Low Doses of Ouabain Protect from Serum Deprivation–Triggered Apoptosis and Stimulate Kidney Cell Proliferation via Activation of NF-κB

Abstract

It now generally is agreed that Na,K-ATPase, in addition to its role in the maintenance of Na ؉ and K ؉ gradients across the cell membrane, plays a role in communicating information from the extracellular environment to intracellular signaling pathways. It was reported recently that interaction between ouabain-bound Na,K-ATPase and the 1,4,5-trisphosphate receptor (IP 3 R) triggers slow calcium oscillations and activation of NF-B. Here it is demonstrated that this signaling pathway can serve to prevent cell death and promote cell growth. Rat renal proximal tubular cells in primary culture first were grown in the presence of 10% serum and then exposed to 0.2% serum for 24 h to induce apoptosis. Serum starvation increased the apoptotic index from 1.21 ؎ 0.26 to 14.01 ؎ 1.17%. Ouabain in concentrations that did not inhibit Na,K-ATPase activity (1 to 10 nM) completely abolished the apoptotic effect of serum starvation. Ouabain protection from apoptosis was not observed when release of calcium from intracellular stores via the IP 3 R was prevented. It was shown that the NH 2 terminal tail of the Na,K-ATPase ␣ subunit plays a key role in ouabain-triggered calcium oscillations. It was found that ouabain did not protect from apoptosis in serum-deprived cells that expressed a mutant Na,K-ATPase ␣ subunit with deletion of the NH 2 terminal tail. . Recent studies suggest that Na,K-ATPase, in addition to being the major determinant of intracellular ion composition, may act as a signal transducer (2-5).Ouabain, a steroid derivative, is a specific ligand of Na,KATPase that dose-dependently inhibits the activity of Na,KATPase (6). Noninhibitory doses of ouabain activate the signaling function of Na,K-ATPase. The signaling cascade that is triggered by Na,K-ATPase is complex, and several different pathways have been implicated (7-9). We recently reported that ouabain can trigger an interaction between Na,K-ATPase and the inositol 1,4,5-trisphosphate (IP 3 ) receptor, which results in low-frequency Ca 2ϩ oscillations and activation of the transcription factor NF-B. This phenomenon was observed in rat renal proximal tubular cells in primary culture and in a kidney cell line (10).A number of recent studies have demonstrated that ouabain in noninhibitory doses can promote cell proliferation (11-13). Because NF-B has an antiapoptotic effect (14), we speculated that ouabain also might act to protect from apoptosis. Here we demonstrate that noninhibitory doses of ouabain can protect rat renal proximal tubular cells from serum deprivation-triggered apoptosis, albeit not from pharmacologically staurosporinetriggered apoptosis. We show that protection from serum-triggered apoptosis depends on NF-B activation. Normal kidney development is critically dependent on a well-controlled balance between cell proliferation and apoptosis (15). Accumulating evidence suggests that ouabain is a mammalian hormone, produced in the adrenal glands and the hypothalamus (16,17). Therefore, ouabain might play an important role as a modulator of kidney growth ...

Ouabain Binds with High Affinity to the Na,K-ATPase in Human Polycystic Kidney Cells and Induces Extracellular Signal–Regulated Kinase Activation and Cell Proliferation

“…The similar origin of the cells suggests that the changes in ouabain response that were observed are a consequence of the cellular phenotype of normal renal epithelial cells versus mural cystic cells and not due to differences in the progenitor cell type. The proliferative effects of ouabain have been described in various cell types, including normal proximal tubule cells of the rat kidney (14,38). Although we could not detect statistically significant differences in the NHK cells, we observed a trend for an increase in ouabain-dependent proliferation in these cells.…”

Chemical biology of inflammatory cytokine signaling

“…105 It has been shown that ouabain signals calcium oscillations through the inositol 1,4,5-triphosphate receptor that interacts with Na + /K + -ATPase and thereby induces NF-kB activation at concentrations that do not inhibit Na + /K + -ATPase activity. [107][108][109] In cardiac myocytes, ouabain has been reported to promote the rapid generation of reactive oxygen species, thereby inducing NF-kB activation. 110 As an early signaling pathway, Na + /K + -ATPase, on binding to cardiac glycosides, activates the cytoplasmic tyrosine kinase SRC, thereby activating the epidermal growth factor receptor and leading to the recruitment of adaptor proteins and the activation of the MAP kinase cascade that ultimately stimulates the mitochondrial production of reactive oxygen species.…”

2011 Homer Smith Award

“…59 We have found in studies performed on primary cultures of rat proximal tubular cells and on COS cells (a cell line derived from embryonic monkey kidney) that treatment with ouabain in concentrations that spare the pump function will result in translocation of NF-kB from the cytoplasm to the nucleus and reduced expression of I-kB, the cytosolic partner of NF-kB. 53,60 NF-kB consists of five subunits and is a pleiotropic transcription factor. 61 Dependent on the mode of activation, it may trigger transcription of the antiapoptotic factor Bcl-xL or the apoptotic factor apoptotic factor Bax.…”

“…61 Dependent on the mode of activation, it may trigger transcription of the antiapoptotic factor Bcl-xL or the apoptotic factor apoptotic factor Bax. 62 Ouabain-triggered calcium oscillations have, in studies on rat renal cells, been found to have an antiapoptotic effect 53,60 and to increase the expression of Bcl-xL.…”

“…60,63,64 These effects depend on ouabain/Na + , K + -ATPase signaling by both Src and IP3R pathways. Na + ,K + -ATPase stimulation of cell proliferation is a protective mechanism during adverse developmental conditions and in renal disease, but it might also have pathologic implications.…”