Chemical structure of known cembrenoids and proposed structure of the compounds in F3.1.1.

<p>(A) Structure of incensole, incensole oxide and cembrene. (B) Hypothetical structure <b>1′</b> and three possible routes of its fragmentation. The structure is proposed based on ¹H NMR and the mass spectrum (EI 70 eV) of the compounds present in F3.1.1.</p

Reduction of intrinsic tryptophan of tubulin by cembrenoids and determination of dissociation constant.

<p>(A) The effects of cembrenoids on the tryptophan fluorescence spectra of tubulin are shown. Spectra are monitored in the absence (Ct) and presence of 5 µM of Cembrene, Incensole oxide, Colchicine or Paclitaxel (B) The change in the fluorescence intensity of tubulin (ΔF) is plotted against concentrations of each compound. The dissociation constant (Kd) for Cembrene, Incensole oxide, Colchicine and Paclitaxel binding to tubulin is estimated using an equation described in the methods. Data are represented as the mean ± MAD of two independent experiments.</p

Cytological analysis of control (Ct) and F3.1.1-treated U373 cells.

<p>All treatments with fraction F3.1.1 were done at 4 µg/mL for 72 h. (A) F-actin filament immunostaining with phalloidin conjugated to Alexa Fluor 488 (red) and visualization of nuclei with DAPI staining (blue). Arrows indicate irregularly shaped nuclei. (B) Flow cytometry analysis of the cell size distribution of the entire cell population as measured by electronic volume (EV). (C) Cytoskeleton immunostaining for F-actin (red) and with anti-α/β-tubulin antibodies for the microtubule arrangement (green). The scale is the same in all panels (bar  = 20 µm).</p

GC-MS analysis of fraction F3.1.1.

<p>(A) GC chromatogram of fraction F3.1.1. (B) Mass spectrum of the peak eluted at 68.43 min.</p

Crude chloroformic extract and sub-fraction F3.1.1. of <i>N. tabacum</i> (leafy galls (LG) and non-infected plants (NIP)) with antiproliferative activity against the glioblastoma U373 cell growth.

<p>Cells were treated for 72± SEM (n = 6).</p

Analysis of cell division-related processes in control (Ct), F3.1.1-, and cembrene-treated U373 cells.

<p>Cells were treated with 4 µg/mL of F3.1.1 or cembrene. (A, B) <i>In vitro</i> cell global growth after 24 and 48 h of culture of (A) cembrene- and (B) F3.1.1-treated cells. (C, D) Cell division duration over a time period of 48 h of growth in (C) cembrene- and (D) F.3.1.1- treated cells. The data are represented as the mean ± SD (n = 3).</p

Tubulin and nuclei visualization in U373 cells treated with F3.1.1 (10 µg/mL), colchicine (16 nM), paclitaxel (13 nM) or cembrene (8 µg/mL).

<p>In the DAPI panel of the F3.1.1, paclitaxel or cembrene-treated cells, the red arrow indicates a nucleus that is deteriorated to micronuclei, the arrowhead points to two nuclei linked by a chromatin bridge, and the arrow to the nucleus of a polyploid cell. In the panels with the merged pictures, the arrows show the cells that are enlarged in the lower panels. Bar  = 20 µm.</p

Analysis of cell division-related processes in control (Ct) and F3.1.1-treated U373 cells.

<p>Treatments with F3.1.1 were done at 4 µg/mL in a–e. (A) <i>In vitro</i> cell imaging by video microscopy after 24 h, 48 h and 72 h of growth. (B) <i>In vitro</i> cell global growth following 24 h, 48 h and 72 h of culture. The represented data are the mean ± SEM (n = 6). (C) Cell division duration over a time period of 72 h of growth in control (Ct) and treated cells (F.3.1.1). (D, E) Cell proportion in the different cell cycle phases at 48 h (D) and 72 h (E) for control (Ct) and F3.1.1-treated cells. The represented data are the mean ± SD (n = 3). (F) <i>In vitro</i> cell global growth at 12 h, 24 h, and 48 h of incubation with 10 µg/mL F3.1.1. The data represented are the mean ± SD (n = 3). (G) The occurrence of apoptopic cells (pre-G1 phase) and polyploid cells after 12 h of growth on 10 µg/mL or 50 µg/mL of F3.1.1. The represented data are the mean ± SD (n = 4).</p

Potent antiproliferative cembrenoids accumulate in tobacco upon infection with Rhodococcus fascians and trigger unusual microtubule dynamics in human glioblastoma cells.

Though plant metabolic changes are known to occur during interactions with bacteria, these were rarely challenged for pharmacologically active compounds suitable for further drug development. Here, the occurrence of specific chemicals with antiproliferative activity against human cancer cell lines was evidenced in hyperplasia (leafy galls) induced when plants interact with particular phytopathogens, such as the Actinomycete Rhodococcus fascians.Journal ArticleResearch Support, Non-U.S. Gov'tSCOPUS: ar.jinfo:eu-repo/semantics/publishe