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The molecular mechanism of apoptosis induced by xenogeneic cytotoxicity
I Fujiwara1, H Nakajima, H Yamagishi
1Second Department of Surgery, Kyoto Prefectural University of Medicine, Japan.
Abstract:
In order to clarify the role of natural killer (NK) cells in delayed xenograft rejection (DXR) of discordant xenotransplantation, we used in vitro xenogeneic combination of human NK cells and pig kidney target cells (PK15), and investigated the mechanism of xenogeneic cytotoxicity caused by human NK cells. In the presence of decomplemented human serum or human IgG, freshly isolated human peripheral blood lymphocytes (PBLs) caused both membrane (51Cr release) and DNA (3H release) damage on PK15. In contrast, only membrane damage was detected in the presence of normal human serum. To clarify the participation of perforin/granzymescell mediated cytotoxicity (P/G-CMC), when EGTA or concanamycin B (CMB) was added to the cytotoxicity assays, both cytotoxicities were completely inhibited by these drugs in a dose-dependent manner. In terms of the involvement of Fas/FasL-based cytotoxicity (F-CMC), while the cytotoxicity assays were performed in the presence of antagonistic anti-human FasL mAb, this antibody was not able to block the cytotoxicity. From these results, it is concluded that xenogeneic cytotoxicity is due to NK cell dependent ADCC (antibody-dependent cell-mediated cytotoxicity), and their effector mechanism can cause apoptosis on target cells via P/G-CMC.
Insights
Natural killer (NK) cells drive delayed xenograft rejection via antibody-dependent cell-mediated cytotoxicity. Their effector mechanism induces apoptosis in pig kidney cells through perforin/granzyme pathways, not Fas/FasL.
Area of Science:
- Immunology
- Transplantation Biology
- Cellular Cytotoxicity
Background:
- Delayed xenograft rejection (DXR) is a major hurdle in xenotransplantation.
- The precise role of natural killer (NK) cells in DXR remains incompletely understood.
- Investigating NK cell mechanisms is crucial for overcoming xenograft rejection.
Purpose of the Study:
- To elucidate the role of human NK cells in xenogeneic cytotoxicity against pig kidney cells.
- To determine the specific effector mechanisms employed by NK cells in this context.
- To clarify the contribution of antibody-dependent cell-mediated cytotoxicity (ADCC) and perforin/granzyme-mediated cytotoxicity (P/G-CMC).
Main Methods:
- In vitro co-culture assays using human peripheral blood lymphocytes (NK cells) and pig kidney cells (PK15).
- Assessment of target cell damage via 51Cr (membrane) and 3H (DNA) release.
- Inhibition studies using EGTA, concanamycin B (CMB), and anti-FasL monoclonal antibody (mAb).
Main Results:
- Human NK cells induced both membrane and DNA damage on pig kidney cells in the presence of decomplemented serum or human IgG.
- Cytotoxicity was significantly inhibited by EGTA and CMB, indicating perforin/granzyme-mediated cytotoxicity (P/G-CMC) involvement.
- Inhibition of Fas/FasL-based cytotoxicity (F-CMC) using anti-FasL mAb did not block the observed xenogeneic cytotoxicity.
Conclusions:
- Xenogeneic cytotoxicity against pig kidney cells is mediated by NK cell-dependent ADCC.
- The effector mechanism involves P/G-CMC, leading to apoptosis in target cells.
- NK cells play a significant role in DXR through these cytotoxic pathways.