Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells

Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells

Aim - the aim of the study was in vitro analysis of biological activity of recombinant human beta-defensin-4 (rec-hBD-4). Methods: hBD-4 cDNA was cloned into pGEX-2T vector, and recombinant plasmid was transformed into E. coli BL21(DE3) cells. To purify soluble fusion GST-hBD-4 protein, affinity chr...

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Видавець:Інститут експериментальної патології, онкології і радіобіології ім. Р.Є. Кавецького НАН України
Дата:2013
Автори: Gerashchenko, O.L., Zhuravel, E.V., Skachkova, O.V., Khranovska, N.N., Filonenko, V.V., Pogrebnoy, P.V., Soldatkina, M.A.
Формат: Стаття
Мова:English
Опубліковано: Інститут експериментальної патології, онкології і радіобіології ім. Р.Є. Кавецького НАН України 2013
Назва видання:Experimental Oncology
Теми:
Original contributions
Онлайн доступ:http://dspace.nbuv.gov.ua/handle/123456789/145208
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Цитувати:Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells / O.L. Gerashchenko, E.V. Zhuravel, O.V. Skachkova, N.N. Khranovska, V.V. Filonenko, P.V. Pogrebnoy, A. Soldatkina // Experimental Oncology. — 2013. — Т. 35, № 2. — С. 76–82. — Бібліогр.: 37 назв. — англ.

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Digital Library of Periodicals of National Academy of Sciences of Ukraine
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spelling irk-123456789-1452082019-01-21T01:23:18Z Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells Gerashchenko, O.L. Zhuravel, E.V. Skachkova, O.V. Khranovska, N.N. Filonenko, V.V. Pogrebnoy, P.V. Soldatkina, M.A. Original contributions Aim - the aim of the study was in vitro analysis of biological activity of recombinant human beta-defensin-4 (rec-hBD-4). Methods: hBD-4 cDNA was cloned into pGEX-2T vector, and recombinant plasmid was transformed into E. coli BL21(DE3) cells. To purify soluble fusion GST-hBD-4 protein, affinity chromatography was applied. Rec-hBD-4 was cleaved from the fusion protein with thrombin, and purified by reverse phase chromatography on Sep-Pack C18. Effects of rec-hBD-4 on proliferation, viability, cell cycle distribution, substrate-independent growth, and mobility of cultured human cancer cells of A431, A549, and TPC-1 lines were analyzed by direct cell counting technique, MTT assay, flow cytofluorometry, colony forming assay in semi-soft medium, and wound healing assay. Rech-BD-4 was expressed in bacterial cells as GST-hBD-4 fusion protein, and purified by routine 3-step procedure (affine chromatography on glutathione-agarose, cleavage of fusion protein by thrombin, and reverse phase chromatography). Analysis of in vitro activity of rec-hBD-4 toward three human cancer cell lines has demonstrated that the defensin is capable to affect cell behaviour in concentration-dependent manner. In 1 - 100 nM concentrations rec-hBD-4 significantly stimulates cancer cell proliferation and viability, and promotes cell cycle progression through G2/M checkpoint, greatly enhances colony-forming activity and mobility of the cells. Treatment of the cells with 500 nM of rec-hBD-4 resulted in opposite effects: significant suppression of cell proliferation and viability, blockage of cell cycle in G1/S checkpoint, significant inhibition of cell migration and colony forming activity. 2013 Article Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells / O.L. Gerashchenko, E.V. Zhuravel, O.V. Skachkova, N.N. Khranovska, V.V. Filonenko, P.V. Pogrebnoy, A. Soldatkina // Experimental Oncology. — 2013. — Т. 35, № 2. — С. 76–82. — Бібліогр.: 37 назв. — англ. 1812-9269 http://dspace.nbuv.gov.ua/handle/123456789/145208 en Experimental Oncology Інститут експериментальної патології, онкології і радіобіології ім. Р.Є. Кавецького НАН України
institution Digital Library of Periodicals of National Academy of Sciences of Ukraine
collection DSpace DC
language English
topic Original contributions
Original contributions
spellingShingle Original contributions
Original contributions
Gerashchenko, O.L.
Zhuravel, E.V.
Skachkova, O.V.
Khranovska, N.N.
Filonenko, V.V.
Pogrebnoy, P.V.
Soldatkina, M.A.
Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
Experimental Oncology
description Aim - the aim of the study was in vitro analysis of biological activity of recombinant human beta-defensin-4 (rec-hBD-4). Methods: hBD-4 cDNA was cloned into pGEX-2T vector, and recombinant plasmid was transformed into E. coli BL21(DE3) cells. To purify soluble fusion GST-hBD-4 protein, affinity chromatography was applied. Rec-hBD-4 was cleaved from the fusion protein with thrombin, and purified by reverse phase chromatography on Sep-Pack C18. Effects of rec-hBD-4 on proliferation, viability, cell cycle distribution, substrate-independent growth, and mobility of cultured human cancer cells of A431, A549, and TPC-1 lines were analyzed by direct cell counting technique, MTT assay, flow cytofluorometry, colony forming assay in semi-soft medium, and wound healing assay. Rech-BD-4 was expressed in bacterial cells as GST-hBD-4 fusion protein, and purified by routine 3-step procedure (affine chromatography on glutathione-agarose, cleavage of fusion protein by thrombin, and reverse phase chromatography). Analysis of in vitro activity of rec-hBD-4 toward three human cancer cell lines has demonstrated that the defensin is capable to affect cell behaviour in concentration-dependent manner. In 1 - 100 nM concentrations rec-hBD-4 significantly stimulates cancer cell proliferation and viability, and promotes cell cycle progression through G2/M checkpoint, greatly enhances colony-forming activity and mobility of the cells. Treatment of the cells with 500 nM of rec-hBD-4 resulted in opposite effects: significant suppression of cell proliferation and viability, blockage of cell cycle in G1/S checkpoint, significant inhibition of cell migration and colony forming activity.
format Article
author Gerashchenko, O.L.
Zhuravel, E.V.
Skachkova, O.V.
Khranovska, N.N.
Filonenko, V.V.
Pogrebnoy, P.V.
Soldatkina, M.A.
author_facet Gerashchenko, O.L.
Zhuravel, E.V.
Skachkova, O.V.
Khranovska, N.N.
Filonenko, V.V.
Pogrebnoy, P.V.
Soldatkina, M.A.
author_sort Gerashchenko, O.L.
title Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
title_short Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
title_full Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
title_fullStr Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
title_full_unstemmed Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
title_sort biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells
publisher Інститут експериментальної патології, онкології і радіобіології ім. Р.Є. Кавецького НАН України
publishDate 2013
topic_facet Original contributions
url http://dspace.nbuv.gov.ua/handle/123456789/145208
citation_txt Biologic activities of recombinant human-beta-defensin-4 toward cultured human cancer cells / O.L. Gerashchenko, E.V. Zhuravel, O.V. Skachkova, N.N. Khranovska, V.V. Filonenko, P.V. Pogrebnoy, A. Soldatkina // Experimental Oncology. — 2013. — Т. 35, № 2. — С. 76–82. — Бібліогр.: 37 назв. — англ.
series Experimental Oncology
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first_indexed 2023-05-20T17:21:31Z
last_indexed 2023-05-20T17:21:31Z
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