Plate centrifugation enhances the efficiency of polyethylenimine-based transfection and lentiviral infection.

PURPOSE: To propose an efficient, reproducible, and consistent transgenic technology based on plate centrifugation, which is particularly useful for polyethylenimine (PEI) transfection and lentiviral infection. METHODS: We optimized multiple factors that could contribute to transfection efficiency, such as the dosage of the PEI or DNA, the working solution buffer used for diluting the PEI or DNA, the incubation time for the PEI/DNA complexes, and the transfection time. RESULTS: Plate centrifugation led to a 5.46-fold increase in the transfection efficiency of PEI-based transfection while maintaining the cell survival rate. Moreover, the average copy number of viral genes in each genome increased 4.96-fold with plate centrifugation. Plate centrifugation alters the spatial arrangement of the PEI/DNA complexes or lentiviruses, increasing the chances of these complexes or viruses coming into contact with target cells, ultimately resulting in improved transfection or infection efficiency. CONCLUSIONS: We present a protocol based on plate centrifugation for transfection or lentiviral infection that is suitable for genetic modification of primary cells or stem cells.

Impact of lipid oxidation products on the digestibility and structural integrity of Myofibrillar proteins during thermal processing.

This study explores the effects of lipid oxidation products (LOPs), specifically CHP, t,t-DDE, and MDA, on the digestibility and structural integrity of myofibrillar proteins (MP) during processing. LOPs were first assessed by heating at 180 °C for 15 min, showing a significant reduction in digestibility in MDA-treated samples (65.40 %), followed by t,t-DDE (45.10 %) and CHP (13.07 %). MALDI-TOF MS analysis revealed decreased peptide abundance and lower average molecular weight in t,t-DDE- and MDA-treated samples. Notably, substantial decreases in α-helix content and increases in random coil structures were detected, particularly in MDA-treated samples. Assessments of surface hydrophobicity and thiol content underscored the detrimental impact of secondary LOPs on MP structure. Higher MDA concentrations led to a substantial reduction in intrinsic fluorescence intensity, along with an increase in Schiff base content. A PLS regression model demonstrated strong predictive capabilities for MP digestibility, highlighting the importance of optimizing meat processing parameters to minimize nutritional degradation.

Veillonella parvula acts as a pathobiont promoting the biofilm virulence and cariogenicity of Streptococcus mutans in adult severe caries.

Adult severe caries (ASC) brings severe oral dysfunction and treatment difficulties to patients, and yet no clear pathogenic mechanism for it has been found. This study is focused on the composition of dental plaque microbiome profiles in order to identify disease-relevant species and to investigate into their interactions with the S. mutans. Samples of dental plaque were collected for metagenomic analysis. The acidification, aciduricity, oxidative stress tolerance, and gtf (glucosyltransferase) gene expression of S. mutans cocultured with V. parvula which was identified as ASC-related dominant bacterium. The biofilm formation and extracellular exopolysaccharide (EPS) synthesis of dual-strain were analyzed with scanning electron microscopy (SEM), crystal violet (CV) staining, live/dead bacterial staining, and confocal laser scanning microscopy (CLSM). Furthermore, rodent model experiments were performed to validate the in vivo cariogenicity of the dual-species biofilm. The most significantly abundant taxon found associated with ASC was V. parvula. In vitro experiments found that V. parvula can effectively promote S. mutans mature biofilm formation with enhanced acid resistance, hydrogen peroxide detoxicity, and biofilm virulence. Rodent model experiments revealed that V. parvula was incapable of causing disease on its own, but it significantly heightened the biofilm virulence of S. mutans when being co-infected and augmented the progression, quantity, and severity of dental caries. Our findings demonstrated that V. parvula may act as a synergistic pathobiont to modulate the metabolic activity, spatial structure, and pathogenicity of biofilms of S. mutans in the context of ASC.IMPORTANCEAdult severe caries (ASC), as a special type of acute caries, is rarely reported and its worthiness of further study is still in dispute. Yet studies on the etiology of severe caries in adults have not found a clear pathogenic mechanism for it. Knowledge of the oral microbiota is important for the treatment of dental caries. We discovered that the interaction between V. parvula and S. mutans augments the severity of dental caries in vivo, suggesting V. parvula may act as a synergistic pathobiont exacerbating biofilm virulence of S. mutans in ASC. Our findings may improve the understanding of ASC pathogenesis and are likely to provide a basis for planning appropriate therapeutic strategies.

Obesity mediates the relationship between depression and infertility: insights from the NHANES 2013-2018 cross-sectional study and a Mendelian randomization study.

Background: Depression is increasingly recognized as a factor affecting infertility and the causal relationship between them remains controversial. The aim of this study was to explore the relationship between depression and infertility using Mendelian randomization (MR) and cross-sectional study, and to explore the potential mediating role of obesity. Methods: The cross-sectional study used data from the National Health and Nutrition Examination Survey (NHANES) 2013-2018. Multivariable-adjusted logistic regression was used to assess the association between depression and infertility risk, and mediation analysis was to examine the mediating effect of obesity. Then, we performed MR analyses to investigate the causal effect of depression on infertility. Instrumental variables for depression were obtained from a genome-wide association meta-analysis (135,458 cases and 344,901 controls), and summary level data for infertility were obtained from the FinnGen database (6,481 cases and 68,969 controls). Results: In the cross-sectional study, a total of 2,915 participants between the ages of 18 and 45 were included, of whom 389 were infertile. We observed that depression was strongly associated with an increased risk of infertility (OR=1.66, 95%CI: 1.19, 2.33), and this relationship remained significant in mild (OR=1.45, 95% CI: 1.09, 1.93), moderate (OR=1.89, 95% CI: 1.26, 2.84), and severe depression (OR=1.74, 95% CI: 1.02, 2.99). Mediation analysis showed that obesity mediated 7.15% and 15.91% of the relationship between depression and infertility for body mass index and waist circumference. Furthermore, depression significantly increased the risk of infertility in both the general obesity (OR=1.81, 95%CI=1.20-2.73, P<0.01) and abdominal obesity populations (OR=1.57, 95%CI=1.08-2.27, P=0.02) populations. In addition, the MR analysis also revealed a significant positive causal relationship between genetically predicted depression and infertility (OR=1.32, 95% CI: 1.03, 1.70). Conclusion: Depression is associated with an increased risk of infertility, with obesity playing a significant mediating role. This study underscores the importance of incorporating mental health and weight management in infertility treatment strategies.

High-dose dexamethasone regulates microglial polarization via the GR/JAK1/STAT3 signaling pathway after traumatic brain injury.

Although microglial polarization and neuroinflammation are crucial cellular responses after traumatic brain injury, the fundamental regulatory and functional mechanisms remain insufficiently understood. As potent anti-inflammatory agents, the use of glucocorticoids in traumatic brain injury is still controversial, and their regulatory effects on microglial polarization are not yet known. In the present study, we sought to determine whether exacerbation of traumatic brain injury caused by high-dose dexamethasone is related to its regulatory effects on microglial polarization and its mechanisms of action. In vitro cultured BV2 cells and primary microglia and a controlled cortical impact mouse model were used to investigate the effects of dexamethasone on microglial polarization. Lipopolysaccharide, dexamethasone, RU486 (a glucocorticoid receptor antagonist), and ruxolitinib (a Janus kinase 1 antagonist) were administered. RNA-sequencing data obtained from a C57BL/6 mouse model of traumatic brain injury were used to identify potential targets of dexamethasone. The Morris water maze, quantitative reverse transcription-polymerase chain reaction, western blotting, immunofluorescence and confocal microscopy analysis, and TUNEL, Nissl, and Golgi staining were performed to investigate our hypothesis. High-throughput sequencing results showed that arginase 1, a marker of M2 microglia, was significantly downregulated in the dexamethasone group compared with the traumatic brain injury group at 3 days post-traumatic brain injury. Thus dexamethasone inhibited M1 and M2 microglia, with a more pronounced inhibitory effect on M2 microglia in vitro and in vivo. Glucocorticoid receptor plays an indispensable role in microglial polarization after dexamethasone treatment following traumatic brain injury. Additionally, glucocorticoid receptor activation increased the number of apoptotic cells and neuronal death, and also decreased the density of dendritic spines. A possible downstream receptor signaling mechanism is the GR/JAK1/STAT3 pathway. Overactivation of glucocorticoid receptor by high-dose dexamethasone reduced the expression of M2 microglia, which plays an anti-inflammatory role. In contrast, inhibiting the activation of glucocorticoid receptor reduced the number of apoptotic glia and neurons and decreased the loss of dendritic spines after traumatic brain injury. Dexamethasone may exert its neurotoxic effects by inhibiting M2 microglia through the GR/JAK1/STAT3 signaling pathway.

Giant viruses as reservoirs of antibiotic resistance genes.

Nucleocytoplasmic large DNA viruses (NCLDVs; also called giant viruses), constituting the phylum Nucleocytoviricota, can infect a wide range of eukaryotes and exchange genetic material with not only their hosts but also prokaryotes and phages. A few NCLDVs were reported to encode genes conferring resistance to beta­lactam, trimethoprim, or pyrimethamine, suggesting that they are potential vehicles for the transmission of antibiotic resistance genes (ARGs) in the biome. However, the incidence of ARGs across the phylum Nucleocytoviricota, their evolutionary characteristics, their dissemination potential, and their association with virulence factors remain unexplored. Here, we systematically investigated ARGs of 1416 NCLDV genomes including those of almost all currently available cultured isolates and high-quality metagenome-assembled genomes from diverse habitats across the globe. We reveal that 39.5% of them carry ARGs, which is approximately 37 times higher than that for phage genomes. A total of 12 ARG types are encoded by NCLDVs. Phylogenies of the three most abundant NCLDV-encoded ARGs hint that NCLDVs acquire ARGs from not only eukaryotes but also prokaryotes and phages. Two NCLDV-encoded trimethoprim resistance genes are demonstrated to confer trimethoprim resistance in Escherichia coli. The presence of ARGs in NCLDV genomes is significantly correlated with mobile genetic elements and virulence factors.

Micro-macro SERS strategy for highly sensitive paper cartridge with trace-level molecular detection.

Surface-enhanced Raman Scattering (SERS) has become a powerful spectroscopic technology for highly sensitive detection. However, SERS is still limited in the lab because it either requires complicated preparation or is limited to specific compounds, causing poor applicability for practical applications. Herein, a micro-macro SERS strategy, synergizing polymer-assisted printed process with paper-tip enrichment process, is proposed to fabricate highly sensitive paper cartridges for sensitive practical applications. The polymer-assisted printed process finely aggregates nanoparticles with a discrete degree of 1.77, and SERS results are matched with theoretical enhancement, indicating small cluster-dominated hotspots at the micro-scale and thus 41-fold SERS increase compared to other aggregation methods. The paper-tip enrichment process moves molecules in a fluid into small tips filled with plasmonic clusters, and molecular localization at hotspots is achieved by the simulation and optimization of fluidic velocity at the macro-scale, generating a 39.5-fold SERS sensibility increase in comparison with other flow methods. A highly sensitive paper cartridge contains a paper-tip and a 3D-printed cartridge, which is simple, easy-to-operate, and costs around 2 US dollars. With a detection limit of 10 -12 M for probe molecules, the application of real samples and multiple analytes achieves single-molecule level sensitivity and reliable repeatability with a 30-min standardized procedure. The micro-macro SERS strategy demonstrates its potential in practical applications that require point-of-care detection.

IRF3 regulates neuroinflammatory responses and the expression of genes associated with Alzheimer's disease.

The pathological role of interferon signaling is emerging in neuroinflammatory disorders, yet, the specific role of Interferon Regulatory Factor 3 (IRF3) in neuroinflammation remains poorly understood. Here, we show that global IRF3 deficiency delays TLR4-mediated signaling in microglia and attenuates the hallmark features of LPS-induced inflammation such as cytokine release, microglial reactivity, astrocyte activation, myeloid cell infiltration, and inflammasome activation. Moreover, expression of a constitutively active IRF3 (S388D/S390D: IRF3-2D) in microglia induces a transcriptional program reminiscent of the Activated Response Microglia and the expression of genes associated with Alzheimer's disease, notably apolipoprotein-e. Using bulk-RNAseq of IRF3-2D brain myeloid cells, we identified Z-DNA binding protein-1 (ZBP1) as a target of IRF3 that is relevant across various neuroinflammatory disorders. Lastly, we show IRF3 phosphorylation and IRF3-dependent ZBP1 induction in response to Aß in primary microglia cultures. Together, our results identify IRF3 as an important regulator of LPS and Aß -mediated neuroinflammatory responses and highlight IRF3 as a central regulator of disease-specific gene activation in different neuroinflammatory diseases.

In vivo Bruton's tyrosine kinase inhibition attenuates alcohol-associated liver disease by regulating CD84-mediated granulopoiesis.

Severe alcohol-associated hepatitis (AH) is a life-threatening form of alcohol-associated liver disease. Liver neutrophil infiltration is a hallmark of AH, yet the effects of alcohol on neutrophil functions remain elusive. Identifying therapeutic targets to reduce neutrophil-mediated liver damage is essential. Bruton's tyrosine kinase (BTK) plays an important role in neutrophil development and function; however, the role of BTK in AH is unknown. Using RNA sequencing of circulating neutrophils, we found an increase in Btk expression (P = 0.05) and phosphorylated BTK (pBTK) in patients with AH compared with healthy controls. In vitro, physiologically relevant doses of alcohol resulted in a rapid, TLR4-mediated induction of pBTK in neutrophils. In a preclinical model of AH, administration of a small-molecule BTK inhibitor (evobrutinib) or myeloid-specific Btk knockout decreased proinflammatory cytokines and attenuated neutrophil-mediated liver damage. We found that pBTK was essential for alcohol-induced bone marrow granulopoiesis and liver neutrophil infiltration. In vivo, BTK inhibition or myeloid-specific Btk knockout reduced granulopoiesis, circulating neutrophils, liver neutrophil infiltration, and liver damage in a mouse model of AH. Mechanistically, using liquid chromatography-tandem mass spectrometry, we identified CD84 as a kinase target of BTK, which is involved in granulopoiesis. In vitro, CD84 promoted alcohol-induced interleukin-1ß and tumor necrosis factor-α in primary human neutrophils, which was inhibited by CD84-blocking antibody treatment. Our findings define the role of BTK and CD84 in regulating neutrophil inflammation and granulopoiesis, with potential therapeutic implications in AH.

Radiomics nomogram based on CT radiomics features and clinical factors for prediction of Ki-67 expression and prognosis in clear cell renal cell carcinoma: a two-center study.

OBJECTIVES: To develop and validate a radiomics nomogram combining radiomics features and clinical factors for preoperative evaluation of Ki-67 expression status and prognostic prediction in clear cell renal cell carcinoma (ccRCC). METHODS: Two medical centers of 185 ccRCC patients were included, and each of them formed a training group (n = 130) and a validation group (n = 55). The independent predictor of Ki-67 expression status was identified by univariate and multivariate regression, and radiomics features were extracted from the preoperative CT images. The maximum relevance minimum redundancy (mRMR) and the least absolute shrinkage and selection operator algorithm (LASSO) were used to identify the radiomics features that were most relevant for high Ki-67 expression. Subsequently, clinical model, radiomics signature (RS), and radiomics nomogram were established. The performance for prediction of Ki-67 expression status was validated using area under curve (AUC), calibration curve, Delong test, decision curve analysis (DCA). Prognostic prediction was assessed by survival curve and concordance index (C-index). RESULTS: Tumour size was the only independent predictor of Ki-67 expression status. Five radiomics features were finally identified to construct the RS (AUC: training group, 0.821; validation group, 0.799). The radiomics nomogram achieved a higher AUC (training group, 0.841; validation group, 0.814) and clinical net benefit. Besides, the radiomics nomogram provided a highest C-index (training group, 0.841; validation group, 0.820) in predicting prognosis for ccRCC patients. CONCLUSIONS: The radiomics nomogram can accurately predict the Ki-67 expression status and exhibit a great capacity for prognostic prediction in patients with ccRCC and may provide value for tailoring personalized treatment strategies and facilitating comprehensive clinical monitoring for ccRCC patients.

Helicobacter pylori-Induced Angiopoietin-Like 4 Promotes Gastric Bacterial Colonization and Gastritis.

Helicobacter pylori infection is characterized as progressive processes of bacterial persistence and chronic gastritis with features of infiltration of mononuclear cells more than granulocytes in gastric mucosa. Angiopoietin-like 4 (ANGPTL4) is considered a double-edged sword in inflammation-associated diseases, but its function and clinical relevance in H. pylori-associated pathology are unknown. Here, we demonstrate both pro-colonization and pro-inflammation roles of ANGPTL4 in H. pylori infection. Increased ANGPTL4 in the infected gastric mucosa was produced from gastric epithelial cells (GECs) synergistically induced by H. pylori and IL-17A in a cagA-dependent manner. Human gastric ANGPTL4 correlated with H. pylori colonization and the severity of gastritis, and mouse ANGPTL4 from non-bone marrow-derived cells promoted bacteria colonization and inflammation. Importantly, H. pylori colonization and inflammation were attenuated in Il17a -/-, Angptl4 -/-, and Il17a -/- Angptl4 -/- mice. Mechanistically, ANGPTL4 bound to integrin αV (ITGAV) on GECs to suppress CXCL1 production by inhibiting ERK, leading to decreased gastric influx of neutrophils, thereby promoting H. pylori colonization; ANGPTL4 also bound to ITGAV on monocytes to promote CCL5 production by activating PI3K-AKT-NF-κB, resulting in increased gastric influx of regulatory CD4+ T cells (Tregs) via CCL5-CCR4-dependent migration. In turn, ANGPTL4 induced Treg proliferation by binding to ITGAV to activate PI3K-AKT-NF-κB, promoting H. pylori-associated gastritis. Overall, we propose a model in which ANGPTL4 collectively ensures H. pylori persistence and promotes gastritis. Efforts to inhibit ANGPTL4-associated pathway may prove valuable strategies in treating H. pylori infection.

Rate and Associated Factors of Fatigue in Chinese Patients with Non-Alcoholic Fatty Liver Disease: A Cross-Sectional Survey.

Purpose: Fatigue was a common symptom of non-alcoholic fatty liver disease (NAFLD), which seriously affected patients' quality of life. The aim of this study was to detect fatigue rate and to evaluate factors associated with fatigue in NAFLD patients. Patients and Methods: A cross-sectional study was carried out from the Huadong Sanatorium between April 2022 and May 2023, and 133 NAFLD patients were included in this study. They completed Fatigue Severity Scale to assess fatigue, the Hospital Anxiety and Depression Scale to estimate psychological status, and the Pittsburgh Sleep Quality Index for sleep quality. Data were analyzed by independent samples t-tests, χ2 tests and logistic regression models. Results: We found that 51.1% of NAFLD patients had fatigue. Exercise, anxiety, depression, subjective sleep quality, sleep latency, sleep duration, habitual sleep efficiency, sleep disorders, daytime dysfunction and overall sleep quality were related to fatigue among NAFLD patients. Moreover, logistic regression models indicated anxiety, habitual sleep efficiency and sleep disorders as important predictors of fatigue. Conclusion: This was the first time to explore demographic, clinical, psychological and sleeping correlated factors for fatigue in Chinese NAFLD patients. Our study showed that more than half of NAFLD patients had fatigue, and anxiety, habitual sleep efficiency and sleep disorders were significantly associated with fatigue in NAFLD. The findings indicated that it was very necessary to pay more attention to fatigue of NAFLD patients, especially those with negative emotions and poor sleep quality by favorable intervention to relieve fatigue symptoms, so as to improve quality of life.

Assembly of ligation chain reaction and DNA triangular prism for miRNA diagnostics.

Controllable assembly of DNA nanostructure provides a powerful way for quantitative analysis of various targets including nucleic acid molecules. In this study, we have designed detachable DNA nanostructures at electrochemical sensing interface and constructed a ligation chain reaction (LCR) strategy for amplified detection of miRNA. A three-dimensional DNA triangular prism nanostructure is fabricated to provide suitable molecule recognition environment, which can be further regenerated for additional tests via convenient pH adjustment. Target triggered LCR is highly efficient and specific towards target miRNA. Under optimal experimental conditions, this approach enables ultrasensitive exploration in a wide linear range with a single-base resolution. Moreover, it shows excellent performances for the analysis of cell samples and clinical serum samples.

A Novel Ectodysplasin a Gene mutation of X-Linked Hypohidrotic Ectodermal Dysplasia.

Introduction: Hypohidrotic ectodermal dysplasia (HED) is a genetic disorder that influences structures of ectodermal origin, such as teeth, hair, and sweat glands. Compared with autosomal recessive and dominant modes of inheritance, the X-linked HED (XLHED) characterized by Hypodontia/Oligodontia teeth, Absent/sparse hair, Anhidrosis/hypohidrosis, and characteristic facial features, is the most frequent and its primary cause is the mutation of ectodysplasin A (EDA) gene. This research aimed to expound the clinical and molecular features of a Chinese male with XLHED and to summarize and compare several previous findings. Methods: Genomic DNA was obtained from the peripheral blood of the proband and his family members, then Sanger sequencing was used to perform a mutational analysis of EDA. Real-time quantitative PCR and Western blotting were used to detect EDA expression. The transcriptional activity of NF-κB was detected using a luciferase assay. Results: The probandwith XLHED was identified a novel EDA mutation, c.1119G>C(p.M373I), that affected the molecular analysis of transmembrane protein exon8 mutations, inherited from the mother. He showed a severe multiple-tooth loss, with over 20 permanent teeth missing and sparse hair and eyebrows, dry, thin, and itching skin. Furthermore, his sweating function was abnormal to a certain extent. Discussion: The functional study showed that this novel mutant led to a significant decrease in the EDA expression level and transcriptional activity of NF-κB. Our findings extend the range of EDA mutations in XLHED patients, which provides the basis and idea for further exploring the pathogenesis of XLHED.

E proteins control the development of NKγδT cells through their invariant T cell receptor.

T cell receptor (TCR) signaling regulates important developmental transitions, partly through induction of the E protein antagonist, Id3. Although normal γδ T cell development depends on Id3, Id3 deficiency produces different phenotypes in distinct γδ T cell subsets. Here, we show that Id3 deficiency impairs development of the Vγ3+ subset, while markedly enhancing development of NKγδT cells expressing the invariant Vγ1Vδ6.3 TCR. These effects result from Id3 regulating both the generation of the Vγ1Vδ6.3 TCR and its capacity to support development. Indeed, the Trav15 segment, which encodes the Vδ6.3 TCR subunit, is directly bound by E proteins that control its expression. Once expressed, the Vγ1Vδ6.3 TCR specifies the innate-like NKγδT cell fate, even in progenitors beyond the normally permissive perinatal window, and this is enhanced by Id3-deficiency. These data indicate that the paradoxical behavior of NKγδT cells in Id3-deficient mice is determined by its stereotypic Vγ1Vδ6.3 TCR complex.

Measurement of Zinc Ions in Seawater Samples Using a Microfluidic System Based on the GR/CeO2/Nafion Material.

Considering that heavy-metal contamination of seawater is getting worse, building a quick, accurate and portable device for detecting trace zinc in seawater in real time would be very beneficial. In this work, a microfluidic system was developed that includes a planar disc electrode, a micro-cavity for detection, an electrochemical workstation, a computer, a container for waste liquid reprocessing, an external pipeline and other components as well as a graphene/cerium oxide/nano-cerium oxide/Nafion composite membrane was used to modify the planar disc electrode (GR/CeO2/Nafion/Au) to investigate the electrochemical behaviour of Zn(II) using cyclic voltammetry, square-wave voltammetry and orthogonal test methods. Under optimal experimental conditions, the peak reaction current of Zn(II) showed a good linear relationship with the concentration of Zn(II) in the range of 1-900 µg/L with a correlation coefficient of 0.998, and the detection limit of the method was 0.87 µg/L. In addition, the microfluidic system had good stability, reproducibility and anti-interference. The system was used for determining zinc ions in real seawater samples, and the results were very similar to those of inductively coupled plasma-emission spectrometry, demonstrating the practicality of the system for the detection of trace zinc.

SMP30 Interacts with ROCK1 to Inhibit HCC Invasiveness by Effecting the Epithelial-Mesenchymal Transition.

BACKGROUND: The senescence marker protein 30 (SMP30) is a calcium-binding protein whose expression decreases with age, and is closely associated with hepatocellular carcinoma (HCC) development. The primary goal of this study was to examine the mechanistic effect of SMP30 on HCC migration and invasion. METHODS: Bioinformatic and immunohistochemical approaches were used to examine the expression of SMP30 in HCC tissues and its relationship to patient survival. We investigated the effects of SMP30 expression on HCC cell proliferation, migration, invasion, and cell cycle dynamics. cDNA microarray technology was used to determine the gene expression profile of SK-Hep-1 cells following recombinant SMP30 overexpression to identify genes downstream of SMP30 that regulate HCC cell migration and invasion. We identified SMP30 interacting proteins by affinity purification-mass spectrometry (AP-MS) and co-immunoprecipitation/western blotting (COIP-WB). RESULTS: SMP30 expression was lower in HCC tissues compared with normal liver tissues, and its expression positively correlated with overall survival in HCC patients. Additionally, SMP30 overexpression effectively blocked the migratory and invasive properties of SK-Hep-1 cells, but did not affect either proliferation rates or cell cycle. cDNA microarray results confirmed that many of the differentially expressed genes identified are involved in the process of epithelial-mesenchymal transition (EMT). AP-MS and COIP-WB experiments confirmed that Rho-associated protein kinase 1 (ROCK1) interacts with SMP30 in SK-Hep-1 cells, and ROCK1 is known to intimately regulate the EMT process. CONCLUSION: SMP30 inhibits HCC metastasis by influencing the expression of EMT-related proteins after interacting with ROCK1.

Combined Insults of a MASH Diet and Alcohol Binges Activate Intercellular Communication and Neutrophil Recruitment via the NLRP3-IL-1ß Axis in the Liver.

Binge drinking in obese patients positively correlates with accelerated liver damage and liver-related death. However, the underlying mechanism and the effect of alcohol use on the progression of metabolic-dysfunction-associated steatotic liver disease (MASLD) remain unexplored. Here, we show that short-term feeding of a metabolic-dysfunction-associated steatohepatitis (MASH) diet plus daily acute alcohol binges for three days induce liver injury and activation of the NLRP3 inflammasome. We identify that a MASH diet plus acute alcohol binges promote liver inflammation via increased infiltration of monocyte-derived macrophages, neutrophil recruitment, and NET release in the liver. Our results suggest that both monocyte-derived macrophages and neutrophils are activated via NLRP3, while the administration of MCC950, an NLRP3 inhibitor, dampens these effects.In this study, we reveal important intercellular communication between hepatocytes and neutrophils. We discover that the MASH diet plus alcohol induces IL-1ß via NLRP3 activation and that IL-1ß acts on hepatocytes and promotes the production of CXCL1 and LCN2. In turn, the increase in these neutrophils recruits chemokines and causes further infiltration and activation of neutrophils in the liver. In vivo administration of the NLRP3 inhibitor, MCC950, improves the early phase of MetALD by preventing liver damage, steatosis, inflammation, and immune cells recruitment.

Exploring quantitative indices to characterize piano timbre with precision validated using measurement system analysis.

Aim: Timbre in piano performance plays a critical role in enhancing musical expression. However, timbre control in current piano performance education relies mostly on descriptive characterization, which involves large variations of interpretation. The current study aimed to mitigate the limitations by identifying quantitative indices with adequate precision to characterize piano timbre. Methods: A total of 24 sounds of G6 were recorded from 3 grand pianos, by 2 performers, and with 4 repetitions. The sounds were processed and analyzed with audio software for the frequencies and volumes of harmonic series in the spectrum curves. Ten quantitative timbre indices were calculated. Precision validation with statistical gage R&R analysis was conducted to gage the repeatability (between repetitions) and reproducibility (between performers) of the indices. The resultant percentage study variation (%SV) of an index must be ≤10% to be considered acceptable for characterizing piano timbre with enough precision. Results: Out of the 10 indices, 4 indices had acceptable precision in characterizing piano timbre with %SV ≤10%, including the square sum of relative volume (4.40%), the frequency-weighted arithmetic mean of relative volume (4.29%), the sum of relative volume (3.11%), and the frequency-weighted sum of relative volume (2.09%). The novel indices identified in the current research will provide valuable tools to advance the measurement and communication of timbre and advance music performance education.