Chloroplast C-to-U editing, regulated by a PPR protein BoYgl-2, is important for chlorophyll biosynthesis in cabbage.

Leaf color is an important agronomic trait in cabbage (Brassica oleracea L. var. capitata), but the detailed mechanism underlying leaf color formation remains unclear. In this study, we characterized a Brassica oleracea yellow-green leaf 2 (BoYgl-2) mutant 4036Y, which has significantly reduced chlorophyll content and abnormal chloroplasts during early leaf development. Genetic analysis revealed that the yellow-green leaf trait is controlled by a single recessive gene. Map-based cloning revealed that BoYgl-2 encodes a novel nuclear-targeted P-type PPR protein, which is absent in the 4036Y mutant. Functional complementation showed that BoYgl-2 from the normal-green leaf 4036G can rescue the yellow-green leaf phenotype of 4036Y. The C-to-U editing efficiency and expression levels of atpF, rps14, petL and ndhD were significantly reduced in 4036Y than that in 4036G, and significantly increased in BoYgl-2 overexpression lines than that in 4036Y. The expression levels of many plastid- and nuclear-encoded genes associated with chloroplast development in BoYgl-2 mutant were also significantly altered. These results suggest that BoYgl-2 participates in chloroplast C-to-U editing and development, which provides rare insight into the molecular mechanism underlying leaf color formation in cabbage.

Astrocyte activation in hindlimb somatosensory cortex contributes to electroacupuncture analgesia in acid-induced pain.

Background: Several studies have confirmed the direct relationship between extracellular acidification and the occurrence of pain. As an effective pain management approach, the mechanism of electroacupuncture (EA) treatment of acidification-induced pain is not fully understood. The purpose of this study was to assess the analgesic effect of EA in this type of pain and to explore the underlying mechanism(s). Methods: We used plantar injection of the acidified phosphate-buffered saline (PBS; pH 6.0) to trigger thermal hyperalgesia in male Sprague-Dawley (SD) rats aged 6-8 weeks. The value of thermal withdrawal latency (TWL) was quantified after applying EA stimulation to the ST36 acupoint and/or chemogenetic control of astrocytes in the hindlimb somatosensory cortex. Results: Both EA and chemogenetic astrocyte activation suppressed the acid-induced thermal hyperalgesia in the rat paw, whereas inhibition of astrocyte activation did not influence the hyperalgesia. At the same time, EA-induced analgesia was blocked by chemogenetic inhibition of astrocytes. Conclusion: The present results suggest that EA-activated astrocytes in the hindlimb somatosensory cortex exert an analgesic effect on acid-induced pain, although these astrocytes might only moderately regulate acid-induced pain in the absence of EA. Our results imply a novel mode of action of astrocytes involved in EA analgesia.

Performance Improvement for the CuCrZr Alloy Produced by Laser Powder Bed Fusion Using the Remelting Process.

Owing to the high optical reflectivity of copper powder, the high-performance fabrication of copper alloys in the laser additive manufacturing (AM) field is problematic. To tackle this issue, this study employs the remelting process during laser powder bed fusion AM to fabricate defect-free and high-performance CuCrZr alloy. Compared to the non-remelting process, the remelting process yields finer grains, smaller precipitates, denser dislocations, and smaller dislocation cells. It realizes not only the dense molding of high laser reflectivity powders but also excellent mechanical properties and electrical conductivity (with an ultimate tensile strength of 329 MPa and conductivity of 96% IACS) without post-heat treatment. Furthermore, this study elucidates the influence of complex thermal gradients and multiple thermal cycles on the manufacturing process under the remelting process, as well as the internal mechanisms of microstructure evolution and performance improvement.

Taxonomic and functional ß-diversity patterns reveal stochastic assembly rules in microbial communities of seagrass beds.

Microorganisms are important members of seagrass bed ecosystems and play a crucial role in maintaining the health of seagrasses and the ecological functions of the ecosystem. In this study, we systematically quantified the assembly processes of microbial communities in fragmented seagrass beds and examined their correlation with environmental factors. Concurrently, we explored the relative contributions of species replacement and richness differences to the taxonomic and functional ß-diversity of microbial communities, investigated the potential interrelation between these components, and assessed the explanatory power of environmental factors. The results suggest that stochastic processes dominate community assembly. Taxonomic ß-diversity differences are governed by species replacement, while for functional ß-diversity, the contribution of richness differences slightly outweighs that of replacement processes. A weak but significant correlation (p < 0.05) exists between the two components of ß-diversity in taxonomy and functionality, with almost no observed significant correlation with environmental factors. This implies significant differences in taxonomy, but functional convergence and redundancy within microbial communities. Environmental factors are insufficient to explain the ß-diversity differences. In conclusion, the assembly of microbial communities in fragmented seagrass beds is governed by stochastic processes. The patterns of taxonomic and functional ß-diversity provide new insights and evidence for a better understanding of these stochastic assembly rules. This has important implications for the conservation and management of fragmented seagrass beds.

Cevanine-type alkaloids from the bulbs of Fritillaria unibracteata var. wabuensis and their antifibrotic activities in vitro.

Steroidal alkaloids are the main bioactive components of the bulbs of Fritillaria, which have been used as traditional Chinese medicine, known as "Beimu", for the treatment of cough for thousands of years in China. Cough and dyspnea are the most common symptoms observed in patients with pulmonary fibrosis. However, the antifibrotic activity of steroidal alkaloids has not been reported yet. In this study, two previously unreported cevanine-type steroidal alkaloids (1 and 2), four previously undescribed cevanine-type alkaloid glycosides (3-6), and 19 known steroidal alkaloids (7-25) were isolated from the bulbs of Fritillaria unibracteata var. wabuensis. The structures of these compounds were elucidated by comprehensive HRESIMS and NMR spectroscopic data analysis, as well as DP4+ NMR calculations. The biological evaluation showed that compounds 2, 7-10, 14, 15, and 17 downregulated fibrotic markers induced by transforming growth factor-ß (TGF-ß) in MRC-5 cells. Moreover, compounds 14 and 17 dose dependently inhibited TGF-ß-induced epithelial-mesenchymal transition in A549 cells, alleviated TGF-ß-induced migration and proliferation of fibroblasts, and decreased the expression of fibrotic markers, fibronectin, and N-cadherin in TGF-ß-induced MRC-5 cells. The research showed the potential of cevanine-type alkaloids as a class of natural antifibrotic agents.

Emerging functions and therapeutic targets of IL-38 in central nervous system diseases.

Interleukin (IL)-38 is a newly discovered cytokine of the IL-1 family, which binds various receptors (i.e., IL-36R, IL-1 receptor accessory protein-like 1, and IL-1R1) in the central nervous system (CNS). The hallmark physiological function of IL-38 is competitive binding to IL-36R, as does the IL-36R antagonist. Emerging research has shown that IL-38 is abnormally expressed in the serum and brain tissue of patients with ischemic stroke (IS) and autism spectrum disorder (ASD), suggesting that IL-38 may play an important role in neurological diseases. Important advances include that IL-38 alleviates neuromyelitis optica disorder (NMOD) by inhibiting Th17 expression, improves IS by protecting against atherosclerosis via regulating immune cells and inflammation, and reduces IL-1ß and CXCL8 release through inhibiting human microglial activity post-ASD. In contrast, IL-38 mRNA is markedly increased and is mainly expressed in phagocytes in spinal cord injury (SCI). IL-38 ablation attenuated SCI by reducing immune cell infiltration. However, the effect and underlying mechanism of IL-38 in CNS diseases remain inadequately characterized. In this review, we summarize the biological characteristics, pathophysiological role, and potential mechanisms of IL-38 in CNS diseases (e.g., NMOD, Alzheimer's disease, ASD, IS, TBI, and SCI), aiming to explore the therapeutic potential of IL-38 in the prevention and treatment of CNS diseases.

Ablation of histone methyltransferase Suv39h2 in hepatocytes attenuates NASH in mice.

AIMS: Non-alcoholic steatohepatitis (NASH) is characterized by aberrant lipid metabolism in hepatocytes. We investigated the involvement of a histone H3K9 methyltransferase Suv39h2 in the pathogenesis of NASH. METHODS AND MATERIALS: NASH is induced by feeding the mice with a high-fat high-carbohydrate (HFHC) diet or a high-fat choline-deficient amino acid defined (HFD-CDAA) diet. The Suv39h2f/f mice were crossbred with the Alb-Cre mice to specifically delete Suv39h2 in hepatocytes. KEY FINDINGS: Ablation of Suv39h2 in hepatocytes improved insulin sensitivity of the mice fed either the HFHC diet or the CDAA-HFD diet. Importantly, Suv39h2 deletion significantly ameliorated NAFLD as evidenced by reduced lipid accumulation, inflammation, and fibrosis in the liver. RNA-seq uncovered Vanin-1 (Vnn1) as a novel transcriptional target for Suv39h2. Mechanistically, Suv39h2 repressed Vnn1 transcription in hepatocytes exposed to free fatty acids. Consistently, Vanin-1 knockdown normalized lipid accumulation in Suv39h2-null hepatocytes. Importantly, a significant correlation between Suv39h2, Vanin-1, and hepatic triglyceride levels was identified in NASH patients. SIGNIFICANCE: Our study uncovers a novel mechanism whereby Suv39h2 may contribute to NASH pathogenesis and suggests that targeting the Suv39h2-Vanin-1 axis may yield novel therapeutic solutions against NASH.

Alkaloid uptake pathways in renal tubular epithelial cells from different processed products of Phellodendri chinensis Cortex.

This study aimed to investigate the absorption of alkaloids from Phellodendri chinensis Cortex (PC) by human renal tubular epithelial cells (HK-2). Cellular uptake and affinity ultrafiltration assays were employed to determine the alkaloid uptake pathway in HK-2 cells. Stemming from the hypothesis that salt-water processed PC introduces these alkaloids into the kidney at a cellular level, this research focused on different processed products of PC that are tailored for renal targeting. Utilizing the UPLC-QqQ-MS method, we quantified variations in the uptake capacity of phellodendrine, magnoflorine, jatrorrhizine, berberrubine, and berberine from raw Phellodendri chinensis Cortex (RPC), salt-water processed Phellodendri chinensis Cortex (SPC), and wine processed Phellodendri chinensis Cortex (WPC) in HK-2 cells. This study also tracked the concentration changes of these five alkaloids in HK-2 cells during the administration phase. Further, we evaluated the influence of two inhibitors on the absorption of these five alkaloids from PC and its processed products into HK-2 cells: the organic anion transporters (OATs) inhibitor-probenecid (PRO), and the organic cationic transporters (OCTs) inhibitor-tetraethylammonium chloride (TEAC). A pivotal component of this research was an investigation into the effects of PC and its processed products on the expression levels of OCT2, OAT1, and OAT3 proteins in HK-2 cells, facilitated by Western blot analysis. Finally, we appraised the binding affinity of PC's alkaloids to OCT2, OAT1, and OAT3 proteins using an ultrafiltration centrifugation technique. The uptake of different processed products of PC by HK-2 cells showed the following trend: SPC group > RPC group > WPC group. When considering inhibitor uptake in HK-2 cells, the group treated with PRO (an OATs inhibitor) demonstrated a higher uptake than the group treated with TEAC (an OCTs inhibitor). It was observed that different processed products of PC elevated the expression of OCT2 and OAT1 proteins in HK-2 cells. Specifically, both the SPC and berberrubine groups displayed enhanced expression of these proteins, with a marked increase noted for OCT2. Through affinity ultrafiltration assays, it was determined that the binding affinity of alkaloids from different processed products of PC to OCT2 and OAT1 significantly exceeded that to OAT3. These results indicate that PC-derived alkaloids are absorbed by HK-2 cells, predominantly through transport mechanisms mediated by OCT2 and OAT1, with OCT2 serving as the dominant transporter. The higher intake of alkaloids in SPC group can likely be linked to the amplified activity of kidney uptake transporters.

Atrachinenins D-S, novel meroterpenoids with geranyl hydroquinone moiety from Atractylodes chinensis by the LC/MS-based molecular decoy and targeted isolation.

To mine fascinating molecules from the rhizomes of Atractylodes chinensis, the known molecular formula of atrachinenin A was used as a bait to search LC-HRMS data in different subfractions. Sixteen new meroterpenoids, atrachinenins D-S (1-16) including three unprecedented carbon skeletons (1-5) and eleven new oxygen-bridged hybrids (6-16) were obtained by the targeted isolation. Their structures and absolute configurations were elucidated by the spectroscopic data and electronic circular dichroism (ECD) calculations. The isolated compounds were evaluated for their inhibitory activity of NO production and compounds 1, 4, 8, and 13 showed moderate anti-inflammatory activity. The proposed biosynthetic pathways of 1-5 were also discussed.

Rapid and sensitive detection of nucleic acids using an RAA-CRISPR/Cas12b one-pot detection assay (Rcod).

Rapid, sensitive and specific methods are crucial for nucleic acid detection. CRISPR/Cas12b has recently been widely used in nucleic acid detection. However, due to its thermophagic property, DNA isothermal recombinase-aided amplification (RAA) and subsequent CRISPR/Cas12b detection require two separate reactions, which is cumbersome and inconvenient and may cause aerosol pollution. In this study, we propose an RAA-CRISPR/Cas12b one-pot detection assay (Rcod) for Bordetella pertussis detection without additional amplification product transfer steps. The time from sample processing to response time was less than 30 min using nucleic acid extraction-free method, and the sensitivity reached 0.2 copies/µL. In this system, Alicyclobacillus acidoterrestris Cas12b protein (AacCas12b) exhibited strong and specific trans-cleavage activity at a constant temperature of 37 °C, while the cis-cleavage activity was weak. This characteristic reduces the interference of AacCas12b with nucleic acids in the system. Compared with real-time PCR, our Rcod system detected B. pertussis in 221 clinical samples with a sensitivity and specificity of 97.96 % and 99.19 %, respectively, with nucleic acid extraction-free method. The rapid, sensitive and specific Rcod system provides ideas for the establishment of CRISPR-based one-step nucleic acid detection and may aid the development of reliable point-of-care nucleic acid tests. IMPORTANCE: Pertussis is an acute respiratory infection caused by B. pertussis that is highly contagious and potentially fatal, and early diagnosis is essential for the treatment of whooping cough. In this study, we found that AacCas12b has high and strongly specific trans-cleavage activity at lower temperatures. A RAA-CRISPR/Cas12b one-step detection platform (Rcod) without interference with amplification was developed. In addition, the combination of Rcod and nucleic acid extraction-free method can quickly and accurately detect the qualitative detection of B. pertussis, and the detection results are visualized, which makes the pathogen nucleic acid detection and analysis process simpler, and provides a new method for the rapid clinical diagnosis of B. pertussis.

Clinical evaluation of premonitory urges in children and adolescents using the Chinese version of Individualized Premonitory Urge for Tics Scale.

Background: Premonitory urges (PUs) have been the focus of recent efforts to assess the severity and develop interventions for tic disorders (TD). We aimed to investigate the PUs in TD and its comorbidities from multiple dimensions, using the Chinese version of the Premonitory Urge for Tics Scale (C-PUTS) and the Chinese version of the Individualized Premonitory Urge for Tics Scale (C-IPUTS), in order to provide perspectives for the diagnosis and management of TD in children and adolescents. Methods: A total of 123 cases were included in the study. The IPUTS was translated, back-translated, culturally adjusted, and pre-investigated to determine the items of the C-IPUTS. The reliability and validity of the C-IPUTS scale were evaluated by a questionnaire survey on children and adolescents with TD at the Developmental Pediatrics Department of the Second Hospital of Jilin University. Meanwhile, the C-PUTS, which had been evaluated and used in China, Yale Global Tic Severity Scale (YGTSS), Yale-Brown Obsessive-Compulsive Scale (Y-BOCS), Depression Self-Rating Scale (DSRS), Screen for Childhood Anxiety-Related Disorders (SCARED), Achenbach Child Behavior Checklist (CBCL), and Swanson, Nolan and Pelham, Version IV (SNAP-IV), were used to assess the association of PUs with tics and comorbidities of TD. Results: All dimensions of the C-IPUTS demonstrated good reliability and validity. Our findings suggested that PUs in children and adolescents in China occurred primarily at the head/face and neck/throat. The different dimensions of the C-IPUTS (number, frequency, and intensity) and C-PUTS were positively correlated with the YGTSS total score, while the C-PUTS was positively correlated with the Y-BOCS, SCARED, DSRS, and SNAP-IV scale total scores. The three dimensions of the C-IPUTS demonstrated correlations with anxiety severity and obsessive-compulsive symptoms. Conclusion: The C-IPUTS can be used to assess PUs reliably and effectively and provide further information for the C-PUTS from various dimensions in a Chinese setting. PUs relate to obsessive-compulsive symptoms, anxiety, attention deficit hyperactivity, and behavioral problems in children and adolescents with TDs. Accordingly, PUs evaluation using the C-IPUTS combined with the PUTS might provide useful information for future therapies for TDs to achieve greater tic reduction.

A Whole-Genome Assembly for Hyaloperonospora parasitica, A Pathogen Causing Downy Mildew in Cabbage (Brassica oleracea var. capitata L.).

Hyaloperonospora parasitica is a global pathogen that can cause leaf necrosis and seedling death, severely threatening the quality and yield of cabbage. However, the genome sequence and infection mechanisms of H. parasitica are still unclear. Here, we present the first whole-genome sequence of H. parasitica isolate BJ2020, which causes downy mildew in cabbage. The genome contains 4631 contigs and 9991 protein-coding genes, with a size of 37.10 Mb. The function of 6128 genes has been annotated. We annotated the genome of H. parasitica strain BJ2020 using databases, identifying 2249 PHI-associated genes, 1538 membrane transport proteins, and 126 CAZy-related genes. Comparative analyses between H. parasitica, H.arabidopsidis, and H. brassicae revealed dramatic differences among these three Brassicaceae downy mildew pathogenic fungi. Comprehensive genome-wide clustering analysis of 20 downy mildew-causing pathogens, which infect diverse crops, elucidates the closest phylogenetic affinity between H. parasitica and H. brassicae, the causative agent of downy mildew in Brassica napus. These findings provide important insights into the pathogenic mechanisms and a robust foundation for further investigations into the pathogenesis of H. parasitica BJ2020.

An extraction-free method for rapid detection of CYP2C19 * 2/3/17 polymorphisms in one tube using melting curve analysis.

Drug-metabolizing enzymes play an important role in the metabolism of drugs in vivo. Their activity is an important factor affecting the rate of drug metabolism, which directly determines the intensity and persistence of drug action. Patients taking medication can be divided into different metabolic types through detection of CYP2C19 drug-metabolizing enzyme gene polymorphisms, which can then be used for medication guidance for clopidogrel. Here, we describe a detection method based on real-time polymerase chain reaction (PCR). This method uses multicolor melting curve analysis to accurately identify different mutation sites and genotypes of CYP2C19 * 2, CYP2C19 * 3, and CYP2C19 * 17. The detection limit of plasmid samples was 1 copies µL-1 ; that of genomic samples was 0.1 ng µL-1 . The system can detect nine types of CYP2C19 * 2/3/17 at three sites in one tube, quickly achieving detection within 1 h. Combined with the sample release agent, sample extraction was completed in 5 s, achieving rapid diagnosis without extraction for timely diagnosis and treatment. Furthermore, the system is not limited to blood samples and can also be applied to oropharyngeal and saliva samples, increasing sampling diversity and convenience. When using clinical blood samples (n = 93), the detection system we established was able to quickly and accurately identify different genotypes, and the accuracy and effectiveness of the detection were confirmed by Sanger sequencing. Due to its accuracy, rapidity, simple operation, and low cost, detection technology based on real-time polymerase amplification combined with melting curve analysis is expected to become a powerful tool for detecting and guiding clopidogrel use in countries with limited resources.

The REM microarousal and REM duration as the potential indicator in paradoxical insomnia.

OBJECTIVE: Although paradoxical insomnia is a prevalent subtype of chronic insomnia, the etiology of it is unclear. Contrary to complaints of little or no sleep, polysomnography (PSG) findings show that paradoxical insomnia patients have near normal sleep macrostructure. The purpose of this study is to determine the changes of microstructure and explore the etiology of paradoxical insomnia. METHODS: The PSG findings of 89 paradoxical insomnia patients were compared with those of 41 gender balanced healthy controls without sleep complaints. All subjects underwent nocturnal PSG recordings. Conventional PSG measures and microarousals were quantified and statistically analyzed. Receiver operating characteristic curve and correlation analysis were used to evaluate the potential of REM sleep microarousals and REM duration as indicators of paradoxical insomnia. RESULTS: Compared with the controls, paradoxical insomnia patients had no significant differences in sleep macrostructures. Statistical analysis showed that non-rapid eye movement (NREM) microarousals revealed no significant differences between paradoxical insomnia patients and controls. Noticeably, more spontaneous microarousals appeared in rapid eye movement (REM) stage for paradoxical insomnia patients. Based on receiver operating characteristic curve (ROC), the optimal cutoff value of REM sleep microarousals could predict paradoxical insomnia. Furthermore, a positive correlation between microarousals in REM sleep and the duration of REM sleep was presented in paradoxical insomnia patients. CONCLUSIONS: The frequency of REM microarousals and the duration of REM sleep could reflect the real sleep state of paradoxical insomnia patients. That suggested PSG investigation extended to microarousal could be helpful to understand the etiology in paradoxical insomnia.

Persistence of human- and cattle-associated Bacteroidales and mitochondrial DNA markers in freshwater mesocosms.

Accurate identification of the origins of non-point source pollution is essential for the effective control of fecal pollution. Host-associated Bacteroidales and mitochondrial DNA (mtDNA) markers have been developed to identify the sources of human and cattle fecal pollution. However, the differences in persistence between these two types of markers under different environmental conditions are still poorly understood. Here, we conducted mesocosm experiments to investigate the influence of indigenous microbiota and nutrients on the decay of Bacteroidales and mtDNA markers associated with humans and cattle. Raw sewage or cattle feces were inoculated into mesocosms containing natural eutrophic water, sterile eutrophic water or artificial freshwater. The Bacteroidales markers HF183 (human) and CowM3 (cattle) and mtDNA markers HcytB (human) and QMIBo (cattle) were quantified using the quantitative polymerase chain reaction (qPCR) assays. All markers but HF183 decreased the fastest in the presence of indigenous microbiota. Nutrients caused a decrease in the persistence of HF183; however, no significant nutrient effects were observed for HcytB, CowM3, and QMIBo. The time to reach one log reduction (T90) for HF183 and HcytB was similar; CowM3 reached T90 earlier than QMIBo in all the treatments but eutrophic water. E. coli persisted longer than both Bacteroidales and mtDNA markers in the mesocosms regardless of inoculum type. Additionally, 16S rRNA gene amplicon sequencing was used to determine the changes in bacterial communities accompanying the marker decay. Analysis using the SourceTracker software showed that bacterial communities in the mesocosms became more dissimilar to those in the corresponding inoculants over time. Our results indicate that environmental factors are important determinants of genetic markers' persistence, but their impact can vary depending on the genetic markers. The cattle Bacteroidales markers may be more suitable for determining recent fecal contamination than cattle mtDNA.

Ectopic over-expression of HaFT-1, a 14-3-3 protein from Haloxylon ammodendron, enhances acquired thermotolerance in transgenic Arabidopsis.

Haloxylon ammodendron, an important shrub utilized for afforestation in desert areas, can withstand harsh ecological conditions such as drought, high salt and extreme heat. A better understanding of the stress adaptation mechanisms of H. ammodendron is vital for ecological improvement in desert areas. In this study, the role of the H. ammodendron 14-3-3 protein HaFT-1 in thermotolerance was investigated. qRT-PCR analysis showed that heat stress (HS) priming (the first HS) enhanced the expression of HaFT-1 during the second HS and subsequent recovery phase. The subcellular localization of YFP-HaFT-1 fusion protein was mainly detected in cytoplasm. HaFT-1 overexpression increased the germination rate of transgenic Arabidopsis seeds, and the survival rate of HaFT-1 overexpression seedlings was higher than that of wild-type (WT) Arabidopsis after priming-and-triggering and non-primed control treatments. Cell death staining showed that HaFT-1 overexpression lines exhibited significantly reduced cell death during HS compared to WT. Transcriptome analysis showed that genes associated with energy generation, protein metabolism, proline metabolism, autophagy, chlorophyll metabolism and reactive oxygen species (ROS) scavenging were important to the thermotolerance of HS-primed HaFT-1 transgenic plants. Growth physiology analysis indicated that priming-and-triggering treatment of Arabidopsis seedlings overexpressing HaFT-1 increased proline content and strengthened ROS scavenging activity. These results demonstrated that overexpression of HaFT-1 increased not only HS priming but also tolerance to the second HS of transgenic Arabidopsis, suggesting that HaFT-1 is a positive regulator in acquired thermotolerance.

Di- and Triterpenoids from the Rhizomes of Isodon amethystoides and Their Anti-inflammatory Activities.

Amethystoidesic acid (1), a triterpenoid with an unprecedented 5/6/6/6 tetracyclic skeleton, and six undescribed diterpenoids, amethystoidins A-F (2-7), were isolated from the rhizomes of Isodon amethystoides along with 31 known di- and triterpenoids (8-38). Their structures were fully elucidated via extensive spectroscopic analysis including 1D and 2D NMR, high-resolution electrospray ionization mass spectrometry (HRESIMS), and electronic circular dichroism (ECD) calculations. Compound 1 is the first example of a triterpenoid possessing a rare ring system (5/6/6/6) derived from a contracted A-ring and the 18,19-seco-E-ring of ursolic acid. Compounds 6, 16, 21, 22, 24, and 27 significantly inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated RAW264.7 cells, which could be partly mediated by the downregulation of LPS-induced inducible nitric oxide synthase (iNOS) protein expression.

P2X7 receptor-mediated depression-like reactions arising in the mouse medial prefrontal cortex.

Major depressive disorder is a frequent and debilitating psychiatric disease. We have shown in some of the acute animal models of major depressive disorder (tail suspension test and forced swim test) that depression-like behavior can be aggravated in mice by the microinjection into the medial prefrontal cortex of the P2X7R agonistic adenosine 5'-triphosphate or its structural analog dibenzoyl-ATP, and these effects can be reversed by the P2X7R antagonistic JNJ-47965567. When measuring tail suspension test, the prolongation of immobility time by the P2YR agonist adenosine 5'-[ß-thio]diphosphate and the reduction of the adenosine 5'-(γ-thio)triphosphate effect by P2Y1R (MRS 2179) or P2Y12R (PSB 0739) antagonists, but not by JNJ-47965567, all suggest the involvement of P2YRs. In order to elucidate the localization of the modulatory P2X7Rs in the brain, we recorded current responses to dibenzoyl-ATP in layer V astrocytes and pyramidal neurons of medial prefrontal cortex brain slices by the whole-cell patch-clamp procedure; the current amplitudes were not altered in preparations taken from tail suspension test or foot shock-treated mice. The release of adenosine 5'-triphosphate was decreased by foot shock, although not by tail suspension test both in the hippocampus and PFC. In conclusion, we suggest, that in the medial prefrontal cortex, acute stressful stimuli cause supersensitivity of P2X7Rs facilitating the learned helplessness reaction.

Extended characterization of IL-33/ST2 as a predictor for wound age determination in skin wound tissue samples of humans and mice.

Interleukin (IL)-33, an important inflammatory cytokine, is highly expressed in skin wound tissue and serum of humans and mice, and plays an essential role in the process of skin wound healing (SWH) dependent on the IL-33/suppression of tumorigenicity 2 (ST2) pathway. However, whether IL-33 and ST2 themselves, as well as their interaction, can be applied for skin wound age determination in forensic practice remains incompletely characterized. Human skin samples with injured intervals of a few minutes to 24 hours (hs) and mouse skin samples with injured intervals of 1 h to 14 days (ds) were collected. Herein, the results demonstrated that IL-33 and ST2 are increased in the human skin wounds, and that in mice skin wounds, there is an increase over time, with IL-33 expression peaking at 24 hs and 10 ds, and ST2 expression peaking at 12 hs and 7 ds. Notably, the relative quantity of IL-33 and ST2 proteins < 0.35 suggested a wound age of 3 hs; their relative quantity > 1.0 suggested a wound age of 24 hs post-mouse skin wounds. In addition, immunofluorescent staining results showed that IL-33 and ST2 were consistently expressed in the cytoplasm of F4/80-positive macrophages and CD31-positive vascular endothelial cells with or without skin wounds, whereas nuclear localization of IL-33 was absent in α-SMA-positive myofibroblasts with skin wounds. Interestingly, IL-33 administration facilitated the wound area closure by increasing the proliferation of cytokeratin (K) 14 -positive keratinocytes and vimentin-positive fibroblasts. In contrast, treating with its antagonist (i.e., anti-IL-33) or receptor antagonist (e.g., anti-ST2) exacerbated the aforementioned pathological changes. Moreover, treatment with IL-33 combined with anti-IL-33 or anti-ST2 reversed the effect of IL-33 on facilitating skin wound closure, suggesting that IL-33 administration facilitated skin wound closure through the IL-33/ST2 signaling pathway. Collectively, these findings indicate that the detection of IL-33/ST2 might be a reliable biomarker for the determination of skin wound age in forensic practice.

Comparative Transcriptome Analysis Reveals a Potential Regulatory Network for Ogura Cytoplasmic Male Sterility in Cabbage (Brassica oleracea L.).

Ogura cytoplasmic male sterility (CMS) lines are widely used breeding materials in cruciferous crops and play important roles in heterosis utilization; however, the sterility mechanism remains unclear. To investigate the microspore development process and gene expression changes after the introduction of orf138 and Rfo, cytological observation and transcriptome analysis were performed using a maintainer line, an Ogura CMS line, and a restorer line. Semithin sections of microspores at different developmental stages showed that the degradation of tapetal cells began at the tetrad stage in the Ogura CMS line, while it occurred at the bicellular microspore stage to the tricellular microspore stage in the maintainer and restorer lines. Therefore, early degradation of tapetal cells may be the cause of pollen abortion. Transcriptome analysis results showed that a total of 1287 DEGs had consistent expression trends in the maintainer line and restorer line, but were significantly up- or down-regulated in the Ogura CMS line, indicating that they may be closely related to pollen abortion. Functional annotation showed that the 1287 core DEGs included a large number of genes related to pollen development, oxidative phosphorylation, carbohydrate, lipid, and protein metabolism. In addition, further verification elucidated that down-regulated expression of genes related to energy metabolism led to decreased ATP content and excessive ROS accumulation in the anthers of Ogura CMS. Based on these results, we propose a transcriptome-mediated induction and regulatory network for cabbage Ogura CMS. Our research provides new insights into the mechanism of pollen abortion and fertility restoration in Ogura CMS.