DEVELOPMENT OF NEUROPROTECTIVE STRATEGY BY THE COMBINATION THERAPY OF IMMUNOSUPPRESANT AND NEURONAL NITRIC OXIDE INHIBITOR ADMINISTRATION AGAINST NEONATAL BRAIN DAMAGE.

本研究的目的是探讨免疫抑制剂FK 506和神经元型一氧化氮抑制剂7-硝基吲唑联合治疗新生大鼠缺氧缺血性脑损伤的临床神经保护策略。我们还研究了FK 506对新生儿缺氧缺血性脑损伤的剂量依赖性和治疗时间窗。采用结扎大鼠左颈总动脉，暴露于8%氧（HI）90 min的方法，HI后24 h取脑进行组织学检查。HI损伤诱导了以下神经元损伤的混合物：尖缩、嗜酸性变化、核破裂、神经元丢失、半胱天冬酶3阳性细胞、海绵状变性和/或海马、额叶和颞叶皮质和/或基底神经节中巨噬细胞的囊性变化。使用脑损伤评分，通过脑中受损神经元的程度来估计脑损伤的严重程度。HI损伤后立即给予2 mg/kg FK 506（高剂量）（而非1 mg/kg（低剂量）），与溶剂注射相比，损伤后24 h神经元损伤和caspase-3活化的严重程度显著降低（剂量依赖性）。与溶剂注射相比，HI损伤后立即和30 min（但非60 min后）给予2 mg/kg FK 506显示出显著的神经保护作用（治疗时间窗）。1 mg/kg（低剂量）FK 506和7-硝基吲唑的联合治疗显示出比FK 506或7 NI单独使用显著更强的神经保护作用。

The goal of this study was to investigate the clinical neuroprotective strategy by means of the combination therapy of immunosuppressant, FK506, and the neuronal nitric oxide inhibitor, 7-nitroindazole, administration against hypoxic-ischemic (HI) brain damage in neonatal rats. We also investigated the dose dependency and therapeutic time window of FK506 against neonatal brain damage due to hypoxia-ischemia. The rats were performed by the transient occlusion of left carotid artery and the exposure to 8 % oxygen (HI) for 90 min. Then the brains were histologically examined 24 hrs after the HI. The HI insult induced the admixtures of the following neuronal damages; piknosis, eosinophilic change, karyorrhexis, loss of neurons, caspase 3-positive cells, spongy degeneration, and/or cystic change with macrophages in the hippocampus, frontal and temporal cortices, and/or basal ganglia. The severity of the brain damage was estimated by the extent of the damaged neurons in the brain, using the brain damage scores. The treatment of 2 mg/kg FK506 (high dose), but not 1 mg/kg (low dose), immediately after HI insult demonstrated significant decreases in the severity of the neuronal damage and caspase-3 activation 24 h after the insult, compared with vehicle injection (dose dependency). The treatment of 2 mg/kg FK506 immediately and 30 min after HI insult, but not 60 min after, demonstrated significant neuroprotective effect, compared with vehicle injection (therapeutic time window). The combination therapy of 1 mg/kg (low dose) FK506 and 7-nitroindazole demonstrated significantly more neuroprotetive effect than the single usage of either FK506 or 7NI.