Revolutionizing aquatic eco-environmental monitoring: Utilizing the RPA-Cas-FQ detection platform for zooplankton.

The integration of recombinase polymerase amplification (RPA) with CRISPR/Cas technology has revolutionized molecular diagnostics and pathogen detection due to its unparalleled sensitivity and trans-cleavage ability. However, its potential in the ecological and environmental monitoring scenarios for aquatic ecosystems remains largely unexplored, particularly in accurate qualitative/quantitative detection, and its actual performance in handling complex real environmental samples. Using zooplankton as a model, we have successfully optimized the RPA-CRISPR/Cas12a fluorescence detection platform (RPA-Cas-FQ), providing several crucial "technical tips". Our findings indicate the sensitivity of CRISPR/Cas12a alone is 5 × 109 copies/reaction, which can be dramatically increased to 5 copies/reaction when combined with RPA. The optimized RPA-Cas-FQ enables reliable qualitative and semi-quantitative detection within 50 min, and exhibits a good linear relationship between fluorescence intensity and DNA concentration (R2 = 0.956-0.974***). Additionally, we developed a rapid and straightforward identification procedure for single zooplankton by incorporating heat-lysis and DNA-barcode techniques. We evaluated the platform's effectiveness using real environmental DNA (eDNA) samples from the Three Gorges Reservoir, confirming its practicality. The eDNA-RPA-Cas-FQ demonstrated strong consistency (Kappa = 0.43***) with eDNA-Metabarcoding in detecting species presence/absence in the reservoir. Furthermore, the two semi-quantitative eDNA technologies showed a strong positive correlation (R2 = 0.58-0.87***). This platform also has the potential to monitor environmental pollutants by selecting appropriate indicator species. The novel insights and methodologies presented in this study represent a significant advancement in meeting the complex needs of aquatic ecosystem protection and monitoring.

Effects of light qualities on growth and physiological-biochemical traits of Scutellaria baicalensis.

Canopy spectral composition significantly affects growth and functional traits of understory plants. In this study, we explored the optimal light condition suitable for enhancing Scutellaria baicalensis's yield and quality, aiming to provide scientific reference for the exploitation and utilization of medicinal plant resources in the understory of forests. We measured the responses of growth, morphology, biomass allocation, physiological traits, and secon-dary metabolites of S. baicalensis to different light qualities. S. baicalensis was cultured under five LED-light treatments including full spectrum light (control), ultraviolet-A (UV-A) radiation, blue, green, and red light. Results showed that UV-A significantly reduced plant height, base diameter, leaf thickness, leaf area ratio, and biomass of each organ. Red light significantly reduced base diameter, biomass, effective quantum yield of photosystem Ⅱ (ФPSⅡ), and total flavonoid concentration. Under blue light, root length and total biomass of S. baicalensis significantly increased by 48.0% and 10.8%, respectively, while leaf number and chlorophyll content significantly decreased by 20.0% and 31.6%, respectively. The other physiological and biochemical traits were consistent with their responses in control. Our results suggested that blue light promoted photosynthesis, biomass accumulation, and secondary metabolite synthesis of S. baicalensis, while red light and UV-A radiation negatively affected physiological and biochemical metabolic processes. Therefore, the ratio of blue light could be appropriately increased to improve the yield and quality of S. baicalensis.

The complete plastomes of thirteen Libanotis (Apiaceae, Apioideae) plants: comparative and phylogenetic analyses provide insights into the plastome evolution and taxonomy of Libanotis.

BACKGROUND: The genus Libanotis Haller ex Zinn, nom. cons., a contentious member of Apiaceae, encompasses numerous economically and medicinally significant plants, comprising approximately 30 species distributed across Eurasia. Despite many previous taxonomic insights into it, phylogenetic studies of the genus are still lacking. And the establishment of a robust phylogenetic framework remains elusive, impeding advancements and revisions in the taxonomic system for this genus. Plastomes with greater variability in their genetic characteristics hold promise for building a more robust Libanotis phylogeny. RESULTS: During our research, we sequenced, assembled, and annotated complete plastomes for twelve Libanotis species belong to three sections and two closely related taxa. We conducted a comprehensive comparative analysis through totally thirteen Libanotis plastomes for the genus, including an additional plastome that had been published. Our results suggested that Libanotis plastome was highly conserved between different subclades, while the coding regions were more conserved than the non-coding regions, and the IR regions were more conserved than the single copy regions. Nevertheless, eight mutation hotspot regions were identified among plastomes, which can be considered as candidate DNA barcodes for accurate species identification in Libanotis. The phylogenetic analyses generated a robustly framework for Libanotis and revealed that Libanotis was not a monophyletic group and their all three sections were polygenetic. Libanotis schrenkiana was sister to L. sibirica, type species of this genus, but the remainders scattered within Selineae. CONCLUSION: The plastomes of Libanotis exhibited a high degree of conservation and was effective in enhancing the support and resolution of phylogenetic analyses within this genus. Based on evidence from both phylogeny and morphology, we propose the recognition of "Libanotis sensu stricto" and provide taxonomic recommendations for other taxa that previously belonged to Libanotis. In conclusion, our study not only revealed the phylogenetic position and plastid evolution of Libanotis, but also provided new insights into the phylogeny of the family Apiaceae and phylogenetic relationships within the tribe Selineae.

Villosol reverses 5-FU resistance in colorectal cancer by inhibiting the CDKN2A gene regulated TP53-PI3K/Akt signaling axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Patrinia villosa (Juss.) (PV) is the drug of choice in traditional Chinese medicine for the treatment of colorectal cancer (CRC) and has achieved reliable efficacy in clinic. Villosol is the active ingredient in PV. However, the molecular mechanism by which Villosol reverses chemoresistance in CRC remains unclear. AIM OF THE STUDY: Analysis of the molecular mechanism by which Villosol, the active ingredient of PV, reverses CRC/5-FU resistance through modulation of the CDKN2A gene was validated by network pharmacology techniques and experiments. MATERIALS AND METHODS: We identified CDKN2A as a gene associated with 5-FU resistance through gene chip analysis. Next, we conducted a series of functional analyses in cell lines, animal samples, and xenograft models to investigate the role, clinical significance, and abnormal regulatory mechanisms of CDKN2A in 5-FU resistance in CRC. In addition, we screened and obtained a raw ingredient called Villosol, which targets CDKN2A, and investigated its pharmacological effects. RESULTS: Analysis of CRC cells and animal samples showed that the upregulation of CDKN2A expression was strongly associated with 5-FU resistance. CRC cells overexpressing CDKN2A showed reduced sensitivity to 5-FU and enhanced tumor biology in vitro. Inhibition of aberrant activation of CDKN2A enhances the expression of TP53. Mechanistically, overexpression of CDKN2A activates the PI3K/Akt pathway and induces resistance to 5-FU. Villosol inhibited CDKN2A, and CRC/5-FU cells regained sensitivity to 5-FU. Villosol effectively reverses 5-FU resistance through the CDKN2A-TP53-PI3K/Akt axis. CONCLUSION: Changes in CDKN2A gene expression can be used to predict the response of CRC patients to 5-FU therapy. Additionally, inhibiting CDKN2A activation with Villosol may present a new approach to overcoming 5-FU resistance in clinical settings.

Transcutaneous electric acupoint stimulation reduced consumption of profopol in patients undergoing laparoscopic surgery: A randomized clinical trial.

BACKGROUND: Propofol is a common regimen for general anesthesia maintenance. But propofol can dose-dependently generate cardiopulmonary depression. Thus, any strategy to reduce propofol dosage during laparoscopic surgery may have underlying beneficial effect for patient prognosis. Previous studies indicated both analgesic and sedative effect of acupoint stimulation. However, its main sedative effect on patients under general anesthesia remains unclear. OBJECTIVE: The aim of this study was to investigate the sedative effect of transcutaneous electrical acupoint stimulation (TEAS) on patients scheduled for laparoscopic surgery under general anesthesia. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: In this randomized clinical trial, patients scheduled for laparoscopic surgery under general anesthesia in Xijing hospital were randomly assigned to 3 groups, receiving electrical stimulation at the Shenmen (HT7)/Ximen (PC4) (TEAS group), stimulation at the shoulder (non-acupoint group) or no stimulation (control group), respectively. MAIN OUTCOME MEASURES: One hundred sixty-two patients completed the study. The primary outcome was the consumption of propofol, and secondary outcomes included features of recovery after surgery, major complications after surgery and by 1 year after surgery. RESULTS: In patients undergoing laparoscopic surgery, the doses of propofol decreased significantly in the TEAS group compared (0.10 ±â€…0.02 mg·kg-1·min-1) with the other 2 groups (both 0.12 ±â€…0.02 mg·kg-1·min-1, P < .001). The mean differences (95% confidence interval) for non-acupoint versus TEAS and control versus TEAS were 0.021 (0.012, 0.030) and 0.024 (0.013, 0.034), respectively. Time to awake and to extubation were not significantly different among the groups. The incidences of major complications after surgery and by 1 year after surgery were not significantly different among the groups. CONCLUSION: TEAS could induce additional sedative effect in patients during laparoscopic surgery and reduce propofol consumption.

SPL16 and SPL23 mediate photoperiodic control of seasonal growth in Populus trees.

Perennial trees in boreal and temperate regions undergo growth cessation and bud set under short photoperiods, which are regulated by phytochrome B (phyB) photoreceptors and PHYTOCHROME INTERACTING FACTOR 8 (PIF8) proteins. However, the direct signaling components downstream of the phyB-PIF8 module remain unclear. We found that short photoperiods suppressed the expression of miR156, while upregulated the expression of miR156-targeted SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE 16 (SPL16) and SPL23 in leaves and shoot apices of Populus trees. Accordingly, either overexpression of MIR156a/c or mutagenesis of SPL16/23 resulted in the attenuation of growth cessation and bud set under short days (SD), whereas overexpression of SPL16 and SPL23 conferred early growth cessation. We further showed that SPL16 and SPL23 directly suppressed FLOWERING LOCUS T2 (FT2) expression while promoted BRANCHED1 (BRC1.1 and BRC1.2) expression. Moreover, we revealed that PIF8.1/8.2, positive regulators of growth cessation, directly bound to promoters of MIR156a and MIR156c and inhibited their expression to modulate downstream pathways. Our results reveal a connection between the phyB-PIF8 module-mediated photoperiod perception and the miR156-SPL16/23-FT2/BRC1 regulatory cascades in SD-induced growth cessation. Our study provides insights into the rewiring of a conserved miR156-SPL module in the regulation of seasonal growth in Populus trees.

Tartary Buckwheat (Fagopyrum tataricum) FtTT8 Inhibits Anthocyanin Biosynthesis and Promotes Proanthocyanidin Biosynthesis.

Tartary buckwheat (Fagopyrum tataricum) is an important plant, utilized for both medicine and food. It has become a current research hotspot due to its rich content of flavonoids, which are beneficial for human health. Anthocyanins (ATs) and proanthocyanidins (PAs) are the two main kinds of flavonoid compounds in Tartary buckwheat, which participate in the pigmentation of some tissue as well as rendering resistance to many biotic and abiotic stresses. Additionally, Tartary buckwheat anthocyanins and PAs have many health benefits for humans and the plant itself. However, little is known about the regulation mechanism of the biosynthesis of anthocyanin and PA in Tartary buckwheat. In the present study, a bHLH transcription factor (TF) FtTT8 was characterized to be homologous with AtTT8 and phylogenetically close to bHLH proteins from other plant species. Subcellular location and yeast two-hybrid assays suggested that FtTT8 locates in the nucleus and plays a role as a transcription factor. Complementation analysis in Arabidopsis tt8 mutant showed that FtTT8 could not recover anthocyanin deficiency but could promote PAs accumulation. Overexpression of FtTT8 in red-flowering tobacco showed that FtTT8 inhibits anthocyanin biosynthesis and accelerates proanthocyanidin biosynthesis. QRT-PCR and yeast one-hybrid assay revealed that FtTT8 might bind to the promoter of NtUFGT and suppress its expression, while binding to the promoter of NtLAR and upregulating its expression in K326 tobacco. This displayed the bidirectional regulating function of FtTT8 that negatively regulates anthocyanin biosynthesis and positively regulates proanthocyanidin biosynthesis. The results provide new insights on TT8 in Tartary buckwheat, which is inconsistent with TT8 from other plant species, and FtTT8 might be a high-quality gene resource for Tartary buckwheat breeding.

Prelimbic area to lateral hypothalamus circuit drives social aggression.

Controlling aggression is a vital skill in social species such as rodents and humans and has been associated with the medial prefrontal cortex (mPFC). In this study, we showed that during aggressive behavior, the activity of GABAergic neurons in the prelimbic area (PL) of the mPFC was significantly suppressed. Specific activation of GABAergic PL neurons significantly curbed male-to-male aggression and inhibited conditioned place preference (CPP) for aggression-paired contexts, whereas specific inhibition of GABAergic PL neurons brought about the opposite effect. Moreover, GABAergic projections from PL neurons to the lateral hypothalamus (LH) orexinergic neurons mediated aggressive behavior. Finally, directly modulated LH-orexinergic neurons influence aggressive behavior. These results suggest that GABAergic PL-orexinergic LH projection is an important control circuit for intermale aggressive behavior, both of which could be targets for curbing aggression.

Neurotoxicity and the potential molecular mechanisms of mono-2-ethylhexyl phthalic acid (MEHP) in zebrafish.

Mono-2-ethylhexyl phthalic acid (MEHP) is the most toxic metabolite of plasticizer di-2-ethylhexyl phthalic acid (DEHP), and there is limited information available on the effects of MEHP on neurotoxicity. This study aims to examine the neurotoxicity of MEHP and preliminarily explore its potential molecular mechanisms. We found that MEHP impeded the growth of zebrafish embryos and the neurodevelopmental-related gene expression at environmentally relevant concentrations. MEHP exposure also induces oxidative stress response and brain cell apoptosis accompanied by a decrease in acetylcholinesterase (AChE) activity in zebrafish larvae. RNA-Seq and bioinformatics analysis showed that MEHP treatment altered the nervous system, neurogenic diseases, and visual perception pathways. The locomotor activity in dark-to-light cycles and phototaxis test confirmed the abnormal neural behavior of zebrafish larvae. Besides, the immune system has produced a large number of differentially expressed genes related to neural regulation. Inflammatory factor IL1ß and IL-17 signaling pathways highly respond to MEHP, indicating that inflammation caused by immune system imbalance is a potential mechanism of MEHP-induced neurotoxicity. This study expands the understanding of the toxicity and molecular mechanisms of MEHP, providing a new perspective for in-depth neurotoxicity exploration of similar compounds.

Akkermansia muciniphila supplementation prevents cognitive impairment in sleep-deprived mice by modulating microglial engulfment of synapses.

The microbiome-gut-brain axis plays a crucial role in many neurological diseases, including mild cognitive impairment. Sleep deprivation (SD) induces cognitive decline accompanied by alterations in the gut microbiota. However, the role of gut microbiota alterations in SD-induced cognitive dysfunction and the underlying mechanisms remain unclear. Here, we found that dysbiosis of the gut microbiota following pretreatment with broad-spectrum antibiotics worsens SD-induced cognitive impairment in mice. Fecal microbiota transplantation from SD mice to healthy mice induced cognitive impairment. Additionally, the abundance of Akkermansia muciniphila (A. muciniphila) in the mouse gut microbiota was significantly reduced after 7 days of SD. A. muciniphila pretreatment alleviated cognitive dysfunction and prevented synaptic reduction in the hippocampus in SD mice. A. muciniphila pretreatment inhibited extensive microglial activation and synaptic engulfment in the hippocampus of SD mice. Metabolomics analysis revealed that A. muciniphila pretreatment increased the serum acetate and butanoic acid levels in SD mice. Finally, pretreatment with short-chain fatty acids (SCFAs) inhibited microglial synaptic engulfment and prevented neuronal synaptic loss in SD mice and primary microglia-neuron co-culture following LPS stimulation. Together, our findings illustrate that gut dysbiosis plays an essential role in SD-induced cognitive impairment by activating microglial engulfment at synapses. A. muciniphila supplementation may be a novel preventative strategy for SD-induced cognitive dysfunction, by increasing SCFAs production and maintaining microglial homeostasis.

Recent advances in micro (nano) plastics in the environment: Distribution, health risks, challenges and future prospects.

Environmental micro(nano)plastics have become a significant global pollution problem due to the widespread use of plastic products. In this review, we summarized the latest research advances on micro(nano)plastics in the environment, including their distribution, health risks, challenges, and future prospect. Micro(nano)plastics have been found in a variety of environmental media, such as the atmosphere, water bodies, sediment, and especially marine systems, even in remote places like Antarctica, mountain tops, and the deep sea. The accumulation of micro(nano)plastics in organisms or humans through ingestion or other passive ways poses a series of negative impacts on metabolism, immune function, and health. Moreover, due to their large specific surface area, micro(nano)plastics can also adsorb other pollutants, causing even more serious effects on animal and human health. Despite the significant health risks posed by micro(nano)plastics, there are limitations in the methods used to measure their dispersion in the environment and their potential health risks to organisms. Therefore, further research is needed to fully understand these risks and their impacts on the environment and human health. Taken together, the challenges of micro(nano)plastics analysis in the environment and organisms must be addressed, and future research prospects need to be identified. Governments and individuals must take action to reduce plastic waste and minimize the negative impact of micro(nano)plastics on the environment and human health.

Proteomic and metabolomic analyses showing the differentially accumulation of NnUFGT2 is involved in the petal red-white bicolor pigmentation in lotus (Nelumbo nucifera).

Bicolor flower lotus is rare with high ornamental value. During the long history of breeding and artificial selection, a very famous lotus cultivar 'Da Sajin' with red and white picotee bicolor petals were obtained. In order to reveal the mechanism underlying the formation of its picotee bicolor pattern in the petal, an integrative metabolomics and proteomics analyses were conducted between red and white parts of its petals. The results showed that the defect of anthocyanidin 3-O-glucosyltransferases (UFGTs) accumulation resulted in the failure of the glycosylation of anthocyanidin, the last step of anthocyanin biosynthesis in white part of the petals. And proteomic data and biochemical analysis showed that the defect of UFGTs accumulation is not related to their transcription, but because of their degradation. Function of one differentially accumulated NnUFGT were proven being involved in anthocyanin biosynthesis through both in-vitro enzyme assay and in-vivo transgenic analyses. This regulation on the protein accumulation of structural genes in anthocyanin biosynthesis was not explored in any other plants, and hence supposed to be a novel mechanism for the formation of picotee bicolor pattern flower. The results not only provide some new insights into the understanding of lotus flower coloration, but also might assist the breeding of flower lotus.

One-stop combined CT angiography of coronary and craniocervical arteries: recommended as the first examination for patients suspected of coronary or craniocervical artery disease.

OBJECTIVES: To investigate the potential diagnostic value of one-stop combined CT angiography (CTA) as the first examination for patients suspected of coronary artery disease (CAD) or craniocervical artery disease (CCAD), and compare its clinical performance with two consecutive CTA scans. METHODS: Patients with suspected but unconfirmed CAD or CCAD were prospectively enrolled and grouped randomly to undergo coronary and craniocervical CTA using the combined protocol (group 1) or the consecutive protocol (group 2). Diagnostic findings were evaluated for both the targeted and non-targeted regions. The objective image quality, overall scan time, radiation dose, and contrast medium dosage were compared between the two groups. RESULTS: Each group enrolled 65 patients. A substantial number of lesions were found in non-targeted regions, which was 44/65 (67.7%) by patients for group 1 and 41/65 (63.1%) for group 2, reiterating the necessity of extending the scan coverage. Specifically, lesions in non-targeted regions were detected more often for patients suspected of CCAD than for those suspected of CAD (71.4% vs 61.7%). With 21.5% (~51.1 s) reduction of scan time and 21.8% (~20.8 mL) less contrast medium as compared to the consecutive protocol, high-quality images were obtained by the combined protocol. CONCLUSIONS: One-stop combined CTA enables effective detection of lesions in non-targeted regions at a lower cost of scan time and contrast medium than two separate examinations and is thus worth taking as the first examination for patients suspected of CAD or CCAD. KEY POINTS: • Extending the scan range for coronary or craniocervical CTA has the potential to reveal lesions in non-targeted regions. • One-stop combined CTA as enabled on high-speed wide-detector CT delivers high-quality images at a lower cost of contrast medium and operational time than two consecutive CTA scans. • Patients with suspected but unconfirmed CAD or CCAD may benefit from the one-stop combined CTA in the first examination.

Integration of metabolomics and transcriptomics analyses investigates the accumulation of secondary metabolites in maturing seed plumule of sacred lotus (Nelumbo nucifera).

Lotus seed plumule (LP) is rich in a variety of antioxidant and anti-inflammatory secondary metabolites, making it a traditional food and medicine widely used in China. Physiological and histological evidences indicated that LP mainly accumulated metabolites in 15-24 days after pollination (DAP) during their development. To systematically investigate the dynamic accumulation of major secondary metabolites, the UPLC-HRMS-based widely targeted metabolomics analyses were performed on maturing LP at 15, 18, 21, and 24 DAP. In total, 767 metabolites were identified, including many secondary metabolites, e.g., 27 % flavonoids and 8 % alkaloids. Among them, 591 were identified as differentially accumulated metabolites (DAMs). The majority of secondary metabolites showed great accumulation after 18 DAP even at the late stage of LP maturation, such as hesperidin, neohesperidin, orobol, serotonin, and lotus special O-nornuciferine, endowing mature LP with effective pharmaceutical properties. The paralleled transcriptomic analysis identified 11,019 differentially expressed genes (DEGs). Based on the comprehensive data, several systematical metabolic regulation maps were established for different secondary metabolites, and 18 DAP was found as a switching point for LP maturing from active primary metabolism to massive secondary metabolites deposition. This study provides valuable information for understanding the mechanism of secondary metabolite accumulation in maturing LP and facilitates its pharmaceutical application.

QTL mapping and candidate gene analysis for yield and grain weight/size in Tartary buckwheat.

BACKGROUND: Grain weight/size influences not only grain yield (GY) but also nutritional and appearance quality and consumer preference in Tartary buckwheat. The identification of quantitative trait loci (QTLs)/genes for grain weight/size is an important objective of Tartary buckwheat genetic research and breeding programs. RESULTS: Herein, we mapped the QTLs for GY, 1000-grain weight (TGW), grain length (GL), grain width (GW) and grain length-width ratio (L/W) in four environments using 221 recombinant inbred lines (XJ-RILs) derived from a cross of 'Xiaomiqiao × Jinqiaomai 2'. In total, 32 QTLs, including 7 for GY, 5 for TGW, 6 for GL, 11 for GW and 3 for L/W, were detected and distributed in 24 genomic regions. Two QTL clusters, qClu-1-3 and qClu-1-5, located on chromosome Ft1, were revealed to harbour 7 stable major QTLs for GY (qGY1.2), TGW (qTGW1.2), GL (qGL1.1 and qGL1.4), GW (qGW1.7 and qGW1.10) and L/W (qL/W1.2) repeatedly detected in three and above environments. A total of 59 homologues of 27 known plant grain weight/size genes were found within the physical intervals of qClu-1-3 and qClu-1-5. Six homologues, FtBRI1, FtAGB1, FtTGW6, FtMADS1, FtMKK4 and FtANT, were identified with both non-synonymous SNP/InDel variations and significantly differential expression levels between the two parents, which may play important roles in Tatary buckwheat grain weight/size control and were chosen as core candidate genes for further investigation. CONCLUSIONS: Two stable major QTL clusters related to grain weight/size and six potential key candidate genes were identified by homology comparison, SNP/InDel variations and qRT‒qPCR analysis between the two parents. Our research provides valuable information for improving grain weight/size and yield in Tartary buckwheat breeding.

Research progress on the intrinsic non-immune function of PD-L1 in tumors (Review).

Programmed death ligand 1 (PD-L1) is widely expressed in human tumors. It is widely known for its immunosuppressive function as it can help tumor cells evade T cell immune killing through the PD-1/PD-L1 signal. A number of clinical trials have proved that the destruction of the combination of PD-1 and PD-L1 by antibodies could significantly affect patients with advanced cancer. However, a number of patients with cancer still cannot benefit from PD-1/PD-L1 blocking therapy. The main reason is that PD-L1 also has some intrinsic regulatory functions to promote the progression of tumors. PD-L1 Protein contains an intrinsic domain that could link to other signal pathways, but the mechanism has not yet been fully revealed. The present review mainly discussed the non-immune checkpoint functions of PD-L1, such as its role in regulating cell proliferation, cell metabolism, drug resistance and maintaining epithelial-mesenchymal transition and stemness.

Electroacupuncture pretreatment induces ischemic tolerance by neuronal TREM2-mediated enhancement of autophagic flux.

The mechanism of electroacupuncture (EA) pretreatment-induced neuroprotection remains unclear. In this study, we found that neuronal Triggering receptor expressed on myeloid cells 2 (TREM2) expression was increased and peaked at 48 h and 72 h after ischemia/reperfusion. After specific knockdown of TREM2 in excitatory neurons, neurological function was damaged, and the infarct volume was enlarged. Furthermore, the expression of LC3II/LC3I and Beclin1 was decreased, while the expression of p62 was increased. EA pretreatment enhanced TREM2, LC3II/LC3I and Beclin1 expression while reducing p62 in the ischemic penumbra area. The EA-induced neuroprotective effects and improvements in autophagic flux were abolished by specific knockdown of TREM2 in excitatory neurons. Taken together, our findings provide novel mechanistic insight into EA-induced ischemic tolerance and suggest a promising therapeutic strategy of targeting neuronal TREM2 to treat brain ischemia.

Competition between H4PteGlu and H2PtePAS Confers para-Aminosalicylic Acid Resistance in Mycobacterium tuberculosis.

Tuberculosis remains a serious challenge to human health worldwide. para-Aminosalicylic acid (PAS) is an important anti-tuberculosis drug, which requires sequential activation by Mycobacterium tuberculosis (M. tuberculosis) dihydropteroate synthase and dihydrofolate synthase (DHFS, FolC). Previous studies showed that loss of function mutations of a thymidylate synthase coding gene thyA caused PAS resistance in M. tuberculosis, but the mechanism is unclear. Here we showed that deleting thyA in M. tuberculosis resulted in increased content of tetrahydrofolate (H4PteGlu) in bacterial cells as they rely on the other thymidylate synthase ThyX to synthesize thymidylate, which produces H4PteGlu during the process. Subsequently, data of in vitro enzymatic activity experiments showed that H4PteGlu hinders PAS activation by competing with hydroxy dihydropteroate (H2PtePAS) for FolC catalysis. Meanwhile, over-expressing folC in ΔthyA strain and a PAS resistant clinical isolate with known thyA mutation partially restored PAS sensitivity, which relieved the competition between H4PteGlu and H2PtePAS. Thus, loss of function mutations in thyA led to increased H4PteGlu content in bacterial cells, which competed with H2PtePAS for catalysis by FolC and hence hindered the activation of PAS, leading to decreased production of hydroxyl dihydrofolate (H2PtePAS-Glu) and finally caused PAS resistance. On the other hand, functional deficiency of thyA in M. tuberculosis pushes the bacterium switch to an unidentified dihydrofolate reductase for H4PteGlu biosynthesis, which might also contribute to the PAS resistance phenotype. Our study revealed how thyA mutations confer PAS resistance in M. tuberculosis and provided new insights into studies on the folate metabolism of the bacterium.