Background/Aim:Circadian rhythm proteins (CRP) play critical roles in both physiological and pathophysiological conditions, including neurodegenerative disorders. As member of CRP, the nuclear receptors REV-ERBα/β regulates circadian rhythm particularly by inhibiting Bmal1 protein and involved in the neuroinflammation- and cell death processes. However, their roles in the development of neuronal injury after traumatic brain injury (TBI) were largely unexplored, which were investigated in the present study.Materials and Methods: For the induction of TBI, animals were submitted to the cryogenic model of TBI, which is a commonly used animal model and shares essential similarities with cerebral ischemia in terms of pathophysiological cascades. To assess the impact of REV-ERB proteins on TBI, both RevErbα and RevErbβ proteins were activated or deactivated, and their expression profiles were determined by Western blot analyzes. Infarct volume and brain swelling were analyzed by Cresyl violet staining. Blood-brain barrier (BBB) permeability was analyzed by immunoglobulin G extravasation. Neuronal survival was analyzed by NeuN immunohistochemistry.Results: Our observations indicate that RevErbβ significantly reduced brain injury after TBI, which was reversed by inhibiting this protein. Not activation but the inhibition of both RevErb proteins increased brain swelling significantly. In addition, both RevErbα and RevErbβ improved BBB permeability and neuronal survival significantly, which were reversed by their inhibitions.Conclusion: Our results show that both RevErbα and particularly RevErbβ play significant roles in the development of neuronal injury after TBI. Our findings suggest that REV-ERB proteins would be a potential therapeutic target for the treatment of neurodegenerative diseases.

Background and Aims: Intergenerational disease transmission, where parental exposures or experiences influence disease susceptibility in offspring, may represent a crucial layer of stroke risk that extends beyond genetics alone. Environmental conditioning, such as intermittent sub-lethal hypoxia, can induce adaptive protective stress responses in the brain. However, whether such parental conditioning enhances offspring resilience to cerebral ischaemia remains unclear. This study investigates whether intermittent hypoxia in parents acts as an intergenerational conditioning stimulus, conferring resilience to ischaemic stroke in offspring, and explores associated molecular mechanisms. Methods: Male and female Balb/C mice (F0) were exposed to intermittent hypoxia (8% O ₂ , 2 hours every other day, 16 cycles) prior to mating. To confirm that intermittent hypoxia induced neuroprotection in the parental generation, a separate cohort of F0 mice underwent transient middle cerebral artery occlusion (tMCAO). Offspring (F1) were generated from hypoxia-exposed F0 breeders and divided into four groups: biparental hypoxia, paternal hypoxia, maternal hypoxia, and normoxic controls. Adult F1 offspring also underwent tMCAO to model ischaemic stroke. Infarct volume and brain swelling were assessed 48 hours post-ischaemia. In a subgroup of F1 offspring, tandem mass tag (TMT)-based proteomic analysis of injured brain tissue was performed post-stroke to identify molecular pathways associated with neuroprotection. Results: Parental intermittent hypoxia significantly reduced infarct size and swelling in F0 mice. These protective effects were inherited by F1 offspring, with biparental exposure producing the greatest reduction in infarct volume, followed by maternal-only and paternal-only groups, and exhibiting sex-specific differences. Proteomic profiling revealed distinct treatment and lineage clusters. Key pathways implicated in offspring neuroprotection included metabolic regulation, immune signalling, cytoskeletal organisation, and cell survival, notably involving PI3K-Akt and EGFR pathways. Conclusions: Intermittent hypoxia in parents acts as an intergenerational conditioning stimulus, conferring offspring resilience to ischaemic stroke. This neuroprotective phenotype is supported by coordinated molecular adaptations in key pathways involved in survival and stress response. These findings highlight the potential for parental environmental conditioning to shape stroke outcomes in offspring, opening new avenues for therapeutic exploration.

BACKGROUND: Intergenerational disease transmission, where parental exposures or experiences influence disease susceptibility in offspring, may represent a crucial layer of stroke risk that extends beyond genetics alone. Environmental conditioning, such as intermittent sublethal hypoxia, can induce adaptive protective stress responses in the brain. However, whether such parental conditioning enhances offspring resilience to cerebral ischemia remains unclear. This study investigates whether intermittent hypoxia in parents acts as an intergenerational conditioning stimulus, conferring resilience to ischemic stroke in offspring, and explores associated molecular mechanisms. METHODS: Male and female inbred mouse strain originally derived from the Bagg albino lineage mice (parental generation [F0]; 8–10 weeks old) were exposed to intermittent hypoxia (8% O ₂ , 2 hours every other day, 16 cycles) before mating. To confirm parental neuroprotection, F0 mice underwent transient middle cerebral artery occlusion. First filial (offspring) generation from hypoxia-exposed F0 were divided into biparental, paternal-only, maternal-only hypoxia, and normoxic groups. Adult first filial (offspring) generation (8–10 weeks old) offspring also underwent transient middle cerebral artery occlusion to model ischemic stroke. Infarct volume and brain swelling were assessed 48 hours post-ischemia. A subgroup of F0 and first filial (offspring) generation brains was analyzed by tandem mass tag proteomics to identify molecular pathways linked to neuroprotection. RESULTS: Parental intermittent hypoxia significantly reduced infarct size and swelling in F0 mice. These protective effects were inherited by the first filial (offspring) generation, with biparental exposure producing the greatest reduction in infarct volume, followed by maternal-only and paternal-only groups, and exhibiting sex-specific differences. Proteomic profiling revealed distinct treatment and lineage clusters. Key pathways implicated in offspring neuroprotection included metabolic regulation, immune signaling, cytoskeletal organization, and cell survival, notably involving PI3K-Akt (phosphoinositide 3-kinase-Akt signalling pathway) and EGFR (epidermal growth factor receptor) pathways. CONCLUSIONS: Intermittent hypoxia in parents acts as an intergenerational conditioning stimulus, conferring offspring resilience to stroke. This neuroprotective phenotype is supported by coordinated molecular adaptations in key pathways involved in survival and stress response. These findings highlight the potential for parental environmental conditioning to shape stroke outcomes in offspring, opening new avenues for therapeutic exploration.