A cell-free system toward deciphering the post-translational modification barcodes of Oct4 in different cellular contexts

AbstractThe octamer-binding transcription factor 4 (Oct4) is essential for maintaining the self-renewal and pluripotency of embryonic stem cells (ESCs). Post-translational modifications (PTMs) of Oct4 critically control its structure, function and intracellular localization. However, determination of Oct4 PTM profiles has largely been restricted by the quantity and purity of the Oct4 protein samples required for mass spectrometric analyses. In this study, by incubating the Escherichia coli-derived His-tagged Oct4 proteins with the whole cell lysates of a variety of human cells followed by retrieving the reacted Oct4 proteins with the Ni–NTA beads, we developed a labor- and cost-effective in vitro PTM method that allowed for mass spectrometric determination of the phosphorylation profiles of Oct4 proteins exposed to various cell-free systems. A number of Oct4 phosphorylation sites that were commonly present in all the cell-free systems or specifically present in a particular cellular context were identified, indicating that Oct4 is controlled by both common and distinct PTM regulatory pathways. Our work provided a proof-of-concept that such a cell-free system-based in vitro PTM approach can be applied to systematically map out the physiologically-relevant PTM sites in Oct4 proteins, which opened up an avenue to fully decipher the Oct4 PTM barcodes in various cellular contexts

Genome mining reveals the prevalence and extensive diversity of toxin–antitoxin systems in Staphylococcus aureus

IntroductionStaphylococcus aureus (S. aureus) is a highly pathogenic and adaptable Gram-positive bacterium that exhibits persistence in various environments. The toxin-antitoxin (TA) system plays a crucial role in the defense mechanism of bacterial pathogens, allowing them to survive in stressful conditions. While TA systems in clinical pathogens have been extensively studied, there is limited knowledge regarding the diversity and evolutionary complexities of TA systems in S. aureus.MethodsWe conducted a comprehensive in silico survey using 621 publicly available S. aureus isolates. We employed bioinformatic search and prediction tools, including SLING, TADB2.0, and TASmania, to identify TA systems within the genomes of S. aureus.ResultsOur analysis revealed a median of seven TA systems per genome, with three type II TA groups (HD, HD_3, and YoeB) being present in over 80% of the strains. Additionally, we observed that TA genes were predominantly encoded in the chromosomal DNA, with some TA systems also found within the Staphylococcal Cassette Chromosomal mec (SCCmec) genomic islands.DiscussionThis study provides a comprehensive overview of the diversity and prevalence of TA systems in S. aureus. The findings enhance our understanding of these putative TA genes and their potential implications in S. aureus ecology and disease management. Moreover, this knowledge could guide the development of novel antimicrobial strategies

Alpha2B-Adrenergic Receptor Regulates Neutrophil Recruitment in MSU-Induced Peritoneal Inflammation

Gout is one of the most common metabolic disorders in human. Previous studies have shown that the disease activity is closely associated with sympathetic nervous system (SNS). α2B-adrenergic receptor (α2BAR), a subtype of α2 adrenergic receptor, plays a critical role in many diseases. However, the role of α2BAR in the pathogenesis of gout remains unclear. Here, we assessed the role of α2BAR in the monosodium urate (MSU) crystals-induced peritonitis that mimics human gout by using the α2BAR-overexpressing mice (α2BAR-Tg). We found that the number of recruited neutrophils was significantly increased in the α2BAR-Tg mice after MSU treatment, when compared with wild type mice. In contrast, the number of macrophages was not changed. Importantly, there is no difference in the IL-1β levels and caspase-1 activity between wild type and α2BAR-Tg mice in the gout animal model. Notably, the enhanced neutrophil migration in α2BAR-Tg mice was dependent on the α2BAR overexpression in neutrophils, but not resulted from other tissues or cells with α2BAR overexpression. In conclusion, our data provide a direct evidence that α2BAR plays a critical role in neutrophil migration and MSU-induced inflammation

TNFRSF10C methylation is a new epigenetic biomarker for colorectal cancer

Background Abnormal methylation of TNFRSF10C was found to be associated with different types of cancers, excluding colorectal cancer (CRC). In this paper, the performance of TNFRSF10C methylation in CRC was studied in two stages. Method The discovery stage was involved with 38 pairs of CRC tumor and paired adjacent non-tumor tissues, and 69 pairs of CRC tumor and paired adjacent non-tumor tissues were used for the validation stage. Quantitative methylation specific PCR (qMSP) method and percentage of methylated reference (PMR) were used to test and represent the methylation level of TNFRSF10C, respectively. A dual-luciferase reporter gene experiment was conducted to evaluate the promoter activity of TNFRSF10C fragment. Results A significant association of TNFRSF10C promoter hypermethylation with CRC was found and validated (discovery stage: 24.67 ± 7.52 vs. 3.36 ± 0.89; P = 0.003; validation stage: 31.21 ± 12.48 vs. 4.52 ± 1.47; P = 0.0005). Subsequent analyses of TCGA data among 46 pairs of CRC samples further confirmed our findings (cg23965061: P = 4E − 6; cg14015044: P = 1E − 7). Dual-luciferase reporter gene assay revealed that TNFRSF10C fragment was able to significantly promote gene expression (Fold change = 2.375, P = 0.013). Our data confirmed that TNFRSF10C promoter hypermethylation can predict shorter overall survival of CRC patients (P = 0.032). Additionally, bioinformatics analyses indicated that TNFRSF10C hypermethylation was significantly associated with lower TNFRSF10C expression. Conclusion Our work suggested that TNFRSF10C hypermethylation was significantly associated with the risk of CRC

Potential Blood Pressure Goals in IgA Nephropathy: Prevalence, Awareness, and Treatment Rates in Chronic Kidney Disease Among Patients with Hypertension in China (PATRIOTIC) Study

Background/Aims: IgA nephropathy is the most prevalent form of primary glomerulonephritis worldwide. Among patients with kidney disease, hypertension is one of the most important risk factors of disease progression. Considering the limited evidence regarding the appropriate blood pressure (BP) goal for patients with IgA nephropathy, our aim was to critically appraise the potential BP goal in IgA nephropathy. Methods: We performed a retrospective analysis of the BP data from 1055 patients with IgA nephropathy, extracted from the database of a nationwide, multi-center, cross-sectional study, including 61 tertiary hospitals in China. Hypertension was defined by a BP ≥140/90 mmHg. Three BP cutoff levels were evaluated as control values: < 140/90 mmHg, < 130/80 mmHg and < 125/75 mmHg. The primary outcome of our study was the prevalence of BP control among patients with a 24-h proteinuria < 1 g/d or ≥ 1 g/d. Multivariate logistic regression analysis was used to identify demographic and clinical factors associated with a decrease in renal function for the different target levels of BP. Results: The overall prevalence of hypertension was 63.3%. BP was controlled under 140/90 mmHg in 49.1% of patients, with 34.3% of patients with proteinuria < 1 g/d reaching the target BP < 130/80 mmHg and only 12.9% of patients with proteinuria > 1 g/d achieving a BP < 125/75 mmHg. Among patients with proteinuria < 1 g/d, the adjusted odds ratios (OR) and 95% confidence interval (95% CI) of a decrease in renal function, for the 3 target BP levels, were as follows (P > 0.05): < 140/90 mmHg, 0.9 (0.5 - 1.6); < 130/80 mmHg, 1.0 (0.5 - 1.8); and < 125/75 mmHg, 1.0 (0.5 - 2.0). With proteinuria ≥1 g/d, the adjusted ORs (95%CI) of attaining the BP targets of < 140/90 mmHg, < 130/80 mmHg and < 125/75 mmHg were 0.4 (0.2 - 0.6), 0.2 (0.1 - 0.4) and 0.3 (0.1 - 0.5), respectively (P < 0.05). Conclusion: Hypertension was common in IgA nephropathy and hypertensive control was suboptimal. Our result supports a benefit of intensive control of BP < 130/80 mmHg for patients with proteinuria ≥1 g/d. However, in patients with proteinuria < 1 g/d, a renoprotective effect of this BP goal was not identified

A STUDY ON THE INHIBITORY EFFECT OF RADIX SEMIAQUILEGIAE EXTRACT ON HUMAN HEPATOMA HEPG-2 AND SMMC-7721 CELLS

The main objective of this paper was to investigate the extraction process of ethanol extract of Radix Semiaquilegiae, as well as its inhibitory activity on human hepatoma HepG-2 and SMMC-7721 cells, and to compare the inhibitory effects of different concentrations of ethanol extracts against these two hepatoma cells. Ethanol reflux extraction and ultrasound-assisted extraction with ethanol at room temperature were used in the extraction process, and MTT assay was mainly used in the activity experiment to perform in-vitro anti HepG-2 and SMMC-7721 cell activity screening of ethanol extract, and to calculate the cell inhibition rates of the extracts. The results showed that among the two types of extracts, ethanol reflux extract had more superior antitumour activity to that of the ultrasonic extract, but all of the extracts obtained had certain anti-cancer activities, and the anti-proliferative activity increased with the increase of concentration