Author:

Keywords:

Science & Technology, Life Sciences & Biomedicine, Food Science & Technology, Cell dissociation, Proteolytic enzyme, Myoblast, Embryonic stem cells, Mesenchymal stromal cells, EMBRYONIC STEM-CELLS, VIABILITY, Subtilisins, Animals, Humans, Cattle, Mesenchymal Stem Cells, Myoblasts, Cell Proliferation, Cell Differentiation, Cell Culture Techniques, Bacillus subtilis, Cell Line, Trypsin, Cell Adhesion, S002821N#55955168, 0904 Chemical Engineering, 0908 Food Sciences, 1111 Nutrition and Dietetics, Food Science, 3006 Food sciences, 3210 Nutrition and dietetics, 4004 Chemical engineering

Abstract:

Cellular agriculture and regenerative medicine strive to replace animal-derived products in their processes as much as possible. In these efforts dissociation enzymes required for cell detachment from their substrate are often overlooked. Cell detachment is an essential part of subculturing and to date it is achieved using either animal pancreas-derived trypsin or expensive recombinant enzymes. We found that nattokinase, a fermentation product of Bacillus subtilis subsp. Natto, can be effectively applied for the detachment of bovine myoblasts, mesenchymal stromal cells (MSCs) and human embryonic cell line (H9). We described the suitable enzyme concentrations, incubation times and inhibition procedures for each cell type. Long-term subculture with nattokinase did not negatively affect proliferation nor differentiation of the tested cells and even led to significantly higher number of CD56+ cells in myoblast culture after six passages (88 ± 3 % vs 73 ± 5 %, p = 0.03, n = 6) compared to trypsin control. Efficacy, affordability, and current "generally recognized as safe" status of nattokinase make it an attractive product for cell-based food manufacturing.