Potential Mechanisms of Methylglyoxal‐Induced Human Embryonic Kidney Cells Damage: Regulation of Oxidative Stress, DNA Damage, and Apoptosis

Abstract

Methylglyoxal (MGO) is a reactive carbonyl species that can cause cellular damage and is closely related to kidney disease, especially diabetic nephropathy. The toxic effect of MGO (0.5, 1, and 2 mM) on human embryonic kidney (HEK293) cells and its underlying mechanisms were explored in this study. Cell viability, apoptosis and the signaling pathways were measured with MTT, fluorescent staining and western blot experiments, the results showed that MGO could induce oxidative stress and cell inflammation, the level of reactive oxygen species (ROS) increased, and p38MAPK, JNK and NF‐κB signaling pathways were activated. Meanwhile, MGO also induced DNA damage. The expression of DNA oxidative damage marker 8‐hydroxy‐2’‐deoxyguanosine (8‐OHdG) increased, the expression of double‐strand break marker γH2AX increased significantly, and ATM/Chk2/p53 DNA damage response signaling pathway was activated. Furthermore, the expression of the receptor for advanced glycation end products (RAGE) also increased. Finally, mitochondrial membrane potential (MMP) decreased, fluorescence intensity of Hoechst33258 increased, and the protein expression ratio of Bax/Bcl‐2 increased significantly after the treatment of MGO. These results demonstrated that MGO might induce HEK293 cells damage by regulating oxidative stress, inflammation, DNA damage, and cell apoptosis, which revealed the specific mechanisms of MGO‐induced damage to HEK293 cells.

Investigating the molecular mechanisms of glyoxal‐induced cytotoxicity in human embryonic kidney cells: Insights from network toxicology and cell biology experiments

“…The results suggested that glyoxal could induce DNA damage, DNA double‐strand breakage, and trigger the DNA damage response ATM/Chk2/p53 signaling pathway. Microcystin‐LR (MC‐LR) and methylglyoxal have been reported to induce increased expression of γ‐H2AX and 8‐OHdG in cells, and up‐regulate the levels of ATM, CHK2, and p53 proteins to activate ATM/Chk2/p53 pathway, which were consistent with our results 24,59 …”

“…Study indicated that overexpression of RAGE could exacerbate mouse glomerular endothelial cells dysfunction and inflammation response, exacerbating the progression of diabetic nephropathy 29 . Similarly, methylglyoxal could induce apoptosis in human embryonic kidney cells and aggravate the development of diabetic nephropathy 24 . These results were similar to our prediction, indicating that the prediction was reliable.…”

“…The results suggested that glyoxal induced ROS production in HEK293 cells, resulting in oxidative stress in HEK293 cells. Methylglyoxal was also shown to increase ROS levels in HEK293 cells 24 . In fact, kidney disease partly occurs due to oxidative stress, manifested as the cumulative effect of ROS leading to cellular damage 37 .…”

“…The results proved that oxidative stress and apoptosis played essential roles in glyoxal toxicity. Oxidative stress and apoptosis are involved in many physiopathological conditions and are considered a crucial regulatory targets 24,25,27 . Previous study has shown that paraquat induced kidney damage through oxidative stress and apoptosis 28 …”

“…HEK293 cells were cultured in 12‐well plates (8 × 10 4 cells/plate) and treated with 0.5, 1, and 2 mM glyoxal. After that, the cells were fixed with cell fixative solution for 30 min, and then 500 μl Hoechst 33258 fluorescent dye was added for incubation in the dark for 10 min 24 . Then, the cells were washed twice with PBS and observed with a fluorescence microscope.…”