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Open Access Highly Accessed Research article

Tissue Tolerable Plasma (TTP) induces apoptosis in pancreatic cancer cells in vitro and in vivo

Lars Ivo Partecke1*, Katja Evert2, Jan Haugk1, Friderike Doering1, Lars Normann1, Stephan Diedrich1, Frank-Ulrich Weiss3, Matthias Evert2, Nils Olaf Huebner4, Cristin Guenther1, Claus Dieter Heidecke1, Axel Kramer4, René Bussiahn5, Klaus-Dieter Weltmann5, Onur Pati1, Claudia Bender4 and Wolfram von Bernstorff1

Author Affiliations

1 Department of General, Visceral, Thoracic and Vascular Surgery, University Medicine Greifswald, Ernst-Moritz-Arndt University, Greifswald, Germany

2 Institute of Pathology, University Medicine Greifswald, Ernst-Moritz-Arndt University, Greifswald, Germany

3 Department of Gastroenterology and Nephrology, University Medicine Greifswald, Ernst-Moritz-Arndt University, Greifswald, Germany

4 Institute of Hygiene and Environmental Medicine, University Medicine Greifswald, Ernst-Moritz-Arndt University, Greifswald, Germany

5 Leibniz Institute of Plasma Science and Technology (INP), Felix Hausdorff-Str. 2, Greifswald, Germany

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BMC Cancer 2012, 12:473  doi:10.1186/1471-2407-12-473

Published: 15 October 2012

Abstract

Background

The rate of microscopic incomplete resections of gastrointestinal cancers including pancreatic cancer has not changed considerably over the past years. Future intra-operative applications of tissue tolerable plasmas (TTP) could help to address this problem. Plasma is generated by feeding energy, like electrical discharges, to gases. The development of non-thermal atmospheric plasmas displaying spectra of temperature within or just above physiological ranges allows biological or medical applications of plasmas.

Methods

We have investigated the effects of tissue tolerable plasmas (TTP) on the human pancreatic cancer cell line Colo-357 and PaTu8988T and the murine cell line 6606PDA in vitro (Annexin-V-FITC/DAPI-Assay and propidium iodide DNA staining assay) as well as in the in vivo tumour chorio-allantoic membrane (TUM-CAM) assay using Colo-357.

Results

TTP of 20 seconds (s) induced a mild elevation of an experimental surface temperature of 23.7 degree Celsius up to 26.63+/−0.40 degree Celsius. In vitro TTP significantly (p=0.0003) decreased cell viability showing the strongest effects after 20s TTP. Also, TTP effects increased over time levelling off after 72 hours (30.1+/−4.4% of dead cells (untreated control) versus 78.0+/−9.6% (20s TTP)). However, analyzing these cells for apoptosis 10s TTP revealed the largest proportion of apoptotic cells (34.8+/−7.2%, p=0.0009 versus 12.3+/−6.6%, 20s TTP) suggesting non-apoptotic cell death in the majority of cells after 20s TTP. Using solid Colo-357 tumours in the TUM-CAM model TUNEL-staining showed TTP-induced apoptosis up to a depth of tissue penetration (DETiP) of 48.8+/−12.3μm (20s TTP, p<0.0001). This was mirrored by a significant (p<0.0001) reduction of Ki-67+ proliferating cells (80.9+/−13.2% versus 37.7+/−14.6%, p<0.0001) in the top cell layers as well as typical changes on HE specimens. The bottom cell layers were not affected by TTP.

Conclusions

Our data suggest possible future intra-operative applications of TTP to reduce microscopic residual disease in pancreatic cancer resections. Further promising applications include other malignancies (central liver/lung tumours) as well as synergistic effects combining TTP with chemotherapies. Yet, adaptations of plasma sources as well as of the composition of effective components of TTP are required to optimize their synergistic apoptotic actions.

Keywords:
Tissue tolerable plasma (TTP); Plasma medicine; Apoptosis; TUM-CAM Assay; Pancreatic cancer