Restoration of responsiveness to phorbol ester by reconstitution of a functional Na/K/Cl cotransporter in cotransporter-deficient BALB/c 3T3 cells

Guo, Yongjun; O’Brien, Thomas G.

doi:10.1002/(sici)1098-2744(199609)17:1<35::aid-mc5>3.0.co;2-j

Cited by 2 publications

(1 citation statement)

References 27 publications

(27 reference statements)

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“…Bumetanidesensitive 86 Rb uptake increased in actively proliferating cells compared with growth-suppressed serum-starved cells and is consistent with results obtained previously in human skin fibroblasts (20). Moreover, Guo and O'Brien (7) showed that the mouse BALB/c 3T3 fibroblast cell line deficient in NKCC1 is not responsive to a phorbol ester mitogen, but that transfection with a shark NKCC1 gene restores responsiveness. Together, this suggests that stimulation of the NKCC cotransporter may be an essential part of the mitogenic signal.…”

Section: Discussionsupporting

confidence: 88%

Na-K-2Cl cotransporter inhibition impairs human lung cellular proliferation

Iwamoto¹,

Fujiwara²,

Nakamura³

et al. 2004

American Journal of Physiology-Lung Cellular and Molecular Phys

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The widespread presence of the Na-K-2Cl (NKCC) cotransporter protein suggests that chronic administration of inhibitors may result in adverse effects. Inhibition of the NKCC cotransporter by loop diuretics is felt to underlie the diuretic and the pulmonary smooth muscle relaxant effects of this drug class. However, the fundamental regulation of salt and water movement by this cotransporter suggests that it may also mediate cell volume changes occurring during cell cycle progression. Thus we hypothesized that NKCC cotransporter inhibition by loop diuretics would decrease cellular proliferation. Normal human bronchial smooth muscle cells (BSMC) showed a significant concentration-dependent decrease in cell counts after 7 days of exposure to both bumetanide (n=5-10) and furosemide (n=6-16) compared with controls. Proliferation was similarly inhibited in normal human lung fibroblasts (n=5-9). To determine whether this was due to loss of cells, we performed apoptosis assays on BSMC. Both annexin V-propidium iodide staining (n=5-10) and single cell gel electrophoresis assays (n=4) were negative for necrosis and apoptosis in BSMC exposed to 10 microM bumetanide. Subsequent analysis of the cell cycle by flow cytometry showed that bumetanide-exposed BSMC were delayed in G1 phase compared with controls (n=4-8). This is the first evidence for loop diuretic inhibition of airway smooth muscle cell proliferation. NKCC cotransporter inhibition impeded G1-S phase transition without facilitating cell death. Thus although inhibition by loop diuretics relaxes airway smooth muscle, the NKCC cotransporter may have a more important role in cell proliferation regulation.

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Section: Discussionsupporting

confidence: 88%