Tissue tolerable plasma (TTP) induces apoptosis in pancreatic cancer cells in vitro and in vivo.
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.
Department of General, Visceral, Thoracic and Vascular Surgery, University Medicine Greifswald, Ernst-Moritz-Arndt University, Greifswald, Germany. partecke@uni-greifswald.de
Lars Ivo
LI
Partecke
Katja
K
Evert
Jan
J
Haugk
Friderike
F
Doering
Lars
L
Normann
Stephan
S
Diedrich
Frank-Ulrich
FU
Weiss
Matthias
M
Evert
Nils Olaf
NO
Huebner
Cristin
C
Guenther
Claus Dieter
CD
Heidecke
Axel
A
Kramer
René
R
Bussiahn
Klaus-Dieter
KD
Weltmann
Onur
O
Pati
Claudia
C
Bender
Wolfram
W
von Bernstorff
D000230:Adenocarcinoma / Q000188:drug therapy*; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D017679:Cryotherapy / Q000379:methods; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D008954:Models, Biological; D018365:Neoplasm, Residual; D010190:Pancreatic Neoplasms / Q000188:drug therapy*; D058626:Plasma Gases / Q000494:pharmacology*
D000970:Antineoplastic Agents; D058626:Plasma Gases