Function of subendothelial basement membranes in Blood-Brain Barrier

我们报道了一个跨膜型免疫球蛋白超家族的新成员的分子克隆，并将其编码的蛋白命名为限制蛋白，因为它选择性地定位于小鼠大脑的胶质细胞限制蛋白。Limitrin cDNA的获得采用减法杂交程序设计，以确定负责血脑屏障功能的分子。使用限制特异性抗体的Western blots结果表明，该基因产物在小鼠脑和小鼠原代星形胶质细胞中显著表达，并分布于质膜。使用共聚焦和电子显微镜的免疫组织化学研究清楚地表明，在体内，星形胶质末梢足在血管周围区域和脑膜下高度极化定位。Limitrin在脊髓和大脑的许多区域表达，但在缺乏血脑屏障的正中隆起或皮层下器官（脑室周围器官）中不表达。冷损伤破坏血脑屏障导致限制蛋白表达急剧减少。此外，在冷损伤恢复过程中，血管周围终足中限制蛋白表达的增加与损伤边缘毛细血管新生的恢复相关。我们的研究结果表明，limitrin在物理和功能上与血脑屏障相关，这意味着该蛋白可能作为屏障完整性的诊断决定因素有用。

We report the molecular cloning of a new member of the transmembrane-type immunoglobulin superfamily and designate the encoded protein as limitrin since it selectively localized to glia limitans in mouse brain. Limitrin cDNA was obtained using a subtractive hybridization procedure designed to identify molecules responsible for blood-brain barrier function. Western blots using a limitrin-specific antibody demonstrated that the gene product is expressed significantly in mouse brain and primary murine astrocytes, and is distributed in the plasma membrane. Immunohistochemical studies using confocal and electron microscopy clearly demonstrated highly polarized localization in astroglial endfeet in the perivascular region and under the pia mater in vivo. Limitrin is expressed in spinal cord and many areas of the brain but not in the median eminence or subfornical organ (the circumventricular organs) where the blood-brain barrier is lacking. Disruption of the blood-brain barrier by cold injury resulted in a drastic reduction in limitrin expression. Furthermore, during retrieval from cold injury the increased expression of limitrin in perivascular endfeet correlated with the recovery of angiogenesis in capillaries within the lesion margins. Our results suggest that limitrin is physically and functionally associated with the blood-brain barrier, implying that this protein may be useful as a diagnostic determinant of barrier integrity.