This study investigated whether L-NAME, a nitric oxide synthase (NOS) inhibitor commonly used to model hypertension-associated endothelial dysfunction, can exert direct pro-atherogenic effects on vascular smooth muscle cells (VSMCs) independent of systemic hemodynamic influences. Mouse aortic VSMCs (MOVAS) were treated with non-cytotoxic concentrations of L-NAME (50 µM) or oxidized low-density lipoprotein (ox-LDL, 100 µg/mL), which served as a positive atherogenic control. Lipid accumulation was assessed by Oil Red O staining. The expression of contractile phenotype markers (ACTA2, MYH11, and α-SMA), the synthetic phenotype marker SPP1, and matrix metalloproteinases (MMP-2 and MMP-9) was quantified by qRT-PCR and immunofluorescence. Both L-NAME and ox-LDL treatments caused marked intracellular lipid accumulation in MOVAS cells, consistent with VSMC-derived foam cell formation. L-NAME and ox-LDL reduced the mRNA expression of contractile phenotype markers (ACTA2 and MYH11, p < 0.05) and decreased α-SMA protein expression (p < 0.001). Conversely, both treatments increased the mRNA expression of the synthetic phenotype marker SPP1 (p < 0.05) and matrix-remodeling genes MMP-2 and MMP-9 (p < 0.05). These effects were directionally similar to those induced by ox-LDL. L-NAME directly promotes lipid accumulation and synthetic phenotype-associated marker changes in MOVAS cells under the present in vitro conditions. Because eNOS/NOS activity, intracellular NO levels, rescue pathways, and primary VSMC validation were not assessed, these data should be interpreted as phenotypic evidence that NOS/NO pathway disruption may contribute to VSMC dysfunction rather than definitive proof of a single eNOS-dependent mechanism. Not applicable.
Chronic dacryocystitis, characterized by epiphora, is primarily treated with dacryocystorhinostomy (DCR), yet postoperative anastomotic re-obstruction remains a significant cause of failure. This study aimed to identify the key molecular mechanisms driving this recurrence. Nasal mucosal tissues were collected from 30 post-DCR patients: 15 with re-obstruction (proliferation group) and 15 with successful outcomes (non-proliferation group). Proteomic analysis based on LC-MS/MS was conducted, with differentially expressed proteins (DEPs) defined by a fold change ≥ 1.5 and p < 0.05. Western blot and immunohistochemistry were used for validation and localization. The role of RBM39 in vascular endothelial cell proliferation was assessed via functional experiments, and its associated pathway was analyzed. Proteomics identified RBM39 as significantly upregulated in re-obstruction tissues, confirmed to be expressed in the vascular endothelium. In vitro, RBM39 overexpression enhanced HMEC-1 proliferation, while its knockdown suppressed it. KEGG pathway analysis suggested the Focal Adhesion pathway may be involved as a potential downstream mechanism. This study observes that RBM39 is upregulated at the anastomotic site and correlates with enhanced vascular endothelial cell proliferation. The Focal Adhesion pathway may be implicated in this process, suggesting RBM39 as a candidate target for mitigating postoperative ostium re-obstruction after En-DCR. However, further mechanistic studies are needed to establish a direct causal relationship and evaluate its therapeutic potential.
Abstract Background When starved, Dictyostelium discoideum cells form clusters that, when constrained in height, exhibit clear collective rotational motion. During this collective rotation, cells secrete and relay the chemoattractant cAMP, resulting in signal propagation in the form of spiral waves, which rotate in the opposite direction of the collective rotation. The quantification of this collective rotation with reference to cAMP signaling and the dependence on cluster size is currently unclear. Results In this study, we use experiments and modeling to investigate how cell motion and cAMP signaling dynamics depend on cluster sizes in Dictyostelium using aggregates that are confined to quasi-2D environments. For rotating clusters with a single cAMP spiral arm, we find that both cAMP wave and cell angular velocities decrease as cluster sizes increase, suggesting that larger clusters signal more slowly. We also show that the angular velocity of the cells ( $$\omega_{\text{cell}}$$ ) is linearly correlated with the angular velocity of the cAMP wave ( $$\omega_{\text{wave}}$$ ). For clusters with multiple spiral arms, we observe a size dependence where larger clusters have a higher probability of having more spiral arms compared to smaller clusters. The qualitative experimental results are consistent with a computational model that couples a reaction diffusion model to cell motility and chemotaxis. Our simulation results suggest that a variable degradation rate of cAMP, which depends on cluster size, is a possible mechanism to explain the cluster size-dependent wave and cell angular velocities. Conclusion Together, these findings identify cluster size as a key regulator of both cAMP wave dynamics and collective cell motion and link slower signaling and enhanced multi-armed spiral formation to larger clusters. More broadly, our work suggests that size-dependent modulation of extracellular cAMP degradation can couple cell number to emergent spatiotemporal patterns in developing Dictyostelium aggregates.
BACKGROUND:Primary cilia are essential sensory organelles, and their dysfunction leads to a broad spectrum of diseases known as ciliopathies. Immunofluorescence imaging of ciliary substructures is a key tool in cilia research, but multiplex staining is often limited by the availability of antibodies from different host species. METHODS:We developed a novel goat-derived gamma tubulin antibody for basal body labelling and validated its specificity through triple-label immunofluorescence in different cell lines. RESULTS:The antibody showed strong, specific staining of basal bodies under methanol fixation, enabling clear visualization in combination with rabbit- and mouse-derived antibodies. CONCLUSIONS:This antibody expands multiplex imaging capabilities for ciliary substructures, supporting both cilia research and diagnostics of ciliopathies.
BACKGROUND:Renal cyst formation, as observed in autosomal dominant polycystic kidney disease (ADPKD), is a life-threatening condition with no effective cure yet. The molecular mechanisms underlying primary cilia dysfunction, which causes cyst formation and disease development, are not well understood. Human kidney tubuloids offer a promising model system to investigate the disease mechanisms of PKD in physiologically relevant 3D structures. However, their inherent cystic morphology poses a challenge in effectively modelling kidney cystogenesis. Therefore, our study aims to refine the culture method of tubuloids and assess the efficacy of these modified cultures in modeling cyst formation and development. RESULTS:We developed human kidney tubuloid models derived from adult kidney tubular cells using different methods for 3D in-vitro cultures. Tubuloids cultured in suspension or an extracellular matrix scaffold manifested distinctly polarized epithelial structures. Bulk RNA sequencing and immunohistochemistry revealed differential transcriptional profiles, highlighting variations in cellular composition and cellular fate within the kidney epithelium between the two types of tubuloids. Notably, the experimental activation of chronic cAMP signalling promoted cyst formation in vitro, validating the suitability of these tubuloids for studying kidney cystogenesis. Furthermore, we demonstrate that tubuloids are amenable to genetic modification through recombinant adeno-associated virus transduction. CONCLUSIONS:Our study identifies different in-vitro tubuloid cultures as relevant model systems for examining the molecular and cellular changes involved in kidney cystogenesis in humans. These models will enhance our capability to discover novel pathogenetic mechanisms underlying ADPKD and validate candidate drugs for clinical application.
Background Alcohol consumption has been a risk factor for nearly 200 diseases. Although swallowing an alcoholic beverage takes only a few seconds, the substance can stick to the esophageal lining longer after drinking. What alcohol does to the esophageal cells is not fully understood. This study explored the issue through in vitro experimentation. Results It was found that < 0.5% alcohol or a < 30-minute exposure benefited cell growth primarily through autophagy, while > 0.5% alcohol or a > 30-minute exposure caused cell death, including apoptosis, necroptosis, and ferroptosis, but not pyroptosis. Autophagy took place within the first 30 min of alcohol exposure, followed by apoptosis, which peaked at 2 h, and then was gradually replaced by necroptosis as the alcohol exposure continued. Ferroptosis initiated immediately after contact with alcohol and became dominant progressively. However, as alcohol exposure continued, cells developed tolerance through anastasis, reviving and reforming the epithelial monolayer. Conclusions Low concentrations of alcohol or a short exposure promote esophageal cell growth owing to autophagy, while higher concentrations or a longer exposure cause multiple forms of cell death, including apoptosis, necroptosis, and ferroptosis. Yet, cells can recover from alcohol insult through anastasis as the exposure extends.
This study investigates the role of Th17 cells and IL-17 signaling in mediating therapeutic resistance and early recurrence in cervical carcinoma. Th17 cells were generated in vitro, and cervical carcinoma cells were subsequently exposed to cisplatin in the presence of Th17-conditioned media or recombinant IL-17. To elucidate the underlying molecular mechanisms, synthetic siRNAs targeting Akt1 (si-Akt1) and Akt2 (si-Akt2) were employed. Gene expression levels were quantified using ABIVII7 qRT-PCR, and phosphorylation of AKT at Thr308 and Ser473 was assessed. Cervical carcinoma cell lines (HeLa and SW756) were cultured and subjected to siRNA-mediated knockdown of AKT1 and AKT2. Cytotoxicity assays were conducted to evaluate cell viability under various treatment conditions. Cells were stimulated with either standard medium or conditioned medium from in vitro-differentiated Th17 cells. Th17 cell-induced resistance to cisplatin and radiation co-treatment was found to be mediated via the AKT signaling pathway. Moreover, Th17 cells promoted chemoradiotherapy tolerance in cervical cancer cells, with a potential contribution from IL-17 signaling. Although chemoradiotherapy significantly decreased cell viability across all cell lines, preconditioning with recombinant IL-17 notably mitigated this effect, resulting in increased cellular survival. These findings suggest a critical role for Th17 cells in modulating the therapeutic response and recurrence risk in cervical carcinoma through AKT pathway activation.
Abstract Background The ribosomal DNA (rDNA) of the myxomycete Didymium iridis is located on a linear, multi-copy, non-Mendelian chromosome. Efforts to determine the complete sequence by short-read sequencing technologies have been prevented by the presence of highly repetitive regions. Here we use high coverage (~10,000 x) long-read Oxford Nanopore Technology to determine the rDNA chromosome sequence in haploid amoebae from telomere-to-telomere. Results The 20 kb rDNA chromosome, which is present at ~ 132 copies per haploid genome, is capped by regular TTAGGG telomeric repeats at both ends and carries an 11.3 kb pre-rRNA transcription unit coding for the small and large subunit rRNAs. The rRNA genes are further interrupted by autocatalytic group I introns, one of which encodes a homing endonuclease and two catalytic RNA domains with different functions in RNA processing. RNA mapping analyses from amoeba, microcyst, flagellate, and plasmodium stages, based on Illumina short-read sequencing, support the presence of a mature intron homing endonuclease mRNA both in haploid and diploid life stages in D. iridis. The non-transcribed sequence region upstream of the transcription unit contains several direct repeat arrays, including a highly complex upstream promoter region likely to be involved in pre-rRNA transcription regulation. Adjacent to the upstream telomere, a 4.2 kb palindromic region with potential for cruciform structure formation is found. Here, two putative replication origin candidates are located. Conclusions High coverage Oxford Nanopore Technology sequencing results in excellent resolution of complex sequence repeat feature in the D. iridis rDNA chromosome. The rRNA genes are interrupted by complex group I introns and RNA sequencing supports intron autocatalytic processing in haploid and diploid life stages. This study provides new insights into structural arrangements of nuclear rDNA in eukaryotic microorganisms.
Three-dimensional (3D) organoid cultures from pancreatic tissue are emerging tools for studying pancreatic epithelial development, differentiation, and disease. While models derived from human fetal tissue or adult murine pancreas have provided key insights, their clinical translatability is limited by ethical and species-related differences. The domestic dog offers a valuable intermediate model for translational research, yet pancreatic organoid systems in this species remain underdeveloped. We successfully established long-term organoid cultures from adult canine pancreas and demonstrated their stable ductal phenotype. These organoids exhibited limited but detectable expression of acinar and endocrine markers, indicating partial trilineage potential. We further developed two-dimensional (2D) monolayers from organoids and assessed epithelial barrier integrity using transepithelial electrical resistance (TEER). Monolayers maintained tight junctions and showed media-dependent changes in morphology and KRT19 expression. Peak TEER values exceeded 4000 Ω·cm², confirming robust epithelial barrier function. This study presents a reproducible protocol for deriving organoids and monolayers from adult canine pancreas, offering a physiologically relevant and ethically accessible model for investigating pancreatic epithelial biology. The combination of 3D and 2D systems enables both lineage characterization and functional assessment, providing a foundation for future applications in regenerative medicine, disease modeling, and comparative translational research.
BACKGROUND: Previous studies have shown that Dictyostelium discoideum cells lacking the actin-regulating protein coronin A have a large hyaline zone at the front of the cell. However, the coronin mutant cells can efficiently navigate in a gradient of chemoattractant by extending rounded protrusions from the hyaline zone. This study examines whether this zone is occupied by actin filaments, as was previously assumed, or if it is free of filamentous actin, as would be the case for typical blebs. RESULTS: The lack of coronin A results in a large hyaline region in the anterior part of mutant cells, from which the endoplasmic reticulum is displaced. This zone is populated with filamentous actin. Despite this broadened front, the coronin-null cells can respond in a gradient of chemoattractant even when they lack actin-based structures such as filopodia or lamellipodia. During re-orientation into a changed direction of the gradient, the mutant cells form rounded protrusions in various directions, of which the one pointing to the direction of the gradient will expand and become the new front. Contact with a substratum is not necessary for a protrusion to be formed in the right direction. These results illustrate the altered but efficient chemotactic responses of cells under conditions of diminished actin filament turnover. CONCLUSIONS: Despite an impaired actin filament turnover, Dictyostelium discoideum cells lacking coronin A respond to a gradient of chemoattractant by extending rounded protrusions from their large hyaline fronts. These protrusions are rich in filamentous actin, and filopodia or lamellipodia are not required. This mode of chemotactic migration is different from a bleb-driven mode described for Dictyostelium and other eukaryotic cells.
Acute kidney injury (AKI) involves complex crosstalk between tubular epithelial cells (TECs) and immune cells. Extracellular vesicles (EVs) and ageing have emerged as modulators of this process. We investigated the effects of circulating plasma EVs from young (Y_EVs) and old (O_EVs) mice on hypoxic TEC stress, using bone marrow–derived macrophages (BMDMs) from young or old mice, which were pre-conditioned with EVs. First, direct effects of EVs on TEC viability were assessed following hypoxia/reoxygenation injury. We then examined macrophage responses to hypoxic TEC secretomes with or without EV priming, and tested whether EV-primed BMDM secretomes modulated TEC damage, in terms of viability, cytokine expression, and epithelial–mesenchymal transition (EMT). EVs exerted no direct protective effect on hypoxic TECs, and no significant differences were observed between Y_EVs and O_EVs in any condition. Instead, macrophage donor age was the most significant determinant of outcomes. Conditioned medium from old BMDMs consistently improved TEC viability compared to young BMDMs. In agreement with this, the old mouse BMDM secretome also suppressed hypoxia-induced cytokine expression (Il6, Ccl2) and EMT marker induction (Acta2, Fn1) in hypoxic TECs. EV pre-treatment of BMDMs did not significantly alter these effects in either macrophage group. Thus, BMDM donor age, rather than EV donor age, was the critical driver of cytoprotection and inflammatory modulation in this model. Circulating plasma EV age had no significant direct or indirect impacts on TEC viability, inflammatory responses or EMT processes under the tested conditions. This contrasts with prior studies which have suggested broadly rejuvenating effects of young EVs and detrimental effects of old EVs. Not applicable.
Background Dictyostelium discoideum has a unique proteome among sequenced organisms that encodes nearly 10,000 homopolymeric amino acid tracts longer than 10 amino acids long. These repeats are composed of every amino acid except tryptophan with asparagine and glutamine being the most prevalent. Interestingly, asparagine and glutamine-rich regions of proteins are the hallmark of prion-like domains and nearly 25% of the Dictyostelium proteome is predicted to be prion-like in nature.ResultsHere we assessed the insoluble proteome upon heat stress or during nutrient stress which induces Dictyostelium development. We found that both heat and nutrient stress induce the accumulation of predicted prion-like proteins in the insoluble fraction; however, the overall amino acid composition of insoluble proteins is different depending on the stress with a greater percentage of predicted prion-like proteins becoming insoluble upon nutrient stress. We further confirmed that endogenous polyglutamine proteins accumulate in the insoluble fraction over the developmental time course and demonstrate that exogenously expressed polyglutamine tracts form puncta and have reduced solubility in a polyglutamine length dependent manner upon nutrient stress. Finally, using a proximity labeling approach, we found that exogenously expressed polyglutamine tracts have enhanced proximity to glutamine-rich proteins.ConclusionDuring heat and nutrient stress Dictyostelium have numerous proteins that have decreased solubility. Among these proteins, predicted prion-like proteins are enriched and the presence of a polyglutamine tract is sufficient to cause decreased solubility in a polyglutamine length dependent manner.
Transfer RNAs (tRNAs) are the decoders of the protein coding genetic information, as they transfer amino acids into nascent proteins during messenger RNA translation. This pivotal role makes tRNAs a source of translation regulation that can affect protein synthesis. Still, we are beginning to understand the upstream mechanisms regulating tRNA pools themselves. In Dictyostelium discoideum, starvation of a sufficient number of individuals, triggers the development of a coordinated sporulation response denominated the social cycle. By using publicly available transcriptomic, epigenomic, and genomic data, we studied two factors contributing to the regulation of tRNA pools throughout this social cycle. First, the tRNA gene repertoire shows that the compact genome of D. discoideum escapes translational selection as even with a relatively high number of tRNA genes, anticodon and codon frequencies greatly mismatch. This disparity is explained by the overrepresentation of anticodons that can be modified in the wobble position. During the social cycle, the vast majority of tRNA genes lie in nucleosome free regions, indicating that most genes are always contributing to the tRNA pools. However, there is a marked variation in expression levels of the proteins involved in tRNA maturation. This modulation is ultimately mirrored by fine-tuned differential composition of tRNA pools at isodecoder, isoacceptor and isotype levels. Particularly, there is an overall down-regulation in the vegetative to streaming transition. Key elements bypass this down-regulation pattern and taken together, this evidence suggests compensatory mechanisms in tRNA regulation that might rescue translation for the following developmental stages, thus allowing D. discoideum to evolve this remarkable strategy under the pressure of an amino acid scarce environment.
Primary cilia are antenna-like organelles that function as cellular hubs for signaling pathways, including Sonic hedgehog and signaling mediated by the Polycystin-1/Polycystin-2 cation channel complex. Proper regulation of signaling output depends on the dynamic control of ciliary protein composition, which involves intraflagellar transport-mediated trafficking, protein retrieval, and the shedding of extracellular vesicles from cilia. Here we identify ALIX, a protein previously linked to the biogenesis of small extracellular vesicles, as a novel component localized at the base of primary cilia in cultured mammalian cells. We show that ALIX retention at this site requires the ciliary kinesin-3 motor protein KIF13B, which physically interacts with ALIX and the E3 ubiquitin ligase ITCH. In turn, ITCH is enriched at the ciliary base and is essential for ALIX stability. Depletion of either ALIX or ITCH results in elevated ciliary levels of Polycystin-2, while ITCH loss additionally leads to constitutive accumulation of Smoothened, a key Sonic hedgehog effector, within the cilium. Collectively, our findings establish ALIX and ITCH as critical regulators of ciliary membrane protein homeostasis and signaling, acting in coordination with KIF13B to maintain proper ciliary function.
Background The airway epithelial barrier, which defends against external pathogens, is compromised in bronchial asthma. This study investigated the roles of adenosine triphosphate (ATP) and specific purinergic receptors in airway epithelial barrier function. Using the 16HBE14o-human airway epithelial cell line, we measured barrier integrity using transepithelial electrical resistance and the apparent paracellular permeability coefficient. Results ATP enhanced barrier integrity in a dose-dependent manner without affecting cell proliferation. Real-time PCR revealed an increased expression of tight and adherens junction proteins, including E-cadherin and occludin, following ATP stimulation. Our findings suggest that P2Y12 contributes to ATP-related purinergic signaling associated with enhanced airway epithelial barrier integrity. Conclusions These findings suggest that ATP-related purinergic signaling via P2Y12 may contribute to the regulation of airway epithelial barrier integrity in vitro. Further studies in more physiologically relevant systems are warranted to determine the clinical relevance of these observations.
Background Insulin signaling is a conserved regulator of growth, metabolism, and lifespan across metazoans. While its systemic roles are well established, the mechanisms by which insulin coordinates tissue-specific transcriptional programs that underlie distinct functional demands remain incompletely understood. In particular, the differential impact of reduced insulin signaling on different tissues has not been systematically explored. Results We performed a comparative transcriptomic analysis of Drosophila melanogaster olfactory sensory neurons (OSNs) and fat body (Fb) to investigate how reduced insulin signaling remodels gene expression in neural and metabolic tissues. Across both tissue types, insulin reduction suppressed key pathways involved in protein synthesis and mRNA surveillance, indicating shared regulatory responses. However, distinct tissue-specific transcriptional adaptations were also observed. In OSNs, insulin reduction led to the upregulation of synaptic and signaling genes, alongside the downregulation of proteostasis-related factors, suggesting enhanced neural plasticity that may come at the cost of long-term neuronal maintenance. In contrast, the Fb exhibited widespread metabolic suppression accompanied by feedback activation of stress-responsive insulin-like peptide genes, consistent with a shift toward hypometabolic adaptation. Network and pathway analyses revealed that these tissue-specific responses involved distinct regulatory architectures affecting core insulin pathway components and gene families. Conclusions Our findings demonstrate that reduced insulin signaling elicits both shared and divergent transcriptional programs in neural and metabolic tissues of Drosophila melanogaster. These findings reveal how insulin signaling orchestrates tissue-specific transcriptional landscapes that may underlie differential resilience or vulnerability to cognitive and metabolic decline.
Background Generalist consumers can eat a variety of foods, but they are not necessarily equally beneficial. Different feeding histories can influence the fitness of genetically similar individuals. Dictyostelium discoideum is a unicellular generalist predator amoeba that eats bacteria. When bacteria are depleted, amoebae enter the social cycle and aggregate to form a fruiting body, in which about a fifth of the amoebae die to form a stalk while the rest form spores at the top of the stalk. Results In this study, we examined the fitness of D. discoideum when grown on three bacterial prey types: poorly palatable wild-type Pseudomonas vindicans, a more palatable mutant of the same species, and the high-quality food bacterium Klebsiella pneumoniae. We used these three bacteria to examine how feeding history shapes fitness throughout the life cycle by measuring proliferation of amoebae, then following spores through germination, plaque formation, and plaque expansion. Amoebae grew on unpalatable P. vindicans only when seeded at high density, and even then, proliferated more slowly. When amoebae with different feeding histories were mixed and allowed to form chimeric fruiting bodies, those cells fed on palatable versus unpalatable P. vindicans did not differ significantly in their spore contribution, except when under an additional stress of the translational inhibitor G418, which makes cells that previously fed on unpalatable P. vindicans contribute fewer cells in spores. Oddly, the amoebae that fed on the good food bacterium Klebsiella pneumoniae contributed fewer spores when mixed with cells that fed on either of the P. vindicans clones. Surprisingly, though feeding on unpalatable prey was not a hindrance in the social stage, downstream effects were observed. Spores produced on either P. vindicans strain germinated more slowly than those made from amoebae fed on Klebsiella pneumoniae. Conclusions Low prey quality strongly affects D. discoideum fitness during the vegetative stage but does not generally lower social competitive ability. Nevertheless, it does affect fitness after the social cycle by reducing the hatching rate of spores.
Clinical and preclinical evidence suggests that inflammation is closely associated with various arterial diseases. Multiple studies have reported the effects of statins on different cell types, yet the effects of atorvastatin (ATV) and its mechanisms on inflamed HUVECs’ cellular responses are still under investigation. This study investigates how different doses of ATV affect inflammatory responses, extracellular matrix (ECM) regulators, and cell migration capacity assessed by an in vitro scratch wound closure assay in lipopolysaccharide (LPS) stimulated endothelial cells. ATV was applied to cells at different doses (5-10-50 µM) with or without LPS (20 ng/mL). Cell proliferation and toxicity were investigated in the indicated groups. Then, the possible effects of ATV on MMP-2, MMP-9, TIMP-1,TIMP-2 expression levels, scratch-closure (cell migration) time were examined. Gene expression of MMP-2, MMP-9, TIMP-1, and TIMP-2 was quantified by qPCR, while protein levels were determined by Western blotting. Cell migration was evaluated using a scratch assay and real-time imaging system. High-dose ATV was more cytotoxic, while wound closure times showed a numerical increase that did not reach statistical significance under LPS stimulation. While low-dose ATV increased MMP and TIMP expressions, high-dose treatment reduced TIMP-1 and disturbed the MMP/TIMP balance. This imbalance was accompanied by reduced cellular recovery capacity under inflammatory stress. This study highlights the complex cellular effects of ATV under inflammatory conditions, supporting its context- and dose-dependent role in endothelial cell behavior and matrix regulation. These findings highlight the importance of dosage optimization in therapeutic contexts targeting vascular inflammation and provide novel insight into how statin dosing influences endothelial recovery mechanisms, beyond cholesterol regulation.
ObjectiveThe objective of this study was to explore the mechanisms by which urine - derived stem cell exosomes (USCexo) alleviate sepsis - related acute kidney injury (SAKI), as acute kidney injury often complicates sepsis with unclear role of USCexo in SAKI while stem cell exosomes show potential in AKI treatment.MethodsHuman urine - derived stem cells (USCs) were isolated from human urine and characterized via flow cytometry. A sepsis - related acute kidney injury mouse model was induced by cecal ligation and puncture (CLP). The mice were divided into a control group receiving phosphate - buffered saline (PBS) and an experimental group receiving USCexo via tail vein injection. Survival rate, renal damage evaluated by HE staining, and renal function assessed by sCr and BUN levels were measured. Renal RNA was extracted to analyze apoptosis and oxidative stress. In vitro, qRT - PCR was used to assess the inflammatory response in macrophages treated with USCexo.ResultsTreatment with USC - derived exosomes led to a reduction in the levels of pathological injury, sCr, and BUN. It inhibited renal cell apoptosis and oxidative stress, decreased the infiltration of inflammatory cells, and protected renal function in SAKI mice. Additionally, USCexo suppressed the inflammatory response of primary peritoneal macrophages induced by lipopolysaccharide (LPS).ConclusionsUSC - derived exosomes protect against SAKI by inhibiting oxidative stress and inflammation, which provides a theoretical basis for the potential use of USCexo in the treatment of SAKI.Clinical trial numberNot applicable.
Eukaryotic cells migrate using pressure-driven blebs or actin polymerization driven pseudopods, with cells preferring to bleb in compressed environments where high protrusion forces are required for movement. In mammals, paxillin is a focal adhesion protein that acts as a scaffold, linking integrins to the actin cytoskeleton and recruiting signaling molecules that regulate adhesion, cytoskeletal remodeling, and migration. Dictyostelium possesses a paxillin ortholog, PaxB, which shares conserved domains with mammalian paxillin and participates in processes such as adhesion, cytokinesis, development, and chemotaxis. However, the role of PaxB in blebbing is not well understood. Our work combines experimental and theoretical methods to elucidate the role of PaxB in blebbing. We use an under-agarose assay to collect data on paxB ^- and wild-type cells under low and high compression and observe blebbing characteristics such as area and frequency. Our experimental results point to a role for PaxB in regulating the relative size of blebs in response to increased compression. Using a mathematical model of bleb expansion, we show that decreasing the assembly rate of the cortex during bleb formation leads to relatively larger blebs, thus providing a potential mechanism that replicates the mutant phenotype. Together, our experiments and theory suggest a new role of PaxB in bleb-based chemotaxis.
