Search Results (392)

Cyclic nucleotide signaling pathways play essential roles in the physiology of the nematode Caenorhabditis elegans, influencing processes such as reproduction, environmental sensing, and cellular homeostasis. The intracellular levels of cAMP and cGMP are tightly regulated by their synthesis by adenylyl and guanylyl cyclases and their degradation catalyzed by 3′,5′-cyclic nucleotide phosphodiesterases (PDEs). Mammals possess eleven PDE families (PDE1 through PDE11), whereas nematode genomes contain six PDE genes orthologous to six of the mammalian PDE families. Despite their evolutionary conservation, the signaling pathways, regulatory mechanisms, and enzymatic properties of nematode PDEs remain incompletely understood. This review synthesizes current knowledge on the regulation of cyclic nucleotide levels in C. elegans, highlighting how dysregulation of nematode PDEs affects a wide range of physiological and behavioral processes, including sensory transduction, development, and locomotion. Full article

Prolactin (PRL) is a hormone primarily associated with lactation, but it plays various roles in both men and women. PRL belongs to the family of peptide hormones, including placental lactogen and growth hormone. Interestingly, PRL is a pleiotropic hormone affecting several physiological and pathological conditions, including fertility. Moreover, several pathophysiological roles have been associated with this hormone, including those of the immune system, autoimmune disorders, asthma, and ageing. Additionally, PRL receptors are ubiquitously expressed in tissues, including the mammary gland, gonads, liver, kidney, adrenal gland, brain, heart, lungs, pituitary gland, uterus, skeletal muscle, skin blood cells, and immune system. Therefore, in the present paper, we cover the potential role that PRL may play in asthma by promoting inflammation and modulating immune responses. The detection of its receptor in lung tissue suggests a direct role in airway smooth muscle contractility through activation of signaling pathways such as JAK2-STAT5, MAPK/ERK1/2, and PI3K/Akt, as well as influencing ionic currents that regulate cell contraction, proliferation, and survival. In this sense, this review aims to explore the potential involvement of PRL in asthma pathophysiology by examining its interactions with intracellular signaling pathways and its possible impact on airway smooth muscle contractility and immune modulation. Full article

Biofilm-associated infections caused by Gram-negative bacteria, especially multidrug-resistant strains, frequently occur in intensive care units and represent a major therapeutic challenge. The economic burden of biofilm-associated infections is considerable, making the search for new treatment approaches a focal point for policymakers and scientific funding bodies. Biofilm formation is regulated by quorum sensing (QS), a population density-dependent communication mechanism between cells mediated by small diffusible signaling molecules. QS modulates various intracellular processes, and some features of QS are common to all Gram-negative bacteria. While there are differences in the QS regulatory networks of different Gram-negative bacterial species, a common feature of most Gram-negative bacteria is the ability of N-acylhomoserine lactones (AHL) as inducers to diffuse across the bacterial membrane and interact with receptors located either in the cytoplasm or on the inner membrane. Targeting QS by inhibiting the synthesis, transport, or perception of signaling molecules using small molecules, quorum quenching enzymes, antibodies, combinatorial therapies, or nanoparticles is a promising strategy to combat virulence. In-depth knowledge of biofilm biology, antibiotic susceptibility, and penetration mechanisms, as well as a deep understanding of anti-QS agents, will contribute to the development of antimicrobial therapies to combat biofilm infections. Advancing antimicrobial therapies against biofilm infections requires a deep understanding of biofilm biology, antibiotic susceptibility, penetration mechanisms, and anti-QS strategies. This can be achieved through in vivo and clinical studies, supported by state-of-the-art tools such as machine learning and artificial intelligence. Full article

The aging of dry meat enhances its flavor and tenderness; however, continuous internal quality monitoring throughout the aging process is challenging. We developed and validated a novel electromagnetic response measurement system for meat aging that enables continuous bioimpedance monitoring under stable, optimal temperature/humidity conditions. The system comprises a temperature-controlled dry aging fridge and a newly designed puncture-type semi-rigid coaxial probe, allowing for minimally invasive internal measurements over a broad frequency range. The probe achieved stable measurements across 10 kHz to 10 MHz, and its small diameter (1.25 mm) enabled almost non-destructive internal sensing. Beef and pork samples were monitored over 14 days via multi-channel bioimpedance measurements. After an initial stabilization period, bioimpedance steadily decreased throughout aging. This decline reflected progressive increases in tissue conductivity as cell membranes broke down and intracellular fluids leaked out. High-frequency measurements (e.g., around 10 MHz) were more sensitive to environmental disturbances. Periodic defrost cycles in the chamber caused temporary impedance dips at these frequencies, highlighting the influence of short-term temperature/humidity fluctuations. The system enables long-term continuous measurement without removing samples from the fridge, thus maintaining aging conditions during monitoring. Overall, the system enables the stable, long-term, and multi-channel electromagnetic monitoring of meat quality under optimal aging conditions—a capability not achieved in previous studies. This new method offers a minimally invasive, frequency-resolved approach for assessing meat quality evolution during aging. This advance demonstrates a new approach for tracking meat quality changes during dry aging. Full article

With the emergence of the SARS-CoV-2 pandemic the process of coronavirus replication has been under increasing scrutiny. During the replication of their genomic RNA, coronaviruses produce a range of other RNAs in addition to the negative-sense replicative intermediates of the genome, which includes a set of subgenomic RNAs. These subgenomic RNAs are nested within the sequence of the complete genome and can be both replicated further and act as templates for protein production. Alongside these functional products of discontinuous replication, coronaviruses produce defective viral genomes that can potentially impact both the virus and infected host cells. These interactions can arise from the ability of these defective viral genomes to impact the production of new infectious virions, through either competition with the wild-type genome for replication or by stimulating an antiviral response. Examining the behaviour of defective viral genomes can also help to elucidate the functional elements of the genome involved in the processes of replication and packaging. This review covers the process of intracellular replication by coronaviruses describing the mechanisms by which the different RNA species are produced. Of particular focus are factors involved in discontinuous replication that produces defective viral genomes, and the behaviour of coronavirus defective viral genomes. Full article

IR-780 is a heptamethine cyanine dye that exhibits strong absorbance in the near-infrared region. Herein, we report IR-780 dye as a dual sensor for chromogenic cyanide detection and azide’s fluorogenic sensing in acetonitrile. Cyanide and hydroxide cause instant, dramatic color changes in the dye solution from green to yellow and dramatic spectral changes in the UV-Vis spectrum. The interaction of cyanide and hydroxide with the dye caused a dramatic decrease in the intensity of the strong absorption band at 780 nm and a concomitant band appearance at 435 nm. Other monovalent ions, including fluoride, chloride, bromide, iodide, dihydrogen phosphate, thiocyanate, acetate, and dihydrogen arsenate, caused no significant color or spectral changes. UV-Vis studies showed that the IR-780 dye is sensitive and selective to both ions. The detection limits for cyanide and azide are 0.39 µM and 0.50 µM, respectively. Interestingly, the IR-780 dye exhibited strong fluorescence at 535nm upon interaction with azide, while its initial emission at 809 nm was quenched. Both UV-Vis and fluorescence spectroscopy accomplished the detection of cyanide and azide using IR-780. Furthermore, the sensor’s effectiveness in fluorescence imaging of intracellular CN⁻ ions is demonstrated in live HeLa cells. Full article

In this paper, we present a simple solvothermal method to synthesize highly fluorescent metal-free elemental selenium quantum dots (SeQDs) using cost-effective bulk selenium powder. The SeQDs exhibit a small and uniform size, excellent aqueous dispersibility, a high photoluminescence quantum yield (PLQY) of 19.3% with stable fluorescence, and scalable production with a 7.2% yield. Owing to the inner filter effect (IFE), these SeQDs function as a highly effective nanoprobe for Cr(VI) detection, exhibiting exceptional sensitivity (detection limit: 145 nM) and selectivity over a wide linear range (5–105 μM), along with rapid response kinetics. Moreover, SeQDs show low cytotoxicity and efficient cellular uptake, enabling cell imaging and intracellular Cr(VI) monitoring. Significant fluorescence quenching in Cr(VI)-exposed cells confirms the potential of SeQDs as a viable fluorescent nanoprobe for Cr(VI) detection in complex cellular environments. This work thus not only establishes a simple method for the preparation of fluorescent SeQDs but also develops a promising fluorescent nanoprobe for cell imaging and Cr(VI) sensing. Full article

Hepatitis E virus (HEV) is a positive-sense, single-stranded RNA virus that poses a significant public health risk, yet its study is hindered by the complexity of conventional RNA-based reverse genetics systems. These systems require multiple steps, including genome cloning, in vitro transcription, and capping, making them labor-intensive and susceptible to RNA degradation. In this study, we developed a single-step, plasmid-based HEV expression system that enabled direct intracellular transcription of the full-length HEV genome under a cytomegalovirus immediate-early (CMV-IE) promoter. The viral genome was flanked by hammerhead (HH) and hepatitis delta virus (HDV) ribozymes to ensure precise self-cleavage and the generation of authentic 5′ and 3′ termini. This system successfully supported HEV genome replication, viral protein expression, and progeny virion production at levels comparable to those obtained using in vitro-transcribed, capped HEV RNA. Additionally, a genetic marker introduced into the plasmid construct was stably retained in progeny virions, demonstrating the feasibility of targeted genetic modifications. However, plasmid-derived HEV exhibited delayed replication kinetics, likely due to the absence of an immediate 5′ cap. Attempts to enhance capping efficiency through co-expression of the vaccinia virus capping enzyme failed to improve HEV replication, suggesting that alternative strategies, such as optimizing the promoter design for capping, may be required. This plasmid-based HEV reverse genetics system simplifies the study of HEV replication and pathogenesis and provides a versatile platform for the genetic engineering of the HEV genome. Full article

Vitamin D has been shown to improve immunity as well as vascular function. We investigated the effect of cholecalciferol on T-cell phenotype in cultured peripheral blood mononuclear cells (PBMCs) from twenty vitamin D-deficient, non-diabetic chronic kidney disease (CKD) subjects. We also studied vitamin D effects on endothelial and vascular function markers in human aortic endothelial cells (HAECs) and in human aortic smooth muscle cells (HASMCs), respectively. We studied endothelial nitric oxide synthase (eNOS), mitogen-activated protein kinase 38 (p38 Map kinase), protein kinase B (Akt), and nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase) in HAECs and α-smooth muscle actin (α-SMA), smooth muscle calponin (SM-Calponin), smooth muscle myosin heavy chain (SM-MHC), and calcium-sensing receptor (CaSR) in HASMCs. Vitamin D receptors (VDRs) and CYP27B1 were studied in both cell types. In cultured PBMCs isolated from CKD subjects, the percentage of T helper 1(TH1) cells significantly decreased while that of T helper 2 (TH2) cells increased after cholecalciferol treatment. No significant change in intracellular and surface markers of T helper 17 (TH17) and T regulatory (Treg) cells was observed. In vitro treatment of HASMCs and HAECs with cholecalciferol led to significant and favorable alterations in mRNA expression of markers of vascular smooth muscle cells, i.e., α-SMA, SM-Calponin, and SM-MHC. Regarding endothelial cell markers, mRNA encoding eNOS, p38 Map kinase, protein kinase B (Akt), NADPH oxidase, VDR, and CYP27B1 were also significantly changed. Finally, the expression levels of the following proteins were notably altered: NADPH oxidase and protein kinase B (Akt) (in HAECs); SM-MHC and SM-Calponin (in HASMCs). In vitro treatment of PBMCs with cholecalciferol led to a favorable change in T-cell population, decreasing TH1 and increasing TH2 cell percentage, along with beneficial alterations in mRNA expression of HASMCs and HAECs’ cell markers. This study provides evidence that cholecalciferol can influence immune and vascular function in CKD. Full article

Molecular analyses of individual cells with high resolution, specificity, and sensitivity can not only reveal cellular heterogeneity but also provide a better understanding of diseases and accelerate drug discoveries. Single-cell endoscopy is an advanced live-cell technique that relies on a smart endoscope that allows minimally invasive probing of the interiors of individual cells. Compared with other single-cell analysis techniques, single-cell endoscopy has shown great promise in applications such as flexible single-cell manipulation, ultrasensitive sensing, and precise intracellular delivery. In this review, we aim to map out the landscape of recent advances in single-cell endoscopy techniques by focusing on both fundamental considerations and significant progress over the past decade. Specifically, we summarize the predominant live-cell endoscopes, including their fabrication and characterization. Furthermore, a series of valuable intracellular molecular sensing events, such as nucleic acids, proteins, ions, etc., are introduced with a main emphasis on how single-cell endoscopy can solve these issues and what merits single-cell endoscopy can provide. Finally, we briefly outline the remaining challenges and directions for the future development of single-cell endoscopy techniques. Full article

Idiopathic pulmonary fibrosis (IPF) is a disease with a poor prognosis and no curative therapies. Fibroblast activation by transforming growth factor β1 (TGFβ1) and disrupted metabolic pathways, including the arginine–polyamine pathway, play crucial roles in IPF development. Polyamines are agonists of the calcium/cation-sensing receptor (CaSR), activation of which is detrimental for asthma and pulmonary hypertension, but its role in IPF is unknown. To address this question, we evaluated polyamine abundance using metabolomic analysis of IPF patient saliva. Furthermore, we examined CaSR functional expression in human lung fibroblasts (HLFs), assessed the anti-fibrotic effects of a CaSR antagonist, NPS2143, in TGFβ1-activated normal and IPF HLFs by RNA sequencing and immunofluorescence imaging, respectively; and NPS2143 effects on polyamine synthesis in HLFs by immunoassays. Our results demonstrate that polyamine metabolites are increased in IPF patient saliva. Polyamines activate fibroblast CaSR in vitro, elevating intracellular calcium concentration. CaSR inhibition reduced TGFβ1-induced polyamine and pro-fibrotic factor expression in normal and IPF HLFs. TGFβ1 directly stimulated polyamine release by HLFs, an effect that was blocked by NPS2143. This suggests that TGFβ1 promotes CaSR activation through increased polyamine expression, driving a pro-fibrotic response. By halting some polyamine-induced pro-fibrotic changes, CaSR antagonists exhibit disease-modifying potential in IPF onset and development. Full article

The inner ear and/or lateral line are responsible for hearing and balance of vertebrate. The otic sensory hair cells (HCs) employ cilium organelles, namely stereocilia and/or kinocilia, to mediate mechanical stimuli to electrical signal transition. Tektins (Tekts) are known as the cilium microtubule stabilizer and inner-space filler, and four Tekt(1-4)-encoding genes are identified in zebrafish HCs, but the subcellular location of Tekts in HCs remains unknown. In the present study, we first found that tekt3 is expressed in the inner ear and lateral line neuromast. Antibody staining revealed that Tekt3 is present in neuromast and utricular HCs. It is absent in the saccule, the authentic hearing end-organ of zebrafish and the crista of semi-circular canals. Furthermore, Tekt3 were enriched at the apical side of neuromast and utricular HCs, mainly in the cytosol. Similar subcellular distribution of Tekt3 was also evident in the outer HCs of mature mouse cochlea, which are not directly linked to the hearing sense. However, only neuromast HCs exerted morphological defect of kinocilia in tekt3 mutant. The disrupted or distorted HC kinocilia of mutant neuromast ultimately resulted in slower vital dye intake, delayed HC regeneration after neomycin treatment, and reduced startle response to vibration stimulation. All functional defects of tekt3 mutant were largely rescued by wild-type tekt3 mRNA. Our study thus suggests that zebrafish Tekt3 maintains the integrity and function of neuromast kinocilia to against surrounding and persistent low-frequency noises, perhaps via the intracellular distribution of Tekt3. Nevertheless, TEKT3/Tekt3 could be used to clarify HC sub-types in both zebrafish and mice, to highlight the non-hearing HCs. Full article

The regulation of intracellular NADPH levels is currently a hotspot for research into bacterial modification and fermentation process optimization, and Corynebacterium glutamicum, an important industrial microorganism, achieves enhanced L-lysine production by regulating intracellular NADPH levels. In previous studies, transcriptome analysis was performed on C. glutamicum with different intracellular NADPH levels. The results showed that the expression level of transcription factor AtrN changed significantly. Moreover, experiments showed that transcription factor AtrN can sense high intracellular levels of NADPH and negatively regulate its synthesis. In this study, we integrated the pntAB gene of Escherichia coli into the genome of C. glutamicum XQ-5, successfully constructing a chassis cell with a high intracellular NADPH level. It was named TQ-1. On this basis, we knocked out and complemented the AtrN in strain TQ-1, resulting in strains TQ-2 and TQ-3, respectively. Then, the changes in cell growth, intracellular redox substances and cell membrane among these three strains were investigated. We found that the growth of TQ-2 was inhibited in the early growth stage and the cell survival rate was decreased because of the high increase in the intracellular NADPH level. In addition, the deletion of the AtrN gene also led to a decrease in the fluidity and an increase in the permeability of the cell membrane. Compared with TQ-1, TQ-3 showed slow growth only in the late growth stage, and the fluidity of its cell membrane was also enhanced. This indicates that AtrN guides the cells to make some adaptive changes to maintain cell growth when facing excessive intracellular reductive stress. This will facilitate future research on how potential upstream regulatory genes regulate AtrN and how AtrN regulates downstream genes to cope with cellular reductive stress. It also provides theoretical guidance for the specific modification of high-yield lysine-producing strains. Full article

Phages, the most abundant and diverse lifeforms on Earth, require strict parasitism for survival. During infection, temperate phages integrate both intracellular and extracellular host information to decide between lysis and lysogeny for replication. While various environmental and physiological factors influence the lysis–lysogeny decision, recent insights into phage–bacterium interactions reveal phages’ ability to communicate with and influence bacteria, leveraging the host’s quorum sensing system or small molecular signals. This article provides a succinct overview of current research advancements in this field, enhancing our understanding of phage–host dynamics and providing insights into bacteria’s multicellular behavior in antiviral defense. Full article

The IκB kinase (IKK) complex plays a central role in many signaling pathways that activate NF-κB, which turns on a battery of genes important for immune response, inflammation, and cancer development. Ubiquitination is one of the most prevalent post-translational modifications of proteins and is best known for targeting substrates for proteasomal degradation. The investigations of NF-κB signaling pathway primed the unveiling of the non-degradative roles of protein ubiquitination. The NF-κB-essential modulator (NEMO) is the IKK regulatory subunit that is essential for IKK activation by diverse intrinsic and extrinsic stimuli. The studies centered on NEMO as a polyubiquitin-binding protein have remarkably advanced understandings of how NEMO transmits signals to NF-κB activation and have laid a foundation for determining the molecular events demonstrating non-degradative ubiquitination as a major driving element in IKK activation. Furthermore, these studies have largely solved the enigma that IKK can be activated by diverse pathways that employ distinct sets of intermediaries in transmitting signals. NEMO and NEMO-related proteins that include optineurin, ABIN1, ABIN2, ABIN3, and CEP55, as non-degradative ubiquitin chain receptors, play a key role in sensing and transmitting ubiquitin signals embodied in different topologies of polyubiquitin chains for a variety of cellular processes and body responses. Studies of these multifaceted proteins in ubiquitin sensing have promoted understanding about the functions of non-degradative ubiquitination in intracellular signaling, protein trafficking, proteostasis, immune response, DNA damage response, and cell cycle control. In this review, I will also discuss how dysfunction in the NEMO family of protein-mediated non-degradative ubiquitin signaling is associated with various diseases, including immune disorders, neurodegenerative diseases, and cancer, and how microbial virulence factors target NEMO to induce pathogenesis or manipulate host response. A profound understanding of the molecular bases for non-degradative ubiquitin signaling will be valuable for developing tailored approaches for therapeutic purposes. Full article