Search Results (145)

Background: Vascular calcification has been highlighted as a prognostic factor for perioperative thrombosis but a protective factor for late restenosis in lower limb peripheral artery disease (LLPAD). The aim of this study was to investigate the association between calcification and twelve-month primary patency in patients with stenting of the common femoral artery (CFA) and its bifurcation for atheromatous stenosis. Materials/Methods: This single-center retrospective study analyzed consecutive limbs (n = 90) that underwent CFA stenting for symptomatic lesions between January 2018 and January 2023. Calcification was assessed using dedicated computed tomography angiography analysis software (EndoSize; Therenva), with blinded evaluation of volume (mm³) and density (Hounsfield Units) across three anatomically distinct zones: proximal CFA (Zone 1); distal CFA (Zone 2); and bifurcation segments (Zone 3). The primary endpoint was twelve-month primary patency, defined as a peak systolic velocity ratio (PSVR) < 2.4 on duplex ultrasound without target lesion revascularization. Secondary endpoints included predictors of restenosis using multivariable logistic regression. Results: Ninety cases of CFA stenting for LLPAD (lower limb peripheral artery disease) were analyzed. A total of 78.9% of CFA lesions were treated for claudication and 21.1% for critical limb-threatening ischemia (CLTI). Lesions were distributed as Azema types I (1%), II (43%), and III (56%). At twelve-month follow-up, primary patency (PSVR < 2.4) was achieved in 77.4% of limbs. Patent CFA stenting demonstrated significantly higher median calcification density in Zone 2 compared to those with restenosis (1122 [IQR: 903–1248] vs. 858 [788–987] HU; p = 0.006; q = 0.021 after false discovery rate correction). ROC curve analysis identified a density threshold of 800 HU with a 76% reduction in restenosis risk (OR 0.24; 95% CI: 0.08–0.72; p = 0.011). Bootstrap validation (1000 replications) confirmed threshold stability at 821 HU (95% CI: 656–990 HU). Conclusions: In this exploratory study, dense calcification (≥800 HU) in the distal CFA appears to be protective against twelve-month restenosis following stenting. These findings suggest that calcification density may serve as a valuable predictor for patient selection and procedural planning in CFA interventions. Full article

Modification of synthetic oligonucleotides and DNA is widely used in many applications in the life sciences. However, in most cases, modified DNA cannot be restored to its native state. Here, we report the preparation of a thymidine-inosine dimer building block (TID) for oligonucleotide synthesis. The TID modification supports the functionalization of synthetic oligonucleotides, which can later be removed to restore the DNA strand to its native state. The TID unit allows for a wide spectrum of postsynthetic modifications of oligonucleotides through click chemistry, including conjugation with fluorescent tags and small molecules, preparation of branched oligonucleotide scaffolds, and anchoring to a solid support. Due to the modification of the thymine base, the TID unit reduces the stability of the DNA duplex. We found that the negative effect of internal TID modification on duplex stability does not exceed the same for a single base mismatch. As long as the TID modification is present in the DNA strand, it disrupts its natural functionality. The “caging” effect of TID in the template strand with respect to DNA polymerase was demonstrated in primer extension experiments. Traceless removal of the temporary functional group occurs through oxidative cleavage of the inosine subunit, resulting in the formation of a native DNA strand with the thymine base left at the cleavage site. An anthracene-modified dodecamer oligonucleotide and a branched oligonucleotide scaffold were used to study the cleavage of the reporter group or the oligonucleotide side strand, respectively. It was shown that aqueous tetramethylguanidine efficiently cleaves the oxidized inosine subunit of TID at 37 °C, forming the native DNA strand. Full article

This study evaluates the application of metal additive manufacturing—specifically the laser powder bed fusion (LPBF) process—for producing aluminium die-casting mould components, comparing 300-grade maraging steel inserts with conventional H13 tool steel. Efficient thermal management and mould durability are critical in aluminium injection moulding. Still, traditional machining limits the design of cooling channels, resulting in hot spots, accelerated wear, and a reduced service life. LPBF allows the fabrication of complex geometries, enabling conformal cooling channels to enhance thermal control. Component samples were manufactured using maraging steel via LPBF, machined to final dimensions, and subjected to duplex surface treatment (plasma nitriding + CrAlN PVD coating). Thermal performance, dimensional stability, mechanical properties, and wear resistance were experimentally assessed under conditions simulating industrial production. The results demonstrate that LPBF components with optimised cooling channels and surface engineering achieve higher thermal efficiency, an extended service life (up to 2.6×), improved hardness profiles (545 HV0.05 core, 1230 HV0.05 on nitrided surface and 2850 HV0.05 after PVD film deposition), and reduced maintenance frequencies compared to H13 inserts. The study confirms that additive manufacturing, combined with tailored surface treatments and optimised cooling design, overcomes the geometric and thermal limitations of conventional manufacturing, offering a reliable and productive solution for aluminium die-casting moulds. Full article

The geometry and quality of a weld seam are critical factors in laser beam welding, influencing mechanical performance and structural integrity. Dynamically modulated laser beams provide a precise means of tailoring energy input in high-power laser welding processes. This study investigates the influence of beam shape and modulated frequency on weld seam geometry, penetration depth, and capillary behaviour using a coherent beam combining (CBC) laser system from Civan Lasers. Three beam intensity distributions—single point, line–point–line (LPL), and boomerang—were applied across a modulation frequency range of 1, 10, and 100 kHz during the welding of duplex and austenitic stainless steels. High-speed imaging captured real-time capillary dynamics, and the data were analysed to assess capillary stability, measure capillary diameter, and determine the capillary front angle as a function of frequency and beam shape. Transverse cross-sections of the welds were prepared to evaluate seam geometry and microstructure. The results show that beam shape significantly affects energy distribution and weld profile, while modulation frequency critically influences capillary behaviour and penetration characteristics. These findings highlight the critical role of dynamic beam shaping and frequency modulation in optimizing laser welding processes for material-specific performance, offering a versatile platform for advancing precision manufacturing using CBC technology. Full article

Background/Objectives: The course of treatment for renal artery stenosis following renal transplantation depends on the severity of the condition. Mild cases are typically managed medically, while more significant stenosis with flow limitation and graft dysfunction requires percutaneous intervention. Surgical treatment is generally reserved as a last resort. This study aimed to evaluate the outcomes of interventional radiology in managing renal artery stenosis at our transplant center. Methods: The electronic medical records of patients who underwent renal transplantation at our center between January 2020 and December 2024 were reviewed to identify cases of renal artery stenosis and their subsequent management through interventional radiology. Sociodemographic and clinical data were collected for both recipients and donors. Data analysis was performed using SPSS version 26. Results: Out of the total 368 patients who received renal allograft at our center from January 2020 to December 2024, 25 patients were confirmed with duplex ultrasound to have renal artery stenosis. The majority of affected patients were African American, had Class I Obesity and presented with cardiovascular co-morbidities. The mean time from transplant to the diagnosis of RAS was 4.25 (SD ± 3.81) months. The mean serum creatinine level at presentation was 2.54 (SD ± 1.21 mg/dL). All 25 patients underwent digital subtraction angiography, and 24 patients were confirmed to have renal artery stenosis requiring further intervention. The creatinine levels at one week, three months and one year post-intervention were 2.12 (SD ± 1.00), 1.83 (SD ± 0.63) and 2.15 (SD ± 1.68) mg/dL, respectively. Conclusions: Percutaneous interventional treatment for renal artery stenosis is associated with improvements in hemodynamic parameters and the stabilization of allograft function. Follow-up is needed to monitor for the potential occurrence of restenosis. Full article

This study investigates the electroplating characteristics of Co-Zn alloy coatings with varying Zn contents (0.55 wt.%~8.8 wt.%) and their influence on intermetallic compound (IMC) formation during reactions with Sn solder. Co-Zn alloy coatings were successfully fabricated by electroplating using cobalt plating solutions with different concentrations of zinc sulfate. The results reveal anomalous co-deposition behavior, where the less noble Zn preferentially deposits over Co. Surface morphologies and microstructures evolve significantly with increasing Zn content, transitioning from columnar to dendritic structures. Zn incorporation into the Co lattice disrupts its crystallinity, leading to decreased crystallinity and partial amorphization. Liquid-state and solid-state interfacial reactions with Sn solder demonstrate that Zn content considerably influences IMC formation. In liquid-state reactions at 250 °C, lower Zn contents (0.55–4.8 wt.%) slightly enhance CoSn₃ growth. It exhibits a dense layered-structure without IMC spallation. In contrast, a higher Zn content (8.8 wt.%) significantly reduces IMC formation by approximately 50% and produces a duplex structure with two distinct layers. In solid-state reactions at 160 °C, the suppression effect becomes even more pronounced. The Co-0.55Zn deposit exhibits significant inhibition of CoSn₃ growth, while the Co-8.8Zn sample forms only a thin IMC layer, achieving a suppression rate exceeding 85%. These findings demonstrate that Zn doping effectively limits CoSn₃ formation during solid-state reactions and improves interfacial stability. Full article

The maintenance of genome stability requires the coordinated actions of multiple proteins and protein complexes. One critical family of proteins is the recombination mediators. Their role is to facilitate the formation of recombinase nucleoprotein filaments on single-stranded DNA (ssDNA). Filament formation can take place on post-replicative ssDNA gaps as well as on 3′-tailed duplexes resulting from helicase–nuclease processing. In prokaryotes, the RecF, O, and R proteins are widely distributed and mediate RecA loading as either the RecFOR or RecOR complexes, depending on the species being studied. In this review, I compare and contrast the available biochemical and structural information to provide insight into the mechanism of action of this critical family of mediators. Full article

Recent advances in molecular dynamics (MD) simulations and the introduction of artificial intelligence (AI) have resulted in a significant increase in accuracy for structure prediction. However, the cell is a highly crowded environment consisting of various macromolecules, such as proteins and nucleic acids. The macromolecular crowding and solution conditions, such as temperature, ion concentration, and the presence of crowders, significantly influence the molecular interactions between and structural changes in proteins and nucleic acids. In this study, we investigate the presence of crowders and their effect on the melting of DNA molecules by analyzing melting profiles of short and long heterogeneous DNA duplexes. In particular, we examine how multiple inert crowders, randomly distributed along long DNA chains, influence DNA melting. We find that the presence of crowders stabilizes double-stranded DNA (dsDNA), with this effect being more pronounced in short DNA duplexes. These findings complement in vitro observations and improve our understanding of dsDNA in cell-like environments. Full article

Long non-coding RNAs (lncRNAs) are transcripts over 200 nucleotides long that do not encode proteins. Although many lncRNAs remain uncharacterized, they are known to play diverse regulatory roles in gene expression. A group of lncRNAs called natural antisense transcripts can form double-stranded structures with their sense partners due to sequence complementarity. These duplexes can become substrates for A-to-I RNA editing, an epitranscriptomic modification mediated by ADAR enzymes. RNA editing is known to influence transcript splicing, affect the resulting gene expression product or alter RNA stability, all of which can impact cancer cell biology. Here, we show a novel natural antisense transcript, UGGT1-AS1, that we have identified and characterized in terms of its cellular localization and sense partner interactions. Furthermore, we demonstrate that UGGT1-AS1 affects cell proliferation and regulates the stability of the UGGT1 sense transcript. Finally, using publicly available RNA sequencing data, we identify A-to-I RNA editing events in the protein-coding gene UGGT1 and further confirm them by RT-PCR and Sanger sequencing in MCF7 cell lines. We hypothesize that UGGT1-AS1 may act as a triggering factor for the A-to-I RNA editing process in its sense partner. Our findings highlight the regulatory role of UGGT1-AS1 and suggest its involvement in RNA editing and cancer biology. Full article

R-loops are nucleic acid structures composed of an RNA-DNA hybrid (RDH) duplex and a displaced single-stranded DNA (ssDNA), which are fundamentally involved in key biological functions, including transcription and the preservation of genome stability. In an R-loop, the RDH duplex is bent by the folded secondary structures of the displaced ssDNA. Previous experiments and simulations indicated the high bendability of DNA below the persistence length. However, the bendability of a short RDH duplex remains unclear. Here, we report that an RDH duplex exhibits higher bendability than a DNA duplex on the short length scale using single-molecule cyclization experiments. Our molecular dynamics simulations show that an RDH duplex has larger intrinsic curvature and structural fluctuations and more easily forms kinks than DNA, which promote the bending flexibility of RDH from unlooped structures. Interestingly, we found that an RDH duplex composed of a C-rich DNA strand and a G-rich RNA strand shows significantly higher bendability than that composed of a G-rich DNA strand and a C-rich RNA strand in the same CpG island promoter regions, which may contribute to the formation of an R-loop. These findings shape our understanding towards biological processes involving R-loops through the high and sequence-dependent bendability of an RDH duplex. Full article

Background/Objective: Oligodeoxynucleotides (ODNs) containing base-labile modifications such as N4-acetyldeoxycytidine (4acC), N6-acetyladenosine (6acA), N2-acetylguanosine (2acG), and N4-methyoxycarbonyldeoxycytidine (4mcC) are highly challenging to synthesize because standard ODN synthesis methods require deprotection and cleavage under strongly basic and nucleophilic conditions, and there is a lack of ideal alternative methods to solve the problem. The objective of this work is to explore the capability of the recently developed 1,3-dithian-2-yl-methoxycarbonyl (Dmoc) method for the incorporation of multiple 4acC modifications into a single ODN molecule and the feasibility of using the method for the incorporation of the 6acA, 2acG and 4mcC modifications into ODNs. Methods: The sensitive ODNs were synthesized on an automated solid phase synthesizer using the Dmoc group as the linker and the methyl Dmoc (meDmoc) group for the protection of the exo-amino groups of nucleobases. Deprotection and cleavage were achieved under non-nucleophilic and weakly basic conditions. Results: The 4acC, 6acA, 2acG, and 4mcC were all found to be stable under the mild ODN deprotection and cleavage conditions. Up to four 4acC modifications were able to be incorporated into a single 19-mer ODN molecule. ODNs containing the 6acA, 2acG, and 4mcC modifications were also successfully synthesized. The ODNs were characterized using RP HPLC, capillary electrophoresis, gel electrophoresis and MALDI MS. Conclusions: Among the modified nucleotides, 4acC has been found in nature and proven beneficial to DNA duplex stability. A method for the synthesis of ODNs containing multiple 4acC modifications is expected to find applications in biological studies involving 4acC. Although 6acA, 2acG, and 4mcC have not been found in nature, a synthetic route to ODNs containing them is expected to facilitate projects aimed at studying their biophysical properties as well as their potential for antisense, RNAi, CRISPR, and mRNA therapeutic applications. Full article

Mismatched T:G base pairs can arise during de novo replication as well as base excision repair (BER). In particular, the action of the gap-filling polymerase β (Polβ) can generate a T:G pair as well as a nick in the DNA backbone. The processing of a nicked T:G mispair is poorly understood. We are interested in understanding whether the T:G-specific DNA glycosylase MBD4 can recognize and process nicked T:G mismatches. We have discovered that MBD4 binds a nicked T:G-containing DNA, but does not cleave thymine opposite guanine. To gain insight into this, we have determined a crystal structure of human MBD4 bound to a nicked T:G-containing DNA. This structure displayed the full insertion of thymine into the catalytic site and the recognition of thymine based on the catalytic site’s amino acid residues. However, thymine excision did not occur, presumably due to the inactivation of the catalytic D560 carboxylate nucleophile via a polar interaction with the 5′-hydrogen phosphate of the nicked DNA. The nicked complex was greatly stabilized by an ordered water molecule that formed four hydrogen bonds with the nicked DNA and MBD4. Interestingly, the arginine finger R468 did not engage in the phosphate pinching that is commonly observed in T:G mismatch recognition complex structures. Instead, the guanidinium moiety of R468 made bifurcated hydrogen bonding interactions with O6 of guanine, thereby stabilizing the estranged guanine. These observations suggest that R468 may sense and disrupt T:G pairs within the DNA duplex and stabilize the flipped-out thymine. The structure described here would be a close mimic of an intermediate in the base extrusion pathway induced by DNA glycosylase. Full article

This study elucidated the effect of Ti–Mo microalloying on the hydrogen embrittlement (HE) resistance and fracture behavior of warm-rolled Fe-5.6Mn-0.16C-1Al (wt%) steel. After intercritical annealing, both steels, i.e., without and with Ti–Mo microalloying, showed ultrafine ferrite (α) and austenite (γ_R) duplex microstructure. The addition of Ti–Mo to 5.6Mn steel reduces the volume fraction of γ_R, facilitating the formation of (Ti, Mo)C carbides in α phase and further refining the final microstructure. The product of ultimate tensile strength (UTS) and total elongation (TEL) of 5.6MnTiMo can be as high as 35 GPa·% with an ultra-high yield strength of above 1.2 GPa. Furthermore, the addition of Ti–Mo also had a significant effect on the resistance to HE of medium Mn steels. Firstly, the limited (Ti, Mo)C carbides precipitated in γ_R could act as irreversibly trap sites to capture a considerable amount of H, effectively increasing the C_H (Diffusible Hydrogen Content). Additionally, 5.6MnTiMo displayed higher γ_R stability, resulting in a reduced susceptibility to HE. The H-assisted microcracks mainly formed inside γ(α′) and extended along γ(α′) grain boundaries, leading to intergranular cracking and premature fracture. Full article

G-quadruplexes and i-motifs are four-stranded non-canonical structures of DNA. They exist in the cell, where they are implicated in the conformational regulation of cellular events, such as transcription, translation, DNA replication, telomere homeostasis, and genomic instability. Formation of the G-quadruplex and i-motif conformations in the genome is controlled by their competition with the pre-existing duplex. The fate of that competition depends upon the relative stabilities of the competing conformations, leading ultimately to a distribution of double helical, tetrahelical, and coiled conformations that coexist in dynamic equilibrium with each other. We previously developed a CD spectroscopy-based procedure to characterize the distribution of conformations adopted by equimolar mixtures of complementary G- and C-rich DNA strands from the promoter regions of the c-MYC, VEGF, and Bcl-2 oncogenes. In those bimolecular systems, duplex-to-tetraplex and duplex-to-coil transitions are accompanied by strand separation and an associated entropic cost. This situation is distinct from the pseudo-monomolecular nature of conformational transformations within the genome, where strand separation does not occur. To mimic better the situation in the genome, we here extend our studies to a monomolecular DNA construct—a hairpin—in which complementary G- and C-rich strands featuring sequences from the promoter region of the c-MYC oncogene are linked by a dT₁₁ loop. We used our CD-based procedure to quantify the distribution of conformational states sampled by the hairpin at pH 5.0 and 7.0 as a function of temperature and the concentration of KCl. The data were analyzed according to a thermodynamic model based on equilibria between the different conformational states to evaluate the thermodynamic properties of the duplex-to-coil, G-quadruplex-to-coil, and i-motif-to-coil transitions of the hairpin. The results have implications for the modulation of such transitions as a means of therapeutic intervention. Full article

Background/Objectives: The inhibitor of growth family member 3 (ING3) acts as an epigenetic reader through physical interactions with histone-modifying enzymes and subsequent chromatin remodelling processes. It is involved in various cellular functions, such as cell cycle control, cell growth, and apoptosis. Although ING3 was assigned tumour suppressor candidate status in some types of cancers, including prostate cancer, some studies suggest it acts to promote growth. To address these contradictory reports regarding its role in the initiation and progression of prostate cancer, we specifically addressed the question of whether ablation of ING3 in the mouse prostate is sufficient to initiate malignant transformation of the prostate and support its (candidate) tumour suppressor status. Methods: To generate the prostate-specific Ing3 knockout mouse, paternal inheritance of the PB-Cre4 transgene was used, while for the generation of a global knockout control, a female mouse harbouring the PB-Cre4 transgene was utilized. To determine the recombination efficiency of the Cre-LoxP system in the prostate at the Ing3 locus, a duplex probe-based digital PCR assay capable of counting undisrupted Ing3 copies was designed. The impact of DNA recombination on the protein level was investigated by immunohistochemical staining of prostate tissue samples. Results: In the prostate-specific knockout, digital PCR analysis revealed mosaic gene deletion. We found recombination efficiencies in the anterior, dorsolateral, and ventral prostate lobes ranging from approximately 15 to 30%. ING3 staining in the prostate was faint with no detectable differences in signal intensity between the knockout specimen and wild-type controls. This low ING3 expression in the prostate is consistent with observations of X-gal staining of an Ing3-LacZ reporter allele. Immunohistochemistry showed increased expression of DNA-damage-associated markers γH2AX and 53BP1. However, no gross anatomical abnormalities or prostate intraepithelial neoplasia (PIN) lesions in the prostate of tissue-specific knockout animals compared to wild-type controls were observed. Conclusions: Altogether, our data provide evidence that disruption of ING3 expression in prostate cells does not lead to malignant transformation and challenges the idea that ING3 acts primarily in a tumour-suppressive manner. Furthermore, this work supports the crucial role of ING3 in maintaining genomic stability, and we confirmed the embryonic lethal phenotype of homozygous Ing3 null mice that is rescued by ectopic expression of ING3. Full article