Polyploidization has provided much genetic variation for plant adaptive evolution, but the mechanisms by which the molecular evolution of polyploid genomes establishes genetic architecture underlying species differentiation are unclear. Brassica is an ideal model to increase knowledge of polyploid evolution. Here we describe a draft genome sequence of Brassica oleracea, comparing it with that of its sister species B. rapa to reveal numerous chromosome rearrangements and asymmetrical gene loss in duplicated genomic blocks, asymmetrical amplification of transposable elements, differential gene co-retention for specific pathways and variation in gene expression, including alternative splicing, among a large number of paralogous and orthologous genes. Genes related to the production of anticancer phytochemicals and morphological variations illustrate consequences of genome duplication and gene divergence, imparting biochemical and morphological variation to B. oleracea. This study provides insights into Brassica genome evolution and will underpin research into the many important crops in this genus.

Colonic chloride secretion is regulated via the neurohormonal and immune systems. Exogenous chemicals (e.g., butyrate, propionate) can affect chloride secretion. Capsaicin, the pungent ingredient of the chili peppers, exerts various effects on gastrointestinal function. Capsaicin is known to activate the transient receptor potential vanilloid type 1 (TRPV1), expressed in the mesenteric nervous system. Recent studies have also demonstrated its presence in epithelial cells but its role remains uncertain. Because capsaicin has been reported to inhibit colonic chloride secretion, we tested whether this effect of capsaicin could occur by direct action on epithelial cells. In mouse colon and model T84 human colonic epithelial cells, we found that capsaicin inhibited forskolin-dependent short-circuit current (FSK-I(sc)). Using PCR and Western blot, we demonstrated the presence of TRPV1 in colonic epithelial cells. In T84 cells, TRPV1 localized at the basolateral membrane and in vesicular compartments. In permeabilized monolayers, capsaicin activated apical chloride conductance, had no effect on basolateral potassium conductance, but induced NKCC1 internalization demonstrated by immunocytochemistry and basolateral surface biotinylation. AMG-9810, a potent inhibitor of TRPV1, did not prevent the inhibition of the FSK-I(sc) by capsaicin. Neither resiniferatoxin nor N-oleoyldopamine, two selective agonists of TRPV1, blocked the FSK-I(sc). Conversely capsaicin, resiniferatoxin, and N-oleoyldopamine raised intracellular calcium ([Ca(2+)](i)) in T84 cells and AMG-9810 blocked the rise in [Ca(2+)](i) induced by capsaicin and resiniferatoxin suggesting the presence of a functional TRPV1 channel. We conclude that capsaicin inhibits chloride secretion in part by causing NKCC1 internalization, but by a mechanism that appears to be independent of TRPV1.

A series of myricetin derivatives containing amide, thioether, and 1,3,4-thiadiazole moieties were designed and synthesized, and their antiviral and antibacterial activities were assessed. The bioassays showed that all the title compounds exhibited potent in vitro antibacterial activities against Xanthomonas citri (Xac), Ralstonia solanacearum (Rs), and Xanthomonas oryzae pv. Oryzae (Xoo). In particular, the compounds 5a, 5f, 5g, 5h, 5i, and 5l, with EC50 values of 11.5⁻27.3 μg/mL, showed potent antibacterial activity against Xac that was better than the commercial bactericides Bismerthiazol (34.7 μg/mL) and Thiodiazole copper (41.1% μg/mL). Moreover, the in vivo antiviral activities against tobacco mosaic virus (TMV) of the target compounds were also tested. Among these compounds, the curative, protection, and inactivation activities of 5g were 49.9, 52.9, and 73.3%, respectively, which were better than that of the commercial antiviral Ribavirin (40.6, 51.1, and 71.1%, respectively). This study demonstrates that myricetin derivatives bearing amide, thioether, and 1,3,4-thiadiazole moieties can serve as potential alternative templates for the development of novel, highly efficient inhibitors against plant pathogenic bacteria and viruses.

Several reports have suggested a possible association between the interleukin (IL)-8-251A/T single-nucleotide polymorphism (SNP) and the susceptibility to coronary artery disease (CAD). Due to inconclusive results of the studies so far, we conducted a meta-analysis to systematically summarize the studies on the association between this SNP and CAD risk. A systematic literature search identified 9 case-control studies (3752 cases and 4219 controls) on the IL-8-251A/T polymorphism. We observed a significant association between different genetic forms of -251A/T SNP and CAD risk, like the allele model (A vs T: odds ratio [OR] 1.14, 95% confidence interval [CI] 1.02-1.27, P = .02), dominant model (AA + AT vs TT: OR 1.20, 95% CI 1.01-1.43, P = .042), recessive model (AA vs AT + TT: OR 1.15, 95% CI 1.03-1.27, P = .01), and homozygous model (AA vs TT: OR 1.26, 95% CI 1.01-1.56, P = .037), whereas the heterozygote model did not show any significant association (AT vs TT: OR 1.16, 95% CI 0.98-1.38, P = .091). Furthermore, significant heterogeneity was observed among studies in terms of all genetic models, except the recessive model. Analysis of the ethnic subgroups revealed a significantly higher risk of CAD in the East Asian population carrying this SNP, and the heterogeneity among the studies regarding the East Asian population was decreased after subgroup analysis. The results of this meta-analysis suggest that the IL-8-251A/T SNP may increase the risk of CAD, especially in people of East Asian ethnicity. Further large-scale, multicenter epidemiological studies are warranted to validate this finding.

Dihydromyricetin (DMY), an important flavanone found in Ampelopsis grossedentata, plays a protective role in liver injury. Our previous research found that DMY protected L02 cells against hepatotoxicity caused by emodin. In this study, serum, urine, and liver samples from rats were systematically used for biochemical analysis, pathological observation, and nontargeted metabolomics to evaluate the toxicity of emodin and DMY intervention. After oral administration of DMY, DMY may alleviate liver injury by improving liver metabolism. Approximately, 8 of 15 metabolites in rat urine and serum were significantly regulated by DMY. Metabolic pathway analysis showed that glutathione metabolism, pyrimidine metabolism, and tryptophan metabolism were the most affected pathways, and 18 proteins were predicted to be potential targets of DMY during the alleviation of liver injury induced by emodin. This research is of great significance in confirming the liver-protective effect of DMY, especially during acute liver injury caused by traditional Chinese medicine.

Ovarian hyperstimulation syndrome (OHSS) is a common complication caused by ovulatory stimulation therapy, which manifests as an increase in ovarian volume, an increase in the number of oocytes retrieved, and increased vascular permeability throughout the body and especially in ovarian tissue. In our previous study, we found that electroacupuncture (EA) could prevent the progression of OHSS, by mainly affecting ovary. However, the specific molecules and the mechanism of this process were still unknown. In order to explore the underlying mechanism, OHSS rat model was established and EA treatment was performed, which was followed by proteomic analysis of ovaries. Results showed a significant increase in the expression level of CD200 in the ovaries of OHSS group treated with EA than those of OHSS group. Clinical data showed that the level of CD200 in follicular fluid was negatively correlated with the number of oocytes retrieved and serum E2 level. Further in vitro experiments showed a concentration-dependent role of human chorionic gonadotropin (hCG) in reducing CD200 and CD200R levels, and increasing inflammatory cytokine levels in cultured KGN cells. In human umbilical vein endothelial cells (HUVECs), the vascular barrier function was improved by CM (cultural medium from KGN cell) which treated with CD200Fc (CD200R agonist). Meanwhile, the results of in vivo experiments indicated that EA reduced the number of ovarian corpora lutea, decreased inflammatory response, and improved the vascular barrier function by increasing the expression of CD200 and CD200R in rat ovaries. These findings suggest that EA treatment may reduce oocyte number and maintain vascular barrier against OHSS through ovarian anti-inflammatory response mediated by CD200. Therefore, this study is the first to identify CD200 as a main of EA in the ovary and elucidate the possible mechanism of EA on preventing and treating OHSS, which provide a scientific basis for CD200 as an effector and indicator in EA treatment.

Cytosine base editors (CBEs) are great additions to the expanding genome editing toolbox. To improve C-to-T base editing in plants, we first compared seven cytidine deaminases in the BE3-like configuration in rice. We found A3A/Y130F-CBE_V01 resulted in the highest C-to-T base editing efficiency in both rice and Arabidopsis. Furthermore, we demonstrated this A3A/Y130F cytidine deaminase could be used to improve iSpyMacCas9-mediated C-to-T base editing at A-rich PAMs. To showcase its applications, we first applied A3A/Y130F-CBE_V01 for multiplexed editing to generate microRNA-resistant mRNA transcripts as well as pre-mature stop codons in multiple seed trait genes. In addition, we harnessed A3A/Y130F-CBE_V01 for efficient artificial evolution of novel ALS and EPSPS alleles which conferred herbicide resistance in rice. To further improve C-to-T base editing, multiple CBE_V02, CBE_V03 and CBE_V04 systems were developed and tested in rice protoplasts. The CBE_V04 systems were found to have improved editing activity and purity with focal recruitment of more uracil DNA glycosylase inhibitors (UGIs) by the engineered single guide RNA 2.0 scaffold. Finally, we used whole-genome sequencing (WGS) to compare six CBE_V01 systems and four CBE_V04 systems for genome-wide off-target effects in rice. Different levels of cytidine deaminase-dependent and sgRNA-independent off-target effects were indeed revealed by WGS among edited lines by these CBE systems. We also investigated genome-wide sgRNA-dependent off-target effects by different CBEs in rice. This comprehensive study compared 21 different CBE systems, and benchmarked PmCDA1-CBE_V04 and A3A/Y130F-CBE_V04 as next-generation plant CBEs with high editing efficiency, purity, and specificity.

CRISPR-Cas12a is a promising genome editing system for targeting AT-rich genomic regions. Comprehensive genome engineering requires simultaneous targeting of multiple genes at defined locations. Here, to expand the targeting scope of Cas12a, we screen nine Cas12a orthologs that have not been demonstrated in plants, and identify six, ErCas12a, Lb5Cas12a, BsCas12a, Mb2Cas12a, TsCas12a and MbCas12a, that possess high editing activity in rice. Among them, Mb2Cas12a stands out with high editing efficiency and tolerance to low temperature. An engineered Mb2Cas12a-RVRR variant enables editing with more relaxed PAM requirements in rice, yielding two times higher genome coverage than the wild type SpCas9. To enable large-scale genome engineering, we compare 12 multiplexed Cas12a systems and identify a potent system that exhibits nearly 100% biallelic editing efficiency with the ability to target as many as 16 sites in rice. This is the highest level of multiplex edits in plants to date using Cas12a. Two compact single transcript unit CRISPR-Cas12a interference systems are also developed for multi-gene repression in rice and Arabidopsis. This study greatly expands the targeting scope of Cas12a for crop genome engineering.

Pyroptosis is inflammation-associated caspase-1-dependent programmed cell death, which confers a crucial role in sepsis. The present study intends to investigate the regulatory network and function of the microarray-predicted YTHDF1 in caspase-1-dependent pyroptosis of sepsis. Peripheral blood of patients with sepsis was collected to determine WWP1 and YTHDF1 expression. An in vitro sepsis cell model was induced in RAW264.7 cells using lipopolysaccharide (LPS) and ATP and an in vivo septic mouse model by cecal ligation and perforation (CLP). After gain- and loss-of-function assays in vitro and in vivo, TNF-α and IL-1β levels and the cleavage of gasdermin-D (GSDMD) were detected by ELISA and Western blot assay, followed by determination of lactate dehydrogenase (LDH) activity. Immunoprecipitation and meRIP assay were performed to detect the ubiquitination of NLRP3 and the m6A modification of WWP1 mRNA. The binding of WWP1 to YTHDF1 was explored using RIP-RT-qPCR and dual luciferase gene reporter assay. It was noted that WWP1 and YTHDF1 were downregulated in clinical sepsis samples, LPS + ATP-treated RAW264.7 cells, and CLP-induced mice. The ubiquitination of NLRP3 was promoted after overexpression of WWP1. WWP1 translation could be promoted by YTHDF1. Then, WWP1 or YTHDF1 overexpression diminished LDH activity, NLRP3 inflammasomes and caspase-1-mediated cleavage of GSDMD in LPS + ATP-induced RAW264.7 cells. Overexpressed YTHDF1 restrained inflammatory response in CLP-induced mice. Collectively, the alleviatory effect of m6A reader protein YTHDF1 may be achieved through promotion of NLRP3 ubiquitination and inhibition of caspase-1-dependent pyroptosis by upregulating WWP1.

Kidney stones have become a worldwide public health problem. The purpose of this research is to study the relationship between plasma cadmium level and the prevalence of kidney stones in an adult population.

A series of novel phosphorylated penta-1,4-dien-3-one derivatives were designed and synthesized. The structures of all title compounds were determined by ¹H-NMR, 13C-NMR, 31P-NMR, and high-resolution mass spectrometry (HRMS). Bioassay results showed that several of the title compounds exhibited remarkable antibacterial and antiviral activities. Among these, compound 3g exhibited substantial antibacterial activity against Xanthomonas oryzae pv. Oryzae (Xoo), with a 50% effective concentration (EC50) value of 8.6 μg/mL, which was significantly superior to bismerthiazol (BT) (58.8 µg/mL) and thiodiazole-copper (TC) (78.7 μg/mL). In addition, compound 3h showed remarkable protective activity against tobacco mosaic virus (TMV), with an EC50 value of 104.2 μg/mL, which was superior to that of ningnanmycin (386.2 μg/mL). Furthermore, the microscale thermophoresis and molecular docking experiments on the interaction of compounds 3h and 3j with TMV coat protein (TMV CP) were also investigated. Compounds 3h and 3j bound to TMV CP with dissociation constants of 0.028 and 0.23 μmol/L, which were better than that of ningnanmycin (0.52 μmol/L). These results suggest that novel phosphorylated penta-1,4-dien-3-one derivatives may be considered as an activator for antibacterial and antiviral agents.

Baicalin has been demonstrated to exert anticancer effects mainly through induction of tumor cell apoptosis and cell cycle arrest. However, the precise mechanisms underlying its anticancer role remain to be elucidated. In the present study, we investigated whether autophagy was involved in the anticancer activity of baicalin in the human hepatocellular carcinoma (HCC) cell line SMMC-7721 and the possible molecular mechanisms. Our data showed that the viability of SMMC-7721 cells was significantly inhibited by baicalin in a dose- and time-dependent manner. Alongside apoptosis, autophagy was also induced by baicalin dose- and time-dependently with the involvement of the autophagy-associated protein Βeclin 1. Moreover, we demonstrated that cell death induced by baicalin was significantly inhibited by the apoptosis inhibitor z-DEVD-fmk or the autophagy inhibitor 3-MA, respectively. In addition, we found that CD147, a key molecule related both to apoptosis and autophagy, was markedly downregulated at the protein level in SMMC-7721 cells treated with baicalin. Collectively, this is the first study to suggest that baicalin induces autophagic cell death in SMMC-7721 cells, which involves the downregulation of CD147. Our study reveals a new mechanism for the anticancer effects of baicalin and puts forward a potential crucial role of CD147 in baicalin-induced cancer cell death.

Animal experiments have found that oil tea reduces body weight and improves blood lipid levels. However, the effect of oil tea on human health has not been confirmed yet. This study aims to explore the relationship between oil tea consumption and obesity and dyslipidemia.

Hand grip strength is a complex phenotype. The current study aimed to identify the effects of the association between APOE rs405509 polymorphisms and gene-environment interactions on hand grip strength among middle-aged and elderly people in a rural population in Gongcheng, southern China.

Background: Advanced glycation end products (AGEs) impair vascular physiology in Diabetes mellitus (DM). However, the underlying mechanisms remain unclear. Vascular large conductance calcium-activated potassium (BK) channels play important roles in coronary arterial function.Purpose: Our study aimed to investigate the regulatory role of AGEs in BK channels.Research Design: Using gavage of vehicle (V, normal saline) or aminoguanidine (A) for 8 weeks, normal and diabetic rats were divided into four groups: C+V group, DM+V group, C+A group, and DM+A group.Study Sample: Coronary arteries from different groups of rats and human coronary smooth muscle cells were used in this study.Data Collection and Analysis: Data were presented as mean ± SEM (standard error of mean). Student's t-test was used to compare data between two groups. One-way ANOVA with post-hoc LSD analysis was used to compare data between multiple groups.Results: Compared to the C+V group, vascular contraction induced by iberiotoxin (IBTX), a BK channel inhibitor, was impaired, and BK channel densities decreased in the DM+V group. However, aminoguanidine administration reduced the impairment. Protein expression of BK-β1, phosphorylation of adenosine 5'-monophosphate-activated protein kinase (AMPK), and protein kinase B (PKB or Akt) were down-regulated, while F-box protein 32 (FBXO32) expression increased in the DM+V group and in high glucose (HG) cultured human coronary smooth muscle cells. Treatment with aminoguanidine in vitro and in vivo could reverse the above protein expression. The effect of aminoguanidine on the improvement of BK channel function by inhibiting the generation of AGEs was reversed by adding MK2206 (Akt inhibitor) or Compound C (AMPK inhibitor) in HG conditions in vitro.Conclusions: AGEs aggravate BK channel dysfunction via the AMPK/Akt/FBXO32 signaling pathway.

In secretory epithelial cells, the basolateral Na(+)-K(+)-2Cl(-) cotransporter (NKCC1) plays a major role in salt and fluid secretion. Our laboratory has identified NKCC1 surface expression as an important regulatory mechanism for Cl(-) secretion in the colonic crypt cell line T84, a process also present in native human colonic crypts. We previously showed that activation of protein kinase C (PKC) by carbachol and phorbol 12-myristate 13-acetate (PMA) decreases NKCC1 surface expression in T84 cells. However, the specific endocytic entry pathway has not been defined. We used a Madin-Darby canine kidney (MDCK) cell line stably transfected with enhanced green fluorescent protein (EGFP)-NKCC1 to map NKCC1 entry during PMA exposure. At given times, we fixed and stained the cells with specific markers (e.g., dynamin II, clathrin heavy chain, and caveolin-1). We also used chlorpromazine, methyl-beta-cyclodextrin, amiloride, and dynasore, blockers of the clathrin, caveolin, and macropinocytosis pathways and the vesicle "pinchase" dynamin, respectively. We found that PMA caused dose- and time-dependent NKCC1 endocytosis. After 2.5 min of PMA exposure, approximately 80% of EGFP-NKCC1 endocytic vesicles colocalized with clathrin and approximately 40% colocalized with dynamin II and with the transferrin receptor, the uptake of which is also mediated by clathrin-coated vesicles. We did not observe significant colocalization of EGFP-NKCC1 endocytic vesicles with caveolin-1, a marker of the caveolae-mediated endocytic pathway. We quantified the effect of each inhibitor on PMA-induced EGFP-NKCC1 endocytosis and found that only chlorpromazine and dynasore caused significant inhibition compared with the untreated control (61% and 25%, respectively, at 2.5 min). Together, these results strongly support the conclusion that PMA-stimulated NKCC1 endocytosis is associated with a clathrin pathway.

PAM-relaxed Cas9 nucleases, cytosine base editors and adenine base editors are promising tools for precise genome editing in plants. However, their genome-wide off-target effects are largely unexplored. Here, we conduct whole-genome sequencing (WGS) analyses of transgenic plants edited by xCas9, Cas9-NGv1, Cas9-NG, SpRY, nCas9-NG-PmCDA1, nSpRY-PmCDA1 and nSpRY-ABE8e in rice. Our results reveal that Cas9 nuclease and base editors, when coupled with the same guide RNA (gRNA), prefer distinct gRNA-dependent off-target sites. De novo generated gRNAs by SpRY editors lead to additional, but insubstantial, off-target mutations. Strikingly, ABE8e results in ~500 genome-wide A-to-G off-target mutations at TA motif sites per transgenic plant. ABE8e's preference for the TA motif is also observed at the target sites. Finally, we investigate the timeline and mechanism of somaclonal variation due to tissue culture, which chiefly contributes to the background mutations. This study provides a comprehensive understanding on the scale and mechanisms of off-target and background mutations occurring during PAM-relaxed genome editing in plants.

MicroRNA168 (MIR168) is a key miRNA that targets the main RNA-induced silencing complex component Argonaute 1 (AGO1) to regulate plant growth and environmental stress responses. However, the regulatory functions of MIR168 need to be further elucidated in rice. In this paper, we generated clean OsMIR168a deletion mutants by CRISPR-Cas9 strategy. We then phenotypically and molecularly characterized these mutants. The rice OsMIR168a mutants grew rapidly at the seedling stage, produced more tillers and matured early. Compared to the wild-type plants, the mutants were shorter at maturity and produced smaller spikelets and seeds. Analysis of gene expression showed that the transcription levels of OsMIR168a's target genes such as OsAGO1a, OsAGO1b and OsAGO1d were elevated significantly in the OsMIR168a mutants. Intriguingly, OsAGO18, a member of a new AGO clade that is conserved in monocots, was confirmed to be a target of OsMIR168a not only by informatic prediction but also by expression analysis and a cell-based cleavage assay in the OsMIR168a mutants. Many protein-coding genes and miRNAs showed differential expression in the OsMIR168a mutants, suggesting OsMIR168a exerts a major transcriptional regulatory role, likely through its potential target genes such as OsAGO1s and OsAGO18. KEGG enrichment analysis of these differentially expressed genes pointed to OsMIR168a's involvement in important processes such as plant hormone signalling transduction and plant-pathogen interaction. These data collectively support that the complex regulation module of OsMIR168a-OsAGO1/OsAGO18-miRNAs-target genes contributes to agronomically important traits, which sheds light on miRNA-mediated crop breeding.

Background: The study aimed to investigate the relationship between transcription factor EB (TFEB) gene polymorphisms, including their haplotypes, and the cognitive functions of a selected population in Gongcheng County, Guangxi. Methods: A case-control study approach was used. The case group comprised 339 individuals with cognitive impairment, as assessed by their Mini-Mental State Examination scores; the control population also comprised 339 individuals who were matched by sex and age (± 5 years) in a 1:1 ratio. TFEB gene polymorphisms were genotyped in 678 participants (190 men and 488 women, aged 30-91 years) by using the Sequenom MassARRAY platform. Results: Multifactorial logistic regression analysis showed that in the dominant model, the risk of developing cognitive impairment was 1.547 times higher in cases with the TFEB rs14063A allele (AG + AA) than in those with the GG genotype (adjusted odds ratio [OR] = 1.547, Bonferroni correction confidence interval = 1.021-2.345). Meanwhile, the presence of the TFEB rs1062966T allele (CT + TT) was associated with a lower risk of cognitive impairment in comparison with the presence of the CC genotype (adjusted OR = 0.636, Bonferroni correction confidence interval = 0.405-0.998). In the co-dominant model, the risk of developing cognitive impairment was 1.553 times higher in carriers of the TFEB rs14063AG genotype than in carriers of the GG genotype (adjusted OR = 1.553, Bonferroni correction confidence interval = 1.007-2.397). After the Bonferroni correction and adjustment for confounding factors, the association of TFEB rs1062966 with cognitive function persisted in the analyses stratified by education level. Ethnically stratified analysis showed a significant association between TFEB rs1062966 and cognitive function in the Yao population. The multilocus linkage disequilibrium analysis indicated that the identified single nucleotide polymorphisms were not inherited independently. The haplotype analysis suggested that the rs14063A-rs1062966C-rs2278068C-rs1015149T haplotype of the TFEB gene increased the risk of cognitive impairment (P < 0.05) and that the rs14063G-rs1062966T-rs2278068C-rs1015149C haplotype was associated with a reduced risk of cognitive impairment (P < 0.05). Conclusion: TFEB rs1062966 polymorphisms and their rs14063A-rs1062966C-rs2278068C-rs1015149T and rs14063G-rs1062966T-rs2278068C-rs1015149C haplotypes are genetic factors that may affect cognitive function among the rural Chinese population.

Fatty liver disease (FLD) is a serious public health problem that is rapidly increasing. Evidence indicates that the transcription factor EB (TFEB) gene may be involved in the pathophysiology of FLD; however, whether TEFB polymorphism has an association with FLD remains unclear.