PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cellsReport as inadecuate

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Radiation Oncology

, 11:126

Radiation Biology and Molecular Radiation Oncology


BackgroundHadron 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. PolyADP-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.

MethodsActivities 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.

ResultsPARP-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.

ConclusionsOur 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.

KeywordsCarbon ion exposure Gamma radiation PARP-1 Matrix metalloproteinases MMPs Tissue inhibitor of matrix metalloproteinases TIMPs Cell death Apoptosis AbbreviationsDAPI4’, 6-diamidino-2-phenylindole

FITCFluorescein isothiocyanate


LETLinear energy transfer

MMPsMatrix metalloproteinases

PARP-1PolyADP-ribose polymerase-1

SDStandard deviation

TIMPsTissue inhibitor of matrix metalloproteinases

Electronic supplementary materialThe online version of this article doi:10.1186-s13014-016-0703-x contains supplementary material, which is available to authorized users.

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Author: Atanu Ghorai - Asitikantha Sarma - Priyanka Chowdhury - Utpal Ghosh


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