• A simple alternative method may replace the laboratory animals in teratogenic studies. • A scoring system evaluates the changes of Hydra attenuata and zebrafish embryos. • A potentially teratogenic substance can be easily classified.

Bone formation is a complex process regulated by a variety of pathways that are not yet fully understood. One of the proteins involved in multiple osteogenic pathways is TID (DNAJA3). The aim of this work was to study the association of TID with osteogenesis. Therefore, the expression profiles of the TID splice variants (TID-L, TID-I) and their protein products were analyzed during the proliferation and differentiation of bone marrow mesenchymal stromal cells (B-MSCs) into osteoblasts. As the reference, the hFOB1.19 cell line was used. The phenotype of B-MSCs was confirmed by the presence of CD73, CD90, and CD105 surface antigens on ~97% of cells. The osteoblast phenotype was confirmed by increased alkaline phosphatase activity, calcium deposition, and expression of ALPL and SPP1. The effect of silencing the TID gene on the expression of ALPL and SPP1 was also investigated. The TID proteins and the expression of TID splice variants were detected. After differentiation, the expression of TID-L and TID-I increased 5-fold and 3.7-fold, respectively, while their silencing resulted in increased expression of SPP1. Three days after transfection, the expression of SPP1 increased 7.6-fold and 5.6-fold in B-MSCs and differentiating cells, respectively. Our preliminary study demonstrated that the expression of TID-L and TID-I changes under differentiation of B-MSCs into osteoblasts and may influence the expression of SPP1. However, for better understanding the functional association of these results with the relevant osteogenic pathways, further studies are needed.

The HeLa 9903 cell line is used in the in vitro endocrine disruption assay (OECD 455). After treatment with an estrogen receptor agonist, the luminescence of these cells increases. However, some autoactivation of this test system may occur. The aim of this study was to try to explain the reason for the autoactivation of the HeLa 9903 cell line. Untreated cells were seeded on days with different moon phases and luminescence was measured the next day. Results from different moon phases were compared and statistically analyzed. An ELISA was also performed to check for the possible expression of 17β-estradiol. Statistical analysis revealed that the increase in luminescence in untreated cells was related to the phase of the moon. When the visibility of the moon was greater than approximately 87.7% (full moon), the likelihood of the cells autoactivating was greater. ELISA revealed that this change was not related to the production of 17β-estradiol by the cells (ligand-independent activation of the estrogen receptor). This is the first study to show a relationship between the phase of the moon and the activation of cells cultured in vitro. The results are important for planning the study using this test system.