Abstract
Human periodontal ligament stem cells (PDLSCs) have been widely applied as seed cells and cell sheets in periodontal tissue regeneration. Despite significant progress in PDLSCs application, it is a major challenge to promote cell proliferation and multiple differentiations of PDLSCs because cell numbers at the initial obtaining are limited. The goal of study was to determine the paracrine effects of human urine-derived stem cells (USCs) on cell proliferation and osteogenic differentiation of PDLSCs when USCs were indirectly-co-cultured with PDLSCs. After indirectly-co-cultured with USCs at different ratios (PDLSC/USC, 1/0.5, 1/1 and 1/2), number of PDLSCs among the three co-cultured groups visibly increased from day 5 to a similar extent, and the expression of osteogenic and cementogenic genes and proteins in the osteogenic medium significantly increased with an increasing proportion of USCs compared to USC-free control group. In addition, osteogenic matrix PDLSC sheets at a PDLSC/USC ratio of 1/2 contained denser collagen layers and exhibited increased osteogenic and cementogenic protein expression. In vivo transplantation showed that PDLSC sheets noncontact cocultured at a PDLSC/USC ratio of 1/2 formed more new and dense structures and expressed higher levels of osteogenic and cementogenic proteins. In conclusion, the present results demonstrate that USCs promote the proliferation and osteogenic and cementogenic differentiation of PDLSCs in a ratio-dependent manner through noncontact coculture and further accelerate the regeneration of new structures by osteogenic matrix PDLSC sheets in vivo. These results suggest their use as a new strategy for application in clinical periodontal tissue repair.