International Journal of Molecular Sciences

19 pages, 3033 KB

Open AccessArticle

Whole-Genome Sequence Analysis and Subtractive Screening of Lactobacilli in the Searching for New Probiotics to Protect the Mammary Glands

by Dobroslava Bujňáková, Tímea Galambošiová and Lívia Karahutová

Int. J. Mol. Sci. 2025, 26(21), 10809; https://doi.org/10.3390/ijms262110809 - 6 Nov 2025

Viewed by 677

Abstract

To discover new probiotics that can protect mammary glands from mastitis, 40 Lactobacillus (Ligilactobacillus) spp. isolates from bovine milk were subjected to a preliminary series of in vitro subtractive analyses. Antibiotic susceptibility testing was performed according to the ISO norm 10932. [...] Read more.

To discover new probiotics that can protect mammary glands from mastitis, 40 Lactobacillus (Ligilactobacillus) spp. isolates from bovine milk were subjected to a preliminary series of in vitro subtractive analyses. Antibiotic susceptibility testing was performed according to the ISO norm 10932. Many lactobacilli had elevated MIC values for kanamycin (35%), but fewer were resistant to chloramphenicol (15%), streptomycin (7.5%) and tetracycline (5%). The enzymic activities of lactobacilli were tested using an API ZYM system. Nearly 27% exhibited undesirable activities (β-glucuronidase, β-glucosidase and N-acetyl-β-glucosaminidase). The safe strains were monitored for antimicrobial activity against Staphylococcus aureus, Escherichia coli, Salmonella enteritidis, and Bacillus cereus using microtiter plates and for their ability to form biofilms using the crystal violet assay. The antimicrobial activity of lactobacilli against indicator bacteria ranged from 29 to 89% and the isolates exhibited moderate-to-high biofilm formation. Suitable strains were selected for whole-genome sequencing analysis. Antibiotic-resistance genes and putative virulence genes were not predicted in the genomic analysis. Moreover, the isolate Ligilactobacillus salivarius 48 carries genetic information responsible for bacteriocin production that is similar to that encoding salivaricin CRL1328. Our study demonstrates the safety of the above mentioned isolate, which has potential to be used as a probiotic, exerting health benefits through production of antimicrobial substances. Full article

(This article belongs to the Section Molecular Microbiology)

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18 pages, 2130 KB

Open AccessArticle

STAT3 Inhibition to Treat Ulcerative Colitis-Associated Colorectal Cancer

by Prema Robinson, Zal Italia, Zara Italia, Tan Hoang, Emma Rodriguez, T. Kris Eckols, Moses Kasembeli, Leticia Hamana Zorrilla, Luisa Maren Solis Soto, Rajasekaran Mahalingam and David J. Tweardy

Int. J. Mol. Sci. 2025, 26(21), 10808; https://doi.org/10.3390/ijms262110808 - 6 Nov 2025

Viewed by 719

Abstract

In patients with inflammatory bowel disease (IBD), colorectal cancer (CRC) occurs with 20-to-30-fold higher frequency, is more advanced at diagnosis, and has a worse prognosis than in the general population. To improve their treatment options, we determined if targeting STAT3 with TTI-101, a [...] Read more.

In patients with inflammatory bowel disease (IBD), colorectal cancer (CRC) occurs with 20-to-30-fold higher frequency, is more advanced at diagnosis, and has a worse prognosis than in the general population. To improve their treatment options, we determined if targeting STAT3 with TTI-101, a small-molecule STAT3 inhibitor, was beneficial in the azoxymethane (AOM)-disodium sulfate (DSS) mouse model of colitis-associated CRC. C57BL/6 mice received a single intraperitoneal injection of AOM followed by three cycles of 5% DSS in drinking water before receiving TTI-101 (50 mg/kg by oral gavage, OG, and daily) or vehicle for 28 days. TTI-101 treatment reduced adenoma numbers by 89% from 1.14 ± 1.07 in vehicle-treated mice to 0.13 ± 0.35 in TTI-101-treated mice (p ≤ 0.05, Kruskal–Wallis test). Levels of activated STAT3 (pY-STAT3) were increased 3.3-fold in the epithelium and stroma of dysplastic mucosa (147 ± 46; mean ± SD; and n = 4) vs. normal mucosa (45 ± 26; n = 7; and p ≤ 0.05, Kruskal–Wallis test) and were correlated with the adenoma number. TTI-101 was detected at pharmacologically relevant levels in the plasma and colons of TTI-101-treated AOM-DSS mice and was concentrated within colon tissue; plasma TTI-101 levels inversely correlated to pY-STAT3 levels. Importantly, TTI-101 normalized the colon transcriptome of AOM-DSS mice and reduced the expression of STAT3- and STAT1-upregulated genes associated with CRC oncogenesis. Thus, TTI-101 treatment may benefit IBD patients with CRC. Full article

(This article belongs to the Special Issue Current Research on Cancer Biology and Therapeutics: Fourth Edition)

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16 pages, 616 KB

Open AccessReview

Natural Sulfur Compounds in Mineral Waters: Implications for Human Health and Disease

by Mauro Vaccarezza, Marco Vitale, Paola Falletta and Orsola di Martino

Int. J. Mol. Sci. 2025, 26(21), 10807; https://doi.org/10.3390/ijms262110807 - 6 Nov 2025

Viewed by 900

Abstract

Natural sulfur compounds found in various mineral spring waters have attracted considerable interest due to their possible health benefits and healing qualities. Key substances such as hydrogen sulfide (H₂S), sulfate (SO₄²⁻), and thiosulfate (S₂O₃²⁻ [...] Read more.

Natural sulfur compounds found in various mineral spring waters have attracted considerable interest due to their possible health benefits and healing qualities. Key substances such as hydrogen sulfide (H₂S), sulfate (SO₄²⁻), and thiosulfate (S₂O₃²⁻) are essential to numerous physiological functions. This overview delves into the biochemical pathways through which these sulfur compounds exert their influence, emphasizing their roles as antioxidants, anti-inflammatories, and detoxifying agents. Furthermore, it investigates the therapeutic promise of mineral waters rich in sulfur for various diseases like arthritis, skin ailments, and heart diseases. Emerging studies indicate that regular use or topical application of these waters could enhance health outcomes and aid in the prevention of a multitude of diseases. Nonetheless, additional research is required to clarify sulfur water’s mechanisms of action and to develop standardized protocols for their therapeutic applications. This descriptive review highlights the significance of integrating natural sulfur compounds into comprehensive health strategies and advocates for ongoing investigation into their advantages in medical contexts. Full article

(This article belongs to the Special Issue The Role of Natural and Synthetic Sulfur Compounds in Human Health and Diseases)

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22 pages, 7929 KB

Open AccessArticle

Therapeutic Modulation of the Nox2–Hv1–ROS Axis by Botulinum Neurotoxin A Confers Protection Against CoCl₂-Induced Retinal Hypoxic Injury

by Hey Jin Lee, Mira Park, Hyun-Ah Shin and Helen Lew

Int. J. Mol. Sci. 2025, 26(21), 10806; https://doi.org/10.3390/ijms262110806 - 6 Nov 2025

Cited by 1 | Viewed by 531

Abstract

Neuroinflammation and oxidative stress are key drivers of various ocular diseases. Experimental hypoxia, modeled using cobalt chloride (CoCl₂), induces hypoxia-inducible factor 1-alpha (HIF-1α) stabilization, mitochondrial dysfunction, and excessive reactive oxygen species (ROS) production, primarily via the NADPH oxidase 2 (Nox2)–voltage-gated proton [...] Read more.

Neuroinflammation and oxidative stress are key drivers of various ocular diseases. Experimental hypoxia, modeled using cobalt chloride (CoCl₂), induces hypoxia-inducible factor 1-alpha (HIF-1α) stabilization, mitochondrial dysfunction, and excessive reactive oxygen species (ROS) production, primarily via the NADPH oxidase 2 (Nox2)–voltage-gated proton channel Hv1 axis. Although Botulinum neurotoxin type A (BoNT/A) is classically recognized for SNAP-25 cleavage, recent studies suggest broader anti-inflammatory and neuroprotective effects. We evaluated BoNT/A in R28 retinal precursor cells and ex vivo retinal explants subjected to CoCl₂-induced hypoxic stress. BoNT/A pretreatment attenuated CoCl₂-induced upregulation of HIF-1α, Hv1, Nox2, NOD-like receptor protein 3 (NLRP3), COX2, and nuclear factor kappa B (NF-κB), while enhancing protective mediators including suppressor of cytokine signaling 3 (SOCS3), Growth Associated Protein 43 (Gap43), and Syntaxin12. Brn3a expression and retinal architecture were preserved, apoptotic cell death reduced, and glial activation suppressed. Moreover, BoNT/A decreased mitochondrial ROS accumulation, restored voltage-dependent anion channel 1 (VDAC1) distribution, and partially stabilized intracellular pH. These findings indicate that BoNT/A mitigates oxidative stress and inflammation in hypoxia-driven retinal injury, at least in part, via modulation of the Nox2–Hv1–ROS axis, and support its potential as a therapeutic candidate for ocular disorders associated with hypoxia and neuroinflammation. Full article

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

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18 pages, 6310 KB

Open AccessArticle

The Chromatin Remodeler Chd8 Regulates Hematopoietic Stem and Progenitor Cell Survival and Differentiation During Zebrafish Embryogenesis

by Abrar Ahmed, Xiaona Wei, Dan Zhong, Rahat Ullah, Wei Li and Lili Jing

Int. J. Mol. Sci. 2025, 26(21), 10805; https://doi.org/10.3390/ijms262110805 - 6 Nov 2025

Viewed by 505

Abstract

Chromodomain helicase DNA-binding protein 8 (CHD8), a frequently mutated gene in autism spectrum disorder (ASD), is an ATP-dependent chromatin remodeler with emerging roles in hematopoiesis. While CHD8 is known to maintain hematopoietic stem and progenitor cells (HSPCs) in the bone marrow, its function [...] Read more.

Chromodomain helicase DNA-binding protein 8 (CHD8), a frequently mutated gene in autism spectrum disorder (ASD), is an ATP-dependent chromatin remodeler with emerging roles in hematopoiesis. While CHD8 is known to maintain hematopoietic stem and progenitor cells (HSPCs) in the bone marrow, its function during developmental hematopoiesis remains undefined. Here, using a zebrafish model, we demonstrate that chd8 loss severely depletes the HSPC pool in the caudal hematopoietic tissue through a p53-dependent apoptotic mechanism. Furthermore, chd8^−/− embryos exhibit a p53-independent expansion of myelopoiesis. chd8 deficiency upregulates brd4, which promotes systemic inflammatory cytokine expression. Inhibiting brd4 alleviates cytokine expression, suppresses excessive myelopoiesis, and restores HSPC development. Our findings reveal a dual regulatory mechanism in which chd8 governs HSPC development by repressing p53-mediated apoptosis and constraining brd4-driven immune cell differentiation. Full article

(This article belongs to the Special Issue Zebrafish as a Model for Biomedical Studies—2nd Edition)

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36 pages, 1826 KB

Open AccessReview

Platelet-Rich Plasma (PRP): Molecular Mechanisms, Actions and Clinical Applications in Human Body

by Wen-Shan Wu, Li-Ru Chen and Kuo-Hu Chen

Int. J. Mol. Sci. 2025, 26(21), 10804; https://doi.org/10.3390/ijms262110804 - 6 Nov 2025

Viewed by 3615

Abstract

Platelet-rich plasma (PRP) is an autologous blood-derived concentrate increasingly utilized in regenerative medicine for its ability to accelerate healing and tissue repair. PRP is broadly classified by leukocyte content, fibrin architecture, and platelet concentration, with classification systems developed to standardize characterization. Preparation methods, [...] Read more.

Platelet-rich plasma (PRP) is an autologous blood-derived concentrate increasingly utilized in regenerative medicine for its ability to accelerate healing and tissue repair. PRP is broadly classified by leukocyte content, fibrin architecture, and platelet concentration, with classification systems developed to standardize characterization. Preparation methods, including single- or double-spin centrifugation and buffy coat techniques, influence the final composition of PRP, determining the relative proportions of platelets, leukocytes, plasma proteins, and extracellular vesicles. These components act synergistically, with platelets releasing growth factors (e.g., VEGF, PDGF, TGF-β) that stimulate angiogenesis and matrix synthesis, leukocytes providing immunomodulation, plasma proteins facilitating scaffolding, and exosomes regulating intercellular signaling. Mechanistically, PRP enhances tissue repair through four key pathways: platelet adhesion molecules promote hemostasis and cell recruitment; immunomodulation reduces pro-inflammatory cytokines and favors M2 macrophage polarization; angiogenesis supports vascular remodeling and nutrient delivery; and serotonin-mediated pathways contribute to analgesia. These processes establish a regenerative microenvironment that supports both structural repair and functional recovery. Clinically, PRP has been applied across multiple specialties. In orthopedics, it promotes tendon, cartilage, and bone healing in conditions such as tendinopathy and osteoarthritis. In dermatology, PRP enhances skin rejuvenation, scar remodeling, and hair restoration. Gynecology has adopted PRP for ovarian rejuvenation, endometrial repair, and vulvovaginal atrophy. In dentistry and oral surgery, PRP accelerates wound closure and osseointegration, while chronic wound care benefits from its angiogenic and anti-inflammatory effects. PRP has also favored gingival recession coverage, regeneration of intrabony periodontal defects, and sinus grafting. Although preparation heterogeneity remains a challenge, PRP offers a versatile, biologically active therapy with expanding clinical utility. Full article

(This article belongs to the Section Biochemistry)

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22 pages, 9081 KB

Open AccessArticle

Hydrophobic Drug Delivery Platforms Based on Covalent Organic Frameworks for Combined Treatment of Alzheimer’s Disease

by Yun Zhao, Ziwei Wang, Enpeng Xi, Fuming Yang and Nan Gao

Int. J. Mol. Sci. 2025, 26(21), 10803; https://doi.org/10.3390/ijms262110803 - 6 Nov 2025

Viewed by 477

Abstract

Alzheimer’s disease (AD) is a complex neurodegenerative disease. The pathogenesis of AD remains incompletely understood. It is characterized by a variety of neuropathological changes, including neuroinflammation, neuronal loss and synaptic damage. Multiple pathological changes make achieving good therapeutic effects with a single drug [...] Read more.

Alzheimer’s disease (AD) is a complex neurodegenerative disease. The pathogenesis of AD remains incompletely understood. It is characterized by a variety of neuropathological changes, including neuroinflammation, neuronal loss and synaptic damage. Multiple pathological changes make achieving good therapeutic effects with a single drug treatment difficult, and using multiple drugs for combination therapy is currently the most effective method. Currently, the mainstay drugs used for AD treatment are hydrophobic drugs, such as curcumin, donepezil, and resveratrol. Because hydrophobic drugs cannot dissolve in bodily fluids and often aggregate or precipitate, their efficacy is greatly reduced. Therefore, there is an urgent need for a drug carrier that can effectively load and continuously release drugs. However, currently, there are few drug carriers that can achieve efficient co-loading of multiple hydrophobic drugs. Therefore, three of two-dimensional imine covalent organic frameworks (COFs) with different monomers were synthesized through rational design and screening. These three synthesized COFs are simultaneously loaded with curcumin (CUR) and benzofurazan (BZ) to achieve combined therapy. The results indicate that among this series of synthesized COFs, the COF synthesized from 4,4′,4″-(1,3,5-Triazine-2,4,6-triyl) trianiline and benzene-1,3,5-tricarboxaldehyde (COF-TB) exhibits optimal hydrophobic drug-loading capacity, enabling effective co-loading of CUR and BZ (BC@COF-TB). After treatment with BC@COF-TB, the cognitive function of 5×FAD mice was significantly improved. The COF platform provides a new way to deliver hydrophobic drugs for AD treatment. Full article

(This article belongs to the Special Issue Nanodiagnosis and Treatment System for Human Health)

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23 pages, 5052 KB

Open AccessArticle

Temporal Analysis of Embryonic Epidermal Morphogenesis in Caenorhabditis elegans

by Fangzheng Li, Peiyue Li, Mao Onishi, Law King Chuen, Yukihiko Kubota and Masahiro Ito

Int. J. Mol. Sci. 2025, 26(21), 10802; https://doi.org/10.3390/ijms262110802 - 6 Nov 2025

Viewed by 609

Abstract

The development of epidermis plays a central role in driving the morphogenesis of the Caenorhabditis elegans embryo. However, current research on epidermal morphogenesis focuses disproportionately on overt phenotypic abnormalities, potentially overlooking the crucial role of developmental timing. In this study, we developed a [...] Read more.

The development of epidermis plays a central role in driving the morphogenesis of the Caenorhabditis elegans embryo. However, current research on epidermal morphogenesis focuses disproportionately on overt phenotypic abnormalities, potentially overlooking the crucial role of developmental timing. In this study, we developed a modular two-step deep learning-based image analysis pipeline. First, we used ResU-Net to extract completely developed embryos and suppress noise; second, ResNet was used to predict the corresponding embryonic stage. The predicted probabilities and their corresponding embryonic time points were subsequently utilized to construct a developmental timeline. Combining this pipeline with differential interference contrast time-lapse microscopy, we dynamically tracked the timeline of epidermal morphogenesis in RNAi-treated embryos (ajm-1, tes-1, leo-1) and mutant embryos (clk-1). By statistically comparing the duration of each embryonic stage, our approach enabled the detection of stage-specific developmental timing without relying on overt phenotypic abnormalities or fluorescent markers, successfully recapitulating and extending the known roles of these genes from a temporal perspective. Our work underscores the importance of incorporating developmental timing into morphogenetic analysis, offering a novel framework for revealing subtle developmental processes, deepening the understanding of morphogenetic dynamics, and bridging the methodological gap in C. elegans embryology. Full article

(This article belongs to the Section Molecular Biology)

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33 pages, 4181 KB

Open AccessArticle

Synthesis, Physicochemical Characterization, and Biocidal Evaluation of Three Novel Aminobenzoic Acid-Derived Schiff Bases Featuring Intramolecular Hydrogen Bonding

by Alexander Carreño, Vania Artigas, Belén Gómez-Arteaga, Evys Ancede-Gallardo, Marjorie Cepeda-Plaza, Jorge I. Martínez-Araya, Roxana Arce, Manuel Gacitúa, Camila Videla, Marcelo Preite, María Carolina Otero, Catalina Guerra, Rubén Polanco, Ignacio Fuentes, Pedro Marchant, Osvaldo Inostroza, Fernando Gil and Juan A. Fuentes

Int. J. Mol. Sci. 2025, 26(21), 10801; https://doi.org/10.3390/ijms262110801 - 6 Nov 2025

Viewed by 727

Abstract

Metal-free aminobenzoic acid-derived Schiff bases are attractive antimicrobial leads because their azomethine (–C=N–) functionality enables tunable electronic properties and target engagement. We investigated whether halogenation on the phenolic ring would modulate the redox behavior and enhance antibacterial potency, and hypothesized that heavier halogens [...] Read more.

Metal-free aminobenzoic acid-derived Schiff bases are attractive antimicrobial leads because their azomethine (–C=N–) functionality enables tunable electronic properties and target engagement. We investigated whether halogenation on the phenolic ring would modulate the redox behavior and enhance antibacterial potency, and hypothesized that heavier halogens would favorably tune physicochemical and electronic descriptors. We synthesized three derivatives (SB-3/Cl, SB-4/Br, and SB-5/I) and confirmed their structures using FTIR, ¹H- and ¹³C-NMR, UV-Vis, and HRMS. For SB-5, single-crystal X-ray diffraction and Hirshfeld analysis verified the intramolecular O–H⋯N hydrogen bond and key packing contacts. Cyclic voltammetry revealed an irreversible oxidation (aminobenzoic ring) and, for the halogenated series, a reversible reduction associated with the imine; peak positions and reversibility trends are consistent with halogen electronic effects and DFT-based MEP/LHS descriptors. Antimicrobial testing showed that SB-5 was selectively potent against Gram-positive aerobes, with low-to-mid micromolar MICs across the panel. Among anaerobes, activity was more substantial: Clostridioides difficile was inhibited at 0.1 µM, and SB-3/SB-5 reduced its sporulation at sub-MICs, while Blautia coccoides was highly susceptible (MIC 0.01 µM). No activity was detected against Gram-negative bacteria at the tested concentrations. In the fungal assay, Botrytis cinerea displayed only a transient fungistatic response without complete growth inhibition. In mammalian cells (HeLa), the compounds displayed clear concentration-dependent behavior. Overall, halogenation, particularly iodination, emerges as a powerful tool to couple redox tuning with selective Gram-positive activity and a favorable cellular tolerance window, nominating SB-5 as a promising scaffold for further antimicrobial optimization. Full article

(This article belongs to the Special Issue Feature Papers in ‘Physical Chemistry and Chemical Physics’: Fourth Edition)

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23 pages, 2878 KB

Open AccessReview

Ceramide as a Biomarker for HFpEF in Women: Menopause, Aging, and Pregnancy

by Ruth R. Magaye, David M. Kaye and Bing H. Wang

Int. J. Mol. Sci. 2025, 26(21), 10800; https://doi.org/10.3390/ijms262110800 - 6 Nov 2025

Viewed by 1092

Abstract

Heart failure with preserved ejection fraction (HFpEF) currently accounts for half of the heart failure (HF) cases world-wide, affecting nearly 32 million people. HFpEF has a skewed prevalence toward females and those older than 65 years old. The pathophysiology of HFpEF is suggestive [...] Read more.

Heart failure with preserved ejection fraction (HFpEF) currently accounts for half of the heart failure (HF) cases world-wide, affecting nearly 32 million people. HFpEF has a skewed prevalence toward females and those older than 65 years old. The pathophysiology of HFpEF is suggestive of a conglomerate of inflammatory, hypertensive, as well as metabolic dysfunction, giving rise to the syndrome. Disruptions in ceramide metabolism do occur in heart failure as well as within the HFpEF-associated risk factors, both modifiable inflammation, obesity, hypertension, diabetes, and non-modifiable-aging, and female sex. The focus of this review is to draw attention to the links between changes in female biophysiology, such as pregnancy, menopause and aging, in which ceramide is dysregulated and consequently gives rise to the same pathologies that are labeled as risk factors for HFpEF. Our objective is to highlight ceramides as potential biomarkers for prevention and initial diagnostic tools for HFpEF, especially for women later in life. Full article

(This article belongs to the Special Issue Sphingolipid Metabolism and Signaling in Health and Diseases)

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Graphical abstract

17 pages, 3076 KB

Open AccessArticle

LoQANT: An ImageJ Plugin for Quantifying Nuclear Staining in Immunohistochemistry and Immunofluorescence

by Katerina Cizkova

Int. J. Mol. Sci. 2025, 26(21), 10799; https://doi.org/10.3390/ijms262110799 - 6 Nov 2025

Viewed by 590

Abstract

A large number of regulatory proteins are found in both the cytoplasm and the nucleus. Changes in their nuclear abundance are important for cellular signalling, biological activity, and disease mechanisms. Accurate quantification of nuclear staining is therefore essential in studies of cellular function, [...] Read more.

A large number of regulatory proteins are found in both the cytoplasm and the nucleus. Changes in their nuclear abundance are important for cellular signalling, biological activity, and disease mechanisms. Accurate quantification of nuclear staining is therefore essential in studies of cellular function, therapeutic targeting, drug design, and drug resistance. However, manual scoring is time-consuming, unsuitable for high-throughput applications, and introduces potential bias. As expected, manual scoring by six observers with varying levels of expertise led to highly variable results. Moreover, it was far from achieving good interobserver reliability. To overcome these limitations, LoQANT (Localisation and Quantification of Antigen Nuclear sTaining), an open, freely available ImageJ plugin, was developed for reliable and efficient quantification of nuclear signals. LoQANT is a single cell-based approach to assess the proportion of cells with a positive nuclear signal, independent of cytoplasmic staining, in both immunohistochemically and fluorescently stained samples across various sample types. It also provides semiquantitative and quantitative measurements of nuclear staining intensity. The script, its version for batch analysis, and complete user guide are available at GitHub. Full article

(This article belongs to the Section Molecular Immunology)

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53 pages, 4809 KB

Open AccessReview

Epigallocatechin Gallate as a Molecular Therapeutic in Heart Failure and Cardio-Oncology: Mechanistic Pathways and Translational Perspectives

by Faika Ajaz, Jewel Haddad, Bintul Huda, Maryam Yousuf, Rajashree Patnaik, Farida Bhurka and Yajnavalka Banerjee

Int. J. Mol. Sci. 2025, 26(21), 10798; https://doi.org/10.3390/ijms262110798 - 6 Nov 2025

Viewed by 893

Abstract

The global burden of heart failure (HF) continues to escalate, with a lifetime risk approaching one in four adults in the United States. Concurrently, advances in cancer therapeutics have created a burgeoning population of long-term survivors, who now face the significant morbidity and [...] Read more.

The global burden of heart failure (HF) continues to escalate, with a lifetime risk approaching one in four adults in the United States. Concurrently, advances in cancer therapeutics have created a burgeoning population of long-term survivors, who now face the significant morbidity and mortality of chemotherapy-induced cardiovascular disease (CVD). This review addresses the critical overlap of these two pathologies, which share fundamental drivers such as oxidative stress, inflammation, and metabolic dysregulation. Epigallocatechin gallate (EGCG), the most abundant and biologically active polyphenol in green tea, has demonstrated pleiotropic bioactivity in preclinical models, encompassing potent antioxidant, anti-inflammatory, and anti-apoptotic properties. The central aim of this review is to provide a critical and comprehensive synthesis of the evidence supporting EGCG’s dual protective role. This review dissects its molecular mechanisms in modulating key pathways in HF and cardio-oncology, evaluates its translational potential, and importantly, delineates the significant gaps that must be addressed for its clinical application. This analysis uniquely positions EGCG not merely as a nutraceutical, but as a multi-target molecular therapeutic capable of simultaneously addressing the convergent pathological cascades of heart failure and cancer-related cardiotoxicity. The synthesis of preclinical evidence with a critical analysis of its translational barriers offers a novel perspective and a strategic roadmap for future research. Full article

(This article belongs to the Section Molecular Biology)

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17 pages, 580 KB

Open AccessArticle

Association of BMAL1 and CLOCK Gene Polymorphisms with Preeclampsia Risk with Subtype Analysis

by Fan Xia, Peiwen Wang, Ziye Li, Jiehua Wei, Jianhui Wei, Yuhang Wu, Chu Liu, Shanyu Lin, Suyan Guo, Linbin He, Mengshi Chen, Lizhang Chen and Tingting Wang

Int. J. Mol. Sci. 2025, 26(21), 10797; https://doi.org/10.3390/ijms262110797 - 6 Nov 2025

Viewed by 515

Abstract

Preeclampsia (PE), a major cause of maternal and perinatal morbidity, is a hypertensive pregnancy disorder with poorly defined pathogenesis. While dysregulation of core circadian genes including brain and muscle ARNT-like 1 (BMAL1; also termed ARNTL) and circadian locomotor output cycles [...] Read more.

Preeclampsia (PE), a major cause of maternal and perinatal morbidity, is a hypertensive pregnancy disorder with poorly defined pathogenesis. While dysregulation of core circadian genes including brain and muscle ARNT-like 1 (BMAL1; also termed ARNTL) and circadian locomotor output cycles kaput (CLOCK) has been implicated in PE, the contribution of their genetic polymorphisms to PE remains unclear. In this case–control study, polymorphisms in BMAL1 and CLOCK were genotyped using MassARRAY in 202 PE patients (97 early-onset [eoPE], 105 late-onset [loPE]) and 400 controls. Following genotyping and linkage disequilibrium-pruning (r² > 0.8) to retain representative tag SNPs, the final set for association analysis comprised three non-redundant BMAL1 SNPs (rs4757144, rs11022780, rs969485) and one CLOCK SNP (rs1048004). After confounder adjustment, no significant associations were detected for CLOCK variants, whereas the BMAL1 rs11022780 variant demonstrated a significant protective effect against PE (TT vs. CC: OR = 0.26 [95% CI 0.09–0.78]; recessive model: OR = 0.25 [95% CI 0.09–0.74]), particularly in the eoPE subgroup. Expression quantitative trait locus (eQTL) analysis confirmed that this SNP correlated with BMAL1 mRNA expression in whole blood, and protein–protein interaction analysis highlighted BMAL1′s central role in circadian networks, implying a genetically influenced regulatory mechanism of PE through BMAL1 expression. Full article

(This article belongs to the Special Issue Molecular Research on Reproductive Physiology and Endocrinology)

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29 pages, 802 KB

Open AccessReview

Endometrial Microbiome and Reproductive Receptivity: Diverse Perspectives

by Galina Stoyancheva, Nikolina Mihaylova, Maria Gerginova and Ekaterina Krumova

Int. J. Mol. Sci. 2025, 26(21), 10796; https://doi.org/10.3390/ijms262110796 - 6 Nov 2025

Viewed by 1562

Abstract

The human endometrium, previously considered a sterile environment, is now recognized as a low-biomass but biologically active microbial niche critical to reproductive health. Advances in sequencing technologies, particularly shotgun metagenomics, have provided unprecedented insights into the taxonomic and functional complexity of the endometrial [...] Read more.

The human endometrium, previously considered a sterile environment, is now recognized as a low-biomass but biologically active microbial niche critical to reproductive health. Advances in sequencing technologies, particularly shotgun metagenomics, have provided unprecedented insights into the taxonomic and functional complexity of the endometrial microbiome. While 16S rRNA sequencing has delineated the distinction between Lactobacillus-dominant and non-dominant microbial communities, shotgun metagenomics has revealed additional diversity at the species and strain level, uncovering microbial signatures that remain undetected by amplicon-based approaches. Current evidence supports the association of Lactobacillus dominance with endometrial homeostasis and favorable reproductive outcomes. Dysbiosis, characterized by increased microbial diversity and enrichment of anaerobic taxa such as Gardnerella, Atopobium, Prevotella, and Streptococcus, is linked to chronic endometritis, implantation failure, and adverse IVF results. Beyond compositional differences, the endometrial microbiome interacts with the host through immunological, metabolic, and epigenetic mechanisms. These interactions modulate cytokine signaling, epithelial barrier integrity, and receptivity-associated gene expression, ultimately influencing embryo implantation. However, discrepancies between published studies reflect the lack of standardized protocols for sampling, DNA extraction, and bioinformatic analysis, as well as the inherent challenges of studying low-biomass environments. Factors such as geography, ethnicity, hormonal status, and antibiotic exposure further contribute to interindividual variability. Culturomics approaches complement sequencing by enabling the isolation of viable bacterial strains, offering perspectives for microbiome-based biotherapeutics. Emerging 3D endometrial models provide additional tools to dissect microbiome–host interactions under controlled conditions. Taken together, the growing body of data highlights the potential of endometrial microbiome profiling as a biomarker for reproductive success and as a target for personalized interventions. Future research should focus on integrating multi-omics approaches and functional analyses to establish causal relationships and translate findings into clinical practice. This review gives a new insight into current knowledge on the uterine microbiome and its impact on implantation success, analyzed through the lenses of microbiology, immunology, and oxidative stress. Full article

(This article belongs to the Special Issue Molecules and Mechanisms of Oxidative Stress Relevant in the Pathophysiology of Diseases)

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11 pages, 844 KB

Open AccessArticle

Urine Progesterone Level as a Diagnostics Tool to Evaluate the Need for Luteal Phase Rescue in Hormone Replacement Therapy Frozen Embryo Transfer Cycles

by Linette Yde Hansen, Takeshi Fujisawa, Betina Boel Povlsen, Rita Jakubcionyte Laursen, Mette Brix Jensen, Peter Humaidan and Birgit Alsbjerg

Int. J. Mol. Sci. 2025, 26(21), 10795; https://doi.org/10.3390/ijms262110795 - 6 Nov 2025

Viewed by 816

Abstract

Additional progesterone administration during the luteal phase enhances reproductive outcomes in Hormone Replacement Therapy Frozen Embryo Transfer (HRT-FET) cycles in patients with low serum progesterone (P4). In this study we wanted to explore the use of urine P4 as a diagnostic tool during [...] Read more.

Additional progesterone administration during the luteal phase enhances reproductive outcomes in Hormone Replacement Therapy Frozen Embryo Transfer (HRT-FET) cycles in patients with low serum progesterone (P4). In this study we wanted to explore the use of urine P4 as a diagnostic tool during the luteal phase. This prospective observational cohort included a total of 464 HRT-FET cycles. The protocol entailed oral oestradiol (6 mg/24 h), followed by vaginal micronised progesterone (400 mg/12 h). On the day of blastocyst transfer, urine and serum samples were collected. Urine samples were analysed using an ARCHITECT automated immunoassay. A significant difference was found in median urine P4 between patients with serum P4 higher or lower than 11 ng/mL: 6400 ng/mL IQR [2528; 11,930] vs. 3408 ng/mL IQR [592; 6688], p < 0.001. The optimal cut-off to achieve live birth was a urine P4 ≥ 4000 ng/mL. The live birth rate was significantly higher in patients with urine P4 ≥ 4000 ng/mL, 48% (107/222) vs. 35% (45/130), respectively (p = 0.013). The odds ratio for live birth was 1.8 in patients with urine P4 ≥ 4000 ng/mL, 95% CI [1.067; 3.018], p = 0.028. The findings of the present study suggest that urine progesterone could be a valuable diagnostic tool to evaluate the need for additional progesterone in HRT-FET cycles. Full article

(This article belongs to the Special Issue Understanding Reproductive Health and Infertility: Molecular and Genetic Insights)

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19 pages, 392 KB

Open AccessReview

MicroRNAs as Emerging Therapeutic Targets Modulating the Tumor Microenvironment in Head and Neck Squamous Cell Carcinoma

by Roxana Daniela Brata, Lavinia Marcut, Alina Cristina Barb, Alexia Manole, Alexandru Ciolofan, Cristina Stefania Dumitru, Flavia Zara and Raul Patrascu

Int. J. Mol. Sci. 2025, 26(21), 10794; https://doi.org/10.3390/ijms262110794 - 6 Nov 2025

Viewed by 788

Abstract

Head and neck squamous cell carcinoma (HNSCC) remains one of the most aggressive solid tumors, characterized by marked molecular heterogeneity and a complex tumor microenvironment (TME). Recent evidence highlights the pivotal role of microRNAs (miRNAs) in regulating tumor progression, immune evasion, angiogenesis, and [...] Read more.

Head and neck squamous cell carcinoma (HNSCC) remains one of the most aggressive solid tumors, characterized by marked molecular heterogeneity and a complex tumor microenvironment (TME). Recent evidence highlights the pivotal role of microRNAs (miRNAs) in regulating tumor progression, immune evasion, angiogenesis, and stromal remodeling. This review synthesizes current insights into miRNA-mediated molecular pathways that modulate the TME in HNSCC and discusses emerging therapeutic strategies, including nanocarrier- and exosome-based miRNA delivery systems, targeting these molecules. Key miRNAs, including miR-21, miR-146a, and miR-221, orchestrate bidirectional signaling between cancer cells, fibroblasts, and immune infiltrates, thereby shaping tumor aggressiveness and therapy resistance. Advances in nanotechnology have facilitated the development of miRNA-based therapeutics—such as mimics, antagomiRs, and exosome-mediated systems—capable of restoring physiological expression patterns and reprogramming the TME toward an anti-tumor state. However, clinical translation remains hindered by challenges in targeted delivery, molecular stability, and tumor heterogeneity. By integrating molecular and translational perspectives, this review underscores how miRNA-targeting strategies may evolve into a new generation of precision therapies, bridging the gap between molecular oncology and personalized treatment of head and neck cancer. Full article

(This article belongs to the Special Issue Emerging Strategies Targeting Solid Tumors and the Tumor Microenvironment)

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18 pages, 11741 KB

Open AccessArticle

HSA^LR Mice Exhibit Co-Expression of Proteostasis Genes Prior to Development of Muscle Weakness

by Dusan M. Lazic, Vladimir M. Jovanovic, Jelena Karanovic, Dusanka Savic-Pavicevic and Bogdan Jovanovic

Int. J. Mol. Sci. 2025, 26(21), 10793; https://doi.org/10.3390/ijms262110793 - 6 Nov 2025

Viewed by 607

Abstract

Myotonic dystrophy type 1 (DM1) is a progressive multisystemic disease caused by a CTG repeat expansion in the DMPK gene. The toxic mutant mRNA sequesters MBNL proteins, disrupting global RNA metabolism. Although alternative splicing in DM1 skeletal muscle pathology has been extensively studied, [...] Read more.

Myotonic dystrophy type 1 (DM1) is a progressive multisystemic disease caused by a CTG repeat expansion in the DMPK gene. The toxic mutant mRNA sequesters MBNL proteins, disrupting global RNA metabolism. Although alternative splicing in DM1 skeletal muscle pathology has been extensively studied, early-stage transcriptomic changes remained uncharacterized. To gain deeper and contextual insight into DM1 transcriptome, we performed the first Weighted Gene Co-expression Network Analysis (WGCNA) on skeletal muscle RNA sequencing data from the widely used DM1 mouse model HSA^LR (~250 CTG repeats). We identified 532 core genes using data from 16-week-old mice, an age before the onset of muscle weakness. Additional differential expression analysis across multiple HSA^LR datasets revealed 42 common up-regulated coding and non-coding genes. Within identified core genes, the pathway gene-pair signature analysis enabled contextual selection of functionally related genes involved in maintaining proteostasis, including endoplasmic reticulum (ER) protein processing, the ubiquitin-proteasome system (UPS), macroautophagy and mitophagy, and muscle contraction. The enrichment of ER protein processing with prevailing core genes related to ER-associated degradation suggests adaptive chaperone and UPS activation, while core genes such as Ambra1, Mfn2, and Usp30 indicate adaptations in mitochondrial quality control. Coordinated early alterations in processes maintaining protein homeostasis, critical for muscle mass and function, possibly reflect a response to cellular stress due to repeat expansion and appears before muscle weakness development. Although the study relies exclusively on transcriptomic analyses, it offers a comprehensive, hypothesis-generating perspective that pinpoints candidate pathways, preceding muscle weakness, for future mechanistic validation. Full article

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

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16 pages, 1040 KB

Open AccessArticle

Cell-Free DNA Hypermethylation in Patients with Acute Pancreatitis

by Hassan Al-Mashat, Daniel Roger Baddoo, Søren Lundbye-Christensen, Poul Henning Madsen, Inge Søkilde Pedersen, Henrik B. Krarup, Benjamin Emil Stubbe, Ole Thorlacius-Ussing and Stine Dam Henriksen

Int. J. Mol. Sci. 2025, 26(21), 10792; https://doi.org/10.3390/ijms262110792 - 6 Nov 2025

Viewed by 433

Abstract

Cell-free DNA (cfDNA) promoter hypermethylation shows promise as a blood-based biomarker for pancreatic cancer, and similar alterations may occur in acute pancreatitis (AP). This study investigated the cfDNA hypermethylation profile of AP patients over time, compared with healthy controls, and its association with [...] Read more.

Cell-free DNA (cfDNA) promoter hypermethylation shows promise as a blood-based biomarker for pancreatic cancer, and similar alterations may occur in acute pancreatitis (AP). This study investigated the cfDNA hypermethylation profile of AP patients over time, compared with healthy controls, and its association with AP severity markers. A prospective longitudinal study including hospitalized AP patients and healthy controls was conducted. Methylation-specific PCR of a 23-gene panel was performed on plasma collected at inclusion (T0), 6 weeks (T6W), 6 months (T6M), and 7–8 years (T8Y). Associations between gene hypermethylation and clinical markers of AP severity—CRP, leukocyte count, creatinine, hospital stay, and complications—were evaluated. AP patients had a significantly higher mean number of hypermethylated genes at T0 (7.4, 95% CI: 6.8–8.0) compared with the controls (3.3, 95% CI: 2.8–3.8; p < 0.01). The mean number decreased over time to 3.2 (95% CI: 2.4–4.1) at T8Y. Total hypermethylation was positively associated with CRP (ρ = 0.39; p = 0.0018), leukocytes (ρ = 0.35; p = 0.0052), and hospital stay (ρ = 0.27; p = 0.0375). AP patients exhibited significantly higher cfDNA hypermethylation at disease onset, which normalized over time. Total hypermethylation showed positive associations with several markers of AP severity. Full article

(This article belongs to the Special Issue Pancreatic Disease: From Molecular Basis to Novel Therapies—2nd Edition)

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12 pages, 752 KB

Open AccessReview

Bilirubin Photoisomers in Neonatal Jaundice

by Dennis Lindqvist, Magnus Hansson, Mercy Thomas, Christian V. Hulzebos, Libor Vitek, Andries Blokzijl and Miranda van Berkel

Int. J. Mol. Sci. 2025, 26(21), 10791; https://doi.org/10.3390/ijms262110791 - 6 Nov 2025

Viewed by 1016

Abstract

Phototherapy is the standard treatment for neonatal hyperbilirubinemia. During phototherapy, the highly lipophilic bilirubin is converted into more hydrophilic photoisomers, which can be more easily excreted from the body. This process typically lowers bilirubin levels to non-harmful concentrations. However, despite decades of research [...] Read more.

Phototherapy is the standard treatment for neonatal hyperbilirubinemia. During phototherapy, the highly lipophilic bilirubin is converted into more hydrophilic photoisomers, which can be more easily excreted from the body. This process typically lowers bilirubin levels to non-harmful concentrations. However, despite decades of research into the formation and role of bilirubin photoisomers, methodological limitations and the compound’s complex biochemistry have hindered comprehensive understanding. This review provides an updated overview of current knowledge on bilirubin photoisomers, including their basic chemistry, analytical quantification, clinical relevance, and future research directions. Improved insight into the mechanism of photoisomer formation and kinetics may inform optimization of phototherapy parameters, including light intensity and wavelength, and offer additional indicators of treatment efficacy beyond total bilirubin concentration. Advances in sensitive and standardized mass spectrometry techniques now enable more accurate measurement of different bilirubin isomers and serve as a first step towards a deeper insight into the clinical relevance of photoisomers. Full article

(This article belongs to the Special Issue Bilirubin: Health Challenges and Opportunities)

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45 pages, 1518 KB

Open AccessReview

Hydrogen Sulfide in Balneology: Physiology, Evidence, and Clinical Translation

by Jose Manuel Carbajo, Francisco Maraver, Lorena Vela and Constantin Munteanu

Int. J. Mol. Sci. 2025, 26(21), 10790; https://doi.org/10.3390/ijms262110790 - 6 Nov 2025

Viewed by 1432

Abstract

This review integrates the biology and clinical translation of hydrogen sulfide (H₂S) in balneology. It frames H₂S as a gasotransmitters with dual chemical and biological actions and summarizes the H₂S/HS⁻ equilibrium as a function of pH, [...] Read more.

This review integrates the biology and clinical translation of hydrogen sulfide (H₂S) in balneology. It frames H₂S as a gasotransmitters with dual chemical and biological actions and summarizes the H₂S/HS⁻ equilibrium as a function of pH, temperature, and oxygenation, which governs bioaccessibility in sulfurous waters. Endogenous and exogenous sources, transport, and mitochondrial catabolism are outlined, together with core cellular mechanisms: protein persulfidation; activation of Nrf2/ARE; modulation of NF-κB; regulation of ion channels; and engagement of PI3K/Akt, MAPK/ERK, and Wnt pathways, plus epigenetic interactions with HDACs and sirtuins. Preclinical and clinical evidence in dermatology, musculoskeletal disease, and respiratory care is synthesized, alongside metabolic, cardiovascular, gastrointestinal, and renal effects. Technical aspects that preserve the bioactive fraction of H₂S while meeting environmental safety limits are highlighted. Routes of administration (bathing, peloids, inhalation, and drinking cures) and key operational parameters are described. Overall, the review links physicochemical and molecular foundations with clinical indications for sulfurous waters and derivatives and identifies opportunities for research and development in H₂S donors and thermal cosmetics without extrapolating beyond the available data. Full article

(This article belongs to the Section Biochemistry)

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13 pages, 555 KB

Open AccessReview

Update on Nicotinamide and Its Application in the Management of Glaucoma

by Ta-Hung Chiu, Shih-Heng Hung, Chiao-Hsin Lan, Wei-Ting Yen and Da-Wen Lu

Int. J. Mol. Sci. 2025, 26(21), 10789; https://doi.org/10.3390/ijms262110789 - 6 Nov 2025

Viewed by 2021

Abstract

Glaucoma continues to be a primary contributor to permanent vision loss worldwide, frequently advancing even when intraocular pressure management is clinically adequate. Accumulating research emphasizes the metabolic susceptibility of retinal ganglion cells (RGCs), specifically concerning the progressive depletion of nicotinamide adenine dinucleotide (NAD [...] Read more.

Glaucoma continues to be a primary contributor to permanent vision loss worldwide, frequently advancing even when intraocular pressure management is clinically adequate. Accumulating research emphasizes the metabolic susceptibility of retinal ganglion cells (RGCs), specifically concerning the progressive depletion of nicotinamide adenine dinucleotide (NAD⁺), a pivotal coenzyme fundamental to mitochondrial energy production and cellular survival mechanisms. As a key biosynthetic precursor in NAD⁺ synthesis pathways, nicotinamide (NAM), a form of vitamin B3, has exhibited significant neuroprotective properties across various preclinical experimental models and preliminary clinical investigations, demonstrating enhanced preservation of RGC morphology and physiological function. This comprehensive review systematically examines the current body of evidence supporting NAM’s therapeutic utility in glaucomatous neuroprotection, focusing particularly on underlying metabolic pathways, obstacles in clinical translation, and prospective therapeutic applications. Through systematic integration of data from cellular and molecular research, animal experimental studies, and population-based epidemiological investigations, we establish a conceptual framework for repurposing NAM as an innovative complementary therapeutic strategy in comprehensive glaucoma care, addressing key considerations for future clinical development including optimal dosing strategies, delivery mechanisms, and patient selection criteria for maximizing therapeutic outcomes in this challenging neurodegenerative condition. Full article

(This article belongs to the Special Issue Molecular Research and Advances in Ocular Disease)

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26 pages, 959 KB

Open AccessReview

Unraveling Type 1 Diabetes: Integrating Microbiome, Metabolomics, and Immunomodulation for Next-Generation Therapies

by Pleun de Groen, Lente C. H. M. Blok, Coco M. Fuhri Snethlage, Nordin M. J. Hanssen, Elena Rampanelli and Max Nieuwdorp

Int. J. Mol. Sci. 2025, 26(21), 10788; https://doi.org/10.3390/ijms262110788 - 6 Nov 2025

Viewed by 1514

Abstract

Type 1 diabetes (T1D) is a chronic autoimmune disease characterized by T-cell-mediated destruction of pancreatic beta cells, resulting in insulin deficiency. Both genetic predisposition and environmental factors contribute to T1D development, with growing evidence implicating the gut microbiome as a critical environmental modulator [...] Read more.

Type 1 diabetes (T1D) is a chronic autoimmune disease characterized by T-cell-mediated destruction of pancreatic beta cells, resulting in insulin deficiency. Both genetic predisposition and environmental factors contribute to T1D development, with growing evidence implicating the gut microbiome as a critical environmental modulator in disease pathogenesis. Gut microbial composition and derived metabolites influence immune homeostasis and autoimmunity. This review summarizes recent advances elucidating immune dysregulations in T1D and novel therapeutic strategies to preserve beta cell function. We discuss approaches such as immune cell engineering, including CAR-Treg therapy, and targeted modulation of immune signaling pathways like JAK-STAT. Furthermore, we explore the role of the gut microbiota and its metabolites in modulating host immunity and describe emerging microbiome-targeting interventions, including fecal microbiota transplantation and metabolite supplementation. These interventions show promise in modulating disease progression in preclinical and early clinical studies. An integrated understanding of immune and microbiome-related mechanisms is critical for developing next-generation therapies. Further research and clinical trials are needed to optimize these approaches and translate them into durable, personalized treatments for individuals with T1D. Full article

(This article belongs to the Special Issue Innovative Targeted Therapies in Inflammatory Diseases)

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18 pages, 2315 KB

Open AccessArticle

Resveratrol Analogs Ameliorate Mitochondrial Impairment and Insulin Resistance in a Streptozotocin-Induced In Vitro Model of Alzheimer’s Disease

by Alexandra Paszternák, Kamilla Varga, Ramá Gyöngyössy, Katinka Tarnóczi, Noémi Sikur, Éva Szökő and Tamás Tábi

Int. J. Mol. Sci. 2025, 26(21), 10787; https://doi.org/10.3390/ijms262110787 - 6 Nov 2025

Viewed by 615

Abstract

Alzheimer’s disease (AD) is characterized by mitochondrial dysfunction, oxidative stress, insulin resistance, and aberrant protein aggregation. Neurodegeneration model with neuronal insulin resistance was induced in SH-SY5Y human neuroblastoma cells by streptozotocin (STZ). We evaluated the neuroprotective effects of resveratrol (RZV) and three structural [...] Read more.

Alzheimer’s disease (AD) is characterized by mitochondrial dysfunction, oxidative stress, insulin resistance, and aberrant protein aggregation. Neurodegeneration model with neuronal insulin resistance was induced in SH-SY5Y human neuroblastoma cells by streptozotocin (STZ). We evaluated the neuroprotective effects of resveratrol (RZV) and three structural analogs: oxyresveratrol (OXI), monomethyl resveratrol (MONO), and trimethyl resveratrol (TRI). Mitochondrial function, plasma membrane integrity, oxidative stress) and autophagy were studied by fluorescent assays. Phosphorylated GSK3 levels were measured by ELISA as an indicator of insulin sensitivity. TRI exhibited significant mitochondrial protective effects and strongly induced autophagy. OXI demonstrated excellent antioxidant activity but showed no detectable mitochondrial protective or autophagy-inducing effects. RZV and MONO exhibited moderate antioxidant effects along with strong insulin-sensitizing and autophagy-inducing properties. Insulin sensitivity was most potently restored by RZV (IC₅₀ = 54 pM) and MONO (IC₅₀ = 50 pM), whereas TRI (IC₅₀ = 160 pM) was less potent, and OXI (IC₅₀ = 97 pM) showed moderate potency. Our findings suggest that the neuroprotective effects of resveratrol analogs significantly depend on their molecular structure and that they exert their beneficial effects through distinct mechanisms. This research may contribute to the development of novel, multi-target compounds for the treatment of neurodegenerative diseases. Full article

(This article belongs to the Special Issue Natural Bioactives and Inflammation, 2nd Edition)

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16 pages, 3402 KB

Open AccessArticle

Butylated Neuropeptide Antagonist Targeting Hypoxia-Induced GRPR Overexpression in Small Cell Lung Cancer

by Suttikiat Deureh, Amira M. Alghamdi, Ayşe Latif, Kaye J. Williams, Roben G. Gieling and Harmesh S. Aojula

Int. J. Mol. Sci. 2025, 26(21), 10786; https://doi.org/10.3390/ijms262110786 - 6 Nov 2025

Viewed by 517

Abstract

Small cell lung cancer (SCLC) is an aggressive neuroendocrine tumour with limited treatment options and a poor prognosis. Hypoxia, a hallmark of solid tumours, contributes to therapeutic resistance and tumour progression. Gastrin-releasing peptide receptor (GRPR) is known to be overexpressed in SCLC; however, [...] Read more.

Small cell lung cancer (SCLC) is an aggressive neuroendocrine tumour with limited treatment options and a poor prognosis. Hypoxia, a hallmark of solid tumours, contributes to therapeutic resistance and tumour progression. Gastrin-releasing peptide receptor (GRPR) is known to be overexpressed in SCLC; however, its regulation under hypoxic conditions is not well described. In this study, we demonstrate that hypoxia significantly enhances GRPR expression in SCLC cell lines, COR-L24 and DMS79, as confirmed by Western blot, immunofluorescence, and flow cytometric analysis of binding with fluorescein isothiocyanate–labelled bombesin (BBN-FITC), a known GRPR ligand. To exploit this upregulation, we synthesised a previously discovered butylated neuropeptide antagonist (BU peptide) using a new method of solid-phase peptide synthesis (SPPS) by Boc chemistry and evaluated its therapeutic potential. BU peptide exhibited potent, dose-dependent cytotoxicity in both cell lines, with significantly greater efficacy under hypoxic conditions compared to normoxia. Mechanistic studies revealed that BU peptide inhibits GRP–GRPR-mediated activation of the PI3K/Akt and MAPK/ERK signalling pathways, known to be key regulators of tumour cell survival and proliferation. Moreover, BU peptide induced robust caspase 3/7-mediated apoptosis, especially under hypoxic conditions. These findings suggest that GRPR is a hypoxia-inducible target in SCLC and demonstrate that a synthetically optimised BU peptide antagonist exerts selective efficacy against hypoxic tumour cells, outperforming conventional chemotherapy agents. These findings provide new mechanistic insights into SCLC and suggest translational potential to inform the development of future treatment strategies for this and other hypoxia-driven malignancies. Full article

(This article belongs to the Section Molecular Pharmacology)

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12 pages, 1410 KB

Open AccessArticle

Promoter Frame Position Affects Strength and Nature of Circadian Oscillations in hPER2 Luciferase Reporters

by Bhavna Kalyanaraman, Gabrielle Villafana, Stephanie R. Taylor and Michelle E. Farkas

Int. J. Mol. Sci. 2025, 26(21), 10785; https://doi.org/10.3390/ijms262110785 - 6 Nov 2025

Viewed by 408

Abstract

The PER2 gene is a crucial component responsible for the proper functioning of the mammalian core circadian clock. The circadian nature of the murine Per2 (mPer2) promoter’s activity has been thoroughly investigated to identify important elements responsible for its oscillatory behavior; [...] Read more.

The PER2 gene is a crucial component responsible for the proper functioning of the mammalian core circadian clock. The circadian nature of the murine Per2 (mPer2) promoter’s activity has been thoroughly investigated to identify important elements responsible for its oscillatory behavior; however, its human counterpart has not. While there are similarities between murine and human core clocks, there are differences and unconserved elements between their promoter sequences that may influence the nature of rhythms. Further, most studies to date have used murine-based sequences in human cell lines. To fully understand the role(s) of and factors involved in the human PER2 (hPER2) gene, human-derived sequences should be used. To this end, we developed two lentiviral luciferase reporters in well-established, circadian model U2OS cells using different hPER2 promoter regions. Their rhythmic nature was compared to that of the standard mPer2 promoter reporter. We found that hPER2 reporters exhibited stronger oscillations than the mPer2 reporter, and that the frame of the hPER2 promoter affected the period and phase. This work introduces a human sequence-based PER2 promoter in U2OS cells, which should be used for further in vitro tracking of hPER2 activity and to understand PER2 gene dynamics, in lieu of the murine iteration. Full article

(This article belongs to the Section Molecular Biology)

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2 pages, 817 KB

Open AccessCorrection

Correction: Zeppieri et al. Isolated and Syndromic Genetic Optic Neuropathies: A Review of Genetic and Phenotypic Heterogeneity. Int. J. Mol. Sci. 2025, 26, 3892

by Marco Zeppieri, Caterina Gagliano, Marco Di Maita, Alessandro Avitabile, Giuseppe Gagliano, Edoardo Dammino, Daniele Tognetto, Maria Francesca Cordeiro and Fabiana D’Esposito

Int. J. Mol. Sci. 2025, 26(21), 10784; https://doi.org/10.3390/ijms262110784 - 6 Nov 2025

Viewed by 267

Abstract

In the original publication [...] Full article

(This article belongs to the Section Molecular Neurobiology)

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19 pages, 1948 KB

Open AccessArticle

Co-Occurrence of RAD21 and TNFAIP3 Mutations in Cornelia de Lange Syndrome with Pustular Psoriasis: Potential Molecular Interactions

by Beatriz E. Orozco, Cindy V. Orozco, Esperanza Meléndez, María F. Mangones, José Valderrama, Adalberto Lobato, Pilar Garavito-Galofre, Jorge I. Vélez and Oscar M. Vidal

Int. J. Mol. Sci. 2025, 26(21), 10783; https://doi.org/10.3390/ijms262110783 - 6 Nov 2025

Viewed by 485

Abstract

Cornelia de Lange Syndrome (CdLS) is a rare multisystem developmental disorder caused primarily by mutations in cohesin complex genes, including RAD21. Psoriasis is a chronic inflammatory skin disease linked to immune dysregulation, notably involving TNFAIP3 (A20), a negative regulator of [...] Read more.

Cornelia de Lange Syndrome (CdLS) is a rare multisystem developmental disorder caused primarily by mutations in cohesin complex genes, including RAD21. Psoriasis is a chronic inflammatory skin disease linked to immune dysregulation, notably involving TNFAIP3 (A20), a negative regulator of NF-κB signaling. Although case reports have suggested a possible coexistence of CdLS and psoriasis, the underlying molecular basis has remained unexplored. Here we report the first case of molecular co-occurrence of CdLS and generalized pustular psoriasis in a patient with novel heterozygous nonsense variant in RAD21 (c.1306C>T, p.Gln436*), pathogenic for CdLS type 4, and a previously unreported truncating variant in TNFAIP3 (c.2199C>A, p.Cys733*), predicted to disrupt NF-κB regulation and classified as a variant of uncertain significance. Structural protein modeling showed significant conformational disruption in RAD21 and partial truncation of the ZnF domains of TNFAIP3, supporting their functional impact. This study is the first to suggest a possible molecular mechanism that may explain the rare co-occurrence of CdLS and psoriasis: RAD21 deficiency disrupts chromatin architecture and immune gene regulation, while TNFAIP3 loss-of-function removes critical NF-κB inhibition, resulting in synergistic developmental and inflammatory phenotypes. Secondary transcriptomic data analysis further suggests that RAD21 knockdown may downregulate TNFAIP3 expression, providing a possible mechanistic intersection. Our findings provide the first molecular evidence linking RAD21 and TNFAIP3, introducing a novel pathogenic hypothesis connecting cohesin dysfunction and immune dysregulation. This work expands the mutational spectrum of both genes and opens a new avenue for understanding developmental-inflammatory disease overlap. Full article

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

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14 pages, 4882 KB

Open AccessArticle

Allogeneic Umbilical Cord Blood Serum Eyedrops for the Treatment of Severe Dry Eye Disease Patients

by Marco Zeppieri, Giuseppe Gagliano, Matteo Capobianco, Caterina Gagliano, Francesco Cappellani, Giuseppa Tancredi, Alessandro Avitabile, Ludovica Cannizzaro and Fabiana D’Esposito

Int. J. Mol. Sci. 2025, 26(21), 10782; https://doi.org/10.3390/ijms262110782 - 6 Nov 2025

Viewed by 508

Abstract

Human allogeneic umbilical cord blood serum stands out as a potent adjunct to conventional therapies for ocular surface disorders related to severe Dry Eye Disease. By expediting ocular surface regeneration and fostering epithelial integrity, umbilical cord blood serum not only enhances subjective patient [...] Read more.

Human allogeneic umbilical cord blood serum stands out as a potent adjunct to conventional therapies for ocular surface disorders related to severe Dry Eye Disease. By expediting ocular surface regeneration and fostering epithelial integrity, umbilical cord blood serum not only enhances subjective patient experiences but also improves objective clinical indicators. This makes it particularly useful in patients with corneal ulcers through ocular surface regeneration and anti-inflammatory activity. This retrospective, interventional, non-randomized clinical study aims to explore the efficacy of allogenic umbilical cord blood serum in patients who had previously received other treatments unsuccessfully. This study was a retrospective, non-comparative, interventional clinical study involving 55 patients (35 females and 20 males) aged 18–82 years with severe Dry Eye Disease who were unresponsive to standard treatments. The study was conducted at Eye Center “G.B. Morgagni-DSV”, Catania, Italy. Patients were categorized based on the etiology of severe Dry Eye Disease into four groups: group I consisted of 26 patients with filamentary keratitis and corneal ulcers associated with rheumatologic diseases such as Sjogren’s syndrome and systemic sclerosis; group II comprised 15 patients with graft-versus-host disease; group III consisted of 10 patients with corneal neurotrophic ulcers; group IV included four patients with Steven–Johnson syndrome. Outcomes evaluated before and after treatment were OSDI (Ocular Surface Disease Index) and SANDE (Symptom Assessment in Dry Eye) Questionnaires, VAS (Visual Analog Scale), Slit-Lamp Examination, Esthesiometry, Lissamine Green Staining, NIBUT (Non-Invasive Break-Up Time) and BUT, Fluorescein Staining with Photography and Oxford Classification, Schirmer Test, Best-Corrected Visual Acuity (BCVA), Meibography. We observed a significant improvement in SANDE, VAS and OSDI questionnaires, Schirmer Test, BUT, BCVA, and Oxford classification after treatment with allogeneic cord blood serum eyedrops. Clinical variables, such as corneal inflammation, conjunctivalization, corneal neovascularization, or pain, were also considered individually. Nevertheless, pain and inflammation reduced markedly over time until completely healed in all cases. Our study highlights the remarkable efficacy of allogeneic cord blood serum eyedrops in patients with severe Dry Eye Disease who have shown absent or inadequate response to usual treatments for dry eye. This underscores the need for further comprehensive investigations in this field. Full article

(This article belongs to the Special Issue Integrated Approaches, Molecular Mechanism and Therapies in Ocular Surface Diseases)

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17 pages, 1308 KB

Open AccessCommunication

Anti-Pneumococcal Properties of the Native Human Milk Oligosaccharide Fraction: A Concentration-Dependent Study

by Oliwia Makarewicz, Tinatini Tchatchiashvili, Lisa Jasef, Mark P. G. van der Linden, Sylwia Jarzynka, Kamila Strom, Nico Ueberschaar, Maciej Mazur, Gabriela Oledzka and Mathias W. Pletz

Int. J. Mol. Sci. 2025, 26(21), 10781; https://doi.org/10.3390/ijms262110781 - 6 Nov 2025

Viewed by 412

Abstract

Streptococcus pneumoniae is a major opportunistic pathogen and a leading cause of severe infections in infants under two years of age. Human milk oligosaccharides (HMOs), key bioactive components of breast milk, possess immunomodulatory and antimicrobial properties. In this study, the antipneumococcal effects of [...] Read more.

Streptococcus pneumoniae is a major opportunistic pathogen and a leading cause of severe infections in infants under two years of age. Human milk oligosaccharides (HMOs), key bioactive components of breast milk, possess immunomodulatory and antimicrobial properties. In this study, the antipneumococcal effects of HMOs are investigated across multiple S. pneumoniae serotypes, focusing on concentration-dependent activity and underlying mechanisms. Growth inhibition and bacterial viability were evaluated using growth curve analysis and colony-forming unit (CFU) assays. HMOs inhibited pneumococcal growth in a concentration-dependent manner, with suppression observed at 1.5–2.5 mg/mL and complete killing at 5 mg/mL for all serotypes. Nonencapsulated strains were more sensitive, with inhibition at 1 mg/mL. In the CFU assays, killing occurred at 1.25–5 mg/mL depending on the strain. At physiologically relevant colostrum concentrations (20–25 mg/mL), HMOs achieved complete bactericidal effects across all the tested strains. In contrast, lactose at equivalent doses showed no measurable antimicrobial activity, confirming the specificity of the observed effects. Overall, HMOs exhibit serotype-independent antipneumococcal activity, possibly through interference with bacterial adhesion or metabolic disruption. These findings suggest a potential role for HMOs as adjunctive agents in the prevention of pneumococcal infections in vulnerable populations, such as infants, and warrant further in vivo studies to validate these effects and explore clinical applications. Full article

(This article belongs to the Special Issue Therapeutic Horizons of Oligosaccharides: Molecular Mechanisms and Therapeutic Potential)

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32 pages, 3100 KB

Open AccessArticle

Network Controllability Reveals Key Mitigation Points for Tumor-Promoting Signaling in Tumor-Educated Platelets

by Özge Osmanoglu, Elif Özer, Shishir K. Gupta, Katrin G. Heinze, Harald Schulze and Thomas Dandekar

Int. J. Mol. Sci. 2025, 26(21), 10780; https://doi.org/10.3390/ijms262110780 - 5 Nov 2025

Viewed by 983

Abstract

Therapeutic strategies targeting “tumor-educated platelets” (TEPs) and platelet–tumor interactions by key signaling pathways (ITAM, P2Y12) may reduce metastasis and cancer. Using a TEP gene expression dataset originally created to study swarm intelligence-enhanced detection of lung cancer cells (GSE89843), we did perform extensive transcriptome [...] Read more.

Therapeutic strategies targeting “tumor-educated platelets” (TEPs) and platelet–tumor interactions by key signaling pathways (ITAM, P2Y12) may reduce metastasis and cancer. Using a TEP gene expression dataset originally created to study swarm intelligence-enhanced detection of lung cancer cells (GSE89843), we did perform extensive transcriptome analysis to integrate these data with directed protein–protein interactions and build a TEP-specific signaling network. We analyze network topology and controllability and identify critical and indispensable nodes, as well as high-weight, usually high-score nodes. We reconstruct (pharmacological) controllable subnetworks of TEP signaling, which we then explore for drugs targets. We found 111 upregulated and 108 downregulated genes compared to control platelets, enriched in pathways related to extracellular matrix interactions, cytoskeleton organization, immune signaling, and platelet activation. Ribosomal function, apoptosis, and immune signaling were among the downregulated processes, highlighting unique TEP profiles in non-small-cell lung cancer (NSCLC). Our integrative analysis of TEPs in NSCLC reveals key transcriptional and network-based alterations harmful for the cancer patient. Using four complementary strategies, we identified five high-confidence genes (Gene symbols always given throughout the paper), ITGA2B, FLNA, GRB2, FCGR2A, and APP, as central to TEP signaling. These can be targeted by FDA-approved drugs. Fostamatinib, an SYK inhibitor, emerged as the top candidate drug to disrupt ITAM-mediated platelet activation selectively; metastasis-promoting metalloprotease and cytoskeletal targets influencing adhesion were also identified. A low-dose combination therapy of fostamatinib, Aducanumab, and acetylsalicylic acid (aspirin) may control TEP effects. In conclusion, our preclinical in silico approach revealed FDA-approved drugs that allow therapeutic targeting of metastasis-promoting TEPs and target NSCLC at the same time. Full article

(This article belongs to the Special Issue Cancer-Driving Molecules: From Molecular Mechanisms to Novel Therapeutics)

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