International Journal of Molecular Sciences

16 pages, 1560 KiB

Open AccessArticle

Electromagnetic Transduction Therapy (EMTT) Enhances Tenocyte Regenerative Potential: Evidence for Senolytic-like Effects and Matrix Remodeling

by Matteo Mancini, Mario Vetrano, Alice Traversa, Carlo Cauli, Simona Ceccarelli, Florence Malisan, Maria Chiara Vulpiani, Nicola Maffulli, Cinzia Marchese, Vincenzo Visco and Danilo Ranieri

Int. J. Mol. Sci. 2025, 26(15), 7122; https://doi.org/10.3390/ijms26157122 (registering DOI) - 24 Jul 2025

Abstract

Tendinopathies are a significant challenge in musculoskeletal medicine, with current treatments showing variable efficacy. Electromagnetic transduction therapy (EMTT) has emerged as a promising therapeutic approach, but its biological effects on tendon cells remain largely unexplored. Here, we investigated the effects of EMTT on [...] Read more.

Tendinopathies are a significant challenge in musculoskeletal medicine, with current treatments showing variable efficacy. Electromagnetic transduction therapy (EMTT) has emerged as a promising therapeutic approach, but its biological effects on tendon cells remain largely unexplored. Here, we investigated the effects of EMTT on primary cultured human tenocytes’ behavior and functions in vitro, focusing on cellular responses, senescence-related pathways, and molecular mechanisms. Primary cultures of human tenocytes were established from semitendinosus tendon biopsies of patients undergoing anterior cruciate ligament (ACL) reconstruction (n = 6, males aged 17–37 years). Cells were exposed to EMTT at different intensities (40 and 80 mT) and impulse numbers (1000–10,500). Cell viability (MTT assay), proliferation (Ki67), senescence markers (CDKN2a/INK4a), migration (scratch test), cytoskeleton organization (immunofluorescence), and gene expression (RT-PCR) were analyzed. A 40 mT exposure elicited minimal effects, whereas 80 mT treatments induced significant cellular responses. Repeated 80 mT exposure demonstrated a dual effect: despite a moderate decrease in overall cell vitality, increased Ki67 expression (+7%, p ≤ 0.05) and significant downregulation of senescence marker CDKN2a/INK4a were observed, suggesting potential senolytic-like activity. EMTT significantly enhanced cell migration (p < 0.001) and triggered cytoskeletal remodeling, with amplified stress fiber formation and paxillin redistribution. Molecular analysis revealed upregulation of tenogenic markers (Scleraxis, Tenomodulin) and enhanced Collagen I and III expressions, particularly with treatments at 80 mT, indicating improved matrix remodeling capacity. EMTT significantly promotes tenocyte proliferation, migration, and matrix production, while simultaneously exhibiting senolytic-like effects through downregulation of senescence-associated markers. These results support EMTT as a promising therapeutic approach for the management of tendinopathies through multiple regenerative mechanisms, though further studies are needed to validate these effects in vivo. Full article

(This article belongs to the Collection Feature Papers in Molecular Pathology, Diagnostics, and Therapeutics)

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16 pages, 6868 KiB

Open AccessArticle

Unnatural Amino Acid Photo-Crosslinking Sheds Light on Gating of the Mechanosensitive Ion Channel OSCA1.2

by Scarleth Duran-Morales, Rachel Reyes-Lizana, German Fernández, Macarena Loncon-Pavez, Yorley Duarte, Valeria Marquez-Miranda and Ignacio Diaz-Franulic

Int. J. Mol. Sci. 2025, 26(15), 7121; https://doi.org/10.3390/ijms26157121 - 23 Jul 2025

Abstract

Mechanosensitive ion channels such as OSCA1.2 enable cells to sense and respond to mechanical forces by translating membrane tension into ionic flux. While lipid rearrangement in the inter-subunit cleft has been proposed as a key activation mechanism, the contributions of other domains to [...] Read more.

Mechanosensitive ion channels such as OSCA1.2 enable cells to sense and respond to mechanical forces by translating membrane tension into ionic flux. While lipid rearrangement in the inter-subunit cleft has been proposed as a key activation mechanism, the contributions of other domains to OSCA gating remain unresolved. Here, we combined the genetic encoding of the photoactivatable crosslinker p-benzoyl-L-phenylalanine (BzF) with functional Ca²⁺ imaging and molecular dynamics simulations to dissect the roles of specific residues in OSCA1.2 gating. Targeted UV-induced crosslinking at positions F22, H236, and R343 locked the channel in a non-conducting state, indicating their functional relevance. Structural analysis revealed that these residues are strategically positioned: F22 interacts with lipids near the activation gate, H236 lines the lipid-filled cavity, and R343 forms cross-subunit contacts. Together, these results support a model in which mechanical gating involves a distributed network of residues across multiple channel regions, allosterically converging on the activation gate. This study expands our understanding of mechanotransduction by revealing how distant structural elements contribute to force sensing in OSCA channels. Full article

(This article belongs to the Special Issue Ion Channels as a Potential Target in Pharmaceutical Designs 2.0)

18 pages, 10699 KiB

Open AccessArticle

Eurycomanone Blocks TGF-β1-Induced Epithelial-to-Mesenchymal Transition, Migration, and Invasion Pathways in Human Non-Small Cell Lung Cancer Cells by Targeting Smad and Non-Smad Signaling

by Pratchayanon Soddaen, Kongthawat Chairatvit, Pornsiri Pitchakarn, Tanongsak Laowanitwattana, Arisa Imsumran and Ariyaphong Wongnoppavich

Int. J. Mol. Sci. 2025, 26(15), 7120; https://doi.org/10.3390/ijms26157120 - 23 Jul 2025

Abstract

Non-small cell lung cancer (NSCLC) is a predominant form of lung cancer that is often diagnosed at an advanced metastatic stage. The processes of cancer cell migration and invasion involve epithelial-to-mesenchymal transition (EMT), which is crucial for metastasis. Targeting cancer aggressiveness with effective [...] Read more.

Non-small cell lung cancer (NSCLC) is a predominant form of lung cancer that is often diagnosed at an advanced metastatic stage. The processes of cancer cell migration and invasion involve epithelial-to-mesenchymal transition (EMT), which is crucial for metastasis. Targeting cancer aggressiveness with effective plant compounds has gained attention as a potential adjuvant therapy. Eurycomanone (ECN), a bioactive quassinoid found in the root of Eurycoma longifolia Jack, has demonstrated anti-cancer activity against various carcinoma cell lines, including human NSCLC cells. This study aimed to investigate the in vitro effects of ECN on the migration and invasion of human NSCLC cells and to elucidate the mechanisms by which ECN modulates the EMT in these cells. Non-toxic doses (≤IC20) of ECN were determined using the MTT assay on two human NSCLC cell lines: A549 and Calu-1. The results from wound healing and transwell migration assays indicated that ECN significantly suppressed the migration of both TGF-β1-induced A549 and Calu-1 cells. ECN exhibited a strong anti-invasive effect, as its non-toxic doses significantly suppressed the TGF-β1-induced invasion of NSCLC cells through Matrigel and decreased the secretion of MMP-2 from these cancer cells. Furthermore, ECN could affect the TGF-β1-induced EMT process in various ways in NSCLC cells. In TGF-β1-induced A549 cells, ECN significantly restored the expression of E-cadherin by inhibiting the Akt signaling pathway. Conversely, in Calu-1, ECN reduced the aggressive phenotype by decreasing the expression of the mesenchymal protein N-cadherin and inhibiting the TGF-β1/Smad pathway. In conclusion, this study demonstrated the anti-invasive activity of eurycomanone from E. longifolia Jack in human NSCLC cells and provided insights into its mechanism of action by suppressing the effects of TGF-β1 signaling on the EMT program. These findings offer scientific evidence to support the potential of ECN as an alternative therapy for metastatic NSCLC. Full article

(This article belongs to the Special Issue Natural Products with Anti-Inflammatory and Anticancer Activity)

34 pages, 2003 KiB

Open AccessReview

Docetaxel Resistance in Breast Cancer: Current Insights and Future Directions

by Fátima Postigo-Corrales, Asunción Beltrán-Videla, Antonio David Lázaro-Sánchez, Ana María Hurtado, Pablo Conesa-Zamora, Ana Belén Arroyo and Ginés Luengo-Gil

Int. J. Mol. Sci. 2025, 26(15), 7119; https://doi.org/10.3390/ijms26157119 - 23 Jul 2025

Abstract

Docetaxel is a chemotherapeutic agent widely used for breast cancer treatment; however, its efficacy is often limited by drug resistance and associated toxicity. This review examines the molecular mechanisms of docetaxel resistance in breast cancer and discusses research advances and future directions for [...] Read more.

Docetaxel is a chemotherapeutic agent widely used for breast cancer treatment; however, its efficacy is often limited by drug resistance and associated toxicity. This review examines the molecular mechanisms of docetaxel resistance in breast cancer and discusses research advances and future directions for overcoming this challenge. Key resistance mechanisms include alterations in drug targets (microtubules), increased drug efflux, suppression of apoptosis, activation of survival signalling pathways, epithelial-to-mesenchymal transition (EMT), and cancer stem cell enrichment. An evolutionary perspective distinguishes between intrinsic and acquired resistance, emphasising the need for adaptive therapeutic strategies. Recent advances in genomic profiling, non-coding RNA research, novel drug combinations, and biomarker-guided therapies have also been reviewed. Emerging approaches, such as targeting the tumour microenvironment, harnessing immunotherapy, and implementing adaptive dosing schedules, have been discussed. This review emphasises the understanding of resistance as a multifactorial phenomenon that requires multipronged interventions. Research has aimed to identify predictive biomarkers, develop targeted agents to reverse resistance, and design rational combination strategies to improve patient outcomes. Progress in deciphering and targeting docetaxel resistance mechanisms holds promise for enhancing treatment responses and extending survival in patients with breast cancer. Full article

(This article belongs to the Special Issue Molecular Research and Cellular Biology of Breast Cancer)

23 pages, 2486 KiB

Open AccessArticle

Comparative Transcriptomic Analysis for Identification of Environmental-Responsive Genes in Seven Species of Threadfin Breams (Nemipterus)

by Zhaoke Dang, Qiaer Wu, Yanbo Zhou, Liangming Wang, Yan Liu, Changping Yang, Manting Liu, Qijian Xie, Cheng Chen, Shengwei Ma and Binbin Shan

Int. J. Mol. Sci. 2025, 26(15), 7118; https://doi.org/10.3390/ijms26157118 - 23 Jul 2025

Abstract

Members of the genus Nemipterus are economically important fish species distributed in the tropical and subtropical Indo-West Pacific region. The majority of species in this genus inhabit waters with sandy–muddy substrates on the continental shelf, although different species are found at slightly varying [...] Read more.

Members of the genus Nemipterus are economically important fish species distributed in the tropical and subtropical Indo-West Pacific region. The majority of species in this genus inhabit waters with sandy–muddy substrates on the continental shelf, although different species are found at slightly varying water depths. In this study, we sequenced seven species within the genus Nemipterus after identifying the specimens using complementary morphological analysis and DNA barcoding. Each species yielded over 40,000,000 clean reads, totaling over 300,000,000 clean reads across the seven species. A total of 276,389 unigenes were obtained after de novo assembly and a total of 168,010 (60.79%) unigenes were annotated in the protein database. The comprehensive functional annotation based on the KOG, GO, and KEGG databases revealed that these unigenes are mainly associated with numerous physiological, metabolic, and molecular processes, and that the seven species exhibit similarity in these aspects. By constructing a phylogenetic tree and conducting divergence time analysis, we found that N. bathybius and N. virgatus diverged most recently, approximately during the Neogene Period (14.9 Mya). Compared with other species, N. bathybius and N. virgatus are distributed in deeper water layers. Therefore, we conducted selection pressure analysis using these two species as the foreground branches and identified several environmental-responsive genes. The results indicate that genes such as aqp1, arrdc3, ISP2, Hip, ndufa1, ndufa3, pcyt1a, ctsk, col6a2, casp2 exhibit faster evolutionary rates during long-term adaptation to deep-water environments. Specifically, these genes are considered to be associated with adaptation to aquatic osmoregulation, temperature fluctuations, and skeletal development. This comprehensive analysis provides valuable insights into the evolutionary biology and environmental adaptability of threadfin breams, contributing to the conservation and sustainable management of these species. Full article

(This article belongs to the Special Issue Molecular Biology and the Genetic Breeding of Aquatic Animals: Breakthroughs and Challenges)

21 pages, 8405 KiB

Open AccessArticle

Distinct Mitochondrial DNA Deletion Profiles in Pediatric B- and T-ALL During Diagnosis, Remission, and Relapse

by Hesamedin Hakimjavadi, Elizabeth Eom, Eirini Christodoulou, Brooke E. Hjelm, Audrey A. Omidsalar, Dejerianne Ostrow, Jaclyn A. Biegel and Xiaowu Gai

Int. J. Mol. Sci. 2025, 26(15), 7117; https://doi.org/10.3390/ijms26157117 - 23 Jul 2025

Abstract

Mitochondria are critical for cellular energy, and while large deletions in their genome (mtDNA) are linked to primary mitochondrial diseases, their significance in cancer is less understood. Given cancer’s metabolic nature, investigating mtDNA deletions in tumors at various stages could provide insights into [...] Read more.

Mitochondria are critical for cellular energy, and while large deletions in their genome (mtDNA) are linked to primary mitochondrial diseases, their significance in cancer is less understood. Given cancer’s metabolic nature, investigating mtDNA deletions in tumors at various stages could provide insights into disease origins and treatment responses. In this study, we analyzed 148 bone marrow samples from 129 pediatric patients with B-cell (B-ALL) and T-cell (T-ALL) acute lymphoblastic leukemia at diagnosis, remission, and relapse using long-range PCR, next-generation sequencing, and the Splice-Break2 pipeline. Both T-ALL and B-ALL exhibited significantly more mtDNA deletions than did the controls, with T-ALL showing a ~100-fold increase and B-ALL a ~15-fold increase. The T-ALL samples also exhibited larger deletions (median size > 2000 bp) and greater heterogeneity, suggesting increased mitochondrial instability. Clustering analysis revealed distinct deletion profiles between ALL subtypes and across disease stages. Notably, large clonal deletions were detected in some B-ALL remission samples, including one affecting up to 88% of mtDNA molecules, which points toward treatment-driven selection or toxicity. A multivariate analysis confirmed that disease type, timepoint, and WHO subtype significantly influenced mtDNA deletion metrics, while age and gender did not. These findings suggest that mtDNA deletion profiling could serve as a biomarker for pediatric ALL and may indicate mitochondrial toxicity contributing to late effects in survivors. Full article

(This article belongs to the Special Issue Mitochondrial Function in Human Health and Disease: 2nd Edition)

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55 pages, 1057 KiB

Open AccessReview

Rice Adaptation to Abiotic Stresses Caused by Soil Inorganic Elements

by Giulia Vitiello, Daniela Goretti, Caterina Marè, Edoardo Delmastro, Giorgia Siviero, Silvio Collani, Erica Mica and Giampiero Valè

Int. J. Mol. Sci. 2025, 26(15), 7116; https://doi.org/10.3390/ijms26157116 - 23 Jul 2025

Abstract

Soil contamination with toxic inorganic elements poses a major challenge to rice cultivation, affecting plant physiology, yield, and grain safety. While natural variation in tolerance exists among rice genotypes and related species, recent advances in genomics, breeding, and biotechnology offer new opportunities to [...] Read more.

Soil contamination with toxic inorganic elements poses a major challenge to rice cultivation, affecting plant physiology, yield, and grain safety. While natural variation in tolerance exists among rice genotypes and related species, recent advances in genomics, breeding, and biotechnology offer new opportunities to enhance adaptation. This review synthesizes the current knowledge on the physiological effects of toxic elements and explores strategies to improve tolerance, from harnessing genetic diversity to genome editing and transgenic approaches. Attention is also paid to the role of microbiota in mitigating toxicity and reducing translocation to seeds, highlighting emerging solutions for sustainable rice production in contaminated environments. Full article

(This article belongs to the Special Issue Plant Resilience: Insights into Abiotic and Biotic Stress Adaptations)

17 pages, 837 KiB

Open AccessArticle

Protective Effects of Regular Physical Activity: Differential Expression of FGF21, GDF15, and Their Receptors in Trained and Untrained Individuals

by Paulina Małkowska, Patrycja Tomasiak, Marta Tkacz, Katarzyna Zgutka, Maciej Tarnowski, Agnieszka Maciejewska-Skrendo, Rafał Buryta, Łukasz Rosiński and Marek Sawczuk

Int. J. Mol. Sci. 2025, 26(15), 7115; https://doi.org/10.3390/ijms26157115 - 23 Jul 2025

Abstract

According to the World Health Organization (WHO), a healthy lifestyle is defined as a way of living that lowers the risk of becoming seriously ill or dying prematurely. Physical activity, as a well-known contributor to overall health, plays a vital role in supporting [...] Read more.

According to the World Health Organization (WHO), a healthy lifestyle is defined as a way of living that lowers the risk of becoming seriously ill or dying prematurely. Physical activity, as a well-known contributor to overall health, plays a vital role in supporting such a lifestyle. Exercise induces complex molecular responses that mediate both acute metabolic stress and long-term physiological adaptations. FGF21 (fibroblast growth factor 21) and GDF15 (growth differentiation factor 15) are recognized as metabolic stress markers, while their receptors play critical roles in cellular signaling. However, the differential gene expression patterns of these molecules in trained and untrained individuals following exhaustive exercise remain poorly understood. This study aimed to examine the transcriptional and protein-level responses in trained and untrained individuals performed a treadmill maximal exercise test to voluntary exhaustion. Blood samples were collected at six time points (pre-exercise, immediately post-exercise, and 0.5 h, 6 h, 24 h, and 48 h post-exercise). Gene expression of FGF21, GDF15, FGFR1 (fibroblast growth factor receptors), FGFR3, FGFR4, KLB (β-klotho), and GFRAL (glial cell line-derived neurotrophic factor receptor alpha-like) was analyzed using RT-qPCR, while plasma protein levels of FGF21 and GDF15 were quantified via ELISA. The results obtained were statistically analyzed by using Shapiro–Wilk, Mann–Whitney U, and Wilcoxon tests in Statistica 13 software. Untrained individuals demonstrated significant post-exercise upregulation of FGFR3, FGFR4, KLB, and GFRAL. FGF21 and GDF15 protein levels were consistently lower in trained individuals (p < 0.01), with no significant correlations between gene and protein expression. Trained individuals showed more stable expression of genes, while untrained individuals exhibited transient upregulation of genes after exercise. Full article

(This article belongs to the Special Issue Cytokines in Inflammation and Health)

15 pages, 1896 KiB

Open AccessCase Report

Pathogenesis of Cardiac Valvular Hemangiomas: A Case Report and Literature Review

by Kimberly-Allisya Neeter, Catalin-Bogdan Satala, Daniela Mihalache, Alexandru-Stefan Neferu, Gabriela Patrichi, Carmen Elena Opris and Simona Gurzu

Int. J. Mol. Sci. 2025, 26(15), 7114; https://doi.org/10.3390/ijms26157114 - 23 Jul 2025

Abstract

Valvular hemangiomas are uncommon vascular anomalies that appear on the surface of heart valves. They can cause an array of non-specific symptoms and are consequently rarely diagnosed, with only 31 such cases (including the present one) reported to date in the literature; the [...] Read more.

Valvular hemangiomas are uncommon vascular anomalies that appear on the surface of heart valves. They can cause an array of non-specific symptoms and are consequently rarely diagnosed, with only 31 such cases (including the present one) reported to date in the literature; the present case is the first report of an arteriovenous hemangioma with a tricuspid localization. During the preoperative echocardiographic examination for a ventricular septal defect, a mass was incidentally discovered on the tricuspid valve of a 9-month-old infant. The involved leaflet was surgically removed and sent to the pathology department for analysis and subsequently diagnosed as an arteriovenous hemangioma. The patient recovered well, with no local tumor recurrence or other complications. The microscopic examination showed multiple blood vessels which stained positive for the endothelial markers CD31 and CD34 and which did not express D2-40, normally found in lymphatic endothelia. Surprisingly, endothelial cells lining the vessels also showed positivity for SMA, a mesenchymal cell marker, indicating a possible involvement of endothelial-to-mesenchymal transition and its opposite process, mesenchymal-to-endothelial transition, in the pathogenesis of these vascular anomalies. Full article

(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)

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28 pages, 3757 KiB

Open AccessArticle

Growth Hormone Signaling in Bladder Cancer: Transcriptomic Profiling of Patient Samples and In Vitro Evidence of Therapy Resistance via ABC Transporters and EMT Activation

by Emily Davis, Lydia J. Caggiano, Hannah Munholland, Reetobrata Basu, Darlene E. Berryman and John J. Kopchick

Int. J. Mol. Sci. 2025, 26(15), 7113; https://doi.org/10.3390/ijms26157113 - 23 Jul 2025

Abstract

Growth hormone (GH) signaling has been implicated in tumor progression and therapy resistance across multiple cancer types, yet its role in bladder cancer remains largely unexplored. In this study, we investigated the impact of GH and its receptor (GHR) on therapy resistance and [...] Read more.

Growth hormone (GH) signaling has been implicated in tumor progression and therapy resistance across multiple cancer types, yet its role in bladder cancer remains largely unexplored. In this study, we investigated the impact of GH and its receptor (GHR) on therapy resistance and disease progression in urothelial carcinoma (UC) through integrated transcriptomic and in vitro analyses. Transcriptomic profiling of The Cancer Genome Atlas bladder cancer cohort revealed that high tumoral GHR expression was associated with differential upregulation of genes involved in drug efflux, epithelial-to-mesenchymal transition (EMT), and extracellular matrix (ECM) remodeling. Notably, elevated GHR levels correlated with significantly reduced overall survival in patients with UC. In parallel, in vitro experiments demonstrated that GH promotes chemoresistance in UC cell lines via upregulation of ATP-binding cassette-containing (ABC) transporters and activation of EMT. GH also modulated ECM-remodeling-associated genes in a chemotherapy-dependent manner, including matrix metalloproteinases and tissue inhibitors of metalloproteinases. Importantly, these effects were abrogated by Pegvisomant, a GHR antagonist, indicating the functional relevance of GH/GHR signaling in the mediation of these phenotypes. Collectively, our findings support a mechanistic role for GH signaling in driving therapy resistance and tumor aggressiveness in bladder cancer and suggest GHR antagonism as a potential therapeutic strategy to improve treatment outcomes. Full article

(This article belongs to the Special Issue Urologic Cancers: Molecular Basis for Novel Therapeutic Approaches)

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23 pages, 4192 KiB

Open AccessArticle

Efficacy of Various Complexing Agents for Displacing Biologically Important Ligands from Eu(III) and Cm(III) Complexes in Artificial Body Fluids—An In Vitro Decorporation Study

by Sebastian Friedrich, Antoine Barberon, Ahmadabdurahman Shamoun, Björn Drobot, Katharina Müller, Thorsten Stumpf, Jerome Kretzschmar and Astrid Barkleit

Int. J. Mol. Sci. 2025, 26(15), 7112; https://doi.org/10.3390/ijms26157112 - 23 Jul 2025

Abstract

Incorporation of lanthanide (Ln) and actinide (An) ions into the human body poses significant chemotoxic and radiotoxic risks, necessitating effective decorporation strategies. This study investigates the displacement of biologically relevant ligands from trivalent ions of europium, Eu(III), and curium, Cm(III), in artificial biofluids [...] Read more.

Incorporation of lanthanide (Ln) and actinide (An) ions into the human body poses significant chemotoxic and radiotoxic risks, necessitating effective decorporation strategies. This study investigates the displacement of biologically relevant ligands from trivalent ions of europium, Eu(III), and curium, Cm(III), in artificial biofluids by various complexing agents, i.e., ethylenediaminetetraacetic acid (EDTA), ethylene glycol-bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid (EGTA), diethylenetriaminepentaacetic acid (DTPA), and spermine-based hydroxypyridonate chelator 3,4,3-LI(1,2-HOPO) (HOPO). Utilizing a modified unified bioaccessibility method (UBM) to simulate gastrointestinal conditions, we conducted concentration-dependent displacement experiments at both room and body temperatures. Time-resolved laser-induced fluorescence spectroscopy (TRLFS) supported by ²H nuclear magnetic resonance (NMR) spectroscopy and thermodynamic modelling revealed the complexation efficacy of the agents under physiological conditions. Results demonstrate that high affinity, governed by complex stability constants and ligand pK_a values, is critical to overcome cation and anion competition and leads to effective decorporation. Additionally, there is evidence that cyclic ligands are inferior to linear ligands for this application. HOPO and DTPA exhibited superior displacement efficacy, particularly in the complete gastrointestinal tract simulation. This study highlights the utility of in vitro workflows for evaluating decorporation agents and emphasizes the need for ligands with optimal binding characteristics for enhanced chelation therapies. Full article

(This article belongs to the Special Issue Toxicity of Heavy Metal Compounds)

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9 pages, 413 KiB

Open AccessReview

Co-Cultivation Assays for Detecting Infectious Human-Tropic Porcine Endogenous Retroviruses (PERVs)

by Joachim Denner

Int. J. Mol. Sci. 2025, 26(15), 7111; https://doi.org/10.3390/ijms26157111 - 23 Jul 2025

Abstract

Porcine endogenous retroviruses (PERVs) are integrated into the genome of all pigs. As they can be released as infectious virus particles capable of infecting human cells in vitro, they pose a potential risk for xenotransplantation involving pig cells or organs. To assess whether [...] Read more.

Porcine endogenous retroviruses (PERVs) are integrated into the genome of all pigs. As they can be released as infectious virus particles capable of infecting human cells in vitro, they pose a potential risk for xenotransplantation involving pig cells or organs. To assess whether pigs produce infectious human-tropic viruses, infection assays with human cells are required. There are three main types of assays. First is the incubation of human target cells with gamma-irradiated pig cells. This method ensures that viral transmission is assessed in the absence of replicating pig cells. However, gamma irradiation may alter gene expression in pig cells, potentially affecting the results. Second is the co-culture in a double-chamber system in which pig and human cells are separated by a porous membrane, preventing direct cell-to-cell contact. While this method allows for the detection of infection by free virus particles, it does not account for infection via cell-to-cell transmission, which is a common mode of retroviral infection. And third is the co-culture of pig cells with human cells expressing a resistance gene. The resistance gene allows selective elimination of pig cells upon the addition of a selection medium. This assay enables both free virus and cell-to-cell transmission as well as complete removal of pig cells, which may not be fully achieved in the first type of assay. The third assay best simulates the conditions of in vivo xenotransplantation. However, in all cases the selection of donor and recipient cells is crucial to the experimental outcome. Results only indicate whether a specific pig cell type releases PERVs and whether a specific human cell type is susceptible to infection. A negative infection result does not necessarily reflect the in vivo situation, in which a transplanted organ consists of multiple pig cell types interacting with a diverse range of human cells within a living organism. Knowledge of these limitations is important for authorities regulating clinical applications for xenotransplantation. Full article

(This article belongs to the Special Issue Microbial Infections and Novel Biological Molecules for Treatment)

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17 pages, 3752 KiB

Open AccessArticle

PriorCCI: Interpretable Deep Learning Framework for Identifying Key Ligand–Receptor Interactions Between Specific Cell Types from Single-Cell Transcriptomes

by Hanbyeol Kim, Eunyoung Choi, Yujeong Shim and Joonha Kwon

Int. J. Mol. Sci. 2025, 26(15), 7110; https://doi.org/10.3390/ijms26157110 - 23 Jul 2025

Abstract

Understanding the interactions between specific cell types within tissue environments is essential for elucidating key biological processes, such as immune responses, cancer progression, inflammation, and development, in both physiological and pathological studies. The predominant methods for analyzing cell–cell interactions (CCI) rely primarily on [...] Read more.

Understanding the interactions between specific cell types within tissue environments is essential for elucidating key biological processes, such as immune responses, cancer progression, inflammation, and development, in both physiological and pathological studies. The predominant methods for analyzing cell–cell interactions (CCI) rely primarily on statistical inference using mapping or network-based techniques. However, these approaches often struggle to prioritize meaningful interactions owing to the high sparsity and heterogeneity inherent in single-cell RNA sequencing (scRNA-seq) data, where small but biologically important differences can be easily overlooked. To overcome these limitations, we developed PriorCCI, a deep-learning framework that leverages a convolutional neural network (CNN) alongside Grad-CAM++, an explainable artificial intelligence algorithm. This study aims to provide a scalable, interpretable, and biologically meaningful framework for systematically identifying and prioritizing key ligand–receptor interactions between defined cell-type pairs from single-cell RNA-seq data, particularly in complex environments such as tumors. PriorCCI effectively prioritizes interactions between cancer and other cell types within the tumor microenvironment and accurately identifies biologically significant interactions related to angiogenesis. By providing a visual interpretation of gene-pair contributions, our approach enables robust inference of gene–gene interactions across distinct cell types from scRNA-seq data. Full article

(This article belongs to the Special Issue New Insights in Translational Bioinformatics: Second Edition)

20 pages, 376 KiB

Open AccessArticle

Exploring the Relationship Between Brain-Derived Neurotrophic Factor Haplotype Variants, Personality, and Nicotine Usage in Women

by Dominika Borowy, Agnieszka Boroń, Jolanta Chmielowiec, Krzysztof Chmielowiec, Milena Lachowicz, Jolanta Masiak, Anna Grzywacz and Aleksandra Suchanecka

Int. J. Mol. Sci. 2025, 26(15), 7109; https://doi.org/10.3390/ijms26157109 - 23 Jul 2025

Abstract

Brain-derived neurotrophic factor (BDNF) is associated with nicotine use behaviours, the intensity of nicotine cravings, and the experience of withdrawal symptoms. Given the established influence of sex, brain-derived neurotrophic factor variants, personality traits and anxiety levels on nicotine use, this study aimed to [...] Read more.

Brain-derived neurotrophic factor (BDNF) is associated with nicotine use behaviours, the intensity of nicotine cravings, and the experience of withdrawal symptoms. Given the established influence of sex, brain-derived neurotrophic factor variants, personality traits and anxiety levels on nicotine use, this study aimed to conduct a comprehensive association analysis of these factors within a cohort of women who use nicotine. The study included 239 female participants: 112 cigarette users (mean age = 29.19, SD = 13.18) and 127 never-smokers (mean age = 28.1, SD =10.65). Study participants were examined using the NEO Five-Factor Inventory and the State–Trait Anxiety Inventory. Genotyping of rs6265, rs10767664, and rs2030323 was performed by real-time PCR using an oligonucleotide assay. We did not observe significant differences in the distribution of either genotype or allele of rs6265, rs10767664 and rs2030323 between groups. However, compared to the never-smokers, cigarette users scored significantly lower on the Agreeableness (5.446 vs. 6.315; p = 0.005767; d_Cohen’s = 0.363; η² = 0.032) and the Conscientiousness (5.571 vs. 6.882; p = 0.000012; d_Cohen’s = 0.591; η²= 0.08) scales. There was significant linkage disequilibrium between all three analysed polymorphic variants—between rs6265 and rs10767664 (D′ = 0.9994962; p < 2.2204 × 10⁻¹⁶), between rs6265 and rs2030323 (D′ = 0.9994935; p < 2.2204 × 10⁻¹⁶) and between rs10767664 and rs20330323 (D′ = 0.9838157; p < 2.2204 × 10⁻¹⁶), but the haplotype association analysis revealed no significant differences. While our study did not reveal an association between the investigated brain-derived neurotrophic factor polymorphisms (rs6265, rs10767664 and rs2030323) and nicotine use, it is essential to acknowledge that nicotine dependence is a complex, multifactorial phenotype. Our study expands the current knowledge of BDNF ’s potential role in addictive behaviours by exploring the understudied variants (rs10767664 and rs2030323), offering a novel contribution to the field and paving the way for future research into their functional relevance in addiction-related phenotypes. The lower Agreeableness and Conscientiousness scores observed in women who use nicotine compared to never-smokers suggest that personality traits play a significant role in nicotine use in women. The observed relationship between personality traits and nicotine use lends support to the self-medication hypothesis, suggesting that some women may initiate or maintain nicotine use as a coping mechanism for stress and negative affect. Public health initiatives targeting women should consider personality and psychological risk factors in addition to biological risks. Full article

(This article belongs to the Special Issue Molecular Insights into Addiction)

22 pages, 3771 KiB

Open AccessArticle

Integrated Transcriptome and Metabolome Analyses Uncover Cholesterol-Responsive Gene Networks

by Ruihao Zhang, Qi Sun, Lixia Huang and Jian Li

Int. J. Mol. Sci. 2025, 26(15), 7108; https://doi.org/10.3390/ijms26157108 - 23 Jul 2025

Abstract

Cholesterol stress profoundly modulates cellular processes, but its underlying mechanisms remain incompletely understood. To investigate cholesterol-responsive networks, we performed integrated transcriptome (RNA-seq) and metabolome (LC-MS) analyses on HeLa cells treated with cholesterol for 6 and 24 h. Through transcriptomic analysis of cholesterol-stressed HeLa [...] Read more.

Cholesterol stress profoundly modulates cellular processes, but its underlying mechanisms remain incompletely understood. To investigate cholesterol-responsive networks, we performed integrated transcriptome (RNA-seq) and metabolome (LC-MS) analyses on HeLa cells treated with cholesterol for 6 and 24 h. Through transcriptomic analysis of cholesterol-stressed HeLa cells, we identified stage-specific responses characterized by early-phase stress responses and late-phase immune-metabolic coordination. This revealed 1340 upregulated and 976 downregulated genes after a 6 h cholesterol treatment, including induction and suppression of genes involved in cholesterol efflux and sterol biosynthesis, respectively, transitioning to Nuclear Factor kappa-B (NF-κB) activation and Peroxisome Proliferator-Activated Receptor (PPAR) pathway modulation by 24 h. Co-expression network analysis prioritized functional modules intersecting with differentially expressed genes. We also performed untargeted metabolomics using cells treated with cholesterol for 6 h, which demonstrated extensive remodeling of lipid species. Interestingly, integrated transcriptomic and metabolic analysis uncovered GFPT1-driven Uridine Diphosphate-N-Acetylglucosamine (UDP-GlcNAc) accumulation and increased taurine levels. Validation experiments confirmed GFPT1 upregulation and ANGPTL4 downregulation through RT-qPCR and increased O-GlcNAcylation via Western blot. Importantly, clinical datasets further supported the correlations between GFPT1/ANGPTL4 expression and cholesterol levels in Non-Alcoholic Steatohepatitis (NASH) liver cancer patients. This work establishes a chronological paradigm of cholesterol sensing and identifies GFPT1 and ANGPTL4 as key regulators bridging glycosylation and lipid pathways, providing mechanistic insights into cholesterol-associated metabolic disorders. Full article

(This article belongs to the Section Molecular Genetics and Genomics)

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14 pages, 1428 KiB

Open AccessArticle

Extraction of Chitin, Chitosan, and Calcium Acetate from Mussel Shells for Sustainable Waste Management

by Chaowared Seangarun, Somkiat Seesanong, Banjong Boonchom, Nongnuch Laohavisuti, Pesak Rungrojchaipon, Wimonmat Boonmee, Sirichet Punthipayanon and Montree Thongkam

Int. J. Mol. Sci. 2025, 26(15), 7107; https://doi.org/10.3390/ijms26157107 - 23 Jul 2025

Abstract

In this paper, mussel shells were used to produce chitin, chitosan, and calcium acetate using chemical processes, searching for an alternative environmentally friendly biopolymer and calcium source. Mussel shells were treated with acetic acid as a demineralizing agent, resulting in separate solid fractions [...] Read more.

In this paper, mussel shells were used to produce chitin, chitosan, and calcium acetate using chemical processes, searching for an alternative environmentally friendly biopolymer and calcium source. Mussel shells were treated with acetic acid as a demineralizing agent, resulting in separate solid fractions and calcium solution. The solid was further purified to produce chitin by deproteinization and decolorization processes, and then the deacetylation process was used to obtain chitosan. The calcium solution was evaporated to produce calcium acetate powder. The yields of extracted chitin, chitosan, and calcium acetate from 100 g of mussel shells were 2.98, 2.70, and 165.23 g, respectively. The prepared chitin, chitosan, and calcium acetate were analyzed by Fourier transform infrared (FTIR) spectrophotometry, X-ray diffraction (XRD), thermogravimetric analysis (TGA), and scanning electron microscope (SEM) to confirm the chemical and physical properties. The analysis results of chitin and chitosan revealed the similarity to chitosan derived from crustaceans and insects in terms of functional group, structure and morphologies. The prepared calcium acetate shows FTIR and XRD data corresponding to calcium acetate monohydrate (Ca(CH₃COO)₂·H₂O) similar to synthesized calcium acetate in previous research. In addition, the mineral contents of calcium acetate identified by X-ray fluorescence (XRF) analysis exhibit 97.8% CaO with non-toxic impurities. This work demonstrated the potential of the production process of chitin, chitosan, and calcium acetate for the development of a sustainable industrial process with competitive functional performance against the commercial chitin and chitosan production process using crustacean shells and supported the implementation of a circular economy. Full article

(This article belongs to the Section Materials Science)

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19 pages, 3656 KiB

Open AccessArticle

Large-Scale Profiling of Coding and Long Noncoding Transcriptomes in the Hippocampus of Mice Acutely Exposed to Vaporized CBD or THC

by Mi Ran Choi, Jihun Kim, Chaeeun Park, Seok Hwan Chang, Han-Na Kim, Yeung Bae Jin and Sang-Rae Lee

Int. J. Mol. Sci. 2025, 26(15), 7106; https://doi.org/10.3390/ijms26157106 - 23 Jul 2025

Abstract

Cannabis vaping, particularly involving cannabidiol (CBD) and delta-9-tetrahydrocannabinol (THC), rapidly delivers highly concentrated cannabinoids to the brain, potentially affecting the hippocampus. This study examined differential expression of long noncoding RNAs (lncRNAs) and mRNAs in the hippocampus after acute exposure to vaporized CBD or [...] Read more.

Cannabis vaping, particularly involving cannabidiol (CBD) and delta-9-tetrahydrocannabinol (THC), rapidly delivers highly concentrated cannabinoids to the brain, potentially affecting the hippocampus. This study examined differential expression of long noncoding RNAs (lncRNAs) and mRNAs in the hippocampus after acute exposure to vaporized CBD or THC. Male ICR mice were exposed to vaporized CBD or THC (50 mg, n = 5/group), and hippocampal tissues were collected at 1, 3, and 14 days post-exposure. Total RNA sequencing was conducted on day 1 samples, and selected transcripts were validated using qRT-PCR across multiple time points. CBD led to significant up- or downregulation of L3mbtl1, Wnt7a, and Camk2b at day 1. However, Wnt7a showed gradual recovery at days 3 and 14. In the THC group, Grin2a, Gria3, and Golga2 were significantly upregulated, while Drd1, Drd2, Gnal, and Adcy5 were significantly downregulated at day 1. Time-course analysis showed that Drd2 expression returned to baseline by day 14, whereas Adcy5 remained persistently downregulated through days 3 and 14. In the CBD group, NONMMUT069014.2 was upregulated, while NONMMUT033147.2 and NONMMUT072606.2 were downregulated at day 1; notably, NONMMUT072606.2 showed a transient increase at day 3 before returning to baseline. In the THC group, NONMMUT085523.1 and NONMMUT123548.1 were upregulated, whereas NONMMUT019734.2, NONMMUT057101.2, and NONMMUT004928.2 were downregulated, with most showing gradual recovery by day 14. Correlation analysis revealed that THC-responsive lncRNAs—including NONMMUT004928.2, NONMMUT057101.2, and NONMMUT019734.2—were strongly associated with downregulated mRNAs such as Drd2 and Adcy5. These findings highlight cannabinoid-specific hippocampal transcriptomic responses and suggest potential regulatory roles for lncRNA–mRNA interactions in cannabinoid-induced neural changes. Full article

(This article belongs to the Special Issue MicroRNA and Non-Coding RNA: From Basic Research to Potential Clinical Application)

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10 pages, 780 KiB

Open AccessArticle

Facile Synthesis of Polysubstituted Pyridines via Metal-Free [3+3] Annulation Between Enamines and β,β-Dichloromethyl Peroxides

by Yangyang Ma, Hua Zhang, Zhonghao Zhou, Chenyang Yang, Wenxiao Chang, Mohan Li, Yapei Zheng, Weizhuang Zhang, Huan Yue, Changdong Chen, Ming La and Yongjun Han

Int. J. Mol. Sci. 2025, 26(15), 7105; https://doi.org/10.3390/ijms26157105 - 23 Jul 2025

Abstract

Our work introduces a facile and efficient metal-free [3+3] annulation approach for the synthesis of polysubstituted pyridines via the reaction between β-enaminonitriles and β,β-dichloromethyl peroxides. This strategy operates under mild conditions, demonstrating broad substrate scope and excellent functional group tolerance. Mechanistic investigations suggest [...] Read more.

Our work introduces a facile and efficient metal-free [3+3] annulation approach for the synthesis of polysubstituted pyridines via the reaction between β-enaminonitriles and β,β-dichloromethyl peroxides. This strategy operates under mild conditions, demonstrating broad substrate scope and excellent functional group tolerance. Mechanistic investigations suggest that the reaction proceeds through a Kornblum–De La Mare rearrangement followed by S_NV-type C-Cl bond cleavage and intramolecular cyclization/condensation. By circumventing the need for transition metal catalysts or radical initiators, our method offers practical utility in organic synthesis and provides a new avenue for the rapid construction of complex pyridine scaffolds. Full article

(This article belongs to the Section Physical Chemistry and Chemical Physics)

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29 pages, 2831 KiB

Open AccessReview

The 3D Language of Cancer: Communication via Extracellular Vesicles from Tumor Spheroids and Organoids

by Simona Campora and Alessandra Lo Cicero

Int. J. Mol. Sci. 2025, 26(15), 7104; https://doi.org/10.3390/ijms26157104 - 23 Jul 2025

Abstract

Extracellular vesicles (EVs) have emerged as key mediators of intercellular communication, gaining recognition as tumor biomarkers and promising therapeutic targets. As the study of EVs advances, it has become increasingly clear that the cellular context in which they are produced significantly influences their [...] Read more.

Extracellular vesicles (EVs) have emerged as key mediators of intercellular communication, gaining recognition as tumor biomarkers and promising therapeutic targets. As the study of EVs advances, it has become increasingly clear that the cellular context in which they are produced significantly influences their composition and function. Traditional two-dimensional in vitro models are being progressively replaced by more advanced three-dimensional systems, such as tumor spheroids and organoids. These 3D models are particularly valuable in cancer research, providing a more accurate representation of the complex cellular and molecular heterogeneity that characterizes tumors, better mimicking the in vivo microenvironment compared to standard monolayer cultures. This review explores the role of EVs derived from tumor spheroids and organoids in key oncogenic processes, including tumor growth, metastasis, and interactions within the tumor microenvironment. We highlight how EVs contribute to the spread of cancer cells, affecting surrounding tissues, and promote immune evasion, which poses significant challenges in cancer therapy. Full article

(This article belongs to the Special Issue Recent Advances in 3D Tumor Models for Cancer Research)

25 pages, 2221 KiB

Open AccessArticle

by Karolina Małas, Ludmiła Polechońska and Katarzyna Kabała

Int. J. Mol. Sci. 2025, 26(15), 7103; https://doi.org/10.3390/ijms26157103 - 23 Jul 2025

Abstract

Chloroplasts, as the organelles primarily responsible for photosynthesis, require a substantial supply of iron ions. Conversely, due to Fe toxicity, the homeostasis of these ions is subject to tight regulation. Permease in chloroplast 1 (PIC1) has been identified as the primary iron importer [...] Read more.

Chloroplasts, as the organelles primarily responsible for photosynthesis, require a substantial supply of iron ions. Conversely, due to Fe toxicity, the homeostasis of these ions is subject to tight regulation. Permease in chloroplast 1 (PIC1) has been identified as the primary iron importer into chloroplasts. However, previous studies suggested the existence of a distinct pathway for Fe transfer to chloroplasts, likely involving mitoferrin-like 1 (MFL1) protein. In this work, Arabidopsis MFL1 (AtMFL1) and its cucumber homolog (CsMFL1) were characterized using, among others, Arabidopsis protoplasts as well as both yeast and Arabidopsis mutants. Localization of both proteins in chloroplasts has been shown to be mediated via an N-terminal transit peptide. At the gene level, MFL1 expression profiles differed between the model plant and the crop plant under varying Fe availability. The expression of other genes involved in chloroplast Fe homeostasis, including iron acquisition, trafficking, and storage, was affected to some extent in both AtMFL1 knockout and overexpressing plants. Moreover, root growth and photosynthetic parameters changed unfavorably in the mutant lines. The obtained results imply that AtMFL1 and CsMFL1, as putative chloroplast iron transporters, play a role in both iron management and the proper functioning of the plant. Full article

(This article belongs to the Special Issue New Insights in Plant Cell Biology)

21 pages, 1985 KiB

Open AccessReview

Neuroplasticity-Based Approaches to Sensory Processing Alterations in Autism Spectrum Disorder

by Maria Suprunowicz, Julia Bogucka, Natalia Szczerbińska, Stefan Modzelewski, Aleksandra Julia Oracz, Beata Konarzewska and Napoleon Waszkiewicz

Int. J. Mol. Sci. 2025, 26(15), 7102; https://doi.org/10.3390/ijms26157102 - 23 Jul 2025

Abstract

Sensory dysregulation represents a core challenge in autism spectrum disorder (ASD), affecting perception, behavior, and adaptive functioning. The brain’s ability to reorganize, known as neuroplasticity, serves as the basic principle for therapeutic interventions targeting these deficits. Neuroanatomical mechanisms include altered connectivity in the [...] Read more.

Sensory dysregulation represents a core challenge in autism spectrum disorder (ASD), affecting perception, behavior, and adaptive functioning. The brain’s ability to reorganize, known as neuroplasticity, serves as the basic principle for therapeutic interventions targeting these deficits. Neuroanatomical mechanisms include altered connectivity in the sensory and visual cortices, as well as in the limbic system and amygdala, while imbalances of neurotransmitters, in particular glutamate and gamma-aminobutyric acid (GABA), contribute to atypical sensory processing. Traditional therapies used in sensory integration are based on the principles of neuroplasticity. Increasingly, new treatments use this knowledge, and modern therapies such as neurofeedback, transcranial stimulation, and immersive virtual environments are promising in modulating neuronal circuits. However, further research is needed to optimize interventions and confirm long-term effectiveness. This review discusses the role of neuroplasticity in the etiopathogenesis of sensory integration deficits in autism spectrum disorder. The neuroanatomical and neurotransmitter basis of impaired perception of sensory stimuli is considered, and traditional and recent therapies for sensory integration are discussed. Full article

(This article belongs to the Special Issue Molecular Investigations in Neurodevelopmental Disorders)

47 pages, 4589 KiB

Open AccessReview

Understanding Sex Differences in Autoimmune Diseases: Immunologic Mechanisms

by Yu Rin Kim, YunJae Jung, Insug Kang and Eui-Ju Yeo

Int. J. Mol. Sci. 2025, 26(15), 7101; https://doi.org/10.3390/ijms26157101 - 23 Jul 2025

Abstract

Autoimmune diseases such as systemic lupus erythematosus and Sjögren’s syndrome show pronounced sex disparities in prevalence, severity, and clinical outcomes, with females disproportionately affected. Emerging evidence highlights sex-based differences in immune and inflammatory responses as key contributors to this bias. Genetic factors—including sex [...] Read more.

Autoimmune diseases such as systemic lupus erythematosus and Sjögren’s syndrome show pronounced sex disparities in prevalence, severity, and clinical outcomes, with females disproportionately affected. Emerging evidence highlights sex-based differences in immune and inflammatory responses as key contributors to this bias. Genetic factors—including sex chromosomes, skewed X chromosome inactivation, and sex-biased microRNAs—as well as sex hormones and pregnancy modulate gene expression and immune cell function in a sex-specific manner. Additionally, sex hormone-dependent epigenetic modifications influence the transcription of critical immune regulators. These genetic and hormonal factors collectively shape the activation, differentiation, and effector functions of diverse immune cell types. Environmental factors—including infections, gut microbiota, environmental chemicals and pollutants, and lifestyle behaviors such as diet, smoking, UV exposure, alcohol and caffeine intake, physical activity, and circadian rhythms—further modulate immune function and autoimmune disease pathogenesis in a sex-dependent manner. Together, these mechanisms contribute to the heightened risk and distinct clinical features of autoimmunity in females. A deeper understanding of sex-biased immune regulation will facilitate the identification of novel biomarkers, enable patient stratification, and inform the development of sex-specific diagnostic and therapeutic strategies for autoimmune diseases. Full article

(This article belongs to the Section Molecular Immunology)

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22 pages, 1630 KiB

Open AccessArticle

Development of Cytisus Flower Extracts with Antioxidant and Anti-Inflammatory Properties for Nutraceutical and Food Uses

by Adela Alvaredo-López-Vizcaíno, Augusto Costa-Barbosa, Paula Sampaio, Pablo G. del Río, Claudia Botelho and Pedro Ferreira-Santos

Int. J. Mol. Sci. 2025, 26(15), 7100; https://doi.org/10.3390/ijms26157100 - 23 Jul 2025

Abstract

Plant flowers are recognized as a rich source of bioactive phenolic compounds. In this study, for the first time, the recovery of antioxidant phenolic compounds from Cytisus striatus flowers (CF) was optimized using microwave-assisted extraction (MAE). The variables (% of ethanol, temperature, and [...] Read more.

Plant flowers are recognized as a rich source of bioactive phenolic compounds. In this study, for the first time, the recovery of antioxidant phenolic compounds from Cytisus striatus flowers (CF) was optimized using microwave-assisted extraction (MAE). The variables (% of ethanol, temperature, and time) were studied using a response surface methodology (RSM). Extraction efficiency was assessed by total phenol content, total flavonoid content, and the antioxidant capacity through DPPH, ABTS, FRAP, and CUPRAC assays. Additionally, cytotoxicity and anti-inflammatory properties were evaluated in different cell lines. The optimal extraction conditions (87.6% ethanol, 160.8 °C and 8.76 min) yielded extracts rich in phenolics (85.9 mg GAE/g CF) and flavonoids (120.3 mg RE/g CF), with strong antioxidant capacity. LC-MS/MS analysis identified 27 phenolic compounds, including chrysin, apigenin, and quercetin derivatives. Cytotoxicity tests showed that CF extract maintained high viability (>80%) in human embryonic kidney (HEK293T) and human lung adenocarcinoma (A549) cells up to 2000 µg/mL, indicating low cytotoxicity. The anti-inflammatory potential was evidenced by a decrease in IL-1β levels and an increase in IL-10 cytokine production in LPS-stimulated macrophages. These results highlight the great potential of CF as a promising bioresource to obtain value-added compounds for the development of functional foods, nutraceuticals, and cosmetic products. Full article

(This article belongs to the Special Issue Isolation and Biological Activities of Natural Products: Focus on Molecular Advance)

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20 pages, 2629 KiB

Open AccessArticle

Neural Progenitor Cell- and Developing Neuron-Derived Extracellular Vesicles Differentially Modulate Microglial Activation

by Tsung-Lang Chiu, Hsin-Yi Huang, Hock-Kean Liew, Hui-Fen Chang, Hsin-Rong Wu and Mei-Jen Wang

Int. J. Mol. Sci. 2025, 26(15), 7099; https://doi.org/10.3390/ijms26157099 - 23 Jul 2025

Abstract

The developmental processes of microglia follow a general pattern, from immature amoeboid (activated) cells to fully ramified (inactivated) surveilling microglia. However, little is known about the mechanisms controlling the transition of microglia from an activated to an inactivated state during brain development. Due [...] Read more.

The developmental processes of microglia follow a general pattern, from immature amoeboid (activated) cells to fully ramified (inactivated) surveilling microglia. However, little is known about the mechanisms controlling the transition of microglia from an activated to an inactivated state during brain development. Due to the complexity of microenvironmentally dynamic changes during neuronal differentiation, interactions between developing nerve cells and microglia might be involved in this process. Extracellular vesicles (EVs) are cell-released particles that serve as mediators of cellular crosstalk and regulation. Using neural progenitor cells (NPCs) and a long-term neuron culture system, we found that EVs derived from NPCs or developing neurons possessed differential capacity on the induction of microglial activation. The exposure of microglia to NPC- or immature neuron (DIV7)-derived EVs resulted in the higher expression of protein and mRNA of multiple inflammatory cytokines (e.g., TNF-α, IL-1β, and IL-6), when compared with mature neuron-derived EVs. Exploration of the intracellular signaling pathways revealed that MAPK signaling, IκBα phosphorylation/degradation, and NF-κB p65 nuclear translocation were strongly induced in microglia treated with NPC- or immature neuron-derived EVs. Using a pharmacological approach, we further demonstrate that Toll-like receptor (TLR) 7-mediated activation of NF-κB and MAPK signaling cascades contribute to EV-elicited microglial activation. Additionally, the application of conditioned media derived from microglia treated with NPC- or immature neuron-derived EVs is found to promote the survival of late-developing dopaminergic neurons. Thus, our results highlight a novel mechanism used by NPCs and developing neurons to modulate the developmental phases and functions of microglia through EV secretion. Full article

(This article belongs to the Section Molecular Biology)

37 pages, 4312 KiB

Open AccessReview

Neutrophils and NETs in Pathophysiology and Treatment of Inflammatory Bowel Disease

by Marina Ortega-Zapero, Raquel Gomez-Bris, Ines Pascual-Laguna, Angela Saez and Jose M. Gonzalez-Granado

Int. J. Mol. Sci. 2025, 26(15), 7098; https://doi.org/10.3390/ijms26157098 - 23 Jul 2025

Abstract

Inflammatory Bowel Disease (IBD), which includes ulcerative colitis (UC) and Crohn’s disease (CD), results from dysregulated immune responses that drive chronic intestinal inflammation. Neutrophils, as key effectors of the innate immune system, contribute to IBD through multiple mechanisms, including the release of reactive [...] Read more.

Inflammatory Bowel Disease (IBD), which includes ulcerative colitis (UC) and Crohn’s disease (CD), results from dysregulated immune responses that drive chronic intestinal inflammation. Neutrophils, as key effectors of the innate immune system, contribute to IBD through multiple mechanisms, including the release of reactive oxygen species (ROS), pro-inflammatory cytokines, and neutrophil extracellular traps (NETs). NETs are web-like structures composed of DNA, histones, and associated proteins including proteolytic enzymes and antimicrobial peptides. NET formation is increased in IBD and has a context-dependent role; under controlled conditions, NETs support antimicrobial defense and tissue repair, whereas excessive or dysregulated NETosis contributes to epithelial injury, barrier disruption, microbial imbalance, and thrombotic risk. This review examines the roles of neutrophils and NETs in IBD. We summarize recent single-cell and spatial-omics studies that reveal extensive neutrophil heterogeneity in the inflamed gut. We then address the dual role of neutrophils in promoting tissue damage—through cytokine release, immune cell recruitment, ROS production, and NET formation—and in supporting microbial clearance and mucosal healing. We also analyze the molecular mechanisms regulating NETosis, as well as the pathways involved in NET degradation and clearance. Focus is given to the ways in which NETs disrupt the epithelial barrier, remodel the extracellular matrix, contribute to thrombosis, and influence the gut microbiota. Finally, we discuss emerging therapeutic strategies aimed at restoring NET homeostasis—such as PAD4 inhibitors, NADPH oxidase and ROS pathway modulators, and DNase I—while emphasizing the need to preserve antimicrobial host defenses. Understanding neutrophil heterogeneity and NET-related functions may facilitate the development of new therapies and biomarkers for IBD, requiring improved detection tools and integrated multi-omics and clinical data. Full article

(This article belongs to the Special Issue Molecular Mechanisms and Therapeutic Targeting of Inflammatory Complications)

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18 pages, 929 KiB

Open AccessArticle

Associations of Serum GIP, GLP-1, and DPP-4 with Metabolic and Hormonal Profiles and Tobacco Exposure in Women with Polycystic Ovary Syndrome

by Anna Bizoń, Julia Borkowska, Grzegorz Franik and Agnieszka Piwowar

Int. J. Mol. Sci. 2025, 26(15), 7097; https://doi.org/10.3390/ijms26157097 - 23 Jul 2025

Abstract

Disorders in glucose metabolism are well-established features of polycystic ovary syndrome (PCOS) and are linked to its clinical severity and phenotypic variability. This study aimed to assess serum concentrations of glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), and dipeptidyl peptidase-4 (DPP-4) and to [...] Read more.

Disorders in glucose metabolism are well-established features of polycystic ovary syndrome (PCOS) and are linked to its clinical severity and phenotypic variability. This study aimed to assess serum concentrations of glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), and dipeptidyl peptidase-4 (DPP-4) and to examine their relationships with glucose and insulin levels, selected sex hormone concentrations, body weight, and exposure to tobacco smoke. Women with PCOS exhibited significantly elevated levels of fasting glucose, insulin, GIP, and GLP-1 compared to controls. Tobacco smoke exposure in women with PCOS was associated with reduced DPP-4 levels, which were approximately two-fold lower in smokers than in non-smokers. A significant negative correlation between DPP-4 and cotinine levels further supported this relationship. Comorbidities such as overweight/obesity or insulin resistance (IR) were also linked to elevated incretin hormone levels. However, no significant age-related trends in incretin levels were identified, despite the known association between age and glucose dysregulation. The notable alterations in incretin hormone profiles in PCOS, along with the consistent patterns of GIP or GLP-1 with metabolic and hormonal parameters, suggest that these hormones may play coordinated regulatory roles in the pathophysiology of PCOS. Full article

(This article belongs to the Special Issue Focus on Metabolic Research Priorities in PCOS)

15 pages, 679 KiB

Open AccessReview

The Influence of Exercise and Physical Activity on Autonomic Nervous System Function Measured by Heart Rate Variability in Individuals with Type 1 Diabetes Mellitus—A Systematic Review

by Isabel Bekker, Arne Kooistra, Peter R. van Dijk, Joop D. Lefrandt, Nic J. G. M. Veeger and André P. van Beek

Int. J. Mol. Sci. 2025, 26(15), 7096; https://doi.org/10.3390/ijms26157096 - 23 Jul 2025

Abstract

Non-pharmacological interventions, such as physical activity and exercise, are essential in managing type 1 diabetes mellitus by improving glycemic control, cardiovascular health and autonomic function. Given the chronic nature and long-term complications associated with type 1 diabetes, strategies beyond pharmacotherapy are essential. This [...] Read more.

Non-pharmacological interventions, such as physical activity and exercise, are essential in managing type 1 diabetes mellitus by improving glycemic control, cardiovascular health and autonomic function. Given the chronic nature and long-term complications associated with type 1 diabetes, strategies beyond pharmacotherapy are essential. This review examines the effects of exercise on heart rate variability, a key indicator of autonomic nervous system activity. A systematic search was conducted in March 2024 across PubMed, Embase, Cochrane and CINAHL databases. Studies evaluating the retrospective or prospective impact of exercise or physical activity on heart rate variability parameters were included. Utilizing best evidence synthesis, the methodological quality of the included studies was evaluated. Seven studies met the inclusion criteria, all of which were rated as methodologically weak. Moderate evidence suggests that exercise may enhance heart rate variability, particularly by increasing parasympathetic activity and improving sympathovagal balance. However, evidence remains limited regarding the optimal type, frequency and intensity of exercise. Exercise appears to support autonomic function in individuals with type 1 diabetes mellitus. Nonetheless, further high-quality research is needed to determine the most effective exercise modalities and to inform evidence-based clinical guidelines. Full article

(This article belongs to the Special Issue Latest Advances in Diabetes Research and Practice)

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33 pages, 1463 KiB

Open AccessReview

Molecular Mechanisms of the Endocannabinoid System with a Focus on Reproductive Physiology and the Cannabinoid Impact on Fertility

by Patrycja Kalak, Piotr Kupczyk, Antoni Szumny, Tomasz Gębarowski, Marcin Jasiak, Artur Niedźwiedź, Wojciech Niżański and Michał Dzięcioł

Int. J. Mol. Sci. 2025, 26(15), 7095; https://doi.org/10.3390/ijms26157095 - 23 Jul 2025

Abstract

The endocannabinoid system (ECS) is a complex neuromodulatory network involved in maintaining physiological balance through interactions with various neurotransmitter and hormonal pathways. Its key components—cannabinoid receptors (CBRs)—are activated by endogenous ligands and exogenous cannabinoids such as those found in the Cannabis sativa plant. [...] Read more.

The endocannabinoid system (ECS) is a complex neuromodulatory network involved in maintaining physiological balance through interactions with various neurotransmitter and hormonal pathways. Its key components—cannabinoid receptors (CBRs)—are activated by endogenous ligands and exogenous cannabinoids such as those found in the Cannabis sativa plant. Although cannabinoids like cannabidiol (CBD) have garnered interest for their potential therapeutic effects, evidence regarding their safety, particularly for reproductive health, remains limited. This review summarizes the structure and molecular mechanisms of the ECS, its role in reproductive physiology—including its interactions with the hypothalamic–pituitary–gonadal axis (HPG axis), gametogenesis, implantation, and lactation—and the possible consequences of cannabinoid exposure for fertility. In addition, we focus on the involvement of the ECS and cannabinoids in breast cancer, highlighting emerging evidence on their dual role in tumor progression and therapy. These insights emphasize the need for further research to better define the therapeutic potential and risks associated with cannabinoid use in reproductive health and breast cancer. Full article

(This article belongs to the Special Issue Exploring Cannabinoids: Molecular Mechanisms and Potential Therapeutic Applications)

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14 pages, 546 KiB

Open AccessReview

Belzutifan-Associated Hypoxia: A Review of the Novel Therapeutic, Proposed Mechanisms of Hypoxia, and Management Recommendations

by John Kucharczyk, Anshini Bhatt, Laura Bauer and Minas Economides

Int. J. Mol. Sci. 2025, 26(15), 7094; https://doi.org/10.3390/ijms26157094 - 23 Jul 2025

Abstract

Belzutifan is a hypoxia-inducible factor-2α (HIF-2α) inhibitor that received FDA approval in 2021 for treating cancers resulting from von Hippel-Lindau (VHL) disease, including clear cell renal cell carcinoma (ccRCC), followed by approval in 2023 for sporadic ccRCC that has progressed through multiple lines [...] Read more.

Belzutifan is a hypoxia-inducible factor-2α (HIF-2α) inhibitor that received FDA approval in 2021 for treating cancers resulting from von Hippel-Lindau (VHL) disease, including clear cell renal cell carcinoma (ccRCC), followed by approval in 2023 for sporadic ccRCC that has progressed through multiple lines of therapy. HIF-2α is a promising drug target, as VHL is commonly inactivated in ccRCC, which results in HIF-2α-mediated signaling that is considered central to tumorigenesis. Belzutifan has demonstrated efficacy in clinical trials in the first-line and subsequent line settings, and in combination with tyrosine kinase inhibitors. Despite being overall well tolerated, belzutifan has a distinct safety profile because of its unique mechanism of action. Anemia was the most common adverse event observed in clinical trials and is considered an on-target effect. Hypoxia is also frequently observed and commonly results in dose reductions, treatment discontinuation, and supplemental oxygen use. This review summarizes the rates of hypoxia seen in clinical trials of belzutifan in ccRCC. As the cause of hypoxia is not well understood, this review also discusses possible mechanisms of hypoxia based on preclinical studies of the HIF pathway and HIF-2α inhibitors. Finally, this review proposes monitoring and management recommendations for clinicians prescribing belzutifan to ccRCC patients. Full article

(This article belongs to the Special Issue Recent Advances in Urological Cancer)

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22 pages, 3350 KiB

Open AccessArticle

De Novo Hybrid Assembly of the Tripterygium wilfordii Mitochondrial Genome Provides the Chromosomal Mitochondrial DNA Structure and RNA Editing Events

by Yisha Cai, Suxin Yang, Haimei Chen, Yang Ni, Jingling Li, Jinghong Zhang and Chang Liu

Int. J. Mol. Sci. 2025, 26(15), 7093; https://doi.org/10.3390/ijms26157093 - 23 Jul 2025

Abstract

Tripterygium wilfordii has extremely important pharmaceutical value in both traditional and modern medicine. The mitogenome of T. wilfordii was subjected to assembly and annotation with Nanopore long reads and Illumina short reads in this study. The mitogenome is 720,306 bp in length and [...] Read more.

Tripterygium wilfordii has extremely important pharmaceutical value in both traditional and modern medicine. The mitogenome of T. wilfordii was subjected to assembly and annotation with Nanopore long reads and Illumina short reads in this study. The mitogenome is 720,306 bp in length and is responsible for encoding 55 specific genes, including 35 protein-coding genes (PCGs), 17 transfer RNA (tRNA) genes, and 3 ribosomal RNA (rRNA) genes. Upon repetitive sequence analysis, 223 simple sequence repeats (SSRs), 24 long tandem repeats (LTRs), and 47 dispersed repetitive sequences (DRSs) were identified. The 24 common PCGs were used for phylogenetic analysis, which revealed that T. wilfordii is more closely related to Euonymus alatus. Moreover, mitochondrial plastid DNA (MTPT) analysis revealed eight MTPTs in the mitochondrial genome. Furthermore, 600 RNA-editing sites were detected in the protein-coding genes according to RNA-seq results. Among these genes, the ccmB gene contained the greatest number of sites, followed by the nad4 gene. This is the first study to report the T. wilfordii mitogenome and illustrate its linear structure. The findings of this study will help elucidate the evolution of the T. wilfordii mitogenome and facilitate its potential application in genetic breeding. Full article

(This article belongs to the Collection Feature Papers in Molecular Informatics)

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