Poor comparability of plasma renin activity measurement in determining patient samples: the status quo and recommendations for harmonization.

OBJECTIVES: This study aims to investigate and update the consistency and comparability of plasma renin activity (PRA) assays in measuring clinical samples. The contributions of recalibration, blank subtraction, and incubation strategies to interchangeability were also explored. METHODS: Five different laboratories were evaluated using forty-six individual plasma samples, including four liquid chromatography-tandem mass spectrometry (LC‒MS/MS) assays and one chemiluminescence immunoassay (CLIA). Spearman correlation coefficient (R), Passing-Bablok regression, and Bland‒Altman plot analyses were used to evaluate the consistency among assays. Consistency before and after recalibration, blank subtraction, and incubation strategy unification was compared. RESULTS: A good correlation was observed among all assays (R>0.93). None of the samples measured by all assays showed coefficient variation (CV) <10 %, and 37 % of samples showed overall CVs >20 %. The 95 % confidence intervals (CIs) for slopes did not contain 1 for most assay pairs. Large relative biases (-85.1-104.2 %) were found, and 76 % (52-93 %) of samples had unacceptable biases. Recalibration reduced the calibration bias. Ignoring blank subtraction improved the comparability across all assays while unifying incubation did not. CONCLUSIONS: The interchangeability of PRA measurement was unsatisfying. Harmonization on calibrator and ignoring blank were recommended. Unifying incubation strategy was unnecessary.

A simple method for rapid screening and diagnosis of common organic acidemias: quantitative detection of serum and urine organic acid profiles based on liquid chromatography-tandem mass spectrometry.

Organic acid (OA) analysis is a specific test for inherited metabolic disorders (IMDs); however, the previous detection methods are laborious and costly. This study aims to develop a rapid method for the simultaneous quantification of serum and urine OA profiles. The method was established based on the liquid chromatography-tandem mass spectrometry (LC-MS/MS) technique. The specificity, sensitivity, robustness, and accuracy of the established method were validated. Fifteen healthy subjects and nine IMD patients were measured for clinical validation. OAs with their intrinsic isomers were completely separated. The LC-MS/MS analysis time was 5.5 min. Calibration curves were linear within the ranges of 27.00 µg/g for all OAs. The average correlation relationship (R) varied from 0.9891 to 0.9998. The limit of detection and limit of quantification varied from 0.003 to 0.07 µg/g and 0.006 to 0.08 µg/g, respectively. No obvious carryover was observed. The intra-assay, inter-assay, and total imprecisions were 1.22-4.14%, 0.90-5.20%, and 1.67-5.90%, respectively. The mean spiked recovery at the three levels varied from 94.31 to 106.68%. The matrix effects can be compensated for by internal standard correction. Nine IMD patients were identified. A robust LC-MS/MS method for the rapid determination of serum and urine OA profiles without derivatization or liquid-liquid extraction was developed and validated. The analysis of five common OAs can be completed in short minutes. This innovative LC-MS/MS method for OA profiles may present its potential in future rapid screening and diagnosis of IMDs.

Therapeutic drug monitoring status of four common antibiotics: vancomycin, meropenem, linezolid and teicoplanin.

Accurate therapeutic drug monitoring (TDM) of vancomycin, meropenem, linezolid and teicoplanin are conducive to developing optimal therapeutic regimes for patients. However, the measurement status of those drugs in different laboratories has not been reported. In this study, four samples including two frozen plasma samples and two lyophilized plasma samples were measured by over 35 laboratories across China. The inter- and intra-laboratory %CV, biases (%) of laboratories and intra- and inter-measurement-system %CV were calculated and analyzed. The short-term stability and homogeneity of those drugs in samples were studied. The results of frozen and lyophilized samples were also compared to determine whether there were significant differences in their matrix effects on various measurement systems. Results showed most laboratories' intra-laboratory %CVs were less than 9% for all drugs, and the mean inter-laboratory %CVs were 18.4%, 86.4%, 19.1% and 37.1% for vancomycin, meropenem, linezolid and teicoplanin measurements, respectively. For vancomycin, the intra-measurement %CV of commercial measurement systems was found to be smaller than that of other measurement systems. For meropenem, linezolid and teicoplanin, the agreement among laboratories using self-developed methods (Liquid chromatography-mass spectrometry [LC-MS] or high-performance liquid chromatography [HPLC]) was not satisfactory as most intra-measurement system CVs% were over 20%. Drugs in lyophilized samples were found to be more stable than in frozen samples, and no obvious differences in matrix effects were found for those two kinds of processed samples on most measurement systems. In conclusion, this study depicted the measurement status of those drugs in clinical laboratories, and found the lyophilized samples were more suitable EQA material for those drugs.

The consistency of aminotransferase analysis in China: a comparison of six mainstream aminotransferase routine methods and recalibration using human pooled serum preparations supplemented with human recombinant aminotransferases.

Background: To evaluate the consistency of six mainstream homogeneous systems for aminotransferase measurements and improve the consistency of measurements by applying uniform calibrators.Methods: 200 individual samples were grouped into four sets for assays with and without pyridoxal-5-phosphate (P-5'-P) for Alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Routine assays for the P-5'-P group were compared with a reference measurement procedure (RMP). In the non-P-5'-P group, four routine assays were analyzed in a pairwise method across six method pairs. Recalibration was performed using human serum pools (HSPs) supplemented with human original recombinant aminotransferases (HOR). Data were analyzed by Passing-Bablok regression and Bland-Altman plots.Results: In the P-5'-P group, the mean biases for Ortho and Dimension assays for ALT were 17.0% and -25.4%, respectively; for AST, the mean biases were -9.5% and -9.6%, respectively. In the non-P-5'-P group, the mean deviations ranged from -5.9% to 5.9% for ALT. For AST, the relative deviations ranged from -19.1% to 6.5%. After recalibration, in the P-5'-P group, the relative biases were reduced to -12.2% to 7.7% for ALT and -6.9% to 0.8% for AST. The mean deviations for the non-P-5'-P AST group were reduced remarkably (-3.0% to 3.3%).Conclusion: Assays supplemented with P-5'-P exhibited poor performance against RMP for both ALT and AST. For assays without P-5'-P, AST results showed non-satisfactory comparability for almost all method pairs. Uniform calibrators such as HSPs supplemented with HOR could improve consistency among the mainstream homogeneous systems for the measurement of aminotransferase activity, particularly for AST measurement.

The spectrum of plasma renin activity and hypertension diseases: Utility, outlook, and suggestions.

BACKGROUND: Plasma renin activity (PRA) is one of the recommended screening indicators for primary aldosteronism (PA) diagnosis and had become increasingly important in hypertension identification, medication guidance, and endocrine disorder confirmation. METHODS: To provide an overview of the PRA measurement progress and clinical value, this review summarizes the main contributing factors related to PRA measurement and necessary precautions during the entire analysis process. We also outline the characteristics of PRA in different endocrine diseases and their clinical utility. RESULTS: Significant inconsistency was observed in PRA measurement methods, including immunoassay and isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC/MS/MS), which could be attributed to preanalytical, analytical, and postanalytical variations. Meanwhile, consensus about environmental and procedural factors during the entire analytical process, including storage temperature, incubation condition, blank subtraction, and standardized operational procedures across different self-developed assay laboratories, could be important to minimize analytical variations. Furthermore, commutable uniform calibrators should be prepared to improve consistency, ultimately achieving accurate and reliable measurement of PRA. CONCLUSION: This review summarizes the clinical utilization of PRA as a biomarker in multiple diseases, elaborating on routine detection methods and the key factors in the analytical process. We also provide feasible strategies for improving standardization and facilitating PRA assessment for larger-scale clinical applications.

Construction of a CRISPR-Biolayer Interferometry Platform for Real-Time, Sensitive, and Specific DNA Detection.

The clustered regularly interspaced short palindromic repeats (CRISPR) technology has been widely applied for nucleic acid detection because of its high specificity. By using the highly specific and irreversible bond between HaloTag and its alkane chlorine ligand, we modified dCas9 (deactivated CRISPR/Cas9) with biotin as a biosensor to detect nucleic acids. The CRISPR biosensor was facilely prepared to adequately maintain its DNA-recognition capability. Furthermore, by coupling biolayer interferometry (BLI) with the CRISPR biosensor, a real-time, sensitive, and rapid digital system called CRISPR-BLI was established for the detection of double-stranded DNA. The CRISPR biosensor immobilised on the biolayer could recruit the target DNA onto the biosensor surface and change its optical thickness, resulting in a shift in the interference pattern and responding signal of the BLI. The CRISPR-BLI system was further applied to detect the ALP gene of Escherichia coli DH5α combined with a polymerase chain reaction, which demonstrated a linear range from 20 to 20 000 pg and a low detection limit (1.34 pg). The CRISPR-BLI system is a promising approach for rapid and sensitive detection of target DNA analytes.

Quantitation of plasma metanephrines using isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS): a candidate reference measurement procedure and its application to evaluating routine ID-LC/MS/MS methods.

Accurate measurement of plasma metanephrines (MNs) including metanephrine (MN) and normetanephrine (NMN) is crucial for the screening and diagnosis in pheochromocytomas and paragangliomas (PPGLs). Although the number of laboratories using liquid chromatography tandem mass spectrometry (LC-MS/MS) method to measure MNs has been increasing rapidly, those laboratory-developed assays showed incomparable results. There are no reference measurement procedures (RMPs) or reference materials (RMs) for MNs in Joint Committee for Traceability in Laboratory Medicine (JCTLM), which hindered the standardization of MNs measurement. We established a candidate RMP (cRMP) based on isotope dilution liquid chromatography tandem mass spectrometry (ID-LC/MS/MS) method for plasma MNs measurement. Plasma samples were spiked with MN-D3 and NMN-D3 as internal standards; protein precipitation and ion-exchange solid phase extraction (SPE) were performed to extract samples, eventually analyzed by LC-MS/MS. The cRMP was applied to evaluate two routine ID-LC/MS/MS methods through split-sample comparisons. Fifty-three individual patient samples were determined by cRMP and two routine ID-LC/MS/MS methods; results were analyzed by ordinary linear regression and Bland-Altman plots. The cRMP exhibited desirable imprecision, with intra-run and total imprecision (coefficient variation, CV) for MN being 0.79-1.36% and 1.53-1.87% and for NMN being 1.10-1.34% and 1.15-1.64%. The analytical recoveries of MN and NMN ranged from 98.3 to 101.7% and from 98.5 to 101.9%, respectively. Significant calibrator biases and sample-specific deviations were observed in method comparison. An accurate, precise, and reliable cRMP for plasma MNs was developed, and RMs with value assigned following the cRMP would help minimize the calibration bias and improve the comparability of different measuring systems.

Purine signaling regulating HSCs inflammatory cytokines secretion, activation, and proliferation plays a critical role in alcoholic liver disease.

Purine signaling pathway plays an important role in inflammation and tissue damage. To investigate the role of purine signaling pathway in acute alcoholic liver injury and chronic alcoholic liver fibrosis, we replicated two animal models and two cellular models. We found that body weights, liver indexes, serum biochemical parameters, serum fibrosis indexes, and pathological and immunohistochemical results had significant changes in two treatment groups compared with two control groups. In addition, gene expressions of purine receptors, inflammatory cytokines, fibrogenic cytokines, and inflammasomes increased obviously in two animal models and two cellular models. Furthermore, purine receptor inhibitors could significantly inhibit protein expressions of purine receptors and reduce protein expressions of inflammatory cytokines, fibrogenic cytokines, and inflammasomes. Besides, P2X7R small interfering ribonucleic acid (siRNA) had the same effects. Meanwhile, we detected protein expressions of inflammatory cytokines secreted by inflammasomes, and we found that purine receptor-mediated inflammasomes activation was a key event in the process of chronic alcoholic liver fibrosis. In summary, this study shows that inhibition of purine receptors can alleviate acute alcoholic liver injury and chronic alcoholic liver fibrosis in mice. Therefore, purine receptor is a potential new target for the treatment of acute alcoholic liver injury and chronic alcoholic fibrosis.

A facile top-down protocol for postsynthesis modification of hierarchical aluminum-rich MFI zeolites.

High aluminum content constitutes a major hurdle for the postsynthesis modification of hierarchical zeolites. A facile protocol comprising fluorination and sequential alkaline treatment is presented for the postsynthesis modification of hierarchical Al-rich MFI zeolites. By virtue of this protocol, uniform intracrystalline mesoporosity is introduced in an Al-rich MFI zeolite (Si/Al = 14.3). The obtained hierarchical zeolites exhibit a significant mesopore size distribution, centered around 6 nm, and show improved conversions in catalytic cracking of bulky aromatic molecules. The fundamental implications of the fluorination-alkaline treatment protocol are related to the formation of F-bearing tetrahedral aluminum species in the antecedent fluorination step, which alleviates the resistance of Al sites to the alkaline medium and causes Al-F complexation for regulated hydrolysis of the Al species during the alkaline treatment process. This top-down protocol and the derived mechanistic understandings are expected to be applied in the synthesis of hierarchical Al-rich zeolites with other framework topologies.

Identification of Immune Infiltration and Iron Metabolism-Related Subgroups in Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a prevalent neurodevelopmental disorder with a broad spectrum of symptoms and prognoses. Effective therapy requires understanding this variability. ASD children's cognitive and immunological development may depend on iron homoeostasis. This study employs a machine learning model that focuses on iron metabolism hub genes to identify ASD subgroups and describe immune infiltration patterns. A total of 97 control and 148 ASD samples were obtained from the GEO database. Differentially expressed genes (DEGs) and an iron metabolism gene collection achieved the intersection of 25 genes. Unsupervised cluster analysis determined molecular subgroups in individuals with ASD based on 25 genes related to iron metabolism. We assessed gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment, gene set variation analysis (GSVA), and immune infiltration analysis to compare iron metabolism subtype effects. We employed machine learning to identify subtype-predicting hub genes and utilized both training and validation sets to assess gene subtype prediction accuracy. ASD can be classified into two iron-metabolizing molecular clusters. Metabolic enrichment pathways differed between clusters. Immune infiltration showed that clusters differed immunologically. Cluster 2 had better immunological scores and more immune cells, indicating a stronger immune response. Machine learning screening identified SELENBP1 and CAND1 as important genes in ASD's iron metabolism signaling pathway. These genes express in the brain and have AUC values over 0.8, implying significant predictive power. The present study introduces iron metabolism signaling pathway indicators to predict ASD subtypes. ASD is linked to immune cell infiltration and iron metabolism disorders.

Altered neopterin and IDO in kynurenine metabolism based on LC-MS/MS metabolomics study: Novel therapeutic checkpoints for type 2 diabetes mellitus.

BACKGROUND: This study assessed the alternations of kynurenine pathway (KP) and neopterin in type 2 diabetes mellitus (T2DM) and explored possible differential metabolites. METHODS: A fresh residual sera panel was collected from 80 healthy control (HC) individuals and 72 T2DM patients. Metabolites/ratios of interest including tryptophan (TRP), kynurenine (KYN), 5-hydroxytryptamine (5HT), kynurenic acid (KA), xanthurenic acid (XA), neopterin (NEO), KA/KYN ratio and KYN/TRP ratio were determined using a targeted liquid chromatography-tandem mass spectrometry (LC-MS/MS) metabolomics approach, and the difference between groups was assessed. Supervised orthogonal partial least squares-discriminant analysis and differential metabolite screening with fold change (FC) were performed to identify distinct biomarkers. The diagnostic performance of KP metabolites in T2DM was evaluated. RESULTS: Significant decreases of TRP, 5HT, KA, XA, and KA/KYN and increases of KYN/TRP and NEO in T2DM compared to HC group were observed (P < 0.05). The KP metabolites panel significantly changed between T2DM and HC groups (Q2: 0.925, P < 0.005). 5HT (FC: 0.63, P < 0.01) and NEO (FC: 3.27, P < 0.01) were proven to be distinct differential metabolites. A combined testing of fasting plasma glucose and KYN/TRP showed good value in the prediction of T2DM (AUC: 0.904, 95% CI 0.843-0.947). CONCLUSIONS: The targeted LC-MS/MS metabolomics study is a powerful tool for evaluating the status of T2DM. This study facilitated the application of KP metabolomics into future clinical practice. 5HT and NEO are promising biomarkers in T2DM. KYN/TRP was highly associated with the development of T2DM and may serve as a potential treatment target.

Impact of locally delivered diphosphonates on dental implants: A systematic review and meta-analysis.

INTRODUCTION: Dental implants are a common solution for edentulous patients. This systematic review and meta-analysis aimed to determine whether locally delivered diphosphonates influence the osseointegration of dental implants in humans. MATERIAL & METHODS: In March 2023, we conducted an electronic systematic literature search using three databases (MEDLINE/PubMed, Embase, Web of Science). We included randomized trials documenting locally delivered diphosphonates in partly edentulous patients. Two independent reviewers evaluated study eligibility, extracted data, and assessed study quality. RESULTS: We have identified 752 studies, out of which 7 studies involving 154 patients met the inclusion criteria. The overall meta-analysis indicates that diphosphonates are associated with marginal bone loss during the pre-loading period (mean difference (MD) of -0.18 mm, 95% CI -0.24 to -0.12, p<0.00001; I²=83%), marginal bone loss after one year (MD -0.35 mm, 95% CI -0.56 to -0.14, p = 0.0009; I²=14%), and five years loading (MD -0.34 mm, 95% CI -0.56 to -0.13, p = 0.002; I²=0%). However, the drug did not seem to affect the implant survival rate (risk ratios (RR) of 1.02, 95% CI 0.98 to 1.08, P = 0.33; I²=9%). DISCUSSION: This study suggests that local use of diphosphonates does not affect implant survival, but it does reduce marginal bone loss and improve the osseointegration of dental implants in humans. However, future research must be more standardized and address methodological biases to draw more conclusive findings.

Effects of fermented Andrographis paniculata on growth performance, carcass traits, immune function, and intestinal health in Muscovy ducks.

The study aimed to examine the effects of unfermented and fermented Andrographis paniculata on growth performance, carcass traits, immune function, and intestinal health in Muscovy ducks. A total of 450 (16-day-old) Muscovy ducks weighing 271.44 ± 8.25 g were randomly assigned to 5 dietary treatments (6 replicate pens of 15 ducks per treatment), consisting of one control treatment (basal diet without A. paniculata), one unfermented A. paniculata treatment (basal diet plus 30 g/kg unfermented A. paniculata) and 3 fermented A. paniculata treatments (basal diet plus 10, 30, and 50 g/kg). 30 g/kg unfermented A. paniculata increased the ADG, thymus index, peripheral blood lymphocyte conversion rate, villi height, intestinal thickness, villi surface area, intraepithelial lymphocytes rate, while decreased the FCR. 10 g/kg fermented A. paniculata markedly boosted ADG, bursa of fabricius index, thymus index, serum lysozyme, lymphocyte conversion rate, villi height, vilii width, intestinal thickness, villi surface area, while decreased the FCR. 30 g/kg fermented A. paniculata clearly improved ADG, bursa of fabricius index, thymus index, serum lysozyme, lymphocyte conversion rate, villi height, vilii width, intestinal thickness, villi surface area, intraepithelial lymphocytes, while decreased FCR. 50 g/kg fermented A. paniculata significantly increased villi height, vilii width, and villi surface area, while clearly reduced BW. Additionally, compared to 30 g/kg unfermented A. paniculata, 30 g/kg fermented A. paniculata obviously increased bursa of fabricius indices, lymphocyte conversion rate, vilii width, villi surface area. On top of that, supplementation with unfermented and fermented A. paniculata (30 g/kg each) decreased the relative abundance of harmful bacteria (Succinivibrio, Succinatimonas, Sphaerochaeta, and Mucispirillum) and increase the abundance of beneficial bacteria (Rikenellaceae, Methanocorpusculum, Fournierella, Ruminococcaceae) in the ceca of the ducks. However, fermented A. paniculata had considerable better effects than unfermented A. paniculate on all above measured indices. Overall, these results revealed that supplementation with unfermented and fermented A. paniculata across different treatments improved growth, immune status, intestinal morphology, and intestinal microbiota composition and structure in Muscovy ducks, making it a potential alternative to antibiotics in poultry production.

CD73 mitigates hepatic damage in alcoholic steatohepatitis by regulating PI3K/AKT-mediated hepatocyte pyroptosis.

BACKGROUND: Alcohol use is a major risk factor for death and disability, resulting in a significant global disease burden. Alcoholic steatohepatitis (ASH) reflects an acute exacerbation of alcoholic liver disease (ALD) and is a growing health care and economic burden worldwide. Pyroptosis plays a central role in the pathogenesis of ASH. Nt5e (CD73) is a cell surface ecto-5'-nucleotidase, which is a key enzyme that converts the proinflammatory signal ATP to the anti-inflammatory mediator adenosine (ADO). Studies have found that CD73 is involved in multiple diseases and can alleviate gasdermin D (GSDMD)-mediated pyroptosis; however, its role and mechanism in ASH are not explicit. AIM: To investigate the role and mechanisms of CD73-mediated hepatocyte pyroptosis in alcohol-induced liver injury through in vivo and in vitro experiments. METHODS: CD73 knockout (CD73-/-) mice, wild-type (WT) mice, and AML-12 cells were used to evaluate the effect of CD73 on hepatocyte pyroptosis in vivo and in vitro. A combination of molecular and histological methods was performed to assess pyroptosis and investigate the mechanism both in vivo and in vitro. RESULTS: The protein expression of CD73 and pyroptosis pathway-associated genes was increased significantly in hepatocyte injury model both in vivo and in vitro. In vivo, CD73 knockout dramatically aggravated inflammatory damage, lipid accumulation, and hepatocyte pyroptosis in the liver. In vitro, overexpression of CD73 by pEGFP-C1/CD73 can decrease NLRP3 inflammasome activation and pyroptosis in hepatocytes. Further analysis revealed that the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway is a possible mechanism of CD73 regulation. Meanwhile, this pathological process was inhibited after the use of PI3K inhibitors. CONCLUSION: Our results show a novel function of CD73 regulates hepatocytes pyroptosis and highlights the therapeutic opportunity for reducing the disease process in ALD.

Development of a designed comparison method based on isotope dilution liquid chromatography-tandem mass spectrometry for determining plasma renin activity and its clinical assessment of renin activity stability in plasma.

Plasma renin activity (PRA) is recommended as the first screening indicator for primary aldosteronism. Immunoassays and liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods have been developed for quantifying PRA, but the interchangeability across assays and laboratories was suboptimal, which predominantly related to the differences in the plasma incubation strategy. This study aims to establish and validate a designed comparison method based on LC-MS/MS. The sensitivity, matrix effect, precision, accuracy, and storage stability were validated according to the Clinical Laboratory Standard Institution (CLSI) C-62A guidelines. The plasma incubation procedure was optimized to achieve maximum PRA results. The short-term stability of PRA plasma was assessed at 4 °C and room temperature (RT) for specific time points. Differences from the baseline were calculated using a one-way analysis of variance. The designed comparison method for PRA measurement exhibits excellent performance characteristics. The results from the 2022 national external quality assessment scheme for PRA showed good consistency of the developed method with other LC-MS/MS methods (relative biases: -6.8% to 4.6%), which demonstrated the reliability of the established method. Two sets of generation buffers were optimized to maximize the renin activity. The acetate buffer was recommended to be used in laboratory practice due to better metrological sensitivity. PRA plasma is stable for one day at 4 °C and RT. In summary, a reliable, traceable, and reproducible LC-MS/MS method for determining PRA was well-established and validated. The recommended incubation protocol is hoped to reduce the discrepancy in Ang1 generation. The evaluated short-term stability for PRA plasma could provide flexibility in clinical practice.

Effects of earthworm hydrolysate in production performance, serum biochemical parameters, antioxidant capacity and intestinal function of Muscovy ducks.

Earthworm has a variety of molecular biological characteristic, for example, growth promotion, antioxidant, and anti-bacteria. Thus, we decomposed earthworm by earthworm's own protease for preparing of earthworm hydrolysate. Muscovy ducks were fed with basal diet that formulated to contain 1.5% and 2.5% earthworm hydrolysate. Then, we investigated the influences of earthworm hydrolysate on growth performance in Muscovy ducks by performance terminology and measurement for poultry (NY/T 823-2020). The morphology of duodenum and number of intraepithelial lymphocytes were tested by HE staining and immunohistochemical method. Serum biochemical parameters and antioxidant capacity were also determined. High-throughput sequencing technology can sequence 16S rDNA of cecal contents from experimental Muscovy ducks. Results showed that 1.5% earthworm hydrolysate increased ADG (16-70 days old), ALB, HDL-C, T-AOC, CAT, SOD, GSH-PX, villi length, intestine thickness and surface area of villi (P < 0.05 or P < 0.01), and reduced FCR (16-70 days old), UREA, CRE, LDL-C, MDA (P < 0.05 or P < 0.01). Meanwhile, 2.5% improved ADG (16-70 days old), abdominal fat yield, breast muscle yield, heart index, spleen index, ALP, UA, T-AOC, CAT, SOD, GSH-PX, villi length, crypt depth, intestine thickness, surface area of villi, the percentage of intraepithelial lymphocytes (P < 0.05 or P < 0.01), and decreased FCR (42-70 days old and 16-70 days old), UREA, UA, MDA (P < 0.05 or P < 0.01). The sequencing results of gut flora demonstrated that earthworm hydrolysate improved variety of the gut flora in the V4 area of ducks immensely. In a word, our results provide the foundation for preliminary researching the potential principles of earthworm hydrolysate in promoting production performance, adjusting antioxidant function and intestinal functions in the Muscovy duck industry.

Abnormal kynurenine-pathway metabolites in gout: Biomarkers exploration based on orthogonal partial least squares-discriminant analysis.

BACKGROUND: This study aims to investigate serological characteristics of kynurenine pathway (KP) metabolites in healthy controls (HC) and gout patients and explore possible differential metabolites. METHODS: A total of 191 individual fresh residual sera was collected from 129 HC and 62 gout patients. A liquid chromatography-tandem mass spectrometry method was fully validated to measure 6 metabolites, including tryptophan (TRP), kynurenine (KYN), 5-hydroxytryptamine (5HT), kynurenic acid (KA), xanthurenic acid (XA), and neopterin (NEO). Supervised orthogonal partial least squares-discriminant analysis (OPLS-DA) and differential metabolite screening with fold change (FC) were performed to identify intrinsic variations and differential levels of KP metabolites between the HC and gout groups. Logistic regression was used to assess the contributions of KP metabolites to gout. RESULTS: There were significant decreases of TRP, 5HT, XA, and NEO and increases of KYN, KA, KA/KYN, and KYN/TRP in gout patients compared to the HC group (all p < 0.05). KP metabolites of the gout group showed good discrimination from those of the HC group (Q2: 0.892). Two distinct different metabolites were identified in gout, i.e., XA (FC: 0.56, p < 0.01) and NEO (FC: 0.34, p < 0.01). Of the KP metabolites, KYN was strongly associated with gout (OR: 7.91, p < 0.01). CONCLUSIONS: Abnormal levels of serum KP metabolites were observed in gout. XA and NEO are promising biomarkers that were relevant to the status of gout. The level of KYN could be an attractive checkpoint for the management of gout. Continuous monitoring of KP metabolism in gout provides new opportunities to predict therapeutic efficacy and prognosis.

Blocking ATP-P1Rs axis attenuate alcohol-related liver fibrosis.

AIMS: The aim of this study was to explore the fibrogenic effects of ATP-P1Rs axis and ATP-P2Rs axis on alcohol-related liver fibrosis (ALF). MATERIALS AND METHODS: C57BL/6J CD73 knock out (KO) mice were used in our study. 8-12 weeks male mice were used as an ALF model in vivo. In conclusion, after one week of adaptive feeding, 5 % alcohol liquid diet was given for 8 weeks. High-concentration alcohol (31.5 %, 5 g/kg) was administered by gavage twice weekly, and 10 % CCl4 intraperitoneal injections (1 ml/kg) were administered twice weekly for the last two weeks. The mice in the control group were injected intraperitoneally with an equivalent volume of normal saline. Fasting for 9 h after the last injection, blood samples were collected, and related indicators were tested. In vitro, rat hepatic stellate cells (HSCs) were treated with 200 µM acetaldehyde to establish an alcoholic liver fibrosis for 48 h, then tested related indicators. KEY FINDINGS: We found that both adenosine receptors including adenosine A1, A2A, A2B, A3 receptors (A1R, A2AR, A2BR, A3R) and ATP receptors including P2X7, P2Y2 receptors (P2X7R, P2Y2R) were expressed increased in ALF. After CD73 was knocked out, we found that adenosine receptors expression decreased, ATP expression increased, and fibrosis degree decreased. SIGNIFICANCE: Based on the research, we discovered that adenosine plays a more important role in ALF. Therefore, blocking the ATP-P1Rs axis represented a potential treatment for ALF, and CD73 will become a potential therapeutic target.

Realigning the melon chains in carbon nitride by rubidium ions to promote photo-reductive activities for hydrogen evolution and environmental remediation.

The photocatalytic efficiency of polymeric carbon nitride (PCN) suffers from unsatisfactory charge separation because of its amorphous structure. Herein, we report a simple bottom-up method to synthesize a novel structure of rubidium ion inserted PCN (Rb-PCN), which involves the regular alignment of melon chains to endow a crystalline feature in PCN. The insertion of Rb+ decreased not only the N p electrons in the heptazine ring but also the plane angle of the heptazine motifs in the melon chain, which promoted the long-range periodicity and crystallinity of carbon nitride. This structurally rearranged crystalline Rb-PCN demonstrated considerably enhanced separation of charge carriers, resulting in six-fold higher photocatalytic hydrogen evolution activity than its amorphous counterpart. Furthermore, the photoexcited electrons can be efficiently trapped by O2 to generate H2O2, which facilitates the production of reactive oxygen species to inactivate bacteria and degrade organic pollutants, showing great potential for use in both energy and environmental applications.

CD73/NT5E-mediated ubiquitination of AURKA regulates alcohol-related liver fibrosis via modulating hepatic stellate cell senescence.

Alcohol-related liver disease (ALD) is the most common chronic liver disease worldwide; however, no effective treatment to prevent the progression of alcohol-related liver fibrosis (ALF) is available. CD73/NT5E, a nucleotidase, controls cellular homeostasis by combining extracellular purinergic signaling with intracellular kinase activity and gene transcription and is associated with cell proliferation, differentiation, and death. In this study, we demonstrated that CD73/NT5E had a more significant regulatory effect on the activation, proliferation, and apoptosis of HSCs compared with that of CD39/ENTPD1. We examined the expression of CD73/NT5E in the normal and fibrotic human livers. The absence of CD73/NT5E was protective in mouse models of ALF. In addition, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses showed that CD73/NT5E overexpression was related to the p53 signaling pathway, which regulates cell senescence. Proteins interacting with p53 were predicted using the STRING database. The overlap between proteomic analysis and STRING databases was for Aurora kinase A (AURKA), a cell cycle-regulated kinase. Coimmunoprecipitation (co-IP) assay and molecular docking confirmed that CD73/NT5E directly interacted with AURKA. We found that overexpression of CD73/NT5E inhibited AURKA ubiquitination, whereas p53 signaling was downregulated. Mechanistically, CD73/NT5E regulated ALF and the activation and senescence of stellate cells by binding to AURKA. These findings indicate that CD73/NT5E is a potential therapeutic target for ALF.