Activation of the N‐methyl‐d‐aspartate receptor is involved in glyphosate‐induced renal proximal tubule cell apoptosis

Abstract

Glyphosate-based herbicides have been used worldwide for decades and have been suggested to induce nephrotoxicity, but the underlying mechanism is not yet clear.In this study, we treated a human renal proximal tubule cell line (HK-2) with glyphosate for 24 hours at concentrations of 0, 20, 40 and 60 μM. Glyphosate was found to reduce cell viability and induce apoptosis and oxidative stress in a dose-dependent manner. Because the chemical structures of glyphosate and those of its metabolite AMPA are similar to glycine and glutamate, which are agonists of the N-methyl-Daspartate receptor (NMDAR), we investigated the potential role of the NMDAR pathway in mediating the proapoptotic effect of glyphosate on proximal tubule cells.We found that NMDAR1 expression, as well as intracellular Ca 2+ ([Ca 2+ ] i ) and reactive oxygen species (ROS) levels, increased after glyphosate treatment. Blocking NMDAR attenuated glyphosate-induced upregulation of [Ca 2+ ] i and ROS levels as well as apoptosis. Meanwhile, inhibition of [Ca 2+ ] i reduced glyphosate-induced ROS and apoptosis, and inhibition of ROS alleviated glyphosate-induced apoptosis. In mice exposed to 400 mg/kg glyphosate, the urine low molecular weight protein levels started to increase from day 7. Upregulation of apoptosis and NMDAR1 expression in renal proximal tubule epithelium and an imbalance of oxidant and antioxidative products were observed. These results strongly suggest that activation of the NMDAR1 pathway, together with its downstream [Ca 2+ ] i and oxidative stress, is involved in glyphosate-induced renal proximal tubule epithelium apoptosis.

Glutamate-Gated NMDA Receptors: Insights into the Function and Signaling in the Kidney

“…Leung et al [ 103 ], using MDCK cells and proximal tubule-like opossum kidney cells, additionally showed that immoderate activation of the NMDAR (10 mM glutamate), as well as the excessive blockade of this receptor with MK-801 or CPP led to harmful effects on survival of kidney cells [ 103 ]. Recent work from Gao et al [ 104 ] showed that NMDAR activation caused by glyphosate treatment led to increased oxidative stress and apoptotic death of renal epithelial cells, while blockade of NMDAR ameliorated glyphosate-induced cell damage [ 104 ].…”

“…Inhibition of NMDAR by MK-801 significantly reduced glomerular Nox activity, Nox-dependent O 2 production and lipid peroxidation induced by hyperhomocysteinemia (hHcys) in rat kidneys, suggesting an important role of the NMDAR in activation of Nox system in the kidney during hHcys and in the development of hHcys-induced glomerulosclerosis [ 59 ]. Furthermore, blockade of NMDAR attenuated glyphosate-induced increase of Ca 2+ influx and reduced activity of major endogenous antioxidant enzymes such as SOD, CAT and GSH-Px in HK-2 cells, strongly suggesting that activation of NMDAR, accompanied by Ca 2+ influx and oxidative stress, is involved in glyphosate-induced renal proximal tubule epithelium apoptosis [ 104 ]. Treatment of Sprague-Dawley rats with gentamicin led to an increase of endothelin type B receptor (ETBR) in the renal cortex and urinary nitrite concentration, while MK-801 managed to normalize the levels of urinary nitrites [ 102 ].…”

Glyphosate’s Synergistic Toxicity in Combination with Other Factors as a Cause of Chronic Kidney Disease of Unknown Origin

“…Gao et al (2019) [150] use a cell culture model to show that the kidney is damaged by glyphosate-based herbicides and that the renal proximal tubule may be a main target. Though the toxicity of commercial formulations is well established, the authors demonstrate that glyphosate, the active ingredient of glyphosate-based herbicides, injures renal tubule epithelial cells via the NMDAR1/calcium influx/oxidative stress pathway, both in vitro and in vivo.…”

Renal tubular injury induced by glyphosate combined with hard water: the role of cytosolic phospholipase A2

“…Previous evidence have implicated these eicosanoids in a host of inflammatory disorders, and thus cPLA 2 may play an important role in the development of inflammatory diseases (19)(20)(21). Furthermore, our previous study (11) demonstrated that oral exposure of glyphosate at a dose of 400 mg/kg in mice resulted in intracellular calcium ([Ca 2+ ] i ) overload and overproduction of reactive oxygen species (ROS). This suggested that cPLA 2 may be involved in renal tubular damage induced by glyphosate.…”

“…Our previous studies demonstrated that glyphosateinduced nephrotoxicity was dependent on the activation of N-methyl-D-aspartate receptors (NMDARs) expressed in the proximal tubular epithelium. This resulted in the elevation of cytoplasmic calcium ions (Ca 2+ ) and oxidative damage (11).…”