Measurement of Rhodamine 123 in Three-Dimensional Organoids: A Novel Model for P-Glycoprotein Inhibitor Screening

Basic Clin Pharmacol Toxicol. 2016 Oct;119(4):349-52. doi: 10.1111/bcpt.12596. Epub 2016 May 27.

Abstract

P-glycoprotein (P-gp), as the most important efflux transporter in intestines, plays the key role to determine the bioavailability of many drugs. The three-dimensional (3D) organoid model is suitable to imitate small intestinal epithelium. In this study, a rapid, sensitive and efficient method to measure rhodamine 123 (Rh123, P-gp substrate) in 3D organoids was developed to analyse P-gp-mediated drug transport. Ultrasonic cell disruptor was used to smash the organoid, and automatic microplate reader was used for detecting the concentration of Rh123 (λex /λem = 485/520 nm). Moreover, verapamil, quinidine and mitotane were used to make validation about this newly developed approach. All three P-gp inhibitors significantly inhibited the transport of Rh123 into 3D organoids. Therefore, the above-mentioned method could serve as a new model for P-gp inhibitor screening in a high-throughput way.

Publication types

  • Validation Study

MeSH terms

  • ATP Binding Cassette Transporter, Subfamily B / antagonists & inhibitors*
  • ATP Binding Cassette Transporter, Subfamily B / genetics
  • ATP Binding Cassette Transporter, Subfamily B / metabolism
  • Animals
  • Automation, Laboratory
  • Biological Transport / drug effects
  • Enterocytes / cytology
  • Enterocytes / metabolism*
  • Enterocytes / radiation effects
  • Fluorescent Dyes / metabolism*
  • High-Throughput Screening Assays
  • Immunohistochemistry
  • Intestinal Mucosa / cytology
  • Intestinal Mucosa / metabolism
  • Intestine, Small / cytology
  • Intestine, Small / metabolism
  • Kinetics
  • Membrane Transport Modulators / pharmacology*
  • Mice, Inbred C57BL
  • Models, Biological*
  • Organoids / cytology
  • Organoids / metabolism*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Rhodamine 123 / metabolism*
  • Spectrometry, Fluorescence
  • Tissue Culture Techniques
  • Ultrasonic Waves

Substances

  • ATP Binding Cassette Transporter, Subfamily B
  • Fluorescent Dyes
  • Membrane Transport Modulators
  • Rhodamine 123
  • multidrug resistance protein 3