Isolated proximal tubular cells from rat kidney as an in vitro model for studies on nephrotoxicity

1989 ◽  
Vol 101 (1) ◽  
pp. 144-157 ◽  
Author(s):  
Pieter J. Boogaard ◽  
Gerard J. Mulder ◽  
J.Fred Nagelkerke
Keyword(s):  
In Vitro Model ◽  
Rat Kidney ◽  
Proximal Tubular Cells ◽  
Tubular Cells ◽  
Proximal Tubular ◽  
Vitro Model

Primary culture of proximal tubular cells from normal rat kidney as an in vitro model to study mechanisms of nephrotoxicity

1990 ◽  
Vol 39 (8) ◽  
pp. 1335-1345 ◽  
Author(s):  
Pieter J. Boogaard ◽  
J.Paul Zoeteweij ◽  
Theo J.C. Van Berkel ◽  
Joost M. Van't Noordende ◽  
Gerard J. Mulder ◽  
...  
Keyword(s):  
Primary Culture ◽  
In Vitro Model ◽  
Rat Kidney ◽  
Proximal Tubular Cells ◽  
Tubular Cells ◽  
Proximal Tubular ◽  
Normal Rat ◽  
Vitro Model

scholarly journals A Non-Targeted Capillary Electrophoresis-Mass Spectrometry Strategy to Study Metabolic Differences in an In Vitro Model of High-Glucose Induced Changes in Human Proximal Tubular HK-2 Cells

Molecules ◽  
2020 ◽  
Vol 25 (3) ◽  
pp. 512 ◽  
Author(s):  
Samuel Bernardo-Bermejo ◽  
Elena Sánchez-López ◽  
María Castro-Puyana ◽  
Selma Benito-Martínez ◽  
Francisco Javier Lucio-Cazaña ◽  
...  
Keyword(s):  
Diabetic Nephropathy ◽  
High Glucose ◽  
In Vitro Model ◽  
Proximal Tubular Cells ◽  
Extracellular Medium ◽  
Metabolic Alterations ◽  
Tubular Cells ◽  
Proximal Tubular ◽  
Vitro Model

Diabetic nephropathy is characterized by the chronic loss of kidney function due to high glucose renal levels. HK-2 proximal tubular cells are good candidates to study this disease. The aim of this work was to study an in vitro model of high glucose-induced metabolic alterations in HK-2 cells to contribute to the pathogenesis of this diabetic complication. An untargeted metabolomics strategy based on CE-MS was developed to find metabolites affected under high glucose conditions. Intracellular and extracellular fluids from HK-2 cells treated with 25 mM glucose (high glucose group), with 5.5 mM glucose (normal glucose group), and with 5.5 mM glucose and 19.5 mM mannitol (osmotic control group) were analyzed. The main changes induced by high glucose were found in the extracellular medium where increased levels of four amino acids were detected. Three of them (alanine, proline, and glutamic acid) were exported from HK-2 cells to the extracellular medium. Other affected metabolites include Amadori products and cysteine, which are more likely cause and consequence, respectively, of the oxidative stress induced by high glucose in HK-2 cells. The developed CE-MS platform provides valuable insight into high glucose-induced metabolic alterations in proximal tubular cells and allows identifying discriminative molecules of diabetic nephropathy.

Isolation and Culture of Proximal Tubular Cells from Porcine Kidney

1993 ◽  
Vol 21 (4) ◽  
pp. 457-465
Author(s):  
Marieke Kruidering ◽  
Frans A. Prins ◽  
Emile de Heer ◽  
Gerard J. Mulder ◽  
J. Fred Nagelkerke
Keyword(s):  
Primary Culture ◽  
Single Cells ◽  
Proximal Tubular Cells ◽  
Isolated Cells ◽  
Serum Free ◽  
Tubular Cells ◽  
Proximal Tubular ◽  
Vitro Model ◽  
Serum Free Conditions

Porcine proximal tubular cells (PPTC) were isolated from kidneys obtained from slaughterhouse pigs. After disruption of the connective tissue by collagenase, purification was achieved by filtration and centrifugation on a discontinuous density gradient. Single cells and clusters of 10–40 cells were obtained, having a viability of 93–99%. More than 81% of the single cells showed γ-glutamyltranspeptidase (GGT) activity and more than 95% showed non-specific esterase (NE) activity, marker enzymes for proximal tubule cells. One kidney yielded 1 x 107 single cells and 3x107 cells in clusters. Cells were kept in primary culture on plastic or collagen-coated dishes. In the presence of 10% serum, confluency was reached within four days. The monolayers could be kept in culture for four days after confluency, in serum-free conditions. When seeded in serum-free conditions, PPTC did not reach confluency, but the cells could be kept in culture for at least 16 days. The cells displayed epithelial morphology, i.e. cobblestone shape, dome formation, microvilli, basal infoldings and abundant mitochondria. PPTC in primary culture still displayed NE activity, while 80% of the cells showed GGT activity. In conclusion, the isolated cells are of proximal tubular origin, reach confluency in 3–4 days in the presence of 10% serum, and can be kept as monolayers in serum-free conditions for four additional days and may provide a suitable in vitro model for long-term nephrotoxicity studies.

Inhibition by insulin of cellular autophagy in proximal tubular cells of rat kidney

1983 ◽  
Vol 244 (2) ◽  
pp. E109-E114 ◽  
Author(s):  
U. Pfeifer ◽  
M. Warmuth-Metz
Keyword(s):  
Volume Fraction ◽  
Rat Kidney ◽  
Insulin Dose ◽  
Ringer Solution ◽  
Numerical Density ◽  
Partial Recovery ◽  
Proximal Tubular Cells ◽  
Tubular Cells ◽  
Cellular Autophagy ◽  
Proximal Tubular

Adult male Sprague-Dawley rats were injected intraperitoneally with 5 U insulin/kg body wt (45 animals). As determined by quantitative electron microscopy, the volume fraction and the numerical density of autophagic vacuoles (AV) in proximal tubular cells decreased within 10 min by 46 and 26%, respectively. A partial recovery of the AV volume fraction was observed 20 and 30 min after the injection contrary to our previous findings with liver (J. Cell Biol. 78: 152-167, 1978). In an additional experiment (12 animals) it was shown that an insulin dose of 0.5 U but not of 0.05 U/kg body wt reduced the AV volume fraction to an extent similar to that of 5 U. To eliminate possible secondary effects, Ringer solution containing 0.8 microM insulin was dropped intravitally for 15 min to one pole of the decapsulated kidney and Ringer solution without additions to the other pole (8 animals). After intravital fixation, the AV volume fraction and numerical density in proximal tubular cells was found to be reduced under the influence of insulin by 22 and 36%, respectively. This data shows that insulin inhibits the process of cellular autophagy in proximal tubular cells of the kidney.

scholarly journals Palmitate aggravates proteinuria-induced cell death and inflammation via CD36-inflammasome axis in the proximal tubular cells of obese mice

2018 ◽  
Vol 315 (6) ◽  
pp. F1720-F1731 ◽  
Author(s):  
Lung-Chih Li ◽  
Jenq-Lin Yang ◽  
Wen-Chin Lee ◽  
Jin-Bor Chen ◽  
Chien-Te Lee ◽  
...  
Keyword(s):  
Cell Death ◽  
Tubular Injury ◽  
Proximal Tubular Cells ◽  
Tubular Cells ◽  
Renal Tubular Cells ◽  
Caspase 1 ◽  
Proximal Tubular ◽  
Renal Tubular ◽  
Nlrp3 Inflammasomes

High levels of serum free fatty acids (FFAs) and proteinuria have been implicated in the pathogenesis of obesity-related nephropathy. CD36, a class B scavenger receptor, is highly expressed in the renal proximal tubules and mediates FFA uptake. It is not clear whether FFA- and proteinuria-mediated CD36 activation coordinates NLRP3 inflammasomes to induce renal tubular injury and inflammation. In this study, we investigated the roles of CD36 and NLRP3 inflammasomes in FFA-induced renal injury in high-fat diet (HFD)-induced obesity. HFD-fed C57BL/6 mice and palmitate-treated HK2 renal tubular cells were used as in vivo and in vitro models. Immunohistochemical staining showed that CD36, IL-1β, and IL-18 levels increased progressively in the kidneys of HFD-fed mice. Sulfo- N-succinimidyl oleate (SSO), a CD36 inhibitor, attenuated the HFD-induced upregulation of NLRP3, IL-1β, and IL-18 and suppressed the colocalization of NLRP3 and ASC in renal tubular cells. In vitro, SSO abolished the palmitate-induced activation of IL-1β, IL-18, and caspase-1 in HK2 proximal tubular cells. Furthermore, treatment with SSO and the knockdown of caspase-1 expression by siRNA both inhibited palmitate-induced cell death and apoptosis in HK2 cells. Collectively, palmitate causes renal tubular inflammation, cell death, and apoptosis via the CD36/NLRP3/caspase-1 axis, which may explain, at least in part, the mechanism underlying FFA-related renal tubular injury. The blockade of CD36-induced cellular processes is therefore a promising strategy for treating obesity-related nephropathy.

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