scholarly journals Quantitative immunoelectron microscopic analysis of the localization and induction of 25-hydroxyvitamin D3 24-hydroxylase in rat kidney.

1995 ◽  
Vol 43 (3) ◽  
pp. 255-262 ◽  
Author(s):  
K Iwata ◽  
A Yamamoto ◽  
S Satoh ◽  
Y Ohyama ◽  
Y Tashiro ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Vitamin D3 ◽  
Particle Density ◽  
Rat Kidney ◽  
Microscopic Analysis ◽  
Proximal Tubules ◽  
Gold Particles ◽  
Immunogold Technique ◽  
S3 Segment ◽  
Ultrathin Sections

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.

scholarly journals Subcellular distribution of adenosine deaminase and adenosine deaminase-complexing protein in rabbit kidney: implications for adenosine metabolism.

1994 ◽  
Vol 42 (6) ◽  
pp. 775-782 ◽  
Author(s):  
W P Schrader ◽  
C A West ◽  
U H Rudofsky ◽  
W A Samsonoff
Keyword(s):  
Epithelial Cells ◽  
Adenosine Deaminase ◽  
Brush Border ◽  
Renal Tissue ◽  
Proximal Tubules ◽  
Rabbit Kidney ◽  
Specific Cell ◽  
Specific Expression ◽  
Age Related ◽  
S3 Segment

We evaluated the age-related distribution of adenosine deaminase (ADA) and adenosine deaminase-complexing protein (CP) in rabbit kidney by immunohistochemical staining procedures. Paraffin- or resin-embedded tissue from rabbits < 1 week-4 years of age were stained by the peroxidase-anti-peroxidase (PAP) method for ADA and CP. With the exception of neonates, the qualitative staining pattern of each protein remained generally constant with age. In the cortex, distal tubules, blood vessels, histiocytes, and epithelial cells lining Bowman's capsule stained for ADA. Proximal tubules and glomeruli were positive for CP. In contrast to the segregated pattern in the cortex, staining for ADA and CP overlapped in the corticomedullary junction. ADA and CP co-localized on the brush border of tubule cells of the S3 segment. In the cytoplasm of these cells, staining for ADA was characterized by scattered punctuate deposits of peroxidase reaction product. In some instances these punctuate deposits also appeared to be positive for CP. In medulla, epithelial cells of the thin limb were positive for both ADA and CP, whereas papillary collecting ducts stained only for CP. These results document the age-related, tissue-specific expression and localization of ADA in renal tissue, features that probably reflect the crucial role played by the enzyme in adenosine/deoxyadenosine catabolism. In addition, colocalization of ADA and CP on the brush border of cells in the S3 segment of proximal tubules provides support for the hypothesis that one function of CP may be to position ADA on the plasma membrane of specific cell populations, further expanding the enzyme's utility in nucleoside metabolism.

scholarly journals Absence of aquaporin-4 water channels from kidneys of the desert rodent Dipodomys merriami merriami

2001 ◽  
Vol 280 (5) ◽  
pp. F794-F802 ◽  
Author(s):  
Yan Huang ◽  
Randall Tracy ◽  
Glenn E. Walsberg ◽  
Anthony Makkinje ◽  
Pingke Fang ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Rat Kidney ◽  
Freeze Fracture ◽  
Proximal Tubules ◽  
Aquaporin 4 ◽  
Urinary Concentration ◽  
Dipodomys Merriami ◽  
Kangaroo Rat ◽  
S3 Segment ◽  
Aqp4 Protein

Recently, we found that aquaporin-4 (AQP4) is expressed in the S3 segment of renal proximal tubules of mice but not in rat proximal tubules. Because mice have relatively larger papillae than rats, it was proposed that the renal distribution of AQP4 in various species could be related to their maximum urinary concentrating ability. Therefore, kidneys and other tissues of Merriam's desert kangaroo rat, Dipodomys merriami merriami, which produce extremely concentrated urine (up to 5,000 mosmol/kgH2O), were examined for AQP4 expression and localization. Contrary to our expectation, AQP4 immunostaining was undetectable in any region of the kidney, and the absence of AQP4 protein was confirmed by Western blotting. By freeze fracture electron microscopy, orthogonal arrays of intramembraneous particles (OAPs) were not detectable in plasma membranes of principal cells and proximal tubules. However, AQP4 protein was readily detectable in gastric parietal and brain astroglial cells. Northern blotting failed to detect AQP4 mRNA in kangaroo rat kidneys, whereas both in situ hybridization and RT-PCR experiments did reveal AQP4 mRNA in collecting ducts and proximal tubules of the S3 segment. These results suggest that renal expression of AQP4 in the kangaroo rat kidney is regulated at the transcriptional or translational level, and the absence of AQP4 may be critical for the extreme urinary concentration that occurs in this species.

scholarly journals Cerebroside sulfotransferase: preparation of antibody and localization of antigen in kidney.

1981 ◽  
Vol 91 (2) ◽  
pp. 332-339 ◽  
Author(s):  
G I Tennekoon ◽  
J Frangia ◽  
S Aitchison ◽  
D L Price
Keyword(s):  
Catalytic Activity ◽  
Epithelial Cells ◽  
Brush Border ◽  
Plasma Membranes ◽  
Immunohistochemical Study ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Enzyme Protein ◽  
Immunocytochemical Studies ◽  
Phosphoadenosine Phosphosulfate

This immunohistochemical study describes the localization of the enzyme cerebroside sulfotransferase (phosphoadenosine phosphosulfate: galactosylceramide sulfotransferase, EC 2.8.2.11) in rat kidney. The enzyme was purified from kidney and the preparation was used to raise antibodies for immunocytochemical investigations. In the kidney, the antigen was present only on the brush border of the epithelial cells of the proximal tubules, suggesting that sulfation of glycolipids occurs in the cytoplasm and plasma membranes of these specific cells. Moreover, biochemical and immunocytochemical studies of cerebroside sulfotransferase during development indicate that catalytic activity is correlated with the appearance of enzyme protein.

scholarly journals Establishment of renal stem/progenitor-like cell line from S3 segment of proximal tubules in adult rat kidney

2005 ◽  
Vol 68 (5) ◽  
pp. 1966
Author(s):  
Shinji Kitamura ◽  
Yasushi Yamasaki ◽  
Hirofumi Makino
Keyword(s):  
Cell Line ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Adult Rat ◽  
S3 Segment

scholarly journals Immunocytochemical localization of cathepsin B in rat kidney. II. Electron microscopic study using the protein A-gold technique.

1986 ◽  
Vol 34 (7) ◽  
pp. 899-907 ◽  
Author(s):  
S Yokota ◽  
H Tsuji ◽  
K Kato
Keyword(s):  
Cathepsin B ◽  
Rat Kidney ◽  
Protein A ◽  
Proximal Tubules ◽  
Gold Complex ◽  
Electron Microscopic ◽  
Thin Sections ◽  
Gold Particles ◽  
Apical Vesicles ◽  
Collecting Tubules

Thin sections of Lowicryl K4M-embedded materials were labeled with protein A-gold complex. Gold particles representing the antigen sites for cathepsin B were exclusively confined to lysosomes of each segment of the nephron. The heaviest labeling was noted in the lysosomes of the S1 segment of the proximal tubules. Labeling intensity varied considerably with the individual lysosomes. Lysosomes of the other tubular segments, such as the S2 and S3 segments of the proximal tubules, distal convoluted tubules, and collecting tubules were weakly labeled by gold particles. Quantitative analysis of labeling density also confirmed that lysosomes in the S1 segment have the highest labeling density and that approximately 65% of labeling in the whole renal segments, except for the glomerulus, was found in the S1 segment. These results indicate that in rat kidney the lysosomes of the S1 segment are a main location of cathepsin B. Further precise observations on lysosomes of the S1 segment revealed that apical vesicles, tubules, and vacuoles were devoid of gold particles, but when the vacuoles contained fine fibrillar materials, gold labeling was detectable in such vacuoles. As the lysosomal matrix becomes denser, the labeling density is increased. Some small vesicles around the Golgi complex were also labeled. These results indicate that the endocytotic apparatus including the apical vesicles, tubules, and vacuoles contains no cathepsin B. When the vacuoles develop into phagosomes, they acquire this enzyme to digest the absorbed proteins.

Immunolocalization of aquaporin-8 in rat kidney, gastrointestinal tract, testis, and airways

2001 ◽  
Vol 281 (6) ◽  
pp. F1047-F1057 ◽  
Author(s):  
Marie-Louise Elkjær ◽  
Lene N. Nejsum ◽  
Veronika Gresz ◽  
Tae-Hwan Kwon ◽  
Uffe B. Jensen ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Southern Blotting ◽  
Rat Kidney ◽  
Cell Types ◽  
Myoepithelial Cells ◽  
Proximal Tubules ◽  
Basal Cells ◽  
Rt Pcr ◽  
Collecting Ducts ◽  
Outer Stripe

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.

Establishment of renal stem/progenitor-like cell line from S3 segment of proximal tubules in adult rat kidney

2005 ◽  
Vol 68 (5) ◽  
pp. 1966-1966
Author(s):  
Shinji Kitamura ◽  
Yasushi Yamasaki ◽  
Hirofumi Makino
Keyword(s):  
Cell Line ◽  
Rat Kidney ◽  
Proximal Tubules ◽  
Adult Rat ◽  
S3 Segment

Intrarenal and subcellular localization of rat CLC5

1998 ◽  
Vol 275 (5) ◽  
pp. F761-F769 ◽  
Author(s):  
Valerie A. Luyckx ◽  
Fatime O. Goda ◽  
David B. Mount ◽  
Toshiyuki Nishio ◽  
Amy Hall ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Epithelial Cells ◽  
Subcellular Localization ◽  
Chloride Channel ◽  
Rat Kidney ◽  
Polyclonal Antiserum ◽  
Thick Ascending Limb ◽  
Subcellular Membrane ◽  
S3 Segment

Dent’s disease, an inherited disorder characterized by hypercalciuria, nephrolithiasis, nephrocalcinosis, rickets, low-molecular-weight proteinuria, Fanconi’s syndrome, and renal failure, is caused by mutations in the renal chloride channel, CLC5. The normal role of CLC5 is unknown. We have investigated the intrarenal and subcellular localization of CLC5 in rat kidney by in situ hybridization and immunohistochemistry. By in situ hybridization, CLC5 mRNA was detected predominantly in cortical medullary ray and outer medullary tubule epithelial cells. Polyclonal antiserum was generated against a CLC5 fusion protein, affinity purified, and immunoadsorbed against CLC3 and CLC4 to yield a CLC5 isoform-specific antiserum. By immunohistochemistry, CLC5 protein was localized to the intracellular domain of tubular epithelial cells in the S3 segment of the proximal tubule and the medullary thick ascending limb. By subcellular membrane fractionation and flow cytometry, CLC5 expression was found in outer medullary endosomes. These findings are consistent with a model in which CLC5 encodes an endosomal chloride channel that facilitates acidification and trafficking of renal epithelial endosomes.

scholarly journals Ultrastructural localization of arylsulfatase C activity in rat kidney.

1987 ◽  
Vol 35 (5) ◽  
pp. 523-530 ◽  
Author(s):  
J Kawano ◽  
E Aikawa
Keyword(s):  
Endoplasmic Reticulum ◽  
Enzymatic Hydrolysis ◽  
Epithelial Cells ◽  
Barium Sulfate ◽  
Rat Kidney ◽  
Barium Chloride ◽  
Ultrastructural Localization ◽  
Proximal Tubules ◽  
Metal Precipitation ◽  
Biochemical Studies

Metal precipitation techniques for ultrastructural demonstration of arylsulfatase C activity were studied in rat kidney. Possible substrates for the techniques were biochemically tested with regard to their velocity of enzymatic hydrolysis and their specificity for arylsulfatase C. Effects of buffers and capturing metals were also examined. The results of these biochemical studies were then verified histochemically. Incubation in a medium containing 1 mM 4-methylumbelliferyl sulfate, 1% barium chloride, 0.1 M imidazole-HCl buffer (pH 7.5), and 5% sucrose achieved identifiable results in adequately fixed kidney. Precipitation of barium sulfate was localized mainly in the endoplasmic reticulum and perinuclear cisterns of the epithelial cells in the descending portions of proximal tubules.

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