Cold Physical Plasma Does Not Promote Metastasis-Like Spread In Human Pancreatic Cancer Cell Lines

Abstract

1. Purpose/Background

Up to 76% of pancreatic resections show histologically positive resection margins leaving patients with residual tumor load after surgery. Cold Physical Plasma (CPP) has shown antitumor effects inducing apoptosis. The application of CPP after resection of pancreatic cancer could significantly decrease the local microscopic tumor load. Yet, it is unknown whether plasmas may support tumor metastasis. Hence, the aim of this study was to characterize the influence of CPP on possible tumor cell spread. The cell adhesion profile of four different human pancreatic cancer cell lines was analyzed. In addition, the effects of CPP on Epithelial-to-Mesenchymal-Transition (EMT) of pancreatic cancer cells were investigated, especially the relation between ZEB-1 and E-Cadherin.

To complete the in-vitro studies, the chicken chorioallantoic membrane model was used as an in-vivo model. In relation to preliminary experiments, the cell adhesion molecules as well as the tumor cell spread were characterized for each cell line.

2. Methods

Using flow cytometry, the pancreatic cancer cell lines PA-TU-8988-T, PA-TU-8988-S, MIA-PaCa-2, PANC-1 were analyzed for CD324 (E-Cadherin), CD326 (EpCam), CD49b (Integrin α-2) and CD49d (Integrin α-4) as well as ZEB-1 after 60sec of CPP treatment. Furthermore, metabolic activity was investigated. All experiments were carried out in a 96-well plate in which adherent cells and supernatants (possibly containing floating cells) were observed. CPP treatment was performed employing the atmospheric pressure argon plasma jet kINPen 11. Examining the mechanic influence of the gas flow was done employing argon gas treatment without plasma.

3. Results/Conclusion

CPP reduced viability and metastasis potential of aggressive pancreatic carcinoma cell lines, whereas less aggressive cell lines were less affected. PATU-T showed significantly decreased viability after 60s of CPP treatment compared to controls (p= 0.0304), whereas PATU-S was less affected (p=0.3372). CPP-treatment increased the expression of E-Cadherin (p=0.0001) in PATU-T and MIA-PaCa-2 (p=0.0001) compared to untreated controls. This might affect potential metastasis in these cell lines whereas PATU-S and PANC-1 were not significantly affected.

Analyzing the number of viable cells after 4 days (as offspring of detached but viable floating tumor after plasma treatment) in all four cell lines, no changes between treated and non-treated cells were observed (p=0.8478).

These results confirm the antitumor effect of Cold Physical Plasma in pancreatic cancer. Furthermore, the results are promising, that CPP is a safe treatment option which may reduce local reoccurrence and moreover has no promoting effects on tumor cell spread.