Search Results (2,650)

Inositol hexakisphosphate (InsP₆) is the most abundant inositol polyphosphate both in plant and animal cells. Exogenous InsP₆ is known to inhibit cell proliferation and induce apoptosis in cancerous cells. However, cellular entry of exogenous InsP₆ is hindered due to the presence of highly negative charge on this molecule. Therefore, to enhance the cellular delivery of InsP₆ in cancerous cells, InsP₆ was encapsulated by chitosan (CS), a natural polysaccharide, via the ionic gelation method. Our hypothesis is that encapsulated InsP₆ will enter the cell more efficiently to trigger its apoptotic effects. The incorporation of InsP₆ into CS was optimized by varying the ratios of the two and confirmed by InsP₆ analysis via polyacrylamide gel electrophoresis (PAGE) and atomic absorption spectrophotometry (AAS). The complex was further characterized by Scanning Electron Microscopy (SEM) and Fourier Transform Infrared Spectroscopy (FTIR) for physicochemical changes. The data indicated morphological changes and changes in the spectral properties of the complex upon encapsulation. The encapsulated InsP₆ enters human breast cancer MCF-7 cells more efficiently than free InsP₆ and triggers apoptosis via a mechanism involving the production of reactive oxygen species (ROS). This work has potential for developing cancer therapeutic applications utilizing natural compounds that are likely to overcome the severe toxic effects associated with synthetic chemotherapeutic drugs. Full article

The success of messenger RNA (mRNA) vaccines in controlling COVID-19 has warranted further developments in new technology. Currently, their quality control process largely relies on low-resolution electrophoresis for detecting chain breaks. Here, we present an approach using multi-primer reverse transcription sequencing (MPRT-seq) to identify degradation fragments in mRNA products. Using this in-house-made mRNA containing two antigens and untranslated regions (UTRs), we analyzed the mRNA completeness and degradation pattern at a nucleotide resolution. We then analyzed the sensitive base sequence and its correlation with the secondary structure. Our MPRT-seq mapping shows that certain sequences on the 5′ of bulge–stem–loop structures can result in preferential chain breaks. Our results agree with commonly used capillary electrophoresis (CE) integrity analysis but at a much higher resolution, and can improve mRNA stability by providing information to remove sensitive structures or sequences in the mRNA sequence design. Full article

High concentrations of advanced glycation end products (AGEs) have been linked to diseases, including diabetic complications. The pathophysiological effects of AGEs are mainly due to oxidative stress and inflammatory processes. Among the proteins most affected by glycation are albumin, the most abundant circulating protein, and collagen, which has a long biological half-life and is abundant in the extracellular matrix. The potential cellular damage caused by AGEs underscores the importance of identifying and developing natural AGE inhibitors. Indeed, despite initial promise, many synthetic inhibitors have been withdrawn from clinical trials due to issues such as cytotoxicity and poor pharmacokinetics. In contrast, natural products have shown significant potential in inhibiting AGE formation. Olea europaea L. leaves, rich in bioactive compounds like oleuropein and triterpenoids, have attracted scientific interest, emphasizing the potential of olive leaf extracts in health applications. This study investigates the anti-glycation properties of two polyphenol-rich extracts (OPA40 and OPA70) and a triterpene-enriched extract (TTP70) from olive leaves. Using in vitro protein glycation methods with bovine serum albumin (BSA)–glucose and gelatin–glucose systems, this study assesses AGE formation inhibition by these extracts through native polyacrylamide gel electrophoresis (N-PAGE) and autofluorescence detection. OPA40 and OPA70 exhibited strong, dose-dependent anti-glycation effects. These effects were corroborated by electrophoresis and further supported by similar results in a gelatin–glucose system. Additionally, TTP70 showed moderate anti-glycation activity, with a synergistic effect of its components. The results support the real possibility of using olive leaf bioproducts in ameliorating diabetic complications, contributing to sustainable bio-economy practices. Full article

The present study aimed to conduct a comparative investigation of the biological properties of phenolic and polysaccharide extracts obtained using an ultrasound-assisted technique from Aloe vera gel and their effects on each stage of the wound healing process in in vitro experimental models. HPLC analysis showed that the phenolic extract contained aloin, ferulic, and caffeic acid, as well as quercetin dihydrate, as major compounds. Capillary zone electrophoresis indicated the prevalence of mannose and glucose in the polysaccharide extract. Cell culture testing revealed the anti-inflammatory properties of the phenolic extract at a concentration of 0.25 mg/mL through significant inhibition of pro-inflammatory cytokines—up to 28% TNF-α and 11% IL-8 secretion—in inflamed THP-1-derived macrophages, while a pro-inflammatory effect was observed at 0.5 mg/mL. The phenolic extract induced 18% stimulation of L929 fibroblast proliferation at a concentration of 0.5 mg/mL, enhanced the cell migration rate by 20%, and increased collagen type I synthesis by 18%. Moreover, the phenolic extract exhibited superior antioxidant properties by scavenging free DPPH (IC₅₀ of 2.50 mg/mL) and ABTS (16.47 mM TE/g) radicals, and 46% inhibition of intracellular reactive oxygen species (ROS) production was achieved. The polysaccharide extract demonstrated a greater increase in collagen synthesis up to 25%, as well as antibacterial activity against Staphylococcus aureus with a bacteriostatic effect at 25 mg/mL and a bactericidal one at 50 mg/mL. All these findings indicate that the phenolic extract might be more beneficial in formulations intended for the initial phases of wound healing, such as inflammation and proliferation, while the polysaccharide extract could be more suitable for use during the remodeling stage. Moreover, they might be combined with other biomaterials, acting as efficient dressings with anti-inflammatory, antioxidant, and antibacterial properties for rapid recovery of chronic wounds. Full article

The vitamin D receptor (VDR) is an important candidate gene in musculoskeletal phenotypes. Polymorphisms in the VDR have been previously associated with several pathologies and muscular strength in athletes and elderly people; however, the literature reported contradictory results. The object of this research was to verify the association between the most studied VDR variants (rs2228570, rs7975232, and rs1544410) and the increase in muscle mass in elite young soccer players. A sample of 55 soccer players (15–18 years old) from a professional team were selected for this study. DNA was extracted by the salting-out method, and polymorphisms were genotyped by PCR-RFLP, followed by 2% agarose gel electrophoresis. To test the effect of the three SNPs (single nucleotide polymorphisms), a logistic regression analysis was applied. The body composition determination was carried out through the skinfold thickness method, and the muscular area of the arm and lower limb were calculated using the Frisancho formula. All three polymorphisms met the Hardy–Weinberg equilibrium (p > 0.05) and their frequencies fell within the worldwide variability. A significant correlation between rs1544410 and the increase in calf muscle mass was observed. Individuals carrying the A allele showed higher calf muscular mass than those carrying the G allele (p = 0.034). Moreover, a haplotype analysis applied to the two SNPs in linkage disequilibrium (rs7975232 and rs1544410) showed that the AG haplotype appeared negatively correlated to the calf muscle area. In conclusion, we confirm an association between VDR polymorphisms and muscular mass that could encourage the genetic screening of the VDR gene to identify a potential risk of injury and for individual nutritional interventions. Full article

The present study was undertaken to evaluate cypermethrin (CYP)-induced oxidative stress [reactive oxygen species (ROS) and lipid peroxidation (LPO) in gills, muscles, brain, and liver tissues] and DNA damage/genotoxicity (peripheral blood erythrocytes) in a freshwater teleost rohu (Labeo rohita) and the protective role of vitamin C. The LC₅₀ of CYP against rohu was found to be 4.5 µg/L in a semi-static culture system through probit analysis. Fingerlings of rohu were distributed into four groups (Group 1st served as a control, fed 35% protein basal diet and was not exposed to CYP; Group 2nd was fed a basal diet and exposed to CYP; Group 3rd and Group 4th were fed diets supplemented with vitamin C at the rate of 100 and 200 mg/kg diet, respectively, and exposed to CYP). Fingerlings were reared on a basal and vitamin C-supplemented diet for 28 days prior to exposure to CYP. The results indicate a time-dependent significant increase in ROS and LPO (indicated by time course increase in TBARS level) as well as DNA damage in terms of number of comets, % DNA in tail, tail moment, tail length, and olive tail moment after exposure to LC₅₀ of CYP. However, statistically comparable results in both Groups 1st and 4th indicate the protective role of vitamin C. The results reveal the effectiveness of vitamin C as a feed additive for countering pesticides toxicity in Labeo rohita. The current study indicates CYP as a potential genotoxicant for fish and classifies SCGE as a reliable and sensitive tool for assessing DNA damage. Full article

Understanding the epidemiology of mecA-positive Staphylococcus pseudintermedius strains, including those that are oxacillin-susceptible but potentially inducible to resistance, is crucial for developing effective treatment strategies and mitigating public health risks. This study characterized 87 mecA-positive S. pseudintermedius isolates obtained from skin lesions and nasal orifices of 46 dogs with pyoderma enrolled at a referral hospital in Thailand between 2019 and 2020. All isolates underwent antibiogram profiling, SCCmec typing, and pulsed-field gel electrophoresis (PFGE) for phenotypic and genetic analysis. Among the 87 isolates, 33 isolates (37.9%) recovered from 15 dogs were oxacillin-resistant (OR-MRSP), while 54 isolates (62.1%) from 31 dogs were oxacillin-susceptible (OS-MRSP). All OR-MRSP isolates exhibited multidrug resistance (MDR), and 44% of the OS-MRSP isolates also showed MDR. SCCmec typing revealed type V as predominant among OR-MRSP isolates (69.7%), while many oxacillin-susceptible isolates (70.4%) were non-typeable. The OR-MRSP isolates from the same dog showed consistent antibiogram and SCCmec types, while OS-MRSP isolates displayed both identical and diverse patterns. No dominant pulsotypes were observed among the OR-MRSP or OS-MRSP strains. Genetic diversity was also noted among the isolates within the same dogs and among the others, highlighting the complexity of S. pseudintermedius colonization and infection dynamics in pyoderma-affected dogs. Full article

Vibrio parahaemolyticus is an important foodborne bacterium that causes severe gastroenteritis following the consumption of contaminated seafood. To identify V. parahaemolyticus and determine its pathogenicity, the U.S. Food and Drug Administration (FDA)’s Bacteriological Analytical Manual (BAM) recommends a multiplex polymerase chain reaction (PCR) protocol to simultaneously detect the species-specific thermolabile hemolysin (tlh) gene and the pathogenic thermostable-related hemolysin (trh) and thermostable-direct hemolysin (tdh) genes. However, this assay has shown two limitations: difficulty in separating the amplicons of the trh (486 bp) and tlh (450 bp) genes due to their highly similar sizes, and the weaker band exhibited by the tdh gene amplicon (270 bp). The present study aimed to improve the BAM’s multiplex PCR assay by separating the three amplicons with similar intensity. A new primer set was applied for the tlh gene (369 bp) alongside the existing primers for the trh and tdh genes. The amplicons for the three genes were effectively separated by electrophoresis on a 2% tris-borate-EDTA (TBE) agarose gel within 45 min. Primer concentrations of 0.25 µM for three genes produced a significant amount of amplicons among various combinations of primer concentrations with 35 PCR cycles. This assay exhibited a detection limit of 10 pg of bacterial DNA, demonstrating its high sensitivity. It did not display amplicons from nine Vibrio species known to be human pathogens or from 18 well-documented foodborne pathogens. Therefore, the present multiplex PCR protocol could help overcome the limitations of existing assays and provide a more reliable method for detecting the three genes of V. parahaemolyticus. Full article

Frailty is a vulnerable state that marks the transition to long-term care for older people. Early detection and prevention of sarcopenia, the main symptom of frailty, are important to ensure an excellent quality of life for older people. Recently, the relationship between frailty, sarcopenia, and oral function has been attracting attention. This study aimed to clarify the changes in metabolites and metabolic pathways due to aging in the masseter muscle of senescence-accelerated mouse-prone 8 (SAMP8) mice. A capillary electrophoresis-mass spectrometry metabolome analysis was performed on the masseter muscle of 12-week-old, 40-week-old, and 55-week-old mice. The expression of enzymes involved in metabolome pathways considered to be related to aging was confirmed using reverse transcription polymerase chain reaction. Clear metabolic fluctuations were observed between 12, 40-week-old, and 55-week-old SAMP8 mice. The extracted metabolic pathways were the glycolysis, polyamine metabolome, and purine metabolome pathways. Nine fluctuated metabolites were common among the groups. Spermidine and Val were increased, which was regarded as a characteristic change in the masseter muscle due to aging. In conclusion, the age-related metabolic pathways in SAMP8 mice were the glycolysis, polyamine metabolome, and purine metabolome pathways. The increased spermidine and Val levels in the masseter muscle compared with the lower limbs are characteristic changes. Full article

Mutagenesis is an important tool in crop improvement and free of the regulatory restrictions imposed on genetically modified organisms. Barley (Hordeum vulgare L.) is a diploid species with a genome smaller than those of other members of the Triticeae crops, making it an attractive model for genetic studies in Triticeae crops. In this study, we report an ethyl methane sulfonate (EMS)-mutagenized population in the Chinese barley landrace TX9425, which is tolerant to both abiotic and biotic stress. A TILLING (Targeting Induced Locus Lesion in Genomes) population consisting of 2000 M₂ lines was also constructed based on the CEL I enzyme with subsequent polyacrylamide electrophoresis, which decreased the cost and labor investment. The mutant phenotypes of the M₂ and M₃ generations were scored and revealed the presence of a wide spectrum of morphological diversity. The population was evaluated by screening for induced mutations in five genes of interest. A detailed analysis was performed for the HvGLR3.5 gene and three mutations were identified by screening in 2000 M₂ lines. Two of three mutations displayed tuft and yellow striped leaves compared to the wild type. Altogether, our study shows the efficiency of screening and the great potential of the new TILLING population for genetic studies in the barley crop model system. Full article

Introduction: Hematopoietic stem cell transplantation (HSCT) comprises one of the two main treatment regimens for patients with mucopolysaccharidoses (MPS). There is a scarcity of literature concerning the process of growth in children with Mucopolysaccharidosis type I (MPS I) and Mucopolysaccharidosis type I (MPS II) after HSCT. The aim of this manuscript was to evaluate the therapeutic effect of HSCT on the heights of patients with MPS I and MPS II. Material and methods: It was an observational, single-center study on patients with MPS I and II treated with HSCT. Results: 6 MPS patients, including 4 MPS I and 2 MPS II, underwent HSCT at a median age of 2 years. All patients are alive to date, with a median age of 7.7 years (range 5.5–12 years) at the last follow-up. In both (MPS I and MPS II) groups of patients treated with HSCT, the growth rate was higher than in untreated patients and was found to be in line with the population norm. In both MPS I and MPS II patients who were treated with HSCT, normalization of urinary GAG excretion was observed. Additionally, no bands of DS and HS in GAG electrophoresis were visible. Conclusions: Both MPS I and MPS II patients presented height gain after HSCT compared to the curves of untreated patients. The absence of dermatan sulphate after HSCT could lead to normal growth in bone length. Full article

The Ikaros zinc finger factor 1 is a transcription factor with a well-known role in B- and T-cell development. The deletions of IKZF1 have an established significance in acute lymphoblastic leukemia, while reports on its prevalence and prognostic significance among ALL subtypes and regions vary. Breakpoint-specific qPCR is a practical method for testing of the most frequent types of IKZF1 deletions, considering there is clustering of the deletion events. The most commonly reported deletions are Δ4–7, Δ4–8, Δ2–7, and Δ2–8, with deletion Δ4–7 being the most common one. We retrospectively administered a breakpoint-specific qPCR design for screening for the most frequent types of IKZF1 deletions to 78 ALL patients that were diagnosed and treated between 2010 and 2022. We observed the products through gel electrophoresis, and we conducted descriptive statistics, EFS, and OS analyses. Our study found 19 patients with IKZF1 deletions, with two subjects manifesting more than one deletion. The prevalence in the different subgroups was as follows: Ph/+/ B-ALL 46%, Ph/−/ B-ALL 30%, T-ALL/LBL 4%. There was a statistically significant difference in EFS of 39 vs. 0% in favor of patients without deletions (p = 0.000), which translated to a difference in OS of 49 vs. 0% (p = 0.001). This difference was preserved in the subgroup of Ph/−/ B-ALL, while there was no significant difference in the Ph/+/ B-ALL. The most frequently observed type of deletion (15 out of 19) was the Δ4–7. There is a strong negative prognostic impact of the IKZF1 deletions at diagnosis in the observed population. IKZF1 deletion testing through breakpoint-specific qPCR is a practical approach in diagnostic testing for this risk factor. IKZF1 deletions may warrant treatment decisions and intensified treatment strategies to overcome the negative prognostic impact. Full article

R-loops, structures that play a crucial role in various biological processes, are integral to gene expression, the maintenance of genome stability, and the formation of epigenomic signatures. When these R-loops are deregulated, they can contribute to the development of serious health conditions, including cancer and neurodegenerative diseases. The detection of R-loops is a complex process that involves several approaches. These include S9.6 antibody- or RNAse H-based immunoprecipitation, non-denaturing bisulfite footprinting, gel electrophoresis, and electron microscopy. Each of these methods offers unique insights into the nature and behavior of R-loops. In our study, we introduce a novel protocol that has been developed based on a single-molecule DNA combing assay. This innovative approach allows for the direct and simultaneous visualization of RNA:DNA hybrids and replication forks, providing a more comprehensive understanding of these structures. Our findings confirm the transcriptional origin of the hybrids, adding to the body of knowledge about their formation. Furthermore, we demonstrate that these hybrids have an inhibitory effect on the progression of replication forks, highlighting their potential impact on DNA replication and cellular function. Full article

Continuous probabilistic genotyping (PG) provides a means to estimate the probative value of DNA mixtures tendered as evidence in court and subject to alternative propositions about the contributors to the mixtures. The weight of that evidence, however, may be valued differently, depending on which forensic laboratory undertook the DNA analysis. There is a need, therefore, to have a means for the comparison of likelihood ratios (LRs) generated by continuous PG amongst different laboratories for the same initial DNA sample. Such a comparison would enable the courts and the public to make judgements about the reliability of this type of evidence. There are particular mixtures and methods for which such a comparison is meaningful, and this study explores them for the short tandem repeat (STR) electropherograms of two-person mixtures obtained from the PROVEDIt Database. We demonstrate a common maximum attainable LR for a given set of common STR loci and a given DNA mixture that is consistent across three different STR profiling assays and two different capillary electrophoresis instruments. Full article

Background/Objectives: The objective of this study was to investigate the metabolomic profiles of patients with colorectal cancer (CRC) across various stages of the disease. Methods: The plasma samples were obtained from 255 subjects, including patients with CRC in stages I-IV, polyps, and controls. We employed capillary electrophoresis time-of-flight mass spectrometry and liquid chromatography triple quadrupole mass spectrometry to analyze hydrophilic metabolites comprehensively. The data were randomly divided into two groups, and consistent differences observed in both groups were analyzed. Results: Acetylated polyamines, such as N¹-acetylspermine and N¹, N¹²-diacetylspermine, consistently showed elevated concentrations in stage IV compared to stages I-III. Non-acetylated polyamines, including spermine and spermidine, exhibited increasing trends from polyp to stage IV. Other metabolites, such as histidine and o-acetylcarnitine, showed decreasing trends across stages. While acetylated polyamines have been reported as CRC detection markers, our findings suggest that they also possess diagnostic potential for distinguishing stage IV from other stages. Conclusions: This study showed stage-specific changes in metabolic profiles, including polyamines, of colorectal cancer. Full article