Glutamate is an important neurotransmitter in the mammalian brain. Among the receptors that glutamate interacts with is metabotropic glutamate (mGlu) receptor 2, a Gαi/o-coupled receptor. These receptors are primarily located on glutamatergic nerve terminals and act as presynaptic autoreceptors to produce feedback inhibition of glutamate release. Abundant mGlu2 receptors are distributed in major glutamatergic pathways in the basal ganglia, especially the corticostriatal and thalamostriatal projections in the striatum. These receptors are involved in the regulation of motivation, reward processing, learning, motor, and cognitive functions. As an inhibitory presynaptic receptor, mGlu2 is linked to the addictive properties of drugs of abuse, a topic summarized in this review. Chronic exposure to multiple addictive drugs and alcohol causes the adaptive downregulation of mGlu2 receptors in their expression and function in the key regions of the limbic reward circuit. This downregulation contributes to the remodeling of limbic excitatory synaptic transmission and plasticity critical for enduring drug-seeking behavior. Normalization of mGlu2 activity by pharmacological or genetic approaches attenuates drug taking and seeking. Here, we highlight that recent progress in mGlu2 biology research demonstrates the pivotal roles of mGlu2 receptors in different aspects of drug addiction. mGlu2 subtype-selective agents (both orthosteric and allosteric compounds) thus have the potential to be developed into novel pharmacotherapies for addictive conditions.
The serine protease 23 (PRSS23) is a highly conserved member of trypsin-like serine proteases, which are associated with numerous essential processes, including digestion, blood coagulation, fibrinolysis, development, fertilization, apoptosis, and immunity. Original reports on PRSS23 unfolded not earlier than 2006 when a molecular biology study characterized and described PRSS23 as an ovarian protease. Then, in 2012, another important study was published linking PRSS23 with proliferation of breast cancer cells by an estrogen receptor 1 (ESR1)-dependent transcriptional activation of the serine protease. Thereafter, a developmental study in zebrafish reported the implication of PRSS23 in endothelial-to-mesenchymal transition (EndMT) during cardiac valve formation. Although these early studies on PRSS23 have revealed its involvement in some critical or fundamental processes, only in recent years an increasing number of studies have evolved describing the expression and functions of PRSS23 in various normal physiological conditions, diseases, and experimental configurations. Besides breast cancer, PRSS23 has been shown to be involved in different types of malignancies, e.g., in gastric cancer, where drug screening found that the protease inhibitor tipranavir impedes cancer-promoting PRSS23 expression. New innovative techniques such as single cell RNA-sequencing (scRNA-seq) and bioinformatics studies accelerated the discovery of gene expression changes in smaller cell populations, which, e.g., led to the identification of marked PRSS23 expression in a myofibroblast-like subpopulation in localized scleroderma. This review compiles major and significant research results that have contributed to our current knowledge of PRSS23 and briefly discusses where prospective studies could add to our understanding of this versatile serine protease.
Histones were once thought to be exclusive to the nucleus, but were recently discovered in the extracellular space, where they play important roles in disease pathogenesis. In addition to their traditional functions in chromatin organization and gene regulation, extracellular histones also serve as damage-associated molecular patterns (DAMPs), drive inflammation and immune responses, and are responsible for the progression of diseases such as sepsis, autoimmune diseases, and inflammatory diseases. To effectively target extracellular histones and improve disease progression, this review begins with the release and pathogenic mechanisms of histones and explains the main pathogenic mechanisms of extracellular histones in many diseases. In addition, common antagonistic methods for targeting extracellular histones are summarized, and the mechanisms that need to be further studied at this stage are discussed, providing new directions for the future development of effective and safe histone-targeting drugs.
The bioenergetic machinery of the cell is protected and structured within two layers of mitochondrial membranes. The mitochondrial inner membrane is extremely rich in proteins, including respiratory chain complexes, substrate transport proteins, ion exchangers, and structural fusion proteins. These proteins participate directly or indirectly in shaping the membrane’s curvature and facilitating its folding, as well as promoting the formation of nanotubes, and proton-rich pockets known as cristae. Recent fluorescent super-resolution images have demonstrated the strong dynamics of these events, with constant remodeling processes. The mitochondrial outer membrane itself is also highly dynamic, interacting with the endoplasmic reticulum and its environment to ensure a rapid diffusion of surface components throughout the mitochondrial networks. All these movements occur besides migration, fusion, and fission of the mitochondria themselves. These dynamic events at the level of mitochondrial membranes are primarily dependent on their unique lipid composition. In this review, we discuss the latest advances in phospholipid research, focusing on their metabolism and role in mitochondrial dynamics. This process emphasizes the importance of interactions with the endoplasmic reticulum and mitochondrial matrix enzymes, extending its relevance to lipid sources, in particular, cardiolipins and phosphatidylethanolamines at the cellular, tissue and even whole-organism level. Given the expanding array of characterized mitochondrial functions, ranging from calcium homeostasis to inflammation and cellular senescence, research in the field of mitochondrial lipids is particularly significant. As mitochondria play a central role in various pathological processes, including cancer and neurodegenerative disorders, lipid metabolism may offer promising therapeutic approaches.
Acute myocardial infarction (AMI) is one of the main causes of mortality worldwide. Currently, the most effective treatment is percutaneous coronary angioplasty (PCA). However, paradoxically, the restoration of blood flow induces myocardial reperfusion injury (MRI), contributing up to 50% of the final infarct size. Oxidative stress, characterized by a burst of reactive oxygen species (ROS) following reperfusion, plays a fundamental role in its pathophysiology, causing inflammation, endothelial dysfunction, and cell death mainly through autophagy, apoptosis, ferroptosis, necroptosis, and pyroptosis. To mitigate these injury mechanisms, numerous antioxidant strategies have been evaluated using both in vitro and in vivo models with promising results, but limited benefit when tested in humans. Several antioxidants have biological properties that counteract ROS-induced damage by acting as ROS scavengers, metal chelators, and antioxidant enzyme enhancers. In this review, we focus on the mechanisms by which oxidative stress induces cell death after AMI and highlight the most promising therapeutic antioxidant agents that could provide comprehensive protection against MRI. A multitarget cardioprotective strategy, combining interventions with strong preclinical evidence, could provide a more effective approach for reducing MRI. Our study aims to bridge the gap between basic and clinical research and explore the potential clinical applications of antioxidants.
The most common endocrine cancer, thyroid carcinoma (TC), has a dismal prognosis when it reaches an advanced stage. Integrin α-2 (ITGA2) has been implicated in cancer progression, influencing both DNA damage and repair mechanisms. However, it is unknown how ITGA2 influences these processes in TC.
ITGA2 was identified as a key prognostic gene for TC from the Cancer Genome Atlas-thyroid carcinoma (THCA), GSE3678, GSE29265, and GSE33630 datasets. Functional assays were used to evaluate the impact of ITGA2 knockdown on cell viability, migration, apoptosis, invasion, pyroptosis (N-terminal fragment of GSDME, GSDME-N), and cytotoxicity (Lactate dehydrogenase, LDH). DNA damage markers (phosphorylated histone H2AX on serine 139 (γ-H2AX), phosphorylated ataxia telangiectasia mutated (p-ATM), phosphorylated checkpoint kinase 2 (p-CHK2)) and the level of Reactive Oxygen Species (ROS) were used to assess oxidative stress. The impact of ITGA2 inhibition on Wnt/β-catenin signaling was evaluated, and a mouse xenograft model assessed tumor growth in vivo.
ITGA2 was significantly overexpressed in TC. Knockdown of ITGA2 significantly reduced cell viability, migration, and invasion, while promoting pyroptosis by upregulating cleaved-poly(ADP-ribose) polymerase (PARP) and GSDME-N. ITGA2 silencing also increased LDH activity, enhanced the expression of DNA damage markers (p-ATM, γ-H2AX, p-CHK2), and increased ROS levels. Furthermore, suppression of ITGA2 activity attenuated the Wnt/β-catenin pathway by reducing the levels of MYC proto-oncogene, bHLH transcription factor (C-myc), CD44 molecule (CD44), slug, snail, β-catenin, and wingless-type MMTV integration site family, member 1 (Wnt-1). ITGA2 silencing significantly inhibited tumor growth in a mouse model.
ITGA2 promotes TC progression by regulating the DNA damage response and inhibiting pyroptosis. Knockdown of ITGA2 increases oxidative stress, exacerbates DNA damage, and inhibits the Wnt/β-catenin pathway, indicating it may have potential as a treatment target in TC.
Preeclampsia (PE) is a serious complication of pregnancy characterized by chronic inflammation and immune dysregulation, which significantly increases the risk of neurodevelopmental disorders in offspring, including the autism spectrum disorder (ASD). This review investigated the potential mechanisms linking PE to ASD, with a particular focus on the role of microglial abnormalities. Epidemiological studies have revealed that prenatal exposure to PE raised the risk of ASD, with affected offspring showing increased odds ratios. Microglia, the prime resident immune cells of the central nervous system (CNS), are critical for normal neurodevelopment, influencing processes such as neural stem cell (NSC) proliferation, synaptic pruning, and normal function of the neural circuit. Early-onset preeclampsia (EOPE) and late-onset preeclampsia (LOPE) may have an impact on the microglia abnormality and ASD through not exactly same pathway. Postmortem studies of ASD have further revealed increased microglial density, altered microglial morphology, and upregulated inflammatory markers in key brain regions, including the hippocampus and prefrontal cortex. Understanding the complex processes and potential mechanisms between EOPE, LOPE, microglial abnormalities, and ASD pathogenesis may highlight the importance of early screening and intervention for children born to mothers with PE. Targeting microglia-mediated pathways may offer novel therapeutic strategies to reduce the risk of ASD in these vulnerable populations.
Transcription factors are significant regulators of gene expression in most biological processes related to diabetes, including beta cell (β-cell) development, insulin secretion and glucose metabolism. Dysregulation of transcription factor expression or abundance has been closely associated with the pathogenesis of type 1 and type 2 diabetes, including pancreatic and duodenal homeobox 1 (PDX1), neurogenic differentiation 1 (NEUROD1), and forkhead box protein O1 (FOXO1). Gene expression is regulated at the transcriptional level by transcription factor binding, epigenetically by DNA methylation and chromatin remodelling, and post-transcriptional mechanisms, including alternative splicing and microRNA (miRNA). Recent data indicate a central role for transcription factors in pancreatic β-cell failure in the context of systemic insulin resistance and chronic inflammation. Therapeutic modulation of transcription factor abundance via gene therapy, small-molecule pharmacology, and epigenetic therapies holds great promise for β-cell restoration and metabolic normalisation. However, further clinical translation will require targeted delivery to appropriate tissues, minimising off-target effects and ensuring long-term safety. This review focuses on the involvement of pancreatic β-cells and transcription factors in diabetes development and their therapeutic implications, intending to develop and consolidate a basis for further research in this area and for the treatment of diabetes in the future.
Adipose stromal cells (ASCs) are perivascular mesenchymal progenitors of adipose tissue. In cancer patients, ASCs can mobilize and migrate to the tumor, where they subsequently play an important role in cancer progression. This biological process involves the conversion of recruited ASCs into cancer-associated fibroblasts (CAFs). ASC-derived CAFs influence the tumor microenvironment through extracellular matrix remodeling, vascularization, and immunomodulation. These and other processes mediated by secreted paracrine factors also affect gene expression in carcinoma cells to promote the epithelial-mesenchymal transition (EMT), metabolic adaptation, survival, and invasiveness of cancer cells. ASC-derived CAFs can enhance tumor aggressiveness, accounting in part for the link between obesity and mortality observed in many cancer types that are surrounded by adipose tissue. In this review, we highlight recent findings on the characteristics and functions of ASCs in cancer and discuss their potential as therapeutic targets.
CysB is a member of the large bacterial LysR-type transcriptional regulator (LTTR) protein family. Like the majority of LTTRs, CysB functions as a homotetramer in which each subunit has an N-terminal winged-helix-turn-helix (wHTH) DNA-binding domain connected to an effector-binding domain by a helical hinge region. CysB is best known for its role in regulating the expression of genes associated with sulfur uptake and biosynthesis of cysteine in Gram-negative species such as Escherichia coli and Salmonella enterica. Activation of CysB target genes generally requires the effector N-acetyl-L-serine, which derives from an intermediate in the cysteine biosynthetic pathway. Here, we outline the established roles of CysB in controlling the cysteine regulon, complemented with an interpretation of DNA binding modes inspired by the recently published structure of full-length CysB that is consistent with the ‘sliding dimer’ model proposed for many LTTRs. Notably, CysB orthologs have been described for which N-acetyl-L-serine does not appear to be required as an effector, and CysB regulons frequently include genes that are not directly related to sulfur assimilation and cysteine biosynthesis. Examples include hslJ, which encodes a predicted membrane protein involved in novobiocin resistance in E. coli, and pqsR, encoding a transcriptional regulator involved in Pseudomonas Quinolone Signal production and virulence in Pseudomonas aeruginosa. These data suggest that CysB orthologs have diverged to ensure optimal function and incorporation in distinct gene regulatory networks.
Mitochondria play crucial roles in maintaining health and influencing disease progression by acting as central regulators of cellular homeostasis and energy production. Dysfunctions in mitochondrial activity are increasingly recognized as key contributors to various pathologies, ultimately impacting healthspan and disease outcomes. However, traditional treatments often do not restore damaged mitochondria to a healthy state. Mitochondrial transplantation, a cellular organelle-based therapy in which mitochondria are introduced into a recipient, has emerged as a novel concept in next-generation therapeutics that overcomes the limitations of current cell-based treatments. This review highlights the unique properties of mitochondria as therapeutic agents, including their ability to restore cellular functions and treat a wide range of diseases. In this review, we focus on the unique role of mitochondria in the regulation of stem cell functions, including stem cell fate, self-renewal, and differentiation. Various perspectives have been explored to better understand mitochondrial transplantation therapy, which harnesses the capacity of mitochondria as living drugs in regenerative medicine, as an innovative strategy to bridge the gap between cell therapy and organelle-based treatments and overcome current clinical barriers.
Sarcopenia is the progressive loss of skeletal muscle mass, strength, and function, significantly contributing to frailty, disability, and mortality in aging populations. As life expectancy rises, sarcopenia presents a growing public health challenge, increasing healthcare costs, and diminishing quality of life. Despite its prevalence, sarcopenia is often underdiagnosed due to limitations in current diagnostic tools, including the lack of standardized cut-off values and reliance on physical performance tests. The causes of sarcopenia are multifactorial, involving oxidative stress, chronic inflammation, mitochondrial dysfunction, satellite cell depletion, and impaired angiogenesis. Recent research highlights the role of microRNAs (miRs) in regulating these molecular pathways. miRs influence muscle homeostasis by modulating gene expression related to muscle atrophy, apoptosis, inflammation, and insulin resistance. While non-pharmacological interventions such as resistance training and blood flow restriction therapy remain the primary treatment strategies, their effectiveness is often limited in older adults with reduced muscle regenerative capacity. The identification of miRs as biomarkers could enhance early diagnosis and enable more personalized treatment approaches. However, further research is required to validate their clinical utility and therapeutic potential. This review comprehensively analyses the molecular mechanisms underlying sarcopenia, current diagnostic challenges, and emerging miR-based strategies that could transform its management. Future efforts should focus on integrating these molecular insights into clinical practice to improve early detection and intervention strategies.
Schizophrenia (SZ) is associated with chronic oxidative stress in the patient’s body. Previous studies revealed an increased copy number of genes for 47S pre-ribosomal RNA (pre-rRNA) in SZ patients. In this study, levels of oxidative stress and factors involved in the adaptive response to chronic stress (rDNA transcription) were, for the first time, compared in blood cells of patients with catatonic SZ(C) and paranoid SZ(P), chronic forms of schizophrenia, as well as healthy controls (HC).
Ribosomal DNA (rDNA) and telomere repeat (TR) were quantified in leukocyte DNA using non-radioactive quantitative hybridization. Fragments of 5′ external transcribed spacer (5′ ETS) and 18S rRNA were assayed in leukocyte RNA using quantitative reverse transcription PCR (RT-qPCR). Proteins γ-histone H2AX (γH2AX), NADPH-oxidase 4 (NOX4), nuclear factor erythroid 2-related factor 2 (NRF2), BCL2-like protein 4 (BAX), BCL2, and oxidation marker 8-oxo-2′-deoxyguanosine (8-oxodG) were quantified in blood lymphocytes using flow cytometry.
SZ(C) cells exhibited higher levels of the oxidative stress markers than SZ(P) and HC cells. The rDNA copy numbers in SZ(C) genomes negatively correlated with the amounts of the oxidative stress markers levels. Thus, genomes of blood cells isolated from catatonic patients harbor more copies of ribosomal genes than those from paranoid schizophrenia patients, correlating with higher levels of rRNA in catatonic patients.
The upregulated ribosome biogenesis appears to be required for adaptive response to the elevated levels of oxidative stress in catatonic compared to paranoid patients.
Parkinson’s disease (PD) is characterized by a progressive decline in dopaminergic neurons within the substantia nigra (SN). Although its underlying cause has yet to be fully elucidated, accumulating evidence suggests that neuroinflammation contributes substantially to disease development. Treatment strategies targeting neuroinflammation could improve PD outcomes. Monocyte-derived tolerogenic dendritic cells (tolDCs) modulate immune responses and induce regulatory T cells (Tregs) during various inflammatory diseases. However, the mechanisms underlying tolDC-mediated immunoregulation in PD remain unclear.
We investigated the immune modulatory role of tolDCs by analyzing gene expression patterns and identified that the C-type lectin domain family 5 member A (Clec5a) was highly induced in tolDCs. To assess its function, we generated Clec5a-knockdown tolDCs and measured cytokine production, including interleukin (IL)-10 and IL-6, forkhead box protein P3 (Foxp3)+ Treg induction, and nuclear factor kappa B (NF-κB) signaling activity. Furthermore, we evaluated the therapeutic effects of Clec5a-expressing dendritic cells (DCs) in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse model. Dopaminergic neuron survival, α-synuclein (α-syn) accumulation, neuroinflammatory markers, and locomotor behavior were analyzed following DC administration.
Clec5a-knockdown tolDCs exhibited reduced immunomodulatory function and IL-10 levels but enhanced IL-6 levels. In addition, these cells induced fewer Foxp3+ Tregs and showed significantly enhanced NF-κB signaling activity. In the MPTP-induced PD model, administration of Clec5a-expressing DCs ameliorated dopaminergic neuron loss and α-syn accumulation. Furthermore, Clec5a-expressing DCs reduced the number of CD45highCD11b+CD86+ macrophages in the brain, reduced brain inflammatory cytokine expression, and improved locomotor activity.
These findings suggest that Clec5a plays a critical role in the immunomodulatory function of tolDCs. The administration of Clec5a-expressing DCs effectively reduced neuroinflammation and protected dopaminergic neurons in an MPTP-induced PD model. Therefore, Clec5a-expressing tolDCs may demonstrate therapeutic potential by managing PD symptoms by suppressing inflammatory responses associated with neurodegeneration.
Mesenchymal stem cells (MSCs) are one of the effective treatments for acute graft-versus-host disease (aGVHD) following allogeneic hematopoietic stem cell transplantation (allo-HSCT) due to their potent immunoregulatory function. Given the limited quantity and high heterogeneity of freshly isolated MSCs, extensive in vitro expansion is essential for clinical application. Prolonged passaging of MSCs leads to a decline in therapeutic efficacy, with the underlying mechanisms remaining unclear. This study aimed to explore the mechanism and intervention strategies of immunoregulatory dysfunction of MSCs during passaging.
We compared the therapeutic effects of MSCs at early passages (passage 6, P6-MSCs) and later passages (passage 12, P12-MSCs) in a mouse GVHD model. We also analyzed the expression of Gal-3 in MSCs at different passages and its role in macrophage polarization. Additionally, the selective Gal-3 inhibitor TD139 was evaluated for its effects on Gal-3 expression and the immunoregulatory function of MSCs.
Our data showed an inverse correlation between passage number and therapeutic efficacy in MSCs. Late-passage MSCs (P12) exhibited significantly reduced efficacy in alleviating GVHD compared to early-passage MSCs (P6). The expression of Gal-3 was markedly upregulated in late-passage MSCs (P12), and it was found to directly inhibit anti-inflammatory M2-like macrophage polarization. Our research demonstrated that TD139 dose-dependently suppresses Gal-3 expression in MSCs and restores their immunoregulatory function.
Gal-3 contributes to the decline in the immunomodulatory capabilities of MSCs. TD139, a Gal-3 inhibitor, has a potentially positive effect on rescuing the immunoregulation dysfunction of MSCs in late-passage and may represent a novel strategy to enhance the therapeutic potential of late-passage MSCs for GVHD treatment.
Mediator complex subunit 10 (MED10) serves as a critical regulator of eukaryotic gene expression by facilitating RNA polymerase II activity. Our investigation aims to characterize MED10’s functional contributions and underlying molecular pathways in hepatocellular carcinoma (HCC) development.
MED10 expression patterns in HCC and their correlation with clinicopathological parameters and patient outcomes were examined using bioinformatics databases and immunohistochemistry. Subsequently, we systematically investigated the biological functions of MED10 in the malignant progression of HCC through comprehensive in vitro experiments, including assessments of cell migration (transwell and wound healing assays), proliferative capacity (cell counting kit-8, colony formation, and 5-Ethynyl-2′-deoxyuridine assays), and cell cycle progression (flow cytometry analysis). Furthermore, we elucidated the underlying molecular mechanisms using real-time quantitative PCR (RT-qPCR), western blotting, immunofluorescence staining, and public database analyses. Furthermore, an in vivo subcutaneous xenograft model was employed to validate MED10’s impact on tumor growth.
The results revealed a marked increase in MED10 expression levels within HCC tissues, showing a strong association with unfavorable clinical outcomes. Mechanistically, MED10 induced the epithelial-mesenchymal transition (EMT) and enhanced HCC cell migration. Moreover, MED10 overexpression drives HCC cell cycle progression and proliferation by activating rapidly accelerated fibrosarcoma 1 (RAF1), a process potentially mediated through the mitogen-activated protein kinase (MEK)/extracellular signal-regulated kinase (ERK)/cellular myelocytomatosis oncogene (c-Myc) signaling axis.
MED10 promotes HCC cell migration and EMT but, more importantly, also drives cell cycle progression and proliferation via RAF1 activation, and is related to the MEK/ERK/c-Myc axis.
Disialoganglioside (GD2) is a tumor-associated antigen that is highly expressed in various neuroectodermal cancers, including melanoma. While chimeric antigen receptor (CAR) T-cell immunotherapy has demonstrated remarkable success in treating hematologic neoplasms, the identification of suitable targets remains a major obstacle in translating this approach to solid tumors.
Peripheral blood T lymphocytes from six healthy donors were used to generate GD2-specific CAR T cells via retroviral transduction. The resulting GD2.CAR T cells were characterized by NanoString transcriptome profiling, flow cytometry with hierarchical stochastic neighbor embedding (HSNE) dimensionality reduction, and in vitro cytotoxicity assays against GD2+ and GD2- melanoma cell lines. In vivo experiments were also performed using GD2+ xenograft models and a single intratumoral dose of 8 × 106 GD2.CAR T cells.
The GD2.CAR T cell population exhibited a predominantly naive phenotype (CD8+CD40L+CD69‒CD107a+4-1BB+FasL+) and effective anti-tumor mechanisms involving the granzyme A/B axis, the Fas/FasL axis, and cytokine release. Transcriptome analysis revealed transduction-related effects on proliferation and a shift towards an effector phenotype during early co-culture with tumor cells, accompanied by upregulation of interferon-gamma (IFN-γ) and cytokine signaling genes. GD2.CAR T cells demonstrated robust cytotoxicity against GD2+ melanoma cells in vitro, while significant in vivo tumor control was observed in xenograft models.
GD2.CAR T cells demonstrate potent anti-tumor activity against melanoma in vitro and in vivo, highlighting their therapeutic potential and warranting further clinical investigation.
Lon protease 1 (LONP1), an adenosine triphosphate (ATP)-dependent protease encoded by nuclear DNA that is highly conserved, maintains the mitochondrial protein balance and regulates adaptive responses to cellular stress. LONP1 dysfunction ultimately results in various forms of cellular and tissue damage. The function of LONP1 in hepatocellular carcinoma (HCC) and how it affects HCC growth were investigated in this work.
The RNA and protein expression levels of LONP1 were determined in paired HCC and adjacent tissue samples through real-time quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC) staining. The correlation between LONP1 expression and clinical features was evaluated via statistical analysis. Overexpression (OE) and knockdown (KD) experiments, small RNA interference, Cell Counting Kit-8 (CCK8) and wound-healing assays, and animal experiments were employed to assess the potential mechanism by which LONP1 promotes the proliferation and migration of HCC cells both in vitro and in vivo.
In HCC samples, LONP1 expression was higher than in the equivalent surrounding tissues. Compared to patients with low LONP1 expression, individuals with high LONP1 expression had shorter disease-free survival and overall survival periods. Functionally, LONP1 facilitated the proliferation and migration of HCC cells, whereas LONP1 knockdown mitigated the growth of HCC subcutaneous tumors. Mechanistically, LONP1 affects the processes of ferroptosis and cuproptosis processes by regulating the stability of aconitase 2 (ACO2). Histological analysis showed that the expression of LONP1 in liver cancer tissues was significantly upregulated, accompanied by a decrease in the level of ACO2 protein (Hematoxylin-Eosin (HE) staining and IHC verification). Mitochondrial function experiments indicated that overexpression of LONP1 led to a significant decrease in mitochondrial membrane potential suggesting mitochondrial dysfunction and reduced susceptibility to ferroptosis.
Our results suggest that LONP1 promotes HCC proliferation and migration by inhibiting ferroptosis and cuproptosis through the degradation of ACO2. Therefore, targeting LONP1 might be an effective therapeutic strategy to inhibit HCC growth.
Adenocarcinoma of Lung (LUAD) remains a leading cause of cancer-related deaths across the globe, and patients harboring epidermal growth factor receptor (EGFR) mutations frequently develop resistance to targeted therapies. While aurora kinase A (AURKA) has been implicated in tumorigenesis, its involvement in regulating ferroptosis via the kelch-like ECH-associated protein 1 (KEAP1)/NF-E2-related factor 2 (NRF2)/heme oxygenase 1 (HO‑1) signaling axis in EGFR-mutant LUAD remains poorly understood.
We analyzed RNA-seq and clinical data from 594 LUAD samples from The Cancer Genome Atlas (TCGA) to explore associations between AURKA expression, EGFR mutation status, and immune cell infiltration. A ferroptosis-focused random forest algorithm was constructed to predict EGFR-mutant cases. In vitro, AURKA was silenced by siRNA in EGFR-mutant NCI-H1975 cells; subsequent assays included transcriptome profiling, measurements of intracellular Fe2⁺, malondialdehyde (MDA), glutathione (GSH), mitochondrial reactive oxygen species (ROS) levels, and ultrastructural examination by electron microscopy. Protein levels of NRF2, HO‑1, solute carrier family 7 member 11 (SLC7A11), glutathione peroxidase 4 (GPX4), and KEAP1 were assessed via western blot.
The ferroptosis gene–based random forest model distinguished EGFR-mutant LUAD with an area under the curve (AUC) of 0.84. Clinically, high AURKA expression was significantly associated with EGFR wild-type status (p = 0.035), reduced overall survival (p = 0.011), increased M1 macrophage infiltration, and decreased CD4⁺ T-cell infiltration. AURKA knockdown triggered hallmark features of ferroptosis—iron overload (p < 0.001), elevated MDA levels (p < 0.01), increased lipid peroxidation (p < 0.05), heightened mitochondrial ROS (p < 0.05), reduced mitochondrial membrane potential, GSH depletion (p < 0.05), and disruption of mitochondrial cristae. Mechanistically, loss of AURKA decreased KEAP1 (p < 0.01) and enhanced NRF2 (p < 0.001) and HO-1 (p < 0.01) and NRF2 nuclear translocation, while downregulating SLC7A11 (p < 0.05) and GPX4 (p < 0.01). Cell cycle analysis revealed G1-phase arrest (p < 0.001).
Our findings demonstrate that AURKA promotes ferroptosis resistance in EGFR-mutant LUAD by modulating the KEAP1/NRF2/HO-1 axis. Notably, this effect was validated in the gefitinib-resistant EGFR T790M-mutant NCI-H1975 cell model. Our results highlight AURKA as a potential therapeutic target for overcoming epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) resistance and suggest that disrupting the AURKA/KEAP1/NRF2/HO‑1 pathway may offer a novel strategy for treating EGFR-mutant LUAD.
After spinal cord injury (SCI), pro-inflammatory microglia accumulate and impede axonal regeneration. We explored whether secreted protein acidic and rich in cysteine (Sparc) restrains microglial inflammation and fosters neurite outgrowth.
Mouse microglial BV2 cells were polarized to a pro-inflammatory phenotype with lipopolysaccharides (LPSs). Sparc mRNA and protein were quantified by reverse transcription quantitative PCR (RT-qPCR). Sparc was overexpressed via plasmid transfection, then inflammatory cytokines, mitochondrial membrane potential (Δψm), reactive oxygen species (ROS), and oxidative-phosphorylation proteins, including voltage-dependent anion channel 1 (VDAC1), cytochrome c oxidase subunit 1 (COX1), and ATP synthase α subunit (ATP5A), were assayed by Western blot, enzyme-linked immunosorbent assay (ELISA), and flow cytometry. Immunoprecipitation plus mass spectrometry, co-immunoprecipitation, and immunofluorescence confirmed the interaction between Sparc and ubiquitin A-52 residue ribosomal protein fusion product 1 (Uba52). Effects of Sparc overexpression alone or combined with Uba52 small interfering RNA (si-Uba52) were compared in LPS-induced BV2 cells. Finally, BV2 cells and a mouse hippocampal neuron (HT-22) were co-cultured in the Transwell chamber, and the changes in proliferation, apoptosis, and III-tubulin content of the latter were detected.
In LPS-induced BV2 cells, the tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and ROS levels were elevated, while the IL-10 and transforming growth factor-β (TGF-β) levels, Δψm, and the proteins levels of the VDAC1, COX1, ATP5A, and Sparc decreased. Sparc overexpression reversed these changes. Mechanistically, Sparc bound Uba52 and upregulated its expression; Uba52 knockdown abolished the anti-inflammatory and mitochondrial-protective effects of Sparc. In co-culture, Sparc overexpression rescued HT-22 neurons apoptosis and enhanced axonal growth, but the effects were also reversed by Uba52 knockdown.
Sparc may maintain mitochondrial homeostasis by interacting with Uba52 to inhibit LPS-induced BV2 inflammatory response, thereby promoting neuronal axonal regeneration. This suggests that Sparc may play a potential role in SCI repair.
Sarcopenia is a complex, multifactorial condition characterized by progressive loss of muscle mass, strength, and function. Despite growing awareness, the early diagnosis and pathophysiological characterization of this condition remain challenging due to the lack of integrative biomarkers.
This study aimed to conduct a comprehensive multilevel profiling of clinical parameters, immune cell phenotypes, extracellular vesicle (EV) signatures, and biochemical markers to elucidate biological gradients associated with different stages of sarcopenia.
A prospective cohort study enrolled adults aged 45–85 years classified as control, presarcopenic, or sarcopenic based on European Working Group on Sarcopenia in Older People 2 (EWGSOP2) criteria. Clinical evaluation included anthropometry, muscle strength, sarcopenia screening (SARC-F) questionnaire/Short Physical Performance Battery (SPPB) questionnaires, and quality-of-life assessment. Flow cytometry was used to characterize blood monocyte/macrophage subsets (cluster of differentiation 14 (CD14), CD68, CD163, CD206). EVs were isolated from plasma and profiled for surface tetraspanins and matrix metalloproteinases (MMP2, MMP9, tissue inhibitor of metalloproteinase-1 (TIMP-1)) using bead-based flow cytometry. Biochemical assays measured metabolic, inflammatory, and extracellular matrix (ECM)-related markers. Data were analyzed via Kruskal–Wallis testing, discriminant analysis, and principal component analysis (PCA).
Sarcopenia, a muscle-wasting condition linked to aging, is characterized by chronic inflammation, proteolytic imbalance, and metabolic disturbances. Clinical deterioration is evident through reduced appendicular lean mass (ALM), appendicular skeletal muscle index (ASMI), SPPB scores, and sarcopenia quality of life (SarQoL) domains. Principal component analysis (PCA) identified four functional marker clusters: ECM degradation (MMP-positive EVs), inflammatory and homeostasis-stabilizing macrophages, and metabolic disruption (glucose, asprosin, triglycerides). Discriminant analysis emphasized vesicular and immune markers with significant classification potential, even when univariate differences were non-significant. Metabolic destabilization and inflammatory activation are detectable in presarcopenia stages. Chronic inflammation, characterized by CD14–CD163+206+ cells releasing pro-inflammatory cytokines, accelerates muscle degradation. Proteolytic dysfunction, with an imbalance between proteases and inhibitors, further contributes to muscle loss. Metabolic disorders impair energy production and nutrient utilization, exacerbating muscle wasting. A comprehensive assessment, including anthropometric, functional, physical activity, and QoL measures, is crucial for identifying high-risk individuals and understanding sarcopenia’s mechanisms. Vesicular biomarkers, regulating tissue remodeling and inflammation, provide valuable insights. Standardized assessment methods are essential for enhancing diagnostic accuracy and intervention effectiveness. Future research should focus on developing and refining biomarkers to improve specificity and sensitivity, enabling targeted therapies and better QoL.
Integrating clinical, immunological, and biochemical markers with EVs helps stratify sarcopenia effectively. Our data shows that EVs and macrophage profiles reflect systemic changes and metabolic stress. However, age- and gender-related variability in our cohort warrants caution in generalizing the findings. Artificial intelligence (AI) enhances patient clustering by combining these data types, enabling precise, personalized sarcopenia management, predicting disease progression, and identifying high-risk patients. AI also standardizes and optimizes analytical protocols, improving diagnostic and monitoring reliability and reproducibility.
Lysosomes serve not only in the degradation of cellular components but also as calcium (Ca2+) stores. In this study, we investigated the effects of trans-Ned19, an inhibitor of lysosomal calcium channels known to block two-pore channels (TPCs), on fertilization and oocyte activation in mice.
Pronuclear formation was assessed via Hoechst 33342 staining, cortical granule release was evaluated using Lens culinaris agglutinin-fluorescein isothiocyanate (LCA-FITC) staining, intracellular Ca2+ levels were monitored with Cal-520 AM, and sperm motility was analyzed using a sperm motility analysis system (SMAS).
In strontium (Sr2+)-induced oocyte activation, trans-Ned19 significantly reduced pronuclear formation at 8 h post-activation. Cortical granule release and Ca2+ oscillations were also markedly suppressed. In contrast, during in vitro fertilization (IVF), trans-Ned19 treatment significantly decreased the fertilization rate; however, pronuclear formation and cortical granule release remained comparable to controls in fertilized embryos. Notably, when IVF was performed using zona pellucida-free oocytes, the fertilization rate in the trans-Ned19 group was similar to that of the controls. However, a significant increase in polyspermy was observed. Furthermore, trans-Ned19 significantly impaired sperm motility parameters, including straight-line velocity, curvilinear velocity, and average path velocity.
These findings suggest that lysosomal TPCs are essential for both normal fertilization and artificial oocyte activation in mice.
Neonatal jaundice affects up to 60% of newborns, with pathological cases frequently associated with impaired bilirubin metabolism and gut microbiota dysbiosis. Although evidence implicates gut microbiota in bilirubin metabolism, the precise mechanisms remain incompletely characterized. This study investigated treatment-associated changes in gut microbiota composition, fecal metabolites, and liver function in neonates with hyperbilirubinemia.
A total of forty-two neonates diagnosed with hyperbilirubinemia were recruited. Fecal samples were collected pre- and post-treatment. Gut microbiota composition was analyzed via 16S rRNA gene sequencing, while fecal metabolites were profiled using untargeted metabolomics. Liver function parameters, including serum bilirubin levels, were measured. Statistical analyses encompassed alpha/beta diversity assessments, Spearman correlation, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment.
Post-treatment gut microbial diversity decreased significantly, marked by increased abundance of Streptococcus thermophilus and Rothia and reduced levels of Pseudomonas and Staphylococcus. Key altered metabolites included 9,11-methane-epoxy PGF1α, prostaglandin E2 isopropyl ester, and 7-methylthioheptyl glucosinolate. Notably, Streptococcus thermophilus abundance inversely correlated with 7-methylthioheptyl glucosinolate but positively correlated with 9,11-methane-epoxy PGF1α and prostaglandin E2 isopropyl ester. Total bilirubin levels decreased significantly post-treatment, alongside improvements in other liver function markers.
This study demonstrates significant treatment-associated shifts in gut microbiota and metabolites in hyperbilirubinemic neonates, suggesting microbial dysbiosis may contribute to altered bilirubin metabolism. These findings highlight the potential of early microbiome-targeted interventions for managing neonatal jaundice and identify candidate therapeutic targets and biomarkers.
Membrane transport proteins are critical determinants of systemic and intracellular drug levels, thereby contributing substantially to drug response and/or adverse drug reactions. Therefore, the U.S. Food and Drug Administration and the European Medicines Agency, the regulatory authorities for drug approval in the U.S. and Europe, respectively, recommend pre-clinical testing of selected drug transporters during the drug development process to elucidate clinically relevant drug–drug interactions (DDIs). In the current proof-of-principle study, we describe the generation of fruit flies expressing human membrane drug transporters in their salivary glands to enable DDI studies in a time-saving manner and at low costs.
Using the Gal4/upstream activation sequence (UAS) expression system, we established fruit flies expressing human organic cation transporters (hOCTs) 1 and 2 and genetic variants thereof. Both transporters are key drug uptake transporters in humans and are recommended for pre-clinical DDI studies. After injecting fluorescent hOCT substrates, their accumulation in salivary gland cells was observed by confocal laser scanning microscopy.
We demonstrate the feasibility of expressing hOCT1 and hOCT2 in the salivary glands of fruit fly embryos and subsequent alteration by clinically relevant genetic variants, corroborating results from mammalian cell experiments. Moreover, we show an OCT-dependent accumulation of the prototypic fluorescent OCT substrates ethidium (Et+) and 4-Di-1-ASP (4-(4-(dimethylamino)styryl)-N-methylpyridinium, ASP+) in the salivary gland cells and subsequent inhibition by clinically-used OCT drug inhibitors.
Based on the handling procedure and the lack of need for Animal Protection Act approval, we propose that the humanized Drosophila melanogaster fruit fly model opens a new avenue for pre-clinical functional transporter studies.
Glioblastoma (GBM) is an extremely aggressive brain tumor, marked by restricted therapeutic possibilities and a generally unfavorable prognosis. GBM’s complexity and heterogeneity necessitate comprehensive genetic and immunological profiling to enhance therapeutic strategies.
The study integrated The Cancer Genome Atlas (TCGA) and Integrative Epidemiology Unit Open Genome-Wide Association Studies (IEU OpenGWAS) data to identify genetic factors influencing GBM using expression quantitative trait loci (eQTL) and genome-wide association studies (GWAS). Mendelian randomization (MR) analysis revealed 250 GBM-associated genes. A GBM risk prediction model was built using Least Absolute Shrinkage and Selection Operator (LASSO) and Cox regression. The research examined immune infiltration, drug response, and mutation profiles to characterize GBM molecular features. Functional enrichment and in vitro experiments validated key findings.
The analysis uncovered significant genetic associations with GBM, emphasizing key genes such as follistatin-like 1 (FSTL1), FXYD domain-containing ion transport regulator 5 (FXYD5), Ras-related protein (RRAS), and ring finger protein 216 pseudogene 1 (RNF216P1). The risk model effectively categorized patients into low-risk and high-risk groups, showing significantly worse survival outcomes in the high-risk group. Immune profiling revealed differential infiltration of cancer-associated fibroblasts (CAFs), macrophages, and T cells, which correlated with the expression levels of the genes that were identified. Patients at high risk showed increased sensitivity to chemotherapeutic drugs such as dasatinib and lapatinib, while those at low risk were more responsive to elesclomol and lisitinib. Notably, key genes such as DCMP Deaminase (DCTD) and RRAS were found to regulate ferroptosis, underscoring their potential as therapeutic targets for GBM treatment.
This study deepens the understanding of GBM by pinpointing critical genetic markers and elucidating their influence on the tumor immune microenvironment (TME) as well as treatment response. The risk model developed in this study holds promise for enhancing prognostic accuracy and facilitating the personalization of GBM therapy.
The global increase in diabetes mellitus has been accompanied by a significant rise in related complications. Diabetic patients frequently experience ocular surface disorders and multiple studies have demonstrated that the diabetic corneal epithelium is characterized by increased cellular fragility and compromised barrier integrity. It has been demonstrated that the processes of oxidative stress and inflammation are pivotal in causing ocular tissue damage in diabetic patients. Numerous studies have explored the protective effects of various antioxidants, especially those sourced from plants. Cynara cardunculus L. var. altilis (DC.), a species widely integrated into the Mediterranean diet and commonly known as cultivated cardoon (CC), is particularly rich in bioactive phenolic compounds, recognized for their antioxidant effects.
The current work focuses on assessing the effect of CC leaf extracts on high glucose-treated human corneal epithelial cells (HCEpiCs). HCEpiCs were cultured for 24 h in a medium supplemented with glucose up to a concentration of 25 mM. Mannitol treatment was included to distinguish whether the observed effects were due to glucose metabolism or solely osmotic stress. To evaluate the effect of CC extracts, corneal cells were pre-incubated with the CC extract 10–20 μg/mL for 24 h before high glucose (HG) treatment. Cell viability, transepithelial electrical resistance, wound healing assay and reactive oxygen species (ROS) measurements were performed after HG treatment. To evaluate the levels of oxidative stress, the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), along with glutathione (GSH) levels were assayed. The mitogen-activated protein kinase ERK1/2/cytosolic phospholipases A2 (cPLA2)/cyclooxygenase-2 (COX-2) signaling pathway, triggering the inflammatory process, was evaluated by western blot analyses.
Our findings indicate that CC extract (i) improved viability, reducing oxidative stress by lowering ROS production and enhancing SOD, CAT activities and GSH content in human corneal epithelial cells exposed to high glucose concentrations; (ii) maintained a high TEER and promoted corneal epithelial wound healing; and (iii) induced down-regulation of the ERK 1/2/cPLA2/COX-2 signaling pathway involved in the inflammatory process and ROS production.
CC leaf extract could improve human corneal epithelial cell function suggesting its potential as a preventive agent against the development of chronic diabetic complications.
Obesity is a chronic condition linked to health issues such as diabetes, heart disease, and increased cancer risk. High body mass index (BMI) is associated with cancers such as breast and colorectal cancer due to hormone imbalances and inflammation from excess fat, whereas a low BMI can raise cancer risk by weakening the immune system. Maintaining a normal BMI improves cancer treatment outcomes, but in some cases, higher BMI might offer protective effects—a phenomenon known as the “obesity paradox”. This study explores how BMI affects gene expression in cancer, using data from The Cancer Genome Atlas (TCGA), aiming to uncover links between BMI and cancer progression while identifying potential treatment targets.
To analyze the data, a two-stage method using overlapping group screening (OGS) was applied. First, gene groups were identified with the “grpregOverlap” R package. Then, their interactions were tested using the sequence kernel association test. Significant gene-gene interactions were selected based on statistical measures. In the second stage, predictive models were built using regularized regression techniques such as ridge regression, lasso, and adaptive lasso, with generalized ridge regression used to improve accuracy and stability in handling high-dimensional data.
The proposed OGS-based method was tested on simulated and real-world datasets. Results showed that combining OGS with generalized ridge regression and adaptive lasso (OGS_G.ridge_ALasso) gave the best prediction performance, with lower error rates and greater stability compared to other models like support vector regression, k-nearest neighbors, and random forests. In practical applications, gene expression and BMI data from TCGA patients (including bladder, cervical, esophageal and liver cancers) were integrated to identify key genes and interactions related to BMI.
Through evaluations on both simulated synthetic datasets and real-world datasets, we demonstrated the effectiveness of the proposed method in terms of predictive accuracy. Additionally, we identified BMI-associated genes and gene-gene interaction biomarkers across different cancer types and presented the corresponding network structures. Based on the key genes and gene interactions identified, we further explored how BMI influences cancer development and prognosis, providing deeper insights into the biological mechanisms underlying these associations.
Zhang et al.’s recent article utilizes comprehensive single-cell data to identify differences in tumor cell populations, highlighting the CKS1B+ malignant cell subcluster as a potential target for immunotherapy. It develops a prognostic and immunotherapeutic signature (PIS) based on this subcluster, demonstrating good performance in predicting lung adenocarcinoma (LUAD) prognosis. The study also validates the role of PSMB7 in LUAD progression. However, there are areas for improvement. There is a lack of clarity regarding the relationship between the CKS1B+ malignant cell subcluster and the PIS, particularly in terms of why PSMB7 was selected for functional studies. The sequencing data are retrospectively obtained from public databases and lack prospective clinical validation. It is suggested to collect LUAD patient tissues for RT-qPCR and RNA-seq analysis and seek external multi-center validations. Additionally, integrating emerging multi-omics methods is recommended to further validate the findings. Despite these limitations, the study represents progress in understanding LUAD and treatment strategies, and continuous evaluation and refinement of multi-omics and machine learning methods are expected for future research and clinical practice.
Vitamin D decomposition products target a myristic acid sidechain of the predominant glycerophospholipid constructed in the biomembranes of Helicobacter pylori, causing gastric cancer in humans, and disrupt the membrane structure, followed by bacteriolysis. No earlier studies, however, elucidate whether vitamin D decomposition products interact with the glycerophospholipids that construct the eukaryotic biomembranes and confer whatever cell disorders.
A gastric cancer cell line, MKN45, and a non-cancer cell line, Vero, were used in this study. Cell injury activities of vitamin D decomposition products (VDP1 and VD2-1) and a VDP1 derivative (VD3-7) were examined by the 3-(4,5-di-methylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Identification of a glycerophospholipid was performed by 1H-nuclear magnetic resonance (NMR). Fatty acid composition and glycerophospholipid molecular species were analyzed by gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/mass spectrometry (LC/MS), respectively. Structure collapse-induction activity of VDP1, VD2-1 and VD3-7 to glycerophospholipid vesicles was examined using the pigment-containing lamellar vesicles.
MKN45 cells exhibited higher susceptibility to the cell injury activity of VDP1 and VD2-1 than Vero cells. In the analysis of biomembrane lipids, the glycerophospholipid phosphatidylcholine (PC) molecular species turned out to conspicuously differ between MKN45 cells and Vero cells. Contents of myristoyl-PC were higher in MKN45 cells than in Vero cells, while contents of oleoyl-PC were higher in Vero cells than in MKN45 cells. Meanwhile, the contents of palmitoyl- and palmitoleoyl-PC were almost equal between these cells. We next examined the structure collapse-induction activity of VDP1, VD2-1 and VD3-7 on the lamellar vesicles prepared with dimyristoyl-PC, dipalmitoyl-PC and dioleoyl-PC. The vitamin D decomposition products and a VDP1 derivative induced the structural collapse of dimyristoyl- and dipalmitoyl-PC lamellar vesicles but almost no structural collapse of dioleoyl-PC lamellar vesicles. These results suggest that the contents of myristoyl-PC in biomembranes are associated with the susceptibility of eukaryotic cells to the cell injury activity of VDP1, VD2-1 and VD3-7. In addition, no VD3-7 affected the viability of Vero cells and selectively decreased the viability of MKN45 cells.
In the future, we will expect to be capable of developing novel antitumor agents targeting the myristic acid sidechain of biomembranal PC using a vitamin D decomposition product as the fundamental structure.