Occurrence of organotin compounds in food: increasing challenge of phenyltin compounds.

Concentrations and distribution for 16 organotin compounds were studied in all kinds of foods, including seafood, agricultural products, and wine. Meanwhile, the degradation of the TBT or TPhT was also evaluated. Concentrations of total organotins in seafood, agricultural products, and wine were 1047.2, 469.4, and 13.5 µg Sn/kg. Meanwhile, the most frequently detected organotin in three kinds of samples were TPhT, MPhT, and MPhT, respectively. The results demonstrated that phenyltin may probably become an emerging organotin pollutant. Regarding seafood, organotin concentrations of fish and mollusks were much higher than those of crustaceans. At the same time, a significant positive correlation was observed between the concentrations of TBT and MBT (p < 0.05), and between DBT and MBT(p < 0.0001). Moreover, TPhT was significantly and positively associated with DPhT (p < 0.0001), suggesting that TPhT was the precursor of DPhT. Apart from the likely illegal use of OTs as biocides in antifouling paints for ships, anthropogenic activity like agricultural activity or industrial activity also caused organotin contamination. Further research and more effective measures should be formulated to protect the food safety. Meanwhile, monitoring of the organotin contamination should not only in Qinhuangdao, but also expand to the cities along Bohai Bay.

Genome-wide identification and characterization of members of the LEA gene family in Panax notoginseng and their transcriptional responses to dehydration of recalcitrant seeds.

BACKGROUND: Late embryogenesis abundant (LEA) proteins play an important role in dehydration process of seed maturation. The seeds of Panax notoginseng (Burkill) F. H. Chen are typically characterized with the recalcitrance and are highly sensitive to dehydration. However, it is not very well known about the role of LEA proteins in response to dehydration stress in P. notoginseng seeds. We will perform a genome-wide analysis of the LEA gene family and their transcriptional responses to dehydration stress in recalcitrant P. notoginseng seeds. RESULTS: In this study, 61 LEA genes were identified from the P. notoginseng genome, and they were renamed as PnoLEA. The PnoLEA genes were classified into seven subfamilies based on the phylogenetic relationships, gene structure and conserved domains. The PnoLEA genes family showed relatively few introns and was highly conserved. Unexpectedly, the LEA_6 subfamily was not found, and the LEA_2 subfamily contained 46 (75.4%) members. Within 19 pairs of fragment duplication events, among them 17 pairs were LEA_2 subfamily. In addition, the expression of the PnoLEA genes was obviously induced under dehydration stress, but the germination rate of P. notoginseng seeds decreased as the dehydration time prolonged. CONCLUSIONS: We found that the lack of the LEA_6 subfamily, the expansion of the LEA_2 subfamily and low transcriptional levels of most PnoLEA genes might be implicated in the recalcitrant formation of P. notoginseng seeds. LEA proteins are essential in the response to dehydration stress in recalcitrant seeds, but the protective effect of LEA protein is not efficient. These results could improve our understanding of the function of LEA proteins in the response of dehydration stress and their contributions to the formation of seed recalcitrance.

Exogenous gibberellic acid shortening after-ripening process and promoting seed germination in a medicinal plant Panax notoginseng.

BACKGROUND: Panax notoginseng (Burk) F.H. Chen is an essential plant in the family of Araliaceae. Its seeds are classified as a type of morphophysiological dormancy (MPD), and are characterized by recalcitrance during the after-ripening process. However, it is not clear about the molecular mechanism on the after-ripening in recalcitrant seeds. RESULTS: In this study, exogenous supply of gibberellic acid (GA3) with different concentrations shortened after-ripening process and promoted the germination of P. notoginseng seeds. Among the identified plant hormone metabolites, exogenous GA3 results in an increased level of endogenous hormone GA3 through permeation. A total of 2971 and 9827 differentially expressed genes (DEGs) were identified in response to 50 mg L-1 GA3 (LG) and 500 mg L-1 GA3 (HG) treatment, respectively, and the plant hormone signal and related metabolic pathways regulated by GA3 was significantly enriched. Weighted gene co-expression network analysis (WGCNA) revealed that GA3 treatment enhances GA biosynthesis and accumulation, while inhibiting the gene expression related to ABA signal transduction. This effect was associated with higher expression of crucial seed embryo development and cell wall loosening genes, Leafy Contyledon1 (LEC1), Late Embryogenesis Abundant (LEA), expansins (EXP) and Pectinesterase (PME). CONCLUSIONS: Exogenous GA3 application promotes germination and shorts the after-ripening process of P. notoginseng seeds by increasing GA3 contents through permeation. Furthermore, the altered ratio of GA and ABA contributes to the development of the embryo, breaks the mechanical constraints of the seed coat and promotes the protrusion of the radicle in recalcitrant P. notoginseng seeds. These findings improve our knowledge of the contribution of GA to regulating the dormancy of MPD seeds during the after-ripening process, and provide new theoretical guidance for the application of recalcitrant seeds in agricultural production and storage.

The mechanism of medial collateral ligament repair in knee osteoarthritis based on the TLR4/MyD88/NF-κB inflammatory signaling pathway.

OBJECTIVES: To explore the role of medial collateral ligament repair in knee osteoarthritis based on TLR4/ MyD88/ NF-κ inflammatory signaling pathway. METHODS: The modified Hulth method was used to establish models, which were divided into a repair group, a model group, and a sham operation group. The repair group was treated with medial ligament repair technology. Synovium and cartilage morphological changes were evaluated by hematoxylin-eosin staining to determine the degree of reparation. The cartilage was evaluated by the Mankin's score, and inflammatory factors in cartilage tissues were determined by ELISA. The changes in TLR4, MyD88, and NF-κB levels were analyzed using the real-time quantitative PCR and Western blot assays. RESULTS: The synovial and cartilage damages in the repair group and the sham operation group were significantly alleviated compared to the model group. The Mankin's score of the model group was significantly lower than the other two groups. The expression of inflammatory factors in the repair group and the sham operation group were significantly lower than in the model group. The expressions of those factors in the repair group and the model group were higher than those in the model group. CONCLUSIONS: Medial ligament repair can improve the cartilage morphology and delay the development and progression of knee osteoarthritis by inhibiting the TLR4/MyD88/NF-κB signaling pathway.

Wide-scope screening of pesticides in fruits and vegetables using information-dependent acquisition employing UHPLC-QTOF-MS and automated MS/MS library searching.

This paper presents an application of ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) for simultaneous screening and identification of 427 pesticides in fresh fruit and vegetable samples. Both full MS scan mode for quantification, and an artificial-intelligence-based product ion scan mode information-dependent acquisition (IDA) providing automatic MS to MS/MS switching of product ion spectra for identification, were conducted by one injection. A home-in collision-induced-dissociation all product ions accurate mass spectra library containing more than 1700 spectra was developed prior to actual application. Both qualitative and quantitative validations of the method were carried out. The result showed that 97.4 % of the pesticides had the screening detection limit (SDL) less than 50 µg kg-1 and more than 86.7 % could be confirmed by accurate MS/MS spectra embodied in the home-made library. Meanwhile, calibration curves covering two orders of magnitude were performed, and they were linear over the concentration range studied for the selected matrices (from 5 to 500 µg kg-1 for most of the pesticides). Recoveries between 80 and 110 % in four matrices (apple, orange, tomato, and spinach) at two spiked levels, 10 and 100 µg kg-1, was 88.7 or 86.8 %. Furthermore, the overall relative standard deviation (RSD, n = 12) for 94.3 % of the pesticides in 10 µg kg-1 and 98.1 % of the pesticides in 100 µg kg-1 spiked levels was less than 20 %. In order to validate the suitability for routine analysis, the method was applied to 448 fruit and vegetable samples purchased in different local markets. The results show 83.3 % of the analyzed samples have positive findings (higher than the limits of identification and quantification), and 412 commodity-pesticide combinations are identified in our scope. The approach proved to be a cost-effective, time-saving and powerful strategy for routine large-scope screening of pesticides.

Comprehensive determination of polycyclic aromatic hydrocarbons in Chinese herbal medicines by solid phase extraction and gas chromatography coupled to tandem mass spectrometry.

A simple and highly sensitive gas chromatography-tandem mass spectrometry (GC-MS/MS) method combined with solid-phase extraction cleanup was established for the comprehensive determination of 16 Environmental Protection Agency (EPA) polycyclic aromatic hydrocarbons (PAHs) in various kinds of Chinese herbal medicines (CHMs). A solid-phase extraction (SPE) purification strategy, including three parallel procedures, was developed depending on sample type, and satisfactory purification performances were achieved for all selected CHMs. The limits of detection ranged from 0.12 to 1.08 µg kg(-1) for the analyzed PAHs. The average recoveries were in the range of 65.9 % to 100.8 %, except for naphthalene (43.8 %-75.9 %), and the relative standard deviations were ≤12.8 %. The proposed method was successfully applied to the analysis of PAHs in 24 CHMs including five roots, three stems, four flowers, two fruits, four seeds, three leaves, and three barks. In the samples analyzed, all 16 PAHs are present. Their sum ranges from 21.1 to 2236.3 µg kg(-1). The entire procedure was shown to be effective and conveniently fast, and may serve as an alternative screening protocol for the determination of PAHs in CHMs.

[Application of flexible laryngeal mask airway in oral & maxillofacial day surgery].

OBJECTIVE: To access the feasibility and safety of application of flexible laryngeal mask airway (FLMA) in oral & maxillofacial day surgery. METHODS: Retrospective study was conducted of 40 oral & maxillofacial day surgery patients (3 to 61 years of age) using FLMA under general anaesthesia in Department of Oral & Maxillofacial Surgery, Peking University School and Hospital of Stomatology. All the patients were with American Society of Anesthesiologists(ASA) physical status I-II,including 19 males and 21 females. The patients'vital signs were recorded at five different time points: baseline before anesthesia (T0), time right after the FLMA insertion (T1), time at incision (T2), 15 min after incision (T3) and time at the end of the operation (T4). The first attempted FLMA insertion successful rate and the number of timed of changing to endotracheal intubation were recorded. During operation, frequencies of movement, hypoxia and obstruction of airway were noted and the operation time, anesthesia time, time from the end of the operation to extubation, movement and coughing following extubation and sore throat within 24 h were taken down. Operation-related complications of bleeding, hematoma and injury of local nerves were recorded within 24 h as well. RESULTS: The operations of all the 40 patients were successfully done under general anaesthesia. The 36(90%, 36/40)patients using FLMA successfully were under steady process, including 16 males and 20 females. The first attempted successful rate of FLMA insertion was 80% (32/40), and the second 50% (4/8). Three out of the 4 failed FLMA patients were changed to endotracheal intubation after the second attempt failed. The other patient was changed to endotracheal intubation before operation because of leak. The average operation time was (46.58 ± 22.57) min, the anesthesia time was (77.97 ± 26.82) min and the time from the end of operation to extubation was (8.31 ± 3.33) min. All the patients were recorded without obvious body movement during the operation procedure. There were 4 patients (11.11%, 4/36) with slight body movement during extubation. The incidence of sore throat was 13.89% (5/36) within 24 h postoperatively. There were no complications of bleeding, hematoma and injury of local nerves. The vital signs of baseline T0 were significantly different from those at other time points T1, T2, T3, and T4 (P<0.01). As to the hearts rate after anesthesia, the values at T1, T2, T3 and T4 for two-two comparison, there was no statistical difference (P>0.05). As to the values of systolic blood pressure and diastolic blood pressure after anesthesia at T1 and T4, T2 and T3, for two-two comparison, there was no statistical difference (P>0.05). As to the respiratory rate from the start of the surgery, the values at T2, T3 and T4 showed no statistical difference (P>0.05). CONCLUSION: Flexible laryngeal mask airway is a supraglottic airway management method. It is suitable and safe for securing the airway in oral & maxillofacial day surgery. The advantages of fewer haemodynamic changes and postoperative complications are confirmed.

Procalcitonin testing for diagnosis and short-term prognosis in bacterial infection complicated by congestive heart failure: a multicenter analysis of 4,698 cases.

INTRODUCTION: Procalcitonin (PCT) is a biomarker for the clinical diagnosis of bacterial infection that is more specific and earlier than fever, changes in white blood cell count, and blood cultures. Congestive heart failure is an important cause of endotoxin resorption from the intestine, which significantly increases PCT expression in noninfected patients with heart failure. The diagnostic performance and cut-off value of PCT in patients with bacterial infection complicated by congestive heart failure needs to be confirmed. METHODS: A total of 4,698 cases from different cities in China, including those with different classes of congestive heart failure, bacterial infection, bacterial infection complicated by heart failure and healthy individuals, were chosen for the diagnostic value analysis of PCT and screening candidate predictors of mortality in subjects with bacterial infection complicated by congestive heart failure. RESULTS: Patients with simple heart failure had significantly higher PCT levels than normal controls (P < 0.01), whereas patients with bacterial infection complicated by congestive heart failure had significantly higher PCT levels than those with simple infection (P < 0.01). Although it was useful for the diagnosis of infection (area under the receiver operating characteristic curve >80%), the positive predictive value of PCT decreased significantly with increasing severity of heart failure (P < 0.05), and the cut-off value of PCT concentrations for infection complicated by classes II, III and IV heart failure were 0.086, 0.192 and 0.657 µg/L, respectively. Heart failure degree, PCT level, and age were the candidate predictors of mortality in patients with bacterial infection complicated by congestive heart failure. CONCLUSIONS: These data suggest that complicated heart failure elevates the PCT level in patients with bacterial infection. Thus, the results of the PCT test must be analyzed correctly in consideration of the severity of heart failure. Close attention should be paid to cardiac function and PCT expression in aged patients with infection complicated by congestive heart failure.

NsdD, a GATA-type transcription factor is involved in regulation and biosynthesis of macromolecules melanin, pullulan, and polymalate in Aureobasidium melanogenum.

In this study, an NSDD gene, which encoded a GATA-type transcription factor involved in the regulation and biosynthesis of melanin, pullulan, and polymalate (PMA) in Aureobasidium melanogenum, was characterized. After the NSDD gene was completely removed, melanin production by the Δnsd mutants was enhanced, while pullulan and polymalate production was significantly reduced. Transcription levels of the genes involved in melanin biosynthesis were up-regulated while expression levels of the genes responsible for pullulan and PMA biosynthesis were down-regulated in the Δnsdd mutants. In contrast, the complementation of the NSDD gene in the Δnsdd mutants made the overexpressing mutants restore melanin production and transcription levels of the genes responsible for melanin biosynthesis. Inversely, the complementation strains, compared to the wild type strains, showed enhanced pullulan and PMA yields. These results demonstrated that the NsdD was not only a negative regulator for melanin biosynthesis, but also a key positive regulator for pullulan and PMA biosynthesis in A. melanogenum. It was proposed how the same transcriptional factor could play a negative role in melanin biosynthesis and a positive role in pullulan and PMA biosynthesis. This study provided novel insights into the regulatory mechanisms of multiple A. melanogenum metabolites and the possibility for improving its yields of some industrial products through genetic approaches.

Epithelial-Mesenchymal Transformation Promotes the Progression of Hepatocellular Carcinoma through NF-κB/MMP9 Axis.

BACKGROUND: Primary liver cancer (PHC) stands as one of the most prevalent malignant diseases in clinical settings. Studies have indicated that transcatheter arterial chemoembolization (TACE) treatment exhibits superior clinical outcomes, potentially increasing the complete necrosis rate in patients with PHC. A correlation exists between the clinical outcomes of TACE surgery and the process of epithelial-mesenchymal transition (EMT), yet the underlying mechanism remains a mystery. Hence, it is crucial to investigate the impact and mechanism of EMT on hepatocellular carcinoma (HCC). METHODS: Retrospectively, patients with advanced liver cancer who underwent TACE were selected and categorized into two groups based on the assessment of clinical efficacy: the effective group and the ineffective group. The expression levels of nuclear factor-kappa B (NF-κB), matrix metalloproteinase 9 (MMP9), Ki-67, B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X (Bax), Vimentin, E-cadherin, and N-cadherin in tumor tissues were evaluated using reverse transcription-polymerase chain reaction (RT-PCR). In vitro, Huh7 cells were cultured, and lentivirus infections were utilized to inhibit the overexpression of NF-κB and MMP9. The determination of EMT and cell viability was conducted through Cell Counting Kit-8 (CCK-8) assays, RT-PCR, and Western blot. RESULTS: Sixty patients diagnosed with advanced liver cancer were selected for the study. Based on their clinical outcomes, 30 patients with advanced hepatocellular carcinoma were categorized into the effective group, while the remaining 30 patients were categorized into the ineffective group. The results of the Western blot analysis indicated that, in comparison to the effective group, the expression levels of NF-κB, MMP9, Ki-67, Bcl-2, Vimentin, and N-cadherin were significantly higher in the tumor tissues of the ineffective group. Conversely, the expression of Bax and E-cadherin was notably lower in the effective group. Following the individual knockdown of NF-κB and MMP9, the cell experiments revealed a remarkable decrease in the expression levels of Ki-67, Bcl-2, Vimentin, and N-cadherin, whereas the expression of Bax and E-cadherin showed significant elevation (p < 0.05). Furthermore, there was a significant increase in cell viability and a decrease in cell apoptosis after the knockdown of NF-κB and MMP9. CONCLUSIONS: The NF-κB/MMP9 signaling axis serves as a pivotal regulator that fosters proliferation and impedes apoptosis in Huh7 cells by modulating the process of EMT.

Ratiometric fluorescence platform for the ultrasensitive detection of kanamycin based on split aptamer co-recognition triggers Mg2+-DNAzyme-driven DNA walker systems.

In this work, a novel 3D-DNA walker signal amplification strategy was designed to construct a fluorescent aptasensor for the detection of kanamycin (KAN). The aptasensor utilizes split aptamers for the synergistic recognition of KAN. The presence of KAN induces the split aptamers recombination to form the Mg2+-DNAzyme structure, which is activated by Mg2+ to drive the 3D-DNA walker process for cascading signal amplification. Employing gold nanoflowers (AuNFs) as walking substrate material increases the local DNA concentration to enhance the walker efficiency. The prepared fluorescent aptasensor achieved efficient and sensitive detection of KAN with satisfactory results in the concentration range of 1 × 10-8 - 1 × 10-3 µg/kg and the detection limit of 5.63 fg/kg. Meanwhile, the designed fluorescent aptasensor exhibited favorable specificity, anti-interference, storage stability and reproducibility, and verified the feasibility of its application in milk samples. The present work provides an effective tool for the regulation of KAN contamination in animal-derived foods with promising prospects.

Insights into serum metabolic biomarkers for early detection of incident diabetic kidney disease in Chinese patients with type 2 diabetes by random forest.

Diabetic kidney disease (DKD) is a leading cause of end-stage renal disease (ESRD) worldwide. Early detection is critical for the risk stratification and early intervention of progressive DKD. Serum creatinine (sCr) and urine output are used to assess kidney function, but these markers are limited by their delayed changes following kidney pathology, and lacking of both sensitivity and accuracy. Hence, it is essential to illustrate potential diagnostic indicators to enhance the precise prediction of early DKD. A total of 194 Chinese individuals include 30 healthy participants (Stage 0) and 164 incidents with type 2 diabetes (T2D) spanning from DKD's Stage 1a to 4 were recruited and their serums were subjected for untargeted metabolomic analysis. Random forest (RF), a machine learning approach, together with univariate linear regression (ULR) and multivariate linear regression (MvLR) analysis were applied to characterize the features of untargeted metabolites of DKD patients and to identify candidate DKD biomarkers. Our results indicate that 2-(α-D-mannopyranosyl)-L-tryptophan (ADT), succinyladenosine (SAdo), pseudouridine and N,N,N-trimethyl-L-alanyl-L-proline betaine (L-L-TMAP) were associated with the development of DKD, in particular, the latter three that were significantly elevated in Stage 2-4 T2D incidents. Each of the four metabolites in combination with sCr achieves better performance than sCr alone with area under the receiver operating characteristic curve (AUC) of 0.81-0.91 in predicting DKD stages. An average of 3.9 years follow-up study of another cohort including 106 Stage 2-3 patients suggested that "urinary albumin-to-creatinine ratio (UACR) + ADT + SAdo" can be utilized for better prognosis evaluation of early DKD (average AUC = 0.9502) than UACR without sexual difference.

[Effect of EGCG on the proliferation and invasion of human hepatoma HepG2 cells].

OBJECTIVE: To investigate the effect and mechanism of EGCG on the proliferation and invasion of human hepatioma HepG2 cell. METHODS: Hepatioma HepG2 cell was treated with EGCG at different concentrations. The effect of EGCG on HepG2 proliferation was examined by CCK-8. The apoptosis of HepG2 treated with EGCG was observed by fluorescence microscopy and FCM via Annexin V-FITC/PI staining. The invasion of HepG2 was detected by Transwell assay. The expression of MMP-2 and VEGF was analyzed by ELISA. RESULTS: HepG2 proliferation was inhibited after treated with EGCG. The IC25 of 24 h and 48 h was 58.19 and 54.19 mg/L. The IC50 of 24 h and 48 h was 133.90 and 78.97 mg/L. The apoptosis of HepG2 was induced significantly after treated with 60 and 135 mg/L EGCG, and the number of cells crossing Matrigel membrane was (28.33+/-7.66) and 0 (P<0.05), and the inhibitory rate of invasion was 69.47% and 100%. The expression of MMP-2 and VEGF decreased significantly. CONCLUSION: EGCG suppressed the proliferation and invasion of HepG2 with the possible mechanism of inducing apoptosis and down-regulating the expression of MMP-2 and VEGF in HepG2.

Efficacy and mechanism of lipoic acid in the treatment of reproductive injury caused by perfluorooctanoic acid.

Environmental pollutants, including perfluorooctanoic acid (PFOA), cause severe reproductive damage to humans and animals. Lipoic acid (LA) is a strong antioxidant that alleviates the oxidative damage caused by heavy metals, environmental toxicants, chemical poisons, etc. Therefore, the present study investigated the curative effect and mechanism of LA in treating spermatogenesis dysfunction caused by PFOA. A rat reproductive injury model was established by gavage of PFOA for consecutive 30 days and then rats were treated with different doses of LA for 42 days. The effects were assessed by ELISA, western blotting, H&E staining and immunofluorescence staining. The results demonstrated that LA had ameliorating effects on PFOA-induced reproductive injury in rats, it increased the sperm counts, and the levels of serum succinate dehydrogenase, lactate dehydrogenase, glutathione peroxidase and superoxide dismutase returned to normal levels following LA treatment. In addition, LA promoted the expression of estradiol, reduced the expression of serum sex hormones, such as follicle-stimulating hormone, androgen receptor (AR) and malondialdehyde in the testes, and restored the structure of seminiferous tubules. Its therapeutic mechanisms include regulating the testicular oxidative stress pathway and hypothalamic-pituitary-testis axis.

INHBA gene silencing inhibits proliferation, migration, and invasion of osteosarcoma cells by repressing TGF-ß signaling pathway activation.

BACKGROUND: Osteosarcoma (OS) is a refractory malignancy. This study aimed to explore the roles and mechanisms of Inhibin subunit beta A (INHBA) in OS. METHODS: INHBA expression levels in OS tissues and cells were assessed using RT-qPCR and western blotting. The impact of INHBA silencing on OS development was then explored by transfecting the OS cell lines U2OS and MG63 with INHBA-small interfering RNA (siRNA). The influence of INHBA silencing on U2OS and MG63 cell proliferation, migration, and invasion was examined using MTT and Transwell assays. Epithelial-mesenchymal transition (EMT) markers (E-cadherin and N-cadherin) were analyzed by RT-qPCR. The expression of genes involved in cell proliferation, migration, invasion, and the TGF-ß signaling pathway was evaluated by western blotting and RT-qPCR. RESULTS: INHBA levels were elevated in the OS tissues and cells. Furthermore, the transforming growth factor-ß (TGF-ß) signaling pathway of OS cells was suppressed in response to INHBA-siRNA, whereas proliferation, migration, and invasion of OS cells were inhibited. Besides, INHBA-siRNA significantly inhibited OS cell EMT, evidenced by enhanced E-cadherin mRNA expression and reduced N-cadherin mRNA expression. Further mechanistic studies revealed that the TGF-ß1 agonist SRI-011381 hydrochloride increased OS cell proliferation, migration, and invasion after INHBA downregulation. CONCLUSION: We found that INHBA silencing could play a vital role in OS via TGF-ß1-regulated proliferation, migration, and invasion.

Exogenous abscisic acid prolongs the dormancy of recalcitrant seed of Panax notoginseng.

The seeds of Panax notoginseng (Burk.) F. H. Chen are typically characterized by their recalcitrance and after-ripening process and exhibit a high water content at harvest as well as a high susceptibility to dehydration. Storage difficulty and the low germination of recalcitrant seeds of P. notoginseng are known to cause an obstacle to agricultural production. In this study, the ratio of embryo to endosperm (Em/En) in abscisic acid (ABA) treatments (1 mg·l-1 and 10 mg·l-1, LA and HA) was 53.64% and 52.34%, respectively, which were lower than those in control check (CK) (61.98%) at 30 days of the after-ripening process (DAR). A total of 83.67% of seeds germinated in the CK, 49% of seeds germinated in the LA treatment, and 37.33% of seeds germinated in the HA treatment at 60 DAR. The ABA, gibberellin (GA), and auxin (IAA) levels were increased in the HA treatment at 0 DAR, while the jasmonic acid (JA) levels were decreased. ABA, IAA, and JA were increased, but GA was decreased with HA treatment at 30 DAR. A total of 4,742, 16,531, and 890 differentially expressed genes (DEGs) were identified between the HA-treated and CK groups, respectively, along with obvious enrichment in the ABA-regulated plant hormone pathway and the mitogen-activated protein kinase (MAPK) signaling pathway. The expression of pyracbactin resistance-like (PYL) and SNF1-related protein kinase subfamily 2 (SnRK2s) increased in the ABA-treated groups, whereas the expression of type 2C protein phosphatase (PP2C) decreased, both of which are related to the ABA signaling pathway. As a result of the changes in expression of these genes, increased ABA signaling and suppressed GA signaling could inhibit the growth of the embryo and the expansion of developmental space. Furthermore, our results demonstrated that MAPK signaling cascades might be involved in the amplification of hormone signaling. Meanwhile, our study uncovered that the exogenous hormone ABA could inhibit embryonic development, promote dormancy, and delay germination in recalcitrant seeds. These findings reveal the critical role of ABA in regulating the dormancy of recalcitrant seeds, and thereby provide a new insight into recalcitrant seeds in agricultural production and storage.

[Effect of acupuncture combined with infantile tuina on intestinal flora in children with tic disorders].

OBJECTIVE: To observe the effect of acupuncture combined with infantile tuina on intestinal flora and its efficacy in children with tic disorders (TD), and to explore its mechanism. METHODS: A total of 15 children with TD were recruited as an observation group and 10 healthy children as a healthy control group. Regulating spleen and stomach acupuncture combined with infantile tuina were received in the observation group. First, acupuncture was applied to Zhongwan (CV 12), Tianshu (ST 25), Guanyuan (CV 4), Hegu (LI 4), Zusanli (ST 36), etc., and then abdominal massage and other tuina techniques were applied, once a day, 6 times a week, 2 weeks as a course of treatment, a total of 2 courses of treatment were required. No intervention was given in the healthy control group. In the observation group, Yale global tic severity scale (YGTSS) score and TCM syndrome score were compared before treatment and after 1 and 2 courses of treatment. 16S rRNA sequencing technology was used to detect the intestinal flora in the healthy control group and before and after treatment in the observation group. RESULTS: After 1 and 2 courses of treatment, the scores of YGTSS and TCM syndrome in the observation group were lower than those before treatment (P<0.01, P<0.05). Compared with the healthy control group, the number of operational taxonomic units (OTU) and indexes of Chao1, Sobs, Ace and Shannon were decreased in the observation group before treatment (P<0.05, P<0.01). Compared with before treatment, the number of OTU and indexes of Chao1, Sobs, Ace and Shannon were increased in the observation group after treatment (P<0.01, P<0.05). Compared with the healthy control group, the relative abundance of Firmicutes in the observation group before treatment was decreased (P<0.001), while the relative abundance of Bacteroidetes, Bacteroides and Erysipelatoclostridium was increased (P<0.001, P<0.05). Compared with before treatment, the relative abundance of Bacteroidetes in the observation group was decreased (P<0.001) after treatment, while the relative abundance of Actinobacteria, Bifidobacterium and Atopobium was increased (P<0.05, P<0.01). CONCLUSION: Acupuncture combined with infantile tuina based on the principle of regulating spleen and stomach could effectively improve TD symptoms in children, which may be related to regulating the diversity of intestinal flora, increasing beneficial bacteria, maintaining intestinal microecological balance, and playing a role in improving neurological disorders.

Artemether Alleviates Diabetic Kidney Disease by Modulating Amino Acid Metabolism.

Diabetes is a worldwide metabolic disease with rapid growing incidence, characterized by hyperglycemia. Diabetic kidney disease (DKD), the leading cause of chronic kidney disease (CKD), has a high morbidity according to the constantly increasing diabetic patients and always develops irreversible deterioration of renal function. Though different in pathogenesis, clinical manifestations, and therapies, both type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM) can evolve into DKD. Since amino acids are both biomarkers and causal agents, rarely report has been made about its metabolism which lies in T1DM- and T2DM-related kidney disease. This study was designed to investigate artemether in adjusting renal amino acid metabolism in T1DM and T2DM mice. Artemether was applied as treatment in streptozotocin (STZ) induced T1DM mice and db/db T2DM mice, respectively. Artemether-treated mice showed lower FBG and HbA1c and reduced urinary albumin excretion, as well as urinary NAG. Both types of diabetic mice showed enlarged kidneys, as confirmed by increased kidney weight and the ratio of kidney weight to body weight. Artemether normalized kidney size and thus attenuated renal hypertrophy. Kidney tissue UPLC-MS analysis showed that branched-chain amino acids (BCAAs) and citrulline were upregulated in diabetic mice without treatment and downregulated after being treated with artemether. Expressions of glutamine, glutamic acid, aspartic acid, ornithine, glycine, histidine, phenylalanine and threonine were decreased in both types of diabetic mice whereas they increased after artemether treatment. The study demonstrates the initial evidence that artemether exerted renal protection in DKD by modulating amino acid metabolism.

iTRAQ and RNA-seq analyses provide an insight into mechanisms of recalcitrance in a medicinal plant Panax notoginseng seeds during the after-ripening process.

Panax notoginseng (Burk) F.H. Chen is an important economic and medicinal plant from the family of Araliaceae, and its seed is characterised by the recalcitrance and after-ripening process. However, the molecular mechanism on the dehydration sensitivity is not clear in recalcitrant seeds. In the present study, isobaric tag for relative and absolute quantification (iTRAQ) and RNA-seq were used to analyse the proteomic and transcriptomic changes in seeds of P. notoginseng in days after-ripening (DAR). A total of 454 differentially expressed proteins (DEPs) and 12000 differentially expressed genes (DEGs) were obtained. The activity of enzymes related to antioxidant system were significantly increased, and the late embryogenesis abundant (LEA) protein family and most members of glutathione metabolism enzymes have been downregulated during the after-ripening process. The lack or inadequate accumulation of LEA proteins in the embryo and the low activity of antioxidant defense in glutathione metabolism might be the key factors leading to the dehydration sensitivity in recalcitrant seeds of P. notoginseng. In addition, the increased activity of elycolysis (EMP), citric acid cycle (TCA) and pentose phosphate pathway (PPP) pathways might be one of important signals to complete the after-ripening process. Overall, our study might provide a new insight into the molecular mechanism on dehydration sensitivity of recalcitrant seeds.

m6A methyltransferase METTL3 promotes oral squamous cell carcinoma progression through enhancement of IGF2BP2-mediated SLC7A11 mRNA stability.

As the key enzyme of the N6-methyladenosine (m6A) in eukaryotic messenger RNA, METTL3 plays an important role in tumor progression, but the exact mechanism by which METTL3 controls oral squamous cell carcinoma (OSCC) progression remains unclear. In this study, METTL3 expression in OSCC samples was analyzed by qPCR and immunohistochemistry. The effects of METTL3 suppression on OSCC cell lines were measured by CCK-8, Ki67 flow cytometry analysis, invasion transwell and wound healing assays. MeRIP-seq and RNA-seq analyses were performed to explore target gene of METTL3. RIP-qPCR and RNA stability assays were performed to explore the mechanism by which METTL3 regulated the target genes. Triptolide was used to evaluate its specific treatment effects on METTL3 in OSCC cells. BALB/c nude mice were used to establish orthotopic and subcutaneous xenograft models to verify the in vitro results. The results showed that METTL3 was upregulated in OSCC tissues compared with OSCC adjacent normal tissues, and its expression was associated with T stage, lymphatic metastasis and prognosis. METTL3 suppression impaired OSCC cells proliferation, invasion, and migration. MeRIP-seq and RNA-seq analysis identified that SLC7A11 mRNA was the m6A target of METTL3, which was verified by meRIP-qPCR, qPCR and western blot. METTL3 depletion decreased the stability of SLC7A11 mRNA, and IGF2BP2 as m6A reader was involved in this process. Moreover, METTL3 knockdown attenuated the binding between SLC7A11 mRNA and IGF2BP2, finally leading to accelerate SLC7A11 mRNA degradation. Triptolide inhibited METTL3-mediated SLC7A11 expression, thus suppressing malignancy of OSCC cells. In conclusion, the new finding of the manuscript is that METTL3 enhances the mRNA stability of SLC7A11 via m6A-mediated binding of IGF2BP2, which thus promotes OSCC progression, and triptolide inhibits OSCC by suppressing METTL3-SLC7A11 axis. Triptolide has a potential to be as an effective anti-OSCC drug targeted to METTL3.