## 3 Results
### Examinations on Tissue Specimen  
Induction of apoptosis by CAP treatment was superior in tumour tissue compared with healthy mucosal tissue. Following CAP triggered cytotoxicity and induction of early apoptotic events in cultured cells we next extended this view to biopsies of human non-malignant mucosa and squamous cell carcinoma of the head and neck area and investigated differences between these tissues after CAP treatment (3 min). The number of apoptotic cells was evaluated in situ in the tumour tissue and in healthy mucosal tissue of the same patient. In healthy tissue, DNA fragmentation after CAP exposure indicative for apoptosis was marginal. Low numbers of TUNEL-positive cells were counted, namely 2.58% versus 0.79% at median in untreated samples and CAP treated tissue specimens, respectively (Figure 5). In contrast, the tissue samples of head and neck squamous cell carcinoma displayed a significantly elevated median number of apoptotic cells of 5.6% compared to untreated tumour tissue (1.4%). In this context, it is noteworthy to mention that individual responsiveness varies (Figure 5). To strengthen the finding of induced apoptosis by CAP in tissue samples, the supernatant medium was analysed for cytochrome c and selected cytokines. As a marker for cell death, cytochrome c is released by mitochondria upon induction of apoptosis. Initially, the tumour tissue even without CAP treatment revealed a much higher amount of cytochrome c than the corresponding healthy tissue (1729 ng per mg protein versus 385 ng per mg protein) (Figure 6). However, the CAP treatment triggered apoptotic pathways and the cytochrome c concentration raised significantly to as high as 5155 ng per mg protein in the supernatant of the tumour tissue (three-fold increase). Strikingly this was not the case for the corresponding healthy tissue. Here the same CAP treatment regimen led to a mean cytochrome c content of 698 ng per mg protein (1,8-fold increase) (Figure 6). \begin{table}
\begin{tabular}{|c c c c|c|} \hline \hline
**Protein Name** & **Uniprot ID** & **HaCaT** & **HNO97** & **Function** \\
**Ki67** & **P46013** & & & \(\rightarrow\uparrow\) & Proliferation \\ \hline
**NOO1** & **P1559** & \(\rightarrow\uparrow\) & & Nuclear factor erythroid 2-related factor 2 (NRF2) \\  & & & & related signalling \\ \hline
**TMX2** & **Q9Y320** & \(\uparrow\) & & NRF2 related signalling \\ \hline
**GSTM3** & **P21266** & \(\uparrow\) & & NRF2 related signalling \\ \hline
**PRDX12,24.6** & **Q06830** & & & Redox regulation, elimination of peroxides \\ \hline
**S8 NO2** & **Q9Y2G9** & \(\uparrow\uparrow\) & \(\uparrow\) & Transcriptional co-regulation, interleukin signalling \\
**IIKAP** & **Q9H0C8** & \(\uparrow\uparrow\) & \(\downarrow\) & Cell adhesion and growth factor signalling \\
**AAMP** & **Q13685** & \(\rightarrow\uparrow\) & \(\downarrow\) & Cell migration, angiogenesis \\
**ROCK2** & **Q75116** & \(\rightarrow\uparrow\) & \(\downarrow\) & Regulation of actin cytoskeleton, cell adhesion and motility \\
**CTTB2** & **Q8v**_/_274_ & \(\uparrow\uparrow\) & \(\rightarrow\uparrow\) & Cytoskeleton rearrangement, cell migration and motility \\
**CFA20** & **Q9Y6A4** & \(\uparrow\) & & Cell motility \\
**SRGP2** & **O75044** & \(\rightarrow\) & \(\uparrow\) & Cell migration inhibition, actin dynamics \\
**ELMO3** & **Q96 BJ8** & & \(\downarrow\) & Cell motility \\
**CASP8/10** & **Q14790**/**Q92851** & & \(\uparrow\) & Pro-apoptotic caspases \\ \hline \hline \end{tabular}
\end{table} 
Table 1: Expression of selected proteins in normal and malignant cells 3 h and 24 h after CAP treatment. Protein changes are indicated as: up to two-fold increased \(\rightarrow\uparrow\); two-fold increased \(\uparrow\); 5 to 12-fold increased \(\uparrow\uparrow\); two-fold decreased \(\downarrow\downarrow\); 5-12-fold decreased \(\downarrow\downarrow\); \(\blacksquare\) not changed. Figure 5: Induction of apoptotic cells by CAP application in tumour tissue in situ.