Representative Time Points Research Articles

Viral epitope profiling of hematopoietic cell transplant recipients with respiratory syncytial virus upper respiratory tract and lower respiratory tract infections

Abstract Background The pre-fusion conformation of the F glycoprotein of respiratory syncytial virus (RSVpreF) has been the target of recent respiratory syncytial virus (RSV) therapies, including monoclonal antibodies for immunocompetent children and vaccines for the elderly. However, the utility of targeted therapies against RSVpreF is unknown in pediatric or adult hematopoietic cell transplant (HCT) recipients, and there are limited natural history studies characterizing humoral immune responses after RSV infection in HCT recipients. We utilized VirScan, a novel high-throughput profiling of antiviral antibody epitopes, to identify differences in global RSV-related epitope hits after an RSV infection. Methods We evaluated adult and pediatric allogeneic HCT recipients who acquired RSV from 12/2011 to 12/2019. Patients were categorized into 3 groups: upper respiratory tract infections (URTI) only, those who progressed from URTI to lower respiratory tract infections (progressors) and lower respiratory tract infections (LRTI) at diagnosis, and tested at baseline prior to infection (pre-RSV), at RSV URTI or LRTI diagnosis (RSV diagnosis), at LRTI progression (LRTI), 4-6 weeks after infection (Response 1) and >8 weeks after infection (Response 2), depending on sample availability. We interrogated the viral antibody repertoire using a serosurvey (VirScan) that detects immunoglobulin G responses to 206 viruses. VirScan metrics evaluated included: (1) number of RSV epitope hits, (2) log10 of the maximum RSV epitope binding signal (EBS), analogous to an antibody titer for each epitope, and (3) geometric mean of all RSV EBS. Kruskal-Wallis tests were used to compare groups for each VirScan metric at the different time points. Results A total of 129 samples from 39 patients were analyzed; 24 with URTI only, 8 progressors, and 7 with LRTI at diagnosis. At each time point, VirScan metrics were similar among the 3 patient groups (Table 1). Though patterns of VirScan metrics following RSV diagnosis varied by patient, most showed a modest increase in values (Figure 1). Conclusion In this preliminary study, we utilized the power of VirScan, a comprehensive serologic profiling tool of the entire human virome, to characterize RSV specific antibody changes after RSV infection in allogeneic HCT recipients. There were no differences in global RSV VirScan metrics between the 3 groups, suggesting that antibodies may not be the primary driver of viral infection attenuation in this population. Future analyses will focus on differences in individual epitope acquisition, including RSVpreF, between the 3 groups, and investigate possible interactions with humoral responses to other viruses. Figure 1. Longitudinal RSV VirScan metrics for each patient separated by 3 groups (top row: RSV URTI only; middle row: progressors from URTI to LRTI; bottom row: LRTI at diagnosis). Each line is an individual patient. The different panels show the changes in RSV related epitope hits (A), the log10 maximum epitope binding signal (EBS) among RSV epitopes (B), and the geometric mean of all the RSV related EBS (C). Open circles represent time points prior to RSV diagnosis, filled circles represent time points just prior to, on or after URTI, and asterisks represent time points just prior to, on, or after LRTI.

In vitro Antibacterial Effect Study of Plasma-Activated Saline on Mycobacterium Tuberculosis.

This study aimed to investigate the antibacterial effects of plasma-activated saline (PAS) on My-cobacterium tuberculosis (Mtb). We conducted a growth assay on 3 strains of Mtb and an antibiotic sensitivity test on 4 strains of Mtb. Both tests included groups treated with normal saline (NS), PAS, and hydrochloric acid (HCl). The test of antibiotic sensitivity consisted of parallel tests with two concentrations of bacteria suspension: 10-2 and 10-4. The selected antibiotics were rifampicin (RIF), isoniazid (INH), ethambutol (EMB), and streptomycin (SM). The number of bacteria was determined after one month of culture under different conditions. The Kruskal-Wallis test was used to analyze the differences in grouping factors at representative time points. The growth assay indicated that PAS significantly inhibited the growth of 3 strains of Mtb compared with NS and HCl treatment groups. Furthermore, except for the initial observation time point, the remaining three observation time points consistently demonstrate no significant differences between the NS group and the HCl group. The antibiotic sensitivity test of INH, SM, and RIF indicated that PAS could inhibit the growth of antibiotic-resistant Mtb, and the antibiotic sensitivity test of INH and SM with bacterial suspension concentration of 10-2 and SM with bacterial suspension concentration of 10-4 showed statistically different results. The antibiotic sensitivity test of EMB indicated that the growth of Mtb in PAS was slower than that in NS and HCl in both antibiotic-resistant and sensitive Mtb, but there was no statistical difference. The study indicates that PAS contains a significant amount of active substances and exhibits high oxidizability and an acidic pH state. The unique physicochemical properties of PAS significantly delayed the growth of Mtb, compared to the NS and the HCl. PAS not only inhibited the growth of drug-sensitive strains but also significantly enhanced the sensitivity of drug-resistant strains to anti-tuberculosis drugs, which may provide a new therapeutic strategy for the treatment of tuberculosis.

Thyroid-stimulating hormone levels are associated with estradiol levels and impact reproductive outcomes in preconceptionally euthyroid women undergoing their first IVF/ICSI cycles.

Controlled ovarian hyperstimulation (COH) has been reported to affect thyroid function; however, the impact of thyroid-stimulating hormone (TSH) levels during COH on embryo development and early reproductive outcomes has largely not been determined. Therefore, the aim of the present study was to investigate whether TSH levels are associated with COH and impact early reproductive outcomes in preconceptionally euthyroid women. This was a prospective cohort study. A total of 338 euthyroid women who underwent their first in vitro fertilization/intracytoplasmic sperm injection (IVF/ICSI) treatment using the gonadotropin releasing hormone agonist (GnRH-a) protocol were included. Samples were collected at different representative time points for TSH and estradiol measurements. TSH levels significantly increased with the administration of Gn and maintained this tendency until the trigger day. Basal TSH levels increased along with basal estradiol levels and remained stable when estradiol levels were higher than 150 pmol/L. On the trigger day, TSH levels changed with increasing estradiol levels in the high-normal basal TSH group but not in the low TSH group. TSH did not impact clinical pregnancy or early pregnancy loss after adjusting for age, stage or number of embryos. Serum TSH levels change significantly during COH and are associated with significant changes in estradiol levels. However, euthyroid women with high-normal TSH levels showed similar development potential for inseminated embryos and early reproductive outcomes compared to those with low TSH levels.

Identification of BMAL1-Regulated circadian genes in mouse liver and their potential association with hepatocellular carcinoma: Gys2 and Upp2 as promising candidates

Identification and functional analysis of key genes regulated by the circadian clock system will provide a comprehensive understanding of the underlying mechanisms through which circadian clock disruption impairs the health of living organisms. The initial phase involved bioinformatics analysis, drawing insights from three RNA-seq datasets (GSE184303, GSE114400, and GSE199061) derived from wild-type mouse liver tissues, which encompassed six distinct time points across a day. As expected, 536 overlapping genes exhibiting rhythmic expression patterns were identified. By intersecting these genes with differentially expressed genes (DEGs) originating from liver RNA-seq data at two representative time points (circadian time, CT: CT2 and CT14) in global Bmal1 knockout mice (Bmal1−/−), hepatocyte-specific Bmal1 knockout mice (L-Bmal1−/−), and their corresponding control groups, 80 genes potentially regulated by BMAL1 (referred to as BMAL1-regulated genes, BRGs) were identified. These genes were significantly enriched in glycolipid metabolism, immune response, and tumorigenesis pathways. Eight BRGs (Nr1d1, Cry1, Gys2, Homer2, Serpina6, Slc2a2, Nmrk1, and Upp2) were selected to validate their expression patterns in both control and L-Bmal1−/− mice livers over 24 h. Real-time quantitative polymerase chain reaction results demonstrated a comprehensive loss of rhythmic expression patterns in the eight selected BRGs in L-Bmal1−/− mice, in contrast to the discernible rhythmic patterns observed in the livers of control mice. Additionally, significant reductions in the expression levels of these selected BRGs, excluding Cry1, were also observed in L-Bmal1−/− mice livers. Chromatin immunoprecipitation (ChIP)-seq (GSE13505 and GSE39860) and JASPAR analyses validated the rhythmic binding of BMAL1 to the promoter and intron regions of these genes. Moreover, the progression of conditions, from basic steatosis to non-alcoholic fatty liver disease, and eventual malignancy, demonstrated a continuous gradual decline in Bmal1 transcripts in the human liver. Combining the aforementioned BRGs with DEGs derived from human liver cancer datasets identified Gys2 and Upp2 as potential node genes bridging the circadian clock system and hepatocellular carcinoma (HCC). In addition, CCK8 and wound healing assays demonstrated that the overexpression of human GYS2 and UPP2 proteins inhibited the proliferation and migration of HepG2 cells, accompanied by elevated expression of p53, a tumor suppressor protein. In summary, this study systematically identified rhythmic genes in the mouse liver, and a subset of circadian genes potentially regulated by BMAL1. Two circadian genes, Gys2 and Upp2, have been proposed and validated as potential candidates for advancing the prevention and treatment of HCC.

2712. Respiratory syncytial virus (RSV) pre-fusion F antibody titers in hematopoietic cell transplant recipients with upper versus lower respiratory tract infections

Abstract Background Humoral immune responses to respiratory syncytial virus (RSV) infection in hematopoietic cell transplant (HCT) recipients have not been well characterized. We measured RSV pre-fusion F antibody (RSVpreF) titers in HCT recipients in 3 groups: upper respiratory tract infections (URTI) only, lower respiratory tract infections (LRTI) only, and those who progressed from URTI to LRTIs (progressors). Methods We evaluated adult and pediatric allogeneic HCT recipients who acquired RSV from 12/2011 to 12/2019. Prospective and leftover clinical serum specimens were collected from the Fred Hutchinson Cancer Center. Samples included baseline prior to infection, at RSV URTI or LRTI diagnosis, and after infection (both 4-6 and >8 weeks). Quantitative RSVpreF titers were determined using the MesoScale Discovery electrochemiluminescence assay. We compared longitudinal log10 RSVpreF titers among groups using linear mixed effect models with restricted cubic splines. Results A total of 129 samples from 39 patients were analyzed; 24 with URTI only, 8 progressors, and 7 with LRTI only. The median age at infection was 45 years, but HCT recipients with LRTI only had an overall higher median age of 54 (range 23-63) (Table 1). Most had acute leukemia (n=18, 46%) and received peripheral blood stem cells (n=27, 69%) as their cell source (Table 1). Median RSVpreF values were similar across the 3 different groups at each timepoint (Table 2). Trajectories of RSVpreF titers over time for each patient trended towards an increase following RSV diagnosis in all groups (Figure 1a). Predicted curves in RSVpreF titers over time did not differ significantly by group (Figure 1b, p=0.38).Table 2.Summary of RSVpreF titers among allogeneic transplant recipients before and after a post-HCT RSV infection. RSVpreF titers were obtained utilizing Meso Scale Discovery chemiluminescence assays. (A) RSVpreF titers for allogeneic HCT recipients with RSV infection. Each line represents a patient. The top panel shows patients with RSV URTI only, the middle panel shows patients who progressed from URTI to LRTI, and the bottom panel shows patients who presented with RSV LRTI. Open circles represent time points prior to RSV diagnosis, filled circles represent time points just prior to, on or after URTI, and asterisks represent time points just prior to, on, or after LRTI. (B) RSVpreF titers plotted by days from RSV diagnosis. Open circles represent observed values and lines represent model predicted trajectories. Black circles and lines correspond to patients with RSV URTI only, blue corresponds to patients who progressed from RSV URTI to LRTI, and dark orange represents patient who presented with RSV LRTI at diagnosis. An outlying observation at day -53 was excluded. Conclusion Among allogeneic HCT recipients with RSV infection, RSVpreF titers increased in response to RSV infection, but did not differ among patients with URTI only, progressors, or those with LRTI only across pre and post infection time points. Our novel immunoassay results are similar to those from previously described RSV neutralization assays, but extended the observation period beyond 4 weeks after infection, revealing a modest increase in RSVpreF titers. Further characterization of RSV humoral immunity, including mucosal immunity, is needed. Disclosures chikara Ogimi, MD, bioMerieux Japan Ltd.: Honoraria|Horiba: Honoraria|Pfizer: Honoraria Janet A. Englund, MD, Ark Biopharma: Advisor/Consultant|AstraZeneca: Advisor/Consultant|AstraZeneca: Grant/Research Support|GlaxoSmithKline: Grant/Research Support|Meissa Vaccines: Advisor/Consultant|Merck: Grant/Research Support|Moderna: Advisor/Consultant|Moderna: Grant/Research Support|Pfizer: Advisor/Consultant|Pfizer: Grant/Research Support|Sanofi Pasteur: Advisor/Consultant Emily T. Martin, PhD, MPH, Merck: Grant/Research Support Michael J. Boeckh, MD PhD, Allovir: Advisor/Consultant|Amazon: Grant/Research Support|Ansun: Grant/Research Support|Merck: Advisor/Consultant|Merck: Grant/Research Support|Moderna: Advisor/Consultant|Symbio: Advisor/Consultant Alpana Waghmare, MD, Allovir: Grant/Research Support|Amazon: Grant/Research Support|Ansun Biopharma: Grant/Research Support|GlaxoKlineSmith/Vir: Grant/Research Support|Kyorin Pharmaceuticals: Advisor/Consultant|Pfizer: Grant/Research Support|Vir Biotechnology: Advisor/Consultant

E-Learning at-Risk Group Prediction Considering the Semester and Realistic Factors

This study focused on predicting at-risk groups of students at the Open University (OU), a UK university that offers distance-learning courses and adult education. The research was conducted by drawing on publicly available data provided by the Open University for the year 2013–2014. The semester’s time series was considered, and data from previous semesters were used to predict the current semester’s results. Each course was predicted separately so that the research reflected reality as closely as possible. Three different methods for selecting training data were listed. Since the at-risk prediction results needed to be provided to the instructor every week, four representative time points during the semester were chosen to assess the predictions. Furthermore, we used eight single and three integrated machine-learning algorithms to compare the prediction results. The results show that using the same semester code course data for training saved prediction calculation time and improved the prediction accuracy at all time points. In week 16, predictions using the algorithms with the voting classifier method showed higher prediction accuracy and were more stable than predictions using a single algorithm. The prediction accuracy of this model reached 81.2% for the midterm predictions and 84% for the end-of-semester predictions. Finally, the study used the Shapley additive explanation values to explore the main predictor variables of the prediction model.

Comparison of the colonization ability of Burkholderia strain B23 in the citrus rhizoplane and rhizosphere and assessment of the underlying mechanisms using full-length 16S rDNA amplicon and metatranscriptomic analyses.

The characterization of bacterial strains with efficient root colonization ability and the mechanisms responsible for their efficient colonization is critical for the identification and application of beneficial bacteria. In this study, we found that Burkholderia strain B23 exhibited a strong niche differentiation between the rhizosphere and rhizoplane (a niche with more abundant easy-to-use nutrients but stronger selective pressures compared with the tightly adjacent rhizosphere) when inoculated into the field-grown citrus trees. Full-length 16S rDNA amplicon analysis demonstrated that the relative abundance of B23 in the rhizoplane microbiome at 3, 5, and 9 days post-inoculation (dpi) was always higher than that at 1 dpi, whereas its relative abundance in the rhizosphere microbiome was decreased continuously, as demonstrated by a 3.18-fold decrease at 9 dpi compared to 1 dpi. Time-series comparative expression profiling of B23 between the rhizoplane and rhizosphere was performed at representative time points (1, 3, and 9 dpi) through metatranscriptomic analysis, and the results demonstrated that multiple genes involved in the uptake and utilization of easy-to-use carbohydrates and amino acids and those involved in metabolism, energy production, replication, and translation were upregulated in the rhizoplane compared with the rhizosphere at 1 dpi and 3 dpi. Several genes involved in resistance to plant- and microbial competitor-derived stresses exhibited higher expression activities in the rhizoplane compared with the rhizosphere. Furthermore, gene loci responsible for the biosynthesis of the key antifungal and antibacterial metabolites occidiofungin and ornibactin were induced, and their expression levels remained relatively stable from 3 dpi to 9 dpi in the rhizoplane but not in the rhizosphere. Collectively, our findings provide novel lights into the mechanisms underlying the root colonization of the inoculated bacterial strains and serve as a basis for the identification of strains with efficient colonization ability, thus contributing to the development of beneficial bacteria applications.

Comparative assessment of plasma cortisol and fecal corticoid metabolites (FCM) of Atlantic salmon (Salmo salar L.) subjected to acute- and long-term stress

Farmed Atlantic salmon (Salmo salar L.) are subject to a variety of stressors throughout production. Elevations of cortisol level in the blood are one of the major endocrine primary stress responses in vertebrates and are widely used as stress indicator. However, blood sampling is invasive and stressful procedures. Cortisol instantly released into the blood at sampling can easily interfere with the initial stress response of interest. Fecal corticoid metabolites (FCM) have been suggested for a less invasive assessment of stress in fish. In the present study, we evaluated stress responses by using plasma cortisol and FCM as stress indicators in Atlantic salmon. Atlantic salmon (with average weight 42 g) were exposed to 3 different stressors: 1) parr-smolt transformation; 2) infestation with salmon lice (Lepeophtheirus salmonis) for two-weeks; 3) infection with infectious salmon anaemia virus (ISAV) for four weeks. The results demonstrated that FCM levels correlated well with the plasma cortisol levels at single-point sampling during long-term stress (p < 0.05). Significant increases of both plasma cortisol and FCM were found two weeks after initiated 24 h light. Each tank was sampled twice, with around 40-min interval. The chasing during the first sampling was used as an acute stressor. The effects of sampling procedures on plasma cortisol and FCM levels are compared. Blood and feces were analyzed by an Enzyme-Linked Immunosorbent Assay (ELISA). Plasma cortisol and FCM levels increased within 40 min after fish perceived an acute stressor when the average weight of fish was below 100 g. Attenuated cortisol stress responses to acute stress were found after fish experiencing long-term stress. Collectively, FCM reflects changes in plasma cortisol and can be an alternative to analysis of plasma cortisol in Atlantic salmon, complementing evidence of using FCM as stress indicators for monitoring and improving fish welfare. The results highlight that standard sampling procedures and good experimental designs should be established to ensure robust and repeatable results in the study of stress based on the FCM measurement. Future study will investigate representative timepoints for FCM sampling, with particular attention to the types of stress and sizes of the fish.

Scaling-up and proteomic analysis reveals photosynthetic and metabolic insights toward prolonged H2 photoproduction in Chlamydomonas hpm91 mutant lacking proton gradient regulation 5 (PGR5)

Clean and sustainable H2 production is crucial to a carbon–neutral world. H2 generation by Chlamydomonas reinhardtii is an attractive approach for solar-H2 from H2O. However, it is currently not large-scalable because of lacking desirable strains with both optimal H2 productivity and sufficient knowledge of underlying molecular mechanism. We hereby carried out extensive and in-depth investigations of H2 photoproduction of hpm91 mutant lacking PGR5 (Proton Gradient Regulation 5) toward its up-scaling and fundamental mechanism issues. We show that hpm91 is at least 100-fold scalable (up to 10 L) with continuous H2 collection of 7287 ml H2/10L-HPBR in averagely 26 days under sulfur deprivation. Also, we show that hpm91 is robust and active during sustained H2 photoproduction, most likely due to decreased intracellular ROS relative to wild type. Moreover, we obtained quantitative proteomic profiles of wild type and hpm91 at four representing time points of H2 evolution, leading to 2229 and 1350 differentially expressed proteins, respectively. Compared to wild type, major proteome alterations of hpm91 include not only core subunits of photosystems and those related to anti-oxidative responses but also essential proteins in photosynthetic antenna, C/N metabolic balance, and sulfur assimilation toward both cysteine biosynthesis and sulfation of metabolites during sulfur-deprived H2 production. These results reveal not only new insights of cellular and molecular basis of enhanced H2 production in hpm91 but also provide additional candidate gene targets and modules for further genetic modifications and/or in artificial photosynthesis mimics toward basic and applied research aiming at advancing solar-H2 technology.

Torque Teno Virus Primary Infection Kinetics in Early Childhood.

Human torque teno viruses (TTVs) are a diverse group of small nonenveloped viruses with circular, single-stranded DNA genomes. These elusive anelloviruses are harbored in the blood stream of most humans and have thus been considered part of the normal flora. Whether the primary infection as a rule take(s) place before or after birth has been debated. The aim of our study was to determine the time of TTV primary infection and the viral load and strain variations during infancy and follow-up for up to 7 years. TTV DNAs were quantified in serial serum samples from 102 children by a pan-TTV quantitative PCR, and the amplicons from representative time points were cloned and sequenced to disclose the TTV strain diversity. We detected an unequivocal rise in TTV-DNA prevalence, from 39% at 4 months of age to 93% at 2 years; all children but one, 99%, became TTV-DNA positive before age 4 years. The TTV-DNA quantities ranged from 5 × 101 to 4 × 107 copies/mL, both within and between the children. In conclusion, TTV primary infections occur mainly after birth, and increase during the first two years with high intra- and interindividual variation in both DNA quantities and virus strains.

Mitochondrial TOM70 in Hyperoxia‐induced Acute Lung Injury

Study objectiveHyperoxia‐induced acute lung injury (HALI) leads to respiratory failure and mitochondrial dysfunction is a critical mediator of HALI. We previously reported Toll‐Like Receptor (TLR)4 prevents oxidant‐induced ALI by maintaining mitochondrial bioenergetics in mammalian lung endothelium (Ec). Translocase of Outer Mitochondrial Membrane (TOM) have recently been identified to play important roles in mitochondrial quality control, primarily in yeast [1]. TOM70 is an essential subunit of TOM and mediates uptake of newly synthesized proteins to mitochondria as well has putative anti‐oxidant and anti‐apoptotic properties. TOM70 appeared to parallel many of the cytoprotective signaling properties of TLR4 [2‐4]. Therefore, our hypothesis is TLR4 regulates TOM70 expression and TOM70 has mitochondrial and lung protective functions in HALI.MethodsC57BL/6J mice (6‐8 wks, both sex) were purchased from Jackson Laboratory, and Tlr4‐/‐ mice were bred at Duke Animal facilities (IACUC A160‐19‐07) [5]. For hyperoxia exposure, mice were placed in a Plexiglas chamber with 5L/min of 100% oxygen continuous flow with food and water ad libitum. We previously reported 72 h of continuous hyperoxia as a time of maximal lung injury [6], which we used as a representative time point. Bodyweight and survival were monitored daily. Bronchoalveolar lavage (BAL) were collected and BAL protein were determined by a BCA assay. Protein and RNA were extracted from lung tissues and expression was determined by western blot and qPCR, respectively. Mouse lung Ec (MLEC) were isolated from lung tissues and treated with control or Tom70siRNA for 48 h. Statistical differences were analyzed by a Student‐t test.ResultsTlr4‐/‐ MLEC decreased TOM70 and increased Parkin, PINK1, and MFN2 protein expression compared to WT. Tom70 siRNA effectively silenced Tom70 with &gt;75% efficiency at 48 h after transfection. Tom70 siRNA‐treated MLEC showed increased Mfn1, Mff, and Fis1mRNA expression, which are markers of mitochondrial fusion‐fission. Hyperoxia‐exposed mice showed marked weight loss and higher levels of BAL proteins compared to control, indicating increased permeability of the alveolar‐capillary membrane. Moreover, TOM70 protein expression was significantly decreased by hyperoxia in both male and female mice.ConclusionTLR4 deficiency decreased TOM70 expression and induced mitophagy and mitochondrial fusion, indicating that TOM70 is regulated by TLR4. Silencing TOM70 resulted in increased mitochondrial fusion and fission in MLEC, suggesting that TOM70 impacts mitochondrial quality control in mammalian, primary cells (MLEC) that are critical for lung airspace barrier function. Hyperoxia decreased TOM70 protein expression in mouse lungs, which may be a potential mechanism whereby hyperoxia leads to respiratory failure. Given that TOM70 maintains mitochondrial health, our data suggest that TOM70 may play a key role in the pathological mechanisms of HALI.1. Liu et al. (2021) bioRxiv, 2. Zhang et al. (2019) Antioxid Redox Signal, 3. Zhang et al. (2016) Antioxid Redox Signal, 4. Takyar et al. (2015) FASEB J. 5. Kim et al. (2019) Aging Cell. 6. Zhang et al. (2021) AJP Lung Cell Mol Physiol.

Experimental investigation of pore structure during high sodium coal ash sintering with and without calcium oxide additive through XCT technology

AbstractThis article investigates the evolution of morphology and pore structure during sintering at 1200°C for a high sodium coal ash, with and without using a calcium oxide additive. The effect of different proportions of calcium oxide additive was studied. The ratios of height and area of samples were calculated, and representative sintering time points of morphological changes were captured. Meanwhile, frequencies of diameter, pore numbers, and porosities were obtained by processing images. The maximum ratio of height and area reached at 15 min, 4 min, and 10 min for the sintering process of coal ash sample without additive, with 5% and 10% CaO, proving that CaO accelerates the sintering process. In comparison, the maximum porosities were 52.4%, 39.5%, and 47.0% for sintering samples after 10 min, 6 min, and 40 min. Besides, X‐ray diffraction analysis of the minerals of the condensed white matter was conducted.

Transcriptional Feedback Loops in the Caprine Circadian Clock System.

The circadian clock system is based on interlocked positive and negative transcriptional and translational feedback loops of core clock genes and their encoded proteins. The mammalian circadian clock system has been extensively investigated using mouse models, but has been poorly investigated in diurnal ruminants. In this study, goat embryonic fibroblasts (GEFs) were isolated and used as a cell model to elucidate the caprine circadian clock system. Real-time quantitative PCR analysis showed that several clock genes and clock-controlled genes were rhythmically expressed in GEFs over a 24 h period after dexamethasone stimulation. Immunofluorescence revealed that gBMAL1 and gNR1D1 proteins were expressed in GEFs, and western blotting analysis further verified that the proteins were expressed with circadian rhythmic changes. Diurnal changes in clock and clock-controlled gene expression at the mRNA and protein levels were also observed in goat liver and kidney tissues at two representative time points in vivo. Amino acid sequences and tertiary structures of goat BMAL1 and CLOCK proteins were found to be highly homologous to those in mice and humans. In addition, a set of goat representative clock gene orthologs and the promoter regions of two clock genes of goats and mice were cloned. Dual-luciferase reporter assays showed that gRORα could activate the promoter activity of the goat BMAL1, while gNR1D1 repressed it. The elevated pGL4.10-gNR1D1-Promoter-driven luciferase activity induced by mBMAL1/mCLOCK was much higher than that induced by gBMAL1/gCLOCK, and the addition of gCRY2 or mPER2 repressed it. Real-time bioluminescence assays revealed that the transcriptional activity of BMAL1 and NR1D1 in goats and mice exhibited rhythmic changes over a period of approximately 24 h in NIH3T3 cells or GEFs. Notably, the amplitudes of gBMAL1 and gNR1D1 promoter-driven luciferase oscillations in NIH3T3 cells were higher than those in GEFs, while mBMAL1 and mNR1D1 promoter-driven luciferase oscillations in NIH3T3 cells had the highest amplitude. In sum, transcriptional and translational loops of the mammalian circadian clock system were found to be broadly conserved in goats and not as robust as those found in mice, at least in the current experimental models. Further studies are warranted to elucidate the specific molecular mechanisms involved.

21st century gender trends of authorship in the British Journal of Oral and Maxillofacial Surgery

Despite forming the majority in undergraduate medicine and dentistry programmes, females remain under-represented in all surgical specialties, including oral and maxillofacial surgery (OMFS). Efforts are ongoing to promote female uptake in surgery. Research output is a key metric in measuring leadership and academic advancement within a specialty. The aim of this study was to assess female authorship in research published in the British Journal of Oral and Maxillofacial Surgery from 2000-2020 with a retrospective cohort study of original articles and reviews published in the journal from five different volumes of the journal (2000, 2005, 2010, 2015, 2020 as representative time points). The gender of the first and last authors were assessed for gender by first name recognition and verified with a web-search of the author using their stated institutional affiliation when this was not clear. Articles were excluded if the gender of one of the first or last author could not be determined. A total of 501 articles were included with the gender of 1002 authors analysed. 173 (17.3%) authors were female, and 828 (82.7%) were male. The most common author combination was male-male (70.3%) and the least common female-female (4.8%). The proportion of female authors increased over time from 9.3% in 2000 to 23.4% in 2020. Females remain under-represented within surgery, but female presence in academic output is increasing at a similar rate to the numbers of female uptake in the clinical workforce which is a promising sign and indicative of progress towards gender parity.

Identification of key proteins related to high-quality fiber in Upland cotton via proteomics analysis.

The dynamics of cotton fiber elongation and microfibirl deposition orientation were delineated; advanced ethylene synthesis and redox reaction homeostasis may be crucial for high-quality fiber formation. Fiber length, strength, and fineness determine the use and commercial value of cotton fiber, but their underlying molecular mechanisms remain obscure. We compared the dynamic change trajectories of length, diameter and microfibril orientation angle of the fibers produced by an introgression line SY6167 which generates high-quality fibers even better than Sea island cotton with those of the common-quality fibers from TM-1 across 5 to 30days post anthesis (DPA). The proteomes were profiled and compared at six representative time points using 2-DE and MS/MS. 14 proteins differentially expressed inside each of cotton line temporally and significantly different tween the two lines were identified. The dynamic change trajectories of fiber length and microfibril angle are close to "s" and reverse "s" growth curves, respectively. SY6167 and TM-1 fibers entered the logarithmic elongation phase simultaneously at 10 DPA, and SY6167 kept elongating logarithmically for 2 more days than TM-1. In parallel to logarithmic elongation, microfibril orientation angles dived sharply, and SY6167 declined faster for a shorter duration than TM-1. 53% of the identified proteins are related to redox homeostasis, and most of them are expressed at higher levels in SY6167 during logarithmic elongation. 1-Aminocyclopropane-1-Carboxylic Acid Oxidase (ACO) started to accumulate at 16 DPA in SY6167, and its encoding genes were highly expressed at this stage, with a much higher level than TM-1. These findings suggest high-quality fibers are associated with high expression of the proteins related to stress and redox homeostasis, the continuously elevated expression of ethylene synthesis ACO gene may play an essential role.

Back analysis of mechanical parameters of rock masses based on multi-point time-dependent monitoring data

The physical and mechanical parameters of rock masses are important factors that affect the numerical simulation and safety evaluation of geotechnical engineering. Due to the discontinuity and heterogeneity of rock mass, the parameters obtained by laboratory test and field test can not represent the actual situation. With the development of computer technology, the parameter identification method based on machine learning provides a new way to solve the above problems. Aiming at the rheological model based on thermodynamics with internal state variables, a back analysis method of mechanical parameters based on IAGA-BP algorithm is proposed. The sensitivity of deformation parameters, damage effect parameters and viscoplastic parameters involved in the model are analyzed using range analysis method. And the parameters which have great influence on the deformation of surrounding rock are selected to reduce the dimension of output layer. The displacement corresponding to the representative time point are selected as the input layer neuron. A numerical model is established to finely simulate the driving process of TBM, which is used to generate training samples and test samples. BP, GA-BP, PSO-BP, APSO-BP and SVM are also used to inverse the parameters of surrounding rock. Two evaluation metrics are used to evaluate the performance of above algorithms, including prediction accuracy and stability. The method is applied to inverse parameters of surrounding rock of a tunnel. The results show that the time-dependent deformation curve of surrounding rock obtained by BP neural network optimized by improved adaptive genetic algorithm (IAGA-BP) fits well with the monitoring curve (the average prediction error is 3.8%, and the prediction stability is 16.3%), which is much better than other algorithms.

The dynamic changes of glycogen molecular structure in Escherichia coli BL21(DE3)

Diurnal alteration of glycogen molecular structure has been identified in healthy mice. Recently, both fragile (disintegration in dimethyl sulfoxide) and stable (not disintegrating in DMSO) glycogen particles were found in Escherichia coli. However, how glycogen structure changes dynamically in E. coli is not clear. The question examined here is whether fragile, stable glycogen α particles occur in bacteria, following a similar pattern as in mice. In this study, we examine the dynamic changes of glycogen molecular structure over 24-h in E. coli BL21(DE3), using transmission electron microscopy, size exclusion chromatography and fluorophore-assisted carbohydrate electrophoresis at representative time points. It was found that glycogen structure was mainly fragile at the synthesis stage and largely stable during the degradation stage. qRT-PCR results indicated that balance of anabolic and catabolic gene expression levels in glycogen metabolism could be a key factor affecting the fragility of glycogen α particles in bacteria.

Tracking mesenchymal stem cells with Ir(III) complex-encapsulated nanospheres in cranium defect with postmenopausal osteoporosis

Osteoporosis (OP) is a significant public health problem with associated fragility fractures, thereby causing large bone defects and difficulty in self-repair. The introduction of human mesenchymal stem cells (hMSCs) is the most promising platform in bone tissue engineering for OP therapy, which induces less side effects than conventional medication. However, the safety and efficiency of the cell-based OP therapy requires the ability to monitor the cell's outcome and biodistribution after cell transplantation. Therefore, we designed an in vivo system to track hMSCs in real time and simultaneously attempted to obtain a significant therapeutic effect during the bone repair process. In this study, we synthesized Ir(III) complex, followed by encapsulation with biodegradable methoxy-poly(ethylene glycol) poly(lactic-co-glycolic acid) nanospheres through double emulsions strategy. The Ir(III) complex nanospheres did not affect hMSC proliferation, stemness, and differentiation and realized highly efficient and long-term cellular labeling for at least 25 days in vivo. The optimal transplantation conditions were also determined first by injecting a gradient number of labeled hMSCs percutaneously into the cranial defect of the nude mouse model. Next, we applied this method to ovariectomy-induced OP mice. Results showed long-term optical imaging with high fluorescence intensity and computed tomography (CT) scanning with significantly increased bone formation between the osteoporotic and sham-operated bones. During the tracking process, two mice from each group were sacrificed at two representative time points to examine the bony defect bridging via micro-CT morphometric analyses. Our data showed remarkable promise for efficient hMSC tracking and encouraging treatment in bioimaging-guided OP stem cell therapy.

Translation Initiation Site Profiling Reveals Widespread Synthesis of Non-AUG-Initiated Protein Isoforms in Yeast.

Genomic analyses in budding yeast have helped define the foundational principles of eukaryotic gene expression. However, in the absence of empirical methods for defining coding regions, these analyses have historically excluded specific classes of possible coding regions, such as those initiating at non-AUG start codons. Here, we applied an experimental approach to globally annotate translation initiation sites in yeast and identified 149 genes with alternative N-terminally extended protein isoforms initiating from near-cognate codons upstream of annotated AUG start codons. These isoforms are produced in concert with canonical isoforms and translated with high specificity, resulting from initiation at only a small subset of possible start codons. The non-AUG initiation driving their production is enriched during meiosis and induced by low eIF5A, which is seen in this context. These findings reveal widespread production of non-canonical protein isoforms and unexpected complexity to the rules by which even a simple eukaryotic genome is decoded.

Using CFD Modelling to Relate Pig Lying Locations to Environmental Variability in Finishing Pens

The purpose of this research was to determine which environmental factors within the pen space differ between the locations where pigs choose to lie and areas they avoid. Data on external weather conditions and the construction parameters for an existing commercial finishing pig building were input into a Dynamic Thermal (DT) model generating heat flow and surface temperature patterns in the structure and these were then input into a Computational Fluid Dynamics (CFD) model to generate data on the theoretical spatial patterns of temperature and air velocity within one room of this building on a specified day. The exact location of each pig in six selected pens within this room was taken from images from ceiling-mounted video cameras at four representative time points across the day. Using extracted air velocity and temperature data at the height of 0.64 m above the floor and a grid of approximately 600 mm to create a series of ‘cells’, the effective draught temperature (TED) was calculated from the models for each cell. Using a sequential regression model, the extent to which the actual lying locations of the pigs could be reliably predicted from the environmental conditions generated by model outputs and other pen factors was explored. The results showed that air velocity, TED and proximity to a solid pen partition (all significant at P &lt; 0.05) had significant predictive value and collectively explained 15.55% of the total explained deviation of 17.13%. When the presence of an adjacent pig was considered, results showed that lying next to an adjacent pig, TED, air velocity and temperature accounted for 53.9%, 1.3%, 1.5% and 0.5% of the deviation in lying patterns, respectively (all P &lt; 0.001). Thus, CFD model outputs could potentially provide the industry with a better understanding of which environmental drivers affect pigs’ lying location choice, even before a building is built and stocked.