Visualization of the antioxidative effects of melatonin at the mitochondrial level during oxidative stress‐induced apoptosis of rat brain astrocytes

Abstract

Oxidative stress-induced mitochondrial dysfunction has been shown to play a crucial role in the pathogenesis of a wide range of diseases. Protecting mitochondrial function, therefore, is vital for cells to survive during these disease processes. In this study, we demonstrate that melatonin, a chief secretory product of the pineal gland, readily rescued mitochondria from oxidative stress-induced dysfunction and effectively prevented subsequent apoptotic events and death in rat brain astrocytes (RBA-1). The early protection provided by melatonin in mitochondria of intact living cells was investigated by the application of time-lapse conventional, confocal, and multiphoton fluorescent imaging microscopy coupled with noninvasive mitochondria-targeted fluorescent probes. In particular, we observed that melatonin effectively prevented exogenously applied H2O2-induced mitochondrial swelling in rat brain astrocytes at an early time point (within 10 min) and subsequently reduced apoptotic cell death (150 min later). Other early apoptotic events such as plasma membrane exposure of phosphatidyl serine and the positive YOPRO-1 staining of the early apoptotic nucleus were also prevented by melatonin. A mechanistic study at the mitochondrial level related to the early protection provided by melatonin revealed that the indole molecule significantly reduced mitochondrial reactive oxygen species (ROS) formation induced by H2O2 stress. Melatonin also prevented mitochondrial ROS generation caused by other organic hydroperoxides including tert-butyl hydroperoxide and cumene hydroperoxide. This antioxidative effect of melatonin is more potent than that of vitamin E. Via its ability to reduce mitochondrial ROS generation, melatonin prevented H2O2-induced mitochondrial calcium overload, mitochondrial membrane potential depolarization, and the opening of the mitochondrial permeability transition (MPT) pore. As a result, melatonin blocked MPT-dependent cytochrome c release, the downstream activation of caspase 3, the condensation and karyorrhexis of the nucleus and apoptotic fragmentation of nuclear DNA. Thus, the powerful mitochondrial protection provided by melatonin reinforces its therapeutic potential to combat a variety of oxidative stress-induced mitochondrial dysfunctions as well as mitochondria-mediated apoptosis in various diseases.

Blue Light Induces Mitochondrial DNA Damage and Free Radical Production in Epithelial Cells

“…Cellular dysfunction via this mechanism is likely to reflect in decreased respiratory function and the accumulation of mtDNA defects. By contrast, more severe ROS damage to mitochondria elicits the apoptosis pathway (41)(42)(43)(44). Generation of ROS by photo-irradiation can lead to mitochondrial swelling (45), increased mitochondrial calcium levels (43), and Bcl-2 inhibition and mitochondrial matrix caspase-2 and -9 activation (44).…”

“…By contrast, more severe ROS damage to mitochondria elicits the apoptosis pathway (41)(42)(43)(44). Generation of ROS by photo-irradiation can lead to mitochondrial swelling (45), increased mitochondrial calcium levels (43), and Bcl-2 inhibition and mitochondrial matrix caspase-2 and -9 activation (44). To complicate matters further, there is some evidence that the ROS-mtDNA damage axis can be regulated via growth factors and altered transcription factor activities (12,46).…”

Alzheimer’s Disease: Focus on the Neuroprotective Role of Melatonin

“…First, it increases the activities of mitochondrial respiratory complexes I and IV in a time dependent manner [57]. This effect of melatonin, namely the improvement of electron transport capacity in mitochondria, is remarkable as these effects are observed in senescence-accelerated mice [58]. In addition to this, other processes perturbing the mitochondrial membrane potential such as calcium overload are also antagonized by melatonin.…”

Melatonin for Protection Against Ionizing Radiation

“…The different regulatory mechanisms of apoptosis and their modification by treatment with melatonin were tested in different cells after irradiation. It is found that the mitochondrial pathway was strongly influenced by melatonin by reducing mitochondrial ROS generation and calcium release as well as inhibiting the opening of the MPTP as shown in rat brain astrocytes (Jou et al 2004), mouse striatal neurons (Andrabi et al 2004) and rat cerebellar granule neurons (Han et al 2006). Moreover, the prevented decreases in the mitochondrial membrane potential resulted from irradiation suggests that melatonin, due to its physiochmeical characters crosses the bloodbrain barrier and biological membranes to easily reach mitochondria, stabilizes oxidative stress-mediated dysfunction and integrity of mitochondria by preserving its membrane potential and increasing the efficiency of mitochondrial electron transfer chain and ATP synthesis (Acuna-Castroviejo et al 2001).…”