Using Multi-fluorinated Bile Acids and In Vivo Magnetic Resonance Imaging to Measure Bile Acid Transport

Summary

Tools to diagnose bile acid malabsorption and measure bile acid transport in vivo are limited. An innovative approach in live animals is described that utilizes combined proton (¹H) plus fluorine (¹⁹F) magnetic resonance imaging; this novel methodology has translational potential to screen for bile acid malabsorption in clinical practice.

Abstract

Along with their traditional role as detergents that facilitate fat absorption, emerging literature indicates that bile acids are potent signaling molecules that affect multiple organs; they modulate gut motility and hormone production, and alter vascular tone, glucose metabolism, lipid metabolism, and energy utilization. Changes in fecal bile acids may alter the gut microbiome and promote colon pathology including cholerrheic diarrhea and colon cancer. Key regulators of fecal bile acid composition are the small intestinal Apical Sodium-dependent Bile Acid Transporter (ASBT) and fibroblast growth factor-19 (FGF19). Reduced expression and function of ASBT decreases intestinal bile acid up-take. Moreover, in vitro data suggest that some FDA-approved drugs inhibit ASBT function. Deficient FGF19 release increases hepatic bile acid synthesis and release into the intestines to levels that overwhelm ASBT. Either ASBT dysfunction or FGF19 deficiency increases fecal bile acids and may cause chronic diarrhea and promote colon neoplasia. Regrettably, tools to measure bile acid malabsorption and the actions of drugs on bile acid transport in vivo are limited. To understand the complex actions of bile acids, techniques are required that permit simultaneous monitoring of bile acids in the gut and metabolic tissues. This led us to conceive an innovative method to measure bile acid transport in live animals using a combination of proton (¹H) and fluorine (¹⁹F) magnetic resonance imaging (MRI). Novel tracers for fluorine (¹⁹F)-based live animal MRI were created and tested, both in vitro and in vivo. Strengths of this approach include the lack of exposure to ionizing radiation and translational potential for clinical research and practice.

Introduction

Along with their classical role as detergents that facilitate fat absorption from the gut, bile acids have emerged as potent signaling molecules affecting multiple organs in addition to those associated with their enterohepatic circulation^1,2. In addition to controlling their own metabolism, bile acids modulate several aspects of gastrointestinal physiology (e.g., gut motility and incretin hormone production, colon physiology, and cancer susceptibility) and have systemic effects on vascular tone, glucose and lipid metabolism, and energy utilization. While some of these effects are mediated in the gut, others are due to postprandial changes in syste....

Protocol

The following protocol adheres to guidelines approved by the Institutional Animal Care and Use Committee (IACUC) at the University of Maryland School of Medicine (IACUC Protocol #0415011, approved June 18, 2015).

1. Gavaging Mice with ¹⁹F-Labeled Bile Acids

1. Gavage mice with 150 mg/kg body weight ¹⁹F-labeled bile acids. Fill a 1-ml syringe to the necessary volume with ¹⁹F-labeled bile acid stock solution [cholic acid-trifluoro-acetyl lysine (CA-lys-TFA; in 1:1 polyethylene glycol 400:Dulbecco's phosphate buffered saline) or cholylsarcosine-trifluoro-N-methyl-acetamide (CA-sar-TFMA; in 60% polyethylene glycol 400 and 4....

Results

The use of MFBA for in vivo MRI to 'see' bile acid transport in real time has great potential for both research and clinical use. Moreover, the methods described here for resection of the gallbladder and biochemical analysis of its contents using liquid chromatography and mass spectrometry provide a means of confirming imaging results. However, the validity of these methods requires precise dosing, timing of assays, and localization of the gallbladder for imaging or surgical remo.......

Discussion

The synthesis of CA-lys-TFA and CA-sar-TFMA and the in vitro analysis of their transport using stably transfected Madin-Darby canine kidney cells expressing ASBT and human embryonic kidney cells expressing the sodium/taurocholate co-transporting polypeptide (NTCP) are detailed elsewhere^13,14. Here, the focus is on oral administration of MFBA by gavage to live animals, followed by harvest of the gallbladder, liver, and blood for analysis of MFBA content, and, notably, imaging MFBA in the gallbladder by.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Duall size-21 all glass tissue grinder	Kimble Chase Life Science, Vineland, NJ	885351-0022
Bruker BioSpec 70/30USR Avance III 7T horizontal bore MR Scanner	Bruker Biospin MRI GmbH, Germany		Use companion Paravision Version 5.1 software (see step 3.5)
Bruker 40 mm 19F/1H dual-tuned linerar volume coil	Bruker Biospin MRI GmbH, Germany		Use companion Paravision Version 5.1 software (see step 3.5)
Waters Acquity UPLC System with Quadrupole Detector	Waters Corporation, Milford, MA
Waters Acquity UPLC ethylene bridged hybrid C8 1.7 μm 2.1 x 50 mm column	Waters Corporation, Milford, MA
Gavage Needle	Braintree Scientific, INC.	N-010	20 G-1.5" curved 2.25 mm ball
2 Stainless Steel Hemostats	VWR	10755-018	4 and 5 inch, straight
Ketamine	MWI Veterinary Supply	501090	Ketamin zetamine 100 mg/ml
Xylazine	Akorn, Inc.		20 mg/ml
Intraperitoneal Catheter	Abbott	AbbocathTM-T.I.V. G720-A01 4535-42	24-G x 0.75"