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PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells
BACKGROUND: Hadron therapy is an innovative technique where cancer cells are precisely killed leaving surrounding healthy cells least affected by high linear energy transfer (LET) radiation like carbon ion beam. Anti-metastatic effect of carbon ion exposure attracts investigators into the field of h...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034624/ https://www.ncbi.nlm.nih.gov/pubmed/27659937 http://dx.doi.org/10.1186/s13014-016-0703-x |
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author | Ghorai, Atanu Sarma, Asitikantha Chowdhury, Priyanka Ghosh, Utpal |
author_facet | Ghorai, Atanu Sarma, Asitikantha Chowdhury, Priyanka Ghosh, Utpal |
author_sort | Ghorai, Atanu |
collection | PubMed |
description | BACKGROUND: Hadron therapy is an innovative technique where cancer cells are precisely killed leaving surrounding healthy cells least affected by high linear energy transfer (LET) radiation like carbon ion beam. Anti-metastatic effect of carbon ion exposure attracts investigators into the field of hadron biology, although details remain poor. Poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors are well-known radiosensitizer and several PARP-1 inhibitors are in clinical trial. Our previous studies showed that PARP-1 depletion makes the cells more radiosensitive towards carbon ion than gamma. The purpose of the present study was to investigate combining effects of PARP-1 inhibition with carbon ion exposure to control metastatic properties in HeLa cells. METHODS: Activities of matrix metalloproteinases-2, 9 (MMP-2, MMP-9) were measured using the gelatin zymography after 85 MeV carbon ion exposure or gamma irradiation (0- 4 Gy) to compare metastatic potential between PARP-1 knock down (HsiI) and control cells (H-vector - HeLa transfected with vector without shRNA construct). Expression of MMP-2, MMP-9, tissue inhibitor of MMPs such as TIMP-1, TIMP-2 and TIMP-3 were checked by immunofluorescence and western blot. Cell death by trypan blue, apoptosis and autophagy induction were studied after carbon ion exposure in each cell-type. The data was analyzed using one way ANOVA and 2-tailed paired-samples T-test. RESULTS: PARP-1 silencing significantly reduced MMP-2 and MMP-9 activities and carbon ion exposure further diminished their activities to less than 3 % of control H-vector. On the contrary, gamma radiation enhanced both MMP-2 and MMP-9 activities in H-vector but not in HsiI cells. The expression of MMP-2 and MMP-9 in H-vector and HsiI showed different pattern after carbon ion exposure. All three TIMPs were increased in HsiI, whereas only TIMP-1 was up-regulated in H-vector after irradiation. Notably, the expressions of all TIMPs were significantly higher in HsiI than H-vector at 4 Gy. Apoptosis was the predominant mode of cell death and no autophagic death was observed. CONCLUSIONS: Our study demonstrates for the first time that PARP-1 inhibition in combination with carbon ion synergistically decreases MMPs activity along with overall increase of TIMPs. These data open up the possibilities of improvement of carbon ion therapy with PARP-1 inhibition to control highly metastatic cancers. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13014-016-0703-x) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-5034624 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-50346242016-09-29 PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells Ghorai, Atanu Sarma, Asitikantha Chowdhury, Priyanka Ghosh, Utpal Radiat Oncol Research BACKGROUND: Hadron therapy is an innovative technique where cancer cells are precisely killed leaving surrounding healthy cells least affected by high linear energy transfer (LET) radiation like carbon ion beam. Anti-metastatic effect of carbon ion exposure attracts investigators into the field of hadron biology, although details remain poor. Poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors are well-known radiosensitizer and several PARP-1 inhibitors are in clinical trial. Our previous studies showed that PARP-1 depletion makes the cells more radiosensitive towards carbon ion than gamma. The purpose of the present study was to investigate combining effects of PARP-1 inhibition with carbon ion exposure to control metastatic properties in HeLa cells. METHODS: Activities of matrix metalloproteinases-2, 9 (MMP-2, MMP-9) were measured using the gelatin zymography after 85 MeV carbon ion exposure or gamma irradiation (0- 4 Gy) to compare metastatic potential between PARP-1 knock down (HsiI) and control cells (H-vector - HeLa transfected with vector without shRNA construct). Expression of MMP-2, MMP-9, tissue inhibitor of MMPs such as TIMP-1, TIMP-2 and TIMP-3 were checked by immunofluorescence and western blot. Cell death by trypan blue, apoptosis and autophagy induction were studied after carbon ion exposure in each cell-type. The data was analyzed using one way ANOVA and 2-tailed paired-samples T-test. RESULTS: PARP-1 silencing significantly reduced MMP-2 and MMP-9 activities and carbon ion exposure further diminished their activities to less than 3 % of control H-vector. On the contrary, gamma radiation enhanced both MMP-2 and MMP-9 activities in H-vector but not in HsiI cells. The expression of MMP-2 and MMP-9 in H-vector and HsiI showed different pattern after carbon ion exposure. All three TIMPs were increased in HsiI, whereas only TIMP-1 was up-regulated in H-vector after irradiation. Notably, the expressions of all TIMPs were significantly higher in HsiI than H-vector at 4 Gy. Apoptosis was the predominant mode of cell death and no autophagic death was observed. CONCLUSIONS: Our study demonstrates for the first time that PARP-1 inhibition in combination with carbon ion synergistically decreases MMPs activity along with overall increase of TIMPs. These data open up the possibilities of improvement of carbon ion therapy with PARP-1 inhibition to control highly metastatic cancers. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s13014-016-0703-x) contains supplementary material, which is available to authorized users. BioMed Central 2016-09-22 /pmc/articles/PMC5034624/ /pubmed/27659937 http://dx.doi.org/10.1186/s13014-016-0703-x Text en © The Author(s). 2016 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Ghorai, Atanu Sarma, Asitikantha Chowdhury, Priyanka Ghosh, Utpal PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells |
title | PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells |
title_full | PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells |
title_fullStr | PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells |
title_full_unstemmed | PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells |
title_short | PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells |
title_sort | parp-1 depletion in combination with carbon ion exposure significantly reduces mmps activity and overall increases timps expression in cultured hela cells |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5034624/ https://www.ncbi.nlm.nih.gov/pubmed/27659937 http://dx.doi.org/10.1186/s13014-016-0703-x |
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