The paracellular permeability of opossum kidney cells, a proximal tubule cell line

Liang, Mingyu; Ramsey, Carla R.; Knox, Franklyn G.

doi:10.1046/j.1523-1755.1999.00787.x

Cited by 20 publications

(19 citation statements)

References 17 publications

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“…Empty Transwells devoid of tissues exhibited much higher permeability, with an average A→B P app value of 308 × 10 −6 ± 3.6 × 10 −6 cm/s, supporting the conclusion that the 3D PT tissues themselves were responsible for the observed barrier function. When compared to literature findings, these results demonstrate that the barrier formed by the RPTEC cells in the 3D PT tissues is leakier than isolated epithelial monolayers but more characteristic of the barrier observed for the PT in vivo (Boulpaep and Seely, 1971; Liang et al, 1999). …”

Section: Resultssupporting

confidence: 59%

“…Monolayer cultures of renal epithelia such as HK-2 and primary RPTEC cells have been shown to exhibit high TEER values of 100–1,000 Ω * cm 2 , with a high degree of variability in those values attributed to differences in culture methods and variations between human kidney donors (Prozialeck et al, 2006). In contrast, in vivo tubules exhibit values between 6.6 and 11.6 Ω * cm 2 (Boulpaep and Seely, 1971; Liang et al, 1999). In this study, 3D PT tissues exhibited TEER values of 18.1 Ω * cm 2 (Table 3), which more closely matches values measured in vivo .…”

Section: Discussionmentioning

confidence: 99%

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3D Proximal Tubule Tissues Recapitulate Key Aspects of Renal Physiology to Enable Nephrotoxicity Testing

et al. 2017

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Due to its exposure to high concentrations of xenobiotics, the kidney proximal tubule is a primary site of nephrotoxicity and resulting attrition in the drug development pipeline. Current pre-clinical methods using 2D cell cultures and animal models are unable to fully recapitulate clinical drug responses due to limited in vitro functional lifespan, or species-specific differences. Using Organovo's proprietary 3D bioprinting platform, we have developed a fully cellular human in vitro model of the proximal tubule interstitial interface comprising renal fibroblasts, endothelial cells, and primary human renal proximal tubule epithelial cells to enable more accurate prediction of tissue-level clinical outcomes. Histological characterization demonstrated formation of extensive microvascular networks supported by endogenous extracellular matrix deposition. The epithelial cells of the 3D proximal tubule tissues demonstrated tight junction formation and expression of renal uptake and efflux transporters; the polarized localization and function of P-gp and SGLT2 were confirmed. Treatment of 3D proximal tubule tissues with the nephrotoxin cisplatin induced loss of tissue viability and epithelial cells in a dose-dependent fashion, and cimetidine rescued these effects, confirming the role of the OCT2 transporter in cisplatin-induced nephrotoxicity. The tissues also demonstrated a fibrotic response to TGFβ as assessed by an increase in gene expression associated with human fibrosis and histological verification of excess extracellular matrix deposition. Together, these results suggest that the bioprinted 3D proximal tubule model can serve as a test bed for the mechanistic assessment of human nephrotoxicity and the development of pathogenic states involving epithelial-interstitial interactions, making them an important adjunct to animal studies.

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Section: Resultssupporting

confidence: 59%

Section: Discussionmentioning

confidence: 99%

3D Proximal Tubule Tissues Recapitulate Key Aspects of Renal Physiology to Enable Nephrotoxicity Testing

et al. 2017

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“…The OK cell line has been extensively used for the study of P i transport, and mimics the function of renal proximal tubular epithelial cells in a variety of ways, including the formation of tight junctions (24). When cultured on permeable supports, OK cell sheets have been shown to have polarized P i transport (24).…”

Section: Methodsmentioning

confidence: 99%

An apical expression signal of the renal type IIc Na⁺-dependent phosphate cotransporter in renal epithelial cells

Ito

Sakurai²,

Hayashi³

et al. 2010

American Journal of Physiology-Renal Physiology

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ϩ -dependent phosphate cotransporter (NaPi-IIc) is specifically targeted to, and expressed on, the apical membrane of renal proximal tubular cells and mediates phosphate transport. In the present study, we investigated the signals that determine apical expression of NaPi-IIc with a focus on the role of the N-and the C-terminal tails of mouse NaPi-IIc in renal epithelial cells [opossum kidney (OK) and Madin-Darby canine kidney cells]. Wild-type NaPi-IIc, the cotransporter NaPi-IIa, as well as several IIa-IIc chimeras and deletion mutants, were fused to enhanced green fluorescent protein (EGFP), and their cellular localization was analyzed in polarized renal epithelial cells by confocal microscopy and by cell-surface biotinylation. Fluorescent EGFP-fused NaPi-IIc transporter proteins are correctly expressed in the apical membrane of OK cells. The apical expression of N-terminal deletion mutants (deletion of N-terminal 25, 50, or 69 amino acids) was not affected by truncation. In contrast, C-terminal deletion mutants (deletion of C-terminal 45, 50, or 62 amino acids) did not have correct apical expression. A more detailed mutational analysis indicated that a domain (amino acids WLHSL) in the cytoplasmic C terminus is required for apical expression of NaPi-IIc in renal epithelial cells. We conclude that targeting of NaPi-IIc to the apical cell surface is regulated by a unique amino acid motif in the cytoplasmic C-terminal domain.Npt2c; SLC34A3; localization; OK cell; hereditary hypophosphatemic rickets with hypercalciuria INORGANIC PHOSPHATE (P i ) reabsorption in renal proximal tubules is required for P i homeostasis in the body. Na ϩ -dependent P i transporters (NaPi) in the brush-border membrane (BBM) of proximal tubule cells mediate the rate-limiting step in the overall P i -reabsorption process (26,28). Type IIa (NaPiIIa) and type IIc (NaPi-IIc) NaPi cotransporters are expressed in the apical membrane of proximal tubule cells and mediate P i transport (27, 38). The extent of P i reabsorption in the proximal tubules is determined largely by the abundance of the NaPi-IIa cotransporter (11,28,29).Several groups have shown that hereditary hypophosphatemic rickets with hypercalciuria (HHRH), a Mendelian disorder of renal P i reabsorption (39, 42), results from the lack of a functional NaPi-IIc protein, leading to severe renal wasting with hypophosphatemia (5,16,19,22,25). These data suggest that the NaPi-IIc cotransporter plays an important role in renal P i reabsorption and may be a key determinant of plasma P i concentrations in humans. However, it is not clear why a loss of function of the less abundant and energetically less favorable electroneutral NaPi-IIc transporter causes hypophosphatemia/rickets and osteomalacia in humans, whereas mutations in the more abundant electrogenic NaPi-IIa elicits a mild skeletal phenotype (3).The plasma membrane of epithelial cells is divided into two separate membrane compartments, the apical and the basolateral domains (36,37). This polarity is maintained by intracellular mach...

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“…Thus either OK or NRK-52E cells are a preferred model of a low resistance epithelia. We chose to use OK cells because the opossum genome has been sequenced and this cell line demonstrates a very low TER (29). OK cells were derived from the proximal tubule, a very loose epithelium.…”

mentioning

confidence: 99%

Claudin-4 forms a paracellular barrier, revealing the interdependence of claudin expression in the loose epithelial cell culture model opossum kidney cells

Borovac

Barker

Rievaj

et al. 2012

American Journal of Physiology-Cell Physiology

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The paracellular permeability of opossum kidney cells, a proximal tubule cell line

Abstract: OK cell sheets cultured on Transwell supports possess a leaky paracellular pathway resembling that of the proximal tubule epithelium in vivo.

Cited by 20 publications

References 17 publications

3D Proximal Tubule Tissues Recapitulate Key Aspects of Renal Physiology to Enable Nephrotoxicity Testing

3D Proximal Tubule Tissues Recapitulate Key Aspects of Renal Physiology to Enable Nephrotoxicity Testing

An apical expression signal of the renal type IIc Na⁺-dependent phosphate cotransporter in renal epithelial cells

Claudin-4 forms a paracellular barrier, revealing the interdependence of claudin expression in the loose epithelial cell culture model opossum kidney cells

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The paracellular permeability of opossum kidney cells, a proximal tubule cell line

Abstract: OK cell sheets cultured on Transwell supports possess a leaky paracellular pathway resembling that of the proximal tubule epithelium in vivo.

Cited by 20 publications

References 17 publications

3D Proximal Tubule Tissues Recapitulate Key Aspects of Renal Physiology to Enable Nephrotoxicity Testing

3D Proximal Tubule Tissues Recapitulate Key Aspects of Renal Physiology to Enable Nephrotoxicity Testing

An apical expression signal of the renal type IIc Na+-dependent phosphate cotransporter in renal epithelial cells

Claudin-4 forms a paracellular barrier, revealing the interdependence of claudin expression in the loose epithelial cell culture model opossum kidney cells

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An apical expression signal of the renal type IIc Na⁺-dependent phosphate cotransporter in renal epithelial cells