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A hybrid substratum for primary hepatocyte culture that enhances hepatic functionality with low serum dependency.

Meng Q, Tao C, Qiu Z, Akaike T, Cui F, Wang X - Int J Nanomedicine (2015)

Bottom Line: A poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide) matrix can recognize cells and promote liver function in a spheroid structure because of a specific galactose-asialoglycoprotein receptor interaction.Meanwhile, a fusion protein, E-cadherin-Fc, when incubated with various cells, has shown an enhancing effect on cellular viability and metabolism.The isolated cells showed a monolayer aggregate morphology on the coimmobilized surface and displayed higher functional expression than cells on traditional matrices.

View Article: PubMed Central - PubMed

Affiliation: School of Materials Science and Engineering, Tsinghua University, Beijing, People's Republic of China ; State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, People's Republic of China ; Biomaterials Center for Regenerative Medical Engineering, Ibaraki, Japan.

ABSTRACT
Cell culture systems have proven to be crucial for the in vitro maintenance of primary hepatocytes and the preservation of hepatic functional expression at a high level. A poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide) matrix can recognize cells and promote liver function in a spheroid structure because of a specific galactose-asialoglycoprotein receptor interaction. Meanwhile, a fusion protein, E-cadherin-Fc, when incubated with various cells, has shown an enhancing effect on cellular viability and metabolism. Therefore, a hybrid substratum was developed for biomedical applications by using both of these materials to combine their advantages for primary hepatocyte cultures. The isolated cells showed a monolayer aggregate morphology on the coimmobilized surface and displayed higher functional expression than cells on traditional matrices. Furthermore, the hybrid system, in which the highest levels of cell adhesion and hepatocellular metabolism were achieved with the addition of 1% fetal bovine serum, showed a lower serum dependency than the collagen/gelatin-coated surface. Accordingly, this substrate may attenuate the negative effects of serum and further contribute to establishing a defined culture system for primary hepatocytes.

No MeSH data available.


Related in: MedlinePlus

The maintenance of primary hepatocytes on the hybrid matrix after 1, 3, 5, 7, and 9 days.Notes: The relative ratios of attached cells in various systems were determined by using the number of cells on PVLA without FBS as the 100% value. The number of primary hepatocytes on the hybrid matrix was higher in the medium with 1% FBS at every time point than the cells on the same surface without or with 10% FBS. The hepatocytes were viable on PVLA (1% FBS), gelatin (10% FBS), and collagen (10% FBS), which was used as a control (A). The relative preservation of hepatocytes was determined by normalizing the cell numbers to the value after 1 day on each matrix (B). The hybrid matrix was made up of EFC and PVLA. The symbol * denotes statistical significance at P<0.05.Abbreviations: FBS, fetal bovine serum; PVLA, poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide); EFC, E-cadherin-Fc.
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f4-ijn-10-2313: The maintenance of primary hepatocytes on the hybrid matrix after 1, 3, 5, 7, and 9 days.Notes: The relative ratios of attached cells in various systems were determined by using the number of cells on PVLA without FBS as the 100% value. The number of primary hepatocytes on the hybrid matrix was higher in the medium with 1% FBS at every time point than the cells on the same surface without or with 10% FBS. The hepatocytes were viable on PVLA (1% FBS), gelatin (10% FBS), and collagen (10% FBS), which was used as a control (A). The relative preservation of hepatocytes was determined by normalizing the cell numbers to the value after 1 day on each matrix (B). The hybrid matrix was made up of EFC and PVLA. The symbol * denotes statistical significance at P<0.05.Abbreviations: FBS, fetal bovine serum; PVLA, poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide); EFC, E-cadherin-Fc.

Mentions: The matrices continued to show effects on cell behavior during hepatocyte maintenance in culture. One day after seeding, the primary hepatocytes revealed a monolayer morphology on coimmobilized (Figure 3A and G) and gelatin-coated surfaces (Figure 3C and I), while they maintained a round shape on PVLA surfaces (Figure 3B and H). During continuous incubation, the hepatocytes clustered in the culture systems because of the presence of PVLA, whereas the cells spread out on the gelatin and other traditional matrices. Finally, the cells formed monolayer aggregates on the hybrid matrix (Figure 3D and J) and multilayer spheroid structures on PVLA-coated dishes (Figure 3E and K) after 5 days in culture. Furthermore, the viability of the maintained cells reached the highest level at each time point (days 1, 3, 5, 7, and 9) with a concentration of 1% FBS on the hybrid matrix and on PVLA (Figure 4A); however, a higher serum concentration could possibly enhance cell maintenance in the gelatin or collagen systems. The highest cell numbers, in ascending order for different time points, occurred on the following surfaces: PVLA < hybrid surface < gelatin and collagen, especially in the early culture period. The total number of cells gradually became smaller over time, and finally remained similar among these systems on days 7 and 9. Moreover, the number of remaining hepatocytes relative to the initial cell number during 1 week was higher on the surfaces with PVLA than on the other matrices supplemented with the optimal serum concentrations (Figure 4B).


A hybrid substratum for primary hepatocyte culture that enhances hepatic functionality with low serum dependency.

Meng Q, Tao C, Qiu Z, Akaike T, Cui F, Wang X - Int J Nanomedicine (2015)

The maintenance of primary hepatocytes on the hybrid matrix after 1, 3, 5, 7, and 9 days.Notes: The relative ratios of attached cells in various systems were determined by using the number of cells on PVLA without FBS as the 100% value. The number of primary hepatocytes on the hybrid matrix was higher in the medium with 1% FBS at every time point than the cells on the same surface without or with 10% FBS. The hepatocytes were viable on PVLA (1% FBS), gelatin (10% FBS), and collagen (10% FBS), which was used as a control (A). The relative preservation of hepatocytes was determined by normalizing the cell numbers to the value after 1 day on each matrix (B). The hybrid matrix was made up of EFC and PVLA. The symbol * denotes statistical significance at P<0.05.Abbreviations: FBS, fetal bovine serum; PVLA, poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide); EFC, E-cadherin-Fc.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4376262&req=5

f4-ijn-10-2313: The maintenance of primary hepatocytes on the hybrid matrix after 1, 3, 5, 7, and 9 days.Notes: The relative ratios of attached cells in various systems were determined by using the number of cells on PVLA without FBS as the 100% value. The number of primary hepatocytes on the hybrid matrix was higher in the medium with 1% FBS at every time point than the cells on the same surface without or with 10% FBS. The hepatocytes were viable on PVLA (1% FBS), gelatin (10% FBS), and collagen (10% FBS), which was used as a control (A). The relative preservation of hepatocytes was determined by normalizing the cell numbers to the value after 1 day on each matrix (B). The hybrid matrix was made up of EFC and PVLA. The symbol * denotes statistical significance at P<0.05.Abbreviations: FBS, fetal bovine serum; PVLA, poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide); EFC, E-cadherin-Fc.
Mentions: The matrices continued to show effects on cell behavior during hepatocyte maintenance in culture. One day after seeding, the primary hepatocytes revealed a monolayer morphology on coimmobilized (Figure 3A and G) and gelatin-coated surfaces (Figure 3C and I), while they maintained a round shape on PVLA surfaces (Figure 3B and H). During continuous incubation, the hepatocytes clustered in the culture systems because of the presence of PVLA, whereas the cells spread out on the gelatin and other traditional matrices. Finally, the cells formed monolayer aggregates on the hybrid matrix (Figure 3D and J) and multilayer spheroid structures on PVLA-coated dishes (Figure 3E and K) after 5 days in culture. Furthermore, the viability of the maintained cells reached the highest level at each time point (days 1, 3, 5, 7, and 9) with a concentration of 1% FBS on the hybrid matrix and on PVLA (Figure 4A); however, a higher serum concentration could possibly enhance cell maintenance in the gelatin or collagen systems. The highest cell numbers, in ascending order for different time points, occurred on the following surfaces: PVLA < hybrid surface < gelatin and collagen, especially in the early culture period. The total number of cells gradually became smaller over time, and finally remained similar among these systems on days 7 and 9. Moreover, the number of remaining hepatocytes relative to the initial cell number during 1 week was higher on the surfaces with PVLA than on the other matrices supplemented with the optimal serum concentrations (Figure 4B).

Bottom Line: A poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide) matrix can recognize cells and promote liver function in a spheroid structure because of a specific galactose-asialoglycoprotein receptor interaction.Meanwhile, a fusion protein, E-cadherin-Fc, when incubated with various cells, has shown an enhancing effect on cellular viability and metabolism.The isolated cells showed a monolayer aggregate morphology on the coimmobilized surface and displayed higher functional expression than cells on traditional matrices.

View Article: PubMed Central - PubMed

Affiliation: School of Materials Science and Engineering, Tsinghua University, Beijing, People's Republic of China ; State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, People's Republic of China ; Biomaterials Center for Regenerative Medical Engineering, Ibaraki, Japan.

ABSTRACT
Cell culture systems have proven to be crucial for the in vitro maintenance of primary hepatocytes and the preservation of hepatic functional expression at a high level. A poly-(N-p-vinylbenzyl-4-O-β-D-galactopyranosyl-D-gluconamide) matrix can recognize cells and promote liver function in a spheroid structure because of a specific galactose-asialoglycoprotein receptor interaction. Meanwhile, a fusion protein, E-cadherin-Fc, when incubated with various cells, has shown an enhancing effect on cellular viability and metabolism. Therefore, a hybrid substratum was developed for biomedical applications by using both of these materials to combine their advantages for primary hepatocyte cultures. The isolated cells showed a monolayer aggregate morphology on the coimmobilized surface and displayed higher functional expression than cells on traditional matrices. Furthermore, the hybrid system, in which the highest levels of cell adhesion and hepatocellular metabolism were achieved with the addition of 1% fetal bovine serum, showed a lower serum dependency than the collagen/gelatin-coated surface. Accordingly, this substrate may attenuate the negative effects of serum and further contribute to establishing a defined culture system for primary hepatocytes.

No MeSH data available.


Related in: MedlinePlus