title: Intracellular Acid-extruding regulators and the effect of lipopolysaccharide in cultured human renal artery smooth muscle cells.
abstract: Homeostasis of the intracellular pH (pHi) in mammalian cells plays a pivotal role in maintaining cell function. Thus far, the housekeeping Na(+)-H(+) exchanger (NHE) and the Na(+)-HCO3(-) co-transporter (NBC) have been confirmed in many mammalian cells as major acid extruders. However, the role of acid-extruding regulators in human renal artery smooth muscle cells (HRASMCs) remains unclear. It has been demonstrated that lipopolysaccharide (LPS)-induced vascular occlusion is associated with the apoptosis, activating calpain and increased [Ca(2+)]i that are related to NHE1 activity in endothelia cells. This study determines the acid-extruding mechanisms and the effect of LPS on the resting pHi and active acid extruders in cultured HRASMCs. The mechanism of pHi recovery from intracellular acidosis (induced by NH4Cl-prepulse) is determined using BCECF-fluorescence in cultured HRASMCs. It is seen that (a) the resting pHi is 7.19 ± 0.03 and 7.10 ± 0.02 for HEPES- and CO2/HCO3(-)- buffered solution, respectively; (b) apart from the housekeeping NHE1, another Na(+)-coupled HCO3(-) transporter i.e. NBC, functionally co-exists to achieve acid-equivalent extrusion; (c) three different isoforms of NBC: NBCn1 (SLC4A7; electroneutral), NBCe1 (SLC4A4; electrogenic) and NBCe2 (SLC4A5), are detected in protein/mRNA level; and (d) pHi and NHE protein expression/activity are significantly increased by LPS, in both a dose- and time- dependent manner, but NBCs protein expression is not. In conclusion, it is demonstrated, for the first time, that four pHi acid-extruding regulators: NHE1, NBCn1, NBCe1 and NBCe2, co-exist in cultured HRASMCs. LPS also increases cellular growth, pHi and NHE in a dose- and time-dependent manner.
authors.affiliation: Department of Pharmacology, National Defense Medical Center, Taipei City, Taiwan.
authors.forename: Shih-Hurng
authors.initials: SH
authors.lastname: Loh
authors.affiliation: Department of Cardiovascular Surgery, Tri-Service General Hospital, Taipei, Taiwan.
authors.forename: Chung-Yi
authors.initials: CY
authors.lastname: Lee
authors.affiliation: Department of Cardiovascular Surgery, Tri-Service General Hospital, Taipei, Taiwan.
authors.forename: Yi-Ting
authors.initials: YT
authors.lastname: Tsai
authors.affiliation: Department of Pharmacology, National Defense Medical Center, Taipei City, Taiwan.
authors.forename: Shou-Jou
authors.initials: SJ
authors.lastname: Shih
authors.affiliation: Department of Pharmacology, National Defense Medical Center, Taipei City, Taiwan.
authors.forename: Li-Wei
authors.initials: LW
authors.lastname: Chen
authors.affiliation: Department of Biological Science and Technology, College of Life Sciences, China Medical University, Taichung, Taiwan.
authors.forename: Tzu-Hurng
authors.initials: TH
authors.lastname: Cheng
authors.affiliation: Department of General Surgery, Cheng-Hsieng General Hospital, Taipei, Taiwan.
authors.forename: Chung-Yi
authors.initials: CY
authors.lastname: Chang
authors.affiliation: Department of Cardiovascular Surgery, Tri-Service General Hospital, Taipei, Taiwan.
authors.forename: Chein-Sung
authors.initials: CS
authors.lastname: Tsai
mesh_terms: D000136:Acid-Base Equilibrium / Q000276:immunology; D000368:Aged; D027682:Cation Transport Proteins / Q000378:metabolism*; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D005260:Female; D015870:Gene Expression / Q000276:immunology; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008297:Male; D008875:Middle Aged; D009131:Muscle, Smooth, Vascular / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000276:immunology / Q000378:metabolism*; D012077:Renal Artery / Q000473:pathology; D027982:Sodium-Bicarbonate Symporters / Q000378:metabolism; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: D027682:Cation Transport Proteins; D008070:Lipopolysaccharides; C498898:SLC4A4 protein, human; C422022:SLC4A7 protein, human; C490998:SLC9A1 protein, human; D027982:Sodium-Bicarbonate Symporters; D000074082:Sodium-Hydrogen Exchanger 1; D017923:Sodium-Hydrogen Exchangers