Basic investigation of hypothermia for a treatment of stroke : ischemic neuronal damage by the disruption of blood-brain barrier and microglial activation

通过检测静脉注射伊文思蓝染料对前脑缺血沙土鼠血脑屏障(BBB)的破坏作用。荧光分光光度法测染料渗出量。缺血后1d前脑可见轻度渗出，4d时渗出最为严重。低温沙土鼠全脑渗出减少。在低温条件下，前脑缺血后侧脑室注射血清，可检测到海马区CA1区神经元死亡。非白蛋白大鼠大脑中动脉闭塞1h后24 h的脑梗塞体积较正常大鼠缩小。本实验观察了血清对培养大鼠小胶质细胞产生超氧化物歧化酶(O_2~-)、一氧化氮(NO)和肿瘤坏死因子-α(肿瘤坏死因子-α)的影响。我们在…中发现了一种血清因子，白蛋白佛波酯诱导的O_2~(2+)产生增加，内毒素诱导的NO和肿瘤坏死因子-α的产生增加。我们在白蛋白分子中确定了这种增强的活性部位。一系列合成肽的活性表明，活性最低的序列为Leu-His-Thr-Leu。激活的小胶质细胞可以释放多种神经毒性介质。这类神经毒性介质已被提出在脑缺血时对神经元造成损害。因此，抑制小胶质细胞的激活将为缺血性卒中的治疗打开新的窗口。我们观察了短时间亚低温联合每日γ-谷氨酰胺(茶氨酸)治疗对沙土鼠短暂性前脑缺血延迟性神经元死亡的保护作用。缺血后每日给予茶氨酸5 mg/kg，连续30d。存活30天后，用组织学方法评价大鼠海马区CA1区神经元的损伤程度。持续时间较短的轻度缺血后低温联合茶氨酸治疗导致存活的CA1神经元显著增加(分别为67%和72%)，与对照组(8%的存活率)相比。这些发现表明，持续时间较短的亚低温联合茶氨酸对缺血性神经元损伤有持续的有益效果。较少

Blood-brain barrier (BBB) breakdown was evaluated by examining extravasation of intravenously injected Evans Blue dye in forebrain ischemic gerbils. Dye extravasation was estimated by the fluorometric method. Slight extravasation was observed in the forebrain 1 days after ischemia, and was most severe 4 days after ischemia. Extravasation decreased in whole of brain in the hypothermic gerbils. When serum were administered through the lateral ventricle after forebrain ischemia under hypothermic conditions, neuronal death was detected in CA1 neurons of the hippocampus. Infarction volume 24 h after 1 h MCA occlusion decreased in none-albumin rats compared with normal rats. These results suggest that infiltration of serum factors into brain parenchyma may play a key role in ischemic neuronal death.We examined the effect of serum on the production of super oxide (O_2^-), nitric oxide (NO) and tumor necrosis factor-α (TNF-α) in rat cultured microglia. We found that a serum factor, albumin, in … More creased phorbol ester (PMA)-induced O_2^- production and that serum also enhanced lipopolysaccharide (LPS)-induced production of NO and TNF-α. We identified the active site of this enhancement within the albumin molecule. The activated of a series of synthesized peptides conclusely indicated that the minimum active sequences was Leu-His-Thr-Leu. Activated microglia can release various neurotoxic mediators. Such neurotoxic mediators have been proposed to exert neuronal damage under brain ischemia. Therefore, suppression of microglial activation will open a new window for treatment of ischemic stroke.We examined the protective effect of short-duration and mild hypothermia in combination with daily γ-glutamylethylamide (theanine) treatment on delayed neuronal death in transient forebrain ischemia of gerbils. Theanine was administered daily 5 mg/kg alter ischemia for 30 days. After survival of 30 days, the extent of CA1 neuronal damage in the hippocampus was assessed histologically. Short-duration and mild postischemic hypothermia combined with theanine treatment led to significant increase (67% and 72%, respectively) in viable CA1 neurons, compared to the control group (8% survival). These findings indicate that short-duration and mild hypothermia in combination with theanine has a beneficial effect on ischemic neuronal damage consistently. Less