International Journal of Molecular Sciences

18 pages, 3145 KB

Open AccessArticle

CRISPR/Cas9-Mediated Targeted Mutagenesis of GmAS1/2 Genes Alters Leaf Shape in Soybean

by Juan Xu, Mengyue Pan, Yu Zhu, Peiguo Wang, Liwei Jiang, Dami Xu, Xinyang Wang, Limiao Chen, Wei Guo, Hongli Yang and Dong Cao

Int. J. Mol. Sci. 2025, 26(19), 9657; https://doi.org/10.3390/ijms26199657 (registering DOI) - 3 Oct 2025

Abstract

ASYMMETRIC LEAVES1 (AS1) and AS2 play essential roles in regulating leaf development in plants. However, their functional roles in soybean remain poorly understood. Here, we identified two members of the soybean AS1 gene family, GmAS1a and GmAS1c, which exhibit high [...] Read more.

ASYMMETRIC LEAVES1 (AS1) and AS2 play essential roles in regulating leaf development in plants. However, their functional roles in soybean remain poorly understood. Here, we identified two members of the soybean AS1 gene family, GmAS1a and GmAS1c, which exhibit high expression levels in stem and leaf tissues. Using the CRISPR/Cas9 system, we targeted four GmAS1 and three GmAS2 genes, generating mutant lines with distinct leaf development phenotypes, including wrinkling (refers to fine lines and creases on the leaf surface, like aged skin texture), curling (describes the inward or outward rolling of leaf edges, deviating from the typical flat shape), and narrow. We found that functional redundancy exists among the four GmAS1 genes in soybean. GmAS1 and GmAS2 cooperatively regulate leaf curling, leaf crinkling phenotypes, and leaf width in soybean, with functional redundancy also observed between these two genes. Transcriptome sequencing analysis of w3 mutant (as1b as1c as1d as2a as2b as2c) identified 1801 differentially expressed genes (DEGs), including 192 transcription factors (TFs). Gene ontology enrichment analysis revealed significant enrichment of DEGs in pathways associated with plant hormone biosynthesis and signal transduction. A detailed examination of the DEGs showed several genes involved in the development of leaf lateral organs, such as KNOX (SHOOT MERISTEMLESS (STM), KNAT1, KNAT2, and KNAT6), LOB (LBD25, LBD30), and ARP5, were down-regulated in w3/WT (wild-type) comparison. CRISPR/Cas9-mediated targeted mutagenesis of the GmAS1/2 genes significantly impairs leaf development and polarity establishment in soybean, providing valuable germplasm resources and a theoretical framework for future studies on leaf morphogenesis. Full article

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

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29 pages, 2059 KB

Open AccessReview

Roles and Applications of Circular RNA in Virus Infection

by Fang Gou, Yanmei Gao, Keke Zhong, Tian Bu, Yinggang Li, Faxiang Li and Rong Yang

Int. J. Mol. Sci. 2025, 26(19), 9656; https://doi.org/10.3390/ijms26199656 (registering DOI) - 3 Oct 2025

Abstract

Circular RNAs (circRNAs) are novel types of covalently closed single-stranded RNA formed by the backsplicing of precursor mRNAs (pre-mRNAs). Recently, circRNAs have been shown to play a crucial role in various diseases, including cancers, cardiovascular and cerebrovascular diseases, and autoimmune diseases. Accumulating evidence [...] Read more.

Circular RNAs (circRNAs) are novel types of covalently closed single-stranded RNA formed by the backsplicing of precursor mRNAs (pre-mRNAs). Recently, circRNAs have been shown to play a crucial role in various diseases, including cancers, cardiovascular and cerebrovascular diseases, and autoimmune diseases. Accumulating evidence has demonstrated that both host-derived and virus-encoded circRNAs play pivotal roles during viral infection, including modulating viral entry, genome replication, latency establishment, and the host antiviral immune responses while simultaneously facilitating viral immune evasion. However, their roles during viral infections and circRNA-host interactions remain to be further investigated. Therefore, this article reviews the key characteristics and biological functions of circRNAs, as well as recent advances in understanding the interactions between circRNAs from different sources and viral infections, which will offer insights for developing therapies targeting virus-associated diseases. Full article

(This article belongs to the Section Molecular Microbiology)

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22 pages, 752 KB

Open AccessArticle

Blood–Brain Barrier Penetration of Novel 4-Trifluoromethyl-Coumarin Hybrids with Antibacterial Properties as Potential Brain Therapeutics in the Context of Spatially Diverse Healthcare Systems

by Paweł Kowalczyk, Dominik Koszelewski, Tomasz Misztal, Michał Szlis, Patrycja Młotkowska, Marcin Gołębiewski, Krzysztof Głowacz, Malwina Kocot, Michał Marczyk, Aleksandra Wypych, Apoloniusz Kurylczyk, Anna Krajewska-Pędzik and Ryszard Ostaszewski

Int. J. Mol. Sci. 2025, 26(19), 9655; https://doi.org/10.3390/ijms26199655 (registering DOI) - 3 Oct 2025

Abstract

Effective treatment of central nervous system (CNS) infections remains a major challenge, as most therapeutic agents do not efficiently cross the blood–brain barrier (BBB) and the blood–cerebrospinal fluid barrier (BCSFB). Coumarin derivatives are of particular interest due to their broad pharmacological activity, favorable [...] Read more.

Effective treatment of central nervous system (CNS) infections remains a major challenge, as most therapeutic agents do not efficiently cross the blood–brain barrier (BBB) and the blood–cerebrospinal fluid barrier (BCSFB). Coumarin derivatives are of particular interest due to their broad pharmacological activity, favorable safety profile, and potential to penetrate biological barriers. Eight novel coumarin-based peptidomimetics functionalized with trifluoromethyl or methyl scaffolds were synthesized and evaluated as antimicrobial agents with the ability to cross the blood–brain barrier. Antimicrobial activity of the investigated compounds was tested against Staphylococcus aureus and multiple Escherichia coli strains (K12, R2, R3, R4) using minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) assays. Cytotoxicity was assessed in vitro in BALB/c-3T3 mouse fibroblasts and αT3-1 pituitary gonadotrope cells using the MTT assay. In vivo studies were performed in sheep to assess transfer of the compounds from blood to cerebrospinal fluid (CSF). All synthesized derivatives demonstrated antimicrobial activity and acceptable cytotoxicity, comparable to those of clinically used antibiotics. CF₃-modified coumarin peptidomimetics show promise as antimicrobial agents with the potential to penetrate the BBB/BCSFB. These findings support further investigation of coumarin-based scaffolds as a platform for the development of novel therapeutics for CNS infections. Full article

(This article belongs to the Special Issue Naturally Occurring Bioactive Compounds, Neuroinflammation and Cognitive Dysfunction: Molecular and Biochemical Mechanisms)

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Graphical abstract

28 pages, 492 KB

Open AccessReview

Inflammatory and Oxidative Biological Profiles in Mental Disorders: Perspectives on Diagnostics and Personalized Therapy

by Izabela Woźny-Rasała and Ewa Alicja Ogłodek

Int. J. Mol. Sci. 2025, 26(19), 9654; https://doi.org/10.3390/ijms26199654 (registering DOI) - 3 Oct 2025

Abstract

Personalized psychiatry represents an innovative therapeutic approach that integrates biological, genetic, and clinical data to optimize the treatment of mental disorders. Laboratory diagnostics play a fundamental role in this process by providing precise biomarkers that characterize pathophysiological mechanisms such as neuroinflammatory processes, oxidative [...] Read more.

Personalized psychiatry represents an innovative therapeutic approach that integrates biological, genetic, and clinical data to optimize the treatment of mental disorders. Laboratory diagnostics play a fundamental role in this process by providing precise biomarkers that characterize pathophysiological mechanisms such as neuroinflammatory processes, oxidative stress, dysfunction of the Hypothalamic–Pituitary–Adrenal (HPA) axis, as well as disturbances in neuroplasticity and neurodegeneration. This article discusses the use of advanced analytical techniques, such as immunoenzymatic assays for pro-inflammatory cytokines (Interleukin-1β- IL-1β; Interleukin-6-IL-6; Interleukin-18-IL-18; and Tumor Necrosis Factor- α - TNF-α). It also emphasizes the role of pharmacogenomic diagnostics in the individualization of psychotropic therapy. Interdisciplinary collaboration between laboratory diagnosticians and clinicians supports the potential for multidimensional analysis of biomarker data in a clinical context, which supports precise patient profiling and monitoring of treatment responses. Despite progress, there are limitations, such as the lack of standardization in measurement methods, insufficient biomarker validation, and limited availability of tests in clinical practice. Development prospects include the integration of multi-marker panels, the use of point-of-care diagnostics, and the implementation of artificial intelligence tools for the analysis of multidimensional data. As a result, laboratory diagnostics are becoming an integral element of personalized psychiatry, enabling a better understanding of the neurobiology of mental disorders and the implementation of more effective therapeutic strategies. Full article

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

19 pages, 3638 KB

Open AccessArticle

Glutaminase Reprogramming in Hepatocellular Carcinoma: Implications for Diagnosis, Prognosis, and Potential as a Novel Therapeutic Target

by Vincent Tambay, Valérie-Ann Raymond, Simon Turcotte and Marc Bilodeau

Int. J. Mol. Sci. 2025, 26(19), 9653; https://doi.org/10.3390/ijms26199653 - 3 Oct 2025

Abstract

Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer, with a poor prognosis due to late diagnosis, limited curative therapies, and underlying liver disease. Glutamine metabolism, a crucial pathway in cancer, remains poorly understood in HCC, which develops in an already metabolically [...] Read more.

Hepatocellular carcinoma (HCC) is the most prevalent primary liver cancer, with a poor prognosis due to late diagnosis, limited curative therapies, and underlying liver disease. Glutamine metabolism, a crucial pathway in cancer, remains poorly understood in HCC, which develops in an already metabolically dynamic organ. This study aimed to characterize glutamine metabolism in HCC. Glutamine metabolism in HCC was explored through in vitro analysis of neoplastic characteristics, experimental hepatocarcinogenesis in C57BL/6 mice, and examination of liver samples from patients with HCC, cirrhosis, and non-diseased liver. The evaluation included metabolite abundance and mRNA/protein expressions. In mice, tumors exhibited hyperactive glutaminolysis compared to adjacent tissue. Notably, glutaminase expression shifted from the liver isoform (GLS2) in normal and cirrhotic livers to the kidney isoform (GLS1) in HCC. In samples from patients, HCC tumors showed overexpression of glutamine synthetase and GLS1 along with a loss of GLS2 expression, providing excellent discrimination of HCC lesions from cirrhotic and normal liver samples. Inhibiting GLS1 with CB-839 significantly impacted glutamine metabolism in HCC cells while showing limited activity on normal hepatocytes. HCC tumors show reprogramming of GLS2 to GLS1, with a concomitant increase in glutamine synthetase. These characteristics can discriminate HCC from cirrhotic and normal liver tissues. Overexpressed GLS1 and loss of GLS2 within tumors convey an unfavorable prognosis in patients with HCC. Pharmacological inhibition of GLS1 in HCC cells successfully harnesses glutamine metabolism, representing an attractive target for novel therapeutic approaches. Full article

(This article belongs to the Special Issue Targeting Cancer Metabolism: From Mechanism to Therapies)

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16 pages, 534 KB

Open AccessReview

The Diagnostic Potential of Urinary Titin Fragment in Neuromuscular Diseases

by Andrea Sipos, Dávid Varga and Endre Pál

Int. J. Mol. Sci. 2025, 26(19), 9652; https://doi.org/10.3390/ijms26199652 - 3 Oct 2025

Abstract

Biomarkers are important for the diagnosis and follow-up of neuromuscular diseases. Creatine kinase (CK) is a widely used marker of active muscle damage; however, it is not suitable for assessing muscle mass loss. Therefore, additional biomarkers are required to monitor skeletal muscle damage [...] Read more.

Biomarkers are important for the diagnosis and follow-up of neuromuscular diseases. Creatine kinase (CK) is a widely used marker of active muscle damage; however, it is not suitable for assessing muscle mass loss. Therefore, additional biomarkers are required to monitor skeletal muscle damage and loss. Titin plays an essential role in the structure and function of muscle fibers. It provides stability and elasticity to the sarcomeres. During sarcomere damage, fragments of titin and other proteins are released from muscle fibers and can be detected in blood and urine. Urinary titin-N fragment (UTN) detection is a noninvasive method for assessing and monitoring the extent of muscle damage. In addition to muscular dystrophies, elevated UTN levels have been observed in patients with sarcopenia. The UTN level increased significantly during eccentric muscle strain, indicating muscle damage, whereas the concentric load was associated with only a minimal increase in UTN. As titin is also present in the heart muscle, UTN can help diagnose cardiomyopathies and predict disease prognosis. In summary, the detection of urinary titin fragments is a promising tool for diagnosing and monitoring neuromuscular and cardiac diseases. While both CK and UTN rise and are related in acute conditions, their relationship is less clear in chronic diseases where muscle tissue damage and muscle mass loss are combined. Full article

(This article belongs to the Section Molecular Neurobiology)

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7 pages, 177 KB

Open AccessEditorial

Special Issue “Plant Responses to Abiotic and Biotic Stresses”

by Longxing Tao and Tingting Chen

Int. J. Mol. Sci. 2025, 26(19), 9651; https://doi.org/10.3390/ijms26199651 - 3 Oct 2025

Abstract

During their growth and development, plants often encounter a variety of biotic and abiotic stresses, such as drought, high salinity, extreme temperatures, heavy metal pollution, and pests and diseases [...] Full article

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

26 pages, 735 KB

Open AccessReview

Protective Effects of PACAP in Diabetic Complications: Retinopathy, Nephropathy and Neuropathy

by Dora Reglodi, Andrea Tamas, Inez Bosnyak, Tamas Atlasz, Edina Szabo, Lina Li, Gabriella Horvath, Balazs Opper, Peter Kiss, Liliana Lucas, Grazia Maugeri, Agata Grazia D’Amico, Velia D’Agata, Eszter Fabian, Gyongyver Reman and Alexandra Vaczy

Int. J. Mol. Sci. 2025, 26(19), 9650; https://doi.org/10.3390/ijms26199650 - 3 Oct 2025

Abstract

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide exerting, among others, strong trophic and protective effects. It plays a role in several physiological functions, including glucose homeostasis. The protective effects of PACAP are mainly mediated via its specific PAC1 receptor by stimulating anti-inflammatory, [...] Read more.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide exerting, among others, strong trophic and protective effects. It plays a role in several physiological functions, including glucose homeostasis. The protective effects of PACAP are mainly mediated via its specific PAC1 receptor by stimulating anti-inflammatory, anti-apoptotic and antioxidant pathways. The aim of the present review is to summarize data on the protective effects of PACAP in the three major complications of diabetes, retinopathy, nephropathy and neuropathy, as well as some other complications. In type 1 and type 2 diabetic retinopathy models and in glucose-exposed cells of the eye, PACAP counteracted the degeneration of retinal layers and inhibited apoptosis and factors leading to abnormal vessel growth. In models of nephropathy, kidney morphology was better retained after PACAP administration, with decreased apoptosis and fibrosis. In diabetic neuropathy, PACAP protected against axonal–myelin lesions and less activation in pain processing centers. This neuropeptide has several other beneficial effects in diabetes-induced complications like altered vascular response, cognitive deficits and atherosclerosis. The promising therapeutic effects of PACAP in several pathological conditions have encouraged researchers to design PACAP-related drugs and to develop ways to enhance tissue delivery. These intentions are expected to result in overcoming the hurdles preventing PACAP from being introduced into therapeutic treatments, including diabetes-related conditions. Full article

(This article belongs to the Special Issue Molecular Research on Type 1 Diabetes and Its Complications: 2nd Edition)

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18 pages, 728 KB

Open AccessReview

Healthcare-Associated Infections (HAI) in the Elderly: Molecular Mechanisms of Immunosenescence and Clinical, Nutritional and Therapeutic Implications

by Livia Moffa and Claudio Tana

Int. J. Mol. Sci. 2025, 26(19), 9649; https://doi.org/10.3390/ijms26199649 - 3 Oct 2025

Abstract

Healthcare-associated infections (HAIs) in the elderly represent a growing clinical and public health concern, primarily driven by age-related biological remodeling. Key mechanisms include immunosenescence, inflammaging, gut microbiota dysbiosis, and profound metabolic and epigenetic alterations, all of which progressively weaken host defense and resilience [...] Read more.

Healthcare-associated infections (HAIs) in the elderly represent a growing clinical and public health concern, primarily driven by age-related biological remodeling. Key mechanisms include immunosenescence, inflammaging, gut microbiota dysbiosis, and profound metabolic and epigenetic alterations, all of which progressively weaken host defense and resilience to pathogens. In this review, we delineate the molecular pathways underlying these processes, with particular attention to impaired innate and adaptive immune responses, dysfunctional cellular signaling, and disrupted immunometabolic networks that increase susceptibility to multidrug-resistant organisms and aggravate clinical outcomes in older patients. We also address the synergistic impact of frailty-related factors such as malnutrition, multimorbidity, and polypharmacy on infection risk. Finally, we discuss emerging translational perspectives, including nutritional interventions and microbiota-targeted strategies aimed at restoring immune competence and reducing infection burden. By integrating molecular mechanisms with clinical implications, this review highlights innovative opportunities for personalized prevention and management of HAIs in the aging population. Full article

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

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26 pages, 4811 KB

Open AccessArticle

Ginkgo Biloba and Green Tea Polyphenols Captured into Collagen–Lipid Nanocarriers: A Promising Synergistically Approach for Apoptosis Activation and Tumoral Cell Cycle Arrest

by Mirela Mihaila, Nicoleta Badea, Marionela Birliga, Marinela Bostan, Madalina Georgiana Albu Kaya and Ioana Lacatusu

Int. J. Mol. Sci. 2025, 26(19), 9648; https://doi.org/10.3390/ijms26199648 - 3 Oct 2025

Abstract

Considering the world’s growing interest in health-promoting phytochemicals, the current research investigated the development of a dual-captured Ginkgo Biloba and Green Tea Extract into Collagen-Nanostructured Lipid Nanocarriers (Col-NLC-GBil-GTE) for an enhanced therapeutic efficacy against hepatic, colon or breast cancer. NLC considerably [...] Read more.

Considering the world’s growing interest in health-promoting phytochemicals, the current research investigated the development of a dual-captured Ginkgo Biloba and Green Tea Extract into Collagen-Nanostructured Lipid Nanocarriers (Col-NLC-GBil-GTE) for an enhanced therapeutic efficacy against hepatic, colon or breast cancer. NLC considerably reduced cell viability; the most advanced cytotoxicity profile was determined on human colon adenocarcinoma cells (LoVo) and liver cancer cells (HepG2), e.g., tumor cell viability was 21.81% in the presence of Col-NLC-GBil-GTE, similar to that determined for Cisplatin. Col-NLC exhibited apoptosis in HepG2 and LoVo cells and no significant apoptosis induction in normal HUVECs. A 20% increase in apoptosis for HepG2 cells was registered for 100 μg/mL NLC-GBil-GTE compared to Cisplatin (Cis-Pt), e.g., a 63.4% total apoptosis for NLC-GBil-GTE versus a 52.6 apoptosis induced by 100 μg/mL of a chemotherapeutic drug. According to the cell cycle outcomes, an accumulation of hepatocyte HepG2 tumor cells in the G0/G1 phase was detected upon treatment with 100 mg/mL of NLC- and Col-NLC-GBil-GTE, simultaneously with a drastic decrease in the S phase, which may indicate a cell number reduction that enters in the division cycle. The simultaneous delivery of GBil and GTE by synchronizing their bioactivities offers several advantages; Col-NLC-GBil-GTE can be viewed as a noteworthy strategy for consideration in connection with antitumor therapeutic protocols. Full article

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

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27 pages, 4073 KB

Open AccessArticle

Thyroid Hormone T4 Alleviates Traumatic Brain Injury by Enhancing Blood–Brain Barrier Integrity

by Mayuri Khandelwal, Zhe Ying and Fernando Gomez-Pinilla

Int. J. Mol. Sci. 2025, 26(19), 9632; https://doi.org/10.3390/ijms26199632 - 3 Oct 2025

Abstract

Traumatic brain injury (TBI) disrupts the blood–brain barrier (BBB), resulting in increased permeability, neuronal loss, and cognitive dysfunction. This study investigates the therapeutic potential of thyroid hormone (T4) to reduce BBB dysfunction following moderate fluid percussion injury. T4 injection (intraperitoneal) after TBI restores [...] Read more.

Traumatic brain injury (TBI) disrupts the blood–brain barrier (BBB), resulting in increased permeability, neuronal loss, and cognitive dysfunction. This study investigates the therapeutic potential of thyroid hormone (T4) to reduce BBB dysfunction following moderate fluid percussion injury. T4 injection (intraperitoneal) after TBI restores the levels of pericytes and endothelial cells vital for BBB integrity, reduces edema by downregulating AQP-4 gene expression, and enhances levels of the tight junction protein ZO-1. T4 counteracts the TBI-related increase in MMP-9 and TLR-4, significantly reducing BBB permeability. Furthermore, T4 enhances the neuroprotective functions of astrocytes by promoting the activity of A2 astrocytes. Additionally, T4 treatment increases DHA levels (important for membrane integrity and function), stimulates mitochondrial biogenesis, and leads to a notable improvement in spatial learning and memory retention. These findings suggest that T4 has significant potential to reduce vascular leakage and inflammation after TBI, thereby improving cognitive function and maintaining BBB integrity. Full article

(This article belongs to the Special Issue The Blood–Brain Barrier and Neuroprotection)

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15 pages, 2315 KB

Open AccessArticle

Coenzyme Q10 Ameliorates Chemotherapy-Induced Neurotoxicity in iPSC-Derived Neurons by Reducing Oxidative Stress

by Nidaa A. Ababneh, Razan AlDiqs, Mohammad H. Gharandouq, Mohammad A. Ismail, Raghda Barham, Fairouz Nairat, Omar Hamdan, Qais Mussa, Momen Sarhan, Amira T. Masri, Anas Abu-Humaidan, Sofian Al Shboul, Areej Abuhammad, Abdalla Awidi and Tareq Saleh

Int. J. Mol. Sci. 2025, 26(19), 9647; https://doi.org/10.3390/ijms26199647 - 2 Oct 2025

Abstract

Chemotherapy-induced neurotoxicity (CIN) is a major barrier against optimal anticancer treatment. This study investigated the neuroprotective effects of the naturally occurring antioxidant, Coenzyme Q10 (CoQ10), against CIN using a model of induced pluripotent stem cell (iPSC)-derived neurons. iPSCs have consistently proven to be [...] Read more.

Chemotherapy-induced neurotoxicity (CIN) is a major barrier against optimal anticancer treatment. This study investigated the neuroprotective effects of the naturally occurring antioxidant, Coenzyme Q10 (CoQ10), against CIN using a model of induced pluripotent stem cell (iPSC)-derived neurons. iPSCs have consistently proven to be reliable for disease modeling and drug discovery. We employed cell viability, oxidative stress, and mitochondrial function assays to measure the effect of 10 μM CoQ10 on iPSC-derived motor neuron progenitors (iPSC-MNPs) that were exposed to five chemotherapeutic agents: 5-Fluorouracil, methotrexate, paclitaxel (0, 1, and 10 μM) and doxorubicin, and vincristine (0, 0.1, and 1 μM). Our findings show that CoQ10 significantly reversed the reduction in cell viability inflicted by the exposure of iPSCs-MNPs to all five chemotherapeutics. Moreover, CoQ10 treatment resulted in a marked reduction in intracellular ROS levels and enhancement of mitochondrial membrane potential (MMP) in a drug- and dose-dependent manners, highlighting its role in preserving mitochondrial health. This study is the first to explore the protective effects of CoQ10 against CIN using an iPSC-derived neuronal platform, offering insights into its potential therapeutic use. Further investigation is essential to validate these findings and to determine the behavioral effects of CoQ10 in in vivo models of CIN. Full article

(This article belongs to the Special Issue Toxicity of Metals, Metal-Based Drugs, and Microplastics)

26 pages, 4520 KB

Open AccessArticle

T-Cadherin Finetunes Proliferation–Differentiation During Adipogenesis via PI3K–AKT Signaling Pathway

by Polina Klimovich, Ilya Brodsky, Valentina Dzreyan, Marianna Ivleva, Olga Grigorieva, Mark Meshcheriakov, Ekaterina Semina, Veronika Sysoeva, Vsevolod Tkachuk and Kseniya Rubina

Int. J. Mol. Sci. 2025, 26(19), 9646; https://doi.org/10.3390/ijms26199646 - 2 Oct 2025

Abstract

Adipose tissue renewal requires precise coordination of stem/progenitor cell proliferation, preadipocyte commitment, and terminal adipocyte differentiation. T-cadherin (CDH13), an atypical GPI-anchored cadherin, is expressed in adipose tissue and functions as a receptor for high-molecular-weight (HMW) adiponectin—a key adipokine produced by adipose tissue and [...] Read more.

Adipose tissue renewal requires precise coordination of stem/progenitor cell proliferation, preadipocyte commitment, and terminal adipocyte differentiation. T-cadherin (CDH13), an atypical GPI-anchored cadherin, is expressed in adipose tissue and functions as a receptor for high-molecular-weight (HMW) adiponectin—a key adipokine produced by adipose tissue and involved in metabolic regulation. While T-cadherin is implicated in cardiovascular and metabolic homeostasis, its role in adipogenesis still remains poorly understood. In this study, we used the 3T3-L1 preadipocyte model to investigate the function of T-cadherin in adipocyte differentiation. We analyzed T-cadherin expression dynamics during differentiation and assessed how T-cadherin overexpression or knockdown affects lipid accumulation, expression of adipogenic markers, and key signaling pathways including ERK, PI3K–AKT, AMPK, and mTOR. Our findings demonstrate that T-cadherin acts as a negative regulator of adipogenesis. T-cadherin overexpression ensured a proliferative, undifferentiated cell state, delaying early adipogenic differentiation and suppressing both lipid droplet accumulation and the expression of adipogenic markers. In contrast, T-cadherin downregulation accelerated differentiation, enhanced lipid accumulation, and increased insulin responsiveness, as indicated by PI3K–AKT pathway activation at specific stages of adipogenesis. These results position T-cadherin as a key modulator of adipose tissue plasticity, regulating the balance between progenitor expansion and terminal differentiation, with potential relevance to obesity and metabolic disease. Full article

(This article belongs to the Special Issue Fundamental and Practical Perspectives in Regenerative Medicine: Proceedings of the VI National Congress of Regenerative Medicine (2024))

18 pages, 280 KB

Open AccessArticle

Connections Between Gene Polymorphism and Fetlock and Hock Measurements in Polish Sport Horses

by Dorota Lewczuk, Maria Wypchło, Mateusz Hecold, Roma Buczkowska and Agnieszka Korwin-Kossakowska

Int. J. Mol. Sci. 2025, 26(19), 9645; https://doi.org/10.3390/ijms26199645 - 2 Oct 2025

Abstract

Finding the causative mutations for musculoskeletal system development and health status is of a higher priority for all sport horse breeders’ associations. Of the regulating proteins involved in animal ossification, 15 gene polymorphisms were chosen to be identified as connected with the nine [...] Read more.

Finding the causative mutations for musculoskeletal system development and health status is of a higher priority for all sport horse breeders’ associations. Of the regulating proteins involved in animal ossification, 15 gene polymorphisms were chosen to be identified as connected with the nine fetlock and 14 hock bone structures measurements of 198 horses. All measurements were taken using X-rays of the limbs, which were available at the beginning and end of the horse training. The analysis of variance (GLM, SAS program) was performed taking into account identified training and horse-connected characteristics, and gene polymorphism. The larger size of the bone structure was achieved in the fetlock for the heterozygotes of COL9A2, AOAH1, BMPER, HYAL3, and ELMO1. The heterozygotes were superior to homozygotes in the hock for the following genes: COL9A2, HYAL3, ANLN, and HYAL1. The lower homozygote values were obtained for GG in CPVL in fetlock measurements, TT for HYAL3 in fetlock, TT for ANLN in fetlock, CC for FRZB in the hock, TT for MATN in the hock, and TT for COL5A2 in the hock than their opposite homozygote and heterozygote variants. COL9A2 and HYAL3 are expressed in the same way for most of the bone structures in both joints. Full article

(This article belongs to the Special Issue Cytokines and Other Biomarkers of Health Status)

20 pages, 1949 KB

Open AccessArticle

Brassinosteroid Synthesis and Perception Differently Regulate Phytohormone Networks in Arabidopsis thaliana

by Yaroslava Bukhonska, Michael Derevyanchuk, Roberta Filepova, Jan Martinec, Petre Dobrev, Eric Ruelland and Volodymyr Kravets

Int. J. Mol. Sci. 2025, 26(19), 9644; https://doi.org/10.3390/ijms26199644 - 2 Oct 2025

Abstract

Brassinosteroids (BRs) are essential regulators of plant development and stress responses, but the distinct contributions of BR biosynthesis and signaling to hormonal crosstalk remain poorly defined. Here, we investigated the effects of the BR biosynthesis inhibitor brassinazole (BRZ) and the BR-insensitive mutant bri1-6 [...] Read more.

Brassinosteroids (BRs) are essential regulators of plant development and stress responses, but the distinct contributions of BR biosynthesis and signaling to hormonal crosstalk remain poorly defined. Here, we investigated the effects of the BR biosynthesis inhibitor brassinazole (BRZ) and the BR-insensitive mutant bri1-6 on endogenous phytohormone profiles in Arabidopsis thaliana. Using multivariate analysis and targeted hormone quantification, we show that BRZ treatment and BRI1 disruption alter hormone balance through partially overlapping but mechanistically distinct pathways. Principal component analysis (PCA) and hierarchical clustering revealed that BRZ and the bri1-6 mutation do not phenocopy each other and that BRZ still alters hormone profiles even in the bri1-6 mutant, suggesting potential BRI1-independent effects. Both BRZ treatment and the bri1-6 mutation tend to influence cytokinins and auxin conjugates divergently. On the contrary, their effects on stress-related hormones converge: BRZ decreases salicylic acid (SA), jasmonic acid (JA), and abscisic acid (ABA) in the WT leaves; similarly, bri1-6 mutants show reduced SA, JA, and ABA. These results indicate that BR biosynthesis and BRI1-mediated perception may contribute independently to hormonal reprogramming, with BRZ eliciting additional effects, possibly via metabolic feedback, compensatory signaling, or off-target action. Hormone correlation analyses revealed conserved co-regulation clusters that reflect underlying regulatory modules. Altogether, our findings provide evidence for a partial uncoupling of BR levels and BR signaling and illustrate how BR pathways intersect with broader hormone networks to coordinate growth and stress responses. Full article

(This article belongs to the Special Issue Emerging Insights into Phytohormone Signaling in Plants)

24 pages, 1426 KB

Open AccessReview

Dietary and Pharmacological Modulation of Aging-Related Metabolic Pathways: Molecular Insights, Clinical Evidence, and a Translational Model

by Antonio Fernando Murillo-Cancho, David Lozano-Paniagua and Bruno José Nievas-Soriano

Int. J. Mol. Sci. 2025, 26(19), 9643; https://doi.org/10.3390/ijms26199643 - 2 Oct 2025

Abstract

Advances in geroscience suggest that aging is modulated by molecular pathways that are amenable to dietary and pharmacological intervention. We conducted an integrative critical review of caloric restriction (CR), intermittent fasting (IF), and caloric restriction mimetics (CR-mimetics) to compare shared mechanisms, clinical evidence, [...] Read more.

Advances in geroscience suggest that aging is modulated by molecular pathways that are amenable to dietary and pharmacological intervention. We conducted an integrative critical review of caloric restriction (CR), intermittent fasting (IF), and caloric restriction mimetics (CR-mimetics) to compare shared mechanisms, clinical evidence, limitations, and translational potential. Across modalities, CR and IF consistently activate AMP-activated protein kinase and sirtuins, inhibit mTOR (mechanistic target of rapamycin) signaling, and enhance autophagy, aligning with improvements in insulin sensitivity, lipid profile, low-grade inflammation, and selected epigenetic aging measures in humans. CR-mimetics, such as metformin, resveratrol, rapamycin, and spermidine, partially reproduce these effects; however, long-term safety and efficacy in healthy populations remain incompletely defined. Methodological constraints—short trial duration, selective samples, intermediate (nonclinical) endpoints, and limited adherence monitoring—impede definitive conclusions on hard outcomes (frailty, disability, hospitalization, mortality). We propose the Active Management of Aging and Longevity (AMAL) model, a three-level biomarker-guided framework that integrates personalized diet, chrono-nutrition, exercise, and the selective use of CR-mimetics, along with digital monitoring and decision support. AMAL emphasizes epigenetic clocks, multi-omics profiling, inflammatory and microbiome metrics, and adaptive protocols to enhance adherence and clinical relevance. Overall, CR, IF, and CR mimetics constitute promising, complementary strategies to modulate biological aging; rigorous long-term trials with standardized biomarkers and clinically meaningful endpoints are needed to enable their scalable implementation. Full article

(This article belongs to the Special Issue The Roles of Diet and Nutrition in Metabolic Diseases: Advancements in Molecular Insights)

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23 pages, 2077 KB

Open AccessArticle

Tailored Reaction Conditions and Automated Radiolabeling of [¹⁷⁷Lu]Lu-PSMA-ALB-56 in a ⁶⁸Ga Setting: The Critical Impact of Antioxidant Concentrations

by Johanne Vanney, Léa Rubira, Jade Torchio and Cyril Fersing

Int. J. Mol. Sci. 2025, 26(19), 9642; https://doi.org/10.3390/ijms26199642 - 2 Oct 2025

Abstract

The growing use of experimental radiopharmaceuticals for targeted radionuclide therapy (TRT) highlights the need for robust “in house” radiolabeling protocols. Among these, PSMA-ALB-56 is a PSMA ligand incorporating an albumin-binding moiety to enhance pharmacokinetics, which showed promise for prostate cancer treatment. This study [...] Read more.

The growing use of experimental radiopharmaceuticals for targeted radionuclide therapy (TRT) highlights the need for robust “in house” radiolabeling protocols. Among these, PSMA-ALB-56 is a PSMA ligand incorporating an albumin-binding moiety to enhance pharmacokinetics, which showed promise for prostate cancer treatment. This study investigated manual radiolabeling conditions of this vector molecule with lutetium-177 and developed a corresponding automated synthesis protocol. Manual experiments on low activities explored buffer systems and antioxidants, identifying sodium acetate buffer and L-methionine as optimal, achieving radiochemical purities above 97% with excellent stability over 48 h. However, when these conditions were transposed directly to an automated process on a GAIA^® module with activities > 2 GBq, radiochemical purity dropped below 70% due to significant radiolysis. This result emphasized that conditions optimized at low activities are not directly transferable to high-activity automated production, and highlighted the crucial role of antioxidant concentration. An optimized automated method was subsequently developed, integrating a solid-phase extraction purification step, higher antioxidant levels during radiolabeling and formulation, and a larger final product volume. These changes led to radiochemical purities above 98.9% and excellent product stability over 120 h for 3 test batches. The presence of high concentrations of methionine and ascorbic acid was essential to protect against radiolysis. This work underscores the importance of adjusting radiolabeling strategies during process scale-up and confirmed that antioxidant concentration is essential for successful ¹⁷⁷Lu radiolabeling. The optimized automated method developed here for [¹⁷⁷Lu]Lu-PSMA-ALB-56 may also be adapted to other radiopharmaceuticals in development for TRT. Full article

(This article belongs to the Special Issue Radiolabeled Compounds for Theranostic Applications in Oncology)

18 pages, 17064 KB

Open AccessArticle

Interplay of the Genetic Variants and Allele Specific Methylation in the Context of a Single Human Genome Study

by Maria D. Voronina, Olga V. Zayakina, Kseniia A. Deinichenko, Olga Sergeevna Shingalieva, Olga Y. Tsimmer, Darya A. Tarasova, Pavel Alekseevich Grebnev, Ekaterina A. Snigir, Sergey I. Mitrofanov, Vladimir S. Yudin, Anton A. Keskinov, Sergey M. Yudin, Dmitry V. Svetlichnyy and Veronika I. Skvortsova

Int. J. Mol. Sci. 2025, 26(19), 9641; https://doi.org/10.3390/ijms26199641 - 2 Oct 2025

Abstract

The methylation of CpG sites with 5mC mark is a dynamic epigenetic modification. However, the relationship between the methylation and the surrounding genomic sequence context remains poorly explored. Investigation of the allele methylation provides an opportunity to decipher the interplay between differences in [...] Read more.

The methylation of CpG sites with 5mC mark is a dynamic epigenetic modification. However, the relationship between the methylation and the surrounding genomic sequence context remains poorly explored. Investigation of the allele methylation provides an opportunity to decipher the interplay between differences in the primary DNA sequence and epigenetic variation. Here, we performed high-coverage long-read whole-genome direct DNA sequencing of one individual using Oxford Nanopore technology. We also used Illumina whole-genome sequencing of the parental genomes in order to identify allele-specific methylation sites with a trio-binning approach. We have compared the results of the haplotype-specific methylation detection and revealed that trio binning outperformed other approaches that do not take into account parental information. Also, we analysed the cis-regulatory effects of the genomic variations for influence on CpG methylation. To this end, we have used available Deep Learning models trained on the primary DNA sequence to score the cis-regulatory potential of the genomic loci. We evaluated the functional role of the allele-specific epigenetic changes with respect to gene expression using long-read Nanopore RNA sequencing. Our analysis revealed that the frequency of SNVs near allele-specific methylation positions is approximately four times higher compared to the biallelic methylation positions. In addition, we identified that allele-specific methylation sites are more conserved and enriched at the chromatin states corresponding to bivalent promoters and enhancers. Together, these findings suggest that significant impact on methylation can be encoded in the DNA sequence context. In order to elucidate the effect of the SNVs around sites of allele-specific methylation, we applied the Deep Learning model for detection of the cis-regulatory modules and estimated the impact that a genomic variant brings with respect to changes to the regulatory activity of a DNA loci. We revealed higher cis-regulatory impact variants near differentially methylated sites that we further coupled with transcriptomic long-read sequencing results. Our investigation also highlights technical aspects of allele methylation analysis and the impact of sequencing coverage on the accuracy of genomic phasing. In particular, increasing coverage above 30X does not lead to a significant improvement in allele-specific methylation discovery, and only the addition of trio binning information significantly improves phasing. We investigated genomic variation in a single human individual and coupled computational discovery of cis-regulatory modules with allele-specific methylation (ASM) profiling. In this proof-of-concept analysis, we observed that SNPs located near methylated CpG sites on the same haplotype were enriched for sequence features suggestive of high-impact regulatory potential. This finding—derived from one deeply sequenced genome—illustrates how phased genetic and epigenetic data analyses can jointly put forward a hypotheses about the involvement of regulatory protein machinery in shaping allele-specific epigenetic states. Our investigation provides a methodological framework and candidate loci for future studies of genomic imprinting and cis-mediated epigenetic regulation in humans. Full article

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

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28 pages, 2759 KB

Open AccessReview

Design and Efficacy of Oncolytic Viruses and Antitumor Vaccines: A Dead End in the Immunotherapy of Pancreatic Cancer?

by Eduard Achim, Elena Pîrlici, Cecilia Cristea and Mihaela Tertis

Int. J. Mol. Sci. 2025, 26(19), 9640; https://doi.org/10.3390/ijms26199640 - 2 Oct 2025

Abstract

Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest malignancies, marked by late diagnosis, limited responsiveness to conventional therapies, and an immunosuppressive tumor microenvironment. While immunotherapy has transformed treatment paradigms in several cancers, its efficacy in PDAC has been minimal. Oncolytic viruses and [...] Read more.

Pancreatic ductal adenocarcinoma (PDAC) remains one of the deadliest malignancies, marked by late diagnosis, limited responsiveness to conventional therapies, and an immunosuppressive tumor microenvironment. While immunotherapy has transformed treatment paradigms in several cancers, its efficacy in PDAC has been minimal. Oncolytic viruses and therapeutic cancer vaccines have emerged as promising immunotherapeutic strategies designed to stimulate robust, tumor-specific immune responses and reshape the immune landscape. However, despite encouraging preclinical data, clinical translation in PDAC has been largely disappointing. This review critically evaluates the design, delivery, and efficacy of oncolytic virotherapy and cancer vaccines in PDAC, examining barriers such as stromal desmoplasia, immune exclusion, and tumor heterogeneity. We also explore combination strategies integrating checkpoint inhibitors, chemotherapy, radiotherapy, and stromal modulation to overcome resistance. Ultimately, the viability of these approaches hinges on a clearer understanding of their mechanistic limitations and the refinement of delivery platforms. These factors will determine whether oncolytic viruses and cancer vaccines can be successfully repositioned within the therapeutic arsenal or warrant reevaluation in the evolving landscape of PDAC treatment. Full article

(This article belongs to the Section Molecular Immunology)

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