Background: Significant alterations in feeding, housing, and physiology are observed in dairy cows during the transition period (3 weeks pre- and post-calving), in addition to changes in the composition and abundance of the endometrial microbiota. Thus, this study aimed to evaluate any changes in the composition and predicted metabolic pathways in the cow uterine microbiome during this transition period. Methods: Scrapings were sampled from the endometrial surface of clinically healthy cows (n = 3) in dynamics as follows: in the 10 Days period before, and on Days 3, 5, and 20 after calving. Total DNA was isolated from the samples, and the composition of the microbial community was assessed using targeted next-generation sequencing (NGS) technology. Based on the subsequent NGS data, the dynamics of the predicted metabolic pathways of the microbiota were evaluated. Results: Seven superphyla and phyla of microorganisms were found in the endometrial microbiota of cows during the transition period. Among these, the phylum Firmicutes (with a dominant class of Clostridia) and the superphylum Fusobacteriota (represented by a single class of Fusobacteriia) can be considered the dominant bacteria in the endometrium, with representation noted from 25.2 to 68.2% and from 12.3 to 51.1%, respectively. The microbiome composition underwent significant changes (p < 0.05) during the transition period. In particular, the high abundance of the Fusobacteriaceae family (up to 68.2%) in the uterus of clinically healthy cows was unexpected, given the potential association of Fusobacteriaceae with the occurrence of metritis in cows. The numbers of microorganisms in two dominant classes, Fusobacteriia and Clostridia, showed generally opposite changes in their relative abundance during the transition period. The predicted functional potential level for 32 pathways in the endometrium changed (p < 0.05) in cows during the transition period. Indeed, the activity of the predicted pathways, such as pyridoxal 5′-phosphate biosynthesis I and teichoic acid (poly-glycerol) biosynthesis, was lowered on day 3 postpartum (p < 0.05). Conclusions: Microbiota composition and the activity of the predicted metabolic pathways in the cow endometrium underwent significant changes at different critical stages in the transition period. Moreover, even clinically healthy cows exhibited signs of dysbiotic disorders.
BACKGROUND:Transient transformation is a convenient and less time-consuming method for investigating gene functions compared with production of stable transformants. Most systems for Agrobacterium-mediated plant transformation, regardless of transient or stable transformation, utilize a combination of Agrobacterium strains carrying a helper Ti-plasmid and a binary vector. However, the helper Ti-plasmids which are mega-plasmids with sizes of 200-250 kbp and are difficult to manipulate directly with conventional molecular cloning techniques due to their large sizes. METHODS:A small helper Ti-plasmid, pCU307D with a size of 46 kbp was constructed from pTiEHA101 which is commonly used for plant genetic engineering. They consist of the virulence (vir) region, the mutated replication origin from pTiEHA101, the gentamicin resistance marker and the replication origin for E. coli cells. The abilities of T-DNA transfer were examined by transient expression in Arabidopsis cultured cells. RESULTS:Efficiencies to transfer T-DNA to plant cells by EHA101 and C58C1 carrying pCU307D were similar, although pCU307D had 2.8-fold higher copy numbers than EHA101. The construction processes of pCU307D eliminated the A281virF gene encoding F-box-like protein which was located outside of the vir region. The A281virF gene was cloned into a binary vector and it was introduced into C58C1 cells with pCU307D which was named as CU307DF. T-DNA transfer efficiencies by EHA101 or CU307DF were examined by GUS activities from transiently transformed T-DNA with the CaMV35S::NLS-GUS cassette in Arabidopsis cells. Co-culture with CU307DF carrying the GUS gene conferred 3.2-fold higher GUS activities, compared to that with its parental strain EHA101. CONCLUSION:A new Agrobacterium strain CU307DF has higher capacity for T-DNA transfer, compared with its parental strain EHA101. pCU307D is as small as 46 kbp and can propagate in E. coli, and that may enable to add further modification to it for further improvement of T-DNA transfer by CU307DF.
Background: Adeno-associated viruses (AAVs) are established vectors for efficient gene delivery to the central nervous system (CNS). Increasingly, strategies aim to restrict transduction to specific neuronal subtypes defined by the associated functional properties, thereby enhancing precision and therapeutic potential. Methods: Recombinant AAV9 vectors carrying fluorescent reporters under the control of cytomegalovirus (CMV), human synapsin (hSyn), or homeobox 9 (Hb9) promoters were delivered intrathecally in Wistar rats. Transgene expression was evaluated 7 days post-injection by confocal microscopy. Neurons in laminae VII–X were quantified across cervical, thoracic, lumbar, and sacral spinal cord levels. Statistical analysis was performed using the Kruskal–Wallis test followed by Mann–Whitney U tests with Bonferroni correction. Results: In lamina VII, consistent neuronal expression was mediated by hSyn across all spinal levels, with significantly higher transduction at cervical compared to thoracic and lumbar regions (p < 0.01). CMV and Hb9 showed no detectable tropism for this lamina. In lamina VIII, CMV drove markedly higher expression than hSyn and Hb9, with a 2.8-fold difference at the lumbar level (p < 0.001). In lamina X, CMV expression exceeded hSyn at the lumbar and sacral levels (p < 0.05), while Hb9 showed no activity. In lamina IX, all promoters mediated motoneuron transduction, but only Hb9 restricted expression specifically to motoneuron somata. Notably, CMV induced off-target expression in glial cells. Conclusions: AAV9-mediated expression patterns in the spinal cord are strongly shaped by promoter choice and segmental level. Hb9 provides high motoneuron specificity, hSyn supports broad neuronal activation across laminae VII–X, whereas CMV drives robust but non-specific expression with significant off-target activity. These findings highlight the importance of rational promoter selection for spinal cord gene therapy and strategies aimed at functional recovery in motor system disorders.
Excessive fructose consumption has emerged as a critical driver of obesity and metabolic dysfunction, with far-reaching implications for multiple organ systems. This review synthesizes current evidence on the biochemical and molecular pathways underlying fructose induced disease mechanisms, discussing how fructose metabolism activates the “survival switch”, promotes fat storage, and generates uric acid, mitochondrial dysfunction, and oxidative stress, thereby disrupting energy homeostasis. Key organ-specific consequences are explored, including hepatic steatosis and progression to non-alcoholic fatty liver disease, pancreatic β-cell dysfunction, renal fibrosis, intestinal barrier disruption with microbial dysbiosis, cardiometabolic impairment, pulmonary inflammation, and neurocognitive decline with relevance to Alzheimer’s disease. Moreover, mechanistic insights highlight the role of fructokinase C activation, adenosine triphosphate (ATP) depletion, leptin resistance, pro-inflammatory signaling (mechanistic target of rapamycin complex-1 (mTORC1), renin angiotensin system (RAS), Toll-like receptor 4 (TLR4)), and cross-talk between fructose metabolism and organ-specific pathophysiology. Animal and human studies consistently reinforce the central role of fructose overload in driving obesity and associated complications. Meanwhile, this review frames fructose not merely as a caloric contributor but as a metabolic disruptor, thereby underscoring the urgent need for public health interventions, dietary regulation, and mechanistic research to mitigate fructose-driven metabolic disease.
Background: Nitrosylcobalamin (NO-Cbl) is a vitamin B12 analog designed to exploit the “Trojan horse” vulnerability created by the heightened need of cancer cells for cobalamin and one-carbon metabolism. Building on our recent biophysical studies confirming the affinity of NO-Cbl for intrinsic factor, this work aimed to investigate the mechanistic basis for the selective anticancer activity of NO-Cbl through the cobalamin transport axis and lysosomal processing. Materials and Methods: Human cancer cell lines (NIH-OVCAR-3, MCF-7, WM9, and DU145) were cultured and transfected to overexpress transcobalamin II (TCII). Cell proliferation and cytotoxicity were measured using the sulforhodamine B (SRB) assay. TCII-R (CD320) expression was quantified by flow cytometry. The impact of anti-CD320 antiserum and lysosomal alkalization (chloroquine) on NO-Cbl activity was assessed. Results: Antiserum inhibition of the TCII receptor resulted in dose-dependent inhibition of NIH-OVCAR-3 and MCF-7 cell proliferation. Lysosomal alkalinization by chloroquine pretreatment abrogated NO-Cbl-induced cytotoxicity in OVCAR-3 cells. Flow cytometric analysis demonstrated an inverse correlation between TCII-R (CD320) expression (MFI ratio) and NO-Cbl ID50. TCII overexpression significantly reduced NO-Cbl ID50 in NIH-OVCAR-3 cells. Conclusion: NO-Cbl utilizes tumor cell cobalamin transport and processing pathways to deliver nitric oxide selectively to cancer cells. These results, integrated with recent binding studies, validate NO-Cbl as a cobalamin-based targeted anticancer agent with efficacy in tumors expressing high levels of TCII and CD320.
Artificial corneas represent a significant breakthrough in addressing global corneal blindness, impacting millions of individuals worldwide. The scarcity of donor tissue and the complications of immune rejection necessitate the development of synthetic alternatives. This review examines key innovations in biomaterials, scaffold design, and regenerative medicine that have informed the development of artificial corneas. Recent studies have demonstrated that polyethylene glycol (PEG)-based hydrogels exhibit 98% light transmittance and an elastic modulus of 1.5 MPa, whereas collagen scaffolds achieve 85% clinical success with <5% inflammatory response. Graphene oxide-based nanocomposites have increased mechanical strength by 25%. Therefore, by synthesizing clinical and preclinical evidence, this article outlines current achievements and unresolved challenges related to scalability, cost, immune compatibility, and regulatory constraints, providing a roadmap for future translational research in corneal tissue engineering.
Background: Deuterium is unevenly distributed in natural waters, while the same applies to the content of deuterium in ice on Mars. Moreover, changes in the deuterium content of drinking water are known to affect the bodies of mammals. Thus, since plans are in place to send people to Mars in the coming years, understanding the effects of water with a Martian isotopic composition is necessary. Therefore, this study aimed to evaluate the impact of water with an increased deuterium content of 1200 ppm on the dynamics of indicators in the body of mammals. Methods: The study was conducted on Wistar rats. The metabolic profile of blood and the content of deuterium in it were studied in dynamics by days using nuclear magnetic resonance (NMR) spectroscopy. Additionally, the isotopic composition of brain tissue was studied in dynamics by days using isotope mass spectrometry. A further study was conducted on the functioning of the antioxidant system in blood plasma and brain tissue using PCR analysis, chemiluminescence, and biochemical analysis methods; the intestinal microbiome was also studied. The durations of the animal experiments were 31 (blood and brain study) and 38 (stress-protective activity study) days. Results: On day 23, the deuterium content in the blood plasma increased to 856 parts per million (ppm), and to 260 ppm in the brain on day 31. This increase led to an imbalance in the antioxidant/prooxidant processes. This effect was accompanied by shifts in the intensity of oxidative processes, alongside changes in enzyme activity and the expression of genes responsible for their synthesis, shifts in amino acid composition, and changes in the concentration of metabolites and microbiome molecules in the blood plasma. By the fifth and eighth days, the number of Bacteroides in the intestines had decreased by 14% and 21.8%, respectively, compared to the values measured on day zero of the experiment. Meanwhile, the population of Firmicutes-type bacteria increased by 12% and 16% on the fifth and eighth days, respectively, compared to the indicators measured on day zero of the experiment. Conclusion: An increase in the concentration of deuterium in the body promotes the development of a stress reaction and the activation of compensatory mechanisms aimed at adaptation.
Background: The global rise of antimicrobial resistance necessitates the development of innovative therapeutic strategies beyond traditional antibiotics. Drug repurposing offers a time- and cost-effective approach by identifying new antimicrobial applications for existing medications. Thus, this study aimed to investigate the antimicrobial and anti-virulence potential of several clinically approved drugs, including fluconazole, buspirone, duloxetine, escitalopram, and finasteride. Methods: We evaluated the antimicrobial efficacy of the selected compounds against a panel of microorganisms comprising two Gram-negative bacteria (Escherichia coli, Serratia marcescens), two Gram-positive bacteria (Bacillus megaterium, Staphylococcus epidermidis), and two opportunistic yeasts (Candida albicans, Rhodotorula mucilaginosa). Antimicrobial activity was evaluated using growth inhibition and viability assays. Additionally, we investigated the effects of the selected drugs on fungal virulence traits, including biofilm formation and filamentation, and assessed infectivity using a Caenorhabditis elegans host model. Results: Duloxetine and escitalopram demonstrated broad-spectrum antimicrobial activity, inhibiting bacterial and fungal growth at concentrations below 512 mg/L. Buspirone exhibited selective antimicrobial effects, particularly against Gram-positive bacteria and C. albicans. Although finasteride exhibited limited direct antifungal activity, it significantly disrupted key virulence traits in yeasts, including biofilm formation, morphological transitions, and host infection capacity. Conclusion: These findings underscore the potential of serotonin reuptake inhibitors and finasteride as candidates for antimicrobial repurposing. By impairing both microbial viability and pathogenicity, these drugs may provide promising avenues for developing adjunct or alternative therapies against resistant bacterial and fungal pathogens.
Acute myeloid leukemia (AML) is a hematologic malignancy with a poor prognosis and high relapse rates, especially in high-risk patients and older adults. Conventional treatment modalities confer limited benefit, specifically in relapsed and refractory cases. Antibody drug conjugates (ADCs) are a rapidly advancing treatment option that provides a novel approach to treating AML. The design and mechanistic aspects of ADCs have also been discussed. ADCs combine cytotoxic chemotherapeutic drugs with the specificity of monoclonal antibodies. This review primarily focuses on the current role of ADCs in the treatment of AML, including approved agents such as gemtuzumab ozogamicin, as well as others. Moreover, challenges associated with the use of ADCs have been explored, including resistance mechanisms, drug stability, immunogenicity, and cost. This review also highlights and summarizes various ongoing and completed clinical trials, which may provide insight into this treatment approach. Future advancements in AML treatment, including the use of nanoparticles or nanostructures, have also been discussed. In conclusion, this comprehensive review sheds light on the current and prospective future directions of ADCs in the treatment of AML, highlighting their potential to significantly alter the therapeutic landscape for this cancer.
Pseudomonas taetrolens is a highly versatile microorganism that has gained significant attention in biotechnology due to its metabolic adaptability and ability to thrive in diverse environmental conditions. Thus, since P. taetrolens efficiently metabolizes organic compounds, P. taetrolens is a promising candidate for sustainable industrial applications. P. taetrolens demonstrates substantial potential in waste utilization by converting dairy byproducts, such as whey, into value-added compounds, including lactobionic acid, thereby advancing the principles of circular economic frameworks. This review provides a comprehensive analysis of the characterization, structural features, and diverse applications of P. taetrolens. In particular, this review explores the enzymatic mechanisms underlying the metabolic pathways of P. taetrolens, focusing on its role in lactose oxidation into lactobionic acid. Additionally, this review underscores the potential of microorganisms in industrial biotechnology and sustainable development practices by showcasing recent advances and ongoing research. This analysis demonstrates how P. taetrolens contributes to innovative solutions in waste utilization, environmental sustainability, and the production of value-added compounds across sectors, including food, pharmaceuticals, and cosmetics.
Background: The unsafe disposal of milk processing effluents has a negative impact on the environment due to their high content of nutrients and organic matter. Green alternatives can be applied to effectively manage and valorize these effluents, reducing their environmental footprint. Methods: The ability of the free-living cyanobacterium Oscillatoria sp. to grow in real cheese whey was evaluated as a potential strategy for integrating dairy wastewater treatment with biomass valorization. Autotrophic and mixotrophic cultures were maintained under controlled laboratory conditions and monitored over 28 days for growth, cell viability, biomass, pigment content, and physicochemical parameters, including pH, protein, carbohydrate, and chemical oxygen demand (COD). Results: Oscillatoria sp. successfully adapted to the initial acidic conditions of the effluent (pH 2.8–2.9), increasing the pH of the treated whey to levels suitable for industrial wastewater disposal (pH 6.0–9.0). A 5-fold increase in dehydrogenase activity was observed after a 28-day culture, with no signs of oxidative damage. Cyanobacterial biomass cultivated under mixotrophic conditions displayed a significant reduction (∼55%) in photosynthetic pigments, including chlorophyll a and total carotenoids, compared to autotrophic cultures. Notably, Oscillatoria sp. biomass increased by 2.3-fold under mixotrophy, compared to the autotrophic control. The higher biomass production was accompanied by a significant reduction in the whey COD from 35,250 mg/L to 8500 mg/L, along with a 65% and 80% decrease in protein and carbohydrate content, respectively. Conclusions: These findings provide new insights into the metabolic behavior of Oscillatoria sp. during cheese whey bioremediation, highlighting the potential of mixotrophic cyanobacteria for managing dairy wastewater management.
The brain malignant tumor Glioblastoma multiforme (GBM) has a median survival of 14–16 months using current treatments; thus, understanding the pathology of GBM is crucial for proposing new therapies and increasing overall survival outcomes. Therefore, this study aimed to analyze different elements, particularly growth factors and the related signal transduction pathways, which play a role in brain neoplastic development, from stem cells to established solid brain tumors, and the application of current immunology techniques, molecular biology, and nanotechnology. Targeting growth factors, especially insulin-like growth factor-1 (IGF-I) (the principal neoplastic development factor) using anti-gene technologies—antisense and triple helix—has previously been shown to produce an immune anti-tumor response (CD8, CD28) through the TK/PI3K/AKT pathway. This immune response was increased using phytochemicals (phenolics), especially nanoparticles (theranostic nanoparticles), by modulating IGF-I through common pathways (IGF-I-R and TK/PI3K/AKT/TLR/MAPK and JAK/STAT). This review demonstrates how studies on central nervous system neoplastic development progressively led to establishing clinical cancer gene therapies, increasing GBM survival by 20–24 months. The presented studies compare the results of cancer gene therapy with other current immunotherapies. Moreover, this research chapter briefly describes the investigations of nanotechnology related to neurotumorigenesis and GBM therapies. The presented studies relate to nanotechnology and compare the results of cancer gene therapy with other current immunotherapies.
Coccinia grandis (L.) Voigt (ivy gourd) is popularly consumed in South Asia for food and therapeutic purposes. C. grandis acts as a remedy for various ailments, such as hypertension, diabetes, cancer, ulcers, diarrhea, jaundice, inflammation, fever, bronchitis, burns, skin eruptions, insect bites, allergies, eye infections, and urinary disorders. Researchers have identified phytoconstituents in diverse chemical classes from this species, including alkaloids, flavonoids, coumarins, esters, ethers, fatty acids, fatty alcohols, terpenoids, and phenolic compounds. Comprehensive research conducted in vitro and in vivo has confirmed the properties of the plant as antidiabetic, anticancer, antiparasitic, antimicrobial, hepatoprotective, analgesic, antipyretic, anti-Alzheimer's, anticataract, antileishmanial, anti-anaphylactic, anti-histaminic, and wound-healing agent, as well as being advantageous for cardiovascular health. Most pharmacological findings are derived from studies on the extracts and the subsequent phytoconstituents from this plant species. Nevertheless, the specific phytoconstituents underlying these biological effects and the mechanisms of action involved are yet to be fully identified. Toxicological evaluations indicate that C. grandis is generally safe, although high doses can cause dose-dependent hepatotoxicity. Moreover, the clinical trials focusing on the antidiabetic effects of C. grandis demonstrate promising effects in managing glucose dysregulation. This review aims to provide a comprehensive update on C. grandis, expanding on previous studies by incorporating a broader ethnomedicinal scope, a more extensive phytochemical profile with detailed chemical structures, and additional clinical trial data. Unlike prior publications, this review emphasizes C. grandis as a functional food, highlighting its potential in chronic disease management. By integrating these aspects, this study offers a more in-depth analysis of the therapeutic potential and future applications of this plant. The functional food aspect of C. grandis, rich in bioactive compounds, supports its role in preventing and managing chronic diseases as a regular vegetable.
BACKGROUND:The E. coli O157:H7 strain has been the subject of many studies. In addition to producing severe abdominal illness in humans and animals, the E. coli O157:H7 strain is characterized by the production of Shiga toxins and demonstrates resistance to multiple antibiotics. METHODS:In this study, 20 fecal samples from patients with typical symptoms of E. coli O157:H7 infection and 20 from animals that tested positive for the same pathogen were analyzed. The bacterium was isolated, identified, and classified using both culture-based and molecular methods, employing the rpoB, stx, waa, and waaO genes. RESULTS:The E. coli O157:H7 strain classification was highly similar to the E. coli O157:H7 strain Sakai. The rpoB, stx, waa, and waaO genes were deposited on the NCBI website under accession numbers PP059841, OR939814, PP059843, and PP059842, respectively. The mutant sequences at the waa sites K, L, and Y were analyzed to determine the alterations in the associated gene function, cell wall formation, and the ability of the mutant E. coli O157:H7 to develop antibiotic resistance compared to the wild-type. CONCLUSIONS:Antibiotic resistance in the mutant E. coli O157:H7 increased significantly regarding some type of theses antimicrobial agents, while in some cases it decreased. This depends on the type of antibiotics and its mode of action and target. This may be explained by the waaK and waaL genes, which prevent the entry of antimicrobial agents into the bacterial cell.
BACKGROUND:This study aimed to investigate the modeling and design of product compositions using grape stem extracts, with an emphasis on phenolic compounds and antioxidant activity. Grape stems, a significant byproduct of winemaking, are a rich source of bioactive phenolic substances. METHODS:Aqueous ethanol extracts were obtained from grape stems of European varieties; the Magarach Institute developed a new selection. Phenolic compounds were quantified using ultraviolet-visible spectrophotometry via the Folin-Ciocalteu method and high-performance liquid chromatography (HPLC). Antioxidant activity was evaluated using an amperometric method. In addition, mathematical models have been developed to predict product composition based on mixing various components. Key HPLC parameters (detection wavelength, flow rate, column temperature, injection volume) and amperometric calibration procedures are described. RESULTS:The mass concentration of phenolic substances in the extracts ranged from 7.95 to 16.40 g/dm3, and the antioxidant activity ranged from 6.30 to 13.90 g/dm3. Identifying the compositions of stem extracts was conducted with a focus on studying substances with a non-flavonoid structure: phenolic acids, esters, and stilbenes. Statistical analysis (analysis of variance (ANOVA) with post-hoc t-tests) confirmed significant differences (p < 0.05) between grape varieties. Furthermore, the developed mathematical model-validated using coefficient of determination (R2) and Root Mean Squared Error (RMSE) metrics-demonstrated robust predictive capabilities for product compositions. CONCLUSIONS:These results indicate that grape stems are a valuable source of bioactive compounds for developing functional products. In addition, the proposed mathematical modeling approach offers a reliable method for designing product compositions. These findings provide a basis for developing software tools to optimize product formulation in the food industry.
Biological therapies have transformed cancer treatment by targeting the molecular mechanisms involved in carcinogenesis. However, higher costs, limited accessibility, and supply chain disruptions-such as those caused by COVID-19 in recent years-underscore the need for cost-effective alternatives. Biosimilars, which are drugs that are highly similar to their reference biologics in terms of safety, efficacy, and quality, offer a viable solution (as these demonstrate clinically meaningful outcomes). This review article examines the role of biosimilars, mainly in gynecological cancers. The primary focus of this article is to compare the efficacy, safety, and cost-effectiveness of biosimilars, as well as to explore the barriers that restrict their widespread adoption. A comprehensive literature review was conducted, analyzing various studies, regulatory guidelines, and the latest data on biosimilars for the treatment of gynecological cancers. Pivotal trials, such as the GOG-0218, ICON7, and RUBY, were reviewed to assess the efficacy, safety, and cost-effectiveness of these biosimilars. This review highlights key oncologic therapies, including bevacizumab, trastuzumab, pembrolizumab, and their biosimilars, mainly for gynecological cancers. Additionally, this review considers the challenges of immunogenicity, interchangeability, and clinician awareness. After reviewing the latest peer-reviewed literature and related online materials, we found that biosimilars demonstrate comparable efficacy and safety to their reference biologics while also being more cost-effective. Recent clinical trials support the role of biosimilars in limiting the progression of disease and improving overall survival while reducing the financial burden of cancer treatments.
BACKGROUND:Fanconi anemia (FA) is an inherited genetic instability syndrome that increases the risk of developing head and neck squamous cell carcinoma, particularly in the oral cavity. These epithelial cancers often arise from visible oral and potentially malignant disorders (OPMD). Research has shown that oral brush biopsies combined with cytology, such as manual DNA cytometry, can facilitate the early detection of OPMDs that require treatment. Thus, this study aimed to evaluate the diagnostic accuracy of a DNA karyometry (DNA-KM) system in the brush biopsy-based diagnostic workup for OPMDs with FA. METHODS:Feulgen-stained liquid-based oral smears were included from 327 independent OPMD cases, which had available cytological diagnoses and clinicopathological reference standards. These samples were automatically analyzed using a DNA-KM system (MotiCyte-auto), which employs digital nuclear classifiers based on expert classification of nuclear images and machine learning algorithms. RESULTS:The detection of (suspected) DNA stemline aneuploidy or single-cell aneuploidy with DNA-KM demonstrated a sensitivity of 69% and a specificity of 96%. In our analysis, when DNA-KM was combined with cytology, we observed a sensitivity of 75% and a specificity of 96%. Meanwhile, additional research using the variation coefficient of a "broad-based" peritetraploid stemline (BPS) as an alternative algorithm further increased the sensitivity to 84%. However, employing this algorithm slightly decreased specificity to 92% at a cut-off of 5.83. CONCLUSIONS:Artificial intelligence (AI)-assisted DNA-KM, with automated slide-scanning and digital classification of nuclei, can serve as a valuable additional method in the brush biopsy-based cytological diagnosis of OPMD in FA. This approach can help identify lesions that require clinical intervention.
Background: Parkinson's disease (PD) is a progressive neurodegenerative disorder with which the leucine-Rich repeat kinase 2 glycine 2019 serine (LRRK2 G2019S) mutation is strongly associated. This mutation elevates kinase activity, disrupts mitochondrial function, increases reactive oxygen species (ROS) production, and impairs DNA repair mechanisms, all of which contribute to the pathogenesis of PD. Thus, addressing these pathological features through targeted delivery systems holds promise for more effective therapies. Methods: This study aimed to investigate the use of Ginkgo biloba leaf extract (EGB) to synthesize sphingomyelin-cholesterol solid lipid nanoparticles (SLNPs) functionalized with poly-L-lysine (EGB–PLL–SLNPs) for siRNA delivery targeting the LRRK2 G2019S mutation. SLNPs suspended in water (H₂O–PLL–SLNPs) served as the comparator. In vitro assays were conducted using either wild-type or LRRK2 G2019S-transformed SH-SY5Y and HEK293 cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was employed to evaluate the cytotoxicity of the SLNPs and nanocomplexes; meanwhile, flow cytometry was used to assess caspase 3/7 activity, mitochondrial membrane potential, DNA damage, and ROS levels. Results: Treatment with EGB–PLL–SLNPs significantly improved mitochondrial health, reducing depolarized and dead cells and enhancing overall cell viability. ROS levels, DNA damage and kinase activity were significantly decreased compared to the control H₂O–PLL–SLNPs. Conclusion: The enhanced therapeutic outcomes observed with the EGB–PLL–SLNPs can be attributed to the bioactive compounds in EGB, particularly the flavonoids and terpenoids, such as quercetin and kaempferol. These molecules play crucial roles in stabilizing mitochondrial membranes, facilitating ATP synthesis, and regulating genes linked to mitochondrial biogenesis. The interaction between EGB and siRNA to mediate gene silencing provides a multifaceted approach to counteracting PD pathophysiology. This study demonstrates that EGB–PLL–SLNPs offer superior gene silencing and cytoprotective effects compared to conventional formulations. The integration of plant-based bioactives with nanomedicine enhances therapeutic delivery and efficacy, positioning biosynthesized PLL–SLNPs as a promising strategy for treating Parkinson's disease.
Pesticides spread into the air, contaminate soil and water, and can affect various objects, contributing to secondary pollution regardless of the employed type or application method. Currently, organophosphorus pesticides (OPs) are widely utilized in agriculture, forestry, and livestock farming worldwide. These chemicals enter the body through multiple exposure routes and can harm the nervous system, endocrine system, and other organs. Owing to the environmental persistence and elevated toxicity exhibited by these pesticides, certain OPs are difficult to break down biologically, thus posing serious threats to human health and ecosystems. Disinfection or destruction of those pesticides remaining in the environment represents one of the important tasks scientists face. This review presents information on OPs, some of their properties, environmental impacts, and mechanisms for the effective decomposition of these pesticide residues by microorganisms. Bacteria and fungi isolated from samples contaminated with various OPs were analyzed. New metabolites formed during OP degradation by these microorganisms, as well as microbial enzymes involved in OP degradation and the molecular mechanisms of the process, are presented. The methods used in these studies and recommendations for future research are also detailed.
Background: Red clover (Trifolium pratense L.) is an important forage crop throughout the world due to its high forage quality, nitrogen fixation capacity and beneficial effects on the soil fertility. But aluminum (Al) toxicity limits significantly red clover production in acid soils, which represent more than one third of the world's agricultural lands. Natural variation for Al3+ ions resistance has been identified in many crop species so development of tolerant accessions and varieties is a promising approach for red clover breeding. In this context the objectives of this article were to select in vitro and evaluate using different DNA markers the tolerant to toxic aluminum breeding samples of red clover. Methods: Seeds of the experimental hybrid population were germinated under various aluminum concentrations, including control without aluminum. Epicotyls of seedlings without roots and with roots not less than 4–5 mm were subcultivated on agar's medium of Gamborg B5 with 2 mg/L of 6-benzylaminopurine and 100 mg/L of Al3+ and then planted in the cassettes with soil. Seedlings and adult plants F2, tolerant to 50 and 100 mg/L of Al3+ were selected, grown in vegetative pots and used further for molecular analyses. Genetic variability between tolerant and susceptible red clover genotypes was evaluated based on DNA markers: sequence-related amplified polymorphism (SRAP), retrotransposon microsatellite amplified polymorphism (REMAP) and inter-primer binding site polymorphism (iPBS). Results: Aluminum-tolerant red clover samples were obtained by in vitro selection on the medium with toxic aluminum ions. F2 seedlings in the variants with 50 mg/L and 100 mg/L of Al3+ were characterized by longer length and roots size compared with F1 seedlings and variety-standard at the same aluminum concentration. Subsequent molecular analysis showed that REMAP and iPBS were efficient markers to detect distinguishes among red clover accessions. The average level of polymorphism was identified as 45.8 using REMAP and 68.2% with iPBS; the average values of polymorphism information content (PIC) were 0.764 and 0.746 accordingly, higher compared to SRAP (0.741). Conclusions: Combination of the biotechnology methods and the current DNA-technologies based on REMAP and iPBS markers is effective approach to improve precision and reliability of selection and assessing of red clover genotypes with tolerance to toxic aluminum ions (Al3+). Breeding samples identified in this study, can be used as a promising initial material for development the new varieties with stable inheritability of the aimed trait.
