Loers Gabriele, Cui Yi-Fang, Neumaier Irmgard, Schachner Melitta, Skerra Arne
*Institut für Biosynthese Neuraler Strukturen, Zentrum für Molekulare Neurobiologie Hamburg, Universitätsklinikum-Hamburg Eppendorf, Hamburg, Germany.
†Munich Center for Integrated Protein Science (CiPS-M) and Lehrstuhl für Biologische Chemie, Technische Universität München, Freising-Weihenstephan, Germany.
Biochem J. 2014 Jun 15;460(3):437-46. doi: 10.1042/BJ20131677.
Lack of permissive mechanisms and abundance of inhibitory molecules in the lesioned central nervous system of adult mammals contribute to the failure of functional recovery, which leads to severe disabilities in motor functions or pain. Previous studies have indicated that the neural cell adhesion molecule L1 constitutes a viable target to promote regeneration. In the present study, we describe the cloning, functional expression in Escherichia coli cells and purification of a recombinant αL1 Fab fragment that binds to L1 with comparable activity as the function-triggering monoclonal antibody 557.B6 and induces neurite outgrowth and neuronal survival in cultured neurons, despite its monovalent function. Infusion of αL1 Fab into the lesioned spinal cord of mice enhanced functional recovery after thoracic spinal cord compression injury. αL1 Fab treatment resulted in reduced scar volume, enhanced number of tyrosine hydroxylase-positive axons and increased linear density of VGLUT1 (vesicular glutamate transporter 1) on motoneurons. Furthermore, the number and soma size of ChAT (choline acetyltransferase)-positive motoneurons and the linear density of ChAT-positive boutons on motoneurons as well as parvalbumin-positive interneurons in the lumbar spinal cord were elevated. Stimulation of endogenous L1 by application of the αL1 Fab opens new avenues for recombinant antibody technology, offering prospects for therapeutic applications after traumatic nervous system lesions.
成年哺乳动物受损中枢神经系统中缺乏允许性机制且存在大量抑制性分子，这导致功能恢复失败，进而导致运动功能严重残疾或疼痛。先前的研究表明，神经细胞粘附分子L1是促进再生的一个可行靶点。在本研究中，我们描述了一种重组αL1 Fab片段的克隆、在大肠杆菌细胞中的功能表达及纯化，该片段与L1结合的活性与触发功能的单克隆抗体557.B6相当，并且尽管其具有单价功能，但仍能在培养的神经元中诱导神经突生长和神经元存活。将αL1 Fab注入小鼠受损脊髓可增强胸段脊髓压迫损伤后的功能恢复。αL1 Fab治疗导致瘢痕体积减小、酪氨酸羟化酶阳性轴突数量增加以及运动神经元上VGLUT1（囊泡谷氨酸转运体1）的线性密度增加。此外，腰段脊髓中胆碱乙酰转移酶（ChAT）阳性运动神经元的数量和胞体大小、运动神经元上ChAT阳性终扣的线性密度以及小白蛋白阳性中间神经元的数量均增加。应用αL1 Fab刺激内源性L1为重组抗体技术开辟了新途径，为创伤性神经系统损伤后的治疗应用提供了前景。
