2α-Hydroxyalantolactone from Pulicaria undulata: activity against multidrug-resistant tumor cells and modes of action.

BACKGROUND: Sesquiterpene lactones having α-methylene-γ-lactone moiety are promising natural metabolites showing various biological activity. One of the major metabolites isolated from Pulicaria undulata, 2α-hydroxyalantolactone (PU-1), has not been investigated in detail yet. Multidrug resistance (MDR) represents a major obstacle for cancer chemotherapy and the capability of novel natural products to overcoming MDR is of great interest. PURPOSE: Exploring the molecular modes of action for potent natural product metabolites. METHODS: The resazurin reduction assay was employed to evaluate the cytotoxicity of PU-1 on sensitive and their corresponding drug-resistant cell lines (overexpressing P-glycoprotein, BCRP, ABCB5, ΔEGFR, or TP53 knockout). Gene expression profiling was performed by transcriptome-wide mRNA microarray in the human CCRF-CEM leukemic cells after treatment with PU-1. The top significantly up- or down-regulated genes were identified by Chipster program and analyzed using Ingenuity Pathway Analysis (IPA) software. Finally, flow cytometry and Western blotting were performed for cell cycle analyses and apoptosis detection. RESULTS: The sesquiterpene lactone, PU-1, showed potent cytotoxicity towards the drug-sensitive and -resistant cell lines. Transcriptome-wide mRNA expression profiling and pathway analysis pointed to genes involved in DNA damage response and G2/M cell cycle arrest. G2/M arrest was verified by flow cytometry and further confirmed by the upregulation of p21 and downregulation of p-CDC25C expression in Western blotting. Moreover, the suggested DNA damage checkpoint regulation was confirmed by immunofluorescence and Western blotting by upregulation of pS345 Chk1, p-H3 and γ-H2AX. Furthermore, PU-1 inhibited PI3K/AKT pathway, which is involved in signaling DNA damage and G2/M arrest. Cells ultimately induced apoptosis upon PU-1 treatment. CONCLUSIONS: PU-1 is a potent natural product inhibiting otherwise drug-resistant human tumor cell growth through DNA damage, G2/M cell cycle arrest and apoptosis.

Polysulfone Membranes Embedded with Halloysites Nanotubes: Preparation and Properties.

In the present study, nanocomposite ultrafiltration membranes were prepared by incorporating nanotubes clay halloysite (HNTs) into polysulfone (PSF) and PSF/polyvinylpyrrolidone (PVP) dope solutions followed by membrane casting using phase inversion method. Characterization of HNTs were conducted using scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy-dispersive X-ray spectroscopy (EDS), X-ray diffraction (XRD), and thermogravimetric (TGA) analysis. The pore structure, morphology, hydrophilicity and mechanical properties of the composite membranes were characterized by using SEM, water contact angle (WCA) measurements, and dynamic mechanical analysis. It was shown that the incorporation of HNTs enhanced hydrophilicity and mechanical properties of the prepared PSF membranes. Compared to the pristine PSF membrane, results show that the total porosity and pore size of PSF/HNTs composite membranes increased when HNTs loadings were more than 0.5 wt % and 1.0 wt %, respectively. These findings correlate well with changes in water flux of the prepared membranes. It was observed that HNTs were homogenously dispersed within the PSF membrane matrix at HNTs content of 0.1 to 0.5 wt % and the PSF/HNTs membranes prepared by incorporating 0.2 wt % HNTs loading possess the optimal mechanical properties in terms of elastic modulus and yield stress. In the case of the PSF/PVP matrix, the optimal mechanical properties were obtained with 0.3 wt % of HNTs because PVP enhances the HNTs distribution. Results of bovine serum albumin (BSA) filtration tests indicated that PSF/0.2 wt % HNTs membrane exhibited high BSA rejection and notable anti-fouling properties.

Novel Insights in the Regulation of Phosphatidylserine Exposure in Human Red Blood Cells.

BACKGROUND/AIMS: In previous publications we were able to demonstrate the exposure of phosphatidylserine (PS) in the outer membrane leaflet after activation of red blood cells (RBCs) by lysophosphatidic acid (LPA), phorbol-12 myristate-13acetate (PMA), or 4-bromo-A23187 (A23187). It has been concluded that three different mechanisms are responsible for the PS exposure in human RBCs: (i) Ca2+-stimulated scramblase activation (and flippase inhibition) by A23187, LPA, and PMA; (ii) PKCα activation by LPA and PMA; and (iii) enhanced lipid flip flop caused by LPA. Further studies aimed to elucidate interconnections between the increased Ca2+ content, scramblase- and PKCα-activation. In addition, the role of the Ca2+-activated K+ channel (Gardos channel) activity in the process of PS exposure needs to be investigated. METHODS: The intracellular Ca2+ content and the PS exposure of RBCs have been investigated after treatment with LPA (2.5 µM), PMA (6 µM), or A23187 (2 µM). Fluo-4 and annexin V-FITC has been used to detect intracellular Ca2+ content and PS exposure, respectively. Both parameters (Ca2+ content, PS exposure) were studied using flow cytometry. Inhibitors of the scramblase, the PKCα, and the Gardos channel have been applied. RESULTS: The percentage of RBCs showing PS exposure after activation with LPA, PMA, or A23187 is significantly reduced after inhibition of the scramblase using the specific inhibitor R5421 as well as after the inhibition of the PKCα using chelerythrine chloride or calphostin C. The inhibitory effect is more pronounced when the scramblase and the PKCα are inhibited simultaneously. Additionally, the inhibition of the Gardos channel using charybdotoxin resulted in a significant reduction of the percentage of RBCs showing PS exposure under all conditions measured. Similar results were obtained when the Gardos channel activity was suppressed by increased extracellular K+ content. CONCLUSION: PS exposure is mediated by the Ca2+-dependent scramblase but also by PKCα activated by LPA and PMA in a Ca2+-dependent and a Ca2+-independent manner. Furthermore, we hypothesize that a hyperpolarisation of RBCs caused by the opening of the Gardos channel is essential for the scramblase activity as well as for a fraction of the LPA-induced Ca2+ entry.

Biovalorization of friedelane triterpenes derived from cork processing industry byproducts.

Here, we describe the synthesis, bioactivity screening, and structure-activity relationships of various synthetic triterpenoids prepared from the cork processing byproducts friedelin (1) and 3-hydroxyfriedel-3-en-2-one (2) via oxidative procedures. The synthesis of compounds 2alpha-trimethylsiloxyfriedelan-3-one (17), friedelin-2,3-lactone (18), friedelin-3-oxime (19), and friedelin-3,4-lactam (20) is also described. We have studied the insecticidal and phytotoxic potential of these compounds, their selective cytotoxic effects on insect and mammalian cells, and their antiparasitic effects. Structural modifications of the A-ring of friedelin (1) improved its insecticidal activity with derivatives 5, 2,3-secofriedelan-2-al-3-oic acid (6), its acetylated derivative 6a, 3beta- and 3alpha-hydroxyfriedelane (9 and 10), 3alpha-hydroxyfriedel-2-one (11), 4beta-hydroxyfriedel-3-one (16), the acetylated 10a, 3,4-secofriedelan-4-oxo-3-oic-acid (14), lactone 18, and the oxime 19 being stronger insecticides than the parent compound. Methyl-3-nor-2,4-secofriedelan-4-oxo-2-oic acid (12) and its acetylated derivative 12a also showed insecticidal activity in contrast to their inactive parent compound 2. The postingestive effects and cytotoxicity of these compounds suggest a multifaceted insecticidal mode of action. These structural modifications did not result in better phytotoxic agents than the parent compounds except for lactam 20 and yielded several moderately active antiparasite derivatives (seco acids 6, 12, 14, and 4beta-hydroxyfriedel-3-one 16) with cytotoxic effects on mammalian cells.