Expression of NOS1 and soluble guanylyl cyclase by human kidney epithelial cells: morphological evidence for an autocrine/paracrine action of nitric oxide.
BACKGROUND
Nitric oxide plays an important role in the kidney through effects on both renal hemodynamics and tubular functions. Tubular epithelial cells are thus a target for nitric oxide. However, as to whether tubular epithelial cells endogeneously produce nitric oxide under physiologic conditions in human kidney is currently unknown. The aim of the present study was to characterize and localize in situ the nitric oxide synthase (NOS) isoforms (NOS1, NOS2, and NOS3) expressed in human normal kidney, and soluble guanylyl cyclase, the well-known target for nitric oxide.


METHODS
Five complementary experimental approaches were used: (1) detection of NOS reductase activity by nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry, (2) immunolocalization of the NOS isoforms (NOS1, NOS2, NOS3), (3) immunoblot analysis, (4) quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis of NOS mRNA, and (5) measurement of NOS activity as the conversion rate of l-[14C]-arginine to l-[14C]-citrulline. In addition, in situ detection of soluble guanylyl cyclase was assessed by immunohistochemistry.


RESULTS
All these techniques led to consistent results showing that epithelial cells of most tubules along the human nephron exhibit functional NOS1, with a corticomedullary gradient observed both at the protein and mRNA levels. Moreover, epithelial cells expressing NOS1 also express soluble guanylyl cyclase, indicating that these cells possess the machinery for autocrine/paracrine effect of nitric oxide.


CONCLUSIONS
The present study demonstrates that NOS1 is strongly expressed in most tubules of the human nephron and therefore invites to consider epithelial cells as one of the major source of nitric oxide in the human kidney under physiologic conditions.
Jarry
Anne
A
INSERM U539, Faculty of Medicine, Nantes, France. ajarry@sante.univ-nantes.fr
Renaudin
Karine
K
Denis
Marc G
MG
Robard
Myriam
M
Buffin-Meyer
Bénédicte
B
Karam
Georges
G
Buzelin
Françoise
F
Paris
Hervé
H
Laboisse
Christian L
CL
Vallette
Geneviève
G
D019898:Autocrine Communication; D004847:Epithelial Cells / Q000378:metabolism; D006162:Guanylate Cyclase / Q000378:metabolism*; D006651:Histocytochemistry; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D007668:Kidney / Q000166:cytology / Q000378:metabolism*; D009252:NADPH Dehydrogenase / Q000378:metabolism; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism*; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D010088:Oxidoreductases / Q000378:metabolism; D019899:Paracrine Communication; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012995:Solubility
D012333:RNA, Messenger; D010088:Oxidoreductases; C496320:NOS1 protein, human; C496319:NOS2 protein, human; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D009252:NADPH Dehydrogenase; D006162:Guanylate Cyclase