Secondary Metabolite Analysis of Phomopsis sp. LGT-5 Based on the Dual Guidance of Whole-Genome Sequencing and HPLC-Q-ToF-MS/MS.

Microorganisms produce a wealth of structurally diverse specialized metabolites with a remarkable range of biological activities. The Phomopsis sp. LGT-5 was obtained through tissue block and repeatedly crossed methods from Tripterygium wilfordii Hook. F. The antibacterial experiments of LGT-5 showed that it has high inhibitory activity against Staphylococcus aureus and Pseudomonas aeruginosa, and moderate inhibitory activity against Candida albicans. To research the generation of the antibacterial phenomenon of LGT-5 and provide support for further research and application, the whole genome sequencing (WGS) of LGT-5 was obtained by single-molecule real-time DNA sequencing platform Pacific Biosciences (PacBio) sequencing and Illumina paired-end sequencing. The final assembled LGT-5 genome is 54.79 Mb with a contig N50 of 290.07 kb; in addition, its secondary metabolites were detected through HPLC-Q-ToF-MS/MS. By comparing its MS/MS data, the secondary metabolites were analyzed based on visual network maps obtained on the Global Natural Products Social Molecular Networking (GNPS). The analysis results showed that the secondary metabolites of LGT-5 were triterpenes and various cyclic dipeptides.

Native Endophytes of Tripterygium wilfordii-Mediated Biotransformation Reduces Toxicity of Celastrol.

Celastrol (1), obtained from the roots of Tripterygium wilfordii Hook F., is most likely to become an antitumor drug, but with severe cytotoxicity. Due to the lack of modifiable sites in the structure of celastrol, the structural diversity of the modified products obtained by synthesis in the previous studies is insufficient, which hinders the pace of its patent medicine. This study describes a method of microbial transformation to increase the modification site of celastrol and reduce its toxicity. The screening of endophytes from native plants was introduced in this context, which led to two novel stereoselective oxidation products such as S-16-hydroxyl celastrol (2) and A-ring aromatized S-16-hydroxyl celastrol (3), along with a rare 7,9-octadecadienoic acid ester of celastrol (4). Their structures were determined by extensive spectroscopic data analysis, especially 1D and 2D NMR. Compared with 1, compounds 3 and 4 exhibited similar antitumor activity in U251, A549, KG-1, and B16 cell lines. Compound 2 had slightly decreased antitumor activity when compared with compound 1. Furthermore, compound 2-4 showed lower cytotoxicity against BV-2 (about 21-fold lower, 2: 92.82 µM, 3: 34.25 µM, and 4: 74.75 µM vs. celastrol: 4.35 µM), and also identical trends against H9c2 and PC12 cell lines.

Classification of diterpenoid alkaloids from Aconitum kusnezoffii Reichb. by liquid chromatography-tandem mass spectrometry-based on molecular networking.

Trace amounts of components in traditional Chinese medicine are considered pharmacological active substances used for treating many serious diseases. However, purifying all the trace substances and making clear their structures are not easy. In this context, high-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry based molecular networking was applied to investigate the chemical constituents of the roots of Aconitum kusnezoffii Reichb., which led to the identification of 33 nodes in different groups (N1-N33). Based on the excremental fragmentation pathway of known diterpenoid alkaloids (1-9) and comparisons of characteristic ions and characteristic loss of analogs in literature, the structures of unknown ions were deduced. This work lays a foundation for the evaluation of the clinical basis and mechanism of traditional Chinese medicine from the aspects of chemistry. In this paper, the method speculation of unknown natural products by means of molecular network method is expected to be applied in the discovery and change law of relevant active components in clinical pharmacology and the change of complex systems caused by trace active compounds.

[A new 9,19-cycloartane glycoside from Asplenium ruprechtii].

Chemical constituents of water extracts of Asplenium ruprechtii were investigated. Five compounds were isolated by silica gel, Sephadex LH-20 gel column chromatographies and preparative HPLC, and their structures were identified by various spectral analyses as aspleniumside G(1), trans-p-coumaric acid(2), trans-p-coumaric acid 4-O-ß-D-glucoside(3), cis-p-coumaric acid 4-O-ß-D-glucoside(4), and(E)-ferulic acid-4-O-ß-D-glucoside(5). Among them, compound 1 is a new 9,19-cycloartane glycoside.

Structural Determination of a New Cycloartane Glycoside from Asplenium ruprechtii.

We characterized a new cycloartane glycoside, herein known as aspleniumside F (1), along with five known compounds as kaempferol-3-O-[(6-O-(E)-feruloyl)-ß-D-glucopyranosyl]-(1→2)-ß-D-galacopyranoside (2), quercetin-3-O-[(6-O-(E)-feruloyl)-ß-D-glucopyranosyl]-(1→2)-ß-D-glucopyranoside (3), kaempferol-3-O-[(6-O-(E)-caffeoyl)-ß-D-glucopyranosyl]-(1→2)-ß-D-glucopyranoside (4), kaempferol-3-O-[(6-O-(E)-caffeoyl)-ß-D-glucopyranosyl]-(1→2)-ß-D-glucopyranosyl-7-O-ß-D-glucopyranoside (5), and kaempferol-3-O-[(6-O-p-coumaroyl)-ß-D-glucopyranosyl]-(1→2)-ß-D-glucopyranosyl-7-O-ß-D-glucopyranoside (6), from Asplenium ruprechtii Sa. Kurata, a folk medicine widely used to treat Thromboangiitis obliterans in China, Japan, and Korea. Based on spectroscopic, mainly 1D-, 2D-NMR and (+)-HR-ESI-MS, analyses as well as through comparisons with previous reports, its chemical structure was determined as 3ß,24,30-tri-ß-D-glucopyranosyl-23,25-dihydroxycycloartane (= (23R,24R)-3ß,24-bis-(ß-D-glucopyranosyloxy)-23,25-dihydroxy-9ß-9,19-cyclolanostan-29-yl ß-D-glucopyranoside). According to the 1 H coupling constant of anomeric protons and co-TLC of the acid hydrolysate with D-glucose, all three glycoside groups in 1 were revealed as ß-D-glucopyranosyl. Furthermore, SOD-like antioxidant activity evaluation via IC50 of 12.43, 6.78, 9.12, 6.94 and 4.85 µM revealed that compounds 2-6 had bioactivity.

Structural determination and in vitro tumor cytotoxicity evaluation of five new cycloartane glycosides from Asplenium ruprechtii Sa. Kurata.

Five new cycloartane glycosides, named aspleniumside A - E, were discovered and characterized by re-investigated the remaining extracts of the whole plant of Asplenium ruprechtii Sa. Kurata, a famous folk medicine for treating thromboangitis obliterans in China, Japan, and Korea. Compounds 3-5 possessed the 9,19-seco-cycloartane-9,11-en triterpene aglycone with 3,7(or 23),24,25,30-highly oxidized methylene, methylene or quaternary carbons, that was found in this species for the first time. The stereo-chemistry of all new compounds were fully discussed by extensive analysis of the 1D and 2D NMR data, and comparisons with those data of known compounds. 24R configuration was determined here which indicated the different growing areas of the same species could influence the secondary metabolic behavior, leading to the differences in chemical composition. All glycoside groups were determined as ß-d-glucopyranosyl by 1H coupling constant of anomeric protons and co-TLC of the acid hydrolysate with d-glucose. All the cycloartane glycosides were evaluated against HL-60 and HepG2 cells for cytotoxicity, compounds 1-3, showed potential cytotoxicity with the IC50 in range of 18-60 µM, while the standard sorafenib showed IC50 value of 10.61 ± 0.43 and 13.43 ± 1.12 µM against HL-60 and HepG2, respectively. The results attained in this study indicated that cycloartane glycosides should be the cytotoxicity substance in A. ruprechtii Sa. Kurata, and had the potential to be developed as tumor cytotoxicity agent applied in clinic.

Clinicopathological Characteristics and Prognosis of Hepatoid Adenocarcinoma of the Stomach: Evaluation of a Pooled Case Series.

Hepatoid adenocarcinoma of the stomach (HAS) is an extremely rare and unique gastric malignancy. The present study aimed to examine the relevance of the clinicopathological characteristics of HAS with patient prognosis. We retrospectively reviewed clinical data of 34 HAS patients treated at our institution between January 2010 and December 2016, as well as 294 cases reported prior to 2017 in research databases. Among these patients, 45.6% (115/252) had lesions in the gastric antrum and 77.0% (235/305) were male. Elevated levels of serum alpha-fetoprotein (AFP) were detected in most patients (75/93, 80.6%). Vascular invasion (199/286, 69.6%), lymph node metastasis (222/283, 78.4%), and preoperative distant metastasis (121/328, 36.9%) were commonly observed. The 5-year disease-free survival (DFS) and disease-specific survival (DSS) were 20.7% and 29.2%, respectively. DFS and DSS of patients receiving neoadjuvant therapy were significantly higher than those of patients receiving postoperative adjuvant therapy [DFS: P<0.001, hazard ratio (HR)=-1.831, 95% confidence interval (CI): 0.060-0.429; DSS: P<0.001, HR=-2.185, 95% CI: 0.032-0.401]. In conclusion, HAS exhibits distinct clinicopathological characteristics and a strikingly worse prognosis when compared with common gastric cancer. Complete surgery, early pTNM stage, and adjuvant therapy may predict a more favorable prognosis. Neoadjuvant therapy is strongly recommended for patients with lymph node metastasis or/and preoperative distant metastasis.

Gene expression pattern of TCR repertoire and alteration expression of IL-17A gene of γδ T cells in patients with acute myocardial infarction.

BACKGROUND: γδ T cells are associated with the pathogenesis of coronary atherosclerotic heart disease, but the relationship between the development of acute myocardial infarction (AMI) and γδ T cells is not clear. So we attempt to investigate the expression pattern and clonality of T cell receptor (TCR) repertoire of γδ T cells in AMI patients, analyze the expression levels of regulatory genes Foxp3 and IL-17A, and characterize the correlation between γδ T cells and the pathogenesis of AMI. METHODS: 25 patients diagnosed with ST-segment-elevation AMI were enrolled and 14 healthy individuals were recruited as the controls. RT-PCR and GeneScan were used to analyze the complementarity-determining region 3 sizes of TCR γδ repertoire genes in sorted γδ T cells from peripheral blood mononuclear cells (PBMCs). RQ-PCR was used to detect the gene expression levels of Foxp3, IL-17A and TCR Vγ subfamilies in sorted γδ T cells. All the patients were followed up for recordings of clinical endpoints. RESULTS: The mRNA gene expression levels of TCR Vγ1, Vγ2, and Vγ3 subfamilies in AMI patients were significantly higher than those in healthy controls. The expression pattern was Vγ1 > Vγ2 > Vγ3 in AMI patients, while Vγ1 > Vγ3 > Vγ2 in healthy controls. The significantly restricted expression of TCR Vδ subfamilies were also found in AMI patients. The expression frequencies of TCR Vδ7 and TCR Vδ6 in AMI patients were significantly lower than those in healthy controls. The high clonal expansion frequencies of the TCR Vδ8, Vδ4 and Vδ3 were determined in AMI patients. High expression of Foxp3 gene was found in AMI PBMCs, while high expression of IL-17A was found in AMI γδ+ cells. CONCLUSIONS: Restrictive expression of TCR γδ repertoire and alteration expression of IL-17A gene are the important characteristics of γδ T cells in AMI patients, which might be related to the immune response and clinical outcome. γδ T cells might play a key role in the pathological progress of AMI and associated with the IL-17A mediated pathway.

[Basic Research of the Adenovirus-mediated hCTR1 Transfection on the Treatment of Cisplatin Resistant Cervical Cancer].

OBJECTIVE: To induce cisplatin-resistant cervical squamous carcinoma cell line and investigate the drug resistant mechanisms and adenovirus trans-gene therapeutical treatment. METHODS: The cisplatin-resistant subline,designated C-33A/cis,was originated by growing parental C-33A cells with gradually increasing doses of cisplatin. The cytotoxicity of cisplatin was determined by CCK-8 assay,and the CTR1 expression was measured by Western blot. Subcutaneous xenograft cervical tumor model was established by cisplatin-resistant C-33A/cis cell line. Recombinant adenovirus ad-hCTR1 was transfected into tumor by intratumoral injection and combined with cisplatin chemotherapy. The changes in the volume of tumor were observed and the mice were executed at 10th day after the last injection,and the expression of CTR1 in tumor tissues was detected by immunohistochemistry. RESULTS: Cisplatin-resistant cervical carcinoma C-33A/cis cells were successfully induced by gradually increased concentration of cisplatin. The cytotoxic IC50 value of cisplatin on C-33A/cis had been upgraded from (1.86±0.08) to (8.11±0.21) µmol/L,while the CTR1 was found decreased by Western blot assay. Immunohistochemical analysis indicated that CTR1 expression was increased by intratumoral injection of adenovirus ad-hCTR1, and the tumor growth of C-33A/cis drug-resistant cervical carcinoma xenograft was inhibited by ad-hCTR1 transfection combined with cisplatin. CONCLUSION: The combination therapy of ad-hCTR1 transfection and cisplatin was effective to inhibit the growth of drug-resistant C-33A/cis tumor.

A New Anti-Inflammatory Alkaloid from Roots of Heracleum dissectum.

Using various chromatographic methods, a new piperidinone alkaloid, (3S)-3-{4-[(1E)-3-hydroxyprop-1-en-1-yl]-2-methoxyphenoxy}piperidin-2-one (1), together with 10 known compounds, bergapten (2), xanthotoxol (3), isopimpinellin (4), isobergapten (5), heratomol-6-O-ß-d-glucopyranoside (6), scopoletin (7), apterin (8), 3-methoxy-4-ß-d-glucopyranosyloxypropiophenone, (praeroside; 9), tachioside (10) and coniferin (11), were isolated from roots of Heracleum dissectum Ledeb. Their structures were elucidated on the basis of physicochemical properties and the detailed interpretation of various spectroscopic data. All the isolated compounds were screened for anti-inflammatory activity in vitro. As the results, compound 1 and 8 showed significantly inhibitory activity on nitric oxide production in RAW264.7 cells.

Four new taraxastane-type triterpenoic acids from Cirsium setosum.

Four new taraxastane-type triterpenoids acids 3ß,22α-dihydroxy-20-taraxasten-30-oic acid (1), 3ß-hydroxy-22-oxo-20-taraxasten-30-oic acid (2), 3-oxo-22α-hydroxy-20- taraxasten-30-oic acid (3), and 3ß,19ß-dihydroxy-20-taraxasten-30-oic acid (4) were isolated and characterized from Cirsium setosum (Willd.) MB. Their structures were determined by the combination of 1D and 2D NMR experiments ((1)H-(1)HCOSY, HSQC, HMBC and ROESY) and mass spectrometry. Compound 2 exhibited potent selective cytotoxicity against human ovarian cancer cell line A2780 with an IC50 value of 3.9 µM.

A self-contrast approach to evaluate the inhibitory effect of chrysosplenetin, in the absence and presence of artemisinin, on the in vivo P-glycoprotein-mediated digoxin transport activity.

In this study, we used a self-contrast method, which excluded the individual difference, to evaluate the inhibitory effect of chrysosplentin (CHR) in the presence or absence of artemisinin (ART) on the P-glycoprotein (P-gp) transport activity. A sensitive and rapid UHPLC-MS/MS method was applied for quantification of digoxin, a P-gp-specific substrate, in rat plasma. A pharmacokinetic study was carried out: first after an oral administration of digoxin at a dose of 0.09 mg/kg (first period), followed by a 20-day wash-out, then after another administration of digoxin (second period). During the second period, test compounds were orally given three times per day for seven consecutive days. Results showed that the t1/2 of digoxin in all the groups had no significant difference between the first and second periods. The AUC0-24 , Cmax , tmax , and Clz /F of the negative control and ART alone groups showed no difference. However, the AUC0-24 and Cmax in the CHR alone, CHR-ART (1:2) and verapamil (positive control) groups showed 2.34-, 3.04-, 1.79-, and 1.81-, 1.99-, 2.06-fold increases along with 3.50-, 3.84- and 4.76-fold decreases for CLz /F, respectively. The tmax in the CHR-ART (1:2) group increased 3.73-fold. In conclusion, our self-contrast study suggested that CHR, especially when combined with ART in a ratio of 1:2, inhibited P-gp activity while ART alone has no effect. Copyright © 2016 John Wiley & Sons, Ltd.

[Determination of Main Active Components and Metal Elements of Hainan Alpinia Katsumadai by Spectral Analysis].

Fourier transform infrared spectroscopy (FT-IR), UV spectroscopy and fluorescence spectroscopy combined with inductively coupled plasma mass spectrometry (ICP-MS) were employed to analyze Alpinia Katsumadai harvested from Hainan Baisha and Bawangling. The results from FT-IR indicated that the emergence of several characteristic absorption peaks around 1 051, 1 390，2 976 and 3 300 cm-1 belong to flavonoids. In light of second derivative spectra, two similar peaks around 2 977.72 and 2 899.94 cm-1 and evident differences peaks around 1 922.36 and 1 650.87 cm-1 were observed, which was obvious to identify and distinguish Hainan Alpinia Katsumadai from different geographical regions. The method of UV spectroscopy were used to determine the different characteristic spectra of Alpinia Katsumadai harvested from Hainan Baisha and Bawangling. An UV-quantitative analysis method for alpinetin and cardamonin in Alpinia Katsumadai was established according to the relevant absorption principle. The results from fluorescence experiments revealed that both ethanol extracts of Alpinia katsumadai Hayata harvestd from Baisha and Bawangling have the similar fluorescence emission spectra and excitation spectra. The difference of fluorescence intensity once again demonstrated the concentration of ethanol extracts of Alpinia katsumadai Hayata harvestd from Baisha was bigger than that of Bawangling. The good relative recovery was in the range of 84.28%～117.41% with the RSDs (n=3) of 0.42%～0.29%. The content of alpinetin and cardamomin in Alpinia katsumadai from various habitats are 4.23% and 3.83% for Baisha, 3.72% and 3.34% for Baiwangling, respectively. Seventeen kinds of metal elements including K, Ca, Mg, Na, Al，Fe, Mn, Cu, Zn et al were determined by using microwave digestion-ICP-MS. Therefore, FT-IR combined with second derivative spectra was an express and comprehensive to analyze and evaluate the subtle difference among different habitats. It was also indicated that the method of UV-absorption spectroscopy was simple and reliable for identifing Alpinia katsumadai from differern geographical regions and cultivation batches, and determining qualitatively and quantitatively their main active components. The determination results of metal elements according to ICP-MS experiments will provide a theoretical foundation for the further development and utilization Alpinia katsumadai and enhancing the food danger consciousness.

[Influence of diethyl sulfate (DES) mutagenesis on growth properties and pigment secondary metabolites of Phellinus igniarius].

The diethyl sulfate (DES) mutagenesis was chosen for the mutagenic treatment to Phellinus igniarius, and the relationship of mutagenesis time and death rate was investigated with 0.5% DES. The differences of mycelial growth speed, liquid fermentation mycelia biomass, morphology and pigment classes of secondary metabolites production speed and antioxidant activities of metabolite products were discussed. The study displayed that DES mutagenesis could change mycelial morphology without obvious effect on mycelium growth, and the DES mutagenesis improved antioxidant activities of the active ingredients of P. igniarius and had more antioxidant activity of hypoxia/sugar PC12 nerve cells than that of P. igniarius.

The gene expression patterns of BMPR2, EP300, TGFß2, and TNFAIP3 in B-Lymphoma cells.

OBJECTIVE: The results of a previous study showed that a clear dysregulation was evident in the global gene expression of the BCL11A-suppressed B-lymphoma cells. In this study, the bone morphogenetic protein receptor, type II (BMPR2), E1A binding protein p300 (EP300), transforming growth factor-ß2 (TGFß2), and tumor necrosis factor, and alpha-induced protein 3 (TNFAIP3) gene expression patterns in B-cell malignancies were studied. METHODS: The relative expression levels of BMPR2, EP300, TGFß2, and TNFAIP3 mRNA in B-lymphoma cell lines, myeloid cell lines, as well as in cells from healthy volunteers, were determined by real-time quantitative reverse transcript-polymerase chain reaction (qRT-PCR) with SYBR Green Dye. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as reference. RESULTS: The expression level of TGFß2 mRNA in B-lymphoma cell lines was significantly higher than those in the cells from the healthy control (P<0.05). However, the expression level of TNFAIP3 mRNA in B-malignant cells was significantly lower than that of the healthy control (P<0.05). The expression levels of BMPR2 and EP300 mRNA showed no significant difference between B-malignant cell lines and the healthy group (P>0.05). In B-lymphoma cell lines, correlation analyses revealed that the expression of BMPR2 and TNFAIP3 (r=0.882, P=0.04) had significant positive relation. The expression levels of BMPR2, EP300, and TNFAIP3 mRNA in cell lines from myeloid leukemia were significantly lower than those in the cells from the healthy control (P<0.05). The expression levels of TGFß2 mRNA showed no significant difference between myeloid leukemia cell lines and the healthy control or B-malignant cell lines (P>0.05). The expression levels of BMPR2, EP300, and TNFAIP3 mRNA in B-lymphoma cells were significantly higher than those of the myeloid leukemia cells (P<0.05). CONCLUSION: Different expression patterns of BMPR2, EP300, TGFß2, and TNFAIP3 genes in B-lymphoma cells exist.

Assessment of atrial fibrillation and vulnerability in patients with Wolff-Parkinson-White syndrome using two-dimensional speckle tracking echocardiography.

PURPOSE: The aim was to assess atrial fibrillation (AF) and vulnerability in Wolff-Parkinson-White (WPW) syndrome patients using two-dimensional speckle tracking echocardiography (2D-STE). METHODS: All patients were examined via transthoracic echocardiography and 2D-STE in order to assess atrial function 7 days before and 10 days after RF catheter ablation. A postoperative 3-month follow-up was performed via outpatient visit or telephone calls. RESULTS: Results showed significant differences in both body mass index (BMI) and supraventricular tachycardia (SVT) duration between WPW patients and DAVNP patients (both P<0.05). Echocardiography revealed that the maximum left atrial volume (LAVmax) and the left ventricular mass index (LVMI) in diastole increased noticeably in patients with WPW compared to patients with DAVNP both before and after ablation (all P<0.05). Before ablation, there were obvious differences in the levels of SRs, SRe, and SRa from the 4-chamber view (LA) in the WPW patients group compared with patients in the DAVNP group (all P<0.05). In the AF group, there were significant differences in the levels of systolic strain rate (SRs), early diastolic strain rate (SRe), and late diastolic strain rate (SRa) from the 4-chamber view (LA) both before and after ablation (all P<0.05). In the non-AF group, there were decreased SRe levels from the 4-chamber view (LA/RA) pre-ablation compared to post-ablation (all P<0.05). CONCLUSION: Our findings provide convincing evidence that WPW syndrome may result in increased atrial vulnerability and contribute to the development of AF. Further, RF catheter ablation of AAV pathway can potentially improve atrial function in WPW syndrome patients. Two-dimensional speckle tracking echocardiography imaging in WPW patients would be necessary in the evaluation and improvement of the overall function of RF catheter ablation in a long-term follow-up period.

[Autotoxicity of aqueous extracts from plant of cultivated Astragalus membranaceus var. mongholicus].

OBJECTIVE: To exploring the relationship between continuous cropping obstacle and autotoxicity of Astragalus membranaceus var. mongholicus, autotoxic effect of plant aqueous extract were determined. METHODS: Distilled water (CK), aqueous extract of plant, including root, stem and leaf (12.5, 25, 50 and 100 mg/mL respectively)were applied to testing their effect on early growth of Astragalus membranaceus var. mongholicus. Specifically, seed germination rate, germination index, emergence rate, elongation of radical and embryo, and seedling vigor index were determined. RESULTS: The aqueous extract of root, stem, and leaf at 25 mg/mL significantly inhibited the seed germination and seedling growth of Astragalus membranaceus var. mongholicus, and this inhibitory effect generally increased with the increase of the concentration of aqueous extracts. To the comprehensive allelopathic effect, the extracts from Astragalus membranaceus var. mongholicus stem were more inhibitory than those from leaf and root. The germination index and seedling vigor index were more sensitive to extract than other determined parameters. CONCLUSION: Aqueous extracts from Astragalus membranaceus var. mongholicus plant gave inhibitory effects on Astragalus. membranaceus var. mongholicus germination and seedling growth, and this inhibitory effect generally increased with the increases of aqueous extract concentration at a certain ranges. In conclusion, there is an autotoxicity in continuous cropping of Astragalus membranaceus var. mongholicus.

A new periplogenin cardenolide from the seeds of Antiaris toxicaria.

A new periplogenin cardenolide, periplogulcoside (1), together with three known cardenolides, was isolated from the seeds of Antiaris toxicaria. The structure of the new compound was characterized as periplogenin-3-O-ß-D-glucopyranosyl-(1 → 4)-ß-D-glucopyranoside (1) by spectroscopic methods including 1D and 2D NMR, HR-TOF-MS, and CD spectrometry, and the known compounds were identified by comparison of their NMR and HR-TOF-MS data with those reported in the literature. Compound 1 showed significant cytotoxicity against Hela and HepG-2 cell lines.

The biophysical property of A549 cells transferred by VEGF-D.

Vascular endothelial growth factor-D (VEGF-D) together with VEGF-C is considered to be associated with lymphangiogenesis and angiogenesis and involve in tumorization. This study aims to investigate the influence of exogenous VEGF-D gene on the biophysical property of cell surface of lung adenocarcinoma cell line. A panel of lung adenocarcinoma cell lines were examined the expression of VEGF-D and VEGF-C by real-time PCR. The VEGF-D recombinant plasmid containing enhanced green fluorescence protein (EGFP) was constructed and transfected to the cell line with no expression of VEGF-D and confirmed by real-time PCR and Western blot analysis. Topographic images of cells were obtained by using atomic force microscope (AFM) in contact mode. Unlike VEGF-C, VEGF-D was found to have a very low expression or undetectable expression in lung adenocarcinoma cell lines. The VEGF-D recombinant plasmid had been constructed successfully and was transferred into the human lung adenocarcinoma cell line A549 cells which had no endogenous expression of VEGF-D, and exogenous VEGF-D could be detected in mRNA and protein expression levels in the gene modified cells, while the VEGF-C gene expression had no change after VEGF-D transfection. After transfection, the irregular microspikes or nano clusters could observe on the surface of A549 cells, and VEGF-D transfected A549 cells became more rigid. The exogenous VEGF-D gene might cause the remarkable biophysical architectural changes in the A549 cells, which might as a novel biomarker for evaluation of its biological function.

Cytotoxic monacolin analogs from Monascus purpureus-fermented rice.

Two new monacolin analogs, monacolins O (1) and P (2), along with three known analogs, have been isolated from the ethanolic extract of Monascus purpureus-fermented rice. Their structures and absolute configurations were elucidated by spectroscopic methods, especially 2D NMR and CD spectral analyses as well as chemical method. Both 1 and 2 were tested against five tumor cell lines, and compound 1 exhibited selective cytotoxic activity against A2780 and A549 cell lines, with IC50 values of 3.7 and 8.0 µM, respectively.