Calpain 1 and -2 play opposite roles in cord formation of lymphatic endothelial cells via eNOS regulation
Calpain 1 and -2 play opposite roles in cord formation of lymphatic endothelial cells via eNOS regulation
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DOI：
10.1007/s13577-012-0042-7
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发表时间：
2012-02
期刊：
影响因子：
4.3
通讯作者：
Orawin Prangsaengtong;K. Senda;Y. Doki;J. Park;M. Jo;H. Sakurai;N. Shibahara;I. Saiki;K. Koizumi-K
中科院分区：
文献类型：
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作者：
Orawin Prangsaengtong;K. Senda;Y. Doki;J. Park;M. Jo;H. Sakurai;N. Shibahara;I. Saiki;K. Koizumi-K
钙蛋白酶是钙依赖性蛋白酶家族。钙蛋白酶 1 和 2 两种亚型与血管生成以及内皮细胞粘附和迁移有关。钙蛋白酶调节 eNOS 的功能；然而，钙蛋白酶和 eNOS 与淋巴管生成的关系仍不清楚。在本研究中，我们评估了钙蛋白酶和 eNOS 在基质胶上淋巴内皮细胞形成索索中的作用。用针对钙蛋白酶 1 或 2 的 siRNA 转染人淋巴微血管真皮源性内皮细胞。钙蛋白酶 2 敲低（而非钙蛋白酶 1 敲低）显着减少了基质胶上的索形成、粘附和迁移。通过免疫沉淀和蛋白质印迹检测，这些降低与 eNOS、磷酸化 eNOS 和 Hsp90 水平的降低相关。相比之下，与对照相比，敲低钙蛋白酶 1（而非钙蛋白酶 2）可增加细胞粘附、增强迁移，并通过增加脊髓长度来稳定晚期脊髓形成。这些差异与磷酸化 eNOS 水平的增加相关。结果表明，钙蛋白酶和 eNOS 的功能对于淋巴内皮细胞的索形成很重要。我们首次发现了calpain 1和2的不同功能。Calpain 1参与eNOS和Hsp90的降解以及eNOS的磷酸化，而calpain 2在Matrigel上调节淋巴内皮细胞在索状形成过程中的eNOS磷酸化。
Calpains are a family of calcium-dependent proteases. Two isoforms, calpain 1 and 2, have been implicated in angiogenesis and endothelial cell adhesion and migration. Calpains regulate the function of eNOS; however, the relation of calpains and eNOS to lymphangiogenesis is still unclear. In the present study, we evaluated the role of calpain and eNOS in the formation of cords by lymphatic endothelial cells on Matrigel. Human lymphatic microvascular dermal-derived endothelial cells were transfected with siRNA against calpain 1 or 2. Calpain 2 knockdown, but not calpain 1 knockdown, significantly reduced cord formation, adhesion, and migration on Matrigel. These decreases correlated with a reduction in eNOS, and phosphorylated eNOS and Hsp90 levels, as assayed by immunoprecipitation and western blotting. In contrast, the knockdown of calpain 1, but not calpain 2, increased cell adhesion, enhanced migration, and stabilized late-stage cord formation by increasing cord length compared to the control. These differences correlated with an increase in the level of phosphorylated eNOS. The results indicated that the functions of calpains and eNOS are important for cord formation by lymphatic endothelial cells. For the first time, we have found different functions of calpain 1 and 2. Calpain 1 is involved in the degradation of eNOS and Hsp90 and the phosphorylation of eNOS, while calpain 2 regulates eNOS phosphorylation during cord formation by lymphatic endothelial cells on Matrigel.