In vivo and in vitro studies on the toxicity of Hoechst 33342 (Ho342). Implications for employing Ho342 for the isolation of haematopoietic stem cells.

The fluorescence dye Hoechst 33342 (Ho342) is employed for isolating early haematopoietic cells and the aim of this study was to evaluate the in vivo and in vitro toxicity of this compound. First, by employing a murine model we studied the influence of this dye on the morphology of the different organs of animals that have been injected intravenously with increasing doses of Ho342. Accordingly, we found that Ho342 at relatively low doses (0,3 M) caused morphological changes in the spleen and lungs and at higher doses (1,5 & 6 M) damaged also the liver. In contrast, kidneys appear to be relatively resistant to this dye. Next, since Ho342 is employed for isolating early haematopoietic cells by FACS, we have been looking for potential toxicity of this dye against normal human haematopoietic progenitors. Accordingly, CD34+ cells isolated from cord blood (CB) samples were exposed to increasing doses of Ho342 (0-50 microM). We found that the low concentration of Ho342 (10 microM) recommended for isolating HSC significantly inhibited the clonogenecity of human erythroid progenitors (BFU-E). The higher doses of Ho342 have also been toxic against normal human myeloid progenitors (CFU-GM). This study shows that Ho342 could potentially damage human cells. This fact should be considered whenever Ho342 has to be employed for isolating living cells.