In Vitro Evaluation of Graft-versus-Graft Alloreactivity as a Tool to Identify the Predominant Cord Blood Unit before Double Cord Blood Transplantation

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Presentation transcript:

In Vitro Evaluation of Graft-versus-Graft Alloreactivity as a Tool to Identify the Predominant Cord Blood Unit before Double Cord Blood Transplantation Antonia Moretta, Gabriella Andriolo, Daniela Lisini, Miryam Martinetti, Annamaria Pasi, Paolo Rebulla, Davide Soligo, Rosaria Giordano, Lorenza Lazzari, Rita Maccario Biology of Blood and Marrow Transplantation Volume 18, Issue 7, Pages 1108-1118 (July 2012) DOI: 10.1016/j.bbmt.2011.12.586 Copyright © 2012 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 1 Experimental design scheme. CBMCs were obtained from each CB unit pair (CB-A and CB-B). (A) In experiments 1-5, a portion of CBMCs from each CB unit were used to obtain PHA-TLCs, and the remaining portion was used to carry out primary and secondary two-way MLCs. Effector cells harvested from MLCs were tested for DNA chimerism, cell subset recovery, and alloantigen-specific cytotoxic activity toward both CB-A and CB-B PHA-TLCs. (B) In experiments 6-8, a portion of CBMCs from each CB unit was used to select CD34+ cells and perform single and mixed CB-HP expansion cultures and to obtain PHA-TLCs and carry out a secondary two-way MLC. Effector cells harvested from MLCs were tested for DNA chimerism, cell subset recovery, and alloantigen-specific cytotoxic activity toward both PHA-TLCs and expanded CB-HPs, and single and mixed CB-HP expansion cultures were tested for DNA chimerism and cell subset recovery. Biology of Blood and Marrow Transplantation 2012 18, 1108-1118DOI: (10.1016/j.bbmt.2011.12.586) Copyright © 2012 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 2 Alloantigen-specific cytotoxic activity generated in primary and secondary two-way MLCs. Cytotoxic activity was measured toward PHA-TLC targets obtained from CBMCs (A and B) or PBMCs (C and D) for each of the two subjects assessed in the two-way MLC. (A) Experiment 2: CB-2A/CB-2B pair. (B) Experiment 5: CB-5A/CB-5B pair, primary MLC. Results from the secondary two-way MLC are not available because of the very low cell recovery after primary culture that precluded the possibility of carrying out the assay. (C) Experiment 1: PB-1A/PB-1B pair. (D) Experiment 3: PB-3A/PB-3B pair. Results are expressed as LU10/million effector cells. Dotted columns represent CB-A and PB-A targets; black columns, CB-B and PB-B targets. Biology of Blood and Marrow Transplantation 2012 18, 1108-1118DOI: (10.1016/j.bbmt.2011.12.586) Copyright © 2012 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 3 Alloantigen-specific cytotoxic activity of secondary two-way MLCs at different E:T ratios. Cytotoxic activity was measured toward PHA-TLC targets obtained from CBMCs (A) or PBMCs (B) for each of the two subjects assessed in the two-way MLC. (A) Experiment 3: CB-3A/CB-3B pair. Black square, CB-3A PHA-TLC target; empty square, CB-3B PHA-TLC target. (B) Experiment 2: PB-2A/PB-2B pair. Black triangle, PB-2A PHA-TLC target; empty triangle, PB-2B PHA-TLC target. Results are expressed as percent-specific lysis measured in an 18-hour (CBMCs) or a 5-hour (PBMCs) 51Cr-release assay. Biology of Blood and Marrow Transplantation 2012 18, 1108-1118DOI: (10.1016/j.bbmt.2011.12.586) Copyright © 2012 American Society for Blood and Marrow Transplantation Terms and Conditions

Figure 4 Alloantigen-specific cytotoxic activity generated in secondary two-way MLCs. Cytotoxic activity was measured toward both PHA-TLC and expanded CB-HP targets obtained from CBMCs for each of the two subjects used in the two-way MLC. (A) Experiment 6: CB-6A/CB-6B pair. (B) Experiment 7: CB-7A/CB-7B pair. (C) Experiment 8: CB-8A/CB-8B pair. Results are expressed as LU10/million effector cells. Dotted columns represent CB-A PHA-TLC targets; black columns, CB-B PHA-TLC targets; striped columns, CB-A expanded HP targets; gray columns, CB-B expanded HP targets. Biology of Blood and Marrow Transplantation 2012 18, 1108-1118DOI: (10.1016/j.bbmt.2011.12.586) Copyright © 2012 American Society for Blood and Marrow Transplantation Terms and Conditions