# Three-Dimensional-Bioprinted Liver Chips and Challenges
## 1 Introduction
Liver cells for OOC cultures can be derived from primary human hepatocytes (PHHs) (they possess intrinsic properties of the liver but are difficult to isolate, costly, and unsuitable for long-term culture), stem cell-derived hepatocytes (a stable source of hepatocytes, but they are difficult to manipulate and require specific induce factors), and hepatic-derived cell lines (which have an indefinite lifespan and are easy to manipulate, but suffer from inaccuracy in drug response and rapid loss of expression of liver-specific enzymes/transporters) [48,73,76]. \begin{table}
\begin{tabular}{c c c c c c} \hline \hline \multirow{2}{*}{**Method**} & **Extursion-Based** & \multirow{2}{*}{**Inkjet Bioprinting**} & \multicolumn{2}{c}{**Stereolithography-**} & \multirow{2}{*}{**Multiphoton-Based** **Bioprinting**} & **Laser-Based** **Bioprinting** \\  & **Bioprinting** & & & **(SLA, DLP)** & & **Bioprinting** \\ \hline \multirow{2}{*}{**Working Principle**} & Pneumatic, mechanical, microfluidics assisted & Thermal, piezoelectric, electrostatic & Point-by-point exposure (SLA), layer-layer & Point-by-point exposure & Laser-induced \\  & & & exposure (DLP) & by a light beam & Laser-induced \\ \hline \multicolumn{2}{c}{**Printable Viscosity**} & 30 to \(6\times 107\) m Pas [55] & 3-12 m Pas [55] & 1-1000 m Pas [56] & - & 1-300 m Pas [57,58] \\ \hline \multicolumn{2}{c}{**Cell Density**} & High [58] & Low [58] & Medium [58] & Medium & Medium [58] \\ \hline \multicolumn{2}{c}{**Print Speed**} & Medium [59,60] & High [59] & High [57] & High & High \\ \hline \multicolumn{2}{c}{**Resolution**} & \textgreater{}100 μm [61] & \textgreater{}10 μm [62] & \textgreater{}6–10 μm [57] & \textgreater{}100 nm [63] & \textgreater{}50–100 μm [64] \\ \hline \multicolumn{2}{c}{**Cell Viability**} & \textgreater{}90\% [65] & \textgreater{}85\% [66] & \textgreater{}80\% [67] & \textgreater{}90\% [68] & \textgreater{}90\% [69,70] \\ \hline \multicolumn{2}{c}{**Adrodability, large**} & \multirow{3}{*}{ \begin{tabular}{c} **Adrodability, good** \\ **bioprinting**, of \\ **bi-viscosity** **biophins** \\ **with high cell** \\ **concentration,** \\ **bioprinting**, scalability** \\ **[60,61]** \\ \end{tabular}  } & \begin{tabular}{c} **High resolution, high** \\ **resolution, fast** \\ **bioprinting**, the ability** \\ **to generate a cell** \\ **concentration gradient** \\ **[57,58]** \\ **[67,58]** \\ **[60,61]** \\ \end{tabular}  & \begin{tabular}{c} **High resolution (single** \\ **cell per droplet, high \\ **cell visibility, no** \\ **clogging, ability to** \\ **can be bioprinted** \\ **[57,58]** \\ **[57,58]** \\ \end{tabular}  & \begin{tabular}{c} **High resolution (single** \\ **cell per droplet, high \\ **cell visibility, no** \\ **clogging, ability to** \\ **bioprint low-viscosity** \\ **bioniths with high cell** \\ **density** [58,64] \\ \end{tabular}  \\ \hline \multicolumn{2}{c}{**Limitations**} & \ \begin{tabular}{c} High shear stress, \\ **nozzleogging, low** \\ **resolution, low** \\ **structural fidelity,** \\ **limited material choice** \\ **(only materials with** **shear-thinning** \\ **spavil for** \\ **property)** [60,61] \\ \end{tabular}  &
\begin{tabular}{c} High-viscosity **biohik** \\ **cannot be bioprinted,** \\ **limited vertical** \\ **bioprinting**,** **dogging,** \\ **shear stress, additional** \\ **cross-linking step is** \\ **sleared-thinning** \\Figure 1: Three-dimensional bioprinting. (**A**) Different 3D bioprinting methods. Reproduced with permission from [8]. 2016, Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim. (**B**) Schematic representation of liver-on-chip 3D bioprinting to form a bioreactor with continuous perfusion for long-term cell culture. (**C**) Implemented liver chip comprised of PDMS and PMMA layers, compressed by nuts and screws to enable disassembly of the setup for examining cell culture anytime [75]. (**B,C**) Reproduced with permission from [75].