Search Results (58)

In mammals, the binding of transcriptional activators leads to the repositioning of silent loci from the nuclear periphery to the nuclear interior. However, it remained unknown whether the same mechanism functions in Drosophila. Here, using FISH and DamID, we have shown that binding the GAL4 activator to the silent loci causes weakening of their interactions with the nuclear lamina and relocalization inside nuclei in Drosophila salivary gland cells and neurons. This mimics the removal from the nuclear periphery of a neuron-specific gene upon its activation in neurons. Salivary gland cells contain polytene chromosomes with mechanical properties, different from chromosomes of diploid cells, while neurons represent predominantly non-dividing cell type. Our results indicate a causal relationship between transcriptional activator binding and changes in the intranuclear position of loci in Drosophila. They also point to the similarity in general chromatin dynamics in mammals and Drosophila, thus strengthening the role of model organisms in studying genome architecture. Full article

Kidney disease causes the retention of uremic metabolites in blood, which is associated with many comorbidities. Hemodialysis does not properly clear many metabolites, including large, middle-sized, and small protein-bound uremic toxins (PBUTs). Adsorption strategies for metabolite removal require the development of engineered adsorbents with tailored surfaces to increase the binding of desired metabolites. Albumin is uniquely positioned for modifying blood-contacting surfaces to absorb uremic metabolites, as it (i) minimizes non-specific protein adsorption and (ii) binds a range of molecules at Sudlow Sites I and II with different affinities. It is unknown if albumin-modified surfaces retain the adsorption qualities of solution-free albumin, namely, adsorption stability or specificity. Herein, albumin was covalently attached to iron oxide nanoparticles and characterized using multiple methods. Metabolite adsorption was conducted by incubating particles in a model solution of thirty-three uremic metabolites associated with kidney failure. Adsorption efficiency, selectivity, and stability were affected by albumin concentration and incubation time. Metabolite adsorption was found to change with time, and it was more effective on albumin-modified particles than unmodified controls. The findings outlined in this paper are crucial for the design of next-generation advanced blood-contacting materials to enhance dialysis and blood purification for patients with kidney disease. Full article

Oral immunotherapy (OIT) is a promising approach for treating food allergy. Here, we elucidated the mechanisms of desensitization induced by OIT in rats sensitized to ovalbumin (OVA). The desensitization was induced by ingestion of OVA three times per week after sensitization in rats. OIT suppressed the decrease in rectal temperature and increase in plasma histamine levels induced by OVA injection immediately and 4 weeks after OIT completion. Plasma OVA-specific IgE (sIgE) levels did not differ between the non-OIT and OIT groups, but OVA-specific IgG₁ levels were higher in the OIT group than in the non-OIT group at both timepoints. To evaluate IgG’s effect on IgE crosslinking with OVA, amplified luminescence proximity homogeneous assay involving crosslinking (AlphaCL) was performed. When IgG was removed using a Protein G column, the AlphaCL signal was significantly increased, especially in the OIT group, indicating that OIT-induced IgG inhibited the sIgE response. The proportions of cluster of differentiation (CD)4⁺ cells and CD4⁺CD25⁺FoxP3⁺ cells in mesenteric lymph nodes and spleen were similar between the two groups. These findings indicate that OIT attenuates systemic allergic responses by inhibiting sIgE binding to OVA through increased IgG. Our model is useful for understanding the mechanisms of OIT and optimizing therapeutic strategies for ameliorating food allergies. Full article

The effectiveness of copper-based composites, specifically cupric ion (Cu²⁺)-modified phyllosilicate minerals, was evaluated in reducing the concentration of infectious agents in the environment while minimizing metal ion release. The phyllosilicate minerals, vermiculite, exfoliated and unexfoliated, and sepiolite, all modified with Cu²⁺, were compared with copper oxide for their antiviral activity against non-enveloped porcine parvovirus (PPV) and enveloped human coronavirus 229E (HCoV). Sepiolite effectively removed PPV and HCoV from the solution, regardless of Cu²⁺ presence, while vermiculite showed substantial viral clearance only when Cu²⁺ was present. The kinetics of viral clearance was fast, with complete clearance within one hour in many cases. To better understand the mechanism of virus clearance, EDTA was added at different times during the clearance study for PPV. EDTA prevented virus clearance in all vermiculite samples, whereas sepiolite containing copper still demonstrated substantial virus clearance. The addition of BSA before the virus binding was able to block binding in all cases. It was determined that binding is the key mechanism, and PPV can be eluted from the minerals with EDTA and still be infectious. This study provides the potent antiviral mechanisms of Cu²⁺-modified phyllosilicate minerals, offering insights for designing paints and plastics for high-touch surfaces to reduce viral transmission and enhance public health significantly. Full article

This study investigates the potential of released polysaccharides (RPS) from the halophilic cyanobacterium Cyanothece sp. CE4 as biosorbents for heavy metals, specifically copper (Cu), nickel (Ni), and zinc (Zn). By combining ICP-OES, SEM-EDX, FT-IR spectroscopy, and XAS techniques, this work provides a comprehensive chemical and spectroscopic analysis of the biosorption mechanisms driving metal removal. The results revealed a strong binding affinity for Cu, followed by Ni and Zn, with RPS functional groups playing a key role in metal coordination. The RPS efficiently removed metals from both monometallic and multimetallic solutions, emphasizing their adaptability in competitive environments. XAS analysis highlighted unique metal-specific coordination patterns. Ni preferentially binds to oxygen donors and Zn to chlorine, and Cu exhibits non-selective binding. Remarkably, the extracted RPS achieved a maximum Cu removal capacity of 67 mg per gram of RPS dry weight, surpassing previously reported biosorption capacities. This study not only advances the understanding of biosorption mechanisms by cyanobacterial RPS but also emphasizes their dual role in environmental remediation and circular resource management. The insights provided here establish a foundation for the development of sustainable, cyanobacteria-based solutions for heavy-metal recovery and environmental sustainability. Full article

We have designed and produced 39 amino acid peptide mimics of the Torpedo and human acetylcholine receptors’ (AChRs) main immunogenic regions (MIRs). These conformationally sensitive regions consist of three non-contiguous segments of the AChR α-subunits and are the target of 50–70% of the anti-AChR autoantibodies (Abs) in human myasthenic serum and in the serum of rats with a model of that disease, experimental autoimmune myasthenia gravis (EAMG), induced by immunizing the rats with the Torpedo electric organ AChR. These MIR segments covalently joined together bind a significant fraction of the monoclonal antibodies (mAbs) raised in rats against electric organ AChR. Many of these mAbs cross react with the rat neuromuscular AChR MIR and induce myasthenic symptoms when injected into naïve rats. The human MIR mimic peptide (H39MIR) is evolutionarily related to that of the Torpedo electric organ MIR mimic peptide (T39MIR) with eight amino acid differences between the two MIR mimics. The mAbs raised to the electric organ AChR MIR cross react with the human and scores of other species’ neuromuscular AChRs. However, the mAbs do not cross react with the H39MIR mimic attached to the N-terminus of an intein–chitin-binding domain (H39MIR-IChBD) even though they do bind to the T39MIR-IChBD construct. To account for this difference in binding anti-MIR mAbs, each of the eight human amino acids was substituted individually into the T39MIR-IChBD, and four of them were found to weaken mAb recognition. Substituting the corresponding four Torpedo amino acids individually and in combination into the homologous positions in H39MIR-IChBD makes chimeric human MIR mimic peptides (T/H39MIR), some of which bind anti-MIR mAbs and anti-MIR Abs from rat EAMG and human MG sera. The best mAb binding chimeric peptide constructs may potentially serve as the basis of a diagnostic anti-MIR Ab titer assay that is both prognostic and predictive of disease severity. Furthermore, the best peptides may also serve as the targeting element of a non-steroidal antigen-specific treatment of MG to remove anti-AChR MIR Abs, either as fused to the N-terminals of the human immunoglobin F_c fragment or as the targeting component of a T cell chimeric autoantibody receptor (CAAR) directed to anti-MIR memory B cells for elimination. Full article

Molecularly imprinted polymers (MIPs) are synthetic receptors designed to selectively bind specific molecules, mimicking natural antibody–antigen interactions. Produced through polymerization around a target molecule (template), MIPs create imprints that confer high specificity and binding affinity upon template removal. Initially developed in the 1970s with organic polymers, MIPs now play critical roles in separation sciences, catalysis, drug delivery, and sensor technology. In forensic science, MIPs offer potential for sample preparation, pre-concentration, and analyte detection, especially with complex biological and non-biological matrices. They exhibit superior stability under extreme conditions, enabling their use in challenging forensic contexts such as detecting new psychoactive substances or trace explosives. Despite advantages like reusability and high selectivity, MIPs face limitations in forensic analysis due to their complex synthesis, potential template leakage, and non-specific binding. Moreover, the lack of standardized protocols limits their mainstream adoption, as forensic applications require validated, reproducible methods. This review systematically assesses MIPs in forensic toxicology, focusing on their current capabilities, limitations, and potential for broader integration into forensic workflows. Future research should address standardization and evaluate MIPs’ effectiveness in diverse forensic applications to realize their full potential. Full article

Gastric cancer (GC) is a malignant tumor of the gastrointestinal tract associated with high mortality rates and accounting for approximately 1 million new cases diagnosed annually. Surgery, particularly radical gastrectomy, remains the primary treatment; however, there are currently no specific approaches to better distinguish malignant from healthy tissue or to differentiate between metastatic and non-metastatic lymph nodes. As a result, surgeons have to remove all lymph nodes indiscriminately, increasing intraoperative risks for patients and prolonging hospital stay. Near-infrared fluorescence imaging with indocyanine green (ICG) can provide real-time visualization of the surgical field using both conventional laparoscopy and robotic mini-invasive precision surgery platforms. However, its application shows some limits, as ICG is a non-targeted contrast agent. Several studies are now investigating the potential efficacy of fluorescent targeted agents that could selectively bind to the tumor tissue, offering a valuable tool for metastatic mapping during robotic gastrectomy. This review aims to summarize the key fluorescent agents that have been developed to recognize GC markers, as well as those targeting the tumor microenvironment (TME) and metabolic features. These agents hold great potential as valuable tools for enhancing precision surgery in robotic gastrectomy procedures improving the clinical recovery of GC patients. Full article

Background: Oncological diseases are a major focus in medicine, with millions diagnosed each year, leading researchers to seek new diagnostic and treatment methods. One promising avenue is the development of targeted therapies and rapid diagnostic tests using recognition molecules. The pharmaceutical industry is increasingly exploring nucleic acid-based therapeutics. However, producing long oligonucleotides, especially aptamers, poses significant production challenges. Objectives: This study aims to demonstrate the efficacy of using molecular modeling, supported by experimental procedures, for altering aptamer nucleotide sequences while maintaining their binding capabilities. The focus is on reducing production costs and enhancing binding dynamics by removing nonfunctional regions and minimizing nonspecific binding. Methods: A molecular modeling approach was employed to elucidate the structure of a DNA aptamer, Gli-55, facilitating the truncation of nonessential regions in the Gli-55 aptamer, which selectively binds to glioblastoma (GBM). This process aimed to produce a truncated aptamer, Gli-35, capable of forming similar structural elements to the original sequence with reduced nonspecific binding. The efficiency of the truncation was proved by flow cytometry, fluorescence polarization (FP), and confocal microscopy. Results: The molecular design indicated that the new truncated Gli-35 aptamer retained the structural integrity of Gli-55. In vitro studies showed that Gli-35 had a binding affinity comparable to the initial long aptamer while the selectivity increased. Gli-35 internalized inside the cell faster than Gli-55 and crossed the blood–brain barrier (BBB), as demonstrated in an in vitro model. Conclusions: The success of this truncation approach suggests its potential applicability in scenarios where molecular target information is limited. The study highlights a strategic and resource-efficient methodology for aptamer development. By employing molecular modeling and truncation, researchers can reduce production costs and avoid trial and error in sequence selection. This approach is promising for enhancing the efficiency of therapeutic agent development, particularly in cases lacking detailed molecular target insights. Full article

The efficacy of glioblastoma treatment is closely associated with complete tumor resection. However, conventional surgical techniques often result in incomplete removal, leading to poor prognosis. A major challenge is the accurate delineation of tumor margins from healthy tissues. Imaging-guided surgery, particularly using fluorescent probes, is a promising solution for intraoperative guidance. The recently developed ‘always-on’ types of targeted fluorescence probes generate signals irrespective of their presence in tumor cells or in blood circulation, hampering their effectiveness. Here, we propose a novel activatable fluorescence imaging probe, Q-cRGD, that targets glioma cells via the specific binding of the cyclic Arg-Gly Asp-containing pentapeptide (cRGD) to integrins. The Q-cRGD probe was synthesized by conjugating a near-infrared (NIR) dye to a tryptophan quencher via a disulfide linkage, including a cRGD-targeting ligand. This activatable probe remained inactive until the redox-responsive cleavage of the disulfide linkage occurred within the target cell. The zwitterionic nature of NIR dyes minimizes nonspecific interactions with serum proteins, thereby enhancing the tumor-to-background signal ratio (TBR). An in vivo fluorescence imaging study demonstrated a TBR value of 2.65 within 3 h of the intravenous injection of Q-cRGD, confirming its potential utility in imaging-guided brain cancer surgery. Full article

A low total plasma vitamin B12 supports a clinical suspicion of B12 deficiency, while the interpretation of an unexpectedly normal/high level is marred by controversies. Here, we critically review current knowledge on B12 in blood plasma, including the presence of the so-called “macro-B12”. The latter form is most often defined as the fraction of B12 that can be removed by precipitation with polyethylene glycol (PEG), a nonspecific procedure that also removes protein polymers and antibody-bound analytes. Plasma B12 includes B12 attached to transcobalamin and haptocorrin, and an increased concentration of one or both proteins almost always causes an elevation of B12. The total plasma B12 is measured by automated competitive binding assays, often incorrectly referred to as immunoassays, since the binding protein is intrinsic factor and not an antibody. An unexpectedly high level of B12 may be further explored using immunological measurements of haptocorrin and transcobalamin (optionally combined with e.g., size-exclusion chromatography). Nonspecific methods, such as PEG precipitation, are likely to give misleading results and cannot be recommended. Currently, the need for evaluation of a high B12 of unknown etiology is limited since other tests (such as measurements of methylmalonic acid) may better guide the diagnosis of B12 deficiency. Full article

Hemochromatosis represents clinically one of the most important genetic storage diseases of the liver caused by iron overload, which is to be differentiated from hepatic iron overload due to excessive iron release from erythrocytes in patients with genetic hemolytic disorders. This disorder is under recent mechanistic discussion regarding ferroptosis, reactive oxygen species (ROS), the gut microbiome, and alcohol abuse as a risk factor, which are all topics of this review article. Triggered by released intracellular free iron from ferritin via the autophagic process of ferritinophagy, ferroptosis is involved in hemochromatosis as a specific form of iron-dependent regulated cell death. This develops in the course of mitochondrial injury associated with additional iron accumulation, followed by excessive production of ROS and lipid peroxidation. A low fecal iron content during therapeutic iron depletion reduces colonic inflammation and oxidative stress. In clinical terms, iron is an essential trace element required for human health. Humans cannot synthesize iron and must take it up from iron-containing foods and beverages. Under physiological conditions, healthy individuals allow for iron homeostasis by restricting the extent of intestinal iron depending on realistic demand, avoiding uptake of iron in excess. For this condition, the human body has no chance to adequately compensate through removal. In patients with hemochromatosis, the molecular finetuning of intestinal iron uptake is set off due to mutations in the high-FE²⁺ (HFE) genes that lead to a lack of hepcidin or resistance on the part of ferroportin to hepcidin binding. This is the major mechanism for the increased iron stores in the body. Hepcidin is a liver-derived peptide, which impairs the release of iron from enterocytes and macrophages by interacting with ferroportin. As a result, iron accumulates in various organs including the liver, which is severely injured and causes the clinically important hemochromatosis. This diagnosis is difficult to establish due to uncharacteristic features. Among these are asthenia, joint pain, arthritis, chondrocalcinosis, diabetes mellitus, hypopituitarism, hypogonadotropic hypogonadism, and cardiopathy. Diagnosis is initially suspected by increased serum levels of ferritin, a non-specific parameter also elevated in inflammatory diseases that must be excluded to be on the safer diagnostic side. Diagnosis is facilitated if ferritin is combined with elevated fasting transferrin saturation, genetic testing, and family screening. Various diagnostic attempts were published as algorithms. However, none of these were based on evidence or quantitative results derived from scored key features as opposed to other known complex diseases. Among these are autoimmune hepatitis (AIH) or drug-induced liver injury (DILI). For both diseases, the scored diagnostic algorithms are used in line with artificial intelligence (AI) principles to ascertain the diagnosis. The first-line therapy of hemochromatosis involves regular and life-long phlebotomy to remove iron from the blood, which improves the prognosis and may prevent the development of end-stage liver disease such as cirrhosis and hepatocellular carcinoma. Liver transplantation is rarely performed, confined to acute liver failure. In conclusion, ferroptosis, ROS, the gut microbiome, and concomitant alcohol abuse play a major contributing role in the development and clinical course of genetic hemochromatosis, which requires early diagnosis and therapy initiation through phlebotomy as a first-line treatment. Full article

Non-heme dioxygenases of the AlkB family hold a unique position among enzymes that repair alkyl lesions in nucleic acids. These enzymes activate the Fe(II) ion and molecular oxygen through the coupled decarboxylation of the 2-oxoglutarate co-substrate to subsequently oxidize the substrate. ALKBH3 is a human homolog of E. coli AlkB, which displays a specific activity toward N1-methyladenine and N3-methylcytosine bases in single-stranded DNA. Due to the lack of a DNA-bound structure of ALKBH3, the basis of its substrate specificity and structure–function relationships requires further exploration. Here we have combined biochemical and biophysical approaches with site-directed mutational analysis to elucidate the role of key amino acids in maintaining the secondary structure and catalytic activity of ALKBH3. Using stopped-flow fluorescence spectroscopy we have shown that conformational dynamics play a crucial role in the catalytic repair process catalyzed by ALKBH3. A transient kinetic mechanism, which comprises the steps of the specific substrate binding, eversion, and anchoring within the DNA-binding cleft, has been described quantitatively by rate and equilibrium constants. Through CD spectroscopy, we demonstrated that replacing side chains of Tyr143, Leu177, and His191 with alanine results in significant alterations in the secondary structure content of ALKBH3 and decreases the stability of mutant proteins. The bulky side chain of Tyr143 is critical for binding the methylated base and stabilizing its flipped-out conformation, while its hydroxyl group is likely involved in facilitating the product release. The removal of the Leu177 and His191 side chains substantially affects the secondary structure content and conformational flexibility, leading to the complete inactivation of the protein. The mutants lacking enzymatic activity exhibit a marked decrease in antiparallel β-strands, offset by an increase in the helical component. Full article

Identifying potential sources of suspended sediment (SS) in headwater catchments is crucial for water quality management. To differentiate these potential SS sources, we investigated the distribution of two fallout radionuclides (FRNs), ¹³⁷Cs and ²¹⁰Pb_ex, using gamma spectrometry along with soil organic matter (SOM) analysis in a headwater catchment with five potential SS sources: cultivated land, non-harvested forest floor, eroded hillslope, harvested forest floor, and stream bank. The ¹³⁷Cs and ²¹⁰Pb_ex concentrations and the SOM content were considerably higher in the harvested forest floor materials than in the other four potential SS source materials. FRN concentrations revealed distinct properties according to the type of potential SS sources. Specifically, the combination of FRNs (with the effect of SOM content removed) associated with the mineral fraction and SOM showed distinguishable differences among the potential SS sources, except for no difference between cultivated land and eroded hillslope. Therefore, SOM and FRNs, or their combination, can be effective indices to differentiate or trace potential SS sources on various land use/land cover types within a catchment. Further field tests will allow the tracing techniques that bind FRNs with SOM to contribute to understanding SS transport from non-point sources within a catchment. Full article

Epigenetic mechanisms, including DNA methylation, histone modifications, and non-coding RNA, play a crucial role in the regulation of gene expression and are pivotal in biological processes like apoptosis, cell proliferation, and differentiation. SIN3a serves as a scaffold protein and facilitates interactions with transcriptional epigenetic partners and specific DNA-binding transcription factors to modulate gene expression by adding or removing epigenetic marks. However, the activation or repression of gene expression depends on the factors that interact with SIN3a, as it can recruit both transcriptional activators and repressors. The role of SIN3a has been extensively investigated in the context of cancer, including melanoma, lung, and breast cancer. Our group is interested in defining the roles of SIN3a and its partners in pulmonary vascular disease. Pulmonary arterial hypertension (PAH) is a multifactorial disease often described as a cancer-like disease and characterized by disrupted cellular metabolism, sustained vascular cell proliferation, and resistance to apoptosis. Molecularly, PAH shares many common signaling pathways with cancer cells, offering the opportunity to further consider therapeutic strategies used for cancer. As a result, many signaling pathways observed in cancer were studied in PAH and have encouraged new research studying SIN3a’s role in PAH due to its impact on cancer growth. This comparison offers new therapeutic options. In this review, we delineate the SIN3a-associated epigenetic mechanisms in cancer and PAH cells and highlight their impact on cell survival and proliferation. Furthermore, we explore in detail the role of SIN3a in cancer to provide new insights into its emerging role in PAH pathogenesis. Full article