Development and characterization of a cholangiocyte cell line from the PCK rat, an animal model of Autosomal Recessive Polycystic Kidney Disease

In the PCK rat, a rodent model of Autosomal Recessive Polycystic Kidney Disease (ARPKD), a spontaneous splicing mutation of Pkhd1 initiates hepatic cyst development. Cystic cholangiocytes possess short and malformed cilia that do not express fibrocystin, the Pkhd1 protein. During the disease course, cysts continue to grow; however, the mechanisms underlying cyst progression are unclear due in part to the lack of suitable cell lines to study cystogenesis. Here, we describe the development of a PCK-derived cholangiocyte cell line (PCK-CCL). Normal rat cholangiocytes (NRCs) were used as a control. The PCK-CCL maintained a cholangiocyte phenotype as assessed by the expression of the CK-19, CK-7 and GGT. PCK-CCL grown on collagen formed a polarized monolayer with well-developed junctional complexes, and distinct apical and basolateral membranes. Compared to NRCs, cilia in the PCK-CCL were short and malformed and did not express fibrocystin. The PCK-CCL exhibited a higher rate of proliferation (P<0.05) with a doubling time approximately half that of NRCs. By RT-PCR analysis of exons 33-37, an approximately 800 bp product of Pkhd1 was amplified in NRCs. In contrast and as expected, in the PCK-CCL, the Pkhd1 amplicon was smaller (∼630 bp) reflecting the IVS35-2A → T mutation. PCK-CCL and NRCs seeded in 3-D cultures formed cystic structures; however, the PCK cysts expanded progressively up to day 21 while cysts formed by NRCs remained the same size after day 9. In summary, we have developed a cholangiocyte cell line from the PCK rat that retains properties of the cholangiocytes lining hepatic cysts in vivo. The cells have been grown continuously for ∼18 month and 45 passages without crisis or senescence. The morphology and growth characteristics of the PCK-CCL are consistent with those seen in vivo in the PCK rat, suggesting that this cell line will be useful in dissecting the mechanisms of hepatic cyst formation.