A study of the protective effect of nicotinamide adenine dinucleotide(NAD+)on post-resuscitation brain injury

Zhang Guowei; Xu Jiefeng; Hu Yufeng; Zhang Mao

doi:10.3969/j.issn.1002-1949.2026.02.006

Abstract：Objective To observe the effect of nicotinamide adenine dinucleotide(NAD+)on cerebral injury after cardiopulmonary resuscitation(CPR)in pigs and to preliminarily explore its potential mechanism.Methods Twenty-two domestic male white pigs were randomly divided into three groups:a Sham group(n=6),a CPR group(n=8),and an NAD+group(n=8).Animals in the Sham group only underwent tracheal intubation and vascular catheterization.Animals in the CPR and NAD+groups were subjected to ventricular fibrillation induced by electrical stimulation of the right ventricle for 10 minutes,followed by 6 minutes of CPR to establish the model.At 5 minutes after resuscitation,animals in the NAD+group received an intravenous infusion of NAD+(20 mg/kg dissolved in 20 ml of normal saline)via the femoral vein using a microinfusion pump over 30 minutes,while animals in the Sham and CPR groups received an equivalent volume of normal saline.Venous blood samples were collected at 1,2,4,and 24 hours after resuscitation.Serum levels of neuron-specific enolase(NSE)and S100β protein were measured using ELISA.Neurological deficit scores(NDS)were assessed at 24 hours post-resuscitation,after which the animals were euthanized,and cortical brain tissues were collected.Western blot was used to detect the expression of cleaved caspase-3(C-CAS3),phosphorylated cuproptosis-related protein ferredoxin 1(FDX1),dihydrolipoamide acetyltransferase(DLAT),and lipoic acid synthetase(LIAS).Intracellular copper ion levels were measured using a copper-specific assay kit and spectrophotometry.One-way analysis of variance(ANOVA)with Bonferroni post-hoc testing was used for comparisons inter-group comparisons.Results Baseline parameters did not differ significantly among the three groups.All animals in the Sham group survived,while two animals died in each of the CPR and NAD+groups.At 24 hours post-resuscitation,compared to the Sham group,both the CPR and NAD+groups showed significantly higher NDS and increased serum levels of NSE and S100β(all P＜0.05).Compared to the CPR group,the NAD+group exhibited lower NDS(283±55 vs.93±50,P＜0.05)and significantly reduced serum NSE and S100β concentrations at 2,4,and 24 hours(all P＜0.05).Compared to the Sham group,both the CPR and NAD+groups showed decreased expression of FDX1,DLAT,and LIAS,along with increased intracellular copper ion levels and apoptosis indicators in the cerebral cortex at 24 hours post-resuscitation.Compared to the CPR group,the NAD+group demonstrated increased expression of FDX1,DLAT,and LIAS,as well as reduced intracellular copper ion levels and apoptosis indicators in the brain tissue.Conclusions NAD+has a protective effect against cerebral injury after CPR in pigs,and its mechanism is associated with mitochondrial cuproptosis in brain cells following cerebral ischemia-reperfusion injury.