Osteocytes are key players in regulating bone mass maintenance and are therefore a major research focus in the field of bone and mineral biology. Analysing osteocyte function often requires in vivo investigations. Therefore, promoter fragments of osteocyte-specific genes i.e. SOST or the 8kb and 10kb fragments of the Dentin-matrix-protein 1 (Dmp1) were used to overexpress genes of interest or Cre-recombinase for conditional deletion studies using the Cre-loxP system. While these tools have been very useful to investigate osteocyte biology, evidence emerged suggesting that these promoters are not osteocyte-specific, which is very important for the data interpretation. To further investigate the specificity of a supposedly osteocyte-specific Cre-loxP-system, we crossed the 8kb-Dmp1-Cre mice with Ai9 reporter mice in which a loxP-flanked STOP cassette prevents the expression of tdTomato (Dmp1-Cre+;Ai9T/wt). At 8-weeks of age, male mice were sacrificed and various organs were harvested. Tibiae were decalcified and all tissues were paraffin-embedded, followed by tomato detection using immunohistochemistry. In bone, tomato was strongly expressed in all osteocytes and osteoblasts covering endocortical and trabecular surfaces, while no expression was found in articular cartilage of Dmp1-Cre+;Ai9T/wt mice compared to Dmp1-Cre-;Ai9T/wt and Dmp1-Cre+;Ai9wt/wt control animals. Furthermore, we detected tomato expression in muscle, brain, testis, and in vessels in the heart, spleen, lung, and intestine. We did not observe tomato expression in the kidney, liver, fat, or skin. These results indicate that in the 8kb-Dmp1-Cre;Ai9 reporter system, Cre-mediated recombination occurring during mouse development and growth is not restricted to osteocytes, but also takes place in other osteolineage cells and in several organs known to be functionally involved in the regulation of bone homeostasis. Our findings therefore suggest that despite the great usefulness of conditional gene deletion systems, the expression pattern of the gene of interest should be determined carefully and the findings need to be interpreted accordingly.
Disclosure: The authors declared no competing interests.