Lab on a chip-based hepatic sinusoidal system simulator for optimal primary hepatocyte culture

Abstract

Primary hepatocyte cultures have been used in studies on liver disease, physiology, and pharmacology. While they are an important tool for in vitro liver studies, maintaining liver-specific characteristics of hepatocytes in vitro is difficult, as these cells rapidly lose their unique characteristics and functions. Portal flow is an important condition to preserve primary hepatocyte functions and liver regeneration in vivo. We have developed a microfluidic chip that does not require bulky peripheral devices or an external power source to investigate the relationship between hepatocyte functional maintenance and flow rates. In our culture system, two types of microfluidic devices were used as scaffolds: a monolayer- and a concave chamber-based device. Under flow conditions, our chips improved albumin and urea secretion rates after 13 days compared to that of the static chips. Reverse transcription polymerase chain reaction demonstrated that hepatocyte-specific gene expression was significantly higher at 13 days under flow conditions than when using static chips. For both two-dimensional and three-dimensional culture on the chips, flow resulted in the best performance of the hepatocyte culture in vitro. We demonstrated that flow improves the viability and efficiency of long-term culture of primary hepatocytes and plays a key role in hepatocyte function. These results suggest that this flow system has the potential for long-term hepatocyte cultures as well as a technique for three-dimensional culture.

Spheroids-on-a-chip: Recent advances and design considerations in microfluidic platforms for spheroid formation and culture

“…Indeed, continuous perfusion culture improves oxygen and nutrition supply for cells, as previously predicted computationally [63][64][65][66][67]. It has also been reported that gene expression profiles collected from hepatocyte cells cultured on a microfluidic chip with dynamic flow conditions showed more hepatocyte-specific characteristics than cells in static conditions [68].…”

“…In general, a microwell-based µSFC needs a specified number of microwells and a microchannel network to deliver the culture medium ( Figure 5A-E). For spheroid generation, the cell suspension is introduced at the inlet, usually with a cell density of 105 to 107 cells/ml [6,68,96], depending on the cell type ( Figure 5A). To achieve uniform cell seeding in microwells and uniformly sized spheroids, the cell suspension should fill the device entirely before cells begin to deposit in the microwells.…”

Inventions and Innovations in Preclinical Platforms for Cancer Research

“…Few minutes are needed that cells deposit on the bottom of the microwells and the microchannel ( Figure 4B). The cells that did not trap in the microwells are pushed out of the chip before the cells make aggregations and clog microchannels [8,24,103] (Figure 4C). Next, the cells begin aggregation and form spheroids ( Figure 4D) and are culture for drug screening ( Figure 4E).…”

Architecture in 3D cell culture: An essential feature for in vitro toxicology

“…Although human embryonic stem cells have been successfully used for differentiation into cholangiocytes with formation of polarized cysts and biliary ducts in 3D cell culture (Dianat et al, 2014), recapitulation of the polarized architecture of hepatic cords with human cells using the sandwich method is awaited. Ultimately, these relatively simple monoculture and coculture systems can serve as basis to include more complex parameters of liver physiology, such as a directional movement of medium to mimic sinusoidal blood flow and shear forces (Burkhardt et al, 2014; Choi et al, 2016). …”