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Necrosis and apoptosis associated with distinct Ca2+ response patterns in target cells attacked by human natural
Y Oshimi1, K Oshimi, S Miyazaki
1Department of Physiology, Tokyo Women's Medical College, Japan.
Abstract:
1. During the process of cell death, rises in cytosolic Ca2+ concentration ([Ca2+]i) together with structural changes were investigated in target cells attacked by purified CD3-,CD16+ human natural killer (NK) cells. 2. In the target cell line K562, a rapid [Ca2+]i rise to 1-2 microM occurred a few minutes after NK cell-target cell contact, immediately followed by leakage of the Ca2+ indicator dye fura-2 from the cell. Cells were permeabilized, but their chromatin was not fragmented. The changes were basically consistent with those seen in necrosis induced by activated complement. 3. In the target cell line MOLT-4, which expressed the apoptosis-inducing surface antigen Fas much more strongly than K562, the majority of attacked cells displayed a [Ca2+]i rise to 0.7-1 microM followed by a slow decline, often associated with diminishing [Ca2+]i oscillations. As a whole, [Ca2+]i remained higher than 150 nM for at least 1.5-3 h (approximately 100 nM in control cells). 4. MOLT-4 cells attacked by NK cells became bubble shaped within 20 min of the main [Ca2+]i rise reaching its peak, and then both the cell and chromatin were fragmented into small pieces. These findings were basically consistent with those in apoptosis induced by a monoclonal antibody against the surface antigen Fas. 5. NK cells induced both necrosis and apoptosis in cell lines insensitive to NK cells in the presence of an antibody against the major histocompatibility complex class I (antibody-dependent cell-mediated cytotoxicity, ADCC). The distinct Ca2+ responses patterns described above corresponded to necrosis or apoptosis in different cells stimulated by the common ADCC pathway. 6. Human NK cells were found to be capable of inducing necrosis (membrane damage) or apoptosis (nuclear damage) depending on the target cell types. The characteristic Ca2+ response profile was a good indicator for distinguishing between the modes of cell death induced by the cytotoxicity of the killer cells.
Insights
Human natural killer (NK) cells induce necrosis or apoptosis in target cells. Distinct cytosolic calcium ([Ca2+]i) rise patterns indicate whether NK cells cause membrane damage (necrosis) or nuclear damage (apoptosis).
Area of Science:
- Immunology
- Cell Biology
- Cytotoxicity
Background:
- Natural killer (NK) cells are crucial for innate immunity, eliminating infected or cancerous cells.
- Cell death can occur via necrosis (uncontrolled) or apoptosis (programmed), with distinct cellular and molecular events.
- Cytosolic calcium concentration ([Ca2+]i) is a key signaling molecule in cellular processes, including cell death.
Purpose of the Study:
- To investigate the role of cytosolic calcium ([Ca2+]i) dynamics during NK cell-mediated target cell death.
- To correlate specific [Ca2+]i changes with distinct modes of cell death (necrosis vs. apoptosis).
- To understand how NK cells differentiate between necrotic and apoptotic pathways.
Main Methods:
- Co-culture of purified human NK cells with target cell lines (K562 and MOLT-4).
- Measurement of intracellular calcium ([Ca2+]i) using fura-2 dye.
- Microscopic observation of cell morphology and chromatin fragmentation.
- Induction of antibody-dependent cell-mediated cytotoxicity (ADCC) using antibodies against MHC class I.
Main Results:
- NK cell contact with K562 cells induced a rapid, high [Ca2+]i rise followed by membrane permeabilization (necrosis).
- NK cell interaction with MOLT-4 cells resulted in a sustained, moderate [Ca2+]i rise, leading to cell and chromatin fragmentation (apoptosis).
- Distinct [Ca2+]i response patterns were observed correlating with either necrosis or apoptosis, even under ADCC conditions.
Conclusions:
- Human NK cells can induce either necrosis or apoptosis depending on the target cell type.
- Specific patterns of cytosolic calcium ([Ca2+]i) flux serve as reliable indicators distinguishing NK cell-induced necrosis from apoptosis.
- Understanding these calcium dynamics provides insight into the mechanisms of cytotoxic lymphocyte-mediated cell death.