Modeling X chromosome inactivation using t5iLA naive human pluripotent stem cells.

BACKGROUND: X chromosome inactivation (XCI) is a critical epigenetic event for dosage compensation of X-linked genes in female mammals, ensuring developmental stability. A robust in vitro model is required for mimicking XCI during the early stages of embryonic development. This methodology article introduces an advanced framework for the in-depth study of XCI using human pluripotent stem cells (hPSCs). By focusing on the transition between naive and primed pluripotent states, we highlight the role of long non-coding RNA X-inactive specific transcript (XIST) and epigenetic alterations in mediating XCI. RESULTS: Our methodology enables the distinction between naive and primed hESCs based on XIST expression and the activity of X-linked reporters, facilitating the investigation of XCI initiation and maintenance. Through detailed experimental procedures, we demonstrate the utility of our hESC lines in modeling the process of human XCI, including the establishment of conditions for random XCI induction and the analysis of X chromosome reactivation. METHODS: The study outlines a comprehensive approach for characterizing the X chromosome status in hPSCs, employing dual fluorescent reporter hESC lines. These reporter lines enable real-time tracking of XCI dynamics through differentiation processes. We detailed protocols for the induction of X chromosome reactivation and inactivation, as well as the X status characterization methods including cultivation of hESCs, flow cytometric analysis, RNA fluorescence in situ hybridization (FISH), and transcriptome sequencing, providing a step-by-step guide for researchers to investigate XCI mechanisms in vitro. CONCLUSIONS: This article provides a detailed, reproducible methodology for studying XCI mechanisms in vitro, employing hPSCs as a model system. It presents a significant advance in our ability to investigate XCI, offering potential applications in developmental biology, disease modeling, and regenerative medicine. By facilitating the study of XCI dynamics, this methodological framework paves the way for deeper understanding and manipulation of this fundamental biological process.

Comprehensive genome-wide analysis of the DREB gene family in Moso bamboo (Phyllostachys edulis): evidence for the role of PeDREB28 in plant abiotic stress response.

Dehydration response element binding (DREB) proteins are vital for plant abiotic stress responses, but the understanding of DREBs in bamboo, an important sustainable non-timber forest product, is limited. Here we conducted a comprehensive genome-wide analysis of the DREB gene family in Moso bamboo, representing the most important running bamboo species in Asia. In total, 44 PeDREBs were identified, and information on their gene structures, protein motifs, phylogenetic relationships, and stress-related cis-regulatory elements (CREs) was provided. Based on the bioinformatical analysis, we further analyzed PeDREBs from the A5 group and found that four of five PeDREB transcripts were induced by salt, drought, and cold stresses, and their proteins could bind to stress-related CREs. Among these, PeDREB28 was selected as a promising candidate for further functional characterization. PeDREB28 is localized in nucleus, has transcriptional activation activity, and could bind to the DRE- and coupling element 1- (CE1) CREs. Overexpression of PeDREB28 in Arabidopsis and bamboo improved plant abiotic stress tolerance. Transcriptomic analysis showed that broad changes due to the overexpression of PeDREB28. Furthermore, 628 genes that may act as the direct PeDREB28 downstream genes were identified by combining DAP-seq and RNA-seq analysis. Moreover, we confirmed that PeDREB28 could bind to the promoter of pyrabactin-resistance-like gene (DlaPYL3), which is a homolog of abscisic acid receptor in Arabidopsis, and activates its expression. In summary, our study provides important insights into the DREB gene family in Moso bamboo, and contributes to their functional verification and genetic engineering applications in the future.

Precisely Constructing Renal-Clearable and LAP-Activatable Ratiometric Molecular Probes for Early Diagnosis of Acute and Chronic Kidney Injury Via Optimizing Asymmetric DPP Dyes.

Fluorescence analysis is an increasingly important contributor to the early diagnosis of kidney diseases. To achieve precise visualization of the kidneys and early diagnosis of related diseases, an asymmetric pyrrolopyrrolidone (DPP) dye platform with C-aromatic substituents and N-lipophilic/hydrophilic modification was constructed. Based on these, we developed the renal-clearable, water-soluble, and kidney injury biomarker leucine aminopeptidase (LAP) activated ratiometric fluorescent probe DPP-S-L. In the mouse model of cisplatin-induced acute kidney injury and during the development of type 2 diabetes to diabetic kidney disease, we visualized for the first time the upregulation of LAP in the kidney and urine by dual-channel ratiometric fluorescence signal and diagnosed the kidney injury earlier and more sensitively than blood/urine enzyme detection and tissue analysis. This study showcases an excellent asymmetric DPP dye platform and renal-clearable ratiometric fluorescent probe design strategy that is extended to determination and visualization of other biomarkers for early disease diagnosis.

Assessing the Post-Activation Performance Enhancement of Upper Limbs in Basketball Athletes: A Sensor-Based Study of Rapid Stretch Compound and Blood Flow Restriction Training.

OBJECTIVE: This study introduces a novel methodology combining rapid stretch compound training with blood flow restriction (BFR) to investigate post activation performance enhancement (PAPE) in basketball players, a field that has been predominantly explored for lower limbs. We aimed to assess the efficacy of this combined approach on upper limb muscle performance in athletes. METHODS: We employed a randomized, self-controlled crossover trial with ten male basketball players. The bench press throw (BPT) served as the primary metric, with players undergoing four interventions post-baseline: (1) STR-plyometric training; (2) BFR-blood flow restriction; (3) COMB-STR integrated with BFR; and (4) CON-control. Innovatively, we utilized an intelligent tracking sensor to precisely measure peak power (PP), peak velocity (PV), mean power (MP), and mean velocity (MV) at 4, 8, and 12 min post-intervention, providing a detailed temporal analysis of PAPE. RESULTS: The COMB intervention demonstrated superior PAPE effects at 4 min, significantly outperforming STR and BFR alone and the control group across all measured indices (p < 0.05). Notably, the COMB group maintained these improvements for PV, PP, and H up to 12 min post-intervention, suggesting a prolonged effect. CONCLUSION: (1) The COMB stimulation has been shown to successfully induce PAPE more effectively than STR and BFR modality alone. (2) It appears that the optimal effects of PAPE are achieved within 4 min of exercising under this COMB. By the 12 min mark, only the COMB group continued to show significant improvements in PV, PP, and H compared to both the baseline and the CON group, while the effects in the STR and BFR groups further diminished. This suggests that although the PAPE effect is maintained over time, its optimal performance may peak at the 4 min mark and then gradually weaken as time progresses.

Synthesis and Properties of Cobalt/Nickel-Iron-Antimony(III, V)-Oxo Tartrate Cluster-Based Compounds.

Two types of isostructural iron-cobalt/nickel-antimony-oxo tartrate cluster-based compounds, namely (H3O)(Me2NH2)[M(H2O)6]2[FeII2SbIII12(µ4-O)3(µ3-O)8(tta)6]·6H2O (M = Co (1); Ni (3)), H5/3[Co2.5FeII4/3FeIII3(H2O)13SbV1/3FeIII2/3(µ4-O)2(µ3-O)4SbIII6(µ3-O)2(tta)6]·2H2O (2) and H2[Ni2.25FeII1.5FeIII3(H2O)14SbV0.25FeIII0.75(µ4-O)2(µ3-O)4SbIII6(µ3-O)2(tta)6]·2H2O (4) (H4tta = tartaric acid) were synthesized via simple solvothermal reactions. All the clusters in the structures adopt sandwich configurations, that is, bilayer sandwich configuration in 1 and 3 and monolayer sandwich configuration in 2 and 4. Interestingly, the monolayer sandwiched compounds 2 and 4 represent rare examples of cluster-based compounds containing mixed-valence Sb(III, V), whose center of the intermediate layer is the co-occupied [FexSbV1-x]. This is different from that of previously reported sandwich-type antimony-oxo clusters in which the center position is either occupied by a transition metal ion or a Sb(V) alone. Thus, the discovery of title compounds 2 and 4 makes the evolution of center metal ion more complete, that is, from M, MxSbV1-x to SbV. All the title compounds were fully characterized, and the photocatalysis, proton conduction and magnetism of compounds 2 and 4 were studied.

Insulin-transferrin-selenium promote formation of tissue-engineered vascular grafts in early stage of culture.

To create tissue-engineered vascular grafts (TEVGs) in vitro, vascular smooth muscle cells (VSMCs) must function effectively and produce sufficient extracellular matrix (ECM) in a three-dimensional space. In this study, we investigated whether the addition of insulin-transferrin-selenium (ITS), a medium supplement, could enhance TEVG formation. PGA fabric was used as the scaffold, and 1% ITS was added to the medium. After two weeks, the tissues were examined using electron microscopy and staining. The ITS group exhibited a denser structure and increased collagen production. VSMCs were cultured in two dimensions with ITS and assessed for collagen production, cell growth, and glucose metabolism. The results showed that ITS supplementation increased collagen production, cell growth, glucose utilization, lactate production, and ATP levels. Furthermore, reducing the amount of fetal bovine serum (FBS) in the medium did not affect the TEVGs or VSMCs when ITS was present. In conclusion, ITS improves TEVG construction by promoting VSMCs growth and reducing the need for FBS.

A Membrane-Anchoring Self-Assembling Peptide Allows Bioorthogonal Coupling of Type-I AIEgens for Pyroptosis-Induced Cancer Therapy.

Enrichment of photosensitizers (PSs) on cancer cell membranes via bioorthogonal reactions is considered to be a very promising therapeutic modality. However, azide-modified sugars-based metabolic labeling processes usually lack targeting and the labeling speed is relatively slow. Moreover, it has been rarely reported that membrane-anchoring pure type-I PSs can induce cancer cell pyroptosis. Here, we report an alkaline phosphatase (ALP) and cholecystokinin-2 receptor (CCK2R) dual-targeting peptide named DBCO-pYCCK6, which can selectively and rapidly self-assemble on cancer cell membrane, and then bioorthogonal enrich type-I aggregation-induced emission luminogens (AIEgen) PSs (SAIE-N3) on the cell membrane. Upon light irradiation, the membrane-anchoring SAIE-N3 could effectively generate type-I reactive oxygen species (ROS) to induce gasdermin E (GSDME)-mediated pyroptosis. In vivo experiments demonstrated that the bioorthogonal combination strategy of peptide and AIEgen PSs could significantly inhibit tumor growth, which is accompanied by CD8+ cytotoxic T cell infiltration. This work provides a novel self-assembly peptide-mediated bioorthogonal reaction strategy to bridge the supramolecular self-assembly and AIE field through strain-promoted azide-alkyne cycloaddition (SPAAC) and elucidates that pure type-I membrane-anchoring PSs can be used for cancer therapy via GSDME-mediated pyroptosis.

[Early identification of acute kidney injury in children with primary nephrotic syndrome]. / å„¿ç«¥åŽŸå‘æ€§è‚¾ç— ç»¼åˆå¾å¹¶å‘æ€¥æ€§è‚¾æŸä¼¤çš„æ—©æœŸè¯†åˆ«.

OBJECTIVES: To investigate the incidence and risk factors for acute kidney injury (AKI) in children with primary nephrotic syndrome (PNS), as well as the role of neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1 (KIM-1) in the early identification of AKI in these children. METHODS: A prospective collection of clinical data from children hospitalized with PNS at the Children's Hospital of the Capital Institute of Pediatrics from January 2021 to October 2022 was conducted. The children were divided into two groups based on the presence of AKI: the AKI group (47 cases) and the non-AKI group (169 cases). The risk factors for AKI in children with PNS were identified by multivariate logistic regression analysis. Urinary KIM-1 and NGAL levels were compared between the AKI and non-AKI groups, as well as among the different stages of AKI. RESULTS: The incidence of AKI in children with PNS was 21.8%. Multivariate logistic regression analysis revealed that steroid-resistant nephrotic syndrome, gastrointestinal infections, and heavy proteinuria were independent risk factors for AKI in these children with PNS (P<0.05). Urinary KIM-1 and NGAL levels were higher in the AKI group compared to the non-AKI group (P<0.05), and the urinary NGAL and KIM-1 levels in the AKI stage 2 and stage 3 subgroups were higher than those in the AKI stage 1 subgroup (P<0.017). CONCLUSIONS: KIM-1 and NGAL can serve as biomarkers for the early diagnosis of AKI in children with PNS. Identifying high-risk populations for AKI in children with PNS and strengthening the monitoring of related risk factors is of significant importance.

Quantitative proteomics reveals PPAR signaling pathway regulates the cardiomyocyte activity of neonatal mouse heart.

Cardiovascular diseases (CVDs) are among the most morbid and deadly types of diseases worldwide, while the existing therapeutic approaches all have their limitations. Mouse heart undergoes a very complex postnatal developmental process, including the 1-week window in which cardiomyocytes (CMs) maintain relatively high cell activity. The underlying mechanism provides an attractive direction for CVDs treatment. Herein, we collected ventricular tissues from mice of different ages from E18.5D to P8W and performed iTRAQ-based quantitative proteomics to characterize the atlas of cardiac development. A total of 3422 proteins were quantified at all selected time points, revealing critical proteomic changes related to cardiac developmental events such as the metabolic transition from glycolysis to beta-oxidation. A cluster of significantly dysregulated proteins containing proteins that have already been reported to be associated with cardiac regeneration (Erbb2, Agrin, and Hmgb) was identified. Meanwhile, the peroxisome proliferator-activated receptor (PPAR) signaling pathway (Cpt1α, Hmgcs2, Plin2, and Fabp4) was also found specifically enriched. We further revealed that bezafibrate, a pan-activator of PPAR signaling pathway markedly enhanced H9C2 cardiomyocyte activity via enhancing Cpt1α expression. This work provides new hint that activation of PPAR signaling pathway could potentially be a therapeutic strategy for the treatment of CVDs.

Quasi-LD-Targeted and ONOO--Responsive Fluorescent Probe for Investigating the Interaction of Nonalcoholic Fatty Liver with Drug-Induced Liver Injury.

Hepatic lipid droplets (LDs) and peroxynitrite (ONOO-) levels are closely related to nonalcoholic fatty liver disease (NAFLD). Additionally, some drug-induced liver injury (DILI) is often associated with ONOO-. Here, we constructed and screened the quasi-LDs-targeted and ONOO--responsive fluorescent probe MBDP-Py+ to investigate the interaction of NAFLD with DILI. By monitoring the upregulation of the ONOO- levels and the accumulation of LDs, MBDP-Py+ was more sensitive and efficient than tissue staining and serum markers detection in evaluating the early toxicity of NAFLD and diagnosing the anticancer-DILI. More importantly, the sensitive enhancement of fluorescence signals demonstrated that in different stages of NAFLD, the dominant element of liver injury was different in the NAFLD combined with DILI mice models. As the degree of NAFLD deepens, the synergistic effect of the two will lead to more serious liver damage.

Stone-Wales Defect in Graphene.

2D graphene the most investigated structures from nanocarbon family studied in the last three decades. It is projected as an excellent material useful for quantum computing, artificial intelligence, and next generation advanced technologies. Graphene exists in several forms and its extraordinary thermal, mechanical, and electronic properties, principally depend on the kind of perfection of the hexagonal atomic lattice. Defects are always considered as undesired components but certain defects in graphene could be an asset for electrochemistry and quantum electronics due to the engineered electronclouds and quantum tunnelling. The authors carefully discuss the Stone-Wales imperfections in graphene and its derivatives comprehensively. A specific emphasis is focused on the experimental and theoretical aspects of the Stone-Wales defects in graphene with respect to structure-property relationships. The corroboration of extrinsic defects like external atomic doping, functionalization, edge distortion in the graphene consisting of Stone-Wales imperfections, which are very significant in designing graphene-based electronic devices, are summarized.

Regulation of Female Germline Specification via Small RNA Mobility in Arabidopsis.

In the ovules of most sexually reproducing plants, one hypodermal cell differentiates into a megaspore mother cell (MMC), which gives rise to the female germline. Trans-acting small interfering RNAs known as tasiR-ARFs have been suggested to act non-cell-autonomously to prevent the formation of multiple MMCs by repressing AUXIN RESPONSE FACTOR3 (ARF3) expression in Arabidopsis (Arabidopsis thaliana), but the underlying mechanisms are unknown. Here, we examined tasiR-ARF-related intercellular regulatory mechanisms. Expression analysis revealed that components of the tasiR-ARF biogenesis pathway are restricted to distinct ovule cell types, thus limiting tasiR-ARF production to the nucellar epidermis. We also provide data suggesting tasiR-ARF movement along the mediolateral axis into the hypodermal cells and basipetally into the chalaza. Furthermore, we used cell type-specific promoters to express ARF3m, which is resistant to tasiR-ARF regulation, in different ovule cell layers. ARF3m expression in hypodermal cells surrounding the MMC, but not in epidermal cells, led to a multiple-MMC phenotype, suggesting that tasiR-ARFs repress ARF3 in these hypodermal cells to suppress ectopic MMC fate. RNA sequencing analyses in plants with hypodermally expressed ARF3m showed that ARF3 potentially regulates MMC specification through phytohormone pathways. Our findings uncover intricate spatial restriction of tasiR-ARF biogenesis, which together with tasiR-ARF mobility enables cell-cell communication in MMC differentiation.

Vagus nerve stimulation paired with rehabilitation for motor function, mental health and activities of daily living after stroke: a systematic review and meta-analysis.

OBJECTIVE: Vagus nerve stimulation (VNS) plus rehabilitation (Rehab) has shown a potential effect on recovery with a stroke. We systematically synthesised studies examining VNS+Rehab for improving motor function, mental health and activities of daily living (ADL) postintervention and at the end of follow-up in patients with a stroke. METHODS: The search was performed in electronic databases EMBASE, Medline, EBSCO, Cochrane Library, PubMed, PsycINFO, CINAHL, CNKI, and WANFANG and three clinical trial registries from inception to February 2022. Randomised controlled trials (RCTs) applied VNS+Rehab in stroke were included. RESULTS: Seven RCTs involving 263 (analysed) participants was included. The effect size of VNS+Rehab over Rehab for motor function was medium postintervention (g=0.432; 95% CI 0.186 to 0.678) and large at the end of follow-up (g=0.840; 95% CI 0.288 to 1.392). No difference was found in the effect of VNS+Rehab over traditional rehabilitation for ADL, mental health or safety outcomes. Subgroup analyses revealed larger effects for patients received taVNS (transcutaneous auricular VNS) devices (at acute/subacute phase of stroke, with lower VNS stimulation frequency or pluses per session, greater VNS on-off time or sessions, higher VNS intervention weekly frequency). CONCLUSION: The results suggest VNS+Rehab showed better motor function outcomes in patients after stroke, while no better than Rehab on mental health or ADL. Combinations of phase of stroke, specific parameters of VNS and VNS intervention frequency are key modulators of VNS effects. TRIAL REGISTRATION NUMBER: CRD42022310194.

Synthesis of 2'-fucosyllactose from apple pomace-derived xyloglucan oligosaccharides by an α-L-fucosidase from Pedobacter sp. CAU209.

2'-Fucosyllactose (2'-FL) is known for its ability to provide various health benefits to infants, such as gut maturation, pathogen resistance, improved immunity, and nervous system development. However, the production of 2'-FL using α-L-fucosidases is hindered by the lack of low-cost natural fucosyl donors and high-efficiency α-L-fucosidases. In this work, a recombinant xyloglucanase from Rhizomucor miehei (RmXEG12A) was applied to produce xyloglucan-oligosaccharide (XyG-oligos) from apple pomace. Then, an α-L-fucosidase gene (PbFucB) was screened from the genomic DNA of Pedobacter sp. CAU209 and expressed in Escherichia coli. The capability of purified PbFucB to catalyze XyG-oligos and lactose to synthesize 2'-FL was further evaluated. The deduced amino acid sequence of PbFucB shared the highest identity (38.4%) with that of other reported α-L-fucosidases. PbFucB showed the highest activity at pH 5.5 and 35 °C. It catalyzed the hydrolysis of 4-nitrophenyl-α-L-fucopyranoside (pNP-Fuc, 20.3 U mg-1), 2'-FL (8.06 U mg-1), and XyG-oligos (0.43 U mg-1). Furthermore, PbFucB demonstrated a high enzymatic conversion rate in 2'-FL synthesis with pNP-Fuc or apple pomace-derived XyG-oligos as donors and lactose as acceptor. Under the optimized conditions, PbFucB converted 50% of pNP-Fuc or 31% of the L-fucosyl residue in XyG-oligos into 2'-FL. This work elucidated an α-L-fucosidase that mediates the fucosylation of lactose and provided an efficient enzymatic strategy to synthesize 2'-FL either from artificial pNP-Fuc or natural apple pomace-derived XyG-oligos. KEY POINTS: • Xyloglucan-oligosaccharide (XyG-oligos) was produced from apple pomace by a xyloglucanase from Rhizomucor miehei. • An α-L-fucosidase (PbFucB) from Pedobacter sp. CAU209 shared the highest identity (38.4%) with reported α-L-fucosidases. •PbFucB synthesized 2'-FL using apple pomace-derived XyG-oligos and lactose with a conversion ratio of 31%.

Three-dimension titanium phosphate aerogel for selective removal of radioactive strontium(II) from contaminated waters.

The effective removal of radioactive strontium (especially 90Sr) from nuclear wastewater is crucial to environmental safety. Nevertheless, materials with excellent selectivity in Sr removal remain a challenge since the similarity with alkaline earth metal ions in the liquid phase. In this work, a novel titanium phosphate (TiP) aerogel was investigated for Sr(II) removal from the radioactive wastewater based on the sol-gel method and supercritical drying technique. The TiP aerogel has amorphous, three-dimensional and mesoporous structures with abundant phosphate groups, which was confirmed by X-ray diffraction (XRD), energy dispersive spectroscopy (EDS), atomic force microscope (AFM) and Fourier transform infrared spectroscopy (FT-IR). The adsorbent exhibited high efficiency and selectivity for the removal of Sr(II) with an extensive distribution coefficient up to 4740.03 mL/g. The adsorption equilibrium reached within 10 min and the maximum adsorption capacity was 373.6 mg/g at pH 5. And the kinetics and thermodynamics data fitted well with the pseudo-second-order model and Langmuir model respectively. It can be attributed to the rapid trapping and slow intraparticle diffusion of Sr(II) inside the mesoporous channels of the TiP aerogel. Furthermore, TiP aerogel exhibited over 80% removal for 50 mg/L Sr2+ in real water systems (seawater, lake water and tap water). X-ray photoelectron spectroscopy (XPS) and Raman spectroscopy revealed that strong ionic bonding formed during Sr(II) adsorption with the phosphate group on TiP aerogel. These results indicated that TiP aerogel is a promising high-capacity adsorbent for the effective and selective capture of Sr(II) from radioactive wastewater.

Ultrasound-assisted reverse micelle extraction and characterization of tea protein from tea residue.

BACKGROUND: In this study, ultrasonic-assisted reverse micelles were used to extract tea protein from tea residues. First, the extraction conditions of ultrasonic power, ionic strength and pH were optimized by response surface methodology. Then, structural comparison of ultrasonic-assisted reverse micelle extraction of tea protein (UARME) and ultrasonic-assisted alkali extraction (UAAE) were performed using scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy and amino acid composition. RESULTS: The optimum process conditions were determined as follows: ultrasonic power 300 W, KCl 0.15 mol L-1 , pH 8. The extraction rate was 46.29%, which was close to the theoretical value (46.44%). SEM showed that the protein particles extracted by UARME were smaller than those by UAAE. The results of FTIR spectroscopy showed that the protein extracted by UARME had higher α-helix, ß-sheet and ß-turn, and the contents were 20%, 62.3% and 17.1%, respectively. The content of random coil was 0%, which was significantly lower than that of alkali extraction, indicating that the secondary structure of protein extracted by UARME was more orderly. By comparing the amino acid composition of the two methods, the amino acid content of tea protein extracted by UARME was significantly higher than that of UAAE. CONCLUSION: The biological activity of tea protein is closely related to its structure. Compared with alkali extraction, reverse micelles can better protect the secondary structure of proteins, which is of great significance for studying their functional properties. © 2022 Society of Chemical Industry.

Recombination events that occur in a poxtA-carrying Enterococcus faecium during the conjugation process.

OBJECTIVES: To investigate transferability of the poxtA-carrying plasmids in Enterococcus faecium and the mechanism of recombination that occurs during the conjugation process. METHODS: MICs were determined by broth microdilution. Transferability of the poxtA-carrying plasmids in E. faecium was investigated by conjugation. The mechanism of recombination that occurred during the conjugation process was explored by S1-PFGE and WGS. RESULTS: E. faecium strain Fac90 carries two plasmids, designated pFac90-154 and pFac90-54, respectively. Six transconjugants with different characteristics were obtained. In transconjugant T90-1, a plasmid-chromosome fusion event led to the integration of plasmid pFac90-154 from the donor E. faecium strain Fac90 into the chromosomal DNA of the recipient strain Enterococcus faecalis JH2-2. In transconjugants T90-2, -3 and -4, losses or additions of different-sized plasmid segments most likely occurred due to IS1216-mediated recombination. In transconjugants T90-5 and -6, two large plasmids with sizes of 101 656 and 149 526 bp were formed by plasmid fusion. CONCLUSIONS: To the best of our knowledge, this is the first report showing the integration of pFac90-154 from E. faecium Fac90 into the chromosomal DNA of recipient E. faecalis JH2-2 via homologous recombination. Besides, we showed that five new plasmid types were formed by genetic rearrangements. These recombination events resulted simultaneously in the formation of various types of mosaic plasmids with multiple resistance genes and/or conjugation characteristics, which might promote the transmission of diverse plasmids encoding resistance genes among enterococci. Thus, these data significantly expand our knowledge regarding conjugative events, establishing a dual role of conjugation in both dissemination of resistance genes and plasmid evolution.

Periodontitis may impair the homeostasis of systemic bone through regulation of gut microbiota in ApoE-/- mice.

AIM: To investigate whether periodontitis impacts bone homeostasis via gut microbiota regulation. MATERIALS AND METHODS: Experimental periodontitis was induced by ligatures (LIG group). ApoE-/- mice were employed as a model with weakened bone homeostasis. Bone turnover was evaluated through micro-computerized tomography, haematoxylin and eosin-stained sections, osteoblast and osteoclast biomarkers in the bone and serum. Gut microbiota was analysed through 16S ribosomal RNA gene sequencing. Serum concentrations of cytokines were detected by enzyme-linked immunosorbent assay. The role of gut microbiota was evaluated through their transplantation into antibiotic-treated mice. RESULTS: Periodontitis significantly increased the number of osteoclasts and the expression of the osteoclast biomarkers in the proximal tibia of ApoE-/- mice, with the RANKL/OPG (receptor activator of nuclear factor-κB ligand/osteoprotegerin) ratio significantly increased, which indicated the osteoclastic activity overwhelmed osteogenesis. Meanwhile, periodontitis altered the composition of gut microbiota and induced low-grade inflammation in the colon and blood circulation. Interestingly, the concentration of circulating tumour necrosis factor-α, interleukin (IL)-6, IL-1ß, IL-17A, and monocyte chemotactic factor-1 were positively correlated with faecal α1-antitrypsin and calprotectin, as well as serum OPG and RANKL. Furthermore, transplantation of gut microbiota from mice with periodontitis to antibiotic-treated mice could partially re-capitulate the phenotypes in the bone and colon. CONCLUSION: Periodontitis may impair systemic bone homeostasis through gut microbiota.

A novel neutral thermophilic ß-mannanase from Malbranchea cinnamomea for controllable production of partially hydrolyzed konjac powder.

Partially hydrolyzed konjac powder (PHKP) can be used to increase the daily intake of dietary fibers of consumers. To produce PHKP by enzymatic hydrolysis, a novel ß-mannanase gene (McMan5B) from Malbranchea cinnamomea was expressed in Pichia pastoris. It showed a low identity of less than 52% with other GH family 5 ß-mannanases. Through high cell density fermentation, the highest ß-mannanase activity of 42200 U mL-1 was obtained. McMan5B showed the maximal activity at pH 7.5 and 75 °C, respectively. It exhibited excellent pH stability and thermostability. Due to the different residues (Phe214, Pro253, and His328) in catalytic groove and the change of ß2-α2 loop, McMan5B showed unique hydrolysis property as compared to other ß-mannanases. The enzyme was employed to hydrolyze konjac powder for controllable production of PHKP with a weight-average molecular weight of 22000 Da (average degree of polymerization 136). Furthermore, the influence of PHKP (1.0%-4.0%) on the qualities of steamed bread was evaluated. The steamed bread adding 3.0% PHKP had the maximum specific volume and the minimum hardness, which showed 11.0% increment and 25.4% decrement as compared to the control, respectively. Thus, a suitable ß-mannanase for PHKP controllable production and a fiber supplement for steamed bread preparation were provided in this study. KEY POINTS: • A novel ß-mannanase gene (McMan5B) was cloned from Malbranchea cinnamomea and expressed in Pichia pastoris at high level. • McMan5B hydrolyzed konjac powder to yield partially hydrolyzed konjac powder (PHKP) instead of manno-oligosaccharides. • PHKP showed more positive effect on the quality of steamed bread than many other dietary fibers including konjac powder.

Association of ABCG2 polymorphisms with susceptibility to anti-tuberculosis drug-induced hepatotoxicity in the Chinese population.

The accumulation of endogenous hepatotoxin protoporphyrin IX (PPIX) in the liver was proposed to be a novel mechanism of anti-tuberculosis drug-induced hepatotoxicity (ATDH). ATP-binding cassette transporter G2 (ABCG2) plays an important role in modulating PPIX concentrations. This study aimed to explore the role of ABCG2 genetic polymorphisms in the risk of ATDH in Chinese patients.A 1:4 matched case-control study was performed among 202 ATDH cases and 808 controls. Conditional logistic regression model was used to estimate the association between genotypes and the risk of ATDH by odds ratios (ORs) with 95% confidence intervals (CIs).Male patients with CC genotype of rs2622605 had an increased risk of ATDH (adjusted OR = 1.615, 95% CI: 1.119-2.332, p = 0.011). The peak value of alkaline phosphatase (ALP) was significantly higher in male patients with CC genotype of rs2622605 than in those with TT + TC genotype during antituberculosis treatment (102.0 U/L vs. 98.0 U/L, p = 0.029).This is the first attempt to evaluate the association between ABCG2 genetic variants and the risk of ATDH. Based on the 1:4 matched case-control study, the polymorphism at rs2622605 in the ABCG2 gene may be associated with the susceptibility to ATDH in Chinese male patients.