The Clinical Impact of Metagenomic Next-Generation Sequencing for the Diagnosis of Periprosthetic Joint Infection.

Background: Synovial fluid metagenomic next-generation sequencing was introduced into the diagnosis of periprosthetic joint infection (PJI) in recent years. However, the clinical impact of mNGS remains unknown. Therefore, we performed a prospective cohort study to evaluate the clinical impact of mNGS for PJI diagnosis. Materials and Methods: Between April 2019 and April 2021, a total of 201 patients with suspected PJI were recruited in a high-volume PJI revision center. All patients underwent joint aspiration before surgeries and the obtained synovial fluids were sent to tests for the diagnosis of PJI. Based on the clinical evaluation of these patients, the patients were categorized into three groups: Group A: the mNGS reports were not acted upon. Group B: mNGS confirmed the standard diagnostic tests of PJI and generated identical clinical impact compared to standard diagnostic tests. Group C: mNGS results guided clinical therapy. Then, the concordance between synovial mNGS and cultures was analyzed. After that, multivariate regressions were performed to explore the "targeted populations" of mNGS tests. Results: A total of 107 patients were diagnosed with PJI based on the 2014 MSIS criteria and there were 33, 123, 45 patients in the group A, B, C respectively. The predictive factors of mNGS inducing clinical impact compared to standard diagnostic tests were negative culture results (adjusted OR: 5.88), previous history of joint infection (adjusted OR: 5.97), polymicrobial PJI revealed by culture (adjusted OR: 4.39) and PJI identified by MSIS criteria (adjusted OR: 17.06). Conclusion: When standard diagnostic tests for PJI were performed, about 22% of synovial fluid mNGS tests can change the treatment protocols built on standard diagnostic tests and affect the clinical practice. Thus, the use of synovial fluid mNGS in some "target" populations is more valuable compared to others such as patients with previous joint infection, polymicrobial PJI, and culture-negative PJI. Evidence Level: Level I.

Quercetin Alleviates Lipopolysaccharide-Induced Cell Oxidative Stress and Inflammatory Responses via Regulation of the TLR4-NF-κB Signaling Pathway in Bovine Rumen Epithelial Cells.

Subacute rumen acidosis (SARA) will cause an increase in endotoxin, which will have a negative effect on the bovine rumen epithelial cells (BREC). Flavonoids are effective in treating inflammation caused by endotoxin. Quercetin is a vital flavonoid widely occurring in fruits and vegetables and has received significant interest as a prospective anti-inflammatory antioxidant. Nonetheless, quercetin's protective machinery against such damage to BREC induced by lipopolysaccharide (LPS) remains unclear. A combined quercetin and LPS-induced BREC inflammation model was utilized to elucidate the effect of quercetin protecting BREC from LPS-induced injury. After treating BREC with different doses of LPS (1, 5, and 10 µg/mL) for 6 h or 24 h, the mRNA expression of inflammatory factors was detected. Our experimental results show the establishment of the BREC inflammation model via mRNA high expression of pro-inflammatory cytokines in BREC following 6 h treatment with 1 µg/mL LPS. The promotive effect of 80 µg/mL quercetin on BREC growth via the cell counting kit-8 (CCK8) assay was observed. The expression of pro-inflammatory cytokines and chemokines, notably tumor necrosis factor α (TNF-α), Interleukin 1ß (IL-1ß), IL-6, CC-motif chemokine ligand 2 (CCL2), CCL20, CCL28, and CXC motif chemokine 9 (CXCL9), etc., was significantly reduced by quercetin supplementation. We also analyzed the mRNA detection of related pathways by qRT-PCR. Our validation studies demonstrated that quercetin markedly curbed the mRNA expression of the toll-like receptor 4 (TLR4) and myeloid differentiation primary response protein (MyD88) and the nuclear factor-κB (NF-κB) in LPS-treated BREC. In addition, western blot result outcomes confirmed, as expected, that LPS significantly activated phosphorylation of p44/42 extracellular regulated protein kinases (ERK1/2) and NF-κB. Unexpectedly, this effect was reversed by adding quercetin. To complement western blot results, we assessed p-ERK1/2 and p-p65 protein expression using immunofluorescence, which gave consistent results. Therefore, quercetin's capacity to bar the TLR4-mediated NF-κB and MAPK signaling pathways may be the cause of its anti-inflammatory effects on LPS-induced inflammatory reactions in BREC. According to these results, quercetin may be utilized as an anti-inflammatory medication to alleviate inflammation brought on by high-grain feed, and it also lays out a conceptual foundation regarding the development and utilization of quercetin in the later stage.

Quercetin Alleviates Lipopolysaccharide-Induced Cell Damage and Inflammation via Regulation of the TLR4/NF-κB Pathway in Bovine Intestinal Epithelial Cells.

Acute diarrhoea and intestinal inflammation represent one of the most prevalent clinical disorders of milk production, resulting in enormous annual financial damage for the dairy sector. In the context of an unsatisfactory therapeutic effect of antibiotics, the natural products of plants have been the focus of research. Quercetin is an important flavonoid found in a variety of plants, including fruits and vegetables, and has strong anti-inflammatory effects, so it has received extensive attention as a potential anti-inflammatory antioxidant. However, the underlying basis of quercetin on inflammatory reactions and oxidative tension generated by lipopolysaccharide (LPS) in bovine intestinal epithelial cells (BIECs) is currently unexplained. This research aimed to determine the influence of quercetin on LPS-induced inflammatory reactions, oxidative tension, and the barrier role of BIECs. Our findings demonstrated that BIEC viability was significantly improved in LPS-treated BIEC with 80 µg/mL quercetin compared with the control group. Indicators of oxidative overload and genes involved in barrier role revealed that 80 µg/mL quercetin efficiently rescued BIECs from oxidative and barrier impairment triggered by 5 µg/mL LPS. In addition, the mRNA expression of pro-inflammatory cytokines TNF-α, IL-1ß, and IL-6, as well as chemokines CXCL2, CXCL5, CCL5, and CXCL8, was diminished in LPS-treated BIECs with 80 µg/mL quercetin compared with LPS alone. Furthermore, the mRNA expression of toll-like receptor 4 (TLR4), CD14, myeloid differential protein-2 (MD2), and myeloid differentiation primary response protein (MyD88) genes associated with the TLR4 signal mechanism was markedly reduced by the addition of quercetin to LPS-modulated BIECs, indicating that quercetin can suppress the TLR4 signal mechanism. We performed Western blotting on the NF-κB signalling mechanism and compared it with immunofluorescence to further corroborate this conclusion. The LPS treatment enhanced the proportions of p-IκBα/GAPDH and p-p65/GAPDH. Compared with the LPS-treated group, quercetin administration decreased the proportions of p-IκBα/GAPDH and p-p65/GAPDH. In addition, immunofluorescence demonstrated that quercetin greatly reduced the LPS-induced nuclear translocation of NF-κB p65 in BIECs. The benefits of quercetin on inflammatory reactions in LPS-induced BIECs may be a result of its capacity to inhibit the TLR4-mediated NF-κB signalling mechanism. These findings suggest that quercetin can be used as an anti-inflammatory reagent to treat intestinal inflammation induced by LPS release.

Astragalus membranaceus Additive Improves Serum Biochemical Parameters and Reproductive Performance in Postpartum Dairy Cows.

The purpose of the study was to assess the recovery, immune function, and breeding efficiency of postpartum dairy cows fed Astragalus membranaceus (AM) as a feed additive. The experiment used a completely randomized design. Cows were randomly assigned to two groups: (1) Control group fed total mixed ration (TMR; CON group, n = 15); (2) AM group fed TMR and AM (AM group, n = 15). The AM group was fed 675 g/day. The experimental results showed that compared with the CON group. The breeding interval of the AM group of dairy cows had a tendency to shorten (0.05 < p < 0.1). Plasma viscosity (PV), Plasma fibrinogen (FIB), the red cell aggregation index (TRCAI), Calcitonin (CT), Immunoglobulin M (IgM), and Luteinizing hormone (LH) results of AM group showed a time-treatment interaction (p < 0.05). Furthermore, the result of the study revealed that feeding AM as feed additives to dairy cows during the postpartum period had positive effects on wound recovery, immune function, endocrine regulation, and breeding efficiency.

Quercetin Alleviates Lipopolysaccharide-Induced Inflammatory Response in Bovine Mammary Epithelial Cells by Suppressing TLR4/NF-κB Signaling Pathway.

Bovine mastitis is one of the most common clinical diseases in dairy cows, causing huge economic losses to the dairy industry. Quercetin is an important flavonoid existing in many food resources, which has attracted widespread attention as a potential anti-inflammatory and antioxidant. However, the molecular mechanism of quercetin on inflammatory responses and oxidative stress in bovine mammary epithelial cells (BMECs) induced by lipopolysaccharide (LPS) remains unknown. The objective of this study was to investigate the effects of quercetin on inflammation responses, oxidative stress, and barrier function of BMEC induced by LPS. Our results showed that BMEC viability was not affected by treatment with 50 and 100 µg/ml of quercetin and 1 µg/ml of LPS compared with control group. The results of oxidative stress indicators and related genes of barrier function indicated that 100 µg/ml of quercetin effectively protected the BMECs from damage of oxidative and barrier induced by 1 µg/ml of LPS. Moreover, the messenger RNA (mRNA) expressions of pro-inflammatory cytokines TNF-α, IL-1ß, IL-6, and chemokines CXCL2, CXCL5, CCL5, and CXCL8 were markedly decreased in the LPS-treated bovine retinal endothelial cells (BRECs) with 100 µg/ml of quercetin relatively to LPS alone. More importantly, the mRNA expressions of toll-like receptor 4 (TLR4), CD14, myeloid differential protein-2 (MD2), and myeloid differentiation primary response protein (MyD88) genes involved in TLR4 signal pathway were significantly attenuated by the addition of quercetin in LPS-treated BMEC, suggesting that quercetin can inhibit the TLR4 signal pathway. In addition, immunocytofluorescence showed that quercetin significantly inhibited the nuclear translocation of NF-κB p65 in BMEC induced by LPS. Therefore, the protective effects of quercetin on inflammatory responses in LPS-induced BMEC may be due to its ability to suppress the TLR4-mediated NF-κB signaling pathway. These findings suggest that quercetin can be used as an anti-inflammatory reagent to treat mastitis induced by exogenous or endogenous LPS release.

Partial Substitution of Alfalfa Hay by Stevia (Stevia rebaudiana) Hay Can Improve Lactation Performance, Rumen Fermentation, and Nitrogen Utilization of Dairy Cows.

The objective of this study was to determine the effect of replacing isonitrogenous and isoenergetic basis alfalfa hay (AH) with stevia (Stevia rebaudiana) hay in dairy cow diets on nutrient digestion, milk performance, rumen fermentation, and nitrogen (N) utilization. In this study, 24 healthy Holstein lactating dairy cattle with a similar milk yield of 33.70 ± 2.75 (mean ± SD) kg, days in milk 95.98 ± 23.59 (mean ± SD) days, and body weight 587.75 ± 66.97 (mean ± SD) kg were selected and randomly allocated into three groups. The constituents of the three treatments were (1) 30.0% AH, and 0% stevia hay (SH) for the AH group; (2) 24.0% AH, and 6% SH for the 6% SH group; (3) 18.0% AH, and 12% SH for the 12% SH group. The substitution of AH with SH did not affect dry matter intake (DMI), gross energy (GE), and other nutrients intake but increased the digestibility of neutral detergent fiber (NDF) and acid detergent fiber (ADF). Compared with the AH diet, the cows fed the 6% SH diet had a higher milk yield and concentration of milk fat. Fecal and urinary nitrogen (N) were lower in cows fed a 6% SH diet than in cows fed the AH diet. Milk N secretion and milk N as a percentage of N intake were higher in cows fed a 6% SH diet than in cows fed AH diets. The concentration of ruminal volatile fatty acids, acetic acid, and ammonia-N were higher in cows fed a 6% SH diet than in cows fed an AH diet. By comparison, the 12% SH group did not affect milk yield, milk composition, N utilization, and rumen fermentation compared with the AH and 6% SH groups. In conclusion, it appears that feeding 6% SH, replacing a portion of AH, may improve lactation performance and N utilization for lactating dairy cows.

Effects of Buffalo Milk and Cow Milk on Lipid Metabolism in Obese Mice Induced by High Fat.

The aim of this study was to evaluate the effects of buffalo milk and cow milk on lipid metabolism in obese mice. Milk composition analysis showed fat, protein, and total solid content in buffalo milk was higher than cow milk, while the lactose content of buffalo milk was lower than cow milk. After milk metabolite extraction and LC-MS/MS analysis, differential metabolites were mainly enriched in "linoleic acid metabolism pathways," "pentose and glucuronate interconversion pathways," and "metabolism of xenobiotics by cytochrome P450 pathways." We fed three groups of C57BL/6J mice (n = 6 per group) for 5 weeks: (1) high-fat diet group (HFD group); (2) high-fat diet + buffalo milk group (HBM group); and (3) high-fat diet + cow milk group (HCM group). Our results showed that body weight of mice was significantly decreased in HBM and HCM groups from 1 to 4 weeks compared with the HFD group. The mRNA expression of ACAA2, ACACB, and SLC27A5 genes involved in the lipid metabolism in liver tissue were significantly elevated in HCM group, relatively to HFD and HBM group. In addition, the adipocyte number, size and lipid accumulation in the liver were significantly decreased in HCM group compared with the HFD group by H&E staining and oil red O staining, but was not change in HBM group. The mRNA levels of TNF-α and IL-1ß inflammatory genes were significantly increased in HBM group, relatively to HFD and HCM group, which is consistent with results from inflammatory cell infiltration and tissue disruption by colon tissue sections. In conclusion, dietary supplementation of cow milk has beneficial effects on loss of weight and lipid metabolism in obese mice.

Growth Performance, Rumen Fermentation and Inflammatory Response on Holstein Growing Cattle Treated with Low and High Non-Fibrous Carbohydrate to Neutral Detergent Fiber Ratio Pelleted Total Mixed Ration.

Twenty-four healthy castrated male Holstein growing cattle, with similar body weight (301 ± 11.5 kg), were enrolled in this study and randomly allocated into two groups (12/pen). Holstein growing cattle in the LPT (low NFC/NDF pelleted TMR) group were fed basal pelleted TMR with a low NFC/NDF ratio (NFC/NDF = 1.07), while the HPT (high NFC/NDF pelleted TMR) group were fed with a high NFC/NDF ratio diet (NFC/NDF = 1.71). The results showed that: (1) Body measurements were found to be increased for the LPT group (p < 0.05); compared with the HPT group, feed intake to gain ratio and feed cost in the LPT group were decreased by 12.24% and 15.35%, respectively (p < 0.01). Compared with the HPT group, the LPT group tended to increase chest girth. (2) Digestibility of DM and NDF in the LPT group was higher (p < 0.05) than in the HPT group, being increased by 3.41% and 4.26%, respectively, and increased digestibility of ADF in the LPT group was significant (p < 0.01). (3) The daily feed consumption of NDF in the LPT group was higher than that in the HPT group and the daily rumination time and chewing time in the LPT group were longer than that in the HPT group (p < 0.05). (4) Compared with the LPT group, the parameter of pH, microbial protein and acetate: propionate (p < 0.05) in the HPT group were decreased by 8.57%, 12.46% and 23.71%, respectively. In contrast, the concentration of total volatile fatty acids, acetate and propionate were higher (p < 0.05) in the HPT group, and increased by 13.49%, 19.59% and 52.70%, respectively. (5) Compared with the LPT group, rumen fluid in the HPT group diet up-regulated the mRNA expression levels of BRECs pro-inflammatory factor IL-1ß and TNF-α (p < 0.05), and meanwhile, up-regulated the mRNA expression levels of BRECs pro-inflammatory factor IL-6 (p < 0.01); compared with the LPT group, rumen fluid in the HPT group diet up-regulated the mRNA expression levels of CCL28 and CCL20 (p < 0.05) chemokines in CCL types of BRECs; in addition, compared with the LPT group, rumen fluid in the HPT group up-regulated the mRNA expression levels of CXCL2, CXCL3, CXCL9 and CXCL14 chemokines in CXCL types of BRECs (p < 0.01), and the mRNA expression levels of the CXCL5 chemokine tended to be increased (p = 0.06).

1 H NMR-based metabonomic evaluation of the pesticides camptothecin and matrine against larvae of Spodoptera litura.

BACKGROUND: Camptothecin (CPT) and matrine (MAT) have potential as botanical pesticides against several pest species. However, the mechanisms of metabolic and physiological changes in pests induced by CPT and MAT are unknown. In this study, a toxicological test, an NMR-based metabolomic study, an enzymatic test, and an RT quantitative PCR (RT-qPCR) experiment were all conducted to examine the effect of CPT and MAT on Spodoptera litura. RESULTS: CPT (0.5-1%) exerted high toxicity against larvae of S. litura and caused growth stagnation and high mortality of larvae. A variety of metabolites were significantly influenced by 0.5% CPT, including several energy-related metabolites such as trehalose, lactate, succinate, citrate, malate, and fumarate. In contrast, MAT showed low toxicity against larvae and induced almost no changes in hemolymph metabolites of S. litura. Enzymatic tests showed that trehalase activity was significantly decreased in larvae after feeding with 0.5% CPT. RT-qPCR showed that the transcription levels of alanine aminotransferase, malate dehydrogenase, and isocitrate dehydrogenase were decreased while lactate dehydrogenase was increased in the 0.5% CPT-treated group. CONCLUSIONS: These data indicate that one of the important mechanisms of CPT against S. litura larvae is via the inhibition of trehalose hydrolysis and glycolysis. Our findings also suggest that CPT exhibits a stronger toxicological effect than MAT against S. litura, which provides basic information for the application of CPT in the control of S. litura or other lepidoptera pests.

Left-lateral thoracotomy for catheter ablation of scar-related ventricular tachycardia in patients with inaccessible pericardial access.

OBJECTIVES: We aimed to describe the feasibility of a surgical left thoracotomy for catheter ablation of scar-related ventricular tachycardia (VT) in patients with inaccessible pericardial access. BACKGROUND: Pericardial adhesion due to prior cardiac surgery or previous epicardial ablation procedures limits epicardial access in patients with drug-refractory VT originated from the epicardium. METHODS: Six patients who underwent a surgical left lateral thoracotomy epicardial access for catheter ablation of VT after failed subxiphoid percutaneous epicardial access were reviewed. Patients' baseline characteristics and procedural characteristics including epicardial access, mapping, and ablation were described. Epicardial access was successfully obtained in all patients by a surgical left lateral thoracotomy. RESULTS: The reasons of pericardial adhesion were prior cardiac surgery (n = 3, 50%) and previous epicardial ablation procedures (n = 3, 50%). Epicardial mapping of the lateral and inferior left ventricle was acquired, and a total of 15 different VTs originated from those regions were abolished. Unless one patient with ST elevation myocardial infarction due to periprocedural occlusion of the posterior descending artery no further complications occurred. All patients were discharged 10.2 ± 4 days after the procedure. VT recurred in 1 patient (17%) and was controlled with oral amiodarone therapy during follow-up (median follow-up: 479 days). CONCLUSIONS: A surgical left lateral thoracotomy is feasible and safe for selected patients. This approach provides epicardial ablation in patients with VT located at the infero-lateral left ventricle and pericardial adhesions due to previous cardiac surgery or previous ablation procedures.

Sanguinarine caused larval lethality and growth inhibition by suppressing energy metabolism in silkworms, Bombyx mori.

Sanguinarine (Sang) is a natural alkaloid and distributed in several plants of Papaveraceae. The antitumor, antioxidant, antimicrobial and anti-inflammatory effects of Sang were extensively reported, but its speciality and mechanism against Lepidoptera insects were still unknown. In this study, detailed toxicological parameters of Sang against silkworms, Bombyx mori (B. mori), were determined by a toxicological test. Then, a nuclear magnetic resonance-based (NMR) metabolomics method was adopted to analyze the changes in hemolymph metabolites of silkworms after feeding Sang. The growth of fourth-instar larvae was significantly ceased by the oral administration of 0.05-0.3% Sang and vast deaths appeared in 0.3% Sang group on Day 4 and Day 5. The quantitative analysis of metabolites indicated that trehalose and citrate levels in hemolymph were increased after 24 h of feeding 0.3% Sang, whereas the concentrations of pyruvate, succinate, malate and fumarate were decreased. In addition, the enzymatic determination and reverse transcription quantitative PCR (RT-qPCR) showed that the trehalase (THL) activity and the transcriptional level of one gene coding THL were uniformly weakened by 0.3% Sang. One of the important mechanisms of Sang against silkworms might be interpreted as follows. Sang impaired trehalose hydrolysis, reduced THL activity and transcription, and led to the inhibition of energy metabolism, consequent antigrowth and high lethality in larvae of B. mori. Our findings offered new insights into the insecticidal effect of Sang from the perspective of energy metabolism and provided the basis for the application of Sang in the control of Lepidoptera pests.

Combination of Curcumin with an Anti-Transferrin Receptor Antibody Suppressed the Growth of Malignant Gliomas In vitro.

AIM: Transferrin receptor (TfR) has been used as a target for the molecular cancer therapy due to its higher expression in a variety of tumors. Anti-TfR antibodies combined with chemotherapeutic drugs has showed great potential as a possible cancer therapeutic strategy. In our study, we investigated the anti-tumor effects of anti-TfR monoclonal antibody (mAb) alone or in combination with curcumin in vitro. MATERIAL AND METHODS: We detected the apoptosis, proliferation and cell cycle of glioma cells after treated with anti-TfR mAb and curcumin alone or the combinations by flow cytometer. RESULTS: Anti-TfR mAb or curcumin could inhibit proliferation of tumor cells. Anti-TfR mAb marked S phase arrest and curcumin induced G2/M arrest in tumor cells. When anti-TfR mAb and curcumin were used simultaneously, a synergistic effect was detected in relation to tumor growth inhibition and the induction of cells necrosis. CONCLUSION: These results provided a potential role of anti-TfR mAb-containing curcumin in the treatment for gliomas.

Identification of a novel NOG mutation in a Chinese family with proximal symphalangism.

Proximal symphalangism (SYM1) is an autosomal dominant disorder, mainly characterized by variable fusion of the proximal interphalangeal joints of the hands and feet. To date, two genes, GDF5 and NOG, have been reported to associate with SYM1. Herein, we clinically characterized a Chinese family with fusions of the bilateral proximal interphalangeal joints in the 2-5 digits without conductive hearing loss. Direct DNA sequencing of the two genes revealed a novel heterozygous missense mutation (c.499C>T, p.R167C) in the NOG gene. This mutation co-segregates with the phenotype in the family and is not present in the 200 control individuals. The c.499C>T mutation is predicted to change the conserved amino acid arginine at codon 167 to cysteine at the protein level. A different mutation in the same codon (R167G) has been described to cause brachydactyly type B2 (BDB2). Our work indicates that the c.499C>T (R167C) mutation is likely to represent the pathogenic mutation in the family. This finding broadens the spectrum of NOG mutations associated with SYM1 and will help to provide genetic counseling to the affected family.

[NF1 mutation analysis in a Chinese family with neuro- fibromatosis type].

A Chinese family affected with autosomal dominant disorder-neurofibromatosis type I was identified in this study. Linkage analysis was performed, and DNA sequencing for whole coding region of NF1 was carried out to identify the disease-causing mutation. The disease gene of the Chinese NF1 family was linked to NF1 locus, and a nonsense mutation, G1336X in the NF1 gene was identified. This mutation truncates the NF1 protein by 1 483 amino acid residues at the C-terminus, and is co-segregate with all the patients, but not present in unaffected individuals in the family. The present study demonstrated that G1336X mutation in the NF1 gene cause Neurofibromatosis type I in the family. To our knowledge, this mutation is firstly reported in Chinese population.