fosters an immunosuppressive microenvironment and facilitates tumor immune escape. T cell factor-1 (TCF-1), encoded by Tcf7 gene, is required for central memory CD8+ T cell (TCM) formation and is closely linked to canonical Wnt/β-catenin signaling. However, whether SHP1 in DCs regulates TCF-1 expression and TCM formation remains unclear. To investigate the role of DC intrinsic SHP1 in T cell immunity, SHP1 deficient DC2.4 cells and primary bone marrow derived dendritic cells (BMDCs) were co-cultured with OT-1 T cells to assess proliferation, TCM formation, cytotoxic activity, and TCF-1 expression. A DC-specific SHP1 knockout mice model was used to evaluate antitumor immunity in vivo, and Tcf7 or Ctnnb1 silencing was used to probe the TCF-1/Wnt/β-catenin axis. SHP1 downregulation in DCs markedly enhanced CD8+ T cell proliferation, promoted the generation of CD62L+ CD44+ central memory T cells, and potentiated B16-F10-OVA tumor cell killing, accompanied by increased TCF-1 expression in OT-1 T cells. In DC-specific SHP1 knockout mice, EO771 tumor growth was suppressed with concurrent increases in intratumoral IFN-γ+ and TCF-1+ CD8+ T cell frequencies. Mechanistically, we found that DC SHP1 regulates TCM formation via TCF-1, as silencing Tcf7 in OT-1 T cells abrogated this effect. SHP1-deficient DCs activated Wnt/β-catenin signaling in CD8+ T cells, as shown by increased active β-catenin, total β-catenin, c-Myc and Cyclin D1, and a reduced phospho β-catenin/total β-catenin ratio. Critically, Ctnnb1 silencing in T cells abrogated the enhanced proliferation, TCM formation, and cytotoxic activity induced by SHP1-deficient DCs. DC-intrinsic SHP1 restrains central memory CD8+ T cell formation via the TCF-1/Wnt/β-catenin axis.