Monovalent ions and stress-induced senescence in human mesenchymal endometrial stem/stromal cells

Abstract

Monovalent ions are involved in growth, proliferation, differentiation of cells as well as in their death. This work concerns the ion homeostasis during senescence induction in human mesenchymal endometrium stem/stromal cells (hMESCs): hMESCs subjected to oxidative stress (sublethal pulse of H2O2) enter the premature senescence accompanied by persistent DNA damage, irreversible cell cycle arrest, increased expression of the cell cycle inhibitors (p53, p21) cell hypertrophy, enhanced β-galactosidase activity. Using flame photometry to estimate K+, Na+ content and Rb+ (K+) fluxes we found that during the senescence development in stress-induced hMESCs, Na+/K+pump-mediated K+ fluxes are enhanced due to the increased Na+ content in senescent cells, while ouabain-resistant K+ fluxes remain unchanged. Senescence progression is accompanied by a peculiar decrease in the K+ content in cells from 800–900 to 500–600 µmol/g. Since cardiac glycosides are offered as selective agents for eliminating senescent cells, we investigated the effect of ouabain on ion homeostasis and viability of hMESCs and found that in both proliferating and senescent hMESCs, ouabain (1 nM–1 µM) inhibited pump-mediated K+ transport (ID50 5 × 10–8 M), decreased cell K+/Na+ ratio to 0.1–0.2, however did not induce apoptosis. Comparison of the effect of ouabain on hMESCs with the literature data on the selective cytotoxic effect of cardiac glycosides on senescent or cancer cells suggests the ion pump blockade and intracellular K+ depletion should be synergized with target apoptotic signal to induce the cell death.

The Role of Intracellular Potassium in Cell Quiescence, Proliferation, and Death

“…Later, during senescence progression, the Na + content is slightly increased and the pumpmediated K + transport is enhanced. The higher K + influxes in "late" stressed senescent eMSCs are not associated with modulation of the Na/K pump properties or an increased number of ion pumps but are due to higher cell Na + content in late senescence [25]. Stressinduced proliferation arrest does not affect the ouabain-resistant K + fluxes associated with the activity of ion channels and ion co-transporters.…”

“…Fluorescent probes have revealed an increased content of both K + and Na + in the senescent human lung fibroblast IMR90 [40]. However, thorough estimations of ions in cells with the flame photometry technique have shown that stressinduced senescent human eMSCs have low Na + and maintain a high K + /Na + ratio typical for functionally active animal cells [25]. Indeed, short oxidative stress leads to a decrease in intracellular K + content and an increase in Na + content in eMSCs; however, within a day, in "early" stress-arrested cells, the ionic gradients and ion pump activity are restored (Figure 2).…”

“…During long-term culture, senescent cells maintain the high K + /Na + ratio typical for functionally active animal cells. The most significant observation regarding ions during the development of stressinduced senescence in eMSCs concerns the intracellular K + content [25]. The senescence progression is accompanied by a gradual decrease in the intracellular K + from 0.7 mmol/g protein ("early" senescent eMSCs) to 0.5-0.6 mmol/g protein ("late" senescent eMSCs) (Figure 2A).…”

“…Grey columns represent cellular content of K + (A) and Na + (B) after treatment of cells with 200 µM H 2 O 2 .For the first 8 days, data are shown as means ± SD for six independent experiments performed in triplicate; significant difference between stress-induced and control proliferating cells (Ctr) was calculated using one-way ANOVA with Tukey's post hoc tests, * p < 0.05. For late senescent cells(18-22 days), data are shown as means ± SD (n = 3) according to[25].…”

Intracellular Potassium in Cell Growth and Proliferation of Human Mesenchymal Stem Cells and Blood Lymphocytes

“…Short treatment with 200 μM H 2 O 2 of subconfluent eMSCs cultures has been shown to induce senescence [31]. eMSCs subjected to sublethal oxidative stress enter into irreversible cell cycle arrest and exhibit a senescent phenotype including p53/p21/Rb mediated cell cycle arrest, cell hypertrophy, enhanced β-galactosidase staining [31,32].…”