RELATIONSHIP OF DECREASED CHEMOTAXIS OF TECHNETIUM-99M-HMPAO-LABELED LYMPHOCYTES TO APOPTOSIS

The purpose of this study was to evaluate chemotaxis and its relations hip to apoptosis in Tc-99m-HMPAO-labeled lymphocytes. Methods: Periphe ral lymphocytes, obtained from 12 healthy volunteers using lymphoprep, were divided in three equal fractions. One fraction was used as the c ontrol, one was labeled with cold HMPAO and one was labeled with 1.5 m Ci (55.5 Mbq) Tc-99m-HMPAO. Chemotaxis of T-lymphocytes was measured b y the Boyden microchamber technique (BMA) (n = a) using human monocyte chemotactic protein-3 (MCP-3) as chemoattractans. A chemotactic index was calculated as the number of HMPAO and Tc-99m-HMPAO-labeled cells that migrated towards the MCP-3 solution, divided by the number of non labeled migrated lymphocytes. Apoptosis evaluation (n = 10) of unlabel ed, HMPAO-labeled and Tc-99m-HMPAO-labeled cells was performed using f lowcytometry (FCM) forward light scatter analysis, 90O light scatter a nalysis, fluorescein-isothiocyanate (FITC)-labeled Annexin V and dye e xclusion of propidium iodide. Results: Chemotaxis of Tc-99m-HMPAO-labe led T-lymphocytes was found to be reduced by approximately 31% (migrat ion index of 0.69) (p = 0.01) as compared to both unlabeled and HMPAO- labeled lymphocytes, both the latter showing no difference in migratio n index. Whereas the mean percentages apoptotic lymphocytes in the unl abeled, 18.5%, and HMPAO-labeled fraction, 16.6%, were more or less co mparable (p = 0.1), the mean percentage apoptotic cells in the Tc-99m- HMPAO-labeled fraction was 51.8%, yielding a difference of 33.3% betwe en Tc-99m-HMPAO-labeled and unlabeled cells (p = 0.003). The procentua l concordance between apoptotic cells (33.3%) and chemotactic impaired cells (31%) in the Tc-99m-HMPAO-labeled fraction may be explained by the formation of a rigid cytoskeleton early in the apoptotic process t hat may theoretically limit chemotaxis. Conclusion: Using the BMA, che motaxis of Tc-99m-HMPAO-labeled lymphocytes was round to be reduced by approximately 31%. Furthermore, the percentage apototic lymphocytes i nduced by irradiation after labeling with Tc-99m-HMPAO concurs well wi th the percentage of chemotaxis impaired cells.