Heme Oxygenase-1 Mediates Neuroprotection Conferred by Argon in Combination with Hypothermia in Neonatal Hypoxia–Ischemia Brain Injury

Abstract

Background Hypoxic–ischemic encephalopathy is a major cause of mortality and disability in the newborn. The authors investigated the protective effects of argon combined with hypothermia on neonatal rat hypoxic–ischemic brain injury. Methods In in vitro studies, rat cortical neuronal cell cultures were challenged by oxygen and glucose deprivation for 90 min and exposed to 70% Ar or N2 with 5% CO2 balanced with O2, at 33°C for 2 h. Neuronal phospho-Akt, heme oxygenase-1 and phospho-glycogen synthase kinase-3β expression, and cell death were assessed. In in vivo studies, neonatal rats were subjected to unilateral common carotid artery ligation followed by hypoxia (8% O2 balanced with N2 and CO2) for 90 min. They were exposed to 70% Ar or N2 balanced with oxygen at 33°, 35°, and 37°C for 2 h. Brain injury was assessed at 24 h or 4 weeks after treatment. Results In in vitro studies, argon–hypothermia treatment increased phospho-Akt and heme oxygenase-1 expression and significantly reduced the phospho-glycogen synthase kinase-3β Tyr-216 expression, cytochrome C release, and cell death in oxygen–glucose deprivation–exposed cortical neurons. In in vivo studies, argon–hypothermia treatment decreased hypoxia/ischemia-induced brain infarct size (n = 10) and both caspase-3 and nuclear factor-κB activation in the cortex and hippocampus. It also reduced hippocampal astrocyte activation and proliferation. Inhibition of phosphoinositide-3-kinase (PI3K)/Akt pathway through LY294002 attenuated cerebral protection conferred by argon–hypothermia treatment (n = 8). Conclusion Argon combined with hypothermia provides neuroprotection against cerebral hypoxia–ischemia damage in neonatal rats, which could serve as a new therapeutic strategy against hypoxic–ischemic encephalopathy.

The Molecular Pathway of Argon-Mediated Neuroprotection

“…They concluded that NF-κB, as a potential modulator of apoptosis, may be a downstream target, exerting distinct effects on neuronal cells. These findings were later on supported by Zhao et al [47] in an in vivo model of neonatal hypoxia and brain injury. This argon-mediated suppression of NF-κB and STAT3 signaling was counteracted by TLR inhibition in vitro, rendering these possible pathways.…”

“…It is tempting to speculate that HO-1 may be the central and common part in different settings of protection. Zhao et al [47] confirmed the findings with respect to HO-1 in their in vitro OGD and hypothermia model as well as in vivo argon-mediated neuroprotective effects through HO-1 induction. These effects were prominent in both, the cortex and the hippocampal area, while the brain infarct volume was reduced.…”

“…Furthermore, the authors demonstrated that IRI-induced caspase-3 cleavage was suppressed by argon 75%, if administered immediately after injury. Extended FACS analysis by Zhao et al and Ulbrich et al confirmed the anti-apoptotic properties of argon, as well as results, proving a reduction in reactive oxygen species (ROS) formation and normalization of mitochondrial membrane potential after argon treatment [38,39,47]. …”

“…In their model of neonatal asphyxia the authors found that argon treatment did not show any effects regarding the phosphorylation of pro-apoptotic proteins BAX and BCL-XL, but a significant increase of the anti-apoptotic acting protein BCL-2. Zhao et al [47] demonstrated in vitro and in vivo that argon administration after neuronal injury may upregulate BCL-2 expression, while caspase-3 expression was cut back to baseline levels in the cortex and the hippocampal area of MCAO injured rats. Ulbrich et al [27] confirmed these findings of argon’s anti-apoptotic properties in a different in vivo model of retinal ischemia and reperfusion injury (IRI).…”

Argon attenuates multiorgan failure following experimental aortic cross‐clamping

“…It has also been demonstrated previously that argon activates protective pathways in the neuronal and cardiac tissue during regional ischaemia. For example, argon increased heme oxygenase 1 expression and Akt phosphorylation in hypoxic neurones . Its effect was found to be blunted by the pharmacological inhibition of phosphoinositide 3‐kinase or extracellular regulated kinase .…”