Troglitazone's rapid and sustained activation of ERK1/2 induces cellular acidosis in LLC-PK1-F+ cells: physiological responses
| Autor: | Ellen Friday, Robert E. Oliver Iii, Francesco Turturro, Tomas Welbourne, Ashley Lacy |
|---|---|
| Rok vydání: | 2005 |
| Předmět: |
medicine.medical_specialty
Physiology Swine Intracellular pH Kidney Tubules Proximal chemistry.chemical_compound Troglitazone Biosynthesis Internal medicine medicine Animals Hypoglycemic Agents Chromans Acidosis chemistry.chemical_classification Acid-Base Equilibrium Mitogen-Activated Protein Kinase 1 Mitogen-Activated Protein Kinase 3 DNA synthesis Chemistry Cell biology Enzyme Activation Sodium–hydrogen antiporter Enzyme Endocrinology LLC-PK1 Cells Thiazolidinediones medicine.symptom Intracellular medicine.drug |
| Zdroj: | American journal of physiology. Renal physiology. 288(6) |
| ISSN: | 1931-857X |
| Popis: | We studied the signal pathway through which troglitazone (TRO) acts in inducing cellular acidosis in LLC-PK1-F+cells in relation to ammoniagenesis and DNA synthesis. Cells were grown to confluent monolayers in 30-mm chambers and monitored for intracellular pH (pHi) by the BCECF assay and activated ERK by phospo-ERK1/2 antibodies. TRO induces a severe cellular acidosis (pHi6.68 ± 0.10 vs. 7.28 ± 0.07 time control at 4 min, P < 0.01), whereas phospho-ERK1/2 to total ERK1/2 ratio increases 3.4-fold ( P < 0.01). To determine whether ERK1/2 was activated by cellular acidosis or TRO was acting via MEK1/2 to activate ERK1/2, cells were pretreated with specific inhibitors of MEK1/2 activity, PD-098059 and U-0126, followed by the addition of TRO or vehicle. With MEK1/2 activity inhibited, TRO treatment failed to activate ERK1/2. Preventing ERK1/2 activation abrogated the TRO-induced cellular acidosis and maintained the pHiwithin the low normal range (7.06 ± 0.11). To determine whether blocking ERK activation prevents TRO's inhibitory effect on NHE activity, cells were acid-loaded and the recovery response was monitored as ΔpHi/ t over a 4-min recovery period. TRO inhibited NHE activity by 85% ( P < 0.01), whereas blocking ERK activation restored the response. We measured activated ERK levels and pHiafter 3- and 18-h exposure to TRO or extracellular acidosis (pHe = 6.95) to determine whether ERK activation was sustained. Whereas both TRO and extracellular acidosis increased activated ERK and decreased pHiafter 3 h, only TRO sustained this response at 18 h. Furthermore, both enhanced ammoniagenesis and decreased DNA synthesis reflected the effect of TRO to induce and sustain a cellular acidosis. |
| Databáze: | OpenAIRE |
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