Aim: To develop a novel hepatocyte serum-free medium based on sericin, and to explore the effect of sericin on the hepatocyte transcriptome.
Methods: A controlled trial comparing novel serum-free medium and other media: C3A cells were cultured in our novel serum-free medium, HepatoZYME, complete medium (DMEM/F12 with 100 mL/L FBS), and DMEM/F12, and then cell attachment, proliferation, and function as well as the biocompatibility of the media were assessed. A comparative study of serum-free media with or without 2 mg/mL sericin: the effect of sericin on C3A growth was assessed by cell viability and proliferation, the effect of sericin on C3A cell cycle distribution was determined by flow cytometry, and the effect of sericin on the C3A transcriptome was assessed by gene-chip array and RT-qPCR.
Results: More C3A cells attached to the plate containing our serum-free medium than to those containing HepatoZYME and DMEM/F12 at 24 h post-seeding. Both the viability and proliferation rate of C3A cells in sericin-based serum-free medium were superior to those of cells in HepatoZYME and DMEM/F12 (P < 0.001). The content of albumin and urea in our serum-free medium was significantly higher than that in HepatoZYME and DMEM/F12 throughout the whole culture period (P < 0.001) and was similar to that in complete medium at day 3, 4, and 5. In part 2, cell viability and proliferation were greater in the presence of 2 mg/mL sericin (P < 0.001), as was the proportion of cells in S phase (16.21% ± 0.98% vs 12.61% ± 0.90%, P < 0.01). Gene-chip array analysis indicated that the expression of CCR6, EGFR, and FOS were up-regulated by 2 mg/mL sericin, and RT-qPCR revealed that the expression of CCR6, EGFR, FOS, AKT1, JNK1, NFkB1, MMP-9, MEK2, ERK1/2 and MYC was up-regulated by 2 mg/mL sericin (P < 0.05).
Conclusion: We developed a novel hepatocyte serum-free medium. Sericin probably enhances cell attachment through the CCR6-Akt-JNK-NF-κB pathway and promotes cell proliferation through CCR6-mediated activation of the ERK1/2-MAPK pathway.
Keywords: Bioartificial liver support system; CCR6; MAPK pathway; Sericin; Serum-free medium.