Characterization of a Long-Lived Alginate Lyase Derived from Shewanella Species YH1

Polysaccharides from seaweeds are widely used in various fields, including the food, biomedical material, cosmetic, and biofuel industries. Alginate, which is a major polysaccharide in brown algae, and the products of its degradation (oligosaccharides) have been used in stabilizers, thickeners, and gelling agents, especially in the food industry. Discovering novel alginate lyases with unique characteristics for the efficient production of oligosaccharides may be relevant for the food and pharmaceutical fields. In this study, we identified a unique alginate lyase derived from an alginate-utilizing bacterium, Shewanella species YH1. The recombinant enzyme (rAlgSV1-PL7) was produced in an Escherichia coli system and it was classified in the Polysaccharide Lyase family 7. The optimal temperature and pH for rAlgSV1-PL7 activity were around 45 °C and 8, respectively. Interestingly, we observed that rAlgSV1-PL7 retained over 80% of its enzyme activity after incubation at 30 °C for at least 20 days, indicating that rAlgSV1-PL7 is a long-lived enzyme. Moreover, the degradation of alginate by rAlgSV1-PL7 produced one to four sugars because of the broad substrate specificity of this enzyme. Our findings suggest that rAlgSV1-PL7 may represent a new commercially useful enzyme

Transplantation of hybrid adipose-derived stem cell sheet with autologous peritoneum: An in vivo feasibility study

Introduction: In regenerative medicine, cell sheet engineering has various advantages, including the retention of cells at the transplantation site for a longer period and the local delivery of growth factors and cytokines. Adipose-derived stem cell (ASC) is widely used owing to their various functions such as wound healing, immunomodulation, and nerve regeneration, in addition to their ability to differentiate into adipocytes, chondrocytes, and osteoblasts. ASC sheet generated using cell sheet engineering is considered effective in preventing anastomotic leakage, a serious postoperative complication in gastrointestinal surgery. However, the ASC sheet is too soft, thin, and brittle to handle with laparoscopic forceps during the operation. Therefore, we considered using the peritoneum, which is stiff and easy to collect while operating, as an alternative support. In this study, we explored the feasibility of using the peritoneum as a support for the precise transplantation of ASC sheets during surgery. Methods: ASCs were isolated from the subcutaneous fat of the inguinal region of Sprague-Dawley (SD) transgenic rats expressing green fluorescent protein. ASCs were cultured until passage 3, seeded in temperature-responsive culture dishes, and the resulting ASC sheet was harvested at more than 80% confluency. Non-transgenic SD rats were used for transplant experiments. The wall peritoneum was harvested from SD rats following laparotomy, and hybrid adipose-derived stem cell (HASC) sheet was prepared by laminating the peritoneum with ASC sheet. The cell sheets were transplanted on the backs of SD rats following the incision. On post-transplantation days 3 and 7, the specimens were extracted. ASC and HASC sheets were then compared macroscopically and histopathologically. Results: HASC sheet transplantation was macroscopically and histopathologically more effective than ASC sheet transplantation. The peritoneum provided sufficient stiffness as a support for precise transplantation. Conclusion: The newly developed HASC sheet, which combine the advantages of ASC sheet with those of the peritoneum, could be more useful for clinical application than the ASC sheet alone