A Short-Term Preservation of Human Cultured Periosteal Sheets, Osteogenic Grafting Materials, Using a Commercial Preservation Solution Containing Epigallocatechin-3-gallate (Theliokeep^®) under Hypothermic Conditions

Abstract

In the past decade, it has increasingly been reported that epigallocatechin-3-gallate (EGCG), a major catechin derivative extracted from Green tea, has various bioactivities, including a cell-protective action on mammalian cells and tissues. In this study, we have tested a commercial preservation solution containing EGCG (Theliokeep(®)) in both two- and three-dimensional cultures of human periosteal sheets, which have been used as an osteogenic grafting material for periodontal regenerative therapy. When periosteal sheets were 3D-cultured on collagen mesh, cell viability was maintained for 2 days using the hypothermic EGCG preservation solution. Replenishment of EGCG solution with 2-day intervals prevented the time-dependent decline in cell viability at 3 days and later. As observed in nonpreserved control cultures, most cells were positive for proliferating cell-nuclear antigen (PCNA) in the cultures preserved at 4°C in the EGCG solution, whereas PCNA-negative cells were increased in the cultures preserved at 4°C in the MesenPRO medium. In periosteal sheets 2D-cultured in plastic dishes, the EGCG solution occasionally was associated with vacuole formation in the cytoplasm, but cells could again expand in the culture medium at 37°C. As observed in the nonpreserved periosteal sheets control, the osteogenic induction upregulated alkaline phosphatase in those cells and tissues preserved in the EGCG solution. The EGCG solution protected cells from the cold shock-induced membrane phospholipid peroxidation. Our data suggest that the EGCG solution acts as an antioxidant to protect periosteal cells from cold shock and preserves cells under chilled conditions. The limited period of preservation time could be expanded by repeating replenishment of the EGCG solution or by optimizing the formula to be more favorable for human periosteal sheets without sacrificing cell viability. This methodology of preserving human cultured periosteal sheets with EGCG would be expected to support and spread the clinical use of regenerative therapy with autologous periosteal sheets.

Synthetic Calcium–Phosphate Materials for Bone Grafting

“…EGCG also inhibits osteoblast migration induced by insulin and growth factor I, potentially contributing to the regulation of bone remodeling, possibly by suppressing p44/p42 MAP kinase [ 359 ]. The ability of EGCG to inhibit osteoblast migration induced by EGF has been proposed as a key mechanism underlying its beneficial effects on proper bone remodeling [ 360 ]. Additionally, EGCG has demonstrated the capacity to enhance cell viability in 3D human periosteal cultures and has been used as an osteogenic graft material for periodontal regenerative therapy [ 360 ].…”

“…The ability of EGCG to inhibit osteoblast migration induced by EGF has been proposed as a key mechanism underlying its beneficial effects on proper bone remodeling [ 360 ]. Additionally, EGCG has demonstrated the capacity to enhance cell viability in 3D human periosteal cultures and has been used as an osteogenic graft material for periodontal regenerative therapy [ 360 ]. Treatment with EGCG at concentrations of 6–10 mM has been shown to upregulate the expression of type I collagen, osteopontin, and osterix in the human periodontal ligament (PDL) cells, suggesting a promising role for this plant polyphenol in periodontal regeneration [ 361 ].…”

Complete human serum maintains viability and chondrogenic potential of human synovial stem cells: suitable conditions for transplantation

“…This also increases manufacturing costs. For the preservation of cells, some alternatives to freezing have been reported, such as green tea polyphenol, also called as epigallocatechin-3-gallate (EGCG) [14, 15]. However, this solution has not been approved for pharmaceutical use in humans.…”