The characterisation and uptake of paraquat in cultured baboon kidney proximal tubule cells (bPTC)

2001 ◽  
Vol 20 (2) ◽  
pp. 90-99 ◽  
Author(s):  
R Machaalani ◽  
V Lazzaro ◽  
G G Duggin
Keyword(s):  
Proximal Tubule ◽  
Mdck Cells ◽  
Distal Tubule ◽  
Genetic Mutation ◽  
Immunoperoxidase Staining ◽  
Renal Tubules ◽  
Trypan Blue Exclusion ◽  
Proximal Tubule Cells ◽  
Morphological Studies ◽  
Tubule Cells

A primary culture of baboon proximal tubule cells (bPTC) was prepared and characterised using LLC-PK1 cells of proximal tubule origin and MDCK cells of distal tubule origin, as positive and negative references, respectively. The proximal tubular origin of the bPTC was determined by morphological studies, immunoperoxidase staining and the expression of proximal tubule markers alkaline phosphatase and gammaglutamyltransferase. The hypothesis that paraquat (PQ) is transported by the bPTC was investigated. The cytotoxic threshold for PQ in these cells was determined and compared to the LLC-PK1 and MDCK cells. Furthermore, this study investigated the transport of the monovalent cation tetraethyl ammonium (TEA) and the polyvalent cation cimetidine in the bPTC and demonstrated their effect on the cellular uptake of PQ. The cytotoxic threshold of PQ in the bPTC, determined by cellular viability studies using the method of Trypan blue exclusion, is 0.05 mM at 2 h incubation. The LC50 after 24 his 76, 61 and 455 pM for the bPTC, LLC-PK1 and MDCK cells, respectively. This indicates that proximal tubule cells are more susceptible to PQ toxicity compared to distal tubule cells, which is consistent with clinical PQ toxicity where renal damage is found predominantly in the proximal renal tubules. The cations PQ and cimetidine were actively transported by the bPTC. The uptake of PQ (0.05 mM) commenced after 15 min whereas cimetidine (0.5 mM) uptake was evident after 2 min. Furthermore, cimetidine was shown to compete with PQ for uptake in the bPTC. Coincubating PQ (0.05 mM) and cimetidine (0.5 mM) for 60 min resulted in an approximate 50% decrease in PQ uptake. The cation TEA was not transported by the bPTC suggesting either a genetic mutation or complete absence of the transporter for TEA in the cells. The results suggest that PQ may be transported by the same cation transporter as cimetidine and not TEA, indicating PQ uptake in the bPTC to be via a polyvalent organic cation transporter.

Aluminium deposition in liver and kidney following acute intravenous administration of aluminium chloride or citrate in conscious rats

1995 ◽  
Vol 14 (10) ◽  
pp. 787-794 ◽  
Author(s):  
AJ Spencer ◽  
JA Wood ◽  
HC Saunders ◽  
MS Freeman ◽  
CJ Lote
Keyword(s):  
Proximal Tubule ◽  
Intravenous Administration ◽  
Distal Tubule ◽  
Dry Weight ◽  
Aluminium Chloride ◽  
Proximal Tubule Cells ◽  
Al Content ◽  
Liver And Kidney ◽  
Wet Weight ◽  
Tubule Cells

1 Plasma, urinary, liver and kidney cell aluminium (Al) levels were monitored in the rat, 1h after intravenous administration of 29630 nmol (800 μg) Al as either Al chloride or as Al citrate (Al chloride plus excess sodium citrate). Al levels were measured in plasma, urine and liver by atomic absorption spectroscopy (AAS). Liver and kidney Al content was measured at the cellular and subcellular level by electron probe X-ray microanalysis (EPXMA). 2 Urinary excretion of Al was significantly higher ( P < 0.01), when Al was given as the citrate than as the chloride. After 1h, plasma Al levels were significantly lower in the Al citrate group than the Al chloride group (59 ± 3.7 vs 877 ± 214 nmol ml-1, respectively; P< 0.01). 3 Al concentrations were significantly higher in the livers of rats receiving Al chloride (818 ± 252 nmol g-1 wet weight; P < 0.05), than in either control or Al citrate groups (122 ± 41 and 107 ± 26 nmol g-1 wet weight, respectively). Al concentrations derived from EPXMA measurements were in agreement with AAS values for the three groups, with significantly higher Al concentra tions in the Al chloride group (1.7 ± 0.4 nmol mg-1 dry weight; P < 0.05) than in the control or Al citrate groups, where Al was not detectable. EPXMA analysis showed that Al was distributed in all liver organelles analysed (cytoplasm, mitochondria, nucleus, ER) and was not preferentially taken up by any one organelle in Al chloride treated rats. 4 Significant amounts of Al were found in cytoplasm and mitochondria of proximal tubule cells of rats given Al citrate (0.64 ± 0.15 and 0.80 ± 0.11 nmol mg-1 dry weight, respectively), but not in nuclei or lysosomes of these cells. Al levels were not detectable in control kid neys, in proximal tubule cells after Al chloride adminis tration or distal tubule cells after either Al treatment.

scholarly journals Antenatal betamethasone attenuates the angiotensin-(1–7)-Mas receptor-nitric oxide axis in isolated proximal tubule cells

2017 ◽  
Vol 312 (6) ◽  
pp. F1056-F1062 ◽  
Author(s):  
Yixin Su ◽  
Jianli Bi ◽  
Victor M. Pulgar ◽  
Mark C. Chappell ◽  
James C. Rose
Keyword(s):  
Nitric Oxide ◽  
Proximal Tubule ◽  
Cell Model ◽  
Primary Cultures ◽  
Specific Effect ◽  
Renal Tubules ◽  
Proximal Tubule Cells ◽  
Mas Receptor ◽  
Tubule Cells

We previously reported a sex-specific effect of antenatal treatment with betamethasone (Beta) on sodium (Na+) excretion in adult sheep whereby treated males but not females had an attenuated natriuretic response to angiotensin-(1–7) [Ang-(1–7)]. The present study determined the Na+ uptake and nitric oxide (NO) response to low-dose Ang-(1–7) (1 pM) in renal proximal tubule cells (RPTC) from adult male and female sheep antenatally exposed to Beta or vehicle. Data were expressed as percentage of basal uptake or area under the curve for Na+ or percentage of control for NO. Male Beta RPTC exhibited greater Na+ uptake than male vehicle cells (433 ± 28 vs. 330 ± 26%; P < 0.05); however, Beta exposure had no effect on Na+ uptake in the female cells (255 ± 16 vs. 255 ± 14%; P > 0.05). Ang-(1–7) significantly inhibited Na+ uptake in RPTC from vehicle male (214 ± 11%) and from both vehicle (190 ± 14%) and Beta (209 ± 11%) females but failed to attenuate Na+ uptake in Beta male cells. Beta exposure also abolished stimulation of NO by Ang-(1–7) in male but not female RPTC. Both the Na+ and NO responses to Ang-(1–7) were blocked by Mas receptor antagonist d-Ala7-Ang-(1–7). We conclude that the tubular Ang-(1–7)-Mas-NO pathway is attenuated in males and not females by antenatal Beta exposure. Moreover, since primary cultures of RPTC retain both the sex and Beta-induced phenotype of the adult kidney in vivo they appear to be an appropriate cell model to examine the effects of fetal programming on Na+ handling by the renal tubules.

Gentamicin suppresses endotoxin-driven TNF-α production in human and mouse proximal tubule cells

2007 ◽  
Vol 293 (4) ◽  
pp. F1373-F1380 ◽  
Author(s):  
Richard A. Zager ◽  
Ali C. M. Johnson ◽  
Adam Geballe
Keyword(s):  
Protein Synthesis ◽  
Proximal Tubule ◽  
Renal Tubules ◽  
Mrna Accumulation ◽  
Chemokine Production ◽  
Proximal Tubule Cells ◽  
Gram Negative ◽  
Essentially Normal ◽  
Tnf Α ◽  
Tubule Cells

Gentamicin is a mainstay in treating gram-negative sepsis. However, it also may potentiate endotoxin (LPS)-driven plasma TNF-α increases. Because gentamicin accumulates in renal tubules, this study addressed whether gentamicin directly alters LPS-driven tubular cell TNF-α production. HK-2 proximal tubular cells were incubated for 18 h with gentamicin (10–2,000 μg/ml). Subsequent LPS-mediated TNF-α increases (at 3 or 24 h; protein/mRNA) were determined. Gentamicin effects on overall protein synthesis ([35S]methionine incorporation), monocyte chemoattractant protein-1 (MCP-1) levels, and LPS-stimulated TNF-α generation by isolated mouse proximal tubules also were assessed. Finally, because gentamicin undergoes partial biliary excretion, its potential influence on gut TNF-α/MCP-1 mRNAs was probed. Gentamicin caused striking, dose-dependent inhibition of LPS-driven TNF-α production (up to 80% in HK-2 cells/isolated tubules). Surprisingly, this occurred despite increased TNF-α mRNA accumulation. Comparable changes in MCP-1 were observed. These changes were observed at clinically relevant gentamicin concentrations and despite essentially normal overall protein synthetic rates. Streptomycin also suppressed LPS-driven TNF-α increases, suggesting an aminoglycoside drug class effect. Gentamicin doubled basal TNF-α mRNA in cecum and in small intestine after LPS. Gentamicin can suppress LPS-driven TNF-α production in proximal tubule cells, likely by inhibiting its translation. Overall preservation of protein synthesis and comparable MCP-1 suppression suggest a semiselective blockade within the LPS inflammatory mediator cascade. These results, coupled with increases in gut TNF-α/MCP-1 mRNAs, imply that gentamicin may exert protean, countervailing actions on systemic cytokine/chemokine production during gram-negative sepsis.

scholarly journals Steroidogenic acute regulatory-related lipid transfer domain protein 5 localization and regulation in renal tubules

2009 ◽  
Vol 297 (2) ◽  
pp. F380-F388 ◽  
Author(s):  
Yu-Chyu Chen ◽  
Renate K. Meier ◽  
Shirong Zheng ◽  
Syed J. Khundmiri ◽  
Michael T. Tseng ◽  
...  
Keyword(s):  
Er Stress ◽  
Proximal Tubule ◽  
Mouse Kidney ◽  
Cell Periphery ◽  
Type I ◽  
Renal Tubules ◽  
Lipid Transfer ◽  
Proximal Tubule Cells ◽  
Cholesterol Levels ◽  
Tubule Cells

STARD5 is a cytosolic sterol transport protein that is predominantly expressed in liver and kidney. This study provides the first report on STARD5 protein expression and distribution in mouse kidney. Immunohistochemical analysis of C57BL/6J mouse kidney sections revealed that STARD5 is expressed in tubular cells within the renal cortex and medullar regions with no detectable staining within the glomeruli. Within the epithelial cells of proximal renal tubules, STARD5 is present in the cytoplasm with high staining intensity along the apical brush-border membrane. Transmission electronmicroscopy of a renal proximal tubule revealed STARD5 is abundant at the basal domain of the microvilli and localizes mainly in the rough endoplasmic reticulum (ER) with undetectable staining in the Golgi apparatus and mitochondria. Confocal microscopy of STARD5 distribution in HK-2 human proximal tubule cells showed a diffuse punctuate pattern that is distinct from the early endosome marker EEA1 but similar to the ER membrane marker GRP78. Treatment of HK-2 cells with inducers of ER stress increased STARD5 mRNA expression and resulted in redistribution of STARD5 protein to the perinuclear and cell periphery regions. Since recent reports show elevated ER stress response gene expression and increased lipid levels in kidneys from diabetic rodent models, we tested STARD5 and cholesterol levels in kidneys from the OVE26 type I diabetic mouse model. Stard5 mRNA and protein levels are increased 2.8- and 1.5-fold, respectively, in OVE26 diabetic kidneys relative to FVB control kidneys. Renal free cholesterol levels are 44% elevated in the OVE26 mice. Together, our data support STARD5 functioning in kidney, specifically within proximal tubule cells, and suggest a role in ER-associated cholesterol transport.

scholarly journals Exosomal transfer from human renal proximal tubule cells to distal tubule and collecting duct cells

2014 ◽  
Vol 47 (15) ◽  
pp. 89-94 ◽  
Author(s):  
John J. Gildea ◽  
Joscelyn E. Seaton ◽  
Ken G. Victor ◽  
Camellia M. Reyes ◽  
Dora Bigler Wang ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Collecting Duct ◽  
Distal Tubule ◽  
Renal Proximal Tubule ◽  
Proximal Tubule Cells ◽  
Duct Cells ◽  
Collecting Duct Cells ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

scholarly journals Expression of gp330 and gp330/alpha 2-macroglobulin receptor-associated protein in renal tubular differentiation.

1994 ◽  
Vol 4 (12) ◽  
pp. 2003-2015 ◽  
Author(s):  
M Abbate ◽  
D R Bachinsky ◽  
R T McCluskey ◽  
D Brown
Keyword(s):  
Ligand Binding ◽  
Proximal Tubule ◽  
Rat Kidney ◽  
Immunoperoxidase Staining ◽  
Apical Surface ◽  
Proximal Tubule Cells ◽  
Receptor Associated Protein ◽  
Tubule Cells

The gp330/alpha 2-macroglobulin receptor-associated protein (RAP) is a 39- to 45-kd protein that binds to the low-density lipoprotein receptor-related protein/alpha 2-macroglobulin receptor and to gp330, a major glycoprotein of the brush border of proximal tubule cells. Despite evidence that gp330 functions as a receptor for several ligands and that soluble RAP inhibits ligand binding to gp330 in vitro, the physiologic function of RAP is unknown. Given the predominant location of RAP within the rough endoplasmic reticulum (RER), RAP might be involved in the intracellular processing and/or transport of gp330. The developing rat kidney was used as a dynamic model to study in detail the relationship between gp330 and RAP in vivo by immunohistochemical techniques. RAP was expressed in the renal vesicle and continued to be present, with a vesicular and perinuclear pattern of staining, in both proximal tubule cells and glomerular cells at subsequent stages. Immunoperoxidase electron microscopy demonstrated RAP in cisternae of the RER and in large subapical vesicles. gp330 was initially expressed in early proximal tubule cells in S-shaped bodies and was located in the perinuclear envelope and cytoplasmic vesicles as well as at the apical surface. Cytoplasmic gp330 staining was more evident at a stage subsequent to the S-shaped body, possibly related to more active biosynthesis. By comparative analysis of the patterns of immunofluorescence and immunoperoxidase staining, gp330 and RAP colocalized in the RER and in some large subapical vacuoles, but no definite RAP staining could be detected at the surface of proximal tubule cells at any stage, despite the presence of abundant gp330 in this location. The expression of gp330 at the apical surface of immature tubular cells was associated with the onset of fluid-phase endocytosis of fluoroscein isothiocyanate-dextran and, therefore, of reabsorption of material from the tubular lumen, in the absence of concomitant changes in RAP expression in the same cells. These findings indicate that the role of endogenous RAP may not be directly related to ligand binding of gp330 at the surface of proximal tubule cells, although RAP may be involved in the processing and the intracellular trafficking of newly synthesized gp330, in particular in the delivery of gp330 to the plasma membrane.

The vitamin D receptor in the proximal renal tubule is a key regulator of serum 1α,25-dihydroxyvitamin D3

2015 ◽  
Vol 308 (3) ◽  
pp. E201-E205 ◽  
Author(s):  
Yongji Wang ◽  
Jinge Zhu ◽  
Hector F. DeLuca
Keyword(s):  
Vitamin D ◽  
Proximal Tubule ◽  
Vitamin D Receptor ◽  
Brush Border ◽  
Distal Tubule ◽  
Proximal Convoluted Tubule ◽  
Proximal Tubule Cells ◽  
Hydroxyvitamin D ◽  
25 Hydroxyvitamin D ◽  
Tubule Cells

It is well established that the mitochondria of proximal convoluted tubule cells of the kidney are the site of production of circulating 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3]. The production of 1,25(OH)2D3 at this site is tightly regulated. Parathyroid hormone markedly stimulates 1,25(OH)2D3 production, whereas 1,25(OH)2D3 itself suppresses production. The mechanism of suppression by 1,25(OH)2D3 has not yet been elucidated. We have now found that in the absence of vitamin D (vitamin D deficiency), the vitamin D receptor (VDR) is found in the interior of the apical brush border of the proximal tubule cells. This is unique for the proximal tubule cells, since this has not been observed in the distal tubule cells or in other epithelial cells, such as intestinal mucosa. Administration of 1,25(OH)2D3 to vitamin D-deficient rats results in the movement of VDR from the brush border to the cytoplasm and nucleus presumably bound to reabsorbed 1,25(OH)2D3. The VDR bound to 1,25(OH)2D3 suppresses expression of 25-hydroxyvitamin D3 1α-hydroxylase and stimulates the 25-hydroxyvitamin D3 24-hydroxylase. Thus, VDR in the apical brush border of the proximal convoluted tubule cells serves to “sense” the level of circulating 1,25(OH)2D3 and modulates the activity of the 1α-hydroxylase and the 24-hydroxylase accordingly.

scholarly journals Immunohistochemical studies on hemoglobin-haptoglobin and hemoglobin catabolism sites.

1987 ◽  
Vol 35 (3) ◽  
pp. 381-386 ◽  
Author(s):  
K Kino ◽  
K Mizumoto ◽  
J Watanabe ◽  
H Tsunoo
Keyword(s):  
Proximal Tubule ◽  
Binding Capacity ◽  
Distal Tubule ◽  
Cell Types ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Immunohistochemical Methods ◽  
Pass Through ◽  
Immunohistochemical Studies ◽  
Parenchymal Cells

In order to clarify the catabolism sites of Hb-Hp and free Hb, the organ distributions of [125I]-Hb-Hp and [125I]-Hb were studied, and the cell types in each organ incorporating them were determined by immunohistochemical methods. After administration of [125I]-Hb-Hp in very small amounts to rats, 84.5% was incorporated into the liver, but the renal uptake was only 0.6%. [125I]-Hb was incorporated into the kidneys rather than into the liver when a fivefold greater amount of [125I]-Hb than the binding capacity of plasma Hp was administered. Parenchymal cells, but not Kupffer cells, in the liver were stained with anti-Hb or anti-Hp IgG after administration of Hb in an amount corresponding to the Hb binding capacity of Hp. The proximal tubule cells, but not the distal tubule cells, in the kidney were stained with anti-Hb IgG after administration of a fivefold greater amount of Hb than the binding capacity of Hp. On the basis of these results, we suggest that Hb-Hp was incorporated mainly into liver parenchymal cells and did not traverse glomeruli in the kidney. In contrast to Hb-Hp, free Hb could pass through the glomeruli easily and was incorporated into the proximal tubule cells.

scholarly journals PROBLEMS OF SPECIFICITY IN THE USE OF A STRONTIUM CAPTURE TECHNIQUE FOR THE CYTOCHEMICAL LOCALIZATION OF OUABAIN-SENSITIVE, POTASSIUM-DEPENDENT PHOSPHATASE IN MAMMALIAN RENAL TUBULES

1974 ◽  
Vol 22 (12) ◽  
pp. 1163-1168 ◽  
Author(s):  
J. A. FIRTH
Keyword(s):  
Alkaline Phosphatase ◽  
Proximal Tubule ◽  
Adenosine Triphosphatase ◽  
Plasma Membranes ◽  
Distal Tubule ◽  
Renal Tubules ◽  
Cytochemical Localization ◽  
Nitrophenyl Phosphate ◽  
Basal Plasma ◽  
Tubule Cells

The p-nitrophenyl phosphate-strontium procedure for the localization of the phosphatase component of Na-K-activated adenosine triphosphatase was evaluated using rat renal cortex as a test tissue. The results obtained by light microscopy were unexpected in that reaction product was found only on the brush borders of proximal tubule cells; this reaction was ouabain-resistant, K-independent and partially Mg-dependent, but could be completely inhibited by l-tetramisole. Electron microscopy showed that a reaction was also present on the cytoplasmic surfaces of the lateral and basal plasma membranes of the proximal and distal tubule cells. That seen in the distal tubule was sensitive to ouabain but not to l-tetramisole, whereas that in the proximal tubule showed a mixture of ouabain-sensitive and l-tetramisole-sensitive components. It is concluded that the procedure as originally described is not specific, demonstrating alkaline phosphatase as well as Na-K-adenosine triphosphatase, but that this problem may be overcome by the use of an alkaline phosphatase inhibitor.

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