脑缺血再灌注损伤中激肽释放酶的蛋白治疗。

The protein therapy of kallikrein in cerebral ischemic reperfusion injury.

机构信息

Department of Neurology, Huashan Hospital, Fudan University, Number 12 Wulumuqi Mid Road, Shanghai, PR China.

出版信息

Curr Med Chem. 2009;16(34):4502-10. doi: 10.2174/092986709789760689.

DOI:10.2174/092986709789760689

PMID:19903154

Abstract

OBJECTIVE

Biomacromolecule like exogenous Kallikrein is difficult to pass through biomembrane and blood brain barrier. So, the use of exogenous Kallikrein for the therapy of nervous system diseases is restricted. We constructed the Protein Transduction Domain-Kallikrein (PTD-Kallikrein), checked its function of penetration and biotoxicity, and observed its influence on neurons and ischemic brain tissues.

METHODS

PTD-Kallikrein (tissue kallikrein) was prepared by chemical synthesis. After PTD-Kallikrein injected 2.5 hours, rats brains were taken out and contents of Kallikrein were quantitated to observe the function of passing through blood brain barrier. Cell survival rate were measured by XTT methods to determine the peptide's biotoxicity. Apoptosis were inspected by TUNEL. PTD-Kallikrein was administrated immediately after cerebral ischemia. 24h later, infarct volume was determined by TTC stain and IL-1beta, TNF-alpha as well as PGE2 were measured by ELISA.

RESULTS

PTD-Kallikrein can pass through the biomembrane and blood brain barrier; 2. PTD-Kallikrein itself has no obviously biotoxicity. 3. PTD-Kallikrein increases cell survival rate, decreases neurons apoptosis during OGD/recovery; 4. HOE140 inhibits the effects of PTD-Kallikrein. 5. PTD-Kallikrein improves neurological impairment, decreases the infarct volume, and inhibits the release of IL-1beta, TNF-alpha, PGE2. 6. HOE140 inhibits the effects of PTD-Kallikrein on ischemia-reperfusion injury.

CONCLUSIONS

PTD-Kallikrein can pass through the biomembrane and BBB efficiently and itself has no obviously biotoxicity. 2. PTD-Kallikrein has neuroprotective effect on neurons and cerebral ischemia injury. 3. PTD-Kallikrein is partially mediated by B2 receptors.

摘要

目的

生物大分子类似外源性激肽释放酶很难穿透生物膜和血脑屏障。因此，外源性激肽释放酶在神经系统疾病的治疗中的应用受到限制。我们构建了蛋白转导结构域-激肽释放酶（PTD-Kallikrein），检查了其穿透和生物毒性的功能，并观察了其对神经元和缺血性脑组织的影响。

方法

化学合成 PTD-Kallikrein（组织激肽释放酶）。在 PTD-Kallikrein 注射 2.5 小时后，取出大鼠大脑并定量测定激肽释放酶的含量，以观察其穿过血脑屏障的功能。通过 XTT 法测定细胞存活率来确定肽的生物毒性。通过 TUNEL 检测细胞凋亡。脑缺血后立即给予 PTD-Kallikrein。24 小时后，通过 TTC 染色测定梗死体积，并通过 ELISA 测定 IL-1β、TNF-α和 PGE2 的含量。

结果

PTD-Kallikrein 可以穿透生物膜和血脑屏障；2. PTD-Kallikrein 本身没有明显的生物毒性。3. PTD-Kallikrein 增加 OGD/恢复期间的细胞存活率，减少神经元凋亡；4. HOE140 抑制 PTD-Kallikrein 的作用。5. PTD-Kallikrein 改善神经功能缺损，减少梗死体积，抑制 IL-1β、TNF-α、PGE2 的释放。6. HOE140 抑制 PTD-Kallikrein 对缺血再灌注损伤的作用。