Substantial evidence now exists to support that formation of DNA G-quadruplexes (G4s) is coupled to altered gene expression. However, approaches that allow us to probe G4s in living cells without ...perturbing their folding dynamics are required to understand their biological roles in greater detail. Herein, we report a G4-specific fluorescent probe (SiR-PyPDS) that enables single-molecule and real-time detection of individual G4 structures in living cells. Live-cell single-molecule fluorescence imaging of G4s was carried out under conditions that use low concentrations of SiR-PyPDS (20 nM) to provide informative measurements representative of the population of G4s in living cells, without globally perturbing G4 formation and dynamics. Single-molecule fluorescence imaging and time-dependent chemical trapping of unfolded G4s in living cells reveal that G4s fluctuate between folded and unfolded states. We also demonstrate that G4 formation in live cells is cell-cycle-dependent and disrupted by chemical inhibition of transcription and replication. Our observations provide robust evidence in support of dynamic G4 formation in living cells.
Four-stranded G-quadruplexes (G4s) are DNA secondary structures in the human genome that are primarily found in active promoters associated with elevated transcription. Here, we explore the ...relationship between the folding of promoter G4s, transcription and chromatin state.
Transcriptional inhibition by DRB or by triptolide reveals that promoter G4 formation, as assessed by G4 ChIP-seq, does not depend on transcriptional activity. We then show that chromatin compaction can lead to loss of promoter G4s and is accompanied by a corresponding loss of RNA polymerase II (Pol II), thus establishing a link between G4 formation and chromatin accessibility. Furthermore, pre-treatment of cells with a G4-stabilising ligand mitigates the loss of Pol II at promoters induced by chromatin compaction.
Overall, our findings show that G4 folding is coupled to the establishment of accessible chromatin and does not require active transcription.
•The superiority of plasma N antigen in early identification of severe COVID-19 patients surpasses that of nasopharyngeal nucleic acid CT values and established COVID-19 blood biomarkers.•Patients ...with elevated concentrations of plasma N antigen during the initial phase are more susceptible to developing severe COVID-19.•The ongoing surveillance of plasma N antigen concentrations may serve as a means to monitor the progression of the disease.
Clear and effective indicators for early detection of severe coronavirus disease 2019 (COVID-19) are insufficient. We investigated the clinical value of the plasma SARS-CoV-2 N antigen (plasma N antigen) for severe COVID-19 early identification and disease progression monitoring.
A cross-sectional study compared the diagnostic value of plasma N antigen levels detected within two days after hospital admission in 957 patients with COVID-19 during the BA2.2 outbreak in Shanghai (April 6–June 15, 2022). A follow-up study analyzed the plasma N antigen prognostic value in 274 non-severe patients, and a longitudinal study evaluated its continuous monitoring value in 16 patients with COVID-19 grade changes.
Plasma N antigen concentrations were significantly higher in severely ill than in non-severely ill patients. The plasma N antigen was superior to nasopharyngeal nucleic acid CT values and established COVID-19 blood biomarkers in identifying severe COVID-19. Patients with high plasma N-antigen concentrations at initial admission were more prone to developing severe COVID-19. The changes in plasma N antigen concentrations were consistent with disease progression. Two logistic regression models, including and excluding plasma N antigen, were established, with model 1 (including plasma N antigen) (AUC = 0.971, 0.958–0.980) yielding a better diagnostic value for severe COVID-19 than Model 2 (plasma N antigen excluded).
The plasma N antigen is superior to nasopharyngeal nucleic acids and established COVID-19 blood biomarkers for severe COVID-19 early recognition and progression monitoring, enabling the most accurate patient triaging and efficient utilization of medical resources.
Various comorbidities associated with COVID-19 add up in severity of the disease and obviously prolonged the time for viral clearance. This study investigated a novel ultrasensitive MAGLUMI
...SARS-CoV-2 Ag chemiluminescent immunoassay assay (MAG-CLIA) for diagnosis and monitoring the infectivity of COVID-19 patients with comorbid conditions during the pandemic of 2022 Shanghai.
Analytical performances of the MAG-CLIA were evaluated, including precision, limit of quantitation, linearity and specificity. Nasopharyngeal specimens from 232 hospitalized patients who were SARS-CoV-2 RT-qPCR positive and from 477 healthy donors were included. The longitudinal studies were performed by monitoring antigen concentrations alongside with RT-qPCR results in 14 COVID-19 comorbid participants for up to 22 days. The critical antigen concentration in determining virus infectivity was evaluated at the reference cycle threshold (Ct) of 35.
COVID-19 patients were well-identified using an optimal threshold of 0.64 ng/L antigen concentration, with sensitivity and specificity of 95.7% (95% CI: 92.2-97.9%) and 98.3% (95% CI: 96.7-99.3%), respectively, while the Wondfo LFT exhibited those of 34.9% (95% CI: 28.8-41.4%) and 100% (95% CI: 99.23-100%), respectively. The sensitivity of MAG-CLIA remained 91.46% (95% CI: 83.14-95.8%) for the samples with Ct values between 35 and 40. Close dynamic consistence was observed between MAG-CLIA and viral load time series in the longitudinal studies. The critical value of 8.82 ng/L antigen showed adequate sensitivity and specificity in evaluating the infectivity of hospitalized convalescent patients with comorbidities.
The MAG-CLIA SARS-CoV-2 Ag detection is an effective and alternative approach for rapid diagnosis and enables us to evaluate the infectivity of hospitalized convalescent patients with comorbidities.
Glioblastoma multiforme (GBM) is the most common form of malignant glioma, characterized by genetic instability and unpredictable clinical behavior. GBM is marked by an extremely poor prognosis with ...median overall survival of 12~14 months. In this study, we detected the CD137L-expressing cells and IL-17-expressing cells in tumor tissues resected from patients with GBM. Expression of CD137L and IL-17 were assessed by immunohistochemistry, and the prognostic value of CD137L and IL-17 expression within the tumor tissues were assessed by Cox regression and Kaplan-Meier analysis. Immunohistochemical detection showed that positive cells of CD137L and IL-17 in glioblastoma tissue samples were 46.3% (19/ 41) and 73.2% (30/41) respectively. Expression of CD137L was not correlated with overall survival of GBM patients (P=0.594), while significantly longer survival rate was seen in patients with high expression of IL-17, compared to those with low expression of IL-17 (P=0.007). In addition, we also found that IL-17 expression was significantly correlated with Progression-free survival (PFS) (P=0.016) and death rate (P=0.01). Furthermore, multivariate Cox proportional hazard analyses revealed that IL-17 (P=0.018) and PFS (P=0.028) were independent factors affecting the overall survival probability. Kaplan-Meier analysis showed that PFS of high expression of IL-17 group were significantly longer (P=0.004) than low expression group with GBM. It is concluded that high levels of IL-17 expression in the tumor tissues may be a good prognostic marker for patients with GBM.
The diagnose instrument of Hess screen is an important instrument for ophthalmic diagnoses. It is mainly used for the measurement of paralytic strabismus. This article is about a digital diagnosis ...instrument of Hess screen for paralytic strabismus, abstract truncated by publisher.
Objective
Unidentified mechanisms largely restrict the viability of effective therapies in pharmacoresistant epilepsy. Our previous study revealed that hyperactivity of the subiculum is crucial for ...the genesis of pharmacoresistance in temporal lobe epilepsy (TLE), but the underlying molecular mechanism is not clear.
Methods
Here, we examined the role of subicular caspase‐1, a key neural pro‐inflammatory enzyme, in pharmacoresistant TLE.
Results
We found that the expression of activated caspase‐1 in the subiculum, but not the CA1, was upregulated in pharmacoresistant amygdaloid‐kindled rats. Early overexpression of caspase‐1 in the subiculum was sufficient to induce pharmacoresistant TLE in rats, whereas genetic ablation of caspase‐1 interfered with the genesis of pharmacoresistant TLE in both kindled rats and kainic acid‐treated mice. The pro‐pharmacoresistance effect of subicular caspase‐1 was mediated by its downstream inflammasome‐dependent interleukin‐1β. Further electrophysiological results showed that inhibiting caspase‐1 decreased the excitability of subicular pyramidal neurons through influencing the excitation/inhibition balance of presynaptic input. Importantly, a small molecular caspase‐1 inhibitor CZL80 attenuated seizures in pharmacoresistant TLE models, and decreased the neuronal excitability in the brain slices obtained from patients with pharmacoresistant TLE.
Interpretation
These results support the subicular caspase‐1‐interleukin‐1β inflammatory pathway as a novel alternative mechanism hypothesis for pharmacoresistant TLE, and present caspase‐1 as a potential target. ANN NEUROL 2021;90:377–390
Objective. Hyperuricemia (HUA) is a common metabolic disease caused by disordered purine metabolism. We aim to reveal the mechanisms underlying the anti-HUA function of Simiao pill and provide ...therapeutic targets. Methods. Simiao pill-related targets were obtained using Herbal Ingredients’ Targets (HIT), Traditional Chinese Medicine Systems Pharmacology (TCMSP), and Traditional Chinese Medicine Integrated Database (TCMID). HUA-associated targets were retrieved from GeneCards, DisGeNET, and Therapeutic Targets Database (TTD). Protein-protein interaction (PPI) network was constructed using the Search Tool for the Retrieval of Interacting Genes/Proteins (STRING) database, ggraph and igraph R packages. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were performed using ClusterProfiler. The top 10 core targets were identified through cytoHubba. Molecular docking was conducted using PyMOL and AutoDock high-performance liquid chromatograph (HPLC) analysis was performed to identify effective compounds of Simiao pill. Results. Simiao pill-HUA target network contained 80 targets. The key targets were mainly involved in inflammatory responses. Insulin (INS), tumor necrosis factor (TNF), interleukin-6 (IL6), interleukin 1 beta (IL1B), vascular endothelial growth factor A (VEGFA), leptin (LEP), signal transducer and activator of transcription 3 (STAT3), C-C motif chemokine ligand 2 (CCL2), interleukin-10 (IL10), and toll like receptor 4 (TLR4) were the top 10 targets in the PPI network. GO analysis demonstrated the main implication of the targets in molecular responses, production, and metabolism. KEGG analysis revealed that Simiao pill might mitigate HUA through advanced glycation end-product- (AGE-) receptor for AGE- (RAGE-) and hypoxia-inducible factor-1- (HIF-1-) associated pathways. IL1B, IL6, IL10, TLR4, and TNF were finally determined as the promising targets of Simiao pill treating HUA. Through molecular docking and HPLC analysis, luteolin, quercetin, rutaecarpine, baicalin, and atractylenolide I were the main active compounds. Conclusions. Simiao pill can mitigate HUA by restraining inflammation, mediating AGE-RAGE- and HIF-1-related pathways, and targeting IL1B, IL6, IL10, TLR4, and TNF.
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