Effect of Bile Pigments on the Compromised Gut Barrier Function in a Rat Model of Bile Duct Ligation

Abstract

BackgroundStudies have shown that the absence of bile in the gut lumen, either by bile duct ligation or bile diversion, induces mucosal injury. However, the mechanism remains elusive. In this study, the role of bile pigments in gut barrier function was investigated in a rat model of bile duct ligation.MethodsMale Sprague Dawley (SD) rats were used in this study. After ligation of bile duct, the animals were administrated with free bilirubin, bilirubin ditaurate, or biliverdin by intragastric gavage. 1, 2, or 3 days later, the animals were sacrificed and the damage of mucosa was assessed by histological staining as well as biochemical parameters such as changes of diamine oxidase (DAO) and D-lactate (D-Lac) in the blood. Trypsin and chymotrypsin of the gut were also measured to determine how these digestive proteases may relate to the observed effects of bile pigments.ResultsBile duct ligation (BDL) caused significant increases in gut trypsin and chymotrypsin along with damage of the mucosa as demonstrated by the histological findings under microscope, the reduced expression of tight junction molecules like occludin, and significant changes in DAO and D-lac in the blood. Free bilirubin but not bilirubin ditaurate or biliverdin showed significant inhibitions on trypsin and chymotrypsin as well as alleviated changes of histological and biochemical parameters related to gut barrier disruption.ConclusionBile may protect the gut from damage through inhibiting digestive proteases like trypsin and chymotrypsin by free bilirubin.

Unconjugated bilirubin ameliorates the inflammation and digestive protease increase in TNBS-induced colitis

“…As an important endogenous substance largely distributed in GI tract, the unconjugated bilirubin (UCB) from heme metabolism by the heme oxygenase-1 (HO-1) is an effective antioxidant ( 18 ). Our recent studies using bile duct ligated rats confirmed the critical role of unconjugated bilirubin in inactivation of digestive proteases and gut protection ( 19 , 20 ). Whereas, the specific effects of UCB on the inflammation in colitis, and the modifications of digestive proteases levels are still unrevealed.…”

“…TNBS group received colonic instillation of 1 ml of 50% ethanol in saline containing 25 mg TNBS, while the control group received 1 ml saline ( 22 , 23 ). After colonic instillation, the UCB treatment group received an intra-gastric gavages of 3.5 ml UCB (40 µM, UCB is dissolved in 0.4% dimethyl sulfoxide at concentrations up to 40 µM) ( 19 , 23 ), while the Control and TNBS groups received equal volume of saline solution. All animals were recorded daily for body weight and total feces were collected daily and stored at −4°C ( 24 ).…”

Role of bilirubin in digestive proteases inactivation in the lower intestine

“…They proposed that inactivation of digestive proteases by deconjugated bilirubin may be the evolutionary driving force for bilirubin or biliverdin predominance in animals [3]. Our recent study also showed increases in trypsin and chymotrypsin in small intestine in rats with bile duct ligation (BDL) [4]. Free bilirubin but not conjugated bilirubin or biliverdin showed an inhibitory effect on the increased pancreatic proteases as well as the compromised gut barrier function after BDL [4].…”

“…Our recent study also showed increases in trypsin and chymotrypsin in small intestine in rats with bile duct ligation (BDL) [4]. Free bilirubin but not conjugated bilirubin or biliverdin showed an inhibitory effect on the increased pancreatic proteases as well as the compromised gut barrier function after BDL [4]. In this study, using the BDL model, we further investigated how the absence of bile will affect pancreatic digestive proteases activity in the large intestine and how treatment with different kinds of bile pigments (free bilirubin, conjugated bilirubin and biliverdin) will affect those changes.…”

A Common Control Group - Optimising the Experiment Design to Maximise Sensitivity

“…Frequently, animal researchers conduct experiments that involve multiple treatments and a common control. For example, a survey of recent PLoS ONE papers identified an R&D drug study involving multiple different treatments versus a vehicle control [10] , a study comparing high cholesterol diets to a low cholesterol diet [11] and a study comparing responses at later time points to a baseline group [12] . This type of study design is also commonly used in toxicology and safety assessment where studies are typically performed so that they can compare increasing doses of a treatment back to a control group.…”