[Analysis of SOX2 gene promoter activity in porcine early embryonic development].

SOX2 (sex determining region Y-box2) is one of the critical pluripotent factors that play a crucial role in the first lineage differentiation and maintenance of pluripotency in inner cell mass during early embryonic development. However, there are few researches about the regulation of the SOX2 promoter, especially in Sus scrofa. To analyzed the activity of SOX2 promoter in early porcine embryos, we determined the control system and established the microinjection system for assessing SOX2 promoter activity by analyzing the embryonic development and the expression of enhanced green fluorescence protein (EGFP) after micro-injected different EGFP plasmids at different times after activation of the oocytes. Then, we analyzed the structure of 5000 bp upstream of the SOX2 translation initiation site and found there were four transcription factor binding site clusters. Next, we designed and constructed promoter-containing plasmids to analyze the function of each cluster. To detect the activity of different promoters, we assessed the mCherry expression in protein levels and mRNA levels by analyzing the mCherry fluorescence intensity and qRT-PCR after injecting plasmids into embryos. These results showed that the activity of the shorted promoter, with the region from 2254 bp to 2442 bp upstream of translation initiation site deleted, decreased to 17.8% at 4-cell and 8-cell stages compared with the full-length promoter. This region included two NF-AT transcription factor binding sites, which indicated that the NF-AT binding site is a key region to regulate the activity of the SOX2 promoter. The results provide important data for determination the mechanism of porcine SOX2 regulation.

A bioactive new protostane-type triterpenoid from Alisma plantago-aquatica subsp. orientale (Sam.) Sam.

A new protostane-type triterpenoid, 5ß,29-dihydroxy alisol A (1) was isolated from Alisma plantago-aquatica subsp. orientale (Sam.) Sam. as well as 12-deoxyphorbol-13α-pentadecanoate (2). We first report the presence of compound 2 in the genus Alisma. Their structures were established on the basis of 1D and 2D NMR, and HRESIMS spectroscopic analyses. All the isolated compounds were assayed for their inhibitory effects against human carboxylesterase 2 (HCE-2). Compounds 1 and 2 displayed inhibitory activities against HCE-2 with IC50 values of 29.2 and 4.6 µM, respectively. The interaction mechanisms of HCE-2 with compounds 1 and 2 were investigated by molecular docking, respectively.

An indole diterpenoid isolated from the fungus Drechmeria sp. and its antimicrobial activity.

One new indole diterpenoid, drechmerin I (1), was isolated from the fermentation broth of Drechmeria sp. isolated from the root of Panax notoginseng. Its structure was elucidated based on 1 D and 2 D nuclear magnetic resonance (NMR), high resolution electrospray ionization mass spectrum (HRESIMS), and electronic circular dichroism (ECD) spectroscopic analyses as well as TD DFT calculations of ECD spectra. Drechmerin I (1) was assayed for its antimicrobial activity against Candida albicans, Staphylococcus aureus, Bacillus cereus, B. subtillis, Pseudomonas aeruginosa, and Klebsiella pneumonia, respectively. Drechmerin I (1) showed antimicrobial activities against B. subtillis with an MIC value of 200 µg/mL. The interaction of S. aureus peptide deformylase with drechmerin I (1) was investigated by molecular docking.

Xylarianins A-D from the endophytic fungus Xylaria sp. SYPF 8246 as natural inhibitors of human carboxylesterase 2.

Eighteen secondary metabolites were isolated from the fermentation broth of the endophytic fungus Xylaria sp. SYPF 8246, including four new compounds, xylarianins A-D (1-4), three new natural products, 6-methoxycarbonyl-2'-methyl-3,5,4',6'-tetramethoxy-diphenyl ether (5), 2-chlor-6-methoxycarbonyl-2'-rnethyl-3,5,4',6'-tetramethoxy-diphenyl ether (6), and 2-chlor-4'-hydroxy-6-methoxy carbonyl-2'-methyl-3,5,6'-trimethoxy-diphenyl ether (7), and eleven known compounds (8-18). Their structural elucidations were conducted by using 1D and 2D NMR, HRESIMS, and Rh2(OCOCF3)4-induced electronic circular dichroism (ECD) spectra analyses. The integrated 1H and 13C NMR data of three new natural products 5-7 were reported for the first time. All the isolated compounds were assayed for their inhibitory activities against human carboxylesterase 2 (hCE 2). Compounds 1, 5-9, and 18 displayed significant inhibitory activities against hCE 2 with IC50 values of 10.43 ±â€¯0.51, 6.69 ±â€¯0.85, 12.36 ±â€¯1.27, 18.25 ±â€¯1.78, 29.78 ±â€¯0.48, 18.86 ±â€¯1.87, and 20.72 ±â€¯1.51 µM, respectively. The interactions between compounds 1 and 5 with hCE 2 were anaylzed by molecular docking.

Uncaria rhynchophylla Ameliorates Parkinson's Disease by Inhibiting HSP90 Expression: Insights from Quantitative Proteomics.

BACKGROUND/AIMS: Uncaria rhynchophylla, known as "Gou-teng", is a traditional Chinese medicine (TCM) used to extinguish wind, clear heat, arrest convulsions, and pacify the liver. Although U. rhynchophylla has a long history of being often used to treat central nervous system (CNS) diseases, its efficacy and potential mechanism are still uncertain. This study investigated neuroprotective effect and the underlying mechanism of U. rhynchophylla extract (URE) in MPP+-induced SH-SY5Y cells and MPTP-induced mice. METHODS: MPP+-induced SH-SY5Y cells and MPTP-induced mice were used to established Parkinson's disease (PD) models. Quantitative proteomics and bioinformatics were used to uncover proteomics changes of URE. Western blotting was used to validate main differentially expressed proteins and test HSP90 client proteins (apoptosis-related, autophagy-related, MAPKs, PI3K, and AKT proteins). Flow cytometry and JC-1 staining assay were further used to confirm the effect of URE on MPP+-induced apoptosis in SH-SY5Y cells. Gait analysis was used to detect the behavioral changes in MPTP-induced mice. The levels of dopamine (DA) and their metabolites were examined in striatum (STR) by HPLC-EC. The positive expression of tyrosine hydroxylase (TH) was detected by immunohischemical staining and Western blotting. RESULTS: URE dose-dependently increased the cell viability in MPP+-induced SH-SY5Y cells. Quantitative proteomics and bioinformatics results confirmed that HSP90 was an important differentially expressed protein of URE. URE inhibited the expression of HSP90, which further reversed MPP+-induced cell apoptosis and autophagy by increasing the expressions of Bcl-2, Cyclin D1, p-ERK, p-PI3K p85, PI3K p110α, p-AKT, and LC3-I and decreasing cleaved caspase 3, Bax, p-JNK, p-p38, and LC3-II. URE also markedly decreased the apoptotic ratio and elevated mitochondrial transmembrane potential (DΨm). Furthermore, URE treatment ameliorated behavioral impairments, increased the contents of DA and its metabolites and elevated the positive expressions of TH in SN and STR as well as the TH protein. CONCLUSIONS: URE possessed the neuroprotective effect in vivo and in vitro, regulated MAPK and PI3K-AKT signal pathways, and inhibited the expression of HSP90. U. rhynchophylla has potentials as therapeutic agent in PD treatment.

Drechmerin H, a novel 1(2), 2(18)-diseco indole diterpenoid from the fungus Drechmeria sp. as a natural agonist of human pregnane X receptor.

A novel 1(2), 2(18)-diseco indole diterpenoid, drechmerin H (1), was isolated from the fermentation broth of Drechmeria sp. together with a new indole diterpenoid, 2'-epi terpendole A (3), and a known analogue, terpendole A (2). Their structures were determined by HRESIMS, 1D and 2D NMR, ECD, and X-ray single crystal diffraction analyses as well as quantum chemical calculation. The abosulte configuration of terpendole A (2) was determined for the first time. Compound 1 displayed the significant agonistic effect on pregnane X receptor (PXR) with EC50 value of 134.91 ±â€¯2.01 nM, and its interaction with PXR was investigated by molecular docking. Meantime, a plausible biosynthetic pathway for compounds 1-3 is also discussed in the present work.

Flavobacterium artemisiae sp. nov., isolated from the rhizosphere of Artemisia annua L. and emended descriptions of Flavobacterium compostarboris and Flavobacterium procerum.

A Gram-stain-negative, strictly aerobic, yellow-coloured, motile by gliding and elongated rod-shaped bacterial strain, designated SYP-B1015T, was isolated from the rhizosphere of Artemisia annua L. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain SYP-B1015T belonged to the genus Flavobacterium and had highest 16S rRNA gene sequence similarity to Flavobacterium compostarboris JCM 16527T (98.1 %) and Flavobacterium procerum JCM 30113T (97.2 %). The predominant respiratory quinone for the strain was MK-6, and the major cellular fatty acids were iso-C15 : 0, iso-C15 : 0 3-OH and iso-C17 : 0 3-OH. The polar lipid profile contained phosphatidylethanolamine as a major compound. The DNA G+C content of strain SYP-B1015T was 33.5 mol%. The DNA-DNA relatedness values between strain SYP-B1015T and F. compostarboris JCM 16527T and F. procerum JCM 30113T were 56.5±0.4 and 48.9±1.2 %, respectively. Combining the data from morphological, physiological, biochemical and chemotaxonomic characterizations presented in this study, strain SYP-B1015T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium artemisiae sp. nov. is proposed. The type strain is SYP-B1015T (=CGMCC 1.16115T=KCTC 62025T).

Indole diterpenoids from the endophytic fungus Drechmeria sp. as natural antimicrobial agents.

A fungal strain, Drechmeria sp., was isolated from the root of Panax notoginseng. Totally, seven new indole diterpenoids, drechmerins A-G (1-7), were isolated from the fermentation broth of Drechmeria sp. together with four known analogues (8-11). Their structures were determined on the basis of 1D and 2D NMR and electronic circular dichroism (ECD) spectroscopic analyses as well as theoretical calculations. All the isolated compounds were evaluated for their antimicrobial activities against Candida albicans, Staphylococcus aureus, Bacillus cereus, B. subtillis, Pseudomonas aeruginosa, and Klebsiella pneumonia, respectively. Drechmerin B (2) displayed antimicrobial activity against C. albicans with an MIC value of 12.5 µg/mL. Molecular docking was used to investigate interactions of peptide deformylase with compounds 1-3, 5-7, 9, and 10.

Chemical constituents from Alisma plantago-aquatica subsp. orientale (Sam.) Sam and their anti-inflammatory and antioxidant activities.

The chemical constituent investigation of Alisma plantago-aquatica subsp. orientale has led to isolation and identification of thirteen compounds, including one new sesquiterpene, (10S)-11-hydroxy-ß-cyperone (1), three sesquiterpenes (2-4), five phenylpropanoids (5-9), and four alkaloids (10-13). We report herein, for the first time, the presence of compounds 2-13 in the genus Alisma. Their structures were established using 1D and 2D NMR and HRESIMS spectroscopic analyses. All the isolated compounds were assayed for their inhibitory activities against nitric oxide production in LPS-induced RAW 264.7 cells and antioxidant activities by DPPH scavenging assay. Compounds 1-13 displayed significant inhibitory effects against NO production at a certain concentration, while compound 5 showed antioxidant activity with IC50 of 55.28 µM. The interactions of compounds 1, 5, and 11 with iNOS were investigated using molecular docking.

Arthrobacter ginkgonis sp. nov., an actinomycete isolated from rhizosphere of Ginkgo biloba L.

A Gram-stain-positive, aerobic actinobacterial strain (designated SYP-A7299T), which displayed a rod-coccus growth lifecycle, was isolated from the rhizosphere of Ginkgo biloba L. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain SYP-A7299T belongs to the genus Arthrobacter and is most closely related to Arthrobacter halodurans JSM 078085T (97.4 % 16S rRNA gene sequence similarity). The DNA-DNA relatedness value between strain SYP-A7299T and A. halodurans JSM 078085T was 37 % ±2.9. The cell-wall peptidoglycan was A4α, and glucose and galactose were whole-cell sugars. The polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, two glycolipids and an unknown polar lipid. The major menaquinone were MK-8(H2) (72 %) and MK-9(H2) (28 %), and the predominant cellular fatty acids were anteiso-C15 : 0, iso-C15 : 0 and anteiso-C17 : 0. The DNA G+C content was 68.9 mol%. Based on the morphological, physiological, biochemical and chemotaxonomic characters presented in this study, strain SYP-A7299T represents a novel species of the genus Arthrobacter, for which the name Arthrobacter ginkgonis sp. nov. is proposed. The type strain is SYP-A7299T (=DSM 100491T=KCTC 39 592T).

[Inactivation Efficiency and Mechanism of Three Dominant Fungal Spores in Drinking Groundwater by Chlorine].

The outbreak of fungi in drinking groundwater sources can produce odor, cause toxicity and form lots of visible flocs, which seriously affect the drinking water quality. In this study, the inactivation efficiency of three dominant fungal spores Trichoderma, Penicillium, Cladosporium by chlorine was conducted, and the inactivation mechanism was explored by monitoring the hydrophobicity, the leakage of intracellular substances, the increase of extracellular adenosine triphosphate (ATP), deoxyribonucleic acid (DNA) and proteins, and the change of spores' morphology. The results showed that the inactivation of fungal spores was consistent with first-order kinetics and satisfied the Chick model. The resistance to chlorine was in sequence of Trichoderma > Penicillium > Cladosporium, the larger size the fungal spores and the more hydrophilicity the fungal spores, the higher the inactivation efficiency. Chlorination resulted in the remarkable leakage of intracellular compounds, the increase of extracellular characteristic compounds (ATP, DNA and protein), the damage of the cell surface, and the fungal spores recessed and wrinkled. In summary, chlorine firstly reacted with spores' surface and reduced their cultivability, and then resulted in the damage to the permeability barrier of the spores and the release of intracellular characteristic compounds, and finally the viability of spores was damaged.

In silico classification of human maximum recommended daily dose based on modified random forest and substructure fingerprint.

A modified random forest (RF) algorithm, as a novel machine learning technique, was developed to estimate the maximum recommended daily dose (MRDD) of a large and diverse pharmaceutical dataset for phase I human trials using substructure fingerprint descriptors calculated from simple molecular structure alone. This type of novel molecular descriptors encodes molecular structure in a series of binary bits that represent the presence or absence of particular substructures in the molecule and thereby can accurately and directly depict a series of local information hidden in this molecule. Two model validation approaches, 5-fold cross-validation and an independent validation set, were used for assessing the prediction capability of our models. The results obtained in this study indicate that the modified RF gave prediction accuracy of 80.45%, sensitivity of 75.08%, specificity of 84.85% for 5-fold cross-validation, and prediction accuracy of 80.5%, sensitivity of 76.47%, specificity of 83.48% for independent validation set, respectively, which are as a whole better than those by the original RF. At the same time, the important substructure fingerprints, recognized by the RF technique, gave some insights into the structure features related to toxicity of pharmaceuticals. This could help provide intuitive understanding for medicinal chemists.