International Journal of Molecular Sciences

15 pages, 2772 KiB

Open AccessArticle

Molecular Detection of Human Papillomavirus (HPV) and Other Sexually Transmitted Pathogens in Cervical and Self-Collected Specimens

by Chiara Giubbi, Marianna Martinelli, Michelle Rizza, Maria Letizia Di Meo, Ruth Chinyere Njoku, Federica Perdoni, Giulio Mannarà, Rosario Musumeci, Robert Fruscio, Fabio Landoni and Clementina Elvezia Cocuzza

Int. J. Mol. Sci. 2025, 26(3), 1296; https://doi.org/10.3390/ijms26031296 (registering DOI) - 3 Feb 2025

Abstract

This study investigated the detection of high-risk Human Papillomavirus (hrHPV) and seven other pathogens associated with sexually transmitted infections (STIs) in matched clinician-collected cervical samples and self-taken vaginal and urine specimens collected from 342 asymptomatic women referred to colposcopy to evaluate (i) the [...] Read more.

This study investigated the detection of high-risk Human Papillomavirus (hrHPV) and seven other pathogens associated with sexually transmitted infections (STIs) in matched clinician-collected cervical samples and self-taken vaginal and urine specimens collected from 342 asymptomatic women referred to colposcopy to evaluate (i) the concordance in the molecular detection of investigated pathogen in three different sample types; (ii) the analytical sensitivity and specificity of STIs detection on self-samples; and (iii) the distribution of STIs in hrHPV-positive and hrHPV-negative women. Pathogens detection was performed using Anyplex™II HR and Anyplex™II STI-7e, respectively. Good/substantial agreement was observed between cervical and self-taken samples in detecting hrHPV (κ = 0.870 and κ = 0.773 for vaginal and urine). The agreement between cervical and self-taken samples for detecting STIs was found to be significant (κ = 0.779 and κ = 0.738 for vaginal and urine), with almost perfect agreement between urine and vaginal specimens (κ = 0.899). The positivity rate for all investigated STIs was found to be higher in hrHPV-positive compared to hrHPV-negative women. In conclusion, self-sampling proved to be a valid alternative to cervical samples to detect hrHPV and STIs, but further studies are required to evaluate the role of STI coinfections in cervical lesions development and progression. Full article

(This article belongs to the Special Issue New Strategies to Improve the Detection of Human Papillomavirus (HPV) Persisting Infections)

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49 pages, 819 KiB

Open AccessReview

Exploring the Immunological Aspects and Treatments of Recurrent Pregnancy Loss and Recurrent Implantation Failure

by Jenny Valentina Garmendia, Claudia Valentina De Sanctis, Marián Hajdúch and Juan Bautista De Sanctis

Int. J. Mol. Sci. 2025, 26(3), 1295; https://doi.org/10.3390/ijms26031295 (registering DOI) - 3 Feb 2025

Abstract

Recurrent pregnancy loss (RPL) is defined as the occurrence of two or more consecutive pregnancy losses before 24 weeks of gestation. It affects 3–5% of women who are attempting to conceive. RPL can stem from a variety of causes and is frequently associated [...] Read more.

Recurrent pregnancy loss (RPL) is defined as the occurrence of two or more consecutive pregnancy losses before 24 weeks of gestation. It affects 3–5% of women who are attempting to conceive. RPL can stem from a variety of causes and is frequently associated with psychological distress and a diminished quality of life. By contrast, recurrent implantation failure (RIF) refers to the inability to achieve a successful pregnancy after three or more high-quality embryo transfers or at least two instances of egg donation. RIF shares several causative factors with RPL. The immunological underpinnings of these conditions involve alterations in uterine NK cells, reductions in M2 macrophages and myeloid-derived suppressor cells, an increased Th1/Th2 ratio, a decreased Treg/Th17 ratio, the presence of shared ≥3 HLA alleles between partners, and autoimmune disorders. Various therapeutic approaches have been employed to address these immunological concerns, achieving varying degrees of success, although some therapies remain contentious within the medical community. This review intends to explore the immunological factors implicated in RPL and RIF and to analyze the immunological treatments employed for these conditions, which may include steroids, intravenous immunoglobulins, calcineurin inhibitors, anti-TNF antibodies, intralipid infusions, granulocyte colony-stimulating factor, and lymphocyte immunotherapy. Full article

(This article belongs to the Section Molecular Immunology)

16 pages, 1749 KiB

Open AccessReview

Biological Significance and Therapeutic Promise of Programmed Ribosomal Frameshifting

by Miora Bruna Marielle Ramamonjiharisoa and Sen Liu

Int. J. Mol. Sci. 2025, 26(3), 1294; https://doi.org/10.3390/ijms26031294 (registering DOI) - 3 Feb 2025

Abstract

Programmed Ribosomal Frameshifting (PRF) is a mechanism that alters the mRNA reading frame during translation, resulting in the production of out-of-frame proteins. PRF plays crucial roles in maintaining cellular homeostasis and contributes significantly to disease pathogenesis, particularly in viral infections. Notably, PRF can [...] Read more.

Programmed Ribosomal Frameshifting (PRF) is a mechanism that alters the mRNA reading frame during translation, resulting in the production of out-of-frame proteins. PRF plays crucial roles in maintaining cellular homeostasis and contributes significantly to disease pathogenesis, particularly in viral infections. Notably, PRF can induce immune responses in the SARS-CoV-2 mRNA vaccine, further extending its biological significance. These multiple aspects of PRF highlight its potential as a therapeutic target. Since PRF efficiency can be modulated by cellular factors, its expression or silencing is context-dependent. Therefore, a deeper understanding of PRF is essential for harnessing its therapeutic potential. This review explores PRF biological significance in disease and homeostasis. Such knowledge would serve as a foundation to advance therapeutic strategies targeting PRF modulation, especially in viral infections and vaccine development. Full article

(This article belongs to the Special Issue Molecular Insights into Macromolecules Structure, Function, and Regulation: 2nd Edition)

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20 pages, 7992 KiB

Open AccessArticle

The Potential of Single-Transcription Factor Gene Expression by RT-qPCR for Subtyping Small Cell Lung Cancer

by Albert Iñañez, Raúl del Rey-Vergara, Fabricio Quimis, Pedro Rocha, Miguel Galindo, Sílvia Menéndez, Laura Masfarré, Ignacio Sánchez, Marina Carpes, Carlos Martínez, Sandra Pérez-Buira, Federico Rojo, Ana Rovira and Edurne Arriola

Int. J. Mol. Sci. 2025, 26(3), 1293; https://doi.org/10.3390/ijms26031293 - 3 Feb 2025

Abstract

Complex RNA-seq signatures involving the transcription factors ASCL1, NEUROD1, and POU2F3 classify Small Cell Lung Cancer (SCLC) into four subtypes: SCLC-A, SCLC-N, SCLC-P, and SCLC-I (triple negative or inflamed). Preliminary studies suggest that identifying these subtypes can guide targeted therapies and [...] Read more.

Complex RNA-seq signatures involving the transcription factors ASCL1, NEUROD1, and POU2F3 classify Small Cell Lung Cancer (SCLC) into four subtypes: SCLC-A, SCLC-N, SCLC-P, and SCLC-I (triple negative or inflamed). Preliminary studies suggest that identifying these subtypes can guide targeted therapies and potentially improve outcomes. This study aims to evaluate whether the expression levels of these three key transcription factors can effectively classify SCLC subtypes, comparable to the use of individual antibodies in immunohistochemical (IHC) analysis of formalin-fixed, paraffin-embedded (FFPE) tumor samples. We analyzed preclinical models of increasing complexity, including eleven human and five mouse SCLC cell lines, six patient-derived xenografts (PDXs), and two circulating tumor cell (CTC)-derived xenografts (CDXs) generated in our laboratory. RT-qPCR conditions were established to detect the expression levels of ASCL1, NEUROD1, and POU2F3. Additionally, protein-level analysis was performed using Western blot for cell lines and IHC for FFPE samples of PDX and CDX tumors, following our experience with patient tumor samples from the CANTABRICO trial (NCT04712903). We found that the analyzed SCLC cell line models predominantly expressed ASCL1, NEUROD1, and POU2F3, or showed no expression, as identified by RT-qPCR, consistently matching the previously assigned subtypes for each cell line. The classification of PDX and CDX models demonstrated consistency between RT-qPCR and IHC analyses of the transcription factors. Our results show that single-gene analysis by RT-qPCR from FFPE-extracted RNA simplifies SCLC subtype classification. This approach provides a cost-effective alternative to IHC staining or expensive multi-gene RNA sequencing panels, making SCLC subtyping more accessible for both preclinical research and clinical applications. Full article

(This article belongs to the Special Issue Recent Trends in Experimental Models for Cancer Research)

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23 pages, 2236 KiB

Open AccessArticle

Development and Characterization of Magnetic Nanoemulsion-Based Senolytic Peptides for Osteoarthritis Treatment

by Camelia-Mihaela Zara-Danceanu, Jenifer García-Fernández, Dumitru-Daniel Herea, Daniel Gherca, Irene de Francisco Carrera, Luminita Labusca and Maria de la Fuente

Int. J. Mol. Sci. 2025, 26(3), 1292; https://doi.org/10.3390/ijms26031292 - 3 Feb 2025

Abstract

The formulation and characterization of a novel nanoemulsion (NE) delivery system for senomodulator peptides aimed at enhancing the treatment of osteoarthritis (OA) are reported, in combination with magnetic nanoparticles (MNPs), for improving targeted delivery and traceability. Osteoarthritis, a prevalent degenerative joint disease associated [...] Read more.

The formulation and characterization of a novel nanoemulsion (NE) delivery system for senomodulator peptides aimed at enhancing the treatment of osteoarthritis (OA) are reported, in combination with magnetic nanoparticles (MNPs), for improving targeted delivery and traceability. Osteoarthritis, a prevalent degenerative joint disease associated with aging, is currently not effectively treated by disease-modifying therapies, posing a consistent health burden on individuals and healthcare systems worldwide. Existing treatments, such as nonsteroidal anti-inflammatory drugs and intra-articular injections, are limited by inadequate local drug concentrations and rapid clearance, often necessitating costly joint replacement. Lipid-based NE composed of biocompatible and biodegradable vitamin E and sphingomyelin, associated with the senolytic peptide NE:TUB1, is able to target senescent cells implicated in OA progression. Improved cellular retention and therapeutic effects of the associated TUB1 peptide, compared to its free form, have been demonstrated, suggesting a significant enhancement in therapeutic potential. The incorporation of MNPs to obtain NE:TUB1-MNP formulations offers the advantage of being traceable in vivo through clinically available imaging technologies, with the potential to enhance targeting capabilities through magnetic guidance. The characterization of NE:TUB1-MNPs involved the assessment of their physical and chemical properties, interaction with cells, cytotoxicity profile, and nanoparticle uptake in vitro using human primary adipose-derived stem cells. NE and NE:TUB1-MNP are shown to be stable, non-toxic, and capable of efficient intracellular uptake. The inclusion of MNPs not only supports cell viability and proliferation but also facilitates medium and long-term product traceability within joints, offering a promising approach for localized treatment. The enhanced anti-senescent role of NE:TUB1-MNP formulations are highlighted, suggesting their potential utility in mitigating OA progression and possibly other degenerative diseases. In conclusion, the study presents a novel therapeutic approach for OA, NE:TUB1-MNPs, leveraging the synergistic effects of peptide-functionalized nanoemulsions and magnetic nanoparticles to improve targeted delivery and therapeutic outcomes. This innovative formulation could pave the way for new treatments for OA and other joint-related conditions, offering significant advancements in regenerative medicine. Full article

(This article belongs to the Special Issue Molecular Mechanisms and Therapeutic Approaches to Osteoarthritis)

18 pages, 34503 KiB

Open AccessArticle

Corydalis Tuber Extract Alleviates Atopic Dermatitis: Transcriptomics-Based Mechanism Prediction and In Vitro/In Vivo Studies

by Seong-Eun Jin, Chang-Seob Seo, Woo-Young Jeon, Yong-Jin Oh, Hyeun-Kyoo Shin and Hyekyung Ha

Int. J. Mol. Sci. 2025, 26(3), 1291; https://doi.org/10.3390/ijms26031291 - 3 Feb 2025

Abstract

Atopic dermatitis (AD) is a common inflammatory skin disease characterized by recurrent eczema and chronic itching, affecting a significant portion of the global population. This study investigated the effects of Corydalis Tuber 70% ethanol extract (CTE) on tumor necrosis factor-α- and interferon-γ (TI)-stimulated [...] Read more.

Atopic dermatitis (AD) is a common inflammatory skin disease characterized by recurrent eczema and chronic itching, affecting a significant portion of the global population. This study investigated the effects of Corydalis Tuber 70% ethanol extract (CTE) on tumor necrosis factor-α- and interferon-γ (TI)-stimulated human keratinocytes (HaCaT) and a house dust mite-induced AD mouse model, elucidating its mechanism via transcriptome analysis. A total of 13 compounds, including columbamine, corydaline, dehydrocorydaline, and glaucine, were identified in CTE using ultra performance liquid chromatography-tandem mass spectrometry. CTE downregulated pathways related to cytokine signaling and chemokine receptors in TI-stimulated HaCaT cells. It significantly inhibited C-C motif chemokine ligand (CCL)5, CCL17, and CCL22 levels by blocking the Janus kinase-signal transducers and activators of transcription and nuclear factor kappa-light-chain-enhancer of activated B cells pathways. In the AD mouse model, topical CTE significantly decreased dermatitis scores, epidermal thickening, and inflammatory cell infiltration. Plasma levels of histamine, immunoglobulin E, CCL17, CCL22, corticosterone, and cortisol were reduced. Lesions showed decreased thymic stromal lymphopoietin, CD4⁺ T cells, interleukin-4, and intercellular adhesion molecule-1 expression. The findings demonstrate that CTE alleviates AD by modulating inflammatory mediators, cytokines, and chemokines, reducing inflammatory cell infiltration, and alleviating stress-related factors. Full article

(This article belongs to the Special Issue Molecular Research on Skin Inflammation)

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17 pages, 7358 KiB

Open AccessArticle

Disrupted Mitochondrial Dynamics Impair Corneal Epithelial Healing in Neurotrophic Keratopathy

by Mengyi Jin, Zeyu Liu, Ruize Shi, Ya Deng, Jingwei Lin, Yuting Zhang, Lexin Lin, Yanzi Wang, Yunyi Shi, Cheng Li and Zuguo Liu

Int. J. Mol. Sci. 2025, 26(3), 1290; https://doi.org/10.3390/ijms26031290 - 3 Feb 2025

Abstract

Neurotrophic keratopathy (NK) is a degenerative corneal disease characterized by impaired corneal sensitivity and epithelial repair that is often linked to sensory nerve dysfunction. To establish a clinically relevant model and explore the mechanisms underlying NK pathogenesis, we developed a novel mouse model [...] Read more.

Neurotrophic keratopathy (NK) is a degenerative corneal disease characterized by impaired corneal sensitivity and epithelial repair that is often linked to sensory nerve dysfunction. To establish a clinically relevant model and explore the mechanisms underlying NK pathogenesis, we developed a novel mouse model through partial transection of the ciliary nerve. This approach mimics the progressive nature of NK, reproducing key clinical features such as corneal epithelial defects, reduced sensitivity, diminished tear secretion, and delayed wound healing. Using this model, we investigated how disruptions in mitochondrial dynamics contribute to corneal epithelial dysfunction and impaired repair in NK. Our findings revealed substantial disruptions in mitochondrial dynamics, including reduced expression of fusion proteins (OPA1), downregulation of fission regulators (FIS1 and MFF), and impaired mitochondrial transport, as evidenced by decreased expression of Rhot1 and Kif5b. Additionally, the downregulation of mitophagy-related genes (Pink1 and Prkn) contributed to the accumulation of dysfunctional mitochondria, leading to DNA damage and impaired corneal epithelial repair. These mitochondrial abnormalities were accompanied by increased γH2AX staining, indicative of DNA double-strand breaks and cellular stress. This study highlights the pivotal role of mitochondrial dynamics in corneal epithelial health and repair, suggesting that therapeutic strategies aimed at restoring mitochondrial function, enhancing mitophagy, and mitigating oxidative stress may offer promising avenues for treating NK. Full article

(This article belongs to the Section Molecular Neurobiology)

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13 pages, 1364 KiB

Open AccessReview

The Human Superficial Fascia: A Narrative Review

by Caterina Fede, Claudia Clair, Carmelo Pirri, Lucia Petrelli, Xiaoxiao Zhao, Yunfeng Sun, Veronica Macchi and Carla Stecco

Int. J. Mol. Sci. 2025, 26(3), 1289; https://doi.org/10.3390/ijms26031289 - 3 Feb 2025

Abstract

In recent years, the interest in the comprehension of the fasciae has significantly grown, together with the necessity of finding a consensus for a terminology of the fasciae in the research and clinical fields. Furthermore, it is becoming necessary to categorize the various [...] Read more.

In recent years, the interest in the comprehension of the fasciae has significantly grown, together with the necessity of finding a consensus for a terminology of the fasciae in the research and clinical fields. Furthermore, it is becoming necessary to categorize the various types of fascia (superficial, deep, visceral, neural) since they possess different anatomical characteristics, and are implicated in different pathophysiological pathways. While in the past we have described the deep/muscular fascia, the aim of this work is to summarize and catalog the information relating to the human superficial fascia (thickness, cellular end extracellular matrix component, innervation, vascularization). Full article

(This article belongs to the Special Issue Fascial Anatomy and Histology: Advances in Molecular Biology)

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19 pages, 3204 KiB

Open AccessArticle

Microbiota Co-Metabolism Alterations Precede Changes in the Host Metabolism in the Early Stages of Diet-Induced MASLD in Wistar Rats

by María Martín-Grau, Pilar Casanova, Laura Moreno-Morcillo, José Manuel Morales, Vannina G. Marrachelli and Daniel Monleón

Int. J. Mol. Sci. 2025, 26(3), 1288; https://doi.org/10.3390/ijms26031288 - 2 Feb 2025

Abstract

Metabolic-dysfunction-associated steatotic liver disease (MASLD) affects around 30% of the global population. The sexual dimorphism and gut microbiota play an important role in the early development of MASLD. The main objective of this research was to investigate metabolic changes during the early subclinical [...] Read more.

Metabolic-dysfunction-associated steatotic liver disease (MASLD) affects around 30% of the global population. The sexual dimorphism and gut microbiota play an important role in the early development of MASLD. The main objective of this research was to investigate metabolic changes during the early subclinical MASLD progression, for identifying the sequence of events and evaluating the impact of sexual dimorphism and the microbiota on the initial stages of MASLD development. Male and female Wistar rats 18 weeks old were randomly divided into different groups and fed a chow diet or a 45% high-fat diet for 21 weeks. Every three weeks, samples of serum, urine, and faeces were collected and studied by metabolomics. Furthermore, the liver was analysed at the endpoint. In addition, the gut microbiota was analysed from faecal samples over time using 16S rRNA gene-targeted group-specific primers. Our results revealed that three weeks on an HFD reduced the bacterial diversity in the faecal microbiota of Wistar rats, accompanied by changes in the faecal and urine metabolome. The HFD-induced alterations in microbiota-related co-metabolites in the liver, blood, urine, and faeces indicate a significant role of host–microbiota co-metabolism changes in the early stages of MASLD. In this study, we provide a comprehensive longitudinal analysis, detailing the sequence of events in the early development of MASLD. Our findings suggest that alterations in the gut microbiota diversity and co-metabolism occur before changes in host metabolism in the early onset of liver steatosis, a subclinical phase of MASLD. Full article

(This article belongs to the Special Issue Research Progress of Metabolomics in Health and Disease)

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21 pages, 3292 KiB

Open AccessArticle

Exploring the Impact of Microgravity on Gene Expression: Dysregulated Pathways and Candidate Repurposed Drugs

by Karina Galčenko, Marilena M. Bourdakou and George M. Spyrou

Int. J. Mol. Sci. 2025, 26(3), 1287; https://doi.org/10.3390/ijms26031287 - 2 Feb 2025

Abstract

Space exploration has progressed from contemporary discoveries to current endeavors, such as space tourism and Mars missions. As human activity in space accelerates, understanding the physiological effects of microgravity on the human body is becoming increasingly critical. This study analyzes transcriptomic data from [...] Read more.

Space exploration has progressed from contemporary discoveries to current endeavors, such as space tourism and Mars missions. As human activity in space accelerates, understanding the physiological effects of microgravity on the human body is becoming increasingly critical. This study analyzes transcriptomic data from human cell lines exposed to microgravity, investigates its effects on gene expression, and identifies potential therapeutic interventions for health challenges posed by spaceflight. Our analysis identified five under-expressed genes (DNPH1, EXOSC5, L3MBTL2, LGALS3BP, SPRYD4) and six over-expressed genes (CSGALNACT2, CSNK2A2, HIPK1, MBNL2, PHF21A, RAP1A), all of which exhibited distinct expression patterns in response to microgravity. Enrichment analysis highlighted significant biological functions influenced by these conditions, while in silico drug repurposing identified potential modulators that could counteract these changes. This study introduces a novel approach to addressing health challenges during space missions by repurposing existing drugs and identifies specific genes and pathways as potential biomarkers for microgravity effects on human health. Our findings represent the first systematic effort to repurpose drugs for spaceflight, establishing a foundation for the development of targeted therapies for astronauts. Future research should aim to validate these findings in authentic space environments and explore broader biological impacts. Full article

(This article belongs to the Special Issue Exploring the Molecular Basis of Cellular Responses to Microgravity-Induced Oxidative Stress)

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21 pages, 4007 KiB

Open AccessArticle

Unveiling the Multitarget Potential of a Rare Caffeoyl Ester from Artemisia capillaris for Diabetes Mellitus: An Integrated In Vitro and In Silico Study

by Md. Nurul Islam, Manh Tuan Ha, Byung-Sun Min, Jae Sue Choi and Hyun Ah Jung

Int. J. Mol. Sci. 2025, 26(3), 1286; https://doi.org/10.3390/ijms26031286 - 2 Feb 2025

Abstract

As a part of our ongoing search for bioactive constituents of Artemisia capillaris, we isolated 4-O-caffeoyl-2-C-methyl-d-threonic acid (PPT-14). This is a rare caffeic acid ester derivative that is reported here for the first time in the [...] Read more.

As a part of our ongoing search for bioactive constituents of Artemisia capillaris, we isolated 4-O-caffeoyl-2-C-methyl-d-threonic acid (PPT-14). This is a rare caffeic acid ester derivative that is reported here for the first time in the Artemisia species, which is the third occurrence in any plant species worldwide. In this study, we evaluated the anti-diabetic potential of PPT-14 using in vitro and in silico approaches. PPT-14 demonstrated significant inhibitory activity against two crucial enzymes linked to diabetes progression and complications: protein tyrosine phosphatase 1B (PTP1B) and aldose reductase (AR). These had IC₅₀ values of 64.92 and 19.50 µM, respectively. Additionally, PPT-14 exhibited free radical scavenging activity with 2,2-diphenyl-2-picrylhydrazyl (IC₅₀ 14.46 µM). Molecular docking and 200 ns molecular dynamics simulations confirmed that there were stable binding interactions with the key residues of PTP1B and AR, highlighting strong affinity and dynamic stability. Pharmacokinetic analyses revealed favorable water solubility, adherence to Lipinski’s Rule of Five, and minimal interactions with cytochrome P450 enzymes, indicating the drug-like potential of PPT-14. Toxicity studies confirmed its safety profile, showing no genotoxicity, hepatotoxicity, or significant toxicity risks, with an acceptable oral LD₅₀ value of 2.984 mol/kg. These findings suggest that PPT-14 could be a promising multitarget lead compound for ameliorating diabetes and its associated complications. Full article

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

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18 pages, 3432 KiB

Open AccessArticle

Rational Identification of Ritonavir as IL-20 Receptor A Ligand Endowed with Antiproliferative Properties in Breast Cancer Cells

by Valentina Maggisano, Adriana Gargano, Jessica Maiuolo, Francesco Ortuso, Francesca De Amicis, Stefano Alcaro and Stefania Bulotta

Int. J. Mol. Sci. 2025, 26(3), 1285; https://doi.org/10.3390/ijms26031285 - 2 Feb 2025

Abstract

Targeting the tumor microenvironment (TME) is an attractive strategy for developing new drugs with anticancer activity against triple-negative breast cancer (TNBC). Interleukins (ILs) are key players in the TME cytokine network promoting cancer progression. Recent studies have highlighted the involvement of IL-20 receptor [...] Read more.

Targeting the tumor microenvironment (TME) is an attractive strategy for developing new drugs with anticancer activity against triple-negative breast cancer (TNBC). Interleukins (ILs) are key players in the TME cytokine network promoting cancer progression. Recent studies have highlighted the involvement of IL-20 receptor subunit alpha (IL-20RA) signalling in several cancers, including BC, in which IL-20RA is highly expressed, correlating with poor prognosis and influencing tumoral characteristics such as proliferation, cell death, invasiveness, and TME activity. Therefore, elucidating the role of the IL-20RA signalling pathway could form the basis for developing new therapeutic strategies. This study aimed to identify selective bioactive ligands able to affect IL-20RA activity. Virtual screening of over 310,000 compounds from both the DrugBank and ZINC15 databases identified four potential hit compounds tested for their anticancer activity against TNBC in vitro cell lines. Notably, Ritonavir, a well-known Human Immunodeficiency Virus Type 1 (HIV-1) protease inhibitor, significantly inhibited cell proliferation (about 40% at 50 µM, p < 0.001). IL-20 preincubation counteracted Ritonavir’s cytostatic effect while IL-20RA knockdown restored proliferation in Ritonavir-treated TNBC cells. In conclusion, these findings demonstrated that Ritonavir reduced TNBC cell proliferation through IL-20RA activity modulation, suggesting its potential repurposing as a therapeutic agent for TNBC management. Full article

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

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18 pages, 4957 KiB

Open AccessArticle

Skin Telocyte Secretome as Conditioned Medium Prevents Profibrotic Differentiation of Skin Fibroblasts into Myofibroblasts

by Irene Rosa, Bianca Saveria Fioretto, Elena Andreucci, Alessio Biagioni, Eloisa Romano and Mirko Manetti

Int. J. Mol. Sci. 2025, 26(3), 1284; https://doi.org/10.3390/ijms26031284 - 2 Feb 2025

Abstract

Telocytes (TCs) are distinctive cells widely localized in the stromal compartment of several human organs, including the skin. By means of their peculiar prolongations named telopodes, skin TCs are organized in networks interconnected with a variety of adjacent cells, being thus supposed to [...] Read more.

Telocytes (TCs) are distinctive cells widely localized in the stromal compartment of several human organs, including the skin. By means of their peculiar prolongations named telopodes, skin TCs are organized in networks interconnected with a variety of adjacent cells, being thus supposed to take part in skin homeostasis through both cell-to-cell contacts and the release of extracellular vesicles. A disarrangement/loss of the TC network was shown in human fibrotic skin as well as in the murine model of bleomycin-induced cutaneous fibrosis, but whether such TC alterations may represent just a consequence or a trigger of the fibrotic process still remains to be clarified. Thus, we investigated the effects of skin TC secretome as conditioned medium (TC-CM) on the transition of skin fibroblasts into myofibroblasts promoted by the master profibrotic cytokine transforming growth factor β1 (TGFβ1). Primary cultures of both adult human skin TCs and fibroblasts were obtained by means of immunomagnetic cell separation. Nanoparticle tracking analysis was carried out to measure extracellular vesicles in TC-CM. The combination of multiple morphological, gene/protein expression, and functional assessments demonstrated that TC-CM was able to significantly prevent TGFβ1-induced fibroblast-to-myofibroblast transition. TC-CM did not influence cell viability, while it effectively inhibited TGFβ1-induced fibroblast proliferation, migration, and morphological changes. Indeed, TC-CM was able to reduce TGFβ1-mediated skin fibroblast phenotypic and functional differentiation into myofibroblasts, as shown by a significant decrease in FAP, ACTA2, COL1A1, COL1A2, FN1, and CTGF gene expression, α-smooth muscle actin, N-cadherin, COL1A1, and FN-EDA protein levels, and collagen gel matrix contraction. Furthermore, TC-CM significantly lowered TGFβ1-mediated ERK1/2 signaling pathway activation. This in vitro study proves for the first time that TCs may play an important role in skin homeostasis through the prevention of fibroblast-to-myofibroblast transition via paracrine mechanisms and affords the necessary basis to investigate in the future the feasibility of TC secretome as an innovative antifibrotic therapeutic tool. Full article

(This article belongs to the Section Molecular Biology)

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24 pages, 1436 KiB

Open AccessReview

Impact of High-Temperature Stress on Maize Seed Setting: Cellular and Molecular Insights of Thermotolerance

by Zhaoyi Fan, Haoqi Song, Mengyuan Qi, Mengqing Wang, Yunfeng Bai, Yuhui Sun and Haidong Yu

Int. J. Mol. Sci. 2025, 26(3), 1283; https://doi.org/10.3390/ijms26031283 - 2 Feb 2025

Abstract

Global warming poses a significant threat to crop production and food security, with maize (Zay mays L.) particularly vulnerable to high-temperature stress (HTS). This review explores the detrimental impacts of elevated temperatures on maize development across various growth stages, analyzed within the [...] Read more.

Global warming poses a significant threat to crop production and food security, with maize (Zay mays L.) particularly vulnerable to high-temperature stress (HTS). This review explores the detrimental impacts of elevated temperatures on maize development across various growth stages, analyzed within the source–sink framework, with a particular focus on seed setting and yield reduction. It provides a broad analysis of maize cellular and molecular responses to HTS, highlighting the key roles of plant hormone abscisic acid (ABA) signaling, calcium signaling, chloroplast, and the DNA damage repair (DDR) system in maize. HTS disrupts ABA signaling pathways, impairing stomatal regulation and reducing water-use efficiency, while calcium signaling orchestrates stress responses by activating heat shock proteins and other protective mechanisms. Chloroplasts, as central to photosynthesis, are particularly sensitive to HTS, often exhibiting photosystem II damage and chlorophyll degradation. Recent studies also highlight the significance of the DDR system, with genes like ZmRAD51C playing crucial roles in maintaining genomic stability during reproductive organ development. DNA damage under HTS conditions emerges as a key factor contributing to reduced seed set, although the precise molecular mechanisms remain to be fully elucidated. Furthermore, the review examines cutting-edge genetic improvement strategies, aimed at developing thermotolerant maize cultivars. These recent research advances underscore the need for further investigation into the molecular basis of thermotolerance and open the door for future advancements in breeding thermotolerant crops. Full article

(This article belongs to the Special Issue Advance in Plant Abiotic Stress: 2nd Edition)

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40 pages, 8055 KiB

Open AccessArticle

Exertional Exhaustion (Post-Exertional Malaise, PEM) Evaluated by the Effects of Exercise on Cerebrospinal Fluid Metabolomics–Lipidomics and Serine Pathway in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome

by James N. Baraniuk

Int. J. Mol. Sci. 2025, 26(3), 1282; https://doi.org/10.3390/ijms26031282 - 1 Feb 2025

Abstract

Post-exertional malaise (PEM) is a defining condition of myalgic encephalomyelitis (ME/CFS). The concept requires that a provocation causes disabling limitation of cognitive and functional effort (“fatigue”) that does not respond to rest. Cerebrospinal fluid was examined as a proxy for brain metabolite and [...] Read more.

Post-exertional malaise (PEM) is a defining condition of myalgic encephalomyelitis (ME/CFS). The concept requires that a provocation causes disabling limitation of cognitive and functional effort (“fatigue”) that does not respond to rest. Cerebrospinal fluid was examined as a proxy for brain metabolite and lipid flux and to provide objective evidence of pathophysiological dysfunction. Two cohorts of ME/CFS and sedentary control subjects had lumbar punctures at baseline (non-exercise) or after submaximal exercise (post-exercise). Cerebrospinal fluid metabolites and lipids were quantified by targeted Biocrates mass spectrometry methods. Significant differences between ME/CFS and control, non-exercise vs. post-exercise, and by gender were examined by multivariate general linear regression and Bayesian regression methods. Differences were found at baseline between ME/CFS and control groups indicating disease-related pathologies, and between non-exercise and post-exercise groups implicating PEM-related pathologies. A new, novel finding was elevated serine and its derivatives sarcosine and phospholipids with a decrease in 5-methyltetrahydrofolate (5MTHF), which suggests general dysfunction of folate and one-carbon metabolism in ME/CFS. Exercise led to consumption of lipids in ME/CFS and controls while metabolites were consumed in ME/CFS but generated in controls. In general, the frequentist and Bayesian analyses generated complementary but not identical sets of analytes that matched the metabolic modules and pathway analysis. Cerebrospinal fluid is unique because it samples the choroid plexus, brain interstitial fluid, and cells of the brain parenchyma. The quantitative outcomes were placed into the context of the cell danger response hypothesis to explain shifts in serine and phospholipid synthesis; folate and one-carbon metabolism that affect sarcosine, creatine, purines, and thymidylate; aromatic and anaplerotic amino acids; glucose, TCA cycle, trans-aconitate, and coenzyme A in energy metabolism; and vitamin activities that may be altered by exertion. The metabolic and phospholipid profiles suggest the additional hypothesis that white matter dysfunction may contribute to the cognitive dysfunction in ME/CFS. Full article

(This article belongs to the Special Issue Molecular Pathologies and Treatment for Myalgic Encephalomyelitis/Chronic Fatigue Syndrome)

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14 pages, 6246 KiB

Open AccessArticle

Subgenomic RNA Detection in SARS-CoV-2 Assessing Replication and Inactivation Through Serial Passages, RT-qPCR, and Electron Microscopy

by Talita da Silva França, Juliana Fernandes Amorim da Silva, Gabriella Christine Neves da Silva, Barbara Oliveira dos Santos, Stephanie Almeida Silva, José Henrique Resende Linhares, Marcos Alexandre Nunes da Silva, Debora Ferreira Barreto-Vieira, Vanessa Salete de Paula, Liliane Monteiro de Morais, Renata Tourinho Santos and Gisela Freitas Trindade

Int. J. Mol. Sci. 2025, 26(3), 1281; https://doi.org/10.3390/ijms26031281 - 1 Feb 2025

Abstract

Subgenomic RNAs (sgRNAs) are potential markers of active SARS-CoV-2 replication, serving as templates for the synthesis of structural and accessory proteins in infectious viral particles. This study aimed to use RT-qPCR to quantify sgRNA and negative RNA intermediates, assessing viral replication in virus [...] Read more.

Subgenomic RNAs (sgRNAs) are potential markers of active SARS-CoV-2 replication, serving as templates for the synthesis of structural and accessory proteins in infectious viral particles. This study aimed to use RT-qPCR to quantify sgRNA and negative RNA intermediates, assessing viral replication in virus samples inactivated by β-propiolactone (βPL). Inactivated viruses subjected to five blind serial passages (BSs) were amplified by RT-qPCR using primers to target the envelope (ENV) and nucleoproteins (N1 and N2) of genomic genes, subgenomic envelope RNA (sgENV), and intermediate envelope RNA (ENV-). All positive controls showed consistent viral titers across passages (10 log10 copies/mL in N1/N2 and 11 log10 copies/mL in ENV) during BSs. Inactivated viral samples for ENV and ENV- targets ranged from 11.34 log10 copies/mL in BS1 to 11.20 log10 copies/mL in BS5. The sgENV was no longer detected in the inactivated SARS-CoV-2 samples after the second passage, suggesting successful inactivation. Replication kinetics showed consistent profiles for N1/N2, ENV, and ENV- targets in the first three post-infection hours (pih) and maintained approximately 5 log10 copies/mL at 1 pih, 2 pih, and 3 pih. A sharp exponential increase in the viral titer was observed from 24 pih onwards, peaking at 11.64 log10 copies/mL at 48 pih. Transmission electron microscopy confirmed viral particles only in cells infected with active SARS-CoV-2. These results support the use of sgRNA as a reliable marker for SARS-CoV-2 replication, especially in distinguishing between active replication and non-viable particles and in the development of diagnostic and therapeutic strategies. Full article

(This article belongs to the Section Molecular Biology)

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15 pages, 3011 KiB

Open AccessArticle

Connexin 43 Affects Pulmonary Artery Reactivity via Changes in Nitric Oxide Production and Influences Proliferative and Migratory Responses in Mouse Pulmonary Artery Fibroblasts

by Saad Wali, Abdmajid Hwej, David J. Welsh, Kathryn Wilson, Simon Kennedy and Yvonne Dempsie

Int. J. Mol. Sci. 2025, 26(3), 1280; https://doi.org/10.3390/ijms26031280 - 1 Feb 2025

Abstract

Pulmonary hypertension (PH) is a complex condition characterized by pulmonary artery constriction and vascular remodeling. Connexin 43 (Cx43), involved in cellular communication, may play a role in PH development. Cx43 heterozygous (Cx43^+/−) mice show partial protection against hypoxia-induced pulmonary remodeling, with [...] Read more.

Pulmonary hypertension (PH) is a complex condition characterized by pulmonary artery constriction and vascular remodeling. Connexin 43 (Cx43), involved in cellular communication, may play a role in PH development. Cx43 heterozygous (Cx43^+/−) mice show partial protection against hypoxia-induced pulmonary remodeling, with prior research highlighting its role in rat pulmonary artery fibroblast (PAF) proliferation and migration. However, inhibiting Cx43 may compromise nitric oxide (NO)-mediated vascular relaxation. This study evaluated the effects of Cx43 on mouse PAF (MPAF) proliferation, migration, NO-dependent and independent pulmonary vascular relaxation, and NO synthesis. Proliferation and migration were assessed in Cx43^+/− MPAFs under normoxic and hypoxic conditions. Vascular responses were analyzed in intra-lobar pulmonary artery rings with acetylcholine (ACh), SNAP, and U46619, while NO production was measured in lung tissue. Both genetic knockdown and pharmacological inhibition of Cx43 significantly reduced serum-induced proliferation but not migration under normoxia, while ^37,43Gap27 inhibited hypoxia-induced proliferation and migration. The effects of genetic knockdown and pharmacological inhibition of Cx43 on vascular reactivity were also investigated. NO-dependent and independent relaxations and NO production were reduced in Cx43^+/− mice by ^37,43Gap27. In conclusion, while Cx43 inhibition may protect against PAF proliferation and migration, it could also impair pulmonary vascular relaxation, at least in part through a reduction in NO signaling. Further studies are needed to fully understand the mechanisms by which Cx43 influences NO signaling. Full article

(This article belongs to the Special Issue Pulmonary Hypertension: Pre-clinical Research on New Molecular Targets and Therapeutic Strategies)

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19 pages, 4925 KiB

Open AccessReview

The Pivotal Role of LACTB in the Process of Cancer Development

by Minghui Zhang, Bowen Wu and Jinke Gu

Int. J. Mol. Sci. 2025, 26(3), 1279; https://doi.org/10.3390/ijms26031279 - 1 Feb 2025

Abstract

The mitochondrial serine β-lactamase-like protein LACTB has emerged as a critical regulator in cancer biology, distinguished by its unique structural and functional attributes. Defined by its conserved penicillin-binding proteins and β-lactamases (PBP-βLs) domain and SXXK catalytic motif, LACTB demonstrates properties distinct from its [...] Read more.

The mitochondrial serine β-lactamase-like protein LACTB has emerged as a critical regulator in cancer biology, distinguished by its unique structural and functional attributes. Defined by its conserved penicillin-binding proteins and β-lactamases (PBP-βLs) domain and SXXK catalytic motif, LACTB demonstrates properties distinct from its prokaryotic homologs, including the ability to polymerize into filaments. These structural characteristics enable LACTB to modulate mitochondrial organization and enzymatic activity, influencing lipid metabolism and indirectly affecting cellular proliferation. Importantly, the expression and functional roles of LACTB exhibit cancer-type-specific variation, underscoring its dual function as both a tumor suppressor and an oncogene. Decreased LACTB expression is associated with poor clinical outcomes in cancers such as breast cancer, lung cancer, and colorectal cancer, while specific mutations and regulatory mechanisms have been linked to its oncogenic activity in osteosarcoma and pancreatic adenocarcinoma. Mechanistically, LACTB regulates key processes in cancer progression, including mitochondrial dynamics, epithelial–mesenchymal transition (EMT), and cell death pathways. This duality highlights LACTB as a promising therapeutic target and underscores its relevance in advancing precision oncology strategies. This review provides a comprehensive analysis of expression level, structure–function relationships, and the diverse roles of LACTB in oncogenesis, underscoring its promise as a focal point for precision cancer therapies. Full article

(This article belongs to the Section Biochemistry)

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17 pages, 1424 KiB

Open AccessArticle

miR-30d Levels Predict Re-Hospitalization in Patients with Acute Cardiogenic Pulmonary Edema: A Preliminary Study

by Giordano Bianchi, Barbara Vizio, Ornella Bosco, Martina Schiavello, Paolo Cagna Vallino, Francesca Rumbolo, Fulvio Morello, Giulio Mengozzi, Giuseppe Montrucchio, Enrico Lupia and Emanuele Pivetta

Int. J. Mol. Sci. 2025, 26(3), 1278; https://doi.org/10.3390/ijms26031278 - 1 Feb 2025

Abstract

Acute cardiogenic pulmonary edema (ACPE) is a common and serious manifestation of heart failure (HF), representing 10–20% of all acute HF admissions. It is associated with elevated in-hospital mortality and high rates of re-hospitalization. MicroRs, like miR-30d, are of particular interest in heart [...] Read more.

Acute cardiogenic pulmonary edema (ACPE) is a common and serious manifestation of heart failure (HF), representing 10–20% of all acute HF admissions. It is associated with elevated in-hospital mortality and high rates of re-hospitalization. MicroRs, like miR-30d, are of particular interest in heart failure due to their regulatory role in gene expression and potential as biomarkers for diagnosing and predicting patient outcomes, especially in high-risk cases such as ACPE. We conducted a cohort study on patients diagnosed with ACPE in the Emergency Department (ED). The circulating levels of miR-30d were analyzed at the time of hospital admission and at one-month follow-up along with other biomarkers. We enrolled 24 ACPE patients and 10 control subjects. Median age was 80.8 years (interquartile range, IQR, 8.2) in ACPE cases, and 78.5 years (IQR 9.8) in controls with a male/female ratio of 2 and 0.66, respectively. In ACPE patients, median cardiac ejection fraction was 42.5%, creatinine 1.63 mg/dl (IQR 1.24), troponin 63.5 ng/dl (58), and NT-proBNP 4243.5 pg/mL (IQR 5846) at ED evaluation. Median concentration of miR30d was 0.81 in controls, and 3.67 and 7.28 in ACPE patients at enrollment time and one month later, respectively. Re-hospitalization occurred in 7 ACPE patients in the following 3 months, and in 9 during the following year. miR-30d had a significant predictive value in assessing the risk of re-hospitalization at both 3 months and 1 year following the initial diagnosis of ACPE, while it did not in assessing the risk of death at 1 year. When compared with the other biomarkers, none of them showed a better accuracy than miR-30d. Our findings suggest that elevated levels of miR-30d are associated with an increased rate of hospital readmission at both 3 months and 1 year after discharge. Larger, multicenter studies will be needed to confirm the validity of circulating miR-30d levels as a potential biomarker useful for risk prediction in ACPE patients and its utility in improving individualized patient care. Full article

(This article belongs to the Special Issue Novel Insights into Molecular Mechanisms of Pulmonary Pathology)

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