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Two isopentenyl resorcinols, peperobtusin B and peperobtusin C, have been isolated from Peperomia tetraphylla. Their structures were determined on the basis of spectroscopic methods, especially 1H ...

PurposeTo develop a risk stratification system that can predict axillary lymph node (LN) metastasis in invasive breast cancer based on the combination of shear wave elastography (SWE) and conventional ultrasound.Materials and MethodsA total of 619 participants pathologically diagnosed with invasive breast cancer underwent breast ultrasound examinations were recruited from a multicenter of 17 hospitals in China from August 2016 to August 2017. Conventional ultrasound and SWE features were compared between positive and negative LN metastasis groups. The regression equation, the weighting, and the counting methods were used to predict axillary LN metastasis. The sensitivity, specificity, and the areas under the receiver operating characteristic curve (AUC) were calculated.ResultsA significant difference was found in the Breast Imaging Reporting and Data System (BI-RADS) category, the “stiff rim” sign, minimum elastic modulusof the internal tumor and peritumor region of 3 mm between positive and negative LN groups (p < 0.05 for all). There was no significant difference in the diagnostic performance of the regression equation, the weighting, and the counting methods (p > 0.05 for all). Using the counting method, a 0–4 grade risk stratification system based on the four characteristics was established, which yielded an AUC of 0.656 (95% CI, 0.617–0.693, p < 0.001), a sensitivity of 54.60% (95% CI, 46.9%–62.1%), and a specificity of 68.99% (95% CI, 64.5%–73.3%) in predicting axillary LN metastasis.ConclusionA 0–4 grade risk stratification system was developed based on SWE characteristics and BI-RADS categories, and this system has the potential to predict axillary LN metastases in invasive breast cancer.

Our previous research found the ethyl acetate extract of Peperomia tetraphylla (EAEPT) inhibited the growth of U937 cells by blocking the cell cycle and prompted apoptosis via the reactive oxygen species (ROS)-medicated mitochondria pathway. While the compounds in EAEPT which possessed the anti-tumor activity were unclear. Peperobtusin A is a phenolic compound, which was isolated from the whole plant of Peperomia tetraphylla. In this work, we found that peperobtusin A had the anti-proliferative effects against human lymphoma U937 cells and induced apoptosis in a dose dependent manner. Peperobtusin A significantly enhanced the formation of intracellular ROS and induced the loss of mitochondrial membrane potential (Δψm). And peperobtusin A could increase the ratio of Bax/Bcl-2, induce the cleavage of Bid, Caspase-3, Caspase-8 and Caspase-9 and enhance the level of P-P38. Moreover, peperobtusin A induced the accumulation of cells at S phase. Through using of inhibitors such as antioxidant NAC, pan-caspase inhibitor Z-VAD-FMK, p38 MAPK specific inhibitor SB203580, we found that intracellular ROS generation, activation of Caspases and p38 MAPK played very important roles in the apoptosis induced by peperobtusin A in U937 cells. Our results indicated that intracellular ROS generation, the Caspase-dependent and p38 MAPK signaling pathways involved in apoptosis induced by peperobtusin A in U937 cells.

The current study evaluated the cytotoxicity and the mechanism of apoptotic induction by Peperomia tetraphylla in U937 lymphoma cells. The results showed that P. tetraphylla ethyl acetate extract (EAEPT) inhibited the cell growth in U937 cells by MTT assay. After the U937 cells were treated with EAEPT, the cells exhibited marked morphological features of apoptosis (Hoechst 33342 staining) and the number of apoptotic cell (Annexin V-FITC/PI staining) increased. The treatment of EAEPT could induce loss of mitochondrial membrane potential (MMP) and increase the ROS level. Moreover, EAEPT treatment resulted in the accumulation of cells at S phase. We found that EAEPT could induce the cleavage of the caspase 3, caspase 8, caspase 9 and Bid. And the treatment of EAEPT could increase expression of Bax and down-regulate the expression of CCNB1, CCND1 and CDK1. The sub-fraction of EAEPT, namely EASub1 demonstrated the highest cytotoxicity activity on U937 cells. It was confirmed that EAEPT could inhibit the growth of U937 cells by blocking the cell cycle and prompted apoptosis via the ROS-medicated mitochondria pathway in vitro.

Neurotensin (NT), an endogenous peptide found in the central nervous system and in peripheral tissues, contributes to the pathophysiology of neurodegenerative and psychiatric diseases, cancer, inflammation, and immunomodulatory disease. NT exerts its physiological effects predominantly through its cognate high-affinity neurotensin receptor-1 (NTS1). NTS1 emerges as a druggable target; however, there are limited numbers of NTS1 active compounds reported to date. Here we reported a label-free cell phenotypic profiling model for screening NTS1 ligands and differentiating their biased agonism. Resonant waveguide grating enabled dynamic mass redistribution (DMR) assay was first optimized against cell confluency and then used to characterize the endogenous NTS1 in HT-29 cell using known agonists and antagonists. Pathway modulators were also used to deconvolute the signaling pathways of endogenous NTS1. Results showed that the NTS1 DMR assay is robust for screening and can differentiate biased agonism; and the activation of NTS1 in HT-29 triggers multiple pathways including Gq signaling and epidermal growth factor receptor transactivation. This study highlighted the power of label-free DMR assay to characterize receptor signaling and pharmacology of distinct classes of ligands for NTS1, G protein-coupled receptors in general.

GPR35受体是G蛋白偶联受体家族中的孤儿受体, 它的很多生物学功能还不是很清楚. 寻找发现GPR35受体的配体对于GPR35受体的生物学及药理学研究具有重要意义. 本研究利用无标记细胞靶点药理学筛选技术, 系统研究了天然黄酮类化合物对GPR35受体的激动活性, 新发现了草质素和异补骨脂二氢黄酮具有GPR35受体激动活性, 其中草质素 EC 50=(1.452±0.093) μmol/L, 是最具药物开发潜力的一个GPR35受体激动剂.

Background: Danshen is an important traditional Chinese medicine (TCM) used for the treatment of cardiovascular and cerebrovascular diseases. Separation and analysis of its components have been widely investigated. However, the systematical two dimensional liquid chromatography (2D-LC) methods have not been developed to comprehensively separate and characterize its components. Objective: In this work, double off-line 2D-LC methods were aimed to develop for the systematical separation of compounds from Danshen. Methods: Using solid phase extraction (SPE), the Danshen extract was divided into a medium-polar fraction (Sample I) and a weak-polar fraction (Sample II) according to their polarities. Based on reversed-phase liquid chromatography (RPLC) and hydrophilic interaction liquid chromatography (HILIC) modes, a 2D-HILIC × RPLC system and a 2D-RPLC × RPLC system were designed for the separation of Sample I and Sample II, respectively. According to reversed-phase and HILIC columns selectivities characterized in our previous reports, ZIC-HILIC and XTerra C18 were employed to build the 2D-HILIC × RPLC system and Click TE-CD and XTerra C18 for the 2D-RPLC × RPLC system, respectively. Results: The 2D-HILIC × RPLC and 2D-RPLC × RPLC systems exhibited excellent orthogonality for the separation of Sample I and Sample II, respectively. Their orthogonalities were 88.42% and 63.24%. Based on these double 2D-LC systems combined with mass spectrometry, at least 200 compounds were found and 33 compounds of them were identified, including 16 phenolic acids and 17 diterpenoid quinines. Conclusion: These results suggest that these two off-line 2D-LC methods are effective for the separation and characterization of components in Danshen.

Traditional Chinese medicines (TCMs) have been used in the clinic for thousands of years. Their reliable therapeutic efficacies are closely related to their multi-target mechanisms of action (MOAs). Discovery of these targets is important for understanding their clinical features. Danshen is the dried root of Salvia miltiorrhiza, a Traditional Chinese Medicine (TCM) used for the treatment of cardiovascular and cerebrovascular diseases. Although its clinical features are well recognized, the targets of its active constituents are poorly understood. Here, a label-free cell phenotypic assay was used to investigate the potential targets of the phenolic acids in danshen. Pharmacological profiling of 10 known phenolic acids in danshen using HT-29 and A431 cells revealed that lithospermic acid, salvianolic acid A, salvianolic acid B, salvianolic acid C and danshensu all displayed agonistic activity towards GPR35; however, salvianolic acid A and salvianolic acid C at high doses were also active in altering intracellular Ca2+ via another unknown target. Since GPR35 has been implicated in inflammation and cardiovascular diseases, the discovery of GPR35 as one target of the phenolic acids in danshen was useful for elucidating their mechanisms of action in the treatment of these diseases. This study also highlights the potential of label-free cell phenotypic assays for discovering multiple targets of TCMs.

A family of 2H-chromen-2-one derivatives were identified as G protein-coupled receptor-35 (GPR35) agonists using dynamic mass redistribution assays in HT-29 cells. The compounds with 1H-tetrazol-5-yl in 3-substituted position displayed higher potency than the corresponding carboxyl analogs, and the hydroxyl group in the 7-position also played an important role in GPR35 agonistic activity. 6-Bromo-7-hydroxy-8-nitro-3-(1H-tetrazol-5-yl)-2H-chromen-2-one (50) was found to be the most potent GPR35 agonist with an EC50 of 5.8 nM. Calculating the physicochemical properties of compounds with moderate to high potency suggested that compounds 30, 50, and 51 showed good druggability. This study provides a novel series of GPR35 agonists, and compound 50 may be a powerful tool to study GPR35.