Enhanced Downstream Processing for a Cell-Based Avian Influenza (H5N1) Vaccine.

H5N1 highly pathogenic avian influenza virus (HPAIV) infections pose a significant threat to human health, with a mortality rate of around 50%. Limited global approval of H5N1 HPAIV vaccines, excluding China, prompted the need to address safety concerns related to MDCK cell tumorigenicity. Our objective was to improve vaccine safety by minimizing residual DNA and host cell protein (HCP). We developed a downstream processing method for the cell-based H5N1 HPAIV vaccine, employing CaptoTM Core 700, a multimodal resin, for polishing. Hydrophobic-interaction chromatography (HIC) with polypropylene glycol as a functional group facilitated the reversible binding of virus particles for capture. Following the two-step chromatographic process, virus recovery reached 68.16%. Additionally, HCP and DNA levels were reduced to 2112.60 ng/mL and 6.4 ng/mL, respectively. Western blot, high-performance liquid chromatography (HPLC), and transmission electron microscopy (TEM) confirmed the presence of the required antigen with a spherical shape and appropriate particle size. Overall, our presented two-step downstream process demonstrates potential as an efficient and cost-effective platform technology for cell-based influenza (H5N1 HPAIV) vaccines.

Serum metabolomics analysis of biomarkers and metabolic pathways in patients with colorectal cancer associated with spleen-deficiency and qi-stagnation syndrome or damp-heat syndrome: a prospective cohort study.

Objective: To profile the serum metabolites and metabolic pathways in colorectal cancer (CRC) patients associated with spleen-deficiency and qi-stagnation syndrome (SDQSS) or damp-heat syndrome (DHS). Methods: From May 2020 to January 2021, CRC patients diagnosed with traditional Chinese medicine (TCM) syndromes of SDQSS or DHS were enrolled. The clinicopathological data of the SDQSS and DHS groups were compared. The serum samples were analyzed by liquid chromatography-mass spectrometry (LC-MS). The variable importance in the projection >1, fold change ≥3 or ≤0.333, and P value ≤0.05 were used to identify differential metabolites between the two groups. Furthermore, areas under the receiver operating characteristic (ROC) curve > 0.9 were applied to select biomarkers with good predictive performance. The enrichment metabolic pathways were searched through the database of Kyoto Encyclopedia of Genes and Genomes. Results: 60 CRC patients were included (30 SDQSS and 30 DHS). The level of alanine aminotransferase was marginally significantly higher in the DHS group than the SDQSS group (P = 0.051). The other baseline clinicopathological characteristics were all comparable between the two groups. 23 differential serum metabolites were identified, among which 16 were significantly up-regulated and 7 were significantly down-regulated in the SDQSS group compared with the DHS group. ROC curve analysis showed that (S)-3-methyl-2-oxopentanoic acid, neocembrene, 1-aminocyclopropanecarboxylic acid, 3-methyl-3-hydroxypentanedioate, and nicotine were symbolic differential metabolites with higher predictive power. The top five enrichment signalling pathways were valine, leucine and isoleucine biosynthesis; lysosome; nicotine addiction; fructose and mannose metabolism; and pertussis. Conclusion: Our study identifies the differential metabolites and characteristic metabolic pathways among CRC patients with SDQSS or DHS, offering the possibility of accurate and objective syndrome differentiation and TCM treatment for CRC patients.

INTRINSIC/EXTRINSIC APOPTOSIS AND PYROPTOSIS CONTRIBUTE TO THE SELECTIVE DEPLETION OF B CELL SUBSETS IN SEPTIC SHOCK PATIENTS.

ABSTRACT: The depletion of peripheral blood B cells is associated with immunosuppression and poor prognosis during sepsis, and selective depletion occurs when B cell subsets are specifically targeted. In this study, we examined the mechanisms underlying the selective depletion of B cell subsets in the immunosuppressive phase of septic shock patients. Thirty-two septic shock patients were recruited as a septic shock group and 10 healthy volunteers as a control group. The expression of Bcl-2, CD95, cleaved caspase-9/8, and activated caspase-3/1 in the B cell subsets were measured by flow cytometry. Another 23 septic shock patients were recruited to test the remission of caspase-3 (Z-DEVD-FMK) and caspase-1 (VX-765) inhibitors on B cell subset depletion in vitro . In septic shock patients, the Bcl-2 levels in immature/transitional (IM) B cells decreased and the levels of cleaved caspase-9 in IM B cells increased; the levels of CD95 in IM, naive, resting memory (RM), and activated memory (AM) B cells and the levels of cleaved caspase-8 in IM, RM, and AM B cells increased; the levels of activated caspase-3 and caspase-1 in IM, RM, and AM B cells increased. Activated caspase-1 levels in IM B cells were higher compared with activated caspase-3 in septic shock patients, whereas the levels of activated caspase-1 in AM B cells were lower compared with activated caspase-3. Moreover, in vitro experiments showed that Z-DEVD-FMK and VX-765 could alleviate the depletion of IM, AM, and RM B cells. The selective reduction of circulating B cell subsets in septic shock patients could be attributed to intrinsic and extrinsic apoptosis as well as pyroptosis.

Insights into the Roles of B Cells in Patients with Sepsis.

Sepsis is a life-threatening yet common disease, still posing high mortality worldwide. Sepsis-related deaths primarily occur during immunosuppression; the disease can hamper the numbers and function of B cells, which mediate innate and adaptive immune responses to maintain immune homeostasis. Dysfunction of B cells, along with aggravated immunosuppression, are closely related to poor prognosis. However, B cells in patients with sepsis have garnered little attention. This article focuses on the significance of B-cell subsets, including regulatory B cells, in sepsis and how the counts and function of circulating B cells are affected in patients with sepsis. Finally, potential B-cell-related immunotherapies for sepsis are explored.

[Non-Invasive Prenatal Testing:Results in 3733 Cases of Twin Pregnancy and Association With Factors Such as Age].

Objective To explore the clinical significance of non-invasive prenatal testing(NIPT)for fetal chromosomal abnormalities in the cases of twin pregnancy and its relationship with age and other related factors.Methods A total of 3733 women with twin pregnancy of 12-26+6 weeks who voluntarily underwent NIPT in the Ningbo Women and Children's Hospital from January 2018 to December 2022 were selected.The results of NIPT and amniocentesis were compared and all the participants were followed up.The detection rate of chromosomal abnormalities by NIPT was calculated,and its correlations with age,gestational weeks,chorionicity,and pregnancy type were analyzed.Results Among the 3733 cases,71 cases of fetal chromosome abnormality were indicated by NIPT,including 13 cases of trisomy 21,19 cases of trisomy 18,5 cases of trisomy 13,18 cases of sex chromosome abnormality,and 16 cases of chromosome microdeletion/duplication(excluding 21,18,13,and sex chromosomes),among which 34 cases were true positive and 37 cases were false positive.The overall sensitivity,specificity,and positive predictive value(PPV)of NIPT for chromosomal abnormalities in the cases of twin pregnancy were 100%,98.99%,and 47.89%(34/71),respectively.NIPT showed the sensitivity,specificity,and PPV of 100%,99.78%,and 78.38%(29/37)for trisomy 21,18,and 13,100%,99.56%,and 16.67%(3/18)for sex chromosome abnormalities,and 100%,99.62%,and 12.5%(2/16)for chromosome microdeletion/duplication,respectively.In the age group of ≥40 years,the NIPT for chromosomal abnormalities showed the PPV of 66.67%,the sensitivity of 100%,and the misdiagnosis rate of 30%。However,the NIPT for trisomy 21,18,and 13 showed the PPV of 100%,the misdiagnosis rate of 0,and the sensitivity and specificity of 100%.In terms of grouping based on gestational weeks,the NIPT for chromosomal abnormalities showed the highest PPV(51.28%)in the women with twin pregnancy for 14-17+6 weeks,followed by that(50.00%)in the women with twin pregnancy for 22-26+6 weeks;the NIPT for trisomy 21,18,and 13 showed the highest PPV of 94.74% in the gestation group of 14-17+6 weeks,followed by that(83.33%)in the gestation group of 18-21+6 weeks.The rate of dichorionic diamniotic twins was higher in assisted pregnancies than in natural pregnancies,and NIPT showed the same detection efficiency for dichorionic diamniotic twins and monochorionic diamniotic twins and the same detection efficiency for different pregnancy types.Conclusions NIPT has high accuracy in the diagnosis of twin pregnancy and high sensitivity and high specificity for different ages and gestational weeks,especially for trisomy 21,18,and 13.NIPT is suitable for assisted pregnancy and natural pregnancy,and it is of high value in clinical application.However,extensive application needs a large population-based study.

Prognostic Value of an Integrin-Based Signature in Hepatocellular Carcinoma and the Identification of Immunological Role of LIMS2.

Objective: Evidence proves that integrins affect almost every step of hepatocellular carcinoma (HCC) progression. The current study aimed at constructing an integrin-based signature for prognostic prediction of HCC. Methods: TCGA-LIHC and ICGC-LIRI-JP cohorts were retrospectively analyzed. Integrin genes were analyzed via univariate Cox regression, followed by generation of a prognostic signature with LASSO approach. Independent factors were input into the nomogram. WGCNA was adopted to select this signature-specific genes. Gene Ontology (GO) enrichment together with Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were conducted to explore the function of the dysregulated genes. The abundance of tumor microenvironment components was estimated with diverse popular computational methods. The relative importance of genes from this signature was estimated through random-forest method. Results: Eight integrin genes (ADAM15, CDC42, DAB2, ITGB1BP1, ITGB5, KIF14, LIMS2, and SELP) were adopted to define an integrin-based signature. Each patient was assigned the riskScore. High-riskScore subpopulation exhibited worse overall survival, with satisfying prediction efficacy. Also, the integrin-based signature was independent of routine clinicopathological parameters. The nomogram (comprising integrin-based signature, and stage) accurately inferred prognostic outcome, with the excellent net benefit. Genes with the strongest positive interaction to low-riskScore were primarily linked to biosynthetic, metabolic, and catabolic processes and immune pathways; those with the strongest association with high-riskScore were principally associated with diverse tumorigenic signaling. The integrin-based signature was strongly linked with tumor microenvironment components. Among the genes from this signature, LIMS2 possessed the highest importance, and its expression was proven through immunohistochemical staining. Conclusion: Altogether, our study defined a quantitative integrin-based signature that reliably assessed HCC prognosis and tumor microenvironment features, which possessed the potential as a tool for prognostic prediction.

Development and Validation of a Nomogram for Adverse Outcomes of Geriatric Trauma Patients Based on Frailty Syndrome.

Purpose: Currently, assessing trauma severity alone in geriatric trauma patients (GTPs) cannot accurately predict the risk of serious adverse outcomes during hospitalization. As an emerging concept in recent years, frailty syndrome is closely related to the poor prognosis of many diseases in elderly patients, including trauma. A logistic model for predicting adverse outcomes in elderly trauma patients during hospitalization was constructed in elderly patients, and the predictive efficacy of the model was verified. Patients and Methods: Trauma patients aged ≥65 years between June 2020 and September 2021 were selected and randomly divided into a training set and validation set at a ratio of 3:1. Mid arm muscle circumference (MAMC) was measured to determine the degree of frailty. LASSO regression was used to screen appropriate variables for the construction of a prognostic model. The logistic regression model was established and presented in the form of a nomogram. Calibration curves and ROC curves were used to verify the performance of the model. Results: A total of 209 patients were enrolled, including 143 (68.4%) males and 66 (31.6%) females, with an average age of 70.8 ± 4.8 years. Ageless Charlson comorbidity index, BT unit, ISS, GCS, MAMC, prealbumin and lactic acid levels were screened by LASSO regression to construct a prognostic model. The AUC of the ROC analysis prediction model was 0.89 (95% CI 0.80-0.97) in the validation set. The results of the Hosmer-Lemeshow test for the validation set were χ2 = 11.23, P = 0.189. Conclusion: The prognostic model of adverse outcomes in GTPs has good accuracy and differentiation, which can improve the prediction results of risk stratification of GTPs during hospitalization by medical staff and provide a new idea for prognostic prediction.

Closed-Surface Multifunctional Antireflective Coating Made from SiO2 with TiO2 Nanocomposites.

An SiO2-TiO2 closed-surface antireflective coating was fabricated by the one-dipping method. TiO2 nanoparticles were mixed with a nanocomposited silica sol, which was composed of acid-catalyzed nanosilica networks and silica hollow nanospheres (HNs). The microstructure of the sol-gel was characterized by transmission electron microscopy. The silica HNs were approximately 40-50 nm in diameter with a shell thickness of approximately 8-10 nm. The branched-chain structure resulting from acidic hydrolysis grew on these silica HNs, and TiO2 was distributed inside this network. The surface morphology of the coating was measured by field emission scanning electron microscopy and atomic force microscopy. After optimization, transmittance of up to 94.03% was obtained on photovoltaic (PV) glass with a single side coated by this antireflective coating, whose refractive index was around 1.30. The short-circuit current gain of PV module was around 2.14-2.32%, as shown by the current-voltage (IV) curve measurements and external quantum efficiency (EQE) tests. This thin film also exhibited high photocatalytic activity. Due to the lack of voids on its surface, the antireflective coating in this study possessed excellent long-term reliability and robustness in both high-moisture and high-temperature environments. Combined with its self-cleaning function, this antireflective coating has great potential to be implemented in windows and photovoltaic modules.

Octyl-2-cyanoacrylate tissue adhesive without subcuticular suture for wound closure after total hip arthroplasty: a prospective observational study on thirty-two cases with controls for 3 months follow-up.

BACKGROUND: Whether using tissue adhesive alone after subcutaneous suture can close the skin incision with safety as well as cosmetic appearance after total hip arthroplasty was not clear. METHODS: A prospective study was conducted. The same surgical methods were consistent throughout the entire study. After implanting prosthesis, the joint capsule was reconstructed. Fascial and subcutaneous layer were respectively closed by continuous running barbed suture. Patients were randomized allocated to group A with octyl-2-cyanoacrylate tissue adhesive alone, to group B with tissue adhesive after continuous subcuticular suture, or to group C with skin staples. Time of closure, drainage, pain, wound complications, and cosmesis were compared. All data were analyzed statistically. RESULTS: There was no significant difference in drainage, Visual Analog Scale score or early wound complications between the three groups. However, there was significant difference in time of closure (P = 0.013). In pairwise comparison, time of closure in groups A and B was significantly longer than those in group C (P = 0.001 and P = 0.023, respectively); time of closure in group A was significantly shorter than those in group B (P = 0.003). Patient and Observer Scar Assessment Scale total scores were not significantly different at 6 weeks and 3 months postoperatively (P = 0.078 and P = 0.284, respectively). CONCLUSION: Tissue adhesive without subcuticular suture was similar with a combination of subcuticular suture and tissue adhesive as well skin staples in terms of safety and cosmetic appearance after total hip arthroplasty.

In vitro activities of telithromycin against Staphylococcus aureus biofilms compared with azithromycin, clindamycin, vancomycin and daptomycin.

Introduction. Staphylococcus aureus biofilms are difficult to treat and the effect of telithromycin treatment is still unclear.Aim. This study aimed to explore the effect of telithromycin against Staphylococcus aureus biofilms compared with azithromycin, clindamycin, vancomycin and daptomycin.Methodology. Eight methicillin-susceptible and eight methicillin-resistant S. aureus isolates (MSSA and MRSA, respectively) were used for this study. Biofilm biomasses were detected by crystal violet staining and the adherent cells in the established biofilms were quantified by determination of colony-forming units (c.f.u.). The RNA levels of biofilm formation-related genes were determined by RT-qPCR.Results. Telithromycin [8× minimum inhibitory concentration (MIC)] eradicated more established biofilms than azithromycin or clindamycin in the four MSSA isolates, and eliminated the established biofilms of six MRSA isolates more effectively than vancomycin or daptomycin. Telithromycin (8× MIC) killed more adherent cells in the established biofilms than azithromycin or clindamycin in the six MSSA isolates, and killed more adherent cells than vancomycin in all eight MRSA isolates. Daptomycin also showed an excellent effect on the adherent cells of MRSA isolates, with similarresults to telithromycin. The effect of a subinhibitory concentration of telithromycin (1/4× MIC) was significantly superior to that of azithromycin or clindamycin, inhibiting the biofilm formation of six MSSA isolates and seven MRSA isolates more effectively than vancomycin or daptomycin. The RNA levels of agrA, agrC, clfA, icaA and sigB decreased when treated with telithromycin (1/4× MIC).Conclusions. Telithromycin is more effective than azithromycin, clindamycin, vancomycin, or daptomycin against S. aureus biofilms.

[Energy Demand and Its Regulatory Mechanism during Folliculogenesis].

The growth and development of follicles are regulated by genes,hormones and growth factors autocrined and paracrined from granulosa cells,theca cells,and oocytes.Products of glycolysis from granulosa cells such as pyruvate and lactate are one of the main energy sources,which play an important role during folliculogenesis and follicle maturity.Studies on the changes of the products and rate-limiting enzymes during granulosa cells' glycolysis help to clarify the molecular mechanism of energy demand in folliculogenesis and guide the clinical treatment of infertility due to abnormal follicular development.This article reviews recent research advances in the energy demand and regulatory mechanism in different states of folliculogenesis.

Effect of Long-Time Annealing at 1000 °C on Phase Constituent and Microhardness of the 20Co-Cr-Fe-Ni Alloys.

The phase constituent and microhardness of the arc-melted 20Co-Cr-Fe-Ni alloys, in both as-cast state and after annealing at 1000 °C for 30 days, were experimentally investigated by scanning electron microscopy (SEM) and X-ray diffraction (XRD). Experiment results indicated that a uniform, stable, single Face-Center Cubic (FCC) phase can be obtained in as-cast 20 Co-Cr-Fe-Ni alloys with less than 30 at.% Cr. Annealing at 1000 °C has no effect on their phase composition and microhardness. When the Cr content is above 40 at.%, the σ phase forms and its volume fraction increases with the Cr content, which leads to an increase in microhardness. Annealing at 1000 °C for 30 days can slightly decrease the volume fraction of the σ phase and slightly decrease the alloy microhardness. Except for the Fe-rich alloys, the alloy microhardness increases with the Cr content when the Co and Ni or the Co and Fe contents were fixed. Moreover, comparing with the thermodynamically calculated phase diagram based on the TCFE database, it has been proved that the calculation can predict the phase stability of the FCC phase and the 1000 °C isothermal section. However, it fails to predict the stability of the σ phase near the liquidus. The present results will help to design and process treatment of the Co-Cr-Fe-Ni based high entropy alloys.

In vitro activities of daptomycin combined with fosfomycin or rifampin on planktonic and adherent linezolid-resistant isolates of Enterococcus faecalis.

PURPOSE: This study aimed to explore daptomycin combined with fosfomycin or rifampin against the planktonic and adherent linezolid-resistant isolates of Enterococcus faecalis. METHODOLOGY: Four linezolid-resistant and four linezolid-sensitive isolates of E. faecalis which formed biofilms were collected for this study. Biofilm biomasses were detected by crystal violet staining and the adherent cells in the mature biofilms were quantified by c.f.u. determination. RESULTS: Daptomycin alone, or combined with fosfomycin or rifampin (4×MIC) demonstrated bactericidal activities on the planktonic cells, and daptomycin combined with fosfomycin killed more planktonic cells (at least 1-log10 c.f.u. ml-1) than daptomycin or fosfomycin alone. Daptomycin alone (16×MIC) showed anti-biofilm activities against the mature biofilms and bactericidal activities on the adherent cells, while daptomycin combined with fosfomycin (16×MIC) demonstrated significantly more anti-biofilm activities than daptomycin or fosfomycin alone and effectively killed the adherent cells in the mature biofilms. The high concentration of daptomycin (512 mg l-1 ) combined with fosfomycin indicated more bactericidal activities on the adherent cells and more anti-biofilm activities against the mature biofilms than daptomycin 64 mg l-1 (16×MIC) combined with fosfomycin. The addition of rifampin increased the anti-biofilm and bactericidal activities of daptomycin against the mature biofilms and the adherent cells of two isolates, however, which was not observed in other isolates. CONCLUSIONS: Daptomycin combined with fosfomycin demonstrated better effect on the planktonic and adherent linezolid-resistant isolates of E. faecalis than daptomycin or fosfomycin alone. The role of rifampin in the treatment of E. faecalis isolates is discrepant and needs more studies.

YHMI: a web tool to identify histone modifications and histone/chromatin regulators from a gene list in yeast.

Post-translational modifications of histones (e.g. acetylation, methylation, phosphorylation and ubiquitination) play crucial roles in regulating gene expression by altering chromatin structures and creating docking sites for histone/chromatin regulators. However, the combination patterns of histone modifications, regulatory proteins and their corresponding target genes remain incompletely understood. Therefore, it is advantageous to have a tool for the enrichment/depletion analysis of histone modifications and histone/chromatin regulators from a gene list. Many ChIP-chip/ChIP-seq datasets of histone modifications and histone/chromatin regulators in yeast can be found in the literature. Knowing the needs and having the data motivate us to develop a web tool, called Yeast Histone Modifications Identifier (YHMI), which can identify the enriched/depleted histone modifications and the enriched histone/chromatin regulators from a list of yeast genes. Both tables and figures are provided to visualize the identification results. Finally, the high-quality and biological insight of the identification results are demonstrated by two case studies. We believe that YHMI is a valuable tool for yeast biologists to do epigenetics research.

Biofilm formation in erythromycin-resistant Staphylococcus aureus and the relationship with antimicrobial susceptibility and molecular characteristics.

PURPOSE: In this study, we aimed to investigate biofilm formation characteristics in clinical Staphylococcus aureus (S. aureus) isolates with erythromycin (ERY) resistance from China and further analyze their correlations with antimicrobial susceptibility and molecular characteristics. METHODOLOGY: A total of 276 clinical isolates of ERY-resistant S. aureus, including 142 methicillin-resistant S. aureus (MRSA) strains and 134 methicillin-susceptible S. aureus (MSSA) strains, were retrospectively collected in China. Biofilms were determined by crystal violet staining and ERY resistance genes (ermA, ermB and ermC) were detected by polymerase chain reaction. Inducible clindamycin resistance was examined by D test and multilocus sequence typing, and clonal complexes (CCs) based on housekeeping genes were further determined. RESULTS: The frequency of biofilm formation among ERY-resistant S. aureus was 40.9% (113/276) in total and no significant difference was found for the frequency of biofilm formation between ERY-resistant MRSA and ERY-resistant MSSA (44.4% vs 37.3%, P > 0.05). In ERY-resistant MRSA isolates, the frequency of biofilm formation in ermA-positive, gentamicin-resistant and ciprofloxacin-resistant isolates was higher than that in ermA-negative, gentamicin-sensitive and ciprofloxacin-sensitive isolates, respectively (63.9% vs 23.6%, P < 0.01; 60.3% vs 27.5%, P < 0.01; 65.2% vs 26.3%, P < 0.01). In addition, tetracycline resistance facilitated biofilm formation in both ERY-resistant MRSA and MSSA and the frequency of biofilm formation in CC239- or CC7S. aureus isolates with ERY resistance was significantly higher compared with that in CC59S. aureus (both P < 0.01). CONCLUSION: The ermA gene, and gentamicin, ciprofloxacin and tetracycline resistance facilitate biofilm formation in ERY-resistant MRSA isolates and, moreover, ERY-resistant S. aureus isolates with positive biofilm formation exhibited clonality clustering regarding CC239 and CC7.

YARG: A repository for arsenic-related genes in yeast.

Arsenic is a toxic metalloid. Moderate levels of arsenic exposure from drinking water can cause various human health problems such as skin lesions, circulatory disorders and cancers. Thus, arsenic toxicity is a key focus area for environmental and toxicological investigations. Many arsenic-related genes in yeast have been identified by experimental strategies such as phenotypic screening and transcriptional profiling. These identified arsenic-related genes are valuable information for studying arsenic toxicity. However, the literature about these identified arsenic-related genes is widely dispersed and cannot be easily acquired by researchers. This prompts us to develop YARG (Yeast Arsenic-Related Genes) database, which comprehensively collects 3396 arsenic-related genes in the literature. For each arsenic-related gene, the number and types of experimental evidence (phenotypic screening and/or transcriptional profiling) are provided. Users can use both search and browse modes to query arsenic-related genes in YARG. We used two case studies to show that YARG can return biologically meaningful arsenic-related information for the query gene(s). We believe that YARG is a useful resource for arsenic toxicity research. YARG is available at http://cosbi4.ee.ncku.edu.tw/YARG/.