Feng Yangzheng, LeBlanc Michael H
Department of Pediatrics, University of Mississippi Medical Center, 2500 North State Street, Jackson, MS 39216-4505, USA.
Eur J Pharmacol. 2006 Dec 3;551(1-3):34-40. doi: 10.1016/j.ejphar.2006.08.070. Epub 2006 Sep 8.
N-tosyl-L-phenylalanyl-chloromethyl ketone (TPCK) suppresses apoptosis and protects neurons from damage in animal models. TPCK is thought to act by inhibiting ceramide production by sphingomyelinase. Ceramide is a proapoptotic intracellular signal that is involved in the cerebral ischemia. We wished to see whether ceramide contributes to TPCK's neuroprotective effects in vivo. Seven-day-old rat pups had the right carotid arteries permanently ligated followed by 2.5 h of hypoxia (8% oxygen). TPCK (10 mg/kg, n=62) or vehicle (n=63) was administered by i.p. 5 min prior to hypoxia. The level of ceramide in brain cortex both in lesioned and unlesioned hemispheres was measured at 8 h, 18 h, 24 h, 2 and 5 days after hypoxia-ischemia using reversed phase high performance liquid chromatography. The level of ceramide significantly increased due to hypoxic-ischemia at 18, 24 h and 2 days after hypoxia (P<0.05 or P<0.01) but not at 8 h or 5 days after hypoxia as compared to the contralateral hemisphere or a sham group. Pretreatment with TPCK reduced this increase. We also examined the level of sphingomyelin and the activities of the ceramide synthesizing sphingomyelinase enzymes by thin layer chromatography. The activities of acidic and neutral sphingomyelinase significantly increased due to hypoxic ischemia at 24 h after hypoxia. TPCK significantly reduced this increase (P<0.05 vs. vehicle) but did not affect the level of sphingomyelin. The results are consistent with the hypothesis that ceramide is involved in TPCK's neuroprotective effects in hypoxic-ischemic brain injury in the newborn rat.
N-对甲苯磺酰-L-苯丙氨酰氯甲基酮（TPCK）在动物模型中可抑制细胞凋亡并保护神经元免受损伤。TPCK被认为是通过抑制鞘磷脂酶产生神经酰胺来发挥作用的。神经酰胺是一种促凋亡的细胞内信号，参与脑缺血过程。我们想了解神经酰胺是否在体内对TPCK的神经保护作用有贡献。将7日龄的大鼠幼崽右侧颈动脉永久性结扎，随后进行2.5小时的缺氧（8%氧气）处理。在缺氧前5分钟腹腔注射TPCK（10mg/kg，n = 62）或赋形剂（n = 63）。使用反相高效液相色谱法在缺氧缺血后8小时、18小时、24小时、2天和5天测量损伤和未损伤半球脑皮质中的神经酰胺水平。与对侧半球或假手术组相比，缺氧后18小时、24小时和2天，由于缺氧缺血，神经酰胺水平显著升高（P<0.05或P<0.01），但在缺氧后8小时或5天未升高。TPCK预处理可减少这种升高。我们还通过薄层色谱法检测了鞘磷脂水平以及神经酰胺合成酶鞘磷脂酶的活性。缺氧后24小时，由于缺氧缺血，酸性和中性鞘磷脂酶的活性显著增加。TPCK显著降低了这种增加（与赋形剂相比，P<0.05），但不影响鞘磷脂水平。这些结果与神经酰胺参与TPCK对新生大鼠缺氧缺血性脑损伤的神经保护作用这一假设一致。
