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Inhibition of ecto-ATPase activity by curcumin in hepatocellular carcinoma HepG2 cells
Takuto Fujii1, Takuma Minagawa, Takahiro Shimizu
1Department of Pharmaceutical Physiology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Japan.
Abstract:
Effects of curcumin, a major constituent of turmeric, on ecto-nucleotidases have not been clarified. Here, we investigated whether curcumin affects ecto-nucleotidase activities in human hepatocellular carcinoma HepG2 cells. In the cells, high levels of Mg(2+)-dependent activity of ecto-nucleotidases were observed in the presence of 1 mM adenosine triphosphate (ATP). The activity was inhibited by ecto-ATPase inhibitors such as suramin, ZnCl(2) and 4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid. On the other hand, the activity was significantly decreased at alkaline pH (pH 9) and was not inhibited by levamisole, an inhibitor of alkaline phosphatase. In the presence of ATP, curcumin inhibited the activity in a concentration-dependent manner (IC(50) = 6.2 μM). In contrast, curcumin had no effects on ecto-nucleotidase activity in the presence of ADP (1 mM) or AMP (1 mM). The K (m) value for ATP hydrolysis of curcumin-sensitive ecto-ATPase was similar to the value of NTPDase2, an isoform of ecto-nucleoside triphosphate diphosphohydrolase. These results suggest that curcumin is a potent inhibitor of ecto-ATPase and may affect extracellular ATP-dependent responses.
Insights
Curcumin, a compound in turmeric, potently inhibits ecto-ATPase activity in liver cancer cells. This suggests curcumin may influence cellular responses mediated by extracellular adenosine triphosphate (ATP).
Area of Science:
- Biochemistry
- Pharmacology
- Cell Biology
Background:
- Ecto-nucleotidases play crucial roles in cellular signaling by hydrolyzing extracellular nucleotides.
- The effects of curcumin, a turmeric-derived compound, on ecto-nucleotidase activity remain largely uncharacterized.
- Human hepatocellular carcinoma HepG2 cells express ecto-nucleotidase activity.
Purpose of the Study:
- To investigate the impact of curcumin on ecto-nucleotidase activities in HepG2 cells.
- To determine if curcumin specifically inhibits ecto-ATPase, ecto-ADPase, or ecto-5'-nucleotidase.
- To characterize the inhibitory effects of curcumin on ATP hydrolysis.
Main Methods:
- Enzyme activity assays were performed on HepG2 cell lysates.
- Adenosine triphosphate (ATP), adenosine diphosphate (ADP), and adenosine monophosphate (AMP) were used as substrates.
- Inhibition kinetics were analyzed using known ecto-nucleotidase inhibitors and varying concentrations of curcumin.
- The Michaelis constant (Km) for ATP hydrolysis was determined.
Main Results:
- HepG2 cells exhibited significant Mg(2+)-dependent ecto-nucleotidase activity with ATP as a substrate.
- Curcumin inhibited this ATP hydrolysis in a concentration-dependent manner, with an IC50 of 6.2 μM.
- Curcumin did not affect ecto-nucleotidase activity when ADP or AMP were used as substrates.
- The kinetic parameters for ATP hydrolysis were consistent with the NTPDase2 isoform.
Conclusions:
- Curcumin is a potent inhibitor of ecto-ATPase activity in human hepatocellular carcinoma HepG2 cells.
- Curcumin's inhibitory action is specific to ATP hydrolysis, suggesting a targeted effect on ecto-ATPases.
- These findings indicate that curcumin may modulate extracellular ATP-dependent cellular processes and signaling pathways.
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