Molecular Mechanism of Sevoflurane Affecting Fetal Nervous System's Development Through the GABAAR/Sirt 1 Pathway.

Objective: To investigate the molecular mechanism of sevoflurane affecting the development of the offspring's nervous system through the GABAAR/Sirt 1 pathway. Methods: Pregnant rats were obtained by mating females and males, and were randomly divided into 3 h sevoflurane (2.3% sevoflurane anesthesia for 3 h), 6 h sevoflurane (2.3% sevoflurane anesthesia for 6 h), Sirt-1 activator-SRT1720 (10 mg/kg SRT1720), 6 h sevoflurane+SRT1720 (10 mg/kg SRT1720) and control groups) group and control group, 31-day-old littermates were taken out and their learning and memory functions were examined by the water maze experiment; the heads were severed to remove the brains, and the kits were used to detect the levels of 5-HT and Ach in the brain tissue; the hippocampal tissues of the littermates were isolated, and neuronal damage in the hippocampal tissues was assessed by Nissen staining; neuronal apoptosis in the hippocampal tissues was detected by TUNEL staining; and GABAAR in the hippocampal tissues was detected by Western blot. GABAAR, Sirt-1, and apoptosis-related proteins (Caspase-3, BCL-2, BAX) in hippocampal tissue. Results: Compared with the control group, the 3 h sevoflurane group and the 6 h sevoflurane group neurons were arranged sparsely, the cells appeared to be swollen, the evasion latency, the apoptosis rate of neurons, the expression of Caspase-3, and BAX increased significantly, and the number of crossing the plateau, the level of 5-HT and Ach in the brain tissues, and the expression of GABAAR, Sirt-1, and BCL-2 were decreased significantly, and the differences existed between the groups (P < .5); compared with the 6 h sevoflurane group, neuronal morphological changes in the hippocampal tissue of the 6 h sevoflurane+SRT1720 group were improved, with a significant decrease in the evasion latency, neuronal apoptosis rate, expression of Caspase-3 and BAX, and a significant increase in the number of traversing platforms, brain tissue 5-HT, Ach level, GABAAR, Sirt-1, and BCL-2 expression (P < .5); compared with the SRT1720 group, the neurons in the 6 h sevoflurane + SRT1720 group were sparsely arranged, with a significant increase in evasion latency, neuronal apoptosis rate, caspase-3, BAX expression, and a significant decrease in the number of traversing platforms, brain tissue 5-HT, Ach level, GABAAR, Sirt-1, and BCL-2 expression (P < .5 ). Conclusion: Sevoflurane can affect the neurological development of rat offspring, which may be related to the inhibition of Sirt-1 expression.

Unraveling Cadmium Toxicity in Trifolium repens L. Seedling: Insight into Regulatory Mechanisms Using Comparative Transcriptomics Combined with Physiological Analyses.

Trifolium repens (T. repens) can accumulate significant amounts of heavy metal ions, and has strong adaptability to wide environmental conditions, and relatively large biomass, which is considered a potential plant for phytoremediation. However, the molecular mechanisms of T. repens involved in Cd tolerance have not yet been studied in detail. This study was conducted to examine the integrative responses of T. repens exposed to a high-level CdCl2 by investigating the physiological and transcriptomic analyses. The results suggested that T. repens seedlings had a high degree of tolerance to Cd treatment. The roots accumulated higher Cd concentration than leaves and were mainly distributed in the cell wall. The content of MDA, soluble protein, the relative electrolyte leakage, and three antioxidant enzymes (POD, SOD, and APX) was increased with the Cd treatment time increasing, but the CAT enzymes contents were decreased in roots. Furthermore, the transcriptome analysis demonstrated that the differentially expressed genes (DEGs) mainly enriched in the glutathione (GSH) metabolism pathway and the phenylpropanoid biosynthesis in the roots. Overexpressed genes in the lignin biosynthesis in the roots might improve Cd accumulation in cell walls. Moreover, the DEGs were also enriched in photosynthesis in the leaves, transferase activity, oxidoreductase activity, and ABA signal transduction, which might also play roles in reducing Cd toxicity in the plants. All the above, clearly suggest that T. repens employ several different mechanisms to protect itself against Cd stress, while the cell wall biosynthesis and GSH metabolism could be considered the most important specific mechanisms for Cd retention in the roots of T. repens.

Luteimonas wenzhouensis Sp. Nov., A Chitinolytic Bacterium Isolated from a Landfill Soil.

A Gram-staining-negative, aerobic, non-motile, rod-shaped bacterium with degradation ability of chitin, designated strain YD-1 T, was isolated from landfill soil sample collected in Wenzhou, Zhejiang province, China. The growth of strain YD-1 T occurred optimally in the tryptone soy broth (TSB) with 1.0% NaCl at pH 7.0-8.0, 30 °C. Ubiquinone-8 (Q-8) was the predominant quinone. The polar lipids of strain YD-1 T consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, five glycolipids and four lipids. The major fatty acids were iso-C15:0 (30.7%), iso-C17:1ω9c (23.2%), iso-C11:0 (18.9%), iso-C11:0 3-OH (6.8%) and iso-C17:0 (5.9%). Phylogenetic analysis based on the 16S rRNA gene sequences indicated that strain YD-1 T was affiliated to the genus Luteimonas with the highest similarity to Luteimonas marina KCTC 12327 T (97.3%), followed by Luteimonas aquatica DSM 22088 T (96.5%) and Luteimonas composti CCUG 53595 T (96.4%). The genomic DNA G + C content of strain YD-1 T was 71.8 mol%. Average nucleotide identity (ANI) and the digital DNA-DNA hybridizations (dDDH) for draft genomes between strain YD-1 T and Luteimonas marina KCTC 12327 T were 82.7% and 26.1%, respectively. On the basis of genotypic, phenotypic and chemotaxonomic data, strain YD-1 T is considered to represent a novel species to degrade chitin in the genus Luteimonas, for which the name Luteimonas wenzhouensis sp. nov. is proposed, with YD-1 T (= KCTC 72425 T = CCTCC AB 2019153 T) as the type strain.

A Stable 2D Zr(IV)-Based Metal-Organic Framework (USTS-7) for Selective Sensing of Cr2O72- in Aqueous Solution.

A novel 2D porous Zr(IV)-based metal-organic framework (USTS-7) was assembled from 2,5-bis[2-(methylthio)ethylthio]terephthalic acid and ZrCl4. USTS-7 retains its stability in water, strong acid, and base; moreover, it is highly luminescent and displays a remarkable selective sensing property toward Cr2O72- in aqueous solution with a very low detection limit.

Immune Regulation by Ubiquitin Tagging as Checkpoint Code.

The immune system is equipped with effective machinery to mobilize its activation to defend invading microorganisms, and at the same time, to refrain from attacking its own tissues to maintain immune tolerance. The balance of activation and tolerance is tightly controlled by diverse mechanisms, since breakdown of tolerance could result in disastrous consequences such as the development of autoimmune diseases. One of the mechanisms is by the means of protein ubiquitination, which involves the process of tagging a small peptide ubiquitin to protein substrates. E3 ubiquitin ligases are responsible for catalyzing the final step of ubiquitin-substrate conjugation by specifically recognizing substrates to determine their fates of degradation or functional modification. The ubiquitination process is reversible, which is carried out by deubiquitinating enzymes to release the ubiquitin molecule from the conjugated substrates. Protein ubiquitination and deubiquitination serve as checkpoint codes in many key steps of lymphocyte regulation including the development, activation, differentiation, and tolerance induction. In this chapter, we will discuss a few E3 ligases and deubiquitinating enzymes that are important in controlling immune responses, with emphasis on their roles in T cells.

Identification and characterization of plant-specific NAC gene family in canola (Brassica napus L.) reveal novel members involved in cell death.

NAC transcription factors are plant-specific and play important roles in plant development processes, response to biotic and abiotic cues and hormone signaling. However, to date, little is known about the NAC genes in canola (or oilseed rape, Brassica napus L.). In this study, a total of 60 NAC genes were identified from canola through a systematical analysis and mining of expressed sequence tags. Among these, the cDNA sequences of 41 NAC genes were successfully cloned. The translated protein sequences of canola NAC genes with the NAC genes from representative species were phylogenetically clustered into three major groups and multiple subgroups. The transcriptional activities of these BnaNAC proteins were assayed in yeast. In addition, by quantitative real-time RT-PCR, we further observed that some of these BnaNACs were regulated by different hormone stimuli or abiotic stresses. Interestingly, we successfully identified two novel BnaNACs, BnaNAC19 and BnaNAC82, which could elicit hypersensitive response-like cell death when expressed in Nicotiana benthamiana leaves, which was mediated by accumulation of reactive oxygen species. Overall, our work has laid a solid foundation for further characterization of this important NAC gene family in canola.

Apple polyphenol extract ameliorates sugary-diet-induced depression-like behaviors in male C57BL/6 mice by inhibiting the inflammation of the gut-brain axis.

To explore whether apple polyphenol extract (APE) ameliorates sugary-diet-induced depression-like behaviors, thirty male C57BL/6 mice (3-4 weeks old) were assigned to three groups randomly to receive different treatments for 8 consecutive weeks: (1) control group (CON), (2) S-HSD group (60% high sucrose diet feeding with 0.1 mg mL-1 sucralose solution as drinking water), and (3) S-APE group (S-HSD feeding with 500 mg per (kg bw day) APE solution gavage). The S-HSD group showed significant depression-like behaviors compared with the CON group, which was manifested by an increased number of buried marbles in the marble burying test, prolonged immobility time in both the tail suspension test and forced swimming test, and cognitive impairment based on the Morris water maze test. However, APE intervention significantly improved the depression-like behaviors by reducing serum levels of corticosterone and adrenocorticotropic hormone, and increasing the serum level of IL-10. Moreover, APE intervention inhibited the activation of the NF-κB inflammatory pathway, elevated colonic MUC-2 protein expression, and elevated the colonic and hippocampal tight junction proteins of occludin and ZO-1. Furthermore, APE intervention increased the richness and diversity of gut microbiota by regulating the composition of microbiota, with increased relative abundance of Firmicutes and Bacteroidota, decreased relative abundance of Verrucomicrobiota at the phylum level, significantly lowered relative abundance of Akkermansia at the genus level, and rebalanced abnormal relative abundance of Muribaculaceae_unclassified, Coriobacteriaceae_UCG-002, and Lachnoclostridium induced by S-HSD feeding. Thus, our study supports the potential application of APE as a dietary intervention for ameliorating depression-like behavioral disorders.

Genome-Wide Identification, Characterization, and Expression Profiling Analysis of SPL Gene Family during the Inflorescence Development in Trifolium repens.

Trifolium repens is the most widely cultivated perennial legume forage in temperate region around the world. It has rich nutritional value and good palatability, seasonal complementarity with grasses, and can improve the feed intake and digestibility of livestock. However, flowering time and inflorescence development directly affects the quality and yield of T. repens, as well as seed production. The Squa promoter binding protein-like (SPL) gene family is a plant specific transcription factor family, which has been proved to play a critical role in regulating plant formation time and development of flowers. In this study, a total of 37 TrSPL genes were identified from the whole genome of T. repens and were divided into nine clades based on phylogenetic tree. Seventeen TrSPL genes have potential target sites for miR156. The conserved motif of squamosa promoter binding protein (SBP) contains two zinc finger structures and one NLS structure. Gene structure analysis showed that all TrSPL genes contained SBP domain, while ankyrin repeat region was just distributed in part of genes. 37 TrSPL genes were relatively dispersedly distributed on 16 chromosomes, and 5 pairs of segmental repeat genes were found, which indicated that segmental duplication was the main way of gene expansion. Furthermore, the gene expression profiling showed that TrSPL11, TrSPL13, TrSPL22, and TrSPL26 were highly expressed only in the early stage of inflorescence development, while TrSPL1 and TrSPL6 are highly expressed only in the mature inflorescence. Significantly, the expression of TrSPL4 and TrSPL12 increased gradually with the development of inflorescences. The results of this study will provide valuable clues for candidate gene selection and elucidating the molecular mechanism of T. repens flowering regulation.

Synthesis of Copper Nanoclusters and Their Application for Environmental Pollutant Probes: A Review.

Copper nanoclusters (CuNCs) as a new type of probe for environmental contaminants are gaining increasing attention because of its low cost, superior water dispersibility, wide availability and excellent optical properties. Compared with the other probes such as quantum dots and organic dyes, CuNCs show much more potential in practical application for their excellent photostability, large Stokes shift, low toxicity and other preponderance, especially in the fields of biosensing and environmental monitoring. Recently, the template-assisted synthesis of metal nanoclusters (MNCs) has been widely studied. A variety of templates such as proteins, small thiol molecules, polymers, and DNA with different spatial configuration have been used for the preparation of MNCs so far. This review primarily described recent advances in CuNCs in terms of the synthesis of CuNCs from different templates, the methods to improve the fluorescence (FL) properties of CuNCs, as well as the basic detection mechanisms based on the FL properties or catalytic properties. Finally, to promote the practical application of CuNCs probes, the challenges and prospects of CuNCs multifunctional probes are also discussed.

The Identification and Characterization of the <i>KNOX</i> Gene Family as an Active Regulator of Leaf Development in <i>Trifolium repens</i>.

Leaves are the primary and critical feed for herbivores. They directly determine the yield and quality of legume forage. <i>Trifolium repens</i> (<i>T. repens</i>) is an indispensable legume species, widely cultivated in temperate pastures due to its nutritional value and nitrogen fixation. Although the leaves of <i>T. repens</i> are typical trifoliate, they have unusual patterns to adapt to herbivore feeding. The number of leaflets in <i>T. repens</i> affects its production and utilization. The <i>KNOX</i> gene family encodes transcriptional regulators that are vital in regulating and developing leaves. Identification and characterization of <i>TrKNOX</i> gene family as an active regulator of leaf development in <i>T. repens</i> were studied. A total of 21 <i>TrKNOX</i> genes were identified from the T. repens genome database and classified into three subgroups (Class I, Class II, and Class M) based on phylogenetic analysis. Nineteen of the genes identified had four conserved domains, except for <i>KNOX5</i> and <i>KNOX9</i>, which belong to Class M. Varying expression levels of TrKNOX genes were observed at different developmental stages and complexities of leaves. <i>KNOX9</i> was observed to upregulate the leaf complexity of T. repens. Research on <i>TrKNOX</i> genes could be novel and further assist in exploring their functions and cultivating high-quality <i>T. repens</i> varieties.

R2R3-MYB gene family: Genome-wide identification provides insight to improve the content of proanthocyanidins in Trifolium repens.

The R2R3-MYB family is one of largest transcription factor families in plants playing significant roles in regulating anthocyanin and proanthocyanidin biosynthesis. Proanthocyanidins are one of major objectives to improve the quality of white clover (Trifolium repens L.), which have a beneficial effect on ruminant to prevent the lethal pasture bloat. A total of 133 TrR2R3-MYB genes were identified and distributed on all 16 chromosomes based on the whole genome information of white clover. Also, by exploring the gene structure, motifs and duplication events of TrR2R3-MYBs, as well as the evolutionary relationship with TrR2R3-MYB genes of other species, 10 TrR2R3-MYB genes with the potential to regulate the anthocyanins and proanthocyanidins biosynthesis were screened. These TrR2R3-MYB genes responded significantly to low temperature in white clover. In addition, they have different expression patterns in leaves, petioles and inflorescences of white clover. Importantly, TrMYB116 and TrMYB118 may positively regulate anthocyanin accumulation and low temperature response in white clover. TrMYB118 may also be associated with anthocyanin pigmentation pattern in Purple leaves. This study provides a basis for verifying the function of TrR2R3-MYB and breeding white clover cultivars with high proanthocyanidins.

[Preparation and chiral recognition of heterosubstituted amylose derivatives-based chiral stationary phases].

Two novel amylose derivatives, namely amylose 2-benzoate-3-(4-methylphenylcarbamate)-6-(3,5-dichlorophenylcarbamate) and amylose 2-benzoate-3-(3,5-dichlorophenylcarbamate)-6-(4-methylphenylcarbamate), were prepared utilizing a serial regioselective process. After coated onto aminopropyl silica gel, they were utilized as chiral stationary phases (CSPs) for high performance liquid chromatography. Investigations indicated that the CSPs exhibited characteristic chiral recognition and their chiral recognition abilities were much higher than those of amylose tris(3,5-dichlorophenylcarbamate) (ADCPC), a homosubstituted derivative. The nature and position of the substituents at 3-, and 6-positions of a glucose unit had great influence on the chiral resolution abilities of the amylose derivatives. Some chiral compounds which were not effectively resolved on the commercial column Chiralpak AD were effectively separated on the new CSPs. Moreover, with all the eight tested racemates resolved, amylose 2-benzoate-3-(4-methylphenylcarbamate)-6-(3,5-dichlorophenylcarbamate) exhibited relatively high chiral recognition and might be a potential useful CSP.

miR-548d-3p/TP53BP2 axis regulates the proliferation and apoptosis of breast cancer cells.

Fast growth and hardly any apoptosis are important characteristics of breast cancer, which assure the spread via invasion and metastasis of breast cancer cells. Inhibition of fast proliferation and induction of apoptosis are critical way to cure this cancer. microRNAs (miRNAs) had been increasingly reported to be the critical regulator of tumorigenesis. In our study, we found that increasing copy number of miR-548d-2-3p is critically involved poor prognosis. We overexpressed miR-548d-3p in MDA-MB-231cells and found that the proliferation was promoted significantly, whereas the inhibition of miR-548d-3p repressed the proliferation of MDA-MB-231 cells and also induced the increase in apoptosis. Additionally, we found that miR-548d-3p downregulated the expression of TP53BP2 by directly targeting the 3'UTR. We also found that knockdown of TP53BP2 significantly resorted the proliferation and apoptosis regulated by miR-548d-3p inhibitor. Our study showed that miR-548d-3p/TP53BP2 pathway is critically involved in the proliferation and apoptosis of breast cancer cells and may be new therapeutic target of breast cancer cells.

Identification, cloning and characterization of R2R3-MYB gene family in canola (Brassica napus L.) identify a novel member modulating ROS accumulation and hypersensitive-like cell death.

The R2R3-MYB proteins comprise one of the largest families of transcription factors in plants. Although genome-wide analysis of this family has been carried out in some plant species, little is known about R2R3-MYB genes in canola (Brassica napus L.). In this study, we have identified 76 R2R3-MYB genes in the canola genome through mining of expressed sequence tags (ESTs). The cDNA sequences of 44 MYB genes were successfully cloned. The transcriptional activities of BnaMYB proteins encoded by these genes were assayed in yeast. The subcellular localizations of representative R2R3-MYB proteins were investigated through GFP fusion. Besides, the transcript abundance level analysis during abiotic conditions and ABA treatment identified a group of R2R3-MYB genes that responded to one or more treatments. Furthermore, we identified a previously functionally unknown MYB gene-BnaMYB78, which modulates reactive oxygen species (ROS)-dependent cell death in Nicotiana benthamiana, through regulating the transcription of a few ROS- and defence-related genes. Taken together, this study has provided a solid foundation for understanding the roles and regulatory mechanism of canola R2R3-MYB genes.

Sevoflurane represses the self-renewal ability by regulating miR-7a,7b/Klf4 signalling pathway in mouse embryonic stem cells.

Sevoflurane is a frequently-used clinical inhalational anaesthetic and can cause toxicity to embryos during foetal development. Embryonic stem cells (ESCs) are derived from the inner cell mass of blastospheres and can be used as a useful model of early development. Here, we found that sevoflurane significantly influenced self-renewal ability of mESCs on stemness maintenance and cell proliferation. The cell cycle was arrested via G1 phase delay. We further found that sevoflurane upregulated expression of miR-7a,7b to repress self-renewal. Next we performed rescue experiments and found that after adding miR-7a,7b inhibitor into mESCs treated with sevoflurane, its influence on self-renewal could be blocked. Further we identified stemness factor Klf4 as the direct target of miR-7a,7b. Overexpression of Klf4 restored self-renewal ability repressed by miR-7a,7b or sevoflurane. In this work, we determined that sevoflurane repressed self-renewal ability by regulating the miR-7a,7b/Klf4 signalling pathway in mESCs. Our study demonstrated molecular mechanism underlying the side effects of sevoflurane during early development, laying the foundation for studies on safe usage of inhalational anaesthetic during non-obstetric surgery.

[Characterization of human lung cancer cell line A549 transfected with human interferon-gamma gene].

OBJECTIVE: To establish a human lung cancer cell line expressing human interferon-gamma. METHODS: The full-length gene of human interferon-gamma (IFN-gamma) was introduced into the human lung adenocarcinoma cell line A549 through retroviral vector pLXSN. The established cell line A549-IFN-gamma was tested for expression of MHC class I and class II by flow cytometer (FCM) and tested for expression of IFN-gamma by enzyme-lined immunoadsorbent assay (ELISA ). The tumorigenesis of cell line A549-IFN-gamma was tested on nude mice. RESULTS: A high level of IFN-gamma protein was detected in the culture supernatants of cell line A549-IFN-gamma. The expressions of MHC class I and class II on A549-IFN-gamma cells increased significantly (P<0.01), when compared with parental cell line A549. However, there was no significant difference (P<0.05) between the growth of cell line A549-IFN-gamma and A549. Finally, the tumorigenesis test showed that A549-IFN-gamma had lower tumorigenetic effects than A549. CONCLUSION: The results indicate that introduction of human IFN-gamma gene into cell line A549 could increase its immunogenicity and decrease its tumorigenesis. With the established cell line A549-IFN-gamma, a tumor vaccine for human lung cancer may be developed.