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Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells

Vinblastine is a vinca alkaloid that binds to tubulin and inhibits microtubule formation in cells. Vinblastine treatment results in the formation of paracrystalline aggregates in the cells, which are formed from tightly packed tubulin molecules. Mitotic spindle assemblies in treated cells are disrup...

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Autores principales: NAKAMURA, YUKA, ISHIGAKI, YASUHITO
Formato: Online Artículo Texto
Lenguaje:English
Publicado: D.A. Spandidos 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4214480/
https://www.ncbi.nlm.nih.gov/pubmed/25364400
http://dx.doi.org/10.3892/ol.2014.2549
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author NAKAMURA, YUKA
ISHIGAKI, YASUHITO
author_facet NAKAMURA, YUKA
ISHIGAKI, YASUHITO
author_sort NAKAMURA, YUKA
collection PubMed
description Vinblastine is a vinca alkaloid that binds to tubulin and inhibits microtubule formation in cells. Vinblastine treatment results in the formation of paracrystalline aggregates in the cells, which are formed from tightly packed tubulin molecules. Mitotic spindle assemblies in treated cells are disrupted and cell cycle progression is arrested at the mitosis phase. Vinblastine is therefore widely used for cancer treatment. However, the mechanism underlying paracrystal formation has not been fully elucidated. The present study attempted to observe paracrystal formation in human A549 cells. Initally, paracrystal formation was detected using the anti-tubulin antibody. Secondly, the exogenousuly expressed RFP-conjugated tubulin also formed paracrystals. Additionally, immunostaining with the anti-RBM8A antibody overlapped with paracrystal images obtained from RFP conjugated tubulin. This suggested that the localization of the RBM8A proteins was adjacent to the tubulin molecules prior to vinblastine treatment. Furthermore, a time-lapse analysis was developed for paracrystal formation in viable human A549 cells. This was achieved using exogenous expression of fluorescent proteins conjugated with tubulin and time-lapse microscopy. It may be concluded that the indicated method was successful for the real-time analysis of paracrystal formation in human cells.
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spelling pubmed-42144802014-10-31 Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells NAKAMURA, YUKA ISHIGAKI, YASUHITO Oncol Lett Articles Vinblastine is a vinca alkaloid that binds to tubulin and inhibits microtubule formation in cells. Vinblastine treatment results in the formation of paracrystalline aggregates in the cells, which are formed from tightly packed tubulin molecules. Mitotic spindle assemblies in treated cells are disrupted and cell cycle progression is arrested at the mitosis phase. Vinblastine is therefore widely used for cancer treatment. However, the mechanism underlying paracrystal formation has not been fully elucidated. The present study attempted to observe paracrystal formation in human A549 cells. Initally, paracrystal formation was detected using the anti-tubulin antibody. Secondly, the exogenousuly expressed RFP-conjugated tubulin also formed paracrystals. Additionally, immunostaining with the anti-RBM8A antibody overlapped with paracrystal images obtained from RFP conjugated tubulin. This suggested that the localization of the RBM8A proteins was adjacent to the tubulin molecules prior to vinblastine treatment. Furthermore, a time-lapse analysis was developed for paracrystal formation in viable human A549 cells. This was achieved using exogenous expression of fluorescent proteins conjugated with tubulin and time-lapse microscopy. It may be concluded that the indicated method was successful for the real-time analysis of paracrystal formation in human cells. D.A. Spandidos 2014-12 2014-09-18 /pmc/articles/PMC4214480/ /pubmed/25364400 http://dx.doi.org/10.3892/ol.2014.2549 Text en Copyright © 2014, Spandidos Publications http://creativecommons.org/licenses/by/3.0 This is an open-access article licensed under a Creative Commons Attribution-NonCommercial 3.0 Unported License. The article may be redistributed, reproduced, and reused for non-commercial purposes, provided the original source is properly cited.
spellingShingle Articles
NAKAMURA, YUKA
ISHIGAKI, YASUHITO
Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells
title Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells
title_full Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells
title_fullStr Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells
title_full_unstemmed Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells
title_short Immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human A549 cells
title_sort immunostaining and time-lapse analysis of vinblastine-induced paracrystal formation in human a549 cells
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4214480/
https://www.ncbi.nlm.nih.gov/pubmed/25364400
http://dx.doi.org/10.3892/ol.2014.2549
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