Cholangiopathies (CPs) are a set of diseases affecting the biliary tree and are a significant cause of mortality in adults and children1. Some CPs including biliary atresia, distal cholangiocarcinoma and iatrogenic injures affect in particular the common bile duct (CBD) with strictures leading to liver failure. Therapeutic options for these disorders are limited and rely mainly on the anastomosis of the CBD to the jejunum, which is associated with duodenal ulcers, retrograde infections, and stenosis. Liver transplantation remains the most effective therapeutic option2, but due to the shortage of donor organ and associated costs1, new therapeutic alternatives are needed. The bioengineering of bile ducts represents a promising strategy to restore bile flow continuity2 for CBD disorders. However, the results of this research have been often inconclusive and poorly translatable. Our research group is working on optimizing both the cellular source and the biomaterials, which represent the main challenges of this research field, with the prospect of creating an implantable bioartificial CBD (BCBD). For its fabrication two techniques are used. First, cholangiocytes are mixed with photocollagen at 3 million cells/mL of polymer. Then, the cell-embedded photocollagen is poured into a plastic mold and incubated at 37°C for 30 minutes. The mold is therefore opened and the hydrogel is photopolymerized by UV exposure for complete crosslinking. An external coating is added by dipping the cell-embedded hydrogel pipe into an agarose solution. The BCBD is therefore transferred into the culture medium. Alternatively, electrospun polyurethane-fibers replace agarose on the outer surface of the photocollagen tube. Optimization of cell culture parameters, including photocollagen concentration, cell density, and length of culture, are conducted in small-scale experiments in 96-multiwell plates using disks of cell-embedded material. Mechanical (uniaxial and circumferential tensile testing, dynamic compliance) and cell characterization (cell viability, functionality, scaffold colonization) are performed to select the best parameters for obtaining a functional BCBD. We obtained a BCBD of dimensions similar to the native CBD: 60mm long, 1mm thick, and inner diameter of 5.5mm. For preliminary characterization, we used HuCCT1 intrahepatic cholangiocarcinoma cells, which are embedded in disks of 4-8mg/mL photocollagen and at 1-5 million cells/mL photocollagen. We demonstrate that cells embedded in 6mg/mL photocollagen and at 3 million cells/mL show the best viability profile overtime (up to 14 days of culture) and express biliary markers including cytokeratin 7 and cystic fibrosis transmembrane regulator. We obtained a human-sized BCBD through a simple, inexpensive and easy-to-scale method. We optimized some cell parameters and many other characterizations of cells and materials are ongoing. We trust that the our BCBD, consisting on materials commonly used in tissue engineering3-5 and a cell source already proven to be suitable for our purpose6, will be ready for a preclinical assessment soon.
1Lazaridis et al. Mayo Clin. Proc. 90, 791–800 (2015)
2Justin et al. Acta (BBA)-Molecular Basis Dis. 1864, 1532–1538 (2018)
3Campos et al. Front. Bioeng. Biotechnol. 8, 596 (2020)
4Alonso et al. J. Surg. Res. 179, 18–21 (2013)
5Mazari-Arrighi et al. Biomaterials, 12120 (2021)
6Sampaziotis et al. Nat. Med. 23, 954–963 (2017)