International Journal of Molecular Sciences

22 pages, 939 KiB

Open AccessReview

Bacteriophages, Antibiotics and Probiotics: Exploring the Microbial Battlefield of Colorectal Cancer

by Cristian Constantin Volovat, Mihai Andrei Cosovanu, Madalina-Raluca Ostafe, Iolanda Georgiana Augustin, Constantin Volovat, Bogdan Georgescu and Simona Ruxandra Volovat

Int. J. Mol. Sci. 2025, 26(16), 7837; https://doi.org/10.3390/ijms26167837 - 13 Aug 2025

Abstract

Colorectal cancer (CRC), a prevalent malignancy, is a significant global health concern. The intricate interplay of genetic mutations, inflammatory processes, and environmental factors underscores the complexity of CRC’s etiology. The human gut harbors a diverse microbial community that plays a key role in [...] Read more.

Colorectal cancer (CRC), a prevalent malignancy, is a significant global health concern. The intricate interplay of genetic mutations, inflammatory processes, and environmental factors underscores the complexity of CRC’s etiology. The human gut harbors a diverse microbial community that plays a key role in maintaining homeostasis and influencing various aspects of host physiology. Perturbations in the gut microbiome (GM) composition and function have been implicated in CRC carcinogenesis. This bidirectional relationship involves microbial contributions to inflammation, DNA damage, and immune modulation, shaping the tumor microenvironment (TME). Bacteriophages, viruses that infect bacteria, contribute to the microbiome’s diversity and function by influencing bacterial abundance and composition. These phages can impact host–microbiome interactions, potentially influencing CRC risk. Furthermore, they can be manipulated to transport targeted medication, without being metabolized. Antibiotics exert selective pressures on the gut microbiome, leading to shifts in bacterial populations and potential dysbiosis. Probiotics can modulate the composition and activity of the GM and could be considered adjunctive therapy in the treatment of CRC. Understanding the intricate balance between bacteriophages, antibiotics–probiotics, and the GM is essential for comprehending CRC etiology and progression. Full article

(This article belongs to the Special Issue Colorectal Cancer: From Pathophysiology to Novel Therapies)

25 pages, 4622 KiB

Open AccessReview

Immunological Landscape and Molecular Therapeutic Targets of the Tumor Microenvironment in Hepatocellular Carcinoma

by Yusra Zarlashat, Abdul Ghaffar, Flora Guerra and Anna Picca

Int. J. Mol. Sci. 2025, 26(16), 7836; https://doi.org/10.3390/ijms26167836 - 13 Aug 2025

Abstract

Hepatocellular carcinoma (HCC) is the most common liver cancer, with poor survival rates in advanced stages due to late diagnosis, tumor heterogeneity, and therapy resistance. The tumor microenvironment (TME) in HCC has a crucial role in tumor progression, characterized by a complex interaction [...] Read more.

Hepatocellular carcinoma (HCC) is the most common liver cancer, with poor survival rates in advanced stages due to late diagnosis, tumor heterogeneity, and therapy resistance. The tumor microenvironment (TME) in HCC has a crucial role in tumor progression, characterized by a complex interaction of immune cells, stromal components, and immunosuppressive signaling pathways. Chronic inflammation driven by viral infections, metabolic dysfunction, and alcohol consumption triggers an immunosuppressive TME, promoting immune evasion and tumor growth. Immune cell populations, such as myeloid-derived suppressor cells, regulatory T cells, and tumor-associated macrophages, contribute to immunosuppression, while cytotoxic T lymphocytes and natural killer cells exert anti-tumor effects. Recent advances in immunotherapy, mainly immune checkpoint inhibitors (ICIs) targeting programmed death-ligand 1 and programmed cell death protein 1 and cytotoxic T-lymphocyte-associated protein 4, have revolutionized HCC treatment, though response rates remain limited. Combined therapies using tyrosine kinase inhibitors, anti-angiogenic agents, and ICIs improve patient outcomes. This review discusses the immunological mechanisms contributing to HCC progression, the role of immune cell subsets in tumor evasion, and therapeutic interventions, from conventional treatments to advanced immunotherapies. Ongoing clinical trials, barriers to effective treatment, and future directions to enhance HCC management and patient survival will also be overviewed. Full article

(This article belongs to the Special Issue Hepatopancreatic and Intestine Diseases: From Molecular Basis to Therapy)

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22 pages, 5708 KiB

Open AccessArticle

Genome-Wide Identification and Expression Analysis of the WRKY Gene Families in Vaccinium bracteatum

by Haijing Du, Jianqiang Zhou, Xiaoran Liang, Yufei Chen, Xiaohui Liu, Cheng Zhen, Hong Zhang, Jiaxin Xiao and Xuan Gao

Int. J. Mol. Sci. 2025, 26(16), 7835; https://doi.org/10.3390/ijms26167835 - 13 Aug 2025

Abstract

The WRKY gene family is a widely distributed and highly conserved transcription factor (TF) family in plants, with its members playing key roles in plant growth and development, stress response, and metabolism. Although WRKY TFs have been extensively studied in many plant species, [...] Read more.

The WRKY gene family is a widely distributed and highly conserved transcription factor (TF) family in plants, with its members playing key roles in plant growth and development, stress response, and metabolism. Although WRKY TFs have been extensively studied in many plant species, research on the WRKY gene family in Vaccinium bracteatum Thunb. remains limited. Therefore, integrating molecular biology and bioinformatics approaches to further explore the WRKY gene family in V. bracteatum is of considerable scientific importance. In this study, we employed various online tools to obtain genomic and expression data, which were subsequently analyzed to determine the composition, evolutionary relationships, and functions of WRKY family genes in V. bracteatum. A total of 66 WRKY genes (VaWRKY) were identified, named based on homology alignment. Phylogenetic analysis classified the 66 VaWRKYs into three major clades and seven subclades. Sequence and structural analyses of VaWRKY genes provided insights into their evolutionary and functional characteristics. Expression profile analysis revealed significant differences in the expression of 12 VaWRKY genes at various stages of fruit development. Protein interaction analysis further indicated that VaWRKY genes are functionally diverse, playing important roles in stress response, seed germination regulation, and plant growth and development. In summary, we have a deeper understanding of VaWRKY genes, and systematic analysis of structure, evolutionary characteristics, and expression patterns plays an important role in analyzing its biological functions, molecular breeding, and enhancing economic value. Full article

(This article belongs to the Section Molecular Plant Sciences)

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21 pages, 4239 KiB

Open AccessArticle

Melatonin-Producing Bacillus aerius EH2-5 Enhances Glycine max Plants Salinity Tolerance Through Physiological, Biochemical, and Molecular Modulation

by Eun-Hae Kwon, Suhaib Ahmad and In-Jung Lee

Int. J. Mol. Sci. 2025, 26(16), 7834; https://doi.org/10.3390/ijms26167834 - 13 Aug 2025

Abstract

Climate change has intensified extreme weather events and accelerated soil salinization, posing serious threats to crop yield and quality. Salinity stress, now affecting about 20% of irrigated lands, is expected to worsen due to rising temperatures and sea levels. At the same time, [...] Read more.

Climate change has intensified extreme weather events and accelerated soil salinization, posing serious threats to crop yield and quality. Salinity stress, now affecting about 20% of irrigated lands, is expected to worsen due to rising temperatures and sea levels. At the same time, the global population is projected to exceed 9 billion by 2050, demanding a 70% increase in food production (UN, 2019; FAO). Agriculture, responsible for 34% of global greenhouse gas emissions, urgently needs sustainable solutions. Microbial inoculants, known as “plant probiotics,” offer a promising eco-friendly alternative by enhancing crop resilience and reducing environmental impact. In this study, we evaluated the plant growth-promoting (PGP) traits and melatonin-producing capacity of Bacillus aerius EH2-5. To assess its efficacy under salt stress, soybean seedlings at the VC stage were inoculated with EH2-5 and subsequently subjected to salinity stress using 150 mM and 100 mM NaCl treatments. Plant growth parameters, the expression levels of salinity-related genes, and the activities of antioxidant enzymes were measured to determine the microbe’s role in promoting plant growth and mitigating salt-induced oxidative stress. Here, our study shows that the melatonin-synthesizing Bacillus aerius EH2-5 (7.48 ng/mL at 24 h after inoculation in Trp spiked LB media) significantly improved host plant (Glycine max L.) growth, biomass, and photosynthesis and reduced oxidative stress during salinity stress conditions than the non-inculcated control. Whole genome sequencing of Bacillus aerius EH2-5 identified key plant growth-promoting and salinity stress-related genes, including znuA, znuB, znuC, and zur (zinc uptake); ptsN, aspA, and nrgB (nitrogen metabolism); and phoH and pstS (phosphate transport). Genes involved in tryptophan biosynthesis and transport, such as trpA, trpB, trpP, and tspO, along with siderophore-related genes yusV, yfhA, and yfiY, were also detected. The presence of multiple stress-responsive genes, including dnaK, dps, treA, cspB, srkA, and copZ, suggests EH2-5′s genomic potential to enhance plant tolerance to salinity and other abiotic stresses. Inoculation with Bacillus aerius EH2-5 significantly enhanced soybean growth and reduced salt-induced damage, as evidenced by increased shoot biomass (29%, 41%), leaf numbers (12% and 13%), and chlorophyll content (40%, 21%) under 100 mM and 150 mM NaCl compared to non-inoculated plants. These results indicate EH2-5′s strong potential as a plant growth-promoting and salinity stress-alleviating rhizobacterium. The EH2-5 symbiosis significantly enhanced a key ABA biosynthesis enzyme-related gene NCED3, dehydration responsive transcription factors DREB2A and NAC29 salinity stresses (100 mM and 150 mM). Moreover, the reduced expression of peroxidase (POD), superoxide dismutase (SOD), and catalase (CAT) by 16%, 29%, and 24%, respectively, and decreased levels of malondialdehyde (MDA) and hydroxy peroxidase (H₂O₂) by 12% and 23% were observed under 100 mM NaCl compared to non-inoculated plants. This study demonstrated that Bacillus aerius EH2-5, a melatonin-producing strain, not only functions effectively as a biofertilizer but also alleviates plant stress in a manner comparable to the application of exogenous melatonin. These findings highlight the potential of utilizing melatonin-producing microbes as a viable alternative to chemical treatments. Therefore, further research should focus on enhancing the melatonin biosynthetic capacity of EH2-5, improving its colonization efficiency in plants, and developing synergistic microbial consortia (SynComs) with melatonin-producing capabilities. Such efforts will contribute to the development and field application of EH2-5 as a promising plant biostimulant for sustainable agriculture. Full article

(This article belongs to the Special Issue Genetics and Novel Techniques for Soybean Pivotal Characters)

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31 pages, 468 KiB

Open AccessArticle

Impact of Soil Drought on Yield and Leaf Sugar Content in Wheat: Genotypic and Phenotypic Relationships Compared Using a Doubled Haploid Population

by Magdalena Grela, Steve Quarrie, Katarzyna Cyganek, Jan Bocianowski, Małgorzata Karbarz, Mirosław Tyrka, Dimah Habash, Michał Dziurka, Edyta Kowalczyk, Wojciech Szarski and Ilona Mieczysława Czyczyło-Mysza

Int. J. Mol. Sci. 2025, 26(16), 7833; https://doi.org/10.3390/ijms26167833 - 13 Aug 2025

Abstract

Improving yield stability under water-limited conditions is a key objective of wheat breeding programmes. One trait of particular interest is carbohydrate accumulation and remobilisation. This study assessed the genetic basis of aspects of yield and flag leaf sugar contents under drought and well-watered [...] Read more.

Improving yield stability under water-limited conditions is a key objective of wheat breeding programmes. One trait of particular interest is carbohydrate accumulation and remobilisation. This study assessed the genetic basis of aspects of yield and flag leaf sugar contents under drought and well-watered conditions using QTL mapping in a population of 90 doubled haploid lines derived from the cross Chinese Spring × SQ1. As well as soluble sugar content, glucose, fructose, sucrose, and maltose, the traits grain yield (Yld), biomass (Bio), and thousand grain weight (TGW) were also analysed. Analysis of variance showed that genotype, environment and their interactions significantly influenced all the traits studied, with environmental effects explaining up to 74.4% of the total variation. QTL analysis identified 40 QTLs for Yld, TGW, and Bio as well as 53 QTLs for soluble carbohydrates, accounting for up to 40% of phenotypic variation. QTLs coincident for more than one trait were identified on 21 chromosome regions, associated with carbohydrate metabolism and yield performance under drought, particularly on chromosomes 2D, 4A, 4B, 5B, 5D, 6B, and 7A. Candidate genes for several yield-related QTLs were identified. These results provide useful genetic markers for the development of more drought-resistant wheat cultivars. Full article

(This article belongs to the Special Issue Molecular Mechanisms of Plant Abiotic Stress Tolerance: 2nd Edition)

27 pages, 6274 KiB

Open AccessArticle

Synergistic Antimicrobial Activity of BrSPR20-P1 Peptide and Silver Nanoparticles Against Pathogenic Bacteria

by Thanyamai Thongin, Somchai Sawatdee, Nuttapon Songnaka, Jumpei Uchiyama, Theanchai Wiwasuku, Teerapol Srichana, Titpawan Nakpheng and Apichart Atipairin

Int. J. Mol. Sci. 2025, 26(16), 7832; https://doi.org/10.3390/ijms26167832 - 13 Aug 2025

Abstract

Bacterial infection is a cause of life-threatening diseases. The emergence of antimicrobial-resistant bacteria exacerbates this situation, highlighting the need for the discovery of new antimicrobial agents. Our previous study identified a novel antimicrobial peptide, BrSPR20-P1 (P1), which showed potential activity against MRSA. Additionally, [...] Read more.

Bacterial infection is a cause of life-threatening diseases. The emergence of antimicrobial-resistant bacteria exacerbates this situation, highlighting the need for the discovery of new antimicrobial agents. Our previous study identified a novel antimicrobial peptide, BrSPR20-P1 (P1), which showed potential activity against MRSA. Additionally, silver nanoparticles (AgNPs) exhibit broad-spectrum antibacterial activity, capable of killing multidrug-resistant bacteria. The combination of antimicrobial agents presents a novel strategy for combating these pathogens. This study aimed to evaluate the antibacterial activity of the combination of P1 and AgNPs. It revealed that the combinations showed synergy. The P1 and AgNP mixture at a concentration of 1 and 8 µg/mL (1:8) doubled the activity against S. aureus and MRSA, while that combination of 64 and 64 µg/mL (64:64) exhibited broad-spectrum activity, expanding to E. coli with a 32-fold increase. These combinations exhibited a bactericidal effect, showing the rapid killing of tested bacteria at 10× MIC, with killing rates during the first 3 h ranging from 4.04 ± 0.01 to 4.31 ± 0.03 h⁻¹. The P1 and AgNP mixtures caused a low risk of antibacterial resistance up to 30 passages. It was demonstrated that the synergistic activity of P1 and AgNPs occurred through the disruption of cell walls and membranes, leakage of intracellular materials, and cell lysis. Additionally, the mixtures appeared to interact with bacterial genomic DNA, as indicated by a gel retardation assay. These activities of the combinations were concentration-dependent. The 1:8 µg/mL mixture caused low hemolysis and cytotoxicity and did not impede the wound healing process. In contrast, although the 64:64 µg/mL mixture showed excellent antibacterial efficacy, it was toxic to erythrocytes and mammalian cells. It implies that dose optimization is required to balance its efficacy and toxicity. Therefore, the P1 and AgNP combinations exhibit synergistic antimicrobial activity and have the potential to resolve bacterial infections. Full article

(This article belongs to the Special Issue Antimicrobial Nanomaterials: Approaches, Strategies and Applications)

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7 pages, 188 KiB

Open AccessEditorial

Special Issue “New Insights into Adipose Tissue Metabolic Function and Dysfunction, 3rd Edition”

by Federica Mannino

Int. J. Mol. Sci. 2025, 26(16), 7831; https://doi.org/10.3390/ijms26167831 - 13 Aug 2025

Abstract

The understanding of adipose tissue has evolved from viewing it as a passive storage depot for excess energy to recognizing it as a central endocrine organ, crucial in regulating metabolic homeostasis, immune responses, and inter-organ communication [...] Full article

(This article belongs to the Special Issue New Insights into Adipose Tissue Metabolic Function and Dysfunction, 3rd Edition)

25 pages, 834 KiB

Open AccessArticle

The Roles of PD-L1, Ki-67, P53, and Cyclin D1 in PitNETs: Diagnostic and Prognostic Implications in a Series of 74 Patients

by Anna Krzentowska, Beata Biesaga, Ryszard Czepko, Anna Merklinger-Gruchała, Dariusz Adamek, Małgorzata Jasińska, Barbara Pluta, Wiktoria Michalska, Katarzyna Wróblewska, Filip Janczy and Filip Gołkowski

Int. J. Mol. Sci. 2025, 26(16), 7830; https://doi.org/10.3390/ijms26167830 - 13 Aug 2025

Abstract

Pituitary neuroendocrine tumors (PitNETs), also known as pituitary adenomas, are rare tumors that are usually benign. At present, the WHO PitNET classification based on transcription factors is in force. A problem is caused by invasive tumors and silent tumors which, despite a lack [...] Read more.

Pituitary neuroendocrine tumors (PitNETs), also known as pituitary adenomas, are rare tumors that are usually benign. At present, the WHO PitNET classification based on transcription factors is in force. A problem is caused by invasive tumors and silent tumors which, despite a lack of obvious clinical symptoms, tend to behave aggressively. Factors influencing the clinical course of these tumors are currently being sought. The aim of our study was to assess the expression of programmed death-ligand 1 (PD-L1) and proliferation biomarkers (Ki-67, cyclin D1, and P53) in PitNETs depending on the transcription factor and adenoma subtype. The analysis was performed in seventy-four patients operated on in a single neurosurgical center for pituitary tumors. Immunohistochemistry was performed for transcription factors and biomarkers—PD-L1, Ki-67, P53, and cyclin D1—in tissue microarray format. Membranous expression of PD-L1 was scored as 0 (no expression) and ≥1%. Nuclear expression of Ki-67 was scored at <3% and ≥3%, and the expression of P53 and cyclin D1 was scored at <10% and ≥10%. The following tumors expressed PD-L1 at ≥1%: gonadotroph, 21 (28.4%); corticotroph, 5 (6.7%); gonadotroph/lactotroph, 2 (2.7%); null cell adenoma, 3 (4.0%); multiple synchronous PitNET, 2 (2.7%); immature PIT-1 tumor, 1 (1.3%); mature PIT-1 tumor, 1 (1.5%). KI ≥ 3% was found in the following PitNETs: gonadotroph, 3 (4.0%); corticotroph, 2 (2.7%); lactotroph, 1 (1.3%); multiple synchronous PitNET, 1 (1.3%); immature PIT-1 tumor, 1 (1.3%); and mature PIT-1 tumor, 1 (1.3%). Patients with K ≥ 3% were statistically significantly younger (p = 0.03). All tumors (100%) with a combination of cyclin D1 ≥ 10% and P53 < 10% were invasive on the Hardy scale. Of the four factors, PD-L1 increased the odds of invasiveness the most (adjusted OR = 2.35; 95% CI: 0.56–9.90). PD-L1 expression was present in some types of PitNETs. PD-L1 expression may help in identifying null cell adenomas. High cyclin D1 with low P53 may indicate greater tumor invasiveness. Full article

(This article belongs to the Special Issue Biomarkers in Cancer Immunology)

14 pages, 1611 KiB

Open AccessReview

TRP Channels in Skin Cancer: Focus on Malignant Melanoma

by Damian Twardak, Vita Havryliuk and Maciej Gagat

Int. J. Mol. Sci. 2025, 26(16), 7829; https://doi.org/10.3390/ijms26167829 - 13 Aug 2025

Abstract

Cutaneous malignant melanoma remains one of the most aggressive forms of skin cancer, characterized by high metastatic potential and resistance to standard therapies. Emerging evidence suggests that transient receptor potential (TRP) channels, non-selective cation channels involved in calcium homeostasis, and cellular stress responses [...] Read more.

Cutaneous malignant melanoma remains one of the most aggressive forms of skin cancer, characterized by high metastatic potential and resistance to standard therapies. Emerging evidence suggests that transient receptor potential (TRP) channels, non-selective cation channels involved in calcium homeostasis, and cellular stress responses play a pivotal role in melanoma development and progression. This review highlights the physiological expression of key TRP subfamilies (TRPM1, TRPM7, TRPM8, TRPV1, TRPV4, and TRPM2) in melanocytes and discusses their dysregulation in melanoma cells. TRPM1 is implicated as a tumor suppressor, whereas TRPM7, TRPV1, and TRPV4 often function as both melanoma suppressor or oncogenic drivers, modulating proliferation, apoptosis, and metastasis. TRPM2, which is responsive to oxidative stress, supports melanoma cell survival under metabolic stress. The potential of TRP channels as diagnostic biomarkers and therapeutic targets is evaluated, with attention paid to current pharmacological approaches and research challenges. The complexity and context-dependency of TRP function in melanoma underscore the need for isoform-specific modulation and personalized therapeutic strategies. Full article

(This article belongs to the Section Molecular Oncology)

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16 pages, 1118 KiB

Open AccessReview

The Role of Receptor Tyrosine Kinase-like Orphan Receptor 1 (ROR1) in Cancer Stem Cell Signaling

by Matthew S. Jung, Won-Young Choi, Wenjing Zhang, Francisco N. Barrera and Rachel S. Perkins

Int. J. Mol. Sci. 2025, 26(16), 7828; https://doi.org/10.3390/ijms26167828 - 13 Aug 2025

Abstract

Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a key regulator of cancer stem cell (CSC) biology and signaling. In CSCs, ROR1 acts as a receptor or co-receptor, interacting with non-canonical WNT ligands, and forming complexes with proteins like CD19 and HER2, to [...] Read more.

Receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a key regulator of cancer stem cell (CSC) biology and signaling. In CSCs, ROR1 acts as a receptor or co-receptor, interacting with non-canonical WNT ligands, and forming complexes with proteins like CD19 and HER2, to activate diverse downstream signaling pathways. ROR1 signaling in CSCs promotes proliferation, maintains stemness, and enhances migration, invasion, and the epithelial-to-mesenchymal transition (EMT). While minimally expressed after embryogenesis, ROR1 is aberrantly upregulated in numerous cancers, including ovarian, breast, pancreatic, and hematologic malignancies. ROR1 overexpression drives tumor progression, resistance to chemotherapies, disease recurrence, and ultimately metastasis. This expression pattern positions ROR1 as a promising target for CSC-specific therapies. High ROR1 expression is consistently linked to aggressive disease and poor patient outcomes. Here, we review ROR1′s role in CSCs and highlight the complex signaling that is observed in the CSC population. Further, we evaluate the gaps in the current understanding of ROR1 signaling in CSCs and describe how ROR1 regulates the associated signaling pathways. Finally, we provide an up-to-date summary of the promising therapeutic strategies targeting ROR1 that overcome conventional cancer treatment limitations. This review highlights the role of ROR1 as a critical, functional driver of CSCs and adverse patient outcomes across various malignancies. Full article

(This article belongs to the Special Issue New Advances in Cancer Stem Cell Research: 2nd Edition)

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15 pages, 1580 KiB

Open AccessArticle

Syringin (Sinapyl Alcohol 4-O-Glucoside) Improves the Wound Healing Capacity of Fibroblasts and Keratinocytes In Vitro

by Andrzej Parzonko, Agnieszka Filipek, Marcin Równicki and Anna K. Kiss

Int. J. Mol. Sci. 2025, 26(16), 7827; https://doi.org/10.3390/ijms26167827 - 13 Aug 2025

Abstract

Wound healing is a complex process in which TGFβ plays a key role. Previous studies have shown that syringin, a phenylpropanoid glycoside present in lilac bark (Syringa vulgaris L.), stimulates TGFβ expression in human monocyte-derived macrophages in addition to inhibiting the secretion [...] Read more.

Wound healing is a complex process in which TGFβ plays a key role. Previous studies have shown that syringin, a phenylpropanoid glycoside present in lilac bark (Syringa vulgaris L.), stimulates TGFβ expression in human monocyte-derived macrophages in addition to inhibiting the secretion of pro-inflammatory cytokines. Here, we investigated the effect of syringin on migration, invasion, and TGFβ production, as well as the effect on the release of pro-inflammatory cytokines in human dermal fibroblasts (NHDF) and keratinocytes (HaCaT) and its mechanism of action. NHDF and HaCaT cells were treated with the tested compound (12.5–100 µM), and a scratch assay was performed. The effect of migration using modified Boyden chambers was analyzed. TGFβ and IL-6 release were also assessed using ELISA kits. Cell proliferation was assessed using MTT and BrdU incorporation tests, while cytotoxicity was assessed using a neutral red uptake test. Smad2 and Smad3 phosphorylation were assessed using Western Blotting. ACTA2, COL1A1, and TIMP3 expression was analyzed using qPCR. Cells treated with syringin showed an increase in invasion potential in the scratch assay. A significant increase in skin fibroblast migration through the porous membrane was also observed. Syringin increased TGFβ release and inhibited IL-6 release by NHDF and HaCaT cells. No effect of syringin on cell proliferation or cytotoxic effects was observed. Western blot analysis showed significant activation of Smad2 and Smad3 in the presence of syringin in NHDF cells, but not in HaCaT. Quantitative PCR analysis revealed a strong increase in ACTA2 and COL1A1 gene expression in fibroblast cells treated with syringin. The present study demonstrated that syringin present in S. vulgaris stem bark increased dermal fibroblasts and keratinocytes’ wound healing function through activation of cell migration. Full article

(This article belongs to the Section Bioactives and Nutraceuticals)

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17 pages, 6704 KiB

Open AccessArticle

Effects of a Novel Mammalian-Derived Collagen Matrix on Human Articular Cartilage-Derived Chondrocytes from Osteoarthritis Patients

by Mingyuan Wang, Toru Iwahashi, Taisuke Kasuya, Mai Konishi, Katsuyuki Konishi, Miki Kawanaka, Takashi Kanamoto, Hiroyuki Tanaka and Ken Nakata

Int. J. Mol. Sci. 2025, 26(16), 7826; https://doi.org/10.3390/ijms26167826 - 13 Aug 2025

Abstract

Osteoarthritis (OA) is the most common joint disorder worldwide. Autologous chondrocyte implantation (ACI) is an established treatment for articular cartilage defects of the knee, but its effectiveness in OA is still under investigation. In this study, we investigated the effects of a newly [...] Read more.

Osteoarthritis (OA) is the most common joint disorder worldwide. Autologous chondrocyte implantation (ACI) is an established treatment for articular cartilage defects of the knee, but its effectiveness in OA is still under investigation. In this study, we investigated the effects of a newly developed mammalian-derived collagen matrix, NC-Col, on the proliferation, migration, adhesion, and gene expression of human articular cartilage-derived chondrocytes from OA patients in vitro, using proliferation assays, wound healing assays, adhesion assays, RT-qPCR, and RNA sequencing, respectively. In addition, the effects of NC-Col were compared with three different commercially available collagen matrices, and the underlying molecular mechanisms through which NC-Col influences these cellular behaviours were explored. Our results showed that NC-Col, used as a coating matrix, enhances cell proliferation, maintains the phenotype, and upregulates Proteoglycan 4 (PRG4) in human articular cartilage-derived chondrocytes. Inhibition of the PI3K-Akt signalling pathway was found to be involved in some of these effects. In conclusion, our findings suggest that NC-Col collagen may offer new strategies for improving therapeutic outcomes in OA, particularly in the context of ACI. Full article

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

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22 pages, 757 KiB

Open AccessReview

Carbon Monoxide as a Molecular Modulator of Ischemia–Reperfusion Injury: New Insights for Translational Application in Organ Transplantation

by Zhouyu Li, Kazuhiro Takeuchi, Yuichi Ariyoshi, Akira Kondo, Takehiro Iwanaga, Yurika Ichinari, Akiyuki Iwamoto, Kenya Shimizu, Kohei Miura, Shiori Miura, Lina Ma, Mitsuhiro Sekijima, Masayoshi Okumi and Hisashi Sahara

Int. J. Mol. Sci. 2025, 26(16), 7825; https://doi.org/10.3390/ijms26167825 - 13 Aug 2025

Abstract

Carbon monoxide (CO) is generally recognized as a toxic gas; however, it has recently been identified as an endogenous gasotransmitter with significant cytoprotective properties. CO modulates key molecular pathways, including anti-inflammatory, anti-apoptotic, antioxidant, and vasodilatory signaling pathways, by targeting heme- and non-heme-containing proteins. [...] Read more.

Carbon monoxide (CO) is generally recognized as a toxic gas; however, it has recently been identified as an endogenous gasotransmitter with significant cytoprotective properties. CO modulates key molecular pathways, including anti-inflammatory, anti-apoptotic, antioxidant, and vasodilatory signaling pathways, by targeting heme- and non-heme-containing proteins. These proteins include soluble guanylate cyclase, cytochrome P450 enzymes, MAPKs, and NLRP3. This review summarizes recent advances in understanding the molecular mechanisms associated with the protective effects of CO, particularly in the context of ischemia–reperfusion injury relevant to organ transplantation. We discuss preclinical data from rodent and large animal models, as well as therapeutic delivery strategies, such as inhalation, CO-releasing molecules, and gas-entrapping materials. We also reviewed early-phase clinical trials. The objective of this review is to provide a thorough exploration of CO as a potential therapeutic gas, with special emphasis on its application in transplantation. Full article

(This article belongs to the Special Issue New Molecular Insights into Ischemia/Reperfusion: 2nd Edition)

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23 pages, 11380 KiB

Open AccessArticle

Integrated Analysis of Physiological Responses and Transcriptome of Cotton Seedlings Under Drought Stress

by Xin Li, Yuhao Zhao, Chen Gao, Xiaoya Li, Kunkun Wu, Meiwei Lin and Weihong Sun

Int. J. Mol. Sci. 2025, 26(16), 7824; https://doi.org/10.3390/ijms26167824 - 13 Aug 2025

Abstract

Investigating the physiological responses and resistance mechanisms in plants under drought stress provides critical insights for optimizing irrigation water utilization efficiency and promoting the development of irrigation science. In this study, cotton seedlings were cultivated in a light incubator. Three drought stress levels [...] Read more.

Investigating the physiological responses and resistance mechanisms in plants under drought stress provides critical insights for optimizing irrigation water utilization efficiency and promoting the development of irrigation science. In this study, cotton seedlings were cultivated in a light incubator. Three drought stress levels were applied: mild (M1, 50–55% field moisture), moderate (M2, 45–50%), and severe (M3, 40–45%). Transcriptome analysis was performed under mild and severe stress. The results revealed that differentially expressed genes (DEGs) related to proline degradation were down-regulated and proline content increased in cotton. Under different stress treatments, cotton exhibited a stress-intensity-dependent regulation of carbohydrate metabolism and soluble sugar content decreased and then increased. And the malondialdehyde content analysis revealed a dose-dependent relationship between stress intensity and membrane lipid peroxidation. Stress activated the antioxidant system, leading to the down-regulation of DEGs for reactive oxygen species production in the mitogen-activated protein kinase (MAPK) signaling pathway. Concurrently, superoxide dismutase activity and peroxidase content increased to mitigate oxidative damage. Meanwhile, the photosynthetic performance of cotton seedlings was inhibited. Chlorophyll content, stomatal conductance, the net photosynthetic rate, the transpiration rate and water use efficiency were significantly reduced; intercellular carbon dioxide concentration and leaf stomatal limitation value increased. But photosynthesis genes (e.g., PSBO (oxygen-evolving enhancer protein 1), RBCS (ribulose bisphosphate carboxylase small chain), and FBA2 (fructose-bisphosphate aldolase 1)) in cotton were up-regulated to coordinate the photosynthetic process. Furthermore, cotton seedlings differentially regulated key biosynthesis and signaling components of phytohormonal pathways including abscisic acid, indoleacetic acid and gibberellin. This study elucidates the significant gene expression of drought-responsive transcriptional networks and relevant physiological response in cotton seedlings and offers a theoretical basis for developing water-saving irrigation strategies. Full article

(This article belongs to the Special Issue Plant Responses to Biotic and Abiotic Stresses)

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16 pages, 918 KiB

Open AccessStudy Protocol

Investigating the Impact of Fusobacterium nucleatum on Oxidative Stress, Chemoresistance, and Inflammation in Inflammatory Bowel Disease and Colorectal Cancer: Rationale and Design of a Clinical Trial

by Pierluigi Consolo, Carlotta Giorgi, Concetta Crisafulli, Francesco Fiorica, Paolo Pinton, Nicola Maurea, Sonia Missiroli, Vincenzo Quagliariello, Beatrice Mantoan, Alessandro Ottaiano, Giovanni Francesco Pellicanò, Germano Orrù, Alessandra Scano, Irene Cacciola, Teresa Pollicino, Giordana Di Mauro, Salvatore Berretta, Alessia Bignucolo, Enrica Toscano, Giuliana Ciappina and Massimiliano Berretta Show full author list Hide full author list

Int. J. Mol. Sci. 2025, 26(16), 7823; https://doi.org/10.3390/ijms26167823 - 13 Aug 2025

Abstract

Fusobacterium nucleatum (F. nucleatum), a Gram-negative anaerobe, is increasingly implicated in the pathogenesis of colorectal cancer (CRC) and inflammatory bowel disease (IBD). Its adhesin FadA enables epithelial adherence and invasion, promoting inflammation and tumorigenesis. F. nucleatum has been shown to activate [...] Read more.

Fusobacterium nucleatum (F. nucleatum), a Gram-negative anaerobe, is increasingly implicated in the pathogenesis of colorectal cancer (CRC) and inflammatory bowel disease (IBD). Its adhesin FadA enables epithelial adherence and invasion, promoting inflammation and tumorigenesis. F. nucleatum has been shown to activate the NLRP3 inflammasome, leading to IL-1β release, and is associated with chemoresistance and poor prognosis in CRC. Additionally, lipid peroxidation markers such as malondialdehyde (MDA) and 4-hydroxy-nonenal (4-HNA) may contribute to inflammation-driven carcinogenesis. This study protocol aims to investigate the role of F. nucleatum in the development and progression of IBD and CRC through integrated clinical, molecular, and imaging approaches. The protocol involves quantifying F. nucleatum in tissue biopsies across disease stages and assessing correlations with inflammatory and oxidative markers. It will explore the bacterium’s involvement in NLRP3 inflammasome activation, IL-1β production, and autophagy, and its potential contribution to chemoresistance. Furthermore, radiomic analysis of computed tomography (CT) images will be performed to identify imaging phenotypes associated with microbial load and inflammatory activity. Although primarily a protocol, the study includes preliminary in vitro data showing that exposure to FadA significantly increases inflammatory markers in Caco-2 cells, supporting the hypothesis that F. nucleatum contributes to a pro-inflammatory, pro-tumorigenic microenvironment relevant to CRC progression. Full article

(This article belongs to the Special Issue Immune-Inflammatory and Oxidative Stress Signaling Pathways Involved in Human Disorders)

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11 pages, 1482 KiB

Open AccessArticle

Deep Learning-Based Imaging Analysis Reveals Radiation-Induced Bystander Effects on Cancer Cell Migration and the Modulation by Cisplatin

by Ryosuke Seino and Hisanori Fukunaga

Int. J. Mol. Sci. 2025, 26(16), 7822; https://doi.org/10.3390/ijms26167822 - 13 Aug 2025

Abstract

Regulating tumor invasion and metastasis is pivotal for improving cancer patient prognosis. While cell migration is a key factor in these processes, the non-targeted effects of chemoradiotherapy on cell motility remain poorly understood. In this study, we employed HeLa-FUCCI cells—a cervical cancer-derived HeLa [...] Read more.

Regulating tumor invasion and metastasis is pivotal for improving cancer patient prognosis. While cell migration is a key factor in these processes, the non-targeted effects of chemoradiotherapy on cell motility remain poorly understood. In this study, we employed HeLa-FUCCI cells—a cervical cancer-derived HeLa cell line integrated with the Fluorescent Ubiquitination-Based Cell Cycle Indicator (FUCCI) probe, enabling the visualization of cell cycle phases—to investigate the radiation-induced impacts, including non-targeted effects, on cell migration. To create irradiated (In-field) and non-irradiated (out-of-field) regions, half of the culture dish was shielded with a lead block during irradiation. Cells were then exposed to 2 Gy X-rays, with or without cisplatin. Following irradiation, the cells were subjected to time-lapse imaging at 15 min intervals for 24 h, and the acquired data were analyzed using cell segmentation and tracking algorithms, Cellpose 2.0 and TrackMate 7. Without cisplatin, the migration velocity and total distance traveled of Out-of-field cells were significantly reduced compared to controls, suggesting a suppressive bystander signal. In contrast, with cisplatin treatment, these parameters significantly increased in both In-field and Out-of-field cells. This suggests that chemoradiotherapy may inadvertently enhance tumor cell motility outside the target volume, a critical finding with significant implications for therapeutic outcomes. Full article

(This article belongs to the Special Issue Effects of Ionizing Radiation in Cancer Radiotherapy: 2nd Edition)

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13 pages, 1730 KiB

Open AccessArticle

Patient Stratification for Serum LDH Levels Reveals Distinct CLA⁺ T-Cell Cytokine Secretion in Response to HDM, Clinical Features and Allergic Comorbidities

by Irene García-Jiménez, Ignasi Figueras-Nart, Lídia Sans-de San Nicolás, Laia Curto-Barredo, Marta Bertolín-Colilla, Montserrat Bonfill-Ortí, Sandra Díez-Ribas, Alex Llobet-del Pino, Antonio Guilabert-Vidal, Anna Ryzhkova, Marta Ferran, Ramon M. Pujol and Luis F. Santamaria-Babí

Int. J. Mol. Sci. 2025, 26(16), 7821; https://doi.org/10.3390/ijms26167821 - 13 Aug 2025

Abstract

Lactate dehydrogenase (LDH) is a serum biomarker well known to correlate with disease severity in atopic dermatitis (AD). The aim of this study was to explore the cutaneous immune responses and the clinical profile of AD patients in relation to serum LDH levels. [...] Read more.

Lactate dehydrogenase (LDH) is a serum biomarker well known to correlate with disease severity in atopic dermatitis (AD). The aim of this study was to explore the cutaneous immune responses and the clinical profile of AD patients in relation to serum LDH levels. To this end, 47 untreated, adult patients with moderate-to-severe AD were stratified by median levels of serum LDH. Circulating memory T-cell responses to house dust mite (HDM) extract, in the presence of autologous lesional epidermal cells, were compared between AD subgroups. The LDH^high group exhibited significantly higher IL-13, IL-5 and IL-9 in vitro responses confined to the cutaneous lymphocyte-associated antigen (CLA)⁺ subset compared to LDH^low patients. Clinically, LDH^high patients were younger and exhibited more severe disease, elevated eosinophil counts in their blood, increased total and specific IgE levels in their plasma, and a higher prevalence of allergic rhinitis. Our data suggests that high LDH levels identify a subgroup of AD patients with a specific immune and clinical profile, and highlight the potential of LDH as a clinical parameter that may enable patient stratification for treatment selection. Full article

(This article belongs to the Special Issue Dermatology: Advances in Pathophysiology and Therapies (3rd Edition))

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27 pages, 1069 KiB

Open AccessReview

Recent Advances and Future Directions in Alzheimer’s Disease Genetic Research

by Mikaela Stancheva, Draga Toncheva and Sena Karachanak-Yankova

Int. J. Mol. Sci. 2025, 26(16), 7819; https://doi.org/10.3390/ijms26167819 - 13 Aug 2025

Abstract

Alzheimer’s disease (AD) is a complex neurodegenerative condition which, despite its high prevalence and socioeconomic impact on the world, has an etiology that remains poorly understood. The genetic causes of AD are complex and have been continuously studied for decades. They range from [...] Read more.

Alzheimer’s disease (AD) is a complex neurodegenerative condition which, despite its high prevalence and socioeconomic impact on the world, has an etiology that remains poorly understood. The genetic causes of AD are complex and have been continuously studied for decades. They range from rare pathogenic, highly penetrant mutations in early-onset (EOAD) forms, which account for 5% of the cases to multiple-risk alleles across different genes in late-onset (LOAD) forms. Monogenic causes of EOAD allocate within APP, PSEN1, and PSEN2 genes in 10–15% of cases. The most significant risk factor in LOAD heritability is the APOE ε4 allele, as well as numerous loci within genes involved in immunity, endocytosis, lipid metabolism, and amyloid and tau processing. LOAD can also be attributed to the accumulation of somatic mutations, which may be detected by analysis of brain-derived cell-free DNA (cfDNA) in plasma. This review offers a comprehensive overview of the genetic architecture of Alzheimer’s disease, with particular focus on the molecular mechanisms underlying both early- and late-onset forms of the condition. An improved understanding of the genetic etiology of AD can aid better prevention, earlier diagnosis, and novel therapeutic approaches. This can be achieved by analyzing understudied populations, performing case-control studies with appropriately matched controls, and surveying brain-derived cell-free DNA in plasma, with the latter having the potential to contribute to the implementation of liquid biopsy in dementology. Full article

(This article belongs to the Special Issue Molecular Progression of Genome-Related Diseases)

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21 pages, 1957 KiB

Open AccessArticle

Importin α3 Is Tolerant to Nuclear Localization Signal Chirality

by Felipe Hornos, Bruno Rizzuti and José L. Neira

Int. J. Mol. Sci. 2025, 26(16), 7818; https://doi.org/10.3390/ijms26167818 - 13 Aug 2025

Abstract

Several carrier proteins are involved in nuclear translocation from the cytoplasm to the nucleus in eukaryotic cells. We have previously demonstrated the binding of several intact folded and disordered proteins to the human isoform importin α3 (Impα3); furthermore, disordered peptides, corresponding to their [...] Read more.

Several carrier proteins are involved in nuclear translocation from the cytoplasm to the nucleus in eukaryotic cells. We have previously demonstrated the binding of several intact folded and disordered proteins to the human isoform importin α3 (Impα3); furthermore, disordered peptides, corresponding to their nuclear localization signals (NLSs), also interact with Impα3. These proteins and their isolated NLSs also bind to the truncated importin species ∆Impα3, which does not contain the N-terminal disordered importin binding domain (IBB). In this work, we added a further ‘layer’ of conformational disorder to our studies, testing whether the isolated D-enantiomers of NLSs of selected proteins, either folded or unfolded, were capable of binding to both Impα3 and ∆Impα3. The D-enantiomers, like their L-form counterparts, were monomeric and disordered in isolation, as shown by nuclear magnetic resonance (NMR). We measured the ability of such D-enantiomeric NLSs to interact with both importin species by using fluorescence, biolayer interferometry (BLI), isothermal titration calorimetry (ITC), and molecular simulations. In all cases, the binding affinities were within the same range as those measured for their L-isomer counterparts for either Impα3 or ∆Impα3, and the binding locations corresponded to the major NLS binding site of the protein. Thus, the stereoisomeric nature is not important in defining the binding of proteins to the main component of classical cellular translocation machinery, although the primary structure of the hot-spot site for NLS binding of importin is well defined. Full article

(This article belongs to the Section Biochemistry)

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26 pages, 627 KiB

Open AccessReview

Seed Amplification Assay for α-Synuclein: Diagnostic Applications in Synucleinopathies

by Alexandros Giannakis, Louisa Pechlivani, Chrissa Sioka, George Alexiou, Spiridon Konitsiotis and Athanasios Kyritsis

Int. J. Mol. Sci. 2025, 26(16), 7817; https://doi.org/10.3390/ijms26167817 - 13 Aug 2025

Abstract

Seed amplification assays (SAA) targeting misfolded α-synuclein have emerged as powerful tools for the diagnosis and study of synucleinopathies, including Parkinson’s disease (PD), dementia with Lewy bodies, and multipßle system atrophy. These assays exploit the prion-like seeding properties of pathological α-synuclein to detect [...] Read more.

Seed amplification assays (SAA) targeting misfolded α-synuclein have emerged as powerful tools for the diagnosis and study of synucleinopathies, including Parkinson’s disease (PD), dementia with Lewy bodies, and multipßle system atrophy. These assays exploit the prion-like seeding properties of pathological α-synuclein to detect minute amounts of misfolded protein in biological specimens. the PubMed database was searched according to our study criteria, and 55 clinical studies comprised the final literature review. the majority of studies have focused on patients at various stages of PD, with cerebrospinal fluid (CSF) being the most commonly investigated biological specimen. Diagnostic utility was most pronounced in the CSF of PD patients, whereas results from other biological samples and across different synucleinopathies have been more modest. α-syn SAA demonstrate significant diagnostic potential in synucleinopathies. Additional applications may include monitoring disease progression. Future studies should explore the utility of α-syn SAA in alternative biological specimens, assess its performance across various synucleinopathies and other neurodegenerative diseases, and determine its comparative diagnostic value. Full article

(This article belongs to the Section Molecular Neurobiology)

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17 pages, 663 KiB

Open AccessArticle

The Association of Adropin with Asymptomatic Coronary Calcification in Patients in Early Stages of Chronic Kidney Disease

by Tetiana A. Berezina, Oleksandr O. Berezin, Evgen V. Novikov and Alexander E. Berezin

Int. J. Mol. Sci. 2025, 26(16), 7816; https://doi.org/10.3390/ijms26167816 - 13 Aug 2025

Abstract

Early stages of chronic kidney disease (CKD) are closely associated with vascular remodeling and coronary artery calcification. The aim of this study is to determine whether adropin is associated with asymptomatic coronary calcification in patients in the early stages of CKD. This study [...] Read more.

Early stages of chronic kidney disease (CKD) are closely associated with vascular remodeling and coronary artery calcification. The aim of this study is to determine whether adropin is associated with asymptomatic coronary calcification in patients in the early stages of CKD. This study enrolled 337 individuals fulfilling the inclusion criteria of the early stages of CKD (G1–2, A1–3) and divided them into two subgroups with (n = 196) and without (n = 141) asymptomatic coronary artery calcification. Native coronary multi-detector computed tomography angiography was conducted to determine coronary artery calcification, which was stratified into four grades according to the Agatston method. Serum levels of adropin were measured by ELISA. The patients with known asymptomatic coronary artery calcification had significantly lower levels of adropin than those without this condition. The levels of adropin in individuals with mild (130–199 HU), moderate (200–299 HU), severe (300–399 HU) and very severe (≥400 HU) calcification were 3.13 (95% CI = 1.92–4.21) ng/mL, 2.3 (95% CI = 1.45–3.6) ng/mL, 2.1 (95% CI = 1.22–3.25) ng/mL and 1.26 (95% CI = 1.13–1.98) ng/mL, respectively. In multivariate logistic regression low adropin (<2.95 ng/mL), a presence of hypertension, type 2 diabetes mellitus (T2DM) exerted their independent potencies to predict asymptomatic coronary calcification. Moreover, adropin demonstrated better discriminative potency than concomitant hypertension and T2DM. Conclusions: Low levels of circulating adropin significantly predicted a risk of coronary artery calcification in patients in the early stages of CKD. Full article

(This article belongs to the Special Issue Atherosclerosis: From Molecular Biology to Therapeutic Perspective: 6th Edition)

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21 pages, 4815 KiB

Open AccessArticle

Native putA Overexpression in Synechocystis sp. PCC 6803 Significantly Enhances Polyhydroxybutyrate Production, Further Augmented by the adc1 Knockout Under Prolonged Nitrogen Deprivation

by Suthira Utharn, Peter Lindblad and Saowarath Jantaro

Int. J. Mol. Sci. 2025, 26(16), 7815; https://doi.org/10.3390/ijms26167815 - 13 Aug 2025

Abstract

This study highlights a new avenue to improve polyhydroxybutyrate (PHB) productivity by optimizing genes related to arginine catabolism, which influences nitrogen metabolism in cyanobacteria based on the carbon/nitrogen metabolism balance. In the Synechocystis sp. PCC 6803 wild type (WT) and its adc1 mutant [...] Read more.

This study highlights a new avenue to improve polyhydroxybutyrate (PHB) productivity by optimizing genes related to arginine catabolism, which influences nitrogen metabolism in cyanobacteria based on the carbon/nitrogen metabolism balance. In the Synechocystis sp. PCC 6803 wild type (WT) and its adc1 mutant (Δadc1), the native putA gene, responsible for the oxidation of proline to glutamate, was overexpressed to create the OXPutA and OXPutA/Δadc1 strains, respectively. PHB accumulation was considerably higher in OXPutA and OXPutA/Δadc1 under the nitrogen-deprived condition than in strains that overexpressed the proC gene, involved in proline synthesis. The increased transcript level of glgX, associated with glycogen degradation, confirmed that glycogen served as the primary carbon source for PHB synthesis under nitrogen stress without any carbon source addition. Furthermore, proline and glutamate level changes helped cells deal with nitrogen stress and considerably improve intracellular carbon/nitrogen metabolism. As indicated by elevated levels of proA and argD transcripts as well as chlorophyll a accumulation, this impact was most noticeable in strains that overexpressed putA, which was crucial for the synthesis of glutamate, a precursor for important metabolic pathways that respond to nitrogen stress. Therefore, our metabolic model presents PHB-producing strains as promising candidates for biomaterial biotechnology applications in medical and agricultural fields. Full article

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

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19 pages, 622 KiB

Open AccessReview

Decoding Pancreatic Neuroendocrine Tumors: Molecular Profiles, Biomarkers, and Pathways to Personalized Therapy

by Linda Galasso, Federica Vitale, Gabriele Giansanti, Giorgio Esposto, Raffaele Borriello, Irene Mignini, Alberto Nicoletti, Lorenzo Zileri Dal Verme, Antonio Gasbarrini, Maria Elena Ainora and Maria Assunta Zocco

Int. J. Mol. Sci. 2025, 26(16), 7814; https://doi.org/10.3390/ijms26167814 - 13 Aug 2025

Abstract

Pancreatic neuroendocrine tumors (pNETs) are rare malignancies, accounting for 1–2% of pancreatic cancers, with an incidence of ≤1 case per 100,000 individuals annually. Originating from pancreatic endocrine cells, pNETs display significant clinical and biological heterogeneity. Traditional classification based on proliferative grading does not [...] Read more.

Pancreatic neuroendocrine tumors (pNETs) are rare malignancies, accounting for 1–2% of pancreatic cancers, with an incidence of ≤1 case per 100,000 individuals annually. Originating from pancreatic endocrine cells, pNETs display significant clinical and biological heterogeneity. Traditional classification based on proliferative grading does not fully capture the complex mechanisms involved, such as oxidative stress, mitochondrial dysfunction, and tumor-associated macrophage infiltration. Recent advances in molecular profiling have revealed key oncogenic drivers, including MEN1 (menin 1), DAXX (death domain–associated protein), ATRX (alpha thalassemia/mental retardation syndrome X-linked), CDKN1B (cyclin-dependent kinase inhibitor 1B) mutations, chromatin remodeling defects, and dysregulation of the mTOR pathway. Somatostatin receptors, particularly SSTR2, play a central role in tumor biology and serve as important prognostic markers, enabling the use of advanced diagnostic imaging (e.g., Gallium-68 DOTATATE PET/CT) and targeted therapies like somatostatin analogs and peptide receptor radionuclide therapy (PRRT). Established biomarkers such as Chromogranin A and the Ki-67 proliferation index remain vital for diagnosis and prognosis, while emerging markers, like circulating tumor DNA and microRNAs, show promise for enhancing disease monitoring and diagnostic accuracy. This review summarizes the molecular landscape of pNETs and highlights genomic, transcriptomic, proteomic, and epigenomic factors that support the identification of novel diagnostic, prognostic, and therapeutic biomarkers, ultimately advancing personalized treatment strategies. Full article

(This article belongs to the Special Issue Novel Molecular Mechanisms Underlying Tumorigenesis and Innovative Therapeutic Approaches for Cancer-Fighting, 3rd Edition)

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22 pages, 3081 KiB

Open AccessArticle

Elucidating the Complex Structural and Molecular Mechanisms Driving P-Glycoprotein-Mediated Transport of Cardiac Glycosides

by Rohit Katti, Amanda M. Kozarich, Gershon A. K. Mensah and Michael G. Bartlett

Int. J. Mol. Sci. 2025, 26(16), 7813; https://doi.org/10.3390/ijms26167813 - 13 Aug 2025

Abstract

P-glycoprotein (Pgp) plays a significant role in the disposition of cardiac glycoside (CG) drugs across the cell membrane. The relatively narrow therapeutic indices of these drugs, coupled with the co-administration of drugs that inhibit Pgp’s transport mechanism, often cause an increased level of [...] Read more.

P-glycoprotein (Pgp) plays a significant role in the disposition of cardiac glycoside (CG) drugs across the cell membrane. The relatively narrow therapeutic indices of these drugs, coupled with the co-administration of drugs that inhibit Pgp’s transport mechanism, often cause an increased level of CG in the patient’s plasma, resulting in fatal arrhythmia. Therefore, understanding the underlying mechanism of the CG–Pgp interaction is necessary to circumvent Pgp-mediated transport and effectively design next-generation CGs. In this study, we conducted a comparative analysis to examine the interaction with Pgp and further understand the Pgp-mediated transport of digoxin, digitoxin, digoxigenin, and digitoxigenin. Through the drug-induced kinetic studies of Pgp, our findings suggest that each of the four drugs tested has a single binding site within Pgp. The CG–Pgp binding studies demonstrated that digoxin, digitoxin, and digoxigenin had relatively higher binding affinities. The CG-mediated conformational changes in Pgp indicated that each of the drugs shifts Pgp to an “outward-open” conformation in a nucleotide-dependent manner. STDD NMR indicated that the protons within the δ-lactone ring and the tri-D-digitoxose sugar moieties (glycones) predominantly interact with Pgp. Finally, a model was proposed for CG-induced Pgp-mediated ATP hydrolysis and transport by integrating our data with previously published Pgp-mediated CG transport results. Full article

(This article belongs to the Section Molecular Biophysics)

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32 pages, 1659 KiB

Open AccessReview

Vagal Oxytocin Receptors as Molecular Targets in Gut–Brain Signaling: Implications for Appetite, Satiety, Obesity, and Esophageal Motility—A Narrative Review

by Agnieszka Nowacka, Maciej Śniegocki and Ewa A. Ziółkowska

Int. J. Mol. Sci. 2025, 26(16), 7812; https://doi.org/10.3390/ijms26167812 - 13 Aug 2025

Abstract

Oxytocin (OT), traditionally associated with reproduction and social bonding, has emerged as a key modulator of gastrointestinal (GI) physiology and appetite regulation behavior through its actions within the gut–brain axis. Central to this regulation are vagal oxytocin receptors (VORs), which are located along [...] Read more.

Oxytocin (OT), traditionally associated with reproduction and social bonding, has emerged as a key modulator of gastrointestinal (GI) physiology and appetite regulation behavior through its actions within the gut–brain axis. Central to this regulation are vagal oxytocin receptors (VORs), which are located along vagal afferent and efferent fibers and within brainstem nuclei such as the nucleus tractus solitarius and dorsal motor nucleus of the vagus. This review presents a comprehensive synthesis of current knowledge on the anatomical distribution, molecular signaling, developmental plasticity, and functional roles of VORs in the regulation of GI motility, satiety, and energy homeostasis. We highlight how VORs integrate hormonal, microbial, and stress-related cues and interact with other neuropeptidergic systems including GLP-1, CCK, and nesfatin-1. Recent advances in spatial transcriptomics, single-nucleus RNA sequencing, chemogenetics, and optogenetics are discussed as transformative tools for mapping and manipulating VOR-expressing circuits. Particular attention is given to sex differences, translational challenges, and the limited understanding of VOR function in humans. This article proposes VORs as promising therapeutic targets in dysphagia, obesity, and functional GI disorders. We outline future research priorities, emphasizing the need for integrative, cross-species approaches to clarify VOR signaling and guide the development of targeted, personalized interventions. Full article

(This article belongs to the Special Issue Recent Research in Gut Microbiota–Gut–Brain Axis)

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17 pages, 10183 KiB

Open AccessArticle

IL-15 Promotes the Survival of Anti-Inflammatory (M2), Immunoinhibitory (IL-10+) Dermal Macrophages in Human Eyelid Skin Under IFNγ-Dominated Inflammatory Conditions

by Dana-Lee Demetrius, Sofia M. Perez, Takahiro Suzuki, Jennifer Gherardini, Wendy Lee, Jérémy Chéret and Ralf Paus

Int. J. Mol. Sci. 2025, 26(16), 7811; https://doi.org/10.3390/ijms26167811 - 13 Aug 2025

Abstract

Interleukin (IL)-15 is primarily known as a pro-inflammatory and anti-apoptotic cytokine, which stimulates the proliferation and survival of key immunocytes, including macrophages (MACs). Yet, it remains unclear how IL-15 specifically impacts MACs in intact human skin, particularly immunoinhibitory, IL-10-producing/secreting M2 MACs (CD206⁺ [...] Read more.

Interleukin (IL)-15 is primarily known as a pro-inflammatory and anti-apoptotic cytokine, which stimulates the proliferation and survival of key immunocytes, including macrophages (MACs). Yet, it remains unclear how IL-15 specifically impacts MACs in intact human skin, particularly immunoinhibitory, IL-10-producing/secreting M2 MACs (CD206⁺IL-10⁺). In the current pilot study, we explored this in organ-cultured healthy human eyelid skin in the presence of IFNγ (100 IU/mL) to mimic a pro-inflammatory signaling milieu found in several chronic immunodermatoses. Quantitative immunohistomorphometry showed that IFNγ significantly reduced the number of CD68⁺MACs, M2 CD206⁺MACs, and immunoinhibitory CD206⁺IL-10⁺MACs. Moreover, co-administering recombinant human (rh) IL-15 after inducing inflammation by IFNγ largely reversed the IFNγ-induced decline in MAC populations. To investigate if this was mediated via the private IL-15 receptor alpha (IL-15Rα), we successfully silenced IL-15Rα in human skin ex vivo. Indeed, co-administration of IL-15Rα siRNA abrogated the rhIL-15 protection of M2 CD206⁺MACs against IFNγ, but not of the CD206⁺IL-10⁺MAC subpopulation. These pilot data suggest that IL-15 maintains immunoinhibitory M2 CD206⁺IL-10⁺MACs in human skin under IFNγ-dominated inflammatory conditions. Therefore, it deserves to be explored whether IL-15 or IL-15Rα agonists can exert therapeutic benefit in chronic inflammatory dermatoses by preserving the intracutaneous pool of anti-inflammatory dermal M2 MACs. Full article

(This article belongs to the Special Issue Molecular and Cellular Mechanisms of Skin Diseases (Second Edition))

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19 pages, 2367 KiB

Open AccessArticle

Integrative High-Throughput RNAi Screening Identifies BRSK1, STK32C and STK40 as Novel Activators of YAP/TAZ

by Mandeep K. Gill, Siyuan Song, Tania Christova and Liliana Attisano

Int. J. Mol. Sci. 2025, 26(16), 7810; https://doi.org/10.3390/ijms26167810 - 13 Aug 2025

Abstract

Disruption of the Hippo pathway leads to activation of the YAP/TAZ transcriptional program which promotes tumor initiation, progression and metastasis in diverse cancers. Aggressive triple-negative breast cancers (TNBC) lack an effective therapy; thus, inactivating YAP and TAZ has emerged as an attractive approach [...] Read more.

Disruption of the Hippo pathway leads to activation of the YAP/TAZ transcriptional program which promotes tumor initiation, progression and metastasis in diverse cancers. Aggressive triple-negative breast cancers (TNBC) lack an effective therapy; thus, inactivating YAP and TAZ has emerged as an attractive approach and a new treatment modality. Thus, we performed two complementary high-throughput RNAi-based kinome screens to uncover cancer-associated activators of YAP/TAZ in two TNBC cell lines, MDA-MB231 and MDA-MB468. Integrated analysis that combined a YAP/TAZ localization screen with a TEAD-luciferase reporter screen, identified novel regulators including BRSK1, STK32C and STK40. The AMPK family members NUAKs, MARKs and SIKs are known to inhibit the Hippo kinase cassette; here, we uncover BRSK1, another AMPK family member as a regulator of YAP/TAZ. We also reveal that two poorly studied kinases, STK32C, a member of the AGC family, and STK40, a pseudokinase, can also inhibit the activity of YAP/TAZ. Thus, our studies expand the repertoire of known AMPK family members and reveal two new kinases that modulate the Hippo pathway and may play a role in YAP/TAZ driven breast cancers. Further analysis of other screen hits may similarly uncover new regulators that could be targeted for therapeutic interventions. Full article

(This article belongs to the Section Molecular Biology)

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52 pages, 3866 KiB

Open AccessReview

Beyond Oxidation: Engineering Functional Anodised Metal Matrices Through Molecular and Surface Modifications

by Mateusz Schabikowski, Agnieszka Stróż and Andrzej Kruk

Int. J. Mol. Sci. 2025, 26(16), 7809; https://doi.org/10.3390/ijms26167809 - 13 Aug 2025

Abstract

Anodised metal matrices represent a versatile and multifunctional platform for the development of advanced materials with tunable physicochemical properties. Through electrochemical oxidation processes—commonly referred to as anodisation—metals such as aluminium, titanium, niobium, zinc and tantalum can be transformed into structured oxide layers with [...] Read more.

Anodised metal matrices represent a versatile and multifunctional platform for the development of advanced materials with tunable physicochemical properties. Through electrochemical oxidation processes—commonly referred to as anodisation—metals such as aluminium, titanium, niobium, zinc and tantalum can be transformed into structured oxide layers with defined porosity, thickness and surface morphology. These methods enable the fabrication of ordered nanoporous arrays, nanotubes and nanowires, depending on the process parameters and the type of metal. The review introduces and outlines the various anodisation techniques and parameters. This is crucial, since each individual metal requires specified optimal conditions to obtain a stable anodised oxide layer. This review provides an overview of recent advances in the design and application of anodised metal substrates, with the focus on their role as functional platforms in catalysis, sensing, energy storage and biomedical engineering. Special attention is given to post-anodisation surface modification strategies, such as chemical functionalisation, thin-film deposition and molecular-level integration, which significantly expand the utility of these materials. The review also highlights the challenges, limitations and future perspectives of anodising technologies, aiming to guide the rational design of next-generation devices based on engineered oxide architectures. Full article

(This article belongs to the Special Issue Functional Nanomaterials: Structures, Compositions and Various Applications)

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25 pages, 4702 KiB

Open AccessPerspective

Integrative Diagnostic and Prognostic Paradigms in Diffuse Axonal Injury: Insights from Clinical, Histopathological, Biomolecular, Radiological, and AI-Based Perspectives

by Alessandro Santurro, Matteo De Simone, Anis Choucha, Donato Morena, Francesca Consalvo, Daniele Romano, Pamela Terrasi, Francesco Corrivetti, Raffaele Scrofani, Nicola Narciso, Ettore Amoroso, Marco Cascella, Vittorio Fineschi and Giorgio Iaconetta

Int. J. Mol. Sci. 2025, 26(16), 7808; https://doi.org/10.3390/ijms26167808 - 13 Aug 2025

Abstract

Diffuse axonal injury (DAI) is one of the most severe consequences of traumatic brain injury (TBI), characterized by widespread axonal damage in the cerebral white matter. DAI plays a crucial role in determining clinical outcomes, significantly contributing to long-term disability and mortality in [...] Read more.

Diffuse axonal injury (DAI) is one of the most severe consequences of traumatic brain injury (TBI), characterized by widespread axonal damage in the cerebral white matter. DAI plays a crucial role in determining clinical outcomes, significantly contributing to long-term disability and mortality in severe cases. Despite advancements in neuroscience and clinical management, the diagnosis and prognosis of DAI remain challenging due to its complex pathophysiology and the difficulty of detecting axonal damage in its early stages. This study critically analyzes the clinical and post-mortem methodologies used to assess DAI, highlighting their strengths and limitations. Traditional histopathological grading systems provide valuable insights into disease progression, yet their correlation with long-term functional outcomes remains controversial. Advanced neuroimaging techniques, such as diffusion-weighted MRI, have improved lesion detection, although their routine clinical application is still limited. Additionally, emerging approaches involving biomarkers and artificial intelligence-based models hold promise for enhancing diagnostic accuracy and prognostic predictions. By synthesizing current knowledge on DAI, this work aims to outline a comprehensive framework for improving diagnosis and outcome assessment. Furthermore, it seeks to foster collaboration among clinicians and researchers, ultimately advancing the understanding of DAI and refining strategies to improve patient care. Full article

(This article belongs to the Special Issue Latest Advances in Oxidative Stress and Brain Injury)

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11 pages, 586 KiB

Open AccessArticle

Fibroblast Activation Protein (FAP) as a Serum Biomarker for Fibrotic Ovarian Aging: A Clinical Validation Study Based on Translational Transcriptomic Targets

by Hyun Joo Lee, Yunju Jo, Shibo Wei, Eun Hee Yu, Sul Lee, Dongryeol Ryu and Jong Kil Joo

Int. J. Mol. Sci. 2025, 26(16), 7807; https://doi.org/10.3390/ijms26167807 - 13 Aug 2025

Abstract

Chronological age is an imprecise proxy for reproductive capacity, necessitating biomarkers that reflect the underlying pathophysiology of the ovary. Fibrotic remodeling of the ovarian stroma is a key hallmark of biological ovarian aging, yet it cannot be assessed by current clinical tools. This [...] Read more.

Chronological age is an imprecise proxy for reproductive capacity, necessitating biomarkers that reflect the underlying pathophysiology of the ovary. Fibrotic remodeling of the ovarian stroma is a key hallmark of biological ovarian aging, yet it cannot be assessed by current clinical tools. This study aimed to identify and validate a novel serum biomarker for fibrotic ovarian aging by applying supervised machine learning (ML) to human ovarian transcriptomic data. Transcriptomic data from the Genotype-Tissue Expression (GTEx) database were analyzed using ML algorithms to identify candidate genes predictive of ovarian aging, and finally, fibroblast activation protein (FAP) and collectin-11 (COLEC11) were selected for clinical validation. In a cross-sectional study, serum levels of FAP and COLEC11, along with key hormonal indices, were measured in two nested patient cohorts, and their associations with ovarian reserve and clinical parameters were analyzed. Serum FAP levels did not correlate with age but showed a strong inverse correlation with anti-Müllerian hormone (AMH) (r = −0.61, p = 0.001), a finding accentuated in women with decreased ovarian reserve (DOR). While COLEC11 correlated with age, it failed to differentiate DOR status. FAP levels were independent of central hormonal regulation, consistent with preclinical fibrotic models. Circulating FAP reflects age-independent, fibrotic ovarian aging, offering stromal-specific information not captured by conventional hormonal markers. This study provides the first clinical validation of FAP as a biomarker for ovarian stromal aging, holding potential for improved reproductive risk assessment. Full article

(This article belongs to the Section Molecular Biology)

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Int. J. Mol. Sci., Volume 26, Issue 16 (August-2 2025) – 176 articles

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