Effects of Paclitaxel-conjugated N-Succinyl-Hydroxyethyl Chitosan Film for Proliferative Cholangitis in Rabbit Biliary Stricture Model / 中华医学杂志(英文版)

<p><b>Background</b>Paclitaxel (PTX) could inhibit the growth of fibroblasts, which occurs in proliferative cholangitis and leads to biliary stricture. However, its use has been limited due to poor bioavailability and local administration for short time. This study designed and synthesized a new PTX-conjugated chitosan film (N-succinyl-hydroxyethyl chitosan containing PTX [PTX-SHEC]) and evaluated its safety and efficiency using in vivo and in vitro experiments.</p><p><b>Methods:</b>The SHEC conjugated with PTX was confirmed by nuclear magnetic resonance (NMR) and Fourier-transform infrared spectroscopy (FT-IR) measurements. Drug releases in vitro and in vivo were determined using high-performance liquid chromatography. Cell viability in vitro was measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide. Rabbit biliary stricture model was constructed. All rabbits randomly divided into five groups (n = 8 in each group): the sham-operated rabbits were used as control (Group A), Groups B received laparotomies and suture, Group C received laparotomies and covered SHEC suture without the PTX coating, Group D received laparotomies and covered PTX-SHEC suture, and Group E received laparotomies and 1000 μmol/L PTX administration. Liver function tests and residual dosage of PTX from each group were measured by enzyme-linked immunosorbent assay. Histological data and α-smooth muscle actin (SMA) immunohistochemical staining of common bile duct were examined.</p><p><b>Results:</b>NMR and FT-IR indicated that PTX was successfully introduced, based on the appearance of signals at 7.41-7.99 ppm, 1.50 ppm, and 1.03 ppm, due to the presence of aromatic protons, methylene protons, and methyl protons of PTX, respectively. No bile leak was observed. The PTX-conjugated film could slowly release PTX for 4 weeks (8.89 ± 0.03 μg at day 30). The in vitro cell viability test revealed significantly different levels of toxicity between films with and without PTX (111.7 ± 4.0% vs. 68.1 ± 6.0%, P < 0.001), whereas no statistically significant difference was observed among the three sets of PTX-contained films (67.7 ± 5.4%, 67.2 ± 3.4%, and 59.1 ± 6.0%, P > 0.05). Histological examinations revealed that after 28 days of implantment, Groups D and E (but not Group C) had less granulation tissue and glandular hyperplasia in the site of biliary duct injury than Group B. The pattern was more obvious in Group D than Group E. Less α-SMA-positive cells were found in tissue from Groups D and E. Comparing with Group E, the liver function was improved significantly in Group D, including total bilirubin (2.69 ± 1.03 μmol/L vs. 0.81 ± 0.54 μmol/L, P = 0.014), alanine aminotransferase (87.13 ± 17.51 U/L vs. 42.12 ± 15.76 U/L, P = 0.012), and alkaline phosphatase (60.61 ± 12.31 U/L vs. 40.59 ± 8.78 U/L, P < 0.001).</p><p><b>Conclusions</b>PTX-SHEC film effectively inhibites the myofibroblast proliferation and extracellular matrix over-deposition during the healing process of biliary reconstruction. This original film might offer a new way for reducing the occurrence of the benign biliary stricture.</p>

Therapeutic effects of antisense gene targeting at PCNA on proliferative cholangitis / 中华肝胆外科杂志

Objective In recent years, high stone recurrence and biliary restenosis rates in hepatolithiasis patients have been confirmed to be closely related to chronic proliferative cholangitis(PC).Since PC is a kind of chronic proliferative disease, we designed this study to investigate the feasibility and effectiveness of applying PCNA shRNA to inhibit the hyperplasia and lithogenic potentiality of PC and to explore a new treatment approach for the prevention of stone recurrence and biliary restenosis.Methods The common bile duct of a PC animal model was given an intralumenal administration of 0.5 ml of PCNA shRNA to investigate its influence on PC.Results The PCNA shRNA could efficiently inhibit the hyperplasia of the biliary epithelium, submucosal gland, and collagen fiber by inhibiting mRNA or protein expressions of PCNA and Procollagen Ⅲ, thus showing promise to control or reverse PC and its secondary biliary stricture.In addition, it could inhibit the lithogenic potentiality of PC by inhibiting the expression of mucin 5AC.Conclusion PCNA shRNA treatment might achieve the effect of controlling or reversing the PC and its subsequent biliary stricture and concurrently assist in preventing the recurrence of intrahepatic stone.

The Effects of Paclitaxel-Coated Nylon Thread on the Proliferative Cholangitis in a Rat / 한국간담췌외과학회지

BACKGROUND/AIMS: Local drug delivery to the bile duct may be effective to prevent proliferative cholangitis (PC) through capability of high dose administeration with minimal systemic side effects. Paclitaxel is an anticancer drug whose side effect on the stabilization of microtubule leads to cell death. The aims of this study were to establish the proliferative cholangitis model in rat, mimicking biliary stricture in human, and to test whether paclitaxel-coated nylon thread prevents biliary stricture in a PC model of rat. METHODS: PC was induced by introducing a fine nylon thread into the bile duct in a rat from 1 week to 4 week. To evaluate the effects of paclitaxel as a locally-delivered anti-proliferative drug, dog gallbladder epithelial cells were exposed to sequential concentrations of paclitaxel (0.1microM, 1microM, 10microM, and 100microM) for 20 min. in vitro, and inhibition of proliferation was measured by (3)H-thymidine uptake assay. Paclitaxel- coated 5-0 nylon threads (1.8+/-0.5 ug/3 cm thread, measured by HPLC) were made by immersion of ethanolic paclitaxel (50 mg/ml) and evaporation of the solvent. Nylon threads were inserted into the bile duct of male Sprague-Dawley rats weighing 200~50 g. Paclitaxel (n=15) and control (n=15) groups were divided with or without paclitaxel-coating procedure. The paclitaxel effects were assessed by histomorphological examination one week after thread implantation. RESULTS: The decrease of (3)H-thymidine uptake was observed at 100microM of paclitaxel exposured for 20 minutes in the presence of epidermal growth factor (50 nM/ml) than control. PC model characterized by epithelial-glandular proliferation and fibrous thickening of the bile duct wall through 1~ week. This model was established at 1 week. The effect of paclitaxel-coated nylon thread into the bile duct were evaluated after 1 week. In paclitaxel treated the luminal area, luminal length and the ratio of lumen to bile duct cross sectional area increased by 276% (p=0.044), 87% (p=0.012) and by 330% (p=0.000), respectively, versus control. The total wall area, epithelial-glandular area, and stromal area were similar between paclitaxel treated group and control (p>0.05). The bile duct wall thickness of paclitaxel group decreased by 33% (p=0.011, 273 (90)microM vs 410 (95)microM, paclitaxel vs control). CONCLUSION: Paclitaxel-coated nylon thread into bile duct was effective for the suppression of luminal stenosis, and may offer a therapeutic option for biliary stricture and biliary stricture associated disease.