Probenecid and N-Acetylcysteine Prevent Loss of Intracellular Glutathione and Inhibit Neuronal Death after Mechanical Stretch Injury In Vitro

Abstract

Probenecid and N-acetylcysteine (NAC) can preserve intracellular levels of the vital antioxidant glutathione (GSH) via two distinct biochemical pathways. Probenecid inhibits transporter-mediated GSH efflux and NAC serves as a cysteine donor for GSH synthesis. We hypothesized that probenecid and NAC alone would maintain intracellular GSH concentrations and inhibit neuronal death after traumatic stretch injury, and that the drugs in combination would produce additive effects. Sex-segregated rat primary cortical neurons were treated with probenecid (100 μM) and NAC (50 μM), alone and in combination (Pro-NAC), then subjected to mechanical stretch (10s strain rate, 50% membrane deformation). At 24 h, both probenecid and NAC inhibited trauma-induced intracellular GSH depletion, lactate dehydrogenase (LDH) release, and propidium iodide (PI) uptake in both XY- and XX-neurons. Combined Pro-NAC treatment was superior to probenecid or NAC alone in maintenance of intracellular GSH and neuronal death assessed by PI uptake. Interestingly, caspase 3 activity 24 h after mechanical trauma was more prominent in XX-neurons, and treatment effects (probenecid, NAC, and Pro-NAC) were observed in XX- but not XY-neurons; however, XY-neurons were ultimately more vulnerable to mechanical stretch-induced injury than their XX counterparts, as was evidenced by more neuronal death detected by LDH release and PI uptake. In addition, after stretch injury in HT22 hippocampal cells, both NAC and probenecid were highly effective at reducing oxidative stress detected by dichlorofluorescein fluorescence. These in vitro data support further testing of this drug combination in models of traumatic neuronal injury in vivo.

Dimethyl fumarate treatment after traumatic brain injury prevents depletion of antioxidative brain glutathione and confers neuroprotection

“…In vitro , loss of cellular GSH after stretch injury in primary cortical neurons was reported to cause cell death and was prevented via preservation of intracellular GSH levels, inhibition of transporter‐mediated GSH efflux, and the cysteine donor N‐acetylcysteine (Du et al . ). Cysteine is the rate‐limiting component of de novo GSH synthesis.…”

Histomorphological evaluations on the frontal cortex extrapyramidal cell layer following administration of N-Acetyl cysteine in aluminum induced neurodegeneration rat model

“…We observed that seven days after Aluminum treatment cessation, the extrapyramidal neurons were able to endogenously produce glutathione reported to be ubiquitously synthesized in all mammalian cells (Shahripour et al 2014) as compared with Group B, this endogenous glutathione, mediates neuron repair by mobbing off ROS and up regulating brain energy metabolism to sustain the stressed neuron repair process. Activation of glial cells may reduce or delay the progression of neurodegeneration (Du et al 2016;Durieux et al 2015) thereby ameliorating pathological features associated with H 2 O 2 -mediated neuronal toxicity and provided protection against apoptotic cell death (Gao et al 2017) due to Aluminum intoxication. It was established that aluminum activated oxidative stress can deplete the cellular level of glutathione (Asher and Guilford 2016) associated with chromatolysis and gliosis seen in Fig.…”

Inhibitory Effect of Probenecid on Osteoclast Formation via JNK, ROS and COX-2

“…In a recent study, probenecid was shown to exhibit antihypertensive action by inhibiting the α-adrenergic receptor (Park and Kim, 2011). Furthermore, it was found that probenecid blocks the efflux of antioxidant glutathione (GSH), as well as GSH conjugates in neurons (Du et al ., 2016). Although the molecular mechanisms involved in oxidative stress-induced osteoclast differentiation are complex and not well characterized (Lee et al ., 2005; Oka et al ., 2012), therapeutic strategies to prevent ROS generation may be useful in overcoming many diseases, including osteoporosis and diabetes.…”