Hypoxia, the major cause of ischemic injury, leads to debilitating disease in infants via birth asphyxia and cerebral palsy, whereas in adults via heart attack and stroke. A widespread, natural protective phenomenon termed ‘hypoxic preconditioning’ (PH) occurs when prior exposures to hypoxia eventually result in robust hypoxia resistance. Accordingly, we have developed and optimized a novel model of hypoxic preconditioning in adult zebrafish to mimic the tolerance of mini stroke(s) in human, which appears to protect against the severe damage inflicted by a major stroke event. Here, we observed a remarkable difference in the progression pattern of neuroprotection between preconditioning hypoxia followed by acute hypoxia (PH) group, and acute hypoxia (AH) only group, with noticeable sex difference when compared with normoxia behaviour upon recovery. Since gender difference has been reported in stroke risk factors and disease history, it was pertinent to investigate whether any such sex difference also exists in PH’s protective mechanism against acute ischemic stroke. In order to elucidate the neural molecular mechanisms behind sex difference in neuroprotection induced by PH, a high throughput proteomics approach utilizing iTRAQ was performed, followed by protein enrichment analysis using ingenuity pathway analysis (IPA) tool. Out of thousands of significantly altered proteins in zebrafish brain, the ones having critical role either in neuroglial proliferation/differentiation or neurotrophic functions were validated by analyzing their expression levels in preconditioned (PH), acute hypoxia (AH), and normoxia groups. The data indicate that female zebrafish brains are more protected against the severity of AH when exposed to the hypoxic preconditioning. The study also sheds light on the involvement of many signalling pathways underlying sex difference in preconditioning-induced neuroprotective mechanism, which can be further validated for the therapeutic approach.

缺氧是缺血性损伤的主要原因，导致婴儿因出生时窒息和脑瘫而使人衰弱，而成年人则因心脏病和中风而使人衰弱。当先前暴露于缺氧环境最终导致强大的抗缺氧性时，就会发生一种广泛的，称为“低氧预处理”（PH）的自然保护现象。因此，我们已经开发和优化了成年斑马鱼的低氧预处理新模型，以模拟人类对迷你中风的耐受性，这似乎可以防止重大中风事件造成的严重损害。在这里，我们观察到在预适应性低氧继之以急性低氧（PH）组和仅急性低氧（AH）组之间神经保护的进展模式存在显着差异，与恢复时的正常氧行为相比，存在明显的性别差异。由于在中风危险因素和疾病史上已报告了性别差异，因此有必要调查在PH的急性缺血性中风保护机制中是否也存在这种性别差异。为了阐明由PH引起的神经保护作用中性别差异背后的神经分子机制，采用iTRAQ的高通量蛋白质组学方法进行了研究，然后使用智能路径分析（IPA）工具进行了蛋白质富集分析。在斑马鱼大脑中成千上万的显着改变的蛋白质中，通过分析其在预处理（PH），急性缺氧（AH）和常氧组中的表达水平，验证了在神经胶质增生/分化或神经营养功能中起关键作用的蛋白质。数据表明，暴露于低氧预处理条件下，雌性斑马鱼的大脑对AH严重程度的保护更大。该研究还揭示了性别差异导致的许多信号传导通路参与了预处理诱导的神经保护机制，这些途径可进一步用于治疗方法的验证。