Abstract
Purpose
In normal physiology, a vacuolar-type proton pump (V-ATPase) maintains an intracellular acid microenvironment in lysosome, endosome, and other endomembrane systems. Cancer cells overexpress V-ATPase compared with normal cells, and disturbances of the acid environment are thought to significantly impact the cancer cell infiltration and growth. Bafilomycin A1 (Baf-A1) is a specific inhibitor of the proton-pump inhibitor (PPI) V-ATPase. Neoplastic cells are reportedly more sensitive to Baf-A1 than normal cells, and the difference between the susceptibility to Baf-A1 in normal cells and that in cancer cells may become a target in the cancer therapy. With this in mind, we used cells of hepatoblastoma, the cancer type accounting for 80% of all childhood liver cancers, to investigate the effects of Baf-A1 as an inducer of cancer cell apoptosis and inhibitor of cancer cell reproduction
Methods and results
Electron microscopy showed significant morphological change of the hepatoblastoma cells of the Baf-A1-treated group compared with hepatoblastoma cells of the Baf-A1-free group. The rate of the apoptotic cell increased, and cell reproduction was inhibited. Moreover, the analysis of hepatoblastoma cells using the gene Chip gene expression analysis arrays showed that three of the 27 V-ATPase-related transcripts (ATP6V0D2, ATP6V1B1, and ATP6V0A1) were more weakly expressed in the Baf-A1-treated cells than in the Baf-A1-free cells. In normal human hepatic cells, on the other hand, the inhibition of cell growth of the Baf-A1-treated cells was negligible compared to that of the cells without Baf-A1 treatment. The result of apoptotic cell detection by morphological observations and flow cytometry revealed that Baf-A1 inhibits hepatoblastoma cellular reproduction by inducing apoptosis. On the other hand, the Baf-A1-conferred inhibition of cell growth was negligible in normal human hepatocytes
Conclusion
The V-ATPase inhibitor Baf-A1 has been proven to selectively inhibit the reproduction and induce the apoptosis of hepatoblastoma cells without adversely influencing normal hepatic cells. With these effects, V-ATPase inhibitors may hold promise as therapeutic agents for hepatoblastoma. Given that three V-ATPase-related genes (ATP6V0D2, ATP6V1B1, and ATP6V0A1) were more weakly expressed in the hepatoblastoma cells of the Baf-A1-treated group than in the Baf-A1-free cells, drug development targeting V-ATPase gene of hepatoblastomas is expected.
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References
Nelson N et al (1989) Structure, molecular genetics, and evolution of vacuolar H+-ATPases. J Bioenerg Biomembr 21:553–571
Ohta T, Numata M, Yagishita H et al (1996) Expression of 16 kDa proteolipid of vacuolar-type H+-ATPase in human pancreatic cancer. Br J Cancer 73:1511–1517
Ohkuma S, Shimizu S, Noto M et al (1993) Inhibition of cell growth by bafilomycin A1, a selective inhibitor of vacuolar H+-ATPase. In vitro Cell Dev Biol Anim 29A:862–866
Manabe T, Yoshimori T, Henomatsu N et al (1993) Inhibitors of vacuolar-type H+-ATPase suppresses proliferation of cultured cells. J Cell Physiol 157:445–452
Ohkuma S, Poole B (1978) Fluorescence probe measurement of the intralysosomal pH in living cells and perturbation of pH by various agents. Proc Natl Acad Sci USA 75:3327–3331
Nishihara T, Akifusa S, Koseki T et al (1995) Specific inhibitors of vacuolar type H+-ATPase induce apoptotic cell death. Biochem Biophys Res Commun 212:255–262
Bunge RP, Wood P (1973) Studies on the transplantation of spinal cord tissue in the rat. I. The development of a culture system for hemisections of embryonic spinal cord. Brain Res 57:261–276
Perilongo G, Shafford E, Plaschkes J (2000) SIOPEL trials using preoperative chemotherapy in hepatoblastoma. Lancet Oncol 1:94–100
Hurtado-Lorenzo A, Skinner M, Annan JEI et al (2006) V-ATPase interacts with ARNO and Arf6 in early endosomes and regulates the protein degradative pathway. Nat Cell Biol 8:124–136
Martinez-Zaguilan R, Lynch RM, Martinez GM et al (1993) Vacuolar-type H+-ATPases are functionally expressed in plasma membranes of human tumor cells. Am J Physiol 265:1015–1029
Finbow ME, Harrison MA (1997) The vacuolar H+-ATPase: a universal proton pump of eukaryotes. Biochem J 324:697–712
Shrode LD, Tapper H, Gristein S (1997) Role of intracellular pH in proliferation, transformation, and apoptosis. Bioenerg Biomembr 29:393–399
Hinton A, Bond S, Forgac M (2007) V-ATPase functions in normal and disease processes. Pflugers Arch. doi: 10.1007/s00424-007-0382-4
Nelson N, Harvey WR et al (1999) Vacuolar and plasma membrane proton adenosinetriphoshatases. Physiol Rev 79:361–385
Yoshimoto Y, Imoto M (2002) Induction of EGF-dependent apoptosis by vacuolar type H+-ATPase inhibitors in A431 cells overexpressing the EGF receptor. Exp Cell Res 279:118–127
Hamada H, Moriyama Y, Maeda N et al (1990) Kinetic studies of chromaffin granule H+-ATPase and effects of bafilomycin A1. Biochem Biophys Res Commun 170:873–878
Huss M, Sasse F, Kunze B, Kunze B et al (2005) Archazolid and apicularen: novel specific V-ATPase inhibitors. BMC Biochem 6:13
Niikura K, Takeshita M, Takano M et al (2005) A vacuolar ATPase inhibitor, FR 167356, prevents bone resorption in ovariectomized rats with high potency and specificity: potential for clinical application. J Bone Miner Res 20:1579–1588
Ohta T, Arakawa H, Futagami F et al (1996) A new strategy for the therapy of pancreatic cancer by proton pump inhibitor. Jpn J Cancer Chemother 23:1660–1664
Nakashima S, Hiraku Y, Oikawa ST (2003) Vacuolar H+-ATPase inhibitor induces apoptosis via lysosomal dysfunction in the human gastric cancer cell line MKN-1. J Biochem 134:359–364
Sennoune SR, Bakunts K, Martinez GM et al (2004) Vacuolar H+-ATPase in human breast cancer cells with distinct metastatic potential: distribution and functional activity. Am J Physiol Cell Physiol 286:1443–1452
Nishi T, Forgac M (2002) The vacuolar H+-ATPases—nature’s most versatile proton pumps. Nat Rev Mol Cell Biol 3:94–103
Stevens TH, Forgac M (1997) Structure, function and regulation of the vacuolar H+-ATPase. Annu Rev Cell Dev Biol 13:779–808
Pali T, Whyteside G, Dixon N et al (2004) Interaction of inhibitors of the vacuolar H+-ATPase with the transmembrane Vo-sector. Biochemistry 43:12297–12305
Bowman EJ, Bowman BJ (2005) V-ATPases as drug targets. J Bioenerg Biomembr 37:431–435
Whyteside G, Meek PJ, Ball SK et al (2005) Concanamycin and indolyl pentadieneamide inhibitors of the vacuolar H+-ATPase bind with high affinity to the purified proteolipid subunit of the membrane domain. Biochemistry 44:15024–15031
Otero-Rey EM, Somoza-Martin M, Barros-Angueira F et al (2008) Intercellular pH regulation in oral squamous cell carcinoma is mediated by increased V-ATPase activity via over expression of the ATP6V1C1 gene. Oral Oncol 44:193–199
Lu X, Qin W, Li J et al (2005) The growth and metastasis of human hepatocellular carcinoma xenografts are inhibited by small interfering RNA targeting to the subunit ATP6L of proton pump. Cancer Res 65:6843–6849
Hinoki A, Yoshimura K, Fujita K et al (2006) Suppression of proinflammatory cytokine production in macrophages by lansoprazol. Pediatr Surg Int 22:915–923
Acknowledgments
We thank Ms. Kumiko Komatsu and Ms. Sachiko Matsumoto from the Division of Morphological Science at Saitama Medical University for their collaborations in this research.
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Morimura, T., Fujita, K., Akita, M. et al. The proton pump inhibitor inhibits cell growth and induces apoptosis in human hepatoblastoma. Pediatr Surg Int 24, 1087–1094 (2008). https://doi.org/10.1007/s00383-008-2229-2
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DOI: https://doi.org/10.1007/s00383-008-2229-2