In teeth with an injured pulp, dentin matrix orchestrates hard tissue repair through the release of dentin extracellular matrix components (dEMCs). dEMCs regulate the differentiation of resident mesenchymal stromal cells (MSCs), thereby affecting mineral deposition. In this study, we show that low-concentration solubilized dEMCs in osteogenic cultures of human umbilical cord mesenchymal stromal cells (UC-MSCs) and dental pulp stromal cells (DPSCs) enhanced mineral deposition, while adipose stromal cells (ASCs) were barely affected. Interestingly, UC-MSCs displayed significantly greater hydroxyapatite formation compared with DPSCs. UC-MSCs and DPSCs showed a dose-dependent viability and proliferation, whereas proliferation of ASCs remained unaffected. Qualitative analysis of the dEMC-supplemented osteogenic cultures through scanning electron microscopy demonstrated differences in the architecture of the deposited mineralized structures. Large-sized mineral accretions on a poorly organized collagen network were the prominent feature of UC-MSC cultures, while mineral nodules interspersed throughout a collagen mesh were observed in the respective DPSC cultures. The ability of dEMCs to induce mineralization varies between different human MSC types in terms of total mineral formation and architecture. Mineral formation by UC-MSCs exposed to low-concentration dEMCs proved to be the most efficient and therefore could be considered a promising combination for mineralized tissue engineering. Impact Statement This research has been conducted with the aim to contribute to the development of treatment modalities for the reconstruction of lost/damaged mineralized tissues. Currently, determining the most appropriate stromal cell population and signaling cues stands at the core of developing effective treatments. We provide new insights into the effect of innate inductive cues found in human dentin matrix components, on the osteogenic differentiation of various human stromal cell types. The effects of dentin extracellular matrix components on umbilical cord mesenchymal stromal cells have not been investigated before. The findings of this study could underpin translational research based on the development of techniques for mineralized tissue engineering and will be of great interest for the readership of Tissue Engineering Part A.