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1 The Water and Salt Research Center, University of Aarhus, DK-8000 Aarhus C; 2 Department of Physiology, Dongguk University, 780 - 714 Kyungju, Korea; and 3 Institute of Human Genetics, University of Aarhus, DK-8000 Aarhus C, Denmark
First published August 8, 2001;
10.1152/ajprenal.00158.2001.
The purpose of this study was to
determine the cellular and subcellular localization of aquaporin-8
(AQP8) in rat kidney and other organs by RT-PCR analyses and by
immunoblotting and immunohistochemistry using peptide-derived rabbit
antibodies to rat AQP8. RT-PCR and Southern blotting revealed the
presence of AQP8 mRNA in all kidney zones. LLC-PK1 cells
transfected with a rat AQP8 construct exhibited strong labeling with
the affinity-purified antibodies, whereas controls using cells
transfected with the vector, but without the insert, were negative. The
labeling was almost exclusively associated with intracellular vesicles.
Immunoblotting of kidney membrane fractions revealed a predominant
single band of 26-28 kDa. AQP8 immunoreactivity was mainly present
in the cortex and outer stripe of the outer medulla. Sequential
ultracentrifugation of rat kidney membrane revealed that AQP8 resides
predominantly in intracellular vesicular fractions. Immunocytochemistry
revealed modest labeling of proximal tubules and weak labeling of
collecting ducts in cortex and medulla of rat kidney. The labeling was
confined to cytoplasmic areas with no labeling of the brush border.
Immunoblotting and RT-PCR/Southern blotting also revealed the presence
of AQP8 protein and mRNA in rat liver, testis, epididymis, duodenum,
jejunum, colon, and bronchi/trachea. Consistent with this,
immunohistochemistry revealed AQP8 labeling in the hepatocytes and
spematogenic cells in testis and in the basal cells in ductus
epididymis, trachea, and bronchial epithelia. Moreover, AQP8 labeling
was observed in the myoepithelial cells in salivary, bronchial, and
tracheal glands with no labeling of acini or ductal epithelial cells.
AQP8 is also present in the surface epithelial cells in duodenum,
jejunum, and colon. In conclusion, AQP8 is expressed at low levels in
rat kidney proximal tubules and collecting ducts, and it is present in
distinct cell types in liver, testis, epididymis, duodenum, jejunum,
colon, trachea, and principal bronchi as well as in multiple glands,
including salivary glands.
intracellular water channel; renal; immunocytochemistry
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