Sex-specific effects of N-acetylcysteine in neonatal rats treated with hypothermia after severe hypoxia-ischemia

Abstract

Approximately half of moderate to severely hypoxic-ischemic (HI) newborns do not respond to hypothermia, the only proven neuroprotective treatment. N-Acetylcysteine (NAC), an antioxidant and glutathione precursor, shows promise for neuroprotection in combination with hypothermia, mitigating post-HI neuroinflammation due to oxidative stress. As mechanisms of HI injury and cell death differ in males and females, sex differences must be considered in translational research of neuroprotection. We assessed the potential toxicity and efficacy of NAC in combination with hypothermia, in male and female neonatal rats after severe HI injury. NAC 50 mg/kg/d administered 1h after initiation of hypothermia significantly decreased iNOS expression and caspase 3 activation in the injured hemisphere versus hypothermia alone. However, only females treated with hypothermia + NAC 50mg/kg showed improvement in short-term infarct volumes compared with saline treated animals. Hypothermia alone had no effect in this severe model. When NAC was continued for 6 weeks, significant improvement in long-term neuromotor outcomes over hypothermia treatment alone was observed, controlling for sex. Antioxidants may provide insufficient neuroprotection after HI for neonatal males in the short term, while long-term therapy may benefit both sexes.

Neuroprotection with hypothermia and allopurinol in an animal model of hypoxic-ischemic injury: Is it a gender question?

“…Despite the implementation of TH as a standard of care in many clinical units for HIE [ 12 ], it does not provide complete neuroprotection [ 36 ], making further investigation into additional therapies to improve neurological outcomes necessary. Xenon [ 16 ], Epo [ 37 , 38 ], N-Acetylcysteine (NAC) [ 34 ], and cannabidiol [ 39 ] are some of the drugs that have been tested in experimental models to potentiate the effects of TH. Historically, one of the first strategies evaluated for HIE treatment was allopurinol, an antioxidant molecule reported to act as a mild neuroprotective agent [ 40 ].…”

“…For histological analysis, western blot and negative geotaxis test, a minimum of n = 4 per sex to observe a 50% difference by Wilcoxon signed rank test (80% power, a = 0.05) was required. This figure was based on previous studies with antioxidant treatments [ 34 ]. For WMT, at least 6 animals per sex and per condition were necessary, as reported in other studies.…”

Sex‐dependent consequences of neonatal brain hypoxia‐ischemia in the rat

“…The developmental delay of physical characteristics, i.e., eye opening, ear unfolding, and incisor eruption, along with neurological reflexes, such as cliff aversion, gait, grasp, and limb placing, seem to be dependent on the severity of the injury and independent of sex (Ashwal et al, ; Huang et al, ; Nie et al, ). Motor skills in males appear to be more vulnerable to HI in the negative geotaxis test (Huang et al, ; Waddell et al, ).…”

“…Nevertheless, HI males did not exhibit significant improvement in sensorimotor behavioral tests after different neuroprotective treatments compared with HI females (Bona et al, ; Thoresen et al, ; Fan et al, ; Pimentel‐Coelho et al, ; Smith et al, ). Characteristics such as strength, swimming capabilities, quality of hindlimb placement, and fine movement coordination in the staircase are not sex dependent (Hill et al, ; Peterson et al, ; Nie et al, ).…”

“…Temperature modulation seems to protect females preferentially (Bona et al, ; Smith et al, ) as well as hypothermia associated with N‐acetylcisteine reduced histological damage in females only (Nie et al, ). Post‐HI rehabilitative training during adolescence also prevented histological damage and spatial memory deficits in females but not in males, as assessed in adulthood (Tsuji et al, ).…”