Cell Biology Research Articles

Modular Fluorescent Cholesterol Naphthalimide Probes And Their Application For Cholesterol Trafficking Studies In Cells.

Development of fluorescent cholesterol analogs to better understand subcellular cholesterol trafficking is of great interest for cell biology and medicine. Our approach utilizes a bifunctional 1,8-naphthalimide scaffold with a push-pull character, modified on one side with a head group and a linker on the other side connecting it to cholesterol via an ester bond. Through structure-function studies, we've explored how different substituents-linkers and head groups-affect the ability of these fluorescent cholesterol naphthalimide analogs (CNDs) to mimic natural cholesterol behavior at both molecular and cellular levels. We categorized the resulting analogs into three groups: neutral, charged, and those featuring a hydroxyl group. Each compound was assessed for its solvatochromic behavior in organic solvents and model membranes. Extensive all-atom molecular dynamics simulations helped us examine how these analogs perform in model membranes compared to cholesterol. Additionally, we investigated the partitioning of these fluorescent probes in phase-separated giant unilamellar vesicles. We evaluated the uptake and distribution of these probes within mouse fibroblast cells and astrocytes, for their subcellular distributions in lysosomes and compared that to BODIPY-cholesterol, a well-regarded fluorescent cholesterol analog. The internalization efficiency of the fluorescent probes varies in different cell types and is affected mainly by the head groups. Our results demonstrate that the modular design significantly simplifies the creation of fluorescent cholesterol probes bearing distinct spectral, biophysical, and cellular targeting features, which makes it a valuable toolkit for the investigation of subcellular distribution and trafficking of cholesterol and its derivatives.

The Low Tumorigenic Risk and Subtypes of Cardiomyocytes Derived from Human-induced Pluripotent Stem Cells

Background: Clinical application of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) is a promising approach for the treatment of heart diseases. However, the tumorigenicity of hiPSC-CMs remains a concern for their clinical applications and the composition of the hiPSC-CM subtypes need to be clearly identified. Methods: In the present study, hiPSC-CMs were induced from hiPSCs via modulation of Wnt signaling followed by glucose deprivation purification. The structure, function, subpopulation composition, and tumorigenic risk of hiPSC-CMs were evaluated by single-cell RNA sequencing (scRNAseq), whole exome sequencing (WES), and integrated molecular biology, cell biology, electrophysiology, and/or animal experiments. Results: The high purity of hiPSC-CMs, determined by flow cytometry analysis, was generated. ScRNAseq analysis of differentiation day (D) 25 hiPSC-CMs did not identify the transcripts representative of undifferentiated hiPSCs. WES analysis showed a few newly acquired confidently identified mutations and no mutations in tumor susceptibility genes. Further, no tumor formation was observed after transplanting hiPSC-CMs into NOD-SCID mice for 3 months. Moreover, D25 hiPSC-CMs were composed of subtypes of ventricular-like cells (23.19%) and atrial-like cells (66.45%) in different cell cycle stages or mature levels, based on the scRNAseq analysis. Furthermore, a subpopulation of more mature ventricular cells (3.21%) was identified, which displayed significantly up-regulated signaling pathways related to myocardial contraction and action potentials. Additionally, a subpopulation of cardiomyocytes in an early differentiation stage (3.44%) experiencing nutrient stress-induced injury and heading toward apoptosis was observed. Conclusions: This study confirmed the biological safety of hiPSC-CMs and described the composition and expression profile of cardiac subtypes in hiPSC-CMs which provide standards for quality control and theoretical supports for the translational applications of hiPSC-CMs.

Spatiotemporal Variations in Co-Occurrence Patterns of Planktonic Prokaryotic Microorganisms along the Yangtze River.

Bacteria and archaea are foundational life forms on Earth and play crucial roles in the development of our planet's biological hierarchy. Their interactions influence various aspects of life, including eukaryotic cell biology, molecular biology, and ecological dynamics. However, the coexistence network patterns of these microorganisms within natural river ecosystems, vital for nutrient cycling and environmental health, are not well understood. To address this knowledge gap, we systematically explored the non-random coexistence patterns of planktonic bacteria and archaea in the 6000-km stretch of the Yangtze River by using high-throughput sequencing technology. By analyzing the O/R ratio, representing the divergence between observed (O%) and random (R%) co-existence incidences, and the module composition, we found a preference of both bacteria and archaea for intradomain associations over interdomain associations. Seasons notably influenced the co-existence of bacteria and archaea, and archaea played a more crucial role in spring as evidenced by their predominant presence of interphyla co-existence and more species as keystone ones. The autumn network was characterized by a higher node or edge number, greater graph density, node degree, degree centralization, and nearest neighbor degree, indicating a more complex and interconnected structure. Landforms markedly affected microbial associations, with more complex networks and more core species found in plain and non-source areas. Distance-decay analysis suggested the importance of geographical distance in shaping bacteria and archaea co-existence patterns (more pronounced in spring). Natural, nutrient, and metal factors, including water temperature, NH4+-N, Fe, Al, and Ni were identified as crucial determinants shaping the co-occurrence patterns. Overall, these findings revealed the dynamics of prokaryotic taxa coexistence patterns in response to varying environmental conditions and further contributed to a broader understanding of microbial ecology in freshwater biogeochemical cycling.

Role and Mechanism of Lactylation in Cancer

Post translational modifications (PTMs) can change the properties of a protein by covalent addition of functional groups to one or more amino acids, and influence almost all aspects of normal cell biology and pathogenesis. Lactylation is a novel identified PTM, and has been found in both histone and non-histone proteins. Since associated with the end product of glycolysis-- lactate, lactylation modification could provide a new perspective for understanding the relationship between metabolic reprogramming and epigenetic modifications. Accumulated evidences suggest that lactylation play important roles in tumor progression and links to poor prognosis in clinical studies. Histone lactylation can affect gene expression in tumor cells and immunological cells, further promoting tumor progression and immune suppression. Lactylation on non-histone proteins can also regulate tumor progression and drug resistance. In this review, we aimed to summarize the roles of lactylation in cancer progression, microenvironment interactions and immune suppression, try to identify new molecular targets for cancer therapy and provide a new direction for combined targeted therapy and immunotherapy. .

Collagen structures of demineralized bone paper direct mineral metabolism.

Bone is a dynamic mineralized tissue that undergoes continuous turnover throughout life. While the general mechanism of bone mineral metabolism is documented, the role of underlying collagen structures in regulating osteoblastic mineral deposition and osteoclastic mineral resorption remains an active research area, partly due to the lack of biomaterial platforms supporting accurate and analytical investigation. The recently introduced osteoid-inspired demineralized bone paper (DBP), prepared by 20-μm thin sectioning of demineralized bovine compact bone, holds promise in addressing this challenge as it preserves the intrinsic bony collagen structure and retains semi-transparency. Here, we report on the impact of collagen structures on modulating osteoblast and osteoclast-driven bone mineral metabolism using vertical and transversal DBPs that exhibit a uniaxially aligned and a concentric ring collagen structure, respectively. Translucent DBP reveals these collagen structures and facilitates longitudinal tracking of mineral deposition and resorption under brightfield microscopy for at least 3wk. Genetically labeled primary osteogenic cells allow fluorescent monitoring of these cellular processes. Osteoblasts adhere and proliferate following the underlying collagen structures of DBPs. Osteoblastic mineral deposition is significantly higher in vertical DBP than in transversal DBP. Spatiotemporal analysis reveals notably more osteoblast adhesion and faster mineral deposition in vascular regions than in bone regions. Subsequent osteoclastic resorption follows these mineralized collagen structures, directing distinct trench and pit-type resorption patterns. In vertical DBP, trench-type resorption occurs at an 80% frequency, whereas transversal DBP shows 35% trench-type and 65% pit-type resorption. Our studies substantiate the importance of collagen structures in regulating mineral metabolism by osteogenic cells. DBP is expected to serve as an enabling biomaterial platform for studying various aspects of cellular and extracellular bone remodeling biology.

Tissue gene expression profiles and communication networks inform candidate blood biomarker identification in psoriasis and atopic dermatitis

Overlapping clinical and pathomechanistic features can complicate the diagnosis and treatment of inflammatory skin diseases, including psoriasis and atopic dermatitis (AD). Spatial transcriptomics allows the identification of disease- and cell-specific molecular signatures that may advance biomarker development and future treatments.This study identified transcriptional signatures in keratinocytes and sub-basal CD4+ and CD8+ T lymphocytes from patients with psoriasis and AD. In silico prediction of ligand:receptor interactions delivered key signalling pathways (interferon, effector T cells, stroma cell and matrix biology, neuronal development, etc.). Targeted validation of selected transcripts, including CCL22, RELB, and JUND, in peripheral blood T cells suggests the chosen approach as a promising tool also in other inflammatory diseases.Psoriasis and AD are characterized by transcriptional dysregulation in T cells and keratinocytes that may be targeted therapeutically. Spatial transcriptomics is a valuable tool in the search for molecular signatures that can be used as biomarkers and/or therapeutic targets.

How Parkinson’s Disease-Linked LRRK2 Mutations Affect Different CNS Cell Types

LRRK2 is a relatively common genetic risk factor for Parkinson’s disease (PD), with six coding variants known to cause familial PD. Non-coding variation at the same locus is also associated with sporadic PD. LRRK2 plays a role in many different intracellular signaling cascades including those involved in endolysosomal function, cytoskeletal dynamics, and Ca2+ homeostasis. PD-causing LRRK2 mutations cause hyperactive LRRK2 kinase activity, resulting in altered cellular signaling. Importantly, LRRK2 is lowly expressed in neurons and prominently expressed in non-neuronal cells in the brain. In this review, we will summarize recent and novel findings on the effects of PD-causing LRRK2 mutations in different nervous system cell types. This review will also provide novel insight into future areas of research at the intersection of LRRK2 cell biology, cell type specificity, and PD.

Photoactivatable Blue Fluorescent Protein.

Photoactivatable and photoswitchable fluorescent proteins (FPs) are sophisticated molecular tools that in combination with super-resolution microscopy are helping to elucidate many biological processes. Through the Y66H mutation in the chromophore of the violet fluorescent protein SumireF, we created the first photoactivatable blue fluorescent protein (PA-BFP). This protein is rapidly activated over ordinary UV transilluminators at 302 or 365 nm in irreversible mode and by direct exposition to sunlight. The maximum excitation and emission wavelengths of this protein, centered at 358 and 445 nm, respectively, resemble the values of DAPI-the blue stain widely used in fluorescence microscopy to visualize nucleic acids in cells. Therefore, the immediate use of PA-BFP in cellular biology is clear because the technology required to follow this new genetically encoded reporter at the microscopic level has already been established. PA-BFP can potentially be used together with other photoactivatable fluorescent proteins of different colors to label multiple proteins, which can be simultaneously tracked by advanced microscopic techniques.

Chemo-Enzymatic Generation of Highly Fluorescent Nucleoside Analogs Using Purine-Nucleoside Phosphorylase

Chemo-enzymatic syntheses of strongly fluorescent nucleoside analogs, potentially applicable in analytical biochemistry and cell biology are reviewed. The syntheses and properties of fluorescent ribofuranosides of several purine, 8-azapurine, and etheno-purine derivatives, obtained using various types of purine nucleoside phosphorylase (PNP) as catalysts, as well as α-ribose-1-phosphate (r1P) as a second substrate, are described. In several instances, the ribosylation sites are different to the canonical purine N9. Some of the obtained ribosides show fluorescence yields close to 100%. Possible applications of the new analogs include assays of PNP, nucleoside hydrolases, and other enzyme activities both in vitro and within living cells using fluorescence microscopy.

Relating Macroscopic PET Radiomics Features to Microscopic Tumor Phenotypes Using a Stochastic Mathematical Model of Cellular Metabolism and Proliferation.

Cancers can manifest large variations in tumor phenotypes due to genetic and microenvironmental factors, which has motivated the development of quantitative radiomics-based image analysis with the aim to robustly classify tumor phenotypes in vivo. Positron emission tomography (PET) imaging can be particularly helpful in elucidating the metabolic profiles of tumors. However, the relatively low resolution, high noise, and limited PET data availability make it difficult to study the relationship between the microenvironment properties and metabolic tumor phenotype as seen on the images. Most of previously proposed digital PET phantoms of tumors are static, have an over-simplified morphology, and lack the link to cellular biology that ultimately governs the tumor evolution. In this work, we propose a novel method to investigate the relationship between microscopic tumor parameters and PET image characteristics based on the computational simulation of tumor growth. We use a hybrid, multiscale, stochastic mathematical model of cellular metabolism and proliferation to generate simulated cross-sections of tumors in vascularized normal tissue on a microscopic level. The generated longitudinal tumor growth sequences are converted to PET images with realistic resolution and noise. By changing the biological parameters of the model, such as the blood vessel density and conditions for necrosis, distinct tumor phenotypes can be obtained. The simulated cellular maps were compared to real histology slides of SiHa and WiDr xenografts imaged with Hoechst 33342 and pimonidazole. As an example application of the proposed method, we simulated six tumor phenotypes that contain various amounts of hypoxic and necrotic regions induced by a lack of oxygen and glucose, including phenotypes that are distinct on the microscopic level but visually similar in PET images. We computed 22 standardized Haralick texture features for each phenotype, and identified the features that could best discriminate the phenotypes with varying image noise levels. We demonstrated that "cluster shade" and "difference entropy" are the most effective and noise-resilient features for microscopic phenotype discrimination. Longitudinal analysis of the simulated tumor growth showed that radiomics analysis can be beneficial even in small lesions with a diameter of 3.5-4 resolution units, corresponding to 8.7-10.0 mm in modern PET scanners. Certain radiomics features were shown to change non-monotonically with tumor growth, which has implications for feature selection for tracking disease progression and therapy response.

A comprehensive analysis of m6A/m7G/m5C/m1A-related gene expression and immune infiltration in liver ischemia–reperfusion injury by integrating bioinformatics and machine learning algorithms

BackgroundLiver ischemia–reperfusion injury (LIRI) is closely associated with immune infiltration, which commonly occurs after liver surgery, especially liver transplantation. Therefore, it is crucial to identify the genes responsible for LIRI and develop effective therapeutic strategies that target immune response. Methylation modifications in mRNA play various crucial roles in different diseases. This study aimed to identify potential methylation-related markers in patients with LIRI and evaluate the corresponding immune infiltration.MethodsTwo Gene Expression Omnibus datasets containing human liver transplantation data (GSE12720 and GSE151648) were downloaded for integrated analysis. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses were conducted to investigate the functional enrichment of differentially expressed genes (DEGs). Differentially expressed methylation-related genes (DEMRGs) were identified by overlapping DEG sets and 65 genes related to N6-methyladenosine (m6A), 7-methylguanine (m7G), 5-methylcytosine (m5C), and N1-methyladenosine (m1A). To evaluate the relationship between DEMRGs, a protein–protein interaction (PPI) network was utilized. The core DEMRGs were screened using three machine learning algorithms: least absolute shrinkage and selection operator, random forest, and support vector machine-recursive feature elimination. After verifying the diagnostic efficacy using the receiver operating characteristic curve, we validated the expression of the core DEMRGs in clinical samples and performed relative cell biology experiments. Additionally, the immune status of LIRI was comprehensively assessed using the single sample gene set enrichment analysis algorithm. The upstream microRNA and transcription factors of the core DEMRGs were also predicted.ResultsIn total, 2165 upregulated and 3191 downregulated DEGs were identified, mainly enriched in LIRI-related pathways. The intersection of DEGs and methylation-related genes yielded 28 DEMRGs, showing high interaction in the PPI network. Additionally, the core DEMRGs YTHDC1, METTL3, WTAP, and NUDT3 demonstrated satisfactory diagnostic efficacy and significant differential expression and corresponding function based on cell biology experiments. Furthermore, immune infiltration analyses indicated that several immune cells correlated with all core DEMRGs in the LIRI process to varying extents.ConclusionsWe identified core DEMRGs (YTHDC1, METTL3, WTAP, and NUDT3) associated with immune infiltration in LIRI through bioinformatics and validated them experimentally. This study may provide potential methylation-related gene targets for LIRI immunotherapy.

Appreciating animal induced pluripotent stem cells to shape plant cell reprogramming strategies.

Animals and plants have developed resilience mechanisms to effectively endure and overcome physical damage and environmental challenges throughout their life span. To sustain their vitality, both animals and plants employ mechanisms to replenish damaged cells, either directly, involving the activity of adult stem cells, or indirectly, via dedifferentiation of somatic cells that are induced to revert to a stem cell state and subsequently redifferentiate. Stem cell research has been a rapidly advancing field in animal studies for many years, driven by its promising potential in human therapeutics, including tissue regeneration and drug development. A major breakthrough was the discovery of induced pluripotent stem cells (iPSCs), which are reprogrammed from somatic cells by expressing a limited set of transcription factors. This discovery enabled the generation of an unlimited supply of cells that can be differentiated into specific cell types and tissues. Equally, a keen interest in the connection between plant stem cells and regeneration has been developed in the last decade, driven by the demand to enhance plant traits such as yield, resistance to pathogens, and the opportunities provided by CRISPR/Cas-mediated gene editing. Here we discuss how knowledge of stem cell biology benefits regeneration technology, and we speculate on the creation of a universal genotype-independent iPSC system for plants to overcome regenerative recalcitrance.

Integrative transcriptomic and proteomic profiling of the effects of cell confluency on gene expression

In this study we examine the impact of cell confluency on gene expression. We focused on Argonaute (AGO) protein dynamics and associated gene and protein expression in HEK293, A375, and SHSY5Y cell lines. As a consequence of cell confluency, AGO2 protein translocates into the nucleus. Therefore, we generated transcriptomic data using RNA sequencing to compare gene expression in subconfluent versus confluent cells, which highlighted significant alterations in gene regulation patterns directly corresponding to changes in cell density. Our study also encompasses miRNA profiling data obtained through small RNA sequencing, revealing miRNA expressional changes dependent on cellular confluency, as well as cellular localization. Finally, we derived proteomic data from mass spectrometry analyses following AGO1-4 immunoprecipitation, providing a comprehensive view of AGO interactome in both nuclear and cytoplasmic compartments under varying confluency. These datasets offer a detailed exploration of the cellular and molecular dynamics, influenced by cell confluency, presenting a valuable resource for further research in cellular biology, particularly in understanding the basic mechanisms of cell density in cancer cells.

Activation of alveolar epithelial ER stress by β-coronavirus infection disrupts surfactant homeostasis in mice: implications for COVID-19 respiratory failure.

COVID-19 syndrome is characterized by acute lung injury, hypoxemic respiratory failure, and high mortality. Alveolar type 2 (AT2) cells are essential for gas exchange, repair, and regeneration of distal lung epithelium. We have shown that the causative agent, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and other members of the β-coronavirus genus induce an endoplasmic reticulum (ER) stress response in vitro; however, the consequences for host AT2 cell function in vivo are less understood. To study this, two murine models of coronavirus infection were used-mouse hepatitis virus-1 (MHV-1) in A/J mice and a mouse-adapted SARS-CoV-2 strain. MHV-1-infected mice exhibited dose-dependent weight loss with histological evidence of distal lung injury accompanied by elevated bronchoalveolar lavage fluid (BALF) cell counts and total protein. AT2 cells showed evidence of both viral infection and increased BIP/GRP78 expression, consistent with activation of the unfolded protein response (UPR). The AT2 UPR included increased inositol-requiring enzyme 1α (IRE1α) signaling and a biphasic response in PKR-like ER kinase (PERK) signaling accompanied by marked reductions in AT2 and BALF surfactant protein (SP-B and SP-C) content, increases in surfactant surface tension, and emergence of a reprogrammed epithelial cell population (Krt8+ and Cldn4+). The loss of a homeostatic AT2 cell state was attenuated by treatment with the IRE1α inhibitor OPK-711. As a proof-of-concept, C57BL6 mice infected with mouse-adapted SARS-CoV-2 demonstrated similar lung injury and evidence of disrupted surfactant homeostasis. We conclude that lung injury from β-coronavirus infection results from an aberrant host response, activating multiple AT2 UPR stress pathways, altering surfactant metabolism/function, and changing AT2 cell state, offering a mechanistic link between SARS-CoV-2 infection, AT2 cell biology, and acute respiratory failure.NEW & NOTEWORTHY COVID-19 syndrome is characterized by hypoxemic respiratory failure and high mortality. In this report, we use two murine models to show that β-coronavirus infection produces acute lung injury, which results from an aberrant host response, activating multiple epithelial endoplasmic reticular stress pathways, disrupting pulmonary surfactant metabolism and function, and forcing emergence of an aberrant epithelial transition state. Our results offer a mechanistic link between SARS-CoV-2 infection, AT2 cell biology, and respiratory failure.

In Situ Magnetometry of Iron in Human Dopaminergic Neurons Using Superresolution MRI and Ion-Beam Microscopy

Paramagnetic transition metals play a crucial role as cofactors in various cellular catalytic processes. However, their high concentrations in reactive oxidation states can induce oxidative stress, resulting in cell dysfunction or death. Hence, it is vital to have methods to monitor metal concentrations and paramagnetic properties in cells for medicine and cell biology. Here we present a novel multimodal method for in-cell magnetometry enabling direct measurement of metal magnetic properties within individual cells in tissue, without prior isolation and at room temperature. Individual cell magnetic moments are measured using superresolution magnetic resonance imaging (MRI) microscopy at 9.4 T by detecting microscopic magnetic-field perturbations around the cells. The cellular metal content is quantified using ion-beam microscopy or synchrotron micro-x-ray fluorescence for the same cells. The metal magnetic susceptibility at 9.4 T is then obtained from the slope of the cell magnetic moments’ dependence on cell metal content. To estimate the susceptibility at lower fields, multifield MR relaxometry and biophysical modeling are employed, extrapolating the 9.4-T susceptibility values to fields as low as 3 T. We apply the new method to determine the susceptibility of iron accumulated in human dopaminergic neurons inside neuromelanin, the by-product of dopamine synthesis. The susceptibility of iron in neuromelanin is measured to be χρ=(2.98±0.19)×10−6 m3/kg providing unique insights into the biochemistry of iron inside dopaminergic neurons. The obtained value reveals a predominant monoatomic low-affinity iron-binding site within neuromelanin, indicating a higher neurotoxicity of iron than previously suggested. Furthermore, the measured susceptibility value establishes a quantitative relationship between cellular iron concentration and iron-sensitive MRI parameters, which can be noninvasively measured . This breakthrough paves the way for the detection of dopaminergic neuron density and iron load, requiring a standard clinical MRI scanner only. It promises to facilitate early diagnosis of Parkinson’s disease. In conclusion, our presented novel method enables the direct measurements of magnetic properties of paramagnetic metals within single cells with high sensitivity and across large cell groups within a macroscopic volume, providing invaluable information about the cellular biology of metals. Published by the American Physical Society 2024

A first-in-human phase I trial of daily oral zelenirstat, a N-myristoyltransferase inhibitor, in patients with advanced solid tumors and relapsed/refractory B-cell lymphomas.

Myristoylation, the N-terminal addition of the fatty acid myristate to proteins, regulates membrane-bound signal transduction pathways important in cancer cell biology. This modification is catalyzed by two N-myristoyltransferases, NMT1 and NMT2. Zelenirstat is a first-in-class potent oral small molecule inhibitor of both NMT1 and NMT2 proteins. Patients with advanced solid tumors and relapsed/refractory (R/R) B-cell lymphomas were enrolled in an open label, phase I dose escalation trial of oral daily zelenirstat, administered in 28-day cycles until progression or unacceptable toxicity. The endpoints were to evaluate dose-limiting toxicities (DLT) to establish a maximum tolerated dose (MTD), pharmacokinetic parameters, and anticancer activity. Twenty-nine patients were enrolled (25 advanced solid tumor; 4 R/R B-cell lymphoma) and 24 were DLT-evaluable. Dosing ranged from 20mg once daily (OD) to 210mg OD without DLT, but gastrointestinal DLTS were seen in the 280mg cohort. MTD and recommended phase 2 dose were 210mg OD. Common adverse events were predominantly Gr ≤ 2 nausea, vomiting, diarrhea, and fatigue. Plasma concentrations peaked at 2h with terminal half-lives averaging 10h. Steady state was achieved by day 15, and higher doses achieved trough concentrations predicted to be therapeutic. Stable disease as best response was seen in eight (28%) patients. Progression-free survival and overall survival were significantly better in patients receiving 210mg OD compared to those receiving lower doses. Zelenirstat is well-tolerated, achieves plasma exposures expected for efficacy, and shows early signs of anticancer activity. Further clinical development of zelenirstat is warranted.

Proceedings of the 69th Congress of the Italian Embryological Group-Italian Society of Development and Cell Biology (GEI-SIBSC) - Naples, 11-14 June 2024

Proceedings of the 69th Congress of the Italian Embryological Group-Italian Society of Development and Cell Biology (GEI-SIBSC) - Naples, 11-14 June 2024

Dual-Stimuli Regulation of DNAzyme Cleavage Reaction by Coordination-Driven Nanoprobes for Cancer Cell Imaging.

Endowing current artificial chemical reactions (ACRs) with high specificity and intricate activation capabilities is crucial for expanding their applications in accurate bioimaging within living cells. However, most of the reported ACR-based evaluations relied on either single biomarker stimuli or dual activators without obvious biological relevance, still limiting their accuracy and fidelity. Herein, taking the metal-ion-dependent DNAzyme cleavage reaction as a model ACR, two regulators, glutathione (GSH) and telomerase (TE) activated DNAzyme cleavage reactions, were exploited for precise discrimination of cancerous cells from normal cells. DNA probe was self-assembled into the ZIF-90 nanoparticle framework to construct coordination-driven nanoprobes. This approach enhances the stability and specificity of tumor imaging by utilizing biomarkers associated with rapid tumor proliferation and those commonly overexpressed in tumors. In conclusion, the research not only paves the way for new perspectives in cell biology and pathology studies but also lays a solid foundation for the advancement of biomedical imaging and disease diagnostic technologies.

The VISTA/VSIG3/PSGL-1 axis: crosstalk between immune effector cells and cancer cells in invasive ductal breast carcinoma

A checkpoint protein called the V-domain Ig suppressor of T cell activation (VISTA) is important for controlling immune responses. Immune cells that interact with VISTA have molecules, or receptors, known as VISTA receptors. Immune system activity can be modified by the interaction between VISTA and its receptors. Since targeting VISTA or its receptors may be beneficial in certain conditions, VISTA has been studied in relation to immunotherapy for cancer and autoimmune illnesses. The purpose of this study was to examine the expression levels and interactions between VISTA and its receptors, VSIG3 and PSGL-1, in breast cancer tissues. IHC analysis revealed higher levels of proteins within the VISTA/VSIG3/PSGL-1 axis in cancer tissues than in the reference samples (mastopathies). VISTA was found in breast cancer cells and intratumoral immune cells, with membranous and cytoplasmic staining patterns. VISTA was also linked with pathological grade and VSIG3 and PSGL-1 levels. Furthermore, we discovered that the knockdown of one axis member boosted the expression of the other partners. This highlights the significance of VISTA/VSIG3/PSGL-1 in tumor stroma and microenvironment remodeling. Our findings indicate the importance of the VISTA/VSIG3/PSGL-1 axis in the molecular biology of cancer cells and the immune microenvironment.

Electro-osmotic Flow Generation via a Sticky Ion Action.

Selective transport of ions through nanometer-sized pores is fundamental to cell biology and central to many technological processes such as water desalination and electrical energy storage. Conventional methods for generating ion selectivity include placement of fixed electrical charges at the inner surface of a nanopore through either point mutations in a protein pore or chemical treatment of a solid-state nanopore surface, with each nanopore type requiring a custom approach. Here, we describe a general method for transforming a nanoscale pore into a highly selective, anion-conducting channel capable of generating a giant electro-osmotic effect. Our molecular dynamics simulations and reverse potential measurements show that exposure of a biological nanopore to high concentrations of guanidinium chloride renders the nanopore surface positively charged due to transient binding of guanidinium cations to the protein surface. A comparison of four biological nanopores reveals the relationship between ion selectivity, nanopore shape, composition of the nanopore surface, and electro-osmotic flow. Guanidinium ions are also found to produce anion selectivity and a giant electro-osmotic flow in solid-state nanopores via the same mechanism. Our sticky-ion approach to generate electro-osmotic flow can have numerous applications in controlling molecular transport at the nanoscale and for detection, identification, and sequencing of individual proteins.