Lymphokine-activated killer cells induced in vivo in mice showing IL-2 toxicity have cytoplasmic granules containing perforin and its hemolytic activity.
Cytoplasmic granules of LAK cells isolated from mice showing side effects of recombinant human IL-2 (rhIL-2) display BLT-serine esterase activity and calcium-dependent hemolytic activity and seem to be involved in the mechanisms of LAK cell-mediated rhIL-2 toxicity. In this report, the hemolytically active substance was partially purified from the LAK granules and shown to be mouse perforin. Mice were implanted with miniosmotic pumps filled with a dose of rhIL-2 that induced known toxicity. The LAK granules were isolated from LAK cell-rich subcutaneous tissue around the infusion sites. The hemolytically active substance in the granule preparation was solubilized by 2 M NaCl and passed through a butyl Sepharose column and then a DEAE Toyopearl column. In immunoblotting, the hemolytically active fractions reacted with an anti-mouse perforin mAb and the reaction depended on the activity. The molecular sizes of the perforin-positive bands were 66 kDa and 51 kDa under reducing and nonreducing conditions, respectively. These results confirmed the existence of mouse perforin and its hemolytic activity in the LAK granules of rhIL-2-treated mice, as has been shown for granules of cytotoxic T or NK cell clones in vitro.
Ueno
M
M
Developmental Research Laboratories, Shionogi, Osaka, Japan.
D000818:Animals; D003594:Cytoplasmic Granules / Q000502:physiology*; D005260:Female; D006460:Hemolysin Proteins / Q000633:toxicity*; D006863:Hydrogen-Ion Concentration; D007376:Interleukin-2 / Q000633:toxicity*; D015979:Killer Cells, Lymphokine-Activated / Q000502:physiology* / Q000648:ultrastructure; D008297:Male; D008562:Membrane Glycoproteins / Q000633:toxicity*; D051379:Mice; D008807:Mice, Inbred BALB C; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D051381:Rats; D017207:Rats, Sprague-Dawley
D006460:Hemolysin Proteins; D007376:Interleukin-2; D008562:Membrane Glycoproteins; D052899:Pore Forming Cytotoxic Proteins; D054353:Perforin