International Journal of Molecular Sciences

27 pages, 619 KiB

Open AccessReview

Nanomaterials in COPD: Emerging Therapeutic and Diagnostic Frontiers with a Focus on Metal–Organic Frameworks

by Antonio Tiralosi, Manuela Cambria, Mariachiara Campanella, Vincenzo Paratore, Cristina Russo, Lucia Malaguarnera, Maria Stella Valle and Maria Teresa Cambria

Int. J. Mol. Sci. 2025, 26(16), 8025; https://doi.org/10.3390/ijms26168025 - 19 Aug 2025

Abstract

Chronic obstructive pulmonary disease (COPD) is one of the leading causes of morbidity and mortality worldwide. Although conventional therapies are effective in controlling symptoms, they remain limited in altering the course of the disease and significantly reducing the chronic inflammation and oxidative stress [...] Read more.

Chronic obstructive pulmonary disease (COPD) is one of the leading causes of morbidity and mortality worldwide. Although conventional therapies are effective in controlling symptoms, they remain limited in altering the course of the disease and significantly reducing the chronic inflammation and oxidative stress underlying it. In this context, nanoparticles and nanomaterials are emerging as innovative tools capable of overcoming traditional pharmacological barriers due to their ability to deliver therapeutic oligonucleotides, antioxidants, and drugs in a targeted manner, modulate immune responses, and improve the bioavailability of active compounds. In particular, metal–organic frameworks (MOFs) stand out as ideal candidates for inhalable drug delivery in COPD, owing to their permanent crystalline porous structure, high specific surface area, and versatile chemical functionalization. This review provides the most recent preclinical evidence on the use of different nanoparticles in COPD, with a focus on the therapeutic and diagnostic potential of MOFs. It discusses their biocompatibility, drug loading strategies, and controlled release mechanisms and explores future perspectives for clinical translation. Full article

(This article belongs to the Section Molecular Nanoscience)

14 pages, 3021 KiB

Open AccessArticle

Formation of Supramolecular Structures in Oxidation Processes Catalyzed by Heteroligand Complexes of Iron and Nickel: Models of Enzymes

by Ludmila Ivanovna Matienko, Elena M. Mil, Anastasia A. Albantova and Alexander N. Goloshchapov

Int. J. Mol. Sci. 2025, 26(16), 8024; https://doi.org/10.3390/ijms26168024 - 19 Aug 2025

Abstract

In some cases, the catalytic processes involve the formation of self-organized supramolecular structures due to H-bonds and other non-covalent interactions. It has been suggested that the construction of self-assembled catalytic systems is a promising strategy to mimic enzyme catalysis at the model level. [...] Read more.

In some cases, the catalytic processes involve the formation of self-organized supramolecular structures due to H-bonds and other non-covalent interactions. It has been suggested that the construction of self-assembled catalytic systems is a promising strategy to mimic enzyme catalysis at the model level. As a rule, the real catalysts are not the primary catalytic complexes, but rather, those that are formed during the catalytic process. In our earlier works, we have established that the effective catalysts M(II)_xL¹_y(L¹_ox)_z(L²)_n(H₂O)_m (M = Ni, Fe, L¹ = acac⁻, L² = activating electron-donating ligand) for the selective oxidation of ethylbenzene to α-phenyl ethyl hydroperoxide are the result of the transformation of primary (Ni(Fe)L¹)_x(L²)_y complexes during the oxidation of ethylbenzene. In addition, the mechanism of the transformation to active complexes is similar to the mechanism of action of NiFeARD (NiFe-acireductone dioxygenase). Based on kinetic and spectrophotometric data, we hypothesized that the high stability of effective catalytically active complexes may be associated with the formation of stable supramolecular structures due to intermolecular hydrogen bonds and possibly other non-covalent bonds. We confirmed this assumption using AFM. In this work, using AFM, we studied the possibility of forming supramolecular structures based on iron complexes with L²-crown ethers and quaternary ammonium salts, which are catalysts for the oxidation of ethylbenzene and are models of FeARD (Fe-acireductone dioxygenase). The formation of supramolecular structures based on complexes of natural Hemin with PhOH and L-histidine or Hemin with L-tyrosine and L-histidine, which are models of heme-dependent tyrosine hydroxylase and cytochrome P450-dependent monooxygenases (AFM method), may indicate the importance of outer-sphere regulatory interactions with the participation of Tyrosine and Histidine in the mechanism of action of these enzymes. Full article

(This article belongs to the Special Issue Design of Functional Supramolecular Architectures Based on Coordination Compounds)

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21 pages, 2101 KiB

Open AccessArticle

Prostaglandins Regulate Urinary Purines by Modulating Soluble Nucleotidase Release in the Bladder Lumen

by Mahsa Borhani Peikani, Alejandro Gutierrez Cruz, Zoe S. Buckley and Violeta N. Mutafova-Yambolieva

Int. J. Mol. Sci. 2025, 26(16), 8023; https://doi.org/10.3390/ijms26168023 - 19 Aug 2025

Abstract

Distention of the urinary bladder wall during filling stretches the urothelium and induces the release of chemical mediators, including adenosine 5′-triphosphate (ATP) and prostaglandins (PGs), that transmit signals between cells within the bladder wall. The urothelium also releases soluble nucleotidases (s-NTDs) that control [...] Read more.

Distention of the urinary bladder wall during filling stretches the urothelium and induces the release of chemical mediators, including adenosine 5′-triphosphate (ATP) and prostaglandins (PGs), that transmit signals between cells within the bladder wall. The urothelium also releases soluble nucleotidases (s-NTDs) that control the availability of ATP and its metabolites at receptor sites in umbrella cells and cells deeper in the bladder wall, as well as in the urine. This study investigated whether PGs regulate the intravesical breakdown of ATP by s-NTDs. Using a murine decentralized mucosa-only bladder model and an HPLC technology with fluorescence detection, we evaluated the decrease in ATP and increase in ADP, AMP, and adenosine (ADO) in intraluminal solutions (ILS) collected at the end of physiological bladder filling. PGD₂, PGE₂, and PGI₂, but not PGF_2α, inhibited the conversion of AMP (produced from ATP) to ADO, likely due to a suppressed intravesical release of s-AMPases. The effects of exogenous PGD₂, PGE₂, and PGI₂ were mediated by DP1/DP2, EP2, and IP prostanoid receptors, respectively. Activation of either DP1 or DP2 receptors by endogenous PGD₂ also led to AMP increase and ADO decrease in ILS-containing ATP substrate. Finally, PGs produced by either COX-1 or COX-2 inhibited the hydrolysis of AMP to ADO. Together, these observations suggest that (1) endogenous PGs (chiefly PGD₂, and to lesser degree PGE₂ and PGI₂) allow release of s-NTDs like s-ATPases and s-ADPases but impede the formation of ADO from intravesical ATP by inhibiting the release of s-NTDs/s-AMPases; (2) it is possible that high concentrations of PGD₂, PGE₂ and PGI₂, as anticipated in inflammation or bladder pain syndrome, delay the ADO production and prolong the action of excitatory purine mediators; and (3) either COX-1 and COX-2 are constitutively expressed in the mouse bladder mucosa or COX-2 is induced by distention of the urothelium during bladder filling. Full article

(This article belongs to the Special Issue Advances in the Purinergic System)

13 pages, 1956 KiB

Open AccessArticle

Differential Immune Checkpoint Expression in CD4⁺ and CD4⁻ NKT Cell Populations During Healthy Pregnancy

by Matyas Meggyes, Nagy U. David, Livia Mezosi, Fanni Vastag, Dora Kevey and Laszlo Szereday

Int. J. Mol. Sci. 2025, 26(16), 8022; https://doi.org/10.3390/ijms26168022 - 19 Aug 2025

Abstract

This study investigated the expression of immune checkpoint molecules on CD4⁺ and CD4⁻ NKT cell subpopulations throughout healthy pregnancy trimesters and in non-pregnant condition to understand their role in maternal–fetal immunotolerance. Using flow cytometry, we found that CD4⁻ NKT cells [...] Read more.

This study investigated the expression of immune checkpoint molecules on CD4⁺ and CD4⁻ NKT cell subpopulations throughout healthy pregnancy trimesters and in non-pregnant condition to understand their role in maternal–fetal immunotolerance. Using flow cytometry, we found that CD4⁻ NKT cells significantly outnumbered CD4⁺ NKT cells in all investigated groups. In the case of the immune checkpoint molecules, PD-1 receptor expression was significantly lower in CD4⁻ NKT cells compared to CD4⁺ counterpart cells only in non-pregnant women, while the PD-L1 ligand expression on CD4⁺ NKT cells significantly decreased in the third trimester. In contrast, LAG-3 and Galectin-3 expressions remained stable across all subsets and trimesters. For the TIGIT/CD226 axis, CD226 expression was significantly higher in CD4⁺ NKT cells in the third trimester and in non-pregnant women. The two ligands CD112 and CD155 were consistently lower on CD4⁻ NKT cells across all groups. The activating receptor NKG2D was significantly higher on CD4⁻ NKT cells in all examined cohorts. These findings suggest that CD4⁺ NKT cells tend towards a more tolerogenic phenotype, while CD4⁻ NKT cells maintain a balanced cytotoxic potential with reduced immunoregulation function. The dynamic regulation of immune checkpoints on NKT cell subsets, particularly the downregulation of PD-L1 and CD226 in late pregnancy, highlights their fine-tuned role in balancing maternal–fetal immune tolerance with readiness for parturition. Full article

(This article belongs to the Special Issue Molecular Pathology of the Placenta in Pregnancy Complications)

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11 pages, 739 KiB

Open AccessArticle

Clonal Hematopoiesis and Outcomes After High-Dose Chemotherapy and Autologous Stem Cell Transplantation in Patients with AML, Myeloma, and Lymphoma

by Corinne Natalie Schmid, Katharina Sponagel, Ulrike Bacher, Katja Seipel, Naomi Porret, Gertrud Wiedemann, Michèle Hoffmann, Michael Daskalakis and Thomas Pabst

Int. J. Mol. Sci. 2025, 26(16), 8021; https://doi.org/10.3390/ijms26168021 - 19 Aug 2025

Abstract

Autologous stem cell transplantation (ASCT) after high-dose chemo-therapy (HDCT) is an option of consolidation therapy in patients with AML, lymphoma, or myeloma. Clonal hematopoiesis (CH) is a premalignant state, associated with an increased risk of hematological cancer. The incidence of CH in patients [...] Read more.

Autologous stem cell transplantation (ASCT) after high-dose chemo-therapy (HDCT) is an option of consolidation therapy in patients with AML, lymphoma, or myeloma. Clonal hematopoiesis (CH) is a premalignant state, associated with an increased risk of hematological cancer. The incidence of CH in patients with AML, myeloma, and lymphoma and its effect on the outcome after HDCT/ASCT remain poorly studied. Here we screened 142 patients treated with HDCT/ASCT between 2002 and 2021 at Bern University Hospital for somatic gene mutations in ASXL1, DNMT3A, JAK2, TET2, and TP53. CH-associated somatic gene mutations were detected in 14/31 AML patients (45%), 13/64 myeloma patients (20%), and 9/47 lymphoma patients (19%). Clinical characteristics, treatment modalities, and responses to treatment were similar in patients with and without CH. Patients with CH-associated gene mutations had higher relapse rates and reduced progression free survival, most evident in lymphoma patients (p = 0.007). Overall survival tended to be shorter in lymphoma patients with CH-associated mutations (p = 0.078), whereas this was not observed in AML and myeloma patients. Survival in lymphoma patients with CH was inferior, which may have an impact on post-transplant surveillance strategies in the future. In contrast, survival outcomes were not associated significantly with CH in AML and myeloma patients in our study. Longer follow-ups and larger cohorts will be needed to validate our observations. Full article

(This article belongs to the Special Issue Hematologic Malignancies: From Molecular Pathology to Novel Therapeutics)

16 pages, 4715 KiB

Open AccessArticle

Comparative Metabolomics Reveals Phosphine-Induced Metabolic Disruptions in Planococcus citri (Risso)

by Junbeom Lee, Soo-Jung Suh, Bong-Su Kim and Dae-Weon Lee

Int. J. Mol. Sci. 2025, 26(16), 8020; https://doi.org/10.3390/ijms26168020 - 19 Aug 2025

Abstract

Phosphine (PH₃) is a fumigant often used to control insect pests, but its metabolic effects on insect physiology remain unclear. In this study, a comparative metabolomics analysis was performed to elucidate the physiological metabolic pathways affected by PH₃ exposure in [...] Read more.

Phosphine (PH₃) is a fumigant often used to control insect pests, but its metabolic effects on insect physiology remain unclear. In this study, a comparative metabolomics analysis was performed to elucidate the physiological metabolic pathways affected by PH₃ exposure in Planococcus citri, and significant changes in the metabolic profiles induced by PH₃ treatment were identified. Principal component analysis and correlation analysis revealed different metabolic changes, and a total of 45 metabolites were identified and mapped to metabolic pathways using the KEGG database. PH₃ exposure inhibited energy metabolism by down-regulating riboflavin and flavin adenine dinucleotide, which are important cofactors in oxidative phosphorylation and reactive oxygen species generation. In addition, purine and pyrimidine metabolism, essential for nucleotide synthesis and cellular energy homeostasis, were also suppressed. Notably, lipid metabolism was significantly altered, and the juvenile hormone biosynthesis pathway was down-regulated. These results suggest that PH₃ inhibits electron transport chain activity, induces oxidative stress, and disrupts lipid homeostasis. This study enhances our understanding of the potential biomarkers of PH₃ exposure, the metabolic processes involved, and the resistance mechanisms that pests may develop in response to such exposure. Full article

(This article belongs to the Section Molecular Endocrinology and Metabolism)

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43 pages, 11428 KiB

Open AccessReview

MXene-Based Electrocatalysts for Water Splitting: Material Design, Surface Modulation, and Catalytic Performance

by Mohammad R. Thalji, Farzaneh Mahmoudi, Leonidas G. Bachas and Chinho Park

Int. J. Mol. Sci. 2025, 26(16), 8019; https://doi.org/10.3390/ijms26168019 - 19 Aug 2025

Abstract

Developing efficient and sustainable hydrogen production technologies is critical for advancing the global clean energy transition. This review highlights recent progress in the design, synthesis, and electrocatalytic applications of MXene-based materials for electrochemical water splitting. It discusses the fundamental mechanisms of the hydrogen [...] Read more.

Developing efficient and sustainable hydrogen production technologies is critical for advancing the global clean energy transition. This review highlights recent progress in the design, synthesis, and electrocatalytic applications of MXene-based materials for electrochemical water splitting. It discusses the fundamental mechanisms of the hydrogen evolution reaction (HER) and oxygen evolution reaction (OER), and the structure–function relationships that govern electrocatalytic behavior. Emphasis is placed on the intrinsic structural and surface properties of MXenes, such as their layered architecture and tunable surface chemistry, which render them promising candidates for electrocatalysis. Despite these advantages, several practical limitations hinder their full potential, including oxidation susceptibility, restacking, and a limited number of active sites. Several studies have addressed these challenges using diverse engineering strategies, such as heteroatom doping; surface functionalization; and constructing MXene-based composites with metal chalcogenides, oxides, phosphides, and conductive polymers. These modifications have significantly improved catalytic activity, charge transfer kinetics, and long-term operational stability under various electrochemical conditions. Finally, this review outlines key knowledge gaps and emerging research directions, including defect engineering, single-atom integration, and system-level design, to accelerate the development of MXene-based electrocatalysts for sustainable hydrogen production. Full article

(This article belongs to the Special Issue Cutting-Edge Innovations in Molecular and Nanoscale Engineering for Energy Applications)

13 pages, 804 KiB

Open AccessCommunication

Preparturient Oral Selenitetriglycerides Supplementation Elevates Erythrocyte Glutathione Peroxidase Activity and Modulates Hepatic TNF-α, PPAR-α, and PPAR-δ mRNA in Postparturient Holstein–Friesian Cows

by Katarzyna Żarczyńska, Katarzyna Różańska, Paweł Brym and Dawid Tobolski

Int. J. Mol. Sci. 2025, 26(16), 8018; https://doi.org/10.3390/ijms26168018 - 19 Aug 2025

Abstract

The transition period in dairy cows, spanning late pregnancy and early lactation, is associated with substantial metabolic and immunological challenges, leading to increased oxidative stress and inflammation. Selenium (Se), particularly in organic forms, supports antioxidant defenses, immune function, and metabolic regulation. This study [...] Read more.

The transition period in dairy cows, spanning late pregnancy and early lactation, is associated with substantial metabolic and immunological challenges, leading to increased oxidative stress and inflammation. Selenium (Se), particularly in organic forms, supports antioxidant defenses, immune function, and metabolic regulation. This study investigated the effects of supplementing periparturient Holstein–Friesian cows with orally administered selenitetriglycerides (0.5 mg Se/kg body weight/day starting 12 days before the expected calving date and continuing until parturition) on antioxidant enzyme activity and on the hepatic expression of key inflammatory and metabolic genes. Serum selenium concentrations and erythrocyte glutathione peroxidase (GSH-Px) activity were assessed before and after parturition, and hepatic gene expression levels of tumor necrosis factor alpha (TNF-α), peroxisome proliferator-activated receptor alpha (PPAR-α) and delta (PPAR-δ) were assessed 24 h and 7 days postpartum. Supplemented cows showed significantly elevated serum Se levels and increased GSH-Px activity, reflecting improved antioxidant capacity. Moreover, hepatic expression of TNF-α and PPAR-δ was significantly reduced postpartum in the supplemented group, whereas PPAR-α expression remained stable. These findings indicate that selenitetriglycerides effectively enhance antioxidant defenses, moderate inflammation, and stabilize metabolic pathways during the periparturient phase, potentially reducing postpartum metabolic disorders and improving dairy-cow health. Full article

(This article belongs to the Special Issue The Role of Metabolism-Related Pathways in Animal Growth, Development, Disease and Behavior)

22 pages, 1378 KiB

Open AccessArticle

Evaluation of the In Vitro Blood–Brain Barrier Transport of Ferula persica L. Bioactive Compounds

by Pouya Mohammadnezhad, Alberto Valdés, Melis Cokdinleyen, Jose A. Mendiola and Alejandro Cifuentes

Int. J. Mol. Sci. 2025, 26(16), 8017; https://doi.org/10.3390/ijms26168017 - 19 Aug 2025

Abstract

Species of the Ferula genus are known for their traditional medicinal applications against diverse illnesses. Our previous study was the first to suggest the cholinesterase inhibitory activity of Ferula persica L. However, the neuroprotective efficacy of therapeutic molecules is often limited by their [...] Read more.

Species of the Ferula genus are known for their traditional medicinal applications against diverse illnesses. Our previous study was the first to suggest the cholinesterase inhibitory activity of Ferula persica L. However, the neuroprotective efficacy of therapeutic molecules is often limited by their ability to cross the blood–brain barrier (BBB) and reach the brain. In the present study, the BBB permeability of the main molecules present in the aerial parts and roots of F. persica L. extracted under optimum conditions was assessed using two well-established methods: the parallel artificial membrane permeability assay (PAMPA) and the HBMEC cell culture in vitro model. The results demonstrated a high permeability of several neuroprotective compounds, such as apigenin, diosmetin, and α-cyperone. Additionally, the neuroprotective potential of F. persica extracts was evaluated using SH-SY5Y neuron-like cells exposed to different insults, including oxidative stress (H₂O₂), excitotoxicity (L-glutamate), and Aβ1-42 peptide toxicity. However, none of the obtained extracts provided significant protection. This study highlights the importance of in vitro cell culture models for a better understanding of BBB permeability mechanisms and reports the tentative identification of newly formed sulfated metabolites derived from the metabolism of ferulic acid, apigenin, and diosmetin by HBMEC cells. Full article

(This article belongs to the Special Issue IJMS 25th Anniversary: Updates and Progress on Bioactives and Nutraceuticals)

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30 pages, 4096 KiB

Open AccessReview

New Frontiers in 3D Printing Using Biocompatible Polymers

by Nagireddy Poluri, Jacob Carter, John Grasso, Walter Miller, Matthew Leinbach, Frederick Durant, Riley Benbrook, Assa John, Allan Wang and Xiao Hu

Int. J. Mol. Sci. 2025, 26(16), 8016; https://doi.org/10.3390/ijms26168016 - 19 Aug 2025

Abstract

Biocompatible polymers have emerged as essential materials in medical 3D printing, enabling the fabrication of scaffolds, tissue constructs, drug delivery systems, and biosensors for applications in and on the human body. This review aims to provide a comprehensive overview of the current state [...] Read more.

Biocompatible polymers have emerged as essential materials in medical 3D printing, enabling the fabrication of scaffolds, tissue constructs, drug delivery systems, and biosensors for applications in and on the human body. This review aims to provide a comprehensive overview of the current state of 3D-printable biocompatible polymers and their composites, with an emphasis on their processing methods, properties, and biomedical uses. The scope of this work includes both natural and synthetic biocompatible polymers, polymer–nanocomposite systems, and bioinks that do not require photo initiators. The relevant literature was critically examined to classify materials by type, evaluate their compatibility with major 3D printing techniques such as stereolithography, selective laser sintering, and fused deposition modeling, and assess their performance in various medical applications. Key findings highlight that reinforced polymer composites, tailored surface chemistries, and hybrid printing strategies significantly expand the range of functional, customizable, and affordable biomedical devices. This review concludes by discussing present-day applications and emerging trends, underscoring that 3D-printable biocompatible polymers are rapidly transitioning from research to clinical practice, offering transformative potential for patient-specific healthcare solutions. Full article

(This article belongs to the Special Issue Latest Review Papers in Macromolecules 2025)

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29 pages, 3388 KiB

Open AccessArticle

A Dual-Template Molecularly Imprinted Polymer to Inhibit Quorum Sensing Molecules: Theoretical Design, Optimized Synthesis, Physicochemical Characterization and Preliminary Microbiological Analysis

by Khonzisizwe Somandi, Tama S. Mwale, Monika Sobiech, Dorota Klejn, Gillian D. Mahumane, Joanna Giebułtowicz, Sandy van Vuuren, Yahya E. Choonara and Piotr Luliński

Int. J. Mol. Sci. 2025, 26(16), 8015; https://doi.org/10.3390/ijms26168015 - 19 Aug 2025

Abstract

Molecularly imprinted polymers (MIPs) have emerged as promising materials for selectively targeting biomolecules, including quorum sensing autoinducers that regulate bacterial communication and biofilm formation. In this study, both single-template and dual-template strategies were employed to design and synthesize MIPs capable of capturing autoinducer-2 [...] Read more.

Molecularly imprinted polymers (MIPs) have emerged as promising materials for selectively targeting biomolecules, including quorum sensing autoinducers that regulate bacterial communication and biofilm formation. In this study, both single-template and dual-template strategies were employed to design and synthesize MIPs capable of capturing autoinducer-2 analogs using (3R,4S)-tetrahydro-3,4-furandiol (T1) or (R/S) 2,2-dimethyl-1,3-dioxolane-4-methanol (T2) as the templates. This approach offers translational potential of a complementary or non-antibiotic strategy to conventional antimicrobial therapies in mitigating biofilm-associated infections. Computational modeling guided the rational selection of functional monomers, predicting favorable interaction energies (ΔE_C up to −135 kcal·mol⁻¹) and optimal hydrogen-bonding patterns to enhance template–polymer affinity. The synthesized MIPs were characterized using spectroscopic and microscopic techniques to confirm imprinting efficiency and structural integrity. The adsorption capacity measurements demonstrated higher adsorption capacity and selectivity of MIPs compared to non-imprinted polymers, with the highest selectivity equal to 3.36 for T1 and 3.14 for T2 on MIPs fabricated from methacrylic acid. Preliminary microbiological evaluations using Chromobacterium violaceum ATCC 12472 reveal that the MIPs prepared from 2-hydroxyethyl methacrylate effectively inhibited violacein production by up to 78.2% at 5.0 mg·mL⁻¹, consistent with quorum sensing interference. These findings highlight the feasibility of employing molecular imprinting to target autoinducer-2 analogs, introducing a novel synthetic strategy for disrupting bacterial communication. This further suggests that molecular imprinting can be leveraged to develop potent quorum-sensing inhibitors, an approach that offers translational potential as an alternative to conventional antimicrobial strategies to mitigate biofilm-associated infections. Full article

(This article belongs to the Section Materials Science)

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19 pages, 1877 KiB

Open AccessArticle

PAM-Independent Cas12a Detection of Specific LAMP Products by Targeting Amplicon Loops

by Konstantin G. Ptitsyn, Leonid K. Kurbatov, Svetlana A. Khmeleva, Daria D. Morozova, Olga S. Timoshenko, Elena V. Suprun, Sergey P. Radko and Andrey V. Lisitsa

Int. J. Mol. Sci. 2025, 26(16), 8014; https://doi.org/10.3390/ijms26168014 - 19 Aug 2025

Abstract

A straightforward approach is suggested to selectively recognize specific products of loop-mediated isothermal amplification (LAMP) with the Cas12a nuclease without a need for a protospacer adjacent motif (PAM) in the sequence of LAMP amplicons (LAMPlicons). This strategy is based on the presence of [...] Read more.

A straightforward approach is suggested to selectively recognize specific products of loop-mediated isothermal amplification (LAMP) with the Cas12a nuclease without a need for a protospacer adjacent motif (PAM) in the sequence of LAMP amplicons (LAMPlicons). This strategy is based on the presence of single-stranded DNA loops in LAMPlicons and the ability of Cas12a to be trans-activated via the binding of guide RNA (gRNA) to single-stranded DNA in the absence of PAM. The approach feasibility is demonstrated on Clavibacter species—multiple bacterial plant pathogens that cause harmful diseases in agriculturally important plants. For Clavibacter species, the detection sensitivity of the developed PAM-independent LAMP/Cas12a system was determined by that of LAMP. The overall detection selectivity was enhanced by the Cas12a analysis of LAMPlicons. It was shown that the LAMP/Cas12a detection system can be fine-tuned by carefully designing gRNA to selectively distinguish C. sepedonicus from other Clavibacter species based on single nucleotide substitutions in the targeted LAMPlicon loop. The suggested loop-based Cas12a analysis of LAMPlicons was compatible with the format of a single test tube assay with the option of naked-eye detection. The findings broaden the palette of approaches to designing PAM-independent LAMP/Cas12a detection systems with potential for on-site testing. Full article

(This article belongs to the Special Issue CRISPR/Cas Systems and Genome Editing—3rd Edition)

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29 pages, 2957 KiB

Open AccessArticle

Bioactive Properties of Selected European Phellinus Species: A Comprehensive Study

by Grzegorz Świderski, Monika Kalinowska, Ewa Zapora, Marek Wołkowycki, Marcin Stocki, Ewa Ciszkowicz, Aleksandra Bocian, Marcin Jaromin, Mirosław Tyrka, Katarzyna Lecka-Szlachta, Elżbieta Wołejko, Urszula Wydro, Małgorzata Pawłowska, Paweł Golianek, Małgorzata Zawadzka, Qëndrim Ramshaj, Carolina Elena Girometta and Mitko Karadelev

Int. J. Mol. Sci. 2025, 26(16), 8013; https://doi.org/10.3390/ijms26168013 - 19 Aug 2025

Abstract

This study conducted a multi-directional evaluation of the chemical potential and biological properties of selected European fungal species of the genus Phellinus. We investigated 30 samples belonging to 22 Phellinus species. Fruiting bodies were collected, among other specimens, in the Białowieża Forest [...] Read more.

This study conducted a multi-directional evaluation of the chemical potential and biological properties of selected European fungal species of the genus Phellinus. We investigated 30 samples belonging to 22 Phellinus species. Fruiting bodies were collected, among other specimens, in the Białowieża Forest (Poland); Village Kozhle (North Macedonia); Estremadura, Sesimbra, and Lagoa de Albufeira (Portugal); Zlatari close to Prishtina (Kosovo); and Spoleto and the Bosco Siro Negri State Nature Reserve (Italy). Morphological identification of the collected fungi was carried out, and genetic tests were performed to confirm the identity of the collected specimens. Methanol extracts for biological activity tests were prepared. Screening of antimicrobial activity of 30 methanolic extracts was performed on strains of bacteria (Klebsiella pneumoniae, Pseudomonas aeruginosa, Staphylococcus aureus, Staphylococcus epidermidis, and Kocuria rhizophila) and fungi (Candida albicans). Antioxidant activity tests (DPPH and ABTS) were also performed. The three most biologically active fungi species were selected (Phellinus igniarius, Fomitiporia robusta, and Porodaedalea pini) for further research. The chemical composition of the extracts was determined using GC-MS analysis. Thermal decomposition studies and spectroscopic analysis of the dry fruiting bodies were performed. The extracts were tested for their antimicrobial activity against antibiotic-resistant bacteria. Cytotoxic activity was also tested. Full article

(This article belongs to the Special Issue Molecular Insight into Plant Bioactive Compounds)

15 pages, 1279 KiB

Open AccessArticle

25-Hydroxycholesterol Induces Intrinsic Apoptosis via Mitochondrial Pathway in BE(2)-C Human Neuroblastoma Cells

by Jaesung Kim, Koanhoi Kim, Dongha Park, Seong-Kug Eo, Bo-Ae Lee and Yonghae Son

Int. J. Mol. Sci. 2025, 26(16), 8012; https://doi.org/10.3390/ijms26168012 - 19 Aug 2025

Abstract

An oxysterol, 25-Hydroxycholesterol (25OHChol), is produced through cholesterol oxidation and is involved in various cellular processes, including apoptosis. However, the precise mechanisms underlying 25OHChol-induced apoptosis in neuroblastoma cells remain unclear. The aim of this study was to elucidate the detailed molecular mechanisms by [...] Read more.

An oxysterol, 25-Hydroxycholesterol (25OHChol), is produced through cholesterol oxidation and is involved in various cellular processes, including apoptosis. However, the precise mechanisms underlying 25OHChol-induced apoptosis in neuroblastoma cells remain unclear. The aim of this study was to elucidate the detailed molecular mechanisms by which 25OHChol induces apoptosis in human neuroblastoma cells. This study explores the apoptotic effects of 25OHChol and the associated signaling pathways in BE(2)-C cells, a widely used human neuroblastoma cell model for neuronal differentiation and cancer research. To evaluate the cytotoxicity of 25OHChol, cell viability was assessed using the CCK-8 assay, which demonstrated a concentration-dependent decline, indicating a potential induction of cell death. Morphological changes characteristic of apoptosis, such as nuclear condensation and fragmentation, were confirmed via DAPI staining. Additionally, Annexin V/PI flow cytometry analysis revealed an increase in late apoptotic cell populations, further corroborating apoptosis induction. To investigate the molecular mechanisms, we analyzed the expression of Bcl-2 family proteins via Western blotting. The results showed an elevated Bax/Bcl-2 ratio, suggesting activation of the intrinsic mitochondrial apoptotic pathway. This was further supported by a reduction in mitochondrial membrane potential (MMP), as measured by flow cytometry. Increased caspase-9 and caspase-3/7 activity provided additional evidence for caspase-mediated apoptosis. Moreover, treatment with the pan-caspase inhibitor Z-VAD-FMK led to a dose-dependent increase in cell viability, confirming the essential role of caspases in 25OHChol-induced apoptosis. In conclusion, this study demonstrates that 25OHChol triggers apoptosis in BE(2)-C neuroblastoma cells through activation of the intrinsic mitochondrial apoptotic pathway. These findings provide new insights into the cytotoxic effects of 25OHChol and its potential role in neuroblastoma cell death. Full article

(This article belongs to the Section Molecular Neurobiology)

64 pages, 1331 KiB

Open AccessReview

Physical Activity, Exerkines, and Their Role in Cancer Cachexia

by Jan Bilski, Aleksandra Szlachcic, Agata Ptak-Belowska and Tomasz Brzozowski

Int. J. Mol. Sci. 2025, 26(16), 8011; https://doi.org/10.3390/ijms26168011 - 19 Aug 2025

Abstract

Cancer-associated cachexia is a multifaceted wasting syndrome characterized by progressive loss of skeletal muscle mass, systemic inflammation, and metabolic dysfunction and is particularly prevalent in gastrointestinal cancers. Physical activity has emerged as a promising non-pharmacological intervention capable of attenuating key drivers of cachexia. [...] Read more.

Cancer-associated cachexia is a multifaceted wasting syndrome characterized by progressive loss of skeletal muscle mass, systemic inflammation, and metabolic dysfunction and is particularly prevalent in gastrointestinal cancers. Physical activity has emerged as a promising non-pharmacological intervention capable of attenuating key drivers of cachexia. Exercise modulates inflammatory signaling (e.g., IL-6/STAT3 and TNF-α/NF-κB), enhances anabolic pathways (e.g., IGF-1/Akt/mTOR), and preserves lean body mass and functional capacity. Exercise-induced signaling molecules, known as exerkines, are key mediators of these benefits, which are released during physical activity and act in an autocrine, paracrine, and endocrine manner. However, many of these molecules also exhibit context-dependent effects. While they exert protective, anti-inflammatory, or anabolic actions when transiently elevated after exercise, the same molecules may contribute to cachexia pathogenesis when chronically secreted by tumors or in systemic disease states. The biological effects of a given factor depend on its origin, timing, concentration, and physiological milieu. This review presents recent evidence from clinical and experimental studies to elucidate how physical activity and exerkines may be harnessed to mitigate cancer cachexia, with particular emphasis on gastrointestinal malignancies and their unique metabolic challenges. Full article

(This article belongs to the Special Issue Molecular Mediators of Exercise: Cross-Organ Communication Promoting Health and Combating Disease)

20 pages, 5412 KiB

Open AccessArticle

Site-Specific Changes in Cytosine Methylation in Promoters of the Genes Encoding the Membrane Subunits of Succinate Dehydrogenase During Germination of Maize Seeds

by Dmitry N. Fedorin, Alexander T. Eprintsev and Abir U. Igamberdiev

Int. J. Mol. Sci. 2025, 26(16), 8010; https://doi.org/10.3390/ijms26168010 - 19 Aug 2025

Abstract

The cytosine methylation status of symmetric and asymmetric sites of promoters of the genes encoding the membrane-bound subunits C and D of succinate dehydrogenase (SDH) was assessed during the germination of maize (Zea mays L.) seeds, when the stored lipids were utilized [...] Read more.

The cytosine methylation status of symmetric and asymmetric sites of promoters of the genes encoding the membrane-bound subunits C and D of succinate dehydrogenase (SDH) was assessed during the germination of maize (Zea mays L.) seeds, when the stored lipids were utilized and the glyoxylate cycle produced succinate. The results of bisulfite sequencing of the promoters of Sdh genes in maize scutella allowed us to determine the cytosine methylation status in the CG, CNG, and CNN sites. The observed site-specific changes in the cytosine methylation status of the Sdh3-1 and Sdh3-2 genes encoding the SDH subunit C indicate an important role in controlling their transcriptional activity. In contrast, no marked changes were observed in the methylation of promoters of the Sdh4 gene, encoding SDH subunit D. The analysis of changes in the activity of the CG, CNG, and CNN DNA methyltransferases revealed the redistribution of activity between CG, CNG, and CNN DNA methyltransferases toward an increase in the proportion of CG DNA methyltransferases. The locus-specific methylation dynamics of SDH membrane subunit promoters during maize germination have been demonstrated. It is concluded that the changes in the cytosine methylation status may play a role in the regulation of the expression of the Sdh genes in the course of the conversion of succinate formed in the glyoxylate cycle. Full article

(This article belongs to the Section Molecular Biology)

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15 pages, 3437 KiB

Open AccessArticle

Adaptation of the Protocol for the Isolation of Biotinylated Protein Complexes for Drosophila melanogaster Tissues

by Igor A. Shokodko, Rustam H. Ziganshin and Nadezhda E. Vorobyeva

Int. J. Mol. Sci. 2025, 26(16), 8009; https://doi.org/10.3390/ijms26168009 - 19 Aug 2025

Abstract

Proximity biotinylation, which utilizes various biotin ligating enzymes (BioID, TurboID, etc.), is widely used as a powerful tool for identifying novel protein–protein interactions. However, this method has a significant limitation: the use of streptavidin on beads for enriching biotinylated proteins often results in [...] Read more.

Proximity biotinylation, which utilizes various biotin ligating enzymes (BioID, TurboID, etc.), is widely used as a powerful tool for identifying novel protein–protein interactions. However, this method has a significant limitation: the use of streptavidin on beads for enriching biotinylated proteins often results in a high background of peptides from streptavidin itself, which interferes with identification by peptide mass fingerprinting. This limitation makes it practically impossible to study samples containing a small amount of material, such as individual insect tissues. In this study, we compared different precipitation and elution conditions for the purification of biotinylated proteins from protein extracts of Drosophila melanogaster S2 cells. We found that biotinylated proteins can be purified using anti-biotin antibodies, although with lower efficiency than streptavidin-based resin. We also demonstrated that protease-resistant streptavidin (prS), previously tested in mammalian cells, can be used effectively to purify biotinylated proteins from tissues of D. melanogaster. In our experiments, prS showed precipitation efficiency comparable to regular streptavidin but generated a lower background in peptide fingerprinting. To further demonstrate the applicability of prS for studying protein–protein interactions in D. melanogaster tissues, we carried out experiments to identify interaction partners of the ecdysone receptor (EcR) in D. melanogaster ovarian tissue using TurboID-based proximity biotinylation. As a result, EcR was found to interact with both previously described and novel protein partners in Drosophila ovaries. Full article

(This article belongs to the Special Issue Drosophila: A Versatile Model in Biology and Medicine—2nd Edition)

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24 pages, 3501 KiB

Open AccessArticle

Gestational and Lactation Exposure to Perfluorohexanoic Acid Results in Sex-Specific Changes in the Cerebellum in Mice

by Elizabeth C. Plunk, Navya Navnith, Hannah Swan, Linh Le, Matthew McCall, Marissa E. Sobolewski and Ania K. Majewska

Int. J. Mol. Sci. 2025, 26(16), 8008; https://doi.org/10.3390/ijms26168008 - 19 Aug 2025

Abstract

Currently regulated per- and polyfluoroalkyl substances (PFAS) have been associated with immune, endocrine, and neurotoxicity following gestational exposures. As a result, industries have effectively replaced them with next-generation PFAS, including perfluorohexanoic acid (PFHxA). PFHxA is increasingly found in the serum of pregnant women [...] Read more.

Currently regulated per- and polyfluoroalkyl substances (PFAS) have been associated with immune, endocrine, and neurotoxicity following gestational exposures. As a result, industries have effectively replaced them with next-generation PFAS, including perfluorohexanoic acid (PFHxA). PFHxA is increasingly found in the serum of pregnant women and in breast milk, and adult human post-mortem studies indicate that PFHxA is found in the brain, with the highest concentrations in the cerebellum and hypothalamus. Despite evidence of gestational, lactational, and nervous system exposure to PFHxA, developmental neurotoxicity (DNT) testing in mammals has not been conducted. For DNT evaluation, we exposed pregnant C57Bl/6J mice daily from gestational day 0 through postnatal day (P) 21 to two PFHxA exposure levels (a lower (0.32 mg/kg of body weight (bw), or higher (50 mg/kg of bw) dose of PFHxA)) or ddH₂O using treat-based administration. Given the high PFHxA levels in the cerebellum in post-mortem studies and the cerebellum’s protracted developmental window, we assessed acute transcriptional dysregulation and cellular morphology in this brain region on the last day of exposure at P21. Using bulk-RNA sequencing, we found that PFHxA exposure had subtle effects on transcripts related to neurons and glia, with females having a greater number of dysregulated transcripts than males. Using immunohistochemistry, we found that Purkinje cell linear frequency was increased in specific lobules in the higher-exposure group and that microglial morphology underwent subtle changes in specific cerebellar layers in the lower-exposure group in both sexes. Together these data suggest that PFHxA exposure may have lobule-specific impacts on the development of both neurons and glia in the cerebellum, highlighting the importance of studying the neurotoxicity of PFHxA in both sexes. Full article

(This article belongs to the Special Issue Toxicity Mechanism of Emerging Pollutants: 2nd Edition)

27 pages, 6117 KiB

Open AccessArticle

Clinical and Proteomic Associations of SARS-CoV-2 Infection and COVID-19 Vaccination in Multimorbid Patients: A Cross-Sectional Observational Study

by Anett Hudák, Aladár Pettko-Szandtner, Annamária Letoha and Tamás Letoha

Int. J. Mol. Sci. 2025, 26(16), 8007; https://doi.org/10.3390/ijms26168007 - 19 Aug 2025

Abstract

Vaccines played a crucial role in the COVID-19 pandemic, but their long-term biological effects and efficacy in vulnerable populations remain under intensive investigation. This study assessed clinical outcomes, comorbidities, and systemic biomarker and proteomic profiles in 366 multimorbid patients, stratified into four groups [...] Read more.

Vaccines played a crucial role in the COVID-19 pandemic, but their long-term biological effects and efficacy in vulnerable populations remain under intensive investigation. This study assessed clinical outcomes, comorbidities, and systemic biomarker and proteomic profiles in 366 multimorbid patients, stratified into four groups based on SARS-CoV-2 infection and vaccination status (COV+ vac+, COV+ vac−, COV− vac+, COV− vac−). Clinical and laboratory data, including comorbidities and relevant biomarkers, were collected. Proteomic analysis using mass spectrometry was performed to identify molecular changes associated with infection and vaccination. Statistical analyses examined associations between clinical status, biomarkers, and patient outcomes. As most participants received mRNA-based vaccines, the results primarily reflect responses to spike protein-expressing platforms. Biomarkers of cardiac and renal stress—namely proBNP and carbamide—were elevated in vaccinated individuals. Five deaths occurred in the COV+ vac+ group and two in the COV+ vac− group, most of which were attributed to exacerbations of pre-existing chronic diseases rather than to COVID-19 pneumonia. Protection against breakthrough infections waned over time, particularly beyond 200 days post-vaccination. Mass spectrometry identified proteins such as actin, fibrinogen chains, and SAA2 as potential diagnostic targets. Although the cross-sectional observational design limits the ability to draw causal inferences, the observed waning immunity and potential systemic alterations in vaccinated multimorbid patients highlight the importance of longitudinal follow-up to guide tailored immunization strategies and post-vaccination monitoring in high-risk groups. Full article

(This article belongs to the Special Issue Coronavirus Disease (COVID-19): Pathophysiology (6th Edition))

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