Developmental effects of exposure to ultraviolet B radiation on the freshwater prawn Macrobrachium olfersi: Mitochondria as a target of environmental UVB radiation.

In South America, increased UVB radiation has become an important environmental issue that is potentially threatening aquatic ecosystems. Considering that species exhibit different degrees of sensitivity to UVB radiation and that embryos are more sensitive than organisms at later life stages, the aim of this study was to characterize the effects of UVB radiation on subcellular compartments of embryos of the freshwater prawn Macrobrachium olfersi. This species lives and reproduces in clear and shallow waters, where UV radiation can fully penetrates. Embryos were irradiated with a UVB 6W lamp for 30min and examined after 1h, 12h, 24h and 48h of exposure. The irradiance of the UVB used simulates the UV radiation that embryos receive in the natural environment. The subcellular compartment most affected by the UVB radiation was the mitochondria, which exhibited a circular shape, a decrease in mitochondrial cristae, rupture of membranes and a morphology compatible with fission. These impairments were observed simultaneously with increased ROS production, just after 1h of UVB exposure. Thus, we investigated proteins related to mitochondrial fission (Drp-1) and fusion (Mfn-1), which are essential to cell maintenance. We found a significant increase in Drp-1 expression at all analyzed time-points and a significant decrease in Mfn-1 expression only after 24h of UVB exposure. Additionally, a decrease in embryonic cell viability was verified via the mitochondrial integrity assay. To conclude, we observed important mitochondrial dysfunctions against the environmental stress caused by UVB radiation. Moreover, the cellular responses found are critical and should not be disregarded, because they impact embryos that can potentially compromise the aquatic ecosystems.

Effect of UVB radiation exposure in the expression of genes and proteins related to apoptosis in freshwater prawn embryos.

Our previous studies showed that embryos of the freshwater prawn Macrobrachium olfersii exposed to ultraviolet B (UVB) radiation exhibited DNA damage, excessive ROS production, mitochondrial dysfunction and increased hsp70 expression, which are able, independently or together, to induce apoptosis. Thus, we attempted to elucidate some key apoptosis-related genes (ARG) and apoptosis-related proteins (ARP) and their expression during different stages of embryonic development, as well as to characterize the chronology of ARG expression and ARP contents after UVB radiation insult. We demonstrate that p53, Bax and Caspase3 genes are active in the embryonic cells at early embryonic developmental stages, and that the Bcl2 gene is active from the mid-embryonic stage. After UVB radiation exposure, we found an increase in ARP such as p53 and Bak after 3h of exposure. Moreover, an increase in ARG transcript levels for p53, Bax, Bcl2 and Caspase3 was observed at 6h after UVB exposure. Then, after 12h of UVB radiation exposure, an increase in Caspase3 gene expression and protein was observed, concomitantly with an increased number of apoptotic cells. Our data reveal that ARG and ARP are developmentally regulated in embryonic cells of M. olfersii and that UVB radiation causes apoptosis after 12h of exposure. Overall, we demonstrate that embryonic cells of M. olfersii are able to active the cell machinery against environmental changes, such as increased incidence of UVB radiation in aquatic ecosystems.