Esculetin, a coumarin derivative, exhibits diverse biological activities; however, its impact on pancreatic β-cell stress responses and survival remains poorly defined. In this study, we investigated the effects of esculetin on cellular stress signaling, apoptosis, and functional gene expression in INS-1 pancreatic β-cells. Following treatment with 3000 µM esculetin for 6 h, cell viability, proliferation, apoptosis, oxidative stress, endoplasmic reticulum (ER) stress markers, MAPK signaling components, cell cycle distribution, and β-cell–specific gene expression were assessed. In silico molecular docking was performed to explore putative interactions between esculetin and proteins involved in ER stress, MAPK signaling, and apoptosis. Treatment with 3000 µM esculetin for 6 h increased apoptotic cell death by approximately 32-fold and elevated total oxidation status and oxidative stress index. Intracellular insulin, Ca²⁺, and CA19-9 levels were increased, accompanied by cell cycle arrest characterized by G0/G1 phase accumulation and a reduction in the G2/M population. Expression of ER stress sensors ATF6, IRE1α, and CHOP was upregulated, whereas PERK expression was reduced. In parallel, MAPK/JNK pathway activation was evidenced by increased levels of p-ATF2, p-c-Jun, and HSP90, along with decreased p-Elk1. Gene expression analysis revealed marked downregulation of β-cell functional and identity–associated genes, including Ins1, Ins2, IR, Akt, MafA, Nkx6.1, Pdx1, NeuroD1, and Pax6, while FoxO1 expression was upregulated. Collectively, these findings suggest the involvement of ER stress-associated MAPK/JNK signaling in esculetin-induced apoptosis and functional impairment in INS-1 pancreatic β-cells, providing mechanistic insight into stress-associated β-cell dysfunction.
Intestinal inflammation and barrier disruption induced by high-altitude exposure are recognized contributors to colitis. Although artificial-enzyme-engineered Bifidobacterium longum (AE-BL) has demonstrated therapeutic potential against inflammatory bowel disease, the precise mechanisms remain elusive. The present study aimed to assess the efficacy of AE-BL against high-altitude colitis and to elucidate its underlying mechanisms. AE-BL was generated through the assembly of single-atom enzymes (SAzymes) with Bifidobacterium longum (BL). In vivo investigations involved housing mice in a hypobaric hypoxic chamber to replicate high-altitude conditions, followed by AE-BL administration. After seven days, colon specimens were collected for histopathological evaluation, inflammatory and ferroptosis-related parameter analyses, and the expression of glycerol-3-phosphate dehydrogenase 2 (GPD2), tight junction proteins, and ferroptosis markers. An in vitro colitis model was also established using Caco-2 cells subjected to lipopolysaccharide (LPS) and hypoxia, followed by AE-BL treatment. Administration of AE-BL markedly ameliorated high-altitude-induced colitis in mice, as reflected by attenuated weight loss, increased colon length, reduced disease activity index (DAI), and diminished histopathological injury. Pro-inflammatory cytokine production was suppressed, and intestinal barrier integrity was preserved. Mechanistic investigations indicated that the protective actions of AE-BL were potentially mediated through suppression of GPD2-driven ferroptosis. Under hypoxic conditions, AE-BL significantly reduced ferroptosis in colon tissue and colonic epithelial cells both in vivo and in vitro, thereby alleviating inflammation and restoring intestinal barrier function.
Plant secondary metabolites are mainly synthesized and stored in secretory tissues. Secretory canal development has been mainly characterized in Apiaceae. The secretory canals of Peucedanum praeruptorum contain pharmacologically active coumarins, but their organ-specific distribution and developmental dynamics remain poorly understood. This study integrated light microscopy (LM), transmission electron microscopy (TEM), X-ray microcomputed tomography (µ-CT), and high-performance liquid chromatography (HPLC) to investigate canal development, distribution, ultrastructure, 3D architecture, and coumarin accumulation in P. praeruptorum roots. Histological analysis showed that canals adjacent to the periderm originate from pericycle cells, whereas those in secondary phloem arise from parenchyma differentiation; both develop schizogenously. Canal quantity and dimensions varied temporally. Canals located in phloem showed density increasing toward the cambial zone, where cross-sectional areas were smaller. The canal density index increased from September to November, peaking on November 15, then declined. HPLC revealed dynamic accumulation of five major coumarins: content increased from September, peaked on November 15, then gradually decreased. TEM showed that epithelial cells surrounding the canal lumen were rich in Golgi, ER, mitochondria, plastids, starch grains, and osmiophilic droplets. µ-CT volumetric analysis and segmentation generated detailed 3D models, revealing spatial organization and enabling size-based grouping of canals (1000–3000 μm). These dimensional characteristics aligned with developmental progression. This study characterizes the ontogeny, distribution, ultrastructure, and 3D architecture of secretory canals, providing a structural foundation for investigating correlations between secretory tissues and compound synthesis.
Soybean is a major oilseed crop of global importance. Although haploid and doubled haploid (DH) production has been attempted in soybean, androgenesis success remains very low (approximately 2
Winter cereals at the etiolated seedling stage possess the ability to undergo cold hardening, after which they develop varying levels of frost tolerance depending on their genotype. However, the functioning of stress-protective systems in etiolated wheat seedlings at the biochemical and cellular levels remains poorly understood. We investigated the contribution of dehydrins, osmoprotective and antioxidant systems to cold adaptation in six Triticum aestivum (L.) cultivars differing in frost resistance. The seedlings were subjected to a 6-day hardening treatment at 3 °C, followed by freezing at − 9 °C. The survival rate of the most frost-tolerant cultivars after 5 h of freezing was 54–73
Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains a major global health challenge due to persistent diagnostic gaps. CRISPR–Cas–based diagnostics have emerged as highly sensitive and programmable platforms for nucleic acid detection, enabling rapid identification of Mtb targets, including drug-resistance–associated mutations. These systems integrate isothermal amplification, diverse Cas effectors, and multiple signal readout strategies to achieve high analytical performance. This review provides a comparative analysis of clinically evaluated CRISPR-based TB diagnostic platforms, highlighting substantial variability in assay design, performance, and translational readiness. While many platforms demonstrate strong analytical sensitivity, their implementation remains constrained by workflow complexity and limited integration into true point-of-care formats. This highlights that successful clinical translation of CRISPR-based TB diagnostics is determined more by real-world adaptability than by analytical performance alone. The current review presents a comparative analysis of CRISPR-based diagnostic platforms for tuberculosis, evaluating the variability in assay design, analytical and clinical performance, and translational readiness across currently available systems. Overview of CRISPR–Cas–based diagnostic platforms for Mycobacterium tuberculosis, illustrating the workflow from sample collection and nucleic acid processing to isothermal amplification, target-specific detection using Cas12/Cas13 systems, and diverse signal readouts (lateral flow, fluorescent, and biosensor-based). Despite strong analytical performance, clinical utility remains limited by sample preparation requirements, workflow complexity, and incomplete integration into deployable point-of-care systems. Created with BioRender.com.
Chromosome movements during meiosis are essential for accurate homolog recognition and segregation. However, two striking phenomena, rapid prophase chromosome movements (RPMs) and the formation of unusual nuclear structures, both involving large-scale chromatin dynamics and occurring simultaneously during early meiotic prophase I, remain poorly understood in plants. RPMs are known to facilitate homolog pairing across diverse taxa, but their regulation and robustness in plants remain unclear. Similarly, a phenomenon characterized by the formation of nuclear protrusions, that extend through intercellular channels into the cytoplasm of neighboring meiocytes, is frequently reported during plant meiosis. As a rule, this process is referred to as cytomixis, yet its nature and biological significance remain unresolved. Here, we integrate the study of these processes by investigating Arabidopsis thaliana male meiocytes at early prophase I using live-cell imaging with ASY1:GFP as a reporter for chromosome dynamics. We show that chromatin in nuclear protrusions remains active and displays RPMs indistinguishable from those of intact nuclei. Remarkably, a process unavoidably involving mechanical stress and disruption of cytoskeletal connections, does not interrupt RPMs. These findings demonstrate that meiotic RPMs are inherently robust and largely independent of the local cytoplasmic environment and support the view that cytomixis is a normal cytological process rather than a pathology or artifact.
Anatomical insights from plant studies offer a unique way to understand the tissue-level adaptations that enable some species to develop aluminum (Al) tolerance. To conduct functional anatomy studies, methods are required that uphold both structural integrity and cellular chemical stability. Previous research on the Al-tolerant species in Fagopyrum esculentum has focused primarily on physiological mechanisms, leaving technical and structural gaps regarding its sequestration in tissues. This research presents a histochemical approach to elucidate Al compartmentalization in mature F. esculentum plants. Samples were preserved with a modified Zamboni fixative to avoid metal leaching before being embedded in glycol methacrylate (GMA). High-quality 12 μm sections were obtained. For anatomical description, Toluidine Blue O was utilized, while Chromazurol S (CAS) was employed for histochemistry. In addition, laser microdissection of the leaves and Al quantification by ICP-MS were performed. The results revealed a progressive acropetal gradient. The marginal papillae of the leaves exhibited a concentration of 4352 ± 0.5 mg Al*kg⁻¹ DW (1.6 times higher than the lamina fraction). This pattern shows that Al sequestration in this region is a permanent trait inherited from the developmental start. This study offers a replicable foundation for exploring metal compartmentalization and detoxification processes in F. esculentum and related metal-tolerant herbaceous plants.
One of the most frequently used plant cell lines is the tobacco cell line BY-2, which was established from Nicotiana tabacum L. cv. Bright Yellow 2 plants. However, surprisingly, after more than 50 years, the original information about the authors, the place and the method of its establishment is quite laborious to obtain. This could be important for any de novo derivation of highly proliferative cell lines. In this brief reflection article, we summarise the publicly available information on the origin of BY-2 in the 1960s, when it was used in applied research at the Central Research Institute of Japan Tobacco Salt Public Corporation (JTS). Thanks largely to Professor Toshiyuki Nagata’s activities, it spread to the rest of the world in the 1980s, becoming a cellular model for basic research into the cell cycle, membrane and cytoskeletal dynamics, and many other processes. Available literature suggests that the BY-2 cell line was derived from calli induced on the stems of seedlings of the Bright Yellow-2 Virginia-type tobacco cultivar. We hope that the information we have gathered will be useful for everyone working with, or planning to use, the BY-2 cell line. We also hope that it will encourage efforts to search for more details in old Japanese patent records that could be used to establish new, highly proliferating lines.
NADPH oxidase, a key enzyme responsible for reactive oxygen species (ROS) production in plants, plays a central role in regulating plant growth and development as well as the transmission of stress signals. However, the regulatory mechanism of ROS mediated by NADPH oxidase in Brassica napus remains unclear. In this study, B. napus 16NTS309 was used as the experimental material, and NADPH oxidase-specific inhibitor (diphenyleneiodonium chloride, DPI) was applied to treat seeds, seedlings, and callus tissues respectively. Combined with nitroblue tetrazolium (NBT) staining, tissue section observation, and superoxide anion (O2−) content determination, the mechanism by which NADPH oxidase-mediated ROS (O2−) regulates seed germination, cell division, and cold stress signal transmission in B. napus was systematically investigated. The results showed that: (1) DPI treatment completely inhibited seed germination (germination rate = 0
Drought represents one of the most pervasive and intensifying abiotic stresses under changing climate regimes severely constraining agricultural productivity, ecosystem stability, and global food security. Water deficit disrupts cellular homeostasis, reduces photosynthetic efficiency, and induces excessive accumulation of reactive oxygen species (ROS), resulting in oxidative damage. To survive under such conditions, plants employ a diverse array of adaptive responses, including osmotic adjustment, antioxidant defense, hormonal signalling, and stress-responsive gene regulation. Among the key signalling molecules involved in drought tolerance, nitric oxide (NO) and abscisic acid (ABA) have emerged as pivotal signalling molecules orchestrating a wide spectrum of physiological and molecular responses under drought. NO functions as a versatile signalling molecule that regulates redox homeostasis, enhances antioxidant activity, and promotes the accumulation of osmoprotectant. ABA maintains drought perception by inducing stomatal closure, and activating stress-responsive pathways. Co-application of NO and ABA regulates seed germination, root-shoot growth, and stomatal movement, thereby improving relative water content (RWC), membrane stability index (MSI), and photosynthetic efficiency while reducing oxidative stress markers such as malondialdehyde (MDA) and hydrogen peroxide (H2O2). This comprehensive review navigates through a clear and integrative overview of the mechanistic role of NO and ABA, and at the molecular level, NO and ABA modulate drought tolerance through transcriptional regulation, mRNA-level control, and translational modification of stress-responsive genes. Additionally, emerging strategies, including plant-growth promoting rhizobacteria (PGPR), marker-assisted selection (MAS) with QTL mapping, and genome editing tools such as CRISPR/Cas systems, offer promising approaches for enhancing drought tolerance and developing climate-resilient crop varieties.
Calcium (Ca2⁺) signatures are central to plant stress signaling, and GLR and CNGC channels are major Ca2⁺ entry routes. We integrated comparative phylogenomics, protein and promoter analyses, localization prediction, protein-interaction inference, miRNA-target prediction, and RT-qPCR validation to examine the regulation of wheat GLR/CNGCs under heat stress. Phylogenies separated GLR and CNGC lineages and revealed wheat-specific expansion consistent with polyploid retention. TaCNGCs were generally alkaline and more often predicted as unstable, whereas TaGLRs showed broader pI variation and greater predicted stability. Most proteins were predicted to localize to the plasma membrane, with a subset predicted to localize to chloroplasts. Promoters were enriched for light, ABA/MeJA/auxin, MYB drought, and anaerobic/energy–stress motifs. Network analyses highlighted tae-miR399, tae-miR167a, tae-miR156a, tae-miR164, and tae-miR171a as inferred regulatory hubs targeting selected TaGLR and TaCNGC transcripts. RT-qPCR across 0–72 h heat exposure showed early miRNA induction, reciprocal repression of many channel transcripts, and genotype-dependent recovery. The tolerant genotype displayed stronger transient miRNA activation and clearer channel transcript rebound by 48–72 h. These data support an inferred, reversible miRNA–channel regulatory model during heat acclimation and nominate candidates for direct target and functional validation.
This review aims to examine the role of probiotics in modulating the gut-derived incretin peptide axis in type 2 diabetes (T2D), with emphasis on GLP-1, related gut peptides such as GLP-2, microbial metabolite signaling, and both conventional and engineered probiotic approaches. Narrative synthesis of literature from PubMed, Scopus, and Google Scholar, using search terms such as ‘probiotics’, ‘GLP-1’, ‘type 2 diabetes’, and ‘gut microbiota’. Recent literature from 2015 to 2025 was prioritized, including randomized controlled trials, meta-analyses, systematic reviews, and relevant preclinical studies. Earlier landmark studies were included only when they established foundational mechanisms related to incretin biology, SCFA–FFAR signaling, or GLP-1 secretion physiology. Probiotics promote GLP-1 secretion via SCFAs binding to FFAR2/3 receptors, leading to improved glycemic control in meta-analyses, though with heterogeneity. Engineered probiotics like LgsGPA show superior preclinical efficacy in alleviating hyperglycemia and restoring β-cell function. While conventional probiotics offer benefits, engineered systems represent a promising advancement, requiring further clinical and regulatory development for personalized T2D therapy.
Despite the marked diversity of carpel closure, stigmatic surfaces, and stylar configurations documented in Alismataceae, no ultrastructural analyses of these tissues have been conducted for the family. Here, we present the first integrated morphological, anatomical, and ultrastructural study of the stigma and style in four species representing the major clades of Alismataceae: Hydrocleys nymphoides, Alisma plantago-aquatica, Sagittaria montevidensis, and Echinodorus grandiflorus. Using light microscopy, SEM, and TEM, we found marked differences in both stigma and style organization among species. Three species exhibited papillose stigmas, whereas E. grandiflorus presented a non-papillose receptive surface. H. nymphoides showed an open style with a well-developed stylar canal, whereas the remaining species exhibited closed styles with reduced or absent transmitting tissue. These structural differences are associated with contrasting secretory patterns, organelle composition, and inferred pollen tube pathways, highlighting clear distinctions between open and closed stylar systems. Our results reveal substantial intrafamilial variation in reproductive structures and provide a framework for interpreting stigma and style characters within the phylogeny of Alismataceae. These findings also contribute to a broader understanding of gynoecium evolution in early-diverging monocots.
Embryo rescue is a crucial technique for developing new seedless grape germplasm; however, its current efficiency remains suboptimal. Optimizing the culture medium is essential for the success of embryo rescue, as it directly influences the development and germination of in vitro embryos. Therefore, this study systematically investigated the effects of parental genotypes and the addition of zeatin (ZT) to the culture medium on the efficiency of embryo rescue, using four self-pollinated maternal parents and 23 seedless×seeded (or seedless) cross combinations. The results showed that among the four self-pollinated parental lines, hybrid ovules exhibited the highest development rate in the ER medium, while a higher proportion of normal plantlets was obtained in the MM3 medium. Although the addition of ZT at various concentrations did not significantly enhance the embryo development rate in the self-pollinated maternal parents, it significantly improved germination outcomes: compared to the control, the rate of normal plantlet formation increased by 8.84–19.03
Low temperature stress (LTS) is a major abiotic constraint that limits wheat (Triticum aestivum L.) growth, physiological performanceand yield particularly as winter severity increases under climate change . This study evaluated seven wheat cultivars, namely VL-2015, VL-1080, VL-967, VL-2028, VL-953, VL-826, and VL-907, under controlled LTS conditions (12/8°C, day/night) to identify contrasting tolerance levels and to examine the role of exogenous hydrogen peroxide (50 µM H₂O₂) in stress mitigation. Significant cultivar dependent variations were observed across physiological and biochemical traits. An integrated tolerance matrix classified cultivars into four groups, identifying VL-2028 as the most tolerant, with superior photosynthesis, growth and antioxidant capacity coupled with minimal oxidative damage, whereas VL-953 was the most sensitive. Intermediate responses were recorded for moderately tolerant (VL-2015, VL-826, VL-1080) and moderately sensitive (VL-967, VL-907) cultivars, and these groupings were supported by heatmap based clustering. LTS reduced biomass, chlorophyll content, photochemical efficiency Fv/Fm, and ETR, while increasing proline accumulation, malondialdehydeand membrane leakage. Correlation analysis highlighted strong positive associations among growth, RWC and photosynthetic traits, whereas stress indicators showed negative relationships. Antioxidant enzymes such as CAT and APX were closely associated with stress responsive traits. Principal component analysis explained 67.1
The currently known favorable properties of soybean have become the target of fraud in meat production. The use of plant-based raw materials in the adulteration of meat products is increasingly recognized by consumers, manufacturers, and researchers. However, the toxicity of soybeans has not been addressed. This article aimed to focus on the following: detection, quantification of soybean adulteration in meat products, analyzing how other adulterants could impact cholesterol levels in the products, and discussing how soybean adulteration could cause conditions such as allergies and toxicity. Utilizing immunohistochemical techniques, 450 different meat product samples were analyzed. Highlighting the confirmed soybean adulteration by analyzing cholesterol levels using HPLC. Results showed that, across all tested cases, soybean toxic doses ranged from 62% to 85%. Meanwhile, there were no indications on their respective nutritional facts labels. Surprisingly, the cholesterol levels were lower than those reported by previous investigators.
Pathogenic fungi that cause rust adversely affect plants, including crops, affecting them differentially depending on the cultivar. This is the case of blackberry (Rubus spp.), a member of Rosaceae, introduced in Brazil by Embrapa during the 1980s and 1990s. Given the advantageous nutritional properties of its fruits and the relatively low management costs associated with its cultivation, blackberry production has emerged as a significant alternative agricultural source for family farmers. We studied nonsymptomatic leaves of Rubus spp. and those infected with Kuehneola uredinis to evaluate structural and histochemical alterations induced by the rust fungi and compared the responses among the three blackberry cultivars. Kuehneola urenidis colonizes the mesophyll region through hyphal growth, leading to the formation of a pseudoparenchyma and to cell hypertrophy. As colonization progresses, the epidermis becomes ruptured, leading to the production of urediniospores and teliospores. While the cultivars exhibit similar anatomical characteristics, their profiles diverge in the presence of K. uredinis. The Guarani cultivar exhibits more intense tissue expansion responses in the presence of the fungus, with an increase in total leaf blade thickness (15%) and adaxial epidermis thickness (25.9%). On the other hand, the BRS Xavante cultivar suffers the greatest structural losses, showing a reduction in mesophyll (9%) and a drastic 40% drop in palisade parenchyma thickness. Finally, the BRS Tupy cultivar proved to be more stable, maintaining better tissue preservation. In regions colonized by K. uredinis, the histochemical profile undergoes significant changes, characterized by the accumulation of phenolic compounds, proteins, lipids, and essential oils, compared with those in nonsymptomatic regions, although essential oils were less detected in the more susceptible BRS Xavante. Ultimately, our results confirm varying degrees of susceptibility among the cultivars, providing insights for future research and breeding programs aimed at developing rust-resistant blackberry varieties.
