Abstract
Cadmium at environmental concentrations is a risk factor for many diseases, including cardiovascular and neurodegenerative diseases, in which macrophages play an important role. The aim of this study was to evaluate the effects of cadmium at low environmental (nanomolar) concentrations on apoptotic processes in THP-1(acute monocytic leukemia cells line)-derived macrophages, with special focus on mitochondrial events involved. Macrophages were incubated with various cadmium chloride (CdCl2) solutions for 48 h at final concentrations of 5 nM, 20 nM, 200 nM and 2 µM CdCl2. Cell viability was measured using flow cytometry. Flow cytometric measurement (annexin V/FITC (annexin V/fluorescein isothiocyanate) and PI (propidium iodide) double staining) was used to quantify the extent of apoptosis. Fluorescence and confocal microscopy were used for imaging of apoptosis process. Changes in mitochondrial membrane potential were monitored using cytofluorimetry after cell staining with JC-1(5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazol-carbocyane iodide) probe. Mitochondrial ROS (reactive oxygen species) levels were measured cytofluorimetrically after incubation of cells with mitochondrial superoxide indicator (MitoSOX) red fluorescent marker. The mRNA expression of Bcl-2 and Bax was analysed with qRT-PCR. Our study demonstrates that cadmium, even at low environmental concentrations, exerts mitochondrial toxicity in THP-1 macrophages. Forty-eight-hour exposure to very low concentrations reduces cell viability and results in cell death by apoptosis and necrosis. The decrease in mitochondrial membrane potential, increased ROS production, increased Bax and decreased Bcl-2 mRNA expression are mitochondrial events involved in cadmium-induced apoptosis.
Highlights
Cadmium is a toxic, mutagenic and carcinogenic heavy metal constituting substantial threat to public health [1,2,3]
THP-1 macrophages were exposed to varying concentrations of cadmium chloride (5 nM, 20 nM, 200 nM and 2 μM) for 48 h
It appeared that THP-1 macrophage viability decreased with increasing cadmium concentration, the significant reduction in cell viability was observed at the highest applied cadmium concentration, i.e., 2 μM Cd (3% reduction vs. control, p < 0.05) (Figure 1)
Summary
Mutagenic and carcinogenic heavy metal constituting substantial threat to public health [1,2,3]. Cadmium was suggested to cause atherosclerosis, the basis of most cardiovascular diseases [5]. A number of studies have suggested that cadmium induces apoptosis in several cell types [6,7,8,9,10,11,12,13,14,15,16,17,18,19], including monocytes and macrophages [20,21,22,23,24,25,26,27,28]. Different apoptotic pathways may be induced by cadmium in different cell types and depending on the cell treatment conditions [6]. The mitochondria are thought to be the primary target in cadmium-induced apoptosis [11]. A very important role in the regulation of these events is played by pro-apoptotic (Bax, Bak, Bid, Bim, Bad, Noxa) and anti-apoptotic (Bcl-2, Bcl-xL, Mcl-1) members of the Bcl-2 family [29]
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