RNA editing events and expression profiles of mitochondrial protein-coding genes in the endemic and endangered medicinal plant, Corydalis saxicola.

Corydalis saxicola, an endangered medicinal plant endemic to karst habitats, is widely used in Traditional Chinese Medicine to treat hepatitis, abdominal pain, bleeding hemorrhoids and other conditions. However, to date, the mitochondrial (mt) genome of C. saxicola has not been reported, which limits our understanding of the genetic and biological mechanisms of C. saxicola. Here, the mt genome of C. saxicola was assembled by combining the Nanopore and Illumina reads. The mt genome of C. saxicola is represented by a circular chromosome which is 587,939 bp in length, with an overall GC content of 46.50%. 40 unique protein-coding genes (PCGs), 22 tRNA genes and three rRNA genes were identified. Codon usage of the PCGs was investigated and 167 simple sequence repeats were identified. Twelve homologous fragments were identified between the mt and ct genomes of C. saxicola, accounting for 1.04% of the entire mt genome. Phylogenetic examination of the mt genomes of C. saxicola and 30 other taxa provided an understanding of their evolutionary relationships. We also predicted 779 RNA editing sites in 40 C. saxicola mt PCGs and successfully validated 506 (65%) of these using PCR amplification and Sanger sequencing. In addition, we transcriptionally profiled 24 core mt PCGs in C. saxicola roots treated with different concentrations of CaCl2, as well as in other organs. These investigations will be useful for effective utilization and molecular breeding, and will also provide a reference for further studies of the genus Corydalis.

Protective effect of ethyl ferulate against hypoxic injury in retinal cells and retinal neovascularization in an oxygen-induced retinopathy model.

BACKGROUND: Pathological neovascularization is a major cause of visual impairment in hypoxia-induced retinopathy. Ethyl ferulate (EF), the natural ester derivative of ferulic acid commonly found in Ferula and Angelica Sinensis, has been shown to exert antioxidant, neuroprotective, and anti-inflammatory properties. However, whether EF exerts a protective effect on retinal neovascularization and the underlying mechanisms are not well known. PURPOSE: The aim of the study was to investigate the effect of EF on retinal neovascularization and explore its underlying molecular mechanisms. STUDY-DESIGN/METHODS: We constructed hypoxia models induced by cobalt chloride (CoCl2) in ARPE-19 cells and Rhesus choroid-retinal vascular endothelial (RF/6A) cells in vitro, as well as a retinal neovascularization model in oxygen-induced retinopathy (OIR) mice in vivo. RESULTS: In this work, we demonstrated that EF treatment inhibited hypoxia-induced vascular endothelial growth factor A (VEGFA) expression in ARPE-19 cells and abrogated hypoxia-induced tube formation in RF/6A cells. As expected, intravitreal injection of EF significantly suppressed retinal neovascularization in a dose-dependent manner in OIR retinas. We also found that hypoxia increased VEGFA expression by blocking autophagic flux, whereas EF treatment enhanced autophagic flux, thereby reducing VEGFA expression. Furthermore, EF activated the sequestosome 1 (p62) / nuclear factor E2-related factor 2 (Nrf-2) pathway via upregulating oxidative stress-induced growth inhibitor 1 (OSGIN1) expression, thus alleviating oxidative stress and reducing VEGFA expression. CONCLUSION: As a result of our findings, EF has an inhibitory effect on retinal neovascularization, implying a potential therapeutic strategy for hypoxia-induced retinopathy.

Transanal Minimally Invasive Surgery for Rectal Anastomotic Stenosis After Colorectal Cancer Surgery.

BACKGROUND: Anastomotic stenosis is a common complication of colorectal cancer surgery with anastomosis. Transanal minimally invasive surgery is a novel approach to the treatment of anastomotic stenosis. OBJECTIVE: This study aimed to evaluate the efficacy and safety of transanal minimally invasive surgery for anastomotic stenosis treatment. DESIGN: This was a retrospective study. SETTINGS: This study was conducted at a comprehensive cancer center. PATIENTS: This study included patients with rectal anastomotic stenosis who after undergoing colorectal surgery were admitted to the Sir Run Run Shaw Hospital between September 2017 and June 2019. MAIN OUTCOME MEASURES: The primary outcome was the operative success rate. The secondary outcomes were intraoperative variables, postoperative complications, stoma closure conditions, and stenosis recurrence risks. RESULTS: Nine patients, aged 52 to 80 years, with a history of colorectal cancer with end-to-end anastomosis underwent transanal minimally invasive surgery for anastomotic stenosis. The distance between the stenosis and the anal verge ranged from 5 to 12 cm. The mean stenosis diameter was 0.3 cm. Four patients had completely obstructed rectal lumens. Eight of 9 patients successfully underwent transanal minimally invasive surgery radial incision and cutting. The average operation time was 50 minutes. After the procedure, 1 patient had symptomatic procedure-associated perforations but recovered with conservative treatment. No perioperative mortality occurred. One patient underwent transverse colostomy 1 month after transanal minimally invasive surgery because of proximal colon ischemia induced by primary rectal surgery. Eight patients underwent protective loop ileostomy. After transanal minimally invasive surgery, stoma closure was performed in 88% of patients with no stenosis recurrence or obstruction at follow-up (21-42 mo). LIMITATIONS: This study was limited by its small sample size and single-center design. CONCLUSIONS: Transanal minimally invasive surgery provides an excellent operative field, good maneuverability, and versatile instrumentation and is a safe and effective treatment for rectal anastomotic stenosis, especially for severe fibrotic stenosis or complete obstruction. See Dynamic Article Video at http://links.lww.com/DCR/B965 .

Research progress on natural cannabinoid receptor type 2( CB2R)regulators and their effects on bone metabolism / 中国中药杂志

Cannabinoid receptor type 2( CB2 R),a member of the G protein-coupled receptor( GPCR) superfamily,has a variety of biological activities,such as regulating pain response,resisting inflammation and fibrosis,and mediating bone metabolism. Some CB2 R regulators exhibit a good regulatory effect on bone metabolism. Cannabinoids in Cannabis sativa can cause psychoactive effects despite various pharmacological actions they exerted by targeting CB2 R. Therefore,it is of great significance to discover CB2 R regulators in non-Cannabis plants for finding new lead compounds without psychoactive effects and elucidating the action mechanism of plant drugs. The present study clarifies the discovery,structure,and physiological functions of CB2 R,especially its regulatory effects on bone metabolism,summarized CB2 R regulators extracted from non-Cannabis plants,and systematically analyzes the regulatory effects of CB2 R regulators on bone metabolism in animals,osteoblasts,and osteoclasts,to provide a scientific basis for the discovery of new CB2 R regulators and the development of anti-osteoporotic drugs.

Analysis on Mechanism of Astragali Radix in Treating IgA Nephropathy Based on Network Pharmacology and in Vitro Cell Experiment / 中国实验方剂学杂志

Objective:To explore the multi-component， multi-target and multi-pathway mechanism of Astragali Radix against immunoglobulin A nephropathy （IgAN） by network pharmacology， aiming to provide evidence for its basic research and clinical application. Method:The active chemical components and targets of Astragali Radix and targets associated with IgAN were obtained by literature mining and GeneCards， Traditinal Chinese Medicine Integrated Database （TCMID）， Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP） databases. Cytoscape 3.7.1 software was used to draw network interaction diagrams. The key targets of Astragali Radix against IgAN were searched by network topology. Gene ontology （GO） analysis and Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway analysis involved in the targets were analyzed by different packages in R programming language. On this basis， cell experiments <italic>in vitro</italic> were carried out to verify the activation effect of astragaloside Ⅳ on phosphatidylinositol 3-kinase/protein kinase B/tumor suppressor gene protein 53 （PI3K/Akt/p53） signaling pathway of human mesangial cells. Result:A total of 25 active components and 49 ingredient-disease targets of Astragali Radix were screened. The GO enrichment analysis included 84 items， which were related to nuclear hormone receptor binding， nuclear receptor activity， deoxyribonucleic acid binding transcriptional activation activity and other aspects. The KEGG pathway enrichment analysis included 88 KEGG pathways， which were closely related to PI3K/Akt signaling pathway， hypoxia inducible factor-1 （HIF-1） signaling pathway， advanced glycation end product/receptor of advanced glycation end product （AGE/RAGE） signaling pathway and others. Cell experiments <italic>in vitro </italic>confirmed that astragaloside Ⅳ could effectively inhibit the platelet derived growth factor-BB （PDGF-BB）-induced proliferation of human mesangial cells by regulating PI3K/Akt/p53 signaling pathway. Conclusion:The active ingredients of Astragali Radix may play a role in the treatment of IgAN by acting on targets and pathways related to apoptosis， oxidative stress， inflammation response and others， providing ideas and directions for the new drug development and mechanism study of IgAN.

Impact of Vitamin E supplementation on vascular function in haptoglobin genotype stratified diabetes patients (EVAS Trial): a randomised controlled trial.

AIMS: Vitamin E (Vit-E) may preferentially improve cardiovascular risk in haptoglobin 2-2 (Hp2-2) genotype diabetes individuals. We studied the impact of Vit-E supplementation on vascular function in diabetes individuals stratified by haptoglobin genotype in Singapore. METHODS: In this 24-week, double blind, placebo-controlled RCT, we recruited 187 subjects (101 Hp2-2, 86 non-Hp2-2). INTERVENTION: alpha-tocopherol-400 IU. PRIMARY OUTCOME: Change in EndoPAT-derived reactive-hyperaemia index (RHI) and augmentation index (AIx); Secondary Outcomes: Pulse-Wave velocity (Sphygmocor-PWV), carotid intima media thickness (CIMT), inflammation (hsCRP), derivatives of reactive-oxygen metabolites (dROMs), biological antioxidant-potential (BAPs), HbA1c, LDL-C, HDL-C and oxidised LDL-C (ox-LDL). RESULTS: Overall, with Vit-E supplementation no significant change in RHI, PWV, CIMT, hsCRP, dROMS, BAPs, HDL-C and HbA1c was observed (p > 0.05); an increase in LDL-C with concomitant decrease in ox-LDL, and incidentally increase in eGFR was observed (p < 0.05). No interaction effect with haptoglobin genotype was seen for all outcomes (p > 0.05). Subgroup analysis: In the non-Hp-2-2 group, Vit-E supplementation led to a higher EndoPAT-derived AIx, accompanied by higher LDL and ox-LDL concentrations (p < 0.05); Hp2-2 group: Vit-E supplementation led to higher eGFR when compared to the non-Hp2-2 group (exploratory) (p < 0.05). We observed an interaction effect for baseline haptoglobin concentration (threshold > 119 mg/dl) with intervention in terms of increased EndoPAT-derived AIx in the Hp > 119 mg/dl group whereas no change in the group with Hp ≤ 119 mg/dl. CONCLUSION: Vit-E supplementation did not show any preferential benefit or deleterious effect on vascular function in Hp2-2 diabetes subjects in Singapore. A possible deleterious effect of an increase in arterial stiffness in individuals with Hp > 119 mg/dl was observed. Future studies should consider personalisation based on baseline Hp concentrations in patients with T2DM rather than just Hp2-2 genotype to evaluate impact on the detailed lipid pathways, cardiac and renal physiology. The impact of ethnic differences needs to be explored in greater details.

A fluorescence resonance energy transfer (FRET) based "Turn-On" nanofluorescence sensor using a nitrogen-doped carbon dot-hexagonal cobalt oxyhydroxide nanosheet architecture and application to α-glucosidase inhibitor screening.

The medicines targeted at α-glucosidase played an important role in anti-diabetes and anti-HIV therapy. Unfortunately, the method based on fluorescent assay strategy for α-glucosidase inhibitor screening remains poorly investigated. In this study, a novel "Turn On" fluorescence sensor platform has been developed for trace α-glucosidase inhibitor screening from natural medicines. Firstly, carbon dots were prepared by one-pot synthesis and used as the signal output. Combining with the carbon dots, cobalt oxyhydroxide (CoOOH) nanoflakes were employed to build the fluorescence resonance energy transfer (FRET) based sensor platform. Secondly, L-ascorbic acid-2-O-α-D-glucopyranosyl (AAG) was innovatively introduced as α-glucosidase substrate. With hydrolysis of AAG by α-glucosidase, ascorbic acids (AA) were released that can rapidly reduce CoOOH nanoflakes to Co(2+), and then FRET was stopped accompanying with the fluorescence recovery of CDs. The sensor platform was ultrasensitive to AA with a detection limit of 5 nM, ensuring the sensitive monitoring of enzyme activity. Acarbose was used as the inhibitor model and its inhibition rate is proportional to the logarithm of concentration in range of 10(-9)-10(-3)M with the correlation coefficient of R(2)=0.996, and an ultralow limit of detection of ~1×10(-9)M was obtained. The inhibiting ability of seven compounds isolated from natural medicines was also evaluated. The constructed sensor platform was proven to be sensitive and selective as well as cost-effective, facile and reliable, making it promising as a candidate for trace α-glucosidase inhibitor screening.

Accurate Analysis and Evaluation of Acidic Plant Growth Regulators in Transgenic and Nontransgenic Edible Oils with Facile Microwave-Assisted Extraction-Derivatization.

Determination of plant growth regulators (PGRs) in a signal transduction system (STS) is significant for transgenic food safety, but may be challenged by poor accuracy and analyte instability. In this work, a microwave-assisted extraction-derivatization (MAED) method is developed for six acidic PGRs in oil samples, allowing an efficient (<1.5 h) and facile (one step) pretreatment. Accuracies are greatly improved, particularly for gibberellin A3 (-2.72 to -0.65%) as compared with those reported (-22 to -2%). Excellent selectivity and quite low detection limits (0.37-1.36 ng mL(-1)) are enabled by fluorescence detection-mass spectrum monitoring. Results show the significant differences in acidic PGRs between transgenic and nontransgenic oils, particularly 1-naphthaleneacetic acid (1-NAA), implying the PGRs induced variations of components and genes. This study provides, for the first time, an accurate and efficient determination for labile PGRs involved in STS and a promising concept for objectively evaluating the safety of transgenic foods.

The Spectroscopy Study of the Binding of an Active Ingredient of Dioscorea Species with Bovine Serum Albumin with or without Co(2+) or Zn(2+).

Diosgenin (DIO) is the active ingredient of Dioscorea species. The interaction of DIO with bovine serum albumin (BSA) was investigated through spectroscopic methods under simulated physiological conditions. The fluorescence quenching data revealed that the binding of DIO to BSA without or with Co(2+) or Zn(2+) was a static quenching process. The presence of Co(2+) or Zn(2+) both increased the static quenching constants K SV and the binding affinity for the BSA-DIO system. In the sight of the competitive experiment and the negative values of ΔH (0) and ΔS (0), DIO bound to site I of BSA mainly through the hydrogen bond and Van der Waals' force. In addition, the conformational changes of BSA were studied by Raman spectra, which revealed that the secondary structure of BSA and microenvironment of the aromatic residues were changed by DIO. The Raman spectra analysis indicated that the changes of conformations, disulfide bridges, and the microenvironment of Tyr, Trp residues of BSA induced by DIO with Co(2+) or Zn(2+) were different from that without Co(2+) or Zn(2+).

A sensitive fluorescence reagent, 2-[2-(7H-dibenzo[a,g]carbazol-7-yl)-ethoxy]ethyl chloroformate, for amino acids determination in Saussurea involucrate and Artemisia capillaris Thunb using high-performance liquid chromatography with fluorescence detection and identification with mass spectroscopy/electrospray ionization source.

Recent researches shows that amino acids (AA) are not only cell signaling molecules but are also regulators of gene expression and the protein phosphorylation cascade. More precise analysis of AA composition is reckoned to be one of the most important applications in the biomedical and pharmaceutical fields. In this paper, we develop a sample, sensitive and mild method using 2-[2-(7H-dibenzo[a,g]carbazol-7-yl)-ethoxy]ethyl chloroformate (DBCEC) as A labeling reagent for AA determination by high-performance liquid chromatography (HPLC) with fluorescence detection (FLD) and identification with mass spectroscopy. The maximum excitation and emission wavelengths for DBCEC-AA derivatives were 300 and 395 nm, respectively. This method, in conjunction with a gradient elution, offered a baseline resolution of 20 AA on a reversed-phase Hypersil BDS C(18) column. LC separation for the derivatized AA showed good reproducibility, and all AA were found to give excellent linear responses with correlation coefficients > 0.9993. The calculated detection limits with a 25.0 fmol injection of each AA (at a signal-to-noise ratio of 3:1) ranged from 2.62 to 22.6 fmol. This method was applied to determine the AA composition in Saussurea involucrate and Artemisia capillaris Thunb. Meanwhile, this method exhibits a powerful potential for trace analysis of AA from biomedicine, foodstuff and other complex samples.

Supercritical CO(2) oil extraction from Chinese star anise seed and simultaneous compositional analysis using HPLC by fluorescence detection and online atmospheric CI-MS identification.

BACKGROUND: Supercritical CO(2) was utilised to extract Chinese star anise seed oil (CSASO), and a three-level Box-Behnken factorial design from response surface methodology was applied to optimise the extraction conditions, including pressure, temperature and amount of modifier (ethanol). The compositional analysis of fatty acids in CSASO was performed by HPLC with fluorescence detection using 2-(11H-benzo[a]carbazol-11-yl)-ethyl-4-methylbenzenesulfonate (BCETS) as labelling reagent. Identification was carried out by online atmospheric chemical ionisation-mass spectrometry. RESULTS: The optimum extraction conditions were as follows: extraction pressure, 27.72 MPa, extraction temperature, 46.22 degrees C, and amount of modifier, 8.58 vol.%. The experimental result showed that the maximum extraction yield was 25.31 +/- 0.22% (w/w) under the conditions proposed. The compositional analysis indicated that CSASO mainly contained C18:2, C18:1, C18:3, C20:4, C16, C18 and C20 fatty acids. CONCLUSION: In this study, a fast, simple and high-efficiency supercritical technique for extracting oil from Chinese star anise seed was developed. Simultaneous determination of fatty acids in CSASO using BCETS as the labelling reagent with HPLC fluorescence detection and online mass spectroscopy identification has been successfully achieved.

[Separation and purification of flavonoids from Smilax glabra by macroporous adsorption resin].

OBJECTIVE: To investigate the process of separating and purifying flavonoids from Smilax glabra. METHOD: With the yield of flavonoids as index, the optimum process of separating and purifying flavonoids from S. glabra Roxb was screened by static and dynamic adsorption tests. RESULT: The static saturated adsorption capacity of D101 macroporous resin to flavonoids of S. glabra was 45.6 mg x g(-1) (dry resin). The optimum conditions of dynamic adsorption and elution were as that the pH, the concentration, the adsorption velocity of the extracting solution, and the adsorption capacity were 6.00 +/- 0.20, 4.2 mg x mL(-1), 2 mL x min(-1) and 15 mL, respectively. The adsorbed resin column was washed by 100 mL 60% ethanol with pH value of 8.00 +/- 0.20 at the eluting velocity of 3 mL x min(-1) after washed by 100 mL distilled water. CONCLUSION: The flavonoids of S. glabra was able to be easily separated and purified by D101 macroporous resin under the optimum conditions above, and the recovery rate was higher than 90%. The content of obtained flavonoids reached 62.6%, which was 2 times of the content before purification.

Separation and purification of flavonoids from Smilax glabra by macroporous adsorption resin / 中国中药杂志

<p><b>OBJECTIVE</b>To investigate the process of separating and purifying flavonoids from Smilax glabra.</p><p><b>METHOD</b>With the yield of flavonoids as index, the optimum process of separating and purifying flavonoids from S. glabra Roxb was screened by static and dynamic adsorption tests.</p><p><b>RESULT</b>The static saturated adsorption capacity of D101 macroporous resin to flavonoids of S. glabra was 45.6 mg x g(-1) (dry resin). The optimum conditions of dynamic adsorption and elution were as that the pH, the concentration, the adsorption velocity of the extracting solution, and the adsorption capacity were 6.00 +/- 0.20, 4.2 mg x mL(-1), 2 mL x min(-1) and 15 mL, respectively. The adsorbed resin column was washed by 100 mL 60% ethanol with pH value of 8.00 +/- 0.20 at the eluting velocity of 3 mL x min(-1) after washed by 100 mL distilled water.</p><p><b>CONCLUSION</b>The flavonoids of S. glabra was able to be easily separated and purified by D101 macroporous resin under the optimum conditions above, and the recovery rate was higher than 90%. The content of obtained flavonoids reached 62.6%, which was 2 times of the content before purification.</p>