1400W对脂多糖诱导少突胶质细胞前体死亡通路的阻断研究

1400W blocks death pathway of LPS-induced activated-microglia to preOLs

摘要:

目的：建立细菌脂多糖（LPS）诱导的2日龄感染型脑室周围白质软化（PVL）新生大鼠动物模型，探讨iNOS抑制剂1400W 对LPS诱导脑白质少突胶质细胞（OL）前体死亡的体内阻断效果。方法：2日龄新生大鼠随机分为假手术组、PVL组、1400W-即刻组、1400W-8h组、1400W-16h组以及1400W-24h组，分别在LPS注射后即刻、8 h、16 h以及24 h经皮下注射1400W 20 mg/kg。于造模后第5天处死取脑，分别进行光镜下脑白质病理评估、硝酸还原法检测一氧化氮（NO）含量、Western blot 法检测诱生型一氧化氮合酶（iNOS）表达量、免疫组织化学染色检测过氧亚硝酸盐（ONOO）含量以及OL前体数量。结果：LPS诱导可引起新生大鼠脑白质明显损伤，脑内iNOS表达明显上调,NO和ONOO-含量显著增加，脑白质内少突胶质细胞前体标记物(O4）标记的OL前体显著减少。与PVL组比较，1400W-即刻组、1400W-8h组以及1400W-16h组新生大鼠的脑白质病理均获明显改善，iNOS蛋白合成量、NO及ONOO-生成量均显著降低，OL前体数量明显增加（P<0.05）。1400W-24h组的各项检测结果与PVL组相似，均无明显改善。结论：体内研究确认1400W通过抑制iNOS的表达上调、减少脑内NO及ONOO-的生成量，从而阻断OL前体的死亡通路，发挥对脑白质的保护效果。在LPS诱导后16 h内应用1400W，均有良好的脑保护作用。［中国当代儿科杂志，2010，12（5）：357-362］

Abstract:

OBJECTIVE: To explore the efficacy of inductible nitric oxide synthase (iNOS) inhibitor 1400W in vivo in blocking the death pathway of lipopolysaccharide (LPS)-induced activated-microglia to preoligodendrocytes (preOLs) in neonatal rats with infective-type periventricular leukomalacia (PVL) induced by LPS. METHODS: Two-day-old neonatal rats were randomly divided into: a sham-operated group, an untreated PVL group, and four 1400W-treated PVL groups that were subcutaneously administrated with 20 mg/kg of 1400W at 0 h, 8 hrs, 16 hrs, and 24 hrs after LPS induction, respectively. The brain specimens were obtained 5 days after LPS induction. The pathological assessment of cerebral white matter was performed under a light microscope. Concentrations of nitric oxide (NO) were measured by nitric acid-deoxidize colorimetry. Synthesis of iNOS was determined by Western blot analysis. Peroxynitrite (ONOO) level and the amount of preOLs were determined by immunocytochemistry. RETHODS: The obvious injuries of periventricular white matter, massive loss of positive O4-labelled preOLs, and increased levels of NO, ONOO and iNOS were observed in neonatal rats with PVL. Compared to the untreated PVL group, the use of 1400W at 0 h, 8 hrs and 16 hrs after LPS induction significantly improved white matter injuries, reduced the levels of NO, ONOO and iNOS, and increased the amount of O4-labelled preOLs. However, the use of 1400W at 24 hrs after LPS induction did not result in the improvements. CONCLUSIONS: iNOS inhibitor 1400W can effectively block the toxicity of LPS-activated microglia to preOLs and protect cerebral white matter through inhibiting iNOS and reducing the production of NO and ONOO. The use of 1400W within 16 hrs after LPS induction may provide cerebral protections in neonatal rats with PVL.［Chin J Contemp Pediatr, 2010, 12(5):357-362］