Lignans from Bursera fagaroides Affect In Vivo Cell Behavior by Disturbing the Tubulin Cytoskeleton in Zebrafish Embryos

Abstract

By using a zebrafish embryo model to guide the chromatographic fractionation of antimitotic secondary metabolites, seven podophyllotoxin-type lignans were isolated from a hydroalcoholic extract obtained from the steam bark of Bursera fagaroides. The compounds were identified as podophyllotoxin (1), β-peltatin-A-methylether (2), 5′-desmethoxy-β-peltatin-A-methylether (3), desmethoxy-yatein (4), desoxypodophyllotoxin (5), burseranin (6), and acetyl podophyllotoxin (7). The biological effects on mitosis, cell migration, and microtubule cytoskeleton remodeling of lignans 1–7 were further evaluated in zebrafish embryos by whole-mount immunolocalization of the mitotic marker phospho-histone H3 and by a tubulin antibody. We found that lignans 1, 2, 4, and 7 induced mitotic arrest, delayed cell migration, and disrupted the microtubule cytoskeleton in zebrafish embryos. Furthermore, microtubule cytoskeleton destabilization was observed also in PC3 cells, except for 7. Therefore, these results demonstrate that the cytotoxic activity of 1, 2, and 4 is mediated by their microtubule-destabilizing activity. In general, the in vivo and in vitro models here used displayed equivalent mitotic effects, which allows us to conclude that the zebrafish model can be a fast and cheap in vivo model that can be used to identify antimitotic natural products through bioassay-guided fractionation.

Lignans from Bursera fagaroides: Chemistry, Pharmacological Effects and Molecular Mechanism. A Current Review

“…The lignans 1-19 have been isolated from specimens of B. fagaroides, collected in México, principally from the bark (Michoacán state) [19,20,26,27,38,39,41,42], two reports from Oaxaca state [25] and one from Guerrero state [42] analyzed the resin. The polarity of the used extracts was chloroform (CHCl 3 ) [25,26], dichloromethane (CH 2 Cl 2 ) [20,38,40,41], ethanol (CH 3 CH 2 OH) [27], CH 3 CH 2 OH 80% (previously treated with hexane) [42], and methanol (CH 3 OH) 70% [19,29,39]. In general, the purification of these extracts was carried out by bioassay-guided chromatographic methods [19].…”

“…In addition, their 13 , respectively [19,20,[25][26][27]29,40]. On the other hand, the absolute configurations of lignans 4, 5, 7, 11, and 12 were determined using vibrational circular dichroism [27], while that of lignan 1 was determined by chemical correlation with D-phenylalanine [44] and by X-ray diffraction analysis of 2 -bromopodophyllotoxin [45].…”

“…In another in vivo study, the hydroalcoholic extract from the bark of B. fagaroides does not affect the number of Histone H3 phosphorylated at serine 10 (H3S10ph)-positive nuclei, with respect to the control without treatment, when measured in whole 24 h post-fertilization (hpf) zebrafish embryos; this indicated that the extract does not induce mitotic cells in the embryos [ 29 ]. This result contrasts with the strong in vivo antitumor activity against L5178Y lymphoma in mice and the authors attributed this to the poor bioavailability because of the low concentration of the active compounds present in the studied extract [ 19 ].…”

“…This result contrasts with the strong in vivo antitumor activity against L5178Y lymphoma in mice and the authors attributed this to the poor bioavailability because of the low concentration of the active compounds present in the studied extract [ 19 ]. Further, chromatographic fractionation afforded two rich-lignans fractions that induced a high amount of cells in mitotic arrest in zebrafish embryos [ 29 ].…”

Lignans

“…The extensive analysis of the potent biological activities of lignans remains a popular avenue of investigation. Antunez-Mojica et al used a zebrafish embryo model to guide the chromatographic fractionation of antimitotic secondary metabolites, ultimately leading to the isolation of several podophyllotoxin-type lignans from the steam bark of Bursera fagaroides [10]. Subsequent to their isolation, the biological effects on mitosis, cell migration, and microtubule cytoskeleton remodeling of the isolated lignans were then further evaluated in zebrafish embryos through various methods.…”