99m-Technetium binding site in bone marrow mononuclear cells

Grazielle Dias Suhett, Sergio Augusto Lopes de Souza, Adriana Bastos Carvalho, Rachel de Pinho Rachid, Narcisa Leal da Cunha-E-Silva, Antonio Carlos Campos de Carvalho, Lea Mirian Barbosa da Fonseca, Regina Coeli dos Santos Goldenberg, Bianca Gutfilen

The increasing interest in 99m-Technetium ((99m)Tc)-labeled stem cells encouraged us to study the (99m)Tc binding sites in stem cell compartments. Bone marrow mononuclear cells were collected from femurs and tibia of rats. Cells were labeled with (99m)Tc by a direct method, in which reduced molecules react with (99m)Tc with the use of chelating agents, and lysed carefully in ultrasonic apparatus. The organelles were separated by means of differential centrifugation. At the end of this procedure, supernatants and pellets were counted and the percentages of radioactivity (MBq) bound to the different cellular fractions were determined. Percentages were calculated by dividing the radioactivity in each fraction by total radioactivity in the sample. The pellets were separated and characterized by their morphology on electron microscopy. The labeling procedure did not affect viability of bone marrow mononuclear cells. Radioactivity distribution in bone marrow mononuclear cell organelles, obtained in five independent experiments, was approximately 38.5 % in the nuclei-rich fraction; 5.3 % in the mitochondria-rich fraction; 2.2 % in microsomes and 54 % in the cytosol. Our results showed that most of the radioactivity remained in the cytosol; therefore this is an intracellular labeling procedure that has ribosomes unbound to membrane and soluble molecules as targets. However, approximately 39 % of the radioactivity remained bound to the nuclei-rich fraction. To confirm that cell disruption and organelle separation were efficient, transmission electron microscopy assays of all pellets were performed. Our results showed that most part of radioactivity was present in the cytosol fraction. More studies to understand the mechanisms involved in the cellular uptake of (99m)Tc in bone marrow cells are now ongoing.