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The COVID-19 pandemic, caused by the SARS-CoV-2 virus, has triggered a protracted global pandemic from 2019 to 2022, and posed a significant threat to human health. One of the non-structural proteins 3CL pro of SARS-CoV-2 is considered as a validated target for the development of inhibitors against the virus. Disulfiram has been reported as a covalent inhibitor of 3CL pro ; however, its structure lacks bonding site with active pockets of 3CL pro and its highly symmetric structure doesn't match well with the irregular cavity of the active center, limiting its therapeutic applications. To enhance their affinity for the 3CL pro target, in this study, two kinds of disulfiram derivatives, designed based on the reevaluation and optimization of disulfiram, have been synthesized through photoredox chemistry, and the novel carbamo(dithioperoxo)thioates 4g-m were found to display 5-17 folds potency against SARS-CoV-2 3CL pro compared to the parent disulfiram, with resulting half-maximal inhibitory concentration (IC 50 ) values ranging from 0.14-0.47 μM. Carbamo(dithioperoxo)thioates 4i containing a 4-hydroxy piperidine and a 4-trifluoromethyl phenyl ring, was identified as the most potent inhibitor to both 3CL pro (IC 50  = 0.14 μM) and PL pro (IC 50  = 0.04 μM). Furthermore, molecular dynamics simulations, binding free energy analysis and mass analysis were performed and provided insights on the stability, conformational behavior, and interactions of 4g with 3CL pro . The green synthetic methodology, the privileged carbamo(dithioperoxo)thioate scaffold, and the molecular mechanisms presented might serve as a useful system for the further discovery of highly potent inhibitors targeting SARS-CoV-2 3CL pro ., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Preventing nonspecific binding is essential for sensitive surface-based quantitative single-molecule microscopy. Here we report a much-simplified RainX-F127 (RF-127) surface with improved passivation. This surface achieves up to 100-fold less nonspecific binding from protein aggregates compared to commonly used polyethylene glycol (PEG) surfaces. The method is compatible with common single-molecule techniques including single-molecule pull-down (SiMPull), super-resolution imaging, antibody-binding screening and single exosome visualization. This method is also able to specifically detect alpha-synuclein (α-syn) and tau aggregates from a wide range of biofluids including human serum, brain extracts, cerebrospinal fluid (CSF) and saliva. The simplicity of this method further allows the functionalization of microplates for robot-assisted high-throughput single-molecule experiments. Overall, this simple but improved surface offers a versatile platform for quantitative single-molecule microscopy without the need for specialized equipment or personnel.

Objectives: Differences in clinical adverse outcomes (CAO) based on different intestinal stricturing definitions in Crohn's disease (CD) are poorly documented. This study aims to compare CAO between radiological strictures (RS) and endoscopic strictures (ES) in ileal CD and explore the significance of upstream dilatation in RS., Methods: This retrospective double-center study included 199 patients (derivation cohort, n = 157; validation cohort, n = 42) with bowel strictures who simultaneously underwent endoscopic and radiologic examinations. RS was defined as a luminal narrowing with wall thickening relative to the normal gut on cross-sectional imaging (group 1 (G1)), which further divided into G1a (without upstream dilatation) and G1b (with upstream dilatation). ES was defined as an endoscopic non-passable stricture (group 2 (G2)). Strictures met the definitions of RS (with or without upstream dilatation) and ES were categorized as group 3 (G3). CAO referred to stricture-related surgery or penetrating disease., Results: In the derivation cohort, G1b (93.3%) had the highest CAO occurrence rate, followed by G3 (32.6%), G1a (3.2%), and G2 (0%) (p < 0.0001); the same order was found in the validation cohort. The CAO-free survival time was significantly different among the four groups (p < 0.0001). Upstream dilatation (hazard ratio, 1.126) was a risk factor for predicting CAO in RS. Furthermore, when upstream dilatation was added to diagnose RS, 17.6% of high-risk strictures were neglected., Conclusions: CAO differs significantly between RS and ES, and clinicians should pay more attention to strictures in G1b and G3. Upstream dilatation has an important impact on the clinical outcome of RS but may not be an essential factor for RS diagnosis., Clinical Relevance Statement: This study explored the definition of intestinal stricture with the greatest significance for the clinical diagnosis and prognosis of patients with CD, and consequently provided effective auxiliary information for clinicians to formulate strategies for the treatment of CD intestinal strictures., Key Points: • The retrospective double-center study showed that clinical adverse outcome is different between radiological strictures and endoscopic strictures in CD. • Upstream dilatation has an important impact on the clinical outcome of radiological strictures but may not be an essential factor for diagnosis of radiological strictures. • Radiological stricture with upstream dilatation and simultaneous radiological and endoscopic stricture were at increased risk for clinical adverse outcomes; thus, closer monitoring should be considered., (© 2023. The Author(s), under exclusive licence to European Society of Radiology.)

Background: Dengue virus (DENV) can be divided into four serotypes-DENV-1, DENV-2, DENV-3, and DENV-4. In humans, infection leads to dengue fever (DF), dengue hemorrhagic fever, and dengue shock syndrome, both widely prevalent in tropical and subtropical regions. In 2019, a severe outbreak of DF occurred in Xishuangbanna, Yunnan province. Objective: To investigate the etiology and genotype of the causative agents of this severe dengue outbreak in Xishuangbanna. Methods: Between October and November 2019, the sera of patients clinically diagnosed with DF were collected in the first People's Hospital of Xishuangbanna. RNA was extracted from the sera and amplified by RT-PCR with flavivirus primers. Flavivirus-positive sera were then used to inoculate Aedes albopictus cells (C6/36); viral RNA was extracted from these cells, amplified, and sequenced with DENV E gene-specific primers. Sequence splicing and nucleotide homology genetic evolution analysis were carried out by biological software (DNAStar). Unique mutations in the E genes of isolated DENV were analyzed by SWISS-MODEL and PyMOL. Results: Of the 60 samples collected from DF patients, 39 tested positively with flavivirus primers. The DENV was isolated from 25 of the 39 positive seras, of which 20 showed cytopathic effects (CPE) and 5 were no CPE. In these 25 isolated nucleic acids, 21 strains of DENV-1, 3 strains of DENV-2, and 1 strain of DENV-3 were identified according to the sequence of E protein. In the four unique mutations (D52, Y149, L312, T386), D52 and Y149 in the E protein of DENV-1 were predicted to be exposed on the surface of the prefusion conformation. Conclusion: The 2019 outbreak of DF in Xishuangbanna area of Yunnan Province consists of at least three serotypes of DENV-1, DENV-2, and DENV-3, and the sources of these virus strains are of mixed and complicated origin.

Giardia duodenum (G. duodenalis) can cause giardiasis and infect a variety of hosts. So far, there have been no detailed data regarding the positive rate of G. duodenalis in sheep and goats in China. Here, a systematic literature review was carried out to investigate the epidemiology of G. duodenalis in sheep and goats in China. To perform the meta-analysis, the databases CNKI, VIP, WanFang, PubMed, Web of science and ScienceDirect were employed for screening studies related to the prevalence of G. duodenalis in sheep and goats in China. The total prevalence of G. duodenalis in sheep and goats was estimated to be 7.00% (95% CI: 4.00-10.00). In the age subgroup, the prevalence of G. duodenalis in sheep and goats of >12 months (11.29%; 95% CI: 8.08-14.97) was higher than that in sheep and goats of ≤12 months (7.57%; 95% CI: 3.95-12.24). An analysis based on seasons showed that the prevalence of G. duodenalis in sheep and goats was higher in summer (11.90%; 95% CI: 0.50-35.05) than that in other seasons. The prevalence of G. duodenalis in sheep and goats after 2016 was 8.57% (95% CI: 5.34-11.79), which was higher than others. The highest prevalence of G. duodenalis in sheep and goats was 13.06% (95% CI: 6.26-19.86) recorded in Southwestern China. The prevalence of Giardia infection in sheep (7.28%; 95% CI: 2.30-14.73) was higher than that in goats (5.43%; 95% CI: 2.73-8.98). The NOAA's National Center for Environmental Information (https://gis.ncdc.noaa.gov/maps/ncei/cdo/monthly) was used to extract relevant geoclimatic data (latitude, longitude, elevation, temperature, precipitation, humidity, and climate). By analyzing the data of each subgroup, it was shown that region, genetype, and climate were potential risk factors for giardiasis prevalence in sheep and goats. Based on the analysis of common factors and geographical factors, it is recommended to strengthen effective management measures (e.g. ventilation and disinfection in warm and humid areas) and formulate relevant policies according to local conditions. Breeders should strengthen the detection of G. duodenalis in sheep and goats, customize corresponding control measures according to the diet and living habits of sheep and goats, and strengthen the protection of sheep and lamb calves, so as to reduce the incidence rate and reduce the economic loss of China's animal husbandry., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

Background: The gut microbiota plays an essential role in maintaining gut homeostasis and improving performance, with the composition of microbial communities visibly differing across different laying stages in hens and significantly correlating with egg production. To gain further insights into the association between microbial community characteristics and laying periods in Hy-Line variety brown and Isa brown laying hens, we conducted a 16S rRNA amplicon sequencing survey., Results: Our result revealed the diversity of bacteria in the early laying period was commonly higher than peak, and in Hy-Line variety brown laying hens were generally higher than Isa brown. Principal coordinate analysis (PCoA) and permutational multivariate analysis of variance (PERMANOVA) revealed that the structure and composition of the gut microbiota of laying hens exhibited significant differences among different groups. Phylum Firmicutes, Bacteroidota, Proteobacteria, and Fusobacteriota were found that dominant in the host's feces. Therein, the abundance of Fusobacteriota was higher in the peak period than in the early period, while the abundance of Cyanobacteria in the early period was higher in two breeds of hens. Furthermore, random forest based on machine learning showed that there were several distinctly abundant genera, which can be used as potential biomarkers to differentiate the different groups of laying periods and breeds. In addition, the prediction of biological function indicated the existing discrepancy in microbial function among the microbiota of four groups., Conclusions: Our findings offer new insights into the bacterial diversity and intestinal flora composition of different strains of laying hens during various laying periods, contributing significantly to the improvement of production performance and the prevention of chicken diseases., (© 2023. The Author(s).)

Malus plants are frequently devastated by the apple rust caused by Gymnosporangium yamadae Miyabe. When rust occurs, most Malus spp. and cultivars produce yellow spots, which are more severe, whereas a few cultivars accumulate anthocyanins around rust spots, forming red spots that inhibit the expansion of the affected area and might confer rust resistance. Inoculation experiments showed that Malus spp. with red spots had a significantly lower rust severity. Compared with M. micromalus , M. 'Profusion', with red spots, accumulated more anthocyanins. Anthocyanins exhibited concentration-dependent antifungal activity against G. yamadae by inhibiting teliospores germination. Morphological observations and the leakage of teliospores intracellular contents evidenced that anthocyanins destroyed cell integrity. Transcriptome data of anthocyanins-treated teliospores showed that differentially expressed genes were enriched in cell wall and membrane metabolism-related pathways. Obvious cell atrophy in periodical cells and aeciospores was observed at the rust spots of M. 'Profusion'. Moreover, WSC , RLM1 , and PMA1 in the cell wall and membrane metabolic pathways were progressively downregulated with increasing anthocyanins content, both in the in vitro treatment and in Malus spp. Our results suggest that anthocyanins play an anti-rust role by downregulating the expression of WSC , RLM1 , and PMA1 to destroy the cell integrity of G. yamadae ., Competing Interests: The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Wang, An, Guo, Wang, Shang, Chen, Liu, Meng, Zhang, Wei and Li.)

Background: More than half of patients with Crohn's disease (CD) require at least one surgery for symptom management; however, approximately half of the patients may experience postoperative anastomotic recurrence (PAR)., Objectives: This study aims to develop and validate a preoperative computed tomography enterography (CTE)-based radiomics signature to predict early PAR in CD., Design: A total of 186 patients with CD (training cohort, n = 134; test cohort, n = 52) who underwent preoperative CTE and surgery between January 2014 and June 2020 were included in this retrospective multi-centre study., Methods: 106 radiomic features were initially extracted from intestinal lesions and peri-intestinal mesenteric fat, respectively; significant radiomic features were selected from them and then used to develop intestinal or mesenteric radiomics signatures, using the least absolute shrinkage and selection operator and a Cox regression model. A radiomics-based nomogram incorporating these signatures with clinical-radiological factors was created for comparison with a model based on clinical-radiological features alone., Results: 68 of 134 patients in training cohort and 16 of 52 patients in test cohort suffered from PAR. The intestinal radiomic signature (hazard ratio [HR]: 2.17; 95% confidence interval [CI]: 1.32-3.58; P = 0.002) and mesenteric radiomic signature (HR: 2.19; 95% CI: 1.14-4.19; P = 0.018) were independent risk factors for PAR in the training cohort as per a multivariate analysis. The radiomics-based nomogram (C-index: 0.710; 95% CI: 0.672-0.748) yielded superior predictive performance than the clinical-radiological model (C-index, 0.607; 95% CI: 0.582-0.632) in the test cohort. Decision curve analysis demonstrated that the radiomics-based nomogram outperformed the clinical-radiological model in terms of clinical usefulness., Conclusions: Preoperative mesenteric and intestinal CTE radiomics signatures are potential non-invasive predictors of PAR in postoperative patients with CD., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

DNA gyrase is an essential nucleoprotein motor present in all bacteria and is a major target for antibiotic treatment of Mycobacterium tuberculosis (MTB) infection. Gyrase hydrolyzes ATP to add negative supercoils to DNA using a strand passage mechanism that has been investigated using biophysical and biochemical approaches. To analyze the dynamics of substeps leading to strand passage, single-molecule rotor bead tracking (RBT) has been used previously to follow real-time supercoiling and conformational transitions in Escherichia coli (EC) gyrase. However, RBT has not yet been applied to gyrase from other pathogenically relevant bacteria, and it is not known whether substeps are conserved across evolutionarily distant species. Here, we compare gyrase supercoiling dynamics between two evolutionarily distant bacterial species, MTB and EC. We used RBT to measure supercoiling rates, processivities, and the geometries and transition kinetics of conformational states of purified gyrase proteins in complex with DNA. Our results show that E. coli and MTB gyrases are both processive, with the MTB enzyme displaying velocities ∼5.5× slower than the EC enzyme. Compared with EC gyrase, MTB gyrase also more readily populates an intermediate state with DNA chirally wrapped around the enzyme, in both the presence and absence of ATP. Our substep measurements reveal common features in conformational states of EC and MTB gyrases interacting with DNA but also suggest differences in populations and transition rates that may reflect distinct cellular needs between these two species., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Introduction: Despite Toxoplasma gondii infection leading to dysbiosis and enteritis, the function of gut microbiota in toxoplasmosis has not been explored., Methods: Here, shotgun metagenomics was employed to characterize the composition and function of mouse microbial community during acute and chronic T. gondii infection, respectively., Results: The results revealed that the diversity of gut bacteria was decreased immediately after T. gondii infection, and was increased with the duration of infection. In addition, T. gondii infection led to gut microbiota dysbiosis both in acute and chronic infection periods. Therein, several signatures, including depression of Firmicutes to Bacteroidetes ratio and infection-enriched Proteobacteria, were observed in the chronic period, which may contribute to aggravated gut inflammation and disease severity. Functional analysis showed that a large amount of Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways and carbohydrate-active enzymes (CAZy) family displayed distinct variation in abundance between infected and healthy mice. The lipopolysaccharide biosynthesis related pathways were activated in the chronic infection period, which might lead to immune system imbalance and involve in intestinal inflammation. Moreover, microbial and functional spectrums were more disordered in chronic than acute infection periods, thus implying gut microbiota was more likely to participate in disease process in the chronically infected mice, even exacerbated immunologic derangement and disease progression., Discussion: Our data indicate that the gut microbiota plays a potentially important role in protecting mice from T. gondii infection, and contributes to better understand the association between gut microbiota and toxoplasmosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Meng, Wei, Guo, Chen, Wang, Geng, Yang, Jiang and Zhang.)

The fungal community, also known as mycobiota, plays pivotal roles in host nutrition and metabolism and has potential to cause disease. However, knowledge of the gut fungal structure in Caprinae is quite limited. In this study, the composition and diversity of the gut mycobiota of Caprinae animals from different geographical locations (Anhui, Jilin, Guangxi, Shandong, Shanxi, and Tibet) were comprehensively characterized by analyzing the internal transcribed spacer 2 (ITS-2) sequences of the fungal community. The results showed that Ascomycota and Basidiomycota were the dominant phyla, which, respectively, accounted for 90.86 to 95.27% and 2.58 to 7.62% of sequences in samples from each region. Nonetheless, the structure of the gut mycobiota was largely different in Caprinae animals in the different provinces. Therein, Sporormiaceae and Thelebolaceae were the dominant fungal families in the samples from Tibet, whereas their abundance was generally low in other regions. The intestinal diversity of individuals from Guangxi was higher than that in other regions. In addition, there were 114 differential genera among all regions. Finally, the co-occurrence network revealed 285 significant correlations in cross-family pairs in the guts of Caprinae animals, which contained 149 positive and 136 negative relationships, with 96 bacterial and 86 fungal participants at the family level. This study has improved the understanding of the mycobiota of ruminants and provided support for the improvement in animal health and productivity. IMPORTANCE In this study, we elucidated and analyzed the structure of the gut mycobiota of Caprinae animals from different regions. This study revealed differences in the structure of the gut mycobiota among Caprinae animals from different geographical environments. Based on previous findings, correlations between fungal and bacterial communities were analyzed. This study adds to previous research that has expanded the present understanding of the gut microbiome of Caprinae animals.

Most microbiome studies regarding the ruminant digestive tract have focused on the rumen microbiota, whereas only a few studies were performed on investigating the gut microbiota of ruminants, which limits our understanding of this important component. Herein, the gut microbiota of 30 Caprinae animals (sheep and goats) from six provinces in China was characterized using ultradeep (>100 Gbp per sample) metagenome shotgun sequencing. An inventory of Caprinae gut microbial species containing 5,046 metagenomic assembly genomes (MAGs) was constructed. Particularly, 2,530 of the genomes belonged to uncultured candidate species. These genomes largely expanded the genomic repository of the current microbes in the Caprinae gut. Several enzymes and biosynthetic gene clusters encoded by these Caprinae gut species were identified. In summary, our study extends the gut microbiota characteristics of Caprinae and provides a basis for future studies on animal production and animal health. IMPORTANCE We constructed a microbiota catalog containing 5,046 MAGs from Caprinae gut from six regions of China. Most of the MAGs do not overlap known databases and appear to be potentially new species. We also characterized the functional spectrum of these MAGs and analyzed the differences between different regions. Our study enriches the understanding of taxonomic, functional, and metabolic diversity of Caprinae gut microbiota. We are confident that the manuscript will be of utmost interest to a wide range of readers and be widely applied in future research.

Toxoplasmosis is an important zoonotic parasitic disease caused by Toxoplasma gondii ( T. gondii ). However, the functions of circRNAs and miRNAs in response to T. gondii infection in the livers of mice at acute and chronic stages remain unknown. Here, high-throughput RNA sequencing was performed for detecting the expression of circRNAs and miRNAs in livers of mice infected with 20 T. gondii cysts at the acute and chronic stages, in order to understand the potential molecular mechanisms underlying hepatic toxoplasmosis. Overall, 265 and 97 differentially expressed (DE) circRNAs were found in livers at the acute and chronic infection stages in comparison with controls, respectively. In addition, 171 and 77 DEmiRNAs were found in livers at the acute and chronic infection stages, respectively. Functional annotation showed that some immunity-related Gene ontology terms, such as "positive regulation of cytokine production", "regulation of T cell activation", and "immune receptor activity", were enriched at the two infection stages. Moreover, the pathways "Valine, leucine, and isoleucine degradation", "Fatty acid metabolism", and "Glycine, serine, and threonine metabolism" were involved in liver disease. Remarkably, DEcircRNA 6:124519352|124575359 was significantly correlated with DEmiRNAs mmu-miR-146a-5p and mmu-miR-150-5p in the network that was associated with liver immunity and pathogenesis of disease. This study revealed that the expression profiling of circRNAs in the livers was changed after T. gondii infection, and improved our understanding of the transcriptomic landscape of hepatic toxoplasmosis in mice., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Zou, Meng, Wei, Gu, Chen, Geng, Yang, Zhang and Cao.)

Echinococcosis is a foodborne parasitic zoonosis caused by the larvae of Echinococcus . This disease can affect goats and other mammals. In this study, a systematic review and meta-analysis for echinococcosis in global goats were performed based on the following five databases (China National Knowledge Infrastructure [CNKI], VIP Chinese Journal Database, Wanfang Data, PubMed, and ScienceDirect). In total, 108,197 samples were collected. The global prevalence of echinococcosis in goats was identified to be 10.85% (3217/108,197). The prevalence of echinococcosis in goats was 6.16% (1369/22,208) and 13.27% (874/5932) in South America and Africa, respectively. The prevalence of echinococcosis in goats before 2010 (9.76%; 112/713) was significantly higher than that from 2010 to 2014 (1.44%; 45/32,145) or after 2014 (2.95%; 154/3889). The prevalence of echinococcosis in goats aged <12 months (4.48%; 70/2911) was higher than that in goats aged ≥12 months (2.88%; 36/819). We also investigated the effects of geographical factors and climates on the prevalence of echinococcosis in goats. The results showed that the prevalence of echinococcosis was higher in the areas with high altitude and cold climate. This meta-analysis indicated that echinococcosis was ubiquitous in goats. Thus, we should improve the feeding conditions for goats, and strengthen the control measures of echinococcosis epidemic in goats, with the aims of reducing the economic losses of animal husbandry and providing protection for humans in the aspects of food security and health.

The diversity of bacteria and fungi in the gut microbiota of commercial broilers that raised in cages from hatch to the end of the production cycle were examined by an analysis of 3,592 and 3,899 amplicon sequence variants (ASVs), respectively. More than 90% sequences in bacterial communities were related to Firmicutes and Proteobacteria. More than 90% sequences in fungal communities were related to Ascomycota, Basidiomycota, and Glomeromycota. A statistical analysis of the microbiota composition succession showed that age was one of the main factors affecting the intestinal microbial communities of broilers. The increasingly complex community succession of transient microbiota occurred along with an increase of age. This dynamic change was observed to be similar between bacteria and fungi. The gut microbiota had a special structure in the first 3 d after birth of broiler. The microbiota structure was quite stable in the period of rapid skeletal growth (d 14-21), and then changed significantly in the period of rapid gaining weight (d 35-42), thus indicating the composition of gut microbiota in broilers had unique structures at different developmental stages. We observed that several bacteria and fungi occupied key functions in the gut microbiota of broilers, suggesting that the gut homeostasis of broilers might be affected by losses of bacteria and fungi via altering interactions between microbiota. This study aimed to provide a data basis for manipulating the microbiota at different developmental stages, in order to improve production and the intestinal health of broilers., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Soluble aggregates of the microtubule-associated protein tau have been challenging to assemble and characterize, despite their important role in the development of tauopathies. We found that sequential hyperphosphorylation by protein kinase A in conjugation with either glycogen synthase kinase 3β or stress activated protein kinase 4 enabled recombinant wild-type tau of isoform 0N4R to spontaneously polymerize into small amorphous aggregates in vitro. We employed tandem mass spectrometry to determine the phosphorylation sites, high-resolution native mass spectrometry to measure the degree of phosphorylation, and super-resolution microscopy and electron microscopy to characterize the morphology of aggregates formed. Functionally, compared with the unmodified aggregates, which require heparin induction to assemble, these self-assembled hyperphosphorylated tau aggregates more efficiently disrupt membrane bilayers and induce Toll-like receptor 4-dependent responses in human macrophages. Together, our results demonstrate that hyperphosphorylated tau aggregates are potentially damaging to cells, suggesting a mechanism for how hyperphosphorylation could drive neuroinflammation in tauopathies., (© 2022. The Author(s).)

Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy has led to unprecedented results to date in relapsed/refractory (R/R) diffuse large B-cell lymphoma (DLBCL), yet its clinical application in elderly patients with R/R DLBCL remains somewhat limited. In this study, a total of 31 R/R DLBCL patients older than 65 years of age were enrolled and received humanized anti-CD19 CAR T-cell therapy. Patients were stratified into a fit, unfit, or frail group according to the comprehensive geriatric assessment (CGA). The fit group had a higher objective response (OR) rate (ORR) and complete response (CR) rate than that of the unfit/frail group, but there was no difference in the part response (PR) rate between the groups. The unfit/frail group was more likely to experience AEs than the fit group. The peak proportion of anti-CD19 CAR T-cells in the fit group was significantly higher than that of the unfit/frail group. The CGA can be used to effectively predict the treatment response, adverse events, and long-term survival.

The "Spring-red-leaf" crabapple cultivar has young red leaves and mature green leaves. However, the mechanism of anthocyanin biosynthesis in crabapple leaves in spring remains unknown. In this study, Illumina RNA sequencing (RNA-Seq) was performed on Malus 'Radiant' leaf tissues in different stages of development. Twenty-two genes in the anthocyanin biosynthesis pathway and 44 MYB transcription factors (TFs) were significantly enriched among differentially expressed genes (DEGs). Three R2R3-MYB TFs in subgroup 22 of the MYB TF family, MrMYB44-like1 , MrMYB44-like2 , and MrMYB44-like3 , were highly expressed in green leaves according to RNA-Seq and quantitative real-time quantitative PCR results. Their expression levels were negatively correlated with anthocyanin content. In transient assays, overexpression of MrMYB44-like1 , MrMYB44-like2 , or MrMYB44-like3 inhibited anthocyanin accumulation and reduced pigment in leaf disks of M . 'Radiant' and fruit peels of M. domestica 'Fuji.' When the conserved region of the three MrMYB44-like s was silenced, the anthocyanin biosynthesis pathway was activated and pigments increased in both tissues. Moreover, bimolecular fluorescence complementation assays showed MrMYB44-like s interacted with MrWRKY6 to form protein complexes that regulated anthocyanin biosynthesis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Meng, Wei, Chi, Qiao, Zhou, Wang, Wang and Li.)

Casuarina cunninghamiana Miq. naturally occurs in eastern Australia from New South Wales to north Queensland. After being introduced to China, it has become an important tree species of ecological shelter plantations in coastal areas of southern China. In this study, the complete chloroplast (cp) genome of C. cunninghamiana was sequenced and analyzed based on the Illumina NovaSeq 6000 platform. The cp genome of C. cunninghamiana was found to be 15,6129 bp in length, including a large single copy (LSC) region of 86,200 bp and a small single copy (SSC) region of 18,457 bp, which were separated by two inverted repeats (IRs) of 25,736 bp. The cp genome contains 132 genes, consisting of 87 protein-coding genes, 37 tRNA genes, and eight rRNA genes. The overall GC content of the cp genome was 36.34%. The phylogenetic analyses indicated that C. cunninghamiana was closely related to C. glauca and C. equisetifolia and clustered with 4 Betulaceae species., Competing Interests: No potential conflict of interest was reported by the author(s)., (© 2021 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.)

Objective: To compare the activity difference of the high affinity humanized CD19 chimeric antigen receptor (CAR)-T cells and murine CD19 CAR-T cells. Methods: Peripheral venous blood T cells from 8 healthy volunteers were collected and infected with humanized and murine CD19 CAR lentivirus. Human and murine CD19 CAR-T cells were prepared and cell proliferation was detected by cell counting kit-8 (CCK-8) method. The cytotoxicity of CD3(+) T cells, humanized and murine CD19 CAR-T cells to NALM-6 cells was detected by lactate dehydrogenase assay. Thirty BAL B/c nude mice transplanted with NALM-6 cells were randomly divided into 3 groups with 10 mice in each group and injected humanized CD19 CAR-T cells, mouse CD19 CAR-T cells and control CD3(+) T cell via tail vein, respectively. The proportion of NALM-6 cells in peripheral blood and the proportion of CD19 CAR-T cells in T cells from the vein of the inner canthus were detected by flow cytometry. The overall survival of BAL B/c nude mice was observed. Results: The proliferation of mouse and humanized CD19 CAR-T cells were (68.50±0.93)% and (80.63±1.41)%, respectively ( t =20.353, P <0.001) after cultured in vitro for 24 hours, and were (91.38±1.41)% and (148.13±1.25)%, respectively ( t =85.364, P <0.001) after cultured for 48 hours. When the effect to target ratio was 1∶1, there was no difference between the humanized and murine CD19 CAR-T cell group after co-culture for 24 hours ( P =0.169), while the killing activity of humanized CD19 CAR-T cells against NALM-6 cells was higher than that of murine CD19 CAR-T cells ( P <0.01) after 48 hours of co-culture. When the effect to target ratio was 4∶1, the cytotoxicity of humanized CD19 CAR-T cells against NALM-6 cells was higher than that of murine CD19 CAR-T cells in co-culture for 24 and 48 hours ( P <0.01). On the seventh day of CD19 CAR-T cell therapy, the proportion of NALM-6 cells in the peripheral blood of BAL B/c nude mice decreased to the lowest level in the humanized CD19 CAR-T cell group and the murine CD19 CAR-T cell group. After 21 days, the proportion of NALM-6 cells in the murine CD19 CAR-T cell group was higher than that in the humanized CD19 CAR-T cell group ( P (21 d )=0.001, P (28 d )<0.001, P (35 d )<0.001). The proportion of humanized and murine CD19 CAR-T cells in the peripheral blood reached the peaks after 7 days of therapy, and the proportion of humanized CD19 CAR-T cells was higher than that of murine CAR-T cells ( P (7 d )=0.002). The CD19 CAR-T cells disappeared in the peripheral blood in the murine CD19 CAR-T cell group after 14 days of therapy, while in the humanized CD19 CAR-T cell group it disappeared after 21 days of therapy. The median survival of BAL B/c nude mice in the murine CD19 CAR-T cell group and the humanized CD19 CAR-T cell group was 42 days and 63 days, respectively ( χ (2)=15.382, P <0.001). Conclusions: High affinity humanized CD19 CAR-T cells have stronger proliferation, higher cytotoxicity and longer survival time compared with those of murine CD19 CAR-T cells. The results indicate that the clinical efficacy of humanized CD19 CAR-T cells would be better than that of murine CD19 CAR-T cells.

Objective: To investigate the effect and involved mechanism of RSL3 on ferroptosis action in acute leukemia cells MOLM13 and its drug-resistant cells., Methods: After MOLM13 treated with RSL3, CCK-8 assay was performed to detect cell viability, flow cytometry was used to detect the reactive oxygen species (ROS) level of the cells, RT-qPCR and Western blot were used to detect the expression of glutathione peroxidase 4 (GPX4). After MOLM13/IDA and MOLM13/Ara-C, the drug-resistant cell lines were constructed, the ferroptosis induced by RSL3 was observed. Bone marrow samples were collected from patients with acute monocytic leukemia. RT-qPCR and Western blot were performed to detect the expression of related genes and proteins involved in ferroptosis pathway., Results: RSL3 significantly inhibited the cell viability of MOLM13 and increased the intracellular ROS level, which were partially reversed by ferrostatin-1. The mRNA and protein expression of GPX4 decreased in MOLM13 treated with RSL3. RSL3 inhibited the viability of MOLM13/IDA and MOLM13/Ara-C cells more strongly than that of non-drug resistant cells, also increased the intracellular ROS level . The cytotoxic effects were partially reversed by ferrostatin-1. The mRNA and protein expressions of GPX4 in MOLM13/IDA and MOLM13/Ara-C cells were higher than those in non-drug resistant cells. The mRNA and protein levels of GPX4 in bone marrow of relapsed/refractory acute mononuclear leukemia patients were higher than those of ordinary acute mononuclear leukemia patients., Conclusion: RSL3 can induce non-drug resistant cells MOLM13 ferroptosis by inhibiting GPX4 activity. MOLM13/IDA and MOLM13/Ara-C are more sensitive to RSL3 compared with non-drug resistant cells MOLM13, which may be caused by the differences in GPX4 expression. The expressions of GPX4 mRNA and protein in relapsed/refractory acute mononuclear leukemia are higher than those in ordinary acute mononuclear leukemia.

Materials with near-infrared (NIR) persistent luminescence (PersL) and NIR-to-NIR photostimulated luminescence (PSL) properties are attractive platforms for photonic energy harvesting and release. In this work, we develop Mg 2 SnO 4 :Cr as a broadband NIR PersL and NIR-to-NIR PSL material (luminescence maxima at ∼800 nm) and reveal the origin of the PersL and PSL properties. The material has an inverse spinel structure with the Mg 2+ and Sn 4+ disorder at the Wyckoff 16d site based on the Rietveld refinement. Cr K-edge X-ray absorption near-edge structure (XANES) spectra uncover that the doped Cr ions have a +3 valence state and occupy the disordered (Mg,Sn) site with octahedral coordination. The disorder results in multiple Cr 3+ centers, and the broadband luminescence originates from the 4 T 2 ( 4 F) → 4 A 2 transition of Cr 3+ at sites with intermediate crystal field strength. The distribution of trap depths is continuous according to the analysis of thermoluminescence (TL) spectra using the initial rising method, which relates to the random distribution of Mg 2+ and Sn 4+ at the second coordination sphere of the Cr 3+ centers rather than the oxygen-related defects. Stimulating the material with a NIR laser, the NIR PersL gets significantly enhanced due to a PSL process. The broadband PersL and PSL are detectable beyond 100 h and have good tissue penetrability and therefore the developed Mg 2 SnO 4 :Cr 3+ has potential in applications of optical information storage/reading and autofluorescence-free bioimaging. Finally, three crystal and electronic structure factors are proposed for screening new Cr 3+ -activated PersL and PSL materials.

Objective: To investigate the characteristics and cytotoxicity in vitro of the residual leukemia cells in the culture system that caused the accidental transfer of CD19 chimeric antigen receptor (CAR) into leukemia cells during the preparation of autologous CD19 CAR-T cells of relapsed/refractory B-cell acute lymphoblastic leukemia. Methods: ①Peripheral blood mononuclear cells (PBMC) of 30 patients with relapsed/refractory B-cell acute lymphoblastic anemia (R/R B-ALL) who accepted CD19 CAR-T cell therapy and six healthy volunteers were collected. ②The residual leukemia cells were analyzed by flow cytometry in the system after the PBMCs of R/R B-ALL patients were sorted by CD3 magnetic beads. ③ CD3(+) T cells from patients and healthy volunteers were transfected with CD19 CAR and CD22 CAR lentivirus to prepare CD19 CAR-T and CD22 CAR-T cells. ④The Nalm-6 cell line was resuscitated and the Nalm-6 cells with CD19 CAR lentivirus were transfected to prepare CD19 CAR-Nalm-6 cells. The patient's primary ALL cells were transfected with CD19 CAR lentivirus at the same time. ⑤The transfection rates were analyzed by flow cytometer, the cell proliferation was analyzed by the CCK-8 method, and the cell-killing activities were detected by the lactate dehydrogenase method. Results: ① Among the 30 R/R B-ALL patients who received CD19 CAR-T cell therapy, two patients had 2.04% and 3.32% residual leukemia cells in CD3(+) T cells. After 4 days in culture, the residual leukemia cells disappeared and could not be detected by a flow cytometer with prolonged cultivation in vitro. ② The proliferation of CD19 CAR-Nalm-6 cells was higher than that of the Nalm-6 cells. ③ The killing activity of the CD19 CAR-T cells on Nalm-6 cells was higher than that of the CD19 CAR-Nalm6 cells at a target ratio of 1∶1 on 24, 48, 72 h, respectively. The cytotoxicity of CD22 CAR-T cells on CD19 CAR-Nalm-6 cells was significantly higher than that of CD19 CAR-T cells. ④ The cytotoxicity of CD22 CAR-T alone on CD19 CAR-Nalm-6 cells was higher than that of CD19 CAR-T combined with CD22 CAR-T at the same target ratio. Conclusion: The residual leukemia cells in the culture system in the preparation of CD19 CAR-T cells may lead to the introduction of CD19 CAR into leukemia cells and results in the failure of the CD19 CAR-T cell therapy. Detecting the residual leukemia cells in the culture system via flow cytometry before transfection with CD19 CAR lentivirus is needed. Thus, CD22 CAR-T cell therapy could be used as one of the salvage treatments.

The flower color of many horticultural plants fades from red to white during the development stages, affecting ornamental value. We selected Malus halliana , a popular ornamental species, and analyzed the mechanisms of flower color fading using RNA sequencing. Forty-seven genes related to anthocyanin biosynthesis and two genes related to anthocyanin transport were identified; the expression of most of these genes declined dramatically with flower color fading, consistent with the change in the anthocyanin content. A number of transcription factors that might participate in anthocyanin biosynthesis were selected and analyzed. A phylogenetic tree was used to identify the key transcription factor. Using this approach, we identified MhMYB10 as directly regulating anthocyanin biosynthesis. MhMYB10 expression was strongly downregulated during flower development and was significantly positively related to the expression of anthocyanin biosynthetic genes and anthocyanin content in diverse varieties of Malus . To analyze the methylation level during flower development, the MhMYB10 promoter sequence was divided into 12 regions. The methylation levels of the R2 and R8 increased significantly as flower color faded and were inversely related to MhMYB10 expression and anthocyanin content. Therefore, we deduce that the increasing methylation activities of these two regions repressed MhMYB10 expression., (Copyright © 2020 Han, Yin, Zhao, Sun, Meng, Zhou, Shen, Li and Zhang.)

Objective: To investigate the factors influencing the efficacy of CD19 chimeric antigen receptor T (CAR-T) cells in the treatment of patients with relapsed refractory B cell lymphoma and to provide evidence for further improvement of CAR-T efficacy. Methods: A total of 34 patients with relapsed and refractory B-cell lymphoma were recruited from the Department of Hematology of Tianjin First Central Hospital from February 2017 to January 2019. All patients received CD19 CAR-T cell therapy. These patients were evaluated for efficacy, factors with poor efficacyand adverse effects. Results: The overall response rate was 58.8% (20/34) and the complete remission rate was 41.2% (14/34) after infusion of CD19 CAR-T cells in 34 patients with relapsed refractory B cell lymphoma. According to the efficacy of CAR-T cells, patients were divided into two groups, 20 in the effective group and 14 in the poorly effective group. The median am ount of CD19 CAR-T cell infusions in these two groups were 8.6 (5.0-12.7)×10(6)/kg and 9.7 (5.8-15.0) × 10(6)/kg, respectively, and the difference was not statistically significant ( P= 0.654). The percentage of CD19 CAR-T cells in the effective group and the poorly treated group was 10.28% (3.92%-44.16%) and 4.05% (0.92%-28.63%), respectively.The effective group had a higher proportion of CAR-T cells than the poorly treated group, but the difference was not statistically significant ( P= 0.371).The presence of massive mass was an unfavorable factor affecting the efficacy of CD19 CAR-T cells and the difference was statistically significant ( P= 0.001). Logistic regression multivariate analysis showed that the characteristics of massive tumors were still independent prognostic factors for poor efficacy of CD19 CAR-T cells ( P= 0.005, OR= 0.039). Of all 34 patients, there were 70.6% (24/34) who showed varying degrees of adverse reactions after the infusion of CD19 CAR-T cells, mainly cytokines release syndrome (CRS). The median time of occurrence of fever was on the third day after infusion (0-11th) day. 16 patients were with grade 1 CRS, 7 with grade 2, and 1 with grade 3. After glucocorticoids and support treatment, they all showed improvements. Conclusions: CD19 CAR-T cell therapy has achieved a certain effect in CD19(+)B cell lymphoma, but has poor efficacy on some patients. Large mass tumors may be an adverse factors to CAR-T cell treatment.

Anti-CD19 chimeric antigen receptor T cell (CAR-T) therapy has changed the typical outcomes of relapsed/refractory B-cell leukemia and lymphoma. However, treatment effectiveness for patients with relapsed/refractory B-cell non-Hodgkin lymphoma has been less satisfactory compared with patients with B-cell acute lymphoblastic leukemia. The present study described a case of refractory follicular lymphoma. A high expression of programmed cell death 1 (PD-1) was measured on CD3 + T cells (80.90%) in peripheral blood samples obtained from the patient enrolled in this study, indicating that treatment with autologous CAR-T 19 cell therapy may not be successful. Therefore, a therapy regimen consisting of CAR-T 19 cells in combination with a reduced dose of nivolumab (1.5 mg/kg) for PD-1 blockade was used. A low dose of PD-1 blockade therapy was used to reduce the adverse effects associated with the combination of a PD-1 inhibitor and CAR-T 19 cells. This salvage therapy resulted in remission that lasted for >10 months., (Copyright: © Wang et al.)

Objective: To observe the changes of PD-1 expression, mRNA level and cytotoxic activity of CD19 CAR-T cells during the culture process of CAR-T cells. Methods: The peripheral blood T cells of 6 lymphoma patients with high expression of PD-1 and 6 healthy volunteers were the source of CAR-T cells. The expression of PD-1 was analyzed by flow cytometry. The mRNA level of PD-1 was analyzed by PCR. The cell proliferation was analyzed by CCK-8 assay. The cytotoxicity was analyzed by LDH assay. Results: ①The transfection efficiency of high PD-1 expression T cells and healthy volunteer T cells were as the same ( P >0.05) . ②The cell proliferation capacity of CD19 CAR-T cells from high PD-1 expression T cells or healthy volunteer T cells, with or without PD-1 inhibitor were as the same ( P >0.05) . ③The cytotoxicity to lymphoma cells of high PD-1 expression T cells and CAR-T cells were lower than that of these two T cells combined with PD-1 inhibitor and the CAR-T cells from healthy volunteer T cells ( P <0.001) . There was no difference of the cytotoxicity between the CAR-T cells from high PD-1 expression T cells combined with PD-1 inhibitor and the CAR-T cells from healthy volunteer ( P >0.05) . ④There was no difference of the expression of PD-1 in all CAR-T cell groups during the culture process ( P >0.05) . There was no difference of mRNA level of PD-1 in all groups during the culture process ( P >0.05) . ⑤The PD-1 expression of CAR-T cells increased by the time of culture after contacting with lymphoma cells ( P <0.001) . The PD-1 inhibitors could antagonize this effect. There was no difference of mRNA level of PD-1 in all groups after contacting with lymphoma cells ( P >0.05) . Conclusion: The PD-1 expression of CAR-T cells from high PD-1 expression T cells increased by the time of culture after contacting with lymphoma cells. However, the mRNA level of PD-1 of all groups did not change, even if PD-1 inhibitor was applied.

Objective: To retrospectively analyze the efficacy and safety of modified cell infusion method in reducing the incidence of febrile non-hemolytic transfusion reaction (FNHTR). Methods: A total of 69 patients were enrolled in the clinical trial of CD(19) chimeric antigen receptor T (CAR-T) cell treatment from February 2017 to October 2018. Study group received the modified cell infusion method, that 1×10(6) CAR-T cells were re-suspended in 2 mg human serum albumin with total volume of 20 ml and injected intravenously. The control group was intravenously administrated with CAR-T cell in 100 ml normal saline. The incidence of FNHTR, cytokine releasing syndrome (CRS) grade, cytokine level and efficacy were compared. Results: (1)The incidence of FNHTR in the study group was 21.1%, significantly lower than that in the control group (71%)( P =0.000). (2)There was no statistical difference in cell proliferation between the study group and the control group on day 4, 7, 14 and 21 after CAR-T cell infusion ( P =10.223, 3.254, 5.551, 7.605). (3)There was no statistical difference in CRS grading between the study group and the control group ( P =0.767). There was no statistical difference in the levels of interleukin 2 receptor (IL-2R), IL-6, tumor necrosis factor (TNF)-α between the two groups. (4)The C-reaction protein (CRP) level of the study group was lower than that of the control group on day 4 and 7 ( P =0.026, 0.007). (5)There was no statistical difference of response rates in acute lymphocytic leukemia (ALL) and non-Hodgkin lymphoma (NHL) patients between the two groups ( P (ALL)=0.842; P (NHL)=0.866). Conclusion: The modified cell infusion method in CD(19) CAR-T cell treatment reduces the incidence of treatment-related FNHTR. It does not affect the proliferation of CAR-T cells in vivo , the grading of CRS and the response rates.

Objective: To investigate the efficacy and safety of CD19 chimeric antigen receptor T (CAR-T) lymphocytes for the treatment of B cell lymphoma. Methods: A total of 22 patients with B-cell lymphoma from February 1, 2017 to July 1, 2018 were reviewed to evaluate the efficacy and adverse reactions of CD19 CAR-T. Results: Of 22 patients with B-cell lymphoma received CD19 CAR-T cells, the median dose of CAR-T cells was 7.2 (2.0-12.0) ×10 6 /kg. Nine of 12 cases of relapse refractory patients were overall response. Complete remission (CR) occurred in 2 of 12 patients, partial remission (PR) in 7 of 12 patients. The overall response in minor residual disease positive (MRD) group was 8 of 10 patients. CD19 CAR-T cells proliferated in vivo and were detectable in the blood of patients. The peak timepoints of CAR-T cells proliferated in the relapsed refractory and MRD positive groups were 12 (5-19) and 4.5 (1-12) days after treatment respectively, and among peripheral blood cells, CAR-T cells accounted for 10.10% (3.55%-24.74%) and 4.02% (2.23%-28.60%) of T lymphocytes respectively. The MRD positive patients achieved sustained remissions during a median follow-up of 8 months (rang 3-18 months) . None of all the patients relapsed during a median follow-up time of 10 months (3-18 months) . However, 7 PR responders of the relapsed refractory patients maintained a good condition for 1.5-6.0 months. One patient bridged to hematopoietic stem cell transplantation, another one sustained remission for 12 months. Cytokine-release syndrome (CRS) occurred in 14 patients with grade 1-2 CRS in MRD positive group and grade 3 CRS in relapsed refractory group. Conclusions: CAR-T cell therapy not only played a role in the rescue treatment of relapsed and refractory patients, but also produced a surprising effect in the consolidation and maintenance of B-cell lymphoma. CD19 CAR-T cells might be more effective in the treatment of MRD positive B-cell lymphoma patients than in the refractory or relapsed cases. High response rate was observed with fewer adverse reactions.

Objective: To observe the effect of electroacupuncture (EA) intervention on hyperplastic mammary glands, serum estradiol (E 2 ) and progesterone (P) contents, estrogen receptor alpha (ERα) and progesterone receptor (PR) expression of breast tissues in mammary gland hyperplasia (MGH) rats, so as to reveal its mechanisms underlying improvement of MGH., Methods: Sixty female SD rats were randomly divided into control, model, EA, EA＋ovariectomy(OVX) and EA＋sham-OVX groups ( n ＝12 in each). The MGH model was established by injection of Estradiol Benzoate injection (0.5 mg/kg, once daily for 25 d) and P injection (0.5 mg/kg, once daily for 5 d after estradiol injection) into the medial hind-leg muscle. After model establishment, bilateral OVX was performed for rats of the EA＋OVX group, and sham OVX (only exposure of ovaries) was performed for rats of the EA＋sham OVX group. EA (2 Hz, 2 mA) was applied to bilateral "Tianzong"(SI 11), "Ganshu"(BL 18), "Zusanli" (ST 36) for point group A, and "Wuyi"(ST 15), "Hegu"(LI 4), "Danzhong" (CV 17) for point group B for 20 min, once daily, 5 days a week for 4 weeks. The two acupoint groups were used alternately. The height of the rats' nipples (the 2 nd pairs) were measured. Serum E 2 and P levels were assayed by ELISA. The expression of ERα and PR in mammary tissues was detected by immunofluorescence staining and Western blot, separately., Results: The height of nipples, serum E 2 content, and mammary ERα protein expression level were significantly higher in the model group than in the control group ( P <0.01), while serum P content and PR expression were remarkably lower in the model group than in the control group ( P <0.01). Following EA intervention, the height of nipples, serum E 2 content and mammary ERα protein expression level in both EA and EA＋sham OVX groups, and ERα expression in the EA＋OVX group were considerably lower in comparison with those of the model group( P <0.01), and serum P content and PR expression in the 3 EA groups were significantly higher than those in the model group ( P <0.01， P <0.05). But there were no significant changes in the nipple height and serum E 2 levels in the EA＋OVX group relevant to the model group ( P >0.05), suggesting an important role of the ovary in producing EA's effect., Conclusion: EA intervention can improve hyperplastic mammary glands by down-regulating serum E 2 content and mammary ERα protein expression, and up-regulating serum P content and PR expression in MGH rats, which has a close relation with the intact ovaries.

Objective: To Evaluation the effect of PD-1 inhibitor Nivolumab on the proliferation and cytotoxicity of anti-CD19 chimeric antigen receptor T cells (CD19-CAR-T) in vitro . Methods: Five patients with high PD-1 expression in peripheral blood and five healthy volunteers were selected. These peripheral blood mononuclear cells were used as the source of T cells to prepare CD19-CAR-T cells. Different doses (72, 36, 18 μg/ml) of Nivolumab was added on day 8 to the culture medium. Patient T cells incubated with 72 μg/ml Nivolumab and CD19-CAR-T cells of healthy volunteers were used as controls. CCK-8, lactate dehydrogenase (LDH) cytotoxicity assay and ELASA were used to detect the proliferation capacity, the specific cytotoxicity and the inflammatory factor secretion. Results: ①T cells from patients with high expression of PD-1 as the source of CD19-CAR-T cells did not affect transfection rate compared with that of healthy volunteers [(32.80±7.22)% vs (35.10±5.84)%, t =-0.554, P =0.593]. ②Incubation of CD19-CAR-T cells with 72 μg/ml Nivolumab did not affect CD19-CAR-T cell proliferation, but its cytotoxicity was significantly higher than that of CD19-CAR-T cells alone or patients' T cells +72 μg/ml Nivolumab (all P <0.001), there was no significant difference in the killing activity between the 72 μg/ml and 36 μg/ml Nivolumab treated CD19-CAR-T cells on Pfeiffer cells ( P =0.281, 0.267, respectively), and they were all higher than those of 18 μg/ml Nivolumab treated CD19-CAR-T cells (all P <0.001). ③Different doses of PD-1 inhibitor Nivolumab combined with CD19-CAR-T cells does not affect the secretion of IFN-γ and IFN-α (all P >0.05). Conclusion: Combination of 36 μg/ml PD-1 inhibitor and CD19-CAR-T cells could reduce the drug toxicity and enhance the cytotoxicity.

Propofol, an intravenous anesthetic agent widely used in clinical practice, is the preferred anesthetic for asthmatic patients. This study was designed to determine the protective effect and underlying mechanisms of propofol on airway inflammation in a mast cell-dependent mouse model of allergic asthma. Mice were sensitized by ovalbumin (OVA) without alum and challenged with OVA three times. Propofol was given intraperitoneally 0.5 h prior to OVA challenge. The inflammatory cell count and production of cytokines in the bronchoalveolar lavage fluid (BALF) were detected. The changes of lung histology and key molecules of the toll-like receptor 4 (TLR4)/reactive oxygen species (ROS)/NF-κB signaling pathway were also measured. The results showed that propofol significantly decreased the number of eosinophils and the levels of IL-4, IL-5, IL-6, IL-13, and TNF-α in BALF. Furthermore, propofol significantly attenuated airway inflammation, as characterized by fewer infiltrating inflammatory cells and decreased mucus production and goblet cell hyperplasia. Meanwhile, the expression of TLR4, and its downstream signaling adaptor molecules--myeloid differentiation factor 88 (MyD88) and NF-κB, were inhibited by propofol. The hydrogen peroxide and methane dicarboxylic aldehyde levels were decreased by propofol, and the superoxide dismutase activity was increased in propofol treatment group. These findings indicate that propofol may attenuate airway inflammation by inhibiting the TLR4/MyD88/ROS/NF-κB signaling pathway in a mast cell-dependent mouse model of allergic asthma.

Objective: To evaluate and compare the clinical efficacy and safety of recombinant human thrombopoietin(rhTPO) and recombinant human interleukin11(rhIL-11) for the treatment of chemotherapy-induced thrombocytopenia in adult acute myeloid leukaemia patients., Methods: Total of 96 adult acute myeloid leukaemia patients were divided into 3 groups according to randomized controlled method: rhTPO group, rhIL-11 group and control group, 32 cases in each group. The patients in rhTPO group and rhIL-11 received rhTPO of 15000 IU/d and rhIL-11 of 1.5 mg/d, respectively after the standard combined chemotherapy within 24 hours, and patients in control group, received nothing drugs to promote thrombocyte recovery. And rhTPO and rhIL-11 should be stopped when the Plt≥100× 10 9 /L. After chemotherapy, the platelet recovery degree, duration of Plt<50× 10 9 /L, ≥50× 10 9 /L and ≥100× 10 9 /L, the count of infusion thrombocytes, and incidence of adverse reactions all were compared., Results: The duration of Plt<50× 10 9 /L was obviously less than that in control group(P<0.01). The duration of rhIL-11 was less than that in control group, but there was no statistical significance(P>0.05). As compared with that in control group, the Plt count in rhTPO and rhIL-11 groups can faster increase to Plt≥50× 10 9 /L (P<0.01, P<0.05), among them the Plt count in rhTPO group faster increase, but there was no statistical signiticance. As compared with that in control group, the Plt count in rhTPO group and rhIL-11 group can increase to Plt≥100× 10 9 /L (P<0.01), the Plt count in rhTPO group was more obviously increase than that in rhIL-11 group(P<0.05). The count of infusion Plt in rhTPO and rhIL-11 groups was lese than that in control group(P<0.01, P<0.05), and the count of infusion Plt in rhTPO group was less than that in rhIL-11 group(P<0.05). After using rhTPO and rhIL-11, the adverse reactions, such as low fever, induration of injection site, athralgia, nausea and vomiting occured in rhTPO group and rhIL-11 group, but all can be tolerated., Conclusion: Both rhTPO and rhIL-11 can reduce the duration of thrombocytopenia and the amount of infused thrombocyte, promote platelet recovery in the patients with acute myeloid leukaemia after chemotherapy, to decreae the risk of bleeding, and reduce incidence of adverse reactions, both of them can be tolerated by patients, and rhTPO is more advantage than rhIL-11, worthy of clinical popularization and application.

The adverse effects of iron overload have raised more concerns as a growing number of studies reported its association with immune disorders. This study aimed to investigate alterations in the immune system by iron overload in patients with myelodysplastic syndrome (MDS) and an iron-overloaded mouse model. The peripheral blood from patients was harvested to test the effect of iron overload on the subsets of T lymphocytes, and the level of reactive oxygen species (ROS) was also evaluated. The data showed that iron-overloaded patients had a lower percentage of CD3 + T cells and disrupted T cell subsets, concomitant with higher ROS level in lymphocytes. In order to explore the mechanism, male C57Bl/6 mice were intraperitoneally injected with iron dextran at a dose of 250 mg/kg every 3 days for 4 weeks to establish an iron-overloaded mouse model and the blood of each mouse was collected for the analysis of the T lymphocyte subsets and T cell apoptosis. The results showed that iron overload could reduce the percentage of CD3 + T cells and the ratio of Th1/Th2 and Tc1/Tc2 but increase the percentage of regulatory T (Treg) cells and the ratio of CD4/CD8. We also found that iron overload induced the apoptosis of T lymphocytes and increased its ROS level. Furthermore, these effects could be partially recovered after treating with antioxidant N-acetyl-L-cysteine (NAC) or iron chelator deferasirox (DFX). Taken together, these observations indicated that iron overload could selectively affect peripheral T lymphocytes and induce an impaired cellular immunity by increasing ROS level.

What Is Known and Objective: Numerous population pharmacokinetic studies of theophylline have been conducted in paediatric and adult patients. The purpose of this review was to summarize the published studies concerning population pharmacokinetics of theophylline in patients of different ages and discuss factors that might cause the large variability in the pharmacokinetics of theophylline., Methods: A literature search was conducted in PubMed using the following keywords: 'theophylline', 'population pharmacokinetic(s)' and 'nonlinear mixed effect model'. Additionally, the relevant references listed in the retrieved articles were manually reviewed. All of the studies that reported the population pharmacokinetics of theophylline in humans were included in this review. However, articles were excluded if they were not written in English., Results and Discussion: Sixteen articles were included in this review. Among them, 11 were conducted on paediatric patients, and five were conducted on adults. A one-compartment model with first-order elimination was employed in most of the included articles. A nonlinear mixed effect modelling approach (NONMEM) was the most commonly used software to develop a population pharmacokinetic model. Body weight and age (post-conceptional age and post-natal age) were the most important factors associated with the clearance (CL) of theophylline in paediatric patients. Body weight (ideal body weight and lean body mass), age and smoking status were most frequently used to estimate the CL of theophylline in adults. The median (range) estimate values of CL for paediatric and adult patients were 0·062 (0·0056-0·0949) L/h/kg and 0·053 (0·0493-0·0517) L/h/kg, respectively. The median values of the interindividual variability of CL were 33·5% in adults and 25·8% in paediatric patients. The mean values of the residual variability were 21% in paediatric patients and 14·3% in adults., What Is New and Conclusion: This review concludes that body weight and age were the most important factors associated with the clearance of theophylline in paediatric patients. Body weight, age and smoking were most frequently used to estimate the clearance of theophylline in adults. Future studies are warranted to detect the influence of new factors, such as cytochrome P450 (CYP) 1A2 gene polymorphisms, on the pharmacokinetics of theophylline because some pharmacokinetic variability was not fully explained., (© 2016 John Wiley & Sons Ltd.)

Aim: To investigate whether plasma miRNAs targeting CYP3A4/5 have an impact on the variance of pharmacokinetics of clopidogrel., Materials & Methods: The contribution of 13 miRNAs to the CYP3A4/5 gene expression and activity was investigated in 55 liver tissues. The association between plasma miRNAs targeting CYP3A4/5 mRNA and clopidogrel pharmacokinetics was analyzed in 31 patients with coronary heart disease who received 300 mg loading dose of clopidogrel., Results: Among 13 miRNAs, miR-142 was accounting for 12.2% (p = 0.002) CYP3A4 mRNA variance and 9.4% (p = 0.005) CYP3A5 mRNA variance, respectively. Plasma miR-142 was negatively associated with H4 Cmax (r = -0.5269; p = 0.0040) and associated with H4 AUC0-4h (r = -0.4986; p = 0.0069) after 300 mg loading dose of clopidogrel in coronary heart disease patients., Conclusion: miR-142 could account for a part of missing heritability of CYP3A4/5 functionality related to clopidogrel activation.

Objective: To investigate the effects of iron overload on apoptosis and function of splenic CD8+ T cells in mice., Methods: Forty C57BL/6 mice were randomly divided into control groups, Iron overload (IO), IO+NAC and IO+DFX groups. The iron overload model was established by intraperitoneal injection of iron dextran, and saline was injected as the control. The levels of intracellular reactive oxygen species (ROS) and labile iron pool (LIP) were analyzed by measuring the mean fluorescence intensity (MFI) of 2-7 dichlorofluorescein (DCF) or calcein. The ratio of CD8+ T cells and the levels of IFN-γ, TNF-α, Granzyme-B, and perforin in CD8+ T cells were detected by flow cytometry. The CD8+ T cell apoptosis was determined by flow cytometry with Annexin V/PI double staining. Real-time PCR was used to detect the expression of IFN-γ, TNF-α, Granzyme-B, perforin, BCL-2, and bax at mRNA level in CD8+ T cells., Results: Iron overload was found by spleen iron staining and flow cytometry. The level of intracellular ROS in iron overload (IO) groups was higher than that of the control groups (P<0.01). The percentage of CD8+ T cells in spleen from mice with IO was lower than that in control groups (P<0.05). The expression of IFN-γ and Granzyme-B in CD8+ T cells in IO group were lower than that in control group, the expression of IFN-γ and Granzyme-B at mRNA level in CD8+ T cells was lower than that of control group (P<0.05). CD8+ T cell apoptosis in iron overload group was significantly higher than that in control groups (P<0.01); the expression of BCL-2 at mRNA level was lower than that in control group, but the expression of BAX at mRNA level was higher than that in control group (P<0.05). These effects could be reversed after treating iron-overloaded mice with DFX or NAC., Conclusion: Iron overload can inhibit the ratio of CD8+ T cells of splenic cells in mice, decrease the expression of IFN-γ, Granzyme-B, increase the apoptosis of CD3+ CD8+/CD8-. These effects may be regulated through increasing the intracellular ROS level, and can be partially reversed after treating iron-overloaded mice with DFX or NAC.

To evaluate the independent contribution of miRNAs to the missing heritability in CYP3A4/5 functionality and atorvastatin metabolism, the relationships among three levels of factors, namely (1) clinical characteristics, CYP3A4/5 genotypes, and miRNAs, (2) CYP3A4 and CYP3A5 mRNAs, and (3) CYP3A activity, as well as their individual impacts on atorvastatin metabolism, were assessed in 55 human liver tissues. MiR-27b, miR-206, and CYP3A4 mRNA respectively accounted for 20.0%, 5.8%, and 9.5% of the interindividual variations in CYP3A activity. MiR-142 was an independent contributor to the expressions of CYP3A4 mRNA (partial R(2) = 0.12, P = 0.002) and CYP3A5 mRNA (partial R(2) = 0.09, P = 0.005) but not CYP3A activity or atorvastatin metabolism. CYP3A activity was a unique independent predictor of variability of atorvastatin metabolism, explaining the majority of the variance in reduction of atorvastatin (60.0%) and formation of ortho-hydroxy atorvastatin (78.8%) and para-hydroxy atorvastatin (83.9%). MiR-27b and miR-206 were found to repress CYP3A4 gene expression and CYP3A activity by directly binding to CYP3A4 3'-UTR, while miR-142 was found to indirectly repress CYP3A activity. Our study indicates that miRNAs play significant roles in bridging the gap between epigenetic effects and missing heritability in CYP3A functionality.

Objective: To establish macrophage iron overload model in vitro by co-culture macrophages with iron, and to explore the effect of iron overload on cell reactive oxygen species (ROS) and the impact of ROS on macrophages., Method: Iron overload group were treated with different concentrations (0, 5, 10, 20, 40, 80 μmol/L respectively) of ferric ammonium citrate (FAC). The control group was the group of macrophages without FAC treatment. We detected the number and state of cells, metabolic activity, the change of phagocytosis, the levels of ROS and reactive nitrogen, and changes of related oxidative stress signaling pathways in different groups. Changes in the above indexes were detected after application of deferasirox (DFX) to remove iron and the antioxidant N -acetylcysteine (NAC) to clear excess oxidative stress., Results: (1)The levels of labile iron pool (LIP) in macrophages co-cultivated with iron was increased with the increase of iron concentration in a dose-dependent manner. The LIP levels was the highest in the macrophages treated with 80 μmol/L. (2)The increase of FAC concentration, the metabolic activity of macrophages in the 5 FAC-treated groups decreased to 51.58%, 40.98%, 16.23%, 3.46%, and 0.05% of the activity level of the control group (all P< 0.05). The group with the metabolic activity decreased to 16.23% (20 μmol/L) was selected as the iron overload group for the following experiments. (3)Compared with the control group, the number of macrophages in the iron overload group reduced to 32.80% (P<0.05), and the state of cells changed from adherence to partial suspension. The phagocytosis of macrophages in the iron overload group reduced to 20.40% of the control group (P<0.05). (4)Our further experiment showed that the levels of ROS and the activity nitrogen in the iron overload group increased by 7.71-and 1.45-fold compared with the control group (both P<0.05). The RT-PCR showed up-regulated mRNA expression of genes related with ROS production, i. e. nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX 4) gene related with ROS production and inducible nitric oxide synthase (iNOS) gene related with reactive nitrogen production, down-regulated mRNA expression of glutathione peroxidase 1 (GPX1) gene which participated in ROS clearance. Moreover, mRNA expression of phosphatidylinositol-3-kinase (PI3K) gene involved in oxidative stress signaling pathway in the iron overload group was up-regulated, while fork head protein O3 (FOXO3) which regulated oxidative stress through negative feedback showed a down-regulation level of mRNA expression compared with the control group. (5)After iron chelation and antioxidant treatment, the above-mentioned damage in the iron overload group were partially reversed., Conclusions: The damages of iron overload on macrophages may be mediated by inducing oxidative stress and activating oxidative stress signaling pathways. Our established model provides a method to explore the mechanism of iron overload on macrophage, and may shed some new light on possible therapeutic target in treating iron overload patients.

Embryonic stem cells (ESCs) are a population of pluripotent cells which can differentiate into different cell types. However, there are few reports with regard to differentiate ESCs into epidermal cells in vitro. In this study, we aimed to investigate differentially methylated promoters involved in process of differentiation from ESCs into epidermal-like cells (ELCs) induced by human amnion. We successfully induced ESCs into ELCs, which expressed the surface markers of CK19, CK15 and β1-integrin. With MeDIP-chip arrays, we identified 3435 gene promoters to be differentially methylated, involving 894 HCP (high CpG-containing promoter), 974 ICP (intermediate CpG-containing promoter) and 1567 LCP (low CpG-containing promoter) among all the 17,500 DNA methylation regions of gene promoters in both ESCs and ELCs. Gene oncology and pathway analysis demonstrated that these genes were involved in all the three categories of GO enrichment analysis, including biological process, molecular function and cellular component. All these data suggested that embryonic stem cells can differentiate into epidermal-like cells and promoter methylation is of great importance in this process., (Copyright © 2015 John Wiley & Sons, Ltd.)

Objective: To establish a mouse model of iron overload by intraperitoneal injection of iron dextran and investigate the impact of iron overload on bone marrow hematopoiesis., Methods: A total of 40 C57BL/6 mice were divided into control group, low-dose iron group (12.5 mg/ml), middle-dose iron group (25 mg/ml), and high-dose iron group (50 mg/ml). The control group received normal saline (0.2 ml), and the rest were injected with intraperitoneal iron dextran every three days for six weeks. Iron overload was confirmed by observing the bone marrow, hepatic, and splenic iron deposits and the bone marrow labile iron pool. In addition, peripheral blood and bone marrow mononuclear cells were counted and the hematopoietic function was assessed., Results: Iron deposits in bone marrow, liver, and spleen were markedly increased in the mouse models. Bone marrow iron was deposited mostly within the matrix with no significant difference in expression of labile iron pool.Compared with control group, the ability of hematopoietic colony-forming in three interventional groups were decreased significantly (P<0.05). Bone marrow mononuclear cells counts showed no significant difference. The amounts of peripheral blood cells (white blood cells, red blood cells, platelets, and hemoglobin) in different iron groups showed no significant difference among these groups;although the platelets were decreased slightly in low-dose iron group [(780.7±39.60)×10(9)/L], middle dose iron group [(676.2±21.43)×10(9)/L], and high-dose iron group [(587.3±19.67)×10(9)/L] when compared with the control group [(926.0±28.23)×10(9)/L], there was no significant difference(P>0.05)., Conclusions: The iron-overloaded mouse model was successfully established by intraperitoneal administration of iron dextran. Iron overload can damage the hepatic, splenic, and bone marrow hematopoietic function, although no significant difference was observed in peripheral blood count.

The fluorescence characteristics of oxidation reaction between MDA and cooked ground meat were analyzed by front face three dimensional synchronous fluorescence spectroscopy, parallel factor and two dimensional correlation technique. The results showed that the reaction system has two synchronous fluorescence peaks, one is Ex 292 nm and deltalambda 50 nm, assigned to the fluorescence characteristics of tryptophan residues in proteins; the other is Ex 400 nm, delta 70 nm, corresponding with the fluorescence characteristics of MDA-protein adducts formed during oxidation; The synchronous fluorescence landscape was analyzed using PARAFAC. The loading profiles of 1st and 2nd components had an optimal lambda 50 and 70 nm, respectively. During oxidation reaction, the synchronous fluorescence intensity of tryptophan gradually decreased, while the synchronous fluorescence intensity of MDA-protein adducts gradually increased. Two dimensional correlation synchronous fluorescence spectroscopy technique showed that the variation ratio of fluorescence intensity of tryptophan preceded that of MDA-protein adducts.

Objective: To explore the effects of iron overload on umbilical cord derived mesenchymal stem cells (UC-MSC) and elucidate the involvement of reactive oxygen species (ROS) in this process., Methods: The iron overload model of MSC was established by in vitro addition of ferric ammonium citrate (FAC) into culture medium. Cell proliferation and apoptosis were determined by Annexin V/PI double staining and population doubling time (DT) respectively. Co-culture system was used to assess the hematopoietic support capacity of UC-MSC in different groups. Thereafter the ROS level was detected with fluorescent probe 2', 7'-dichlorofluorescin diacetate (DCFH-DA). And the ROS related signaling factors of p-p38MAPK, p38 MAPK, P53 were measured by Western blot., Results: (1) The DT of UC-MSC in iron overload group was significantly longer than that of control ((24.43 ± 2.72) h vs (16.03 ± 2.31) h, P < 0.05). But the difference was insignificant after two passages (P > 0.05). (2) Apoptosis in iron overload group was higher than that of control (12.75% ± 0.32% vs 3.63% ± 0.80%, P < 0.05). (3) The colony forming capacity of mononuclear cell (MNC) co-cultured with UC-MSC of iron overload group for 1/2 weeks significantly decreased. (4) The ROS level of UC-MSC with iron overload was higher than that of control in time and concentration-dependent fashions and it peaked at 400 µmol/L of FAC for 12 h (1499 ± 86 vs 548 ± 97, P < 0.05). (5) The expressions of p-p38MAPK and P53 increased in response to FAC compared with control. But such an effect was partially inhibited after the use of antioxidants., Conclusions: Iron overload may impair the proliferation, survival and hematopoiesis supportive function of UC-MSC by enhancing the generation of ROS. And ROS stimulates the signaling pathways of p-p38MAPK and P53.

Background: Sequence variants in the β-adrenergic receptor (ADRB) genes have a close relationship with the development of coronary artery disease (CAD) and the patient's prognosis. However, there is a lack of data on the role of the variants in ADRBs genes in Han Chinese patients with CAD. We aimed to investigate the association of genetic variants in the ADRB1 and ADRB2 genes with the incidence of major adverse cardiac event (MACE) in Han Chinese patients with CAD., Methods: A total of 545 Han Chinese patients with CAD undergoing percutaneous coronary intervention (PCI) were recruited to the study and followed for one year. Three variant sites in ADRB1 (rs1801253) and ADRB2 (rs1042713 and rs1042714) were genotyped. The effect of the ADRB1 and ADRB2 genotypes on MACE within one year was assessed., Results: There were 47 cases of MACE during follow-up. There was no significant difference in the incidence of MACE among patients carrying different genotypes of the three variants in ADRB1 and ADRB2 (Log-rank, all P > 0.05). Cox regression analysis showed no association between three variants in ADRB1 and ADRB2 genes and the incidence of MACE during one-year follow-up, the adjusted hazard ratios (95% confidence interval) for rs1801253, rs1042713 and rs1042714 were 1.05 (0.54-2.02), 1.24 (0.58-2.64) and 1.66 (0.81-3.42), respectively., Conclusion: Our data did not support a relationship between the three polymorphisms of ADRB1 (rs1801253) and ADRB2 (rs1042713 and rs1042714) genes and risk of subsequent cardiovascular events after PCI in Han Chinese patients with CAD.

Reactive oxygen species (ROS) is a kind of molecules derived by oxygen in the metabolic process of aerobic cells, which mainly includes superoxide, hydroxyl radicals, alkoxyl, hydrogen peroxide, hypochlorous acid, ozone, etc. They can destroy the structure and function of cells through the damage of biological macromolecules such as DNA, proteins and the lipid peroxidation. ROS also can regulate the proliferation, differentiation and apoptosis of cells through several signaling pathways and participate in fibrogenesis of many organs including hepatic and pulmonary fibrosis. Recent study shows that ROS might have an important effect on the forming of myelofibrosis. Consequently, ROS plays a significant role in the fibrogenesis of tissues and organs. In this review, the relevance between ROS and common tissues and organs fibrosis is summarized.