Three Capsular Polysaccharide Synthesis-Related Glucosyltransferases, GT-1, GT-2 and WcaJ, Are Associated With Virulence and Phage Sensitivity of Klebsiella pneumoniae

Klebsiella pneumoniae (K. pneumoniae) spp. are important nosocomial and community-acquired opportunistic pathogens, which cause various infections. We observed that K. pneumoniae strain K7 abruptly mutates to rough-type phage-resistant phenotype upon treatment with phage GH-K3. In the present study, the rough-type phage-resistant mutant named K7RR showed much lower virulence than K7. Liquid chromatography-tandem mass spectrometry (LC-MS-MS) analysis indicated that WcaJ and two undefined glycosyltransferases (GTs)- named GT-1, GT-2- were found to be down-regulated drastically in K7RR as compared to K7 strain. GT-1, GT-2, and wcaJ are all located in the gene cluster of capsular polysaccharide (CPS). Upon deletion, even of single component, of GT-1, GT-2, and wcaJ resulted clearly in significant decline of CPS synthesis with concomitant development of GH-K3 resistance and decline of virulence of K. pneumoniae, indicating that all these three GTs are more likely involved in maintenance of phage sensitivity and bacterial virulence. Additionally, K7RR and GT-deficient strains were found sensitive to endocytosis of macrophages. Mitogen-activated protein kinase (MAPK) signaling pathway of macrophages was significantly activated by K7RR and GT-deficient strains comparing with that of K7. Interestingly, in the presence of macromolecular CPS residues (>250 KD), K7(ΔGT-1) and K7(ΔwcaJ) could still be bounded by GH-K3, though with a modest adsorption efficiency, and showed minor virulence, suggesting that the CPS residues accumulated upon deletion of GT-1 or wcaJ did retain phage binding sites as well maintain mild virulence. In brief, our study defines, for the first time, the potential roles of GT-1, GT-2, and WcaJ in K. pneumoniae in bacterial virulence and generation of rough-type mutation under the pressure of bacteriophage

Plumbagin ameliorates ferroptosis of ovarian granulosa cells in polycystic ovary syndrome by down-regulating SLC7A5 m6A methylation modification through inhibition of YTHDF1

Abstract Background and objective Polycystic ovary syndrome (PCOS) is a common endocrine-metabolic disease in women of reproductive age. One of its core pathologies is ovarian granulosa cell (GC) dysfunction, and ferroptosis, as a novel cell death mode dependent on iron ions and lipid peroxidation, may be involved in the PCOS process, but the exact mechanism is unknown. Plumbagin (PLB) shows potential in PCOS treatment due to its antioxidant properties. The present study aimed to elucidate the molecular mechanisms by which PLB ameliorates mitochondrial dysfunction and ferroptosis in PCOS GCs through the YTH N6-methyladenosine RNA binding protein 1/L-type amino acid transporter 1 (YTHDF1/SLC7A5) axis. Methods An in vitro model of PCOS was constructed by treating KGN cells with dihydrotestosterone (DHT), and PLB treatment, YTHDF1 knockdown (si-YTHDF1), and SLC7A5 overexpression (pcDNA 3.1-SLC7A5) were intervened respectively. Cell viability was measured by cell counting kit-8. Lactate dehydrogenase (LDH) release, adenosine triphosphate (ATP) level, iron ion, and lipid peroxidation (LPO) content were detected by commercial kits. Mitochondrial membrane potential (MMP) was analyzed by JC-1 staining combined with flow cytometry. Reactive oxygen species (ROS) levels were assessed by C11-BODIPY probe, oxidative stress indicators including malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase were measured by kits, and Cytochrome C, Ferritin, mitochondrial transcription factor A (TFAM), glutathione peroxidase 4 (GPX4) and SLC7A5 expression were detected by Western blot. Fluorescence in situ hybridization, RNA immunoprecipitation, and m6A quantitative real-time polymerase chain reaction verified the interaction and translational regulation of YTHDF1 and SLC7A5. Results DHT treatment significantly decreased KGN cell viability, MMP and ATP levels, increased LDH release, ROS, MDA, iron ions and LPO content, up-regulated Cytochrome C expression, and down-regulated Ferritin, TFAM, and GPX4 expression. Both PLB treatment and YTHDF1 knockdown significantly reversed the above changes, but YTHDF1 overexpression reversed the protective effect of PLB. YTHDF1 co-localized with SLC7A5 mRNA and enhanced its translation through m6A modification. YTHDF1 knockdown reduced SLC7A5 protein levels without affecting mRNA expression. SLC7A5 overexpression weakened the protective effect of YTHDF1 knockdown, resulting in decreased cell viability, deterioration of mitochondrial function, and increased ferroptosis. Conclusion PLB ameliorates DHT-induced mitochondrial dysfunction and ferroptosis in KGN cells by inhibiting YTHDF1 expression, and its action is dependent on the mechanism by which YTHDF1 regulates SLC7A5 translation through m6A modification. Downregulating YTHDF1 or SLC7A5 significantly enhances GC survival and function

Preparation of Am-MSN/PVDF Mixed Matrix Membranes for Enhanced Removal of Reactive Black 5

The discharge of large volumes of textile dyeing wastewater, characterized by poor biodegradability and high toxicity, poses severe threats to the environment. In this study, polyvinylidene difluoride (PVDF) membranes were prepared using the nonsolvent-induced phase separation (NIPS) method, with porous amino-functionalized mesoporous silica nanoparticles (Am-MSNs) mixed into the casting solution to fabricate the Am-MSN/PVDF mixed matrix membranes. By varying the amount of Am-MSNs added, the microstructure and overall performance of the membranes were comprehensively analyzed. The results demonstrated that the addition of Am-MSNs significantly enhanced the hydrophilicity of the membranes. The high specific surface area and amino groups of Am-MSNs facilitated interactions with dye molecules, such as Reactive Black 5 (RB5), through hydrogen bonding, electrostatic attraction, and physical adsorption, resulting in a marked improvement in RB5 rejection rates. Static adsorption tests further validated the superior adsorption capacity of the Am-MSN/PVDF mixed matrix membranes for RB5. Additionally, the nanoscale mesoporous structure of Am-MSNs enhanced the mechanical strength of the membranes. The synergistic effects of the mesoporous structure and amino groups significantly increased the efficiency and stability of the Am-MSN/PVDF mixed matrix membranes in dye removal applications, providing an effective and sustainable solution for the treatment of dye-contaminated wastewater

Exploring the dilemma of enterprises participating in the old community renewal: Perspective of managers

Exploring the dilemma of enterprises participating in the old community renewal: Perspective of manager

The Antifungal Activity of Loquat (Eriobotrya japonica Lindl.) Leaves Extract Against Penicillium digitatum

This study was performed to determine the antifungal activity of loquat (Eriobotrya japonica Lindl) leaf extract (LLE) against the citrus postharvest pathogen Penicillium digitatum (P. digitatum). The LLE exhibited an antifungal activity against P. digitatum, with a minimum inhibitory concentration (MIC) of 0.625 mg/ml and a minimum fungicidal concentration (MFC) of 1.25 mg/ml. Significant inhibitory effects of LLE on mycelial growth and spore germination of P. digitatum were seen in a dose-dependent manner. Simultaneously, to investigate possible antifungal mechanisms by LLE, we analyzed their influence on morphological changes, cell membrane permeability, cell wall and cell membrane integrity, and adenosine phosphates (ATP, ADP, and AMP) levels. Alterations, such as sunken surface and malformation, occurred in the LLE-treated P. digitatum spores. Furthermore, intracellular inclusion content decreased after LLE treatment, indicating an increase in cell membrane permeability. Besides, the LLE treatment induced a significant decline in the level of adenosine monophosphate (AMP), adenosine diphosphate (ADP), and adenosine triphosphate (ATP) with a noticeable addition of extracellular ATP, ADP, and AMP during the entire treatment period. Overall, the results manifested that the antifungal activity of LLE against P. digitatum can be attributed to the derangement of cell membrane permeability and disordered energy metabolism. This is the first report on the mechanism of antifungal activity of LLE and could be useful in the development of targeted fungicides from natural origin.</jats:p

Glucuronidation and its effect on the bioactivity of amentoflavone, a biflavonoid from <i>Ginkgo biloba</i> leaves

Abstract Objectives Ginkgo biloba leaves contain amentoflavone (AMF), a dietary flavonoid that possesses antioxidant and anticancer activity. Flavonoids are extensively subjected to glucuronidation. This study aimed to determine the metabolic profile of AMF and the effect of glucuronidation on AMF bioactivity. Methods A pharmacokinetic study was conducted to determine the plasma concentrations of AMF and its metabolites. The metabolic profile of AMF was elucidated using different species of microsomes. The antioxidant activity of AMF metabolites was determined using DPPH/ABTS radical and nitric oxide assays. The anticancer activity of AMF metabolites was evaluated in U87MG/U251 cells. Key findings Pharmacokinetic studies indicated that the oral bioavailability of AMF was 0.06 ± 0.04%, and the area under the curve of the glucuronidated AMF metabolites (410.938 ± 62.219 ng/ml h) was significantly higher than that of AMF (194.509 ± 16.915 ng/ml h). UGT1A1 and UGT1A3 greatly metabolized AMF. No significant difference was observed in the antioxidant activity between AMF and its metabolites. The anticancer activity of AMF metabolites significantly decreased. Conclusions A low AMF bioavailability was due to extensive glucuronidation, which was mediated by UGT1A1 and UGT1A3. Glucuronidated AMF metabolites had the same antioxidant but had a lower anticancer activity than that of AMF. </jats:sec

Comparison of miRNA and mRNA Expression in Sika Deer Testes With Age

To elucidate the complex physiological process of testis development and spermatogenesis in Sika deer, this study evaluated the changes of miRNA and mRNA profiles in the four developmental stages of testis in the juvenile (1-year-old), adolescence (3-year-old), adult (5-year-old), and aged (10-year-old) stages. The results showed that a total of 198 mature, 66 novel miRNAs, and 23,558 differentially expressed (DE) unigenes were obtained; 14,918 (8,413 up and 6,505 down), 4,988 (2,453 up and 2,535 down), and 5,681 (2,929 up and 2,752 down) DE unigenes, as well as 88 (43 up and 45 down), 102 (44 up and 58 down), and 54 (18 up and 36 down) DE miRNAs were identified in 3- vs. 1-, 5- vs. 3-, and 10- vs. 5-year-old testes, respectively. By integrating miRNA and mRNA expression profiles, we predicted 10,790 mRNA–mRNA and 69,883 miRNA–mRNA interaction sites. The target genes were enriched by GO and KEGG pathways to obtain DE mRNA (IGF1R, ALKBH5, Piwil, HIF1A, BRDT, etc.) and DE miRNA (miR-140, miR-145, miR-7, miR-26a, etc.), which play an important role in testis development and spermatogenesis. The data show that DE miRNAs could regulate testis developmental and spermatogenesis through signaling pathways, including the MAPK signaling pathway, p53 signaling pathway, PI3K-Akt signaling pathway, Hippo signaling pathway, etc. miR-140 was confirmed to directly target mutant IGF1R-3′UTR by the Luciferase reporter assays. This study provides a useful resource for future studies on the role of miRNA regulation in testis development and spermatogenesis.</jats:p