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Characterization of the Aquaporin-9 Inhibitor RG100204 In Vitro and in db/db Mice

Florio, Marilina ; Engfors, Angelica LU ; Gena, Patrizia ; Larsson, Jessica ; Massaro, Alessandro ; Timpka, Stella ; Kvist Reimer, Martina LU ; Kjellbom, Per LU ; Beitz, Eric and Johanson, Urban LU orcid , et al. (2022) In Cells 11(19).
Abstract

Aquaporin-9 (AQP9) is a facilitator of glycerol and other small neutral solute transmembrane diffusion. Identification of specific inhibitors for aquaporin family proteins has been difficult, due to high sequence similarity between the 13 human isoforms, and due to the limited channel surface areas that permit inhibitor binding. The few AQP9 inhibitor molecules described to date were not suitable for in vivo experiments. We now describe the characterization of a new small molecule AQP9 inhibitor, RG100204 in cell-based calcein-quenching assays, and by stopped-flow light-scattering recordings of AQP9 permeability in proteoliposomes. Moreover, we investigated the effects of RG100204 on glycerol metabolism in mice. In cell-based assays,... (More)

Aquaporin-9 (AQP9) is a facilitator of glycerol and other small neutral solute transmembrane diffusion. Identification of specific inhibitors for aquaporin family proteins has been difficult, due to high sequence similarity between the 13 human isoforms, and due to the limited channel surface areas that permit inhibitor binding. The few AQP9 inhibitor molecules described to date were not suitable for in vivo experiments. We now describe the characterization of a new small molecule AQP9 inhibitor, RG100204 in cell-based calcein-quenching assays, and by stopped-flow light-scattering recordings of AQP9 permeability in proteoliposomes. Moreover, we investigated the effects of RG100204 on glycerol metabolism in mice. In cell-based assays, RG100204 blocked AQP9 water permeability and glycerol permeability with similar, high potency (~5 × 10 -8 M). AQP9 channel blocking by RG100204 was confirmed in proteoliposomes. After oral gavage of db/db mice with RG100204, a dose-dependent elevation of plasma glycerol was observed. A blood glucose-lowering effect was not statistically significant. These experiments establish RG100204 as a direct blocker of the AQP9 channel, and suggest its use as an experimental tool for in vivo experiments on AQP9 function.

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organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
membrane transport, aquaglyceroporins, metabolic homeostasis, energy balance, glycerol metabolism, gluconeogenesis, calcein quenching, Pichia pastoris, proteoliposomes, stopped-flow light scattering
in
Cells
volume
11
issue
19
article number
3118
pages
11 pages
publisher
MDPI AG
external identifiers
  • pmid:36231080
  • scopus:85139751545
ISSN
2073-4409
DOI
10.3390/cells11193118
language
English
LU publication?
yes
id
83681040-4b63-4c80-9ce2-b6a9d8dd42e4
date added to LUP
2022-10-17 08:31:21
date last changed
2024-06-09 16:03:11
@article{83681040-4b63-4c80-9ce2-b6a9d8dd42e4,
  abstract     = {{<p>Aquaporin-9 (AQP9) is a facilitator of glycerol and other small neutral solute transmembrane diffusion. Identification of specific inhibitors for aquaporin family proteins has been difficult, due to high sequence similarity between the 13 human isoforms, and due to the limited channel surface areas that permit inhibitor binding. The few AQP9 inhibitor molecules described to date were not suitable for in vivo experiments. We now describe the characterization of a new small molecule AQP9 inhibitor, RG100204 in cell-based calcein-quenching assays, and by stopped-flow light-scattering recordings of AQP9 permeability in proteoliposomes. Moreover, we investigated the effects of RG100204 on glycerol metabolism in mice. In cell-based assays, RG100204 blocked AQP9 water permeability and glycerol permeability with similar, high potency (~5 × 10 <sup>-8</sup> M). AQP9 channel blocking by RG100204 was confirmed in proteoliposomes. After oral gavage of <i> db/db</i> mice with RG100204, a dose-dependent elevation of plasma glycerol was observed. A blood glucose-lowering effect was not statistically significant. These experiments establish RG100204 as a direct blocker of the AQP9 channel, and suggest its use as an experimental tool for in vivo experiments on AQP9 function. </p>}},
  author       = {{Florio, Marilina and Engfors, Angelica and Gena, Patrizia and Larsson, Jessica and Massaro, Alessandro and Timpka, Stella and Kvist Reimer, Martina and Kjellbom, Per and Beitz, Eric and Johanson, Urban and Rützler, Michael and Calamita, Giuseppe}},
  issn         = {{2073-4409}},
  keywords     = {{membrane transport; aquaglyceroporins; metabolic homeostasis; energy balance; glycerol metabolism; gluconeogenesis; calcein quenching; Pichia pastoris; proteoliposomes; stopped-flow light scattering}},
  language     = {{eng}},
  month        = {{10}},
  number       = {{19}},
  publisher    = {{MDPI AG}},
  series       = {{Cells}},
  title        = {{Characterization of the Aquaporin-9 Inhibitor RG100204 In Vitro and in <i>db/db</i> Mice}},
  url          = {{http://dx.doi.org/10.3390/cells11193118}},
  doi          = {{10.3390/cells11193118}},
  volume       = {{11}},
  year         = {{2022}},
}