Identification of the major in vivo ABl-42-degrading catabolic-system in brain parenchyma.

淀粉样蛋白β肽（Ap）在大脑中的积累触发了导致阿尔茨海默病的病理级联。为了研究AB分解代谢的机制，我们分析了注射到大鼠海马的多重放射性标记的3H/ 14c - a - βl-42的结局，并使用一组肽酶抑制剂确定了与neprilysin相似或相同的中性内肽酶作为限速肽酶。为了进一步鉴定硫孤儿敏感肽酶，我们分析了中性内肽酶家族中6种重组硫孤儿敏感肽酶对ABl-42的分解代谢。其中Neprilysin降解AB的速度最快，效率最高。neprilysin缺陷小鼠分解AB的能力显著降低。我们还观察到小鼠大脑中内源性AB水平（-/- > +/- > +/+）的基因剂量依赖性分化。neprilysin缺乏小鼠大脑中AB42水平的增加与携带PSI突变的小鼠相当。此外，野生型小鼠脑内的区域Ap水平依次为海马>皮质>纹状体/丘脑>小脑，且这种趋势在神经元素缺乏小鼠中明显放大。在老龄小鼠的实验中，与2月龄相比，30月龄时海马中neprilysin活性及其蛋白质含量降低了200%。随着年龄的增长，大脑皮层neprilysin的活性也有轻微但显著的下降，而其他区域的活性则没有变化。这些观察结果表明，尽管neprilysin家族存在分子冗余，但neprilysin是大脑中主要的AB降解酶，而且neprilysin活性的降低，特别是在AB病理易感区域，将通过促进AB沉积而促进阿尔茨海默病的发展。

Accumulation of amyloid-pβ peptide (Ap) in brain triggers the pathological cascade leading to Alzheimer' s disease. To examine the mechanism of AB catabolism, we analysed the fate of multiply radio-labeled 3H/14C-Aβl-42 injected into rat hippocampus and identified neutral endopeptidase (s) similar or identical to neprilysin as the rate-1imiting peptidase using a panel of peptidase inhibitors. Chronic infusion of thiorphan, a specific inhibitor of neprilysin, into rat hippocampus caused accumulation of endogenous AB. To further identify the thiorphan-sensitive peptidase, we analysed ABl-42 catabolism by six recombinant thiorphan-sensitive peptidases in neutral endopeptidase family. Neprilysin degraded AB most rapidly and efficiently among them. Neprilysin-deficient mice showed significantly reduced ability to catabolize AB. We also observed a gene dosage-dependent clevation of the endogenous AB levels (-/- > +/- > +/+) in the mouse brains. The increase in the AB42 level in the neprilysin-deficient mouse brain was comparable to that in the mice carrying PSI mutation. Moreover, the regional Ap levels in wild-type mouse brain were in the order of hippocampus > cortex > striatum/thalamus > cerebellum and this tendency was markedly exaggerated in the neprilysin-deficient mice. In the experiments using aged mice, neprilysin activity and its protein content in hippocampus were reduced by 200% at 30 months-old ages, compared to 2 months-old ages. The neprilysin activity in cortex was also slightly but significantly reduced with aging, whereas those in other regions were not changed. These observations suggest that the neprilysin is the major AB-degrading enzyme in brain despite the molecular redundancy of the neprilysin family and that reduction of neprilysin activity particularly in the regions vulnerable to AB pathology will contribute to Alzheimer' s disease development by promoting AB deposition.