Frizzled 6 endows high-grade serous ovarian cancer with stem-like properties and chemoresistance.

Stem-like properties contribute to tumor growth, metastasis, and chemoresistance. High-grade serous ovarian cancer (HGSOC) exhibits a very aggressive phenotype characterized by extensive metastasis, rapid progression, and therapy resistance. Frizzled 6 (FZD6) is overexpressed in HGSOC, and higher levels of FZD6 have been associated with shorter survival times in patients with HGSOC. Functionally, FZD6 promotes HGSOC growth and peritoneal metastasis. It endues HGSOC cells with stem-like properties by modulating POU5F1, ALDH1, and EPCAM. It can also desensitize HGSOC cells to certain chemical drugs. As a putative ligand for FZD6, WNT7B is also implicated in cell proliferation, stem-like properties, invasion and migration, and chemoresistance. SMAD7 is a downstream component of FZD6 signaling that is thought to mediate FZD6-associated phenotypes, at least in part. Therefore, FZD6/WNT7B-SMAD7 can be considered a tumor-promoting signaling pathway in HGSOC that may be responsible for tumor growth, peritoneal metastasis, and chemoresistance.

Association of the LEP gene with immune infiltration as a diagnostic biomarker in preeclampsia.

Objective: Preeclampsia (PE) is a serious condition in pregnant women and hence an important topic in obstetrics. The current research aimed to recognize the potential and significant immune-related diagnostic biomarkers for PE. Methods: From the Gene Expression Omnibus (GEO) data sets, three public gene expression profiles (GSE24129, GSE54618, and GSE60438) from the placental samples of PE and normotensive pregnancy were downloaded. Differentially expressed genes (DEGs) were selected and determined among 73 PE and 85 normotensive control pregnancy samples. The DEGs were used for Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Disease Ontology (DO) enrichment analysis, and Gene Set Enrichment Analysis (GSEA). The candidate biomarkers were identified by the least absolute shrinkage and selection operator (LASSO) and support vector machine recursive feature elimination (SVM-RFE) analysis. The receiver operating characteristic curve (ROC) was applied to evaluate diagnostic ability. For further confirmation, the expression levels and diagnostic value of biomarkers in PE were verified in the GSE75010 data set (80 PE and 77 controls) and validated by qRT-RCR, Western blot, and immunohistochemistry (IHC). The CIBERSORT algorithm was used to calculate the compositional patterns of 22 types of immune cells in PE. Results: In total, 15 DEGs were recognized. The GO and KEGG analyses revealed that the DEGs were enriched in the steroid metabolic process, receptor ligand activity, GnRH secretion, and neuroactive ligand-receptor interaction. The recognized DEGs were primarily implicated in cell-type benign neoplasm, kidney failure, infertility, and PE. Gene sets related to hormone activity, glycosylation, multicellular organism process, and response to BMP were activated in PE. The LEP gene was distinguished as a diagnostic biomarker of PE (AUC = 0.712) and further certified in the GSE75010 data set (AUC = 0.850). The high expression of LEP was associated with PE in clinical samples. In addition, the analysis of the immune microenvironment showed that gamma delta T cells, memory B cells, M0 macrophages, and regulatory T cells were positively correlated with LEP expression (P < 0.05). Conclusion: LEP expression can be considered to be a diagnostic biomarker of PE and can offer a novel perspective for future studies regarding the occurrence and molecular mechanisms of PE.

Wogonoside inhibits tumor growth and metastasis in endometrial cancer via ER stress-Hippo signaling axis.

Wogonoside, a bioactive flavonoid component derived from Scutellaria baicalensis Georgi, has been reported to inhibit tumor growth in mice bearing various types of cancer cells such as breast cancer, lung cancer, and leukemia cells. However, whether wogonoside could inhibit tumor growth of endometrial cancer has not been elucidated. In this study, we explored the function of wogonoside on tumor growth and the underlying mechanism on endometrial cancer. Firstly, we investigated the effect of wogonoside on endometrial cancer cells and found that wogonoside could significantly decrease cell proliferation and metastasis. Mechanistically, wogonoside could aggravate the extent of ER stress and upregulate the phosphorylation level of Mammalian Ste20-like kinase 1, leading to the activation of the Hippo signaling pathway. Taken together, in vitro and in vivo data demonstrated that wogonoside could be a potent inducer of ER stress and could be further developed into a promising therapy for endometrial cancer.

Clinical diagnosis and genetic counseling of atypical ataxia­telangiectasia in a Chinese family.

Ataxia­telangiectasia (A­T) is an autosomal recessive chromosome breakage disorder caused by mutations in the ATM serine/threonine kinase (ATM) gene. Typically, it presents in early childhood with progressive cerebellar dysfunction, accompanied by immunodeficiency and oculocutaneous telangiectasia. In the present study, the clinical and genetic findings of a Chinese family affected with A­T in two live siblings, the proband (II­2) and his elder brother (II­1), as well as a fetus (II­3) were reported. General health, clinical neurological, electrophysiological (motor and sensory nerve conduction) and magnetic resonance imaging evaluations revealed that patients II­1 and II­2 had similar symptoms of ataxia, dysarthria, conjunctival hyperemia and elevated serum α­fetoprotein, whereas patient II­1 had earlier A­T onset at 2 years old and more serious problems with movement and intelligence. Targeted sequencing followed by Sanger sequencing revealed that these two patients carried the compound heterozygotes of a novel nonsense mutation c.5170G>T (p.Glu1724Ter) and a known nonsense mutation c.748C>T (p.Arg250Ter) in the ATM gene. Each mutation was inherited from an asymptomatic parent, which therefore confirmed the diagnosis of A­T. Given this, proband's mother performed prenatal diagnosis in her third pregnancy. Unfortunately, the fetus had the same causal mutations as its siblings and the pregnancy was terminated. The findings of the present study expanded the mutation spectrum of the ATM gene and may help in understanding the genetic basis of A­T, in order to guide genetic counseling and prenatal diagnosis.

An ex vivo model for anti-angiogenic drug testing on intact microvascular networks.

New models of angiogenesis that mimic the complexity of real microvascular networks are needed. Recently, our laboratory demonstrated that cultured rat mesentery tissues contain viable microvascular networks and could be used to probe pericyte-endothelial cell interactions. The objective of this study was to demonstrate the efficacy of the rat mesentery culture model for anti-angiogenic drug testing by time-lapse quantification of network growth. Mesenteric windows were harvested from adult rats, secured in place with an insert, and cultured for 3 days according to 3 experimental groups: 1) 10% serum (angiogenesis control), 2) 10% serum + sunitinib (SU11248), and 3) 10% serum + bevacizumab. Labeling with FITC conjugated BSI-lectin on Day 0 and 3 identified endothelial cells along blood and lymphatic microvascular networks. Comparison between day 0 (before) and 3 (after) in networks stimulated by 10% serum demonstrated a dramatic increase in vascular density and capillary sprouting. Growing networks contained proliferating endothelial cells and NG2+ vascular pericytes. Media supplementation with sunitinib (SU11248) or bevacizumab both inhibited the network angiogenic responses. The comparison of the same networks before and after treatment enabled the identification of tissue specific responses. Our results establish, for the first time, the ability to evaluate an anti-angiogenic drug based on time-lapse imaging on an intact microvascular network in an ex vivo scenario.

Chemometrics-based approach to modeling quantitative composition-activity relationships for Radix Tinosporae.

Quantitative composition-activity relationship (QCAR) study makes it possible to discover active components in traditional Chinese medicine (TCM) and to predict the integral bioactivity by its chemical composition. In the study, 28 samples of Radix Tinosporae were quantitatively analyzed by high performance liquid chromatography, and their analgesic activities were investigated via abdominal writhing tests on mice. Three genetic algorithms (GA) based approaches including partial least square regression, radial basis function neural network, and support vector regression (SVR) were established to construct QCAR models of R. Tinosporae. The result shows that GA-SVR has the best model performance in the bioactivity prediction of R. Tinosporae; seven major components thereof were discovered to have analgesic activities, and the analgesic activities of these components were partly confirmed by subsequent abdominal writhing test. The proposed approach allows discovering active components in TCM and predicting bioactivity by its chemical composition, and is expected to be utilized as a supplementary tool for the quality control and drug discovery of TCM.

Anti-inflammatory and anti-nociceptive activities of compounds from Tinospora sagittata (Oliv.) Gagnep.

Radix Tinosporae is a herb widely used in traditional Chinese medicine for the treatment of various inflammatory diseases. In the present study, its anti-inflammatory and antinociceptive activities were investigated. The ethanol extract of Radix Tinosporae exhibited significant inhibitory effects on xylene-induced ear edema and acetic acid-induced writhing in mice. Using bioassay-guided fractionation, the n-butanol fraction was determined as the active fraction. Further purification of the most active n-butanol fraction led to the isolation of three compounds, palmatine, columbamine and columbinyl glucoside. All three compounds showed inhibitory activities on xylene-induced ear edema, but only palmatine and columbamine exhibited significant inhibitory effects on acetic acid-induced writhing. In addition, palmatine and columbamine markedly inhibited in vitro production of nitric oxide and nuclear factor-kappaB activation in RAW264.7 macrophage cells in response to lipopolysaccharide or tumor necrosis factor alpha stimulation. These results provide justification for the utilization of Radix Tinosporae in Chinese folk medicine for the treatment of inflammatory diseases.

A new flavanol from Daphne genkwa.

A new flavanol, 5, 7, 4'-trihydroxy-8-ethoxycarbonylflavanol (1), was isolated from the ethanol extract of Daphne genkwa Sieb. et Zucc. Its structure was defined by spectroscopic methods.

Quantitative LC/MS/MS method and pharmacokinetic studies of columbin, an anti-inflammation furanoditerpen isolated from Radix Tinosporae.

Columbin is an important component isolated from Radix Tinosporae. It has been demonstrated to possess many pharmacological activities, including anti-inflammation, antitumor and inhibition of enzyme activity in vivo. The purpose of the present study was to examine in vivo pharmacokinetics and bioavailability of columbin in rats using a high-performance liquid chromatography coupled with tandem mass spectrometry quantitative detection method. The columbin was extracted from rat plasma samples by methyl tert-butyl ether, evaporated and reconstituted in 100 microL methanol prior to analysis. The separation was performed using a Luna reversed-phase analytical column (5 microm, 100 x 2.0 mm) and an SB-C18 guard column (5 microm, 20 x 4.0 mm). The mobile phase was a mixture of methanol and water containing 25 mmoL/L NH(4)Ac (80:20, v/v). The method was validated within the concentration range of 5-5000 ng/mL, and the calibration curves were linear with correlation coefficients (r) >0.999. It was further applied to assess pharmacokinetics and oral bioavailability of columbin after i.v. and oral administration to rats. The oral bioavailability of columbin was only 3.18%, which indicated that columbin had poor absorption or underwent extensive first-pass metabolism.

Simultaneous determination of eight components in Radix Tinosporae by high-performance liquid chromatography coupled with diode array detector and electrospray tandem mass spectrometry.

High-performance liquid chromatography (HPLC) coupled with electrospray tandem mass spectrometry (ESI-MS-MS) and diode array detection (DAD) was used to identify and simultaneously determine eight major ingredients in Radix Tinosporae. The assay was performed on a Diamonsil C(18) analytical column with a gradient solvent system of A (water containing 0.2% formic acid, 20mM ammonium acetate) and B (methanol/acetonitrile=1/1, v/v). The 217, 248, 270 and 347 nm, respectively, were chosen as the monitoring wavelengths to determine four structural types of components, say columbin, phytoecdysteroids (including 20-hydroxyecdysone, 2-deoxy-20-hydroxyecdysone 3-O-beta-d-glucopyranoside and 2-deoxy-20-hydroxyecdysone), menisperine and protoberberine alkaloids (including columbamine, jatrorrhizine and palmatine). This method was validated in respect to precision, repeatability and accuracy, and was successfully applied to quantify the eight components in 39 batches of R. Tinosporae for quality control purpose. The results indicated that the proposed method could be readily utilized as a quality control method for traditional Chinese medicine (TCM).

Characterization of isoquinoline alkaloids, diterpenoids and steroids in the Chinese herb Jin-Guo-Lan (Tinospora sagittata and Tinospora capillipes) by high-performance liquid chromatography/electrospray ionization with multistage mass spectrometry.

This study sought to determine the primary components (isoquinoline alkaloids, diterpenoids and steroids) in crude extracts of the Chinese herb Jin-Guo-Lan, prepared from the roots of Tinospora sagittata and T. capillipes, by liquid chromatography/electrospray ionization multistage mass spectrometry coupled with diode-array detection (LC-DAD/ESI-MS(n)). After separation on a reversed-phase C(18) column using gradient elution, positive and negative ESI-MS experiments were performed. In positive ion mode, the three types of compounds showed very different characteristic ions: strong [M](+) or [M+H](+) ions were observed for isoquinoline alkaloids; [M+NH(4)](+) and/or [M+H-CO(2)](+) for diterpenoids; [M+H-nH(2)O](+) (n=1-3) for steroids. These adduct ions and/or fragments were used to deduce the mass and categories of known and unknown components in crude extracts, and their structures were further confirmed by ESI-MS(n) in positive ion mode. Moreover, UV absorption peaks obtained from DAD provided useful functional group information to aid the MS(n)-based identification. As a result, 11 compounds were unambiguously identified by comparing with standard compounds and 13 compounds were tentatively identified or deduced according to their MS(n) data. Two of these compounds (13-hydroxycolumbamine and 13-hydroxyjatrorrhizine) were found to be new compounds and another one (13-hydroxypalmatine) was detected for the first time as a natural product. In addition, a [M-*CH(3)-H(2)O](*+) ion in MS(2) of [M](+) after in-source collision-induced dissociation was used to differentiate positional isomers of protoberberine alkaloids, columbamine and jatrorrhizine. Although the roots of T. sagittata and T. capillipes contain almost identical compounds, the content of the compounds in them is dramatically different, suggesting the necessity for further comparison of the bioactivities of the two species.