Mesenchymal stem cells (MSCs), a class of adult stem cells, are considered a promising source for bone regeneration. Although combining MSCs with biomaterial scaffolds offers an interesting clinical strategy for bone tissue engineering, the presence of the scaffolds could induce an undesirable effect on cell–cell interactions. Moreover, before the application of scaffold materials in bone tissue reconstruction, cells must be manipulated with proteolytic enzymes, such as trypsin or dispase that degrade extracellular matrix (ECM) molecules and cell surface proteins, which can result in the cell damage and loss of cellular activity. Therefore, the development of alternative strategies for bone regeneration is required to solve these problems. Recently, a novel tissue engineering technology named ‘cell sheet’ has been efficaciously utilized in the regeneration of bone, corneal, cardiac, tracheal and periodontal ligament-like tissues. The cell sheet is a layer of cells, which contains intact ECM and cell surface proteins such as growth factor receptors, ion channels and cell-to-cell junction proteins. MSC sheets can be easily fabricated by layering the recovered cell sheets without any scaffolds or complicated manipulation. This review summarizes the current state of the literature regarding the use of MSCs to produce cell sheets and assesses their applicability in bone tissue regeneration and repair.