Quantitative immunoelectron microscopic analysis of the localization and induction of 25-hydroxyvitamin D3 24-hydroxylase in rat kidney.
25-Hydroxyvitamin D3 24(R)-hydroxylase (24-hydroxylase) is involved in the metabolism and regulation of vitamin D3 and is markedly induced by administration of vitamin D3. We detected this enzyme by electron microscopy and an immunogold technique along nephrons of normal and vitamin D3-administered rats. After the rats were administered vitamin D3, 50,000 IU/day for 1 week, they were perfusion-fixed with a paraformaldehyde solution. The fixed kidneys were then removed and embedded in LR White resin. Ultrathin sections were prepared and labeled by the immunogold technique using a mouse anti-rat 25-hydroxyvitamin D3 24-hydroxylase monoclonal antibody. We counted the number of gold particles bound per micron 2 of the mitochondria (particle density) of the tubule epithelial cells along the nephrons. In normal and vitamin D3-administered rats, gold particles were observed in the mitochondria of epithelial cells along the tubules. In normal rats, gold labeling for 24-hydroxylase was statistically significant (p < 0.05), in the S1-S2 segments, the S3 segment of the proximal tubules, and in the distal convoluted tubules. In the rats administered vitamin D3, the particle density increased significantly (p < 0.05) by about 12-fold in the S1-S2 segments of the proximal tubules, whereas it increased less markedly in other parts of the nephron. The marked induction of the S1-S2 segments of the proximal tubules suggests that these segments play an important role in the regulation of vitamin D3 metabolism.
Iwata
K
K
Department of Surgery I, Miyazaki Medical College, Japan.
Yamamoto
A
A
Satoh
S
S
Ohyama
Y
Y
Tashiro
Y
Y
Setoguchi
T
T
D000818:Animals; D000911:Antibodies, Monoclonal; D002762:Cholecalciferol / Q000494:pharmacology*; D003577:Cytochrome P-450 Enzyme System / Q000032:analysis* / Q000096:biosynthesis*; D004790:Enzyme Induction; D005260:Female; D015151:Immunoblotting; D007074:Immunoglobulin G; D007668:Kidney / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D007685:Kidney Tubules, Collecting / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D007686:Kidney Tubules, Distal / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D007687:Kidney Tubules, Proximal / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D008138:Loop of Henle / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D016253:Microscopy, Immunoelectron / Q000379:methods; D008928:Mitochondria / Q000201:enzymology / Q000648:ultrastructure; D008970:Molecular Weight; D009399:Nephrons / Q000187:drug effects / Q000201:enzymology / Q000648:ultrastructure; D051381:Rats; D017208:Rats, Wistar; D013250:Steroid Hydroxylases / Q000032:analysis* / Q000096:biosynthesis*; D065668:Vitamin D3 24-Hydroxylase
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D002762:Cholecalciferol; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D065668:Vitamin D3 24-Hydroxylase