## 3 Results  
### Effect of Simulated Acidosis on Cell Proliferation Activity  
Figure 1 A, B demonstrate the \"behavior\" of cancer cells cultured on a medium with different pH values (from 6.0 to 8.0) achieved by lactic acid (1%) and hydrochloric acid (0.1 mol/L), respectively. The results shown in Figure 1A indicate an abrupt increase in the viability and proliferative activity of colon cancer cells after 48 h of incubation in the pH-corrected cell medium (from pH = 6.6 to pH = 7.9) adjusted with lactic acid as compared with the control group cells (pH = 8.0). An exception to the general tendency was noticed at pH = 6.0, where the proliferative activity was lower than in the control cells incubated in the normal medium. A decrease in cancer activity was observed after 72 h of incubation, as compared with incubation for 48 h, but these results are related to over-crowding of the cell mass due to the rapid cell proliferation. A deviation from this behavior was observed at pH = 6.0, where the proliferative activity of the colon cancer cells after 48 h of incubation was lower than that at 72 h. There was no change in the proliferative activity and viability after 24 h of incubation of cancer cells exposed to the medium containing lactic acid (from pH = 6.0 to pH = 7.9) compared to the control group (pH = 8.0), where a slight elevation in the proliferative activity was observed.  
Figure 1: Cancer cell proliferation activity (Colon 26) after correction of pH values of the culture medium by 1% lactic acid (**A**) and by 0.1 mol/L hydrochloric acid (**B**) after 24, 48, and 72 h of cultivation. The levels of statistical significance (**_p_ < 0.001) in the individual group experimental series (n = 3) with lactic acid (group 1) or HCl (group 2) for 48 and for 72 h were determined. The level of statistical significance (* \(p\) < 0.05) between two periods of incubation (48 h and 72 h) with HCl was also determined.  
A tendency for increased cell division and viability of colon cancer cells was also demonstrated after incubation with the medium containing 0.1 mol/L hydrochloric acid (Figure 1B). Generally, the obtained results in the medium with HCl (Figure 1B) are lower than those noticed in the medium containing lactic acid (Figure 1A). According to Figure 1B, the acceleration in cell proliferative activity occurred after 48 h of incubation in all tested pH-corrected cell medium ranges (from pH = 6.0 to pH = 7.9) compared with the absence of such activity under the same conditions but after 24 h of treatment. Compared to the results shown in Figure 1A, the cancer cells exposed to HCl did not show dramatic variations in cell proliferative activity and viability after 48 and 72 h of incubation, but a slight suppression of proliferative activity was noticed at pH = 7.0, pH = 7.8, and pH = 7.9. A special case was observed at pH=7.4, where a suppression of cell proliferative activity was established after 48 and 72 h compared with other tested conditions.  
The data in Figure 2 A, B show the viability of colon cancer cells after incubation in the pH-corrected cell medium adjusted with lactic acid and hydrochloric acid, respectively. The obtained results are similar to those observed in the previous analysis.  
The characteristic sign of the viability of epithelial colon cancer cells is the formation of colonies by adhesion-molecule-mediated cell forces. A surprising tendency for elevated survival, but in a cell suspension, was observed for epithelial colon cancer cells exposed to the pH-corrected cell medium (under acidosis conditions) after 48 and 72 h of incubation. This finding was registered during the microscopic study (Figure 3), in which the cell suspension was formed from live cells, and a dramatic change in the shape of cells was observed (round cells). Such a morphological change, provoked by extracellular signals, is indicative of an increased tendency for metastasis formation and could be a possible reason for the release of cancer cells into the lymphatic system or blood stream.