Substrate-dependent nitric oxide synthesis by secreted endoplasmic reticulum aminopeptidase 1 in macrophages
Substrate-dependent nitric oxide synthesis by secreted endoplasmic reticulum aminopeptidase 1 in macrophages
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DOI：
10.1093/jb/mvv001
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发表时间：
2015-06-01
影响因子：
2.7
通讯作者：
Tsujimoto, Masafumi
中科院分区：
文献类型：
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作者：
Goto, Yoshikuni;Ogawa, Kenji;Tsujimoto, Masafumi
在这项研究中，我们研究了氨肽酶的作用，参考内质网氨肽酶1（ERAP 1）在一氧化氮（NO）的合成采用小鼠巨噬细胞系RAW 264.7细胞激活的脂多糖（LPS）和干扰素（IFN）-γ和LPS激活的腹腔巨噬细胞来自ERAP 1基因敲除小鼠。当在具有N-末端Arg的肽的存在下测量NO合成时，观察到与在Arg存在下测量的NO合成相当的NO合成。在氨肽酶抑制剂amastatin的存在下，NO的合成在激活的RAW 264.7细胞显着减少。这些结果表明，氨肽酶参与了激活的RAW 264.7细胞的NO合成。随后，与野生型细胞相比，在ERAP 1敲除细胞中观察到NO合成的显著减少。这种减少被外源性添加的ERAP 1挽救。此外，当使用从ERAP 1敲除小鼠制备的腹腔巨噬细胞时，敲除小鼠巨噬细胞中NO合成的减少也归因于ERAP 1。在amastatin的存在下，在敲除小鼠来源的巨噬细胞中观察到进一步减少。总之，这些结果表明，几种氨肽酶在活化的巨噬细胞中以底物肽依赖的方式最大化NO合成中起重要作用，ERAP 1是参与NO合成的氨肽酶之一。
In this study, we examined the role of aminopeptidases with reference to endoplasmic reticulum aminopeptidase 1 (ERAP1) in nitric oxide (NO) synthesis employing murine macrophage cell line RAW264.7 cells activated by lipopolysaccharide (LPS) and interferon (IFN)-gamma and LPS-activated peritoneal macrophages derived from ERAP1 knockout mouse. When NO synthesis was measured in the presence of peptides having N-terminal Arg, comparative NO synthesis was seen with that measured in the presence of Arg. In the presence of an aminopeptidase inhibitor amastatin, NO synthesis in activated RAW264.7 cells was significantly decreased. These results suggest that aminopeptidases are involved in the NO synthesis in activated RAW264.7 cells. Subsequently, significant reduction of NO synthesis was observed in ERAP1 knockdown cells compared with wild-type cells. This reduction was rescued by exogenously added ERAP1. Furthermore, when peritoneal macrophages prepared from ERAP1 knockout mouse were employed, reduction of NO synthesis in knockout mouse macrophages was also attributable to ERAP1. In the presence of amastatin, further reduction was observed in knockout mouse-derived macrophages. Taken together, these results suggest that several aminopeptidases play important roles in the maximum synthesis of NO in activated macrophages in a substrate peptide-dependent manner and ERAP1 is one of the aminopeptidases involved in the NO synthesis.