Department of Biomedicine, Aarhus University, Aarhus, Denmark.
J Nucl Med. 2013 Jan;54(1):159-65. doi: 10.2967/jnumed.112.110528. Epub 2012 Dec 11.
(99m)Tc-labeled dimercaptosuccinic acid ((99m)Tc-DMSA) accumulates in the kidney cortex and is widely used for imaging of the renal parenchyma. Despite its extensive clinical use, the mechanism for renal targeting of the tracer is unresolved. Megalin and cubilin are cooperating receptors essential to the proximal tubule endocytic uptake of proteins from the glomerular ultrafiltrate. We have used megalin/cubilin-deficient mice produced by gene knockout to determine whether receptor-mediated endocytosis is responsible for the renal uptake of (99m)Tc-DMSA.
Control or megalin/cubilin-deficient mice were injected intravenously with 0.5 MBq of (99m)Tc-DMSA or (99m)Tc-mercaptoacetyltriglycine (MAG3). Whole-body scintigrams and the activity in plasma, urine, and the kidneys were examined 6 h after injection. The size and identity of (99m)Tc-DMSA-bound proteins in urine were analyzed by fractionation by centrifugation and separation by sodium dodecyl sulfate polyacrylamide gel electrophoresis, followed by autoradiography and mass spectrometry.
No renal accumulation of (99m)Tc-DMSA was identified in scintigrams of megalin/cubilin-deficient mice. The renal accumulated activity of the tracer was reduced to 11.4% (± 2.5%, n = 7) of the normal uptake in control mice, correlating with a reduction in renal megalin/cubilin expression in knockout mice to about 10% of normal. The reduced renal uptake in megalin/cubilin-deficient mice was accompanied by an increase in the urinary excretion of (99m)Tc-DMSA. Size separation of the urine by ultracentrifugation and sodium dodecyl sulfate polyacrylamide gel electrophoresis demonstrated that in megalin/cubilin-deficient mice an increased amount of (99m)Tc-DMSA was excreted in an approximately 27-kDa form, which by mass spectrometry was identified as the plasma protein α1-microglobulin, an established megalin/cubilin ligand.
(99m)Tc-DMSA is filtered bound to α1-microglobulin and accumulates in the kidneys by megalin/cubilin-mediated endocytosis of the (99m)Tc-DMSA protein complex. Renal accumulation of (99m)Tc-DMSA is thus critically dependent on megalin/cubilin receptor function and therefore is a marker of proximal tubule endocytic activity.
（99m）Tc 标记的二巯丁二酸（（99m）Tc-DMSA）积聚在肾脏皮质中，广泛用于肾脏实质的成像。尽管其在临床上广泛应用，但示踪剂肾靶向的机制仍未解决。巨球蛋白和 cubilin 是协同受体，对于从肾小球超滤液中摄取蛋白质的近端小管内吞作用至关重要。我们使用基因敲除产生的巨球蛋白/cubilin 缺陷小鼠来确定受体介导的内吞作用是否负责（99m）Tc-DMSA 的肾脏摄取。
对照或巨球蛋白/cubilin 缺陷小鼠静脉注射 0.5MBq（99m）Tc-DMSA 或（99m）Tc-巯基乙酰三甘氨酸（MAG3）。注射后 6 小时检查全身闪烁扫描和血浆、尿液和肾脏中的活性。通过离心分离和十二烷基硫酸钠聚丙烯酰胺凝胶电泳分离分析尿液中（99m）Tc-DMSA 结合蛋白的大小和身份，然后进行放射自显影和质谱分析。
在巨球蛋白/cubilin 缺陷小鼠的闪烁扫描中未发现（99m）Tc-DMSA 的肾脏积聚。示踪剂在肾脏中的累积活性减少到正常摄取的 11.4%（±2.5%，n=7），与在敲除小鼠中肾脏巨球蛋白/cubilin 表达减少到正常的 10%相关。巨球蛋白/cubilin 缺陷小鼠的肾脏摄取减少伴随着（99m）Tc-DMSA 的尿液排泄增加。通过超速离心和十二烷基硫酸钠聚丙烯酰胺凝胶电泳对尿液进行大小分离表明，在巨球蛋白/cubilin 缺陷小鼠中，（99m）Tc-DMSA 以大约 27kDa 的形式排泄增加，通过质谱鉴定为血浆蛋白α1-微球蛋白，这是一种公认的巨球蛋白/cubilin 配体。
（99m）Tc-DMSA 与 α1-微球蛋白结合并通过巨球蛋白/cubilin 介导的（99m）Tc-DMSA 蛋白复合物内吞作用积聚在肾脏中。因此，（99m）Tc-DMSA 的肾脏积聚严重依赖于巨球蛋白/cubilin 受体功能，因此是近端小管内吞活性的标志物。
