Diglycolic Acid Is the Nephrotoxic Metabolite in Diethylene Glycol Poisoning Inducing Necrosis in Human Proximal Tubule Cells In Vitro

Abstract

Diethylene glycol (DEG), a solvent and chemical intermediate, can produce an acute toxic syndrome, the hallmark of which is acute renal failure due to cortical tubular degeneration and proximal tubular necrosis. DEG is metabolized to two primary metabolites, 2-hydroxyethoxyacetic acid (2-HEAA) and diglycolic acid (DGA), which are believed to be the proximate toxicants. The precise mechanism of toxicity has yet to be elucidated, so these studies were designed to determine which metabolite was responsible for the proximal tubule cell death. Human proximal tubule (HPT) cells in culture, obtained from normal cortical tissue and passaged 3-6 times, were incubated with increasing concentrations of DEG, 2-HEAA, or DGA separately and in combination for 48 h at pH 6 or 7.4, and various parameters of necrotic and apoptotic cell death were measured. DEG and 2-HEAA did not produce any cell death. DGA produced dose-dependent necrosis at concentrations above 25 mmol/l. DGA did not affect caspase-3 activity and increased annexin V staining only in propidium iodide-stained cells. Hence, DGA induced necrosis, not apoptosis, as corroborated by severe depletion of cellular adenosine triphosphate levels. DGA is structurally similar to citric acid cycle intermediates that are taken up by specific transporters in kidney cells. HPT cells, incubated with N-(p-amylcinnamoyl)anthranilic acid, a sodium dicarboxylate-1 transporter inhibitor showed significantly decreased cell death compared with DGA alone. These studies demonstrate that DGA is the toxic metabolite responsible for DEG-induced proximal tubular necrosis and suggest a possible transporter-mediated uptake of DGA leading to toxic accumulation and cellular dysfunction.

Cardiotoxicity testing of diglycolic acid using In Vitro and In Vivo models

“…The identification and characterization of DGA as a renal and hepatic toxicant has been unequivocally established using in vitro cellular, in vivo rat, and even human systems [3,[7][8][9][10]. A single case report of human exposure indicated that the heart could be negatively affected by DGA, but this effect has never been previously captured in vitro or in vivo animal models.…”

“…Interestingly, it has been found that during the chemical synthesis of CMC, a byproduct called diglycolic acid (DGA) is also made and that DGA is a difficult impurity to eliminate [2,3]. DGA is a small four-carbon molecule consisting of two carboxylic acids connected together by an ether linkage (PubChem CID: 8088).…”

A 15-year retrospective review of brake fluid exposures in children from a single poison center

“…Like the toxic alcohols methanol and ethylene glycol, the metabolites of DEG (and not DEG itself) are responsible for the majority of its toxicity. As with other toxic alcohols, however, the parent compound causes altered mental status [14]. Like other toxic alcohols, DEG undergoes metabolism by both alcohol dehydrogenase and aldehyde dehydrogenase ( Figure 7) to form 2-hydroxyethoxyacetic acid (HEAA) [6,15] and diglycolic acid (DGA).…”

“…Like other toxic alcohols, DEG undergoes metabolism by both alcohol dehydrogenase and aldehyde dehydrogenase ( Figure 7) to form 2-hydroxyethoxyacetic acid (HEAA) [6,15] and diglycolic acid (DGA). While HEAA appears to be responsible for the acidosis in DEG poisoning, DGA is the metabolite responsible for kidney injury [14].…”

Case Files of the University of California, San Francisco Medical Toxicology Fellowship: Seizures and a Persistent Anion Gap Metabolic Acidosis

“…Patients who survive initial toxicity may develop peripheral neuropathies, particularly of cranial nerve VII [21]. The exact cause of the metabolic acidosis and end-organ toxicity associated with DEG is unclear, but may be the result of accumulation of the DEG metabolites 2-hydroxyethoxyacetic acid and diglycolic acid [21][22][23]. Triethylene glycol (TEG) may also be found in brake fluid, though ingestions are rare.…”