MeHg Developing Exposure Causes DNA Double-Strand Breaks and Elicits Cell Cycle Arrest in Spinal Cord Cells

Abstract

The neurotoxicity caused by methylmercury (MeHg) is well documented; however, the developmental neurotoxicity in spinal cord is still not fully understood. Here we investigated whether MeHg affects the spinal cord layers development. Chicken embryos at E3 were treated in ovo with 0.1 μg MeHg/50 μL saline solution and analyzed at E10. Thus, we performed immunostaining using anti-γ-H2A.X to recognize DNA double-strand breaks and antiphosphohistone H3, anti-p21, and anti-cyclin E to identify cells in proliferation and cell cycle proteins. Also, to identify neuronal cells, we used anti-NeuN and anti-βIII-tubulin antibodies. After the MeHg treatment, we observed the increase on γ-H2A.X in response to DNA damage. MeHg caused a decrease in the proliferating cells and in the thickness of spinal cord layers. Moreover, we verified that MeHg induced an increase in the number of p21-positive cells but did not change the cyclin E-positive cells. A significantly high number of TUNEL-positive cells indicating DNA fragmentation were observed in MeHg-treated embryos. Regarding the neuronal differentiation, MeHg induced a decrease in NeuN expression and did not change the expression of βIII-tubulin. These results showed that in ovo MeHg exposure alters spinal cord development by disturbing the cell proliferation and death, also interfering in early neuronal differentiation.

Oxidative Biochemistry Disbalance and Changes on Proteomic Profile in Salivary Glands of Rats Induced by Chronic Exposure to Methylmercury

“…Studies appoint that MeHg induces multiple effects within cell homeostasis. MeHg is able to induce cell death [ 37 ], impairment on cell proliferation and differentiation [ 38 ], disorganization of cytoskeleton [ 39 , 40 ], DNA mutation [ 41 ], impairments on transcriptome [ 42 ], and proteome [ 43 ]. In addition, according to the literature, these damages on biological processes and structures could be associated with oxidative/nitrosative biochemistry misbalance due to oxidative stress.…”

MeHg Causes Ultrastructural Changes in Mitochondria and Autophagy in the Spinal Cord Cells of Chicken Embryo

“…In response to such damage, other types of programmed cell death can be triggered, for example, death by autophagy, previously observed in systems exposed to heavy metals [ 46 – 48 ]. We observed DNA damage associated with cell death in the spinal cord of chicken embryos in an earlier study [ 30 ]. Although we did not observe morphological changes, the reduction in the thickness of the spinal cord layers was significant, demonstrating MeHg developing neurotoxicity, even at a very low dose.…”

“…Studies of developmental neurotoxicity generally use brain structures (brain, cerebellum, and hippocampus) as an organ model, given the well-documented behavioral (learning and memory deficit) and motor changes caused by MeHg poisoning [ 16 , 17 , 21 , 27 – 29 ]. However, the spinal cord is also an interesting model for neurodevelopmental toxicity studies because its structural organization is less complex than the encephalon and it has fewer tissue layers, allowing observation of the neurotoxic effects on cells at different stages of differentiation [ 14 , 15 , 30 ].…”

“…The effects of MeHg can be observed in different stages of neurogenesis; these effects can be observed in neural progenitors at the initial stage [ 31 ] which had reduced proliferation after exposure to the metal [ 15 , 20 , 30 , 32 ] in later stages; when the neural and glial lineages are being established, MeHg disrupts the cell differentiation process, causing alterations in the expression of genes related to maintaining the characteristics of the neuronal and glial cell lineages [ 15 , 33 , 34 ]. An increase in programmed cell death also occurs in neural cells exposed to MeHg [ 15 , 31 , 33 , 35 ], especially in the precursors that appear to be more susceptible than more differentiated cells [ 21 , 31 ].…”

“…Mitochondria appear to be an important target of MeHg cytotoxicity and considering that developing nervous tissue makes high energetic demands, damage to the mitochondria may result in risks to cell survival [ 13 , 20 , 27 , 30 ].…”