Inhibition of the Na+-H+ exchanger isoform-1 and the extracellular signal-regulated kinase induces apoptosis: a time course of events.
OBJECTIVE
The present study attempts to shed light on the role and the relative position of the Na(+)/H(+) exchanger isoform 1 (NHE1) and the extracellular signal-regulated kinase (ERK) in HEp-2 cell signaling pathways concerning a diverse range of cellular functions such as regulation of intracellular pH (pHi), DNA synthesis, production of reactive oxygen species (ROS) and apoptosis.


METHODS
Pharmacological inhibition with cariporide (highly specific inhibitor of NHE1) and PD98059 (specific inhibitor of the upstream activator of ERK) was implemented. Fluorescence spectrometry, atomic absorption spectrometry and ELISA methods were used in order to obtain the results.


RESULTS
NHE1 and ERK take part in all of the aforementioned cellular functions, as their inhibition had an effect on all of them. Additionally, inhibition of NHE1 resulted in ERK inhibition as well. Moreover, continuous inhibition of NHE1 or ERK for up to 24h led HEp-2 cells to apoptosis, as assessed through caspase-3 activation, DNA fragmentation and annexin-V binding levels.


CONCLUSIONS
Our data shows a time course of events in relation to NHE1 and ERK and suggests the existence of a positive feedback loop between NHE1 and ERK which could pose a barrier against apoptosis.
Konstantinidis
Diamantis
D
Laboratory of Animal Physiology, School of Biology, Aristotle University of Thessaloniki, Greece.
Koliakos
George
G
Vafia
Katerina
K
Liakos
Panagiotis
P
Bantekas
Christos
C
Trachana
Varvara
V
Kaloyianni
Martha
M
D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D027682:Cation Transport Proteins / Q000037:antagonists & inhibitors*; D002478:Cells, Cultured; D053938:DNA Fragmentation; D004261:DNA Replication / Q000187:drug effects; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors*; D005419:Flavonoids / Q000494:pharmacology; D006146:Guanidines / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D020033:Protein Isoforms / Q000037:antagonists & inhibitors; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017382:Reactive Oxygen Species / Q000378:metabolism; D012964:Sodium / Q000378:metabolism; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors*; D013450:Sulfones / Q000494:pharmacology; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors
D017304:Annexin A5; D027682:Cation Transport Proteins; D005419:Flavonoids; D006146:Guanidines; D020033:Protein Isoforms; D047428:Protein Kinase Inhibitors; D017382:Reactive Oxygen Species; C490998:SLC9A1 protein, human; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers; D013450:Sulfones; C093373:cariporide; D012964:Sodium; D048049:Extracellular Signal-Regulated MAP Kinases; D053148:Caspase 3; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D013936:Thymidine