Characterization of the Na/HCO3(-) cotransport in human neutrophils.
BACKGROUND
Bicarbonate transport has crucial roles in regulating intracellular pH (pHi) in a variety of cells. The purpose of this study was to evaluate its participation in the regulation of pHi in resting and stimulated human neutrophils.


METHODS
Freshly isolated human neutrophils acidified by an ammonium prepulse were used in this study.


RESULTS
We demonstrated that resting neutrophils have a bicarbonate transport mechanism that prevents acidification when the Na(+)/H(+) exchanger is blocked by EIPA. Neutrophils acidified by an ammonium prepulse showed an EIPA-resistant recovery of pHi that was inhibited by the blocker of the anionic transporters SITS or the Na(+)/HCO3(-) cotransporter (NBC) selective inhibitor S0859, and abolished when sodium was removed from the extracellular medium. In western blot and RT-PCR analysis the expression of NBCe2 but not NBCe1 or NBCn1 was detected in neutrophils Acidified neutrophils increased the EIPA-insensitive pHi recovery rate when its activity was stimulated with fMLF/ cytochalasin B. This increase in the removal of acid equivalents was insensitive to the blockade of the NADPH oxidase with DPI.


CONCLUSIONS
It is concluded that neutrophils have an NBC that regulates basal pHi and is modulated by chemotactic agents.
Centro de Investigaciones Cardiovasculares, CCT La Plata, CONICET. Facultad de Ciencias Médicas, Universidad Nacional de La Plata, La Plata, Argentina.
Miriam S
MS
Giambelluca
María C
MC
Ciancio
Alejandro
A
Orlowski
Oscar A
OA
Gende
Marc
M
Pouliot
Ernesto A
EA
Aiello
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D001549:Benzamides / Q000494:pharmacology; D001639:Bicarbonates / Q000494:pharmacology; D003571:Cytochalasin B / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport / Q000187:drug effects; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D027982:Sodium-Bicarbonate Symporters / Q000235:genetics / Q000378:metabolism*; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology
D001549:Benzamides; D001639:Bicarbonates; D020033:Protein Isoforms; D012333:RNA, Messenger; C528718:S 0859 compound; D027982:Sodium-Bicarbonate Symporters; D017923:Sodium-Hydrogen Exchangers; D013449:Sulfonamides; D000643:Ammonium Chloride; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D003571:Cytochalasin B; D009240:N-Formylmethionine Leucyl-Phenylalanine; D000584:Amiloride; D019255:NADPH Oxidases; C039614:ethylisopropylamiloride