gms | German Medical Science

Objectives: Hepatic osteodystrophy refers to disordered bone homeostasis associated with liver damage, mainly manifested as osteopenia and osteoporosis. When liver damage occurs, it can indirectly or directly affect bone homeostasis. To identify/investigate underlying pathogenic mechanisms, this stud aimed at generating a stable, reproducible, and reliable 3D Liver-Bone (L-B) organ-on-a-chip system that mimics the altered bone metabolism and liver damage associated with chronic diclofenac exposure.

Methods: Human hepatic progenitor HepaRG cell spheroids were used as a model for the liver compartment. 3D co-cultures of human immortalized mesenchymal stem SCP-1 cells and human monocytic THP-1 cell line were used as a model for the bone compartment. Bone scaffolds were co-cultured with the HepaRG spheroids in an agarose platform, to allow direct medium exchange without cell-cell contact. The effects of different L-B medium combinations (100%, 75%, and 25) and the single media supplements on HepaRG cells and bone cells viability and function were investigated.Diclofenac was chronically applied at a concentration of 6 µM. After 7, 14, and 21 days of co-culture, bone and liver cells were assessed for viability (resazurin conversion, total DNA), function (alkaline phosphatase [AP], tartrate-resistant acid phosphatase [TRAP], cytochrome P450 2C9 [CYP2C9], and uridine diphosphate-glucuronosyltransferases [UGT] activity, and secreted proteins)

Results and conclusion: Our results indicate that osteogenic media supplements did not have a significant impact on HepaRG cell viability and function. Nevertheless, Dimethyl sulfoxide (induces hepatocyte differentiation) negatively affected bone cell viability and function (p<0.001). The different L-B media mixes did not negatively affect HepaRG spheroids viability and function. However, the L-B medium mix containing 75% liver cell culture medium affected bone cell viability (p<0.001 at day 21). Chronic Diclofenac exposure significantly increased CYP2C9 activity in liver spheroids co-cultured with bone cells. Furthermore, it significantly increased TRAP activity in the respective bone co-cultures. This effect on TRAP activity was not observed when the bone-co-cultures were not in interaction with the functional liver compartment.

Our data indicate that the tested L-B medium mix is suitable to maintain our established liver-bone co-culture system for up to 21 days. In our co-culture system, bone homeostasis was disrupted after chronic diclofenac exposure similar to that in humans. The 3D L-B in vitro model presented here shows for the first time a functional liver culture in combination with a functional bone culture. In addition, it not only allows to reduce animal experiments, but also to study the disturbed bone metabolism caused by liver damage.