Porcine Bile Duct Research Articles

Tu1664 Cholangioscopy With Image-Guided Intervention in the Human Bile Duct Using Scanning Fiber Endoscope System (Sfe): Results of First Clinical Experience

Currently available endoscopic imaging devices used in the biliary tree are low resolution with poor image quality, leading to inadequate evaluation of biliary tract disorders and limiting image guided therapy. However, a new ultrathin and flexible cholangioscope system has been developed and successfully demonstrated in a porcine bile duct. This mini-cholangioscope system uses a scanning fiber endoscope (SFE) as a forward-imaging guidewire, dimensions of 1.2-mm diameter and 3-m length. Full color video (500-line resolution at 30Hz) is the standard SFE imaging mode using spiral scanning of red, green, and blue laser light at low power.

Safety evaluation of self‐expanding metallic biliary stents eluting gemcitabine in a porcine model

Palliative biliary decompression by metal stent is the treatment of choice for unresectable malignant biliary obstruction; however, conventional stents provide only mechanical palliation and exert no anti-tumor effects. Gemcitabine (GEM) has been reported to be more effective in unresectable pancreatic cancer and biliary cancer compared with other chemotherapeutic drugs. We evaluated the safety of a GEM-eluting stent by analyzing histologic responses of the porcine bile duct. Stents containing GEM (0%, 10%, 15%, and 20% [w/v]) were surgically inserted into bile ducts of pigs (each group, n = 2). The animals were euthanized after 4 weeks, and the stented bile duct segment underwent gross and microscopic examination. Laboratory assay was performed for aspartate transaminase (AST), alanine transaminase (ALT), total bilirubin, and gamma-glutamyl transferase (γ-GTP). Moderate to severe inflammation was observed in the bile ducts in contact with stents containing 15 and 20% GEM, compared with no inflammation with 0% GEM and mild inflammation with 10% GEM. Fibrous reactions observed in the submucosal layer did not differ among groups. Transmural necrosis and perforations were not observed in any animal. No abnormal laboratory test findings were directly caused by GEM. Our newly developed GEM eluting stents can be used safely in normal bile ducts. Our results indicated that 10% GEM produced mild histologic changes in the stented segment and adjacent tissue; this concentration may be appropriate for clinical application.

Culture of porcine hepatocytes or bile duct epithelial cells by inductive serum-free media

A serum-free, feeder cell-dependent, selective culture system for the long-term culture of porcine hepatocytes or cholangiocytes was developed. Liver cells were isolated from 1-wk-old pigs or young adult pigs (25 and 63 kg live weight) and were placed in primary culture on feeder cell layers of mitotically blocked mouse fibroblasts. In serum-free medium containing 1% DMSO and 1 μM dexamethasone, confluent monolayers of hepatocytes formed and could be maintained for several wk. Light and electron microscopic analysis showed hepatocytes with in vivo-like morphology, and many hepatocytes were sandwiched between the feeder cells. When isolated liver cells were cultured in medium without dexamethasone but with 0.5% DMSO, monolayers of cholangioctyes formed that subsequently self-organized into networks of multicellular ductal structures, and whose cells had monocilia projecting into the lumen of the duct. Gamma-glutamyl transpeptidase (GGT) was expressed by the cholangiocytes at their apical membranes, i.e., at the inner surface of the ducts. Cellular GGT activity increased concomitantly with the development of ductal structures. Cytochrome P-450 was determined in microsomes following addition of metyrapone to the cultures. In vivo-like levels of P-450s were found in hepatocyte monolayers while levels of P-450 were markedly reduced in cholangiocyte monolayers. Serum protein secretion in conditioned media was analyzed by Western blot and indicated that albumin, transferrin, and haptoglobin levels were maintained in hepatocytes while albumin and haptoglobin declined over time in cholangiocytes. Quantitative RT-PCR analysis showed that serum protein mRNA levels were significantly elevated in the hepatocytes monolayers in comparison to the bile ductule-containing monolayers. Further, mRNAs specific to cholangiocyte differentiation and function were significantly elevated in bile ductule monolayers in comparison to hepatocyte monolayers. The results demonstrate an in vitro model for the study of either porcine hepatocytes or cholangiocytes with in vivo-like morphology and function.

Holmium: YAG Laser Safety Data On Bile Duct Epithelium in the Porcine Model

Background: Laser therapy has been increasing used for gastrointestinal and urological applications. Due to a very short wavelength, holmium: YAG can generate high-energy focused shock waves while reducing surrounding thermal injury. This characteristic has led to recent interest in using this technology for the fragmentation of large bile duct stones that have been refractory to conventional endoscopic therapy. Aim: The aim of this study was to establish safety data for holmium: YAG laser lithotripsy within the bile duct in the porcine model. Methods: 6 female 50 kg domestic pigs underwent necropsy and the bile ducts were explanted. The bile ducts were excised longitudinally exposing the epithelial surface and submerged under tension in liquid media. A Slimline holmium YAG laser probe (Lumenis, Sunnyvale, CA) was placed in perpendicular contact with the bile duct epithelium and activated at various energy density settings to evaluate perforation rates. The power density settings ranged from 8 to 15 watts (W) and were accomplished using various combinations of frequencies (Hz) and joules (J). The laser was activated until perforation of the bile duct occurred or for a maximum of 60 seconds. For each setting the probe was placed in similar locations on both the proximal and distal aspects of the duct. Both location and wattage were randomized to avoid physiologic differences that may affect outcomes. The median perforation times at each power setting were then compared using the Kruskal-Wallis One Analysis of Variance. Results: 104 data points were collected from 6 porcine bile ducts using 6 different power settings: 8 W (n=22); 9 W (n=22); 10 W (n=16); 12 W (n=20); 14 W (n=8); 15 W (n=16). The median perforation times were: 8 W (60 s); 9 W (24.5 s); 10 W (28 s); 12 W (24.5 s); 14 W (14 s); 15 W (5 s). Pairwise multiple comparison (Dunn's Method) revealed statistically significant differences (P<0.05) in the perforation times between the following settings: 8 W vs 15 W; 9 W vs 15 W; 10 W vs 15 W; 12 W vs 15 W. Conclusions: Holmium: YAG laser appears to be safe when used in short bursts (< 5 sec) at power settings ranging from 8 W to 12 W. Further studies are required to determine the ideal power settings balancing effective stone fragmentation and perforation risk.

The effect on porcine bile duct of a metallic stent covered with a paclitaxel-incorporated membrane

Biliary metallic stents are covered with a membrane to prevent tumor ingrowth and to prolong patency. The only function of these stents is to promote biliary drainage; they have no antitumor effect. A metallic stent was developed that is covered with a paclitaxel-incorporated membrane. The metallic stents were coated with one of 3 concentrations of paclitaxel (0, 10, and 20 % wt/v) and polyurethane. A stent with each concentration was surgically inserted in the bile duct of two pigs. Four weeks after insertion, the segment of bile duct containing the stent was examined histologically. To determine the efficacy of the drug release, stents were placed in phosphate buffered saline solution for 6 weeks, and the amount of paclitaxel released was measured by high-performance liquid chromatography. The histologic changes in the pig biliary epithelium were acceptable with respect to safety and included inflammatory cell infiltration and fibrous reactions. The changes corresponded to the amount of paclitaxel incorporated within the stent in contact with the bile duct. Epithelial denudation, mucin hypersecretion, and epithelial metaplasia were noted in the bile ducts that were in contact with stents containing 20 % wt/v paclitaxel. Transmural necrosis and perforation were not observed in any animal. In the in vitro experiment, the amounts of paclitaxel released over 1 week and over 6 weeks were similar, regardless of the concentration of paclitaxel incorporated in the stent. The stent with 10% (wt/v) paclitaxel in the covering membrane was found to be better than that with 20 % (wt/v) with respect to histologic changes and the effectiveness of drug release. A paclitaxel-incorporated metallic stent could serve as a basis for the development of a new and safe treatment modality for malignant biliary obstruction. Clinical trials of this stent with other adjuvant therapy are warranted.

The nuclear receptor SHP mediates inhibition of hepatic stellate cells by FXR and protects against liver fibrosis

The farnesoid X receptor (FXR) is an endogenous sensor for bile acids and inhibits bile acid synthesis by inducing small heterodimer partner (SHP) gene expression. The aim of this study was to investigate whether FXR is expressed by and modulates function of hepatic stellate cells (HSCs). The antifibrotic activity of FXR ligand was tested in 2 rodent models: the porcine serum and bile duct ligation (BDL). Twelve-week administration of 1-10 mg/kg 6-ethyl chenodeoxycholic acid (6-ECDCA), a synthetic FXR ligand, to porcine serum-treated rats prevented liver fibrosis development and reduced liver expression of alpha1(I) collagen, TGF-beta1 and alpha-SMA mRNA by approximately 90%. Therapeutic administration of 6-ECDCA, 3 mg/kg, to BDL rats reduced liver fibrosis and alpha1(I) collagen, transforming growth factor (TGF)-beta1, alpha-SMA, and tissue metalloproteinase inhibitor (TIMP)-1 and 2 messenger RNA (mRNA) by 70%-80%. No protection was observed in BDL rats treated with CDCA, 3 mg/kg, and ursodeoxycholic acid, 15 mg/kg. FXR expression was detected in HSCs. Exposure of HSCs to FXR ligands caused a 3-fold increase of SHP, reduced alpha1(I)collagen and TGF-beta1 by approximately 60%-70% and abrogates alpha1(I) collagen mRNA up-regulation induced by thrombin and TGF-beta1. By retrovirus infection and small interference RNA, we generated SHP overexpressing and SHP-deficient HSC-T6. Using these cell lines, we demonstrated that SHP binds JunD and inhibits DNA binding of adaptor protein (AP)-1 induced by thrombin. By demonstrating that an FXR-SHP regulatory cascade promotes resolution of liver fibrosis, this study establish that FXR ligands might represent a novel therapeutic option to treat liver fibrosis.

Mechanical properties of the porcine bile duct wall.

Background and AimThe function of the common bile duct is to transport bile from the liver and the gall bladder to the duodenum. Since the bile duct is a distensible tube consisting mainly of connective tissue, it is important to obtain data on the passive mechanical wall properties. The aims of this study were to study morphometric and biomechanical wall properties during distension of the bile duct.MethodsTen normal porcine common bile ducts were examined in vitro. A computer-controlled volume ramp infusion system with concomitant pressure recordings was constructed. A video camera provided simultaneous measurement of outer dimensions of the common bile duct. Wall stresses and strains were computed.ResultsThe common bile duct length increased by 25% from 24.4 ± 1.8 mm at zero pressure to 30.5 ± 2.0 mm at 5 kPa (p < 0.01). The diameter increased less than 10% in the same pressure range from 8.6 ± 0.4 mm to 9.3 ± 0.4 mm (p < 0.01). The stress-strain relations showed an exponential behavior with a good fit to the equation: σ = α . (exp(βε) - 1). The circumferential stress-strain curve was shifted to the left when compared to the longitudinal stress-strain curve, i.e. the linear constants (α values) were different (p < 0.01) whereas the exponential constants (β values) did not differ (p > 0.5).ConclusionThe porcine bile duct exhibited nonlinear anisotropic mechanical properties.

Luminal cross‐sectional area and tension–strain relation of the porcine bile duct

The bile duct is a distensible tube serving to transport bile from the liver and gallbladder to the duodenum. The purpose of this study was to characterize the luminal cross-sectional area (CSA) and tension-strain properties during distension of the normal isolated porcine common bile duct in vitro. An impedance planimetric system located inside a balloon was used. Eleven porcine bile ducts were examined in two locations, in the hepatic duct and in the common bile duct. The CSAs obtained in the common bile duct were significantly higher than those in the hepatic duct in the pressure range 0-8 kPa (P < 0.001). The circumferential wall tension (T)-strain (epsilon) relations for both locations fitted to the exponential equation T = a.eb. epsilon with determination coefficients of 0.97 +/- 0.01. The a and b constants were not statistically different between the two locations indicating that the elastic properties did not differ. In conclusion, the luminal CSAs were larger in the common bile duct when compared to the hepatic duct and the tension-strain relations did not differ between the two segments of the bile duct.

Division of donor liver for successful split-liver transplantation in pigs.

Split Liver Transplantation (SLT) is an attractive method to solve the problem of a shortage of liver grafts. A through knowledge of the anatomy of the porcine liver vessels and bile duct is essential in performing the experimental SLT. This study was undertaken to decide the split line for successful SLT in pigs by examining the main branching patterns both vessels and bile duct in 30 porcine livers macroscopically and angiographically. The hepatic arterial branching patterns were divided into three types and bile duct patterns into two types. There was no exception in branching patterns of the portal vein and the hepatic vein. We conclude it is desirable that the donor liver should be divided into two grafts between the left medial lobe and quadrate lobe.

Experimental evaluation of the optimal in-vitro conditions for bipolar electrocoagulation in the bile duct.

Although there have been many reports concerning the therapeutic use and potential of laser and monopolar electrocoagulation therapy, little has been published on the effects of bipolar electrocoagulation on biliary tissue. Although it has been reported that endoscopic recanalization of malignant stent obstruction using bipolar electrocoagulation is an advantageous form of treatment, the optimal parameters for this type of treatment have not yet been determined. The purpose of this prospective study was therefore to evaluate, under in-vitro conditions, the maximum applicable energy required; the influence of probe angle and duration time on the extent of the coagulation achieved; and whether multiple intermittent short pulses show any significant differences in the extent of coagulation achieved, in comparison with the use of a single continuous pulse. In this in-vitro study, experiments were carried out in 84 common bile ducts from freshly slaughtered swine. The extension of tissue injury after electrocoagulation treatment was measured at 60 combinations of varying levels of power output, probe angles, duration times, and pulse patterns. All of the results were correlated with the diameter of the untreated and normal bile duct walls under in-vitro conditions. When energy was applied at levels up to a maximum of 16 J, the extension of tissue coagulation was about 58% of the untreated bile duct wall diameter. Energy settings exceeding 16 J involved the whole wall diameter in most cases, often including the periductal fatty tissue. Additional energy applications higher than 16 J, 32 J, and 100 J, showed undesirable tissue changes, such as an increasing diameter in the treated bile duct walls of up to 56.8% of the wall diameter of control specimens. The study evaluated the standard settings in an in-vitro model. The results indicate that protective and effective use of bipolar electrocoagulation on normal porcine bile duct tissue requires: the use of a low power setting (less than 16 J); the use of continuous short pulse patterns; placement of the probe tip at an obtuse angle (0 degree); and the use of short application times.