Search Results (138)

‘Korlaxiangli’ pear occupies a pivotal position in the pear industry of Xinjiang, with both its cultivation area and total output ranking first in the region. However, ‘Korlaxiangli’ is vulnerable to freezing injury and has suffered frequent frost damage in recent years. A cold-tolerant superior individual plant was identified via preliminary field surveys and designated as cold-tolerant superior line 15-25-15 (abbreviated as ‘HY15’). To facilitate the large-scale application of this germplasm, tissue culture technology was adopted for its rapid propagation. Using spring-collected young shoots and autumn-collected dormant buds as explants, the optimal proliferation medium was determined to be DKW supplemented with 1.0 mg/L of 6-BA and 0.5 mg/L of NAA, while the optimal rooting medium was identified as 1/2 MS containing 2.5 mg/L of NAA and 15 g/L of sucrose. Transcriptome analysis revealed eight candidate genes potentially associated with shoot proliferation, among which ARF3 and ARR12 might be associated with high proliferation efficiency. This study provides a theoretical foundation and practical technical support for the conservation, genetic improvement, and establishment of efficient propagation systems of elite pear germplasm resources. Full article

Background/Objectives: Histological examination constitutes a fundamental methodology for establishing the vitality of a lesion. In cases where the corpse is preserved for an extended duration of time prior to the post-mortem evaluation, particularly if the body has undergone freezing and thawing cycles, post-mortem changes may obscure or alter evidence of traumatic injuries. Consequently, the reliability of hematoxylin and eosin (H&E) staining for the reliable detection of intralesional erythrocytes in suspected traumatic fatalities is potentially severely compromised. The primary objective of this study is to rigorously underscore the detrimental influence of freeze–thaw processes on histologic examination and to advocate the indispensable incorporation of immunohistochemical analysis, specifically employing anti-human glycophorin A antibodies, to ascertain the presence of red blood cells. Methods: Skin samples from 10 autopsy cases were subjected to serial freeze–thaw cycles and analyzed using anti-human Glycophorin A (GPA) immunohistochemistry staining to evaluate skin lesion vitality in freeze–thawed tissues compared to fresh controls. Results: Results indicated that while H&E reliability was limited to fresh tissue, anti-GPA staining remained stable across all freeze–thaw cycles. Conclusions: Forensic pathologists must remain acutely cognizant of the potential artifacts produced by freeze–thaw cycles. In these cases, anti-GPA staining proved to be a reliable asset for evaluating the vitality of a lesion. Full article

Skeletal muscle injuries are common and some are able to regenerate due to satellite cells, the muscle stem cell population. However, in cases of severe muscle injury, complete tears, or muscle loss via trauma, muscles can undergo fibrosis and long-term compromise of their structure and function. The development of animal models has been key to understanding the pathways involved in muscle injury, fibrosis, and repair. In this review, we discuss the animal models currently used, with a focus on those most applicable to studying muscle fibrosis after traumatic injury. We summarize the approach, findings, and limitations of the most widely used models, including volumetric muscle loss, laceration, and myotoxin injection studies, and provide a brief description of ischemia/reperfusion, crush injury, freeze injury, and dystrophy models. We summarize the histological, cellular, molecular, and functional outcome measures commonly used in the field and outline areas for translation and future work. An expansion of current studies to specifically focus on muscle fibrosis will surely elucidate novel mechanisms for reducing debilitating fibrosis and promoting regeneration. Full article

Short-term hypothermic storage at 4 °C represents a promising non-freezing alternative for transporting bovine embryos and synchronizing assisted reproductive procedures. However, chilling induces oxidative stress, mitochondrial dysfunction, and apoptosis, which markedly impair post-preservation embryonic viability. Glutathione (GSH), a key intracellular antioxidant, may mitigate these damaging effects, yet its protective mechanisms during bovine blastocyst hypothermic preservation remain unclear. Here, we investigated the impact of exogenous GSH supplementation on the survival, hatching ability, cellular integrity, mitochondrial function, and developmental potential of bovine blastocysts preserved at 4 °C for seven days. Optimization experiments revealed that 4 mM GSH provided the highest post-chilling survival and hatching rates. Using DCFH-DA, TUNEL, and γ-H2AX staining, we demonstrated that 4 °C preservation significantly increased intracellular reactive oxygen species (ROS), DNA fragmentation, and apoptosis. GSH supplementation markedly alleviated oxidative injury, reduced apoptotic cell ratio, and decreased DNA double-strand breaks. MitoTracker and JC-1 staining indicated severe chilling-induced mitochondrial suppression, including decreased mitochondrial activity and membrane potential (ΔΨm), which were largely restored by GSH. Gene expression analyses further revealed that chilling downregulated antioxidant genes (SOD2, GPX1, TFAM, NRF2), pluripotency markers (POU5F1, NANOG), and IFNT, while upregulating apoptotic genes (BAX, CASP3). GSH effectively reversed these alterations and normalized the BAX/BCL2 ratio. Moreover, SOX2/CDX2 immunostaining, total cell number, and ICM/TE ratio confirmed improved embryonic structural integrity and developmental competence. Collectively, our findings demonstrate that exogenous GSH protects bovine blastocysts from chilling injury by suppressing ROS accumulation, stabilizing mitochondrial function, reducing apoptosis, and restoring developmental potential. This study provides a mechanistic foundation for improving 4 °C embryo storage strategies in bovine reproductive biotechnology. Full article

Cryotherapy and radiofrequency (RF) treatments modulate tissue temperature to induce therapeutic effects; however, improper application can result in thermal injury. Traditional temperature-based monitoring methods rely on multiple thermal sensors whose accuracy strongly depends on their number and spatial positioning, often failing to detect early tissue crystallization. This study introduces a fractional order bioimpedance modelling framework for the early detection of tissue freezing during cryogenic and thermal medical treatments, with the feasibility and effectiveness of this approach having been reported in our prior publications. While bioimpedance spectroscopy itself is a well-est. The corresponablished technique in biomedical engineering, its novel application to predict and identify premature freezing events provides a new pathway for safe and efficient energy-based therapies. Fractional-order models derived from the Cole family accurately reproduce the complex electrical behavior of biological tissues using fewer parameters than classical integer-order models, thus reducing both hardware requirements and computational cost. Experimental impedance data from human abdominal, gluteal, and femoral regions were modelled to extract fractional parameters that serve as sensitive indicators of phase-transition onset. The results demonstrate that the proposed approach enables real-time identification of freezing-induced electrical transitions, offering a physiologically grounded alternative to conventional temperature-based monitoring. Furthermore, the fractional order bioimpedance method exhibits high reproducibility and selectivity, and its analytical figures of merit, including the limits of detection and quantification, support its use for reliable real-time tissue monitoring and early injury detection. Overall, the proposed fractional order bioimpedance framework enhances both safety and control precision in cryogenic and thermal medical applications. Full article

Cryotherapy involves flash freezing of tissue and removing unwanted tissue. Mechanism of injury is causing cell membrane rupture by rapid multiple freeze–thaw cycles, while reserving tissue architecture and the collagen matrix. This promotes favorable wound healing. In recent years, it has gained increasing attention as a treatment option for upper gastrointestinal diseases (Barrett’s Esophagus and early cancer). Currently, two FDA-approved delivery methods are available in the GI tract: Cryoballoon and spray cryotherapy, which will be discussed. In this review, we also propose to examine the expanding role of cryotherapy in gastrointestinal practice, drawing from both clinical studies and illustrative vignettes. In addition, we will highlight its established role in eradicating Barrett’s with low and high-grade dysplasia and compare its outcomes and safety profile with radiofrequency ablation (RFA). We will also discuss the application and safety of spray cryotherapy in the palliation of malignant esophageal strictures when compared with Esophageal stent placement. Cryotherapy may have immunological potential, and it may shrink both primary and metastatic diseases. Ongoing research in this field of Cryo-immunology will be highlighted. Beyond esophageal neoplasia, cryotherapy is increasingly utilized in other upper gastrointestinal precancerous conditions. Through this synthesis, our goal is to provide a timely and comprehensive overview of advancements in cryotherapy and its potential to reshape novel therapeutic approaches in upper gastrointestinal cancers. Finally, we highlight the evolution of a novel platform using nitrous oxide delivered by a handheld device, a contact balloon, and a small replaceable cartridge. This approach may make delivery of cryogen application favorable and a first-line approach in the management of Barrett’s esophagus and early cancer. In addition, Cryoballoon therapy for dysphagia palliation for malignant esophageal strictures may become a preferred approach as more data evolves. Full article

During their natural growth, plants encounter adverse environmental conditions, such as chilling injury, freezing injury, drought, and salt damage, collectively known as abiotic stresses. Several studies have shown that WRKY proteins regulate various abiotic stress responses and plant developmental processes. However, researchers have rarely investigated WRKY genes associated with the stress response in apples. Within this research, Malus baccata (L.) Borkh as the experimental material. We isolated and cloned MbWRKY63 and investigated its function in low-temperature stress tolerance. Subcellular localization analysis shows that MbWRKY63 localizes to the cell nucleus. Tissue-specific expression analysis revealed that MbWRKY63 is relatively highly expressed in the young leaves and root tissues of apples. Under low-temperature treatment at 4 °C, Arabidopsis thaliana plants that overexpressed MbWRKY63 showed greater cold stress resistance than the wild type (WT) and the empty vector (UL) control. In transgenic plants, the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) were significantly enhanced; meanwhile, the contents of proline, malondialdehyde (MDA), and chlorophyll also changed significantly. In addition, by regulating the expression levels of AtKIN1, AtCBF1, AtCBF2, AtCBF3, AtCOR47, and AtCOR15a, MbWRKY63 enhanced the low-temperature stress tolerance in transgenic Arabidopsis. The results suggest that MbWRKY63 in apples may be involved in the response to low-temperature stress, laying a foundation for understanding the role of WRKY transcription factors (TFs) in abiotic stress responses. Full article

Commercial turkey breeding relies almost entirely on artificial insemination, yet avian sperm are unusually vulnerable to cooling and freezing injury. As a result, extender chemistry and processing steps, especially low-temperature equilibration, are pivotal for post-thaw performance. We evaluated how extender choice, paired with equilibration time, shapes the post-thaw quality of turkey semen. Ejaculates were diluted in Beltsville, Sperm Motility Medium (SMM), Botucrio, or Kobidil⁺, then equilibrated for 20 or 40 min before freezing; samples were cryostored for one month and assessed immediately after thawing. The outcomes included motility/kinematics, membrane integrity, mitochondrial activity and membrane potential, apoptosis/necrosis, reactive oxygen species (ROS), DNA fragmentation, and bacteriological load. Overall, 20 min equilibration improved post-thaw motility and membrane integrity, and reduced DNA fragmentation and ROS. Among extenders, Beltsville delivered the best overall sperm quality. Considering the extender × time interaction, Beltsville, Botucrio, and Kobidil⁺ performed best at 20 min, whereas SMM performed best at 40 min. Thus, Beltsville and SMM provide strong, time-specific options for turkey semen cryopreservation—Beltsville at 20 min and SMM at 40 min. Full article

Tendon injuries affect millions of people globally and are among the most prevalent musculoskeletal conditions, frequently resulting in chronic pain, reduced mobility, and functional impairment. While conservative and surgical treatments are available, limitations such as low healing capacity, scar formation, and reduced biomechanics necessitate alternative approaches. Tissue engineering offers a promising solution by combining cells, scaffolds, and bioactive molecules to regenerate tendon tissue. This review presents key concepts and emerging trends, highlighting the cellular components, scaffold materials, and manufacturing processes. Tenocytes and mesenchymal stem cells are fundamental for tissue regeneration, as they synthesize extracellular matrix components and regulate inflammatory responses. Various natural and synthetic polymers have been fabricated into scaffolds that mimic the structure and biomechanics of natural tendons. Composite and hybrid scaffolds are utilized to improve the biocompatibility of natural materials with the mechanical stability of synthetic materials. Advanced technologies, such as electrospinning, freeze-drying, and 3D bioprinting, enable the creation of scaffolds with defined architecture and functional gradients, improving cell alignment, differentiation, and tendon–bone integration. Although promising preclinical data exists, major challenges remain in translating these strategies clinically, particularly vascularization, immune rejection, and mechanical stability. Continued interdisciplinary attempts in biomaterials science, cellular biology, and engineering are crucial to advancing clinically viable tendon tissue engineering. Full article

Anubias (Araceae) is a globally important group of ornamental aquatic plants. However, when temperatures drop to 10 °C, most species suffer obvious frostbite from cold stress, restricting winter cultivation and broader application. This study focused on two Anubias genotypes with distinct cold tolerance, adopting an integrated approach combining phenotypic, physiological, metabolomic, and transcriptomic analyses to reveal the mechanisms underlying their differential cold tolerance. Under 10 °C cold stress, compared with normal temperatures, the leaves of cold-tolerant Anubias sp. ‘Long Leaf’ (Jian) showed no significant frostbite, while cold-sensitive Anubias barteri var. nana ‘Coin Leaf’ (Jin) had clear frost damage. Both genotypes exhibited increased leaf relative electrical conductivity, malondialdehyde (MDA) content, soluble sugar content, and activities of superoxide dismutase (SOD) and catalase (CAT); “Jian” had more notable rises in SOD/CAT activities and maintained higher levels, whereas “Jin” showed greater increases in conductivity, MDA, and soluble sugar. Metabolomic and transcriptomic analyses revealed “Jian” specifically upregulated metabolites in pathways like flavone and flavonol biosynthesis and tryptophan metabolism, as well as genes related to valine, leucine, isoleucine degradation and phenylpropanoid biosynthesis pathways. ERFs, WRKYs, NACs and other transcription factors correlated with these differentially expressed genes, suggesting potential transcriptional regulation. These results provides insights for breeding cold-tolerant Anubias and optimizing low-temperature cultivation. Full article

Late spring frosts represent a major threat to grapevine (Vitis vinifera L.), a risk increasingly exacerbated by climate change-driven shifts in phenology. To explore sustainable strategies for frost mitigation, this study investigated the effect of a natural polysaccharide-based biogel, derived from carob (Ceratonia siliqua L.), on the molecular response of grapevine buds exposed to severe cold stress. To this aim, a preliminary RNA-Seq analysis was carried out to compare the transcriptomes of biogel-treated frozen buds (BIOGEL), untreated frozen buds (NTF), and unstressed controls (TNT). The transcriptomic analysis revealed extensive reprogramming of gene expression under freezing stress, highlighting the involvement of pathways related to membrane stabilization, osmotic adjustment, and metabolic regulation. Interestingly, the biogel treatment appeared to attenuate the modulation of several cold-responsive genes, particularly those associated with membrane functionality. Based on these preliminary transcriptomic data, twelve candidate genes, representative of the functional classes affected by biogel treatment, were selected for qRT-PCR validation. The expression patterns confirmed the RNA-Seq trends, further suggesting that biogel application might mitigate the typical transcriptional activation induced by frost, while supporting genes involved in cellular protection and integrity maintenance. The overall analyses suggest that the biogel may act through a dual mechanism: (i) providing a physical barrier that reduces cold-induced cellular damage and stress perception, and (ii) promoting a selective adjustment of gene expression that restrains excessive defense activation while enhancing membrane stability. Although further field validation is required, this natural and biodegradable formulation represents a promising and sustainable tool for mitigating late frost injuries in viticulture. Full article

In various European regions, white lupin (Lupinus albus L) production could increase by autumn sowing of winter-hardy varieties. This study aimed to explore the genetic variation, the genetic architecture, and the genomic prediction of frost resistance in two reference populations, one including 144 landrace and cultivar genotypes, and the other comprising 144 breeding lines. These populations were genotyped by 40,914 and 32,951 SNP markers, respectively, issued by genotyping-by-sequencing. The genotypes were phenotyped for mortality and a biomass injury score at freezing temperature of −11 °C under controlled conditions. Both traits, highly correlated, exhibited large genetic variation and high broad-sense heritability (H² = 0.76–0.82). A genome-wide association study highlighted their polygenic architecture and detected markers linked to candidate genes. The intra-population predictive ability of plant mortality achieved 0.41 for landrace and cultivar germplasm, and 0.67 for breeding lines. The cross-population predictive ability was higher when using the model constructed for landrace and cultivar germplasm to predict breeding lines (0.39) than the reverse (0.26). Landrace field survival was largely influenced by late phenology in addition to frost resistance. Our results revealed frost-resistant germplasm, confirmed the polygenic control of frost resistance, and highlighted genomic prediction opportunities for line selection and the identification of elite genetic resources. Full article

Background/Objectives: Urinary angiotensin-converting enzyme (uACE) activity has long been regarded as a promising biomarker for kidney and cardiovascular diseases; however, its clinical applicability has been limited by the presence of endogenous urinary inhibitors and technically demanding assay protocols. We aimed to establish a fast and reproducible method for measuring uACE activity to identify the inhibitory compounds responsible for previous assay failures and to define practical preanalytical conditions suitable for routine laboratory implementation. Methods: A fluorescence-based kinetic assay was optimized for urine samples. Endogenous inhibitors were isolated by membrane filtration and chemically characterized, while the effect of sample dilution was evaluated as a simplified alternative for eliminating inhibitory interference. We assessed the stability of ACE activity under various storage conditions to support reliable measurement. Results: Urea (IC₅₀ = 1.18 M), uric acid (IC₅₀ = 3.61 × 10⁻³ M), and urobilinogen (IC₅₀ = 2.98 × 10⁻⁴ M) were identified as the principal reversible inhibitors, jointly accounting for up to 90% suppression of uACE activity. Their inhibitory effect was effectively eliminated by a 128-fold dilution. ACE activity remained stable for 24 h at 25 °C but was completely lost after freezing. A strong positive correlation between uACE activity and creatinine concentration (r = 0.76, p < 0.0001) justified normalization. ACE activity-to-creatinine ratio turned out to be significantly lower in ACE inhibitor-treated patients than in untreated controls (6.49 vs. 36.69 U/mol, p < 0.0001). Conclusions: Our findings demonstrate that accurate measurement of uACE activity is feasible using a rapid dilution-based protocol. The normalized ACE activity can serve as a practical biomarker for detecting pharmacological ACE inhibition and monitoring therapy adherence in cardiovascular care and may also provide insight into renal pathophysiology such as tubular injury or local RAAS-related processes. Full article

To investigate how sweating–drying processing affects the components, antioxidant activity, and hepatoprotective mechanisms of Eucommia ulmoides (EUB) against acute liver injury (ALI), this study constructed a “processing–active components–ALI targets” network. Eight processed EUB samples were analyzed using HPLC fingerprinting, multi-assay antioxidant tests (DPPH/ABTS·+/pyrogallol), network pharmacology, and molecular docking. Sweating–drying significantly altered EUB’s chemical profile, with HPLC fingerprint similarities ranging from 0.715 to 1.000, the lowest being for FG4 (40 °C dried after sweating) and FD (freeze-dried after sweating). Key components (chlorogenic acid (CA), pinoresinol diglucoside (PDG), aucubin (AU), geniposidic acid (GPA)) varied: XS (sun-dried) had the highest CA/PDG, while FG4 showed increased AU/GPA. FY (shade-dried after sweating) exhibited the strongest free radical scavenging (DPPH/ABTS·+/pyrogallol IC₅₀ = 0.828, 0.134, 14.200 mg/mL), which correlated with CA/PDG/liriodendrin (PD) synergy. Network pharmacology identified 205 EUB-ALI intersection targets (core: TNF, PTGS2, GAPDH) and the AGE-RAGE pathway; molecular docking confirmed strong CA/PDG binding to GAPDH/PTGS2. This study clarifies how processing regulates EUB’s components and their links to antioxidant and hepatoprotective effects, providing scientific support for EUB’s clinical application against ALI. Full article

Background: Freeze tolerance is an uncommon but highly effective strategy that allows certain vertebrates to survive prolonged exposure to subzero temperatures in a frozen, ischemic state. While past studies have characterized the metabolic and biochemical adaptations involved, including cryoprotectant accumulation and metabolic rate suppression, the contribution of post-transcriptional gene regulation by microRNAs (miRNAs) remains largely unexplored. This study investigated freeze-responsive miRNAs in cardiac tissue of the gray tree frog, Dryophytes versicolor, to better understand the molecular mechanisms that support ischemic survival and tissue preservation. Methods: Adult frogs were subjected to controlled freezing at −2.5 °C, and cardiac tissue was collected from frozen and control animals. Total RNA was extracted and analyzed via small RNA sequencing to identify differentially expressed miRNAs, followed by target gene prediction and KEGG pathway enrichment analysis. Results: A total of 3 miRNAs were differentially expressed during freezing, with significant upregulation of miR-93-5p and let-7b-5p and downregulation of miR-4485-3p. Predicted targets of upregulated miRNAs included genes involved in immune signaling pathways (e.g., cytokine–cytokine receptor interaction), steroid hormone biosynthesis, and neuroactive ligand–receptor interaction, suggesting suppression of energetically costly signaling processes. Downregulation of miRNAs targeting cell cycle, insulin signaling, and WNT pathways indicates possible selective preservation of cytoprotective and repair functions. Conclusion: Overall, these results suggest that D. versicolor employs miRNA-mediated regulatory networks to support metabolic suppression, maintain essential signaling, and prevent damage during prolonged cardiac arrest. This work expands our understanding of freeze tolerance at the molecular level and may offer insights into biomedical strategies for cryopreservation and ischemia–reperfusion injury. Full article