Characterization of inorganic phosphate transport in osteoclast-like cells

Ito, Mikiko; Matsuka, Naoko; Izuka, Michiyo; Haito, Sakiko; Sakai, Yuko; Nakamura, Rie; Segawa, Hiroko; Kuwahata, Masashi; Yamamoto, Hironori; Pike, Wesley J.; Miyamoto, Ken‐ichi

doi:10.1152/ajpcell.00412.2004

Cited by 30 publications

(35 citation statements)

References 40 publications

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“…Although PAA and AsO 4 3-are frequently described as inhibitors of Na + -dependent P i transport, they have previously been shown also to exert effects against Na + -independent systems [33]. In the present study, these competitive compounds were able to inhibit effectively the Na + -dependent component -albeit with low affinitywhereas a sizeable fraction of the Na + -independent component was resistant.…”

Section: Discussionmentioning

confidence: 44%

“…To initiate measurements of P i uptake, a known volume of chondrocyte suspension in a flux solution (37°C) was transferred into an Eppendorf tube which contained an equivalent volume of flux solution supplemented with a known concentration of sodium phosphate and approximately 2 µCi ml -1 of 33 [P]HPO 4 2-. For certain experiments, the uptake solution was also supplemented with putative inhibitors of P i uptake, at doses previously shown to be effective in other studies [33,34].…”

Section: Measurement Of P I Influxmentioning

confidence: 99%

“…For certain experiments, the uptake solution was also supplemented with putative inhibitors of P i uptake, at doses previously shown to be effective in other studies [33,34]. Immediately after addition of the chondrocyte suspension (at time 0) the tubes were mixed for approximately 2 s prior to incubation in a water bath at 37°C or, for certain experiments, on ice (4°C).…”

Section: Measurement Of P I Influxmentioning

confidence: 99%

“…Counts recorded at t=0 min were subtracted from values measured at all subsequent timepoints. Standard tubes containing known amounts of 33 [P]HPO 4 2-from each separate influx solution were prepared and subjected to scintillation counting. The concentration of P i taken up into chondrocytes was determined from an estimate of cell volume (assuming chondrocytes are spherical with a diameter of 10 µm).…”

Section: Measurement Of P I Influxmentioning

confidence: 99%

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Characterisation of Inorganic Phosphate Transport in Bovine Articular Chondrocytes

Solomon¹,

Wilkins²,

Meredith³

et al. 2007

Cell Physiol Biochem

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In mineralising tissues such as growth plate cartilage extracellular organelles derived from the chondrocyte membrane are present. These matrix vesicles (MV) possess membrane transporters that accumulate Ca²⁺ and inorganic phosphate (P_i), and initiate the formation of hydroxyapatite crystals. MV are also present in articular cartilage, and hydroxyapatite crystals are believed to promote cartilage degradation in osteoarthritic joints. In the present study, P_i transport pathways in isolated bovine articular chondrocytes have been characterised. P_i uptake was temperature-sensitive and could be resolved into Na⁺-dependent and Na⁺-independent components. The Na⁺-dependent component saturated at high concentrations of extracellular P_i, with a K_m for P_i of 0.17mM. In solutions lacking Na⁺, uptake did not fully saturate, implying that under these conditions carrier-mediated uptake is supplemented by a diffusive pathway. Both Na⁺-dependent and Na⁺-independent components were sensitive to the P_i transport inhibitors phosphonoacetate and arsenate, although a fraction of Na⁺-independent P_i uptake was resistant to these anions. Total P_i uptake was optimal at pH 7.4, and reduced as pH was made more acidic or more alkaline, an effect that represented reduced Na⁺-dependent influx. RT-PCR analysis confirmed that two members of the NaPi III family, Pit-1 and Pit-2, are expressed, but that NaPi II transporters are not.

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Section: Discussionmentioning

confidence: 44%

Section: Measurement Of P I Influxmentioning

confidence: 99%

Section: Measurement Of P I Influxmentioning

confidence: 99%

Section: Measurement Of P I Influxmentioning

confidence: 99%

See 2 more Smart Citations

Characterisation of Inorganic Phosphate Transport in Bovine Articular Chondrocytes

Solomon¹,

Wilkins²,

Meredith³

et al. 2007

Cell Physiol Biochem

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“…The cells were exposed to FBS-free assay medium containing 14 C mannitol in the apical or the basolateral compartments and to the FBS-free assay medium containing 0.25 mM unlabeled mannitol in the opposite compartment (2). P i transport was measured using a previously described procedure (18) and was calculated as nanomoles of 32 P per milligram of protein taken up by the cells over 6 min. The experiments were performed in triplicate.…”

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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Phosphate: Known and potential roles during development and regeneration of teeth and supporting structures

Foster

Tompkins

Rutherford

et al. 2008

Birth Defects Research Pt C

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Inorganic phosphate (Pi) is abundant in cells and tissues as an important component of nucleic acids and phospholipids, a source of high-energy bonds in nucleoside triphosphates, a substrate for kinases and phosphatases, and a regulator of intracellular signaling. The majority of the body’s Pi exists in the mineralized matrix of bones and teeth. Systemic Pi metabolism is regulated by a cast of hormones, phosphatonins, and other factors via the bone-kidney-intestine axis. Mineralization in bones and teeth is in turn affected by homeostasis of Pi and inorganic pyrophosphate (PPi), with further regulation of the Pi/PPi ratio by cellular enzymes and transporters. Much has been learned by analyzing the molecular basis for changes in mineralized tissue development in mutant and knock-out mice with altered Pi metabolism. This review focuses on factors regulating systemic and local Pi homeostasis and their known and putative effects on the hard tissues of the oral cavity. By understanding the role of Pi metabolism in the development and maintenance of the oral mineralized tissues, it will be possible to develop improved regenerative approaches.

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Characterization of inorganic phosphate transport in osteoclast-like cells

Cited by 30 publications

References 40 publications

Characterisation of Inorganic Phosphate Transport in Bovine Articular Chondrocytes

Characterisation of Inorganic Phosphate Transport in Bovine Articular Chondrocytes

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

Phosphate: Known and potential roles during development and regeneration of teeth and supporting structures

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Characterization of inorganic phosphate transport in osteoclast-like cells

Cited by 30 publications

References 40 publications

Characterisation of Inorganic Phosphate Transport in Bovine Articular Chondrocytes

Characterisation of Inorganic Phosphate Transport in Bovine Articular Chondrocytes

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

Phosphate: Known and potential roles during development and regeneration of teeth and supporting structures

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