Induction of Surgical Obstructive Cholestasis in rats: morphological, biochemical and immunohistochemical changes

Document Type : Original Articles

Authors

Department of Surgery, Anesthesiology, and Radiology, Faculty of Veterinary, Medicine, Mansoura University, 35516 Mansoura, Egypt

Abstract

Objective: To detail an easy approach for surgical induction of obstructive cholestasis in a rat model. Additionally, to assess the role of serum and tissue biomarkers for determination of the extent of cholestasis induced hepatocellular injury and to quantify the apoptotic liver cell using fluorescein-labeled Annexin V.
Design: Randomized controlled experimental study.
Animals: Sixty-five healthy Male Sprague- Dawley rats weighing 268.18 ± 23.3 grams.
Procedures: The rats were randomly allocated into two main groups: 1) sham group (SO; n= 15) and 2) bile duct ligation group (BL; n= 50). Three SO rats and 8 BL rats were euthanized at 10, 20, 30, 45 and 60 post-surgery. Blood and liver samples were collected for serum and tissue biochemical analysis. Liver apoptosis was quantified using Annexin V, meanwhile liver fibrosis was quantified using both TNF-β and histological examination.
Results: There was a tendency towards less complications and higher surgeon satisfaction when the common bile duct was approached by the stomach approach. Serum biochemical analysis revealed a significant time-dependent decreases in hepatic serum biomarkers (alanine aminotransferease, aspartate aminotransferase, serum glutamic oxaloacetic transaminase, alkaline phosphatase, gamma-glutamyltransferase, serum albumin, the concentration of total bilirubin, and direct bilirubin) after induced obstructive cholestasis (P = 0.01-0.001), whereas AST: ALT ratio conversely was elevated (p < 0.05 value). Flow cytometer analysis revealed time-dependent significant increase in the percent of apoptotic cells after BL (P < 0.001) and consequently the state of oxidative stress (malondialdehyde) was (P < 0.001). The antioxidant tissue markers (superoxide dismutase, Catalase and glutathione reductase) were significantly decreased (P < 0.05), meanwhile TNF-β, the fibrosis marker showed time dependent significant increases (P = 0.001).
Conclusion and clinical relevance: Stomach approach provided an easy, fast and reliable procedure for induction of obstructive cholestasis in rats. Liver Annexin V and TGF-β were diagnostic tools in further apoptosis and fibrosis.

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