The loss of viable osteocytes in the elderly is a critical issue upon the development of osteoporosis. Previous work in our group suggested a positive osteocyte survival effect by TGFβ₃ supplemented to a serum containing medium during long-term ex vivo culture of human cancellous bone explants. In this study, no beneficial effect by TTGFβ₃ addition to serum-free cultured ex wVo osteocytes was observed. Therefore, the effect of TGFβ₃ on osteocyte survival was further investigated in vitro.

The most commonly used culture model for osteoblast-lineage cells is 2D/monolayer culture. However, monolayer culture only resembles the in vivo situation of osteoblast-lineage cells in bone to a limited extent, omitting the extensive 3D cell-to-cell and cell-to-matrix interconnectivity. In order find a more relevant culture model to be used for in vitro investigations of osteoblast biology;​ this study evaluated a variety of cell culture systems.

In all systems evaluated, the effect of TGFβ₃ was strongly dependent upon the presence of serum. Only in the presence of serum did TGFβ₃ increase osteocyte survival both ex vivo and in vitro. These investigations, also, led to the generation of different 3D in vitro culture models. Within a multilayer and pellet culture model for human primary osteoblasts, a reduction of initial proliferation in combination with a more rapid cell differentiation in comparison to monolayer culture was identified. Such 3D culture models are of great benefit compared to the existing in vitro osteoblast monolayer culture as they create more active/mature osteoblasts or potentially early osteocytes in a shorter time course.

This study showed, firstly TGFβ₃ is an in vitro survival factor for osteocytes, if cultured in the presence of serum, and secondly the choice of 3D culture models for osteoblast-lineage cells should be intended more frequently to properly evaluate cellular reactions prior to conducting animal studies.