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Early acute necrosis and delayed apoptosis induced by methyl mercury in murine peritoneal neutrophils
Tsun-Cheng Kuo1, Shoei-Yn Lin-Shiau
1Institute of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan.
Abstract:
There is growing evidence that heavy metals in general, and mercurial compounds in particular, are immunotoxic. The purpose of this study was to explore the mechanism of MeHg in inducing cell death of mouse peritoneal neutrophils. In this paper we demonstrate that MeHg induces apoptosis and necrosis depending on MeHg concentration. In vitro exposure of mouse peritoneal neutrophils to MeHg resulted in a time- and concentration-dependent cell death. MeHg (15 microM) induced neutrophil necrosis in 13 min. The type of cell death was attributed to necrosis based on cells permeable to the fluorescent dye, propidium iodide and DNA appeared as a smear. With fura-2 microfluorimetric technique, we found that the entry of external Ca2+ into the cytosol played a crucial role in inducing cell necrosis by 15 microM MeHg. However, at lower concentrations, MeHg (10 microM)-induced apoptosis is confirmed by the observation of morphological features characterised by apoptotic bodies and fragmented DNA ladder. MeHg (10 microM) caused an immediate fall in pHi as revealed by the pH-sensitive fluorescent probe 2'7'-bis (carboxyethyl)-5(6)-carboxyfluorescein. We have found that MeHg induced cellular acidification prior to DNA fragmentation so as the other two apoptosis-inducing agents (ZnCl(2) and EGTA). Furthermore, acid-activated endonuclease was increased by MeHg in neutrophils, which we considered to play a possible role in chromatin digestion leading to apoptosis. Taken together, these findings indicate that MeHg induces necrosis at higher concentrations by a rapid increase of [Ca2+]i and apoptosis at lower concentrations by acid activation of endonuclease.
Insights
Methylmercury (MeHg) induces different cell death pathways in neutrophils. High MeHg concentrations cause necrosis via calcium influx, while lower concentrations induce apoptosis through cellular acidification and endonuclease activation.
Area of Science:
- Immunotoxicology
- Cellular Biology
- Environmental Health
Background:
- Heavy metals, particularly mercury compounds, are increasingly recognized for their immunotoxic effects.
- Understanding the specific mechanisms of mercury-induced cell death is crucial for assessing its health risks.
Purpose of the Study:
- To investigate the mechanisms by which methylmercury (MeHg) induces cell death in mouse peritoneal neutrophils.
- To differentiate between apoptosis and necrosis pathways triggered by MeHg exposure.
Main Methods:
- In vitro exposure of mouse peritoneal neutrophils to varying concentrations of MeHg.
- Assessment of cell death using propidium iodide staining and DNA fragmentation analysis (gel electrophoresis).
- Measurement of intracellular calcium ([Ca2+]i) and intracellular pH (pHi) using fura-2 and BCECF probes, respectively.
- Evaluation of acid-activated endonuclease activity.
Main Results:
- MeHg induced both apoptosis and necrosis in a concentration-dependent manner.
- Neutrophil necrosis occurred rapidly at high MeHg concentrations (15 microM), associated with increased cytosolic calcium ([Ca2+]i).
- Apoptosis was observed at lower MeHg concentrations (10 microM), characterized by cellular acidification, DNA fragmentation, and increased acid-activated endonuclease activity.
Conclusions:
- MeHg triggers distinct cell death mechanisms in neutrophils based on concentration.
- Necrosis is mediated by rapid calcium influx at higher MeHg levels.
- Apoptosis is induced by MeHg at lower concentrations via cellular acidification and subsequent activation of endonuclease.
