Colorectal cancer is influenced by genetic mutations, lifestyle factors, and diet, particularly high fat intake, which raises bile acid levels in the intestinal lumen. This study hypothesized that bile acids contribute to tumorigenesis by disrupting ion transport and ATPase activity in the intestinal mucosa. The effects of 3-sulfo-taurolithocholic acid (TLC–S) on ATPase activity were investigated in colorectal cancer samples from 10 patients, using adjacent healthy tissue as controls, and in rodent liver function. ATPase activity was measured spectrophotometrically by determining inorganic phosphorus (P_i) in postmitochondrial fractions. Ca²⁺ dynamics were assessed in isolated mouse hepatocytes with fluorescence imaging, and rat liver mitochondria were studied using polarographic methods to evaluate respiration and oxidative phosphorylation. TLC–S increased Na⁺/K⁺ ATPase activity by 1.5 times in colorectal cancer samples compared to controls (p ≤ 0.05). In healthy mucosa, TLC–S decreased Mg²⁺ ATPase activity by 3.6 times (p ≤ 0.05), while Mg²⁺ ATPase activity in cancer tissue remained unchanged. TLC–S had no significant effect on Ca²⁺ ATPase activity in healthy colon mucosa but showed a trend toward decreased activity in cancer tissue. In rat liver, TLC–S decreased Ca²⁺ ATPase and Na⁺/K⁺ ATPase activities while increasing basal Mg²⁺ ATPase activity (p ≤ 0.05). Additionally, TLC–S induced cytosolic Ca²⁺ signals in mouse hepatocytes, partially attenuated by NED-19, an NAADP antagonist (p ≤ 0.05). TLC–S also reduced the V3 respiration rate of isolated rat liver mitochondria during α-ketoglutarate oxidation. These findings suggest that TLC–S modulates ATPase activity differently in cancerous and healthy colon tissues, playing a role in colorectal cancer development. In rat liver, TLC–S affects mitochondrial activity and ATPase function, contributing to altered cytosolic calcium levels, providing insight into the mechanistic effects of bile acids on colorectal cancer and liver function.

Bafilomycin A1 inhibits V-type H+ ATPases on the molecular level, which acidifies endolysosomes. The main objective of the study was to assess the effect of bafilomycin A1 on Ca2+content, NAADP-induced Ca2+ release, and ATPase activity in rat hepatocytes and human colon
cancer samples. Chlortetracycline (CTC) was used for a quantitative measure of stored calcium in permeabilized rat hepatocytes. ATPase activity was determined by orthophosphate content released after ATP hydrolysis in subcellular post-mitochondrial fraction obtained from rat liver as well as from patients’ samples of colon mucosa and colorectal cancer samples. In rat hepatocytes, bafilomycin A1 decreased stored Ca2+ and prevented the effect of NAADP on stored Ca2+. This effect was dependent on EGTA–Ca2+ buffers in the medium. Bafilomycin A1 significantly increased the activity of Ca2+ATPases of endoplasmic reticulum (EPR), but not plasma membrane (PM) Ca2+ ATPases in rat liver.
Bafilomycin A1 also prevented the effect of NAADP on these pumps. In addition, bafilomycin A1 reduced Na+/K+ ATPase activity and increased basal Mg2+ ATPase activity in the subcellular fraction of rat liver. Concomitant administration of bafilomycin A1 and NAADP enhanced these effects. Bafilomycin A1 increased the activity of the Ca2+ ATPase of EPR in the subcellular fraction of normal human colon mucosa and also in colon cancer tissue samples. In contrast, it decreased Ca2+ ATPase PM activity in samples of normal human colon mucosa and caused no changes in colon cancer. Bafilomycin A1 decreased Na+/K+ ATPase activity and increased basal Mg2+ ATPase activity
in normal colon mucosa samples and in human colon cancer samples. It can be concluded that bafilomycin A1 targets NAADP-sensitive acidic Ca2+ stores, effectively modulates ATPase activity, and assumes the link between acidic stores and EPR. Bafilomycin A1 may be useful for cancer therapy.
Keywords: molecular mechanisms; colon cancer; ATPase; autophagy; hepatocytes; liver; NAADP; biomarkers; bafilomycin A1; Ca2+ store