Transient poration and cell surface receptor removal from human lymphocytes in vitro by 1 MHz ultrasound

TRANSIENT PORATION AND CELL SURFACE RECEPTOR REMOVAL FROM HUMAN LYMPHOCYTES IN VITRO BY 1 MHZ ULTRASOUND.
AA Brayman, ML Coppage, S Vaidya, MW Miller. School of Medicine and Denistry, University of Rochester, Rochester, NY and The University of Texas Medical Branch at Galveston, Galveston, TX.

The study objective was to gain insight into ultrasound-induced, sub-lytic cell surface modifications. Two primary hypotheses were tested by flow cytometric methods; viz., sonication will: (1) remove all or part of a specific cell surface marker in lymphocytes surviving insonation, and (2) induce transient pores in the cell membranes of some surviving cells. RPMI 1788 human lymphocytes were exposed in vitro to 1 MHz, continuous wave ultrasound ([US];~8 W/cm² I_SP) for 30 s, which lysed ~50% of the cells. Insonation: (1) altered cell morphology, increasing the population of cells of reduced size but high structure [R2], many of which were nonviable, and diminishing the population of cells of large size and high structure [R1], most of which were viable, (2) diminished the fluorescence signal from the pan B lymphocyte marker CD19 in population R1 and R2 to equivalent extents, and (3) increased by ~7-fold the number of transiently permaeabilized cells in R1, as evidenced by simultaneous uptake of popidium iodide and fluorescein diacetate. The results indicate that US-induced CD19 removal from R1 cells can occur without accompanying gross membrane loss. The cell morphology/mortality shifts indicate that the US-induced morphologic change is associated with lethal membrane poration, suggesting that the diminished CD19 fluorescence signal from insonated R2 cells arises partly by simultaneous loss of membrane fragments, CD19, and cytoplasm.